EA045973B1 - TRIAZACYCLODODECANESULFONAMIDE (“TCD”) PROTEIN SECRETION INHIBITORS - Google Patents

Description

State of the art Field of technology

The present invention relates to triazacyclododecanesulfonamide (TCD) protein secretion inhibitors, including methods for their preparation and use.

Incorporation of material submitted electronically by reference

The present invention contains as a separate part a sequence listing in machine-readable form (file name: 40056_SeqListing.txt; 20275 bytes; created February 14, 2019), the contents of which are incorporated by reference in their entirety.

Description of related technology

Protein translocation into the endoplasmic reticulum (ER) is the first step in protein secretion. Protein import into the ER is key for all eukaryotic cells and is especially important for rapidly growing tumor cells. Thus, the protein secretion process may serve as a target for both possible anticancer drugs and bacterial virulence factors. See Kalies and Romisch, Traffic, 16(10):1027–1038 (2015).

Protein transport into the ER begins in the cytosol when N-terminal hydrophobic signal peptides are exposed to the surface of the ribosome. Binding of a signal recognition particle (SRP) to a signal sequence targets the ribosome-nascent chain-SRP complex to the ER membrane, where contact of SRP with its receptor triggers signal peptide transmission to Sec61. Sec61 is an ER membrane protein translocator (called translocon) that is donut-shaped with three main subunits (heterotrimeric). It involves a traffic jam that blocks transport into or out of the ER. The plug is displaced when the hydrophobic region of the growing polypeptide interacts with the Sec61 seam region, allowing translocation of the polypeptide into the lumen of the ER. In mammals, only short proteins (<160 amino acids) can enter the ER after translation, and proteins composed of <120 amino acids inevitably use this route. The ability to translocation is maintained to some extent by the binding of calmodulin to the signal sequence. Upon entry into the Sec61 channel, the signal peptide or signal anchor peptide is inserted between transmembrane domains (TMDs) 2 and 7 of Sec61a, which form the side of the gate, thereby allowing the channel to open for soluble secretory proteins. Because the Sec61 channel consists of 10 TMDs (Sec61a) surrounded by a hydrophobic vice formed by Sec6fy, channel opening is dependent on conformational changes that affect virtually all TMDs.

Inhibiting protein transport across the ER membrane has the potential to treat or prevent diseases such as cancer cell growth and inflammation. Known secretion inhibitors, ranging from broad spectrum inhibitors to highly substrate specific inhibitors, can disrupt virtually any step of this multistep process and even the transport of endocytosed antigens into the cytosol for cross-presentation. These inhibitors interact with signal peptide, chaperones, or the Sec61 channel to block substrate binding or prevent conformational changes required for protein import into the ER. Examples of protein secretion inhibitors include calmodulin inhibitors (eg, E6 berbamine and ophiobolin A), lanthanum, sterols, cyclodepsipeptides (eg, HUN-7293, CAM741, NFI028, cotransine, apratoxin A, decatransin, valinomycin), CADA, mycolactone, Eeyarestatin I ( ESI) and exotoxin A. However, the above secretion inhibitors suffer from one or more of the following: lack of selectivity for the Sec61 channel, difficulty in preparation due to complex structure, and molecular weight that imposes limitations on administration, bioavailability and distribution.

Thus, there is a need for new small molecule inhibitors of protein secretion.

Brief description of the invention

In one aspect, provided herein is a compound of formula (I) or a pharmaceutically acceptable salt thereof:

where each of R ^a and R ^b independently represents H or C _1-3 alkyl;

R ¹ is H, OH, C1-3alkyl, CC1-3alkyl, =CH2 or = ^NOR5 or ^R1 is C3-6cycloalkyl or _C3-6 heterocycloalkyl and forms a spiro group together with the carbon atom in the ring to which it is attached ;

R ^1a and R ^1b are each independently H or C _1-3 alkyl;

R ² represents C1-6 alkyl, N(R ⁵ )2, C _3-8 cycloalkyl, C _3-9 heterocycloalkyl, C _3-9 heterocycloalkenyl or C _6-10 aryl;

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R ³ represents H, C 1-6 alkyl, C _3-8 cycloalkyl, C _3-8 cycloalkenyl, C _3-7 heterocycloalkyl, C _6-10 aryl or C _2-6 heteroaryl;

R ⁴ represents C _3-8 cycloalkyl, C _3-9 heterocycloalkyl, C _6-10 aryl or C _2-9 heteroaryl;

each R ⁵ independently represents H, C _1-3 alkyl or C _0-2 alkylene-C _6-10 aryl;

X is absent or represents C _1-3 alkylene, C=O or (C=O)O;

Y is SO or SO2;

each heterocycloalkyl, heterocycloalkenyl and heteroaryl group independently contains 1, 2 or 3 heteroatoms on the ring selected from N, O and S.

In some embodiments, each Y is an SO. In various embodiments, each Y is SO2.

In various cases, R ^a is H. In some cases, R ^a is C1-3 alkyl. In some embodiments, R ^a is CH3. In some embodiments, R ^b is H. In various embodiments, R ^b is C1-3 alkyl. In various cases, R ^b represents CH3.

In different cases, R ^a and R ^b each represent H.

In some cases, R ¹ represents H, OH or =NOR ⁵ . In various cases, each R ⁵ independently represents H, €1.3alkyl or C0-2alkylene-C6-10aryl. In some cases, R ⁵ is H or CH3. In some embodiments, R ¹ is C1-3alkyl or OC1-3alkyl. In various embodiments, R ¹ is CH3 or OCH ₃ . In some cases, R ¹ is CH3 and has S stereochemistry. In various embodiments, R ¹ is C3-6 cycloalkyl or C _3-6 heterocycloalkyl and forms a spiro group together with the carbon atom on the ring to which it is attached. In some cases, R ^1, together with the atom in the ring to which it is attached, forms or In other cases, R ¹ represents =CH ₂ .

In some cases, each of R ^1a and R ^1b is H. In some cases, at least one of R ^1a and R ^1b is C1-3 alkyl. In some cases, R ^1a and R ^1b are each CH3.

In some embodiments, R ² is C1-6 alkyl or N(R ⁵ )2. In various cases, each R ⁵ independently contains H, C1-3 alkyl or benzyl. In various embodiments, R ² represents Et, iPr, N(CH3) ₂ , ^λ J A or . In some cases

R ² contains C3-8cycloalkyl. In various cases, R ² contains cyclopentyl or cyclohexyl. In some embodiments, R ² is C3-9 heterocycloalkyl or C _3-9 heterocycloalkenyl. In various embodiments, R ² contains oxetanyl, azetidinyl, tetrahydrofuranyl, pyrrolidinyl, tetrahydropyranyl, pyranyl, piperidinyl, piperazinyl, azepanyl, morpholinyl, or tetrahydropyridinyl. In various embodiments, the C _3-9 heterocycloalkyl or C _3-9 heterocycloalkenyl contains a bridging or spiro group. In some cases, the C _3-9 heterocycloalkyl containing a bridged or spiro group is selected from the group consisting of *. · both A and R6 represent

C _1-6 alkyl, C _3-8 cycloalkyl or C _6-10 aryl. In some cases, R ⁶ is phenyl, optionally substituted with one to three groups independently selected from halogen, C _1-3 alkyl, C _1-3 alkoxy and CN. In different cases, R ² is selected from the group consisting of

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group consisting of

CI •

In various cases, R ² represents C _6-10 aryl. In some embodiments ^{, R2} represents In some cases, it is chosen to be itself or ^0MV . In different implementations, X is missing. In some embodiments, R ³ contains C1-6 alkyl or H. In some cases, R ³ is 2-methylbutyl, isopropyl, isopentyl, CH2CH ₂ OCH ₃ , CH ₂ C(CH ₃ ) ₂ CN, CH ₂ CF ₃ or CH ₂ CH ₂ CF ₃ . In various cases, R ³ is isobutyl.

In some embodiments, X is C _1-3 alkylene. In some cases, X is CH ₂ , CH ₂ CH ₂ or CH(CH ₃ ). In various cases, R ³ contains C _3-8 cycloalkyl, C _3-8 cycloalkenyl, C _3-7 heterocycloalkyl, C _6-10 aryl or C _2-6 heteroaryl. In some embodiments, R ³ contains cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopentenyl, cyclohexenyl, tetrahydropyranyl, phenyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl, furanyl, thiophenyl, thiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, pyridinyl, pyrazinyl or pyrimidinyl .

In various embodiments, XR ³ is selected from the group consisting of

In some embodiments, X is C=O or (C=O)O. In various embodiments, R ³ contains C _1-6 alkyl or C _6-10 aryl. In some cases, XR ³ is

or

In various cases, R ⁴ contains C _3-8 cycloalkyl or C _3-9 heterocycloalkyl. In some embodiments, R ⁴ is or ⁴ -Λ In different embodiments, R ⁴ contains

C _6-10 aryl or C _2-9 heteroaryl, and R ⁴ is optionally substituted with one to three groups independently selected from halogen, C 1-3 alkyl, C 1-3 alkoxy, C(O)N(R ⁿ ) 2 and N(R ⁿ ) ₂ and each RN independently represents H or C _1-3 alkyl. In some embodiments, R ⁴ is selected from the group consisting of,

MeO _h

Ome

-hl

E / g υλ / \ Ηγγ\ ’ and

. In various embodiments, R ⁴ is 'P/X. In some embodiments, R ^a , R ^b , R ^1a , and R ^1b are each H; R ¹ represents =CH ₂ or CH ₃ ;

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each Y represents SO2.

In some embodiments, R ¹ is in the ^ configuration.

Also provided herein are the compounds listed in Table 1. A and B, and their pharmaceutically acceptable salts. In some embodiments, provided herein is a compound selected from the group consisting of:

This document further provides a pharmaceutical composition containing

- 5 045973 compound of formula (I), the compound listed in table. A, the compound listed in table. B, or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

Also provided herein is a method of inhibiting protein secretion in a cell, comprising contacting the cell with a compound or salt of formula (I), a compound listed in Table. A, or a pharmaceutically acceptable salt thereof, a compound listed in table. B, or a pharmaceutically acceptable salt thereof, or with a pharmaceutical composition described herein, in an amount effective to inhibit secretion. In some cases, contacting involves administering the compound or composition to a subject.

Further provided herein is a method of treating inflammation in a subject, comprising administering to the subject a therapeutically effective amount of a compound or salt of Formula (I), a compound listed in Table. A, or a pharmaceutically acceptable salt thereof, a compound listed in table. B, or a pharmaceutically acceptable salt or pharmaceutical composition thereof described herein.

Also provided herein is a method of treating or preventing cancer or precancerous conditions in a subject, comprising administering to the subject a therapeutically effective amount of a compound or salt of formula (I), a compound listed in Table. A, or a pharmaceutically acceptable salt thereof, a compound listed in table. B, or a pharmaceutically acceptable salt or pharmaceutical composition thereof described herein.

Additional aspects and advantages will become apparent to those of ordinary skill in the art upon examination of the following detailed description as well as the drawings. The following description includes specific embodiments, it being understood that the description is illustrative and does not limit the invention to the specific embodiments described herein.

Brief description of drawings

The figure depicts B16F10 efficacy data for compound A87 compared to PD-1-targeted therapy. Compound A87 demonstrated improved efficacy when administered once weekly compared to PD-1-targeted therapy in a difficult-to-treat disease model in terms of tumor growth inhibition and percentage change in body weight.

Additional aspects and advantages will become apparent to those of ordinary skill in the art upon examination of the following detailed description. Although the compounds and methods described herein are capable of embodiments in various forms, specific embodiments are included in the following description, it being understood that the description is illustrative and does not limit the invention to the specific embodiments described herein. document.

Detailed Description of the Invention

Provided herein are compounds that inhibit protein secretion. The compounds described herein can be used to treat or prevent diseases associated with excess protein secretion, such as inflammation and cancer, and thereby improve the quality of life of affected individuals.

The compounds described herein have the structure of formula (I)

where the substituents are described in detail below.

Without being bound by any particular theory, the compounds described herein are believed to inhibit protein secretion by binding to and inactivating components of the translocon, including, but not limited to, Sec61, and in some cases interrupting, in a sequence-specific manner, interactions between formed the signal sequence of translated proteins and translocon components, including, but not limited to, Sec61. The compounds described herein can specifically bind to a signal sequence with little or no interaction with the translocon itself.

The compounds described herein preferably can inhibit TNFa secretion with an IC ₅₀ of up to 5 μM, or up to 3 μM, or up to 1 μM. In various cases, the compounds described herein can inhibit IL-2 secretion with an IC ₅₀ of up to 5 μM, or up to 3 μM, or up to 1 μM. In some cases, the compounds described herein can inhibit PD-1 secretion with an IC ₅₀ of up to 5 μM, or up to 3 μM, or up to 1 μM.

The compounds described herein are effective in reducing tumor growth and body weight.

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For example, compound A87 demonstrated improved tumor growth reduction efficacy when administered once weekly in a difficult-to-treat disease model compared to PD-1-targeted therapy. See examples section and figure.

Chemical definitions

As used herein, the term alkyl refers to linear and branched saturated hydrocarbon groups containing one to thirty carbon atoms, such as one to twenty carbon atoms or one to ten carbon atoms.

The term C _n means that the alkyl group contains n carbon atoms. For example, C ₄ alkyl refers to an alkyl group containing 4 carbon atoms. C _1-7 alkyl refers to an alkyl group containing carbon atoms in numbers covering the entire range (i.e., 1 to 7 carbon atoms) as well as all subgroups thereof (e.g., 1-6, 2-7, 1- 5, 3-6, 1, 2, 3, 4, 5, 6 and 7 carbon atoms). Non-limiting examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl (2-methylpropyl), tert-butyl (1,1-dimethylethyl), 3,3-dimethylpentyl and 2-ethylhexyl. Unless otherwise specified, the alkyl group may be an unsubstituted alkyl group or a substituted alkyl group.

As used herein, the term alkylene refers to a divalent saturated aliphatic radical. The term C _n means that the alkylene group contains n carbon atoms. For example, Ci-balkylene refers to an alkylene group containing carbon atoms in numbers covering the entire range, as well as all its subgroups, as described above for alkyl groups.

As used herein, the term alkenyl has the same definition as alkyl, except that it contains at least one carbon-carbon double bond and 2 to 30 carbon atoms, such as 2 to 20 carbon atoms or 2 to 10 carbon atoms. The term Cn means that the alkenyl group contains n carbon atoms. For example, C4alkenyl refers to an alkenyl group containing 4 carbon atoms. CZ-alkenyl refers to an alkenyl group containing carbon atoms in numbers covering the entire range (i.e., 2 to 7 carbon atoms) as well as all subgroups thereof (e.g., 2-6, 2-5, 3-6, 2, 3, 4, 5, 6 and 7 carbon atoms). Specifically proposed alkenyl groups include ethenyl, 1-propenyl, 2-propenyl and butenyl. Unless otherwise specified, an alkenyl group may be an unsubstituted alkenyl group or a substituted alkenyl group.

As used herein, the term cycloalkyl refers to an aliphatic cyclic hydrocarbon. The term C _n means that the cycloalkyl group contains n carbon atoms. For example, C ₅ cycloalkyl refers to a cycloalkyl group containing 5 carbon atoms in the ring. C _5-8 cycloalkyl refers to cycloalkyl groups containing carbon atoms in numbers including the entire range (i.e., 5 to 8 carbon atoms) as well as all subgroups thereof (e.g., 5-6, 6-8, 7- 8, 5-7, 5, 6, 7 and 8 carbon atoms). Non-limiting examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl. Unless otherwise specified, a cycloalkyl group may be an unsubstituted cycloalkyl group or a substituted cycloalkyl group.

As used herein, the term cycloalkenyl has the same definition as cycloalkyl, except that it contains at least one carbon-carbon double bond but is not aromatic. The term C _n means that the cycloalkenyl group contains n carbon atoms. For example, _C5 cycloalkenyl refers to a cycloalkenyl group containing 5 carbon atoms in the ring. C _5-8 cycloalkenyl refers to cycloalkenyl groups containing carbon atoms in numbers including the entire range (i.e. 5 to 8 carbon atoms) as well as all subgroups thereof (e.g. 5-6, 6-8, 7- 8, 5-7, 5, 6, 7 and 8 carbon atoms). Non-limiting examples of cycloalkenyl groups include cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl and cyclooctenyl. Unless otherwise specified, a cycloalkenyl group may be an unsubstituted cycloalkenyl group or a substituted cycloalkenyl group.

As used herein, the term heterocycloalkyl has the same definition as cycloalkyl, except that the ring contains from one to three heteroatoms independently selected from oxygen, nitrogen or sulfur. Non-limiting examples of heterocycloalkyl groups include piperidine, tetrahydrofuran, tetrahydropyran, dihydrofuran, morpholine, oxazepanyl, etc. Cycloalkyl and heterocycloalkyl groups may be saturated or partially unsaturated ring systems, optionally substituted with, for example, one to three groups independently selected from alkyl, alkylene-OH, C(O)NH2, NH2, oxo (=O), aryl, haloalkyl , halogen and OH. The heterocycloalkyl groups may optionally be further N-substituted, as described herein.

As used herein, the term aryl refers to monocyclic or polycyclic (eg, fused bicyclic and fused tricyclic) carbocyclic aromatic ring systems. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, tetrahydronaphthyl, phenanthrenyl, biphenylethyl, indanyl, indenyl, anthracenyl, fluorenyl, tetralinyl. Unless otherwise specified, the aryl group may be unsubstituted

- 7 045973 aryl group or substituted aryl group.

As used herein, the term heteroaryl refers to monocyclic or polycyclic (e.g., fused bicyclic and fused tricyclic) aromatic ring systems wherein one to four ring atoms are selected from oxygen, nitrogen, or sulfur and the remaining ring atoms are carbon, wherein the specified ring system is connected to the remainder of the molecule through any atom in the ring. Non-limiting examples of heteroaryl groups include, but are not limited to, pyridyl, pyridazinyl, pyrazinyl, pyrimidinyl, pyrrolyl, pyrazolyl, imidazolyl, thiazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiadiazolyl, oxadiazolyl, furanyl, thienyl, quinolinyl, isoquinolinyl, benzoxazolyl, benzimidazolyl , benzofuranyl , benzothiazolyl, triazinyl, triazolyl, purinyl, pyrazinyl, purinyl, indolinyl, phthalazinyl, indazolyl, quinolinyl, isoquinolinyl, cinnolinyl, quinazolinyl, naphthyridinyl, pyridopyridinyl, indolyl, 3H-indolyl, pteridinyl and quinoxalinyl. Unless otherwise specified, a heteroaryl group may be an unsubstituted heteroaryl group or a substituted heteroaryl group.

As used herein, the term hydroxy or hydroxyl refers to the group -OH.

As used herein, the term alkoxy or alkoxy refers to the group -O-alkyl.

As used herein, the term halogen is defined to include fluorine, chlorine, bromine and iodine.

As used herein, the term carboxy or carboxyl refers to the group -COOH.

As used herein, the term amino refers to a -NH2 or -NH- group, wherein any hydrogen atom may be replaced by an alkyl, cycloalkyl, aryl, heteroaryl or heterocycloalkyl group.

about Ph

As used herein, the term sulfonyl refers to group ⁰

A substituted functional group (eg, substituted alkyl, alkylenyl, cycloalkyl, aryl or heteroaryl) is a functional group in which at least one hydrogen radical is replaced by a radical other than hydrogen (ie, a substituent). Examples of non-hydrogen radicals (or substituents) include, but are not limited to, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, ether, aryl, heteroaryl, heterocycloalkyl, hydroxyl, oxy (or oxo), alkoxy, ester, thioester , acyl, carboxyl, cyano, nitro, amino, sulfhydryl and halogen. If the substituted alkyl group includes more than one radical other than hydrogen, then the substituents may be bonded to one carbon atom or to two or more different carbon atoms.

Protein secretion inhibitors.

In one aspect, the compounds of the invention have the structure of formula (I) or a pharmaceutically acceptable salt thereof:

R ^{2 R4} YX k R ^{1b Y} 'N ^A R' V ^X -A (ί) where each of R ^a and R ^b independently represents H or C _1-3 alkyl;

R ¹ is H, OH, C1-3alkyl, OC1-3alkyl, =CH2 or = ^NOR5 or ^R1 is C3-6cycloalkyl or _C3-6 heterocycloalkyl and forms a spiro group with a carbon atom in the ring to which it is attached;

R ^1a and R ^1b are each independently H or C _1-3 alkyl;

R2 is C _1-6 alkyl, N(R ⁵ )2, C _3-8 cycloalkyl, C _3-9 heterocycloalkyl, C _3-9 heterocycloalkenyl or C _6-10 aryl;

R ³ represents H, C _1-6 alkyl, C _3-8 cycloalkyl, C _3-8 cycloalkenyl, C _3-7 heterocycloalkyl, C _6-10 aryl or C _2-6 heteroaryl;

R ⁴ represents C _3-8 cycloalkyl, C _3-9 heterocycloalkyl, C _6-10 aryl or C _2-9 heteroaryl;

each R ⁵ independently represents H, C _1-3 alkyl or C _0-2 alkylene-C _6-10 aryl;

X is absent or represents C _1-3 alkylene, C=O or (C=O)O;

Y represents SO or SO2;

each heterocycloalkyl, heterocycloalkenyl and heteroaryl group independently contains 1, 2 or 3 heteroatoms on the ring selected from N, O and S.

In some embodiments, each Y is an SO. In various embodiments, each Y is SO ₂ .

In various cases, R ^a is H. In some cases, R ^a is C1-3 alkyl. In some embodiments, R ^a is CH3. In some embodiments, R ^b is H. In various embodiments, R ^b is C1-3 alkyl. In various cases, R ^b represents CH3.

In some cases, R ^a and R ^b are both H.

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In some cases, R ¹ represents H, OH or =NOR ⁵ . In various cases, R ⁵ represents H or CH3. Thus, suitable R ¹ groups may include H, OH, =NOH and =NORCH3. In some embodiments, R ¹ is C1-3alkyl or OC1-3αalkyl. For example, R ¹ may be CH3 or OCH ₃ . In some cases, R ¹ is CH3 and has S stereochemistry. In various embodiments, R ¹ is C3-6 cycloalkyl or C _3-6 heterocycloalkyl and forms a spiro group together with the carbon atom on the ring to which it is attached.

1

Suitable R ¹ spiro groups may include -X. In different cases, R ¹ is =CH2. In some embodiments, each of R ^1a and R ^1b is H. In some embodiments, at least one of R ^1a and R ^1b is C1-3 alkyl. In some embodiments, R ^1a and R ^1b are each CH3. In some embodiments, R ^a is H, R ^b is H, and R ¹ is CH3. In various embodiments, R ^a is CH3, R ^b is H, and R ¹ is H. In some cases, R ^a is H, R ^b is CH3, and R ¹ is H. In various cases, R ^{a is} is CH3, R ^b is H, and R ¹ is CH3. In some embodiments, R ^a is H, R ^b is CH3, and R ¹ is CH3. In various embodiments, R ^a is CH3, R ^b is CH3, and R ¹ is H.

In some embodiments, R2 is _C1-6 alkyl or N( ^R5 )2. In some cases, each R ⁵ independently contains H, C1-3 alkyl or benzyl. Suitable R2 groups may include Et, iPr, N(CH3)2J~ ⁴ '^ ^NH ' and ^ΗΝΐ -333^ο. in some cases R2 contains

C _3-8 cycloalkyl. In different cases, R2 contains cyclopentyl or cyclohexyl. In some embodiments, R2 is C _3-9 heterocycloalkyl or C _3-9 heterocycloalkenyl. In various embodiments, R2 contains oxetanyl, azetidinyl, tetrahydrofuranyl, pyrrolidinyl, tetrahydropyranyl, pyranyl, piperidinyl, piperazinyl, azepanyl, morpholinyl, or tetrahydropyridinyl. In various embodiments, the C _3-9 heterocycloalkyl or C _3-9 heterocycloalkenyl contains a bridging or spiro group. In some cases, the C _3-9 heterocycloalkyl containing a bridged or spiro group is selected from the group consisting of

and b ^N 3 ^:—R , and R6 is and CN. Examples of suitable R2 groups include

C _1-6 alkyl, C _3-8 cycloalkyl or C _6-10 aryl. In some cases, R6 is phenyl, optionally substituted with one to three groups independently selected from halogen, C _1-3 alkyl, C _1-3 alkoxy

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ci

IN

selected cases

In various cases, R ² represents ^-^aryl. In some embodiments, R ² is represented by

represents some or from a group consisting of. In different implementations, X is missing.

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In some embodiments, R ³ contains C1-6 alkyl or H. In some cases, R ³ is C1-6 alkyl, such as methyl, ethyl, propyl, butyl, pentyl, or hexyl. In some cases, R ³ is 2-methylbutyl, isopropyl, isopentyl, CH ₂ CH ₂ OCH ₃ , CH ₂ C(CH ₃ ) ₂ CN, CH2CF3 or CH ₂ CH ₂ CF ₃ . In various cases, R ³ is isobutyl.

In some embodiments, X is C _1-3 alkylene. In some cases, X is CH ₂ , CH2CH2 or CH(CH ₃ ). In various cases, R ³ contains ^^cycloalkyl, ^^cycloalkenyl, ^^heterocycloalkyl, ^-^aryl or ^^heteroaryl. In some embodiments, R ³ contains cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopentenyl, cyclohexenyl, tetrahydropyranyl, phenyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl, furanyl, thiophenyl, thiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, pyridinyl, pyrazinyl or pyrimidinyl . Examples of suitable XR ³ include ₅

In some cases the XR ³ is in some cases the XR ³ is

or

or

. In some way. In some embodiments, X is C=O or (C=O)O. In various embodiments, R ³ contains C _1-6 alkyl or O-yuaryl. In some cases, XR ³ is

or

In various cases, R ⁴ contains ^^cycloalkyl or ^^heterocycloalkyl, such as or . In various embodiments, R ⁴ contains ^-^ aryl or ^^ heteroaryl and R ⁴ is optionally substituted with one to three groups independently selected from halogen, C _1-3 alkyl, Ci-zalkoxy, C(O)N(R ⁿ )2 and N(R ⁿ )2, each R ^N independently representing H or C _1-3 alkyl.

In some embodiments, R ⁴ is selected from the group consisting of

In different embodiments, R ⁴ is

In some embodiments, R ^a , R ^b , R ^1a , and R ^1b are each H; R ¹ represents =CH ₂ or CH ₃ ;

or

;

R ⁴ represents

R ² represents

MeO

XR ³ is isobutyl,

or

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R ² is each Y is SO ₂ .

In some cases, R ¹ is in the ^-configuration. In some cases, this document

or , where or mente compounds are proposed that have the structure and ^C| and R ³ is cyclobutyl, cyclopentyl or cyclohexyl.

Examples of compounds of formula (I) are shown in table. And as compounds A1-A210 or pharmaceutically acceptable salts of these compounds. In some cases, the compound is compound A1-A151 or a pharmaceutically acceptable salt thereof.

Table A

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In some embodiments, the compounds of the invention include

or pharmaceutically acceptable salts of these compounds.

Additional compounds according to the invention are shown in table. B as compounds B1-B22 or pharmaceutically acceptable salts of these compounds. In some cases, the compound is compound B1-B20 or a pharmaceutically acceptable salt thereof.

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Table B

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On chemical structures containing one or more stereocenters, represented by dotted and bold bonds (i.e.......and -) are intended to indicate the absolute stereochemistry of the stereocenter(s) present in the chemical structure. Bonds indicated by a simple line do not indicate a preferred stereochemistry. Unless otherwise indicated, chemical structures that include one or more stereocenters and that are illustrated herein without specifying absolute or relative stereochemistry include all possible stereoisomeric forms of the compound (eg, diastereomers, enantiomers) and mixtures thereof. Structures that have one bold or dashed line and at least one additional single line include distinct enantiomer groups for all possible diastereomers.

Synthesis of protein secretion inhibitors.

The compounds provided herein can be synthesized by conventional methods using readily available starting materials known to those skilled in the art. In general, the compounds proposed herein are efficiently prepared by standard chemical methods of organic synthesis.

Synthesis of final compounds.

In some cases, the compounds described herein can be synthesized by reacting a sulfonyl chloride containing the target group R ² with propane-1,3diamine and reacting the resulting product with an aldehyde containing the target group R ³ to produce N- substituted propylamine compound. A propylamino group can be incorporated into N-substituted propylamine at the ^R4 position by reacting with 2-(3-bromopropyl)isoindoline-1,3-dione, then with hydrazine, and the target ^R4 group can be added to the compound by reacting the compound with interaction with sulfonyl chloride functionalized with the target group R ⁴ . Finally, the compound can be cyclized using a cyclization agent containing the desired R ¹ group, such as 3-chloro-2-chloromethyl-1-propene, 2-methylpropane-1,3diyl-diethanesulfonate, cyclopropane-1,1-diyl-bis- (methylene)diethanesulfonate, 1,3-dichloropropane or 1,4-dichlorobutane.

In some embodiments, compounds containing a methylene group as R ¹ can be synthesized by coupling a (3-aminopropyl)propane-1,3-diamino derivative containing the target R ³ and R ⁴ groups with the target R ² -sulfamoyl chloride or R ² -sulfanyl chloride group and then reacting the resulting compound with 3-chloro-2-chloromethyl1-propene to obtain the desired triazacyclododecanesulfonamide compound as suggested in Method 1 in the Examples section.

In various embodiments, compounds containing a methylene group as R ¹ can be synthesized by coupling a protected butane-1,3-diamine with an aldehyde containing the target R ³ group to produce 3-(azanyl)-N-butane-1- amine, bringing the amino group at the R ² position into interaction with a chloride group functionalized with the target group R ² , inserting a propylamino group into 3-(azanyl)-N-butan-1-amine at the R ⁴ position by reacting with 2-(3- bromopropyl)isoindoline-1,3-dione, then with hydrazine, reacting the resulting amino group at the R ⁴ position with a chloride group functionalized with the target group R ⁴ , and then reacting the resulting compound with 3-chloro-2-chloromethyl-1propene to obtain the desired triazacyclododecanesulfonamide compound as proposed in Method 2 in the Examples section.

In some cases, compounds containing a spiro group as R ¹ can be synthesized by reacting a derivative of ^(3-(azanyl)propyl)propane-1,3-diamino group containing the target groups R ² , R ³ and R ⁴ with a 1,1-diyl-bis-(methylene)diethanesulfonate derivative containing the desired spiro group as suggested in Methods 3, 8 and 9 in the Examples section.

In various cases, the compounds proposed herein can be synthesized by reducing the benzyl group at the R ² position of a ^(3-(azanyl)propyl)propane1,3-diamino derivative functionalized with the target groups R ³ and R ⁴ , and then reducing the resulting the product into interaction with the target aldehyde derivative containing the target R ² group (eg, dimethylcyclohexyl), as proposed in method 4 in the Examples section.

In some embodiments, compounds containing a carboxylate group as ^R2 ,

- 31 045973 can be prepared by reacting a derivative of a triazacyclododecanesulfonyl group containing the target groups R ¹ , R ³ and R ⁴ with the target alkyl group and triphosgene under basic conditions, as proposed in method 5 in the Examples section.

In various embodiments, compounds containing a piperidinyl group as R ² can be prepared by reacting DBU/mercaptoethanol with a triazacyclododecanesulfonyl compound containing the desired groups R ¹ , R ³ and R ⁴ and functionalized at the R ² position with a nitrophenyl group, and then combining the resulting product with piperidinyl sulfonyl chloride as suggested in Method 6 in the Examples section.

In some embodiments, compounds containing a hydrogen group as R ¹ can be synthesized by coupling N(3-(azanyl)propyl)propane-1,3-diamino, functionalized with the target groups R ² , R ^{3 ,} and R ⁴ , with 1 ,3-dichloropropane or 1,4-dichlorobutane, as suggested in Method 7 and Method 17 in the Examples section.

In some cases, compounds containing a chlorophenylmethanone group as R ² can be synthesized by reacting a triazacyclododecanesulfonyl functionalized with the desired groups R ¹ , R ³ and R ⁴ with 4-chlorobenzoyl chloride under basic conditions as suggested in Method 10 in the Examples section .

In various cases, compounds containing a 4-(dimethylamino)-2-methylbenzyl group as R ⁴ and a spiro group as R ¹ can be prepared by reducing N(3-(azanyl)propyl)propane-1,3-diamino-containing a compound functionalized with the target groups R ² and R ³ into interaction with 4-(dimethylamino)-2-methylbenzenesulfonyl chloride and subsequent cyclization of the compound by coupling with a 1,1-diyl-bis-(methylene)diethanesulfonate derivative containing the target spiro group using the methods described above herein, as proposed in Method 11 in the Examples section.

In some embodiments, the compounds described herein can be prepared by reacting a sulfonyl chloride containing the target group R ² with propane-1,3-diamine and reacting the resulting product with an aldehyde containing the target group R ³ to obtaining 3-(azanyl)-N-butan-1-amine compound. The propylamino group can be incorporated into 3-(azanyl)-N-butan-1-amine at the ^R4 position by reacting with 2-(3-bromopropyl)isoindoline-1,3-dione, then with hydrazine, and the target group R ⁴ can be added to a compound by reacting the compound with a target sulfonyl chloride (eg, 4-(dimethylamino)-2-methylbenzenesulfonyl chloride). Finally, the compound can be cyclized using an appropriate cyclization agent such as 3-chloro-2-chloromethyl-1-propene, 2-methylpropane-1,3-diyl-diethanesulfonate, cyclopropane-1,1-diyl-bis-(methylene) diethanesulfonate, 1,3 dichloropropane or 1,4-dichlorobutane, as described above herein and as suggested in Method 12 in the Examples section.

In various embodiments, compounds containing a 4-phenylpiperidin-1-ylsulfonyl group as R ² can be synthesized by reacting a triazacyclododecanesulfonyl group functionalized with the desired groups R ¹ , R ³ and R ⁴ with 3-methyl-1- trifluoromethanesulfonate ((4-phenylpiperidin-1-yl)sulfonyl)-III-imidazol-3-ium as suggested in Method 14 in the Examples section.

In some cases, compounds containing a 4-phenylpiperidin-1-ylsulfonyl group as ^R2 can be synthesized by reacting N(3-(azanyl)propyl)propan-1,3diamino functionalized with the desired ^R3 and ^R4 groups with 3-methyl-1-((4-phenylpiperidin-1-yl)sulfonyl)-III-imidazol-3-ium trifluoromethanesulfonate and subsequent cyclization of the compound using an appropriate cyclization agent such as 2-methylpropan-1,3-diyl- diethanesulfonate, 3-chloro-2-chloromethyl-1-propene, cyclopropane-1,1-diyl-bis(methylene)diethanesulfonate, 1,3-dichloropropane, or 1,4-dichlorobutane, as previously described herein and as suggested in method 15 in the Examples section. Similar compounds containing a methyl group adjacent to the nitrogen atom at the R ⁴ position can be synthesized in a similar manner using 3-chloro-2-(chloromethyl)prop-1-ene as suggested in Method 18 in the Examples section.

In some cases, compounds containing an alcohol group as R ¹ can be synthesized by cyclizing an N(3-(azanyl)propyl)propan-1,3-diamino-containing compound with a protected 1,3-dichloropropan-2-ol under basic conditions, as suggested in method 19 in the Examples section. The alcohol group at the R ¹ position can be methylated by reacting with MeI and NaH in toluene as suggested in Method 21 in the Examples section. The alcohol group at the R ¹ position can be oxidized to a carbonyl by reaction with DessaMartin periodinane as suggested in Method 22 in the Examples section. The carbonyl group at the R ¹ position can be reacted with hydroxylamine hydrochloride to produce an oxime compound as proposed in Method 25 in the Examples section.

In various cases, compounds containing an S-methyl group as R ¹ can be synthesized by reacting an amino group functionalized with the target R ² group with 2,3-dimethyl-1-((2-methyl-1H-imidazol- 1-yl)sulfonyl)-1H-imidαzol-3

- 32 045973 ia, then with CF ₃ SO ₃ Me to obtain 2,3-dimethyl-1H-imidazolium-3-trifluoromethanesulfonate, functionalized with the target group R ² . The product can be reacted with (K.)-'H-(3-amino-2methylpropyl) functionalized with the target group R ⁴ to obtain ^)-^^-(2-methylpropane1,3-diamino) functionalized with the target groups R ² and ^R4 . The resulting product can then be reacted with (azandiyl)-bis-(propan-3,1-diyl)dimethanesulfonate functionalized with the target group R ³ to obtain the target compound as taught in Method 27 in the Examples section.

Synthesis of intermediate compounds.

The intermediates used to prepare the compounds described herein can also be prepared by standard methods known to those skilled in the art, as described below in the Examples section (e.g., Method 13, Method 16, Method 20, Method 24, Method 26 , method 28, method 30, method 31, method 32, method 33).

Methods of application.

The compounds described herein (eg, the compounds of formula (I), the compounds listed in Tables A and B, and pharmaceutically acceptable salts of these compounds) can inhibit the secretion of a protein of interest. The compounds described herein can disrupt the secretion mechanism of the Sec61 protein in the cell. In some cases, a compound such as described herein inhibits the secretion of one or more of TNFa, VCAM, PRL, IL-2, INFy, CD4, insulin and PD-1 (mouse and/or human) or each of TNFa, VCAM , PRL, IL-2, INFy, CD4, insulin and PD-1 (mouse and/or human). In some cases, a compound such as described herein can inhibit the secretion of a checkpoint protein or inhibits the secretion of a cell surface protein, an endoplasmic reticulum associated protein, or a secreted protein involved in the regulation of antitumor immune responses. In various cases, a compound described herein may inhibit the secretion of one or more of PD-1, PD-L1, TIM-1, LAG-3, CTLA4, BTLA, OX-40, B7H1, B7H4, CD137, CD47, CD96 , CD73, CD40, VISTA, TIGIT, LAIR1, CD160, 2B4, TGFRP and their combinations. Protein secretion activity can be assessed in a manner such as that described below in the Examples section.

As used herein, the term inhibitor describes a compound that blocks or reduces the activity of a pharmacological target (eg, a compound that inhibits the function of Sec61 in the protein secretion pathway). The inhibitor may act through competitive, noncompetitive, or noncompetitive inhibition. The inhibitor may bind reversibly or irreversibly, and thus the term includes compounds that are suicidal substrates of the protein or enzyme. An inhibitor can modify one or more sites in or near the active site of a protein or can cause conformational changes in other regions of the enzyme. The term inhibitor is used herein in a broader sense than that used in the scientific literature and thus covers other classes of pharmacologically or therapeutically effective agents such as agonists, antagonists, stimulants, cofactors, etc.

Thus, methods are provided herein for inhibiting protein secretion in a cell. In these methods, the cell is brought into contact with a compound described herein (for example, a compound of formula (I) or a compound listed in Tables A or B, or pharmaceutically acceptable salts of these compounds), or a pharmaceutical composition containing the same in an amount , which effectively inhibits the secretion of the protein of interest. In some embodiments, the cell is brought into contact in vitro. In various embodiments, the cell is brought into contact in vivo. In some embodiments, contacting includes administering the compound or pharmaceutical composition to the subject.

The biological consequences of Sec61 inhibition are numerous. For example, inhibition of Sec61 has been proposed to treat or prevent inflammation and/or cancer in a subject. Thus, pharmaceutical formulations of compounds having specificity for Sec61 provide a means for administering a drug to a subject and treating these conditions. In this document, the terms treat, treatment, method of treatment, etc. refer to the elimination, reduction or amelioration of a disease or condition and/or its associated symptoms. Although possible, treatment of a disease or condition does not require complete elimination of the disease, condition, or symptoms associated with it. In this document, the terms treat, treatment, method of treatment, etc. may include prophylactic treatment, which refers to reducing the likelihood of recurrence of a disease or condition, or the recurrence of a disease or condition that has been previously treated, in a subject who does not have the disease or condition but who is at risk of or is susceptible to recurrence of the disease or condition or reappearance of a disease or condition. The term treat and its synonyms mean the administration of a therapeutically effective amount of a compound according to the invention to an individual in need of said treatment. Within the scope of the invention, treatment also includes prevention of relapses or prophylaxis

- 33 045973 tic of a certain phase of the disease, as well as treatment of acute or chronic signs, symptoms and/or disorders. Treatment may be aimed at symptoms, such as symptom suppression. It may be carried out over a short period, may be aimed at the medium term or may be a long-term treatment, for example in the context of maintenance therapy. As used herein, the terms prevent, prevent, prevent are those as are customary in the art and when used in relation to a condition such as a local relapse (eg, pain), a disease such as cancer, a complex syndrome such as heart failure , or any other medical condition, they are as is generally known in the art and involve administering a composition that reduces the frequency or delays the onset of symptoms of the medical condition in a subject compared to a subject not administering the composition. Thus, cancer prevention involves, for example, reducing the number of detectable growing cancers in a population of patients receiving prophylactic treatment compared with a control population without treatment and/or delaying the appearance of detectable cancers in a population receiving treatment compared with control population without treatment, for example, at a statistically and/or clinically significant level. Prevention of infection includes, for example, reducing the number of diagnoses of infection in a post-treatment population compared to a control population without treatment and/or delaying the onset of symptoms of infection in a population after treatment compared to a control population without treatment. Prevention of pain includes, for example, reducing the severity or, alternatively, delaying the onset of pain experienced by subjects in a treated population compared to an untreated control population. As used herein, the terms patient and subject may be used interchangeably to refer to animals such as dogs, cats, cows, horses and sheep (ie, non-human animals) and humans. Specific patients are mammals (eg, humans). The term patient includes men and women.

Inhibition of Sec61-mediated secretion of inflammatory proteins (eg, TNF-α) may impair inflammatory signaling. Thus, provided herein is a method of treating inflammation in a subject by administering to the subject a therapeutically effective amount of a compound described herein (i.e., a compound of formula (I) or a compound listed in Table A or B, or a pharmaceutically acceptable salt thereof connections).

In addition, the viability of cancer cells is based on increased protein secretion in the ER, which determines their survival. Thus, nonselective or partially selective inhibition of Sec61-mediated protein secretion may inhibit tumor growth. Alternatively, in immuno-oncology, selective inhibitors of known secreted or transmembrane immune checkpoint proteins (eg, PD-1, TIM-3, LAG3, etc.) can lead to activation of the immune system against various cancers.

Accordingly, also provided herein is a method of treating cancer in a subject by administering to the subject a therapeutically effective amount of a compound described herein (e.g., a compound of formula (I), a compound listed in Tables A or B, or a pharmaceutically acceptable salt thereof) . Specific anticipated cancers that may be treated with the compounds and compositions described herein include, but are not limited to, melanoma, multiple myeloma, prostate cancer, lung cancer, non-small cell lung carcinoma, squamous cell carcinoma, leukemia, acute myelogenous leukemia , chronic myelogenous leukemia, lymphoma, anaplastic large cell lymphoma with NPM/ALK transformation, diffuse large B-cell lymphoma, neuroendocrine tumors, breast cancer, mantle cell lymphoma, renal cell carcinoma, rhabdomyosarcoma, ovarian cancer, endometrial cancer, small cell carcinoma, adenocarcinoma , gastric carcinoma, hepatocellular carcinoma, pancreatic cancer, thyroid carcinoma, anaplastic large cell lymphoma, hemangioma, head and neck cancer, bladder cancer and colorectal cancer. In some cases, the cancer is a solid tumor. In different cases, the cancer is head and neck cancer, squamous cell carcinoma, gastric carcinoma or pancreatic cancer.

The compounds described herein are also contemplated for the prevention and/or treatment of a variety of diseases, including, but not limited to, proliferative diseases, neurotoxic/degenerative diseases, ischemic conditions, autoimmune and autoinflammatory disorders, inflammation, immune system-related diseases , HIV, cancers, organ transplant rejection, septic shock, viral and parasitic infections, conditions associated with acidosis, macular degeneration, pulmonary conditions, muscle wasting diseases, fibrotic diseases, bone diseases and hair growth diseases .

Examples of proliferative diseases or conditions include diabetic retinopathy, macular degeneration, diabetic nephropathy, glomerulosclerosis, IgA nephropathy, cirrhosis,

- 34 045973 bile duct resection, congestive heart failure, scleroderma, radiation fibrosis and pulmonary fibrosis (idiopathic pulmonary fibrosis, collagen vascular disease, sarcoidosis, interstitial lung diseases and external lung disorders).

Inflammatory diseases include acute (eg, bronchitis, conjunctivitis, myocarditis, pancreatitis) and chronic conditions (eg, chronic cholecystitis, bronchiectasis, aortic valve stenosis, restenosis, psoriasis and arthritis), as well as conditions associated with inflammation such as fibrosis, infection and ischemia.

Immunodeficiency disorders occur when part of the immune system does not work properly or is missing. They can affect B lymphocytes, T lymphocytes, or phagocytes and can be either inherited (eg, IgA deficiency, severe combined immunodeficiency (SCID), thymic dysplasia, and chronic granulomatosis) or acquired (eg, acquired immunodeficiency syndrome (AIDS), virus human immunodeficiency (HIV) and drug-induced immunodeficiencies). Conditions related to the immune system include allergic disorders such as allergies, asthma and atopic dermatitis such as eczema. Other examples of these immune system-related conditions include lupus, rheumatoid arthritis, scleroderma, ankylosing spondylitis, dermatomyositis, psoriasis, multiple sclerosis, and inflammatory bowel disease (such as ulcerative colitis and Crohn's disease).

Tissue/organ transplant rejection occurs when the immune system mistakenly attacks cells being introduced into the host's body. Graft-versus-host disease (GVHD) after allogeneic transplantation occurs when T cells from the donor tissue become aggressive and attack the host tissue. In all three cases, autoimmune disease, transplant rejection and GVHD, modulation of the immune system by treating the subject with a compound or composition of the invention may provide a beneficial effect.

Also provided herein is a method of treating an autoimmune disease in a patient, comprising administering a therapeutically effective amount of a compound described herein. An autoimmune disease as used herein is a disease or disorder that occurs in and is directed against an individual's own tissues. Examples of autoimmune diseases include, but are not limited to, inflammatory responses such as inflammatory skin diseases, including psoriasis and dermatitis (eg, atopic dermatitis); systemic scleroderma and sclerosis; responses associated with inflammatory bowel disease (such as Crohn's disease and ulcerative colitis); respiratory distress syndrome (including adult respiratory distress syndrome (ARDS)); dermatitis; meningitis; encephalitis; uveitis; colitis; glomerulonephritis; allergic conditions such as eczema and asthma, and other conditions in which T cell infiltration and chronic inflammatory responses occur; atherosclerosis; leukocyte adhesion deficiency; rheumatoid arthritis; systemic lupus erythematosus (SLE); diabetes mellitus (for example, type I diabetes mellitus or insulin-dependent diabetes mellitus); multiple sclerosis; Raynaud's syndrome; autoimmune thyroiditis; allergic encephalomyelitis; Sjögren's syndrome; diabetes that manifests itself in adolescence; and immune responses associated with acute and delayed hypersensitivity mediated by cytokines and T lymphocytes, typically occurring in tuberculosis, sarcoidosis, polymyositis, granulomatosis and vasculitis; pernicious anemia (Addison's disease); diseases in which diapedesis of leukocytes occurs; inflammatory disorder of the central nervous system (CNS); multiple organ failure syndrome; hemolytic anemia (including, but not limited to, cryoglobinemia or anemia with a positive Coombs test); myasthenia gravis; diseases mediated by the antigen-antibody complex; a disease with the formation of antibodies to the glomerular basement membrane; antiphospholipid syndrome; allergic neuritis; Graves' disease; Lambert-Eaton myasthenic syndrome; bullous pemphigoid; pemphigus; autoimmune polyendocrinopathies; Reiter's disease; muscle stiffness syndrome; Behçet's disease; giant cell arteritis; immune complex nephritis; IgA nephropathy; IgM polyneuropathy; immune thrombocytopenic purpura (ITP) or autoimmune thrombocytopenia. The compounds provided herein may be suitable for the treatment of conditions associated with inflammation, including, but not limited to, COPD, psoriasis, asthma, bronchitis, emphysema and cystic fibrosis.

The present document also provides the use of a compound such as described herein for the treatment of neurodegenerative diseases. Neurodegenerative diseases and conditions include, but are not limited to, stroke, ischemic nervous system injury, neuronal injury (eg, traumatic brain injury, spinal cord injury, and traumatic nervous system injury), multiple sclerosis, and other immune system-mediated neuropathies (eg, Guillain-Barré and its variants, acute motor axonal neuropathy, acute inflammatory demyelinating polyneuropathy and Fisher's syndrome), HIV/AIDS dementia complex, axonomia, diabetic neuropathy, Parkinson's disease, Huntington's disease, multiple sclerosis, bacterial, parasitic, fungal and viral meningitis , encephalitis, vascular dementia, multi-infarct dementia, dementia with Lewy bodies, frontal lobe dementia such as Pick's disease, subcortical dementias (such as Hahn's disease

- 35 045973 Tingtones or progressive supranuclear palsy), focal cortical atrophy syndromes (such as primary aphasia), metabolic-toxic dementias (such as chronic hypothyroidism or B12 deficiency) and dementias caused by infections (such as syphilis or chronic meningitis).

Additional recommendations for the use of the compounds and compositions described herein (eg, a compound of formula (I), a compound listed in Tables A or B, or a pharmaceutically acceptable salt thereof) to inhibit protein secretion can be found below in the Examples section.

Pharmaceutical Formulations and Introduction.

The methods provided herein include the preparation and use of pharmaceutical compositions that include one or more of the compounds provided herein. Also included are pharmaceutical compositions as such. Pharmaceutical compositions typically include a pharmaceutically acceptable carrier. Thus, provided herein are pharmaceutical compositions that include a compound described herein (e.g., a compound of formula (I), a compound listed in Table A or B, or a pharmaceutically acceptable salt thereof) as described above herein. document, and one or more pharmaceutically acceptable carriers.

The phrase pharmaceutically acceptable as used herein refers to ligands, materials, compositions and/or dosage forms that, within the bounds of sound medical judgment, are suitable for use in contact with human and animal tissues without causing excessive toxicity, irritation, allergic response or other problem. or complication, and have an acceptable benefit/risk ratio.

The phrase pharmaceutically acceptable carrier as used herein means a pharmaceutically acceptable material, composition or excipient, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material. As used herein, the expression pharmaceutically acceptable carrier includes buffers, sterile water for injection, solvents, dispersing media, coating materials, antibacterial and antifungal agents, isotonic agents and absorption delaying agents, etc., compatible with pharmaceutical administration. Each carrier must be acceptable in terms of compatibility with the other ingredients of the formulation and harmlessness to the patient. Some examples of materials that can be used as pharmaceutically acceptable carriers include (1) sugars such as lactose, glucose and sucrose; (2) starches such as corn starch, potato starch and substituted or unsubstituted β-cyclodextrin; (3) cellulose and its derivatives, such as sodium carboxymethylcellulose, ethylcellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter or suppository waxes; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols such as propylene glycol; (11) polyols such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline solution; (18) Ringer's solution; (19) ethyl alcohol; (20) phosphate buffer solutions and (21) other non-toxic compatible substances used in pharmaceutical formulations. In certain embodiments, the pharmaceutical compositions provided herein are pyrogen-free, i.e. do not cause a significant increase in temperature when administered to a patient.

The term pharmaceutically acceptable salt refers to the relatively non-toxic inorganic and organic acid addition salts of a compound provided herein. These salts may be prepared in situ during the final isolation and purification of the compound provided herein, or by separately reacting the free base form of the compound with a suitable organic or inorganic acid and isolating the salt thus obtained. Typical salts include hydrobromide, hydrochloride, sulfate, bisulfate, phosphate, nitrate, acetate, valerate, oleate, palmitate, stearate, laurate, benzoate, lactate, phosphate, tosylate, citrate, maleate, fumarate, succinate, tartrate, naphthylate, mesylate, glu coheptonate , lactobionate, lauryl sulfonate salts and amino acid salts, etc. (See, for example, Berge et al. (1977), Pharmaceutical Salts, J. Pharm. Sci. 66:1-19.)

In some embodiments, a compound provided herein may contain one or more acidic functional groups and thus may form pharmaceutically acceptable salts with pharmaceutically acceptable bases. The term pharmaceutically acceptable salts as used herein refers to the relatively non-toxic inorganic and organic base addition salts of a compound provided herein. These salts can likewise be prepared in situ during the final isolation and purification of the compound or by separately reacting the purified compound in free acid form with a suitable base such as a hydroxide, carbonate or bicarbonate pharmaceutically at

- 36 045973 acceptable metal cation, with ammonia and a pharmaceutically acceptable organic primary, secondary or tertiary amine. Typical alkali or alkaline earth metal salts include lithium, sodium, potassium, calcium, magnesium and aluminum salts, etc. Typical organic amines suitable for preparing base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, etc. (see, for example, Berge et al., above).

The compositions may also contain humectants, emulsifiers and lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as colorants, release agents, coating agents, sweeteners, flavoring agents, preservatives and antioxidants.

The compositions prepared as described above herein may be administered in different forms depending on the disorder being treated and the age, condition and body weight of the patient, as is well known in the art. For example, if the compositions are intended for oral administration, they may be in the form of tablets, capsules, granules, powders or syrups; if they are intended for oral administration, they can be prepared as injections (intravenous, intramuscular or subcutaneous), drips or suppositories. They can be obtained in the form of eye drops or eye ointments for application through the mucous membrane of the eye. These compositions can be prepared using traditional means using the methods described herein, and, if desired, the active ingredient can be mixed with any traditional additive or excipient such as a binder, disintegrant, lubricant, flavoring agent, agent, solubility enhancer, suspending agent, emulsifier or coating agent.

The actual dosages of the active ingredients in the pharmaceutical compositions provided herein can be varied to obtain a therapeutically effective amount, which is an amount of the active ingredient that effectively provides the target therapeutic response for a particular patient, composition and route of administration and is not toxic to the patient.

The concentration of a compound provided herein in a pharmaceutically acceptable mixture may vary depending on several factors, including the dosage of the compound administered, the pharmacokinetic characteristics of the compound(s) administered, and the route of administration. In some embodiments, the compositions provided herein may be provided in an aqueous solution containing about 0.1-10% (w/v) of a compound described herein, among other substances, for parenteral administration. Typical dosage ranges may include from about 0.01 to about 50 mg/kg body weight per day, administered in 1 to 4 separate doses. Each individual dose may contain the same or different compounds. The dosage is a therapeutically effective amount depending on several factors, including the general health of the patient and the formulation and route of administration of the selected compound(s).

Dosage forms or compositions can be prepared containing a compound such as described herein in an amount ranging from 0.005 to 100%, the balance being a non-toxic carrier. Methods for preparing these compositions are known to those skilled in the art. The proposed compositions may contain 0.001-100% of the active ingredient, in one embodiment 0.1-95%, in another embodiment 75-85%. Although the dosage may vary depending on the symptoms, age and weight of the patient, the nature and severity of the disorder being treated or prevented, the route of administration and the form of the drug, in general, the daily dosage of the compound is from 0.01 to 2000 mg recommended for the adult human patient and may be administered as a single dose or multiple divided doses. The amount of active ingredient that can be combined with a carrier material to form a unit dosage dosage form is generally the amount of compound that provides a therapeutic effect.

In jurisdictions that prohibit the patenting of methods administered to the human body, the meaning of the term administration of a composition to a human subject shall be limited to the administration of a controlled substance that the human subject will self-administer by any means (e.g., orally, inhalation, topical application, injection , introductions, etc.). The broadest permissible interpretation consistent with the law or regulation defining patentable subject matter is intended. In jurisdictions that do not prohibit the patenting of methods applied to the human body, the introduction of compositions includes both methods applied to the human body and the above operations.

Other implementation options.

Although the invention has been construed in detail in the specification, it should be understood that the foregoing description is intended to illustrate, but not limit, the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages and modifications are included within the scope of the following claims.

- 37 045973

Examples

The following examples are provided for illustration and are not intended to limit the scope of the invention.

Method 1.

The starting material was synthesized by the method described in J. Med. Chem. 2016, 59, 2633-2647.

To Na(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4-methylbenzenesulfonamide (0.200 g, 0.523 mmol) in DCM (2.0 mL) was added DIEA (2 eq., 1.05 mmol, 179 µl), then piperidine-1sulfonyl chloride (0.096 g, 0.523 mmol). After 16 h, quench the reaction with sodium bicarbonate (sat), extract with DCM, dry with sodium sulfate, filter and concentrate.

To crude Na(3-((cyclohexylmethyl)(3-((4-methylphenyl)sulfonamido)propyl)amino)propyl)piperidine-1-sulfonamide in DMF (10 mL) was added NaH (2.5 eq., 1.31 mmol, 0.052 g 60% dispersion in mineral oil). Stir the mixture for 1 hour, then warm to 85°C and add 3-chloro-2-chloromethyl-1-propene (0.9 eq., 0.471 mmol, 55 ml, dissolved in 0.5 ml DMF) over 4 h. After heating for 16 hours, cool the mixture to ambient temperature, dilute with brine and water, extract with ethyl acetate (3x), wash with brine, dry with sodium sulfate, filter and concentrate. Flash column chromatography (0-60% hexanes/ethyl acetate+1% DEA) provided the product in free base form.

Diethyl ether (1.0 mL) was added to the free base to dissolve and HCl (4N in dioxane, 1.046 mmol, 262 μL). The HCl salt was filtered and dried in high vacuum to obtain compound A5.

MS (ESI) for C ₂₉ H ₄₈ N ₄ O ₄ S ₂ , experiment 581 [M+H] ⁺ . The following compounds were synthesized similarly:

No. IU PAK [M+N] ⁺ Ab 9-(cyclohexylmethyl)-1-(ethanesulfonyl)-5-(4methylbenzenesulfonyl)-3-methylidene-1,5,9triazacyclododecane 526 A7 9-(cyclohexylmethyl)-1-(4-methylbenzenesulfonyl)-3methylidene-5-(propane-2-sulfonyl)-1,5,9-triazacyclododecane 540 A8 1 -(cyclohexane fonyl)-9-(cyclohexylmethyl)-5-(4methylbenzenesulfonyl)-3-methylidene-1,5,9triazacyclododecane 580

Method 2.

- 38 045973

To tert-butyl-(3-aminobutyl)carbamate (2.5 g, 13.3 mmol) in DCM (12.5 ml), sat. Na ₂ CO ₃ (12.5 ml) and sat. NaCl (12.5 ml) was added to TsCl (13.3 mmol, 2.54 g). After 1 hour, the organic layer was washed with brine, dried with sodium sulfate, filtered and concentrated to obtain tert-butyl (3-((4methylphenyl)sulfonamido)butyl)-carbamate, which was used without further purification.

TFA (66.5 mmol, 5.09 mL) and DCM (5.09 mL) were added to the tosylate (13.3 mmol). After 2 h, the reaction mixture was concentrated, diluted with NaOH (1N, 30 ml), extracted with DCM (3x30 ml), dried with sodium sulfate, filtered and concentrated to give the amine.

Toluene (49 ml) and cyclohexanecarboxaldehyde (1.34 g, 12.0 mmol) were added to the amine (10.94 mmol). Reflux the mixture using a Dean-Stark trap overnight (external temperature 185°C), then concentrate, dilute with ethanol (absolute, 19 ml) and add NaBH4 (0.827 g, 21.8 mmol) portionwise, maintaining at this internal temperature is <20°C. After stirring for 2 hours, dilute the mixture with water (10 ml), stir for 20 minutes, extract with DCM (3x), wash with brine, dry with sodium sulfate, filter and concentrate to give the amine. The amine was diluted with diethyl ether (20 ml) and HCl (conc., ~0.3 ml) was added dropwise. The HCl salt precipitated within 5 minutes and was filtered off using diethyl ether as a wash. The product was dried in vacuo overnight to obtain N(4-((cyclohexylmethyl)amino)butan-2-yl)-4-methylbenzenesulfonamide hydrochloride.

Acetonitrile (9 mL), sodium carbonate (0.99 g, 9.2 mmol), LiI (0.114 g) and 3-bromopropylphthalimide (2.30 g) were added to HCl salt (3.5 mmol). The mixture was heated to 70°C for 5 hours, after which it was filtered and concentrated. The crude phthalimide was used further without further purification.

Hydrazine monohydrate (4.2 ml) and ethanol (15 ml) were added to phthalimide (3.5 mmol). After heating to 80°C for 30 min, the reaction was completed, and the precipitate was filtered off, washing with ethanol. Concentrate the filtrate and dilute with HCl (2N, 30 ml), then leave to settle for 1 hour. NaOH (1N) is added to the aqueous mixture to alkalinize to pH 10. The mixture is then extracted with DCM (3x), dried with sodium sulfate, filtered and concentrated to obtain crude N(4-((3-aminopropyl)-(cyclohexylmethyl)amino)butan-2-yl)-4-methylbenzenesulfonamide.

To the amine (695 mg, 1.76 mmol) was added DCM (9 ml), sat. Na2CO3 (9 ml), sat. NaCl (9 ml), TsCl (1.76 mmol, 336 mg). After stirring for 1 hour, the mixture was extracted with DCM (3x), dried with sodium sulfate, filtered and concentrated to give the free base. The HCl salt was prepared by diluting the free base in diethyl ether (50 ml) and adding HCl (conc., 300 µl). After placing under high vacuum for 2 hours, the HCl salt (1.02 g) was collected.

To HCl salt (0.400 g, 0.683 mmol) in DMF (14 ml) was added NaH (3.5 eq., 2.39 mmol, 60% dispersion in mineral oil, 80.3 mg). After stirring for 1 hour, 3-chloro-2-chloromethyl-1-propene (0.9 eq, 0.615 mmol, 71 µl in 7 ml DMF) was added dropwise over 3 hours. After heating for a further 2 hours h cooled the mixture to ambient temperature, diluted with saline, water and ethyl acetate. The mixture was extracted with ethyl acetate (2x), washed with brine, dried with sodium sulfate, filtered and concentrated. Flash column chromatography (090% hexanes/ethyl acetate + 1% DEA) provided the product. The HCl salt was prepared by diluting the free base in diethyl ether (20 vol) and adding HCl (4N in dioxane, 4 eq.) to give 5-(cyclohexylmethyl)-2-methyl-11-methylene-1,9-ditosyl-hydrochloride. 1,5,9-triazacyclododecane, compound B1.

MS (ESI) for C ₃₂ H ₄₇ N ₃ O ₄ S ₂ , experiment 602 [M+H]+.

The following compounds were synthesized similarly:

No. IUPAC [M+N] ⁺ AT 2 1-(cyclohexylmethyl)-3-methyl-5,9-bis(4methylbenzenesulfonyl)-7-methylidene-1,5,9triazacyclododecane 602 A9 N, X-dimethyl-4-{[5-(4-methylbenzenesulfonyl)-3-methylidene9-[(oxan-4-yl)methyl]-1,5,9-triazacyclododecan-1yl] sulfonyl} aniline 619

Method 3.

EiOsSC^X^OSOjEl ”] \ NaH, DMF b HCI (4N in /A dioxane), /O._ DIETHYL Q l-b ether _; b

- 39 045973

Sodium hydride (0.350 mmol 60 % dispersion in mineral oil). Stir the mixture at ambient temperature for 30 minutes, then warm to 80°C, after which cyclopropane-1,1-diyl-bis(methylene)diethanesulfonate (0.099 mmol, dissolved in 1.0 ml DMF) was added over 2 h. After stirring for another 4 hours at 80°C, the mixture was cooled to ambient temperature, the reaction was quenched with saline, and extracted with ethyl acetate (3x). Wash the combined organic layers with brine (2x), dry with sodium sulfate, filter and concentrate. Flash column chromatography (0-60% hexanes (1% DEA)/ethyl acetate) gave the free base product, which was dissolved in diethyl ether (2 ml) and HCl (0.1 ml 4N in 1.4 -dioxane). The mixture was concentrated to obtain 4-{[9-(cyclohexylmethyl)-13-(4methylbenzenesulfonyl)-5,9,13-triazaspiro[2.11]tetradecan-5-yl]sulfonyl}-Ni-dimethylaniline hydrochloride, compound A11.

MS (ESI) for C33H50N4O4S2, experiment 631 [M+H]+.

Method 4.

/

The starting material was synthesized by the method described in J. Med. Chem. 2016, 59, 2633-2647. Pd/C (20 mg) was added to the starting material (0.327 mmol) in methanol (2.0 ml) under argon. A hydrogen atmosphere was introduced and the mixture was stirred at ambient temperature. After 3 hours, the reaction mixture was filtered and purified by flash column chromatography (0-80% hexanes (1% DEA)/ethyl acetate) to give the product.

Toluene (5 ml) was added to the starting material (0.028 mmol), then aldehyde (0.028 mmol). The mixture was stirred for 3 hours at reverse condensation temperature, then concentrated. Ethanol was added to the imine, then sodium borohydride. Stir the mixture at ambient temperature for 30 min, then dilute with water (1 ml), extract with DCM (3 x 1 ml), dry with sodium sulfate, filter and concentrate. The product was purified by flash column chromatography (060% hexanes (1% DEA)/ethyl acetate). Dilute the free base with diethyl ether (1 ml) and add HCl (7 µl, 4N in dioxane). The mixture was filtered to obtain 4-((9-((4,4dimethylcyclohexyl)methyl)-3-methylene-5-tosyl-1,5,9-triazacyclododecan-1-yl)sulfonyl)-HN dimethylaniline hydrochloride (compound A10) .

MS (ESI) for C34H52N4O4S2, experiment 645 [M+H]+.

Method 5.

// -N -N

To triphosgene (0.0316 mmol, 9.4 mg) in DCM (100 µl) was added 1-propanol (0.190 mmol, 14 µl). After stirring for 45 min, the resulting solution was added to amine (0.0158 mmol, 8.2 mg), DIEA (0.126 mmol, 22 μl) and DCM (100 μl). Stir the mixture at ambient temperature for 1 hour, then purify directly by flash column chromatography (0-70% hexanes/ethyl acetate) to give propyl-5-((4-(dimethylamino)phenyl)sulfonyl)- 7-methylene-9-tosyl-1,5,9triazacyclododecane-1-carboxylate (compound A12).

MS (ESI) for C29H42N4O6S2, experiment 607 [M+H]+.

Method 6.

The starting material, 4-((9-(cyclohexylmethyl)-3-methylene-5-((2-nitrophenyl)sulfonyl)-1,5,9triazacyclododecan-1-yl)sulfonyl)-X^dimethylaniline, was synthesized in a manner similar to that described in method 2, replacing with the corresponding sulfonyl chlorides.

To the starting material (1.09 g, 1.68 mmol) in ACN (12.3 mL), DBU (0.780 mL, 5.22 mmol) was added, followed by 2-mercaptoethanol (0.135 mL, 1.92 mmol). Stir the mixture at ambient temperature for 2 hours, then add an additional aliquot of 2-mercaptoethanol (1.14 eq.). After stirring for a further 2 hours, the mixture was concentrated and purified directly by FCC (0-100% hexanes (1% DEA)/ethyl acetate); 4-((9-(cyclohexylmethyl)-3-methylene-1,5,9-triazacyclododecan-1yl)sulfonyl)dimethylaniline.

To the amine (70 mg, 0.152 mmol) in DCM (2 ml) was added DIEA (0.303 mmol, 52 μl), then piperidine-1-sulfonyl chloride (0.152 mmol, 28 mg). After standing overnight, the reaction mixture was purified directly by flash column chromatography (hexanes (1% DEA)/ethyl acetate 0-60%). The HCl salt of the product was prepared by dissolving the free base in diethyl ether (1 ml) and adding HCl (4N in dioxane, 38 µl). The filtrate was collected to obtain 4-((9-(cyclohexylmethyl)-3-methylene-5-(piperidin-1-ylsulfonyl)-1,5,9-triazacyclododecan-1yl)sulfonyl)N-X-dimethylaniline hydrochloride (compound A15) .

MS (ESI) for C ₃₀ H ₅₁ N ₅ O ₄ S ₂ , experiment 610 [M+H] ⁺ .

The following compounds were synthesized similarly:______________________________

No. IUPAC [M+H1+ A16 9-(cyclohexylmethyl)-5-[4-(dimethylamino)benzenesulfonyl]X, Nd have l-3-methylidene-1,5,9-triazacyclic od odekai-1-sulfonamide 570 A13 4-{[9-(cyclohexylmethyl)-3-methylidene-5-(piperidine-1-sulfonyl)1,5,9-triazacyclododecan-1-yl]sulfonyl}-N,N-dimethylaniline 610

Method 7.

The starting material, N(3-((cyclohexylmethyl)(3-((4-(dimethylamino)phenyl)sulfonamido)propyl)amino)propyl)-4-methylbenzenesulfonamide, was synthesized in the same way as described in method 1, replacing the corresponding sulfonyl chloride.

NaH (0.22 mmol, 9 mg) was added to the diamine (49 mg, 0.087 mmol) in DMF (2.5 mL). After stirring for 30 min at ambient temperature, 1,3-dichloropropane (0.087 mmol, 8.3 μL in 0.5 mL DMF) was slowly added over 1 h while heating at 80°C. After 3 hours, cool the mixture to ambient temperature, dilute with saline (10 ml), water (5 ml) and ethyl acetate (10 ml). Extract the aqueous layer with ethyl acetate (3X5 ml), combine the organic layers and wash with brine (1X5 ml), dry with sodium sulfate and concentrate. Flash column chromatography (0-50% hexanes (1% DEA/ethyl acetate) gave the free base product, which was converted to the HCl salt by adding diethyl ether (1 ml) then HCl

- 41 045973 (20 µl, 4N in dioxane). The filtrate was collected and concentrated to give 4-((5-(cyclohexylmethyl)9-tosyl-1,5,9-triazacyclododecan-1-yl)sulfonyl)-N,Ndimethylaniline (compound A14).

MS (ESI) for C ₃₁ H ₄₈ N ₄ O ₄ S ₂ , experiment 605 [M+H]+.

The following compound was synthesized in a similar manner: ________________________

No. IU PAK [M+N] ⁺ A17 4-{[10-(cyclohexylmethyl)-14-(piperidin-1-sulfonyl)-2-oxa6,10,14-triazaspiro[3.11]pentadecan-6-yl]sulfonyl}-N, Ndimethylaniline 640

Method 8.

The starting material was synthesized by the method described in J. Med. Chem. 2006, 49, 1291-1312. To Na(3-aminopropyl)-N1-benzylpropane-1,3-diamine (363 mg, 1.64 mmol) in DCM (2 mL) was added DIEA (1.14 mL, 6.56 mL), then sulfonyl chloride ( 345 µl, 2.46 mmol). The solution was left to settle overnight at ambient temperature. Concentration and purification by flash column chromatography (0-80% hexanes (1% DEA)/ethyl acetate) gave the product.

NaH (50 mg 60% dispersion in mineral oil, 1.26 mmol) was added to the starting material (260 mg, 504 µl) in DMF (20 ml). Stir the mixture at 80°C for 20 min, then add the bisulfonate (144 mg, 504 µl) over 1 h. After heating for 4 h, the reaction is complete and the mixture is diluted with ethyl acetate (40 ml) and saline (40 ml) , extracted with ethyl acetate (2X20 ml). Wash the combined organic layers with brine (20 ml), dry with sodium sulfate, filter and concentrate. Flash column chromatography (hexanes (1% DEA)/ethyl acetate 0-50%) gave the product, which was diluted with diethyl ether (5 ml) and added HCl (4N in dioxane, 100 µl). Concentrate the mixture, wash with diethyl ether and dry to give 9-benzyl-5,13-bis-(piperidin-1-ylsulfonyl)-5,9,13 triazaspiro[2.11]tetradecane hydrochloride (compound A18).

MS (ESI) for C ₂₈ H ₄₇ N ₅ O ₄ S ₂ , experiment 581 [M+H] ⁺ .

Method 9.

χι ₀ V

X. ₀ -A 9 /—\ Et^SO-XX-OSOjEt

0 « f ^{0 0} , ⁰ g .

\_^ X DIEA.HXM v ) NaH, DMF h b

- 42 045973

The starting material, N(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4-(dimethylamino)benzenesulfonamide, was synthesized in a manner similar to that described in Method 2.

DIEA (0.71 mL, 4 .08 mmol), then sulfonyl chloride (250 mg, 1.36 mmol). The solution was left to settle overnight at ambient temperature. Concentration and purification by flash column chromatography (0-80% hexanes (1% DEA)/ethyl acetate) provided the product.

The cyclization of N(3-((cyclohexylmethyl)(3-((4-(dimethylamino)phenyl)sulfonamido)propyl)amino)propyl)piperidine-1-sulfonamide and the preparation of the salt were carried out similarly to method 8 to obtain 4-((9- (cyclohexylmethyl)-13-(piperidin-1-ylsulfonyl)-5,9,13-triazaspiro[2.11]tetradecan-5-yl)sulfonyl)-H^dimethylaniline (compound A19).

MS (ESI) for C31H53N5O4S2, experiment 624 [M+H]+.

The following compounds were synthesized similarly. The cyclization of compounds A25-A29 was carried out using NMP. as a solvent._______________________________________

No. IUPAC [M+N] ⁺ A20 4-{ [9-(cyclohexylmethyl)-13 -[(2-methylpiperidin-1yl)sulfonyl]-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl }-N, N-dimethylaniline 638 A21 4-{[9-(cyclohexylmethyl)-13-[(3-methylpiperidin-1yl)sulfonyl]-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl}-N, N-dimethylaniline 638 A22 4-{ [9-(cyclohexylmethyl)-13 -[(4-methylpiperidin-1yl)sulfonyl]-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl }-N, N-dimethylaniline 638 A23 4-{ [9-(cyclohexylmethyl)-13 -(pyrrolidine-1-sulfonyl)-5,9,13triazaspiro[2.11]tetradecan-5-yl]sulfonyl }-N, Ndimethylaniline 610 VZ 4-{[8-(cyclohexylmethyl)-4-(piperidine-1-sulfonyl)-!,4,8triazacycloundecan-1-yl]sulfonyl}-N,N-dimethylaniline 584 AT 4 4-{[7-(cyclohexylmethyl)-3-(piperidin-1-sulfonyl)-1,3,7triazecan-1-yl]sulfonyl}-M, N-dimethylaniline 570 A25 4-{[ 13 -(azepan-1-sulfonyl)-9-(cyclohexylmethyl)-5,9,13triazaspiro[2.11]tetradecan-5-yl]sulfonyl}-M, Ndimethylaniline 638 A26 4-{[9-(cyclohexylmethyl)-13-[(4,4-dimethylpiperidin-1yl)sulfonyl]-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl}-N, N-dimethylaniline 652 A27 4-{[9-(cyclohexylmethyl)-13 - [(4-methoxypiperide in-1 yl)sulfonyl]-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl}-N, N-dimethylaniline 654 A28 1 -{[9-(cyclohexylmethyl)-13-[4-(dimethylamino)benzenesulfonyl]5,9,13-triazaspiro[2.11]tetradecan-5-yl]sulfonyl}piperidin-4ol 640 A29 4-{[9-(cyclohexylmethyl)-13-[(4-phenylpiperidin-1yl)sulfonyl]-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl}-N, N-dimethylaniline 700

Method 10.

C.I.

To the amine (67 mg, 0.14 mmol) in DCM (12 ml) was added DIEA (35 mg, 0.27 mmol), then 4-chlorobenzoyl chloride (24 mg, 0.14 mmol). After standing at ambient temperature for 30 min, quench the reaction with sodium bicarbonate (sat), extract the mixture with DCM (3x), dry with sodium sulfate and filter. Flash column chromatography (0-60% hexanes (1% DEA)/ethyl acetate) gave (5,13-bis-(piperidin-1-ylsulfonyl)-5,9,13triazaspiro[2.11]tetradecan-9-yl) (4-chlorophenyl)methanone (compound A24).

MS (ESI) for C ₂₈ H ₄₄ ClN ₅ O ₅ S ₂ , experiment 630 [M+H]+.

- 43 045973

Method 11.

1. NaH, DMF

2. HCI/dioxane

Chlorosulfonic acid (4 ml) was added to a solution of XNi-3-trimethylaniline (1.35 g, 10.0 mmol) in CHCl ₃ (4 ml). Heat the reaction mixture at 80°C for 18 hours. Cool the mixture to ambient temperature and dilute with DCM (20 ml). The resulting mixture was poured into ice-water (30 ml) and then adjusted to pH 7-8 with saturated aqueous Na ₂ CO ₃ . The organic layer was separated, washed with saturated aqueous NaHCO ₃ , dried over anhydrous Na ₂ SO ₄ and concentrated. The residue was recrystallized from petroleum ether/EtOAc (10:1, 10 ml) to give 4-(dimethylamino)-2methylbenzenesulfonyl chloride.

1H NMR (400 MHz, CDCl3): δ 7.88 (d, J=9.6 Hz, 1H), 6.53 (m, 2H), 3.09 (s, 6H), 2.70 (s, 3H).

4-{[9-(Cyclohexylmethyl)-13-(piperidine-1-sulfonyl)-5,9,13-triazaspiro[2.11]tetradecane-5yl]sulfonyl}-NaX,3-trimethylaniline was prepared according to the method described in Method 12, compound A30 was obtained.

1H NMR (400 MHz, DMSO-06): δ 9.57 (b, 1H), 7.50 (d, J=8.8 Hz, 2H), 6.85 (m, 2H), 3, 95 (broad, 1H), 2.89~3.55 (m, 24H), 1.45~1.90 (m, 15H), 1.28 (m, 4H), 0.95 (m, 2H), 0.66 (br.z, 4H).

MS (ESI) for C ₃₂ H ₅₅ N ₅ O ₄ S ₂ , experiment 638 [M+H] ⁺ .

The following compound was synthesized similarly: _________________________

IUPAC

M+H1+

5-(cyclohexane sulfonyl)-9(cyclohexylmethyl)-13-(piperidine-1 sulfonyl)-5,9,13triazaspiro[2.11]tetradecane

Ή NMR (400 MHz,

DMSOA): δ 9.46 (yninp.s, 1H), 3.37-2.95 (m, 15H), 2.01-1.01 (m, 33H), 0.95 (m, 2H), 0.73 (m, 4H).

Method 12.

_{ 1. NaH, DMF

2. 2. HCI/dioxane

To a solution of piperidine-1-sulfonyl chloride (68.3 g, 0.37 mol) in toluene (350 ml) was added dropwise a solution of propane-1,3-diamine (83.4 g, 1.12 mol) in toluene (150 ml). Stir the reaction mixture overnight at ambient temperature.

The resulting suspension was filtered and the filtrate was concentrated. Purify the residue by silica gel column chromatography (DCM/MeOH = 50:1 to 20:1) to give N(3-aminopropyl)piperidine-1-sulfonamide.

- 44 045973 ¹ H NMR (400 MHz, CDC13): δ 3.17 (m, 6H), 2.88 (t, J=6.0 Hz, 2H), 1.65 (m, 6H), 1, 54 (m, 2H).

A mixture of H-(3-aminopropyl)piperidine-1-sulfonamide (22.62 mmol) and cyclohexanecarbaldehyde (2.8 g, 24.89 mmol) in toluene (90.0 ml) was heated at reflux temperature for nights, removing water using a Dean-Stark trap. Cool the mixture to ambient temperature and concentrate in vacuo. The resulting product was dissolved in ethanol (35 ml) and NaBH ₄ (1.66 g, 43.7 mmol) was added. Stir the reaction mixture at ambient temperature overnight. We extinguished the reaction 1n. aqueous HCl (30 ml) and then adjusted to pH 10 with aqueous NaOH. The resulting mixture was extracted with ethyl acetate (50 ml) and the organic layer was concentrated. Purify the residue by silica gel column chromatography (DCM/MeOH = 50:1 to 40:1) to give N(3-(cyclohexylmethylamino)propyl)piperidine-1-sulfonamide.

1H NMR (400 MHz, CDC13): δ 3.17 (m, 6H), 2.78 (t, J=6.0 Hz, 2H), 2.45 (d, J=6.4 Hz, 2H) , 1.75 (m, 12H), 1.44 (m, 1H), 1.23 (m, 3H), 0.96 (m, 2H).

MS (ESI) for Ci ₅ H ₃ iN ₃ O ₂ S, experiment 318 [M+H]+.

To a solution of Na(3-(cyclohexylmethylamino)propyl)piperidine-1-sulfonamide (2.8 g, 8.83 mmol) in CH3CN (50 ml) was added Na ₂ CO ₃ (0.97 g, 9.19 mmol), LiI (0.28 g, 2.12 mmol) and 3-bromopropylphthalimide (5.66 g, 21.19 mmol). The reaction mixture was heated at reflux temperature overnight. Cool the mixture to ambient temperature and filter. Concentrate the filtrate and purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:3) to give N(3-((cyclohexylmethyl)(3-(1,3-dioxoisoindolin-2yl)propyl)amino)propyl)-piperidine -1-sulfonamide.

1H NMR (400 MHz, CDC13): δ 7.85 (m, 2H), 7.73 (m, 2H), 5.82 (br, 1H), 3.68 (t, J=7.2 Hz, 2H), 3.17 (m, 6H), 2.46 (m, 4H), 2.14 (m, 2H), 1.82 (m, 2H), 1.67 (m, 12H), 1, 45 (m, 3H), 1.18 (m, 2H), 0.87 (m, 2H).

A solution of N-(3 -((cyclohexylmethyl)(3 -(1,3-dioxoisoindolin-2-yl)propyl)amino)-propyl)piperidine-1-sulfonamide (500 mg, 0.10 mmol) in EtOH (5 ml) NH2NH2 (1.5 ml, 2.55 mmol) was added. Stir the reaction mixture at ambient temperature overnight. The solvent was removed under reduced pressure and the residue was diluted with ethyl acetate (20 ml). The resulting mixture was washed with water (2x10 ml). Dry the organic layer over anhydrous sodium sulfate and concentrate to give N(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)piperidine-1sulfonamide (380 mg, quantitative yield).

1H NMR (400 MHz, CDC13): δ 3.17 (m, 6H), 2.78 (t, J=6.8 Hz, 2H), 2.45 (m, 4H), 2.14 (d, J=7.2 Hz, 2H), 1.82 (m, 2H), 1.67 (m, 12H), 1.45 (m, 3H), 1.18 (m, 2H), 0.87 ( m, 2H).

MS (ESI) for Ci ₈ H ₃₈ N ₄ O ₂ S, experiment 375 [M+H]+.

A solution of N-(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)piperidine-1-sulfonamide (2.42 g, 6.49 mmol) and DIEA (1.31 g, 10.3 mmol) in DCM (50 ml) was added with sulfonyl chloride (1.69 g, 7.13 mmol). Stir the reaction mixture at ambient temperature for 2 hours. Wash the mixture with water (50 ml). The organic phase was dried over anhydrous Na ₂ SO ₄ and concentrated. Purify the residue by silica gel column chromatography (MeOH/DCM = 1:80 to 1:30) to give A(3-((cyclohexylmethyl)(3-(4-(dimethylamino)-2-fluorophenylsulfonamido)propyl)amino)propyl )piperidine-1-sulfonamide.

1H NMR (400 MHz, CDC13): δ 7.66 (m, 1H), 6.43 (m, 2H), 5.58 (bb, 2H), 3.15 (m, 6H), 3, 03 (s, 6H), 2.99 (broadband, 2H) 2.46 (broadband, 4H), 2.12 (broadband, 2H), 1.30~1.70 (m, 18H ), 1.21 (m, 4H), 0.82 (m, 2H).

MS (ESI) for C ₂₆ H ₄₆ FN5O ₄ S ₂ , experiment 575 [M+H]+.

To a solution of N(3-((cyclohexylmethyl)(3-(4-(dimethylamino)-2-fluorophenylsulfonamido)propyl)amino)propyl)piperidine-1-sulfonamide (2.28 g, 3.97 mmol) in DMF (20 ml) sodium hydride (400 mg, 3.97 mmol) was added. Stir the mixture for 30 minutes at ambient temperature. Add cyclopropane-1,1-diyl-bis(methylene)diethanesulfonate (1.3 g, 3.97 mmol) and stir the reaction mixture at 80°C for 2 hours. Cool the mixture to ambient temperature and pour into water ( 150 ml). The resulting mixture was extracted with ethyl acetate (100 ml). The organic layer was washed with water (2 x 50 ml), dried over anhydrous sodium sulfate and concentrated. Purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:15 to 1:8) to give the free base, which is dissolved in diethyl ether (5 ml). Hydrochloric acid (1.5 ml of 3N solution in 1,4-dioxane) was added and the mixture was stirred for 0.5 hours. The solvent was removed in vacuo and the residue was triturated with diethyl ether three times to obtain compound A40.

1H NMR (400 MHz, DMSO-b6): δ 9.89 (bb, 1H), 7.54 (m, 1H), 6.63 (m, 2H), 3.67 (bb, 1H), 3.552.89 (m, 23H), 1.90-1.45 (m, 15H), 1.28 (m, 4H), 0.95 (m, 2H), 0.69 (m, 4H).

MS (ESI) for C ₃ iH ₅₂ FN ₅ O ₄ S ₂ , experiment 643 [M+H]+.

- 45 045973

The following compounds were synthesized similarly:___________________________ No. IUPAC [M+N] + Sh NMR A31 9-(cyclohexylmethyl)-5-(4methoxybenzene sulfonyl)-13(piperidine-1-sulfonyl)5,9,13triazaspiro [2. 11]tetradecane 611 Ή NMR (400 MHz, DMSO-7 ₆ ): δ 9.75 (b, 1H), 7.73 (d, ./=8.8 Hz, 2H), 7.16 (d, ./= 8.8 Hz, 2H), 3.85 (s, ZN), 3.552.89 (m, 18H), 1.90-1.45 (m, 15H), 1.28 (t, 4H), 0, 95 (t, 2H), 0.66 (t, 4H). A32 9-(cyclohexylmethyl)-5-(3methoxybenzene sulfonyl)-13(piperidine-1-sulfonyl)5,9,13triazaspiro [2. 11]tetradecane 611 Ή NMR (400 MHz, DMSO-7 ₆ ): δ 9.89 (b, 1H), 7.57 (t, 1H), 7.32 (t, 2H), 7.23 (s, 1H) , 3.85 (s, ZN), 3.55-2.89 (t, 18N), 2.04-1.45 (t, 15N), 1.28 (t, 4N), 0.95 (t , 2H), 0.67 (t, 4H). AZZ 9-(cyclohexylmethyl)-5-(2methoxybenzene sulfonyl)-13(piperidine-1-sulfonyl)- 611 ^H NMR (400 MHz, DMSO-7 ₆ ): 6 9.35 (b, 1H), 7.80 (t, 1H), 7.68 (t, 1H), 7.28 (t, 1H ), 7.13 (t, 1N), 3.89 (z, ZN), 5,9,13triazaspiro[2.11]tetradecane 3.55-2.89 (t, 18N), 1.90-1.45 (t, 15N), 1.28 (t, 4H), 0.95 (t, 2H), 0.66 (broad. h, 2H), 0.51 (broad z, 2H). A34 5-(benzenesulfonyl)-9(cyclohexylmethyl)-13(piperidine-1-sulfonyl)5,9,13triazaspiro[2.11]tetradecane 581 ^XH NMR (400 MHz, DMSOG ₆ ): δ 10.26 (b, 1H), 7.76 (t, 4H), 3.41-2.89 (t, 18H), 1.90-1 .45 (t, 15H), 1.28 (t, 4H), 0.95 (t, 2H), 0.73 (broadz, 4H). Method 13.

To 1-((1H-imidazol-1-yl)sulfonyl)-3-methyl-1H-imidazol-3-ium trifluorosulfonate (1.69 g, 4.65 mmol, see: J. Org. Chem. 2002, 68, 115-119.) and 4-phenylpiperidine (750 mg, 4.65 mmol), ACN (16 ml) was added and the solution was left to settle for 2 hours, then concentrated and purified directly by flash column chromatography (0-80% hexanes /ethyl acetate) to obtain the product.

To 1-((N-imidazol-1-yl)sulfonyl)-4-phenylpiperidine (600 mg, 2.06 mmol) in DCM at 0°C was added methyl triflate (0.23 mL, 2.06 mmol) over 5 min. The mixture was left to settle for 30 minutes, after which the mixture was concentrated to obtain 3-methyl-1-((4-phenylpiperidin-1yl)sulfonyl)-III-imidazolium-3-ium trifluoromethanesulfonate, which was used further without further purification.

MS (ESI) for C ₁₅ H ₂₀ N ₃ O ₂ S, experiment 307 [M] ⁺ .

The following compounds were synthesized similarly:

1-((8-chloro-3,4-dihydroisoooquinolin-2(1H)-yl)sulfonyl)3-methyl-1H-imidazol-3-ium trifluoromethanesulfonate (intermediate for compound A51).

1-((7-chloro-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)3-methyl-1H-imidazolium-3-trifluoromethanesulfonate (intermediate for compound A52).

1-((6-chloro-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)3-methyl-1H-imidazolium-3-ium trifluoromethanesulfonate (intermediate for compound A53).

1-((5-chloro-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)3-methyl-1H-imidazolium-3-trifluoromethanesulfonate (intermediate for compound A54).

1-((6-fluoro-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)3-methyl-1H-imidazolium-3-trifluoromethanesulfonate (intermediate for compound A59).

1-((6-bromo-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)3-methyl-1H-imidazolium-3-trifluoromethanesulfonate (intermediate for compound A60).

1-((4-isopropoxypiperidine-1) trifluoromethanesulfonate hydrochloride

- 46 045973 yl)sulfonyl)-3-methyl-1H-imidazol-3-ium (intermediate for compound A66) trifluoromethanesulfonate 1 -((4-(cyclopent-1 -en-1 -yl)-3,6- d in-1(2H)yl)sulfonyl)-3-methyl-1H-imidazol-3-ium hydropyridide (intermediate for compound A67).

trifluoromethanesulfonate 1 -((4-(cyclohex-1-en-1-yl)-3,6-d ihydropyrid in-1 (2H)yl)sulfonyl)-3-methyl-1H-imidazol-3-ium (intermediate connection for connection A68).

trifluoromethane 1-((4-(3-chlorophenyl)-3,6-d ihydropyrid in-1 (2H)yl)sulfonyl)-3-methyl-1H-imidazol-3-ium (intermediate for compound A69) trifluoromethanesulfonate 1-((4-(2-chlorophenyl)-3,6-dihydropyridin-1(2H)yl)sulfonyl)-3-methyl-1H-imidazol-3-ium phonate (intermediate for compound A73).

1-((4-(4-chlorophenyl)-3,6-dihydropyridin-1(2H)yl)sulfonyl)-3-methyl-1H-imidazol-3-ium trifluoromethanesulfonate (intermediate for compound A74).

1 -((4-( 1 H-pyrazol-1 -yl)piperidin-1 -yl)sulfonyl)-3 methyl-1H-imidazol-3-ium trifluoromethanesulfonate (intermediate for compound A79).

1-((4-(benzo[c1]oxazol-2-yl)piperidin-1 -yl)sulfonyl)3-methyl-1H-imidazol-3-ium trifluoromethanesulfonate (intermediate for compound A80).

3-methyl-1-((4-(pyrimidin-2-yl)piperidin-1 yl)sulfonyl)-1H-imidazol-3-ium trifluoromethane lphonate (intermediate for compound A81).

1-((2-chloro-7,8-dihydro-1,6-naphthyr idin-6(5H)yl)sulfonyl)-3-methyl-1H-imidazol-3-ium trifluoromethanesulfonate (intermediate for compound A82) .

1-((5-chloro isoind olin-2-yl)su fonyl)-3-methyl-1Nimidazol-3-ium trifluoromethanes fonate (intermediate for compound A89).

Method 14.

/

To 4-((9-(cyclohexylmethyl)-3-methylene-1,5,9-triazacyclododecan-1-yl)sulfonyl)-K,K-dimethylaniline (97 mg, 0.21 mmol) in ACN (0.8 ml) 3-methyl-1-((4phenylpiperidin-1-yl)sulfonyl)-1H-imidazol-3-ium trifluoromethanesulfonate (95 mg, 0.21 mmol) was added and the mixture was heated to 80°C for 8 hours, then concentrated and purified directly by flash column chromatography (0-70% hexanes (1% DEA)/ethyl acetate) to give the free base. Diethyl ether (5 ml) and HCl (52 µl 4N HCl in 1,4-dioxane) were added to the free base and the mixture was allowed to stand for 10 min, then concentrated to give 4-{[9-(cyclohexylmethyl)-3- hydrochloride methylidene-5-[(4-phenylpiperidin-1-yl)sulfonyl]-1,5,9-triazacyclododecan-1-yl]sulfonyl}-^^dimethylaniline (compound A35).

MS (ESI) for C36H55N5O4S2, experiment 686 [M+H]+.

- 47 045973 connections similar to:

No. 1IIOIIAK [M+N] ⁺ A36 A37 A38 4-{[9-(cyclohexylmethyl)-3-methylidene-5-[(4-phenylpiperazin-1yl)sulfonyl]-1,5,9-triazacyclododecan-1 -yl]sulfonyl} -N, Ndimethylaniline 4-{[9-(cyclohexylmethyl)-3-methylidene-5-[(4-phenoxypiperidin-1yl)sulfonyl]-1,5,9-triazacyclododecan-1 -yl]sulfonyl} -N, Ndimethylaniline 4-{[9-(cyclohexylmethyl)-3-methylidene-5 -[(3-phenylpiperidin-1 yl)sulfonyl]-1,5,9-triazacyclododecan-1 -yl]sulfonyl} -N, Ndimethylaniline 687 702 686 A39 1-{[9-(cyclohexylmethyl)-5-[4-(dimethylamino)benzenesulfonyl]-3methylidene-1,5,9-triazacyclododecan-1-yl]sulfonyl} piper ide in-4carbonitrile 635 A42 4-{[5-(cyclohexylmethyl)-9-[(4-phenylpiperidin-1-yl)sulfonyl]1,5,9-triazacyclic ododecai-1-yl]sulfonyl} -N, N-d methylaniline 674 A43 4-{[9-(cyclohexylmethyl)-3-methylidene-5-{[(3 S)-3-phenylpiperidin1-yl]sulfonyl}-1,5,9-triazacyclododecan-1-yl]su fonyl} -N, Ndimethylaniline 686 A46 4- {[9-(cyclohexylmethyl)-5 - {[(3R)-3 -methyl-4-phenylpiperazin-1 yl]sulfonyl}-3-methylidene-1,5,9-triazacyclododecan-1yl]sulfonyl}-M , N-dimethylaniline 701 A47 4- {[9-(cyclohexylmethyl)-5 - {[(3 S)-3 -methyl-4-phenylpiperazin-1 yl]sulfonyl}-3-methylidene-1,5,9-triazacyclododecan-1yl]sulfonyl}- M,N-dimethylaniline 701 A48 4-{[9-(cyclohexylmethyl)-5-[4-(dimethylamino)benzenesulfonyl]-3methylidene-1,5,9-triazacyclododo ekan-1 -yl]sulfonyl}-1 phenylpiperazin-2-one 701 A49 4-{[9-(cyclohexylmethyl)-3-methylidene-5-{[(3K)-3-phenylpiperidin1 -yl]sulfonyl}-1,5,9-triazacyclododecan-1 -yl]sulfonyl}-N, Ndimethylaniline 686 A50 4-{[9-(cyclohexylmethyl)-3-methylidene-5-(1,2,3,4tetrahydroisoquinoline-2-cy fonyl)-1,5,9-triazacyclododo ecan-1 yl]sulfonyl}-N, N- dimethylaniline 658

Method 15.

To Na(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4-(dimethylamino)-benzenesulfonamide (838 mg, 1.84 mmol) in ACN (5 ml) was added 3-methyl-1-( trifluoromethanesulfonate (4-phenylpiperidine-1

- 48 045973 yl)sulfonyl)-1H-imidazol-3-ium (838 mg, 1.84 mmol) and heated the mixture to 80°C for 8 hours, then concentrated and purified directly by flash column chromatography (0-70 % hexanes (1% DEA)/ethyl acetate) to obtain the product.

¹ H NMR (400 MHz, CDCl ₃ ): δ 7.68 (d, J=8.8 Hz, 2H), 7.33 (m, 5H), 6.68 (d, J=8.8 Hz, 2H), 5.75 (bb, 2H), 3.84 (m, 2H), 3.18 (m, 2H), 3.05 (s, 6H), 2.91 (m, 4H), 2.65 (m, 1H), 2.43 (m, 4H), 2.08 (m, 2H), 1.73 (m, 12H), 1.40 (m, 1H), 1.15 (m , 4H), 0.85 (m, 2H).

To 4-((9-(cyclohexylmethyl)-3-methyl-5-((4-phenylpiperidin-1-yl)sulfonyl)-1,5,9-triazacyclododecan-1-yl)sulfonyl)-K,K-dimethylaniline (123 mg, 0.194 mmol) to NMP (10 ml) was added sodium hydride (19 mg 60% dispersion in mineral oil, 0.485 mmol) over 20 min. The mixture was stirred at ambient temperature for 30 minutes, then at 80°C for another 20 minutes. The bisulfonate was then added over 1 hour. After 8 hours at 80°C, cool the mixture to ambient temperature, dilute with brine and ethyl acetate, extract with ethyl acetate (3x), wash the combined organic layers with brine (3x), dry with sodium sulfate , filtered and concentrated. Flash column chromatography (0-50% hexanes (1% DEA)/ethyl acetate) provided the free base. The free base was converted to the HCl salt by adding diethyl ether (5 ml) and HCl (4N in dioxane, 50 µl). Concentration for 10 minutes gave 4-{[9-(cyclohexylmethyl)-3-methyl-5-[(4-phenylpiperidin-1-yl)sulfonyl]1,5,9-triazacyclododecan-1-yl]sulfonyl} hydrochloride -H,Ndimethylaniline (compound A44).

MS (ESI) for C ₃₆ H ₅₇ N ₅ O ₄ S ₂ , experiment 688 [M+H]+.

The following compound was synthesized in a similar manner: ________________________

IUPAC |M +N] ⁺ 7 N- { 3 -[(cyclohexylmethyl)(3 - {[(4-phenylpiperidin-1 yl)sulphonyl] amino} propyl)amino]propyl} -4- (dimethylamino)benzene-1-sulfonamide 634

Method 16.

EtSChCI,

V7 thea, dkhm V7

HO^X^OH _____> EtOjSO^zX^OSOzEt

To 1,1-bis-(hydroxymethyl)cyclopropane (5.00 g, 49.0 mmol) in acetone (20 ml) was added TEA (2.2 eq., 108 mmol) and the mixture was cooled to 0-5°C. Ethanesulfonyl chloride was added at a rate to maintain the internal temperature below 10°C (~1 hour). After stirring for a further 2 hours at ambient temperature, the mixture was diluted with 150 ml water, extracted with ethyl acetate (2X50 ml), washed with brine, dried with sodium sulfate, filtered and concentrated to give the product.

¹ H NMR (400 MHz, CDCl ₃ ): δ 4.16 (s, 4H), 3.20 (m, 4H), 1.46 (t, J=7.2 Hz, 6H), 0.83 ( s, 4H).

MS (ESI) for C9H ₁₈ O ₆ S ₂ , experiment 287 [M+H] ⁺

The following compound was synthesized similarly: 2-methylpropane-1,3-diyl-diethanesulfonate (intermediate for compound A44).

Method 17.

HCI (4N in dioxane), diethyl ether

To Na(3-((cyclohexylmethyl)(3-((4-(dimethylamino)phenyl)sulfonamido)propyl)amino)propyl)-4phenylpiperidine-1-sulfonamide (89 mg, 0.14 mmol) in NMP (7 ml) NaH (14 mg 60% dispersion in mineral oil, 0.35 mmol) was added piecemeal over 20 min. Stir the mixture at ambient temperature for 30 minutes, then at 80°C for 20 minutes. Dichloride was then added and the mixture was heated at 80°C. After 8 hours at 80°C, cool the mixture to ambient temperature, dilute with brine and ethyl acetate, extract with ethyl acetate (3x),

- 49 045973 washed the combined organic layers with brine (3x), dried with sodium sulfate, filtered and concentrated. Flash column chromatography (0-50% hexanes (1% DEA)/ethyl acetate) provided the product. The product was converted to the HCl salt by adding diethyl ether (5 ml) and HCl (4N in dioxane, 35 µl). The mixture was concentrated to obtain 4-{[5-(cyclohexylmethyl)-9-[(4-phenylpiperidin-1-yl)sulfonyl]-1,5,9-triazacyclotridecan-1yl]sulfonyl}-I,Ni-dimethylaniline hydrochloride (compound B5 ).

MS (ESI) for C ₃₆ H ₅₇ N ₅ O ₄ S2, experiment 688 [M+H]+.

The following compounds were synthesized similarly: ______________________

No. IUPAC [M+N] ⁺ AT 8 4-{[(11 Z)-5 -(cyclohexylmethyl)-9-[(4-phenylpiperidin-1 yl)sulfonyl]-1,5,9-triazacyclotridec-11-en-1 -yl]sulfonyl} N, N -dimethylaniline 686 AT 9 4-{[(11 E)-5 -(cyclohexylmethyl)-9-[(4-phenylpiperidin-1yl)sulfonyl]-1,5,9-triazacyclotridec-11-en-1-yl]sulfonyl} N, N -dimethylaniline 686

Method 18.

Na(4-((3-Aminopropyl)(cyclohexylmethyl)amino)butan-2-yl)-4-(dimethylamino)benzenesulfonamide was synthesized according to method 2 with replacement of the corresponding sulfonyl chloride.

4-((5-(Cyclohexylmethyl)-2-methyl-11-methylene-9-((4-phenylpiperidin-1-yl)sulfonyl)-1,5,9triazacyclododecan-1-yl)sulfonyl)-I^-dimethylaniline synthesized according to the method described in method 15, replacing 3-chloro-2-(chloromethyl)prop-1-ene to give 4-((5-(cyclohexylmethyl)-2methyl-11-methylene-9-((4- phenylpiperidin-1-yl)sulfonyl)-1,5,9-triazacyclododecan-1-yl)sulfonyl)^^dimethylaniline (compound B6).

MS (ESI) for C37H57N5O4S2, experiment 700 [M+H]+.

The following compounds were synthesized similarly:___________________________

No. IUPAC [M+N] + Ή NMR B19 4-{[5-(cyclohexylmethyl)-2-methyl11-methylidene-9-[(4- phenylpiperidin-1 -yl)sulfonyl]1,5,9-triazacyclic od ecan-1 yl]sulfonyl }-N, Ndimethylaniline 700 A51 4-({5-[(8-chloro-1,2,3,4tetrahydroisoquinolin-2yl)sulfonyl]-9(cyclohexylmethyl)-3-methyl ide en1,5,9-triazacyclic od ecan-1 yl}sulfonyl) -K, Ndimethylaniline 692

- 50 045973

A52 - T4, Ndimethylaniline 692 A53 4-({5-[(6-chloro-1,2,3,4tetrahydroisoquinolin-2yl)sulfonyl]-9(cyclohexylmethyl)-3-methylide επί, 5,9-triazacyclic ecan-1yl}sulfonyl)- T4, Ndimethylaniline 692 Ή NMR (400 MHz, CDC1 ₃ ): δ 7.59 (d, 7=8.8 Hz, 2H), 7.15 (m, 2H), 7.04 (m, 1H), 6.70 (d , 7=8.8 Hz, 2H), 5.25 (s, 1H), 5.13 (s, 1H), 4.34 (s, 2H), 4.12 (s, 2H), 4.10 (s, 2H), 3.51 (w, 6H), 3.05 (s, 6H), 2.88 (m, 4H), 2.41 (m, 2H), 2.25 (m, 2H) , 1.95 (m, 2H), 1.85 (m, 2H), 1.67 (m, 6H), 1.23 (w, 4H), 0.85 (m, 1H). A54 4-({5-[(5-chloro-1,2,3,4tetrahydroisoquinolin-2yl)sulfonyl]-9(cyclohexylmethyl)-3-methylidene1,5,9-triazacyclic ecan-1yl}sulfonyl)-T4 , Ndimethylaniline 692 A56 - 3-fluoro-T4, Ndimethylaniline 710 A57 4-[(5 - {[4-(2-chlorophenyl)piperide in1 -yl]sulfonyl } -9(cyclohexylmethyl)-3-methylide επί , 5, 9-triazacycle odo ecan-1yl)sulfonyl]-X, N-dimotyl aniline 720 A58 4-[(5 - {[4-(3-chlorophenyl)p and perid in1 -yl] sulfonyl } -9(cyclohexylmethyl)-3-methylidene1,5,9-triazacycle odo ecan-1 yl)sulfonyl] -T4, N-dimethyl aniline 720 A59 4-{[9-(cyclohexylmethyl)-5 - [(6fluoro-1,2,3,4-tetrahydroisoquinolin2-yl)sulfonyl]-3-methylidene-1,5,9triazacyclododecan-1 yl]sulfonyl}-N, Ν - 676

- 51 045973

dimethylaniline A60 4-({5-[(6-bromo-1,2,3,4tetrahydroisoquinolin-2yl)sulfonyl]-9(cyclohexylmethyl)-3-methylidene1,5,9-triazacyclic odoecan-1yl}sulfonyl)-H Ndimethylaniline 736 A64 4- {[9-(cyclohexylmethyl)-5 - [(4cyclopentylpiperidin-1 yl)sulfonyl]-3-methylidene-1,5,9triazacyclododecan-1 yl]sulfonyl }-N, Ndimethylaniline 678 A65 4- {[9-(cyclohexylmethyl)-5 - [(4cyclohexylpiperidin-1yl)sulfonyl]-3-methylidene-1,5,9triazacyclododecan-1yl]sulfonyl}-N, Ndimethylaniline 692 A66 4-{[9-(cyclohexylmethyl)-3 methylidene-5-{[4-(propan-2yloxy)piperidin-1yl]sulfonyl)-1,5,9triazacyclododecan-1yl]sulfonyl}-N, Ndimethylaniline 668 A67 4-{ [9-(cyclohexylmethyl)-5-{ [4(cyclopent-1-eu-1-yl)-1,2,3,6tetrahydropyridin-1yl] sulfonyl }-3-methylidene-1,5 ,9triazacyclododecan-1yl] sulfonyl }-N, Ndimethylaniline 674 A68 4-[(5 - {[4-(cyclohex-1-ey-1-yl)1,2,3,6-tetrahydropyridin-1yl]sulfonyl}-9(cyclohexylmethyl)-3-methylide επί, 5,9- triazacyclic od od ecan-1 yl)sulfonyl]-N, N-dimethyl aniline 688 A69 4-[(5-{[4-(3-chlorophenyl)-1,2,3,6tetrahydropyridin-1 yl]sulfonyl}-9(cyclohexylmethyl)-3-methylidene1,5,9-triazacycle od odecan-1 - 718

- 52 045973

yl)sulfonyl]-H N-dimethyl aniline A70 2-{[9-(cyclohexylmethyl)-5-[4(d imethyl amino)benzene sulfonyl] 3-methylidene-1,5,9triazacyclododecan-1yl]sulfonyl}-1,2,3,4tetrahydroisoquinoline-6carbonitrile 683 A71 4-{[9-(cyclohexylmethyl)-5 - [(6methoxy-1,2,3,4tetrahydroisoquinolin-2yl)sulfonyl]-3-methylidene-1,5,9triazacyclododecan-1yl]sulfonyl}-N, Ndimethylaniline 688 A72 4-[(5 - {[4-(4-chlorophenyl)piperide in1 -yl] sulfonyl } -9(cyclohexylmethyl)-3 - methyl ide en1,5,9-triazacycle od ecan-1 yl)sulfonyl]-No , N-dimethyl aniline 720 A73 4-[(5-{[4-(2-chlorophenyl)-1,2,3,6tetrahydropyridin-1 yl]sulfonyl}-9(cyclohexylmethyl)-3 - methyl ide επί, 5,9-triazacycle odoecane -1 yl)sulfonyl]-T\G, N-dimethyl aniline 718 A74 4-[(5-{[4-(4-chlorophenyl)-1,2,3,6tetrahydropyridin-1 yl]sulfonyl}-9(cyclohexylmethyl)-3 - methyl ide επί, 5,9-triazacycle odoecane -1 yl)sulfonyl]-T\G, N-dimethyl aniline 718 A78 4- {[9-(cyclohexylmethyl)-3 methylidene-5-{ [4-(oxan-4yl)piperid in-1 -yl] sulfonyl } - 1,5,9-triazacycle odo ecan-1yl] sulfonyl }-N, Νdimethylaniline 694 A79 4- {[9-(cyclohexylmethyl)-3 methylidene-5 - {[4-( 1 H-pyrazol-1 yl)piperid in-1 -yl] sulfonyl } - 1,5,9-triazacycle odo ecan-1yl] sulfonyl }-N, Νdimethylaniline 676 A80 4- [(5 - {[4-( 1,3 -benzoxazole-2- 727

- 53 045973

yl)piperidin-1 -yl]sulfonyl } -9(cyclohexylmethyl)-3-methylide ene1,5,9-triazacyclic ododecan-1 yl)sulfonyl]-T4, N-dimethyl aniline A81 4- {[9-(cyclohexylmethyl)-3 methylidene-5-{ [4-(pyrimidin-2yl)piperid in-1 -yl] sulfonyl } - 1,5,9-triazacycle od od ecan-1 yl]sulfonyl }-N, Ndimethylaniline 688 A82 4-( { 5 - [(2-chloro-5,6,7,8-tetrahydro1,6-naphthyridin-6-yl)sulfonyl]-9(cyclohexylmethyl)-3-methylide επί, 5,9-triazacyclic ekan-1 yl}sulfonyl)-18G, Ndimethylaniline 693 A88 (32)-9-(cyclohexylmethyl)-1 - [4(d imethyl amino)benzene sulfonyl] M-methoxy-5-[(4-phenylpiperidin1 -yl)sulfonyl]-1,5,9triazacyclododoecan- 3 -imin 717 A83 4-[(5 - {[4-(2-chlorophenyl)piperazine- 1 -yl]sulfonyl } -9(cyclohexylmethyl)-3-methylide επί, 5, 9-triazacyclic ododcan-1yl)sulfonyl]-T4, N-dimothyl aniline 721 Ή NMR (400 MHz, CDC1 ₃ ): δ 7.61 (d, 7=8.8 Hz, 2H), 7.39 (m, 2H), 7.05 (t, 2H), 6.82 (d , 7=8.8 Hz, 2H), 5.35 (t, 2H), 4.10 (t, 2H), 3.41 (t, 6H), 3.10 (t, 4H), 3.05 (s, 6H), 2.88 (t, 2H), 2.41 (t, ZN), 2.21 (t, 4H), 2.01 (t, ZN), 1.85 (t, 6H) , 1.55 (t, IN), 1.15 (t, 9N), 0.82 (t, ZN). A84 4-[(5 - {[4-(3-chlorofonyl)piporazine- 1 -yl]sulfonyl } -9(cyclohxylmstil)-3-methylide сН1,5,9-triazacyclododecan-1yl)sulfonyl]-T4, N-dimethyl aniline 721 Ή NMR (400 MHz, CDCh): δ 7.59 (d, 7=8.8 Hz, 2H), 7.18 (t, 1H), 6.85 (t, ZH), 6.78 (d, 7=8.8 Hz, 2H), 5.25 (t, 2H), 4.10 (t, 2H), 3.51 (t, 2H), 3.20 (t, UN), 3.05 ( s, 6H), 2.88 (t, 2H), 2.41 (t, 2H), 2.01 (t, 2H), 1.85 (t, 4H), 1.65 (t, 6H), 1.25 (t, 5H), 0.82 (t, 1H). A85 4-[(5 - {[4-(4-chlorophenyl)piperazine- 1 -yl]sulfonyl } -9(cyclohexylmethyl)-3-methylide ene1,5,9-triazacyclic ododecan-1yl)sulfonyl]-M, N-dimethyl aniline 721 Ή NMR (400 MHz, CDCI3): δ 7.59 (d, 7=8.8 Hz, 2H), 7.26 (d, 7=8.0 Hz, 2H), 6.87 (d, 7= 8.0 Hz, 2H), 6.70 (d, 7=8.8 Hz, 2H), 5.25 (t, 2H), 4.10 (t, 2H), 3.51 (t, 2H) , 3.20 (t, JN), 3.05 (s, 6H), 2.88 (t, 2H), 2.41 (t, 2H), 2.01 (t, 2H), 1.85 ( t, 4H), 1.65 (t, 6H), 1.27 (t, 5H), 0.85 (t, 1H). A89 4-( { 5 - [(5-chloro-2,3-dihydro-1Nisoindol-2-yl)sulfonyl]-9(cyclohexylmethyl)-3-methylide επί , 5, 9-triazacyclic odocai-1 yl }sulfonyl )-T4, Ndimothylaniline 678 Ή NMR (400 MHz, CDCI3): δ 7.59 (d, 7=8.8 Hz, 2H), 7.25 (t, 4H), 6.68 (d, 7=8.8 Hz, 2H) , 5.28 (s, 1H), 5.12 (s, 1H), 4.64 (t, 4H), 4.12 (s, 2H), 3.45 (t, 4H), 3.05 ( s, 6H), 2.88 (t, 2H), 2.25 (t, 2H), 1.95 (t, 2H), 1.65 (t, 8H), 1.23 (t, 8H), 0.85 (t, 1H).

Method 19.

- 54 045973

To Na(3-((cyclohexylmethyl)(3-((4-(dimethylamino)phenyl)sulfonamido)propyl)amino)propyl)-4phenylpiperidine-1-sulfonamide (33 mg, 0.052 mmol) in NMP (3 ml) was added sodium (4.6 mg 60% dispersion in mineral oil, 0.11 mmol). The mixture was stirred at ambient temperature for 30 minutes, then at 80°C for another 20 minutes. Dichloride (19 mg, 0.052 mmol) was then added. After 2 hours at 80°C, cool the mixture to ambient temperature, dilute with brine and ethyl acetate, extract with ethyl acetate (3x), wash the combined organic layers with brine (3x), dry with sodium sulfate, filter and concentrate. Flash column chromatography (0-50% hexanes (1% DEA)/ethyl acetate) gave 9-(cyclohexylmethyl)-1-[4-(dimethylamino)benzenesulfonyl]-5-[(4-phenylpiperidin-1-yl) sulfonyl]1,5,9-triazacyclododecan-3-ol (compound A55).

MS (ESI) for C ₃₅ H ₅₅ N ₅ O ₅ S ₂ , experiment 690 [M+H]+.

Method 20.

/ HCHO / / /=< NaBH ₃ CN \ /=< BULL hso ₃ ci \ /=\

Η,ΝΥ·;., —I -----------y ^N y, //— ¹ --------------” / ^N y. y ^S0 2 ^CI

To a solution of 3-fluoro-4-iodoaniline (1.11 g, 4.68 mmol) in acetonitrile (15.00 ml) was added 37% aqueous formaldehyde (8.0 ml, 99.9 mmol) and sodium cyanoborohydride (1. 88 g, 29.97 mmol). Acetic acid (1 ml) was then added dropwise over 10 min and the reaction mixture was stirred at ambient temperature overnight. Aqueous NaOH (1N, 30 ml) was added and the resulting mixture was extracted with ethyl acetate (50 ml). The organic layer was washed with brine, dried over anhydrous sodium sulfate and concentrated to give 3-fluoro-4-iodo-H^dimethylaniline.

¹ H NMR (400 MHz, CDCl ₃ ): δ 7.47 (m, 1H), 6.43 (m, 1H), 6.28 (m, 1H), 2.94 (s, 6H).

To a solution of 3-fluoro-4-iodo-H^dimethylaniline (500 mg, 1.89 mmol) in diethyl ether (5 ml) was added n-BuLi (0.8 ml, 1.89 mmol) at -78°C. The mixture was stirred for 0.5 h at the indicated temperature. Sulfuryl dichloride (405 mg, 3 mmol) was added and the reaction mixture was stirred for 1 hour. Quench the reaction with water (10 ml) and then adjust the mixture to pH 8 with saturated aqueous NaHCO ₃ . The resulting mixture was extracted with ethyl acetate (10 ml). The organic phase was separated and concentrated. The residue was purified by silica gel column chromatography to give 4-(dimethylamino)-2-fluorobenzene-1-sulfonyl chloride.

1H NMR (400 MHz, CDCla): δ 7.73 (m, 1H), 6.43 (m, 2H), 3.15 (s, 3H).

Method 21.

To 9-(cyclohexylmethyl)-1-((4-(dimethylamino)phenyl)sulfonyl)-5-((4-phenylpiperidin-1yl)sulfonyl)-1,5,9-triazacyclododecan-3-ol (30 mg, 0.043 mmol) in toluene (0.5 ml) was added NaH (1.6 mg of 60% dispersion in mineral oil, 0.065 mmol), then iodomethane (14 μl, 0.22 mmol). After stirring for 1 hour, quench the reaction with water, extract the mixture with DCM (2x), combine the organic layers and dry with sodium sulfate, filter and concentrate. Flash column chromatography (0-50% hexanes/ethyl acetate + 1% DEA) gave

4-{[9-(cyclohexylmethyl)-3-methoxy-5-[(4-phenylpiperidin-1-yl)sulfonyl]-1,5,9-triazacyclododecan1-yl]sulfonyl}-H^dimethylaniline (compound A61).

MS (ESI) for C36H57N5O5S2, experiment 704 [M+H] ⁺ .

Method 22.

To 9-(cyclohexylmethyl)-1-((4-(dimethylamino)phenyl)sulfonyl)-5-((4-phenylpiperidin-1yl)sulfonyl)-1,5,9-triazacyclododecan-3-ol (171 mg, 0.248 mmol) DCM (10 ml) was added, then Dess-Martin periodinane (126 mg, 0.297 mmol). The mixture was allowed to sit for 2 hours at RT and then concentrated and purified directly by FCC (0-60% hexanes (1% DEA)/ethyl acetate) to give 9-(cyclohexylmethyl)-1-((4-(dimethylamino )phenyl)sulfonyl)-5-((4-phenylpiperidin-1yl)sulfonyl)-1,5,9-triazacyclododecan-3-one (compound B20).

MS (ESI) for C35H53N5O5S2, experiment 688 [M+H] ⁺ .

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Method 23.

K 4-((5-((6-bromo-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)-9-(cyclohexylmethyl)-3-methylene1,5,9-triazacyclododecan-1-yl)sulfonyl )-HRP-dimethylaniline (70 mg, 0.095 mmol) in DMF (0.7 ml) was added water (0.1 ml), phenylboronic acid (13 mg, 0.10 mmol), P6(6pp^Cl ₂ -DCM ( 7.8 mg, 0.001 mmol) and sodium carbonate (30 mg, 0.29 mmol).The mixture was heated to 80°C for 1 hour, then diluted with brine, extracted with ethyl acetate (2x), dried with sodium sulfate, filtered and concentrated Flash column chromatography (0-50% hexanes (1% DEA)/ethyl acetate) gave 4-((9-(cyclohexylmethyl)-3-methylene-5-((6-phenyl-3,4-dihydroisoquinoline- 2(III)yl)sulfonyl)-1,5,9-triazacyclododecan-1-yl)sulfonyl)^X-dimethylaniline (compound A63).

MS (ESI) for C ₄₀ H ₅₃ N ₅ O ₄ S ₂ , experiment 734 [M+H]+.

Method 24.

σΒ· ^soGdhm -/ Pd(PPh3)4, No. ₂ CO3,

1,4-dioxane, water

To a mixture of cyclopentanone (5.50 g, 65.5 mmol) and Na ₂ CO ₃ (10.4 g, 98.2 mmol) in DCM (130 ml) was added Tf ₂ O (18.6 g, 72.0 mmol ) at -20°C. The reaction mixture was allowed to warm to ambient temperature and stirred overnight. The solid was filtered off and the filtrate was concentrated to give cyclopentenyl trifluoromethanesulfonate.

A solution of cyclopentenyl trifluoromethanesulfonate (7.7 g, 35.6 mmol) in 1,4-dioxane (105 ml) and water (50 ml) was degassed and then filled with argon. Add tert-butyl 4-(4,4,5,5-tetramethyl-1,3dioxozan-2-yl)-5,6-di1ldro11iridin-1(211)-carboxysate (6.0 g, 19.4 mmol) , Pd(PPh ₃ ) ₄ (1.02 g, 0.88 mmol) and sodium carbonate (10.3 g, 97.1 mmol). Stir the reaction mixture overnight at 90°C and then cool to ambient temperature. The mixture was poured into water and extracted with ethyl acetate. The combined organic layers were washed with brine, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure. Purification by silica gel flash column chromatography (ethyl acetate/hexanes = 1:99 to 5:95) afforded tert-butyl4-cyclopentenyl-5,6-dihydropyridine-1(2H)-carboxylate.

1H NMR (400 MHz, CDCl ₃ ): δ 5.73 (br, 1H), 5.56 (br, 1H), 4.00 (br, 2H), 3.45 (br, 2H) , 2.47 (m, 4H), 2.33 (bb, 2H), 1.94 (m, 2H), 1.48 (s, 9H).

Pd/C (0.6 g) was added to a solution of tert-butyl-4-cyclopentenyl-5,6-dihydropyridine-1(2H)-carboxylate (3.3 g, 15.3 mmol) in methanol (30 ml). Hydrogenate the mixture under 15 psi for 5 hours. Filter the mixture and concentrate the filtrate under reduced pressure to give tert-butyl 4-cyclopentylpiperidine-1-carboxylate.

To a solution of tert-butyl 4-cyclopentylpiperidine-1-carboxylate (2.9 g, 11.5 mmol) in ethyl acetate (15 ml) was added EtOAc/HCl (4N, 20 ml). The mixture was stirred for 2 hours and the solvent was removed. The resulting product was washed with diethyl ether to obtain 4-cyclopentylpiperidine hydrochloride.

1H NMR (400 MHz, D ₂ O): δ 3.25 (m, 2H), 2.75 (m, 2H), 1.84 (m, 2H), 1.61 (m, 2H), 1. 45 (m, 8H), 0.96 (m, 2H).

Mix 4-cyclopentylpiperidine hydrochloride (1.4 g, 9.5 mmol) and 1(1H-imidazol-1-ylsulfonyl)-3-methyl-1H-imidαzol-3-ium trifluoromethanesulfonate (3.45 g, 9.5 mmol ) in CH3CN (20 ml). Stir the mixture at ambient temperature overnight. The solvent was removed. Purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:9 to 1:3) to give 1-(III-imidazol-1-ylsulfonyl)-4-cyclopentylpiperidine.

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1H NMR (400 MHz, CDCl ₃ ): δ 7.91 (s, 1H), 7.24 (s, 1H), 7.14 (s, 1H), 3.87 (m, 2H), 2.51 (m, 2H), 1.83 (m, 4H), 1.54 (m, 5H), 1.33 (m, 2H), 1.08 (m, 3H).

To a solution of 1-(1I-imidazol-1-ylsulfonyl)-4-cyclopentylpiperidine (500 mg, 1.8 mmol) in DCM (5 ml) was added MeOTf (0.3 g, 1.8 mmol). Stir the reaction mixture at ambient temperature overnight. The solvent was removed and the resulting product was washed with diethyl ether to give 1-((4-cyclopentylpiperidin-1-yl)sulfonyl)-3-methyl1H-imidazol-3-ium trifluoromethanesulfonate.

1H NMR (400 MHz, CD ₃ OD): δ 9.59 (s, 1H), 8.02 (s, 1H), 7.78 (s, 1H), 4.00 (s, 3H), 3. 95 (m, 2H), 2.87 (m, 2H), 1.81 (m, 4H), 1.54 (m, 6H), 1.23 (m, 4H).

The following compounds were synthesized similarly:

1-((4-cyclohexylpiperidin-1-yl)sulfonyl)-3-methyl-1H-imidazol-3-ium trifluoromethanesulfonate (intermediate for compound A65);

3-methyl-1-((4-(tetrahydro-2H-pyran-4-yl)piperidin-1 -yl)sulfonyl)-1Nimidazol-3-ium trifluoromethanesulfonate (intermediate for compound A78).

Method 25.

methanol, hydroxylamine-HC1, DCM, sodium acetate

9-(cyclohexylmethyl)-1-((4-(dimethylamino)phenyl)sulfonyl)-5-((4-phenylpiperidin-1yl)sulfonyl)-1,5,9-triazacyclododecan-3-one (19 mg, 0.028 mmol ) in DCM (0.5 ml) and methanol (0.05 ml), hydroxylamine hydrochloride (5.8 mg, 0.083 mmol) and sodium acetate (6.8 mg, 0.083 mmol) were added. Stir the mixture for 6 hours at ambient temperature, then concentrate and purify by flash column chromatography (0-60% hexanes (1% DEA)/ethyl acetate) to give the oxime 9-(cyclohexylmethyl)-1-((4-( dimethylamino)phenyl)sulfonyl)-5-((4-phenylpiperidin-1-yl)sulfonyl)1,5,9-triazacyclododecan-3-one (compound A75).

MS (ESI) for C ₃₅ H ₅₄ N ₆ O ₅ S ₂ , experiment 703 [M+H]+.

Method 26.

A solution of 1-chloro-2-iodobenzene (5.0 g, 21.1 mmol) in DMF (150 ml) was degassed and then filled with argon. Add tert-butyl 4-(4,4,5,5-tetramethyl-1,3-dioxolan-2-yl)-5,6-dihydropyridine-1(2I)carboxylate (7.13 g, 23.0 mmol ), Pd(dppf)Cl ₂ (1.7 g, 2.1 mmol) and potassium carbonate (8.69 g, 63.0 mmol). Stir the reaction mixture overnight at 110°C and then cool to ambient temperature. The mixture was poured into water and extracted with ethyl acetate. The combined organic layers were washed with brine, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure. Purification by flash column chromatography on silica gel (ethyl acetate/hexanes = 1:9 to 1:4) gave tert-butyl 4-(2-chlorophenyl)-5,6-dihydropyridine-1(2H)-carboxylate.

1H NMR (400 MHz, CDCl ₃ ): δ 7.37 (m, 1H), 7.18 (m, 3H), 5.67 (m, 1H), 4.06 (m, 2H), 3.64 (m, 2H), 2.46 (usirh 2H), 1.51 (s, 9H).

To a solution of tert-butyl 4-(2-chlorophenyl)-5,6-dihydropyridine-1(2H)-carboxylate (2.0 g, 6.8 mmol) in MeOH (20 ml) was added HCl/MeOH (3 M , 20 ml). The mixture was stirred at ambient temperature for 2 hours. The resulting solution of 4-(2-chlorophenyl)-1,2,3,6-tetrahydropyridine hydrochloride was used directly in the next step.

PtO ₂ (0.1 g) was added to a solution of 4-(2-chlorophenyl)piperidine hydrochloride in MeOH/HCl. Hydrogenate the mixture at ambient temperature under 15 psi (103 kPa) for 3 hours. Filter the catalyst and concentrate the filtrate to give the product.

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1H NMR (400 MHz, DMSO-06): δ 9.19 (bb, 2H), 7.37 (m, 1H), 7.36 (m, 4H), 3.44 (m, 1H), 3.33 (m, 2H), 3.08 (m, 2H), 1.95 (m, 4H).

Mix the free base (0.95 g, 4.8 mmol) and 1-(III-imidazol-1ylsulfonyl)-3-methyl-1H-imidazol-3-ium trifluoromethanesulfonate (1.76 g, 4.8 mmol) in CH3CN (10 ml). Stir the mixture at ambient temperature overnight. The solvent was removed and the residue was purified by silica gel column chromatography to give 1-(1H-imidazol-1-ylsulfonyl)-4-(2chlorophenyl)piperidine.

1H NMR (400 MHz, CDCl ₃ ): δ 8.01 (s, 1H), 7.36 (m, 2H), 7.18 (m, 4H), 4.07 (m, 2H), 3.06 (m, 1H), 2.74 (m, 2H), 1.99 (m, 2H), 1.81 (m, 2H).

To a solution of 1-(III-imidazol-1-ylsulfonyl)-4-(2-chlorophenyl)piperidine (501 mg, 1.5 mmol) in DCM (5 ml) was added MeOTf (0.27 g, 1.63 mmol) . Stir the mixture at ambient temperature overnight. The solvent was removed. The resulting product was washed with diethyl ether to obtain 1-((4-(2-chlorophenyl)piperidin-1-yl)sulfonyl)-3-methyl-1H-imidazol-3ium trifluoromethanesulfonate (intermediate for compound A57).

1H NMR (400 MHz, CD3OD): δ 9.65 (s, 1H), 8.08 (s, 1H), 7.82 (s, 1H), 7.39 (m, 4H), 4.16 ( m, 2H), 4.02 (s, 3H), 3.31 (m, 3H), 2.03 (m, 2H), 1.85 (m, 2H).

The following compounds were synthesized similarly:

1-((4-(3-chlorophenyl)piperidin-1-yl)sulfonyl)-3-methyl-111-imidazolium-3 trifluoromethanesulfonate (intermediate for compound A58);

1-((4-(4-chlorophenyl)piperidin-1-yl)sulfonyl)-3-methyl-1 H-imidazolium-3 trifluoromethanesulfonate (intermediate for compound A72).

Method 27.

To a solution of 2-(3-chlorophenyl)ethanamine (149 g, 0.95 mol) and TEA (144 g, 1.42 mol) in DCM (1.5 L) was added dropwise ethyl carbonochloridate (107 g, 1. 14 mol) at 0°C. The reaction mixture was stirred for 3 hours and then washed with 1N. aqueous HCl (1 l) and saturated aqueous NaHCO ₃ (600 ml). Dry the layer in DCM over anhydrous sodium sulfate and concentrate to give methyl 3chlorophenethyl carbamate.

1H NMR (400 MHz, CDCl ₃ ): δ 7.27 (m, 3H), 7.08 (m, 1H), 4.77 (m, 1H), 3.78 (s, 3H), 3.44 (m, 2H), 2.80 (m, 2H).

Dissolved methyl 3-chlorophenethylcarbamate (193 g, 0.9 mol) in CF ₃ SO ₃ H (1.36 kg). Heated the mixture at 120°C overnight. Cool the mixture to ambient temperature and then pour into ice-water mixture (4 L). The resulting product was collected by filtration and washed with diethyl ether to give 6-chloro-3,4-dihydroisoquinolin-1(2H)-one.

1H NMR (400 MHz, CDCl ₃ ): δ 8.01 (m, 1H), 7.35 (m, 1H), 7.25 (m, 2H), 6.21 (bb, 1H), 3 .60 (m, 2H), 3.01 (m, 2H).

To a solution of 6-chloro-3,4-dihydroisoquinolin-1(2H)-one (33 g, 182 mmol) in THF (330 ml) was added BH ₃ -Me ₂ S (73 ml, 729 mmol) dropwise. The reaction mixture was refluxed overnight. The mixture was cooled to ambient temperature, and then the reaction was quenched with 6H. aqueous HCl (300 ml). Remove THF under reduced pressure and reflux the remaining solution overnight. The mixture was concentrated to a certain volume, and then made alkaline with 2N. aqueous NaOH. The resulting mixture was extracted with DCM. Dry the layer in DCM over anhydrous sulfate

- 58 045973 sodium and concentrated to give 6-chloro-1,2,3,4-tetrahydroisoquinoline.

To a solution of 6-chloro-1,2,3,4-tetrahydroisoquinoline (18.5 g, 0.11 mol) in CH3CN (180 ml) was added the compound, 2,3-dimethyl-1-((2-methyl- 1 H-imidazol-1 -yl)sulfonyl)-1 H-imidazol3-ium (43.5 g, 0.11 mol, see: J. Org. Chem. 2002, 68, 115-119.). The reaction mixture was stirred overnight at 30°C. The solvent was removed and the residue was purified by silica gel column chromatography (EtOAc/petroleum ether = 1:1) to give 6-chloro-2-(2-methyl-N-imidazol-1-ylsulfonyl)1,2,3,4-tetrahydroisoquinoline .

1H NMR (400 MHz, CDCl ₃ ): δ 7.21 (m, 1H), 7.18 (s, 1H), 7.02 (m, 1H), 6.94 (m, 1H), 4.45 (s, 2H), 3.62 (m, 2H), 2.92 (m, 2H), 2.67 (s, 3H).

To a solution of 6-chloro-2-(2-methyl-III-imidazol-1-ylsulfonyl)-1,2,3,4-tetrahydroisoquinoline (25.5 g, 82 mmol) in DCM (260 ml) was added CF ₃ SO ₃ Me (13.45 g, 82 mmol). Stir the reaction mixture at ambient temperature overnight. The solvent was removed and the residue was washed with diethyl ether to obtain 1-(6-chloro-3,4-dihydroisoquinolin-2(1H)ylsulfonyl)-2,3-dimethyl-N-imidazol-3-ium trifluoromethanesulfonate.

In a solution of 1-(6-chloro-3,4-dihydroisoquinolin-2(III)-ylsulfonyl)-2,3dimethyl-III-imidazol-3-ium trifluoromethanesulfonate (8.7 g, 18.4 mmol) in CH ₃ CN (100 ml) (I.)-K-(3-amino-2methylpropyl)-4-(dimethylamino)benzenesulfonamide (5.0 g, 18.4 mmol) was added. The reaction mixture was stirred overnight at 30°C. The solvent was removed and the residue was purified by silica gel column chromatography (EtOAc/petroleum ether = 1:3 to 1:1) to give N)-6-chloro-N-(3-((4(dimethylamino)phenyl)sulfonamido)- 2-methylpropyl)-3,4-dihydroisoquinoline-2(1H)-sulfonamide (compound B10).

1H NMR (400 MHz, CDCl ₃ ): δ 7.85 (m, 2H), 7.43 (m, 2H), 7.18 (m, 2H), 7.03 (m, 1H), 4.35 (s, 2H), 3.48 (m, 2H), 3.18 (s, 6H), 3.15 (m, 1H), 2.95 (m, 6H), 2.85 (m, 1H) , 1.89 (m, 1H), 0.89 (m, 3H).

Dissolved ^)-6-chloro-M-(3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-3,4-dihydroisoquinoline-2(III)-sulfonamide (300 mg, 0.6 mmol ) in NMP (3 ml) and NaH (60%, 96 mg, 2.4 mmol) was added. The mixture was stirred for 30 min at ambient temperature and then heated at 80°C for 20 min. Cool the mixture to ambient temperature. A solution of ((cyclohexylmethyl)azandiyl)-bis-(propane-3,1-diyl)dimethanesulfonate (231 mg, 0.6 mmol) in NMP (1 ml) was added. The reaction mixture was heated at 80°C for 1 hour, then cooled to ambient temperature and quenched with water (10 ml). The resulting mixture was extracted with ethyl acetate (15 ml). The extract was washed with saline and concentrated. Purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:8 to 1:3) to give compound A87.

MS (ESI) for C ₃₄ H ₅₂ ClN ₅ O ₄ S ₂ , experiment 695 [M+H]+.

1H NMR (400 MHz, CDCl ₃ ): δ 7.59 (d, J=8.8 Hz, 2H), 7.15 (m, 2H), 7.00 (m, 1H), 6.68 (d , J=8.8 Hz, 2H), 4.34 (m, 2H), 3.71 (m, 2H), 3.45 (m, 2H), 3.17 (m, 3H), 3.05 (s, 6H), 2.88 (m, 4H), 2.25 (m, 2H), 2.15 (m, 6H), 1.95 (m, 2H), 1.65 (m, 8H) , 1.23 (m, 6H), 0.95 (m, 5H).

The following compounds were synthesized similarly:

No. IUPAC [M+N] + Sh NMR Bll N-[(2R)-3-{[(6-xnop-l,2,3,4tetrahydroisoquinolin-2yl)sulfonyl]amino}-2methylpropyl]-4(dimethylamino)benzene-1 sulfonamide 501 A90 4- {[(3 8)-9-(cyclohexylmethyl)3-methyl-5 - {[(3 S)-3 phenylpyrrolidin-1 yl]sulfonyl}-1,5,9triazacycle ododecan-1yl]sulfonyl}- N, Ndimethylaniline 674 A91 4-{[(3 8)-9-(cyclohexylmethyl)3-MeTmi-5-{[(3R)-3phenylpyrrolidin-1 yl]sulfonyl}-1,5,9triazacyclic ododecan-1yl]sulfonyl}-N, Ndimethylaniline 674 A92 4-{[(3 8)-9-(cyclohexylmethyl)3-methyl-5-[(4phenoxypiperidine-1 - 704

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yl)sulfonyl]-1,5,9triazacycle ododecan-1yl]sulfonyl}-N, Ndimethylaniline A93 4- {[(3 8)-9-(cyclohexylmethyl)3-methyl-5-{[4-(pyrimidin-2yl)piperazin-1 -yl]sulfonyl } - 1,5,9-triazacycle od od ekai-1 yl] sulfonyl }-N, Ndimethylaniline 691 A94 4- {[(3 8)-9-(cyclohexylmethyl)5 - {[4-(2-fluorophenyl)piperazine- 1-yl]sulfonyl}-3-methyl-1,5,9triazacyclode decan-1yl]sulfonyl}-N, Ndimethylaniline 707 A95 4-{[(3 8)-9-(cyclohexylmethyl)5 - {[4-(3-fluorophenyl)piperazin1 -yl]sulfonyl}-3-methyl-1,5,9triazacyclododecan-1yl]sulfonyl}-N , Ndimethylaniline 707 A96 4- {[(3 8)-9-(cyclohexylmethyl)5 - {[4-(4-fluorophenyl)piperazine- 1-yl]sulfonyl}-3-methyl-1,5,9triazacyclode decan-1yl]sulfonyl}-N, Ndimethylaniline 707 A97 4- {[(3 8)-9-(cyclohexylmethyl)3-methyl-5 - {[4-(2methylphenyl)piperazin-1yl]sulfonyl}-1,5,9triazacyclodecan-1yl]sulfonyl}-N , Ndimethylaniline 703 A98 4-{[(3 8)-9-(cyclohexylmethyl)3-methyl-5-{[4-(3methylphenyl)piperazin-1 yl]sulfonyl}-1,5,9triazacyclododecan-1yl]sulfonyl}-N , Ndimethylaniline 703 A99 4-{[(3 8)-9-(cyclohexylmethyl)3-methyl-5-{[4-(4methylphenyl)piperazine-1 - 703

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yl]sulfonyl}-1,5,9triazacycle ododecan-1yl]sulfonyl}-N, Ndimethylaniline A100 4-{[(3S)-5-[(4- benzoylpiperazin-1 yl)sulfonyl]-9(cyclohexylmethyl)-3-methyl1,5,9-triazacyclododo ekan-1 yl]sulfonyl}-N, Ndimethylaniline 717 A101 4-{[(3 8)-9-(cyclohexylmethyl)3-methyl-5-[(3-phenylazetidin-1yl)sulfonyl]-1,5,9triazacyclododecan-1yl]sulfonyl}-N, Ndimethylaniline 660 A102 4-{[(3 8)-9-(cyclohexylmethyl)5-{[4-(4-fluoro-2methylphenyl)piperazin-1yl]sulfonyl}-3-methyl-1,5,9triazacyclodeodecan-1yl]sulfonyl }-N, Ndimethylaniline 721 A76 4-{ [(ZK)-9-(cyclohexylmethyl)3 -methyl-5 - [(4-phenylpiperide in1 -yl)sulfonyl]-1,5,9triazacyclododecan-1yl]sulfonyl }-N, Ndimethylaniline 688 Ή NMR (400 MHz, CDC1 ₃ ): δ 7.60 (d, 7=8.8 Hz, 2H), 7.33 (m, 5H), 6.82 (d, 7=8.8 Hz, 2H ), 3.77 (w, 4H), 3.28 (m, 1H), 3.18 (m, ZN), 3.05 (s, 6H), 2.88 (m, 4H), 2.61 (w, 1H), 2.41 (m, 1H), 2.21 (m, 4H), 2.01 (m, ZN), 1.85 (m, 8H), 1.25 (w, 6H) , 0.93 (w, ZN), 0.82 (m, 4H). A77 4- {[(3 8)-9-(cyclohexylmethyl)3 -methyl-5 - [(4-phenylpiperide in1 -yl)sulfonyl]-1,5,9triazacyclic odecane-1yl]sulfonyl}-N, Ndimethylaniline 688 Ή NMR (400 MHz, CDCh): δ 7.60 (d, 7=8.8 Hz, 2H), 7.33 (m, 5H), 6.82 (d, 7=8.8 Hz, 2H) , 3.77 (w, 4H), 3.28 (m, 1H), 3.18 (w, ZN), 3.05 (s, 6H), 2.88 (m, 4H), 2.61 ( w, 1H), 2.41 (m, 1H), 2.21 (m, 4H), 2.01 (w, ZN), 1.85 (m, 8H), 1.25 (w, 6H), 0.93 (w, ZN), 0.82 (m, 4H).

Method 28.

To a solution of benzylamine (37.7 g, 0.350 mol) in CH ₃ CN (1 L) was added 3-chloropropan-1-ol (100 g, 1.06 mol) and Na ₂ CO ₃ (131 g, 1.24 mol ). The reaction mixture was refluxed for 50 hours and then cooled to ambient temperature. The mixture was filtered and the filtrate was concentrated under reduced pressure. Purify the residue by silica gel column chromatography (MeOH/DCM = 1:99 to 3:97) to give 3,3'-(benzylazandiyl)dipropan-1-ol.

1H NMR (400 MHz, CDCl ₃ ): δ 7.34 (m, 5H), 3.70 (m, 4H), 3.61 (s, 2H), 3.48 (s, 2H), 2.68 (m, 4H), 1.78 (m, 4H).

Pd/C (10 g) was added to a solution of 3,3'-(benzylazandiyl)dipropan-1-ol (55 g) in MeOH (500 ml). The mixture was hydrogenated at ambient temperature under a pressure of 15 kg for 6 hours. The mixture was filtered and the filtrate was concentrated to obtain 3,3'-azandiyldipropan-1-ol.

1H NMR (400 MHz, CDCl ₃ ): δ 3.78 (m, 4H), 3.46 (s, 2H), 2.84 (m, 4H), 1.75 (m, 4H).

To a solution of 3,3'-azandiyldipropan-1-ol (15 g, 113 mmol) in DCM (300 ml) was added cyclohexanecarbaldehyde (18.8 g, 141 mmol) and NaBH(OAc) ₃ (72.5 g, 0. 34 mol). Stir the mixture for 0.5 h and add HOAc (20.4 g, 0.34 mol). Stir the reaction mixture at ambient temperature

- 61 045973 Wednesdays overnight. Quench the reaction with water (150 ml), and then adjust the pH of the mixture to 12 using NaOH. The organic layer was separated, dried over anhydrous sodium sulfate and concentrated. Purify the residue by silica gel column chromatography (MeOH/DCM = 3:97 to 1:9) to give 3,3'-((cyclohexylmethyl)azandiyl)-bis-(propan-1-ol).

^1H NMR (400 MHz, CDCl3): δ 3.75 (m, 4H), 2.64 (m, 4H), 2.18 (m, 2H), 1.75 (m, 9H), 1.55 (m, 1H), 1.21 (m, 3H), 0.89 (m, 1H).

In a solution of the compound, 3,3'-((cyclohexylmethyl)azandiyl)-bis-(propan-1-ol) (1.2 g, 5.2 mmol) and TEA (1.0 g, 10.4 mmol) in DCM (15 ml) was added MsCl (1.18 g, 10.4 mmol) at 0°C. The reaction mixture was stirred for 4 hours and then the reaction was quenched with water (15 ml). The layer was separated in DCM, dried over anhydrous sodium sulfate and concentrated to give the compound, ((cyclohexylmethyl)azandiyl)-bis-(propan-3,1-diyl)dimethanesulfonate. The dimesylate was used directly without further purification.

Method 29.

BosjtO

N _g M' ^Lch ^ ^L MN _g

THF

TEA, DMF

To a solution of propane-1,3-diamine (100 g, 1.35 mol) in THF (800 ml) was added a solution of Boc ₂ O (73.6 g, 0.34 mol) in THF (200 ml) at 0~10 °C. The reaction mixture was stirred for 3 hours and water (1 L) was added. The resulting mixture was extracted with ethyl acetate (500 mlx2). Combine the organic layers and concentrate to a volume of 400 ml. Hexane (300 ml) was added and 15% aqueous oxalic acid (1 L) was added to the resulting solution. The mixture was stirred for 0.5 hours. The organic layer was decanted and the pH of the aqueous layer was adjusted to 10 3N. aqueous NaOH. The resulting mixture was extracted with DCM (600 mlx2). Dry the combined extracts over anhydrous sodium sulfate and concentrate to give tert-butyl-3-aminopropylcarbamate.

^1H NMR (400 MHz, CDCi ₃ ): δ 5.03 (b.e., 1H), 3.23 (b.e., 2H), 2.69 (m, 2H), 1.71 (m, 2H ), 1.44 (s.9H).

To a stirred mixture of benzyl 3-bromopropyl carbamate (4.2 g, 15.5 mmol) triethylamine (4.70 g, 46.5 mmol) was added dropwise to give tert-butyl 3-aminopropyl carbamate (2.7 g, 15 .5 mmol) in DMF (50 ml) at ambient temperature. Heat the reaction mixture at 70°C for 1 hour. Remove most of the DMF in vacuo and dilute the remaining mixture with water (120 ml) and wash with diethyl ether (150 ml x 4) to remove most of the dialkylated by-product. The pH of the aqueous layer was adjusted to 11 1N. aqueous NaOH and extracted with diethyl ether (200 ml). Wash the extract with water (3 x 150 ml) to remove unreacted tert-butyl-3aminopropylcarbamate, dry with anhydrous sodium sulfate and concentrate to obtain tert-butyl (3-((3-(((benzyloxy)carbonyl)amino)propyl)amino)propyl)carbamate .

1H NMR(400 MHz, CDCl ₃ ): δ 7.36 (m, 5H), 5.60 (b, 1H), 5.11 (m, 3H), 3.30 (m, 2H), 3 .20 (m, 2H), 2.68 (m, 4H), 1.70 (m, 5H), 1.44 (s, 9H).

To a solution of tert-butyl-(3-((3-(((benzyloxy)carbonyl)amino)propyl)amino)propyl)-carbamate (100 mg, 0.27 mmol) in DCM (5 ml) was added cyclohexanecarbaldehyde (31 mg , 0.27 mmol) and NOAc (30 mg, 1.1 mmol). The mixture was stirred for 0.5 h and NaBH(OAc) ₃ (235 g, 1.1 mmol) was added in one portion. Stir the reaction mixture for 4 hours. TLC analysis indicated 50% conversion and another portion of cyclohexanecarbaldehyde (15 mg, 0.13 mmol) was added. Stir the reaction mixture overnight at ambient temperature. Add water (10 ml) to quench

- 62 045973 reaction and adjusted the pH of the mixture to 2 by adding HOAc. The mixture was stirred for another 0.5 hour and then adjusted to pH=12 with aqueous NaOH (1N). The organic layer was separated and concentrated to give tert-butyl (3((3-(((benzyloxy)carbonyl)amino)propyl)(cyclohexylmethyl)amino)propyl)carbamate.

1H NMR (400 MHz, CDCl ₃ ): δ 7.27 (m, 5H), 6.12 (br, 1H), 5.55 (br, 1H), 5.01 (s, 3H), 3.34 (m, 2H), 3.18 (m, 2H), 3.08 (m, 2H), 2.80 (m, 4H), 1.65 (m, 9H), 1.44 (s, 9H) , 1.12 (m, 6H).

In a solution of tert-butyl-(3-((3-(((benzyloxy)carbonyl)amino)propyl)-(cyclohexylmethyl)amino)propyl)carbamate (0.50 g, 1.1 mmol) in MeOH (5 ml) Pd/C (0.1 g) was added. Hydrogenate the mixture under 15 psi for 3 hours. Filter the mixture and concentrate the filtrate to give tert-butyl (3-((3-aminopropyl)-(cyclohexylmethyl)amino)propyl)carbamate.

1H NMR (400 MHz, CDCl3): δ 5.78 (br, 1H), 3.19 (br, 2H), 2.78 (m, 2H), 2.40 (m, 4H), 2.13 ( m, 2H), 1.98 (broad, 2H), 1.67 (m, 9H), 1.44 (s, 9H), 1.12 (m, 4H), 0.89 (m, 2H).

3-Methyl-1-(4 -phenylpiperidin-1-ylsulfonyl)-1H-imidazol-3-ium (1.47 g, 3.7 mmol). Stir the reaction mixture at ambient temperature overnight. The solvent was removed and the residue was purified by silica gel column chromatography to give tert-butyl-3-((cyclohexylmethyl)(3-(4-phenylpiperidin-1-sulfonamido)propyl)amino)propylcarbamate.

TFA (10 ml). Stir the reaction mixture at ambient temperature for 3 hours. Remove the solvent and dissolve the residue in DCM (50 ml). The solution was made alkaline with 1N. aqueous NaOH. Dry the layer in DCM over anhydrous Na ₂ SO ₄ and concentrate to give ^(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4phenylpiperidine-1-sulfonamide.

To a solution of N(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4-phenylpiperidine-1sulfonamide (90 mg, 0.20 mmol) in DCM (5 ml) was added TEA (40 mg, 0.4 mmol ). Cool the mixture to 0°C and add a solution of 4-(dimethylamino)-2,6-difluorobenzene-1-sulfonyl chloride (60 mg, 0.24 mmol) in DCM (1 ml). Stir the reaction mixture at ambient temperature for 2 hours. Dilute the mixture with DCM (30 ml) and then wash with water (20 ml). Dry the organic layer over anhydrous Na ₂ SO ₄ and concentrate. The residue was purified by column chromatography on silica gel to obtain N(3-((cyclohexylmethyl)(3-(4-(dimethylamino)phenylsulfonamido)propyl)amino)propyl)4-phenylpiperidine-1-sulfonamide.

To a solution of Na(3-((cyclohexylmethyl)(3-(4-(dimethylamino)phenylsulfonamido)propyl)amino)propyl)-4-phenylpiperidine-1-sulfonamide (80 mg, 0.123 mmol) in NMP (2 ml) was added NaH (12 mg, 0.29 mmol). Heat the mixture at 80°C for 0.5 h. Cool the mixture to ambient temperature and add a solution of 3-chloro-2-(chloromethyl)prop-1-ene (15 mg, 0.12 mmol) in NMP (0. 1 ml). Heat the reaction mixture at 80°C for 2 hours. Cool the mixture to ambient temperature and add water (20 ml). The resulting mixture was extracted with ethyl acetate (15 ml). Dry the organic layer over anhydrous sodium sulfate and concentrate. Purify the residue by silica gel column chromatography (ethyl acetate/hexane = 10:30 to 20:10) to give compound A86.

MS (ESI) for C36H53F2N5O4S2, experiment 733 [M+H]+.

1H NMR (400 MHz, CDCl ₃ ): δ 7.31 (m, 5H), 6.18 (m, 2H), 5.25 (m, 2H), 4.05 (s, 2H), 3.87 (m, 4H), 3.20 (m, 4H), 3.05 (s, 6H), 2.88 (m, 2H), 2.41 (m, 4H), 2.05 (m, 5H) , 1.75 (m, 9H), 1.27 (m, 6H), 0.85 (m, 2H).

Method 30.

To a solution of 3,5-difluoro-4-iodo-X-dimethylaniline (500 mg, 1.77 mmol) in diethyl ether (5 ml), n-BuLi (0.71 ml, 1.77 mmol) was added dropwise at - 78°C. The mixture was stirred for 0.5 h at -78°C. Sulfuryl dichloride (358 mg, 1.5 mmol) was added. The reaction mixture was stirred for another 0.5 hours. The reaction was quenched with water (10 ml), and then the mixture was extracted with ethyl acetate (15 ml). Dry the organic layer over anhydrous Na ₂ SO ₄ and concentrate. Purify the residue by silica gel column chromatography to give the crude product, which is further recrystallized from hexane/EtOAc (10:1) to give 4-(dimethylamino)-2,6-difluorobenzene-1-sulfonyl chloride.

1H NMR (400 MHz, CDCl3): δ 6.21 (s, 1H), 6.18 (s, 1H), 3.09 (s, 6H).

Method 31.

Hydrochloride was added to a suspension of 1,3-dichloropropan-2-one (5.0 g, 39.5 mmol) in H ₂ O (50 ml).

- 63 045973

O-methylhydroxylamine (3.5 g, 41.5 mmol) and stir the reaction mixture at ambient temperature for 1 hour. Separate the organic layer, dilute with diethyl ether and concentrate in vacuo (<30°C) to give O-methyloxime 1 ,3-dichloropropan-2-one.

'll NMR (400 MHz, CDC1 ₃ ): δ 4.33 (s, 2H), 4.27 (s, 2H), 3.94 (s, 3H).

Method 32.

K (8)-6-chloro-K-(3 -((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-3,4-dihydroisoquinoline-2(1H)-sulfonamide (100.0 mg, 0.200 mmol) in dry NMP (3 ml) at 0°C, add NaH (24.0 mg, 0.600 mmol) and stir the mixture at ambient temperature for 20 min, during which time the cloudy mixture becomes almost clear. Iodomethane (27 µl, 0.440 mmol) was added and the mixture was left to stir at ambient temperature for 1 hour, the completeness of the reaction did not exceed ~50%, so the same amounts of NaH and methyl iodide were added again, and after another 1 hour at ambient temperature the reaction was ending. Saline (10 ml) and EA (10 ml) were added. Extract the aqueous layer with EA (1x5 ml) and combine the organic layers, wash with brine (3x10 ml), dry with sodium sulfate, filter and concentrate. FCC (hexanes/EA 0-70%) resulted in compound B12.

MS (ESI) for C ₂₃ H ₃₃ C1N ₄ O ₄ S ₂ , experiment 529 [M+H]+.

Method 33.

SOOM"

neither.

MeOH

TsOH cross-crystal

_Ng . Pd/C acetone mixture)

THF

ΡΡΙΊ3, toluene none soome TEA, DHM

To a solution of methyl methacrylate (300 g, 3.0 mol) in MeOH (1.5 L) was added N)-1-phenylethanamine (363 g, 3 mol). The reaction mixture was refluxed for 5 days. Concentrate the mixture and purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:9 to 1:1) to give methyl 2-methyl-3-(^)-1-phenylethylamino)propanoate.

^1H NMR (400 MHz, CDC13): δ 7.25 (m, 5H), 3.75 (m, 1H), 3.69 (s, 3H), 2.75 (m, 3H), 1.39 (m, 1H), 1.28 (m, 3H), 1.09 (m, 3H).

TsOH-H 2 O (378 g _, 1.99 mol) was added to a solution of methyl 2-methyl-3-(^)-1-phenylethylamino)propanoate (440 g, 1.99 mol) in acetone (6 L). Stir the mixture at ambient temperature for 1 hour. Collect the precipitated solid by filtration and dry to obtain a salt (300 g), which is suspended in acetone (2 L) and reflux for 1 hour. Cool the mixture to ambient temperature medium and collected the resulting precipitate by filtration to obtain a salt, which was suspended in DCM (2 L) and saturated aqueous K ₂ CO ₃ (1.5 L). The mixture was stirred for 0.5 h, the solid dissolved. The layer was separated in DCM, dried over anhydrous Na ₂ SO ₄ and concentrated to give ^)-methyl-2-methyl-3-(^)-1-phenylethylamino)propanoate.

To a solution of N)-methyl-2-methyl-3-(N)-1-phenylethylamino)propanoate (140 g, 0.630 mol) in MeOH (1.4 L) was added HOAc (30 ml) and Pd/C (15 g ). The mixture was hydrogenated under a pressure of 20 kg at 60°C for 6 hours. The catalyst was removed by filtration and the filtrate was concentrated to obtain N-methyl-3amino-2-methylpropanoate.

^1H NMR (400 MHz, CDCI3): δ 5.09 (br, 3H), 3.72 (s, 3H), 3.05 (m, 1H), 2.93 (m, 1H), 2. 75 (m, 1H), 1.23 (d, J=7.2 Hz, 3H).

To a solution of N)-methyl-3-amino-2-methylpropanoate (112 g, 0.63 mol) in dichloromethane (1 L) was added

- 64 045973

TEA (160 g, 1.58 mol) and Boc2O (152 g, 0.69 mol) at 0-5°C. The reaction mixture was allowed to warm to ambient temperature and stirred overnight. The mixture was washed with water (1 l), 2N. aqueous HCl (1 l) and saturated aqueous NaHCO ₃ (1 l), respectively. Dry the organic layer over anhydrous Na ₂ SO ₄ and concentrate. The residue was purified by column chromatography on silica gel to obtain N)-methyl-3-(tert-butoxycarbonylamino)-2-methylpropanoate.

^1H NMR (400 MHz, CDCl3): δ 4.94 (br-s, 1H), 3.70 (s, 3H), 3.31 (m, 2H), 2.70 (m, 1H), 1 .52 (s, 9H), 1.18 (d, J=7.2 Hz, 3H).

LiBH 4 (1.12 g, ₁₀₂ mmol) was added to a solution of N)-methyl-3-(tert-butoxycarbonylamino)-2-methylpropanoate (6 g, 51.3 mmol) in THF (60 ml). Stir the reaction mixture overnight at ambient temperature. The reaction was quenched with water (30 ml), then 1N was added. aqueous HCl, bringing to pH 2-3. The resulting mixture was extracted with ethyl acetate and the extract was washed with saturated aqueous NaHCO ₃ , dried over anhydrous Na ₂ SO ₄ and concentrated to obtain (I)-tert-butyl-3-hydro1<si-2methylpropylcarbamate.

^1H NMR (400 MHz, CDCl3): δ 3.57 (m, 1H), 3.37 (m, 2H), 3.05 (m, 1H), 1.77 (m, 1H), 1.48 (s, 9H), 0.88 (d, J=7.2 Hz, 3H).

PPh ₃ (4.15 g, 15.8 mmol) and isoindoline- 1,3-dione (2.72 g, 18.5 mmol). Cool the mixture to 0~5°C and add dropwise DIAD (3.2 g, 15.8 mmol). Stir the reaction mixture at ambient temperature for 3 hours. Filter off the precipitated solid and concentrate the filtrate. Purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:9 to 1:1) to give N)-tert-butyl-3-(1,3-dioxoisoindolin-2-yl)-2-methylpropylcarbamate.

^1H NMR (400 MHz, CDCl3): δ 7.85 (m, 2H), 7.47 (m, 2H), 5.23 (br 8, 1H), 3.64 (m, 2H), 3 .03 (broad 8, 2H), 2.13 (m, 1H), 1.43 (s, 9H), 0.96 (d, J=6.4 Hz, 3H).

A solution of methylamine (7.8 g, 30 % in MeOH). The reaction mixture was heated at 60°C for 6 hours. The solvent was removed under reduced pressure. Suspend the residue in diethyl ether (300 ml) and then stir for 0.5 hours. Filter the solid and concentrate the filtrate under reduced pressure to give N)-tert-butyl-3-amino-2methylpropylcarbamate.

To a solution of N)-tert-butyl-3-amino-2-methylpropylcarbamate (12.5 g, 66.5 mmol) in DCM (250 ml) was added TEA (10.3 g, 101.7 mmol). Cool the mixture to 0~5°C and add 4-(dimethylamino)benzene-1sulfonyl chloride (16 g, 72 mmol). The reaction mixture was stirred at ambient temperature for 3 hours. The mixture was washed with water and the organic phase was concentrated under reduced pressure. Purify the residue by silica gel column chromatography (EtOAc/petroleum ether = 1:9 to 3:7) to give da-tert-butyl-3-(4-(dimethylamino)phenylsulfonamido)-2-methylpropylcarbamate.

^1H NMR (400 MHz, CDCls): δ 7.70 (d, J=8.8 Hz, 2H), 6.71 (d, J=8.8 Hz, 2H), 5.45 (m, 1H ), 4.72 (m, 1H), 3.18 (m, 1H), 3.04 (s, 6H), 2.98 (m, 2H), 2.75 (m, 1H), 1.78 (m, 1H), 1.41 (s, 9H), 0.87 (m, J=6.8 Hz, 3H).

To a solution of da-tert-butyl-3-(4-(dimethylamino)phenylsulfonamido)-2-methylpropylcarbamate (21 g, 59.3 mmol) in MeOH (150 ml) was added 4N. HCl/MeOH (150 ml). The mixture was stirred for 3 hours at ambient temperature. The solvent was removed under reduced pressure. Suspend the resulting solid in DCM (200 ml) and add saturated aqueous K ₂ CO ₃ (150 ml). Stir the mixture for 5 minutes and separate the organic layer, dry over anhydrous Na ₂ SO ₄ and concentrate to give the compound, N)-Na-(3-amino-2-methylpropyl)-4-(dimethylamino)benzenesulfonamide.

¹ H NMR (400 MHz, CDCl ₃ ): δ 7.70 (d, J=8.8 Hz, 2H), 6.68 (d, J=8.8 Hz, 2H), 3.04 (s, 6H), 2.98 (m, 1H), 2.75 (m, 2H), 2.58 (m, 1H), 1.78 (m, 2H), 0.87 (m, J=7.2 Hz, 3H).

N)-N-(3-amino-2-methylpropyl)-4-(dimethylamino)benzenesulfonamide (intermediate for compound A76) was synthesized in a similar manner.

Method 34.

To 6-chloro-N(^)-3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-1,2,3,4-tetrahydronaphthalene-2-sulfonamide (100 mg, 200 µmol) in dry NMP (4 ml) at 0°C, add NaH (24 mg, 600 µmol) and stir the mixture at ambient temperature for 20 min, during which time

- 65 045973 the cloudy mixture became almost transparent. Dichloride (21.3 µl, 200 µmol) was added and the mixture was heated to 80°C. After 40 minutes, quench the reaction with brine and extract the aqueous layer with ethyl acetate (2x), wash the combined organic layers with brine (3x), dry with sodium sulfate, filter and concentrate. Column chromatography (0-50% hexanes+1% diethylamine)/ethyl acetate) gave the product (compound B21).

MS (ESI) for C ₂₅ H ₃₅ ClN ₄ O ₄ S ₂ , experiment 555 [M+H]+.

Method 35.

To a solution of 1-(4-fluoro-2-methoxyphenyl)piperazine (1.4 g, 6.8 mmol) in CH3CN (15 ml) was added 1-(III-imidazol-1-ylsulfonyl)-3-methyl-III trifluoromethanesulfonate -imidazol-3-ium (2.7 g, 6.8 mmol). Stir the reaction mixture at ambient temperature overnight. The mixture was then concentrated and the residue was purified by column chromatography to give 1-((1H-imidazol-1yl)sulfonyl)-4-(4-fluoro-2-methoxyphenyl)-piperazine.

To a solution of 1-((III-imidazol-1-yl)sulfonyl)-4-(4-fluoro-2-methoxyphenyl)-piperazine (0.92 g, 2.7 mmol) in DCM (10 ml) was added CF ₃ SO ₃ Me (0.45 g, 2.7 mmol) at 0°C. Stir the reaction mixture at ambient temperature for 5 hours, then concentrate and wash the resulting product with diethyl ether to give 1-((4-(4-fluoro-2methoxyphenyl)piperazin-1-yl)sulfonyl)-3-methyl-111 trifluoromethanesulfonate -imidazol-3-ium.

1-((4-(4 -fluoro-2-methoxyphenyl)piperazin-1yl)sulfonyl)-3-methyl-III-imidazol-3-ium (1.1 g, 2.2 mmol). Stir the reaction mixture at ambient temperature overnight. Concentrate the mixture and purify the residue by column chromatography to obtain ^)-N-(3-((4-(dimethylamino)phenyl)sulfonamido)-2methylpropyl)-4-(4-fluoro-2-methoxyphenyl)piperazine-1-sulfonamide.

In solution of ^)-M-(3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-4-(4-fluoro-2methoxyphenyl)piperazine-1-sulfonamide (771 mg, 1.42 mmol) NaH (140 mg, 3.48 mmol, 60%) was added to NMP (7 ml). The mixture was stirred for 0.5 h at 80°C. Cool the mixture to 70°C and add ((cyclohexylmethyl)azandiyl)-bis-(propane-3,1-diyl)dimethanesulfonate (820 mg, 2.1 mmol). The reaction mixture was stirred for 2 hours at 70°C, then cooled and poured into water. The resulting mixture was extracted with ethyl acetate. Concentrate the extract and purify the residue by column chromatography (petroleum ether/EtOAc = 10:1) to give compound A137.

MS (ESI) for C ₃₆ H ₅₇ FN ₆ O ₅ S ₂ , experiment 737 [M+H]+.

1H NMR (400 MHz, CDCI3): δ 7.62 (d, J=8.8 Hz, 2H), 7.16 (m, 2H), 7.05 (m, 1H), 6.86 (d, J=8.8 Hz, 2H), 4.35 (m, 2H), 3.62 (m, 4H), 3.19 (m, 2H), 3.04 (s, 6H), 2.87 ( m 4H), 2.46 (m, 5H), 1.92 (m, 1H), 1.79 (m, 2H), 1.66 (m, 4h).

- 66 045973

The following compounds were synthesized similarly:

No. IUPAC [M+N] ⁺ A153 tert-butyl-4-{[(3 8)-9-(cyclohexylmethyl)-5-[4- (d imethylamino)benzenesulfonyl]-3 -methyl-1,5,9- triazacyclododecan-1-yl]sulfonyl } piperazine-1-carboxylate 714 A118 2-(4-{ [(3 8)-9-(cyclohexylmethyl)-5-[4- (d ymethylamino)benzenesulfonyl]-3 -methyl-1,5,9triazacyclode decan-1 -yl]sulfonyl } piperazin-1 yl)benzonitrile 715 A154 4- {[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(2methoxyphenyl)piperazin-1 -yl]sulfonyl } -3 -methyl-1,5,9triazacyclododecan-1 -yl]sulfonyl } - N,N-dimethylaniline 720 A155 4- {[(3 8)-9-(cyclohexylmethyl)-3 -methyl-5 -(piperazine-1 sulfonyl)-1,5,9-triazacyclod odekai-1 -yl] sulfonyl } -N, N- dimethylaniline 613 A119 4-{ [(3 8)-9-(cyclohexylmethyl)-5-{ [4-(2,6dimethylphenyl)piperazin-1 -yl]sulfonyl } -3 -methyl-1,5,9triazacyclododecan-1 -yl]sulfonyl }-N,N-dimethylaniline 717 A136 4-{[(3 8)-9-(cyclohexylmethyl)-5 - [(6-ethyl-1,2,3,4tetrahydroisoquinolin-2-yl)sulfonyl]-3-methyl-1,5,9triazacyclododecane-1 - yl]sulfonyl}-N, N-dimethylaniline 688 A161 4-{ [(3 8)-9-(cyclohexylmethyl)-5-{ [6-(4-fluoro-2-methylphenyl)-2,6diazaspiro[3.3] heptan-2-yl]sulfonyl } -3 -methyl- 1,5,9triazacyclododecan-1 -yl]sulfonyl}-N, N-dimethylaniline 733 A162 4-{ [(3 8)-9-(cyclohexylmethyl)-5-{ [(28)-4-(4-fluoro-2methylphenyl)-2-methylpiperazin-1 -yl] sulfonyl } -3 -methyl-1, 5,9triazacyclododecan-1 -yl]sulfonyl} -N,N-dimethylaniline 735 A163 4-{ [(3 8)-9-(cyclohexylmethyl)-5-{ [(2R)-4-(4-fluoro-2- 735

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methylphenyl)-2-methylpiperazin-1 -yl]sulfonyl}-3-methyl-1,5,9triazacyclododecan-1-yl]sulfonyl}-T\G, N-dimethylaniline A166 4- {[(3 8)-9-(cyclohexylmethyl)-3 -methyl-5 - [(5 -methyl-1,2,3,4tetrahydroisoquinolin-2-yl)sulfonyl] -1,5,9- triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethylaniline 674 A138 4- {[(3 8)-9-(cyclohexylmethyl)-5 - [(5-methoxy-1,2,3,4tetrahydroisoquinolin-2-yl)sulfonyl]-3 -methyl-1,5,9triazacyclododecane-1- yl]sulfonyl)-X, N-dimethylaniline 690 A151 2-{ [(3 8)-9-(cyclohexylmethyl)-5-[4- (d imethylamino)benzenesulfonyl]-3-methyl-1,5,9triazacyclod decan-1-yl]sulfonyl)-1,2,3,4tetrahydroisoquinoline-5-carbonitrile 685 A170 4- {[(3 8)-9-(cyclohexylmethyl)-3 -methyl-5 - [(1 -methyl-1,2,3,4tetrahydroisoquinolin-2-yl)sulfonyl] -1,5,9- triazacyclododecan-1-yl]sulfonyl)-X, N-dimethylaniline 674 A172 4- {[(3 8)-9-(cyclohexylmethyl)-3 -methyl-5 - [(3 -methyl-1,2,3,4tetrahydroisoquinolin-2-yl)sulfonyl] -1,5,9- triazacyclododecan-1-yl]sulfonyl)-18G, N-dimethylaniline 674 A173 4- {[(3 8)-9-(cyclohexylmethyl)-3 -methyl-5 -({ 4-[2(trifluoromethoxy)phenyl]piperazin-1 -yl } sulfonyl)-1,5,9triazacyclododecan-1-yl ]sulfonyl)-X, N-dimethylaniline 773 A174 4- {[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(3,4-d iforo-2methoxyphenyl)piperazin-1 -yl] sulfonyl } -3 -methyl-1,5,9triazacyclododecane- 1-yl]sulfonyl)-18G, N-dimethylaniline 755 A175 4- {[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(3-fluoro-2 methoxyphenyl)piperazin-1 -yl] sulfonyl } -3 -methyl-1,5,9triazacyclododecane-1- yl]sulfonyl)-X, N-dimethylaniline 737 A176 4-{[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(5-fluoro-2methoxyphenyl)piperazin-1-yl]sulfonyl} -3-methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl)-18G, N-dimethylaniline 737 A177 4-{ [(3 8)-9-(cyclohexylmethyl)-5-{ [(3H)-4-(4-fluoro-2methoxyphenyl)-3 -methylpiperazin-1 -yl] sulfonyl } -3 -methyl- 1,5,9-triazacycle od od ekan-1 -yl] sulfonyl } -N,N-dimethylaniline 751 A178 4- {[(3 8)-9-(cyclohexylmethyl)-5 - {[3 -(4-fluoro-2methoxyphenyl)azetidin-1 -yl] sulfonyl } -3 -methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl)-18G, N-dimethylaniline 708 A181 4- {[(3 S)-5 -[(5-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-3-methyl-1,5,9triazacyclododecane-1- yl]sulfonyl)-H N-dimethylaniline 694 A187 (38)-M-benzyl-9-(cyclohexylmethyl)-5-[4(d ymethylamino)benzenesulfonyl]-3-methyl-1,5,9triazacyclodedecane-1-sulfonamide 634 A198 4-{[(3 8)-5-{[4-(2-chlorophenyl)piperazin-1-yl]sulfonyl)-9(cyclohexylmethyl)-3-methyl-1,5,9-triazacyclododecan-1 yl] sulfonyl}-N,N-dimethylaniline 723

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A199 4- {[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(4- methoxyphenyl)piperazin-1-yl]sulfonyl} -3 -methyl-1,5,9- triazacyclododecan-1-yl]sulfonyl}-T4, N-dimethylaniline 719 A201 4-{[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(3 methoxyphenyl)piperazin-1-yl]sulfonyl} -3-methyl-1,5,9triazacyclododecan-1-yl]sulfonyl} -18G, N-dimethylaniline 719 A202 4-{[(3 8)-9-(cyclohexylmethyl)-5 - {[4-(3-fluoro-4methoxyphenyl)piperazin-1-yl]sulfonyl} -3-methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl}-18G, N-dimethylaniline 737 A206 4-{[(3 8)-5-{[4-(2-chloro-4-fluorophenyl)piperazin-1-yl]sulfonyl}- 9-(cyclohexylmethyl)-3-methyl-1,5,9-triazacyclododecan-1yl]sulfonyl}-N,N-dimethylaniline 741 A147 4- {[(4 S)-5 -[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-4-methyl-1,5,9triazacyclododecane-1- yl]sulfonyl}-18G, N-dimethylaniline 694 A148 4-{[(4K)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-4-methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl}-18G, N-dimethylaniline 694 A150 4- {[(2 S)-5 -[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-2-methyl-1,5,9triazacyclododecane-1- yl]sulfonyl}-18G, N-dimethylaniline 694 A149 4-{[(2K)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-2-methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl}-18G, N-dimethylaniline 694 A190 4-{[(3 8)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-[(1S)-1-cyclohexylethyl]-3-methyl-1, 5,9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethylaniline 708 A191 4-{[(3 8)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-[(1R)-1-cyclohexylethyl]-3-methyl-1, 5,9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethylaniline 708 A207 4-{[(3 S)-9-[fl 8)-1-cyclohexylethyl]-5-[[(3C)-4-(4-fluoro-2methoxyphenyl)-3-methylpiperazin-1-yl]sulfonyl} - 3 -methyl- 1,5,9-triazacyclododecan-1-yl]sulfonyl}-N,N-dimethylaniline 765 A205 N, X-dimethyl-4-{ [(38)-3-methyl-9-[(18)-1-phenylethyl]-5-(1,2,3,4- tetrahydroisoquinoline-2-sulfonyl)-1,5,9-triazacyclododecan1 -yl] sulfonyl } aniline 668

Method 36.

LiBH4 (0.7 g, 33 mmol) was added to a solution of N)-methyl-2-((tert-butoxycarbonylamino)methyl)butanoate (4.8 g, 20.8 mmol) in THF (50 mL). Stir the reaction mixture overnight at ambient temperature. The reaction was quenched with water (30 ml), then 1N was added dropwise. aqueous HCl, bringing the pH to 2-3. The resulting mixture was extracted with ethyl acetate and the extract was washed with saturated aqueous NaHCO ₃ , dried over anhydrous sodium sulfate and concentrated to give (I)-tert-butyl-2-(hydroxymethyl)butylcarbamate.

To a solution of N)-tert-butyl-2-(hydroxymethyl)butylcarbamate (3.4 g, 16.7 mmol) in toluene (40 ml) was added PPh ₃ (6.2 g, 17.4 mmol) and isoindoline-1 ,3-dione (3.7 g, 25 mmol). Cool the mixture to 0-5°C and add dropwise DIAD (4.3 g, 22 mmol). The reaction mixture was stirred at ambient temperature for 3 hours. The resulting precipitate was filtered off and the filtrate was concentrated. Eyes

- 69 045973 purify the residue by column chromatography (petroleum ether/EtOAc = 9:1 to 1:1) to give 1-(4-fluoro-2-methoxyphenyl)-4-(2-methyl-1H-imidazol-1- ylsulfonyl)piperazine.

Hydrazine hydrate was added to a solution of 1-(4-fluoro-2-methoxyphenyl)-4-(2-methyl-III-imidazol-1-ylsulfonyl)piperazine (3.8 g, 11.4 mmol) in EtOH (70 ml). (1.1 g, 80%). The reaction mixture was refluxed for 6 hours. The resulting product was filtered and the filtrate was concentrated under reduced pressure. Dissolve the residue in 1N. aqueous NaOH (20 ml), then extracted with DCM (50 ml). Dry the organic layer over anhydrous sodium sulfate and concentrate to give N)-tert-butyl-2-(aminomethyl)butylcarbamate.

TEA (1.01 g, 10.1 mmol) was added to a solution of N-tert-butyl-2-(aminomethyl)butylcarbamate (1.87 g, 9.2 mmol) in DCM (20 mL). Cool the mixture to 0~5°C and add 4-(dimethylamino)benzene-1sulfonyl chloride (2.0 g, 9.2 mmol). The reaction mixture was stirred at ambient temperature for 3 hours. The mixture was washed with water and then concentrated. Purify the residue by column chromatography (petroleum ether/EtOAc = 9:1 to 7:3) to give N)-tert-butyl-2-((4(dimethylamino)phenylsulfonamido)methyl)butylcarbamate.

To a solution of N)-tert-butyl-2-((4-(dimethylamino)phenylsulfonamido)methyl)-butylcarbamate (2.9 g, 7.5 mmol) in MeOH (10 ml) was added 4N. HCl/MeOH (15 ml). The mixture was stirred for 3 hours at ambient temperature. The solvent was removed under reduced pressure. The resulting product was suspended in DCM (100 ml) and saturated aqueous K ₂ CO ₃ (450 ml) was added. Stir the mixture for 5 minutes. The layer was separated in DCM, dried over anhydrous sodium sulfate and concentrated to give N)-NX2-(aminomethyl)butyl)-4-(dimethylamino)benzenesulfonamide.

To a solution of N)-N-(2-(aminomethyl)butyl)-4-(dimethylamino)benzenesulfonamide (0.9 g, 3.2 mmol) in CH3CN (9 ml) was added 1-((6-chloro-3 ,4-dihydroisoquinolin-2(III)yl)sulfonyl)-2,3-dimethyl-III-imidazol-3-ium (0.59 g, 3.2 mmol). Stir the reaction mixture overnight at ambient temperature. Concentrate the mixture and purify the residue by column chromatography (petroleum ether/EtOAc = 8:1 to 2:1) to give ^)-6-χλορ-Ν-(2-((4(dimethylamino)-phenylsulfonamido)methyl)butyl) -3,4-dihydroisoquinoline-2(III)-sulfonamide.

In a solution of N)-6-chloro-N2-((4-(dimethylamino)phenylsulfonamido)methyl)butyl)-3,4-dihydroisoquinoline-2(III)-sulfonamide (0.8 g, 1.5 mmol) in NMP (3 ml) NaH (159 mg, 3.9 mmol, 60%) was added. The mixture was stirred for 0.5 h at 80°C. Cool the mixture to 70°C and add cyclopropane-1,1diyl-bis(methylene) dimethanesulfonate (510 mg, 2.0 mmol). The reaction mixture was stirred for 2 hours at 70°C. The solvent was removed and the residue was purified by column chromatography (petroleum ether/EtOAc = 15:1) to give compound A182.

LC-MS: m/z 708.2 [M+H]+.

^1H NMR (400 MHz, CDClj: δ 7.59 (m, 2H), 7.15 (m, 2H), 7.05 (m, 1H), 6.68 (m, 2H), 4.34 ( m, 2H), 3.55 (m, 4H), 3.19 (m, 6H), 3.04 (s, 6H), 2.87 (m, 4H), 2.15 (m, 4H), 1.89 (m, 10H), 1.32 (m, 8H), 0.88 (m, 3H).

Method 37.

K benzyl-(S)-5-((6-chloro-3,4-dihydroisoquinolin-2(1H)-yl)sulfonyl)-9-((4-(dimethylamino)phenyl)sulfonyl)-7-methyl-1 ,5,9-triazacyclododecane-1-carboxylate (25 mg) was added with HBr (33% in acetic acid, 0.5 ml). After standing for 30 minutes at ambient temperature, the mixture was diluted with diethyl ether and water (1:1 mixture, 10 ml) and the organic layer was removed. The aqueous layer was washed with diethyl ether (1X5 ml), made alkaline with NaOH (4N) to pH ~12, then extracted with DCM (3X2 ml). The organic layers were dried over sodium sulfate, filtered and concentrated. The addition of hexanes (~2 ml) and subsequent concentration was used to accelerate the precipitation of the product. Drying in vacuum gave compound A183.

MS (ESI) for C27H40CN5O4S2, experiment 598 [M+H]+.

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Method 38.

Isobutyraldehyde (1.22 g, 11.3 mmol) was added to a solution of 3,3'-azandiyldipropan-1-ol (1.0 g, 7.5 mmol) in DCM (20 mL). Stir the mixture for 30 minutes at ambient temperature. Acetic acid (1.8 g, 30.1 mmol) was added and the mixture was stirred for an additional 30 minutes. Sodium triacetoxyborohydride (6.40 g, 30.1 mmol) was then added and the reaction mixture was allowed to stir overnight at ambient temperature. The pH of the mixture was adjusted to 1-2 with 3N. aqueous HCl and then stirred for 1 hour. 20% aqueous NaOH was added to the mixture (adjusting pH to 10-11), then extracted with DCM (50 ml). The extract was washed with saline, dried over anhydrous sodium sulfate and concentrated to give 3,3'-(isobutylazandiyl)-bis-(propan-1-ol).

NEt ₃ (820 mg, 8.2 mmol) was added to a solution of 3,3'-(isobutylazandiyl)-bis-(propan-1-ol) (780 mg, 4.1 mmol) in DCM (10 mL). MsCl (935 mg, 8.2 mmol) was then added slowly at 0°C. The reaction mixture was stirred for 3 hours at ambient temperature. The mixture was diluted with DCM (20 ml) and then washed with water (25 ml). Dry the organic layer over anhydrous sodium sulfate and concentrate to give 3,3'-(isobutylazandiyl)bis(propan-3,1-diyl)dimethanesulfonate.

In a solution of ^)-6-chloro-I-(3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-3,4-dihydroisoquinoline-2(III)-sulfonamide (0.515 g, 1.03 mmol) in dry N-methylpyrrolidone, NaH (0.25 g, 2.26 mmol) was added at ambient temperature. The mixture was heated at 80°C with stirring for 30 minutes and then cooled to ambient temperature. Add 3,3'-(isobutylazandiyl)-bis-(propane-3,1-diyl)dimethanesulfonate (0.5 g, 1.5 mmol) and heat the reaction mixture at 80°C for 2 hours. Quench the reaction with water, the mixture was then extracted with ethyl acetate. The extract was dried over anhydrous sodium sulfate and concentrated. The residue was purified by flash column chromatography to give compound A152.

MS (ESI) for C ₃₁ H ₄₈ ClN ₅ O ₄ S ₂ , experiment 654.4 [M+H]+.

^1H NMR (400 MHz, CDCl ₃ ): δ 7.59 (d, 2H), 7.16 (m, 2H), 7.04 (m, 1H), 6.69 (d, 2H), 4. 32 (m, 2H), 3.60 (m, 4H), 3.31 (m, 3H), 3.04 (s, 6H), 2.90 (m, 4H), 2.27 (m, 4H ), 1.93 (m, 2H), 1.79 (m, 1H), 0.87 (m, 9H).

The following compounds were synthesized similarly:

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No. IUPAC [M+N] ⁺ A123 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-(3-methylbutyl)-1,5,9triazacyclododecane-1 -yl]sulfonyl}-T\G, N-dimethyl aniline 668 A156 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-[(3-methylphenyl)methyl]-1.5, 9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethyl aniline 702 A131 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-[(4-methylphenyl)methyl]-1.5, 9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethyl aniline 702 A157 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclopentylmethyl)-3-methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl}-18G, N-dimethyl aniline 680 A106 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-(2-methylbutyl)-1,5,9triazacyclododecane-1 -yl]sulfonyl}-H N-dimethyl aniline 668 A108 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclobutylmethyl)-3-methyl-1,5,9triazacyclododecan-1-yl ]sulfonyl}-18G, N-dimethyl aniline 666 A134 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(2-cyclohexylethyl)-3-methyl-1,5,9triazacyclododecane-1 -yl]sulfonyl}-18G, N-dimethyl aniline 708 A122 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(2-cyclopropylethyl)-3-methyl-1,5,9triazacyclododecane-1 -yl]sulfonyl}-H N-dimethyl aniline 666 A120 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(2-cyclopentylethyl)-3-methyl-1,5,9triazacyclododecane-1 -yl]sulfonyl}-18G, N-dimethyl aniline 694 A165 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-(3,3,3-trifluoropropyl)-1, 5,9triazacyclododecan-1-yl]sulfonyl}-M, N-dimethyl aniline 694 A141 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinoline-2- yl)sulfonyl]-3-methyl-9-(propan-2-yl)-1,5,9-triazacyclododecane- 1 -yl ] sulphonyl} -N, N-dimethyl aniline 640 A176 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinoline-2- 678

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yl)sulfonyl]-9-[(cyclopent-1 -en-1 -yl)methyl]-3 -methyl-1,5,9triazacyclododecan-1-yl]sulfonyl}-X, N-dimethyl aniline A135 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-[(cyclohex-1-en-1-yl)methyl]-3- methyl-1,5,9triazacyclododecan-1-yl]sulfonyl}-T4, N-dimethyl aniline 692 A167 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-(2,2,2-trifluoroethyl)-1,5 ,9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethyl aniline 680 A169 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-[(3,3-difluorocyclobutyl)methyl]-3-methyl-1, 5,9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethyl aniline 702 A133 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-[(2-methylphenyl)methyl]-1.5, 9triazacyclododecan-1-yl]sulfonyl}-X, N-dimethyl aniline 702 A140 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(2-methoxyethyl)-3-methyl-1,5,9triazacyclododecane-1 -yl]sulfonyl}-18G, N-dimethyl aniline 656 A125 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-[(1,3-oxazol-2-yl)methyl ]-1,5,9triazacyclododecan-1-yl]sulfonyl}-X, N-dimethyl aniline 679 A124 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-9-[(1,2-oxazol-3-yl)methyl ]-1,5,9triazacyclododecan-1-yl]sulfonyl}-X, N-dimethyl aniline 679 A183 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-3-methyl-1,5,9-triazacyclododecan-1yl]sulfonyl} -N,N-dimethylaniline 598 A185 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinoline-2- yl)sulfonyl]-9-[(4,4-difluorocyclohexyl)methyl]-3-methyl-1,5,9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethylaniline 730 A107 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(2,2-dimethylpropyl)-3-methyl-1,5,9triazacyclododecane -1-yl]sulfonyl}4, N-dimethylaniline 668 A204 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinoline-2- yl)sulfonyl]-9-[(4-fluorocyclohexyl)methyl]-3-methyl-1,5,9triazacyclododecan-1-yl]sulfonyl}-18G, N-dimethylaniline 712

Method 39.

A mixture of 4-fluoro-2-methoxyaniline (25.0 g, 177 mmol), K ₂ CO ₃ (36.7 g, 266 mmol) and NaI (10.6 g, 0.07 mmol) in i-PrOH ( 250 ml) bis-(2-chloroethyl)amine hydrochloride (47 g, 177 mmol) was added. Boil

- 73 045973 reflux the reaction mixture overnight. The solvent was removed and the residue was purified by column chromatography (petroleum ether/EtOAc = 10:1) to give 1-(4-fluoro-2methoxyphenyl)piperazine.

MS: m/z 212.36 [M+H]+.

To a solution of 1-(4-fluoro-2-methoxyphenyl)piperazine (4.0 g, 19 mmol) in MeCN (40 ml) was added 2,3-dimethyl-1-(2-methyl-III-imidazol-1-ylsulfonyl )-III-imidazole-3 (7.5 g, 19 mmol). Stir the reaction mixture overnight at ambient temperature. The solvent was removed and the residue was purified by column chromatography (petroleum ether/EtOAc = 20:1) to give 1-(4-fluoro-2-methoxyphenyl)-4-(2-methyl-1H-imidazol-1-ylsulfonyl)piperazine.

MS: m/z 355.36 [M+H] ⁺ .

To a solution of 1-(4-fluoro-2-methoxyphenyl)-4-(2-methyl-III-imidazol-1-ylsulfonyl)-piperazine (6.0 g, 17 mmol) in DCM (60 ml) was added CF ₃ SO ₃ Me (2.78 g, 17 mmol). The reaction mixture was stirred for 2 hours at ambient temperature. The solvent was removed and the residue was washed with diethyl ether to obtain 1-(4-(4-fluoro-2-methoxyphenyl)piperazin-1ylsulfonyl)-2,3-dimethyl-111-imidazol-3-ium trifluoromethanesulfonate.

A solution of 1-(4-(4-fluoro-2-methoxyphenyl)piperazin-1-ylsulfonyl)-2,3dimethyl-III-imidazol-3-ium trifluoromethanesulfonate (740 mg, 2.3 mmol) in CH3CN (8 ml) tert-butyl 3-((3aminopropyl)(cyclohexylmethyl)amino)propylcarbamate (1.17 g, 2.3 mmol) was added. Stir the reaction mixture overnight at ambient temperature. Concentrate the mixture and purify the residue by column chromatography (petroleum ether/EtOAc = 8:1 to 2:1) to give tert-butyl-3-((cyclohexylmethyl)(3-(4-(4-fluoro-2-methoxyphenyl)piperazine- 1-sulfonamido)propyl)amino)propylcarbamate.

In a solution of tert-butyl-3-((cyclohexylmethyl)(3-(4-(4-fluoro-2-methoxyphenyl)-piperazine-1sulfonamido)propyl)amino)propylcarbamate (600 mg, 1 mmol) in DCM (30 ml) TFA (30 ml) was added. Stir the mixture for 2 hours at ambient temperature. The solvent was removed and the residue was dissolved in ethyl acetate, then washed with saturated aqueous NaHCO ₃ . Dry the organic phase over anhydrous sodium sulfate and concentrate to give N(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4-(4-fluoro-2-methoxyphenyl)piperazine-1-sulfonamide.

To a solution of N(3-((3-aminopropyl)(cyclohexylmethyl)amino)propyl)-4-(4-fluoro-2-methoxyphenyl)piperazine-1-sulfonamide (416 mg, 0.83 mmol) in DCM (5 ml ) Et ₃ N (170 mg, 1.66 mmol) and 4(dimethylamino)benzenesulfonyl chloride (365 mg, 1.66 mmol) were added. Stir the reaction mixture overnight at ambient temperature. Concentrate the mixture and purify by column chromatography (petroleum ether/EtOAc = 8:1 to 2:1) to give N(3-((cyclohexylmethyl)(3-(4(dimethylamino)phenylsulfonamido)propyl)amino)propyl)-4 -(4-fluoro-2-methoxyphenyl)piperazine-1sulfonamide.

In a solution of N(3-((cyclohexylmethyl)(3-(4-(dimethylamino)phenylsulfonamido)-propyl)amino)propyl)-4-(4-fluoro-2-methoxyphenyl)piperazine-1-sulfonamide (320 mg, 0 .45 mmol) to NMP (3 ml) was added NaH (53 mg, 1.32 mmol, 60% dispersion). Stir the mixture for 0.5 h at 80°C, then cool to 70°C and add cyclopropane-1,1-diyl-bis(methylene) dimethanesulfonate (170 mg, 0.66 mmol). The reaction mixture was stirred for 2 hours at 70°C. The solvent was removed and the residue was purified by column chromatography (petroleum ether/EtOAc = 15:1) to give compound A193.

MS (ESI) for C37H57FN6O5S2, experiment 745.0 [M+H] ⁺ .

^1H NMR (400 MHz, CDCl ₃ ): δ 7.57 (t, 2H), 6.64 (t, 1H), 6.63 (br.e, 4H), 3.86 (s, 3H), 3.19 (m, 4H), 3.04 (m, 8H), 2.87 (m, 12H), 2.36 (m, 2H), 1.56 (m, 15H), 0.43 (m , 4H).

Method 40.

CtaOSti TFUK

BocHN. NHj ---------► _6l)[Wi f №Oz —————-*·

- 74 045973

CbzOSu (5.83 g, 23.5 mmol). Aqueous NaOH (4N) was added to adjust the pH to 11. The reaction mixture was stirred at ambient temperature for 2 hours. The mixture was diluted with water (15 ml) and then extracted with ethyl acetate (50 ml). The organic phase was washed with 1N. aqueous HCl, dried over anhydrous sodium sulfate and concentrated. The residue was purified by column chromatography to obtain benzyl-tert-butyl-(2-methylpropan-1,3-diyl)^)-dicarbamate.

To a solution of benzyl tert-butyl-(2-methylpropan-1,3-diyl)-^)-dicarbamate (4.5 g, 14.0 mmol) in DCM (25 ml) was added TFA (20 ml). Stir the reaction mixture at ambient temperature for 3 hours. Concentrate the mixture and dissolve the residue in DCM (50 ml). Wash the resulting solution with saturated aqueous K ₂ CO ₃ and dry the layer in DCM over anhydrous sodium sulfate and concentrate to give N)-benzyl-3-amino-2-methylpropylcarbamate.

To a solution of N)-benzyl-3-amino-2-methylpropylcarbamate (2.7 g, 12.2 mmol) in CH ₃ CN (25 ml) was added 1-(6-chloro-3,4-dihydroisoquinoline-2(III )-ylsulfonyl)-2,3-dimethyl-III-imidazol-3-ium (5.7 g, 12.2 mmol). Stir the reaction mixture at ambient temperature overnight. Concentrate the mixture and purify the residue by column chromatography to give N)-benzyl3-(6-chloro-1,2,3,4-tetrahydroisoquinolino-2-sulfonamido)-2-methylpropylcarbamate.

Dissolve N)-benzyl-3-(6-chloro-1,2,3,4-tetrahydroisoquinoline-2-sulfonamido)-2-methylpropylcarbamate (2.7 g, 6.0 mmol) in HBr (48%)/HOAc (20 ml). Stir the mixture at ambient temperature for 1 hour. Dilute the mixture with water (100 ml) and then wash with diethyl ether (50 ml). The aqueous layer was made alkaline with NaOH to pH 10. The resulting mixture was extracted with DCM (100 ml). Dry the layer in DCM over anhydrous sodium sulfate and concentrate to give N)-N-(3-amino2-methylpropyl)-6-chloro-3,4-dihydroisoquinoline-2(III)-sulfonamide.

In a solution of N)-N3-amino-2-methylpropyl)-6-chloro-3,4-dihydroisoquinoline-2(III)-sulfonamide (0.6 g, 1.9 mmol) and TEA (270 mg, 2.7 mmol) in DCM (6 ml) was added benzo[isoxazole-5sulfonyl chloride (420 mg, 1.9 mmol) at 0°C. The reaction mixture was stirred at ambient temperature for 2 hours. The mixture was washed with water and then concentrated. The residue was purified by column chromatography to obtain ^)-M-(3-(6-chloro-1,2,3,4-tetrahydroisoquinoline-2sulfonamido)-2-methylpropyl)benzo^]isoxazole-5-sulfonamide.

In a solution of ^)-X(3-(6-chloro-1,2,3,4-tetrahydroisoquinoline-2-sulfonamido)-2-methylpropyl)benzo^]isoxazole-5-sulfonamide (0.61 g, 1.22 mmol) in NMP (6 ml) was added NaH (122 mg, 3.05 mmol, 60%). The mixture was stirred for 0.5 h at 80°C. Cool the mixture to 70°C and add 3,3'-(cyclohexylmethylazandiyl)bis(propan-3,1-diyl)dimethanesulfonate (0.74 g, 1.83 mmol). The reaction mixture was stirred for 2 hours at 70°C. Cool the mixture and pour it into water. The resulting mixture was extracted with ethyl acetate. Concentrate the organic phase and purify the residue by column chromatography (petroleum ether/EtOAc = 12:1) to give compound A180.

LC-MS (ESI): m/z 692.2 [M+H]+.

1H NMR (400 MHz, CDCl ₃ ): δ 7.84 (s, 1H), 7.61 (m, 1H), 7.13 (m, 3H), 6.69 (m, 2H), 4.32 (m, 2H), 3.66 (m, 5H), 3.21 (m, 5H), 2.91 (m, 4H), 2.77 (m, 4H), 1.91 (m, 4H) , 1.57 (m, 6H), 1.13 (m, 6H), 0.89 (m, 3H).

The following compounds were synthesized similarly:______________________________

No. IUPAC 3 - {[(3 8)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinoline-2- [M+N] ⁺ A168 yl)sulfonyl]-9-(cyclohexylmethyl)-3-methyl-1,5,9triazacyclododecan-1-yl]sulfonyl}-T\G, N-dimethylaniline 6-chloro-2-{[(3 8)-9- (cyclohexylmethyl)-3-methyl-5- 694 A171 phenylmethanesulfonyl-1,5,9-triazacyclododecane-1yl]sulfonyl)-1,2,3,4-tetrahydroisoquinoline 4-{[(38)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinoline-2- 665 A179 yl)sulfonyl]-9-(cyclohexylmethyl)-3-methyl-1,5,9triazacyclododecan-1-yl]sulfonyl)-X, N-dimethylbenzamide (38)-X-benzyl-9-(cyclohexylmethyl)-5-[4- 722 A184 (dimethylamino)benzenesulfonyl]-X,3-dimethyl-1,5,9- triazacyclododecane-1-sulfonamide 6-chloro-2- {[(3 8)-9-(cyclohexylmethyl)-3 -methyl-5 -[(1 -methyl-1H- 648 A188 indol-5-yl)sulfonyl]-1,5,9-triazacycle od odekai-1 yl]sulfonyl)-1,2,3,4-tetrahydroisoquinoline 704 A189 (38)-9-(cyclohexylmethyl)-5-[4-(dimethylamino)benzenesulfonyl]N-[(4-ethylphenyl)methyl]-3-methyl-1,5,9-triazacyclic odecane-1 sulfonamide 662 N- { 4- [( {[(3 8)-9-(cyclohexylmethyl)-5 - [4- A192 (dimethylamino)benzenesulfonyl]-3-methyl-1,5,9- triazacyclododecan-1-yl]sulfonyl}amino)methyl]phenyl}-X- 705 methylacetamide

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Method 41.

about CF3SO3

To a suspension of Mg (4.4 g, 181 mmol) in THF (200 ml) was added 1-bromo-2-methoxybenzene (40 g, 214 mmol). Stir the mixture at 80°C for 0.5 hour. Cool the mixture to 0°C and add a solution of tert-butyl-3-oxopiperidine-1-carboxylate (35.5 g, 181 mmol) in THF. Stir the reaction mixture at ambient temperature for 2 hours. Purify the residue by column chromatography to give tert-butyl 3-hydroxy-3-(2-methoxyphenyl)piperidine-1-carboxylate.

^1H NMR (400 MHz, CDCl3): δ 7.48 (m, 1H), 7.32 (m, 1H), 6.98 (m, 2H), 4.08 (m, 1H), 3.96 (s, 3H), 3.82 (m, 1H), 3.31 (s, 1H), 2.98 (m, 1H), 2.25 (m, 1H), 1.99 (m, 2H) , 1.62 (m, 2H), 1.63 (m, 9H).

Cool a solution of tert-butyl-3-hydroxy-3-(2-methoxyphenyl)piperidine-1-carboxylate (23.0 g, 75 mmol) and triethylsilane (44 ml) in DCM (230 ml) to -30°C and add TFA (27 ml). The reaction mixture was stirred at -30°C for 2.5 hours and then allowed to warm to ambient temperature and stirred for another 3.5 hours. The mixture was poured into ice water and the pH of the solution was adjusted to 9 with saturated aqueous sodium hydroxide. The resulting mixture was extracted with DCM three times. The combined organic phases were dried over anhydrous sodium sulfate and concentrated. The residue was purified by column chromatography to give 3-(2-methoxyphenyl)piperidine.

A solution of 3-(2-methoxyphenyl)piperidine (6.5 g, 34 mmol) and D-tartaric acid (5.1 g, 34 mmol) in MeOH (25 ml) was refluxed for 2 hours. The mixture was cooled to ambient temperature and filtered to obtain the tartrate salt. The resulting tartrate salt was recrystallized twice from MeOH and then treated with aqueous NaOH. The resulting mixture was extracted with DCM. Dry the layer in DCM over anhydrous sodium sulfate and concentrate to give N)-3-(2-methoxyphenyl)piperidine.

A solution of N)-3-(2-methoxyphenyl)piperidine (500 mg, 2.6 mmol) and 2,3-dimethyl-1-((2-methyl-III-imidazol-1-yl)sulfonyl)-III trifluoromethanesulfonate was stirred -imidazol-3-ium (1.1 g, 2.7 mmol) in MeCN (15 ml) overnight at ambient temperature. The solvent was removed. Purify the residue by column chromatography (petroleum ether/EtOAc = 20:1) to give (S)-3-(2methoxyphenyl)-1-((2-methyl-III-imidazol-1-yl)sulfonyl)piperidine.

LC-MS (ESI): m/z 336.06 [M+H]+.

CF was added to a solution of N)-3-(2-methoxyphenyl)-1-((2-methyl-N-imidazol-1-yl)sulfonyl)-piperidine (800 g, 2.4 mmol) in DCM (10 ml). ₃ SO ₃ Me (398 mg, 2.4 mmol). The reaction mixture was stirred for 2 hours at ambient temperature. The solvent was removed and the residue was washed with diethyl ether to obtain Ι-1-(3-(2-methoxyphenyl)piperidin-1-yl)-2,3dimethyl-11-im idazol-3-ium trifluoromethanesulfonate.

To a solution of ^)-1-(3-(2-methoxyphenyl)piperidin-1-yl)-2,3-dimethyl-shimidazol-3-ium trifluoromethanesulfonate (1 g, 2 mmol) in CH3CN (24 ml) was added ^) -N-(3-amino-2-methylpropyl)-4(dimethylamino)benzenesulfonamide (0.55 g, 2 mmol). Stir the reaction mixture overnight at ambient temperature. The mixture was concentrated. Purify the residue by column chromatography (petroleum ether/EtOAc = 8:1 to 2:1) to give (S)-N-((S)-3-((4(dimethylamino)phenyl)sulfonamido)-2-methylpropyl) -3-(2-methoxyphenyl)piperidine-1-sulfonamide.

In a solution of ^)-M-(^)-3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-3-(2methoxyphenyl)piperidine-1-sulfonamide (454 mg, 0.86 mmol) in NMP (14 ml) added NaH (104 mg,

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2.6 mmol, 60%). The mixture was stirred for 0.5 h at 80°C. Cool the mixture to 70°C and add ((cyclohexylmethyl)azandiyl)-bis-(propane-3,1-diyl)dimethanesulfonate (500 mg, 1.3 mmol). The reaction mixture was stirred for 2 hours at 70°C. The mixture was cooled to 0-5°C and poured into water. The resulting mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated. The residue was purified by preparative HPLC to obtain compound A210.

LC-MS (ESI): m/z 718.17 [M+H]+.

^1H NMR (400 MHz, CDCl3): δ 7.60 (m, 2H), 7.23 (m, 2H), 6.95 (m, 2H), 6.68 (m, 2H), 3.82 (s, 3H), 3.73 (m, 5H), 3.26 (m, 4H), 3.12 (s, 6H), 2.76 (m, 5H), 2.36 (m, 4H) , 1.76 (m, 9H), 1.33 (m, 10H), 0.86 (m, 6H).

The following compound was synthesized similarly:___________________________

No. IUPAC [M+N] ⁺ A208 4-{ [(3 8)-9-(cyclohexylmethyl)-5-{ [(3 8)-3-(4methoxyphenyl)piperidin-1-yl] sulfonyl } -3 -methyl-1,5,9triazacyclododecane-1- yl]sulfonyl}-K, N-dimethylaniline 718

Method 42.

In a solution of tert-butyl-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-5,6-dihydropyridine-1(2H)carboxylate (10.92 g, 35 mmol), 1-bromo-3-methoxybenzene (6.6 g, 35 mmol) and Cs ₂ CO ₃ (3.44 g, 106 mmol) in 1,4-dioxane (50 ml) and H ₂ O (10 ml ) Pd(dppf)Cl ₂ (1.32 g, 1.75 mmol) was added. The reaction mixture was stirred overnight at 80°C. The solvent was removed. The residue was purified by column chromatography (petroleum ether/EtOAc = 10:1) to give tert-butyl 3-(3-methoxyphenyl)-5,6dihydropyridine-1(2H-carboxylate.

LC-MS (ESI): m/z 290.12 [M+H]+.

^1H NMR (400 MHz, CDCl ₃ ): δ 7.26 (m, 1H), 6.96 (m, 1H), 6.89 (s, 1H), 6.81 (m, 1H), 6. 19 (s, 1H), 4.25 (s, 2H), 3.81 (s, 3H), 3.54 (m, 2H), 2.30 (s, 2H), 1.49 (s, 9H ).

Pd/ C (2 g). The mixture was stirred at ambient temperature for 4 hours under an H ₂ atmosphere. Filter the mixture and concentrate the filtrate to obtain tert-butyl 3-(3-methoxyphenyl)piperidine-1-carboxylate.

^1H NMR (400 MHz, CDCl ₃ ): δ 7.25 (m, 1H), 6.81 (m, 3H), 3.99 (m, 2H), 3.73 (s, 3H), 2. 65 (m, 1H), 2.55 (m, 1H), 2.50 (m, 2H), 1.98 (m, 1H), 1.65 (m, 2H), 1.44 (s, 9H ).

Stir a solution of tert-butyl 3-(3-methoxyphenyl)piperidine-1-carboxylate (7.85 g, 27 mmol) in HCl/MeOH (80 ml) at ambient temperature for 4 hours. Treat the mixture with water and then brought to pH 12 3N. aqueous NaOH. The resulting mixture was extracted with EtOAc (2x100 ml). Dry the combined extracts over anhydrous sodium sulfate and concentrate to give 3-(3-methoxyphenyl)piperidine.

LC-MS (ESI): m/z 192.09 [M+H]+.

To a solution of P-(-)-tartaric acid (2.5 g, 16.7 mmol) in MeOH (12.5 ml) was added 3-(3-methoxyphenyl)piperidine (3.2 g, 16.7 mmol). The mixture was stirred at 80°C for 24 hours, then cooled to ambient temperature. The resulting precipitate was collected by filtration and then recrystallized three times from MeOH to give N)-3-(3-methoxyphenyl)piperidine.

To a solution of N)-3-(3-methoxyphenyl)piperidine (1.28 g, 6.7 mmol) in CH3CN (10 ml) was added 1-(III-imidazol-1-ylsulfonyl)-3-methyl-III-trifluoromethanesulfonate imidazol-3-ium (2.87 g, 7.3 mmol). Stir the reaction mixture at ambient temperature overnight. Removed the solution

- 77 045973 tel. Purify the residue by column chromatography (petroleum ether/EtOAc = 9:1 to 8:2) to give N)-3-(3-methoxyphenyl)-1-(2-methyl-N-imidazol-1-ylsulfonyl)piperidine.

^1H NMR (400 MHz, CDCl3): δ 7.27 (m, 1H), 7.21 (s, 1H), 6.93 (s, 1H), 6.79 (m, 2H), 6.72 (s, 1H), 3.93 (m, 2H), 3.82 (s, 3H), 2.68 (m, 1H), 2.65 (m, 2H), 2.62 (s, 3H) , 2.06 (m, 1H), 1.95 (m, 1H), 1.82 (m, 1H), 1.62 (m, 1H).

To a solution of N)-3-(3-methoxyphenyl)-1-(2-methyl-N-imidazol-1-ylsulfonyl)-piperidine (1.6 mg, 4.7 mmol) in DCM (20 ml) was added CF ₃ CO ₂ Me (782 mg, 4.7 mmol). The reaction mixture was stirred at ambient temperature for 2 hours. The solvent was removed. The resulting product was washed with diethyl ether to obtain N)-1-(3-(3-methoxyphenyl)piperidin1-ylsulfonyl)-2,3-dimethyl-III-imidazol-3-ium trifluoromethanesulfonate.

In a solution of trifluoromethanesulfonate ^)-1-(3-(3-methoxyphenyl)piperidin-1-ylsulfonyl)-2,3dimethyl-III-imidazol-3-ium (1.1 g, 2.2 mmol) in CH3CN (11 ml ) (I)-K-(3-amino-2methylpropyl)-4-(dimethylamino)benzenesulfonamide (896 mg, 33 mmol) was added. Stir the mixture at ambient temperature overnight. The solvent was removed. The residue was purified by column chromatography to obtain N)-M-(N)-3-(4-(dimethylamino)phenylsulfonamido)-2-methylpropyl)-3-(3methoxyphenyl)piperidine-1-sulfonamide.

^1H NMR (400 MHz, CDCl3): δ 7.68 (m, 2H), 7.28 (m, 1H), 6.85 (m, 1H), 6.80 (m, 2H), 6.68 (m, 2H), 4.81 (m, 2H), 3.82 (s, 3H), 3.76 (m, 2H), 3.12 (m, 1H), 3.07 (s, 6H) , 2.82 (m, 4H), 2.11 (m, 1H), 1.93 (m, 2H), 1.77 (m, 1H), 1.29 (m, 3H), 0.91 ( m, 3H).

A solution of ^)-M-(^)-3-(4-(dimethylamino)phenylsulfonamido)-2-methylpropyl)-3-(3-methoxyphenyl)piperidine-1-sulfonamide (550 mg, 1.05 mmol) in NMP (6 ml) sodium hydride (75.6 mg, 3.15 mmol) was added. The mixture was stirred for 30 minutes at 80°C. Cool the mixture to 70°C and add a solution of ((cyclohexylmethyl)azandiyl)-bis-(propane-3,1-diyl)dimethanesulfonate (606 mg, 1.57 mmol) in NMP (3 ml). Stir the reaction mixture at 70°C for 1 hour. Cool the mixture to ambient temperature and pour into water (10 ml). The resulting mixture was extracted with ethyl acetate (20 ml). Wash the organic layer with water (2x15 ml), dry over anhydrous sodium sulfate and concentrate. Purify the residue by column chromatography (EtOAc/petroleum ether = 1:15 to 1:5) to give compound A209.

LC-MS (ESI): m/z 719.2 [M+H]+.

^1H NMR (400 MHz, CDCl3): δ 7.61 (m, 2H), 7.28 (m, 2H), 6.84 (m, 3H), 6.70 (m, 2H), 3.83 (s, 3H), 3.70 (m, 2H), 3.55 (m, 2H), 3.30 (m, 1H), 3.11 (s, 9H), 2.79 (m, 5H) , 2.41 (m, 1H), 2.33 (m, 3H), 2.05 (m, 8H), 1.67 (m, 4H), 1.27 (m, 7H), 0.94 ( m, 3H), 0.85 (m, 2H).

Method 43.

In a solution of ^)-N3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-4-(4-fluoro-2methoxyphenyl)piperazine-1-sulfonamide (900 mg, 1.65 mmol) in NMP ( 5 ml) NaH (198 mg, 4.92 mmol, 60%) was added. The mixture was stirred for 0.5 h at 80°C. Cool the mixture to 70°C and add ^)-3,3'-(1-cyclohexylethylazanediyl)-bis(propane-3,1-diyl)dimethanesulfonate (844 mg, 2.49 mmol). The reaction mixture was stirred for 2 hours at 70°C, then cooled to 0-5°C and poured into water. The resulting mixture was extracted with ethyl acetate. The extract was concentrated. Purify the residue by column chromatography (petroleum ether/EtOAc = 15:1) to give compound A203.

LC-MS (ESI): m/z 751.33 [M+H]+.

1H NMR (400 MHz, CDCl3): δ 7.68 (m, 2H), 6.92 (m, 1H), 6.78 (m, 4H), 3.92 (s, 4H), 3.86 ( m, 1H), 3.62 (m, 1H), 3.40 (m, 2H), 3.19 (m, 3H), 3.04 (s, 9H), 2.87 (m, 2H), 2.26 (m, 3H), 1.92 (m, 3H), 1.79 (m, 4H), 1.66 (m, 2H), 1.54 (m, 2H), 1.38 (m , 4H), 1.26 (m, 5H), 0.98 (m, 3H), 0.83 (m, 4H).

The following compounds were synthesized similarly:

No. IUPAC [M+N] ⁺ B22 4-{[(1S, 12R)-10-[(6-chloro-1,2,3,4-τeτpahydroisoquinolin-2yl)sulfonyl]-6-(cyclohexylmethyl)-2,6,10- triazabicyclo[10.2.0]tetrad ekan-2-yl]sulfonyl} -N, N- dimethylaniline 706 A160 4-{[(3S,7S,l 18)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-3,7,11 -trimethyl- 1,5,9triazacyclododecan-1-yl]sulfonyl}-X, N-dimethyl aniline 722 B16 4-{[(3S,7R,l 18)-5-[(6-chloro-1,2,3,4-tetrahydroisoquinolin-2yl)sulfonyl]-9-(cyclohexylmethyl)-3,7,11 -trimethyl- 1,5,9triazacyclododecan-1-yl]sulfonyl}-X, N-dimethyl aniline 722

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Method 44.

mpem-butyl-(3R, 5R)-3,5-dimethylpiperazine-1-carboxylate (950 mg, 4.4 mmol). Stir the reaction mixture at 100°C for 3 hours. Remove the solvent and purify the residue by column chromatography (petroleum ether/EtOAc = 20:1) to give tert-butyl-(3R,5R)-4-(3-fluoro-5-methoxyphenyl) )-3,5-dimethylpiperazine-1-carboxylate.

LC-MS (ESI): m/z 339.0 [M+H]+.

TFA was added to a solution of tert-butyl-(3R,5R)-4-(3-fluoro-5-methoxyphenyl)-3,5-dimethylpiperazine-1-carboxylate (800 mg, 2.4 mmol) in DCM (4 mL). (4 ml). The mixture was stirred at ambient temperature for 1 hour. The solvent was removed. Dissolve the residue in DCM (50 ml) and then wash with saturated aqueous K ₂ CO ₃ . Dry the organic layer over anhydrous sodium sulfate and concentrate to give (2R,6R)-1-(3-methoxy-5-methylphenyl)-2,6-dimethylpiperazine.

To a solution of (2R,6R)-1-(3-methoxy-5-methylphenyl)-2,6-dimethylpiperazine (500 mg, 2.1 mmol) in MeCN (5 ml) was added 2,3-dimethyl-1-trifluoromethanesulfonate ((2-methyl-1H-imidazol-1-yl)sulfonyl)-1Nimidazol-3-ium (825 mg, 2.1 mmol). Stir the reaction mixture overnight at ambient temperature. The solvent was removed. Purify the residue by column chromatography (petroleum ether/EtOAc = 20:1) to give (2R,6R)-1-(3-fluoro-5-methoxyphenyl)-2,6-dimethyl-4-((2-methyl1H- imidazol-1-yl)sulfonyl)piperazine.

LC-MS (ESI): m/z 383.53 [M+H]+.

CF ₃ SO ₃ Me (193 mg, 1.2 mmol). The reaction mixture was stirred for 2 hours at ambient temperature. The solvent was removed and the residue was washed with diethyl ether to give 1-(((3R,5R)-4-(3-fluoro-5-methoxyphenyl)-3,5-dimethylpiperazin-1yl)sulfonyl)-2,3-dimethyl-1 trifluoromethanesulfonate H-imidazol-3-ium.

Into a solution of l-(((3R,5R)-4-(3-fluoro-5-methoxyphenyl)-3,5-dimethylpiperazin1-yl)sulfonyl)-2,3-dimethyl-III-imidazolium-3-trifluoromethanesulfonate ( 500 mg, 0.91 mmol) in CH3CN (5 ml) was added N)-A(3-amino-2-methylpropyl)-4-(dimethylamino)-benzenesulfonamide (248 mg, 0.91 mmol). Stir the reaction mixture overnight at ambient temperature. The mixture was concentrated. Purify the residue by column chromatography (petroleum ether/EtOAc = 8:1 to 2:1) to give ^^)-A(^)-3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)- 4-(3-fluoro-5-methoxyphenyl)-2,6-dimethylpiperazine-1-sulfonamide.

LC-MS (ESI): m/z 572.4 [M+H]+.

Into a solution of ^^)-A(^)-3-((4-(dimethylamino)phenyl)sulfonamido)-2-methylpropyl)-4-(3-fluoro-5methoxyphenyl)-2,6-dimethylpiperazine-1-sulfonamide ( 430 mg, 0.75 mmol) to NMP (4 ml) was added NaH (91 mg, 2.3 mmol, 60%). The mixture was stirred for 0.5 h at 80°C. Cool the mixture to 70°C and add ((cyclohexylmethyl)azandiyl)bis(propane-3,1-diyl)dimethanesulfonate (435 mg, 1.1 mmol). The reaction mixture was stirred for 2 hours at 70°C. The solvent was removed and the residue was purified by column chromatography (petroleum ether/EtOAc = 15:1) to give compound A200.

LC-MS (ESI): m/z 764.3 [M+H]+.

^1H NMR (400 MHz, CDCl ₃ ): δ 7.60 (d, 2H), 6.69 (d, 2H), 6.32 (d, 3H), 3.78 (s, 3H), 3. 06 (q, 2H), 3.04 (s, 8H), 3.00 (s, 2H), 2.87 (d, 2H), 2.76 (s, 2H), 2.34 (br.B , 4H), 2.09 (bb, 2H), 1.33 (s, 17H), 1.04 (d, 7H), 0.93 (d, 3H).

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^)-3,3'-(1-Cyclohexylethylazandiyl)dipropan-1-ol (intermediate for compounds A203, A190, A207)

Methyl acrylate (8.1 g, 94.38 mmol) was added to a solution of N)-1-cyclohexylethanamine (4.0 g, 31.46 mmol) in methanol (50 mL). The reaction mixture was stirred for 72 hours at 45°C. Concentrate the mixture and purify the residue by column chromatography (EtOAc/petroleum ether = 30:1) to give N)-dimethyl-3,3'-(1-cyclohexylethylazanediyl)dipropanoate.

LiBH4 (1.45 g, 66.84 mmol) was added to a solution of N)-dimethyl-3,3'-(1-cyclohexylethylazanediyl)dipropanoate (4.0 g, 13.37 mmol) in THF (50 ml). The reaction mixture was stirred at 60°C for 2 hours. The mixture was cooled to 0-5°C and the reaction was quenched with water (30 ml). The pH of the mixture was adjusted to 3 with 1N. aqueous hydrochloric acid and stirred for 30 minutes. The resulting mixture was washed with dichloromethane. Adjust the pH of the aqueous layer to 9-10 with saturated aqueous sodium carbonate and then extract with DCM (3x). Dry the combined organic layers over anhydrous sodium sulfate and concentrate to give N)-3,3'-(1-cyclohexylethylazandiyl)dipropan-1-ol.

^1H NMR (400 MHz, CDCl ₃ ): δ 3.66 (s, 1H), 2.82 (m, 2H), 2.42 (m, 2H), 2.21 (m, 1H), 1. 96 (m, 2H), 1.67 (m, 4H), 1.22 (m, 4H), 0.91 (m, 3H), 0.81 (m, 1H), 0.72 (m, 1H ).

The following compounds were synthesized similarly: d-3,3'-(1-cyclohexylethylazandiyl)dipropan1-ol (intermediate for compound A191) and d)-3,3'-((1-phenylethyl)azandiyl)-bis-(propane1- ol) (intermediate for connection A205).

6-Chloro-K-(((^^)-2-(4-(dimethylamino)phenylsulfonamido)cyclobutyl)methyl)-3,4-dihydroisoquinoline-2(III)-sulfonamide (intermediate for compound B22)

Benzyl alcohol was added dropwise to a suspension of cis-3-oxabicyclo[3.2.0]heptane-2,4-dione (32.8 g, 200 mmol) and quinine (71.4 g, 220 mmol) in toluene (1 L). (64.9 g, 600 mmol) for 0.5 h at -55°C. The reaction mixture was stirred at -55°C for 96 hours. The resulting clear solution was concentrated to dryness and the residue was dissolved in diethyl ether (1.2 L). The solution was washed with 2N. aqueous HCl (1 L), extracted the organic layer with saturated aqueous sodium bicarbonate (500x5 ml) and washed the combined aqueous phases with diethyl ether (500 ml) to remove traces of benzyl alcohol. The aqueous phase was acidified with 8N. aqueous HCl, then extracted with DCM (2x 1.0 L). Dry the combined organic layers over anhydrous sodium sulfate and concentrate to give (1S,2R)-2(benzyloxycarbonyl)cyclobutanecarboxylic acid.

DPPA (51, 8 g, 188.5 mmol) at 0°C. The mixture was stirred for 2 hours at ambient temperature, then washed with water and dried over anhydrous NaSO ₄ . BnOH (18.5 g, 171 mmol) and triethylamine (34.6 g, 342 mmol) were added to the organic solution. Reflux the reaction mixture overnight and then dilute with DCM (500 ml). The mixture was washed with 1N. aqueous HCl (1 l) and saturated aqueous NaHCO ₃ . The organic phase was dried over anhydrous sodium sulfate, concentrated and purified by column chromatography to give (^^)-benzyl-2-(benzyloxycarbonylamino)cyclobutanecarboxylate.

- 80 045973

To a solution of (1K^)-benzyl-2-(benzyloxycarbonylamino)cyclobutanecarboxylate (20 g, 59 mmol) in THF (100 ml) was added 4N. aqueous NaOH (50 ml) at 0°C. Stir the reaction mixture overnight at ambient temperature. The mixture was diluted with water (30 ml), then washed with diethyl ether (2x50 ml). The aqueous layer was acidified with 3N. aqueous HCl and extracted with ethyl acetate. The extracts were dried over anhydrous sodium sulfate and concentrated to give (1R,2S)-2(benzyloxycarbonylamino)cyclobutanecarboxylic acid.

1H NMR (400 MHz, CDC13): δ 7.34 (m, 5H), 5.12 (m, 2H), 4.62 (m, 1H), 3.58 (m, 1H), 2.28 ( m, 2H), 1.89 (m, 2H).

_{MeI (} 2, 96 g, 20.8 mmol). Stir the reaction mixture at ambient temperature overnight. The mixture was poured into water (150 ml), then extracted with diethyl ether (200 ml). The organic phase was dried over anhydrous sodium sulfate and concentrated. The residue was purified by column chromatography to obtain (III)-methyl-2-(benzyloxycarbonylamino)cyclobutanecarboxylate.

Ή NMR (400 MHz, CDCl ₃ ): δ 7.34 (m, 5H), 5.69 (m, 1H), 5.08 (s, 2H), 4.62 (m, 1H), 3.66 (s, 3H), 3.38 (m, 1H), 2.32 (m, 2H), 1.99 (m, 2H).

NaOMe (9.1 g, 169 mmol) was added to a solution of (III)-methyl-2-(benzyloxycarbonylamino)cyclobutanecarboxylate (8.9 g, 33.8 mmol) in MeOH (250 ml). Heat the mixture at 45°C for 4 hours, then cool to 0°C and add dropwise HOAc (10.2 g, 169 mmol). The mixture was then poured into water (500 ml) and extracted with ethyl acetate (300 ml). The extract was washed with saturated aqueous NaHCO ₃ and dried over anhydrous sodium sulfate. The organic solution was concentrated and the residue was purified by column chromatography to give (N^)-methyl-2-(benzyloxycarbonylamino)cyclobutanecarboxylate.

Ή NMR (400 MHz, CDC13): δ 7.34 (m, 5H), 5.08 (m, 3H), 4.38 (m, 1H), 3.72 (s, 3H), 3.38 ( m, 1H), 2.32 (m, 2H), 1.89 (m, 2H).

LiBH ₄ (0.38 g, 9.8 mmol) was added to a solution of (N^)-methyl-2-(benzyloxycarbonylamino)cyclobutanecarboxylate (1.3 g, 4.9 mmol) in THF (10 ml). The reaction mixture was stirred for 3 hours at ambient temperature, then poured into water (15 ml) and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated to give benzyl-(NH)-2(hydroxymethyl)cyclobutylcarbamate.

Ή NMR (400 MHz, CDCl ₃ ): δ 7.34 (m, 5H), 5.08 (m, 3H), 3.72 (m, 1H), 3.57 (m, 2H), 2.32 (m, 2H), 1.89 (m, 2H), 1.45 (m, 1H).

In a solution of benzyl-(Ni^)-2-(hydroxymethyl)cyclobutylcarbamate (1.1 g, 4.94 mmol), PPh ₃ (1.5 g, 5.9 mmol) and isoindoline-1,3-dione (1 .0 g, 6.9 mmol) in toluene (10 ml) was added DIAD (1.2 g, 5.9 mmol) at 0°C. The reaction mixture was stirred for 3 hours. The precipitate was filtered off and the filtrate was concentrated and purified by column chromatography to give benzyl (1S,2R)-2-((1,3-dioxoisoindolin-2-yl)methyl)cyclobutylcarbamate.

Hydrazine hydrate (0. 64 g). The reaction mixture was refluxed for 3 hours. The mixture was cooled, filtered, and the filtrate was concentrated. Dissolve the residue in 1N. aqueous HCl (30 ml) and then washed with diethyl ether (20 ml). The pH of the aqueous layer was adjusted to 10. The resulting mixture was extracted with DCM (2x50 ml). Dry the combined organic layers over anhydrous sodium sulfate and concentrate to give benzyl (1S,2R)-2(aminomethyl)cyclobutylcarbamate.

Trifluoromethanesulfonate 1 -((6-chloro-3,4-dihydroisoquinoline-2(1H) )-yl)sulfonyl)-2,3dimethyl-1H-imidαzol-3-ium (2.4 g, 5.12 mmol). Stir the reaction mixture at ambient temperature overnight. Concentrate the mixture and purify the residue by column chromatography to give benzyl (1S,2R)-2-((6-chloro-1,2,3,4-tetrahydroisoquinoline-2-sulfonamido)methyl)cyclobutylcarbamate.

To a 40% solution of HBr in HOAc (10 ml) was added benzyl-(i.2K)-2-((6-chloro-1.2.3.4tetrahydroisoquinoline-2-sulfonamido)methyl)cyclobutylcarbamate (1.2 g, 2.6 mmol ). The mixture was stirred for 3 hours, then poured into water (50 ml) and washed with diethyl ether. The pH of the aqueous layer was adjusted to 10 8N. aqueous NaOH. The mixture was extracted with DCM (50 ml). The organic phase was dried over anhydrous sodium sulfate and concentrated to give Na(((III)-2-aminocyclobutyl)methyl)6-chloro-3,4-dihydroisoquinoline-2(1H)-sulfonamide.

Into a solution of Na(((III^)-2-aminocyclobutyl)methyl)-6-chloro-3,4-dihydroisoquinoline-2(1H)sulfonamide (0.66 g, 2.0 mmol) and NEt ₃ (350 mg, 3.5 mmol) in DCM (5 ml) was added 4-(dimethylamino)benzenesulfonyl chloride (426 mg, 2.0 mmol) at 0°C. The reaction mixture was stirred at ambient temperature for 2 hours, then concentrated. Dissolve the residue in DCM and wash with water. The organic phase was dried over anhydrous sodium sulfate and concentrated

- 81 045973 was performed to obtain 6-chloro-N-(((^^)-2-(4-(dimethylamino)phenylsulfonamido)cyclobutyl)methyl)-3,4dihydroisoquinoline-2(1H)-sulfonamide.

1H NMR (400 MHz, CDCI3): δ 7.77 (m, 2H), 7.18 (m, 2H), 7.05 (m, 1H), 6.75 (m, 2H), 5.28 ( m, 1H), 5.05 (m, 1H), 4.36 (s, 2H), 3.50 (m, 2H), 3.35 (m, 1H), 3.15 (m, 1H), 3.09 (s, 6H), 2.92 (m, 3H), 2.42 (m, 1H), 2.03 (m, 1H), 1.65 (m, 2H), 1.25 (m , 1H).

^)-3-((Cyclohexylmethyl)(^)-2-methyl-3-(methylsulfonyloxy)propyl)amino)-2-methylpropylmethanesulfonate (intermediate for compound A178):

Methanesulfonyl chloride (5, 6 g, 10.20 mmol) at 0-5°C. The reaction mixture was stirred for 1 hour at ambient temperature. The mixture was then diluted with DCM (100 ml) and washed with water (50 ml). Dry the organic layer over anhydrous sodium sulfate and concentrate to give N)-methyl-2-methyl-3-(methylsulfonyloxy)propanoate.

To a solution of N)-methyl-2-aminopropanoate hydrochloride (1.60 g, 10.20 mmol) in acetonitrile (200 ml) was added N)-methyl-2-methyl-3-(methylsulfonyloxy)propanoate (4.2 g, 21.43 mmol) and potassium carbonate (4.20 g, 30.60 mmol). The reaction mixture was refluxed for 16 hours, then filtered and concentrated to give (2R,2'S)-dimethyl-2,2'-azandiyl-bis(methylene)dipropanoate.

To a solution of (2^2^)-dimethyl-2,2'-azandiyl-bis-(methylene)dipropanoate (650 mg, 3.0 mmol) in DCM (10 ml) was added cyclohexanecarbaldehyde (336 mg, 3.0 mmol) and sodium triacetoxyborohydride (954 mg, 4.50 mmol). Stir the reaction mixture overnight at ambient temperature. The reaction was quenched with water and the pH of the mixture was adjusted to 10. The resulting mixture was extracted with DCM. The extract was concentrated and the residue was purified by column chromatography to give (2R,2'S)-dimethyl-2,2'-(cyclohexylmethylazandiyl)bis(methylene)dipropanoate.

To a solution of (2R,2'S)-dimethyl-2,2'-(cyclohexylmethylazandiyl)-bis-(methylene)dipropanoate (350 mg, 1.28 mmol) in THF (10 ml) was added LiBH4 (486 mg, 12.80 mmol ). The reaction mixture was stirred at 80°C overnight. The mixture was cooled and the reaction was quenched with water. The pH of the resulting mixture was adjusted to 5 with 1N. aqueous hydrochloric acid and stirred for 0.5 h, then adjusted to pH 10 and extracted three times with ethyl acetate. Dry the combined organic layers over anhydrous sodium sulfate and concentrate to give (2R,2'S)-2,2'-(cyclohexylmethylazandiyl)-bis(methylene)dipropan-1-ol.

In a solution of (2^2^)-2,2'-(cyclohexylmethylazandiyl)-bis-(methylene)dipropan-1-ol (153 mg, 0.59 mmol) and triethylamine (0.12 g, 1.2 mmol) in DCM (5 ml) was added MsCl (135 mg, 1.18 mmol) at 0°C. The reaction mixture was stirred for 3 hours at ambient temperature. Dilute the mixture with DCM (15 ml) and then wash with water (20 ml x 2). Dry the organic layers over anhydrous sodium sulfate and concentrate to give ^)-3-((cyclohexylmethyl)(^)-2-methyl-3(methylsulfonyloxy)propyl)amino)-2-methylpropyl methanesulfonate.

MS (ESI) C ₁₇ H ₃₅ NO ₆ S ₂ , experiment 414 [M+H]+.

The following compound was synthesized similarly: DO,2'b)-((cyclohexylmethyl)-azandiyl)-bis-(2methylpropan-3,1-diyl)dimethanesulfonate (intermediate for compound A160).

^)-6-chloro-NX4-(4-(dimethylamino)phenylsulfonamido)butan-2-yl)-3,4-dihydroisoquinoline-2(III)sulfonamide (intermediate for compound A148):

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A solution of N)-tert-butyl-1-cyanopropan-2-ylcarbamate (13.0 g, 71 mmol) in ethanol (250 mL), saturated with anhydrous ammonia, was treated with Raney Ni (24 g) at 55 psi (380 kPa) H ₂ overnight. The mixture was filtered through a pad of celite and the filtrate was concentrated. The residue was purified by column chromatography to obtain N)-tert-butyl-4-aminobutan-2-ylcarbamate.

A solution of 4 -(dimethylamino)benzene-1-sulfonyl chloride (2.93 g, 13.3 mmol) in DCM (15 ml) at 0-5°C. The reaction mixture was stirred for 1 hour at ambient temperature. Dilute the mixture with DCM (50 ml) and wash with water (30 ml). Dry the organic layer over anhydrous sodium sulfate and concentrate. The residue was purified by column chromatography to obtain N)-tert-butyl-4-(4-(dimethylamino)phenylsulfonamido)-butan-2ylcarbamate.

To a solution of N)-tert-butyl-4-(4-(dimethylamino)phenylsulfonamido)butan-2-ylcarbamate (1.2 g, 3.2 mmol) in DCM (15 ml) was added TFA (7 ml). The mixture was stirred for 2 hours and the solvent was removed. Dissolve the residue in DCM (30 ml) and then wash with saturated aqueous NaHCO ₃ . Dry the organic phase over anhydrous sodium sulfate and concentrate to give (R)-N-(3aminobutyl)-4-(dimethylamino)benzenesulfonamide (0.88 g, quantitative yield).

6-chloro-1,2,3,4-tetrahydroisoquinoline was added to a solution of N)-N-(3-aminobutyl)-4-(dimethylamino)benzenesulfonamide (0.88 g, 4.0 mmol) in CH3CN (10 ml). (1.98 g, 4.0 mmol). Stir the reaction mixture at ambient temperature overnight. Concentrate the mixture and purify the residue by column chromatography to obtain N)-6-chloro-N(4-(4-(dimethylamino)phenylsulfonamido)butan-2-yl)-3,4-dihydroisoquinoline-2(III)-sulfonamide.

The following compounds were synthesized similarly:

^)-6-chloro-N(4-((4-(dimethylamino)phenyl)sulfonamido)butan-2-yl)-3,4-dihydroisoquinoline2(1H)-sulfonamide (intermediate for compound A147);

^)-6-chloro-N(3-((4-(dimethylamino)phenyl)sulfonamido)butyl)-3,4-dihydroisoquinoline-2(III)sulfonamide (intermediate for compound A150) and

^)-6-chloro-N(3-((4-(dimethylamino)phenyl)sulfonamido)butyl)-3,4-dihydroisoquinoline-2(III)sulfonamide (intermediate for compound A149).

^)-1-(4-Fluoro-2-methoxyphenyl)-2-methylpiperazine (intermediate for compound A177):

To tert-butyl-^)-3-methylpiperazine-1-carboxylate (5.37 g, 26.8 mmol) and 1-bromo-4-fluoro-2methoxybenzene (5.00 g, 24.4 mmol) in toluene ( 50 ml) sodium tert-butoxide (4.69 g, 48.8 mmol) was added, followed by BrettPhos G3 palladacyclic (66.4 mg, 0.0732 mmol). The mixture was sealed under nitrogen and stirred at 100°C, and after 1 hour the mixture was cooled to ambient temperature, diluted with ethyl acetate and water. The organic layer was dried over sodium sulfate, filtered and concentrated. Purify the crude product by column chromatography (0-40%, ethyl acetate/hexanes) to give the product.

MS (ESI) for C ₁₇ H ₂₅ FN ₂ O ₃ , experiment 325 [M+H]+.

To tert-butyl-^)-4-(4-fluoro-2-methoxyphenyl)-3-methylpiperazine-1-carboxylate (0.51 g, 1.6 mmol) was added HCl (1.6 ml, 4N in dioxane ) and left the mixture to settle at ambient temperature for 16 hours, then diluted with water (5 ml) and ethyl acetate (5 ml). The organic layer was removed and washed with water (1X5 ml).

The combined aqueous layers were washed with ethyl acetate (1X5 ml), basified with NaOH (5N, to pH~12), extracted with DCM (3X5 ml), dried with sodium sulfate, filtered and concentrated to give ^)-1-(4-fluoro-2- methoxyphenyl)-2-methylpiperazine, which was further used without further purification.

MS (ESI) for C ₁₂ H ₁₇ FN ₂ O, experiment 225 [M+H] ⁺ .

The following compounds were synthesized similarly:

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1-(4-fluoro-2-methoxyphenyl)piperazine (intermediate A137) 1-(2-methoxyphenyl)piperazine (intermediate A154) 2-(piperazin-1-yl)benzonitrile (intermediate A118) 1-( 2,6-dimethylphenyl)piperazine (A119 intermediate) 2-(4-fluoro-2-methyl)-2,6-diazaspiro[3.3]heptane (A161 intermediate) (8)-1-(4-fluoro) -2-methylphenyl)-3-methylpiperazine (intermediate for

A162) (C)-1-(4-fluoro-2-methylphenyl)-3-methylpiperazine (intermediate for A163)

1-(2-(trifluoromethoxy)phenyl)piperazine (intermediate for A173) 1-(3,4-difluoro-2-methoxyphenyl)piperazine (intermediate for A174) 1-(3-fluoro-2-methoxyphenyl)piperazine ( intermediate for A175) 1-(5-fluoro-2-methoxyphenyl)piperazine (intermediate for A176) 1-(2-chlorophenyl)piperazine (intermediate for A198) 1-(4-methoxyphenyl)piperazine (intermediate for A199 ) 1-(3-methoxyphenyl)piperazine (intermediate for A201) 1-(3-fluoro-4-methoxyphenyl)piperazine (intermediate for A202) 1-(2-chloro-4-fluorophenyl)piperazine (intermediate for A206 ) (8)-1-(3-fluoro-5-methoxyphenyl)-2-methylpiperazine (intermediate for A194) (R)-l-(3-fluoro-5-methoxyphenyl)-2-methylpiperazine (intermediate for

A195) (28.6K)-1-(3-fluoro-5-methoxyphenyl)-2,6-dimethylpiperazine (intermediate for A197) (2R, 6R)-1 -(3-fluoro-5-methoxyphenyl)-2 ,6-d methylpiperazine (intermediate for A200)

Research

Dox-induced PD1-ss-Gluc study.

Flp-In 293 T-REx™ cells were transfected with the plasmid pcDNA™5/FRT/TO, into which a cDNA encoding Gaussia luciferase hybridized to the 3' end of a cDNA encoding the PD1 signal sequence plus 10 amino acids was inserted (N-MQIPQAPWPVVWAVLQLGWRPGWFLDSPDR-C ) (SEQ ID NO: 1). Transfected cells were selected based on resistance to selected markers hygromycin and blasticidin to generate a stable cell line that contained the PD1-ss+10aa/Gaussia luciferase cDNA insert, the expression of which was regulated under the conditions of the T-REx™ system. The day before the study, cells were trypsinized and placed in 384-well tissue culture plates. The next day, compound diluted in DMSO/medium containing doxycycline was added to the wells and incubated at 37°C, 5% CO ₂ . After 24 h, coelenterazine substrate was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine activity.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

Dox-induced PD1-FL-Gluc study.

Flp-In 293 T-REx™ cells were transfected with the plasmid pcDNA™5/FRT/TO, into which a cDNA encoding firefly luciferase hybridized to the 3' end of a cDNA encoding full-length PD1 (amino acids 1-288) was inserted. Transfected cells were selected based on resistance to selected markers hygromycin and blasticidin to create a stable cell line that contained a PDl-FL/firefly luciferase cDNA insert, the expression of which was regulated under the conditions of the T-REx™ system. The day before the study, cells were trypsinized and placed in 384-well tissue culture plates. The next day, compound diluted in DMSO/medium containing doxycycline was added to the wells and incubated at 37°C, 5% CO2. After 24 h, coelenterazine substrate was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine activity.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

- 84 045973

Study of Dox-induced TNF-FL-Gluc.

Flp-In 293 T-REx™ cells were transfected with the plasmid pcDNA™5/FRT/TO, into which a cDNA encoding Gaussia luciferase hybridized to the 3' end of a cDNA encoding full-length TNF-α (amino acids 1-233) was inserted. Transfected cells were selected based on resistance to selected markers hygromycin and blasticidin to generate a stable cell line that contained a TNFα-FL/Gaussia luciferase cDNA insert, the expression of which was regulated under the conditions of the T-REx™ system. The day before the study, cells were trypsinized and placed in 384-well tissue culture plates. The next day, compound diluted in DMSO/medium containing doxycycline was added to the wells and incubated at 37°C, 5% CO ₂ . After 24 h, coelenterazine substrate was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine activity.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

Dox-induced IL2-FL-Gluc study.

Flp-In 293 T-REx™ cells were transfected with the plasmid pcDNA™5/FRT/TO, into which a cDNA encoding Gaussia luciferase hybridized to the 3' end of a cDNA encoding full-length IL-2 (amino acids 1-153) was inserted. Transfected cells were selected based on resistance to selected markers hygromycin and blasticidin to create a stable cell line that contained the I-2-PE/Gaussia luciferase cDNA insert, the expression of which was regulated under the conditions of the T-REx™ system. The day before the study, cells were trypsinized and placed in 384-well tissue culture plates. The next day, compound diluted in DMSO/medium containing doxycycline was added to the wells and incubated at 37°C, 5% CO ₂ . After 24 h, coelenterazine substrate was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine activity.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

Dox-induced HER3-ss-Gluc study.

Flp-In 293 T-REx™ cells were transfected with the plasmid pcDNA™5/FRT/TO, into which a cDNA encoding Gaussia luciferase hybridized with the 3' end of a cDNA encoding the HER3 signal sequence plus 4 amino acids was inserted (N-MRANDALQVLGLLFSLARGSEVG-C ) (SEQ ID NO: 2). Transfected cells were selected based on resistance to selected markers hygromycin and blasticidin to create a stable cell line that contained the HER3-ss+4aα/Gaussia luciferase cDNA insert, the expression of which was regulated under the conditions of the T-REx™ system. The day before the study, cells were trypsinized and placed in 384-well tissue culture plates. The next day, compound diluted in DMSO/medium containing doxycycline was added to the wells and incubated at 37°C, 5% CO ₂ . After 24 h, coelenterazine substrate was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine activity.

The results for the individual compounds proposed herein are shown in the table below. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

- 85 045973

HER3 Activity Table

No. Dox-induced Her3(ss+4) Glue 293FRT/TO at 24 hours: Average IC50 (nM) A122 110.63 A120 31.42 A137 10.85 A151 276.03 A177 10.86 A179 399.69 A124 912.67 A182 203.11 A183 2145.26 A184 155.56 A185 85.5 A107 44.21 A187 802.1 A188 88.04 A189 959.52 A190 35.6 A191 116.79 A192 1709.12 A193 50.85 A194 303.25 A195 381.8 A196 78.19 A197 77.48 A198 44.08 A199 131.52 A200 29.08 A201 211.04 A202 129.54 A203 34.13 A204 134.41 A205 230.38 A206 38.19 A207 17.98 A208 427.53 A209 497.91 A210 342.58 B22 57.77

H929 cell viability assay.

The human multiple myeloma cell line NCI-H929 was grown in Advanced RPMI 1640 medium (Gibco®) supplemented with 6% fetal bovine serum, 2 mM glutamine and 1x penicillin/streptomycin. On the day of the assay, cells were resuspended in RPMI 1640 medium supplemented with 10% fetal bovine serum, 2 mM glutamine and 1x penicillin/streptomycin and plated in 384-well tissue culture plates and treated with compound diluted in DMSO/medium. The plates were incubated at 37°C, 5% CO ₂ for 48 hours. After 48 hours, Celltiter-Glo® (Promega) was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine cell viability.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

U266 cell viability assay.

The human multiple myeloma cell line U266B1 was grown in RPMI 1640 medium supplemented with 10% fetal bovine serum, 2 mM glutamine and 1x penicillin/streptomycin. Cells were plated in 384-well tissue culture plates and treated with compound diluted in DMSO/medium. The plates were incubated at 37°C, 5% CO ₂ for 48 hours. After 48 hours, Celltiter-Glo® (Promega) was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine cell viability.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

In vitro microsomal stability studies.

Potassium phosphate (423 μl of a 0.1 M solution) was added to strips of 8 deep well tubes, followed by human, mouse, rat, or monkey microsomes (25 μl of a 20 mg/ml solution). Then the tubes were placed in ice and the test sample (2 μl of a 0.25 mM solution in DMSO) was added. The mixture was pre-incubated at 37°C for 3-5 min (shaking at 150 rpm), then the reaction was started

- 86 045973 tion by adding 50 µl of NADPH. Aliquots of samples (100 μL) were collected at 0 and 30 min, and 200 μL of acetonitrile mixture containing IS (compound 914) was added to quench the reaction. After centrifugation for 10 min at 4000 rpm, a mixed solution sample of 20 μL supernatant (20 μL) and 100 μL ACN/H2O (1:1) was injected for LC-MS/MS analysis. The stability of liver microsomes was assessed by the content of the remaining compound in % after 30 min, which was calculated using the following equation:

[^^analyte T=30/AUCIS)/(AUCanalyte T=0/AUCIS)]x100.

The results for the individual compounds proposed herein are shown in Table 1 below. 1. For chemical structures that include one or more stereoisomers but are illustrated without specifying the stereochemistry, the studies refer to the mixture of stereoisomers.

Investigation of the transiently expressed Dox-induced Gluc signal sequence.

Flp-In 293 T-REx™ cells were transfected with 500 ng of pcDNA™5/FRT/TO plasmids, into which a cDNA encoding Gaussia luciferase hybridized to the 3' end of a cDNA encoding the target signal sequences plus 10 amino acids was inserted. Transfected cells were incubated overnight, then the cells were trypsinized and plated into 384-well tissue culture plates. The next day, compound diluted in DMSO/medium containing doxycycline was added to the wells and incubated at 37°C, 5% CO2. After 24 h, coelenterazine substrate was added to each well and the luciferase signal was quantified using a Tecan Infinite M1000 Pro to determine activity.

Target Signal peptide sequence +10 amino acid T A87 A137 A151 TNFR1 MGLSTVPDLLLPLVLLELLVGIYPSG VIG (SEQ ID NO: 3) LVPHLGDR EK (SEQ ID NO: 4) 307 152 1625 TNFR2 MAPVAVWAALAVGLELWAAAHA (SEQ ID NO: 5) LPAQVAFTP Y (SEQ ID NO: 6) 564 186 2114 IL-17a MTPGKTSLVSLLLLLSLEAIVKA (SEQ ID NO: 7) GITIPRNPGC (SEQ ID NO: 8) 447 216 1816 IL-17Ra MGAARSPPSAVPGPLLGLLLLLLGVL APGGAS (SEQ ID NO: 9) LRLLDHRA LV (SEQ ID NO: 10) 424 176 1862 IL-23 a MLGSRAVMLLLLLPWTAQG (SEQ ID NO: 11) RAVPGGSSP A (SEQ ID NO: 12) 567 223 1924 IL-12b (p40) MCHQQLVISWFSLVFLASPLVA (SEQ ID NO: 13) IWELKKDV YV (SEQ ID NO: 14) 155 74 913 IL-23R MNQ VTIQWD A VIAL YILF S WCHG (SEQ ID NO: 15) GITNINCSG H 111 47 666

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(SEQ ID NO: 16) IL-6 MNSFSTSAFGPVAFSLGLLLVLPAAF PAP (SEQ ID NO: 17) VPPGEDSKD V (SEQ ID NO: 18) 66 38 437 IL-6R MLAVGCALLAALLAAPGAA (SEQ ID NO: 19) LAPRRCPAQ E (SEQ ID NO: 20) 78 48 445 GP130 MLTLQTWLVQALFIFLTTESTG (SEQ ID NO: 21) ELLDPCGYI S (SEQ ID NO: 22) 201 161 969 IL-1R1 MKVLLRLICFIALLIS S (SEQ ID NO: 23) LEADKCKE RE (SEQ ID NO: 24) 374 282 1474 IL-1R2 MLRLYVLVMGVSA (SEQ ID NO: 25) FTLQPAAHT G (SEQ ID NO: 26) 23 17 168 IL-IRa (al isoform) MEICRGGLRSHLITLLLFLFHSETIC (SEQ ID NO: 27) RPSGRKSSK M (SEQ ID NO: 28) 111 47 666 IL-2Ra (CD25) MDSYLLMWGLLTFIMVPGCQA (SEQ ID NO: 29) ELCDDDPPE I (SEQ ID NO: 30) 159 55 876 IFN-al MASPFALLMVLVVLSCKSSCSLG (SEQ ID NO: 31) CDLPETHSL D (SEQ ID NO: 32) 128 57 820 IFN-a2 MALTFALLVALLVLSCKSSCSVG (SEQ ID NO: 33) CDLPQTHSL G (SEQ ID NO: 34) 51 44 404 IFN-a4 MALSF SLLMAVLVLS YKSIC SLG (SEQ ID NO: 35) CDLPQTHSL G (SEQ ID NO: 36) 221 89 1129 IFN-b MTNKCLLQIALLLCF STTALS (SEQ ID NO: 37) MSYNLLGF LQ (SEQ ID NO: 818 288 4305

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38) IFNAR1 MMVVLLGATTLVLVAVAPWVLSAA AGG (SEQ ID NO: 39) KNLKSPQK V.E. (SEQ ID NO: 40) 14 6 145 IFNAR2 MLLSQNAFIFRSLNLVLMVYISLVFG (SEQ ID NO: 41) ISYDSPDYT D (SEQ ID NO: 42) 133 42 778 CTLA-4 MACLGFQRHKAQLNLATRTWPCTLL FFLLFIPVFC (SEQ ID NO: 43) KAMHVAQP AV (SEQ ID NO: 44) 259 112 1145 PD-L1 MRIFAVFIFMTYWHLLNA (SEQ ID NO: 45) FTVTVPKDL Y (SEQ ID NO: 46) 56 19 366 LAG3 MWEAQFLGLLFLQPLWVAPVKPLQP GAE (SEQ ID NO: 47) VPVVWAQE GA (SEQ ID NO: 48) 75 48 464 TIM3 MF SHLPFDC VLLLLLLLLTRS (SEQ ID NO: 49) SEVEYRAE VG (SEQ ID NO: 50) 282 132 1354 TIGIT MRWCLLLIWAQGLRQAPLASG (SEQ ID NO: 51) MMTGTIETT G (SEQ ID NO: 52) 162 79 737 CD96 MEKKWKYCAVYYIIQIHFVKG (SEQ ID NO: 53) VWEKTVNT E.E. (SEQ ID NO: 54) 12 7 107 VISTA MGVPTALEAGSWRWGSLLFALFLAA SLGPVAA (SEQ ID NO: 55) FKVATPYSL Y (SEQ ID NO: 56) 211 150 911 B7H3 MLRRRGSPGMGVHVGAALGALWFC LTGA (SEQ ID NO: 57) LEVQVPEDP V (SEQ ID NO: 58) 27 9 201 CD73 MCPRAARAPATLLLALGAVLWPAA GA (SEQ ID NO: 59) WELTILHTN D (SEQ ID NO: 60) 58 44 375

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PDGFRa MGTSHPAFLVLGCLLTGLSLILC (SEQ ID NO: 61) QLSLPSILPN (SEQ ID NO: 62) 90 37 565 VGFR2 MQSKVLLAVALWLCVETRA (SEQ ID NO: 63) ASVGLPSVS L (SEQ ID NO: 64) 684 213 2591 IL-7R MTILGTTFGMVFSLLQVVSG (SEQ ID NO: 65) ESGYAQNG DL (SEQ ID NO: 66) 58 16 392 EGFR MRPSGTAGAALLALLAALCPASRA (SEQ ID NO: 67) LEEKKVCQ GT (SEQ ID NO: 68) 60 47 412 HER3 MRAND ALQ VLGLLF SLARG (SEQ ID NO: 69) SEVGNSQA VC (SEQ ID NO: 70) 89 57 522 VEGF MNFLLSWVHWSLALLLYLHHAKWS QA (SEQ ID NO: 71) APMAEGGG QN (SEQ ID NO: 72) 111 51 645 TACE (ADAMI Ό MRQSLLFLTSVVPFVLA (SEQ ID NO: 73) PRPPDDPGF G (SEQ ID NO: 74) 66 40 391 ADAM10 MVLLRVLILLLSWAAGMGG (SEQ ID NO: 75) QYGNPLNK YI (SEQ ID NO: 76) 231 127 849

Study of cytokines in peripheral blood mononuclear cells (PBMCs).

Fresh human PBMCs were isolated from whole blood collected from 3 healthy donors using density gradient centrifugation followed by red blood cell lysis. PBMCs from each donor were examined separately. Cells were suspended in RPMI 1640 medium supplemented with 5% fetal bovine serum, 2 mM L-glutamine, 10 mM HEPES, and 1X penicillin/streptomycin. PBMCs were seeded at 200,000 cells/well in 96-well round-bottom plates either without stimulation or by stimulation with lipopolysaccharide (LPS, 1 μg/ml) or antibodies to human CD3 (plate-bound, 2 μg/ml) and CD28 human (soluble, 2 µg/ml). At the same time, the compound, serially diluted in DMSO/medium, was added and PBMCs were incubated at 37°C, 5% CO ₂ for 24 hours. After 24 hours, the plates were centrifuged and half the volume of supernatant was removed for the study of cytokines in an electrochemiluminescent immunoassay (U- PLEX® Biomarker Multiplex, Meso Scale Discovery). For viability assays, CellTiter-Glo® (Promega) was added to the remaining material in each well and quantitated on fluorescent plates.

Average IC50 (nM) [stimulation conditions] A87 A137 A133 A177 GM-CSF [anti-CD3/CD28] 165 25 404 19 IFNy [anti-CD3/CD28 antibody] 152 21 234 19 IL-Ιβ[LPS] >25000 >25000 >25000 >25000 IL-2 [anti-CD3/CD28] 31 8 197 10 IL-6 [LPS] 61 12 193 24 IL-23 [LPS] 200 79 1054 54 TNFa [anti-CD3/CD28 antibody] 82 17 216 13 Viability for CellTiter-Glo [no stimulation] >25000 >25000 >25000 >25000

Efficacy Study: Effect of Compound A87 on Tumor Growth and Body Weight in the B16F10 Model Compared to PD-1 Targeted Therapy.

Female C57BL/6 mice (7-8 weeks old) received a 0.1 ml subcutaneous injection of 5x106 B16F10 cells in the flank on day 0. On day 3, Compound A87 co-formulated with 10% EtOH/10% Kolliphor EL was administered at .V. 30 mg/kg QW, i.v. 10 mg/kg D1D2, or i.p. 15 mg/kg QOD. Anti-PD1 antibody (RMP1-14 from Bioxcell, 200 μg) was administered i.p. according to the QODx3 administration regimen. Tumor size was monitored using a digital caliper (Fowler) every 2-3 days until the size reached 1.5-2.0 cm and reported as volume (length x width x height). B16F10, a melanoma line, was obtained in

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ATSS. C57/BL6 mice were purchased from Charles Rivers Laboratories.

Table 1

Activity of compounds

No. Doxindus. PDlssGlu with 293FRT/T O after 24h: average IC50 (nM) Doxindus. PDIFLFlu with 293FRT/T O after 24h: average IC50 (nM) Doxindus. OHOaFL Glue 293FRT/T O after 24h: average IC50 (nM) Doxindus. IL2FLG1U with 293FRT/T O at 24h: average IC50 (nM) Lifesp. H929 CelltiterGlo after 24h: average EC50 (nM) Lifesp. U266 CelltiterGlo after 24h: average EC50 (nM) Stab. in liver microsomes - % rest. after 30 min with NADPH (%) Mouse Rat Monkey Man A1 1026.3 1396.1 ON THE. 1170.58 ON THE. ON THE. 0.21 0.12 0.18 0.18 A2 344.75 807.39 ON THE. 10141 ON THE. 0.09 0.59 0.09 0.04 AZ 241.49 637.58 11412 5080.51 ON THE. 0.09 0.37 0.83 0.25 A4 93.87 310.79 3763.69 1220.45 ON THE. 0.56 1D7 1.17 0.25 A5 651.38 ON THE. 8429.37 ON THE. 0.27 0.24 0.31 0.24 Ab 4747.33 ON THE. 24073.2 ON THE. 0.12 0.17 0.44 0.53 A7 2314.7 ON THE. 22428.6 ON THE. 0.17 0.23 0.99 0.29 A8 2681.93 ON THE. ON THE. ON THE. 0.17 0.61 0.19 0.09 A9 1106.83 ON THE. ON THE. ON THE. 0.34 0.14 0.66 0.09 AY 123.42 4687.09 2863.07 ON THE. 0.2 8.85 0.14 0.77 AN 365.31 865.78 ON THE. 623.7 ON THE. ON THE. 0.18 0.17 0.08 0.21

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A12 857.59 ON THE. ON THE. ON THE. 0.1 0.07 0.22 0.11 A13 25.01 1471.73 987.14 ON THE. 0.04 0.1 0.1 0.65 A14 640.11 1234.03 ON THE. 1163.27 ON THE. ON THE. 5.39 1.13 1.4 0.08 A15 95.1 3063.1 2955.87 14604.7 0.29 0.061 0.13 0.07 A16 108.07 6341.65 4977.83 ON THE. 2.09 2.62 41.1 o,i A17 942.97 ON THE. ON THE. ON THE. 0.41 1.06 0.36 0.49 A18 ON THE. ON THE. ON THE. 15473.2 0.05 o, and 0.05 0.04 A19 325.83 ON THE. ON THE. 21895.5 0.24 0.1 0.08 0.05 A20 206.77 ON THE. 12798.2 ON THE. 0.07 0.19 0.06 0.05 A21 184.02 10002.6 8288.81 ON THE. 1.77 0.62 0.11 0.14 A22 155.66 404.42 6377.93 370.18 6673.8 ON THE. 0.12 0.4 o, and 0.16

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A23 758.88 ON THE. ON THE. ON THE. 0.17 0.13 0.03 0.04 A24 ON THE. ON THE. ON THE. ON THE. 0.13 0.04 0.03 0.03 A25 435.17 ON THE. ON THE. ON THE. 0.32 4.16 0.2 0.12 A26 224.49 ON THE. 16068.5 ON THE. 3.4 1.84 0.13 6.65 A27 492.55 ON THE. ON THE. ON THE. 2.15 0.98 2.07 2.52 A28 5251.11 ON THE. 10426 ON THE. 0.06 0.14 0.06 A29 241.1 591.29 ON THE. 446.59 ON THE. ON THE. 32.4 43.3 19.5 39.6 AZO 792.84 ON THE. ON THE. ON THE. 0.35 0.28 0.03 0.12 A31 619.25 ON THE. ON THE. ON THE. 0.25 0.09 0.03 0.05 A32 ON THE. ON THE. ON THE. ON THE. 0.51 0.27 0.09 0.06 AZZ ON THE. ON THE. ON THE. ON THE. 0.08 0.13 0.07 0.05

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A34 ON THE. ON THE. ON THE. ON THE. 0.24 0.12 0.09 0.03 AZ 5 83.03 243.99 > 11897.98 ON THE. ON THE. 20.9 30 13.6 24.8 AZ 6 118.49 319.35 > 21809.41 300.59 > 15385.93 ON THE. 1.85 0.94 1.54 6.69 AZ 7 202.16 ON THE. ON THE. ON THE. 6.03 13.5 5.69 8.26 AZ 8 75.12 5490.99 > 11844.59 ON THE. 12.6 24.2 9.04 I.4 AZ 9 761.18 ON THE. ON THE. 10421.2 0.26 6 o, and 0.05 A40 308.26 > 15070.18 ON THE. ON THE. 0.19 0.12 0.04 0.04 A41 17532.8 ON THE. ON THE. ON THE. 0.2 0.14 0.1 0.08 A42 696.43 ON THE. ON THE. ON THE. 0.79 7.69 0.81 2.65 A43 46.87 160.55 1208.05 132.63 2974.53 ON THE. 20.1 22.3 12 15.4 A44 132.1 12652.5 ON THE. ON THE. 43.8 54 38 47.6

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A46 50.13 1758.33 3336.62 ON THE. 3.63 4.46 1.44 6.12 A47 258.84 ON THE. ON THE. ON THE. 5.68 10.7 1.68 12.6 A48 991.35 ON THE. ON THE. ON THE. 0.07 0.4 0.1 0.19 A49 407.05 ON THE. ON THE. ON THE. I.8 16.4 9.36 3.5 A50 22.21 74.12 660.73 56.97 1007.74 6536.29 70.1 3.6 0.79 1.32 A51 61.36 3068.97 6153.41 ON THE. 28 34.7 23.6 26.7 A52 45.56 1035.16 2105.88 ON THE. 29.3 40.1 38.3 31 A53 35.33 1014.07 1717.71 > 10260.36 39.2 50.5 39 38 A54 26.51 952.37 1907.23 ON THE. 34.6 42.4 30.5 33.8 A55 1540.39 ON THE. ON THE. ON THE. 1.18 0.97 0.34 4.87 A56 32.76 1255.87 1634.05 ON THE. 53.2 57.3 42.6 44.5

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A57 73.41 7807.65 ON THE. ON THE. 60.2 84 28.5 31.9 A58 72.14 8500.05 ON THE. ON THE. 72.8 66.7 53.1 31.6 A59 24.23 615.54 1442.63 ON THE. 3.73 3.9 1.94 1.89 A60 18.54 886.17 1483.8 ON THE. 60.8 50.3 38.7 46.7 A61 955.94 ON THE. ON THE. ON THE. 17.9 19.6 8.96 13.3 A63 43.65 1873.85 7433.97 ON THE. 85.2 88.2 84.4 76.4 A64 74.98 3293.48 6402.59 ON THE. 53.1 63.1 48 53.3 A65 153.77 8715.63 ON THE. ON THE. 73.4 78 57.8 53.1 A66 187.59 3495.74 9211 ON THE. 0.02 0.39 0.35 0.13 A67 40.77 937.29 2398.75 ON THE. 28 45.4 27.6 33.2 A68 156.12 6440.61 ON THE. ON THE. 55.1 57.1 57.1 50.5

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A69 136.39 ON THE. ON THE. ON THE. 64.6 74.3 58.2 63 A70 46.08 1301.94 3023.7 ON THE. 3.14 6.75 3.26 1.36 A71 63.32 2014.74 7724.78 ON THE. I.5 17.2 6.75 5.55 A72 96.18 6972.14 10286.3 ON THE. 43.3 54.6 53.2 62.5 A73 37.27 1861.53 3471.61 ON THE. 16.3 71.5 25.4 18.2 A74 77.38 257.97 ON THE. 248.26 ON THE. ON THE. 53.7 55.7 60.6 56.3 A75 104.41 255.94 3782.3 233.05 3029.96 ON THE. 2.75 7.54 0.28 3.08 A76 ON THE. ON THE. ON THE. ON THE. ON THE. ON THE. 28.6 33.9 35.7 53.1 A77 95.05 338 5544.21 359.68 4012.61 ON THE. 37.3 32.5 20.8 30.1 A78 359.83 ON THE. ON THE. ON THE. 0.15 0.25 o, and 0.2 A79 408.23 ON THE. ON THE. ON THE. 10.6 0.1 0.06 0.16

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A80 272.95 ON THE. ON THE. ON THE. 13.4 14.1 2.56 10.8 A81 476.93 ON THE. ON THE. ON THE. 0.12 0.12 0.03 0.2 A82 30.52 1547.55 2298.14 ON THE. 0.08 0.16 0.23 o,w A83 28.17 961.05 959.61 ON THE. 23.4 47.6 34.9 39.3 A84 175.94 ON THE. ON THE. ON THE. 68.3 67.3 65.8 60.6 A85 87.46 ON THE. ON THE. ON THE. 52.2 64.9 42.9 25.6 A86 428.72 ON THE. 394.88 5625.91 ON THE. 34.1 39.9 29.2 41.3 A87 19.15 492.19 50.23 223.57 ON THE. 49.6 43.6 32.7 39.7 A88 4330.54 ON THE. ON THE. ON THE. ON THE. 15.9 15.2 1.33 I.7 A89 82.05 1208.13 186.53 1751.25 ON THE. 17.1 22.2 13.5 18.5 A90 207.02 2413.57 493.18 3786.86 ON THE. 28.2 20.2 14.1 20.1

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A91 193.42 2459.27 376.81 3011.92 14653.1 25.4 21.6 21.8 23.7 A92 304.02 ON THE. 518.65 ON THE. ON THE. 40.8 51.8 40.4 44.3 A93 670.27 ON THE. 911.37 10427.9 ON THE. 0.4 0.51 0.18 0.84 A94 36.37 550.17 78.29 800.16 ON THE. 45.7 45.5 30.3 42.1 A95 198.02 4897.04 390.92 6711.7 ON THE. 54.2 43.1 46.1 43.9 A96 121.85 1847.65 231.78 5104.13 ON THE. 37.6 41.9 32.8 40.5 A97 41.78 565.68 87.04 725.76 ON THE. 77.7 71.4 66.1 71.1 A98 201.79 7335.28 351.2 ON THE. ON THE. 55.2 32.8 26.9 57.5 A99 115.58 3535.19 349.8 ON THE. ON THE. 60.8 50.8 37 41.9 AYU 0 1150.93 ON THE. 1560.04 ON THE. ON THE. 0.1 2.18 0.04 0.22 AY 1 220.23 3425.89 564.3 ON THE. ON THE. 20.8 25.4 18.8 30.2

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AYU 2 33.39 477.31 90.53 558.29 ON THE. 86.6 101.9 70 62.3 AYU 3 445.68 ON THE. 698.23 ON THE. ON THE. 42.2 36.5 38.5 31.6 AYU 6 52.35 1118.61 126.18 902.63 ON THE. 8.81 18.9 30.7 7.18 AYU 7 26.11 609.25 72.97 1176.64 ON THE. 16.1 19.3 25.0 83.9 AYU 8 29.58 546.31 68.29 420.52 ON THE. 0.62 2.05 0.88 1.06 AN 8 67.79 2093.86 172.7 5345.11 ON THE. 22.1 18.3 14.7 14.2 AN 9 52.34 1077.79 170.3 7675.81 ON THE. 50.8 66.1 53.9 71.7 A12 0 15.58 238.67 27.34 672.16 ON THE. 19.2 28.5 16.3 4.78 A12 2 51.93 885.26 134.57 2420.21 ON THE. 36.4 0.17 0.15 0.38 A12 3 13.98 424.92 45.93 834.51 ON THE. 6.13 7.41 3.47 5.02 A12 4 606.76 ON THE. 802.05 ON THE. ON THE. 1.01 1.25 1.91 18.9

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A12 5 829.42 11524.98 1090.55 ON THE. ON THE. 0.14 0.37 8.67 0.2 A13 1 145.44 2403.07 426.19 4744.68 ON THE. 101 73.9 70.2 84.5 A13 3 36.84 748.55 134.51 5121.95 ON THE. 82.2 87.3 67.3 71.3 A13 4 11.04 309.22 27.28 540.66 ON THE. 27.9 41.3 9.76 31.9 A13 5 21.58 625.08 48.58 1239.26 ON THE. 26.1 56.3 39.2 9.36 A13 6 26.46 543.65 101.39 1726.93 ON THE. 14.3 26.6 23.2 65.3 A13 7 8.30 191.24 21.45 281.93 ON THE. 28 31.6 14.2 48.8 A13 8 73.91 947.28 156.27 2396.92 ON THE. 13.2 17.1 12.5 8.84 A14 0 491.56 3009.42 808.08 ON THE. ON THE. 3.68 0.56 1.56 0.41 A14 1 431.02 4235.2 862.9 9106.16 ON THE. 0.48 1.14 0.66 0.26 A14 2 38.25 741.05 67.02 1446.85 ON THE. 16 22.6 3.89 11.8

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A14 7 4122.89 ON THE. ON THE. ON THE. ON THE. 0.74 1.16 0.99 2.76 A14 8 925.61 ON THE. 1117.32 ON THE. ON THE. 1.59 1.29 0.67 0.52 A14 9 ON THE. ON THE. ON THE. ON THE. ON THE. 0.83 0.47 0.96 0.44 A15 0 30.29 467.79 41.34 2000.66 ON THE. 1.38 2.02 2.2 4.6 A15 1 152.57 1216.54 305.64 2711.98 ON THE. 4.55 4.2 3.98 4.28 A15 2 81.5 1028.9 173.82 2171.97 ON THE. 5.06 3.6 3.22 3.69 A15 3 1293.98 ON THE. 1255.35 ON THE. ON THE. 0.87 6.23 1.26 2.57 A15 4 14.94 504.45 58.66 749.43 ON THE. 17.5 12.3 10.2 7.33 A15 5 1350.05 3484.03 7795.99 ON THE. 16.8 38.8 4.86 17.8 A15 6 92.46 1825.18 206.72 3707.48 ON THE. 75.6 82.1 80.6 81.3 A15 7 23.33 584.47 77.41 858.16 ON THE. 23.3 32.2 27.3 21.1

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A16 0 152.83 10687.02 452.72 ON THE. ON THE. 61.3 51.1 46.1 71.8 A16 1 577.46 ON THE. 725.52 ON THE. ON THE. 26.5 27.3 16.7 20.2 A16 2 101.25 6263.53 200.98 ON THE. ON THE. 62.5 69.6 60.3 70.5 A16 3 55.56 1928.36 81.21 13622.1 ON THE. 72.8 54.9 57.5 64.2 A16 4 454.69 ON THE. 974.47 ON THE. ON THE. 69.0 57.9 40.9 64.8 A16 5 131.14 1857.05 293.51 4086.76 ON THE. 14.9 I.9 12.8 2.23 A16 6 16.68 278.28 30.25 547.83 ON THE. 26.4 31.3 18.7 14.0 A16 7 200.61 2656.44 412.45 6495.51 ON THE. 18.3 9.19 73 5.68 A16 8 1491.98 ON THE. 1144.17 ON THE. ON THE. 42 42.1 73 25.7 A16 9 81.52 1048.04 192.27 3272.71 ON THE. 22.5 13.1 4.23 34.7 A17 0 56.13 797.63 133.66 1775.44 ON THE. 3.65 10.8 1.37 7.2

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A17 1 ON THE. ON THE. ON THE. ON THE. ON THE. 0.91 0.90 0.26 0.60 A17 2 29.18 436.2 66.4 1750.08 ON THE. 3.65 2.23 2.63 3.46 A17 3 78.83 5638.24 248.3 ON THE. ON THE. 49.2 53.8 55.9 56.1 A17 4 15.79 565.56 53.31 1861.24 ON THE. 36.9 44.4 28.4 41.7 A17 5 9.35 393.21 40.35 1436.98 ON THE. 6.73 6.28 2.27 4.26 A17 6 17.11 631.2 62.1 3109.99 ON THE. 32.4 23.0 15.2 31.9 A17 7 4.95 154.54 12.57 179.88 ON THE. I.6 14.5 2.8 13.7 A17 8 134.76 4820.51 255.26 ON THE. ON THE. 22.8 29.2 25.8 24.0 A17 9 269.79 3249.91 72.57 11347.74 ON THE. 0.5 0.18 0.26 0.04 A18 0 2742.03 ON THE. 13843.24 20179.62 ON THE. 0.44 1.97 0.08 0.80 A18 1 38.69 983.85 122.41 1479.93 ON THE. 17.1 22.0 27.9 23.3

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A18 2 78.84 1957.98 255.55 ON THE. ON THE. 61.7 41.9 41.5 53.7 A18 3 1443.55 2524.48 6116.1 ON THE. 7.39 3.17 0.75 Yu.1 A18 4 100.13 1079.53 232.68 6916.08 ON THE. 5.64 7.47 1.23 2.56 A18 5 43.18 573.1 85.38 1468.19 ON THE. 30.1 37.8 10.0 21.8 A18 7 427.09 6171.01 1087.5 3866.88 ON THE. 0.03 0.07 0.15 0.05 A18 8 38.94 714.46 99.28 1555.52 ON THE. 4.48 27.8 12.4 27 A18 9 512.4 8138.23 1358.47 10609.19 ON THE. 0.14 0.07 0.01 0.06 A19 0 16.23 294.54 45.7 867.11 ON THE. 69.3 85.3 71.6 74.1 A19 1 57.62 1095.62 144.64 6734.93 ON THE. 55.3 65.0 68.3 48.6 A19 2 1153.54 ON THE. 2332.92 9362.89 ON THE. 0.18 0.08 0.10 0.02 A19 3 23.8 748.16 81.77 6806.9 ON THE. Yu,1 and,z 7.72 34.2

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A19 4 261.32 ON THE. 401.51 ON THE. ON THE. 68.5 59.2 65.0 63.6 A19 5 210.96 11201.43 668.05 ON THE. ON THE. 58.5 63.3 32.8 53.6 A19 6 30.96 1474.7 104.78 ON THE. ON THE. A19 7 34.25 666.07 95.53 4125.97 ON THE. 65.4 76.6 92.8 70.7 A19 8 18.54 372.95 53.28 1170.31 ON THE. 68.2 61.4 65.1 39.2 A19 9 80.8 1420.53 162.14 ON THE. ON THE. 23.7 28.3 25.5 8.0 A20 0 11.23 284.85 27.87 435.72 ON THE. 71.9 83.1 73.9 78.1 A20 1 117.7 7086.36 282.88 ON THE. ON THE. 39.1 35.4 26.5 32.8 A20 2 56.59 2052.14 175.52 2914.63 ON THE. 55.0 50.8 38.3 13.3 A20 14.76 434.38 30.44 396.26 ON THE. 80.3 78.7 77.9 83.4 A20 4 58.42 981.88 157.89 1487.11 ON THE. 62.6 58.3 49.7 48.4 A20 5 128.76 1806.9 305.32 2362.23 ON THE. 38.9 38.7 31.7

- 106 045973

37.2 A20 6 17.37 395.41 38.95 657.05 ON THE. 84.3 80.5 65 54.7 A20 7 7.93 221.47 10.6 105.74 ON THE. 84.3 98.8 63.0 62.7 A20 8 343 ON THE. 615.95 ON THE. ON THE. 51.8 68.9 37.5 23.3 A20 9 326.73 ON THE. 672.98 ON THE. ON THE. 31.8 44.7 45.7 22.6 A21 0 202.76 ON THE. 446.86 ON THE. ON THE. 76.0 54.6 55.4 47.4 Bl 423.7 ON THE. ON THE. ON THE. 0.02 0.01 0.02 0.04 B2 1721.99 ON THE. ON THE. ON THE. 0.55 1.46 0.23 0.35 VZ 7382.27 ON THE. 9441.64 12597.4 0.15 0.1 0.08 0.26 AT 4 ON THE. ON THE. ON THE. 16627.8 2.21 2.79 1.7 12.1 AT 5 1497.99 ON THE. ON THE. ON THE. 0.24 7.08 0.21 0.54

- 107 045973

Wb 416.05 ON THE. ON THE. ON THE. 81.8 13.2 4.57 7.66 AT 7 6900.19 24323 6858.59 7481.69 1.47 0.58 0.04 0.03 AT 8 ON THE. ON THE. ON THE. ON THE. 17.1 23.8 15.5 22.1 AT 9 3642.93 ON THE. ON THE. ON THE. 0.36 3.74 0.26 0.49 AT 10 16476.4 ON THE. 6923.68 ON THE. ON THE. IN AND ON THE. ON THE. ON THE. ON THE. ON THE. AT 12 5655.74 ON THE. ON THE. ON THE. ON THE. B16 449.25 13885 825.91 ON THE. ON THE. 58.4 72.3 44.7 55.2 B19 220.99 15364.2 ON THE. ON THE. 13.5 15.7 0.98 9.46 IN 20 794.07 ON THE. ON THE. ON THE. 3.41 ID 1.73 5.43 AT 21 3951.77 ON THE. ON THE. ON THE. ON THE. 0.16 0.06 0.03 0.17 B22 31.31 519.36 80.48 1648.2 ON THE. 8.31 Yu.1 1.77 1.77

*ON THE. indicates that the IC50 or EC50 is greater than 25 µM.

Claims (39)

1. A compound of formula (I) or a pharmaceutically acceptable salt thereof: where each of R ^a and R ^b independently represents H or C _1-3 alkyl; R ¹ represents H, OH, C _1-3 alkyl, OS _1-3 alkyl or =CH ₂ or R ¹ together with the atom in the ring to which it is attached forms or ; R ^1a and R ^1b are each independently H or C _1-3 alkyl; R ² is oxetanyl, azetidinyl, tetrahydrofuranyl, pyrrolidinyl, tetrahydropyranyl, pyranyl, piperidinyl, piperazinyl, azepanyl or tetrahydropyridinyl, or R ² has the structure: - 108 045973 - 109 045973 where R ⁶ represents C _1-6 alkyl, C _3-8 cycloalkyl or phenyl, optionally substituted with one to three groups independently selected from halogen, C _1-3 alkyl, C _|-3 alkoxy and CN; R ³ represents cyclopropyl, cyclobutyl, 3,3-difluorocyclobutyl, 4,4-dimethylcyclohexyl, 4-fluorocyclohexyl, 4,4-difluorocyclohexyl, cyclopentyl, cyclohexyl, cyclopentenyl, cyclohexenyl, tetrahydropyranyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl, furanyl, thiophenyl, thiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, pyridinyl, pyrazinyl or pyrimidinyl; R ⁴ is phenyl, optionally substituted with one to three groups independently selected from halogen, C 1-3 alkyl, C 1-3 alkoxy, C(O)N(R ⁿ ) 2 and N(R ⁿ ) 2 , and each R ^N is independently is H or C1-3αalkyl; X represents C _1-3 alkylene; Y represents SO or SO2. 2. The compound or salt according to claim 1, characterized in that each Y is SO. 3. The compound or salt according to claim 1, characterized in that each Y is SO2. 4. A compound according to any one of claims 1-3, characterized in that R ^a represents H. 5. A compound according to any one of claims 1-3, characterized in that R ^a represents C _1-3 alkyl. 6. A compound according to claim 5, characterized in that R ^a represents CH3. 7. A compound according to any one of claims 1 to 6, characterized in that R ^b represents H. 8. A compound according to any one of claims 1-6, characterized in that R ^b represents C _1-3 alkyl. 9. A compound according to claim 8, characterized in that R ^a represents CH3. 10. A compound or salt according to any one of claims 1 to 9, characterized in that R ¹ represents H or OH. 11. A compound or salt according to claim 10, characterized in that R ¹ represents H. 12. A compound or salt according to any one of claims 1 to 9, characterized in that R ¹ represents C1 _-3 alkyl or OC1 _-3 alkyl. 13. The compound or salt according to claim 12, characterized in that R ¹ represents CH3 or OCH ₃ . 14. The compound or salt according to claim 13, characterized in that R ¹ represents CH3 and has S-stereochemistry. 15. A compound or salt according to any one of claims 1 to 9, characterized in that R ¹ together with the atom in the ring to which it is attached forms, or 16. A compound or salt according to any one of claims 1 to 9, characterized in that R ¹ represents =CH ₂ . 17. The compound or salt according to any one of claims 1 to 16, characterized in that each of R ^1a and R ^1b is - 110 045973 is N. 18. A compound or salt according to any one of claims 1 to 16, characterized in that at least one of R ^1a and R ^1b is C ₁ _ ₃ alkyl. 19. The compound or salt according to claim 18, characterized in that R ^1a and R ^1b are each CH3. 20. A compound or salt according to any one of claims 1 to 19, characterized in that R ² contains oxetanyl, azetidinyl, tetrahydrofuranyl, pyrrolidinyl, tetrahydropyranyl, pyranyl, piperidinyl, piperazinyl, azepanyl, morpholinyl or tetrahydropyridinyl. 21. A compound according to any one of claims 1 to 19, characterized in that R ² is selected from the group consisting of and and ^R and R ⁶ is C ₁ . ₆ alkyl, C _3-8 cycloalkyl or phenyl. 22. The compound according to claim 21, characterized in that R ⁶ is phenyl substituted with one or three groups independently selected from halogen, C ₁ . ₃ alkyl, C _|-3 alkoxy and CN. 23. A compound or salt according to any one of claims 1 to 19, characterized in that R ² is selected from the group consisting of: - 111 045973 24. The compound or salt according to claim 23, characterized in that R ² is selected from the group consisting of: 25. A compound or salt according to any one of claims 1 to 24, characterized in that X represents CH ₂ , CH ₂ CH ₂ or CH(CH ₃ ). 26. A compound or salt according to claim 25, characterized in that R ³ contains cyclopropyl, cyclobutyl, - 112 045973 cyclopentyl, cyclohexyl, cyclopentenyl, cyclohexenyl, tetrahydropyranyl, phenyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl, furanyl, thiophenyl, thiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, pyridinyl, pyrazinyl or pyrimidinyl. 27. The compound or salt according to claim 26, characterized in that XR ³ is selected from the group consisting of: O--# 28. The compound or salt according to claim 27, characterized in that XR ³ is or 29. The compound or salt according to claim 28, characterized in that XR ³ is or . 30. A compound or salt according to any one of claims 1 to 29, characterized in that R ⁴ contains C6-10 aryl or C 2-9 heteroaryl and R ⁴ is optionally substituted with one to three groups independently selected from halogen, C 1-3 alkyl, C _{1- 3} alkoxy, C(O)N(R ⁿ )2 and N(R ⁿ )2, and each RN independently represents H or C _1-3 alkyl. 31. A compound or salt according to any one of claims 1 to 29, characterized in that R ⁴ is selected from the group consisting of: 32. The compound or salt according to claim 31, characterized in that R ⁴ represents 33. The compound or salt according to claim 1, characterized in that each R ^a , R ^b , R ^1a and R ^1b represents H; R ¹ represents =CH ₂ or CH ₃ ; / _Λ V Cl F. R ² represents ^{x Cl} > , ^- _ , H ^N _V _ _/ ^N “vj ^> , ; XR ³ is R ⁴ is or each Y represents SO ₂ . 34. A compound or salt according to any one of claims 1 to 33, characterized in that R ¹ has a ^-configuration. 35. Compound or pharmaceutically acceptable salt thereof: - 113 045973 No. Structure A5 Q.Pi A13 ~P. .tf} · Pi Ox , ^z— \ θ Al 5 Q P 00 dl Q z— / Al 7 J o-j d'N Al 9 Vx 3 -,-ΟΆγ A2 0 Ua Eyo 1J ⁰ 1 A21 I A22 to ₄ ri ,,,O da / A2 3 A° Z'-ZiSQ - 114 045973 - 115 045973 AZO уХХ Υ /'Ν'Υ А31 Α _ζ ^ζ '-ζ ° .€Ο b ο / AZ 2 ΥΧ?' _Ο AZZ χΟθ AZ 4 ^ ^Ν Υ/ Αν4° ^? Ό U'° - 116 045973 - 117 045973 - 118 045973 - 119 045973 - 120 045973 A61Ά. LA:-----. O .-----1 1-----·. -z—f zwz }—? 0 _chu AO b A63 0’ - G A64 T. X Ck fA \ X ^ъ A65 X.o A66 4---1 to L? <0 A O. i - 121 045973 - 122 045973 - 123 045973 - 124 045973 - 125 045973 - 126 045973 - 127 045973 A97 Ql 0 £?-° A98 QT Q 0=5=0 ...0jO A99 Φ 0 0=(=0 J7 ^Xj A100 ΟγΟ 0 0=^=0 ,JU' AIOI ζΊ] 0=5=0 - 128 045973 - 129 045973 - 130 045973 A112 °y-iX A Ν Q=5=Q ^i ^iХз jXX % 1 A113 M 0 χ blood pressure Azhu A114 Оу 0 0-4 □ Жж VA:?YX / О 4ΛΝ A115 1 Xj o+A o=s=o Q 0E - 131 045973 - 132 045973 - 133 045973 - 134 045973 - 135 045973 - 136 045973 - 137 045973 - 138 045973 Al 5 6 o I A ¹ *'' g---- °' χφ o -g X A157 vX ⁰ Т 1 ΓΥΎ ^ъ Г A160 O .. ,—\ ^g— X EE ^- * ( y — ^/ )— ⁷ o ^v A161 <1 m N O=S=O χτ^ 1 Al 62 Α, Ρ u l UK .9' b η - 139 045973 - 140 045973 - 141 045973 - 142 045973 - 143 045973 - 144 045973 - 145 045973 - 146 045973 36. A compound or salt according to claim 35, characterized in that said compound is selected from the group consisting of: - 147 045973 37. A compound or salt according to claim 36, having the structure 38. Compound or salt according to item 36, having the structure 39. A compound or salt according to claim 36, having the structure