Linkers for use in antibody drug conjugates

Abstract

The present disclosure provides traceless linkers capable of linking inducers of protein-protein interactions to cell binding agents. Also provided are compositions comprising the linked compounds. Such compounds and compositions are useful for treating diseases, such as cancer, in subjects in need thereof.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Application No. 63 / 241,914, filed September 8, 2021, U.S. Provisional Application No. 63 / 293,591, filed December 23, 2021, U.S. Provisional Application No. 63 / 351,639, filed June 13, 2022, and U.S. Provisional Application No. 63 / 374,282, filed September 1, 2022, which are incorporated by reference in their entireties.

[0002] Reference to an electronically submitted sequence listing The electronically submitted Sequence Listing (Name: 4547_020PC03_Seqlisting_ST26; Size: 52,902 bytes; Created: September 6, 2022) submitted with this application is hereby incorporated by reference in its entirety.

[0003] The present disclosure provides traceless linkers capable of linking inducers of protein-protein interactions to cell-binding agents. Also provided are compositions containing the linked compounds. Such compounds and compositions are useful for treating diseases in subjects in need thereof. [Background technology]

[0004] Protein-protein interactions (PPIs) represent a large class of therapeutic targets both inside and outside the cell. PPIs are central to all biological processes and are often dysregulated in disease. Despite the importance of PPIs in human biology, this target class has been extremely difficult to translate into therapeutic drugs. Thus, there is a continuing need for new compounds that can effectively induce PPIs and treat diseases such as cancer.

[0005] Targeted protein degraders are currently being investigated as a class of molecules that can be used to target and degrade difficult-to-druggable proteins. The most well-known targeted protein degraders utilize proteolysis-directing chimeras (PROTACs), a highly promising emerging field for targeting "undruggable" proteins that lack enzymatic activity and are resistant to classical inhibition. PROTACs are heterobifunctional small molecules that simultaneously bind to a target protein and an E3 ligase, resulting in the ubiquitination and subsequent degradation of the target. Molecular glues are another type of targeted protein degrader that differs from PROTACs in that they bind to an E3 ligase and alter the surface geometry of the ligase, allowing the ligase to bind to the target protein, resulting in the ubiquitination and degradation of the target. Targeted protein degraders offer exciting opportunities to modulate proteins in a manner independent of enzymatic or signaling activity; however, preparing compounds that distinguish between different cell types can be challenging.

[0006] Antibody-drug conjugates are an innovative therapeutic application that combines the inherent high specificity of monoclonal antibodies with the potent activity of small molecule drugs, which are often not suitable for systemic administration. The two entities are tethered via a linker that is typically cleaved at the target site.

[0007] The use of traceless linkers in antibody-drug conjugates allows for the release of therapeutic payloads without evidence of linker attachment. Ideally, the linker should be stable enough to allow the attached molecule to localize to its intended location without premature cleavage. The linker should also be capable of rapid cleavage to release the payload after internalization into the target cell. Known examples of traceless linkers use chemical triggers that require UV light for activation, or have a stability half-life of minutes to hours, or that produce a mixture of less active by-products upon activation.

[0008] Despite ongoing research, there remains a need to selectively target diseased tissues and cells and proteins previously considered "undruggable" with compounds that can induce desirable protein-protein interactions, such as compounds that induce protein degradation. Summary of the Invention

[0009] In certain aspects, the present disclosure provides compounds of formula (XX): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein: a is 1 to 10; n is 0 or 1, R 1 is a compound that induces protein-protein interactions, R 2 is hydrogen, -(CH2CH2O) v -CH3, C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl), and v is 1 to 24; each Y is independently S or O; L is a cleavable linker, Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0010] In some embodiments, the present disclosure provides a compound of formula (XXXII): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein: a is 1 to 10; A' is [ka] or [ka] wherein: n is 0 or 1, each Y is independently S or O; [ka] is R 1 indicates the point of attachment to [ka] indicates the point of attachment to the methylene group, R 1 is a compound that induces protein-protein interaction together with A', R 2 is hydrogen, R 1 -A', a group that provides stability to R 1 -A', a group that provides solubility to A', and R 1 -A' is selected from groups that provide stability and solubility to A'; L is a cleavable linker, Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0011] In certain aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the binding moiety is an antibody, an antibody fragment, or an antigen-binding fragment.

[0012] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein a is 2-8.

[0013] In some aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the linker is cleavable by a protease.

[0014] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein L is [ka] is selected from the group consisting of During the ceremony, q is 2 to 10; Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are each independently absent or a naturally occurring amino acid residue in the L-configuration or the D-configuration, with the proviso that Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are amino acid residues, [ka] is the point of attachment to the parent molecular moiety, [ka] is the point of attachment to the binding moiety.

[0015] In some embodiments, Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 is independently absent or selected from the group consisting of L-valine, D-valine, L-citrulline, D-citrulline, L-alanine, D-alanine, L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-asparagine, D-asparagine, L-phenylalanine, D-phenylalanine, L-lysine, D-lysine, and glycine, with the proviso that Z 1 , Z 2 , Z 3 , Z 4, and Z 5 At least two of the residues are amino acid residues.

[0016] In some embodiments, Z 1 is absent or is glycine, Z 2 is absent or selected from the group consisting of L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-alanine, D-alanine, and glycine; Z 3 is selected from the group consisting of L-valine, D-valine, L-alanine, D-alanine, L-phenylalanine, D-phenylalanine, and glycine; Z 4 is selected from the group consisting of L-citrulline, D-citrulline, L-asparagine, D-asparagine, L-lysine, D-lysine, L-phenylalanine, D-phenylalanine, and glycine; Z 5 is absent or is glycine.

[0017] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein L is [ka] is.

[0018] In some embodiments, q is 4.

[0019] In some aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein L is a bioreducible linker.

[0020] In some embodiments, L is [ka] wherein: q is 2 to 10; R, R', R'', and R''' are each independently selected from hydrogen, C-C alkoxyC-C alkyl, (C-C) NC-C alkyl, and C-C alkyl, or two geminal R groups together with the carbon atoms to which they are attached can form a cyclobutyl or cyclopropyl ring; [ka] is the point of attachment to the parent molecular moiety, [ka] is the point of attachment to the binding moiety.

[0021] In some embodiments, L is [ka] is.

[0022] In some embodiments, q is 2.

[0023] In some aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein L is a click-to-release linker.

[0024] In some embodiments, L is [ka] and During the ceremony, q is 2 to 10; [ka] is the point of attachment to the parent molecular moiety, [ka] is the point of attachment to the binding moiety.

[0025] In some aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein L is a beta-glucuronidase cleavable linker.

[0026] In some embodiments, L is [ka] During the ceremony, q is 2 to 10; ---- is absent or a bond, [ka] is the point of attachment to the parent molecular moiety, [ka] is the point of attachment to the binding moiety.

[0027] In certain embodiments, L is linked to a cysteine, lysine, tyrosine, or glutamine within Bm. In certain embodiments, the cysteine ​​or lysine is an engineered cysteine ​​or lysine. In some embodiments, the cysteine ​​or lysine is endogenous to Bm.

[0028] In some embodiments, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein Bm is an antibody or antigen-binding portion thereof. In certain embodiments, L is attached to an engineered cysteine ​​at position S239 and / or K334 of the heavy chain of the antibody or antigen-binding portion thereof, according to EU numbering. In some embodiments, L is attached to a glutamine at position 295 of the heavy chain of the antibody or antigen-binding portion thereof, according to EU numbering.

[0029] In some aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the protein to which Bm binds is a surface antigen, and optionally, binding of Bm to the surface antigen results in internalization of the conjugate or a pharmaceutically acceptable salt thereof into a cell.

[0030] In some embodiments, the surface antigen is selected from the group consisting of 5T4, ACE, ADRB3, AKAP-4, ALK, AOC3, APP, Axin1, AXL, B7H3, B7-H4, BCL2, BCMA, bcr-abl, BORIS, BST2, C242, C4.4a, CA 125, CA6, CA9, CAIX, CCL11, CCR5, CD123, CD133, CD138, CD142, CD15, CD15-3, CD171, CD17 9a, CD18, CD19, CD19-9, CD2, CD20, CD22, CD23, CD24, CD25, CD27L, CD28, CD3, CD30, CD31, C D300LF, CD33, CD352, CD37, CD38, CD4, CD40, CD41, CD44, CD44v6, CD5, CD51, CD52, CD54, CD 56, CD62E, CD62P, CD62L, CD70, CD71, CD72, CD74, CD79a, CD79b, CD80, CD90, CD97, CD125, C D138, CD141, CD147, CD152, CD154, CD326, CEA, CEACAM5, CFTR, clumping factor, cKit, claudin 3, claudin 18.2, CLDN6, CLEC12A, CLL-1, cll3, c-MET, Crypto1 growth factor, CS1, CTLA-4, CXCR2, CXORF61, cyclin B1, CYP1B1, cadherin-3, cadherin-6, DLL3, E7, EDNRB, EFNA4, EGFR, EGFRvIII, ELF2M, EMR2, ENPP3, EPCAM, EphA2, ephrin-A4, ephrin-B2, EPHB4, ERBB2 (Her2 / neu), ErbB3, ERG (TMPRSS2) ETS fusion gene), ETBR, ETV6-AML, FAP, FCAR, FCRL5, FGFR1, FGFR2, FGFR3, FGFR4, FLT3, folate receptor alpha, folate receptor beta, FOLR1, Fos-related antigen 1, fucosyl GM1, GCC, GD2, GD3, GloboH, GM3, GPC1, GPC2, GPC3, gplOO, GPNMB, GPR20, GPRC5D, GUCY2C, HAVCR1, HER2, HER3, HGF, HMI.24, HMWMAA, HPV E6, hTERT, human telomerase reverse transcriptase, ICAM, ICOS-L, IFN-α, IFN-γ, IGF-I receptor, IGLL1, IL-2 receptor, IL-4 receptor, IL-13Ra2, IL-l lRa, IL-1 receptor, IL-12 receptor, IL-23 receptor, IL-13 receptor, IL-22 receptor, IL-4 receptor, IL-5 receptor, IL-6 receptor, interferon receptor, integrin (α4, α). v β3, α v β5, α v β6, α1β4, α4β1, α4β7, α5β1, α6β4, α IIbIntegrin β3), integrin alpha V, intestinal carboxylesterase, KIT, LAGE-1a, LAIR1, LAMP-1, LCK, legumain, LewisY, LFA-1 (CD11a), L-selectin (CD62L), LILRA2, LIV-1, LMP2, LRRC15, LY6E, LY6K, LY75, MAD-CT-1, MAD-CT-2, MAGE Al, MelanA / MARTl, mesothelin, ML-IAP, MSLN, mucin, MUC1, MUC16, mut hsp70-2, MYCN, myostatin, NA17, NaPi2b, NCA-90, NCAM, nectin-4, NGF, NOTCH1, NOTCH2, NOTCH3, NOTCH4, NY-BR-1, NY-ESO-1, o-acetyl-GD2, OR51E2, OY-TES1, p53, p53 mutant, PANX3, PAP, PAX3, PAX5, p-CAD, PCTA-1 / galectin 8, PD-L1, PD-L2, PDGFR, PDGFR-beta, phosphatidylserine, PIK3CA, PLAC1, polysialic acid, protease, prostate cancer cells, prostein, Pseudomonas aeruginosa, rabies, survivin and telomerase, PD-1, PRSS21, PSCA, PSMA, PTK7, RAGE-1, RANKL, Ras mutants, respiratory syncytial virus, rhesus factor, RhoC, RON, ROR1, ROR2, RU1, RU2, sarcoma translocation breakpoints, SART3, SLAMF7, SLC44A4, sLe, SLITRK6, sperm protein 17, sphingosine-1-phosphate, SSEA-4, SSX2, STEAP1, TAG72, TARP, TCRβ, TEM1 / CD248, TEM7R, tenascin-C, TF, TGF-1, TGF-β2, TNF-α, TGS5, Tie2, TIM-1, Tn Ag, TRAC, TRAIL-R1, TRAIL-R2, TROP-2, TRP-2, TRPV1, TSHR, tumor antigen CTAA16.88, tyrosinase, UPK2, VEGF, VEGFR1, VEGFR2, vimentin, WT1, XAGE1, or a combination thereof.

[0031] In some embodiments, the surface antigen comprises HER2, CD20, CD38, CD33, BCMA, CD138, EGFR, FGFR4, GD2, PDGFR, or a combination thereof. In some embodiments, the surface antigen comprises CD79b. In some embodiments, the surface antigen comprises PSMA.

[0032] In some embodiments, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein Bm binds to a nuclear hormone receptor. In some embodiments, the nuclear hormone receptor is an androgen receptor (AR) or an estrogen receptor (ER).

[0033] In some embodiments, the antibody is selected from the group consisting of rituximab, trastuzumab, gemtuzumab, pertuzumab, obinutuzumab, ofatumumab, daratumumab, STI-6129, lintuzumab, huMy9-6, balantamab, indatuximab, cetuximab, dinutuximab, anti-CD38 A2 antibody, huAT15 / 3 antibody, alemtuzumab, ibritumomab, tositumomab, bevacizumab, panitumumab, tremelimumab, ticilimumab, catumaxomab, oregovomab, and veltuzumab.

[0034] In some embodiments, the antibody is rituximab, trastuzumab, pertuzumab, huMy9-6, lintuzumab, or gemtuzumab. In some embodiments, the antibody is polatuzumab, J591, or belantamab. In some embodiments, the antibody is CD33-D.

[0035] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII) or a pharmaceutically acceptable salt thereof, wherein R 2 is a group that provides stability to the complex. In some embodiments, R 2is selected from C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl).

[0036] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 2 is hydrogen or C1-C6 alkyl. In some embodiments, R 2 is methyl.

[0037] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 2 is methyl.

[0038] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 2 is a group that provides solubility to the complex. In some embodiments, R 2 teeth, [ka] [ka] is selected from During the ceremony, each n is independently 1, 2, 3, 4, or 5; each y is independently 1 or 2; Each R is independently hydrogen, CH 11 O5, C 12 H 21 O 10 , C 18 H 31 O 15 , or C 24 H 41 O 20 is.

[0039] In some aspects, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein each Y is O.

[0040] In some embodiments, the present disclosure provides a conjugate of formula (XX) or a pharmaceutically acceptable salt thereof, wherein R 1 In some embodiments, the present disclosure provides a conjugate of formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 1 -A' is a PPI modulator.

[0041] In some embodiments, the present disclosure provides a conjugate of formula (XX) or a pharmaceutically acceptable salt thereof, wherein R 1 In some aspects, the present disclosure provides a conjugate of formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 1 -A' is a targeted proteolytic agent. In some embodiments, the targeted proteolytic agent is a substituted isoindoline. In some embodiments, the targeted proteolytic agent is a 5'-substituted isoindoline.

[0042] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 1 is the expression: [ka] wherein [ka] indicates the point of attachment to the parent molecular moiety, A is phenyl or C4-C 10 is a cycloalkyl ring, R 10 is independently selected from hydrogen and halo; U is selected from NH and CF; R 20 -CH3, -C(O)R 3 , -N(R4 )2, -(CH2) n OH, -(CH2) n N(R 4 )2, -(CH2) n Q'(CH2) m OH, -(CH2) n Q'(CH2) m SH, and -(CH2) n Q'(CH2) m N(R 4 )2, wherein: R 3 is hydrogen or C1-C6 alkyl, Each R 4 are independently hydrogen or C1-C6 alkyl; Q' is O, S, or NR 4 and n is 1 to 6; m is 2 to 5.

[0043] In some embodiments, A is phenyl; U is NH, R 10 is a halo, R 20 is methyl.

[0044] In some embodiments, A is phenyl; U is NH, R 10 is a halo, R 20 is -(CH2)2O(CH2)2NHCH3.

[0045] In some embodiments, the present disclosure provides a conjugate of formula (XX) or a pharmaceutically acceptable salt thereof, wherein R 1 is a proteolysis targeting chimera (PROTAC). In some aspects, the present disclosure provides a conjugate of formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 1 -A' is a proteolysis-directed chimera (PROTAC).

[0046] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 1 is the expression: POI-L 100 -CBN wherein POI is a compound that binds to a protein of interest, L 100 is the PROTAC linker, CBN is the binding moiety of cereblon.

[0047] In some embodiments, the protein of interest is a nuclear hormone receptor, a translation termination factor, a transcription factor, a cyclin-dependent kinase, a tyrosine kinase, a serine / threonine kinase, or an E3 ligase. In some embodiments, the protein of interest is selected from CD33, GSPT1, BRD4, AR, ER, IKZF1 / 3, CK1a, BCL-XL, IKZF2, IRAK4, BTK, STAT3, BTK and iMiD, BRD9, TRK, MDM2, CDK2 / CDK9, CD97b, and EGFR.

[0048] In some embodiments, L 100 comprises one or more functional groups selected from glycol, alkyl, alkynyl, triazolyl, piperazinyl, piperidinyl, and combinations thereof.

[0049] In some embodiments, CBN is [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A', [ka] L 100 indicates the attachment point to

[0050] In some aspects, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein R 1 teeth, [ka] [ka] [ka] [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A'.

[0051] In some embodiments, the present disclosure provides a compound of formula (XXI): [ka] A complex of or a pharmaceutically acceptable salt thereof, During the ceremony, a is 1 to 10;

[0052] Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0053] In some embodiments, the present disclosure provides a conjugate of formula (XXI), or a pharmaceutically acceptable salt thereof, wherein a is 2 to 8.

[0054] In some aspects, the present disclosure provides a conjugate of formula (XXI), or a pharmaceutically acceptable salt thereof, wherein Bm is an antibody or an antigen-binding portion thereof.

[0055] In some aspects, the disclosure provides a conjugate of Formula (XXI), or a pharmaceutically acceptable salt thereof, wherein the protein that specifically binds to the binding moiety is a surface antigen.

[0056] In some aspects, the disclosure provides a conjugate of Formula (XXI), or a pharmaceutically acceptable salt thereof, wherein the surface antigen is selected from the group consisting of 5T4, ACE, ADRB3, AKAP-4, ALK, AOC3, APP, Axin1, AXL, B7H3, B7-H4, BCL2, BCMA, bcr-abl, BORIS, BST2, C242, C4.4a, CA 125, CA6, CA9, CAIX, CCL11, CCR5, CD123, CD133, CD138, CD142, CD15, CD15-3, CD171, CD17 9a, CD18, CD19, CD19-9, CD2, CD20, CD22, CD23, CD24, CD25, CD27L, CD28, CD3, CD30, CD31, C D300LF, CD33, CD352, CD37, CD38, CD4, CD40, CD41, CD44, CD44v6, CD5, CD51, CD52, CD54, CD 56, CD62E, CD62P, CD62L, CD70, CD71, CD72, CD74, CD79a, CD79b, CD80, CD90, CD97, CD125, C D138, CD141, CD147, CD152, CD154, CD326, CEA, CEACAM5, CFTR, clumping factor, cKit, claudin 3, claudin 18.2, CLDN6, CLEC12A, CLL-1, cll3, c-MET, Crypto1 growth factor, CS1, CTLA-4, CXCR2, CXORF61, cyclin B1, CYP1B1, cadherin-3, cadherin-6, DLL3, E7, EDNRB, EFNA4, EGFR, EGFRvIII, ELF2M, EMR2, ENPP3, EPCAM, EphA2, ephrin-A4, ephrin-B2, EPHB4, ERBB2 (Her2 / neu), ErbB3, ERG (TMPRSS2) ETS fusion gene), ETBR, ETV6-AML, FAP, FCAR, FCRL5, FGFR1, FGFR2, FGFR3, FGFR4, FLT3, folate receptor alpha, folate receptor beta, FOLR1, Fos-related antigen 1, fucosyl GM1, GCC, GD2, GD3, GloboH, GM3, GPC1, GPC2, GPC3, gplOO, GPNMB, GPR20, GPRC5D, GUCY2C, HAVCR1, HER2, HER3, HGF, HMI.24, HMWMAA, HPV E6, hTERT, human telomerase reverse transcriptase, ICAM, ICOS-L, IFN-α, IFN-γ, IGF-I receptor, IGLL1, IL-2 receptor, IL-4 receptor, IL-13Ra2, IL-l lRa, IL-1 receptor, IL-12 receptor, IL-23 receptor, IL-13 receptor, IL-22 receptor, IL-4 receptor, IL-5 receptor, IL-6 receptor, interferon receptor, integrin (α4, α). v β3, α v β5, α v β6, α1β4, α4β1, α4β7, α5β1, α6β4, α IIbIntegrin β3), integrin alpha V, intestinal carboxylesterase, KIT, LAGE-1a, LAIR1, LAMP-1, LCK, legumain, LewisY, LFA-1 (CD11a), L-selectin (CD62L), LILRA2, LIV-1, LMP2, LRRC15, LY6E, LY6K, LY75, MAD-CT-1, MAD-CT-2, MAGE Al, MelanA / MARTl, mesothelin, ML-IAP, MSLN, mucin, MUC1, MUC16, mut hsp70-2, MYCN, myostatin, NA17, NaPi2b, NCA-90, NCAM, nectin-4, NGF, NOTCH1, NOTCH2, NOTCH3, NOTCH4, NY-BR-1, NY-ESO-1, o-acetyl-GD2, OR51E2, OY-TES1, p53, p53 mutant, PANX3, PAP, PAX3, PAX5, p-CAD, PCTA-1 / galectin 8, PD-L1, PD-L2, PDGFR, PDGFR-beta, phosphatidylserine, PIK3CA, PLAC1, polysialic acid, protease, prostate cancer cells, prostein, Pseudomonas aeruginosa, rabies, survivin and telomerase, PD-1, PRSS21, PSCA, PSMA, PTK7, RAGE-1, RANKL, Ras mutants, respiratory syncytial virus, rhesus factor, RhoC, RON, ROR1, ROR2, RU1, RU2, sarcoma translocation breakpoints, SART3, SLAMF7, SLC44A4, sLe, SLITRK6, sperm protein 17, sphingosine-1-phosphate, SSEA-4, SSX2, STEAP1, TAG72, TARP, TCRβ, TEM1 / CD248, TEM7R, tenascin-C, TF, TGF-1, TGF-β2, TNF-α, TGS5, Tie2, TIM-1, Tn Ag, TRAC, TRAIL-R1, TRAIL-R2, TROP-2, TRP-2, TRPV1, TSHR, tumor antigen CTAA16.88, tyrosinase, UPK2, VEGF, VEGFR1, VEGFR2, vimentin, WT1, XAGE1, or a combination thereof.

[0057] In some embodiments, the surface antigen comprises HER2, CD20, CD38, CD33, BCMA, CD138, EGFR, FGFR, GD2, PDGFR, or a combination thereof. In some embodiments, the surface antigen comprises CD79b. In some embodiments, the surface antigen comprises PSMA.

[0058] In some embodiments, the antibody comprises rituximab, trastuzumab, gemtuzumab, pertuzumab, obinutuzumab, ofatumumab, daratumumab, STI-6129, lintuzumab, huMy9-6, belantamab, indatuximab, cetuximab, dinutuximab, anti-CD38 A2 antibody, huAT15 / 3 antibody, alemtuzumab, ibritumomab, tositumomab, bevacizumab, panitumumab, tremelimumab, ticilimumab, catumaxomab, oregovomab, or veltuzumab. In some embodiments, the antibody comprises lorvotuzumab. In some embodiments, the antibody comprises sacituzumab.

[0059] In some embodiments, the antibody is rituximab, trastuzumab, pertuzumab, huMy9-6, lintuzumab, or gemtuzumab. In some embodiments, the antibody is polatuzumab, J591, or belantamab. In some embodiments, the antibody is CD33-D.

[0060] In some aspects, the disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein (a) the protein to which Bm binds is CD33, and R 1 (b) Bm binds to mouse double minute 2 homolog (MDM2), or (b) the protein that Bm binds is prostate-specific membrane antigen (PSMA), and R 1 binds to the androgen receptor (AR), or (c) the protein that Bm binds to is CD33, and R 1 binds to bromodomain-containing protein 4 (BRD4), or (d) the protein to which Bm binds is HER2, and R 1binds to G1 to S Phase Transition 1 (GSPT1), or (e) the protein that Bm binds is CD33, and R 1 In some embodiments, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the protein to which Bm binds is CD79b, and R 1 In some embodiments, the present disclosure provides a conjugate of Formula (XX) or Formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the protein to which Bm binds is HER2, and R 1 In some embodiments, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the protein to which Bm binds is BCMA, and R 1 In some embodiments, the present disclosure provides a conjugate of formula (XX) or formula (XXXII), or a pharmaceutically acceptable salt thereof, wherein the protein to which Bm binds is HER2, and R 1 binds to the ER.

[0061] In some aspects, the present disclosure provides pharmaceutical compositions comprising a conjugate described herein or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers.

[0062] In some aspects, the present disclosure provides a method of treating cancer in a subject in need thereof, the method comprising administering to the subject a pharmaceutically acceptable amount of a conjugate or composition described herein, or a pharmaceutically acceptable salt thereof. In some aspects, the cancer is a solid tumor. In some aspects, the cancer is a hematological tumor. In some aspects, the cancer is breast cancer, gastric cancer, lymphoma, acute myeloid leukemia, multiple myeloma, head and neck cancer, squamous cell carcinoma, and / or hepatocellular carcinoma. In some aspects, the cancer is prostate cancer, breast cancer, gastric cancer, non-small cell lung cancer, cholangiocarcinoma, colon cancer, ovarian cancer, lung cancer, or neuregulin-1 (NRG1)-positive cancer. In some aspects, the cancer is non-Hodgkin's lymphoma (NHL). In some aspects, the cancer is B-cell non-Hodgkin's lymphoma. In some aspects, the cancer is diffuse large B-cell lymphoma (DLBCL). In some embodiments, the method further comprises administering to the subject a pharmaceutically acceptable amount of an additional agent before, after, or simultaneously with the conjugate or composition described herein, or a pharmaceutically acceptable salt thereof. In some embodiments, the additional agent is a cytotoxic agent or an immune response modifier. In some embodiments, the immune response modifier is a checkpoint inhibitor. In some embodiments, the checkpoint inhibitor comprises a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a TIM3 inhibitor, and / or a LAG-3 inhibitor.

[0063] In some embodiments of the method, (a) the protein to which Bm binds is CD33, and R 1 (b) Bm binds to MDM2 or G1 to S Phase Transition 1 (GSPT1), and the cancer is acute myeloid leukemia, or (b) the protein that Bm binds to is prostate-specific membrane antigen (PSMA), and R 1 binds to the androgen receptor (AR) and the cancer is prostate cancer, or (c) the protein to which Bm binds is CD33 and R 1 binds to bromodomain-containing protein 4 (BRD4), and the cancer is acute myeloid leukemia, or (d) the protein to which Bm binds is HER2, and R 1In some embodiments of the method, the protein that Bm binds is CD79b, and R binds to G1 to S Phase Transition 1 (GSPT1), and the cancer is breast cancer, gastric cancer, non-small cell lung cancer, cholangiocarcinoma, colon cancer, ovarian cancer, or neuregulin-1 (NRG1)-positive cancer. 1 In some embodiments, the protein that Bm binds to is HER2, and the R binds to IRAK4, and the cancer is non-Hodgkin's lymphoma (NHL), e.g., B-cell non-Hodgkin's lymphoma or diffuse large B-cell lymphoma (DLBCL). In some embodiments, the protein that Bm binds to is HER2, and the R binds to IRAK4. 1 In some embodiments, the protein that Bm binds to is BCMA, and R binds to BRD4, and the cancer is breast cancer, gastric cancer, non-small cell lung cancer, cholangiocarcinoma, colon cancer, ovarian cancer, or neuregulin-1 (NRG1)-positive cancer. 1 binds to BRD4 and the cancer is multiple myeloma.

[0064] In some aspects of the methods, the methods further comprise administering to the subject a pharmaceutically acceptable amount of an additional agent before, after, or simultaneously with the conjugate or composition described herein. In some aspects, the additional agent is a cytotoxic agent or an immune response modifier. In some aspects, the immune response modifier is a checkpoint inhibitor.

[0065] In some embodiments, the present disclosure provides a compound of formula (XXII): [ka] The compound or a pharmaceutically acceptable salt thereof, wherein: n is 0 or 1, R 1 is a compound that induces protein-protein interactions, R 2 is hydrogen, -(CH2CH2O) v -CH3, C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl), and v is 1 to 24; each Y is independently S or O; L * is a cleavable linker precursor attached to the binding moiety.

[0066] In some embodiments, the present disclosure provides a compound of formula (XXXI): [ka] The compound or a pharmaceutically acceptable salt thereof, wherein: A' is [ka] or [ka] wherein: n is 0 or 1, each Y is independently S or O; [ka] [ka] indicates the point of attachment to the methylene group, R 1 is a compound that induces protein-protein interaction together with A', R 2 is hydrogen, R 1 -A', a group that provides stability to R 1 -A', a group that provides solubility to A', and R 1 -A' is selected from groups that provide stability and solubility to A'; L is a cleavable linker precursor that is attached to the binding moiety.

[0067] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * is a protease cleavable linker precursor.

[0068] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * teeth, [ka] is selected from the group consisting of During the ceremony, q is 2 to 10; Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are each independently absent or a naturally occurring amino acid residue in the L-configuration or the D-configuration, with the proviso that Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are amino acid residues, [ka] is the point of attachment to the parent molecular moiety.

[0069] In some embodiments, Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 is independently absent or selected from the group consisting of L-valine, D-valine, L-citrulline, D-citrulline, L-alanine, D-alanine, L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-asparagine, D-asparagine, L-phenylalanine, D-phenylalanine, L-lysine, D-lysine, and glycine, with the proviso that Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 At least two of the residues are amino acid residues.

[0070] In some embodiments, Z 1 is absent or is glycine, Z 2 is absent or selected from the group consisting of L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-alanine, D-alanine, and glycine; Z 3 is selected from the group consisting of L-valine, D-valine, L-alanine, D-alanine, L-phenylalanine, D-phenylalanine, and glycine; Z 4 is selected from the group consisting of L-citrulline, D-citrulline, L-asparagine, D-asparagine, L-lysine, D-lysine, L-phenylalanine, D-phenylalanine, and glycine; Z 5 is absent or is glycine.

[0071] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * teeth [ka] and During the ceremony, [ka] is the point of attachment to the parent molecular moiety.

[0072] In some embodiments, q is 4.

[0073] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * is a bioreducible linker precursor.

[0074] In some aspects, the present application provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * teeth, [ka] wherein: q is 2 to 10; R, R', R'', and R''' are each independently selected from hydrogen, C-C alkoxyC-C alkyl, (C-C) NC-C alkyl, and C-C alkyl, or two geminal R groups together with the carbon atoms to which they are attached can form a cyclobutyl or cyclopropyl ring; [ka] is the point of attachment to the parent molecular moiety.

[0075] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * teeth [ka] is.

[0076] In some embodiments, q is 2.

[0077] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * is the click-to-release linker precursor.

[0078] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * teeth [ka] and During the ceremony, q is 2 to 10; [ka] is the point of attachment to the parent molecular moiety.

[0079] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * is a beta-glucuronidase cleavable linker precursor.

[0080] In some aspects, the present disclosure provides a compound of formula (XXII) or formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein L * teeth [ka] and During the ceremony, q is 2 to 10; ---- is absent or a bond, [ka] is the point of attachment to the parent molecular moiety.

[0081] In some aspects, the present disclosure provides compounds of formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 2 is R 1 -A' is a group that provides stability to -A'. In some embodiments, R 2 is selected from C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl).

[0082] In some aspects, the disclosure provides a compound of Formula (XXII) or Formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 2 is hydrogen or C1-C6 alkyl. In some embodiments, R2 is methyl.

[0083] In some aspects, the present disclosure provides compounds of formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 2 is R 1 -A' is a group that provides solubility to A'.

[0084] In some embodiments, R 2 teeth,

[0085] [ka] [ka] is selected from During the ceremony, each n is independently 1, 2, 3, 4, or 5; each y is independently 1 or 2; Each R is independently hydrogen, CH 11 O5, C 12 H 21 O 10 , C 18 H 31 O 15 , or C 24 H 41 O 20 is.

[0086] In some aspects, the disclosure provides compounds of Formula (XXII) or Formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein each Y is O.

[0087] In some aspects, the disclosure provides compounds of formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 1 -A' is a PPI modulator.

[0088] In some aspects, the present disclosure provides a compound of formula (XXII), or a pharmaceutically acceptable salt thereof, wherein R 1In some aspects, the present disclosure provides a compound of formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 1 -A' is a targeted proteolytic agent. In some embodiments, the targeted proteolytic agent is a substituted isoindoline. In some embodiments, the targeted proteolytic agent is a 5'-substituted isoindoline.

[0089] In some aspects, the disclosure provides compounds of formula (XXII) or (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 1 is the formula (XXX): [ka] is a compound of During the ceremony, [ka] indicates the point of attachment to the parent molecular moiety, A is phenyl or C4-C 10 is a cycloalkyl ring, R 10 is independently selected from hydrogen and halo; U is selected from NH and CF; R 20 -CH3, -C(O)R 3 , -N(R 4 )2, -(CH2) n OH, -(CH2) n N(R 4 )2, -(CH2) n Q'(CH2) m OH, -(CH2) n Q'(CH2) m SH, and -(CH2) n Q'(CH2) m N(R 4 )2, wherein: R 3 is hydrogen or C1-C6 alkyl, Each R 4 are independently hydrogen or C1-C6 alkyl; Q' is O, S, or NR 4 and n is 1 to 6; m is 2 to 5.

[0090] In some embodiments, A is phenyl; U is NH, R 10 is a halo, R 20 is methyl.

[0091] In some embodiments, A is phenyl; U is NH, R 10 is a halo, R 20 is -(CH2)2O(CH2)2NHCH3.

[0092] In some aspects, the present disclosure provides a compound of formula (XXII), or a pharmaceutically acceptable salt thereof, wherein R 1 is a proteolysis-directed antigen-activating chimera (PROTAC).

[0093] In some aspects, the present disclosure provides a compound of formula (XXII), or a pharmaceutically acceptable salt thereof, wherein R 1 -A' is a proteolysis-directed chimera (PROTAC).

[0094] In some embodiments, R 1 is the expression: POI-L 100 -CBN wherein POI is a compound that binds to a protein of interest, L 100 is the PROTAC linker, CBN is the binding moiety of cereblon.

[0095] In some embodiments, the protein of interest is a nuclear hormone receptor, a translation termination factor, a transcription factor, a cyclin-dependent kinase, a tyrosine kinase, a serine / threonine kinase, or an E3 ligase. In some embodiments, the protein of interest is selected from CD33, GSPT1, BRD4, AR, ER, IKZF1 / 3, CK1a, BCL-XL, IKZF2, IRAK4, BTK, STAT3, BTK and iMiD, BRD9, TRK, MDM2, CDK2 / CDK9, CD97b, and EGFR.

[0096] In some embodiments, L 100 comprises one or more functional groups selected from glycol, alkyl, alkynyl, triazolyl, piperazinyl, piperidinyl, and combinations thereof.

[0097] In some embodiments, CBN is [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A', [ka] L 100 indicates the attachment point to

[0098] In some aspects, the disclosure provides a compound of Formula (XXII) or Formula (XXXI), or a pharmaceutically acceptable salt thereof, wherein R 1 teeth, [ka] [ka] [ka] [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A'.

[0099] In certain aspects, the present disclosure provides compounds of formula (XXXII): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein: a is 1 to 10; A' is [ka] or [ka] wherein: n is 0 or 1, each Y is independently S or O; [ka] is R 1 indicates the point of attachment to [ka] indicates the point of attachment to the methylene group, R 1 is a compound that induces protein-protein interaction together with A', R 2 is hydrogen, R 1 -A', a group that provides stability to R 1 - a group that provides solubility to A, and R 1 -A' is selected from groups that provide stability and solubility to A'; L is a cleavable linker, Bm is a binding moiety capable of specifically binding to a protein, Such a method comprises: (XXXI) [ka] The compound or a pharmaceutically acceptable salt thereof [wherein: A', R 1 , and R 2 is as defined above, L * is a cleavable linker precursor], This involves reacting with a binding moiety that is capable of specifically binding to the protein.

[0100] In some embodiments, L is substituted for cysteine, lysine, tyrosine, or glutamine in Bm. * In some embodiments, the cysteine ​​or lysine is an engineered cysteine ​​or lysine. In some embodiments, the cysteine ​​or lysine is endogenous to Bm.

[0101] In some embodiments, the binding moiety is an antibody or an antigen-binding portion thereof. * is attached to an engineered cysteine ​​at position S239 and / or K334 of the heavy chain of the antibody, or antigen-binding portion thereof, according to EU numbering. * is attached to glutamine at position 295 of the heavy chain of the antibody or antigen-binding portion thereof according to EU numbering. In some embodiments, attachment is via site-specific attachment.

[0102] In some embodiments of the method, (a) the protein to which Bm binds is CD33, and R 1 (b) Bm binds to mouse double minute 2 homolog (MDM2), or (b) the protein that Bm binds is prostate-specific membrane antigen (PSMA), and R 1 binds to the androgen receptor (AR), or (c) the protein that Bm binds to is CD33, and R 1binds to bromodomain-containing protein 4 (BRD4), or (d) the protein to which Bm binds is HER2, and R 1 binds to G1 to S Phase Transition 1 (GSPT1), or (e) the protein that Bm binds is CD33, and R 1 In some embodiments of the method, the protein to which Bm binds is CD79b, and R 1 In some embodiments of the method, the protein to which Bm binds is HER2, and R 1 In some embodiments of the method, the protein to which Bm binds is BCMA, and R 1 In some embodiments of the method, the protein to which Bm binds is HER2, and R 1 binds to the ER.

[0103] In some embodiments of the method, R 1 -A' is a targeted proteolytic agent. In some embodiments, R 1 is the expression: POI-L 100 -CBN wherein POI is a compound that binds to a protein of interest, L 100 is the PROTAC linker, CBN is the binding moiety of cereblon.

[0104] In some embodiments, the protein of interest is a nuclear hormone receptor, a translation termination factor, a transcription factor, a cyclin-dependent kinase, a tyrosine kinase, a serine / threonine kinase, or an E3 ligase. In some embodiments, the protein of interest is selected from CD33, GSPT1, BRD4, AR, ER, IKZF1 / 3, CK1a, BCL-XL, IKZF2, IRAK4, BTK, STAT3, BTK and iMiD, BRD9, TRK, MDM2, CDK2 / CDK9, CD97b, and EGFR.

[0105] In some embodiments, L100 comprises one or more functional groups selected from glycol, alkyl, alkynyl, triazolyl, piperazinyl, piperidinyl, and combinations thereof.

[0106] In some embodiments, CBN is [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A', [ka] L 100 indicates the attachment point to

[0107] In some embodiments, R 1 teeth, [ka] [ka] [ka] [ka] is selected from

[0108] During the ceremony, [ka] indicates the point of attachment to A'.

[0109] In some aspects, the present disclosure provides a composite produced by the methods described herein.

[0110] In some aspects, the present disclosure provides methods of delivering a complex that induces a protein-protein interaction to a cell, such methods comprising contacting the cell with a complex or composition described herein, or a pharmaceutically acceptable salt thereof.

[0111] In some embodiments, the present disclosure provides a compound of formula (XXXII): [ka] A complex of or a pharmaceutically acceptable salt thereof to a cell, wherein: a is 1 to 10; A' is [ka] or [ka] wherein: n is 0 or 1, each Y is independently S or O; [ka] is R 1 indicates the point of attachment to [ka] indicates the point of attachment to the methylene group, R 1 is a compound that induces protein-protein interaction together with A', R 2 is hydrogen, R 1 -A', a group that provides stability to R 1 -A', a group that provides solubility to A', and R 1 -A' is selected from groups that provide stability and solubility to A', and L is a cleavable linker; Bm is a binding moiety capable of specifically binding to a protein, Such methods include contacting the cell with a complex of formula (XXXII), or a pharmaceutically acceptable salt thereof. [Brief explanation of the drawings]

[0112] [Figure 1] Figure 1 shows the LCMS spectra of Pertuzumab-Compound (I) conjugate (DAR=8) at 4°C, pH 5.5 and after 24 hours of incubation at 37°C, pH 7.5. [Figure 2A] Figure 1 shows the LCMS spectrum of Pertuzumab-Compound (VIII) conjugate (DAR=8) after 24 hours of incubation at 37°C and pH 7.4. [Figure 2B] Figure 1 shows the LCMS spectrum of Pertuzumab-Compound (VIII) conjugate (DAR=8) after 24 hours of incubation at 37°C and pH 7.4. [Figure 3] Figure 1 shows the LCMS spectrum of Pertuzumab-Compound (X) conjugate (DAR=3.66) after 24 hours of incubation at 37°C and pH 7.5. [Figure 4] Figure 1 shows the LCMS spectrum of Pertuzumab-Compound (XI) conjugate (DAR=3.74) after 24 hours of incubation at 37°C and pH 7.5. [Figure 5] RP-LC-UV profiles of pertuzumab-compound (XI) conjugate (DAR=3.74) and neoDegrader P1 before and after treatment with the cysteine ​​protease papain are shown, as well as LCMS of the conjugate before and after treatment with the cysteine ​​protease papain. [Figure 6] Figure 1 shows the in vitro activity of representative conjugates against the BT-474 breast cancer cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of BT-474 cells when treated with pertuzumab-compound (X) conjugates (triangles, solid line) and rituximab-compound (X) conjugates (circles, dotted line). [Figure 7]1 shows the in vitro activity of representative conjugates against the NCI-N87 cancer cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of NCI-87 cells when treated with pertuzumab-compound (X) conjugates (triangles, solid line) and rituximab-compound (X) conjugates (circles, dotted line). [Figure 8] Figure 1 shows the in vitro activity of representative conjugates against the BT-474 breast cancer cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of BT-474 cells when treated with pertuzumab-compound (XI) conjugates (triangles) and rituximab-compound (XI) conjugates (circles). [Figure 9]

[0023] Figure 1 shows the in vitro activity of representative conjugates against the NCI-H929 cancer cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of NCI-H929 cells when treated with belantamab-compound (XIV) conjugate (circles), Synagis-compound (XIV) conjugate (squares), belantamab (upward triangles), and unconjugated compound (XIII) (downward triangles). [Figure 10] Figure 1 shows the in vitro activity of representative conjugates against Jurkat HiBiT labeled cells (HER2-) in the absence and presence of SK-BR-3 cells (HER2+). The X-axis shows the log payload concentration (M). The Y-axis shows the % viability of Jurkat HiBiT cells when treated with pertuzumab-compound (X) conjugates in the presence (circles, solid line) and absence (squares, dotted line) of SK-BR-3 cells. [Figure 11] 1 shows the in vitro activity of representative conjugates against the BT-474 breast cancer cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of BT-474 cells when treated with pertuzumab-compound (XVIII) conjugate (circles), pertuzumab-compound (XI) conjugate (squares), and rituximab-compound (XVIII) conjugate (triangles). [Figure 12]1 shows the in vitro activity of representative conjugates against the BT-474 breast cancer cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of BT-474 cells when treated with pertuzumab-compound (XLI) conjugate (circle), pertuzumab-compound (Ie) conjugate (square), pertuzumab-compound (XIX) conjugate (upward triangle), pertuzumab-compound (Ii) conjugate (downward triangle), pertuzumab-compound (Ia) conjugate (diamond), rituximab-compound (XLI) conjugate (hexagon), and rituximab-compound (XIX) conjugate (star). [Figure 13]

[0033] Figure 1 shows the in vitro activity of representative conjugates against the MV-411 AML cell line. The X-axis shows the log antibody concentration (M). The Y-axis shows the % viability of BT-474 cells when treated with gemtuzumab-compound (XL) conjugate (circle), gemtuzumab-compound (XL) conjugate + gemtuzumab (square), gemtuzumab-compound (XL) conjugate + trastuzumab (upward triangle), compound 40-3 (downward triangle), gemtuzumab (diamond), and trastuzumab (star). [Figure 14] 1 shows the degradation of BRD4 protein by compound (XL) and compound 40-3. [Figure 15A] Figure 1 shows the change in HCC1569 (breast cancer) tumor volume over time when treated with 3 mg / kg or 10 mg / kg of pertuzumab-compound (Ia) (squares and upward triangles, respectively), and 3 mg / kg or 10 mg / kg of pertuzumab-compound (XI) (downward triangles and diamonds, respectively). [Figure 15B] Figure 1 shows the change in body weight over time in mice bearing HCC1569 (breast cancer) tumors when treated with 3 mg / kg or 10 mg / kg of pertuzumab-compound (Ia) (squares and upward triangles, respectively), and 3 mg / kg or 10 mg / kg of pertuzumab-compound (XI) (downward triangles and diamonds, respectively). [Figure 16] SEC chromatogram of an anti-PSMA antibody with a DAR of 4 and a Cys-mutant-compound (XV) endogenous cysteine ​​conjugate. [Figure 17]1 shows the SEC chromatogram of J591 antibody with the S239C mutation-compound (XV) site-directed engineered cysteine ​​conjugate with a DAR of 1.85. [Figure 18] The HPLC chromatogram of compound (XIX) is shown. [Figure 19A] HPLC chromatogram of the reaction mixture when compound (XIX) was treated with cysteine: Compound (XIX) was completely consumed, and the only identified product had a retention time of 2.41 min. [Figure 19B] MS data of the peak at retention time 2.4 minutes corresponding to compound 18-6 is shown. [Figure 20] 1 shows the SEC chromatogram of the antibody HER2-A (wild-type sequence)-compound (XLII) conjugate with a DAR of 3.3. [Figure 21] 1 shows the SEC chromatogram of antibody HER2-A (containing cysteine ​​mutants for site-specific conjugation)-compound (XLII) conjugate with a DAR of 2.0. [Figure 22] The change in tumor volume over time of MV-4-11 tumors treated with 10 mg / kg of antibody CD33-D-compound (XL) conjugate (squares), 0.4 mg / kg of a BRD4 heterobifunctional degrader small molecule (compound 15 from Xiamg, W. et al., Biorganic Chemistry 2021, volume 115) (upward triangles), 10 mg / kg of ARV-825 (downward triangles), and vehicle (circles) is shown. [Figure 23] Individual tumor volumes over time for each group shown in Figure 22 are shown. [Figure 24] FIG. 23 shows the change in body weight over time in the mice used in the study shown in FIG. [Figure 25] 1 shows the CD79b binding affinity of antibody CD79b-A-compound (XLIII) conjugates of three different DARs (filled circles, upward pointing triangles, and diamonds), Synagis N297A (open circles), and antibody CD79b-A (squares). [Figure 26]1 shows a Western blot demonstrating degradation of IRAK4 (compared to a β-actin control) by antibody CD79b-A-compound (XLIII) conjugate, unconjugated payload, and unconjugated antibody CD79b-A. [Figure 27] 1 shows a Western blot showing the amount of IRAK4 (compared to a β-actin control) in the presence of antibody CD79b-A-compound (XLIII) conjugate, unconjugated antibody CD79b-A, or both antibody CD79b-A-compound (XLIII) conjugate and unconjugated antibody CD79b-A. DETAILED DESCRIPTION OF THE INVENTION

[0113] The present disclosure provides a compound of formula (XX): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein:

[0114] a is 1 to 10;

[0115] n is 0 or 1,

[0116] R 1 is a compound that induces protein-protein interactions,

[0117] R 2 is hydrogen, -(CH2CH2O) v -CH3, C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl), and v is 1 to 24;

[0118] each Y is independently S or O;

[0119] L is a cleavable linker,

[0120] Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0121] In some embodiments, R 2 is methyl.

[0122] The present disclosure further provides a compound of formula (XXXII): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein:

[0123] a is 1 to 10;

[0124] A' is [ka] or [ka] wherein:

[0125] n is 0 or 1,

[0126] each Y is independently S or O;

[0127] [ka] is R 1 indicates the point of attachment to

[0128] [ka] indicates the point of attachment to the methylene group,

[0129] R 1 is a compound that induces protein-protein interaction together with A',

[0130] R2 is hydrogen, R 1 -A', a group that provides stability to R 1 -A', a group that provides solubility to A', and R 1 -A' is selected from groups that provide stability and solubility to A';

[0131] L is a cleavable linker,

[0132] Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0133] The disclosure also provides compositions comprising the conjugates, methods of using such conjugates, and the above compounds conjugated to binding moieties.

[0134] The inclusion of a spacer capable of undergoing the retro-Mannich reaction described herein is a suitable general solution for linking and releasing gluturamide- or dihydrouracil-containing degradable agents to antibodies or other cell-binding agents. This technique avoids the need for additional chemical processing for antibody-based delivery to cancer cells.

[0135] I. Definition In order that this specification may be more readily understood, certain terms are first defined. Additional definitions are set forth throughout the detailed description.

[0136] It should be noted that the term "a" entity or "an" entity refers to one or more of that entity; for example, "a nucleotide sequence" is understood to represent one or more nucleotide sequences. Accordingly, the terms "a" (or "an"), "one or more," and "at least one" may be used interchangeably herein. It should further be noted that the claims may be drafted to exclude any optional element. Accordingly, this statement is intended to be used as a basis for using limiting language such as "only," "only," and the like in connection with the recitation of claim elements or as a predicate for using "negative" limitations.

[0137] Furthermore, when "and / or" is used herein, it should be construed as a specific disclosure of each of the two specified features or components, with or without the other. Thus, the term "and / or" when used herein in expressions such as "A and / or B" is intended to include "A and B," "A or B," "A" (alone), and "B" (alone). Similarly, the term "and / or" when used in expressions such as "A, B, and / or C" is intended to encompass each of the following aspects: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0138] Wherever embodiments are described herein using the word "comprising," it is understood that other similar embodiments described with the terms "consisting of" and / or "consisting essentially of" are also provided.

[0139] Unless otherwise defined, 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 disclosure pertains. For example, *Concise Dictionary of Biomedicine and Molecular Biology*, Juo, Pei-Show, 2nd ed., 2002, CRC Press; *The Dictionary of Cell and Molecular Biology*, 3rd ed., 1999, Academic Press; and *Oxford Dictionary of Biochemistry And Molecular Biology*, Revised, 2000, Oxford University Press provide those of ordinary skill in the art with a general dictionary of many of the terms used in this disclosure.

[0140] Units, prefixes, and symbols are indicated in the format accepted by the Systeme International de Unites (SI). Numeric ranges are inclusive of the numbers defining the range. When a range of values ​​is described, it is to be understood that each intervening integer and fractional value between the stated upper and lower limits of that range is also specifically disclosed, along with each subrange between such values. The upper and lower limits of any range may independently be included or excluded, and ranges where either limit, neither limit, or both limits are included are also encompassed within the disclosure. Thus, ranges described herein are understood to be shorthand notations for any value within that range, including the recited endpoints.

[0141] Where a value is explicitly recited, it should be understood that values ​​of approximately the same quantity or amount as the recited value are also within the scope of the disclosure. Where a combination is disclosed, each subcombination of the elements of that combination is also specifically disclosed and within the scope of the disclosure. Conversely, where different elements or groups of elements are individually disclosed, combinations thereof are also disclosed. Where any element of the disclosure is disclosed as having multiple alternatives, examples of that disclosure in which each alternative is excluded, alone or in any combination with other alternatives, are also hereby disclosed; multiple elements of the disclosure may have such exclusions, and all combinations of elements with such exclusions are hereby disclosed.

[0142] As used herein, the terms "targeted protein degrader" and "neoDegrader" refer to molecules that form ternary complexes with E3 ubiquitin ligases and can target proteins for degradation. Examples include, but are not limited to, molecular glues and PROTACs. Examples of molecular glues include, but are not limited to, CC-90009, lenalidomide, pomalidomide, DKY709, and compound P1 described in WO2021 / 198965.

[0143] As used herein, the term "antibody" also refers to a full-length immunoglobulin molecule or an immunologically active portion of a full-length immunoglobulin molecule, i.e., a molecule that contains an antigen-binding site that immunospecifically binds to a target antigen of interest or a portion thereof, such targets including, but not limited to, cancer cells or cells that produce autoimmune antibodies associated with autoimmune diseases. The immunoglobulins disclosed herein can be of any type (e.g., IgG, IgE, IgM, IgD, and IgA), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass of immunoglobulin molecule. The immunoglobulin can be derived from any species. However, in one aspect, the immunoglobulin is of human, murine, or rabbit origin.

[0144] The term "single domain antibody", also known as nanobody, refers to an antibody fragment consisting of a single monomeric variable antibody domain with a molecular weight of about 12 kDa to about 15 kDa. Single body antibodies can be based on the heavy chain variable domain or the light chain. Examples of single domain antibodies include V H H fragment and V fragment NAR These include, but are not limited to, fragments.

[0145] An "antibody fragment" comprises a portion of an intact antibody, generally the antigen-binding or variable region thereof. Examples of antibody fragments include Fab, Fab', F(ab'), and Fv fragments; diabodies; linear antibodies; fragments produced by an Fab expression library; anti-idiotypic (anti-Id) antibodies; CDRs (complementarity-determining regions); and epitope-binding fragments of any of the above that immunospecifically bind to a cancer cell antigen, a viral antigen, or a microbial antigen; single-chain antibody molecules; and multispecific antibodies formed from antibody fragments.

[0146] An "intact antibody" is one which comprises an antigen-binding variable region and a light chain constant domain (CL) and heavy chain constant domains, CH1, CH2 and CH3. The constant domains may be native sequence constant domains (e.g., human native sequence constant domains) or amino acid sequence variants thereof.

[0147] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous population of antibodies, i.e., a population in which the individual antibodies comprising the population are identical except for possible naturally occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations that include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. In addition to their specificity, monoclonal antibodies are advantageous in that they can be synthesized uncontaminated by other antibodies. The modifier "monoclonal" indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies and is not to be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present disclosure may be produced by hybridoma or recombinant DNA methods. "Monoclonal antibodies" may also be isolated from phage antibody libraries.

[0148] As used herein, monoclonal antibodies specifically include "chimeric" antibodies in which a portion of the heavy and / or light chain is identical to or homologous to corresponding sequences in antibodies from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain(s) is identical to or homologous to corresponding sequences in antibodies from another species or belonging to another antibody class or subclass, so long as the desired biological activity is exhibited, as well as fragments of such antibodies. Chimeric antibodies of interest herein include "primatized" antibodies comprising antigen-binding variable domain sequences derived from a non-human primate (e.g., Old World Monkey, Ape, etc.) and human constant region sequences.

[0149] Various methods are used to generate monoclonal antibodies (MAbs). Hybridoma technology, which refers to a cloned cell line that produces a single type of antibody, uses cells from various species, including mice (murine), hamsters, rats, and humans. Another method for preparing MAbs uses genetic engineering, such as recombinant DNA technology. Monoclonal antibodies produced by these techniques include chimeric and humanized antibodies, among others. Chimeric antibodies combine DNA coding regions from multiple species. For example, in chimeric antibodies, the variable region may be derived from mice and the constant region from humans. Humanized antibodies are primarily derived from humans, although they contain nonhuman portions. Like chimeric antibodies, humanized antibodies may contain fully human constant regions. However, unlike chimeric antibodies, the variable region may be partially derived from humans. The nonhuman synthetic portions of humanized antibodies are often derived from the CDRs of mouse antibodies. In either case, these regions are crucial for enabling the antibody to recognize and bind to a specific antigen. While mouse antibodies are useful for diagnosis and short-term therapy, they cannot be administered to people long-term without increasing the risk of a harmful immunogenic reaction. This reaction, called human anti-mouse antibody (HAMA), occurs when the human immune system recognizes the mouse antibody as foreign and attacks it. The HAMA reaction can lead to toxic shock and even death.

[0150] Chimeric and humanized antibodies reduce the likelihood of a HAMA reaction by minimizing the non-human portion of the administered antibody, and may have the added benefit of activating secondary human immune responses such as antibody-dependent cellular cytotoxicity.

[0151] An intact antibody may have one or more "effector functions," which refer to biological activities attributable to the Fc region of an antibody (a native sequence Fc region or an amino acid sequence variant Fc region). Examples of antibody effector functions include C1q binding, complement-dependent cytotoxicity, Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, cell surface receptors (e.g., B cell receptors; BCR), etc.

[0152] Intact antibodies can be assigned to different "classes" depending on the amino acid sequences of the constant domains of their heavy chains. There are five major classes of intact antibodies: IgA, IgD, IgE, IgG, and IgM, and some of these can be further divided into "subclasses" (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA, and IgA2. The heavy-chain constant domains that correspond to the different classes of antibodies are called α, δ, ε, γ, and μ, respectively. The structures and three-dimensional configurations of the subunits of the different classes of immunoglobulins are well known.

[0153] The term "about" is used herein to mean approximately, roughly, in the region of, or within the region. When the term "about" is used in conjunction with a numerical range, the term modifies that range by extending the boundaries above and below the stated numerical values. In general, the term "about" can modify a numerical value by varying above and below the stated value, for example, by 10 percent above or below (high or low).

[0154] The terms "administration," "administering," and grammatical variations thereof refer to the introduction of a composition, such as an EV (e.g., exosome) of the present disclosure, into a subject via a pharmaceutically acceptable route. Introduction of a composition, such as an EV (e.g., exosome) of the present disclosure, into a subject may be by any suitable route, such as intratumoral, oral, pulmonary, intranasal, parenteral (intravenous, intraarterial, intramuscular, intraperitoneal, or subcutaneous), rectal, intralymphatic, intrathecal, periocular, or topical. Administration includes self-administration and administration by another. A suitable route of administration enables the composition or agent to perform its intended function. For example, if the suitable route is intravenous, the composition is administered by introducing the composition or agent into the subject's vein.

[0155] As used herein, the term "antibody" includes immunoglobulins (whether natural or partially or wholly synthetically produced) and fragments thereof. The term also encompasses any protein having a binding domain homologous to an immunoglobulin binding domain. "Antibody" further includes polypeptides comprising a framework region from an immunoglobulin gene or fragments thereof that specifically bind and recognize an antigen. The use of the term antibody is intended to encompass whole, polyclonal, monoclonal, and recombinant antibodies, fragments thereof, as well as single-chain antibodies, humanized antibodies, murine antibodies, mouse-human, mouse-primate, and primate-human chimeric monoclonal antibodies, anti-idiotypic antibodies, antibody fragments such as scFv, (scFv)2, Fab, Fab', and F(ab')2, F(ab1)2, Fv, dAb, and Fd fragments, diabodies, and antibody-related polypeptides. Antibodies also include bispecific and multispecific antibodies, so long as they exhibit the desired biological activity or function. In some aspects of the present disclosure, the biologically active molecule is an antibody or a molecule comprising an antigen-binding fragment thereof.

[0156] The terms "antibody drug conjugate" and "ADC" are used interchangeably and refer to an antibody linked, e.g., covalently, to a therapeutic agent (which may be referred to herein as a drug, agent, or active pharmaceutical ingredient) or agent. In some aspects of the present disclosure, the biologically active molecule is an antibody drug conjugate.

[0157] As used herein, the term "approximately" when applied to one or more values ​​of interest refers to a value similar to the stated reference value. In certain embodiments, unless otherwise stated or otherwise clear from the context, the term "approximately" refers to a range of values ​​that is 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (above or below) from the stated reference value (except where such number exceeds 100% of possible values).

[0158] A "conservative amino acid substitution" is a substitution in which the amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues with similar side chains have been defined in the art and include basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, if an amino acid in a polypeptide is replaced with another amino acid from the same side chain family, the substitution is considered conservative. In another embodiment, a series of amino acids can be conservatively replaced with a series of structurally similar side chain family members that differ in order and / or composition.

[0159] As used herein, the term "conserved" refers to nucleotide or amino acid residues of a given polynucleotide or polypeptide sequence, respectively, that occur unchanged in the same position in two or more sequences being compared. A relatively conserved nucleotide or amino acid is one that is more conserved between related sequences than a nucleotide or amino acid that appears elsewhere in the sequence.

[0160] In some embodiments, two or more sequences are said to be "completely conserved" or "identical" if they are 100% identical to each other. In some embodiments, two or more sequences are said to be "highly conserved" if they are at least about 70% identical, at least about 80% identical, at least about 90% identical, or at least about 95% identical to each other. In some embodiments, two or more sequences are said to be "conserved" if they are at least about 30% identical, at least about 40% identical, at least about 50% identical, at least about 60% identical, at least about 70% identical, at least about 80% identical, at least about 90% identical, or at least about 95% identical to each other. Sequence conservation can apply to the entire length of a polynucleotide or polypeptide, or to a portion, region, or feature thereof.

[0161] As used herein, the terms "linking" and "binding" are used interchangeably and refer to the covalent or non-covalent attachment, respectively, of two or more moieties, including one or more compounds that induce protein-protein interactions, to a binding moiety. In some embodiments, the linkage or bond can include a linker.

[0162] The term "amino acid sequence variant" refers to a polypeptide having an amino acid sequence that differs to some extent from a native sequence polypeptide. Typically, an amino acid sequence variant will have at least about 70% sequence identity with at least one receptor-binding domain of a native antibody or at least one ligand-binding domain of a native receptor, and will typically have at least about 80%, and more typically at least about 90%, sequence identity with such receptor-binding domain or ligand-binding domain. Amino acid sequence variants have substitutions, deletions, and / or insertions at certain positions within the amino acid sequence of the native amino acid sequence. Amino acids are designated by conventional names, single-letter codes, and three-letter codes.

[0163] "Sequence identity" is defined as the percentage of residues in amino acid sequence variants that are identical after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity. Methods and computer programs for alignment are well known in the art. One such computer program is "Align 2," created by Genentech, Inc., which was submitted with user documentation to the United States Copyright Office, Washington, DC 20559 on December 10, 1991.

[0164] The term "Fc receptor" or "FcR" is used to refer to a receptor that binds to the Fc region of an antibody. An exemplary FcR is a native-sequence human FcR. Additionally, an FcR can be one that binds IgG antibodies (gamma receptors) and includes receptors of the subclasses FcγRI, FcγRII, and FcγRIII, including allelic variants and alternatively spliced ​​forms of these receptors. FcγRII receptors include FcγRIIA (an "activating receptor") and FcγRIIB (an "inhibiting receptor"), which have similar amino acid sequences but differ primarily in their cytoplasmic domains. Activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. Inhibiting receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic domain. Other FcRs, including those yet to be identified, are encompassed by the term "FcR" herein. The term also includes the neonatal receptor FcRn, which is involved in the transport of maternal IgG to the fetus.

[0165] "Complement-dependent cytotoxicity" or "CDC" refers to the ability of a molecule to lyse a target in the presence of complement. The complement activation pathway is initiated by the binding of the first component of the complement system (C1q) to a molecule (e.g., an antibody) complexed with a cognate antigen. To assess complement activation, a CDC assay can be performed.

[0166] "Native antibodies" are typically heterotetrameric glycoproteins of about 150,000 daltons, composed of two identical light (L) chains and two identical heavy (H) chains. Each light chain is linked to a heavy chain by one covalent disulfide bond; the number of disulfide bonds varies among the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has a variable domain (VH) at one end followed by several constant domains. Each light chain has a variable domain (VL) at one end and a constant domain at the other end. The constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the light chain variable domain is aligned with the variable domain of the heavy chain. Particular amino acid residues are believed to form an interface between the light-chain variable domain and the heavy-chain variable domain.

[0167] The term "variable" refers to the fact that certain portions of the variable domains vary significantly in sequence among different antibodies and are responsible for the binding and specificity of each particular antibody to its specific antigen. However, variability is not evenly distributed throughout the variable domains of antibodies. In both the light-chain and heavy-chain variable domains, it is concentrated in three segments called hypervariable regions. The more highly conserved portions of the variable domains are called framework regions (FRs). Naturally occurring heavy and light-chain variable domains each contain four FRs that primarily adopt a β-sheet configuration, connected by three hypervariable regions that form loops that connect, and in some cases form part of, the β-sheet structure. The hypervariable regions of each chain are held in close proximity by the FRs and, together with the hypervariable regions of the other chain, contribute to the formation of the antigen-binding site of antibodies. The constant domains are not directly involved in binding an antibody to an antigen but exhibit various effector functions, such as the participation of antibodies in antibody-dependent cellular cytotoxicity (ADCC).

[0168] The term "hypervariable region" as used herein refers to the amino acid residues of an antibody that are involved in antigen binding. Hypervariable regions generally include amino acid residues from the "complementarity determining regions" or "CDRs" (e.g., residues 24-34 (L1), 50-56 (L2), and 89-97 (L3) in the light-chain variable domain and residues 31-35 (H1), 50-65 (H2), and 95-102 (H3) in the heavy-chain variable domain; Kabat et al., supra) and / or residues from the "hypervariable loops" (e.g., residues 26-32 (L1), 50-52 (L2), and 91-96 (L3) in the light-chain variable domain and residues 26-32 (H1), 53-55 (H2), and 96-101 (H3) in the heavy-chain variable domain). "Framework region" or "FR" residues are those variable domain residues other than the hypervariable region residues as defined herein.

[0169] Papain digestion of antibodies produces two identical antigen-binding fragments called "Fab" fragments, each with a single antigen-binding site, and a residual "Fc" fragment, so named because of its ability to crystallize readily. Pepsin treatment yields an F(ab')2 fragment that has two antigen-binding sites and is still capable of cross-linking antigen.

[0170] An "Fv" is the minimum antibody fragment which contains a complete antigen-recognition and antigen-binding site. This region consists of a dimer of one heavy- and one light-chain variable domain in tight, non-covalent association. It is in this configuration that the three hypervariable regions of each variable domain interact to define an antigen-binding site on the surface of the VH-VL dimer. Together, the six hypervariable regions confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv, containing only three antigen-specific hypervariable regions) has the ability to recognize and bind antigen, although with lower affinity than the entire binding site.

[0171] Fab fragments also contain the constant domain of the light chain and the first constant domain (CH1) of the heavy chain. Fab' fragments differ from Fab fragments by the addition of a few residues at the carboxy terminus of the heavy chain CH1 domain, including one or more cysteines from the antibody hinge region. Fab'-SH is the designation used herein for Fab' in which the cysteine ​​residue(s) of the constant domains bear at least one free thiol group. F(ab')2 antibody fragments were originally produced as pairs of Fab' fragments with hinge cysteines between them. Other chemical couplings of antibody fragments are also known.

[0172] The "light chains" of antibodies from any vertebrate species can be assigned to one of two clearly distinct types, called kappa (κ) and lambda (λ), based on the amino acid sequences of their constant domains.

[0173] "Single-chain Fv" or "scFv" antibody fragments comprise the VH and VL domains of antibody, wherein these domains are present in a single polypeptide chain. The Fv polypeptide may further comprise a polypeptide linker between the VH and VL domains that enables the scFv to form the desired structure for antigen binding.

[0174] The term "diabody" refers to small antibody fragments with two antigen-binding sites, which comprise a variable heavy domain (VH) connected to a variable light domain (VL) on the same polypeptide chain (VH-VL). By using a linker that is too short to allow pairing between the two domains on the same chain, the domains are forced to pair with the complementary domains of another chain and create two antigen-binding sites.

[0175] "Humanized" forms of non-human (e.g., rodent) antibodies are chimeric antibodies that contain minimal sequence derived from non-human immunoglobulin. Humanization is a method for transferring murine antigen-binding information into non-immunogenic human antibody acceptors, resulting in many therapeutically useful drugs. The humanization process generally begins with the transfer of all six murine complementarity-determining regions (CDRs) onto a human antibody framework. These CDR-grafted antibodies generally do not retain their original affinity for antigen binding; in fact, affinity is often significantly impaired. In addition to the CDRs, select non-human antibody framework residues must also be incorporated to maintain proper CDR conformation. Introduction of key murine framework residues into the human acceptor to support the structural conformation of the CDRs has been shown to restore antigen binding and affinity. For the most part, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a hypervariable region of the recipient are replaced by residues from a hypervariable region of a non-human species (donor antibody) such as mouse, rat, rabbit, or non-human primate having the desired specificity, affinity, and capacity. In some instances, framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, humanized antibodies may comprise residues that are not found in the recipient or donor antibody. These modifications are made to improve antibody performance. In general, humanized antibodies will comprise substantially all of at least one, and typically two, variable domains, with all or substantially all of the hypervariable loops corresponding to those of a non-human immunoglobulin and all or substantially all of the FRs being those of a human immunoglobulin sequence. The humanized antibody also optionally will comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin.

[0176] An "isolated" antibody is one that has been identified and separated and / or recovered from a component of its natural environment. Contaminant components of its natural environment are substances that would interfere with diagnostic or therapeutic uses for the antibody, and may include enzymes, hormones, and other proteinaceous or nonproteinaceous solutes. In certain aspects, the antibody is (1) purified to greater than 95% by weight or greater than 99% by weight as determined by the Lowry method, (2) purified sufficiently to obtain at least 15 residues of N-terminal or internal amino acid sequence by use of a gas-phase protein sequencer, or (3) purified to homogeneity by SDS-PAGE under reducing or non-reducing conditions using Coomassie blue or silver staining. Isolated antibodies include antibodies in situ within recombinant cells, since at least one component of the antibody's natural environment will not be present. Ordinarily, however, isolated antibodies will be prepared by at least one purification step.

[0177] "Cancer" refers to a broad group of diverse diseases characterized by the uncontrolled growth of abnormal cells in the body. Unregulated cell division and growth leads to the formation of malignant tumors that can invade neighboring tissues and metastasize to distant parts of the body via the lymphatic system or bloodstream. As used herein, "cancer" refers to primary, metastatic, and recurrent cancers.

[0178] As used herein, the term "immune response" refers to a biological response in a vertebrate to foreign substances, which protects the organism from these substances and the diseases they cause. Immune responses are mediated by the action of cells of the immune system (e.g., T lymphocytes, B lymphocytes, natural killer (NK) cells, macrophages, eosinophils, mast cells, dendritic cells, or neutrophils) and soluble macromolecules (including antibodies, cytokines, and complement) produced by either these cells or the liver, which result in the selective targeting, binding to, damaging, destroying, and / or eliminating from the vertebrate body invading pathogens, pathogen-infected cells or tissues, cancerous or other abnormal cells, or, in the case of autoimmunity or pathological inflammation, normal human cells or tissues. Immune responses include, for example, T cells, e.g., effector T cells or Th cells, e.g., CD4 + T cells or CD8 + These include activating or inhibiting T cells, or inhibiting Treg cells. As used herein, the terms "T cell" and "T lymphocyte" are synonymous and refer to any lymphocyte produced or processed by the thymus. In some embodiments, the T cell is a CD4+ T cell. In some embodiments, the T cell is a CD8+ T cell. In some embodiments, the T cell is a NKT cell.

[0179] A "subject" includes any human or non-human animal. The term "non-human animal" includes, but is not limited to, vertebrates such as non-human primates, sheep, dogs, and rodents such as mice, rats, and guinea pigs. In some embodiments, the subject is a human. The terms "subject" and "patient" are used interchangeably herein.

[0180] The term "therapeutically effective amount" or "therapeutically effective dosage" refers to an amount of an agent (e.g., a conjugate disclosed herein) that provides a desired biological, therapeutic, and / or prophylactic result. The result can be a decrease, amelioration, alleviation, reduction, delay, and / or alleviation of one or more of the signs, symptoms, or causes of a disease, or any other desired alteration of a biological system. With respect to solid tumors, an effective amount includes an amount sufficient to shrink the tumor and / or reduce the rate of tumor growth (such as inhibiting tumor growth) or prevent or delay other undesirable cell proliferation. In some embodiments, an effective amount is an amount sufficient to slow tumor growth. In some embodiments, an effective amount is an amount sufficient to prevent or delay tumor recurrence. An effective amount can be administered in one or more administrations. An effective amount of the composition can, for example, (i) reduce the number of cancer cells, (ii) reduce tumor size, (iii) inhibit, delay, slow, or stop to some extent cancer cell invasion into peripheral organs, (iv) inhibit (i.e., slow to some extent and stop) tumor metastasis, (v) inhibit tumor growth, (vi) prevent or delay the onset and / or recurrence of tumors, and / or (vii) alleviate to some extent one or more symptoms associated with cancer.

[0181] In some embodiments, a "therapeutically effective amount" is the amount of conjugate clinically proven to affect a significant reduction in cancer or a slowing of progression (regression) of cancer, such as an advanced solid tumor. The ability of a therapeutic agent to promote disease regression can be assessed using a variety of methods known to those of skill in the art, for example, by assessing the activity of the agent in human subjects in clinical trials, in animal model systems predictive of efficacy in humans, or in in vitro assays.

[0182] As used herein, the term "standard of care" refers to a treatment that is accepted by medical professionals as an appropriate treatment for a particular type of disease and that is widely used by medical professionals. This term can be used interchangeably with any of the terms "best medical care," "standard of care," and "standard therapy."

[0183] By way of example, an "anti-cancer drug" promotes regression of cancer or prevents further tumor growth in a subject. In certain embodiments, a therapeutically effective amount of the drug promotes regression of cancer to the point where the cancer disappears.

[0184] The terms "effective" and "efficacy" in reference to treatment include both pharmacological effectiveness and physiological safety. Pharmacological effectiveness refers to the ability of a drug to promote cancer regression in a patient. Physiological safety refers to the level of toxicity or other adverse physiological effects (adverse effects) at the cellular, organ, and / or organism level resulting from the administration of the drug.

[0185] As used herein, the term "immune checkpoint inhibitor" refers to a molecule that completely or partially reduces, inhibits, interferes with, or modulates one or more checkpoint proteins. Checkpoint proteins control the activation or function of T cells. Many checkpoint proteins are known, such as CTLA-4 and its ligands CD80 and CD86; and PD-1 and its ligands PD-L1 and PD-L2. Pardoll, DM, Nat Rev Cancer 12(4):252-64(2012). These proteins are involved in costimulatory or inhibitory interactions in T cell responses. Immune checkpoint proteins control and maintain self-tolerance and the duration and magnitude of physiological immune responses. Immune checkpoint inhibitors include or are derived from antibodies.

[0186] The term "treat" or "treatment" refers to both therapeutic treatment and prophylactic or preventative measures, the purpose of which is to prevent or slow (reduce) the development or spread of an undesirable physiological change or disorder, such as cancer. For purposes of this disclosure, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, attenuation of the extent of the disease, stabilized (i.e., not worsening) disease, delayed or slowed disease progression, improvement or palliation of the disease state, and remission (whether partial or complete), whether detectable or undetectable. "Treatment" can also mean prolonging survival as compared to expected survival if not receiving treatment. Those in need of treatment include those already with the condition or disorder, as well as those prone to have the condition or disorder or those in whom the condition or disorder is to be prevented.

[0187] II. Protein-protein interaction inducers The present disclosure provides a compound of formula (XX): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein R 1 is a compound that induces protein-protein interactions.

[0188] The present disclosure further provides a compound of formula (XXXII): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein:

[0189] A' is [ka] or [ka] wherein:

[0190] n is 0 or 1,

[0191] each Y is independently S or O;

[0192] [ka] is R 1 indicates the point of attachment to

[0193] [ka] indicates the point of attachment to the methylene group,

[0194] R 1 is a compound that, together with A', induces protein-protein interaction.

[0195] Compounds that induce protein-protein interactions include protein-protein interaction regulators such as those described in Biophysical Reviews 2019, 11:559-581.

[0196] In certain embodiments, the protein-protein interaction inducer comprises a targeted protein degrader that can disaggregate and degrade unwanted proteins.

[0197] In some embodiments, the targeted protein degrader comprises a substituted isoindole compound. In some embodiments, the targeted protein degrader comprises a 5'-substituted isoindole compound. In certain embodiments, R 1 is the formula (XXX) shown below: [ka] (XXX) is a compound of During the ceremony,

[0198] [ka] is the point of attachment to the parent molecular moiety,

[0199] A is phenyl or C4-C 10 is a cycloalkyl ring,

[0200] U is selected from NH and CF;

[0201] R 10 is independently selected from hydrogen and halo;

[0202] R 20 -CH3, -C(O)R 3 , -N(R 4 )2, -(CH2) n OH, -(CH2) n N(R 4 )2, -(CH2) n Q'(CH2) m OH, -(CH2) n Q'(CH2) m SH, and -(CH2) n Q'(CH2) m N(R 4 )2, wherein:

[0203] R 3 is hydrogen or C1-C6 alkyl,

[0204] Each R 4 are independently hydrogen or C1-C6 alkyl;

[0205] Q' is O, S, or NR 4 and

[0206] n is 1 to 6;

[0207] m is 2 to 5.

[0208] In certain aspects, the present disclosure provides a compound of formula (XXX), or a pharmaceutically acceptable salt thereof, wherein:

[0209] A is a phenyl ring or a C4-C 10is a cycloalkyl ring,

[0210] U is NH,

[0211] R 10 is selected from hydrogen and halo;

[0212] R 20 is -(CH2) n Q'(CH2) m N(R 4 )2, -(CH2) n OH, -N(R 4 )2, and -C(O)R 3 is selected from:

[0213] m is 2,

[0214] n is 2,

[0215] Q' is -O-;

[0216] R 3 is methyl,

[0217] Each R 4 is independently selected from hydrogen and methyl.

[0218] As used herein, the term "C1-C6 alkoxy," as used herein, refers to a C1-C6 alkyl group attached to the parent molecular moiety through an oxygen atom.

[0219] The term "C1-C6 alkoxy C1-C6 alkyl," as used herein, refers to a C1-C6 alkoxy group attached to the parent molecular moiety through a C1-C6 alkyl group.

[0220] As used herein, the term "C1-C6 alkyl" refers to a group derived from a straight or branched chain saturated hydrocarbon containing from 1 to 6 carbon atoms.

[0221] As used herein, "C4-C 10 The term "cycloalkyl" refers to a saturated monocyclic hydrocarbon ring system having from 4 to 10 carbon atoms and 0 heteroatoms. Representative examples of cycloalkyl groups include, but are not limited to, cyclobutyl, cyclopentyl, and cyclohexyl. Cycloalkyl groups containing from 7 to 10 atoms can be monocyclic or fused, spirocyclic, or bridged bicyclic structures.

[0222] As used herein, the term "halo" refers to F, Cl, Br, or I.

[0223] In some embodiments, the compound of formula (XXX) is [ka] The compound is selected from the group consisting of:

[0224] In some embodiments, the targeted proteolytic agent comprises a proteolysis-directed chimera (PROTAC). Examples of PROTACs are known in the art (see, e.g., Acta Pharmaceutica Sinica B, 2020; 10(2): 207-238).

[0225] In certain embodiments, the PROTAC has the formula: POI-L 100 -CBN wherein

[0226] POI is a compound that binds to a protein of interest,

[0227] L 100 is the PROTAC linker,

[0228] CBN is the binding moiety of cereblon.

[0229] Several different protein classes have been reported as targets for PROTACs. In certain embodiments, the protein of interest may be a nuclear hormone receptor, a translation termination factor, a transcription factor, a cyclin-dependent kinase, a tyrosine kinase, a serine / threonine kinase, or an E3 ligase.

[0230] Among various protein classes, there are several proteins of interest that can be targeted by the PROTACS methods described herein. In certain embodiments, the protein of interest can be selected from CD33, GSPT1, BRD4, androgen receptor (AR), estrogen receptor (ER), IKZF1 / 3, CK1a, BCL-XL, IKZF2, IRAK4, BTK, STAT3, BTK and iMiD, BRD9, TRK, MDM2, CDK2 / CDK9, CD97b, and EGFR. In some embodiments, the protein of interest can be selected from BRD4, ER, and IRAK4. In some embodiments, the protein of interest can be selected from BTK, BRD9, TRK, CDK2 / CDK9, and STAT3.

[0231] In certain embodiments, the PROTAC comprises a linker L 100 PROTAC linkers have been well studied in the art (see, e.g., Troup RI, Fallan C, Baud MGJ. "Current strategies for the design of PROTAC linkers: a critical review." Explor Target Antitumor Ther. 2020;1:273-312. https: / / doi.org / 10.37349 / etat.2020.00018).

[0232] In certain embodiments, L 100can comprise an alkyl linker. In some embodiments, the alkyl linker can comprise 2 to 30 atoms. In some embodiments, the alkyl linker can comprise 5 to 25 atoms. In some embodiments, the alkyl linker can comprise 10 to 20 atoms. In some embodiments, the alkyl linker can comprise 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 atoms.

[0233] In certain embodiments, L 100 can comprise a glycol linker. In some embodiments, the glycol linker can comprise 3 to 30 atoms. In some embodiments, the glycol linker can comprise 5 to 25 atoms. In some embodiments, the alkyl linker can comprise 10 to 20 atoms. In some embodiments, the alkyl linker can comprise 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 atoms.

[0234] In certain embodiments, L 100 can comprise a glycol and an alkyl linker. In some embodiments, the linker can comprise 5 to 35 atoms. In some embodiments, the alkyl linker can comprise 10 to 30 atoms. In some embodiments, the alkyl linker can comprise 15 to 25 atoms. In some embodiments, the alkyl linker can comprise 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 atoms.

[0235] In certain embodiments, L 100 can comprise one or more functional groups selected from polyethylene glycol (PEG), alternative glycol groups such as propylene glycol, alkyl, alkynyl, triazolyl, piperazinyl, piperidinyl, and mixtures thereof. It should be understood that suitable PROTAC linkers can be selected using methods known to those of skill in the art. In some embodiments, the linker can comprise 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 atoms.

[0236] Typically, in certain embodiments, a PROTAC can comprise a cereblon-binding moiety (CNB). In some aspects, the cereblon-binding moiety is [ka] may be selected from During the ceremony, [ka] indicates the point of attachment to A', [ka] L 100 indicates the attachment point to

[0237] In some embodiments, the PROTAC has the formula:

[0238] [ka] [ka] [ka] [ka] and

[0239] During the ceremony, [ka] indicates the point of attachment to A'.

[0240] III. Stability and Solubility Enhancers The stability and / or solubility of the conjugates described herein may be determined by the R 2 In certain embodiments, when improved stability and / or solubility is not required, functionalization at R 2 R can be hydrogen. In other embodiments, additional stability and / or solubility is desired. 2 can be a group other than hydrogen.

[0241] In certain embodiments, R 2 can be a group that confers stability to the complex. In some embodiments, R 2 is selected from C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl (C1-C3 alkyl). In some embodiments, R 2 is C1-C6 alkyl. In some embodiments, R 2 is methyl.

[0242] In certain embodiments, R 2 can be a group that confers solubility to the complex. In some embodiments, R 2 teeth, [ka] [ka] may be selected from During the ceremony, each n is independently 1, 2, 3, 4, or 5; each y is independently 1 or 2; Each R is independently hydrogen, CH 11 O5, C 12 H 21 O 10 , C 18 H 31 O 15 , or C 24 H 41 O 20 is.

[0243] IV. Complex The present disclosure provides conjugates of one or more protein-protein interaction inducers, a linker, and a binding moiety.

[0244] In some aspects, the present disclosure provides a compound of formula (I): [ka] The compound or a pharmaceutically acceptable salt thereof, wherein:

[0245] a is 1 to 10;

[0246] n is 0 or 1,

[0247] R 1 is a compound that induces protein-protein interactions,

[0248] R 2 is hydrogen, -(CH2CH2O) v -CH3, C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl), and v is 1 to 24;

[0249] each Y is independently S or O;

[0250] L is a cleavable linker,

[0251] Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0252] In some embodiments, the present disclosure provides a compound of formula (XXXII): [ka] A complex of or a pharmaceutically acceptable salt thereof, wherein:

[0253] a is 1 to 10;

[0254] A' is [ka] or [ka] wherein:

[0255] n is 0 or 1,

[0256] each Y is independently S or O;

[0257] [ka] is R 1 indicates the point of attachment to

[0258] [ka] indicates the point of attachment to the methylene group,

[0259] R 1 is a compound that induces protein-protein interaction together with A',

[0260] R 2 is hydrogen, R 1 -A', a group that provides stability to R 1-A', a group that provides solubility to A', and R 1 -A' is selected from groups that provide stability and solubility to A';

[0261] L is a cleavable linker,

[0262] Bm is a binding moiety capable of specifically binding to a protein. In some embodiments, the protein is on a cell surface.

[0263] In some embodiments, the protein to which Bm binds is HER2, and R 1 In some embodiments, the protein that Bm binds to is CD33, and R 1 Bm binds to mouse double minute 2 homolog (MDM2). In some embodiments, the protein that Bm binds is prostate-specific membrane antigen (PSMA), and R 1 Bm binds to the androgen receptor (AR). In some embodiments, the protein to which Bm binds is CD33, and R 1 binds to bromodomain-containing protein 4 (BRD4). In some embodiments, the protein to which Bm binds is CD33, and R 1 In some embodiments, the protein that Bm binds to is CD79b, and R 1 In some embodiments, the protein that Bm binds to is HER2, and R 1 In some embodiments, the protein that Bm binds to is BCMA, and R 1 In some embodiments, the protein that Bm binds to is HER2, and R 1 binds to the ER.

[0264] In some embodiments, the complexes described herein have in vitro antiproliferative activity against tumor cell lines. In some embodiments, complexes comprising one or more compounds that induce protein-protein interactions and binding moieties have in vitro antiproliferative activity that is at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, or at least about 100% greater than the compound(s) or binding moiety alone. In some embodiments, complexes comprising one or more compounds that induce protein-protein interactions and binding moieties have in vitro antiproliferative activity that is at least about 2-fold, at least about 3-fold, at least about 4-fold, at least about 5-fold, at least about 6-fold, at least about 7-fold, at least about 8-fold, at least about 9-fold, or at least about 10-fold greater than the compound(s) or binding moiety alone.

[0265] In some embodiments, a complex comprising one or more compounds that induce protein-protein interactions and a binding moiety has in vitro anti-proliferative activity against a BT-474 breast cancer cell line, e.g., higher anti-proliferative activity against a BT-474 breast cancer cell line, compared to the compound(s) alone or the binding moiety alone. In some embodiments, a complex comprising one or more compounds that induce protein-protein interactions and a binding moiety has in vitro anti-proliferative activity against a SK-BR-3 breast cancer cell line, e.g., higher anti-proliferative activity against a SK-BR-3 breast cancer cell line, compared to the compound(s) alone or the binding moiety alone. In some embodiments, a complex comprising one or more compounds that induce protein-protein interactions and a binding moiety has in vitro anti-proliferative activity against a NCI-N87 gastric cancer cell line, e.g., higher anti-proliferative activity against a NCI-N87 gastric cancer cell line, compared to the compound(s) alone or the binding moiety alone. In some embodiments, a complex comprising one or more compounds that induce protein-protein interactions and a binding moiety has in vitro antiproliferative activity against the lymphoma cell line Daudi, e.g., higher antiproliferative activity against the lymphoma cell line Daudi, compared to the compound(s) alone or the binding moiety alone. In some embodiments, a complex comprising one or more compounds that induce protein-protein interactions and a binding moiety has in vitro antiproliferative activity against the HL-60 acute myeloid leukemia cell line, e.g., higher antiproliferative activity against the HL-60 acute myeloid leukemia cell line, compared to the compound(s) or the binding moiety alone. In some embodiments, a complex comprising one or more compounds that induce protein-protein interactions and a binding moiety has in vitro antiproliferative activity against the non-Hodgkin's lymphoma cell line Ramos, e.g., higher antiproliferative activity against the non-Hodgkin's lymphoma cell line Ramos, compared to the compound(s) alone or the binding moiety alone. In some embodiments, the complexes described herein can maintain their antiproliferative activity in the presence of human serum. In some aspects, the conjugates described herein can be used in the treatment of cancer.

[0266] In some embodiments, the conjugates provided herein can be used to treat breast cancer, gastric cancer, non-small cell lung cancer, cholangiocarcinoma, colon cancer, ovarian cancer, or neuregulin-1 (NRG1) positive cancer, for example, the protein to which Bm binds is HER2 and R 1 In some embodiments, the conjugates provided herein can be used to treat acute myeloid leukemia, for example, when the protein to which Bm binds is CD33 and R 1 R binds to MDM2, GSPT1, or bromodomain-containing protein 4 (BRD4). In some embodiments, the conjugates provided herein can be used to treat prostate cancer, for example, when the protein to which Bm binds is prostate-specific membrane antigen (PSMA), R 1 In some embodiments, the conjugates provided herein can be used to treat NHL, e.g., B-cell NHL or DLBCL, e.g., the protein to which Bm binds is CD79b and AR binds. 1 binds to IRAK4.

[0267] III.A. Linkers The compound(s) that induces protein-protein interaction can be linked to the binding moiety via a glutarmide or dihydrouracil ring as described herein. As used herein, the term "linker" refers to any chemical moiety that can connect the binding moiety (Bm) to the nitrogen atom of the glutaramide or dihydrouracil ring in a compound of formula (XX) or formula (XXX).

[0268] In certain embodiments, the linker can contain a heterobifunctional group. In the present disclosure, the term "heterobifunctional group" refers to a chemical moiety that connects the linker of which it is a part to the binding moiety. A heterobifunctional group is characterized by having different reactive groups at both ends of the chemical moiety. Conjugation to "Bm" can be achieved by chemical or enzymatic conjugation, or a combination of both. Chemical conjugation involves the controlled reaction of accessible amino acid residues on the surface of the binding moiety with reactive handles on the heterobifunctional group. Examples of chemical conjugation include, but are not limited to, lysine amide coupling, cysteine ​​coupling, and coupling via genetically engineered non-natural amino acids, where a non-natural amino acid residue bearing a desired reactive handle is placed on "Bm." In enzymatic conjugation, an enzyme mediates the coupling of the linker to an accessible amino acid residue on the binding moiety. Examples of enzymatic conjugation include, but are not limited to, transpeptidation using sortase, transpeptidation using microbial transglutaminase, and N-glycan engineering. Chemical and enzymatic conjugation may also be used sequentially. For example, enzymatic conjugation can also be used to place unique reactive handles on "Bm" that are utilized in subsequent chemical conjugation.

[0269] In some aspects, the heterobifunctional group is [ka] is selected from During the ceremony, [ka] is the point of attachment to the rest of the linker, [ka] is the point of attachment to Bm.

[0270] In certain aspects, the linker may be cleavable. In some aspects, the linker may be susceptible to acid-induced cleavage, photo-induced cleavage, bioreductive cleavage, enzymatic cleavage, etc., under conditions that allow the compound(s) inducing the protein-protein interaction and / or the binding moiety to remain active.

[0271] In some embodiments, the cleavable linker can be cleaved by an enzyme, ie, a protease, peptidase, esterase, β-glucuronidase, glycosidase, phosphodiesterase, phosphatase, pyrophosphatase, or lipase.

[0272] In some embodiments, the cleavable linker can be cleaved by a protease, examples of which include, but are not limited to, cathepsin B, VAGP tetrapeptide, and the like.

[0273] In certain embodiments, the cleavable linker contains a peptide. In some embodiments, the peptide is the cleavage site of the linker, thereby facilitating drug release upon exposure to intracellular proteases, such as lysosomal enzymes. Peptides can be designed and optimized for enzymatic cleavage by specific enzymes, such as tumor-associated proteases, cathepsins B, C, and D, or plasmin proteases. Examples of peptides containing two amino acids include, but are not limited to, alanine-alanine (ala-ala), valine-alanine (val-ala), valine-citrulline (vc or val-cit), alanine-phenylalanine (af or ala-phe); phenylalanine-lysine (fk or phe-lys); phenylalanine-homlysine (phe-homolys); and N-methyl-valine-citrulline (Me-val-cit). Examples of peptides having three amino acids include, but are not limited to, glycine-valine-citrulline (gly-val-cit), aspartic acid-valine-citrulline (asp-val-cit), alanine-alanine-asparagine (ala-Ala-asn), alanine-phenylalanine-lysine (ala-phe-lys), glycine-glycine-phenylalanine (gly-gly-phe), and glycine-glycine-glycine (gly-gly-gly). Examples of peptides having four amino acids include, but are not limited to, glycine-glycine-valine-citrulline (gly-gly-val-cit) and glycine-glycine-phenylalanine-glycine (gly-gly-phe-gly). Examples of peptides having five amino acids include, but are not limited to, glycine-glycine-valine-citrulline-glycine (gly-gly-val-cit-gly) and glycine-glycine-phenylalanine-glycine-glycine (gly-gly-phe-gly-gly). The above amino acid combinations can also occur in the reverse order (i.e., cit-val).

[0274] The peptides of the present disclosure can include L- or D-isomers of amino acid residues. The term "naturally occurring amino acid" refers to Ala, Asp, Asx, Cit, Cys, Glu, Phe, Glx, Gly, His, Ile, Lys, Leu, Met, Asn, Pro, Gln, Arg, Ser, Thr, Val, Trp, and Tyr. "D-" denotes an amino acid having a "D" (dextrorotatory) configuration, as opposed to the configuration in naturally occurring ("L-") amino acids. The amino acids described herein can be purchased commercially (Sigma Chemical Co., Advanced Chemtech) or synthesized using methods known in the art.

[0275] In certain embodiments, the linker (“L”) is [ka] is a protease-cleavable linker selected from During the ceremony,

[0276] q is 2 to 10;

[0277] Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are each independently absent or a naturally occurring amino acid residue in the L-configuration or the D-configuration, with the proviso that Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are amino acid residues,

[0278] [ka] is the point of attachment to the parent molecular moiety,

[0279] [ka] is the point of attachment to the binding moiety.

[0280] In certain embodiments, Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 is independently absent or selected from the group consisting of L-valine, D-valine, L-citrulline, D-citrulline, L-alanine, D-alanine, L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-asparagine, D-asparagine, L-phenylalanine, D-phenylalanine, L-lysine, D-lysine, and glycine, with the proviso that Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 At least two of the residues are amino acid residues.

[0281] In some embodiments, Z 1 is absent or is glycine, and Z 2 is absent or is selected from L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-alanine, D-alanine, and glycine; Z 3 is selected from L-valine, D-valine, L-alanine, D-alanine, L-phenylalanine, D-phenylalanine, and glycine; Z 4 is selected from L-citrulline, D-citrulline, L-asparagine, D-asparagine, L-lysine, D-lysine, L-phenylalanine, D-phenylalanine, and glycine; Z 5 is absent or is glycine.

[0282] In some embodiments, L is [ka] is.

[0283] In some embodiments, q is 4.

[0284] In certain embodiments, L is a beta-glucuronidase cleavable linker.

[0285] In some embodiments, L is [ka] is a beta-glucuronidase cleavable linker which is During the ceremony,

[0286] q is 2 to 10;

[0287] ---- is absent or a bond,

[0288] [ka] is the point of attachment to the parent molecular moiety,

[0289] [ka] is the point of attachment to the binding moiety.

[0290] In some embodiments, the linker is bioreductive. The bioreductive linker utilizes the difference between the reduction potential of intracellular compartments and that of plasma. The reduced glutathione present in the cytoplasm of tumor cells is up to 1000 times higher than that present in the cytoplasm of normal cells, and tumor cells also contain enzymes that can contribute to reduction in the cellular compartment. The linker keeps the conjugate intact during systemic circulation, and is selectively cleaved by the high concentration of intracellular glutathione, releasing the active drug from the non-toxic prodrug at the tumor site.

[0291] In some embodiments, L is [ka] is a bioreducible linker selected from the group consisting of:

[0292] q is 2 to 10;

[0293] R, R', R'', and R''' are each independently selected from hydrogen, C-C alkoxyC-C alkyl, (C-C) NC-C alkyl, and C-C alkyl, or two geminal R groups together with the carbon atoms to which they are attached can form a cyclobutyl or cyclopropyl ring;

[0294] [ka] is the point of attachment to the parent molecular moiety,

[0295] [ka] is the point of attachment to the binding moiety.

[0296] In certain embodiments, L is [ka] is a bioreducible linker

[0297] In certain embodiments, L is a click-to-release linker, in which case release of the compound that induces protein-protein interactions is chemically triggered by a tetrazine or related compound.

[0298] In some embodiments, L is [ka] a click-to-release linker which is During the ceremony,

[0299] q is 2 to 10;

[0300] [ka] is the point of attachment to the parent molecular moiety,

[0301] [ka] is the point of attachment to the binding moiety.

[0302] In some embodiments, the point of attachment to the binding moiety is a cysteine, lysine, tyrosine, or glutamine within the binding moiety. In some embodiments, the point of attachment to the binding moiety is a cysteine. In some embodiments, the point of attachment to the binding moiety is a lysine. In some embodiments, the point of attachment to the binding moiety is a tyrosine. In some embodiments, the point of attachment to the binding moiety is a glutamine.

[0303] The cysteine ​​or lysine can be, for example, an engineered (i.e., not endogenous to the binding moiety) cysteine ​​or lysine for site-specific conjugation. Site-specific conjugation refers to conjugation via a unique, defined site on the binding moiety (e.g., an antibody or its antigen-binding portion). Site-specific conjugation is discussed, for example, in Zhou, Qun. "Site-Specific Antibody Conjugation for ADC and Beyond," Biomedicines vol. 5, 4 64. 9 Nov. 2017, doi:10.3390 / biomedicines5040064, the entire contents of which are incorporated herein by reference.

[0304] The attachment point cysteine ​​or lysine can be a cysteine ​​or lysine endogenous to the attachment moiety.

[0305] III.B. Joining part The present disclosure provides one or more compounds that induce protein-protein interactions conjugated to a binding moiety. As used herein, the term "binding moiety" refers to any molecule that recognizes and binds to a cell surface marker or receptor. In certain embodiments, the binding moiety binds to a protein, including but not limited to a polypeptide moiety. In addition to directing a compound(s) to a specific cell, tissue, or location, the binding moiety may also have a specific therapeutic effect, such as antiproliferative (cytostatic and / or cytotoxic) activity against the target cell or pathway. In certain embodiments, the binding moiety can include or be engineered to include at least one chemically reactive group, such as a carboxylic acid, amine, thiol, or a chemically reactive amino acid moiety or side chain. In some embodiments, the binding moiety can include a targeting moiety for a given target cell population that binds to or complexes with a cell surface molecule, such as a cell surface receptor or antigen. After specific binding or complex formation with the receptor, the cell is able to take up the targeting moiety or complex, which is then internalized within the cell.

[0306] In some embodiments, the "Bm" group can be a peptide or protein that binds to a cell surface receptor or antigen.

[0307] In certain embodiments, the "Bm" group can be an antibody, antibody fragment, or antigen-binding fragment. Antibodies are proteins produced by the immune system that can recognize and bind to a specific antigen. A target antigen generally has multiple binding sites, also called epitopes, recognized by CDRs on multiple antibodies. Each antibody that specifically binds to a different epitope has a different structure. Thus, there can be multiple corresponding antibodies for one antigen. The term "antibody" is used in the broadest sense herein and specifically encompasses monoclonal antibodies, single-domain antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, so long as they exhibit the desired biological activity. Antibodies can be murine, human, humanized, chimeric, or derived from other species.

[0308] Monoclonal antibodies, which can be conjugated to a compound(s), are homogeneous populations of antibodies directed against a particular antigenic determinant (e.g., cancer cell antigens, viral antigens, microbial antigens, proteins, peptides, carbohydrates, chemicals, nucleic acids, or fragments thereof). Monoclonal antibodies (mAbs) directed against an antigen of interest can be prepared by using any technique known in the art that provides for the production of antibody molecules by continuous cell lines in culture. These include, but are not limited to, hybridoma technology, human B-cell hybridoma technology, and EBV hybridoma technology. Such antibodies can be of any immunoglobulin class, including IgG, IgM, IgE, IgA, and IgD, and any subclass thereof. Hybridomas producing mAbs used in the present disclosure can be cultured in vitro or in vivo.

[0309] Useful monoclonal antibodies include, but are not limited to, human monoclonal antibodies, humanized monoclonal antibodies, antibody fragments, or human-mouse (or other species) chimeric monoclonal antibodies. Human monoclonal antibodies can be made by any of a number of techniques known in the art.

[0310] The antibody may also be a bispecific antibody. Methods for producing bispecific antibodies are known in the art. Traditional production of full-length bispecific antibodies is based on the co-expression of two pairs of immunoglobulin heavy and light chains, where the two chains have different specificities. Due to the random combination of immunoglobulin heavy and light chains, these hybridomas (quadromas) may produce a mixture of 10 different antibody molecules, of which only one has the correct bispecific structure. Purification of the correct molecule is usually performed using an affinity chromatography step, which is quite cumbersome and results in low product yields.

[0311] According to a different approach, antibody variable domains with the desired binding specificities (antibody-antigen combining sites) are fused to immunoglobulin constant domain sequences. Fusions can be with immunoglobulin heavy chain constant domains, including at least part of the hinge, CH2, and CH3 regions. The first heavy chain constant region (CH1) can contain the site necessary for light chain binding, present in at least one of the fusions. Nucleic acids encoding the immunoglobulin heavy chain fusions and, if desired, the immunoglobulin light chain, are inserted into separate expression vectors and co-transfected into a suitable host organism. This allows for greater flexibility in adjusting the relative proportions of the three polypeptide fragments relative to one another, in embodiments where optimal yields are achieved by using unequal ratios of the three polypeptide chains in the construction. However, if expression of equal ratios of at least two polypeptide chains results in high yields, or if the ratio is not particularly important, it is possible to insert the coding sequences for two or all three polypeptide chains into a single expression vector.

[0312] Bispecific antibodies can have a hybrid immunoglobulin heavy chain with a first binding specificity in one arm and a hybrid immunoglobulin heavy chain-light chain pair (providing a second binding specificity) in the other arm. This asymmetric structure facilitates separation of the desired bispecific compound from undesired immunoglobulin chain combinations, as the presence of an immunoglobulin light chain in only one half of the bispecific molecule facilitates this separation mode. Such techniques can be used to prepare bispecific antibodies for binding to compounds that induce protein-protein interactions in the treatment or prevention of diseases as defined herein.

[0313] Hybrid or bifunctional antibodies can be derived biologically, i.e., by cell fusion techniques, or chemically, particularly using cross-linking or disulfide bridge-forming reagents, and can include whole antibodies or fragments thereof.

[0314] The antibody can be a functionally active fragment, derivative, or analog of an antibody that immunospecifically binds to a cancer cell antigen, a viral antigen, or a microbial antigen, or to another antibody bound to a tumor cell or matrix. In this context, "functionally active" means that the fragment, derivative, or analog is capable of eliciting anti-anti-idiotypic antibodies that recognize the same antigen as that recognized by the antibody from which the fragment, derivative, or analog is derived. Specifically, in exemplary embodiments, the idiotypic antigenicity of an immunoglobulin molecule can be enhanced by deletion of framework and CDR sequences C-terminal to the CDR sequences that specifically recognize the antigen. To determine which CDR sequences bind to an antigen, synthetic peptides containing the CDR sequences can be used in binding assays with the antigen using any binding assay method known in the art.

[0315] Other useful antibodies include, but are not limited to, antibody fragments such as F(ab')2 fragments containing the variable region, light chain constant region, and heavy chain CH1 domain, which can be produced by pepsin digestion of an antibody molecule, and Fab fragments, which can be produced by reducing the disulfide bridges of F(ab')2 fragments. Other useful antibodies are dimers of antibody heavy and light chains, or any minimal fragments thereof, such as Fv or single-chain antibodies (SCAs), or any other molecules having the same specificity as an antibody.

[0316] Additionally, recombinant antibodies containing both human and non-human portions, such as chimeric and humanized monoclonal antibodies, which can be produced using standard recombinant DNA techniques, are useful antibodies. Chimeric antibodies are molecules in which different portions are derived from different animal species, such as those having a variable region derived from a murine monoclonal and a human immunoglobulin constant region. Humanized antibodies are antibody molecules from non-human species that have one or more complementarity-determining regions (CDRs) from the non-human species and a framework region from a human immunoglobulin molecule. Such chimeric and humanized monoclonal antibodies can be produced by recombinant DNA techniques known in the art.

[0317] Fully human antibodies can be produced using transgenic mice that are incapable of expressing endogenous immunoglobulin heavy and light chain genes but that can express human heavy and light chain genes. The transgenic mice are immunized in the usual manner with a selected antigen, e.g., all or a portion of a polypeptide of the present disclosure. Monoclonal antibodies directed against the antigen can be obtained using conventional hybridoma technology. The human immunoglobulin transgenes harbored by the transgenic mice rearrange during B cell differentiation and subsequently undergo class switching and somatic mutation. Thus, such technology can be used to produce therapeutically useful IgG, IgA, IgM, and IgE antibodies. For an overview of this technology for producing human antibodies, see Lonberg and Huszar (1995, Int. Rev. Immunol. 13:65-93). Other human antibodies are commercially available, for example, from Abgenix, Inc. (Freemont, Calif.) and Genpharm (San Jose, Calif.).

[0318] Fully human antibodies that recognize a selected epitope can be generated using a technique called "guided selection." In this approach, a selected non-human monoclonal antibody, e.g., a murine antibody, is used to guide the selection of a fully human antibody that recognizes the same epitope. Human antibodies can also be generated using various techniques known in the art, such as phage display libraries.

[0319] The antibody may be a fusion protein of an antibody, or a functionally active fragment thereof, e.g., an antibody fused via a covalent bond (e.g., a peptide bond) at either the N- or C-terminus to the amino acid sequence (or a portion thereof, e.g., at least a 10, 20, or 50 amino acid portion of the protein) of another protein that is not an antibody. The antibody or fragment thereof may be covalently linked to the other protein at the N-terminus of the constant domain.

[0320] Antibodies include analogs and derivatives, all of which have been modified, i.e., by the covalent attachment of any type of molecule, provided that such covalent attachment allows the antibody to retain its antigen-binding immunospecificity. For example, without limitation, antibody derivatives and analogs include those that have been further modified, e.g., by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization with known protecting / blocking groups, proteolytic cleavage, conjugation to cellular antibody units or other proteins, etc. Any of a number of chemical modifications can be performed by known techniques, including, but not limited to, specific chemical cleavage, acetylation, formylation, metabolic synthesis in the presence of tunicamycin, etc. Additionally, analogs or derivatives can contain one or more unnatural amino acids.

[0321] The antibodies of the complexes can include antibodies with modifications (e.g., substitutions, deletions, or additions) in amino acid residues that interact with Fc receptors. In particular, the antibodies include antibodies with modifications in amino acid residues identified as being involved in the interaction between the anti-Fc domain and the FcRn receptor. Antibodies immunospecific for cancer cell antigens can be obtained commercially, for example, from Genentech (San Francisco, Calif.), or can be produced by any method known to those of skill in the art, for example, by chemical synthesis or recombinant expression techniques. Nucleotide sequences encoding antibodies immunospecific for cancer cell antigens can be obtained, for example, from the GenBank database or a database like it, literature publications, or by routine cloning and sequencing.

[0322] In certain embodiments, the antibody of the conjugate can be a monoclonal antibody, e.g., a murine monoclonal antibody, a chimeric antibody, or a humanized antibody. In some embodiments, the antibody can be an antibody fragment, e.g., a Fab fragment.

[0323] Antibodies for the treatment or prevention of cancer can be conjugated to the compound(s) described herein. Antibodies immunospecific for cancer cell antigens can be commercially obtained or produced by any method known to those skilled in the art, such as, for example, recombinant expression technology. Nucleotide sequences encoding antibodies immunospecific for cancer cell antigens can be obtained, for example, from the GenBank database or a database like it, literature publications, or by routine cloning and sequencing.Examples of antibodies available for the treatment of cancer include a humanized anti-HER2 monoclonal antibody for the treatment of patients with metastatic breast cancer; RITUXAN® (rituximab; Genentech), a chimeric anti-CD20 monoclonal antibody for the treatment of patients with non-Hodgkin's lymphoma; OVAREX® (oregovomab; AltaRex Corporation, MA), a murine antibody for the treatment of ovarian cancer; Panorex (edrecolomab; GlaxoWellcome, NC), a murine IgG2a antibody for the treatment of colorectal cancer; cetuximab-erbitux (cetuximab; Imclone Systems Inc., NY), an anti-EGFR IgG chimeric antibody for the treatment of epidermal growth factor-positive cancers such as head and neck cancer; Vitaxin (etaracizumab; MedImmune, Inc., MD), a humanized antibody for the treatment of sarcoma; and Campath, a humanized IgG1 antibody for the treatment of chronic lymphocytic leukemia (CLL). I / H (alemtuzumab, Leukosite, MA); Smart MI95 (Protein Design Labs, Inc., CA), a humanized anti-CD33 IgG antibody for the treatment of acute myeloid leukemia (AML); LymphoCide (epratuzumab, Immunomedics, Inc., NJ), a humanized anti-CD22 IgG antibody for the treatment of non-Hodgkin's lymphoma; Smart ID10 (Protein Design Labs, Inc., CA), a humanized anti-HLA-DR antibody for the treatment of non-Hodgkin's lymphoma; Oncolym (Techniclone, Inc., CA), a radiolabeled murine anti-HLA-Dr10 antibody for the treatment of non-Hodgkin's lymphoma; and humanized anti-CD2 for the treatment of Hodgkin's disease or non-Hodgkin's lymphoma. These include, but are not limited to, Allomune (BioTransplant, CA), a mAb; Avastin (bevacizumab, Genentech, Inc., CA), a humanized anti-VEGF antibody for the treatment of lung and colorectal cancer; epratuzumab (Immunomedics, Inc., NJ and Amgen, CA), an anti-CD22 antibody for the treatment of non-Hodgkin's lymphoma; and CEAcide (Immunomedics, NJ), a humanized anti-CEA antibody for the treatment of colorectal cancer.

[0324] Other antibodies useful in the conjugate include, but are not limited to, trastuzumab, gemtuzumab, pertuzumab, obinutuzumab, ofatumumab, daratumumab, STI-6129, lintuzumab, huMy9-6, belantamab, indatuximab, dinutuximab, anti-CD38 A2 antibody, buAT15 / 3 H3s antibody, ibritumomab, tositumomab, panitumumab, tremelimumab, ticilimumab, catumaxomab, and veltuzumab. In certain embodiments, the antibody is selected from the group consisting of rituximab, trastuzumab, pertuzumab, huMy9-6, lintuzumab, and gemtuzumab. Other antibodies useful in conjugates include, but are not limited to, polatuzumab, J591, lorvotuzumab, and sacituzumab.

[0325] Other antibodies useful in conjugates include, but are not limited to, antibodies against the following antigens: CA125 (ovarian), CA15-3 (carcinoma), CA19-9 (carcinoma), L6 (carcinoma), Lewis Y (carcinoma), Lewis X (carcinoma), alpha fetoprotein (carcinoma), CA242 (colon), placental alkaline phosphatase (carcinoma), prostate specific antigen (prostate), prostatic acid phosphatase (prostate), epidermal growth factor (carcinoma), MAGE-1 (carcinoma), MAGE-2 (carcinoma), MAGE-3 (carcinoma), MAGE-4 (carcinoma), anti-transferrin receptor (carcinoma), p97 (melanoma), MUC1-KLH (breast cancer), CEA (colon). , gp100 (melanoma), MART1 (melanoma), PSA (prostate), IL-2 receptor (T-cell leukemia and lymphoma), CD20 (non-Hodgkin's lymphoma), CD52 (leukemia), CD33 (leukemia), CD22 (lymphoma), human chorionic gonadotropin (carcinoma), CD38 (multiple myeloma), CD40 (lymphoma), mucin (carcinoma), P21 (carcinoma), MPG (melanoma), and Neu oncogene product (carcinoma). Some particularly useful antibodies include, but are not limited to, BR96 mAb (Trail, PA, et al Science (1993) 261, 212-215), BR64 (Trail, PA, et al Cancer Research (1997) 57, 100-105), mAbs against the CD40 antigen such as S2C6 mAb (Francisco, JA, et al Cancer Res. (2000) 60:3225-3231), mAbs against the CD70 antigen such as 1F6 mAb, and mAbs against the CD30 antigen such as AC10. Many other internalizing antibodies that bind to tumor-associated antigens can be used and are being considered.

[0326] Other antigens that the complex can bind include 5T4, ACE, ADRB3, AKAP-4, ALK, AOC3, APP, Axin1, AXL, B7H3, B7-H4, BCL2, BCMA, bcr-abl, BORIS, BST2, C242, C4.4a, CA 125, CA6, CA9, CAIX, CCL11, CCR5, CD123, CD133, CD138, CD142, CD15, CD15-3, CD171, CD17 9a, CD18, CD19, CD19-9, CD2, CD20, CD22, CD23, CD24, CD25, CD27L, CD28, CD3, CD30, CD31, C D300LF, CD33, CD352, CD37, CD38, CD4, CD40, CD41, CD44, CD44v6, CD5, CD51, CD52, CD54, CD 56, CD62E, CD62P, CD62L, CD70, CD71, CD72, CD74, CD79a, CD79b, CD80, CD90, CD97, CD125, C D138, CD141, CD147, CD152, CD154, CD326, CEA, CEACAM5, CFTR, clumping factor, cKit, claudin 3, claudin 18.2, CLDN6, CLEC12A, CLL-1, cll3, c-MET, Cripto protein, CS1, CTLA-4, CXCR2, CXORF61, cyclin B1, CYP1B1, cadherin-3, cadherin-6, DLL3, E7, EDNRB, EFNA4, EGFR, EGFRvIII, ELF2M, EMR2, ENPP3, EPCAM, EphA2, ephrin A4, ephrin B2, EPHB4, ERBB2 (Her2 / neu), ErbB3, ERG (TMPRSS2) ETS fusion gene), ETBR, ETV6-AML, FAP, FCAR, FCRL5, FGFR1, FGFR2, FGFR3, FGFR4, FLT3, folate receptor alpha, folate receptor beta, FOLR1, Fos-related antigen 1, fucosyl GM1, GCC, GD2, GD3, GloboH, GM3, GPC1, GPC2, GPC3, gplOO, GPNMB, GPR20, GPRC5D, GUCY2C, HAVCR1, HER2, HER3, HGF, HMI.24, HMWMAA, HPVE6, hTERT, human telomerase reverse transcriptase, ICAM, ICOS-L, IFN-α, IFN-γ, IGF-I receptor, IGLL1, IL-2 receptor, IL-4 receptor, IL-13Ra2, IL-l lRa, IL-1 receptor, IL-12 receptor, IL-23 receptor, IL-13 receptor, IL-22 receptor, IL-4 receptor, IL-5 receptor, IL-6, interferon receptor, integrin (including α4, αvβ3, αvβ5, αvβ6, α1β4, α4β1, α4β7, α5β1, α6β4, and αIIbβ3 integrins), integrin alpha V, intestinal carboxylesterase, KIT, LAGE-la, LAIR1, LAMP-1, LCK, legumain, LewisY, LFA-1 (CD11a), L-selectin (CD62L), LILRA2, LIV-1, LMP2, LRRC15, LY6E, LY6K, LY75, MAD-CT-1, MAD-CT-2, MAGE Al, MelanA / MARTl, mesothelin, ML-IAP, MSLN, mucin, MUC1, MUC16, mut hsp70-2, MYCN, myostatin, NA17, NaPi2b, NCA-90, NCAM, nectin-4, NGF, NOTCH1, NOTCH2, NOTCH3, NOTCH4, NY-BR-1, NY-ESO-1, o-acetyl-GD2, OR51E2, OY-TES1, p53, p53 mutant, PANX3, PAP, PAX3, PAX5, p-CAD, PCTA-1 / galectin-8, PD-L1, PD-L2, PDGFR, PDGFR-beta, phosphatidylserine, PIK3CA, PLAC1, polysialic acid, protease, prostate cancer cells, prostein, Pseudomonasaeruginosa, rabies, survivin and telomerase, PD-1, PRSS21, PSCA, PSMA, PTK7, RAGE-1, RANKL, Ras mutants, respiratory syncytial virus, rhesus factor, RhoC, RON, ROR1, ROR2, RU1, RU2, sarcoma translocation breakpoints, SART3, SLAMF7, SLC44A4, sLe, SLITRK6, sperm protein 17, sphingosine-1-phosphate, SSEA-4, SSX2, STEAP1, TAG72, TARP, TCRβ, TEM1 / CD248, TEM7R, tenascin-C, TF, TGF-1, TGF-β2, TNF-α, TGS5, Tie2, TIM-1, Tn These include, but are not limited to, Ag, TRAC, TRAIL-R1, TRAIL-R2, TROP-2, TRP-2, TRPV1, TSHR, tumor antigen CTAA16.88, tyrosinase, UPK2, VEGF, VEGFR1, VEGFR2, vimentin, WT1, and / or XAGE1.

[0327] Antibodies that bind to antigens associated with antigen-presenting cells, such as CD40, OX40L, endoglin, DEC-205, 4-1BBL, CD36, CD36, CD204, MARCO, DC-SIGN, CLEC9A, CLEC5A, Dectin-2, CLEC10A, CD206, CD64, CD32A, CD1A, HVEM, CD32B, PD-L1, BDCA-2, XCR-1, and CCR2, can also be conjugated to a compound(s) that induces a protein-protein interaction.

[0328] The antibodies of the conjugates described herein can bind to both receptors or receptor complexes expressed on activated lymphocytes. The receptor or receptor complex can include an immunoglobulin gene superfamily member, a TNF receptor superfamily member, an integrin, a cytokine receptor, a chemokine receptor, a major histocompatibility protein, a lectin, or a complement regulatory protein. Non-limiting examples of suitable immunoglobulin superfamily members are CD2, CD3, CD4, CD8, CD19, CD22, CD28, CD79, CD90, CD152 / CTLA-4, PD-1, and ICOS. Non-limiting examples of suitable TNF receptor superfamily members are CD27, CD40, CD95 / Fas, CD134 / OX40, CD137 / 4-1BB, TNF-R1, TNFR-2, RANK, TACI, BCMA, osteoprotegerin, Apo2 / TRAIL-R1, TRAIL-R2, TRAIL-R3, TRAIL-R4, and APO-3. Non-limiting examples of suitable integrins are CD11a, CD11b, CD11c, CD18, CD29, CD41, CD49a, CD49b, CD49c, CD49d, CD49e, CD49f, CD103, and CD104. Non-limiting examples of suitable lectins are C-type, S-type, and I-type lectins.

[0329] In some embodiments, antibodies useful in the present disclosure include 3F8, 8H9, abagovomab, abciximab (REOPRO®), abituzumab, abrezekimab, abrilumab, actoxumab, adalimumab (HUMIRA®), adecatumumab, aducanumab, afacevicumab, afelimomab, afutuzumab, alacizumab, ALD518, alemtuzumab (CAMPATH®), (trademark), alirocumab (PRALUENT®), altumomab, amatuximab, anatumomab, andecaliximab, anetumab, anifrolumab, anrukinzumab, apolizumab, aprutumab, arcitumomab (CEA-SCAN®), asclinbacumab, acelizumab, atidortoxumab, atlizumab (tocilizumab, ACTEMRA®, ROACTEMRA®), atezolizumab (TECENTRIQ®), (Registered Trademark), atinumab, atorolimumab, avelumab (BAVENCIO), azintuxizumab, belantamab, bapineuzumab, basiliximab (SIMULECT®), bavituximab, BCD-100, bectumomab (LYMPHOSCAN®), begelomab, belantamab, belimumab (BENLYSTA®), bemarituzumab, benralizumab FASENRA®, bermekimab, bersanlimab, bertilimumab, besilesomab (SCINITIMUN®), bevacizumab (AVASTIN®), bezlotoxumab (ZINPLAVA®), biciromab (FIBRISCINT®), bimagrumab, bimekizumab, viltamimab, bivatuzumab, bleselumab, blinatumomab, brontuvetmab, brosozumab, bococizumab, brazikumab,Brentuximab, briakinumab, brodalumab (SILIQ™), brolucizumab (BEOVU®), brontiximab, burosumab (CRYSVITA®), cabiralizumab, caplacizumab (CABLIVI®), camidanlumab, camrelizumab, canakinumab (ILARIS®), cantuzumab, capromab, carlumab, carotuximab, catumaxomab (REMOVAB®), cBR96, CC49, cedelizumab, cemiplimab (LIBTAY®), O (registered trademark), sergituzumab, certrelimab, certolizumab, cetuximab (ERBITUX (registered trademark), civisatamab, cirmtuzumab, sitatuzumab, cizutumumab, clazakizumab, clenoliximab, clivatuzumab, cotuzumab, cofetuzumab, coltuximab, conatumumab, concizumab, cosfrobiximab, CR6261, crenezumab, crizanlizumab (ADAKVEO (registered trademark), clotede tumab, cusatuzumab, dacetuzumab, daclizumab (ZINBRYTA®), dalotuzumab, dapirolizumab, daratumumab (DARZALEX®), dectrekumab, demcizumab, denituzumab, denosumab (PROLIA®), depatuximab, derlotuximab, detumomab, dezamizumab, dinutuximab (UNITUXIN®), diridavumab, domaglotuzumab, dostarlimab, dorlimomab, dorlixizumab, drozizumab, DS-8201, durigotuzumab, dupilumab (DUPIXENT®), durvalumab (IMFINZI®), dusigizumab, eclumeximab, eculizumab (SOLIRIS®), edovacomab, edrecolomab (PANOREX®), efalizumab (RAPTIVA®), efangumab (MYCOGRAB®), eldelumab,Elezanumab, elgemtumab, elotuzumab (EMPLICITI®), elsilimomab, emactuzumab, emapalumab (GAMIFANT®), emibetuzumab, emicizumab (HEMLIBRA®), enapotamab, enavatuzumab, enfortumab (Padcev®), enlimomab, enoblitzumab, enokizumab, enoticumab, ensituximab, epitumomab momab), eptinezumab (VYEPTI®), epratuzumab, erenumab (AIMOVIG®), erlizumab, ertumaxomab (REXOMUN®), etaracizumab (ABEGRIN®), etigilimab, etrolizumab, evinacumab, evolocumab (REPATHA®), exbivirumab, fanolesomab (NEUTROSPEC®), faralimomab, faricimab, farletuzumab , fasinumab, FBTA05, felvizumab, fezakinumab, fibatuzumab, ficlatuzumab, fizituzumab, firivumab, framvotumab, fretikumab, flotetuzumab, fontolizumab (HUZAF®), foralumab, foravirumab, fremanezumab (AJOVY®), frezolimumab, froboximab, flunevetomab, furanumab, futuximab ab), galcanezumab (EMGALITY®), galiximab, gancotamab, ganitumab, gantenerumab, gavilimomab, gezivumab, gemtuzumab, gevokizumab, gilvetmab, dimcirumab, girentuximab, glenbatumumab, golimumab (SIMPONI®), gomiliximab, guselkumab (TREMFYA®), huMy9-6, ianalumab, ibalizumab (TROGARZO®), IBI308, ibritumomab, icrucumab,Idarucizumab (PRAXBIND®), ifabotuzumab, igovomab (INDIMACIS-125), iladatuzumab, IMAB362, imalumab, imaprelimab, imciromab (MYOSCINT®), imgatuzumab, inlacumab, indatuximab, indusatumab, inebilizumab, infu Riximab (REMICADE®), intetumumab, inolimomab, inotuzumab, iomab-B, ipilimumab, iratumumab, isatuximab (SARCLISA®), iscalimab, istiratumab, itolizumab, ixekizumab (TALTZ®), keliximab, labetuzumab (CEA-CIDE™), lacunotuzumab, radilatuzumab, lampalizumab, lanadelumab (TAKHZYRO®), landgrossima, rapamycin, Rituximab (laprituximab), ralcabiximab, lebrikizumab, remaresomab (lemalesomab), lendalizumab, lenvervimab, lenzilumab, lerdelimumab, leronlimab, lesofabumab, letolizumab, lexatumumab, ribivirumab (libivirumab), rifastuzumab, ligelizumab, rilotomab, lintuzumab, lirilumab, rodelcizumab, lokivetmab, loncastuximab , lorvotuzumab, rosatuxizumab, lucatumumab, lulizumab, lumiliximab, lumletuzumab, rupartumab, lutikizumab, mapatumumab, marjetuximab, marstacimab, maslimomab, matuzumab, mavrilimumab, mepolizumab (NUCALA®), metelimumab, milatuzumab, minretumomab, mirikizumab, mirvetuximab, mitumomab,Modotuximab, molalizumab, mogamulizumab (POTELIGEO®), morolimumab, mosunetuzumab, motavizumab (NUMAX®), moxetumomab (LUMOXITI®), muromonab-CD3 (ORTHOCLONE®), OKT3®), nacolomab, namilumab, naptumomab, naratuximab, nalnatumab, natalizumab (TYSABRI®), navicixizumab, navivumab, naxitamab, nebacumab, necitumumab (PORTRAZZA®), nemolizumab, NEOD001, nerelimomab, nesbacumab, netakimab (netakimab), nimotuzumab (THERACIM®), nirsevimab, nivolumab, nofetumomab, obinutuzumab, ocaratuzumab, ocrelizumab (OCREVUS®), odulimomab, ofatumumab (ARZERRA®), olaratumumab (LARTRUVO®), olecurumab, orendali Ibuprofen, olokizumab, omalizumab (XOLAIR®), omburtamab, OMS721, onartuzumab, ontecizumab, ontuxizumab, onvatilimab, opicinumab, oportuzumab, oregovomab (OVAREX), orticumab, otelixizumab, otilimab, otlertuzumab, oxelumab, ozanezumab, ozogamicin ozoralizumab, pagibaximab, palivizumab (SYNAGIS®), pamrevlumab, panitumumab (VECTIBIX®), pankomab, panobacumab, palsatuzumab, pascolizumab, pasotuxizumab, pateclizumab, patritumab, PDR001, pembrolizumab, pemtumomab (THERAGYN®),Perakizumab, pertuzumab (OMNITARG®), pexelizumab, pidilizumab, pinatuzumab, pintumomab, placurumab, polatuzumab (Polivy), prezalumab (preza, lumab), prozalizumab, pogalizumab, ponezumab, polgabiximab, prasinezumab, prezalizumab, priliximab, pritoxaximab, pritumumab, PRO 140, kilimumab, racotumomab, radretumab, rafivirumab, ralpancizumab, ramucirumab, ranevetomab, ranibizumab (LUCENTIS®), ravagalimab, ravulizumab (ULTOMIRIS®), raxibacumab, rafanezumab, regavirumab, REGN-EB3, lenatolizumab (r enatlimab), remtolumab, reslizumab (CINQAIR®), rilotumumab, rinucumab, risankizumab (SKYRIZI®), rituximab (RITUXAN®), rivabazumab, rmab, lobatumumab, roledumab, romilkimab, romosozumab (EVENITY®), lontalizumab, rosmantuzumab , rovalpituzumab, rovelizumab (LEUKARREST®), rozanolixizumab, ruplizumab (ANTOVA), SA237, sacituzumab, samalizumab, samrotamab, sarilumab (KEVZARA®), satralizumab, satumomab pendetide, secukinumab (COSENTYX®), selicrelumab, seribantumab, setoxaximab, setrusumab ab), sevirumab, SGN-CD19A, SHP647, sibrotuzumab, sifalimumab, siltuximab, simtuzumab, siplizumab, sirtratumab, sirukumab, sofituzumab, solanezumab, solitomab, sonepcizumab, sontuzumab, spartalizumab, stamulumab, STI-6129,Sulesomab (LEUKOSCAN®), sputumab, stimulimab, suvizumab, subratoxumab, tabalumab, tacatuzumab (AFP-CIDE®), tadocizumab, talacotuzumab, talizumab, tamtuvetmab, tanezumab, taplitumomab paptox paptox, talexuzumab, tabolimab, tefibazumab (AUREXIS®), telimomab, telisotuzumab, tesidolumab, tetraxetan, tetulomab, tenatumomab, teneliximab, teprotumumab (TEPEZZA®), teplizumab, tezepelumab, TGN1412, tiburizumab, ticiririm Mab (TREMELIMUMAB®), tigatuzumab, timigutuzumab, timolumab, tiragolumab, tiragotumab, tislelizumab, tisotumab, tiuxetan, tildrakizumab (ILUMYA®), TNX-650, tocilizumab (atlizumab, ACTEMRA®), tomuzotuximab b), toralizumab, tosatoxumab, tositumomab (BEXXAR®), tobetumab, tralokinumab, trastuzumab (HERCEPTIN®), TRBS07, tregalizumab, tremelimumab, trevoglumab, tucotuzumab, tuvilumab, urtoxazumab, ustekinumab (STELERA®), ublituximab Mab, urocupulumab, urelumab, utomilumab, vadastuximab, banalimab, bundletuzumab, vanticutuzumab, vanucizumab, vapaliximab, varisacumab, valilumab, vatelizumab, vedolizumab, veltuzumab, bepalimomab, besencumab, visilizumab (NUVION®), bovalilizumab,Antibodies useful in the present disclosure include, but are not limited to, volociximab (HUMASPECT®), bonlerolizumab, vopratelimab, borsetuzumab, votumumab, bunakizumab, xentuzumab, XMAB-5574, zalutumumab (HuMEX-EGFr), zanolimumab (HuMAX-CD4), zatuximab, xenoctuzumab, ziralimumab, zolbetuximab, or zolimomab. In some embodiments, antibodies useful in the present disclosure include, but are not limited to, J591 and belantamab.

[0330] In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that comprises six CDRs of an antibody of Table A (i.e., three CDRs from the variable heavy chain or heavy chain and three CDRs from the variable light chain or light chain of the same antibody).

[0331] The term "Kabat numbering" and similar terms are recognized in the art and refer to a numbering system for amino acid residues in the variable regions of the heavy and light chains of an antibody or an antigen-binding fragment thereof. In some embodiments, CDRs can be determined according to the Kabat numbering system (see, e.g., Kabat EA & Wu TT (1971) Ann NY Acad Sci 190:382-391 and Kabat EA et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USDapartment of Health and Human Services, NIH Publication No. 91-3242). Using the Kabat numbering system, the CDRs within an antibody heavy chain molecule are typically located at amino acids 31-35 (optionally including one or two additional amino acids after 35, designated 35A and 35B in the Kabat numbering scheme) (CDR1), 50-65 (CDR2), and 95-102 (CDR3). Using the Kabat numbering system, the CDRs within an antibody light chain molecule are typically located at amino acids 24-34 (CDR1), 50-56 (CDR2), and 89-97 (CDR3). In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof comprising six Kabat-defined CDRs (i.e., three Kabat-defined CDRs for the variable heavy chain or heavy chain and three Kabat-defined CDRs for the variable light chain or light chain of the same antibody) of an antibody of Table A.

[0332] The CDRs of an antibody or antigen-binding fragment thereof can be determined according to the Chothia numbering scheme, which refers to the position of the loops in the immunoglobulin structure (see, e.g., Chothia C & Lesk AM, (1987), J Mol Biol 196:901-917; Al-Lazikani B et al., (1997) J Mol Biol 273:927-948; Chothia C et al., (1992) J Mol Biol 227:799-817; Tramontano A et al., (1990) J Mol Biol 215(1):175-82; and U.S. Patent No. 7,709,226). Typically, using the Kabat numbering system, the Chothia CDR-H1 loop is located at heavy chain amino acids 26 through 32, 33, or 34; the Chothia CDR-H2 loop is located at heavy chain amino acids 52-56; the Chothia CDR-H3 loop is located at heavy chain amino acids 95-102; the Chothia CDR-L1 loop is located at light chain amino acids 24-34; the Chothia CDR-L2 loop is located at light chain amino acids 50-56; and the Chothia CDR-L3 loop is located at light chain amino acids 89-97. When numbered using Kabat numbering, the ends of the Chothia CDR-H1 loop vary from H32 to H34 depending on the length of the loop (this is because the Kabat numbering scheme places insertions at H35A and H35B; if neither 35A nor 35B are present, the loop ends at 32; if only 35A is present, the loop ends at 33; and if both 35A and 35B are present, the loop ends at 34).

[0333] In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that comprises six CDRs according to the Chothia definition of an antibody of Table A (i.e., three CDRs according to the Chothia definition of the variable heavy or heavy chain and three CDRs according to the Chothia definition of the variable light or light chain of the same antibody). In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that comprises one or more CDRs where the Chothia and Kabat CDRs are identical in amino acid sequence. In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that comprises a combination of Kabat and Chothia CDRs of an antibody of Table A.

[0334] In some embodiments, the CDRs of an antibody or antigen-binding fragment thereof may be determined according to the IMGT numbering system described in Lefranc MP, (1999) The Immunologist 7:132-136 and Lefranc MP et al., (1999) Nucleic Acids Res 27:209-212. According to the IMGT numbering scheme, VH-CDR1 is located at positions 26-35, VH-CDR2 is located at positions 51-57, VH-CDR3 is located at positions 93-102, VL-CDR1 is located at positions 27-32, VL-CDR2 is located at positions 50-52, and VL-CDR3 is located at positions 89-97. In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof, comprising six CDRs according to the IMGT definition of an antibody of Table A (i.e., three CDRs according to the IMGT definition of the variable heavy chain or heavy chain and three CDRs according to the IMGT definition of the variable light chain or light chain of the same antibody), e.g., as described in Lefranc MP (1999) (supra) and Lefranc MP et al., (1999) (supra)).

[0335] In some embodiments, the CDRs of an antibody or antigen-binding fragment thereof can be determined according to MacCallum RM et al., (1996) J Mol Biol 262:732-745. See also, e.g., Martin A. "Protein Sequence and Structure Analysis of Antibody Variable Domains," in Antibody Engineering, Kontermann and Dubel, eds., Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001). In some embodiments, the binding portion is an antibody or antigen-binding portion thereof that comprises six MacCallum CDRs (i.e., three MacCallum CDRs of the variable heavy chain or heavy chain and three MacCallum CDRs of the variable light chain or light chain of the same antibody), as determined, e.g., by the method of MacCallum RM et al.

[0336] In some embodiments, the CDRs of an antibody or antigen-binding fragment thereof may be determined according to the AbM numbering scheme, which refers to AbM hypervariable regions that are a compromise between the Kabat CDRs and Chothia structural loops, and is used by Oxford Molecular's AbM antibody modeling software (Oxford Molecular Group, Inc.). In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that comprises six AbM-defined CDRs of an antibody of Table A (i.e., three AbM-defined CDRs of the variable heavy chain or heavy chain and three AbM-defined CDRs of the variable light chain or light chain of the same antibody) as determined by the AbM numbering scheme.

[0337] In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that binds to CD33. In some embodiments, the antibody or antigen-binding portion thereof binds to CD33 and comprises the six CDRs of the anti-CD33 antibody disclosed in U.S. Patent No. 10,711,062, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to CD33 and comprises the VH and VL of the anti-CD33 antibody disclosed in U.S. Patent No. 10,711,062. In some embodiments, the antibody or antigen-binding portion thereof binds to CD33 and comprises the six CDRs of the anti-CD33 antibody disclosed in U.S. Patent Application Publication No. 2021 / 0047404, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to CD33 and comprises the VH and VL of the anti-CD33 antibody disclosed in U.S. Patent Application Publication No. 2021 / 0047404. In some embodiments, the antibody or antigen-binding portion thereof binds CD33 and comprises the six CDRs of the anti-CD33 antibody disclosed in U.S. Patent Application Publication No. 2020 / 0297764, which is incorporated by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds CD33 and comprises the VH and VL of the anti-CD33 antibody disclosed in U.S. Patent Application Publication No. 2020 / 0297764. In some embodiments, the antibody or antigen-binding portion thereof binds CD33 and comprises the six CDRs of an anti-CD33 antibody (e.g., CD33-A, CD33-B, or CD33-C) listed in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds CD33 and comprises the six CDRs of the anti-CD33 antibody CD33-D listed in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds CD33 and comprises the six CDRs of the anti-CD33 antibody CD33 huMy9-6 listed in Table A. In some embodiments, the antibody, or antigen-binding portion thereof, binds to CD33 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 1 and a VL comprising the amino acid sequence of SEQ ID NO: 2. In some embodiments, the antibody, or antigen-binding portion thereof, binds to CD33 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 3 and a VL comprising the amino acid sequence of SEQ ID NO: 4.In some embodiments, the antibody, or antigen-binding portion thereof, binds to CD33 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 5 and a VL comprising the amino acid sequence of SEQ ID NO: 6. In some embodiments, the antibody, or antigen-binding portion thereof, binds to CD33 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 27 and a VL comprising the amino acid sequence of SEQ ID NO: 28. In some embodiments, the antibody, or antigen-binding portion thereof, binds to CD33 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 22 and a VL comprising the amino acid sequence of SEQ ID NO: 23. In some embodiments, the antibody, or antigen-binding portion thereof, binds to CD33 and comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 24 and a light chain comprising the amino acid sequence of SEQ ID NO: 25.

[0338] In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that binds to PSMA. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the six CDRs of the anti-PSMA antibody disclosed in U.S. Patent Application Publication No. 2019 / 0022205, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the VH and VL of the anti-PSMA antibody disclosed in U.S. Patent Application Publication No. 2019 / 0022205. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the six CDRs of the anti-PSMA antibody disclosed in U.S. Patent Application Publication No. 10,100,126, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the VH and VL of the anti-PSMA antibody disclosed in U.S. Patent Application Publication No. 10,100,126. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the six CDRs of the anti-PSMA antibody disclosed in U.S. Patent No. 8,470,330, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the VH and VL of the anti-PSMA antibody disclosed in U.S. Patent No. 8,470,330. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the six CDRs of an anti-PSMA antibody (i.e., PSMA-A, PSMA-B, or PSMA-C) listed in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises the six CDRs of the anti-PSMA antibody PSMA-D listed in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises a VH comprising the amino acid sequence of SEQ ID NO:7 and a VL comprising the amino acid sequence of SEQ ID NO:8. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises a VH comprising the amino acid sequence of SEQ ID NO:9 and a VL comprising the amino acid sequence of SEQ ID NO:10. In some embodiments, the antibody or antigen-binding portion thereof binds to PSMA and comprises a VH comprising the amino acid sequence of SEQ ID NO:11 and a VL comprising the amino acid sequence of SEQ ID NO:12.In some embodiments, the antibody, or antigen-binding portion thereof, binds to PSMA and comprises a VH comprising the amino acid sequence of SEQ ID NO:29 and a VL comprising the amino acid sequence of SEQ ID NO:30.

[0339] In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that binds to HER2. In some embodiments, the antibody or antigen-binding portion thereof binds to HER2 and comprises the six CDRs of the anti-HER2 antibody disclosed in U.S. Patent No. 7,862,817, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to HER2 and comprises the VH and VL of the anti-HER2 antibody disclosed in U.S. Patent No. 7,862,817. In some embodiments, the antibody or antigen-binding portion thereof binds to HER2 and comprises the six CDRs of the anti-HER2 antibody disclosed in U.S. Patent No. 7,850,966, which is incorporated herein by reference in its entirety. In some embodiments, the antibody or antigen-binding portion thereof binds to HER2 and comprises the VH and VL of the anti-HER2 antibody disclosed in U.S. Patent No. 7,850,966. In some embodiments, the antibody or antigen-binding portion thereof binds to HER2 and comprises the six CDRs of the anti-HER2 antibody disclosed in PCT International Publication No. WO 2016 / 201051, which is incorporated herein by reference in its entirety. In some embodiments, the antibody, or antigen-binding portion thereof, binds to HER2 and comprises the VH and VL of an anti-HER2 antibody disclosed in PCT International Publication No. WO 2016 / 201051. In some embodiments, the antibody, or antigen-binding portion thereof, binds to HER2 and comprises the six CDRs of an anti-HER2 antibody set forth in Table A (i.e., HER2-A, HER2-B, or HER2-C). In some embodiments, the antibody, or antigen-binding portion thereof, binds to HER2 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 13 and a VL comprising the amino acid sequence of SEQ ID NO: 14. In some embodiments, the antibody, or antigen-binding portion thereof, binds to HER2 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 15 and a VL comprising the amino acid sequence of SEQ ID NO: 16. In some embodiments, the antibody, or antigen-binding portion thereof, binds to HER2 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 17 and a VL comprising the amino acid sequence of SEQ ID NO: 18.

[0340] In some embodiments, the binding portion is an antibody or antigen-binding portion thereof that binds to CD20. In some embodiments, the antibody or antigen-binding portion thereof binds to CD20 and comprises the six CDRs of anti-CD20 antibody CD20-A, as set forth in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds to CD20 and comprises a VH comprising the amino acid sequence of SEQ ID NO: 31 and a VL comprising the amino acid sequence of SEQ ID NO: 32.

[0341] In some embodiments, the binding portion is an antibody or antigen-binding portion thereof that binds to CD79b. In some embodiments, the antibody or antigen-binding portion thereof binds to CD79b and comprises the six CDRs of anti-CD79b antibody CD79b-A, as set forth in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds to CD79b and comprises a VH comprising the amino acid sequence of SEQ ID NO: 33 and a VL comprising the amino acid sequence of SEQ ID NO: 34.

[0342] In some embodiments, the binding moiety is an antibody or antigen-binding portion thereof that binds to BCMA. In some embodiments, the antibody or antigen-binding portion thereof binds to BCMA and comprises the six CDRs of BCMA-A, an anti-BCMA antibody described in Table A. In some embodiments, the antibody or antigen-binding portion thereof binds to BCMA and comprises the amino acid sequences of SEQ ID NO:35 and SEQ ID NO:36. [Table 6-1] [Table 6-2] [Table 6-3] [Table 6-4] [Table 6-5]

[0343] In some embodiments, the antibody, or antigen-binding portion thereof, comprises a constant region. The linker may be attached to an amino acid within the constant region. In some embodiments, the antibody, or antigen-binding portion thereof, comprises a CH1 domain. The linker may be attached to an amino acid within the CH1 domain. In some embodiments, the antibody, or antigen-binding portion thereof, comprises a CH2 domain. The linker may be attached to an amino acid within the CH2 domain. In some embodiments, the antibody, or antigen-binding portion thereof, comprises a CH3 domain. The linker may be attached to an amino acid within the CH3 domain. In some embodiments, the antibody, or antigen-binding portion thereof, comprises a CL domain. The linker may be attached to an amino acid within the CL domain.

[0344] In some embodiments, the constant region, CH1 domain, CH2 domain, CH3 domain, or CL domain is an engineered constant region, CH1 domain, CH2 domain, CH3 domain, or CL domain.

[0345] In some embodiments, the antibody, or antigen-binding portion thereof, comprises a heavy chain constant region, e.g., a human heavy chain constant region. The linker may be attached to an amino acid within the heavy chain constant region, e.g., a human heavy chain constant region. In some embodiments, the antibody, or antigen-binding portion thereof, comprises an IgG heavy chain constant region, e.g., a human IgG heavy chain constant region. The linker may be attached to an amino acid within the IgG heavy chain constant region, e.g., a human IgG heavy chain constant region. In some embodiments, the antibody, or antigen-binding portion thereof, comprises an IgG1 heavy chain constant region, e.g., a human IgG1 heavy chain constant region. The linker may be attached to an amino acid within the IgG1 heavy chain constant region, e.g., a human IgG1 heavy chain constant region. In some embodiments, the antibody, or antigen-binding portion thereof, comprises an IgG4 heavy chain constant region. The linker may be attached to an amino acid within the IgG4 heavy chain constant region, e.g., a human IgG4 heavy chain constant region.

[0346] In some aspects, the antibody, or antigen-binding portion thereof, comprises a light chain constant region, e.g., a human light chain constant region. The linker can be attached to an amino acid within the light chain constant region, e.g., a human light chain constant region. In some aspects, the antibody, or antigen-binding portion thereof, comprises a kappa light chain constant region, e.g., a human kappa light chain constant region. The linker can be attached to an amino acid within the kappa light chain constant region, e.g., a human kappa light chain constant region. In some aspects, the antibody, or antigen-binding portion thereof, comprises a gamma light chain constant region, e.g., a human gamma light chain constant region. The linker can be attached to an amino acid within the gamma light chain constant region, e.g., a human gamma light chain constant region.

[0347] In some embodiments, the antibody, or antigen-binding portion thereof, comprises an engineered cysteine ​​at heavy chain position S239 according to EU numbering. A linker can be attached to S239C. In some embodiments, the antibody, or antigen-binding portion thereof, comprises an engineered cysteine ​​at heavy chain position K334 according to EU numbering. A linker can be attached to K334C.

[0348] Thus, the antibody or antigen-binding portion thereof can comprise the heavy chain constant region of SEQ ID NO:19, SEQ ID NO:20, or SEQ ID NO:21. IgG1 heavy chain constant region ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG (SEQ ID NO: 19) [ka] [ka]

[0349] The antibody or antigen-binding portion thereof can comprise the heavy chain constant region of SEQ ID NO:26. IgG4 heavy chain constant region S228P ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK (SEQ ID NO: 26)

[0350] The antibody or antigen-binding portion thereof can comprise the heavy chain constant region of SEQ ID NO:37. IgG1 N297A constant region (CH1-hinge-CH2-CH3) ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG (SEQ ID NO: 37)

[0351] In some embodiments, the linker can be attached to heavy chain Q295 of the antibody or antigen-binding portion thereof according to EU numbering.

[0352] An antibody that "binds" to a molecular target or antigen of interest is one that can bind to that antigen with sufficient affinity so that such an antibody is useful in targeting cells that express that antigen.

[0353] In the present disclosure, the "Bm" group can be bound to multiple compounds that induce protein-protein interactions. In some embodiments, "Bm" can be bound to 1 to 10 compounds. In some embodiments, "Bm" can be bound to 1 to 9 compounds. In some embodiments, "Bm" can be bound to 1 to 8 compounds. In some embodiments, "Bm" can be bound to 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 compounds. In some embodiments, "Bm" can be bound to 7 or 8 compounds. In some embodiments, "Bm" is bound to 5 compounds. In some embodiments, "Bm" is bound to 6 compounds. In some embodiments, "Bm" is bound to 7 compounds. In some embodiments, "Bm" is bound to 8 compounds. In some embodiments, "Bm" is bound to 9 compounds.

[0354] V. Composition and Methods of Use The conjugates and / or compounds described herein may be in the form of pharmaceutical or pharmaceutically acceptable salts. In some embodiments, such salts are derived from inorganic or organic acids or bases.

[0355] Examples of suitable acid addition salts include acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, lucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, oxalate, pamoate, pectinate, persulfate, 3-phenyl-propionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, tosylate, and undecanoate salts.

[0356] Examples of suitable base addition salts include ammonium salts; alkali metal salts, such as sodium salts and potassium salts; alkaline earth metal salts, such as calcium salts and magnesium salts; salts with organic bases, such as dicyclohexylamine salts, N-methyl-D-glucamine salts; and salts with amino acids such as arginine and lysine.

[0357] For example, Berge lists the following FDA-approved commercially available salts of the anions acetate, besylate (benzenesulfonate), benzoate, bicarbonate, bitartrate, bromide, calcium edetate (ethylenediaminetetraacetate), camsylate (camphorsulfonate), carbonate, chloride, citrate, dihydrochloride, edetate (ethylenediaminetetraacetate), edisylate (1,2-ethanedisulfonate), estolate (lauryl sulfate), esylate (ethanesulfonate), fumarate, gluceptate (glucoheptonate), gluconate, glutamate, glycollylarsanilate (glycolamidophenylarsonate), hexylresorcinate, hydrabamine (N,N'-di(dehydroabietyl)ethylenediamine), hydrobromide, hydrochloride, hydroxynaphthoate, and iodide. esters, isethionate (2-hydroxyethanesulfonate), lactate, lactobionate, malate, maleate, mandelate, mesylate|mesylate (methanesulfonate), methyl bromide, methyl nitrate, methyl sulfate, mucate, napsylate (2-naphthalenesulfonate), nitrate, pamoate (embonate), pantothenate, phosphate / diphosphate, polygalacturonate, salicylate, stearate, diacetate, succinate, sulfate, tannate, tartrate, teoclate (8-chlorotheophyllinate), and triethiodide; the organic cations benzathine (N,N'-dibenzylethylenediamine), chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N-methylglucamine), and procaine; and the metal cations aluminum, calcium, lithium, magnesium, potassium, sodium, and zinc.

[0358] Berge further lists the following commercially available (outside the United States) salts that are not FDA approved: anionic adipate, alginate, aminosalicylic acid, anhydromethylene citrate, arecoline, aspartic acid, bisulfate, butyl bromide, camphorate, digluconate, dibromide, disuccinate, glycerophosphate, hemisulfate, hydrofluoride, hydroiodide, methylenebis(salicylate), napadisilate (1,5-naphthalenedisulfonate), ), oxalate, pectinate, persulfate, phenylethylbarbiturate, picrate, propionate, thiocyanate, tosylate, and undecanoate; the organic cations benethamine (N-benzylphenethylamine), clemizole (1-p-chlorobenzyl-2-pyrrolizin-1'-ylmethylbenzimidazole), diethylamine, piperazine, and tromethamine (tris(hydroxymethyl)aminomethane); and the metal cations barium and bismuth.

[0359] Pharmaceutical compositions containing the conjugates described herein may also contain suitable carriers, excipients, and adjuvants, which may vary depending on the method of administration.

[0360] In some embodiments, pharmaceutical compositions can be formulated into suitable parenteral dosage forms. Such formulations can be prepared by various methods known in the art. Pharmaceutical compositions can be administered directly into the bloodstream, intramuscularly, or directly into an organ. Suitable means for parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, ​​intrasternal, intracranial, intramuscular, and subcutaneous. Suitable devices for parenteral administration include needle-type injectors, needle-free injectors, and infusion techniques.

[0361] Parenteral compositions are typically aqueous solutions which may contain excipients such as salts, carbohydrates, and buffers, however, the compositions may also be formulated in sterile non-aqueous solutions or in dry form in combination with a suitable vehicle such as sterile pyrogen-free water.

[0362] The preparation of parenteral compositions under sterile conditions, for example, by lyophilization, may be readily accomplished using standard techniques well known to those skilled in the art.

[0363] Compositions for parenteral administration can be formulated for immediate and / or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted-, and programmed-release. Thus, compositions can be formulated as solids, semi-solids, or thixotropic liquids for administration as implanted depots that provide modified release of the active agent.

[0364] The parenteral preparation may be mixed with other suitable pharmaceutically acceptable excipients used in parenteral dosage forms, including, but not limited to, preservatives.

[0365] In another aspect, the pharmaceutical composition may be formulated as a suitable oral dosage form such as a tablet, capsule, powder, pellet, suspension, solution, emulsion, etc. Other suitable carriers may be present such as disintegrants, diluents, chelating agents, binders, glidants, lubricants, extenders, fillers, anti-adherents, etc.

[0366] Oral administration formulations may also contain other suitable pharmaceutical excipients such as sweeteners, vehicles / humectants, colorants, flavoring agents, preservatives, thickening / thickening agents, and the like.

[0367] The conjugates described herein can be used to treat various cancers. Certain conjugates disclosed herein may be useful as pharmaceuticals because they may be superior in terms of efficacy, pharmacokinetics (e.g., absorption, distribution, metabolism, excretion), solubility (e.g., solubility in water), interactions with other pharmaceuticals (e.g., drug-metabolizing enzyme inhibitory activity), safety (e.g., acute toxicity, chronic toxicity, genotoxicity, reproductive toxicity, cardiotoxicity, carcinogenicity, central nervous system toxicity), and / or stability (e.g., chemical stability, enzymatic stability).

[0368] The conjugates of the present disclosure can be used to treat diseases, such as cancers, including colorectal cancer (e.g., colon cancer, rectal cancer, anal cancer, familial colorectal cancer, hereditary non-polyposis colorectal cancer, gastrointestinal stromal tumors), lung cancer (e.g., non-small cell lung cancer, small cell lung cancer, malignant mesothelioma), mesothelioma, pancreatic cancer (e.g., pancreatic ductal carcinoma, pancreatic endocrine tumor), pharyngeal cancer, laryngeal cancer, esophageal cancer, stomach cancer / gastric cancer, and the like. cancer) (e.g., papillary adenocarcinoma, mucinous adenocarcinoma, adenosquamous carcinoma), duodenal cancer, small intestine cancer, breast cancer (e.g., invasive ductal carcinoma, ductal carcinoma in situ, inflammatory breast cancer), ovarian cancer (e.g., epithelial ovarian cancer, extragonadal germ cell tumor, ovarian germ cell tumor, ovarian low malignant potential tumor), testicular tumor, prostate cancer (e.g., hormone-dependent prostate cancer, non-hormone-dependent prostate cancer, castration-resistant prostate cancer), liver cancer (e.g., hepatocellular carcinoma, primary liver cancer, extrahepatic bile duct carcinoma), thyroid cancer (e.g., medullary thyroid carcinoma), kidney cancer (e.g., renal cell carcinoma (e.g., clear cell renal cell carcinoma), transitional cell carcinoma of the renal pelvis and ureter), uterine cancer (e.g., cervical cancer, endometrial carcinoma, uterine sarcoma), gestational choriocarcinoma, brain tumor (e.g., medulloblastoma, glioma, pine and pituitary adenoma), retinoblastoma, skin cancer (e.g., basal cell carcinoma, malignant melanoma), sarcoma (e.g., rhabdomyosarcoma, leiomyosarcoma, soft tissue sarcoma, spindle cell sarcoma), malignant bone tumor, bladder cancer, hematological / blood cancer (e.g., multiple myeloma, leukemia (e.g., acute myeloid leukemia), malignant lymphoma, Hodgkin's disease, chronic myeloproliferative disorder), cancer of unknown primary; cancer growth inhibitors; cancer metastasis inhibitors; apoptosis promoters; drugs for treating precancerous lesions (e.g., myelodysplastic syndromes); and the like.

[0369] In certain embodiments, the conjugates of the present disclosure can be used as pharmaceuticals for breast cancer, gastric cancer, ovarian cancer, uterine cancer, lung cancer, pancreatic cancer, liver cancer, lymphoma, or blood cancer. In certain embodiments, the conjugates of the present disclosure can be used as pharmaceuticals for prostate cancer, breast cancer, gastric cancer, non-small cell lung cancer, bile duct cancer, colon cancer, ovarian cancer, or neuregulin-1 (NRG1)-positive cancer. In certain embodiments, the conjugates of the present disclosure can be used as pharmaceuticals for non-Hodgkin's lymphoma (NHL), such as B-cell non-Hodgkin's lymphoma or diffuse large B-cell lymphoma (DLBCL).

[0370] Furthermore, the conjugates of the present disclosure can be used simultaneously with non-drug therapies, specifically, the conjugates can be combined with non-drug therapies such as (1) surgery, (2) hypertensive chemotherapy using angiotensin II or the like, (3) gene therapy, (4) hyperthermia, (5) cryotherapy, (6) laser ablation, and (7) radiation therapy.

[0371] For example, by using the conjugate of the present disclosure before or after the above-mentioned surgery, etc., effects such as prevention of the emergence of resistance, extension of disease-free survival, suppression of cancer metastasis or recurrence, extension of survival time, etc. may be obtained.

[0372] Furthermore, treatment with the conjugate of the present disclosure can be combined with supportive care, namely, (i) administration of antibiotics (e.g., β-lactams such as pansporin, and macrolides such as clarithromycin) for complications due to various infections, (ii) administration of high-calorie infusions, amino acid preparations, or multivitamins for improving malnutrition, (iii) administration of morphine for pain relief, (iv) administration of pharmaceutical agents for improving side effects such as nausea, vomiting, anorexia, diarrhea, leukopenia, thrombocytopenia, decreased hemoglobin concentration, hair loss, liver damage, kidney damage, DIC, fever, etc., and (v) administration of pharmaceutical agents for suppressing multidrug resistance of cancer, etc.

[0373] In some embodiments, the conjugates of the present disclosure can be used in combination with standard therapy, e.g., one or more therapeutic agents (e.g., anti-cancer agents and / or immunomodulatory agents). Thus, in certain embodiments, the methods of treating tumors disclosed herein include administering a conjugate of the present disclosure in combination with one or more additional therapeutic agents. In some embodiments, the conjugates of the present disclosure can be used in combination with one or more anti-cancer agents to target multiple elements of the immune pathway. In some embodiments, the anti-cancer agent comprises an immune checkpoint inhibitor (i.e., which blocks signaling through a specific immune checkpoint pathway). Non-limiting examples of immune checkpoint inhibitors that can be used in the present methods include a CTLA-4 antagonist (e.g., an anti-CTLA-4 antibody), a PD-1 antagonist (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody), a TIM-3 antagonist (e.g., an anti-TIM-3 antibody), or a combination thereof. Additional examples of immune checkpoint inhibitors include T-cell immunoglobulin and ITIM domain (TIGIT) antagonists, V-domain Ig suppressor of T-cell activation (VISTA) antagonists, B-cell and T-cell lymphocyte attenuator (BTLA) antagonists, and lymphocyte-activation gene-3 (LAG-3) antagonists. A comprehensive, non-limiting list of combinations is disclosed in detail in the "Combination Therapies" section of this application.

[0374] In some aspects, the conjugate of the present disclosure is administered to a subject before or after administration of an additional therapeutic agent. In other aspects, the conjugate of the present disclosure is administered to a subject simultaneously with the additional therapeutic agent. In certain aspects, the conjugate of the present disclosure and the additional therapeutic agent may be administered simultaneously as a single composition dissolved in a pharmaceutically acceptable carrier. In other aspects, the conjugate of the present disclosure and the additional therapeutic agent are administered simultaneously as separate compositions.

[0375] In some aspects, subjects that can be treated with the conjugates of the present disclosure are non-human animals, such as rats or mice, hi some aspects, subjects that can be treated are humans.

[0376] VI. Methods of Preparing Compounds and Conjugates The present disclosure provides a method for preparing a conjugate, such method comprising: combining a binding moiety with a compound of formula (XXII): [ka] The compound or a pharmaceutically acceptable salt thereof, wherein

[0377] n is 0 or 1,

[0378] R 1 is a compound that induces protein-protein interactions,

[0379] R 2 is hydrogen, -(CH2CH2O) v -CH3, C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl(C1-C3 alkyl), and v is 1 to 24;

[0380] each Y is independently S or O;

[0381] L * is a cleavable linker precursor attached to the binding moiety.

[0382] The present disclosure also provides a compound of formula (XXXII): [ka] The compound or a pharmaceutically acceptable salt thereof,

[0383] a is 1 to 10;

[0384] A' is [ka] or [ka] wherein:

[0385] n is 0 or 1,

[0386] each Y is independently S or O;

[0387] [ka] is R 1 indicates the point of attachment to

[0388] [ka] indicates the point of attachment to the methylene group,

[0389] R 1 is a compound that induces protein-protein interaction together with A',

[0390] R 2 is hydrogen, R 1 -A', a group that provides stability to R 1 - a group that provides solubility to A, and R 1 -A' is selected from groups that provide stability and solubility to A';

[0391] L is a cleavable linker,

[0392] Bm is a binding moiety capable of specifically binding to a protein, the method comprising the steps of:

[0393] (XXXI) [ka] The compound or a pharmaceutically acceptable salt thereof [wherein:

[0394] A', R 1 , and R 2 is as defined above,

[0395] L * is a cleavable linker precursor], This involves reacting with a binding moiety that is capable of specifically binding to the protein.

[0396] As described herein, the linker precursor contains a heterobifunctional group that connects to a binding moiety.

[0397] In some aspects, the linker precursor is cleavable by a protease. [ka] is selected from the group consisting of During the ceremony,

[0398] q is 2 to 10;

[0399] Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are each independently absent or a naturally occurring amino acid residue in the L-configuration or the D-configuration, with the proviso that Z 1 , Z 2 , Z 3 , and Z 4 are amino acid residues,

[0400] [ka] is the point of attachment to the parent molecular moiety.

[0401] Z 1 , Z 2 , Z 3 , Z 4 , and Z 5is independently absent or selected from the group consisting of L-valine, D-valine, L-citrulline, D-citrulline, L-alanine, D-alanine, L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-asparagine, D-asparagine, L-phenylalanine, D-phenylalanine, L-lysine, D-lysine, and glycine, with the proviso that Z 1 , Z 2 , Z 3 , and Z 4 , and Z 5 At least two of the residues are amino acid residues.

[0402] In some embodiments, Z 1 is absent or is glycine, and Z 2 is absent or is selected from the group consisting of L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-alanine, D-alanine, and glycine; Z 3 is selected from the group consisting of L-valine, D-valine, L-alanine, D-alanine, L-phenylalanine, D-phenylalanine, and glycine; Z 4 is selected from the group consisting of L-citrulline, D-citrulline, L-asparagine, D-asparagine, L-lysine, D-lysine, L-phenylalanine, D-phenylalanine, and glycine; Z 5 is absent or is glycine.

[0403] In some embodiments, L * teeth [ka] and During the ceremony, [ka] is the point of attachment to the parent molecular moiety.

[0404] In some embodiments, q is 4.

[0405] In some embodiments, L * is a bioreducible linker precursor. In some embodiments, the bioreducible linker precursor is [ka] wherein:

[0406] q is 2 to 10;

[0407] R, R', R'', and R''' are each independently selected from hydrogen, C-C alkoxyC-C alkyl, (C-C) NC-C alkyl, and C-C alkyl, or two geminal R groups together with the carbon atoms to which they are attached can form a cyclobutyl or cyclopropyl ring;

[0408] [ka] is the point of attachment to the parent molecular moiety.

[0409] In some embodiments, L * teeth, [ka] is a bioreducible linker In certain embodiments, L * is a click-to-release linker precursor. In some embodiments, L * teeth, [ka] and During the ceremony, q is 2 to 10; [ka] is the point of attachment to the parent molecular moiety.

[0410] In certain embodiments, L * is a beta-glucuronidase cleavable linker precursor. In some embodiments, L * teeth [ka] and During the ceremony, q is 2 to 10; ---- is absent or a bond, [ka] is the point of attachment to the parent molecular moiety.

[0411] In certain embodiments, R 2 is a group that provides stability to the complex. In some embodiments, R 2 is selected from C2-C6 alkenyl, C1-C6 alkyl; C2-C6 alkynyl, benzyl, C3-C6 cycloalkyl, and C3-C6 cycloalkyl (C1-C3 alkyl). In some embodiments, R 2 is C1-C6 alkyl. In some embodiments, R 2 is methyl.

[0412] In certain embodiments, R 2 is a group that provides solubility to the complex. In some embodiments, R 2 teeth, [ka] [ka] is selected from During the ceremony,

[0413] each n is independently 1, 2, 3, 4, or 5;

[0414] each y is independently 1 or 2;

[0415] Each R is independently hydrogen, CH 11 O5, C 12 H 21 O 10 , C 18 H 31 O 15 , or C 24 H 41 O 20 In some embodiments, R 1 is the formula (XXX): [ka] is a compound of During the ceremony,

[0416] [ka] indicates the point of attachment to the parent molecular moiety,

[0417] A is phenyl or C4-C 10 is a cycloalkyl ring,

[0418] R 10 is independently selected from hydrogen and halo;

[0419] U is selected from NH and CF;

[0420] R 20 is -C(O)R 3 , -N(R 4 )2, -(CH2) n OH, -(CH2) n N(R 4 )2, -(CH2) n Q'(CH2) m OH, -(CH2) n Q'(CH2) m SH, and -(CH2) n Q'(CH2) m N(R 4 )2, wherein:

[0421] R3 is hydrogen or C1-C6 alkyl,

[0422] Each R 4 are independently hydrogen or C1-C6 alkyl;

[0423] Q' is O, S, or NR 4 and

[0424] n is 1 to 6;

[0425] m is 2 to 5.

[0426] In some embodiments,

[0427] A is phenyl;

[0428] U is NH,

[0429] R 10 is a halo,

[0430] R 20 is methyl.

[0431] In some embodiments, A is phenyl;

[0432] U is NH,

[0433] R 10 is a halo,

[0434] R 20 is -(CH2)2O(CH2)2NHCH3.

[0435] In some embodiments, the compound of formula (XXX) is [ka] The compound is selected from the group consisting of:

[0436] In some embodiments, R1 -A' is a proteolysis targeting chimera (PROTAC). In certain embodiments, R 1 is the expression: POI-L 100 -CBN wherein

[0437] POI is a compound that binds to a protein of interest,

[0438] L 100 is the PROTAC linker,

[0439] CBN is the binding moiety of cereblon.

[0440] In some embodiments, the protein of interest is a nuclear hormone receptor, a translation termination factor, a transcription factor, a cyclin-dependent kinase, a tyrosine kinase, a serine / threonine kinase, or an E3 ligase. In some embodiments, the protein of interest is selected from CD33, GSPT1, BRD4, AR, ER, IKZF1 / 3, CK1a, BCL-XL, IKZF2, IRAK4, BTK, STAT3, BTK and iMiD, BRD9, TRK, MDM2, CDK2 / CDK9, CD97b, and EGFR.

[0441] In certain embodiments, L 100 comprises one or more functional groups selected from glycol, alkyl, alkynyl, triazolyl, piperazinyl, piperidinyl, and combinations thereof.

[0442] In some embodiments, CBN is [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A', [ka] L 100 indicates the attachment point to

[0443] In certain embodiments, R 1 teeth, [ka] [ka] [ka] [ka] is selected from During the ceremony, [ka] indicates the point of attachment to A'.

[0444] In some embodiments, the binding moiety is pretreated prior to reacting with a compound of Formula (XXII) or (XXXI). In certain embodiments, a compound of Formula (XXII) or (XXXI) is reacted with a binding moiety comprising an antibody or an antigen-binding portion thereof. In embodiments in which the binding moiety is an antibody, the antibody may be pretreated to reduce interchain disulfides prior to reaction with a compound of Formula (XXII) or (XXXI).

[0445] A general method for attaching a compound of formula (XXII) or formula (XXXI) to a cysteine ​​in Bm via the maleimide moiety of the linker is shown in Scheme II. 1 , R 2 and Y are defined herein; L ** is part of a linker as defined herein. [ka]

[0446] A general method for attaching a compound of formula (XXII) or formula (XXXI) to a lysine in Bm via an N-hydroxysuccinimide moiety of the linker is shown in Scheme I-2. 1 , R 2 and Y are defined herein; L ** is part of a linker as defined herein. [ka]

[0447] A general method for preparing compounds of formula (XX) and formula (XXX) and their activation in cells to release compounds that induce protein-protein interactions is shown in Scheme I-3. 1 , R 2 and Y are defined herein; L ** is part of a linker as defined herein.

[0448] In step 1, the heterobifunctional group within the linker is activated. In step 2, a protected linker is attached to the nitrogen of ring A. Step 3 shows the deprotection of the amine, step 4 shows the attachment of the Bm group, and step 5 shows the attachment of Bm to the PPI-linker moiety (which is shown in Schemes I-1 and I-2). Step 6 shows the activation of the complex within the cancer cell via a retro-Mannich reaction, releasing the active compound that induces protein-protein interactions. [ka] [Example]

[0449] General Synthetic Methods and Intermediates The compounds of the present disclosure can be prepared by one of ordinary skill in the art in light of this disclosure and knowledge in the art, and / or by reference to the schemes and synthetic examples set forth below. Some reagents and intermediates are known in the art. Other reagents and intermediates can be made by methods known in the art using readily available materials. Exemplary synthetic routes are described in the schemes and examples below. It should be understood that variables (e.g., "R" groups) appearing in the schemes and examples below are to be interpreted independently of their appearance elsewhere in this specification. One of ordinary skill in the art will readily understand how the schemes and examples set forth below describe the preparation of the compounds described herein.

[0450] Abbreviations used in the schemes generally follow conventions used in the art. Chemical abbreviations used in the specification and examples are defined as follows:

[0451] "Et3N" and "TEA" trimethylamine; "DMF" N,N-dimethylformamide; "rt" or "rt" or "RT" room temperature or retention time (depending on the context); "h" hour; "min" minute; "CDI" 1,1'-carbonyldiimidazole; "DMAP" N,N-dimethylaminopyridine; "TBAI" tetrabutylammonium bromide; "HATU" 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate or N-[(dimethylamino)-1H-1,2,3-triazolo-[4,5-b]pyridin-1-ylmethylene]-N-methylmethanaminium hexafluorophosphate N-oxide; "DIEA" and "iPrNEt2" diisopropylethylamine; "ACN" acetonitrile; "DCM" Dichloromethane; "MeOH" methanol; "Me" methyl; "PE" petroleum ether; "TFA" trifluoroacetic acid; "BOC" or "Boc", "DMSO" dimethyl sulfoxide; "Cbz" carbobenzyloxy; "EtOH" ethanol; "HOBt" or "HOBT" 1-hydroxybenzotriazole hydrate; "NBS" N-bromosuccinimide; "TMS" trimethylsilyl; and "THF" tetrahydrofuran.

[0452] [Table 7-1] [Table 7-2] [Table 7-3] [Table 7-4] [Table 7-5] [Table 7-6] [ka] Step 1. Synthesis of Compounds 1-3 To a stirred mixture of Gly-Gly-Gly (compound 1-1, 5.00 g, 25.1 mmol, 1.00 equiv.) in HO (25 mL), TEA (7.6 g, 75.1 mmol, 2.99 equiv.) was added dropwise at 0° C. To the above mixture, a DMF solution (25 mL) of 2,5-dioxopyrrolidin-1-yl 6-(2,5-dioxopyrrol-1-yl)hexanoate (compound 1-2, 9.29 g, 30.1 mmol, 1.20 equiv.) was added dropwise at 0° C. The resulting mixture was stirred at room temperature for an additional 3 hours. LCMS indicated the reaction was complete. The mixture was acidified to pH 4 with HCl (2N, aq.). The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel (330 g, 20-40 μm); mobile phase, water (containing 0.05% TFA), ACN (0% to 20% ACN gradient over 30 min); detector, UV 220 nm. This afforded (2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido]-acetamido)acetic acid (compound 1-3, 1 g, 8%) as a white solid. LCMS (ES, m / s): 383 [M+H] +

[0453] Step 1A: Synthesis of Compounds 1-4 Step a: 1: 3-(5-bromo-1-oxoisoindolin-2-yl)piperidine-2,6-dione To a stirred DMF solution (120 mL) of a mixture of methyl 4-bromo-2-(bromomethyl)benzoate (60.0 g, 194.8 mmol, 1.00 equiv.) and 3-aminopiperidine-2,6-dione hydrochloride (38.48 g, 233.8 mmol, 1.20 equiv.) was added TEA (67.70 mL, 669.0 mmol, 2.50 equiv.) dropwise at 25°C under a nitrogen atmosphere. The mixture was stirred at 25°C for 16 hours. After this time, HO (120 mL), AcOH (46 mL), and EtO (120 mL) were added sequentially at 25°C. The mixture was stirred at 25°C for 2 hours. LCMS indicated the reaction was complete. The precipitated solid was collected by filtration and washed with EtO (60 mL). This gave 3-(5-bromo-1-oxo-3H-isoindol-2-yl)piperidine-2,6-dione (40.0 g, 63%) as a white solid. LCMS (ESI, ms): 323,325 (M+H) + . 1 H NMR (300 MHz, DMSO-d6) δ 11.00(s, 1H), 7.90 (d, J = 1.5 Hz, 1H), 7.74-7.66 (m, 2H), 5.15-5.10(m, 1H), 4.51-4.32(m, 2H), 2.93-2.85(m, 1H), 2.74-2.56(m, 1H), 2.43-2.32(m, 1H), 2.06-1.99(m, 1H).

[0454] Step b: 2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-carbonitrile To a stirred solution of 3-(5-bromo-1-oxo-3H-isoindol-2-yl)piperidine-2,6-dione (2.00 g, 6.19 mmol, 1.00 equiv.) in DMF (40.00 mL) was added Zn(CN) (872 mg, 7.42 mmol, 1.20 equiv.) and Pd(PPh) (715 mg, 0.62 mmol, 0.10 equiv.) in a nitrogen atmosphere at room temperature in portions. The resulting mixture was stirred overnight at 80° C. under a nitrogen atmosphere. LCMS indicated the reaction was complete. The mixture was cooled to room temperature. The mixture was added to water (120.00 mL) and stirred for 30 minutes. The precipitated solid was collected by filtration and washed with water (3×20 mL) and EtOAc (3×20 mL). The resulting solid was dried under a sun lamp. This gave 2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindole-5-carbonitrile (1.2 g, 72%) as a white solid. LCMS (ESI, ms): 270 (M+H) + ; 1 H NMR (400 MHz, DMSO-d6) δ 11.03(s, 1H), 8.17 (d, J = 1.6 Hz, 1H), 8.00-7.91 (m, 2H), 5.18-5.13(m, 1H), 4.57-4.40(m, 2H), 2.92-2.88(m, 1H), 2.74-2.56(m, 1H), 2.48-2.37(m, 1H), 2.07-1.99(m, 1H).

[0455] Step c: 3-(5-(aminomethyl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione hydrochloride To a slurry of 2-(2,6-dioxopiperidin-3-yl)1-oxo-3H-isoindole-5-carbonitrile (8.00 g, 29.7 mmol, 1.00 equiv) in MeOH (67.00 mL) was added HCl (12 M, 9.60 mL) and PtO2 (3.30 g, 14.5 mmol, 0.49 equiv) at 25 °C. The mixture was stirred at room temperature under a hydrogen atmosphere using a hydrogen balloon for 16 h. LCMS showed the reaction was complete. The reaction was filtered and washed with MeOH (2 × 30 mL). The filtrate was evaporated to dryness under reduced pressure. The resulting solid was washed with DCM:MeOH (3:1) (3 × 30 mL). This gave 3[5-(aminomethyl)-1-oxo-3H-isoindol-2-yl]piperidine-2,6-dione hydrochloride (5.08 g, 57%) as a white solid. LCMS (ESI, ms): 274 (M+H-HCl). + ; 1 H NMR (400 MHz, CD3OD) δ 7.88(d, J = 8.0Hz, 1H), 7.68(s, 1H), 7.62(d, J = 8.0Hz, 1H), 5.19-5.15(m, 1H), 4.55-4.53(m, 2H), 4.26(s, 2H), 2.95-2.88(m, 1H), 2.81-2.80(m, 1H), 2.55-2.45(m, 1H), 2.22-2.16(m, 1H).

[0456] Step 2. Synthesis of Compounds 1-5 To a stirred mixture of 3-[5-(aminomethyl)-1-oxo-3H-isoindol-2-yl]piperidine-2,6-dione (compound 1-4, 1.00 g, 3.66 mmol, 1.00 equiv.) in DMF (10.00 mL), CDI (0.59 g, 3.66 mmol, 1.00 equiv.) and TEA (0.37 g, 3.66 mmol, 1.00 equiv.) were added portionwise at 0° C. under a nitrogen atmosphere. The resulting mixture was stirred at 0° C. for 2 hours under a nitrogen atmosphere. To the above mixture, DMAP (1.34 g, 10.98 mmol, 3.00 equiv.) and 3-chloro-p-toluidine (0.52 g, 3.66 mmol, 1.00 equiv.) were added portionwise at room temperature. The resulting mixture was stirred at 60° C. overnight under a nitrogen atmosphere. LCMS showed the reaction was complete. The reaction was quenched with water / ice at room temperature. The precipitated solid was collected by filtration and washed with DCM and water. This gave 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 1-5, 1.0 g, 51%) as a light brown solid. LCMS (ES, m / s): 441,443 [M+1] + .

[0457] Step 3. Synthesis of Compounds 1-6 To a stirred mixture of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound I-5, 500.00 mg, 1.13 mmol, 1.00 equiv) in DMF (5.00 mL) was added KCO (500.0 mg, 3.62 mmol, 3.19 equiv) in portions under a nitrogen atmosphere at 0° C. The resulting mixture was stirred at 0° C. for 30 minutes under a nitrogen atmosphere. To the above mixture, TBAI (100.0 mg, 0.27 mmol, 0.24 equiv), NaI (200.0 mg, 1.34 mmol, 1.18 equiv), and chloromethyl 4-nitrophenyl carbonate (801.0 mg, 3.46 mmol, 3.05 equiv) were added portionwise at 0° C. The resulting mixture was stirred in the dark at 0° C. for an additional 1 h. LCMS showed the reaction was complete. The reaction mixture was used in the next step without treatment. LCMS (ES, m / s): 636,638 [M+H] +

[0458] Step 4. Synthesis of compounds 1-7 To a stirred DMF solution (5.00 mL) of a mixture of [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 4-nitrophenyl carbonate (compound 1-6, 500 mg, 0.78 mmol, 1.00 equiv.) and K2CO3 (500 mg, 3.62 mmol, 4.60 equiv.) was added tert-butyl N-(2-aminoethyl)carbamate (400 mg, 2.50 mmol, 3.18 equiv.) in a nitrogen atmosphere at 0 °C in portions. The resulting mixture was stirred at room temperature for 1 hour. LCMS showed the reaction was complete. The reaction was quenched with water. The resulting mixture was extracted with diethyl ether (3 × 20 mL). The combined organic layer was washed with brine (20 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (EA) to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-[2-[(tert-butoxycarbonyl)amino]ethyl]-carbamate (compound 1-7, 270 mg, 44%) as a white solid. LCMS (ES, m / s): 657,659 [M+H] + .

[0459] Step 5. Synthesis of Compounds 1-8 A mixture of [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-[2-[(tert-butoxycarbonyl)amino]ethyl]carbamate (compound 1-7, 100 mg, 0.13 mmol, 1.00 equiv.) in HCl (gas)·1,4-dioxane (4 M) solution (1.5 mL) was stirred at 0° C. for 30 minutes. LCMS showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. This gave [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-(2-aminoethyl)carbamate hydrochloride (compound 1-8, 100 mg, 61%) as a white solid. LCMS (ES, m / s): 557,559 [M+H] + .

[0460] Step 6. Synthesis of Compound (I) To a stirred mixture of (2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido]-acetamido)acetic acid (compound 1-3, 64 mg, 0.17 mmol, 1.10 equiv) in DMF (3.5 mL) was added HATU (69 mg, 0.18 mmol, 1.20 equiv) in portions at 0° C. The resulting mixture was stirred at 25° C. for 20 min. To the above mixture, [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-(2-aminoethyl)carbamate hydrochloride (compound 1-8, 100 mg, 0.15 mmol, 1.00 equiv.) and DIEA (49 mg, 0.37 mmol, 2.50 equiv.) were added at 0 °C. The resulting mixture was stirred at 25 °C for an additional 16 hours. LCMS indicated the reaction was complete. The crude product was purified by preparative HPLC using the following conditions: column, XSelect CSH preparative C18 OBD column, 19 × 250 mm, 5 μm; mobile phase, water (containing 0.05% TFA) and ACN (24% to 43% in 7 min); detector, UV 254 nm. The collected fractions were lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-[2-[2-(2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]-acetamido]acetamido)acetamido]ethyl]carbamate (Compound (I), 18.2 mg, 12%). LCMS (ES, m / z): 921,923 [M+H] + . 1H-NMR (CD3OD, 400 MHz) δ (ppm): 7.76 (d, J = 7.6 Hz, 1H), 7.56-7.48 (m, 3H), 7.13-7.12 (m, 2H), 6.76 (s, 2H), 5.75-5.71 (m, 2H), 5.25-5.20 (m, 1H), 4.51-4.46 (m, 4H), 3.85-3.80 (m, 6H), 3.46-3.42 (m, 2H), 3.28-3.26 (m, 2H), 3.23-3.21 (m, 2H), 3.08-2.86 (m, 2H), 2.66-2.39 (m, 1H), 2.27-2.21 (m, 6H), 1.61-1.51 (m, 4H), 1.28-1.24 (m, 2H).

[0461] [ka] Step 1. Synthesis of compound 2-2 To a stirred mixture of 2-methyl-2-sulfanylpropan-1-ol (compound 2-1, 1.00 g, 9.41 mmol, 1.00 equiv) in DCM (3.00 mL) and MeOH (3.00 mL), 5-nitro-2-[(5-nitropyridin-2-yl)disulfanyl]pyridine (1.46 g, 4.70 mmol, 0.50 equiv) was added at room temperature. The resulting mixture was stirred at room temperature overnight. To the above mixture, MnO (1.50 g, 17.25 mmol, 1.83 equiv) was added portionwise at room temperature. The resulting mixture was stirred at room temperature for an additional 2 hours. LCMS showed the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with DCM (20 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (8:1) to give 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propan-1-ol (compound 2-2, 1.4 g, 57%) as an orange solid. LCMS (ESI, ms): 261 [M+H]

[0462] Step 2. Synthesis of Compound 2-3 To a stirred DCM solution (20.00 mL) of a mixture of 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propan-1-ol (compound 2-2, 1.20 g, 4.61 mmol, 1.00 equiv.) and pyridine (0.90 g, 11.38 mmol, 2.47 equiv.) was added a DCM solution (2.00 mL) of chloromethyl chloroformate (0.60 g, 4.65 mmol, 1.01 equiv.) dropwise at 0 °C. The resulting mixture was stirred at room temperature overnight. 30% of the desired product was detected by LCMS. The reaction was quenched with water / ice. The resulting mixture was extracted with DCM (3 × 20 mL). The combined organic layers were washed with brine (20 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (10:1) to give chloromethyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (compound 2-3, 330 mg, 20%) as a pale yellow oil. LCMS (ESI, ms): 353,355 [M+H]

[0463] Step 3. Synthesis of Compounds 2-4 To a stirred mixture of 3-chloro-p-toluidine (102 mg, 0.72 mmol, 0.99 equiv.) in THF (10.00 mL) was added diphosgene (145.00 mg, 0.73 mmol, 1.00 equiv.) dropwise at 0° C. under a nitrogen atmosphere. The resulting mixture was stirred at room temperature under a nitrogen atmosphere for 1 hour. The resulting mixture was concentrated under reduced pressure. To a stirred DMF solution (10.00 mL) of a mixture of 3-[5-(aminomethyl)-1-oxo-3H-isoindol-2-yl]piperidine-2,6-dione (INT, prepared according to the procedure described for Compound 1-4, 200 mg, 0.73 mmol, 1.00 equiv.) and TEA (61.00 mg, 0.60 mmol, 0.82 equiv.) was added dropwise to a DMF solution (15.00 mL) of the above mixture under a nitrogen atmosphere at 0° C. The resulting mixture was stirred under a nitrogen atmosphere at room temperature for 3 hours. LCMS indicated the reaction was complete. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), gradient from 0% to 80% in 40 minutes; detector, UV 254 nm. This afforded 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (8 compound 2-45 mg, 26.34%) as a white solid. The product was purified by preparative HPLC using the following conditions: Column: XBridge Shield RP18 OBD column, 19 x 250 mm, 10 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 25 B to 38 B in 18 min; 220 nm; RT 1:14.25; Collected fractions were lyophilized to give 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 2-4, 21.4 mg, 7%) as a white solid. LCMS (ESI, ms): 441,443 [M+H] +

[0464] Step 4. Synthesis of Compounds 2-5 To a stirred mixture of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 2-5, 300.00 mg, 0.68 mmol, 1.00 equiv.) and K2CO3 (180 mg, 1.30 mmol, 1.91 equiv.) in DMF (0.50 mL) was added chloromethyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (600 mg, 1.70 mmol, 2.50 equiv.) and TBAI (119.00 mg, 0.46 mmol, 0.67 equiv.) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature overnight. LCMS showed the reaction was complete. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), gradient from 0% to 85% over 40 min; detector, UV 254 nm. This afforded [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (85 mg, 14.85%) as a brown solid. The crude product was further purified using the following conditions: Column: XBridge Preparative OBD C18 column, 19 x 250 mm, 5 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 53 B to 68 B in 10 min; 220 nm; RT: 9.80; Collected fractions were lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (compound 2-5, 8.5 mg) as a white solid. LCMS (ESI): 757,757 [M+H] + ; 1H NMR (300 MHz, DMSO-d6) 9.23 (d, J = 2.4Hz, 1H), 8.75 (s, 1H), 8.58 (d, J = 2.7Hz, 1H), 8.05 (d, J = 8.7Hz, 1H), 7.72-7.6 (m, 2H), 7.53 (s, 1H), 7.46(d, J = 6.3Hz, 1H), 7.25-7.11 (m, 2H), 6.82-6.78 (m, 1H), 5.68-5.63 (m, 2H), 5.35-5.28 (m, 1H), 4.52-4.30 (m, 4H), 4.08 (s, 2H), 3.12-3.06 (m, 1H), 2.87-2.73 (m, 1H), 2.49-2.44 (m, 1H), 2.28 (s, 3H), 2.10-2.00 (m, 1H), 1.33 (s, 6H).

[0465] ステップ5. Synthesis of Compound (II) To a stirred solution of [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (Compound 2-5, 70.00 mg, 0.092 mmol, 1.00 equiv.) and sodium methylsulfinate (28.00 mg, 0.27 mmol, 2.97 equiv.) in DCM (14.00 mL) was added Br (14 mg, 0.087 mmol, 0.95 equiv.) dropwise at 0 °C. The resulting mixture was stirred at room temperature for 3 hours. To the above stirred mixture, sodium methylsulfinate (28.00 mg, 0.27 mmol, 2.97 equiv.) and Br2 (14 mg, 0.087 mmol, 0.95 equiv.) were added at room temperature. The resulting mixture was stirred at room temperature overnight. LCMS showed the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with DCM (20 mL). The filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water, gradient from 5% to 85% in 40 min; detector, UV 254 nm. The crude product was purified under the following conditions: Column: YMC-Actus Triart C18, 30 x 250, 5 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 55 B to 75 B in 7 min; 220 nm; RT1: 6.35. The collected fractions were lyophilized to give 1-(3-chloro-4-methylphenyl)-3-[(2-[1-[([[2-(methanesulfonylsulfanyl)-2-methylpropoxy]carbonyl]oxy)-methyl]-2,6-dioxopiperidin-3-yl]-1-oxo-3H-isoindol-5-yl)methyl]urea (Compound (II), 3.3 mg, 5.05%) as a white solid. LCMS (ESI): 681,683 [M+H] + 1H NMR (300 MHz, DMSO-d6) 8.75(s, 1H), 7.73-7.66(m, 2H), 7.53(s, 1H), 7.47(d, J = 7.8Hz, 1H), 7.20-7.12 (m, 2H), 6.82-6.79(m, 1H), 5.75-5.66(m, 2H), 5.33-5.27(m, 1H), 4.51-4.29(m, 6H), 3.35(s, 3H), 3.11-3.06(m, 1H), 2.87-2.73(m, 1H), 2.49-2.44 (m, 1H), 2.28(s, 3H), 2.10-2.00(m, 1H), 1.48(s, 6H)

[0466] [ka] Step 1. Synthesis of compound 3-2 To a stirred mixture of 2-methyl-2-sulfanylpropan-1-ol (1.00 g, 9.41 mmol, 1.00 equiv) in DCM (3.00 mL) and MeOH (3.00 mL), 5-nitro-2-[(5-nitropyridin-2-yl)disulfanyl]pyridine (1.46 g, 4.70 mmol, 0.50 equiv) was added at room temperature. The resulting mixture was stirred at room temperature overnight. To the above mixture, MnO2 (1.50 g, 17.25 mmol, 1.83 equiv) was added portionwise at room temperature. The resulting mixture was stirred at room temperature for an additional 2 hours. LCMS showed the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with DCM (20 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (8:1) to give 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propan-1-ol (compound 3-2, 1.4 g, 57%) as an orange solid. LCMS (ESI, ms): 261 [M+H].

[0467] Step 2. Synthesis of compound 3-3 To a stirred mixture of 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propan-1-ol (compound 3-2, 1.20 g, 4.61 mmol, 1.00 equiv.) and pyridine (0.90 g, 11.38 mmol, 2.47 equiv.) in DCM (20.00 mL) was added dropwise a solution of chloromethyl chloroformate (0.60 g, 4.65 mmol, 1.01 equiv.) in DCM (2.00 mL) at 0 °C. The resulting mixture was stirred at room temperature overnight. LCMS showed 30% of the desired product. The reaction was quenched with water / ice. The resulting mixture was extracted with DCM (3 × 20 mL). The combined organic layers were washed with brine (20 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (10:1) to give chloromethyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (compound 3-3, 330 mg, 20%) as a pale yellow oil. LCMS (ESI, ms): 353,355 [M+H] + .

[0468] Step 3. Synthesis of Compounds 3-4 To a stirred mixture of 3-chloro-p-toluidine (102 mg, 0.72 mmol, 0.99 equiv.) in THF (10.00 mL) was added diphosgene (145.00 mg, 0.73 mmol, 1.00 equiv.) dropwise at 0° C. under a nitrogen atmosphere. The resulting mixture was stirred at room temperature under a nitrogen atmosphere for 1 hour. The resulting mixture was concentrated under reduced pressure. To a stirred DMF solution (10.00 mL) of a mixture of 3-[5-(aminomethyl)-1-oxo-3H-isoindol-2-yl]piperidine-2,6-dione (INT, prepared according to the procedure described for Compound 1-4, 200 mg, 0.73 mmol, 1.00 equiv.) and TEA (61.00 mg, 0.60 mmol, 0.82 equiv.) was added dropwise to a DMF solution (15.00 mL) of the above mixture under a nitrogen atmosphere at 0° C. The resulting mixture was stirred under a nitrogen atmosphere at room temperature for 3 hours. LCMS indicated the reaction was complete. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), gradient from 0% to 80% in 40 minutes; detector, UV 254 nm. This afforded 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 3-4, 85 mg, 26.34%) as a white solid. The product was purified by preparative HPLC using the following conditions: Column: XBridge Shield RP18 OBD column, 19 x 250 mm, 10 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 25 B to 38 B in 18 min; 220 nm; RT 1:14.25; Collected fractions were lyophilized to give 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 3-4, 21.4 mg, 7%) as a white solid. LCMS (ESI, ms): 441,443 [M+H] +

[0469] Step 4. Synthesis of compound (III) To a stirred mixture of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 3-4, 300.00 mg, 0.68 mmol, 1.00 equiv.) and K2CO3 (180 mg, 1.30 mmol, 1.91 equiv.) in DMF (0.50 mL), chloromethyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (compound 3-3, 600 mg, 1.70 mmol, 2.50 equiv.) and TBAI (119.00 mg, 0.46 mmol, 0.67 equiv.) were added at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature overnight. LCMS showed the reaction was complete. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), gradient from 0% to 85% over 40 min; detector, UV 254 nm. This afforded [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (Compound (III), 85 mg, 14.85%) as a brown solid. The crude product was further purified using the following conditions: Column: XBridge Preparative OBD C18 column, 19 x 250 mm, 5 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 53 B to 68 B in 10 min; 220 nm; RT1: 9.80; Collected fractions were lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-methyl-2-[(5-nitropyridin-2-yl)disulfanyl]propyl carbonate (8.5 mg) as a white solid. LCMS (ESI): 757,757 [M+H] + ; 1H NMR (300 MHz, DMSO-d6) 9.23 (d, J = 2.4Hz, 1H), 8.75 (s, 1H), 8.58 (d, J = 2.7Hz, 1H), 8.05(d, J = 8.7Hz, 1H), 7.72-7.6 (m, 2H), 7.53 (s, 1H), 7.46 (d, J = 6.3Hz, 1H), 7.25-7.11 (m, 2H), 6.82-6.78 (m, 1H), 5.68-5.63 (m, 2H), 5.35-5.28 (m, 1H), 4.52-4.30 (m, 4H), 4.08 (s, 2H), 3.12-3.06 (m, 1H), 2.87-2.73 (m, 1H), 2.49-2.44 (m, 1H), 2.28 (s, 3H), 2.10-2.00 (m, 1H), 1.33 (s, 6H).

[0470]

change

[0471] [ka] Step 1. Synthesis of compound 5-2 To a stirred mixture of 2-carboxybenzaldehyde (compound 5-1, 2.00 g, 12.65 mmol, 1.00 equiv) in MeOH (27.00 mL) was added CHNH 2. A solution of HCl (0.79 g, 25.30 mmol, 2.00 equiv) in HO (4.00 mL) was added at 0 °C. The resulting mixture was stirred at 25 °C for 1 h. To the above mixture, NaBH (0.24 g, 6.33 mmol, 0.50 equiv) was added at 25 °C. The resulting mixture was stirred at 25 °C for an additional 0.5 h. LCMS showed that the reaction was complete. The reaction was quenched by adding acetone (20 mL) at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by trituration with acetone (30 mL). This afforded 2-[(methylamino)methyl]benzoic acid (compound 5-2, 2 g, 86%) as a white solid. LCMS (ES, m / z): 166 [M+H] +

[0472] Step 2. Synthesis of compound 5-3 To a stirred solution of 2-[(methylamino)methyl]benzoic acid (compound 5-2, 1.00 g, 5.44 mmol, 1.00 equiv.) and 20.00 mL of 1 M NaOH in HO in dioxane in a stirred dioxane solution in dioxane was added (Boc)O (2.38 g, 10.88 mmol, 2.00 equiv.) at 0° C. The resulting mixture was stirred at 25° C. for 2 h. LCMS showed that the reaction was complete. The resulting mixture was concentrated under reduced pressure. The residue was acidified to pH 3 with HCl (1 N, aq.). The resulting mixture was extracted with EtOAc (3×30 mL). The combined organic layers were washed with brine (30 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The crude product was used in the next step without further purification. LCMS (ES, m / z): 266 [M+H] +

[0473] Step 3. Synthesis of compound 5-4 To a stirred mixture of 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoic acid (compound 5-3, 500 mg, 1.70 mol, 1 equiv) in DCM (6 mL) and HO (7.5 mL) was added NaHCO (570 mg, 6.78 mmol, 4 equiv) and tetrabutylammonium hydrogen sulfate (57 mg, 0.17 mmol, 0.10 equiv) at 0 °C. The resulting mixture was stirred at 0 °C for 10 min. To the above mixture was added chloromethanesulfonyl chloride (303 mg, 2.04 mol, 1.20 equiv) at 0 °C. The resulting mixture was stirred at 25 °C for an additional 3 h. LCMS showed the reaction was complete. The reaction was quenched by adding water (20 mL) at room temperature. The resulting mixture was extracted with CHCl (3 × 30 mL). The combined organic layers were washed with brine (21 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure to give chloromethyl 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoate (compound 5-4, 250 mg, 42%) as a yellow oil. LCMS (ES, m / z): 314,316 [M+H] + , 214,216 [M+H-100] +

[0474] Step 4. Synthesis of compound (V) To a stirred DMF solution (2.00 mL) of a mixture of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-2,3-dihydro-1H-isoindol-5-yl]methyl]urea (compound 5, prepared according to the procedure described for compound 1-5, 100 mg, 0.20 mmol, 1.00 equiv.) and KCO (84 mg, 0.61 mmol, 3.00 equiv.), chloromethyl 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoate (compound 5-4, 128 mg, 0.40 mmol, 2 equiv.) was added at 0 °C. The resulting mixture was stirred at 25 °C for 16 h. LCMS indicated the reaction was complete. The reaction was quenched by adding water (10 mL) at room temperature. The resulting mixture was extracted with EtOAc (3 × 20 mL). The combined organic layers were washed with brine (20 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (1:1). The crude product was purified by preparative HPLC using the following conditions: column, XSelect CSH Fluorophenyl, 30 mm × 150 mm, 5 μm; mobile phase, water (0.1% FA) and ACN (45% to 58% in 10 min); detector, UV 254 nm. The collected fractions were lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-2,3-dihydro-1H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoate (compound (V), 9.4 mg, 6%) as a white solid. LCMS (ES, m / z): 716,718 [MH] - 1H-NMR (DMSO-d6, 400 MHz) δ (ppm): 8.83 (br s, 1H), 7.83-7.80 (m, 1H), 7.71-7.61 (m, 3H), 7.51-7.38 (m, 3H), 7.19-7.11 (m, 3H), 6.90 (br s, 1H), 5.93-5.82 (m, 2H), 5.35-5.30 (m, 1H), 4.67 (d, J = 6.8 Hz, 2H), 4.46-4.29 (m, 4H), 3.20-3.05 (m, 1H), 2.96-2.86 (m, 4H), 2.44-2.43 (m, 1H), 2.22 (s, 3H), 2.08-2.06 (m, 1H), 1.43-1.26 (m, 9H).

[0475] [ka] Step 1. Synthesis of compound 6-2 To a stirred mixture of tert-butyl N-[2-(methylamino)ethyl]carbamate (compound 6-1, 2.00 g, 11.48 mmol, 1.00 equiv.) and TEA (1.40 g, 13.83 mmol, 1.21 equiv.) in DCM (20.00 mL) was added dropwise a solution of CbzCl (2.05 g, 12.01 mmol, 1.05 equiv.) in DCM (5 mL) at 0 °C. The resulting mixture was stirred at room temperature for 1 h. LCMS showed that the reaction was complete. The reaction was quenched by adding water. The resulting mixture was extracted with CHCl (3 × 20 mL). The combined organic layers were washed with brine (20 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. This gave tert-butyl N-(2-[[(benzyloxy)carbonyl](methyl)amino]ethyl)carbamate (compound 6-2, 3.5 g, 98%) as a pale yellow oil. LCMS (ms, ESI): 309 [M+H] + ,331 [M+Na] +

[0476] Step 2. Synthesis of compound 6-3 To a stirred solution of tert-butyl N-(2-[[(benzyloxy)carbonyl]-(methyl)amino]ethyl)carbamate (compound 6-2, 1.50 g) in DCM (20.00 mL) was added dropwise HCl (4N) in 1,4-dioxane (20.00 mL) at 0° C. under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 1 hour. LCMS showed that the reaction was complete. The resulting mixture was concentrated under reduced pressure to give benzyl N-(2-aminoethyl)-N-methylcarbamate hydrochloride (compound 6-3, 1.4 g, crude) as a white solid. LCMS (ESI, ms): 209 [M+H] +

[0477] Step 3. Synthesis of Compound 6-5 To a stirred solution of benzyl N-(2-aminoethyl)-N-methylcarbamate hydrochloride (compound 6-3, 1.20 g, 4.90 mmol, 1.00 equiv.) and K2CO3 (2.03 g, 14.71 mmol, 3.00 equiv.) in ACN (150 mL) was added KI (0.41 g, 2.45 mmol, 0.50 equiv.) and ethanol, 2-(2-chloroethoxy)- (0.73 g, 5.88 mmol, 1.20 equiv.) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred overnight at 60 °C under a nitrogen atmosphere. LCMS showed the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with MeCN (3 × 100 mL). The filtrate was concentrated under reduced pressure. The resulting mixture was used directly in the next step without further purification. LCMS (ESI, ms): 297 [M+H] +

[0478] Step 4. Synthesis of compound 6-6 To a stirred solution of benzyl N-(2-[[2-(2-hydroxyethoxy)ethyl]amino]ethyl)-N-methylcarbamate (compound 6-5, 1.40 g, 4.72 mmol, 1.00 equiv.) and NaHCO3 (396 mg, 4.72 mmol, 1.00 equiv.) in THF (14.00 mL) and HO (14.00 mL) was added Boc2O (1.03 g, 4.72 mmol, 1.00 equiv.) at 0 °C under a nitrogen atmosphere. The resulting mixture was stirred at room temperature under a nitrogen atmosphere overnight. LCMS showed the reaction was complete. The resulting mixture was extracted with EtOAc (3 × 20 mL). The combined organic layers were washed with brine (3 × 20 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to give benzyl N-[2-[(tert-butoxycarbonyl)[2-(2-hydroxyethoxy)ethyl]amino]ethyl]-N-methylcarbamate (compound 6-6, 1.4 g, 74%) as a white solid. LCMS (ESI, ms): 397 [M+H] + 1 H NMR (300 MHz, chloroform-d) δ 7.42-7.29 (m, 5H), 5.13 (s, 2H), 3.81-3.31 (m, 12H), 2.97 (t, J = 2.7 Hz, 3H), 1.46 (s, 9H).

[0479] Step 5. Synthesis of compounds 6-7 To a stirred solution of benzyl N-[2-[(tert-butoxycarbonyl)[2-(2-hydroxyethoxy)ethyl]amino]ethyl]-N-methylcarbamate (compound 6-6, 1.30 g, 3.28 mmol, 1.00 equiv.) in EtOH (65.00 mL) was added Pd / C (26 mg, 10%) at room temperature. The resulting mixture was stirred overnight at room temperature under an H atmosphere. LCMS showed the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with EtOH (3 × 10 mL). The filtrate was concentrated under reduced pressure to give tert-butyl N-[2-(2-hydroxyethoxy)ethyl]-N-[2-(methylamino)ethyl]carbamate (800 mg, 93%) as an off-white solid. 1H NMR (300 MHz, クロロホルム-d) δ 3.70-3.64 (m, 2H), 3.59 (d, J = 5.7 Hz, 2H), 3.55-3.50 (m, 2H), 3.39 (s, 4H), 3.11 (s, 1H), 2.77 (t, J = 6.3 Hz, 2H), 2.41 (s, 3H), 1.44 (s, 9H).

[0480] ステップ6. Synthesis of Compound (VI) To a stirred DMF solution (2.00 mL) of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 6-7, 200 mg, 0.45 mmol, 1.00 equiv.) was added KCO (188 mg, 1.36 mmol, 3.00 equiv.) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 30 minutes under a nitrogen atmosphere. To the above mixture, chloromethyl 4-nitrophenyl carbonate (105 mg, 0.45 mmol, 1.00 equiv.), NaI (34 mg, 0.22 mmol, 0.50 equiv.), and TBAI (167 mg, 0.45 mmol, 1.00 equiv.) were added at room temperature. The resulting mixture was stirred at room temperature for an additional 1 hour. Then, tert-butyl N-[2-(2-hydroxyethoxy)ethyl]-N-[2-(methylamino)ethyl]carbamate (238 mg, 0.90 mmol, 2.00 equiv.) was added. The resulting mixture was stirred overnight at room temperature under a nitrogen atmosphere. LCMS showed the reaction was complete. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), 10% to 80% gradient over 40 min; detector, UV 254 nm. The collected fractions were lyophilized. This gave [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-[2-[(tert-butoxycarbonyl)[2-(2-hydroxyethoxy)ethyl]amino]ethyl]-N-methylcarbamate (Compound (VI), 50 mg, 14.52%) as a white solid. The crude product (50 mg) was purified by preparative HPLC using the following conditions: column, XSelect CSH Fluorophenyl, 30 mm × 150 mm, 5 μm; mobile phase, water (0.05% FA) and ACN (phase B from 43% to 63% in 7 min); detector, UV 254 nm.The collected fractions were lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl N-[2-[(tert-butoxycarbonyl)[2-(2-hydroxyethoxy)-ethyl]amino]ethyl]-N-methylcarbamate (13.3 mg, 3.86%) as a white solid. LCMS (ESI, ms): 759,761 [M+H]. + , 559,561[M+H-100] + 1 H NMR (400 MHz, DMSO-d6) δ 8.75 (s, 1H), 7.75 - 7.60 (m, 2H), 7.54 - 7.41 (m, 2H), 7.23 - 7.10 (m, 2H), 6.80 (t, J = 6.0 Hz, 1H), 5.64 - 5.47 (m, 2H), 5.26-5.22 (m, 2H), 4.56 (br s, 1H), 4.50 - 4.28 (m, 4H), 3.46 (d, J = 5.2 Hz, 4H), 3.42-3.41 (m, 2H), 3.29-3.28 (m, 2H), 3.28 - 3.17 (m, 4H), 3.07-3.05 (m, 1H), 2.87 - 2.76 (m, 4H), 2.49-2.47(m, 1H), 2.23 (s, 3H), 2.07 (s, 1H), 1.36 (s, 9H).

[0481] [ka] Step 1. Synthesis of compound 7-3 To a stirred solution of 3-[5-(aminomethyl)-1-oxo-3H-isoindol-2-yl]piperidine-2,6-dione (compound 7-1, prepared according to the procedure described for compound 1-4, 1.00 g, 3.66 mmol, 1.00 equiv.) and TEA (0.37 g, 3.66 mmol, 1.00 equiv.) in DMF (10 mL) was added CDI (0.59 g, 3.66 mmol, 1.00 equiv.) at 0 °C under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 2 hours under a nitrogen atmosphere. To the above mixture, DMAP (1.34 g, 10.98 mmol, 3.00 equiv.) and 3-chloro-p-toluidine (0.52 g, 3.66 mmol, 1.00 equiv.) were added at room temperature. The resulting mixture was further stirred at 60 °C overnight. LCMS showed the reaction was complete. The mixture was cooled to room temperature. The reaction mixture was poured into ice / water. The resulting mixture was then filtered, and the filter cake was washed with MeCN (3 x 50 mL). The filter cake was dried under infrared light. This afforded 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 7-3, 1.1 g, 68%) as a white solid. LCMS (ESI, ms): 441,443 [M+H] + .

[0482] Step 2. Synthesis of compound (VII) To a stirred DMF solution (2.00 mL) of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 7-3, 200.00 mg, 0.45 mmol, 1.00 equiv.) was added KCO (188 mg, 1.36 mmol, 3.00 equiv.) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 30 minutes under a nitrogen atmosphere. To the above mixture, chloromethyl 4-nitrophenyl carbonate (compound 7-4, 315 mg, 1.36 mmol, 3.00 equiv.), TBAI (84 mg, 0.23 mmol, 0.50 equiv.), and NaI (68 mg, 0.45 mmol, 1.00 equiv.) were added at room temperature. The resulting mixture was stirred at room temperature for an additional 1 hour. Then, (2S)-tert-butyl 2-[(methylamino)methyl]pyrrolidine-1-carboxylate (compound 7-5, 194 mg, 0.91 mmol, 2.00 equiv.) was added. The final reaction mixture was stirred at room temperature for 1 h. LCMS showed the reaction was complete. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), 10% to 80% gradient over 40 min; detector, UV 254 nm. The collected fractions were concentrated under vacuum to give tert-butyl (2S)-2-([[([3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methoxy)carbonyl](methyl)amino]methyl)pyrrolidine-1-carboxylate (compound (VII), 5 mg, 2%) as a white solid. LCMS (ESI, ms): 711,713 [M+H] + ,611,613[M+H-100] + . 1H NMR (400 MHz, DMSO-d6) δ 8.75 (s, 1H), 7.71-7.60 (m, 2H), 7.52-7.46 (m, 2H), 7.19-7.13 (m, 2H), 6.80 (s, 1H), 5.61-5.47 (m, 2H),5.32-5.18 (m, 1H), 4.60 (s, 1H), 4.52-4.26 (m, 4H), 3.46-3.40(m, 4H), 3.39(s, 1H), 3.30-3.22(m, 4H), 3.14-3.08(m, 1H), 2.91-2.78(m, 4H), 2.33(s, 1H), 2.22(s, 3H), 2.07(s, 1H), 1.36(s, 9H).

[0483] [ka] Step 1. Synthesis of compound 8-2 To a stirred mixture of 2-carboxybenzaldehyde (compound 8-1, 10 g, 63.27 mmol, 1.00 equiv) in MeOH (100 mL) was added CHNH (65.0 mL, 129.72 mmol, 2N in THF, 2.05 equiv) in HO (20 mL) at 0 °C. The resulting mixture was stirred at 25 °C for 1 h. To the above mixture was added NaBH (1.20 g, 31.72 mmol, 0.50 equiv) at 25 °C. The resulting mixture was stirred at 25 °C for an additional 2 h. LCMS showed the reaction was complete. The reaction was quenched by adding acetone (100 mL) at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by trituration with acetone (100 mL). This gave 2-[(methylamino)methyl]benzoic acid (compound 8-2, 10.4 g, 84%) as an off-white solid. LCMS (ES, m / z): 166 [M+H] + .

[0484] Step 2. Synthesis of compound 8-3 To a stirred mixture of 2-[(methylamino)methyl]benzoic acid (compound 8-2, 9 g, 54.48 mmol, 1.00 equiv.) in dioxane (90 mL) was added 1 M NaOH in HO (90 mL) and (Boc)O (24 g, 108.96 mmol, 2.00 equiv.) at 0 °C. The resulting mixture was stirred at 25 °C for 4 h. LCMS showed the reaction was complete. The residue was acidified to pH 3 with HCl (aq.). The resulting mixture was extracted with CHCl (3 × 300 mL). The combined organic layers were washed with brine (30 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure to give 2-[[(tert-butoxycarbonyl)(methyl)amino]methyl]benzoic acid (compound 8-3, 12 g, 81%) as a yellow oil. LCMS (ES, m / z): 266 [M+H] + , 166 [M+H-100] + .

[0485] Step 3. Synthesis of compound 8-4 To a stirred mixture of 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoic acid (compound 8-3, 8 g, 27.14 mmol, 1.00 equiv) in DCM (80 mL) and HO (80 mL) was added NaHCO (9 g, 108.55 mmol, 4.00 equiv), tetrabutylammonium hydrogen sulfate (0.92 g, 2.71 mmol, 0.10 equiv), and chloromethanesulfonyl chloride (4.9 g, 32.82 mmol, 1.2 equiv) at 0 °C. The resulting mixture was stirred at 25 °C for an additional 5 h. LCMS showed the reaction was complete. The reaction was quenched by adding water (20 mL) at room temperature. The resulting mixture was extracted with CHCl (3 × 100 mL). The residue was purified by silica gel column chromatography eluting with PE / EtOAc (3:1) to give chloromethyl 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoate (compound 8-4, 6.6 g, 65%) as a yellow oil. LCMS (ES, m / z): 314 [M+H] + , 214 [M+H-100] + . 1H NMR( 300MHz, CDCl3): 8.06 (t, J=3Hz,1H), 7.62-7.57 (m, 1H), 7.39-7.30 (m, 2H), 5.95 (s, 2H), 4.87 (s, 2H), 2.92 (d, J=3Hz, 3H), 1.61-1.25 (m, 9H).

[0486] Step 4. Synthesis of Compound 8-6 To a stirred mixture of 1-(3-chloro-4-methylphenyl)-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-2,3-dihydro-1H-isoindol-5-yl]methyl]urea (compound 8-5, prepared according to the procedure described for compound 1-5, 500 mg, 1.13 mmol, 1.00 equiv.) and KCO (470 mg, 3.40 mmol, 3.00 equiv.) in DMF (10 mL), 2-(chloromethyl 2-([[(tert-butoxy)carbonyl](methyl)amino]methyl)benzoate (compound 8-4, 712 mg, 2.29 mmol, 2.00 equiv.) was added at room temperature. The resulting mixture was stirred at room temperature for 16 hours. The desired product could be detected by LCMS. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA, ACN in water, 10% to 80% gradient over 40 min; detector, UV 254 nm. The collected fractions were concentrated to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-2,3-dihydro-1H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-([[(tert-butoxy)carbonyl](methyl)amino]-methyl)benzoate (compound 8-6, 200 mg, 24%) as a semi-solid. LCMS (ES, m / z): 618,620 [M+H-100] + , 718,720 [M+H] + .

[0487] Step 5. Synthesis of Compounds 8-7 [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-[[(tert-butoxycarbonyl)(methyl)amino]methyl]benzoate (compound 8-6, 200 mg, 0.28 mmol, 1.00 equiv) was added portionwise to a stirred mixture of HCl (gas)·1,4-dioxane (4 mL) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 1 hour under a nitrogen atmosphere. LCMS indicated the reaction was complete. The resulting mixture was concentrated in vacuo. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA, ACN in water, 10% to 50% gradient over 30 minutes; detector, UV 254 nm. The mixture was lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-[(methylamino)methyl]benzoate (compound 8-7, 80 mg, 41%) as a white solid. LCMS (ES, m / z): 618,620 [M+H] + , 640,642[M+Na] + .

[0488] Step 6. Synthesis of Compounds 8-9 To a stirred mixture of (2S)-2-(2-[2-[(tert-butoxycarbonyl)amino]acetamido]-acetamido)-3-phenylpropanoic acid (compound 8-8, 1.50 g, 3.95 mmol, 1.00 equiv.) in DMF (15 mL), HATU (2.25 g, 5.92 mmol, 1.50 equiv.), HOBT (0.53 g, 3.92 mmol, 0.99 equiv.), glycine (0.36 g, 4.79 mmol, 1.21 equiv.), and DIEA (1.53 g, 11.84 mmol, 2.99 equiv.) were added at 0° C. The resulting mixture was stirred at 25° C. overnight. LCMS showed 12% of the desired product. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, FA (0.1%), ACN in water, 10% to 50% gradient over 40 min; detector, UV 254 nm. The resulting mixture was concentrated in vacuo to give [(2S)-2-(2-[2-[(tert-butoxycarbonyl)amino]acetamido]acetamido)-3-phenylpropanamido]acetic acid (compound 8-9, 300 mg, 15%) as a white solid. LCMS (ES, m / z): 437 [M+H] + , 337 [M+H-100] + .

[0489] Step 7. Synthesis of compounds 8-10 To a stirred mixture of [(2S)-2-(2-[2-[(tert-butoxycarbonyl)amino]-acetamido]acetamido)-3-phenylpropanamido]acetic acid (compound 8-9, 290 mg, 0.66 mmol, 1.00 equiv) in HCl (gas)·1,4-dioxane (6.0 mL) was added at 0° C. The resulting mixture was stirred at 25° C. for an additional 3 hours. LCMS indicated the reaction was complete. The resulting mixture was concentrated in vacuo to afford [(2S)-2-[2-(2-aminoacetamido)acetamido]-3-phenylpropanamido]acetic acid hydrochloride (compound 8-10, 330 mg, 93%) as a white solid. The crude product was used in the next step without purification. LCMS (ES, m / z): 337 [M+H] + .

[0490] Step 8. Synthesis of Compounds 8-12 To a stirred mixture of [(2S)-2-[2-(2-aminoacetamido)acetamido]-3-phenylpropanamido]acetic acid hydrochloride (Compound 8-10, 320 mg, 0.86 mmol, 1.00 equiv.) in DMSO (6 mL), 2,5-dioxopyrrolidin-1-yl 6-(2,5-dioxopyrrol-1-yl)hexanoate (Compound 8-11, 318 mg, 1.03 mmol, 1.20 equiv.) and DIEA (333 mg, 2.58 mmol, 3.0 equiv.) were added portionwise at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for an additional 3 hours. LCMS indicated the reaction was complete. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA (0.5%), ACN in water, 10% to 50% gradient over 30 min; detector, UV 254 nm. The collected fractions were lyophilized to afford [(2S)-2-(2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido]acetamido)-3-phenylpropanamido]acetic acid (compound 8-12, 330 mg, 68%) as a white solid. LCMS (ES, m / z): 530 [M+H] + , 552 [M+Na] + . 1 H-NMR (300 MHz, DMSO-d6) δ: 8.36-8.30 (m, 1H), 8.11-8.06 (m, 2H), 8.06-7.97 (m, 1H), 7.26-7.15 (m, 5H), 6.99 (s, 2H), 4.57-4.49 (m, 1H), 3.79-3.59 (m, 6H), 3.37 (t, J=6 Hz, 2H), 3.04 (t, J=9 Hz, 1H), 2.82-2.77 (m, 1H), 2.11 (t, J=9 Hz, 2H), 1.52-1.44 (m, 4H), 1.24-1.16 (m, 2H).

[0491] Step 9. Synthesis of compound (VIII) To a stirred mixture of [(2S)-2-(2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]-acetamido]acetamido)-3-phenylpropanamido]acetic acid (compound 8-7, 60 mg, 0.11 mmol, 1.00 equiv.) and HATU (65 mg, 0.17 mmol, 1.50 equiv.) in DMF (2 mL) was added HOBT (15 mg, 0.11 mmol) under a nitrogen atmosphere. , 1.0 equiv.), [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-[(methylamino)methyl]benzoate (70 mg, 0.11 mmol, 1.00 equiv.), and DIEA (44 mg, 0.34 mmol, 3.0 equiv.) were added portionwise at room temperature. The resulting mixture was stirred at room temperature under a nitrogen atmosphere for 2 hours. LCMS showed the reaction was complete. The crude product was purified by preparative HPLC using the following conditions: Column: YMC-Actus Triart C18, 30 mm × 150 mm, 5 μm; Mobile Phase A: water (0.05% TFA), Mobile Phase B: ACN; Flow Rate: 60 mL / min. The collected fractions were lyophilized to give [3-[5-([[(3-chloro-4-methylphenyl)carbamoyl]amino]methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl]methyl 2-([2-[(2S)-2-(2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido]acetamido)-3-phenylpropanamido]-N-methylacetamido]methyl)benzoate (compound (VIII), 40 mg, 30%) as a white solid. LCMS (ES, m / z): 566 [M / 2+1] + , 1129, 1131[M+1] + , 1151,1153[M+Na] + . 1H-NMR (300 MHz, DMSO-d6) δ: 8.77 (s, 1H), 8.28-7.80 (m, 5H), 7.73-7.40 (m, 6H), 7.30-7.10 (m, 8H), 6.99 (s, 2H), 6.85-6.80 (m, 1H), 5.95-5.84 (m, 2H), 5.36-5.30 (m, 1H), 4.88-4.82 (m, 2H), 4.62-4.25 (m, 5H), 4.12 (d, J=3 Hz, 1H), 3.89 (d, J=3 Hz, 1H), 3.70-3.65 (m, 5H), 3.36 (t, J=6 Hz, 2H), 3.20-2.70 (m, 7H), 2.23 (s, 3H), 2.10 (t, J=9 Hz, 3H), 1.50-1.44 (m, 4H), 1.28-1.10 (m, 2H).

[0492] [ka] Step 1. Synthesis of compound 9-2 To a stirred solution of (2-chloro-4-nitrophenyl)acetic acid (compound 9-1, 10 g, 46.38 mmol, 1.00 equiv.) in THF (100 mL) was added BH3-Me2S (8.8 g, 115.97 mmol, 2.50 equiv.) portionwise at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at 70 °C for 2 h under a nitrogen atmosphere. TLC showed the reaction was complete. The reaction mixture was cooled to room temperature and concentrated to dryness. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (2:1) to give 2-(2-chloro-4-nitrophenyl)ethanol (compound 9-2, 6.4 g, 68%) as a red oil. 1 H NMR (300 MHz, CDCl3) δ 8.22 (s, 1H), 8.07-8.03 (m, 1 H), 7.51 (d, J = 3 Hz, 1H), 3.92 (t, J = 6 Hz, 2H), 3.09 (t, J = 6 Hz, 2H).

[0493] Step 2. Synthesis of compound 9-3 To a stirred solution of 2-(2-chloro-4-nitrophenyl)ethanol (compound 9-2, 6.4 g, 31.74 mmol, 1.00 equiv.) in DCM (120 mL), NBS (8.48 g, 47.64 mmol, 1.50 equiv.) and PPh3 (12.50 g, 47.62 mmol, 1.50 equiv.) were added portionwise at room temperature. The resulting mixture was stirred overnight at room temperature. TLC showed the reaction was complete. The reaction was concentrated to dryness under vacuum. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (4:1) to give 1-(2-bromoethyl)-2-chloro-4-nitrobenzene (7.0 g, 66%) as a red oil. 1 H NMR (Compound 9-3, 300 MHz, CDCl3) δ 8.28 (d, J = 2.4 Hz, 1H), 8.13 (d, J = 9.0 Hz, 1H), 7.51 (d, J = 3 Hz, 1H), 3.66 (t, J = 6.0 Hz, 2H), 3.42 (t, J = 6.0 Hz, 2H).

[0494] Step 3. Synthesis of compound 9-4 To a stirred mixture of 1-(2-bromoethyl)-2-chloro-4-nitrobenzene (compound 9-3, 6 g, 22.68 mmol, 1.00 equiv.) in EtOH (60 mL) was added sodium methanethiolate (1.92 g, 27.45 mmol, 1.21 equiv.) portionwise at 0 °C under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 3 h under a nitrogen atmosphere. LCMS failed to detect the desired product, but TLC (PE:EA = 10:1) showed a new spot. The resulting mixture was concentrated in vacuo. The residue was purified by silica gel column chromatography eluting with PE / EtOAc (9:1) to give 2-chloro-1-[2-(methylsulfanyl)ethyl]-4-nitrobenzene (compound 9-4, 1.7 g, 32%) as a yellow solid. 1H NMR (300MHz, CDCl3): 8.28 (t, J=3Hz,1H), 8.10 (d, J=3Hz,1H), 7.45 (d, J=9Hz,1H), 3.14 (t, J=6Hz, 2H), 2.80 (t, J=3Hz, 2H), 2.18 (s, 3H).

[0495] Step 4. Synthesis of compound 9-5 To a stirred mixture of 2-chloro-1-[2-(methylsulfanyl)ethyl]-4-nitrobenzene (compound 9-4, 2 g, 8.63 mmol, 1.00 equiv.) and Fe (1.45 g, 25.96 mmol, 3.01 equiv.) in EtOH (60 mL) was added NH4Cl (4.6 g, 86.32 mmol, 10.00 equiv.) in HO (20 mL). The resulting mixture was stirred at 90 °C for 3 h. LCMS showed the reaction was complete. The mixture was cooled to room temperature. The resulting mixture was filtered, and the filter cake was washed with CHCl2. The resulting mixture was concentrated in vacuo. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA (0.05%), ACN in water, gradient from 10% to 50% over 40 min; detector, UV 254 nm. The collected fractions were concentrated to give 3-chloro-4-[2-(methylsulfanyl)ethyl]aniline (compound 9-5, 2.0 g, 69%) as a yellow oil. LCMS (ESI, ms): 202, 204 [M+H] + ,243,245[M+H+ACN] + .

[0496] Step 5. Synthesis of compound 9-6 To a stirred mixture of 3-[5-(aminomethyl)-1-oxo-3H-isoindol-2-yl]piperidine-2,6-dione (INT1, prepared according to the procedure described for Compound 1-4, 1.4 g, 4.96 mmol, 1.00 equiv.) in DMF (25 mL), CDI (0.80 g, 4.96 mmol, 1.00 equiv.) and TEA (0.50 g, 4.94 mmol, 1.00 equiv.) were added portionwise at 0 °C under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 3 hours under a nitrogen atmosphere. To the above mixture, 3-chloro-4-[2-(methylsulfanyl)ethyl]aniline (Compound 9-5, 1 g, 4.96 mmol, 1.00 equiv.) and DMAP (1.82 g, 14.90 mmol, 3.00 equiv.) were added portionwise at room temperature. The resulting mixture was stirred overnight at 60 °C under a nitrogen atmosphere. LCMS revealed 58% of the desired product. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA (0.05%), ACN in water, gradient from 10% to 70% in 40 min; detector, UV 254 nm. The collected fractions were concentrated to give 1-[3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 9-6, 700 mg, 27%) as a solid. LCMS (ES, m / z): 501, 503 [M+H] + . 1 H-NMR (300 MHz, DMSO-d6) δ 10.98 (s, 1H), 8.79 (s, 1H), 7.71-7.67 (m, 2H), 7.52-7.25 (m, 2H), 7.25-7.15 (m, 2H), 6.82 (t, J=6 Hz, 1H), 5.14-5.08 (m, 1H), 4.49-4.29 (m, 4H), 2.98-2.84 (m, 3H), 2.84-2.63 (m, 3H), 2.42-2.34 (m, 1H), 2.09 (s, 3H), 2.03-1.90 (m, 1H).

[0497] Step 6. Synthesis of Compound 9-7 To a stirred mixture of 1-[3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]-3-[[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl]urea (compound 9-6, 700 mg, 1.40 mmol, 1.00 equiv.) and KCO (579 mg, 4.19 mmol, 3.00 equiv.) in DMF (10 mL), 2-[[(tert-butoxycarbonyl)(methyl)amino]methyl]benzoic acid chloromethyl ester (compound 8-4, 526 mg, 1.68 mmol, 1.20 equiv.) was added portionwise at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 2 days under a nitrogen atmosphere. LCMS indicated the reaction was complete. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA (0.05%), ACN in water, 30% to 80% gradient over 40 min; detector, UV 254 nm. The collected fractions were lyophilized to afford [3-(5-[[([3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]carbamoyl)amino]methyl]-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl 2-[[(tert-butoxycarbonyl)(methyl)amino]methyl]benzoate (compound 9-7, 260 mg, 21%) as a white solid. LCMS (ES, m / z): 778,780 [M+H] + ,678,780[M+H-100] + .

[0498] Step 7. Synthesis of Compounds 9-8 [3-(5-[[([3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]carbamoyl)amino]methyl]-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl 2-[[(tert-butoxycarbonyl)(methyl)amino]methyl]benzoate (compound 9-7, 250 mg, 0.32 mmol, 1.00 equiv) was added portionwise to a stirred mixture of HCl (gas)·1,4-dioxane (5 mL) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at room temperature for 1 hour under a nitrogen atmosphere. LCMS showed the reaction was complete. The resulting mixture was concentrated in vacuo. The crude product was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, TFA (0.05%), ACN in water, 10% to 50% gradient over 40 min; detector, UV 254 nm. The mixture was lyophilized to afford [3-(5-[[([3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]carbamoyl)amino]methyl]-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl 2-[(methylamino)methyl]benzoate (compound 9-8, 132 mg, 56%) as a yellow solid. LCMS (ES, m / z): 678,680 [M+H] + ,700,702[M+Na] + .

[0499] Step 8. Synthesis of compound (IX) To a stirred DMF solution (2.00 mL) of a mixture of [(2S)-2-(2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido]acetamido)-3-phenylpropanamido]acetic acid (Compounds 8-12, 100 mg, 0.19 mmol, 1.00 equiv.) and HATU (108 mg, 0.28 mmol, 1.5 equiv.) was added HOBT (26 mg, 0.19 mmol, 1.0 equiv.), [3 -(5-[[([3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]carbamoyl)amino]methyl]-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl 2-[(methylamino)methyl]benzoate (compound 9-8, 115 mg, 0.17 mmol, 0.90 equiv.) and DIEA (73 mg, 0.57 mmol, 3.0 equiv.) were added portionwise at room temperature. The resulting mixture was stirred at room temperature under a nitrogen atmosphere for 2 hours. LCMS showed the reaction was complete. The crude product was purified by preparative HPLC using the following conditions (Column: XSelect CSH preparative C18 OBD column, 19 × 250 mm, 5 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min). The collected fractions were lyophilized to give [3-(5-[[([3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl]carbamoyl)amino]methyl]-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl 2-([2-[(2S)-2-(2-[2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido]acetamido)-3-phenylpropanamido]-N-methylacetamido]methyl)benzoate (Compound (IX), 52.1 mg, 23%) as a white solid. LCMS (ES, m / z): 596 [M / 2+1] + ,1189,1191[M+1] + . 1H-NMR (300 MHz, DMSO-d6) δ 8.82 (s, 1H), 8.12-7.83 (m, 5H), 7.73-7.38 (m, 6H), 7.26-7.15 (m, 8H), 6.99 (s, 2H), 6.83 (t, J=6 Hz, 1H), 5.98-5.84 (m, 2H), 5.32 (t, J=6 Hz, 1H), 4.884.81 (m, 2H), 4.65-4.30 (m, 5H), 4.12 (d, J=3 Hz, 1H), 3.94-3.85 (m, 4H), 3.72-3.59 (m, 3H), 3.36 (t, J=6 Hz, 2H), 3.20-2.95 (m, 4H), 2.89-2.74 (m, 4H), 2.62-2.50 (m, 2H), 2.12-2.05 (m, 6H), 1.58-1.40 (m, 4H), 1.28-1.10 (m, 2H).

[0500] [ka] Step 1. Synthesis of compound 10-2 To a stirred mixture of Gly-Gly (10 g, 75.69 mmol, 1.00 equiv.) and NaHCO (12.72 g, 151.3 mmol, 2 equiv.) in HO (70 mL) was added chloro(prop-2-en-1-yloxy)methanone (10.95 g, 90.82 mmol, 1.2 equiv.) in THF (35 mL) dropwise at 0 °C. The resulting mixture was stirred at 25 °C for 5 h. LCMS showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. The mixture was acidified to pH 5 with HCl (aqueous, 1N). The precipitated solid was collected by filtration and washed with HCl (aqueous, 1N) (2 × 5 mL). This afforded (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)acetic acid (compound 10-2, 9 g, 55%) as a white solid. LCMS (ES, m / z): 217 [M+H] +

[0501] Step 2. Synthesis of compound 10-3 A mixture of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)acetic acid (compound 10-2, 9 g, 41.62 mmol, 1.00 equiv.) and Cu(OAc) (0.76 g, 4.16 mmol, 0.1 equiv.) in THF (220 mL) was stirred at 60 °C for 1 h under a nitrogen atmosphere. The mixture was cooled to room temperature. To the above mixture, Pb(OAc) (22.15 g, 49.9 mmol, 1.2 equiv.) was added at room temperature. The resulting mixture was stirred at 25 °C for an additional 1 h. LCMS showed that the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with MeOH (3 × 50 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (1:10) to give (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamide)methyl acetate (compound 10-3, 5 g, 52%) as a white solid. LCMS (ES, m / z): 231 [M+H] +

[0502] Step 3. Synthesis of compound 10-4 To a stirred mixture of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl acetate (compound 10-3, 1 g, 4.34 mmol, 1.00 equiv) in DCM (40 mL) was added TMSCl (1.89 g, 17.37 mmol, 4 equiv) dropwise at 0 °C. The resulting mixture was stirred at 0 °C for 1 h. LCMS (quenched with MeOH by LCMS) showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. The crude product was used directly in the next step without further purification. This afforded prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 10-3, 1 g, 77%) as a yellow solid. LCMS (ES, m / z): 203 [M+H] + (quench with MeOH)

[0503] Step 4. Synthesis of compound 10-6 A mixture of 1-(3-chloro-4-methylphenyl)-3-{[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl}urea (compound 10-5, prepared according to the procedure described for compound 1-5, 750 mg, 1.70 mmol, 1.00 equiv.) and AgCO (938 mg, 3.40 mmol, 2 equiv.) in NMP (15.00 mL) was stirred at 50 °C for 1 h. The mixture was cooled to room temperature. To the above mixture was added prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 10-4, 703 mg, 3.40 mmol, 2.00 equiv.) at room temperature. The resulting mixture was stirred at 60 °C for an additional 36 h. LCMS showed the reaction was complete. The residue was purified on a YMC-Actus Triart C18 ExRS column; 30 × 150 mm; mobile phase A: water (0.05% TFA), mobile phase B: ACN; flow rate: 60 mL / min; gradient: 27% B to 53% B, 53% B in 10 min; wavelength: 254 nm; RT1 (min): 9.22 min. This afforded prop-2-en-1-yl N-{[({3-[5-({[(3-chloro-4-methylphenyl)carbamoyl]amino}methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamate (compound 10-6, 100 mg, 8%) as a yellow solid. LCMS (ES, m / z): 611,613 [M+H] +

[0504] Step 5. Synthesis of compound 10-7 To a stirred mixture of prop-2-en-1-yl N-{[({3-[5-({[(3-chloro-4-methylphenyl)carbamoyl]amino}methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamate (compound 10-6, 100 mg, 0.17 mmol, 1.00 equiv.) and Pd(PPh3)4 (19 mg, 0.017 mmol, 0.10 equiv.) in THF (1.50 mL) was added phenylsilane (36 mg, 0.34 mmol, 2.00 equiv.) dropwise at 25 °C under a nitrogen atmosphere. The resulting mixture was stirred at 25 °C for 2 hours under a nitrogen atmosphere. LCMS showed the reaction was complete. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), ACN (5% to 50% gradient in 30 min); detector, UV 254 nm. This afforded 2-amino-N-({3-[5-({[(3-chloro-4-methylphenyl)carbamoyl]amino}methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)acetamide (compound 10-7, 70 mg, 79%) as a yellow solid. LCMS (ES, m / z): 527,529 [M+H] +

[0505] Step 6. Synthesis of Compound X [(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetic acid (compound 10-8, prepared according to the procedure described for compounds 8-12, 65 mg, 0.12 mmol, 1 equiv.) and a mixture of HATU (56 mg, 0.14 mmol, 1.2 equiv.), HOBT (20 mg, 0.14 mmol, 1.2 equiv.). To a stirred DMF solution (650 μL) was added 2-amino-N-({3-[5-({[(3-chloro-4-methylphenyl)carbamoyl]amino}methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)acetamide (compound 10-7, 65 mg, 0.12 mmol, 1.00 equiv.) and DIEA (47 mg, 0.36 mmol, 3 equiv.) at 0 °C. The resulting mixture was stirred at 25 °C for 16 h. LCMS indicated the reaction was complete. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.5% TFA), 10% to 50% ACN gradient over 30 min; detector, UV 254 nm. The crude product was re-purified by preparative HPLC using the following conditions: Column: Kinetex EVO Prep C18, 30 x 150, 5 μm; Mobile phase A: Water (0.05% TFA), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 25% B to 35% B, 35% B in 17 min; Wavelength: 254 nm; RT1 (min): 16. The collected fractions were lyophilized to give N-{[({[(1S)-1-{[({[({3-[5-({[(3-chloro-4-methylphenyl)carbamoyl]amino}methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamoyl)methyl]carbamoyl}-2-phenylethyl]carbamoyl}methyl)carbamoyl]methyl}-6-(2,5-dioxopyrrol-1-yl)hexanamide (compound (X), 11.4 mg, 8.84%) as a white solid. LCMS (ES, m / z): 1038,1040 [M+H] + . 1H-NMR (DMSO, 400 MHz) δ (ppm): 8.75 (s, 1H), 8.31-8.29 (m, 1H), 8.19-8.16 (m, 1H), 8.12-8.05 (m, 2H), 7.99-7.94 (m, 2H), 7.71-7.66 (m, 2H), 7.52 (s, 1H), 7.44 (d, J = 8.0 Hz, 1H), 7.24-7.08 (m, 7H), 6.99-6.86 (m, 2H), 6.82-6.79 (m, 1H), 5.20-5.13 (m, 2H), 4.99-4.95 (m, 1H), 4.49-4.40 (m, 4H), 4.31-4.27 (m, 1H), 3.76-3.56 (m, 8H), 3.37-3.30 (m, 2H), 3.07-2.97 (m, 2H), 2.79-2.67 (m, 2H), 2.40-2.30 (m, 1H), 2.22 (s, 3H), 2.11-2.02 (m, 3H), 1.49-1.42 (m, 4H), 1.23-1.14 (m, 2H).

[0506]

change

[0507] Step 2. Synthesis of compound 11-4 To a stirred mixture of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl acetate (compound 11-3, 1 g, 4.34 mmol, 1.00 equiv.) in DCM (40 mL) was added TMSCl (1.89 g, 17.37 mmol, 4 equiv.) dropwise at 0 °C. The resulting mixture was stirred at 0 °C for 2 h. LCMS showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. This afforded prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 11-4, 0.8 g, 89%) as a white solid. LCMS (ES, m / z): 203 [M+H] + (Quenched with MeOH for LCMS)

[0508] Step 3. Synthesis of compound 11-5 To a stirred mixture of tert-butyl N-[2-(2-{2-chloro-4-[({[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl}carbamoyl)amino]phenyl}ethoxy)ethyl]-N-methylcarbamate (compound 11-2, 1 g, 1.59 mmol, 1.00 equiv.) and K2CO3 (0.44 g, 3.18 mmol, 2 equiv.) in NMP (16 mL) was added prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 11-4, 0.82 g, 3.98 mmol, 2.5 equiv.) in portions at 25 °C. The resulting mixture was stirred at 60 °C for 16 h. LCMS showed the reaction was complete. The mixture was cooled to room temperature. The reaction was quenched by adding water (20 mL) at room temperature. The resulting mixture was extracted with EtOAc (3×30 mL). The combined organic layers were washed with brine (30 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), ACN (5% to 50% gradient in 30 min; detector, UV 254 nm. This afforded tert-butyl N-(2-{2-[2-chloro-4-({[(2-{2,6-dioxo-1-[(2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl]piperidin-3-yl}-1-oxo-3H-isoindol-5-yl)methyl]carbamoyl}amino)phenyl]ethoxy}ethyl)-N-methylcarbamate, compound 11-5 (0.5 g, 39%), as a yellow solid. LCMS (ES, m / z): 798,800 [M+H]+

[0509] Step 4. Synthesis of compound 11-6 To a stirred mixture of tert-butyl N-(2-{2-[2-chloro-4-({[(2-{2,6-dioxo-1-[(2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl]piperidin-3-yl}-1-oxo-3H-isoindol-5-yl)methyl]carbamoyl}amino)phenyl]ethoxy}ethyl)-N-methylcarbamate (compound 11-5, 500 mg, 0.62 mmol, 1.00 equiv) in THF (6 mL) was added Pd(PPh3)4 (72 mg, 0.063 mmol, 0.1 equiv) and phenylsilane (136 mg, 1.25 mmol, 2 equiv) at 0 °C under a nitrogen atmosphere. The resulting mixture was stirred at 25 °C for 3 h under a nitrogen atmosphere. LCMS showed the reaction was complete. The mixture was cooled to room temperature. The reaction was quenched by adding water (20 mL) at room temperature. The resulting mixture was extracted with EtOAc (3 × 30 mL). The combined organic layers were washed with brine (30 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), ACN (gradient from 5% to 50% in 30 min); detector, UV 254 nm. This gave tert-butyl N-(2-{2-[4-({[(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-5-yl)methyl]carbamoyl}amino)-2-chlorophenyl]ethoxy}ethyl)-N-methylcarbamate (compound 11-6, 400 mg, 89%) as a yellow solid. LCMS (ES, m / z): 714,716 [M+H]+

[0510] Step 5. Synthesis of Compound 11-8 A stirred solution of a mixture of [(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetic acid (compounds 11-7, prepared according to the procedure described for compounds 8-12, 163 mg, 0.30 mmol, 1.1 equiv.), HATU (159 mg, 0.42 mmol, 1.5 equiv.), and HOBT (57 mg, 0.42 mmol, 1.5 equiv.) in DMF (3 mL) To the resulting solution was added tert-butyl N-(2-{2-[4-({[(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-5-yl)methyl]carbamoyl}amino)-2-chlorophenyl]ethoxy}ethyl)-N-methylcarbamate (compound 11-6, 200 mg, 0.28 mmol, 1.00 equiv.), DIEA (108 mg, 0.84 mmol, 3 equiv.) at 0° C. The resulting mixture was stirred at 25° C. for 5 hours. LCMS showed that the reaction was complete. The reaction was quenched by adding water (10 mL). The precipitated solid was collected by filtration and washed with water (11 mL). This gave tert-butyl N-(2-{2-[2-chloro-4-({[(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-5-yl)methyl]carbamoyl}amino)phenyl]ethoxy}ethyl)-N-methylcarbamate (compound 11-8, 100 mg, 29%) as a yellow solid. LCMS (ES, m / z): 1225,1227 [M+H] +

[0511] Step 6. Synthesis of compound XI To a stirred mixture of tert-butyl N-(2-{2-[2-chloro-4-({[(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-5-yl)methyl]carbamoyl}amino)phenyl]ethoxy}ethyl)-N-methylcarbamate (compound 11-8, 100 mg, 0.08 mmol, 1.00 equiv) in DCM (0.8 mL) was added TFA (0.2 mL) at 0° C. The resulting mixture was stirred at 25° C. for 4 hours. LCMS showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. The crude product was purified by preparative HPLC using the following conditions: Column: XBridge Shield RP18 OBD column, 30 × 150 mm, 5 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 20% B to 40% B, 40% B in 7 min; Wavelength: 254 nm; RT1 (min): 5.7 min. The collected fractions were lyophilized to give N-{[({[(1S)-1-{[({[({3-[5-({[(3-chloro-4-{2-[2-(methylamino)ethoxy]ethyl}phenyl)carbamoyl]amino}methyl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamoyl)methyl]carbamoyl}-2-phenylethyl]carbamoyl}methyl)carbamoyl]methyl}-6-(2,5-dioxopyrrol-1-yl)hexanamide; trifluoroacetic acid (Compound (XI), 30.5 mg, 29%) as a white solid. LCMS (ES, m / z): 1125,1127 [M+H] + . 1H-NMR (DMSO, 400 MHz) δ (ppm): 8.88 (s, 1H), 8.39 (br s, 2H), 8.33-8.29 (m, 1H), 8.21-8.17 (m, 1H), 8.13-8.06 (m, 2H), 8.01-7.94 (m, 2H), 7.71 (d, J = 7.6 Hz, 1H), 7.72 (d, J = 2.0 Hz, 1H), 7.51 (s, 1H), 7.44 (d, J = 8.0 Hz, 1H), 7.26-7.14 (m, 7H), 6.99 (s, 2H), 6.93-6.90 (m, 1H), 5.26-5.08 (m, 2H), 5.02-4.88 (m, 1H), 4.58-4.39 (m, 4H), 4.35-4.18 (m, 1H), 3.76-3.56 (m, 12H), 3.37-3.34 (m, 2H), 3.10-3.00 (m, 4H), 2.90-2.87 (m, 2H), 2.81-2.75 (m, 2H), 2.57-2.54 (m, 3H), 2.42-2.32 (m, 1H), 2.11-2.08 (m, 2H), 2.06-1.99 (m, 1H), 1.49-1.43 (m, 4H), 1.19-1.16 (m, 2H).

[0512]

change

[0513] Step 2. Synthesis of compound 12-5 To a stirred mixture of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl acetate (compound 12-4, prepared according to the procedure described for compound 10-3, 0.9 g, 3.90 mmol, 1.00 equiv) in DCM (38 mL) was added TMSCl (1.9 mL, 14.86 mmol, 3.80 equiv) dropwise at 0 °C. The resulting mixture was stirred at 0 °C for 2 h. LCMS showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. The crude product was used directly in the next step without further purification. This afforded prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 12-5, 0.8 g, 99%) as a white solid. LCMS (ES, m / z): 203 [M+H] + (methanol-induced)

[0514] Step 3. Synthesis of compound 12-6 To a stirred mixture of tert-butyl N-{5-[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-4-yl]pent-4-yn-1-yl}carbamate (compound 12-3, 1 g, 2.35 mmol, 1.00 equiv.) and prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 12-5, 0.73 g, 3.52 mmol, 1.5 equiv.) in NMP (24 mL) was added K2CO3 (0.65 g, 4.70 mmol, 2 equiv.) at room temperature. The resulting mixture was stirred at 60 °C for 16 h. LCMS showed the reaction was complete. The reaction was quenched by adding water (30 mL) at room temperature. The resulting mixture was extracted with EtOAc (3 × 30 mL). The combined organic layers were washed with brine (3 × 30 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), ACN (5% to 50% gradient in 30 min); detector, UV 254 nm. This afforded tert-butyl N-[5-(2-{2,6-dioxo-1-[(2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl]piperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamate (compound 12-6, 1.2 g, 85%) as a yellow oil. LCMS (ES, m / z): 596 [M+H] +

[0515] Step 4. Synthesis of compound 12-7 To a stirred solution of tert-butyl N-[5-(2-{2,6-dioxo-1-[(2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl]piperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamate (compound 12-6, 1.4 g, 2.35 mmol, 1.00 equiv.) and Pd(PPh3)4 (0.27 g, 0.23 mmol, 0.1 equiv.) in THF (30 mL) was added phenylsilane (0.51 g, 4.70 mmol, 2 equiv.) under a nitrogen atmosphere. The resulting mixture was stirred at 25 °C under a nitrogen atmosphere for 3 hours. LCMS showed the reaction was complete. The reaction was quenched by adding water (1 mL) at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), 30 min gradient of 5% to 50% ACN; detector, UV 254 nm. This afforded tert-butyl N-[5-(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamate; trifluoroacetate (compound 12-7, 500 mg, 34%) as a yellow solid. LCMS (ES, m / z): 512 [M+H] +

[0516] Step 5. Synthesis of Compound 12-9 To a stirred mixture of [(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetic acid (compound 12-8, prepared according to the procedure described for compound 8-12, 279.33 mg, 0.52 mmol, 1.1 equiv) in DMF (6.00 mL) was added HATU (218.80 mg, 0.57 mmol, 1.2 equiv) and HOBT (77.76 mg, 0.57 mmol, 1.2 equiv) at 0° C. The resulting mixture was stirred at 25° C. for 30 min. To the above mixture, tert-butyl N-[5-(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamate; trifluoroacetic acid (compound 12-7, 300 mg, 0.48 mmol, 1.00 equiv.), and DIEA (185.93 mg, 1.44 mmol, 3 equiv.) were added at 0 °C. The resulting mixture was stirred at 25 °C for an additional 16 h. LCMS showed the reaction was complete. The reaction was quenched by adding water (0.5 mL) at room temperature. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), ACN (5% to 50% gradient in 30 min); detector, UV 254 nm. This gave tert-butyl N-[5-(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamate (compound 12-9, 300 mg, 61%) as a yellow solid. LCMS (ES, m / z): 1023 [M+H] +

[0517] Step 6. Synthesis of Compounds 12-10 To a stirred mixture of tert-butyl N-[5-(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamate (compound 12-9, 300 mg, 0.29 mmol, 1.00 equiv) in DCM (3.00 mL), TFA (0.75 mL) was added dropwise at 0° C. The resulting mixture was stirred at 25° C. for 3 hours. LCMS showed the reaction was complete. The resulting mixture was concentrated under reduced pressure. This gave N-{[({[(1S)-1-{[({[({3-[4-(5-aminopent-1-yn-1-yl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamoyl)methyl]carbamoyl}-2-phenylethyl]carbamoyl}methyl)carbamoyl]methyl}-6-(2,5-dioxopyrrol-1-yl)hexanamide; trifluoroacetic acid (compound 12-10, 300 mg, 98%) as a yellow solid. LCMS (ES, m / z): 923 [M+H] +

[0518] Step 7. Synthesis of compound 12-12 To a stirred mixture of (3'S,4'R,5'S)-6''-chloro-4'-(3-chloro-2-fluorophenyl)-2''-oxo-1''H-dispiro[cyclohexane-1,2'-pyrrolidine-3',3''-indole]-5'-carboxylic acid (compound 12-11, prepared as described in J. Med. Chem. 2014, 57, 10486-10498, 300 mg, 0.64 mmol, 1.00 equiv.) and methyl 4-aminobenzoate (117 mg, 0.77 mmol, 1.2 equiv.) in DMA (8 mL) was added DIEA (100 mg, 0.77 mmol, 1.2 equiv.) at 0 °C. The resulting mixture was stirred at 0 °C for 30 min. To the above mixture was added HATU (295 mg, 0.77 mmol, 1.2 equiv.) at 0 °C. The resulting mixture was stirred at 25 °C for an additional 16 hours. LCMS indicated the reaction was complete. The reaction was quenched by adding water (0.5 mL) at room temperature. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), ACN (gradient of 5% to 50% over 30 minutes); detector, UV 254 nm. This afforded methyl 4-[(3'S,4'R,5'S)-6''-chloro-4'-(3-chloro-2-fluorophenyl)-2''-oxo-1''H-dispiro[cyclohexane-1,2'-pyrrolidine-3',3''-indole]-5'-ylamido]benzoate (compound 12-12, 200 mg, 51%) as a yellow solid. LCMS (ES, m / z): 596,598 [M+H] +

[0519] Step 8. Synthesis of compounds 12-13 To a stirred mixture of methyl 4-[(3'S,4'R,5'S)-6''-chloro-4'-(3-chloro-2-fluorophenyl)-2''-oxo-1''H-dispiro[cyclohexane-1,2'-pyrrolidin-3',3''-indole]-5'-ylamido]benzoate (compound 12-12, 190 mg, 0.31 mmol, 1.00 equiv) in THF (2 mL), NaOH (12 mg, 0.31 mmol, 1 equiv) and LiOH (15 mg, 0.63 mmol, 2 equiv) in HO (2 mL) were added dropwise at 0 °C. The resulting mixture was stirred at 25 °C for 4 h. LCMS indicated the reaction was complete. The resulting mixture was concentrated under reduced pressure. The mixture was acidified to pH 6 with HCl (aq). The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), gradient of 5% to 50% ACN over 30 min; detector, UV 254 nm. This afforded 4-[(3'S,4'R,5'S)-6''-chloro-4'-(3-chloro-2-fluorophenyl)-2''-oxo-1''H-dispiro[cyclohexane-1,2'-pyrrolidine-3',3''-indole]-5'-ylamido]benzoic acid (compound 12-13, 50 mg, 26%) as a yellow solid. LCMS (ES, m / z): 582,584 [M+H] +

[0520] Step 9. Synthesis of Compound XII To a stirred mixture of 4-[(3'S,4'R,5'S)-6''-chloro-4'-(3-chloro-2-fluorophenyl)-2''-oxo-1''H-dispiro[cyclohexane-1,2'-pyrrolidine-3',3''-indole]-5'-ylamido]benzoic acid (compound 12-13, 50 mg, 0.086 mmol, 1.00 equiv) in DMF (0.9 mL) was added HATU (36 mg, 0.095 mmol, 1.1 equiv) in portions at 0° C. The resulting mixture was stirred at 0° C. for 10 min. To the above mixture, N-{[({[(1S)-1-{[({[({3-[4-(5-aminopent-1-yn-1-yl)-1-oxo-3H-isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamoyl)methyl]carbamoyl}-2-phenylethyl]carbamoyl}methyl)carbamoyl]methyl}-6-(2,5-dioxopyrrol-1-yl)hexanamide; trifluoroacetic acid (compound 12-10, 98 mg, 0.095 mmol, 1.10 equiv.) and DIEA (33 mg, 0.25 mmol, 3 equiv.) were added at 0° C. The resulting mixture was stirred at 25° C. for an additional 16 hours. LCMS showed the reaction was complete. The reaction was quenched by adding water (0.1 mL) at room temperature. The crude product was purified by preparative HPLC using the following conditions: Column: XBridge Shield RP18 OBD column, 30 × 150 mm, 5 μm; Mobile phase A: water (0.1% FA), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 37% B to 53% B in 10 min, 53% B to 53% B in 11 min; Wavelength: 254 nm; RT1 (min): 10.52.The collected fractions were lyophilized to give (3'R,4'S,5'R)-6''-chloro-4'-(3-chloro-2-fluorophenyl)-N-(4-{[5-(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-1-oxo-3H-isoindol-4-yl)pent-4-yn-1-yl]carbamoyl}phenyl)-2''-oxo-1''H-dispiro[cyclohexane-1,2'-pyrrolidine-3',3''-indole]-5'-carboxamide (compound (XII), 24.1 mg, 18%)) as a white solid. LCMS (ES, m / z): 1486,1488 [M+H]. + 1 H-NMR (DMSO, 400 MHz) δ (ppm): 10.59 (s, 1H), 10.23 (s, 1H), 8.47-8.44 (m, 1H), 8.36-8.26 (m, 1H), 8.25-7.90 (m, 5H), 7.80 (d, J = 8.8 Hz, 2H), 7.76-7.58 (m, 5H), 7.53-7.45 (m, 2H), 7.37-7.34 (m, 1H), 7.26-7.13 (m, 6H), 7.05-6.99 (m, 3H), 6.68 (d, J = 2.0 Hz, 1H), 5.32-5.10 (m, 2H), 5.02-4.90 (m, 1H), 4.77-4.68 (m, 2H), 4.50-4.46 (m, 2H), 4.36-4.32 (m, 1H), 3.76-3.60 (m, 9H), 3.42-3.36 (m, 4H), 3.06-3.02 (m, 2H), 2.82-2.70 (m, 2H), 2.55-2.54 (m, 2H), 2.46-2.43 (m, 1H), 2.11-2.04 (m, 4H), 1.84-1.80 (m, 3H), 1.68-1.52 (m, 4H), 1.49-1.36 (m, 6H), 1.19-1.15 (m, 2H), 1.05-0.96 (m, 1H), 0.92-0.82 (m, 1H).

[0521] [ka] Step 1. Synthesis of compound 13-3 A solution of [(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetic acid (compound 13-1, 1.00 g, 2.50 mmol, 1.00 equiv.) and HATU (1.90 g, 4.99 mmol, 2.00 equiv.) in DMF (10 mL) was stirred at room temperature for 0.5 h. Then, methyl 6-aminohexanoate hydrochloride (compound 13-2, 0.54 g, 2.994 mmol, 1.2 equiv.) and DIEA (1.93 g, 14.97 mmol, 6.00 equiv.) were added. The resulting mixture was stirred at room temperature for 2 h. LCMS showed the reaction was complete. The resulting mixture was diluted with water (100 mL) and extracted with EA (50 mL × 3). The combined organic layers were washed with water (50 mL), brine (50 mL), dried over anhydrous sodium sulfate, and concentrated to dryness under vacuum. The residue was purified by silica gel column chromatography (DCM:MeOH = 13:1) to afford methyl 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexanoate (compound 13-3, 1.22 g, 91%) as a brown solid. LCMS (ES, m / z): 528,530 [M+H] +

[0522] Step 2. Synthesis of compound 13-4 To a solution of methyl 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexanoate (compound 13-3, 1.20 g, 2.27 mmol, 1.00 equiv.) in THF (10 mL) and HO (5 mL) was added LiOH.HO (65 mg, 2.73 mmol, 1.20 equiv.) at room temperature. The resulting mixture was stirred at room temperature for 2 hours. LCMS showed the reaction was complete. The solvent was removed in vacuo. The residue was dissolved in EA (100 mL) and water (300 mL). The organic layer was separated. The aqueous phase was extracted with EA (100 mL × 3). The combined organic layers were dried over anhydrous sodium sulfate and concentrated to dryness under vacuum to give 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexanoic acid (compound 13-4, 1 g, 84%) as a yellow solid. LCMS (ES, m / z): 514,516 [M+H] +

[0523] Step 3. Synthesis of compound 13-6 IR[DF(CF3)PPY]2(DTBPY)PF6 (166 mg, 0.15 mmol, 0.10 equiv.), nickel(2+), tetraaqua[4,4'-bis(1,1-dimethylethyl)-2,2'-bipyridine-κN1,κN1']-chloride (69 mg, 0.15 mmol, 0.10 equiv.), 4-bromo-2-(2,6-dioxopiperidin-3-yl)isoindole- To a DMSO solution (25.00 mL) of 1,3-dione (compound 13-5, 500 mg, 1.48 mmol, 1.00 equiv.) and [(tert-butoxycarbonyl)amino]acetic acid (390 mg, 2.22 mmol, 1.50 equiv.) was added 2-tert-butyl-1,1,3,3-tetramethylguanidine (381 mg, 2.22 mmol, 1.50 equiv.) at room temperature. The reaction vial was sealed, and nitrogen gas was bubbled through the reaction mixture for 5 minutes. The stirred reaction mixture was then irradiated with a blue LED (365 nm) for 24 hours. LCMS showed the reaction was complete. Two parallel reactions were performed. The reaction was diluted with water (250 mL) and extracted with EA (75 mL × 3). The combined organic layers were washed with water (75 mL), brine (75 mL), dried over anhydrous sodium sulfate, and concentrated to dryness in vacuo. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), 0% to 60% gradient over 30 min; detector, UV 254 nm. The collected fractions were concentrated to give tert-butyl N-{[2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindol-4-yl]methyl}carbamate (compound 13-6, 300 mg, 22%) as a yellow solid. LCMS (ES, m / z): 388 [M+H] + , 288 [M+H-Boc] + , 329 [M+H-Boc+ACN] +

[0524] Step 4. Synthesis of compound 13-7 A solution of tert-butyl N-{[2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindol-4-yl]methyl}carbamate (compound 13-6, 300 mg, 0.77 mmol, 1.00 equiv.) in HCl (gas)·1,4-dioxane (15 mL) was stirred at room temperature overnight. LCMS showed the reaction was complete. The reaction was concentrated to dryness in vacuo to give 4-(aminomethyl)-2-(2,6-dioxopiperidin-3-yl)isoindole-1,3-dione (compound 13-7, 380 mg, crude) as a yellow solid. The crude product was used in the next step without further purification. LCMS (ES, m / z): 288 [M+H] +

[0525] Step 5. Synthesis of compound XIII 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexanoic acid (compound 13-4, 390 mg, 0.76 mmol, 1.00 equiv.) dissolved in DMF To a solution (5 mL) of 4-(aminomethyl)-2-(2,6-dioxopiperidin-3-yl)isoindole-1,3-dione (compound 13-7, 376 mg, 0.76 mmol, 1.00 equiv, 58%), HATU (577 mg, 1.52 mmol, 2.00 equiv), and DIEA (294 mg, 2.28 mmol, 3.00 equiv) were added in air at room temperature. The resulting mixture was stirred at room temperature for 2 hours. LCMS showed that the reaction was complete. The resulting mixture was diluted with water (50 mL) and extracted with EA (50 mL × 3). The combined organic layers were washed with brine (50 mL), water (50 mL), dried over anhydrous sodium sulfate, and concentrated to dryness in vacuo. The residue was purified by preparative TLC (DCM:MeOH=10:1) to give 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}-N-{[2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindol-4-yl]methyl}hexanamide (compound (XIII), 330 mg, 52%)) as a yellow solid. MS: (ES, m / s): 783,785 [M+H] + 1H NMR (400 MHz, DMSO-d6) δ11.14 (s, 1H), 8.45 (t, J= 5.6Hz, 1H), 8.18 (t, J=4.4Hz, 1H), 7.85-7.77 (m, 2H), 7.67 (d, J=7.6 Hz, 1H), 7.50-7.40 (dd, J=8.8 Hz, 20.8 Hz, 4H), 5.22-5.16 (m, 1H), 4.71-4.72 (m, 2H), 4.53-4.49 (m, 1H), 3.30-3.06 (m, 4H), 2.95-2.87 (m, 1H), 2.67-2.51 (m, 5H), 2.41 (s, 1H), 2.20 (t, J=7.2 Hz, 2H), 2.08-2.04 (m, 2H), 1.62 (s, 1H), 1.53-1.59 (m, 2H), 1.48-1.42 (m, 2H), 1.35-1.29 (m, 2H).

[0526] [ka] Step 1. Synthesis of compound 14-2 To a solution of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)-methyl acetate (compound 14-1, prepared according to the procedure described for compound 10-3, 100 mg, 0.44 mmol, 1.00 equiv.) in DCM (10 mL) was added TMSCl (70 mg, 0.66 mmol, 1.50 equiv.) at room temperature. The reaction was stirred at room temperature for 0.5 h. LCMS showed the reaction was complete. The reaction was concentrated to dryness in vacuo, and the residue was used directly in the next step. LCMS (ES, m / z): 203 [M+H] + (methanol-induced)

[0527] Step 2. Synthesis of compound 14-4 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}-N-{[2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindol-4-yl]methyl To a solution (5 mL) of {N-(chloromethyl)hexanamide (compound (XIII), 100 mg, 0.12 mmol, 1.00 equiv.) and prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 14-3, 106 mg, 0.52 mmol, 4.00 equiv.) in DMF, KCO (70 mg, 0.52 mmol, 4.00 equiv.) was added at room temperature. The resulting mixture was stirred at room temperature for 48 h. LCMS indicated the reaction was complete. The reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), gradient from 0% to 60% in 30 min; detector, UV 254 nm–220 nm. The collected fractions were concentrated to give prop-2-en-1-yl N-({[(3-{4-[(6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexane)methyl]-1,3-dioxoisoindol-2-yl}-2,6-dioxopiperidin-1-yl)methyl]carbamoyl}methyl)carbamate (compound 14-4, 110 mg, 83%) as a yellow solid. LCMS (ES, m / z): 953,955 [M+H] +

[0528] Step 3. Synthesis of compound 14-5 Propylene-2-en-1-yl N-({[(3-{4-[(6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexane)methyl]-1,3-dioxoisoin To a solution of {(2- ...3-(2-(2-(2-(2-(2-(2-(3-(2-(2-(2-(2-(2-(2-(3-(2-(2-(2-(2-(3-(2 LCMS (ES, m / z): 869,871 [M+H] +

[0529] Step 4. Synthesis of compound XIV N-[(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-1,3-dioxoisoindol-4-yl)methyl]-6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamido}hexanamide (compound 14-5, 50 mg, 0.06 mmol, 1.00 equiv.), [(2S To a solution of )-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetic acid (compound 14-4, prepared according to the procedure described for compounds 8-12, 34 mg, 0.06 mmol, 1.10 equiv.) in DMF (3 mL), HOBT (16 mg, 0.12 mmol, 2.00 equiv.), HATU (44 mg, 0.12 mmol, 2.00 equiv.), and DIEA (67 mg, 0.52 mmol, 9.00 equiv.) were added at room temperature. The resulting mixture was stirred overnight at room temperature. The resulting mixture was purified by column: XSelect CSH Prep C18 OBD column, 19 * 150 mm, 5 μm; Mobile phase A: water (0.05% FA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 29% B to 59% B in 7 min; Wavelength: 254 nm; RT1 (min): 6.8, purified with 6-{2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0^{2,6}]trideca-2(6),4,7,10,12-pentaene- 9-yl]acetamido}-N-[(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-1,3-dioxoisoindol-4-yl)methyl]hexanamide (compound (XIV), 14.4 mg, 17%) was obtained as a light yellow solid. LCMS (ES, m / z): 1380,1382 [M+H] + 1H NMR (400 MHz, DMSO-d6) δ8.47-8.44 (m, 1H), 8.29-8.21 (m, 2H), 8.19-8.15 (m, 1H), 8.13-8.04 (m, 2H), 8.00-7.92 (m, 2H), 7.89-7.72 (m, 2H), 7.68-7.66 (m, 1H), 7.50-7.41 (m, 4H), 7.35-7.23 (m, 4H), 7.20-7.11 (m, 1H), 6.99 (s, 2H), 5.25-5.12 (m, 2H), 5.08-5.00 (m, 1H), 4.72 (d, J=6Hz, 2H), 4.53-4.50 (m, 2H), 3.76-3.69 (m, 5H), 3.68-3.65 (m, 4H), 3.36 (t, J=7.2 Hz, 2H), 3.24-3.20 (m, 2H), 3.10-3.03 (m, 4H), 2.82-2.78 (m, 2H), 2.60 (s, 3H), 2.41 (s, 3H), 2.23-2.19 (m, 2H), 2.19-2.01 (m, 3H), 1.62 (s, 3H), 1.59-1.55 (m, 2H), 1.50-1.42 (m, 6H), 1.33-1.31 (m, 2H), 1.20–1.16 (m, 2H).

[0530]

change

[0531] Step 2. Synthesis of compound 15-4 To a stirred mixture of 6-[4-({4-[2-(2,6-dioxopiperidin-3-yl)-6-fluoro-1,3-dioxoisoindol-5-yl]piperazin-1-yl}methyl)piperidin-1-yl]-N-[(1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl]pyridazine-3-carboxamide (compound 15-3, 500 mg, 0.61 mmol, 1 equiv) in DMF (8 mL) was added NaH (37 mg, 0.92 mmol, 1.50 equiv, 60%) in portions at 0° C. under a nitrogen atmosphere. The resulting mixture was stirred at 0° C. for 30 minutes under a nitrogen atmosphere. To the above mixture was added prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 15-2, 254 mg, 1.23 mmol, 2 equiv.) at 0 °C. The resulting mixture was stirred at 25 °C for an additional 16 h. LCMS showed the reaction was complete. The reaction was quenched by adding water (1 mL) at room temperature. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), gradient of 5% to 50% ACN in 30 min; detector, UV 254 nm. This gave prop-2-en-1-yl N-{[({3-[5-fluoro-1,3-dioxo-6-(4-{[1-(6-{[(1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl]carbamoyl}pyridazin-3-yl)piperidin-4-yl]methyl}piperazin-1-yl)isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamate (compound 15-4, 130 mg, 21%) as a yellow solid. LCMS (ES, m / z): 982,984 [M+H] +

[0532] Step 3. Synthesis of compound 15-5 To a stirred mixture of prop-2-en-1-yl N-{[({3-[5-fluoro-1,3-dioxo-6-(4-{[1-(6-{[(1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl]carbamoyl}pyridazin-3-yl)piperidin-4-yl]methyl}piperazin-1-yl)isoindol-2-yl]-2,6-dioxopiperidin-1-yl}methyl)carbamoyl]methyl}carbamate (Compound 15-4, 120 mg, 0.12 mmol, 1 equiv.) and Pd(PPh3)4 (14 mg, 0.012 mmol, 0.1 equiv.) in THF (1.2 mL) was added phenylsilane (26 mg, 0.24 mmol, 2 equiv.) dropwise at room temperature under a nitrogen atmosphere. The resulting mixture was stirred under a nitrogen atmosphere at 25 °C for 3 hours. LCMS indicated the reaction was complete. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, water (0.05% TFA), gradient of 5% to 50% ACN over 30 minutes; detector, UV 254 nm. This afforded 6-(4-{[4-(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-6-fluoro-1,3-dioxoisoindol-5-yl)piperazin-1-yl]methyl}piperidin-1-yl)-N-[(1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl]pyridazine-3-carboxamide (compound 15-5, 40 mg, 36%) as a yellow solid. LCMS (ES, m / z): 899,901 [M+H] +

[0533] Step 4. Synthesis of compound XV To a stirred DMF solution (0.4 mL) of [(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetic acid (compound 15-6, prepared according to the procedure described for compounds 8-12, 21 mg, 0.040 mmol, 1 equiv.) and a mixture of HATU (18 mg, 0.048 mmol, 1.2 equiv.) and HOBT (7 mg, 0.048 mmol, 1.2 equiv.) was added 6-(4-{ [4-(2-{1-[(2-aminoacetamido)methyl]-2,6-dioxopiperidin-3-yl}-6-fluoro-1,3-dioxoisoindol-5-yl)piperazin-1-yl]methyl}piperidin-1-yl)-N-[(1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl]pyridazine-3-carboxamide (compound 15-5, 40 mg, 0.040 mmol, 1 equiv.) and DIEA (16 mg, 0.12 mmol, 3 equiv.) were added at 0° C. The resulting mixture was stirred under a nitrogen atmosphere at 25° C. for 16 hours. LCMS showed the reaction was complete. The crude product was purified by preparative HPLC using the following conditions: Column: XBridge Shield RP18 OBD column, 30 × 150 mm, 5 μm; Mobile phase A: water (0.1% FA), Mobile phase B: ACN; Flow rate: 60 mL / min; Gradient: 23% B to 43% B, 43% B in 10 min; Wavelength: 254 nm; RT1 (min): 8.92. The collected fractions were lyophilized to give 6-(4-{[4-(2-{1-[(2-{2-[(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetamido}acetamido)methyl]-2,6-dioxopiperidin-3-yl}-6-fluoro-1,3-dioxoisoindol-5-yl)piperazin-1-yl]methyl}piperidin-1-yl)-N-[(1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl]pyridazine-3-carboxamide (compound (XV), 7.2 mg, 10%)) as a yellow solid. LCMS (ES, m / z): 1409,1411 [M+H] + . 1H-NMR (DMSO-d6, 400 MHz) δ (ppm): 8.59 (d, J = 8.0 Hz, 1H), 8.28-8.24 (m, 2H), 8.13-8.06 (m, 2H), 7.99-7.92 (m, 2H), 7.86-7.73 (m, 3H), 7.39-7.31 (m, 3H), 7.24-7.21 (m, 4H), 7.17-7.12 (m, 2H), 6.99 (s, 2H), 5.17-5.14 (m, 3H), 4.53-4.47 (m, 4H), 3.74-3.50 (m, 11H), 3.37-3.59 (m, 2H), 3.30-3.22 (m, 3H), 3.07-2.76 (m, 7H), 2.62-2.52 (m, 3H), 2.32-2.00 (m, 7H), 1.91-1.83 (m, 5H), 1.65-1.62 (m, 2H), 1.52-1.43 (m, 6H), 1.20-1.14 (m, 4H).

[0534] [ka] Step 1. Synthesis of compound 17-7 To a solution of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl acetate (compound 17-6, 100 mg, 0.43 mmol, 1.00 equiv) in DCM (5 mL) was added TMSCl (71 mg, 0.65 mmol, 1.50 equiv) at room temperature. The reaction was stirred at room temperature for 0.5 h. LCMS showed the reaction was complete. The reaction was concentrated to dryness in vacuo, and the residue was used directly in the next step. LCMS (ES, m / z): 203 [M+H] + (methanol-induced)

[0535] Step 2. Synthesis of compound 17-8 A solution of 1-{3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl}-3-{[2-(2,6-dioxopiperidin-3-yl)-1-oxo-3H-isoindol-5-yl]methyl}urea (compound 9-6, 50 mg, 0.100 mmol, 1.00 equiv.), prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (compound 17-7, 82 mg, 0.40 mmol, 4.00 equiv.), and KCO (41 mg, 0.30 mmol, 3.00 equiv.) in NMP (2500 μL) was stirred at 50 °C for 24 h. Then, prop-2-en-1-yl N-[(chloromethylcarbamoyl)methyl]carbamate (82 mg, 0.40 mmol, 4.00 equiv.) and K2CO3 (14 mg, 0.10 mmol, 1.00 equiv.) in NMP (0.5 mL) were added. The resulting mixture was stirred at 50 °C for 24 h. LCMS showed the reaction was complete. The reaction was run eight times in parallel. The resulting mixture was purified by column: Sunfire preparative C18 OBD column, 19 * Purification by preparative HPLC using the following conditions: 250 mm, 10 μm; Mobile phase A: water (0.1% FA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 41% B to 57% B, 57% B in 10 min; Wavelength: 254 nm; RT 1 (min) gave the product prop-2-en-1-yl N-[({[3-(5-{[({3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl}carbamoyl)amino]methyl}-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl}carbamoyl)methyl]carbamate (compound 17-8, 64 mg, 11%) as a yellow solid. LCMS (ES, m / z): 671 [M+H] +

[0536] Step 3. Synthesis of compound 17-9 To a solution of prop-2-en-1-yl N-[({[3-(5-{[({3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl}carbamoyl)amino]methyl}-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl}carbamoyl)methyl]carbamate (compound 17-8, 64 mg, 0.10 mmol, 1.00 equiv) in THF (5 mL) and phenylsilane (21 mg, 0.19 mmol, 2.00 equiv) was added Pd(PPh3)4 (11 mg, 0.01 mmol, 0.10 equiv) under N2. The resulting mixture was stirred at room temperature for 3 hours. LCMS showed the reaction was complete. After filtration, the reaction mixture was purified by reverse-phase flash chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN in water (0.1% FA), 0% to 60% gradient in 30 min; detector, UV 254 nm. The collected fractions were lyophilized to give 2-amino-N-{[3-(5-{[({3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl}carbamoyl)amino]methyl}-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl}acetamide (compound 17-9, 32 mg, 51%) as an off-white solid. LCMS (ES, m / z): 587 [M+H] +

[0537] Step 4. Synthesis of compound XVII A solution of [(2S)-2-(2-{2-[6-(2,5-dioxopyrrol-1-yl)hexane]acetamido}acetamido)-3-phenylpropanamido]acetic acid (compound 17-9, 32 mg, 0.06 mmol, 1.20 equiv.), HOBT (7 mg, 0.05 mmol, 1.00 equiv.), and HATU (19 mg, 0.05 mmol, 1.00 equiv.) in DMF (3 mL) was stirred at room temperature for 1 hour. Then, 2-amino-N-{[3-(5-{[({3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl}carbamoyl)amino]methyl}-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl}acetamide (compound 17-10, prepared according to the procedure described for compound 8-12, 30 mg, 0.05 mmol, 1.00 equiv.) and DIEA (26 mg, 0.20 mmol, 4.00 equiv.) were added at room temperature. The resulting mixture was stirred at room temperature overnight. LCMS showed the reaction was complete. The reaction was purified by reverse-phase flash chromatography using the following conditions: Column: Kinetex EVO C18, 21.2 * 250 mm, 5 μm; Mobile phase A: water (0.1% FA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 34% B to 64% B, 64% B in 7 min; Wavelength: 254 nm; RT1 (min): 6. The collected fractions were lyophilized to give N-{[({[(1S)-1-[({[({[3-(5-{[({3-chloro-4-[2-(methylsulfanyl)ethyl]phenyl}carbamoyl)amino]methyl}-1-oxo-3H-isoindol-2-yl)-2,6-dioxopiperidin-1-yl]methyl}carbamoyl)methyl]carbamoyl}methyl)carbamoyl]-2-phenylethyl]carbamoyl}methyl)carbamoyl]methyl}-6-(2,5-dioxopyrrol-1-yl)hexanamide (compound (XVII), 6.5 mg, 10.57%) as a white solid. LCMS (ES, m / z): 1098 [M+H] + 1H NMR (400 MHz, DMSO-d6) δ 8.80 (s, 1H), 8.35-8.28 (m, 1H), 8.19-8.17 (m, 1H), 8.12-8.10 (m, 1H), 8.07-8.06 (m, 1H), 8.02-7.98 (m, 1H), 7.97-7.92 (m, 1H), 7.73-7.67 (m, 2H),7.53 (s, 1H), 7.47-7.45 (m, 1H), 7.24-7.22 (m, 5H), 7.20-7.15 (m, 2H), 7.00 (s, 2H), 6.84-6.80 (m, 1H), 5.22-4.96 (m, 3H), 4.55-4.50 (m, 1H), 4.44-4.41 (m, 2H), 4.34-4.28 (m, 1H), 3.73-3.66 (m, 7H), 3.37-3.36 (m, 4H), 3.10-2.95 (m, 2H), 2.90-2.70 (m, 4H), 2.69-2.62 (m, 3H), 2.15-2.00 (m, 5H), 2.09-2.00 (m, 1H), 1.50-1.45 (m, 4H), 1.21-1.19 (m, 2H).

[0538]

change

[0539] Step 2. Synthesis of compound 18-3 A mixture of (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)acetic acid (compound 18-2, 9 g, 41.62 mmol, 1.00 equiv.) and Cu(OAc) (0.76 g, 4.16 mmol, 0.1 equiv.) in THF (220 mL) was stirred at 60 °C for 1 h under a nitrogen atmosphere. The mixture was cooled to room temperature. To the above mixture, Pb(OAc) (22.15 g, 49.95 mmol, 1.2 equiv.) was added at room temperature. The resulting mixture was stirred at 25 °C for an additional 1 h. LCMS showed that the reaction was complete. The resulting mixture was filtered, and the filter cake was washed with MeOH (3 × 50 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (1:10) to give (2-{[(prop-2-en-1-yloxy)carbonyl]amino}acetamido)methyl acetate (compound 18-3, 5 g, 52%) as a white solid...

Claims

1. Formula (XXXII): 【Chemistry 175】 A complex of or a pharmaceutically acceptable salt thereof, wherein: a is 1 to 10; A' is 【176】 or 【Chemical 177】 where: n is 0 or 1; each Y is independently S or O; 【178】 is R １ indicates the point of attachment to 【179】 indicates the point of attachment to the methylene group, R １ is a compound that induces protein-protein interaction together with A', R ２ is hydrogen, R １ -A', a group that provides stability to R １ -A', a group that provides solubility to R １ - selected from groups that provide stability and solubility to A'; L is a cleavable linker; The conjugate or a pharmaceutically acceptable salt thereof, wherein Bm is a binding moiety capable of specifically binding to a protein.

2. Formula (XX): 【Chemistry 180】 A complex of or a pharmaceutically acceptable salt thereof, wherein: a is 1 to 10; n is 0 or 1; R １ is a compound that induces protein-protein interactions, R ２ is hydrogen, -(CH ２ CH ２ O) ｖ -CH ３ , C ２ -C ６ Alkenyl, C １ -C ６ Alkyl; C ２ -C ６ Alkynyl, benzyl, C ３ -C ６ Cycloalkyl, and C ３ -C ６ Cycloalkyl (C １ -C ３ alkyl), and v is 1 to 24; each Y is independently S or O; L is a cleavable linker; Bm is a binding moiety capable of specifically binding to a protein, preferably said binding moiety is an antibody, antibody fragment, or antigen-binding fragment, and preferably a is 2 to 8.

3. 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein the linker is cleavable by a protease.

4. L, 【Chemistry 181】 is selected from the group consisting of During the ceremony, q is 2 to 10; Z １ , Z ２ , Z ３ , Z ４ , and Z ５ are each independently absent or a naturally occurring amino acid residue in the L- or D-configuration, with the proviso that Z １ , Z ２ , Z ３ , Z ４ , and Z ５ are amino acid residues, 【Chemistry 182】 is the point of attachment to the parent molecular moiety, 【Chemistry 183】 is the point of attachment to the binding moiety, preferably Z 1 , Z 2 , Z 3 , Z 4 and Z 5 are independently absent or selected from the group consisting of L-valine, D-valine, L-citrulline, D-citrulline, L-alanine, D-alanine, L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-asparagine, D-asparagine, L-phenylalanine, D-phenylalanine, L-lysine, D-lysine and glycine, with the proviso that at least two of Z 1 , Z 2 , Z 3 , Z 4 and Z 5 are amino acid residues, and more preferably Z 1 is absent or glycine; Z 2 is absent or selected from the group consisting of L-glutamine, D-glutamine, L-glutamic acid, D-glutamic acid, L-aspartic acid, D-aspartic acid, L-alanine, D-alanine, and glycine; Z 3 is selected from the group consisting of L-valine, D-valine, L-alanine, D-alanine, L-phenylalanine, D-phenylalanine, and glycine; Z 4 is selected from the group consisting of L-citrulline, D-citrulline, L-asparagine, D-asparagine, L-lysine, D-lysine, L-phenylalanine, D-phenylalanine, and glycine; The conjugate of claim 3, or a pharmaceutically acceptable salt thereof, wherein Z 5 is absent or glycine.

5. L, 【Chemistry 184】 and preferably q is 4, or a pharmaceutically acceptable salt thereof.

6. L is a bioreducible linker, preferably L is 【Chemistry 185】 is selected from the group consisting of During the ceremony, q is 2 to 10; R, R', R'', and R''' are each independently selected from hydrogen, C 1 -C 6 alkoxyC 1 -C 6 alkyl, (C 1 -C 6 ) 2 NC 1 -C 6 alkyl, and C 1 -C 6 alkyl, or two geminal R groups together with the carbon atoms to which they are attached can form a cyclobutyl or cyclopropyl ring; 【Chemistry 186】 is the point of attachment to the parent molecular moiety, 【187】 is the point of attachment to the binding moiety, or a pharmaceutically acceptable salt thereof.

7. L, 【Chemistry 188】 and preferably q is 2, or a pharmaceutically acceptable salt thereof.

8. L is a click-to-release linker, preferably L is 【Chemistry 189】 and During the ceremony, q is 2 to 10; 【190】 is the point of attachment to the parent molecular moiety, 【Chemistry 191】 is the point of attachment to the binding moiety, or a pharmaceutically acceptable salt thereof.

9. L is a beta-glucuronidase cleavable linker, preferably L is 【Chemistry 192】 and During the ceremony, q is 2 to 10; --- is absent or a bond, 【Chemistry 193】 is the point of attachment to the parent molecular moiety, 【Chemistry 194】 is the point of attachment to the binding moiety, or a pharmaceutically acceptable salt thereof.

10. 2. The complex of claim 1, or a pharmaceutically acceptable salt thereof, wherein L is bound to a cysteine, lysine, tyrosine, or glutamine within the Bm, and preferably the cysteine ​​or lysine is an engineered cysteine ​​or lysine, or the cysteine ​​or lysine is endogenous to the Bm.

11. 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein Bm is an antibody or antigen-binding portion thereof, and preferably L is attached to an engineered cysteine ​​at position S239 and / or K334 of the heavy chain of the antibody or antigen-binding portion thereof according to EU numbering, or L is attached to the glutamine at position 295 of the heavy chain of the antibody or antigen-binding portion thereof according to EU numbering.

12. The protein to which the Bm binds is a surface antigen, and optionally, binding of the Bm to the surface antigen results in internalization of the complex or a pharmaceutically acceptable salt thereof into a cell, and preferably the surface antigen is selected from the group consisting of 5T4, ACE, ADRB3, AKAP-4, ALK, AOC3, APP, Axin1, AXL, B7H3, B7-H4, BCL2, BCMA, bcr-abl, BORIS, BST2, C242, C4.4a, CA1, 125, CA6, CA9, CAIX, CCL11, CCR5, CD123, CD133, CD138, CD142, CD15, CD15-3, CD171, CD17 9a, CD18, CD19, CD19-9, CD2, CD20, CD22, CD23, CD24, CD25, CD27L, CD28, CD3, CD30, CD31, C D300LF, CD33, CD352, CD37, CD38, CD4, CD40, CD41, CD44, CD44v6, CD5, CD51, CD52, CD54, CD 56, CD62E, CD62P, CD62L, CD70, CD71, CD72, CD74, CD79a, CD79b, CD80, CD90, CD97, CD125, C D138, CD141, CD147, CD152, CD154, CD326, CEA, CEACAM5, CFTR, clumping factor, cKit, claudin 3, claudin 18.2, CLDN6, CLEC12A, CLL-1, cll3, c-MET, Crypto1 growth factor, CS1, CTLA-4, CXCR2, CXORF 61, cyclin B1, CYP1B1, cadherin-3, cadherin-6, DLL3, E7, EDNRB, EFNA4, EGFR, EGFRvIII, ELF2M, EMR2, ENPP3, EPCAM, EphA2, ephrinA4, ephrinB2, EPHB4, ERBB2 (Her2 / neu), ErbB3, ERG (TMPRSS2ETS fusion gene), ETBR, ETV6-AML, FAP, FCAR, FCRL5, FGFR1, FGFR2, FGFR3, FGFR4, FLT3, folate receptor alpha, folate receptor beta, FOLR1, Fos-related antigen 1, fucosyl GM1, GCC, GD2, GD3, GloboH, GM3, GPC1, GPC2, GPC3, gplOO, GPNMB, GPR20, GPRC5D, GUCY2C, HAVCR1, HER2, HER3, HGF, HMI. 24, HMWMAA, HPV E6, hTERT, human telomerase reverse transcriptase, ICAM, ICOS-L, IFN-α, IFN-γ, IGF-I receptor, IGLL1, IL-2 receptor, IL-4 receptor, IL-13Ra2, IL-l lRa, IL-1 receptor, IL-12 receptor, IL-23 receptor, IL-13 receptor, IL-22 receptor, IL-4 receptor, IL-5 receptor, IL-6 receptor, interferon receptor, integrin (α 4 , α v β 3 , α v β 5 , α v β 6 , α 1 β 4 , α 4 β 1 , α 4 β 7 , α 5 β 1 , α 6 β 4 , α IIb β 3 Integrins (including integrins), integrin alpha V, intestinal carboxylesterase, KIT, LAGE-la, LAIR1, LAMP-1, LCK, legumain, LewisY, LFA-1 (CD11a), L-selectin (CD62L), LILRA2, LIV-1, LMP2, LRRC15, LY6E, LY6K, LY75, MAD-CT-1, MAD-CT-2, MAGE Al, MelanA / MARTl, mesothelin, ML-IAP, MSLN, mucin, MUC1, MUC16, mut hsp70-2, MYCN, myostatin, NA17, NaPi2b, NCA-90, NCAM, nectin-4, NGF, NOTCH1, NOTCH2, NOTCH3, NOTCH4, NY-BR-1, NY-ESO-1, o-acetyl-GD2, OR51E2, OY-TES1, p53, p53 mutant, PANX3, PAP, PAX3, PAX5, p-CAD, PCTA-1 / galectin 8, PD-L1, PD-L2, PDGFR, PDGFR-beta, phosphatidylserine, PIK3CA, PLAC1, polysialic acid, protease, prostate cancer cells, prostein, Pseudomonasaeruginosa, rabies, survivin and telomerase, PD-1, PRSS21, PSCA, PSMA, PTK7, RAGE-1, RANKL, Ras mutants, respiratory syncytial virus, rhesus factor, RhoC, RON, ROR1, ROR2, RU1, RU2, sarcoma translocation breakpoints, SART3, SLAMF7, SLC44A4, sLe, SLITRK6, sperm protein 17, sphingosine-1-phosphate, SSEA-4, SSX2, STEAP1, TAG72, TARP, TCRβ, TEM1 / CD248, TEM7R, tenascin-C, TF, TGF-1, TGF-β2, TNF-α, TGS5, Tie2, TIM-1, Tn Ag, TRAC, TRAIL-R1, TRAIL-R2, TROP-2, TRP-2, TRPV1, TSHR, tumor antigen CTAA16.88, tyrosinase, UPK2, VEGF, VEGFR1, VEGFR2, vimentin, WT1, XAGE1, or a combination thereof, and optionally the Bm that binds to CD33 comprises the amino acid sequence of SEQ ID NOs: 1 and 2, 3 and 4, 5 and 6, 22 and 23, 24 and 25, or 27 and 28, or the Bm that binds to PSMA comprises the amino acid sequence of SEQ ID NOs: 7 and 8, 9 and 10, 11 and 12, or 29 and 30, or the Bm that binds to HER2 The complex of claim 1, or a pharmaceutically acceptable salt thereof, wherein the Bm that binds to CD20 comprises the amino acid sequence of SEQ ID NOs: 13 and 14, 15 and 16, or 17 and 18, the Bm that binds to CD20 comprises the amino acid sequence of SEQ ID NOs: 31 and 32, the Bm that binds to CD79b comprises the amino acid sequence of SEQ ID NOs: 33 and 34, or the Bm that binds to BCMA comprises the amino acid sequence of SEQ ID NOs: 35 and 36, and more preferably the surface antigen comprises HER2, CD20, CD38, CD33, BCMA, CD138, EGFR, FGFR4, GD2, PDGFR, or a combination thereof.

13. The antibody is selected from the group consisting of rituximab, trastuzumab, gemtuzumab, pertuzumab, obinutuzumab, ofatumumab, daratumumab, STI-6129, lintuzumab, huMy9-6, belantamab, indatuximab, cetuximab, dinutuximab, anti-CD38 12. The conjugate of claim 11, or a pharmaceutically acceptable salt thereof, wherein the antibody is selected from the group consisting of A2 antibody, huAT15 / 3 antibody, alemtuzumab, ibritumomab, tositumomab, bevacizumab, panitumumab, tremelimumab, ticilimumab, catumaxomab, oregovomab, veltuzumab, polatuzumab, J591, lorvotuzumab and sacituzumab, preferably wherein the antibody is rituximab, trastuzumab, pertuzumab, huMy9-6, lintuzumab, gemtuzumab, or CD33-D.

14. R ２ is a group that provides stability to the conjugate, and preferably R 2 is selected from C 2 -C 6 alkenyl, C 1 -C 6 alkyl; C 2 -C 6 alkynyl, benzyl, C 3 -C 6 cycloalkyl, and C 3 -C 6 cycloalkyl (C 1 -C 3 alkyl), more preferably R 2 is C 1 -C 6 alkyl, and more preferably R 2 is methyl, or a pharmaceutically acceptable salt thereof.

15. R ２ is a group that provides solubility to the complex, and preferably R 2 is 【Chemistry 195】 is selected from During the ceremony, each n is independently 1, 2, 3, 4, or 5; each y is independently 1 or 2; 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein each R is independently hydrogen, C6H11O5, C12H21O10, C18H31O15, or C24H41O20.

16. 2. The conjugate of claim 1, wherein each Y is O, or a pharmaceutically acceptable salt thereof.

17. R １ 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein -A' is a PPI modulator, preferably a targeted protein degrader, more preferably a molecular glue, more preferably a substituted isoindoline, more preferably a 5'-substituted isoindoline.

18. R １ is represented by formula (XXX): 【Chemistry 196】 is a compound of During the ceremony, 【Chemistry 197】 indicates the point of attachment to the parent molecular moiety, A is phenyl or C ４ -C １０ is a cycloalkyl ring, R １０ is independently selected from hydrogen and halo; U is NH and CF ２ is selected from R ２０ is -CH ３ , -C(O)R ３ , -N(R ４ ) ２ , -(CH ２ ) ｎ OH, -(CH ２ ) ｎ N (R ４ ) ２ , -(CH ２ ) ｎ Q'(CH ２ ) ｍ OH, -(CH ２ ) ｎ Q'(CH ２ ) ｍ SH, and -(CH ２ ) ｎ Q'(CH ２ ) ｍ N (R ４ ) ２ is selected from: R ３ is hydrogen or C １ -C ６ is alkyl, Each R ４ are independently hydrogen or C １ -C ６ is alkyl, Q' is O, S, or NR ４ and n is 1 to 6; m is 2 to 5, preferably A is phenyl; U is NH; R 10 is halo; R 20 is methyl; or A is phenyl; U is NH; R 10 is halo; 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein R 20 is —(CH 2 ) 2 O(CH 2 ) 2 NHCH 3 .

19. R １ 2. The conjugate of claim 1, wherein -A' is a proteolysis-directed chimera (PROTAC), or a pharmaceutically acceptable salt thereof.

20. R １ But the formula: POI-L １００ -CBN wherein POI is a compound that binds to a protein of interest, L １００ is the PROTAC linker, 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein CBN is a cereblon-binding moiety.

21. The conjugate according to claim 20, or a pharmaceutically acceptable salt thereof, wherein the target protein is selected from a nuclear hormone receptor, a translation termination factor, a transcription factor, a cyclin-dependent kinase, a tyrosine kinase, a serine / threonine kinase, or an E3 ligase, or the target protein is selected from CD33, GSPT1, BRD4, AR, ER, IKZF1 / 3, CK1a, BCL-XL, IKZF2, IRAK4, BTK, STAT3, BTK and iMiD, BRD9, TRK, MDM2, CDK2 / CDK9, CD97b, and EGFR.

22. L １００ 21. The conjugate of claim 20, or a pharmaceutically acceptable salt thereof, wherein comprises one or more functional groups selected from glycol, alkyl, alkynyl, triazolyl, piperazinyl, piperidinyl, and combinations thereof.

23. CBN, 【Chemistry 198】 is selected from During the ceremony, 【Chemistry 199】 indicates the point of attachment to A', 【Chemistry 200】 Is, L １００ 21. The conjugate of claim 20, or a pharmaceutically acceptable salt thereof, showing a point of attachment to:

24. R １ but, 【Chemistry 201】 【Chemistry 202】 【Chemistry 203】 【Chemistry 204】 is selected from During the ceremony, 【Chemistry 205】 2. The conjugate of claim 1, or a pharmaceutically acceptable salt thereof, wherein: indicates the point of attachment to A'.

25. (a) the protein to which the Bm binds is CD33, and R １ (b) the protein to which Bm binds is prostate-specific membrane antigen (PSMA), and R １ binds to the androgen receptor (AR), or (c) the protein to which the Bm binds is CD33, and R １ binds to bromodomain-containing protein 4 (BRD4), or (d) the protein to which the Bm binds is HER2, and R １ binds to G1 to S Phase Transition 1 (GSPT1), or (e) the protein to which the Bm binds is CD33, and R １ binds to GSPT1, or (f) the protein to which the Bm binds is CD79b, and R １ binds to IRAK4, or (g) the protein to which the Bm binds is HER2, and R １ binds to BRD4, or (h) the protein to which the Bm binds is BCMA, and R １ binds to BRD4, or (i) the protein to which the Bm binds is HER2, and R １ The conjugate of claim 1 , or a pharmaceutically acceptable salt thereof, wherein

26. A pharmaceutical composition comprising the conjugate of claim 1 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers.

27. 27. A composition comprising the conjugate of any one of claims 1 to 25, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 26, for use in a method for treating cancer, wherein the cancer is preferably breast cancer, gastric cancer, lymphoma, acute myeloid leukemia, multiple myeloma, head and neck cancer, squamous cell carcinoma, hepatocellular carcinoma, prostate cancer, non-small cell lung cancer, colon cancer, ovarian cancer, neuregulin-1 (NRG1) positive cancer, lung cancer, or non-Hodgkin's lymphoma.

28. (a) the protein to which the Bm binds is CD33, and R １ (b) the protein to which the Bm binds is prostate-specific membrane antigen (PSMA), and R １ (c) the protein to which Bm binds is CD33, and R １ (d) the protein to which the Bm binds is HER2, and R １ (e) the protein to which Bm binds is CD79b, and R １ (f) the protein to which the Bm binds is HER2, and R １ binds to BRD4, and the cancer is breast cancer, gastric cancer, non-small cell lung cancer, cholangiocarcinoma, colon cancer, ovarian cancer, or neuregulin-1 (NRG1) positive cancer, or (g) the protein to which the Bm binds is BCMA, and R １ binds to BRD4 and the cancer is multiple myeloma.

29. The composition of claim 27, wherein the composition is for use in combination with an additional agent, wherein the additional agent is a cytotoxic agent or an immune response modifier, preferably wherein the immune response modifier is a checkpoint inhibitor, more preferably wherein the checkpoint inhibitor comprises a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a TIM3 inhibitor, and / or a LAG-3 inhibitor.

30. Formula (XXII): 【Chemistry 206】 The compound or a pharmaceutically acceptable salt thereof, wherein: n is 0 or 1; R １ is a compound that induces protein-protein interactions, R ２ is hydrogen, -(CH ２ CH ２ O) ｖ -CH ３ , C ２ -C ６ Alkenyl, C １ -C ６ Alkyl; C ２ -C ６ Alkynyl, benzyl, C ３ -C ６ Cycloalkyl, and C ３ -C ６ Cycloalkyl (C １ -C ３ alkyl), and v is 1 to 24; each Y is independently S or O; L ＊ is a cleavable linker precursor attached to the binding moiety, or a pharmaceutically acceptable salt thereof.

31. Formula (XXXI): 【Chemistry 207】 The compound or a pharmaceutically acceptable salt thereof, wherein: A' is 【Chemistry 208】 or 【Chemistry 209】 where: n is 0 or 1; each Y is independently S or O; 【Chemistry 210】 is R １ indicates the point of attachment to 【Chemistry 211】 indicates the point of attachment to the methylene group, R １ is a compound that induces protein-protein interaction together with A', R ２ is hydrogen, R １ -A', a group that provides stability to R １ -A', a group that provides solubility to R １ - selected from groups that provide stability and solubility to A'; wherein L is a cleavable linker precursor attached to the binding moiety, or a pharmaceutically acceptable salt thereof.

32. Formula (XXXII): 【Chemistry 235】 A complex of or a pharmaceutically acceptable salt thereof, a is 1 to 10; A' is 【Chemical 236】 or 【Chemical 237】 where: n is 0 or 1; each Y is independently S or O; 【Chemical 238】 is R １ indicates the point of attachment to 【Chemical 239】 indicates the point of attachment to the methylene group, R １ is a compound that induces protein-protein interaction together with A', R ２ is hydrogen, R １ -A', a group that provides stability to R １ -A', a group that provides solubility to R １ - selected from groups that provide stability and solubility to A'; L is a cleavable linker; Bm is a binding moiety capable of specifically binding to a protein, (XXXI) 【Chemistry 240】 The compound or a pharmaceutically acceptable salt thereof, A', R １ , and R ２ is as defined above, L ＊ is a cleavable linker precursor] The method further comprises reacting L* with a binding moiety capable of specifically binding to a protein, and preferably the method further comprises binding L* to a cysteine, lysine, tyrosine, or glutamine within the Bm, more preferably the cysteine ​​or lysine is an engineered cysteine ​​or lysine, or the cysteine ​​or lysine is endogenous to the Bm.

33. 33. The method of claim 32, wherein the binding moiety is an antibody or an antigen-binding portion thereof.

34. L ＊ The method of claim 32, wherein L* is linked to an engineered cysteine ​​at position S239 and / or K334 of the heavy chain of the antibody or its antigen-binding portion according to EU numbering, or L* is linked to a glutamine at position 295 of the heavy chain of the antibody or its antigen-binding portion according to EU numbering.

35. 33. The method of claim 32, wherein the binding is via site-specific binding.

36. (a) the protein to which the Bm binds is CD33, and R １ (b) the protein to which Bm binds is prostate-specific membrane antigen (PSMA), and R １ binds to the androgen receptor (AR), or (c) the protein to which the Bm binds is CD33, and R １ binds to bromodomain-containing protein 4 (BRD4), or (d) the protein to which the Bm binds is HER2, and R １ binds to G1 to S Phase Transition 1 (GSPT1), or (e) the protein to which the Bm binds is CD33, and R １ binds to GSPT1, or (f) the protein to which the Bm binds is CD79b, and R １ binds to IRAK4, or (g) the protein to which the Bm binds is HER2, and R １ binds to BRD4, or (h) the protein to which the Bm binds is BCMA, and R １ binds to BRD4, or (i) the protein to which the Bm binds is HER2, and R １ The method of claim 32, wherein the

37. A composition comprising the complex of any one of claims 1 to 25 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 26, for use in a method for delivering a complex that induces protein-protein interaction to a cell, the method comprising contacting the cell with the complex or the pharmaceutically acceptable salt thereof, or the composition.

38. Formula (XXXII): 【Chemical 249】 A complex of or a pharmaceutically acceptable salt thereof, a is 1 to 10; A' is 【Chemistry 250】 or 【Chemistry 251】 where: n is 0 or 1; each Y is independently S or O; 【Chemical 252】 is R １ indicates the point of attachment to 【Chemistry 253】 indicates the point of attachment to the methylene group, R １ is a compound that induces protein-protein interaction together with A', R ２ is hydrogen, R １ -A', a group that provides stability to R １ -A', a group that provides solubility to R １ - selected from groups that provide stability and solubility to A'; L is a cleavable linker; Bm is a binding moiety capable of specifically binding to a protein; The method comprises contacting the cell with the complex of formula (XXXII) or the pharmaceutically acceptable salt thereof.

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