WO2024078581A1

WO2024078581A1 - Selective bcl-xl protac compounds and uses thereof

Info

Publication number: WO2024078581A1
Application number: PCT/CN2023/124262
Authority: WO
Inventors: Zhengying Pan
Original assignee: Appicine Therapeutics (Hk) Limited
Priority date: 2022-10-12
Filing date: 2023-10-12
Publication date: 2024-04-18

Abstract

The present application relates to novel PROTAC compounds, and tautomers, stereoisomers, or pharmaceutically acceptable salts thereof, which degrade and/or inhibit the level or activity of BCL-XL proteins. The present application also relates to pharmaceutical compositions comprising one or more of the PROTAC compounds and tautomers, stereoisomers, or pharmaceutically acceptable salts thereof as an active ingredient, and to the use of the PROTAC compounds and tautomers, stereoisomers, or pharmaceutically acceptable salts thereof in the treatment of BCL-XL associated diseases, disorders or conditions, including cancers.

Description

SELECTIVE BCL-XL PROTAC COMPOUNDS AND USES THEREOF

FIELD OF THE DISCLOSURE

The present application relates to compounds comprising a BCL-XL proteins modulating moiety covalently linked to a degradation signaling moiety that binds to a degradation protein or degradation protein complex, e.g., an E3 ubiquitin ligase or an E3 ubiquitin ligase complex, the pharmaceutical compositions comprising the same and the use of the same in the treatment of diseases, disorders or conditions associated with BCL-XL proteins, including cancers.

BACKGROUND OF THE DISCLOSURE

BCL-2 (B-cell lymphoma 2) protein, encoded in humans by the BCL2 gene, is the founding member of the BCL-2 family of regulator proteins that regulate cell death (apoptosis) . B-cell lymphoma-extra large (BCL-XL) , encoded by the BCL2-like 1 gene, is a transmembrane molecule in the mitochondria. BCL-XL is a member of the BCL-2 family of proteins, and acts as an anti-apoptotic protein by preventing the release of mitochondrial contents such as cytochrome c, which leads to caspase activation and ultimately, programmed cell death (SJ Korsmeyer, “Regulators of Cell Death” , Trends in Genetics 11 (3) : 101-105, March 1995) .

The mechanism of action for BCL-XL modulators can be converted to degradation via PROTAC (Proteolysis-targeting chimeras) approach. These PROTAC degraders may be compounds comprising (i) a ligand targeting a protein of interest to be degraded and/or inhibited, (ii) an E3 ubiquitin ligase recruitment ligand (for example, Cereblon (CRBN) or Von Hippel-Lindau (VHL) ligands) , and (iii) a chemical linker covalently connecting the two ligands.

There remains a need for developing novel compounds as potent PROTACs that selectively degrade the BCL-XL protein.

SUMMARY OF THE DISCLOSURE

Disclosed herein are novel compounds that possess potent BCL-XL degradation activity. As a result, the compounds of the present application are particularly useful in the treatment of BCL-XL associated diseases, disorders or conditions.

In one aspect, the present disclosure provides compounds of Formula I:
D-L-E (I) ,

or tautomers, stereoisomers, or pharmaceutically acceptable salts thereof, wherein,

E is a degradation signaling moiety;

L is a linker that covalently attaches E to D; and

D is a drug moiety, e.g., a BCL-XL modulator of Formula II:

wherein

W is absent or selected from alkyl, alkenyl, or alkynyl, each of which is optionally substituted with one or more R’;

Q is selected from the group consisting of cycloalkyl, heterocyclyl, aryl and heteroaryl, wherein each of the cycloalkyl, heterocyclyl, aryl and heteroaryl is optionally substituted with one or more R” ;

each R’ is independently selected from the group consisting of halogen, cyano, hydroxyl, sulfhydryl, -NH₂, -NO₂, -SO₂-alkyl, -SO₂-haloalkyl, alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, haloalkyl, alkoxyl, haloalkoxyl, cycloalkyl, heterocyclyl, aryl and heteroaryl;

each R” is independently selected from the group consisting of halogen, cyano, hydroxyl, sulfhydryl, -NH₂, -NO₂, alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, haloalkyl, alkoxyl, haloalkoxyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -alkyl-R^a1, -alkyl-C (O) -R^a1, -C (O) -R^a1, -S (O) ₂-R^a1, -R^a2-NHR^a3 and -R^a2-NHC (O) R^a3;

R^a1, R^a2 and R^a3 are each independently selected from the group consisting of hydrogen, hydroxyl, halogen, alkyl, haloalkyl, alkoxyl, cycloalkyl and -alkyl-NH₂.

In another aspect, the present disclosure provides compounds of Formula I:
D-L-E (I) ,

E is a degradation signaling moiety;

L is a linker that covalently attaches E to D; and

D is a BCL-XL modulator of Formula (I-a) :

wherein Ring A is cyclohexyl or 6-membered heterocyclyl.

In a further aspect, the present disclosure provides a pharmaceutical composition, comprising the compound of Formula (I) or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

In a further aspect, the present disclosure provides a method of modulating the level or activity of BCL-XL protein in a cell, comprising exposing the cell to the compound of Formula (I) , or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition provided herein.

In a further aspect, the present disclosure provides a method of treating a subject having or suspected of having a cancer, comprising administering to the subject a therapeutically effective amount of the compound of Formula (I) or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition provided herein.

DETAILED DESCRIPTION OF THE DISCLOSURE

Reference will now be made in detail to certain embodiments of the present disclosure, examples of which are illustrated in the accompanying structures and formulas. While the present disclosure will be described in conjunction with the enumerated embodiments, it will be understood that they are not intended to limit the present disclosure to those embodiments. On the contrary, the present disclosure is intended to cover all alternatives, modifications, and equivalents, which may be included within the scope of the present disclosure as defined by the claims. One skilled in the art will recognize many methods and materials similar or equivalent to those described herein, which could be used in the practice of the present disclosure. The present disclosure is in no way limited to the methods and materials described. In the event that one or more of the incorporated references and similar materials differs from or contradicts this application, including but not limited to defined terms, term usage, described techniques, or the like, the present disclosure controls. All references, patents, patent applications cited in the present disclosure are hereby incorporated by reference in their entireties.

It is appreciated that certain features of the present disclosure, which are, for clarity, described in the context of separate embodiments, can also be provided in combination in a single embodiment. Conversely, various features of the present disclosure, which are, for brevity, described in the context of a single embodiment, can also be provided separately or in any suitable sub-combination. It must be noted that, as used in the specification and the appended claims, the singular forms “a, ” “an, ” and “the” include plural forms of the same unless the context clearly dictates otherwise. Thus, for example, reference to “a compound” includes both a single compound and a plurality of different compounds.

Definitions

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

Definitions of specific functional groups and chemical terms are described in more detail below. For purposes of this disclosure, the chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75^th Ed., inside cover, and specific functional groups are generally defined as described therein. Additionally, general principles of organic chemistry, as well as specific functional moieties and reactivity, are described in Organic Chemistry, Thomas Sorrell, 2^nd Edition, University Science Books, Sausalito, 2006; Smith and March March’s Advanced Organic Chemistry, 6^th Edition, John Wiley & Sons, Inc., New York, 2007; Larock, Comprehensive Organic Transformations, 3^rd Edition, VCH Publishers, Inc., New York, 2018; Carruthers, Some Modern Methods of Organic Synthesis, 4^th Edition, Cambridge University Press, Cambridge, 2004; the entire contents of each of which are incorporated herein by reference.

At various places in the present disclosure, linking substituents are described. It is specifically intended that each linking substituent includes both the forward and backward forms of the linking substituent. For example, -NR (CR’R”) -includes both -NR (CR’R”) -and - (CR’R”) NR-. Where the structure clearly requires a linking group, the Markush variables listed for that group are understood to be linking groups. For example, if the structure requires a linking group and the Markush group definition for that variable lists “alkyl” , then it is understood that the “alkyl” represents a linking alkylene group.

Unless defined otherwise, e.g., in the absence of symbols indicating specific point (s) of connectivity, when a structure or fragment of a structure is drawn, it may be used on its own or attached to other components of a compound, and it may do so with any orientation. When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, then such substituent may be bonded to any atom in the ring. When a substituent is listed without indicating the atom via which such substituent is bonded to the rest of the compound of a given formula, then such substituent may be bonded via any atom in such formula. Combinations of substituents and/or variables are permissible, but only if such combinations result in stable compounds.

When any variable (e.g., Rⁱ) occurs more than one time in any constituent or formula for a compound, its definition at each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group is shown to be substituted with 0-2 Rⁱ moieties, then the group may optionally be substituted with up to two Rⁱ moieties and Rⁱ at each occurrence is selected independently from the definition of Rⁱ. Also, combinations of substituents and/or variables are permissible, but only if such combinations result in stable compounds.

As used herein, the term “about” indicates that the values quoted are not to be construed as absolute, and measurement error, inter-batches variation and/or inter-apparatus variation should also be taken into account.

As used herein, the term “C_i-j” indicates a range of the carbon atoms numbers, wherein i and j are integers and the range of the carbon atoms numbers includes the endpoints (i.e. i and j) and each integer point in between, and wherein j is greater than i. For examples, C_1-6 indicates a range of one to six carbon atoms, including one carbon atom, two carbon atoms, three carbon atoms, four carbon atoms, five carbon atoms and six carbon atoms. In some embodiments, the term “C_1-12” indicates 1 to 12, particularly 1 to 10, particularly 1 to 8, particularly 1 to 6, particularly 1 to 5, particularly 1 to 4, particularly 1 to 3 or particularly 1 to 2 carbon atoms.

As used herein, the term “alkyl” , whether as part of another term or used independently, refers to a saturated linear or branched-chain hydrocarbon radical, which may be optionally substituted independently with one or more substituents described below. The term “C_i-j alkyl” refers to an alkyl having i to j carbon atoms. In some embodiments, alkyl groups contain 1 to 10 carbon atoms. In some embodiments, alkyl groups contain 1 to 9 carbon atoms. In some embodiments, alkyl groups contain 1 to 8 carbon atoms, 1 to 7 carbon atoms, 1 to 6 carbon atoms, 1 to 5 carbon atoms, 1 to 4 carbon atoms, 1 to 3 carbon atoms, or 1 to 2 carbon atoms. Examples of “C_1-10 alkyl” include, but are not limited to, methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, and decyl. Examples of “C_1-6 alkyl” are methyl, ethyl, propyl, isopropyl, n-butyl, i-butyl, s-butyl, t-butyl, n-pentyl, 2-pentyl, 3-pentyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 3-methyl-1-butyl, 2-methyl-1-butyl, 1-hexyl, 2-hexyl, 3-hexyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 3-methyl-3-pentyl, 2-methyl-3-pentyl, 2, 3-dimethyl-2-butyl, 3, 3-dimethyl-2-butyl, and the like.

As used herein, the term “alkenyl” , whether as part of another term or used independently, refers to linear or branched-chain hydrocarbon radical having at least one carbon-carbon double bond, which may be optionally substituted independently with one or more substituents described herein, and includes radicals having “cis” and “trans” orientations, or alternatively, “E” and “Z” orientations. In some embodiments, alkenyl groups contain 2 to 12 carbon atoms. In some embodiments, alkenyl groups contain 2 to 11 carbon atoms. In some embodiments, alkenyl groups contain 2 to 11 carbon atoms, 2 to 10 carbon atoms, 2 to 9 carbon atoms, 2 to 8 carbon atoms, 2 to 7 carbon atoms, 2 to 6 carbon atoms, 2 to 5 carbon atoms, 2 to 4 carbon atoms, 2 to 3 carbon atoms, and in some embodiments, alkenyl groups contain 2 carbon atoms. Examples of alkenyl group include, but are not limited to, ethylenyl (or vinyl) , propenyl, butenyl, pentenyl, 1-methyl-2 buten-1-yl, 5-hexenyl, and the like.

As used herein, the term “alkynyl” , whether as part of another term or used independently, refers to a linear or branched hydrocarbon radical having at least one carbon-carbon triple bond, which may be optionally substituted independently with one or more substituents described herein. In some embodiments, alkynyl groups contain 2 to 12 carbon atoms. In some embodiments, alkynyl groups contain 2 to 11 carbon atoms. In some embodiments, alkynyl groups contain 2 to 11 carbon atoms, 2 to 10 carbon atoms, 2 to 9 carbon atoms, 2 to 8 carbon atoms, 2 to 7 carbon atoms, 2 to 6 carbon atoms, 2 to 5 carbon atoms, 2 to 4 carbon atoms, 2 to 3 carbon atoms, and in some embodiments, alkynyl groups contain 2 carbon atoms. Examples of alkynyl group include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, and the like.

As used herein, the term “alkoxyl” , whether as part of another term or used independently, refers to an alkyl group, as previously defined, attached to the parent molecule through an oxygen atom. The term “C_i-j alkoxyl” means that the alkyl moiety of the alkoxy group has i to j carbon atoms. In some embodiments, alkoxy groups contain 1 to 10 carbon atoms. In some embodiments, alkoxy groups contain 1 to 9 carbon atoms. In some embodiments, alkoxy groups contain 1 to 8 carbon atoms, 1 to 7 carbon atoms, 1 to 6 carbon atoms, 1 to 5 carbon atoms, 1 to 4 carbon atoms, 1 to 3 carbon atoms, or 1 to 2 carbon atoms. Examples of “C_1-6 alkoxyl” include, but are not limited to, methoxy, ethoxy, propoxy (e.g. n-propoxy and isopropoxy) , t-butoxy, neopentoxy, n-hexoxy, and the like.

As used herein, the term “amino” refers to -NH₂ group. Amino groups may also be substituted with one or more groups such as alkyl, aryl, carbonyl or other amino groups.

As used herein, the term “aryl” , whether as part of another term or used independently, refers to monocyclic and polycyclic ring systems having a total of 5 to 20 ring members, wherein at least one ring in the system is aromatic and wherein each ring in the system contains 3 to 12 ring members. Examples of “aryl” include, but are not limited to, phenyl, biphenyl, naphthyl, anthracyl and the like, which may bear one or more substituents. Also included within the scope of the term “aryl” , as it is used herein, is a group in which an aromatic ring is fused to one or more additional rings. In the case of polycyclic ring system, only one of the rings needs to be aromatic (e.g., 2, 3-dihydroindole) , although all of the rings may be aromatic (e.g., quinoline) . The second ring can also be fused or bridged. Examples of polycyclic aryl include, but are not limited to, benzofuranyl, indanyl, phthalimidyl, naphthimidyl, phenanthridinyl, or tetrahydronaphthyl, and the like. Aryl groups can be substituted at one or more ring positions.

As used herein, the term “cycloalkyl” , whether as part of another term or used independently, refer to a monovalent non-aromatic, saturated or partially unsaturated monocyclic and polycyclic ring system, in which all the ring atoms are carbon and which contains at least three ring forming carbon atoms. In some embodiments, the cycloalkyl may contain 3 to 12 ring forming carbon atoms, 3 to 10 ring forming carbon atoms, 3 to 9 ring forming carbon atoms, 3 to 8 ring forming carbon atoms, 3 to 7 ring forming carbon atoms, 3 to 6 ring forming carbon atoms, 3 to 5 ring forming carbon atoms, 4 to 12 ring forming carbon atoms, 4 to 10 ring forming carbon atoms, 4 to 9 ring forming carbon atoms, 4 to 8 ring forming carbon atoms, 4 to 7 ring forming carbon atoms, 4 to 6 ring forming carbon atoms, 4 to 5 ring forming carbon atoms. Cycloalkyl groups may be saturated or partially unsaturated. Cycloalkyl groups may be substituted. In some embodiments, the cycloalkyl group may be a saturated cyclic alkyl group. In some embodiments, the cycloalkyl group may be a partially unsaturated cyclic alkyl group that contains at least one double bond or triple bond in its ring system. In some embodiments, the cycloalkyl group may be monocyclic or polycyclic. Examples of monocyclic cycloalkyl group include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopent-1-enyl, 1-cyclopent-2-enyl, 1-cyclopent-3-enyl, cyclohexyl, 1-cyclohex-1-enyl, 1-cyclohex-2-enyl, 1-cyclohex-3-enyl, cyclohexadienyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cycloundecyl and cyclododecyl. In some embodiments, the cycloalkyl group may be saturated or partially unsaturated polycyclic (e.g., bicyclic and tricyclic) carbocyclic ring system, which can be arranged as a fused-, spiro-or bridged-ring system. As used herein, the term “fused-ring” refers to a ring system having two rings sharing two adjacent atoms, the term “spiro-ring” refers to a ring systems having two rings connected through one single common atom, and the term “bridged-ring” refers to a ring system with two rings sharing three or more atoms. Examples of fused carbocyclyl include, but are not limited to, naphthyl, benzopyrenyl, anthracenyl, acenaphthenyl, fluorenyl and the like. Examples of spiro carbocyclyl include, but are not limited to, spiro [5.5] undecanyl, spiro-pentadienyl, spiro [3.6] -decanyl, and the like. Examples of bridged carbocyclyl include, but are not limited to bicyclo [1, 1, 1] pentenyl, bicyclo [2, 2, 1] heptenyl, bicyclo [2, 2, 1] heptanyl, bicyclo [2, 2, 2] octanyl, bicyclo [3, 3, 1] nonanyl, bicyclo [3, 3, 3] undecanyl, and the like.

As used herein, the term “cyano” refers to –CN.

As used herein, the term “halogen” refers to an atom selected from fluorine (or fluoro) , chlorine (or chloro) , bromine (or bromo) and iodine (or iodo) .

As used herein, the term “haloalkyl” , whether as part of another term or used independently, refers to an alkyl group having one or more halogen substituents. Examples of haloalkyl group include, but are not limited to, trifluoromethyl (-CF₃) , pentafluoroethyl (-C₂F₅) , difluoromethyl (-CHF₂) , trichloromethyl (-CCl₃) , dichloromethyl (-CHCl₂) , pentachloroethyl (-C₂Cl₅) , and the like.

As used herein, the term “haloalkoxyl” , whether as part of another term or used independently, refers to an alkoxyl group having one or more halogen substituents. As a result, the term “halo-C_i-j alkoxyl” , whether as part of another term or used independently, refers to a C_i-j alkoxyl group having one or more halogen substituents. Examples of haloalkoxyl include, but are not limited to, -O-CF₃, -O-C₂F₅, -O-CHF₂, -O-CCl₃, -O-CHCl₂, -O-C₂Cl₅, and the like.

As used herein, the term “heteroatom” refers to nitrogen, oxygen, or sulfur, and includes any oxidized form of nitrogen or sulfur, and any quaternized form of a basic nitrogen (including N-oxides) .

As used herein, the term “heteroalkyl” , “heteroalkenyl” , or “heteroalkynyl” , whether as part of another term or used independently, refers to an alkyl, alkenyl, or alkynyl group containing one or more heteroatoms. As a result, the term “hetero-C_i-j alkyl” , “hetero-C_i-j alkenyl” , or “hetero-C_i-j alkynyl” , whether as part of another term or used independently, refers to a C_i-j alkyl, C_i-j alkenyl, or C_i-j alkynyl containing one or more heteroatoms. For example, the term “hetero-C_1-6 alkyl” , whether as part of another term or used independently, refers to a C_1-6 alkyl containing one or more heteroatoms.

As used herein, the term “heteroaryl” , whether as part of another term or used independently, refers to an aryl group having, in addition to carbon atoms, one or more heteroatoms. The heteroaryl group can be monocyclic. Examples of monocyclic heteroaryl include, but are not limited to, thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, isothiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, indolizinyl, purinyl, naphthyridinyl, benzofuranyl and pteridinyl. The heteroaryl group also includes polycyclic groups in which a heteroaromatic ring is fused to one or more aryl, cycloaliphatic, or heterocyclyl rings, where the radical or point of attachment is on the heteroaromatic ring. Examples of polycyclic heteroaryl include, but are not limited to, indolyl, isoindolyl, benzothienyl, benzofuranyl, benzo [1, 3] dioxolyl, dibenzofuranyl, indazolyl, benzimidazolyl, benzthiazolyl, quinolyl, isoquinolyl, dihydroquinolinyl, dihydroisoquinolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 4H-quinolizinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, phenoxazinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, and the like.

As used herein, the term “heterocyclyl” refers to a saturated or partially unsaturated carbocyclyl group in which one or more ring atoms are heteroatoms independently selected from oxygen, sulfur, nitrogen, phosphorus, and the like, the remaining ring atoms being carbon, wherein one or more ring atoms may be optionally substituted independently with one or more substituents. In some embodiments, the heterocyclyl is a saturated heterocyclyl. In some embodiments, the heterocyclyl is a partially unsaturated heterocyclyl having one or more double bonds in its ring system. In some embodiments, the heterocyclyl may contains any oxidized form of carbon, nitrogen or sulfur, and any quaternized form of a basic nitrogen. “Heterocyclyl” also includes radicals wherein the heterocyclyl radicals are fused with a saturated, partially unsaturated, or fully unsaturated (i.e., aromatic) carbocyclic or heterocyclic ring. The heterocyclyl radical may be carbon linked or nitrogen linked where such is possible. In some embodiments, the heterocycle is carbon linked. In some embodiments, the heterocycle is nitrogen linked. For example, a group derived from pyrrole may be pyrrol-1-yl (nitrogen linked) or pyrrol-3-yl (carbon linked) . Further, a group derived from imidazole may be imidazol-1-yl (nitrogen linked) or imidazol-3-yl (carbon linked) .

In some embodiments, the term “3-to 12-membered heterocyclyl” refers to a 3-to 12-membered saturated or partially unsaturated monocyclic or polycyclic heterocyclic ring system having 1 to 3 heteroatoms independently selected from nitrogen, oxygen, or sulfur. The fused, spiro and bridged ring systems are also included within the scope of this definition. Examples of monocyclic heterocyclyl include, but are not limited to oxetanyl, 1, 1-dioxothietanylpyrrolidyl, tetrahydrofuryl, tetrahydrothienyl, pyrrolyl, furanyl, thienyl, pyrazolyl, imidazolyl, triazolyl, oxazolyl, thiazolyl, piperidyl, piperazinyl, piperidinyl, morpholinyl, pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl, pyridonyl, pyrimidonyl, pyrazinonyl, pyrimidonyl, pyridazonyl, pyrrolidinyl, triazinonyl, and the like. Examples of fused heterocyclyl include, but are not limited to, phenyl fused ring or pyridinyl fused ring, such as quinolinyl, isoquinolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, quinoxalinyl, quinolizinyl, quinazolinyl, azaindolizinyl, pteridinyl, chromenyl, isochromenyl, indolyl, isoindolyl, indolizinyl, indazolyl, purinyl, benzofuranyl, isobenzofuranyl, benzimidazolyl, benzothienyl, benzothiazolyl, carbazolyl, phenazinyl, phenothiazinyl, phenanthridinyl, imidazo [1, 2-a] pyridinyl, [1, 2, 4] triazolo [4, 3-a] pyridinyl, [1, 2, 3] triazolo [4, 3-a] pyridinyl groups, and the like. Examples of spiro heterocyclyl include, but are not limited to, spiropyranyl, spirooxazinyl, and the like. Examples of bridged heterocyclyl include, but are not limited to, morphanyl, hexamethylenetetraminyl, 3-aza-bicyclo [3.1.0] hexane, 8-aza-bicyclo [3.2.1] octane, 1-aza-bicyclo [2.2.2] octane, 1, 4-diazabicyclo [2.2.2] octane (DABCO) , and the like.

As used herein, the term “hydroxyl” or “hydroxy” refers to –OH.

As used herein, the term “nitro” refers to -NO₂.

As used herein, the term “sulfhydryl” refers to -SH.

As used herein, the term “sulfonyl” refers to –SO₂R’, wherein R’ is selected from hydrogen, alkyl, alkenyl, alkynyl, haloalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl.

As used herein, the term “Boc” refers to t-butoxyl carbonyl.

As used herein, the term “partially unsaturated” refers to a radical that includes at least one double or triple bond. The term “partially unsaturated” is intended to encompass rings having multiple sites of unsaturation, but is not intended to include aromatic (i.e., fully unsaturated) moieties.

As used herein, the term “substituted” , whether preceded by the term “optionally” or not, means that one or more hydrogens of the designated moiety are replaced with a suitable substituent. It will be understood that “substitution” or “substituted with” includes the implicit proviso that such substitution is in accordance with permitted valence of the substituted atom and that the substitution results in a stable or chemically feasible compound, e.g., which does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc. Unless otherwise indicated, an “optionally substituted” group may have a suitable substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted with more than one substituent selected from a specified group, the substituent may be either the same or different at every position. It will be understood by those skilled in the art that substituents can themselves be substituted, if appropriate. Unless specifically stated as “unsubstituted” , references to chemical moieties herein are understood to include substituted variants. For example, reference to an “aryl” group or moiety implicitly includes both substituted and unsubstituted variants.

Compounds

The present disclosure provides novel PROTAC compounds or tautomers, stereoisomers, or pharmaceutically acceptable salts thereof, synthetic methods for making the compounds, pharmaceutical compositions containing them and various uses of the disclosed compounds.

The PROTAC compounds of the present disclosure include a degradation signaling moiety covalently attached by a linker to a drug moiety (e.g., a BCL-XL modulator) .

The drug moiety when not conjugated to a degradation signaling moiety is capable of reducing the expression and/or activity of BCL-XL and/or one or more upstream modulators or downstream targets thereof.

The term “degradation signaling moiety” , as used herein, refers to degradation signaling compounds or moieties derived therefrom that induce degradation of target proteins, such as BCL-XL. The degradation signaling moiety of the present disclosure degrades targeted proteins by binding or recruiting at least one degradation protein, which is usually associated with the proteasome, the ubiquitin-proteasome pathways, or lysosomal proteolysis. In some embodiments, the degradation signaling moiety of the present disclosure is E3 ubiquitin ligase recognition or recruitment ligand.

By conjugating the drug moiety to a degradation signaling moiety (e.g., an E3 ubiquitin ligase) , an E3 ubiquitin ligase can ubiquitinate a target protein (e.g., BCL-XL) once the E3 ligase and the target protein are brought into proximity by the PROTAC compounds as described herein, which binds the E3 ubiquitin ligase via the degradation signaling moiety and the target protein, and accordingly, the target protein can be degraded by the E3 ubiquitin ligase. Without being bound by theory, by conjugating the drug moiety to a degradation signaling moiety that binds to an E3 ubiquitin ligase, the PROTAC compounds as described herein may provide improved activity, better cytotoxic specificity, and/or reduced off-target killing as compared to the drug moiety when administered alone.

In some embodiments, a PROTAC compound of the present disclosure is a compound of Formula I:
D-L-E (I) ,

E is a degradation signaling moiety;

L is a linker that covalently attaches E to D; and

D is a drug moiety (e.g., BCL-XL modulator) .

Drug moiety

In some embodiments, the drug moiety D in the PROTAC compounds of the present disclosure is a BCL-XL modulator.

In certain embodiments, the drug moiety D in the PROTAC compounds of the present disclosure is a BCL-XL inhibitor.

In some embodiments, the drug moiety D in the PROTAC compounds of the present disclosure is a BCL-XL modulator of Formula II:

wherein

In some embodiments, the drug moiety D in the PROTAC compounds of the present disclosure is a BCL-XL modulator Formula I-a:

wherein Ring A is cyclohexyl or 6-membered heterocyclyl.

In some embodiments, Ring A is cyclohexyl.

In certain embodiments, Ring A is

In some embodiments, Ring A is piperidinyl.

In some embodiments, Ring A is selected from

Degradation signaling moiety

Degradation signaling compounds and moieties of the present disclosure include compounds and moieties thereof that induce degradation of the targeted proteins (e.g., BCL-XL) .

In some embodiments, the degradation signaling moiety E in the PROTAC compounds of the present disclosure is an E3 ligase recognition agent.

In some embodiments, the degradation signaling moiety E in the PROTAC compounds of the present disclosure is a ligand of RING (Really Interesting New Gene) finger protein.

In some embodiments, the degradation signaling moiety E in the PROTAC compounds of the present disclosure is a VHL (Von Hippel-Lindau) ligand, a MDM2 (mouse double minute 2 homolog) ligand, a CRBN (Cereblon) ligand or an IAP (inhibitor of apoptosis) ligand.

In some embodiments, the degradation signaling moiety E in the PROTAC compounds of the present disclosure is an VHL ligand.

In certain embodiments, the degradation signaling moiety E in the PROTAC compounds of the present disclosure has a structure as following:

wherein

end of E is connected to L,

W¹ is

each W² is independently selected from the group consisting of hydrogen, halogen, cyano, hydroxyl, nitro, alkoxyl, alkyl, -O-cycloalkyl, aryl, heteroaryl, cycloalkyl and heterocyclyl, wherein the alkyl, aryl, heteroaryl, cycloalkyl and heterocyclyl is optionally substituted by one or more groups independently selected from halogen, cyano, hydroxyl, nitro, alkyl, haloalkyl, alkoxyl or haloalkoxyl;

each W³ is independently selected from the group consisting of hydrogen, haloalkyl, hydroxyalkyl, alkyl and cycloalkyl,

W^a and W^b are each independently selected from the group consisting of hydrogen, haloalkyl, hydroxyalkyl, alkyl and cycloalkyl, or W^a and W^b together with the carbon atom to which they are attached form a cycloalkyl,

W^c is selected from the group consisting of -NH-, aryl, heteroaryl, cycloalkyl and heterocyclyl, wherein the aryl, heteroaryl, cycloalkyl and heterocyclyl is optionally substituted by one or more groups independently selected from hydrogen, halogen, cyano, hydroxyl, nitro, alkyl, haloalkyl, alkoxyl or haloalkoxyl.

In some embodiments, one of W^a and W^b is hydrogen, the other is selected from hydrogen, alkyl or cycloalkyl. In certain embodiments, one of W^a and W^b is hydrogen, the other is C_1-6 alkyl.

In some embodiments, W^c is -NH-.

In some embodiments, each W² is independently alkyl, cycloalkyl, heterocyclyl or heteroaryl, each optionally substituted with one or more groups independently selected from halogen, cyano, hydroxyl, nitro, alkyl, haloalkyl or cycloalkyl.

In some embodiments, each W³ is independently selected from hydrogen, alkyl or cycloalkyl. In certain embodiments, one of W³ is hydrogen, and the other is alkyl or cycloalkyl.

In certain embodiments, the degradation signaling moiety E is

In some embodiments, the degradation signaling moiety E in the PROTAC compounds of the present disclosure is an CRBN ligand.

wherein

R^a is selected from hydrogen or alkyl; and

each R^b is hydrogen or two R^b together with the carbon atom to which they are attached form -C (O) -.

In some embodiments, R^a is selected from hydrogen or alkyl.

In some embodiments, R^a is selected from hydrogen or methyl.

In some embodiments, both R^b are hydrogen.

In some embodiments, two R^b together with the carbon atom to which they are attached form -C (O) -.

In some embodiments, the degradation signaling moiety E is selected from

Linker

In some embodiments, a linker compound is used to covalently attach a degradation signaling compound to a BCL-XL modulator compound to form the PROTAC compounds of the present disclosure comprising a degradation signaling moiety E and a BCL-XL modulator moiety D. The linker compound has at one end a reactive group that can react with the BCL-XL modulator compound and at the other end another reactive group that can react with the degradation signaling compound.

In some embodiments, the linker L in the PROTAC compounds of the present disclosure is of Formula (I-b) :
-L¹-L²-L³-L⁴- (I-b) ;

wherein,

L¹ is selected from the group consisting of a bond, - (CH₂) _pC (O) -, alkyl, alkenyl and alkynyl, wherein the alkyl, alkenyl and alkynyl is optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano, nitro or -N (R²) ₂;

L² is selected from the group consisting of a bond, -NH-C (O) -C (R¹) ₂-*, -N (R²) -*, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclyl and heteroaryl, wherein the alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclyl and heteroaryl is optionally substituted with one or more groups independently selected from halogen, oxo, hydroxyl, cyano, nitro or -N (R²) ₂, and *end of L² is connected to L³;

L³ is selected from the group consisting of a bond, - (OCH₂CH₂) _m-#, - (CH₂CH₂O) _m (CH₂) _n-#, alkyl, alkenyl and alkynyl, and # end of L³ is connected to L⁴;

L⁴ is selected from the group consisting of a bond, -N (R²) -, -O-, -C (O) -cycloalkyl and heterocyclyl;

each R¹ is independently selected from the group consisting of hydrogen, halogen, hydroxyl, cyano, nitro, alkyl and - (CH₂) _pN (R³) ₂;

each R² is independently selected from the group consisting of hydrogen, -C (O) R⁴, alkyl, alkenyl and alkynyl wherein the alkyl, alkenyl and alkynyl is optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano or nitro;

each R³ is independently selected from hydrogen or -C (O) R⁴;

each R⁴ is independently selected from -O-alkyl or alkyl;

m is 0, 1, 2, 3, 4, 5, 6, 7 or 8;

n is 0, 1, 2 or 3; and

p is 0, 1, 2, 3 or 4.

In some embodiments, L¹ is a bond.

In some embodiments, L¹ is alkyl optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano, nitro or -N (R²) ₂.

In some embodiments, L¹ is linear C_1-6 alkyl.

In some embodiments, L¹ is - (CH₂) _pC (O) -.

In some embodiments, L² is a bond.

In some embodiments, L² is -NH-C (O) -C (R¹) ₂-*.

In some embodiments, each R¹ is independently selected from hydrogen, alkyl or - (CH₂) _pN (R³) ₂.

In some embodiments, one R¹ is hydrogen and the other R¹ is - (CH₂) _pN (R³) ₂.

In some embodiments, both R³ are hydrogen.

In some embodiments, one R³ is hydrogen, the other R³ is -C (O) R⁴, wherein R⁴ is alkyl.

In some embodiments, R⁴ is methyl.

In some embodiments, L² is selected from the group consisting of:

In some embodiments, L² is -N (R²) -.

In some embodiments, R² is selected from hydrogen, -C (O) R⁴ or alkyl optionally substituted with one or more groups independently selected from halogen, cyano or hydroxyl.

In some embodiments, R² is hydrogen, -C (O) R⁴, or -alkyl-OH.

In some embodiments, R⁴ is -O-alkyl.

In some embodiments, R⁴ is -O-butyl.

In some embodiments, L² is selected from:

In some embodiments, L² is alkyl optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano, nitro or -N (R²) ₂.

In some embodiments, L² is methyl optionally substituted with one or more groups independently selected from halogen or -N (R²) ₂.

In some embodiments, each R² is independently selected from hydrogen or -C (O) R⁴.

In some embodiments, R⁴ is alkyl or -O-alkyl.

In some embodiments, R⁴ is methyl or -O-butyl.

In some embodiments, L² is selected from:

In some embodiments, L² is selected from cycloalkyl, aryl, heterocyclyl or heteroaryl.

In some embodiments, L² is selected from

In some embodiments, L³ is a bond.

In some embodiments, L³ is alkyl.

In some embodiments, L³ is - (OCH₂CH₂) _m-or - (CH₂CH₂O) _m (CH₂) _n-.

In some embodiments, L³ is selected from the group consisting of:

In some embodiments, L⁴ is a bond.

In some embodiments, L⁴ is -N (R²) -.

In some embodiments, R² is hydrogen.

In some embodiments, L⁴ is -O-.

In some embodiments, L⁴ is -C (O) -.

In some embodiments, L⁴ is heterocyclyl.

In some embodiments, L⁴ is

In some embodiments, L is selected from the group consisting of:

wherein **end of L is connected to E.

Exemplary PROTAC compounds of the present disclosure are set forth in Table 1 below.

Table 1. Structures of Exemplary PROTAC Compounds

Compounds provided herein are described with reference to both generic formulae and specific compounds. In addition, the compounds of the present disclosure may exist in a number of different forms or derivatives, including but not limited to, stereoisomers, racemic mixtures, regioisomers, tautomers, salts, prodrugs, soft drugs, active metabolic derivatives (active metabolites) , solvated forms, different crystal forms or polymorphs, all within the scope of the present disclosure.

The compounds of present disclosure can comprise one or more asymmetric centers, and thus can exist in various stereoisomeric forms, e.g., enantiomers and/or diastereomers. Thus, the compounds of present disclosure and compositions thereof may be in the form of an individual enantiomer, diastereomer or geometric isomer, or may be in the form of a mixture of stereoisomers. In certain embodiments, the compounds of the present disclosure are enantiopure compounds. In certain embodiments, mixtures of enantiomers or diastereomers are provided.

The term “enantiomer” refers to two stereoisomers of a compound which are non-superimposable mirror images of one another. The term “diastereomer” refers to a pair of optical isomers which are not mirror images of one another. Diastereomers have different physical properties, e.g. melting points, boiling points, spectral properties, and reactivities.

Furthermore, certain compounds, as described herein may have one or more double bonds that can exist as either the Z or E isomer, unless otherwise indicated. The present disclosure additionally encompasses the compounds as individual isomers substantially free of other isomers and alternatively, as mixtures of various isomers, e.g., racemic mixtures of enantiomers. In addition to the above-mentioned compounds per se, this disclosure also encompasses compositions comprising one or more compounds.

As used herein, the term “isomers” includes any and all geometric isomers and stereoisomers. For example, “isomers” include cis-and trans-isomers, E-and Z-isomers, R-and S-enantiomers, diastereomers, (D) -isomers, (L) -isomers, racemic mixtures thereof, and other mixtures thereof, as falling within the scope of the invention. For instance, a stereoisomer may, in some embodiments, be provided substantially free of one or more corresponding stereoisomers, and may also be referred to as “stereochemically enriched” .

Where a particular enantiomer is preferred, it may, in some embodiments be provided substantially free of the opposite enantiomer, and may also be referred to as “optically enriched” . “Optically enriched” , as used herein, means that the compound is made up of a significantly greater proportion of one enantiomer. In certain embodiments, the compound is made up of at least about 90%by weight of a preferred enantiomer. In other embodiments, the compound is made up of at least about 95%, 98%, or 99%by weight of a preferred enantiomer. Preferred enantiomers may be isolated from racemic mixtures by any method known to those skilled in the art, including chiral high performance liquid chromatography (HPLC) and the formation and crystallization of chiral salts or prepared by asymmetric syntheses. See, for example, Jacques, et al., Enantiomers, Racemates and Resolutions (Wiley Interscience, New York, 1981) ; Wilen, S.H., et al., Tetrahedron 33: 2725 (1977) ; Eliel, E.L. Stereochemistry of Carbon Compounds (McGraw-Hill, NY, 1962) ; Wilen, S.H. Tables of Resolving Agents and Optical Resolutions p. 268 (E.L. Eliel, Ed., Univ. of Notre Dame Press, Notre Dame, IN 1972) .

The compounds of the present disclosure may also exist in different tautomeric forms, and all such forms are embraced within the scope of the present disclosure. The term “tautomer” or “tautomeric form” refers to structural isomers of different energies which are interconvertible via a low energy barrier. The presence and concentrations of the isomeric forms will depend on the environment the compound is found in and may be different depending upon, for example, whether the compound is a solid or is in an organic or aqueous solution. By way of examples, proton tautomers (also known as prototropic tautomers) include interconversions via migration of a proton, such as keto-enol, amide-imidic acid, lactam-lactim, imine-enamine isomerizations and annular forms where a proton can occupy two or more positions of a heterocyclic system. Valence tautomers include interconversions by reorganization of some of the bonding electrons. Tautomers can be in equilibrium or sterically locked into one form by appropriate substitution. Compounds of the present disclosure identified by name or structure as one particular tautomeric form are intended to include other tautomeric forms unless otherwise specified.

As used herein, the term “prodrug” refers to compounds or pharmaceutically acceptable salts thereof which, when metabolized under physiological conditions or when converted by solvolysis, yield the desired active compound. Prodrugs include, without limitation, esters, amides, carbamates, carbonates, ureides, solvates, or hydrates of the active compound. Typically, the prodrug is inactive, or less active than the active compound, but may provide one or more advantageous handling, administration, and/or metabolic properties. For example, some prodrugs are esters of the active compound; during metabolism, the ester group is cleaved to yield the active drug. Also, some prodrugs are activated enzymatically to yield the active compound, or a compound which, upon further chemical reaction, yields the active compound. Prodrugs may proceed from prodrug form to active form in a single step or may have one or more intermediate forms which may themselves have activity or may be inactive. Preparation and use of prodrugs are discussed in T. Higuchi and V. Stella, “Pro-drugs as Novel Delivery Systems” , Vol. 14 of the A.C.S. Symposium Series, in Bioreversible Carriers in Drug Design, ed. Edward B. Roche, American Pharmaceutical Association and Pergamon Press, 1987; in Prodrugs: Challenges and Rewards, ed. V. Stella, R. Borchardt, M. Hageman, R. Oliyai, H. Maag, J. Tilley, Springer-Verlag New York, 2007, all of which are hereby incorporated by reference in their entireties.

As used herein, the term “soft drug” refers to compounds that exert a pharmacological effect but break down to inactive metabolites degradants so that the activity is of limited time. See, for example, “Soft drugs: Principles and methods for the design of safe drugs” , Nicholas Bodor, Medicinal Research Reviews, Vol. 4, No. 4, 449-469, 1984, which is hereby incorporated by reference in its entirety.

As used herein, the term “metabolite” , e.g., active metabolite overlaps with prodrug as described above. Thus, such metabolites are pharmacologically active compounds or compounds that further metabolize to pharmacologically active compounds that are derivatives resulting from metabolic process in the body of a subject. For example, such metabolites may result from oxidation, reduction, hydrolysis, amidation, deamidation, esterification, deesterification, enzymatic cleavage, and the like, of the administered compound or salt or prodrug. Of these, active metabolites are such pharmacologically active derivative compounds. For prodrugs, the prodrug compound is generally inactive or of lower activity than the metabolic product. For active metabolites, the parent compound may be either an active compound or may be an inactive prodrug.

Prodrugs and active metabolites may be identified using routine techniques known in the art. See, e.g., Bertolini et al., 1997, J Med Chem 40: 2011-2016; Shan et al., J Pharm Sci 86: 756-757; Bagshawe, 1995, Drug Dev Res 34: 220-230.

As used herein, the term “active intermediate” refers to an intermediate compound in the synthetic process, which exhibits the same or essentially the same biological activity as the final synthesized compound.

The compounds of the present disclosure are also intended to include all isotopic forms of the compounds. Isotopes of an atom include atoms having the same atomic number but different mass numbers. For example, unless otherwise specified, hydrogen, carbon, nitrogen, oxygen, phosphorous, sulfur, fluorine, chlorine, bromide or iodine in the compounds of present disclosure are meant to also include their isotopes, such as but not limited to ¹H, ²H, ³H, ¹¹C, ¹²C, ¹³C, ¹⁴C, ¹⁴N, ¹⁵N, ¹⁶O, ¹⁷O, ¹⁸O, ³¹P, ³²P, ³²S, ³³S, ³⁴S, ³⁶S, ¹⁷F, ¹⁸F, ¹⁹F, ³⁵Cl, ³⁷Cl, ⁷⁹Br, ⁸¹Br, ¹²⁴I, ¹²⁷I and ¹³¹I. In some embodiments, hydrogen includes protium, deuterium and tritium. In some embodiments, carbon includes ¹²C and ¹³C.

Compounds of the present disclosure can be formulated as or be in the form of pharmaceutically acceptable salts. Unless specified to the contrary, a compound provided herein includes pharmaceutically acceptable salts of such compound.

It is also to be understood that the compounds of present disclosure can exist in unsolvated forms, solvated forms (e.g., hydrated forms) , and solid forms (e.g., crystal or polymorphic forms) , and the present disclosure is intended to encompass all such forms.

As used herein, the term “solvate” or “solvated form” refers to solvent addition forms that contain either stoichiometric or non-stoichiometric amounts of solvent. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thus forming a solvate. If the solvent is water, then the solvate formed is a hydrate; and if the solvent is alcohol, then the solvate formed is an alcoholate. Hydrates are formed by the combination of one or more molecules of water with one molecule of the substance in which the water retains its molecular state as H₂O. Examples of solvents that form solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine.

As used herein, the terms “crystal form” , “crystalline form” , “polymorphic forms” and “polymorphs” can be used interchangeably, and mean crystal structures in which a compound (or a salt or solvate thereof) can crystallize in different crystal packing arrangements, all of which have the same elemental composition. Different crystal forms usually have different X-ray diffraction patterns, infrared spectral, melting points, density hardness, crystal shape, optical and electrical properties, stability and solubility. Recrystallization solvent, rate of crystallization, storage temperature, and other factors may cause one crystal form to dominate. Crystal polymorphs of the compounds can be prepared by crystallization under different conditions.

Synthesis of Compounds

Synthesis of the compounds provided herein, including tautomers, stereoisomers, pharmaceutically acceptable salts thereof, are illustrated in the synthetic schemes in the examples. The compounds provided herein can be prepared using any known organic synthesis techniques and can be synthesized according to any of numerous possible synthetic routes, and thus these schemes are illustrative only and are not meant to limit other possible methods that can be used to prepare the compounds provided herein. Additionally, the steps in the schemes are for better illustration and can be changed as appropriate. The embodiments of the compounds in examples were synthesized for the purposes of research and potentially submission to regulatory agencies.

The reactions for preparing compounds of the present disclosure can be carried out in suitable solvents, which can be readily selected by one skilled in the art of organic synthesis. Suitable solvents can be substantially non-reactive with the starting materials (reactants) , the intermediates, or products at the temperatures at which the reactions are carried out, e.g. temperatures that can range from the solvent’s freezing temperature to the solvent’s boiling temperature. A given reaction can be carried out in one solvent or a mixture of more than one solvent. Depending on the particular reaction step, suitable solvents for a particular reaction step can be selected by one skilled in the art.

Preparation of compounds of the present disclosure can involve the protection and deprotection of various chemical groups. The need for protection and deprotection, and the selection of appropriate protecting groups, can be readily determined by one skilled in the art. The chemistry of protecting groups can be found, for example, in T.W. Greene and P.G.M. Wuts, Protective Groups in Organic Synthesis, 3rd Ed., Wiley & Sons, Inc., New York (1999) , which is incorporated herein by reference in its entirety.

Reactions can be monitored according to any suitable method known in the art. For example, product formation can be monitored by spectroscopic means, such as nuclear magnetic resonance spectroscopy (e.g. ¹H or ¹³C) , infrared spectroscopy, spectrophotometry (e.g. UV-visible) , mass spectrometry, or by chromatographic methods such as high performance liquid chromatography (HPLC) , liquid chromatography-mass spectroscopy (LCMS) , or thin layer chromatography (TLC) . Compounds can be purified by one skilled in the art by a variety of methods, including high performance liquid chromatography (HPLC) ( “Preparative LC-MS Purification: Improved Compound Specific Method Optimization” Karl F. Blom, Brian Glass, Richard Sparks, Andrew P. Combs J. Combi. Chem. 2004, 6 (6) , 874-883, which is incorporated herein by reference in its entirety) , and normal phase silica chromatography.

The structures of the compounds in the examples are characterized by nuclear magnetic resonance (NMR) or/and liquid chromatography-mass spectrometry (LC-MS) . NMR chemical shift (δ) is given in the unit of 10^-6 (ppm) . ¹H-NMR spectra is recorded in CDCl₃, CD₃OD or DMSO-d₆ solutions (reported in ppm) on a Bruker instrument (400 MHz or 500 MHz) , using tetramethylsilane (TMS) as the reference standard (0.0 ppm) .

Unless otherwise specified, the reactions of the present disclosure were typically done under a positive pressure of nitrogen or argon or with a drying tube in anhydrous solvents, and the reaction flasks were typically fitted with rubber septa for the introduction of substrates and reagents via syringe. Glassware was oven dried and/or heat dried.

Pharmaceutical Compositions

The present disclosure provides pharmaceutical compositions comprising one or more compound of the present disclosure, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof. In some embodiments, the pharmaceutical composition comprises one or more compounds of the present disclosure, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, and at least one pharmaceutical acceptable excipient.

A “pharmaceutical composition” , as used herein, is a formulation containing the compounds of the present disclosure in a form suitable for administration to a subject. In some embodiments, the pharmaceutical composition is in bulk or in unit dosage form. The unit dosage form is any of a variety of forms, including, for example, tablets, capsules, pills, powders, granules, sachets, cachets, lozenges, suspensions, emulsions, solutions, syrups, aerosols (as a solid or in a liquid medium) , spray, ointment, paste, cream, lotion, gel, patch, inhalant, or suppository. The quantity of active ingredient (e.g., a formulation of the disclosed compound or salt, hydrate, solvate or isomer thereof) in a unit dose of composition is a therapeutically effective amount and is varied according to the particular treatment involved. One skilled in the art will appreciate that it is sometimes necessary to make routine variations to the dosage depending on the age and condition of the patient. The dosage will also depend on the route of administration. A variety of routes are contemplated, including oral, pulmonary, rectal, parenteral, transdermal, subcutaneous, intravenous, intramuscular, intraperitoneal, inhalational, buccal, sublingual, intrapleural, intrathecal, intranasal, and the like. Dosage forms for the topical or transdermal administration of a compound of this invention include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. In some embodiments, the compound of the present disclosure is mixed under sterile conditions with a pharmaceutically acceptable excipient, and with any preservatives, buffers or propellants that are required.

As used herein, the term “pharmaceutically acceptable excipient” means an excipient that is useful in preparing a pharmaceutical composition that is generally safe, non-toxic and neither biologically nor otherwise undesirable, and includes excipient that is acceptable for veterinary use as well as human pharmaceutical use. A “pharmaceutically acceptable excipient” as used in the specification and claims includes both one and more than one such excipient.

The term “pharmaceutically acceptable salt, ” as used herein, refers to a salt which does not abrogate the biological activity and properties of the compounds of the present disclosure, and does not cause significant irritation to a subject to which it is administered. Examples of such salts include, but are not limited to: (a) acid addition salts formed with inorganic acids, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid and the like; and salts formed with organic acids, for example, acetic acid, oxalic acid, tartaric acid, succinic acid, maleic acid, fumaric acid, gluconic acid, citric acid, malic acid, ascorbic acid, benzoic acid, tannic acid, palmitic acid, alginic acid, polyglutamic acid, naphthalenesulfonic acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, polygalacturonic acid, and the like; and (b) salts formed from elemental anions such as chlorine, bromine, and iodine. See, e.g., Haynes et al., “Commentary: Occurrence of Pharmaceutically Acceptable Anions and Cations in the Cambridge Structural Database, ” J. Pharmaceutical Sciences, vol. 94, no. 10 (2005) , and Berge et al., “Pharmaceutical Salts, ” J. Pharmaceutical Sciences, vol. 66, no. 1 (1977) , which are incorporated by reference herein.

As used herein, the term “therapeutically effective amount” refers to an amount of a pharmaceutical agent to treat, ameliorate, or prevent an identified disease or condition, or to exhibit a detectable therapeutic or inhibitory effect. The effect can be detected by any assay method known in the art. The precise effective amount for a subject will depend upon the subject’s body weight, size, and health; the nature and extent of the condition; and the therapeutic or combination of therapeutics selected for administration. Therapeutically effective amounts for a given situation can be determined by routine experimentation that is within the skill and judgment of the clinician.

In some embodiments, the pharmaceutical compositions can be formulated so that a dosage of between 0.01-500 mg/kg body weight/day, for example, 0.05-500 mg/kg body weight/day, 0.1-500 mg/kg body weight/day, 0.1-400 mg/kg body weight/day, 0.1-300 mg/kg body weight/day, 0.1-200 mg/kg body weight/day, 0.1-100 mg/kg body weight/day, 0.1-80 mg/kg body weight/day, 1-100 mg/kg body weight/day or 1-80 mg/kg body weight/day of the compounds of the present disclosure, or a pharmaceutically acceptable salt thereof, can be administered.

In some embodiments, the pharmaceutical compositions comprise one or more compounds of the present disclosure, or a pharmaceutically acceptable salt thereof, as a first active ingredient, and further comprise a second active ingredient. The second active ingredient can be any anti-tumor agent known in the art, for example, antineoplastic agents, antiangiogenic agents, immunotherapy approaches, efficacy enhancers, and the like.

Examples of the antineoplastic agents include, but are not limited to, DNA alkylating agents (for example cisplatin, oxaliplatin, carboplatin, cyclophosphamide, nitrogen mustards like ifosfamide, bendamustine, melphalan, chlorambucil, busulphan, temozolamide and nitrosoureas like carmustine) ; antimetabolites (for example gemcitabine and antifolates such as fluoropyrimidines like 5-fluorouracil and tegafur, raltitrexed, methotrexate, cytosine arabinoside, and hydroxyurea) ; anti-tumor antibiotics (for example anthracyclines like adriamycin, bleomycin, doxorubicin, liposomal doxorubicin, pirarubicin, daunomycin, valrubicin, epirubicin, idarubicin, mitomycin, dactinomycin, amrubicin and mithramycin) ; antimitotic agents (for example vinca alkaloids like vincristine, vinblastine, vindesine and vinorelbine and taxoids like taxol and taxotere and polokinase inhibitors) ; and topoisomerase inhibitors (for example epipodophyllotoxins like etoposide and teniposide, amsacrine, irinotecan, topotecan and camptothecin) ; inhibitors of DNA repair mechanisms such as CHK kinase; DNA-dependent protein kinase inhibitors; inhibitors of poly (ADP-ribose) polymerase (PARP inhibitors, including Olaparib, Rucaparib, Niraparib, Talazoparib, Pamiparib and Fluzoparib) ; and Hsp90 inhibitors such as tanespimycin and retaspimycin, inhibitors of ATR kinase (such as AZD6738) ; and inhibitors of WEE 1 kinase (such as AZD1775/MK-1775) .

Examples of antiangiogenic agents include those that inhibit the effects of vascular endothelial growth factor, such as but not limited to, the anti-vascular endothelial cell growth factor antibody bevacizumab, a VEGF receptor tyrosine kinase inhibitor such as vandetanib (ZD6474) , sorafenib, vatalanib (PTK787) , sunitinib (SU11248) , axitinib (AG-013736) , pazopanib (GW 786034) and cediranib (AZD2171) ; compounds such as those disclosed in International Patent Applications WO 97/22596, WO 97/30035, WO 97/32856 and WO 98/13354; and compounds that work by other mechanisms (for example linomide, inhibitors of integrin ανβ3 function and angiostatin) , or inhibitors of angiopoietins and their receptors (Tie-1 and Tie-2) , inhibitors of PLGF, inhibitors of delta-like ligand (DLL-4) .

Examples of immunotherapy approaches include, but are not limited to, ex-vivo and in-vivo approaches to increase the immunogenicity of patient tumor cells, such as transfection with cytokines such as interleukin 2, interleukin 4 or granulocyte -macrophage colony stimulating factor; approaches to decrease T-cell anergy or regulatory T-cell function; approaches that enhance T-cell responses to tumors, such as blocking antibodies to CTLA4 (for example ipilimumab and tremelimumab) , B7H1, PD-1 (for example BMS-936558 or AMP-514) , PD-L1 (for example MEDI4736) and agonist antibodies to CD 137; approaches using transfected immune cells such as cytokine-transfected dendritic cells; approaches using cytokine-transfected tumor cell lines, approaches using antibodies to tumor associated antigens, and antibodies that deplete target cell types (e.g., unconjugated anti-CD20 antibodies such as Rituximab, radiolabeled anti-CD20 antibodies Bexxar and Zevalin, and anti-CD54 antibody Campath) ; approaches using anti-idiotypic antibodies; approaches that enhance Natural Killer cell function; and approaches that utilize antibody-toxin conjugates (e.g. anti-CD33 antibody Mylotarg) ; immunotoxins such as moxetumumab pasudotox; agonists of toll-like receptor 7 or toll-like receptor 9.

Examples of efficacy enhancers include leucovorin.

Therefore, in some embodiments, there is provided pharmaceutical composition comprising a PROTAC compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable salt thereof, and at least one additional anti-tumor agent. In some embodiments, there is one additional anti-tumor agent. In some embodiments, there are two additional anti-tumor agents. In some embodiments, there are three or more additional anti-tumor agents.

In some embodiments, the amount of additional anti-tumor agent present in the composition of the present disclosure can be no more than the amount that would normally be administered in a composition comprising that anti-tumor agent as the only active agent. In certain embodiments, the amount of the additional anti-tumor agent in the composition of the present disclosure will range from about 50%to 100%of the amount normally present in a composition comprising that anti-tumor agent as the only therapeutically active agent.

Therefore, in another aspect, there is provided a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof in combination with one or more anti-tumor agents listed above.

In some embodiments, the additional anti-tumor agent is selected from the group consisting of doxorubicin, irinotecan, topotecan, etoposide, mitomycin, bendamustine, chlorambucil, cyclophosphamide, ifosfamide, carmustine, melphalan and bleomycin.

As used herein, the term “combination” refers to simultaneous, separate or sequential administration. In some embodiments, “combination” refers to simultaneous administration. In some embodiments, “combination” refers to separate administration. In some embodiments, “combination” refers to sequential administration. Where the administration is sequential or separate, the delay in administering the second component should not be such as to lose the beneficial effect of the combination.

In a further aspect, there is provided a pharmaceutical composition comprising a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof in combination with one or more anti-tumor agents listed above, in association with a pharmaceutically acceptable excipient.

In a further aspect, there is provided a kit comprising a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof in combination with one or more anti-tumor agents listed above.

In a further aspect, there is provided a kit comprising:

(a) a PROTAC compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof in a first unit dosage form;

(b) an anti-tumor agent selected from those listed above in a second unit dosage form; and

(c) container for containing the first and second unit dosage forms.

Uses and Methods for Treatment

The drug moiety of the PROTAC compounds of the present disclosure, i.e., the BCL-XL modulator moiety which is covalently attached to the degradation signaling moiety via the linker, retains essentially the same, similar, or enhanced biological function or activity as compared to the original BCL-XL modulator compound. Also, the degradation signaling moiety of the PROTAC compounds of the present disclosure make it capable of inducing the degradation of BCL-XL proteins.

In some embodiments, there is provided a method of inhibiting the activity of BCL-XL, comprising contacting the BCL-XL with a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof.

In some embodiments, there is provided a method of inducing degradation of BCL-XL, comprising contacting the BCL-XL with a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof.

As a result of their BCL-XL inhibitory and/or degradation activity, the PROTAC compounds, or tautomers, stereoisomers, or pharmaceutically acceptable salts thereof provided herein are useful in therapy, for example in the treatment of diseases, disorders or medical conditions mediated at least in part by BCL-XL, including cancers.

As used herein, the term “cancer” is intended to encompass both non-metastatic cancer and metastatic cancer. In this context, treating cancer involves treatment of both primary tumors and tumor metastases.

As used herein, the term “therapy” is intended to have its normal meaning of dealing with a disease in order to entirely or partially relieve one, some or all of its symptoms, or to correct or compensate for the underlying pathology. The term “therapy” also includes “prophylaxis” unless there are specific indications to the contrary. The terms “therapeutic” and “therapeutically” should be interpreted in a corresponding manner.

As used herein, the term “prophylaxis” is intended to have its normal meaning and includes primary prophylaxis to prevent the development of the disease and secondary prophylaxis whereby the disease has already developed and the patient is temporarily or permanently protected against exacerbation or worsening of the disease or the development of new symptoms associated with the disease.

The term “treatment” , “treat” or “treating” is used synonymously with “therapy” . Similarly the term “treat” can be regarded as “applying therapy” where “therapy” is as defined herein.

Therefore, in one aspect, there is provided a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, for use in therapy.

In some embodiments, there is provided a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, for use as a medicament.

In some embodiments, there is provided a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, for use in the treatment of diseases, disorders or conditions. In some embodiments, the diseases, disorders or conditions are related to an increased level or activity of BCL-XL proteins.

In some embodiments, there is provided a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, for use in the manufacture of a medicament for the treatment of diseases, disorders or conditions. In some embodiments, the diseases, disorders or conditions are related to an increased level or activity of BCL-XL proteins.

In some embodiments, the BCL-XL associated disease, disorder or condition is cancer. In some embodiments, a cancer is a solid tumor or a hematological cancer. In some embodiments, a cancer is a breast cancer, multiple myeloma, plasma cell myeloma, leukemia, lymphoma, gastric cancer, acute myeloid leukemia, bladder cancer, brain cancer, bone marrow cancer, cervical cancer, chronic lymphocytic leukemia, colorectal cancer, esophageal cancer, hepatocellular cancer, lymphoblastic leukemia, follicular lymphoma, lymphoid malignancies of T-cell or B-cell origin, melanoma, myelogenous leukemia, myeloma, oral cancer, ovarian cancer, non-small cell lung cancer, chronic lymphocytic leukemia, prostate cancer, small cell lung cancer, or spleen cancer. In some embodiments, the cancer is selected from the group consisting of leukemia, Hodgkin lymphoma, Non-Hodgkin lymphoma, diffuse large B-cell lymphoma, chronic lymphocytic leukaemia, acute myeloid leukaemia, mantle cell lymphomas, gastro-intestinal cancer, gastric cancer, vascular cancer, biliary carcinomas, pancreatic cancer, colorectal cancer, esophageal cancer, hepatocellular cancer, melanoma, myeloma, oral cancer, ovarian cancer, small cell lung cancer, non-small cell lung cancer, myeloma, prostate cancer, bladder cancer, brain cancer, breast cancer, bone marrow cancer, cervical cancer, spleen cancer, glioblastoma, head and neck squamous cell carcinoma. In some embodiments, the cancer is head and neck squamous cell carcinoma, including but not limited to, lip carcinoma, oral cavity carcinoma, oropharynx carcinoma, hypopharynx carcinoma, glottic larynx carcinoma, supraglottic larynx carcinoma, ethmoid sinus carcinoma, maxillary sinus carcinoma, and occult primary carcinoma. In some embodiments, the cancer is leukemia, including but not limited to, lymphatic leukemia, lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse large B-cell lymphoma, acute myeloid leukemia, lymphoblastic leukemia, follicular lymphoma, lymphoid malignancies of T-cell or B-cell origin, myelogenous leukemia, granulocytic leukemia, polycythemia vera, erythremia. In some embodiments, the cancer is metastatic cancer. In some embodiments, the metastatic cancer comprises metastases of the central nervous system. In some embodiments, the metastases of the central nervous system comprise brain metastases. In some embodiments, the metastases of the central nervous system comprise leptomeningeal metastases. “Leptomeningeal metastases” occur when cancer spreads to the meninges, the layers of tissue that cover the brain and the spinal cord. Metastases can spread to the meninges through the blood or they can travel from brain metastases, carried by the cerebrospinal fluid (CSF) that flows through the meninges.

As used herein, the term “subject in need thereof” is a subject having a BCL-XL associated disease, disorder or condition (e.g., cancer) , or a subject having an increased risk of developing BCL-XL associated disease, disorder or condition (e.g., cancer) relative to the population at large. In some embodiments, a subject is a subject having or suspected of having a cancer. In the case of cancer, a subject in need thereof can have a precancerous condition. A “subject” includes a warm-blooded animal. In some embodiments, the warm-blooded animal is a mammal, e.g. human.

In this context, the term “therapeutically effective amount” refers to an amount of a compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof which is effective to provide “therapy” in a subject, or to “treat” a BCL-XL associated disease, disorder or condition in a subject. In the case of cancer, the therapeutically effective amount may cause any of the changes observable or measurable in a subject as described in the definition of “therapy” , “treatment” and “prophylaxis” above. For example, the effective amount can reduce the number of cancer or tumor cells; reduce the overall tumor size; inhibit or stop tumor cell infiltration into peripheral organs including, for example, the soft tissue and bone; inhibit and stop tumor metastasis; inhibit and stop tumor growth; relieve to some extent one or more of the symptoms associated with the cancer; reduce morbidity and mortality; improve quality of life; or a combination of such effects. An effective amount may be an amount sufficient to decrease the symptoms of a disease responsive to inhibition and/or degradation of BCL-XL. For cancer therapy, efficacy in-vivo can, for example, be measured by assessing the duration of survival, time to disease progression (TTP) , the response rates (RR) , duration of response, and/or quality of life. As recognized by those skilled in the art, effective amounts may vary depending on route of administration, excipient usage, and co-usage with other agents. For example, where a combination therapy is used, the amount of the compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof described in this specification and the amount of the other pharmaceutically active agent (s) are, when combined, jointly effective to treat a targeted disorder in the animal patient. In this context, the combined amounts are in a “therapeutically effective amount” if they are, when combined, sufficient to decrease the symptoms of a disease responsive to inhibition and/or degradation of BCL-XL as described above.

In generally, “therapeutically effective amount” may be determined by one skilled in the art by, for example, starting with the dosage range described in this specification for the compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof and an approved or otherwise published dosage range (s) of the other pharmaceutically active compound (s) .

The method of treating BCL-XL associated diseases, disorders or conditions described herein may be used as a monotherapy. As used herein, the term “monotherapy” refers to the administration of a single active or therapeutic compound to a subject in need thereof. In some embodiments, monotherapy will involve administration of a therapeutically effective amount of one of the compounds of the present disclosure, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, to a subject in need of such treatment.

Depending upon the particular diseases or conditions to be treated, the method of treating BCL-XL associated diseases, disorders or conditions described in this specification may involve, in addition to administration of the compound of the present disclosure, one or more additional therapies, for example, conventional surgery, radiotherapy, chemotherapy, immunotherapy, or a combination of such additional therapies. As used herein, the term “combination therapy” refers to the administration of a combination of multiple active compounds.

The additional therapies, such as additional anti-tumor agents, may be administered separately from the compounds of the present disclosure, as part of a multiple dosage regimen. Alternatively, these additional therapies may be part of a single dosage form, mixed with the compounds of the present disclosure in a single composition.

In some embodiments, the compounds of the present disclosure may be administered simultaneously, sequentially or separately to treatment with the conventional surgery, radiotherapy, chemotherapy or immunotherapy.

Radiotherapy may include one or more of the following categories of therapy: (i) external radiation therapy using electromagnetic radiation, and intraoperative radiation therapy using electromagnetic radiation; (ii) internal radiation therapy or brachytherapy; including interstitial radiation therapy or intraluminal radiation therapy; or (iii) systemic radiation therapy, including but not limited to iodine 131 and strontium 89.

Chemotherapy may include anti-tumor agents known in the art, for example, antineoplastic agents, antiangiogenic agents, efficacy enhancers, and the like described in this specification.

Immunotherapy may include, for example, immune checkpoint modulator. Immune checkpoints are regulators of the immune system, and belong to immunoinhibitory pathway or immunostimulatory pathway, responsible for co-stimulatory or inhibitory interactions of T-cell responses, and regulate and maintain self-tolerance and physiological immune responses. Non-limiting immunoinhibitory checkpoint molecules found in the immunoinhibitory pathways can include LAG3 (CD223) , A2AR, B7-H3 (CD276) , B7-H4 (VTCN1) , BTLA (CD272) , BTLA, CD160, CTLA-4 (CD152) , IDO1, IDO2, TDO, KIR, LAIR-1, NOX2, PD-1, PD-L1, PD-L2, TIM-3, VISTA, SIGLEC-7 (CD328) , TIGIT, PVR (CD155) , TGFβ, or SIGLEC9 (CD329) , among others. Non-limiting immunostimulatory checkpoint molecules found in the immunostimulatory pathways can include CD2, CD3, CD7, CD16, CD27, CD30, CD70, CD83, CD28, CD80 (B7-1) , CD86 (B7-2) , CD40, CD40L (CD154) , CD47, CD122, CD137, CD137L, OX40 (CD134) , OX40L (CD252) , NKG2C, 4-1BB, LIGHT, PVRIG, SLAMF7, HVEM, BAFFR, ICAM-1, 2B4, LFA-1, GITR, ICOS (CD278) , or ICOSLG (CD275) , among others.

Therefore, in one aspect, there is provided a method of treating BCL-XL associated diseases, disorders or conditions in a subject in need thereof, wherein the compound of Formula I, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof is administered simultaneously, separately or sequentially with a second therapy.

In some embodiments, the second therapy is chemotherapy or immunotherapy. In some embodiments, the second therapy is selected from the group consisting of a chemotherapeutic agent, an anti-tumor agent, a radiation therapy agent, an immunotherapy agent, an anti-angiogenesis agent, a targeted therapy agent, a cellular therapy agent, a gene therapy agent, a hormonal therapy agent, an antiviral agent, an antibiotic, an analgesics, an antioxidant, a metal chelator, and cytokines. In some embodiments, the second therapy is a BTK inhibitor, a BCR-ABL inhibitor, a JAK1 inhibitor, a JAK2 inhibitor, a JAK3 inhibitor, a TYK2 inhibitor, a PARP inhibitor, a MEK inhibitor, an ERK inhibitor, or a RAF inhibitor.

EXAMPLES

For the purpose of illustration, the following examples are included. However, it is to be understood that these examples do not limit the invention and are only meant to suggest a method of practicing the present disclosure. Persons skilled in the art will recognize that the chemical reactions described may be readily adapted to prepare a number of other compounds of the present disclosure, and alternative methods for preparing the compounds of the present disclosure are deemed to be within the scope of the present disclosure. For example, the synthesis of non-exemplified compounds according to the present disclosure may be successfully performed by modifications apparent to those skilled in the art, e.g., by appropriately protecting interfering groups, by utilizing other suitable reagents known in the art other than those described, and/or by making routine modifications of reaction conditions. Alternatively, other reactions disclosed herein or known in the art will be recognized as having applicability for preparing other compounds of the present disclosure.

For illustrative purposes, the following shows general synthetic schemes for preparing the compounds of the present disclosure as well as key intermediates. Those skilled in the art will appreciate that other synthetic schemes may be used to synthesize the inventive compounds. Although specific starting materials and reagents are depicted in the General Schemes and discussed below, other starting materials and reagents can be easily substituted to provide a variety of derivatives and/or reaction conditions. In addition, many of the compounds prepared by the methods described below can be further modified in light of this disclosure using conventional chemistry well known to those skilled in the art.

Example 1. Preparation of Drug Moieties

1.1 Synthesis of D-1

Compound D-1-18 is prepared by method of the following scheme, whereinisand R_D2 is

General Procedure

Preparation of Compound D-1-2

A solution of 2M BH₃ ^. Me₂S in THF (206.0 mmol, 1.5 eq) and 1M (R) -3, 3-Diphenyl-1-methylpyrrolidino [1, 2-c] -1, 3, 2-oxazaborole in toluene (274.0 mmol, 2.0 eq) was stirred at 0 ℃ for 1 h. Then D-1-1 (137 mmol, 1.0 eq) in THF (100 mL) was added and the reaction mixture was stirred 0 ℃ for 2 h. Methanol was added to quench the reaction. The solvent was removed under vacuum. H₂O (300 mL) was added and the mixture was extracted with DCM (200 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (EA/PE = 0～30%) to give D-1-2.

Preparation of Compound D-1-3

To a solution of D-1-2 (12.44 mmol, 1.0 eq) in toluene (20 mL) was added DPPA (24.88 mmol, 2 eq) and DBU (18.66 mmol, 1.5 eq) . The reaction mixture was stirred under N2 atmosphere at 50 ℃ for 4 hours. H₂O (50 mL) was added and the mixture was extracted with DCM (30 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (PE = 100%) to give D-1-3.

Preparation of Compound D-1-4

Method Ⅰ: To a solution of D-1-3 (15.79 mmol, 1.0 eq) in THF (50 mL) and H₂O (5 mL) was added PPh₃ (31.58 mmol, 2.0 eq) . The reaction mixture was stirred at 50 ℃ overnight. H₂O (100 mL) was added and the mixture was extracted with DCM (50 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was dissolved in DCM (60 mL) and concentrated HCl (4 mL) was added. The solution was filtrated to give D-1-4.

Method Ⅱ: To a solution of D-1-3 (4.51 mmol, 1.0 eq) in methanol (20 mL) was added NiCl₂ (4.96 mmol, 1.1 eq) and NaBH₄ (6.76 mmol, 1.5 eq) at 0 ℃ and the reaction mixture was stirred at 0 ℃ for 2 h. H₂O (20 mL) was added and the mixture was extracted with EA (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (ME/DCM = 0～10%) to give D-1-4.

Preparation of Compound D-1-6

To a stirred solution of D-1-5 (20.00 g, 108 mmol, 1.0 eq) in DCM (200 mL) was added DMAP (6.60 g, 54.02 mmol, 0.5 eq) , DIPEA (54 mL, 324 mmol, 3.0 eq) and (Boc) ₂O (35.37 g, 162 mmol, 1.5 eq) . The mixture was stirred at rt overnight. It was quenched with aqueous HCl solution (50 mL) , and the aqueous solution was extracted with DCM (50mL X 3) . The combined organic extracts were washed with brine (50 mL) , dried over anhydrous sodium sulfate and concentrated under the reduced pressure. The residue was purified by combine flash (EA/PE = 0～15%) to give D-1-6 (22 g, yield 84%) .

Preparation of Compound D-1-8

To a stirred solution of D-1-6 (21.00 g, 87.06 mmol, 1.0 eq) , D-1-7 (17.52 g, 130.59 mmol, 1.5 eq) in NMP (200 mL) was added Cs₂CO₃ (42.55 g, 130.59 mmol, 1.5 eq) . The mixture was stirred at rt for 15 h under N₂ and LC-MS shows total consumption of D-1-6. The reaction mixture was filtered. The filtrate was diluted with water (200 mL) and was extracted with EA (100mL X 3) . The combined organic extracts were washed with brine (100mL X 7) , dried it over sodium sulfate and concentrated under reduced pressure. The residue was purified by combine flash (EA/PE = 0～30%) to give D-1-8 (21 g, 68%) . MS (ESI) : m/z 356.0 (M+H⁺) .

Preparation of Compound D-1-9

To a stirred solution of D-1-8 (21.00 g, 59.10 mmol, 1.0 eq) in THF (210 mL) was added 60%NaH (3.07 g, 76.83mmol, 1.5 eq) at 0 ℃. The mixture was stirred at 0 ℃ for 1h and then SEMCl (12.81 g, 76.83mmol, 1.5 eq) was added. The mixture was stirred at rt for 2h and LC-MS shows the total consumption of D-1-8. It was quenched by water (20 mL) and extracted with EA (3×100mL) . The combined organic layer was washed brine (1×50mL) , dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by combine flash (EA/PE = 0～30%) to give D-1-9 (25 g, yield 87%) . MS (ESI) : m/z 486.2 (M+H⁺) .

Preparation of Compound D-1-10

To a stirred solution of D-1-9 (23.00 g, 47.36 mmol, 1.0 eq) in Ethanol (200 mL) and saturated NH₄Cl aqueous solution (80 mL) was added Fe (13.22 g, 236.81 mmol, 5.0 eq) . The resulting mixture was stirred at 90 ℃ under N₂ for 3 h. The reaction mixture was filtered and filtrate was diluted with water (200 mL) and extracted with EtOAc (3×100mL) . The combined organic extracts were washed with brine (1×50mL) , dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by combine flash (EA/PE = 0～40%) to give D-1-10 (21 g, yield 97%) . MS (ESI) : m/z 456.2 (M+H⁺) .

Preparation of Compound D-1-11

To a stirred solution of D-1-10 (21 g, 46.09 mmol, 1.0 eq) in DCM (200mL) was added and TEA (25.6 mL, 184.36 mmol, 4.0 eq) and 2-chloroacetyl chloride (10.41 g, 92.18 mmol, 2.0 eq) at 0 ℃. The resulting mixture was stirred at rt for 2 h and LC-MS shows the total consumption of D-1-10. It was quench by aqueous NH₄HCO₃ solution and extracted with DCM (3×100 mL) . The combined organic layer was washed with brine (1×50mL) , dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by flash chromatography (EA/PE =0～60%) to give D-1-11 (22 g, 90%) . MS (ESI) : m/z 532.1 (M+H⁺) .

Preparation of Compound D-1-12

To a solution of D-1-4 (1.13 mmol, 1.0 eq) , D-1-11 (1.13 mmol, 1.0 eq) in acetonitrile (30 mL) was added NaI (3.38 mmol, 3.0 eq) , K₂CO₃ (3.38 mmol, 3.0 eq) . The mixture was heated at 90 ℃ for 5 h. H₂O (40 mL) was added and the mixture was extracted with EA (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (EA/PE = 0～40%) to give D-1-12.

Preparation of Compound D-1-13

To a solution of D-1-12 (1.01 mmol, 1.0 eq) in DCM (15 mL) was added TEA (4.04 mmol, 4.0 eq) and 2-chloroacetyl chloride (2.02 mmol, 2.0 eq) at 0 ℃. The reaction mixture was stirred at rt for 1 h. The organic phase was washed with saturated NaHCO₃ (15 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (EA/PE = 0～40%) to give D-1-13.

Preparation of Compound D-1-14

To a solution of D-1-13 (0.98 mmol, 1.0 eq) in acetonitrile (15 mL) was added NaI (2.95 mmol, 3.0 eq) , K₂CO₃ (2.95 mmol, 3.0 eq) . The reaction mixture was heated at 90 ℃ for 4 h. EA (50 mL) was added and the organic phase was washed with H₂O (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (EA/PE = 0～40%) to give D-1-14.

Preparation of Compound D-1-15

To a solution of D-1-14 (1.0 mmol, 1.0 eq) in THF (10 mL) was added 1M BH₃ ^. THF (10.0 mmol, 10.0 eq) and the mixture was stirred at rt for 2 h. MeOH (30 mL) was added slowly and the mixture was stirred at 60 ℃ overnight. The solvent was removed under vacuum. H₂O (40 mL) was added and the mixture was extracted with EA (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude residue was purified by combine flash (EA/PE = 0～40%) to give D-1-15.

Preparation of Compound D-1-16

To a solution of D-1-15 (0.27 mmol) in DCM (2 mL) was added TFA (2 mL) . The reaction mixture was stirred at rt for 2 h. LC-MS monitored and the starting material was consumed completely. The mixture was concentrated under vacuum. The residue was dissolved in MeOH (8 mL) and K₂CO₃ aqueous solution was added to adjust pH >8. The mixture was stirred at rt overnight. 1M HCl aqueous solution was added to adjust the pH to 5 and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated to give D-1-16.

Preparation of Compound D-1-18

To a solution of D-1-16 (1.0 mmol, 1.0 eq) and D-1-17 (1.0 mmol, 1.0 eq) in DMF (5 mL) was added DIPEA (3.0 mmol, 3.0 eq) , DMAP (3.0 mmol, 3.0 eq) and EDCI (1.3 mmol, 1.3 eq) . The reaction mixture was stirred at rt overnight. The crude mixture was purified by prep-HPLC to give D-1-18.

Drug Moiety D-1 is prepared by the following method.

Preparation of Compound D-1

To a solution of Compound D-1-18 (180 mg, 0.36 mmol) in DCM (10 ml) was added 4M HCL/dioxane (10 ml) and the mixture was stirred for 2h at RT. LCMS showed reaction was completed. Concentrated and residue was purified by prep-HPLC to give Compound D-1 (30 mg, Yield: 18%) . MS (ESI) : m/z 896.2 (M+H⁺) .

1.2 Synthesis of D-2

Compound D-2 is prepared by method of the following scheme, wherein isand R_D2 is

General Procedure

Preparation of Compound D-2-1

To a solution of D-1-15 (10.0 mmol, 1.0 eq) in EtOH (100 mL) was added 2M NaOH aqueous solution (50 mL, 100.0 mmol, 10 eq) . The reaction mixture was stirred at 80 ℃ overnight. Ethanol was removed under vacuum. HCl aqueous solution was added to adjust pH to 6 and it was extracted with DCM (3×100 mL) . The combined organic phase was washed with brine (1×50mL) , dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by flash chromatography (ME/DCM = 0～10%) to give D-2-1.

Preparation of Compound D-2-2

To a solution of D-2-1 (1.0 mmol, 1.0 eq) and D-2-2 (1.0 mmol, 1.0 eq) in DMF (5 mL) was added DIPEA (3.0 mmol, 3.0 eq) , DMAP (3.0 mmol, 3.0 eq) and EDCI (1.3 mmol, 1.3 eq) . The reaction mixture was stirred at rt overnight. EA (50 mL) was added and the mixture was washed with water (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by flash chromatography (ME/DCM = 0～10%) to give D-2-3.

Preparation of Compound D-2

To a solution of D-2-3 (0.27 mmol) in DCM (4 mL) was added TFA (2 mL) . The reaction mixture was stirred at rt for 2 h. LC-MS monitored and the starting material was consumed completely. The mixture was concentrated under vacuum. The residue was dissolved in MeOH (8 mL) and DCM (4 mL) . K₂CO₃ aqueous solution was added to adjust pH >8. The mixture was stirred at RT overnight. 1M HCl aqueous solution was added to adjust the pH to 7 and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated. The crude mixture was purified by Prep-HPLC to give D-2. MS (ESI) : m/z 853.0 (M+H+) .

1.3 1H NMR data of Drug Moieties

D-1

1H NMR (400 MHz, DMSO) δ 11.72 (d, J = 14.8 Hz, 2H) , 10.78 (s, 1H) , 8.66 –8.50 (m, 2H) , 8.43 (s, 2H) , 8.06 (s, 1H) , 7.82 (d, J = 9.2 Hz, 1H) , 7.67 –7.42 (m, 5H) , 7.27 (s, 1H) , 7.10 (d, J = 9.5 Hz, 1H) , 6.76 (d, J = 8.6 Hz, 1H) , 6.40-6.30 (m, 2H) , 4.72 (s, 1H) , 3.50-3.30 (m, 12H) , 3.10-3.00 (m, 6H) , 2.70-2.60 (m, 2H) , 2.19 –1.82 (m, 8H) , 1.81 –1.54 (m, 6H) , 1.40 (s, 2H) .

D-2

1H NMR (500 MHz, DMSO) δ 11.49 (s, 1H) , 8.36 (d, J = 2.0Hz, 1H) , 8.23 (s, 1H) , 7.91 (d, J = 2.5 Hz, 1H) , 7.58 (dd, J = 19.0, 9.0 Hz, 2H) , 7.40 (s, 1H) , 7.26 (dd, J = 21.5, 5.0 Hz, 2H) , 7.15 –7.02 (m, 2H) , 6.71-6.68 (m, 2H) , 6.30 (s, 2H) , 3.46 (s, 2H) , 3.24 (d, J = 6.0 Hz, 2H) , 3.08 (s, 5H) , 2.94 (d, J = 35.0 Hz, 4H) , 2.57 (dd, J = 18.0, 10.0 Hz, 1H) , 2.13 (d, J = 5.5 Hz, 3H) , 1.96 (dd, J = 21.5, 11.5 Hz, 3H) , 1.85 (s, 1H) , 1.68 (d, J = 28.0 Hz, 2H) , 1.62-1.58 (m, 5H) , 1.50 (t, J = 13.0 Hz, 1H) , 1.21-1.18 (m, 3H) .

Example 2. Preparation of PROTAC Compounds

2.1 General Synthetic Scheme 1

General Procedure

Preparation of compound 1-3

To a solution of compound 1-1 (0.46 mmol, 1.0 eq) in MeCN (10 ml) was added HATU (0.47 mmol, 1.02 eq) , DIEA (1.39 mmol, 3.0 eq) and compound 1-2 (0.93mmol, 2.0 eq) . The reaction mixture was stirred at rt for 16 h. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (50 mL) and washed with brine (2 x 20 mL) , dried over Na₂SO₄, concentrated to afford 400 mg of crude product. The crude product was purified by HPLC (H₂O: MeCN=0-50%, at 254nm) to give compound 1-3.

Preparation of compound 1-4

To a solution of compound 1-3 (0.14 mmol, 1.0 eq) in DMF (10 ml) was added HATU (0.21 mmol, 1.5 eq) at rt. Then the reaction mixture was stirred at rt for 0.5 h. DIEA (0.42 mmol, 3.0 eq) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2-aminoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-1) (0.14 mmol, 1.0 eq) was added, the reaction mixture was stirred at rt for 16 h. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (50 mL) and washed with brine (2 x 20 mL) , dried over Na₂SO₄, concentrated to afford crude product. The crude product was purified by HPLC to give compound 1-4.

Preparation of compound P-003:

Preparation of 7- ( ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) amino) -7-oxoheptanoic acid (P-003-2) :

To a solution of (2S, 4R) -1- ( (S) -2-amino-3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (1-1) (200 mg, 0.46 mmol) in MeCN (10 ml) was added HATU (176 mg) , DIEA (180 mg) and heptanedioic acid (P-003-1) (148 mg, 0.93 mmol) . The reaction mixture was stirred at rt for 16 h. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (50 mL) and washed with brine (2 x 20 mL) , dried over Na₂SO₄, concentrated to afford 400 mg of crude product. The crude product was purified by HPLC (H₂O: MeCN=0-50%, at 254nm) to give 7- ( ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) amino) -7-oxoheptanoic acid (P-003-2) . (245 mg, yield 92%) . MS (ESI) : m/z 573.2 (M+H⁺) .

Preparation of N1- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) -N7- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) heptanediamide (P-003) :

To a solution of 7- ( ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) amino) -7-oxoheptanoic acid (P-003-2) (80 mg, 0.14 mmol) in DMF (10 ml) was added HATU (80 mg) at rt. Then the reaction mixture was stirred at rt for 0.5 h. DIEA (54 mg,0.42 mmol) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2-aminoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-1) (126 mg, 0.14 mmol) was added, the reaction mixture was stirred at rt for 16 h. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (50 mL) and washed with brine (2 x 20 mL) , dried over Na₂SO₄, concentrated to afford 150 mg of crude product, The crude product was purified by HPLC to give N1- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) -N7- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl- 1-oxobutan-2-yl) heptanediamide (P-003) as a yellow solid (30 mg, yield 15%) . MS (ESI) : m/z 725.8 (M/2+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 1.

2.2 General Synthetic Scheme 2

General Procedure

Preparation of compound 2-3

To a stirred solution of compound 2-2 (0.80 mmol, 1.0 eq) , 2- (2, 6-dioxopiperidin-3-yl) -4-fluoroisoindoline-1, 3-dione (2-1) (1.20 mmol, 1.5 eq) and DIEA (2.40 mmol, 3.0 eq) in DMSO (2 mL) was stirred at 130 ℃ under microwave for 2 h. The reaction mixture was filtered off. The filtrate was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to give compound 2-3.

Preparation of compound 2-4

To a solution of compound 2-3 (0.11 mmol, 1.0 eq) in DMF (5 mL) was added HATU (0.17 mmol, 1.5 eq) at rt. Then the reaction mixture was stirred at rt for 0.5 h. DIEA (0.33 mmol, 3.0 eq) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2-aminoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3- nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-1) (60 mg, 0.13mmol) was added and the reaction mixture was stirred at rt for 16 h. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (50 mL) and washed with brine (2 x 20 mL) , dried over Na₂SO₄, concentrated to afford 150 mg of crude product. The crude product was purified by HPLC to give compound 2-4.

Preparation of compound P-008:

Preparation of 10- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) decanoic acid (P-008-2) :

To a stirred solution of 10-aminodecanoic acid (P-008-1) (150 mg, 0.80 mmol) , 2- (2, 6-dioxopiperidin-3-yl) -4-fluoroisoindoline-1, 3-dione (2-1) (330 mg, 1.20 mmol) and DIEA (312 mg, 2.40 mmol) in DMSO (2 mL) was stirred at 130 ℃under microwave for 2 h. The reaction mixture was filtered off. The filtrate was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to give 10- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3- dioxoisoindolin-4-yl) amino) decanoic acid (P-008-2) as yellow oil (200 mg, yield 56%) . MS (ESI) : m/z 444.2 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (10- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) decanamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-008) :

To a solution of 10- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) decanoic acid (P-008-2) (100 mg, 0.11 mmol) in DMF (5 mL) was added HATU (64 mg, 0.17 mmol) at rt. Then the reaction mixture was stirred at rt for 0.5 h. DIEA (43 mg, 0.33 mmol) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2-aminoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-1) (60 mg, 0.13mmol) was added and the reaction mixture was stirred at rt for 16 h. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (50 mL) and washed with brine (2 x 20 mL) , dried over Na₂SO₄, concentrated to afford 150 mg of crude product. The crude product was purified by HPLC to give 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (10- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) decanamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-008) as a yellow solid (15 mg, yield 10%) . MS (ESI) : m/z 1321.3 (M+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 2.

2.3 General Synthetic Scheme 3

General Procedure

Preparation of compound 3-2

To a solution of compound 3-1 (0.41 mmol, 1.0 eq) in DCM (5 mL) was added TEA (0.82 mmol, 2.0 eq) and MsCl (0.61 mmol, 1.5 eq) at 0 ℃. The reaction mixture was stirred at RT for 1 h. DCM (20 mL) was added and the organic phase was washed with saturated H₂O (10 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum to give compound 3-2. The crude product was used in the next step directly.

Preparation of compound 3-4

To a solution of compound 3-2 (0.41 mmol, 1.0 eq) and compound 3-3 (1.23 mmol, 3.0 eq) in DMF (6 mL) was added K₂CO₃ (1.02 mmol, 2.5 eq) . The reaction mixture was stirred at 80 ℃ overnight. LC-MS monitored and there was desired product produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give compound 3-4.

Preparation of compound 3-5

To a solution of compound 3-4 (0.21 mmol) in DCM (2 mL) was added TFA (2 mL) . The reaction mixture was stirred at RT for 1 h. LC-MS monitored and there was desired product produced. The solvent was removed under vacuum. The residue was purified by Prep-HPLC to compound 3-5.

Preparation of compound 3-6

To a solution of compound 3-5 (0.17 mmol, 1.0 eq) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2-aminoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-1) (0.20 mmol, 1.2 eq) in DMF (5 mL) was added TEA (0.85 mmol, 5.0 eq) and HATU (0.25 mmol, 1.5 eq) . The reaction mixture was stirred at RT overnight. The mixture was purified by Prep-HPLC to give compound 3-6.

Preparation of compound P-005:

Preparation of tert-butyl 2- (2- (2- ( (methylsulfonyl) oxy) ethoxy) ethoxy) acetate (P-005-2) :

To a solution of tert-butyl 2- (2- (2-hydroxyethoxy) ethoxy) acetate (P-005-1) (90 mg, 0.41 mmol) in DCM (5 mL) was added TEA (83 mg, 0.82 mmol) and MsCl (70 mg, 0.61 mmol) at 0 ℃. The reaction mixture was stirred at RT for 1 h. DCM (20 mL) was added and the organic phase was washed with saturated H2O (10 mL x 1) , brine, dried over anhydrous Na2SO4 and concentrated under vacuum to give tert-butyl 2- (2- (2- ( (methylsulfonyl) oxy) ethoxy) ethoxy) acetate (P-005-2) (122 mg, yield 100%) . The crude product was used in the next step directly. MS (ESI) : m/z 321.0 (M+Na+) .

Preparation of tert-butyl 2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetate (P-005-3) :

To a solution of tert-butyl 2- (2- (2- ( (methylsulfonyl) oxy) ethoxy) ethoxy) acetate (P-005-2) (122 mg, 0.41 mmol) and 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (3-3) (336 mg, 1.23 mmol) in DMF (6 mL) was added K₂CO₃ (141 mg, 1.02 mmol) . The reaction mixture was stirred at 80 ℃ overnight. LC-MS monitored and there was desired product produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give tert-butyl 2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetate (P-005-3) (100 mg, yield 51%) . MS (ESI) : m/z 499.0 (M+Na⁺) .

Preparation of 2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetic acid (P-005-4) :

To a solution of tert-butyl 2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetate (P-005-3) (100 mg, 0.21 mmol) in DCM (2 mL) was added TFA (2 mL) . The reaction mixture was stirred at RT for 1 h. LC-MS monitored and there was desired product produced. The solvent was removed under vacuum. The residue was purified by Prep-HPLC to give 2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetic acid (P-005-4) (70 mg, yield 79%) . MS (ESI) : m/z 421.1 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-005) :

To a solution of 2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetic acid (P-005-4) (70 mg, 0.17 mmol) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2-aminoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-1) (179 mg, 0.20 mmol) in DMF (5 mL) was added TEA (0.12 mL, 0.83 mmol) and HATU (95 mg, 0.25 mmol) . The reaction mixture was stirred at RT overnight. The mixture was purified by Prep-HPLC to give 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (2- (2- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethoxy) ethoxy) acetamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-005) (42.3 mg, yield 20%) . MS (ESI) : m/z 649.8 (M/2+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 3.

2.4 General Synthetic Scheme 4

General Procedure

Preparation of compound 4-2

To a solution of compound 4-1 (2.13 mmol, 1.0 eq) in DCM (100 mL) was added Dess-Martin (4.26 mmol, 2.0 eq) . The mixture was stirred at RT for 5 h. LC-MS monitored. The desired product was produced. DCM (100 mL) was added and the mixture was washed with Na₂S₂O₃ aqueous solution (100 mL x 1) , Na₂CO₃ aqueous solution (100 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give compound 4-2.

Preparation of compound 4-3

To a solution of compound 4-2 (1.29 mmol, 1.0 eq) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (0.70 mmol, 0.54 eq) in NMP (15 ml) was added two drops AcOH and NaBH (AcO) ₃ (3.88 mmol, 3.0 eq) . The reaction mixture was stirred at RT overnight. LCMS showed the reaction was completed. EA (100 mL) was added and the mixture was washed with H₂O (50 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give compound 4-3.

Preparation of compound 4-4

To a solution of compound 4-3 (0.37 mmol) in ethanol (10 ml) was added 1M NaOH aqueous solution (5 mL) . The reaction mixture was stirred at RT overnight. LCMS showed the reaction was completed. DCM (100 mL) was added and the mixture was washed with H₂O (30 mL x 3) , concentrated to give compound 4-4.

Preparation of compound 4-5

To a solution of compound 4-4 (0.12 mmol, 1.0 eq) and (2S, 4R) -1- ( (S) -2-amino-3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (1-1) (0.12 mmol, 1.0 eq) in DCM (5 mL) was added HATU (0.18 mmol, 1.5 eq) and DIPEA (0.60 mmol, 5.0 eq) . The reaction mixture was stirred at RT overnight. LCMS showed the reaction was completed. The solvent was removed under vacuum and the residue was purified by Prep-HPLC to give compound 4-5.

Preparation of compound P-011:

Preparation of tert-butyl 3- (2- (2-oxoethoxy) ethoxy) propanoate (P-011-2) :

To a solution of tert-butyl 3- (2- (2-hydroxyethoxy) ethoxy) propanoate (P-011-1) (500 mg, 2.13 mmol) in DCM (100 mL) was added Dess-Martin (1.8 g, 4.26 mmol) . The mixture was stirred at RT for 5 h. LC-MS monitored. The desired product was produced. DCM (100 mL) was added and the mixture was washed with Na₂S₂O₃ aqueous solution (100 mL x 1) , Na₂CO₃ aqueous solution (100 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give tert-butyl 3- (2- (2-oxoethoxy) ethoxy) propanoate (P-011-2) (300 mg, yield 61%) . MS (ESI) : m/z 233.2 (M+H⁺) .

Preparation of tert-butyl (R) -3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanoate (P-011-3) :

To a solution of tert-butyl 3- (2- (2-oxoethoxy) ethoxy) propanoate (P-011-2) (300 mg, 1.29 mmol) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (600 mg, 0.70 mmol) in NMP (15 ml) was added two drops AcOH and NaBH (AcO) ₃ (822 mg, 3.88 mmol) . The reaction mixture was stirred at RT overnight. LCMS showed the reaction was completed. EA (100 mL) was added and the mixture was washed with H₂O (50 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give tert-butyl (R) -3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanoate (P-011-3) (420 mg, yield 56%) . MS (ESI) : m/z 1069.4 (M+H⁺) .

Preparation of (R) -3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanoic acid (P-011-4) :

To a solution of tert-butyl (R) -3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanoate (P-011-3) (400 mg, 0.37 mmol) in ethanol (10 ml) was added 1M NaOH aqueous solution (5 mL, 5.0 mmol) . The reaction mixture was stirred at RT overnight. LCMS showed the reaction was completed. DCM (100 mL) was added and the mixture was washed with H₂O (30 mL x 3) and concentrated to give (R) -3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7- yl) ethoxy) ethoxy) propanoic acid (P-011-4) (350 mg, yield 92%) . The crude product was used in the next step directly. MS (ESI) : m/z 1013.3 (M+H⁺) .

Preparation of (2S, 4R) -1- ( (S) -2- (3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (P-011) :

To a solution of (R) -3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanoic acid (P-011-4) (120 mg, 0.12 mmol) and (2S, 4R) -1- ( (S) -2-amino-3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (1-1) (51 mg, 0.12 mmol) in DCM (5 mL) was added HATU (68 mg, 0.18 mmol) and DIPEA (77 mg, 0.60 mmol) . The reaction mixture was stirred at RT overnight. LCMS showed the reaction was completed. The solvent was removed under vacuum and the residue was purified by Prep-HPLC to give (2S, 4R) -1- ( (S) -2- (3- (2- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethoxy) ethoxy) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (P-011) (6.8 mg, yield 4%) . MS (ESI) : m/z 713.4 (M/2+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 4.

2.5 General Synthetic Scheme 5

General Procedure

Preparation of compound 5-2

To a solution of compound 5-1 (3.16 mmol, 1.0 eq) in DCM (20 mL) was added pyridine (12.63 mmol, 4.0 eq) and TsCl (4.74 mmol, 1.5 eq) . The reaction mixture was stirred at RT overnight. The solvent was removed under vacuum. Methanol (15 mL) was added and a lot of white solid generated. Filtrate and the filtrate were removed under vacuum. The crude residue was purified by combine flash (CH₃CN/H₂O = 0～50%) to give compound 5-2.

Preparation of compound 5-4

To a solution of compound 5-2 (0.63 mmol, 1.0 eq) , compound 5-3 (1.90 mmol, 3.0 eq) in DMF (5 mL) was added K₂CO₃ (1.58 mmol, 2.5 eq) . The reaction mixture was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and there was desired product produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give compound 5-4.

Preparation of compound 5-5

To a solution of compound 5-4 (0.26 mmol, 1.0 eq) in DCM (5 mL) and DMF (1 mL) was added pyridine (1.29 mmol, 5.0 eq) and TsCl (0.52 mmol, 2.0 eq) . The reaction mixture was stirred at RT overnight. LC-MS monitored and the desired product was produced. H₂O (30 mL) was added and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude was purified by combine flash (ME/DCM = 0 ～ 10%) to give compound 5-5.

Preparation of compound 5-6

To a solution of compound 5-5 (0.13 mmol, 1.0 eq) , (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (0.13 mmol, 1.0 eq) , K₂CO₃ (0.26 mmol, 2.0 eq) and NaI (0.26 mmol, 2.0 eq) in NMP (5 mL) was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and the desired product was produced. H₂O (20 mL) was added and the mixture was extracted with DCM (20 mL x 2) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude was purified by Prep-TLC (ME/DCM = 1/20) or Prep-HPLC to give compound 5-6.

Preparation of compound P-022:

Preparation of ( (1S, 3R) -3- (hydroxymethyl) cyclopentyl) methyl 4-methylbenzenesulfonate (P-022-2) :

To a solution of ( (1R, 3S) -cyclopentane-1, 3-diyl) dimethanol P-022-1) (411 mg, 3.16 mmol) in DCM (20 mL) was added pyridine (1.0 mL, 12.63 mmol) and TsCl (903 mg, 4.74 mmol) . The reaction mixture was stirred at RT overnight. The solvent was removed under vacuum. Methanol (15 mL) was added and a lot of white solid generated. Filtrate and the filtrate were removed under vacuum. The crude residue was purified by combine flash (CH₃CN/H₂O = 0～50%) to give ( (1S, 3R) -3- (hydroxymethyl) cyclopentyl) methyl 4-methylbenzenesulfonate (P-022-2) (430 mg, yield 48%) . MS (ESI) : m/z 307.1 (M+Na⁺) .

Preparation of 2- (2, 6-dioxopiperidin-3-yl) -4- ( ( (1S, 3R) -3- (hydroxymethyl) cyclopentyl) methoxy) isoindoline-1, 3-dione (P-022-3) :

To a solution of ( (1S, 3R) -3- (hydroxymethyl) cyclopentyl) methyl 4-methylbenzenesulfonate (P-022-2) (180 mg, 0.63 mmol) , 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (3-3) (521 mg, 1.90 mmol) in DMF (5 mL) was added K₂CO₃ (219 mg, 1.58 mmol) . The reaction mixture was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and there was desired product produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give 2- (2, 6-dioxopiperidin-3-yl) -4- ( ( (1S, 3R) -3- (hydroxymethyl) cyclopentyl) methoxy) isoindoline-1, 3-dione (P-022-3) (100 mg, yield 41%) . MS (ESI) : m/z 387.0 (M+H⁺) .

Preparation of ( (1R, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclopentyl) methyl 4-methylbenzenesulfonate (P-022-4) :

To a solution of 2- (2, 6-dioxopiperidin-3-yl) -4- ( ( (1S, 3R) -3- (hydroxymethyl) cyclopentyl) methoxy) isoindoline-1, 3-dione (P-022-3) (100 mg, 0.26 mmol) in DCM (5 mL) and DMF (1 mL) was added pyridine (102 mg, 1.29 mmol) and TsCl (99 mg, 0.52 mmol) . The reaction mixture was stirred at RT overnight. LC-MS monitored and the desired product was produced. H₂O (30 mL) was added and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude was purified by combine flash (ME/DCM = 0 ～ 10%) to give ( (1R, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclopentyl) methyl 4-methylbenzenesulfonate (P-022-4) (70 mg, yield 50%) . MS (ESI) : m/z 541.2 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( ( (1R, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclopentyl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-022) :

To a solution of ( (1R, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclopentyl) methyl 4-methylbenzenesulfonate (P-022-4) (70 mg, 0.13 mmol) , (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (110 mg, 0.13 mmol) , K₂CO₃ (36 mg, 0.26 mmol) and NaI (39 mg, 0.26 mmol) in NMP (5 mL) was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and the desired product was produced. H₂O (20 mL) was added and the mixture was extracted with DCM (20 mL x 2) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude was purified by Prep-TLC (ME/DCM = 1/20) to give 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( ( (1R, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclopentyl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-022) (9.9 mg, yield 6%) . MS (ESI) : m/z 611.3 (M/2+H⁺) .

2.6 General Synthetic Scheme 6

General Procedure

Preparation of compound 6-2

To a solution of compound 6-1 (0.31 mmol, 1.0 eq) and 2- (2, 6-dioxopiperidin-3-yl) -4-fluoroisoindoline-1, 3-dione (2-1) (0.38 mmol, 1.2 eq) in DMSO (1 mL) was added DIPEA (0.93 mmol, 3.0 eq) . The reaction mixture was stirred at RT overnight. LC-MS monitored and there was desired product produced. The mixture was purified by Prep-HPLC to give compound 6-2.

Preparation of compound 6-3

To a solution of compound 6-2 (0.22 mmol, 1.0 eq) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (0.22 mmol, 1.0 eq) in NMP (5 mL) was added DIPEA (1.10 mmol, 5.0 eq) and HATU (0.33 mmol, 1.5 eq) . The reaction mixture was stirred at RT overnight. LC-MS monitored and there was desired product produced. The mixture was purified by combine flash (H₂O (0.1%TFA) /CH₃CN = 0～50%) or Prep-HPLC to give compound 6-3.

Preparation of compound P-028:

Preparation of (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) methyl) cyclohexane-1-carboxylic acid (P-028-2) :

To a solution of (1s, 4s) -4- (aminomethyl) cyclohexane-1-carboxylic acid (P-028-1) (50 mg, 0.31 mmol) and 2- (2, 6-dioxopiperidin-3-yl) -4-fluoroisoindoline-1, 3-dione (2-1) (105 mg, 0.38 mmol) in DMSO (1 mL) was added DIPEA (0.16 mL, 0.95 mmol) . The reaction mixture was stirred at RT overnight. LC-MS monitored and there was desired product produced. The mixture was purified by Prep-HPLC to give (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) methyl) cyclohexane-1-carboxylic acid (P-028-2) (90 mg, yield 68%) . MS (ESI) : m/z 414.1 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) methyl) cyclohexane-1-carbonyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-028) :

To a solution of (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) methyl) cyclohexane-1-carboxylic acid (P-028-2) (90 mg, 0.22 mmol) and (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (186 mg, 0.22 mmol) in NMP (5 mL) was added DIPEA (0.18 mL, 1.09 mmol) and HATU (124 mg, 0.33 mmol) . The reaction mixture was stirred at RT overnight. LC-MS monitored and there was desired product produced. The mixture was purified by combine flash (H₂O (0.1%TFA) /CH₃CN = 0～50%) to give 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) methyl) cyclohexane-1-carbonyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-028) (256.2 mg, yield 94%) . MS (ESI) : m/z 624.7 (M/2+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 6.

2.7 General Synthetic Scheme 7

General Procedure

Preparation of compound 7-2

To a solution of compound 7-1 (1.26 mmol, 1.0 eq) in DCM (20 mL) was added Pyridine (3.79 mmol, 3.0 eq) and TsCl (1.90 mmol, 1.5 eq) at RT. Then the mixture was stirred at RT for 16 h. DCM (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated. The crude product was purified by HPLC to give compound 7-2.

Preparation of compound 7-3

To a solution of compound 7-2 (0.32 mmol, 1.0 eq) in DMF (10 mL) was added (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (0.32 mmol, 1.0 eq) , HATU (0.48 mmol, 1.5 eq) and DIEA (0.96 mmol, 3.0 eq) at RT. Then the mixture was stirred at RT for 16 h. EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by TLC (DCM: MeOH=50: 1, at 254nm) to give compound 7-3.

Preparation of compound 7-4

To a solution of compound 7-3 (0.31 mmol, 1.0 eq) in DMF (20 mL) was added 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (3-3) (0.31 mmol, 1.0 eq) , NaI (0.35 mmol, 1.1 eq) and K₂CO₃ (0.47 mmol, 1.5 eq) at RT. Then the mixture was stirred at 80 ℃ for 6 h. EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by HPLC to give compound 7-4.

Preparation of compound P-029:

Preparation of (1s, 4s) -4- ( (tosyloxy) methyl) cyclohexane-1-carboxylic acid (P-029-2) :

To a solution of (1s, 4s) -4- (hydroxymethyl) cyclohexane-1-carboxylic acid (P-029-1) (200 mg, 1.26 mmol) in DCM (20 mL) was added Pyridine (300 mg, 3.79 mmol) and TsCl (362 mg, 1.90 mmol) at RT. Then the mixture was stirred at RT for 16 h. DCM (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated. The crude product was purified by HPLC to give (1s, 4s) -4- ( (tosyloxy) methyl) cyclohexane-1-carboxylic acid (P-029-2) (190 mg, yield 48%) as a white solid. MS (ESI) : m/z 313.1 (M+H⁺) .

Preparation of ( (1s, 4s) -4- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonane-7-carbonyl) cyclohexyl) methyl 4-methylbenzenesulfonate (P-029-3) :

To a solution of (1s, 4s) -4- ( (tosyloxy) methyl) cyclohexane-1-carboxylic acid (P-029-2) (100 mg, 0.32 mmol) in DMF (10 mL) was added (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (273 mg, 0.32 mmol) , HATU (182 mg, 0.48 mmol) and DIEA (124 mg, 0.96 mmol) at RT. Then the mixture was stirred at RT for 16 h. EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by TLC (DCM: MeOH=50: 1, at 254nm) to give ( (1s, 4s) -4- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonane-7-carbonyl) cyclohexyl) methyl 4-methylbenzenesulfonate (P-029-3) (360 mg, yield 98%) as a yellow solid. MS (ESI) : m/z 1147.3 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclohexane-1-carbonyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-029) :

To a solution of ( (1s, 4s) -4- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonane-7-carbonyl) cyclohexyl) methyl 4-methylbenzenesulfonate (P-029-3) (360 mg, 0.31 mmol) in DMF (20 mL) was added 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (3-3) (86 mg, 0.31 mmol) , NaI (52 mg, 0.35 mmol) and K₂CO₃ (65 mg, 0.47 mmol) at RT. Then the mixture was stirred at 80 ℃ for 6 h. EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by HPLC to give 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (1s, 4s) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclohexane-1-carbonyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-029) (22 mg, yield 6%) as a yellow solid. MS (ESI) : m/z 625.2 (M/2+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 7.

2.8 General Synthetic Scheme 8

General Procedure

Preparation of compound 8-2

To a solution of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (0.12 mmol, 1.0 eq) and compound 8-1 (0.12 mmol, 1.0 eq) in NMP (5 mL) was added NaBH (AcO) ₃ (0.24 mmol, 2.0 eq) . The mixture was stirred at RT overnight. LC-MS monitored and desired product produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 4) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give compound 8-2.

Preparation of compound 8-3

To a solution of compound 8-2 (0.085 mmol) in DCM (6 mL) was added TFA (2 mL) . The mixture was stirred at RT for 2 h. LC-MS monitored and desired product produced. The solvent was removed under vacuum to give compound 8-3.

Preparation of compound 8-5

To a solution of compound 8-3 (0.083 mmol, 1.0 eq) and 2- (2, 6-dioxopiperidin-3-yl) -5-fluoroisoindoline-1, 3-dione (8-4) (0.083 mmol, 1.0 eq) in DMSO (4 mL) was added DIPEA (0.25 mmol, 3.0 eq) . The mixture was stirred at 130 ℃ under microwave for 2 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give compound 8-5.

Preparation of compound P-034:

Preparation of tert-butyl (R) -4- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) piperidine-1-carboxylate (P-034-2) :

To a solution of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (100 mg, 0.12 mmol) and tert-butyl 4- (2-oxoethyl) piperidine-1-carboxylate (P-034-1) (27 mg, 0.12 mmol) in NMP (5 mL) was added NaBH (AcO) ₃ (50 mg, 0.24 mmol) . The mixture was stirred at RT overnight. LC-MS monitored and desired product produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 4) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give tert-butyl (R) -4- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) piperidine-1-carboxylate (P-034-2) (90 mg, yield 72%) . MS (ESI) : m/z 1064.5 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (2- (piperidin-4-yl) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-034-3) :

To a solution of tert-butyl (R) -4- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) piperidine-1-carboxylate (P-034-2) (90 mg, 0.085 mmol) in DCM (6 mL) was added TFA (2 mL) . The mixture was stirred at RT for 2 h. LC-MS monitored and desired product produced. The solvent was removed under vacuum to give (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (2- (piperidin-4-yl) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-034-3) (82 mg, yield 100%) . The crude product was used in the next step directly. MS (ESI) : m/z 964.3 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (1- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) piperidin-4-yl) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-034) :

To a solution of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (2- (piperidin-4-yl) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-034-3) (80 mg, 0.083 mmol) and 2- (2, 6-dioxopiperidin-3-yl) -5-fluoroisoindoline-1, 3-dione (8-4) (23 mg, 0.083 mmol) in DMSO (4 mL) was added DIPEA (32 mg, 0.25 mmol) . The mixture was stirred at 130 ℃ under microwave for 2 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (1- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) piperidin-4-yl) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-034) (18.7 mg, yield 18%) . MS (ESI) : m/z 610.7 (M/2+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 8.

2.9 General Synthetic Scheme 9

General Procedure

Preparation of compound 9-2

To a stirred solution of compound 9-1 (0.93 mmol, 1eq) , Dess-Martin (1.41 mmol, 1.5eq) in DCM (20 mL) was stirred at RT for 16 h. The reaction mixture was added NaHCO₃. The mixture was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated to give P-010-2 as yellow oil.

Preparation of compound 9-3

To a solution of compound 9-2 (0.93 mmol, 1eq) in THF/MeOH (3/3 ml) was added 2-aminoethan-1-ol (1.86 mmol, 2 eq) , AcOH (0.96 mmol, 1 eq) and NaBH₃CN (1.4 mmol, 1.5 eq) at RT. Then the reaction mixture was stirred at RT for 4 h. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford crude product compound 9-3.

Preparation of compound 9-4

A solution of compound 9-3 (0.93 mmol, 1eq) in DCM (20 ml) was added (Boc) ₂O (1.40 mmol, 1.5 eq) , DIEA (1.86 mmol, 2eq) at RT. Then the reaction mixture was stirred at RT for 4 h. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford crude compound 9-4.

Preparation of compound 9-5

To a stirred solution of compound 9-4 (0.93 mmol, 1 eq) , Dess-martin (1.41 mmol, 1.5 eq) in DCM (20 mL) was stirred at RT for 16 h. The reaction mixture was added NaHCO₃. The mixture was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated to give compound 9-5.

Preparation of compound 9-6

To a solution of compound 9-5 (0.54 mmol, 1 eq) in NMP (10 ml) was added (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (0.29 mmol, 0.5 eq) , AcOH (0.16 mmol, 0.3 eq) and NaBH (AcO) ₃ (0.49 mmol, 0.9 eq) at RT. Then the reaction mixture was stirred at RT for 4 h. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford 350 mg of crude product. The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give compound 9-6.

Preparation of compound 9-7

To a stirred solution of compound 9-6 (0.19 mmol, 1 eq) , NaOH (5 ml) in EtOH (10 mL) was stirred at rt for 5 h. The reaction mixture was filtered off. The filtrate was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to compound 9-7.

Preparation of compound 9-9

To a stirred solution of compound 9-7 (0.096 mmol, 1eq) , (2S, 4R) -1- ( (S) -2-amino-3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (9-8) (0.106 mmol, 1.1 eq) , DIEA (0.29 mmol, ) and HATU (0.14 mmol, 1.5 eq) in NMP (5 mL) was stirred at RT for 16 h. The reaction mixture was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to give compound 9-9.

Preparation of compound 9-10

To a stirred solution of compound 9-9 (0.018 mmol, 1 eq) , Dioxane/HCl (2 ml) in DCM (5 mL) was stirred at RT for 5 h. The reaction mixture was concentrated and lyophilize to give compound 9-10.

Preparation of compound P-010:

Preparation of tert-butyl 1-oxo-3, 6, 9, 12-tetraoxapentadecan-15-oate (P-010-2) :

To a stirred solution of tert-butyl 1-hydroxy-3, 6, 9, 12-tetraoxapentadecan-15-oate (P-010-1) (300 mg, 0.93 mmol) , Dess-Martin (600 mg, 1.41 mmol) in DCM (20 mL) was stirred at RT for 16 h. The reaction mixture was added NaHCO₃. The mixture was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated to give tert-butyl 1-oxo-3, 6, 9, 12-tetraoxapentadecan-15-oate (P-010-2) (298 mg, yield 100%) as yellow oil. MS (ESI) : m/z 321.0 (M+H⁺) .

Preparation of tert-butyl 1-hydroxy-6, 9, 12, 15-tetraoxa-3-azaoctadecan-18-oate (P-010-3) :

To a solution of tert-butyl 1-oxo-3, 6, 9, 12-tetraoxapentadecan-15-oate (P-010-2) (298 mg, 0.93 mmol) in THF/MeOH (3/3 ml) was added 2-aminoethan-1-ol (114 mg, 1.86 mmol) , AcOH (56 mg, 0.96 mmol) and NaBH₃CN (88 mg, 1.4 mmol) at RT. Then the reaction mixture was stirred at RT for 4 h. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford crude product tert-butyl 1-hydroxy-6, 9, 12, 15-tetraoxa-3-azaoctadecan-18-oate (P-010-3) (340 mg, yield 100%) .

Preparation of tert-butyl 5- (2-hydroxyethyl) -2, 2-dimethyl-4-oxo-3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oate (P-010-4) :

A solution of tert-butyl 1-hydroxy-6, 9, 12, 15-tetraoxa-3-azaoctadecan-18-oate (P-010-3) (340 mg, 0.93 mmol) in DCM (20 ml) was added (Boc) ₂O (305 mg, 1.40 mmol) , DIEA (240 mg, 1.86 mmol) at RT. Then the reaction mixture was stirred at RT for 4 h. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford crude tert-butyl 5- (2-hydroxyethyl) -2, 2-dimethyl-4-oxo-3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oate (P-010-4) (430 mg, yield 99%) .

Preparation of tert-butyl 2, 2-dimethyl-4-oxo-5- (2-oxoethyl) -3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oate (P-010-5) :

To a stirred solution of tert-butyl 5- (2-hydroxyethyl) -2, 2-dimethyl-4-oxo-3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oate (P-010-4) (430 mg, 0.93 mmol) , Dess-martin (600 mg, 1.41 mmol) in DCM (20 mL) was stirred at RT for 16 h. The reaction mixture was added NaHCO₃. The mixture was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated to give tert-butyl 2, 2-dimethyl-4-oxo-5- (2-oxoethyl) -3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oate (P-010-5) (278 mg, yield 65%) as yellow oil. MS (ESI) : m/z 486.0 (M+Na⁺) .

Preparation of tert-butyl (R) -1- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -6, 9, 12, 15-tetraoxa-3-azaoctadecan-18-oate (P-010-6) :

To a solution of tert-butyl 2, 2-dimethyl-4-oxo-5- (2-oxoethyl) -3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oate (P-010-5) (250 mg, 0.54 mmol) in NMP (10 ml) was added (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (D-2) (250 mg, 0.29 mmol) , AcOH (10 mg, 0.16 mmol) and NaBH (AcO) ₃ (102 mg, 0.49 mmol) at RT. Then the reaction mixture was stirred at RT for 4 h. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford 350 mg of crude product. The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give tert-butyl (R) -1- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -6, 9, 12, 15-tetraoxa-3-azaoctadecan-18-oate (P-010-6) (275 mg, yield 42%) as a yellow solid. MS (ESI) : m/z 1201.0 (M+H⁺) .

Preparation of (R) -5- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) -2, 2-dimethyl-4-oxo-3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oic acid (P-010-7) :

To a stirred solution of tert-butyl (R) -1- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -6, 9, 12, 15-tetraoxa-3-azaoctadecan-18-oate (P-010-6) (250 mg, 0.19 mmol) , NaOH (5 ml) in EtOH (10 mL) was stirred at rt for 5 h. The reaction mixture was filtered off. The filtrate was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to give (R) -5- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) -2, 2-dimethyl-4-oxo-3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oic acid (P-010-7) (190 mg, yield 79%) as yellow oil. MS (ESI) : m/z 1244.5 (M+H⁺) .

Preparation of tert-butyl (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) ( (S) -17- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidine-1-carbonyl) -18, 18-dimethyl-15-oxo-3, 6, 9, 12-tetraoxa-16-azanonadecyl) carbamate (P-009) :

To a stirred solution of (R) -5- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) -2, 2-dimethyl-4-oxo-3, 8, 11, 14, 17-pentaoxa-5-azaicosan-20-oic acid (P-010-7) (120 mg, 0.096 mmol) , (2S, 4R) -1- ( (S) -2-amino-3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (46 mg, 0.106 mmol) , DIEA (37 mg, 0.29 mmol) and HATU (55 mg, 0.14 mmol) in NMP (5 mL) was stirred at RT for 16 h. The reaction mixture was diluted with water (20 mL) and was extracted with EtOAc (10mL X 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to give tert-butyl (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) ( (S) -17- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidine-1-carbonyl) -18, 18-dimethyl-15-oxo-3, 6, 9, 12-tetraoxa-16-azanonadecyl) carbamate (P-009) (5.9 mg, yield 4%) as yellow oil. MS (ESI) : m/z 829.3 (M/2+H⁺) .

Preparation of (2S, 4R) -1- ( (S) -1- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -20- (tert-butyl) -18-oxo-6, 9, 12, 15-tetraoxa-3, 19-diazahenicosan-21-oyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (P-010) :

To a stirred solution of tert-butyl (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) ethyl) ( (S) -17- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidine-1-carbonyl) -18, 18-dimethyl-15-oxo-3, 6, 9, 12-tetraoxa-16-azanonadecyl) carbamate (P-009) (30 mg, 0.018 mmol) , Dioxane/HCl (2 ml) in DCM (5 mL) was stirred at RT for 5 h. The reaction mixture was concentrated to give (2S, 4R) -1- ( (S) -1- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -20- (tert-butyl) -18-oxo-6, 9, 12, 15-tetraoxa-3, 19-diazahenicosan-21-oyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (P-010) (27 mg, yield 94%) as yellow solid. MS (ESI) : m/z 519.8 (M/3+H⁺) .

The following compounds were synthesized by the method of general synthetic scheme 9.

2.10 Synthesis of other PROTAC Compounds

The following compounds were prepared by following procedures.

Preparation of compound P-025:

Preparation of cyclobutane-1, 3-diyldimethanol (P-025-2) :

To a solution of compound P-025-1 (500 mg, 3.47 mmol) in THF (25 mL) was added BH3 (1M in THF) of 10 mL) at 0℃. It was warmed to rt and stirred for 3h. Quenched with methanol (3mL) . Solvent was removed under vacuum. The crude residue was purified by combine flash (CH₃CN/H₂O = 0～50%) to give compound P-025-2 (338 mg, yield 84%) . MS (ESI) : m/z 117.1 (M+H+) .

Preparation of (3- (hydroxymethyl) cyclobutyl) methyl 4-methylbenzenesulfonate (P-025-3) :

To a solution of compound P-025-2 (338 mg, 2.91 mmol) in DCM (20 mL) was added pyridine (0.94 mL, 11.64 mmol) and TsCl (832 mg, 4.36 mmol) . The reaction mixture was stirred at RT overnight. The solvent was removed under vacuum. Methanol (15 mL) was added and a lot of white solid generated. Filtrate and the filtrate was removed under vacuum. The crude residue was purified by combine flash (CH3CN/H2O = 0～50%) to give compound P-025-3 (380 mg, yield 48%) . MS (ESI) : m/z 293.1 (M+Na⁺) .

Preparation of 2- (2, 6-dioxopiperidin-3-yl) -4- ( (3- (hydroxymethyl) cyclobutyl) methoxy) isoindoline-1, 3-dione (P-025-4) :

To a solution of compound P-025-3 (380 mg, 1.41 mmol) , 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (1.16 g, 4.22 mmol) in DMF (20 mL) was added K₂CO₃ (486 mg, 3.51 mmol) . The reaction mixture was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and there was desired product produced. The mixture was filtrated and the solvent was removed under vacuum. The residue was purified by combine flash (CH₃CN/H₂O = 0～60%) to give the compound P-025-4 (530 mg, yield 87%) . MS (ESI) : m/z 373.1 (M+H⁺) .

Preparation of 3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclobutane-1-carbaldehyde (P-025-5) :

To a solution of compound P-025-4 (160 mg, 0.43 mmol) in THF (20 mL) was added Dess-Martin periodinane (365 mg, 0.86 mmol) . The reaction mixture was stirred at RT for 5 h. DCM (50 mL) was added and the organic phase was washed with saturated Na₂SO₃ solution (20 mL x 1) , saturated NaHCO₃ solution (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude was purified by combine flash (ME/DCM = 0 ～ 10%) to give compound P-025-5 (160 mg, yield 100%) . MS (ESI) : m/z 371.1 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclobutyl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-025) :

To a solution of compound P-025-5 (200 mg, 0.54 mmol) and compound D-2 (230 mg, 0.27 mmol) in NMP (5 mL) was added NaBH (AcO) ₃ (343 mg, 1.62 mmol) . The reaction mixture was stirred at RT overnight. LC-MS monitored and there was desired product produced. H₂O (30 mL) was added and the mixture was extracted with DCM (30 mL x 3) . The combined organic phase was removed under vacuum. The residue was purified by Prep-HPLC to give product P-025 (68.8 mg, yield 21%) . MS (ESI) : m/z 604.3 (M/2+H⁺) .

Preparation of compound P-030:

Preparation of ( (1R, 3S) -cyclohexane-1, 3-diyl) dimethanol (P-030-2) :

To a solution of compound P-030-1 (800 mg, 4.65 mmol) in THF (25 mL) was added 15 mL BH₃ (1M in THF) at 0℃. It was warmed to rt and stirred for 3h. Quenched with methanol (3mL) . Solvent was removed under vacuum. The crude residue was purified by combine flash (CH3CN/H2O = 0～50%) to give compound P-030-2 (520 mg, yield 78%) . MS (ESI) : m/z 145.2 (M+H⁺) .

Preparation of ( (1S, 3R) -3- (hydroxymethyl) cyclohexyl) methyl 4-methylbenzenesulfonate (P-030-3) :

To a solution of compound P-030-2 (520 mg, 3.61 mmol) in DCM (50 mL) was added pyridine (0.94 mL, 11.64 mmol) and TsCl (1.4 g, 7.22 mmol) . The reaction mixture was stirred at RT overnight. The solvent was removed under vacuum. Methanol (15 mL) was added and a lot of white solid generated. Filtrate and the filtrate was removed under vacuum. The crude residue was purified by combine flash (CH3CN/H2O = 0～50%) to give compound P-030-3 (550 mg, yield 51%) . MS (ESI) : m/z 299.1 (M+H⁺) .

Preparation of 2- (2, 6-dioxopiperidin-3-yl) -4- ( ( (1S, 3R) -3- (hydroxymethyl) cyclohexyl) methoxy) isoindoline-1, 3-dione (P-030-4) :

To a solution of compound P-030-3 (550 mg, 1.84 mmol) , 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (1.01 g, 3.68 mmol) in DMF (20 mL) was added K₂CO₃ (507 mg, 3.68 mmol) . The reaction mixture was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and there was desired product produced. The mixture was filtrated and the solvent was removed under vacuum. The residue was purified by combine flash (CH₃CN/H₂O = 0～60%) to give the compound P-030-4 (401 mg, yield 54%) . MS (ESI) : m/z 400.1 (M+H⁺) .

Preparation of (1R, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclohexane-1-carbaldehyde (P-030-5) :

To a solution of compound P-030-4 (200 mg, 0.5 mmol) in THF (20 mL) was added Dess-Martin (318 mg, 0.75 mmol) . The reaction mixture was stirred at RT for 5 h. DCM (50 mL) was added and the organic phase was washed with saturated Na₂SO₃ solution (20 mL x 1) , saturated NaHCO₃ solution (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude was purified by combine flash (ME/DCM = 0 ～ 10%) to give compound P-030-5 (265 mg, yield 83%) . MS (ESI) : m/z 399.1 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( ( (1S, 3S) -3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) cyclohexyl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-030) :

To a solution of compound P-030-5 (100 mg, 0.25 mmol) and compound D-2 (100 mg, 0.12 mmol) in NMP (5 mL) was added NaBH (AcO) ₃ (52 mg, 0.25 mmol) and HOAc (1 drops) . The reaction mixture was stirred at RT overnight. LC-MS monitored and there was desired product produced. H₂O (30 mL) was added and the mixture was extracted with DCM (30 mL x 3) . The combined organic phase was removed under vacuum. The residue was purified by Prep-HPLC to give product P-030 (75 mg, yield 24%) . MS (ESI) : m/z 618.3 (M/2+H⁺) .

Preparation of compound P-037:

Preparation of 2- (2, 6-dioxopiperidin-3-yl) -5- ( ( (1r, 4r) -4- (hydroxymethyl) cyclohexyl) methoxy) isoindoline-1, 3-dione (P-037-3) :

A mixture of compound P-037-2 (140 mg, 0.97 mmol) , P-037-1 (200 mg, 0.72 mmol) , PPh3 (210 mg, 0.8 mmol) , DIAD (147 mg, 0.72 mmol) in DMF (5 ml) was stirred at RT for 16h. H2O (20 mL) and DCM (50 mL) was added to the mixture. The organic layer was washed with brine, dried with Na2SO4. It was concentrated and the residue was purified by combine flash (DCM: MEOH=10: 1) to give product P-037-3 (60 mg, yield 21%) . MS (ESI) : m/z 400.1 (M+H⁺) .

Preparation of ( (1r, 4r) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) methyl) cyclohexyl) methyl methanesulfonate (P-037-4) :

A mixture of P-037-3 (60 mg, 0.15) , MsCl (17 mg, 0.15) , TEA (0.1 mL) in DCM (25 ml) was stirred at RT for 16h. H₂O (20 ml) was added to the mixture. The organic layer was separated, dried with Na2SO4, concentrated and the residue was purified by combine flash (DCM: MEOH=20: 1) to give product P-037-4 (60 mg, yield 84%) . MS (ESI) : m/z 479.1 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( ( (1r, 4r) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) methyl) cyclohexyl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-037) :

A mixture of P-037-4 (40 mg, 0.08mmol) , D-2 (70 mg, 0.08 mmol) , DIEA (0.2 mL) in DMSO (2 ml) was heated at 130℃ under MW for 1h. DCM (50 ml) was added to the mixture. The mixture was washed with water (10mL X2) , dried with Na2CO3. The organic layer was concentrated under vacuum and the residue was purified by combine flash (DCM: MEOH=10: 1) to give product P-037 (10 mg, yield 10%) . MS (ESI) : m/z 618.2 (M/2+H⁺) .

Preparation of compound P-042:

Preparation of 3- (4- ( ( (1r, 4r) -4- (hydroxymethyl) cyclohexyl) methoxy) -1-oxoisoindolin-2-yl) piperidine-2, 6-dione (P-042-3) :

To a solution of compound P-042-1 (150 mg, 0.50 mmol) and compound P-042-1 (196 mg, 0.75 mmol) in DMF (15 mL) was added K₂CO₃ (83 mg, 0.60 mmol) . The mixture was stirred at 80 ℃ under microwave for 3 h. LC-MS monitored and desired product was produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give the compound P-042-3 (60 mg, yield 28%) . MS (ESI) : m/z 387.3 (M+H⁺) .

Preparation of ( (1r, 4r) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-4-yl) oxy) methyl) cyclohexyl) methyl 4-methylbenzenesulfonate (P-042-4) :

To a solution of compound P-042-3 (60 mg, 0.15 mmol) in DCM (15 mL) was added pyridine (0.13 mL, 1.55 mmol) and TsCl (44 mg, 0.23 mmol) . The mixture was stirred at RT for 2 days. H₂O (50 mL) was added and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na2SO4 and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give the compound P-042-4 (50 mg, yield 60%) . MS (ESI) : m/z 541.3 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( ( (1r, 4r) -4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-4-yl) oxy) methyl) cyclohexyl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-042) :

To a solution of compound P-042-4 (50 mg, 0.092 mmol) and compound D-2 (79 mg, 0.092 mmol) in NMP (5 mL) was added NaI (28 mg, 0.18 mmol) and K₂CO₃ (26 mg, 0.18 mmol) . The mixture was stirred at 80 ℃ under microwave for 4 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give the product-042 (9.6 mg, yield 9%) . MS (ESI) : m/z 1221.4 (M+H⁺) .

Preparation of compound P-017

Preparation of 2- (4-hydroxybutyl) isoindoline-1, 3-dione (P-017-2) :

A mixture of 4-aminobutan-1-ol (1 g, 11.2 mmol) , P-017-1 (1.7 g, 11.3 mmol) , TEA (0.5 ml) in toluene (30 ml) was heated at reflux for 3h. LCMS showed reaction was complete. EA (50mL) was added and washed with water (20 mLX2) . The organic layer was dried and concentrated, the residue was purified by combine flash (PE: EA=1: 1, normal phase silica, UV254) to give the product P-017-2 (1 g, yield 41%) . MS (ESI) : m/z 220.1 (M+H⁺) .

Preparation of 4- (1, 3-dioxoisoindolin-2-yl) butanal (P-017-3) :

A mixture of compound P-017-2 (300 mg, 1.37 mmol) , Dess-Martin (700 mg, 1.65mmol) in DCM (25 ml) was stirred at RT 3h. LCMS showed reaction was complete. DCM (50mL) was added and washed with water (20 mLX2) . The organic layer was dried and concentrated, the residue was purified by combine flash (PE: EA=1: 2, normal phase silica, UV254) to give the product P-017-3 (250 mg, yield 83%) . MS (ESI) : m/z 218.1 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- (1, 3-dioxoisoindolin-2-yl) butyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-017-4) :

To a solution of P-017-3 (70 mg, 0.32 mmol) and compound D-2 (275 mg, 0.32 mmol) in NMP (5 ml) was added two drops AcOH and NaBH (OAc) ₃ (203 mg, 0.94) . The reaction mixture was stirred at RT overnight. LCMS showed reaction was complete. EA (50mL) was added and washed with water (20 mLX2) . The organic layer was dried and concentrated, the residue was purified by combine flash (PE : EA=1: 1, normal phase silica, UV254) to give the product P-017-4 (150 mg, yield 45%) . MS (ESI) : m/z 1054.3 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (4-aminobutyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-017-5) :

A mixture of compound P-017-4 (50 mg, 0.05 mmol) , NH2NH2. H2O (4.8 mg, 0.1 mmol) in EtOH (10 ml) was heated at 70℃ for 2h. LCMS showed reaction was complete. Concentrated and the residue was purified by combine flash (DCM : MeOH = 1 : 1, normal phase silica, UV254) to give the product P-017-5 (40 mg, yield 87%) . MS (ESI) : m/z 924.3 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) butyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-017) :

A mixture of P-017-5 (40 mg, 0.04 mmol) , compound 2-1 (15 mg, 0.05 mmol) , DIEA (17.5 mg, 0.13 mmol) in DMSO (3 ml) was heated at 130℃ for 1h under MW. LCMS showed reaction was complete. DCM (30mL) was added, washed with water (20 mLX2) . The organic layer was dried and concentrated, the residue was purified by combine flash (DCM : MEOH=10: 1, normal phase silica, UV254) to give the product P-017 (15 mg, yield 32%) . MS (ESI) : m/z 1180.7 (M+H⁺) .

Preparation of compound P-031:

Preparation of 4-fluoro-2- (1-methyl-2, 6-dioxopiperidin-3-yl) isoindoline-1, 3-dione (P-031-1) :

CH₃I (153 mg, 1.08 mmol) was added to a solution of 2-1 (200 mg, 0.72 mmol) , K₂CO₃ (150 mg, 1.08 mmol) in DMF (10 ml) was stirred for 2h.. LC-MS monitored and desired product was produced. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give the compound P031-1 (190 mg, yield 91%) . MS (ESI) : m/z 291.0 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- ( (2- (1-methyl-2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) butyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-031) :

To a solution of compound P-017-5 (150 mg, 0.16 mmol) and compound P-031-1 (50 mg, 0.17 mmol) in DMSO (5 ml) , and was added to DIEA (0.5 ml) . The reaction mixture was stirred at 130℃ 2h under MW. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (DCM/CH₃OH = 10: 1) to give the compound P-031 (20 mg, yield 10.5%) . MS (ESI) : m/z 597.6 (M/2+H⁺) .

Preparation of compound P-032:

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) amino) butyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-032) :

To a solution of compound P-017-5 (150 mg, 0.16 mmol) and compound P-032-2 (45 mg, 0.16 mmol) in DMSO (5 ml) , and was added to DIEA (0.5 ml) . The reaction mixture was stirred at 130℃ 2 h MW. DCM (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (DCM/CH₃OH = 10: 1) to give the compound P-032 (15 mg, yield 7.9%) . MS (ESI) : m/z 590.6 (M/2+H⁺) .

Preparation of compound P-020:

Preparation of 3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) benzaldehyde (P-020-3) :

A mixture of compound P-020-1 (200 mg, 0.98 mmol) , compound P-020-2 (332 mg, 1.20 mmol) , K₂CO₃ (167 mg, 1.20 mmol) in DMF (10 ml) was stirred at RT for 3h. LCMS showed reaction was complete. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (DCM/CH₃OH = 15: 1) to give the compound P-020-3 (150 mg, yield 39%) . MS (ESI) : m/z 393.1 (M/2+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (3- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) benzyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-020) :

To a solution of compound P-020-3 (45 mg, 0.12 mmol) and D-2 (98 mg, 0.12 mmol) in NMP (5 ml) was added two drops AcOH and NaBH (OAc) ₃ (73 mg, 0.35 mmol) . The reaction mixture was stirred at RT overnight. DCM (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by prep-TLC (DCM/CH₃OH = 15: 1) to give the compound P-020 (10 mg, yield 6.7%) . MS (ESI) : m/z 1229.7 (M+H⁺) .

Preparation of compound P-021:

Preparation of 4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) benzaldehyde (P-021-3) :

A mixture of compound P-021-1 (200 mg, 0.98 mmol) , compound P-021-2 (332 mg, 1.20 mmol) , K₂CO₃ (167 mg, 1.20 mmol) in DMF (10 ml) was stirred at RT for 3h. LCMS showed reaction was complete. EA (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (DCM/CH₃OH = 15: 1) to give the compound P-021-3 (160 mg, yield 42%) . MS (ESI) : m/z 393.1 (M/2+H) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- ( ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) methyl) benzyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-021) :

To a solution of compound P-021-3 (150 mg, 0.38 mmol) and D-2 (323 mg, 0.38 mmol) in NMP (5 ml) was added two drops AcOH and NaBH (OAc) ₃ (120 mg, 0.57 mmol) . The reaction mixture was stirred at RT overnight. DCM (100 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by prep-TLC (DCM/CH3OH = 15: 1) to give the compound P-021 (75 mg, yield 16%) . MS (ESI) : m/z 615.3 (M/2+H⁺) .

Preparation of compound P-040:

Preparation of N2-acetyl-N6- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) -L-lysine (P-040-2) :

A mixture of compound P-040-1 (100 mg, 0.53 mmol) , compound 2-1 (147 mg, 0.53 mmol) , DIEA (0.1 mL) in DMSO (5 ml) was stirred at 130℃ for 1h under MW. LCMS showed product was found. The solution was purified by reverse phase combine flash (CH3CN/H2O = 0～50%) to give compound P-040-2 (80 mg, yield 34%) . MS (ESI) : m/z 445.1 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2- ( (2S) -2-acetamido-6- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) hexanamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-040) :

A mixture of P-040-2 (20 mg, 0.104mmol) , D-1 (40 mg, 0.04 mmol) , EDCI (11.4 mg, 0.06mmol) , HOBT (12 mg, 0.08 mmol) in DMF (5 ml) was stirred at RT for 16h. LCMS showed product was found, DCM (50mL) was added and washed with water (50 ml x 2) . Organic layer was dried and concentrated and residue was purified by prep-HPLC to give compound P-040 (20 mg , Yueld: 38%) as white solid. MS (ESI) : m/z 662.0 (M/2+H⁺) .

Preparation of compound P-043:

Preparation of methyl N6-acetyl-L-lysinate (P-043-2) :

To a solution of compound P-043-1 (300 mg, 1.59 mmol) in MeOH (15 mL) was added SO₂Cl₂ (1.5 mL) slowly. The reaction mixture was stirred at 60℃ for 1 h. NaHCO₃ aqueous solution was added to adjust pH to 7. The solvent was removed under vacuum. The residue was washed with ME/DCM (1/10, 100 mL) . Filtrate and the organic phase were removed under vacuum to give the product compound P-043-2 (140 mg, yield 43%) . MS (ESI) : m/z 203.1 (M+H⁺) .

Preparation of methyl N6-acetyl-N2- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) -L-lysinate (P-043-4) :

To a solution of compound P-043-2 (40 mg, 0.20 mmol) and compound P-043-3 (66 mg, 0.24 mmol) in DMSO (2 mL) was added DIPEA (76 mg, 0.59 mmol) . The mixture was stirred at 130 ℃ under microwave for 3 h. LC-MS monitored and desired product was produced. DCM (30 mL) was added and the mixture was washed with H₂O (10 mL x 2) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by Prep-TLC (ME/DCM = 1/10) to give the compound P-043-4 (30 mg, yield 33%) . MS (ESI) : m/z 459.2 (M+H⁺) .

Preparation of N6-acetyl-N2- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) -L-lysine (P-043-5) :

To a solution of compound P-043-4 (10 mg, 0.022 mmol) in dioxane (2 mL) and H₂O (2 mL) was added two drops concentrated HCl. The mixture was stirred at 50 ℃ overnight. LC-MS monitored and desired product was produced. The solvent was removed under vacuum to give the compound P-043-5 (10 mg, yield 100%) . The crude product was used in the next step directly. MS (ESI) : m/z 445.1 (M+H⁺) .

2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2- ( (2S) -6-acetamido-2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) amino) hexanamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-043) :

To a solution of compound P-043-5 (10 mg, 0.022 mmol) , compound D-1 (20 mg, 0.022 mmol) in DMF (2 mL) was added EDCI (8 mg, 0.045 mmol) and HOBt (9 mg, 0.066 mmol) . The reaction mixture was stirred at RT overnight. The mixture was purified by Prep-HPLC to give the product P-043 (1.9 mg, yield 6%) . MS (ESI) : m/z 662.0 (M+H⁺) .

Preparation of compound P-045:

Preparation of N6- (tert-butoxycarbonyl) -N2- (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-5-yl) -L-lysine (P-045-3) :

To a solution of compound P-045-1 (300 mg, 0.93mmol) in DMSO (10 mL) was added compound P-045-2 (275 mg, 1 mmol) , CuI (106 mg, 0.56 mmol) and K₃PO₄ (591 mg, 2.78 mmol) at rt. Then the mixture was stirred at 120℃ for 2 h, LCMS showed ok, EA (50 mL) was added and the organic phase was washed with H2O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated., The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give the product P-045-3 (350 mg, yield 77%) . MS (ESI) : m/z 489.1 (M+H⁺) .

Preparation of N6- (tert-butoxycarbonyl) -N2- (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-5-yl) -L-lysine (P-045-4) :

To a solution of compound P-045-3 (100 mg, 0.2 mmol) , compound D-1 (179mg, 0.2 mmol) in DMF (5 mL) was added EDCI (57 mg, 0.3 mmol) and HOBt (41 mg, 0.3 mmol) . The reaction mixture was stirred at RT overnight. DCM (30 mL) was added and the mixture was washed with H₂O (10 mL x 2) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by Prep-TLC (ME/DCM = 1/10) to give the compound P-045-4 (90 mg, yield 33%) . MS (ESI) : m/z 683.5 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2- ( (2S) -6-amino-2- ( (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-5-yl) amino) hexanamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-045) :

To a solution of Compound P-045-4 (90 mg, 0.06) in DCM (10 mL) was added TFA (3 ml) at rt, Then the mixture was stirred at rt for 4 h, LCMS showed OK, concentrated , The crude product was purified by HPLC to P-045 (40 mg, yield 45%) . MS (ESI) : m/z 633.5 (M+H⁺) .

Preparation of compound P-046:

Preparation of N6- (tert-butoxycarbonyl) -N2- (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-5-yl) -L-lysine (P-046-2) :

To a stirred solution of compound P-046-1 (500 mg, 2.02 mmol) , compound (bromomethyl) benzene (378 mg, 2.23 mmol) TBAI (74 mg, 0.2 mmol) and KHCO3 (303 mg, 3.03 mmol) in DMSO (20 mL) was stirred at 80C for 6 h. The mixture was diluted with water (50 mL) and was extracted with EtOAc (50mL X 3) . The combined organic extracts were concentrated. The crude product was purified by TLC (DCM: MeOH=0-10%, at 254nm) to give 600 mg of Compound P-046-2 as a yellow oil. LCMS: [M+H] ⁺: 338. Yelid: 88%

Preparation of benzyl (S) -2- ( (tert-butoxycarbonyl) amino) -6- ( (methylsulfonyl) oxy) hexanoate (P-046-3) :

To a solution of A (300 mg, 0.89 mmol) in DCM (30 mL) was added DIEA (270 mg, 2.67 mmol) and MsCl (205 mg, 1.78 mmol) at rt, Then the mixture was stirred at rt for 16 h, DCM (50 mL) was added and the organic phase was washed with H2O (50 mL x 1) , brine, dried over anhydrous Na2SO4 and concentrated , The crude product was purified by TLC (DCM: MeOH=20: 1, at 254nm) to give 360 mg of Compound P-046-3 as a white solid. LCMS: [M+H] ⁺: 416.1. Yelid: 98%

Preparation of benzyl N2- (tert-butoxycarbonyl) -N6- (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-4-yl) -L-lysinate (P-046-4) :

To a solution of P-046-3 (300 mg, 0.72 mmol) in NMP (20 mL) was added DIEA (139 mg, 1.08mmol) , NaI (108 mg, 0.72 mmol) and 3- (4-amino-1-oxoisoindolin-2-yl) piperidine-2, 6-dione (205 mg, 0.79 mmol) at rt, Then the mixture was stirred 120C for 6 h, LCMS showed OK, DCM (50 mL) was added and the organic phase was washed with H2O (50 mL x 1) , brine, dried over anhydrous Na2SO4 and concentrated , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 300 mg of Compound P-046-4 as a white solid. LCMS: [M+H] +: 579.1, Yelid: 72%

Preparation of N2- (tert-butoxycarbonyl) -N6- (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-4-yl) -L-lysine (P-046-5) :

To a solution of P-046-4 (190 mg, 0.33 mmol) in MeOH (20 mL) was added Pd/C (190 mg) at rt, Then the mixture was stirred at rt for 16 h under H2, LCMS showed OK, filleted and concentrated to give 150 mg of P-046-5 as a white solid. LCMS: [M+H] +: 489.1. Yelid: 93%

Preparation of tert-butyl ( (2S) -1- ( (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7- azaspiro [3.5] nonan-7-yl) ethyl) amino) -6- ( (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-4-yl) amino) -1-oxohexan-2-yl) carbamate (P-046-6) :

To a solution of compound P-046-5 (100 mg, 0.20 mmol) , compound D-1 (55 mg, 0.061 mmol) in DMF (5 mL) was added EDCI (78 mg, 0.41 mmol) and HOBt (83 mg, 0.61 mmol) . The reaction mixture was stirred at RT overnight. EA (100 mL) was added and the mixture was washed with H₂O (30 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give the compound P-046-6 (70 mg, yield 25%) . MS (ESI) : m/z 1367.2 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (2- ( (2S) -2-amino-6- ( (2- (2, 6-dioxopiperidin-3-yl) -1-oxoisoindolin-4-yl) amino) hexanamido) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-046) :

To a solution of compound P-046-6 (70 mg, 0.051 mmol) in DCM (3 mL) was added TFA (1 mL) . The reaction mixture was stirred at RT for 2 h. LC-MS monitored and the starting material was consumed completely. The mixture was purified by Prep-HPLC to give the product P-046 (7.4 mg, yield 11%) . MS (ESI) : m/z 633.9 (M/2+H⁺) .

Preparation of compound P-047 and P-049:

Preparation of tert-butyl bis (3-hydroxypropyl) carbamate (P-047-2) :

To a stirred solution of P-047-1 (500 mg, 3.76 mmol) , (Boc) ₂O (1.23 g, 5.64 mmol) and DIEA (970 mg, 7.52 mmol) in DCM (20 mL) was stirred at RT for 16 h. The mixture was diluted with water (50 mL) and was extracted with DCM (50mL X 3) . The combined organic extracts were concentrated. The crude product was purified by TLC (DCM: MeOH=0-10%, at 254nm) to give 600 mg of Compound P-047-2 as a yellow oil. LCMS: [M+H] ⁺: 234. Yelid: 68%

Preparation of tert-butyl (3- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) propyl) (3-hydroxypropyl) carbamate (P-047-3) :

To a solution of P-047-2 (100 mg, 0.43 mmol) in THF (10 mL) was added 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (129 mg, 0.47 mmol) , PPh₃ (123 mg, 0.47 mmol) and DIAD (86 mg 0.43 mmol) at rt, Then the mixture was stirred at rt for 16 h, LCMS showed OK, DCM (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (DCM: MeOH=20: 1, at 254nm) to give 130 mg of Compound P-047-3 as a white solid. LCMS: [M+H] +: 490.1. Yelid: 62%

Preparation of tert-butyl (3- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) propyl) (3-oxopropyl) carbamate (P-047-4) :

To a solution of P-047-3 (100 mg, 0.20 mmol) in DCM (20 mL) was added NaOAc (82 mg, 0.60 mmol) and PCC (129 mg, 0.60 mmol) at rt, Then the mixture was stirred at rt for 4 h, LCMS showed OK, DCM (50 mL) was added and the organic phase was washed with H2O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 90 mg of Compound P-046-4 as a white solid. LCMS: [M+H] +: 488.1, Yelid: 92%

Preparation of tert-butyl (3- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) propyl) (3- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) propyl) carbamate (P-047-5) :

To a solution of P-047-4 (90 mg, 0.18 mmol) in DMF (20 mL) was added D-2 (158 mg, 0.18 mmol) and HOAc (32 mg, 0.54 mmol) at rt, Then the mixture was stirred at rt for 1 h under N₂, NaBH (OAc) ₃ (21 mg, 0.54 mmol) was added, the mixture was stirred at rt for 16 h under N2, LCMS showed OK, EA (50 mL) was added and the organic phase was washed with H2O (50 mL x 1) , brine, dried over anhydrous Na2SO4 and concentrated , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 120 mg of Compound P-047-5 as a white solid. LCMS: [M+H] +: 1324.5, Yelid: 50%

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (3- ( (3- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) propyl) amino) propyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-047) :

To a solution of compound P-047-5 (60 mg, 0.045 mmol) in DCM (4 mL) was added TFA (1 mL) . The reaction mixture was stirred at RT for 1 h. LC-MS monitored and the desired product was produced. The solvent was removed under vacuum. The residue was dissolved in DCM (50 mL) and the organic phase was washed with saturated NaHCO₃ solution (30 mL x 1) . The organic phase was removed under vacuum to give the product compound P-047 (55 mg, yield 100%) . MS (ESI) : m/z 612.8 (M/2+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (3- ( (3- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) propyl) (3-hydroxypropyl) amino) propyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-049) :

To a solution of compound P-047 (55 mg, 0.045 mmol) in DMF (2 mL) was added 3-bromopropan-1-ol (37 mg, 0.27 mmol) and DIPEA (116 mg, 0.90 mmol) . The reaction mixture was stirred at 80 ℃ under microwave for 1.5 h. LC-MS monitored and the desired product was produced. The mixture was purified by Prep-HPLC to give the product P-049 (18.3 mg, yield 32%) . MS (ESI) : m/z 641.8 (M/2+H⁺) .

Preparation of compound P-055:

Preparation of 2-azidoethyl 4-methylbenzenesulfonate (P-055-2) :

To a solution of P-055-1 (200 mg, 2.30 mmol) in DCM (10 mL) was added TEA (464 mg, 4.60mmol) and TsCl (437 mg, 2.30 mmol) at rt, Then the mixture was stirred at rt for 16 h, LCMS showed OK, DCM (30 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (PE: EA=4: 1, at 254nm) to give 280 mg of Compound P-055-2 as a white solid. LCMS: [M+H] +: 242.0. Yelid: 51%

Preparation of 4- (2-azidoethoxy) -2- (2, 6-dioxopiperidin-3-yl) isoindoline-1, 3-dione (P-055-3) :

A mixture of P-052-2 (280 mg, 1.15 mmol) , 2- (2, 6-dioxopiperidin-3-yl) -4-hydroxyisoindoline-1, 3-dione (315 mg, 1.15 mmol) , Na₂CO₃ (159 mg, 1.50 mmol) in DMF (10 mL) was heated at 80℃ for 3h. DCM (100 mL) was added and the organic phase was washed with H₂O (50 mL x 2) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (PE: EA=1: 1, at 254nm) to give 200 mg of Compound P-055-3 as a white solid. LCMS: [M+H] +: 344.1. Yelid: 51%

Preparation of 3-nitro-4- ( ( (7- (prop-2-yn-1-yl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) benzenesulfonamidebenzenesulfonamide (P-055-5) :

A mixture of P-052-4 (150 mg, 0.82 mmol) , 3-bromoprop-1-yne (96 mg, 0.82 mmol) , TEA (163 mg, 1.62 mmol) in DMF (10 mL) was heated at 60℃ for 2h. DCM (100 mL) was added and the organic phase was washed with H2O (50 mL x 2) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 180 mg of Compound P-055-5 as a white solid. LCMS: [M+H] +: 393.1. Yelid: 46%

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (prop-2-yn-1-yl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-055-7) :

To a solution of P-055-6 (200 mg, 0.38 mmol) in DMF (3 mL) was added EDCI (110 mg, 0.57 mmol ) , DMAP (69 mg, 0.57 mmol) and DIEA (142 mg, 1.10 mmol) at rt, folowwed by P-055-5 was added. Then the mixture was stirred at rt for 16 h, LCMS showed OK, DCM (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 190 mg of P-055-7 as a yellow solid. LCMS: [M+H] +: 891.3. Yelid: 56%

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (1- (2- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) ethyl) -1H-1, 2, 3-triazol-4-yl) methyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-055) :

A mixture of compound P-055-7 (80 mg, 0.09 mmol) , P-055-3 (30 mg, 0.09 mmol) , Sodium ascorbate (35 mg, 0.18 mmol) , CuSO₄.5H₂O (10 mg, 0.04mmol) in DCM (1 mL) , t-BuOH (2mL) and water (1mL) was heated at 40℃ for 5h. DCM (50 mL) was added and the organic phase was washed with H2O (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 30 mg of P-055 as a yellow solid. LCMS: [M+H] +: 617.5. Yelid: 28%

Preparation of compound P-048:

Preparation of dimethyl 3- ( (tert-butoxycarbonyl) amino) pentanedioate (P-048-2) :

A mixture of P-048-1 (3 g, 17.2 mmol) , Ammonium acetate (1.98 g, 25.8 mmol) , NaBH₃CN (1.60 g, 25.8 mmol) in DMF (50 mL) was stirred for 5h. O (Boc) ₂ (5.5 g, 25.8 mmol) was added and stirred at rt for 16h. DCM (200 mL) was added and the organic phase was washed with H₂O (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated , The crude product was purified by combine flash (PE: EA=3: 1, at 254nm) to give 2g of P-048-2 as a white solid. LCMS: [M+H] +: 276.1. Yelid: 42%

Preparation of tert-butyl (1, 5-dihydroxypentan-3-yl) carbamate (P-048-3) :

NaBH₄ (273 mg, 7.2 mmol ) was added to a solution of P-048-2 (1 g, 3.6 mmol) in methanol (50 mL) at rt. Then the mixture was heated at 40℃ for 16h. Concentrated and residue was purified by combine flash (DCM: CH₃OH=15: 1) to give 600 mg of P-048-3 as a white solid. LCMS: [M+H] +: 220.1. Yelid: 76%.

Preparation of tert-butyl (1- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) -5-hydroxypentan-3-yl) carbamate (P-048-5) :

A mixture of compound P-048-3 (600 mg, 2.72 mmol) , P-048-4 (290 mg, 1.05 mmol) , PPh₃ (700 mg, 2.72 mmol) and DIAD (550 mg, 2.72 mmol) in THF (25 ml) was stirred at rt for 16h. LCMS showed reaction was completed. EA (50 ml) was added. Washed with water (20ml x 2) . The organic layer was dried and concentrated, residue was purified by combine flash (DCM: CH3OH=15: 1) to give 750 mg of P-048-5 as a white solid. LCMS: [M+H] +: 476.2. Yelid: 58%.

Preparation of tert-butyl (1- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) -5-oxopentan-3-yl) carbamate (P-048-6) :

To a solution of compound P-048-5 (100 mg, 0.21 mmol) in DCM (20 mL) was added NaOAc (77 mg, 0.93 mmol) and PCC (202 mg, 0.93 mmol) . The mixture was stirred at RT for 2 h. LC-MS monitored and the starting material was consumed completed. DCM (50 mL) was added and the mixture was washed with saturated Na₂SO₃ aqueous solution (30 mL x 1) , saturated NaHCO₃ aqueous solution (30 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum to give the compound P-048-6 (100 mg, 100%) . The crude product was used in the next step directly. MS (ESI) : m/z 474.1 (M+H⁺) .

Preparation of tert-butyl (1- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) -5-oxopentan-3-yl) carbamate (P-048-7) :

A mixture of D-2 (50 mg, 0.06 mmol) , P-048-6 (60 mg, 0.13 mmol) , NaBH (OAc) ₃ (20 mg) in DMF (5 ml) and AcOH (0.5 ml) was stirred at RT for 16h. LCMS showed reaction was completed. DCM (100 mL) was added and the mixture was washed with H₂O (30 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0～10%) to give the compound P-048-7 (100 mg, yield 38%) . MS (ESI) : m/z 6545.6 (M/2+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (3-amino-5- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) oxy) pentyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-048) :

A mixture of P-048-7 (60 mg, 0.05) in DCM (10 ml) and TFA (2 ml) was stirred at RT for 2h. LCMS showed reaction was completed. Concentrated and the residue was purified by prep-HPLC to give the compound P-048 (11 mg, yield 17%) . MS (ESI) : m/z 605.5 (M/2+H⁺) .

Preparation of compound P-033:

Preparation of 7-azaspiro [3.5] nonane-2-carbonitrile hydrochloride (P-033-2) :

To a stirred solution of compound P-033-1 (1.0 g, 4 mmol) in DCM (20 mL) was added Dioxane/HCl (3 ml) was stirred at RT for 2 h. LCMS ok, The combined organic extracts were concentrated to give 900 mg of compound P-033-2 as white solid . LCMS: [M+H] ⁺: 151

Preparation of tert-butyl (4- (2-cyano-7-azaspiro [3.5] nonan-7-yl) -3-hydroxybutyl) carbamate (P-033-4) :

To a solution of compound P-033-2 (350 mg, 1.88 mmol) in EtOH (10 mL) was added compound P-033-3 (704 mg, 3.74 mmol) and K₂CO₃ (518 mg, 3.75 mmol) , Then the mixture was stirred at 80c for 1.5 h, LCMS showed OK, concentrated under vacuum , EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The residue was purified by combine flash (ME/DCM = 0～10%) to give the compound P-033-4 (600 mg, yield 94%) . MS (ESI) : m/z 338 (M+H⁺) .

Preparation of tert-butyl (4- (2-cyano-7-azaspiro [3.5] nonan-7-yl) -3-hydroxybutyl) carbamate (P-033-5) :

To a solution of compound P-033-4 (1.0 g, 2.9 mmol) and 2, 6-dimethylpyridine (962 mg, 10.10 mmol) in DCM (100 mL) was added DAST (955 mg, 5.90 mmol) at -78 C, Then the mixture was stirred at 0c for 2 h, LCMS showed ok, NaHCO₃ (50 ml) was added, DCM (50 mL) was added and the organic phase was washed with H₂O (100 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum . The crude product was purified by TLC (DCM: MeOH=5%-10%) to give compound P-033-5 (500 mg, yield 83%) . MS (ESI) : m/z 340 (M+H⁺) .

Preparation of tert-butyl (4- (2- (aminomethyl) -7-azaspiro [3.5] nonan-7-yl) -3-fluorobutyl) carbamate (P-033-6) :

To a solution of P-033-5 (500 mg, 1.47 mmol) in Dioxane (50 mL) was added Ni (500 mg) , Pd/C (500 mg) and LiOH/H₂O (5ml) , Then the mixture was stirred at 50c for 24 h under H₂, LCMS showed OK, filted, concentrated under vacuum , EA (10 mL) was added and the organic phase was concentrated under vacuum to give crude compound P-033-6 (400 mg) as a yellow oil. The crude product was used for next step .

Preparation of tert-butyl (3-fluoro-4- (2- ( ( (2-nitro-4-sulfamoylphenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) butyl) carbamate (P-033-8) :

To a solution of compound P-033-6 (500 mg, 1.45 mmol) in MeCN (50 mL) was added compound P-033-7 (516 mg, 2.19 mmol) and DIEA (565 mg) , Then the mixture was stirred at 90℃for 4 h, LCMS showed OK, concentrated under vacuum , EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum . The crude product was purified by HPLC to give 100 mg of Compound P-033-8 as a yellow solid. LCMS: [M+H] +: 544.3.

Preparation of tert-butyl (4- (2- ( ( (4- (N- (4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -2- ( (1- ( (2- (trimethylsilyl) ethoxy) methyl) -1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -3-fluorobutyl) carbamate (P-033-9) :

To a solution of Compound P-033-8 (100 mg, 0.18 mmol) and (R) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -2- ( (1- ( (2- (trimethylsilyl) ethoxy) methyl) -1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) benzoic acid (238 mg, 0.37 mmol) in anhydrous DMF (5 mL) was added EDCI (88 mg, 0.46 mmol) , DMAP (174 mg, 0.78 mmol) and DIEA (119 mg) at rt. The reaction mixture was stirred at 30 ℃ for 16 h under N₂ atmosphere. LCMS showed the reaction was completed. The reaction mixture was poured into EtOAc (300 mL) and washed with H₂O (2 x 250 mL) . The extract was washed with brine (1 x 200mL) , dried over Na₂SO₄, concentrated to afford 400 mg of crude product. The crude product was purified by TLC (DCM: MeOH=20: 1, at 254nm) to give 200 mg of Compound P-033-9 as a yellow solid. LCMS: [M+H] ⁺: 1172.3.

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (4-amino-2-fluorobutyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-033-10) :

To a solution of Compound P-033-10 (200 mg, 0.17 mmol) and TFA (3 ml) in anhydrous DCM (10 mL) was stirred at RT for 4 h under N₂ atmosphere. LCMS showed the reaction was completed. The reaction mixture was concentrated to afford 150 mg of crude product. Then MeOH (50 ml) and K₂CO₃ (30ml, aq) was added, the mixture was stirred for 16h, LCMS showed the reaction was completed. 1N HCl was added to PH=6, The reaction mixture was poured into EtOAc (30 mL) and washed with H₂O (2 x 25 mL) . The extract was washed with brine (1 x 200mL) , dried over Na₂SO₄, concentrated to afford 150 mg of crude product. The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 100 mg of Compound P-033-10 as a yellow solid. LCMS: [M+H] ⁺: 942.3.

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) -2-fluorobutyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-033) :

To a solution of Compound P-033-10 (100 mg, 0.11 mmol) in DMSO (2 mL) was added Compound 12 (44 mg, 0.16 mmol) and DIEA (43 mg) , Then the mixture was stirred at 130℃ for 2 h under Mw, LCMS showed OK , EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by TLC (DCM: MeOH=20: 1) to give 30 mg of Compound P-033 as a yellow solid LCMS: [M+H] +: 1198.3.

Preparation of compound P-038:

Preparation of 2- (2, 2-difluoro-3-hydroxypropyl) isoindoline-1, 3-dione (P-038-2) :

To a solution of compound P-038-1 (200 mg, 1.80 mmol) and compound isobenzofuran-1, 3-dione (267 mg, 1.80 mmol) in toluene (10 mL) was added TEA (0.25 mL, 1.80 mmol) . The mixture was stirred at 120 ℃ for 3 h. LC-MS monitored and desired product produced. The solvent was removed under vacuum and the residue was purified by combine flash (EA/PE = 0～50%) to give the compound P-038-2 (260 mg, yield 60%) . MS (ESI) : m/z 242.0 (M+H⁺) .

Preparation of 3- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropropyl trifluoromethanesulfonate (P-038-3) :

To a solution of compound P-038-2 (100 mg, 0.42 mmol) in DCM (8 mL) was added 2, 6-dimethylpyridine (0.1 mL, 0.84 mmol) and TfOTf (0.1 mL, 0.62 mmol) , and the mixture was stirred at 0 ℃ for 2 h. LC-MS monitored. H₂O (20 mL) was added and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0 ～ 50%) to give the compound P-038-3 (130 mg, yield 84%) . MS (ESI) : m/z 374.1 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (3- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropropyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-038-4) :

To a solution of compound P-038-3 (460 mg, 1.23 mmol) and compound D-2 (316 mg, 0.37 mmol) in DMSO (15 mL) was added DIPEA (0.82 mL, 4.93 mmol) . The mixture was stirred at 40 ℃ overnight. LC-MS monitored and desired product was produced. DCM (100 mL) was added and the mixture was washed with H₂O (30 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (ME/DCM = 0 ～10%) to give the compound P-038-4 (360 mg, yield 90%) . MS (ESI) : m/z 1076.7 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (3- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropropyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-038-5) :

To a solution of compound P-038-4 (100 mg, 0.092 mmol) in EtOH (25 mL) was added 85%hydrazinium hydroxide solution (47 mg, 0.92 mmol) . The mixture was stirred at 80 ℃ for 2 h. LC-MS monitored and desired product was produced. The solvent was removed under vacuum. The residue was purified by combine flash (ME/DCM = 0～10%) to give the compound P-038-5 (50 mg, yield 57%) . MS (ESI) : m/z 946.2 (M+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (3- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) -2, 2-difluoropropyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-038) :

To a solution of compound P-038-5 (10 mg, 0.01 mmol) and) and compound 2-1 (4 mg, 0.016 mmol) in DMSO (2 mL) was added DIPEA (4 mg, 0.031 mmol) . The mixture was stirred at 130 ℃ under microwave for 2 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give the product P-038 (2.3 mg, yield 18%) . MS (ESI) : m/z 1202.3 (M+H⁺) .

Preparation of compound P-039:

Preparation of tert-butyl (R) -4- (2- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) acetyl) piperazine-1-carboxylate (P-039-2) :

A mixture of D-2 (50 mg, 0.05 mmol) , P-039-1 (16 mg, 0.06 mmol) , NaI (8 mg, 0.05 mmol) in THF (3 ml) and DMSO (5 ml) was heated at 60℃ for 6h. DCM (50 mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (DCM/CH3OH = 10: 1) to give the compound P-039-2 (60 mg, yield 100%) . MS (ESI) : m/z 540.8 (M/2+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (2-oxo-2- (piperazin-1-yl) ethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-039-3) :

A mixture of P-039-2 (60 mg, 0.06 mmol) , TFA (1ml) , DCM (10 ml) was stirred at RT for 2h. Concentrated and DCM (50 mL) was added. pH of solution was adjusted to 7-8 with Na₂CO₃ solution. The organic layer was separated, washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated to give the compound P-039-3 (50 mg, yield 89%) . MS (ESI) : m/z 490.8 (M/2+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (2- (4- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) piperazin-1-yl) -2-oxoethyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-039) :

A mixture of P-039-3 (50 mg, 0.05 mmol) , 8-4 (24 mg, 0.08 mmol) , DIEA (0.5 ml) in DMSO (3 ml) was heated at 130℃ for 3h under MW. The reaction solution was purified by prep-HPLC in reverse and lyophilized to obtain P-039 (11.3 mg, yield: 18%) . MS (ESI) : m/z 618.3 (M/2+H⁺) .

Preparation of compound P-041:

Preparation of ethyl 4- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoro-3-hydroxybutanoate (P-041-3) :

To a stirred solution of compound P-041-2 (11.0 g, 0.05 mmol) in THF (100 mL) was added Zn (3.5 g) was stirred at 60-rt for 1 h. then compound P-041- 1 (5.0 g, 0.03 mmol) in THF (50 ml) was added dropwise, the mixture was stirred at rt for 2h, LCMS ok, NaHCO3 (aq) was added, EA (200 mL) was added and the organic phase was washed with H2O (150 mL x 2) , brine, dried over anhydrous Na2SO4 and concentrated under vacuum . The crude product was purified by TLC (DCM: MeOH=5%-10%) to give 4.8 g of Compound P-041-3 as a colourless oil. LCMS: [M+H] +: 314.3.

Preparation of ethyl 3- ( (1H-imidazole-1-carbonothioyl) oxy) -4- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluorobutanoatehydroxybutanoate (P-041-5) :

To a solution of compound P-041-3 (4.8 g, 15.3 mmol ) in EtOH (50 mL) was added compound P-041-4 (4.1 g, 23 mmol) and DMAP (187 mg) , Then the mixture was stirred at RT for 2 h, LCMS showed OK, , DCM (50 mL) was added and the organic phase was washed with H2O (50 mL x 2) , brine, dried over anhydrous Na2SO4 and concentrated under vacuum, The crude product was purified by TLC (DCM: MeOH=20: 1) to give 4.8 g of Compound P-041-5 as a yellow oil. LCMS: [M+H] +: 424.3.

Preparation of ethyl 4- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluorobutanoate (P-041-6) :

To a solution of compound P-041-5 (3.0 g, 7.09 mmol) and Et₃SiH (50 ml) in Toluene (50 mL) was added stirred at 90 C , the BPO (2.3 g, 9.5 mmol) in Toluene (20 mL) was added, , Then the mixture was stirred at 90 C for 4 h, LCMS showed ok, concentrated under vacuum . The crude product was purified by TLC (PE: EA=0%-50%) to give 1.4g of Compound P-041-6. LCMS: [M+H] +: 298.1.

Preparation of 4- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluorobutanoic acid (P-041-7) :

To a solution of compound P-041-6 (1.5 g, 5.01 mmol ) in MeOH (20 mL) was added LiOH (7 ml, 1N) , Then the mixture was stirred at RT for 2 h under H₂, LCMS showed OK, HCl (1N) was added, , DCM (50 mL x 2) was added and the organic phase was washed with H₂O (50 mL) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum, The crude product was purified by HPLC to give 500 mg of Compound P-041-7 as a white solid. LCMS: [M+H] +: 270.1.

Preparation of 2- (3, 3-difluoro-4-hydroxybutyl) isoindoline-1, 3-dione (P-041-8) :

To a solution of compound P-041-7 (500 mg, 1.85 mmol) in THF (50 mL) was added BH₃/Me₂S (1.1 ml, 1N) , Then the mixture was stirred at RT for 4 h, LCMS showed OK, MeOH (5 mL) was added and concentrated under vacuum . The crude product was purified by HPLC to give 100 mg of Compound P-041-8 as a colourless oli. LCMS: [M+H] +: 256.1.

Preparation of 4- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluorobutyl trifluoromethanesulfonate (P-041-9) :

To a solution of compound P-041-8 (100 mg, 0.39 mmol) in DCM (20 mL) was added 2, 6-dimethylpyridine (0.12 mL, 1.04 mmol) and trifluoroacetic anhydride (0.13 mL, 0.78 mmol) , and the mixture was stirred at 0 ℃ for 2 h. LC-MS monitored. H₂O (50 mL) was added and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0 ～20%) to give the compound P-041-9 (100 mg, yield 66%) . MS (ESI) : m/z 388.0 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (4- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluorobutyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-041-10) :

To a solution of compound P-041-9 (100 mg, 0.26 mmol) and compound D-2 (220 mg, 0.26 mmol) in DMSO (5 mL) was added DIPEA (0.31 mL, 1.87 mmol) . The mixture was stirred at 40 ℃ overnight. LC-MS monitored and desired product was produced. DCM (50 mL) was added and the mixture was washed with H₂O (30 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by Prep-TLC (ME/DCM = 1/10) to give the compound P-041-10 (150 mg, yield 57%) . MS (ESI) : m/z 545.8 (M/2+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (4-amino-2, 2-difluorobutyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3- nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-041-11) :

To a solution of compound P-041-10 (150 mg, 0.16 mmol) and in EtOH (40 mL) was added 85%hydrazinium hydroxide solution (104 mg, 2.08 mmol) . The mixture was stirred at 80 ℃ for 5 h. LC-MS monitored and the desired product was produced. The solvent was removed under vacuum and the residue was purified by Prep-TLC (ME/DCM = 1/10) to give the compound P-041-11 (80 mg, yield 52%) . MS (ESI) : m/z 960.3 (M+H⁺) .

Preparation of 2- (3, 3-difluoro-4-hydroxybutyl) isoindoline-1, 3-dione (P-041) :

To a solution of compound P-041-11 (80 mg, 0.08 mmol) in DMSO (2 mL) was added compound 2-1 (35 mg, 0.13 mmol) and DIEA (31 mg, 0.24 mmol) , Then the mixture was stirred at 130℃ for 2 h under Mw, LCMS showed OK , EA (50 mL) was added and the organic phase was washed with H₂O (50 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum , The crude product was purified by TLC (DCM: MeOH=10: 1, at 254nm) to give 65 mg of Compound P-041 as a yellow solid. Yield: 67%. LCMS: [M+H] ⁺: 1216.3.

Preparation of compound P-050:

Preparation of ethyl 5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoro-3-hydroxypentanoate (P-050-3) :

To a solution of Zn (3.22 g, 49.21 mmol) in THF (100 mL) was added compound P-050-2 (9.99 g, 49.21 mmol) . The mixture was stirred at RT for 30 min. Then compound P-050-1 (5.0 g, 24.61 mmol) was added and the mixture was stirred at 70 ℃ for 2 h. Filter to remove unreacted Zn. EA (100 mL) was added the mixture was washed with saturated NH₄Cl solution (40 mL x 2) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～50%) to give the compound P-050-3 (3.5 g, yield 43%) . MS (ESI) : m/z 328.1 (M+H⁺) .

Preparation of ethyl 3- ( (1H-imidazole-1-carbonothioyl) oxy) -5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropentanoate (P-050-5) :

To a solution of compound P-050-3 (3.5 g, 10.69 mmol) in DCM (100 mL) was added compound P-050-4 (2.86 g, 16.04 mmol) and DMAP (131 mg, 1.07 mmol) . The mixture was stirred at RT for 2 h. The solvent was removed under vacuum. The residue was purified by combine flash (EA/PE = 0～50%) to give the compound P-050-5 (3.4 g, yield 73%) . MS (ESI) : m/z 438.1 (M+H⁺) .

Preparation of ethyl 5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropentanoate (P-050-6) :

To a solution of compound P-050-5 (3.4 g, 7.77 mmol) in toluene (16 mL) and Et₃SiH (16 mL) was added benzoic peroxyanhydride (2.07 g, 8.55 mmol) slowly. The mixture was stirred at 120 ℃ for 3 h. LC-MS monitored and desired product was produced. The solvent was removed under vacuum. The residue was purified by combine flash (EA/PE = 0 ～ 30%) to give the compound P-050-6 (2.2 g, yield 91%) . MS (ESI) : m/z 312.1 (M+H⁺) .

Preparation of 5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropentanoic acid (P-050-7) :

To a solution of compound P-050-6 (1.4 g, 4.5 mmol) in AcOH (15 mL) was added 6M HCl aqueous solution (1.5 mL, 9.0 mmol) . The mixture was stirred at 90 ℃ for 2 h. LC-MS monitored and desired product was produced. The solvent was removed under vacuum. H₂O (100 mL) was added and the mixture was extracted with EA (50 mL x 2) . The combined organic solvent was removed under vacuum. The residue was purified by combine flash (ME/DCM = 0～10%) to give the compound P-050-7 (1.17 g, yield 91%) . MS (ESI) : m/z 284.1 (M+H⁺) .

Preparation of ethyl 3- ( (1H-imidazole-1-carbonothioyl) oxy) -5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropentanoate (P-050-8) :

To a solution of compound P-050-7 (900 mg, 3.18 mmol) in THF (30 mL) was added 2M BH₃ ^. Me₂S (2.4 mL, 4.8 mmol) . The mixture was stirred at RT for 24 h. LC-MS monitored and desired product was produced. MeOH (20 mL) was added slowly to quench the reaction. The solvent was removed under vacuum. The residue was purified by combine flash (EA/PE = 0～30%) to give the compound P-050-8 (560 mg, yield 51%) . MS (ESI) : m/z 270.1 (M+H⁺) .

Preparation of 5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropentyl trifluoromethanesulfonate (P-050-9) :

To a solution of compound P-050-8 (280 mg, 1.04 mmol) in DCM (20 mL) was added 2, 6-dimethylpyridine (0.24 mL, 2.08 mmol) and TfOTf (0.26 mL, 1.56 mmol) , and the mixture was stirred at 0 ℃ for 2 h. LC-MS monitored. H₂O (50 mL) was added and the mixture was extracted with DCM (20 mL x 3) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0 ～ 20%) to give the compound P-050-9 (150 mg, yield 36%) . MS (ESI) : m/z 402.0 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (5- (1, 3-dioxoisoindolin-2-yl) -2, 2-difluoropentyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-050-10) :

To a solution of compound P-050-9 (150 mg, 0.37 mmol) and compound D-2 (160 mg, 0.19 mmol) in DMSO (5 mL) was added DIPEA (0.31 mL, 1.87 mmol) . The mixture was stirred at 40 ℃ overnight. LC-MS monitored and desired product was produced. DCM (50 mL) was added and the mixture was washed with H₂O (30 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by Prep-TLC (ME/DCM = 1/10) to give the compound P-050-10 (166 mg, yield 80%) . MS (ESI) : m/z 552.8 (M/2+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (5-amino-2, 2-difluoropentyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-050-11) :

To a solution of compound P-050-10 (230 mg, 0.21 mmol) and in EtOH (40 mL) was added 85%hydrazinium hydroxide solution (104 mg, 2.08 mmol) . The mixture was stirred at 80 ℃ for 5 h. LC-MS monitored and the desired product was produced. The solvent was removed under vacuum and the residue was purified by Prep-TLC (ME/DCM = 1/10) to give the compound P-050-11 (30 mg, yield 15%) . MS (ESI) : m/z 487.8 (M/2+H⁺) .

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- (5- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-4-yl) amino) -2, 2-difluoropentyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-050) :

To a solution of compound P-050-11 (30 mg, 0.031 mmol) and compound 2-1 (17 mg, 0.062 mmol) in DMSO (3 mL) was added DIPEA (20 mg, 0.15 mmol) . The mixture was stirred at 130 ℃ under microwave for 1 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give the product P-050 (6.3 mg, yield 17%) . MS (ESI) : m/z 615.8 (M/2+H⁺) .

Preparation of compound P-051:

Preparation of ethyl 2- (2-cyanospiro [3.5] nonan-7-ylidene) acetate (P-051-3) :

To a solution of 60%NaH (85 mg, 2.11 mmol) in THF (15 mL) was added compound P-051-2 (433 mg, 1.93 mmol) in 0 ℃. The mixture was stirred at 0 ℃ for 1 h. Then compound P-051-1 (300 mg, 1.84 mmol) was added at 0 ℃ and the mixture was stirred at RT for 2 h. H₂O (30 mL) was added and the mixture was extracted with EA (20 mL x 2) . The combined organic phase was washed with brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～15%) to give compound P-051-3 (400 mg, yield 93%) . MS (ESI) : m/z 234.3 (M+H⁺) .

Preparation of ethyl 2- (2- (aminomethyl) spiro [3.5] nonan-7-yl) acetate (P-051-4) :

To a solution of compound P-051-3 (400 mg, 1.71 mmol) in MeOH (25 mL) was added Pd/C (300 mg) and the mixture was stirred at RT for 3 h under H₂ atmosphere. LC-MS monitored and the starting material was consumed completed. Filtrate to remove the Pd/C and the solvent was removed under vacuum to give compound P-051-4 (340 mg, yield 83%) . MS (ESI) : m/z 240.3 (M+H⁺) .

Preparation of ethyl 2- (2- ( ( (tert-butoxycarbonyl) amino) methyl) spiro [3.5] nonan-7-yl) acetate (P-051-5) :

To a solution of compound P-051-4 (340 mg, 1.42 mmol) in DCM (25 mL) was added TEA (431 mg, 4.26 mmol) and (Boc) ₂O (465 mg, 2.13 mmol) . The mixture was stirred at RT for 2 h. LC-MS monitored and the starting material was consumed completed. The mixture was washed with H₂O (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～50%) to give compound P-051-5 (370 mg, yield 77%) . MS (ESI) : m/z 240.3 (M-100+H⁺) .

Preparation of 2- (2- ( ( (tert-butoxycarbonyl) amino) methyl) spiro [3.5] nonan-7-yl) acetic acid (P-051-6) :

To a solution of compound P-051-5 (350 mg, 1.03 mmol) in MeOH (20 mL) was added 1 M LiOH aqueous solution (10 mL, 10.0 mmol) . The mixture was stirred at RT overnight. LC-MS monitored and the starting material was consumed completed. Saturated NaH₂PO₄ aqueous solution was added to adjust pH to 6. The mixture was extracted with EA (30 mL x 3) and the combined organic was washed with brine, dried over anhydrous Na₂SO₄ and concentrated to give compound P-051-6 (321 mg, yield 100%) . The product was used in the next step without further purification. MS (ESI) : m/z 256.1 (M-56+H⁺) .

Preparation of tert-butyl ( (7- (2-hydroxyethyl) spiro [3.5] nonan-2-yl) methyl) carbamate (P-051-7) :

To a solution of compound P-051-6 (321 mg, 1.03 mmol) in THF (5 mL) was added 2 M BH₃ ^. Me₂S (5 mL, 10.3 mmol) . The mixture was stirred at RT for 4 h. LC-MS monitored and the starting material was consumed completed. MeOH (20 mL) was added slowly to quench the reaction. The solvent was removed under vacuum to give compound P-051-7 (300 mg, yield 98%) . The product was used in the next step without further purification. MS (ESI) : m/z 298.2 (M+H⁺) .

Preparation of 2- (2- (aminomethyl) spiro [3.5] nonan-7-yl) ethan-1-ol (P-051-8) :

To a solution of compound P-051-7 (300 mg, 1.01 mmol) in DCM (6 mL) was added 4 M HCl/dioxane (6 mL, 24.0 mmol) . The mixture was stirred at RT for 1 h. LC-MS monitored and the starting material was consumed completed. The solvent was removed under vacuum to give compound P-051-8 (199 mg, yield 100%) . The product was used in the next step without further purification. MS (ESI) : m/z 198.3 (M+H⁺) .

Preparation of 4- ( ( (7- (2-hydroxyethyl) spiro [3.5] nonan-2-yl) methyl) amino) -3-nitrobenzenesulfonamide (P-051-9) :

To a solution of compound P-051-8 (200 mg, 1.01 mmol) and compound 4-chloro-3-nitrobenzenesulfonamide (360 mg, 1.52 mmol) in CH₃CN (20 mL) was added DIPEA (1.7 mL, 10.14 mmol) . The mixture was stirred at 90 ℃ for 7 h. LC-MS monitored and the starting material was consumed completed. The solvent was removed under vacuum. The residue was purified by combine flash (ME/DCM = 0～10%) to give compound P-051-9 (100 mg, yield 25%) . MS (ESI) : m/z 398.2 (M+H⁺) .

Preparation of 3-nitro-4- ( ( (7- (2-oxoethyl) spiro [3.5] nonan-2-yl) methyl) amino) benzenesulfonamide (P-051-10) :

To a solution of compound P-051-9 (90 mg, 0.23 mmol) in DCM (30 mL) was added NaOAc (56 mg, 0.68 mmol) and PCC (146 mg, 0.68 mmol) . The mixture was stirred at RT for 2 h. LC-MS monitored and the starting material was consumed completed. DCM (60 mL) was added and the mixture was washed with saturated Na₂SO₃ aqueous solution (30 mL x 1) , saturated NaHCO₃ aqueous solution (30 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated under vacuum to give compound P-051-10 (90 mg, 100%) . The crude product was used in the next step directly. MS (ESI) : m/z 396.1 (M+H⁺) .

Preparation of tert-butyl 4- (2- (2- ( ( (2-nitro-4-sulfamoylphenyl) amino) methyl) spiro [3.5] nonan-7-yl) ethyl) piperazine-1-carboxylate (P-051-11) :

To a solution of compound P-051-10 (90 mg, 0.23 mmol) and compound tert-butyl piperazine-1-carboxylate (85 mg, 0.46 mmol) in 1, 2-dichloroethane (20 mL) was added NaBH (OAc) ₃ (96 mg, 0.46 mmol) . The mixture was stirred at RT for 2 h. LC-MS monitored and the desired product was produced. DCM (20mL) was added and the mixture was washed with H₂O (20 mL x 1) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by Prep-TLC (ME/DCM=1/20) to give compound P-051-11 (100 mg, yield 70%) . MS (ESI) : m/z 566.3 (M+H⁺) .

Preparation of tert-butyl (R) -4- (2- (2- ( ( (4- (N- (4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -2- ( (1- ( (2- (trimethylsilyl) ethoxy) methyl) -1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) spiro [3.5] nonan-7-yl) ethyl) piperazine-1-carboxylate (P-051-12) :

To a solution of compound P-051-11 (100 mg, 0.18 mmol) and compound (R) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -2- ( (1- ( (2- (trimethylsilyl) ethoxy) methyl) -1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) benzoic acid (149 mg, 0.23 mmol) in DMF (3 mL) and DCM (1.5 mL) was added EDCI (51 mg, 0.27 mmol) , DMAP (65 mg, 0.53 mmol) and DIPEA (69 mg, 0.53 mmol) . The mixture was stirred at RT overnight. LC-MS monitored and the desired product was produced. EA (50mL) was added and the mixture was washed with H₂O (20 mL x 3) , brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (EA/PE = 0～100%) to give compound P-051-12 (110 mg, yield 52%) . MS (ESI) : m/z 597.5 (M/2+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (2- (piperazin-1-yl) ethyl) spiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-051-13) :

To a solution of compound P-051-12 (100 mg, 0.084 mmol) in DCM (9 mL) was added TFA (3 mL) . The mixture was stirred at RT for 1 h. LC-MS monitored and the desired product was produced. The solvent was removed under vacuum. The residue was dissolved in DCM (5 mL) and MeOH (20 mL) and saturated K₂CO₃ aqueous solution (10 mL) was added. The mixture was stirred at RT for 2 h. LC-MS monitored and the desired product was produced. The solvent was removed under vacuum. DCM (50 mL) was added and the mixture was washed with H₂O (30 mL x 2) . The solvent was removed under vacuum to give compound P-051-13 (80 mg, yield 100%) . The crude product was used in the next step directly. MS (ESI) : m/z 964.4 (M+H⁺) .

Preparation of (R) -2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- (1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (3-nitro-4- ( ( (7- (2- (piperazin-1-yl) ethyl) spiro [3.5] nonan-2-yl) methyl) amino) phenyl) sulfonyl) benzamide (P-051) :

To a solution of compound P-051-13 (80 mg, 0.083 mmol) and compound 8-4 (46 mg, 0.17 mmol) in DMSO (3 mL) was added DIPEA (54 mg, 0.41 mmol) . The mixture was stirred at 120 ℃ under microwave for 1 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give the product P-051 (43.5 mg, yield 43%) . MS (ESI) : m/z 610.3 (M/2+H⁺) .

Preparation of compound P-054:

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) -N- ( (4- ( ( (7- ( (4- (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) piperazin-1-yl) methyl) spiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) benzamide (P-054) :

To a solution of compound P-054-1 (50 mg, 0.05 mmol) and) and compound 8-4 (22 mg, 0.08 mmol) in DMSO (2 mL) was added DIPEA (40 mg, 0.31 mmol) . The mixture was stirred at 130℃ under microwave for 2 h. LC-MS monitored and desired product was produced. The mixture was purified by Prep-HPLC to give the product P-054 (23 mg, yield 38%) . MS (ESI) : m/z 603.8 (M/2+H⁺) .

Preparation of compound P-052:

Preparation of benzyl N, N, O-tribenzylhomoserinate (P-052-2) :

To a stirred solution of compound P-052-1 (1.0 g, 8.4 mmol) , compound (bromomethyl) benzene (7.2 g, 42 mmol) and K₂CO₃ (5.8 g) in H₂O (50 mL) was stirred at 80C for 2 h. The mixture was diluted with water (20 mL) and was extracted with EtOAc (50mL x 3) . The combined organic extracts were concentrated. The crude product was purified by TLC (DCM: MeOH=0-10%, at 254nm) to give 275 mg of Compound P-052-2 as a yellow oil. LCMS: [M+H] +: 480.

Preparation of 4- (benzyloxy) -2- (dibenzylamino) butan-1-ol (P-052-3) :

A solution of compound P-052-2 (800 mg, 1.67 mmol) in THF (20 ml) was added LiAlH₄ (190 mg) at RT. Then the reaction mixture was stirred at rt for 1 h. LCMS showed the reaction was completed. Na2SO4.10H2O was added, filtered, the reaction mixture was concentrated to afford 800 mg of crude P-052-3.

Preparation of N, N-dibenzyl-4- (benzyloxy) -1-chlorobutan-2-amine (P-052-4) :

A solution of compound P-052-3 (800 mg, 2.13 mmol) in DCM (20 ml) was added MsCl (150 mg, 1.31 mmol) , DIEA (200 mg) at rt. Then the reaction mixture was stirred at rt for 4 h. LCMS showed the reaction was completed. The mixture was diluted with water (100 mL) and was extracted with DCM (50mL X 3) . The combined organic extracts were concentrated. The crude product was purified by TLC (DCM: MeOH=0-10%, at 254nm) to give 700 mg of Compound P-052-4 as a yellow oil. LCMS: [M+H] +: 394.

Preparation of 5- (4- (benzyloxy) -2- (dibenzylamino) butoxy) -2- (2, 6-dioxopiperidin-3-yl) isoindoline-1, 3-dione (P-052-6) :

To a stirred solution of compound P-052-4 (630 mg, 1.61 mmol) , P-052-5 (876 mg, 3.18 mmol) , K₂CO₃ (331 mg, 2.39 mmol) and NaI (10 mg) in DMF (10 mL) was stirred at 60℃ for 2 h. The mixture was diluted with water (100 mL) and was extracted with DCM (50mL X 3) . The combined organic extracts were concentrated. The crude product was purified by TLC (DCM: MeOH=0-10%, at 254nm) to give 520 mg of Compound P-052-6 as a yellow solid. LCMS: [M+H] ⁺: 632.

Preparation of tert-butyl (1- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) -4-hydroxybutan-2-yl) carbamate (P-052-7) :

A solution of compound P-052-6 (50 mg) in dioxnae (10 ml) was added Pd/C (10 mg) , HCl (0.01 ml) at rt under H₂. Then the reaction mixture was stirred at 50c for 3 h.. LCMS showed the reaction was completed. The reaction mixture was filtered and concentrated to afford 50 mg of crude product. The crude product was added DCM (10 ml) , O (Boc) ₂ (20 mg) and DIEA (20 mg) , then the mixture was stirred at rt for 16 h. LCMS showed the reaction was completed, The mixture was diluted with water (20 mL) and was extracted with DCM (20mL X 3) . The combined organic extracts were concentrated. The crude product was purified by TLC (DCM: MeOH=0-10%, at 254nm) to give 20 mg of Compound P-052-7 as a yellow solid. LCMS: [M+H] +: 462.

Preparation of tert-butyl (1- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) -4-oxobutan-2-yl) carbamate (P-052-8) :

A mixture of compound P-052-7 (100 mg) , PCC (140 mg) , NaOAc (53 mg) in DCM (15 ml) was stirred at RT for 3h. LCMS showed reaction was completed. Na₂SO₃ (100 mg) was added and stirred for 30 min. DCM (50 mL) was added. Washed with water (20mL x 2) . The organic layer was dried and concentrated, The residue was purified by combine flash (DCM: CH₃OH=0-10%, normal phase silica, UV254) to give 90 mg P-052-8 as yellow solid.

Preparation of tert-butyl (4- (2- ( ( (4- (N- (2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzoyl) sulfamoyl) -2-nitrophenyl) amino) methyl) -7-azaspiro [3.5] nonan-7-yl) -1- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) butan-2-yl) carbamate (P-052) :

A solution of compound P-052-8 (100 mg) in DMF (10 ml) was added D-2 (278 mg) , AcOH (48 mg) and NaBH₃Ac (121 mg) at RT. Then the reaction mixture was stirred at rt for 4 h. LCMS showed the reaction was no reaction. The reaction mixture was diluted with water (50 mL) and was extracted with EtOAc (30 mL x 3) . The combined organic extracts were concentrated. The residue was purified by flash chromatography (0 to 10 %MeOH in DCM) to give 130 mg of P-052 as yellow solid LCMS: [M+H] ⁺: 1297

Preparation of compound P-053:

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (3-amino-4- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) butyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-053) :

A mixture of compound P-052 (30 mg, 0.023 mmol) TFA (0.5 ml) in DCM (50 ml) was stirred at rt for 2h. Concentrated and residue was purified by prep-HPLC to give the product compound P-053 (15 mg, yield 51%) . MS (ESI) : m/z 598.9 (M/2+H⁺) .

Preparation of compound P-056:

Preparation of 2- ( (1H-pyrrolo [2, 3-b] pyridin-5-yl) oxy) -N- ( (4- ( ( (7- (3-acetamido-4- ( (2- (2, 6-dioxopiperidin-3-yl) -1, 3-dioxoisoindolin-5-yl) oxy) butyl) -7-azaspiro [3.5] nonan-2-yl) methyl) amino) -3-nitrophenyl) sulfonyl) -4- (4- ( (R) -1-chloro-6, 7, 8, 9-tetrahydro-5H-benzo [7] annulen-5-yl) piperazin-1-yl) benzamide (P-056) :

A mixture of compound P-053 (30 mg, 0.023 mmol) , 2, 5-dioxopyrrolidin-1-yl acetate (5 mg, 0.03 mmol) and TEA (0.5 ml) in DMF (5 ml) was stirred at rt for 2h. DCM (50 ml) was added, It was washed with brine, dried over anhydrous Na₂SO₄ and concentrated. The residue was purified by combine flash (DCM/MeOH = 1: 10) to give the compound P-056 (20 mg, yield 70%) . MS (ESI) : m/z 619.6 (M/2+H⁺) .

The following compounds were prepared by above-described synthesis routes with modifications.

2.11 1H NMR data of PROTAC Compounds

P-001

1H NMR (400 MHz, DMSO) δ 11.51 (s, 1H) , 11.11 (s, 1H) , 8.36 (s, 1H) , 8.30 –8.18 (m, 4H) , 7.91 (d, J = 2.4 Hz, 1H) , 7.86 (s, 1H) , 7.63 –7.53 (m, 3H) , 7.39 (s, 1H) , 7.31 –7.21 (m, 2H) , 7.14 –7.00 (m, 4H) , 6.73 –6.59 (m, 3H) , 6.32 –6.26 (m, 2H) , 5.05 (dd, J = 12.8, 5.4 Hz, 1H) , 3.45 –2.97 (m, 16H) , 2.59 –2.54 (m, 2H) , 2.44 –2.26 (m, 5H) , 2.19 –2.02 (m, 5H) , 2.01 –1.94 (m, 2H) , 1.89 –1.74 (m, 5H) , 1.59 –1.45 (m, 9H) , 1.25 –1.15 (m, 2H) .

P-002

1H NMR (500 MHz, DMSO) δ 11.50 (s, 1H) , 8.98 (s, 1H) , 8.57 (s, 1H) , 8.37 (s, 1H) , 8.22 (s, 3H) , 7.94 –7.84 (m, 2H) , 7.76 (s, 1H) , 7.58 (t, J = 9.6 Hz, 2H) , 7.44 –7.36 (m, 5H) , 7.30 –7.22 (m, 2H) , 7.14 –7.04 (m, 2H) , 6.69 (dd, J = 36.7, 8.6 Hz, 2H) , 6.29 (s, 2H) , 4.56 –4.18 (m, 6H) , 3.69 –3.59 (m, 4H) , 3.31 –3.04 (m, 13H) , 2.44 (s, 3H) , 2.20 –1.45 (m, 27H) , 1.25 –1.13 (m, 1H) , 0.94 (s, 9H) .

P-003

1H NMR (400 MHz, DMSO) δ 11.59 (s, 1H) , 8.98 (s, 1H) , 8.57 (t, J = 6.0 Hz, 1H) , 8.48 –8.34 (m, 2H) , 8.14 (s, 1H) , 7.99 –7.90 (m, 2H) , 7.86 (d, J = 9.3 Hz, 1H) , 7.72 –7.65 (m, 1H) , 7.56 (d, J = 8.7 Hz, 1H) , 7.47 –7.35 (m, 6H) , 7.30 –7.25 (m, 1H) , 7.12 –7.04 (m, 2H) , 6.89 (s, 1H) , 6.67 (d, J = 9.0 Hz, 1H) , 6.33 (s, 1H) , 6.26 (s, 1H) , 5.13 (s, 1H) , 4.54 (d, J = 9.2 Hz, 1H) , 4.47 –4.39 (m, 2H) , 4.35 (s, 1H) , 4.21 (dd, J = 15.7, 4.9 Hz, 1H) , 3.69 –3.61 (m, 2H) , 3.25 –3.20 (m, 2H) , 3.15 –3.00 (m, 7H) , 2.87 –2.68 (m, 4H) , 2.44 (s, 3H) , 2.34 –2.22 (m, 2H) , 2.14 –1.99 (m, 9H) , 1.97 –1.80 (m, 7H) , 1.74 –1.69 (m, 1H) , 1.66 –1.54 (m, 6H) , 1.52 –1.41 (m, 6H) , 1.28 –1.14 (m, 4H) , 0.93 (s, 9H) .

P-004

1H NMR (500 MHz, DMSO) δ 11.50 (s, 1H) , 8.98 (s, 1H) , 8.56 (t, J = 5.8 Hz, 1H) , 8.36 (s, 1H) , 8.28 –8.17 (m, 4H) , 7.91 (d, J = 2.2 Hz, 1H) , 7.83 (d, J = 9.6 Hz, 1H) , 7.74 –7.69 (m, 1H) , 7.62 –7.54 (m, 2H) , 7.44 –7.36 (m, 5H) , 7.30 –7.21 (m, 2H) , 7.13 –7.05 (m, 2H) , 6.68 (dd, J = 29.2, 8.5 Hz, 2H) , 6.29 (d, J = 7.6 Hz, 2H) , 4.54 (d, J = 9.3 Hz, 1H) , 4.45 –4.32 (m, 3H) , 4.22 (dd, J = 15.9, 5.4 Hz, 1H) , 3.41 –3.02 (m, 15H) , 2.44 (s, 3H) , 2.38 –2.25 (m, 5H) , 2.18 –1.82 (m, 14H) , 1.51 (t, J = 30.0 Hz, 13H) , 1.27 –1.14 (m, 8H) , 0.93 (s, 9H) .

P-005

1H NMR (400 MHz, DMSO) δ 11.50 (s, 1H) , 11.11 (s, 1H) , 8.37 (s, 1H) , 8.21 (s, 3H) , 7.91 (d, J = 2.5 Hz, 1H) , 7.84 –7.77 (m, 1H) , 7.62 –7.51 (m, 4H) , 7.45 (d, J = 7.2 Hz, 1H) , 7.39 (s, 1H) , 7.31 –7.23 (m, 2H) , 7.14 –7.04 (m, 2H) , 6.72 (d, J = 9.1 Hz, 1H) , 6.65 (d, J = 8.5 Hz, 1H) , 6.32 –6.27 (m, 2H) , 5.08 (dd, J = 12.6, 5.4 Hz, 1H) , 4.38 –4.32 (m, 2H) , 3.86 (s, 2H) , 3.84 –3.79 (m, 2H) , 3.73 –3.68 (m, 2H) , 3.63 –3.57 (m, 2H) , 3.45 –3.37 (m, 2H) , 3.36 –3.29 (m, 2H) , 3.27 –3.16 (m, 4H) , 3.12 –3.01 (m, 4H) , 2.92 –2.83 (m, 2H) , 2.63 –2.55 (m, 1H) , 2.41 –2.27 (m, 4H) , 2.15 –1.80 (m, 9H) , 1.63 –1.43 (m, 10H) , 1.26 –1.12 (m, 2H) .

P-006

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 11.11 (s, 1H) , 8.37 (s, 1H) , 8.28 –8.13 (m, 3H) , 7.92 (d, J = 2.2 Hz, 1H) , 7.83 –7.75 (m, 1H) , 7.65 –7.56 (m, 3H) , 7.52 (d, J = 8.5 Hz, 1H) , 7.45 (d, J = 7.2 Hz, 1H) , 7.40 (s, 1H) , 7.31 –7.22 (m, 2H) , 7.13 –7.04 (m, 2H) , 6.74 (d, J = 9.1 Hz, 1H) , 6.66 (d, J = 7.1 Hz, 1H) , 6.29 (s, 2H) , 5.08 (dd, J = 12.5, 5.3 Hz, 1H) , 4.35 –4.30 (m, 2H) , 3.85 (s, 2H) , 3.82 –3.77 (m, 2H) , 3.66 –3.60 (m, 2H) , 3.57 –3.49 (m, 8H) , 3.44 –3.31 (m, 6H) , 3.26 –3.20 (m, 4H) , 3.12 –3.02 (m, 4H) , 2.93 –2.82 (m, 2H) , 2.63 –2.53 (m, 2H) , 2.15 –1.82 (m, 12H) , 1.62 –1.43 (m, 10H) , 1.25 –1.15 (m, 2H) .

P-007

1H NMR (400 MHz, DMSO) δ 11.56 (s, 1H) , 11.09 (s, 1H) , 8.44 (s, 1H) , 8.33 (s, 1H) , 7.99 –7.84 (m, 2H) , 7.66 (d, J = 8.5 Hz, 1H) , 7.60 –7.53 (m, 2H) , 7.45 –7.25 (m, 3H) , 7.13 –6.99 (m, 4H) , 6.87 (s, 1H) , 6.67 (d, J = 6.6 Hz, 1H) , 6.52 (t, J = 5.7 Hz, 1H) , 6.32 (s, 1H) , 6.27 (s, 1H) , 5.05 (dd, J = 12.9, 5.3 Hz, 1H) , 3.44 –3.37 (m, 4H) , 3.15 –3.01 (m, 6H) , 2.92 –2.83 (m, 2H) , 2.64 –2.55 (m, 7H) , 2.17 –1.82 (m, 13H) , 1.69 –1.41 (m, 15H) , 1.35 –1.13 (m, 9H) .

P-008

1H NMR (400 MHz, DMSO) δ 11.72 (s, 2H) , 11.11 (s, 1H) , 9.34 (s, 1H) , 8.60 –8.52 (m, 2H) , 8.17 –8.00 (m, 2H) , 7.82 (d, J = 10.9 Hz, 1H) , 7.62 –7.49 (m, 4H) , 7.13 –6.98 (m, 4H) , 6.75 (s, 1H) , 6.53 (s, 1H) , 6.40 (s, 1H) , 6.27 (s, 1H) , 5.05 (dd, J = 12.8, 5.3 Hz, 1H) , 3.49 –3.05 (m, 12H) , 2.98 –2.79 (m, 6H) , 2.68 –2.56 (m, 4H) , 2.16 –1.98 (m, 7H) , 1.94 –1.83 (m, 3H) , 1.76 –1.45 (m, 14H) , 1.36 –1.18 (m, 13H) .

P-009

1H NMR (400 MHz, MeOD) δ 8.85 (s, 1H) , 8.59 (s, 1H) , 8.24 (s, 1H) , 7.96 (s, 1H) , 7.66 (t, J = 9.4 Hz, 2H) , 7.47 –7.35 (m, 6H) , 7.32 (d, J = 3.4 Hz, 1H) , 7.21 (dd, J = 6.2, 3.1 Hz, 1H) , 7.03 –6.96 (m, 2H) , 6.66 (t, J = 9.4 Hz, 2H) , 6.34 –6.26 (m, 2H) , 4.65 (s, 1H) , 4.61 –4.53 (m, 2H) , 4.52 –4.46 (m, 2H) , 4.39 –4.30 (m, 1H) , 3.93 –3.86 (m, 1H) , 3.81 –3.68 (m, 4H) , 3.61 –3.51 (m, 18H) , 3.46 –3.38 (m, 3H) , 3.20 –3.06 (m, 8H) , 2.59 –2.53 (m, 1H) , 2.45 (s, 3H) , 2.24 –2.12 (m, 4H) , 2.10 –1.97 (m, 4H) , 1.90 –1.69 (m, 5H) , 1.67 –1.56 (m, 4H) , 1.46 (s, 9H) , 1.35 –1.24 (m, 6H) , 1.05 –0.97 (m, 14H) .

P-010

1H NMR (500 MHz, MeOD) δ 10.07 (s, 1H) , 8.60 (s, 1H) , 8.28 (s, 2H) , 7.91 (s, 1H) , 7.74 (s, 1H) , 7.65 –7.44 (m, 8H) , 7.28 (s, 1H) , 7.09 –6.62 (m, 3H) , 4.82 –4.41 (m, 5H) , 4.36 –4.29 (m, 1H) , 3.97 –3.77 (m, 4H) , 3.71 –3.44 (m, 30H) , 3.35 (s, 2H) , 2.62 (s, 3H) , 2.35 –2.21 (m, 3H) , 2.12 –1.88 (m, 6H) , 1.87 –1.65 (m, 5H) , 1.62 –1.42 (m, 3H) , 1.36 –1.24 (m, 6H) , 1.04 (s, 9H) .

P-011

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 8.98 (s, 1H) , 8.57 (t, J = 5.7 Hz, 1H) , 8.41 –8.37 (m, 1H) , 8.31 –8.22 (m, 1H) , 8.19 –8.13 (m, 2H) , 7.96 –7.90 (m, 1H) , 7.63 –7.56 (m, 2H) , 7.45 –7.35 (m, 5H) , 7.31 –7.24 (m, 2H) , 7.14 –7.05 (m, 2H) , 6.81 –6.74 (m, 1H) , 6.66 (d, J = 8.1 Hz, 1H) , 6.33 –6.26 (m, 2H) , 4.55 (d, J = 9.4 Hz, 1H) , 4.47 –4.38 (m, 2H) , 4.35 (s, 1H) , 4.26 –4.17 (m, 1H) , 3.69 –3.56 (m, 6H) , 3.54 –3.46 (m, 5H) , 3.14 –3.00 (m, 5H) , 2.90 –2.67 (m, 5H) , 2.44 (s, 3H) , 2.42 –2.31 (m, 4H) , 2.18 –2.00 (m, 6H) , 1.95 –1.81 (m, 6H) , 1.72 –1.65 (m, 2H) , 1.62 –1.45 (m, 7H) , 1.26 –1.14 (m, 3H) , 0.93 (s, 9H) .

P-012

1H NMR (400 MHz, DMSO) δ 11.53 (s, 1H) , 8.98 (s, 1H) , 8.57 (t, J = 5.3 Hz, 1H) , 8.39 (s, 1H) , 8.27 (s, 1H) , 8.18 (s, 1H) , 7.96 –7.90 (m, 2H) , 7.64 –7.56 (m, 2H) , 7.46 –7.35 (m, 5H) , 7.30 –7.24 (m, 2H) , 7.13 –7.03 (m, 2H) , 6.77 (d, J = 9.0 Hz, 1H) , 6.66 (d, J = 8.5 Hz, 1H) , 6.29 (s, 2H) , 4.55 (d, J = 9.6 Hz, 1H) , 4.49 –4.39 (m, 2H) , 4.35 (s, 1H) , 4.27 –4.19 (m, 1H) , 3.72 –3.56 (m, 6H) , 3.53 –3.40 (m, 9H) , 3.36 –3.19 (m, 5H) , 3.12 –3.02 (m, 6H) , 2.91 –2.66 (m, 7H) , 2.44 (s, 3H) , 2.38 –2.31 (m, 2H) , 2.17 –1.98 (m, 5H) , 1.94 –1.82 (m, 5H) , 1.73 –1.44 (m, 9H) , 1.26 –1.14 (m, 2H) , 1.05 (s, 1H) , 0.93 (s, 9H)

P-013

1H NMR (400 MHz, DMSO) δ 11.57 (s, 1H) , 11.11 (s, 1H) , 8.43 (s, 1H) , 8.33 (s, 1H) , 8.14 (s, 1H) , 7.96 (s, 1H) , 7.66 (d, J = 8.5 Hz, 1H) , 7.62 –7.53 (m, 2H) , 7.46 –7.41 (m, 1H) , 7.39 –7.33 (m, 1H) , 7.28 (d, J = 5.9 Hz, 1H) , 7.16 –7.00 (m, 4H) , 6.85 (d, J = 9.4 Hz, 1H) , 6.74 –6.62 (m, 2H) , 6.30 (d, J = 20.7 Hz, 2H) , 5.05 (dd, J = 12.8, 5.1 Hz, 1H) , 3.58 –3.44 (m, 3H) , 3.28 –3.17 (m, 2H) , 3.14 –2.99 (m, 6H) , 2.94 –2.82 (m, 3H) , 2.77 –2.64 (m, 3H) , 2.64 –2.54 (m, 3H) , 2.47 –2.38 (m, 3H) , 2.18 –2.06 (m, 3H) , 2.05 –1.95 (m, 2H) , 1.95 –1.80 (m, 4H) , 1.72 –1.44 (m, 9H) , 1.28 –1.11 (m, 3H) .

P-014

1H NMR (400 MHz, DMSO) δ 11.56 (s, 1H) , 11.12 (s, 1H) , 8.42 (s, 1H) , 8.32 (s, 1H) , 8.13 (s, 1H) , 7.95 (d, J = 2.4 Hz, 1H) , 7.68 –7.52 (m, 3H) , 7.46 –7.42 (m, 1H) , 7.33 (s, 1H) , 7.30 –7.25 (m, 1H) , 7.12 –7.05 (m, 3H) , 6.97 (d, J = 5.6 Hz, 1H) , 6.89 –6.82 (m, 2H) , 6.67 (d, J = 7.3 Hz, 1H) , 6.29 (d, J = 19.2 Hz, 2H) , 5.08 (dd, J = 12.8, 5.3 Hz, 1H) , 3.95 (d, J = 5.3 Hz, 2H) , 3.23 (d, J = 5.7 Hz, 2H) , 3.17 –3.01 (m, 6H) , 2.94 –2.84 (m, 2H) , 2.67 –2.55 (m, 5H) , 2.16 –1.81 (m, 11H) , 1.66 –1.45 (m, 9H) , 1.30 –1.13 (m, 5H) .

P-015

1H NMR (400 MHz, DMSO) δ 11.56 (s, 1H) , 11.10 (s, 1H) , 8.43 (s, 1H) , 8.32 (s, 1H) , 7.96 (d, J = 2.5 Hz, 2H) , 7.65 (d, J = 8.3 Hz, 1H) , 7.61 –7.53 (m, 2H) , 7.45 –7.40 (m, 1H) , 7.34 (s, 1H) , 7.28 (dd, J = 7.3, 2.0 Hz, 1H) , 7.14 –7.00 (m, 4H) , 6.84 (d, J = 8.6 Hz, 1H) , 6.67 (dd, J = 8.9, 2.1 Hz, 1H) , 6.55 (t, J = 5.9 Hz, 1H) , 6.34 –6.25 (m, 2H) , 5.05 (dd, J = 12.8, 5.3 Hz, 1H) , 3.28 –3.19 (m, 2H) , 3.15 –3.01 (m, 6H) , 2.94 –2.83 (m, 2H) , 2.80 –2.65 (m, 3H) , 2.62 –2.54 (m, 4H) , 2.29 (t, J = 6.8 Hz, 1H) , 2.17 –1.98 (m, 7H) , 1.95 –1.82 (m, 4H) , 1.72 –1.65 (m, 2H) , 1.63 –1.48 (m, 11H) , 1.30 –1.16 (m, 4H) , 1.10 (d, J = 6.6 Hz, 2H) .

P-016

1H NMR (400 MHz, DMSO) δ 11.49 (s, 1H) , 11.11 (s, 1H) , 8.34 (d, J = 1.8 Hz, 1H) , 8.19 (t, J = 4.7 Hz, 1H) , 7.90 (d, J = 2.5 Hz, 1H) , 7.83 –7.76 (m, 1H) , 7.60 (d, J = 8.7 Hz, 1H) , 7.57 –7.48 (m, 2H) , 7.43 (d, J = 7.2 Hz, 1H) , 7.40 –7.37 (m, 1H) , 7.28 (d, J = 7.5 Hz, 1H) , 7.22 (d, J = 2.2 Hz, 1H) , 7.13 –7.04 (m, 2H) , 6.71 –6.63 (m, 2H) , 6.29 (d, J = 9.6 Hz, 2H) , 5.08 (dd, J = 12.9, 5.4 Hz, 1H) , 4.01 (d, J = 6.0 Hz, 2H) , 3.46 –3.38 (m, 2H) , 3.24 (d, J = 5.8 Hz, 2H) , 3.13 –3.02 (m, 5H) , 2.90 –2.83 (m, 1H) , 2.68 –2.56 (m, 2H) , 2.24 –1.98 (m, 10H) , 1.92 –1.67 (m, 10H) , 1.64 –1.43 (m, 10H) , 1.25 –1.16 (m, 2H) , 1.14 –1.03 (m, 2H) , 0.91 –0.81 (m, 2H) .

P-017

1H NMR (400 MHz, DMSO) δ 11.62 (s, 1H) , 11.11 (s, 1H) , 8.52 –8.38 (m, 2H) , 7.99 (s, 1H) , 7.72 (d, J = 8.4 Hz, 1H) , 7.62 –7.51 (m, 2H) , 7.48 –7.39 (m, 2H) , 7.28 (dd, J = 7.1, 2.1 Hz, 1H) , 7.14 –7.02 (m, 4H) , 6.99 –6.90 (m, 1H) , 6.68 (d, J = 8.7 Hz, 1H) , 6.63 (t, J = 6.3 Hz, 1H) , 6.35 (s, 1H) , 6.25 (s, 1H) , 5.06 (dd, J = 12.8, 5.4 Hz, 1H) , 3.46 –3.36 (m, 5H) , 3.23 (d, J = 6.0 Hz, 2H) , 3.15 –2.84 (m, 10H) , 2.65 –2.54 (m, 4H) , 2.15 –1.81 (m, 10H) , 1.76 –1.66 (m, 5H) , 1.64 –1.45 (m, 7H) , 1.22 –1.13 (m, 1H) .

P-018

1H NMR (500 MHz, DMSO) δ 11.74 –11.50 (m, 2H) , 11.11 (s, 1H) , 8.75 (s, 1H) , 8.60 –8.50 (m, 2H) , 8.05 (s, 1H) , 7.85 –7.79 (m, 2H) , 7.59 –7.49 (m, 4H) , 7.46 (d, J = 7.3 Hz, 1H) , 7.35 –7.19 (m, 1H) , 7.10 (d, J = 9.5 Hz, 2H) , 6.74 (s, 1H) , 6.39 (s, 1H) , 6.25 (s, 1H) , 5.08 (dd, J = 12.5, 5.2 Hz, 1H) , 4.13 (d, J = 6.0 Hz, 2H) , 3.49 –3.41 (m, 4H) , 3.40 –3.27 (m, 3H) , 3.23 –2.99 (m, 9H) , 2.95 –2.74 (m, 4H) , 2.68 –2.57 (m, 3H) , 2.11 –1.96 (m, 6H) , 1.94 –1.84 (m, 3H) , 1.80 –1.69 (m, 4H) , 1.69 –1.40 (m, 12H) .

P-019

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 8.98 (s, 1H) , 8.58 (t, J = 6.2 Hz, 1H) , 8.37 (s, 1H) , 8.24 (s, 1H) , 8.01 –7.88 (m, 2H) , 7.59 (d, J = 8.6 Hz, 2H) , 7.45 –7.35 (m, 5H) , 7.31 –7.22 (m, 2H) , 7.15 –7.04 (m, 2H) , 6.75 (d, J = 9.3 Hz, 1H) , 6.65 (d, J = 7.4 Hz, 1H) , 6.29 (s, 2H) , 4.56 (d, J = 9.4 Hz, 1H) , 4.49 –4.32 (m, 3H) , 4.27 –4.17 (m, 1H) , 3.74 –3.56 (m, 9H) , 3.45 –3.32 (m, 5H) , 3.24 (d, J = 5.3 Hz, 2H) , 3.13 –2.94 (m, 10H) , 2.44 (s, 3H) , 2.41 –2.27 (m, 6H) , 2.19 –1.80 (m, 11H) , 1.63 –1.57 (m, 3H) , 1.55 –1.44 (m, 5H) , 1.26 –1.14 (m, 3H) , 0.97 –0.88 (m, 10H) .

P-020

1H NMR (400 MHz, DMSO) δ 11.77 –11.57 (m, 2H) , 11.14 (s, 1H) , 9.50 (s, 1H) , 8.62 –8.48 (m, 2H) , 8.05 (d, J = 2.5 Hz, 1H) , 7.87 –7.79 (m, 2H) , 7.65 –7.45 (m, 9H) , 7.24 –6.99 (m, 3H) , 6.74 (d, J = 7.3 Hz, 1H) , 6.43 –6.36 (m, 1H) , 6.28 (s, 1H) , 5.41 (s, 2H) , 5.10 (dd, J = 12.7, 5.4 Hz, 1H) , 4.35 –4.07 (m, 4H) , 3.68 –3.41 (m, 4H) , 3.28 –2.55 (m, 15H) , 2.09 –1.55 (m, 15H) .

P-021

1H NMR (400 MHz, DMSO) δ 11.72 (s, 2H) , 11.13 (s, 1H) , 9.65 (s, 1H) , 8.62 –8.51 (m, 2H) , 8.05 (d, J = 2.6 Hz, 1H) , 7.89 –7.78 (m, 2H) , 7.64 –7.46 (m, 9H) , 7.42 –7.17 (m, 2H) , 7.09 (d, J = 9.5 Hz, 1H) , 6.75 (d, J = 9.4 Hz, 1H) , 6.39 (dd, J = 3.3, 1.8 Hz, 1H) , 6.29 (s, 1H) , 5.41 (s, 2H) , 5.10 (dd, J = 12.8, 5.3 Hz, 1H) , 4.43 –4.13 (m, 6H) , 3.51 –2.81 (m, 15H) , 2.65 –2.55 (m, 2H) , 2.13 –1.30 (m, 15H) .

P-022

1H NMR (400 MHz, DMSO) δ 11.58 (s, 1H) , 11.11 (s, 1H) , 8.47 (s, 1H) , 8.37 (s, 1H) , 7.98 (d, J = 2.4 Hz, 1H) , 7.87 –7.77 (m, 1H) , 7.70 (d, J = 8.6 Hz, 1H) , 7.54 (dd, J = 21.4, 8.7 Hz, 2H) , 7.46 (d, J = 7.4 Hz, 2H) , 7.39 (s, 1H) , 7.27 (dd, J = 7.0, 2.1 Hz, 1H) , 7.12 –7.04 (m, 2H) , 6.90 (d, J = 8.9 Hz, 1H) , 6.68 (d, J = 7.2 Hz, 1H) , 6.34 (s, 1H) , 6.26 (s, 1H) , 5.08 (dd, J = 12.7, 5.4 Hz, 1H) , 4.17 –4.08 (m, 2H) , 3.26 –2.84 (m, 16H) , 2.65 –2.55 (m, 2H) , 2.47 –2.28 (m, 4H) , 2.16 –1.90 (m, 10H) , 1.87 –1.79 (m, 4H) , 1.76 –1.70 (m, 2H) , 1.65 –1.55 (m, 3H) , 1.53 –1.43 (m, 3H) , 1.17 –1.06 (m, 2H) .

P-023

1H NMR (400 MHz, DMSO) δ 11.74 –11.63 (m, 2H) , 11.11 (s, 1H) , 8.59 –8.52 (m, 2H) , 8.06 (d, J = 2.6 Hz, 1H) , 7.85 –7.79 (m, 1H) , 7.61 –7.50 (m, 5H) , 7.35 –7.18 (m, 3H) , 7.11 –7.02 (m, 2H) , 6.76 (d, J = 7.9 Hz, 1H) , 6.40 (dd, J = 3.3, 1.8 Hz, 1H) , 6.30 (s, 1H) , 6.15 (d, J = 8.2 Hz, 1H) , 5.05 (dd, J = 12.8, 5.4 Hz, 1H) , 4.15 –3.87 (m, 5H) , 3.59 –3.12 (m, 9H) , 2.95 –2.83 (m, 2H) , 2.71 –2.55 (m, 4H) , 2.10 –1.90 (m, 7H) , 1.80 –1.23 (m, 19H) .

P-024

1H NMR (400 MHz, DMSO) δ 11.79 –11.65 (m, 2H) , 11.12 (s, 1H) , 8.63 –8.50 (m, 2H) , 8.07 (d, J = 2.3 Hz, 1H) , 7.83 (d, J = 9.1 Hz, 1H) , 7.63 –7.43 (m, 5H) , 7.28 (d, J = 29.4 Hz, 2H) , 7.14 –7.07 (m, 2H) , 7.02 (d, J = 7.0 Hz, 1H) , 6.77 (d, J = 8.6 Hz, 1H) , 6.60 (s, 1H) , 6.40 (dd, J = 3.0, 1.7 Hz, 1H) , 6.31 (s, 1H) , 5.06 (dd, J = 12.8, 5.3 Hz, 2H) , 4.90 –4.42 (m, 5H) , 3.48 –3.25 (m, 8H) , 2.96 –2.83 (m, 2H) , 2.66 –2.54 (m, 3H) , 2.08 –1.75 (m, 9H) , 1.68 –1.22 (m, 17H) , 1.15 –1.01 (m, 3H) .

P-025

1H NMR (400 MHz, DMSO) δ 11.51 (s, 1H) , 11.11 (s, 1H) , 8.40 (d, J = 1.9 Hz, 1H) , 8.27 (s, 1H) , 8.15 (s, 1H) , 7.93 (d, J = 2.5 Hz, 1H) , 7.86 –7.77 (m, 1H) , 7.65 –7.44 (m, 5H) , 7.43 –7.38 (m, 1H) , 7.31 –7.25 (m, 2H) , 7.13 –7.05 (m, 2H) , 6.80 (d, J = 9.3 Hz, 1H) , 6.66 (d, J = 6.6 Hz, 1H) , 6.33 –6.25 (m, 2H) , 5.13 –5.04 (m, 1H) , 4.26 (d, J = 6.2 Hz, 1H) , 4.12 (d, J = 4.9 Hz, 1H) , 3.14 –3.03 (m, 6H) , 3.01 –2.96 (m, 2H) , 2.93 –2.79 (m, 4H) , 2.71 –2.60 (m, 2H) , 2.30 –1.87 (m, 15H) , 1.80 –1.46 (m, 13H) , 1.26 –1.14 (m, 2H) .

P-026

1H NMR (400 MHz, DMSO) δ 11.71 (s, 1H) , 11.59 (s, 1H) , 11.11 (s, 1H) , 8.63 –8.48 (m, 2H) , 8.06 (s, 1H) , 7.86 –7.75 (m, 2H) , 7.57 –7.49 (m, 4H) , 7.46 –7.02 (m, 6H) , 6.73 (d, J = 8.7 Hz, 1H) , 6.39 (s, 1H) , 6.26 (s, 1H) , 5.08 (d, J = 10.1 Hz, 1H) , 4.02 (d, J = 4.5 Hz, 2H) , 3.35 –3.23 (m, 4H) , 3.18 –3.03 (m, 4H) , 2.95 –2.83 (m, 3H) , 2.67 –2.56 (m, 3H) , 2.09 –1.66 (m, 14H) , 1.63 –1.33 (m, 12H) , 1.26 –1.11 (m, 6H) .

P-027

1H NMR (400 MHz, DMSO) δ 11.71 (s, 1H) , 11.65 (s, 1H) , 11.10 (s, 1H) , 8.60 –8.51 (m, 2H) , 8.06 (d, J = 2.6 Hz, 1H) , 7.82 (dd, J = 9.2, 2.0 Hz, 1H) , 7.62 –7.49 (m, 5H) , 7.46 –7.02 (m, 6H) , 6.75 (d, J = 8.2 Hz, 1H) , 6.52 (d, J = 7.9 Hz, 1H) , 6.40 (dd, J = 3.3, 1.9 Hz, 1H) , 6.29 (s, 1H) , 5.07 (dd, J = 12.9, 5.4 Hz, 1H) , 3.49 –3.28 (m, 8H) , 3.19 –3.05 (m, 4H) , 2.93 –2.82 (m, 2H) , 2.75 –2.55 (m, 5H) , 2.20 –1.87 (m, 7H) , 1.84 –1.66 (m, 6H) , 1.63 –1.36 (m, 14H) .

P-028

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.66 (s, 1H) , 11.10 (s, 1H) , 8.59 –8.51 (m, 2H) , 8.06 (d, J = 2.6 Hz, 1H) , 7.84 –7.78 (m, 1H) , 7.62 –7.15 (m, 8H) , 7.12 –7.05 (m, 2H) , 7.02 (d, J = 7.0 Hz, 1H) , 6.75 (d, J = 9.2 Hz, 1H) , 6.60 (s, 1H) , 6.39 (dd, J = 3.3, 1.8 Hz, 1H) , 6.29 (s, 1H) , 5.05 (dd, J = 12.9, 5.4 Hz, 1H) , 3.86 –3.68 (m, 4H) , 3.51 –3.22 (m, 12H) , 3.16 –3.08 (m, 1H) , 2.93 –2.83 (m, 1H) , 2.73 –2.54 (m, 4H) , 2.10 –1.76 (m, 8H) , 1.69 –1.32 (m, 18H) .

P-029

1H NMR (400 MHz, DMSO) δ 11.68 (s, 1H) , 11.52 (s, 1H) , 11.11 (s, 1H) , 8.61 –8.44 (m, 2H) , 8.04 (s, 1H) , 7.85 –7.73 (m, 2H) , 7.61 –7.41 (m, 5H) , 7.27 (s, 1H) , 7.13 –6.99 (m, 3H) , 6.70 (d, J = 9.2 Hz, 1H) , 6.38 (s, 1H) , 6.23 (s, 1H) , 5.12 –5.03 (m, 1H) , 4.13 (d, J = 4.7 Hz, 2H) , 3.49 –3.40 (m, 2H) , 3.25 –3.01 (m, 8H) , 2.94 –2.83 (m, 1H) , 2.76 –2.56 (m, 3H) , 2.13 –2.00 (m, 5H) , 1.97 –1.88 (m, 4H) , 1.87 –1.81 (m, 1H) , 1.78 –1.70 (m, 2H) , 1.67 –1.54 (m, 8H) , 1.51 –1.37 (m, 6H) , 1.29 –1.12 (m, 5H) .

P-030

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.68 (s, 1H) , 11.13 (s, 1H) , 8.94 (s, 1H) , 8.61 –8.52 (m, 2H) , 8.06 (d, J = 2.6 Hz, 1H) , 7.82 (t, J = 7.9 Hz, 2H) , 7.59 –7.02 (m, 9H) , 6.76 (d, J = 9.3 Hz, 1H) , 6.42 –6.28 (m, 2H) , 5.08 (dd, J = 12.7, 5.4 Hz, 1H) , 4.12 –4.00 (m, 2H) , 3.50 –2.53 (m, 19H) , 2.09 –0.82 (m, 29H) .

P-031

1H NMR (400 MHz, DMSO) δ 11.78 (s, 1H) , 11.72 (s, 1H) , 10.20 (s, 1H) , 8.61 –8.50 (m, 2H) , 8.06 (d, J = 2.6 Hz, 1H) , 7.82 (dd, J = 9.2, 1.9 Hz, 1H) , 7.63 –7.34 (m, 6H) , 7.25 –7.02 (m, 4H) , 6.79 –6.60 (m, 2H) , 6.40 (dd, J = 3.2, 1.8 Hz, 1H) , 6.29 (s, 1H) , 5.13 (dd, J = 13.0, 5.4 Hz, 1H) , 3.50 –3.41 (m, 3H) , 3.38 –3.25 (m, 7H) , 3.12 –2.99 (m, 7H) , 2.93 –2.71 (m, 4H) , 2.64 –2.54 (m, 7H) , 2.10 –2.01 (m, 3H) , 1.95 –1.83 (m, 2H) , 1.80 –1.69 (m, 6H) , 1.67 –1.55 (m, 5H) , 1.48 –1.29 (m, 3H) .

P-032

1H NMR (400 MHz, DMSO) δ 11.73 (s, 1H) , 11.67 (s, 1H) , 11.07 (s, 1H) , 9.30 (s, 1H) , 8.61 –8.51 (m, 2H) , 8.06 (d, J = 2.2 Hz, 1H) , 7.83 (d, J = 8.8 Hz, 1H) , 7.61 –7.41 (m, 5H) , 7.36 –7.06 (m, 4H) , 6.97 (s, 1H) , 6.87 (d, J = 8.1 Hz, 1H) , 6.75 (d, J = 8.1 Hz, 1H) , 6.40 (s, 1H) , 6.29 (s, 1H) , 5.04 (dd, J = 12.9, 5.3 Hz, 2H) , 3.51 –3.29 (m, 6H) , 3.25 –3.18 (m, 3H) , 3.16 –3.02 (m, 5H) , 2.95 –2.74 (m, 4H) , 2.66 –2.54 (m, 4H) , 2.09 –1.85 (m, 6H) , 1.82 –1.54 (m, 12H) , 1.51 –1.29 (m, 3H) .

P-033

1H NMR (400 MHz, DMSO) δ 11.67 (s, 1H) , 11.10 (s, 1H) , 8.55 –8.46 (m, 2H) , 8.02 (d, J = 2.6 Hz, 1H) , 7.78 (d, J = 9.2 Hz, 1H) , 7.64 –7.57 (m, 1H) , 7.54 –7.44 (m, 3H) , 7.27 (dd, J = 7.0, 2.3 Hz, 1H) , 7.14 –7.01 (m, 5H) , 6.73 –6.66 (m, 2H) , 6.37 (dd, J = 3.3, 1.9 Hz, 1H) , 6.22 (d, J = 1.8 Hz, 1H) , 5.05 (dd, J = 12.7, 5.4 Hz, 2H) , 3.50 –3.37 (m, 6H) , 3.23 –3.00 (m, 9H) , 2.94 –2.82 (m, 2H) , 2.64 –2.54 (m, 3H) , 2.13 –2.01 (m, 4H) , 1.98 –1.81 (m, 5H) , 1.76 –1.43 (m, 8H) , 1.27 –1.13 (m, 5H) .

P-034

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 11.09 (s, 1H) , 8.36 (d, J = 1.9 Hz, 1H) , 8.24 (t, J = 5.3 Hz, 1H) , 8.21 –8.16 (m, 2H) , 7.91 (d, J = 2.6 Hz, 1H) , 7.65 (d, J = 8.5 Hz, 1H) , 7.62 –7.55 (m, 2H) , 7.42 –7.39 (m, 1H) , 7.33 –7.26 (m, 2H) , 7.26 –7.20 (m, 2H) , 7.13 –7.05 (m, 2H) , 6.74 (d, J = 9.3 Hz, 1H) , 6.65 (d, J = 8.8 Hz, 1H) , 6.32 –6.27 (m, 2H) , 5.07 (dd, J = 13.0, 5.4 Hz, 1H) , 4.03 (d, J = 12.5 Hz, 2H) , 3.46 –3.32 (m, 5H) , 3.24 (d, J = 5.7 Hz, 1H) , 3.07 –3.02 (m, 2H) , 2.98 –2.83 (m, 4H) , 2.77 –2.64 (m, 4H) , 2.62 –2.53 (m, 3H) , 2.16 –2.07 (m, 3H) , 2.04 –1.97 (m, 1H) , 1.96 –1.82 (m, 4H) , 1.76 –1.65 (m, 4H) , 1.63 –1.45 (m, 11H) , 1.26 –1.12 (m, 5H) .

P-035

1H NMR (400 MHz, DMSO) δ 11.74 (s, 1H) , 11.60 (s, 1H) , 11.10 (s, 1H) , 8.61 –8.53 (m, 2H) , 8.05 (d, J = 2.6 Hz, 1H) , 7.77 (dd, J = 35.3, 9.7 Hz, 2H) , 7.58 –7.43 (m, 5H) , 7.33 (d, J = 8.8 Hz, 2H) , 7.17 –7.08 (m, 3H) , 6.73 (d, J = 9.7 Hz, 1H) , 6.39 (d, J = 3.2 Hz, 1H) , 6.24 (s, 1H) , 5.09 (dd, J = 12.6, 5.4 Hz, 1H) , 3.50 –3.44 (m, 6H) , 3.41 –3.32 (m, 6H) , 3.24 –3.13 (m, 4H) , 3.11 –3.00 (m, 3H) , 2.96 –2.82 (m, 3H) , 2.68 –2.56 (m, 5H) , 2.36 –2.31 (m, 1H) , 2.11 –1.93 (m, 6H) , 1.91 –1.75 (m, 7H) , 1.69 –1.59 (m, 5H) , 1.31 –1.13 (m, 2H) .

P-036

1H NMR (400 MHz, DMSO) δ 11.74 (s, 1H) , 11.67 (s, 1H) , 11.10 (s, 1H) , 9.60 (s, 1H) , 8.62 –8.51 (m, 2H) , 8.06 (d, J = 2.5 Hz, 1H) , 7.83 (d, J = 9.3 Hz, 1H) , 7.67 (d, J = 8.5 Hz, 1H) , 7.59 –7.49 (m, 3H) , 7.46 –7.07 (m, 6H) , 6.75 (d, J = 8.4 Hz, 1H) , 6.43 –6.37 (m, 1H) , 6.28 (s, 1H) , 5.07 (dd, J = 13.0, 5.3 Hz, 1H) , 4.07 (d, J = 12.0 Hz, 4H) , 3.52 –3.27 (m, 8H) , 3.14 –2.76 (m, 10H) , 2.65 –2.53 (m, 2H) , 2.19 –1.16 (m, 23H) .

P-037

1H NMR (400 MHz, DMSO) δ 11.54 (s, 1H) , 11.12 (s, 1H) , 8.38 (s, 1H) , 8.28 (s, 1H) , 7.93 (d, J = 2.5 Hz, 1H) , 7.83 (d, J = 8.3 Hz, 1H) , 7.59 (d, J = 8.7 Hz, 2H) , 7.45 –7.24 (m, 6H) , 7.13 –7.03 (m, 2H) , 6.77 (d, J = 9.2 Hz, 1H) , 6.66 (d, J = 8.7 Hz, 1H) , 6.33 –6.24 (m, 2H) , 5.12 (dd, J = 12.8, 5.4 Hz, 1H) , 4.00 (d, J = 6.1 Hz, 2H) , 3.23 (d, J = 5.8 Hz, 2H) , 3.16 –3.01 (m, 6H) , 2.96 –2.82 (m, 2H) , 2.66 –2.53 (m, 4H) , 2.19 –1.45 (m, 28H) , 1.21 –0.82 (m, 6H) .

P-038

1H NMR (400 MHz, DMSO) δ 11.67 (s, 1H) , 11.51 (s, 1H) , 11.11 (s, 1H) , 8.56 –8.45 (m, 2H) , 8.03 (d, J = 2.3 Hz, 1H) , 7.76 (d, J = 8.7 Hz, 1H) , 7.65 –7.58 (m, 1H) , 7.55 –7.46 (m, 3H) , 7.32 –7.26 (m, 2H) , 7.13 –7.06 (m, 3H) , 7.04 –6.98 (m, 1H) , 6.77 (t, J = 6.8 Hz, 1H) , 6.70 (d, J = 7.2 Hz, 1H) , 6.37 (s, 1H) , 6.24 (s, 1H) , 5.07 (dd, J = 12.9, 5.3 Hz, 1H) , 4.04 –3.84 (m, 2H) , 3.45 –3.38 (m, 3H) , 3.23 (d, J = 6.1 Hz, 1H) , 3.18 –3.01 (m, 5H) , 2.94 –2.84 (m, 1H) , 2.76 (t, J = 13.9 Hz, 2H) , 2.64 –2.53 (m, 3H) , 2.46 –2.35 (m, 4H) , 2.15 –1.98 (m, 5H) , 1.96 –1.82 (m, 4H) , 1.64 –1.44 (m, 9H) , 1.21 –1.15 (m, 1H) .

P-039

1H NMR (400 MHz, DMSO) δ 11.59 (s, 1H) , 11.10 (s, 1H) , 8.50 –8.31 (m, 2H) , 8.14 (s, 1H) , 7.97 (d, J = 2.4 Hz, 1H) , 7.71 (d, J = 8.5 Hz, 1H) , 7.66 (d, J = 8.6 Hz, 1H) , 7.56 (d, J = 8.8 Hz, 1H) , 7.47 –7.42 (m, 1H) , 7.40 –7.33 (m, 2H) , 7.30 –7.23 (m, 2H) , 7.13 –7.04 (m, 2H) , 6.87 (d, J = 8.5 Hz, 1H) , 6.67 (d, J = 8.9 Hz, 1H) , 6.33 (s, 1H) , 6.27 (s, 1H) , 5.08 (dd, J = 12.9, 5.3 Hz, 1H) , 3.69 –3.42 (m, 14H) , 3.27 –3.02 (m, 11H) , 2.94 –2.53 (m, 9H) , 2.15 –1.81 (m, 10H) , 1.73 –1.15 (m, 12H) .

P-040

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 11.11 (s, 1H) , 8.37 (s, 1H) , 8.24 (s, 1H) , 8.17 (s, 1H) , 8.10 –7.99 (m, 2H) , 7.91 (s, 2H) , 7.62 –7.54 (m, 3H) , 7.40 (s, 1H) , 7.31 –7.23 (m, 2H) , 7.14 –6.99 (m, 4H) , 6.73 (d, J = 8.5 Hz, 1H) , 6.65 (d, J = 8.9 Hz, 1H) , 6.53 (t, J = 4.5 Hz, 1H) , 6.29 (s, 2H) , 5.06 (d, J = 12.1 Hz, 1H) , 4.17 (s, 1H) , 3.11 –2.71 (m, 5H) , 2.19 –1.95 (m, 8H) , 1.91 –1.79 (m, 6H) , 1.68 –1.44 (m, 15H) , 1.37 –1.11 (m, 16H) .

P-041

1H NMR (400 MHz, DMSO) δ 11.68 (s, 1H) , 11.50 (s, 1H) , 11.10 (s, 1H) , 8.57 –8.46 (m, 2H) , 8.04 (d, J = 2.5 Hz, 1H) , 7.78 (d, J = 8.5 Hz, 1H) , 7.62 –7.55 (m, 1H) , 7.50 (dd, J = 10.2, 5.9 Hz, 3H) , 7.28 (dd, J = 6.9, 2.3 Hz, 1H) , 7.17 (d, J = 8.6 Hz, 1H) , 7.11 –7.00 (m, 4H) , 6.78 –6.67 (m, 2H) , 6.38 (s, 1H) , 6.23 (s, 1H) , 5.06 (dd, J = 12.8, 5.3 Hz, 1H) , 3.50 (d, J = 6.6 Hz, 2H) , 3.45 –3.37 (m, 3H) , 3.22 (d, J = 6.0 Hz, 1H) , 3.18 –3.02 (m, 5H) , 2.94 –2.83 (m, 1H) , 2.80 –2.67 (m, 2H) , 2.63 –2.54 (m, 3H) , 2.47 –2.34 (m, 5H) , 2.30 –2.18 (m, 2H) , 2.14 –1.99 (m, 3H) , 1.98 –1.82 (m, 3H) , 1.62 –1.45 (m, 8H) , 1.27 –1.13 (m, 4H) .

P-042

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.62 (s, 1H) , 10.99 (s, 1H) , 8.85 (s, 1H) , 8.61 –8.51 (m, 2H) , 8.05 (d, J = 2.6 Hz, 1H) , 7.82 (dd, J = 9.1, 2.1 Hz, 1H) , 7.56 (d, J = 2.5 Hz, 1H) , 7.55 –7.49 (m, 2H) , 7.47 (d, J = 7.8 Hz, 1H) , 7.34 –7.07 (m, 5H) , 6.74 (d, J = 8.0 Hz, 1H) , 6.39 (dd, J = 3.3, 1.8 Hz, 1H) , 6.28 (s, 1H) , 5.13 (dd, J = 13.3, 5.1 Hz, 1H) , 4.37 (d, J = 17.4 Hz, 1H) , 4.22 (d, J = 17.4 Hz, 1H) , 3.94 (d, J = 6.0 Hz, 4H) , 3.50 –3.43 (m, 2H) , 3.40 –3.28 (m, 3H) , 3.24 –3.02 (m, 5H) , 2.97 –2.86 (m, 4H) , 2.83 –2.75 (m, 1H) , 2.68 –2.53 (m, 3H) , 2.47 –2.37 (m, 2H) , 2.08 –1.57 (m, 21H) , 1.20 –0.94 (m, 5H) .

P-043

1H NMR (400 MHz, DMSO) δ 11.61 (s, 1H) , 11.07 (s, 1H) , 8.48 (s, 1H) , 8.34 (d, J = 35.3 Hz, 2H) , 8.14 (s, 1H) , 7.99 (s, 1H) , 7.82 (t, J = 5.6 Hz, 1H) , 7.72 (d, J = 7.9 Hz, 1H) , 7.61 –7.52 (m, 2H) , 7.48 –7.39 (m, 2H) , 7.30 –7.23 (m, 2H) , 7.13 –7.04 (m, 2H) , 7.00 –6.86 (m, 3H) , 6.68 (d, J = 8.8 Hz, 1H) , 6.34 (s, 1H) , 6.25 (s, 1H) , 5.04 (dd, J = 12.8, 5.4 Hz, 1H) , 4.15 –3.89 (m, 2H) , 3.26 –2.96 (m, 12H) , 2.89 –2.55 (m, 9H) , 2.16 –1.81 (m, 9H) , 1.77 (s, 3H) , 1.74 –1.13 (m, 17H) .

P-044

1H NMR (400 MHz, DMSO) δ 11.55 (s, 1H) , 8.40 (s, 1H) , 8.30 (s, 1H) , 7.94 (d, J = 2.4 Hz, 1H) , 7.84 –7.77 (m, 1H) , 7.66 –7.39 (m, 5H) , 7.34 –7.25 (m, 2H) , 7.12 –7.03 (m, 2H) , 6.81 (d, J = 8.7 Hz, 1H) , 6.66 (d, J = 8.6 Hz, 1H) , 6.30 (d, J = 9.2 Hz, 2H) , 5.15 (dd, J = 13.0, 5.4 Hz, 1H) , 4.02 (d, J = 6.0 Hz, 2H) , 3.45 –3.40 (m, 2H) , 3.23 (d, J = 5.8 Hz, 2H) , 3.14 –3.05 (m, 5H) , 3.02 (s, 3H) , 2.97 –2.89 (m, 2H) , 2.80 –2.54 (m, 7H) , 2.17 –1.45 (m, 25H) , 1.24 –1.06 (m, 4H) , 1.02 –0.89 (m, 2H) .

P-045

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.66 (s, 1H) , 10.94 (s, 1H) , 9.62 (s, 1H) , 8.57 (d, J = 2.1 Hz, 1H) , 8.55 –8.46 (m, 2H) , 8.06 (d, J = 2.5 Hz, 1H) , 7.88 –7.74 (m, 4H) , 7.59 –7.49 (m, 4H) , 7.43 (d, J = 6.8 Hz, 2H) , 7.10 (d, J = 9.3 Hz, 1H) , 6.80 –6.63 (m, 4H) , 6.43 –6.38 (m, 1H) , 6.28 (s, 1H) , 5.04 (dd, J = 13.0, 4.5 Hz, 1H) , 4.28 (dd, J = 17.0, 4.6 Hz, 2H) , 4.14 (d, J = 16.8 Hz, 1H) , 3.85 –3.78 (m, 1H) , 3.52 –3.37 (m, 5H) , 3.33 –3.01 (m, 11H) , 2.95 –2.66 (m, 6H) , 2.63 –2.54 (m, 2H) , 2.40 –2.28 (m, 1H) , 2.10 –1.90 (m, 5H) , 1.88 –1.70 (m, 7H) , 1.68 –1.31 (m, 12H) .

P-046

1H NMR (400 MHz, DMSO) δ 11.71 (s, 1H) , 11.61 (s, 1H) , 11.03 (s, 1H) , 9.43 (s, 1H) , 8.78 (d, J = 6.2 Hz, 1H) , 8.57 (d, J = 2.1 Hz, 1H) , 8.56 –8.51 (m, 1H) , 8.24 –8.12 (m, 3H) , 8.05 (d, J = 2.5 Hz, 1H) , 7.83 (d, J = 9.0 Hz, 1H) , 7.56 (d, J = 2.5 Hz, 1H) , 7.54 –7.48 (m, 2H) , 7.39 –7.06 (m, 5H) , 6.97 –6.90 (m, 1H) , 6.81 (d, J = 7.9 Hz, 1H) , 6.74 (d, J = 8.2 Hz, 1H) , 6.39 (dd, J = 3.2, 1.8 Hz, 1H) , 6.27 (s, 1H) , 5.13 (dd, J = 13.2, 4.8 Hz, 1H) , 4.28 –4.07 (m, 2H) , 3.76 –3.68 (m, 2H) , 3.39 –3.30 (m, 3H) , 3.17 –3.06 (m, 5H) , 3.01 –2.77 (m, 6H) , 2.68 –2.56 (m, 4H) , 2.36 –2.25 (m, 2H) , 2.11 –1.98 (m, 4H) , 1.94 –1.55 (m, 17H) , 1.47 –1.38 (m, 3H) , 1.33 –1.20 (m, 3H) .

P-047

1H NMR (400 MHz, DMSO) δ 11.51 (s, 1H) , 11.15 (s, 1H) , 8.37 (s, 1H) , 8.23 (t, J = 4.9 Hz, 1H) , 7.91 (d, J = 2.5 Hz, 1H) , 7.89 –7.81 (m, 1H) , 7.62 –7.47 (m, 4H) , 7.42 –7.38 (m, 1H) , 7.30 –7.21 (m, 2H) , 7.14 –7.04 (m, 2H) , 6.73 (d, J = 9.6 Hz, 1H) , 6.65 (d, J = 8.8 Hz, 1H) , 6.29 (s, 2H) , 5.09 (dd, J = 12.6, 5.3 Hz, 1H) , 4.30 (s, 2H) , 3.35 –2.85 (m, 15H) , 2.68 –2.55 (m, 3H) , 2.44 –2.27 (m, 5H) , 2.17 –1.94 (m, 8H) , 1.90 –1.80 (m, 3H) , 1.78 –1.68 (m, 2H) , 1.64 –1.43 (m, 8H) , 1.24 –1.14 (m, 3H) .

P-048

1H NMR (400 MHz, DMSO) δ 11.50 (s, 1H) , 11.11 (s, 1H) , 8.35 (d, J = 2.1 Hz, 1H) , 8.24 –8.18 (m, 2H) , 7.92 –7.82 (m, 2H) , 7.63 –7.48 (m, 4H) , 7.41 –7.38 (m, 1H) , 7.28 (d, J = 7.5 Hz, 1H) , 7.22 (d, J = 2.7 Hz, 1H) , 7.14 –7.05 (m, 2H) , 6.68 (dd, J = 24.8, 9.1 Hz, 2H) , 6.29 (d, J = 7.0 Hz, 2H) , 5.10 (dd, J = 12.6, 5.4 Hz, 1H) , 4.34 (d, J = 30.8 Hz, 2H) , 3.29 –3.02 (m, 9H) , 2.95 –2.83 (m, 2H) , 2.67 –2.53 (m, 2H) , 2.46 –2.32 (m, 6H) , 2.18 –1.98 (m, 10H) , 1.91 –1.71 (m, 6H) , 1.63 –1.42 (m, 10H) .

P-049

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.61 (s, 1H) , 11.13 (s, 1H) , 9.95 (d, J = 47.0 Hz, 2H) , 8.62 –8.50 (m, 2H) , 8.05 (d, J = 2.5 Hz, 1H) , 7.90 –7.79 (m, 2H) , 7.62 –7.46 (m, 5H) , 7.35 (s, 1H) , 7.22 –7.05 (m, 3H) , 6.73 (d, J = 8.2 Hz, 1H) , 6.39 (dd, J = 3.2, 1.8 Hz, 1H) , 6.25 (s, 1H) , 5.10 (dd, J = 12.7, 5.4 Hz, 1H) , 4.33 (t, J = 5.2 Hz, 2H) , 3.40 –3.03 (m, 20H) , 2.98 –2.77 (m, 4H) , 2.68 –2.53 (m, 4H) , 2.26 –2.00 (m, 9H) , 1.97 –1.49 (m, 15H) , 1.32 –1.18 (m, 1H) .

P-050

1H NMR (400 MHz, DMSO) δ 11.70 (s, 1H) , 11.56 (s, 1H) , 11.11 (s, 1H) , 9.91 (s, 1H) , 8.58 –8.51 (m, 2H) , 8.05 (d, J = 2.5 Hz, 1H) , 7.81 (d, J = 8.9 Hz, 1H) , 7.64 –7.48 (m, 5H) , 7.26 –7.02 (m, 6H) , 6.76 –6.65 (m, 2H) , 6.41 –6.37 (m, 1H) , 6.25 (s, 1H) , 5.06 (dd, J = 12.8, 5.4 Hz, 1H) , 3.42 –3.33 (m, 7H) , 3.18 –2.84 (m, 12H) , 2.64 –2.55 (m, 3H) , 2.12 –1.97 (m, 9H) , 1.90 –1.70 (m, 9H) , 1.66 –1.57 (m, 3H) , 1.26 –1.21 (m, 1H) .

P-051

1H NMR (400 MHz, DMSO) δ 11.56 (s, 1H) , 11.08 (s, 1H) , 8.42 (s, 1H) , 8.30 (s, 1H) , 8.15 (s, 1H) , 7.95 (s, 1H) , 7.70 –7.54 (m, 3H) , 7.42 (d, J = 2.8 Hz, 1H) , 7.36 –7.22 (m, 4H) , 7.08 (q, J = 7.9 Hz, 2H) , 6.83 (d, J = 9.5 Hz, 1H) , 6.67 (d, J = 6.8 Hz, 1H) , 6.29 (d, J = 18.2 Hz, 2H) , 5.07 (dd, J = 12.9, 5.4 Hz, 1H) , 3.47 –3.38 (m, 5H) , 3.24 (d, J = 5.7 Hz, 2H) , 3.15 –2.84 (m, 7H) , 2.66 –2.55 (m, 3H) , 2.41 –2.32 (m, 3H) , 2.16 –1.70 (m, 10H) , 1.63 –1.14 (m, 16H) , 1.04 –0.85 (m, 3H) .

P-052

1H NMR (400 MHz, DMSO) δ 11.57 (s, 1H) , 11.15 (s, 1H) , 8.46 (s, 1H) , 8.31 (s, 1H) , 7.98 (s, 1H) , 7.84 (d, J = 8.1 Hz, 1H) , 7.65 (dd, J = 24.1, 8.2 Hz, 2H) , 7.44 (s, 2H) , 7.35 (s, 2H) , 7.26 (s, 1H) , 7.06 (s, 3H) , 6.83 (d, J = 8.6 Hz, 1H) , 6.66 (d, J = 7.8 Hz, 1H) , 6.30 (d, J = 12.3 Hz, 2H) , 5.15 (dd, J = 12.7, 5.2 Hz, 1H) , 4.12 (s, 2H) , 3.86 (s, 1H) , 3.46 –3.32 (m, 4H) , 3.23 –3.16 (m, 1H) , 3.13 –3.01 (m, 5H) , 2.97 –2.76 (m, 5H) , 2.67 –2.40 (m, 6H) , 2.15 –2.01 (m, 4H) , 1.99 –1.77 (m, 6H) , 1.74 –1.51 (m, 7H) , 1.39 (s, 9H) , 1.29 –1.11 (m, 2H) .

P-053

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.64 (s, 1H) , 11.14 (s, 1H) , 9.60 (s, 1H) , 8.63 –8.53 (m, 2H) , 8.37 (s, 2H) , 8.05 (d, J = 2.6 Hz, 1H) , 7.93 (d, J = 8.2 Hz, 1H) , 7.87 –7.79 (m, 1H) , 7.60 –7.48 (m, 4H) , 7.41 (dd, J = 8.3, 2.1 Hz, 1H) , 7.10 (d, J = 9.4 Hz, 1H) , 6.74 (d, J = 7.6 Hz, 1H) , 6.39 (dd, J = 3.3, 1.9 Hz, 1H) , 6.28 (s, 1H) , 5.14 (dd, J = 12.8, 5.3 Hz, 1H) , 4.44 –4.37 (m, 2H) , 4.28 –4.21 (m, 2H) , 3.78 –3.68 (m, 2H) , 3.51 –3.44 (m, 3H) , 3.42 –3.31 (m, 3H) , 3.28 –3.02 (m, 8H) , 2.97 –2.78 (m, 4H) , 2.67 –2.54 (m, 4H) , 2.18 –2.03 (m, 6H) , 1.95 –1.85 (m, 3H) , 1.83 –1.58 (m, 8H) , 1.55 –1.42 (m, 1H) .

P-054

1H NMR (400 MHz, DMSO) δ 11.71 (s, 1H) , 11.64 (s, 1H) , 11.11 (s, 1H) , 9.52 (s, 1H) , 8.61 –8.48 (m, 2H) , 8.06 (d, J = 2.5 Hz, 1H) , 7.85 –7.74 (m, 2H) , 7.59 –7.16 (m, 8H) , 7.08 (d, J = 9.3 Hz, 1H) , 6.75 (d, J = 7.5 Hz, 1H) , 6.40 (d, J = 1.7 Hz, 1H) , 6.28 (s, 1H) , 5.10 (dd, J = 13.0, 5.4 Hz, 1H) , 4.24 –4.15 (m, 4H) , 3.62 –3.27 (m, 9H) , 3.16 –2.85 (m, 10H) , 2.64 –2.55 (m, 2H) , 2.11 –1.72 (m, 9H) , 1.68 –1.40 (m, 7H) , 1.37 –1.20 (m, 3H) , 1.11 –0.90 (m, 3H) .

P-055

1H NMR (400 MHz, DMSO) δ 11.49 (s, 1H) , 11.13 (s, 1H) , 8.38 –8.24 (m, 3H) , 8.18 –8.08 (m, 2H) , 7.90 (d, J = 2.5 Hz, 1H) , 7.82 –7.74 (m, 1H) , 7.64 –7.52 (m, 2H) , 7.50 –7.43 (m, 2H) , 7.40 –7.36 (m, 1H) , 7.28 (d, J = 7.5 Hz, 1H) , 7.22 (d, J = 2.5 Hz, 1H) , 7.14 –7.05 (m, 2H) , 6.72 –6.61 (m, 2H) , 6.29 (dd, J = 12.4, 2.6 Hz, 2H) , 5.10 (dd, J = 12.7, 5.5 Hz, 1H) , 4.87 –4.77 (m, 2H) , 4.58 (d, J = 4.5 Hz, 2H) , 3.32 –3.22 (m, 5H) , 3.15 –3.02 (m, 6H) , 2.96 –2.85 (m, 1H) , 2.64 –2.54 (m, 2H) , 2.32 –1.95 (m, 7H) , 1.90 –1.82 (m, 3H) , 1.80 –1.72 (m, 3H) , 1.64 –1.36 (m, 9H) , 1.26 –1.14 (m, 2H) .

P-056

1H NMR (400 MHz, DMSO) δ 11.55 (s, 1H) , 11.12 (s, 1H) , 8.41 (d, J = 1.9 Hz, 1H) , 8.31 (t, J = 4.3 Hz, 1H) , 8.15 (s, 1H) , 8.06 (d, J = 6.8 Hz, 1H) , 7.95 (d, J = 2.5 Hz, 1H) , 7.85 (d, J = 8.3 Hz, 1H) , 7.63 (d, J = 9.5 Hz, 1H) , 7.58 (d, J = 8.7 Hz, 1H) , 7.46 (d, J = 1.9 Hz, 1H) , 7.44 –7.40 (m, 1H) , 7.36 (dd, J = 8.3, 2.1 Hz, 1H) , 7.32 (d, J = 2.0 Hz, 1H) , 7.28 (dd, J = 7.3, 1.9 Hz, 1H) , 7.12 –7.03 (m, 2H) , 6.82 (d, J = 9.3 Hz, 1H) , 6.68 –6.63 (m, 1H) , 6.33 –6.24 (m, 2H) , 5.13 (dd, J = 12.9, 5.3 Hz, 1H) , 4.19 –4.07 (m, 3H) , 3.45 –3.34 (m, 5H) , 3.23 (d, J = 5.9 Hz, 2H) , 3.13 –3.01 (m, 6H) , 2.94 –2.73 (m, 6H) , 2.65 –2.54 (m, 3H) , 2.16 –2.04 (m, 4H) , 1.99 –1.89 (m, 4H) , 1.85 (s, 3H) , 1.74 –1.44 (m, 9H) , 1.26 –1.12 (m, 1H) .

P-057

1H NMR (400 MHz, DMSO) δ 11.70 (s, 1H) , 11.62 –11.53 (m, 1H) , 11.09 (s, 1H) , 9.14 (s, 1H) , 8.62 –8.49 (m, 2H) , 8.05 (s, 1H) , 7.82 (d, J = 9.1 Hz, 1H) , 7.71 (d, J = 8.5 Hz, 1H) , 7.61 –7.48 (m, 3H) , 7.40 (s, 1H) , 7.31 (d, J = 8.7 Hz, 2H) , 7.10 (d, J = 9.3 Hz, 2H) , 6.74 (s, 1H) , 6.39 (s, 1H) , 6.25 (s, 1H) , 5.12 –5.04 (m, 1H) , 4.23 (d, J = 12.3 Hz, 2H) , 3.55 –3.43 (m, 4H) , 3.41 –3.30 (m, 3H) , 3.22 –3.05 (m, 5H) , 3.03 –2.78 (m, 7H) , 2.66 –2.55 (m, 4H) , 2.17 –1.96 (m, 8H) , 1.91 –1.78 (m, 4H) , 1.73 –1.54 (m, 8H) .

P-058

1H NMR (400 MHz, DMSO) δ 11.72 (s, 1H) , 11.63 (s, 1H) , 11.10 (s, 1H) , 9.77 (s, 1H) , 8.62 –8.49 (m, 2H) , 8.05 (d, J = 2.6 Hz, 1H) , 7.83 (dd, J = 9.1, 2.1 Hz, 1H) , 7.73 (d, J = 8.2 Hz, 1H) , 7.58 –7.50 (m, 3H) , 7.30 –7.06 (m, 3H) , 6.88 (s, 1H) , 6.78 –6.70 (m, 2H) , 6.43 –6.37 (m, 1H) , 6.28 (s, 1H) , 5.12 –5.03 (m, 1H) , 4.49 –4.04 (m, 9H) , 3.61 –3.33 (m, 8H) , 3.28 –3.05 (m, 5H) , 3.00 –2.74 (m, 6H) , 2.67 –2.54 (m, 4H) , 2.36 –2.27 (m, 2H) , 2.10 –1.95 (m, 5H) , 1.92 –1.57 (m, 12H) .

P-059

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 10.99 (s, 1H) , 8.37 (s, 1H) , 8.27 (s, 1H) , 8.15 (s, 1H) , 7.92 (d, J = 2.6 Hz, 1H) , 7.59 (d, J = 8.8 Hz, 2H) , 7.47 (t, J = 7.9 Hz, 1H) , 7.41 (t, J = 2.9 Hz, 1H) , 7.32 –7.21 (m, 4H) , 7.13 –7.05 (m, 2H) , 6.80 –6.74 (m, 1H) , 6.65 (d, J = 8.8 Hz, 1H) , 6.29 (s, 2H) , 5.12 (dd, J = 13.1, 4.7 Hz, 1H) , 4.37 (d, J = 17.3 Hz, 1H) , 4.22 (d, J = 17.7 Hz, 1H) , 4.01 (d, J = 6.7 Hz, 2H) , 3.45 –3.38 (m, 2H) , 3.24 (d, J = 5.3 Hz, 1H) , 3.14 –3.01 (m, 5H) , 2.97 –2.85 (m, 1H) , 2.70 –2.55 (m, 5H) , 2.44 –2.33 (m, 2H) , 2.28 –2.19 (m, 1H) , 2.16 –1.45 (m, 25H) , 1.37 –1.12 (m, 3H) , 1.04 –0.95 (m, 1H) .

P-060

1H NMR (400 MHz, DMSO) δ 11.52 (s, 1H) , 11.02 (s, 1H) , 8.36 (s, 1H) , 8.28 –8.14 (m, 2H) , 7.92 (s, 1H) , 7.63 –7.52 (m, 2H) , 7.40 (s, 1H) , 7.32 –7.21 (m, 3H) , 7.14 –7.04 (m, 2H) , 6.90 (d, J = 7.4 Hz, 1H) , 6.77 –6.61 (m, 3H) , 6.29 (s, 2H) , 5.63 (s, 1H) , 5.11 (dd, J = 13.3, 5.0 Hz, 1H) , 4.17 (dd, J = 45.3, 17.3 Hz, 2H) , 3.44 –3.39 (m, 1H) , 3.24 (d, J = 6.0 Hz, 2H) , 3.14 –2.88 (m, 9H) , 2.67 –2.56 (m, 2H) , 2.36 –2.24 (m, 3H) , 2.17 –1.74 (m, 14H) , 1.68 –1.34 (m, 13H) , 1.25 –1.15 (m, 2H) , 1.01 –0.77 (m, 4H) .

P-061

1H NMR (400 MHz, DMSO) δ 11.58 (s, 1H) , 11.09 (s, 1H) , 8.43 (s, 1H) , 8.33 (s, 1H) , 8.15 (s, 1H) , 7.96 (s, 1H) , 7.64 (d, J = 8.2 Hz, 2H) , 7.56 (d, J = 8.7 Hz, 1H) , 7.43 (s, 1H) , 7.37 –7.25 (m, 2H) , 7.13 –7.04 (m, 2H) , 6.90 –6.74 (m, 2H) , 6.66 (t, J = 9.6 Hz, 2H) , 6.29 (d, J = 19.5 Hz, 2H) , 5.06 (dd, J = 12.9, 5.3 Hz, 1H) , 4.03 (t, J = 7.7 Hz, 2H) , 3.85 –3.55 (m, 5H) , 3.27 –2.82 (m, 12H) , 2.63 –2.53 (m, 2H) , 2.39 –1.80 (m, 14H) , 1.65 –1.41 (m, 9H) , 1.26 –1.12 (m, 2H) .

Example 3. Biological Assays for Drug Moieties and PROTAC compounds

The HTRF BCL-2/BAK or BCL-XL/BAK assay from Cisbio (63ADK000CB01PEG; 63ADK000CB04PEG) was used to test compounds disclosed herein for blocking of BCL-2 or BCL-XL protein with its ligand, BAK. Recombinant human 2nM Tag1-BCL-2, 2nM Tag1-BCL-XL protein, 10nM Tag2-BAK/5nM Tag2-BAK (corresponding to BCL-2 and BCL-XL assay, respectively) were pre-incubated with a serial dilution of compounds disclosed herein (maximum concentration and dilution ratio determined by the results of pre-experiment may vary) at room temperature for 15 minutes in an assay buffer from BCL-2/BAK or BCL-XL/BAK assay kits, respectively. Then the pre-mixed anti-Tag1-Eu³⁺ and anti-tag2-XL665 were added to the plate and further incubated at room temperature for another 2 hours. The signals (665nM, 615nM) were read on Envision 2104 instrument. The IC₅₀ for each compound was derived from fitting the signal of 665/615nM to the increasing compound concentration.

Using the above assays, the drug moieties and PROTAC compounds provided herein and a control compound (Venetoclax) were tested. In Table 2 below, for IC₅₀ data, “***” means the compound had an IC₅₀ of greater than zero but less than or equal to about 20nM; “**” means the compound had an IC₅₀ of greater than about 20nM but less than or equal to about 200nM; “*” means the compound had an IC₅₀ of greater than about 200nM but less than or equal to about 2000nM; “-” means the compound had an IC₅₀ of greater than about 2000nM.

Table 2. IC₅₀ data of Exemplary Drug Moieties and PROTAC compounds

Example 4. CYP2C9 Activity Assay

In this assay, the drug moieties provided herein and a control compound (Venetoclax) were used as the test compounds. In general, the assay was conducted by the following steps:

1. The solution of test compound, human liver microsomes solution, and substrate (diclofenac) solution were mixed in a 96-well assay plate on ice to, and the final concentration of each test compound is 1 μM;

2. The 96-well assay plate and NADPH solution were pre-incubated at 37℃ for 5 minutes;

3. NADPH solution was added to into the assay plates to initiate the reaction;

4. The assay plate was incubated at 37 ℃ for 10 min;

5. The reaction was stopped; and after quenching, a portion of the supernatant was taken from each well for LC/MS analysis;

6. The inhibition rate was calculated by the following equation: Inhibition rate (%) = (1 –Value_test/Value_control) x 100%, where Value_test refers to the LC/MS data obtained from wells with test compounds and Value_control refers to the LC/MS data obtained from wells without test compound.

The inhibition rates of several test compounds were shown in Table 3 below.

Table 3. Inhibition Rate (%) of Exemplary Drug Moieties against CYP2C9 Enzyme

As demonstrated in table above, the drug moieties of the present disclosure showed a significantly decreased inhibition rate compared to the reference compound Venetoclax.

Example 5. Efficacy Study

Cell Proliferation Assay for Drug Moieties

TheLuminescent Cell Viability Assay (Promega, G7573) was used to study the cellular potency of disclosed compounds herein. The cells were harvested during the logarithmic growth period and counted with hemocytometer. The DOHH2 cells were seeded at 1.6*10⁴ in 90ul DMEM medium supplemented with 10%fetal bovine serum (FBS) (as RS4; 11 cells were seeded at 4000 in 90ul RPMI-1640 medium with 10%FBS) per well in 96-well plates and treated with 10ul of a serial dilution of the drug moieties provided herein (maximum concentration and dilution ratio determined by the results of pre-experiment may vary) for 72 hours in a 5%CO₂ incubator at 37℃. Cell viability was assessed according to the manufacturer’s recommendations. After the plates return to the room temperature, 100ul of reagent was added to 100ul of cell culture. The mixture was agitated on an orbital shaker for 2 minutes or placed it at room temperature for 10 minutes to allow cell lysis and stabilization of luminescent signals. Luminescent signals were recorded using Envision 2104 instrument. And GI₅₀ values were then calculated.

The GI₅₀ values of the tested compounds are shown in Table 4A below. In Table 4A below, for GI₅₀ data, “***” means the compound had an GI₅₀ of greater than zero but less than or equal to about 50nM; “**” means the compound had an GI₅₀ of greater than about 50nM but less than or equal to about 500nM; “*” means the compound had an GI₅₀ of greater than about 500nM but less than or equal to about 5000nM; “-” means the compound had an GI₅₀ of greater than about 5000nM.

Table 4A. GI₅₀ Data of Exemplary Drug Moieties

Cell Proliferation Assay for PROTAC compounds

TheLuminescent Cell Viability Assay (Promega, G7573) was used to study the cellular potency of disclosed compounds herein. The cells were harvested during the logarithmic growth period and counted with hemocytometer. Molt-4 cells were seeded at 5*10³ in 90ml PRMI 1640 medium supplemented with 10%fetal bovine serum (FBS) per well in 96-well plates and treated with 10μl of a serial dilution of the drug moieties provided herein (maximum concentration and dilution ratio determined by the results of pre-experiment may vary) for 72 hours in a 5%CO₂ incubator at 37℃. Cell viability was assessed according to the manufacturer’s recommendations. After the plates return to the room temperature, 40ml of reagent was added to 100ml of cell culture. The mixture was agitated on an orbital shaker for 2 minutes or placed it at room temperature for 10 minutes to allow cell lysis and stabilization of luminescent signals. Luminescent signals were recorded using Envision 2104 instrument. And GI₅₀ values were then calculated.

The GI₅₀ values of the tested compounds are shown in Table 4B below. In Table 4B below, for GI₅₀ data, “***” means the compound had an GI₅₀ of greater than zero but less than or equal to about 50nM; “**” means the compound had an GI₅₀ of greater than about 50nM but less than or equal to about 500nM; “*” means the compound had an GI₅₀ of greater than about 500nM.

Table 4B. GI₅₀ Data of Exemplary PROTAC compounds

In vivo Pharmacodynamic Study

In general, NOD/SCID RS4; 11 subcutaneous xenograft tumor model was established by inoculating 5*10⁶/0.1ml/mouse subcutaneously in the right back of the NOD/SCID female mouse. The animals were checked daily for any effects of treatments on behaviors such as mobility, food and water consumption, body weight gain/loss, eyes, hairs and any other abnormalities. Mortality and clinical signs observed during the study were recorded in the raw data. Animal weight and tumor size were measured every two days during the study. Tumor volume (TV) was calculated as: TV=0.5*a*b², wherein a and b represent the measured length and width of tumor, respectively. Relative tumor proliferation inhibition rate (TGI_RTV (%) ) , as an indication of anti-tumor effectiveness, was calculated as: TGI_RTV (%) = (1-T_RTV/C_RTV) *100%, wherein T_RTV and C_RTV were relative tumor volume (RTV) in treatment group and vehicle control group, respectively. RTV was calculated as: RTV = V_t/V₀, wherein V_t and V₀ represented the tumor volume measured on day t after dosing and on the first day of dosing. At the end of last dosing, plasma and tumor tissue were collected, weighed, and photographed according to requirements of the study protocol. Similarly, a MOLT4 mice xenograft model and an H1963 mice xenograft model were used.

Compared to the vehicle group, the drug treatment groups showed anti-tumor proliferation effect. The TGI_RTV (%) values after 10 or 20 days dosing were shown in Table 5 below, where “+++ ” represented TGI_RTV (%) ≥ 80%; “++” represented 30%<TGI_RTV (%) <80%. p.o. stands for “per os” ; I.P. stands for “intraperitoneal” ; q.d. stands for “once daily” ; q3d stands for “every 3 days” .

Table 5. TGI_RTV of Exemplary Drug Moieties and PROTAC compounds

As shown in Table 5 above, the drug moieties and PROTAC compounds of the present disclosure showed potent inhibition of tumor growth.

Example 6. Degradation Assays for PROTAC compounds

The ability of the PROTACs to degrade endogenous BCL-XL was assessed using the conventional western blot assays.

The cells were harvested during the logarithmic growth period and counted with hemocytometer. Molt4 cells (30000 cells/well) were treated with DMSO and compounds at indicated concentrations for 16 h. Cells were harvested and washed with PBS, and then were lysed in RIPA buffer (50mM Tris (pH 7.4) , 150mM NaCl, 1%NP-40, 1%sodium deoxycholate, 0.1%SDS) supplemented with 1%protease inhibitor cocktail, followed by centrifugation at 14,000xg for 10 min at 4℃. Supernatant was removed and protein concentration were determined by BCA protein assay kit. Cell lysates (20-30 μg) were subjected to SDS-PAGE and the expression levels of indicated protein were examined by Western blot analysis. The immunoblots were quantified by densitometry using e-blot software and the data were expressed as relative band intensities normalized to equal loading control. And DC₅₀ values were calculated with Graphpad Prism software.

The DC₅₀ values of the tested PROTAC compounds are shown in Table 2 below, in which , for DC₅₀ data, “++++” represents DC₅₀<15nM; “+++” represents 15nM≤DC_50≤100nM; “++” represents DC₅₀>100nM.

Table 6. Degradation Ability of Exemplary PROTAC Compounds

The foregoing description is considered as illustrative only of the principles of the present disclosure. Further, since numerous modifications and changes will be readily apparent to those skilled in the art, it is not desired to limit the invention to the exact construction and process shown as described above. Accordingly, all suitable modifications and equivalents maybe considered to fall within the scope of the invention as defined by the claims that follow.

The words “comprise, ” “comprising, ” “include, ” “including, ” and “includes” when used in this specification and in the following claims are intended to specify the presence of stated features, integers, components, or steps, but they do not preclude the presence or addition of one or more other features, integers, components, steps, or groups thereof.

Claims

A compound of Formula I:
D-L-E (I) ,

or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein,

E is a degradation signaling moiety;

L is a linker that covalently attaches E to D; and

D is a BCL-XL modulator moiety of Formula I-a

wherein Ring A is cyclohexyl or 6-membered heterocyclyl.
The compound of claim 1, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein Ring A is piperidinyl.
The compound of claim 2, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein Ring A is selected from
The compound of claim 1, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein E is an E3 ligase recognition agent.
The compound of claim 4, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein E is a VHL ligand or a CRBN ligand.
The compound of claim 5, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein E is
The compound of claim 5, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein E is

wherein

R^a is selected from hydrogen or alkyl; and

each R^b is independently hydrogen or alkyl, or two R^b together with the carbon atom to which they are attached form -C (O) -.
The compound of claim 7, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R^a is selected from hydrogen or alkyl.
The compound of claim 8, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R^a is selected from hydrogen or methyl.
The compound of claim 7, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein both R^b are hydrogen.
The compound of claim 7, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein two R^b together with the carbon atom to which they are attached form -C (O) -.
The compound of claim 7, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein E is selected from
The compound of claim 1, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L is a linker of Formula (I-b) :
-L¹-L²-L³-L⁴- (I-b) ;

wherein,

L¹ is selected from the group consisting of a bond, - (CH₂) _pC (O) -, alkyl, alkenyl and alkynyl, wherein the alkyl, alkenyl and alkynyl is optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano, nitro or -N (R²) ₂;

L² is selected from the group consisting of a bond, -NH-C (O) -C (R¹) ₂-*, -N (R²) -*, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclyl and heteroaryl, wherein the alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclyl and heteroaryl is optionally substituted with one or more groups independently selected from halogen, oxo, hydroxyl, cyano, nitro or -N (R²) ₂, and *end of L² is connected to L³;

L³ is selected from the group consisting of a bond, - (OCH₂CH₂) _m-#, - (CH₂CH₂O) _m (CH₂) _n-#, alkyl, alkenyl and alkynyl, and #end of L³ is connected to L⁴;

L⁴ is selected from the group consisting of a bond, -N (R²) -, -O-, -C (O) -, cycloalkyl and heterocyclyl;

each R¹ is independently selected from the group consisting of hydrogen, halogen, hydroxyl, cyano, nitro, alkyl and - (CH₂) _pN (R³) ₂;

each R² is independently selected from the group consisting of hydrogen, -C (O) R⁴, alkyl, alkenyl and alkynyl wherein the alkyl, alkenyl and alkynyl is optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano or nitro;

each R³ is independently selected from hydrogen or -C (O) R⁴;

each R⁴ is independently selected from -O-alkyl or alkyl;

m is 0, 1, 2, 3, 4, 5, 6, 7 or 8;

n is 0, 1, 2 or 3; and

p is 0, 1, 2, 3 or 4.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L¹ is a bond.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L¹ is alkyl optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano, nitro or -N (R²) ₂.
The compound of claim 15, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L¹ is linear C_1-6 alkyl.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L¹ is - (CH₂) _pC (O) -.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is a bond.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is -NH-C (O) -C (R¹) ₂-*.
The compound of claim 19, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein each R¹ is independently selected from hydrogen, alkyl or - (CH₂) _pN (R³) ₂.
The compound of claim 20, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein one R¹ is hydrogen and the other R¹ is - (CH₂) _pN (R³) ₂.
The compound of claim 21, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein both R³ are hydrogen.
The compound of claim 21, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein one R³ is hydrogen, the other R³ is -C (O) R⁴, wherein R⁴ is alkyl.
The compound of claim 23, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R⁴ is methyl.
The compound of claim 19, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is selected from the group consisting of:
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is -N (R²) -.
The compound of claim 26, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R² is selected from hydrogen, -C (O) R⁴ or alkyl optionally substituted with one or more groups independently selected from halogen, cyano or hydroxyl.
The compound of claim 27, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R² is hydrogen, -C (O) R⁴, or -alkyl-OH.
The compound of claim 28, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R⁴ is -O-alkyl.
The compound of claim 29, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R⁴ is -O-butyl.
The compound of claim 26, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is selected from:
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is alkyl optionally substituted with one or more groups independently selected from halogen, oxo, amino, hydroxyl, cyano, nitro or -N (R²) ₂.
The compound of claim 32, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is methyl optionally substituted with one or more groups independently selected from halogen or -N (R²) ₂.
The compound of claim 33, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein each R² is independently selected from hydrogen or -C (O) R⁴.
The compound of claim 34, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R⁴ is alkyl or -O-alkyl.
The compound of claim 35, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R⁴ is methyl or -O-butyl.
The compound of claim 32, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is selected from:
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is selected from cycloalkyl, aryl, heterocyclyl or heteroaryl.
The compound of claim 38, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L² is selected from
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L³ is a bond.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L³ is alkyl.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L³ is - (OCH₂CH₂) _m-or - (CH₂CH₂O) _m (CH₂) _n-.
The compound of claim 41 or 42, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L³ is selected from the group consisting of:
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L⁴ is a bond.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L⁴ is -N (R²) -.
The compound of claim 45, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein R² is hydrogen.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L⁴ is -O-.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L⁴ is -C (O) -.
The compound of claim 13, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L⁴ is heterocyclyl.
The compound of claim 49, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L⁴ is
The compound of any one of claims 1 to 50, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein L is selected from the group consisting of:

wherein **end of L is connected to E.
The compound of claim 1, or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, wherein the compound is selected from the group consisting of:
A pharmaceutical composition, comprising the compound of any one of claims 1 to 52 or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.
A method of treating a subject having or suspected of having a cancer, comprising administering to the subject a therapeutically effective amount of the compound of any one of claims 1 to 52 or a tautomer, a stereoisomer, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 53.
The method of claim 54, wherein the cancer is a solid tumor or a hematological cancer.
The method of claim 54, wherein the cancer is a breast cancer, multiple myeloma, plasma cell myeloma, leukemia, lymphoma, gastric cancer, acute myeloid leukemia, bladder cancer, brain cancer, bone marrow cancer, cervical cancer, chronic lymphocytic leukemia, colorectal cancer, esophageal cancer, hepatocellular cancer, lymphoblastic leukemia, follicular lymphoma, lymphoid malignancies of T-cell or B-cell origin, melanoma, myelogenous leukemia, myeloma, oral cancer, ovarian cancer, non-small cell lung cancer, chronic lymphocytic leukemia, prostate cancer, small cell lung cancer, or spleen cancer.
The method of any one of claims 54 to 56, wherein the compound is administered as monotherapy.
The method of any one of claims 54 to 56, wherein the compound is administered simultaneously, separately or sequentially with a second therapy.
The method of claim 58, wherein the second therapy is chemotherapy or immunotherapy.
The method of claim 59, wherein the second therapy is selected from the group consisting of a chemotherapeutic agent, an anti-tumor agent, a radiation therapy agent, an immunotherapy agent, an anti-angiogenesis agent, a targeted therapy agent, a cellular therapy agent, a gene therapy agent, a hormonal therapy agent, an antiviral agent, an antibiotic, an analgesics, an antioxidant, a metal chelator, and cytokines.
The method of claim 60, wherein the second therapy is a BTK inhibitor, a BCR-ABL inhibitor, a JAK1 inhibitor, a JAK2 inhibitor, a JAK3 inhibitor, a TYK2 inhibitor, a PARP inhibitor, a MEK inhibitor, an ERK inhibitor, or a RAF inhibitor.