OA21166A

OA21166A - Modulators of cystic fibrosis transmembrane conductance regulator.

Info

Publication number: OA21166A
Application number: OA1202300043
Authority: OA
Inventors: Mark Thomas Miller; Thomas Cleveland; Jinglan Zhou; Minson BAEK; Prasuna PARASELLI; Jeremy J. Clemens; Brett C. Bookser; William Schulz BECHARA; Timothy R. Coon; Michel Gallant; Sara Sabina MADIDA RUAH; Julie LATERREUR; Vito Melillo; Yeeman K. Ramtohul; Thumkunta Jagadeeswar Reddy; Claudio Sturino; Lino Valdez
Original assignee: Vertex Pharmaceuticals Incorporated
Priority date: 2020-08-07
Filing date: 2021-08-06
Publication date: 2024-02-02

Abstract

This disclosure provides modulators of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), pharmaceutical compositions containing at least one such modulator, methods of treatment of cystic fibrosis using such modulators and pharmaceutical compositions, and processes for making such modulators.

Description

MODULATORS OF CYSTIC FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR

This application daims the benefit of U.S. Provisional Application No. 63/063,194, filed on August 7, 2020, the contents of which are incorporated by reference in its entirety.

The invention relates to modulators of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), pharmaceutical compositions containing the modulators, methods of treating cystic fibrosis and CFTR-mediated disorders using such modulators and pharmaceutical compositions, and processes for making such modulators.

Cystic fibrosis (CF) is a récessive genetic disease that affects approximately 83,000 children and adults worldwide. Despite progress in the treatment of CF, there is no cure.

In patients with CF, mutations in CFTR endogenously expressed in respiratory epithelia lead to reduced apical anion sécrétion causing an imbalance in ion and fluid transport. The resulting decrease in anion transport contributes to increased mucus accumulation in the lung and accompanying microbial infections that ultimately cause death in CF patients. In addition to respiratory disease, CF patients typically suffer from gastrointestinal problems and pancreatic insufficiency that, if left untreated, resuit in death. In addition, the majority of males with cystic fibrosis are infertile, and fertility is reduced among females with cystic fibrosis.

Sequence analysis of the CFTR gene has revealed a variety of disease-causing mutations (Cutting, G. R. et al. (1990) Nature 346:366-369; Dean, M. et al. (1990) Cell 61:863:870; and Kerem, B-S. et al. (1989) Science 245:1073-1080; Kerem, B-S et al. (1990) Proc. Natl. Acad. Sci. USA 87:8447-8451). To date, greater than 2000 mutations in the CF gene hâve been identified; currently, the CFTR2 database contains information on only 322 of these identified mutations, with suffîcient evidence to define 281 mutations as disease-causing. The most prévalent disease-causing mutation is a délétion of phenylalanine at position 508 of the CFTR amino acid sequence and is commonly referred to as the F508del mutation. This mutation occurs in many of the cases of cystic fibrosis and is associated with severe disease.

CFTR is a cAMP/ATP-mediated anion channel that is expressed in a variety of cell types, including absorptive and secretory epithelia cells, where it régulâtes anion flux across the membrane, as well as the activity of other ion channels and proteins. In epithelia! cells, normal functioning of CFTR is critical for the maintenance of electrolyte transport throughout the body, including respiratory and digestive tissue. CFTR is composed of 1480 amino acids that encode a protein which is made up of a tandem repeat of transmembrane domains, each containing six transmembrane helices and a nucléotide binding domain. The two transmembrane domains are linked by a large, polar, regulatory (R)-domain with multiple phosphorylation sites that regulate channel activity and cellular trafficking.

Chloride transport takes place by the coordinated activity of ENaC (épithélial sodium channel) and CFTR présent on the apical membrane and the Na⁺-K⁺-ATPase pump and Cl’ channels expressed on the basolateral surface of the cell. Secondary active transport of chloride from the luminal side leads to the accumulation of intracellular chloride, which can then passively leave the cell via Cl’ channels, resulting in a vectorial transport. Arrangement of Na⁺/2C17K⁺ co-transporter, Na⁺-K⁺-ATPase pump and the basolateral membrane K⁺ channels on the basolateral surface and CFTR on the luminal side coordinate the sécrétion of chloride. Because water is probably never actively transported itself, its flow across epithelia dépends on tiny transepithelial osmotic gradients generated by the bulk flow of sodium and chloride.

A number of CFTR modulators hâve recently been identified. These modulators can be characterized as, for example, potentiators, correctors, potentiator enhancers/co-potentiators, amplifiera, readthrough agents, and nucleic acid thérapies. CFTR modulators that increase the channel gating activity of mutant and wild-type CFTR at the épithélial cell surface are known as potentiators. Correctors improve faulty protein processing and resulting trafficking to the épithélial surface. Ghelani and Schneider-Futschik (2020) ACS Pharmacol. Transi. Sci. 3:4-10. There are three CFTR correctors approved by the U.S. FDA for treatment of cystic fîbrosis. However, monotherapy with some CFTR correctors has not been found to be effective enough and as a resuit combination therapy with a potentiator is needed to enhance CFTR activity. There is currently only one CFTR potentiator that is approved for the treatment of cystic fîbrosis. Thus, although the treatment of cystic fîbrosis has been transformed by these new small molécule CFTR modulators, new and better modulators are needed to prevent disease progression, reduce the severity of the cystic fîbrosis and other CFTR-mediated diseases, and to treat the more severe forms of these diseases.

One aspect of the invention provides novel compounds, including compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Hlb, IIIc, Hld, IHe, and Hlf, Compounds 1 to 53, Compounds 54 to 77, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing.

For example, compounds of Formula I can be depicted as:

and deuterated dérivatives and pharmaceutically acceptable salts thereof, wherein: X is selected from -O-, -S-, -SO-, and -SO2-;

£ B J each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and * ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, Cô-Cio aryl, 5- to 10-membered heteroaryl, -OR^Yt, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^YI-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R^YI is independently selected from hydrogen and Ci-Cô alkyl, or two instances of R^YI bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Ci-Cô alkyl, and Ci-Cô alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

3- to 6-membered heterocyclyl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl);

each Q is independently selected from:

Ci-Cô alkyl optionally substituted with 1-3 groups independently selected from: o halogen, o oxo, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen and -OCF3), and o C3-C8 cycloalkyl,

C3-C8 cycloalkyl optionally substituted with 1-3 groups independently selected from:

o halogen, ο CN, ο Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH2, and -NHCOMe), o Ci-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl), and o C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen and hydroxy), o Ci-Cô alkoxy optionally substituted with 1-4 groups independently selected from: halogen,

C3-C8 cycloalkyl (optionally substituted with CF3), o C3-C8 cycloalkyl (optionally substituted with 1-3 groups independently selected from halogen, CF3, OCF3, and Ci-Cô alkyl), and o Cô-Cio aryl,

5- to 10-membered heteroaryl optionally substituted with 1-3 groups independently selected from: o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3-to 10-membered heterocyclyl,

3- to 10-membered heterocyclyl optionally substituted with 1-3 groups independently selected from:

o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from oxo and C3-C8 cycloalkyl), and o oxo;

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -OR², -N(R²)2, CO2R², -CO-N(R²)2, -CN, phenyl, benzyl, Ci-Ci, alkoxy, C3-C8 cycloalkyl, 5- to 64 membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

each R² is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

/^□21 '^h^,.R²³ ^l ( θ ÿ (R²²)^

J^R⁷² J^'R²³ v ·

Z is selected from , ' , and , wherein

Ring C is selected from Cô-Cio aryl and 5- to 10-membered heteroaryl;

R²¹ is selected from hydrogen, -CN, Ci-C₆ alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^z2 is selected from hydrogen, halogen, and hydroxy, or R²* and R^Z2 taken together form a group selected from oxo and =N-OH;

each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, Ci-Cô haloalkyl, and Cô-Cio aryl; or two instances of R²³ are taken together to form a 3- to 6membered heterocyclyl;

n is selected from 4, 5, 6, 7, and 8; and m is selected from 0,1,2, and 3.

In some embodiments, X is -O-.

In some embodiments, each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, and -OR^Y1, wherein Q and R^Y1 are as defined above. In some embodiments, -OR^Y1is -OH.

In some embodiments, each Q is independently selected from C3-C8 cycloalkyl and CôCio aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Cô alkyl. In some embodiments, each Q is independently selected from:

In some embodiments, each R^v is independently selected from:

In some embodiments, Ring B is selected from C3-C8 cycloalkyl and phenyl optionally substituted with 1-3 groups independently selected from halogen. In some embodiments, Ring B is selected from:

In some embodiments, n is selected from 4, 5, and 6.

In some embodiments, -(Y)_n- is a group selected from:

In some embodiments, each R¹ is independently selected from Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -N(R²)2, and CO2R², wherein R² is as defined above. In some embodiments, each R¹ is independently selected from -CF₃, -NH₂, -NH(CH₂CH3), CO₂H, and CH2OH.

In some embodiments, each R² is independently selected from hydrogen and Ci-Cô alkyl.

X _RZ1 ( C 4s/ RZ2

In some embodiments, Z is selected from and , wherein R^Z1, ^C^rZ1

R^Z2

R^Z2, and Ring C are as defined above. In some embodiments, Z is , wherein R^Z1 and ^R^Z1 's/ ^rZ2

R^Z2 are as defined above. In some embodiments, Z is , wherein R^Z1 and R^z2 are as 'Cr²¹ •x/ R²² defined above. In some embodiments, Z is , wherein R^Z1 and R²² are as defined above.

VR^Z1

W<_RZ2 .

In some embodiments, Z is , wherein R^Z1 and R^Z2 are as defined above, and wherein (7?) refers to the stereochemical désignation of the central carbon atom under the Cahn-Ingold<R^Z1

R²²

Prelog convention. In some embodiments, Z is . , wherein R^Z1 and R^Z2 are as defïned above, and wherein (5) refers to the stereochemical désignation of the central carbon atom under the Cahn-Ingold-Prelog convention.

In some embodiments, the group:

In some embodiments, R^Z1 is selected from hydrogen and Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen). In some embodiments, R^Z1 is selected from hydrogen and -CF3. In some embodiments, R^zt is -CF3.

In some embodiments, R²² is hydroxy.

In some embodiments, Z is selected from:

In some embodiments, Z is

In some embodiments, m is selected from 1 and 2.

In some embodiments, compounds of the invention are encompassed by Formula I’

and deuterated dérivatives and pharmaceutically acceptable salts thereof, wherein:

X is -O-;

each Y is independently selected from -C(R^Y)2-, -O-, and each R^Y is independently selected from hydrogen and Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q);

Ring B is selected from C3-C8 cycloalkyl groups:

each Q is independently selected from C3-C8 cycloalkyl and Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Cô alkyl, each R¹ is independently selected from Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen) and -NH2;

R^Z1 is selected from Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen);

R^Z2 is hydroxy;

n is selected from 5 and 6; and m is 2.

In some embodiments, each Q of Formula l’is independently selected from:

In some embodiments, each R^Y of Formula l’is independently selected from:

In some embodiments, -(Y)_n- of Formula l’is a group selected from:

In some embodiments, R^Z1 in Formula l’is -CF3.

In some embodiments, Z in Formula l’is

In some embodiments, Z in

Formula l’is

In some embodiments, n in Formula l’is 5. In some embodiments, n in Formula l’is 6.

In some embodiments, compounds of the invention are encompassed by Formula I”:

X is selected from -O-, -S-, -SO-, and -SO2-;

( B j each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' f ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^YI, -COR^YI, -CON(R^Y1)2, and -NR^Y1-; _or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R^Y1 is independently selected from hydrogen and Ci-Cô alkyl, or two instances of R^YI bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

Ci-Cô alkyl optionally substituted with 1-3 groups independently selected from: o halogen,

Ο 0X0, ο Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen and -OCF3), and o C3-C8 cycloalkyl,

C3-C8 cycloalkyl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH2, and -NHCOMe), o Ci-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl), and o C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen and hydroxy), o Ci-Cô alkoxy optionally substituted with 1-4 groups independently selected from:

halogen,

C3-C8 cycloalkyl (optionally substituted with CF3), o C3-C8 cycloalkyl (optionally substituted with 1-3 groups independently selected from halogen, CF3, OCF3, and Ci-Ce alkyl), and o Cô-Cio aryl,

5- to 10-membered heteroaryl optionally substituted with 1-3 groups independently selected from:

o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3-to 10-membered heterocyclyl,

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)2, -CN, phenyl, benzyl, Ci-Cô alkoxy, Ci-Cô alkyl, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

R^Z1 is selected from hydrogen, -CN, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R^Z1 and R^Z2 taken together form a group selected from oxo and =N-0H;

each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CôCio aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl;

n is selected from 4, 5, 6, and 7; and m is selected from 0, 1,2, and 3.

In some embodiments, X in Formula I” is -O-.

In some embodiments, each Y in Formula I” is independently selected from -C(R^Y)2-, ( B 1

CO-, and ' ' , wherein R^Y and Ring B are as defined for Formula I”.

In some embodiments, each Y in Formula I” is -C(R^Y)2-, wherein R^Y is as defined for Formula I”.

Τη some embodiments, each R^Y in Formula I” is independently selected from hydrogen, Ci-Ce alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1, wherein R^Y1 and Q are as defrned for Formula I”.

In some embodiments, each R^Y in Formula I” is independently selected from:

CH₃ OH hydrogen, —1— , —1— ,

H₃C^,CH₃

OH , and

wherein Q is as defrned for Formula I”.

In some embodiments, each Q in Formula I” is independently selected from:

Cs-Cs cycloalkyl,

C6-C10 aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Ce alkyl.

In some embodiments, each Q in Formula I” is independently selected from:

In some embodiments, Ring B in Formula I” is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

In some embodiments, Ring B in Formula I” is selected from:

In some embodiments, each R* in Formula I” is independently selected from Ci-Cô alkyl 5 (optionally substituted with 1-3 groups independently selected from halogen) and -N(R²)2, wherein R² is as defined for Formula I”. In some embodiments, each R¹ in Formula I” is independently selected from -CF3 and -N(R²)2, wherein R² is as defined for Formula I”.

In some embodiments, each R² in Formula I” is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), and 10 Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy optionally substituted with 1-3 groups independently selected from halogen). In some embodiments, each R² in Formula I” is independently selected from hydrogen and Ci-Cô alkyl. In some embodiments, each R² in Formula I” is hydrogen.

R^Z1

R^Z2

In some embodiments, Z in Formula I” is , wherein R^ZI and R^Z2 are as defined

W*_RZ2 .

for Formula I”. In some embodiments, Z in Formula I” is , wherein R^Z1 and R²² are as defined for Formula I”. In some embodiments, Z in Formula I” is

In some embodiments, R^Z1 in Formula I” is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen). In some embodiments, R^Z1 in Formula I” is -CF₃.

In some embodiments, R²² in Formula I” is hydroxy.

In some embodiments, n in Formula I, I’, and/or I” is selected from 4, 5, and 6. In some embodiments, n in Formula I, I’, and/or I” is 5. In some embodiments, n in Formula I, I’, and/or I” is 6.

In some embodiments, m in Formula I, I’, and/or I” is selected from 1 and 2. In some embodiments, m in Formula I, I’, and/or I” is 1. In some embodiments, m in Formula I, I’, and/or I” is 2.

Another aspect of the invention provides pharmaceutical compositions comprising at least one compound chosen from the novel compounds disclosed herein, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing, and at least one pharmaceutically acceptable carrier, which compositions may further include at least one additional active pharmaceutical ingrédient. Thus, another aspect of the invention provides methods of treating the CFTR-mediated disease cystic fibrosis comprising administering at least one of compound chosen from the novel compounds disclosed herein, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing, and at least one pharmaceutically acceptable carrier, optionally as part of a pharmaceutical composition comprising at least one additional component, to a subject in need thereof.

In certain embodiments, the pharmaceutical compositions of the invention comprise at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, compositions comprising at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof may optionally further comprise at least one compound chosen from Compound II, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof.

Another aspect of the invention provides methods of treating the CFTR-mediated disease cystic fibrosis comprising administering to a patient in need thereof at least one compound chosen from the novel compounds disclosed herein, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing, and optionally further administering one or more additional CFTR modulating agents selected from (A)-l-(2,2-difluorobenzo[d][l,3]dioxol5-yl)-7V-(l-(2,3-dihydroxypropyl)-6-fluoro-2-(l-hydroxy-2-methylpropan-2-yl)-lZ/-indol-5yl)cyclopropanecarboxamide (Compound II), 7V-[2,4-bis(l,l-dimethylethyl)-5-hydroxyphenyl]l,4-dihydro-4-oxoquinoline-3-carboxamide (Compound III) or 7V-(2-(tert-butyl)-5-hydroxy-4-(2(methyl-d3)propan-2-yl-l,l,l,3,3,3-d6)phenyl)-4-oxo-l,4-dihydroquinoline-3-carboxamide (Compound ΙΠ-d), 3-(6-(1-(2,2-difluorobenzo[d][l,3]dioxol-5-yl)cyclopropane carboxamido)-3methylpyridin-2-yl)benzoic acid (Compound IV), JV-(l,3-dimethylpyrazol-4-yl)sulfonyl-6-[3(3,3,3-trifluoro-2,2-dimethyl-propoxy)pyrazol-1 -yl]-2-[(45)-2,2,4-trimethylpyrrolidin-1 yl]pyridine-3-carboxamide (Compound V), 7V-(benzenesulfonyl)-6-[3-[2-[l-(trifluoromethyl) cyclopropyl]ethoxy]pyrazol-l-yl]-2-[(4>S)-2,2,4-trimethylpyrrolidin-l-yl]pyridine-3-carboxamide (Compound VI), (145)-8-[3-(2-{dispiro[2.0.2.1]heptan-7-yl}ethoxy)-127-pyrazol-l-yl]-12,12dimethyl-2X⁶-thia-3,9,l 1,18,23-pentaazatetracyclo [17.3.1.111,14.05,10]tetracosa1(22),5,7,9,19(23),20-hexaene-2,2,4-trione (Compound VII), (117î)-6-(2,6-dimethylphenyl)-ll(2-methylpropyl)-12-{spiro[2.3]hexan-5-yl}-9-oxa-2X⁶-thia-3,5,12,19tetraazatricyclo[12.3.1.14,8]nonadeca-l(17),4(19),5,7,14(18),15-hexaene-2,2,13-trione (Compound VIII); 7V-(benzenesulfonyl)-6-(3-fluoro-5-isobutoxy-phenyl)-2-[(4S)-2,2,4trimethylpyrrolidin-l-yl]pyridine-3-carboxamide (Compound IX), and 7V-[(6-amino-2pyridyl)sulfonyl]-6-(3-fluoro-5-isobutoxy-phenyl)-2-[(45)-2,2,4-trimethylpyrrolidin-lyl]pyridine-3-carboxamide (Compound X).

Another aspect of the invention provides methods of treating the CFTR-mediated disease cystic fibrosis comprising administering to a patient in need thereof at least one compound chosen from the novel compounds disclosed herein, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing, and optionally further administering one or more additional CFTR modulating agents selected from:

N

O

N <^O

HN

(ASP-11), disclosed in Journal of Cystic

Fibrosis (2018), 17(5), 595-606, and:

(nesolicaftor or PTI-428), disclosed in WO 2016/105485.

In one embodiment, the additional CFTR modulating agent is ASP-11. In one embodiment, the additional CFTR modulating agent is PTI-428.

Another aspect of the invention provides methods of treating the CFTR-mediated disease 5 cystic fibrosis comprising administering to a patient in need thereof at least one compound chosen from the novel compounds disclosed herein, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing, and optionally further administering one or more additional CFTR modulating agents selected from:

O (galicaftor or ABB V-2222), disclosed in United States Patent

Application Publication No. 2016-0120841;

(ABBV-3221), disclosed in WO 2018/065921;

COoH

(posenacaftor or PTI-801), disclosed in WO 2017/062581; ABBV2851, disclosed in WO 2017/009804; GLPG2737, disclosed in United States Patent Application Publication No. 2017-0101405; ABBV-3748; ABBV-3903; and ABBV-119.

Another aspect of the invention provides compounds of Formulae I, I’, I”, I’”, la, lia, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof, for use in any of the methods described herein.

Brief Description of the Figures

FIG. 1 provides an X-ray power diffraction (XRPD) pattern of Compound 11 heptane solvaté.

FIG. 2 provides an overlay of X-ray power diffraction (XRPD) patterns of Compound 11 heptane solvaté prepared under three different drying conditions.

FIG. 3 provides a DSC analysis of Compound 11 heptane solvaté.

FIG. 4 provides a ¹³C solid-state NMR spectrum of Compound 11 heptane solvaté.

FIG. 5 provides a ¹⁹F solid-state NMR spectrum of Compound 11 heptane solvaté.

FIG. 6A provides a thermogravimetric analysis (TGA) curve for Compound 11 heptane solvaté (Drying Condition 1). FIG. 6B provides a thermogravimetric analysis (TGA) curve for Compound 11 heptane solvaté (Drying Condition 2). FIG. 6C provides a thermogravimetric analysis (TGA) curve for Compound 11 heptane solvaté (Drying Condition 3).

Définitions “Compound II” as used herein, refers to (R)-l-(2,2-difluorobenzo[d][l,3]dioxol-5-yl)-7V(1 -(2,3-dihydroxypropyl)-6-fluoro-2-(l -hydroxy-2-methylpropan-2-yl)- l//-indol-5yl)cyclopropanecarboxamide, which can be depicted with the following structure:

Compound II may be in the form of a pharmaceutically acceptable sait. Compound II and methods of making and using Compound II are disclosed in WO 2010/053471, WO 2011/119984, WO 2011/133751, WO 2011/133951, and WO 2015/160787, each incorporated herein by reference.

“Compound III” as used throughout this disclosure refers to 7V-(5-hydroxy-2,4-di-/ez7butyl-phenyl)-4-oxo-lH-quinoline-3-carboxamide which is depicted by the structure:

Compound III may also be in the form of a pharmaceutically acceptable sait. Compound III and methods of making and using Compound III are disclosed in WO 2006/002421, WO 2007/079139, WO 2010/108162, and WO 2010/019239, each incorporated herein by reference.

In some embodiments, a deuterated dérivative of Compound III (Compound III-d) is employed in the compositions and methods disclosed herein. A Chemical name for Compound III-d is7V-(2-(terf-butyl)-5-hydroxy-4-(2-(methyl-d3)propan-2-yl-l,l,l,3,3,3-d6)phenyl)-4-oxol,4-dihydroquinoline-3-carboxamide, as depicted by the structure:

Compound III-d may be in the form of a pharmaceutically acceptable sait. Compound III-d and methods of making and using Compound III-d are disclosed in WO 2012/158885, WO 2014/078842, and US Patent No. 8,865,902, incorporated herein by reference.

“Compound IV” as used herein, refers to 3-(6-(1-(2,2-difluorobenzo[d][l,3]dioxol-5yl)cyclopropanecarboxamido)-3-methylpyridin-2-yl)benzoic acid, which is depicted by the Chemical structure:

IV.

Compound IV may be in the form of a pharmaceutically acceptable sait. Compound IV and methods of making and using Compound IV are disclosed in WO 2007/056341, WO 2009/073757, and WO 2009/076142, incorporated herein by reference.

“Compound V” as used herein, refers to 7V-(l,3-dimethylpyrazol-4-yl)sulfonyl-6-[3(3,3,3-trifluoro-2,2-dimethyl-propoxy)pyrazol-l-yl]-2-[(45)-2,2,4-trimethylpynOlidin-lyl]pyridine-3-carboxamide, which is depicted by the Chemical structure:

Compound V may be in the form of a pharmaceutically acceptable sait. Compound V and methods of making and using Compound V are disclosed in WO 2018/107100 and WO 2019/113476, incorporated herein by reference.

“Compound VI” as used herein, refers to 7V-(benzenesulfonyl)-6-[3-[2-[l(trifluoromethyl) cyclopropyl]ethoxy]pyrazol-l-yl]-2-[(4S)-2,2,4-trimethylpyrrolidin-lyl]pyridine-3-carboxamide, which is depicted by the Chemical structure:

Compound VI may be in the form of a pharmaceutically acceptable sait. Compound VI and methods of making and using Compound VI are disclosed in WO 2018/064632, incorporated herein by reference.

“Compound VU” as used herein, refers to (145)-8-[3-(2-{dispiro[2.0.2.1]heptan-7yl} ethoxy)-17/-pyrazol-1 -yl] -12,12-dimethyl-2X⁶-thia-3,9,11,18,23-pentaazatetracyclo

[17.3.1.111,14.05,10]tetracosa-l(22),5,7,9,19(23),20-hexaene-2,2,4-trione, which is depicted by the Chemical structure:

Compound VII may be in the form of a pharmaceutically acceptable sait. Compound VII and 5 methods of making and using Compound VII are disclosed in WO 2019/152940 and United

States Provisional Patent Application No. 62/886,660, incorporated herein by reference.

“Compound VIII” as used herein, refers to (117?)-6-(2,6-dimethylphenyl)-l l-(2methylpropyl)-12-{spiro[2.3]hexan-5-yl}-9-oxa-2X⁶-thia-3,5,12,19tetraazatricyclo[12.3.1.14,8]nonadeca-l(17),4(19),5,7,14(18),15-hexaene-2,2,13-trione, which is depicted by the Chemical structure:

Compound VIII may be in the form of a pharmaceutically acceptable sait. Compound VIII and methods of making and using Compound VIII are disclosed in PCT/US2020/026331, incorporated herein by reference.

“Compound IX” as used herein, refers to 7V-(benzenesulfonyl)-6-(3-fluoro-5-isobutoxyphenyl)-2-[(4₁S)-2,2,4-trimethylpyrrolidin-l-yl]pyridine-3-carboxamide, which is depicted by the Chemical structure:

Compound IX may be in the form of a pharmaceutically acceptable sait. Compound IX and methods of making and using Compound IX are disclosed in WO 2016/057572, incorporated herein by reference.

“Compound X” as used herein, refers to 7V-[(6-amino-2-pyridyl)sulfonyl]-6-(3-fluoro-5isobutoxy-phenyl)-2-[(4S)-2,2,4-trimethylpyrrolidin-l-yl]pyridine-3-carboxamide, which is depicted by the Chemical structure:

F X.

Compound X may be in the form of a pharmaceutically acceptable sait. Compound X and methods of making and using Compound X are disclosed in WO 2016/057572, incorporated herein by reference.

As used herein, the term “alkyl” refers to a saturated, branched or unbranched aliphatic hydrocarbon containing carbon atoms (such as, for example, 1, 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 carbon atoms). Alkyl groups may be substituted or unsubstituted.

As used herein, the term “pi bond” refers to a covalent bond formed by the p orbitals of adjacent atoms. Pi bonds exist where there is a multiple bond, i.e., a double or triple bond, between two atoms. For example, a carbon-carbon double bond consists of one pi bond, and a carbon-carbon triple bond consists of two pi bonds.

As used herein, the term “haloalkyl group” refers to an alkyl group substituted with one or more halogen atoms.

The term “alkoxy” as used herein refers to an alkyl or cycloalkyl covalently bonded to an oxygen atom. Alkoxy groups may be substituted or unsubstituted.

As used herein, the term “haloalkoxyl group” refers to an alkoxy group substituted with one or more halogen atoms.

As used herein, “cycloalkyl” refers to a cyclic, bicyclic, tricyclic, or polycyclic nonaromatic hydrocarbon groups having 3 to 12 carbons (such as, for example 3-10 carbons). “Cycloalkyl” groups encompass monocyclic, bicyclic, tricyclic, bridged, fused, and spiro rings, including mono spiro and dispiro rings. Non-limiting examples of cycloalkyl groups are 23 cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, adamantyl, norbomyl, and dispiro[2.0.2.1]heptane. Cycloalkyl groups may be substituted or unsubstituted.

The term “heteroaryl ring” as used herein refers to an aromatic ring comprising at least one ring atom that is a heteroatom, such as O, N, or S.

As used herein, the terms “heterocyclyl ring” and “heterocyclyl” refer to a non-aromatic hydrocarbon containing 3 to 12 atoms in a ring (such as, for example 3-10 atoms) comprising at least one ring atom that is a heteroatom, such as O, N, S, or Si. “Heterocyclyl” rings encompass monocyclic, bicyclic, tricyclic, polycyclic, bridged, fused, and spiro rings, including mono spiro and dispiro rings.

“Substituted” indicates that at least one hydrogen of the “substituted” group is replaced by a substituent. Unless otherwise indicated, an “optionally substituted” group may hâve a suitable substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted with more than one substituent chosen from a specified group, the substituent may be either the same or different at each position.

Examples of protecting groups for nitrogen include, for example, Z-butyl carbamate (Boc), benzyl (Bn),para-methoxybenzyl (PMB), tetrahydropyranyl (THP), 9-fluorenylmethyl carbamate (Fmoc), benzyl carbamate (Cbz), methyl carbamate, ethyl carbamate, 2,2,2trichloroethyl carbamate (Troc), 2-trimethylsilylethyl carbamate (Teoc), allyl carbamate (Aloc or Alloc), formamide, acetamide, benzamide, allylamine, trifluoroacetamide, triphenylmethylamine, benzylideneamine, and p-toluenesulfonamide. A comprehensive list of nitrogen protecting groups can be found in Wuts, P. G. M. “Greene’s Protective Groups in Organic Synthesis: Fifth Edition,” 2014, John Wiley and Sons.

As used herein, “deuterated derivative(s)” means the same Chemical structure, with one or more hydrogen atoms replaced by a deuterium atom. In some embodiments, the deuterated dérivatives are compounds where one or more hydrogen atoms of an alkyl group are replaced by a deuterium atom.

As used herein, “CFTR” means cystic fîbrosis transmembrane conductance regulator.

As used herein, the term “CFTR modulator” refers to a compound that increases the activity of CFTR. The increase in activity resulting from a CFTR modulator includes but is not limited to compounds that correct, potentiate, stabilize and/or amplify CFTR.

As used herein, the term “CFTR corrector” refers to a compound that facilitâtes the processing and trafficking of CFTR to increase the amount of CFTR at the cell surface.

As used herein, the term “CFTR potentiator” refers to a compound that increases the channel activity of CFTR protein located at the cell surface, resulting in enhanced ion transport. The novel compounds disclosed herein are CFTR potentiators.

As used herein, the term “CFTR potentiator enhancer”, “CFTR potentiation enhancer”, and “CFTR co-potentiator” are used interchangeably and refer to a compound that enhances CFTR potentiation.

As used herein, the term “active pharmaceutical ingrédient” (“API”) or “therapeutic agent” refers to a biologically active compound.

As used herein, the term “one or more additional therapeutic agent(s) comprise(s),” includes the possibility that there is only one therapeutic agent.

The terms “patient” and “subjecf ’ are used interchangeably and refer to an animal including humans.

The terms “effective dose” and “effective amount” are used interchangeably herein and refer to that amount of a compound that produces the desired effect for which it is administered (e.g., improvement in CF or a symptom of CF, or lessening the severity of CF or a symptom of CF). The exact amount of an effective dose will dépend on the purpose of the treatment and will be ascertainable by one skilled in the art using known techniques (see, e.g., Lloyd (1999) The Art, Science and Technology of Pharmaceutical Compounding).

As used herein, the terms “treatment,” “treating,” and the like generally mean the improvement in one or more symptoms of CF or lessening the severity of CF or one or more symptoms of CF in a subject. “Treatment,” as used herein, includes, but is not limited to, the following: increased growth of the subject, increased weight gain, réduction of mucus in the lungs, improved pancreatic and/or liver function, réduction of chest infections, and/or réductions in coughing or shortness of breath. Improvements in or lessening the severity of any of these symptoms can be readily assessed according to standard methods and techniques known in the art.

As used herein, the term “in combination with,” when referring to two or more compounds, agents, or additional active pharmaceutical ingrédients, means the administration of two or more compounds, agents, or active pharmaceutical ingrédients to the patient prior to, concurrent with, or subséquent to each other.

The terms “about” and “approximately”, when used in connection with doses, amounts, or weight percent of ingrédients of a composition or a dosage form, include the value of a specified dose, amount, or weight percent or a range of the dose, amount, or weight percent that is recognized by one of ordinary skill in the art to provide a pharmacological effect équivalent to that obtained from the specified dose, amount, or weight percent. The terms “about” and “approximately” may refer to an acceptable error for a particular value as determined by one of skill in the art, which dépends in part on how the values is measured or determined. In some embodiments, the terms “about” and “approximately” mean within 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, or 0.5% of a given value or range. As used herein, the symbol appearing immediately before a numerical value has the same meaning as the terms “about” and “approximately.”

As used herein, the term “solvent” refers to any liquid in which the product is at least partially soluble (solubility of product >1 g/L).

As used herein, the term “room température” or “ambient température” means 15 °C to 30 °C.

It will be appreciated that certain compounds of this invention may exist as separate stereoisomers or enantiomers and/or mixtures of those stereoisomers or enantiomers. As used in the Chemical structures disclosed herein, a “wedge” (^) or “hash” (··*'' ) bond to a stereogenic atom indicates a chiral center of known absolute stereochemistry (i.e., one stereoisomer). As used in the Chemical structures disclosed herein, a “wavy” bond ( ) to a stereogenic atom indicates a chiral center of unknown absolute stereochemistry (i.e., one stereoisomer). As used in the Chemical structures disclosed herein, a “wavy” bond ( ) to a double-bonded carbon indicates a mixture of E/Z isomers. As used in the Chemical structures disclosed herein, a s'' (“straight”) bond to a stereogenic atom indicates where there is a mixture (e.g., a racemate or enrichment). As used herein, two (“straight”) bonds to a double-bonded carbon indicates that the double bond possesses the ΕΓΖ stereochemistry as drawn. As used in the Chemical

V^A structures disclosed herein, a ' (a “wavy” line perpendicular to a “straight” bond to group “A”) indicates that group “A” is a substituent whose point of attachment is at the end of the bond that terminâtes at the ”wavy” line. As used herein, a stereogenic atom that is notated with an (R) or (5) indicates the stéréo Chemical désignation of the stereogenic atom under the Cahn-IngoldPrelog convention.

Certain compounds disclosed herein may exist as tautomers and both tautomeric forms are intended, even though only a single tautomeric structure is depicted. For example, a description of Compound A is understood to include its tautomer Compound B and vice versa, as well as mixtures thereof:

Compound A Compound B

As used herein, “minimal fonction (MF) mutations” refer to CFTR gene mutations associated with minimal CFTR fonction (little-to-no fonctioning CFTR protein) and include, for example, mutations associated with severe defects in ability of the CFTR channel to open and close, known as defective channel gating or “gating mutations”; mutations associated with severe defects in the cellular processing of CFTR and its delivery to the cell surface; mutations associated with no (or minimal) CFTR synthesis; and mutations associated with severe defects in channel conductance.

As used herein, the term “pharmaceutically acceptable sait” refers to a sait form of a compound of this disclosure wherein the sait is nontoxic. Pharmaceutically acceptable salts of the compounds of this disclosure include those derived from suitable inorganic and organic acids and bases. A “free base” form of a compound, for example, does not contain an ionically bonded sait.

The phrase “and pharmaceutically acceptable salts and deuterated dérivatives thereof’ is used interchangeably with “and pharmaceutically acceptable salts thereof and deuterated dérivatives of any of the forgoing” in reference to one or more compounds or formulae of the invention. These phrases are intended to encompass pharmaceutically acceptable salts of any one of the referenced compounds, deuterated dérivatives of any one of the referenced compounds, and pharmaceutically acceptable salts of those deuterated dérivatives.

One of ordinary skill in the art would recognize that, when an amount of “a compound or a pharmaceutically acceptable sait thereof’ is disclosed, the amount of the pharmaceutically acceptable sait form of the compound is the amount équivalent to the concentration of the free base of the compound. It is noted that the disclosed amounts of the compounds or their pharmaceutically acceptable salts thereof herein are based upon their free base form.

Suitable pharmaceutically acceptable salts are, for example, those disclosed in S. M. Berge, et al. J. Pharmaceutical Sciences, 1977, 66, 1-19. For example, Table 1 of that article provides the following pharmaceutically acceptable salts:

Table 1:

Acetate	lodide	Benzathine
Benzenesulfonate	Isethionate	Chloroprocaine
Benzoate	Lactate	Choline
Bicarbonate	Lactobionate	Diethanolamine
Bitartrate	Malate	Ethylènediamine
Bromide	Maleate	Meglumine
Calcium edetate	Mandelate	Procaine
Camsylate	Mesylate	Aluminum
Carbonate	Methylbromide	Calcium
Chloride	Methylnitrate	Lithium
Citrate	Methylsulfate	Magnésium
Dihydrochloride	Mucate	Potassium
Edetate	Napsylate	Sodium
Edisylate	Nitrate	Zinc
Estolate	Pamoate (Embonate)
Esylate	Pantothenate
Fumarate	Phosphate/diphosphate
Gluceptate	Polygalacturonate
Gluconate	Salicylate
Glutamate	Stéarate
Glycollylarsanilate	Subacetate
Hexylresorcinate	Succinate
Hydrabamine	Sulfate
Hydrobromide	Tannate
Hydrochloride	Tartrate
Hydroxynaphthoate	Teociate
	Triethiodide

Non-limiting examples of pharmaceutically acceptable acid addition salts include: salts formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, or perchloric acid; salts formed with organic acids, such as acetic acid, oxalic acid, 5 maleic acid, tartaric acid, citric acid, succinic acid or malonic acid; and salts formed by using other methods used in the art, such as ion exchange. Non-limiting examples of pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate,

0 glycérophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stéarate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, and valerate

5 salts. Pharmaceutically acceptable salts derived from appropriate bases include alkali métal, alkaline earth métal, ammonium, and N⁺(Ci-4alkyl)4 salts. This disclosure also envisions the quatemization of any basic nitrogen-containing groups of the compounds disclosed herein. Suitable non-limiting examples of alkali and alkaline earth métal salts include sodium, lithium, potassium, calcium, and magnésium. Further non-limiting examples of pharmaceutically acceptable salts include ammonium, quatemary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate. Other suitable, non-limiting examples of pharmaceutically acceptable salts include besylate and glucosamine salts.

As used herein, the terni “amorphous” refers to a solid material having no long-range order in the position of its molécules. Amorphous solids are generally supercooled liquids in which the molécules are arranged in a random manner so that there is no well-defmed arrangement, e.g., molecular packing, and no long-range order. Amorphous solids are generally isotropie, i.e., exhibit similar properties in ail directions and do not hâve definite melting points. For example, an amorphous material is a solid material having no sharp characteristic crystalline peak(s) in its X-ray power diffraction (XRPD) pattern (i.e., is not crystalline as determined by XRPD). Instead, one or several broad peaks (e.g., halos) appear in its XRPD pattern. Broad peaks are characteristic of an amorphous solid. See, US 2004/0006237 for a comparison of XRPDs of an amorphous material and crystalline material. In some embodiments, a solid material may comprise an amorphous compound, and the material may, for example, be characterized by a lack of sharp characteristic crystalline peak(s) in its XRPD spectrum (i.e., the material is not crystalline, but is amorphous, as determined by XRPD). Instead, one or several broad peaks (e.g., halos) may appear in the XRPD pattern of the material. See US 2004/0006237 for a comparison of XRPDs of an amorphous material and crystalline material. A solid material, comprising an amorphous compound, may be characterized by, for example, a glass transition température which is lower than the melting point of a pure crystalline solid. Other techniques, such as, for example, solid State NMR may also be used to characterize crystalline or amorphous forms.

As used herein, the terms “crystal form,” “crystalline form,” and “Form” interchangeably refer to a crystal structure (or polymorph) having a particular molecular packing arrangement in the crystal lattice. Crystalline forms can be identified and distinguished from each other by one or more characterization techniques including, for example, X-ray powder diffraction (XRPD), single crystal X-ray diffraction, and ^,3C solid State nuclear magnetic résonance (¹³C SSNMR). Accordingly, as used herein, the terms “crystalline Form [X] of Compound I” refer to unique crystalline forms that can be identified and distinguished from other crystalline forms by one or more characterization techniques including, for example, XRPD, single crystal X-ray diffraction, and ¹³C SSNMR. In some embodiments, the novel crystalline forms are characterized by an Xray powder diffractogram having one or more signais at one or more specified two-theta values (°20).

As used herein, the term “free form” refers to a non-ionized version of the compound in the solid State. Examples of free forms include free bases and free acids.

As used herein, the term “solvaté” refers to a crystal form comprising one or more molécules of a compound of the présent disclosure and, incorporated into the crystal lattice, one or more molécules of a solvent or solvents in stoichiometric or nonstoichiometric amounts. When the solvent is water, the solvaté is referred to as a “hydrate.”

In some embodiments, a solid material may comprise a mixture of crystalline solids and amorphous solids. A solid material comprising an amorphous compound may also, for example, contain up to 30% of a crystalline solid. In some embodiments, a solid material prepared to comprise an amorphous compound may also, for example, contain up to 25%, 20%, 15%, 10%, 5%, or 2% of a crystalline solid. In embodiments wherein the solid material contains a mixture of crystalline solids and amorphous solids, the characterizing data, such as XRPD, may contain indicators of both crystalline and amorphous solids. In some embodiments, a crystalline form of this disclosure may contain up to 30% amorphous compound. In some embodiments, a crystalline préparation of a compound of Formula I may contain up to 25%, 20%, 15%, 10%, 5%, or 2% of an amorphous solid.

As used herein, the term substantially amorphous refers to a solid material having little or no long-range order in the position of its molécules. For example, substantially amorphous materials hâve less than 15% crystallinity (e.g., less than 10% crystallinity, less than 5% crystallinity, or less than 2% crystallinity). It is also noted that the term “substantially amorphous” includes the descriptor, “amorphous,” which refers to materials having no (0%) crystallinity.

As used herein, the term substantially crystalline refers to a solid material having little or no amorphous molécules. For example, substantially crystalline materials hâve less than 15% amorphous molécules (e.g., less than 10% amorphous molécules, less than 5% amorphous molécules, or less than 2% amorphous molécules). It is also noted that the term “substantially crystalline” includes the descriptor “crystalline,” which refers to materials that are 100% crystalline form.

As used herein, a crystalline form is substantially pure when it accounts for an amount by weight equal to or greater than 90% of the sum of ail solid form(s) in a sample as determined by a method in accordance with the art, such as quantitative XRPD. In some embodiments, the solid form is substantially pure when it accounts for an amount by weight equal to or greater than 95% of the sum of ail solid form(s) in a sample. In some embodiments, the solid form is substantially pure when it accounts for an amount by weight equal to or greater than 99% of the sum of ail solid form(s) in a sample. It is also noted that the tenu “substantially pure” includes the descriptor “pure,” which refers to materials that are 100% pure.

As used herein, the term “XRPD” refers to the analytical characterization method of Xray powder diffraction. XRPD patterns disclosed herein were recorded at ambient conditions in transmission or reflection geometry using a diffractometer.

As used herein, the term “ambient conditions” means room température, open air condition and uncontrolled humidity condition. The terms “room température” and “ambient température” mean 15 °C to 30 °C.

As used herein, the terms “X-ray powder diffractogram,” “X-ray powder diffraction pattern,” “XRPD pattern,” “XRPD spectrum” interchangeably refer to an experimentally obtained pattern plotting signal positions (on the abscissa) versus signal intensities (on the ordinate). For an amorphous material, an X-ray powder diffractogram may include one or more broad signais; and for a crystalline material, an X-ray powder diffractogram may include one or more signais, each identified by its angular value as measured in degrees 20 (° 20), depicted on the abscissa of an X-ray powder diffractogram, which may be expressed as “a signal at... degrees two-theta,” “a signal at [a] two-theta value(s)of...” and/or “a signal at at least... twotheta value(s) selected from ....”

A “signal” or “peak” as used herein refers to a point in the XRPD pattem where the intensity as measured in counts is at a local maximum. One of ordinary skill in the art would recognize that one or more signais (or peaks) in an XRPD pattem may overlap and may, for example, not be apparent to the naked eye. Indeed, one of ordinary skill in the art would recognize that some art-recognized methods are capable of and suitable for determining whether a signal exists in a pattem, such as Rietveld refmement.

As used herein, “a signal at... degrees two-theta” refer to X-ray reflection positions as measured and observed in X-ray powder diffraction experiments (° 20).

The repeatability of the measured angular values is in the range of ±0.2° 20, i.e., the angular value can be at the recited angular value + 0.2 degrees two-theta, the angular value - 0.2 31 degrees two-theta, or any value between those two end points (angular value +0.2 degrees twotheta and angular value -0.2 degrees two-theta).

The terms “signal intensifies” and “peak intensities” interchangeably refer to relative signal intensities within a given X-ray powder diffractogram. Factors that can affect the relative signal or peak intensities include sample thickness and preferred orientation (e.g., the crystalline particles are not distributed randomly).

As used herein, an X-ray powder diffractogram is “substantially similar to that in [a particular] Figure” when at least 90%, such as at least 95%, at least 98%, or at least 99%, of the signais in the two diffractograms overlap. In determining “substantial similarity,” one of ordinary skill in the art will understand that there may be variation in the intensities and/or signal positions in XRPD diffractograms even for the same crystalline form. Thus, those of ordinary skill in the art will understand that the signal maximum values in XRPD diffractograms (in degrees two-theta) generally mean that value is identifïed as ±0.2 degrees two-theta of the reported value, an art-recognized variance.

As used herein, a solid State nuclear magnetic résonance (SSNMR) spectrum is “substantially similar to that in [a particular] Figure” when at least 90%, such as at least 95%, at least 98%, or at least 99%, of the signais in the two spectra overlap. In determining “substantial similarity,” one of ordinary skill in the art will understand that there may be variation in the intensities and/or signal positions in SSNMR spectra even for the same crystalline form. Thus, those of ordinary skill in the art will understand that the Chemical shifts in SSNMR spectra (in parts per million (ppm) referred to herein) generally mean that value is identifïed as ± 0.2 ppm of the reported value, an art-recognized variance.

The term “X-ray powder diffractogram having a signal at... two-theta values” as used herein refers to an XRPD pattern that contains X-ray reflection positions as measured and observed in X-ray powder diffraction experiments (° two-theta).

As used herein, the term “DSC” refers to the analytical method of Differential Scanning Calorimetry.

As used herein, the term “onset of décomposition” refers to the intersection point of the baseline before transition and the interflection tangent.

As used herein, the term “glass transition température” or “Tg” refers to the température above which a hard and brittle “glassy” amorphous solid becomes viscous or rubbery.

As used herein, the term “TGA” refers to the analytical method of Thermo Gravimétrie (or thermogravimetric) Analysis.

Detailed Description of Embodiments

In addition to compounds of Formula I, I’, and I”, pharmaceutically acceptable salts thereof, and deuterated dérivatives of those compounds and salts, the invention provides compounds of Formulae I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof.

For example, in some embodiments, the compound of Formula I is selected from compounds of Formula la:

(b J each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Ce alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^YI, -COR^YI, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R^Y1 is independently selected from hydrogen and Ci-Cr, alkyl, or two instances of R^Y1 bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

halogen,

5- to 10-membered heteroaryl optionally substituted with 1-3 groups independently selected from: o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3- to 10-membered heterocyclyl,

each R² is independently selected from hydrogen, Ci-Ce alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Ciq aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

R^z* is selected from hydrogen, -CN, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^z2 is selected from hydrogen, halogen, and hydroxy, or R^Z1 and R^Z2 taken together form a group selected from oxo and =N-OH;

each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CôC10 aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl; and n is selected from 4, 5, 6, and 7.

In some embodiments, X in Formula la is -O-.

In some embodiments, each Y in Formula la is independently selected from -C(R^Y)2-, ( B 2

CO-, and ' ' , wherein R^Y and Ring B are as defined for Formula la.

In some embodiments, each Y in Formula la is -C(R^Y)2-, wherein R^Y is as defined for Formula la.

In some embodiments, each R^Y in Formula la is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1, wherein Q and R^YI are as defined for Formula la.

In some embodiments, each R^Y in Formula la is independently selected from:

CH₃ OH hydrogen, -~L- , —1— ,

In some embodiments, each Q in Formula la is independently selected from:

C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Cô alkyl.

In some embodiments, each Q in Formula la is independently selected from:

In some embodiments, Ring B in Formula la is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

In some embodiments, Ring B in Formula la is selected from:

In some embodiments, -(Y)_n- in Formula la is a group selected from:

In some embodiments, each R¹ in Formula la is independently Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen) and -N(R²)2, wherein R² is as defined for Formula la. In some embodiments, each R¹ in Formula la is independently selected from -CF3 and -N(R²)2, wherein R² is as defined for Formula la.

In some embodiments, each R² in Formula la is independently selected from hydrogen,

C1-C6 alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Ce alkoxy, which is optionally substituted with 1-3 groups independently selected from halogen). In some embodiments, each R² in Formula la is independently selected from hydrogen and Ci-Cf, alkyl. In some embodiments, each R² in

Formula la is hydrogen.

*^v^rZ1

R⁷²

In some embodiments, Z in Formula la is , wherein R^Z1 and R^Z2 are as defined for Formula la.

In some embodiments, R^Z1 in Formula la is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen). In some embodiments, R^ZI in Formula la is -CF₃.

In some embodiments, R^Z2 in Formula la is hydroxy.

In some embodiments, n in Formula la is selected from 4, 5, and 6. In some embodiments, n in Formula la is 6.

In some embodiments, the compound of Formula I is selected from compounds of Formulae

Ha, Ilb, and Ile:

X is selected from -O-, -S-, -SO-, and -SO2-;

( B ) each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^YI, -CON(R^YI)2, and -NR^YI-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R^Y1 is independently selected from hydrogen and Ci-Cô alkyl, or two instances of R^Y1 bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

o halogen, .

o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH2, and -NHCOMe), o Ci-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl), and o C3-C8 cycloalkyl,

halogen,

C3-C8 cycloalkyl (optionally substituted with CF3), o C₃-C₈ cycloalkyl (optionally substituted with 1-3 groups independently selected from halogen, CF3, OCF3, and Ci-Cô alkyl), and o Cô-Cio aryl,

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R^Z1 and R²² taken together form a group selected from oxo and =N-OH;

each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CôCio aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl; and m is selected from 0, 1,2, and 3.

In some embodiments, m in Formulae Ha, Hb, or Hc is selected from 1 and 2. In some embodiments, m in Formulae Ha, Ilb, or Ile is 2.

Ild, Ile, and Ilf:

( B j each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

Ring B is selected from:

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Ci-Ce alkyl, and Ci-Cô alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)₂, -CN, phenyl, benzyl, Ci-C₆ alkoxy, Ci-C₆ alkyl, C₃-C₈ cycloalkyl, 5- to 642 membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)₂;

each R² is independently selected from hydrogen, Ci-Cf, alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R^Z1 and R^Z2 taken together form a group selected from oxo and =N-OH; and each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CôC10 aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl.

In some embodiments, X in Formulae Ha, Ilb, Ile, Hd, Ile, or Ilf is -O-.

In some embodiments, each Y in Formulae Ha, Ilb, Ile, Ild, Ile, or Ilf is independently ( ^B ) selected from -C(R^Y)2-, -CO-, and V / , wherein R^Y and Ring B are as defined for Formulae Ha, Ilb, Ile, Hd, Ile, or Ilf.

In some embodiments, each Y in Formulae Ha, Hb, Ile, Hd, Ile, or Ilf is -C(R^Y)2-, wherein R^Y is as defined for Formulae Ha, Hb, Ile, Hd, Ile, or Hf.

In some embodiments, each R^Y in Formulae Ha, Hb, Hc, Ild, Ile, or Hf is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1, wherein Q and R^Y1 are as defined for Formulae Ha, Ilb, Ile, Ild, Ile, or Ilf.

In some embodiments, each R^Y in Formulae Ha, Hb, Hc, Hd, Ile, or Hf is independently selected from:

H₃C. ,CH₃ ,OH

CH₃ OH ³ <

hydrogen, -4— , -4~~ , -4— , -4— , and —L- , wherein Q is as defined for Formulae Ha, Hb, Ile, Hd, Ile, or Hf.

In some embodiments, each Q in Formulae Ha, Hb, Hc, Hd, Ile, or Hf is independently selected from:

C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Ce alkyl.

In some embodiments, each Q in Formulae Ha, Hb, Ile, Hd, Ile, or Hf is independently selected from:

In some embodiments, Ring B in Formulae lia, Hb, Ile, Hd, Ile, or Hf is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

In some embodiments, Ring B in Formulae Ha, Hb, Hc, Hd, Ile, or Hf is selected from:

F

In some embodiments, each R¹ in Formulae Ha, Hb, Ile, Hd, Ile, or Hf is independently Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen) and N(R²)2 wherein R² is as defïned for Formulae Ha, Ilb, Ile, Ild, Ile, or Hf. In some embodiments, each R¹ in Formulae Ha, Hb, Hc, Hd, Ile, or Hf is independently selected from CF3 and -N(R²)2 wherein R² is as defïned for Formulae Ha, Hb, Hc, Hd, Ile, or Hf.

In some embodiments, each R² in Formulae Ha, Hb, Ile, Hd, Ile, or Hf is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-3 groups independently selected from halogen). In some embodiments, each R² in Formulae Ha, Hb, Hc, Hd, Ile, or Hf is independently selected from hydrogen and Ci-C₆ alkyl. In some embodiments, each R² in Formulae Ha, Hb, Hc, Hd, Ile, or Hf is hydrogen.

R^Z1

R²²

In some embodiments, Z in Formulae Ha, Ilb, Ile, Ild, Ile, or Hf is , wherein

R^Z1 and R²² are as defined for Formulae Ha, Hb, Hc, Ild, Ile, or Hf.

In some embodiments, R^Z1 in Formulae Ha, Hb, Hc, Ild, Ile, or Hf is selected from CiCô alkyl (optionally substituted with 1-3 groups selected from halogen). In some embodiments, R^ZI in Formulae Ha, Hb, Hc, Hd, Ile, or Ilf is -CF3.

In some embodiments, R^Z2 in Formulae Ha, Hb, Hc, Ild, Ile, or Hf is hydroxy.

Ilia, HIb, and Hic:

X is selected from -O-, -S-, -SO-, and -SO2-;

each Y is independently selected from -C(R^V)2-, -O-, -CO-, and each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

halogen,

5- to 10-membered heteroaryl optionally substituted with 1-3 groups independently selected from: o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o Cj-Cs cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3- to 10-membered heterocyclyl,

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)₂, -CN, phenyl, benzyl, Ci-Cô alkoxy, Ci-Cô alkyl, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2î each R² is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

R^ZI is selected from hydrogen, -CN, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

In some embodiments, m in Formulae Ilia, IHb, or Hic is selected from 1 and 2. In some embodiments, m in Formulae Ilia, IHb, and IIIc is 2.

Illd, IHe, and IHf:

X is selected from -O-, -S-, -SO-, and -SO2-;

‘ L ^B J each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' * ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CÛ2R^Y1, -COR^YI, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

Ci-Cô alkyl optionally substituted with 1-3 groups independently selected from: o halogen, o oxo, o Cô-Cio aryl (optionally substituted with 1 -3 groups independently selected from halogen and -OCF3), and o C3-C8 cycloalkyl,

o halogen, ο CN, ο Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH2, and -NHCOMe), o Ci-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Ce alkyl), and o C3-C8 cycloalkyl,

5- to 10-membered heteroaryl optionally substituted with 1 -3 groups independently selected from: o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3- to 10-membered heterocyclyl,

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)₂, -CN, phenyl, benzyl, Ci-Cô alkoxy, Ci-Cô alkyl, C3-C8 cycloalkyl, 5- to 649 membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)₂;

each R² is independently selected from hydrogen, Ci-Ce alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Ce-Cio aryl (optionally substituted with Ci-Ce alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

-<.R^Z1 -Cr^Z3 'S ÛO

V^R²² x/^Sl'R^Z3 V^R^{Z1 X} Jz2

Z is selected from , ' , and ^R ;

R^Z1 is selected from hydrogen, -CN, Ci-Ce alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R^ZI and R^Z2 taken together form a group selected from oxo and =N-OH;

each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CâC10 aryl; or two instances of R⁷³ are taken together to form a 3- to 6-membered heterocyclyl.

In some embodiments, X in Formulae Ilia, IHb, IIIc, Hld, IHe, or Hlf is -O-.

In some embodiments, each Y in Formulae Ilia, IHb, IIIc, Hld, Ille, or IHf is

L ^B J independently selected from -C(R^Y)₂-, -CO-, and \ ' , wherein R^Y and Ring B as defined for Formulae Ilia, IHb, IIIc, Hld, IHe, or IHf.

In some embodiments, each Y in Formulae Ilia, IHb, IIIc, Hld, IHe, or Hlf is -C(R^Y)2-, wherein R^Y is as defined for Formulae Ilia, IHb, IIIc, Hld, Ille, or IHf.

In some embodiments, each R^Y in Formulae Ilia, IHb, IIIc, Hld, Ille, or IHf is independently selected from hydrogen, Ci-C₆ alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1, wherein Q and R^YI are as defined for Formulae Ilia, IHb, IIIc, Hld, Ille, or IHf.

In some embodiments, each R^Y in Formulae Ilia, IHb, IIIc, Hld, Ille, or Hlf is independently selected from:

ch₃ oh hydrogen, -J— , —

wherein Q is as defined for Formulae Ilia, IHb, IIIc, IHd, IHe, or IHf.

In some embodiments, each Q in Formulae Ilia, IHb, IIIc, IHd, IHe, or IHf is independently selected from:

C3-C8 cycloalkyl,

In some embodiments, Ring B in Formulae HIa, HIb, IIIc, IHd, IHe, or Hlf is selected from Ce-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

In some embodiments, Ring B in Formulae HIa, IHb, IIIc, IHd, IHe, or IHf is selected from:

In some embodiments, each R¹ in Formulae HIa, HIb, IIIc, IHd, IHe, or Hlf is independently Ci-Ce alkyl (optionally substituted with 1-3 groups independently selected from halogen) and -N(R²)2, wherein R² is as defined for Formulae HIa, HIb, IIIc, IHd, IHe, or IHf. In some embodiments, each R¹ in Formulae HIa, IHb, IIIc, IHd, IHe, or IHf is independently selected from -CF3 and -N(R²)2, wherein R² is as defined for Formulae HIa, HIb, IIIc, IHd, HIe, or IHf.

In some embodiments, each R² in Formulae HIa, HIb, IIIc, IHd, HIe, or Hlf is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-3 groups independently selected from halogen). In some embodiments, each R² in Formulae HIa, HIb, IIIc, IHd, HIe, or IHf is independently selected from hydrogen and Ci-Cô alkyl.

In some embodiments, each R² in Formulae Ilia, Illb, IIIc, Illd, Ille, or Illf is hydrogen.

'Cr²¹ /^_rz2

In some embodiments, Z in Formulae Ilia, Illb, IIIc, Illd, Ille, or Illf is , wherein Z is as defined for Formulae Ilia, Illb, IIIc, Illd, Ille, or Illf.

In some embodiments, R^Z1 in Formulae Ilia, Illb, IIIc, Illd, Ille, or Illf is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen). In some embodiments, R^ZI in Formulae Ilia, Illb, IIIc, Illd, Ille, or Illf is -CF3.

In some embodiments, R^Z2 in Formulae Ilia, Illb, IIIc, Illd, Ille, or Illf is hydroxy.

In some embodiments, the compound of Formula I is selected from compounds of Formula Γ”:

X------(Y)n

0L°\'^rZ’ ,_r1; v /Tr⁷²

Im N-N I’” and deuterated dérivatives and pharmaceutically acceptable salts thereof, wherein:

X is selected from -O-, -S-, -SO-, and -SO2-;

Ç B ) each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, Cô-Cio aryl, 5- to 10-membered heteroaryl, -OR^Y1, -CO₂R^Y1, -COR^YI, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Yon the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Ci-C₆ alkyl, and Ci-Ce, alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

5- to 10-membered heteroaryl optionally substituted with 1-3 groups independently selected from: o halogen, o Ci-Cô alkyl (optionally substituted with 1 -3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3-to 10-membered heterocyclyl,

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -OR², -N(R²)2, CO2R², -CO-N(R²)2, -CN, phenyl, benzyl, Ci-Cô alkoxy, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

n is selected from 4, 5, 6, 7, and 8; and m is selected from 0, 1,2, and 3.

In some embodiments, X in Formula I’” is -O-.

In some embodiments, each R^Y in Formula I’” is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C₃-Cs cycloalkyl, and -OR^YI, wherein Q and R^Y1 are as defrned for Formula I’”. In some embodiments, -OR^Y1 in Formula I’” is -OH.

In some embodiments, each Q in Formula I’” is independently selected from C3-C8 cycloalkyl and Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Cô alkyl. In some embodiments, each Q in Formula I’” is independently selected from:

In some embodiments, each R^Y in Formula Γ” is independently selected from:

CH₃ OH hydrogen, fluorine, —1— , -4—

In some embodiments, Ring B in Formula I’” is selected from Cj-Cs cycloalkyl and phenyl optionally substituted with 1-3 groups independently selected from halogen. In some embodiments, Ring B in Formula I’” is selected from:

F

In some embodiments, n in Formula I’” is selected from 4, 5, and 6.

In some embodiments, -(Y)n- in Formula I’” is a group selected from:

In some embodiments, each R¹ in Formula I’” is independently selected from Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -N(R²)2, and -CO2R², wherein R² is as defined for Formula I’”. In some embodiments, each R¹ in Formula I’” is independently selected from -CF₃, -NH2, -NH(CH₂CH3), CO₂H, and CH2OH.

In some embodiments, each R² in Formula I’” is independently selected from hydrogen and Ci-Cô alkyl.

In some embodiments, R^Z1 in Formula I’” is selected from hydrogen and Ci-Cf, alkyl (optionally substituted with 1-6 groups selected from halogen). In some embodiments, R^Z1 in Formula I’” is -CF3.

In some embodiments, R²² in Formula I’” is hydroxy.

In some embodiments, R^Z1 in Formula I’” is Ci-Cô alkyl (optionally substituted with 1-6 groups selected from halogen) and R^Z2 in Formula I’” is hydroxy. In some embodiments, R^Z1in Formula I’” is -CF3 and R^Z2 in Formula I’” is hydroxy.

In some embodiments, m in Formula I’” is selected from 1 and 2.

In some embodiments, the compound of Formula I is selected from compounds of Formula lia’:

( B ) each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Ce alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^YI-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Ci-Ci, alkyl, and Ci-Cô alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

o halogen, .

halogen,

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)₂, -CN, phenyl, benzyl, Ci-Cô alkoxy, Ci-Cô alkyl, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2', each R² is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

R^zl is selected from hydrogen, -CN, Ci-C₆ alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^z2 is selected from hydrogen, halogen, and hydroxy, or R^Z1 and R^z2 taken together form a group selected from oxo and =N-OH; and m is selected from 0, 1,2, and 3.

In some embodiments, m in Formula Ha’ is selected from 1 and 2. In some embodiments, m in Formula Ha’ is 2.

In some embodiments, X in Formula lia’ is -O-.

In some embodiments, each Y in Formula lia’ is independently selected from -C(R^Y)2-, ί B 1

CO-, and ' ' , wherein R^Y and Ring B are as defined for Formula lia’.

In some embodiments, each Y in Formula Ha’ is -C(R^Y)2-, wherein R^Y is as defined for

Formula lia’.

In some embodiments, each R^Y in Formula Ha’ is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1, wherein Q and R^YI are as defined for Formula Ha’.

In some embodiments, each R^Y in Formula Ha’ is independently selected from:

CH₃ OH hydrogen, —L- , ~J— ,

and

wherein Q is as defined for Formula Ha’.

In some embodiments, each Q in Formula lia’ is independently selected from:

C3-C8 cycloalkyl,

In some embodiments, each Q in Formula Ha’ is independently selected from:

In some embodiments, Ring B in Formula Ha’ is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

In some embodiments, Ring B in Formula Ha’ is selected from:

F

In some embodiments, each R¹ in Formula Ha’ is independently Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen) and -N(R²)2 wherein R² is as defined for Formula Ha’. In some embodiments, each R¹ in Formula Ha’ is independently selected from -CF3 and -N(R²)2 wherein R² is as defined for Formula Ha’.

In some embodiments, each R² in Formula lia’ is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-3 groups independently selected from halogen). In some embodiments, each R² in Formula Ha’ is independently selected from hydrogen and Ci-Cô alkyl. In some embodiments, each R in Formula Ha’ is hydrogen.

In some embodiments, R^Z1 in Formula Ha’ is selected from Ci-Cô alkyl (optionally substituted with 1-6 groups selected from halogen). In some embodiments, R^Z1 in Formula Ha’ is -CF3.

In some embodiments, R²² in Formula Ha’ is hydroxy.

In some embodiments, R^Z1 in Formula Ha’ is Ci-Cô alkyl (optionally substituted with 1-6 groups selected from halogen) and R^Z2 in Formula Ha’ is hydroxy. In some embodiments, R^Z1in Formula Ha’ is -CF3 and R^Z2 in Formula Ha’ is hydroxy.

In some embodiments, the compound of Formula I is selected from compounds of Formula Ilia’:

L ^B Z each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^YI is independently selected from hydrogen and Ci-Cô alkyl, or two instances of R^Y1 bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

halogen,

5- to 10-membered heteroaryl optionally substituted with 1-3 groups independently selected from: o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and ο 3- to 10-membered heterocyclyl,

R²² is selected from hydrogen, halogen, and hydroxy, or R^Z1 and R²² taken together form a group selected from oxo and =N-0H; and m is selected from 0, 1,2, and 3.

In some embodiments, X in Formula Ilia’ is -O-.

In some embodiments, each Y in Formula Ilia’ is independently selected from -C(R^Y)2-, ( B J

-CO-, and \ ' , wherein R^Y and Ring B as defined for Formula Ilia’.

In some embodiments, each Y in Formula Ilia’ is -C(R^Y)2-, wherein R^Y is as defined for Formula Ilia’.

In some embodiments, each R^Y in Formula Ilia’ is independently selected from hydrogen, Ci-C₆ alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Yl, wherein Q and R^Y1 are as defined for Formula Ilia’.

In some embodiments, each R^Y in Formula Ilia’ is independently selected from:

ch₃ oh hydrogen, —L~ , —1— ,

wherein Q is as defined for Formula Ilia’.

In some embodiments, each Q in Formula Ilia’ is independently selected from:

C3-C8 cycloalkyl,

In some embodiments, each Q in Formula Ilia’ is independently selected from:

In some embodiments, Ring B in Formula Ilia’ is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

In some embodiments, Ring B in Formula Ilia’ is selected from:

F

In some embodiments, each R¹ in Formula Ilia’ is independently Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen) and -N(R²)2, wherein R² is as defined for Formula Ilia’. In some embodiments, each R¹ in Formula Ilia’ is independently selected from -CF3 and -N(R²)2, wherein R² is as defined for Formula Ilia’.

In some embodiments, each R² in Formula Ilia’ is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-C₆ alkoxy, which is optionally substituted with 1-3 groups independently selected from halogen). In some embodiments, each R² in Formula Ilia’ is independently selected from hydrogen and Ci-Cô alkyl.

In some embodiments, each R² in Formula Ilia’ is hydrogen.

In some embodiments, R^Z1 in Formula Ilia’ is selected from Ci-Cô alkyl (optionally substituted with 1-6 groups selected from halogen). In some embodiments, R^ZI in Formula Ilia’ is -CF3.

In some embodiments, R^z2 in Formula Ilia’ is hydroxy.

In some embodiments, R^Z1 in Formula Ilia’ is Ci-Cô alkyl (optionally substituted with Ιό groups selected from halogen) and R^Z2 in Formula Ilia’ is hydroxy. In some embodiments, R^Z1 in Formula Ilia’ is -CF3 and R^Z2 in Formula Ilia’ is hydroxy.

Compounds of the invention include Compounds 1-53 and 54-77, and deuterated dérivatives and pharmaceutically acceptable salts thereof.

1	2	3
Ο'^^χΧΧ ^F2C-A_n s \\ // OH N-N	o^^^A FiC^ A. L Il ^N 1 \\ // OH N-N enantiomer 1	O'^^A ^fMn \\ // OH N-N enantiomer 2
4	5	6
O^^A ^n X^A/°\A-CF₃T X // OH nh₂ n-n	O^^X ^F3CY^n Ux/°VA^CF3 T X // OH nh₂ n-n	O^^'A ^F3C-A s CA^/oxA^cf3 I v II OH nh₂ n-n
7	8	9
CF-i^ A. k ki n η •HCl Y, OH nh₂ n-n diastereomer pair 1	CF>. A k >i n > Un u₃ HCl V U OH nh₂ n-n diastereomer pair 2	0 o=s η FqC^ A \ XXx/O.to^' ^CF3 T X 11 oh nh₂ n-n Enantiomer 1
10	11	12
0 o=s η ^γ3^ύ^_νWv^³T X v OH nh₂ N-n Enantiomer 2	o^^A ^CF3-An s ^A/°xAc^CF3 T X // OH nh₂ n-n	o^^X ^CF3Y^n X A^X/°xX^CF₃ T \\ // OH ^HCI nh₂ n-n

13	14	15
υΧΧ, •HCl T '' ” OH nh₂ n-n	‘UïA, T \\ // OH nh₂ n-n	0 II O=S—\ ^r,V^. nh₂ n-n enantiomer 1
16	17	18
0 II O=S—\ ^,,cXXLv-. nh₂ n-n enantiomer 2	o^=L ( UAv°v''^CFj •HCI 1 v ' OH ^HCI nh₂ n-n	nh₂ n-n ^ohdiastereomer pair
19	20	21
^f=^c->n > Vvrt? nh₂ n-n	^ρ=^εΆ S vwb nh₂ N-n	'tikX, T \\ // OH nh₂ n-n
22	23	24
T \\ // OH nh₂ n-n	U'o kU ^oh enantiomer 1	^f3cA7~\ nh₂ n-n ^oh enantiomer 2

37	38	39
HO^ k tSîVî-^cf3 NH₂ N-n OH	- D _oA^d ^CF3Y^n k k^k/°xA-CF₃ T v H OH nh₂ n-n	D^³O CF.^ Λ D D L Il ^N I k^k/°x>rCF₃ T \\ // OH nh₂ n-n
40	41	42
^F3^CA^0H k LA^°xA-cf₃T V // OH nh₂ n-n diastereomer 1	^f3cJIn ^oh \ K^^/O^AtCFs T \\ // OH nh₂ n-n diastereomer 2	θΑ^γΟΗ ^CF3Y^n S k^k/°\UcCF₃ T \\ // OH nh₂ n-n diastereomer 1
43	44	45
_Ξ OH ^CF3Y^n k kk^°xkv^CF3 T \\ // OH nh₂ n-n regioisomeric diastereomer 1	o ^^ ^cf3-^à_n d-K U/J^CFj T \\ // oh nh₂ n-n	οΛ^γΟΗ ^cfYn kk\x°x^/VCF3 T \\ // OH nh₂ n-n diastereomer 2
46	47	48
_Ξ OH ^CF3Y^n k T v # OH nh₂ n-n diastereomer 2	^y^N k kA^O L^CF₃ NH₂ N-n ^oh enantiomer 1	O'^^/X} ^F3CY^n k k^k^Oq J^CF₃t » ³ nh₂ n-n oh enantiomer 2

62 63

enantiomer 3

diastereomer 1

Methods of Treatment

Any of the novel compounds disclosed herein, such as for example, compounds of Formulae I, I’, I”, F”, la, Ha, lia’, Hb, Ile, Ild, Ile, Hf, HIa, HIa’, HIb, IIIc, IHd, HIe, and Hlf, Compounds 1 to 53, Compounds 54 to ΊΊ, pharmaceutically acceptable salts thereof, and deuterated dérivatives of such compounds and salts can act as a CFTR modulator, i.e., it modulâtes CFTR activity in the body. Individuals suffering from a mutation in the gene encoding CFTR may benefit from receiving a CFTR modulator. A CFTR mutation may affect the CFTR quantity, i.e., the number of CFTR channels at the cell surface, or it may impact CFTR fiinction, i.e., the functional ability of each channel to open and transport ions. Mutations affecting CFTR quantity include mutations that cause defective synthesis (Class I defect), mutations that cause defective processing and trafficking (Class II defect), mutations that cause reduced synthesis of CFTR (Class V defect), and mutations that reduce the surface stability of CFTR (Class VI defect). Mutations that affect CFTR fiinction include mutations that cause defective gating (Class III defect) and mutations that cause defective conductance (Class IV defect). Some CFTR mutations exhibit characteristics of multiple classes. Certain mutations in the CFTR gene resuit in cystic fibrosis.

Thus, in some embodiments, the invention provides methods of treating, lessening the severity of, or symptomatically treating cystic fibrosis in a patient comprising administering to 71 the patient an effective amount of any of the novel compounds disclosed herein, such as for example, compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Hd, Ile, Hf, Ilia, IHa’, HIb, Hic, HId, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, pharmaceutically acceptable salts thereof, and/or deuterated dérivatives of such compounds and salts, alone or in 5 combination with another active ingrédient, such as another CFTR modulating agent. In some embodiments, the patient has an F508del/minimal fonction (MF) génotype, F508del/F508del génotype (homozygous for the F508del mutation), F508del/gating génotype, or F508del/residual fonction (RF) génotype. In some embodiments the patient is heterozygous and has one F508del mutation. In some embodiments the patient is homozygous for the NI303K mutation.

In some embodiments, 1 mg to 1000 mg of a compound disclosed herein, a pharmaceutically acceptable sait thereof, or a deuterated dérivative of such compound or sait are administered daily.

In some embodiments, the patient is heterozygous and has an F508del mutation on one allele and a mutation on the other allele selected from Table 2:

Table 2: CFTR Mutations

Mutation

Q2X	L218X	Q525X	R792X	E1104X
S4X	Q220X	G542X	E822X	W1145X
W19X	Y275X	G550X	W882X	R1158X
G27X	C276X	Q552X	W846X	R1162X
Q39X	Q290X	R553X	Y849X	S1196X
W57X	G330X	E585X	R851X	W1204X
E60X	W401X	G673X	Q890X	L1254X
R75X	Q414X	Q685X	S912X	S1255X
L88X	S434X	R709X	Y913X	W1282X
E92X	S466X	K710X	Q1042X	Q1313X
Q98X	S489X	Q715X	W1089X	Q1330X
Y122X	Q493X	L732X	Y1092X	E1371X
E193X	W496X	R764X	W1098X	Q1382X
W216X	C524X	R785X	R1102X	Q1411X
185+1 G->T	711+5G—>A	1717-8G—»A	2622+1 G—» A	3121-1G—»A
296+1 G—A	712-1G—>T	1717-1G—A	2790-1 G->C	3500-2A—»G
296+1 G—>T	1248+1 G—» A	1811+1G—»C	3040G—>C	3600+2insT
405+1G—>A	1249-1 G—A	1811+1.6kbA—»G	(G970R)	3850-1 G—> A
405+3A—C	1341+1G—>A	1811+1643G—>T	3120G—>A	4005+1 G-> A
406-1 G—» A	1525-2A—>G	1812-1G—>A	3120+1G—>A	4374+1 G—>T
621+1 G—»T	1525-1 G—> A	1898+1G—>A	3121-2 A—> G
711+1G—♦ T		1898+1 G-»C
182delT	1078delT	1677delTA	2711delT	3737delA
306insA	1119delA	1782delA	2732insA	3791delC
306delTAGA	1138insG	1824delA	2869insG	3821delT

Mutation
365-366insT	1154insTC	1833delT	2896insAG	3876delA
394delTT	llôldelC	2043delG	2942insT	3878delG
442delA	1213delT	2143delT	2957delT	3905insT
444delA	1259insA	2183 AA—»G	3007delG	4016insT
457TAT—>G	1288insTA	2184delA	3028delA	4021dupT
541delC	1343delG	2184insA	3171delC	4022insT
574delA	1471delA	2307insA	3171insC	4040delA
663delT	1497delGG	2347delG	3271delGG	4279insA
849delG	1548delG	2585delT	3349insT	4326delTC
935delA	1609delCA	2594delGT	3659delC
CFTRdelel		CFTRdelel 6-17b	1461ins4
CFTRdele2		CFTRdelel 7a, 17b	1924del7
CFTRdele2,3		CFTRdelel 7a-l 8	2055del9—»	A
CFTRdele2-4		CFTRdelel 9	2105-2117del 13insAGAAA
CFTRdele3-10,14b-16	CFTRdelel9-21	2372del8
CFTRdele4-7		CFTRdele21	2721delll
CFTRdele4-ll		CFTRdele22-24	2991del32
CFTR50kbdel		CFTRdele22,23	3667ins4
CFTRdup6b-10	124del23bp	4010del4
CFTRdelel 1		602dell4	4209TGTT-	->AA
CFTRdelel3,14a	852del22
CFTRdelel4b-17b	991del5
A46D	V520F	Y569D	N1303K
G85E	A559T	L1065P
R347P	R560T	R1066C
L467P	R560S	L1077P
I507del	A561E	M1101K

In some embodiments, the disclosure also is directed to methods of treatment using isotope-labelled compounds of the afore-mentioned compounds, or pharmaceutically acceptable salts thereof, wherein the formula and variables of such compounds and salts are each and independently as described above or any other embodiments described above, provided that one 5 or more atoms therein hâve been replaced by an atom or atoms having an atomic mass or mass number which differs from the atomic mass or mass number of the atom which usually occurs naturally (isotope labelled). Examples of isotopes which are commercially available and suitable for the disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphores, fluorine and chlorine, for example ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, ^l70,³¹P, ³²P, ³⁵S, ¹⁸F and ³⁶C1, respectively.

The isotope-labelled compounds and salts can be used in a number of bénéficiai ways.

They can be suitable for médicaments and/or varions types of assays, such as substrate tissue distribution assays. For example, tritium (³H)- and/or carbon-14 (¹⁴C)-labelled compounds are particularly useful for varions types of assays, such as substrate tissue distribution assays, due to relatively simple préparation and excellent detectability. For example, deuterium (²H)-labelled ones are therapeutically useful with potential therapeutic advantages over the non-²H-labelled compounds. In general, deuterium (²H)-labelled compounds and salts can hâve higher metabolic stability as compared to those that are not isotope-labelled owing to the kinetic isotope effect described below. Higher metabolic stability translates directly into an increased in vivo half-life or lower dosages, which could be desired. The isotope-labelled compounds and salts can usually be prepared by carrying out the procedures disclosed in the synthesis schemes and the related description, in the example part and in the préparation part in the présent text, replacing a nonisotope-labelled reactant by a readily available isotope-labelled reactant.

In some embodiments, the isotope-labelled compounds and salts are deuterium (²H)labelled ones. In some spécifie embodiments, the isotope-labelled compounds and salts are deuterium (²H)-labelled, wherein one or more hydrogen atoms therein hâve been replaced by deuterium. In Chemical structures, deuterium is represented as “²H” or “D.”

When discovering and developing therapeutic agents, the person skilled in the art attempts to optimize pharmacokinetic parameters while retaining désirable in vitro properties. It may be reasonable to assume that many compounds with poor pharmacokinetic profiles are susceptible to oxidative metabolism.

The deuterium (²H)-labelled compounds and salts can modulate the oxidative metabolism of the compound by way of the primary kinetic isotope effect. The primary kinetic isotope effect is a change of the rate for a Chemical reaction that results from exchange of isotopic nuclei, which in tum is caused by the change in ground state energies necessary for covalent bond formation after this isotopic exchange. Exchange of a heavier isotope usually results in a lowering of the ground state energy for a Chemical bond and thus causes a réduction in the ratelimiting bond breakage. If the bond breakage occurs in or in the vicinity of a saddle-point région along the coordinate of a multi-product reaction, the product distribution ratios can be altered substantially. For explanation: if deuterium is bonded to a carbon atom at a non-exchangeable position, rate différences of kM/ko= 2-7 are typical. For a further discussion, see S. L. Harbeson and R. D. Tung, Deuterium In Drug Discovery and Development, Ann. Rep. Med. Chem. 2011, 46,403-417, which is incorporated herein by reference.

The concentration of the isotope(s) (e.g., deuterium) incorporated into the isotopelabelled compounds and sait of the disclosure may be defined by the isotopic enrichment factor. The term “isotopic enrichment factor” as used herein means the ratio between the isotopic abundance and the naturel abundance of a specified isotope. In some embodiments, if a substituent in a compound of the disclosure is denoted deuterium, such compound has an 74 isotopic enrichment factor for each designated deuterium atom of at least 3500 (52.5% deuterium incorporation at each designated deuterium atom), at least 4000 (60% deuterium incorporation), at least 4500 (67.5% deuterium incorporation), at least 5000 (75% deuterium incorporation), at least 5500 (82.5% deuterium incorporation), at least 6000 (90% deuterium incorporation), at least 6333.3 (95% deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5% deuterium incorporation).

Combination Thérapies

One aspect disclosed herein provides methods of treating cystic fibrosis and other CFTRmediated diseases using any of the novel compounds disclosed herein, such as for example, compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Hd, Ile, Hf, Ilia, IIIa’,IIIb, IIIc, Hld, Hle, and IHf, Compounds 1 to 53, Compounds 54 to 77, pharmaceutically acceptable salts thereof, and deuterated dérivatives of such compounds and salts in combination with at least one additional active pharmaceutical ingrédient.

Thus, in some embodiments, the invention provides methods of treating, lessening the severity of, or symptomatically treating cystic fibrosis in a patient comprising administering to the patient an effective amount of any of the novel compounds disclosed herein, such as for example, compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Hd, Ile, Hf, HIa, HIa’, HIb, IIIc, Hld, Hle, and IHf, Compounds 1 to 53, Compounds 54 to 77, pharmaceutically acceptable salts thereof, and/or deuterated dérivatives of such compounds and salts, alone or in combination with at least one additional active pharmaceutical ingrédient, such as, e.g., a CFTR modulating agent.

In some embodiments, at least one additional active pharmaceutical ingrédient is selected from mucolytic agents, bronchodilators, antibiotics, anti-infective agents, and antiinflammatory agents.

In some embodiments, the additional therapeutic agent is an antibiotic. Exemplary antibiotics usefiil herein include tobramycin, including tobramycin inhaled powder (TIP), azithromycin, aztreonam, including the aerosolized form of aztreonam, amikacin, including liposomal formulations thereof, ciprofloxacin, including formulations thereof suitable for administration by inhalation, levoflaxacin, including aerosolized formulations thereof, and combinations of two antibiotics, e.g., fosfomycin and tobramycin.

In some embodiments, the additional agent is a mucolyte. Exemplary mucolytes useful herein includes Pulmozyme®.

In some embodiments, the additional agent is a bronchodilator. Exemplary bronchodilators include albuterol, metaprotenerol sulfate, pirbuterol acetate, salmeterol, or tetrabuline sulfate.

In some embodiments, the additional agent is an anti-inflammatory agent, i.e., an agent that can reduce the inflammation in the lungs. Exemplary such agents useful herein include ibuprofen, docosahexanoic acid (DHA), sildenafil, inhaled glutathione, pioglitazone, hydroxychloroquine, or simavastatin.

In some embodiments, the additional agent is a nutritional agent. Exemplary nutritional agents include pancrelipase (pancreating enzyme replacement), including Pancrease®, Pancreacarb®, Ultrase®, or Creon®, Liprotomase® (formerly Trizytek®), Aquadeks®, or glutathione inhalation. In one embodiment, the additional nutritional agent is pancrelipase.

In some embodiments, at least one additional active pharmaceutical ingrédient is selected from CFTR modulating agents. In some embodiments, the CFTR modulating agent is a CFTR corrector. In some embodiments, the CFTR modulating agent is a CFTR potentiator enhancer/co-potentiator (for example, ASP-11). In some embodiments, the at least one i additional active pharmaceutical ingrédient is a CFTR amplifier. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR readthrough agent. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR nucleic acid therapy.

In some embodiments, the at least one additional active pharmaceutical ingrédient is a ENaC inhibitor. In some embodiments, the at least one additional active pharmaceutical ingrédient is a TMEM16A modulator. In some embodiments, the at least one additional active pharmaceutical ingrédient is a GPR39 agonist.

In some embodiments, the at least one additional active pharmaceutical ingrédient is chosen from (a) Compound II and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) Compound IV and pharmaceutically acceptable salts and deuterated dérivatives thereof; (c) Compound V and pharmaceutically acceptable salts and deuterated dérivatives thereof; (d) Compound VI and pharmaceutically acceptable salts and deuterated dérivatives thereof; (e) Compound VII and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (f) Compound VIII and pharmaceutically acceptable salts and deuterated dérivatives thereof. Thus, in some embodiments, the combination thérapies provided herein comprise a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Hlb, IIIc, Illd, Hle, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and at least one compound chosen from Compound II, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, the combination thérapies provided herein comprise (a) at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, lia, lia’, Hb, Ile, Hd, Ile, Ilf, Ilia, IIIa’,IIIb, IIIc, IHd, IHe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound II, Compound IV, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one compound chosen from Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, the combination thérapies provided herein comprise (a) at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, lia, Ha’, Ilb, Ile, Ild, Ile, Ilf, Ilia, IIIa’,IIIb, IIIc, IHd, IHe, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound selected from Compound II and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one compound chosen from Compound VII and pharmaceutically acceptable salts and deuterated dérivatives thereof.

In some embodiments, the combination thérapies provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Ilb, Ile, Hd, Ile, Ilf, HIa, HIa’JHb, IIIc, IHd, IHe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one compound chosen from compounds disclosed in WO 2016/105485, United States Patent Application Publication No. 2016-0120841, United States Patent Application Publication No. 2017-0101405, WO 2017/009804, WO 2018/065921, WO 2017/062581, or Journal of Cystic Fibrosis (2018), 17(5), 595-606.

In some embodiments, the combination thérapies provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Hd, Ile, Ilf, HIa, HIa’JHb, IIIc, IHd, IHe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one compound chosen from PTI-428, ASP-11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and PTI-801.

In some embodiments, the combination thérapies provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Hf, Ilia, HIa’JIIb, Hic, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (b) at least two compounds chosen from compounds disclosed in WO 2019/195739, WO 2019/200246, WO 2021/030555, WO 2021/030556, WO 2017/173274, WO 2019/010092, WO 2019/018353, WO 2010/053471, WO 2011/119984, WO 2011/133751, WO 2011/133951, WO 2015/160787, WO 2007/056341, WO 2009/073757, WO 2009/076142, WO 2018/107100, WO 2019/113476, WO 2018/064632, WO 2019/152940, WO 2016/057572, WO 2021/030554, WO 2020/206080, WO 2016/105485, United States Patent Application Publication No. 2016-0120841, United States Patent Application Publication No. 2017-0101405, WO 2017/009804, WO 2018/065921, WO 2017/062581, Journal of Cystic Fîbrosis (2018), 17(5), 595-606, Pedemonte, N. et al. Sci. Adv. 2020, 6 (8), eaay9669, Phuan, P.-W. et al. Sci. Rep. 2019, 9 (1), 17640, Bose, S. et al. J. Cyst. Fibros. 2020,19 Suppl 1, S25-S32, Crawford, D.K. J. Pharmacol. Exp. Ther. 2020, 374 (2), 264-272, Brasell, E.J. et al. PLoS One 2019,14 (12), e0223954, Smith, N.J, Solovay, C.F., Pharm. Pat. Anal. 2017, 6 (4), 179-188, Kunzelmann, K. et al., Front. Pharmacol. 2019,10, 3, or Son, J.-H. et al., Eur. J. ofMed. Chem. 2020, 112888.

In some embodiments, the combination thérapies provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, lia, lia’, Ilb, Hc, Ild, Ile, Ilf, Ilia, HIa’JIIb, IIIc, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (b) at least two compounds chosen from Compound II, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, PTI-428, ASP-11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and PTI-801, and pharmaceutically acceptable salts and deuterated dérivatives thereof.

In some embodiments, the combination thérapies provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Hc, Ild, Ile, Hf, Ilia, IHa’, IHb, IIIc, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound II, Compound III, Compound ΗΙ-d, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, PTI-428, ASP

11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV119, FDL-169, ARN5562, ARN21586, ARN22081, ARN22652, ARN23765, ARN23766, PTI801, FDL-176, PTI-808 (dirocaftor), GLPG1837, GLPG2451/ABBV-2451, QBW251 (icenticaftor), GLPG3067/ABBV-3067 (Navocaftor), ABBV-191, ELX-02, MRT5005, LunarCF, RCT223, amiloride, ETD001, CF552, GS-9411, GS-5737, P-1037 (VX-371), P-1055 (VX551), AZD5634, SPX-101, Ionis-ENaC-2.5 Rx, BI 1265162, AZ5634, ARO-ENaClOOl, ETD002, and DS-1039, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one pharmaceutically acceptable carrier.

In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Ild, Ile, Hf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in combination with at least one compound chosen from Compound II and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Hc, Ild, Ile, Hf, IHa, HIa’, Illb, IIIc, IHd, Ille, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in combination with at least one compound chosen from Compound IV and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Ild, Ile, Hf, HIa, HIa’, Illb, IIIc, IHd, HIe, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in combination with at least one compound chosen from Compound V and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Ild, Ile, Hf, Ilia, HIa’, HIb, IIIc, HId, Ille, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in combination with at least one compound chosen from Compound VI and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Hc, Ild, Ile, Hf, IHa, HIa’, Illb, IIIc, HId, Ille, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in combination with at least one compound chosen from Compound VII and pharmaceutically acceptable salts and deuterated dérivatives thereof. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Ild, Ile, Hf,

Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in combination with at least one compound chosen from Compound VIII and pharmaceutically acceptable salts and deuterated dérivatives thereof.

Each of the compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, Compound II, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and their pharmaceutically acceptable salts and deuterated dérivatives thereof, independently can be administered once daily, twice daily, or three times daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, Γ”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof are administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound IV and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound IV and pharmaceutically acceptable salts thereof are administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, F”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound V and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound V and pharmaceutically acceptable salts thereof are administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound VI and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound VI and pharmaceutically acceptable salts thereof are administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, lia, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I”’, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof are administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, F”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound VIII and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least one compound chosen from Compound VIII and pharmaceutically acceptable salts thereof are administered twice daily.

In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Hd, Ile, Hf, Ilia, Ilia’, IHb, Hic, IHd, IHe, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; at least one compound chosen from Compound II, Compound IV, and pharmaceutically acceptable salts thereof; and at least one compound chosen from Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Hd, Ile, Ilf, Ilia, IHa’, IHb, Hic, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; at least one compound chosen from Compound H, Compound IV, and pharmaceutically acceptable salts thereof; and at least one compound chosen from Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and pharmaceutically acceptable salts thereof are administered twice daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Hc, Hd, Ile, Ilf, Ilia, HIa’, IHb, Hic, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof; and at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof are administered once daily. In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Hc, Hd, Ile, Hf, HIa, HIa’, IHb, IHc, HId, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof; and at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof are administered twice daily.

Compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Hc, Hd, Ile, Ilf, IHa, HIa’, IHb, IHc, HId, IHe, and Hlf, Compounds 1 to 53, Compounds 54 to 77, Compound II, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and their pharmaceutically acceptable salts and deuterated dérivatives thereof can be administered in a single pharmaceutical composition or separate pharmaceutical compositions. Such pharmaceutical compositions can be administered once daily or multiple times daily, such as twice daily. As used herein, the phrase that a given amount of API (e.g.,

Compound II, Compound VII, or pharmaceutically acceptable salts thereof) is administered once or twice daily or per day means that said given amount is administered per dosing, which may occur once or twice daily.

In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Hd, Ile, Hf, IHa, HIa’, IHb, IIIc, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in a first pharmaceutical composition; and at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof is administered in a second pharmaceutical composition.

In some embodiments, at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Ild, Ile, Ilf, IHa, HIa’, HIb, IIIc, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof is administered in a first pharmaceutical composition; at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof is administered in a second pharmaceutical composition; and at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof is administered in a third pharmaceutical composition.

Any suitable pharmaceutical compositions known in the art can be used for compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Hd, Ile, Ilf, HIa, Ilia’, IHb, IIIc, Hld, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof. Some exemplary pharmaceutical compositions for Compound II and its pharmaceutically acceptable salts can be found in WO 2011/119984 and WO 2014/014841, incorporated herein by reference. Some exemplary pharmaceutical compositions for Compound III and its pharmaceutically acceptable salts can be found in WO 2007/134279, WO 2010/019239, WO 2011/019413, WO 2012/027731, and WO 2013/130669, and some exemplary pharmaceutical compositions for Compound IH-d and its pharmaceutically acceptable salts can be found in US 8,865,902, US 9,181,192, US 9,512,079, WO 2017/053455, and WO 2018/080591, ail of which are incorporated herein by reference. Some exemplary pharmaceutical compositions for Compound IV and its pharmaceutically acceptable salts can be found in WO 2010/037066, WO 2011/127421, and WO 2014/071122, incorporated herein by reference. Some exemplary pharmaceutical compositions for Compound V and its pharmaceutically acceptable salts can be found in WO 2019/152940, incorporated herein by reference. Some exemplary pharmaceutical compositions for Compound VI and its pharmaceutically acceptable salts can be found in WO 2019/079760, incorporated herein by reference.

Pharmaceutical Compositions

Another aspect of the invention provides a pharmaceutical composition comprising at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, lia, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to ΊΊ, and pharmaceutically acceptable salts and deuterated dérivatives thereof, and at least one pharmaceutically acceptable carrier.

In some embodiments, the invention provides pharmaceutical compositions comprising at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, lia, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to ΊΊ, and pharmaceutically acceptable salts and deuterated dérivatives thereof in combination with at least one additional active pharmaceutical ingrédient. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR modulator. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR corrector. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR potentiator. In some embodiments, the at least one additional active pharmaceutical ingrédient is a compound that enhances CFTR potentiation, i.e., a CFTR potentiator enhancer/co-potentiator. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR amplifier. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR readthrough agent. In some embodiments, the at least one additional active pharmaceutical ingrédient is a CFTR nucleic acid therapy. In some embodiments, the at least one additional active pharmaceutical ingrédient is a ENaC inhibitor. In some embodiments, the at least one additional active pharmaceutical ingrédient is a TMEM16A modulator. In some embodiments, the at least one additional active pharmaceutical ingrédient is a GPR39 agonist. In some embodiments, the pharmaceutical composition comprises at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least two additional active pharmaceutical ingrédients, each of which is a CFTR corrector. In some embodiments, the pharmaceutical composition comprises at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Ilb, Ile, Ild, Ile, Ilf, Ilia, Ilia’, Illb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof and at least two additional active pharmaceutical ingrédients, one of which is a CFTR corrector and one of which is a CFTR potentiator enhancer.

In some embodiments, the invention provides a pharmaceutical composition comprising (a) at least one compound chosen from compounds of Formulae I, I’, I”, Γ”, la, Ha, lia’, Hb, Ile, Hd, Ile, Ilf, HIa, IHa’, IHb, IIIc, IHd, HIe, and Hlf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof, (b) at least one compound chosen from Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and pharmaceutically acceptable salts thereof, and (c) at least one pharmaceutically acceptable carrier.

In some embodiments, the invention provides a pharmaceutical composition comprising (a) at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Ild, Ile, Ilf, HIa, Ilia’, IHb, IIIc, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof, (b) at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof, and (c) at least one pharmaceutically acceptable carrier.

In some embodiments, the invention provides a pharmaceutical composition comprising (a) at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Ild, Ile, Ilf, HIa, Ilia’, HIb, IIIc, IHd, HIe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof, (b) at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof, and (c) at least one pharmaceutically acceptable carrier.

In some embodiments, the disclosure provides a pharmaceutical composition comprising (a) at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Hd, Ile, Ilf, HIa, HIa’, HIb, IIIc, IHd, IHe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof, (b) at least one compound chosen from Compound H, Compound IV, and pharmaceutically acceptable salts thereof, (c) at least one compound chosen from Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and pharmaceutically acceptable salts thereof, and (d) at least one pharmaceutically acceptable carrier.

In some embodiments, the disclosure provides a pharmaceutical composition comprising (a) at least one compound chosen from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Ild, Ile, Ilf, Ilia, HIa’, IHb, IIIc, IHd, IHe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof, (b) at least one compound chosen from Compound II and pharmaceutically acceptable salts thereof, (c) at least one compound chosen from Compound VII and pharmaceutically acceptable salts thereof, and (d) at least one pharmaceutically acceptable carrier.

In some embodiments, the pharmaceutical compositions provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Ilb, Hc, Hd, Ile, Ilf, IHa, HIa’, HIb, IIIc, Illd, Ille, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (c) at least one compound chosen from compounds disclosed in WO 2016/105485, United States Patent Application Publication No. 2016-0120841, United States Patent Application Publication No. 2017-0101405, WO 2017/009804, WO 2018/065921, WO 2017/062581, Journal of Cystic Fibrosis (2018), 17(5), 595-606, Pedemonte, N. et al. Sci. Adv. 2020, 6 (8), eaay9669, Phuan, P.-W. et al. Sci. Rep. 2019, 9 (1), 17640, Bose, S. et al. J. Cyst. Fibros. 2020,19 Suppl 1, S25-S32, Crawford, D.K. J. Pharmacol. Exp. Ther. 2020, 374 (2), 264-272, Brasell, E.J. et al. PLoS One 2019,14 (12), e0223954, Smith, N.J, Solovay, C.F., Pharm. Pat. Anal. 2017, 6 (4), 179-188, Kunzelmann, K. et al., Front. Pharmacol. 2019,10, 3, or Son, J.-H. et al., Eur. J. ofMed. Chem. 2020,112888; and (d) at least one pharmaceutically acceptable carrier.

In some embodiments, the pharmaceutical compositions provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, lia’, Hb, Ile, Hd, Ile, Ilf, HIa, IHa’, HIb, IIIc, Illd, HIe, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (c) at least one compound chosen from PTI-428, ASP-11, ABBV2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and ΡΊΊ801; and (d) at least one pharmaceutically acceptable carrier.

In some embodiments, the pharmaceutical compositions provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Hd, Ile, Ilf, IHa, HIa’, HIb, IIIc, Illd, HIe, and Illf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least two compounds chosen from compounds disclosed in WO 2019/195739, WO 2019/200246, WO 2021/030555, WO 2021/030556, WO 2017/173274, WO 2019/010092, WO 2019/018353, WO 2010/053471, 86

WO 2011/119984, WO 2011/133751, WO 2011/133951, WO 2015/160787, WO 2007/056341, WO 2009/073757, WO 2009/076142, WO 2018/107100, WO 2019/113476, WO 2018/064632, WO 2019/152940, WO 2016/057572, WO 2021/030554, WO 2020/206080, WO 2016/105485, United States Patent Application Publication No. 2016-0120841, United States Patent Application Publication No. 2017-0101405, WO 2017/009804, WO 2018/065921, WO 2017/062581, Journal of Cystic Fibrosis (2018), 17(5), 595-606, Pedemonte, N. et al. Sci. Adv. 2020, 6 (8), eaay9669, Phuan, P.-W. et al. Sci. Rep. 2019, 9 (1), 17640, Bose, S. et al. J. Cyst. Fibros. 2020,19 Suppl 1, S25-S32, Crawford, D.K. J. Pharmacol. Exp. Ther. 2020, 374 (2), 264-272, Brasell, E.J. et al. PLoS One 2019,14 (12), e0223954, Smith, N.J, Solovay, C.F., Pharm. Pat. Anal. 2017, 6 (4), 179-188, Kunzelmann, K. et al., Front. Pharmacol. 2019,10, 3, or Son, J.-H. et al., Eur. J. ofMed. Chem. 2020, 112888; and (c) at least one pharmaceutically acceptable carrier.

In some embodiments, the pharmaceutical compositions provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Hd, Ile, Hf, HIa, HIa’, Illb, IIIc, Hld, Hle, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least two compounds chosen from Compound H, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, PTI-428, ASP-11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and PTI-801, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one pharmaceutically acceptable carrier.

In some embodiments, the pharmaceutical compositions provided herein comprise (a) a compound selected from compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Hc, Hd, Ile, Hf, HIa, Ilia’, IHb, IIIc, Hld, Hle, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts and deuterated dérivatives thereof; (b) at least one compound chosen from Compound H, Compound III, Compound Hl-d, Compound IV, Compound V, Compound VI, Compound VII, Compound VIH, Compound IX, Compound X, PTI-428, ASP11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV119, FDL-169, ARN5562, ARN21586, ARN22081, ARN22652, ARN23765, ARN23766, PTI801, FDL-176, PTI-808 (dirocaftor), GLPG1837, GLPG2451/ABBV-2451, QBW251 (icenticaftor), GLPG3067/ABBV-3067 (Navocaftor), ABBV-191, ELX-02, MRT5005, LunarCF, RCT223, amiloride, ETD001, CF552, GS-9411, GS-5737, P-1037 (VX-371), P-1055 (VX551), AZD5634, SPX-101, Ionis-ENaC-2.5 Rx, BI 1265162, AZ5634, ARO-ENaClOOl,

ETD002, and DS-1039, and pharmaceutically acceptable salts and deuterated dérivatives thereof; and (c) at least one pharmaceutically acceptable carrier.

Any pharmaceutical composition disclosed herein may comprise at least one pharmaceutically acceptable carrier. In some embodiments, the at least one pharmaceutically acceptable carrier is chosen from pharmaceutically acceptable vehicles and pharmaceutically acceptable adjuvants. In some embodiments, the at least one pharmaceutically acceptable is chosen from pharmaceutically acceptable fillers, disintegrants, surfactants, binders, lubricants.

The pharmaceutical compositions described herein are useful for treating cystic fibrosis and other CFTR-mediated diseases.

As described above, pharmaceutical compositions disclosed herein may optionally further comprise at least one pharmaceutically acceptable carrier. The at least one pharmaceutically acceptable carrier may be chosen from adjuvants and vehicles. The at least one pharmaceutically acceptable carrier, as used herein, includes any and ail solvents, diluents, other liquid vehicles, dispersion aids, suspension aids, surface active agents, isotonie agents, thickening agents, emulsifying agents, preservatives, solid binders, and lubricants, as suited to the particular dosage form desired. Remington; The Science and Practice of Pharmacy, 21st édition, 2005, ed. D.B. Troy, Lippincott Williams & Wilkins, Philadelphia, and Encyclopedia of Pharmaceutical Technology, eds. J. Swarbrick and J. C. Boylan, 1988-1999, Marcel Dekker, New York discloses varions carriers used in formulating pharmaceutical compositions and known techniques for the préparation thereof. Except insofar as any conventional carrier is incompatible with the compounds of this disclosure, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutical composition, its use is contemplated to be within the scope of this disclosure. Non-limiting examples of suitable pharmaceutically acceptable carriers include, but are not limited to, ion exchangers, alumina, aluminum stéarate, lecithin, sérum proteins (such as human sérum albumin), buffer substances (such as phosphates, glycine, sorbic acid, and potassium sorbate), partial glyceride mixtures of saturated vegetable fatty acids, water, salts, and electrolytes (such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, and zinc salts), colloïdal silica, magnésium trisilicate, polyvinyl pyrrolidone, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, wool fat, sugars (such as lactose, glucose and sucrose), starches (such as corn starch and potato starch), cellulose and its dérivatives (such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate), powdered tragacanth, malt, gelatin, talc, excipients (such as cocoa butter and suppository waxes), oils (such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil), glycols (such as propylene glycol and polyethylene glycol), esters (such as ethyl oleate and ethyl laurate), agar, buffering agents (such as magnésium hydroxide and aluminum hydroxide), alginic acid, pyrogen-free water, isotonie saline, Ringer's solution, ethyl alcohol, phosphate buffer solutions, non-toxic compatible lubricants (such as sodium lauryl sulfate and magnésium stéarate), coloring agents, releasing agents, coating agents, sweetening agents, flavoring agents, perfuming agents, preservatives, and antioxidants.

Compound 11 Heptane Solvaté

In some embodiments, the invention provides solvated crystalline forms of Compound 11. In some embodiments, the solvated crystalline form is a heptane solvaté. In some embodiments, the invention provides Compound 11 heptane solvaté. FIG. 1 provides an X-ray powder diffractogram of Compound 11 heptane solvaté at room température.

In some embodiments, Compound 11 heptane solvaté is substantially pure. In some embodiments, Compound 11 heptane solvaté is substantially crystalline. In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram generated by an X-ray powder diffraction analysis with an incident beam of Cu Ka radiation. In some embodiments, Compound 11 heptane solvaté has many molécules in an asymmetric unit. In some embodiments, Compound 11 heptane solvaté is a physical mixture of crystal lattices. In some embodiments, Compound 11 heptane solvaté has a variable amount of heptane in the crystal lattice. In some embodiments, Compound 11 heptane solvaté has a stoichiometric amount of heptane in the crystal lattice. In some embodiments, Compound 11 heptane solvaté has a nonstoichiometric amount of heptane in the crystal lattice.

In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram having a signal at 5.8 ± 0.2 degrees two-theta. In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram having a signal at 10.1 ± 0.2 degrees two-theta. In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram having a signal at 11.7 ± 0.2 degrees two-theta. In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram having one, two, or three signais selected from 5.8 ± 0.2 degrees two-theta, 10.1 ± 0.2 degrees two-theta, and 11.7 ± 0.2 degrees two-theta.

In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram having (a) one, two, or three signais selected from 5.8 ± 0.2 degrees twotheta, 10.1 ± 0.2 degrees two-theta, and 11.7 ± 0.2 degrees two-theta, and (b) one, two, three, or four signais selected from 5.6 ± 0.2 degrees two-theta, 18.1 ± 0.2 degrees two-theta, 20.5 ± 0.2 degrees two-theta, and 20.9 ± 0.2 degrees two-theta. In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram having signais at 5.6 ± 0.2 degrees two-theta, 5.8 ± 0.2 degrees two-theta, 10.1 ± 0.2 degrees two-theta, 11.7 ± 0.2 degrees two-theta, 18.1 ± 0.2 degrees two-theta, 20.5 ± 0.2 degrees two-theta, and 20.9 ± 0.2 degrees two-theta.

In some embodiments, Compound 11 heptane solvaté is characterized by an X-ray powder diffractogram substantially similar to FIG. 1.

In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 166.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 165.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 164.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 163.4 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 154.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 154.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 152.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 151.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 140.2 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 139.4 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 138.5 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 138.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 135.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^I3C SSNMR spectrum with a peak at 134.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 131.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 130.2 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 129.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 128.5 ± 0.2 ppm. Τη some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 125.7 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 123.7 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^I3C SSNMR spectrum with a peak at 123.2 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 122.9 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 121.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 120.2 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 119.2 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 117.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 76.2 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 74.4 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 73.7 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 73.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 40.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 38.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 37.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 36.9 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^I3C SSNMR spectrum with a peak at 35.7 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 33.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 32.5 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 32.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 30.4 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 30.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 29.5 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^I3C SSNMR spectrum with a peak at 28.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 28.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 27.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 25.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 23.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 22.7 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 22.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 21.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 20.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 19.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 18.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 17.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 13.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with a peak at 13.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^l3C SSNMR spectrum with a peak at 12.5 ± 0.2 ppm.

In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹³C SSNMR spectrum with one, two, three, four, fïve, six, seven, eight, nine, ten, or more peaks selected from 166.3 ± 0.2 ppm, 165.8 ± 0.2 ppm, 164.6 ± 0.2 ppm, 163.4 ± 0.2 ppm, 154.8 ± 0.2 ppm, 154.0 ± 0.2 ppm, 152.1 ± 0.2 degppm, 151.6 ± 0.2 ppm, 140.2 ± 0.2 ppm, 139.4 ± 0.2 ppm, 138.5 ± 0.2 ppm, 138.0 ± 0.2 ppm, 135.1 ± 0.2 ppm, 134.6 ± 0.2 ppm, 131.3 ± 0.2 ppm, 130.2 ± 0.2 ppm, 129.6 ± 0.2 ppm, 128.5 ± 0.2 ppm, 125.7 ± 0.2 ppm, 123.7 ± 0.2 ppm, 123.2 ± 0.2 ppm, 122.9 ± 0.2 ppm, 121.1 ± 0.2 ppm, 120.2 ± 0.2 ppm, 119.2 ± 0.2 ppm, 117.8 ± 0.2 ppm, 76.2 ± 0.2 ppm, 74.4 ± 0.2 ppm, 73.7 ± 0.2 ppm, 73.3 ± 0.2 ppm, 40.0 ± 0.2 ppm, 38.6 ± 0.2 ppm, 37.6 ± 0.2 ppm, 36.9 ± 0.2 ppm, 35.7 ± 0.2 ppm, 33.6 ± 0.2 ppm, 32.5 ± 0.2 ppm, 32.0 ± 0.2 ppm, 30.4 ± 0.2 ppm, 30.1 ± 0.2 ppm, 29.5 ± 0.2 ppm, 28.8 ± 0.2 ppm, 28.1 ± 0.2 ppm, 27.1 ± 0.2 ppm, 25.3 ± 0.2 ppm, 23.1 ± 0.2 ppm, 22.7 ± 0.2 ppm, 22.0 ± 0.2 ppm, 21.6 ± 0.2 ppm, 20.3 ±

0.2 ppm, 19.6 ± 0.2 ppm, 18.3 ± 0.2 ppm, 17.6 ± 0.2 ppm, 13.8 ± 0.2 ppm, 13.1 ± 0.2 ppm, and 12.5 ± 0.2 ppm.

In some embodiments, Compound 11 heptane solvaté is characterized by a ^I3C SSNMR spectrum substantially similar to FIG. 3.

In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with a peak at -63.5 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with a peak at -63.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^,9F SSNMR spectrum with a peak at -65.1 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with a peak at -65.8 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^,9F SSNMR spectrum with a peak at -66.3 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with a peak at -67.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ^I9F SSNMR spectrum with a peak at -74.0 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with a peak at -74.9 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with a peak at -76.6 ± 0.2 ppm.

In some embodiments, Compound 11 heptane solvaté is characterized as having a ^I9F SSNMR spectrum with one, two, or three peaks selected from -65.1 ± 0.2 ppm, -67.0 ± 0.2 ppm, and -76.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with one, two, three, four, or five peaks selected from -63.5 ± 0.2 ppm, -65.1 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, and -76.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with one, two, three, four, five, or more peaks selected from -63.5 ± 0.2 ppm, -63.8 ± 0.2 ppm, 65.1 ± 0.2 ppm, -65.8 ± 0.2 ppm, -66.3 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, and -76.6 ± 0.2 ppm. In some embodiments, Compound 11 heptane solvaté is characterized as having a ¹⁹F SSNMR spectrum with one, two, three, four, five, or more peaks selected from -63.5 ± 0.2 ppm, -63.8 ± 0.2 ppm, -65.1 ± 0.2 ppm, -65.8 ± 0.2 ppm, -66.3 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, -76.6 ± 0.2 ppm, and -77.6 ± 0.2 ppm.

In some embodiments, Compound 11 heptane solvaté is characterized by a ^l9F SSNMR spectrum substantially similar to FIG. 4.

Another aspect of the invention provides a process for preparing a solvated crystalline solid form of Compound 11 comprising dissolving Compound 11 in one or more solvents to form a mixture and crystallising the compound from the mixture. In some embodiment the one or more solvents comprises heptane. In some embodiment the one or more solvents comprises heptane and dichloromethane.

Another aspect of the invention provides a method of making Compound 11 heptane solvaté. In some embodiments; the method of making Compound 11 heptane solvaté comprises: (i) dissolving Compound 11 in heptane and dichloromethane to form a mixture; (ii) concentrating the mixture; (iii) collecting solids from the mixture; and (iv) drying the collected solids. In some embodiments, (ii) optionally comprises swirling the mixture at room température. In some embodiments, (iii) optionally comprises rinsing the collected solids with cold heptane. In some embodiments, the method of making Compound 11 heptane solvaté comprises dissolving Compound 11 in heptane and dichloromethane, concentrating under rotary évaporation, swirling at room température, filtering the solids, washing the solids with cold heptane, and drying under vacuum to provide Compound 11 heptane solvaté.

Non-limiting Exemplary Embodiments

1. A compound selected from compounds of Formula I:

( B j each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, Cô-Cio aryl, 5- to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Yon the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -OR², -N(R²)2, CO2R², -CO-N(R²)2, -CN, phenyl, benzyl, Ci-Cô alkoxy, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)₂;

rZ1 R^Z3 |] ( C -)-(R^Z2)o.2

JV -s7^S''R^Z3 V^R^Z1

Z is selected from , ' , and ; wherein

Ring C is selected from Cô-Cio aryl and 5- to 10-membered heteroaryl;

n is selected from 4, 5, 6, 7, and 8; and m is selected from 0, 1,2, and 3.

2. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 1, wherein:

X is selected from -O-, -S-, -SO-, and -SO₂-;

( b 2 each Y is independently selected from -C(R^Y)₂-, -O-, -CO-, and * ' ;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

halogen,

o halogen, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3- to 10-membered heterocyclyl,

each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)i, -CO2R², -CON(R²)2, -CN, phenyl, benzyl, Ci-Cô alkoxy, C3-Cs cycloalkyl, 5- to 6-membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and PO(R²)2;

R²² is selected from hydrogen, halogen, and hydroxy, or R²¹ and R²² taken together form a group selected from oxo and =N-OH;

n is selected from 4, 5, 6, and 7; and m is selected from 0, 1,2, and 3.

3. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 1 or 2, wherein X is -O-.

4. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-3, wherein each Y is independently selected from -C(R^Y)2-, - ( B 2

CO-, and ' ' .

5. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-4, wherein each R^Y is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1.

6.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-5, wherein each R^Y is independently selected from:

H₃C. ,CH₃ ,OH

CH₃ OH ³ Y ³ (

7.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-6, wherein each Q is independently selected from:

C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from

8.

halogen and Ci-Cô alkyl.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-7, wherein each Q is independently selected from:

9.

10.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-8, wherein Ring B is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-9, wherein Ring B is selected from:

F

11.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-10, wherein -(Y)_n- is a group selected from:

100

12. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-11, wherein each Y is -C(R^Y)2-.

13. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-12, wherein each R¹ is independently selected from -CF3 and -N(R²)2-

14. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-13, wherein each R² is independently selected from hydrogen and Ci-Cô alkyl.

15. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-14, wherein each R² is hydrogen.

16. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to ^^RZ1 , · · V^rZ2 any one of Embodiments 1-15, wherein Z is .

17. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-16, wherein R^Z1 is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen).

18. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-17, wherein R^Z1 is -CF3.

19. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-18, wherein R²² is hydroxy.

101

20. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-19, wherein n is selected from 4, 5, and 6.

21. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-20, wherein n is 6.

22. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-21, wherein m is selected from 1 and 2.

23. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1-22, wherein m is 2.

24. A compound selected from compounds of Formula la:

( B ) each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ^Y ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^YI, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

102

C3-C8 cycloalkyl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH2, and -NHCOMe), o Ci-Cf, alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl), and o C3-C8 cycloalkyl,

halogen,

103

each R² is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

A,R^Z1 „R^Z3 j]

V^R^Z2 -s/^Sl'R^Z3 VR^Z1 Z2

Z is selected from ' , and R ;

R²¹ is selected from hydrogen, -CN, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen or 1-3 hydroxy), 3- to 6-membered heterocyclyl, 3- to 6-membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R²¹ and R^Z2 taken together form a group selected from oxo and =N-OH;

each R²³ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CôCio aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl; and n is selected from 4, 5, 6, and 7.

25. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 24, wherein X is -O-.

26. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 24 or 25, wherein each Y is independently selected from -C(R^Y)2-, -CO-, and

104

27. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-26, wherein each R^Y is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1.

28. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-27, wherein each R^Y is independently selected from:

29. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-28, wherein each Q is independently selected from:

C3-C8 cycloalkyl,

30. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-29, wherein each Q is independently selected from:

, and

31. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-30, wherein Ring B is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

32. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-31, wherein Ring B is selected from:

' f , ' ' , and ' ' .

33. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-32, wherein -(Y)_n- is a group selected from:

105

34. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-33, wherein each Y is -C(R^Y)2-.

35. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-34, wherein each R¹ is independently selected from -CF3 and -N(R²)2.

36. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-35, wherein each R² is independently selected from hydrogen and Ci-Cô alkyl.

37. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to 15 any one of Embodiments 24-36, wherein each R² is hydrogen.

106

38. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to <^R^Z1 /^_RZ2 any one of Embodiments 24-37, wherein Z is .

39. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-38, wherein R^Z1 is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen).

40. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-39, wherein R^Z1 is -CF3.

41. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-40, wherein R^Z2 is hydroxy.

42. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-41, wherein n is selected from 4, 5, and 6.

43. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 24-42, wherein n is 6.

44. A compound selected from compounds of Formulae Ha, Hb, and Ile:

(b ) each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and x ' ;

Ring B is selected from:

107

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Ci-Cô alkyl, and Ci-Ci, alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

3- to 6-membered heterocyclyl (optionally substituted with 1-3 groups independently selected from Ci-Cf, alkyl);

each Q is independently selected from:

halogen,

108

each R¹ is independently selected from halogen, -CF3, -OR², -N(R²)2, -CO2R², -CON(R²)2, -CN, phenyl, benzyl, Ci-Cô alkoxy, Ci-Ce, alkyl, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

45.

R^Z1 is selected from hydrogen, -CN, Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen and hydroxy), 3- to 6-membered heterocyclyl, 3- to 6membered cycloalkyl, 5- to 6-membered aryl, and 5- to 6-membered heteroaryl;

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R^ZI and R²² taken together form a group selected from oxo and =N-OH;

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 44, wherein m is selected from 1 and 2.

109

46.

47.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 44 or 45, wherein m is 2.

A compound selected from compounds of Formulae Ild, Ile, and Ilf:

( B ] each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ' ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C6-C10 aryl, 5to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

110

C3-C8 cycloalkyl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o Ci-Cf, alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH₂, and -NHCOMe), o Ci-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl), and o C3-C8 cycloalkyl,

halogen,

111 each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)2, -CN, phenyl, benzyl, Ci-Cô alkoxy, Ci-Cô alkyl, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

R^Z2 is selected from hydrogen, halogen, and hydroxy, or R^ZI and R^Z2 taken together form a group selected from oxo and =N-OH; and each R²⁵ is independently selected from hydroxy, Ci-Cô alkoxy, Ci-Cô alkyl, and CôCio aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl.

48. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-47, wherein X is -O-.

49. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-48, wherein each Y is independently selected from -C(R^Y)2-, £ B )

-CO-, and ' < .

50. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-49, wherein each R^Y is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1.

51. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-50, wherein each R^Y is independently selected from:

112

52. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-51, wherein each Q is independently selected from:

C3-C8 cycloalkyl,

53. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-52, wherein each Q is independently selected from:

, and

54. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-53, wherein Ring B is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

55. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-54, wherein Ring B is selected from:

56. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-55, wherein each Y is

-C(R^v)2-.

57. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-56, wherein each R¹ is independently selected from -CF3 and -N(R²)₂.

58. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-57, wherein each R² is independently selected from hydrogen and Ci-Cô alkyl.

59. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-58, wherein each R² is hydrogen.

113

60.

61.

62.

63.

64.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to ^^RZ1A_rZ2 any one of Embodiments 44-59, wherein Z is .

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-60, wherein R^Z1 is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen).

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-61, wherein R^Z1 is -CF3.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 44-62, wherein R^Z2 is hydroxy.

A compound selected from compounds of Formulae HIa, Illb, and IIIc:

( B J each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ^γ ' ;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

114

each Q is independently selected from:

halogen,

o halogen,

115 o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C3-C8 cycloalkyl (optionally substituted with 1-3 CF3 groups), and o 3- to 10-membered heterocyclyl,

65.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 64, wherein m is selected from 1 and 2.

116

66. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 64 or 65, wherein m is 2.

67. A compound selected from compounds of Formulae IHd, HIe, and IHf:

R¹ N-N IHd, R¹ N-N me, and R¹ N-N nif, and deuterated dérivatives and pharmaceutically acceptable salts thereof, wherein:

X is selected from -O-, -S-, -SO-, and -SO2-;

(b J each Y is independently selected from -C(R^Y)2-, -O-, -CO-, and ^Y ' ;

each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), Cô-Cio aryl, 5to 10-membered heteroaryl, -OR^Y1, -CÛ2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3- to 6-membered heterocyclyl; or two instances of R^Y, one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

Ring B is selected from:

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

117

halogen,

118 each R¹ is independently selected from halogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), -OR², -N(R²)2, -CO2R², -CON(R²)2, -CN, phenyl, benzyl, Ci-Cf, alkoxy, Ci-Cf, alkyl, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

each R² is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Ci-Ce alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

each R²³ is independently selected from hydroxy, Ci-Ce alkoxy, Ci-Cf, alkyl, and Cf,C10 aryl; or two instances of R²³ are taken together to form a 3- to 6-membered heterocyclyl.

68. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-67, wherein X is -O-.

69. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-68, wherein each Y is independently selected from -C(R^Y)2-, ( B J

-CO-, and ' f .

70. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-69, wherein each R^Y is independently selected from hydrogen, Ci-Cf, alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), and -OR^Y1.

71. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-70, wherein each R^Y is independently selected from:

119 ch₃ oh hydrogen, -A , Ά ,

and

72. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-71, wherein each Q is independently selected from:

C3-C8 cycloalkyl,

73. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-72, wherein each Q is independently selected from:

a ju jO ' , ^x , and ' .

74. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-73, wherein Ring B is selected from Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen.

75. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-74, wherein Ring B is selected from:

F .A

76. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-75, wherein each Y is -C(R^Y)2-.

77. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-76, wherein each R¹ is independently selected from -CF3 and -N(R²)₂.

78. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-77, wherein each R² is independently selected from hydrogen and Ci-Cô alkyl.

79. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-78, wherein each R² is hydrogen.

80. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to

A\r^z1

2^- rZ2 any one of Embodiments 64-79, wherein Z is .

120

81.

82.

83.

84.

85.

86.

87.

88.

89.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-80, wherein R^Z1 is selected from Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen).

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-81, wherein R^Z1 is -CF3.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 64-82, wherein R^Z2 is hydroxy.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 1, wherein X is -O-.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to Embodiment 1 or 84, wherein each R^Y is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, and -OR^YI.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1, 84, and 85, wherein -OR^Y1 is -OH.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-86, wherein each Q is independently selected from:

C3-C8 cycloalkyl,

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-87, wherein each Q is independently selected from:

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-88, wherein each R^Y is independently selected from:

CH₃ OH hydrogen, fluorine, —L- , —L-

121

90. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-89, wherein Ring B is selected from C3-C8 cycloalkyl and phenyl optionally substituted with 1-3 groups independently selected from halogen.

91. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-90, wherein Ring B is selected from:

F

92. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-91, wherein n is selected from 4, 5, and 6.

93. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-92, wherein -(Y)_n- is a group selected from:

F F

z OH

122

94.

95.

96.

97.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-93, wherein each R¹ is independently selected from Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -N(R²)2, and -CO2R².

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 85-94, wherein each R² is independently selected from hydrogen and Ci-Cô alkyl.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-95, wherein each R¹ is independently selected from CF₃, -NH2, -NH(CH₂CH3), CO2H, and CH₂OH.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to 'sZ R²² any one of Embodiments 1 and 84-96, wherein Z is selected from and

123

98.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-97, wherein the group:

^)0-2 is selected from:

R²²,

N

H

N H

N H N

F F

99.

N H

L '^NN H , and

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-98, wherein the group:

(R^Z2)o-2

R^Z2 and is selected from:

NH N

100.

101.

102.

103.

104.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-99, wherein R^ZI is selected from hydrogen and Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen).

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-100, wherein R^Z1 is selected from hydrogen and -CF3. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-101, wherein R²² is hydroxy.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-102, wherein Z is selected from:

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1 and 84-103, wherein m is selected from 1 and 2.

124

105.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1,2, and 84-104, wherein:

X is -O-;

Ring B is selected from C3-C8 cycloalkyl groups:

each Q is independently selected fronrCs-Cs cycloalkyl and Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen and Ci-Cô alkyl, each R* is independently selected from Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen) and -NH2;

106.

R²² is hydroxy;

n is selected from 5 and 6; and m is 2.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1, 2, and 84-105, wherein each Q is independently selected from:

107.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1, 2, and 84-106, wherein each R^Y is independently selected from:

125

108. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1, 2, and 84-107, wherein Ring B is .

109. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1, 2, and 84-108, wherein -(Y)_n- is a group selected from:

110. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to 10 any one of Embodiments 1, 2, and 84-109, wherein R^Z1 is -CF3.

111. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1,2, and 84-110, wherein n is 5.

112. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of Embodiments 1,2, and 84-110, wherein n is 6.

113. A compound selected from compounds of Table 12, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing.

114. A compound selected from compounds of Table 13, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing.

115. A compound according to Embodiment 113, wherein the compound is selected from:

126

127

deuterated dérivatives thereof, and pharmaceutically acceptable salts of any of the foregoing.

116. A compound according to Embodiment 114, wherein the compound is selected from:

128

117. A pharmaceutical composition comprising a compound, deuterated dérivative, or pharmaceutically acceptable sait of any one of Embodiments 1-116 and a pharmaceutically acceptable carrier.

118. The pharmaceutical composition according to Embodiment 117, further comprising one or more additional therapeutic agent(s).

119. The pharmaceutical composition according to Embodiment 118, wherein the one or more additional therapeutic agent(s) comprise(s) a compound with CFTR modulating activity 10 or a sait or deuterated dérivative thereof.

120. The pharmaceutical composition according to Embodiment 118 or 119, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR corrector.

121. The pharmaceutical composition according to any one of Embodiments 118-120, wherein the one or more additional therapeutic agent(s) comprise(s) (R)-1-(2,215 difluorobenzo[d][l,3]dioxol-5-yl)-7V-(l-(2,3-dihydroxypropyl)-6-fluoro-2-(l-hydroxy-2methylpropan-2-yl)-l//-indol-5-yl)cyclopropanecarboxamide (Compound II):

129

122. The pharmaceutical composition according to any one of Embodiments 118-121, wherein the one or more additional therapeutic agent(s) comprise(s) 3-(6-(1-(2,2difluorobenzo[d][l,3]dioxol-5-yl)cyclopropanecarboxamido)-3-methylpyridin-2yl)benzoic acid (Compound IV):

123. The pharmaceutical composition according to any one of Embodiments 118-122, wherein the one or more additional therapeutic agent(s) comprise(s) 7V-(l,3-dimethylpyrazol-4yl)sulfonyl-6-[3-(3,3,3-trifluoro-2,2-dimethyl-propoxy)pyrazol-l-yl]-2-[(4S)-2,2,4trimethylpyrrolidin-l-yl]pyridine-3-carboxamide (Compound V):

/

124. The pharmaceutical composition according to any one of Embodiments 118-123, wherein the one or more additional therapeutic agent(s) comprise(s) 7V-(benzenesulfonyl)-6-[3-[2[ 1 -(trifluoromethyl) cyclopropyl]ethoxy]pyrazol-l -yl]-2-[(4S)-2,2,4-trimethylpyrrolidinl-yl]pyridine-3-carboxamide (Compound VI):

125. The pharmaceutical composition according to any one of Embodiments 118-124, wherein the one or more additional therapeutic agent(s) comprise(s) (14S)-8-[3-(2{dispiro[2.0.2.1]heptan-7-yl}ethoxy)-177-pyrazol-l-yl]-12,12-dimethyl-2X⁶-thia-

130

3,9,11,18,23-pentaazatetracyclo [17.3.1.11 l,14.05,10]tetracosa-l(22),5,7,9,19(23),20hexaene-2,2,4-trione (Compound Vil):

126. The pharmaceutical composition according to any one of Embodiments 118-125, wherein the one or more additional therapeutic agent(s) comprise(s) (117?)-6-(2,6dimethylphenyl)-ll-(2-methylpropyl)-12-{spiro[2.3]hexan-5-yI}-9-oxa-2X⁶-thia3,5,12,19-tetraazatricyclo[ 12.3.1.14,8]nonadeca-1 ( 17),4( 19),5,7,14( 18), 15-hexaene2,2,13-trione (Compound VIII):

127. The pharmaceutical composition according to any one of Embodiments 118-126, wherein the one or more additional therapeutic agent(s) comprise(s) at least one compound selected from PTI-428, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV3748, ABBV-3903, ABBV-119, and PTI-801.

128. The pharmaceutical composition according to any one of Embodiments 118-127, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR potentiator enhancer.

129. The pharmaceutical composition according to any one of Embodiments 118-128, wherein the one or more additional therapeutic agent(s) comprise(s) ASP-11.

130. A method of treating cystic fibrosis, comprising administering an effective amount of the compound, sait, or deuterated dérivative according to any one of Embodiments 1-116 or the pharmaceutical composition according to any one of Embodiments 117-129 to a patient in need thereof.

131. The method according to Embodiment 130, further comprising administering one or more additional therapeutic agent(s).

131

132. The method according to Embodiment 131, wherein the one or more additional therapeutic agent(s) comprise(s) a compound with CFTR modulating activity or a sait or deuterated dérivative thereof.

133. The method according to Embodiment 131 or 132, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR corrector.

134. The method according to any one of Embodiments 131-133, wherein the one or more additional therapeutic agent(s) comprise(s) (Æ)-l-(2,2-difIuorobenzo[d][l,3]dioxol-5-yl)7V-(1 -(2,3-dihydroxypropyl)-6-fluoro-2-( 1 -hydroxy-2-methylpropan-2-yl)-177-indol-5yl)cyclopropanecarboxamide (Compound II):

135. The method according to any one of Embodiments 131-134, wherein the one or more additional therapeutic agent(s) comprise(s) 3-(6-(1-(2,2-difluorobenzo[d][l,3]dioxol-5yl)cyclopropanecarboxamido)-3-methylpyridin-2-yl)benzoic acid (Compound IV):

136. The method according to any one of Embodiments 131-135, wherein the one or more additional therapeutic agent(s) comprise(s) 7V-(l,3-dimethylpyrazol-4-yl)sulfonyl-6-[3(3,3,3 -trifluoro-2,2-dimethyl-propoxy)pyrazol-1 -yl]-2-[(45)-2,2,4-trimethylpyrrolidin-1 yl]pyridine-3-carboxamide (Compound V):

137. The method according to any one of Embodiments 131-136, wherein the one or more additional therapeutic agent(s) comprise(s) 7V-(benzenesulfonyl)-6-[3-[2-[l(trifluoromethyl) cyclopropyl]ethoxy]pyrazol-l-yl]-2-[(45)-2,2,4-trimethylpyrrolidin-lyl]pyridine-3-carboxamide (Compound VI):

132

/ VI.

138. The method according to any one of Embodiments 131-137, wherein the one or more additional therapeutic agent(s) comprise(s) (145)-8-[3-(2-{dispiro[2.0.2.1]heptan-7yl}ethoxy)-177-pyrazol-l-yl]-12,12-dimethyl-2X⁶-thia-3,9,l 1,18,23-pentaazatetracyclo [17.3.1.11 l,14.05,10]tetracosa-l(22),5,7,9,19(23),20-hexaene-2,2,4-trione (Compound VII):

139. The method according to any one of Embodiments 131-138, wherein the one or more additional therapeutic agent(s) comprise(s) (1 UÎ)-6-(2,6-dimethylphenyl)-l l-(2methyIpropyl)-12-{spiro[2.3]hexan-5-yl}-9-oxa-2X⁶-thia-3,5,12,19tetraazatricyclo[12.3.1.14,8]nônadeca-l(17),4(19),5,7,14(18),15-hexaene-2,2,13-trione (Compound VIII):

140. The method according to any one of Embodiments 131-139, wherein the one or more additional therapeutic agent(s) comprise(s) at least one compound selected from PTI-428, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and PTI-801.

141. The method according to any one of Embodiments 131-140, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR potentiator enhancer.

142. The method according to any one of Embodiments 131-141, wherein the one or more additional therapeutic agent(s) comprise(s) ASP-11.

133

143. The compound, deuterated dérivative, or pharmaceutically acceptable sait of any one of Embodiments 1-116 or the pharmaceutical composition according to any one of Embodiments 117-129 for use in the treatment of cystic fibrosis.

144. Use of the compound, deuterated dérivative, or pharmaceutically acceptable sait of any one of Embodiments 1-117 in the manufacture of a médicament for the treatment of cystic fibrosis.

145. Use of the pharmaceutical composition according to any one of Embodiments 117-129 in the manufacture of a médicament for the treatment of cystic fibrosis.

146. Substantially crystalline Compound 11 heptane solvaté (i.e., wherein less than 15% of Compound 11 is in amorphous form, wherein less than 10% of Compound 11 is in amorphous form, wherein less than 5% of Compound 11 is in amorphous form).

147. The Compound 11 according to Embodiment 146, wherein Compound 11 is 100% crystalline heptane solvaté.

148. The crystalline Compound 11 heptane solvaté according to Embodiment 146 or 147, characterized by an X-ray powder diffractogram having one, two, or three signais selected from 5.8 ± 0.2 degrees two-theta, 10.1 ± 0.2 degrees two-theta, and 11.7 ± 0.2 degrees two-theta.

149. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146148, characterized by an X-ray powder diffractogram having (a) one, two, or three signais selected selected from 5.8 ± 0.2 degrees two-theta, 10.1 ± 0.2 degrees two-theta, and 11.7 ± 0.2 degrees two-theta, and (b) one, two, three, or four signais selected from 5.6 ± 0.2 degrees two-theta, 18.1 ± 0.2 degrees two-theta, 20.5 ± 0.2 degrees two-theta, and 20.9 ± 0.2 degrees two-theta.

150. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146149, characterized by an X-ray powder diffractogram having signais at 5.6 ± 0.2 degrees two-theta, 5.8 ± 0.2 degrees two-theta, 10.1 ± 0.2 degrees two-theta, 11.7 ± 0.2 degrees two-theta, 18.1 ± 0.2 degrees.

151. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146150, characterized by an X-ray powder diffractogram substantially similar to FIG. 1.

152. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146151, characterized by a ¹³C ssNMR spectrum having one, two, three, four, five, six, seven, eight, nine, ten, or more peaks selected from 166.3 ± 0.2 ppm, 165.8 ± 0.2 ppm, 164.6 ± 0.2 ppm, 163.4 ± 0.2 ppm, 154.8 ± 0.2 ppm, 154.0 ± 0.2 ppm, 152.1 ± 0.2 ppm, 151.6 ± 0.2 ppm, 140.2 ± 0.2 ppm, 139.4 ± 0.2 ppm, 138.5 ± 0.2 ppm, 138.0 ± 0.2 ppm, 135.1 ± 0.2 ppm, 134.6 ± 0.2 ppm, 131.3 ± 0.2 ppm, 130.2 ± 0.2 ppm, 129.6 ± 0.2 ppm, 134

128.5 ± 0.2 ppm, 125.7 ± 0.2 ppm, 123.7 ± 0.2 ppm, 123.2 ± 0.2 ppm, 122.9 ± 0.2 ppm, 121.1 ± 0.2 ppm, 120.2 ± 0.2 ppm, 119.2 ± 0.2 ppm, 117.8 ± 0.2 ppm, 76.2 ± 0.2 ppm, 74.4 ± 0.2 ppm, 73.7 ± 0.2 ppm, 73.3 ± 0.2 ppm, 40.0 ± 0.2 ppm, 38.6 ± 0.2 ppm, 37.6 ± 0.2 ppm, 36.9 ± 0.2 ppm, 35.7 ± 0.2 ppm, 33.6 ± 0.2 ppm, 32.5 ± 0.2 ppm, 32.0 ± 0.2 ppm, 30.4 ± 0.2 ppm, 30.1 ± 0.2 ppm, 29.5 ± 0.2 ppm, 28.8 ± 0.2 ppm, 28.1 ± 0.2 ppm, 27.1 ± 0.2 ppm, 25.3 ± 0.2 ppm, 23.1 ± 0.2 ppm, 22.7 ± 0.2 ppm, 22.0 ± 0.2 ppm, 21.6 ± 0.2 ppm, 20.3 ± 0.2 ppm, 19.6 ± 0.2 ppm, 18.3 ± 0.2 ppm, 17.6 ± 0.2 ppm, 13.8 ± 0.2 ppm, 13.1 ± 0.2 ppm, and 12.5 ± 0.2 ppm.

153. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146152, characterized by a ^I3C SSNMR spectrum substantially similar to FIG. 3.

154. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146153, characterized as having a ¹⁹F SSNMR spectrum with one, two, three, four, five, or more peaks selected from -63.5 ± 0.2 ppm, -63.8 ± 0.2 ppm, -65.1 ± 0.2 ppm, -65.8 ± 0.2 ppm, -66.3 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, and -76.6 ± 0.2 ppm.

155. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146154, characterized as having a ¹⁹F SSNMR spectrum with one, two, three, four, five, or more peaks selected from -63.5 ± 0.2 ppm, -63.8 ± 0.2 ppm, -65.1 ± 0.2 ppm, -65.8 ± 0.2 ppm, -66.3 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, -76.6 ± 0.2 ppm, and -77.6 ± 0.2 ppm.

156. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146155, characterized as having a ¹⁹F SSNMR spectrum with a peak at -67.0 ± 0.2 ppm.

157. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146156, characterized as having a ^I9F SSNMR spectrum with a peak at -65.1 ± 0.2 ppm.

158. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146157, characterized as having a ¹⁹F SSNMR spectrum with a peak at -76.6 ± 0.2 ppm.

159. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146158, characterized as having a ¹⁹F SSNMR spectrum with a peak at -63.5 ± 0.2 ppm.

160. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146159, characterized as having a ¹⁹F SSNMR spectrum with a peak at -74.9 ± 0.2 ppm.

161. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146160, characterized as having a ¹⁹F SSNMR spectrum with at least one peak selected from -65.1 ± 0.2 ppm, -67.0 ± 0.2 ppm, and -76.6 ± 0.2 ppm.

135

162. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146161, characterized as having a ^,9F SSNMR spectrum with peaks at -65.1 ± 0.2 ppm, -67.0 ± 0.2 ppm, and -76.6 ± 0.2 ppm.

163. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146162, characterized as having a ^,9F SSNMR spectrum with at least one peak selected from -63.5 ± 0.2 ppm, -65.1 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, and -76.6 ± 0.2 ppm.

164. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146163, characterized as having a ^l9F SSNMR spectrum with peaks at -63.5 ± 0.2 ppm, -65.1 ± 0.2 ppm, -67.0 ± 0.2 ppm, -74.9 ± 0.2 ppm, and -76.6 ± 0.2 ppm.

165. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146164, characterized by a ^l9F SSNMR spectrum substantially similar to FIG. 4.

166. The crystalline Compound 11 heptane solvaté according to any one of Embodiments 146165, prepared by a process comprising dissolving Compound 11 in heptane and dichloromethane, concentrating under rotary évaporation, swirling at room température, filtering the solids, washing the solids with cold heptane, and drying under vacuum to provide Compound 11 heptane solvaté.

167. A method of preparing the crystalline Compound 11 heptane solvaté according to any one of Embodiments 146-166, prepared by a process comprising dissolving Compound 11 in heptane and dichloromethane, concentrating under rotary évaporation, swirling at room température, filtering the solids, washing the solids with cold heptane, and drying under vacuum to provide Compound 11 heptane solvaté.

168. The pharmaceutical composition according to any one of Embodiments 118-126, wherein the one or more additional therapeutic agent(s) comprise(s) at least one compound selected from Compound II, Compound III, Compound ΙΠ-d, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, PTI-428, ASP-11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, FDL-169, ARN5562, ARN21586, ARN22081, ARN22652, ARN23765, ARN23766, PTI-801, FDL-176, PTI-808 (dirocaftor), GLPG1837, GLPG2451/ABBV-2451, QBW251 (icenticaftor), GLPG3067/ABBV-3067 (Navocaftor), ABBV-191, ELX-02, MRT5005, Lunar-CF, RCT223, amiloride, ETD001, CF552, GS-9411, GS-5737, P-1037 (VX-371), P-1055 (VX-551), AZD5634, SPX-101, Ionis-ENaC-2.5 Rx, BI 1265162, AZ5634, ARO-ENaClOOl, ETD002, and DS-1039.

169. The method according to any one of Embodiments 131-139, wherein the one or more additional therapeutic agent(s) comprise(s) at least one compound selected

136

Compound II, Compound III, Compound ΙΙΙ-d, Compound IV, Compound V, Compound VI, Compound VII, Compound VIII, Compound IX, Compound X, PTI-428, ASP-11, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV3903, ABBV-119, FDL-169, ARN5562, ARN21586, ARN22081, ARN22652, ARN23765, ARN23766, PTI-801, FDL-176, PTI-808 (dirocaftor), GLPG1837, GLPG2451/ABBV-2451, QBW251 (icenticaftor), GLPG3067/ABBV-3067 (Navocaftor), ABBV-191, ELX-02, MRT5005, Lunar-CF, RCT223, amiloride, ETD001, CF552, GS9411, GS-5737, P-1037 (VX-371), P-1055 (VX-551), AZD5634, SPX-101, lonis-ENaC2.5 Rx, BI 1265162, AZ5634, ARO-ENaClOOl, ETD002, and DS-1039.

170. Substantially crystalline Compound 6 (free form) (i.e., wherein less than 15% of Compound 6 is in amorphous form, wherein less than 10% of Compound 6 is in amorphous form, wherein less than 5% of Compound 6 is in amorphous form).

171. The Compound 6 (free form) according to Embodiment 170, wherein Compound 6 is 100% crystalline Compound 6 (free form).

172. The crystalline Compound 6 (free form) according to Embodiment 170 or 171, characterized by a monoclinic crystal System, a P2\ space group, and unit cell dimensions measured at 100 K on a Bruker diffractometer equipped with Cu Ka radiation (λ=1.5478 Â) of:

a 9.6 ± 0.1 Â a 90° b 13.6 ± 0.1 Â β 1O5.3°±O.1° c 13.8 ± 0.1 Â γ 90°

173. Substantially crystalline Compound 19 (free form) (i.e., wherein less than 15% of Compound 19 is in amorphous form, wherein less than 10% of Compound 19 is in amorphous form, wherein less than 5% of Compound 19 is in amorphous form).

174. The Compound 19 (free form) according to Embodiment 173, wherein Compound 19 is 100% crystalline Compound 19 (free form).

175. The crystalline Compound 19 (free form) according to Embodiment 173 or 174, characterized by a tetragonal crystal System, a P4i2i2 space group, and unit cell dimensions measured at 100 K on a Bruker diffractometer equipped with Mo Ka radiation (λ=0.71073 Â) of:

a 9.8 ± 0.1 Â a90° b 9.8 ± 0.1 Â β90° c 37.1 ± 0.1 Â γ90°

137

176. Substantially crystalline Compound 20 (free form) (i.e., wherein less than 15% of Compound 20 is in amorphous form, wherein less than 10% of Compound 20 is in amorphous form, wherein less than 5% of Compound 20 is in amorphous form).

177. The Compound 20 (free form) according to Embodiment 176, wherein Compound 20 is 100% crystalline Compound 20 (free form).

178. The crystalline Compound 20 (free form) according to Embodiment 176 or 177, characterized by an orthorhombic crystal System, a P212121 space group, and unit cell dimensions measured at 100 K on a Bruker diffractometer equipped with Mo Ka radiation (λ=0.71073 Â) of:

a 10.7 ± 0.1 Â a90° b 13.7 ± 0.1 Â β90° c 25.5 ± 0.1 Â γ90°

Examples

General Experimental Procedures

Abbreviations

ACN: Acetonitrile

AcOH: Acetic acid

BCh: Boron trichloride

Boc anhydride ((BocjiO): Di-teri-butyl dicarbonate

CDCI3: Chloroform-tf

CDI: 1,1 '-Carbonyldiimidazole

CD3OD: Methyl-i/3 alcohol-J

CH2CI2: Dichloromethane

CH3CN: Acetonitrile

CO2: Carbon dioxide

CS2CO3: Césium carbonate

CuBn: Copper(II) bromide

Cul: Copper(I)iodide

DCE: 1,2-Dichloroethane

DCM: Dichloromethane

DDQ: 2,3-Dichloro-5,6-dicyano-l ,4-benzoquinone

DI: Deionized

DIAD: Diisopropyl azodicarboxylate

138

DIEA: DIPEA; A/A^-Diisopropylcthylaminc

DMAP: 4-Dimethylaminopyridine

DMF: W.W-Dimethylformamide

DMSO: Dimethyl sulfoxide

DMSO-Jô: Dimethyl sulfoxide-cfc

EA: Ethyl acetate

ELSD: Evaporative light scattering detector

EtiO: Diethyl ether

EtOAc: Ethyl acetate

EtOH: Ethanol

ESI-MS: Electrospray ionization mass spectrometry

Grubbs l^st Génération catalyst: Dichloro(benzylidene)bis(tricyclohexylphosphine)ruthenium(II)

Grubbs 2^nd Génération catalyst: [l,3-Bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]dichloro-[(2-isopropoxyphenyl)methylene]ruthenium

Fh: Hydrogen

HATU: 7V-[(Dimethylamino)-lH-l,2,3-triazolo-[4,5-b]pyridin-l-ylmethylene]-7Vmethylmethanaminium hexafluorophosphate TV-oxide

HCl: Hydrochloric acid

HFIP: Hexafluoroisopropanol

Hoveyda-Grubbs 2^nd Génération catalyst: Dichloro[l,3-bis(2,4,6-trimethylphenyl)-2imidazolidinylidene](2-isopropoxyphenylmethylene)ruthenium(II)

HPLC: High performance liquid chromatography

IP A: Isopropanol

IP AC: Isopropyl acetate iPrOH: Isopropanol

KHSO4: Potassium bisulfate

LC: Liquid chromatography

LCMS: Liquid chromatography mass spectrometry

LD A: Lithium diisopropylamide

LiOH: Lithium hydroxide

MeCN: Acetonitrile

MeTHF or 2-MeTHF: 2-Methyltetrahydrofuran

MeOH: Methanol

MTBE: Methyl teri-butyl ether

139

MgSO* Magnésium sulfate n-BuLi: n-Butyllithium

NaBH4: Sodium borohydride

NaHCCh: Sodium bicarbonate

NaHMDS: Sodium bis(trimethylsilyl)amide

NaOH: Sodium hydroxide

Na2S2C>3: Sodium thiosulfate

Na2SO4: Sodium sulfate

NBS: 7V-Bromosuccinimide

NMP: 7V-Methyl-2-pyrrolidone

NMR: Nuclear magnetic résonance

Pd/C: Palladium on carbon

Pd(OAc)2: Palladium(II) acetate rt: Room température

SFC: Supercritical fluid chromatography

Silica Cat Pd: Palladium on Silica

SilicaMetS: Silica Supported Métal Scavenger

S1O2: Silica gel

T3P: 1-Propanephosphonic anhydride

TBAI: Tetrabutylammonium iodide

TEA: Triethylamine

TFA: Trifluoroacetic acid

THF: Tetrahydrofuran

UPLC: Ultra Performance Liquid Chromatography

Xantphos: 4,5-Bis(diphenylphosphino)-9,9-dimethylxanthene

XPhos Pd G3: (2-Dicyclohexylphosphino-2',4',6'-triisopropyl-l,l'-biphenyl)[2-(2'-amino-l,rbiphenyl)]palladium(II) methanesulfonate

Zhan catalyst-1 B: Dichloro[l,3-bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene][[5[(dimethylamino)sulfonyl]-2-(l-methylethoxy-O)phenyl]methylene-C]ruthenium(II)

General Methods

Reagents and starting materials were obtained by commercial sources unless otherwise stated and were used without purification.

140

Proton and carbon NMR spectra were acquired on either a Broker Biospin DRX 400 MHz FTNMR spectrometer operating at a ’H and ¹³C résonant frequency of400 and 100 MHz respectively, or on a 300 MHz NMR spectrometer. One dimensional proton and carbon spectra were acquired using a broadband observe (BBFO) probe with 20 Hz sample rotation at 0.1834 and 0.9083 Hz/Pt digital resolution respectively. Ail proton and carbon spectra were acquired with température control at 30 °C using standard, previously published puise sequences and routine processing parameters.

NMR (1D & 2D) spectra were also recorded on a Broker AVNEO 400 MHz spectrometer operating at 400 MHz and 100 MHz respectively equipped with a 5 mm multinuclear Iprobe.

NMR spectra were also recorded on a Varian Mercury NMR instrument at 300 MHz for *H using a 45 degree puise angle, a spectral width of4800 Hz and 28860 points of acquisition. FID were zero-filled to 32k points and a line broadening of 0.3Hz was applied before Fourier transform. ¹⁹F NMR spectra were recorded at 282 MHz using a 30 degree puise angle, a spectral width of 100 kHz and 59202 points were acquired. FID were zero-filled to 64k points and a line broadening of 0.5 Hz was applied before Fourier transform.

NMR spectra were also recorded on a Broker Avance III HD NMR instrument at 400 MHz for *H using a 30 degree puise angle, a spectral width of 8000 Hz and 128k points of acquisition. FID were zero-filled to 256k points and a line broadening of 0.3Hz was applied before fourrier transform. ¹⁹F NMR spectra were recorded at 377 MHz using a 30 deg puise angle, a spectral width of 89286 Hz and 128k points were acquired. FID were zero-filled to 256k points and a line broadening of 0.3 Hz was applied before Fourier transform.

NMR spectra were also recorded on a Broker AC 250MHz instrument equipped with a: 5mm QNP(H1/C13/F19/P31) probe (type: 250-SB, s#23055/0020) or on a Varian 500MHz instrument equipped with a ID PFG, 5 mm, 50-202/500 MHz probe (model/part# 99337300).

Unless stated to the contrary in the following examples, final purity of compounds was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient ron from 1-99% mobile phase B over 3.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C. Final purity was calculated by averaging the area under the curve (AUC) of two UV traces (220 nm, 254 nm). Low-resolution mass spectra were reported as [M+l]⁺ species obtained using a single quadropole mass spectrometer equipped with an electrospray ionization

141 (ESI) source capable of achieving a mass accuracy of 0.1 Da and a minimum resolution of 1000 (no units on resolution) across the détection range.

Solid-state NMR (SSNMR) spectra were recorded on a Bruker-Biospin 400 MHz widebore spectrometer equipped with Bruker-Biospin 4mm HFX probe. Samples were packed into 4mm ZrO2 rotors and spun under Magic Angle Spinning (MAS) condition with spinning speed typically set to 12.5 kHz. The proton relaxation time was measured using *H MAS Ti saturation recovery relaxation experiment in order to set up proper recycle delay of the ¹³C crosspolarization (CP) MAS experiment. The fluorine relaxation time was measured using ¹⁹F MAS Ti saturation recovery relaxation experiment in order to set up proper recycle delay of the ^l9F MAS experiment. The CP contact time of carbon CPMAS experiment was set to 2 ms. A CP proton puise with linear ramp (from 50% to 100%) was employed. The carbon Hartmann-Hahn match was optimized on extemal reference sample (glycine). Both carbon and fluorine spectra were recorded with proton decoupling using TPPM15 decoupling sequence with the field strength of approximately 100 kHz.

General Synthetic Schemes

Another aspect of the disclosure provides methods for making compounds of Formulae I, I’, I”, I’”, la, Ha, Ha’, Hb, Ile, Hd, Ile, Hf, Ilia, HIa’, Illb, IIIc, IHd, IHe, and IHf, Compounds 1 to 53, Compounds 54 to 77, and pharmaceutically acceptable salts of any of those compounds, deuterated dérivatives of any of the foregoing, and intermediates for making any of the foregoing. In some embodiments of the following Schemes and Examples, each nitrogen and oxygen atom may optionally hâve, in addition to or in place of a specified variable substituent, one or more protecting groups selected from the range of protecting groups disclosed herein. In some embodiments of the following Schemes and Examples, each compound may be replaced with its deuterated dérivative.

142

Scheme 1

S1-2

S1-3

S1-4

S1-1

(Y)l-2 °\ !

h₂n-^{nh OBn}

S1-7

Scheme 1 refers to processes for preparing an intermediate compound of Formula Sl-7 from a compound of Formula Sl-1. Alk is selected from Ci-Cô linear or branched alkyl groups.

X¹ is selected from halogens such as Cl, I, or Br. Y and R^Y are as defined for Formula I above.

Any suitable conditions for a Grignard addition can be used to react a compound of Formula Sl-1 with a compound of Formula Sl-2 to form a compound of Formula Sl-3. For example, the Grignard addition of a compound of Formula Sl-1 with a compound of Formula Sl-2 may be performed in EtiO at -78 °C, followed by addition of 1 N aqueous HCl to yield a compound of Formula Sl-3. Conversion of a compound of Formula Sl-3 to a compound of

Formula Sl-4 may be accomplished by any suitable benzylation procedure. Conversion of an ester of Formula Sl-4 to a carboxylic acid of Formula Sl-5 may be accomplished by any suitable hydrolysis conditions. For example, conversion of a carboxylic acid of Formula Sl-5 to a compound of Formula Sl-6 may be accomplished by reacting a compound of Formula Sl-5 with

HATU and Et₃N in DMF, followed by addition of teri-butyl 7V-aminocarbamate. Any suitable hydrolysis conditions may be used to couvert a carbamate of Formula Sl-6 to a hydrazide of Formula Sl-7. For example, a compound of Formula Sl-7 may be obtained by reacting a compound of Formula Sl-6 with HCl in CH2CI2 at ambient température.

143

Scheme 2

rrcF₃h₂n-^{nh OBn}

R^Y (Y)i-2 O. /

S1-7

S2-3

Scheme 2 refers to processes for preparing an intermediate compound of Formula S2-3 from a compound of Formula S2-1. R^A1 is selected from -X-(Y)2-4-C(R^y)=C(R^y)2, -OH, -OPG (wherein PG is a suitable protecting group), and halogen. R¹, m, X, Y, and R^Y are as defined for Formula I above.

Any suitable conditions to form an amide bond can be used to produce a compound of Formula S2-2 from a compound of Formula S2-1 and a compound of Formula S1-7. For example, a compound of Formula S2-1 can be reacted with CDI in acetonitrile and DMF, followed by addition of a compound of Formula Sl-7, to yield a compound of Formula S2-2. A compound of Formula S2-2 can be converted to a compound of Formula S2-3 using any conditions suitable for oxadiazole formation. For example, a compound of Formula S2-2 can be reacted with DIPEA in acetonitrile, followed by addition ofp-toluenesulfonyl chloride, to yield an oxadiazole of Formula S2-3.

144

Scheme 3

Scheme 3 refers to processes for preparing a compound of Formula S3-8 from a compound of Formula S3-1. Alk is selected from Ci-Cô linear or branched alkyl groups. LG is 5 selected from halogens and oxygen-based leaving groups such as OTf. R¹, m, Y and R^Y are as defined for Formula I above.

The reaction of a compound of Formula S3-1 with a compound of Formula S3-2 to yield a compound of Formula S3-3 may be accomplished by any suitable aromatic substitution conditions. For example, a compound of Formula S3-2 may be reacted with sodium hydride in 10 DMF, followed by addition to a compound of Formula S3-1, to yield a compound of Formula S3-3. Conversion of an ester of Formula S3-3 to a carboxylic acid of Formula S3-4 may be accomplished by any suitable hydrolysis conditions. A compound of Formula S3-5 may be prepared from a compound of Formula S3-4 and a compound of Formula S1-7 using any suitable amide bond formation conditions. A compound of Formula S3-5 can be converted to a compound of Formula S3-6 using any conditions suitable for oxadiazole formation. For example, a compound of Formula S3-5 can be reacted with methoxycarbonyl(triethylammonio)sulfonyl-azanide in THF to yield an oxadiazole of Formula S3-6.

145

Macrocyclization of a compound of Formula S3-6 may be accomplished by any suitable ringclosing metathesis conditions. For example, a compound of Formula S3-6 may be reacted in the presence of [l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]-dichloro-[(2-isopropoxy-5nitro-phenyl)methylene]ruthenium in DCE to yield a macrocycle of Formula S3-7 as a mixture of EIZ isomers (as denoted by the bond). Conversion of an unsaturated compound of Formula S3-7 to a macrocycle of Formula S3-8 can be accomplished using any suitable procedure for olefin réduction and benzyl deprotection.

Scheme 4

Scheme 4 refers to processes for preparing a compound of Formula S4-4 from a compound of Formula S4-1. LG is selected from halogens, hydroxy, and oxygen-based leaving groups such as OTf. R¹, m, Y and R^Y are as defined for Formula I above.

The reaction of a compound of Formula S4-1 with a compound of Formula S3-2 to yield a compound of Formula S4-2 may be accomplished by any suitable aromatic substitution conditions or Mitsunobu conditions. For example, a compound of Formula S4-1 may be reacted an alcohol of Formula S3-2 with césium carbonate and iodocopper in DMSO. Macrocyclization of a compound of Formula S4-2 may be accomplished by any suitable ring-closing metathesis conditions. For example, a compound of Formula S4-2 may be reacted in the presence of Grubbs 2^nd génération catalyst in DCE to yield a macrocycle of Formula S4-3 as a mixture of E/Z isomers (as denoted by the bond). Conversion of an unsaturated compound of Formula S4-3 to a macrocycle of Formula S4-4 can be accomplished using any suitable procedure for olefin réduction and benzyl deprotection.

146

S5-3

S5-6

S5-7

Scheme 5 refers to processes for preparing a compound of Formula S5-3, a compound of Formula S5-6, and a compound of Formula S5-7 from a compound of Formula S5-1. R¹, m, and 5 Y are as defined for Formula I above.

The conversion of a compound of Formula S5-1 to a deuterated compound of Formula S5-2 may be accomplished by any suitable catalytic deutération conditions. For example, a compound of Formula S5-1 may be reacted with 10% palladium on carbon in CD3OD under a deuterium gas atmosphère to yield a compound of Formula S5-2. Conversion of a benzyl10 protected compound of Formula S5-2 to a free alcohol of Formula S5-3 may be accomplished by any suitable deprotection conditions.

Conversion of an unsaturated compound of Formula S5-1 to an alcohol of Formula S5-4 may be accomplished by any suitable hydroboration/oxidation conditions. For example, a compound of Formula S5-1 may be reacted with borane dimethylsulfide complex in THF, 15 followed by quenching with aqueous NaOH and a subséquent addition of hydrogen peroxide to yield an alcohol of Formula S5-4 as a mixture of regioisomers. Débenzylation of a compound of Formula S5-4 to yield a compound of Formula S5-5 may be accomplished using any suitable

147 benzyl deprotection conditions. Conversion of a compound of Formula S5-5 to a compound Formula S5-7 may be accomplished by any suitable oxidation conditions. For example, a compound of Formula S5-5 may be reacted with NalICCh and Dess-Martin periodinane in CH2CI2 to yield a compound of Formula S5-7.

In an alternative route, conversion of a compound of Formula S5-4 to a compound of Formula S5-6 may be accomplished by any suitable oxidation conditions. For example, a compound of Formula S5-4 may be reacted with Dess-Martin periodinane in CH2CI2 to yield a compound of Formula S5-6. Débenzylation of a compound of Formula S5-6 to yield a compound of Formula S5-7 may be accomplished using any suitable benzyl deprotection conditions.

Scheme 6

S6-4 (E/Z mixture)

S6-5

Scheme 6 refers to processes for preparing a compound of Formula S6-5 from a compound of Formula S6-1. LG is selected from halogens and oxygen-based leaving groups such as OTf. R¹, m, Y, and R^Y are as defined for Formula I above.

The conversion of a compound of Formula S6-1 and a compound of Formula S6-2 to a compound of Formula S6-3 may be accomplished by any suitable aromatic substitution conditions. For example, a compound of Formula S6-1 may be reacted with a compound of Formula S6-2 and DMSO. Macrocyclization of a compound of Formula S6-3 may be accomplished by any suitable ring-closing metathesis conditions. For example, a compound of Formula S6-3 may be reacted in the presence of benzylidene-[l,3-bis(2,4,6trimethylphenyl)imidazolidin-2-ylidene]-dichloro-ruthenium;tricyclohexylphosphane in DCE to yield a macrocycle of Formula S6-4 as a mixture of E/Z isomers (as denoted by the bond).

148

Conversion of an unsaturated compound of Formula S6-4 to a macrocycle of Formula S6-5 can be accomplished using any suitable procedure for olefin réduction and benzyl deprotection.

Scheme 7

S7-4 (E/Z mixture)

S7-5 S7-6

Scheme 7 refers to processes for préparation of a compound of Formula S7-6 from a compound of Formula S7-1. R¹, m, Y, and R^Y are as defined for Formula I above. LG is selected from halogens and oxygen-based leaving groups such as OTf.

Reaction of a compound of Formula S7-1 with a compound of Formula S7-2 to form a compound of Formula S7-3 can be accomplished by any suitable lithiation procedure. For example, the reaction of a compound of Formula S7-1 with a compound of Formula S7-2 may be performed in ether at -78 °C with π-BuLi to form a compound of Formula S7-3. Conversion of a compound of Formula S7-3 to a compound of Formula S7-4 may be accomplished by any suitable ring-closing metathesis procedure. For example, the ring-closing metathesis reaction of the compound of Formula S7-3 may be accomplished in the presence ofbenzylidene-[l,3bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]-dichloro-ruthenium;tricyclohexylphosphane in DCE to yield a compound of Formula S7-4 as a mixture of E/Z isomers (as denoted by the bond). Conversion of a compound of Formula S7-4 to a compound of Formula S7-5 may be accomplished by any suitable procedure for olefin réduction and benzyl deprotection. Conversion of a compound of Formula S7-5 to a compound of Formula S7-6 may be accomplished by any suitable procedure for oxidizing a thioether to a sulfoxide.

149

Scheme 8

S8-7 (E/Z mixture)

S8-8

Scheme 8 refers to processes for preparing a compound of Formula S8-8 from a compound of Formula S8-1. Alk is selected from Ci-Cô linear or branched alkyl groups. LG is 5 selected from oxygen-based leaving groups such as OTf and halogens such Cl, I, and Br. R¹, m, Y, R^Y, and Ring B are as defined for Formula I above.

Any suitable conditions for synthesizing an aryl ether from an alcohol and an aryl halide can be used to react a compound of Formula S8-1 with a compound of Formula S8-2 to yield a compound of Formula S8-3. Any suitable conditions for condensation of a hydrazide with a carboxylic acid can be used to react a compound of Formula S8-3 with a compound of Formula S8-4 to form a compound of Formula S8-5. Any suitable conditions for oxadiazole formation from a hydrazide can be used to convert a compound of Formula S8-5 to a compound of Formula S8-6. Conversion of a compound of Formula S8-6 to a compound of Formula S8-7 may be accomplished by any suitable ring-closing metathesis procedure. For example, the ring-closing metathesis reaction of the compound of Formula S8-6 may be accomplished in the presence of benzylidene-[l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]-dichlororuthenium;tricyclohexylphosphane in DCE to yield a compound of Formula S8-7 as a mixture of 150

E/Z isomers (as denoted by the bond). Conversion of a compound of Formula S8-7 to a compound of Formula S8-8 may be accomplished by any suitable procedure for olefin réduction and benzyl deprotection.

Scheme 9

S9-1

macrocyclization

S9-3

S9-4 S9-5

S9-6

S9-7

Scheme 9 refers to processes for preparing a compound of Formula S9-6 and a compound of Formula S9-7 from a compound of Formula S9-1. R¹, m, Y, R^Y, and Ring B are as defined for Formula I above. LG is selected from oxygen-based leaving groups such as OTf and halogens such Cl, I, and Br. L^x is selected from halogens such as Cl, I, or Br.

Any suitable conditions for synthesizing an aryl ether from an alcohol and an aryl halide can be used to react a compound of Formula S9-1 with a compound of Formula S9-2 to form a compound of Formula S9-3. Conversion of a compound of Formula S9-3 to a compound of Formula S9-4 and/or a compound of Formula S9-5 may be accomplished by any suitable crosscoupling procedure. For example, the macrocyclization reaction of the compound of Formula

S9-3 may be accomplished in the presence of palladium (II) acetate, tris-o-tolylphosphane, and triethylamine in acetonitrile to yield the compound of Formula S9-4 and/or the compound of 151

Formula S9-5. Conversion of a compound of Formula S9-4 to a compound of Formula S9-6 and conversion of a compound of Formula S9-5 to a compound of Formula S9-7 may be accomplished by any suitable procedure olefin réduction and benzyl deprotection.

Scheme 10

S10-1

S10-4 (E/Z mixture)

S10-5

Scheme 10 refers to processes for preparing a compound of Formula S10-6 from a compound of Formula S10-1. R¹, m, Y, R^Y, and Ring B are as defined for Formula I above. L* is selected from halogens such as Cl, I, or Br.

Reaction of a compound of Formula S10-1 with a compound of Formula S10-2 to yield a compound of Formula S10-3 may be accomplished using any suitable oxadiazole formation procedure. For example, a compound of Formula S10-1 may be reacted with a compound of Formula S10-2 and (isocyanoimino)triphenylphosphorane to yield a compound of Formula S103. Conversion of a compound of Formula S10-3 to a compound of Formula S10-4 may be accomplished by any suitable cross-coupling procedure. For example, the macrocyclization reaction of the compound of Formula S10-3 may be accomplished in the presence of palladium (II) acetate, tris-o-tolylphosphane, and triethylamine in acetonitrile to yield a compound of Formula S10-4 as a mixture of E/Z isomers (as denoted by the ***“ bond). Conversion of a compound of Formula S10-4 to a compound of Formula S10-5 may be accomplished by any suitable procedure for reducing olefins.

152

Procedures for the Synthesis of Common Intermediates

Intermediate 1: Préparation of methyl 6-chIoro-3-nitro-5-(trifluoromethyI)pyridine-2carboxylate

Step 1

Step 2

Step 3

Step 4

Step 1: Methyl l-oxido-5-(trifluoromethyl)pyridin-l-ium-2-carboxylate

Urea hydrogen peroxide (62.7 g, 646.53 mmol) was added portion-wise to a stirred solution of methyl 5-(trifluoromethyl)pyridine-2-carboxylate (40 g, 191.09 mmol) in 1,2dichloroethane (300 mL) at 0 °C. Trifluoroacetic anhydride (107.70 g, 72 mL, 507.65 mmol) was then added over 30 minutes at a température of-10 °C, with cooling bath (CCh/acetone bath). The reaction mixture was then stirred for a further 30 minutes at a température of 0 °C and then for 1 hour at ambient température. The reaction mixture was then poured into cooled icewater (600 mL). The mixture was diluted with dichloromethane (300 mL) and then layers were separated. The aqueous phase was extracted with dichloromethane (2 X 200 mL). The combined organic phase was washed with water (2 X 300 mL) and brine (IX 200 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to give methyl l-oxido-5-(trifluoromethyl)pyridin-l-ium-2-carboxylate (47.6 g, 90%) as light yellow solid. ’H NMR (300 MHz, DMSO-d₆) δ 8.89 (s, 1H), 8.02 - 7.90 (m, 1H), 7.86 - 7.72 (m, 1H), 3.89 (s, 3H) ppm. ¹⁹F NMR (282 MHz, DMSO-d₆) δ -62.00 (s, 3F) ppm. ESI-MS m/z cale. 221.02998, found 222.1 (M+l)⁺; Rétention time: 1.24 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: Methyl 6-hydroxy-5-(trifluoromethyl)pyridine-2-carboxylate

153

Trifluoroacetic anhydride (291.62 g, 193 mL, 1.3885 mol) was added drop-wise to a mixture of methyl l-oxido-5-(trifluoromethyl)pyridin-l-ium-2-carboxylate (51.058 g, 230.66 mmol) in DMF (305 mL) at 0 °C. The mixture was then stirred at room température ovemight. The mixture was concentrated under reduced pressure to remove excess of trifluoroacetic acid. The residual DMF solution was poured dropwise to a 0 °C cooled and stirring water volume (1000 mL). The precipitated solid was collected by filtration and then washed with water (300 mL). The solid was dried over high vacuum to afford methyl 6-hydroxy-5(trifluoromethyl)pyridine-2-carboxylate (45.24 g, 86%) as white solid. *H NMR (300 MHz, DMSO-dô) δ 7.90 (d, J= 7.2 Hz, 1H), 7.03 (d, J= 7.2 Hz, 1H), 4.02 (s, 3H) ppm. ¹⁹F NMR (282 MHz, DMSO-dô) δ -66.39 (s, 3F) ppm. ESI-MS m/z cale. 221.03, found 222.1 (M+l)⁺; Rétention time: 1.43 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 3: Methyl 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylate

To an ice-cooled solution of methyl 6-hydroxy-5-(trifluoromethyl)pyridine-2-carboxylate (33.04 g, 149.41 mmol) in sulfuric acid (200 mL of 18.4 M, 3.6800 mol) was added nitric acid (13 mL of 15.8 M, 205.40 mmol) dropwise. After 5 min, the ice bath was removed, and the reaction mixture was stirred at 38 °C ovemight. The reaction was not completed, nitric acid (3 mL of 15.8 M, 47.400 mmol) was added dropwise at room température and the reaction was heated at 38 °C for 4.5 hours. The reaction was poured slowly into ice-cold water (900 mL) and the mixture was cooled at 0 °C for 15 minutes. Then the résultant solid was isolated by filtration and washed with water (600 mL). The solid was dried ovemight under high vacuum to give methyl 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylate (39.49 g, 99%) as white solid. ’H NMR (300 MHz, DMSO-d₆) δ 8.54 (s, 1H), 3.95 (s, 3H) ppm. ¹⁹F NMR (282 MHz, DMSO-dô) δ -64.56 (s, 3F) ppm. ESI-MS m/z cale. 266.0151, found 267.1 (M+l)⁺; Rétention time: 1.64 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 4: Methyl 6-chloro-3-nitro-5-(trifluoromethyl)pyridme-2-carboxylate

154

A mixture of methyl 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylate (10 g, 37.575 mmol) and phenyl dichlorophosphate (48.008 g, 34 mL, 227.55 mmol) was heated at 170 °C for 90 minutes. After cooling to room température, the mixture was diluted with ethyl acetate (400 mL) and washed with brine (2 X 200 mL). The organic phase was dried on anhydrous sodium sulfate, filtered and concentrated under reduced pressure. Purification by silica gel chromatography (0% to 15% of ethyl acetate in heptanes) provided methyl 6-chloro-3-nitro-5(trifluoromethyl)pyridine-2-carboxylate (5.45 g, 50%) as a yellow solid. *H NMR (300 MHz, CDC1₃) δ 8.75 (s, 1H), 4.07 (s, 3H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -64.12 (s, 3F) ppm. ESI-MS m/z cale. 283.9812, found 285.0 (M+l)⁺; Rétention time: 1.95 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Intermediate 2: Préparation of 6-hydroxy-3-nitro-5-(trifIuoromethyl)pyridine-2-carboxylic acid

Step 1: 6-Hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylic acid

A mixture of methyl 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylate (32 g, 120.24 mmol) in THF (180 mL) and water (180 mL) was treated with lithium hydroxide monohydrate (15.14 g, 360.79 mmol) and stirred at 27 °C ovemight. The crude reaction mixture was cooled at room température and the pH adjusted to 2 with a 0.5 M aqueous solution of hydrochloric acid (380 mL), then transferred to a 1-L separatory tunnel with 2-methyl THF and extracted. The layers were separated and the organic layer was then washed with water (150 mL), brine (150 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to afford 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylic acid (29.61 g, 96%) as off-white solid. Ή NMR (300 MHz, DMSO-d₆) δ 8.45 (s, 1H) ppm. ¹⁹F NMR (282 MHz, DMSO-dâ) δ -64.53 (s, 3F) ppm. ESI-MS m/z cale. 251.9994, found 253.0 (M+l)⁺; Rétention time: 0.79 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

155

Intermediate 3: Préparation of 6-chloro-3-nitro-5-(trifluoromethyI)pyridine-2-carboxyIic acid

Step 1

Step 1: 6-Chloro-3-nitro-5-(trifluoromethyI)pyridine-2-carboxylic acid

To a solution of methyl 6-chloro-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylate (1.14 g, 4.006 mmol) in THF (48.51 mL) and water (24.26 mL) at 0 °C was added lithium hydroxide monohydrate (201.7 mg, 4.807 mmol). The reaction was allowed to warm to room température then stirred for 2 hours. The solution was acidified to pH ~2 - 3 by the addition of 1 N HCl, then extracted with EtOAc. The organic phase was washed with water and brine, then dried over sodium sulfate, filtered and concentrated to afford, as a clear syrup, 6-chloro-3-nitro-5(trifluoromethyl)pyridine-2-carboxylic acid (1.05 g, 97%). ESI-MS m/z cale. 269.9655, found 271.0 (M+l)⁺; Rétention time: 0.37 minutes determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 4: Préparation of methyl 3-[bis(tert-butoxycarbonyl)amino]-6-bromo-5(trifluoromethyl)pyridine-2-carboxyIate

156

Step 1: Methyl 3-(benzhydrylideneamino)-5-(trifluoromethyl)pyridine-2-carboxylate

A mixture of methyl 3-chloro-5-(trifluoromethyl)pyridine-2-carboxylate (47.3 g, 197.43 mmol), diphenylmethanimine (47 g, 259.33 mmol), Xantphos (9.07 g, 15.675 mmol), and césium carbonate (131 g, 402.06 mmol) in dioxane (800 mL) was degassed with bubbling nitrogen for 30 minutes. Pd(OAc)2 (3.52 g, 15.679 mmol) was added and the System was purged with nitrogen three times. The reaction mixture was heated at 100 °C for 18 h. The reaction was cooled to room température and filtered on a pad of Celite. The cake was washed with EtOAc and solvents were evaporated under reduced pressure to give methyl 3-(benzhydrylideneamino)5-(trifluoromethyl)pyridine-2-carboxylate (90 g, 84%) as yellow solid. ESI-MS m/z cale. 384.10855, found 385.1 (M+l)⁺; Rétention time: 2.24 minutes. LCMS Method: Kinetex Cis 4.6 X 50mm 2.6 μΜ, 2.0 mL/min, 95% H2O (0.1% formic acid) + 5% acetonitrile (0.1% formic acid) to 95% acetonitrile (0.1% formic acid) gradient (2.0 min) then held at 95% acetonitrile (0.1% formic acid) for 1.0 min.

Step 2: Methyl 3-amino-5-(trifluoromethyl)pyridine-2-carboxylate

To a suspension of methyl 3-(benzhydrylideneamino)-5-(trifluoromethyl)pyridine-2carboxylate (65 g, 124.30 mmol) in methanol (200 mL) was added HCl (3 M in methanol) (146 mL of 3 M, 438.00 mmol). The mixture was stirred at room température for 1.5 hour then the solvent was removed under reduced pressure. The residue was taken up in ethyl acetate (2 L) and dichloromethane (500 mL). The organic phase was washed with 5% aqueous sodium bicarbonate solution (3 X 500 mL) and brine (2 X 500 mL), dried over anhydrous sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was triturated with heptanes (2 X 50 mL) and the mother liquors were discarded. The solid obtained was triturated with a mixture of dichloromethane and heptanes (1:1,40 mL) and filtered to afford methyl 3-amino-5-(trifluoromethyl)pyridine-2-carboxylate (25.25 g, 91%) as yellow solid. 'H NMR (300 MHz, CDCI3) δ 8.24 (s, 1H), 7.28 (s, 1H), 5.98 (br. s, 2H), 4.00 (s, 3H) ppm. ¹⁹F

157

NMR (282 MHz, CDC1₃) δ -63.23 (s, 3F) ppm. ESI-MS m/z cale. 220.046, found 221.1 (M+l)⁺;

Rétention time: 1.62 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 3: Methyl 3-amino-6-bromo-5-(trifluoromethyl)pyridine-2-carboxylate

To a solution of methyl 3-amino-5-(trifluoromethyl)pyridine-2-carboxylate (18.75 g, 80.91 mmol) in acetonitrile (300 mL) at 0 °C was added portion wise TV-bromosuccinimide (18.7g, 105.3 mmol). The mixture was stirred ovemight at 25 °C. Ethyl acetate (1000 mL) was added. The organic layer was washed with 10% sodium thiosulfate solution (3 X 200 mL) which were back extracted with ethyl acetate (2 X 200mL). The combined organic extracts were washed with saturated sodium bicarbonate solution (3 X 200 mL), brine (200 mL), dried over sodium sulfate and concentrated in vacuo to provide methyl 3-amino-6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (25.46 g, 98%). ^!H NMR (300 MHz, CDCI3) δ 3.934.03 (m, 3H), 6.01 (br. s., 2H), 7.37 (s, 1H) ppm. ¹⁹F NMR (282 MHz, CDCI3) ppm -64.2 (s, 3F). ESI-MS m/z cale. 297.9565, found 299.0 (M+l)⁺; Rétention time: 2.55 minutes. LCMS Method: Kinetex Cis 4.6 X 50 mm 2.6 pM. Temp: 45 °C, Flow: 2.0 mL/min, Run Time: 6 min. Mobile Phase: Initial 95% H2O (0.1% formic acid) and 5% acetonitrile (0.1% formic acid) linear gradient to 95% acetonitrile (0.1% formic acid) for 4.0 min then held at 95% acetonitrile (0.1% formic acid) for 2.0 min.

Step 4: Methyl 3-[bis(Z‘i’rAbutoxycarbonyl)amino]-6-bromo-5-(trifIuoromethyl)pyridine-2carboxylate

A mixture of methyl 3-amino-6-bromo-5-(trifluoromethyl)pyridine-2-carboxylate (5 g, 15.549 mmol), (Boc)2Û (11 g, 11.579 mL, 50.402 mmol), DMAP (310 mg, 2.5375 mmol) and CH2CI2 (150 mL) was stirred at room température ovemight. The reaction mixture was concentrated under reduced pressure and purification by silica gel chromatography (0 - 15% ethyl acetate in heptane) provided methyl 3-[bis(ter/-butoxycarbonyl)amino]-6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (6.73 g, 87%) as light yellow solid. *H NMR (300 158

MHz, CDCh) δ 1.42 (s, 18H), 3.96 (s, 3H), 7.85 (s, 1H) ppm. ¹⁹F NMR (282 MHz, CDC1₃) δ 63.9 (s, 3F) ppm. ESI-MS m/z cale. 498.06134, Rétention time: 2.34 minutes. LCMS Method: Kinetex Cis 4.6 X 50 mm 2.6 μΜ. Temp: 45 °C, Flow: 2.0 mL/min, Run Time: 3 min. Mobile Phase: Initial 95% H2O (0.1% formic acid) and 5% acetonitrile (0.1% formic acid) linear gradient to 95% acetonitrile (0.1% formic acid) for 2.0 min then held at 95% acetonitrile (0.1% formic acid) for 1.0 min.

Intermediate 5: Préparation of 6-bromo-3-(tert-butoxycarbonylamino)-5(trifluoromethyl)pyridine-2-carboxylic acid

Step 1

Step 1: 6-Bromo-3-(n?/7-butoxycarbonylamino)-5-(trifluoromethyI)pyridine-2-carboxyIic acid

To a mixture of methyl 3-[bis(/er/-butoxycarbonyl)amino]-6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (247 g, 494.7 mmol) in THF (1.0 L) was added a solution of LiOH (47.2 g, 1.971 mol) in water (500 mL). The mixture was stirred at ambient température for 18 h affording a yellow slurry. The mixture was cooled with an ice-bath and slowly acidified with HCl (1000 mL of 2 M, 2.000 mol) keeping the reaction température <15 °C. The mixture was diluted with heptane (1.5 L), mixed and the organic phase separated. The aqueous phase was extracted with heptane (500 mL). The combined organic phases were washed with brine, dried over MgSO j, filtered and concentrated in vacuo. The crude oil was dissolved in heptane (600 mL), seeded and stirred at ambient température for 18 h affording a thick slurry. The slurry was diluted with cold heptane (500 mL) and the precipitate collected using a medium frit. The filter cake was washed with cold heptane and air dried for 1 h, then in vacuo at 45 °C for 48 h to afford 6-bromo-3-(ter/-butoxycarbonylamino)-5(trifluoromethyl)pyridine-2-carboxylic acid (158.3 g, 83%). *H NMR (400 MHz, DMSO-dô) δ 10.38 (s, 1H), 9.01 (s, 1H), 1.50 (s, 9H) ppm. ESI-MS m/z cale. 383.99326, found 384.9 (M+l)⁺; Rétention time: 2.55 minutes. LCMS Method Detail: Final purity was determined by 159 reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60

Intermediate 6: Préparation of methyl 3-amino-6-bromo-5-fluoro-pyridine-2-carboxylate

nh₂

Step 1

NH₂ O

Step 2

Step 1: Methyl 3-amino-5-fluoro-pyridine-2-carboxylate

In an autoclave (600 mL) was added 2-bromo-5-fluoro-pyridin-3-amine (22 g, 115.18 mmol), methanol (250 mL), triethylamine (23.232 g, 32 mL, 229.59 mmol) and [Ι,Γbis(diphenylphosphino)ferrocene]dichloropalladium(II) (2.1 g, 2.8700 mmol). The autoclave was purged with nitrogen, then with carbon monoxide. The mixture was heated to 130 °C and the carbon monoxide pressure was adjusted to 120 psi. The mixture was stirred for 3 h at 130 °C, then cooled to 25 °C. The mixture was purged with nitrogen and concentrated under vacuum. The resulting solid was diluted with ethyl acetate (500 mL). Water (200 mL) and sodium carbonate (15 g) were added and the mixture was vigorously stirred for 20 minutes. The layers were separated. The organic layer was washed with water (100 mL) and brine (100 mL), dried over sodium sulfate, filtered and evaporated under reduced pressure to provide methyl 3-amino5-fluoro-pyridine-2-carboxylate (14.4 g, 53%) as brown solid. 'H NMR (400 MHz, CDCI3) δ 7.90 (s, 1H), 6.72 (d, J= 9.8 Hz, 1H), 5.94 (br. s, 2H), 3.96 (s, 3H) ppm. ESI-MS m/z cale. 170.04915, found 171.1 (M+l)⁺; Rétention time: 1.35 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: Methyl 3-amino-6-bromo-5-fluoro-pyridine-2-carboxylate

To a solution of methyl 3-amino-5-fluoro-pyridine-2-carboxylate (2.03 g, 11.931 mmol) in acetonitrile (40 mL), A-bromosuccinimide (2.34 g, 13.147 mmol) was added portion-wise. After stirring at room température for 2 h, the reaction mixture was diluted with EtOAc (150 160 mL), washed with a saturated aqueous NaHCCfi (150 mL) and brine (150 mL), then dried over sodium sulfate and concentrated under reduced pressure. Purification by silica gel chromatography (20% to 60% ethyl acetate in heptanes) provided methyl 3-amino-6-bromo-5fluoro-pyridine-2-carboxylate (2.9 g, 98%) as white solid. ’H NMR (300 MHz, CDCh) δ 6.80 (d, J= 8.5 Hz, 1H), 5.98 (br. s., 2H), 4.22 - 3.72 (m, 3H) ppm. ¹⁹F NMR (282 MHz, CDCh) δ 105.70 (d, J - 9.2 Hz, 1F) ppm. ESI-MS m/z cale. 247.9597, found 248.9 (M+l)⁺; Rétention time: 1.73 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Intermediate 7: Préparation of 2-benzyIoxy-2-(trifluoromethyl)hex-5-enehydrazide (hydrochloride sait)

To a solution of ethyl 3,3,3-trifluoro-2-oxo-propanoate (25.15 g, 147.87 mmol) in EtzO (270 mL) at -78 °C was added bromo(but-3-enyl)magnesium in THF (190 mL of 0.817 M, 155.23 mmol) dropwise over a period of 1.5 h (inner température -72 °C to -76 °C). The mixture was stirred at -78 °C for 20 min. The dry ice-acetone bath was removed. The mixture was slowly warm to 5 °C during 1 h, added to a mixture of 1 N aqueous HCl (170 mL) and crushed ice (150 g) (pH = 4). The two layers were separated. The organic layer was concentrated, and the residue was combined with aqueous phase and extracted with EtOAc (2 x 150 mL). The combined organic phase was washed with 5% aqueous NaHCCfi (50 mL) and brine (20 mL), dried with Na2SC>4. The mixture was filtered and concentrated, and co-evaporated with THF (2 X 40 mL) to give ethyl 2-hydroxy-2-(trifluoromethyl)hex-5-enoate (37.44 g, 96%) as colorless oil. 'H NMR (300 MHz, CDCI3) δ 5.77 (ddt, J= 17.0, 10.4, 6.4 Hz, 1H), 5.15 - 4.93 (m, 2H),

161

4.49-4.28 (m, 2H), 3.88 (s, 1H), 2.35 - 2.19 (m, 1H), 2.17- 1.89 (m, 3H), 1.34 (t,7= 7.0 Hz, 3H) ppm. ¹⁹F NMR (282 MHz, CDC1₃) δ -78.74 (s, 3F) ppm.

Step 2: Ethyl 2-benzyIoxy-2-(trifluoromethyI)hex-5-enoate

To a solution of ethyl 2-hydroxy-2-(trifluoromethyl)hex-5-enoate (24.29 g, 87.6% purity, 94.070 mmol) in DMF (120 mL) at 0 °C was added NaH (60% in minerai oil, 5.64 g, 141.01 mmol) portion-wise. The mixture was stirred at 0 °C for 10 min. Benzyl bromide (24.13 g, 141.08 mmol) and TBAI (8.68 g, 23.500 mmol) were added. The mixture was stirred at room température ovemight. NH4CI (3 g, 0.6 eq) was added. The mixture was stirred for 10 min. 30 mL of EtOAc was added, then ice-water was added (400 g). The mixture was extracted with CH2CI2 and the combined organic layers were concentrated. Purification by silica gel chromatography (0 - 20% CH2CI2 in heptanes) provided ethyl 2-benzyloxy-2(trifhioromethyl)hex-5-enoate (26.05 g, 88%) as pink oil. *H NMR (300 MHz, CDCI3) δ 1.34 (t, J =7.2 Hz, 3H), 2.00-2.19 (m, 3H), 2.22-2.38 (m, 1H), 4.33 (q, 7=7.2 Hz, 2H), 4.64 (d, J=10.6 Hz, 1H), 4.84 (d, 7=10.9 Hz, 1H), 4.91-5.11 (m, 2H), 5.62-5.90 (m, 1H), 7.36 (s, 5H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -70.5 (s, 3F) ppm. ESI-MS m/z cale. 316.12863, found 317.1 (M+l)⁺; Rétention time: 2.47 minutes. LCMS Method: Kinetex Cis 4.6 X 50mm 2.6 μΜ. Temp: 45 °C, Flow: 2.0 mL/min, Run Time: 3 min. Mobile Phase: Initial 95% H2O (0.1% formic acid) and 5% acetonitrile (0.1% formic acid) linear gradient to 95% acetonitrile (0.1% formic acid) for 2.0 min then held at 95% acetonitrile (0.1% formic acid) for 1.0 min.

Step 3: 2-BenzyIoxy-2-(trifluoromethyI)hex-5-enoic acid

A solution of sodium hydroxide (7.86 g, 196.51 mmol) in water (60 mL) was added to a solution of ethyl 2-benzyloxy-2-(trifluoromethyl)hex-5-enoate (24.86 g, 78.593 mmol) in methanol (210 mL). The reaction was heated at 50 °C ovemight. The reaction was concentrated to remove methanol, diluted with water (150 mL) and the carboxylate sodium sait was washed with heptane (1 X 100 mL). The aqueous solution was acidified to pH = 2 with aqueous 3N

162 solution of HCl. The carboxylic acid was extracted with dichloromethane (3 X lOOmL) and dried over sodium sulfate. The solution was filtered and concentrated to give 2-benzyloxy-2(trifluoromethyl)hex-5-enoic acid (22.57 g, 97%) as pale yellow oil. *H NMR (300 MHz, DMSO-dô) δ 14.31 (br. s., 1H), 7.55 - 7.20 (m, 5H), 5.93 - 5.70 (m, 1H), 5.17 - 4.91 (m, 2H), 4.85 - 4.68 (m, 1H), 4.67 - 4.55 (m, 1H), 2.32 - 1.94 (m, 4H) ppm. ¹⁹F NMR (282 MHz, DMSOdô) δ -70.29 (s, 3F) ppm. ESI-MS m/z cale. 288.09732, found 287.1 (M-l); Rétention time: 3.1 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 4: tert-Butyl 7V-[[2-benzyloxy-2-(trifluoromethyI)hex-5-enoyl]amino]carbamate

To a solution of 2-benzyloxy-2-(trifluoromethyl)hex-5-enoic acid (21.92 g, 92.4% purity, 70.263 mmol) in DMF (130 mL) was added HATU (37.2 g, 97.836 mmol) and EtsN (15 g, 148.24 mmol). The mixture was stirred for 10 minutes then ierZ-butyl 7V-aminocarbamate (12.2 g, 92.312 mmol) was added. The mixture was stirred at 25 °C ovemight and at 40 °C for 1 h. The mixture was diluted with ice-water (500g) and extracted with CH2CI2. The organic layer dried over anhydrous sodium sulfate and was concentrated. Purification by silica gel chromatography (0-30% EtOAc in heptanes) provided ter/-butyl 7V-[[2-benzyloxy-2(trifluoromethyl)hex-5-enoyl]amino]carbamate (26.08 g, 92%) as white solid. 'H NMR (300 MHz, CDCI3) δ 1.46 (s, 9H), 2.10-2.31 (m, 3H), 2.34-2.51 (m, 1H), 4.60-4.72 (m, 1H), 4.73-4.86 (m, 1H), 4.95-5.19 (m, 2H), 5.83 (ddt, J=16.7, 10.4, 6.1 Hz, 1H), 6.28 (br. s., 1H), 7.30-7.51 (m, 5H), 8.34 (d, J =2.6 Hz, 1H) ppm. ¹⁹F NMR (282 MHz, CDCI3) ppm -73.6 (s, 3F) ppm.

Step 5: 2-Benzyloxy-2-(trifluoromethyI)hex-5-enehydrazide

To a solution of Zer/-butyl 7V-[[2-benzyloxy-2-(trifluoromethyl)hex-5enoyl]amino]carbamate (43.12 g, 107.2 mmol) in CH2Q2 (200 mL) was added HCl (100 mL of 4 M, 400.0 mmol) and the mixture was stirred at ambient température for 7 h. The solvent was

163 removed in vacuo, the residue stripped 2 times from heptane and the résultant solid was dried in vacuo using a high vac for 20 h giving 2-benzyloxy-2-(trifluoromethyl)hex-5-enehydrazide (Hydrochloride sait) (35 g, 96%). Ή NMR (400 MHz, Chloroform-d) δ 9.92 (s, 2H), 7.41 - 7.31 (m, 2H), 7.30 - 7.24 (m, 2H), 7.24 - 7.16 (m, 1H), 5.72 - 5.57 (m, 1H), 5.02 - 4.87 (m, 2H), 4.71 (d, J= 10.9 Hz, 1H), 4.62 (d, J= 11.0 Hz, 1H), 3.70 (s, 2H), 2.34 - 1.85 (m, 4H) ppm. ESI-MS m/z cale. 302.1242, found 303.2 (M+l)⁺; Rétention time: 1.5 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 8: Préparation of [6-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazoI-2-yl]-5-nitro-3-(trifluoromethyl)-2-pyridyl] trifluoromethanesulfonate

Step 2

Step 1 : N -[2-Benzyloxy-2-(trifluoromethyl)hex-5-enoyl]-6-hydroxy-3-nitro-5(trifluoromethyl)pyridine-2-carbohydrazide

To a solution of 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylic acid (29.92 g, 102.66 mmol) in acetonitrile (300 mL) and DMF (60 mL) was added CDI (17.48 g, 107.80 mmol). The mixture was stirred for 0.5 h at room température, then 2-benzyloxy-2(trifluoromethyl)hex-5-enehydrazide (hydrochloride sait) (33.04 g, 97.534 mmol) was added in portions. The reaction mixture was stirred at 26 °C for 19 hours. The reaction mixture was 164 transferred to an extraction funnel rinsing with water (300 mL) and 2-Me THF (400 mL). The mixture was extracted with 2-Me THF (3 X 400 mL). The combined organic layer was washed with 0.5 N aqueous solution of HCl (3 X 300 mL), brine (3 X 250 mL), dried over anhydrous NaiSCh, filtered and concentrated by évaporation under reduced pressure. It was then solubilized twice in dichloromethane (2 X 300 mL) and the volatiles were removed by évaporation under reduced pressure to provide 7V'-[2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (58.5 g, 94%) as brown foam residue. ESI-MS m/z cale. 536.11304, found 537.2 (M+l)⁺. Rétention time: 2.03 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: [6-[5-[l-Benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3j4-oxadiazol-2-yl]-5-nitro-3(trifluoromethyl)-2-pyridyl] trifluoromethanesulfonate

To a 0 °C solution of7V'-[2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]-6-hydroxy-3nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (9.76 g, 16.922 mmol) in dichloromethane (190 mL) was added DIPEA (8.0136 g, 10.8 mL, 62.004 mmol) followed by trifluoromethylsulfonyl trifluoromethanesulfonate (12.410 g, 7.4 mL, 43.985 mmol). The icecold bath was removed after 20 min and the reaction was stirred at room température for 2.5 hours. The mixture was transferred to a separatory funnel provided with ice-cold aqueous 1.0 N solution of HCl, and EtOAc (300 mL). The organic layer was separated, and the aqueous phase extracted with ethyl acetate (2 X 150 mL). The combined organic layer was washed again with ice-cold HCl 1.0 N aqueous solution (60 mL) and brine (3 X 40 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. Purification by silica gel chromatography (0-10% EtOAc in heptanes) provided [6-[5-[l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)-2-pyridyl] trifluoromethanesulfonate (5.334 g, 40%) as an orange oil. ’H NMR (300 MHz, CDCI3) δ 8.74 (s, 1H), 7.50 - 7.27 (m, 5H), 5.87 - 5.68 (m, 1H), 5.12 - 4.96 (m, 2H), 4.88 (d, J= 10.6 Hz, 1H), 4.67 (d, J = 10.9 Hz, 1H), 2.60 - 2.16 (m, 4H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -62.68 (s, 3F), -71.80 (s, 3F), -73.04 (s, 3F) ppm. ESI-MS m/z cale. 650.0518, found 651.1 (M+l)⁺;

165

Rétention time: 3.94 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Intermediate 9: Préparation of tert-butyl A-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

Step 1

Step 2

Step 1: tert-Butyl7V-[2-[[[2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]amino]carbamoyI]-6bromo-5-(trifluoromethyI)-3-pyridyl]carbamate

To a mixture of 6-bromo-3-(terZ-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid (239.2 g, 621.1 mmol) and 2-benzyloxy-2-(trifluoromethyl)hex-5-enehydrazide (hydrochloride sait) (230.1 g, 761.2 mmol) in EtOAc (2.2 L) at ambient température was added pyridine (200 mL, 2.473 mol) which afforded a precipitate. To the mixture was added 1propanephosphonic anhydride (500 g of 50 % w/w, 785.7 mmol) and the reaction mixture was stirred at ambient température for 12 h. The reaction was quenched with the slow addition of NaOH (149 g of 50 % w/w, 1.863 mol) in water (2 L) and the mixture was stirred for 15 min. The organic phase was separated, and the aqueous phase extracted with EtOAc (1 L). The combined organic phases washed with brine, dried over MgSÜ4, filtered and concentrated in vacuo. After half of the solvent was removed, the organic phase was washed 2 times with aqueous HCl (1000 mL of 1 Μ, 1.000 mol). The organic phase was dried over MgSÜ4, filtered and concentrated in vacuo. The crude product was slurried in warm heptane (2.5 L) and MTBE (0.25 L) and the mixture stirred at ambient température for 12 h affording a light yellow slurry. The slurry was filtered, and the résultant filter cake was washed 2 times with IL 10%

166

MTBE/heptane. The off-white solid was air dried for 2h, then in vacuo at 40 °C for 20 h giving ZerZ-butyl 7V-[2-[[[2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (379.9 g, 91%). ’H NMR (400 MHz, DMSO-dô) δ 11.09 (s, 1H), 10.92 (s, 1H), 10.35 (s, 1H), 9.15 (s, 1H), 7.50 (d, J= 7.4 Hz, 2H), 7.36 (dt, J= 24.4, 7.2 Hz, 3H), 5.87 (ddt, J= 16.0, 10.4, 5.2 Hz, 1H), 5.09 (d, J- 16.9 Hz, 1H), 5.02 (d, J = 10.1 Hz, 1H), 4.84 (q, J= 11.4 Hz, 2H), 2.35 - 2.12 (m, 4H), 1.49 (s, 9H) ppm. ESI-MS m/z cale. 668.1069, found 670.9 (M+l)⁺; Rétention time: 3.5 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 5.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60

Step 2: teri-Butyl A-[2-[5-|l-benzyIoxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2 yI]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

ZerZ-Butyl jV-[2-[[[2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]amino]carbamoyl]-6bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (102 g, 150.8 mmol) was dissolved in anhydrous acetonitrile (1000 mL) and DIPEA (92 mL, 528.2 mmol) was added. The résultant orange solution was heated to 70 °C (internai temp) making a clear yellow solution. Then ptoluenesulfonyl chloride (37.4 g, 196.2 mmol) was added in 3 equal portions of 12.47 g separated by 10 minutes and then the reaction was heated for another 30 min. The reaction was cooled to room température and the acetonitrile was concentrated under reduced pressure. To the mixture was added 1000 mL MTBE, then 800 mL water, and the mixture was stirred, and the layers were separated. The organic layer was washed with a solution of citric acid (36.3 g, 188.9 mmol) in 700 mL water, then 400 mL saturated NaHCO₃, then 300 mL brine. The organic layer was then dried over anhydrous MgSCL and concentrated under reduced pressure. The material was purified using silica gel chromatography using a gradient of 15% to 50% of 8% EtOAc in hexanes (B) and Hexanes (A) to provide Zeri-butyl 7V-[2-[5-[l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3pyridyl]carbamate (91.7 g, 93%). ‘H NMR (400 MHz, Chloroform-d) δ 10.18 (s, 1H), 9.35 (s,

167

1H), 7.55 - 7.47 (m, 2H), 7.45 - 7.37 (m, 2H), 7.36 - 7.28 (m, 1H), 5.83 - 5.68 (m, 1H), 5.10 4.93 (m, 2H), 4.82 (d, J = 10.5 Hz, 1H), 4.69 (d, J= 10.5 Hz, 1H), 2.59 - 2.13 (m, 4H), 1.56 (s, 9H) ppm. ESI-MS m/z cale. 650.0963, found 651.0 (M+l)⁺; Rétention time: 3.81 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intcrmcdiate 10: Préparation of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyI)pent-4enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-teri-butoxycarbonylcarbamate

Step 1

Step 1: tert-Butyl A-[2-|5-[l-benzyloxy-l-(trifluoromethyI)pent-4-enyI]-l,3,4-oxadiazol-2yl]-6-bromo-5-(trifluoromethyI)-3-pyridyl]-A-ZerZ-butoxycarbonyl-carbamatc

Into a solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (30 g, 41.910 mmol) in MTBE (300 mL) was added DIEA (6.6780 g, 9 mL, 51.670 mmol), DMAP (0.28 g, 2.2919 mmol) and Boc anhydride (20.1 g, 21.158 mL, 92.097 mmol). The resulting yellow cloudy solution was stirred at 35 °C ovemight. After cooling to room température, the solvent was evaporated. The yellow oily residue was then dissolved in 300 mL DCM and was washed with water (300 mL), followed by brine (300 mL). The organic layer was separated, dried over anhydrous sodium sulfate, filtered, and concentrated. Purification by silica gel chromatography (0% to 20% EtOAc in hexanes) provided ZerZ-butyl 2V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (28.68 g, 87%) as white solid. 'H NMR (500 MHz, DMSO-d₆) δ 8.89 (s, 1H), 7.51 (d, J= 7.4 Hz, 2H), 7.43 (t, J= 7.5 Hz, 2H), 7.35 (t, J = 7.3 Hz, 1H), 5.96 - 5.76 (m, 1H), 5.11 (d, J= 17.2 Hz, 1H),

168

5.01 (d, J=10.1 Hz, 1H), 4.73 (d, 7= 10.7 Hz, 1H), 4.66 (d, 7= 10.6 Hz, 1H), 2.65-2.51 (m, 2H), 2.36-2.17 (m, 2H), 1.27 (d, 7= 23.5 Hz, 18H)ppm.

Intermediate 11: Préparation of 2-benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (hydrochloride sait)

Step 1: Ethyl 2-hydroxy-2-(trifluoromethyI)pent-4-enoate

To a solution of ethyl 3,3,3-trifluoro-2-oxo-propanoate (30 g, 176.38 mmol) in diethyl ether (300 mL) at -78 °C was added allyl(bromo)magnesium (185 mL of 1 M, 185.00 mmol) dropwise over a period of 3 hours (internai température: -74 °C — -76 °C). The mixture was stirred at -78 °C for 45 min. The dry ice-acetone bath was removed. The mixture was warmed to about 10 °C over a period of 1 h and added to a mixture of 1 N aqueous HCl (210 mL) and crushed ice (400 g) (pH 4). The mixture was extracted with EtOAc, washed with 5% aqueous NaHCCh, brine and dried over anhydrous Na2SÜ4. The mixture was filtered, concentrated and co-evaporated with hexane to give ethyl 2-hydroxy-2-(trifluoromethyl)pent-4-enoate (42.2 g, 90%) as light yellow oil. 'H NMR (300 MHz, CDCI3) δ 1.33 (t, 7= 7.1 Hz, 3H), 2.60 - 2.79 (m, 2H), 3.84 (br. s., 1H), 4.24 - 4.48 (m, 2H), 5.09 - 5.33 (m, 2H), 5.59 - 5.82 (m, 1H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -78.5 (s, 3F) ppm.

Step 2: Ethyl 2-benzyloxy-2-(trifluoromethyl)pent-4-enoate

169

To a solution of ethyl 2-hydroxy-2-(trifluoromethyl)pent-4-enoate (18.56 g, 83.105 mmol) in DMF (100 mL) was added NaH (5.3 g, 60 % w/w, 132.51 mmol) at 0 °C. The reaction was stirred for 15 minutes and benzyl bromide (21.14 g, 15 mL, 121.12 mol) and tetrabutyl ammonium iodide (8.5 g, 23.012 mmol) were added. The mixture was stirred at room température ovemight. The reaction was quenched with water (300 mL) and extracted with ethyl acetate (3 X 300 mL) before being washed with brine (500 mL) and dried over sodium sulfate. Purification by silica gel chromatography (20 to 60% DCM in hexanes) provided ethyl 2benzyloxy-2-(trifluoromethyl)pent-4-enoate (22.01 g, 70%) as colorless oil. ’H NMR (250 MHz, CDCb) δ 7.55 - 7.25 (m, 5H), 6.00 - 5.80 (m, 1H), 5.30 - 5.10 (m, 2H), 4.86 (d, J= 10.5 Hz, 1H), 4.68 (d, J= 10.5 Hz, 1H), 4.33 (q, J= 7.0 Hz, 2H), 2.81 (d, J= 7.0 Hz, 2H), 1.34 (t, J= 7.1 Hz, 3H) ppm. ESI-MS m/z cale. 302.113, found 303.5 (M+l)⁺; Rétention time: 4.14 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 3: 2-Benzyloxy-2-(trifluoromethyl)pent-4-enoic acid

Into a solution of ethyl 2-benzyloxy-2-(trifluoromethyl)pent-4-enoate (28.99 g, 95.902 mmol) in methanol (150 mL) was added a solution of NaOH (7.6714 g, 191.80 mmol) in water (50 mL). The reaction mixture was stirred at 40 °C for 3 hours. The reaction mixture was concentrated under vacuum, the residue was diluted with water (200 mL) and washed with diethyl ether (200 mL). The aqueous layer was acidified with concentrated HCl to pH 1 and extracted with diethyl ether (3 X 200 mL). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate, and concentrated under vacuum to fumish 2benzyloxy-2-(trifluoromethyl)pent-4-enoic acid (28.04 g, 99%) as a light yellow liquid. ’H NMR (250 MHz, CDCI3) δ 7.55 - 7.28 (m, 5H), 5.97 - 5.69 (m, 1H), 5.33 - 5.17 (m, 2H), 4.95 4.66 (m, 2H), 2.91 (d, J = 7.1 Hz, 2H) ppm.

Step 4: fert-Butyl A^r-[[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamate

170

To a solution of 2-benzyloxy-2-(trifluoromethyl)pent-4-enoic acid (300 g, 1.094 mol) in DMF (2 L) was added HATU (530 g, 1.394 mol) and DIEA (400 mL, 2.296 mol) and the mixture was stirred at ambient température for 10 min. To the mixture was added Ze/7-butyI Naminocarbamate (152 g, 1.150 mol) and the mixture stirred at ambient température for 36 h. The reaction was quenched with cold water (4 L) and the mixture extracted 2X with EtOAc (2 L). The organic phase was washed brine, dried over MgSCU, filtered and concentrated in vacuo. Purification by silica gel chromatography (0 - 40% EtOAc/hexanes) provided teri-butyl 7V-[[2benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamate (386.49 g, 91%) as an oil which slowly crystallized to an off-white solid. 'H NMR (400 MHz, DMSO) δ 10.00 (d, J— 37.9 Hz, 1H), 8.93 (s, 1H), 7.46 - 7.39 (m, 2H), 7.38 - 7.29 (m, 3H), 6.01 - 5.64 (m, 1H), 5.32 (d, J= 17.1 Hz, 1H), 5.17 (d, J= 10.1 Hz, 1H), 4.77 (s, 2H), 2.96 (qd, J= 15.4, 6.8 Hz, 2H), 1.39 (d, J= 17.3 Hz, 9H) ppm. ESI-MS m/z cale. 388.16098, found 389.0 (M+l)⁺; Rétention time: 2.51 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 5: 2-Benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (hydrochloride sait)

To a solution of ZerZ-butyl JV-[[2-benzyloxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamate (98.5 g, 240.94 mmol) in DCM (400 mL) was added HCl in dioxane (200 mL of 4 M, 800.00 mmol). The mixture was stirred at room température for 2 hours, concentrated and co-evaporated with DCM and hexanes to give 2-benzyloxy-2(trifluoromethyl)pent-4-enehydrazide (hydrochloride sait) (81.15 g, 97%) as an offwhite solid. 'H NMR (500 MHz, DMSO-d₆) δ 11.07 (s, 1H), 7.70 - 7.16 (m, 5H), 5.87 - 5.61 (m, 1H), 5.45 5.09 (m, 2H), 4.79 (s, 2H), 3.6 - 3.4 (m, 2H), 3.23 - 3.07 (m, 1H), 3.04 - 2.87 (m, 1H) ppm. ESI-MS m/z cale. 288.10855, found 289.2 (M+l)⁺; Rétention time: 2.0 minutes. LCMS Method: Waters Cortex 2.7u Cis (3.0mm X 50mm), 55 °C; flow: 1.2 mL/min; mobile phase: 100% water with 0.1% trifluoroacetic acid then 100% acetonitrile with 0.1% trifluoroacetic acid, gradient of 5% to 100% B over 4 min, with équilibration at 100% B for 0.5 min, then 5% B over 1.5 min.

171

Intermediate 12: Préparation of [6-[5-[l-Benzyloxy-l-(trifIuoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)-2-pyridyI] trifluoromethanesulfonate

Step 2

Step 1 : N -[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyI]-6-hydroxy-3-nitro-5(trifluoromethyl)pyridine-2-carbohydrazide

OH

To a solution of 6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylic acid (8.5 g, 29.165 mmol) in acetonitrile (90 mL) and DMF (18 mL) was added CDI (5 g, 30.836 mmol). The mixture was stirred for 0.5 h at room température, then 2-benzyloxy-2(trifluoromethyl)pent-4-enehydrazide (hydrochloride sait) (9 g, 27.716 mmol) was added. The reaction mixture was stirred at room température ovemight. The reaction mixture was transferred to an extraction tunnel rinsing with water (300 mL) and 2-Me THF (400 mL). The mixture was extracted with 2-methyl tetrahydrofuran (3 X 400 mL). The combined organic layer was washed with 0.5 N aqueous solution of HCl (3 X 300 mL), brine (3 X 250 mL), dried over anhydrous Na2SÜ4, filtered and concentrated under reduced pressure. It was then solubilized twice in dichloromethane (2 X 300 mL) and the volatiles were removed under reduced pressure gi ving N -[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl] -6-hydroxy-3 -nitro-5(trifluoromethyl)pyridine-2-carbohydrazide (14.7 g, 75%) as yellow solid. ESI-MS m/z cale. 522.0974, found 523.1 (M+l)⁺; Rétention time: 2.08 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

172

Step 2: [6-[5-[l-Benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-5-nitro-3(trifluoromethyl)-2-pyridyl] trifluoromethancsulfonate

OH

Trifluoromethylsulfonyl trifluoromethancsulfonate (14.758 g, 8.8 mL, 52.308 mmol) was added to7V-[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]-6-hydroxy-3-nitro-5(trifluoromethyl)pyridine-2-carbohydrazide (14.7 g, 20.712 mmol) and DIPEA (9.79 g, 13.2 mL, 75.783 mmol) in dichloromethane (175 mL) at 0 °C. The ice-cold bath was removed after 20 min and the reaction was stirred at room température for 2.5 h. The mixture was transferred to a separatory tunnel with ice-cold aqueous 1.0 N solution of HCl (180 mL), and EtOAc (500 mL). The organic layer was separated, and the aqueous phase extracted with ethyl acetate (2 X 120 mL). The combined organic layer was washed again with ice-cold HCl 1.0 N aqueous solution (120 mL) and brine (3 X 120 mL), dried over anhydrous Na2SÜ4, filtered and concentrated by évaporation under reduced pressure. Purification by silica gel chromatography (0% to 20% of ethyl acetate in heptanes) provided [6-[5-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)-2-pyridyl] trifluoromethancsulfonate (5.425 g, 40%) as an orange viscous oil. *H NMR (300 MHz, CDCh) δ 8.65 (s, 1H), 7.36 - 7.21 (m, 5H), 5.93 5.74 (m, 1H), 5.28 - 5.10 (m, 2H), 4.78 (d, J= 10.9 Hz, 1H), 4.60 (d, J= 10.6 Hz, 1H), 3.21 3.05 (m, 2H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -62.69 (s, 3F), -71.82 (s, 3F), -73.32 (s, 3F) ppm. ESI-MS m/z cale. 636.03613, found 637.1 (M+l)⁺; Rétention time: 4.0 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

173

Intermediate 13: Préparation of tert-butyl A-[2-[5-[l-benzyIoxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

Step 1

Br

Step 2

Step 1: tert-Butyl 7V-[2-[[[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamoyl] 6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

To a mixture of 6-bromo-3-(7er/-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid (53 g, 137.6 mmol) and 2-benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (hydrochloride sait) (55 g, 169.4 mmol) in EtOAc (500 mL) at ambient température was added pyridine (44 mL, 544.0 mmol). To the mixture was added 1-propanephosphonic anhydride (111 g of 50 % w/w, 174.4 mmol) and the reaction mixture stirred at ambient température for 12 h. The reaction was quenched with slow addition of NaOH (35 g of 50 % w/w, 437.5 mmol) in water (500 mL) and the mixture stirred for 15 min. The organic phase was separated, and the aqueous phase extracted with EtOAc (500 mL). The combined organic phases washed with HCl (250 mL of 1 M, 250.0 mmol), brine, dried over MgSÛ4, filtered and concentrated in vacuo. Purification by silica gel chromatography (0 - 20% EtOAc/hexanes) provided ier/-butyl 7V-[2[[[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)3-pyridyl]carbamate (66 g, 73%) as pale pink solid. *H NMR (400 MHz, DMSO) δ 11.11 (s, 1H), 10.91 (s, 1H), 10.40 (s, 1H), 9.16 (s, 1H), 7.47 (d, J= 6.9 Hz, 2H), 7.42 - 7.29 (m, 3H), 5.91 (ddt, J= 17.1, 10.6, 7.1 Hz, 1H), 5.37 (dd,J= 17.2, 1.9 Hz, 1H), 5.22 (dd, J= 10.4, 1.8 Hz, 1H), 4.85 (d, J= 2.1 Hz, 2H), 3.20 - 2.91 (m, 2H), 1.50 (s, 9H) ppm. ESI-MS m/z cale. 654.09125, found 657.0 (M+l)⁺; Rétention time: 3.49 minutes. Final purity was determined by reversed 174 phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: fërt-Butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazoI-2-yl] 6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

Br

A solution of /eri-butyl 7V-[2-[[[2-benzyloxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (2.15 g, 3.2641 mmol) and DIPEA (1.12 g, 1.5 mL, 8.6117 mmol) in acetonitrile (43 mL) was heated at 50 °C, thenp-toluenesulfonyl chloride (765 mg, 4.0127 mmol) was added portion wise at 50 °C. Résultant mixture was stirred at 70 °C for 2 hours. The reaction mixture was cooled, then basified with a saturated solution of sodium bicarbonate (100 mL) and extracted with ethyl acetate (3 X 50 mL). The combined organic layers were washed with brine (50 mL), dried over sodium sulfate, filtered and concentrated. Purification by silica gel chromatography (0% to 10% of ethyl acetate in heptanes) afforded ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (1.7 g, 80%) as yellow viscous oil. Ή NMR (300 MHz, CDCI3) δ 10.18 (br. s, 1H), 9.33 (br. s, 1H), 7.53 - 7.27 (m, 5H), 6.00 - 5.83 (m, 1H), 5.32 - 5.13 (m, 2H), 4.86 - 4.76 (m, 1H), 4.73 - 4.64 (m, 1H), 3.27 - 3.11 (m, 2H), 1.55 (s, 9H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -63.78 (s, 3F), -72.93 (s, 3F) ppm. No ionization by regular ESI method was observed, but ionization was observed using an APCI method: (M-C4Hs+l)⁺⁺ = 580.8. ESI-MS m/z cale. 636.0807, Rétention time: 2.7 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 4 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

175

Intermediate 14: Préparation of terf-butyl A-[2-[5-[(lJ?)-l-benzyIoxy-l(trifluoromethyI)but-3-enyI]-l,3,4-oxadiazoI-2-yl]-6-bromo-5-(trifluoromethyl)-3pyridyl] carbamate

Br

Step 2

Step 1: terf-Butyl A-[2-[[[(2R)-2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino] carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyI]carbamate

The racemic ZerZ-butyl 7V-[2-[[[2-henzyloxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (24.5 g, 37.38 mmol) was purified by préparative chiral SFC by 500 pL injections of a 32 mg/mL solution onto a ChiralPak IC (250 X 21.2 mm), 5pm column eluted at 40 °C at 70 mL/min with 8 % MeOH (20mM NH3) and 92 % CO2. First eluting enantiorner-1 (Peak 1 at rétention time = 4.17 min) to provided ierZ-butyl 7V-[2-[[[(25)-2-benzyloxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (11.73 g, 96%). 'H NMR (400 MHz, Chloroform-d) δ 10.59 (s, 1H), 9.83 (s, 1H), 9.28 (s, 1H), 9.02 (d, J= 29.6 Hz, 1H), 7.48 - 7.33 (m, 5H), 5.96 - 5.77 (m, 1H), 5.41 (d, J= 1.6 Hz, 1H), 5.36 - 5.29 (m, 1H), 4.86 (s, 2H), 3.19 (dd, J= 15.5, 5.9 Hz, 1H), 3.03 (dd, J= 15.5, 7.8 Hz, 1H), 1.53 (s, 9H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.89, -73.76 ppm. ESI-MS m/z cale. 654.09125, found 655.3 (M+l)⁺; Rétention time: 0.53 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 X 2.1 mm, 1.7 pm particle) made by

176

Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 or·

The later eluting enantiomer 2 (Peak 2 at rétention time = 6.63 min) provided ZerZ-butyl 7V-[2-[[[(2Æ)-2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (11.62 g, 95%). ’H NMR (400 MHz, Chloroform-d) δ 10.59 (s, 1H), 9.74 (s, 1H), 9.28 (s, 1H), 9.06 (s, 1H), 7.39 (d, J= 4.4 Hz, 5H), 6.02 - 5.79 (m, 1H), 5.44 - 5.36 (m, 1H), 5.34 (dd, J= 10.3, 1.3 Hz, 1H), 4.91 - 4.81 (m, 2H), 3.19 (dd, J= 15.4, 5.8 Hz, 1H), 3.03 (dd, J= 15.5, 7.8 Hz, 1H), 1.53 (s, 9H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.89, -73.76 ppm. ESI-MS m/z cale. 654.09125, found 657.2 (M+l)⁺; Rétention time: 0.53 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (30 X 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 2: tert-Butyl 7V-[2-[5-[(LR)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazoI 2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

Br

teri-Butyl 7V-[2-[[[(27?)-2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (enantiomer 2) (24.97 g, 38.10 mmol) was dissolved in anhydrous acetonitrile (200 mL) under nitrogen, making a clear yellow solution. DIPEA (19.91 mL, 114.3 mmol) was added, and the solution tumed orange. The solution was heated to 70 °C, then j>-toluenesulfonyl chloride (7.99 g, 41.91 mmol) was added in 3 portions at 30 min intervals and heated for about 3 h. The reaction mixture was cooled to room température and evaporated majority of the acetonitrile at 45 °C. Added 145 mL MTBE, followed by a solution of citric acid (11.0 g, 57.25 mmol) in 250 mL water, stirred, then added 73 mL hexanes. Separated the layers and water layer extracted with MTBE. Combined the organic layers dried over MgSCU, concentrated in vacuo at 45 °C. Purification by silica gel chromatography (15% to 80% of hexanes (as solvent A) in 10% EtOAc/hexanes (as solvent B)) provided ZerZ-butyl N-[2[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5177 (trifluoromethyl)-3-pyridyl]carbamate (20.47 g, 84%). 'H NMR (400 MHz, Chloroform-d) δ 10.18 (s, 1H), 9.34 (s, 1H), 7.48 (d, 7= 7.1 Hz, 2H), 7.39 (t, 7= 7.5 Hz, 2H), 7.31 (t, 7= 7.3 Hz, 1H), 6.00 - 5.81 (m, 1H), 5.25 (d,7= 17.1, 1.6 Hz, 1H), 5.20 (d,7= 10.1, 1.5 Hz, 1H), 4.82 (d,7 = 10.6 Hz, 1H), 4.70 (d, 7= 10.6 Hz, 1H), 3.30 - 3.09 (m, 2H), 1.56 (s, 9H) ppm. ESI-MS m/z cale. 636.0807, found 637.3 (M+l)⁺; Rétention time: 3.81 minutes. LCMS Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 15: Préparation of teri-butyl7V-[2-[5-[(LR)-l-benzyloxy-l(trifIuoromethyl)but-3-enyl]-l,3,4-oxadiazoI-2-yl]-6-hydroxy-5-(trifluoromethyl)-3pyridyl]-7V-fôrt-butoxycarbonyl-carbamate

Step 1

Step 1: tert-Butyl 7V-[2-[5-[(Lff)-l-benzyloxy-l-(trifluorometliyl)but-3-enyl]-l,3,4-oxadiazol 2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-terf-butoxycarbonyl-carbamate

To a stirring solution of ZerZ-butyl 7V-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-terZ-butoxycarbonylcarbamate (5.52 g, 7.485 mmol) in DMSO (35.86 mL) at room température was added césium acetate (1.437 g, 7.486 mmol) and the mixture was capped and heated under nitrogen atmosphère at 80 °C ovemight. Cooled to room température and diluted with saturated aqueous NH4CI then extracted with EtOAc (2X). Combined the organic fractions, dried over MgSCfi, filtered and concentrated to a yellow oil. Purification by silica gel chromatography (100% hexanes to 100% EtOAc) giving as a white solid, ter/butyl A-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-Zeri-butoxycarbonylcarbamate (1.8 g, 36%). ESI-MS m/z cale. 674.2175, found 575.2 (M-Boc)⁺. Rétention time:

178

0.45 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 16: Préparation of te/7-butyI ;V-[2-[5-[(17?)-l-benzyloxy-l(trifluoromethyl)but-3-enyI]-l,3j4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]A-terAbutoxycarbonyl-carbamate

Step 1

Step 1: terf-Butyl A-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol 2-yl]-6-bromo-5-(trifluoromethyI)-3-pyridyl]-7V-teri-butoxycarbonyl-carbamate

di-terZ-Butyl dicarbonate (208 mg, 0.9530 mmol) and triethylamine (400 pL, 2.870 mmol) were added to a solution of fôrAbutyl 7V-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)but3-enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (500 mg, 0.7531 mmol) dissolved in dioxane (5 mL) followed by DMAP (14 mg, 0.1146 mmol). The reaction mixture was stirred for 3 hours at room température. The mixture was concentrated to half of its volume and water was added. Extracted with ethyl acetate and combined organics washed with brine. The organics were separated, dried over sodium sulfate, and evaporated. Purification by silica gel chromatography (0 to 50% EtOAc in hexanes) provided ZerZ-butyl 7V-[2-[5-[( 1 R)-1 benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3pyridyl]-7V-ter/-butoxycarbonyl-carbamate (487 mg, 88%) as white solid. *H NMR (400 MHz, DMSO-dô) δ 8.90 (s, 1H), 7.49 (d, J= 7.5 Hz, 2H), 7.42 (t, J= 7.4 Hz, 2H), 7.35 (t, J= 7.3 Hz, 1H), 5.93 (dq, J= 17.1, 7.6 Hz, 1H), 5.38 (d,J= 17.0 Hz, 1H), 5.25 (d, J= 10.2 Hz, 1H), 4.78 (d, J= 10.6 Hz, 1H), 4.65 (d, J= 10.6 Hz, 1H), 2.50 (p, J= 1.8 Hz, 2H), 1.27 (d, J= 21.4 Hz, 18H) ppm. ESI-MS m/z cale. 736.1331, found 739.2 (M+l)⁺; Rétention time: 1.66 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 179

- 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 17: Préparation of (2/?)-2-benzyIoxy-2-(trifluoromethyl)hex-5-enoic acid

Step 1

Step 2

Step-1: (2Æ)-2-BenzyIoxy-2-(trifluoromethyl)hex-5-enoic acid; (Æ)-4-quinolyI-[(25',45)-5vinylquinuclidin-2-yI] methanol

To a N2 purged jacketed reactor set to 20 °C was added isopropyl acetate (IPAC, 100 L, 0.173 M, 20 Vols), followed by previously melted 2-benzyloxy-2-(trifluoromethyl)hex-5-enoic acid (5.00 kg, 17.345 mol) and cinchonidine (2.553 kg, 8.67 mol) made into a slurry with minor amount of the reaction solvent. The reactor was set to ramp internai température to 80 °C over 1 hour, with solids going in solution upon heating to set température, then the solution was held at température for at least 10 minutes, then cooled to 70 °C held and seeded with chiral sait (50g, 1.0 % by wt). The mixture was stirred for 10 minutes, then ramped to 20 °C internai température over 4 hours, then held ovemight at 20 °C. The mixture was filtered, cake washed with isopropyl acetate (10.0 L, 2.0 vols) and dried under vacuum. The cake was then dried in vacuo (50 °C, vacuum) to afford 4.7 kg of sait. The resulting solid sait was retumed to the reactor by making a slurry with a portion of isopropyl acetate (94 L, 20 vol based on current sait wt), and pumped into reactor and stirred. The mixture was then heated to 80 °C internai, stirred hot slurry for at least 10 minutes, then ramped to 20 °C over 4-6 h, then stirred ovemight at 20 °C. The material was then filtered and cake washed with isopropyl acetate (9.4 L, 2.0 vol), pulled dry, cake scooped out and dried in vacuo (50 °C, vacuum) to afford 3.1 kg of solid. The solid (3.1 kg) and 180 isopropyl acetate (62 L, 20 vol based on sait solid wt) was slurried and added to a reactor, stirred under N2 purge and heated to 80 °C and held at température at least 10 minutes, then ramped to 20 °C over 4-6 hours, then stirred ovemight. The mixture was filtered, cake washed with isopropyl acetate (6.2 L, 2 vol), pulled dry, scooped out and dried in vacuo (50 °C, vac) to afford 2.25 kg of solid sait. The solid (2.25 kg) and isopropyl acetate (45 L, 20 vol based on sait solid wt) was slurried and added to a reactor, stirred under N2 purge and heated to 80 °C, held at température at least 10 minutes, then ramped to 20 °C over 4-6 hours, then stirred ovemight. The mixture was filtered, cake washed with isopropyl acetate (4.5 L, 2 vol), pulled dry, scooped out and dried in vacuo (50 °C to afford (27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoic acid;(7?)-4-quinolyl-[(25',45)-5-vinylquinuclidin-2-yl]methanol (1.886 kg, > 98.0 % ee ) as offwhite to tan solid. Chiral purity was determined by Agilent 1200 HPLC instrument using Phenomenex Lux i-Amylose-3 column (3 pm, 150 X 4.6 mm) and a dual, isocratic gradient run 30% to 70% mobile phase B over 20.0 minutes. Mobile phase A = H2O (0.1 % CF3CO2H). Mobile phase B = MeOH (0.1 % CF3CO2H). Flow rate =1.0 mL/min, injection volume = 2 pL, and column température = 30 °C, sample concentration: 1 mg/mL in 60% acetonitrile/40% water.

Step 2: (2Æ)-2-Benzyloxy-2-(trifluoromethyl)hex-5-enoic acid

A suspension of (27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoic acid; (A)-4-quinolyl[(25',4S)-5-vinylquinuclidin-2-yl]methanol (50 g, 87.931 mmol) in ethyl acetate (500.00 mL) was treated with an aqueous solution of hydrochloric acid (200 mL of 1 M, 200.00 mmol). After stirring 15 minutes at room température, the two phases were separated. The aqueous phase was extracted twice with ethyl acetate (200 mL). The combined organic layer was washed with 1 N HCl (100 mL). The organic layer was dried over sodium sulfate, filtered and concentrated. The material was dried over high vacuum ovemight to give (27?)-2-benzyIoxy-2(trifluoromethyl)hex-5-enoic acid (26.18 g, 96%) as pale brown oil. *H NMR (400 MHz, CDCI3) δ 7.46 - 7.31 (m, 5H), 5.88 - 5.73 (m, 1H), 5.15 - 4.99 (m, 2H), 4.88 (d, J = 10.3 Hz, 1H), 4.70 (d, J= 10.3 Hz, 1H), 2.37 - 2.12 (m, 4H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -71.63 (br s, 3F) ppm. ESI-MS m/z cale. 288.0973, found 287.0 (M-l)'; Rétention time: 2.15 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

181

Intermediate 18: Préparation of (2Æ)-2-benzyloxy-2-(trifluoromethyl)hex-5-enehydrazide

Step 1

Step 2

Step 1: tert-Butyl 7V-[[(2.K)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoyI]amino]carbamate

To a solution of (27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoic acid (365 g, 1.266 mol) in DMF (2 L) was added HATU (612 g, 1.610 mol) and DIEA (450 mL, 2.584 mol) and the mixture was stirred at ambient température for 10 min. To the mixture was added ZerZ-butyl Naminocarbamate (200 g, 1.513 mol) (slight exotherm upon addition) and the mixture was stirred at ambient température for 16 h. The reaction was poured into ice water (5 L). The résultant precipitate was collected by filtration and washed with water. The solid was dissolved in EtOAc (2 L) and washed with brine. The organic phase was dried over MgSC>4, filtered and concentrated in vacuo. The oil was diluted with EtOAc (500 mL) followed by heptane (3 L) and stirred at ambient température for several hours affording a thick slurry. The slurry was diluted with additional heptane and filtered to collect fluffy white solid (343 g). The filtrate was concentrated and purification by silica gel chromatography (0 - 40% EtOAc/hexanes) provided terributyl 7V-[[(27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]amino]carbamate (464 g, 91%, combined with product from crystallization). ESI-MS m/z cale. 402.17664, found 303.0 (M+lBoc)⁺; Rétention time: 2.68 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350) and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: (27?)-2-Benzyloxy-2-(trifluoromethyI)hex-5-enehydrazide

182

To a solution of /er/-butyl 7V-[[(2J?)-2-benzyloxy-2-(trifluoromethyl)hex-5enoyl]amino]carbamate (464 g, 1.153 mol) in DCM (1.25 L) and was added HCl (925 mL of 4 M, 3.700 mol) and the mixture stirred at ambient température for 20 h. The mixture was concentrated in vacuo removing most of the DCM. The mixture was diluted with isopropyl acetate (1 L) and basified to pH = 6 with NaOH (140 g of 50 % w/w, 1.750 mol) in IL of ice water. The organic phase was separated and washed with IL of brine and the combined aqueous phases were extracted with isopropyl acetate (1 L). The combined organic phases were dried over MgSO4, filtered and concentrated in vacuo affording a dark yellow oil of (2/?)-2-benzyloxy2-(trifluoromethyl)hex-5-enehydrazide (358 g, quant.). ’H NMR (400 MHz, CDCh) δ 8.02 (s, 1H), 7.44 - 7.29 (m, 5H), 5.81 (ddt, J= 16.8, 10.1, 6.4 Hz, 1 H), 5.13 - 4.93 (m, 2H), 4.75 (dd, J= 10.5, 1.5 Hz, 1H), 4.61 (d, J= 10.5 Hz, 1H), 3.78 (s, 2H), 2.43 (ddd, J= 14.3, 11.0, 5.9 Hz, 1H), 2.26 - 1.95 (m, 3H) ppm. ESI-MS m/z cale. 302.1242, found 303.0 (M+l)⁺; Rétention time: 2.0 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 19: Préparation of tert-butyl/V-[2-[5-[(LÆ)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yI]-6-bromo-5-(trifluoromethyi)-3pyridyl] carbamate

Br F,(k /L H ^NW\zOH + T ii \.O^NH O ' O i^r RCX /L / Il N ( N S p 0 > JX >-£-cf₃ X-O nH HN-NH 0 k

°0t^CF3 ^{516Ρ 1} 7 O H₂N-NH Br % ^F3%A s Step 2 ------ T λ\ H O \XL·_NH N-N I Π °

183

Step 1: tert-Butyl 7V-[2-[[[(2Æ)-2-benzyloxy-2-(trifluoromethyl)hex-5 enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate

To a mixture of 6-bromo-3-(teri-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid (304 g, 789.3 mmol) and (2/?)-2-benzyloxy-2-(trifluoromethyl)hex-5enehydrazide (270 g, 893.2 mmol) in EtOAc (2.25 L) at ambient température was added DIEA (425 mL, 2.440 mol). To the mixture was slowly added T3P (622 g of 50 % w/w, 977.4 mmol) using an ice-water bath to keep the température < 35 °C (température rose to 34 °C) and the reaction mixture was stirred at ambient température for 18 h. Added additional DIEA (100 mL, 574.1 mmol) and T3P (95 g, 298.6 mmol) and stirred at ambient température for 2 days. Starting material was still observed and an additional T3P (252 g, 792 mmol) was added and stirred for 5 days. The reaction was quenched with the slow addition of water (2.5 L) and the mixture stirred for 30 min. The organic phase was separated, and the aqueous phase extracted with EtOAc (2 L). The combined organic phases were washed with brine, dried over MgSÛ4, filtered and concentrated in vacuo. The crude product was dissolved in MTBE (300 mL) and diluted with heptane (3 L), the mixture stirred at ambient température for 12 h affording a light yellow slurry. The slurry was filtered, and the résultant solid was air dried for 2 h, then in vacuo at 40 °C for 48 h. The filtrate was concentrated in vacuo and purified by silica gel chromatography (0 - 20% EtOAc/hexanes) and combined with material obtained from crystallization providing /er/-butyl 7V-[2-[[[(27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (433 g, 82%). 'H NMR (400 MHz, DMSO) δ 11.07 (s, 1H), 10.91 (s, 1H), 10.32 (s, 1H), 9.15 (s, 1H), 7.53 - 7.45 (m, 2H), 7.45 - 7.28 (m, 3H), 5.87 (ddt, J= 17.0, 10.2, 5.1 Hz, 1H), 5.09 (dq, J= 17.1, 1.3 Hz, 1H), 5.02 (dd, J= 10.3, 1.9 Hz, 1H), 4.84 (q, J= 11.3 Hz, 2H), 2.37 - 2.13 (m, 4H), 1.49 (s, 9H) ppm. ESI-MS m/z cale. 668.1069, found 669.0 (M+l)⁺; Rétention time: 3.55 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

184

Step 2: tert-Butyl A-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromcthyl)pcnt-4-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyI]carbamate

To a solution of teri-butyl 7V-[2-[[[(27?)-2-benzyloxy-2-(trifluoromethyI)hex-5enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (240 g, 358.5 mmol) in anhydrous acetonitrile (1.5 L) under nitrogen was added DIEA (230 mL, 1.320 mol) and the orange solution heated to 70 °C. To the mixture was added p-toluenesulfonyl chloride (80.5 g, 422.2 mmol) in 3 equal portions over 1 h. The mixture was stirred at 70 °C for 9 h then additional j?-toluenesulfonyl chloride (6.5 g, 34.09 mmol) was added. The mixture was stirred for a total of 24 h then allowed to cool to ambient température. Acetonitrile was removed in vacuo affording a dark orange oil which was diluted with EtOAc (1.5 L) and water (1.5 L). The organic phase was separated and washed with 500 mL of IM HCl, 500 mL of brine, dried over MgSO4, filtered and concentrated in vacuo. Purification by silica gel chromatography (0 - 20% EtOAc/hexanes) provided /eri-butyl jV-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (200 g, 86%). 'H NMR (400 MHz, DMSO) δ 10.11 (s, 1H), 9.10 (s, 1H), 7.55 - 7.48 (m, 2H), 7.47 - 7.28 (m, 3H), 5.87 (ddt, J= 16.7, 10.2, 6.4 Hz, 1H), 5.11 (dt, J= 17.2, 1.7 Hz, 1H), 5.01 (dt, J= 10.2, 1.5 Hz, 1H), 4.74 (d, J= 10.6 Hz, 1H), 4.65 (d, J= 10.6 Hz, 1H), 2.55 - 2.42 (m, 2H), 2.30 (qd, J= 11.3, 10.3, 6.9 Hz, 2H), 1.52 (s, 9H) ppm. ESI-MS m/z cale. 650.0963, found 650.0 (M+l)⁺; Rétention time: 3.78 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A - H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermcdiate 20: Préparation of tert-butyl 7V-[2-[5-[(LR)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifIuoromethyl)-3-pyridyI]TV-tert-butoxycarbonyl-carbamate

Step 1

185

Step 1: te/T-Butyl A-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)peiit-4-enyl]-l,3,4 oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-fôrt-butoxycarbonyl-carbamate

To a solution of Zeri-butyl JV-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (222 g, 340.8 mmol) in MTBE (1.333 L) was added DIPEA (65.3 mL, 374.9 mmol) followed DMAP (2.09 g, 17.11 mmol). Added a solution of di-ZerZ-butyl dicarbonate (111.6 g, 511.3 mmol) in MTBE (250 mL) over approx. 8 minutes, and the resulting mixture was stirred for additional 30 min. Added 1 L of water and separated the layers. The organic layer was washed with KHSO4 (886 mL of 0.5 M, 443.0 mmol), 300 mL brine, dried with MgSCL and most (>95%) of the MTBE was evaporated by rotary évaporation at 45 °C, leaving a thick oil. Added 1.125 L of heptane, spun in the 45 °C rotovap bath until dissolved, then evaporated out 325 mL of solvent by rotary évaporation. The rotovap bath temp was allowed to drop to room température and product started crystallizing out during the évaporation. Then put the flask in a -20 °C freezer ovemight. The résultant solid was filtered and washed with cold heptane and dried at room température for 3 days to give Zeri-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-N-/erZ-butoxycarbonyl-carbamate (240.8 g, 94%). ‘H NMR (400 MHz, Chloroform-d) δ 7.95 (s, 1H), 7.52 - 7.45 (m, 2H), 7.44 - 7.36 (m, 2H), 7.36 - 7.29 (m, 1H), 5.83 - 5.67 (m, 1H), 5.08 - 5.00 (m, 1H), 5.00 - 4.94 (m, 1H), 4.79 (d, J= 10.4 Hz, 1H), 4.64 (d, J= 10.4 Hz, 1H), 2.57 - 2.26 (m, 3H), 2.26 - 2.12 (m, 1H), 1.41 (s, 18H) ppm. ESI-MS rn/z cale. 750.14874, found 751.1 (M+l)⁺; Rétention time: 3.76 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

186

Intermediate 21: Préparation of tert-butyl A-[2-[5-[(17î)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazoI-2-yl]-6-hydroxy-5-(trifluoromethyl)-3pyridyl]-7V-tert-butoxycarbonyl-carbamate

Step 1

Step 1: teri-Butyl A-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4 oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-teri-butoxycarbonyl-carbamate

ZerZ-Butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-ier/-butoxycarbonyl-carbamate (280 g, 372.6 mmol) was dissolved in DMSO (1.82 L) (yellow solution) and treated with césium acetate (215 g, 1.120 mol) under stirring at room température. The yellow suspension was heated at 80 °C for 5 h. The reaction mixture was cooled to room température and added to a stirred cold émulsion of water (5.5 L) with 1 kg ammonium chloride dissolved in it and a 1:1 mixture of MTBE and heptane (2 L) (in 20 L). The phases were separated and the organic phase washed water (3X3 L) and with brine (1 X 2.5 L). The organic phase was dried with MgSO4, filtered and concentrated under reduced pressure. The résultant yellow solution was diluted with heptane (~1 L) and seeded with ter/-butyl TV-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-/er/-butoxycarbonyl-carbamate and stirred on the rotavap at 100 mbar pressure at room température for 1.5 h. The solid mass was stirred mechanically for 2 h at room température, résultant thick fine suspension was filtered, washed with dry ice cold heptane and dried under vacuum at 45 °C with a nitrogen bleed for 16 h to give terZ-butyl 7V-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ieri-butoxycarbonyl-carbamate (220 g, 85%) as an off white solid. Ή NMR (400 MHz, DMSO-d₆) δ 13.28 (s, 1H), 8.43 (s, 1H), 7.58 - 7.26 (m, 5H), 5.85 (ddt, J= 16.8, 10.3, 6.5 Hz, 1H), 5.10 (dq, J= 17.2, 1.6 Hz, 1H), 5.01 (dq, J= 10.2, 1.3 Hz, 1H), 4.76 (d, J= 11.0 Hz, 1H), 4.65 (d, J= 11.0 Hz, 1H), 2.55 (dd, J= 9.6, 5.2 Hz, 2H), 2.23 (td, J= 13.2, 10.0, 5.7 Hz, 2H), 1.27 (d, J= 3.8 Hz, 18H) ppm. ESI-MS m/z cale. 688.23315, found 689.0 (M+l)⁺; Rétention time: 3.32 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by 187

Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes.

Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H).

Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Intermediate 22: Préparation of (27?)-2-benzyIoxy-2-(trifIuoromethyl)pent-4-enehydrazide

Step 1

Step 2

Step 1: 2-Benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide

te/7-Butyl 7V-[[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamate (386.49 g, 995.1 mmol) was dissolved in DCM (1.25 L) and toluene (250 mL) and treated with HCl (750 mL of 4 M, 3.000 mol) at room température and the yellow solution was stirred at room température for 18 h. The mixture was concentrated in vacuo and diluted with EtOAc (2 L). The mixture was treated with NaOH (600 mL of 2 Μ, 1.200 mol) and stirred at ambient température for 10 min. The organic phase was separated, washed with 1 L of brine, dried over MgSO4, filtered and concentrated in vacuo and used directly in the ensuing step (trace toluene présent), 2-benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (286 g, 100%). ’H NMR (400 MHz, DMSO) δ 9.34 (s, 1H), 7.40 - 7.22 (m, 5H), 5.69 (ddt, J= 17.1, 10.3, 6.9 Hz, 1H), 5.33 5.23 (m, 1H), 5.15 (dd,J= 10.3, 1.8 Hz, 1H), 4.73 (s, 2H), 4.51 (s, 2H), 3.05 - 2.87 (m, 2H) ppm. ESI-MS m/z cale. 288.10855, found 289.0 (M+l)⁺; Rétention time: 1.32 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 2: (27?)-2-Benzyloxy-2-(trifluoromethyI)pent-4-enehydrazide

188

Racemic 2-benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (5.0 g, 17.35 mmol) was separated by chiral SFC using a ChiralPak IG column (250 X 21.2 mm; 5 pm) at 40 °C using a mobile phase 7% MeOH (plus 20 mM NH3), 93% CO2 at a 70 mL/min flow and concentration of the sample was 111 mg/mL in methanol (no modifier), injection volume = 160 pL with an outlet pressure of 136 bar, détection wavelength of 210 nm providing as the second eluting enantiomer, (27î)-2-benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (1.7 g, 68%) as a white solid. *H NMR (400 MHz, DMSO-d₆) δ 9.31 (s, 1H), 7.48 - 7.39 (m, 2H), 7.39 - 7.25 (m, 3H), 5.77 - 5.62 (m, 1H), 5.28 (dq, J= 17.1,1.6 Hz, 1H), 5.15 (dq, J= 10.2, 1.5 Hz, 1H), 4.73 (s, 2H), 4.51 (s, 2H), 3.00 (dd, J= 15.3, 7.5 Hz, 1H), 2.91 (dd, J= 15.3, 6.4 Hz, 1H) ppm. ESI-MS m/z cale. 288.10855, found 289.2 (M+l)⁺; Rétention time: 1.28 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60

Intermediate 23: Préparation of tert-butyl A-[2-[5-[(17î)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyI)-3 pyridyl] carbamate

Step 1

Step 1: terf-Butyl A^r-[2-[5-[(17î)-l-benzyIoxy-l-(trifIuoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yI]-6-hydroxy-5-(trifluoromethyI)-3-pyridyl]carbamate

To a solution of Zeri-butyl 2V-[2-[5-[(lR)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-tert-butoxycarbonyl-carbamate (1.089 g, 1.449 mmol) in DMSO (13.61 mL) was added (25)-pent-4-en-2-ol (745.5 pL, 7.244 mmol), césium carbonate (1.496 g, 4.591 mmol) and iodocopper (63.93 mg, 0.3357 mmol) and the reaction mixture was heated at 80 °C for 1 h. The reaction mixture poured into saturated

189 aqueous NH4CI and extracted with EtOAc. The organic layer was washed with brine, dried (MgSCU), filtered and concentrated to an orange oil which was dissolved in THF (10.89 mL) and formic acid (10.89 mL, 288.7 mmol) was added, stirred 20 min then added formic acid (10.93 mL, 289.7 mmol) and stirred for 45 min. The reaction was quenched by slowly adding to saturated aqueous NaHCCh (vigorous gas évolution was observed) and EtOAc in a separatory funnel bringing the aqueous layer eventually to pH ~2 - 3. The aqueous layer was removed and the EtOAc layer was washed with saturated aqueous NaHCOs (still vigorous gas évolution) then dried over MgSO4, filtered and concentrated to a yellow syrup which was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc giving several products including ZerZ-butyl 7V-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]carbamate in an impure form. This impure material was further purified by chromatography on a 275 g reverse phase Cis column eluting with 50 - 100% acetonitrile/water giving as a clear syrup, ZerZ-butyl 7V-[2-[5-[(17?)-1 benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)3-pyridyl]carbamate (126 mg, 15%). ESI-MS m/z cale. 588.1807, found 589.2 (M+l)⁺; Rétention time: 0.51 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 -99% mobile phase B over 1.0 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

190

Examples

Example 1: Préparation of 6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-oI, Compound 1

Step 1: Methyl 6-bromo-5-(trifluoromethyl)pyridine-2-carboxylate

A mixture of methyl 6-oxo-5-(trifluoromethyl)-l/Z-pyridine-2-carboxylate (21.15 g, 95.64 mmol), POBn (41.14 g, 143.5 mmol) and DMF (350 mg, 4.788 mmol) in toluene (200 mL) was heated at 110 °C ovemight. The mixture was cooled to 0 °C and poured on crushed ice (200 g). The mixture was neutralized to pH = 7 with KHCO3 (100 g, 10.5 eq) at < 2 °C and extracted with EtOAc (2 X 200 mL). The combined organic layers were washed with 5% aqueous NaHCCh (100 mL) and brine (100 mL) and dried with NaiSCU. The mixture was filtered, and the solvent was removed by évaporation. The residue was triturated with heptanes/EtOAc (20:1) to give 22.92 g of pure product. The filtrate was concentrated to give 4.02 g, which was purified by flash chromatography (heptanes/EtOAc 0 - 30%) to give an additional 3.32 g of product. The two crops were combined to give methyl 6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (26.24 g, 96%) as a white solid. ’H NMR (300 MHz, CDCI3) δ 4.04 (s, 3H), 8.05-8.30 (m, 2H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -63.8 (s, 3F) ppm. ESI-MS m/z cale. 282.9456, found 284.0 (M+l)⁺; Rétention time: 4.04 minutes. LCMS Method: Symmetry, 4.6 X 75 mm 3.5 pm. Temp: 45 °C, Flow: 2.0 mL/min, run time: 8 min. Mobile Phase: Initial 95% H2O (0.1% formic acid) and 5% CH3CN (0.1% formic acid) linear gradient to

191

95% CH3CN (0.1% formic acid) for 6.0 min then held at 95% CH3CN (0.1% formic acid) for 2.0 min.

Step 2: 6-Bromo-5-(trifIuoromethyl)pyridine-2-carboxyIic acid

To a solution of methyl 6-bromo-5-(trifluoromethyl)pyridine-2-carboxylate (10.5 g, 36.968 mmol) in THF (100 mL) was added a solution of lithium hydroxide monohydrate (1.8 g, 42.894 mmol) in water (70 mL). This mixture was stirred 45 min at room température. The THF was evaporated in vacuo. Water (60 mL) was added to the remaining aqueous solution and the pH was adjusted to 3 - 4 by addition of 3 N hydrochloric acid (T < 5 °C) leading to précipitation of the desired product. The solid was recovered by filtration and dried in-vacuo to give 6-bromo-5-(trifluoromethyl)pyridine-2-carboxylic acid (9.46 g, 95%) as a white solid. *H NMR (300 MHz, CDCI3) δ 8.18 - 8.28 (m, 1H), 8.28 - 8.39 (m, 1H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -63.8 (s, 3F) ppm. ESI-MS m/z cale. 268.92993, found 270.0 (M+l)⁺; Rétention time: 2.08 minutes; LCMS Method: Kinetex Cis 4.6 X 50 mm 2.6 μΜ. Temp: 45 °C, Flow: 2.0 mL/min, Run Time: 6 min. Mobile Phase: Initial 95% H2O (0.1% formic acid) and 5% CH3CN (0.1% formic acid) linear gradient to 95% CH3CN (0.1% formic acid) for 4.0 min then held at 95% CH3CN (0.1% formic acid) for 2.0 min.

Step 3: 6-Pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carboxylic acid

To a solution of pent-4-en-l-ol (1.5 mL, 14.79 mmol) in DMF (12 mL) was added sodium hydride (1.05 g of 60 % w/w, 26.25 mmol) and the reaction mixture was stirred at room température for 0.5 h. The mixture was cooled to 0 °C and 6-bromo-5-(trifluoromethyl)pyridine2-carboxylic acid (2 g, 7.407 mmol) in DMF (12 mL) was added. The reaction mixture was warmed to room température and stirred for another 2 h. The mixture was carefiilly quenched with water (2 mL). The solvent was removed in vacuo and the reaction mixture was diluted with MeOH and filtered. The filtrate was evaporated and purified by reverse phase chromatography on Cis column using a gradient elution of 20% - 70% water/acetonitrile and a flow rate of 80 mL/min over 20 min to afford 6-pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carboxylic acid (1.73 g, 85%). ESI-MS m/z cale. 275.07693, found 274.0 (M-l)⁺; Rétention time: 0.83 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column 192 made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes.

Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 4: 7V-[2-BenzyIoxy-2-(trifluoromethyl)pent-4-enoyl]-6-pent-4-enoxy-5(trifluoromethyl)pyridine-2-carbohydrazide

To a stirred solution of 6-pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carboxylic acid (427.6 mg, 1.540 mmol) and HATU (761.2 mg, 2.002 mmol) in DMF (5 mL) was added DIPEA (697.4 pL, 4.004 mmol) (exotherm). After 5 min, 2-benzyloxy-2-(trifluoromethyl)pent-4enehydrazide (hydrochloride sait) (500 mg, 1.540 mmol) was added in one portion and the mixture was stirred for 30 min. The reaction mixture was diluted with water (15 mL), extracted with ethyl acetate (3X15 mL) and the combined extracts were washed with brine then dried (MgSCU). The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (10% to 33%) to afford N-[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]-6pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carbohydrazide (725 mg, 86%) as colorless gum. 'H NMR (400 MHz, Chloroform-d) δ 9.71 (s, 1H), 9.16 (s, 1H), 7.96 (d, J= 7.7 Hz, 1H), 7.77 - 7.70 (m, 1H), 7.38 - 7.26 (m, 5H), 5.88 - 5.72 (m, 2H), 5.36 - 5.22 (m, 2H), 5.05 - 4.90 (m, 2H), 4.85 - 4.73 (m, 2H), 4.40 (t, 6.2 Hz, 2H), 3.11 (dd, J= 15.5, 5.9 Hz, 1H), 2.96 (dd, J= 15.5, 7.8 Hz,

1H), 2.25 - 2.13 (m, 2H), 1.87 (dt, J= 7.9, 6.3 Hz, 2H) ppm. ESI-MS m/z cale. 545.1749, found 546.21 (M+l)⁺; Rétention time: 1.12 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B ~ acetonitrile (with formic acid).

Step 5: 2-[l-Benzyloxy-l-(trifluoromethyl)but-3-enyI]-5-[6-pent-4-enoxy-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazoIe

To a degassed solution of JV’-[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]-6-pent-4enoxy-5-(trifluoromethyl)pyridine-2-carbohydrazide (100 mg, 0.1818 mmol) in THF (2 mL) was 193 added methoxycarbonyl-(triethylammonio)sulfonyl-azanide (130 mg, 0.5455 mmol) in one portion. The resulting solution was heated in a sealed vial at 80 °C for 2 hours, solvent was evaporated then diluted the residue with ethyl acetate (10 mL), washed with 2N NaOH solution and 0.5 N HCl, brine, then dried (MgSCU), filtered and concentrated. The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (5% to 26%, 8 column volumes) which afforded 2-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-5-[6-pent-4enoxy-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (76 mg, 79%) as an oil. ’H NMR (400 MHz, Chloroform-d) δ 8.07 - 8.03 (m, 1H), 7.84 (dd, J= 7.8, 0.8 Hz, 1H), 7.43 - 7.29 (m, 5H), 6.06 - 5.80 (m, 2H), 5.34 - 5.20 (m, 2H), 5.13 - 5.00 (m, 2H), 4.85 (d, J= 10.8 Hz, 1H), 4.65 (d, J= 10.9 Hz, 1H), 4.56 (t, J= 6.3 Hz, 2H), 3.30 - 3.21 (m, 2H), 2.33 - 2.22 (m, 2H), 2.02 - 1.91 (m, 2H) ppm. ESI-MS m/z cale. 527.16437, found 528.21 (M+l)⁺; Rétention time: 1.23 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 6: 6-BenzyIoxy-6,15-bis(trifIuoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaene (£7Z mixture)

To a degassed stirred solution of 2-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-5-[6pent-4-enoxy-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (265 mg, 0.5024 mmol) in DCE (21 mL) was added [l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]-dichloro-[(2isopropoxy-5-nitro-phenyl)methylene]ruthenium (68 mg, 0.1012 mmol), résultant mixture was purged with nitrogen and heated at 80 °C for 16 hours. The reaction mixture was concentrated and the residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (5% to 40%, 12 column volumes) which afforded 6-benzyloxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,8,14(18),15-hexaene (E/Zmixture) (35 mg, 14%) as an oil and followed by dimeric side-product (85 mg) as white solid. *H NMR (400 MHz, Chloroform-d) δ 8.06 (dd, J—Ί.Ί, 0.8 Hz, 1H), 7.86 - 7.79 (m, 1H), 7.28 (d, J= 2.4 Hz, 4H), 7.23 - 7.16 (m, 1 H), 6.13 - 5.99 (m, 1H), 5.90 - 5.77 (m, 1H), 4.91 (d, J= 11.6 Hz, 1H), 4.79 - 4.46 (m, 3H), 3.18 (dd, J= 14.6, 5.3 Hz, 1H), 2.81 (dd, J= 14.5, 8.9 Hz, 1H), 2.24 - 1.90 (m, 4H) ppm. ESI-MS m/z cale. 499.13306, found 500.2 (M+l)⁺; Rétention time: 1.17 minutes. Final purity was determined by reversed

194 phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 7: 6,15-bis(Trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol, Compound 1

To a solution of 6-benzyloxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaene (E/Zmixture) (30 mg, 0.06007 mmol) in MeOH (3 mL) was added Silica Cat Pd (70 mg of 0.2 mmol/g, 0.01400 mmol) and stirred for 16 hours under hydrogen balloon. The mixture was diluted with ethyl acetate, filtered through a pad of Celite eluting with ethyl acetate and DCM then concentrated. The residue was purified by silica gel chromatography eluted with a gradient of ethyl acetate in hexanes (5% to 40%, 15 column volumes) followed by lyophilization using acetonitrile and water which afforded racemic 6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (17.5 mg, 70%) as a white solid. 'H NMR (400 MHz, Chloroform-d) δ 8.08 (dd, J= 7.7, 0.8 Hz, 1H), 7.85 (dd, J= 7.7, 0.8 Hz, 1H), 4.69 - 4.60 (m, 1H), 4.54 - 4.42 (m, 1H), 3.94 (d, J= 1.3 Hz, 1H), 2.52 - 1.34 (m, 10H) ppm. ESI-MS m/z cale. 411.10175, found 411.54 (M+l)⁺; Rétention time: 3.7 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 5 - 85% mobile phase B over 6.0 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Example 2: Préparation of 6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricycio[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (enantiomer 1), Compound 2, and 6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18- triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (enantiomer 2), Compound 3

Step 1

enantiomer 1

N-N enantiomer 2

Step 1: 6,15-bis(Trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol (enantiomer 1), Compound 2, and 6,15195 bis(Trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol (enantiomer 2), Compound 3

N-N

enantiomer 1

enantiomer 2

Racemic 6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (9.5 mg, 0.02310 mmol) was separated by SFC using isocratic 3% methanol over 30 min on a Lux2 10 X 250 mm column to provide two single enantiomers. The first enantiomer to elute, 3.6 mg of 6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 17),2,4,14( 18), 15pentaen-6-ol (enantiomer 1), was enantiomerically pure however LCMS of this compound showed a polar impurity. This material was re-purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (5% to 40%, 15 column volumes) affording 6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(17),2,4,14(18),15pentaen-6-ol (enantiomer 1) (2.7 mg, 54%), chiral purity, > 99.9%. ESI-MS m/z cale.

411.10175, found 412.21 (M+l)⁺; Rétention time: 3.76 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 5 - 85% mobile phase B over 6.0 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid). The second enantiomer to elute from the SFC séparation was 6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (3.3 mg, 69%), chiral purity, >99.9%, ESI-MS m/z cale. 411.10175, found 412.2 (M+l)⁺; Rétention time: 0.96 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

196

Example 3: Préparation of 17-Amino-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (racemic), Compound 4

To methyl 3-amino-6-bromo-5-(trifluoromethyl)pyridine-2-carboxylate (approximately 2.5 g, 8.36 mmol) and CuBn (approximately 2.801 g, 12.54 mmol) in acetonitrile (60.90 mL) at room température was added /er/-butyl nitrite (approximately 1.293 g, 1.491 mL, 12.54 mmol) dropwise and the reaction was stirred at room température for 16 h in a closed atmosphère.

Complété consumption of the SM was observed. The reaction was diluted with saturated aqueous NH4CI and the aqueous layer was extracted with CH2CI2. The combined organic layers were dried over sodium sulfate and the solvent was removed under reduced pressure. The crude residue, methyl 3,6-dibromo-5-(trifluoromethyl)pyridine-2-carboxylate (1.6 g, 53%) was used in the next reaction without further purification.

197

Step 2: 3,6-Dibromo-5-(trifIuoromethyl)pyridine-2-carboxylic acid

To a solution of methyl 3,6-dibromo-5-(trifluoromethyl)pyridine-2-carboxylate (1 g, 2.755 mmol) in THF (10 mL) was added a solution of lithium hydroxide monohydrate (2 mL of 1.7 M, 3.400 mmol) in water (2.8 mL). This mixture was stirred room température for 15 hours, concentrated by rotary évaporation, acidified with aqueous 2 N HCl and extracted with methylene chloride (3X15 mL). The combined extracts were passed through a phase separator and concentrated to afford 3,6-dibromo-5-(trifluoromethyl)pyridine-2-carboxylic acid (920 mg, 93%) as a brown solid. *H NMR (400 MHz, Chloroform-d) δ 8.36 (s, 1H) ppm. ESI-MS m/z cale. 346.84042, found 347.91 (M+l)⁺; Rétention time: 0.51 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 3: 3-Bromo-6-pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carboxylic acid

To a stirred suspension of sodium hydride (310 mg of 60 % w/w, 7.751 mmol) in DMF (6 mL) was added pent-4-en-l-ol (550 pL, 5.421 mmol) in DMF (2 mL) stirred at room température for 0.5 h. The mixture was cooled to 0 °C, 3,6-dibromo-5-(trifluoromethyl)pyridine2-carboxylic acid (920 mg, 2.566 mmol) in DMF (6 mL) was added, the reaction mixture was warmed to room température and stirred for a further 2 h. The mixture was carefully quenched with water (10 mL). Aqueous basic solution was washed with hexanes and ether (1:1), acidified with aqueous 2 N HCl, extracted with ether (3 X 20 mL) and combined extracts were washed with brine, dried (MgSCU) and concentrated. Purification by column chromatography afforded still impure 3-bromo-6-pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carboxylic acid (600 mg, 66%) as an oil which was used as such in the next step without further purification. ESI-MS m/z cale. 352.98743, found 354.01 (M+l)⁺; Rétention time: 0.86 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

198

Step 4:7V-[2-Benzyloxy-2-(trifIuoromethyl)pent-4-enoyl]-3-bromo-6-pent-4-enoxy-5(trifluoromethyl)pyridine-2-carbohydrazide

To a stirred solution of 3-bromo-6-pent-4-enoxy-5-(trifluoromethyl)pyridine-2carboxylic acid (640 mg, 1.807 mmol) and HATU (885 mg, 2.328 mmol) in DMF (6 mL) was added DIPEA (1.1 mL, 6.315 mmol) (exotherm). After 5 min, 2-benzyloxy-2(trifluoromethyl)pent-4-enehydrazide (hydrochloride sait) (600 mg, 1.791 mmol) was added in one portion and the mixture was stirred for 16 h. The reaction mixture was diluted with water (25 mL), extracted with ethyl acetate (3 X 25 mL) and combined extracts were washed with brine and dried (MgSCh) then filtered and concentrated. The residue was purifïed by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (5% to 29%, 10 column volumes) to afford 7V-[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]-3-bromo-6-pent-4-enoxy5-(trifluoromethyl)pyridine-2-carbohydrazide (340 mg, 26%) as colorless gum. *H NMR (400 MHz, Chloroform-d) δ 9.81 (s, 1H), 9.31 (s, 1H), 8.14 (s, 1H), 7.40 - 7.35 (m, 5H), 5.88 - 5.75 (m, 2H), 5.38 - 5.26 (m, 2H), 5.07 - 4.95 (m, 2H), 4.83 (s, 2H), 4.43 (t, J= 5.9 Hz, 2H), 3.20 2.97 (m, 2H), 2.28 - 2.19 (m, 2H), 1.96 - 1.87 (m, 2H) ppm. ESI-MS m/z cale. 623.08545, found 624.13 (M+l)⁺; Rétention time: 1.1 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 5: 2-[l-BenzyIoxy-l-(trifluoromethyl)but-3-enyl]-5-[3-bromo-6-pent-4-enoxy-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole

To a degassed solution of 7V-[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]-3-bromo-6pent-4-enoxy-5-(trifluoromethyl)pyridine-2-carbohydrazide (340 mg, 0.4609 mmol) in THF (6 mL) was added methoxycarbonyl-(triethylammonio)sulfonyl-azanide (330 mg, 1.385 mmol) in one portion. Résultant solution was heated in a sealed vial at 80 °C for 2 hours. The mixture was

199 quenched with 2 N NaOH (~1.5 mL) and water (5 mL), most of the solvent was evaporated then diluted with ethyl acetate (20 mL). The organic solution was washed with 0.5 N HCl, brine, dried (MgSCU), filtered and concentrated. The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (0% to 12%, 12 column volumes) affording 2[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-5-[3-bromo-6-pent-4-enoxy-5-(trifluoromethyl)-2pyridyl]-l,3,4-oxadiazole (240 mg, 86%) as an oil. 'H NMR (400 MHz, Chloroform-d) δ 8.17 (s, 1H), 7.39 - 7.27 (m, 5H), 6.00 - 5.88 (m, 1H), 5.86 - 5.75 (m, 1H), 5.29 - 5.21 (m, 1H), 5.21 5.16 (m, 1H), 5.06 - 4.95 (m, 2H), 4.84 (d, J= 10.8 Hz, 1H), 4.63 (d, J= 10.8 Hz, 1H), 4.44 (t, J= 6.3 Hz, 2H), 3.20 (t, J= 6.0 Hz, 2H), 2.20 (q, J= 7.2 Hz, 2H), 1.93 - 1.84 (m, 2H) ppm. ESI-MS m/z cale. 605.0749, found 607.12 (M+l)⁺; Rétention time: 1.22 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 6: 6-Benzyloxy-17-bromo-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaene (E/Z mixture)

To a degassed stirred solution of 2-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-5-[3bromo-6-pent-4-enoxy-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (247 mg, 0.4074 mmol) in DCE (20 mL) was added [l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]-dichloro-[(2isopropoxy-5-nitro-phenyl)methylene]ruthenium (50 mg, 0.07445 mmol) and the resulting mixture was purged with nitrogen and heated at 80 °C for 30 min. SiliaMetS (150 mg) was added, stirred for 30 min, filtered and rinsed with DCM then concentrated. The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (0% to 20%, 15 column volumes) to afford 6-benzyloxy-17-bromo-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 17),2,4,8,14( 18), 15-hexaene (E/Z mixture) (100 mg, 42%) as a white solid. 'H NMR (400 MHz, Chloroform-d) δ 8.17 (brs, 1H), 7.25 7.24 (m, 2H), 7.24 - 7.23 (m, 2H), 7.18 - 7.12 (m, 1H), 5.88 - 5.80 (m, 2H), 4.88 (d, J= 11.6 Hz, 1H), 4.69 - 4.57 (m, 1H), 4.53 (d, J= 11.6 Hz, 1H), 4.50 - 4.41 (m, 1 H), 3.12 (dd, 14.4, 4.3

Hz, 1H), 2.72 (dd, J= 14.6, 8.0 Hz, 1H), 2.19 - 1.83 (m, 4H) ppm. ESI-MS m/z cale. 577.0436, found 579.97 (M+l)⁺; Rétention time: 1.16 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run

200 from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid).

Mobile phase B = acetonitrile (with formic acid).

Step 7: 6-Benzyloxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaen-17-amme (ΕΓΖ, mixture)

To a degassed mixture of 6-benzyloxy-17-bromo-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaene (E/Zmixture) (35 mg, 0.06052 mmol), Xantphos (4 mg, 0.006913 mmol), diphenylmethanimine (14 pL) and césium carbonate (40 mg, 0.1228 mmol) in dioxane (700 pL) was added Pd(OAc)z (1.8 mg, 0.008017 mmol). Degassed using vacuum/nitrogen and heated in a sealed vial at 100 °C for 2.5 hours, cooled to room température, diluted with water (2 mL), extracted with ethyl acetate (4X5 mL) and combined extracts were dried (MgSCh), filtered and concentrated to afford the intermediate 7V-[6-benzyloxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaen-17-yl]-l,l-diphenylmethanimine intermediate as a yellow liquid. ESI-MS m/z cale. 678.20654, found 678.38 (M+l)⁺; Rétention time: 2.16 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 50 100% mobile phase B over 3.0 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid). To a stirred solution of the crude 7V-[6-benzyloxy6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,8,14(18),15-hexaen-17-yl]-l,l-diphenyl-methanimine (E/Zmixture) (60 mg, 129%) in THF (2 mL) was added HCl (2 mL of 2 M, 4.000 mmol) at room température, stirred for 10 min, concentrated by rotary évaporation, diluted with water (1 mL), extracted with methylene chloride (3X5 mL) and the combined extracts were dried (MgSCU), filtered and concentrated. The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (10% to 40%, 9 column volumes) which afforded 6-benzyloxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,8,14(18),15-hexaen-17-amine (E/Zmixture) (24 mg, 77%) as a brown solid. *H NMR (400 MHz, Chloroform-d) δ 7.40 (brs, 1H), 7.27 - 7.22 (m, 4H), 7.20 - 7.14 (m, 1H), 6.09 - 5.98 (m, 1H), 5.78 (dt, J= 14.9, 7.0 Hz, 1H), 5.32 (s, 2H), 4.86 (d, J= 11.5 Hz, 1H), 4.56 (d, J= 11.5 Hz, 1H), 4.53 - 4.32 (m, 2H), 3.14 (dd, J= 14.6, 5.4 Hz, 1H), 2.78 (dd, J= 14.5, 8.7 Hz, 1H), 2.18

201

- 1.84 (m, 4H) ppm. ESI-MS m/z cale. 514.144, found 515.15 (M+l)⁺; Rétention time: 1.14 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 15 - 98% mobile phase B over 1.5 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

Step 8: 17-Amino-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 4

To a solution of 6-benzyloxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaen-17-amine (E/Zmixture) (22 mg, 0.04277 mmol) in MeOH (3 mL) was added SiliaCat Pd (49 mg, 0.1944 mmol), stirred for 2 hours under hydrogen balloon then added an additional amount of SiliaCat Pd (20 mg of 0.2 mmol/g, 0.004000 mmol). Stirred for 2 hours and then the mixture was heated at 50 °C for 2 hours. The reaction was diluted with ethyl acetate, filtered through a pad of Celite eluting with ethyl acetate then concentrated. The residue was purified by silica gel chromatography eluting with a gradient of ethyl acetate in hexanes (10% to 50%, 15 column volumes) followed by lyophilization using acetonitrile and water which afforded 17-amino-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (14.5 mg, 77%) as a light yellow solid. ’H NMR (400 MHz, Chloroform-d) δ 7.36 (s, 1H), 5.18 (brs, 2H), 4.44 _ 4.24 (m, 2H), 3.48 (brs, 1H), 2.35 - 2.11 (m, 2H), 2.09 - 1.94 (m, 1H), 1.92 - 1.69 (m, 1H), 1.67 - 1.33 (m, 6H) ppm. ESI-MS m/z cale. 426.11267, found 427.29 (M+l)⁺; Rétention time: 3.83 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC HSS T3 column made by Waters, and a dual gradient run from 5 - 85% mobile phase B over 6.0 minutes. Mobile phase A = water (with formic acid). Mobile phase B = acetonitrile (with formic acid).

202

Example 4: Préparation of (61î)-17-aniino-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI, Compound 5

Step 1

Step 2

Step 1: tfrf-Butyl 7V-[2-[5-[(lR)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol 5 2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyI]carbamate

To a stirring solution of /eri-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (10 g, 15.69 mmol) in DMSO (64.97 mL) at room température was added césium acetate (3.012 g, 15.69 mmol) and the mixture was capped and heated under nitrogen atmosphère to 80 °C and stirred for 160 min. Reaction was stopped as it was progressing to the undesired 7V-acetyl product. Diluted the reaction mixture with water and extracted with EtOAc. Washed the organic layer with saturated aqueous NaHCCh (IX), saturated aqueous NH4CI (IX) and brine (IX), then dried (MgSCU), filtered and concentrated to a residue which was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc. Mixed fractions were combined, concentrated and purified by Cis reverse phase chromatography using a gradient run from 50% 99% mobile phase B over 15.0 minutes (mobile phase A = H2O (5 mM HCl), mobile phase B = acetonitrile. Isolated ier/-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4203 oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]carbamate (297.7 mg, 3%) as a minor product which was used directly in the ensuing step.

Step 2: ZerZ-Butyl 7V-[2-[5-[(l.R)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol 2-yI]-6-pent-4-enoxy-5-(trifluoromethyI)-3-pyridyl]carbamate

To a solution of Zeri-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]carbamate (295 mg, 0.5135 mmol) and pent-4-en-l-ol (78.14 pL, 0.7702 mmol) in toluene (6.21 mL) was added triphenylphosphine (178.4 pL, 0.7700 mmol). After stirring at room température for 1 min, DIAD (161.8 pL, 0.8218 mmol) was added and the mixture was stirred at room température for 5 minutes. Diluted the reaction mixture with EtOAc then washed with saturated aqueous NaHCCh (IX), saturated aqueous NH4CI (IX) and brine (IX) then dried over MgSO4, filtered and concentrated to a yellow oil which was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc giving as a clear, slightly yellow syrup, terZ-butyl7V-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6pent-4-enoxy-5-(trifluoromethyl)-3-pyridyl]carbamate (271.9 mg, 82%). ESI-MS m/z cale. 642.22766, found 643.3 (M+l)⁺; Rétention time: 0.83 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 ^x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60

c.

Step 3: tert-Butyl A-[(6Æ)-6-benzyIoxy-6,15-bis(trifluoroniethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaen-17-yl]carbamate (L7Z mixture)

To a two neck flask bubbling in nitrogen added ZerZ-butyl 7V-[2-[5-[(lÀ)-l-benzyloxy-l(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-pent-4-enoxy-5-(trifluoromethyl)-3204 pyridyl]carbamate (271.9 mg, 0.4231 mmol) in DCE (62.54 mL) and heated to 60 °C. Then added via syringe, Zhan catalyst-lB (77.63 mg, 0.1058 mmol) in dichloroethane (1 mL), heated reaction to 60 °C and stirred while bubbling nitrogen through the solution. Added dichloroethane intermittently as the reaction progressed to maintain volume. After 80 min, added Zhan catalyst-lB (46.56 mg, 0.06345 mmol) and continued stirring at 60 °C for 160 min. Allowed the reaction mixture to cool to room température then added 2-sulfanylpyridine-3carboxylic acid (26.26 mg, 0.1692 mmol) and stirred for 5 min. Concentrated the reaction mixture by rotary évaporation then purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc giving as a yellow solid, ZerZ-butyl N-[(6R)-6benzyloxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,8,14(18),15-hexaen-17-yl]carbamate (E/Zmixture) (169 mg, 65%). ESI-MS m/z cale. 614.1964, found 615.2 (M+l)⁺; Rétention time: 0.75 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60

Step 4: ZtvZ-Butyl A^r-[(6R)-6-hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate

To a solution of terributyl 7V-[(6Æ)-6-benzyloxy-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,8,14(18),15-hexaen-17-yl]carbamate (E/Z mixture) (167 mg, 0.2717 mmol) in AcOH (5.566 mL) was added Pd/C (88.83 mg of 10 % w/w, 0.08347 mmol) and hydrogen gas was bubbled through the stirring mixture for 15 minutes then the reaction was sealed and capped with a hydrogen balloon and stirred for 2.5 h. Added palladium (28.91 mg of 10 % w/w, 0.02717 mmol), stirred for 1 h then purged the flask with nitrogen and filtered over Celite eluting with EtOAc. The filtrate was concentrated then purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc giving as a white foam, ZerZ-butyl 7V-[(67?)-6-hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (143 mg, 100%). ESI-MS m/z cale. 526.1651, found 527.2 (M+l)⁺; Rétention time: 0.52 minutes.

205

Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 5: (67?)-17-Amino-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 5

To a stirring solution of ier/-butyl 7V-[(67?)-6-hydroxy-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (143 mg, 0.2716 mmol) in DCM (1.43 mL) was added TFA (522.9 pL, 6.787 mmol) and the resulting mixture was stirred at room température for 2 h then concentrated by rotary évaporation to a yellow residue which was dissolved in EtOAc and washed with saturated aqueous NaHCCfi (IX), dried (MgSCh), filtered and concentrated to a pale yellow residue. This material was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc giving as a pale yellow solid, (67î)-17-amino-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (88.7 mg, 76%). 'H NMR (400 MHz, DMSO) δ 7.80 - 7.76 (m, 1H), 7.55 (s, 1H), 6.37 (s, 2H), 4.44 - 4.27 (m, 2H), 2.20 (q, J= 7.2 Hz, 1H), 2.10 (dd, J = 14.7, 7.1 Hz, 1H), 2.06 - 1.96 (m, 1H), 1.80 (dd, J= 11.3, 5.8 Hz, 1H), 1.69 - 1.53 (m, 4H), 1.42 (d, J= 7.2 Hz, 2H) ppm. ESI-MS m/z cale. 426.11267, found 427.4 (M+l)⁺; Rétention time: 1.86 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cie column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Example 5: Préparation of (6S)-17-amino-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI, Compound 6

Step 1

enantiomer 2

206

Step 1 : (6S)-17-Amino-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 6

nh₂ n-n enantiomer 2

Racemic 17-amino-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (32.6 mg, 0.07647 mmol) was separated by préparative SFC using a LUX-4 (25 cm X 2.1 cm, 5 μΜ) column using methanol as solvent to give as the second enantiomer to elute, (65)-17-amino-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (11.8 mg, 72%). Ή NMR (400 MHz, DMSO-d₆) δ 7.78 (s, 1H), 7.61 (s, 1H), 6.39 (s, 2H), 4.35 (s, 2H), 2.18 (s, 1H), 2.08 (d, J= 39.0 Hz, 2H), 1.82 (s, 1H), 1.64 (s, 4H), 1.42 (s, 2H) ppm. ESI-MS m/z cale. 426.11267, found 427.0 (M+l)⁺; Rétention time: 1.86 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 2: Solid form chracterization of crystalline Compound 6 (neat form)

Single crystals of crystalline Compound 6 (neat form) were grown by vapor diffusion of pentane into a solution of Compound 6 in 1,2-dicholorethane. X-ray diffraction data were acquired at 100 K on a Broker diffractometer equipped with Cu Ka radiation (λ=1.5478 Â) and a CCD detector. The structure was solved and refined using SHELX programs (Sheldrick, G.M., Acta Cryst., (2008) A64, 112-122). The results are summarized in Table 3 below.

Table 3: Single crystal élucidation of crystalline Compound 6 (neat form)

Crystal System	Monoclinic
Space Group	P2i
a(Â)	9.5564(4)
b(Â)	13.5953(5)
c(Â)	13.8474(5)
a(°)	90
β(°)	105.3070(10)
y(°)	90
V(Â³)	1735.26(12)
Z/Z'	4/1

207

Température 100 K

Example 6: Préparation of 17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (diastereomer pair 1), Compound 7, and 17-amino-12-methyl-6,15-bis(trifluoromethyl)-

13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (diastereomer pair 2), Compound 8

Step 1

Step 2

Step 3

Step 1: Methyl 3-chloro-l-oxido-5-(trifluoromethyl)pyridin-l-ium-2-carboxylate

To a solution of methyl 3-chloro-5-(trifluoromethyl)pyridine-2-carboxylate (25 g, 102.26 mmol) in dichloromethane (250 mL) cooled to 0 °C was added urea hydrogen peroxide (34 g, 361.43 mmol) followed by the slow addition of trifluoroacetic anhydride (72.528 g, 48 mL, 208

345.32 mmol). The reaction mixture was stirred at room température for 3 h. The reaction mixture was then poured into ice-water (200 mL) and adjusted to pH = 7 - 8 with 25 % aqueous sodium hydroxide solution. The mixture was diluted with dichloromethane (100 mL) and then the layers were separated. The aqueous phase was extracted with dichloromethane (2 X 100 mL). The combined organic phases were washed with brine (250 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to afford methyl 3-chloro-loxido-5-(trifluoromethyl)pyridin-l-ium-2-carboxylate (24.5 g, 94%) as a white solid which was used directly in the ensuing step. ’H NMR (300 MHz, DMSO-dô) δ 9.00 (s, 1H), 8.25 (s, 1H), 3.97 (s, 3H) ppm. ¹⁹F NMR (282 MHz, DMSO-d₆) δ -61.75 (br. s., 3F) ppm. ESI-MS m/z cale. 254.99101, found 256.0 (M+l)⁺; Rétention time: 1.66 minutes; LCMS Method: Kinetex Polar Ci8 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in water (0.1% formic acid) 1.2 mL/min.

Step 2: Methyl 3,6-dichloro-5-(trifluoromethyl)pyridine-2-carboxylate ci

Methyl 3-chloro-l-oxido-5-(trifluoromethyl)pyridin-l-ium-2-carboxylate (18.52 g, 72.463 mmol) was added in portions to phosphoryl trichloride (121.73 g, 74 mL, 793.90 mmol) at 0 °C and the resulting mixture was stirred at 50 °C ovemight. Removal of the solvent in vacuo gave a black oil which was dissolved in ethyl acetate (200 mL) and carefully neutralized with a saturated aqueous solution of sodium carbonate until pH ~ 8. The mixture was extracted with ethyl acetate (2 X 500 mL) and the combined organic phases were washed with brine (250 mL), dried over anhydrous sodium sulfate, filtered and concentrated. The black oil was purified by flash chromatography on silica gel, eluting with a gradient from 0% to 20% ethyl acetate in heptanes to afford methyl 3,6-dichloro-5-(trifluoromethyl)pyridine-2-carboxylate (16.43 g, 83%) as a light yellow oil. *H NMR (300 MHz, CDCI3) δ 8.13 (s, 1H), 4.04 (d, J= 1.8 Hz, 3H) ppm. ¹⁹F NMR (282 MHz, CDCI3) ppm -64.2 (s, 3F) ppm. ESI-MS m/z cale. 272.95712, found 274.0 (M+l)⁺; Rétention time: 2.02 minutes; LCMS Method: Kinetex Cis 4.6 X 50 mm 2.6 pM. Temp: 45 °C, Flow: 2.0 mL/min, Run Time: 3 min. Mobile Phase: Initial 95% H2O (0.1% formic acid) and 5% acetonitrile (0.1% formic acid) linear gradient to 95% acetonitrile (0.1% formic acid) for 2.0 min then held at 95% acetonitrile (0.1% formic acid) for 1.0 min.

209

Step 3: 3,6-Dichloro-5-(trifluoromethyl)pyridine-2-carboxyIic acid

A mixture of methyl 3,6-dichloro-5-(trifluoromethyl)pyridine-2-carboxylate (14.63 g, 52.428 mmol) in THF (150 mL) and water (150 mL) was treated with lithium hydroxide monohydrate (4.5 g, 107.24 mmol) added portion-wise and the mixture was stirred vigorously at room température for 1.5 h. The crude reaction mixture was transferred to a 2 L separatory funnel with 5% citric acid (400 mL) and ethyl acetate (800 mL) and the layers were separated. The aqueous phase was extracted further with ethyl acetate (2 X 200 mL). The combined organic layers were then washed with water (120 mL), brine (2 X 120 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to afford 3,6-dichloro-5(trifluoromethyl)pyridine-2-carboxylic acid (12.58 g, 92%) as a pale pink solid which was used directly in the ensuing step. *H NMR (300 MHz, DMSO-dô) δ 14.64 (br. s, 1H), 8.68 (s, 1H) ppm. ¹⁹F NMR (282 MHz, DMSO-d₆) δ -62.62 (s, 3F) ppm. ESI-MS m/z cale. 258.94147, found 257.9 (M+l)⁺; Rétention time: 1.51 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in water (0.1% formic acid) 1.2 mL/min.

Step 4: 7V'-[2-Benzyloxy-2-(trifluoromethyl)hex-5-enoyI]-3,6-dichloro-5(trifIuoromethyl)pyridine-2-carbohydrazide

To a solution of 3,6-dichloro-5-(trifluoromethyl)pyridine-2-carboxylic acid (8.5 g, 32.693 mmol) in DMF (80 mL) was added triethylamine (10.890 g, 15 mL, 107.62 mmol) and HATU (15 g, 39.450 mmol). The mixture was stirred for 10 min, then 2-benzyloxy-2(trifluoromethyl)hex-5-enehydrazide (hydrochloride sait) (11.51 g, 33.978 mmol) was added. The mixture was stirred at room température for 18 h, then poured into ice-cold water (200 g) and extracted with ethyl acetate (2 X 220 mL). The combined organic layers were washed with aqueous saturated sodium bicarbonate solution (2 X 80 mL), water (1 X 80 mL) and brine (2 X 80 mL). The organic layer was concentrated by évaporation under reduced pressure to give a dark yellow oil residue (19.2 g) that was combined with dichloromethane (100 mL) and preadsorbed on silica gel. Flash chromatography on silica gel (220 g column, crude dry loaded on 50 g of silica gel) using a gradient from 0% to 20% EtOAc in heptanes afforded V-[2

210 benzyloxy-2-(trifluoromethyl)hex-5-enoyl]-3,6-dichloro-5-(trifluoromethyl)pyridine-2carbohydrazide (11.37 g, 60%) as a white solid. Ή NMR (300 MHz, CDC1₃) δ 9.18 (br. s, 1H), 8.17 (s, 1H), 7.49 - 7.32 (m, 5H), 5.95 - 5.75 (m, 1H), 5.18 - 5.01 (m, 2H), 4.86 (d, J= 10.3 Hz, 1H), 4.73 (d, J= 10.6 Hz, 1H), 2.55 - 2.38 (m, 1H), 2.36 - 2.16 (m, 3H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -64.15 (s, 3F), -73.62 (s, 3F) ppm. ESI-MS m/z cale. 543.0551, found 544.1 (M+l)⁺; Rétention time: 2.25 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in water (0.1% formic acid) 1.2 mL/min.

Step 5: 2-[l-Benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3,6-dichloro-5-(trifluoromethyl)2-pyridyl]-l,3,4-oxadiazole

To a solution of?/-[2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]-3,6-dichloro-5(trifluoromethyl)pyridine-2-carbohydrazide (10.37 g, 17.891 mmol) andTV.TVdiisopropylethylamine (6.5378 g, 8.9 mL, 50.079 mmol) in acetonitrile (240 mL) at 50 °C was added /7-toluenesulfonyl chloride (4.15 g, 21.768 mmol) portion-wise. The mixture was stirred at 70 °C. Upon completion (1 h), the reaction mixture was concentrated. The residue was dissolved in dichloromethane and washed with 5% aqueous sodium bicarbonate (20 mL), dried with anhydrous sodium sulfate. Flash chromatography on silica gel (40 g column, gradient from 0% to 15% EtOAc in heptanes) afforded 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3,6dichloro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (8.1 g) as a white solid, containing residual p-toluenesulfonyl chloride. A 1.9 g fraction was dissolved in dichloromethane (40 mL) and added ammonium hydroxide (2.5 mL, 28 - 30% NH3 basis) under stirring. The mixture was stirred at room température for 1 h, then transferred to a separatory tunnel with ethyl acetate (160 mL) and separated. The organic layer was further washed with water (2X30 mL) and brine (30 mL), dried over anhydrous sodium sulfate and filtered. The volatiles of the fïltrate were removed by évaporation under reduced pressure. The residue was dry loaded on silica gel (50 g) and purified by silica gel chromatography (80 g column) using a gradient from 0% to 10% EtOAc in heptanes giving 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3,6-dichloro-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (1.7 g, 18%) as a white solid. The remaining 6.2 g fraction from the initial silica gel column was dissolved in dichloromethane (60 mL) and added ammonium hydroxide (5.0 mL, 28.0-30.0% NH3 basis) under stirring. The mixture was stirred at room température for 1 h, then transferred to a separatory funnel with dichloromethane (100

211 mL) and separated. The organic layer was washed with water (2X30 mL) and brine (30 mL), dried over anhydrous sodium sulfate then fïltered. The volatiles of the filtrate were removed by évaporation under reduced pressure. The residue was dry loaded on silica gel (50 g) and purified by silica gel chromatography on a 120 g column using a gradient from 0% to 10% ethyl acetate in heptanes giving 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3,6-dichloro-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (5.87 g, 62%) as a white solid, calculated overall yield of product from both columns was 80%. *H NMR (300 MHz, CDCI3) δ 8.25 (s, 1H), 7.53 7.28 (m, 5H), 5.87 - 5.68 (m, 1H), 5.13 - 4.94 (m, 2H), 4.85 (d, J= 10.6 Hz, 1H), 4.66 (d, J= 10.9 Hz, 1H), 2.61 - 2.15 (m, 4H) ppm. ¹⁹F NMR (282 MHz, CDCI3) δ -64.11 (s, 3F), -72.85 (s, 3F) ppm. ESI-MS m/z cale. 525.04456, found 526.1 (M+l)⁺; Rétention time: 2.42 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in water (0.1% formic acid) 1.2 mL/min.

Step 6: 2-[l-Benzyloxy-l-(trifluoromethyl)pent-4-enyI]-5-[3-chloro-6-(l-methyIbut-3 enoxy)-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole

To a solution of 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3,6-dichloro-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (1 g, 1.900 mmol) in DMSO (10 mL) was added pent-4-en-2-ol (502 mg, 5.828 mmol), CS2CO3 (3.2 g, 9.821 mmol) and iodocopper (215 mg, 1.129 mmol) then the mixture was heated at 60 °C for 6 h. The reaction mixture was poured onto crushed ice and the résultant pasty material was dissolved in ethyl acetate, washed with brine, dried over anhydrous sodium sulphate, fïltered and concentrated. The résultant brown residue was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 50% EtOAc in hexanes to afford 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3-chloro6-(l-methylbut-3-enoxy)-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (427 mg, 39%). ESIMS m/z cale. 575.14105, found 567.1 (M+l)⁺; Rétention time: 0.79 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

212

Step 7: tert-Butyl N-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2yl]-6-(l-methylbut-3-enoxy)-5-(trifluoromethyl)-3-pyridyl]carbamate

To a nitrogen degassed solution of2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[3chloro-6-(l-methylbut-3-enoxy)-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (425 mg, 0.7379 mmol) in dioxane (5 mL), was added teri-butyl carbamate (262 mg, 2.237 mmol), XPhos Pd G3 (8.6 mg, 0.01016 mmol), palladium (II) acetate (4.2 mg, 0.01871 mmol) and CS2CO3 (375 mg, 1.151 mmol) and heated the mixture in a sealed vial at 100 °C ovemight. Cooled to room température, diluted with water (8 mL), extracted with ethyl acetate (3 X 70 mL) then combined extracts were washed with brine (15 mL), dried (sodium sulfate), filtered and concentrated. The résultant brown residue was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 100% EtOAc to afford Zer/-butyl 7V-[2-[5-[l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-1,3,4-oxadiazol-2-yl]-6-( 1 -methylbut-3 -enoxy)-5(trifluoromethyl)-3-pyridyl]carbamate (152 mg, 31%) as a mixture of diastereomers. ESI-MS m/z cale. 656.24335, found 657.4 (M+l)⁺; Rétention time: 0.78 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 8: terf-Butyl 7V-[6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (E7Z mixture)

To a degassed solution of Zeri-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4enyl]-l,3,4-oxadiazol-2-yl]-6-(l-methylbut-3-enoxy)-5-(trifIuoromethyl)-3-pyridyl]carbamate (150 mg, 0.2284 mmol) in DCE (50 mL) was added dichloro[l,3-bis(2,4,6-trimethylphenyl)-2imidazolidinylidene][[5-[(dimethylamino)sulfonyl]-2-(l-methylethoxy-O)phenyl]methylene213

C]ruthenium(II) (Zhan catalyst-lB, 50 mg, 0.06221 mmol) and the reaction was heated at 70 °C ovemight while continuously bubbling nitrogen into the solution with a gas outlet. The reaction mixture was then cooled to room température and concentrated under reduced pressure. Diluted with 1:4 EtOAc/hexanes and filtered through Celite. The filtrate was concentrated and the résultant brown residue was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 30% EtOAc in hexanes to afford teri-butyl 7V-[6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16hexaen-17-yl]carbamate (E/Z mixture) (52 mg, 36%) as a yellow oil. ESI-MS m/z cale. 628.21204, found 629.3 (M+l)⁺; Rétention time: 0.73 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 9: terf-Butyl 7V-[6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate

Combined teri-butyl JV-[6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (E/Z mixture) (52 mg, 0.08273 mmol), Pd/C (47 mg of 10 % w/w, 0.04416 mmol), and AcOH (1 mL) in a Parr pressure vessel and sealed. Subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. Filled the vessel with hydrogen gas at 150 psi then stirred the mixture for 15 h. Subjected to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated and purified by reverse phase HPLC-MS using a gradient from 30% to 99% acetonitrile in water (+ 5 mM HCl) over 15.0 minutes to afford teri-butyl 7V-[6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16-pentaen-17-yl] carbamate (12.7 mg, 28%) as light brown solid. ESI-MS m/z cale. 540.1807, found 541.2 (M+l)⁺; Rétention time: 2.07 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % 214

CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 10:17-Amino-12-methyI-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (diastereomer pair 1), Compound 7, and 17-amino-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (diastereomer pair 2), Compound 8

-cf₃OH

-CF₃OH

Ck _NH diastereomer pair 1 diastereomer pair 2

To a solution of ZerZ-butyl 7V-[6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (12 mg, 0.02220 mmol) was added TFA (100 pL, 1.298 mmol) and dichloromethane (300 pL) (pre made solution of 1:4 TFA/dichloromethane) and the reaction was stirred at room température for about 1 h. Solvents were removed and dissolved in DMSO (1 mL) and the residue was purified by reverse phase HPLC-MS using a gradient from 1% to 99% acetonitrile in water (+ 5 mM HCl) over 15.0 minutes to afford as a light brown solid and the first eluting diastereomer pair, 17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (diastereomer pair 1) (2.1 mg, 38%). ESI-MS m/z cale. 440.1283, found 441.16 (M+l)⁺ ; Rétention time: 1.46 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C. The second eluting diastereomer pair, isolated as a light brown solid, was 17-amino-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (hydrochloride sait) (diastereomer pair 2) (3.6 mg, 67%). 'H NMR (500 MHz, DMSO-dô) δ 7.76 (s, 1H), 7.63 (s, 1H), 6.36 (s, 2H), 4.77 - 4.65 (m, 1H), 2.55 (dd, J= 8.8, 4.4 Hz, 1H), 2.17 (t, J= 12.2 Hz, 1H), 2.09 (ddd, J= 14.2, 10.5, 6.9 Hz, 1H), 1.68 (s, 1H), 1.59 (d, J = 7.9 Hz, 2H), 1.48 (d, J= 6.6 Hz, 3H), 1.33 (d, J= 6.3 Hz, 3H), 1.14 (q, J = 9.0, 8.0 Hz, 1H) ppm. ESI-MS m/z cale. 440.1283, found 441.2 (M+l)⁺; Rétention time: 1.51 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x

215

2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Example 7: Préparation of (6Æ)-17-amino-13,13-dioxo-6,15-bis(trifluoromethyl)-19-oxa13I⁶-thia-3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 9, and (6S)-17-amino-13,13-dioxo-6,15-bis(trifIuoromethyl)-19oxa-13X⁶-thia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 10

Enantiomer 1

Enantiomer 2

Step 1: ZerZ-Butyl A-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyI]-l,3,4-oxadiazol-2 yl]-6-but-3-enylsulfonyl-5-(trifluoromethyl)-3-pyridyl]carbamate

A mixture of ZerZ-butyl 2V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,415 oxadiazol-2-yl]-6-bromo-5-(trifIuoromethyl)-3-pyridyl]carbamate (545 mg, 0.84 mmol), but-3ene-l-sulfinate (sodium sait) (351 mg, 2.47 mmol), and Cul (472 mg, 2.48 mmol) in DMSO (5

216 mL) was heated at 100 °C for 3 h, then diluted with ether and water, the mixture filtered, the layers partitioned and the organic layer washed with water, brine, dried (MgSCh) and evaporated. The residue was purified by silica gel chromatography (24 g S1O2, 0 - 20% EtOAc in hexanes over 15 min) to provide ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4enyl]-l,3,4-oxadiazol-2-yl]-6-but-3-enylsulfonyl-5-(trifluoromethyl)-3-pyridyl]carbamate (327 mg, 57%). ’H NMR (400 MHz, Chloroform-d) δ 10.36 (s, 1H), 9.57 (s, 1H), 7.42 - 7.28 (m, 5H), 5.75 (tdt, J= 17.0, 10.2, 6.5 Hz, 2H), 5.06 (dt, J= 17.1, 1.4 Hz, 2H), 5.01 (d, J= 10.2 Hz, 2H), 4.84 (d, J= 10.9 Hz, 1H), 4.66 (d, J= 10.8 Hz, 1H), 3.69 (hept, J= 7.0 Hz, 2H), 2.65 - 2.58 (m, 2H), 2.56 - 2.31 (m, 2H), 2.30 - 2.18 (m, 1H), 1.59 (s, 9H), 1.55 - 1.51 (m, lH)ppm. ¹⁹F NMR (376 MHz, Chloroform-d) δ -58.48, -72.86 ppm. ESI-MS m/z cale. 690.1947, found 691.2 (M+l)⁺; Rétention time: 0.89 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: tert-Butyl 7V-[6-(benzyloxy)-13,13-dioxo-6,15-bis(trifhioromethyl)-19-oxa-13Z⁶-thia3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (E!Z mixture)

A solution of Ze/7-butyl jV-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-but-3-enylsulfonyl-5-(trifluoromethyl)-3-pyridyl]carbamate (300 mg, 0.4344 mmol) in 5 mL DCE was added dropwise over 5 min to a solution ofbenzylidene-[l,3-bis(2,4,6trimethylphenyl)imidazolidin-2-ylidene]-dichloro-ruthenium;tricyclohexylphosphane (55 mg, 0.06478 mmol) in DCE (42 mL) heated at 70 °C with constant N₂ bubbling for 1 h. The solvent was evaporated and the residue purified by silica gel chromatography (24 g S1O2, 0 - 20% EtOAc in hexane over 15 min) to provide ZerZ-butyl 7V-[6-(benzyloxy)-13,13-dioxo-6,15bis(trifluoromethyl)-19-oxa-l 3X⁶-thia-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,9,14,16-hexaen-17-yl]carbamate (EIZmixture) (157 mg, 55%). ’H NMR (400 MHz, Chloroform-d) δ 10.05 (s, 1H), 9.54 (s, 1H), 7.36 - 7.20 (m, 5H), 5.67 (q, J= 8.3, 7.8 Hz, 1H), 5.48 (q, J= 9.1, 8.7 Hz, 1H), 4.90 (s, 2H), 3.65 (ddd, J= 14.6, 12.6, 4.4 Hz, 1H), 3.58 - 3.47 (m, 217

1H), 3.06 - 2.84 (m, 2H), 2.60 - 2.34 (m, 2H), 2.23 (t, J= 10.5 Hz, 1H), 2.12 - 2.06 (m, 1H), 1.58 (d, J= 2.9 Hz, 9H)ppm. ¹⁹F NMR (376 MHz, Chloroform-d) δ -58.72, -74.24 ppm. ESI-MS m/z cale. 662.1634, found 663.2 (M+l)⁺; Rétention time: 0.85 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 ^x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 3: fert-Butyl A-[6-hydroxy-13,13-dioxo-6,15-bis(trifIuoromethyl)-19-oxa-13L⁶-thia3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate

A mixture of teri-butyl Æ-[6-(benzyloxy)-13,13-dioxo-6,15-bis(trifluoromethyl)-19-oxa13X⁶-thia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (E/Z mixture) (154 mg, 0.232 mmol), and Pd/C (74 mg of 10 % w/w, 0.070 mmol) in AcOH (1.5 mL) was stirred at room température under 180 psi H2 in a stainless Steel pressure vessel for 15 h. Then the mixture was filtered and the filtrate evaporated to provide the target terrtbutyl N-[6hydroxy- 13,13-dioxo-6,l 5-bis(trifluoromethyl)-19-oxa-13L⁶-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (136 mg, 96%), ESI-MS m/z cale. 574.1321, found 575.1 (M+l)⁺; Rétention time: 0.72 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 ^χ 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 4: (6/?)-17-Amino-6-hydroxy-6,15-bis(trifluoromethyI)-19-oxa-13I⁶-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-13,13-dione (enantiomer 1), Compound 9, and (6S)-17-amino-6-hydroxy-6,15-bis(trifluoromethyl)-19-oxa-13I⁶-thia218

3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-13,13-dione (enantiomer 2), Compound 10

A mixture of terZ-butyl V-[6-hydroxy-13,13-dioxo-6,15-bis(trifluoromethyl)-19-oxa13X⁶-thia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (134 mg, 0.22 mmol), TFA (2 mL), triisopropylsilane (67 pL, 0.33 mmol) and water (100 pL) was stirred at room température for 30 min and then solvent evaporated. The residue was coevaporated from acetonitrile (2X). The residue, dissolved into 2 mL acetonitrile, was subjected to préparative SFC with 330 pL injections through a préparative SFC eluting a gradient of 5 mM NH3 in methanol to CO2 (5-15% over 10 min) though a 21.2 X 250 mm AD column, 5 pm particle giving the first eluent (6Æ)-17-amino-6-hydroxy-6,15-bis(trifluoromethyl)-19-oxa-13X⁶thia-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaene-13,13 -dione (enantiomer 1) (38 mg, 37%). ’H NMR (400 MHz, DMSO-dô) δ 7.93 (s, 1H), 7.66 (s, 3H, D2O exchanged), 3.77 - 3.64 (m, 1H), 3.61 - 3.49 (m, 1H), 2.21 (t, J= 12.1 Hz, 2H), 2.08 (d, J= 15.0 Hz, 1H), 1.92 (dd, J= 12.5, 7.2 Hz, 1H), 1.56 (m, 6H) ppm; ¹⁹F NMR (376 MHz, DMSO-d6) δ 57.96, -78.11 ppm. ESI-MS m/z cale. 474.07965, found 475.0 (M+l)⁺; Rétention time: 1.16 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Ci8 column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C. UV/vis Xmax 231, 277, 356 nm.

Further elution provided the second eluent (65)-17-amino-6-hydroxy-6,15bis(trifluoromethyl)-19-oxa-13X⁶-thia-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaene-l3,13-dione (enantiomer 2) (36 mg, 35%). ’H NMR (400 MHz, DMSO-dô) δ 7.93 (s, 1H), 7.66 (s, 3H, D2O exchanged), 3.77 - 3.63 (m, 1H), 3.56 (td, J= 14.9, 13.7, 4.1 Hz, 1H), 2.21 (t, J= 11.9 Hz, 2H), 2.08 (dd,J= 14.1, 8.0 Hz, 1H), 1.97- 1.84 (m, 1H), 1.56 (m, 6H) ppm; ¹⁹F NMR (376 MHz, DMSO-d6) δ -57.96, -78.11. ESI-MS m/z cale. 474.07965, found 475.0 (M+l)⁺; Rétention time: 1.16 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by

219

Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 8: Préparation of (6R,12R)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 11

Step 1

Step 2

E/Z mixture

Step 1: tert-Butyl A-[2-[5-[(LR)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazoI-2-yl]-6-[(LR)-l-methyIbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7V-tertbutoxycarbonyl-carbamate

Dissolved ter/-butyl7V-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ter/-butoxycarbonyl-carbamate (159.3 g, 231.3 mmol) and triphenylphosphine (72.9 g, 277.9 mmol) in toluene (1 L), then added (2S)-pent-4-en-2-ol (28.7 mL, 278.9 mmol). Heated this mixture to 45 °C, then added DIAD (58.3 mL, 296.1 mmol) (exotherm) slowly over 40 min. For the next approximately 2 h, the mixture was cooled to room température. During this cooling period, after the first 10 minutes, triphenylphosphine (6.07 g, 23.14 mmol) was added. After a further 1 h, additional triphenylphosphine (3.04 g, 11.59 mmol) was added. After a further 23 min, DIAD (2.24 mL, 11.57 mmol) was added. After the ~2 h cooling to room température period, the mixture was cooled to 15 °C, and seed crystals of DIAD-triphenylphosphine oxide complex were added

220 which caused précipitation to occur, then added 1000 mL heptane. Stored the mixture at -20 °C for 3 days. Filtered out and discarded the precipitate and concentrated the filtrate to give a red residue/oil. Dissolved the residue in 613 mL heptane at 45 °C, then cooled to 0 °C, seeded with DIAD-triphenylphosphine oxide complex, stirred at 0 °C for 30 min, then filtered the solution. The filtrate was concentrated to a smaller volume, then loaded onto a 1.5 kg silica gel column (column volume = 2400 mL, flow rate = 600 mL/min). Ran a gradient of 1% to 6% EtOAc in hexanes over 32 minutes (8 column volumes), then held at 6% EtOAc in hexanes until the product finished eluting which gave ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[(17î)-l-methylbut-3-enoxy]-5(trifluoromethyl)-3-pyridyl]-jV-/e/7-butoxycarbonyl-carbamate (163.5 g, 93%). 'H NMR (400 MHz, Chloroform-d) δ 7.82 (s, 1H), 7.43 - 7.27 (m, 5H), 5.88 - 5.69 (m, 2H), 5.35 (h, J= 6.2 Hz, 1H), 5.16 - 4.94 (m, 4H), 4.81 (d, J= 10.7 Hz, 1H), 4.63 (d, J= 10.7 Hz, 1H), 2.58 - 2.15 (m, 6H), 1.42 (s, 18H), 1.36 (d, J= 6.2 Hz, 3H) ppm. ESI-MS m/z cale. 756.2958, found 757.3 (M+l)⁺; Rétention time: 4.0 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 2: terf-Butyl 7V-[(6Æ,12Æ)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yI]-A-ft?r/butoxycarbonyl-carbamate (E!Z mixture)

E/Z mixture

The following reaction was run, split equally between two, 12 L reaction flasks run in parallel. Mechanical stirring was employed, and reactions were subjected to a constant nitrogen gas purge using a course porosity gas dispersion tube. To each flask was added tert-butyl 7V-[2[5-[(12î)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[(17î)-lmethyIbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-A^r-/er/-butoxycarbonyl-carbamate (54 g, 71.36 mmol in each flask) dissolved in DCE (8 L in each flask) and both flasks were strongly purged with nitrogen at room température. Both flasks were heated to 62 °C and Grubbs l^stGénération Catalyst (9 g, 10.94 mmol in each flask) was added to each reaction and stirred at 221

400 rpm while setting an internai température control to 75 °C with strong nitrogen purging (both reactions reached -75 °C after approximately 20 min). After 5 h 15 min, the internai température control was set to 45 °C. After approximately 2 h, 2-sulfanylpyridine-3-carboxylic acid (11 g, 70.89 mmol in each flask) was added to each flask followed by triethylamine (10 mL, 71.75 mmol in each flask). On completion of addition, the nitrogen purge was tumed off and both reaction flasks were stirred at 45 °C open to air ovemight. The reactions were then removed from heat and 130 g of silica gel was added to each reaction and each was stirred at room température. After approximately 2 h, the green mixtures were combined and filtered over Celite then concentrated by rotary évaporation at 43 °C. The obtained residue was dissolved in dichloromethane/heptane 1:1 (400 mL) and the formed orange solid was removed by filtration. The greenish mother liquor was evaporated to give 115.5 g of a green foam. Dissolved this material in 500 mL of 1:1 dichloromethane/hexanes then loaded onto a 3 kg silica gel column (column volume = 4800 mL, flow rate = 900 mL/min). Ran a gradient of 2% to 9% EtOAc in hexanes over 43 minutes (8 column volumes), then ran at 9% EtOAc until the product finished eluting giving 77.8 g of impure product. This material was co-evaporated with methanol (-500 mL) then diluted with methanol (200 mL) to give 234.5 g of a methanolic solution, which was halved and each half was purified by reverse phase chromatography (3.8 kg Cis column, column volume = 3300 mL, flow rate = 375 mL/min, loaded as solution in methanol). Ran the column at 55% acetonitrile for -5 minutes (0.5 column volumes), then at a gradient of 55% to 100% acetonitrile in water over -170 minutes (19-20 column volumes), then held at 100% acetonitrile until the product and impurities finished eluting. Clean product fractions from both columns were combined and concentrated by rotary évaporation then transferred with éthanol into 5 L flask, evaporated and carefully dried (becomes a foam) to give as a mixture of olefin isomers, ZerZ-butyl N-[(6R, 127?)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5] nonadeca-1 ( 18),2,4,9,14,16-hexaen-17-yl]-A-ZerZ-butoxycarbonylcarbamate (E!Zmixture) (55.5 g, 53%). ESI-MS m/z cale. 728.26447, found 729.0 (M+l)⁺; Rétention time: 3.82 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

222

Step 3: tert-Butyl A-[(6/?,127?)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-7V-ter/‘butoxycarbonyl-carbamate

E/Z mixture

ter/-Butyl 7V-[(67î,12R)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaen-17-yl] -N-tertbutoxycarbonyl-carbamate {E/Z mixture) (11.7 g, 16.06 mmol) was dissolved in stirring éthanol (230 mL) and cycled the flask 3 times vacuum/nitrogen and treated with 10% Pd/C (50% water wet, 2.2 g of 5 % w/w, 1.034 mmol). The mixture was cycled 3 times between vacuum/nitrogen and 3 times between vacuum/hydrogen. The mixture was then stirred strongly under hydrogen (balloon) for 7.5 h. The catalyst was removed by filtration, replaced with fresh 10% Pd/C (50% water wet, 2.2 g of 5% w/w, 1.034 mmol) and stirred vigorously under hydrogen (balloon) ovemight. Then, the catalyst was removed again by filtration, the filtrate evaporated and the residue (11.3 g, 1 g set aside) was dissolved in éthanol (230 mL) charged with fresh 10% Pd/C (50% water wet, 2.2 g of 5 % w/w, 1.034 mmol) and stirred vigorously under hydrogen (balloon) for 6 h, recharged again with fresh 10% Pd/C (50% water wet, 2.2 g of 5 % w/w, 1.034 mmol) and stirred vigorously under hydrogen (balloon) ovemight. The catalyst was removed by filtration and the filtrate was evaporated (10 g of residue obtained). This crude material (10 g ΤΙ g set aside above) was purified by silica gel chromatography (330 g column, liquid load in dichloromethane) with a linear gradient of 0% to 15% ethyl acetate in hexane until the product eluted followed by 15% to 100% ethyl acetate in hexane to giving, as a colorless foam, ZerZ-butyl N-[(6R, 12R)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclof 12.3.1.12,5]nonadeca-1 (18),2,4,14,16-pentaen-l 7-yl]-7V-ter/-butoxycarbonylcarbamate (9.1 g, 78%). ESI-MS m/z cale. 730.2801, found 731.0 (M+l)⁺ ; Rétention time: 3.89 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

223

Step 4: (6Æ,127?)-17-Amino-12-mcthyI-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI, Compound 11

Zert-Butyl N-[(6R, 127?)-6-bcnzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[l 2.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-l l-y\]-N-tertbutoxycarbonyl-carbamate (8.6 g, 11.77 mmol) was dissolved in éthanol (172 mL) then the flask was cycled 3 times between vacuum/nitrogen. Treated the mixture with 10% Pd/C (50% water wet, 1.8 g of 5 % w/w, 0.8457 mmol) then cycled 3 times between vacuum/nitrogen and 3 times between vacuum/hydrogen and then stirred vigorously under hydrogen (balloon) at room température for 18 h. The mixture was cycled 3 times between vacuum/nitrogen, fïltered over Celite washing with éthanol and then the filtrate was evaporated to give 7.3 g of ZerZ-butyl NZer/-butoxycarbonyl-7V-[(6/?, 12/?)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate an offwhite solid. This material was dissolved in dichloromethane (69 mL), cooled in an ice bath under nitrogen and slowly treated with TFA (23 mL, 298.5 mmol). The solution was stirred in the ice bath for 5 min and then at room température for 1 h. The pale-yellow solution was diluted with heptane (-100 mL) and evaporated to give a yellow solid mass. The residue was diluted again with heptane (-100 - 200 mL) and dichloromethane was added under warming until a yellow solution was obtained. Most of the dichloromethane was removed by rotary évaporation (35 °C water bath, 100 mbar pressure) to give a fine yellow suspension. The suspension was swirled for -1 h at room température, fïltered washing the solid with dry ice chilled heptane and then dried over 3 days under vacuum with a nitrogen leak at 50 °C to give as a pale yellow solid, (6R,127î)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (4.68 g, 90%). *H NMR (400 MHz, DMSO-dô) δ 7.77 (s, 1H), 7.55 (s, 1H), 6.34 (s, 2H), 4.90 - 4.70 (m, 1H), 2.47 (dd, J= 7.8, 5.5 Hz, 1H), 2.29 (t, J= 11.2 Hz, 1H), 2.11 (ddd, J= 14.4, 8.7, 6.1 Hz, 1H), 1.73 (dt, J= 12.7, 7.6 Hz, 2H), 1.59 - 1.38 (m, 4H), 1.35 (d, J= 6.3 Hz, 3H), 1.18 (ddt, J= 12.4, 9.6, 6.2 Hz, 1H) ppm. Ή NMR (400 MHz, Chloroform-d) δ 7.42 (d, J= 0.8 Hz, 1H), 5.20 (s, 2H), 4.75 (dtt, J = 12.6, 6.3, 3.2 Hz, 1H), 3.98 (s, 1H), 2.68 (dtd, J= 12.9, 7.6, 2.3 Hz, 1H), 2.38-2.18 (m, 2H), 2.03 (d, J= 7.9 Hz, 1H), 1.75 - 1.46 (m, 5H), 1.41 (d, J= 6.3 Hz, 3H), 1.35 - 1.27 (m, 1H) ppm. ^,9F NMR (376 MHz, Chloroform-d) δ -63.95, -77.34 ppm. ESI-MS m/z cale. 440.1283, found

224

441.0 (M+l)⁺; Rétention time: 2.87 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes.

Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 5: Solid form chracterization of Compound 11 heptane solvaté

A. X-Ray Powder Diffraction

The X-ray powder diffraction (XRPD) diffractogram of the product of Step 4, Compound 11 heptane solvaté, was acquired at room température in transmission mode using a PANalytical Empyrean system equipped with a sealed tube source and a PIXcel 1D Medipix-3 detector (Malvem PANalytical Inc, Westborough, Massachusetts). The X-ray generator operated at a voltage of 45 kV and a current of 40 mA with copper radiation (1.54060 Â). The powder sample was placed on a 96 well sample holder with mylar film and loaded into the instrument. The sample was scanned over the range of about 3° to about 40°20 with a step size of 0.0131303° and 49s per step.

The XRPD diffractogram for Compound 11 heptane solvaté is provided in FIG. 1, and the XRPD data are summarized below in Table 4.

Table 4: XRPD signais for Compound 11 heptane solvaté

XRPD Peak No.	Angle (degrees 2Theta ± 0.2)	Intensity %
1	5.8265	100
2	10.1389	30.54
3	5.5705	22.28
4	18.1061	21
5	20.5379	15.76
6	11.7247	12.43
7	20.9306	11.25

XRPD diffractograms for Compound 11 heptane solvaté samples prepared under three different drying conditions are provided in FIG. 2. The XRPD diffractograms were recorded at room température in continuons mode using a PANalytical Empyrean X-ray Diffract meter (Almelo, The Netherlands). The X-Ray was generated using Cu tube operated at 45 kV and 40 mA. Pixel Id detector was used with anti-scatter slit P8. The Divergence optics is Bragg Brentano High Définition (BBHD) with a 10mm mask, 1/8 divergence slit, and '/? anti-scatter slit. The continuous scan mode utilized a 0.0131 degree step size and count time of 13.77 225 seconds per step, integrated over the range from 4 to 40 degrees two-theta. The powder sample was placed on an indented area within a zéro background holder and flattened with a glass slide.

Under Drying Condition 1, Compound 11 heptane solvaté was dried over the weekend under house vacuum with a nitrogen leak at 50 °C. Under Drying Condition 2,

Compound 11 heptane solvaté was dried over the weekend at 40-45 °C. Under Drying Condition 3, Compound 11 heptane solvaté was dried for 4 days under house vacuum with a nitrogen bleed at 40-45 °C.

The XRPD diffractograms for Compound 11 heptane solvaté samples prepared under Drying Condition 1, Drying Condition 2, and Drying Condition 3 are provided in FIG. 2, and the

XRPD data are summarized below in Tables 5, 6, and 7. In FIG. 2, the top curve corresponds to Drying Condition 2, the middle curve corresponds to Drying Condition 1, and the bottom curve corresponds to Drying Condition 3. Each curve is substantially similar to each other and to the XRPD ofFIG. 1.

Table 5: XRPD signais for Compound 11 heptane solvaté, Drying Condition 1

XRPD Peak No.	Angle (degrees 2-Theta ± 0.2)	Intensity %
1	5.8572	100
2	6.0844	62.33
3	10.189	34.55
4	10.5918	21.42
5	18.5355	10.59
6	18.1599	10.55
7	12.2176	10.14

Table 6: XRPD signais for Compound 11 heptane solvaté, Drying Condition 2

XRPD Peak No.	Angle (degrees 2-Theta ± 0.2)	Intensity %
1	5.9314	100
2	6.1405	52.48
3	10.2746	33.73
4	10.6383	18.32
5	11.9036	10

Table 7: XRPD signais for Compound 11 heptane solvaté, Drying Condition 3

XRPD Peak No.	Angle (degrees 2-Theta ± 0.2)	Intensity %
1	6.0778	100
2	5.812	81.49
3	10.6059	29.34
4	10.1304	21.77
5	12.27	12.64

226

B. Differential Scanning Calorimetry Analysis

The melting point of the product of Step 4, Compound 11 heptane solvaté, was measured using the TA Instruments Q2000 DSC.

The DSC thermogram for Compound 11 heptane solvaté is provided in FIG. 2. The thermogram for Compound 11 heptane solvaté shows an endotherm at -93.45 °C and recrystallization at -103 °C.

C. Solid-State ¹³C NMR

The ¹³C SSNMR of the product of Step 4, Compound 11 heptane solvaté, was acquired using the procedure described in the General Methods. The ^l3C SSNMR spectrum for

Compound 11 heptane solvaté Form is provided in FIG. 3, and the data are summarized below in Table 8.

Table 8:¹³C SSNMR signais for Compound 11 heptane solvaté

Peak #	Chem Shift [ppm]	Intensity [rel]
1	166.3	26.5
2	165.8	19.3
3	164.6	48.8
4	163.4	18.2
5	154.8	12.9
6	154.0	21.1
7	152.1	24.5
8	151.6	50.5
9	140.2	12.3
10	139.4	17.5
11	138.5	23.1
12	138.0	18.9
13	135.1	17.6
14	134.6	20.4
15	131.3	26.2
16	130.2	24.2
17	129.6	24.3
18	128.5	18.0
19	125.7	31.6
20	123.7	15.1
21	123.2	14.6
22	122.9	16.7
23	121.1	21.9
24	120.2	40.6
25	119.2	15.1
26	117.8	48.6
27	76.2	31.3

227

Peak#	Chem Shift [ppm]	Intensity [rel]
28	74.4	84.8
29	73.7	84.5
30	73.3	73.1
31	40.0	22.3
32	38.6	12.0
33	37.6	27.9
34	36.9	32.7
35	35.7	12.6
36	33.6	13.5
37	32.5	57.6
38	32.0	69.7
39	30.4	56.4
40	30.1	50.3
41	29.5	44.7
42	28.8	36.63
43	28.1	21.1
44	27.1	29.6
45	25.3	27.1
46	23.1	90.9
47	22.7	88.1
48	22.0	46.4
49	21.6	48.1
50	20.3	100.0
51	19.6	83.0
52	18.3	24.6
53	17.6	47.9
54	13.8	51.6
55	13.1	39.3
56	12.5	82.3

D. Solid-State ^,9F NMR

The ¹⁹F SSNMR of the product of Step 4, Compound 11 heptane solvaté, was acquired using the procedure described in the General Methods. The ¹⁹F SSNMR spectrum for

Compound 11 heptane solvaté is provided in FIG. 4, and the data are summarized below in

Table 9.

Table 9: ¹⁹F SSNMR signais for Compound 11 heptane solvaté

Peak#	Chem Shift [ppm]	Intensity [rel]
1	-63.5	4.4
2	-63.8	3.4
3	-65.1	12.2
4	-65.8	7.6
5	-66.3	7.8

228

6	-67.0	12.5
7	-74.0	2.1
8	-74.9	4.2
9	-76.6	10.0
10	-77.6	5.9

E. Thermogravimetric Analysis (TGA)

TGA was used to investigate the presence of residual solvents in the lots characterized and identify the température at which décomposition of the sample occurs. TGA data were 5 collected on a Mettler Toledo TGA/DSC 3+ STARe System.

The TGA curve for Compound 11 heptane solvaté prepared under Drying Condition 1 is provided in FIG. 6A. The TGA curve for Compound 11 heptane solvaté prepared under Drying Condition 2 is provided in FIG. 6B. The TGA curve for Compound 11 heptane solvaté prepared under Drying Condition 3 is provided in FIG. 6C. Each of the curves in FIGS. 6A, 10 6B, and 6C are substantially similar to each other.

Example 9: Préparation of (65’,127?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19- dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait), Compound 12

Step 1

Step 1 : fôrt-Butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2yl]-6-[(lÆ)-l-methyIbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyI]carbamate

229

To a solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-/eri-butoxycarbonyl-carbamate (500 mg, 0.6653 mmol) in DMSO (5 mL) was added (2/?)-pent-4-en-2-ol (350 pL, 3.401 mmol), césium carbonate (751 mg, 2.305 mmol) and iodocopper (31 mg, 0.1628 mmol) and the reaction mixture was heated at 100 °C for 6 h in an oil bath. The reaction mixture was poured onto crushed ice and extracted with ethyl acetate and washed with brine. The organics were separated, dried over sodium sulfate, filtered and evaporated. The résultant brown residue was purified by silica gel chromatography using a shallow gradient 100% hexanes to 50% EtOAc in hexanes to afford as light brown viscous oil, ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[(lR)-l-methylbut-3-enoxy]-5(trifluoromethyl)-3-pyridyl]carbamate (120 mg, 27%). ’H NMR (400 MHz, DMSO-dô) δ 9.71 (s, 1H), 8.86 (s, 1H), 7.46 - 7.28 (m, 5H), 5.91 - 5.72 (m, 2H), 5.24 (qt, J= 6.1, 3.1 Hz, 1H), 5.16 - 4.97 (m, 4H), 4.75 (d, J= 11.0 Hz, 1H), 4.66 (d, J= 11.0 Hz, 1H), 2.50 - 2.40 (m, 4H), 2.32 (d, J = 8.1 Hz, 2H), 1.49 (s, 9H), 1.32 (d, J= 6.2 Hz, 3H) ppm. ESI-MS m/z cale. 656.24335, found 657.3 (M+l)⁺ ; Rétention time: 0.84 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60

Step 2: tert-Butyl A^r-[(12Æ)-6-benzyloxy-12-methyI-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (EIZ mixture)

To a degassed solution of [l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]dichloro-[(2-isopropoxyphenyl)methylene]ruthenium (32 mg, 0.05107 mmol) (Grubbs 2nd génération catalyst) in DCE (50 mL) was added a degassed solution of ZerZ-butyl 7V-[2-[5-[lbenzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[(lR)-l-methylbut-3-enoxy]5-(trifluoromethyl)-3-pyridyl]carbamate (150 mg, 0.2284 mmol) in DCE (50 mL) slowly dropwise under a stream of nitrogen flow bubbling through the solution over 30 min and the

230 reaction mixture was then heated at 50 °C for 5 h. The reaction was stopped, and the solvents were removed by rotary évaporation. The résultant brown residue was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 30% EtOAc in hexanes to afford as a mixture of olefin isomers, terrtbutyl A-[(12/?)-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclof 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16hexaen-17-yl]carbamate (E/Z mixture) (102 mg, 36%) which co-eluted with some unreacted starting material. ESI-MS m/z cale. 628.21204, found 629.2 (M+l)⁺; Rétention time: 0.36 minutes. This material was used directly in the ensuing step. ESI-MS m/z cale. 628.21204, found 629.2 (M+l)⁺; Rétention time: 0.36 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 3: tert-Butyl A-[(127?)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate

To a solution of teri-butyl 7V-[(127?)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-l ( 18),2,4,9,14,16-hexaen-1Ίyl]carbamate (E/Z mixture) (102 mg, 0.08114 mmol) in AcOH (5 mL) was added 10% w/w Pd/C (54 mg, 0.05074 mmol) in a 250 mL flask equipped with a hydrogen balloon using a 3-way adaptor. Subjected the flask to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. Filled the vessel with hydrogen gas and the mixture was stirred at room température for 15 h. Subjected the vessel to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated to afford ZerZ-butyl JV-[( 12//)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (120 mg, 96%) which was used directly in the ensuing step. ESI-MS m/z cale. 540.1807, found 541.2 (M+l)⁺; Rétention time: 0.57 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn:

231

186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 4: (65,12/?)-17-Amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait), Compound 12

To a solution of ZerZ-butyl 7V-[(12Æ)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-17yl]carbamate (120 mg, 0.2220 mmol) was added TFA (500 mL, 6.490 mol) and dichloromethane (1.5 mL) (pre made solution of 1:4 TFA/dichloromethane) and the reaction was stirred at room température for about 1 h. The solvents were removed by évaporation and the residue was dissolved in DMSO (1 mL) and purified by reverse phase HPLC using a gradient run from 40% to 85% acetonitrile in water (+ 5 mM HCl) over 30.0 minutes to afford as a light brown solid and the second diastereomer to elute, (65',121î)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (14.7 mg, 28%). Ή NMR (400 MHz, DMSO-d₆) δ 7.78 (s, 1H), 4.72 (ddt, J= 11.5, 7.7, 3.8 Hz, 1H), 2.61 - 2.52 (m, 1H), 2.24 - 2.03 (m, 2H), 1.68 (s, 1H), 1.59 (h, J= 7.5, 6.4 Hz, 2H), 1.54 - 1.38 (m, 3H), 1.34 (d, J= 6.3 Hz, 3H), 1.21 - 1.10 (m, 1H) ppm. ESI-MS m/z cale. 440.1283, found 441.1 (M+l)⁺; Rétention time: 2.87 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °n

232

Example 10: Préparation of (6S',125)-17-aniino-12-methyl-6,15-bis(trifluoromethyl)-13,19- dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait), Compound 13

Step 1: tert-Butyl 7V-[2-[5-[l-benzyloxy-l-(trifIuoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2yl]-6-[(15)-l-methylbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7V-fôr/-butoxycarbonylcarbamate

To a solution of ZerZ-butyl A-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-A^r-ZerZ-butoxycarbonyl-carbamate (800 mg, 1.065 mmol) in DMSO (10 mL) was added (2S)-pent-4-en-2-ol (550 pL, 5.345 mmol), césium carbonate (1.1 g, 3.376 mmol) and iodocopper (47 mg, 0.2468 mmol) and the reaction mixture was heated at 80 °C for 3 h. The reaction mixture was poured onto crushed ice and extracted with ethyl acetate and the organic phase was washed with brine. The organic layer was dried over sodium sulfate, filtered and evaporated. The résultant brown residue was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 30% EtOAc in hexanes to afford tert-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-[(15)-l-methylbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-V-/erZbutoxycarbonyl-carbamate (303 mg, 38%). ’H NMR (400 MHz, DMSO-dô) δ 8.21 (s, 1H), 7.41 - 7.30 (m, 5H), 5.90 - 5.75 (m, 2H), 5.43 (qt, J= 7.7, 3.9 Hz, 1H), 5.12 - 4.98 (m, 4H), 4.81 (d, J = 11.1 Hz, 1H), 4.68 (d, J= 11.0 Hz, 1H), 2.60 - 2.47 (m, 4H), 2.39 - 2.26 (m, 2H), 1.38 - 1.26

233 (m, 21H) ppm. ESI-MS m/z cale. 756.2958, found 757.47 (M+l)⁺; Rétention time: 0.78 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H).

Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température — 60 °C.

Step 2: teri-Butyl A^r-[(125)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo [12.3.1.12,5] nonadeca-1 (18),2,4,9,14,16-hexaen-17-yl]carbamate (EtZ mixture)

To a degassed solution of [l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]dichloro-[(2-isopropoxyphenyl)methylene]ruthenium (43 mg, 0.06862 mmol) (Grubbs 2nd génération catalyst) in DCE (50 mL) was added a degassed solution of ZerZ-butyl 7V-[2-[5-[lbenzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[(15)-l-methylbut-3-enoxy]5-(trifluoromethyl)-3-pyridyl]-A-ier/-butoxycarbonyl-carbamate (200 mg, 0.2643 mmol) in DCE (50 mL) slowly dropwise under a stream of nitrogen flow bubbling through the solution over 30 min and on completion of addition the reaction mixture was heated at 50 °C for 5 h. The reaction was stopped, and the solvents removed by rotary évaporation. The résultant brown residue was purified by silica gel chromatography using a shallow gradient from 100% hexanes to 30% EtOAc in hexanes to afford as a mixture of olefin isomers, ZerZ-butyl 7V-[(12S)-6-benzyloxy-12methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,9,14,16-hexaen-17-yl] carbamate (E/Z mixture) (150 mg, 45%) along with some unreacted starting material which co-eluted with the product. This material was used directly in the ensuing step without further purification. ESI-MS m/z cale. 628.21204, found 629.3 (M+l)⁺; Rétention time: 0.7 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

234

Step 3: (65,125)-17-Amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait), Compound 13

To a solution of ter/-butyl 7/-[(125)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16-hexaen-17yl]carbamate (E/Zmixture) (150 mg, 0.2386 mmol) in AcOH (5 mL) was added 10 % w/w Pd/C (78 mg, 0.07329 mmol) in a 250 mL flask equipped with a hydrogen balloon using a 3-way adapter. Subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. Filled the vessel with hydrogen gas and the mixture was stirred at room température for 15 h. Subjected to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated to afford 125 mg of fôrt-butyl N[(125)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate as a mixture of diastereomers. This material was dissolved in a 1:3 mixture of a premixed solution of TFA (500 pL, 6.490 mmol) and dichloromethane (1.5 mL) and the mixture was stirred for 30 min at room température. The solvent was removed by rotary évaporation and the resulting residue was purified by reverse phase HPLC using a gradient from 40% to 85% acetonitrile in water (+ 5 mM HCl) over 30.0 minutes to afford as a light brown solid and the first enantiomer to elute, (65,125)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait) (20.6 mg, 36%). Ή NMR (400 MHz, Chloroform-d) δ 7.42 (s, 1H), 4.75 (ddt, J= 10.6, 6.7, 3.4 Hz, 1H), 4.06 - 3.76 (m, 1H), 2.76 - 2.63 (m, 1H), 2.28 (t, J= 7.6 Hz, 2H), 2.01 (d, J= 5.7 Hz, 1H), 1.61 (m, 4H), 1.50 (dd, J= 12.0, 6.5 Hz, 1H), 1.41 (d, J= 6.3 Hz, 3H), 1.33 - 1.26 (m, 1H), 0.89 (m, 1H) ppm. ESI-MS m/z cale. 440.1283, found 441.1 (M+l)⁺ ; Rétention time: 2.82 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

235

Example 11: Préparation of (6R,12S)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 14

Step 1

Step 2

Step 1: tert-Butyl A-[2-[5-[(17?)-l-benzyIoxy-l-(trifluoromethyl)pcnt-4-enyI]-l,3,4 oxadiazol-2-yl]-6-[(l>y)-l-methylbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]carbamate

To a solution of /er/-butyl 7V-[2-[5-[(lA)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]carbamate (417.3 mg, 0.7091 mmol) and (22?)-pent-4-en-2-ol (109.5 pL, 1.064 mmol) in toluene (8.784 mL) was added triphenylphosphine (246.6 pL, 1.064 mmol). After stirring at room température for 1 min, DIAD (223.5 pL, 1.135 mmol) was added and the mixture was stirred at room température for 5 min. Diluted the reaction mixture with EtOAc then washed with saturated aqueous sodium bicarbonate (IX), saturated aqueous NH4CI (IX) and brine (IX) then dried over magnésium sulfate, filtered and concentrated to a yellow oil which was purified by silica gel chromatography using a gradient from 100% hexanes to 100% EtOAc giving as a clear, slightly yellow syrup, ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6[(15)-l-methylbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]carbamate (433.8 mg, 93%). *H NMR (400 MHz, Chloroform-d) δ 9.81 (s, 1H), 9.16 (s, 1H), 7.42 - 7.37 (m, 2H), 7.36 - 7.28 (m, 3H), 5.90 - 5.72 (m, 2H), 5.26 (q, J= 6.1 Hz, 1H), 5.13 - 4.97 (m, 4H), 4.83 (d, J= 10.8 Hz, 1H), 4.68

236 (d, J= 10.9 Hz, 1H), 2.51 (m, 2H), 2.46 - 2.36 (m, 3H), 2.27 (d, J= 11.2 Hz, 1H), 1.55 (s, 9H), 1.36 (d, J = 6.2 Hz, 3H) ppm. ESI-MS m/z cale. 656.24335, found 657.3 (M+l)⁺; Rétention time: 0.86 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 2: terf-Butyl A-[(67?,125)-6-benzyloxy-12-methyl-6,15-bis(trifluoromcthyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (E/Z mixture)

Ck .NH

E/Z mixture

To a degassed solution of [l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]dichloro-[(2-isopropoxyphenyl)methylene]ruthenium (378.2 mg, 0.6036 mmol) (Hoveyda Grubbs 2nd Gen catalyst) in toluene (229 mL) stirring at 100 °C with a reflux condenser and nitrogen bubbling through the solution was added a degassed solution of Zeri-butyl 7V-[2-[5[( 1 R)-1 -benzyloxy-1 -(trifluoromethyl)pent-4-enyl]-1,3,4-oxadiazol-2-yl]-6-[( 15)-1 -methylbut-3 enoxy]-5-(trifluoromethyl)-3-pyridyl]carbamate (1.5269 g, 2.325 mmol) in toluene (229 mL) slowly dropwise under a stream of nitrogen flow bubbling through the solution over 10 min and the reaction mixture was heated at 100 °C for 60 min. The mixture was removed from the heating bath and 2-sulfanylpyridine-3-carboxylic acid (180.5 mg, 1.163 mmol) was added. The resulting mixture was stirred for 10 min then concentrated by rotary évaporation to a residue which was chromatographed on a 275 g reverse phase Cis column eluting with a gradient from 50% to 100% acetonitrile in water giving as an off-white/yellow foam, ZerZ-butyl //-((67^,125)-6benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclof 12.3.1.12,5]nonadeca-l ( 18),2,4,9,14,16-hexaen-l 7-yl]carbamate (E/Z mixture) (324.3 mg, 22%). ESI-MS m/z cale. 628.21204, found 629.2 (M+l)⁺; Rétention time: 0.82 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 %

237

CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 3: tert-Butyl 7V-[(6Æ,125)-6-hydroxy-12-methyI-6,15-bis(trifluoromethyI)-13,19-dioxa 3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate

To a solution of Zer/-butyl A-[(67?,120)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16-hexaen-17yl]carbamate (E/Z mixture) (324.3 mg, 0.5159 mmol) in AcOH (10.81 mL) was added 10% w/w Pd/C (168.7 mg, 0.1585 mmol, 50% water wet) and hydrogen gas was bubbled through the stirring mixture for 15 minutes then the reaction was sealed and capped with a hydrogen balloon and stirred for 16 h. Added 10 % w/w Pd/C (54.9 mg, 0.05159 mmol, 50% water wet), stirred for 1 h then purged the flask with nitrogen and filtered over Celite eluting with EtOAc. The filtrate was concentrated then purified by silica gel chromatography using a gradient from 100% hexanes to 100% EtOAc giving as a white foam, teri-butyl A-[(67?,125)-6-hydroxy-12-methyI6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (216 mg, 77%). ESI-MS m/z cale. 540.1807, found 541.2 (M+l)⁺; Rétention time: 0.61 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 «r¹

Step 4: (6/?,12y)-17-Amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 14

238

To a stirring solution of /err-butyl V-[(67?,125)-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (216 mg, 0.3997 mmol) in dichloromethane (2.16 mL) was added TFA (769.6 pL, 9.989 mmol) and the resulting mixture was stirred at room température for 1 h then concentrated by rotary évaporation to a yellow residue. The residue was chromatographed on a 100 g reverse phase Cis column eluting with a gradient from 50% to 100% acetonitrile in water giving as a pale yellow solid, (6?î,12iS)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (177.1 mg, 100%). ’H NMR (400 MHz, DMSO-d₆) δ 7.76 (s, 1H), 7.59 (s, 1H), 6.36 (s, 2H), 4.73 (dq, J= 6.4, 3.1, 2.4 Hz, 1H), 2.56 (d, J= 5.5 Hz, 1H), 2.22 - 2.04 (m, 2H), 1.74 - 1.64 (m, 1H), 1.59 (d, J= 7.9 Hz, 2H), 1.54 - 1.43 (m, 3H), 1.34 (d, J= 6.3 Hz, 3H), 1.22 - 1.10 (m, 1H) ppm. ESI-MS m/z cale. 440.1283, found 441.1 (M+l)⁺; Rétention time: 2.02 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Example 12: Préparation of 16-amino-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa-12L⁶thia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-oI (enantiomer 1), Compound 15 and 16-amino-12,12-dioxo-6,14-bis(trifluoromethyI)-18-oxa-12I⁶-thia-3,4,17-

239 triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (enantiomer 2), Compound 16

Step 1

Step 4

Step 3

enantiomer 1 enantiomer 2

Step 1: tert-Butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazoI-2-yl] 6-but-3-enylsulfonyl-5-(trifluoromethyl)-3-pyridyI]carbamate

A mixture of /eri-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (225 mg, 0.3530 mmol), but3-ene-l-sulfmate (sodium sait) (150 mg, 1.055 mmol), and iodocopper (202 mg, 1.061 mmol) in

DMSO (2.2 mL) was heated at 100 °C for 3 h, then diluted with ether and water, filtered through Celite, the layers partitioned and the organic layer washed with water, brine, dried (MgSCU) and evaporated. Purification by silica gel chromatography (0 - 20% EtOAc in hexanes) provided ZerZ-butyl jV-[2-[5-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-but-3enylsulfonyl-5-(trifluoromethyl)-3-pyridyl]carbamate (99 mg, 41%). *H NMR (400 MHz,

Chloroform-d) δ 10.36 (s, 1H), 9.56 (s, 1H), 7.41 - 7.28 (m, 5H), 5.91 (dt, J= 17.0, 8.5 Hz, 1H), 5.74 (ddt, J = 16.8, 10.2, 6.5 Hz, 1H), 5.32 - 5.17 (m, 2H), 5.10 - 5.04 (m, 1H), 5.02 (dq, J=

240

10.2, 1.3 Hz, 1H), 4.83 (d, J= 10.9 Hz, 1H), 4.67 (d, J= 10.9 Hz, 1H), 3.80 - 3.60 (m, 2H), 3.29 - 3.14 (m, 2H), 2.65 - 2.56 (m, 2H), 1.54 (s, 9H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ 58.49, -73.15 ppm. ESI-MS m/z cale. 676.179, found 677.2 (M+l)⁺; Rétention time: 0.87 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: terf-Butyl 7V-[6-benzyloxy-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa-12I⁶-thia3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,8,13(17),14-hexaen-16-yl]carbamate (E/Z mixture)

In a 3-neck round bottom flask, a solution of teri-butyl 7V-[2-[5-[l-benzyloxy-l(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-but-3-enylsulfonyl-5-(trifluoromethyl)-3pyridyl]carbamate (206 mg, 0.3045 mmol) in DCE (10 mL) was slowly added dropwise from an addition tunnel over 45 min to a solution of dichloro[l,3-bis(2,4,6-trimethylphenyl)-2imidazolidinylidene][[5-[(dimethylamino)sulfonyl]-2-(l-methylethoxy-O)phenyl]methyleneC]ruthenium(II) (34 mg, 0.04634 mmol) in DCE (25 mL) heated at 70 °C with N2 bubbling through the solution. The mixture was stirred at 70 °C with N2 bubbling a further 90 min and then the solvent was evaporated. Purification by silica gel chromatography (0 - 20% EtOAc in hexanes) provided ZerZ-butyl 7V-[6-benzyloxy-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa12X⁶-thia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,8,13(17),14-hexaen-16yl]carbamate (E/Z mixture) (92 mg, 44%). ’H NMR (400 MHz, Chloroform-d) δ 9.46 (s, 1H), 9.42 (s, 1H), 7.37 - 7.27 (m, 2H), 7.25 - 7.16 (m, 3H), 5.88 - 5.64 (m, 2H), 4.92 (d, J= 11.6 Hz, 1H), 4.54 (d, J= 11.6 Hz, 1H), 4.13 - 3.98 (m, 1H), 3.59 (dt, J= 15.6,5.9 Hz, 1H), 3.12 (dd, J= 14.6, 4.8 Hz, 1H), 2.87 - 2.76 (m, 2H), 2.69 - 2.62 (m, 1H), 1.58 (s, 9H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -58.84, -74.24 ppm; UV/vis k_max 233, 268, 322 nm. ESI-MS m/z cale. 648.1477, found 649.1 (M+l)⁺; Rétention time: 0.83 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0

241 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 %

CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60

Step 3: ZerZ-Butyl A-[6-hydroxy-12,12-dioxo-6,14-bis(trifIuoromethyI)-18-oxa-12X⁶-thia 3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,13(17),14-pentaen-16-yl]carbamate

A mixture of terZ-butyl 7V-[6-benzyloxy-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa12X⁶-thia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,8,13(17),14-hexaen-16yljcarbamate (ElZ mixture) (90 mg, 0.1304 mmol) and Pd/C (42 mg of 10 % w/w, 0.03947 mmol) in AcOH (850 pL) was stirred at room température under 180 psi H2 in a stainless Steel pressure vessel for 36 h. Then the mixture was filtered and the filtrate was evaporated to provide ter/-butyl 7V-[6-hydroxy-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa-12X⁶-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,13(17),14-pentaen-16-yl]carbamate (81 mg, 102%). ESI-MS m/z cale. 560.1164, found 561.2 (M+l)⁺; Rétention time: 0.69 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C; UV/vis X_max 233, 268, 323 nm.

Step 4: 16-Amino-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa-12L⁶-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (enantiomer 1) and 16amino-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa-12I⁶-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-oI (enantiomer 2)

enantiomer 1

enantiomer 2 /eri-Butyl7V-[6-hydroxy-12,12-dioxo-6,14-bis(trifluoromethyl)-18-oxa-12X⁶-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,13(17),14-pentaen-16-yl]carbamate (81 mg, 0.13

242 mmol) was dissolved into TFA (1.27 mL), water (68 pL) and triisopropylsilane (40 pL, 0.1953 mmol), and stirred at room température for 15 min, then solvents evaporated. The residue was subjected to chiral séparation by SFC chromatography using a ChiralPak AD (250 X 21.2 mm column, 5 pm particle size) with 5% to 30% methanol (5 mM NH3)/and carbon dioxide mobile phase at 10 mL/min over 10.0 min (injection volume = 330 pL of 25mg/mL solution in methanol) giving as the first enantiomer to elute 16-amino-12,12-dioxo-6,14bis(trifluoromethyl)-18-oxa-12X⁶-thia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15pentaen-6-ol (16 mg, 27%) as a white crystalline solid. *H NMR (400 MHz, DMSO-dô) δ 7.84 (s, 1H), 7.66 (s, 1H, D2O exchanged), 7.39 (s, 2H, D2O exchanged), 3.87 - 3.69 (m, 1H), 3.57 3.41 (m, 1H), 2.30 - 2.02 (m, 4H), 1.70 (m, 2H), 1.58 - 1.45 (m, 2H) ppm; ¹⁹F NMR (376 MHz, DMSO-dô) δ -58.27, -77.73; UV/vis Xmax 230, 275, 353 nm. ESI-MS m/z cale. 460.064, found 461.0 (M+l)⁺; Rétention time: 1.04 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CH07644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

The second enantiomer to elute was 16-amino-12,12-dioxo-6,14-bis(trifluoromethyl)-18oxa-12X⁶-thia-3,4,17-triazatricyclo[l 1.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (14.4 mg, 24%) as a white crystalline solid. *H NMR (400 MHz, DMSO-dô) δ 7.84 (s, 1H), 7.66 (s, 1H, D2O exchanged), 7.39 (s, 2H, D₂O exchanged), 3.84 - 3.70 (m, 1H), 3.55 - 3.43 (m, 1H), 2.31 2.00 (m, 4H), 1.70 (s, 2H), 1.56 - 1.45 (m, 2H) ppm; ¹⁹F NMR (376 MHz, DMSO-dô) δ -58.27, 77.73 ppm; UV/vis Xmax 230, 275, 353 nm. ESI-MS m/z cale. 460.064, found 460.9 (M+l)⁺; Rétention time: 1.04 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 13: Préparation of21-amino-6,19-bis(trifluoromethyl)-17,23-dioxa-3,4,22triazatetracyclo[16.3.1.12,5.011,16]tricosa-l(22),2,4,ll(16),12,14,18,20-octaen-6-ol (hydrochloride sait), Compound 17

243

Step 7

Step 1: Methyl 6-(2-alIylphenoxy)-3-[bis(tert-butoxycarbonyl)amino]-5(trifIuoromethyI)pyridine-2-carboxylate

To a solution of methyl 3-[bis(terZ-butoxycarbonyl)amino]-6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (1.2 g, 2.403 mmol) in DMSO (12 mL) was added 2allylphenol (380 pL, 2.911 mmol), césium carbonate (2.43 g, 7.458 mmol) and iodocopper (100 mg, 0.5251 mmol) and the reaction mixture was heated at 100 °C for 30 min in an oil bath. LCMS shows completion of the reaction. The reaction mixture was poured on crushed ice and extracted with ethyl acetate and washed with brine. The organics were separated, dried over sodium sulfate, and evaporated. The résultant brown residue was purified by silica gel column

244 chromatography using a gradient of 100% hexanes to 100% EtOAc to afford methyl 6-(2allylphenoxy)-3-[bis(/eri-butoxycarbonyl)amino]-5-(trifluoromethyl)pyridine-2-carboxylate (612 mg, 46%). Ή NMR (400 MHz, Chloroform-d) δ 7.85 (s, 1H), 7.30 - 7.25 (m, 2H), 7.20 (t, 7.6 Hz, 2H), 5.88 (ddt, J= 16.8, 10.1, 6.6 Hz, 1H), 5.02 - 4.94 (m, 2H), 3.79 (s, 3H), 3.32 (d, J= 6.7 Hz, 2H), 1.42 (s, 18H) ppm; ESI-MS m/z cale. 552.2083, found 453.1 (M+l)⁺; Rétention time: 0.75 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 30 - 99% mobile phase B over 1.0 minutes. Mobile phase A — H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 2: 6-(2-AIIylphenoxy)-3-(terf-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid

To a solution of methyl 6-(2-allylphenoxy)-3-[bis(ter/-butoxycarbonyl)amino]-5(trifluoromethyl)pyridine-2-carboxylate (612 mg, 1.108 mmol) in THF (8 mL), MeOH (8 mL), and water (8 mL) was added LiOH (86 mg, 3.591 mmol). The mixture was stirred at room température for 30 min.

THF and methanol were removed under reduced pressure and then 10 mL HCl (10%) was carefully added to pH ~ 6 and the product was extracted by EtOAc (2 X 50 mL). The organic phases were combined, washed with brine (5 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo and placed under high vacuum for 12 hours to afford as a yellow solid 6-(2-allylphenoxy)-3-(fôr/-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid (482 mg, 99%). ESI-MS m/z cale. 438.14026, found 439.13 (M+l)⁺; Rétention time: 0.69 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 30 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

245

Step 3: ter/-Butyl A-[6-(2-allylphenoxy)-2-[[[2-benzyloxy-2-(trifIuoromethyl)pent-4enoyl] amino] carbamoyl]-5-(trifluoromethyl)-3-pyridyl] carbamate

To a solution of 6-(2-allylphenoxy)-3-(te/7-butoxycarbonylamino)-5(trifluoromethyl)pyridine-2-carboxylic acid (394.8 mg, 0.9006 mmol) and 2-benzyloxy-2(trifluoromethyl)pent-4-enehydrazide (312.5 mg, 1.084 mmol) in DMF (5 mL) was added DIEA (527.0 pL, 3.026 mmol), followed by HATU (526.6 mg, 1.385 mmol). The reaction mixture was stirred at room température for 4 h. The mixture was diluted with water and extracted with ethyl acetate. The organic phases combined and dried over MgSCh, filtered, and concentrated in vacuo. The résultant brown residue was purified by silica gel column chromatography using a gradient of 100% hexanes to 30% EtOAc-hexanes to afford Zeri-butyl 7V-[6-(2-allylphenoxy)-2[[[2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]-5-(trifluoromethyl)-3pyridyl]carbamate (543 mg, 85%). ESI-MS m/z cale. 708.2383, found 808.0 (M+l)⁺; Rétention time: 0.7 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 4: terZ-Butyl 7V-[6-(2-allylphenoxy)-2-[5-[l-benzyIoxy-l-(trifIuoromethyl)but-3-enyI] l,3,4-oxadiazoI-2-yI]-5-(trifluoromethyl)-3-pyridyI]carbamate

To a solution of teri-butyl 7V-[6-(2-allylphenoxy)-2-[[[2-benzyloxy-2(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]-5-(trifluoromethyl)-3-pyridyl]carbamate (630 mg, 0.8890 mmol) in acetonitrile (75 mL) was added DIEA (500 pL, 2.871 mmol) and was heated to 70 °C, then 4-methylbenzenesulfonyl chloride (255 mg, 1.338 mmol) was added in 3

246 portions (85 mg each portion in 10 min intervals). The resulted mixture was heated at 70 °C for 16 hours. The reaction mixture was cooled and quenched with saturated solution of sodium bicarbonate and extracted with ethyl acetate. The combined organic layers were washed with brine, dried over sodium sulfate, filtered, and evaporated in vacuo. The résultant brown residue was purifïed by silica gel column chromatography using a gradient from 100% hexanes to 30% EtOAc to afford Zeri-butyl 7V-[6-(2-allylphenoxy)-2-[5-[l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3-pyridyl]carbamate (336 mg, 55%). ’H NMR (400 MHz, DMSO-dô) δ 9.72 (s, 1H), 8.99 (s, 1H), 7.35 - 7.29 (m, 4H), 7.25 (m, 5H), 5.90 - 5.72 (m, 2H), 5.23 - 5.10 (m, 2H), 4.96 - 4.87 (m, 2H), 4.66 (d, J= 10.9 Hz, 1H), 4.52 (d, J= 10.9 Hz, 1H), 3.29 (dd, J= 11.3, 6.8 Hz, 2H), 3.10 (d, J= 7.1 Hz, 2H), 1.49 (s, 9H) ppm. ESI-MS m/z cale. 690.22766, found 691.3 (M+l)⁺; Rétention time: 0.81 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 ^x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 5: teri-Butyl A-[6-(benzyIoxy)-6,19-bis(trifluoromethyI)-17,23-dioxa-3,4,22triazatetracyclo[16.3.1.12,5.011,16]tricosa-l(22),2,4,8,ll(16),12,14,18,20-nonaen-21yl] carbamate (E!Z mixture)

To a degassed solution of benzylidene-[l,3-bis(2,4,6-trimethylphenyl)imidazolidin-2ylidene]-dichloro-ruthenium;tricyclohexylphosphane (6 mg, 0.007067 mmol) (Grubbs-2nd Gen catalyst) in DCE (30 mL) was added degassed solution of ter/-butyl 7V-[6-(2-allylphenoxy)-2-[5[ 1 -benzyloxy-1 -(trifluoromethyl)but-3-enyl]-1,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3pyridyl]carbamate (212 mg, 0.3070 mmol) in DCE (30 mL) slowly dropwise under a stream of N2 flow bubbling through the solution over 30 min and the reaction mixture was heated at 50 °C for 5h. The température was increased to 70 °C and the reaction mixture was heated ovemight. The reaction was stopped, and the solvents removed in vacuo. The résultant brown residue was purifïed by silica gel column chromatography using a shallow gradient of 100% hexanes to 30%

247

EtOAc-hexanes to afford Zeri-butyl 7V-[6-(benzyloxy)-6,19-bis(trifluoromethyl)-17,23-dioxa3,4,22-triazatetracyclo[16.3.1.12,5.01 l,16]tricosa-l(22),2,4,8,ll(16),12,14,18,20-nonaen-21yl]carbamate {E/Zmixture) (126 mg, 62%). ESI-MS m/z cale. 662.1964, found 663.19 (M+l)⁺; Rétention time: 0.74 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 6: tert-Butyl 7V-[6-hydroxy-6,19-bis(trifluoromethyl)-17,23-dioxa-3,4,22triazatetracyclo[16.3.1.12,5.011,16]tricosa-l(22),2,4,ll(16),12,14,18,20-octaen-21yljcarbamate

To a solution of a ~1:1 isomeric mixture of Ze/7-butyl /V-[6-(benzyloxy)-6,19bis(trifluoromethyl)-17,23-dioxa-3,4,22-triazatetracyclo[ 16.3.1.12,5.011,16]tricosa1(22),2,4,8,11(16),12,14,18,20-nonaen-21-yl]carbamate (£7Z mixture) (120 mg, 0.1811 mmol) and Zeri-butyl 7V-[6-(2-allylphenoxy)-2-[5-[l-hydroxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-5-(trifluoromethyl)-3-pyridyl]carbamate (120 mg, 0.1998 mmol) in AcOH (5 mL) was added Pd/C (42 mg of 10 % w/w, 0.03947 mmol) in a round-bottomed flask equipped with a H2 balloon using a 3-way adaptor. Subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. Filled the flask with hydrogen gas then stirred the mixture for 15 hours. Subjected to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. Filtrate was concentrated and dried under high vacuum. The résultant brown residue was purified by silica gel column chromatography using a shallow gradient of 100% hexanes to 30% EtOAc-hexanes to afford ZerZ-butyl 7V-[6-hydroxy-6,19bis(trifluoromethyl)-17,23 -dioxa-3,4,22-triazatetracyclo[ 16.3.1.12,5.011,16]tricosa1(22),2,4,11(16),12,14,18,20-octaen-21-yl]carbamate (14.2 mg, 27%); ESI-MS m/z cale. 574.1651, found 575.2 (M+l)⁺; Rétention time: 1.31 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle)

248 made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60

Step 7: 21-Amino-6,19-bis(trifluoromethyl)-17,23-dîoxa-3,4,22triazatetracyclo[16.3.1.12,5.011,16]tricosa-l(22),2,4,ll(16),12,14,18,20-octaen-6-oI (hydrochloride sait), Compound 17

To a solution of Zeri-butyl JV-[6-hydroxy-6,19-bis(trifIuoromethyl)-17,23-dioxa-3,4,22triazatetracyclo[16.3.1.12,5.011,16]tricosa-l(22),2,4,ll(16),12,14,18,20-octaen-21-yl]carbamate (13 mg, 0.02263 mmol) was added TFA (100 pL, 1.298 mmol) and DCM (400 pL) (pre made solution of 1: 4 TFA-DCM) and the reaction was stirred at room température for about 1 h. LCMS shows the completion of reaction. Solvents were removed and dissolved in DMSO (1 mL) and the residue was purified by a reverse phase HPLC-MS method using a dual gradient run from 50 - 99% mobile phase B over 15.0 minutes (mobile phase A = H2O (5 mM HCl); mobile phase B = CH3CN) to afford 2l-amino-6,19-bis(trifluoromethyl)-17,23-dioxa-3,4,22triazatetracyclo[ 16.3.1.12,5.011,16] tricosa-1 (22),2,4,11(16),12,14,18,20-octaen-6-ol (Hydrochloride sait) (2.6 mg, 22%) as an off-white amorphous solid. 'H NMR (400 MHz, Chloroform-d) δ 7.49 (s, 1H), 7.35 - 7.33 (m, 1H), 7.21 - 7.14 (m, 3H), 5.36 (s, 2H), 3.63 (s, 1H), 3.06 - 2.89 (m, 2H), 2.29 (m, 1H), 2.22 - 2.14 (m, 1H), 2.13 - 2.05 (m, 2H), 1.92 (d, J= 5.2 Hz, 2H) ppm. ESI-MS m/z cale. 474.11267, found 475.13 (M+l)⁺; Rétention time: 2.76 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = HzO (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

249

Example 14: Préparation of (15Æ)-20-amino-8-fluoro-15-methyI-6,18-bis(trifluoromethyl)16,23-dioxa-3,4,21-triazatetracyclo[15.3.1.12,5.17,ll]tricosa-l(21),2,4,7(22),8,10,17,19octaen-6-ol (diastereomer pair), Compound 18

diastereomer pair

Step 1: 3-[bis(tert-Butoxycarbonyl)amino]-6-[(172)-l-methylbut-3-enoxy]-5(trifluoromethyl)pyridine-2-carboxylic acid

To a solution of methyl 3-[bis(/erZ-butoxycarbonyl)amino]-6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (500 mg, 1.001 mmol) in DMSO (2 mL) was added (27î)-pent-4-en-2-ol (160 pL, 1.555 mmol), césium carbonate (504 mg, 1.547 mmol) and iodocopper (47 mg, 0.2468 mmol) and the reaction mixture was heated at 100 °C for 3 h. The reaction mixture was cooled to RT and poured on crushed ice and extracted with ethyl acetate and washed with brine. The organics were separated, dried over sodium sulfate, and evaporated. The résultant brown residue was purified by silica gel column chromatography using 100% hexanes to 30% EtOAc-hexanes to afford 3-[bis(Zert-butoxycarbonyl)amino]-6-[(17?)-lmethylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carboxylic acid (64 mg, 26%). ’H NMR (400 MHz, Chloroform-d) δ 10.17 (s, 1H), 7.63 (s, 1H), 5.73 (ddt, J= 17.2,10.2, 7.1 Hz, 1H), 5.20 (p, J= 6.4 Hz, 1H), 5.17 - 5.07 (m, 2H), 2.48 (dt, J= 13.5, 6.6 Hz, 1H), 2.37 (dt, J= 14.1, 6.9 Hz, 1H), 1.45 (d, J= 3.0 Hz, 18H), 1.34 (d, J= 6.3 Hz, 3H) ppm. ESI-MS m/z cale. 490.1927, found 250

391.2 (M-Boc)⁺. Rétention time: 0.5 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 30 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B — acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: tert-Butyl A-ZerZ-butoxycarbonyI-A-|6-[(17?)-l-niethylbut-3-cnoxy]-2-[5-[2,2,2triiluoro-l-(2-fluoro-5-iodo-phenyI)-l-hydroxy-ethyI]-l,3,4-oxadiazol-2-yl]-5(trifIuoromethyI)-3-pyridyl]carbamate

To a pre-heated at 50 °C stirred solution of 3-[bis(ZerZ-butoxycarbonyl)amino]-6-[(17?)-lmethylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carboxylic acid (120 mg, 0.2447 mmol) and 2,2,2-trifluoro-l-(2-fluoro-5-iodo-phenyl)ethanone (121 mg, 0.3805 mmol) in DMF (2 mL) was added (isocyanoimino)triphenylphosphorane (113 mg, 0.3738 mmol) at once. The mixture was stirred at room température ovemight then it was diluted with EtOAc (50 mL), washed with water and brine consecutively, then dried over sodium sulfate and filtered. The filtrate was concentrated to dryness. The résultant brown residue was purified by silica gel column chromatography using 100% hexanes to 50% EtOAc-hexanes to provide a light brown viscous oil, ZerZ-butyl A-ZerZ-butoxycarbonyl-7V-[6-[(17?)-l-methylbut-3-enoxy]-2-[5-[2,2,2-trifluoro-l(2-fluoro-5-iodo-phenyl)-l-hydroxy-ethyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3pyridyl]carbamate (103 mg, 51%). ^lH NMR (400 MHz, Chloroform-d) δ 8.17 (d, J= 2.2 Hz, 1H), 7.81 (s, 1H), 7.76 (ddd, 7= 8.6, 4.6, 2.2 Hz, 1H), 6.83 (ddd, 7= 10.4, 8.7, 1.2 Hz, 1H), 5.79 (ddtd, 7= 17.4, 10.9, 7.1,4.0 Hz, 1H), 5.38 (hept, 7= 6.2 Hz, 1H), 5.14-5.01 (m, 2H), 4.84 (d,7 = 3.9 Hz, 1H), 2.53 (dtd, 7= 13.7, 6.7, 2.9 Hz, 1H), 2.42 (dt, 7= 13.7, 6.7 Hz, 1H), 1.66 (s, 3H), 1.38 (dd, 7= 4.1, 2.1 Hz, 18H) ppm. ESI-MS m/z cale. 832.12036, found 733.1 (M-Boc)⁺. Rétention time: 0.79 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 30 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

251

Step 3: tert-Butyl A-[(15/î)-8-fluoro-6-hydroxy-15-methyl-6,18-bis(trifluoromethyl)-16,23dioxa-3,4,21-triazatetracyclo[15.3.1.12,5.17,ll]tricosa-l(21),2,4,7(22),8,10,12,17,19-nonaen20-yl]carbamate (E!Z mixture)

To a stirred solution of teri-butyl zV-Zer/-butoxycarbonyl-7V-[6-[(17?)-l-methylbut-3enoxy]-2-[5-[2,2,2-trifluoro-l-(2-fluoro-5-iodo-phenyl)-l-hydroxy-ethyl]-l,3,4-oxadiazol-2-yl]5-(trifluoromethyl)-3-pyridyl]carbamate (100 mg, 0.1201 mmol) in acetonitrile (10 mL) was added Palladium (II) acetate (8 mg, 0.03563 mmol) followed by tris-o-tolylphosphane (21 mg, 0.06900 mmol) and triethylamine (60 pL, 0.4305 mmol) and the solution was bubbled with N₂for 1 min then heated by microwave irradiation at 120 °C for 0.5 h. The mixture was cooled to room température then diluted with EtOAc and washed with saturated aqueous NH4CI (IX) and brine (IX) then dried over sodium sulfate, fïltered and concentrated to a yellow oil. The resulting material was dissolved in DMSO, fïltered and purified using a reverse phase HPLC-MS method using a Luna Cis column (75 x 30 mm, 5 pm particle size) sold by Phenomenex (pn: 00C-4252-U0-AX), and a dual gradient run from 50 - 99% mobile phase B over 15.0 minutes (mobile phase A = H₂O (5 mM HCl), mobile phase B = acetonitrile, flow rate — 50 mL/min, injection volume = 950 pL and column température = 25 °C giving a yellow solid, ZerZ-butyl N[(15/?)-8-fluoro-6-hydroxy-15-methyl-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.17,ll]tricosa-l(21),2,4,7(22),8,10,12,17,19-nonaen-20-yl]carbamate (E/Z mixture) (21 mg, 29%). ESI-MS m/z cale. 604.15564, found 605.03 (M+l)⁺; Rétention time: 0.55 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF₃CO₂H). Mobile phase B = acetonitrile (0.035 % CF₃CO₂H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

252

Step 4: (15Æ)-20-Amino-8-fluoro-15-methyl-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.17,ll]tricosa-l(21),2,4,7(22),8,10,17,19-octaen-6-ol (diastereomer pair), Compound 18

diastereomer pair

Part 1: To a solution of /errtbutyl 7V-[(152î)-8-fluoro-6-hydroxy-15-methyl-6,18bis(trifluoromethyl)-16,23-dioxa-3,4,21 -triazatetracyclof 15.3.1.12,5.17,1 1 ] tricosal(21),2,4,7(22),8,10,12,17,19-nonaen-20-yl]carbamate (E/Zmixture) (20 mg, 0.03309 mmol) in éthanol (2 mL) was added Pd/C (18 mg of 10 % w/w, 0.01691 mmol) in a round bottom flask equipped with a Fb balloon using a 3-way adaptor. The mixture was subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. The flask was filled with hydrogen gas then stirred the mixture for 15 hours. The mixture was subjected to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated and dried under high vacuum.

Part 2: The material from Step 1 was dissolved in TFA (100 pL, 1.298 mmol) and DCM (400 pL) (pre-made solution of 1: 4 TFA-DCM) and the reaction was stirred at room température for about Ih. Solvents were removed and dissolved in DMSO (1 mL) and the residue was purifîed by a reverse phase HPLC-MS method using a dual gradient run from 30 99% mobile phase B over 15.0 minutes using a mobile phase A = H2O (5 mM HCl) and a mobile phase B = acetonitrile to afford (15A)-20-amino-8-fluoro-15-methyl-6,18-bis(trifluoromethyl)16,23-dioxa-3,4,21-triazatetracyclo[15.3.1.12,5.17,ll]tricosa-l(21),2,4,7(22),8,10,17,19-octaen6-0I (2 mg, 12%) as diastereomeric mixture. ESI-MS m/z cale. 506.1189, found 507.1 (M+l)⁺; Rétention time: 2.93 minutes; Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Example 15: Préparation of (6Æ,13S)-17-Amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa13-thionia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 19, and (6Æ,13/î)-17-aniino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa-13-

253 thionia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 20

Step 1

Step 1: (6R,135)-17-Amino-13-oxido-6,15-bis(trifluoromethyI)-19-oxa-13-thionia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 19, and (6R,13R)-17-amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa-13-thionia-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 20

To a solution of (67î)-17-amino-6,15-bis(trifluoromethyl)-19-oxa-13-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (56 mg, 0.127 mmol) in EtOAc (1.5 mL) cooled by ice bath was added 3-chlorobenzenecarboperoxoic acid (802 pL of 0.11 M, 0.088 mmol) as a solution in EtOAc and the mixture stirred at 0 °C for 15 min. Then 3chlorobenzenecarboperoxoic acid (227 pL of 0.11 M, 0.025 mmol) as a solution in EtOAc was added and the mixture was stirred at 0 °C for 15 additional minutes. Then more 3chlorobenzenecarboperoxoic acid (85 pL of 0.11 M, 0.0094 mmol) as a solution in EtOAc was added and the mixture stirred at 0 °C for 15 min. Then the mixture was diluted with EtOAc (30 mL) and MeOH (1 mL) and washed with 5% NaiSiOs, 1 M NaHCOj, dried (MgSO4) and evaporated. The residue was purified by silica gel chromatography (12 g S1O2, 10-50% of a solution (10% MeOH in EtOAc) to hexanes over 20 min) eluted first 2.3 mgs (4%) of recovered starting material (6R)-17-amino-6,15-bis(trifluoromethyl)-l9-oxa-l3-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol. Continued elution provided (6R, 13 R)-17-amino-13 -oxido-6,15-bis(trifluoromethyl)- 19-oxa-13 -thionia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (2.6 mg, 4%). Continued elution provided (6R, 135)-17-amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa- 13-thionia3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol and then a mixture of (6R, 135)-17-amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa-13-thionia-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16-pentaen-6-ol and (6R, 13R)~ 17-amino-13oxido-6,15-bis(trifluoromethyl)-19-oxa-l 3-thionia-3,4,18-triazatricyclo[l 2.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-6-ol.

254

The mixture of (6R,135)-17-amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa-13thionia-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-6-ol and (6R, 13R)17-amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa-13-thionia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol was dissolved in 1:1 MeOH/acetonitrile and subjected to préparative HPLC eluting with 30-70% acetonitrile vs 5 mM HCl in water at 50 mL/min over 14 min through a Luna 5 μΜ Cis 100Â 75 X 30 mm column to provide the first eluent as a white solid (6R,13R)-17-amino-13-oxido-6,15-bis(trifluoromethyl)19-oxa-13-thionia-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-6-ol (18 mg, 31%): Ή NMR (400 MHz, Methanol-d₄) δ 7.82 (s, 1H), 3.80 (td, J= 12.5, 5.1 Hz, 1H), 3.15 (td, J= 12.4, 3.8 Hz, 1H), 2.37 (td, J= 13.0, 11.5,3.3 Hz, 1H), 2.29-2.17 (m, 1H), 2.091.97 (m, 1H), 1.95 - 1.84 (m, 1H), 1.83 - 1.65 (m, 4H), 1.59 (t, J= 8.6 Hz, 2H) ppm; ¹⁹FNMR (376 MHz, Methanol-d4) δ -60.52, -80.84 ppm; ESI-MS m/z cale. 458.08472, found 459.1 (M+l)⁺; Rétention time: 0.94 minutes. Final purity was determined by reversed phase HPLCMS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =12 mL/min, injection volume = 50 pL, and column température = 25 °C. Continued elution provided second eluent (6R,135)-17-amino-13-oxido-6,15-bis(trifluoromethyl)-19-oxa-13thionia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (19 mg, 33%): *H NMR (400 MHz, Methanol-d4) δ 7.81 (s, 1H), 3.75 (td, J= 12.7,4.0 Hz, 1H), 3.28 - 3.17 (m, 1H), 2.48 - 2.27 (m, 3H), 2.25 - 2.11 (m, 1H), 1.72 (dddd, J= 32.5, 20.0, 13.7, 8.0 Hz, 6H) ppm; ¹⁹F NMR (376 MHz, Methanol-d₄) δ -60.41, -80.81; ESI-MS m/z cale. 458.08472, found 459.0 (M+l)⁺; Rétention time: 1.09 minutes. Final purity was determined by reversed phase HPLCMS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Step 2: Solid form chracterization of crystalline Compound 19 (neat form)

Compound 19(15 mg) was dissolved in 0.45 mL of methanol. The solution was allowed to sit at room température for 2 hours. Cubes with slightly opaque faces formed.

Single crystals of crystalline Compound 19 (neat form) were grown by slow cooling a methanol solution from 80 °C to 25 °C. X-ray diffraction data were acquired at 100 K on a Bruker diffractometer equipped with Mo Ka radiation (λ=0.71073 Â) and a CCD detector. The

255 structure was solved and refined using SHELX programs (Sheldrick, G.M., Acta Cryst., (2008)

A64, 112-122). The results are summarized in Table 10 below.

Table 10: Single crystal élucidation of crystalline Compound 19 (neat form)

Crystal System	Tetragonal
Space Group	P41212
a(Â)	9.8237(4)
b(Â)	9.8237(4)
c(Â)	37.0548(18)
a(°)	90
β(°)	90
y(°)	90
V(Â³)	3576.0(3)
Z/Z'	8/1
Température	100 K

Step 3: Solid form chracterization of crystalline Compound 20 (neat form)

Compound 20 (10 mg) was dissolved in 0.3 mL of methanol. The solution was heated, then cooled to room température over 2 hours. Clear rectangular prisms were obtained.

Single crystals crystalline Compound 20 (neat form) were grown by slow cooling of a 10 methanol solution. X-ray diffraction data were acquired at 100 K on a Bruker diffractometer equipped with Mo Ka radiation (λ=0.71073 Â) and a CCD detector. The structure was solved and refined using SHELX programs (Sheldrick, G.M., Acta Cryst., (2008) A64, 112-122) and results are summarized in Table 11 below.

Table 11: Single crystal élucidation of crystalline Compound 20 (neat form) ____________________

Crystal System	Orthorhombic
Space Group	P2i2i2i
a(Â)	10.6547(4)
b(Â)	13.7046(5)
c(Â)	25.5376(11)
a(°)	90
β(°)	90
y(°)	90
V(Â³)	3729.0(3)
Z/Z'	8/1
Température	100 K

256

Example 16: (6Æ)-17-Amino-6,15-bis(trifluoromethyl)-19-oxa-13-thia-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 21

Step 5

Step 1: l-But-3-enylsulfany!sulfonyl-4-methyl-benzene

A mixture of l-methyl-4-sulfidosulfonyl-benzene (potassium sait) (2000 mg, 8.836 mmol), and 4-iodobut-l-ene (913 pL, 8.026 mmol) in DMF (20 mL) was stirred at 60 °C for 30 min, then diluted with ether and washed with water (2X) and brine, dried (MgSCU) and evaporated. The residue was purifïed by silica gel chromatography (0-15% EtOAc in hexanes 10 over 15 min) to provide l-but-3-enylsulfanylsulfonyl-4-methyl-benzene (1.516 g, 78%). ’H

NMR (400 MHz, Chloroform-d) δ 7.87 - 7.78 (m, 2H), 7.35 (d, J= 8.0 Hz, 2H), 5.68 (ddt, J= 17.0, 10.4, 6.7 Hz, 1H), 5.06 - 4.98 (m, 2H), 3.05 (t, J= 7.3 Hz, 2H), 2.46 (s, 3H), 2.40 - 2.29 (m, 2H) ppm. ESI-MS m/z cale. 242.04352, Rétention time: 0.6 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 15 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 1 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température - 60 °C.

257

Step 2: te/7-Butyl A-[2-|5-[(U?)-l-benzyIoxy-l-(trifluoromethyI)pent-4-enyI]-l,3,4oxadiazol-2-yl]-6-but-3-enylsuIfanyl-5-(trifluoromethyI)-3-pyridyl]carbamate

To terZ-butyl TV- [2-[5-[1-benzyloxy-1 -(trifluoromethyl)pent-4-enyl]-1,3,4-oxadiazol-2yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (57 mg, 0.08750 mmol) in ether (1.15 mL) at -78 °C was added n-BuLi (76 pL of 2.5 M, 0.1900 mmol) as a solution in hexanes and the mixture stirred at -78 °C for 15 min, then a solution of l-but-3-enylsulfanylsulfonyl-4-methylbenzene (28 mg, 0.1155 mmol) in ether (285 pL) was added dropwise. The mixture was stirred at -78 °C for 15 min and at 0 °C for 15 min. The mixture was diluted with 1 M NH4CI in water and ether then partitioned. The organic layer was separated and washed with 1 M NaHCCL, dried (MgSCU) and evaporated. The residue was purified by silica gel chromatography (0-10% EtOAc in hexanes over 15 min) provided teri-butyl 7V-[2-[5-[(17î)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-but-3-enylsulfanyl-5-(trifluoromethyl)-3pyridyl]carbamate (33 mg, 57%). *H NMR (400 MHz, Chloroform-d) δ 9.95 (s, 1H), 9.18 (s, 1H), 7.45 - 7.28 (m, 5H), 5.94 - 5.65 (m, 2H), 5.13 - 4.97 (m, 4H), 4.84 (d, J= 10.8 Hz, 1H), 4.68 (d, J= 10.9 Hz, 1H), 3.36 - 3.27 (m, 2H), 2.62 - 2.16 (m, 6H), 1.56 (s, 9H) ppm; ^I9F NMR (376 MHz, Chloroform-d) δ -63.55, -72.90 ppm. ESI-MS m/z cale. 658.20483, found 659.3 (M+l)⁺; Rétention time: 2.09 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (50 ^χ 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 3: teri-Butyl A-[(6J?)-6-benzyloxy-6,15-bis(trifluoromethyl)-19-oxa-13-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]carbamate (E/Z mixture)

A solution of terrtbutyl 7V-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-but-3-enylsulfanyl-5-(trifluoromethyl)-3-pyridyl]carbamate (209 mg, 0.3173 258 mmol) in DCE (15 mL) was dropwise added to a 80 °C preheated solution of benzylidene-[l,3bis(2,4,6-trimethylphenyl)imidazolidin-2-ylidene]-dichloro-ruthenium;tricyclohexylphosphane (40 mg, 0.04712 mmol) in DCE (15 mL) and the resulting mixture heated at 80 °C for 45 min. Then the solvent was evaporated. The residue was purified by silica gel chromatography (0-5% EtOAc in hexanes over 15 min) to provide ZerZ-butyl jV-[(67?)-6-benzyloxy-6,15bis(trifluoromethyl)-l 9-oxa-l 3-thia-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,9,14(18),15-hexaen-17-yl]carbamate (E/Zmixture) (158 mg, 79%). ESI-MS m/z cale. 630.1735, found 631.2 (M+l)⁺; Rétention time: 0.67 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 4: ZerZ-Butyl A-[(67?)-6-benzyloxy-6,15-bis(trifIuoromethyl)-19-oxa-13-thia-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate

A mixture of ZerZ-butyl 7V-[(6R)-6-benzyloxy-6,15-bis(trifluoromethyl)-19-oxa-13-thia3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaen-17-yl]carbamate (E/Z mixture) (158 mg, 0.2506 mmol) and Pd/C (50 mg of 10 % w/w, 0.04698 mmol) in EtOAc (800 pL) and MeOH (800 pL) was stirred at room température under 200 psi H2 in a stainless Steel pressure vessel for 23 h. Then the mixture was filtered and the filtrate evaporated to provide Ze/7-butyl 7V-[(6Z?)-6-benzyloxy-6,15-bis(trifluoromethyl)-l 9-oxa-l 3-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (160 mg, 100%). Ή NMR (400 MHz, Chloroform-d) δ 9.70 (s, 1H), 9.14 (s, 1H), 7.33 - 7.22 (m, 5H), 4.86 (d, J= 11.2 Hz, 1H), 4.79 (d, J= 11.2 Hz, 1H),3.O6 (ddt, J= 13.8, 9.5, 4.8 Hz, 2H), 2.44 (dt, J= 15.8, 8.5 Hz, 1H), 2.27 (dt, J= 14.5, 7.0 Hz, 1H), 2.04 (s, 1H), 1.97 - 1.84 (m, 1H), 1.78 (dq,J=13.7, 6.7 Hz, 2H), 1.64 (dt, J = 12.8, 6.7 Hz, 1H), 1.59-1.51 (m, 13H)ppm; ¹⁹FNMR (376 MHz, Chloroform-d) δ -63.64, -74.38 ppm. ESI-MS m/z cale. 632.1892, found 633.3 (M+l)⁺; Rétention time: 0.7 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile

259 phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 5: (67?)-17-Amino-6,15-bis(trifluoromethyl)-19-oxa-13-thia-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI, Compound 21

To a solution of Ze/7-butyl ;V-[(6/?)-6-benzyloxy-6,15-bis(trifluoromcthyl)-19-oxa-13thia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (57 mg, 0.090 mmol) in DCM (2 mL) at 0 °C was added BCI3 (2 mL of 1 M, 2.00 mmol) as a solution in DCM and the mixture stirred at room température for 22 h. Then the mixture was evaporated, and the residue dissolved in EtOAc (70 mL) and methanol (0.5 mL) and washed with 1 M NaHCO3, brine, dried (MgSÜ4) and evaporated. The residue was purified by silica gel chromatography (0-25% EtOAc in hexanes over 15 min) to provide (67?)-17-amino-6,15bis(trifluoromethyl)-19-oxa-13-thia-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (61 mg, 62%). ^!H NMR (400 MHz, Methanol-d4) δ 7.62 (s, 1H), 3.16 (ddd, J= 13.7, 12.1, 4.2 Hz, 1H), 2.97 (td, J= 13.2, 4.1 Hz, 1H), 2.35 (ddd, J= 14.4, 11.2, 3.3 Hz, 1H), 2.20 (dddt, J= 19.8, 14.3, 10.6, 5.1 Hz, 2H), 1.83 - 1.50 (m, 7H) ppm; ¹⁹FNMR(376 MHz, Methanol-ck) δ -64.96, -80.84 ppm. ESI-MS m/z cale. 442.0898, found 443.0 (M+l)⁺; Rétention time: 1.61 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =12 mL/min, injection volume = 50 pL, and column température = 25 °C.

260

Example 17: (6jR)-16-amino-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol, Compound 22

Step2

Step 3

Step 1: tert-Butyl A-|2-[5-[(lZ?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyI]-l,3,4-oxadiazol 2-yl]-6-but-3-enylsulfanyl-5-(trifluoromethyI)-3-pyridyl]carbamate

To ZerZ-butyl 7V-[2-[5-[( 1 R)-1 -benzyloxy-1 -(trifluoromethyl)but-3 -enyl] -1,3,4-oxadiazol2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (96 mg, 0.1506 mmol) in ether (1.8 mL) at -78 °C was added n-BuLi (130 pL of 2.5 M, 0.3250 mmol) as a solution in hexanes, then a solution of l-but-3-enylsulfanylsulfonyl-4-methyl-benzene (48 mg, 0.1981 mmol) in ether (500 pL) was added dropwise. The mixture was stirred at -78 °C for 15 min and at 0 °C for 15 min. The mixture was diluted with 1 M NH4CI in water and ether then partitioned. The organic layer was separated and washed with 1 M NaHCCfi, dried (MgSCL) and evaporated. The residue was purified by silica gel chromatography (0-5% EtOAc in hexanes over 15 min) to provide tertbutyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-but-3enylsulfanyl-5-(trifluoromethyl)-3-pyridyl]carbamate (42 mg, 43%). 'H NMR (400 MHz, Chloroform-d) δ 9.94 (s, 1H), 9.17 (s, 1H), 7.40 - 7.28 (m, 5H), 6.09 - 5.76 (m, 2H), 5.25 (dd, J= 17.0, 1.5 Hz, 1H), 5.22 - 5.18 (m, 1H), 5.07 (dd, J= 17.2, 1.7 Hz, 1H), 5.01 (dd, J= 10.2, 1.6 Hz, 1H), 4.83 (d, J= 10.9 Hz, 1H), 4.68 (d, J= 10.8 Hz, 1H), 3.31 (td, J= 7.1, 2.8 Hz, 2H), 3.21 (t, J = 6.4 Hz, 2H), 2.48 (q, J= 7.0 Hz, 2H), 1.56 (s, 9H) ppm; ¹⁹F NMR (376 MHz, Chloroformd) δ -63.56, -73.16 ppm. ESI-MS m/z cale. 644.1892, found 645.2 (M+l)⁺; Rétention time: 0.7

261 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = HzO (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: terf-Butyl 7V-[(6J?)-6-benzyloxy-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,8,13,15-hexaen-16-yl]carbamate (JE/Z mixture)

A solution of ierZ-butyl7V-[2-[5-[(lJ?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-but-3-enylsulfanyl-5-(trifluoromethyl)-3-pyridyl]carbamate (140 mg, 0.2172 mmol) in toluene (10 mL) was added dropwise to a 120 °C preheated solution of l,3-bis(o-tolyl)4,5-dihydroimidazole;dichloro-[(2-isopropoxyphenyl)methylene]ruthenium (19 mg, 0.03330 mmol) in toluene (10 mL) and the mixture heated at 120 °C for 45 min. Then more l,3-bis(otolyl)-4,5-dihydroimidazole;dichloro-[(2-isopropoxyphenyl)methylene]ruthenium ( 12 mg, 0.02103 mmol) was added and heating continued at 120 °C for 55 min and this process was carried out onemore timeprecisely. Thenmore l,3-bis(o-tolyl)-4,5-dihydroimidazole;dichloro[(2-isopropoxyphenyl)methylene]ruthenium (2.7 mg, 0.004733 mmol) was added and heating at 120 °C continued for 60 min and this process was carried out two more times precisely. The solvent was evaporated and the residue purified by silica gel chromatography (12 g S1O2, 0-30% of a solution of 10% EtOAc in hexanes from 100% hexanes over 20 min) to provide ZerZ-butyl N[(6/?)-6-benzyloxy-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17triazatricyclofl 1.3.1.12,5]octadeca-l(16),2,4,8,13(17),14-hexaen-16-yl]carbamate (E/Zmixture) (20 mg, 13%). ESI-MS m/z cale. 616.1579, found 617.1 (M+l)⁺; Rétention time: 0.62 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

262

Step 3: ZerZ-Butyl A-[(67?)-6-benzyIoxy-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17 triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-16-yl]carbamate

A mixture of ZerZ-butyl 7V-[(6R)-6-benzyloxy-6,14-bis(trifluoromethyl)-18-oxa-12-thia3,4,17-triazatricyclo[l 1.3.1.12,5]octadeca-l(16),2,4,8,13(17),14-hexaen-16-yl]carbamate (E/Z mixture) (24 mg, 0.03309 mmol), and palladium on carbon (11 mg of 10 % w/w, 0.01034 mmol) in EtOAc (300 pL) and MeOH (300 pL) was stirred at room température under 200 psi H2 for 40 h, filtered and solvent evaporated. The residue was purified by silica gel chromatography (030% of a solution of 10% EtOAc in hexanes from 100% hexanes over 18 min) to provide tertbutyl 7V-[(6Æ)-6-benzyloxy-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17triazatricyclofl 1.3.1.12,5]octadeca-l(16),2,4,13(17),14-pentaen-16-yl]carbamate (16 mg, 78%). ESI-MS m/z cale. 618.1735, found 619.2 (M+l)⁺; Rétention time: 0.64 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 4: (67î)-16-Amino-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17triazatricycIo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol, Compound 22

nh₂ n-n

To a solution of ZerZ-butyl N-[(6R)-6-benzyloxy-6,14-bis(trifluoromethyl)-18-oxa-12thia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(16),2,4,13(17),14-pentaen-16-yl]carbamate (16 mg, 0.02587 mmol) in DCM (160 pL) at 0 °C was added BCI3 (520 pL of 1 M, 0.5200 mmol) as a solution in DCM and the mixture stirred at room température for 24 h. Then the mixture was evaporated and the residue dissolved in EtOAc (20 mL) and methanol (2 mL) and washed with 1 M NaHCO3, brine, dried (MgSO.4) and evaporated. The residue was purified by silica gel chromatography (5-25% EtOAc in hexanes over 15 min) to provide (6R)-16-amino-6,14bis(trifluoromethyl)-18-oxa-12-thia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15263 pentaen-6-ol (4.5 mg, 41 %). *H NMR (400 MHz, Methanol-cU) δ 7.47 (s, 1H), 3.05 (td, J= 13.1, 4.3 Hz, 1H), 2.87 (td, J = 13.6, 13.1, 4.5 Hz, 1H), 2.25 - 2.00 (m, 3H), 1.91 - 1.84 (m, 1H), 1.79 (dq, J= 10.9, 6.3, 5.9 Hz, 2H), 1.50 (p, J= 5.5 Hz, 2H) ppm; ^,9F NMR (376 MHz, Methanol-cU) δ -64.87, -80.28ppm. ESI-MS m/z cale. 428.07416, found 429.0 (M+l)⁺;

Rétention time: 1.49 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF3CO₂H). Mobile phase B = acetonitrile (0.035 % CF3CO₂H). Flow rate =12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 18: Préparation of (6E)-16-amino-12-oxido-6,14-bis(trifIuoromethyl)-18-oxa-12thionia-3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (enantiomer 1), Compound 23, and (6E)-16-amino-12-oxido-6,14-bis(trifluoromethyl)-18-oxa-12-thionia3,4,17-triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (enantiomer 2), Compound 24

Step 1

enantiomer 1

enantiomer 2

Step 1: (6/î)-16-Amino-12-oxido-6,14-bis(trifluoromethyI)-18-oxa-12-thionia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (enantiomer 1), Compound 23, and (6/î)-16-amino-12-oxido-6,14-bis(trifluoromethyl)-18-oxa-12-thionia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-oI (enantiomer 2), Compound 24

enantiomer 1

enantiomer 2

To a solution of (67?)-16-amino-6,14-bis(trifluoromethyl)-18-oxa-12-thia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (3.7 mg, 0.0086 mmol) in EtOAc (222.0 pL) cooled by ice bath was added 3-chlorobenzenecarboperoxoic acid (44 pL of 0.11 M, 0.0048 mmol) as a solution in EtOAc and the mixture stirred at 0 °C for 15 min. Then 3-chlorobenzenecarboperoxoic acid (20 pL of 0.11 M, 0.0022 mmol) as a solution in EtOAc was added and the mixture stirred at 0 °C for 15 additional minutes. Then more 3chlorobenzenecarboperoxoic acid (8 pL of 0.11 M, 0.00088 mmol) as a solution in EtOAc was added with the mixture stirred at 0 °C for 15 min. Then the mixture was diluted with EtOAc (30

264 mL) and MeOH (1 mL) and washed with 5% NasSzOj, 1 M NaHCCfi, dried (MgSCfi) and evaporated. The mixture residue was dissolved into 1:1 MeOH/acetonitrile and subjected to préparative HPLC eluting with 30 - 99% acetonitrile vs 5 mM HCl in water at 50 mL/min over 14 min through a Luna 5 μΜ Ci8 100Â 75X30 mm column to provide first eluting isomer as a white solid (67?)-16-amino-12-oxido-6,14-bis(trifluoromethyl)-18-oxa-12-thionia-3,4,17triazatricyclo[11.3.1.12,5]octadeca-l(17),2,4,13,15-pentaen-6-ol (enantiomer 1) (2 mg, 52%). *H NMR (400 MHz, Methanol-d₄) δ 7.77 (s, 1H), 3.83 (ddd, J= 13.1, 11.3, 5.5 Hz, 1H), 3.06 (ddd, J= 13.2, 11.1, 3.8 Hz, 1H), 2.38(dt,J= 15.3, 7.8 Hz, 2H), 2.14 - 2.04 (m, 1H), 2.01 - 1.87 (m, 3H), 1.85 - 1.67 (m, 2H) ppm; ¹⁹F NMR (376 MHz, Methanol-d₄) δ -60.67, -80.30 ppm. ESI-MS m/z cale. 444.0691, found 445.0 (M+l)⁺; Rétention time: 1.0 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CH0-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Continued elution provided as a white solid (67?)-16-amino-12-oxido-6,14bis(trifluoromethyl)-l 8-oxa-l 2-thionia-3,4,17-triazatricyclo[l 1.3.1.12,5]octadeca1 (17),2,4,13,15-pentaen-6-ol (enantiomer 2) (1.6 mg, 42%). *H NMR (400 MHz, Methanol-ck) δ 7.76 (s, 1H), 3.64 (td, J= 12.5, 5.0 Hz, 1H), 3.22 (dd, J= 13.1, 3.8 Hz, 1H), 2.43 (dp,./-9.1, 6.4, 4.3 Hz, 1H), 2.28 (t, J= 7.2 Hz, 2H), 2.09 - 1.87 (m, 2H), 1.75 (ddt, J= 34.2, 7.8, 3.7 Hz, 3H) ppm; ¹⁹F NMR (376 MHz, Methanol-tL) δ -60.71, -80.20 ppm. ESI-MS m/z cale. 444.0691, found 445.0 (M+l)⁺; Rétention time: 0.81 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 19: Préparation of (67?,127î)-17-amino-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-10-one (hydrochloride sait), Compound 25, and (67?,127?)-17-amino-6-hydroxy-12265 methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-9-one (hydrochloride sait), Compound 26

Step 1: tert-Butyl A-[(67?,127?)-6-benzyloxy-9-hydroxy-12-methyl-6,15-bis(trifluoromethyl)5 13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-y^-Λ^,terf-butoxycarbonyl-carbamate and ZerZ-butyl A^r-[(6/?,127?)-6-benzyloxy-10-hydroxy-12methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]-A^r-te/7-butoxycarbonyl-carbamate (mixture of regioisomeric diastereomers)

To a solution of ZerZ-butyl 7V-[(67?,127?)-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16hexaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate (E/Z mixture) (190 mg, 0.2607 mmol) in THF (3 mL) at 0 °C was added dropwise borane dimethylsulfide complex (200 pL of 2 M, 0.4000 mmol) and let the resulting mixture stir for 15 min at 0 °C. Allowed the reaction warm to room température and stirred for 1 h. Added additional borane dimethylsulfide complex (200 pL of 2

266

Μ, 0.4000 mmol) and stirred at room température for an additional 30 minutes. Cooled the reaction to 0 °C before quenching with aqueous NaOH (1.5 mL of 1 Μ, 1.500 mmol) followed by the addition of hydrogen peroxide (600 pL of 30 % w/v, 5.292 mmol). Allowed the resulting mixture stir for 30 min at room température then the mixture was extracted with ethyl acetate (2 X 80 mL). The organic layers were combined, washed with brine (80 mL), dried over sodium sulfate, filtered and concentrated. The crude residue was then purified by silica gel chromatography (80 gram column) using a gradient from 100% hexanes to 100% ethyl acetate to collect as a ~1:1 inséparable mixture of regioisomeric diastereomers, Ze/7-butyl jV-[(6/?,12/?)-6benzyloxy-9-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-17-yl]-7V-ter/-butoxycarbonylcarbamate and terZ-butyl 7V-[(6R,12R)-6-benzyloxy-10-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]-7V-tert-butoxycarbonyl-carbamate (mixture of regioisomeric diastereomers) (128 mg, 66%). ESI-MS m/z cale. 746.275, found 747.4 (M+l)⁺; Rétention time: 1.89 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: terAButyl A-[(6J?,12/?)-6-benzyloxy-12-methyl-9-oxo-6,15-bis(trifluoromcthyI)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-A^rte/7-butoxycarbonyl-carbamate and ZerZ-butyl A-[(67?,12/?)-6-benzyIoxy-12-methyl-10-oxo267

6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate

To a solution of a -1:1 mixture of regioisomeric diastereomers, ZerZ-butyl N-[(6R,T2R)-6benzyloxy-9-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16-pentaen-l 7-yl]-7V-ZerZ-butoxycarbonylcarbamate and ZerZ-butyl 7V-[(6Æ,12Æ)-6-benzyloxy-10-hydroxy-12-methyl-6,15bis(trifluorom ethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]-jV-ZerZ-butoxycarbonyl-carbamate (mixture of regioisomeric diastereomers) (106 mg, 0.142 mmol) in dichloromethane (2 mL) was added Dess-Martin Periodinane (92 mg, 0.2169 mmol) at 0 °C and the mixture was stirred for 1 h allowing the reaction to warm up to room température. The reaction was diluted with ether (10 mL) and filtered through Celite. The filtrate was washed with saturated aqueous sodium bicarbonate and with brine, dried over sodium sulfate, filtered and concentrated. The resulting material was dissolved in 2 mL of DMSO, filtered and the filtrate was purified by reverse phase HPLC using a gradient from 50% to 99% acetonitrile in water (+5 mM HCl) giving as an inséparable -1:1 mixture of regioisomers, ZerZ-butyl N-[(6R, 12/?)-6-benzyloxy-12-methyl-9-oxo-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-JV-ZerZbutoxycarbonyl-carbamate and ZerZ-butyl 7V-[(6R,12R)-6-benzyloxy-12-methyl-10-oxo-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate (44 mg, 42%). ESI-MS m/z cale. 744.2594, found 645.2 (M-Boc+1)⁺; Rétention time: 2.05 minutes. Final purity was determined by

268 reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 3: (6R,12R)-17-Amino-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-10-one, Compound 25, and (6R,12R)-17-amino-6-hydroxy-12-methyI-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-9-one, Compound 26

To a solution of a ~1:1 mixture of regioisomers, ZerZ-butyl //-[(ôÆ^Æj-ô-benzyloxy-^methyl-9-oxo-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate and ZerZ-butyl N-[(6R,12R)-6benzyloxy-12-methyl-l 0-oxo-6,l 5-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-17-yl]-JV-teri-butoxycarbonylcarbamate (34 mg, 0.0457 mmol) in éthanol (5 mL) was added Pd/C (116 mg of 10 % w/w, 0.1090 mmol) in a flask equipped with a hydrogen balloon using a 3-way adaptor. The material was subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. The flask was filled with hydrogen gas then stirred the mixture for 15 h. The vessel was subjected to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated and dried under reduced pressure. The resulting residue was dissolved in TFA (1.5 mL, 19.47 mmol) and dichloromethane (4.5 mL) (pre-made solution of 1:3 TFA/dichloromethane) and the reaction was stirred at room température for about Ih. The solvents were removed by rotary évaporation and the residue was

269 dissolved in DMSO (1 mL) then purified by reverse phase HPLC using a gradient from 30% to 99% acetonitrile in water (+5 mM HCl) to afford the first eluting regioisomer (based on H NMR AB pattern of -O CH (CH3)-CH2-CO) as (6R, 12/?)-17-amino-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-10-one (Hydrochloride sait) (2.3 mg, 41%). 'H NMR (400 MHz, Chloroform-d) δ 7.61 (s, 1H), 6.92 (dq, J= 16.0, 6.9 Hz, 1H), 6.13 (dq, J= 15.8, 1.7 Hz, 1H), 2.67 (t, J= 6.7 Hz, 2H), 2.46 (ddd, J= 15.1, 10.2, 5.7 Hz, 1H), 2.13 (ddd, J= 14.7, 10.3,5.0 Hz, 1H), 1.90 (dd, J= 6.9, 1.6 Hz, 3H), 1.83 (dd, J= 10.4, 4.8 Hz, 2H), 1.71-1.42 (m, 2H) ppm. ESI-MS m/z cale.

454.10757, found 455.1 (M+l)⁺ ; Rétention time: 1.34 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C and the later eluting regioisomer as (6/?,127?)-17-amino-6-hydroxy-12-mcthyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-9-one (Hydrochloride sait) (3.0 mg, 54%). ’H NMR (400 MHz, Chloroform-d) δ 7.41 (s, 1H), 6.27 (s, 1H), 5.42 - 5.35 (m, 1H), 2.99 (ddd, J= 18.3, 11.2,2.1 Hz, 1H), 2.89 - 2.80 (m, 1H), 2.79 - 2.63 (m, 2H), 2.52 (ddd, J= 18.1, 7.2, 2.2 Hz, 1H), 2.35 - 2.26 (m, 1H), 2.13 - 2.01 (m, 1H), 1.99 - 1.89 (m, 1H), 1.42 (d, J= 6.4 Hz, 3H) ppm. ESI-MS m/z cale. 454.10757, found 455.0 (M+l)⁺; Rétention time: 1.7 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Example 20: Préparation of20-amino-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-ol (enantiomer 1), Compound 27, 20-amino-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracycIo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-oI (enantiomer 2), Compound 28,19-amino-9-methyI-6,17-bis(trifluoromethyI)-15,22-dioxa3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaen-6-ol (diastereomer pair 1), Compound 29, and 19-amino-9-methyl-6,17-bis(trifluoromethyl)270

15,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20), 2,4,10(21), 11,13,16,18octaen-6-ol (diastereomer pair 2), Compound 30

Step 1: 2-[l-Benzyloxy-l-(trifluoromethyl)pent-4-enyI]-5-[6-(3-iodophenoxy)-3-nitro-5 5 (trifluoromethyl)-2-pyridyI]-l,3,4-oxadiazole

The reagent 3-iodophenol (30 mg, 0.1364 mmol) was added to a mixture of [6-[5-[lbenzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)-2pyridyl] trifluoromethanesulfonate (69 mg, 0.1061 mmol) and CS2CO3 (35 mg, 0.1074 mmol) in

DMF (1.4 mL) and was stirred at 0 °C for 1 h and then at room température for 2 h. The mixture was diluted with ether, washed with water (2X), brine, dried (MgSOfi and evaporated. The residue was purified by silica gel chromatography (12 g S1O2, 0-10% EtOAc in hexanes over 20 min) to provide 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(3-iodophenoxy)-3-nitro-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (51 mg, 67%). *H NMR (400 MHz, Chloroform15 d) δ 8.76 (s, 1H), 7.67 - 7.56 (m, 2H), 7.40 - 7.28 (m, 5H), 7.23 (ddd, J= 8.3, 2.3, 1.0 Hz, 1H), 7.12 (t, J= 8.0 Hz, 1H), 5.71 (ddt,J=16.7, 10.1,6.3 Hz, 1H), 5.00 (dd, J= 17.1, 1.6 Hz, 1H),

271

4.94 (dd, J= 10.3, 1.6 Hz, 1H), 4.77 (d, J= 10.6 Hz, 1H), 4.61 (d, J= 10.6 Hz, 1H), 2.55 - 2.25 (m, 3H), 2.24 - 2.11 (m, 1H) ppm; ^,9F NMR (376 MHz, Chloroform-d) δ -64.07, -73.02 ppm. ESI-MS m/z cale. 720.03046, found 721.0 (M+l)⁺; Rétention time: 0.56 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: (9£)-6-(Benzyloxy)-20-nitro-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,9,ll(22),12,14,17,19-nonaene and 6(benzyloxy)-9-methyIidene-19-nitro-6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaene (inséparable mixture of isomers)

inséparable mixture of isomers

A mixture of 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(3-iodophenoxy)-3nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (126 mg, 0.1749 mmol), palladium (II) acetate (10 mg, 0.04454 mmol), tris-o-tolylphosphane (27 mg, 0.08871 mmol) and triethylamine (51 pL, 0.3659 mmol) in acetonitrile (6.3 mL) was bubbled with N2 for 1 min then heated at 100 °C for Ih. The mixture was diluted with ether and washed with 1 M NH4CI, 1 M NaHCCh, brine then dried (MgSCU) and evaporated. The residue was purified by silica gel chromatography (24 g S1O2, 0-50% of a solution (20% EtOAc in hexanes) to hexanes over 20 min) to provide as a 2:1 inséparable mixture of isomers, (9/:)-6-(bcnzyloxy)-20-nitro-6,18-bis(trifluoromethyl)-16,23dioxa-3,4,21 -triazatetracyclof 15.3.1.12,5.111,15] tricosa-1 (21 ),2,4,9,11 (22), 12,14,17,19-nonaene (69 mg, 66% purity, 44%). ESI-MS m/z cale. 592.11816, found 593.1 (M+l)⁺; Rétention time: 0.55 minutes and 6-(benzyloxy)-9-methylidene-19-nitro-6,17-bis(trifluoromethyl)-15,22-dioxa3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),l 1,13,16,18-octaene (69 mg, 33% purity, 22%). ESI-MS m/z cale. 592.11816, found 593.1 (M+l)⁺; Rétention time: 0.54 minutes. Final purities were determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 %

272

Step 3: 20-Amino-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracycIo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-ol (enantiomer 1), Compound 27,20-amino-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-ol (enantiomer 2), Compound 28,19-amino-9-methyl-6,17-bis(trifluoromethyl)-15,22-dioxa3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaen-6-ol (diastereomer pair 1), Compound 29, and 19-amino-9-methyl-6,17-bis(trifluoromethyl)15,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18octaen-6-oI (diastereomer pair 2), Compound 30

inséparable mixture of isomers

enantiomer 1 enantiomer 2 diastereomer pair 1 diastereomer pair 2

A 2:1 inséparable mixture of isomers, (9£)-6-(benzyloxy)-20-nitro-6,18bis(trifluoromethyl)-16,23 -dioxa-3,4,21 -triazatetracyclo[ 15.3.1.12,5.111,15] tricosa1(21),2,4,9,1 l(22),12,14,17,19-nonaene (47 mg, 66% purity, 0.052 mmol) and 6-(benzyloxy)-9methylidene-19-nitro-6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaene (23 mg, 33% purity, 0.013 mmol) and Pd/C (38 mg of 10 % w/w, 0.03571 mmol) in EtOAc (1.3 mL) was stirred at room température under 200 psi H2 in a stainless Steel pressure vessel for 16 h. Then the mixture was fïltered and the filtrate evaporated. The residue was mixed with iron (50 mg, 0.8953 mmol), THF (0.5 mL), EtOH (0.25 mL) and HCl (125 pL of 4 M, 0.5000 mmol) as a solution in water at 60 °C for 30 min then diluted with EtOAc, fïltered and the filtrate washed with 1 M NaHCO3, dried (MgSO-i) and evaporated. The residue was purified by silica gel chromatography (24 g S1O2, 5-40% of a solution (20% EtOAc in hexanes) to hexanes over 20 min) to provide, eluting first, an inséparable mixture of 6-(benzyloxy)-6,18-bis(trifluoromethyl)

273

16,23-dioxa-3,4,21 -triazatetracyclo[ 15.3.1.12,5.111,15]tricosa-l (21 ),2,4,11 (22), 12,14,17,19octaen-20-amine (30 mg, 92%) and the first diastereomeric pair of isomers as 6-(benzyloxy)-9methyl-6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa1(20),2,4,10(21),11,13,16,18-octaen-19-amine (diastereomer pair 1) in an undetermined ratio. Continued elution provided the second diastereomeric pair isomers as 6-(benzyloxy)-9-methyl6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosal(20),2,4,10(21),ll,13,16,18-octaen-19-amine (diastereomer pair 2) (10 mg, 34%).

The inséparable mixture of 6-(benzyloxy)-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[ 15.3.1.12,5.111,15]tricosa-1 (21 ),2,4,11 (22), 12,14,17,19-octaen-20-amine and the first diastereomeric pair isomers of 6-(benzyloxy)-9-methyl-6,17-bis(trifluoromethyl)-15,22dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaen19-amine (diastereomer pair 1) was dissolved into TFA (0.3 mL) and water (0.015 mL) and heated at 60 °C for Ih, then diluted with EtOAc and washed with 1 M NaHCCL, dried and evaporated. The residue was purified by silica gel chromatography (12 g SiO₂, 5-40% EtOAc in hexanes over 20 min) to provide first, racemic 20-amino-6,18-bis(trifluoromethyl)-16,23-dioxa3,4,21-triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-ol, the séparation of which into single enantiomers is described below. ESI-MS m/z cale. 474.11267, found 475.2 (M+l)⁺; Rétention time: 0.71 minutes. Continued elution provided the first diastereomeric pair isomers of 19-amino-9-methyl-6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20triazatetracyclo[ 14.3.1.12,5.110,14] docosa-1 (20),2,4,10(21),11,13,16,18-octaen-6-ol (diastereomer pair 1) (1.4 mg, 6%). ’H NMR (400 MHz, Chloroform-d) δ 7.50 (s, 1H), 7.32 (t, J = 7.8 Hz, 1H), 7.23 (d, J= 2.0 Hz, 1H), 7.09 - 7.00 (m, 2H), 5.17 (s, 2H), 3.74 - 3.59 (m, 1H), 2.88 - 2.69 (m, 1H), 2.26 - 1.94 (m, 3H), 1.49 - 1.35 (m, 1H), 1.25 (d, J= 7.0 Hz, 3H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.36, -80.78 ppm. ESI-MS m/z cale. 474.11267, found 475.1 (M+l)⁺; Rétention time: 1.54 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Ci8 column (50 x 4.6 mm) sold by Phenomenex (pn: CH07644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

The racemic mixture of20-amino-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.11 l,15]tricosa-l(21),2,4,l l(22),12,14,17,19-octaen-6-ol obtained above was dissolved into 1:1 acetonitrile/MeOH and purified by préparative SFC eluting a gradient of 5 mM NH3 in methanol to CO2 (5-15% over 10 min at 60 mL/min) though a 21.2 X 250mm AS3 column, 5μιη particle, providing as the first enantiomer to elute, 20-amino-6,18

274 bis(trifluoromethyl)-16,23 -dioxa-3,4,21 -triazatetracyclo[ 15.3.1.12,5.111,15] tricosa1(21),2,4,11(22), 12,14,17,19-octaen-6-ol (enantiomer 1) (3.5 mg, 14%). *H NMR (400 MHz, Chloroform-d) δ 7.51 (s, 1H), 7.48 (d, J = 2.0 Hz, 1H), 7.31 (t, J= 7.8 Hz, 1H), 7.06 (d, J= 7.6 Hz, 1H), 6.98 (dd, J= 8.1, 2.3 Hz, 1H), 5.45 (s, 2H), 3.71 (s, 1H), 2.82 (ddd, J= 14.1, 6.7, 3.8 Hz, 1H), 2.70 (ddd, J= 13.7, 8.7, 4.0 Hz, 1H), 2.29 (ddd, 7= 14.3, 12.6, 5.1 Hz, 1H), 2.07 - 2.03 (m, 1H), 2.00 - 1.92 (m, 1H), 1.86 - 1.66 (m, 3H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ 63.64, -78.87 ppm. ESI-MS m/z cale. 474.11267, found 475.2 (M+l)⁺; Rétention time: 1.57 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Ci8 column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C and as the second enantiomer to elute, 20-amino-6,18bis(trifluoromethyl)-16,23 -dioxa-3,4,21 -triazatetracyclof 15.3.1.12,5.111,15] tricosal(21),2,4,ll(22),12,14,17,19-octaen-6-ol (enantiomer 2) (2.9 mg, 12%). ¹H NMR (400 MHz, Chloroform-d) δ 7.52 (s, 1H), 7.48 (d, J= 2.0 Hz, 1H), 7.31 (t, 7= 7.8 Hz, 1H), 7.06 (d, 7= 7.6 Hz, 1H), 7.02 - 6.95 (m, 1H), 5.45 (s, 2H), 3.69 (d, 7= 6.2 Hz, 1H), 2.82 (ddd, J= 14.0, 6.7, 3.8 Hz, 1H), 2.70 (ddd, 7= 13.7, 8.7, 4.0 Hz, 1H), 2.29 (td,7= 13.4, 12.9, 5.1 Hz, 1H), 2.08 - 1.91 (m, 3H), 1.87 - 1.64 (m, 2H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.64, -78.87 ppm. ESI-MS m/z cale. 474.11267, found 475.2 (M+l)⁺; Rétention time: 1.58 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =12 mL/min, injection volume = 50 pL, and column température = 25 °C.

The second diastereomeric pair of isomers described above, 6-(benzyloxy)-9-methyl6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20-triazatetracyclo[ 14.3.1.12,5.110,14]docosal(20),2,4,10(21),ll,13,16,18-octaen-19-amine (diastereomer pair 2) was dissolved into TFA (0.3 mL) and water (0.015 mL) and heated at 60 C for Ih, then diluted with EtOAc and washed with 1 M NaHCÜ3, dried and evaporated. The residue was purified by silica gel chromatography (12 g S1O2, 5-40% EtOAc in hexanes over 20 min) to provide 19-amino-9-methyl-6,17bis(trifluoromethyl)-l 5,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa1(20),2,4,10(21),11,13,16,18-octaen-6-ol (diastereomer pair 2) (2.6 mg, 10%). ’H NMR (400 MHz, Chloroform-d) δ 7.49 (s, 1H), 7.34 (t, 7= 2.2 Hz, 1H), 7.32 (d, 7= 7.8 Hz, 1H), 7.07 (d, 7 = 7.6 Hz, 1H), 7.01 (dd, 7= 8.1, 2.2 Hz, 1H), 5.16 (s, 2H),3.54(s, 1H), 2.93 (t, 7= 7.4 Hz, 1H),

275

2.25 (t, J= 13.4 Hz, 1H), 2.08 (td, J= 13.3,4.6 Hz, 1H), 1.98 (dt, J= 13.0, 10.7 Hz, 1H), 1.81 (t, J= 13.2 Hz, 1H), 1.33 (d, J= 7.1 Hz, 3H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.34, 77.95 ppm. ESI-MS m/z cale. 474.11267, found 475.1 (M+l)⁺; Rétention time: 1.54 minutes.

Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column 5 (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 21: Préparation of 20-amino-13-fluoro-6,18-bis(trifluoromethyl)-16,23-dioxa-

3,4,21-triazatetracycIo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-ol (enantiomer 1), Compound 31, and 20-amino-13-fluoro-6,18-bis(trifluoromethyl)-16,23dioxa-3,4,21-triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen6-0I (enantiomer 2), Compound 32

enantiomer 1 enantiomer 2

Step 1: 2-[l-Benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(3-fluoro-5-iodo-phenoxy)-3nitro-5-(trifluoromethyI)-2-pyridyl]-l,3,4-oxadiazole

276

The reagent CS2CO3 (127 mg, 0.3898 mmol) was added to a mixture of [6-[5-[lbenzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)-2pyridyl] trifluoromethanesulfonate (230 mg, 0.3536 mmol) and 3-fluoro-5-iodo-phenol (134 mg, 0.565 mmol) in DMF (4 mL) and was stirred at 0 °C for 1 h. The mixture was diluted with ether, washed with water (2X), brine, dried (MgSO₄) and evaporated. The residue was purified by silica gel chromatography (12 g S1O2, 0-30% of a solution (20% EtOAc in hexanes) to hexanes over 15 min) to provide 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(3-fluoro-5-iodophenoxy)-3-nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (180 mg, 55%). *H NMR (400 MHz, Chloroform-d) δ 8.76 (s, 1H), 7.43 - 7.41 (m, 1H), 7.39 (dd, J= 7.7, 1.6 Hz, 1H), 7.36 7.29 (m, 5H), 7.01 (dt, J= 8.7, 2.3 Hz, 1H), 5.73 (ddd, J= 17.0, 10.4, 6.4 Hz, 1H), 5.01 (d, J= 17.4 Hz, 1H), 4.95 (d, J= 10.2 Hz, 1H), 4.80 (d, J= 10.6 Hz, 1H), 4.62 (d, J= 10.6 Hz, 1H), 2.54 - 2.28 (m, 3H), 2.20 (dt, J= 17.3, 7.9 Hz, 1H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ 63.97, -73.02, -108.26 ppm. ESI-MS m/z cale. 738.021, found 739.0 (M+l)⁺; Rétention time: 0.6 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: 6-(Benzyloxy)-13-fIuoro-20-nitro-6,18-bis(trifluoromethyI)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,9,ll(22),12,14,17,19-nonaene {ElZ mixture)

A mixture of 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(3-fluoro-5-iodophenoxy)-3-nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (180 mg, 0.2438 mmol), palladium (II) acetate (10 mg, 0.04454 mmol), tris-o-tolylphosphane (27 mg, 0.08871 mmol) and triethylamine (51 pL, 0.3659 mmol) in acetonitrile (6.3 mL) was bubbled with N2 for 1 min then heated to 100 °C for Ih. The mixture was diluted with ether and washed with 1 M NH₄C1,1 M NaHCCh, brine then dried (MgSO₄) and evaporated. The residue was purified by silica gel chromatography (24 g S1O2, 0-50% of a solution (20% EtOAc in hexanes) to hexanes over 20

277 min) to provide 6-(benzyloxy)-13-fluoro-20-nitro-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[ 15.3.1.12,5.111,15]tricosa-1 (21 ),2,4,9,11 (22), 12,14,17,19-nonaene (E/Z mixture) (35 mg, 18%). Ή NMR (400 MHz, Chloroform-d) δ 8.79 (s, 1H), 7.35 - 7.27 (m, 6H), 6.89 - 6.77 (m, 2H), 6.50 (d, J= 11.5 Hz, 1H), 5.79 - 5.65 (m, 1H), 4.90 (d, J= 10.8 Hz, 1H), 4.67 (d, J= 11.0 Hz, 1H), 2.88 - 2.71 (m, 1H), 2.66 - 2.52 (m, 1H), 2.39 (td, J= 13.5, 3.2 Hz, 1H), 2.34 - 2.14 (m, 1H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.53, -63.87, -73.43, 73.64, -110.37, -110.61 ppm. ESI-MS m/z cale. 610.1087, found 611.2 (M+l)⁺; Rétention time: 0.55 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 3: 20-Amino-13-fIuoro-6,18-bis(trifluoromethyI)-16,23-dioxa-3,4,21triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6-ol (enantiomer 1), Compound 31, and 20-amino-13-fluoro-6,18-bis(trifluoromethyl)-16,23dioxa-3,4,21-triazatetracyclo[15.3.1.12,5.111,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen6-0I (enantiomer 2), Compound 32

Part 1: A mixture of 6-(benzyloxy)-13-fluoro-20-nitro-6,18-bis(trifluoromethyl)-16,23dioxa-3,4,21-tri azatetracyclo[l 5.3.1.12,5.111,15]tricosa-l(21 ),2,4,9,11(22),12,14,17,19-nonaene (E/Z mixture) (35 mg, 0.05734 mmol) and Pd/C (15 mg of 10 % w/w, 0.01410 mmol) in AcOH (400 pL), MeOH (400 pL) and EtOAc (800 pL) was stirred at room température under 200 psi H2 in a stainless steel pressure vessel for 20 h. Then the mixture was filtered and the filtrate evaporated to provide 6-(benzyloxy)-13-fhioro-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[ 15.3.1.12,5.111,15]tricosa-1(21),2,4,11(22), 12,14,17,19-octaen-20-amine which was taken directly to the next reaction. ESI-MS m/z cale. 582.1502, found 583.2 (M+l)⁺; Rétention time: 0.58 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile

278 phase A = H₂O (0.05 % CF₃CO₂H). Mobile phase B = acetonitrile (0.035 % CF₃CO₂H). Flow rate =1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Part 2: 6-(Benzyloxy)-13-fluoro-6,18-bis(trifluoromethyl)-16,23 -dioxa-3,4,21 triazatetracyclo[ 15.3.1.12,5.111,15] tricosa-1 (21 ),2,4,11 (22), 12,14,17,19-octaen-20-amine was dissolved in TFA (300 pL) and water (15 pL) and heated at 60 °C for Ih, then diluted with EtOAc and washed with 1 M NaHCO₃, dried and evaporated. The residue was purified by silica gel chromatography (12 g SiO₂, 5-40% EtOAc in hexanes over 20 min) to provide 20-amino-13fluoro-6,18-bis(trifluoromethyl)-16,23 -dioxa-3,4,21 -triazatetracyclof 15.3.1.12,5.111,15] tricosa1(21),2,4,11(22), 12,14,17,19-octaen-6-ol (2.4 mg, 9%). *H NMR (400 MHz, Chloroform-d) δ 7.53 (s, 1H), 7.28 (s, 1H), 6.79 (dd, J= 9.2, 2.2 Hz, 1H), 6.73 (dd, .7 = 9.2, 2.3 Hz, 1H), 5.51 (s, 2H), 3.68 (s, 1H), 2.80 (ddd, J= 14.0, 6.9, 3.7 Hz, 1H), 2.69 (ddd, J= 13.8, 8.8, 3.8 Hz, 1H), 2.29 (td, J= 13.3, 5.1 Hz, 1H), 2.05 (d, J= 13.1 Hz, 1H), 2.01 -1.91 (m, 1H), 1.85 - 1.67 (m, 3H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.62, -78.86, -112.30 ppm. ESI-MS m/z cale. 492.10324, found 493.1 (M+l)⁺; Rétention time: 1.62 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF₃CO₂H). Mobile phase B = acetonitrile (0.035 % CF₃CO₂H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Part 3: Racemic 20-amino-13-fluoro-6,18-bis(trifluoromethyl)-16,23-dioxa-3,4,21triazatetracyclo[ 15.3.1.12,5.111,15]tricosa-1 (21 ),2,4,11 (22), 12,14,17,19-octaen-6-ol was dissolved into 1:1 acetonitrile/MeOH and purified by préparative SFC eluting a gradient of 5 mM NH₃ in methanol to CO₂ (5-15% over 10 min at 60 mL/min) though a 21.2 X 250mm AS3 column, 5pm particle, providing first eluent 20-amino-13-fluoro-6,18-bis(trifluoromethyl)-16,23dioxa-3,4,21-triazatetracyclo[15.3.1.12,5.11 l,15]tricosa-l(21),2,4,ll(22),12,14,17,19-octaen-6ol, (enantiomer 1) (1.7 mg, 5%). Ή NMR (400 MHz, Chloroform-d) δ 7.53 (s, 1H), 7.28 (d, J= 1.7 Hz, 1H), 6.79 (dt, J= 9.3, 2.0 Hz, 1H), 6.73 (dt, J= 9.1, 2.3 Hz, 1H), 5.51 (s, 2H), 3.67 (s, 1H), 2.80 (ddd, J= 14.1, 6.8, 3.7 Hz, 1H), 2.68 (ddd, J= 20.2, 9.7, 5.0 Hz, 1H), 2.29 (ddd, J= 14.1, 12.5, 5.1 Hz, 1H), 2.10 - 2.02 (m, 2H), 2.01 - 1.89 (m, 1H), 1.85 - 1.67 (m, 2H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.62, -78.86, -112.30 ppm; ESI-MS m/z cale. 492.10324, found 493.1 (M+l)⁺; Rétention time: 1.61 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF₃CO₂H). Mobile phase B = acetonitrile (0.035 % CF₃CO₂H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

279

Continued elution provided as the second eluent 20-amino-13-fluoro-6,18bis(trifluoromethyl)-16,23-dioxa-3,4,21 -triazatetracyclo[ 15.3.1.12,5.111,15]tricosa1(21),2,4,11(22), 12,14,17,19-octaen-6-ol (enantiomer 2) (1.4 mg, 7%). 'H NMR (400 MHz, Chloroform-d) δ 7.53 (s, 1H), 7.28 (d, J= 1.8 Hz, 1H), 6.79 (dt, J= 9.1, 2.0 Hz, 1H), 6.73 (dt, J = 9.2, 2.3 Hz, 1H), 5.51 (s, 2H), 3.63 (s, 1H), 2.80 (ddd, J= 14.0, 6.8,3.7 Hz, 1H), 2.69 (ddd, J = 13.8, 8.7, 3.8 Hz, 1H), 2.29 (ddd, J = 14.1, 12.4, 5.1 Hz, 1H), 2.11 - 2.02 (m, 2H), 2.01 - 1.89 (m, 1H), 1.85 - 1.66 (m, 2H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.62, -78.87, -112.30 ppm; ESI-MS m/z cale. 492.10324, found 493.1 (M+l)⁺; Rétention time: 1.62 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x

4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 22: Préparation of 19-amino-12-fluoro-9-methyl-6,17-bis(trifluoromethyI)-15,2215 dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaen-

6-0I (diastereomer pair 1), Compound 33

diastereomer pair 1

280

Step 1: 6-(Benzyloxy)-12-fluoro-9-methylidene-19-nitro-6,17-bis(trifluoromethyl)-15,22dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaene

A mixture of 2-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(3-fluoro-5-iodophenoxy)-3-nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (180 mg, 0.2438 mmol), palladium (II) acetate (10 mg, 0.04454 mmol), tris-o-tolylphosphane (27 mg, 0.08871 mmol) and triethylamine (51 pL, 0.3659 mmol) in acetonitrile (6.3 mL) was bubbled with N₂ for 1 min then heated at 100 °C for Ih. The mixture was diluted with ether and washed with 1 M NH4CI, 1 M NaHCCh, brine then dried (MgSCL) and evaporated. The residue was purified by silica gel chromatography (24 g SiO₂, 0-50% of a solution (20% EtOAc in hexanes) to hexanes over 20 min) to provide 6-(benzyloxy)-12-fluoro-9-methylidene-19-nitro-6,17-bis(trifluoromethyl)15,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18octaene (9.4 mg, 6%). 'H NMR (400 MHz, Chloroform-d) δ 8.65 (s, 1H), 7.32 - 7.27 (m, 2H), 7.23 (d, J= 15.5 Hz, 4H), 6.84 (dt, J= 8.9, 2.4 Hz, 2H), 5.94 - 5.76 (m, 1H), 5.53 (dt, J= 14.9, 6.7 Hz, 1H), 4.83 (d, J= 11.3 Hz, 1H), 4.41 (d, J= 11.3 Hz, 1H), 3.37 (d, J= 5.9 Hz, 2H), 3.11 (dd, J= 14.9, 5.6 Hz, 1H), 2.78 (dd, J = 15.2, 7.9 Hz, 1H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.81, -73.39, -110.54 ppm. ESI-MS m/z cale. 610.1087, found 611.2 (M+l)⁺; Rétention time: 0.52 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H₂O (0.05 % CF₃CO₂H). Mobile phase B = CH3CN (0.035 % CF₃CO₂H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

281

Step 2: 19-Amino-12-fluoro-9-methyl-6,17-bis(trifluoromethyI)-15,22-dioxa-3,4,20triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaen-6-oI (diastereomer pair 1), Compound 33

F

Diastereomer pair 1

Part 1: A mixture of 6-(benzyloxy)-12-fluoro-9-methylidene-19-nitro-6,17bis(trifluoromethyl)-15,22-dioxa-3,4,20-triazatetracyclo[ 14.3.1.12,5.110,14] docosal(20),2,4,10(21),ll,13,16,18-octaene (9.4 mg, 0.01540 mmol) and Pd/C (15 mg of 10 % w/w, 0.01410 mmol) in AcOH (400 pL), MeOH (400 pL) and EtOAc (800 pL) was stirred at room température under 200 psi H2 in a stainless Steel pressure vessel for 20 h. Then the mixture was filtered and the filtrate evaporated to provide 6-(benzyloxy)-12-fluoro-9-methyl-6,17bis(trifluoromethyl)-15,22-dioxa-3,4,20-triazatetracyclo[14.3.1.12,5.110,14]docosa1(20),2,4,10(21),1 l,13,16,18-octaen-19-amine. ESI-MS m/z cale. 582.1502, found 583.2 (M+l)⁺; Rétention time: 0.59 minutes (yield missing). Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (30 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002349), and a dual gradient run from 50 - 99% mobile phase B over 1.0 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.5 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Part 2: 6-(Benzyloxy)-12-fluoro-9-methyl-6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),ll,13,16,18-octaen-19-amine was dissolved in TFA (300 pL) and water (15 pL) and heated at 60 °C for Ih, then diluted with EtOAc and washed with 1 M NaHCÛ3, dried and evaporated. The residue was purified by silica gel chromatography (12 g S1O2, 5-40% EtOAc in hexanes over 20 min) to provide diastereomer pair 1,19-amino-12-fluoro-9-methyl-6,17-bis(trifluoromethyl)-15,22-dioxa-3,4,20triazatetracyclo[14.3.1.12,5.110,14]docosa-l(20),2,4,10(21),11,13,16,18-octaen-6-ol (1.3 mg, 4%). ’H NMR (400 MHz, Chloroform-d) δ 7.50 (s, 1H), 7.13 (d, J= 2.1 Hz, 1H), 6.82 (dt, J= 9.6, 1.9 Hz, 1H), 6.77 (dt, J= 9.1, 2.2 Hz, 1H), 5.20 (s, 2H), 3.44 (s, 1H), 2.95 (d, J= 8.8 Hz, 1H), 2.25 (t, .7=13.3 Hz, 1H), 2.13- 1.90 (m, 2H), 1.79 (t, J= 13.3 Hz, 1H), 1.32 (d, .7=7.1 Hz, 3H) ppm; ¹⁹F NMR (376 MHz, Chloroform-d) δ -63.31, -78.21, -112.26 ppm. ESI-MS m/z cale.

282

492.10324, found 493.1 (M+l)⁺; Rétention time: 1.83 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 %

CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 23: Préparation of (6JÎ)-17-amino-12-isopropyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1) (hydrochloride sait), Compound 34, and (67?)-17-amino-12-isopropyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,1610 pentaen-6-ol (enantiomer 2) (hydrochloride sait), Compound 35

Step 1: terf-Butyl A-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazoI-2-yl]-6-(l-isopropyIbut-3-enoxy)-5-(trifluoromethyl)-3-pyridyl]-7V-te/7butoxycarbonyl-carbamate

283

To a solution of ZerZ-butyl 7V-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(triiluoromethyl)-3-pyridyl]-jV-/eri-butoxycarbonylcarbamate (200 mg, 0.2904 mmol) and 2-methylhex-5-en-3-ol (104 mg, 0.9108 mmol) in toluene (2 mL) was added triphenylphosphine (162 mg, 0.6176 mmol). After stirring at room température for 1 min, DIAD (120 pL, 0.6095 mmol) was added and the mixture stirred at room température for 30 min. The reaction mixture was diluted with EtOAc then washed with saturated aqueous sodium bicarbonate (IX), saturated aqueous NH4CI (IX) and brine (IX) then dried over magnésium sulfate, filtered and concentrated to a yellow oil which was purified by silica gel chromatography using a gradient of 0% to 50% EtOAc in hexanes giving as a diastereomeric mixture and clear, slightly yellow syrup, ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-(l-isopropylbut-3-enoxy)-5(trifluoromethyl)-3-pyridyl]-JV-terAbutoxycarbonyl-carbamate (168 mg, 74%). ESI-MS m/z cale. 784.3271, found 785.3 (M+l)⁺; Rétention time: 2.23 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: terf-Butyl 7V-[(6.R)-6-benzyloxy-12-isopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]-7V-tert‘butoxycarbonyl-carbamate (£7Z mixture)

To a degassed solution of terAbutyl A-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromcthyl)pent4-enyl]-l,3,4-oxadiazol-2-yl]-6-(l-isopropylbut-3-enoxy)-5-(trifluoromethyl)-3-pyridyl]-7V-terZbutoxycarbonyl-carbamate (164 mg, 0.2090 mmol) in DCE (100 mL) was added dichloro[l,3bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene][[5-[(dimethylamino)sulfonyl]-2-(lmethylethoxy-O)phenyl]methylene-C]ruthenium(II) (Zhan catalyst-lB, 18 mg, 0.02453 mmol) ail at once and the reaction mixture was heated at 60 °C for ~6 h. The reaction was quenched with few drops of DMSO and the solvents were removed by rotary évaporation. The resulting brown residue was purified by silica gel chromatography using a gradient of 0 to 50% EtOAc in 284 hexanes giving as a mixture of diastereomers and as a colorless viscous oil, ZerZ-butyl N-[(6R)-6benzyloxy-12-isopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,9,14,16-hexaen-l 7-yl]-7V-ter/-butoxycarbonylcarbamate (E/Z mixture) (97 mg, 61%). ESI-MS m/z cale. 756.2958, found 757.3 (M+l)⁺;

Rétention time: 2.08 minutes and 2.17 (major fragment 657.3 (M-Boc)⁺. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 3: tert-Butyl A-re/-r-butoxycarbonyl-A-[(6/?)-6-hydroxy-12-isopropyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl] carbamate

To a solution of ZerZ-butyl jV-[(67?)-6-benzyloxy-12-isopropyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]-7V-teributoxycarbonyl-carbamate (E/Z mixture) (97 mg, 0.1282 mmol) in EtOH (5 mL) was added Pd/C (46 mg of 10 % w/w, 0.04322 mmol, 50% water by weight) in a 250 mL flask equipped with a hydrogen balloon using a 3-way adapter. The flask was subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum again. The vessel was filled with hydrogen gas and the mixture was stirred at room température ovemight. The flask was subjected to vacuum and backfilled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated to give, as a colorless viscous oil and a mixture of diastereomers, Zeri-butyl 7V-/er/-butoxycarbonyl-7V-[(67?)-6-hydroxy-12-isopropyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (85 mg, 99%); ESI-MS m/z cale. 668.26447, found 569.2 (M-Boc+1)⁺; Rétention time: 1.62 minutes and Rétention time: 1.65 minutes (inséparable diastereomers). Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % 285

Step 4: (6/?)-17-Amino-12-isopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1) (hydrochloride sait), Compound 34, and (67î)-17-amino-12-isopropyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (enantiomer 2) (hydrochloride sait), Compound 35 v-cf₃OH rCF₃OH

C^cf3 OH

O. .N.

enantiomer 1 enantiomer 2 iert-Butyl 7V-teri-butoxycarbonyl-7V-[(6Æ)-6-hydroxy-12-isopropyl-6,15bis(trifluorom ethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (80 mg, 0.1196 mmol) was dissolved in a pre-made solution (1:3 TFA/dichloromethane) of TFA (250 pL, 3.245 mmol) and dichloromethane (750 pL). The reaction was stirred for ~1 h and the solvents were evaporated. The résultant residue was dissolved in 2 mL of MeOH and purified by reverse phase HPLC using a gradient from 40% to 80% acetonitrile in water (+5 mM HCl) over 30.0 minutes giving as an off-white solid and the first enantiomer to elute, (6/?)-17-amino-12-isopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (hydrochloride sait, enantiomer 1) (18.3 mg, 61%). *H NMR (400 MHz, DMSO-d₆) δ 7.77 (s, 1H), 7.57 (s, 1H), 6.35 (s, 2H), 4.59 (dt, J= 5.9, 2.7 Hz, 1H), 2.34 - 2.22 (m, 2H), 2.18 - 2.07 (m, 1H), 2.01 - 1.88 (m, 1H), 1.69 (m, 2H), 1.58 - 1.35 (m, 5H), 0.99 (d, J= 2.7 Hz, 3H), 0.98 (d, J= 2.7 Hz, 3H) ppm; ESI-MS m/z cale. 468.1596, found 469.1 (M+l)⁺ ; Rétention time: 1.74 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C. The second enantiomer to elute, as an off-white solid, was (6Æ)-17-amino-12-isopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (hydrochloride sait, enantiomer 2) (19.0 mg, 63%). Ή NMR (400 MHz, DMSO-d₆) δ 7.77 (s, 1H), 7.57 (s, 1H), 6.35 (s, 2H), 4.60-4.39 (m, 1H), 2.34 - 2.24 (m, 1H), 2.18 (d, J= 8.6 Hz, 1H), 2.14-2.04 (m, 1H), 1.99-1.89

286 (m, 1H), 1.63 (s, 3H), 1.42 (d,J= 23.7 Hz, 4H), 1.00 (d, J= 2.5 Hz, 3H), 0.98 (d, J= 2.4 Hz, 3H) ppm; ESI-MS m/z cale. 468.1596, found 469.2 (M+l)⁺; Rétention time: 1.79 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 5 99% mobile phase B over 2.9 minutes. Mobile phase A = water (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Example 24: Préparation of (6R,12/î)-17-amino-12-(hydroxymethyl)-6,15- bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,1610 pentaen-6-ol, Compound 36

Step 1: (25)-l-BenzyIoxypent-4-en-2-ol

To a solution of bromo(vinyl)magnesium (37 mL of 1 M, 37.000 mmol) was added copper bromide (350 mg, 2.4399 mmol) at 0 °C, then the black mixture was stirred at -78 °C. After 5 min, a solution of (2S)-2-(benzyloxymethyl)oxirane (2 g, 12.180 mmol) in THF (20 mL) was added dropwise over 30 min at -78 °C. Then the black mixture was stirred at -78 °C for 30 min. Then methanol (5 mL) was added at -78 °C, followed by aqueous hydrogen chloride

287 (2M, 24 mL) and then the mixture was stirred at room température for 5 min. MTBE (40 mL) was then added, the aqueous layer was separated and extracted with MTBE (2 X 20 mL). The organic layers were washed with aqueous hydrogen chloride (IM, 40 mL), water (40 mL), aqueous sodium thiosulfate (10%, 40 mL) and again with water (40 mL). The organic layers were dried over sodium sulfate and concentrated under vacuum to give crude (25)-1benzyloxypent-4-en-2-ol (2.46 g, 102%) as a yellow oil. ESI-MS m/z cale. 192.11504, found 193.0 (M+l)⁺; Rétention time: 1.92 minutes; LCMS Method: Kinetex Polar Cis, 3.0 x 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% Formic Acid) 1.2 ml/min.

Step 2: te/7-ButyI A-[6-[(17?)-l-(benzyloxymethyl)but-3-enoxy]-2-[5-[(17?)-l-benzyIoxy-l(trifIuoromethyI)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3-pyridyl]-7V-fôr£butoxycarbonyl-carbamate

A solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-/erZ-butoxycarbonyl-carbamate (1.2 g, 1.7426 mmol) and (25)-1-benzyloxypent-4-en-2-ol (1.42 g, 7.3861 mmol) in toluene (12 mL) was treated with triphenylphosphine (1.35 g, 5.1471 mmol) followed by DIAD (1.0815 g, 1.03 mL, 5.3485 mmol) at rt. The yellow solution was stirred at room température ovemight. The yellow suspension was concentrated under vacuum, then dry-packed on silica with DCM. Purification by chromatography over a 120 g silica column (1 - 30% EtOAc/heptanes) gave tertbutyl 7V-[6-[( 17?) -1 -(benzyloxymethyl)but-3 -enoxy]-2-[5- [ 1 -benzyloxy-1 -(trifluoromethyl)pent4-enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3-pyridyl]-7V-/erZ-butoxycarbonyl-carbamate (1.5 g, 99%) as a yellow oil. Ή NMR (400 MHz, CDCI3) δ 7.85 (s, 1H), 7.41 - 7.24 (m, 10H), 5.78 (m, 2H), 5.61 - 5.52 (m, 1H), 5.14 - 4.95 (m, 4H),4.80 (d, J= 10.6 Hz, 1H), 4.66 - 4.51 (m, 3H), 3.80 - 3.64 (m, 2H), 2.64 - 2.16 (m, 6H), 1.43 (d, J= 2.3 Hz, 18H) ppm. ^I9F NMR (377 MHz, CDCI3) δ -64.05 (s, 3F), -73.04 (s, 3F) ppm.

288

Step 3: ZerZ-Butyl A-[(67?,127?)-6-benzyloxy-12-(benzyloxymethyl)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaen-17-yl]/V-ZerZ-butoxycarbonyl-carbamate (E/Z mixture)

Nitrogen was bubbled throughout a solution of ZerZ-butyl A-[6-[(l A)-l(benzyloxymethyl)but-3-enoxy]-2-[5-[(17î)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (200 mg, 0.2304 mmol) in DCE (62 mL) for 30 min. Zhan catalyst-lB (18 mg, 0.0245 mmol) was then added at rt, and nitrogen was bubbled again for 5 min. The light yellow solution was stirred at 60 °C (pre-heated oil bath) for 2.5 h. The brown solution was cooled to rt, then DMSO (~5 drops) was added to quench the catalyst. The solvent was removed under vacuum and the residue was dry-packed on silica with DCM. The product was purified by chromatography on a 40 g silica column (1 - 30% EtOAc/heptanes) to give ZerZ-butyl A-[(67?,127?)-6-benzyloxy-12(benzyloxymethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclof 12.3.1.12,5]nonadeca-1 ( 17),2,4,9,14(18), 15-hexaen-l 7-yl]-A-ZerZ-butoxycarbonylcarbamate (E/Z mixture) (134 mg, 46%) as a light yellow oil. ESI-MS m/z cale. 834.30634, Rétention time: 4.54 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 4: ZerZ-Butyl A^r-ZerZ-butoxycarbonyI-A-[(67?,12J?)-6-hydroxy-12-(hydroxymcthyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-17-yl]carbamate

Palladium over charcoal (280 mg, 10 % w/w, 0.2631 mmol) was added to a degassed solution of ZerZ-butyl A-[(6R, 12Æ)-6-benzyloxy-12-(benzyloxymethyl)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaen-17-yl]-A289

ZerZ-butoxycarbonyl-carbamate (E/Z mixture) (880 mg, 0.8296 mmol) in methanol (40 mL) at rt. The black suspension was degassed with nitrogen for 5 min, then hydrogen was bubbled throughout the suspension for 5 min. Then the mixture was stirred at room température for three days under a hydrogen atmosphère. The black suspension was filtered through Celite with DCM, and concentrated under vacuum to give crude ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V[(6/?,12/?)-6-hydroxy-12-(hydroxymethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,l 8triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (630 mg, 87%) as a white solid. ¹⁹F NMR (377 MHz, CDC1₃) δ -63.73 (s, 3F), -80.72 (s, 3F) ppm. ESIMS m/z cale. 656.2281, found 501.0 (M-155)⁺; Rétention time: 3.47 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 5: (6/î,121î)-17-Amino-12-(hydroxymethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa 3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 36

ZerZ-Butyl 7V-ZerZ-butoxycarbonyl-A^z-[(67?,127?)-6-hydroxy-12-(hydroxymethyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15pentaen-17-yl]carbamate (630 mg, 0.7196 mmol) was dissolved in a solution of hydrogen chloride in 1,4-dioxane (4 mL of 4 M, 16.000 mmol) at rt. The solution was stirred at room température ovemight. T he solvent was evaporated under vacuum and the residual orange oil was purified by reverse phase chromatography over a 50 g Cis column (5-80% acetonitrile/0.1% formic acid in water) to give (6Æ,12/?)-17-amino-12-(hydroxymethyl)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (199 mg, 59%) as a light yellow solid. Ή NMR (400 MHz, DMSO-d6) δ 7.77 (s, 1H), 7.59 (s, 1H), 6.38 (br s, 2H), 4.68 (t, J= 5.3 Hz, 1H), 4.65 - 4.57 (m, 1H), 3.68 - 3.56 (m, 2H), 2.28 - 2.05 (m, 3H), 1.69 - 1.30 (m, 7H) ppm. ¹⁹F NMR (377 MHz, DMSO-d6) δ -62.23 (s, 3F), -79.01 (s, 3F) ppm. ESI-MS m/z cale. 456.12323, found 457.1 (M+l)⁺; Rétention time: 2.91 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

290

Example 25: Préparation of (6R,12S)-17-amino-12-(hydroxymethyl)-6,15- bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol, Compound 37

Step 1: (2R)-l-Benzyloxypent-4-en-2-ol

To a solution of bromo(vinyl)magnesium in THF (92 mL of 1 M, 92.000 mmol) was added copper bromide (870 mg, 6.0648 mmol) at 0 °C, then the black mixture was stirred at -78 °C. After 5 min, a solution of (2/?)-2-(benzyloxymethyl)oxirane (5 g, 30.450 mmol) in THF (50 10 mL) was added dropwise with a dropping funnel over 15 min at -78 °C. Then the black mixture was stirred at -78 °C for 40 min. Then methanol (13 mL) was added at -78 °C, followed by aqueous hydrogen chloride (2M, 80 mL) and then stirred at room température for 5 min. MTBE (80 mL) was then added, the aqueous layer was separated and extracted with MTBE (2 X 40 mL). The organic layer was washed with aqueous hydrogen chloride (IM, 50 mL), water (50 15 mL), aqueous sodium thiosulfate (10%, 50 mL) and again with water (50 mL). The organic layer was dried over sodium sulfate, filtered and concentrated under vacuum to give a yellow oil (6.01 g). The crude oil was purified by chromatography over a 120 g silica column (1-50%

291

MTBE/heptanes) to give (27?)-l-benzyloxypent-4-en-2-ol (5.89 g, 97%) as a light yellow oil. ’H NMR (400 MHz, CDC1₃) δ 7.42 - 7.28 (m, 5H), 5.84 (ddt, J= 17.2, 10.1, 7.1 Hz, 1H), 5.20 5.05 (m, 2H), 4.57 (s, 2H), 3.90 (qd, J= 6.7, 3.5 Hz, 1H), 3.53 (dd, J= 9.5, 3.4 Hz, 1H), 3.39 (dd, 7= 9.5, 7.4 Hz, 1H), 2.35 (br s, 1H), 2.28 (t, 7= 6.7 Hz, 2H) ppm. ESI-MS m/z cale.

192.11504, found 193.2 (M+l)⁺; Rétention time: 2.51 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: ZerZ-Butyl A-[6-[(lY)-l-(benzyIoxymethyl)but-3-enoxy]-2-[5-[(ljR)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyI)-3-pyridyl]-A''-ZerZbutoxycarbonyl-carbamate

A solution of ZerZ-butyl A-[2-[5-[(17?)-l-bcnzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (1 g, 1.4522 mmol) and (27?)-l-benzyloxypent-4-en-2-ol (837 mg, 4.3536 mmol) in toluene (10 mL) was treated with triphenylphosphine (796 mg, 3.0349 mmol) followed by DIAD (616.20 mg, 0.6 mL, 3.0474 mmol) at room température. The yellow solution was stirred at room température ovemight. The yellow suspension was concentrated under vacuum, then dry-packed on silica with DCM. Purification by chromatography over a 120 g silica column (0-30% ethyl acetate in heptanes) gave ZerZ-butyl7V-[6-[(lS)-l-(benzyloxymethyl)but-3-enoxy]-2-[5-[(17î)-lbenzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3-pyridyl]7V-ZerZ-butoxycarbonyl-carbamate (1.21 g, 96%) as a yellow oil. *H NMR (400 MHz, CDCI3) δ 7.85 (s, 1H), 7.41 - 7.27 (m, 10H), 5.85 - 5.69 (m, 2H), 5.59 - 5.52 (m, 1H), 5.14 - 4.94 (m, 4H), 4.80 (d, J= 10.5 Hz, 1H), 4.62 (d, 7= 10.8 Hz, 1H), 4.60 - 4.51 (m, 2H), 3.78 - 3.64 (m, 2H), 2.65 - 2.14 (m, 6H), 1.43 (s, 18H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -64.05 (s, 3F), -73.00 (s, 3F) ppm. ESI-MS m/z cale. 862.33765, found 864.1 (M+l)⁺; Rétention time: 4.65 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

292

Step 3: terf-Butyl 7V-[(6Æ,12*S)-6-benzyloxy-12-(benzyIoxymethyl)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaen-17-yl]A-zZ'rZ-butoxycarbonyl-carbamate (E!Z mixture)

E/Z mixture

A stirring solution of teri-butyl 7V-[6-[(15)-l-(benzyloxymethyl)but-3-enoxy]-2-[5-[(17?)l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3pyridyl]-7V-terLbutoxycarbonyl-carbamate (1.21 g, 1.3911 mmol) in 1,2-dichloroethane (500 mL) was degassed with bubbling with nitrogen gas for 20 hours. To the solution was added Zhan catalyst-lB (118 mg, 0.1608 mmol) then the reaction was heated in an oil bath set at 60 °C for 5.5 hours. Once cooled at room température, the catalyst was quenched with a few drops of DMSO (about 5-6 drops) and the reaction was concentrated under reduced pressure. The residue was dry loaded on silica gel and purified by silica gel liquid chromatography eluting from 0% to 30% ethyl acetate in heptane, to afford ZerZ-butyl 7V-[(67?,12S)-6-benzyloxy-12(benzyloxymethyl)-6,l 5-bis(trifluoromethyl)-13,l 9-dioxa-3,4,l 8triazatricyclo[ 12.3.1.12,5]nonadeca-l ( 17),2,4,9,14(18), 15-hexaen-l 7-yl]-7V-/er/-butoxycarbonylcarbamate (E/Z mixture)(543 mg, 45%) as colorless oil. ’H NMR (400 MHz, CDCh) δ 7.93 7.84 (m, 1H), 7.42 - 7.23 (m, 10H), 5.65 - 5.40 (m, 2H), 5.16 - 4.93 (m, 1H), 4.76 - 4.67 (m, 3H), 4.67 - 4.44 (m, 1H), 3.91 - 3.71 (m, 2H), 3.54 - 3.43 and 2.83 - 2.70 (m, 1H), 2.65 - 2.08 (m, 5H), 1.53 - 1.38 (m, 18H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.45 to -63.84 (m, 3F), -73.75 to -74.24 (m, 3F) ppm. Rétention time: 4.51 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid), flow =1.2 mL/min.

293

Step 4: ZerZ-Butyl 7V-/eri-butoxycarbonyl-7V-[(61î,12S)-6-hydroxy-12-(hydroxymethyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca1 (17),2,4,14(18),15-pentaen-l 7-yl] carbamate

E/Z mixture

HO^

Palladium over charcoal (214 mg, 10 % w/w, 0.2011 mmol) was added to a degassed solution of ZerZ-butyl /V-[(67?,12S)-6-benzyloxy-12-(benzyloxymethyl)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaen-17-yl]-7VZerZ-butoxycarbonyl-carbamate (E/Z mixture) (543 mg, 0.6264 mmol) in methanol (25 mL) at room température. The black suspension was degassed with nitrogen for 5 min, then hydrogen was bubbled through the suspension for 5 min. Then the mixture was stirred at room température ovemight under hydrogen atmosphère. The black suspension was filtered through Celite with DCM, and concentrated under vacuum to give crude ZerZ-butyl N-tertbutoxycarbonyl-7V-[(67?, 125)-6-hydroxy-12-(hydroxymethyl)-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (446 mg, 96%) as a light yellow oil. 'H NMR (400 MHz, CDCh) δ 7.88 (s, 1H), 4.92 - 4.83 (m, 1H), 3.96 - 3.82 (m, 2H), 3.60 (br. s, 1H), 2.59 - 2.47 (m, 1H), 2.38 - 2.12 (m, 3H), 2.08 - 1.93 (m, 2H), 1.73 - 1.33 (m, 23H) ppm. ^I9F NMR (377 MHz, CDCI3) δ -63.78 (s, 3F), -77.56 (s, 3F) ppm. ESI-MS m/z cale. 656.2281, found (M+)+;501.1 (M-155)+; Rétention time: 3.54 minutes; LCMS Method: Kinetex Polar Cis, 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H₂O (0.1% formic acid), flow =1.2 mL/min.

Step 5: (6E,125)-17-Amino-12-(hydroxymethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa 3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 37

To a 0 °C stirring solution of ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(67?,125)-6-hydroxy12-(hydroxymethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18294 triazatricyclo[ 12.3.1.12,5]nonadeca-1(17),2,4,14( 18), 15-pentaen-17-yl]carbamate (65 mg, 0.0483 mmol) in 1,4-dioxane (2 mL) was added dropwise a solution of HCl in 1,4-dioxane (0.5 mL of 4 M, 2.0000 mmol). The ice-water cooling bath was removed 2 minutes after the addition and the mixture was stirred at room température ovemight. Then, more HCl solution in 1,4dioxane (1.5 mL of 4 M, 6.0000 mmol) was added at room température and stirring continued for 4 hours. The volatiles were removed by évaporation under reduced pressure and HCl (2 mL of 4 M, 8.0000 mmol) was added at room température. The mixture was stirred ovemight with gentle heating at 30 °C. The volatiles were removed by évaporation under reduced pressure. The crude was solubilized in dichloromethane (3 mL) and was furthermore concentrated by évaporation under reduced pressure. Combined lots of crude (67?,12S)-17-amino-12(hydroxymethyl)-6,l 5-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol were purified by reverse phase Cis chromatography on a 50 g column, eluting with a gradient of acetonitrile (0 - 50%) in water containing 0.1 w/w % of formic acid. Pure fractions were combined and concentrated by évaporation under reduced pressure, then transferred in a 25 mL flask and submitted for freezedrying (acetonitrile/water) ovemight to finally isolate (67?,125)-17-ammo-12-(hydroxymethyl)6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-6-ol (127 mg, 49%) as light yellow solid. *H NMR (400 MHz, DMSOd₆) δ 7.77 (s, 1H), 7.58 (s, 1H), 6.44 - 6.27 (m, 2H), 4.78 - 4.60 (m, 2H), 3.74 - 3.52 (m, 2H), 2.35 - 2.24 (m, 1H), 2.21 - 2.08 (m, 2H), 1.82 - 1.33 (m, 7H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.24 (s, 3F), -76.43 (s, 3F) ppm. ESI-MS m/z cale. 456.12323, found 457.1 (M+l)⁺; Rétention time: 3.02 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid), flow =1.2 mL/min.

Example 26: Préparation of (6_JR,12R)-17-amino-9,10-dideuterio-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol, Compound 38

Step 2 CF₃.

Step 1

V-CF₃OH

O O

Step 1: tert-Butyl A^r-tert-butoxycarbonyl-7V-[(6/î,12/?)-9,10-dideuterio-6-hydroxy-12-methyI6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate

295

To a solution of Ze/7-butyl 7V-[(67î,127?)-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16hexaen-17-yl]-7V-ier/-butoxycarbonyl-carbamate (E/Z mixture) (200 mg, 0.2745 mmol) in CD3OD (8 mL) under nitrogen was added 10% palladium on carbon (50 mg, 0.0470 mmol). Nitrogen was replaced with deuterium gas through vacuum for 3 times. The mixture was stirred at room température under deuterium atmosphère (balloon) ovemight. The mixture was filtered through diatomaceous earth and washed EtOAc and concentrated to give ZerZ-butyl 7V-ter/-butoxycarbonyl-7V-[(6Æ, 127?)-9,10-dideuterio-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (184 mg, 100%) as a colorless oil. 'H NMR (400 MHz, CDCI3) δ 7.82 (s, 1H), 4.97 - 4.86 (m, 1H), 3.72 - 3.54 (m, 1H), 2.71 - 2.59 (m, 1H), 2.34 - 2.24 (m, 1 H), 2.21 2.11 (m, 1H), 2.08 - 1.96 (m, 1H), 1.67 - 1.19 (m, 25H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ 63.99 (s, 3F), -77.58 (s, 3F) ppm.

Step 2: (6R,12R)-17-Amino-9,10-dideuterio-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 38

To a solution of tert-butyl V-rerZ-butoxycarbonyl-7V-[(67?,127?)-9,10-dideuterio-6hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (184 mg, 0.2749 mmol) in CH2Q2 (2 mL) was added TFA (2.9600 g, 2 mL, 25.960 mmol). The mixture was stirred at room température for 1.5 h. The mixture was concentrated and co-evaporated with EtOAc (3X5 mL). The residue was dissolved in EtOAc (20 mL), washed with saturated NaHCO3 (5 mL) and dried with Na2SO4, filtered and concentrated. The residue was purifïed

296 by silica gel chromatography (24 g S1O2, eluting 0 to 30% EtOAc/heptanes) and the product was dissolved in minimum of acetonitrile and water and freeze-dried ovemight to affbrd (6R,12R)~ 17-amino-9,10-dideuterio-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (103 mg, 83%) as a yellow solid. ESI-MS m/z cale. 442.1409, found 443.1 (M+l)⁺; Rétention time: 3.63 minutes. ’H NMR (400 MHz, DMSO-de) δ 7.76 (s, 1H), 7.57 (s, 1H), 6.35 (s, 2H), 4.84 - 4.74 (m, 1H), 2.49 - 2.42 (m, 1H), 2.33 - 2.21 (m, 1H), 2.15 - 2.04 (m, 1H), 1.77 - 1.65 (m, 1H), 1.56 - 1.36 (m, 3H), 1.34 (d, J= 6.4 Hz, 3H), 1.25 - 1.12 (m, 1H) ppm. ^I9F NMR (377 MHz, DMSO-d₆) δ -62.50 (s, 3F), 76.38 (br s, 3F) ppm. ESI-MS m/z cale. 442.14087, found 443.1 (M+l)⁺; Rétention time: 3.63 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Example 27: Préparation of (61î,121î)-17-amino-ll,ll,12-trideuterio-12- (trideuteriomethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI, Compound 39

Step 1: (5)-2-(Methyl-d3)oxirane-2,3,3-d3

Acetic acid (0.94 mL, 16.41 mmol, 0.021 eq.) was added to a solution of (15,25)-(+)[l,2-cyclohexanediamino-M/V⁷-bis-(3,5-di-ter/-butylsalicylidene)]cobalt(II) ((5,5)-Co(salen) Jacobsen catalyst) (0.943 g, 1.56 mmol) in toluene (25 mL). The resulting solution was stirred at room température open to air for 30 minutes over which time the color changed from orange-red to dark-brown. The solution was concentrated under reduced pressure to give a crude brown solid. Racemic 2-(methyl-d₃)oxirane-2,3,3-d3 (50 g, 781.25 mmol, 1 equiv) was added to

297 dissolve the crude catalyst at room température and then cooled in an ice bath to 0 °C.

Deuterium oxide (8.6 mL, 429.69 mmol, 0.55 equiv) was added dropwise over 10 minutes. The reaction mixture was allowed to warm to room température and stirred ovemight. Distillation of the mixture under atmosphère pressure gave (5)-2-(methyl-d3)oxirane-2,3,3-d3 (17.23 g, boiling point = 31.5-34.5 °C, 34.5% yield) as a colorless oil which was stored in a freezer.

Step 2: (.V)-Pent-4-en-l,l,l,2,3,3-d6-2-ol

ho _D

CD

A solution of IM vinyl magnésium bromide in THF (100 mL, 100 mmol) was added dropwise while maintaining the température below -10 °C to a mixture of copper(I) chloride (0.20 g, 2 mmol) and lithium chloride (0.17 g, 4 mmol) in anhydrous THF (40 mL). The mixture was stirred at -5 to -10 °C for 15 minutes, and then cooled to -15 °C. A solution of (5)-2(methyl-d3)oxirane-2,3,3-d3 (6.4 g, 100 mmol) in anhydrous THF (100 mL) was added dropwise while maintaining the température below -5 °C. The resulting mixture was stirred at -10 °C to 0 °C for 4 hours. Saturated ammonium chloride in deuterium oxide (50 mL) was added slowly to quench the reaction while maintaining the température below 10 °C. Deuterium oxide (50 mL) was added and the mixture was stirred in an ice bath for 30 minutes. The layers were separated, and the organic layer was washed with a saturated ammonium chloride in deuterium oxide (30 mL). The combined aqueous layers were extracted with diethyl ether (3 X 50 mL). The combined organic layers were washed with saturated brine (50 mL), dried over sodium sulfate, filtered and concentrated by distillation under atmosphère pressure to give the desired (5)-pent-4en-l,l,l,2,3,3-dô-2-ol as a colorless oil (Fraction A: 1.29 g, boiling point = 80-100 °C; Fraction B: 2.27 g, boiling point = 100-80 °C; Fraction C: 0.40 g, boiling point = 80-65 °C, 40% combined yield). ’H NMR indicated that fraction A contained 0.25 molar ratio of THF, both fraction B and C were solvent free. Additional compound (5)-2-(methyl-d3)oxirane-2,3,3-d3 (2 X 5 g) was processed as described above to give compound (5)-pent-4-en-l,l,l,2,3,3-d6-2-ol as a colorless oil (Fraction A: 1.46 g, boiling point = 75-95 °C; Fraction B: 4.30 g, boiling point = 95100-75 °C; Fraction C: 0.83 g, boiling point = 80-65 °C, 40% combined yield). *H NMR indicated that fraction A contained 0.50 molar ratio of THF, both fraction B and C were solvent free.

298

Step 3: terZ-Butyl A^r-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyI)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-[(l.R)-l,2,2-trideuterio-l-(trideuteriomethyI)but-3-enoxy]-5(trifIuoromethyl)-3-pyridyl]-7V-ter/-butoxycarbonyI-carbamate

HO n

CD₃

A solution of ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (6.2 g, 9.04 mmol), triphenylphosphine (2.9 g, 10.9 mmol) and (S)-pent-4-en-l,l,l,2,3,3-d6-2-ol (1.0 g, 10.9 mmol) in toluene (45.0 mL) was stirred under a nitrogen atmosphère was heated to 45 °C. Diisopropyl azodicarboxylate (2.3 mL, 11.8 mmol) was added slowly over 20 minutes, maintaining an internai température of less than 55 °C. The reaction mixture was cooled to room température and diluted with ethyl acetate (50 mL) and water (50 mL). The organic layer was separated, washed with water (50 mL), saturated brine (50 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by silica gel chromatography (120 g S1O2), eluting with a gradient of 0 to 10% ethyl acetate in hexanes to give ZerZ-butyl7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]6-[(lR)-l,2,2-trideuterio-l-(trideuteriomethyI)but-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7VZerZ-butoxycarbonyl-carbamate as a clear, colorless oil (5.7 g, 83% yield). 'H NMR (400 MHz, DMSO-dô) δ 8.56 (d, J= 0.7 Hz, 1H), 7.44 - 7.29 (m, 5H), 5.92 - 5.70 (m, 2H), 5.10 (dd, J = 4.5, 1.9 Hz, 1H), 5.06 (dd, J =4.5,1.9 Hz, 1H), 5.01 (ddd, J =9.7, 7.7, 1.9 Hz, 2H),4.77 (d, J= 11.1 Hz, 1H), 4.67 (d, J= 11.1 Hz, 1H), 2.63-2.52 (m, 1H), 2.47 (d, J= 15.0 Hz, 1H), 2.27 (ddt, J= 16.0, 10.4, 6.0 Hz, 2H), 1.28 (d, J= 12.6 Hz, 18H) ppm. ²H NMR (400 MHz, DMSO) δ 5.33 (bs, 1D), 1.28 (s, 6D) ppm. ^l9F NMR (400 MHz, DMSO-d₆) δ -62.70, -73.26 ppm. ESIMS m/z cale. 762.33, found 785.3 (M+Na)⁺; Rétention time: 14.5 minutes. Final purity was determined by reversed phase HPLC using an Atlantis T3, 3 pm, 2.1 X 50 mm made by Waters (pn: 186003717), and a dual gradient run from 5 - 95% mobile phase B over 14 minutes with a 4 min hold at 95% B. Mobile phase A = H2O (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H). Flow rate = 0.7 mL/min, injection volume = 2 pL and column température = 40 °C.

299

Step 4: ZrrZ-Butyl A-[(67?,12/?)-6-benzyIoxy-ll,ll,12-trideuterio-12-(trideuteriomethyl)6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,9,14,16-hexaen-17-yl]-7V-tert-butoxycarbonyl-carbamate (E/Z mixture)

E/Z mixture

In a 3-necked flask charged with stirring bar, a gas dispersion needle, a condenser and a bubbler, /er/-butyl7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2yl]-6-[(17?)-l,2,2-trideuterio-l-(trideuteriomethyl)but-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7VZe/7-butoxycarbonyl-carbamate (5.4 g, 7.079 mmol) in DCE (810 mL, 0.009 M) was purged with N2 for 1 hour. The mixture was heated to 75 °C, then benzylidenebis(tricyclohexylphosphine)dichlororuthenium (1.201 g, 1.416 mmol) was added as a solid. The reaction was stirred at 75 °C with N2 purging. After 5 hours additional benzylidenebis(tricyclohexylphosphine)dichlororuthenium (0.45 g, 0.531 mmol) was added and the mixture was heated at 75 °C for 2 additional hours. The internai température was decreased to 50 °C, and 2-sulfanylpyridine-3-carboxylic acid (1.091 g, 7.033 mmol, 0.993 equiv.) was added followed by triethylamine (0.72 g, 7.115 mmol) and the reaction température was set to 45 °C. The material was allowed to stir ovemight. The mixture was removed from heating and 13 g of silica gel was added and stirred for 0.5 h at ambient température. The mixture was filtered over Celite, rinsed with DCE, and concentrated under reduced pressure. To the residue was added DCM/heptane (1:3, 50 mL), followed by filtration and the insoluble precipitate was washed with heptane. The filtrate was concentrated to give the crude material. The crude compound was dissolved in DCM/heptane (1:1,4 vols), and chromatographed on a 120 g normal phase silica column using a gradient of 0 % to 5 % EtOAc/hexanes over 13 min, followed by a gradient of 5% EtOAc for 23 min affording the crude product as colorless oil. The collected oil was then dissolved in MeOH (3 vols) and loaded onto a 50 g Cis reverse phase column which was eluted using a gradient of 40% to 100% acetonitrile/water over 20 min. The resulting pure fractions were concentrated and dried under house vacuum ovemight to give ZerZ-butyl 7V-[(6R,127î)-6-benzyloxy-l 1,11,12trideuterio-12-(trideuteriomethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaen-17-yl]-7V-ter/-butoxycarbonylcarbamate (E/Z mixture) as a white foam (2.21 g, 42.5 %) which was used directly in the next step.

300

Step 5: (6/?,12Æ)-17-Amino-ll,ll,12-trideuterio-12-(trideuteriomethyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol, Compound 39

E/Z mixture

terZ-Butyl 7V-[(6Æ,127î)-6-benzyloxy-l 1,1 l,12-trideuterio-12-(trideuteriomethyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16hexaen-17-yl]-7V-Zeri-butoxycarbonyl-carbamate (E/Zmixture) (2.21 g, 3.008 mmol) was dissolved in EtOH (44.2 mL, 0.068 M), and the vacuum and nitrogen was cycled 3X then treated with 10 % Pd/C (50% wet palladium) (0.445 g, 4.181 mmol). The mixture was stirred at ambient température under a hydrogen balloon for 23 h. The vacuum and hydrogen was cycled 3X, then filtered over Celite, washed with éthanol and evaporated under reduced pressure to give the crude ZerZ-butyl AZ-/er/-butoxycarbonyl-/V-[(67?,127?)-l 1,1 l,12-trideuterio-6-hydroxy-12(trideuteriomethyl)-6,15-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18) ,2,4,14,16-pentaen-17-yl]carbamate. The white foam was dissolved in DCM (17.68 mL, 0.17 M), cooled in an ice bath and treated with trifluoroacetic acid (5.294 mL, 69.182 mmol) under Ni. Then the ice bath was removed and the pale yellow solution was stirred for 2 h. The yellow solution was diluted with heptane (10 mL), filtered, and the filtrate was evaporated under reduced pressure. The crude material was treated with DCM (2.4 mL) in 50 °C water bath, then the yellow clear solution was diluted with hot (60 °C) heptane (12 mL), stirred in the warm water bath and allowed to slowly cool to ambient. The formed suspension was stirred at ambient température ovemight, and the white solid was collected and rinsed with heptane to give the first crop of product. The compound was dried in the vacuum oven at 40 °C purging with nitrogen ovemight. The mother liquor was concentrated and loaded on a Cis reverse phase chromatography (50 g) eluting with acetonitrile/water (30 - 80%) over 25 min. The obtained pale yellow solid was treated with hot DCM/heptane (1 mL/5 mL) then allowed to cool to ambient température ovemight. The formed solid was collected and rinsed with heptane to give 0.5 g of product as the second crop. The combined crops were dried in the vacuum oven at 45 °C under nitrogenN2 giving the off-white solid (6/?,12/?)-17-amino-l 1,11,12trideuterio-12-(trideuteriomethyl)-6,15-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (1.14 g, 84.9 %). *H NMR 301 (400 MHz, DMSO-dô) δ 7.76 (s, 1H), 7.58 (s, 1H), 6.35 (s, 2H), 2.29 (t, J= 11.7 Hz, 1H), 2.16 2.04 (m, 1H), 1.73 (s, 2H), 1.56 - 1.44 (m, 1H), 1.44 (s, 3H) ppm. ESI-MS m/z cale. 446.166, found 447.225 (M+l)⁺; Rétention time: 2.895 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by

Waters (pn: 186002350), and a dual gradient run from 1 - 99% mobile phase B over 4.5 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Example 28: Préparation of(67?,12Æ)-17-amino-12-methyI-6,15-bis(trifluoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,ll-diol (diastereomer 1), Compound 40, and (6.R,12.Æ)-17-ammo-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaene-6,ll-diol (diastereomer 2), Compound 41

Step 10

302

Step 12

Step 13

diastereomer 1 diastereomer 2

Step 1: Methyl (25)-2-triisopropyIsilyloxypropanoate

Into a 500 mL round bottom flask was added a solution of methyl (25)-25 hydroxypropanoate (10.702 g, 10 mL, 100.74 mmol) and imidazole (16.5 g, 242.37 mmol) in DCM (220 mL). The solution was cooled to 0 °C, and chloro(triisopropyl)silane (22.957 g, 26 mL, 116.69 mmol) was added to the reaction mixture dropwise over 30 minutes. The reaction mixture was warmed to room température as the ice melted and stirred ovemight. The reaction mixture was poured into a separatory funnel and washed with water (100 mL), saturated sodium 10 bicarbonate (100 mL), brine (100 mL), 1 N HCl (100 mL) and brine (100 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by silica gel chromatography using 0 to 5% ether in hexane to fumish methyl (25)-2triisopropylsilyloxypropanoate (25.4 g, 97%) as a clear liquid. *H NMR (500 MHz, Chloroformd) δ 4.44 (q, J = 6.7, 6.7, 6.7 Hz, 1H), 3.72 (s, 3H), 1.43 (d, J= 6.7 Hz, 3H), 1.19 - 0.99 (m,

21H)ppm.

303

Step 2: (2S)-2-Triisopropylsilyloxypropanal

Into a solution of methyl (2S)-2-triisopropylsilyloxypropanoate (1.008 g, 3.8703 mmol) in anhydrous DCM (18 mL) was added 1.0 M DIBAL-H in toluene (7.8 mL of 1 M, 7.8000 mmol) dropwise at -78 °C. Aller the addition, the reaction was stirred at the same température for 0.5 hour. It was quenched with ethyl acetate (4 mL) at the same température, and the reaction was warmed to 0 °C in an ice bath. Saturated sodium potassium tartrate aqueous solution (10 mL) was added. The reaction was stirred ovemight. Two layers were separated. The aqueous layer was extracted with DCM (3X15 mL). The combined organic layers were dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was loaded onto a column directly and purified by silica gel chromatography using 0 to 10% diethyl ether in hexane to fiimish (2S)-2-triisopropylsilyloxypropanal (838 mg, 94%) as a clear liquid. *H NMR (500 MHz, Chloroform-d) δ 9.66 (d, 1.8 Hz, 1H), 4.18 (qd, J= 6.8, 6.8, 6.8, 1.7 Hz, 1H),

1.30 (d, J= 6.8 Hz, 3H), 1.17 - 0.97 (m, 21H) ppm.

Step 3: (25)-2-Triisopropylsilyloxyhex-5-en-3-oI

OH

Into a solution of (25)-2-triisopropylsilyloxypropanal (18.24 g, 79.160 mmol) in anhydrous DCM (350 mL) was added allyl(bromo)magnesium in diethyl ether (90 mL of 1 M, 90.000 mmol) dropwise at -30 °C. The reaction was stirred at the same température for 45 minutes, then raised to 0 °C and stirred for another 15 minutes. The reaction was quenched with 10% ammonium chloride (300 mL). Two layers were separated, and the aqueous layer was extracted with DCM (2 X 250 mL). The combined organic layers were washed with brine (150 mL), dried over anhydrous magnésium sulfate and concentrated under vacuum. The residue was purified by silica gel chromatography using 0 to 20% ether in hexane to fiimish a 1:1 mixture of diastereomers, (2S)-2-triisopropylsilyloxyhex-5-en-3-ol (18 g, 83%) as a clear liquid. *H NMR (500 MHz, Chloroform-d) δ 6.01 - 5.75 (m, 1H), 5.21 - 4.99 (m, 2H), 4.00 - 3.82 (m, 1H), 3.75 - 3.37 (m, 1H), 2.53 - 2.03 (m, 3H), 1.28 - 0.90 (m, 24H) ppm.

304

Step 4: [(15)-2-Benzyloxy-l-methyl-pent-4-enoxy]-triisopropyl-silane

A slurry of NaH (110 mg, 60 % w/w, 2.7503 mmol) in anhydrous DMF (10 mL) was cooled to 0 °C. A solution of (25)-2-triisopropylsilyloxyhex-5-en-3-ol (672 mg, 2.4661 mmol) in anhydrous DMF (10 mL) was added to the reaction mixture dropwise. The reaction mixture was stirred at room température for 4 hours. Bromomethylbenzene (504.00 mg, 0.35 mL, 2.9468 mmol) was added to the reaction mixture at 0 °C dropwise. The reaction was then stirred at room température for 2 hours. The reaction was poured over 10% ammonium chloride aqueous solution (30 mL) and extracted with diethyl ether (3 X 30 mL). The combined organic layers were washed with brine (3X30 mL), dried over anhydrous magnésium sulfate and concentrated under vacuum. The residue was purified by silica gel chromatography using 0 to 50% DCM in hexane to fumish a 1:1 mixture of diastereomers, [(15)-2-benzyloxy-l-methyl-pent-4-enoxy]triisopropyl-silane (957 mg, 100%) as a clear liquid. 'H NMR (500 MHz, Chloroform-d) δ 7.42 - 7.22 (m, 5H), 6.01 - 5.78 (m, 1H), 5.15 - 4.88 (m, 2H), 4.85 - 4.53 (m, 2H), 4.20 - 3.90 (m, 1H), 3.63-3.35 (m, 1H), 2.58-2.08 (m, 2H), 1.31 - 1.10 (m, 3H), 1.11-0.78 (m, 21H)ppm.

Step 5: (25)-3-Benzyloxyhex-5-en-2-ol

Into a solution of [(15)-2-benzyloxy-l-methyl-pent-4-enoxy]-triisopropyl-silane (14.76 g, 40.704 mmol) in anhydrous THF (140 mL) was added IM TBAF in THF (41 mL of 1 M, 41.000 mmol) dropwise at 0 °C. The reaction was slowly raised to room température and stirred ovemight. The reaction was concentrated under vacuum to remove THF. The residue was diluted with ethyl acetate (400 mL) and washed with water (100 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by silica gel chromatography using 0 to 20% ethyl acetate in hexane to fumish a mixture of diastereomers, (25)-3-benzyloxyhex-5-en-2-ol (7.363 g, 88%) as a clear liquid. *H NMR (500

305

MHz, Chloroform-d) δ 7.45 - 7.20 (m, 5H), 5.96 - 5.77 (m, 1H), 5.20-4.99 (m, 2H), 4.78 4.44 (m, 2H), 3.98 - 3.67 (m, 1H), 3.47 - 3.20 (m, 1H), 2.56 - 1.94 (m, 3H), 1.24 - 1.12 (m, 3H) ppm.

Step 6: Methyl 6-[(l/?)-2-benzyIoxy-l-methyl-pent-4-enoxy]-3-nitro-5(trifluoromethyl)pyridine-2-carboxylate

Into a reaction vial was charged with methyl 6-hydroxy-3-nitro-5(trifluoromethyl)pyridine-2-carboxylate (67 mg, 0.2518 mmol), (2S)-3-benzyloxyhex-5-en-2-ol (71 mg, 0.3442 mmol) and triphenylphosphine (99 mg, 0.0875 mL, 0.3775 mmol) in anhydrous THF (1 mL). DIAD (69.948 mg, 0.067 mL, 0.3459 mmol) was added to the reaction mixture dropwise at 0 °C. The reaction was stirred at room température for 24 hours. The solvent was removed under vacuum. The residue was purified by silica gel chromatography directly using 0 to 10% ethyl acetate in hexane to fiimish methyl 6-[(l/?)-2-benzyloxy-l-methyl-pent-4-enoxy]3-nitro-5-(trifluoromethyl)pyridine-2-carboxylate (71 mg, 60%) as a clear gel. *H NMR (500 MHz, Chloroform-d) δ 8.73 - 8.46 (m, 1H), 7.43 - 7.16 (m, 5H), 5.98 - 5.72 (m, 1H), 5.72 5.50 (m, 1H), 5.19-4.93 (m, 2H), 4.73-4.61 (m, 1 H), 4.61-4.54 (m, 1H), 4.12-3.91 (m, 3H), 3.84 - 3.61 (m, 1H), 2.54 - 2.26 (m, 2H), 1.47 - 1.35 (m, 3H) ppm. ESI-MS m/z cale. 454.13516, found 455.1 (M+l)⁺; Rétention time: 3.96 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 7: Methyl 3-amino-6-[(l/?)-2-benzyIoxy-l-methyl-pent-4-enoxy]-5(trinuoromethyl)pyridine-2-carboxyIate

nh₂ o

Into a solution of methyl 6-[(l/?)-2-benzyloxy-l-mcthyl-pent-4-cnoxy]-3-nitro-5(trifluoromethyl)pyridine-2-carboxylate (1.255 g, 2.6514 mmol) in acetic acid (15 mL) was added iron (740.34 mg, 13.257 mmol) at rt. The reaction was stirred at room température for 3 hours. The reaction mixture was diluted with methanol (15 mL) and filtered through a pad of

306

Celite. The filtrate was concentrated under vacuum. The residue was diluted with ethyl acetate (50 mL) and saturated sodium bicarbonate (100 mL). The solution was filtered through a pad of Celite. Two layers were separated and the aqueous layer was extracted with ethyl acetate (2 X 50 mL). The combined organic layers were washed with brine (50 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purifîed by silica gel chromatography using 0 to 15% ethyl acetate in hexane to fumish methyl 3-amino-6-[(17?)-2benzyloxy-l-methyl-pent-4-enoxy]-5-(trifluoromethyl)pyridine-2-carboxylate (1.073 g, 93%) as a yellow gel. ESI-MS m/z cale. 424.161, found 425.0 (M+l)⁺; Rétention time: 3.89 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 8: Methyl 6-[(17?)-2-benzyIoxy-l-methyl-pent-4-enoxy]-3-(Zi?rz‘-butoxycarbonyIamino) 5-(trifluoromethyl)pyridine-2-carboxyIate

Into a solution of methyl 3-amino-6-[(17î)-2-benzyloxy-l-methyl-pent-4-enoxy]-5(trifluoromethyl)pyridine-2-carboxylate (1.073 g, 2.4776 mmol) and BOC2O (1.34 g, 6.1398 mmol) in anhydrous THF (20 mL) was added 1.0 M NaHMDS in THF (5 mL of 1 M, 5.000 mmol) dropwise at -78 °C. The reaction was stirred at the same température for 1 hour. The reaction was quenched cold with 10% ammonium chloride (20 mL). The reaction was warmed up to room température and extracted with ethyl acetate (3X30 mL). The combined organic layers were washed with brine (20 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purifîed by silica gel chromatography using 0 to 15% ethyl acetate in hexane to fumish methyl 6-[(12î)-2-benzyloxy-l-methyl-pent-4-enoxy]-3-(teributoxycarbonylamino)-5-(trifluoromethyl)pyridine-2-carboxylate (1.328 g, 100%) as a clear gel. ESI-MS m/z cale. 524.21344, found 525.3 (M+l)⁺; Rétention time: 4.51 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

307

Step 9: 6-[(lÆ)-2-Benzyloxy-l-methyl-pent-4-enoxy]-3-(teri-butoxycarbonylamino)-5(trifluoromethyl)pyridine-2-carboxyIic acid

Into a solution of methyl 6-[(17?)-2-benzyloxy-l-methyl-pent-4-enoxy]-3-(7er/butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2-carboxylate (1.328 g, 2.4812 mmol) in THF (15 mL) was added a solution of LiOH (299 mg, 12.485 mmol) in water (5 mL). The reaction was stirred at room température for 1 hour. The reaction was acidified with 1 N HCl to pH 1. The reaction was diluted with ethyl acetate (30 mL) and water (10 mL). Two layers were separated, and the aqueous layer was extracted with ethyl acetate (2 X 30 mL). The combined organic layers were washed with brine (30 mL), dried over anhydrous sodium sulfate and concentrated under vacuum to fumish 6-[(lR)-2-benzyloxy-l-methyl-pent-4-enoxy]-3-(fôrtbutoxycarbonylamino)-5-(trifluoromethyl)pyridine-2-carboxylic acid (1.067 g, 83%) as a light yellow gel. ESI-MS m/z cale. 510.19778, found 511.2 (M+l)⁺; Rétention time: 4.21 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 10: fôrf-Butyl A-[6-[(l/?)-2-benzyloxy-l-methyl-pent-4-enoxy]-2-[[[(27?)-2-benzyloxy-2(trifluoromethyl)pent-4-enoyl] amino] carbamoyl]-5-(trifluoromethyl)-3-pyridyl] carbamate

Into a solution of 6-[(17?)-2-benzyloxy-l-methyl-pent-4-enoxy]-3-(/er/butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2-carboxylic acid (1.067 g, 2.0483 mmol), (27?)-2-benzyloxy-2-(trifluoromethyl)pent-4-enehydrazide (600 mg, 2.0814 mmol) and pyridine (733.50 mg, 0.75 mL, 9.2731 mmol) in ethyl acetate (10 mL) was added 1-propanephosphonic anhydride (T3P) (913.99 mg, 1.71 mL of 50 % w/w, 1.4363 mmol) in ethyl acetate. The reaction was then heated to 50 °C and stirred for 1 hour. The reaction was diluted with ethyl acetate (100 mL) and washed with 10% ammonium chloride (30 mL) and brine (30 mL). The organic solution was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue 308 was purified by silica gel chromatography using 0 to 30% ethyl acetate in hexane to fumish tertbutyl TV- [ 6 - [ ( 17?)-2-benzyloxy-1 -methyl-p ent-4-enoxy] -2- [ [ [(27?)-2-benzyloxy-2(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]-5-(trifluoromethyl)-3-pyridyl]carbamate (1.506 g, 92%) as a white solid. ESI-MS m/z cale. 780.2958, found 781.7 (M+l)⁺; Rétention time: 4.54 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 11: terf-Butyl A-[6-[(17î)-2-benzyloxy-l-methyl-pent-4-enoxy]-2-[5-[(lÆ)-l-benzyloxyl-(trifluoromethyI)but-3-enyl]-l,3,4-oxadiazol-2-yl]-5-(trifluoromethyl)-3pyridyl] carbamate

To a solution of tert-butyl jV-[6-[(17?)-2-benzyloxy-l-methyl-pent-4-cnoxy]-2-[[[(27?)-2benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamoyl]-5-(trifluoromethyl)-3pyridyl]carbamate (1.506 g, 1.8903 mmol) and DIEA (742.00 mg, 1 mL, 5.7411 mmol) in acetonitrile (25 mL) was addedp-toluenesulfonyl chloride (440 mg, 2.3079 mmol) at 50 °C. The reaction was heated to 70 °C and stirred for 3 hours. The reaction was cooled to room température and diluted with ethyl acetate (100 mL). The organic solution was washed with 10% ammonium chloride (30 mL) and brine (30 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by silica gel chromatography using 0 to 5% ethyl acetate in hexane to provide ZerZ-butyl 7V-[6-[(lR)-2-benzyloxy-l-methyl-pent-4enoxy]-2-[5-[(lÆ)-l -benzyloxy-1 -(trifluoromethyl)but-3-enyl]-l ,3,4-oxadiazol-2-yl]-5(trifluoromethyl)-3-pyridyl]carbamate (1.278 g, 87%) as a yellow gel. ESI-MS m/z cale. 762.2852, found 763.6 (M+l)⁺; Rétention time: 4.79 minutes; Merck Millipore Chromolith, SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A - water (0.1 % CF3CO2H). Mobile phase B = acetonitrile.

309

Step 12: ZerZ-Butyl 7V-[(6R,121î)-6,ll-dibenzyIoxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]îionadeca-l(18),2,4,8,14,16-hexaen-17yljcarbamate (E!Z mixture)

E/Z mixture

A reaction vial was charged with ZerZ-butyl 7V-[6-[(lR)-2-benzyloxy-l-mcthyl-pent-4enoxy] -2 - [ 5 - [( 1 R) -1 -benzyloxy-1 -(trifluoromethyl)but-3 -enyl] -1,3,4-oxadiazol-2-yl] -5 (trifluoromethyl)-3-pyridyl]carbamate (108 mg, 0.1388 mmol) and anhydrous DCE (20 mL). The reaction mixture was purged with argon for 2 minutes. The vial was sealed and heated to 50 °C. Zhan catalyst-lB (10 mg, 0.0131 mmol) was added to the reaction mixture. The reaction was stirred at 70 °C for 2 days. The reaction was concentrated under reduced pressure. The residue was purified by silica gel chromatography using 0 to 10% ethyl acetate in hexane to fumish ZerZ-butyl N-[(6R, 12R)-6,11 -dibenzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yl]carbamate (E/Z mixture) (88 mg, 56%) as a clear gel. ESI-MS m/z cale. 734.2539, found 735.0 (M+l)⁺; Rétention time: 4.77 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 13: ZerZ-Butyl A-[(67?,127?)-6,ll-dihydroxy-12-methyl-6,15-bis(trinuoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate

diastereomer 1 diastereomer 2

NH N-N

A reaction flask was charged with ZerZ-butyl ;V-[(6/?, 12/?)-6,11-dibenzyloxy-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,8,14,16-hexaen-17-yl]carbamate (E/Zmixture) (88 mg, 0.0779 mmol) in éthanol (5 mL). Then 10% Pd/C (50 mg, 10 % w/w, 0.0470 mmol) was added to the reaction mixture. The reaction was hydrogenated under 1 atm of hydrogen gas ovemight. The catalyst was filtered off through a pad of Celite. The solvent was evaporated under vacuum. The residue was purified by 310 silica gel chromatography (0 to 20% ethyl acetate in hexane) to fumish as the first isomer to elute, teri-butyl Λ^Α-[(6//,12//)-6,1 l-dihydroxy-12-mcthyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-17-yl] carbamate (diastereomer 1) (20 mg, 46%). ESI-MS m/z cale. 556.1757, found 557.2 (M+l)⁺; Rétention time: 3.61 minutes and as the second isomer to elute, terZ-butyl N-[(6R, 12//)-6,1 l-dihydroxy-12methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2) (12 mg, 28%). ESI-MS m/z cale. 556.1757, found 557.3 (M+l)⁺; Rétention time: 3.65 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5-100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H.

Step 14: (67/,127/)-17-Amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,ll-diol (diastereomer 1), Compound 40 v-cf₃OH r^CF3 OH

O. -NH N-N

NH₂ N-N diastereomer 1 diastereomer 1

A micro wave vial was charged with Zeri-butyl N-[(6R, 127/)-6,1 l-dihydroxy-12-methyl6,15-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 1) (133 mg, 0.2295 mmol) and hexafluoroisopropanol (5 mL). The vial was sealed and heated at 100 °C for 2.5 hours in a microwave reactor. The solvent was removed under vacuum. Purification by silica gel chromatography (0 to 30% ethyl acetate in hexane) provided (67/,127/)-17-amino-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1 (18),2,4,14,16-pentaene-6,11-diol (diastereomer 1) (75.8 mg, 71%) as a yellow oil. *H NMR (500 MHz, DMSO-dô) δ 7.77 (s, 1H), 7.59 (s, 1H), 6.35 (s, 2H), 4.88 - 4.69 (m, 1H), 4.59 - 4.52 (m, 1H), 4.50 (d, J= 6.1 Hz, 1H), 2.36 (t,J= 12.5 Hz, 1H), 2.12-1.99 (m, 1H), 1.76-1.60 (m, 3H), 1.54 - 1.40 (m, 2H), 1.35 - 1.27 (m, 1H), 1.24 (d, J= 6.6 Hz, 3H) ppm. ESI-MS m/z cale. 456.12323, found 457.3 (M+l)⁺; Rétention time: 2.17 minutes; LCMS Method: Waters Cortex 2.7u Cis (3.0mm X 50mm), 55 °C; flow: 1.2mL/min; mobile phase: 100% water with 0.1% trifluoroacetic acid then 100% acetonitrile with 0.1% trifluoroacetic acid, gradient of 5% to 100% B over 4 min, with équilibration at 100% B for 0.5 min, then 5% B over 1.5 min.

311

Step 15: (6Æ,12Æ)-17-Amino-12-methyl-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,ll-diol (diastereomer 2), Compound 41

diastereomer 2

A microwavable vial was charged with teri-butyl N-[(6R, 12/?)-6,11 -dihydroxy-12methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2) (75 mg, 0.1321 mmol) and hexafluoroisopropanol (5 mL). The reaction was heated at 100 °C in a microwave reactor for 2.5 hours. The solvent was removed under vacuum. Purification by silica gel chromatography (0 to 10 30% ethyl acetate in hexane) provided (67?,127?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,ll-diol (diastereomer 2) (39.2 mg, 63%) as a yellow oil. *H NMR (500 MHz, DMSO-dô) δ 7.75 (s, 1H), 7.57 (s, 1H), 6.37 (s, 2H), 5.09 - 4.92 (m, 1H), 4.50 (d, J= 6.4 Hz, 1H), 3.75 - 3.58 (m, 1H), 2.36-2.26 (m, 1H), 2.24-2.13 (m, 1H), 2.08 - 1.95 (m, 1H), 1.75-1.62 (m, 1H), 1.62-1.50 (m, 1H), 1.44 - 1.35 (m, 2H), 1.34 (d, J= 6.7 Hz, 3H), 1.22 - 1.07 (m, 1H) ppm. ESI-MS m/z cale. 456.12323, found 457.3 (M+l)⁺; Rétention time: 2.23 minutes; LCMS Method: Waters Cortex 2.7u Cis (3.0mm X 50mm), 55 °C; flow: 1.2mL/min; mobile phase: 100% water with 0.1% trifluoroacetic acid then 100% acetonitrile with 0.1% trifluoroacetic acid, gradient of 5% to 100% B over 4 min, with équilibration at 100% B for 0.5 min, then 5% B over 1.5 min.

312

Example 29: Préparation of (6Æ,127?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,9-diol (diastereomer 1), Compound 42

Step 3

diastereomer 1

313

Step 1: tert-Butyl 7V-[(6.R,121î)-6-benzyloxy-9-hydroxy-12-methyI-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17yl] carbamate (diastereomer 1)

mixture of regioisomeric diastereomers

To a solution of ZerZ-butyl 7V-[(67?,127?)-6-benzyloxy-9-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pcntaen-17-yl]-7V-Zeri-butoxycarbonyl-carbamate and teri-butyl 7V-[(67?,127?)-6-benzyloxy-10hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-7V-fërf-butoxycarbonylcarbamate (mixture of regioisomeric diastereomers) (0.45 g, 0.6027 mmol) in DCM (30 mL) was added silica gel (4.5 g, 74.895 mmol). The mixture was stirred at room température for 2 days. The mixture was concentrated and purified by silica gel chromatography (80 g S1O2, eluting 0 to 10% EtOAc/DCM) twice and silica gel chromatography (80 g S1O2, eluting 10% to 30% EtOAc/heptanes) to provide a 1:3 mixture of two régional isomers, ZerZ-butyl N-[(6R,12R)-6benzyloxy-9-hydroxy-12-methyl-6,15 -bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (minor diastereomer) and ferf-butyl 7V-[(67?,127?)-6-benzyloxy-10-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (major diastereomer) (215 mg, 55%) as a semi solid. *H NMR (400 MHz, CDCI3) δ 9.29 - 9.17 (m, 1H), 9.16 - 9.11 (m, 1H), 7.38 - 7.28 (m, 5H), 4.92 - 4.79 (m, 1H), 4.79 - 4.67 (m, 2H), 4.16 - 3.92 (m, 1H), 2.77 (br dd, J= 13.1, 3.8 Hz, 1H), 2.67 - 2.55 (m, 1H), 2.48 - 2.25 (m, 1H), 1.98 - 1.80 (m, 2H), 1.78 - 1.63 (m, 3H), 1.59 (d, J= 6.4 Hz, 3H), 1.56

314 (s, 9H), 1.50 - 1.46 (m, 1H) ppm. ¹⁹F NMR for the major product (377 MHz, CDCh) δ -63.88 (s, 3F), -74.11 (s, 3F). ^I9F NMR for the minor product (377 MHz, CDCI3) δ -63.86 (s, 3F), 74.07 (s, 3F) ppm; as well as pure terLbutyl 7V-[(67?, 12/?)-6-benzyloxy-9-hydroxy-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 1) (175 mg, 45%) as a colorless oil. *H NMR (400 MHz, CDCI3) δ 9.17 - 9.08 (m, 2H), 7.37 - 7.28 (m, 5H), 5.07 - 4.96 (m, 1H), 4.79 (d, J= 10.8 Hz, 1H), 4.47 (d, J= 10.8 Hz, 1H), 4.40 - 4.29 (m, 1H), 2.69 - 2.56 (m, 1H), 2.30 (dt, J = 14.8, 5.8 Hz, 1H), 2.24 - 2.14 (m, 1H), 2.06 - 1.89 (m, 2H), 1.77 - 1.60 (m, 3H), 1.57 (s, 9H), 1.53 - 1.50 (m, 1H), 1.48 (d, J= 6.1 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCh) δ -63.84 (s, 3F), -73.91 (s, 3F) ppm.

Step 2: tert-Butyl A-[(67?,127?)-6,9-dihydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 1)

diastereomer 1

To a solution of terLbutyl A^z-[(67?,127?)-6-bcnzyloxy-9-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 1) (60 mg, 0.0928 mmol) in MeOH (5 mL) was added 10% palladium on carbon 50% wet (30 mg, 0.0141 mmol). The mixture was stirred under hydrogen (balloon) at room température ovemight. The mixture was filtered through diatomaceous earth and washed with EtOAc. The filtrate was concentrated and the residue was purified by silica gel chromatography (24 g S1O2, eluting 10 to 30% EtOAc/CH2Ch) to afford /eri-butyl 7V-[(67?,127?)-6,9-dihydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 1) (48 mg, 93%) as a white solid. *H NMR (400 MHz, CDCh) δ 9.15 (s, 1H), 9.07 (s, 1H), 6.65 (br s, 1H), 5.07 - 4.94 (m, 1H), 4.15 - 4.06 (m, 1H), 3.29 (br d, J= 10.0 Hz, 1H), 2.58 (br t, J= 11.9 Hz, 1H), 2.52 - 2.37 (m, 2H), 2.21 - 2.08 (m, 1H), 2.05 - 1.96 (m, 1H), 1.82 - 1.66 (m, 2H), 1.56 (s, 9H), 1.54 - 1.47 (m, 4H) ppm. ^I9F NMR (377 MHz, CDCh) δ -63.96 (s, 3F), -78.47 (s, 3F) ppm. ESI-MS m/z cale. 556.17566, found 557.1 (M+l)⁺; Rétention time: 3.62 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

315

Step 3: (67?,1277)-17-Amino-12-niethyl-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,9-diol (diastereomer 1), Compound 42

diastereomer 1

To a solution of Zerf-butyl N-[(6R, 1277)-6,9-dihydroxy-12-methyl-6,15bis(trifluoromethyl)-l 3,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5] nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 1) (38 mg, 0.0683 mmol) in DCM (2 mL) was added TFA (2 mL) dropwise. The mixture was stirred at room température for 1 h. The mixture was concentrated under reduced pressure at room température and co-evaporated with MeOH (2X3 mL). The residue was dissolved in EtOAc (20 mL) and washed with saturated NalICOa (5 mL), dried with Na2SÛ4, filtered and concentrated. The residue was purified by flash chromatography (12 g S1O2, eluting 10 to 50% EtOAc/CFbCh) to afford (67?,127?)-17-amino-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaene-6,9-diol (diastereomer 1) (23 mg, 74%) as a pale-yellow solid. 'H NMR (400 MHz, DMSO-dô) δ 7.76 (s, 1H), 7.59 (s, 1H), 6.35 (s, 2H), 5.00 - 4.89 (m, 1H), 4.57 (d, J= 5.1 Hz, 1H), 3.98 - 3.87 (m, 1H), 2.37 - 2.23 (m, 3H), 2.07 - 1.98 (m, 1H), 1.62 - 1.48 (m, 2H), 1.42 1.28 (m, 5H) ppm. ^I9F NMR (377 MHz, DMSO-d₆) δ -62.51 (s, 3F), -76.36 (s, 3F) ppm. ESIMS m/z cale. 456.12323, found 457.1 (M+l)⁺; Rétention time: 2.94 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

316

Example 30: Préparation of (6/î,12/î)-17-amino-12-methyl-6,15-bis(trifluoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,10-diol (diastereomer 1), Compound 43

(1:3) mixture of regioisomeric diastereomers

regioisomeric diastereomer 1 regioisomeric diastereomer 1

diastereomer 1

Step 1: ter/-Butyl7V-[(6/î,12Æ)-6,10-dihydroxy-12-methyl-6,15-bis(trifluoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (regioisomeric diastereomer 1)

- OH

(1:3) mixture of regioisomeric diastereomers

regioisomeric diastereomer 1

To a solution of a 1:3 mixture of Ze/7-butyl 7V-[(6/?,12Æ)-6-benzyloxy-9-hydroxy-1210 methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[l 2.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate and /ert-butyl A^L[(6/?,12/?)-6-benzyloxy-10-hydroxy12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (1:3 mixture of regioisomeric diastereomers) (80 mg, 0.1237 mmol) in MeOH (3 mL) was added 10% palladium on carbon 50% wet (30 mg, 0.0141 mmol). Air was replaced by nitrogen through vacuum for 3 times. The mixture was stirred under hydrogen (balloon) at room température ovemight. The mixture was filtered through diatomaceous earth and washed with EtOAc. The filtrate was concentrated and purified by silica gel chromatography (40 g S1O2, eluting 0 to 30% EtOAc/hcptanes) twice to afford Ze/7-butyl N317

[ (62?, 12 J?)-6,10-dihydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclofl 2.3.1.12,5]nonadeca-l ( 18),2,4,14,16-pentaen-17-yl]carbamate (regioisomeric diastereomer 1) (30 mg, 44%) as a colorless oil. *H NMR (400 MHz, CDCh) δ 9.11 (s, 1H), 9.06 (s, 1H), 4.87 - 4.74 (m, 1H), 4.25 - 4.09 (m, 1H), 3.94 (br s, 1H), 2.85 (br dd, J= 13.3, 3.8 Hz, 1H), 2.47 - 2.25 (m, 2H), 2.01 - 1.79 (m, 3H), 1.73 - 1.65 (m, 1H), 1.62 - 1.47 (m, 14H) ppm. ^,9F NMR (377 MHz, CDCI3) δ -63.88 (s, 3F), -77.40 (s, 3F) ppm. ESI-MS m/z cale. 556.17566, found 557.1 (M+l)⁺ ; Rétention time: 3.62 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: [(6Æ,127?)-17-Amino-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-10-yl] 2,2,2trifluoroacetate (regioisomeric diastereomer 1)

regioisomeric diastereomer 1

To a solution of ZerZ-butyl 7V-[(67?,127?)-6,10-dihydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (regioisomeric diastereomer 1) (30 mg, 0.0539 mmol) in DCM (2 mL) was added TFA (2.9600 g, 2 mL, 25.960 mmol). The mixture was stirred at room température for 1 h. The mixture was concentrated under reduced pressure at 40 °C. The residue was purifîed by silica gel chromatography (24 g S1O2, eluting 0 to 30% EtOAc/heptanes) to afford [(67?, 127?)-17-amino-6-hydroxy-12-methyl-6,l 5-bis(trifluoromethyl)-l 3,19-dioxa-3,4,l 8triazatricyclo[ 12.3.1.12,5]nonadeca-l (18),2,4,14,16-pentaen-10-yl] 2,2,2-trifluoroacetate (regioisomeric diastereomer 1) (27 mg, 91%) as a pale-yellow oil. *H NMR (400 MHz, CDCI3) δ 7.46 (s, 1H), 5.59 (tdd, J= 8.5, 3.9, 2.3 Hz, 1H), 5.37 (s, 2H), 4.58 (quin, J= 6.7 Hz, 1H), 3.82 (s, 1H), 3.01 (dd, J= 14.3, 3.8 Hz, 1H), 2.64 (br t, J= 12.2 Hz, 1H), 2.39 - 2.28 (m, 1H), 2.11 2.01 (m, 1H), 2.00 - 1.71 (m, 4H), 1.51 (d, J = 6.4 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCh) δ -64.00 (s, 3F), -75.28 (s, 3F), -77.43 (s, 3F) ppm.

318

Step 3: (67?,127?)-17-Amino-12-methyI-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,10-diol (diastereomer 1), Compound 43

regioisomeric diastereomer 1

diastereomer 1

To a solution of [(67?,127?)-17-amino-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-10-yl] 2,2,2trifluoroacetate (regioisomeric diastereomer 1) (27 mg, 0.0489 mmol) in THF (2 mL) at 0 °C was added a solution of NaOH (8.6 mg, 0.2150 mmol) in H2O (0.5 mL). The mixture was stirred at 0 °C for 1 h, treated with saturated NaHCCh (5 mL) and brine (3 mL). The mixture was extracted with EtOAc (3X10 mL). The combined organic layers were dried with Na2SO4, filtered and concentrated. The residue was purifïed by silica gel chromatography (12 g S1O2, eluting 20 to 50% EtOAc/CHzCh) to afford (67?,127?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaene-6,10-diol (diastereomer 1) (17 mg, 76%) as a pale-yellow solid. *H NMR (400 MHz, DMSO-dô) δ 7.77 (s, 1H), 7.61 (s, 1H), 6.37 (s, 2H), 4.76 - 4.65 (m, 1H), 4.52 (d, J= 5.1 Hz, 1H), 3.97 - 3.84 (m, 1H), 2.60 (br dd, J= 12.3,4.5 Hz, 1H), 2.43-2.32 (m, 1H), 2.16-2.02 (m, 1H), 1.75- 1.50 (m, 4H), 1.44 (d, J= 6.4 Hz, 3H), 1.31-1.22 (m, 1H) ppm. ^,9F NMR (377 MHz, DMSO-d₆) δ -62.55 (s, 3F), -76.26 (s, 3F) ppm. ESI-MS m/z cale. 456.12323, found 457.1 (M+l)⁺; Rétention time: 2.87 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

319

Example 31: Préparation of (67?,127?)-17-amino-8,9-dideuterio-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol, Compound 44

Step 1

Step 3

E/Z mixture

Step 5

Step 1: (25)-Hex-5-en-2-ol

To a yellow solution of allyl(bromo)magnesium (103 mL of 1 M, 103.00 mmol) in diethyl ether was added copper bromide (975 mg, 6.7968 mmol) at 0 °C, then the black mixture was stirred at -78 °C. After 5 min, a solution of (2S)-2-methyloxirane (1.9896 g, 2.4 mL, 34.257 mmol) in THF (30 mL) was added dropwise with a dropping tunnel over 20 min at -78 °C. Then the black mixture was stirred at -78 °C for 30 min. Then methanol (16 mL) was added at -78 °C, followed by aqueous hydrogen chloride (2M, 80 mL) and then stirred at room température for 5 min. MTBE (120 mL) was then added, the aqueous layer was separated and extracted with MTBE (2 X 120 mL). The organic layers were washed with aqueous hydrogen chloride (IM, 50 mL), water (50 mL), aqueous sodium thiosulphate (10%, 50 mL) and again with water (50 mL). The organic layers were dried over sodium sulphate and concentrated under vacuum. The oil residue was dry loaded with silica gel and purified by liquid chromatography on silica gel eluting with portions of ethyl acetate (0-30%) in heptanes to afford (25)-hex-5-en-2-ol (1.66 g, 44%) as

320 a light yellow oil. 'H NMR (400 MHz, CDC1₃) δ 5.93 - 5.75 (m, 1H), 5.14 - 4.89 (m, 2H), 3.92 3.72 (m, 1H), 2.28 - 2.01 (m, 2H), 1.66 - 1.45 (m, 2H), 1.24 - 1.18 (m, 3H) ppm.

Step 2: tert-Butyl A-[2-[5-[(17?)-l-benzyloxy-l-(trifIuoromethyl)but-3-enyl]-l,3,4-oxadiazoI2-yl]-6-[(LR)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-3-pyridyl]-A^r-tert-butoxycarbonyIcarbamate

To a solution of teri-butyl 7V-[2-[5-[(17?)-l-benzyIoxy-l-(trifluoromethyl)but-3-enyl]l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ieri-butoxycarbonylcarbamate (325 mg, 0.4818 mmol) and (2S)-hex-5-en-2-ol (160 mg, 1.4377 mmol) in toluene (4 mL) was treated with triphenylphosphine (264 mg, 1.0065 mmol) followed by DIAD (205.40 mg, 0.2 mL, 1.0158 mmol) at room température. The yellow solution was stirred at room température for ovemight. The yellow suspension was concentrated under vacuum, then drypacked on silica with DCM. Purification by silica gel chromatography (0 - 30% EtOAc/heptanes) gave teri-butyl N-[2-[5-[(lJ?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]l,3,4-oxadiazol-2-yl]-6-[(17î)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7V-terrbutoxycarbonyl-carbamate (320 mg, 74%) as a yellow oil. 'H NMR (400 MHz, CDCI3) δ 7.83 (s, 1H), 7.35 (d, J= 8.3 Hz, 5H), 5.93 (td, 17.0, 7.1 Hz, 1H), 5.86 - 5.74 (m, 1H), 5.37 - 5.28 (m, 1H), 5.28 - 5.15 (m, 2H), 5.03 - 4.88 (m, 2H), 4.82 (d, J= 10.8 Hz, 1H), 4.63 (d, J= 10.8 Hz, 1H), 3.28 - 3.11 (m, 2H), 2.27 - 2.02 (m, 2H), 1.97 - 1.84 (m, 1H), 1.79 - 1.66 (m, 1H), 1.43 (s, 18H), 1.38 (d, J= 6.1 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -64.13 to -64.28 (m, 3F), -73.32 to -73.43 (m, 3F) ppm.

Step 3: ferf-Butyl A-[(6Æ,127?)-6-benzyloxy-12-methyI-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yl]-7V-fertbutoxycarbonyl-carbamate (E!Z mixture)

E/Z mixture

321

To a stirring solution of ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-[(l/?)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7VZerZ-butoxycarbonyl-carbamate (320 mg, 0.3582 mmol) in 1,2-dichloroethane (175 mL) was degassed with bubbling with nitrogen gas for 24 hours. To the solution at 60 °C was added Zhan catalyst-lB (18 mg, 0.0245 mmol) then the reaction was stirred at this température for 40 minutes. Then, an equal amount of Zhan catalyst-lB (18 mg, 0.0245 mmol) was added and stirring continued at 60 °C for 2.5 hours. Once cooled to room température, the catalyst was quenched with a few drops of DMSO (about 5-6) and the reaction was concentrated under reduced pressure. The residue was purified by silica gel chromatography using a gradient from 0% to 90% ethyl acetate in heptanes to afford ZerZ-butyl 7V-[(67?,127?)-6-benzyloxy-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,8,14,16-hexaen-17-yl]-7V-/erZ-butoxycarbonyl-carbamate (E/Zmixture) (290 mg, 73%) as white solid which still contained a small amount of an unknown impurity. 'H NMR (400 MHz, CDCh) δ 7.82 (s, 1H), 7.34 - 7.24 (m, 5H), 5.84 (dt, J= 14.9, 7.2 Hz, 1H), 5.76 5.64 (m, 1H), 5.28-5.15 (m, 1H), 4.79 (d, J = 11.0 Hz, 1H), 4.50 (d,J= 11.2 Hz, lH),3.13(dd, J= 14.4, 7.3 Hz, 1H), 2.77 (dd, J= 14.4, 6.8 Hz, 1H), 2.33 (dt, J= 13.7, 6.8 Hz, 1H), 2.13 - 2.03 (m, 1H), 1.98 (dd, J= 13.0, 7.1 Hz, 1H), 1.72 - 1.60 (m, 1H), 1.50 (d, J= 6.4 Hz, 3H), 1.46 (s, 18H)ppm. ¹⁹F NMR (377 MHz, CDCh) δ -63.96 (s, 3F), -74.55 (s, 3F) ppm. ESI-MS m/z cale. 728.26447, found 573.2 (M-155, -Boc, -'Bu)’'·; Rétention time: 4.48 minutes; LCMS Method: XBridge Cis 4.6 x 75mm 5pm, initial gradient at 95% NH4HCO3 / 5% acetonitrile 6 min run with 1 min équilibration, gradient 0 to 3 min at 95% acetonitrile and held for 3 minutes, with a flow rate of 1.5 mL/min.

Step 4: ZerZ-Butyl A-ZerZ-butoxycarbonyI-A-[(67?,127?)-8,9-dideuterio-6-hydroxy-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-17-yl]carbamate

To a solution of ZerZ-butyl ^-[(ô/L^/ij-ô-benzyloxy-U-methyl-ô.lSbis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,8,14,16hexaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate (E/Z mixture) (140 mg, 0.1921 mmol) in

322

CD3OD (5.6 mL) under nitrogen was added 10% palladium on carbon (36 mg, 0.0338 mmol). Nitrogen was replaced with deuterium gas through vacuum for 3 cycles. The mixture was stirred at room température under deuterium atmosphère (balloon) ovemight. The mixture was filtered through a pad of Celite and washed with EtOAc (30 mL) and then concentrated by évaporation under reduced pressure to give ZerZ-butyl /V-ZerZ-butoxycarbonyl-A-[(67î,127?)-8,9-dideuterio-6hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (127 mg, 88%) as a colorless oil. 'H NMR (400 MHz, CDCI3) δ 7.83 (s, 1H), 5.00 - 4.85 (m, 1H), 3.95 3.65 (m, 1H), 2.73 - 2.55 (m, 1H), 2.34 - 2.22 (m, 1H), 2.21 - 2.10 (m, 1H), 2.08 - 1.96 (m, 1H), 1.67 - 1.19 (m, 25H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.99 (s, 3F), -77.58 (s, 3F) ppm. ESI-MS m/z cale. 642.2457, found 487.1 (M-155)⁺; Rétention time: 4.01 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) at 1.2 mL/min.

Step 5: (6/î,12R)-17-Amino-8,9-dideuterio-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa 3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol, Compound 44

To a solution of ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(67?,127?)-8,9-dideuterio-6-hydroxy12-methyl-6,15 -bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-17-yl]carbamate (127 mg, 0.1682 mmol) in dichloromethane (1.4 mL) was added 2,2,2-trifluoroacetic acid (2.0720 g, 1.4 mL, 18.172 mmol). The mixture was stirred at room température for 2.5 hours. The mixture was then diluted with dichloromethane (5 mL), then concentrated by évaporation under reduced pressure. The residue was dissolved in ethyl acetate (100 mL), washed with aqueous saturated solution of sodium bicarbonate (3X15 mL) and brine (1X15 mL) then dried with anhydrous sodium sulphate, filtered and concentrated by évaporation under reduced pressure. The residue was purified by silica gel chromatography (12 g S1O2, 0 to 25% portions of ethyl acetate in heptanes). The product was dissolved in a minimum amount of acetonitrile and water, and freeze-dried ovemight to afford (67?, 127?)-17amino-8,9-dideuterio-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (70 mg, 94%) as a yellow solid. ’H NMR (400 MHz, DMSO-d₆) δ 7.77 (s, 1H), 7.56 (s, 1H), 6.35 (s, 2H), 4.85 - 4.73 (m,

323

1H), 2.49 - 2.43 (m, 1H), 2.32 - 2.20 (m, 1H), 2.14 - 2.04 (m, 1H), 1.73 - 1.66 (m, 1H), 1.58 1.37 (m, 3H), 1.34 (d, J= 6.1 Hz, 3H), 1.22 - 1.12 (m, 1H) ppm. ¹⁹F NMR (377 MHz, DMSOd₆) δ -62.50 (s, 3F), -76.40 (s, 3F) ppm. ESI-MS m/z cale. 442.1409, found 443.1 (M+l)⁺; Rétention time: 3.53 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5

- 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Example 32: Préparation of (67?,12./?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,9-dioI (diastereomer 2), Compound 45

diastereomer 1

Step 1

diastereomer 1

Step 3

diastereomer 2

Step 4

diastereomer 2

Step 1: /‘<?rZ‘-ButyIA^r-[(67?,12Æ)-6-benzyloxy-12-methyI-9-oxo-6,15-bis(trifluoromethyI)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17yl] carbamate

324

To a solution of ZerZ-butyl 2V-[(6R,12R)-6-benzyloxy-9-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (147 mg, 0.2274 mmol) in DCM (12 mL) was added NaHCCh (320 mg, 3.8092 mmol). The mixture was cooled to 0 °C and Dess-Martin Periodinane (108 mg, 0.2546 mmol) was added. The mixture was slowly warmed to room température and stirred at room température ovemight. The mixture was treated with saturated NaHCCL (8 mL) and extracted with CH2CI2 (3X10 mL). The combined organic layers were dried with Na2SOi, fïltered and concentrated. The residue was purified by flash chromatography (24 g S1O2, eluting 10 to 30% EtOAc/heptanes) to afford ZerZ-butyl 7V-[(6R,12R)-6-benzyloxy-12-methyl-9-oxo6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (120 mg, 82%) as a colorless oil. 'H NMR (400 MHz, CDCI3) δ 9.48 (s, 1H), 9.14 (s, 1H), 7.36 - 7.27 (m, 5H), 5.10 - 5.00 (m, 1H), 4.73 (d, 11.2

Hz, 1H), 4.57 (d, J= 11.0 Hz, 1 H), 3.06 - 2.91 (m, 3H), 2.71 (ddd,J=16.9, 10.1,2.3 Hz, 1H), 2.60 - 2.47 (m, 2H), 2.29 - 2.18 (m, 1H), 1.88 - 1.76 (m, 1H), 1.55 (s, 9H), 1.48 (d, J= 6.4 Hz, 3H) ppm. ^l9F NMR (377 MHz, CDCI3) δ -63.85 (s, 3F), -74.57 (s, 3F) ppm.

Step 2: terZ-Butyl A-[(67?,12R)-6-benzyloxy-9-hydroxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17yljcarbamate (diastereomer 2)

To a solution of ie/7-butyl A^r-[(6R,12R)-6-benzyloxy-12-methyl-9-oxo-6,15bis(trifluorom ethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (120 mg, 0.1862 mmol) in EtOH (10 mL) at 0 °C was added a solution of NaBH4 (4.8 mg, 0.1269 mmol) in EtOH (1 mL) dropwise. The mixture was stirred at room température for 1 h. More NaBH4 (4 mg, 0.1057 mmol) was added. The mixture was stirred at room température for 20 min. Acetone (1 mL) was added. The mixture was stirred at room température for 5 min. Sat. NaHCÜ3 (1 mL) was added. The mixture was concentrated to remove EtOH. The residue was treated with water (2 mL) and extracted with CH2CI2 (3X10 mL). The combined organic layers were dried with Na2SÜ4, fïltered and concentrated. The residue was purified by silica gel chromatography (80 g S1O2, eluting 10 to 30% EtOAc/heptanes) to afford two diastereomers. The desired diastereomer 2, ZerZ-butyl N325

[(67?, 127?)-6-benzyloxy-9-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2) (60 mg, 50%) as a colorless oil. Ή NMR (400 MHz, CDC1₃) δ 9.18 (s, 1H), 9.12 (s, 1H), 7.36 7.27 (m, 5H), 4.85 - 4.67 (m, 3H), 4.01 - 3.90 (m, 1H), 2.79 (t, J= 12.3 Hz, 1H), 2.51 - 2.31 (m, 2H), 2.25 - 2.12 (m, 1H), 1.97 - 1.78 (m, 2H), 1.73 - 1.63 (m, 2H), 1.56 (s, 9H), 1.54-1.51 (m, 1H), 1.49 (d, J= 6.4 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.87 (s, 3F), -74.08 (s, 3F) ppm. ESI-MS m/z cale. 646.2226, found 647.2 (M+l)⁺ ; Rétention time: 4.14 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

The other enantiomer, diastereomer 1, was the first to elute Zeri-butyl 7V-[(67?,127?)-6benzyloxy-9-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (60 mg, 50%) as a colorless oil. Ή NMR (400 MHz, CDCI3) δ 9.16 - 9.07 (m, 2H), 7.36 - 7.28 (m, 5H), 5.01 (ddd, J= 9.5, 6.4, 3.2 Hz, 1H), 4.79 (d, J= 10.8 Hz, 1H), 4.47 (d, J= 10.5 Hz, 1H), 4.40 - 4.27 (m, 1H), 2.63 (dt, J= 14.7, 7.7 Hz, 1H), 2.30 (dt, J= 14.8, 5.7 Hz, 1H), 2.24 - 2.14 (m, 1H), 2.05 1.92 (m, 2H), 1.77 - 1.68 (m, 3H), 1.57 (s, 9H), 1.54-1.51 (m, 1H), 1.48 (d, J= 6.4 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.85 (s, 3F), -73.92 (s, 3F) ppm. ESI-MS m/z cale. 646.2226, found 647.2 (M+l)⁺; Rétention time: 4.16 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 3: tert-Butyl 7V-[(67?,12Æ)-6,9-dihydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2)

diastereomer 2 diastereomer 2

To a solution of tert-butyl A^-[(6/?,12/?)-6-benzyloxy-9-hydroxy-12-mcthyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 2) (60 mg, 0.0928 mmol) in MeOH (5 mL) was added 10% palladium on carbon 50% wet (30 mg, 0.0141 mmol). The mixture was stirred under hydrogen (balloon) at room température ovemight. The mixture was filtered through diatomaceous earth and washed with EtOAc. The filtrate was concentrated and purification by silica gel chromatography (24 g S1O2, eluting 10 to 30% EtOAc/CHiCh) provided ZerZ-butyl N326

[(67?, 127?)-6,9-dihydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2) (48 mg, 93%) as a white solid. Ή NMR (400 MHz, CDCh) δ 9.15 (s, 1H), 9.07 (s, 1H), 6.65 (br s, 1H), 5.07 - 4.94 (m, 1H), 4.15 - 4.06 (m, 1H), 3.29 (br d, J= 10.0 Hz, 1H), 2.58 (br t, J= 11.9 Hz, 1H), 2.52 - 2.37 (m, 2H), 2.21 - 2.08 (m, 1H), 2.05 - 1.96 (m, 1H), 1.82 - 1.66 (m, 2H), 1.56 (s, 9H), 1.54 - 1.47 (m, 4H) ppm. ¹⁹F NMR (377 MHz, CDCh) δ -63.96 (s, 3F), -78.47 (s, 3F) ppm. ESI-MS m/z cale. 556.17566, found 557.1 (M+l)⁺; Rétention time: 3.62 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H₂O (0.1% formic acid) 1.2 mL/min.

Step 4: (67?,127?)-17-Amino-12-methyI-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,9-diol (diastereomer 2), Compound 45

diastereomer 2 diastereomer 2

To a solution of ZerZ-butyl jV-[(67?,127?)-6,9-dihydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 2) (38 mg, 0.0683 mmol) in CH₂C1₂ (2 mL) at 0 °C was added TFA (1.4800 g, 1 mL, 12.980 mmol). The mixture was stirred at 13-15 °C for 1 h. MeOH (2 mL) was added. The mixture was concentrated and co-evaporated with MeOH (2X3 mL). The residue was dissolved in EtOAc (20 mL) and washed with saturated NaHCOj (4 mL), dried with Na₂SO4, filtered and concentrated. The residue was purified by silica gel chromatography (12 g SiO₂,10 to 30% EtOAc/CH₂Cl₂) and the product was freeze dried to afford (6R,\2R)-\1amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,9-diol (diastereomer 2) (19 mg, 57%) as a pale-yellow solid. ^lH NMR (400 MHz, DMSO-d6) δ 7.75 (s, 1H), 7.61 (s, 1H), 6.33 (s, 2H), 4.70 - 4.55 (m, 2H), 3.73 - 3.62 (m, 1H), 2.46 - 2.29 (m, 2H), 2.19 - 2.08 (m, 1H), 2.02 1.90 (m, 1H), 1.71 - 1.45 (m, 3H), 1.35 (d, J= 6.1 Hz, 3H), 1.30 - 1.21 (m, 1H) ppm. ¹⁹FNMR (377 MHz, DMSO-dô) δ -62.47 (s, 3F), -76.20 (br. s., 3F) ppm. ESI-MS m/z cale. 456.1232, found 457.1 (M+l)⁺; Rétention time: 2.89 minutes; Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

327

Example 33: Préparation of (61î,12/2)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,10-diol (diastereomer 2), Compound 46

Step 2

Step 3

diastereomer 2

Step 1: te/7-Butyl A-[(67?,127?)-6-hydroxy-12-methyl-10-oxo-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17yl]carbamate

To a solution of ZerAbutyl N-[(6R, 1272)-6,10-dihydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]carbamate (65 mg, 0.1168 mmol) in CH2CI2 (8 mL) was added NaHCCh (168 mg, 1.9998 mmol). The mixture was cooled to 0 °C and Dess-Martin periodinane (51 mg, 0.1202 mmol) was added. The mixture was allowed to slowly warm to room température and stirred at room température ovemight. The mixture was treated with saturated NaHCOj (10 mL) and extracted with CH2CI2 (3X10 mL). The combined organic layers were dried with Na2SÛ4, filtered and concentrated. The residue was purified by silica gel chromatography (12 g S1O2, eluting 10 to 60% EtOAc/heptanes) to afford Zc/7-butyl /V-[(6/?,127?)-6-hydroxy-12-mcthyl-10oxo-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (38 mg, 59%) as a white solid. 'H NMR (400 MHz, CDCI3) δ 9.24 (s, 1H), 9.14 (s, 1H), 5.24 (quind, J= 6.4, 2.0 Hz, 1H), 4.75 (s, 1H), 3.56 (dd, J= 18.3,1.7 Hz, 1H), 2.75 - 2.59 (m, 3H), 2.47 (dd, J= 18.3, 6.1 Hz, 1H), 2.25 - 2.07 (m, 2H), 2.00

328

- 1.87 (m, 1H), 1.55 (s, 9H), 1.53 (d, J= 6.6 Hz, 3H) ppm. ^,9F NMR (377 MHz, CDCh) δ 63.83 (s, 3F), -78.16 (s, 3F) ppm. ESI-MS m/z cale. 554.16003, found 499.1 (M-55)⁺; Rétention time: 2.36 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 3 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: terf-Butyl A-[(6R,12R)-6,10-dihydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2)

To a solution of Ze/7-butyl ;V-[(67?,12/?)-6-hydroxy-12-methyl-10-oxo-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]carbamate (33 mg, 0.0595 mmol) in EtOH (4 mL) at 0 °C was added a solution of NaBH4 (2.1 mg, 0.0555 mmol) in EtOH (0.5 mL). The mixture was at 0 °C for 4 h. Acetone (1 mL) was added. The mixture was stirred at 0 °C for 5 min. Sat. NaHCCh (5 mL) was added. The mixture was extracted with EtOAc (3X10 mL). The combined organic layers were dried with Na2SO4, filtered and concentrated. The residue was purified by silica gel chromatography (12 g S1O2, eluting 0 to 50% EtOAc/heptanes) to afford terZ-butyl 7V-[(6R,12R)-6,10-dihydroxy12-methyl-6,15 -bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 2) (10 mg, 30%) as a pale-yellow solid. ’H NMR (400 MHz, CDCI3) δ 9.13 (s, 2H), 5.27 - 5.16 (m, 1H), 4.06 - 3.95 (m, 1H), 3.85 (s, 1H), 2.58 - 2.38 (m, 3H), 2.34 - 2.23 (m, 1H), 2.04 - 1.90 (m, 1H), 1.78 - 1.63 (m, 4H), 1.56 (s, 9H), 1.49 (d, J= 6.4 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.85 (s, 3F), -77.65 (s, 3F) ppm. ESI-MS m/z cale. 556.17566, found 557.2 (M+l)⁺; Rétention time: 3.5 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

329

Step 3: (67?,127/)-17-Amino-12-methyI-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,10-diol (diastereomer 2), Compound 46

To a solution of Zeri-butyl 2V-[(67/,12Æ)-6,10-dihydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 2) (20 mg, 0.0359 mmol) in CH2CI2 (2 mL) at 0 °C was added TFA (740.00 mg, 0.5 mL, 6.4899 mmol) dropwise. The mixture was stirred at 0 °C for 1 h and at 9-13 °C for 1 h. The mixture was treated with MeOH (1 mL) and concentrated at rt, coevaporated with MeOH (2X2 mL). The residue was treated with saturated NaHCOj (5 mL) and extracted with CH2CI2 (3X10 mL). The combined organic layers were dried with Na2SO4, filtered and concentrated. Purification by silica gel chromatography (24 g S1O2, eluting 10 to 30% EtOAc/CH2C12) provided trifluoroacetate formed with the secondary alcohol (about 3 mg) and the first crop of the desired product. Trifluoroacetate was dissolved in THF (1 mL) and treated with 1 N aqueous NaOH (0.1 mL) at room température for 20 min. The mixture was treated with saturated NaHCOa (1 mL) and extracted with CH2CI2 (2X5 mL). The combined organic layers were dried with Na₂SO4, filtered and concentrated. Purification by silica gel chromatography (24 g SiO₂, eluting 10 to 30% EtOAc/CH₂Cl₂) provided the second crop. The two crops were combined and freeze dried to give (6/?,127?)-17-amino-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaene-6,10-diol (diastereomer 2) (10 mg, 59%) as a pale-yellow solid. *H NMR (400 MHz, DMSO-de) δ 7.76 (s, 1H), 7.58 (s, 1H), 6.37 (s, 2H), 5.23 - 5.14 (m, 1H), 4.38 (d, J= 4.9 Hz, 1H), 3.63 - 3.52 (m, 1H), 2.47 - 2.42 (m, 1H), 2.34 - 2.25 (m, 1H), 2.14 - 2.00 (m, 2H), 1.84 1.70 (m, 1H), 1.55 -1.65 (m, 1H), 1.45 - 1.37 (m, 1H), 1.35 (d, J= 6.6 Hz, 3H), 1.32 - 1.25 (m, 1H) ppm. ¹⁹F NMR (377 MHz, DMSO-d6) δ -62.50 (s, 3F), -77.01 (s, 3F) ppm. ESI-MS m/z cale. 456.12323, found 457.1 (M+l)⁺; Rétention time: 2.76 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Example 34: Préparation of (6R)-17-amino-12-(cyclopropyImethyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (enantiomer 1), Compound 47, and (6Æ)-17-amino-12-(cyclopropylmethyl)330

6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 48

enantiomer 1 enantiomer 2

Step 1: 2-CycIopropyl-7V-methoxy-2V-methyl-acetamide

To a stirred solution of 2-cyclopropylacetic acid (10 g, 99.884 mmol) in DCM (100 mL) was added CDI (18.5 g, 114.09 mmol) portion wise and the reaction mixture was stirred for 2 h at 25 °C. Then, TV.O-dimethyl hydroxylamine (hydrochloride sait) (10.000 g, 102.52 mmol) was added in one portion. The reaction mixture was stirred for 12 h at 25 °C, poured into ice-cold water (100 mL) and extracted with ethyl acetate (300 X 2). The combined organic layer was washed with brine (100 mL), dried over anhydrous NaiSCU and concentrated under reduced pressure. The crude product was purified by column chromatography over silica gel (230-400 mesh) using 12% (ethyl acetate-hexane) as eluent to afford 2-cyclopropyl-7V-methoxy-7V-methylacetamide (12 g, 84%). Ή NMR (400 MHz, DMSO-d₆) δ 3.63 (s, 3H), 3.08 (s, 3H), 2.29 (d, J =

6.8 Hz, 2H), 0.991-0.92 (m, 1H), 0.46 - 0.42 (m, 2H), 0.12 - 0.09 (m, 2H) ppm. ESI-MS m/z

331 cale. 143.09464, found 144.0 (M+l)⁺; Rétention time: 2.29 minutes; LCMS Method: TCG method 10: Zorbax Ext Cis, 50 X 4.6 mm 5pm, 5 min run, 10 - 90% acetonitrile in water (10 μΜ NH4OAC modifier), 1.2mL/min.

Step 2: l-Cyclopropylpent-4-en-2-one

A solution of 2-cyclopropyl-7V-methoxy-JV-methyl-acetamide (0.5 g, 3.4920 mmol) in THF (10.5 mL) was cooled to -78 °C and treated with a solution of allylmagnesium bromide (5 mL of 1 M, 5.0000 mmol) in Et2Ü at -78 °C over 22 minutes. The reaction was held at this température for 2 h and 10 min. and then treated with saturated aqueous NH4CI (50 mL) at -78° C. The organic layer was decanted, and the aqueous phase was extracted with DCM (4 X 40 mL). The combined organics were washed with NH4CI (30 mL), dried over Na2SÜ4 and concentrated in vacuo carefully at 25 °C, to give a crude residue (0.564 g). The residue contained the product l-cyclopropylpent-4-en-2-one (564 mg, 77%) as a clear oil. 'H NMR (500 MHz, Chloroform-d) δ 6.00 - 5.86 (m, 1H), 5.22 - 5.08 (m, 2H), 3.22 (d, J= 7.0 Hz, 2H), 2.32 (d, J= 6.9 Hz, 2H), 1.07 - 0.93 (m, 1H), 0.61 - 0.53 (m, 2H), 0.11 (q, J= 5.2 Hz, 2H) ppm.

Step 3: l-Cyc!opropylpent-4-en-2-ol

A slurry of lithium aluminum hydride (1.24 g, 32.671 mmol) in THF (36 mL) was cooled to -78 °C under nitrogen. The slurry was treated dropwise with a solution of 1-cyclopropylpent4-en-2-one (3 g, 21.743 mmol) in THF (7 mL) over 40 minutes and held at this température an additional 2 h. The slurry was treated carefully dropwise with H2O (20 mL) at -78 °C and then diluted with DCM (150 mL) and warmed to room température by removing the bath, and stirred for 30 minutes at rt. A white gel was generated as the second component of the biphasic mixture. The mixture was filtered through Celite and rinsed with CH2Q2 (2 X 200 mL). The filtrate was concentrated in vacuo (28° C, 140 mmHg) to obtain a light yellow crude oil (2.99 g). The crude residue was purified by silica gel chromatography (120 g SiCh, loaded in hexanes, eluted with 0 - 10% Et2Û in hexanes over a 40 minute gradient). The pure fractions were combined and the solvents were carefully removed under reduced pressure (28 °C, 140 mm Hg) to obtain the target l-cyclopropylpent-4-en-2-ol (1.41 g, 51%) as a transparent colorless oil. *H NMR (500 MHz, Chloroform-d) δ 5.85 (dddd, J= 17.0, 10.4, 7.8, 6.5 Hz, 1H), 5.18 - 5.10 (m, 2H), 3.77 (tt, J= 7.6, 4.7 Hz, 1H), 2.40-2.31 (m, 1H), 2.19 (dtt, J= 14.0, 7.8, 1.2 Hz, 1H), 1.47

332

-1.31 (m, 2H), 0.82-0.70 (m, 1H), 0.55-0.41 (m, 2H), 0.12 (dtd, J=9.2, 4.8, 3.3 Hz, 1H), 0.08 - 0.03 (m, 1H) ppm.

Step 4: terf-Butyl 7V-[2-[5-[(117)-l-benzyIoxy-l-(trifluoromethyI)pent-4-enyl]-l,3,4oxadiazol-2-yI]-6-[l-(cyclopropylmethyl)but-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7V-fôrtbutoxycarbonyl-carbamate

A solution of terZ-butyl 7V-[2-[5-[(l7?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-terAbutoxycarbonyl-carbamate (500 mg, 0.7116 mmol) and l-cyclopropylpent-4-en-2-ol (200 mg, 1.7830 mmol) in toluene (5 mL) was treated with DIAD (261.00 mg, 0.25 mL, 1.2908 mmol) dropwise over one minute followed by triphenylphosphine (289 mg, 1.1019 mmol) in one portion at room température under argon. The reaction stirred for a total of 24 h at rt. The reaction was treated with NaHCCh (20 mL) and diluted with EtOAc (100 mL). Then organic layer was washed with saturated aqueous NaHCO₃(1 X 20 mL), saturated aqueous NH4CI (1 X 20 mL) and brine (1 X 20 mL) then dried over NaiSO4, filtered and concentrated in vacuo to obtain a yellow crude oil (1.13 g). The crude residue was purified by silica gel chromatography (80 g S1O2, dry loaded, eluted with 0-50% EtOAc in hexanes over a 20 min. gradient). The pure fractions were combined and the solvent was evaporated in vacuo to obtain the product ZerZ-butyl 7V-[2-[5-[( 127)-1 -benzyloxy-1(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[l-(cyclopropylmethyl)but-3-enoxy]-5(trifluoromethyl)-3-pyridyl]-jV-/erZ-butoxycarbonyl-carbamate (465 mg, 76%) as a colorless oil. ESI-MS m/z cale. 796.3271, found 797.8 (M+l)⁺; Rétention time: 9.44 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 -100% mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

333

Step 5: teri-Butyl 7V-[(6.R)-6-benzyIoxy-12-(cyclopropylmethyI)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaen-17-yl]TV-terZ-butoxycarbonyl-carbamate (E!Z mixture)

A flask was charged with ierAbutyl 7V-[2-[5-[(lA)-l-benzyloxy-l-(trifluoromethyl)pent4-enyl]-l,3,4-oxadiazol-2-yl]-6-[l-(cyclopropylmethyl)but-3-enoxy]-5-(trifluoromethyl)-3pyridyl]-7V-teri-butoxycarbonyl-carbamate (465 mg, 0.5836 mmol) followed by 1,2dichloroethane (280 mL) under argon. The reaction solution was treated with Zhan catalyst-lB (52 mg, 0.0708 mmol) at rt. The reaction was heated at 70 °C for 16 hours. LCMS indicated greater than 95% conversion of the starting material to products. LCMS indicated no additional conversion of tentative starting material peak. The reaction was treated with a drop of DMSO and the contents of the reaction vessel were concentrated onto S1O2. The material was purified by silica gel chromatography (40 g S1O2, dry loaded, eluted with 0 - 15% EtOAc in hexanes over a 20 min. gradient). The material was recovered impure and the fractions containing the target were combined and the solvent was evaporated in vacuo to obtain the product teri-butyl 7V-[(67?)6-benzyloxy-12-(cyclopropylmethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 (17),2,4,9,14(18), 15-hexaen-l 7-yl]-A-/er/-butoxycarbonylcarbamate (E/Z mixture) (272 mg, 52%) as a mixture of diastereomers; the material appeared as a transparent oil. Both peaks at 9.10 and 9.25 min. respectively: ESI-MS m/z cale. 768.2958, found 769.8 (M+l)⁺; Rétention time: 9.1 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100% mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

334

Step 6: terf-Butyl A^r-ZerZ-butoxycarbonyl-A-[(6JÎ)-12-(cyclopropylmcthyl)-6-hydroxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-17-yl]carbamate

Ε/Ζ mixture

A flask was charged with ZerZ-butyl A-[(6Æ)-6-benzyloxy-12-(cyclopropylmethyl)-6,15bis(trifluoromethyl)-l 3,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,9,14(18),15-hexaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate (E/Zmixture) (88 mg, 0.1145 mmol) in EtOH (4.8 mL). Then N2 was bubbled through the solution for 5 minutes. Then the solution was treated with Pd/C (24 mg, 10 % w/w, 0.0226 mmol) at room température and N2 was bubbled through the solution for 5 minutes. Then H2 gas was bubbled through the mixture for 20 min. and held under an atmosphère of H2 with a balloon. The reaction was complété in 11 h. The mixture was diluted with EtOAc and filtered through Celite. The filtrate was concentrated in vacuo to obtain the product ZerZ-butyl A-ZerZ-butoxycarbonyl-A-[(67î)-12(cyclopropylmethyl)-6-hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (74 mg, 94%) as a mixture of diastereomers; the material appeared as a light yellow oil which became a resinlike foam when thoroughly dried. ESI-MS m/z cale. 680.26447, found 581.3 (M-99)⁺; Rétention time: 8.01 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100% mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 7: (6J?)-17-Ammo-12-(cyclopropylmethyl)-6,15-bis(trifluoromethyI)-13,19-dioxa3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI (trifluoroacetate sait)

335

A vial was charged with ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(67?)-12(cyclopropylmethyl)-6-hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-17-yl]carbamate (155 mg, 0.2277 mmol) and treated with a solution of TFA (518.00 mg, 0.35 mL, 4.5429 mmol) in DCM (1.4 mL) at 0 °C. The reaction was warmed to room température over 3 hours. LCMS indicated the transformation was complété. The reaction was concentrated in vacuo to obtain a crude residue of the target product (128 mg). The material was further purified by reverse phase HP LC (Higgins Analytical Ci8 250 X 20mm (50-95%B over a 70 min. gradient), 25mL/min, 254 nm, Buffer A: 0.1% CF3CO2H in H2O, Buffer B: 0.1% CF3CO2H in acetonitrile). The pure fractions were combined and the solvent was evaporated in vacuo to obtain the product (67?)-17-amino-12(cyclopropylmethyl)-6,l 5-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (trifluoroacetate sait) (71 mg, 51%) as a dark yellow resin. ESI-MS m/z cale. 480.1596, found 481.3 (M+l)⁺; Rétention time: 3.27 minutes; LCMS Method: Water Cortex 2.7u Cis (3.0mm X 50mm), temp: 55 °C; Flow: 1.2mL/min; mobile phase: 100% water with 0.1% CF3CO2H then 100% acetonitrile with 0.1% CF3CO2H, grad: 5% to 100% B over 4 min, held at 100% B for 0.5 min, then equilibrated to 5% B over 1.5 min.

Step 8: (67?)-17-Amino-12-(cyclopropyImethyl)-6,15-bis(trifluoromethyI)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI (enantiomer 1), Compound 47, and (67?)-17-amino-12-(cycIopropyImethyl)-6,15-bis(trifIuoroniethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 48

enantiomer 1

enantiomer 2

The two diastereomers of (6Æ)-17-amino-12-(cyclopropylmethyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-l ( 18),2,4,14,16pentaen-6-ol (trifluoroacetate sait) (83 mg, 0.1728 mmol) were separated by supercritical fluid chromatography (SFC) using 0.1% diethylamine in EtOH as co-solvents. Fractions containing each diastereomer were then concentrated under reduced pressure and freeze-dried (acetonitrile/water mixture) to afford (67?)-17-amino-12-(cyclopropylmcthyl)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-6-ol (enantiomer 1) (45 mg) and (67?)-17-amino-12-(cyclopropylmethyl)-6,15

336 bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (enantiomer 2) (31 mg). Co-evaporation of the enantiomer 1 (2 mg) with acetonitrile (2X3 mL) could not remove isopropanol. Enantiomer 1 (2 mg) was dissolved in EtOAc and washing with 1 N HCl could remove isopropanol but a byproduct was observed by ’H NMR. Enantiomer 1 (about 39 mg) was purifïed by silica gel chromatography (24 g S1O2, eluting 10 to 30% EtOAc/CIhCh) and freeze dried to afford (6/?)-17-amino-12(cyclopropylmethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1) (34 mg, 40%) as a pale-yellow solid. *H NMR (400 MHz, DMSO-dô) δ 7.77 (s, 1H), 7.58 (s, 1H), 6.36 (s, 2H), 4.69 - 4.58 (m, 1H), 2.48 - 2.43 (m, 1H), 2.26 - 2.15 (m, 1H), 2.12 - 2.00 (m, 1H), 1.79 (ddd,J= 14.5, 9.0, 5.3 Hz, 1H), 1.72 - 1.54 (m, 3H), 1.54 - 1.40 (m, 3H), 1.35 (ddd, J= 14.5, 8.2, 2.4 Hz, 1H), 1.30 -1.21 (m, 1H), 0.94 - 0.83 (m, 1H), 0.50 - 0.41 (m, 1H), 0.35 (tt, J= 8.7, 4.4 Hz, 1H), 0.16 (dq, J= 9.0, 4.6 Hz, 1H), 0.08 to -0.01 (m, 1H) ppm. ¹⁹F NMR (377 MHz, DMSO-dô) δ -62.38 (s, 3F), -79.03 (s, 3F) ppm. ESI-MS m/z cale. 480.1596, found 481.2 (M+l)⁺ ; Rétention time: 3.83 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Product enantiomer 2 (about 29 mg) was purifïed by flash chromatography (24 g S1O2, eluting 10 to 30% EtOAc/C^Ch) and freeze dried to afford (67?)-17-amino-12(cyclopropylmethyl)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2) (27 mg, 32%) as a pale-yellow solid. ESI-MS m/z cale. 480.1596, found 481.1 (M+l)⁺; Rétention time: 3.81 minutes. Ή NMR (400 MHz, DMSO-d₆) δ 7.77 (s, 1H), 7.57 (s, 1H), 6.36 (s, 2H), 4.70 (t, J= 9.3 Hz, 1H), 2.44 - 2.35 (m, 1H), 2.35 - 2.24 (m, 1H), 2.15 - 2.03 (m, 1H), 1.84 - 1.64 (m, 3H), 1.58 - 1.33 (m, 5H), 1.32 - 1.21 (m, 1H), 0.94 - 0.82 (m, 1H), 0.51 - 0.42 (m, 1H), 0.35 (tt, J = 8.5, 4.5 Hz, 1H), 0.16 (dq, J= 9.0, 4.6 Hz, 1H), 0.08 to -0.01 (m, 1H) ppm. ¹⁹F NMR (377 MHz, DMSO-dô) δ -62.39 (s, 3F), -76.37 (s, 3F) ppm. ESI-MS m/z cale. 480.1596, found 481.1 (M+l)⁺; Rétention time: 3.81 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Example 35: Préparation of (6Æ)-17-amino-12-[(4-fluorophenyI)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (enantiomer 1), Compound 49, and (6R)-17-amino-12-[(4fluorophenyl)methyl]-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18337 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 50

enantiomer 1 enantiomer 2

Step 1: l-(4-Fluorophenyi)pent-4-en-2-ol

A flame dried round bottom flask was charged with 2-(4-fluorophenyl)acetaldehyde (7.0224 g, 6.6 mL, 48.294 mmol) and diethyl ether (57 mL) and cooled to 0 °C. Then allyl(bromo)magnesium (60.4 mL of 1 M, 60.400 mmol) was added dropwise over 30 minutes. The cooling bath was removed and the reaction mixture was allowed to warm to room température and stir for one hour. A saturated aqueous solution of ammonium chloride was added to the flask until the white precipitate disappears. The layers were separated and the

338 aqueous layer was washed with ethyl acetate (3 X 200 mL). The organic layers were combined and washed with a brine solution (2 X 50 mL). The organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated in vacuo. The crude reaction mixture was dry-loaded on silica gel and purifîed by liquid chromatography on silica gel, eluting with portions of ethyl acetate (0 - 30%) in heptanes. Isolated l-(4-fluorophenyl)pent-4-en-2-ol (4.034 g, 44%) as light yellow oil. *H NMR (400 MHz, CDCh) δ 7.24 - 7.15 (m, 2H), 7.06 - 6.95 (m, 2H), 5.94 - 5.78 (m, 1H), 5.23 - 5.10 (m, 2H), 3.86 (qd, J= 7.9, 4.6 Hz, 1H), 2.85 - 2.66 (m, 2H), 2.40 - 2.28 (m, 1H), 2.27-2.15 (m, 1H), 1.66 (d, J=3.4Hz, 1H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ-116.73 to -116.86 (m, 1F) ppm. ESI-MS m/z cale. 180.095, found 163.1 (M-17)⁺; Rétention time: 2.65 minutes. LCMS Method: Kinetex Polar Ci8, 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min. A second fraction was isolated also, slightly less pure, 1(4-fluorophenyl)pent-4-en-2-ol (0.575 g, 6%) as light yellow oil. ESI-MS m/z cale. 180.095, found 163.1 (M-17)⁺; Rétention time: 2.65 minutes. LCMS Method: Kinetex Polar C18.3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Step 2: tert-Butyl JV-[2-[5-[(LR)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yI]-6-[l-[(4-fluorophenyl)methyl]but-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]TV-tert-butoxycarbonyl-carbamate

To a solution of ter/-butylN-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-fôr/-butoxycarbonylcarbamate (250 mg, 0.3630 mmol) and l-(4-fluorophenyl)pent-4-en-2-ol (102 mg, 0.5660 mmol) in toluene (3 mL) was added triphenylphosphine (204 mg, 0.7778 mmol). After stirring at room température for 1 min, DIAD (150 pL, 0.7618 mmol) was added and the mixture stirred at room température for 30 minutes. Diluted the reaction mixture with EtOAc then washed with saturated aqueous NaHCO₃ (IX), saturated aqueous NH₄C1 (IX) and brine (IX) then dried over MgSO4, filtered and concentrated to a yellow oil which was purifîed by silica gel chromatography using a shallow gradient from 100% hexanes to 50% EtOAc-hexanes giving as diastereomeric mixture, a clear, slightly yellow syrup, terZ-butyl 7V-[2-[5-[(lR)-l-benzyIoxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[l-[(4-fluorophenyl)methyl]but-3-enoxy]5-(trifluoromethyl)-3-pyridyl]-7V-teri-butoxycarbonyl-carbamate (160 mg, 52%). ’H NMR (400 339

MHz, Chloroform-d) δ 7.80 (s, 1H), 7.35 (ddd, J= 6.9, 5.2,1.9 Hz, 2H), 7.33 - 7.25 (m, 3H), 7.16 (ddd, J= 8.8, 5.4, 2.1 Hz, 2H), 6.90 - 6.84 (m, 2H), 5.88 - 5.69 (m, 2H), 5.61 - 5.48 (m, 1H), 5.11 - 4.97 (m, 4H), 4.79 (dd, J= 10.8, 1.9 Hz, 1H), 4.60 (dd, J= 10.7, 7.1 Hz, 1H), 3.05 (ddd, J= 13.8, 6.9, 5.3 Hz, 1H), 2.91 (dd, J= 13.9, 6.0 Hz, 1H), 2.64 - 2.08 (m, 5H), 1.40 (d, J= 6.8 Hz, 19H) ppm. ESI-MS m/z cale. 850.3176, found 851.5 (M+l)⁺; Rétention time: 2.16 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 3: tert-Butyl A-[(6/?)-6-benzyloxy-12-[(4-fluorophenyI)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,9,14(18),15-hexaen-17-yl]-7V-ter/-butoxycarbonyl-carbamate (E!Z mixture)

E/Z mixture

To a degassed solution of ieri-butyl N-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)pent4-enyl]-l,3,4-oxadiazol-2-yl]-6-[l-[(4-fluorophenyl)methyl]but-3-enoxy]-5-(trifluoromethyl)-3pyridyl]-7V-terZ-butoxycarbonyl-carbamate (150 mg, 0.1763 mmol) in DCE (75 mL) was added Zhan catalyst-lB (24 mg, 0.03271 mmol) at once and the reaction mixture was heated at 60 °C for about 1 h. Cooled the reaction mixture to room température and quenched the reaction with few drops of DMSO and solvents were removed. The résultant brown residue was purified by silica gel column chromatography using a shallow gradient 100% hexanes to 50% EtOAc giving a diastereomeric mixture of ZerZ-butyl 7V-[(6R)-6-benzyloxy-12-[(4-fluorophenyl)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,9,14(18),15-hexaen-17-yl]-N-tert-butoxycarbonyl-carbamate (E/Z mixture) (92 mg, 63%). ESI-MS m/z cale. 822.2864, found 823.5 (M+l)⁺; Rétention time: 2.03 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 50 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile

340 phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 4: tert-Butyl /V-teri-butoxycarbonyl-7V-[(67î)-12-[(4-fluorophenyl)methyl]-6-hydroxy6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-17-yl]carbamate

E/Z mixture

To a solution of terZ-butyl 7V-[(6R)-6-benzyloxy-12-[(4-fluorophenyl)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,9,14(18),15-hexaen-17-yl]-7V-teri-butoxycarbonyl-carbamate (E/Zmixture) (92 mg, 0.1118 mmol) in EtOH (3 mL) was added, Pd/C (38 mg of 10 % w/w, 0.03571 mmol) (50% water) in a 250 mL vessel equipped with a Fh-balloon using 3-way adaptor. Subjected to vacuum and backfilled with nitrogen gas three times then subjected to vacuum. Filled the vessel with hydrogen gas and the mixture was stirred at room température ovemight. Subjected to vacuum and filled with nitrogen gas three times then diluted with ethyl acetate and filtered over Celite. The filtrate was concentrated to give a colorless viscous oil as a diastereomeric mixture Zeri-butyl V-/er/-butoxycarbonyl-7V-[(6/?)-12-[(4-fiuorophenyl)methyl]-6-hydroxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15pentaen-17-yl]carbamate (72 mg, 88%). ESI-MS m/z cale. 734.25507, found 735.4 (M+l)⁺; Rétention time: 2.07 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Ci8 column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate =1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

341

Step 5: (67?)-17-Amino-12-[(4-fluorophenyl)methyl]-6,15-bis(trifluoromethyl)-13,19-dioxa 3,4,18-triazatricyclo [ 12.3.1.12,5] nonadeca-1 (18),2,4,14,16-pentaen-6-ol

terAbutyl N-teri-butoxycarbonyl-7V-[(67î)-12-[(4-fluorophenyl)methyl]-6-hydroxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 17),2,4,14(18), 15pentaen-17-yl]carbamate (72 mg, 0.09800 mmol) was dissolved in a pre-made solution of TFA (250 pL, 3.245 mmol) and DCM (750 pL). Stirred the reaction for about 0.5 h and solvents removed. The résultant residue was dissolved in 2 mL of MeOH and was purified by a reverse phase HPLC-MS method using a dual gradient run from 30 - 99% mobile phase B over 15.0 minutes. Mobile phase A = FfrO (5 mM formic acid) afforded (6Æ)-17-amino-12-[(4fluorophenyl)methyl] -6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (42 mg, 80%); ESI-MS m/z cale. 534.1502, found 535.2 (M+l)⁺; Rétention time: 1.71 minutes as light yellow amorphous solid. ESI-MS m/z cale. 534.1502, found 535.2 (M+l)⁺; Rétention time: 1.71 minutes. Final purity was determined by reversed phase UPLC using an Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 30 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

Step 6: (6Æ)-17-Amino-12-|(4-fluorophenyl)methyl]-6,15-bis(trifIuoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 49, and (6R)-17-amino-12-[(4-fluorophenyl)methyl]-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI (enantiomer 2), Compound 50

enantiomer 1

enantiomer 2

342

The diastereomeric mixture of (6R)-17-amino-12-[(4-fluorophenyl)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-6-ol (60 mg, 0.1123 mmol) was purified by préparative SFC eluting with 15% methanol to CO2 though a 2 X 25 cm OJ-H column, providing the Ist eluent (67?)-17-amino-12-[(4fluorophenyl)methyl]-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1) (3.6 mg, 6%). Ή NMR (400 MHz, Methanol-Λ) δ 7.62 (s, 1H), 7.36 - 7.29 (m, 2H), 7.03 - 6.93 (m, 2H), 4.86 4.78 (m, 1H), 3.13 (dd, J= 14.3, 2.9 Hz, 1H), 2.95 (dd, J- 14.5, 8.6 Hz, 1H), 2.66 - 2.52 (m, 1H), 2.40 (dd, J= 14.4, 9.0 Hz, 1H), 2.15 (dt, J= 14.5, 8.0 Hz, 1H), 1.77 - 1.57 (m, 6H), 1.41 - 1.29 (m, 1H) ppm. ¹⁹F NMR (376 MHz, Methanol-d4) δ -64.79, -81.78, -119.61 ppm. ESI-MS m/z cale. 534.1502, found 535.4 (M+l)⁺; Rétention time: 2.21 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 x 4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Continued elution provided the 2nd eluent, (6R)-17-amino-12-[(4-fluorophenyl)methyl]6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-6-ol (enantiomer 2) (2.8 mg, 5%). 'il NMR (400 MHz, Methanol-d4) δ 7.63 (s, 1H), 7.32 (dd, J= 8.6, 5.6 Hz, 2H), 7.06 - 6.91 (m, 2H), 5.02 - 4.91 (m, 1H), 3.14 (dd, J= 14.4, 2.9 Hz, 1H), 2.95 (dd, J= 14.5, 8.7 Hz, 1H), 2.54 (ddd, J= 13.9, 8.5, 6.2 Hz, 1H), 2.40 (t, J= 12.0 Hz, 1H), 2.21 (dt, J= 14.5, 7.7 Hz, 1H), 1.94 - 1.72 (m, 3H), 1.68 - 1.52 (m, 3H), 1.41 (ddd, J= 13.0, 9.7, 6.5 Hz, 1H) ppm. ¹⁹F NMR (376 MHz, Methanol-d4) δ -64.76, -78.82, -119.60 ppm. ESI-MS m/z cale. 534.1502, found 535.2 (M+l)⁺; Rétention time: 2.23 minutes. Final purity was determined by reversed phase HPLC-MS using an Onyx Monolithic Cis column (50 ^χ4.6 mm) sold by Phenomenex (pn: CHO-7644), and a dual gradient run from 1 - 99% mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H). Flow rate = 12 mL/min, injection volume = 50 pL, and column température = 25 °C.

Example 36: Préparation of (6R)-17-amino-12-[(3-terf-butyIphenyl)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (enantiomer 1), Compound 51, and (6R)-17-amino-12-[(3-ferfbutylphenyl)methyl]-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18343 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 52

enantiomer 1

Step 1: l-(3-terMJutyiphenyl)pent-4-en-2-ol

enantiomer 2

To a dry vial charged with 2-(3-Zer/-butylphenyl)acetaldehyde (140 mg, 0.7546 mmol) was added anhydrous diethyl ether (4 mL). T he solution was cooled to 0 °C before adding allyl(bromo)magnesium (1 mL of 1 Μ, 1.0000 mmol) dropwise. After the addition, the solution was allowed to warm up to room température and stirred for 1 h. A saturated aqueous 10 solution of NH4CI was added until the white précipitâtes disappeared. The aqueous layer was then extracted with EtOAc (3X10 mL). The organic layers were then combined, washed with brine, dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. ’ 344 21166

The crude was purified via silica gel column chromatography (eluting 0 - 20% EtOAc in hexanes) to yield l-(3-ZerZ-butylphenyl)pent-4-en-2-ol (100 mg, 58%) as a clear oil. ’H NMR (500 MHz, DMSO-dô) δ 7.25 - 7.15 (m, 3H), 7.02 - 6.98 (m, 1H), 5.95 - 5.79 (m, 1H), 5.03 (d, J = 1.4 Hz, 1H), 5.02 - 4.99 (m, 1H), 4.58 (d, J= 5.4 Hz, 1H), 3.74 - 3.68 (m, 1H), 2.63 (dd, J = 13.1, 6.3 Hz, 2H), 2.19 - 2.04 (m, 2H), 1.27 (d, J= 1.2 Hz, 9H) ppm.

Step 2: ZerZ-Butyl A-|2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-6-[l-[(3-Zer/-butylphenyl)methyl]but-3-enoxy]-5-(trifluoromethyl)-3pyridyl]-A-fôr/-butoxycarbonyl-carbamate

A flask was charged with ZerZ-butyl 7/-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent4-enyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonylcarbamate (100 mg, 0.1452 mmol), l-(3-ZerZ-butylphenyl)pent-4-en-2-ol (30 mg, 0.1374 mmol), triphenylphosphine (76 mg, 0.2898 mmol) and flushed with N2 gas. Then the mixture was treated with toluene (0.3 mL). The solution was treated with DIAD (62.640 mg, 0.06 mL, 0.3098 mmol) in toluene (0.2 mL) over 2 h at rt. The reaction stirred an additional 1 h at rt; LCMS indicated almost no alcohol présent. The reaction stirred an additional 72 h; LCMS indicated no alcohol présent. The reaction was diluted with EtOAc (15 mL) and washed with saturated NaHCO₃ (4 mL), then washed with saturated NH4CI (4 mL), then washed with brine (4 mL). The organics were dried over Na2SÛ4, filtered, concentrated in vacuo to obtain a crude residue. The crude residue was purified by silica gel chromatography (12 g S1O2, loaded with minimal DCM, eluted with 0 - 10% EtOAc in hexanes over a 10 column volume gradient). The pure fractions were combined and the solvent was evaporated in vacuo to obtain the product tertbutyl A/-[2-[5-[(lJ?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-6-[l-[(3ZerZ-butylphenyl)methyl]but-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonylcarbamate (59 mg, 43%) as a transparent colorless residue. ESI-MS m/z cale. 888.3897, found 789.7 (M-99)+; Rétention time: 9.77 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100% mobile phase B

345 over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 3: tert-Butyl 7V-[(6R)-6-benzyloxy-12-[(3-terZ-butylphenyl)methyI]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,9,14,16-hexaen-17-yl|-A⁷-Zi’ri‘-butoxycarbonyI-carbamate (E/Zmixture)

E/Z mixture

A flask was charged with ZerZ-butyl 7V-[2-[5-[(lR)-l-benzyloxy-l-(trifluoromethyl)pent4-enyl]-l,3,4-oxadiazol-2-yl]-6-[l-[(3-/erZ-butylphenyl)methyl]but-3-enoxy]-5(trifluoromethyl)-3-pyridyl]-A^r-ter/-butoxycarbonyl-carbamate (25 mg, 0.0281 mmol) followed by 1,2-dichloroethane (15 mL) under argon. The reaction solution was treated with Zhan catalyst-lB (2.5 mg, 0.0034 mmol) at room température. The reaction was heated at 70 °C for 15 h. The vessel was removed from heat. LCMS indicated SM remained 15%. The reaction was heated at 70 °C for a total of 24 h. LCMS indicated no additional conversion of the starting material to products. The contents of the reaction vessel were concentrated onto S1O2. The material was purified by silica gel chromatography (4 g S1O2, dry loaded, eluted with 0 - 15% EtOAc in hexanes over a 20 min. gradient). The material was recovered impure and the fractions were combined and the solvent was evaporated in vacuo to obtain the product, tertbutyl A^r-[(6/?)-6-bcnzyloxy-l 2-[(3-ZerZ-butylphcnyl)methyl]-6,15-bis(trifluoromethyl)-l 3,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaen-1 l-y\\-N-tertbutoxycarbonyl-carbamate (E7Z mixture) (10.7 mg, 38%) as a mixture of diastereomers as a transparent oil. The material was used without further purification in the next step. ESI-MS m/z cale. 860.3584, found 789.7 (M-O'Bu)⁺; Rétention time: 9.58 minutes; LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 100% mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

346

Step 4: ZerZ-Butyl A⁷-ZerZ-butoxycarbonyl-A-[(6/?)-12-[(3-ZerZ-butyIphenyl)methyl]-6hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate

E/Z mixture

A vial was charged with ZerZ-butyl 7V-[(6/?)-6-benzyloxy-12-[(3-ZerZ-butylphenyl)methyl]6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,9,14,16-hexaen-17-yl]-jV-zerZ-butoxycarbonyl-carbamate (E/Zmixture) (11 mg, 0.0109 mmol) in EtOH (0.61 mL). Then N₂ was bubbled through the solution for 5 minutes. Then the solution was treated with Pd/C (2.4 mg, 10 % w/w, 0.0023 mmol) at room température and N₂was bubbled through the solution for 5 minutes. Then H₂ gas was bubbled through the mixture for 20 min. and held under an atmosphère of H₂ with a balloon. The reaction was complété in 13 h. The mixture was diluted with EtOAc and filtered through Celite. The filtrate was concentrated in vacuo to obtain the product ZerZ-butyl A-ZerZ-butoxycarbonyl-N-[(6Æ)-12-[(3-ZerZbutylphenyl)methyl]-6-hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (10 mg, 100%) as a light yellow oil which became a resin-like foam when thoroughly dried. ESI-MS m/z cale. 772.3271, found 673.5 (M-99)+; Rétention time: 8.89 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100% mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO₂H). Mobile phase B = acetonitrile (0.1 % CF3CO₂H).

347

Step 5: (6Æ)-17-Amino-12-[(3-ZÉ’rZ-butylphenyl)methyl]-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol

A thick walled glass microwave vial was charged with ZerZ-butyl 7V-ZerZ-butoxycarbonylN-[(6R)-12-[ (3 -ZerZ-butylphenyl)methyl]-6-hydroxy-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (10 mg, 0.0129 mmol) in a solution of hexafluoro-2-propanol (0.25 mL). The reaction was heated at 100 °C in a microwave vessel for 3 hours. LCMS indicated the transformation was complété. The reaction was concentrated in vacuo to obtain a crude residue of the target (67?)-17-amino-12-[(3ZerZ-butylphenyl)methyl]-6,l 5-bis(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (7.4 mg, 98%) as a dark yellow resin. ESI-MS m/z cale. 572.2222, found 573.1 (M+l)⁺; Rétention time: 3.68 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100% mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 6: (6/?)-17-Amino-12-[(3-ZerZ-butyIphenyl)methyl]-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 51, and (6R)-17-amino-12-[(3-tert-butylphenyI)methyI]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (enantiomer 2), Compound 52

enantiomer 1 enantiomer 2 (672)-17-Amino-l 2-[(3-ZerZ-butylphenyl)methyl]-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (79.3 mg, 0.1338

348 mmol) was purified by préparative SFC eluting with 15% methanol to CO2 though a Cellulose-4 column (40 °C ; 100 Bar; Flow rate: 4 mL/min; 10% MeOH ; 8.5 min run), providing the two diastereoisomers. First elution afforded (6/?)-17-amino-12-[(3-fer/-butylphenyl)mcthyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-6-ol (enantiomer 1) (30.0 mg, 39%) as an off-white solid with a purity of 99.3%. *H NMR (400 MHz, DMSO-d₆) δ 7.72 (s, 1H), 7.59 (s, 1H), 7.28 (s, 1H), 7.24 - 7.12 (m, 2H), 7.11 7.02 (m, 1H), 6.41 - 6.32 (m, 2H), 4.93 - 4.70 (m, 1H), 3.07 (d, J= 14.1 Hz, 1H), 2.96 - 2.81 (m, 1H), 2.55 - 2.39 (m, 1H), 2.29 - 2.15 (m, 1H), 2.14 - 1.98 (m, 1H), 1.73 - 1.43 (m, 6H), 1.36 1.14 (m, 10H) ppm. ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.12 (s, 3F), -79.06 (s, 3F) ppm. ESIMS m/z cale. 572.2222, found 573.0 (M+l)⁺; Rétention time: 3.95 minutes. LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

Continued elution afforded (6R)-17-amino-12-[(3-fôrt-butylphenyl)methyl]-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-6-ol (enantiomer 2) (28.6 mg, 37%) as an off-white solid with a purity of 99.9%. *H NMR (400 MHz, DMSO-d₆) δ 7.72 (s, 1H), 7.59 (br. s, 1H), 7.28 (s, 1H), 7.24 - 7.15 (m, 2H), 7.13 - 7.00 (m, 1H), 6.42 - 6.28 (m, 2H), 4.87 (t, J = 9.0 Hz, 1H), 3.09 (d, J= 12.8 Hz, 1H), 2.89 (dd, J= 14.4, 8.4 Hz, 1H), 2.42 - 2.26 (m, 2H), 2.15 -1.99 (m, 1H), 1.83 - 1.59 (m, 3H), 1.57 1.39 (m, 3H), 1.37 - 1.28 (m, 1H), 1.25 (s, 9H) ppm. ¹⁹F NMR (377 MHz, DMSO-d6) δ -62.12 (s, 3F), -76.37 (s, 3F) ppm. ESI-MS m/z cale. 572.2222, found 573.0 (M+l)⁺; Rétention time: 3.94 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm 2.6 pm, 6 min, 5 - 95% acetonitrile in H2O (0.1% formic acid) 1.2 mL/min.

349

Example 37: Préparation of (6J?,12/?)-17-amino-12-methyl-6,15-bis(trifluoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,8-diol (diastereomer 1), Compound 53

E/Z mixture

Step 3

Step 4

Step 5

diastereomer 1

Step 1: ter/-Butyl A-[(6Æ,127?)-6-benzyloxy-8-hydroxy-12-methyl-6,15-bis(trifluoromethyI)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17yl] car b a ma te (diastereomer 1)

E/Z mixture

To a solution of ZerZ-butyl 7V-[(67?,127?)-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16hexaen-17-yl]-7V-/er?-butoxycarbonyl-carbamate (E/Z mixture) (230 mg, 0.2964 mmol) in THF (3.7 mL) at 0 °C under nitrogen was added dropwise borane dimethyl sulfide complex (80.100 mg, 0.1 mL, 1.0544 mmol). The mixture was stirred at 0 °C for 10 min and at room température

350 for 1 h and re-cooled to 0 °C. NaOH in water (1.8 mL of 1 M, 1.8000 mmol) was added, followed by H2O2 in water (688.93 mg, 0.6207 mL of 30% w/w, 6.0762 mmol). The mixture was stirred at room température for 50 min. A 10% aqueous solution of sodium thiosulfate (9 mL) and brine (7 mL) were added. The mixture was stirred at 0 °C for 10 min and extracted with ethyl acetate (3 X 20 mL). The combined organic layers were dried with anhydrous sodium sulfate, filtered and concentrated. The residue was purified by silica gel chromatography (24 g S1O2, eluting 0% to 10% ethyl acetate/dichloromethane) to provide a mixture containing the mono-boc diastereomers, ZerZ-butyl A^L[(67?,127?)-6-benzyloxy-8-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-l ( 18),2,4,14,16pentaen-17-yl]carbamate (59 mg, 16%) as clear oil with some impurities still présent, ESI-MS m/z cale. 646.2226, found 647.2 (M+l)⁺; Rétention time: 4.16 minutes (Kinetex polar Cis (3.0 X 50 mm) 2.6 pm, 6 min, 5% - 95% acetonitrile in H2O (0.1% formic acid), flow =1.2 mL/min). This material was dry-loaded on silica gel and purified by liquid chromatography on a 12 g silica gel column eluting with 0% to 30% portions of ethyl acetate in heptanes. At 16% ethyl acetate in heptanes, the target product eluted, ZerZ-butyl A-[(67?,12A)-6-benzyloxy-8-hydroxy-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (diastereomer 1) (17 mg, 26%) and was isolated as a sticky white foam which still contained some minor impurities. *H NMR (400 MHz, CDCI3) δ 9.28 (s, 1H), 9.13 (s, 1H), 7.40 - 7.28 (m, 5H), 5.10 - 4.99 (m, 1H), 4.73 (d, J= 11.0 Hz, 1H), 4.65 (d, J= 10.3 Hz, 1H), 4.24 - 4.15 (m, 1H), 2.72 - 2.61 (m, 1H), 2.57 - 2.48 (m, 1H), 2.47 2.36 (m, 1H), 2.25 - 2.13 (m, 1H), 1.85 - 1.61 (m, 5H), 1.56 (s, 9H), 1.46 (d, J= 6.1 Hz, 3H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.93 (s, 3F), -74.31 (s, 3F) ppm. ESI-MS m/z cale. 646.2226, found 647.2 (M+l)+; Rétention time: 4.15 minutes; LCMS Method: Kinetex Polar Cis (3.0 X 50 mm) 2.6 pm, 6 min, 5 - 95% MeCN in H2O (0.1% formic acid), flow =1.2 mL/min.

Step 2: [(6/?,12/7)-6-Bcnzyloxy-17-(ZerZ-butoxycarbonylamino)-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-8-yl] acetate (diastereomer 1)

diastereomer 1

351

To a solution of terf-butyl 7V-[(6/?,12/?)-6-benzyloxy-8-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 1) (15 mg, 0.0232 mmol) in CH2CI2 (1 mL) at 0 °C was added DMAP (8 mg, 0.0655 mmol), followed by acetic anhydride (6 mg, 0.0055 mL, 0.0588 mmol). The mixture was stirred at rt for 2.5 h and concentrated. The residue was purified by silica gel chromatography (12 g S1O2, eluting 0 to 25% EtOAc/heptanes) to afford [(6/?,127?)-6benzyloxy-17-(/er/-butoxycarbonylamino)-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-8-yl] acetate (diastereomer 1) (14 mg, 88%) as a pale-yellow oil. ^!H NMR (400 MHz, CDCI3) δ 9.23 (s, 1H), 9.13 (s, 1H), 7.36 - 7.27 (m, 5H), 5.44 - 5.33 (m, 1H), 4.95 - 4.84 (m, 1H), 4.78 (d, J= 11.0 Hz, 1H), 4.60 (d, 11.0 Hz, 1H), 2.68 - 2.56 (m, 2H), 2.55 - 2.45 (m, 1H), 2.20 - 2.10 (m, 1H), 2.01 (s, 3H), 1.74 - 1.64 (m, 1H), 1.56 (s, 9H), 1.53 - 1.46 (m, 1H), 1.44 (d, J = 6.4 Hz, 3H), 1.41 - 1.30 (m, 2H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -63.94 (s, 3F), -74.58 (s, 3F) ppm.

Step 3: [(6/?,127?)-17-Aniino-6-benzyloxy-12-niethyI-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-yl] acetate (diastereomer 1)

diastereomer 1

To a solution of [(6Æ,12Æ)-6-benzyloxy-17-(tert-butoxycarbonylamino)-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-8-yl] acetate (diastereomer 1) (14 mg, 0.0203 mmol) in CH2CI2 (2 mL) at 0 °C was added TFA (740.00 mg, 0.5 mL, 6.4899 mmol). The mixture was stirred at 10 - 13 °C for 1.5 h. The solvents were removed by a gentle flow of nitrogen. The residue was treated with 2 drops of 28% aqueous ammonia and purified by silica gel chromatography (4 g S1O2, eluting 10% to 40% EtOAc/heptanes) to afford [(6Z?,12/?)-17-amino-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16pentaen-8-yl] acetate (diastereomer 1) (9 mg, 75%) as a pale-yellow oil. 'H NMR (400 MHz, CDCI3) δ 7.45 (s, 1H), 7.37 - 7.28 (m, 5H), 5.45 - 5.37 (m, 1H), 5.34 (s, 2H), 4.90 - 4.80 (m, 1H), 4.77 (d, J= 10.8 Hz, 1H), 4.61 (d, J= 11.0 Hz, 1H), 2.70 - 2.46 (m, 3H), 2.23 - 2.12 (m, 1H), 2.01 (s, 3H), 1.73 - 1.62 (m, 1H), 1.55 - 1.43 (m, 2H), 1.41 (d, J= 6.4 Hz, 3H), 1.37 -1.29 (m, 1H) ppm. ¹⁹F NMR (377 MHz, CDCI3) δ -64.02 (s, 3F), -74.67 (s, 3F) ppm.

352

Step 4: (6Æ,127?)-17-Amino-6-benzyloxy-12-methyl-6,15-bis(trifIuoromethyI)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-ol (diastereomer 1)

To a solution of [(67?,12E)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16-pentaen-8-yl] acetate (diastereomer 1) (9 mg, 0.0153 mmol) in THF (2 mL) was added aqueous NaOH (0.4 mL of 1 M, 0.4000 mmol). The mixture was stirred at rt for 1 h. MeOH (395.50 mg, 0.5 mL, 12.343 mmol) was added. The mixture was stirred at room température ovemight. The mixture was concentrated and the residue was treated with saturated aqueous NaHCO₃ (3 mL). The mixture was extracted with EtOAc (3X8 mL). The combined organic layers were dried with Na₂SÛ4, filtered and concentrated. The residue was purified by silica gel chromatography (4 g SiO₂, eluting 20% to 50% EtOAc/heptanes) to afford (67?,12E)-17-amino-6-benzyloxy-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-8-ol (diastereomer 1) (8 mg, 96%) as a pale-yellow oil. ’H NMR (400 MHz, CDCh) δ 7.45 (s, 1H), 7.38 - 7.28 (m, 5H), 5.36 (s, 2H), 5.04 - 4.92 (m, 1H), 4.76 - 4.69 (m, 1H), 4.68 - 4.61 (m, 1H), 4.27 - 4.17 (m, 1H), 2.71 - 2.62 (m, 1H), 2.56 - 2.38 (m, 2H), 2.29 - 2.18 (m, 1H), 1.85 - 1.69 (m, 2H), 1.67 - 1.61 (m, 1H), 1.54 - 1.47 (m, 1H), 1.43 (d, J= 6.4 Hz, 3H), 1.39 - 1.33 (m, 1H) ppm. ’⁹F NMR (377 MHz, CDCh) δ -63.99 (s, 3F), -74.38 (s, 3F) ppm. ESI-MS m/z cale. 546.17017, found 546.9 (M+l)⁺; Rétention time: 2.22 minutes; LCMS Method: Kinetex Polar Cis (3.0 X 50 mm), 2.6 pm, 3 min, 5 - 95% MeCN in H₂O (0.1% formic acid), flow = 1.2 mL/min.

Step 5: (6Æ,12Æ)-17-Amino-12-methyl-6,15-bis(triiluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,8-diol (diastereomer 1), Compound 53

diastereomer 1

353

A mixture of (67?,12Jî)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-8-ol (diastereomer 1) (8 mg, 0.0146 mmol) and palladium on carbon 5% wet (7 mg, 0.0033 mmol) in MeOH (2 mL) was stirred under hydrogen (balloon) at room température ovemight. The mixture was fïltered through diatomaceous earth and washed with EtOAc. The filtrate was concentrated and the residue was purified by silica gel chromatography (12 g S1O2, eluting 30% to 50% EtOAc/pentane) and freeze dried to afford (67?,127?)-17-amino-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaene-6,8-diol (diastereomer 1) (4.6 mg, 68%) as a pale-yellow solid. *H NMR (400 MHz,

DMSO-de) δ 7.76 (s, 1H), 7.52 (s, 1H), 6.37 (s, 2H), 5.01 - 4.89 (m, 1H), 4.55 (d, J= 6.8 Hz, 1H), 3.94 - 3.79 (m, 1H), 2.39 - 2.09 (m, 4H), 1.83 - 1.69 (m, 1H), 1.45 - 1.37 (m, 1H), 1.34 (d, J = 6.4 Hz, 3H), 1.27 - 1.13 (m, 2H) ppm. ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.51 (s, 3F), -76.79 (s, 3F) ppm. ESI-MS m/z cale. 456.12323, found 456.9 (M+l)⁺; Rétention time: 2.78 minutes; LCMS Method: Kinetex Polar Cis (3.0 X 50 mm), 2.6 pm, 6 min, 5 - 95% MeCN in H2O (0.1% formic acid), flow =1.2 mL/min.

Example 38: Préparation of (61î)-17-aniino-12-ethyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 54, and (6Æ)-17-amino-12-ethyl-6,15-bis(trifIuoromethyl)-13,19-dioxa-3,4,18354 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 55

Step 2

Step 3

Step 4

E/Z mixture

Step 5

enantiomer 1

enantiomer 2

Step 1: ZerZ-Butyl 7V-[2-[5-[(LR)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol5 2-yl]-6-(l-ethylpent-4-enoxy)-5-(trifluoromethyl)-3-pyridyI]-A-ZerZ-butoxycarbonylcarbamate

A solution of ZerZ-butyl 7V-[2-[5-[(lÆ)-l-benzyloxy-l-(trifluoiOmethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-A-ZerZ-butoxycarbonyl-carbamate (1 g, 10 1.4824 mmol), hept-6-en-3-ol (260 mg, 2.2770 mmol), and triphenylphosphine (800 mg, 0.7067

355 mL, 3.0501 mmol) in THF (40 mL) at 0 °C was stirred for 30 min and DIAD (626.40 mg, 0.6 mL, 3.0978 mmol) was added dropwise. The reaction mixture was warmed to rt and stirred for 2 h. The reaction mixture was concentrated and the residue was purifîed by silica gel chromatography (40 g column, 0 to 15 % EtOAc in hexanes for 30 min) provided as a clear oil, ZerZ-butyl ?V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoroniethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-(lethylpent-4-enoxy)-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (1.07 g, 94 %). ’H NMR (500 MHz, CDCh) δ 7.82 (s, 1H), 7.41 - 7.18 (m, 5H), 5.98 - 5.86 (m, 1H), 5.85 5.72 (m, 1H), 5.32 - 5.21 (m, 2H), 5.18 (d, J= 10.2 Hz, 1H), 4.99 - 4.93 (m, 1H), 4.91 (dd, J= 10.2, 2.4 Hz, 1H), 4.80 (dd, J= 10.7, 2.7 Hz, 1H), 4.62 (dd, J= 10.6, 5.7 Hz, 1H), 3.28 - 3.08 (m, 2H), 2.22-2.03 (m, 2H), 1.93 - 1.83 (m, 1 H), 1.81 - 1.70 (m, 3H), 1.42 (s, 18H), 0.960.88 (m, 3H). ESI-MS m/z cale. 770.3114, found 671.5 (M+H-Boc)⁺; Rétention time: 4.74 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 2: ZcrZ-Butyl A-[(67?)-6-benzyIoxy-12-ethyI-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yI]-7V-tertbutoxycarbonyl-carbamate (E/Z mixture)

A solution of ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-(l-ethylpent-4-enoxy)-5-(trifluoromethyl)-3-pyridyl]-7V-Zer/-butoxycarbonylcarbamate (0.88 g, 1.0846 mmol) in DCE (200 mL) was degassed for 15 min and heated at 50 °C under nitrogen atmosphère for 15 min. Zhan catalyst-lB (200 mg, 0.2722 mmol) was then added and the mixture was heated at 70 °C ovemight. The reaction mixture was cooled and concentrated under reduced pressure. The residue was purifîed by silica gel chromatography (40 g column, 0-10 % EtOAc in hexanes) to provide as a white solid, ZerZ-butyl N-[{6R)-6benzyloxy-12-ethyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yl]-7V-ZerZ-butoxycarbonylcarbamate (E/Zmixture) (0.85 g, 91 %). ESI-MS m/z cale. 742.2801, found 643.3 (M+H-Boc)⁺; Rétention time: 4.68 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis

356 column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 3: ZerZ-Butyl 7V-tert-butoxycarbonyl-7V-[(6.R)-12-ethyl-6-hydroxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl] carbamate

A solution of Ze/7-butyl A^L[(6Z?)-6-bcnzyloxy-12-ethyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yl]-AZ-ZerZbutoxycarbonyl-carbamate (E/Z mixture) (0.746 g, 1.0044 mmol) in MeOH (40 mL) was purged with nitrogen three times. It was then backfilled with hydrogen two times before subjecting it to hydrogénation at 60 psi for 22 h using a Parr shaker. After the reaction was done, the reaction mixture was filtered over a pad of Celite and the filter cake was rinsed with MeOH (3 X 20 mL). The combined filtrate was concentrated and the residue was dried in vacuo ovemight provided as a clear oil, ZerZ-butyl ?/-ZerZ-butoxycarbonyl-7V-[(67?)-12-ethyl-6-hydroxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]carbamate (660 mg, 98 %). ESI-MS m/z cale. 654.24884, found 555.3 (M+HBoc)⁺.

Step 4: (6Æ)-17-Amino-12-ethyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol

To a solution of ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(6R)-12-ethyl-6-hydroxy-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (660 mg, 0.9881 mmol) in DCM (50 mL) was added TFA (14.800 g,

357 mL, 129.80 mmol). The resulting yellow solution was stirred at room température for 2 h.

The reaction mixture was concentrated and purified by reverse phase chromatography (buffer A: water buffered with 5 mM HCl; buffer B: 100 % ACN, 55 % to 100 % over 40 min) provided after lyophilization as a yellow powder, (6A)-17-amino-12-ethyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (hydrochloride sait ) (255.7 mg, 51 %). 'H NMR (500 MHz, DMSO-dô) δ 7.76 (s, 1H), 7.57 (s, 1H), 6.36 (s, 2H), 4.52 (dt, J= 26.9, 9.2 Hz, 1H), 2.48-2.39 (m, 1H), 2.18 (t, J= 11.4 Hz, 1H), 2.12-2.01 (m, 1H), 1.83-1.72 (m, 1H), 1.70-1.54 (m, 4H), 1.55-1.37 (m, 3H), 1.29-1.15 (m, 1H), 0.96 (t, J= 7.3 Hz, 3H). ESI-MS m/z cale. 454.14395, found 97.27 (M+l)⁺; Rétention time: 3.16 minutes. LCMS Method: Waters Cortex 2.7 pm particle size Cis (3.0 mm X 50 mm), 55 °C; flow: 1.2 mL/min; mobile phase: 100 % water with 0.1 % trifluoroacetic acid then 100 % acetonitrile with 0.1 % trifluoroacetic acid, gradient of 5 % to 100 % B over 4 min, with équilibration at 100 % B for 0.5 min, then 5 % B over 1.5 min.

Step 5: (6R)-17-Amino-12-ethyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 54, and (67?)-17-amino-12-ethyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 55

enantiomer 1

enantiomer 2

The diastereomeric mixture, (6/?)-17-amino-12-cthyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (255.7 mg, 0.5622 mmol) was purified by SFC using Cellulose 4 column (250 X 30 mm, 5 pm particle size) using a dual gradient run from 10 % EtOH (0.1 % diethylamine) and 90 % CO2.Both isomers were redissolved in EtOAc (5 mL) then washed with 1 M HCl (1X10 mL), saturated sodium bicarbonate (1X10 mL) and brine (1X10 mL). The organic layer was dried over anhydrous sodium sulfate, concentrated under vacuum and lyophilized. SFC peak 1 provided as a light yellow solid, (67?)-17-amino-12-ethyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1) (134 mg, 52%). ’H NMR (400 MHz, DMSO-dô) δ 7.77 (s, 1H), 7.60 (s, 1H), 6.36 (s, 2H), 4.51 (t, J= 9.0 Hz, 1H), 2.54 - 2.41 (m, 1H), 2.24 - 2.14 (m, 1H), 2.12 - 2.00 (m, 1H), 1.85 - 1.73 (m, 1H), 1.71 1.38 (m, 7H), 1.31-1.16 (m, 1H), 0.97 (t, J= 7.2 Hz, 3H). ¹⁹F NMR (377 MHz, DMSO-dô) δ 358

62.45 (s, 3F), -79.05 (s, 3F). ESI-MS m/z cale. 454.144, found 455.2 (M+l)⁺; Rétention time: 3.54 minutes. SFC peak 2 provided as a light yellow solid (6R)-17-amino-12-ethyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,14,16pentaen-6-ol (enantiomer 2) (126 mg, 49%). 'H NMR (400 MHz, DMSO-dô) δ 7.78 (s, 1H), 7.59 (s, 1H), 6.36 (s, 2H), 4.56 (t, J= 9.4 Hz, 1H), 2.46 - 2.36 (m, 1H), 2.35 - 2.24 (m, 1H), 2.14 2.05 (m, 1H), 1.87 - 1.66 (m, 3H), 1.64 - 1.35 (m, 5H), 1.33 - 1.20 (m, 1H), 0.97 (t, J= 7.2 Hz, 3H). ^I9F NMR (377 MHz, DMSO-d₆) δ -62.46 (s, 3F), -76.36 (s, 3F). ESI-MS m/z cale. 454.144, found 455.2 (M+l)⁺; Rétention time: 3.5 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate = 1.2 mL/min).

Example 39: Préparation of (12R)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 1), Compound 56, and (12/?)-17-amino-12-methyl-6,15-bis(trifluoromethyI)359

13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 2), Compound 57

Step 12

enantiomer 1 enantiomer 2

Step 1: Ethyl 3-tert-butoxy-2-hydroxy-2-(trifluoromethyl)pent-4-enoate

A flame-dried flask was charged with 2-allyloxy-2-methylpropane (5.55 g, 43.259 mmol), THF (100 mL) and TMEDA (4.4175 g, 5.7 mL, 38.015 mmol). The flask was cooled in a

360 dry-ice acetone bath and treated dropwise with a cyclohexane solution of sec-butyllithium (27 mL of 1.4 M, 37.800 mmol). After 45 minutes, heptane solution of trimethylaluminum (19 mL of 2 M, 38.000 mmol) was added and the reaction mixture was stirred for another 45 minutes. Ethyl 3,3,3-trifluoro-2-oxo-propanoate (5.1320 g, 4 mL, 30.173 mmol) was added and the reaction was stirred for about 4.0 hours in the cold bath. The reaction was quenched slowly with 1 N aqueous HCl (150 mL) and stirred vigorously at room température for few minutes.

Transferred to a 1.0 L separatory tunnel with 1 N HCl (200 mL) and extracted with diethyl ether (1 X 300 mL, 2 X 150 mL). The combined organic layers were washed with saturated aqueous sodium bicarbonate (200 mL), brine (200 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (330-g column, eluting from 0 % to 20 % ethyl acetate in heptanes) to afford as a colorless oil and single diastereomer pair, ethyl 3-/er/-butoxy-2-hydroxy-2-(trifluoromethyl)pent-4-enoate (3.24 g, 38 %). ’H NMR (400 MHz, CDC1₃) δ 5.87 (ddd, J= 17.7, 10.1, 7.9 Hz, 1H), 5.32 - 5.21 (m, 2H), 4.55 (d, J= 8.1 Hz, 1H), 4.39 - 4.23 (m, 2H), 3.85 (s, 1H), 1.33 (t, J= 7.2 Hz, 3H), 1.23 (s, 9H). ¹⁹F NMR (377 MHz, CDCI3) δ -73.35 (s, 3F).

Step 2: Ethyl 2-benzyloxy-3-ft?rZ-butoxy-2-(trifluoromethyl)pent-4-enoate

A solution of ethyl 3-/crZ-butoxy-2-hydroxy-2-(trifluoromethyl)pcnt-4-cnoate (3.24 g, 11.398 mmol) in DMF (50 mL) was cooled in an ice bath and treated with a minerai oil suspension of sodium hydride (563 mg, 60 %w/w, 14.076 mmol). After 40 minutes, bromomethyl benzene (2.6459 g, 1.84 mL, 15.470 mmol) was added and the reaction was gradually warm to room température and stirred ovemight. Transferred to a 1.0 L separatory funnel with water (450 mL) and the aqueous layer was extracted with MTBE (4 X 150 mL). The combined organic layers washed with water (2 X 150 mL), brine (150 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (220 g column, eluting from 0 % to 20 % ethyl acetate in heptanes) to afford as a colorless oil, ethyl 2-benzyloxy-3-teri-butoxy-2-(trifluoromethyl)pent-4-enoate (3.86 g, 83 %). *H NMR (400 MHz, CDCI3) δ 7.46 - 7.41 (m, 2H), 7.39 - 7.32 (m, 2H), 7.32 - 7.28 (m, 1H), 5.98 (ddd, J= 17.9, 9.3, 8.6 Hz, 1H), 5.27 - 5.22 (m, 1H), 5.21 (s, 1H), 4.92 - 4.85 (m, 1H), 4.79 4.73 (m, 1H), 4.59 (d, J= 8.1 Hz, 1H), 4.28 (q, J= 7.1 Hz, 2H), 1.32 (t, J= 7.1 Hz, 3H), 1.20 (s,

361

9H). ¹⁹F NMR (377 MHz, CDCI3) δ -65.50 (s, 3F). ESI-MS m/z cale. 374.1705, found 397.2 (M+23)⁺; Rétention time: 2.45 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 3: 2-Benzyloxy-3-tert-butoxy-2-(trifluoromethyl)pent-4-enoic acid

A solution of sodium hydroxide (1.13g, 28.252 mmol) in water (10 mL) was added to a solution of ethyl 2-benzyloxy-3-/er/-butoxy-2-(trifluoromethyl)pent-4-enoate (3.4 g, 8.6908 mmol) in methanol (30 mL). The reaction mixture was stirred in an oil bath at 70 °C for 4 days. The reaction was cooled to room température and concentrated under reduced pressure to remove methanol. Added water (100 mL) and acidified to pH of 1-2 with 1 N aqueous HCl. Transferred to a 500 mL separatory tunnel and the aqueous layer was extracted with ethyl acetate (3 X 100 mL). The combined organic layers washed with water (100 mL), brine (100 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to afford as a yellow oil, 2benzyloxy-3-/er/-butoxy-2-(trifluoromethyl)pent-4-enoic acid (3.29 g, 97 %). ’H NMR (400 MHz, DMSO-d₆) δ 14.11 (br. s., 1H), 7.45 - 7.26 (m, 5H), 5.86 (ddd, J= 17.6, 10.0, 8.1 Hz, 1H), 5.34 - 5.20 (m, 2H), 4.88 (d, J= 11.2 Hz, 1H), 4.67 - 4.56 (m, 2H), 1.15 (s, 9H). ¹⁹FNMR (377 MHz, DMSO-dô) δ -64.77 (s, 3F). ESI-MS m/z cale. 346.1392, found no ionization; Rétention time: 2.15 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate = 1.2 mL/min).

Step 4: 2-BenzyIoxy-3-/er/-butoxy-2-(trifluoromethyl)pent-4-enehydrazide

A solution of 2-benzyloxy-3-/errtbutoxy-2-(trifluoromethyI)pcnt-4-enoic acid (3.29 g, 8.4354 mmol) and triethylamine (2.5410 g, 3.5 mL, 25.111 mmol) in DMF (50 mL) was treated with HATU (6.46 g, 16.990 mmol) and stirred at room température for 20 minutes. Cooled in an 362 ice bath and added hydrazine.IbO (6.7080 g, 10 mL, 87.099 mmol). After about 10 minutes, removed the ice bath and the reaction was stirred at room température for about 18 hours. Transferred to a 1.0 L separatory tunnel with water (450 mL) and the aqueous layer was extracted with ethyl acetate (4 X 150 mL). The combined organic layers washed with water (2 X 250 mL), brine (200 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (220 g column, eluting from 0 % to 50 % ethyl acetate in heptanes) to afford as a colorless oil, 2-benzyloxy-3-ZerZ-butoxy-2(trifluoromethyl)pent-4-enehydrazide (2.836 g, 91 %) that solidified on standing. *H NMR (400 MHz, DMSO-dô) δ 9.08 (br. s., 1H), 7.45 - 7.25 (m, 5H), 5.81 (ddd, J= 17.4, 10.5, 7.1 Hz, 1H), 5.27 (d, J= 17.1 Hz, 1H), 5.21 (d, J= 10.5 Hz, 1H), 5.11 - 4.99 (m, 2H), 4.90 (d, J= 7.3 Hz, 1H), 4.36 (d, J= 4.4 Hz, 2H), 1.18 (s, 9H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -67.28 (s, 3F). ESI-MS m/z cale. 360.1661, found 305.1 (M-55)⁺; Rétention time: 2.13 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 5: ZerZ-Butyl A-[2-[[[2-benzyloxy-3-Z<?rZ-butoxy-2-(trifIuoromethyI)pent-4enoyl] amino] carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl] carbamate

A mixture of 6-bromo-3-(ZerZ-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid (661 mg, 1.7163 mmol) and 2-benzyloxy-3-ZerZ-butoxy-2-(trifluoromethyl)pent4-enehydrazide (601 mg, 1.6260 mmol) in ethyl acetate (10 mL) was treated successively with pyridine (508.56 mg, 0.52 mL, 6.4293 mmol) and an ethyl acetate solution of T₃P (1.86 g, 50 %w/w, 2.9229 mmol) at room température., The reaction mixture was stirred for 23 hours, transferred to a 250 mL separatory fiinnel with saturated aqueous ammonium chloride (100 mL) and extracted with ethyl acetate (2 X 75 mL). The combined organic layers were washed with saturated aqueous sodium bicarbonate (100 mL), brine (100 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (80-g column, eluting from 0 % to 50 % ethyl acetate in heptanes) to afford as a thick pale amber oil, ZerZ-butyl 7V-[2-[[[2-benzyloxy-3-ZerZ-butoxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (1.1 g, 92 %). *H NMR (400 MHz, DMSO-d₆) δ 11.04 (s, 1H), 10.84 (s, 1H), 10.10 (s, 1H), 9.13 (s, 1H), 7.47 363

7.41 (m, 2H), 7.40 - 7.33 (m, 2H), 7.33 - 7.28 (m, 1H), 6.02 (ddd, J= 17.1, 10.6, 6.4 Hz, 1H), 5.46 (d, .7=17.1 Hz, 1H), 5.35 (d, J= 10.5 Hz, 1H), 5.19 (d, J= 11.7 Hz, 1H), 5.10 (d, J= 12.5 Hz, 1H), 4.95 (d, J= 6.4 Hz, 1H), 1.49 (s, 9H), 1.22 (s, 9H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ 62.86 (s, 3F), -67.31 (s, 3F). ESI-MS m/z cale. 726.1488, found 571.0 (M-155)⁺; Rétention time: 2.85 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 6: ierf-Butyl A-[2-[5-[l-benzyloxy-2-Zerr-butoxy-l-(trifluoromethyI)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyI]carbamate

A solution of ZerZ-butyl 7V-[2-[[[2-benzyloxy-3-ZerZ-butoxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (1.1 g, 1.4969 mmol) and DIPEA (682.64 mg, 0.92 mL, 5.2818 mmol) in acetonitrile (24 mL) was heated in an oil bath at 60 °C. Addedp-TsCl (322 mg, 1.6890 mmol) and the reaction was heated in the oil bath for 90 minutes. Once cooled to room température, the reaction was concentrated under reduced pressure to remove the acetonitrile. Transferred to a 250 mL separatory funnel with saturated aqueous sodium bicarbonate (100 mL) and the aqueous layer was extracted with ethyl acetate (2 X 75 mL). The combined organic layers were washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (80 g column, eluting from 0 % to 30 % ethyl acetate in heptanes) to afford as a pale amber oil, ZerZ-butyl A-[2-[5-[l-benzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3-enyl]l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (0.96 g, 88 %). *H NMR (400 MHz, CDCh) δ 10.26 (s, 1H), 9.37 (s, 1H), 7.57 - 7.51 (m, 2H), 7.46 - 7.38 (m, 2H), 7.36 7.30 (m, 1H), 6.01 (ddd, J= 17.5, 9.4, 8.3 Hz, 1H), 5.32 - 5.20 (m, 2H), 4.90 - 4.83 (m, 1H), 4.83 - 4.75 (m, 2H), 1.58 (s, 9H), 1.12 (s, 9H). ^,9F NMR (377 MHz, CDCI3) δ -63.73 (s, 3F), 66.73 (s, 3F). ESI-MS m/z cale. 708.1382, found 597.0 (M-l 11)⁺; Rétention time: 4.85 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

364

Step 7: ZerZ-Butyl 7V-[2-[5-[l-benzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazoI-2-yl]-6-bromo-5-(trifluoromethyI)-3-pyridyl]-7V-/eri-butoxycarbonyl-carbamate

A solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (1.21 g, 1.6697 mmol), DIPEA (237.44 mg, 0.32 mL, 1.8372 mmol) and DMAP (10 mg, 0.0819 mmol) in MTBE (25 mL) was treated with di-ZerZ-butyl dicarbonate (479 mg, 0.5042 mL, 2.1948 mmol) and stirred at room température for about 5 hours. The reaction mixture was transferred to a 125 mL separatory funnel with water (75 mL) and MTBE (100 mL). After extraction, the layers were separated, and the organic layer was washed with brine (75 mL). The organic layer was dried over sodium sulfate, fïltered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (80m g column, eluting from 0 % to 25 % ethyl acetate in heptanes) to afford as a colorless oil, ZerZ-butyl 7V-[2-[5-[ 1 -benzyloxy-2-ZerZ-butoxy-1(trifluoromethyl)but-3 -enyl] -1,3,4-oxadiazol-2-yl] -6-bromo-5-(trifluoromethyl)-3 -pyridyl] -NZerZ-butoxycarbonyl-carbamate (1.26 g, 91 %) that solidified on standing to a white solid. *H NMR (400 MHz, CDC1₃) δ 7.95 (s, 1H), 7.57 - 7.48 (m, 2H), 7.45 - 7.37 (m, 2H), 7.36 - 7.30 (m, 1H), 5.98 (ddd, J= 17.4, 9.7, 7.9 Hz, 1H), 5.28 - 5.15 (m, 2H), 4.86 - 4.72 (m, 3H), 1.44 (s, 18H), 1.08 (s, 9H). ¹⁹F NMR (377 MHz, CDCI3) δ -63.74 (s, 3F), -66.63 (s, 3F). ESI-MS m/z cale. 808.1906, found 597.0 (M-211)⁺; Rétention time: 4.57 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 8: ZerZ-Butyl 7V-[2-[5-[l-benzyloxy-2-/er/-butoxy-l-(trifIuoromethyl)but-3-enyl]-l,3,4oxadiazoI-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-A-ZerZ-butoxycarbonyl-carbamate

365

To a mixture of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonylcarbamate (1.18 g, 1.4575 mmol) in DMSO (12 mL) was added césium acetate (899 mg, 4.6835 mmol). The mixture was stirred at 85 °C for 5 h and cooled to rt. The mixture was transferred to a 250 mL separatory tunnel with saturated aqueous ammonium chloride (25 mL) and water (125 mL) and extracted with 1:1 mixture of MTBE and heptanes (3 X 50 mL). The combined organic layers were washed with water (2 X 50 mL), brine (50 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to afford as an off-white foamy solid, ZerZ-butyl N-[2[5-[l-benzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (1.2 g, 105 %, contaminated with residual MTBE). Ή NMR (400 MHz, CDC1₃) δ 7.75 (s, 1H), 7.45 - 7.30 (m, 5H), 7.16 7.08 (m, 1H), 6.03 - 5.91 (m, 1H), 5.31 - 5.21 (m, 2H), 4.91 - 4.75 (m, 3H), 1.47 (s, 9H), 1.42 (s, 9H), 1.07 (s, 9H). ¹⁹F NMR (377 MHz, CDCI3) δ -65.88 (br. s., 3F), -66.39 (s, 3F). ESI-MS m/z cale. 746.275, found 535.2 (M-211)⁺; Rétention time: 3.35 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 mm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Step 9: ZerZ-Butyl 7\-[2-[5-[l-benzyIoxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3-cnyl]-l,3,4oxadiazol-2-yl]-6-[(lÆ)-l-mcthylpent-4-enoxy]-5-(trifIuoromethyl)-3-pyridyI]-A-ZerZbutoxycarbonyl-carbamate

A solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-/V-ZerZ-butoxycarbonylcarbamate (1.0883 g, 1.4575 mmol) and (2S)-hex-5-en-2-ol (494 mg, 4.4389 mmol) in toluene (24 mL) was treated successively with triphenylphosphine (883 mg, 3.3666 mmol) and DIAD (677.82 mg, 0.66 mL, 3.3521 mmol) and the reaction was stirred at room température for 19 hours. Concentrated under reduced pressure and purified by silica gel chromatography (80 g column, eluting from 0 % to 20 % ethyl acetate in heptanes) to afford as a colorless oil, ZerZ-butyl A-[2-[5-[l-bcnzyloxy-2-ZerZ-butoxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol-2-yl]-6366

[(17î)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7V-/er/-butoxycarbonyl-carbamate (1.07 g, 68 %). ESI-MS m/z cale. 828.3533, found 617.2 (M-211)⁺; Rétention time: 5.09 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 10: 4-[5-[3-Amino-6-[(lR)-l-methylpent-4-enoxy]-5-(trifluoromethyI)-2-pyridyl]-l,3,4 oxadiazol-2-yl]-4-benzyIoxy-5,5,5-trifluoro-pent-l-en-3-oI

A solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-2-Zer/-butoxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-[(lR)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-3-pyridyl]-7VZe/7-butoxycarbonyl-carbamate (1.07 g, 1.2910 mmol) in dichloromethane (15 mL) and TFA (22.200 g, 15 mL, 194.70 mmol) was stirred at room température for 4.5 hours. The reaction was concentrated under reduced pressure and the residue was transferred to a 250 mL separatory tunnel with dichloromethane (50 mL) and saturated aqueous sodium bicarbonate (75 mL). After extraction, the layers were separated, and the aqueous layer was extracted again with dichloromethane (2 X 50 mL). The combined organic layers were washed with brine (50 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (40 g column, eluting from 0 % to 30 % ethyl acetate in heptanes) to afford as a yellow oil and about 1:1 diastereomers mixture of 4-[5-[3-amino-6[(17?)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazol-2-yl]-4-benzyloxy5,5,5-trifluoro-pent-l-en-3-ol (585 mg, 72 %). Ή NMR (400 MHz, CDCI3) δ 7.51 - 7.43 (m, 3H), 7.41 - 7.31 (m, 3H), 6.06 - 5.92 (m, 1H), 5.91 - 5.76 (m, 1H), 5.63 (br. s., 2H), 5.36 (d, J = 17.1 Hz, 1H), 5.25 (d, J= 10.3 Hz, 1H), 5.22 - 5.14 (m, 1H), 5.13 - 5.04 (m, 1H), 5.04 - 4.85 (m, 3H), 4.84 - 4.76 (m, 1H), 3.01 - 2.75 (m, 1H), 2.29 - 2.07 (m, 2H), 1.93 - 1.82 (m, 1H), 1.76 1.64 (m, 1H), 1.37 (d, J= 6.1 Hz, 3H, diastereomer A), 1.32 (d, J= 6.1 Hz, 3H, diastereomer B). ^I9F NMR (377 MHz, CDCI3) δ -64.19 (s, 3F), -68.24 (s, 3F, diastereomer A), -68.30 (s, 3F, diastereomer B). ESI-MS m/z cale. 572.1858, found 573.2 (M+l)⁺; Rétention time: 4.12 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow = 1.5 mL/min).

367

Step 11: (121î)-17-Amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Z mixture, diastereomer pair 1)

A solution of 4-[5-[3-amino-6-[(17?)-l-methylpent-4-enoxy]-5-(trifluoromethyl)-2pyridyl]-l,3,4-oxadiazol-2-yl]-4-benzyloxy-5,5,5-trifluoro-pent-l-en-3-ol (98.7 mg, 0.1572 mmol) in dichloroethane (50 mL) was bubbled with nitrogen gas for 21 hours. The solution was heated in an oil bath at 70 °C and treated with Zhan-IB catalyst (8 mg, 0.0109 mmol). After 30 minutes, a second portion of Zhan-IB catalyst (8 mg, 0.0109 mmol) was added and heated for 3 hours. The reaction mixture was cooled to room température, DMSO (4 drops) was added and concentrated under reduced pressure. The residue was purified by silica gel chromatography (24 g column, eluting from 0 % to 40 % ethyl acetate in heptanes) to afford as a dark amber oil, (127?)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Zmixture, diastereomer pair 1) (79.6 mg, 76 %). ESI-MS m/z cale. 544.1545, found 545.2 (M+l)⁺; Rétention time: 3.85 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 12: (127î)-17-Amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 1), Compound 56, and (12Z2)-17-amino-12-inethyl-6,15-bis(trifluoromethyI)-13,19-dioxa3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 2), Compound 57

enantiomer 1 enantiomer 2

A solution of (122?)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Z mixture, diastereomer pair 1) (25 mg, 0.0459 mmol) in methanol (2 mL) was purged three times with

368 nitrogen gas. Added palladium on carbon (30 mg, 5 %w/w, 0.0141 mmol), purged the reaction three times with hydrogen gas and the reaction was stirred at room température for 5 hours. The reaction was purged twice with nitrogen, filtered over a pad of celite and washed with methanol. The solvent was removed under reduced pressure and the residue was purified by silica gel chromatography (4 g column, eluting from 0 % to 40 % ethyl acetate in heptanes). The résultant solid was purified by SFC using Cellulose 4 column (150 X 12.2 mm, 5 pm particle size) using a dual gradient run from 10 % MeOH (no modifier) and 90 % CO2 with flow rate of 75 mL/min giving as a pale-yellow solid, (12J?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 1) (10.5 mg, 48 %). ^!H NMR (400 MHz, DMSO-d6) δ 7.75 (s, 1H), 7.50 (s, 1H), 6.32 (s, 2H), 5.38 (d, J= 8.8 Hz, 1H), 4.83 - 4.66 (m, 1H), 4.42 - 4.25 (m, 1H), 2.48 - 2.40 (m, 1H), 1.84 - 1.71 (m, 1H), 1.66 - 1.56 (m, 2H), 1.55 - 1.46 (m, 1H), 1.45 - 1.37 (m, 2H), 1.34 (d, J= 6.4 Hz, 3H), 1.21 - 1.10 (m, 1H). ^I9F NMR (377 MHz, DMSO-d6) δ -62.50 (s, 3F), -73.40 (s, 3F). ESI-MS m/z cale. 456.1232, found 457.1 (M+l)⁺; Rétention time: 3.38 minutes. Also, isolated as an off white solid, ( 12R)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 2) (9 mg, 42 %). ¹ H NMR (400 MHz, DMSO-d₆) δ 7.74 (s, 1H), 7.52 (s, 1H), 6.24 (s, 2H), 5.62 (d, J = 6.6 Hz, 1H), 5.04 - 4.86 (m, 1H), 4.19 - 3.99 (m, 1H), 2.43 - 2.29 (m, 1H), 2.20 - 2.08 (m, 1H), 2.07 - 1.95 (m, 1H), 1.64 - 1.43 (m, 3H), 1.32 (d, J= 6.4 Hz, 3H), 1.30 - 1.20 (m, 2H). ¹⁹F NMR (377 MHz, DMSO-dô) δ -62.46 (s, 3F), -73.70 (s, 3F). ESI-MS m/z cale. 456.1232, found 457.1 (M+l)⁺; Rétention time: 3.47 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

369

Example 40: Préparation of (67?,127?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,8-diol (diastereomer 2), Compound 58

Step 1: tert-Butyl A-[(6/?,127?)-6-benzyloxy-12-methyI-8-oxo-6,15-bis(trifluoromethyI)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17yljcarbamate

diastereomer 1

To a solution of /er/-butyl 7V-[(6R,127?)-6-benzyloxy-8-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaen-17-yl]carbamate (diastereomer 1) (120 mg, 0.1856 mmol) in CH2CI2 (20 mL) at 0 °C was added NaHCCfl (294 mg, 3.4997 mmol), followed by Dess-Martin periodinane (102 mg, 0.2405 mmol) and the mixture was stirred at rt ovemight. The mixture was treated with 10 % aq. Na2S2Û3 (5 mL) and stirred at rt for 5 min. The resulting mixture was extracted with CH2CI2 (3 X 15 mL). The combined organic layers were dried over Na2SÛ4, filtered and concentrated. The residue was purified by silica gel chromatography (24 g column, eluting from 0 % to 30 % EtOAc/heptanes) to afford as a colorless oil, tert-butyl 7V-[(67î,127?)-6-benzyloxy-12-methyl-8oxo-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (110 mg, 92 %). ’H NMR (400 MHz, CDCI3) δ 9.10 (br. s, 1H), 9.09 (s, 1H), 7.34-7.23 (m, 5H), 4.91 (d,J= 11.0 Hz, 1H), 4.89 - 4.81 (m, 1H),4.49 (d, J= 11.0 Hz, 1H), 3.41 (d, J= 15.2 Hz, 1H), 3.16 (ddd, J= 19.1, 10.2, 2.7 Hz, 1H), 3.08 (d, J = 15.2 Hz, 1H), 2.64 - 2.54 (m, 1H), 2.43 - 2.32 (m, 1H), 1.98 - 1.86 (m, 1H), 1.56 (s, 9H), 1.50 1.35 (m, 5H). ¹⁹F NMR (377 MHz, CDCI3) δ -63.89 (s, 3F), -74.36 (s, 3F). ESI-MS m/z cale.

370

644.207, found 589.1 (M-55)⁺; Rétention time: 5.46 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 mm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Step 2: (67?,12/î)-17-Amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-one

To a solution of ZerAbutyl 7V-[(6R,12R)-6-benzyloxy-12-methyl-8-oxo-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-17-yl]carbamate (26 mg, 0.0403 mmol) in CH2CI2 (2 mL) at 0 °C was added TFA (740.00 mg, 0.5 mL, 6.4899 mmol) dropwise. The mixture was stirred at 8 to 13 °C for 1 h. The solvents were removed by a gentle nitrogen flow at 10 °C with stirring. The residue was treated with sat. NaHCO₃ (5 mL) and extracted with CH2CI2 (3X8 mL). The combined organic layers were dried over Na2SO4, filtered and concentrated. The residue was purified by silica gel chromatography (12 g column, eluting from 0 % to 35 % EtOAc/heptanes) to afford as a paleyellow solid, (6/?,12/?)-17-amino-6-bcnzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-one (21 mg, 96 %). ‘H NMR (400 MHz, CDC1₃) δ 7.43 (s, 1H), 7.35 - 7.27 (m, 5H), 5.27 (br. s, 2H), 4.89 (d, J= 11.2 Hz, 1H), 4.87 - 4.80 (m, 1H), 4.47 (d, J= 11.0 Hz, 1H), 3.39 (d, J= 15.2 Hz, 1H), 3.30 - 3.20 (m, 1H), 3.06 (d, J= 14.9 Hz, 1H), 2.62 - 2.53 (m, 1H), 2.43 - 2.32 (m, 1H), 1.98 - 1.87 (m, 1H), 1.47 - 1.41 (m, 1H), 1.39 (d, J= 6.4 Hz, 3H), 1.37 - 1.32 (m, 1H). ^I9FNMR (377 MHz, CDC1₃) δ -63.99 (s, 3F), -74.33 (s, 3F).

Step 3: (6R,12Z2)-17-Amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-ol (diastereomer 2)

371

To a solution of (67?,127î)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-8-one (21 mg, 0.0386 mmol) in MeOH (3 mL) at 0 °C was added tétraméthylammonium borohydride (9.6 mg, 0.1079 mmol). The mixture was stirred at 0 °C for 1.5 h. Acetone (0.5 mL) was added and the reaction mixture was stirred at 0 °C for 5 min. The mixture was treated with sat. NalICCh (5 mL) and extracted with EtOAc (3X10 mL). The combined organic layers were dried with Na2SO₄, filtered and concentrated. The residue was purified by silica gel chromatography (24 g column, eluting from 0 % to 35 % EtOAc/heptanes) to afford as a pale-yellow oil, (67?,127?)-17-amino-6benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-ol (diastereomer 2) (20 mg, 95 %). Ή NMR (400 MHz, CDC1₃) δ 7.45 (s, 1H), 7.33 - 7.27 (m, 2H), 7.27 - 7.20 (m, 3H), 5.25 (br. s, 2H), 5.04 - 4.94 (m, 2H), 4.93 - 4.82 (m, 1H), 4.57 (d, J= 11.2 Hz, 1H), 2.79 (br. s, 1H), 2.57 (dd, J= 15.2, 8.3 Hz, 1H), 2.35 - 2.24 (m, 1H), 2.12 (d, J= 15.2 Hz, 1H), 1.90 - 1.74 (m, 2H), 1.71 -1.58 (m, 3H), 1.40 (d, J = 6.4 Hz, 3H). ^I9F NMR (377 MHz, CDCI3) δ -64.03 (s, 3F), -74.22 (s, 3F).

Step 4: (67?,127?)-17-Amino-12-mcthyI-6,15-bis(trifliioromethyI)-13,19-dioxa-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,8-diol (diastereomer 2), Compound 58

A mixture of (67?, 127?)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-ol (diastereomer 2) (23 mg, 0.0421 mmol) and palladium on carbon (9 mg, 10 % 50 % wet, 0.0042 mmol) in MeOH (4 mL) was stirred under hydrogen balloon at rt ovemight. The mixture was filtered through Celite and washed with EtOAc. The filtrate was concentrated and the residue was purified by silica gel chromatography (24 g column, eluting from 20 % to 50 % EtOAc/pentane) and freeze dried to afford as a pale-yellow solid (67?,127î)-17-amino-12-methyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[l 2.3.1. 12,5]nonadeca1(18),2,4,14,16-pentaene-6,8-diol (diastereomer 2) (15 mg, 78 %). *H NMR (400 MHz, DMSOd₆) δ 7.76 (s, 1H), 7.69 (s, 1H), 6.29 (s, 2H), 5.06 - 4.97 (m, 1H), 4.82 (d, J= 5.1 Hz, 1H), 4.47 4.37 (m, 1H), 2.29 - 2.20 (m, 1H), 2.13 - 1.92 (m, 3H), 1.72 - 1.52 (m, 2H), 1.49 - 1.37 (m, 1H), 1.33 (d, J= 6.4 Hz, 3H), 1.30 - 1.23 (m, 1H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.48 (s, 3F), 372

78.47 (s, 3F). ESI-MS m/z cale. 456.1232, found 457.2 (M+l)⁺; Rétention time: 3.04 minutes.

LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Example 41: Préparation of (6/?,12Æ)-17-amino-12-methyI-6,15-bis(trifluoromethyI)5 10,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI,

Compound 59

° X °^Λΐ° o \ / 1 \ H | Step 5 HN'^NYtc^F3 ‘ Ο^ΛΟ° O -X-Si f ___NH HN-NH A ° O \ \z , J\-Si f L J*’’. yY°x^CF3 \_O. \ \ zf T r X, / ° WV — O^N^qN-N 1 °η< U¹ :

^H0> JL/CX \ ^HO \ Step 2 H 1 Step 3 > Step 4 Λ —‘ ' ° ?° ° M x M 0 0 Br H L· (λ O ^S,ep6 »··^!Υ * —- o X '___O.___NH OH k < T o ° \ \z , X-Si f Step 7 ^CF3V^N > Step8 T X Y \X)^NH N-N ° O ”^^Sl'°'^_o ^HO'^-O OH > Step 10 | Step 11 ^CF3yU VS°0^cf3 TxVb^F3 .ΟγΝγΟΝ'Ν °'\ ΧΓ°Υ^ΝΎ°^Ν-Ν 1 O O^ 0 O O^x 0 :f₃^A_n \ ?'^°Ί 1 1 „ 1 Step 13 _rF T I YX^O^-CF₃ ---- ^cf3-₁A_n X | \\ // OH H S ο X-rr O^N ON-N VXVXÎ³ ^o ο^χ n^h2 n-n

373

Step 1: tert-Butyl 7V-[[(21î)-2-benzyIoxy-2-(trifluoromethyl)pent-4-enoyl] amino]carbamate

A reaction vial was charged with (2R)-2-benzyloxy-2-(trifluoromethyl)pent-4enehydrazide (3.479 g, 12.069 mmol) and TEA (3.7026 g, 5.1 mL, 36.591 mmol) in DCM (30 mL). BociO (3.26 g, 14.937 mmol) was added and the reaction mixture was stirred at rt ovemight. The reaction was diluted with DCM (100 mL) and washed with 1 N HCl (30 mL), saturated sodium bicarbonate (30 mL) and brine (30 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by silica gel chromatography using 0 % to 30 % ethyl acetate in hexane to fumish as a white solid, ZerZ-butyl 7/-[[(27?)-2-benzyloxy-2-(trifluoromethyl)pent-4-enoyl]amino]carbamate (3.914 g, 84 %). ESIMS m/z cale. 388.161, found 333.2 (M-55)⁺; Rétention time: 3.46 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 2: tert-Butyl A-[[(27?)-2-benzyloxy-5-hydroxy-2-(trifluoromcthyl)pcntanoyI] amino] carbamate

Into a solution of ZerAbutyl 2V-[[(2J?)-2-benzyloxy-2-(trifluoromethyl)pent-4enoyl]amino]carbamate (3.75 g, 9.6555 mmol) in anhydrous THF (53 mL) was added 9Borabicyclo[3.3.1]nonane (105 mL of 0.5 M in THF, 52.500 mmol) dropwise at rt and the reaction was stirred at rt for 80 minutes. The reaction was quenched with methanol (16 mL) at 0 °C slowly and NaOH (105 mL of 1 M, 105.00 mmol), H2O2 (17.649 g, 53 mL of 30 %w/w, 155.66 mmol) was added. The reaction was stirred at rt for 1 hour and diluted with ethyl acetate (320 mL). Two layers were separated, and the aqueous layer was extracted with ethyl acetate (2 X 95 mL). The combined organic layers were washed with saturated sodium thiosulfate (130 mL) and brine (130 mL), dried over anhydrous magnésium sulfate and concentrated. The residue was purified by silica gel chromatography using 10 % to 50 % acetone in hexane to provide as a

374 white solid, ZerZ-butyl A-[[(27?)-2-benzyloxy-5-hydroxy-2-(trifluoromethyl)pentanoyl] amino]carbamate (3.76 g, 96 %). ESI-MS m/z cale. 406.1716, found 407.5 (M+l)⁺; Rétention time: 3.02 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 3: ZerZ-Butyl 7V-[[(2Æ)-2-benzyloxy-5-[(25)-2-hydroxypropoxy]-2(trifluoromethyl)pentanoyl]amino] carbamate

To a solution of ZerZ-butyl A-[[(27?)-2-benzyloxy-5-hydroxy-2(trifluoromethyl)pentanoyl] amino]carbamate (3.34 g, 8.2186 mmol) in THF (35 mL) was added Z-BuOK in THF (33 mL of 1 M, 33.000 mmol) at 0 °C and stirred for 10 minutes. (25)-2methyloxirane (994.80 mg, 1.2 mL, 17.128 mmol) was added slowly at 0 °C and stirred for 10 minutes. The reaction mixture was stirred at 45 °C for 1.5 hours. The reaction mixture was cooled to room température and quenched with saturated aqueous NH4CI (70 mL). The mixture was extracted with EtOAc (3 X 50 mL), washed with brine (70 mL), dried over Na2SÜ4, filtered, and concentrated. The residue was purified by silica gel chromatography using 0 % to 50 % EtOAc in hexane to provide as pale yellow oil, ZerZ-butyl 7V-[[(2Æ)-2-benzyloxy-5-[(25)-2hydroxypropoxy]-2-(trifluoromethyl)pentanoyl]amino] carbamate (1.65 g, 42 %) ESI-MS m/z cale. 464.2134, found 465.4 (M+l)+; Rétention time: 3.06 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 4: ZerZ-Butyl A-[[(2Æ)-2-benzyIoxy-5-[(25')-2-[ZerZ-butyl(dimethyl)silyl]oxypropoxy]-2(trifluoromethyl)pentanoyl] amino] carbamate

375

To a solution of ZerZ-butyl 7V-[[(27?)-2-benzyloxy-5-[(25)-2-hydroxypropoxy]-2(trifluoromethyl)pentanoyl]amino]carbamate (1.65 g, 3.4458 mmol) in DMF (35 mL) was added TBSC1 (1.08 g, 7.1655 mmol) followed by Imidazole (972.5 mg, 14.285 mmol). The reaction mixture was stirred at room température for 3 hours. The reaction mixture was quenched with saturated aqueous NH4CI (70 mL), extracted with EtOAc (3 X 50 mL), washed with brine (70 mL), dried over NaiSO4, filtered, and concentrated. The residue was purified by silica gel chromatography using 0 % to 15 % EtOAc in hexane to provide as light yellow gel, ZerZ-butyl N[[(2Æ)-2-benzyloxy-5-[(2S')-2-[terf-butyl(dimethyl)silyl]oxypropoxy]-2(trifluoromethyl)pentanoyl]amino]carbamate (1.683 g, 84 %). ESI-MS m/z cale. 578.2999, found 579.7 (M+l)⁺; Rétention time: 4.47 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 5: (2R)-2-BenzyIoxy-5-[(2S')-2-[terZ-butyl(dimethyl)siIyl]oxypropoxy]-2(trifluoromethyl)pentanehydrazide

A solution of ZerZ-butyl 7V-[[(27?)-2-benzyloxy-5-[(2S)-2-[ZerZbutyl(dimethyl)silyl]oxypropoxy]-2-(trifluoromethyl)pentanoyl]amino]carbamate (1.772 g, 3.0618 mmol) in HFIP (32 mL) was heated in a microwave reactor at 100 °C for 1.5 h. The reaction was concentrated and the residue was purified by silica gel chromatography by using 0 % to 80 % EtOAc in hexanes to afford as clear gel (27?)-2-benzyloxy-5-[(25)-2-[/erZbutyl(dimethyl)silyl]oxypropoxy]-2-(trifluoromethyl)pentanehydrazide (1.442 g, 98 %). ESI-MS m/z cale. 478.2475, found 479.2 (M+l)⁺; Rétention time: 3.96 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

376

Step 6: tert-Butyl A-[2-[[[(2Æ)-2-benzyloxy-5-[(2S)-2-[ri’/7‘-butyl(dimethyI)silyl|oxypropoxy]2-(trifluoromethyl)pentanoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3pyridyl] carbamate

Into a solution of (27î)-2-benzyloxy-5-[(25)-2-[ZerZ-butyl(dimethyl)silyl]oxypropoxy]-2(trifluoromethyl)pentanehydrazide (1.44 g, 3.0086 mmol) and 6-bromo-3-(terZbutoxycarbonylamino)-5-(trifluoromethyl)pyridine-2-carboxylic acid (1.38 g, 3.5832 mmol) in anhydrous DMF (50 mL) was added TEA (3.6518 g, 5.03 mL, 36.089 mmol), followed by T3P (2.213 g, 2.07 mL of 50 %w/w, 3.4776 mmol) in ethyl acetate. The reaction was stirred at rt for ovemight. Again, to the reaction mixture was added TEA (3.6518 g, 5.03 mL, 36.089 mmol), followed by T3P (2.213 g, 2.07 mL of 50 %w/w, 3.4776 mmol) in ethyl acetate. The reaction mixture was stirred at room température for 2 hours. The reaction was quenched with saturated ammonium chloride (100 mL) and ethyl acetate (150 mL). The aqueous layer was extracted with ethyl acetate (2 X 100 mL). The combined organic layers were washed with brine (2 X 100 mL), dried over anhydrous sodium sulfate and concentrated. The residue was purified by silica gel chromatography using 0 % to 20 % acetone in hexane to provide as off white foam, ier/-butyl N[2-[[[(2Æ)-2-benzyloxy-5-[(2S)-2-[tert-butyl(dimethyl)silyl]oxypropoxy]-2(trifluoromethyl)pentanoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (1.4724 g, 56 %). ESI-MS m/z cale. 844.2302, found 845.5 (M+l)⁺; Rétention time: 4.88 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 7: tert-Butyl A^r-[2-[5-[(lR)-l-benzyIoxy-4-[(2S)-2-[fertbutyI(dimethyl)silyl]oxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-bromo5-(trifluoromethyl)-3-pyridyl]carbamate

377

Into a solution of ZerZ-butyl 7V-[2-[[[(27?)-2-benzyloxy-5-[(2S)-2-[ZerZbutyl(dimethyl)silyl]oxypropoxy]-2-(trifluoromethyl)pentanoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (1.457 g, 1.6711 mmol) and DIEA (934.92 mg, 1.26 mL, 7.2338 mmol) in acetonitrile (25 mL) was addedp-TsCl (359.8 mg, 1.8873 mmol). The reaction was stirred at 70 °C for 1.5 hours. Solvent was removed and the residue was purified by silica gel chromatography using 0 % to 20 % ethyl acetate in hexane to provide as yellow gel, ZerZbutyl7V-[2-[5-[(17?)-l-benzyloxy-4-[(2S)-2-[ZerZ-butyl(dimethyl)silyl]oxypropoxy]-l(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (1.2036 g, 87 %). ESI-MS m/z cale. 826.2196, found 827.5 (M+l)⁺; Rétention time: 5.25 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 8: fôr/-Butyl 7V-[2-[5-[(llî)-l-benzyloxy-4-[(25)-2-[ieributyI(dimethyl)siIyl]oxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-bromo5-(trifluoromethyl)-3-pyridyl]-7V-teri-butoxycarbonyI-carbamate

To a mixture of ZerZ-butyl 7V-[2-[5-[(lÆ)-l-benzyloxy-4-[(2S)-2-[/erZbutyl(dimethyl)silyl]oxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (1.2036 g, 1.4541 mmol) and DIEA (296.80 mg, 0.4 mL, 2.2964 mmol) in MTBE (14 mL) at room température was added DMAP (17.8 mg, 0.1457 mmol) followed by (Boc)2O (539.8 mg, 2.4733 mmol). The reaction mixture was stirred at room température for 17 hours. The mixture was diluted with EtOAc (60 ml), washed with water (30 mL), brine (30 mL), dried over Na2SO4, filtered and concentrated. The residue was purified by silica gel chromatography (40 g column, from 0 % toi0 % EtOAc in Hexanes,) to afford as light yellow gel, ZerZ-butyl _/V-[2-[5-[(17?)-l-benzyloxy-4-[(2S)-2-[ZerZbutyl(dimethyl)silyl]oxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (1.34 g, 99 %). ESI-MS m/z cale. 926.272, found 827.3 (M+H-Boc)⁺; Rétention time: 5.01 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 %

378 to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 9: férZ-Butyl7V-[2-[5-[(LR)-l-benzyIoxy-4-[(2.S)-2-[tert‘butyI(dimethyI)siIyI]oxypropoxy]-l-(trifIuoromethyl)butyl]-l,3,4-oxadiazoI-2-yl]-6hydroxy-5-(trifluoromethyI)-3-pyridyl]-7V-teri-butoxycarbonyl-carbamate

To a solution of terAbutyl 7V-[2-[5-[(17?)-l-benzyloxy-4-[(25)-2-[rerZbutyl(dimethyl)silyl] oxypropoxy] -1 -(trifluoromethyl)butyl]-1,3,4-oxadiazol-2-yl]-6-bromo-5(trifluoromethyl)-3-pyridyl]-7V-terZ-butoxycarbonyl-carbamate (1.3 g, 1.4011 mmol) in DMSO (14 mL) was added césium acetate (808.8 mg, 4.2136 mmol). The reaction mixture was stirred at 80 °C for 7 hours. The reaction mixture was cooled to room température and diluted with EtOAc (120 mL) and water (40 mL). Layers were separated, and the organic layer was washed with water (3 X 30 mL), brine (30 mL), dried over Na2SÛ4, fïltered, and concentrated. The residue was purified by silica gel chromatography using 0 % to 50 % EtOAc in hexanes to provide as white foam, terAbutyl 7V-[2-[5-[(lR)-l-benzyloxy-4-[(2S)-2-[terZbutyl(dimethyl)silyl]oxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5(trifluoromethyl)-3-pyridyl]-jV-/erZ-butoxycarbonyl-carbamate (1.0947 g, 90 %). 'H NMR (500 MHz, CDCI3) δ 7.75 (s, 1H), 7.35 - 7.28 (m, 5H), 4.84 (d, J= 10.9 Hz, 1H), 4.62 (d, J= 10.8 Hz, 1H), 4.00 - 3.91 (m, 1H), 3.51 - 3.42 (m, 2H), 3.37 (dd, J= 9.5, 6.1 Hz, 1H), 3.24 (dd, J= 9.5, 5.1 Hz, 1H), 2.51 (t, J= 8.2 Hz, 2H), 1.86-1.77 (m, 1H), 1.76-1.67 (m, 1H), 1.40 (d,J= 4.5 Hz,18H), 1.15 (d, J= 6.2 Hz, 3H), 0.88 (s, 9H), 0.07 (d, J= 7.2 Hz, 6H). ESI-MS m/z cale. 864.3564, found 765.8 (M+H-Boc)⁺; Rétention time: 4.75 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

379

Step 10: fert-Butyl 7V-[2-[5-[(lE)-l-benzyloxy-4-[(2S)-2-hydroxypropoxy]-l(trifluoromethyl)butyl]-l,3,4-oxadiazoI-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-AterZ-butoxycarbonyl-carbamate

To a solution of ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-4-[(25)-2-[ZerZbutyl(dimethyl)silyl]oxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (1.079 g, 1.2475 mmol) in THF (10 mL) was added TBAF in THF (12.5 mL of 1 M, 12.500 mmol), and stirred at 35 °C for 3 hours. To the reaction mixture was added aqueous saturated NH4CI (80 mL) and extracted with EtOAc (3 X 50 mL). The combined organic layers were washed with water (50mL), brine (50mL), dried over Na₂SO4, filtered, and concentrated. The residue was purified by silica gel chromatography using 0 % to 80 % ethyl acetate in hexane to provide as white foam, ZerZ-butyl 7V-[2-[5-[(lE)-l-benzyloxy-4-[(2S)-2-hydroxypropoxy]-l-(trifluoromethyl)butyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-terZ-butoxycarbonyl-carbamate (916.6 mg, 98 %). ’H NMR (500 MHz, CDCI3) δ 7.81 (s, 1H), 7.36 - 7.27 (m, 5H), 4.90 (d, J= 10.8 Hz, 1H), 4.78 (d, J= 10.8 Hz, 1 H), 4.17 - 4.10 (m, 1H), 3.57-3.51 (m, 1H), 3.47 (dd, J= 9.0,3.0 Hz, 1H), 3.45-3.39 (m, 1H), 3.24 (t,J= 8.9 Hz, 1H), 2.64-2.44 (m, 2H), 1.70-1.59 (m, 2H), 1.40 (d, J= 7.0 Hz, 18H), 1.23 (d, J= 6.4 Hz, 3H). ESI-MS m/z cale. 750.2699, found 651.4 (M+H-Boc)⁺; Rétention time: 3.75 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Ci8 column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2II). Mobile phase B = acetonitrile (0.1 % CF₃CO₂H).

Step 11: ZerZ-Butyl A-[(6E,12E)-6-benzyloxy-12-mcthyl-6,15-bis(trifluoromethyl)-10,13,19trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-7V-ZerZbutoxycarbonyl-carbamate

380

A solution of ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-4-[(2S)-2-hydroxypropoxy]-l(trifluoromethyl)butyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZbutoxycarbonyl-carbamate (845.7 mg, 1.1266 mmol) and PPha (778.2 mg, 2.9670 mmol) in THF (75 mL) at room température was added DIAD (605.52 mg, 0.58 mL, 2.9945 mmol) drop-wise. The reaction mixture was stirred at room température for 5 hours. The reaction mixture was concentrated and the residue was purified directly by silica gel chromatography using 0 % to 15 % ethyl acetate in hexane to provide as white foam, ZerZ-butyl /V-[(67?,127?)-6-benzyloxy-12methyl-6,15-bis(trifluoromethyl)-l 0,13,19-trioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-17-yl]-N-tert-butoxycarbonyl-carbamate (416.8 mg, 50 %). ESI-MS m/z cale. 732.2594, found 733.5 (M+l)⁺; Rétention time: 4.36 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 12: terf-Butyl A^r-ter/-butoxycarbonyI-7V-[(6R,12Jî)-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-10,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate

To a solution of ZerZ-butyl 7V-[(6R,127î)-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-10,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate (391.2 mg, 0.5339 mmol) in éthanol (45 mL) was added 10 % Pd/C (200.3 mg, 1.8822 mmol). The reaction mixture was degassed and refilled hydrogen gas (X 2), and the reaction mixture was stirred under hydrogen atmosphère at room température for 16 hours. The reaction mixture was filtered through Celite, rinsed with EtOAc (30 mL), and concentrated to provide as a white foam, ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V[(67?, 12/?)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-10,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (352.4 mg, 100 %). ESI-MS m/z cale. 642.2124, found 643.2 (M+l)⁺; Rétention time: 3.85 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

381

Step 13: (67?,127?)-17-Amino-12-methyI-6,15-bis(trifluoromethyI)-10,13,19-trioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 59

A solution of ZerZ-butyl jV-/er/-butoxycarbonyl-A-[(67?,127?)-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-10,13,19-trioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-l ( 18),2,4,14,16pentaen-17-yl]carbamate (352.4 mg, 0.5320 mmol) in HFIP (15 mL) was placed in a microwave vial and sealed. It was heated at 100 °C in microwave synthesizer for 2 hours. The reaction was concentrated and the residue was purified by silica gel chromatography by using 0 % to 35 % EtOAc in hexanes to provide as off white solid, (67?,127?)-I7-amino-12-methyl-6,15bis(trifluoromethyl)-10,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (212.6 mg, 88 %). *H NMR (500 MHz, CDC1₃) δ 7.43 (s, 1H), 5.20 (s, 2H), 4.90 (q, .7=6.9 Hz, 1H), 4.49 (d, <7=8.9 Hz, 1H), 3.97-3.88 (m, 1H), 3.62 - 3.53 (m, 2H), 3.19 (t, J = 8.5 Hz, 1H), 2.45 (t, J= 12.1 Hz, 1H), 2.30-2.14 (m, 2H), 1.77-1.68 (m, 1H), 1.52 (d, J =6.4 Hz, 3H). ESI-MS m/z cale. 442.10757, found 443.2 (M+l)⁺; Rétention time: 2.59 minutes. LCMS Method: Cortess Cis 2.7 pm particle size column (3.0 X 50 mm) sold by Waters (pn: 186007370), and a dual gradient run from 5 % to 100 % mobile phase B over 6.0 minutes. Mobile phase A = water (+ 0.1% CF3CO2H), mobile phase B = acetonitrile (0.1% CF3CO2H), flow rate =1.2 mL/min, injection volume = 2 pL and column température = 55 °C.

Example 42: Préparation of (6R,12R)-17-amino-6-hydroxy-12-methyl-6,15- bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-8-one, Compound 60

Step 1: (6R,12R)-17-Amino-6-hydroxy-12-methyI-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-one, Compound 60

[0001] To a solution of (6Æ,12Æ)-17-amino-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-8-one (32 mg, 0.0588 mmol) in MeOH (4 mL) was added palladium on carbon (9.5 mg, 382 %, 50 % wet 0.0045 mmol). The mixture was stirred under hydrogen atmosphère at rt ovemight. The mixture was filtered through diatomaceous earth and washed with EtOAc. The filtrate was concentrated and the residue was purified by silica gel chromatography (24 g column, eluting 20 % to 40 % EtOAc/pentane) to afford as a pale-yellow solid, (67?, 127?)-175 amino-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-8-one (21 mg, 76 %). *H NMR (400 MHz, DMSO-dô) δ 8.03 (s, 1H), 7.76 (s, 1H), 6.38 (s, 2H), 4.94 - 4.81 (m, 1H), 3.50 (d, J= 15.4 Hz, 1H), 3.30 - 3.23 (m, 1H), 3.01 (d, J= 15.4 Hz, 1H), 2.70 (ddd, J= 19.0, 6.4, 2.9 Hz, 1H), 2.35 - 2.20 (m, 1H), 1.89 - 1.74 (m, 1H), 1.44 - 1.27 (m, 5H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ

-62.58 (s, 3F), -76.69 (s, 3F). ESI-MS m/z cale. 454.1076, found 455.1 (M+l)⁺; Rétention time:

3.02 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Example 43: Préparation of (6J?)-17-amino-ll,ll-dimethyI-6,15-bis(trifluoromethyl)-13,1915 dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound

EJZ. mixture

383

Step 1: fôrt-Butyl 7V-[2-[5-[(LR)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4-oxadiazol2-yl]-6-(2,2-dimethylpent-4-enoxy)-5-(trifIuoromethyl)-3-pyridyI]-7V-terZ-butoxycarbonylcarbamate

A solution of Ze/7-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZ-butoxycarbonyl-carbamate (1.38 g, 1.8473 mmol) and 2,2-dimethylpent-4-en-l-ol (630 mg, 5.5174 mmol) in toluene (17 mL) was treated with triphenylphosphine (998 mg, 3.8050 mmol) followed by DIAD (770.25 mg, 0.75 mL, 3.8092 mmol) at room température. The yellow solution was stirred at room température for 24 hours. The yellow suspension was concentrated and the residue was purified by reverse phase chromatography (100 g Cl 8 column, eluting with 0 % to 90 % acetonitrile in water with 0.1 w/w % of formic acid for 5 column volumes followed by isocratic elution with 90 % acetonitrile in water with 0.1 w/w % of formic acid for 10 column volumes) to provide as a light yellow oil, ZerZ-butyl 7V-[2-[5-[(l/?)-l-benzyloxy-l-(trifluoromethyl)but-3-enyl]-l,3,4oxadiazol-2-yl]-6-(2,2-dimethylpent-4-enoxy)-5-(trifluoromethyl)-3-pyridyl]-7V-terrbutoxycarbonyl-carbamate (730 mg, 51 %). ESI-MS m/z cale. 770.3114, found 615.2 (M-155)⁺; Rétention time: 4.87 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 2: tert-Butyl A-[(67?)-6-benzyIoxy-ll,ll-dimethyI-6,15-bis(trifluoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yl]-7V-tertbutoxycarbonyl-carbamate (E/Z mixture)

E/Z mixture

A dried 250 mL flask was charged with toluene (100 mL) and the solvent was bubbled with nitrogen for 30 min. Zhan IB catalyst (40 mg, 0.0545 mmol) was added under gentle flow 384 of nitrogen. The mixture was bubbled with nitrogen for more 10 min and heated to 110 °C. A nitrogen-bubbled solution of ZerZ-butyl 7V-[2-[5-[(17?)-l-benzyloxy-l-(trifluoromethyl)but-3enyl]-l,3,4-oxadiazol-2-yl]-6-(2,2-dimethyIpent-4-enoxy)-5-(trifluoromethyl)-3-pyridyl]-A⁷-ZerZbutoxycarbonyl-carbamate (200 mg, 0.2595 mmol) in toluene (20 mL) was added dropwise over 0.5 h. After addition was completed, the mixture was bubbled with nitrogen for 5 min. The mixture was continued to stir at 110 °C for 1.5 hours. Again, Zhan IB catalyst (40 mg, 0.0545 mmol) was added in two portions while heating the reaction mixture at 111 °C for 2 hours. The mixture was then cooled to room température and the catalyst was quenched by the addition of DMSO (4-5 drops). The mixture was concentrated and the residue was purifîed by reverse phase chromatography (50 g Cl 8 column, eluting with 5 % to 90 % acetonitrile in water containing 0.1 w/w % formic acid for 10 CV then with 90 % acetonitrile in water with 0.1 w/w % of formic acid for 10 CV) to provide as brown oil, ZerZ-butyl 7V-[(67?)-6-benzyloxy-l 1,1 l-dimethyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,8,14,16hexaen-17-yl]-7V-ZerZ-butoxycarbonyl-carbamate (E/Z mixture) (23 mg, 10 %). ESI-MS m/z cale. 742.2801, found 587.2 (M-155)⁺; Rétention time: 4.61 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 3: Z^rZ-Butyl 7V-fôrt-butoxycarbonyl-7V-[(6.R)-6-hydroxy-ll,ll-dimethyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate rCF₃OH

Ck JD

E/Z mixture

A solution of ZerZ-butyl 7V-[(67?)-6-benzyloxy-l 1,1 l-dimethyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-17-yl]-7V-ZerZbutoxycarbonyl-carbamate (E/Z mixture) (21 mg, 0.0283 mmol) in methanol (5 mL) was purged three times (vacuum then nitrogen gas). Added palladium on carbon (5 mg, 10% w/w, 50 % wet 0.0023 mmol), purged twice with hydrogen gas and stirred under hydrogen atmosphère for ovemight. The reaction mixture was purged with nitrogen gas, filtered with a nylon 0.45 micron filter and concentrated to provide as a colorless oil, ZerZ-butyl A^r-ZerZ-butoxycarbonyl-A^L[(6/?)-6hydroxy-11,11 -dimethyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18385 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (14 mg, 76 %).

ESI-MS m/z cale. 654.2488, found 499.2 (M-155)⁺; Rétention time: 4.23 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 4: (6/?)-17-Amino-ll,ll-dimethyI-6,15-bis(trifIuoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI, Compound 61

To a solution of ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(67?)-6-hydroxy-l 1,11-dimethyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate (14 mg, 0.0214 mmol) in dichloromethane (1 mL) was added 2,2,2-trifluoroacetic acid (1.4800 g, 1 mL, 12.980 mmol). The mixture was stirred at room température for 2 hours. The mixture was diluted with dichloromethane (5 mL) and concentrated. The residue was dissolved in ethyl acetate (40 mL), washed with aqueous saturated solution of sodium bicarbonate (3X10 mL), brine (1 X 20 mL), dried over anhydrous sodium sulphate, filtered and concentrated. The material was purified by reverse phase chromatography (15.5g C18 column, using a gradient from 0 % to 95 %water (+0.1v% FA) and MeCN) to provide as an off-white solid, (6/?)-17-amino-l 1,1 l-dimethyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (2.6 mg, 26 %). Ή NMR (400 MHz, CD3OD) δ 7.65 (s, 1H), 4.47 (br. s., 1H), 4.17 (br. s., 1H), 2.44 - 2.30 (m, 2H), 2.14 - 2.09 (m, 1H), 2.05 - 1.92 (m, 2H), 1.46 - 1.40 (m, 2H), 1.26 - 1.21 (m, 1H), 0.98 (s, 3H), 0.96 (s, 3H). ¹⁹F NMR (377 MHz, CD3OD) δ -65.62 (s, 3F), -80.82 (br. s., 3F). ESI-MS m/z cale. 454.144, found 455.1 (M+l)⁺; Rétention time: 3.95 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

386

Example 44: Préparation of (12Æ)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeea-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 3), Compound 62

diastereomer pair 1

EZZ mixture

diastereomer pair 2 E/Z mixture, enantiomer 3 enantiomer 3

Step 1: (12R)-17-Amino-6-benzyIoxy-12-methyl-6,15-bis(trifluoromethyI)-13,19-dioxa 3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-one (E/Z mixture)

diastereomer pair 1

E/Z mixture

To a solution of (127?)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Z mixture, diastereomer pair 1) (30 mg, 0.0551 mmol) in CH2CI2 (3 mL) was added NaHCCh (90 mg, 1.0713 mmol), followed by Dess-Martin periodinane (30 mg, 0.0707 mmol). The mixture was stirred at room température for 3 h. Saturated NaHCCh (3 mL) and 10 % aq. Na2S2Os (3 mL) were added. The mixture was stirred at room température for 5 min and extracted with CH2CI2 (3X10 mL). The combined organic layers were dried with Na2SÜ4, filtered and concentrated. The residue was purifîed by silica gel chromatography (eluting from 0 % to 30 % EtOAc/heptanes) to afford as a pale-yellow oil, (12Æ)-17-amino-6-benzyloxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,8,14,16hexaen-7-one (E/Z mixture) (24 mg, 80 %). *H NMR (400 MHz, CDCI3) δ 7.45 - 7.26 (m, 4H), 7.27 - 7.13 (m, 3H), 7.06 - 6.76 (m, 1H), 5.35 - 5.19 (m, 2H), 5.07 - 4.88 (m, 2H), 4.76 - 4.69 (m,

387

1H). 2.51 - 2.08 (m, 3H), 1.76 - 1.66 (m, 1H), 1.47 - 1.42 (m, 3H). ¹⁹F NMR (377 MHz, CDC1₃) δ -63.98 to -64.01 (m, 3F), -70.18 to -70.70 (m, 3F).

Step 2: (127?)-17-Amino-6-benzyIoxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-oI (E/Z mixture, diastereomer pair 2)

E/Z mixture

E/Z mixture diastereomer pair 2

To a solution of (127?)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-one (E/Z mixture) (28 mg, 0.0516 mmol) in MeOH (2.5 mL) at -5 °C was added tétraméthylammonium borohydride (10 mg, 0.1124 mmol). The mixture was stirred at -5 °C for 10 min. Acetone (0.5 mL) was added. The mixture was stirred at -5 °C for 5 min and then treated with saturated aqueous NaHCOs (5 mL). The mixture was extracted with EtOAc (3X10 mL). The combined organic layers were dried with Na₂SO4, filtered and concentrated. The residue was purified by silica gel chromatography (eluting 5 % EtOAc/CH₂Cl₂) to afford as a pale-yellow oil, (127?)17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Zmixture, diastereomer pair 2) (22 mg, 78 %). ESI-MS m/z cale. 544.15454, found 545.2 (M+l)⁺; Rétention time: 3.49 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 3: (127?)-17-Amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/L mixture, enantiomer 3)

E/Z mixture diastereomer pair 2

E/Z mixture, enantiomer 3 (127î)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Zmixture, diastereomer

388 pair 2) (40 mg, 0.0735 mmol) was purified by silica gel chromatography (eluting 10 to 30 % EtOAc/heptanes) to afford two products. The less polar diastereomer was isolated as a paleyellow oil, (1222)-17-amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,8,14,16-hexaen-7-ol (E/Z mixture, enantiomer 3) (16 mg, 37 %). ESI-MS m/z cale. 544.15454, found 545.2 (M+l)⁺ ; Rétention time: 3.5 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 4: (12/2)-17-Amino-12-methyl-6,15-bis(trifluoroinethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-6,7-diol (enantiomer 3), Compound 62

EfZ mixture, enantiomer 3 enantiomer 3 ( 1272) -17-Amino-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaen-7-ol (E/Zmixture, enantiomer 3) (16 mg, 0.0272 mmol) and 10 % palladium on carbon (50 % wet, 6 mg, 0.0028 mmol) in EtOAc (3 mL) was stirred under hydrogen (balloon) at room température ovemight. The mixture was filtered through diatomaceous earth and the cake was washed with EtOAc (about 10 mL). The filtrate was concentrated and the residue was purified by silica gel chromatography (eluting 20 % to 50 % EtOAc/pentane) and freeze dried to afford (1272)-17-amino-12-methyl-6,15bis(trifluorom ethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaene-6,7-diol (enantiomer 3) (7 mg, 52 %) as a pale-yellow solid. ^!H NMR (400 MHz, DMSO-dô) δ 7.75 (s, 1H), 7.57 (s, 1H), 6.27 (s, 2H), 5.51 (d, J = 6.8 Hz, 1H), 4.88 - 4.78 (m, 1H), 4.35 - 4.26 (m, 1H), 2.63 - 2.51 (m, 2H), 1.96 - 1.83 (m, 1H), 1.75 - 1.64 (m, 1H), 1.63 1.53 (m, 1H), 1.52 - 1.43 (m, 1H), 1.37 - 1.28 (m, 5H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.43 (s, 3F), -73.86 (s, 3F). ESI-MS m/z cale. 456.12323, found 457.2 (M+l)⁺; Rétention time: 2.99 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Example 45: Préparation of (6/2,132?)-18-amino-13-methyl-6,16-bis(trifluoromethyl)-14,20dioxa-3,4,19-triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-6-ol (enantiomer 1), Compound 63, and (672,1372)-18-amino-13-methyl-6,16389 bis(trifluoromethyl)-14,20-dioxa-3,4,19-triazatetracyclo[13.3.1.12,5.09,ll]icosal(19),2,4,15,17-pentaen-6-ol (enantiomer 2), Compound 64

E/Z mixture

Step 1: tert-Butyl 7V-[(6J?,13R)-6-benzyIoxy-13-methyl-6,16-bis(trifluoromethyl)-14,205 dioxa-3,4,19-triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-18-yl]-7V-terfbutoxycarbonyl-carbamate

E/Z mixture

A flask was charged with potassium hydroxide (800 mg, 14.259 mmol) and water (3.4 mL) was added. The mixture was stirred for 2 min until KOH was dissolved. Diethyl ether (14 mL) was added. The mixture was cooled to 0 °C and 1-methyl-l-nitroso-urea (700 mg, 6.1118 mmol) was added. The mixture was stirred at 0 °C for 30 min. Stirring was stopped and the mixture was cooled to -78 °C. Once the aqueous phase was frozen, the yellow ethereal layer was transferred by a plastic pipette to another flask with two pellets (about 220 mg) of potassium hydroxide. The flask stayed at 0 °C for 5 min. Half of the formed diazomethane solution in ether (~7 mL) was added by a plastic pipette to a solution of ter/-butyl 7V-[(6Æ,12Æ)-6-benzyloxy-12methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,9,14,16-hexaen-17-yl]-7V-terZ-butoxycarbonyl-carbamate {E/Zmixture) (200 mg, 0.2536 mmol) in THF (10 mL) at 0 °C. A solution of palladium diacetate (12 mg, 0.0535

390 mmol) in THF (3.4 mL) was added dropwise. The resulting mixture was stirred at 0 °C for 15 min and then the remainder of the ethereal diazomethane solution (~7 mL) was added. The brown solution was allowed to warm to room température and stirred for 1 h. For a second time, a flask was charged with potassium hydroxide (800 mg, 14.259 mmol) and water (4 mL) was added. The mixture was stirred for 2 min until KOH was dissolved. Diethyl ether (14 mL) was added. The mixture was cooled to 0 °C and 1-methyl-1-nitroso-urea (700 mg, 6.7908 mmol) was added. The mixture was stirred at 0 °C for 30 min. Stirring was stopped and the mixture was cooled to -78 °C. Once the aqueous phase was frozen, the yellow ethereal layer was transferred to another flask with 220 mgs of potassium hydroxide. The flask stayed at 0 °C for 5 min. Half of this diazomethane solution in ether (~7 mL) was added to the previous reaction solution. A solution of palladium diacetate (12 mg, 0.0535 mmol) in THF (3 mL) was added dropwise to the reaction mixture. The resulting mixture was stirred at 0 °C for 15 min and then the remainder of the ethereal diazomethane solution (~7 mL) was added. The brown solution was allowed to warm to room température and stirred for 1 h. For a third time, a flask was charged with potassium hydroxide (800 mg, 14.259 mmol) and water (3.5 mL) was added. The mixture was stirred for 2 min until KOH was dissolved. Diethyl ether (14 mL) was added. The mixture was cooled to 0 °C and 1-methyl-1-nitroso-urea (700 mg, 6.7908 mmol) was added. The mixture was stirred at 0 °C for 30 min. Stirring was stopped and the mixture was cooled to -78 °C. Once the aqueous phase was frozen, the yellow ethereal layer was transferred to another flask with 220 mgs of potassium hydroxide. The flask stayed at 0 °C for 5 min. Half of this diazomethane solution in ether (~7 mL) was added by a plastic pipette to the previous reaction solution. A solution of palladium diacetate (12 mg, 0.0535 mmol) in THF (3 mL) was added dropwise. The resulting mixture was stirred at 0 °C for 15 min and then the remainder of the ethereal diazomethane solution (~7 mL) was added. The brown solution was allowed to warm to room température and stirred for 1 h. Nitrogen was bubbled through the black mixture for 10 min, then the solvents were removed under vacuum (at 25 °C). The residue was purified by reverse phase chromatography over a 50 g silica column (5 % to 90 % CH3CN /0.01 % formic acid in water) to give a diastereomeric mixture of Zeri-butyl TV-[(6Z?,13/?)-6-benzyloxy-13-mcthyl-6,16bis(trifluoromethyl)-14,20-dioxa-3,4,19-triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17pentaen-18-yl]-jV-/er/-butoxycarbonyl-carbamate (123 mg, 64 %) as a white solid. ESI-MS m/z cale. 742.2801, found 587.2 (M-155)⁺; Rétention time: 4.6 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

391

Step 2: (6/?,13/?)-6-Benzyloxy-13-methyl-6,16-bis(trifluoromethyl)-14,20-dioxa-3,4,19 triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-18-amine

Trifluoroacetic acid (740.00 mg, 0.5 mL, 6.4899 mmol) was added to a yellow solution of teri-butyl N-[(6R, 13/?)-6-benzyloxy-13-methyl-6,16-bis(trifluoromethyl)-14,20-dioxa-3,4,19triazatetracyclo[ 13.3.1.12,5.09,11 ]icosa-l (19),2,4,15,17-pentaen-18-yl]-7V-ier/-butoxycarbonylcarbamate (120 mg, 0.1582 mmol) in DCM (2.4 mL) at room température and the mixture was stirred for 1 h. The orange solution was concentrated under vacuum. The residual TFA was coevaporated with CH3CN (3X4 mL), CFhCN/toluene (1X4 mL) and toluene (3X4 mL) then dried under high vacuum to give crude (6/?,13A)-6-bcnzyloxy-13-methyl-6,l 6bis(trifluorom ethyl)-14,20-dioxa-3,4,19-triazatetracyclo[ 13.3.1.12,5.09,11 ]icosa-1 ( 19),2,4,15,17pentaen-18-amine (103 mg, quant.) as a light orange oil. ESI-MS m/z cale. 542.17523, found 543.2 (M+l)⁺; Rétention time: 4.35 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow = 1.5 mL/min).

Step 3: (6R,13R)-18-Amino-13-methyl-6,16-bis(trifluoromethyI)-14,20-dioxa-3,4,19triazatetracycIo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-6-ol (enantiomer 1), Compound 63, and (6R,13/î)-18-amino-13-methyl-6,Î6-bis(trifluoromethyl)-14,20-dioxa3,4,19-triazatetracycIo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-6-ol (enantiomer 2), Compound 64

Palladium on carbon (10 % w/w, 18 mg, 0.0169 mmol) was added to a degassed solution of (6R, 13 J?)-6-benzyloxy-13 -methyl-6,16-bis(trifluoromethyl)-14,20-dioxa-3,4,19triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-18-amine (90 mg, 0.1659 mmol) in methanol (2 mL) at room température. The black suspension was purged with nitrogen

392 for 5 min, then hydrogen was bubbled through the suspension for 5 min. Then, the mixture was stirred at room température ovemight under hydrogen atmosphère. The black suspension was filtered over Celite and the cake was washed with DCM (3X5 mL) then concentrated the filtrate under vacuum to give a fluorescent yellow oil. Purified by reverse phase chromatography over a 50 g Cis column (5 % to 90 % CH3CN/O.O2 % HCl in water) to give a yellow solid (57 mg, 98.6 % of purity). This solid was subjected to SFC séparation using the following conditions: Lux Cellulose 3 column, (250 X 21.2 mm), 5 pm column at 40 °C, eluant: 10 % éthanol (0.1 % diethylamine), 90 % CO₂, flow rate: 75 mL/min, concentration: 5.7 mg/mL in éthanol (0.1.% diethylamine), injection volume: 200 uL, pressure: 100 bar, wavelength: 250 nm. Evaporation of the solvents provided two isomers. Both were redissolved in EtOAc (5 mL) then washed with 0.5 M hydrogen chloride aqueous solution (1X10 mL), saturated sodium bicarbonate aqueous solution (1X10 mL) and brine (1X10 mL). The organic layer of each was dried over anhydrous sodium sulfate and then concentrated under vacuum. Both isomers were then lyophilized to afford as the first compound to elute under the SFC conditions and a light yellow solid, (6R, 13R)-18-amino-13-methyl-6,16-bis(trifluoromethyl)-14,20-dioxa-3,4,19triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-6-ol (enantiomer 1) (15.6 mg, 20 %). ‘H NMR (400 MHz, MeOD) δ 7.66 (s, 1H), 5.34 - 5.19 (m, 1H), 2.96 - 2.79 (m, 1H), 2.52 (dd, J= 15.2, 8.1 Hz, 1H), 1.96-1.81 (m, 2H), 1.45 (d, J = 6.4 Hz, 3H), 1.04-0.96 (m, 2H), 0.91 (t, J= 12.5 Hz, 1H), 0.59 - 0.44 (m, 1H), 0.36 - 0.27 (m, 1H), 0.26 - 0.17 (m, 1H). ^I9FNMR (377 MHz, MeOD) δ -65.42 (s, 3F), -80.26 (s, 3F). ESI-MS m/z cale. 452.1283, found 453.1 (M+l)⁺; Rétention time: 3.8 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

The second peak to elute under the above SFC conditions was repurified by SFC using the following conditions: Lux Cellulose 3 column, (250 X 21.2 mm), 5 pm column at 40 °C, eluant: 10 % éthanol (0.1 % diethylamine), 90 % CO2, flow rate: 75 mL/min, concentration: 21 mg/mL in éthanol (0.1.% diethylamine), injection volume: 200 pL, pressure: 100 bar, wavelength: 250 nm. Evaporation of the solvent and lyophilization provided as a light yellow solid, ($R, 13Æ)-18-amino-13 -methyl-6,16-bis(trifluoromethyl)-14,20-dioxa-3,4,19triazatetracyclo[13.3.1.12,5.09,ll]icosa-l(19),2,4,15,17-pentaen-6-ol (enantiomer 2) (9.61 mg, 13 %). Ή NMR (400 MHz, MeOD) δ 7.64 (s, 1H), 5.56 - 5.38 (m, 1H), 2.63 - 2.48 (m, 1H), 2.41 -2.17 (m, 2H), 2.09- 1.87 (m, 1H), 1.32 (d, J= 6.6 Hz, 3H), 1.24- 1.14 (m, 1H), 1.13 1.04 (m, 1H), 0.94 - 0.68 (m, 2H), 0.27 - 0.10 (m, 2H). ^,9F NMR (377 MHz, MeOD) δ -65.27 (s,

393

3F), -78.81 (s, 3F). ESI-MS m/z cale. 452.1283, found 453.1 (M+l)⁺; Rétention time: 3.79 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Example 46: Préparation of (6/?)-12-cyclopropyI-17-(ethylamino)-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (enantiomer 1), Compound 65, and (6R)-12-cyclopropyl-17-(ethylamino)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol (enantiomer 2), Compound 66

BrMg

Step 1

OH

N0₂ OH

E/Z mixture

Step 6

enantiomer 1 enantiomer 2

394

Step 1: l-Cyclopropylbut-3-en-l-ol

To a stirred solution of cyclopropanecarbaldehyde (5.0427 g, 5.6 mL, 69.068 mmol) in diethyl ether (50 mL) in a three neck 250 mL flask was added a solution of allyl(bromo)magnesium (69.5 mL of 1 M, 69.5 mmol) in diethyl ether at -20 °C. The resulting solution was stirred for 1 h from -20 °C to 0 °C. The reaction was then quenched with the addition of 25 mL aqueous saturated solution of ammonium chloride and extracted with 2 X 50 mL of diethyl ether. The combined organic extracts were washed with 1 X 50 mL of brine, dried over anhydrous sodium sulfate and concentrated under vacuum to afford as a light yellow oil, 1cyclopropylbut-3-en-l-ol (6.008 g, 74 %). Ή NMR (400 MHz, DMSO-d₆) δ 5.88 (ddt, J = 17.2, 10.2, 7.1 Hz, 1H), 5.08 - 4.94 (m, 2H), 4.54 - 4.35 (m, 1H), 2.91 (q, J = 6.5 Hz, 1H), 2.28 - 2.14 (m, 2H), 0.84 - 0.73 (m, 1H), 0.40 - 0.28 (m, 2H), 0.27 - 0.19 (m, 1H), 0.18 - 0.09 (m, 1H).

Step 2: A^r’-[(27?)-2-BenzyIoxy-2-(trifluoromethyI)hex-5-enoyl]-6-hydroxy-3-nitro-5(trifluoromethyl)pyridine-2-carbohydrazide

OH

NO₂ OH

6-Hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carboxylic acid (203 mg, 0.8052 mmol) and (2/?)-2-benzyloxy-2-(trifluoromethyl)hcx-5-enehydrazide (hydrochloride sait) (260 mg, 0.7675 mmol) were dissolved in a mixture of ethyl acetate (5 mL) and triethylamine (326.7 mg, 0.45 mL, 3.2286 mmol). Propylphosphonic anhydride (0.7 mL of 1.68 M, 1.176 mmol) solution in ethyl acetate was then added at room température (20 to 25 °C) and the reaction stirred for 3.5 hours at room température. It was then quenched with an aqueous saturated solution of ammonium chloride (5 mL). The phases were separated and the organic phase was washed with an aqueous saturated solution of ammonium chloride (5 mL) and then with an aqueous (5 % w/v) solution of sodium bicarbonate (2X5 mL), dried over anhydrous sodium sulfate, filtered and concentrated by évaporation under reduced pressure. Drying on a vacuum pump ovemight gave as a yellow solid, 7V-[(27?)-2-benzyloxy-2-(trifluoromethyl)hex-5enoyl]-6-hydroxy-3-nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (430 mg, 96 %). ESIMS m/z cale. 536.1131, found 537.2 (M+l)⁺; Rétention time: 3.04 minutes. LCMS Method:

395

Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 3: 6-[5-[(llî)-l-Benzyloxy-l-(trifIuoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5nitro-3-(trifluoromethyl)pyridin-2-ol

To a solution of7V-[(27î)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]-6-hydroxy-3nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (6.47 g, 8.4557 mmol) and N,Ndiisopropylethylamine (5.8618 g, 7.9 mL, 45.355 mmol) in acetonitrile (200 mL) at 0 °C was added 4-methylbenzenesulfonyl chloride (3.71 g, 19.460 mmol) in portions. After the addition, the cooling bath was removed and the reaction was stirred at 26 °C for 44 hours. The volatiles were removed by évaporation under reduced pressure. The residue was dissolved in ethyl acetate (180 mL) and washed with 0.5 N aqueous solution of hydrochloric acid (3 X 25 mL) and brine (2X15 mL). The organic layer was dried over anhydrous sodium sulfate, fïltered and concentrated by évaporation under reduced pressure which gave 6-[5-[(l/?)-l-benzyloxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)pyridin-2-ol (6.67 g, 70 %) as brown sticky residue. ESI-MS m/z cale. 518.1025, found 519.1 (M+l)⁺; Rétention time: 3.1 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 4: 2-[(17?)-l-Benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(l-cyclopropylbut-3 enoxy)-3-nitro-5-(trifluoromethyl)-2-pyridyI]-l,3,4-oxadiazole

To a 25 mL round bottom flask containing 6-[5-[(l/?)-l-benzyIoxy-l(trifluoromethyl)pent-4-enyl]-l,3,4-oxadiazol-2-yl]-5-nitro-3-(trifluoromethyl)pyridin-2-ol (418

396 mg, 0.7395 mmol) was added triphenylphosphine (395 mg, 1.506 mmol) and 1-cyclopropylbut3-en-l-ol (245.1 mg, 0.2 mL, 2.0758 mmol), then toluene (8.5 mL). To the mixture stirring at room température was added dropwise DIAD (308.1 mg, 0.3 mL, 1.5237 mmol) then continued stirring for 4 hours. After standing at room température for 72 hours, the mixture was concentrated by évaporation under reduced pressure. The residue was purified by reverse phase chromatography on a Cis 50 g aqueous column, eluting with a 5 % to 80 % gradient of acetonitrile in basic water (pH =10, ammonium bicarbonate) for 8 column volumes, then with 80 % acetonitrile in basic water (pH = 10, ammonium bicarbonate) over 10 column volumes. Selected fractions were concentrated by évaporation under reduced pressure to afford 2-[(17?)-lbenzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(l-cyclopropylbut-3-enoxy)-3-nitro-5(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (250 mg, 54 %) as a light yellow solid. ESI-MS m/z cale. 612.1807, found 613.2 (M+l)⁺; Rétention time: 4.08 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 5: (6/?)-6-BenzyIoxy-12-cyclopropyl-17-nitro-6,15-bis(trifluoromethyl)-13,19-dioxa 3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaene (E/Z mixture)

EfZ mixture

A dried 250-mL flask was charged with 1,2-dichloroethane (98 mL). The solvent was bubbled with nitrogen for 30 min. Zhan catalyst-lB (29 mg, 0.0395 mmol) was added under a gentle flow of nitrogen. The mixture was bubbled with nitrogen for 10 min and heated to 60 °C. A nitrogen-bubbled solution of 2-[(17?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5-[6-(lcyclopropylbut-3-enoxy)-3-nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (98 mg, 0.1578 mmol) was added dropwise over 1 h. The mixture was stirred at 70 °C for 1.25 h and then cooled to room température. Then, 5 drops of DMSO were added to quench the catalyst. The mixture was concentrated on silica gel (3 g) and purified by silica gel chromatography (eluting with a gradient of 5 % to 25 % ethyl acetate in heptanes) to afford (6R)-6-benzyloxy-12-cyclopropyl17-nitro-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo [12.3.1.12,5]nonadecal(17),2,4,9,14(18),15-hexaene (E/Zmixture) (60 mg, 59 %) as a light yellow solid. ESI-MS m/z cale. 584.1494, found 585.2 (M+l)⁺; Rétention time: 4.22 minutes. LCMS Method: XBridge Cis 397 column (4.6 X 75 mm, 5 mm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 6: (6Æ)-12-CycIopropyl-17-(ethylamino)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol

E/Z mixture

A solution of (67?)-6-benzyloxy-12-cyclopropyl-17-nitro-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,9,14(18),15-hexaene (E/Z mixture) (60 mg, 0.1027 mmol) in éthanol (4 mL) was bubbled with nitrogen gas for 5 minutes. After adding palladium on carbon (63 mg, 5 % w/w, 0.0296 mmol), hydrogen gas was bubbled in the reaction mixture for 5 minutes and then the reaction was left to stir under one atmosphère of hydrogen for about 22 hours. The reaction was purged twice with nitrogen gas then filtered over a pad of celite and the cake was washed with éthanol (30 mL). The volatiles were removed under reduced pressure to afford a 54 mg residue as an intense yellow oil. The residue was dry loaded on silica gel and purified by silica gel chromatography eluting with gradient of ethyl acetate (0 % to 30 %) in heptanes which afforded (67?)-12-cyclopropyl-17-(ethyIamino)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 17),2,4,14( 18), 15pentaen-6-ol (22 mg, 43 %) as a light yellow solid. ESI-MS m/z cale. 494.1753, found 495.2 (M+l)⁺; Rétention time: 3.8 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 7: (6/?)-I2-CyclopropyI-17-(ethylamino)-6,15-bis(trifhioromethyl)-13,19-dioxa-3,4,18triazatricycIo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (enantiomer 1), Compound 65, and (67?)-12-cyclopropyl-17-(ethylamino)-6,15-bis(trifluoromethyl)-13,19

398 dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15-pentaen-6-ol (enantiomer 2), Compound 66

enantiomer 1 enantiomer 2

A diastereomeric mixture of (67?)-12-cyclopropyl-17-(ethylamino)-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(17),2,4,14(18),15pentaen-6-ol (22 mg, 0.0438 mmol) was subjected to SFC séparation using the following conditions: Lux 5 pm Cellulose 4 column, (250 X 21.2 mm, 6.67 mg/injection) at 40 °C, eluant: 7 % EtOH (0.1 % diethylamine), 93 % CO₂, flow rate: 55 mL/min, injection volume: 400 pL, pressure: 100 bar, wavelength: 250 nm. Evaporation of the solvents and lyophilization provided two isomers:

The first isomer to elute was isolated as a yellow solid, (67î)-12-cyclopropyl-17(ethylamino)-6,l 5-bis(trifluoromethyl)-l 3,19-dioxa-3,4,l 8-triazatricyclo[l 2.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol (enantiomer 1) (3.8 mg, 17 %, 98.4 % de). Ή NMR (400 MHz, DMSO-dô) δ 7.63 (s, 1H), 7.59 (s, 1H), 6.37 (t, J = 5.5 Hz, 1H), 4.17 - 4.06 (m, 1H), 3.46 - 3.35 (m, 2H), 2.62 - 2.53 (m, 1H), 2.21 - 2.04 (m, 2H), 1.77 - 1.41 (m, 7H), 1.23 (t, J= 7.1 Hz, 3H), 1.17-1.11 (m, 1H), 0.64 - 0.55 (m, 1H), 0.52 - 0.39 (m, 2H), 0.31-0.23 (m, 1H). ^,9FNMR(377 MHz, DMSO-dô) δ -62.11 (s, 3F), -79.02 (s, 3F). ESI-MS m/z cale. 494.17526, found 495.2 (M+l)⁺; Rétention time: 3.81 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

The second peak to elute was isolated as a yellow solid, (6/?)-12-cyclopropyl-17(ethylamino)-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(17),2,4,14(18),15-pentaen-6-ol (enantiomer 2) (4.6 mg, 21 %, 99.9 % de). ’H NMR (400 MHz, DMSO-dô) δ 7.63 (s, 1H), 7.58 (s, 1H), 6.36 (t, J = 5.6 Hz, 1H), 4.26 - 4.15 (m, 1H), 3.48 - 3.33 (m, 2H), 2.48 - 2.44 (m, 1H), 2.31 - 2.22 (m, 1H), 2.17 - 2.06 (m, 1H), 1.83 - 1.61 (m, 3H), 1.57 - 1.36 (m, 4H), 1.24 (t, J = 7.1 Hz, 3H), 1.18- 1.14 (m, 1H), 0.66 - 0.58 (m, 1H), 0.52 - 0.37 (m, 2H), 0.32 - 0.23 (m, 1H). ’⁹F NMR (377 MHz, DMSO-dô) δ -62.13 (s, 3F), -76.35 (s, 3F). ESIMS m/z cale. 494.17526, found 495.2 (M+l)⁺; Rétention time: 3.8 minutes. LCMS Method:

399

Example 47: Préparation of (67?,127?)-17-amino-15-(difluorometliyI)-12-methyI-6(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,165 pentaen-6-ol, Compound 67

Step 1: Methyl 5-(difIuoromethyl)pyridine-2-carboxylate

In an autoclave was added 2-bromo-5-(difluoromethyl)pyridine (24 g, 115.38 mmol), methanol (240 mL), triethylamine (27.588 g, 38 mL, 272.64 mmol) and [Ι,Γ400 bis(diphenylphosphino)ferrocene]dichloropalladium(II) (2.4 g, 3.28 mmol). The autoclave was purged with nitrogen, then with carbon monoxide. The mixture was heated to 130 °C and the carbon monoxide pressure was adjusted to 120 psi. The mixture was stirred 3 h at 130 °C, then cooled down to 25 °C ovemight. The mixture was purged with nitrogen and concentrated under vacuum. The resulting solid was diluted with ethyl acetate (500 mL). Water (200 mL) and sodium carbonate (20 g) were added and the mixture was vigorously stirred for 10 minutes. The layers were separated and the organic layer was washed with water (200 mL) and brine (200 mL), dried over sodium sulfate, then filtered and evaporated under reduced pressure. The residue was purified by silica gel column chromatography (eluting from 20 % to 50 % of ethyl acetate in heptanes) to afford methyl 5-(difluoromethyl)pyridine-2-carboxylate (8.1 g, 38 %) as a white solid. Ή NMR (400 MHz, CDC1₃) δ 8.89 (s, 1H), 8.25 (d, J = 8.1 Hz, 1H), 8.02 (d, J = 8.1 Hz, 1H), 6.97 - 6.61 (m, 1H), 4.05 (s, 3H). ¹⁹F NMR (377 MHz, CDCI3) δ -113.58 (d, J = 54.5 Hz, 2F). ESI-MS m/z cale. 187.04448, found 188.2 (M+l)⁺; Rétention time: 1.48 minutes. LCMS Method: Kinetex Polar Ci8 column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 2: Methyl 5-(difluoromethyl)-l-oxido-pyridin-l-ium-2-carboxylate ^f’^hcY^n

Urea hydrogen peroxide (13.7 g, 145.64 mmol) was stirred into a solution of methyl 5(difluoromethyl)pyridine-2-carboxylate (8.1 g, 43.282 mmol) in DCE (70 mL). Trifluoroacetic anhydride (24.025 g, 15.9 mL, 114.39 mmol) was then added over 20 minutes at a température of-10 °C, using a cooling bath (CCh/acetone bath). The reaction mixture was then stirred for a further 30 minutes at a température of 0 °C and then for 1 hour at ambient température. The reaction mixture was poured into ice-water (150 mL) and adjusted to pH = 2 - 3 with around 150 mL of aqueous IN sodium hydroxide solution. The mixture was diluted with dichloromethane (200 mL) and then layers were separated. The aqueous phase was extracted with dichloromethane (2 X 150 mL). The combined organic phase was washed with brine (150 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to give methyl 5-(difluoromethyl)-l-oxido-pyridin-l-ium-2-carboxylate (8.39 g, 87 %) as a yellow solid. ’H NMR (400 MHz, CDCI3) δ 8.41 (s, 1H), 7.72 (d, J = 8.1 Hz, 1H), 7.43 - 7.36 (m, 1H), 6.84 - 6.47 (m, 1H), 4.03 (s, 3H). ¹⁹F NMR (377 MHz, CDCI3) δ -115.27 (d, J = 55.9 Hz, 2F). ESI-MS m/z cale. 203.0394, found 204.1 (M+l)⁺; Rétention time: 0.73 minutes. LCMS Method:

401

Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 3: Methyl 5-(difluoromethyl)-6-hydroxy-pyridine-2-carboxylate

Trifluoroacetic anhydride (84.616 g, 56 mL, 402.87 mmol) was added dropwise to a mixture of methyl 5-(difluoromethyl)-l-oxido-pyridin-l-ium-2-carboxylate (11.63 g, 47.06 mmol) in DMF (130 mL) at a température of 0 °C for 30 minutes. The mixture was then heated at 48 °C and was stirred for a further 4 hours then the reaction was stirred at room température ovemight. The reaction mixture was concentrated under reduced pressure to remove trifluoroacetic anhydride. The residual DMF solution was poured into water (1 L) at 0 °C. The precipitated solid was collected by filtration and washed with water (200 mL). The solid was dried under high vacuum which gave as an off-white solid, methyl 5-(difluoromethyl)-6hydroxy-pyridine-2-carboxylate (5.74 g, 60 %). ¹II NMR (400 MHz, DMSO-de) δ 12.29 (br. s., 1H), 7.88 (d, J = 7.3 Hz, 1H), 7.13 (s, 1H), 7.07 - 6.76 (m, 1H), 3.87 (s, 3H). ¹⁹F NMR (377 MHz, DMSO-dô) δ -118.60 (br. s., 2F). ESI-MS m/z cale. 203.0394, found 204.1 (M+l)⁺; Rétention time: 1.34 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 4: Methyl 5-(difluoromethyl)-6-hydroxy-3-nitro-pyridine-2-carboxylate

To an ice-cooled solution of methyl 5-(difluoromethyl)-6-hydroxy-pyridine-2carboxylate (7.43 g, 36.575 mmol) in sulfuric acid (48 mL of 18.4 M, 883.2 mmol) was added nitric acid (2.5 mL of 15.8 M, 39.5 mmol) dropwise. After 5 min, the ice bath was removed and the reaction mixture was stirred at 45 °C ovemight. The reaction was precipitated on ice-water (300 mL). The solution was cooled at 0 °C for 15 minutes then the solid was isolated by filtration and washed with water (200 mL). The solid was dried ovemight under high vacuum to give methyl 5-(difluoromethyl)-6-hydroxy-3-nitro-pyridine-2-carboxylate (5.47 g, 56 %) as an off-white solid. ’H NMR (400 MHz, DMSO-d₆) δ 8.41 (s, 1H), 7.19 - 6.75 (m, 1H),

402

3.94 (s, 3H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -118.87 (d, J = 54.5 Hz, 2F). ESI-MS m/z cale.

248.02448, found 249.1 (M+l)⁺; Rétention time: 1.6 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 5: Methyl 5-(difluoromethyl)-6-[(lR)-l-methylbut-3-enoxy]-3-nitro-pyridine-2carboxylate

To a solution of methyl 5-(difluoromethyl)-6-hydroxy-3-nitro-pyridine-2-carboxylate (1.6 g, 6.448 mmol) and (2>S)-pent-4-en-2-ol (837 mg, 1 mL, 9.7176 mmol) in toluene (31 mL) was added triphenyl phosphine (2.5 g, 9.5316 mmol). After stirring at room température for 10 minutes, DIAD (1.9513 g, 1.9 mL, 9.65 mmol) was added and the mixture was stirred at room température for 16 h. About half of the toluene was evaporated under reduced pressure and the crude was directly loaded onto a 120 g silica cartridge. Silica gel chromatography purification was performed using a gradient of 0 % to 5 % EtOAc in heptanes. Evaporation of the volatiles from the fractions containing the product afforded methyl 5-(difluoromcthyl)-6-[(l/?)-lmethylbut-3-enoxy]-3-nitro-pyridine-2-carboxylate (1.7 g, 82 %) as a light yellow oil. 'H NMR (400 MHz, CDCh) δ 8.61 (s, 1H), 6.80 (t, J = 54.5 Hz, 1H), 5.86 - 5.72 (m, 1H), 5.51 (m, J = 6.2 Hz, 1H), 5.20 - 5.07 (m, 2H), 4.03 (s, 3H), 2.58 - 2.42 (m, 2H), 1.41 (d, J= 6.4 Hz, 3H). ¹⁹F NMR (377 MHz, CDCh) δ -118.39 (s, 2F). Rétention time: 2.1 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 6: Methyl 3-amino-5-(difluoromethyl)-6-[(17?)-l-methylbut-3-enoxy]pyridine-2carboxylate

NO₂ nh₂

Iron (2.4 g, 42.976 mmol) and ammonium chloride (300 mg, 5.6084 mmol) were added to a solution of methyl 5-(difluoromethyl)-6-[(l/?)-l-mcthylbut-3-enoxy]-3-nitro-pyridine-2carboxylate (1.4 g, 4.3471 mmol) in éthanol (40 mL) and water (12 mL) at room température. The mixture was heated to 80 °C for 16 hours, then cooled to room température. The reaction 403 was filtered over Celite, washing the cake with EtOH (100 mL). The volatiles were removed under reduced pressure. The crude residue was purified by reverse phase chromatography on a 120 g Cis cartridge using a 40 % to 100 % gradient of CH3CN in acidic water (0.1 % v/v of formic acid in water). The fractions containing the product were concentrated under reduced pressure until only water was left as solvent. The remaining aqueous phase was extracted with EtOAc (4 X 30 mL). The combined organic layers were washed with brine, dried over sodium sulfate, filtered and concentrated under reduced pressure which afforded methyl 3amino-5-(difluoromethyl)-6-[(17?)-l-methylbut-3-enoxy]pyridine-2-carboxylate (727 mg, 58 %) as a brown oil. ESI-MS m/z cale. 286.11288, found 287.2 (M+l)⁺; Rétention time: 2.05 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 7: Methyl 3-[bis(ter/-butoxycarbonyl)amino]-5-(difluoromethyI)-6-[(llî)-l-methyIbut3-enoxy]pyridine-2-carboxylate

To a solution of methyl 3-amino-5-(difluoromethyl)-6-[(17?)-l-methylbut-3enoxy]pyridine-2-carboxylate (100 mg, 0.3409 mmol) in DCM (3 mL) was added (Boc)zO (298 mg, 1.3654 mmol) followed by DMAP (4 mg, 0.0327 mmol). The reaction mixture was stirred at room température for 20 h and the volatiles were removed under reduced pressure. The crude residue was purified by flash-chromatography on a 24 g silica gel cartridge, using a gradient of 0 % to 10 % ethyl acetate in heptanes which afforded methyl 3-[bis(ier/-butoxycarbonyl)amino]-5(difluoromethyl)-6-[(lÆ)-l-methylbut-3-enoxy]pyridine-2-carboxylate (64 mg, 25 %) as a light yellow oil. ESI-MS m/z cale. 486.21774, found 387.2 (M-99)⁺; Rétention time: 2.29 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

404

Step 8: 3-(tert-ButoxycarbonyIamino)-5-(difluoromethyl)-6-[(llî)-l-methylbut-3enoxy]pyridine-2-carboxylic acid

To a solution of methyl 3-[bis(terrtbutoxycarbonyl)amino]-5-(difluoromethyl)-6-[(17?)-lmethylbut-3-enoxy]pyridine-2-carboxylate (800 mg, 1.6444 mmol) in MeOH (6 mL) and THF (6 mL) was added a solution of lithium hydroxide monohydrate (300 mg, 7.1491 mmol) in water (3 mL). The mixture was stirred at 50 °C for 16 h. The reaction mixture was then cooled down to room température and 5 mL of aqueous 1 N HCl was added. The volatiles were removed under reduced pressure and water (5 mL) was added. The pH was adjusted to 2 using aqueous 1 N HCl and the product was extracted with EtOAc (3X15 mL). The combined organic phases were washed with brine (15 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure which afforded 3-(ZerZ-butoxycarbonylamino)-5-(difluoromethyl)-6-[(lJ?)-lmethylbut-3-enoxy]pyridine-2-carboxylic acid (650 mg, 98 %) as a yellow oil. Rétention time: 2.23 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 9: tert-Butyl 7V-[2-[[[(21î)-2-benzyloxy-2-(trifluoromethyl)hex-5enoyl]aimno]carbamoyl]-5-(difluoromethyI)-6-[(U?)-l-methyIbut-3-enoxy]-3pyridyl] carbamate

HATU (790 mg, 2.0777 mmol) was added to an orange solution of ?>-(tertbutoxycarbonylamino)-5-(difluoromethyl)-6-[(12?)-l-methylbut-3-enoxy]pyridine-2-carboxylic acid (650 mg, 1.7334 mmol), (2Æ)-2-benzyloxy-2-(trifluoromethyl)hex-5-enehydrazide (hydrochloride sait) (705 mg, 2.0812 mmol) and DIPEA (1.1872 g, 1.6 mL, 9.1858 mmol) in DMF (11 mL) at room température. The solution was stirred at room température for 2

405

h. The reaction was directly loaded on a 275 g Cis cartridge and the purification was run eluting with a gradient of 50 % to 100 % CH3CN in acidic water (0.1 % v/v of formic acid in water). The fractions containing the product were concentrated under reduced pressure, coevaporated with a 1:1 mixture of CH₃CN/water and lyophilized which afforded ZerZ-butyl 7V-[2[[[(22?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]amino]carbamoyl]-5-(difluoromethyl)-6[(lR)-l-methylbut-3-enoxy]-3-pyridyl]carbamate (690 mg, 61 %) as a white solid. ’H NMR (400 MHz, DMSO-dô) δ 10.74 (s, 1H), 10.66 (s, 1H), 10.30 (s, 1H), 8.91 (s, 1H), 7.55 - 7.48 (m, 2H), 7.42 - 7.35 (m, 2H), 7.35 - 7.30 (m, 1H), 7.07 (t, J = 54.5 Hz, 1H), 6.03 - 5.79 (m, 2H), 5.79 5.67 (m, 1H), 5.15 - 5.06 (m, 2H), 5.05 - 4.99 (m, 2H), 4.91 - 4.77 (m, 2H), 2.39 (t, J = 6.5 Hz, 2H), 2.36 - 2.27 (m, 2H), 2.26 - 2.14 (m, 2H), 1.47 (s, 9H), 1.26 (d, J = 6.4 Hz, 3H). ¹⁹F NMR (377 MHz, DMSO-dô) δ -71.86 (s, 3F), -117.49 (d, J = 54.5 Hz, 2F). ESI-MS m/z cale. 656.2633, found 557.3 (M+l)⁺; Rétention time: 5.36 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water +0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Step 10: ZerZ-Butyl 2V-[2-[5-[(LR)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yI]-5-(difluoromethyI)-6-[(17?)-l-methylbut-3-enoxy]-3-pyridyI]carbamate

To a solution of ZerZ-butyl 7V-[2-[[[(27?)-2-benzyloxy-2-(trifluoromethyl)hex-5enoyl]amino]carbamoyl]-5-(difluoromethyl)-6-[(lR)-l-methylbut-3-enoxy]-3-pyridyl]carbamate (490 mg, 0.7455 mmol) in 1,2-dichloroethane (15 mL) and TVJV-diisopropylethylamine (742 mg, 1 mL, 5.7411 mmol) was added toluenesulfonyl chloride (430 mg, 2.2555 mmol). The reaction was stirred at 50 °C for 20 h then cooled down to room température. The volatiles were removed under reduced pressure and the crude residue was purified by reverse phase chromatography on a 50 g Cis cartridge using a gradient of 50 % to 100 % CH3CN in acidic water (0.1 % v/v of formic acid in water). The fractions containing the product were concentrated under reduced pressure which afforded ZerZ-butyl 7V-[2-[5-[(lR)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]l,3,4-oxadiazol-2-yl]-5-(difluoromethyl)-6-[(17?)-l-methylbut-3-enoxy]-3-pyridyl]carbamate (351 mg, 73 %) as a tan oil. ESI-MS m/z cale. 638.25275, found 639.3 (M+l)⁺; Rétention time: 406

4.82 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow = 1.5 mL/min).

Step 11 : terf-Butyl A^r-[(6/?,12Z?)-6-benzyloxy-15-(difluoromethyl)-12-methyl-6(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16hexaen-17-yl]carbamate (E/Z mixture)

E/Z mixture

A solution of /er/-butyl 7V-[2-[5-[(12?)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-l,3,4oxadiazol-2-yl]-5-(difluoromethyl)-6-[(17?)-l -methylbut-3-enoxy]-3-pyridyl]carbamate (350 mg, 0.5442 mmol) in dichloroethane (180 mL) was bubbled with nitrogen gas for 1.5 h. The solution was then placed in an oil bath set at 60 °C and a first portion of Zhan catalyst-lB (28 mg, 0.0382 mmol) was added. After 1 h, a second lot of Zhan catalyst-lB (20 mg, 0.0273 mmol) was added and heating was continued for another 1 h. Once cooled to room température, the reaction was quenched with DMSO (8 drops), the volatiles were removed under reduced pressure and the residue was directly purified by reverse phase chromatography on a 50 g Cis cartridge, eluting with a gradient of 65 % to 100 % CH3CN in acidic water (0.1 % v/v of formic acid in water). The fractions containing the product were concentrated under reduced pressure which afforded icr/-butyl N-[(6R, 127?)-6-benzyloxy-15-(difluoromethyl)-12-methyl-6(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16hexaen-17-yl]carbamate (E/Z mixture) (87 mg, 26 %) as a brown oil. ESI-MS m/z cale. 610.22144, found 611.3 (M+l)⁺; Rétention time: 4.55 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

407

Step 12: ZerZ-Butyl A-[(61?,127?)-15-(difluoromethyl)-6-hydroxy-12-methyl-6(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl] carbamate

EfZ. mixture

To palladium on carbon (85 mg, 5 % w/w, 0.0399 mmol) under nitrogen was added a solution of ZerZ-butyl 7V-[(67?,12Z?)-6-benzyloxy-15-(difluoromethyl)-12-methyl-6(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16hexaen-17-yl]carbamate (E/Z mixture) (85 mg, 0.1357 mmol) in THF (10 mL) acidified with a drop of acetic acid (1.056 mg, 1 pL, 0.0176 mmol). Hydrogen was bubbled in for 1 min and the reaction was stirred at room température for 16 h. Nitrogen was bubbled in for 5 min and the reaction mixture was filtered over Celite, washing the cake with CH3CN (40 mL). The filtrate was filtered again and the resulting filtrate was concentrated under reduced pressure which afforded ZerZ-butyl N-[(67?, 127?)-15-(difluoromethyl)-6-hydroxy-12-methyl-6-(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-17yl]carbamate (77 mg, 99 %) as a tan solid. ESI-MS m/z cale. 522.1902, found 467.2 (M-56)⁺; Rétention time: 2.37 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 13: (67?,127?)-17-Amino-15-(difIuoromethyI)-12-methyl-6-(trifluoromethyI)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 67

To a solution of ZerZ-butyl 7V-[(67î,122î)-15-(difluoromethyl)-6-hydroxy-12-methyl-6(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (75 mg, 0.1397 mmol) in DCM (10 mL) was added TFA (2.96 g, 2 mL,

408

25.96 mmol). The reaction mixture was stirred at room température for 1 h. The volatiles were removed under reduced pressure and the crude material was co-evaporated twice with toluene (10 mL each time). The resulting oil was purified by reverse phase chromatography on a 15.5 g Cis cartridge, eluting from 5 % to 100 % acetonitrile in water (pH = 7). The fractions containing the product were concentrated under reduced pressure and lyophilized which afforded (6R,12R)17-amino-l 5-(difluoromethyl)-l 2-methyl-6-(trifluoromethyl)-l 3,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (39 mg, 65 %) as a yellow solid. 'H NMR (400 MHz, DMSO-d₆) δ 7.60 (s, 1H), 7.56 (s, 1H), 7.01 (t, J= 54.5 Hz, 1H), 6.29 (s, 2H), 4.81 - 4.70 (m, 1H), 2.48 - 2.42 (m, 1H), 2.35 - 2.23 (m, 1H), 2.16 - 2.05 (m, 1H), 1.73 (br. s„ 2H), 1.60 - 1.39 (m, 4H), 1.35 (d, J = 6A Hz, 3H), 1.22 - 1.11 (m, 1H). ¹⁹F NMR (377 MHz, DMSO-dô) δ -76.38 (s, 3F), -115.10 to -116.53 (m, 1F), -116.69 to -118.00 (m, 1F). ESI-MS m/z cale. 422.13773, found 423.1 (M+l)⁺; Rétention time: 3.22 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Example 48: Préparation of (67î)-17-amino-12-cyclopropyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 68, and (6_JR)-17-amino-12-cyclopropyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 69

no₂ n-n E/Z mixture ^CF3\pi^-_N X Step 2 ^cf3\< Tï°r\^CF3 ----- NH₂ N-n OH

. Step 1 (çCF₃ ---------- o-^ NH₂ N-N OH nh₂ n-n OH

409

Step 1: (67?)-17-Amino-12-cyclopropyl-6,15-bis(trifluoromethyl)-I3,19-dioxa-3,4,18 triazatricycIo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol

EZZ mixture

A solution of (67?)-6-benzyloxy-12-cyclopropyl-17-nitro-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(17),2,4,9,14( 18), 15-hexaene (E/Z mixture) (48 mg, 0.0812 mmol) in tetrahydrofuran (3 mL) was purged with nitrogen gas for 5 minutes, then added palladium on carbon (53 mg, 5 % w/w, 0.0249 mmol) and hydrogen gas was bubbled in for 5 minutes. The reaction was left to stir under one atmosphère of hydrogen for about 22 hours. The reaction was purged with nitrogen gas then filtered over a pad of Celite and the cake was washed with ethyl acetate (30 mL). The volatiles were removed under reduced pressure. The residue was solubilized in tetrahydrofuran (3 mL) and the resulting solution was purged with nitrogen gas for 5 minutes. Added palladium on carbon (53 mg, 5 % w/w, 0.0249 mmol) then hydrogen gas was bubbled in for 5 minutes. The reaction was left to stir under one atmosphère of hydrogen for another 22 hours. The reaction was purged again with nitrogen gas, then added more palladium on carbon (25 mg, 5 % w/w, 0.0117 mmol) and hydrogen gas was bubbled in for 5 minutes. The reaction was left to stir under one atmosphère of hydrogen for 4 more hours. The reaction was purged with nitrogen gas then filtered over a pad of celite and the cake was washed with ethyl acetate (30 mL). The volatiles were removed under reduced pressure and the resulted yellow oil residue was dry loaded on silica gel and purified by silica gel chromatography eluting with a 0 % to 30 % gradient of ethyl acetate in heptanes which affbrded (67?)-17-amino-12cyclopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-6-ol (24 mg, 63 %) as a light yellow solid. ESI-MS m/z cale. 466.14395, found 467.2 (M+l)⁺; Rétention time: 3.48 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 2: (6R)-17-Amino-12-cyclopropyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 1), Compound 68, and (6/?)-17-amino-12-cycIopropyl-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18410 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (enantiomer 2), Compound 69

enantiomer 1 enantiomer 2

A diastereomeric mixture of (67?)-17-amino-12-cyclopropyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (24 mg, 0.0514 mmol) was subjected to SFC séparation using the following conditions: Lux 5 pm Cellulose 4 column, (250 X 21.2 mm, 4.8 mg/injection) at 40 °C, eluant: 7 % reagent alcohol (+ 0.1 % diethylamine)/ 93 % CO2, flow rate: 75 mL/min, injection volume: 400 uL, pressure: 100 bar, wavelength: 250 nm. Evaporation of the solvents and lyophilization provided two isomers.

The first isomer to elute under the above SFC conditions was further purified using reverse phase chromatography eluting with 70 % acetonitrile in acidic water (containing 0.1 % v/v of formic acid) which gave as a yellow solid, (67î)-17-amino-12-cyclopropyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16pentaen-6-ol (enantiomer 1) (7.1 mg, 29 %, 98.6 % de). ¹H NMR (400 MHz, DMSO-dô) δ 7.75 (s, 1H), 7.58 (s, 1H), 6.34 (s, 2H), 4.15 - 4.07 (m, 1H), 2.61 - 2.53 (m, 1H), 2.22 - 2.03 (m, 2H), 1.80 - 1.37 (m, 7H), 1.18 -1.08 (m, 1H), 0.63 - 0.55 (m, 1H), 0.52 - 0.38 (m, 2H), 0.31 - 0.22 (m, 1H). ^,9F NMR (377 MHz, DMSO-d₆) δ -62.51 (s, 3F), -79.02 (s, 3F). ESI-MS m/z cale. 466.14395, found 467.1 (M+l)⁺; Rétention time: 4.9 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

The second peak to elute gave SFC peak 2 followed by reverse phase chromatography eluting with 70 % acetonitrile in acidic water (containing 0.1 % v/v of formic acid) gave as a light yellow solid (67?)-17-amino-12-cyclopropyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16-pentaen-6-ol (enantiomer 2) (5.36 mg, 22 %, 97.7 % de). Ή NMR (400 MHz, DMSO-d₆) δ 7.75 (s, 1H), 7.57 (s, 1H), 6.33 (s, 2H), 4.24 - 4.14 (m, 1H), 2.48 - 2.42 (m, 1H), 2.31 - 2.21 (m, 1H), 2.17 - 2.07 (m, 1H), 1.82 - 1.62 (m, 3H), 1.57- 1.38 (m, 4H), 1.19 - 1.08 (m, 1H), 0.67 - 0.56 (m, 1H), 0.52

411

- 0.36 (m, 2H), 0.32 - 0.22 (m, 1H). ^I9F NMR (377 MHz, DMSO-d₆) δ -62.53 (s, 3F), -76.34 (s, 3F). ESI-MS m/z cale. 466.14395, found 467.1 (M+l)⁺; Rétention time: 4.85 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Example 49: Préparation of (12R)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-8,13,19trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 1), Compound 70, and (12R)-17-aimno-12-methyl-6,15-bis(trifluoromethyI)10 8,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 2), Compound 71

diastereomer 1 diastereomer 2

412

Step 1: Benzyl 2-(trifIuoromethyl)oxirane-2-carboxylate

Benzyl 2-(trifluoromethyl)prop-2-enoate (50.45 g, 214.79 mmol) was dissolved in a mixture of dioxane (1000 mL) and water (200 mL). The mixture was cooled in an ice-water bath. With vigorous stirring, NaHCCh (91.2 g, 1.0856 mol) was added, followed by portion-wise addition of oxone (135.5g). The addition took 80 minutes then the mixture was stirred at the same température for 20 min before 90 minutes of stirring at room température. Water (300 mL) and ethyl acetate (300 mL) were added and the layers were separated. The organic layer was washed with brine, dried over anhydrous MgSCU, filtered and concentrated. The residue was purified by silica gel chromatography using a gradient from 5 % to 50 % ethyl acetate in hexanes to afford the product as colorless oil, benzyl 2-(trifluoromethyl)oxirane-2-carboxylate (57.49 g, 98 %). 'H NMR (500 MHz, Chloroform-d) δ 7.62 - 7.28 (m, 5H), 5.33 (d, J = 12.3 Hz, 1H), 5.28 (d, J = 12.2 Hz, 1H), 3.30 - 3.18 (m, 2H).

Step 2: Benzyl 3,3j3-trifluoro-2-hydroxy-2-[[(4S)-4-[(4methoxyphenyl)methoxy]pentoxy] methyl] propanoate

To a solution of (45)-4-[(4-methoxyphenyl)methoxy]pentan-l-ol (1.48 g, 6.5918 mmol) in EtOAc (6.5 mL) was added benzyl 2-(trifluoromethyl)oxirane-2-carboxylate (2.2 g, 8.9364 mmol) then magnésium triflate (2.13 g, 6.6058 mmol) and the mixture stirred at 85 °C for 24 h. To the reaction was added EtOAc (60 mL) and water (20 mL) and then extracted the mixture with EtOAc (2 X 50 mL), washed the organic fractions with brine (25 mL), dried over NazSOzi, filtered and concentrated under vacuum. The residue was purified by silica gel flash chromatography (loaded on silica gel with benzene and eluted with 0 % to 15 % EtOAc in hexanes over a 45 min gradient) to provide as a colorless oil, benzyl 3,3,3-trifluoro-2-hydroxy-2[[(4S)-4-[(4-methoxyphenyl)methoxy]pentoxy]methyl]propanoate (1.1 g, 35 %). ESI-MS m/z cale. 470.1916, found 471.1 (M+l)⁺; Rétention time: 6.5 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 413

100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 3: Benzyl 2-benzyloxy-3,3,3-trifluoro-2-[[(45)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanoate

A solution of benzyl 3,3,3-trifluoro-2-hydroxy-2-[[(45)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanoate (1.1 g, 2.3380 mmol) in DMF (5.5 mL) was cooled to 0 °C and then added benzyl bromide (720 mg, 0.5 mL, 4.21 mmol) and tetrabutyl ammonium iodide (171 mg, 0.463 mmol) followed by sodium hydride (125 mg, 3.1253 mmol) in one portion and the resulting mixture was stirred for 4 h at 0 °C. Warmed the mixture to room température and stirred for 12 h. Added more sodium hydride (38 mg, 0.9501 mmol) and benzyl bromide (288 mg, 0.2 mL, 1.6839 mmol) at 0 °C then stirred at room température for 30 minutes. The reaction was quenched with NH4CI (30 mL) at room température and extracted with EtOAc (3 X 75 mL). The combined organic phases were then washed with brine (1 X 100 mL), dried over Na2SÛ4, filtered and concentrated under vacuum. The residue was purified by silica gel flash chromatography (eluted with a gradient from 0 % to 10 % EtOAc in hexanes over 50 min) to provide as a colorless oil, benzyl 2-benzyloxy-3,3,3-trifluoro-2-[[(45)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanoate (1 g, 76 %). ESI-MS m/z cale. 560.2386, found 561.2 (M+l)⁺; Rétention time: 7.88 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 4: 2-BenzyIoxy-3,3,3-trifluoro-2-[[(4S)-4-[(4 methoxyphenyl)methoxy]pentoxy]methyl]propanehydrazide

To a solution of benzyl 2-benzyloxy-3,3,3-trifluoro-2-[[(4S)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanoate (1.64 g, 2.9254 mmol) in methanol (1 mL) 414 was added 3,4,6,7,8,9-hexahydro-277-pyrimido[l,2-a]pyrimidine (164 mg, 1.1782 mmol), then hydrazine hydrate (193.99 mg, 0.19 mL, 3.8751 mmol) and stirred at room température for 10 min. The reaction was stirred for 14 h at room température then additional hydrazine hydrate (204.20 mg, 0.2 mL, 4.0791 mmol) was added and the mixture was stirred for 30 minutes. The reaction was then quenched with water (30 mL), extracted with TBME (3 X 40 mL) and the combined organic layers were washed with saturated aqueous NaHCCh (50 mL), brine (100 mL) dried over MgSO₄, filtered and concentrated under vacuum. The residue was purified by silica gel flash chromatography (loaded with benzene/DCM and eluted with a gradient from 0 % to 10 % MeOH in DCM over 40 min) to provide clean product and mixed fractions containing product. The mixed fractions were concentrated and repurified by silica gel flash chromatography (loaded with benzene and eluted with a gradient from 0 % to 5 % methanol in dichloromethane over 40 min) and combined with the clean product from the first column to provide as an amber oil, 2-benzyloxy-3,3,3-trifluoro-2-[[(45)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanehydrazide (500 mg, 35 %). ESI-MS m/z cale. 484.2185, found 485.5 (M+l)⁺; Rétention time: 5.52 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 5: terf-Butyl A-[2-[[[2-benzyloxy-3,3,3-trifluoro-2-[[(4S)-4-[(4methoxyphenyl)methoxy] pentoxy] methyl] propan oyl] amino] carbamoyl]-6-bromo-5 (trifluoromethyl)-3-pyridyl]carbamate

To a solution of 6-bromo-3-(ter/-butoxycarbonylamino)-5-(trifluoromethyl)pyridine-2carboxylic acid (416 mg, 1.0801 mmol) and 2-benzyloxy-3,3,3-trifluoro-2-[[(4₁S)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanehydrazide (520 mg, 1.0733 mmol) in EtOAc (5.2 mL) was added T3P (1.13 g, 50 % w/w, 1.7757 mmol) then pyridine (391.2 mg, 0.4 mL, 4.9456 mmol) at room température and the reaction was stirred for 5 h. The reaction was diluted with EtOAc (20 mL), washed with saturated aqueous NH₄C1 (5 mL), then NaHCO3 (20 mL) and the aqueous layers were back-extracted with EtOAc (2 X 40 mL). The combined organic layers were washed with brine, dried over Na2SO₄, filtered and concentrated under vacuum. The residue was purified by silica gel flash chromatography (eluted with a gradient from 0 % to 20 % 415

EtOAc in hexanes over 40 min) to provide, as a pale yellow oil, terf-butyl 7V-[2-[[[2-benzyloxy3,3,3-trifluoro-2-[[(4S)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (586.2 mg, 64 %). ESI-MS m/z cale. 850.2012, found 851.6 (M+l)⁺; Rétention time: 8.46 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Ci8 column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 6: tert-Butyl 7V-[2-[5-[l-benzyIoxy-2,2,2-trifluoro-l-[[(45)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate

To a solution of ter/-butyl 7V-[2-[[[2-benzyloxy-3,3,3-trifluoro-2-[[(4.S)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]propanoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (586 mg, 0.6881 mmol) in acetonitrile (7.6 mL) was added tosyl chloride (152 mg, 0.7973 mmol) then DIPEA (267.12 mg, 0.36 mL, 2.0668 mmol) and the reaction mixture was stirred for 5 h at room température. The reaction was diluted with EtOAc (40 mL), washed with saturated aqueous NH4CI (15 mL) and brine (30 mL) then dried over Na2SO4, filtered and concentrated under vacuum. The residue was purified by silica gel flash chromatography (loaded with benzene and eluted with 0 % to 10 % EtOAc in hexanes over a 30 min gradient) to provide as a colorless oil, teri-butyl 7V-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l[[(4S)-4-[(4-methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (425 mg, 73 %). ESI-MS m/z cale. 832.1906, found 833.5 (M+l)⁺; Rétention time: 9.08 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 %

CF3CO2H).

416

Step 7: tert-Butyl 7V-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(4S)-4-[(4methoxyphenyI)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazoI-2-yI]-6-bromo-5(trifluoromethyl)-3-pyridyl]-7V-ter/-butoxycarbonyl-carbamate

To a solution of /er/-butyl 77-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(4S)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (425 mg, 0.5098 mmol) in TBME (4.35 mL) was added DIPEA (161.76 mg, 0.218 mL, 1.2516 mmol) and DMAP (22 mg, 0.1801 mmol). Di-teri-butyl dicarbonate (351.5 mg, 0.37 mL, 1.6106 mmol) was then added and the reaction stirred for 12 h at room température. The reaction was added to water (10 mL) and then the organics were extracted with TBME (50 mL). The organic solution was washed with water (50 mL), brine (50 mL), dried over Na2SO4, filtered and concentrated under vacuum. The residue was purified by silica gel flash chromatography (loaded with benzene and eluted with 0 % to 10 % EtOAc in hexanes over a 30 min gradient) to provide as a colorless oil, Zeri-butyl 7V-[2-[5-[l-benzyloxy2,2,2-trifluoro-l-[[(45)-4-[(4-methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-/eri-butoxycarbonyl-carbamate (476 mg, 99 %). ESI-MS m/z cale. 932.2431, found 933.7 (M+l)⁺; Rétention time: 8.76 minutes. LCMS Method:

Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H).

Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 8: tert-Butyl 7V-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(45)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5(trifluoromethyl)-3-pyridyl]-7V-ter/-butoxycarbonyl-carbamate

To a solution of ie/7-butyl 7V-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(45)-4-[(4methoxyphenyl)methoxy] pentoxy] methyl] ethyl] -1,3,4-oxadiazol-2-yl] -6-bromo-5

417 (trifluoromethyl)-3-pyridyl]-7V-ferZ-butoxycarbonyl-carbamate (476 mg, 0.5098 mmol) in DMSO (4.75 mL) was added césium acetate (395 mg, 2.0578 mmol) and placed in a 86 °C reaction plate for 6 h. The reaction was cooled to room température and diluted with TB ME (20 mL) and aqueous NH4CI (15 mL) and extracted with TBME (5 X 20 mL). The combined organic layers were washed with brine (2X15 mL), dried over NazSCU, filtered and concentrated under vacuum to provide as a colorless oil, /er/-butyl 7V-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(45)-4[(4-methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5(trifluoromethyl)-3-pyridyl]-7V-ier/-butoxycarbonyl-carbamate (444 mg, 95 %). ESI-MS m/z cale. 870.3275, found 871.8 (M+l)⁺; Rétention time: 7.85 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 9: tert-Butyl A-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(4S)-4hydroxypentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3pyridyl]-7V-te/7-butoxycarbonyl-carbamate

To a solution of teri-butyl JV’-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(4S)-4-[(4methoxyphenyl)methoxy]pentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5(trifluoromethyl)-3-pyridyl]-7V-terZ-butoxycarbonyl-carbamate (440 mg, 0.5053 mmol) in DCM (3.9 mL) was added DDQ (128 mg, 0.5639 mmol) then water (0.22 mL). The colorless reaction solution instantly became a grey suspension upon addition of DDQ. The mixture was stirred 1.5 h at 30 °C then diluted with DCM (20 mL), washed with saturated aqueous NaHCCh (15 mL), dried over MgSCM, filtered and concentrated under vacuum (400 mg crude obtained). The residue was purified by silica gel flash chromatography (loaded with benzene and eluted with 0 % to 50 % EtOAc in hexanes over a 30 min gradient) to provide ter/-butyl 7V-[2-[5-[l-benzyloxy2,2,2-trifluoro-l-[[(45)-4-hydroxypentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5(trifluoromethyl)-3-pyridyl]-7V-ter/-butoxycarbonyl-carbamate (310 mg, 81 %). ESI-MS m/z cale. 750.2699, found 651.3 (M-100+H)⁺; Rétention time: 6.95 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 418

Step 10: ZerZ-Butyl 7V-[(12/î)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-8,13,19trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-A-ZerZbutoxycarbonyl-carbamate

To a solution of ZerZ-butyl 7V-[2-[5-[l-benzyloxy-2,2,2-trifluoro-l-[[(45)-4hydroxypentoxy]methyl]ethyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]7V-ZerZ-butoxycarbonyl-carbamate (274 mg, 0.3650 mmol) in toluene (60 mL) was added triphenylphosphine (196 mg, 0.7473 mmol) followed by DIAD (146.16 mg, 0.14 mL, 0.7228 mmol) at room température. After 6 h, the reaction was diluted with EtOAc (30 mL) and washed with saturated aqueous NaHCÛ3 (20 mL), saturated aqueous NH4CI (20 mL) and brine (2 X 50 mL) then dried over Na2SÜ4, filtered and concentrated under vacuum (650 mg crude obtained). The residue was purified by silica gel flash chromatography (loaded with minimal benzene and eluted with 0 % to 5 % EtOAc in hexanes over an 80 minute gradient, then 5 % to 10 % EtOAc in hexanes over a 10 minute gradient) to provide both diastereomers of ZerZ-butyl 7V-[(122î)-6benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-8,13,19-trioxa-3,4,18triazatricyclof 12.3.1.12,5]nonadeca-l (18),2,4,14,16-pentaen-l 7-yl]-7V-ZerZ-butoxycarbonylcarbamate (160 mg, 59 %). ESI-MS m/z cale. 732.2594, found 733.6 (M+l)⁺; Rétention time: 8.57 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 11: ZerZ-Butyl A^r-ZerZ-butoxycarbonyl-A-[(12/?)-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-8,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate

419

To a solution of ZerZ-butyl A-[(12/?)-6-bcnzyloxy-12-methyl-6,15-bis(trifluoromethyl)8,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-7V-ZerZbutoxycarbonyl-carbamate (40 mg, 0.0546 mmol) in éthanol (1.36 mL) was added 10 % palladium on carbon (22 mg, 0.0207 mmol) under nitrogen atmosphère, then the reaction atmosphère was exchanged for hydrogen gas. The reaction was stirred at room température under a balloon of hydrogen for 45 minutes. The reaction was filtered through packed celite and the filtrate was concentrated under vacuum to provide as a yellow foam, ZerZ-butyl N-tertbutoxycarbonyl-7V-[( 127?)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-8,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (17 mg, 48 %). ESI-MS m/z cale. 642.2124, found 543.1 (M-100+H)⁺; Rétention time: 7.39 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 - 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 12: (12Æ)-17-Amino-12-methyI-6,15-bis(trifluoromethyl)-8,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol

A solution of ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(12/?)-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-8,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-17-yl]carbamate (17.5 mg, 0.0272 mmol) was heated at 100 °C in a microwave reactor for 2 h. The reaction was concentrated under vacuum to provide as a yellow foam, (1272)-17 amino-12-methyl-6,15-bis(trifluoromethyl)-8,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (12 mg, 95 %). ESI-MS m/z cale. 442.1076, found 443.5 (M+l)⁺; Rétention time: 6.08 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 100 % mobile phase B over 12 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 13: (12R)-17-Amino-12-methyl-6,15-bis(trifluoromethyI)-8,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI (diastereomer 1), Compound 70, and (12Æ)-17-amino-12-methyl-6,15-bis(trifIuoromethyl)-8,13,19-trioxa420

3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI (diastereomer 2), Compound 71

diastereomer 1 diastereomer 2 ( 127?)-1 7-Amino-12-methyl-6,15-bis(trifluoromethyl)-8,13,19-trioxa-3,4,l 8triazatricyclo[12.3.l.l2,5]nonadeca-l(l8),2,4,l4,l6-pentaen-6-ol (19 mg, 0.04296 mmol) was separated into the individual diastereomers by chiral SFC using a normal phase SFC-MS method with a Phenomenex LUX-4 column (250 X 10 mm; 5pm) at 50 °C (mobile phase was 8 % MeOH (+ 20 mM NH3) / 92 % CO2 at a 10 mL/min flow, concentration of the sample was 20.3 mg/mL in methanol, injection volume = 70 pL with an outlet pressure of 128 bar and détection wavelength of 224 nm). The first peak to elute afforded as a light yellow solid, (127?)-17-amino12-methy 1-6,15-bis(trifluoromethy l)-8,13,19-trioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-6-ol (diastereomer 1) (2.5 mg, 26 %). ’H NMR (400 MHz, Chloroformd) δ 7.41 (d, J = 0.8 Hz, 1H), 5.16 (s, 2H), 4.85-4.71 (m, 1H), 4.12 (d, J = 10.6 Hz, 1 H), 3.993.88 (m, 2H), 3.64 (ddd, J = 8.9, 6.5, 4.6 Hz, 1H), 2.85 - 2.77 (m, 1H), 2.07 - 1.96 (m, 2H), 1.78 - 1.67 (m, 1H), 1.40 (d, J =6.4 Hz, 3H), 1.35- 1.20 (m, 1H). ESI-MS m/z cale. 442.10757, found 443.2 (M+l)⁺; Rétention time: 1.76 minutes. ESI-MS m/z cale. 442.10757, found 443.2 (M+l)+ ; Rétention time: 1.76 minutes. LCMS Method: Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 % to 99 % mobile phase B over 2.9 minutes. Mobile phase A = H₂O (0.05 % CF₃CO₂H). Mobile phase B = acetonitrile (0.035 % CF₃CO₂H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

The second peak to elute afforded as a light yellow solid, (12Æ)-17-amino-12-methyl6,15-bis(trifluoromethyl)-8,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca1(18),2,4,14,16-pentaen-6-ol (diastereomer 2) (1.9 mg, 19 %). ’H NMR (400 MHz, Chloroformd) δ 7.45 - 7.38 (m, 1H), 5.21 (s, 2H), 4.86 - 4.77 (m, 1H), 4.16 (d, J = 10.4 Hz, 1H), 4.02 (d, J = 10.3 Hz, 1H), 3.97 - 3.91 (m, 1H), 3.74 - 3.67 (m, 1H), 2.79 - 2.67 (m, 1H), 1.96 - 1.87 (m, 1H), 1.87 - 1.75 (m, 1H), 1.41 (d, J = 6.5 Hz, 3H), 1.39- 1.34 (m, 1H), 1.34 - 1.24 (m, 1H). ESI-MS m/z cale. 442.10757, found 443.2 (M+l)⁺; Rétention time: 1.78 minutes. LCMS Method: Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 % to 99 % mobile phase B over 2.9 minutes.

421

Mobile phase A = H₂O (0.05 % CF3CO2H). Mobile phase B = acetonitrile (0.035 % CF3CO2H).

Example 50: Préparation of (127?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-9,13,19trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 1), Compound 72, and (12Æ)-17-amino-12-methyl-6,15-bis(trifluoromethyl)9,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 2), Compound 73

422

Step 1: l,l,l-Trifluoro-2-(2-furyl)pent-4-en-2-oI

Allylmagnesium chloride in THF (53 mL of 2 M, 106 mmol) was diluted with ether (60 mL) and cooled in an ice-water bath under nitrogen. With rapid stirring, 2,2,2-trifluoro-l-(2furyl)ethanone (4.2 g, 20.478 mmol) in ether (10 mL) was added quickly. The mixture was stirred at 0 °C for 30 min then cooled to -20 °C. Aqueous hydrochloric acid (10 mL, 2 N) was added and the layers were separated. The ether layer was washed with brine, dried over anhydrous MgSCU, filtered and concentrated to afford as a crude yellow oil, l,l,l-trifluoro-2-(210 furyl)pent-4-en-2-ol (5 g, quant.). 'H NMR (500 MHz, Chloroform-d) δ 7.45 (s, 1H), 6.48 (d, J = 3.4, 1H), 6.41 (d, J = 3.3, 1H), 5.70 - 5.57 (m, 1H), 5.29 - 5.15 (m, 2H), 3.05 (s, 1H), 3.00 2.89 (m, 1H), 2.81 -2.71 (m, 1H).

423

Step 2: 2-[l-Benzyloxy-l-(trifluoromethyl)but-3-enyI]furan

l,l,l-Trifluoro-2-(2-furyl)pent-4-en-2-ol (3 g, 11.641 mmol) was dissolved in DMF (30 mL) and the solution was cooled in an ice-water bath. Sodium hydride in minerai oil (822 mg, 20.552 mmol) was added and the solution became a light brownish mixture. The mixture was stirred 5 minutes then bromomethylbenzene (3.024 g, 2.1 mL, 17.681 mmol) was added followed by tetrabutylammonium iodide (878 mg, 2.377 mmol). The mixture was stirred at 30 °C for 16 h. Saturated aqueous NH4CI (15 mL) was added followed by EtOAc (30 mL). The layers were separated and the organic layer was washed with water then brine, dried over anhydrous MgSO4, filtered and concentrated. The residue was purified by silica gel chromatography (40g column), using a gradient from 0 % to 15 % EtOAc in hexanes to afford as a pale yellow oil, 2-[l-benzyloxy-l-(trifluoromethyl)but-3-enyl]furan (3.4 g, 89 %). 'H NMR (500 MHz, Chloroform-d) δ 7.59 - 7.44 (m, 1H), 7.43 - 7.23 (m, 5H), 6.60 (d, J = 3.4 Hz, 1H), 6.51 - 6.36 (m, 1H), 5.26 - 5.03 (m, 3H), 4.55 - 4.35 (m, 2H), 3.17 - 2.91 (m, 2H).

Step 3: 3-Benzyloxy-4,4,4-trifluoro-3-(2-furyl)butanal

2-[l-Benzyloxy-l-(trifluoromethyl)but-3-enyl]furan (5.3 g, 16.994 mmol) was mixed with THF (40 mL) and water (20 mL) at room température. NaIO₄ (10.9 g, 50.960 mmol), 2,6 lutidine (4.4304 g, 4.8 mL, 41.346 mmol) and OsO4 in tert-butanol (1.2 g, 2.5 % w/w, 0.118 mmol) were added. After 10 minutes of stirring, more water (5 mL) was added. The mixture was stirred at room température for 15 h. EtOAc (15 mL) and water (15 mL) were added and the layers were separated. The organic layer was washed with brine, dried over anhydrous MgSO4, filtered and concentrated. The residue was purified by silica gel chromatography (80g column), using a gradient from 0 % to 15 % EtOAc in hexanes, to afford as a pale yellow oil, 3benzyloxy-4,4,4-trifluoro-3-(2-furyl)butanal (3.72 g, 70 %). Ή NMR (500 MHz, Chloroform-d) δ 9.87 (t, J = 2.2 Hz, 1H), 7.54 (s, 1H), 7.44 - 7.12 (m, 5H), 6.69(d, J = 3.1 Hz, 1H), 6.60 - 6.41 (m, 1H), 4.57 (d, J = 11.0 Hz, 1H), 4.39 (d, J = 11.0 Hz, 1H), 3.12 (d, J= 2.7 Hz, 2H).

424

Step 4: 3-Benzyloxy-4,4,4-trifIuoro-3-(2-furyI)butan-l-ol

3-Benzyloxy-4,4,4-trifluoro-3-(2-furyl)butanal (3.72 g, 11.849 mmol) was dissolved in MeOH (40 mL) and cooled in an ice-water bath. NaBLL (480 mg, 0.5079 mL, 12.688 mmol) was added and the mixture was stirred at the same température for 15 minutes. Saturated aqueous NH4CI (5 mL) was added and the mixture was concentrated to 1/3 of its original volume. EtOAc (40 mL) and water (40 mL) were added and the resulting layers were separated. The organic layer was washed with brine, dried over anhydrous MgSO4, filtered and concentrated to afford as a colorless oil, 3-benzyloxy-4,4,4-trifluoro-3-(2-furyl)butan-l-ol (3.7 g, 94 %). ’H NMR (500 MHz, Chloroform-d) δ 7.52 (s, 1H), 7.41 - 7.27 (m, 5H), 6.67 - 6.54 (m, 1H), 6.51 - 6.39 (m, 1H), 4.54 (d, J = 10.9 Hz, 1H), 4.43 (d, J = 10.9 Hz, 1H), 3.95 - 3.80 (m, 2H), 2.57 - 2.40 (m,

2H), 2.00 (s, 1H).

Step 5: [(l.S)-3-[3-Benzyloxy-4,4,4-trifluoro-3-(2-furyl)butoxy]-l-methyl-propoxy]-fertbutyl-dimethyl-silane

3-Benzyloxy-4,4,4-trifluoro-3-(2-furyl)butan-l-ol (980 mg, 2.9373 mmol) was dissolved in DMF (8 mL) and the mixture was briefly chilled with an ice-water bath. Sodium hydride in minerai oil (202 mg, 5.0505 mmol) was added. The mixture was stirred at room température for 15 minutes. /er/-Butyl-[(15)-3-iodo-l-methyl-propoxy]-dimethyl-silane (2.15 g, 6.499 mmol) in DMF (2 mL) was added and the mixture was stirred at room température under nitrogen balloon for 72 h. Saturated aqueous NH4CI (5 mL) was added followed by EtOAc (20 mL) and water (20 mL). The layers were separated and the organic layer was washed with more water (2 X 20 mL) and brine then dried over anhydrous MgSO4, filtered and concentrated. The residue was purified by silica gel chromatography (40g column), using a gradient from 0 % to 10 % EtOAc in hexanes to afford as a colorless oil, [(15)-3-[3-benzyloxy-4,4,4-trifluoro-3-(2-furyl)butoxy]-lmethyl-propoxy]-ierZ-butyl-dimethyl-silane (760 mg, 51 %). ’H NMR (500 MHz, Chloroform-d) δ 7.49 (s, 1H), 7.37 to 7.25 (m, 5H), 6.58 (d, J = 3.5 Hz, 1H), 6.47 to 6.36 (m, 1H), 4.55 to 4.36

425 (m, 2H), 3.98 to 3.85 (m, 1H), 3.61 to 3.51 (m, 2H), 3.51 to 3.37 (m, 2H), 2.69 to 2.35 (m, 2H), 1.60 to 1.50 (m, 2H), 1.15 to 1.09 (m, 3H), 0.94 to 0.84 (m, 9H), 0.06 to -0.02 (m, 6H).

Step 6: 2-Benzyloxy-4-[(35)-3-[ter/-butyI(dimethyl)silyl]oxybutoxy]-2(trifluoromethyl)butanoic acid

[(15)-3-[3-benzyloxy-4,4,4-trifluoro-3-(2-furyl)butoxy]-l-methyl-propoxy]-/er/-butyldimethyl-silane (700 mg, 1.2946 mmol) was mixed in CH3CN (5.5 mL), CCI4 (5.5 mL) and water (9 mL) at room température. NalCfi (1.68 g, 7.8544 mmol) was added, followed by ruthenium(III) chloride (15 mg, 0.0723 mmol). The mixture was stirred at room température for 15 h then more NaICh (278 mg, 1.2997 mmol) was added and the mixture was stirred for an additional hour. Ethyl acetate (30 mL) and water (30 mL) were added and the layers were separated. The organic layer was washed with brine, dried over anhydrous MgSCfi, filtered and concentrated to afford crude 2-benzyloxy-4-[(35)-3-[Zer/-butyl(dimethyl)silyl]oxybutoxy]-2(trifluoromethyl)butanoic acid (800 mg, quant.). ESI-MS m/z cale. 464.2206, found 465.6 (M+l)⁺; Rétention time: 4.33 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 7: teri-Butyl A-|[2-benzyloxy-4-[(35)-3-[Zer/-butyI(dimethyl)siIyl]oxybutoxy]-2(trifluoromethyl)butanoyl] amino] carbamate

2-Benzyloxy-4-[(35)-3-[ierZ-butyl(dimethyl)silyl]oxybutoxy]-2-(trifluoromethyl)butanoic acid (610 mg, 1.116 mmol) was dissolved in DMF (15 mL) at room température. ZerZ-Butyl Naminocarbamate (221 mg, 1.6722 mmol) was added, followed by HATU (638 mg, 1.6779 mmol) and TEA (217.8 mg, 0.3 mL, 2.1524 mmol). After 30 min, water (40 mL) and EtOAc (40 mL) were added and the layers were separated. The organic layer was washed with more water (3 X 30 mL) and brine then dried over anhydrous MgSÜ4, filtered and concentrated to fumish as a crude colorless oil, terZ-butyl A-[[2-benzyloxy-4-[(35)-3-[ter/-butyl(dimethyl)silyl]oxybutoxy]426

2-(trifluoromethyl)butanoyl]amino]carbamate (260 mg, 38 %). ESI-MS m/z cale. 578.2999, found 579.7 (M+l)⁺; Rétention time: 5.1 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 8: 2-Benzyloxy-4-[(35)-3-[fôr/-butyl(dimethyl)siIyI]oxybutoxy]-2(trifluoromethyl)butanehydrazide

teri-Butyl 7V-[[2-benzyloxy-4-[(3S)-3-[rer/-butyI(dimethyl)silyl]oxybutoxy]-2(trifluoromethyl)butanoyl]amino]carbamate (268 mg, 0.4399 mmol) and 1,1,1,3,3,3-hexafluoro2-propanol (32 g, 20 mL, 190.43 mmol) were sealed in a microwave tube and heated at 100 °C for 110 minutes. The mixture was then concentrated and the residue was purified by silica gel chromatography (40 g column) using a gradient from 0 % to 15 % EtOAc in hexanes to afford as a white foam, 2-benzyloxy-4-[(3S)-3-[ZerZ-butyl(dimethyl)silyl]oxybutoxy]-2(trifluoromethyl)butanehydrazide (127 mg, 57 %). ESI-MS m/z cale. 478.2475, found 479.3 (M+l)⁺; Rétention time: 3.89 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 %

CF3CO2H).

Step 9: tert-Butyl 7V-[2-[[[2-benzyloxy-4-[(3.S')-3-[tert-butyl(dimethyI)silyl]oxybutoxy]-2(trifluoromethyl)butanoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3pyridyl] carbamate

2-Benzyloxy-4-[(3S)-3-[ZerZ-butyl(dimethyl)silyl]oxybutoxy]-2(trifluoromethyl)butanehydrazide (127 mg, 0.2521 mmol) was dissolved in EtOAc (2 mL) at room température. 6-Bromo-3-(ier/-butoxycarbonylamino)-5-(trifluoromethyl)pyndine-2carboxylic acid (118 mg, 0.3064 mmol) was added, followed by pyridine (97.8 mg, 0.1 mL, 1.2364 mmol) and T3P in EtOAc (160.35 mg, 0.3 mL of 50 % w/w, 0.2520 mmol). The mixture 427 was stirred at room température for 14 h. The mixture was concentrated and the residue was purified by silica gel chromatography (40g column) using a gradient from 0 % to 50 % EtOAc in hexanes to afford as a white foam, ZerZ-butyl A/-[2-[[[2-benzyloxy-4-[(3S)-3-[Zeributyl(dimethyl)silyl]oxybutoxy]-2-(trifluoromethyl)butanoyl]amino]carbamoyl]-6-bromo-5(trifluoromethyl)-3-pyridyl]carbamate (200 mg, 84 %). ESI-MS m/z cale. 844.2302, found 791.5 (M-53)⁺; Rétention time: 5.09 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 10: tert-Butyl A-[2-[5-[l-benzyloxy-3-[(35)-3-[terf-butyI(dimethyl)silyl]oxybutoxy]-l(trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3pyridyl] carbamate

ZerZ-Butyl7V-[2-[[[2-benzyloxy-4-[(3S)-3-[terZ-butyl(dimethyl)silyl]oxybutoxy]-2(trifluoromethyl)butanoyl]amino]carbamoyl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (200 mg, 0.2128 mmol) was dissolved in CH3CN (2 mL) at room température. DIEA (96.46 mg, 0.13 mL, 0.7463 mmol) was added followed by TsCl (60 mg, 0.3147 mmol). The mixture was stirred at room température for 5 h then concentrated to 1/3 of its volume and diluted with EtOAc (20 mL) and saturated aqueous NH4CI (20 mL). The resulting layers were separated and the organic layer was washed with brine, dried over anhydrous MgSÜ4, filtered and concentrated. The residue was purified by silica gel chromatography (40g column) using a gradient from 0 % to 10 % EtOAc in hexanes to afford as a colorless oil, feri-butyl 7V-[2-[5-[lbenzyloxy-3-[(3S)-3-[terZ-butyl(dimethyl)silyl]oxybutoxy]-l-(trifluoromethyl)propyl]-l,3,4oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (110 mg, 59 %). ESI-MS m/z cale. 826.2196, found 771.5 (M-55)⁺; Rétention time: 5.25 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

428

Step 11: ZerZ-Butyl7V-[2-[5-[l-benzyloxy-3-[(3S)-3-[ZerZ-butyl(dimethyl)silyI]oxybutoxy]-l(trifliioromethyl)propyl]-l,3,4-oxadiazoI-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7VZerZ-butoxycarbonyl-carbamate

ZerZ-Butyl 7V-[2-[5-[l-benzyloxy-3-[(35)-3-[ZerZ-butyl(dimethyl)silyl]oxybutoxy]-l(trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]carbamate (110 mg, 0.1262 mmol) was mixed with MTBE (2 mL) at room température. Di-ZerZ-butyl dicarbonate (51 mg, 0.0537 mL, 0.2337 mmol) was added followed by DMAP (1 mg, 0.0082 mmol) and DIEA (22.26 mg, 0.03 mL, 0.1722 mmol). The mixture was stirred at room température for 14 h and was then concentrated and purified by silica gel chromatography (24g column) using a gradient from 0 % to 20 % EtOAc in hexanes to afford as a white foam, ZerZbutyl 7V-[2-[5-[ 1 -benzyloxy-3-[(3S)-3-[ZerZ-butyl(dimethyl)silyl]oxybutoxy]-1 (trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZbutoxycarbonyl-carbamate (122 mg, 99 %). ESI-MS m/z cale. 926.272, found 773.5 (M-154)⁺; Rétention time: 5.17 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 %

CF3CO2H).

Step 12: ZerZ-Butyl A^r-[2-[5-[l-benzyIoxy-3-[(35)-3-[ZerZ-butyl(dimethyl)siIyl]oxybutoxy]-l(trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyI)-3-pyridyl]-7VZerZ-butoxycarbonyl-carbamate

ZerZ-Butyl/V-[2-[5-[l-benzyloxy-3-[(3S)-3-[ZerZ-butyl(dimethyl)silyl]oxybutoxy]-l(trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-bromo-5-(trifluoromethyl)-3-pyridyl]-7V-ZerZbutoxycarbonyl-carbamate (122 mg, 0.1249 mmol) was dissolved in DMSO (1 mL). Césium acetate (72 mg, 0.3751 mmol) was added and the mixture was placed in a 70 °C oil bath and stirred under a balloon of nitrogen for 20 h. The mixture was cooled to room température and 429 diluted with EtOAc (20 mL) and saturated aqueous NaHCOj (20 mL). The layers were separated and the organic layer was washed with more water (2 X 20 mL) and brine then dried over anhydrous MgSO4, filtered and concentrated to afford as a pale yellow oil, Zeri-butyl 7V-[2-[5-[ 1 benzyloxy-3-[(3S)-3-[teri-butyl(dimethyl)silyl]oxybutoxy]-l-(trifluoromethyl)propyl]-l,3,4oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-teri-butoxycarbonyl-carbamate (120 mg, 94 %). ESI-MS m/z cale. 864.3564, found 765.7 (M-99)⁺; Rétention time: 4.86 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 13: tert-Butyl A-[2-[5-[l-benzyloxy-3-I(3X)-3-hydroxybutoxy]-l(trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7Vtert-butoxycarbonyl-carbamate

ier/-Butyl7V-[2-[5-[l-benzyloxy-3-[(3S)-3-[ier/-butyl(dimethyl)silyl]oxybutoxy]-l(trifluoromethyl)propyl]-l,3,4-oxadiazol-2-yl]-6-hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-/<?r/butoxycarbonyl-carbamate (120 mg, 0.1179 mmol) was dissolved in THF (2 mL) at room température. Tetrabutylammonium fluoride in THF (88.7 mg, 0.1 mL, 0.3392 mmol) was added. The mixture was stirred at room température for 5 h and then placed in a 40 °C oil bath. Stirring continued for 14 h then more tetrabutylammonium fluoride in THF (354.8 mg, 0.4 mL, 1.357 mmol) was added. After 6 h, DMF (2 mL) was added and stirring was continued for 15 h. More tetrabutylammonium fluoride in THF (354.8 mg, 0.4 mL, 1.357 mmol) was added. After another 16 h, the mixture was diluted with EtOAc (30 mL) and water (40 mL) and the layers were separated. The organic layer was washed with brine, dried over anhydrous MgSÛ4, filtered and concentrated. The residue was purified by silica gel chromatography (12 g column), using a gradient from 10 % to 90 % EtOAc in hexanes to afford as a white foam, ieri-butyl 7V-[2-[5-[lbenzyloxy-3-[(3S)-3 -hydroxybutoxy]-1 -(trifluoromethyl)propyl]-1,3,4-oxadiazol-2-yl]-6hydroxy-5-(trifluoromethyl)-3-pyridyl]-7V-terZ-butoxycarbonyl-carbamate (73 mg, 81 %). ESIMS m/z cale. 750.2699, found 651.7 (M-99)⁺; Rétention time: 3.8 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

430

Step 14: toï-Butyl A-[(12J?)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-9,13,19trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-7V-terZbutoxycarbonyl-carbamate

terZ-Butyl 7V-[2-[5-[l-benzyloxy-3-[(35)-3-hydroxybutoxy]-l-(trifluoromethyl)propyl]1,3,4-oxadiazol-2-yl] -6-hydroxy-5-(trifluoromethyl)-3 -pyridyl] -A-ZerZ-butoxycarbonylcarbamate (70 mg, 0.0914 mmol) was dissolved in toluene (9 mL) at room température and PPI13 (77 mg, 0.068 mL, 0.2936 mmol) was added in one portion followed by DIAD (62.64 mg, 0.06 mL, 0.3098 mmol) via syringe. The mixture was stirred at room température for 7 h then concentrated. The residue was purified by silica gel chromatography (12g column), using a gradient from 0 % to 15 % EtOAc in hexanes to afford as a white solid, ZerZ-butyl 7V-[(127?)-6benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-9,13,19-trioxa-3,4,18triazatricyclof 12.3.1.12,5]nonadeca-1 ( 18),2,4,14,16-pentaen-17-yl]-7V-ZerZ-butoxycarbonylcarbamate (40 mg, 57 %). ESI-MS m/z cale. 732.2594, found 733.6 (M+l)⁺; Rétention time: 4.75 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 15: ZerZ-Butyl A^LZi’rZ-butoxycarbonyl-A-[(127?)-6-liydroxy-12-methyl-6,15bis(trifluoromethyl)-9,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-17-yl]carbamate

ZerZ-Butyl N-[( 12/?)-6-bcnzyloxy-12-methyl-6,15-bis(trifluoromethyl)-9,13,19-trioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]-7V-ZerZbutoxycarbonyl-carbamate (40 mg, 0.0519 mmol) was dissolved in EtOH (2 mL) and 10 % palladium on carbon (20 mg, 0.1879 mmol) was added. The mixture was purged with hydrogen gas several times and hydrogenated at 50 psi on a Pair shaker for 20 h. The mixture was then

431 fïltered through a celite pad, washed with MeOH (~15 mL) and the filtrate was concentrated thoroughly to afford as a crude white solid, ZerZ-butyl 7V-ZerZ-butoxycarbonyl-7V-[(12/?)-6hydroxy-12-methyl-6,15-bis(trifluoromethyl)-9,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-17-yl]carbamate (33 mg, 94 %). ESI-MS m/z cale. 642.2124, found 643.4 (M+l)⁺; Rétention time: 4.88 minutes. LCMS Method: Merck Millipore Chromolith SpeedROD Cis column (50 X 4.6 mm) and a dual gradient run from 5 % to 100 % mobile phase B over 6 minutes. Mobile phase A = water (0.1 % CF3CO2H). Mobile phase B = acetonitrile (0.1 % CF3CO2H).

Step 16: (127?)-17-Amino-12-methyI-6,15-bis(trifIuoromethyl)-9,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI

ZerZ-Butyl 7V-Zert-butoxycarbonyl-7V-[( 12R)-6-hydroxy-12-methyL6,15bis(trifluoromethyl)-9,13,19-trioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-l (18),2,4,14,16pentaen-17-yl]carbamate (33 mg, 0.0488 mmol) was mixed with 1,1,1,3,3,3-hexafluoro-2propanol (3.192 g, 2 mL, 18.996 mmol) in a microwave vessel then sealed and heated at 100 °C for 130 minutes. The mixture was then cooled to room température and concentrated. The residue was purified by silica gel chromatography (12g column), using a gradient from 5 % to 60 % EtOAc in hexanes to afford as a pale yellow solid, (12R)-17-amino-12-methyl-6,15bis(trifluoromethyl)-9,13,19-trioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol (21 mg, 89 %). ESI-MS m/z cale. 442.1076, found 443.5 (M+l)⁺; Rétention time: 2.71 minutes (average of two diastereomeric peaks). LCMS Method: Waters Cortex 2.7 pm particle size Cis (3.0 mm X 50 mm), 55 °C; flow: 1.2 mL/min; mobile phase: 100 % water with 0.1 % trifluoroacetic acid then 100 % acetonitrile with 0.1 % trifluoroacetic acid, gradient of 5 % to 100 % B over 4 min, with équilibration at 100 % B for 0.5 min, then 5 % B over 1.5 min.

Step 17: (12Æ)-17-amino-12-methyI-6,15-bis(trifluoromethyl)-9,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-oI (diastereomer 1), Compound 72, and (12/?)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-9,13,19-trioxa432

3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 2), Compound 73

diastereomer 1 diastereomer 2 (12R)-17-amino-12-methyl-6,15-bis(trifluoromethyl)-9,13,19-trioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (20 mg, 0.04522 mmol) was separated into the individual diastereomers by chiral SFC using a normal phase SFC-MS method utilizing a ChiralCel OJ column (250 X 10 mm; 5 pm particle size) at 50 °C (mobile phase = 7 % MeOH (+ 20 mM NH3) / 93 % CO2 at a 10 mL/min flow, concentration of the sample was 22.0 mg/mL in methanol, injection volume = 70 pL with an outlet pressure of 140 bar, détection wavelength of 224 nm). The first peak to elute afforded as a pale yellow solid, ( 127/)-17-amino12-methyl-6,15-bis(trifluoromethyl)-9,13,19-trioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-6-ol (diastereomer 1) (7.2 mg, 69 %). ’H NMR (400 MHz, Chloroformd) δ 7.40 (s, 1H), 5.39 (pd, J = 6.5,4.3 Hz, 1H), 5.14 (s, 2H), 3.99 (s, 1H), 3.86 - 3.78 (m, 1H), 3.67 - 3.61 (m, 1H), 3.60 - 3.51 (m, 2H), 2.68 (ddd, J = 14.9, 8.9, 5.9 Hz, 1H), 2.52 (dt, J = 15.2, 4.0 Hz, 1H), 2.40 - 2.30 (m, 1H), 1.64 - 1.58 (m, 1H), 1.42 (d, J = 6.5 Hz, 3H). ESI-MS m/z cale. 442.10757, found 443.2 (M+l)⁺; Rétention time: 1.76 minutes. LCMS Method: Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 % to 99 % mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

The second peak to elute afforded as a pale yellow solid, ( 127/)-17-amino-12-methyl6,15-bis(trifluoromethyl)-9,13,19-trioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadecal(18),2,4,14,16-pentaen-6-ol (diastereomer 2) (5.2 mg, 51 %). 'H NMR (400 MHz, Chloroformd) δ 7.42 (s, 1H), 5.25 (ddt, J = 10.2, 6.5, 3.3 Hz, 1H), 5.10 (s, 2H), 4.51 (s, 1H), 3.95 (td, J = 9.2, 4.5 Hz, 1H), 3.86 (ddd, J = 12.1, 10.3, 1.7 Hz, 1H), 3.62 - 3.51 (m, 2H), 2.75 - 2.62 (m, 2H), 2.35 (dt, J = 14.9, 4.7 Hz, 1H), 1.49 - 1.37 (m, 4H). ESI-MS m/z cale. 442.10757, found 443.2 (M+l)⁺; Rétention time: 1.76 minutes. LCMS Method: Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle) made by Waters (pn: 186002350), and a dual gradient run from 1 % to 99 % mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume = 1.5 pL, and column température = 60 °C.

433

Example 51: Préparation of (157?)-15-methyl-18-(trifluoromethyl)-16,22-dioxa-3,4,7,8,21pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,17,19-heptaen-20-amine, Compound 74

434

Step 1: Methyl 3-[bis(terf-butoxycarbonyl)aimno]-6-[(LÆ)-l-methylbut-3-enoxy]-5 (trifluoromethyl)pyridine-2-carboxylate

To a solution of methyl 3-[bis(tert-butoxycarbonyl)amino]-6-bromo-5(trifluoromethyl)pyridine-2-carboxylate (500 mg, 1.001 mmol) in DMSO (5 mL) was added (27?)-pent-4-en-2-ol (160 pL, 1.555 mmol), césium carbonate (521 mg, 1.599 mmol) and iodocopper (54 mg, 0.2835 mmol) and the reaction mixture was heated at 100 °C for 3 h. The reaction mixture was cooled to room température and poured on crushed ice and extracted with ethyl acetate and washed with brine. The organics were separated, dried over sodium sulfate, filtered and evaporated. The résultant brown residue was purified by silica gel column chromatography using a shallow gradient from 100 % hexanes to 30 % ethyl acetate in hexanes to afford as a colorless oil, methyl 3-[bis(tert-butoxycarbonyl)amino]-6-[(17?)-l-methylbut-3enoxy]-5-(trifhioromethyl)pyridine-2-carboxylate (216 mg, 43 %). ESI-MS m/z cale. 504.20834, found 505.0 (M+l)⁺; Rétention time: 1.73 minutes. LCMS Method: Acquity UPLC BEH Cis column (50 x 2.1 mm, 1.7 pm particle size) made by Waters (pn: 186002350), and a dual gradient run from 30 - 99 % mobile phase B over 2.9 minutes. Mobile phase A = H2O (0.05 % CF3CO2H). Mobile phase B = CH3CN (0.035 % CF3CO2H). Flow rate = 1.2 mL/min, injection volume =1.5 pL, and column température = 60 °C.

Step 2: 3-(terf-ButoxycarbonyIamino)-6-[(l.R)-l-methylbut-3-enoxy]-5(trifIuoromethyl)pyridine-2-carboxylic acid

To a solution of methyl 3-[bis(tert-butoxycarbonyl)amino]-6-[(17?)-l-methylbut-3enoxy]-5-(trifluoromethyl)pyridine-2-carboxylate (3.3 g, 6.5412 mmol) in MeOH (24 mL) and THF (24 mL) was added a solution of lithium hydroxide monohydrate (1.2 g, 28.596 mmol) in water (12 mL). The mixture was stirred at 50 °C for 16 h. The reaction mixture was then cooled down to room température and aqueous IN HCl was added until pH = 2 was reached. The volatiles were removed under reduced pressure and the product was extracted with ethyl acetate (3 X 20 mL). The combined organic phases were washed with brine (20 mL), dried over sodium

435 sulfate, filtered and concentrated under reduced pressure which afforded as a yellow oil, 3-(tertbutoxycarbonylamino)-6-[( 17?)-1 -methylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carboxylic acid (2.66 g, 89 %). ‘H NMR (400 MHz, DMSO-d₆) δ 12.12 - 11.38 (m, 1H), 8.78 (s, 1H), 5.92 5.69 (m, 1H), 5.42 - 5.33 (m, 1H), 5.14 - 5.06 (m, 1H), 5.06 - 5.01 (m, 1H), 2.44 - 2.37 (m, 2H), 1.46 (s, 9H), 1.25 (d, J = 6.4 Hz, 3H). ^,9F NMR (377 MHz, DMSO-d₆) δ -62.33 (s, 3F). ESI-MS m/z cale. 390.1403, found 389.0 (M-l)'; Rétention time: 2.28 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 3: Ethyl l-benzylpyrazole-3-carboxylate

To a suspension of ethyl 177-pyrazole-5-carboxylate (20 g, 142.71 mmol) and potassium carbonate (60 g, 434.13 mmol) in dimethyl formamide (200 mL) was added benzyl bromide (25.92 g, 18 mL, 151.55 mmol) at room température and the mixture was stirred for 3 hours at room température. The mixture was diluted with ethyl acetate (500 mL) and washed with water (500 mL). The aqueous layer was extracted with ethyl acetate (250 mL). The combined organic layers were washed with water (500 mL), brine (500 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluted with a gradient of 100 % heptane to 30 % ethyl acetate in heptane) to give as the second eluting regioisomeric product and a white solid, ethyl l-benzylpyrazole-3carboxylate (21.5 g, 65 %). ’H NMR (400 MHz, CDC1₃) δ 7.40 - 7.30 (m, 4H), 7.26 - 7.21 (m, 2H), 6.83 (d, J = 2.4 Hz, 1H), 5.40 (s, 2H), 4.42 (q, J = 7.1 Hz, 2H), 1.41 (t, J = 7.1 Hz, 3H). ESIMS m/z cale. 230.1055, found 231.2 (M+l)⁺; Rétention time: 1.85 minutes; LCMS Method: Kinetex Polar Cis 3.0 X 50 mm, 2.6 pm, 3 min, 5-95 % acetonitrile in water (0.1 % formic acid), flow =1.2 mL/min.

Step 4: Ethyl l-benzyl-4-bromo-pyrazole-3-carboxylate

To a solution of ethyl l-benzylpyrazole-3-carboxylate (7 g, 30.4 mmol) in acetonitrile (140 mL) was added bromine (9.3069 g, 3 mL, 58.238 mmol) at room température. After stirring at room température ovemight, additional bromine (4.9637 g, 1.6 mL, 31.06 mmol) was added and the mixture was stirred at room température for 1 day. The mixture was concentrated under

436 reduced pressure, diluted with ethyl acetate (20 mL), and washed with aqueous 10 % sodium thiosulfate (200 mL). The aqueous layer was extracted with ethyl acetate/tetrahydrofuran (1/1, 200 mL). The combined organic layer was dried over sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluted with a gradient from 0 % to 35 % ethyl acetate in heptane) to give as a white solid, ethyl 1benzyl-4-bromo-pyrazole-3-carboxylate (9.1 g, 97 %); ’H NMR (400 MHz, CDC13) δ 7.44 7.33 (m, 4H), 7.29 - 7.24 (m, 2H), 5.36 (s, 2H), 4.45 (q, J = 7.2 Hz, 2H), 1.43 (t, J = 7.1 Hz, 3H). ESI-MS m/z cale. 308.016, found 309.1 (M+l)⁺; Rétention time: 1.98 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 5: Ethyl 4-alIyI-l-benzyI-pyrazole-3-carboxylate

A solution of ethyl l-benzyl-4-bromo-pyrazole-3-carboxylate (8.7 g, 28.141 mmol) in tetrahydrofuran (130 mL) and water (25 mL) was degassed by bubbling nitrogen for 15 minutes. l-Allyl-3,3,4,4-tetramethyl-borolane (5.6 g, 6.25 mL, 34.127 mmol) and 1,1'bis(diphenylphosphino)ferrocene palladium(II) chloride, complex with dichloromethane (3 g, 3.6736 mmol) were added followed by the addition of césium carbonate (42 mL of 2 M, 84 mmol) under nitrogen. The reaction mixture was heated ovemight at 100 °C. After cooling down to room température, the reaction mixture was diluted with ethyl acetate (150 mL) filtrated through a pad of celite and rinsed with ethyl acetate (2 X 100 mL). The filtrate was washed with water (2 X 200 mL) and brine (400 mL). The organic phase was then dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The resulting mixture was purified by silica gel column chromatography (gradient of 0 % to 30 % ethyl acetate in heptanes). The desired fractions were collected and the solvent was concentrated under reduced pressure to afford as a white solid, ethyl 4-allyl-l-benzyl-pyrazole-3-carboxylate (5.02 g, 64 %). 1H NMR (400 MHz, CDC13) δ 7.41 - 7.31 (m, 3H), 7.26 - 7.21 (m, 2H), 7.15 (s, 1H), 5.94 (ddt, J = 17.0, 10.2, 6.6 Hz, 1H), 5.35 (s, 2H), 5.09 - 4.98 (m, 2H), 4.42 (q, J = 7.2 Hz, 2H), 3.50 (d, J = 6.6 Hz, 2H), 1.41 (t, J = 7.2 Hz, 3H). ESI-MS m/z cale. 270.1368, found 271.2 (M+l)+; Rétention time: 2.0 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate = 1.2 mL/min).

437

Step 6: 4-AIlyI-l-benzyl-pyrazole-3-carboxylic acid

A solution of lithium hydroxide (5 g, 208.78 mmol) in water (10 mL) was added to a solution of ethyl 4-allyl-l-benzyl-pyrazole-3-carboxylate (5 g, 18.015 mmol) in methanol (50 mL) and tetrahydrofuran (25 mL). The reaction was stirred at room température for 3 hours. The reaction was then concentrated to remove the methanol. The crude material was diluted in water (100 mL) and the carboxylate sodium sait was washed with heptane (50 mL) and MTBE (50 mL). The aqueous solution was acidified to pH = 2 with a 3 N aqueous hydrochloric acid solution. The carboxylic acid was extracted with dichloromethane (4 X 100 mL) and dried over sodium sulfate. The solution was filtered and concentrated to give as a light-yellow solid, 4-allyl1-benzyl-pyrazole-3-carboxylic acid (4.45 g, 99 %). 1H NMR (400 MHz, CDC13) δ 12.40 10.19 (m, 1H), 7.48 - 7.39 (m, 3H), 7.37 - 7.31 (m, 2H), 7.27 (s, 1H), 6.01 (ddt, J = 17.0, 10.2, 6.6 Hz, 1 H),5.43 (s, 2H), 5.20 - 5.04 (m, 2H), 3.59 (d, J = 6.6 Hz, 2H). ESI-MS m/z cale. 242.1055, found 243.1 (M+l)+; Rétention time: 1.79 minutes; LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 7: 4-Allyl-l-benzyl-pyrazole-3-carbohydrazide

A solution of 4-allyl-l-benzyl-pyrazole-3-carboxylic acid (4.15 g, 16.616 mmol) and triethylamine (5.0820 g, 7 mL, 50.222 mmol) in DMF (60 mL) was treated with HATU (13 g, 34.19 mmol) and stirred at room température for 20 minutes. The reaction mixture was cooled in an ice bath and hydrazine hydrate (13.416 g, 20 mL, 174.2 mmol) was added. After about 10 minutes, the ice bath was removed and the reaction was stirred at room température for about 18 hours. The reaction mixture was transferred to a 1 L separatory tunnel with water (450 mL) and the aqueous layer was extracted with ethyl acetate (4 X 100 mL). The combined organic layers were washed with water (2 X 200 mL), brine (250 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. Silica gel chromatography (gradient from 0 % to 60 % ethyl acetate in heptanes) afforded as a colorless oil, 4-allyl-l-benzyl-pyrazole-3-carbohydrazide

438 (2.45 g, 52 %). 1H NMR (400 MHz, DMSO-d6) δ 9.11 (s, 1H), 7.66 (s, 1H), 7.44 - 7.15 (m, 5H), 5.95 (ddt, J = 17.0, 10.2, 6.6 Hz, 1H), 5.30 (s, 2H), 5.10 - 4.89 (m, 2H), 4.34 (s, 2H), 3.45 (d, J = 6.6 Hz, 2H). ESI-MS m/z cale. 256.1324, found 257.2 (M+l)+; Rétention time: 3.23 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Step 8: terf-Butyl /V-[2-[[(4-allyl-l-benzyI-pyrazole-3-carbonyl)amino]carbamoyl]-6-[(LR)l-methylbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]carbamate

To a solution of 3-(terZ-butoxycarbonylamino)-6-[(lR)-l-methylbut-3-enoxy]-5(trifluoromethyl)pyridine-2-carboxylic acid (200 mg, 0.5088 mmol) in DMF (1.5 mL) was added 4-allyl-l-benzyl-pyrazole-3-carbohydrazide (160 mg, 0.5662 mmol), HATU (235 mg, 0.618 mmol) and DIEA (371 mg, 0.5 mL, 2.8706 mmol) at room température. The solution was stirred at room température for 2 h and directly loaded on a reversed-phase Cis preparatory column. Purification was performed by reversed-phase chromatography (100 gram column. Gradient from 5 % to 100 % acetonitrile in water with 0.1 % formic acid) to afford as a white foam, tert butyl7V-[2-[[(4-allyl-l-benzyl-pyrazole-3-carbonyl)amino]carbamoyl]-6-[(lR)-l-methylbut-3enoxy]-5-(trifluoromethyl)-3-pyridyl]carbamate (238 mg, 71 %). 1H NMR (400 MHz, DMSOd6) δ 10.83 (s, 1H), 10.65 (s, 1H), 10.15 (s, 1H), 9.01 (s, 1H), 7.77 (s, 1H), 7.50 - 7.18 (m, 5H), 5.97 (ddt, J = 17.0, 10.2, 6.6 Hz, 1H), 5.89 - 5.74 (m, 2H), 5.39 (s, 2H), 5.18 - 4.94 (m, 4H), 3.46 (d, J = 6.6 Hz, 2H), 2.47 - 2.32 (m, 2H), 1.47 (s, 9H), 1.25 (d, J = 6.1 Hz, 3H). ¹⁹F NMR (377 MHz, DMSO-dô) δ -63.11 (s, 3F). ESI-MS m/z cale. 628.2621, found 529.3 (M-99)+; Rétention time: 4.3 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow = 1.5 mL/min).

439

Step 9: tert-Butyl A-|2-[5-(4-allyI-l-benzyl-pyrazol-3-yl)-l,3,4-oxadiazol-2-yl]-6-[(lÆ)-l methyIbut-3-enoxy]-5-(trifluoromethyl)-3-pyridyl]carbamate

To a solution of ZerAbutyl 7V-[2-[[(4-allyl-l-benzyl-pyrazole-3carbonyl)amino]carbamoyl]-6-[(17?)-l-methylbut-3-enoxy]-5-(trifluoromethyl)-3pyridyl]carbamate (235 mg, 0.3577 mmol) and toluenesulfonyl chloride (220 mg, 1.154 mmol) in DCE (7 mL) was added V,7V-diisopropylethylamine (0.5 mL, 2.8706 mmol) and toluenesulfonyl chloride (220 mg, 1.154 mmol). The reaction was stirred at 50 °C for 20 h then cooled down to room température. The volatiles were removed under reduced pressure. Purification was performed by reversed-phase chromatography (50 g column. Gradient from 5 % to 95 % acetonitrile in water with 0.1 % formic acid) to afford as a tan oil, ter/-butyl 7V-[2-[5-(4allyl-1 -benzyl-pyrazol-3 -yl)-1,3,4-oxadiazol-2-yl]-6-[( 1 A)-1 -methylbut-3 -enoxy] -5(trifluoromethyl)-3-pyridyl]carbamate (202 mg, 81 %). 1H NMR (400 MHz, DMSO-d6) δ 9.93 (s, 1H), 8.92 (s, 1H), 7.96 (s, 1H), 7.42 - 7.26 (m, 5H), 6.04 (ddt, J = 16.8,10.2, 6.4 Hz, 1H), 5.84 (ddt, J = 17.2, 10.1, 7.0 Hz, 1H), 5.46 (s, 2H), 5.34 (m, J = 6.1 Hz, 1H), 5.20 - 5.13 (m, 1H), 5.12 - 5.00 (m, 3H), 3.59 (d, J = 6.6 Hz, 2H), 2.50 - 2.38 (m, 2H), 1.50 (s, 9H), 1.34 (d, J = 6.1 Hz, 3H). 19F NMR (377 MHz, DMSO-d6) δ -62.80 (s, 3F). ESI-MS m/z cale. 610.2515, found 555.2 (M-55)⁺; Rétention time: 2.56 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow =1.5 mL/min, run time = 3.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 2.2 minute at 95 % acetonitrile + 0.1 % formic acid.

Step 10: tert-Butyl 7V-[(15R)-8-benzyl-15-methyl-18-(trifluoromethyl)-16,22-dioxa3,4,7,8,21-pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,12,17,19-octaen-20yljcarbamate (E/Z mixture)

440

Nitrogen gas was bubbled through a light yellow solution of fer/-butyl 7V-[2-[5-(4-allyl-1 benzyl-pyrazol-3 -yl)-l ,3,4-oxadiazol-2-yl]-6-[( 1 R)-1 -methylbut-3-enoxy]-5-(trifluoromethyl)-3 pyridyl]carbamate (200 mg, 0.2879 mmol) in dichloroethane (100 mL) for 2 hours. The solution was then placed in an oil bath set at 70 °C and a first portion of Zhan catalyst-lB (20 mg, 0.0273 mmol) was added. The reaction was stirred for 2 hours. DMSO (2 drops) was added and the reaction mixture was cooled down to room température. The volatiles were removed under reduced pressure. Purification by silica gel chromatography (gradient from 0 % to 30 % ethyl acetate in heptanes) afforded as a yellow oil, teri-butyl JV-[(15Æ)-8-benzyl-15-methyl-18(trifluoromethyl)-16,22-dioxa-3,4,7,8,21-pentazatetracyclo[15.3.1.12,5.06,10]docosal(21),2,4,6,9,12,17,19-octaen-20-yl]carbamate (E/Z mixture) (65 mg, 37 %). ¹⁹F NMR (377 MHz, DMSO-d6) δ -62.17 - -62.44 (m, 3F). ESI-MS m/z cale. 582.2202, found 583.3 (M+l)+; Rétention time: 4.51 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 11: tert-Butyl 7V-[(15R)-8-benzyl-15-methyl-18-(trifluoromethyI)-16,22-dioxa3,4,7,8,21-pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,17,19-heptaen-20yl] carbamate

To palladium hydroxide on carbon (21 mg, 20 % w/w, 0.0299 mmol) under nitrogen was added a solution of fer/-butyl 7V-[(15R)-8-benzyl-15-methyl-18-(trifluoromethyl)-16,22-dioxa3,4,7,8,21 -pentazatetracyclo[ 15.3.1.12,5.06,10]docosa-1 (21 ),2,4,6,9,12,17,19-octaen-20yl]carbamate (E/Z mixture) (60 mg, 0.0987 mmol) in methanol (5 mL). Hydrogen gas was bubbled into the mixture for 5 min and then the reaction was stirred at room température for 4 h under hydrogen atmosphère (balloon). The mixture was filtered through celite, washing with ethyl acetate (50 mL) and the filtrate was concentrated under reduced pressure. Purification of the residue by silica gel chromatography (Gradient from 5 % to 60 % ethyl acetate in heptanes) afforded as an off-white solid, ZerZ-butyl 7V-[(15R)-8-benzyl-15-methyl-18-(trifluoromethyl)16,22-dioxa-3,4,7,8,21 -pentazatetracyclof 15.3.1.12,5.06,10]docosa-1 (21 ),2,4,6,9,17,19-heptaen20-yl]carbamate (55 mg, 88 %). ESI-MS m/z cale. 584.2359, found 585.3 (M+l)+; Rétention time: 4.6 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6

441 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow = 1.5 mL/min).

Step 12: tert-Butyl 7V-[(15R)-15-methyl-18-(trifluoromethyl)-16,22-dioxa-3,4,7,8,21pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,17,19-heptaen-20-yl]carbamate

To palladium on carbon (60 mg, 5 % w/w, 0.0282 mmol) under nitrogen was added a solution of/er/-butyl7V-[(157?)-8-benzyl-15-methyl-18-(trifluoromethyl)-16,22-dioxa-3,4,7,8,21pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,17,19-heptaen-20-yl]carbamate (50 mg, 0.0787 mmol) in tetrahydrofuran (10 mL). The tube was sealed and pressurized with hydrogen gas (50 psi). The reaction mixture was stirred at 50 °C for 24 hours. The mixture was then cooled, degassed with nitrogen purge then filtered through celite, washing with ethyl acetate (25 mL). The filtrate was concentrated under reduced pressure. Purification of the residue by silica gel chromatography (Gradient from 0 % to 90 % ethyl acetate in heptanes) afforded as a yellow solid, terZ-butyl 7V-[(157?)-15-methyl-18-(trifluoromethyl)-16,22-dioxa-3,4,7,8,21pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,17,19-heptaen-20-yl]carbamate (18 mg, 45 %). 1H NMR (400 MHz, CDC13) δ 13.31 - 12.26 (m, 1H), 9.21 (s, 1H), 9.10 (s, 1H), 7.92 (s, 1H), 5.07 - 4.91 (m, 1H), 3.12 - 2.95 (m, 1H), 2.69 - 2.51 (m, 2H), 2.42 - 2.25 (m, 1H), 1.72 - 1.62 (m, 3H), 1.60 (s, 9H), 1.55 - 1.48 (m, 1H), 1.45 (d, J = 6.1 Hz, 3H). 19F NMR (377 MHz, CDC13) δ -63.70 (s, 3F). ESI-MS m/z cale. 494.1889, found 495.2 (M+l)+; Rétention time: 4.21 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

Step 13: (157?)-15-Methyl-18-(trifIuoromethyl)-16,22-dioxa-3,4,7,8,21pentazatetracyclo[15.3.1.12,5.06,10]docosa-l(21),2,4,6,9,17,19-heptaen-20-amine, Compound 74

442

To a solution of Zeri-butyl tV-[(1 5//)-15-methyl-l 8-(trifluoromcthyl)-l 6,22-dioxa3,4,7,8,21 -pentazatetracyclof 15.3.1.12,5.06,10]docosa-l (21 ),2,4,6,9,17,19-heptaen-20yl]carbamate (18 mg, 0.0357 mmol) in dichloromethane (1.8 mL) was added TFA (0.6 mL, 7.7879 mmol) and the mixture was stirred at room température for 2 hours. A saturated aqueous 5 NaHCCh solution was then added dropwise until pH = 6 - 7 was reached. Water (10 mL) and DCM (10 mL) were added and the phases were separated. The aqueous phase was extracted with DCM (3X10 mL). The combined organic layers were concentrated under reduced pressure. Purification of the residue by reversed-phase chromatography (15.5 g column. Gradient from 5 % to 90 % acetonitrile in water) afforded as a yellow solid after lyophilization, ( 157/)-15-methyl10 18-(trifluoromethyl)-16,22-dioxa-3,4,7,8,21 -pentazatetracyclof 15.3.1.12,5.06,10] docosal(21),2,4,6,9,17,19-heptaen-20-amine (11.7 mg, 83 %). 1H NMR (400 MHz, DMSO-d6) δ 13.44 (br. s., 1H), 7.86 (br. s., 1H), 7.74 (s, 1H), 6.34 (s, 2H), 4.95 - 4.80 (m, 1H), 2.99 - 2.81 (m, 1H), 2.62 - 2.52 (m, 1H), 2.47 - 2.37 (m, 1H), 2.34 - 2.17 (m, 1H), 1.65 - 1.41 (m, 3H), 1.40 - 1.28 (m, 4H). 19F NMR (377 MHz, DMSO-d6) δ -62.32 (s, 3F). ESI-MS m/z cale. 394.1365, found 395.1 15 (M+l)⁺; Rétention time: 3.53 minutes. LCMS Method: XBridge Cis column (4.6 X 75 mm, 5 pm particle size, 6 minute run with 1 minute équilibration, initial mobile phase at 95 % aqueous NH4HCO3 / 5 % acetonitrile, gradient from 0 to 3 minutes to 95 % MeCN and held for 3 minutes, flow =1.5 mL/min).

443

Example 52: Préparation of (67?,127?)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyI)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-17- carboxylic acid, Compound 75

Step 1: (2Æ)-2-BenzyIoxy-2-(trifluoromethyI)hex-5-enehydrazide (hydrochloride sait)

To a 0 °C solution of (27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enehydrazide (3.672 g, 11.722 mmol) in diethyl ether (30 mL) was added dropwise a hydrogen chloride solution (8.8 mL of 2 M, 17.6 mmol) in diethyl ether. More diethyl ether (40 mL) was then added at 0 °C.

444

After the addition, the ice-water cooling bath was removed and the mixture was stirred at room température over 20 hours. The suspension was filtered on a fritted tunnel and the white precipitate was rinsed with diethyl ether (2X15 mL), collected and dried under vacuum to give as a white solid, (27?)-2-benzyloxy-2-(trifluoromethyl)hex-5-enehydrazide (hydrochloride sait) (3.71 g, 90 %). Ή NMR (400 MHz, DMSO-d₆) δ 10.95 (br s, 1H), 10.45 - 8.71 (m, 3H), 7.51 - 7.28 (m, 5H), 5.90 - 5.77 (m, 1H), 5.12 - 4.97 (m, 2H), 4.79 (s, 2H), 2.40 - 1.92 (m, 4H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -72.72 (s, 3F). ESI-MS m/z cale. 302.1242, found 303.1 (M+l)⁺; Rétention time: 2.83 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 6 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 2: Methyl 3-bromo-6-hydroxy-5-(trifluoromethyl)pyridme-2-carboxylate

l-Bromopyrrolidine-2,5-dione (18.3 g, 102.82 mmol) was added in three equal portions about 15 minutes apart to methyl 6-hydroxy-5-(trifluoromethyl)pyridine-2-carboxylate (20.6 g, 93.157 mmol) dissolved in DMF (160 mL) and the mixture was left stirring at room température 60 minutes after the addition was complété. Deionized water (1 L) was added and the resulting precipitate was filtered and washed with deionized water (3 X 100 mL) then dried under high vacuum to provide as a white powder, methyl 3-bromo-6-hydroxy-5-(trifluoromethyl)pyridine-2carboxylate (26.61 g, 95 %). Ή NMR (300 MHz, CDC1₃) ppm 4.03 (s, 3H), 7.98 (s, 1H), 11.53 (br. s., 1H). ¹⁹F NMR (282 MHz, CDCI3) ppm -66.4 (s, 3F). ESI-MS m/z cale. 298.9405, found 300.0 (M+l)⁺; Rétention time: 1.64 minutes. LCMS Method: Kinetex Polar Cis column (4.6 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate = 2.0 mL/min).

Step 3: Methyl 3-bromo-6-[(lÆ)-l-methylbut-3-enoxy]-5-(trifIuoromethyI)pyridine-2carboxylate

To a solution of methyl 3-bromo-6-hydroxy-5-(trifluoromethyl)pyridine-2-carboxylate (1 g, 3.3297 mmol) and (2S)-pent-4-en-2-ol (420 mg, 4.8762 mmol) in toluene (20 mL) was added triphenyl phosphine (1.3 g, 4.9564 mmol). After stirring at room température for 10 minutes,

445

DIAD (1.1297 g, 1.1 mL, 5.5868 mmol) was added and the mixture was stirred at room température for 2.5 h. Toluene was evaporated under reduced pressure. Purification by silica gel chromatography (40 g column, Gradient: 0 % to 5 % ethyl acetate in heptanes) yielded methyl 3bromo-6-[(l/?)-l-methylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carboxylatc (1.15 g, 94 %) as a light yellow oil. *H NMR (400 MHz, CDC1₃) δ 8.04 (s, 1H), 5.89 - 5.76 (m, 1H), 5.37 (m, J = 6.1 Hz, 1H), 5.15 - 5.04 (m, 2H), 4.00 (s, 3H), 2.55 - 2.38 (m, 2H), 1.36 (d, J = 6.1 Hz, 3H). ¹⁹F NMR (377 MHz, CDCI3) δ -64.41 (s, 3F). ESI-MS m/z cale. 367.0031, found 368.0 (M+l)⁺; Rétention time: 2.11 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow =1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.

Step 4: 3-Bromo-6-[(lE)-l-methylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carboxyIic acid

To a solution of methyl 3-bromo-6-[(lE)-l-methylbut-3-enoxy]-5(trifluoromethyl)pyridine-2-carboxylate (1.11 g, 3.0151 mmol) in MeOH (10 mL) and THF (10 mL) was added a solution of lithium hydroxide hydrate (550 mg, 13.107 mmol) in water (5 mL). The mixture was stirred at room température for 2 hours. To the mixture was added aqueous 1 N HCl until pH = 2 was reached. The volatiles were removed under reduced pressure and the product was extracted with ethyl acetate (3 X 40 mL). The combined organic phases were washed with brine (40 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure which gave as a yellow solid, 3-bromo-6-[(lA)-l-methylbut-3-enoxy]-5(trifluoromethyl)pyridine-2-carboxylic acid (1.1 g, 98 %). ’H NMR (400 MHz, CDCI3) δ 8.21 (s, 1H), 5.89 - 5.76 (m, 1H), 5.28 (m, J = 6.2 Hz, 1H), 5.19 - 5.09 (m, 2H), 2.61 - 2.43 (m, 2H), 1.43 (d, J = 6.1 Hz, 3H). ’⁹F NMR (377 MHz, CDCI3) δ -64.44 (s, 3F). ESI-MS m/z cale. 352.9874, found 354.1 (M+l)⁺; Rétention time: 2.02 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

446

Step 5: 7V’-[(22î)-2-benzyIoxy-2-(trifluoromethyl)hex-5-enoyl]-3-bromo-6-[(ll?)-l methylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carbohydrazide

To a solution of 3-bromo-6-[(12î)-l-methylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2carboxylic acid (1.709 g, 4.7619 mmol) and (27?)-2-benzyloxy-2-(trifluoromethyl)hex-5enehydrazide (hydrochloride sait) (1.7 g, 5.0184 mmol) in ethyl acetate (30 mL) was added triethylamine (1.8876 g, 2.6 mL, 18.654 mmol) and a solution of T3P (5.2 mL of 50 % w/v, 8.1714 mmol) in ethyl acetate. The reaction was stirred at room température (20 — 25 °C) for 72 hours. The mixture was quenched with an aqueous saturated solution of ammonium chloride (20 mL). Ethyl acetate (70 mL) was added to the mixture. The phases were separated and the organic phase was washed with an aqueous saturated solution of ammonium chloride (20 mL) and with an aqueous saturated solution of sodium bicarbonate (2 X 20 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to afford as a white solid, /V-[(27?)-2benzyloxy-2-(trifluoromethyl)hex-5-enoyl] -3 -bromo-6- [( 1 R)-1 -methylbut-3 -enoxy] -5(trifluoromethyl)pyridine-2-carbohydrazide (2.856 g, 94 %). ESI-MS m/z cale. 637.1011, found 638.1 (M+l)⁺; Rétention time: 2.14 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow =1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.

Step 6: 2-[(LR)-l-Benzyloxy-l-(trifluoromethyI)pent-4-enyl]-5-[3-bromo-6-[(l.Æ)-lmethylbut-3-enoxy]-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole

To a solution of7V-[(27î)-2-benzyloxy-2-(trifluoromethyl)hex-5-enoyl]-3-bromo-6-[(17?) l-methylbut-3-enoxy]-5-(trifluoromethyl)pyridine-2-carbohydrazide (1.49 g, 2.1753 mmol) in

1,2-dichloroethane (25 mL) andTVJV-diisopropylethylamine (2.59 g, 3.5 mL, 20.04 mmol) was

447 added toluenesulfonyl chloride (1.3 g, 6.8189 mmol). The reaction was then stirred at 50 °C for 22 h. The reaction was cooled down to room température and the volatiles were removed under reduced pressure. The crude residue was purified by reverse phase chromatography (50 gram Cis gel cartridge) using a gradient of 5 % - 100 % acetonitrile in water (+ 0.1 % v/v of formic acid in water). The fractions containing the product were concentrated under reduced pressure to afford as a yellow oil, 2-[(17?)-l-bcnzyloxy-l-(trifluoromethyl)pcnt-4-enyl]-5-[3-bromo-6-[(17?)-lmethylbut-3-enoxy]-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (1.3 g, 86 %). ESI-MS m/z cale. 619.0905, found 620.2 (M+l)⁺; Rétention time: 2.596 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 7: (67?,127î)-6-BenzyIoxy-17-bromo-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa 3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaene (E/Z mixture)

To a nitrogen purged solution of 2-[(lR)-l-benzyloxy-l-(trifluoromethyl)pent-4-enyl]-5[3-bromo-6-[(17?)-l-methylbut-3-enoxy]-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (525 mg, 0.7591 mmol) in dichloroethane (350 mL) at 60 °C was added a first portion of Zhan catalyst-lB (25 mg, 0.0341 mmol). After 1 hour, a second lot of Zhan catalyst-lB (25 mg, 0.0341 mmol) was added and heating was continued for another 1 h. DMSO (2 drops) was added and the reaction mixture was cooled down to room température. The volatiles were removed under reduced pressure. The crude material was purified by reversed-phase chromatography (Cis column, gradient: 0 % to 100 % acetonitrile in water containing 0.1 % formic acid). The fractions containing the product were concentrated under reduced pressure and lyophilized which gave as a tan solid, (6/?,12/?)-6-benzyloxy-17-bromo-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16hexaene (E/Z mixture) (210 mg, 47 %). ESI-MS m/z cale. 591.0592, found 592.0 (M+l)⁺; Rétention time: 2.31 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow =1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1% formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.

448

Step 8: Methyl (67î,122î)-6-benzyIoxy-12-methyl-6,15-bis(trifluoromethyI)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaene-17-carboxylate (E/Z mixture)

To a solution of (6R,12R)-6-benzyloxy-17-bromo-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaene (E/Z mixture) (200 mg, 0.3262 mmol) in anhydrous methanol (6 mL) in an autoclave was added Pd(dppf)Ch dichloromethane adduct (40 mg, 0.0482 mmol) and triethylamine (108.9 mg, 0.15 mL, 1.0762 mmol). The autoclave was purged with nitrogen, then with carbon monoxide. The mixture was heated to 100 °C and the carbon monoxide pressure was adjusted to 80 psi. The mixture was stirred for 17 h. The solution was cooled down to 25 °C, purged with nitrogen and concentrated under reduced pressure. The crude material was purified by silica gel chromatography (40 g column, gradient: 0 % to 20 % ethyl acetate in heptanes) to afford as a colorless oil, methyl (6R, 127?)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[ 12.3.1.12,5]nonadeca-1 (18),2,4,9,14,16-hexaene-l 7-carboxylate (E/Z mixture) (144 mg, 77 %). ESI-MS m/z cale. 571.1542, found 572.2 (M+l)⁺; Rétention time: 2.2 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow = 1.5 mL/min, run time - 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.

Step 9: Methyl (62?,121f)-6-hydroxy-12-methyI-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18 triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-17-carboxylate

To a solution of methyl (6R,12R)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaene-1Ίcarboxylate (E/Z mixture) (165 mg, 0.2887 mmol) in MeOH (8 mL) under nitrogen atmosphère was added palladium on carbon (62 mg, 10 % w/w, 0.0583 mmol). Hydrogen gas was bubbled in

449 for 5 minutes and the reaction was stirred at room température for 24 h. The mixture was purged with nitrogen, filtered over celite, washed with methanol (40 mL) and concentrated under reduced pressure to afford as a light brown oil, methyl (6/?,12/?)-6-hydroxy-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,14,16pentaene-17-carboxylate (121 mg, 85 %). *H NMR (400 MHz, DMSO-dô) δ 8.53 (s, 1H), 7.75 (s, 1H), 5.10 - 5.00 (m, 1H), 3.87 (s, 3H), 2.34 - 2.27 (m, 1H), 2.20 - 2.13 (m, 2H), 1.67 - 1.57 (m, 1H), 1.39 (d, J = 6.6 Hz, 9H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.63 (s, 3F), -76.86 (s, 3F). ESI-MS m/z cale. 483.1229, found 484.2 (M+l)⁺; Rétention time: 1.99 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow =1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.% CH3CN 0.1 % FA,. T: 45C, Flow: 1.5 mL/min.

Step 10: (6J?,12R)-6-Hydroxy-12-methyI-6,15-bis(trifluoromethyI)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-17-carboxylic acid, Compound 75

To a solution of methyl (6R,127?)-6-hydroxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaene-17-carboxylate (21 mg, 0.0434 mmol) in THF (0.75 mL) was added a solution of lithium hydroxide monohydrate (20 mg, 0.4766 mmol) in water (0.25 mL). The mixture was stirred at 0 °C for 15 minutes. To the mixture was added saturated aqueous ammonium chloride solution (2 mL) and aqueous 10 % citric acid until pH = 4 was reached. The product was extracted with ethyl acetate (3X15 mL). The combined organic phases were washed with brine (20 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. Purification of the residue by reversed-phase chromatography (15.5 g Cis column, gradient: 5 % to 100 % acetonitrile in water containing 0.1 % formic acid) afforded as a white solid after lyophilization, (6/?,12/?)-6-hydroxy-12-mcthyl6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadecal(18),2,4,14,16-pentaene-17-carboxylic acid (7.3 mg, 36 %). ’H NMR (400 MHz, DMSO-dô) δ 13.96 (br. s., 1H), 8.49 (s, 1H), 7.77 (s, 1H), 5.04 - 4.96 (m, 1H), 2.31 - 2.25 (m, 1H), 2.20 - 2.12 (m, 2H), 1.60 - 1.53 (m, 1H), 1.49 - 1.43 (m, 1 H), 1.41 - 1.29 (m, 8H). ¹⁹F NMR (377 MHz, DMSO-dô) δ -62.63 (s, 3F), -77.03 (s, 3F). ESI-MS m/z cale. 469.1072, found 468.1 (M-l)';

450

Rétention time: 4.69 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 μκη particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Example 53: Préparation of (6R,12R)-17-(hydroxymethyl)-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16pentaen-6-ol, Compound 76

Step 1 : [(6R,12Æ)-6-Benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,9,14,16-hexaen-17-yl]methanol (E/Zmixture)

To a solution of (6R,12R)-6-benzyloxy-17-bromo-12-methyl-6,15-bis(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaene (E/Z mixture) (150 mg, 0.2532 mmol) in 1,4-dioxane (3 mL) was added tributylstannylmethanol (163 mg, 0.5077 mmol) and Pd(dppf)Cb dichloromethane adduct (30 mg, 0.0367 mmol). Nitrogen was bubbled in for 5 minutes. The tube was sealed and the reaction mixture was stirred at 95 °C for 16 hours. The volatiles were removed under reduced pressure and the residue was purified by silica gel chromatography (24 g column, gradient: 0 % to 20 % ethyl acetate in heptanes) which afforded as a white solid, [(67?,127?)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,9,14,16-hexaen-17-yl]methanol (E/Z mixture) (58 mg, 42 %). Ή NMR (400 MHz, CDC1₃) δ 8.08 (s, 1H), 7.36 - 7.28 (m, 5H), 5.67 5.57 (m, 1H), 5.54 - 5.44 (m, 1H), 5.07 - 4.93 (m, 2H), 4.92 - 4.82 (m, 1H), 4.81 - 4.72 (m, 2H), 4.05 (t, J = 7.5 Hz, 1H), 3.67 - 3.58 (m, 1H), 2.80 - 2.68 (m, 1H), 2.48 - 2.40 (m, 2H), 2.36 - 2.25 (m, 1H), 1.90-1.81 (m, 1H), 1.53 (s, 3H). ¹⁹F NMR (377 MHz, CDCI3) δ-63.81 (s, 3F),-73.92 (s, 3F). ESI-MS m/z cale. 543.1593, found 544.2 (M+l)⁺; Rétention time: 2.13 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow = 1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic

451 acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.

Step 2: (6R,12R)-17-(Hydroxymethyl)-12-methyI-6,15-bis(trifluoromethyl)-13,19-dioxa3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol, Compound 76

To a solution of [(61/,127/)-6-benzyloxy-12-methyl-6,15-bis(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1 ( 18),2,4,9,14,16-hexaen-17-yl]methanol (E/Z mixture) (69 mg, 0.1268 mmol) in methanol (5 mL) under nitrogen atmosphère was added palladium on carbon (30 mg, 10 % w/w, 0.0282 mmol). Nitrogen was bubbled in for 5 minutes followed by hydrogen gas bubbled in for 5 minutes and the reaction was stirred at room température for 16 hours. The mixture was purged with nitrogen gas, filtered over celite, washed with methanol (40 mL) and concentrated under reduced pressure. Purification by reversed-phase chromatography (15.5 g Cis column, Gradient: 5 % to 100 % acetonitrile in water containing 0.1 % formic acid) afforded as a white solid, (67/,12/?)-17-(hydroxymethyl)-12-methyl-6,15bis(trifluoromethyl)-13,19-dioxa-3,4,l 8-triazatricyclo[l 2.3.1.12,5]nonadeca-l (18),2,4,14,16pentaen-6-ol (19.7 mg, 33 %). Ή NMR (400 MHz, DMSO-d₆) δ 8.34 (s, 1H), 7.63 (s, 1H), 5.66 (t, J = 5.6 Hz, 1H), 5.06 - 4.94 (m, 3H), 2.48 - 2.43 (m, 1H), 2.31 - 2.21 (m, 1H), 2.16 - 2.08 (m, 1H), 1.77 - 1.63 (m, 2H), 1.53 - 1.42 (m, 4H), 1.40 (d, J = 6.4 Hz, 3H), 1.32 - 1.22 (m, 1H). ¹⁹F NMR (377 MHz, DMSO-d₆) δ -62.28 (s, 3F), -76.50 (s, 3F). ESI-MS m/z cale. 455.128, found 456.2 (M+l)⁺; Rétention time: 4.5 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow = 1.5 mL/min).

452

Example 54: Préparation of (12Æ)-17-amino-7,7-difluoro-12-methyl-15-(trifluoromethyl)13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 1), Compound 77

Step 2

Step 1: Methyl 6-[(lÆ)-l-methylpent-4-enoxy]-3-nitro-5-(trifluoromethyl)pyridine-2carboxylate

To a suspension of methyl 6-hydroxy-3-mtro-5-(trifluoromethyl)pyridine-2-carboxylate (5 g, 18.675 mmol), (25)-hex-5-en-2-ol (6.45 g, 46.559 mmol) and triphenylphosphine (10.2 g,

38.889 mmol) in toluene (150 mL) was added DIAD (9.5 g, 46.981 mmol) dropwise. The reaction mixture was stirred at 22 °C ovemight. Triphenylphosphine (1.95 g) was added to quench the excessive DIAD. The mixture was stirred at 22 °C for 15 min and concentrated on silica gel (50 g). Purification by silica gel chromatography (220 g column; gradient: 0 % to 10 % ‘BuOCHj in heptanes) afforded as a colorless oil, methyl 6-[(lÆ)-l-methylpent-4-enoxy]-3-nitro15 5-(trifluoromethyl)pyridine-2-carboxylate (6.32 g, 92 %, > 95 % ee). ’H NMR (400 MHz,

453

CDC1₃) δ 8.64 (s, 1H), 5.87 - 5.73 (m, 1H), 5.55 - 5.44 (m, 1H), 5.05 - 4.93 (m, 2H), 4.03 (s, 3H), 2.26 - 2.07 (m, 2H), 1.99 - 1.87 (m, 1H), 1.83 - 1.71 (m, 1H), 1.41 (d, J = 6.4 Hz, 3H). ¹⁹F NMR (377 MHz, CDCh) δ -64.58 (s, 3F). Rétention time: 4.82 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min). The measurement of the optical purity was carried out under the following conditions: Isocratic mode, Column: ChiralCel OJ-H, S/No: OJH0CE-MJ029, Flow rate 1 mL/min. Starting condition: 90 % hexanes + DEA: 10 %: (10 % IPA/Hexane), Run time =16 minutes, injection = 3 pl, Room température = 24°C, Wavelength: 215 + 280 nm.

Step 2: 6-[(l/?)-l-MethyIpent-4-enoxy]-3-nitro-5-(trifIuoromethyl)pyridine-2carbohydrazide

no₂ o no₂ o

To a solution of methyl 6-[(lÆ)-l-methylpent-4-enoxy]-3-nitro-5(trifluoromethyl)pyridine-2-carboxylate (660 mg, 1.8022 mmol) in methanol (14 mL) in a pressure tube was added hydrazine monohydrate (670 mg, 13.384 mmol). The pressure tube was flushed with nitrogen and sealed. The reaction mixture was stirred at 80 °C for 30 min and cooled to room température then concentrated under reduced pressure to about 5 g. The residue was purified by reversed-phase chromatography (100 g Cis column; gradient: 5 % to 80 % acetonitrile in water). The desired fractions were concentrated under reduced pressure at 30 °C/38 Torr until a white precipitate appeared. The residue was extracted with TluOCTh (3 X 30 mL). The combined organic layers were washed with brine (5 mL), dried with NaiSOi, filtered and concentrated under reduced pressure to give as a pale-yellow solid, 6-[(U?)-l-methylpent-4enoxy]-3-nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (550 mg, 85 %). ESI-MS m/z cale. 348.1045, found 267.2 (M-81)⁺; Rétention time: 4.1 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

454

Step 3: Ethyl 3,3-difluoro-2-hydroxy-pent-4-enoate

O OH

To a solution of 3-bromo-3,3-difluoro-prop-l-ene (4.8 g, 30.582 mmol) and ethyl 2oxoacetate in toluene (5.9 mL of 50 % w/v, 28.896 mmol) in DMF (60 mL) and water (18 mL) at 10 °C was added indium (7 g, 60.966 mmol, ground before use). The mixture was stirred at room température ovemight. Ice-water (200 mL) was added. The resulting mixture was stirred for 30 min, diluted with 'BuOCHa (100 mL), filtered through diatomaceous earth and the cake was washed with 'BuOŒL. The aqueous phase was back extracted with 'BuOCHa (2 X 100 mL). The combined organic layers were dried with Na2SO4, filtered and concentrated. The residue (5.8 g) was purified by silica gel flash chromatography (gradient from 50 % to 100 % dichloromethane/pentane) to afford as a colorless oil, ethyl 3,3-difluoro-2-hydroxy-pent-4-enoate (4.283 g, 82 %). *H NMR (400 MHz, CDC1₃) δ 6.10 - 5.93 (m, 1H), 5.82 - 5.72 (m, 1H), 5.58 (d, J = 11.0 Hz, 1H), 4.43 - 4.27 (m, 3H), 3.22 (d, J = 6.4 Hz, 1H), 1.34 (t, J = 7.1 Hz, 3H). ¹⁹F NMR (377 MHz, CDCI3) δ -107.00 to -107.85 (m, 1F), -108.89 to -109.75 (m, 1F). ESI-MS m/z cale. 180.0598, found 181.2 (M+l)⁺; Rétention time: 1.47 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate = 1.2 mL/min).

Step 4: Ethyl 2-benzyloxy-3,3-difluoro-pent-4-enoate o %

To a solution of ethyl 3,3-difluoro-2-hydroxy-pent-4-enoate (2.45 g, 11.832 mmol) in dichloromethane (19 mL) and anhydrous heptane (38 mL) at 0 °C was added benzyl 2,2,2trichloroacetimidate (7 g, 27.72 mmol). The mixture was stirred at 0 °C for 5 min and triflic acid (400 mg, 2.6653 mmol) was added dropwise, a large amount of white precipitate appeared. The mixture was allowed to slowly warm up to room température and stirred at room température (5 — 19 °C) ovemight and then cooled to 0 °C. Diluted with dichloromethane (50 mL) then saturated NaHCCfi (20 mL) was added. The two layers were separated and the aqueous layer was extracted with dichloromethane (2 X 30 mL). The combined organic layers were dried with Na2SC>4, filtered and concentrated under reduced pressure. Purification by silica gel chromatography (gradient from 0 % to 10 % ethyl acetate in heptanes) afforded ethyl 2benzyloxy-3,3-difluoro-pent-4-enoate as a clear oil (1.56 g, 46 %). ’H NMR (400 MHz, CDCI3)

455 δ 7.45 - 7.31 (m, 5Η), 6.16 - 6.00 (m, 1H), 5.80 - 5.71 (m, 1H), 5.56 (d, J = 11.2 Hz, 1H), 4.79 (d, J = 12.0 Hz, 1H), 4.61 (d, J = 12.0 Hz, 1H), 4.33 - 4.16 (m, 3H), 1.31 (t, J = 7.1 Hz, 3H). ¹⁹F NMR (377 MHz, CDC1₃) δ -102.91 to -103.81 (m, 1F), -106.39 to -107.27 (m, 1F). Rétention time: 4.4 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Step 5: 2-Benzyloxy-3,3-difIuoro-pent-4-enoic acid

To a solution of ethyl 2-benzyloxy-3,3-difluoro-pent-4-enoate (1.55 g, 5.735 mmol) in DCE (60 mL) was added trimethyltin hydroxide (1.88 g, 10.397 mmol). The mixture was stirred at 82 °C for 30 h and cooled to room température. Silica gel (8 g) was added. The mixture was concentrated under reduced pressure then purified by silica gel chromatography (gradient from 0 % to 15 % methanol in dichloromethane) to give as a colorless oil, 2-benzyloxy-3,3-difluoropent-4-enoic acid (1.14 g, 78 %). ’H NMR (400 MHz, CDCI3) δ 7.42 - 7.31 (m, 5H), 6.13 - 5.97 (m, 1H), 5.77 (dt, J = 17.4, 2.2 Hz, 1H), 5.58 (d, J = 11.0 Hz, 1H), 4.83 - 4.70 (m, 2H), 4.25 (t, J = 9.4 Hz, 1H). ¹⁹F NMR (377 MHz, CDCI3) δ -103.47 to -104.44 (m, 1F), -106.04 to -106.95 (m, 1F). Rétention time: 1.86 minutes. LCMS Method: Kinetex Polar Cis column (3.0 X 50 mm, 2.6 pm particle size, 3 minute run, 5 % to 95 % acetonitrile in water (0.1 % formic acid modifier), flow rate =1.2 mL/min).

Step 6: 7V’-(2-Benzyloxy-3,3-difluoro-pent-4-enoyl)-6-[(lR)-l-methylpent-4-enoxy]-3-nitro 5-(trifIuoromethyl)pyridine-2-carbohydrazide

To a solution of 2-benzyloxy-3,3-difluoro-pent-4-enoic acid (850 mg, 3.3338 mmol) in dichloromethane (24 mL) was added oxalyl chloride (549 mg, 0.3773 mL, 4.3254 mmol), followed by DMF (291 mg, 0.3083 mL, 3.9812 mmol) dropwise. The mixture was stirred at

456 room température for 2 h and concentrated under reduced pressure to about 14 mL. The residue was added over a period of 15 min to a solution of 6-[(17?)-l-methylpent-4-enoxy]-3-nitro-5(trifluoromethyl)pyridine-2-carbohydrazide (1.57 g, 4.3817 mmol) and DIPEA (1.75 g, 2.3585 mL, 13.54 mmol) in dichloromethane (24 mL) at 0 °C. The reaction mixture was stirred at room température for 1 h and cooled to 0 °C. Saturated NaHCCh (25 mL) was added. The mixture was extracted with dichloromethane (3 X 40 mL). The combined organic layers were dried with Na2SÛ4, filtered and concentrated under reduced pressure. Purification of the residue by reversed-phase chromatography (150 g Cis column; gradient: 5 % to 95 % acetonitrile in water) afforded as a pale-yellow oil, V-(2-benzyloxy-3,3-difluoro-pent-4-enoyl)-6-[(17?)-l-methylpent4-enoxy]-3-nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (1.32 g, 69 %). *H NMR (400 MHz, CDCh) δ 8.35 (s, 1H), 7.43 - 7.32 (m, 5H), 6.11 - 5.95 (m, 1H), 5.88 - 5.71 (m, 2H), 5.57 (d, J = 11.0 Hz, 1H), 5.50 - 5.39 (m, 1H), 5.05 - 4.94 (m, 2H), 4.86 - 4.76 (m, 2H), 4.23 (t, J = 9.8 Hz, 1H), 2.28 - 2.08 (m, 2H), 2.00 - 1.87 (m, 1H), 1.84 - 1.73 (m, 1H), 1.43 (d, J = 6.1 Hz, 3H). Two protons of hydrazide not observed. ¹⁹F NMR (377 MHz, CDCh) δ -64.54 (s, 3F), 104.49 to -105.35 (m, 1F), -106.38 to -107.26 (m, 1F). ESI-MS m/z cale. 572.1694, found 573.2 (M+l)⁺; Rétention time: 2.02 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow = 1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1% formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1% formic acid.

Step 7: 2-(l-Benzyloxy-2,2-difluoro-but-3-enyl)-5-[6-[(lÆ)-l-methylpent-4-enoxy]-3-nitro-5 (trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole

To a solution of V-(2-benzyloxy-3,3-difluoro-pent-4-enoyl)-6-[(12î)-l-methylpent-4enoxy]-3-nitro-5-(trifluoromethyl)pyridine-2-carbohydrazide (573 mg, 0.9739 mmol) in dichloromethane (17 mL) at 0 °C was added DIPEA (761 mg, 1.0256 mL, 5.8881 mmol), followed by trifluoromethanesulfonic anhydride (420 mg, 1.4886 mmol) dropwise. The mixture was stirred at 0 °C for 30 min. Morpholine (520 mg) was added to quench the reaction. The mixture was stirred at 0 °C for 5 min then saturated NaHCCh (20 mL) was added. The two layers were separated and the aqueous layer was extracted with dichloromethane (2 X 20 mL). The

457 combined organic layers were dried with Na₂SÛ4, filtered and concentrated under reduced pressure. Purification of the residue by silica gel chromatography (gradient: 0 % to 30 % ethyl acetate in heptanes) afforded as a pale-yellow oil, 2-(l-benzyloxy-2,2-difluoro-but-3-enyl)-5-[6[(lE)-l-methylpent-4-enoxy]-3-nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (430 mg, 80 %). ’H NMR (400 MHz, CDCh) δ 8.67 (s, 1H), 7.40 - 7.31 (m, 5H), 6.16-6.01 (m, 1H), 5.86 - 5.73 (m, 2H), 5.60 (d, J = 11.2 Hz, 1H), 5.57 - 5.48 (m, 1H), 5.07 (dd, J = 10.0, 7.6 Hz, 1H), 5.03 - 4.93 (m, 2H), 4.81 (d, J = 11.7 Hz, 1H), 4.60 (d, J = 11.7 Hz, 1H), 2.26 - 2.08 (m, 2H), 2.01 - 1.89 (m, 1H), 1.85 - 1.74 (m, 1H), 1.43 (d, J = 6.4 Hz, 3H). ’⁹F NMR (377 MHz, CDCI3) δ -64.62 (s, 3F), -103.56 to -104.43 (m, 1F), -107.41 to -108.26 (m, 1F). ESI-MS m/z cale.

554.1589, found 555.2 (M+l)⁺; Rétention time: 2.17 minutes. LCMS Method: Luna Cis column (50 X 3 mm, 3 pm particle size, température = 45 °C, flow =1.5 mL/min, run time = 2.5 minutes. Mobile phase conditions: Initial 95 % water + 0.1 % formic acid / 5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile + 0.1 % formic acid over 1.3 minutes then held for 1.2 minute at 95 % acetonitrile + 0.1 % formic acid.

Step 8: (127?)-6-benzyloxy-7,7-difluoro-12-methyI-17-nitro-15-(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16-hexaene (E/Z mixture, diastereomer 1)

E/Z mixture, diastereomer 1

A 1 liter dried flask was charged with 2-(l-benzyloxy-2,2-difluoro-but-3-enyl)-5-[6[(17?)-l-methylpent-4-enoxy]-3-nitro-5-(trifluoromethyl)-2-pyridyl]-l,3,4-oxadiazole (400 mg, 0.7185 mmol) and DCE (400 mL). The mixture was bubbled with nitrogen for 1 h and then stirred at 75 °C for 10 min. A solution of Grubbs 2nd génération catalyst (200 mg, 0.2356 mmol) in DCE (4 mL) was added quickly by a syringe. After the mixture was stirred at 75 °C for 15 min, a solution of Grubbs 2nd génération catalyst (80 mg, 0.0942 mmol) in DCE (2 mL) was added quickly by a syringe. The mixture was stirred at 75 °C for 40 min and cooled to room température. DMSO (0.3 mL) was added. The mixture was stirred at room température for 1 h and then concentrated on silica gel (2 g). Purification by silica gel chromatography (gradient: 0 % to 20 % ethyl acetate in heptanes) afforded a crude product (225 mg) which was further purified by reversed-phase chromatography (120 g Cis column; gradient: 5 % to 95 % acetonitrile in water). The desired fractions were collected and concentrated to remove most of

458 the acetonitrile. The residue was extracted with ethyl acetate (3 X 20 mL). The combined organic layers were dried with Na2SO4, filtered and concentrated under reduced pressure to give as a colorless oil, ( 12Z?)-6-benzyloxy-7,7-difluoro-12-methyl-17-nitro-15-(trifluoromethyl)-13,19dioxa-3,4,18-triazatricyclo[ 12.3.1.12,5]nonadeca-1(18),2,4,8,14,16-hexaene (E/Z mixture, diastereomer 1) (127 mg, 30 %). Note: only a single chiral outcome was obtained at the benzyl alcohol carbon, stereochemistry of that center unknown. ESI-MS m/z cale. 526.1276, found 527.2 (M+l)⁺; Rétention time: 4.96 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1% formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow =1.5 mL/min).

Step 9: (127?)-17-Amino-7,7-difluoro-12-methyI-15-(tnfluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 1), Compound 77

E/Z mixture, diastereomer 1

diastereomer 1

To a solution of (127?)-6-benzyloxy-7,7-difluoro-12-methyl-17-nitro-15(trifluoromethyl)-13,19-dioxa-3,4,18-triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,8,14,16hexaene (E/Z mixture, diastereomer 1) (250 mg, 0.4165 mmol) in methanol (6 mL) and ethyl acetate (2 mL) in a flask was added 10 % palladium on carbon (86 mg, 50 % wet, 5 % w/w, 0.0404 mmol). The mixture was cooled with ice-water bath. Air in the flask was replaced by nitrogen through vacuum twice. Nitrogen in the flask was replaced by hydrogen through vacuum five times. Ammonia in methanol (0.4 mL of 2 M, 0.8 mmol) was added by a syringe. The reaction mixture was stirred at room température for 20 h. The mixture was filtered through Celite, washing with ethyl acetate and the filtrate was concentrated under reduced pressure. Purification of the residue by silica gel chromatography under the same conditions two successive times (gradient: 0 % to 40 % ethyl acetate in heptanes) afforded as a pale-yellow solid, (127?)-17-amino-7,7-difluoro-12-methyl-15-(trifluoromethyl)-13,19-dioxa-3,4,18triazatricyclo[12.3.1.12,5]nonadeca-l(18),2,4,14,16-pentaen-6-ol (diastereomer 1) (76 mg, 44 %). ’H NMR (400 MHz, DMSO-d₆) δ 7.75 (s, 1H), 7.21 (d, J = 5.6 Hz, 1H), 6.27 (s, 2H), 5.24 5.15 (m, 1H), 5.01 - 4.90 (m, 1H), 2.32 - 2.09 (m, 3H), 2.04 - 1.87 (m, 1H), 1.75 - 1.62 (m, 1H), 1.59- 1.47 (m, 1H), 1.45 - 1.37 (m, 1H), 1.34 (d, J = 6.4 Hz, 3H), 1.32- 1.22 (m, 1H). ¹⁹FNMR

459 (377 MHz, DMSO-dô) δ -62.41 (s, 3F), -92.20 to -93.10 (m, 1F), -99.62 to -100.68 (m, 1F). ESIMS m/z cale. 408.1221, found 409.2 (M+l)⁺; Rétention time: 4.42 minutes. LCMS Method: SunFire Cis column (75 X 4.6 mm, 3.5 pm particle size, 6 minute run, mobile phase conditions: initial 95 % water + 0.1 % formic acid/5 % acetonitrile + 0.1 % formic acid, linear gradient to 95 % acetonitrile for 4 min, then held for 2 min at 95 % acetonitrile, température = 45 °C, flow = 1.5 mL/min).

Example 55: Bioactivity Assay

Ussing Chamber Assay of CFTR-mediated short-circuit currents

Ussing chamber experiments were performed using human bronchial épithélial (HBE) cells derived from CF subjects heterozygous for F508del and a minimal function CFTR mutation (F508del/MF-HBE) and cultured as previously described (Neuberger T, Burton B, Clark H, Van Goor F Methods Mol Biol 2011:741:39-54). After four days the apical media was removed, and the cells were grown at an air liquid interface for >14 days prior to use. This resulted in a monolayer of fully differentiated columnar cells that were ciliated, features that are characteristic of human bronchial airway epithelia.

To isolate the CFTR-mediated short-circuit (Isc) current, F508del/MF-HBE grown on Costar® Snapwell™ cell culture inserts were mounted in an Ussing chamber and the transepithelial Isc was measured under voltage-clamp recording conditions (Vhoid= 0 mV) at 37 °C. The basolateral solution contained (in mM) 145 NaCl, 0.83 K2HPO4,3.3 KH2PO4, 1.2 MgCh, 1.2 CaCb, 10 Glucose, 10 HEPES (pH adjusted to 7.4 with NaOH) and the apical solution contained (in mM) 145 NaGluconate, 1.2 MgCb, 1.2 CaCh, 10 glucose, 10 HEPES (pH adjusted to 7.4 with NaOH) and 30 pM amiloride to block the épithélial sodium channel. Forskolin (20 pM) was added to the apical surface to activate CFTR, followed by apical addition of a CFTR inhibitor cocktail consisting of BPO, GlyH-101 and CFTR inhibitor 172 (each at 20 pM final assay concentration) to specifîcally isolate CFTR currents. The CFTRmediated Isc (pA/cm²) for each condition was determined from the peak forskolin response to the steady-state current following inhibition.

Identification ofPotentiator Compounds

The activity of the CFTR potentiator compounds on the CFTR-mediated Isc was determined in Ussing chamber studies as described above. The F508del/MF-HBE cell cultures were incubated with the potentiator compounds at a range of concentrations in combination with 10 pM (145)-8-[3-(2-{dispiro[2.0.2.1]heptan-7-yl}ethoxy)-lH-pyrazol-l-yl]-12,12-dimethyl2X⁶-thia-3,9,ll,18,23-pentaazatetracyclo [17.3.1.111,14.05,10]tetracosa-l(22),5,7,9,19(23),20460 hexaene-2,2,4-trione for 18 — 24 hours at 37 °C and in the presence of 20% human sérum. The concentration of potentiator compounds and (145)-8-[3-(2-{dispiro[2.0.2.1]heptan-7-yl}ethoxy)1 H-pyrazol-1 -yl] -12,12-dimethyl-2X⁶-thia-3,9,11,18,23-pentaazatetracyclo

[17.3.1.111,14.05,10]tetracosa-l(22),5,7,9,19(23),20-hexaene-2,2,4-trione used during the 18-24 hours incubations was kept constant throughout the Ussing chamber measurement of the CFTRmediated Isc to ensure compounds were présent throughout the entire experiment. The efficacy and potency of the putative F508del potentiators was compared to that of the known Vertex potentiator, ivacaftor (TV-[2,4-bis(l,l-dimethylethyl)-5-hydroxyphenyl]-l,4-dihydro-4oxoquinoline-3-carboxamide).

Table 12 and Table 13 represent CFTR modulating activity for représentative compounds of the invention generated using the assay described in this example (EC50: +++ is < 500 nM; ++ is 500 nM — 1 μΜ; + is > 1 μΜ; and ND is “not determined in this assay”).

Table 12: Bioactivity

Comp. No.	Structure	ECso
1	-Tl ω O y—z > zA \ ¹ 0 ) z^/ / O'rt T 0 Tl ω	+
2	^F3C-An s V // OH N~N enantiomer 1	ND
3	^F3CY^n s ^î^^/°x^Vcf₃\\ // OH N-N enantiomer 2	+
4	O^^> f₃c^ A. L ³ >\| n η ^>k/^O\>cCF3 T \\ // OH NH₂ N-N

461

Comp. No.	Structure	ECso
5	0^ ^F3CY^n s ^^/°\Ur^CF3 T v // OH nh₂ n-n
6	O^X''^X'^/'^X1 ^F3Cy^_n s T v h 'oh nh₂ N-N	+
7	^CF3V^n ^JL/\Jç-cf₃ HCl I V U OH NH₂ N-N diastereomer pair 1
8	cf₃^A_m L n y I l!JL o_sJ-cf₃ •HCl I v U OH NH₂ N-N diastereomer pair 2
9	O o=s > ^F3^C\^N S-\^°^r^CF3nh2 νΓ^0Η Enantiomer 1
10	O o=s η F₃C^ U\ \ ³ n η L1 ° h^cf ₃X, V II ÔH NH₂ n-n Enantiomer 2	ND

462

Comp. No.	Structure	ECso
11	^CF3Y^n T \\ // OH nh₂ n-n
12	^cf=>A s upi T V // OH ^HGI nh₂ N-N
13	O^^^i ^CF--A < •HCl I '' <3H nh₂ n-n
14	CF₃^ Us U ³ >\| n η LU\/^OxU^^CF3 VOH nh₂ n-n
15	o II o=s^ il 7 T V U OH nh₂ n-n enantiomer 1
16	0 II O=S-\ ^F3CY^n^^ ^î^x^O\J^^CF3 T \\ H OH nh₂ n-n enantiomer 2	ND

463

Comp. No.	Structure	ECso
17	Ό --x ^CF’yk ( ^CF3 HCI । v OH ^HCI nh₂ n-n
18	H r\ LX Λ Γ ^F nh₂ n-n ^oh diastereomer pair
19	0« s η S o L-^CF3 J X v OH nh₂ n-n
20	Oz s η ^f’^c/» s kA,o, zk 1 X II OH nh₂ n-n
21	^F3CY^n X vV°xJrCF₃ T X // OH nh₂ n-n
22	^SA ^F3CY^n^k X^A/°\Jv^cf3 T X H OH nh₂ n-n

464

Comp. No.	Structure	ECso
23	θχ. /K S ^x--y ^f=^c'/n A T Il OH nh₂ n-n enantiomer 1	+
24	Οχ. /X S ^X--y kJk,^OxJt-^CF3 I V Il OH nh₂ n-n enantiomer 2	+
25	z O ^CF3-^ki k y \| kk^°xkv^CF3 ‘^HCI nh₂ n-n ^oh	+
26	_oA^yO ^cfYn Ayv-A •HCl T '' OH nh₂ n-n
27	jkk ---^y-N / T^xk>^Dx_rk''^CF3 nh₂ Vn ^oh enantiomer 1

465

Comp. No.	Structure	ECso
28	/ T^V°V^î ^CF3 nh₂ ^Xn-N ^oh enantiomer 2	+
29	F-iCL L Y^V°vA~^cf3 NH₂ OH diastereomer pair 1
30	^Ρ3θγ\ k Τ^θΥΤ ^CF3 nh₂ n-N ^oh diastereomer pair 2
31	F Jil ? ^CFanh₂ W ^oh enantiomer 1

466

Comp. No.	Structure	ECso
32	F --< ^Ρ’%Λν / T^V°V^'^CF3 nh₂ Vn ^ohenantiomer 2	+
33	F ~'Τ^Χ'^Ο·'Τ'^CFjnh₂ ft-N ^oh Diastereomer pair 1
34	cf₃. â. L ³ >\| n η _μρ\| [ \\ // OH ^HCI nh₂ n-n enantiomer 1
35	^CF3^xAm k Il 7 I ^lLA^-°x^-CF₃ Lipi T v U OH nh₂ N-N enantiomer 2
36	HO. ^F3^CY^N < Yv>4-^cf3 NH₂ N-n OH

467

Comp. No.	Structure	ECso
37	HO^ ^f=^c/n < ŸV/K nh₂ n-_n oh
38	~ D o-A^{D c}^n LY°Y^CF3 T V // OH nh₂ n-n
39	O m ω z—Z )-o σ r? Y'iO / °x/ o z</ _/ ' οΊ I o ω
40	f₃c^A_m oh L Y ? 1 \Y\x^OxYrCF₃ ΊΤΥ Yoh nh₂ n-n diastereomer 1
41	θ^ΥΊ ^F3cYï>m OH L Y Y ] ^liY^x⁰YvCF3 T \\ // OH NH₂ N-N diastereomer 2
42	θ+^γ°Η ^CF3yL_n < ‘YY/°x^X-CF₃ΊΓ\\ Ïoh nh₂ n-n diastereomer 1

468

Comp. No.	Structure	EC50
43	_Ξ OH ^CF3^A-_m k H ? 1 ^lL_iA^zO_sJr-CF3 T \\ // OH nh₂ N-N regioisomeric diastereomer 1
44	₀A^D ^0Ρ3γΑ_Ν dA VL°vA^CF3 \| \\ // OH nh₂ n-n
45	□A^oh ^CF3Y^n ^L^k/O.J-CF₃T \\ V OH nh₂ n-n diastereomer 2
46	= OH cf₃^L· k Il 7 l ^Îk^^/^o\^rCF₃ T \\ Il OH nh₂ n-n diastereomer 2
47	O^^^N ^F=^CvAn L ŸïU^CFinh₂ n-_n oh enantiomer 1

469

Comp. No.	Structure	ECso
48	L Lcf₃I » /XV ³NH₂ N-n ^oh enantiomer 2
49	^Ρ'Ί^Ί Ux^O LcF₃ NH₂ N-n OH enantiomer 1
50	XX ^ρ3θγ^Ν < V^Oy^CFa NH₂ N-n OH enantiomer 2
51	Υύ⁰^^0Ρ3 NH₂ N-n oh enantiomer 1

470

Comp. No.	Structure	ECso
52	CX y^°U-cF₃nh₂ n-n ^oh enantiomer 2
53	CF₃. A X\|N HO η ^Lî^/^OnAvCF₃ T \\ // OH nh₂ N-N diastereomer 1

Table 13: Bioactivity

Comp. No.	Structure	ECso
54	o^A ^cf3xA_m k kk/^Okr^CF3 T V // OH nh₂ n-n enantiomer 1	ND
55	qAA ^cf-A S kxk/°xkc^CF3 kk\^ Ô'Sh nh₂ n-n enantiomer 2	ND

471

Comp. No.	Structure	EC₅o
56	^CF3^4k, k^OH H ? 1 «OH nh₂ n-n enantiomer 1	+
57	^CF3^A_n v^h T v // OH nh₂ n-n enantiomer 2
58	CF_a. A. X >\| N HO η T V // OH nh₂ n-n diastereomer 2
59	o—°> ^CF3Y^n s L^k/OxA-CF₃ T \\ V OH NH₂ N-N	+
60	O^^^U CFq. Us ³>\| N T \\ // OH nh₂ n-n	ND
61	^CF3-^n^{Z X} k H ? i Χ^Κ/°\Χγ^0Ρ3 T V II OH nh₂ n-n

472

Comp. No.	Structure	ECso
62	^CF3xX_n <λΟΗ nh₂ N-N OH enantiomer 3
63	s NH₂ N-N OH enantiomer 1
64	CF-,χ Jx \ ³>^n η ^JkxO^i-cFa T X nh₂ n-n oh enantiomer 2
65	CF-x Jx X ³>^N N T\i^oy^<^CF3 \^NH N-N OH enantiomer 1	ND
66	^cf-A > T^'^CV''\^CF3χ^,ΝΗ N-N OH enantiomer 2	ND

473

Comp. No.	Structure	ECso
67	T^X°z<\^CF3 NH₂ N-N ^oh	ND
68	CF_?. \ ³>^n b VSn°>^^cf3 nh₂ n-n OH enantiomer 1	ND
69	^CF’>An b kA^°ybçCF₃nh₂ n-n OH enantiomer 2	ND
70	^F3CY^n <b ^7 OH nh₂ n-n diastereomer 1	ND
71	FgC^A-M O. H ? Ί X^^/°\JrCF₃ T \\ // OH nh₂ n-n diastereomer 2	ND

474

Comp. No.	Structure	ECso
72	^f’^cŸ*_n < L /°_K A- cf₃ V(DH NH₂ N-N diastereomer 1	ND
73	o'^x^^^x'o ^F3CyL_n k 1¾ KoH NH₂ N-N diastereomer 2	ND
74	^F3^C^A_M XX o x> Tvr^N NH₂ N-N	ND
75	^f’^cX>n s k^k/\A-CF₃ T \\ // OH X. N-N HO^O	ND
76	^f3C^X. k H ? I ^s^k^OxA-CFa T \\ // OH > N-N HO	ND
77	F₃C. A, k/^F ³ >\| N ^f-F Tv ^0H NH₂ N-N diastereomer 1	ND

475

Other Embodiments

The foregoing discussion discloses and describes merely exemplary embodiments of this disclosure. One skilled in the art will readily recognize from such discussion and from the accompanying drawings and claims, that various changes, modifications and variations can be made therein without departing from the spirit and scope of this disclosure as defined in the following claims.

Claims

1. A compound selected from compounds of Formula I:

( B J each Y is independently selected from-C(R^Y )2-,-O-,-CO-, and ' f ;

each R is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, Cô-Cio aryl, 5- to 10-membered heteroaryl, -OR^Y1, -CO2R^YI, -COR^Y1, -CON(R^Y1)2, and -NR^YI-; or two instances of R^Yon the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3to 6-membered heterocyclyl; or two instances of R\ one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R'¹ is independently selected from hydrogen and Ci-Cô alkyl, or two instances of R'¹ bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Cj-Cô alkyl, and Ci-Cô alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

each Q is independently selected from:

Cj-Cô alkyl optionally substituted with 1-3 groups independently selected from: o halogen, o oxo, o Cô-Cjo aryl (optionally substituted with 1-3 groups independently selected from halogen and -OCF3), and

477 o C3-C8 cycloalkyl.

C₃-Cs cycloalkyl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o C|-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH₂, and -NHCOMe), o Cj-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Ci-Cô alkyl), and o C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from: o halogen, .

o CN, .

o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen and hydroxy), o C]-Cô alkoxy optionally substituted with 1-4 groups independently selected from:

halogen,

C3-C8 cycloalkyl (optionally substituted with CF3), o C3-C8 cycloalkyl (optionally substituted with 1-3 groups independently selected from halogen, CF3, OCF3, and Ci-Cô alkyl), and o C₆-Cio aryl,

478 each R is independently selected from halogen, Cj-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -OR², -N(R²)2, CO2R², -CO-N(R²)2, -CN. phenyl, benzyl, C|-Cô alkoxy, C3-C8 cycloalkyl, 5- to 6membered heteroaryl, 3- to 6-membered heterocyclyl, -SO₂R², -SR², -SOR², -PO(OR²)2, and -PO(R²)2;

each R² is independently selected from hydrogen, Ci-Cô alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cio aryl (optionally substituted with Cj-Cô alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen);

'C R^Z1 R^Z3 C ^c J/^-(R^Z2)o-2 . -Xr⁷² V^Si'R^Z3 V^R^Z1

Z is selected from , \ , and ; wherein

Ring C is selected from Cô-Cio aryl and 5- to 10-membered heteroaryl;

71 72

R is selected from hydrogen, halogen, and hydroxy, or R and R taken together form a group selected from oxo and =N-OH;

each R is independently selected from hydroxy, Ci-Cô alkoxy, C]-Cô alkyl, Ci-Cô haloalkyl, and Cô-Cio aryl; or two instances of R²³ are taken together to form a 3- to 6membered heterocyclyl;

n is selected from 4, 5, 6, 7, and 8; and m is selected from 0, 1,2, and 3.

2. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to claim 1, wherein X is -O-.

3. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to claim 1 or claim 2, wherein each R' is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, and -OR^YI.

4. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 3, wherein -OR'¹ is -OH.

5. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 4, wherein each Q is independently selected from:

479

6.

C₃-C₈ cycloalkyl.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 5, wherein each Q is independently selected from:

7.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 6, wherein each R' is independently selected from:

CH₃ OH hydrogen, fluorine, —L~ , -J— ,

8.

9.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 7, wherein Ring B is selected from C₃-C₈ cycloalkyl and phenyl optionally substituted with 1-3 groups independently selected from halogen.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 8, wherein Ring B is selected from:

10.

11.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 9, wherein n is selected from 4, 5, and 6.

The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 10, wherein -(Y)_n- is a group selected from:

480

481

12. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 11, wherein each R¹ is independently selected from Cj-Cô alkyl 5 (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -N(R²)₂, and -CO₂R².

13. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 12, wherein each R² is independently selected from hydrogen and Cj-Cô alkyl.

10 14. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 13, wherein each R¹ is independently selected from -CF3, -NH₂, NH(CH₂CH₃), CO₂H, and CH₂OH.

15. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to

X _RZ1 £ C 4-(^)0-2

Y---⁷ any one of daims 1 to 14, wherein Z is selected from and .

15 16. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 15, wherein the group:

£c is selected from:

482

17. The compound. deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 16, wherein the group:

18. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 17, wherein R^Z1 is selected from hydrogen and Ci-Cô alkyl (optionally substituted with 1-3 groups selected from halogen).

19. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 18, wherein R^ZI is selected from hydrogen and -CF3.

20. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 19, wherein R^Z2 is hydroxy.

21. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 20, wherein Z is selected from:

22. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 21, wherein m is selected from 1 and 2.

23. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 22, wherein:

X is -O-;

each Y is independently selected from -C(R^Y)2-, -O-, and each R is independently selected from hydrogen and Ci-Cô alkyl (optionally substituted with 1 -3 groups independently selected from hydroxy and Q);

Ring B is selected from C₃-Cg cycloalkyl groups:

each Q is independently selected from:C3-Cs cycloalkyl and Cô-Cio aryl optionally substituted with 1-3 groups independently selected from halogen and Cj-Cô alkyl,

483 each R is independently selected from Cj-Cf, alkyl (optionally substituted with 1-6 groups independently selected from halogen) and -NH2;

R^/J is selected from Cj-Ct, alkyl (optionally substituted with 1-6 groups independently selected from halogen);

R^Z2 is hydroxy;

n is selected from 5 and 6; and m is 2.

24. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 23, wherein each Q is independently selected from:

25. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 24, wherein each R is independently selected from:

26. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 25, wherein Ring B is

27. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 26, wherein -(Y)_n- is a group selected from:

484

5 28. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 27, wherein R^Z1 is -CF3.

29. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 28, wherein n is 5.

30. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to 10 any one of daims 1 to 29, wherein n is 6.

31. A compound selected from compounds of Table 12, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing.

32. A compound selected from compounds of Table 13, pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing.

15 33. A compound according to claim 31, wherein the compound is selected from:

Comp. No. Structure

485

486

Comp. No.

Structure

enantiomer 1

enantiomer 2

enantiomer 2 pharmaceutically acceptable salts thereof, and deuterated dérivatives of any of the foregoing.

34.

A compound according to claim 32, wherein the compound is selected from:

487

Comp. No. Structure

61 oAA Mn i T X // OH NH₂ N-n 9

63 CF·,^ A \ ³ Yn > Τ^θΧ'χ³nh₂ n-n OH enantiomer 1

64 ^cf3-An s ΧΧγ°\ΧςθΡ₃nh₂ n-n OH enantiomer 2 9

488

40. The pharmaceutical composition according to any one of daims 36 to 39, wherein the one or more additional therapeutic agent(s) comprise(s) 3-(6-(1-(2,2difluorobenzo[d][l,3]dioxol-5-yl)cyclopropanecarboxamido)-3-methylpyridin-2-

5 yl)benzoic acid (Compound IV):

41. The pharmaceutical composition according to any one of daims 36 to 40, wherein the one or more additional therapeutic agent(s) comprise(s) 7V-(l,3-dimethylpyrazol-4yl)sulfonyl-6-[3-(3,3,3-trifluoro-2,2-dimethyl-propoxy)pyrazol-l-yl]-2-[(4S)-2,2,4-

10 trimethylpyrrolidin-1 -yl]pyridine-3-carboxamide (Compound V):

42. The pharmaceutical composition according to any one of daims 36 to 41, wherein the one or more additional therapeutic agent(s) comprise(s) 7V-(benzenesulfonyl)-6-[3-[2-[l(trifluoromethyl) cyclopropyl]ethoxy]pyrazol-l-yl]-2-[(4S)-2,2,4-trimethylpyrrolidin-l-

15 yl]pyridine-3-carboxamide (Compound VI):

43. The pharmaceutical composition according to any one of daims 36 to 42, wherein the one or more additional therapeutic agent(s) comprise(s) (145)-8-(3-(2{dispiro[2.0.2.1]heptan-7-yl}ethoxy)-17/-pyrazol-l-yl]-12,12-dimethyl-2L⁶-thia-

489

3,9,11,18,23-pentaazatetracyclo [17.3.1.111,14.05.10]tetracosa-1(22),5,7,9,19(23),20hexaene-2,2,4-trione (Compound VII):

44. The pharmaceutical composition according to any one of daims 36 to 43, wherein the one or more additional therapeutic agent(s) comprise(s) (117?)-6-(2,6-dimethylphenyl)11 -(2-methylpropyl)-12- { spiro [2.3]hexan-5-yl} -9-oxa-2Z⁶-thia-3,5,12,19tetraazatricyclo [ 12.3.1.14,8] nonadeca-1 ( 17),4( 19),5,7,14( 18), 15-hexaene-2,2,13 -tri one (Compound VIII):

45. The pharmaceutical composition according to any one of daims 36 to 44, wherein the one or more additional therapeutic agent(s) comprise(s) at least one compound selected from PTI-428, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and PTI-801.

46. The pharmaceutical composition according to any one of daims 36 to 45, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR potentiator enhancer.

47. The pharmaceutical composition according to any one of daims 36 to 46, wherein the one or more additional therapeutic agent(s) comprise(s) ASP-11.

48. The compound, deuterated dérivative, or pharmaceutically acceptable sait according to any one of daims 1 to 34 or the pharmaceutical composition according to any one of daims 35 to 47 for use in the treatment of cystic fïbrosis.

49. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to claim 48, wherein the use further comprises administering one or more additional therapeutic agent(s).

50. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to claim 49, wherein the one or more

490 additional therapeutic agent(s) comprise(s) a compound with CFTR modulating activity or a sait or deuterated dérivative thereof.

51. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to claim 49 or 50, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR corrector.

52. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 51, wherein the one or more additional therapeutic agent(s) comprise(s) (7?)-l-(2,2- difluorobenzo[d][l,3]dioxol-5-yl)-V-(l-(2,3-dihydroxypropyl)-6-fluoro-2-(l-hydroxy-2methylpropan-2-yl)-l 7/-indol-5-yl)cyclopropanecarboxamide (Compound II):

53. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 52, wherein the one or more additional therapeutic agent(s) comprise(s) 3-(6-(1-(2,2difluorobenzo [d] [ 1,3] dioxol-5-yl)cyclopropanecarboxamido)-3 -methylpyridin-2yl)benzoic acid (Compound IV):

54. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 53, wherein the one or more additional therapeutic agent(s) comprise(s) V-(l,3-dimethylpyrazol-4yl)sulfonyl-6-[3-(3,3,3-trifluoro-2,2-dimethyl-propoxy)pyrazol-l-yl]-2-[(4<S)-2,2,4trimethylpyrrolidin-l-yl]pyridine-3-carboxamide (Compound V):

491

55. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 54, wherein the one or more additional therapeutic agent(s) comprise(s) A^r-(benzenesulfonyl)-6-[3-[2-[l(trifluoromethyl) cyclopropyl]ethoxy]pyrazol-l-yl]-2-[(4S)-2,2,4-trimethylpyrrolidin-l-

5 yl]pyridine-3-carboxamide (Compound VI):

56. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 55, wherein the one or more additional therapeutic agent(s) comprise(s) (14S)-8-[3-(2-

10 {dispiro[2.0.2.1 ]heptan-7-yl} ethoxy)-1 77-pyrazol-1 -yl] -12,12-dimethyl-2X⁶-thia3,9,11,18,23-pentaazatetracyclo [17.3.1.111,14.05,10]tetracosa-l(22),5,7,9,19(23),20hexaene-2,2,4-trione (Compound VII):

57. The compound, deuterated dérivative, pharmaceutically acceptable sait, or

15 pharmaceutical composition for use according to any one of daims 49 to 56, wherein the one or more additional therapeutic agent(s) comprise(s) (1172)-6-(2,6-dimethylphenyl)1 l-(2-methylpropyl)-12-{spiro[2.3]hexan-5-yl}-9-oxa-2Z⁶-thia-3,5,12,19tetraazatricyclo[12.3.1.14,8]nonadeca-l (17),4(19),5,7,14(18),15-hexaene-2,2,13-trione (Compound VIII):

492

58. The compound. deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 57, wherein the one or more additional therapeutic agent(s) comprise(s) at least one compound selected from PTI-428, ABBV-2222, ABBV-2851, GLPG2737, ABBV-3221, ABBV-3748, ABBV-3903, ABBV-119, and PTI-801.

59. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 58, wherein the one or more additional therapeutic agent(s) comprise(s) a CFTR potentiator enhancer.

60. The compound, deuterated dérivative, pharmaceutically acceptable sait, or pharmaceutical composition for use according to any one of daims 49 to 59, wherein the one or more additional therapeutic agent(s) comprise(s) ASP-11.

61. Use of the compound, deuterated dérivative, or pharmaceutically acceptable sait of any one of daims 1 to 34 or the pharmaceutical composition according to any one of daims 35 to 47 in the manufacture of a médicament for the treatment of cystic fibrosis.

62. A process for preparing a compound of Formula S4-4

comprising converting a compound of Formula S4-3:

into a compound of Formula S4-4, wherein:

ί B J each Y is independently selected from -C(R' )2-, -O-, -CO-, and ' ' ;

each R^x is independently selected from hydrogen, halogen, Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from hydroxy and Q), C3-C8 cycloalkyl, Cô-Cio aryl, 5- to 10-membered heteroaryl, -OR^Y1, -CO2R^Y1, -COR^Y1, -CON(R^Y1)2, and -NR^Y1-; or two instances of R^Y

493 on the same atom are taken together to form a ring selected from C3-C8 cycloalkyl and 3to 6-membered heterocyclyl; or two instances of R\ one of which is on one atom and the second of which is on an adjacent atom, are taken together to form a pi bond;

each R'¹ is independently selected from hydrogen and Ci-C₆ alkyl, or two instances of R'¹ bonded to the same nitrogen taken together form a 3- to 6-membered heterocyclyl;

Ring B is selected from:

Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen, Ci-Cô alkyl, and C]-Cô alkoxy),

C3-C8 cycloalkyl,

5- to 10-membered heteroaryl, and

3- to 6-membered heterocyclyl (optionally substituted with 1-3 groups independently selected from C]-Cô alkyl);

each Q is independently selected from:

Ci-Cô alkyl optionally substituted with 1-3 groups independently selected from:

o halogen, o oxo, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from halogen and -OCF3), and o C3-C8 cycloalkyl,

C₃-Cs cycloalkyl optionally substituted with 1-3 groups independently selected from: o halogen, o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen, -NH2, and -NHCOMe), o Ci-Cô alkoxy, o Cô-Cio aryl (optionally substituted with 1-3 groups independently selected from Cj-Cô alkyl), and o C3-C8 cycloalkyl,

Cô-Cio aryl optionally substituted with 1-3 groups independently selected from:

o halogen, o CN, o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen and hydroxy), o Ci-Cô alkoxy optionally substituted with 1-4 groups independently selected from: halogen,

494

C₃-C₈ cycloalkyl (optionally substituted with CF₃), o C₃-C₈ cycloalkyl (optionally substituted with 1-3 groups independently selected from halogen, CF₃, OCF₃, and Ci-C₆ alkyl), and o C₆-C₁₀ aryl,

o halogen, o C|-Cô alkyl (optionally substituted with 1-3 groups independently selected from halogen), o C₃-C₈ cycloalkyl (optionally substituted with 1-3 CF₃ groups), and o 3-to 10-membered heterocyclyl,

o Ci-Cô alkyl (optionally substituted with 1-3 groups independently selected from oxo and C₃-C₈ cycloalkyl), and o oxo;

each R* is independently selected from halogen, CpCe alkyl (optionally substituted with 1-6 groups independently selected from halogen and hydroxy), -OR², -N(R²)3, -CO2R², CO-N(R²)2, -CN, phenyl, benzyl, Ci-Cg alkoxy, C3-C₈ cycloalkyl, 5- to 6-membered heteroaryl, 3- to 6-membered heterocyclyl, -SO2R², -SR², -SOR², -PO(OR²)2, and PO(R²)₂;

each R² is independently selected from hydrogen, Ci-Ce alkyl (optionally substituted with 1-6 groups independently selected from halogen), and Cô-Cjo aryl (optionally substituted with Ci-Ce alkoxy, which is optionally substituted with 1-6 groups independently selected from halogen); and m is selected from 0, 1,2, and 3.

63. A compound selected from:

substantially crystalline Compound 11 heptane solvaté, substantially crystalline Compound 6 (free form), substantially crystalline Compound 19 (free form), and substantially crystalline Compound 20 (free form).