Duocarmycin analogs and applications thereof

Abstract

The present disclosure relates to macrocyclic ketone compounds that are analogs of DNA-alkylating agent duocarmycin, compositions thereof and methods of making and using the same. The compounds may be used to treat a disease caused or characterized by undesired cell proliferation, such as a tumor.

Description

DUOCARMYCIN ANALOGS AND APPLICATIONS THEREOFCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 552,714, filed February 13, 2024, the entire content of which is incorporated herein by reference.BACKGROUND

[0002] Duocarmycins, including CC-1065, are antitumor antibiotics that may alkylate deoxynucleic acid (DNA) at the N3 position of adenine in the minor groove. Such alkylation may disrupt the nucleic acid architecture and initiate an apoptotic cell death mechanism, thereby becoming a cytotoxin. However, CC-1065 exhibits hepatotoxicity that outweighs its benefit as a cytotoxic drug. Hence, its synthetic analogs are studied to retain similar cytotoxicity but with reduced hepatotoxicity. Duocarmycins can be potent cancer cell killers and may act on both dividing and non-dividing cancer cells.SUMMARY

[0003] The present disclosure provides linker-drug compounds, which are synthetic analogues of duocarmycins, and derivatives thereof as mitotic DNA alkylating reagent, and compositions and applications thereof. These disclosed linker-drug compounds, and compositions and applications thereof, may effectively prevent or treat tumors or cancers.

[0004] In an aspect, provided herein is a compound of Formula I or Formula F:Formula I Formula I' or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:Rc is a reactive group;Linker is a cleavable linker or a non-cleavable linker;R2is H or CH3; andR3is H, OH, or OCH3.

[0005] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R1isR2is H or CH3; and R3is H, OH, or OCH3.

[0006] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is CH3; and R3is H. In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is CH3; and R3is OH. In some embodiments of the compound of Formula I d or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is CH3; and R3is OCH3.In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is H; and R3is H. In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is H; and R3is OH, In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is H; and R3is OCH3.

[0007] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R1is H,R2is CH3; and R3is H.

[0008] In some embodiments of the compound of Formula I or a pharmaceutically acceptable| _ LQ salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, Rc is *halide, nonaflate, triflate, fluorosulfonate, tosylate, mesylate, or besylate; RAis independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; RBis independently a protecting group, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each of R6, R7and R8is independently hydrogen, C1-C6 alkyl, cycloalkyl- alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxy carbonyl, or alkylsulfonyl; each R9is independently halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, C1-C4 heteroalkyl, -C(O)H, - C(O)OH, -CN, C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyd, C6-C10 aryl, 5- to 10- membered heteroaryl, -C(O)(C1-C4 alkyl), -C(O)O(C1-C4 alkyl), -C(O)NH2, -C(O)NH(C1-C4 alkyl), -C(O)N(C1-C4 alkyl)2, -NH2, -NH(C1-C4 alkyl), -N(C1-C4 alkyl)2, -NH(C2-C4alkylene)-OH, -NH(C2-C4alkylene)-O-(C1-C4 alkyl), -OH, -O(C1-C4 alkyl), -O(C1-C4 haloalkyl), -O(C2-C4alkylene)-NH2, -O(C2-C4alkylene)-NH-(C1-C4 alkyl), -O(C2-C4alkylene)-N-(C1-C4 alkyl)2, -O(C1-C4 alkylene)-C(O)OH, -O(C1-C4 alkylene)-C(O)O-(C1-C4 alkyl), - O(C2-C4 alkenyl), -O(C1-C4 alkylene)-( C6-C10 aryl), -O(C1-C4 alkylene)-(5- to 10- membered heteroaryl), _O(C6-C 10 aryl), SH, S(O)2OH, -S(O)2(C1-C4 alkyl), -S(O)2NH2, - S(O)2NH(C1-C4 alkyl), or -S(O)2N(C1-C4 alkyl)2; or two R9, together with atoms to which theyare attached, form a C3-C10 cycloalkyl or a 3- to 10-membered heterocycloalkyl ring; and each of p and q is independently an integer of 0-3.

[0009] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, Linker is — ( L2),— ( L1)s— *; * denotes a connection to Rc; each L1and L2is independently a bond, -O-, -S-, -NH-, - NRD- -C(=O)-, -C(=O)O-, -OC(=O)-, -C(=O)NH- -NHC(=O)-, -C(=O)NRD-, - NRDC(=O)-, -(CH2-O-CH2)m-, C1-C6 alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, Ci-Cg heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, - C6-C10 aryl-, or 5- to 10-membered heteroarylene, wherein each C1-C6 alkylene, Ci-Cg haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, -C6-C10 aryl-, and 5- to 10-membered heteroarylene is optionally substituted with 1, 2, or 3 R9; RDis independently -CR3R4R5; each R3, R4, and R5is independently hydrogen, halogen, -U, or -G; -U is independently C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl; wherein each C1-C6 alkyl, Ci- C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl is optionally substituted with 1, 2, or 3 R9and / or 1 or 2 -G; -G is independently C3-C10 cycloalkyl, 3- to 10- membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl; wherein each C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl is optionally substituted with 1, 2, or 3 R9; each R9is independently as defined above; and each of m, r and s is independently an integer of 1-12.

[0010] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, Linker comprisesthereof.

[0011] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, RAis independently as defined above; n is independently an integer of 1-8; and # denotes a connection to Rc or a group connected to Rc.

[0012] In some embodiments, the compound is according to Formulas II, II’, III, IIP, IV or IV’:R2is H or CH3;R3is H, OH, or OCH3;LG is halide, nonaflate, Inflate, fluorosulfonate, tosylate, mesylate, or besylate;RAis independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl- alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxy carbonyl, or alkylsulfonyl;RBis independently a protecting group, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each of R6, R7and R8is independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each R9is independently halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, C1-C4 heteroalkyl, -C(O)H, -C(O)OH, -CN, C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, 5- to 10-membered heteroaryl, - C(O)(C1-C4alkyl), -C(O)O(C1-C4 alkyl), -C(O)NH2, -C(O)NH(C1-C4 alkyl), -C(O)N(C1-C4 alkyl)2, -NH2, -NH(C1-C4 alkyl), -N(C1-C4 alkyl)2, -NH(C2-C4alkylene)-OH, -NH(C2-C4alkylene)-O-(C1-C4 alkyl), -OH, -O(C1-C4 alkyl), -O(C1-C4 haloalkyl), -O(C2-C4alkylene)- NH2, -O(C2-C4alkylene)-NH-(C1-C4 alkyl), -O(C2-C4alkylene)-N-(C1-C4 alkyl)2, -O(C1-C4 alkylene)-C(O)OH, -O(C1-C4 alkylene)-C(O)O-(C1-C4 alkyl), -O(C2-C4alkenyl), -O(C1-C4 alkylene)-(C6-C10 aryl), -O(C1-C4 alkylene)-(5- to 10-membered heteroaryl), 0(C6-C10 aryl), - SH, S(O)2OH, -S(O)2(C1-C4 alkyl), -S(O)2NH2, -S(O)2NH(C1-C4 alkyl), or -S(0)2N(C1-C4 alky 1)2; or two R9, together with atoms to which they are attached, form a C3-C10 cycloalkyl or a 3- to 10-membered heterocycloalkyl ring; each L1and L2is independently a bond, -O-, -S-, -NH-, -NRD-, -C(=O)-, -C(=O)O- -OC(=O)-, -C(=O)NH-, -NHC(=O)-, -C(=O)NRD-, -NRDC(=O)-, -(CH2-O-CH2)m-, C1-C6 alkylene, C1-C6 haloalkylene, - C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, -C6-C10 aryl-, or 5- to 10-membered heteroarylene, wherein each C1-C6 alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, - C6-C10 aryl-, and 5- to 10-membered heteroarylene is optionally substituted with 1, 2, or 3 R9;RDis independently -CR3R4R5; each R3, R4, and R5is independently hydrogen, halogen, -U, or -G;— U is independently C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2- C6 alkenyl, or C2-C6 alkynyl; wherein each C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl is optionally substituted with 1, 2, or 3 R9and / or 1 or 2 -G;-G is independently C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl; wherein each C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl is optionally substituted with 1, 2, or 3 R9; m is independently an integer of 1-12; each of p and q is independently an integer of 0-3; n is independently an integer of 0-8; and each t and u is independently an integer of 1-4.

[0013] In some embodiments, the compound is according to Formulas Ila, II’ a, Illa, Ill’a,, IVa or IV ’a:wherein:R1is independently as defined above;R2is H or CH3;R3is H, OH, or OCH3; each of RAand Rc is independently as defined above in all possible combinations;L1is independently as defined above; n is independently an integer of 0-8; and u is independently an integer of 1-4.

[0014] In some embodiments, the compound is according to Formulas lib, Il’b, Illb, Ill’b, IVb or IV’b:wherein:R1is independently as defined above;R2is H or CH3;R3is H, OH, or OCH3; each of RAand Rc is independently as defined above; m is independently an integer of 1-8; and n is an integer of 0-8.

[0015] In some embodiments, the compound is according to Formulas lie, II’c, IIIc, III’c, IVc or IV’c:Formula II’cFormula IVc , orFormula IV'c wherein:Rc is independently as defined above;R2is H or CH3;R3is H, OH, or OCH3; x is an integer of 0-4; and y is independently an integer of 1-8.

[0016] In some embodiments of the compound of Formulas disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is CH3; and R3is H. In some embodiments of the compound of Formulas disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is CH3; and R3is OH. In some embodiments of the compound of Formulas disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is CH3; and R3is OCH3. In some embodiments of the compound of Formulas disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is H; and R3is H. In some embodiments of the compound of Formulas disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is H; and R3is OH. In some embodiments of the compound of Formulas disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, R2is H; and R3is OCH3.

[0017] In some embodiments, the compound is:wherein:R1is as defined above; andRc is as defined in above.

[0018] In some embodiments of the compound of Formulas disclosed herein or any compound disclosed above, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, wherein R1is:

[0019] In some embodiments of the compound of Formula I or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, wherein the compound is:wherein:Rc is as defined above.

[0020] In some embodiments of the compound of Formulas disclosed herein or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, wherein Rc is:

[0021] In some embodiments of the compound of Formulas disclosed herein or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof disclosed herein, wherein Rc is:

[0022] In another aspect, provided herein is a pharmaceutical composition compnsing a compound disclosed herein, or pharmaceutically acceptable salt, solvate, diastereomeric mixture, or individual enantiomers thereof, and a pharmaceutically acceptable carrier.

[0023] In still another aspect, provided herein is a method of treating a cancer in a mammal suffering therefrom, comprising administering to the mammal a therapeutically effective amount of a compound disclosed herein or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, or a pharmaceutical composition disclosed herein.

[0024] In another aspect, provided herein is a method of treating a cancer in a mammal suffering therefrom with a conjugate of a compound disclosed herein linked with (i) a cell surface targeting agent, or (ii) a long-acting reagent, comprising: administering to the mammal a therapeutically effective amount of (i) the conjugate of the compound disclosed herein or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, or (ii) a pharmaceutical composition comprising the conjugate of the compound disclosed herein, or pharmaceutically acceptable salt, solvate, diastereomeric mixture, or individual enantiomers thereof, and a pharmaceutically acceptable carrier, wherein the cell surface targeting agent is a small molecule, a polypeptide, or an antibody, and wherein the long-acting reagent comprising a polymer, human serum albumin (HSA), or a nanoparticle.

[0025] In another aspect, provided herein is a conjugate comprising: (i) a compound disclosed herein or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof; and (ii) a cell surface targeting agent or a long-acting reagent linked to the compound, wherein the cell surface targeting agent is a small molecule, a polypeptide, or an antibody, and wherein the long-acting reagent comprising a polymer, human serum albumin (HSA), or a nanoparticle.

[0026] In another aspect, provided herein is a conjugate comprising a cell surface targeting agent or a long-acting reagent, wherein the cell surface targeting agent or the long-acting reagent is attached to a compound disclosed herein or a derivative of a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the cell surface targeting agent is a small molecule, a polypeptide, or an antibody or an antigen- binding portion thereof, and wherein the long-acting reagent comprising a polymer, human serum albumin (HSA), or a nanoparticle.

[0027] In some embodiments of the conjugate, the cell surface targeting agent is a monoclonal antibody, a Fab, a Fab’, a F(ab’), a Fv, a disulfide linked Fc, a scFv, a single domain antibody, adiabody, a bi-specific antibody, or a multi-specific antibody. In some embodiments of the conjugate, the cell surface targeting agent binds to a target molecule. In some embodiments of the conjugate, the target molecule comprises a tumor suppressor, a metabolic enzyme, a protein aggregate, or a haploinsufficient protein. In some embodiments of the conjugate, the target molecule comprises p53 mutant or Von Hippel-Lindau tumor suppressor (VHL). Disclosed herein is a pharmaceutical composition comprising a conjugate disclosed herein, and a pharmaceutically acceptable carrier. Disclosed herein is a method of treating a cancer in a mammal suffering therefrom, comprising administering to the mammal a therapeutically effective amount of a conjugate of disclosed herein, or a pharmaceutical composition disclosed herein, wherein the conjugate binds to a target antigen associated with the cancer.

[0028] Additional aspects and advantages of the present disclosure will become readily apparent to those skilled in this art from the following detailed description, wherein only illustrative instances of the present disclosure are shown and described. As will be realized, the present disclosure is capable of other and different instances, and its several details are capable of modifications in various obvious respects, all without departing from the disclosure.Accordingly, the drawings and description are to be regarded as illustrative in nature, and not as restrictive.INCORPORATION BY REFERENCE

[0029] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. To the extent publications and patents or patent applications incorporated by reference contradict the disclosure contained in the specification, the specification is intended to supersede and / or take precedence over any such contradictory material.BRIEF DESCRIPTION OF THE DRAWINGS

[0030] The novel features of the present disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the present disclosure are utilized, and the accompanying drawings (also “Figure” and “FIG.” herein) of which:

[0031] FIG. 1 shows cell viability data for example compounds to treat a human prostate cancer cell line DU 145.

[0032] FIG. 2 shows cell viability data for example compounds to treat a human prostate cancer cell line DU145 with or without a pretreatment with P-D-galactosidase.

[0033] FIG. 3 shows a possible mechanism of prodrug transformation when an example prodrug is conjugated to a targeting moiety.DETAILED DESCRIPTION

[0034] While various embodiments of the present disclosure have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions may occur to those skilled in the art without departing from the present disclosure. It should be understood that various alternatives to the embodiments of the present disclosure described herein may be employed.

[0035] Compounds are generally described herein using standard nomenclature. For compounds having asymmetric centers, it should be understood that (unless otherwise specified) all of the optical isomers and mixtures thereof are encompassed. In addition, compounds with carbon- carbon double bonds may occur in Z- and E- forms, with all isomeric forms of the compounds being included in the present invention unless otherwise specified. Where a compound exists in various tautomeric forms, a recited compound is not limited to any one specific tautomer, but rather is intended to encompass all tautomeric forms.Definitions

[0036] As used herein, the singular forms “a,” “an,” and “the” include the plural reference unless the context clearly dictates otherwise.

[0037] When a range of values is provided, it is to be understood that each intervening value between the upper and lower limit of that range, and any other stated or intervening value in that stated range is encompassed within the scope of the present disclosure. Where the stated range includes upper or lower limits, ranges excluding either of those included limits are also included in the present disclosure. When ranges are used herein for physical properties, such as molecular weight, or chemical properties, such as chemical formulas, all combinations and subcombinations of ranges and specific embodiments therein are intended to be included.

[0038] As used herein, the term “about” or “nearly” when referring to a number or a numerical range means that the number or numerical range generally referred to is within + / - 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of the stated number or numerical range.

[0039] As used herein, the term “comprising” (and related terms such as “comprise” or “comprises” or “having” or “including”) is not intended to exclude that in other certain embodiments, for example, an embodiment of any composition of matter, composition, method, or process, or the like, described herein.

[0040] As used in the specification and appended claims, unless specified to the contrary, the following terms have the meaning indicated below.

[0041] As used herein, the term “C1-C6 alkyl” generally refers to a straight or branched hydrocarbon chain having from 1 to 6 carbon atoms, and the straight or branched hydrocarbon chain is attached to the rest of the molecule by a single bond. Likewise, an alkyl group comprising up to 3 carbon atoms is a C1-C3 alkyl group, and an alkyl group comprising up to 4 carbon atoms is a C1-C4 alkyl group. Examples of a C1-C6 alkyl group include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, s-butyl, tert-butyl, n-pentyl, isopentyl, 2-methylbutyl, neopentyl, 1 -ethylpropyl, n-hexyl, isohexyl, 4-methylpentyl, 3- methylpentyl, 2-methylpentyl, 1 -methylpentyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1- dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,3 -dimethylbutyl, and 2-ethylbutyl. In some instances, a substituent of an alkyl group is specifically indicated. For example, “cyanoalkyl” refers to an alkyd group substituted with at least one cyano substituent.

[0042] The C1-C6 alkyl group may be optionally substituted with a C1-C3 alkoxy group. Examples include, but are not limited to, methoxyethyl, methoxypropyl, methoxyisopropyl, ethoxyethyl, ethoxypropyl, ethoxyisopropyl, propoxyethyl, propoxypropyl, and propoxyisopropyl.

[0043] The C1-C6 alkyl group may be optionally substituted with a C3-C6 cycloalkyl group. Examples include, but are not limited to, 1 -methylcyclopropyl, 1 -methylcyclobutyl, and 1- methylcyclohexyl.

[0044] As used herein, the term “C1-C6 alkoxy” generally refers to a radical of the formula -OR wherein R is a C1-C6 alkyl group as defined. Likewise, an alkoxy group comprising up to 3 carbon atoms is a C1-C3 alkoxy group. Examples include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, s-butoxy, tert-butoxy, n-pentoxy, isopentoxy, 2-methylbutoxy, neopentoxy, 1 -ethylpropoxy, n-hexyloxy, isohexyloxy, 4- methylpentoxy, 3 -methylpentoxy, 2-methylpentoxy, 1 -methylpentoxy, 3.3-dimethylbutoxy, 2,2- dimethylbutoxy, 1,1 -dimethylbutoxy, 1,2-dimethylbutoxy, 1.3 -dimethylbutoxy, 2,3- dimethylbutoxy, and 2-ethylbutoxy.

[0045] The C1-C3 alkoxy group may be optionally substituted with a C1-C3 alkoxy group. Examples include, but are not limited to, methoxy methoxy, methoxy ethoxy, methoxypropoxy, methoxyisopropoxy, ethoxymethoxy, ethoxyethoxy, ethoxypropoxy, ethoxyisopropoxy, propoxymethoxy, propoxyethoxy, propoxypropoxy, and propoxy isopropoxy.

[0046] As used herein, the term “C3-C6 cycloalkylamino” is, for example, azacyclobutyl, pyrrolidine, pipendino, or hexamethylenimino.

[0047] As used herein, the term “C3-C6 cycloalkyl” generally refers to a monocyclic non- aromatic radical having from 3 to 6 ring atoms, wherein each of the atoms forming the ring (i.e. skeletal atoms) is a carbon atom. Examples include, but are not limited to, cyclopropyl,cyclobutyl, cyclopentyl, and cyclohexyl. The term “cycloalkenyl” as used herein generally refers to a group that comprises one or more unsaturated rings in which all ring members are carbon. Polycyclic cycloalkyls or carbocycles include, for example, adamantyl, norbomyl, decalinyl, bicyclo[3.3.0]octane, bicyclo[4.3.0]nonane, cis-decalin, trans-decalin, bicyclo[2.1.1]hexane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2.2]nonane, and bicyclo [3.3.2] decane, and 7,7-dimethyl-bicyclo[2.2.1]heptanyl. Unless stated otherwise specifically in the specification, a cycloalkyl is optionally substitutedo

[0048] As used herein, the term “alkenyl” generally refers to straight or branched chain alkene groups, which comprise at least one unsaturated carbon-carbon double bond. Alkenyl groups include C2-s alkenyl, C2-6 alkenyl and CM alkenyl groups, which have from 2 to 8, 2 to 6, or 2 to 4 carbon atoms, respectively, including, for example, ethenyl, allyl or isopropenyl. The term “alkynyl” as used herein generally refers to straight or branched chain alkyne groups, which have one or more unsaturated carbon-carbon bonds, at least one of which is a triple bond. Alkynyl groups include C2-8 alkynyl, C2-6 alkynyl and C2-4 alkynyl groups, which have from 2 to 8, 2 to 6 or 2 to 4 carbon atoms, respectively.

[0049] As used herein, the term “halogen” or “halide” generally refers to fluorine, chlorine, bromine, and iodine. The term “haloalkyl” as used herein generally refers to an alkyl group that is substituted with one or more independently chosen halogens (e.g., “C1-C6 haloalkyl” groups have from 1 to 6 carbon atoms and at least one halogen). Examples of haloalkyl groups include, but are not limited to, mono-, di- or tri -fluoromethyl; mono-, di- or tri-chloromethyl; mono-, di-, tri-, tetra- or penta-fluoroethyl; mono-, di-, tri-, tetra- or penta-chloroethyl; 2,2,2-trifluoroethyl; 1,2-difluoroethyl; 3-bromo-2-fluoropropyl; 1,2-dibromoethyl; and 1,2,2,2-tetrafluoro-l- trifluoromethyl-ethyl.

[0050] As used herein, the term “heteroalkyl” generally refers to an alkyl group in which one or more skeletal atoms of the alkyl are selected from an atom other than carbon, e.g., oxygen, nitrogen (e.g., -NH-, -N(alkyl)-), sulfur, or combinations thereof. In some instances, a heteroalkyl is attached to the rest of the molecule at a carbon atom of the heteroalkyl. In one aspect, a heteroalkyl is a C1-C6 heteroalkyl wherein the heteroalkyl is comprised of 1 to 6 carbon atoms and one or more atoms other than carbon, e.g., oxygen, nitrogen, sulfur, or combinations thereof. In some instances, a carbon atom or heteroatom is optionally oxidized (e.g., - C(O)OCH2-, -CH2OCH2-, -CH2S(O)2NHCH2-, -NHC(O)NHCH2-, -CH2NHC(O)CH2-). Further examples of such heteroalkyl are, for example, -CFbOCFE, -CH2CH2OCH3, - CH2CH2OCH2CH2OCH3, or -CH(CH3)OCH3. Unless stated otherwise specifically in the specification, a heteroalkyl is optionally substituted for example, with oxo, halogen, amino, nitrile, nitro, hydroxy, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl,heterocy cloalkyl, heteroaryl, and the like. In some embodiments, a heteroalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a heteroalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, - CF3, -OH, or -OMe.

[0051] As used herein, the term “heteroaryl” generally refers to a monocyclic aryl group that includes one or more ring heteroatoms selected from nitrogen, oxy gen and sulfur. Examples include, but are not limited to, pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, pyridazinyl, triazinyl, oxadiazolyl, thiadiazolyl, and furazyl.

[0052] The terms “heterocyclic” or “heterocycle” or “heterocyclyl” or “cycloheteroalkyl” or “heterocycloalkyl” as used herein generally refer to a ring structure (monocycle or poly cycle) containing 3-12 ring atoms (3-12 membered heterocycle), 3-8 ring atoms (3-8 membered heterocy cle or 3-8 membered cycloheteroalkyl), 3-6 ring atoms (3-6 membered heterocycle or 3- 6 membered cycloheteroalkyl), or 5-6 ring atoms (5-6 membered heterocycle or 5-6 membered cycloheteroalkyl), in which at least one ring atom is carbon, and at least one ring atom is a heteroatom selected from N, 0, and S, or a heteroatom group selected from C(=O), S(=O), and S(=O)2. A heterocyclic group may be aromatic or non-aromatic. Piperidine and oxetane are non- limiting examples of non-aromatic heterocycles. Thiazole and pyridine are non-limiting examples of aromatic heterocycles. Other examples of heterocycle include: azindinyl, azetidinyl, oxetanyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothienyl, piperidinyl, morpholinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, thiomorpholinyl, tetrahydropyranyl, 1,1-dioxothiomorpholinyl, butyrolactam, valerolactam, caprolactam, butyrolactone, valerolactone and caprolactone. Similarly, the term “cycloheteroalkenyl” refers to a monocycle or poly cycle ring structure comprising carbon atom(s) and heteroatom(s) / heteroatom group(s), wherein the cycloheteroalkenyl comprises at least one C=C double bond, at least one ring atom that is carbon, and at least one ring atom that is a heteroatom selected from N, 0, and S or a heteroatom group selected from C(=O), S(=O), and S(=O)2. Unless stated otherwise specifically in the specification, a heterocycle or heterocy cloalkyl is optionally substituted, for example, with oxo, halogen, amino, nitrile, nitro, hydroxy, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, and the like. In some embodiments, a heterocycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a heterocy cloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or - OMe.

[0053] As used herein, the term “aryl” generally refers to an all-carbon monocyclic or fused- ring polycyclic groups of 6 to 12 (Ce-12 aryl) or 6 to 10 carbon atoms (Ce-io aryl) having a completely conjugated pi-electron system. Examples include, but are not limited to, phenyl, naphthalenyl, tetrahydronaphthyl, indanyl, biphenyl, and anthracenyl. The aryl group may be substituted or unsubstituted. Typical substituents include halo, trihalomethyl, alkyl, hydroxy, alkoxy, aryloxy, mercapto, alkylthio, arylthio, cyano, nitro, carbonyl, thiocarbonyl, C-carboxy, O-carboxy, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, sulfinyl, sulfonyl, amino and -NRXRY, wherein Rxand RYare independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl, acetyl, sulfonyl, trifluoromethanesulfonyl and, combined, a five- or six-membered heteroalicyclic ring. Illustrative substituted alkyl group include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, aminomethyl, aminoethyl, hydoxymethyl, methoxymethyl, 2-fluoroethyl, and 2- methoxyethyl, etc.

[0054] The term “heteroaryl” as used herein generally refers to an aromatic group in which at least one aromatic ring comprises at least one heteroatom selected from N, O and S. Heteroaryls include, for example, 5-12 membered heteroaryls, 5-10 membered heteroaryls, 5-7 membered monocyclic structures or 7-12 membered bicyclic structures. The number of heteroatoms in a heteroaryl can be 1, 2, 3, 4, or more. Examples include, but are not limited to, thienyl, pyridyl, pyrimidinyl, pyrazinyl, pyrazolyl, pyridazinyl, pyridine-2(lH)-keto, pyridine-4(lH)-keto, pyrrolyl, pyrazolyl, thiazolyl, 1,2 ,3-triazolyl, 1,2,4-triazolyl, 1,2,5-oxadiazolyl, imidazolyl, furanyl, tetrazolyl, isothiazolyl, oxazolyl, isoxazolyl, thiadiazolyl, oxadiazolyl, naphthyl ,benzothienyl, indolyl, benzimidazolyl, benzothiazolyl, benzofuranyl, quinolinyl, isoquinolinyl, and quinazolinyl. The heteroary l group may be substituted or unsubstituted. Typical substituents include halo, trihalomethyl, alkyl, hydroxy, alkoxy, aryloxy, mercapto, alkylthio, arylthio, cyano, nitro, carbonyl, thiocarbonyl, C-carboxy, O-carboxy, O-carbamyl, N-carbamyl, O- thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, sulfinyl, sulfonyl, amino and -NRXRY, with Rxand RYas defined above.

[0055] As used herein, the term “amino” generally refers to primary amino group (-NH2), / — N secondary amino group (-NH-), and tertiary ammo group ( \ ).

[0056] As used herein, the term “alkylamino” generally refers to a secondary or tertiary amine that has the general structure -NH-R1or -NfR1)(R2), respectively, wherein R1and R2are selected independently from alkyl, cycloalkyl and (cycloalkyl)alkyl groups. Such alkylamino groups include, but are not limited to, mono- and di-(Ci-6 alkyl)amino groups, in which each C1-6 alkyl may be the same or different. In this case, the definition of “alkyl” as used in the term“alkylamino” differs from the definition of “alkyl” used for all other alkyl-containing groups, in the inclusion of cycloalkyl and (cycloalkyl)alkyl groups.

[0057] The term “alkylthio” as used herein generally refers to an alkyl-substituted thio group, wherein the term alkyl is as defined above o

[0058] The terms “substituent” and “substituted,” as used herein, generally denote that a molecular moiety is covalently bonded to an atom within a molecule of interest. For example, a ring substituent may be a moiety such as a halogen, alkyl group, haloalkyl group or other group that is covalently bonded to an atom (preferably a carbon or nitrogen atom) that is a ring member. Substituents of aromatic groups are generally covalently bonded to a ring carbon atom. A straight chain substituent may be a moiety such as a halogen, alkyl group, haloalkyl group or other group that is covalently bonded to an atom (preferably a carbon or nitrogen atom) that is a member of a straight chain.

[0059] The term “cycloalkylamine” as used herein generally refers to either a ring structure with an ammo group attached to a carbon atom in the ring or a ring structure with a nitrogen atom as member of the ring.

[0060] As used herein, the term “C1-C4 alkylcarbonyl” generally refers to a carbonyl radical that is substituted by a C1-C4 alkyl radical as defined above. Examples include, but are not limited to, methylcarbonyl, ethylcarbonyl, n-propylcarbonyl, isopropylcarbonyl, buty lcarbonyl, and tert- butylcarbonyl.

[0061] As used herein, the term “C1-C3 alkylsulfonyl” generally refers to a sulfonyl radical that is substituted by a C1-C3 alkyl radical as defined above. Examples include, but are not limited to, methanesulfonyl, ethanesulfonyl, n-propanesulfonyl, and isopropanesulfonyl.

[0062] As used herein, the term “C1-C4 alkoxy carbonyl” generally refers to a carbonyl radical that is substituted by C1-C4 alkoxy radical, as defined above. Examples thereof include methoxycarbonyl, ethoxycarbonyl, n-propoxycarbonyl, isopropoxycarbonyl, butoxycarbonyl, and tert-butoxy carbonyl.

[0063] As used herein, the term “leaving group” generally refers to molecular fragment or stable species that can be detached from a molecule in a bond-breaking step. The leaving group, in accordance with the specification, is not particularly limited. The ability of a leaving group to depart is correlated with the pKa of the conjugate acid, with lower pKa being associated with better leaving group ability. Examples of leaving group include, without limitation, halide or a sulfonate. Halides is as defined above. Examples of sulfonates can include, without limitation, nonaflate, triflate, fluorosulfonate, tosylate, mesylate or besylate. In one embodiment, for example and without limitation, the leaving group is chloride, mesylate or tosylate. The functional groups that can be converted into leaving groups, in accordance with thespecification, are not particularly limited. In one embodiment, for example the functional group can be a hydroxy group that can be converted into a leaving group as described above.

[0064] As used herein, the term “linker” generally refers to a molecule that joins two other molecules, either covalently, or through ionic, van der Waals or hydrogen bonds. In some cases, the linker uses covalent bonds to join the two other molecules. The term “cleavable linker” as used herein generally refers to a linker that can be degraded or otherwise severed to separate the two components connected by the cleavable linker. Cleavable linkers are generally cleaved by enzy mes, typically peptidases, proteases, nucleases, lipases, and the like. Cleavable linkers may also be cleaved by environmental cues, such as, for example, changes in temperature, pH, salt concentration, etc., when there is such a change in environment following transcytosis of the compound disclosed herein across a polarized epithelial membrane.

[0065] As used herein, the term “conjugate” generally refers to a compounds disclosed herein linked to a cell surface targeting agent via a linker. In some cases, the linker can be a cleavable linker. In some cases, the linker can be a non-cleavable linker. In some embodiments, a drug- linker-Rc intermediate can react with a targeting moiety via the Rc (reactive group) on the drug- linker-Rc and form a covalent bond between Rc (or a derivative of the reactive group (Rc) after the conjugation reaction) and the targeting moiety. In the above context, Rc comprises both the reactive group before the conjugation reaction (as in the drug-linker-Rc) and the derivative of the reactive group after the reaction (as in the drug-linker-Rc-targeting moiety).

[0066] As used herein, the term “cell surface targeting agent” generally refers to an agent that binds to a cell surface, in particular, selectively binds to a particular cell surface, for example, by targeting a specific cell surface receptor or a unique cell surface motif. In some cases, the cell surface targeting agent can be a receptor binding domain. For example, the receptor binding domain can be any receptor binding domain known to one of skill in the art without limitation to bind to a cell surface receptor that is present on the apical membrane of an epithelial cell. In some cases, the receptor binding domain can bind specifically to the cell surface receptor. In some cases, the receptor binding domain can bind to the cell surface receptor with sufficient affinity to allow endocytosis of the conjugate. In some cases, the cell surface targeting agent can bind selectively to the surface of targeted cells. For example, the cell surface targeting agent may be a ligand that binds to the cell surface receptor found on a particular type of cell or expressed at a higher frequency on target cells than on other cells.

[0067] In some cases, the “cell surface targeting agent” can comprise a peptide, a polypeptide, a protein, a lipid, a carbohydrate, or a small organic molecule, or a combination thereof. In some cases, examples of each of these molecules can bind to cell surface receptors present on the apical membrane of epithelial cells. Examples of peptides or polypeptides include, but are notlimited to, RGD-containing peptides, bombesin or gastrin-releasing peptide, bacterial toxin receptor binding domains, such as the receptor binding domains from Pseudomonas aeruginosa (PE), cholera toxin, Cholix toxin, botulinum toxin, diptheria toxin, shiga toxin, shiga-like toxin, etc.; fusion proteins (e.g., albumin fusions); antibodies, including monoclonal, polyclonal, and single-chain antibodies, or derivatives thereof (e.g., isotype immunoglobulin G (IgG) or derivatives thereof, or integrin alpha-10 specific antibody), grow th factors, such as EGF, IGF-I, IGF-II, IGF-III etc.; cytokines, such as IL-1, IL-2, IL-3, IL-6, etc.; chemokines, such as MIP-la, MIP-lb, MCAF, IL-8, etc.; and other ligands, such as CD4, cell adhesion molecules from the immunoglobulin superfamily, integrins, ligands specific for the IgA receptor, etc.

[0068] Similarly, as used herein, the term ‘‘targeting moiety” or “targeting agent” generally refers to any moiety that specifically binds or reactively associates or complexes with a moiety specifically or in relative excess present at or near the target site, on, in, or near the target cell, or in (the proximity of) the target tissue or organ, e.g., a receptor, a receptor complex, substrate, antigenic determinant, or other receptive moiety, or that can target the conjugate to the target site via other mechanisms by virtue of its nature, e.g., through the EPR effect. Examples of a targeting moiety include, but are not limited to, a cell surface targeting agent, an aptamer, an antibody or antibody fragment or derivative, a peptide, a polymer, a dendrimer, a lectin, a biologic response modifier, an enzy me, a vitamin, a growth factor, a steroid, a sugar residue, an oligosaccharide residue, a carrier protein, and a hormone, or any combination thereof. The targeting moiety' or targeting agent can bind to a target molecule.

[0069] As used herein, the term “target molecule” when used in association with a targeting moiety generally refers to the binding target for the targeting moiety, including, but not limited to, a cancer-associated antigen such as CD19, CD20, CD30, CD33, CD38, CA125, MUC-1, prostate-specific membrane antigen (PSMA), CD44 surface adhesion molecule, mesothelin (MLSN), carcinoembryonic antigen (CEA), epidermal growth factor receptor (EGFR), EGFRvIII, vascular endothelial growth factor receptor-2 (VEGFR2), HER2, high molecular weight-melanoma associated antigen (HMW-MAA), MAGE-A1, IL- 13R-a2, GD2, lpl9q, ABL1, AKT1, ALK, APC, AR, ATM, BRAF, BRCA1, BRCA2, cKIT, cMET, CSF1R, CTNNB1, FGFR1, FGFR2, FLT3, GNA11, GNAQ, GNAS, HRAS, IDH1, IDH2, JAK2, KDR (VEGFR2), KRAS, MGMT, MGMT-Me, MLH1, MPL, NOTCH1, NRAS, PDGFRA, Pgp, PIK3CA, PR, PTEN, RET, RRM1, SMO, SPARC, TLE3, TOP2A, TOPO1, TP53, TS, TUBB3, VHL, CDH1, ERBB4, FBXW7, HNF1A, JAK3, NPM1, PTPN11, RBI, SMAD4, SMARCB1, STK1, MLH1, MSH2, MSH6, PMS2, ROS1, ERCC1, 5T4 (TPBG), B7-H3, CCR7, CD105, CD22, CD46, CD47, CD56, CD70, CD71, CD79b, CDH6, CLDN6, CLDN18.2, CLEC12A, DLL3, DR5, ERBB3 (HER3), EPCAM, FOLR1, IGF1R, IL2RA (CD25), IL3RA, ITGB6, LIV-1, LRRC15, mesothelin (MSLN), NaPi2b (SLC34A2), nectin-4, PTK7, ROR1, SEZ6, SLC44A4, SLITRK6, Tissue Factor (TF), TROP2 or B7-H4.

[0070] As used herein, the term “biological half-life” of a substance generally refers to a pharmacokinetic parameter which specifies the time required for one half of the substance to be removed from an organism following introduction of the substance into the organism.

[0071] As used herein, the term “long-acting reagent” generally refers to reagents, when linked to another compound, can increase the biological half-life of the other compound. In some cases, an albumin protein or an albumin protein-binding peptide. The albumin protein or albumin protein-binding peptide may extend the half-life of the conjugate. In some cases, water soluble polymers such as polyethylene glycol (PEG) can be conjugated to the compounds disclosed herein. An increase in half-life, an increased solubility, and decreased clearance by the kidney and decreased enzymatic degradation may be attributed to conjugates of a variety of water soluble polymers and functional agents, including PEG conjugates to the compounds disclosed herein. In some cases, another approach to maintaining substance stability in the plasma is to form three-dimensional (3D) drug nanoparticles, which promote the formation of 3D intramolecular and / or intermolecular structures to block intramolecular cleavage sites. Encapsulation of substances in nanoparticles can reduce the apparent drug clearance from plasma, thereby enhancing the apparent drug circulation half-life and potential cumulative drug delivery to the target tissues.

[0072] The term “pharmaceutically acceptable” as used herein generally refers to a form of the compound that is safe for administration to a subject. For example, a free base, a salt form, a solvate, a hydrate, a prodrug or derivative form of a compound described herein, which has been approved for mammalian use, via oral ingestion or any other route of administration, by a governing authority or regulatory agency, such as the Food and Drug Administration (FDA) of the United States, is pharmaceutically acceptable.

[0073] Included in the compounds of Formulas I, T Ila-IIc, Illa-IIIc, IVa-IVc, Il’a-II’c, Ill’a- III’c, and IV’a-IV’c are the pharmaceutically acceptable salt forms of the free-base compounds. The term “pharmaceutically-acceptable salts” as used herein generally refers to salts, commonly used to form alkali metal salts and to form addition salts of free acids or free bases, which have been approved by a regulatory agency. Salts are formed from ionic associations, charge-charge interactions, covalent bonding, complexation, coordination, etc. The nature of the salt is not critical, provided that it is pharmaceutically acceptable.

[0074] As used herein, the term “pharmaceutically acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of subjects without undue toxicity, irritation, allergic response and the like, and are commensuratewith a reasonable benefit / risk ratio. For example, Berge et al. describes pharmaceutically acceptable salts in detail in Pharmaceutical Sciences (1977) 66: 1-19. Pharmaceutically acceptable salts of the compounds provided herein include those derived from suitable inorganic and organic acids and bases. Inorganic acids from which salts can be derived include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Organic acids from which salts can be derived include, but are not limited to, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like. Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, besylate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like. In some embodiments, organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p- toluenesulfonic acid, salicylic acid, and the like.

[0075] Pharmaceutically acceptable salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and other amine salt. Inorganic bases from which salts can be derived include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Organic bases from which salts can be derived include, but are not limited to, primary' , secondary', and tertiary amines, substituted amines, including naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like, examples include, but are not limited to, isopropylamine, trimethylamine, diethylamine, tri ethylamine, tri propyl amine, and ethanolamine. In some embodiments, the pharmaceutically acceptable base addition salt is ammonium, potassium.sodium, calcium, or magnesium salts. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary' amines, substituted animes including naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like, such as isopropylamine, trimethylamine, di ethyl amine, tn ethylamine, tri propyl amine, and ethanolamine. In some embodiments, the pharmaceutically acceptable base addition salt is chosen from ammonium, potassium, sodium, calcium, and magnesium salts. Bis salts (i.e. , two counterions) and higher salts (e.g. , three or more counterions) are encompassed within the meaning of pharmaceutically acceptable salts.

[0076] As used herein, the term “solvate” refers to compounds that further include a stoichiometric or non-stoichiometric amount of solvent bound by non-covalent mtermolecular forces. The solvate can be of a disclosed compound or a pharmaceutically acceptable salt thereof. Where the solvent is water, the solvate is a “hydrate”. Other solvates include, but are not limited to, methanol, ethanol, isopropanol, ethyl acetate, tetrahydrofuran, dimethyl sulfoxide, and N,N-dimethylfonnamide. Pharmaceutically acceptable solvates and hydrates are complexes that, for example, can include I to about 100, or I to about 10, or one to about 2, 3 or 4, solvent or water molecules.

[0077] As used herein, and unless otherwise specified, “prodrug” refers to a compound that can be converted under physiological conditions or by solvolysis to a biologically active compound described herein. Thus, the term “prodrug” refers to a precursor of a biologically active compound that is pharmaceutically acceptable. A prodrug can be inactive when administered to a subject, but is converted in vivo to an active compound, for example, by hydrolysis. A discussion of prodrugs is provided in Higuchi, T., et al , “Pro-drugs as Novel Delivery Systems,” A.C.S. Symposium Series, Vol.14, and in Bioreversible Carriers in Drug Design, ed. Edward B. Roche, American Pharmaceuti cal Association and Pergamon Press, 1987, both of which are incorporated in full by reference herein. The term “prodrug” is also meant to include any covalently bonded carriers, which release the active Formulas I, I’ Ila-IIc, Illa-IIIc, IVa-IVc, Il’a-II’c, IIFa-IITc, and IV’a-IV’c in vivo when such prodrug is administered to a mammalian subject. Prodrugs of an active compound, as described herein, can be prepared by modifying functional groups present in the active Formulas I, I’ Ila-IIc, Illa-IIIc, IVa-IVc, Il’a-II’c, Ill’a- III’ c, and IV’a-IV’c in such a way that the modifications are cleaved, either in routine manipulation or in vivo, to the parent active compound. Prodrugs include compounds wherein ahydroxy, amino or mercapto group is bonded to any group that, when the prodrug of the active Formulas I, I’ Ila-IIc, Illa-IIIc, IVa-IVc, Il’a-II’c, Ill’a-III’c, and IV’a-IV’c is administered to a mammalian subject, cleaves to form a free hydroxy, free ammo or free mercapto group, respectively.

[0078] The term ‘‘isomers” as used herein generally refers to different compounds that have the same molecular formula, including any and all geometric isomers and stereoisomers. “Stereoisomers” are isomers that differ only in the way the atoms are arranged in space. For example, “isomers” include geometric double bond cis- and trans-isomers, also termed E- and Z- isomers; R- and S-enantiomers; diastereomers, (d)-isomers and (l)-isomers, racemic mixtures thereof; and other mixtures thereof, as falling within the scope of this disclosure, unless specified otherwise. As used herein, the term “tautomer” is a type of isomer that includes two or more interconvertible compounds resulting from at least one formal migration of a hydrogen atom and at least one change in valency (e.g., a single bond to a double bond, a triple bond to a single bond, or vice versa).

[0079] hi some embodiments, the compound(s) of Formulas I, I’ Ila-IIc, Illa-IIIc, IVa-IVc, Il’a- II’c, Iira-III’c, and IV’a-IV’c is used to treat a subject by administering the compound(s) as a pharmaceutical composition. To this end, the compound(s), in one embodiment, is combined with one or more pharmaceutically acceptable excipients, including earners, diluents or adjuvants, to form a suitable composition, which is described in more detail herein.

[0080] The terra “excipient” as used herein generally refers io any pharmaceutically acceptable additive, carrier, adjuvant, or other suitable ingredient, other than the acti ve pharmaceutical ingredient (API), which is Apically included for formulation and / or administration purposes.

[0081] The term “diluent” as used herein generally refers to an agent used as filler in order to achieve the desired composition volume or weight. The diluent may be present in the pharmaceutical composition within granules in the form of a single compound or in the form of a mixture of compounds. Non-limiting examples of diluent include lactose, starch, pregelatinized starch, microcrystalline cellulose, silicified microcrystalline cellulose, cellulose acetate, dextrose, mannitol, sodium phosphate, potassium phosphate, calcium phosphate, fructose, maltose, sorbitol, or sucrose.

[0082] The term “adjuvant,” as used herein generally refers to any substance or mixture of substances that increases the efficacy or potency of a compound disclosed herein on a target where the adjuvant is used together with the compound disclosed herein. However, when the adjuvant is used alone, no pharmacological effect is observed on the same target.

[0083] The phrase “effective amount” as used herein generally refers to quantifying the amount of each agent, which will achieve the goal of improvement in disorder severity and thefrequency of incidence over treatment of each agent by itself, while avoiding adverse side effects typically associated with alternative therapies. The effective amount, in one embodiment, is administered in a single dosage form or in multiple dosage forms.

[0084] The terms “treat,” “treating,” “treatment,” and “therapy” as used herein generally refer to therapy, including without limitation, curative therapy, prophylactic therapy, and preventative therapy. Prophylactic treatment generally constitutes either preventing the onset of disorders altogether or delaying the onset of a pre-clinically evident stage of disorders in individuals. Treatment includes the medical management of a patient with the intent to cure, ameliorate, stabilize, or prevent a disease, pathological condition, or disorder. This term includes active treatment, that is, treatment directed specifically toward the improvement of a disease, pathological condition, or disorder, and also includes causal treatment, that is, treatment directed toward removal of the cause of the associated disease, pathological condition, or disorder. In addition, this term includes palliative treatment, that is, treatment designed for the relief of symptoms rather than the curing of the disease, pathological condition, or disorder; preventative treatment, that is, treatment directed to minimizing or partially or completely inhibiting the development of the associated disease, pathological condition, or disorder; and supportive treatment, that is, treatment employed to supplement another specific therapy directed toward the improvement of the associated disease, pathological condition, or disorder.

[0085] As used herein, the term “prevent” or “preventing” refers to precluding, averting, obviating, forestalling, stopping, or hindering something from happening, especially by advance action. It is understood that where reduce, inhibit or prevent are used herein, unless specifically indicated otherwise, the use of the other two words is also expressly disclosed.

[0086] Regardless of the route of administration selected, the compounds of the present invention, which may be used in a suitable hydrated form, and / or the pharmaceutical compositions of the present invention, are formulated into pharmaceutically acceptable dosage forms or by other conventional methods know to those of skill in the art.Cytotoxic Properties

[0087] A possible mechanism that may explain the biological properties of the compounds disclosed herein is their alkylation of certain nucleic acids in a duplex DNA. For example, duocarmycins, such as duocarmycin SA, are the most powerful anticancer agents and may exhibit low picomolar IC50 values across several cancer cell lines. Duocarmycins and their analogs are small-molecule DNA minor groove binding agents that may selectively alkylate adenine at the N3 position. Subsequently, this irreversible alky lation of DNA by duocarmycins and their analogs may result in over stabilization of the DNA double helix, which may lead to impaired repair, transcription and replication of DNA, and even G2-M phase cell cycle arrest. Anumber of duocarmycins have entered clinical trials, including adozelesin, carzelesin and bizelesin. The compounds disclosed herein may be DNA alkylating agents that may cause tumor cell cycle arrest. The compounds disclosed herein may be cytotoxic.Pharmaceutical Compositions / F o rm illations

[0088] One embodiment provides a pharmaceutical composition comprising a compound of Formulas I, I’ Ila-IIc, Illa-IIIc, IVa-IVc, Il’a-II’c, IIFa-IIFc, and IV’a-IV’c, or a stereoisomer, tautomer, hydrate, solvate or pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable excipient.

[0089] In some embodiments, the compounds described herein are formulated into pharmaceutical compositions. Pharmaceutical compositions are formulated in a conventional manner using one or more pharmaceutically acceptable inactive ingredients that facilitate processing of the active compounds into preparations that can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen. A summary of pharmaceutical compositions described herein can be found, for example, in Remington: The Science and Practice of Pharmacy, Nineteenth Ed., Easton, Pa.: Mack Publishing Company (1995); Hoover, John E., Remington’s Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania (1975); Liberman, H.A. and Lachman, L„, Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y. (1980); and Pharmaceutical Dosage Forms and Drug Delivery Systems, Seventh Ed., Lippincott Williams & Wilkins (1999), herein incorporated by reference for such disclosure.

[0090] A pharmaceutical composition, as used herein, refers to a mixture of a compound of Formulas I, I’ Ila-IIc, Illa-IIIc, IVa-IVc, Il’a-II’c, IIFa-IIFc, and IV’a-IV’c with other chemical components (i.e. pharmaceutically acceptable inactive ingredients), such as carriers, excipients, binders, filling agents, suspending agents, flavoring agents, sweetening agents, disintegrating agents, dispersing agents, surfactants, lubricants, colorants, diluents, solubilizers, moistening agents, plasticizers, stabilizers, penetration enhancers, wetting agents, anti-foaming agents, antioxidants, preservatives, or one or more combination thereof. The pharmaceutical composition facilitates administration of the compound to an organism. In practicing the methods of treatment or use provided herein, therapeutically effective amounts of compounds described herein are administered in a pharmaceutical composition to a mammal having a disease, disorder, or condition to be treated. In some embodiments, the mammal is a human. A therapeutically effective amount can vary widely depending on the severity of the disease, the age and relative health of the subject, the potency of the compound used and other factors. The compounds can be used singly or in combination with one or more therapeutic agents as components of mixtures.

[0091] The pharmaceutical formulations described herein are administered to a subject by appropriate administration routes, including but not limited to, oral, parenteral (e.g., intravenous, subcutaneous, intramuscular), intranasal, buccal, topical, rectal, or trans dermal administration routes. The pharmaceutical formulations described herein include, but are not limited to, aqueous liquid dispersions, self-emulsifying dispersions, solid solutions, liposomal dispersions, aerosols, solid dosage forms, powders, immediate release formulations, controlled release formulations, fast melt formulations, tablets, capsules, pills, delayed release formulations, extended release formulations, pulsatile release formulations, multiparticulate formulations, and mixed immediate and controlled release formulations.

[0092] All formulations for oral administration are in dosages suitable for such administration. Examples of such dosage units are tablets or capsules. In some embodiments, these contain an amount of active ingredient from about 1 to 2000 mg, advantageously from about 1 to 500 mg, and typically from about 5 to 150 mg. A suitable daily dose for a human or other mammal vary widely depending on the condition of the patient and other factors, but, once again, can be determined using routine methods and practices.

[0093] Conventional formulation techniques include, e.g., one or a combination of methods: (1) dry mixing, (2) direct compression, (3) milling, (4) dry' or non-aqueous granulation, (5) wet granulation, or (6) fusion. Other methods include, e.g., spray drying, pan coating, melt granulation, granulation, fluidized bed spray drying or coating (e.g., wurster coating), tangential coating, top spraying, tableting, extruding and the like.Numbered Embodiments

[0094] The following embodiments recite non-limiting permutations of combinations of features disclosed herein. Other permutations of combinations of features are also contemplated. In particular, each of these numbered embodiments is contemplated as depending from or relating to every previous or subsequent numbered embodiment, independent of their order as listed.

[0095] Embodiment 1. A compound of Formula I or Formula F:Formula I Formula I' or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:Rc is a reactive group;Linker is a cleavable linker or a non-cleavable linker; andR2is H or CH3; andR3is H, OH, or OCH3.

[0096] Embodiment 2. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H or CH3; andR3is H, OH, or OCH3.

[0097] Embodiment 3. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is H.

[0098] Embodiment 4. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is OH.

[0099] Embodiment 5. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is OCH3.

[0100] Embodiment 6. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is H.

[0101] Embodiment 7. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is OH.

[0102] Embodiment 8. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is OCH3.

[0103] Embodiment 9. The compound of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is H.

[0104] Embodiment 10, The compound of any one of Embodiments 1 to 9, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:AX is halide or hydroxy, or C(O)-AX is an active ester;LG is halide, nonaflate, tritiate, fluorosulfonate, tosylate, mesylate, or besylate;RAis independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl- alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxy carbonyl, or alkylsulfonyl;RBis independently a protecting group, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each of R6, R7and R8is independently hy drogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each R9is independently halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, C1-C4 heteroalkyl, -C(O)H, -C(O)OH, -CN, C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyd, C6-C10 aryl, 5- to 10-membered heteroaryl, - C(O)(C1-C4 alkyl), -C(O)O(C1-C4 alkyl), -C(O)NH2, -C(O)NH(C1-C4 alkyl), -C(O)N(C1-C4 alkyl)2, -NH2, -NH(C1-C4 alkyl), -N(C1-C4 alkyl)2, -NH(C2-C4alkylene)-OH, -NH(C2-C4alkylene)-O-(C1-C4 alkyl), -OH, -O(C1-C4 alkyl), -O(C1-C4 haloalkyl), -O(C2-C4alkylene)- NH2, -O(C2-C4alkylene)-NH-(C1-C4 alkyl), -O(C2-C4alkylene)-N-(C1-C4 alkyl)2, -O(C1-C4 alkylene)-C(O)OH, -O(C1-C4 alkylene)-C(O)O-(C1-C4 alkyl), -O(C2-C4alkenyl), -O(C1-C4 alkylene)-(C6-C10 aryl), -O(C1-C4 alkylene)-(5- to 10-membered heteroaryl), -0(C6-C10 aryl), - SH, S(O)2OH, -S(O)2(C1-C4 alkyl), -S(O)2NH2, -S(O)2NH(C1-C4 alkyl), or -S(0)2N(C1-C4 alky 1)2; or two R9, together with atoms to which they are attached, form a C3-C10 cycloalkyl or a 3- to 10-membered heterocycloalkyl ring; and each of p and q is independently an integer of 0-3.

[0105] Embodiment 11. The compound of any one of Embodiments 1 to 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof:Linker is -(L^r-tE1^-*;* denotes a connection to Rc;each L1and L2is independently a bond, -O-, -S-, -NH-, -NRA-, -C(=O)-, -C(=O)O-, -OC(=O)-, -C(=O)NH- -NHC(=O)-, -C(=O)NRD- -NRDC(=O)-, -(CH2-O-CH2)m-, Ci-C6alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, -C6-C10 aryl-, or 5- to 10-membered heteroarylene, wherein each C1-C6 alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, - C6-C10 aryl-, and 5- to 10-membered heteroarylene is optionally substituted with 1, 2, or 3 R9;RDis independently -CR3R4R5; each R3, R4, and R5is independently hydrogen, halogen, -U, or -G;-U is independently C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkeny l, or C2-C6 alkynyl; wherein each C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, Ci-Cg heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl is optionally substituted with 1, 2, or 3 R9and / or 1 or 2 -G;-G is independently C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, Ce-Cw aryl, or 5- to 10-membered heteroaryl; wherein each C3-C10 cycloalkyl. 3- to 10-membered heterocy cloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl is optionally substituted with 1, 2, or 3 R9; each R9is independently as defined in Embodiment 1; and each of m, r and s is independently an integer of 1-12.

[0106] Embodiment 12. The compound of any one of Embodiments 1-10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:RAis independently as defined in Embodiment 2; n is independently an integer of 1-8; and# denotes a connection to Rc or a group connected to Rc.

[0107] Embodiment 13 The compound of any one of Embodiment 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas II, II’, III, III’, IV or IV’:LG is halide, nonaflate, inflate, fluorosulfonate, tosylate, mesylate, or besylate;RAis independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl- alkylene, alkylamino-alkylene, alkydthio-alkylene, alkylcarbonyl, alkoxy carbonyl, or alkylsulfonyl;RBis independently a protecting group, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each of R6, R7and R8is independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each R9is independently halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, C1-C4 heteroalkyl, -C(O)H, -C(O)OH, -CN, C3-C10 cycloalkyl, 3- to 10-membered heterocycloalky l, C6-C10 aryl, 5- to 10-membered heteroaryl, - C(O)(C1-C4 alkyl), -C(O)O(C1-C4 alkyl), -C(O)NH2, -C(O)NH(C1-C4 alkyl), -C(O)N(C1-C4 alkyl)2, -NH2, -NH(C1-C4 alkyl), -N(C1-C4 alkyl)2, -NH(C2-C4alkylene)-OH, -NH(C2-C4alkylene)-O-(C1-C4 alkyl), -OH, -O(C1-C4 alkyl), -O(C1-C4 haloalkyl), -O(C2-C4alkylene)- NH2, -O(C2-C4alkydene)-NH-(C1-C4 alkyl), -O(C2-C4alkylene)-N-(C1-C4 alkyl)2, -O(C1-C4 alkylene)-C(O)OH, -O(C1-C4 alkylene)-C(O)O-(C1-C4 alkyl), -O(C2-C4alkenyl), -O(C1-C4alkylene)-(C6-C10 aryl), -0(C1-C4 alkylene)-(5- to 10-membered heteroaryl), -0(C6-C10 aryl), - SH, S(O)2OH, -S(O)2(C1-C4 alkyl), -S(O)2NH2, -S(O)2NH(C1-C4 alkyl), or -S(O)2N(C1-C4 alky 1)2; or two R9, together with atoms to which they are attached, form a C3-C10 cycloalkyl or a 3- to 10-membered heterocycloalkyl ring; each L1and L2is independently a bond, -O-, -S-, -NH-, -NRD-, -C(=O)-, -C(=O)O- -OC(=O)-, -C(=O)NH-, -NHC(=O)-, -C(=O)NRD-, -NRDC(=O)-, -(CH2-O-CH2)m- Ci-Cg alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, -C6-C10 aryl-, or 5- to 10-membered heteroarylene, wherein each C1-C6 alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, Ci-Cg heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, - C6-C10 aryl-, and 5- to 10-membered heteroarylene is optionally substituted with 1, 2, or 3 R9;RDis independently -CR3R4R5; each R3, R4, and R5is independently hydrogen, halogen, -U, or -G;— U is independently C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2- Cg alkenyl, or C2-C6 alkynyl; wherein each C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl is optionally substituted with 1, 2, or 3 R9and / or 1 or 2 -G;-G is independently C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl; wherein each C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl is optionally substituted with 1, 2, or 3 R9; m is independently an integer of 1-12; each of p and q is independently an integer of 0-3; n is independently an integer of 0-8; and each t and u is independently an integer of 1-4.

[0108] Embodiment 14. The compound of Embodiment 13, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas Ila, Il’a, Illa, Ill’a,, IVa or IV’a::wherein:R1is independently as defined in Embodiment 13;R2is H or CH3;R3is H, OH, or OCH3; each of RAand Rc is independently as defined in Embodiment 13;L1is independently as defined in Embodiment 13; n is independently an integer of 0-8; and u is independently an integer of 1-4.

[0109] Embodiment 15. The compound of Embodiments 13, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas lib, Il’b, Illb, Ill’b, IVb or IV’b:wherein:R1is independently as defined in Embodiment 13;R2is H or CH3;R3is H, OH, or OCH3; each of RAand Rc is independently as defined in Embodiment 13; m is independently an integer of 1-8; and n is an integer of 0-8.

[0110] Embodiment 16. The compound of Embodiment 15, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according toFormulas lie, II’c, IIIc, III’c, IVc or IV’c:Formula ll'cFormula IVc , orFormula IV'c wherein:Rc is independently as defined in Embodiment 13;R2is H or CH3;R3is H, OH, or OCH3; x is an integer of 0-4; and y is independently an integer of 1-8.[OHl] Embodiment 17. The compound of any one of Embodiments 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein. R2is CH3; and R3is H.

[0112] Embodiment 18, The compound of any one of Embodiments 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein R2is CH3; and R3is OH.

[0113] Embodiment 19. The compound of any one of Embodiments 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein: R2is CH3; and R3is OCH3.

[0114] Embodiment 20. The compound of any one of Embodiments 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein: R2is H; and R3isH.

[0115] Embodiment 21. The compound of any one of Embodiments 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein: R2is H; and R3is OH.

[0116] Embodiment 22. The compound of any one of Embodiments 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein: R2is H; and R3is OCH3.

[0117] Embodiments 23. The compound of Embodiment 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is:wherein:R1is as defined in Embodiment 1; andRc is as defined in Embodiment 10.

[0118] Embodiment 24. The compound of any one of Embodiments 1 or 3-23, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein R1is:

[0119] Embodiment 25. The compound of Embodiment 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is:Rc is as defined in Embodiment 10.

[0120] Embodiment 26, The compound of any one of Embodiments 1-25, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein Rc is:

[0121] Embodiment 27. The compound of Embodiment 26, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein Rc is:

[0122] Embodiment 28. A pharmaceutical composition comprising a compound of any one of Embodiments 1-27, or pharmaceutically acceptable salt, solvate, diastereomeric mixture, or individual enantiomers thereof, and a pharmaceutically acceptable carrier.

[0123] Embodiment 29. A method of treating a cancer in a mammal suffering therefrom, comprising administering to the mammal a therapeutically effective amount of a compound of any one of Embodiments 1-27 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, or a pharmaceutical composition of Embodiment 28.

[0124] Embodiment 30. A conjugate comprising a cell surface targeting agent or a long-acting reagent, wherein the cell surface targeting agent or the long-acting reagent is attached to a compound of any one of Embodiments 1 to 27 or a derivative of a compound of any one of Embodiments 1 to 27, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the cell surface targeting agent is a small molecule, a polypeptide, or an antibody or an antigen-binding portion thereof, and wherein the long-acting reagent comprising a polymer, human serum albumin (HSA), or a nanoparticle.

[0125] Embodiment 31 . The conjugate of Embodiment 30, wherein the cell surface targeting agent is a monoclonal antibody, a Fab, a Fab’, a F(ab’), a Fv, a disulfide linked Fc, a scFv, a single domain antibody, a diabody, a bi-specific antibody, or a multi-specific antibody.

[0126] Embodiment 32. The conjugate of Embodiment 30, wherein the cell surface targeting agent binds to a target molecule.

[0127] Embodiment 33. The conjugate of Embodiment 32, wherein the target molecule comprises a tumor suppressor, a metabolic enzyme, a protein aggregate, or a haploinsufficient protein.

[0128] Embodiment 34. The conjugate of Embodiment 32, wherein the target molecule comprises p53 mutant or Von Hippel-Lindau tumor suppressor (VHL).

[0129] Embodiment 35. A pharmaceutical composition comprising a conjugate of any one of Embodiments 30-34, and a pharmaceutically acceptable carrier.

[0130] Embodiment 36. A method of treating a cancer in a mammal suffering therefrom, comprising administering to the mammal a therapeutically effective amount of a conjugate of any one of Embodiments 30-34, or a pharmaceutical composition of Embodiment 35, wherein the conjugate binds to a target antigen associated with the cancer.EXAMPLESChemical Synthesis

[0131] Methods of the present invention may include the use of at least one compound of Formulas I-IV, Ila-IIc, Illa-IIIc, and IVa-IVc, which inhibits programmed necrosis in the regulation of repair and / or functional performance of a wide range of cells, tissues and organs, and have therapeutic and cosmetic applications ranging from regulati on of neural tissues, bone and cartilage formation and repair, regulation of spermatogenesis, regulation of smooth muscle, regulation of lung, liver and other organs arising from the primitive gut, regulation of hematopoietic function, regulation of skin and hair growth, etc. Accordingly, the methods and compositions of the present invention include the use of the subject mln bi tors for all such uses as inhibitors of programmed necrosis may be implicated. Moreover, the subject methods can be performed on cells which are provided in culture (in vitro), or on cells in a whole animal (in vivo).

[0132] The examples and preparations provided below illustrated and exemplify the compounds described herein and methods of preparing such compounds. In general, the compounds described herein may be prepared by processes known in the general chemical arts.

[0133] The compounds of the present invention can be prepared using various synthetic routes, including those described below, starting from commercially available materials. Starting materials of the invention, are either known, commercially available, or can be synthesized in analogy to or according to methods that are known in the art. Many starting materials may be prepared according to known processes and, in particular, can be prepared using processes described in the examples. In synthesizing starting materials, functional groups in some casesare protected with suitable protecting groups when necessary. Functional groups may be removed according to known procedures in the art.

[0134] The protection of functional groups by protecting groups, the protecting groups themselves, and their removal reactions (commonly referred to as “deprotection”) are described, for example, in standard reference works, such as J.F.W. McOmie, Protective Groups in Organic Chemistry, Plenum Press, London and New York (1973), in T.W. Greene, Protective Groups in Organic Synthesis, Wiley, New' York (1981), in The Peptides, Volume 3, E. Gross and J. Meienhofer editors, Academic Press, London and New' York (1981).

[0135] All synthetic procedures described herein can be carried out under known reaction conditions, advantageously under those described herein, either in the absence or in the presence (usually) of solvents or diluents.

[0136] The invention further encompasses “intermediate” compounds, including structures produced from the synthetic procedures described, whether isolated or not, prior to obtaining the finally desired compound. Structures resulting from carrying out steps from a transient starting material, structures resulting from divergence from the described method(s) at any stage, and structures forming starting materials under the reaction conditions are all “intermediates” included in the invention. Further, structures produced by using starting materials in the form of a reactive derivative or salt, or produced by a compound obtainable by means of the process according to the invention and structures resulting from processing the compounds of the invention in situ are also within the scope of the invention.

[0137] New starting materials and / or intermediates, as well as processes for the preparation thereof, are likewise the subject of this invention. In select embodiments, such stalling materials are used and reaction conditions so selected as to obtain the desired compound(s).

[0138] Starting materials of the invention, are either known, commercially available, or can be synthesized in analogy to or according to methods that are known in the art. Many starting materials may be prepared according to known processes and, in particular, can be prepared using processes described in the examples. In synthesizing starting materials, functional groups in some cases are protected with suitable protecting groups when necessary. Protecting groups, their introduction and removal are described above.

[0139] All reagents and solvents were obtained commercially unless stated otherwise. All commercial reagents and solvent were used without purification unless stated otherwise. When required, some reagents and solvents were purified by standard techniques. For example, tetrahydrofuran may be purified by distillation from sodium.

[0140] Reaction progress was monitored by reverse-phase HPLC and / or thin-layer chromatography (TLC), Liquid chromatography -mass spectrometry was performed using eitherWaters or Shimadzu 201 OEV LCMS instruments using water and acetonitrile or methanol doped with 0.1% formic acid. TLC was performed using silica gel 60 F254 pre-coated plates (0.25 mm). Flash chromatography was performed using silica gel (32-63 pm particle size) or aluminum oxide (activated, basic, aboutl50 mesh size). Automated chromatographic purification was carried out using pre-packed silica or Cl 8 cartridges (from RediSep and Luknova) and eluted using an ISCO Companion system. Reverse phase purifications were conducted using water and acetonitrile or methanol doped with 0.1% formic acid. All final product compounds were purified using one of these two chromatographic methods. Purity and characterization of compounds was established by a combination of TLC, liquid chromatography-mass spectroscopy (LC-MS) and Nuclear Magnetic Resonance (NMR) analytical techniques.JH and13C NMR spectra were obtained on a Joel 400 spectrometer at 400 MHz and 101 MHz, respectively. Chemical shifts are reported in 6 (ppm) and were internally referenced to deuterated solvent signals.

[0141] The size and scale of the synthetic methods will vary depending on the desired amount of end product. It is understood that while specific reactants and amounts are provided in the Examples, one of skill in the art knows other alternative and equally feasible sets of reactants that will also yield the same compounds. Thus, where general oxidizers, reducers, solvents of various nature (aprotic, nonpolar, polar, etc.) are utilized, equivalents will be known in the art and are herein contemplated for use in the present methods.

[0142] Many of the steps below indicate various workups following termination of the reaction. A work-up involves generally quenching of a reaction to terminate any remaining catalytic activity and starting reagents. This is generally followed by addition of an organic solvent and separation of the aqueous layer from the organic layer. The product is typically obtained from the organic layer and unused reactants and other spurious side products and unwanted chemicals are generally trapped in the aqueous layer and discarded. The work-up in standard organic synthetic procedures found throughout the literature is generally followed by drying the product by exposure to a drying agent, such as anhydrous NazSCh, to remove any excess water or aqueous byproducts remaining partially dissolved in the organic layer and concentration of the remaining organic layer. Concentration of product dissolved in solvent may be achieved by any known means, such as evaporation under pressure, evaporation under increased temperature and pressure, and the like. Such concentration may be achieved by use of standard laboratoiy equipment such as rotary-evaporator distillation, and the like. This is optionally followed by one or more purification steps which may include, but is not limited to, flash column chromatography, filtration through various media and / or other preparative methods known in the art and / or crystallization / recrystallization. (See, for instance, Addison Ault, “Techniques andExperiments for Organic Chemistry,” 6th Ed., University Science Books, Sausalito, Calif, 1998, Ann B. McGuire, Ed., pp.45-59).LC-MS Conditions

[0143] HPLC-MS analyses are performed on a Waters ACQUITY UPLC with SQ mass detector and PDA eZ detector. The column used is a Phenomenex Kinetex Cl 8 column (1 ,7um, 2. 1 x 50 mm). The mobile phase consists of eluent A (water, 0.05% TFA) and eluent B (CH3CN, 0.05% TFA), and the elution proceeds at 0.5 mL / min. The initial conditions are 90% A, then 90% A to 10% A linearly decreased within 1.75 min, then from 10% A to 90% A within 0.25 min. The total run time is 2 minutes.General Synthetic Routes

[0144] There are many routes available to synthesize compounds of Formulas I-IV, Ila-IIc, Illa- IIIc, and IVa-IVc. Below are synthetic routes leading to specific compounds of Formulas I-IV, Ila-IIc, Illa-IIIc, and IVa-IVc. The synthetic routes shown are not limiting but are examples from many available routes.

[0145] In one example, as shown in Scheme A, the free hydroxy group on the indole (M-ll) is alkylated via a Mitsunobu reaction with another alcohol (M-12). The benzyl protecting group is removed using palladium catalyzed hydrogenation to provide the secondary amine (M-13). Alkylation of the secondary amine (M-13) with a tosylate (M-14) affords ester (M-15) which is hydrolyzed under basic condition to provide the carboxylic acid (M-16). Coupling reaction between the carboxylic acid (M-16) and 2,3-dihydro-lH-benzo[e]indol (M-16B) forms a protected duocarmycin analog (M-17). Deprotection affords duocarmycin analog (M-18) which is a member of Formula I.

[0146] Scheme A: Synthesis of Compound M-18Note: R1is as defined in Embodiment 1; Rc is as defined in Embodiment 10; L1and s are as defined in Embodiment 11; n is an integer of 0-8; Rdis Rc or a precursor of Rc; and Reis R1or a precursor of R1.

[0147] The synthetic route shown in Scheme A can be adapted to synthesize compounds ofFormula I and Formula I’:Formula I Formula I1with regard to the R1, R2, and R3groups.

[0148] For various R1groups, a properly protected Reprecursor can used according to Scheme A and other synthetic routes disclosed herein. For various disaccharide groups, the hydroxy groups are preferably protected as acetates in Re; the carboxylate groups are preferably protected as esters. Other protecting group may be employed as well; and the free anomeric hydroxy group can be activated for the ensuing coupling with the appropriate heteroaryl group to form the precursor such as(similar to M-16B in Scheme A).

[0149] For various R2groups, a precursor such a(similar to M-16B in SchemeA) can be used with the appropriate R2group installed on the heteroaryl group, then follow similar steps disclosed in Scheme A.

[0150] For various R3groups, a precursor such as(similar to M-ll in Scheme A) can be used according to the synthetic route disclosed in Scheme A. Below is an example synthetic route shown in Scheme B and Scheme C to illustrate how to install an R3group (either OH or OCH3) on the indole ring. Using an appropriate aniline (LUX118-8), a hydrazine (LUX118-9) can be made. Then a Fischer indole synthesis can be employed to prepare the Redecorated indole (LUX118-12A and LUX118-13A)

[0151] Scheme B: Synthesis of Compound LUX118-12A and LUX118-13ALUX118-12A LUX118-13A

[0152] Taking LUX118-13A as the starting material, an example Compound LUX118-1 can be made according to Scheme C The reactions include protection of the hydroxy group; conversion of the nitro group into an amine group using Pd / C and hydrogen; coupling of the amine group with a carboxylate (Int A) on an aryl group; deprotection of the methyl ester to free a carboxylic acid group; coupling of the carboxylic group with benzo[e]indol-5-ol derivative (Int C) to complete the synthesis. The IC50 for Example Compound LUX118-1 is 5.4 nM for the DU145 cell line.

[0153] Scheme :C Synthesis of Example Compound LUX-118-1

[0154] In another example, as shown in Scheme D, the phenol group in M-20 reaction with primary alcohol M-21 via a Mitsunobu reaction to afford benzoate M-22, which is hydrolyzed to provide the carboxylic acid M-23. Coupling reaction between the carboxylic acid M-23 and amine M-24 afford the amide M-25. Another hydrolysis reaction reveals the carboxylic acid M- 26, which couples with 2,3-dihydro-lH-benzo[e]indol (M-16B) to form a protected duocarmycin analog (M-27). Deprotection affords duocarmycin analog (M-28) which is a member of Formula I.

[0155] Scheme D: Synthesis of Compound M-27Note: R1is as defined in Embodiment 1; Rc is as defined in Embodiment 10; L1and s are as defined in Embodiment 11; m is an integer of 1-8; Rdis Rc or a precursor of Rc; and Reis R1or a precursor of R1.

[0156] Scheme D can be modified similarly according to Scheme B and Scheme C for the synthesis of compound of Formula I and Formula I’.Example 1; Synthesis of LUX107-1

[0157] Scheme 1 : Synthesis of 2,2-dimethyl-4-oxo-3,6,9,12-tetraoxa-5-azatetradecan-14-yl 4- methylbenzenesulfonateLUX107-4B LUX107-5B[0158| Synthesis of LUX107-5B

[0159] Step 1 : 2-(2-(2-Hydroxyethoxy)ethoxy)ethyl 4-methylbenzenesulfonate (LUX107-3)

[0160] To a solution of PEG 3 (30 g, 199.8 mmol) in DCM (600 mL) was added TsCI (22.4 g, 130 mmol) at room temperature, followed by the addition of TEA (55.5 mL, 400 mmol) and DMAP (2. 10 g, 16.9 mmol). The reaction was stirred at room temperature for 16 h. The mixture was then concentrated under reduced pressure directly, and the residue was purified by silica column ( / 7-heptane then / 7-heptane / EtOAc (v / v) = from 3: 1 to 2: 1 to 1 : 1) to afford LUX107-3 (16.6 g, 42% yield) as light-yellow oil. LCMS (ESI): m / z 304.9 [M + H]+, 326.8 [M + Na+];XH NMR (400 MHz, CDCI3) δ 7.78 (d, J= 8.3 Hz, 2H), 7.33 (d, J= 8.4 Hz, 2H), 4.18 - 4.13 (m,2H), 3.74 - 3.63 (m, 6H), 3,60 (d, J= 5.1 Hz, 2H), 3,59 (bs, 1H), 3.57 - 3,53 (m, 2H), 2.43 (s, 3H).

[0161] Step 2: Tert-butyl 2-(2-(2-hydroxyethoxy)ethoxy)ethoxycarbamate (LUX107-4B)LUX107-3 LUX107-4B

[0162] A solution of LUX107-3 (15 g, 49.28 mmol) and BocNHOH (9.84 g, 73.9 mmol) in DBU (15.0 mL) was stirred at room temperature under aN2 atmosphere for 16 h. LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 288.0 [M + Na+]). The mixture was purified by silica column (o-heptane) directly to remove DBU to obtain the crude for the first time. The crude was re-purified by silica column (n-heptane / EtOAc (v / v) = from 3:1 to 2: 1 to 1 : 1 then EtOAc) to afford LUX107-4B (6.7 g, 51% yield) as yellow oil with very weak UV absorption; 'H NMR (400 MHz, CDCI3) δ 7.82 (s, 1H), 4.06 - 3.98 (m, 2H), 3.77 - 3.70 (m, 4H), 3.69 - 3.65 (m, 4H), 3.64 - 3.60 (m, 2H), 1.84 (s, 1H), 1.47 (s, 9H).

[0163] STEP 3: 2,2-Dimethyl-4-oxo-3,6,9,12-tetraoxa-5-azatetradecan-14-yl 4- methylbenzenesulfonate (LUX107-5B)LUX107-4B LUX107-5B

[0164] To a solution of LUX107-4B (6.30 g, 23.8 mmol) in DCM (200 mL) was added TsCI (6.79 g, 35.6 mmol) at room temperature, followed by the addition of TEA (7.21 g, 71.2 mmol) and DMAP (290 mg, 2.37 mmol). The reaction was stirred at room temperature for 16 h. LCMS showed the expected product molecule was formed (LCMS (ESI): m / z 442.8 [M + Na+]).The mixture was concentrated under reduced pressure directly, and the residue was purified by silica column (w-heptane / EtOAc (v / v) = from 6: 1 to 5: 1 to 3:1) twice to afford LUX107-5B (4.3 g, 43% yield) as yellow oil. LCMS (ESI): m / z 436.8 [M + NH4+], 441.9 [M + Na+]; 'H NMR (400 MHz, CDCI3) δ 7.80 (d, J= 8.3 Hz, 2H), 7.47 (s, 1H), 7.34 (d, J= 8.0 Hz, 2H), 4.21 - 4. 14 (m,2H), 4.04 - 3.97 (m, 2H), 3.72 - 3.66 (m, 4H), 3.60 (s, 4H), 2.44 (s, 3H), 1.47 (s, 9H).

[0165] Scheme 2: 5-((2,2,15-trimethyl-4-oxo-3,6,9,12-tetraoxa-5,15-diazaheptadecan-17- yl)oxy)-lH-indole-2-carboxylic acid (LUX107-13B)

[0166] Step 1 : Ethyl 5-(2-(benzyl(methyl)amino)ethoxy)-lH-indole-2-carboxylate (LUX107-18)

[0167] To a stirred solution of LUX107-11 (3.70 g, 18.4 mmol) and Ph-P (10 g, 36.8 mmol) in dry THF (80 mL) was added DIAD (7.50 g, 36.8 mmol) at 0 °C under a N2 atmosphere. The mixture was stirred at 0 °C for 20 min, followed by the addition of LUX107-6Bn (3.80 g, 23.0 mmol). The reaction was allowed to warm up to room temperature and stirred for 2 days. LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 353.0 [M + H]+). The mixture was concentrated, and the residue was purified by silica column (w-heptane / EtOAc (v / v) = from 15:1 to 10:1 to 6: 1 to 4: 1) to afford crude LUX107-18 (30 g, containing much Ph ,P=O). which was used directly without further purification.

[0168] Step 2: Ethyl 5-(2-(methylamino)ethoxy)-lH-indole-2-carboxylate (LUX107-19)

[0169] To a solution of the above crude LUX107-18 (30 g, containing much PhsP=O) in 100 mL THF was added 10% Pd / C (5 g). The reaction was then stirred under a H2 atmosphere (1 atm) at room temperature for 16 h. LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 263.0 [M + H]+). The mixture was filtrated, and the filter cake was washed with THF (25 mLx3). The combined filtrate was concentrated, and the residue waspurified by silica column (DCM / MeOH (v / v) = from 60:1 to 50:1 to 40:1 to 20: 1) twice to afford LUX107-19 (920 mg, 19% yield for 2 steps) as brown solid. LCMS (ESI): m / z 263,1 [M + H]+; 'H NMR (400 MHz, CDCL) δ 8.90 (s, 1H), 7.30 (d, J= 8.9 Hz, 1H), 7.15 - 7.11 (m, 1H), 7.09 (d, J= 2.3 Hz, 1H), 7.00 (dd, J= 8.9, 2.4 Hz, 1H), 4.39 (q, J= 6.7 Hz, 2H), 4.11 (t, J = 4.0 Hz, 2H), 3.00 (t, J= 4.5 Hz, 2H), 2.52 (s, 3H), 2.43 (s, 1H), 1.40 (t, J= 7.1 Hz, 3H).

[0170] Step 3: Ethyl 5-((2,2,15-trimethyl-4-oxo-3,6,9,12-tetraoxa-5,15-diazaheptadecan-17- yl)oxy)-lH-indole-2-carboxylate (LUX107-12B)

[0171] To a stirred solution of LUX107-5B (1 g, 2.38 mmol) and LUX107-19 (750 mg, 2.86 mmol) in CH3CN (25 mL) was added DIEA (922 mg, 7.14 mmol) under a N2 atmosphere. The mixture was then warmed up to 65 °C and stirred for 6 h. LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 509.9 [M + H]+). The mixture was concentrated, and the residue was purified by silica column (DCM / MeOH (v / v) = from 100: 1 to 80: 1 to 50: 1 to 40: 1 to 30: 1) to afford crude LUX107-12B (1.4 g) as brown solid. LCMS (ESI): m / z 510.0 [M + N]+.

[0172] Step 4: 5-((2,2,15-Trimethyl-4-oxo-3,6,9,12-tetraoxa-5,15-diazaheptadecan-17-yl)oxy)- lH-indole-2-carboxylic acid (LUX107-13B)

[0173] To a solution of the above crude LUX107-12B (1.4 g, 2.78 mmol) in 14 mL was added 21 mL10% LiOH aqueous solution. The reaction was stirred at room temperature for 16 h. TLC showed completion (DCM / MeOH = 20: 1, Rf of the starting material 12B = 0.5). The mixture was quenched by adding TFA to adjust the PH to 5-6. The resulting mixture was concentrated, and the residue was purified by Reverse Phase column (0.1% TFA in H2O / CH3CN) to afford LUX107-13B (870 mg, 47% yield for 2 steps) as grey solid. LCMS (ESI): m / z 482.9 [M + H]+.

[0174] Scheme 3: (S)-(5-(2-((2-(2-(2-(aminooxy)ethoxy)ethoxy)ethyl)(methyl)amino)-ethoxy)- lH-indol-2-yl)(l-(chloromethyl)-5-hydroxy-9-methyl-lH-benzo[e]indol-3(2H)-yl)methanone(LUX107-1) and (9aS)-2-(5-(2-((2-(2-(2-(aminooxy)ethoxy)ethoxy)ethyl)(methyl)amino)-ethoxy)- lH-indole-2-carbonyl)-8-methyl-9,9a-dihydro-lH-benzo[e]cyclopropa[c]indol-4(2H)- one (LUX107-1A)

[0175] Step 1: (S)-tert-butyl 2-(2-(2-((2-((2-(l-(chloromethyl)-5-hydroxy-9-methyl-2,3-dihydro- lH-benzo[e]indole-3-carbonyl)-lH-indol-5- yl)oxy)ethyl)(methyl)amino)ethoxy)ethoxy)ethoxycarbamate (LUX107-14B)

[0176] To a solution of LUX107-13B (600 mg, 1.25 mmol) and (6)-l-(chloromethyl)-9-methyl- 2,3-dihydro-l#-benzo[e]indol-5-ol hydrochloride LUX105-6 (354 mg, 1.25 mmol) in 15 mL DMF was added EDCI (478 mg, 2.49 mmol) under a N2 atmosphere. The reaction was stirred at room temperature in dark for 16 h, LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 711.0 [M + H]+). The mixture was purified by reverse phase column(0.1% TFA in FbChCHsCN) directly then freeze-dried to afford LUX107-14B (290 mg, 33% yield) as off-white solid. LCMS (ESI): m / z 733.0 [M + H]+, 733.0 [M + Na+]; HPLC: 92.1% @210 nm, Rt = 24.4 min.

[0177] Step 2: (S)-(5-(2-((2-(2-(2-(aminooxy)ethoxy)ethoxy)ethyl)(methyl)amino)ethoxy)-lH- indol-2-yl)(l-(chloromethyl)-5-hydroxy-9-methyl-lH-benzo[e]indol-3(2H)-yl)methanone(LUX107-1)

[0178] To a solution of LUX107-14B (30 mg, 42.2 pmol) in 2 mL dry DCM at 0 °C was added 4 M HCl / dioxane (25 pL, 100 pmol) under aN2 atmosphere. The reaction was stirred at 0 °C in dark for 30 min, LCMS showed completion (LCMS (ESI): m / z 611.0 [M + H]+for LUX107-1; m / z 575.0 [M + H]+for LUX107-1A; ). The mixture was concentrated, and the residue was purified by reverse phase column (H2O / CH3CN), then freeze-dried to afford LUX107-1 (3 mg,11% yield) as light yellow solid. For LUX107-1: LCMS (ESI): m / z 610.9 [M + H]+; HPLC:60.7% @210 nm, Rt= 9.20 min. For LUX107-1-A: LCMS (ESI): m / z 574.9 [M + H]+; HPLC: 33.2% @210 nm, Rt = 8.47 min.

[0179] Step 3: (9aS)-2-(5-(2-((2-(2-(2-(aminooxy)ethoxy)ethoxy)ethyl)(methyl)amino)ethoxy)- lH-indole-2-carbonyl)-8-methyl-9,9a-dihydro-lH-benzo[e]cyclopropa[c]indol-4(2H)-one(LUX107-1A)LUX107-1-A

[0180] To a solution of LUX107-14B (150 mg, 211 pmol) in 10 mL dry DCM at 0 °C was added 4 M HCl / dioxane (125 pL, 500 pmol) under aN2 atmosphere. The reaction was stirred at 0 °C in the dark for 30 min, TLC showed completion (DCM / MeOH = 13: 1, Rf of starting material = 0.5). The mixture was concentrated, and the residue was purified by reverse phase column (H2O / CH3CN), then re-punfied by Prep-TLC (DCM / MeOH = 13:1) to afford LUX107- 1-A (16 mg, 11% yield) as hght yellow solid. LCMS (ESI): m / z 575.3 [M + H]+; HPLC: 91.7% @210 nm, Rt = 7.42 min.Example 2; Synthesis of LUX107-1PAF

[0181] Scheme 4: Synthesis of (S)-2-amino-3-(4-((E)-15-((2-((S)-l-(chloromethyl)-5-hydroxy- 9-methyl-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)oxy)-13-methyl-4,7,10- trioxa-3,13-diazapentadec-2-en-2-yl)phenyl)propanoic acid (LUX107-1PAF)

[0182] Step 1: (S)-(5-(2-((2-(2-(2-(aminooxy)ethoxy)ethoxy)ethyl)(methyl)amino)ethoxy)-lH- indol-2-yl)(l-(chloromethyl)-5-hydroxy-9-methyl-lH-benzo[e]indol-3(2H)-yl)methanone dihydrochloride (LUX107-1 HC1)

[0183] To a solution of LUX107-14B (30 mg, 42.2 pmol) in 2 mL dry DCM at 0 °C was added 4 M HCI / dioxane (25 pL. 100 pmol) under aN2 atmosphere. The reaction was stirred at 0 °C in dark for 30 min, TLC showed completion (DCM / MeOH = 13:1, Rf of starting material = 0.5).The mixture was concentrated, and the residue was slurred with cold MTBE (2 mL*2), filtrated to afford about 30 mg crude LUX107 HC1 (LUX107-1 as 2HC1 salt form,) which was used directly without further punfication.

[0184] Step 2: (S)-2-amino-3-(4-((E)-15-((2-((S)-l-(chloromethyl)-5-hydroxy-9-methyl-2,3- dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)oxy)-13-methyl-4,7,10-trioxa-3,13- diazapentadec-2-en-2-yl)phenyl)propanoic acid (LUX107-1PAF)

[0185] A mixture of the above crude LUX107-1 and para-acetyl-phenylalanine (PAF) (9 mg, 43 pmol) in 0.75 mL H2O / I.5 mL CH3CN was stirred at room temperature in dark for 16 h, LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 800.3 [M + H]+;798.0 [M']). The reaction was purified by Reverse Phase column chromatography (H2O / CH3CN) directly, and then freeze-dried to afford LUX107-1PAF (9 mg, 27% yield for 2 steps) as off- white solid. For LUX107-1PAF: LCMS (ESI): m / z 800.3 [M + H]+; HPLC: 38.5% @210 nm, Rt= 8.35 mm. For LUX107-1PAF-A: LCMS (ESI): m / z 764.3 [M + H]+; HPLC: 55.5% @210 nm, Rt = 7.36 min.

[0186] Example 3: Synthesis of LUX108-1

[0187] Scheme 5: Synthesis of 5-(4-((2,2-dimethyl-4-oxo-3,6,9,12,15,18,21-heptaoxa-5- azatricosan-23-yl)oxy)benzamido)-lH-indole-2-carboxylic acid (LUX108-10B),PEG 6 LUX108-4

[0189] To a solution of PEG 6 (47.8 g, 169.3 mmol) in DCM (950 mL) was added TsCI (19.4 g, 101.6 mmol) at room temperature, followed by the addition of TEA (47 mL, 339 mmol) and DMAP (2.1 g, 16.9 mmol). The reaction was stirred at room temperature for 16 h. LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 454.0 [M + NH?]). The mixture was concentrated under reduced pressure directly, and the residue was purified by silica column (w-heptane / EtOAc (v / v) = from 5: 1 to 1: 1; then DCM to DCM / MeOH (v / v) = from 100:1 to 60: l) to afford LUX108-4 (34.7 g, 47% yield) as light-yellow oil. LCMS (ESI):m / z 454.0 [M + NH4+]; 98% purity @210 nm ; 'H NMR (400 MHz, CDCI3) 6 7.79 (d, J= 8.3 Hz, 2H), 7.33 (d, J= 8.0 Hz, 2H), 4.20 - 4.08 (m, 2H), 3.71 - 3.66 (m, 4H), 3.66 - 3.63 (m, 8H), 3.63 - 3.60 (m, 4H), 3.60 - 3.58 (m, 2H), 3.58 - 3.56 (m, 4H), 2.44 (s, 3H).

[0190] Step 2: Tert-butyl (17-hydroxy-3,6,9,12,15-pentaoxaheptadecyl)oxycarbamate(LUX108-2B)

[0191] Step 3: Tert-butyl (17-hydroxy-3,6,9,12,15-pentaoxaheptadecyl)oxycarbamate(LUX108-2B)

[0192] To a stirred solution of BocNHOH (22.9 g, 171.8 mmol) and DBU (50 mL) was added dry THF (100 mL) under a N2 atmosphere. The mixture was stirred at room temperature for 5 min, followed by the addition of LUX108-4 (10 g, 21.9 mmol). The reaction was stirred at room temperature for 3 days. HPLC showed completion. The mixture was purified by silica column ( / ?-heptane) directly to remove DBU to obtain the crude for the first time. The crude was re- purified by silica column (w-heptane / EtOAc (v / v) = from 100: 1 to 50: 1 to 10: 1 to 1:1; then DCM to DCM / MeOH (v / v) = from 50: 1 to 30: 1) to afford impure LUX108-2B. This impure product was further purified by Reverse Phase column (0.1% TFA in H2O / CH3CN) for the third time to afford LUX108-2B (6.4 g, 74% yield) as light-yellow oil. LCMS (ESI): m / z 415.0 [M + NHL]; almost no UV absorption on HPLC ; 'H NMR (400 MHz, DMSO-d6) δ 9.97 (s, 1H), 4.59 (s, 1H), 4.11 - 4.07 (m, 1H), 3.83 - 3.77 (m, 2H), 3.66 - 3.62 (m, 1H), 3.56 - 3.51 (m, 10H), 3.51 - 3.47 (m, 6H), 3.43 - 3.39 (m, 3H), 1.40 (s, 9H).

[0193] Step 4: Ethyl 4-((2,2-dimethyl-4-oxo-3,6,9,12,15,18,21-heptaoxa-5-azatricosan-23- yl)oxy)benzoate (LUX108-6B)LUX108-6B

[0194] To a stirred solution of ethyl 4-hydroxybenzoate LUX108-5 (4.01 g, 24.15 mmol) and PI13P (12.67 g, 48.31 mmol) in dry THF (90 mL) was added DEAD (8.11 mL, 51.5 mmol) under a N2 atmosphere. The mixture was stirred at room temperature for 20 min, followed by the addition of LUX108-2B (6.4 g, 16.1 mmol). The reaction was stirred at room temperature for 2 days. LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 563.0[M + NH4+]). The mixture was concentrated, and the residue was purified by Reverse Phase column (0.1% TFA in H2O / CH3CN) twice to afford LUX108-6B (850 mg, 9.7% yield) as light- yellow oil. LCMS (ESI): m / z 563.0 [M + NH4+];JH NMR (400 MHz, CDCI3) δ 8.01 - 7.95 (m, 2H), 6.95 - 6.89 (m, 2H), 4.33 (q, J= 7.1 Hz, 2H), 4.20 - 4.15 (m, 2H), 4.03 - 3.96 (m, 2H), 3.88 - 3.85 (m, 2H), 3.76 (bs, 1H), 3.73 - 3.71 (m, 2H), 3.70 - 3.66 (m, 4H), 3.66 - 3.63 (m, 12H), 1.46 (s, 8H), 1.37 (t, J= 7.1 Hz, 3H).

[0195] Step 5: 4-((2,2-Dimethyl-4-oxo-3,6,9,12,15,18,21-heptaoxa-5-azatricosan-23- yl)oxy)benzoic acid (LUX108-7B)

[0196] To a solution of LUX108-6B (650 mg, 1.19 mmol) in 6.5 mL THF / 26 mL H2O was added LiOH*H2O (1.15 g, 27.4 mmol). The reaction was stirred at room temperature for 16 h. LCMS showed completion (expected product molecule-LCMS (ESI): m / z 535.0 [M + NH4+]). The mixture was quenched by adding TFA to adjust the pH to 5-6. The resulting mixture was concentrated, and the residue was purified by Reverse Phase column (0.1% TFA in H2O / CH3CN) to afford LUX108-7B (375 mg, 61% yield) as light-yellow oil. LCMS (ESI): m / z 535.0 [M + NH4+],

[0197] Step 6: Ethyl 5-(4-((2,2-dimethyl-4-oxo-3,6,9,12,15,18,21-heptaoxa-5-azatricosan-23- yl)oxy)benzamido)-lH-indole-2-carboxylate (LUX108-9B)

[0198] To a solution of LUX108-7B (345 mg, 667 pmol), ethyl 5-amino-lH-indole-2- carboxylate LUX108-8 (204 mg, 1.00 mmol) and HATU (634 mg, 1.67 mmol) in 4 mL DMF was added DIEA (0.35 mL, 2 mmol) under aN2 atmosphere. The reaction was stirred at room temperature for 16 h, LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 604.3 [M - 5oc]). The mixture was purified by reverse phase column (0.1% TFA inH2O:CH3CN) directly to afford LUX108-9B (190 mg, 41% yield) as brown oil. LCMS (ESI): m / z 604.3 [M - Boe],

[0199] Step 7: 5-(4-((2,2-Dimethyl-4-oxo-3,6,9,12,15,18,21-heptaoxa-5-azatricosan-23- yl)oxy)benzamido)-lH-indole-2-carboxylic acid (LUX108-10B)

[0200] To a solution of LUX108-9B (190 mg, 0.27 mmol) in 1.9 mL THF / 7.6 mL H2O was added LiOEHFEO (170 mg, 4.05 mmol). The reaction was stirred at room temperature for 16 h. LCMS showed completion (expected product molecule-LCMS (ESI): m / z 576.3 [M - 5oc]).The mixture was quenched by adding TFA to adjust the PH to 5-6. The resulting mixture was concentrated, and the residue was purified by Reverse Phase column (0.1% TFA in H2O / CH3CN) to afford LUX108-10B (150 mg, 82% yield) as red oil. LCMS (ESI): 576.3 [M - Boe],

[0201] Scheme 6: (S)-4-((17-(aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-(l- (chloromethyl)-5-hydroxy-9-methyl-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5- yl)benzamide hydrochloride (LUX108-1)

[0202] Step 1 : (S)-Tert-butyl (17-(4-((2-(l-(chloromethyl)-5-hydroxy-9-methyl-2,3-dihydro-lH- benzo|e|indole-3-carbonyl)- I H-indol-5-yl)carbamoyl)phenoxy)-3.6.9. l 2. 15- pentaoxaheptadecyl)oxy carbamate (LUX108-1 IB)

[0203] To a solution of LUX108-10B (50 mg, 74 pmol) and (5)- 1 -(chi oromethyl)-9-methy 1-2,3 - dihydro- IH-benzo [e]indol-5-ol hydrochloride LUX105-6 (31 mg, 111 pmol) in 2 mL DMF was added EDCI (42 mg, 222 pmol) under a N2 atmosphere. The reaction was stirred at room temperature in dark for 16 h, LCMS showed that the expected product molecule was formed(LCMS (ESI): m / z 805.3 [M - Boe]). The mixture was purified by reverse phase column (0.1% TFA in FECFCHsCN) firstly, then purified by prep-TLC (DCM / MeOH = 9: 1, Rf = 0.5) to afford LUX108-11B (20 mg, 48% yield) as red oil. LCMS (ESI): m / z 804.8 [M - Boe] 928.8 [M + Na+]; HPLC: 96.6% @210 nm, 7?t= 12.51 mm.

[0204] Step 2A: (S)-4-((17-(aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-(l- (chloromethyl)-5-hydroxy-9-methyl-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5- yl)benzamide, hydrochloride salt (LUX108-1 HC1)

[0205] To a solution of LUX108-11B (18 mg, 19.9 pmol) in 1.8 mL dry DCM at 0 °C was added 4 M HCl / di oxane (10 pL, 40 pmol) under a N2 atmosphere. The reaction was stirred at 0 °C in dark for 1.5 h, TLC showed completion (DCM / MeOH = 9: 1, Rf of starting material = 0.5). The mixture was concentrated, and the residue was purified by reverse phase column (pH at about 5, aqueous HC1 :CHsCN), then freeze-dried to afford LUX108-1 HC1 (4 mg, 25% yield) as off-white solid. LCMS (ESI): m / z 804.9 [M + H]+; HPLC: 92.4% @210 nm, R{= 23.4 mm.

[0206] Step 2B: (S)-4-((17-(aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-(l- (chloromethyl)-5-hydroxy-9-methyl-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5- yl)benzamide, trifluoroacetic acid salt (LUX108-1 TFA)

[0207] To a solution of LUX108-11B (20 mg, 22 pmol) in 0.8 mL dry CH3CN at 0 °C was added 0.4 mL TFA under a N2 atmosphere. The reaction was stirred at 0 °C in dark for 1.5 h, TLC showed completion (DCM / MeOH = 9: 1, Rf of starting material = 0.5). The mixture was purified by reverse phase column (0.1% TFA in FFOiCFLCN) directly, then freeze-dried to afford LUX108-1 TFA (16 mg, 80% yield) as bright yellow solid. LCMS (ESI): m / z 805.3 [M + H]+, 827.3 [M + Na+]; HPLC: 92.3% @210 nm, Rt= 10.6 mm; 1H NMR (400 MHz, DMSO- d6) 8 11.73 (s, 1H), 10.48 (s, 4H), 10.06 (s, 1H), 8.17 (s, 1H), 8.05 (d, J= 8.3 Hz, 1H), 8.04 - 7.94 (m, 3H), 7.56 (d, J= 9.0 Hz, 1H), 7.46 (d, J= 8.9 Hz, 1H), 7.34 (d, J= 6.8 Hz, 1H), 7.27 - 7.22 (m, 1H), 7.19 (s, 1H), 7.08 (d, J= 8.6 Hz, 2H), 4.75 - 4.66 (m, 1H), 4.61 - 4.54 (m, 1H), 4.32 (t, J= 8.1 Hz, 1H), 4.22 - 4.17 (m, 2H), 4.09 - 4.06 (m, 2H), 3.86 - 3.75 (m, 4H), 3.64 - 3.59 (m, 4H), 3.58 - 3.54 (m, 2H), 3.54 - 3.47 (m, 13H), 2.79 (s, 3H);19F NMR (400 MHz, DMSO-@) 8 74.03.

[0208] Example 4; Synthesis of LUX108-1PAF

[0209] Scheme 7: Synthesis of (S)-2-amino-3-(4-((E)-21-(4-((2-((S)-l-(chloromethyl)-5- hydroxy-9-methyl-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5- yl)carbamoyl)phenoxy )-4,7, 10, 13 , 16, 19-hexaoxa-3 -azahenicos-2-en-2-yl)pheny l)propanoic acid (LUX108-1PAF)

[0210] Step 1 : (S)-4-((17-(aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-(l-(chloromethyl)-5-hydroxy-9-methyl-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5- yl)benzamide, hydrochloride (LUX108-1 HC1)

[0211] To a solution of LUX108-11B (15 mg, 16.6 pmol) in 1 mL dry DCM at 0 °C was added 4 M HCl / dioxane (10 pL, 40 pmol) under a N2 atmosphere. The reaction was stirred at 0 °C in dark for 1.5 h, LCMS showed completion. The mixture was concentrated, and the residue was slurred with cold MTBE (2 mLx2), filtrated and concentrated to dry to afford crude LUX108-1 HC1, which was used directly without further purification. LCMS (ESI): m / z 805.3 [M + H]+.

[0212] Step 2: (S)-2-amino-3-(4-((E)-21-(4-((2-((S)-l-(chloromethyl)-5-hydroxy-9-methyl-2,3- dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)carbamoyl)phenoxy)-4,7,10,13,16,19- hexaoxa-3-azahenicos-2-en-2-yl)phenyl)propanoic acid (LUX108-1PAF)

[0213] A mixture of the above crude LUX108-1 HC1 and PAF (4 mg, 20 pmol) in 0.65 mL H2O / I.3 mL CH3CN was stirred at room temperature in dark for 16 h, LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 1016.2 [M + Na+]). The reaction mixture was purified by Reverse Phase column (H2O / CH3CN) directly, and then freeze-dried to afford LUX108-1PAF (4 mg, 24% yield for 2 steps) as white foam solid. For LUX108-1PAF: LCMS (ESI): m / z 1016.3 [M + Na+], 497.9 [M / 2 + 1]+; HPLC: 92.8% @210 nm, Rt= 9.88 min. For LUX108-1PAF-A: LCMS (ESI): m / z 479.5 [M / 2 + 1]+; HPLC: 6.1% @210 nm, Rt= 8.45 min.

[0214] Example 5; Synthesis of LUX108-2 HC1

[0215] Scheme 8: Synthetic Route A for 4-((17-(aminooxy)-3, 6, 9, 12,15- pentaoxaheptadecyl)oxy)-N-(2-((S)-l-(chloromethyl)-9-methyl-5-(((2S, 3R, 4S,5R, 67?)-3,4,5- trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3- carbonyl)-lH-indol-5-yl)benzamide, hydrochloride (LUX108-2 HC1)

[0216] Step 1 : (2i?,3S,4S,5A, 6S)-2-(acetoxymethyl)-6-(((S)-3-(5-(4-((17-(aminooxy)- 3,6,9,12,15-pentaoxaheptadecyl)oxy)benzamido)-lH-indole-2-carbonyl)-l-(chloromethyl)-9- methyl-2,3-dihydro-lH-benzo[e]indol-5-yl)oxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (LUX108-12B)

[0217] To a solution of LUX108-11B (40 mg, 44 pmol) and (2R,3S,4S,5R,6R)-2- (acetoxymethyl)-6- (2.2.2-trichloro- l -iminoethoxy)tetrahydro-2 / / -pyran-3.4.5-tnyl triacetate LUX107-15 (32 mg, 66 pmol) in dry DCM (8 mL) was added activated 3A MS (200 mg) under an argon atmosphere. The mixture was stirred at room temperature for 20 min and then cooled to 0 °C with an ice-bath. To the resulting solution, BFs’OEt (22 mg, 154 pmol) was added. The reaction was allowed to warm up to room temperature naturally and stirred for 16 h. HPLCshowed the starting material was consumed with only LUX108-1 detected. The mixture was filtrated, and concentrated. The residue was purified by Reverse Phase column (H2O / CH3CN) to afford 25 mg LUX108-1. HPLC: 78.4% @210 nm, 7?t= 23.3 min.

[0218] To a solution of the above purified 25 mg LUX108-1 in dry DCM (6 mL) were added (2R ,35,45,5R ,6R )-2-(acetoxymethyl)-6-(2,2,2-trichloro-l-iminoethoxy)tetrahydro-2F7-pyran- 3,4,5-triyl triacetate LUX107-15 (20 mg, 41 pmol) and activated 4A MS (150 mg) under an argon atmosphere. The mixture was stirred at room temperature for 20 min and then cooled to 0 °C with an ice-bath. To the resulting solution, BF3«OEt (15 mg, 105 pmol) was added. The reaction was allowed to warm up to room temperature and stirred for 16 h. LCMS showed targeted molecule formed (LCMS (ESI): m / z 1135.8 [M + H]+). The mixture was filtrated, and concentrated. The residue was purified by Reverse Phase column (H2O / CH3CN) for the first time, and then re-purified by prep-TLC (DCM: MeOH = 9: 1, v / v; Rf = 0.4) to afford LUX108- 12B (9 mg, 18% yield) as grey solid. LCMS (ESI): m / z 1135.7 [M + H]+; HPLC: 92.4% @210 nm, R[ - 12.15 min.

[0219] Step 2: 4-((17-(Aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-((5)-l-(chloromethy l)-9-methy l-5-(((2S', 3R, 4S, 5R, 6R )-3.4.5-trihydroxy-6-(hydroxy methyl )tetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)benzamide, hydrochloride (LUX108-2 HC1)

[0220] To a solution of LUX108-12B (9 mg, 44 pmol ) in dry MeOH (2.9 mL) at 0 °C was added 10% MeONa / MeOH (51 mg, 95 pmol). The mixture was stirred at 0 °C for 30 min. LCMS showed completion (LCMS (ESI): m / z 966.7 [M + H]+). The reaction was quenched by adding 4 M HCl / dioxane to adjust the pH to from about 6 to about 7. The resulting solution was purified by Reverse Phase column (about pH 4 HC1 aq. / CH3CN) directly to afford LUX108-2 HC1 (4 mg, 52% yield) as yellow solid. LCMS (ESI): m / z 968,9 [M (37C1) + H]+; HPLC: 94.1% @210 nm, Rt = 9.50 min.

[0221] Example 6; Synthesis of LUX108-2

[0222] Scheme 9: Synthetic Route B for 4-((17-(aminooxy)-3,6,9,12,15- pentaoxaheptadecyl)oxy)-N-(2-((S)-l-(chloromethyl)-9-methyl-5-(((2S, 3R, -IS.5R.6R )-3.4.5- trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3- carbonyl)-lH-indol-5-yl)benzamide (LUX108-2)

[0223] Step 1 : (2R,3S,4S,5R, 6SS-2-(acetoxymethyl)-6-(((<S)-l-(chloromethyl)-3-(5-(4-((2,2- dimethyl-4-oxo-3,6,9, 12, 15, 18,21-heptaoxa-5-azatricosan-23-yl)oxy)benzamido)-lH-indole-2- carbonyl)-9-methyl-2,3-dihydro-lH-benzo[e]indol-5-yl)oxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (LUX108-12C)

[0224] To a solution of the crude LUX108-15 (460 mg, 802 pmol) and LUX108-10B (200 mg, 296 pmol) in 5 mL DMF was added EDCI (307 mg, 1.62 mmol) under aN2 atmosphere. The reaction was stirred at room temperature for 16 h, LCMS showed that the expected product molecule was formed (LCMS (ESI): m / z 1135.3 [M - Boc] observed). The mixture was purified by reverse phase column (0.1% TFA in FEOCFLCN) firstly, then purified by prep-TLC (DCM / MeOH = 15:1, Rf= 0.5), and finally punfied by prep-TLC (DCM / THF = 2: 1, Rf= 0.5) to afford LUX108-12C (133 mg, 48% yield) as yellow solid. LCMS (ESI): m / z 1136.2 [M - 5oc]; HPLC: 96.5% @210 nm, Rt= 23.74 min.

[0225] Step 2: (2R ,3S,4S,5R , 6S)-2-(acetoxymethyl)-6-(((S)-3-(5-(4-((17-(aminooxy)- 3,6,9,12,15-pentaoxaheptadecyl)oxy)benzamido)-lH-indole-2-carbonyl)-l-(chloromethyl)-9- methyl-2,3-dihydro-lH-benzo[e]indol-5-yl)oxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate, 2,2,2- trifluoroacetate salt (LUX108-12B TFA)

[0226] To a solution of LUX108-12B (100 mg, 80.9 pmol) in 3 mL dry CH3CN at 0 °C was added 2 mL TFA under a N2 atmosphere. The reaction was stirred at 0 °C for 2 h, TLC & HPLC showed completion (DCM / MeOH = 15:1, Rf of starting material = 0.55). The mixture was purified by reverse phase column (0.1% TFA in FLCkCFLCN) firstly to afford 60 mg, 74% purity LUX108-12B , then purified by prep-TLC (DCM: MeOH = 9:1, v / v; Rf = 0.4) to afford LUX108-12B TFA (40 mg, 40% yield) as bright yellow solid. LCMS (ESI): m / z 1136.1 [M + H]+; HPLC: 91.8% @210 nm, Rt= 10.6 min.

[0227] Step 3: 4-((17-(Aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-((S)-l- (chloromethy l)-9-methy l-5-(((2S.3R.4S, 5R, 6R )-3.4.5-trihydroxy-6-(hydroxymethyl)tetrahydro- 2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)benzamide (LUX108-2)

[0228] To a solution of LUX108-12B TFA (15 mg, 73 gmol) in dry MeOH (3.9 mL) at 0 °C was added 10% MeONa / MeOH (100 mg, 190 nmol). The mixture was stirred at 0 °C for 30 min. TLC showed completion (DCM / MeOH = 9: 1, Rf of starting material = 0.4). The reaction was quenched by adding TFA to adjust the pH to from about 6 to about 7. The resulting solution was purified by Reverse Phase column (H2O / CH3CN) directly to afford LUX108-2 (7 mg, 60% yield) as white foam solid. LCMS (ESI): m / z 969.4 [M (37C1) + H]+; HPLC: 96.2% @210 nm, Rt = 9.52 mm; 'H NMR (400 MHz, DMSO) 8 11.80 (s, 1H), 10.42 (s, 2H), 10.05 (s, 1H), 8.38 - 8.24 (m, 2H), 8.18 (s, 1H), 7.99 (d, J= 8.6 Hz, 2H), 7.55 (d, J= 9.9 Hz, 1H), 7.47 (d, J= 8.8 Hz, 1H), 7.40 (d, 6.8 Hz, 1H), 7.35 - 7.28 (m, 1H), 7.20 (s, 1H), 7.08 (d, J= 8.7 Hz, 2H),4.93 (d, J= 7.7 Hz, 1H), 4.79 - 4.71 (m, 1H), 4.61 (d, J= 10.6 Hz, 1H), 4.38 (t, J= 7.6 Hz, 1H), 4.23 - 4.17 (m, 2H), 4.07 - 4.03 (m, 2H), 3.88 - 3.73 (m, 6H), 3.68 - 3.65 (m, 1H), 3.63 - 3.60 (m, 4H), 3.58 - 3.55 (m, 4H), 3.53 - 3.51 (m, 12H), 2.82 (s, 3H).

[0229] Example 7; Synthesis of LUX108-2

[0230] Scheme 10: Synthetic Route C for 4-((17-(aminooxy)-3,6,9,12,15- pentaoxaheptadecyl)oxy)-N-(2-((5)-l-(chloromethyl)-9-methyl-5-(((2S, 3R, -IS.5R.6R )-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3- carbonyl)-lH-indol-5-yl)benzamide, hydrochloride salt (LUX108-2 HC1)

[0231] Step 1: Tert-butyl (17-(4-((2-((5)-l-(chloromethyl)-9-methyl-5-(((25,3R ,4S 5R,6R )-3,4,5- trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3- carbonyl)- lH-indol-5-yl)carbamoyl)phenoxy)-3, 6, 9, 12, 15-pentaoxaheptadecyl)oxy carbamate (LUX108-13B)

[0232] To a solution of LUX108-12C (15 mg, 13 pmol) in dry MeOH (5 mL) at 0 °C was added 10% MeONa / MeOH (16 mg, 30 pmol). The mixture was stirred at 0 °C for 30 min. LCMS showed completion (LCMS (ESI): 969.0 [M (37C1) - Boe]). The reaction was quenched by adding 0. 1 M HCl / MeOH to adjust the pH to from about 6 to about 7, then concentrated under reduced pressure to remove the solvent to about 1 mL. The resulting solution was purified by Reverse Phase column (H2O / CH3CN) to afford LUX108-13B (8 mg, 62% yield) as white foam solid. LCMS (ESI): m / z 967.3 [M - Boc]; HPLC: 92.2% @210 nm, Rt= 10.57 min; 'H NMR (400 MHz, DMSO) 8 11.80 (s, 1H), 10.05 (s, 1H), 9.99 (s, 1H), 8.34 - 8.24 (m, 2H), 8.18 (s, 1H), 7.98 (d, J= 8.5 Hz, 2H), 7.55 (d, J= 9.0 Hz, 1H), 7.46 (d, J= 8.8 Hz, 1H), 7.40 (d, J= 7.1 Hz, 1H), 7.34 - 7.28 (m, 1H), 7.21 (s, 1H), 7.08 (d, J= 8.6 Hz, 2H), 5.35 (d, J= 5.2 Hz, 1H), 4.93 (d, J= 6.9 Hz, 2H), 4.79 - 4.70 (m, 1H), 4.69 - 4.55 (m, 3H), 4.42 - 4.35 (m, 1H), 4.24 - 4.14 (m, 2H), 3.86 - 3.76 (m, 6H), 3.69 - 3.63 (m, 1H), 3.63 - 3.58 (m, 3H), 3.58 - 3.54 (m, 4H), 3.54 - 3.46 (m, 12H), 2.82 (s, 3H), 1.39 (s, 9H).

[0233] Step 2: 4-((17-(Aminooxy)-3,6,9,12,15-pentaoxaheptadecyl)oxy)-N-(2-((S)-l- (chloromethy l)-9-methy l-5-(((2S', 3R, 4S, 5R 6 / ?)-3.4.5-trihydroxy-6-(hydroxy methyl jtetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)benzamide, hydrochloride salt (LUX108-2 HC1)

[0234] To a solution of LUX108-13B (6 mg in 0.5 mL DMSO-rfe, 22 pmol) in 1.5 mL dry DCM at 0 °C was added 0.5 mL 4 M HCl / di oxane under a N2 atmosphere. The reaction was stirred at 0 °C for 2 h. LCMS showed completion with no desired product molecule detected. The reaction was concentrated, and the residue was purified by Reverse Phase column (H2O / CH3CN) to afford one major peak LCMS (ESI): m / z 839 / 840.

[0235] Example 8: Alternative Synthesis of LUX108-13B

[0236] Scheme 1: Synthesis of Tert-butyl (17-(4-((2-((5)-l-(chloromethyl)-9-methyl-5-(((25, 3R.4S,5R, 6R )-3.4.5-lnhydroxy-6-(hydroxymelhyl)letrahydro-2H-pyran-2-yl)oxy)-2.3- dihydro-lH-benzo[e]indole-3-carbonyl)-lH-indol-5-yl)carbamoyl)phenoxy)-3,6,9,12,15- pentaoxaheptadecyljoxy carbamate (LUX108-13B)

[0237] Step 1 : (2S, 3R, 4S,5R, 6 / ?)-2-((fS')- l -(chloromethyl)-9-methyl-2, 3 -dihydro- 1H- benzo[e]indol-5-yl)oxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (LUX108-16)

[0238] To a solution of LUX108-15 (20 mg, 35 pmol) in dry MeOH (5 mL) at 0 °C was added 10% MeONa / MeOH (38 mg, 70 pmol). The mixture was stirred at 0 °C for 30 min. LCMS showed completion (LCMS (ESI): m / z 374.3 [M - Cl]). The reaction was quenched by adding 0. 1 M HCl / MeOH to adjust the pH to from about 6 to about 7, then concentrated under reduced pressure to dry. The residue was slurred with ^-heptane, filtrated to afford 20 mg crude LUX108-16, which was used directly without further purification.

[0239] Step 2: Tert-butyl (17-(4-((2-((5)-l-(chloromethyl)-9-methyl-5-(((2S, 3R, 4S,5R, 6R)-3.4,5- trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-2,3-dihydro-lH-benzo[e]indole-3-carbonyl)- lH-indol-5-yl)carbamoyl)phenoxy)-3, 6, 9, 12, 15-pentaoxaheptadecyl)oxy carbamate(LUX108-13B)

[0240] To a solution of the above 20 mg crude LUX108-16 and LUX108-10B (18 mg, 27 pmol) in 1 mL DMF was added EDCI (12 mg, 63 pmol) under a N2 atmosphere. The reaction was stirred at room temperature for 16 h, LCMS showed that the target material was formed. The mixture was purified by reverse phase column (JEOiCHsCN) firstly, then purified by prep-TLC (DCM / MeOH = 5: 1, Rf= 0.45) to afford LUX108-13B (8 mg, 27% yield) as grey solid. LCMS (ESI): m / z 968.3 [M - Boc],Example 9: Biological Assays

[0241] Materials and Methods

[0242] The human prostate cancer cell lines DU145 and PC-3 were purchased from American Type Culture Collection (HTB81 and CRL1435) and maintained according to ATCC recommendations.

[0243] Enzyme: |3-D-galactosidase (E.C. 3.2.1.23) von Escherichia coli G 5635 (Sigma- Aldrich, G5635); activity: 250-600 units per mg of protein at pH = 7.3 and 37 °C; 1 unit = conversion of 1 pmol substrate per min.

[0244] Measurement of Cell Viability

[0245] Cell proliferation was measured by the CellTiter-Glo® (CTG) viability assay. Prostate cancer (PCa) cells (e.g., DU145) were seeded in 96 well plates (e.g., about 10,000 cells per well) and treated for 72 h with various concentrations of testing compounds (Table 1) at the indicated concentrations untreated (in the absence of bgal) or pretreated for 2 hours with 0-D- galactosidase (in the presence of bgal at 2 unit / mL, 4 unit / mL or 6 unit / mL) in media to release the cytotoxic agents from the prodrug. Then cell viability was determined using the CellTiter- Glo® luminescent assay (Promega, Madison, WI, USA), according to the manufacturer’s instructions (CellTiter-Glo® Luminescent Cell Viability Assay, Technical Bulletin, TB 288 Promega). Luminescence for determination of cell viability (expressed in percentages) was measured using the Spectramax iD3 microplate reader. IC50 were calculated using GraphPad Prism version 5.00 (GraphPad Software, San Diego, CA, USA). Dose response curves were generated to calculate the half-maximal inhibitory concentration (IC50) for each testing compound. See FIG. 1 and FIG. 2, Table 2 and Table 3.

[0246] Table 1. Testing compounds.

[0247] Cell viability results

[0248] Table 2. Testing compounds untreated

[0249] Table 3. Testing compounds pretreated with |3-D-galactosidase:

[0250] As shown in FIG. 1 and Table 2, Comp A3 and Comp A4 exhibit sub-nanomolar IC50 values in the cell viability assay. In contrast, the prodrug Comp A5 exhibits much weaker activity (347 nM) in the same cell viability assay, demonstrating that the free hydroxy is critical to the cy totoxic agent.

[0251] As shown in FIG. 2 and Table 3, pretreatment with P-D-galactosidase with Comp A5 increases the potency of the prodrug Comp A5. When comparing the results of Experiment # 1 and Experiment #3 in Table 3, the presence of 4 unit / mL of galactosidase increases the cytotoxicity potency of Comp A5 by more than 60-fold. From Experiment 2 to 4, the appeared cytotoxic activity increase with the increase of P-D-galactosidase concentration. One possible explanation for the increased cytotoxicity of Comp A5 in the presence of P-D-galactosidase is that the galactosidase removes the galactose protecting group on the hydroxy group, thereby converting the prodrug Comp A5 to the parent drug Comp A4. In control experiments (Experiment #5 and Experiment #6), the presence of 4 unit / mL of galactosidase increases the cytotoxicity potency of Comp A3 by about 2-fold, far less than the 60-fold increase for Comp A5 with the same amount of enzy me.

[0252] Without limiting the scope of the present disclosure, FIG. 3 shows a possible mechanism to explain the prodrug transformation for the prodrug conjugated to a targeting moiety.

[0253] As shown in FIG. 3, the prodrug of a compound disclosed herein can conjugate to a targeting moiety via the reactive group Rc to afford a conjugated prodrug. Note: in FIG. 3, Rc in the conjugate represents a derivative of the reactive group after conjugating the prodrug with the targeting moiety'. The conjugated prodrug is non-cytotoxic due to the mask of the free hydroxygroup by an R1protecting group. The targeting moiety can be an antibody, a peptide, a small molecule, etc., that can guide the conjugated prodrug through a trafficking mechanism to internalize into the target cell. Upon internalization, an intercellular complex comprising the internalized conjugated prodrug can undergo a few conversions: (1) the R1protecting group on the prodrug can be cleaved by an enzyme in or around the target cell; and (2) an intra-cellular degradation can digest the targeting moiety, be it an antibody, a peptide, or a small molecule. The end results of these intra-cellular conversions can be the formation of either converted drug A or converted drug B. Converted drug A comprises both the released and activated cytotoxic pay load and an AA moiety connected to the Linker-Rc group. AA as used herein refers to (1) a natural or unnatural ammo acid used for antibody conjugation or peptide conjugation or (2) a modified small molecule when the targeting moiety employs a small molecule. Converted drug B comprises both the released / activated cytotoxic payload and a partial linker. The partial linker can be a product of the intra-cellular degradation. Converted drug A and converted drug B can be cytotoxic to the target cells. The small molecule as a targeting moiety can target or bind to

[0254] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. It is not intended that the invention be limited by the specific examples provided within the specification. While the invention has been described with reference to the aforementioned specification, the descriptions and illustrations of the embodiments herein are not meant to be construed in a limiting sense. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. Furthermore, it shall be understood that all aspects of the invention are not limited to the specific depictions, configurations or relative proportions set forth herein which depend upon a variety of conditions and variables. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention. It is therefore contemplated that the invention shall also cover any such alternatives, modifications, variations or equivalents. It is intended that the following claims define the scope of the invention and that methods and structures within the scope of these claims and their equivalents be covered thereby.

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A compound of Formula I:Formula I' or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:Rc is a reactive group;Linker is a cleavable linker or a non-cleavable linker;R2is H or CH3; andR3is H, OH, or OCH3.

2. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H or CH3; andR3is H, OH, or OCH3.

3. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is H.

4. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is OH.

5. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is OCH3.

6. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is H.

7. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is OH.

8. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is OCH3.

9. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is H.

10. The compound of any one of claims 1 to 9, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:LG is halide, nonaflate, inflate, fluorosulfonate, tosylate, mesylate, or besylate;RAis independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl- alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxy carbonyl, or alkylsulfonyl;RBis independently a protecting group, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each of R6, R7and R8is independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each R9is independently halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, C1-C4 heteroalkyl, -C(O)H, -C(O)OH, -CN, C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, 5- to 10-membered heteroaryl, - C(O)(C1-C4alkyl), -C(O)O(C1-C4 alkyl), -C(O)NH2, -C(O)NH(C1-C4 alkyl), -C(O)N(C1-C4 alkyl)2, -NH2, -NH(C1-C4 alkyl), -N(C1-C4 alkyl)2, -NH(C2-C4alkylene)-OH, -NH(C2-C4alkylene)-O-(C1-C4 alkyl), -OH, -O(C1-C4 alkyl), -O(C1-C4 haloalkyl), -O(C2-C4alkylene)- NH2, -O(C2-C4alkylene)-NH-(C1-C4 alkyl), -O(C2-C4alkylene)-N-(C1-C4 alkyl)2, -O(C1-C4 alkylene)-C(O)OH, -O(C1-C4 alkylene)-C(O)O-(C1-C4 alkyl), -O(C2-C4alkenyl), -O(C1-C4 alkylene)-(C6-C10 aryl), -O(C1-C4 alkylene)-(5- to 10-membered heteroaryl), 0(C6-Cio aryl), > SH, S(O)2OH, -S(O)2(C1-C4 alkyl), -S(O)2NH2, -S(O)2NH(C1-C4 alkyl), or -S(0)2N(C1-C4alky 1)2; or two R9, together with atoms to which they are attached, form a C3-C10 cycloalkyl or a 3- to 10-membered heterocycloalkyl ring; and each of p and q is independently an integer of 0-3.

11. The compound of any one of claims 1 to 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:Linker is -(L^r-CL^s-*;* denotes a connection to Rc; each L1and L2is independently a bond, -O-, -S-, -NH-, -NRD-, -C(=O)-, -C(=O)O- -OC(=O)-, -C(=O)NH- -NHC(=O)-, -C(=O)NRD-, -NRDC(=O)-, -(CH2-O-CH2)m-, Ci-C6alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, -C6-C10 aryl-, or 5- to 10-membered heteroarylene, wherein each C1-C6 alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, - C6-C10 aryl-, and 5- to 10-membered heteroarylene is optionally substituted with 1, 2, or 3 R9;RDis independently -CR3R4R5; each R3, R4, and R5is independently hydrogen, halogen, -U, or -G;— U is independently C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2- C6 alkenyl, or C2-C6 alkynyl; wherein each C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl is optionally substituted with 1, 2, or 3 R9and / or 1 or 2 -G;-G is independently C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl; wherein each C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl is optionally substituted with 1, 2, or 3 R9; each R9is independently as defined in claim 1; and each of m, r and s is independently an integer of 1-12.

12. The compound of any one of claims 1 to 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:Linker comprisescombination thereof;RAis independently as defined in claim 2; n is independently an integer of 1-8; and# denotes a connection to Rc or a group connected to Rc.

13. The compound of claims 1, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas II, II’, III, III’, IV or IV’:R2is H or CH3;R3is H, OH, or OCH3;LG is halide, nonaflate, triflate, fluorosulfonate, tosylate, mesylate, or besylate;RAis independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl- alkylene, alkylamino-alkylene, alkydthio-alkylene, alkylcarbonyl, alkoxy carbonyl, or alkylsulfonyl;RBis independently a protecting group, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each of R6, R7and R8is independently hydrogen, C1-C6 alkyl, cycloalkyl-alkylene, C1-C6 haloalkyl, heteroalkyl-alkylene, heteroaryl-alkylene, heterocycloalkyl-alkylene, aryl-alkylene, heteroaryl-alkylene, alkylamino-alkylene, alkylthio-alkylene, alkylcarbonyl, alkoxycarbonyl, or alkylsulfonyl; each R9is independently halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C1-C4 haloalkyl, C1-C4 alkoxy, C1-C4 haloalkoxy, C1-C4 heteroalkyl, -C(O)H, -C(O)OH, -CN, C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyd, C6-C10 aryl, 5- to 10-membered heteroaryl, -C(O)(C1-C4 alkyl), -C(O)O(C1-C4 alkyl), -C(O)NH2, -C(O)NH(C1-C4 alkyl), -C(O)N(C1-C4 alkyl)2, -NH2, -NH(C1-C4 alkyl), -N(C1-C4 alkyl)2, -NH(C2-C4alkylene)-OH, -NH(C2-C4alkylene)-O-(C1-C4 alkyl), -OH, -O(C1-C4 alkyl), -O(C1-C4 haloalkyl), -O(C2-C4alkylene)- NH2, -O(C2-C4alkylene)-NH-(C1-C4 alkyl), -O(C2-C4alkylene)-N-(C1-C4 alkyl)2, -O(C1-C4 alkylene)-C(O)OH, -O(C1-C4 alkylene)-C(O)O-(C1-C4 alkyl), -O(C2-C4alkenyl), -O(C1-C4 alkylene)-(C6-C10 aryl), -O(C1-C4 alkylene)-(5- to 10-membered heteroaryl), 0(C6-Cio aryl), > SH, S(O)2OH, -S(O)2(C1-C4 alkyl), -S(O)2NH2, -S(O)2NH(C1-C4 alkyl), or-S(O)2N(C1-C4 alky 1)2; or two R9, together with atoms to which they are attached, form a C3-C10 cycloalkyl or a 3- to 10-membered heterocycloalkyl ring; each L1and L2is independently a bond, -O-, -S-, -NH-, -NRD-, -C(=O)-, -C(=O)O- -OC(=O)-, -C(=O)NH-, -NHC(=O)-, -C(=O)NRD-, -NRDC(=O)-, -(CH2-O-CH2)m-, Ci-C6alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, -C6-C10 aryl-, or 5- to 10-membered heteroarylene, wherein each C1-C6 alkylene, C1-C6 haloalkylene, -C1-C6 alkoxy-, C1-C6 heteroalkylene, C2-C6 alkenylene, C2-C6 alkynylene, alkylamino, alkylthio, alkylcarbonyl, alkoxy carbonyl, alkylsulfonyl, C3-C10 cycloalkylene, 3- to 10-membered heterocycloalkylene, - C6-C10 aryl-, and 5- to 10-membered heteroarylene is optionally substituted with 1, 2, or 3 R9;RDis independently -CR3R4R5; each R3, R4, and R5is independently hydrogen, halogen, -U, or -G;— U is independently C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2- C6 alkenyl, or C2-C6 alkynyl; wherein each C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 heteroalkyl, C2-C6 alkenyl, or C2-C6 alkynyl is optionally substituted with 1, 2, or 3 R9and / or 1 or 2 -G;-G is independently C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 aryl, or 5- to 10-membered heteroaryl; wherein each C3-C10 cycloalkyl, 3- to 10-membered heterocycloalkyl, C6-C10 ary l . or 5- to 10-membered heteroaryl is optionally substituted with 1, 2, or 3 R9; m is independently an integer of 1-12; each of p and q is independently an integer of 0-3; n is independently an integer of 0-8; and each t and u is independently an integer of 1-4.

14. The compound of claim 13, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas Ila, II’ a, Illa, III’ a„ IV a or IV’ a:wherein:R1is independently as defined in claim 13;R2is H or CH3;R3is H, OH, or OCH3; each of RAand Rc is independently as defined in claim 13;L1is independently as defined in claim 13; n is independently an integer of 0-8; and u is independently an integer of 1-4.

15. The compound of claim 13, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas lib, Il’b, Illb, Ill’b, IVb or IV’b:wherein:R1is independently as defined in claim 13;R2is H or CH3;R3is H, OH, or OCH3; each of RAand Rc is independently as defined in claim 13; m is independently an integer of 1-8; and n is an integer of 0-8.

16. The compound of claim 15, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is according to Formulas lie, II’ c, IIIc, life, IVc or lV’c:Formula ll'cFormula IVc , orFormula IV'c wherein:Rc is independently as defined in claim 13;R2is H or CH3;R3is H, OH, or OCH3; x is an integer of 0-4; and y is independently an integer of 1-8.

17. The compound of any one of claims 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is H.

18. The compound of any one of claims 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is OH.

19. The compound of any one of claims 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is CH3; andR3is OCH3.

20. The compound of any one of claims 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is H.

21. The compound of any one of claims 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is OH.

22. The compound of any one of claims 13-16 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein:R2is H; andR3is OCH3.

23. The compound of claim 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is:wherein:R1is as defined in claim 1; andRc is as defined in claim 10.

24. The compound of any one of claims 1 or 3 to 23, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein R1is:

25. The compound of claim 10, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the compound is:wherein:Rc is as defined in claim 10.

26. The compound of any one of claims 1 to 25, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein Rc is:

27. The compound of claim 26, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein Rc is:

28. A pharmaceutical composition comprising a compound of any one of claims 1 to 27, or pharmaceutically acceptable salt, solvate, diastereomeric mixture, or individual enantiomers thereof, and a pharmaceutically acceptable carrier.

29. A method of treating a cancer in a mammal suffering therefrom, comprising administering to the mammal a therapeutically effective amount of a compound of any one of claims 1 to 27 or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, or a pharmaceutical composition of claim 28.

30. A conjugate comprising a cell surface targeting agent or a long-acting reagent, wherein the cell surface targeting agent or the long-acting reagent is attached to a compound of any one of claims 1 to 27 or a derivative of a compound of any one of claims 1 to 27, or apharmaceutically acceptable salt, solvate, prodrug, stereoisomer or tautomer thereof, wherein the cell surface targeting agent is a small molecule, a polypeptide, or an antibody or an antigen- binding portion thereof, and wherein the long-acting reagent comprising a polymer, human serum albumin (HSA), or a nanoparticle.

31. The conjugate of claim 30, wherein the cell surface targeting agent is a monoclonal antibody, a Fab, a Fab’, a F(ab’), a Fv, a disulfide linked Fc, a scFv, a single domain antibody, a diabody, a bi-specific antibody, or a multi-specific antibody.

32. The conjugate of claim 20, wherein the cell surface targeting agent binds to a target molecule.

33. The conjugate of claim 32, wherein the target molecule comprises a tumor suppressor, a metabolic enzyme, a protein aggregate, or a haploinsufficient protein.

34. The conjugate of claim 32, wherein the target molecule comprises p53 mutant or Von Hippel-Lindau tumor suppressor (VHL).

35. A pharmaceutical composition comprising a conjugate of any one of claims 30-34, and a pharmaceutically acceptable carrier.

36. A method of treating a cancer in a mammal suffering therefrom, comprising administering to the mammal a therapeutically effective amount of a conjugate of any one of claims 30-34, or a pharmaceutical composition of claim 35, wherein the conjugate binds to a target antigen associated with the cancer.

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