Compounds for treating kinase-dependent disorders

Abstract

To provide a compound for modulating protein kinase enzymatic activity.SOLUTION: The present invention provides a compound of formula (I). (R1: -H, -CN, -CO-NR5R6 or the like. R2: -H, halo, NR5R6 or the like. R3: H, optionally substituted C1-C6 alkyl or the like. R4: H, halo. R5, R6: H, optionally substituted C1-C6 alkyl or the like. Y: -O-, -S-, -SO-, -SO2-, -NH-, and -N((C1-C6) alkyl)-).SELECTED DRAWING: None

Description

Technical Field

[0001] The present invention relates to compounds that regulate cell activities such as proliferation, differentiation, programmed cell death, migration, and chemotaxis by modulating protein kinase enzyme activity. Even more specifically, the present invention relates to compounds that inhibit, control, and / or modulate Axl and Mer receptor tyrosine kinases, compositions containing these compounds, methods of using them for treating kinase-dependent diseases and conditions, the synthesis of these compounds, and the process of formulating these compounds for pharmaceutical purposes.

[0002] Cross - reference to related applications This application claims priority to U.S. Provisional Patent Application No. 62 / 622,702, filed on January 26, 2018, and U.S. Provisional Patent Application No. 62 / 758,321, filed on November 9, 2018, the entire contents of which are incorporated herein by reference.

Background Art

[0003] Human Axl belongs to the TAM subfamily of receptor tyrosine kinases, including Mer. TAM kinases are characterized by an extracellular ligand - binding domain consisting of two immunoglobulin - like domains and two fibronectin type III domains. Axl is overexpressed in several tumor cell types and was initially cloned from patients with chronic myeloid leukemia. When overexpressed, Axl exhibits transforming ability. Axl signaling is thought to induce tumor growth through the activation of proliferative and anti - apoptotic signaling pathways. Axl is associated with cancers such as lung cancer, myeloid leukemia, uterine cancer, ovarian cancer, glioma, melanoma, thyroid cancer, renal cell carcinoma, osteosarcoma, gastric cancer, prostate cancer, and breast cancer. Overexpression of Axl results in poor prognosis for patients with the indicated cancers.

[0004] Activation of Mer, like Axl, transmits downstream signaling pathways that induce tumor growth and activation. Mer binds to ligands such as the soluble protein Gas-6. Gas-6 binding to Mer induces auto-phosphorylation of Mer in its intracellular domain, leading to downstream signal activation. Overexpression of Mer in cancer cells likely leads to increased metastasis through the production of soluble Mer extracellular domain proteins as decoy receptors. Tumor cells secrete soluble forms of extracellular Mer receptors that reduce the ability of the soluble Gas-6 ligand to activate Mer on endothelial cells, resulting in cancer progression.

[0005] Therefore, compounds that inhibit TAM receptor tyrosine kinases such as Axl and Mer are needed to treat selected cancers.

SUMMARY OF THE INVENTION

MEANS FOR SOLVING THE PROBLEM

[0006] In one aspect, the present invention provides a compound of formula I:

Chemical formula

Chemical formula

[0007] Another aspect provides a compound of formula A:

Chemical formula

[0008] Another aspect provides a method of using a compound of formula I or a pharmaceutically acceptable salt thereof for treating a disease, disorder, or syndrome that is at least partially mediated by modulating the in vivo activity of a protein kinase.

[0009] Yet another aspect provides a process for making the compounds of formula A and formula I.

[0010] These and other aspects and embodiments are described below. DETAILED DESCRIPTION OF THE INVENTION

[0011] Abbreviations and Definitions The following abbreviations and terms have the meanings set forth below throughout this specification: [Table 5-1] [Table 5-2]

[0012] The symbol “—” means a single bond, and “═” means a double bond.

[0013] As used herein, the singular forms “a,” “an,” and “the” include references to the plural unless the context clearly dictates otherwise.

[0014] When a variable is generally defined with several possible substituents, each individual radical may be defined with or without a bond. For example, R z can be hydrogen, which means that R zIn the definition, it may be indicated as "-H" or "H".

[0015] When a chemical structure is illustrated or described, unless otherwise explicitly stated, all carbons are assumed to have hydrogen substitution according to a valence of 4. For example, in the structure on the left side of the following figure, it is meant that 9 hydrogens are present. Those 9 hydrogens are illustrated in the structure on the right side. Sometimes, a specific atom in the structure has, in a formula by letters, one hydrogen or a plurality of hydrogens (clearly defined hydrogens) as a substitution, for example, -CH 2 CH 2 - is described. It will be understood by those skilled in the art that the above-described description technique is common in the chemical field where it simplifies and concisely describes the description of complex structures without doing so.

Chem.

[0016] When the group "R" is illustrated as "floating" on a ring system, as in, for example, the formula:

Chem.

[0017] For example, the formula:

Chem.

[0018] "Halogen" or "halo" refers to fluorine, chlorine, bromine, or iodine.

[0019] "C n~m " or "C n ~C m " The term indicates a range including the endpoints, where n and m are integers and indicate the number of carbons. Examples include C 1~4 , C 1 ~C 4 , C1~6 , C 1 ~C 6 etc. are included.

[0020] "Alkyl" refers to a branched or straight-chain hydrocarbon chain of 1 to 8 carbon atoms such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, pentyl, hexyl, and heptyl. (C 1 ~C 6 ) alkyl is preferred. "C n~m alkyl" or (C n ~C m ) alkyl refers to an alkyl group having n to m carbon atoms. When optionally substituted, one or more (e.g., 1 to 4, 1 to 2, or 1) hydrogen atoms of the alkyl group may be replaced by moieties as described below in "optional substitution". In some embodiments, the alkyl group is unsubstituted or optionally unsubstituted.

[0021] "Alkylene" refers to an optionally substituted divalent saturated aliphatic radical having 1 to 10 carbon atoms, 1 to 8 carbon atoms, 1 to 6 carbon atoms, 1 to 4 carbon atoms, or 1 to 2 carbon atoms. When optionally substituted, one or more (e.g., 1 to 4, 1 to 2, or 1) hydrogen atoms of the alkylene group may be replaced by moieties as described below in "optional substitution". In some embodiments, the alkylene group is unsubstituted or optionally unsubstituted. "C n~m alkylene" refers to an alkylene group having n to m carbon atoms. Examples of alkylene groups include, but are not limited to, methylene, ethane-1,2-diyl, propane-1,3-diyl, propane-1,2-diyl, butane-1,4-diyl, butane-1,3-diyl, butane-1,2-diyl, 2-methyl-propane-1,3-diyl, etc.

[0022] The term "alkenyl" refers to a straight-chain or branched hydrocarbon group corresponding to an alkyl group having one or more double carbon-carbon bonds. An alkenyl group formally corresponds to an alkene having one C-H bond replaced by the point of attachment of the alkenyl group to the rest of the compound. "C n~m alkenyl" or (C n ~C m ) alkenyl refers to an alkenyl group having n to m carbons. In some embodiments, the alkenyl moiety contains 2 to 6, 2 to 4, or 2 to 3 carbon atoms. Examples of alkenyl groups include, but are not limited to, ethenyl, n-propenyl, isopropenyl, n-butenyl, sec-butenyl, and the like.

[0023] The term "alkynyl" refers to a straight-chain or branched hydrocarbon group corresponding to an alkyl group having one or more triple carbon-carbon bonds. An alkynyl group formally corresponds to an alkyne having one C-H bond replaced by the point of attachment of the alkyl group to the rest of the compound. "C n~m alkynyl" or (C n ~C m ) alkynyl refers to an alkynyl group having n to m carbons. Examples of alkynyl groups include, but are not limited to, ethynyl, propyn-1-yl, propyn-2-yl, and the like. In some embodiments, the alkynyl moiety contains 2 to 6, 2 to 4, or 2 to 3 carbon atoms.

[0024] "Alkoxy" refers to a moiety of the formula -OR', where R' is an (C 1 ~C 6 ) alkyl moiety as defined herein. "C n~m alkoxy" or (C n ~C m ) alkoxy refers to an alkoxy group whose alkyl group has n to m carbons. Examples of alkoxy moieties include, but are not limited to, methoxy, ethoxy, isopropoxy, and the like.

[0025] The alkoxy group may be unsubstituted or may be optionally substituted. When it is optionally substituted, one or more (for example, 1 to 4, 1 to 2, or 1) hydrogen atoms of the alkoxy group may be replaced by moieties as described below in "optional substitution", provided that the hydrogen atom alpha to the ether oxygen is not replaced by a hydroxy, amino, or thio group. In some embodiments, the alkoxy group is unsubstituted or is not optionally substituted.

[0026] "Alkoxycarbonyl" refers to the group -C(O)-R', where R' is (C 1 ~C 6 ) alkoxy as defined herein.

[0027] The term "amino" refers to a group of the formula -NH 2 .

[0028] The term "carbamyl" refers to a group of the formula -C(O)NH 2 .

[0029] The term "carbonyl", used alone or in combination with other terms, refers to the -C(=O)- group, which may also be written as C(O).

[0030] The term "cyano" or "nitrile" refers to a group of the formula -C≡N, which may also be written as -CN or CN.

[0031] The term "oxo" refers to an oxygen atom as a divalent substituent, which forms a carbonyl group when bonded to carbon, or when bonded to a heteroatom, forms a sulfoxide or sulfone group, or an N-oxide group. In some embodiments, the heterocyclic group may be optionally substituted by one or two oxo (=O) substituents.

[0032] The term "sulfide" refers to a sulfur atom as a divalent substituent, which forms a thiocarbonyl group (C=S) when bonded to carbon.

[0033] As used herein, the term "heteroatom" is intended to include boron, phosphorus, sulfur, oxygen, and nitrogen.

[0034] As used herein, the term "haloalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by halogen atoms. "C n~m haloalkyl" or (C n ~C m ) haloalkyl refers to a C n~m alkyl group having from n to m carbon atoms and at least 1 to {2(n - m)+1} halogen atoms, which may be the same or different. In some embodiments, the halogen atom is a fluoro atom. In some embodiments, the haloalkyl group has 1 to 6 or 1 to 4 carbon atoms. Examples of haloalkyl groups include CF 3 , C 2 F 5 , CHF 2 , CCl 3 , CHCl 2 , C 2 Cl 5 and the like. In some embodiments, the haloalkyl group is a fluoroalkyl group.

[0035] The term "haloalkoxy", used alone or in combination with other terms, refers to a group of the formula -O-haloalkyl, wherein the haloalkyl group is as defined above. "C n~m haloalkoxy" or (C n ~C m ) haloalkoxy refers to a haloalkoxy group in which the haloalkyl group has from n to m carbon atoms. Examples of haloalkoxy groups include trifluoromethoxy and the like. In some embodiments, the haloalkoxy group has 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

[0036] "Aryl" means a monocyclic or bicyclic carbocyclic ring having 6 to 14 ring carbon atoms (e.g., having two fused rings), where the monocyclic ring is aromatic and at least one of the rings of the bicyclic ring is aromatic. "C n~m aryl" or "(C n ~C m )aryl" refers to an aryl group having n to m ring carbon atoms. In some embodiments, the aryl group has 6 to about 10 carbon atoms. In some embodiments, the aryl group has 6 carbon atoms. In some embodiments, the aryl group has 10 carbon atoms. Unless otherwise stated, and as permitted by valence rules, the valence of the group may be located on any atom of any ring within the radical. Representative examples include phenyl, naphthyl, and indanyl.

[0037] The aryl group may be unsubstituted or optionally substituted. When optionally substituted, one or more (e.g., 1 to 5, 1 to 2, or 1) hydrogen atoms of the aryl group may be replaced by moieties as described under "Optional Substitution". In some aspects, the alkoxy group is unsubstituted or not optionally substituted.

[0038] "Arylene" means a divalent 6 to 14 membered monocyclic or bicyclic carbocyclic ring, where the monocyclic ring is aromatic and at least one of the rings of the bicyclic ring is aromatic. Representative examples include phenylene, naphthylene, and indanylene.

[0039] "Cycloalkyl" refers to a non-aromatic hydrocarbon ring system (monocyclic, bicyclic, or polycyclic) including cycloalkyl and alkenyl groups. "C n~m cycloalkyl" or "(C n ~C m) The term "cycloalkyl" refers to cycloalkyl having n to m ring member carbon atoms. The cycloalkyl group may include monocyclic or polycyclic (e.g., having 2, 3, or 4 fused rings) groups and spiro rings. The cycloalkyl group may have 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 ring-forming carbons (C 3~14 ) and may be. In some embodiments, the cycloalkyl group has 3 to 14 members, 3 to 10 members, 3 to 6 ring members, 3 to 5 ring members, or 3 to 4 ring members. In some embodiments, the cycloalkyl group is monocyclic. In some embodiments, the cycloalkyl group is monocyclic or bicyclic. In some embodiments, the cycloalkyl group is a C 3~6 monocyclic cycloalkyl group. The ring-forming carbon atoms of the cycloalkyl group may optionally be oxidized to form oxo or sulfide groups. The cycloalkyl group also includes cycloalkylidene. In some embodiments, the cycloalkyl is cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl. Examples of the cycloalkyl group include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclopentenyl, cyclohexenyl, cyclohexadienyl, cycloheptatrienyl, norbornyl, norpinyl, norcarnyl, bicyclo[1.1.1]pentanyl, bicyclo[2.1.1]hexanyl, and the like. In some embodiments, the cycloalkyl group is cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl. In some embodiments, the cycloalkyl includes a single saturated carbon cyclic ring of 3 to 8 ring carbons such as cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. The cycloalkyl may optionally be substituted with one or more substituents such as 1, 2, or 3 substituents. In some embodiments is that the cycloalkyl substituent is (C 1 ~C 6 ) alkyl, hydroxy, (C 1 ~C 6 ) alkoxy, halo(C 1 ~C 6 ) alkyl, halo(C 1 ~C 6) Alkoxy, halo, amino, mono- and di(C 1 ~C 6 ) alkylamino, hetero(C 1 ~C 6 ) is selected from the group consisting of alkyl, acyl, aryl, and heteroaryl.

[0040] The cycloalkyl group may be unsubstituted or optionally substituted. When optionally substituted, one or more (e.g., 1 to 4, 1 to 2, or 1) hydrogen atoms of the cycloalkyl group may be replaced by moieties as described in "Optional Substitutions". In some embodiments, the substituted cycloalkyl group can incorporate exo- or endocyclic alkenes (e.g., cyclohex-2-en-1-yl). In some embodiments, the cycloalkyl group is unsubstituted or optionally unsubstituted.

[0041] "Cycloalkyloxycarbonyl" means the group -C(O)-OR', where R' is (C 3 ~C 6 ) cycloalkyl as defined herein.

[0042] "Phenyloxycarbonyl" refers to the group -C(O)-O phenyl.

[0043] "Heteroaryl" is -O-, -S(O) n- (where n is 0, 1, or 2), -N-, and -N(R')-, independently selected one or more, preferably 1, 2, 3, or 4 ring heteroatoms, and the remaining ring atoms are carbon, a monovalent monocyclic, fused bicyclic, or fused tricyclic radical of 5 to 14 ring atoms, wherein the ring containing the monocyclic radical is aromatic and at least one of the fused rings containing the bicyclic or tricyclic radical is aromatic. One or two ring carbon atoms of any non-aromatic ring containing a bicyclic or tricyclic radical may be replaced by a -C(O)-, -C(S)-, or -C(=NH)- group. R' is hydrogen, alkyl, hydroxy, alkoxy, acyl, or alkylsulfonyl. Unless otherwise stated, and if permitted by the valence rules, the valence may be located on any atom of any ring of the heteroaryl group. In particular, when the valence point is located on nitrogen, no additional nitrogen substituents are present.More specifically, the term heteroaryl includes, but is not limited to, 1,2,4-triazolyl, 1,3,5-triazolyl, phthalimidyl, pyridinyl, pyrrolyl, imidazolyl, thienyl, furanyl, indolyl, 2,3-dihydro-1H-indolyl (including, for example, 2,3-dihydro-1H-indol-2-yl or 2,3-dihydro-1H-indol-5-yl, etc.), isoindolyl, indolinyl, isoindolinyl, benzimidazolyl, benzodioxol-4-yl, benzofuranyl, cinnolinyl, indolizinyl, naphthyridin-3-yl, phthalazin-3-yl, phthalazin-4-yl, pteridinyl, purinyl, quinazolinyl, quinoxalinyl, tetrazolyl, pyrazolyl, pyrazinyl, pyrimidinyl, pyridazinyl, oxazolyl, isoxazolyl, oxadiazolyl, benzoxazolyl, quinolinyl, isoquinolinyl, tetrahydroisoquinolinyl (including, for example, tetrahydroisoquinolin-4-yl or tetrahydroisoquinolin-6-yl, etc.), pyrrolo[3,2-c]pyridinyl (including, for example, pyrrolo[3,2-c]pyridin-2-yl or pyrrolo[3,2-c]pyridin-7-yl, etc.), benzopyranyl, thiazolyl, isothiazolyl, thiadiazolyl, benzothiazolyl, benzothienyl, and derivatives thereof, and N-oxides or protected derivatives thereof are included.

[0044] A 5-membered heteroaryl ring is a heteroaryl group having 1 or more (e.g., 1, 2, 3, or 4) ring atoms independently selected from 5 ring atoms consisting of N, O, and S. Exemplary 5-membered ring heteroaryls include thienyl, furyl, pyrrolyl, imidazolyl, thiazolyl, oxazolyl, pyrazolyl, isothiazolyl, isoxazolyl, 1,2,3-triazolyl, tetrazolyl, 1,2,3-thiadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-triazolyl, 1,2,4-thiadiazolyl, 1,2,4-oxadiazolyl, 1,3,4-triazolyl, 1,3,4-thiadiazolyl, and 1,3,4-oxadiazolyl. is included.

[0045] The 6-membered heteroaryl ring is a heteroaryl group having 1 or more (e.g., 1, 2, 3, or 4) ring atoms independently selected from N, O, and S. Exemplary 6-membered ring heteroaryls are pyridyl, pyrazinyl, pyrimidinyl, triazinyl, and pyridazinyl.

[0046] "Heteroarylene" means a divalent monocyclic, fused bicyclic, or fused tricyclic radical of 5 to 14 ring atoms containing 1 or more, preferably 1, 2, 3, or 4 ring heteroatoms independently selected from -O-, -S(O) n -(n is 0, 1, or 2), -N-, and -N(R 19 ), with the remaining ring atoms being carbon, wherein the ring containing the monocyclic radical is aromatic and at least one of the fused rings containing the bicyclic or tricyclic radical is aromatic. One or two ring carbon atoms of any non-aromatic ring containing a bicyclic or tricyclic radical may be replaced by a -C(O)-, -C(S)-, or -C(=NH)- group. R 19 is hydrogen, alkyl, or alkenyl. Unless otherwise stated, and subject to the valence rules, the valence may be located on any atom of any ring of the heteroarylene group. Specifically, when the valence point is located on nitrogen, no additional nitrogen substituents are present. More specifically, the term heteroaryl includes, but is not limited to, thien-diyl, benzo[d]isoxazole-diyl, benzo[d]isothiazole-diyl, 1H-indazole-diyl (at the N1 position, optionally substituted with R 19 ), benzo[d]oxazole-diyl, benzo[d]thiazole-diyl, 1H-benzo[d]imidazole-diyl (at the N1 position, optionally substituted with R 19 ), 1H-benzo[d][1,2,3]triazole-diyl (at the N1 position, optionally substituted with R 19(optionally substituted), imidazo[1,2-a]pyridin-diyl, cinnolin-diyl, quinolin-diyl, pyridin-diyl, 1-oxide-pyridin-diyl, [1,2,4]triazolo[4,3-a]pyridin-diyl, and 2,3-dihydroimidazo[1,2-a]pyridin-diyl, etc. are included.

[0047] As used herein, "heterocycloalkyl" or "heterocycle" may optionally contain one or more alkenylene groups as part of the ring structure, and has at least one heteroatom ring member independently selected from boron, nitrogen, sulfur, oxygen, and phosphorus, and refers to a non-aromatic ring or ring system having 4 to 14 ring members, 4 to 10 ring members, 4 to 7 ring members, or 4 to 6 ring members. The term "heterocycloalkyl" includes monocyclic 4-, 5-, 6-, and 7-membered heterocycloalkyl groups. Heterocycloalkyl groups can include monocyclic, bicyclic, or polycyclic (e.g., having two or three fused or bridged rings) ring systems or spiro rings. In some embodiments, the heterocycloalkyl group is a monocyclic group having 1, 2, or 3 heteroatoms independently selected from nitrogen, sulfur, and oxygen. The ring-forming carbon atoms and heteroatoms of the heterocycloalkyl group may optionally be oxidized to form oxo or sulfide groups or other oxidized bonds (e.g., C(O), S(O), C(S), S(O) 2 , N-oxide, etc.) or the nitrogen atom may be quaternized. The heterocycloalkyl group may be bonded via a ring-forming carbon atom or a ring-forming heteroatom. In some embodiments, the heterocycloalkyl group contains 0 to 3 double bonds. In some embodiments, the heterocycloalkyl group contains 0 to 2 double bonds. The definition of heterocycloalkyl also includes a portion having one or more aromatic rings fused to (i.e., having a common bond with) a heterocycloalkyl ring, such as a benzo or thienyl derivative of piperidine, morpholine, azepine, etc. A heterocycloalkyl group containing a fused aromatic ring may be bonded via any ring-forming atom including the ring-forming atoms of the fused aromatic ring. Heterocycloalkyl Examples of groups include azetidinyl, azepanyl, dihydrobenzofuranyl, dihydrofuranyl, dihydropyranyl, morpholino, 3-oxa-9-azaspiro[5.5]undecanyl, 1-oxa-8-azaspiro[4.5]decanyl, piperidinyl, piperazinyl, oxopiperazinyl, pyranyl, pyrrolidinyl, quinuclidinyl, tetrahydrofuranyl, tetrahydropyranyl, 1,2,3,4-tetrahydroquinolinyl, tropanyl, 4,5,6,7-tetrahydrothiazolo[5,4-c]pyridinyl, and thiomorpholino.

[0048] "Heterocycloalkyl" or "heterocycle" may be unsubstituted or optionally substituted. When optionally substituted, one or more (e.g., 1-4, 1-2, or 1) hydrogen atoms of the group may be replaced by moieties independently selected from fluoro, hydroxy, alkoxy, amino, alkylamino, acylamino, thio, and alkylthio. In some embodiments, a substituted heterocyclic group can incorporate an exo- or endocyclic alkene (e.g., cyclohex-2-en-1-yl). In some embodiments, the heterocyclic group is unsubstituted or optionally unsubstituted.

[0049] Optional substitution Unless otherwise indicated, the group is optionally substituted. The term "optionally substituted" refers to being substituted or unsubstituted. In certain embodiments, alkyl, alkenyl, alkynyl, carbocycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups are optionally substituted. "Optionally substituted" refers to a group that may or may not be substituted (e.g., a "substituted" or "unsubstituted" alkyl, "substituted" or "unsubstituted" alkenyl, "substituted" or "unsubstituted" alkynyl, "substituted" or "unsubstituted" cycloalkyl, "substituted" or "unsubstituted" heterocycloalkyl, "substituted" or "unsubstituted" aryl, or "substituted" or "unsubstituted" heteroaryl group). Generally, the term "substituted" means that at least one hydrogen present on the group is replaced by an acceptable substituent, e.g., a substituent that results in a stable compound, e.g., a compound that does not spontaneously undergo transformation such as rearrangement, cyclization, elimination, or other reactions. Unless otherwise indicated, a "substituted" group has a substituent at one or more substitutable positions of the group, and if more than one position in any given structure is substituted, the substituents may be the same or different at each position. The term "substituted" is intended to include substitution with all acceptable substituents of organic compounds and includes any of the substituents described herein that result in the formation of a stable compound. The present invention contemplates any and all such combinations in order to obtain stable compounds. For the purposes of the present invention, a heteroatom such as nitrogen may have a hydrogen substituent and / or any suitable substituent as described herein that satisfies the valence of the heteroatom and results in the formation of a stable moiety. The present invention is not intended to be limited in any way by the exemplary substituents described herein.

[0050] Exemplary carbon atom substituents include, but are not limited to, halogen (halo), -CN, -NO 2 , -N 3 , -SO 2H, -SO 3 H, -OH, -OR aa , -ON(R bb ) 2 , -N(R bb ) 2 , -N(R bb ) 3 + X -、 -N(OR cc )R bb , -SH, -SR aa , -SSR cc , -C(=O)R aa , -CO 2 H, -CHO, -C(OR cc ) 2 , -CO 2 R aa , -OC(=O)R aa , -OCO 2 R aa , -C(=O)N(R bb ) 2 , -OC(=O)N(R bb ) 2 , -NR bb C(=O)R aa , -NR bb CO 2 R aa , -NR bb C(=O)N(R bb ) 2 , -C(=NR bb )R aa , -C(=NR bb )OR aa , -OC(=NR bb )R aa , -OC(=NR bb )OR aa , -C(=NR bb )N(R bb ) 2 , -OC(=NR bb )N(R bb ) 2 , -NR bb C(=NR bb )N(R bb ) 2 , -C(=O)NR bb SO 2 R aa , -NR bbSO 2 R aa 、 -SO 2 N(R bb ) 2 、 -SO 2 R aa 、 -SO 2 OR aa 、 -OSO 2 R aa 、 -S(=O)R aa 、 -OS(=O)R aa 、 -Si(R aa ) 3 、 -OSi(R aa ) 3 、 -C(=S)N(R bb ) 2 、 -C(=O)SR aa 、 -C(=S)SR aa 、 -SC(=S)SR aa 、 -SC(=O)SR aa 、 -OC(=O)SR aa 、 -SC(=O)OR aa 、 -SC(=O)R aa 、 -P(=O) 2 R aa 、 -OP(=O) 2 R aa 、 -P(=O)(R aa ) 2 、 -OP(=O)(R aa ) 2 、 -OP(=O)(OR cc ) 2 、 -P(=O) 2 N(R bb ) 2 、 -OP(=O) 2 N(R bb ) 2 、 -P(=O)(NR bb ) 2 、 -OP(=O)(NR bb ) 2 、 -NR bb P(=O)(OR cc ) 2 、 -NR bb P(=O)(NR bb ) 2 、 -OP(R cc ) 2 、 -OP(R cc ) 3, -B(OR cc ) 2 , -BR aa (OR cc ), C 1~10 alkyl, C 1~10 perhaloalkyl, C 2~10 alkenyl, C 2~10 alkynyl, (C 3 ~C 10 )carbocycloalkyl, 3- to 14-membered heterocycloalkyl, (C 6 ~C 14 )aryl, and 5- to 14-membered heteroaryl are included, where each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R dd groups or two geminal hydrogens on one carbon atom are replaced by the group =O, =S, =NN(R bb ) 2 , =NNR bb C(=O)R aa , =NNR bb C(=O)OR aa , =NNR bb S(=O) 2 R aa , =NR bb , or =NOR cc and each example of R aa is independently (C 1 ~C 10 )alkyl, (C 1 ~C 10 )perhaloalkyl, (C 2 ~C 10 )alkenyl, (C 2 ~C 10 )alkynyl, (C 3 ~C 10 )cycloalkyl, 3- to 14-membered heterocycloalkyl, (C 6 ~C 14 )aryl, and 5- to 14-membered heteroaryl, or two R aaThe groups together form a 3- to 14-membered heterocycloalkyl or 5- to 14-membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R dd groups, Each instance of R bb is independently selected from hydrogen, (C 1 ~C 10 ) perhaloalkyl, (C 2 ~C 10 ) alkenyl, (C 2 ~C 10 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, C 6~14 aryl, and 5- to 14-membered heteroaryl, or two R bb groups together form a 3- to 14-membered heterocycloalkyl or 5- to 14-membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R dd groups, Each instance of R cc is independently selected from hydrogen, (C 1 ~C 10 ) alkyl, (C 1 ~C 10 ) perhaloalkyl, (C 2 ~C 10 ) alkenyl, (C 2 ~C 10 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, 3- to 14-membered heterocycloalkyl, (C 6 ~C 14 ) aryl, and 5- to 14-membered heteroaryl, or two R cc groups together form a 3- to 14-membered heterocycloalkyl or 5- to 14-membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 Rdd is substituted with a base, R dd each example of which is independently halogen, -CN, -NO 2 、-SO 2 H, -SO 3 H, -OH, -OR ee 、-ON(R ff ) 2 、-N(R ff ) 2 、-N(R ff ) 3 + X -、 -N(OR ee )R ff 、-SH, -SR ee 、-SSR ee 、-C(=O)R ee 、-CO 2 H, -CO 2 R ee 、-OC(=O)R ee 、-OCO 2 R ee 、-C(=O)N(R ff ) 2 、-OC(=O)N(R ff ) 2 、-NR ff C(=O)R ee 、-NR ff CO 2 R ee 、-NR ff C(=O)N(R ff ) 2 、-C(=NR ff )OR ee 、-OC(=NR ff )R ee 、-OC(=NR ff )OR ee 、-C(=NR ff )N(R ff ) 2 、-OC(=NR ff )N(R ff ) 2 、-NR ff C(=NR ff )N(R f f ) 2 、-NR ff SO 2R ee 、 -SO 2 N(R ff ) 2 、 -SO 2 R ee 、 -SO 2 OR ee 、 -OSO 2 R ee 、 -S(=O)R ee 、 -Si(R ee ) 3 、 -OSi(R ee ) 3 、 -C(=S)N(R ff ) 2 、 -C(=O)SR ee 、 -C(=S)SR ee 、 -SC(=S)SR ee 、 -P(=O) 2 R ee 、 -P(=O)(R ee ) 2 、 -OP(=O)(R ee ) 2 、 -OP(=O)(OR ee ) 2 、 (C 1 ~C 10 ) alkyl, (C 1 ~C 10 ) perhaloalkyl, (C 2 ~C 10 ) alkenyl, (C 2 ~C 10 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, 3 - to 10 - membered heterocycloalkyl, (C 6 ~C 10 ) aryl, 5 - to 10 - membered heteroaryl, and are selected from, where each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R gg groups, or two geminal R dd substituents may combine together to form =O or =S, R ee Each example of is independently (C 1 ~C 6 ) alkyl, (C 1 ~C6 ) Perhaloalkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, (C 6 ~C 10 ) aryl, 3- to 10-membered heterocycloalkyl, and 3- to 10-membered heteroaryl, wherein each alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R gg groups, R ff each example of which is independently hydrogen, (C 1 ~C 6 ) alkyl, (C 1 ~C 6 ) perhaloalkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, (C 6 ~C 10 ) aryl, and 5- to 10-membered heteroaryl, or two R ff groups together form a 3- to 10-membered heterocycloalkyl or 5- to 10-membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R gg groups, and R gg each example of which is independently halogen, -CN, -NO 2 , -SO 2 H, -SO 3 H, -OH, -OC 1~6 alkyl, -ON(C 1~6 alkyl) 2 , -N(C 1~6 alkyl) 2 , -N(C 1~6(alkyl) 3 + X -、 -NH(C 1~6 (alkyl) 2 + X -、 -NH 2 (C 1~6 (alkyl) + X -、 -NH 3 + X -、 -N(OC 1~6 (alkyl)(C 1~6 (alkyl), -N(OH)(C 1~6 (alkyl), -NH(OH), -SH, -SC 1~6 (alkyl), -SS(C 1~6 (alkyl), -C(=O)(C 1~6 (alkyl), -CO 2 H, -CO 2 (C 1~6 (alkyl), -OC(=O)(C 1~6 (alkyl), -OCO 2 (C 1~6 (alkyl), -C(=O)NH 2 , -C(=O)N(C 1~6 (alkyl) 2 , -OC(=O)NH(C 1~6 (alkyl), -NHC(=O)(C 1~6 (alkyl), -N(C 1~6 (alkyl)C(=O)(C 1~6 (alkyl), -NHCO 2 (C 1~6 (alkyl), -NHC(=O)N(C 1~6 (alkyl) 2 , -NHC(=O)NH(C 1~6 (alkyl), -NHC(=O)NH 2 , -C(=NH)O(C 1~6 (alkyl), -OC(=NH)(C 1~6 (alkyl), -OC(=NH)OC 1~6 (alkyl), -C(=NH)N(C 1~6 (alkyl) 2 , -C(=NH)NH(C 1~6 (alkyl), -C(=NH)NH 2 , -OC(=NH)N(C 1~6(alkyl) 2 ,-OC(NH)NH(C 1~6 alkyl),-OC(NH)NH 2 ,-NHC(NH)N(C 1~6 alkyl) 2 ,-NHC(=NH)NH 2 ,-NHSO 2 (C 1~6 alkyl),-SO 2 N(C 1~6 alkyl) 2 ,-SO 2 NH(C 1~6 alkyl),-SO 2 NH 2 ,-SO 2 C 1~6 alkyl,-SO 2 OC 1~6 alkyl,-OSO 2 C 1~6 alkyl,-SOC 1~6 alkyl,-Si(C 1~6 alkyl) 3 ,-OSi(C 1~6 alkyl) 3 ,-C(=S)N(C 1~6 alkyl) 2 ,C(=S)NH(C 1~6 alkyl),C(=S)NH 2 ,-C(=O)S(C 1~6 alkyl),-C(=S)SC 1~6 alkyl,-SC(=S)SC 1~6 alkyl,-P(=O) 2 (C 1~6 alkyl),-P(=O)(C 1~6 alkyl) 2 ,-OP(=O)(C 1~6 alkyl) 2 ,-OP(=O)(OC 1~ 6 alkyl) 2 ,(C 1 ~C 6 )alkyl,(C 1 ~C 6 )perhaloalkyl,(C 2 ~C 6 )alkenyl,(C 2 ~C6 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, (C 6 ~C 10 ) aryl, 3- to 10-membered heterocycloalkyl, 5- to 10-membered heteroaryl, or two geminal R gg substituents may together form =O or =S, where X - is the counterion.

[0051] As already noted, the nitrogen atom may be substituted or unsubstituted, if valency permits, and includes primary, secondary, tertiary, and quaternary nitrogen atoms. Exemplary nitrogen atom substituents include, but are not limited to, hydrogen, -OH, -OR aa , -N(R cc ), 2 , -CN, -C(=O)R aa , -C(=O)N(R cc ), 2 , -CO 2 R aa , -SO 2 R aa , -C(=NR bb )R aa , -C(=NR cc )OR aa , -C(=NR cc )N(R cc ), 2 , -SO 2 N(R cc ), 2 , -SO 2 R cc , -SO 2 OR cc , -SOR aa , -C(=S)N(R cc ), 2 , -C(=O)SR cc , -C(=S)SR cc , -P(=O) 2 R aa , -P(=O)(R aa ), 2 , -P(=O) 2 N(R cc ), 2 , -P(=O)(NR cc ),2 , (C 1 ~C 10 ) alkyl, (C 1 ~C 10 ) perhaloalkyl, (C 2 ~C 10 ) alkenyl, (C 2 ~C 10 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, 3- to 14-membered heterocycloalkyl, (C 6 ~C1 4 ) aryl, and 5- to 14-membered heteroaryl are included, or two R cc groups bonded to the N atom together form a 3- to 14-membered heterocycloalkyl or 5- to 14-membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R dd groups, and R aa , R bb , R cc and R dd are as defined above.

[0052] In certain embodiments, the substituent present on the nitrogen atom is a nitrogen protecting group (also referred to herein as an "amino protecting group"). Nitrogen protecting groups include, but are not limited to, -OH, -OR aa , -N(R cc ) 2 , -C(=O)R aa , -C(=O)N(R cc ) 2 , -CO 2 R aa , -SO 2 R aa , -C(=NR cc )R aa , -C(=NR cc )OR aa , -C(=NR cc )N(R cc ) 2 , -SO 2 N(R cc ) 2 , -SO 2R cc 、 -SO 2 OR cc 、 -SOR aa 、 -C(=S)N(R cc ) 2 、 -C(=O)SR cc 、 -C(=S)SR cc 、 (C 1 ~C 10 ) alkyl (e.g., aralkyl, heteroaralkyl), (C 2 ~C 10 ) alkenyl, (C 2 ~C 10 ) alkynyl, (C 3 ~C 10 ) cycloalkyl, 3- to 14-membered heterocycloalkyl, (C 6 ~C 14 ) aryl, and 5- to 14-membered heteroaryl groups are included, where each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aralkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R dd groups, and R aa , R bb , R cc , and R dd are as defined herein. Nitrogen protecting groups are well known in the art and include those described in detail in Protecting Groups in Organic Synthesis, T.W. Greene and P.G.M. Wuts, 3 rd edition, John Wiley & Sons, 1999, which is incorporated herein by reference.

[0053] For example, nitrogen protecting groups such as amide groups (e.g., -C(=O)R aa) includes, but is not limited to, formamide, acetamide, chloroacetamide, trichloroacetamide, trifluoroacetamide, phenylacetamide, 3-phenylpropanamide, picolinamide, 3-pyridylcarboxamide, N-benzoylphenylalanyl derivative, benzamide, p-phenylbenzamide, o-nitrophenylacetamide, o-nitrophenoxyacetamide, acetoacetamide, (N'-dithiobenzyl oxyacylamino)acetamide, 3-(p-hydroxyphenyl)propanamide, 3-(o-nitrophenyl)propanamide, 2-methyl-2-(o-nitrophenoxy)propanamide , 2-methyl-2-(o-phenylazophenoxy)propanamide, 4-chlorobutanamide, 3-methyl-3-nitrobutanamide, o-nitrocinnamide, N-acetylmethionine derivative, o-nitrobenzamide, and o-(benzoyloxymethyl)benzamide.

[0054] Nitrogen protecting groups such as carbamate groups (e.g., -C(=O)OR aa) includes, but is not limited to, methylcarbamate, ethylcarbamate, 9-fluorenylmethylcarbamate (Fmoc), 9-(2-sulfo)fluorenylmethylcarbamate, 9-(2,7-dibromo)fluorenylmethylcarbamate, 2,7-di-t-butyl-[9-(10,10-dioxo-10,10,10,10-tetrahydrothioxanthyl)]methylcarbamate (DBD-Tmoc), 4-methoxyphenacylcarbamate (Phenoc), 2,2,2-trichloroethylcarbamate (Troc), 2-trimethylsilylethylcarbamate (Teoc), 2-phenylethylcarbamate (hZ), 1-(1-adamantyl)-1-methylethylcarbamate (Adpoc), 1,1-dimethyl-2-haloethylcarbamate, 1,1-dimethyl-2,2-dibromoethylcarbamate (DB-t-BOC), 1,1-dimethyl-2,2,2-trichloroethylcarbamate (TCBOC), 1-methyl-1-(4-biphenylyl)ethylcarbamate (Bpoc), 1-(3,5-di-t-butylphenyl)-1-methylethylcarbamate (t-Bumeoc), 2-(2’- and 4’-pyridyl)ethylcarbamate (Pyoc), 2-(N,N-dicyclohexylcarboxamido)ethylcarbamate, t-butylcarbamate (BOC or Boc), 1-adamantylcarbamate (Adoc), vinylcarbamate (Voc), allylcarbamate (Alloc), 1-isopropylallylcarbamate (Ipaoc), cinnamylcarbamate (Coc), 4-nitrocinnamylcarbamate (Noc), 8-quinolylcarbamate, N-hydroxypiperidinylcarbamate, alkyldithiocarbamate, benzylcarbamate (Cbz), p-methoxybenzylcarbamate (Moz), p-nitrobenzylcarbamate, p-bromobenzylcarbamate, p-chlorobenzylcarbamate, 2,4-dichlorobenzylcarbamate, 4-methylsulfinylbenzylcarbamate (Msz), 9-anthrylmethylcarbamate, diphenylmethylcarbamate, 2-methylthioethylcarbamate, 2-methylsulfonylethylcarbamate, 2-(p-toluenesulfonyl)ethylcarbamate, [2-(1,3-dithianyl)methylcarbamate (Dmoc), 4-methylthiophenylcarbamate (Mtpc), 2,4-dimethylthiophenylcarbamate (Bmpc), 2-phosphonioethylcarbamate (Peoc), 2-triphenylphosphonioisopropylcarbamate (Ppoc), 1,1-dimethyl-2-cyanoethylcarbamate, m-chloro-p-acetyloxybenzylcarbamate, p-(dihydroxyboryl)benzylcarbamate, 5-benzisoxazolylmethylcarbamate, 2-(trifluoromethyl)-6-chromonylmethylcarbamate (Tcroc), m-nitrophenylcarbamate, 3,5-dimethoxybenzylcarbamate, o-nitrobenzylcarbamate, 3,4-dimethoxy-6-nitrobenzylcarbamate, phenyl(o-nitrophenyl)methylcarbamate, t-amylcarbamate, S-benzylthiocarbamate, p-cyanobenzylcarbamate, cyclobutylcarbamate, cyclohexylcarbamate, cyclopentylcarbamate, cyclopropylmethylcarbamate, p-decyloxybenzylcarbamate, 2,2-dimethoxyacylvinylcarbamate, o-(N,N-dimethylcarboxamide)benzylcarbamate, 1,1-dimethyl-3-(N,N-dimethylcarboxamide)propylcarbamate, 1,1-dimethylpropynylcarbamate, di(2-pyridyl)methylcarbamate, 2-furanylmethylcarbamate, 2-iodoethylcarbamate, isoborynl carbamate, isobutylcarbamate, isonicotinylcarbamate, p-(p'-methoxyphenylazo)benzylcarbamate, 1-methylcyclobutylcarbamate, 1-methylcyclohexylcarbamate, 1-methyl-1-cyclopropylmethylcarbamate, 1- It includes methyl-1-(3,5-dimethoxyphenyl)ethylcarbamate, 1-methyl-1-(p-phenylazophenyl)ethylcarbamate, 1-methyl-1-phenylethylcarbamate, 1-methyl-1-(4-pyridyl)ethylcarbamate, phenylcarbamate, p-(phenylazo)benzylcarbamate, 2,4,6-tri-t-butylphenylcarbamate, 4-(trimethylammonium)benzylcarbamate, and 2,4,6-trimethylbenzylcarbamate.

[0055] Nitrogen protecting groups such as sulfonamide groups (e.g., -S(=O) 2 R aa ) include, but are not limited to, p-toluenesulfonamide (Ts), benzenesulfonamide, 2,3,6-trimethyl-4-methoxybenzenesulfonamide (Mtr), 2,4,6-trimethoxybenzenesulfonamide (Mtb), 2,6-dimethyl-4-methoxybenzenesulfonamide (Pme), 2,3,5,6-tetramethyl-4-methoxybenzenesulfonamide (Mte), 4-methoxybenzenesulfonamide (Mbs), 2,4,6-trimethylbenzenesulfonamide (Mts), 2,6-dimethoxy-4-methylbenzenesulfonamide (iMds), 2,2,5,7,8-pentamethylchroman-6-sulfonamide (Pmc), methanesulfonamide (Ms), β-trimethylsilylethanesulfonamide (SES), 9-anthracenesulfonamide, 4-(4’,8’-dimethoxynaphthylmethyl)benzenesulfonamide (DNMBS), benzylsulfonamide, trifluoromethylsulfonamide, and phenacylsulfonamide.

[0056] Other nitrogen protecting groups include, but are not limited to, phenothiazinyl-(10)-acyl derivatives, N'-p-toluenesulfonylaminoacyl derivatives, N'-phenylaminothioacyl derivatives, N-benzoylphenylalanyl derivatives, N-acetylmethionine derivatives, 4,5-diphenyl-3-oxazolin-2-one, N-phthalimide, N-dithiasuccinimide (Dts), N-2,3-diphenylmaleimide, N-2,5-dimethylpyrrole, N-1,1,4,4-tetramethyldisilazacyclopentane adduct (STABASE), 5-substituted 1,3-dimethyl-1,3,5-triazacyclohexan-2-one, 5-substituted 1,3-dibenzyl-1,3,5-triazacyclohexan-2-one, 1-substituted 3,5-dinitro-4-pyridone, N-methylamine, N-allylamine, N-[2-(trimethylsilyl)ethoxy]methylamine (SEM), N-3-acetoxypropylamine, N-(1-isopropyl-4-nitro-2-oxo-3-pyrroline-3-yl)amine, quaternary ammonium salts, N-benzylamine, N-di(4-methoxyphenyl)methylamine, N-5-dibenzosuberilylamine, N-triphenylmethylamine (Tr), N-[(4-methoxyphenyl)diphenylmethyl]amine (MMTr), N-9-phenylfluorenylamine (PhF), N-2,7-dichloro-9-fluorenylmethyleneamine, N-ferrocenylmethylamino (Fcm), N-2-picolylamino N'-oxide, N-1,1-dimethylthiomethyleneamine, N-benzylideneamine, N-p-methoxybenzylideneamine, N-diphenylmethyleneamine, N-[(2-pyridyl)mesityl]methyleneamine, N-(N',N'-dimethylaminomethylene)amine, N,N'-isopropylidenediamine, N-p-nitrobenzylideneamine, N-salicylideneamine, N-5-chlorosalicylideneamine, N-(5-chloro-2-hydroxyphenyl)phenylmethyleneamine, N-cyclohexylideneamine, N-(5,5-Dimethyl-3-oxo-1-cyclohexenyl)amine, N-borane derivative, N-diphenylboric acid derivative, N-[phenyl(pentaacylchromium- or tungsten)acyl]amine, N-copper chelate, N-zinc chelate, N-nitroamine, N-nitrosoamine, amine N-oxide, diphenylphosphine amide (Dpp), dimethylthiophosphine amide (Mpt), diphenylthiophosphine amide (Ppt), dialkyl phosphoramidate, dibenzyl phosphoramidate, diphenyl phosphoramidate, benzenesulfenamide, o-nitrobenzenesulfenamide (Nps), 2,4-dinitrobenzenesulfenamide, pentachlorobenzenesulfenamide, 2-nitro, 4-methoxybenzenesulfenamide, triphenylmethylsulfenamide, and 3-nitropyridinesulfenamide (Npys) are included.

[0057] In certain embodiments, the substituent present on the oxygen atom is an oxygen protecting group (also referred to herein as a "hydroxyl protecting group"). Oxygen protecting groups include, but are not limited to, -R aa , -N(R bb ) 2 , -C(=O)SR aa , -C(=O)R aa , -CO 2 R aa , -C(=O)N(R bb ) 2 , -C(=NR bb )R aa , -C(=NR bb )OR aa , -C(=NR bb )N(R bb ) 2 , -S(=O)R aa , -SO 2 R aa , -Si(R aa ) 3 , -P(R cc ) 2 , -P(R cc ) 3 , -P(=O) 2 R aa , -P(=O)(R aa) 2 ,-P(=O)(OR cc ) 2 ,-P(=O) 2 N(R bb ) 2 ,and -P(=O)(NR bb ) 2 is included, where R aa , R bb , and R cc are as defined herein. Oxygen protecting groups are well known in the art and include those described in detail in Protecting Groups in Organic Synthesis, T.W. Greene and P.G.M. Wuts, 3 rd edition, John Wiley & Sons, 1999, which is incorporated herein by reference.

[0058] Exemplary oxygen protecting groups include, but are not limited to, methyl, methoxymethyl (MOM), methylthiomethyl (MTM), t-butylthiomethyl, (phenyldimethylsilyl)methoxymethyl (SMOM), benzyloxymethyl (BOM), p-methoxybenzyloxymethyl (PMBM), (4-methoxyphenoxy)methyl (p-AOM), guaiacolmethyl (GUM), t-butoxymethyl, 4-pentenyl oxymethyl (POM), siloxymethyl, 2-methoxyethoxymethyl (MEM), 2,2,2-trichloroethoxymethyl, bis(2-chloroethoxy)methyl, 2-(trimethylsilyl)ethoxymethyl (SEMOR), tetrahydropyranyl (THP), 3-bromotetrahydropyranyl, tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetrahydropyranyl (MTHP), 4-methoxytetrahydrothiopyranyl, 4-methoxytetrahydrothiopyranyl S,S-dioxide, 1-[(2-chloro-4-methyl)phenyl]-4-methoxypiperidin-4-yl (CTMP), 1,4-dioxan-2-yl, tetrahydrofuranyl, tetrahydrothiofuranyl, 2,3,3a,4,5,6,7,7a-octahydro-7,8,8-trimethyl-4,7-methanobenzofuran-2-yl, 1-ethoxyethyl, 1-(2-chloroethoxy)ethyl, 1-methyl-1-methoxyethyl, 1-methyl-1-benzyloxyethyl, 1-methyl-1-benzyloxy-2-fluoroethyl, 2,2,2-trichloroethyl, 2-trimethylsilylethyl, 2-(phenylselenyl)ethyl, t-butyl, allyl, p-chlorophenyl, p-methoxyphenyl, 2,4-dinitrophenyl, benzyl (Bn), p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2-picolyl, 4-picolyl, 3-methyl-2-picolyl N-oxide, diphenylmethyl, p,p'-dinitrobenzhydryl, 5-dibenzosuberyl, triphenylmethyl, α-naphthyldiphenylmethyl, p-methoxyphenyldiphenylmethyl, di(p-methoxyphenyl)phenylmethyl, tri(p-methoxyphenyl)methyl, 4-(4'-bromophenacyloxyphenyl)diphenylmethyl, 4,4',4''-tris(4,5-dichlorophthalimidophenyl)methyl, 4,4',4''-tris(levulinoyloxyphenyl)methyl, 4,4',4''-tris(benzoyloxyphenyl)methyl, 3-(imidazol-1-yl)bis(4',4''-dimethoxyphenyl)methyl, 1,1-bis(4-methoxyphenyl)-1'-pyrenylmethyl, 9-anthryl, 9-(9-phenyl)xanthenyl, 9-(9-phenyl-10-oxo)anthryl, 1,3-benzodithiolan-2-yl, benzisothiazolyl S,S-dioxide, trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropyl, Cyril (IPDMS), diethylisopropylsilyl (DEIPS), dimethyl t-hexylsilyl, t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl (TBDPS), tribenzylsilyl, tri-p-xylylsilyl, triphenylsilyl, diphenylmethylsilyl (DPMS), t-butylmethoxyphenylsilyl (TBMPS), formate, benzoylformate, acetate, chloroacetate, dichloroacetate, trichloroacetate, trifluoroacetate, methoxyacetate, triphenylmethoxyacetate, phenoxyacetate, p-chlorophenoxyacetate, 3-phenylpropionate, 4-oxopentanoate (levulinate), 4,4-(ethylenedithio)pentanoate (levulinoyldithioacetal), pivaloate, adamantoate, crotonate, 4-methoxycrotonate, benzoate, p-phenylbenzoate, 2,4,6-trimethylbenzoate (mesitoate), methyl carbonate, 9-fluorenylmethyl carbonate (Fmoc), ethyl carbonate, 2,2,2-trichloroethyl carbonate (Troc), 2-(trimethylsilyl)ethyl carbonate (TMSEC), 2-(phenylsulfonyl)ethyl carbonate (Psec), 2-(triphenylphosphonio)ethyl carbonate (Peoc), isobutyl carbonate, vinyl carbonate, allyl carbonate, t-butyl carbonate (BOC or Boc), p-nitrophenyl carbonate, benzyl carbonate, p-methoxybenzyl carbonate, 3,4-dimethoxybenzyl carbonate, o-nitrobenzyl carbonate, p-nitrobenzyl carbonate, S-benzylthiocarbonate, 4-ethoxy-1-naphthyl carbonate, methyl dithiocarbonate, 2-iodobenzoate, 4-azidobutyrate, 4-nitro-4-methylpentanoate, o-(dibromomethyl)benzoate, 2-formylbenzenesulfonate, 2-(methylthiomethoxy)ethyl, 4-(methylthiomethoxy)butyrate, 2-(methylthiomethoxymethyl)benzoate, 2,6-dichloro-4-methylphenoxyacetate, 2,6-dichloro-4-(1,1,3,3-Tetramethylbutyl)phenoxyacetate, 2,4-bis(1,1-dimethylpropyl)phenoxyacetate, chlorodiphenylacetate, isobutyrate, monosuccinoate, (E)-2-methyl-2-butenoate, o-(methoxyacyl)benzoate, α-naphthoate, nitrate, alkyl N,N,N’,N’-tetramethylphosphorodiamidate, alkyl N-phenylcarbamate, borate, dimethylphosphinothioyl, alkyl 2,4-dinitrophenylsulfenate, sulfate, methanesulfonate (mesylate), benzylsulfonate, and tosylate (Ts) are included.,

[0059] In certain embodiments, the substituent present on the sulfur atom is a sulfur protecting group (also referred to as a “thiol protecting group”). Sulfur protecting groups include, but are not limited to, -R aa , -N(R bb ) 2 , -C(=O)SR aa , -C(=O)R aa , -CO 2 R aa , -C(=O)N(R bb ) 2 , -C(=NR bb )R aa , -C(=NR bb )OR aa , -C(=NR bb )N(R bb ) 2 , -S(=O)R aa , -SO 2 R aa , -Si(R aa ) 3 , -P(R cc ) 2 , -P(R cc ) 3 , -P(=O) 2 R aa , -P(=O)(R aa ) 2 , -P(=O)(OR cc ) 2 , -P(=O) 2 N(R bb ) 2 , and -P(=O)(NR bb) 2 is included, where R aa , R bb , and R cc are as defined herein. Sulfur protecting groups are well known in the art and include those described in detail in Protecting Groups in Organic Synthesis, T.W. Greene and P.G.M. Wuts, 3 rd edition, John Wiley & Sons, 1999, which is incorporated herein by reference.

[0060] As used herein, "leaving group" (LG) is a term understood in the art to refer to a molecular fragment that departs with a pair of electrons upon heterolytic bond cleavage, where the molecular fragment is an anion or a neutral molecule. As used herein, a leaving group may be an atom or group that can be replaced by a nucleophile. See, for example, Smith, March Advanced Organic Chemistry 6th ed. (501-502). Exemplary leaving groups include, but are not limited to, halo (e.g., chloro, bromo, iodo), -OR (when the O atom is bonded to a carbonyl group, where R aa is as defined herein), -O(C=O)R aa , or -O(SO) LG R 2 (e.g., tosyl, mesyl, besyl), where R LG is optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, the leaving group is a halogen. LG

[0061] The terms defined above are specifically illustrated in the examples.

[0062] The "yield" for each of the reactions described herein is expressed as a percentage of the theoretical yield.

[0063] As used herein, the term "patient" in the context of the present invention includes humans and any other animals, particularly mammals, and other living organisms. Accordingly, the methods are applicable to both human therapy and veterinary use. In a preferred embodiment, the patient is a mammal, and in the most preferred embodiment, the patient is a human. Examples of preferred mammals include mice, rats, other rodents, rabbits, dogs, cats, pigs, cows, sheep, horses, and primates.

[0064] "Kinase-dependent disease or condition" refers to a pathological condition that depends on the activity of one or more kinases. Kinases are directly or indirectly involved in signal transduction pathways for various cellular activities including proliferation, adhesion, migration, differentiation, and invasion. Diseases associated with kinase activity include tumor growth and pathological angiogenesis associated with other diseases such as solid tumor growth and excessive local angiogenesis associated with eye diseases (such as diabetic retinopathy, age-related macular degeneration, etc.), and inflammation (such as psoriasis, rheumatoid arthritis, etc.).

[0065] "Therapeutically effective amount" is the amount of a compound of the present invention that, when administered to a patient, alleviates the symptoms of a disease. The amount of the compound of the present invention that constitutes a "therapeutically effective amount" should vary depending on the compound, the condition and its severity, the age of the patient being treated, etc. A therapeutically effective amount can be routinely determined by one of ordinary skill in the art, taking into account the knowledge of the person skilled in the art and the present disclosure.

[0066] "Cancer" includes, but is not limited to: Heart: Sarcoma (angiosarcoma, fibrosarcoma, rhabdomyosarcoma, liposarcoma), myxoma, rhabdomyoma, fibroma, lipoma and teratoma; Head and Neck: Squamous cell carcinoma of the head and neck, laryngeal and hypopharyngeal cancer, nasal and paranasal sinus cancer, nasopharyngeal cancer, salivary gland cancer, oral and oropharyngeal cancer; Lung: Bronchogenic carcinoma (squamous cell, undifferentiated small cell, undifferentiated large cell, adenocarcinoma, non-small cell lung cancer), alveolar (bronchioloalveolar) carcinoma, bronchial adenoma, sarcoma, lymphoma, chondroid hamartoma, mesothelioma; Colon: Colorectal cancer, adenocarcinoma, gastrointestinal stromal tumor, lymphoma, carcinoid tumor, Turcot syndrome; Gastrointestinal: Gastric cancer, gastroesophageal junction adenocarcinoma, esophagus (squamous cell carcinoma, adenocarcinoma, leiomyosarcoma, lymphoma), stomach (cancer, lymphoma, leiomyosarcoma), pancreas (ductal adenocarcinoma, insulinoma, glucagonoma, gastrinoma, carcinoid tumor, VIPoma), small intestine (adenocarcinoma, lymphoma, carcinoid tumor, Kaposi sarcoma, leiomyoma, hemangioma, lipoma, neurofibroma, fibroma), large intestine (adenocarcinoma, tubular adenoma, villous adenoma, hamartoma, leiomyoma); Breast: Metastatic breast cancer, ductal carcinoma in situ, invasive ductal carcinoma, tubular carcinoma, medullary carcinoma, mucinous carcinoma, lobular carcinoma in situ, triple-negative breast cancer; Urinary and Reproductive Tract: Kidney (adenocarcinoma, Wilms tumor [nephroblastoma], lymphoma, leukemia, renal cell carcinoma), bladder and urethra (squamous cell carcinoma, transitional cell carcinoma, adenocarcinoma, urothelial carcinoma), prostate (adenocarcinoma, sarcoma, castration-resistant prostate cancer), testis (seminoma, teratoma, embryonal carcinoma, teratocarcinoma, choriocarcinoma, sarcoma, interstitial cell carcinoma, fibroma, fibroadenoma, adenomatoid tumor, lipoma), clear cell carcinoma, papillary carcinoma; Liver: Hepatocellular carcinoma (hepatocarcinoma), cholangiocarcinoma, hepatoblastoma, angiosarcoma, hepatocellular adenoma, hemangioma; Bone: Osteogenic sarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma, chondrosarcoma, Ewing sarcoma, malignant lymphoma (microglioma), multiple myeloma, malignant giant cell tumor, chordoma, etc. Osteochondroma (osteochondroma), benign chondroma, chondroblastoma, chondromyxofibroma, osteoid osteoma, and giant cell tumor, thyroid: medullary thyroid carcinoma, differentiated thyroid carcinoma, papillary thyroid carcinoma, follicular thyroid carcinoma, Hürthle cell carcinoma, and anaplastic thyroid carcinoma, nervous system: skull (osteoma, angioma, granuloma, xanthoma, Paget's disease), meninges (meningioma, meningiosarcoma, gliosis), brain (glioblastoma, medulloblastoma, glioma, ependymoma, germ cell tumor [pineal tumor], glioblastoma multiforme, oligodendroglioma, schwannoma, retinoblastoma, congenital tumor), spinal nerve fibroma, meningioma, glioma, sarcoma), gynecology: uterus (endometrial cancer), cervix (cervical cancer, pre-tumor cervical dysplasia), ovary (ovarian cancer [serous cystadenocarcinoma, mucinous cystadenocarcinoma, unclassified cancer], granulosa-theca cell tumor, Sertoli-Leydig cell tumor, undifferentiated germ cell tumor, malignant teratoma), vulva (squamous cell carcinoma, intraepithelial carcinoma, adenocarcinoma, fibrosarcoma, melanoma), vagina (clear cell carcinoma, squamous cell carcinoma, sarcoma botryoides [fetal rhabdomyosarcoma], fallopian tube (cancer), hematological system: blood (myeloid leukemia [acute and chronic], acute lymphoblastic leukemia, chronic lymphocytic leukemia, myeloproliferative disorders, multiple myeloma, myelodysplastic syndrome), Hodgkin's disease, non-Hodgkin lymphoma [malignant lymphoma], skin: malignant melanoma, basal cell carcinoma, squamous cell carcinoma, Kaposi's sarcoma, dysplastic nevus, lipoma, angioma, dermatofibroma, keloid, psoriasis, and adrenal gland: refers to cell proliferative disorders including neuroblastoma. Therefore, the term "cancerous cell" as provided herein includes cells affected by any one of the conditions identified above.

[0067] "Pharmaceutically acceptable salts" include "pharmaceutically acceptable acid addition salts" and "pharmaceutically acceptable base addition salts". "Pharmaceutically acceptable acid addition salts" are formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and further with organic acids such as acetic acid, trifluoroacetic 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, etc., and refer to salts that maintain the biological effectiveness of the free base and are not biologically or otherwise undesirable.

[0068] The term "pharmaceutically acceptable basic addition salts" includes those derived from inorganic bases such as sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum salts and the like. Exemplary salts are ammonium, potassium, sodium, calcium, and magnesium salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include, but are not limited to, primary, secondary, and tertiary amines such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, methylglucosamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins, substituted amines including naturally occurring substituted amines, cyclic amines, and salts of basic ion exchange resins. Exemplary organic bases are isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine (see, for example, S.M. Berge, et al., "Pharmaceutical Salts," J. Pharm. Sci., 1977; 66: 1-19, which is incorporated herein by reference).

[0069] As used herein, the term "compound" is intended to include all stereoisomers, geometric isomers, tautomers and isotopes of the structures shown. This term is also intended to refer to the compounds of the present invention regardless of the method by which they are prepared, for example, by synthesis, by biological processes (e.g., metabolism or enzymatic conversion), or by a combination thereof.

[0070] The compounds of the present invention may include all isotopes of atoms that occur in the intermediates or final compounds. Isotopes include atoms having the same atomic number but different mass numbers. For example, isotopes of hydrogen include tritium and deuterium.

[0071] Any one of the process steps or sequences disclosed and / or claimed herein can be carried out under an inert gas atmosphere, more particularly under argon or nitrogen. In addition, the methods of the present invention may be carried out as semi - continuous or continuous processes, more preferably as continuous processes.

[0072] Furthermore, many of the process steps and sequences described herein can be shortened.

[0073] Generally, the nomenclature used in this application is based on the naming rules adopted by the International Union of Pure and Applied Chemistry (IUPAC). The chemical structures shown herein were created using CHEMDRAW®. Any open valences present on carbon, oxygen, or nitrogen atoms in the structures within this specification indicate the presence of hydrogen atoms.

[0074] Embodiments of the present invention One aspect provides a compound of formula A:

Chemical formula

[0075] In one embodiment of this aspect, the compound of formula A is a compound of formula A - 1:

Chemical formula

[0076] In another embodiment of this aspect, the compound of formula A is a compound of formula A - 2:

Chemical formula

[0077] In a further embodiment of this aspect, the compound of formula A is a compound of formula A-3: [Chemical formula] wherein R a1 is -H or (C 1 ~C 6 ) alkyl].

[0078] In a further embodiment, R 1 in the compound of formula A-3 is -H.

[0079] In a further embodiment of this aspect, the compound of formula A is a compound of formula A-4: [Chemical formula] wherein ring A is a 5- to 14-membered heteroaryl, and the subscript r is 1, 2, 3, or 4.

[0080] In this embodiment, R 2 is -H.

[0081] In a further embodiment, r in formula A-4 is 1 or 2.

[0082] In a further embodiment of formula A: R 1 is -H, optionally substituted (C 1 ~C 6 ) alkyl, halo, -OR a , -NO 2 , -NH 2 , -NHR a , -NR a R a , -SR a , -SOR a , or -S(O) 2 R a and R 2 is (C 2 ~C6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, -CN, -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)NR a R a , NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOR a )R a , -C(=NOH)NR a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NR a )NR a R a , -NR a C(=NR a )NR a R a , -NR a S(O)R a , -NR a S(O) 2 R a , -NR a S(O) 2 NR a R a , -S(O)R a, -S(O)NR a R a , -S(O) 2 R a , -S(O) 2 NR a C(O)R a , -P(O)R a R a , -P(O)(OR a )(OR a ), -B(OH) 2 , -B(OR a ) 2 , and -S(O) 2 NR a R a is selected from the group consisting of.

[0083] In one embodiment of this embodiment, R 1 is -H.

[0084] In a further embodiment: R 1 is (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 10-membered heteroaryl, 4- to 10-membered heterocycloalkyl, -CN, -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a Ra , -C(=NR a )R a , -C(=NOH)R a , -C(=NOH)NR a , -C(=NOR a )R a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NR a )NR a R a , -NR a C(=NR a )NR a R a , -NR a S(O)R a , -NR a S(O) 2 R a , -NR a S(O) 2 NR a R a , -S(O)R a , -S(O)NR a R a , -S(O) 2 R a , -S(O) 2 NR a C(O)R a , -P(O)R a R a , -P(O)(OR a )(OR a ), -B(OH) 2 , -B(OR a ) 2 , and -S(O) 2 NR a R a selected from the group consisting of, and R 2 is -H, optionally substituted (C 1 ~C 6 ) alkyl, halo, -OR a , -NO 2 , -NH 2 , -NHR a , -NR a R a , -SRa 、 -SOR a 、 or -S(O) 2 R a is.

[0085] In one embodiment of this embodiment, R 2 is -H.

[0086] In another embodiment: R 1 is (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5 - 10 membered heteroaryl, 4 - 10 membered heterocycloalkyl, -CN, -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOR a )R a , -C(=NOH)NR a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NRa )NR a R a 、 -NR a C(=NR a )NR a R a 、 -NR a S(O)R a 、 -NR a S(O) 2 R a 、 -NR a S(O) 2 NR a R a 、 -S(O)R a 、 -S(O)NR a R a 、 -S(O) 2 R a 、 -S(O) 2 NR a C(O)R a 、 -P(O)R a R a 、 -P(O)(OR a )(OR a )、 -B(OH) 2 、 -B(OR a ) 2 、 or -S(O) 2 NR a R a and R 2 is -H, halo, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 1 ~C 6 ) haloalkyl, (C 1 ~C 6 ) haloalkoxy, (C 6 ~C 10 ) aryl-(C 1 ~C 4 ) alkylene-, (C 3 ~C 10 ) cycloalkyl-(C 1 ~C 4 ) alkylene-, (5 - 14 membered heteroaryl)-(C 1 ~C4 ) alkylene-, (4- to 14-membered heterocycloalkyl)-(C 1 ~C 4 ) alkylene-, -CN, -NO 2 , -OR a , -SR a , -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)R a , -OC(O)NR a R a , -NHR a , -NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOH)NR a , -C(=NOR a )R a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NR a )NR a R a , -NR a C(=NR a )NR a R a , -NR a S(O)R a , -NR a S(O) 2 R a , -NR aS(O) 2 NR a R a , -S(O)R a , -S(O)NR a R a , -S(O) 2 R a , -S(O) 2 NR a C(O)R a , -P(O)R a R a , -P(O)(OR a )(OR a ), -B(OH) 2 , -B(OR a ) 2 , and -S(O) 2 NR a R a is selected from the group consisting of:

[0087] In one embodiment of this embodiment, R 2 is -H.

[0088] In another embodiment: R 1 -H, halo, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 1 ~C 6 ) haloalkyl, (C 1 ~C 6 ) haloalkoxy, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5-14 membered heteroaryl, 4-14 membered heterocycloalkyl, (C 6 ~C 10 )Aryl-(C 1 ~C 4 ) alkylene-, (C 3 ~C 10 )Cycloalkyl-(C 1 ~C 4 ) alkylene-, (5-14 membered heteroaryl)-(C1 ~C 4 ) alkylene-, (4- to 14-membered heterocycloalkyl)-(C 1 ~C 4 ) alkylene-, -CN, -NO 2 , -OR a , -SR a , -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)R a , -OC(O)NR a R a , -NHR a , -NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOR a )R a , -C(=NOH)NR a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NR a )NR a R a , -NR a C(=NR a )NR a R a , -NR a S(O)R a , -NR a S(O) 2 R a, -NR a S(O) 2 NR a R a , -S(O)R a , -S(O)NR a R a , -S(O) 2 R a , -S(O) 2 NR a C(O)R a , -P(O)R a R a , -P(O)(OR a )(OR a ), -B(OH) 2 , -B(OR a ) 2 , and -S(O) 2 NR a R a is selected from the group consisting of.

[0089] In one embodiment of this embodiment, R 1 is -H.

[0090] In another further embodiment: R 1 and R 2 together with the atoms to which they are attached form a fused (C 3 ~C 7 ) cycloalkyl ring or a fused 4- to 10-membered heterocycloalkyl ring, where the fused (C 3 ~C 7 ) cycloalkyl ring or the fused 4- to 10-membered heterocycloalkyl ring is each optionally substituted with 1, 2, or 3 independently selected R b substituents, provided that the compound is not 1-[2-(4-fluoro-phenyl)-acetyl]-cyclopropanecarboxylic acid [3-fluoro-4-(7,8,10,11,13,14-hexahydro-6,9,12,15-tetraoxa-1-aza-cyclododeca[b]naphthalen-4-yloxy)-phenyl]-amide.

[0091] In yet another embodiment: R 1 and R2 together with the atoms to which they are attached, form a fused (C 3 ~C 7 ) cycloalkyl ring or a fused 4- to 10-membered heterocycloalkyl ring, where the fused (C ~C 3 ~C 7 ) cycloalkyl ring or fused 4- to 10-membered heterocycloalkyl ring is each optionally substituted with one, two, or three independently selected R b substituents, provided that the compound is not a compound having the formula:

Chemical formula

[0092] In another embodiment, R in the compound of formula A, A-1, or A-3 1 is -H, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, -C(=NO-(C 1 ~C 6 ) alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a , phenyl, 5- to 6-membered heteroaryl, (C 3 ~C 6 ) cycloalkyl, and 4- to 6-membered heterocycloalkyl, selected from the group consisting of.

[0093] In another embodiment, R 1 is -H, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C6 ) alkynyl, -C(=NO-(C 1 ~C 6 ) alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a , phenyl, 5- to 6-membered heteroaryl, (C 3 ~C 6 ) cycloalkyl, and 4- to 6-membered heterocycloalkyl, selected from the group consisting of, and R 2 is H, optionally substituted (C 1 ~C 6 ) alkyl, halo, -OR a , -NO 2 , -NH 2 , -NHR a , -NR a R a , -SR a , -SOR a , or -S(O) 2 R a is.

[0094] In another embodiment, R 1 is -H, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, -C(=NO-(C 1 ~C 6 ) alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a , phenyl, 5- to 6-membered heteroaryl, (C 3 ~C 6 ) cycloalkyl: and 4- to 6-membered heterocycloalkyl, selected from the group consisting of, and R 2 is -H, halo, (C 1 ~C 6 )alkyl, (C 2 ~C 6 )alkenyl, (C 2 ~C 6 )alkynyl, (C 1 ~C 6 )haloalkyl, (C 1 ~C 6 )haloalkoxy, (C 6 ~C 10 )aryl, (C 3 ~C 10 )cycloalkyl, (C 6 ~C 10 )aryl-(C 1 ~C 4 )alkylene-, (C 3 ~C 10 )cycloalkyl-(C 1 ~C 4 )alkylene-, (5 - 14 membered heteroaryl)-(C 1 ~C 4 )alkylene-, (4 - 14 membered heterocycloalkyl)-(C 1 ~C 4- )alkylene-, -CN, -NO 2 , -OR a , -SR a , -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)R a , -OC(O)NR a R a , -NHR a , -NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)ORa , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOH)NR a , -C(=NOR a )R a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NR a )NR a R a , -NR a C(=NR a )NR a R a , -NR a S(O)R a , -NR a S(O) 2 R a , -NR a S(O) 2 NR a R a , -S(O)R a , -S(O)NR a R a , -S( O) 2 R a , -S(O) 2 NR a C(O)R a , -P(O)R a R a , -P(O)(OR a )(OR a ), -B(OH) 2 , -B(OR a ) 2 , and -S(O) 2 NR a R a is selected from the group consisting of.

[0095] In another embodiment, R 1 is -H, (C 1 ~C 6 ) alkyl, (C 2 ~C6 ) alkenyl, (C 2 ~C 6 ) alkynyl, -C(=NO-(C 1 ~C 6 ) alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a , phenyl, 5- to 6-membered heteroaryl, -(C 3 ~C 6 ) cycloalkyl, and 4- to 6-membered heterocycloalkyl, selected from the group consisting of, and R 2 is, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, -CN, -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOH)NR a , -C(=NOR a )R a , -C(=NCN)NR a R a , -NR a C(=NCN)NRa R a 、 -C(=NR a )NR a R a 、 -NR a C(=NR a )NR a R a 、 -NR a S(O)R a 、 -NR a S(O) 2 R a 、 -NR a S(O) 2 NR a R a 、 -S(O)R a 、 -S(O)NR a R a 、 -S(O) 2 R a 、 -S(O) 2 NR a C(O)R a 、 -P(O)R a R a 、 -P(O)(OR a )(OR a )、 -B(OH) 2 、 -B(OR a ) 2 、 or -S(O) 2 NR a R a is.

[0096] In a further embodiment, R 1 is -H, R a NHC(O)-, R a OC(O)-, (C 1 ~C 6 ) alkyl, (C 1 ~C 6 ) alkoxy, or -C(=NO-CH 3 )R a and R 2is selected from 2-methoxyethylamino, azetidin-1-yl, methylamino, 3-morpholinopropoxy, 2-methoxyethoxy, 2-hydroxyethoxy, propoxy, 2-hydroxypropoxy, methoxycarbonyl, carboxy, carbamoyl, methylcarbamoyl, (2-hydroxyethoxy)carbamoyl, (2,2-dihydroxyethoxy)carbamoyl, (oxetan-3-yloxy)carbamoyl, methoxycarbamoyl, 2-trimethylsilylethynyl, ethynyl, sulfamoyl, acetyl, and -C(=NOCH 3 )CH 3 selected from.

[0097] In a further embodiment of Formula A and A-2, R 2 is -H, (C 1 ~C 6 )alkyl, (C 2 ~C 6 )alkenyl, (C 2 ~C 6 )alkynyl, -C(=NO-(C 1 ~C 6 )alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , and -S(O) 2 NR a R a selected from the group consisting of.

[0098] In a further embodiment, R 1 is -H, optionally substituted (C 1 ~C 6 )alkyl, halo, -OR a , -NO 2 , -NH 2 , -NHR a , -NR a R a , -SR a , -SOR a , or S(O) 2 R a and R 2 is -H, (C1 ~C 6 )alkyl, (C 2 ~C 6 )alkenyl, (C 2 ~C 6 )alkynyl, -C(=NO-(C 1 ~C 6 )alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a , phenyl, and (C 3 ~C 6 )cycloalkyl selected from the group consisting of.

[0099] In a further embodiment, R 1 is -H, optionally substituted (C 1 ~C 6 )alkyl, halo, -OR a , -NO 2 , -NH 2 , -NHR a , -NR a R a , -SR a , -SOR a , and -S(O) 2 R a selected from the group consisting of, and R 2 is -H, (C 1 ~C 6 )alkyl, (C 2 ~C 6 )alkenyl, (C 2 ~C 6 )alkynyl, -C(=NO-(C 1 ~C 6 )alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a from the group consisting of or are selected.

[0100] In a further embodiment, R 1 is (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 10-membered heteroaryl, 4- to 10-membered heterocycloalkyl, -CN, -NHOR a , -C(O)R a , -C(O)NR a R a , -C(O)NHOR a , -C(O)OR a , -C(O)NR a S(O) 2 R a , -OC(O)NR a R a , -NR a C(O)R a , -NR a C(=NR a )R a , -NR a C(O)OR a , -NR a C(O)NR a R a , -C(=NR a )R a , -C(=NOH)R a , -C(=NOH)NR a , -C(=NOR a )R a , -C(=NCN)NR a R a , -NR a C(=NCN)NR a R a , -C(=NR a )NR a R a , -NR a C(=NR a )NR a R a , -NR a S(O)R a, -NR a S(O) 2 R a , -NR a S(O) 2 NR a R a , -S(O)R a , -S(O)NR a R a , -S(O) 2 R a , -S(O) 2 NR a C(O)R a , -P(O)R a R a , -P(O)(OR a )(OR a ), -B(OH) 2 , -B(OR a ) 2 , and -S(O) 2 NR a R a selected from the group consisting of, and R 2 is -H, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, -C(=NO-(C 1 ~C 6 ) alkyl)R a , -CN, -C(O)OR a , -C(O)NR a R a , -C(O)NHOR a , -S(O) 2 NR a R a selected from the group consisting of.

[0101] In a further embodiment, R 1 is -H, halo, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 1 ~C 6) haloalkyl, (C 1 ~C 6 ) haloalkoxy, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 14-membered heteroaryl, 4- to 14-membered heterocycloalkyl, (C 6 ~C 10 ) aryl-(C 1 ~C 4 ) alkylene-, (C 3 ~C 10 ) cycloalkyl-(C 1 ~C 4 ) alkylene-, (5- to 14-membered heteroaryl)-(C 1 ~C 4 ) alkylene-, (4- to 14-membered heterocycloalkyl)-(C 1 ~C 4 ) alkylene-, -CN, -NO 2 、-OR a 、-SR a 、-NHOR a 、-C(O)R a 、-C(O)NR a R a 、-C(O)NHOR a 、-C(O)OR a 、-C(O)NR a S(O) 2 R a 、-OC(O)R a 、-OC(O)NR a R a 、-NHR a 、-NR a R a 、-NR a C(O)R a 、-NR a C(=NR a )R a 、-NR a C(O)OR a 、-NR a C(O)NR a R a 、-C(=NR a )R a 、-C(=NOH)R a 、-C(=NOR a )Ra ,-C(=NOH)NR a ,-C(=NCN)NR a R a ,-NR a C(=NCN)NR a R a ,-C(=NR a )NR a R a ,-NR a C(=NR a )NR a R a ,-NR a S(O)R a ,-NR a S(O) 2 R a ,-NR a S(O) 2 NR a R a ,-S(O)R a ,-S(O)NR a R a ,-S(O) 2 R a ,-S(O) 2 NR a C(O)R a ,-P(O)R a R a ,-P(O)(OR a )(OR a ),-B(OH) 2 ,-B(OR a ) 2 ,-and -S(O) 2 NR a R a selected from the group consisting of, and R 2 is -H, (C 1 ~C 6 ) alkyl, (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, -C(=NO-(C 1 ~C 6 ) alkyl)R a ,-CN, -C(O)OR a ,-C(O)NR a R a ,-C(O)NHOR a, -S(O) 2 NR a R a is selected from the group consisting of

[0102] In a further embodiment, R 1 is 2-methoxyethylamino, azetidin-1-yl, methylamino, 3-morpholinopropoxy, 2-methoxyethoxy, 2-hydroxyethoxy, propoxy, 2-hydroxypropoxy, methoxycarbonyl, carboxy, carbamoyl, methylcarbamoyl, 2-oxazolyl, pyrazol-3-yl, pyrazol-4-yl, 4-isoxazolyl, 3,5-dimethylisoxazol-4-yl, 1-methyl-pyrazol-4-yl, 2-methyl-pyrazol-3-yl, 2-ethyl-pyrazol-3-yl, 2-(2-hydroxyethyl)-pyrazol-3-yl, 2-( 2,2,2-trifluoroethyl)-pyrazol-3-yl, 2-(2-fluoroethyl)-pyrazol-3-yl, 2-(2,2-difluoroethyl)-pyrazol-3-yl, 2-trifluoromethyl-pyrazol-3-yl, 2-difluoromethyl-pyrazol-3-yl, 1-methyl-imidazol-4-yl, 1-methyl-imidazol-2-yl, 1H-imidazol-2-yl, (2-hydroxyethoxy)carbamoyl, (2,2-dihydroxyethoxy)carbamoyl, (oxetan-3-yloxy)carbamoyl, methoxycarbamoyl, 2-trimethylsilylethynyl, ethynyl, 1,3,4-oxadiazol-3-yl, 1H-1,2,3-triazol-5-yl, sulfamoyl, acetyl, and -C(=NOCH 3 )CH 3 is selected from the group consisting of and R 2 is -H, -R a NHC(O)-, -R a OC(O)-, (C 1 ~C 6 )alkyl, (C 1 ~C 6 )alkoxy, or -C(=NO-CH 3 )R a is.

[0103] In a further embodiment of Formula A-4, the subscript r is 1 or 2.

[0104] In a further embodiment of the above aspects and embodiments, R 10 and R 11 are each -H.

[0105] In a further embodiment of the above aspects and embodiments, the subscript n is 1.

[0106] In a further embodiment of the above aspects and embodiments, the subscript m is 1.

[0107] In a further embodiment of the above aspects and embodiments, the subscript p is 1.

[0108] In another embodiment, the compound of Formula A is a compound of Formula B:

Chemical formula

[0109] In one embodiment of Formulas A and B: (i) R 1 is: (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 10-membered heteroaryl, and 4- to 10-membered heterocycloalkyl (wherein the (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 10-membered heteroaryl, and 4- to 10-membered heterocycloalkyl are each independently optionally substituted), -CN, -P(O)R a R a , P(O)(OR a ) 2 , B(OH) 2 , B(OR a ) 2 , X 2 R a (wherein X 2 is -NHO-, -NH-S(O)-, -N-(C 1 ~C 6 ) alkyl-S(O)-, -NH-S(O) 2 -, -N-(C 1 ~C 6 ) alkyl-S(O) 2 R a -, -NH-S(O)-NH-, -N-(C 1 ~C 6 ) alkyl-S(O)NH-, -NH-S(O) 2 NH-, -N-(C 1 ~C 6 ) alkyl-S(O) 2 NH-, -S(O) 2 NHC(O)-), and

Chemical formula

[0110] In another embodiment of Formulas A and B: (ii) R 1 is: (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 10-membered heteroaryl, and 4- to 10-membered heterocycloalkyl (wherein (C 2 ~C 6 ) alkenyl, (C 2 ~C 6 ) alkynyl, (C 6 ~C 10 ) aryl, (C 3 ~C 10 ) cycloalkyl, 5- to 14-membered heteroaryl, and 4- to 14-membered heterocycloalkyl are each independently optionally substituted), P(O)R a R a , P(O)(OR a )(OR a ), B(OH) 2 , B(OR a ) 2 , CN, X 2 R a (wherein X 2 is -NHO-, -NH-S(O)-, -N-(C 1 ~C 6 ) alkyl-S(O)-, -NH-S(O) 2 -, -N-(C 1 ~C 6 ) alkyl-S(O) 2 R a -, -NH-S(O)-NH-, -N-(C 1 ~C 6 ) alkyl-S(O)NH-, -NH-S(O) 2NH-, -N-(C 1 ~C 6 )alkyl-S(O) 2 NH-, -S(O)-, -S(O) 2 -, -S(O) 2 NHC(O)), and

Chem.

Chem.

Chem.

Chemical formula

[0111] In another embodiment of formulas A and B: (iii)R 1 and R 2 together with the atom to which they are attached form a 4- to 10-membered heterocycloalkyl ring optionally substituted with 1, 2, or 3 groups independently selected from the group consisting of halo, (C 1 ~C 6 )alkyl, (C 1 ~C 6 )haloalkyl, (C 1 ~C 6 )haloalkoxy, -CN, -OH, -NH 2 provided that the compound is not 1-[2-(4-fluoro-phenyl)-acetyl]-cyclopropanecarboxylic acid [3-fluoro-4-(7,8,10,11,13,14-hexahydro-6,9,12,15-tetraoxa-1-aza-cyclododeca[b]naphthalen-4-yloxy)-phenyl]-amide.

[0112] In yet another embodiment of the compounds of formulas A and B: R 1 is -H, -CN, (C 1 ~C 6 )alkyl, (C 3 ~C 10 )cycloalkyl, (C6 ~C 10 ) aryl, 4- to 10-membered heterocycloalkyl, 5- to 10-membered heteroaryl, -S(O) 2 NHR a , -P(O)R a R a , -OR a , or [Chemical formula] and where [Chemical formula] indicates the point of attachment: Y 1 is absent or is -NH-, -N-(C 1 ~C 6 )alkyl-, or -O-, Y 2 is absent or is -O-, -NH-, -NHO-, -N-(C 1 ~C 6 )alkyl-, -NH-NH-, -NH-S(O)-, or NH-S(O) 2 and Z 1 is -O, -NH, -N-(C 1 ~C 6 )alkyl, -N-OH, or -N-O(C 1 ~C 6 )alkyl.

[0113] In another embodiment of Formulas A and B: R 2 is -H, halo, -X 1 R a , (C 2 ~C 6 )alkenyl, (C 2 ~C 6 )alkynyl, or [Chemical formula] and where [Chemical formula] indicates a junction point: Y 1 is absent or is NH, N-(C 1 ~C 6 )alkyl, or O, Y 2 is absent or is O, NH, NHO, N-(C 1 ~C 6 )alkyl, N 2 H 2 , NH-S(O), or NH-S(O) 2 and Z 1 is -O, -NH, -N-(C 1 ~C 6 )alkyl, -NOH-, or -N-O(C 1 ~C 6 )alkyl.

[0114] In another embodiment of Formulas A and B, R 3 is -H or halo.

[0115] In another embodiment of Formulas A and B, R 4 is -H or halo.

[0116] In another embodiment of Formulas A and B, R 14 is -H or halo.

[0117] In another embodiment of Formulas A and B, Y is -O-.

[0118] In another embodiment, the compound of B is a compound of either Formula B-1 or B-2:

Chemical formula

[0119] In one embodiment of Formula B-1: R a1 is optionally substituted (C 1 ~C 6 )alkyl, R 1 is -H, -CN, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted phenyl, optionally substituted 4- to 6-membered heterocycloalkyl, optionally substituted 5- to 6-membered heteroaryl, -SO 2 -(C 1 ~C 6 )alkyl, -SO 2 NH 2 、-SO 2 -NH(C 1 ~C 6 )alkyl, P(O)((C 1 ~C 6 )alkyl) 2 、or

Chemical formula

Chemical formula

Chemical formula

Chemical formula

[0120] In another embodiment of Formula B-1: R a1 is (C 1 ~C 6 )alkyl, R 1 is -H, -CN, optionally substituted cyclopropyl, optionally substituted phenyl, optionally substituted 4- to 6-membered azetidinyl, optionally substituted pyrrolidinyl, optionally substituted piperidinyl, optionally substituted oxetanyl, optionally substituted oxazolyl, optionally substituted pyridinyl, optionally substituted imidazolyl, optionally substituted pyrrolyl, optionally substituted furanyl, optionally substituted pyrazolyl, optionally substituted oxadiazolyl, -SO 2 -(C 1 ~C 6 )alkyl, -SO 2 NH 2 , -SO 2 -NH(C 1 ~C 6 )alkyl, or P(O)((C1 ~C 6 )alkyl 2 or R 1 is [Chemical formula] and here: Y 1 is absent Y 2 is O, NH, NHO, NH-NH, or N-(C 1 ~C 6 )alkyl or Y 2 is optionally substituted azetidinyl Z 1 is O, NH, or N-(C 1 ~C 6 )alkyl and R a is H, (C 1 ~C 6 )alkyl, -(C 2 ~C 6 )alkylene-OH, -CH 2 CHOH-(C 2 ~C 6 )alkylene-OH, -(C 2 ~C 6 )alkylene-NH 2 , -(C 2 ~C 6 )alkylene-NH(C 1 ~C 6 )alkyl, -(C 2 ~C 6 )alkylene-N((C 1 ~C 6 )alkyl) 2 , -(C 2 ~C 6 )alkylene-heterocycloalkyl), or 4-6 membered heterocycloalkyl, where the heterocycloalkyl is optionally substituted.

[0121] In one embodiment of formula B-2: R a2 is optionally substituted (C1 ~C 6 ) is alkyl, R 2 is,

Chem.

Chem.

[0122] In another embodiment of Formula B-2: R a2 is (C 1 ~C 6 ) alkyl, R 2 is,

Chem.

Chem.

[0123] In another embodiment of Formula B-1, R a1 is methoxy.

[0124] In another embodiment of Formula B-2, R a2 is methoxy.

[0125] In another embodiment, the compound of Formula B is either a compound of Formula B-3 or B-4:

Chemical formula

Chemical formula

[0126] In one embodiment of Formula B-3: R 1 is -H or (C 1 ~C 6 ) alkyl, and Y 1 is absent, Y 2 is absent or is -O-, -NHO-, or -NH-, and Z 1 is O or NO-(C 1 ~C 6 ) alkyl, and R a is -H or -(C 1 ~C 6 ) alkyl.

[0127] In another embodiment of Formula B-3: R 1 is -H or methyl, Y 1 is absent, Y 2 is absent or is -O-, -NHO-, or -NH-, and Z 1 is O or NO-Me, and R a is -H or Me.

[0128] In another embodiment of formula B-3: R 1 and R a together with the atoms to which they are attached form a 4- to 6-membered heterocycloalkyl ring optionally substituted with halo, (C ~C 1 ~C 6 )alkyl, or (C 1 ~C 6 )haloalkyl.

[0129] In one embodiment of formula B-4: Y 1 is absent, Y 2 is O, NH, NHO, NH-NH, or N-(C 1 ~C 6 )alkyl or alternatively, Y 2 is optionally substituted azetidinyl, Z 1 is O, NH, NO-(C 1 ~C 6 )alkyl, or N-(C 1 ~C 6 )alkyl and R a is H, (C 1 ~C 6 )alkyl, -(C 2 ~C 6 )alkylene-OH, -CH 2 CHOH-(C 2 ~C 6 )alkylene-OH, -(C 2 ~C 6 )alkylene-NH 2 -(C 2 ~C 6 )alkylene-NH(C 1 ~C 6 )alkyl, -(C 2 ~C 6 )alkylene-N((C 1 ~C 6 )alkyl) 2 -(C 2 ~C6 ) alkylene - optionally substituted 4 - 6 - membered heterocycloalkyl), or optionally substituted 4 - 6 - membered heterocycloalkyl, and R 2 is -H, -F, -Cl, -Br, -(C 1 ~C 6 ) alkoxy, -O-(C 2 ~C 6 ) alkylene -OH, -O-(C 2 ~C 6 ) alkylene -O-(C 1 ~C 6 alkyl), (C 2 ~C 6 ) alkylene -O-(C 1 ~C 6 ) alkyl, -NH 2 , -NH-(C 1 ~C 6 alkyl), -NH-(C 1 ~C 6 ) alkylene -(optionally substituted 4 - 6 - membered heterocycloalkyl), or -NH-(C 2 ~C 6 ) alkylene -O-(C 1 ~C 6 alkyl).

[0130] In another embodiment of formula B - 4: Y 1 is absent, Y 2 is O, NH, NHO, NH - NH, or N-(C 1 ~C 6 ) alkyl or Y 2 is optionally substituted azetidinyl, Z 1 is O, NH, NO-(C 1 ~C 6 ) alkyl, N-(C 1 ~C 6 ) alkyl, and R a is -H, methyl, ethyl, -(C 2 ~C 6 ) alkylene -OH, -CH2 CHOH-(C 2 ~C 6 ) alkylene-OH, -(C 2 ~C 6 ) alkylene-NH 2 、-(C 2 ~C 6 ) alkylene-NHMe, -(C 2 ~C 6 ) alkylene-N(Me) 2 、-(C 1 ~C 6 ) alkylene-morpholinyl), -(C 1 ~C 6 ) alkylene-piperidinyl), (C 1 ~C 6 ) alkylene-(optionally substituted pyrrolidinyl), optionally substituted azetidinyl, or optionally substituted oxetanyl, and R 2 is -H, -F, -Cl, -Br, methoxy, -O-(C 2 ~C 6 ) alkylene-OH, -O-(C 2 ~C 6 ) alkylene-OMe, -NH 2 、-NH-(C 1 ~C 6 alkyl), -NH-(C 2 ~C 6 ) alkylene-OMe, -NH-(C 2 ~C 6 ) alkylene-(optionally substituted morpholinyl), or -NH-(C 2 ~C 6 ) alkylene-O-(C 1 ~C 6 alkyl).

[0131] In another embodiment of Formula B-4: R 2 and R a together with the atom to which they are attached are halo, (C 1 ~C 6 ) alkyl, or (C 1 ~C 6)It forms a 4- to 6-membered heterocycloalkyl ring optionally substituted with haloalkyl.

[0132] In another embodiment, the compound of formula B is a compound of formula B-5:

Chemical formula

[0133] In one embodiment of formula B-5: Ring A is optionally substituted (C 6 ~C 10 ) aryl, optionally substituted (C 3 ~C 10 ) cycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 4- to 10-membered heterocycloalkyl, and R 2 is H, or (C 1 ~C 6 ) alkoxy.

[0134] In another embodiment of formula B-5: Ring A is optionally substituted phenyl, optionally substituted cyclopropyl, optionally substituted pyridyl, optionally substituted imidazolyl, optionally substituted pyrrolyl, optionally substituted furanyl, optionally substituted pyrazolyl, optionally substituted oxazolyl, optionally substituted azetidinyl, or optionally substituted oxetanyl, and R 2 is H, or methoxy.

[0135] Another embodiment of the compounds of formula A and B is a compound of formula C:

Chemical formula

[0136] In another embodiment of Formula C: Y 1 is absent, Y 2 is O, NH, NHO, NH-NH, or N-(C 1 ~C 6 ) alkyl, or Y 2 is optionally substituted azetidinyl, Z 1 is O, NH, NO-(C 1 ~C 6 ) alkyl, or N-(C 1 ~C 6 ) alkyl, R a is -H, methyl, ethyl, -(C 2 ~C 6 ) alkylene-OH, -CH 2 CHOH-(C 2 ~C 6 ) alkylene-OH, -(C 2 ~C 6 ) alkylene-NH 2 , -(C 2 ~C 6 ) alkylene-NHMe, -(C 2 ~C 6 ) alkylene-N(Me) 2 , -(C 1 ~C6 ) (alkylene-morpholinyl), -(C 1 ~C 6 ) (alkylene-piperidinyl), (C 1 ~C 6 ) (alkylene-(optionally substituted pyrrolidinyl)), optionally substituted azetidinyl, or optionally substituted oxetanyl, and R 2 is -H, -F, -Cl, -Br, methoxy, -O-(C 2 ~C 6 ) (alkylene-OH), -O-(C 2 ~C 6 ) (alkylene-OMe), -NH 2 , -NH-(C 1 ~C 6 alkyl), -NH-(C 2 ~C 6 ) (alkylene-OMe), -NH-(C 2 ~C 6 ) (alkylene-(optionally substituted morpholinyl)), or -NH-(C 2 ~C 6 ) (alkylene-O-(C 1 ~C 6 alkyl), and n and m are each 0 or 1.

[0137] In another embodiment of Formula C: R 2 and R a together with the atom to which they are attached form a 4- to 6-membered heterocycloalkyl ring optionally substituted with halo, (C 1 ~C 6 ) alkyl, and (C 1 ~C 6 ) haloalkyl, and n and m are each independently an integer from 0 to 3.

[0138] Another embodiment of Formula C is a compound of Formula C-1:

Chemical Formula

[0139] Another embodiment of Formulas A and B is a compound of Formula D:

Chemical formula

[0140] In one embodiment of Formula D: R 1 is -H or (C 1 ~C 6 ) alkyl, and Y 1 is absent, Y 2 is absent or is -O-, -NHO-, or -NH-, and Z 1 is O or NO-(C 1 ~C 6 ) alkyl, R a is -H or -(C 1 ~C 6 ) alkyl, and n and m are each independently an integer from 0 to 3.

[0141] In another embodiment of Formula D: R 1 is -H or methyl, Y 1 is absent, Y 2 is absent or is -O-, -NHO-, or -NH-, and Z 1 is O or NO-Me, and R a is -H, or -Me, n and m are each independently an integer from 0 to 1.

[0142] In another embodiment of formula D: R 1 and R a together with the atom to which they are attached form a 4 - to 6 - membered heterocycloalkyl ring optionally substituted with halo, (C 1 ~C 6 )alkyl , or (C 1 ~C 6 )haloalkyl and n and m are each independently an integer from 0 to 1. n and m are each independently an integer from 0 to 1.

[0143] Another embodiment of formula D is a compound of formula D - 1

Chemical formula

[0144] Another embodiment of formulas A and B is a compound of formula E:

Chemical formula

[0145] In another embodiment of formula E: Ring A is phenyl optionally substituted, cyclopropyl optionally substituted, pyridyl optionally substituted, imidazolyl optionally substituted, pyrrolyl optionally substituted, furanyl optionally substituted, pyrazolyl optionally substituted, oxazolyl optionally substituted, azetidinyl optionally substituted, or oxetanyl optionally substituted, and R 2 is H or methoxy.

[0146] Another embodiment of Formulas A and B is a compound of Formula F:

Chemical formula

Chemical formula

Chemical formula

Chemical formula

Chemical formula

[0147] In another embodiment of formula F: R a1is methyl, R 1 is H, -CN, optionally substituted cyclopropyl, optionally substituted phenyl, optionally substituted 4- to 6-membered azetidinyl, optionally substituted pyrrolidinyl, optionally substituted piperidinyl, optionally substituted oxetanyl, optionally substituted oxazolyl, optionally substituted pyridinyl, optionally substituted imidazolyl, optionally substituted pyrrolyl, optionally substituted furanyl, optionally substituted pyrazolyl, optionally substituted oxadiazolyl, -SO 2 -(C 1 ~C 6 )alkyl, -SO 2 NH 2 , -SO 2 -NH(C 1 ~C 6 )alkyl, or P(O)((C 1 ~C 6 )alkyl) 2 or R 1 is

Chemical Structure

[0148] Another embodiment of Formulas A and B is a compound of Formula G:

Chemical formula

Chemical formula

Chemical formula

[0149] In another embodiment of formula G: R a2 is methyl, R 2 is

Chemical formula

Chemical formula

[0150] Another embodiment of formulas A and B is a compound of formula H for modulating kinase activity:

Chemical formula

[0151] In one embodiment of the compound of formula I, Y is O.

[0152] In another embodiment, R 3 is -H.

[0153] In another embodiment, [Chemical] is unsubstituted.

[0154] In another embodiment, R 4 is a halo.

[0155] In another embodiment, R 4 is parafluoro.

[0156] In another embodiment, R 2 is -H, halo, or optionally substituted (C 1 ~C 6 )-alkoxy.

[0157] In another embodiment, R 1 is -CN.

[0158] In another embodiment, R 1 is -CO 2 H.

[0159] In another embodiment, R 1 is -CO 2 -Me.

[0160] In another embodiment, R 1 is -CO-NHR 6 .

[0161] In another embodiment, R 1 is -CO-NH2.

[0162] In another embodiment, R 1 is -CO-NMeR 6 .

[0163] In another embodiment, R 3 is -H or halo.

[0164] In another embodiment, R 1 is -CN, -(SO 2 )NH 2, -OMe, -(SO 2 )CH 3

Chem.

Chem.

[0165] In another embodiment, R 1 is

Chem.

Chem.

[0166] In another embodiment, R 1 is

Chem.

Chem.

[0167] In another embodiment, R 2 is -H, -CN, -Br, -F, -Cl, -OMe, -CH 3 ,

Chem.

Chem.

[0168] In another embodiment, R 1 is -H, methyl, or methoxy.

[0169] In another embodiment, R 2 is -CO 2 H.

[0170] In another embodiment, R 1 is -CO 2 -Me.

[0171] In another embodiment, R 1 is -CO-NHR 6 .

[0172] In another embodiment, R 1 is -CO-NH 2 .

[0173] In another embodiment, R 1 is -CO-NMeR 6 .

[0174] In another embodiment, R 1 is

Chemical formula

[0175] In another embodiment, R 1 and R 2 together form

Chemical formula

[0176] In a further embodiment, the compound of formula I is a compound of formula I-1:

Chemical formula

[0177] In another aspect, the present invention provides a compound of formula A or A-I presented in Table 1 below.

Table 1-1

Table 1-2

Table 1-3

Table 1-4

Table 1-5

Table 1-6

Table 1-7

Table 1-8

Table 1-9

Table 1-10

Table 1-11

Table 1-12

Table 1-13

Table 1-14

Table 1-15

Table 1-16

Table 1-17

Table 1-18

Table 1-19

Table 1-20

Table 1-21

Table 1-22

Table 1-23

Table 1-24

Table 1-25

Table 1-26

Table 1-27

Table 1-28

[0178] Systemic administration Administration of the compounds of the present invention, or pharmaceutically acceptable salts thereof, in pure form or in suitable pharmaceutical compositions, can be effected via any of the acceptable modes of administration or agents to obtain similar usefulness. Thus, administration can be, for example, oral, nasal, parenteral (intravenous, intramuscular, or subcutaneous), topical, transdermal, intravaginal, intravesical, intraosseous, or rectal, in the form of, for example, tablets, suppositories, pills, soft and hard gelatin capsules, powders, solutions, suspensions, aerosols, etc., in solid, semi-solid, lyophilized powder, or liquid dosage forms, preferably in unit dosage forms suitable for easy administration of exact dosages.

[0179] The compositions contain conventional pharmaceutical carriers or additives and the compounds of the present invention as the active agent, and may in addition contain other pharmaceutical agents, pharmaceuticals, carriers, adjuvants, etc. The compositions of the present invention may be used in combination with anti-cancer drugs or other agents commonly administered to patients undergoing cancer treatment. Adjuvants include preservatives, wetting agents, suspending agents, sweetening agents, flavoring agents, perfumes, emulsifying agents, and dispersing agents. Prevention of the action of microorganisms can be ensured by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, etc. It is also desirable to include isotonic agents, for example, sugars, sodium chloride, etc. Prolonged absorption of injectable pharmaceutical forms can be brought about by using agents that retard absorption, for example, aluminum monostearate and gelatin.

[0180] If desired, the pharmaceutical compositions of the present invention may contain small amounts of auxiliary substances such as wetting or emulsifying agents, pH buffering agents, antioxidants, etc., for example, citric acid, sorbitan monolaurate, triethanolamine oleate, butylated hydroxytoluene, etc.

[0181] Compositions suitable for parenteral injection may include physiologically acceptable aqueous or non-aqueous sterile solutions, dispersions, suspensions or emulsions, and sterile powders that are reconstituted into sterile injection solutions or dispersions. Examples of suitable aqueous or non-aqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols (such as propylene glycol, polyethylene glycol, glycerol, etc.), suitable mixtures thereof, vegetable oils (such as olive oil), and injectable organic esters such as ethyl oleate. Suitable fluidity can be maintained, for example, by the use of coating agents such as lecithin, by maintaining the required particle size in the case of dispersions, and by the use of surfactants.

[0182] One of the preferred routes of administration is oral, using a convenient daily dosing schedule that can be adjusted according to the severity of the disease state being treated.

[0183] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one conventional inert additive (or carrier), such as sodium citrate or dicalcium phosphate, or (a) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, and silicic acid, (b) binders, such as cellulose derivatives, starch, alginate, gelatin, polyvinylpyrrolidone, sucrose, and acacia, (c) wetting agents, such as glycerol, (d) disintegrating agents, such as agar, calcium carbonate, potato or tapioca starch, alginic acid, croscarmellose sodium, complex silicates, and sodium carbonate, (e) dissolution retardants, such as paraffin, (f) absorption promoters, such as quaternary ammonium compounds, (g) wetting agents, such as cetyl alcohol, and glycerol monostearate, magnesium stearate, etc., (h) adsorbents, such as kaolin and bentonite, and (i) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may contain buffering agents.

[0184] The solid dosage forms as described above can be prepared using coating agents and shells, such as enteric coatings, and others well-known in the art. This may contain sedatives and may also be a composition that releases the active compound(s) in a delayed manner in a specific part of the intestinal tract. Examples of embedding compositions that can be used are polymeric substances and waxes. The active compound, if suitable, may be in microencapsulated form containing one or more of the above-mentioned additives.

[0185] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs. Such dosage forms can be prepared, for example, by dissolving or dispersing the compound(s) of the present invention or its pharmaceutically acceptable salts, and optional pharmaceutical adjuvants, in a carrier such as water, physiological saline, aqueous dextrose, glycerol, ethanol, etc.; solubilizing and emulsifying agents such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, and dimethylformamide; oils, especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol, and fatty acid esters of sorbitan; or mixtures of these substances, etc., thereby forming a solution or suspension. 。

[0186] In addition to the active compound, the suspension may contain suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol, and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, and tragacanth, or mixtures of these substances.

[0187] Compositions for rectal administration are, for example, suppositories that are solid at room temperature but liquid at body temperature and thus melt while in a suitable body cavity and release the active ingredient therein, which can be prepared by mixing the compound of the present invention with a suitable non-irritating additive or carrier, such as cocoa butter, polyethylene glycol or suppository wax.

[0188] Dosage forms for topical administration of the compounds of the present invention include ointments, powders, sprays, and inhalants. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and, if necessary, any preservative, buffer or propellant. Ophthalmic formulations, eye ointments, powders and solutions are also contemplated to be within the scope of the present disclosure.

[0189] In general, depending on the intended mode of administration, pharmaceutically acceptable compositions contain from about 1% to about 99% by weight of the compound(s) of the present invention, or a pharmaceutically acceptable salt thereof, and from 99% to 1% by weight of suitable pharmaceutical additives. In one example, the composition contains from about 5% to about 75% by weight of the compound(s) of the present invention or a pharmaceutically acceptable salt thereof, the balance being suitable pharmaceutical additives.

[0190] The actual methods for preparing such dosage forms are known or apparent to those skilled in the art; see, for example, Remington’s Pharmaceutical Sciences, 18th Ed., (Mack Publishing Company, Easton, Pa., 1990). In any case, the administered composition contains a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof, for treating a disease state in accordance with the teachings of the present invention.

[0191] The compounds of the present invention, or pharmaceutically acceptable salts thereof, are administered in a therapeutically effective amount that will vary depending on a variety of factors including the activity of the specific compound employed, the metabolic stability and length of action of the compound, age, body weight, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, the severity of the particular disease state, and the recipient of treatment. The compounds of the present invention can be administered to a patient at dosage levels ranging from about 0.1 to about 1,000 mg per day. For a normal human adult of about 70 kilograms body weight, an example of a dosage range is from about 0.01 to about 100 mg per kilogram of body weight per day. However, the specific dosage employed may vary. For example, the dosage may depend on many factors including the needs of the patient, the severity of the condition being treated, and the pharmacological activity of the compound employed. Methods for determining the optimal dosage for a particular patient are well known to those of ordinary skill in the art.

[0192] Combination therapy The compounds as disclosed herein can be administered as monotherapy or in combination ( "co - administration") with one or more additional therapies for treating a disease or disorder, such as a disease or disorder associated with hyper - proliferation, e.g., cancer. Therapies that can be used in combination with the compounds disclosed herein include (i) surgery, (ii) radiation therapy (e.g., gamma radiation, neutron beam radiation therapy, electron beam radiation therapy, proton beam radiation therapy, brachytherapy, and systemic radioisotopes), (iii) endocrine therapy, (iv) adjuvant therapy, immunotherapy, CAR T - cell therapy, and (v) other chemotherapeutic agents.

[0193] The term "co - administered" ( "co - administer") refers to any method of simultaneous administration, or separate sequential administration, of a compound of formula I' or a salt thereof and one or more additional pharmaceutical active ingredient(s) including cytotoxic agents and radiation treatment. When the administrations are not simultaneous, the compounds are administered in close temporal proximity to each other. Further, it is not important whether the compounds are administered in the same dosage form; for example, one compound may be administered topically and the other compound may be administered orally.

[0194] Typically, any agent that is active against the disease or condition being treated can be co-administered. Examples of such agents for cancer treatment can be found, for example, in publicly available information sources such as https: / / www.cancer.gov / about-cancer / treatment / drugs (last accessed January 22, 2019) and Cancer Principles and Practice of Oncology by V.T. Devita and S.Hellman (editors), 11 th edition (2018), Lippincott Williams & Wilkins Publishers. One of ordinary skill in the art will be able to identify which combinations of agents are useful based on the specific characteristics of the drug and the disease in question.

[0195] In one embodiment, the treatment method includes co-administration of a compound as disclosed herein or a pharmaceutically acceptable salt thereof with at least one immunotherapy. Immunotherapy (also referred to as biological response modifier therapy, biologic therapy, biotherapy, immune therapy, or biological therapy) is a treatment that uses part of the immune system to fight disease. Immunotherapy can assist the immune system in recognizing cancer cells or enhance the response against cancer cells. Immunotherapy includes active and passive immunotherapy. Active immunotherapy stimulates the body's own immune system, while passive immunotherapy generally uses immune system components made outside the body.

[0196] Examples of active immunotherapy include, but are not limited to, cancer vaccines, tumor cell vaccines (autologous or allogeneic), dendritic cell vaccines, antigen vaccines, anti-idiotype vaccines, DNA vaccines, viral vaccines, or vaccines including tumor-infiltrating lymphocyte (TIL) vaccines containing interleukin-2 (IL-2), or lymphokine-activated killer (LAK) cell therapy.

[0197] Examples of passive immunotherapy include, but are not limited to, monoclonal antibodies and toxin-containing targeted therapeutic agents. Monoclonal antibodies include naked antibodies and conjugated monoclonal antibodies (also called tagged, labeled or loaded antibodies). Naked monoclonal antibodies have no drugs or radioactive substances added, while conjugated monoclonal antibodies are conjugated to, for example, chemotherapeutic drugs (chemical labels), radioactive particles (radioactive labels), or toxins (immunotoxins). Examples of these naked monoclonal antibody drugs include, but are not limited to, rituximab (Rituxan), an antibody against the CD20 antigen used to treat, for example, B-cell non-Hodgkin lymphoma, trastuzumab (Herceptin), an antibody against the HER2 protein used to treat, for example, advanced breast cancer, alemtuzumab (Campath), an antibody against the CD52 antigen used to treat, for example, B-cell chronic lymphocytic leukemia (B-CLL), cetuximab (Erbitux), an antibody against the EGFR protein used, for example, in combination with irinotecan to treat advanced colorectal cancer and head and neck cancer, and bevacizumab (Avastin), an anti-angiogenic therapy that acts against the VEGF protein and is used, for example, in combination with chemotherapy to treat metastatic colorectal cancer. Conjugated mono Examples of monoclonal antibodies include, but are not limited to, radiolabeled antibody ibritumomab tiuxetan (Zevalin), which is used to deliver radioactivity directly to cancerous B lymphocytes, for example, to treat B-cell non-Hodgkin lymphoma; radiolabeled antibody tositumomab (Bexxar), which is used to treat certain types of non-Hodgkin lymphoma; and immunotoxin gemtuzumab ozogamicin (Mylotarg), which contains calicheamicin and is used to treat, for example, acute myeloid leukemia (AML). BL22 is a conjugated monoclonal antibody, for example, for treating hairy cell leukemia, an immunotoxin for treating leukemia, lymphoma, and brain tumors, and radiolabeled antibodies such as OncoScint for colorectal and ovarian cancers, and ProstaScint for prostate cancer, for example.

[0198] Further examples of therapeutic antibodies that can be used include, but are not limited to, HERCEPTIN™ (trastuzumab) (Genentech, Calif.), a humanized anti-HER2 monoclonal antibody for treating patients with metastatic breast cancer, and REOPRO®™, an anti-glycoprotein IIb / IIIa receptor on platelets for preventing blood clot formation.Abciximab (Centocor), ZENAPAX™ (daclizumab), an immunosuppressive humanized anti-CD25 monoclonal antibody for preventing acute renal allograft rejection (Roche Pharmaceuticals, Switzerland), PANOREX™, a murine anti-17-IA cell surface antigen IgG2a antibody (Glaxo Wellcome / Centocor), BEC2 (ImClone System), a murine anti-idiotype (GD3 epitope) IgG antibody, IMC-C225 (ImClone System), a chimeric anti-EGFR IgG antibody, VITAXIN™, a humanized anti-alphaVbeta3 integrin antibody (Applied Molecular Evolution / MedImmune), Campath 1H / LDP-03 (Leukosite), a humanized anti-CD52 IgG1 antibody, Smart M195 (Protein Design Lab / Kanebo), a humanized anti-CD33 IgG antibody, RITUXAN™, a chimeric anti-CD20 IgG1 antibody (IDEC Pharm / Genentech, Roche / Zettyaku), LYMPHOCIDE™, a humanized anti-CD22 IgG antibody (Immunomedics), LYMPHOCIDE™ Y-90 (Immunomedics), Lymphoscan (Tc-99m labeled, radioimaging, Immunomedics), Nuvion (against CD3, Protein Design Labs), CM3 (ICOS Pharm), a humanized anti-ICAM3 antibody, IDEC-114 (IDEC Pharm / Mitsubishi), a primatized anti-CD80 antibody, ZEVALIN™, a radiolabeled murine anti-CD20 antibody (IDEC / Schering AG), IDEC-131 (IDEC / Eisai), a humanized anti-CD40L antibody, IDEC-151 (IDEC), a primatized anti-CD4 antibody, IDEC-152 (IDEC / Seikagaku), a primatized anti-CD23 antibody, SMART anti-CD3 (Protein Design Lab), a humanized anti-CD3 IgG, 5G1, a humanized anti-complement factor 5 (C5) antibody.1 (Alexion Pharm), D2E7 (CAT / BASF), a humanized anti-TNF-alpha antibody, CDP870 (Celltech), a humanized anti-TNF-alpha Fab fragment, IDEC-151 (IDEC Pharm / SmithKline Beecham), a primatized anti-CD4 IgG1 antibody, MDX-CD4 (Medarex / Eisai / Genmab), a human anti-CD4 IgG antibody, CD20 streptavidin (+ biotin-yttrium 90, NeoRx), CDP571 (Celltech), a humanized anti-TNF-alpha IgG4 antibody, LDP-02 (LeukoSite / Genentech), a humanized anti-alpha4beta7 antibody, OrthoClone OKT4A (Ortho Biotech), a humanized anti-CD4 IgG antibody, ANTOVA (trademark) (Biogen), a humanized anti-CD40L IgG antibody, ANTEGREN (trademark) (Elan), a humanized anti-VLA-4 IgG antibody, and CAT-152 (Cambridge Ab Tech), a human anti-TGF-beta antibody. Others are shown in subsequent paragraphs. ta 2 is included. Others are shown in subsequent paragraphs.

[0199] Immunotherapies that can be used in combination with the compounds as disclosed herein include adjuvant immunotherapies. Examples include cytokines such as granulocyte macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), macrophage inflammatory protein (MIP)-1-alpha, interleukins (including IL-1, IL-2, IL-4, IL-6, IL-7, IL-12, IL-15, IL-18, IL-21 and IL-27), tumor necrosis factor (including TNF-alpha) and interferons (including IFN-alpha, IFN-beta and IFN-gamma), aluminum hydroxide (alum), Bacillus Calmette-Guerin (BCG), keyhole limpet hemocyanin (KLH), incomplete Freund's adjuvant (IFA), QS-21, DETOX, levamisole, and dinitrophenyl (DNP), and combinations thereof such as combinations of interleukins, such as combinations of other cytokines such as IL-2 and INF-alpha.

[0200] In various embodiments, an immunological therapy or immunotherapeutic agent may include one or more of the following: adoptive cell transfer, angiogenesis inhibitors, Bacillus Calmette-Guerin therapy, biochemical therapy, cancer vaccines, chimeric antigen receptor (CAR) T cell therapy, cytokine therapy, gene therapy, immune checkpoint regulators, immune complexes, radio conjugates, oncolytic virus therapy, or targeted drug therapy. The function or at least one function of an immunological therapy or immunotherapeutic agent is generally referred to herein as an "immunotherapeutic agent".

[0201] The present disclosure provides a method for preventing, treating, reducing, inhibiting, or controlling a neoplasm, tumor, or cancer in a subject in need thereof, comprising administering a therapeutically effective amount of a combination comprising a compound of formula I' and an immunotherapeutic agent. In one non-limiting embodiment, the method comprises administering a therapeutically effective amount of a combination comprising a compound of formula I' in combination with an immunotherapeutic agent. In various embodiments, the combination provides a synergistic, additive, or supra-additive effect in reducing the number of cancer cells when administered in combination, as compared to treatment with each agent alone. In some embodiments, administration of a therapeutically effective amount of a combination comprising a compound of formula I' and an immunotherapeutic agent provides anti-tumor activity that is more potent than the supra-additive anti-tumor activity and / or the additive effect of administration of the compound of formula I' or the immunotherapeutic agent alone.

[0202] Human cancers have numerous genetic and epigenetic mutations that generate neoantigens that are potentially recognizable by the immune system (Sjoblom et al. (2006) Science 314:268-74). The adaptive immune system, composed of T and B lymphocytes, has a powerful anti-cancer capacity with a broad ability and exquisite specificity to respond to diverse tumor antigens. Furthermore, the immune system demonstrates considerable plasticity and memory elements. By successfully harnessing all of these contributions of the adaptive immune system, immunotherapy will be unique among all cancer treatment modalities.

[0203] The present disclosure provides a combination of a compound of formula I' and an immunotherapeutic agent. These exemplary combinations can be used to treat a subject having cancer. In various embodiments, the immunotherapeutic agents utilized in the present compositions, formulations, and methods can include one or more agents or treatments including adoptive cell transfer, angiogenesis inhibitors, Bacillus Calmette-Guerin treatment, biochemotherapy, cancer vaccines, chimeric antigen receptor (CAR) T cell therapy, cytokine therapy, gene therapy, immune checkpoint regulators such as immune checkpoint inhibitors, immune complexes, radio-conjugates, oncolytic virus therapy, or targeted drug therapy.

[0204] In certain embodiments of the present disclosure, a therapeutically effective combination comprises a compound of formula I' and an immunotherapeutic agent. In various related embodiments, the compound of formula I' enhances the activity of the immunotherapeutic agent.

[0205] In certain embodiments of each of the above-described aspects, as well as other aspects, and the embodiments described elsewhere herein, the immunotherapeutic agent enhances the activity of the compound of formula I'.

[0206] In certain embodiments of each of the above-described aspects, as well as other aspects, and the embodiments described elsewhere herein, the compound of formula I' and the immunotherapeutic agent act synergistically. In various embodiments described herein, exemplary immunotherapeutic agents are immunocyte (e.g., T cells, dendritic cells, natural killer cells, etc.) regulators selected from agonists or activators of costimulatory molecules, where the regulators are monoclonal antibodies known in the art, bispecific antibodies comprising one or more immune checkpoint antigen-binding portions, trispecific antibodies, or immunocyte-engaging multivalent antibodies / fusion proteins / constructs). In some embodiments, the immunotherapeutic agent may be an antibody that modulates a costimulatory molecule and binds to an antigen on the surface of an immunocyte or a cancer cell. In each of these various embodiments, the antibody regulator may be a monoclonal antibody, a polyclonal antibody, a bispecific antibody, a trispecific or multispecific format antibody, a fusion protein, or a fragment thereof, such as a diabody, a single-chain (sc)-diabody (scFv)2, a miniantibody, a minibody, a barnase-barstar, an scFv-Fc, an sc(Fab)2, a trimeric antibody construct, a tribody antibody construct, a trimerbody antibody construct, a tribody antibody construct, a Collabody antibody construct, an (scFv-TNFa)3, or an F(ab)3 / DNL antibody construct.

[0207] In certain specific embodiments of each of the above-described aspects, and further of other aspects, and in the embodiments described elsewhere in this specification, the immunotherapeutic agent is an agent that modulates an immune response, such as, for example, a checkpoint inhibitor or a checkpoint agonist. In some embodiments, the immunotherapeutic agent is an agent that enhances an anti-tumor immune response. In some embodiments, the immunotherapeutic agent is an agent that increases cellular immunity. In some embodiments, the immunotherapeutic agent is an agent that elevates T cell activity. In some embodiments, the immunotherapeutic agent is an agent that elevates cytotoxic T lymphocyte (CTL) activity. In some embodiments, the immunotherapeutic agent is an antibody modulator that targets, among those known in the art, PD-1, PD-L1, PD-L2, CEACAM (such as, for example, CEACAM-1, -3 and / or -5), CTLA-4, TIM-3, LAG-3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4, TGF beta, OX40, 41BB, LIGHT, CD40, GITR, TGF-beta, TIM-3, SIRP-alpha, VSIG8, BTLA, SIGLEC7, SIGLEC9, ICOS, B7H3, B7H4, FAS, and / or BTNL2. In some embodiments, the immunotherapeutic agent is an agent that elevates natural killer (NK) cell activity. In some embodiments, the immunotherapeutic agent is an agent that inhibits the suppression of an immune response. In some embodiments, the immunotherapeutic agent is an agent that inhibits suppressor cells or suppressor cell activity. In some embodiments, the immunotherapeutic agent is an agent or treatment that inhibits Treg activity. In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of an inhibitory immune checkpoint receptor. In some embodiments, the combination of the present disclosure comprises a compound of formula I' and an immunotherapeutic agent, wherein the immunotherapeutic agent comprises a T cell regulator selected from an agonist or activator of a costimulatory molecule.In one embodiment, the agonist of the costimulatory molecule is an agonist of GITR, OX40, ICOS, SLAM (e.g., SLAMF7), HVEM, LIGHT, CD2, CD27, CD28, CDS, ICAM-1, LFA-1 (CD11a / CD18), ICOS (CD278), 4-1BB (CD137), CD30, CD40, BAFFR, CD7, NKG2C, NKp80, CD160, B7-H3, or CD83 ligand (e.g., an agonist antibody or an antigen-binding fragment thereof, or a soluble fusion). In other embodiments, the effector cell combination includes a bispecific T cell engager (e.g., a bispecific antibody molecule that binds CD3 and a tumor antigen (e.g., particularly EGFR, PSCA, PSMA, EpCAM, HER2)).

[0208] In some embodiments, the immunotherapeutic agent is a regulator of PD-1 activity, a regulator of PD-L1 activity, a regulator of PD-L2 activity, a regulator of CTLA-4 activity, a regulator of CD28 activity, a regulator of CD80 activity, a regulator of CD86 activity, a regulator of 4-1BB activity, a regulator of OX40 activity, a regulator of KIR activity, a regulator of Tim-3 activity, a regulator of LAG3 activity, a regulator of CD27 activity, a regulator of CD40 activity, a regulator of GITR activity, a regulator of TIGIT activity, a regulator of CD20 activity, a regulator of CD96 activity, a regulator of IDO1 activity, a regulator of SIRP-alpha activity, a regulator of TIGIT activity, a regulator of VSIG8 activity, a regulator of BTLA activity, a regulator of SIGLEC7 activity, a regulator of SIGLEC9 activity, a regulator of ICOS activity, a regulator of B7H3 activity, a regulator of B7H4 activity, a regulator of FAS activity, a regulator of BTNL2 activity, a cytokine, a chemokine, an interferon, an interleukin, a lymphokine, a member of the tumor necrosis factor (TNF) family, or an immunostimulatory oligonucleotide. In some embodiments, the immunotherapeutic agent is an immune checkpoint regulator (e.g., an immune checkpoint inhibitor, e.g., an inhibitor of PD-1 activity, a regulator of PD-L1 activity, a regulator of PD-L2 activity, a regulator of CTLA-4, or a CD40 agonist (e.g., an anti-CD40 antibody molecule), (xi) an OX40 agonist (e.g., an anti-OX40 antibody molecule), or (xii) a CD27 agonist (e.g., an anti-CD27 antibody molecule). In one embodiment, the immune regulator is an inhibitor of PD-1, PD-L1, PD-L2, CTLA-4, TIM-3, LAG-3, CEACAM (e.g., CEACAM-1, -3 and / or -5), VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4 and / or TGF beta. In one embodiment, the inhibitor of the immune checkpoint molecule inhibits PD-1, PD-L1, LAG-3, TIM-3, CEACAM (e.g., CEACAM-1, -3 and / or -5), CTLA-4, or any combination thereof.

[0209] Inhibition of inhibitory molecules can be performed at the DNA, RNA, or protein level. In embodiments, inhibitory nucleic acids (e.g., dsRNA, siRNA, or shRNA) can be used to inhibit the expression of inhibitory molecules. In other embodiments, inhibitors of inhibitory signals are polypeptides, such as soluble ligands (e.g., PD-1-Ig or CTLA-4 Ig), or antibodies or antigen-binding fragments thereof, such as monoclonal antibodies, bispecific antibodies containing one or more immune checkpoint antigen-binding portions, trispecific antibodies, or immunocyte-engaging multivalent antibodies / fusion proteins / constructs known in the art that bind to inhibitory molecules, e.g., antibodies or fragments thereof (also referred to herein as "antibody molecules") that bind to PD-1, PD-L1, PD-L2, CEACAM (e.g., CEACAM-1, -3, and / or -5), CTLA-4, TIM-3, LAG-3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4, TGF beta, or combinations thereof.

[0210] In some embodiments, when the combination comprises a compound of Formula I' and an immunotherapeutic agent, the immunotherapeutic agent is a monoclonal antibody or a bispecific antibody. For example, the monoclonal or bispecific antibody can specifically bind to a member of the c-Met pathway and / or an immune checkpoint regulator (e.g., the bispecific antibody binds to both the hepatocyte growth factor receptor (HGFR) and an immune checkpoint regulator described herein, PD-1, PD-L1, PD-L2, or CTLA-4, LAG-3, OX40, 41BB, LIGHT, CD40, GITR, TGF-beta, TIM-3, SIRP-alpha, T IGIT, VSIG8, BTLA, SIGLEC7, SIGLEC9, ICOS, B7H3, B7H4, FAS, BTNL2, or an antibody that binds to CD27, etc.). In certain embodiments, the bispecific antibody specifically binds to the human HGFR protein and one of PD-1, PD-L1, and CTLA-4.

[0211] In some embodiments, the immunotherapeutic agent is a cytokine, such as a chemokine, interferon, interleukin, lymphokine, or a member of the tumor necrosis factor family. In some embodiments, the cytokine is IL-2, IL15, or interferon-gamma.

[0212] In some embodiments of any of the above aspects or the embodiments described elsewhere in this specification, the cancer is selected from the group consisting of lung cancer, pancreatic cancer, breast cancer, colon cancer, colorectal cancer, melanoma, gastrointestinal cancer, gastric cancer, kidney cancer, ovarian cancer, liver cancer, endometrial cancer, kidney cancer, prostate cancer, thyroid cancer, neuroblastoma, glioma, glioblastoma, glioblastoma multiforme, cervical cancer, gastric cancer, bladder cancer, head and neck cancer, and hepatocellular carcinoma.

[0213] In some embodiments of any of the above aspects or the embodiments described elsewhere in this specification, the subject's cancer or tumor does not respond to immune checkpoint inhibition (e.g., to any of the immune checkpoint inhibitors described herein such as a PD-1 antagonist or a PD-L1 antagonist), or the subject's cancer or tumor progresses after an initial response to immune checkpoint inhibition (e.g., to any of the immune checkpoint inhibitors described herein such as a PD-1 antagonist or a PD-L1 antagonist).

[0214] In some embodiments of any of the above aspects or the embodiments described elsewhere in this specification, the subject is human.

[0215] Checkpoint inhibitors can be any molecule, agent, treatment, and / or method that inhibits immune checkpoints, and / or promotes inhibitors of immune checkpoints, for example, by promoting endogenous immune checkpoint inhibitors, by inhibiting transcription factors involved in the expression of immune checkpoints, and / or by acting in cooperation with several additional exogenous factors. For example, checkpoint inhibitors may include treatments that inhibit transcription factors involved in the expression of immune checkpoint genes, or that promote the expression of transcription factors for tumor-suppressor genes, such as BACH2 (Luan et al., (2016). Transcription Factors and Checkpoint Inhibitor Expression with Age: Markers of Immunosenescence. Blood, 128(22), 5983). Furthermore, checkpoint inhibitors can inhibit the transcription of immune checkpoint genes, the modification and / or processing of immune checkpoint mRNAs, the translation of immune checkpoint proteins, and / or the activation of common oncogenic processes of molecules involved in the immune or immune checkpoint pathways, such as PD-1 transcription factors, for example, HIF-1, STAT3, NF-κΒ, and AP-1, or common oncogenic processes such as JAK / STAT, RAS / ERK, or PI3K / AKT / mTOR (Zerdes et al., Genetic, transcriptional and post-translational regulation of the programmed death protein ligand 1 in cancer: biology and clinical correlations, Oncogene volume 37, pages 4639-4661 (2018), the entire disclosure of which is incorporated herein by reference).

[0216] Checkpoint inhibitors can include treatments, molecules, agents, and / or methods that regulate immune checkpoints at the transcriptional level, for example, using RNA interference pathway co-suppression and / or post-transcriptional gene silencing (PTGS) (e.g., microRNA, miRNA, silencing RNA, small interfering RNA, or short interfering RNA (siRNA). Transcriptional regulation of checkpoint molecules has been shown to be related to mir-16, which has been shown to target the 3’UTRs of checkpoint mRNAs CD80, CD274 (PD-L1), and CD40 (Leibowitz et al., Post-transcriptional regulation of immune checkpoint genes by mir-16 in melanoma, Annals of Oncology (2017) 28; v428-v448). Mir-33a has also been shown to be related to the regulation of PD-1 expression in the case of lung adenocarcinoma (Boldini et al., Role of microRNA-33a in regulating the expression of PD-1 in lung adenocarcinoma, Cancer Cell Int. 2017;17:105), the entire disclosure of which is incorporated herein by reference).

[0217] T cell-specific aptamer-siRNA chimeras have been suggested as a highly specific method of inhibiting molecules in the immune checkpoint pathway (Hossain et al., The aptamer-siRNA conjugates: reprogramming T cells for cancer therapy, Ther. Deliv. 2015 Jan;6(1):1-4), the entire disclosure of which is incorporated herein by reference).

[0218] Alternatively, members of the immune checkpoint pathway can be inhibited using treatments that affect related pathways, such as metabolism. For example, the over-supply of the glycolytic intermediate pyruvate in mitochondria from CAD macrophages promoted the expression of PD-L1 via the induction of the bone morphogenetic protein 4 / phosphorylated SMAD1 / 5 / IFN regulatory factor 1 (BMP4 / p-SMAD1 / 5 / IRF1) signaling pathway. Thus, the implementation of treatments that regulate metabolic pathways can result in subsequent regulation of the immune-inhibitory PD-1 / PD-L1 checkpoint pathway (Watanabe et al., Pyruvate controls the checkpoint inhibitor PD-L1 and suppresses T cell immunity, J Clin Invest. 2017 Jun 30;127(7):2725-2738).

[0219] Checkpoint immunity can be regulated by means of oncolytic viruses that selectively replicate within tumor cells and induce an acute immune response in the tumor - microenvironment, i.e., oncolytic viruses acting as gene vectors that carry specific agents (e.g., antibodies, miRNAs, siRNAs, etc.) to cancer cells and cause their tumor lysis as well as the secretion of cytokines and chemokines, in combination with immune checkpoint inhibition (Shi et al., Cancer Immunotherapy: A Focus on the Regulation of Immune Checkpoints, Int J Mol Sci. 2018 May;19(5):1389). Currently, clinical trials are underway using the following viruses as checkpoint inhibitors: poliovirus, measles virus, adenovirus, poxvirus, herpes simplex virus (HSV), coxsackievirus, reovirus, Newcastle disease virus (NDV), T-VEC (herpesvirus encoded together with GM-CSF (granulocyte - macrophage colony-stimulating factor)), and H101 (Shi et al., supra).

[0220] Checkpoint inhibitors can act at the translational level of checkpoint immunity. The translation of mRNA into protein represents an important event in the regulation of gene expression, and thus, inhibition of immune checkpoint translation can be a way to inhibit the immune checkpoint pathway. It is a method that can be used.

[0221] Inhibition of the immune checkpoint pathway can occur at any stage of the immune checkpoint translation process. For example, a drug, molecule, agent, treatment, and / or method can inhibit the initiation process (whereby the 40S ribosomal subunit is recruited to the 5’ end of the mRNA and scans the 5’ UTR of the mRNA towards its 3’ end. Inhibition can occur by targeting the anticodon of the initiator methionyl-transfer RNA (tRNA)(Met-tRNAi), its base pairing with the start codon, or the recruitment of the 60S subunit to initiate amino acid elongation and consecutive addition in the translation of immune checkpoint-specific genes. Alternatively, the checkpoint inhibitor can inhibit at the translational level by preventing the formation of the ternary complex (TC), i.e., eukaryotic translation initiation factor (eIF)2 (or one or more of its α, β, and γ subunits), GTP, and Met-tRNAi.

[0222] Checkpoint inhibition can occur through destabilization of eIF2α by preventing phosphorylation of eIF2α via protein kinase R (PKR), PERK, GCN2, or HRI, or by preventing the TC from associating with the 40S ribosome and / or other initiation factors, thereby preventing the formation of the pre-initiation complex (PIC); by inhibiting the eIF4F complex and / or its cap-binding protein eIF4E, scaffold protein eIF4G, or eIF4A helicase. Methods of discussing translational control of cancer are discussed in Truitt et al., New frontiers in translational control of the cancer genome, Nat Rev Cancer. 2016 Apr 26;16(5):288-304, the entire disclosure of which is incorporated herein by reference.

[0223] Checkpoint inhibitors can also include treatments, molecules, agents, and / or methods that regulate immune checkpoints at the cellular and / or protein level, for example, by inhibiting immune checkpoint receptors. Inhibition of checkpoints can occur through the use of antibodies, antibody fragments, antigen-binding fragments, small molecules, and / or other drugs, agents, treatments, and / or methods.

[0224] Immune checkpoints refer to inhibitory pathways in the immune system that are responsible for maintaining self-tolerance and regulating the degree of immune system response to minimize damage to peripheral tissues. However, tumor cells can also activate immune system checkpoints to reduce the effectiveness of the immune response against tumor tissue (to "block" the immune response). In contrast to most anti-cancer drugs, checkpoint inhibitors do not directly target tumor cells, but rather target lymphocyte receptors or their ligands, in order to enhance the intrinsic anti-tumor activity of the immune system (Pardoll, 2012, Nature Reviews Cancer 12:252-264).

[0225] Until recently, cancer immunotherapy has focused substantial efforts on approaches to enhance the antitumor immune response by adoptive transfer of activated effector cells, immunization against relevant antigens, or provision of nonspecific immunostimulatory agents such as cytokines. However, in the past decade, intensive efforts to develop specific immune checkpoint pathway inhibitors have begun to provide new immunotherapy approaches for treating cancer, including the development of an antibody (Ab) that binds to and inhibits CTLA-4, ipilimumab (YERVOY®), for treating patients with advanced melanoma (Hodi et al. (2010) N Engl J Med 363:711-23), as well as the development of antibodies such as nivolumab and pembrolizumab (previously lambrolizumab, USAN Council S tatement(2013)Pembrolizumab:Statement on a nonproprietary name adopted by the USAN Council(ZZ-165),Nov.27,2013) that specifically bind to the programmed death-1 (PD-1) receptor and block the inhibitory PD-1 / PD-1 ligand pathway (Topalian et al. (2012a) N Engl J Med 366:2443-54; Topalian et al. (2012b) Curr Opin Immunol 24:207-12, Topalian et al. (2014) J Clin Oncol 32(10):1020-30, Hamid et al. (2013) N Engl J Med 369:134-144, Hamid and Carvajal(2013) Expert Opin Biol Ther 13(6):847-61, McDermott and Atkins(2013) Cancer Med 2(5):662-73).

[0226] PD-1 is an important immune checkpoint receptor expressed by activated T and B cells and mediates immunosuppression. Nivolumab (previously called 5C4, BMS-936558, MDX-1106, or ONO-4538) is a fully human IgG4 (S228P) PD-1 immune checkpoint inhibitory antibody that selectively blocks the interaction with PD-1 ligands (PD-L1 and PD-L2), thereby blocking the downregulation of anti-tumor T cell function (U.S. Patent No. 8,008,449, Wang et al. (2014) In vitro characterization of the anti-PD-1 antibody nivolumab,BMS-936558,and in vivo toxicology in non-human primates. Nivolumab is approved for the treatment of patients with unresectable or metastatic melanoma and patients who have disease progression after BRAF inhibitors if BRAF V600 mutation positive, as well as for the treatment of squamous non-small cell lung cancer.

[0227] Recent data suggest a secondary mechanism of anti-CTLA-4 antibodies that can occur within the tumor itself. CTLA-4 has been found to be expressed at high levels in regulatory T cells (also referred to herein as "Treg cells") compared to effector T cells within the tumor (also referred to herein as "Teff cells"), giving rise to the hypothesis that anti-CTLA-4 preferentially and strongly affects Treg cells ("Therapeutic use of anti-CTLA-4 antibodies", Christian U. Blank and Alexander Enk, International Immunology, Vol. 27, No. 1, pp. 3-10). Recent studies of the combination of PD-1 and CTLA-4 have shown that the combined blockade of the CTLA-4 and PD-1 pathways also cooperate to increase the ratio of Teff cells to both regulatory T cells and MDSC, thereby reducing suppression and promoting inflammation in the tumor microenvironment (the entire disclosure of which is incorporated herein by reference, "Combination of CTLA-4 and PD-1 blockade expands infiltrating T-cells and reduces regulatory T and myeloid cells within B16 melanoma tumors", Curran et al., PNAS | Mar. 2, 2010; vol. 107(no. 9); pp. 4275-4280). Combinations of checkpoint inhibitors and other therapeutic agent(s) can enhance and / or prolong the anti-tumor response of the checkpoint inhibitor and / or the effect of the therapeutic agent. In this regard, WO2015 / 069770 discloses combination treatments based on the activation of an adaptive immune response for treating cancer, specifically, combinations of CTLA-4 and PD-1 inhibitors. The disclosure of WO2015 / 069770 is incorporated herein by reference in its entirety into the disclosure of the present application.

[0228] One mechanism by which checkpoint-blocking anti-CTLA-4 antibodies mediate an anti-tumor effect is by reducing regulatory T cells. By a distinct mechanism of action of the anti-CTLA-4 antibody, anti-C The TLA-4 antibody can be successfully combined with an anti-PD1 checkpoint blockade antibody that acts to relieve the inhibitory signaling imparted to effector T cells. Dual blockade with these antibodies has jointly improved antitumor responses in both preclinical (Proc Natl Acad Sci USA 2010,107,4275-4280) and clinical (N Engl J Med 2013,369,122-133; N Engl J Med 2015,372,2006-2017) settings.

[0229] CTLA-4 attenuates the initial activation of naive and memory T cells via its interaction with the ligands B7-1 (CD80) and B7-2 (CD86) (Figure 1A). PD-1 is a receptor expressed on the surface of activated mature T cells, activated NK cells, B cells, monocytes, and many normal tissues and plays an important role in the maintenance of peripheral tolerance [20-21] (Figure 1A). In contrast to CTLA-4, PD-1 acts via its interaction with the ligand PD-L1 (also known as B7-H1 or CD274), is mainly involved in the regulation of T cell activity in peripheral tissues, and further provides a major immune tolerance mechanism within the tumor microenvironment.

[0230] In some embodiments, the immunotherapeutic agent is a regulator of PD-1 activity, a regulator of PD-L1 activity, a regulator of PD-L2 activity, a regulator of CTLA-4 activity, a regulator of CD28 activity, a regulator of CD80 activity, a regulator of CD86 activity, a regulator of 4-1BB activity, a regulator of OX40 activity, a regulator of KIR activity, a regulator of Tim-3 activity, a regulator of LAG3 activity, a regulator of CD27 activity, a regulator of CD40 activity, a regulator of GITR activity, a regulator of TIGIT activity, a regulator of CD20 activity, a regulator of CD96 activity, a regulator of IDO1 activity, a cytokine, a chemokine, an interferon, an interleukin, a lymphokine, a member of the tumor necrosis factor (TNF) family, or an immunostimulatory oligonucleotide. In some embodiments, the immune checkpoint regulator is, i.e., an inhibitor or antagonist, or an activator or agonist, e.g., a CD28 regulator, a 4-1BB regulator, an OX40 regulator, a CD27 regulator, a CD80 regulator, a CD86 regulator, a CD40 regulator, or a GITR regulator, a Lag-3 regulator, a 41BB regulator, a LIGHT regulator, a CD40 regulator, a GITR regulator, a TGF-beta regulator, a TIM-3 regulator, a SIRP-alpha regulator, a TIGIT regulator, a VSIG8 regulator, a BTLA regulator, a SIGLEC7 regulator, a SIGLEC9 regulator, an ICOS regulator, a B7H3 regulator, a B7H4 regulator, a FAS regulator, and / or a BTNL2 regulator. In some embodiments, the immunotherapeutic agent is an immune checkpoint regulator as described above (e.g., a monoclonal antibody known in the art, a bispecific antibody comprising one or more immune checkpoint antigen-binding portions, a trispecific antibody, or an immune checkpoint regulatory antibody which may be in the form of an immunocyte-engaging multivalent antibody / fusion protein / construct).

[0231] Combination treatments with immune checkpoint inhibitory immunotherapeutic agents that may include antibodies that specifically target immune system checkpoints such as CTLA4, PD1, and PD-L1 are one of the most promising new avenues for immunotherapy for cancer and other diseases. Additional checkpoint targets such as TIM-3, LAG-3, various B-7 ligands, CHK1 and CHK2 kinases, BTLA, A2aR, etc. are also under investigation. Currently, three checkpoint inhibitors, including ipilimumab (Yervoy®), a CTLA-4 inhibitor, and pembrolizumab (Keytruda®) and nivolumab (Opdivo®), both of which are PD-1 inhibitors, have received early approval from the US Food and Drug Administration for cancer treatment. In addition, several checkpoint inhibitors are in clinical trials.

[0232] Programmed cell death protein 1, (PD-1 or CD279), a 55-kD type I transmembrane protein, is a member of the CD28 family of T cell costimulatory receptors that includes CD28, CTLA- 4, inducible costimulatory molecule (ICOS), and BTLA. PD-1 is highly expressed on activated T cells and B cells. PD-1 expression can also be detected at various levels on subsets of memory T cells. Two ligands specific for PD-1 have been identified: programmed death-ligand 1 (PD-L1, also known as B7-H1 or CD274) and PD-L2 (also known as B7-DC or CD273). PD-L1 and PD-L2 have been shown to downregulate T cell activation when they bind to PD-1 in both mouse and human systems (Okazaki et al., Int Immunol., 2007;19:813-824). The interaction of PD-1 with its ligands, PD-L1 and PD-L2, expressed on antigen-presenting cells (APCs) and dendritic cells (DCs), transmits a negative regulatory stimulus to downregulate the activated T cell immune response. Blockade of PD-1 suppresses this negative signal and amplifies the T cell response.

[0233] Numerous studies have shown that the cancer microenvironment manipulates the PD-L1- / PD-1 signaling pathway, and the induction of PD-L1 expression is associated with the inhibition of the immune response against cancer, enabling cancer progression and metastasis. The PD-L1 / PD-1 signaling pathway is, for several reasons, the primary mechanism of cancer immune evasion. First and most importantly, this pathway is involved in the negative regulation of the immune response of activated T effector cells found peripherally. Second, while PD-L1 is upregulated in the cancer microenvironment, PD-1 is also upregulated in activated tumor-infiltrating T cells, which can strengthen the vicious cycle of inhibition. Third, this pathway is intricately related to both innate and adaptive immune regulation through bidirectional signaling. Due to these factors, the PD-1 / PD-L1 complex has become the central point where cancer can manipulate the immune response and promote its own progression.

[0234] CTLA-4 (also known as cytotoxic T lymphocyte-associated protein 4, CTLA4, CTLA-4, CD152, surface antigen classification 152, ALPS5, CD, CELIAC3, GRD4, GSE, and IDDM12). CTLA-4 is a single-pass type I membrane protein of approximately 24.6 kDa that plays an inhibitory role in T cell function. CTLA-4 was originally identified by differential screening of a mouse cytotoxic T cell cDNA library (see Brunet et al., A new member of the immunoglobulin superfamily--CTLA-4, Nature. 1987 Jul 16-22;328(6127):267-70). CTLA- has been shown to interact with the b7 family ligands CD80 (also known as surface antigen classification 80 and B7-1), and CD86 (also known as surface antigen classification 86 or B7-2). Linsley et al., CTLA-4 is a second receptor for the B cell activation antigen B7, see J Exp Med. 1991 Sep 1;174(3):561-9. Sequence comparison between the region encoding human CTLA-4 DNA and the region encoding CD28 has revealed significant homology between the two sequences, along with a maximum similarity between the membrane-proximal region and the cytoplasmic region. Thus, CTLA-4 is involved in the elimination / reduction of T cell activity and antagonizes the activity of CD28. CTLA-4-deficient mice have been shown to exhibit massive lymphocyte proliferation. Chambers et al., Lymphoproliferation in CTLA-4-deficient mice is mediated by costimulation-dependent activation of CD4+ T cells, Immunity. 1997 Dec;7(6):885-95. CTLA-4 blockade has been reported to enhance T cell responses both in vitro and in vivo, enhance induced autoimmune diseases, and exacerbate antitumor immunity (Luhder, J. Exp. Med. 1998;187:427-432, Walunas et al., Immunity. 1994;1:405-413, Kearney, J. Immunol. 199 5;155:1032-1036), Leach, Science 1996;271:1734-1736). CTLA-4 has also been reported to have alternative and / or additional effects on the initial characteristics of the T cell immune response (Chambers, Curr. Opin. Immunol. 1997;9:396-404, Bluestone, J. Immunol. 1997;158:1989-1993, Thompson, Immunity 1997;7:445-450).

[0235] The first immune checkpoint inhibitor to be tested in clinical trials was ipilimumab (Yervoy, Bristol-Myers Squibb), a CTLA-4 mAb. CTLA-4 belongs to the immunoglobulin superfamily of receptors that also includes PD-1, BTLA, TIM-3, and the V-domain immunoglobulin suppressor of T cell activation (VISTA). Anti-CTLA-4 mAbs are potent checkpoint inhibitors that release the "brake" from both naive and antigen-experienced cells. Treatment enhances the antitumor function of CD8+ T cells, increases the ratio of CD8+ T cells to Foxp3+ T regulatory cells, and inhibits the suppressive function of T regulatory cells. The main drawback of anti-CTLA-4 mAb treatment is the generation of autoimmune toxicity due to off-target effects on the overproliferative immune system that has lost its ability to reduce itself. Up to 25% of patients treated with ipilimumab develop severe grade 3-4 adverse events / autoimmune-type side effects including dermatitis, enterocolitis, hepatitis, endocrine disorders (including hypophysitis, thyroiditis, and adrenalitis), arthritis, uveitis, nephritis, and aseptic meningitis. In contrast to anti-CTLA-4 experience, anti-PD-1 treatment is considered to be better tolerated and induces a relatively low rate of autoimmune-type side effects.

[0236] In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of PD-1. In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of PD-L1 and / or PD-L2. In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of CTLA-4. In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of CD80 and / or CD86. In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of TIGIT. In some embodiments, the immunotherapeutic agent is an agent that inhibits the activity of KIR. In some embodiments, the immunotherapeutic agent is an agent that enhances or stimulates the activity of activating an immune checkpoint receptor.

[0237] In some embodiments of the methods described herein, the immunotherapeutic agent is a PD-1 antagonist, a PD-L1 antagonist, a PD-L2 antagonist, a CTLA-4 antagonist, a CD80 antagonist, a CD86 antagonist, a KIR antagonist, a Tim-3 antagonist, a LAG3 antagonist, a TIGIT antagonist, a CD20 antagonist, a CD96 antagonist, or an IDO1 antagonist.

[0238] In some embodiments, the PD-1 antagonist is an antibody that specifically binds to PD-1. In some embodiments, the antibody that binds to PD-1 is pembrolizumab (KEYTRUDA®, MK-3475, Merck), pidilizumab (CT-011, Curetech Ltd.), nivolumab (OPDIVO®, BMS-936558, MDX-1106, Bristol Myer Squibb), MEDI0680 (AMP-514, AstraZenenca / MedImmune), REGN2810 (Regeneron Pharmaceuticals), BGB-A317 (BeiGene Ltd.), PDR-001 (Novartis), or STI-A1110 (Sorrento Therapeutics). In some embodiments, the antibody that binds to PD-1 is described in PCT Publication WO2014 / 179664 and is, for example, an antibody identified as APE2058, APE1922, APE1923, APE1924, APE1950, or APE1963 (Anaptysbio), or an antibody containing the CDR regions of any of these antibodies. In other embodiments, the P D-1 antagonist is a fusion protein comprising the extracellular domain of PD-L1 or PD-L2, for example, AMP-224 (AstraZeneca / MedImmune). In other embodiments, the PD-1 antagonist is a peptide inhibitor, for example, AUNP-12 (Aurigene).

[0239] In some embodiments, the PD-L1 antagonist is an antibody that specifically binds to PD-L1. In some embodiments, the antibody that binds to PD-L1 is atezolizumab (RG7446, MPDL3280A, Genentech), MEDI4736 (AstraZeneca / MedImmune), BMS-936559 (MDX-1105, Bristol Myers Squibb), avelumab (MSB0010718C, Merck KGaA), KD033 (Kadmon), the antibody portion of KD033, or STI-A1014 (Sorrento Therapeutics). In some embodiments, the antibody that binds to PD-L1 is described in PCT Publication WO2014 / 055897, the entire disclosure of which is incorporated herein by reference, and includes, for example, Ab-14, Ab-16, Ab-30, Ab-31, Ab-42, Ab-50, Ab-52, or Ab-55, or an antibody containing the CDR regions of any of these antibodies.

[0240] In some embodiments, the CTLA-4 antagonist is an antibody that specifically binds to CTLA-4. In some embodiments, the antibody that binds to CTLA-4 is ipilimumab (YERVOY®, Bristol Myer Squibb) or tremelimumab (CP-675,206, Pfizer). In some embodiments, the CTLA-4 antagonist, CTLA-4 fusion protein or soluble CTLA-4 receptor, such as, KARR-102 (Kahr Medical Ltd.).

[0241] In some embodiments, the LAG3 antagonist is an antibody that specifically binds to LAG3. In some embodiments, antibodies that bind to LAG3 are IMP701 (Prima BioMed), IMP731 (Prima BioMed / GlaxoSmithKline), BMS-986016 (Bristol Myer Squibb), LAG525 (Novartis), and GSK2831781 (GlaxoSmithKline). In some embodiments, the LAG3 antagonist comprises a soluble LAG3 receptor, such as IMP321 (Prima BioMed).

[0242] In some embodiments, the KIR antagonist is an antibody that specifically binds to KIR. In some embodiments, the antibody that binds to KIR is lirilumab (Bristol Myer Squibb / Innate Pharma).

[0243] In some embodiments, the immunotherapeutic agent used in the combinations disclosed herein (e.g., in combination with a compound of Formula I’) is an activator or agonist of a co-stimulatory molecule. In one embodiment, the agonist of the co-stimulatory molecule is selected from agonists (e.g., agonist antibodies or antigen-binding fragments thereof, or soluble fusions) of OX40, CD2, CD27, CD28, CDS, ICAM-1, LFA-1 (CD11a / CD18), ICOS (CD278), 4-1BB (CD137), GITR, CD30, CD40, BAFFR, HVEM, CD7, LIGHT, NKG2C, SLAMF7, NKp80, CD160, B7-H3, or the CD83 ligand.

[0244] In some embodiments, the OX40 agonist comprises an OX40 ligand, or an OX40-binding portion thereof. For example, the OX40 agonist may be MEDI6383 (AstraZeneca). In some embodiments, the OX40 agonist is an antibody that specifically binds to OX40. In some embodiments, the antibody that binds to OX40 is MEDI6469 (AstraZeneca / MedImmune), MEDI0562 (As is traZeneca / MedImmune), or MOXR0916 (RG7888, Genentech). In some embodiments, the OX40 agonist is a vector capable of expressing the OX40 ligand (e.g., an expression vector or a virus, such as an adenovirus). In some embodiments, the OX40 expression vector is Delta-24-RGDOX (DNAtrix) or DNX2401 (DNAtrix).

[0245] In some embodiments, the 4-1BB (CD137) agonist is a binding molecule such as an anti-carin. In some embodiments, the anti-carin is PRS-343 (Pieris AG). In some embodiments, the 4-1BB agonist is an antibody that specifically binds to 4-1BB. In some embodiments, the antibody that binds to 4-1BB is PF-2566 (PF-05082566, Pfizer) or urelumab (BMS-663513, Bristol Myer Squibb).

[0246] In some embodiments, the CD27 agonist is an antibody that specifically binds to CD27. In some embodiments, the antibody that binds to CD27 is balstilimab (CDX-1127, Celldex).

[0247] In some embodiments, the GITR agonist comprises the GITR ligand or a GITR-binding portion thereof. In some embodiments, the GITR agonist is an antibody that specifically binds to GITR. In some embodiments, the antibody that binds to GITR is TRX518 (GITR, Inc.), MK-4166 (Merck), or INBRX-110 (Five Prime Therapeutics / Inhibrx).

[0248] TIM-3 has been identified as another important inhibitory receptor expressed by dysfunctional CD8+ T cells. In mouse models of cancer, many dysfunctional tumor-infiltrating CD8+ T cells have been found to actually co-express PD-1 and TIM-3.

[0249] LAG-3 is another recently identified inhibitory receptor that acts to limit effector T cell function and enhance the suppressive activity of regulatory T cells. Recently, it has been shown that PD-1 and LAG-3 are widely co-expressed by tumor-infiltrating T cells in mice, and that combined blockade of PD-1 and LAG-3 induces a potent synergistic anti-tumor immune response in murine models of cancer.

[0250] Blocking the PD-1 pathway can be combined with a vaccine or other compound of formula I', an antibody, for improved therapeutic efficacy (Hirano, F. et al, Cancer Res., 65(3):1089-1096(2005), Li, B. et al, Clin. Cancer Res., 15:1507-1509(2009), and Curran, M. A. et al, Proc. Natl. Acad. Set, 107(9):4275-4280(2010)).

[0251] In some embodiments, immunotherapeutic agents useful in the compositions and methods described herein can include monoclonal antibodies known in the art that specifically target both PD-1 and its ligand PD-L1, bispecific antibodies, trispecific antibodies, or immune cell-engaging multivalent antibodies / fusion proteins / constructs that include one or more immune checkpoint antigen-binding portions.

[0252] PD-1 (programmed death 1, also known as CD279, PDCD1) is a cell surface receptor that plays a critical role in regulating the balance between stimulatory and inhibitory signals in the immune system and maintaining peripheral tolerance (Ishida, Y et al. 1992 EMBO J. 11 3887, Kier, Mary E et al. 20 08 Annu Rev Immunol 26 677-704, Okazaki, Taku et al. 2007 International Immunology 19 813-824). PD-1 is an inhibitory member of the immunoglobulin superfamily that has homology to CD28. The structure of PD-1 consists of a monomeric type I transmembrane protein with one immunoglobulin variable-like extracellular domain and a cytoplasmic domain containing an immunoreceptor tyrosine-based inhibitory motif (ITIM) and an immunoreceptor tyrosine-based switch motif (ITSM). The expression of PD-1 is inducible in T cells, B cells, natural killer (NK) cells, and monocytes upon lymphocyte activation, for example, via T cell receptor (TCR) or B cell receptor (BCR) signaling (Kier, Mary E et al. 2008 Annu Rev Immunol 26 677-704, Agata, Y et al 1996 Int Immunol 8 765-72). PD-1 is a receptor for the ligands CD80, CD86, PD-L1 (B7-H1, CD274), and PD-L2 (B7-DC, CD273), which are cell surface-expressed members of the B7 family (Freeman, Gordon et al. 2000 J Exp Med 192 1027, Latchman, Y et al. 2001 Nat Immunol 2 261). When the ligand engages, PD-1 recruits phosphatases such as SHP-1 and SHP-2 to the intracellular tyrosine motifs of PD-1, and then dephosphorylates effector molecules activated by TCR or BCR signaling (Chemnitz, J et al. 2004 J Immunol 173 945-954, Riley, James L 2009 Immunological Reviews 229 114-125). Thus, PD-1 transmits inhibitory signals to T and B cells only when it engages simultaneously with the TCR or BCR.

[0253] PD-1 has been demonstrated to downregulate effector T cell responses through both intrinsic and extrinsic cellular mechanisms. Inhibitory signaling through PD-1 induces a state of unresponsiveness in T cells, resulting in cells that are unable to clonally expand or produce optimal levels of effector cytokines. PD-1 can also induce apoptosis in T cells through its ability to suppress survival signals from co-stimulation, leading to a decrease in the expression of important anti-apoptotic molecules such as Bcl-XL (Kier, Mary E et al. 2008 Annu Rev Immunol 26 677-704). In addition to these direct effects, recent publications have implicated PD-1 in the suppression of effector cells by promoting the induction and maintenance of regulatory T cells (TREG). For example, PD-L1 expressed on dendritic cells has been shown to synergize with TGF-β to promote the induction of CD4+ FoxP3+ TREG with enhanced suppressive function (Francisco, Loise M et al. 2009 J Exp Med 206 3015-3029).

[0254] TIM-3 (also known as T cell immunoglobulin and mucin-domain containing-3, TIM-3, hepatitis A virus cellular receptor 2, HAVCR2, HAVcr-2, KIM-3, TIMD-3, TIMD3, Tim-3, and CD366) is a single-pass type I membrane protein of approximately 33.4 kDa that is involved in the immune response (Sanchez-Fueyo et al., Tim-3 inhibits T helper type 1-mediated auto- and alloimmune responses and promotes immunological tolerance, Nat. Immunol. 4:1093-1101 (2003)).

[0255] TIM-3 is selectively expressed on Th1 cells and phagocytes (e.g., macrophages and dendritic cells). The use of siRNA or blocking antibodies to reduce human expression resulted in increased secretion of interferon γ (IFN-γ) from CD4+ T cells, which implies an inhibitory role of TIM-3 in human T cells. Analysis of clinical samples from autoimmune disease patients showed no expression of TIM-3 in CD4+ cells. In particular, the expression level of TIM-3 in T cell clones obtained from the cerebrospinal fluid of patients with multiple sclerosis was lower than that in clones obtained from normal healthy humans, and the secretion of IFN-γ was higher (Koguchi K et al., J Exp Med. 203:1413-8. (2006)).

[0256] TIM-3 is a receptor for the ligands galectin-9 (a member of the galectin family that is ubiquitously expressed on various cell types and binds to β-galactoside), phosphatidylserine (PtdSer) (DeKryff et al., T cell / transmembrane, Ig, and mucin-3 allelic variants differentially recognize phosphatidylserine and mediate phagocytosis of apoptotic cells, J Immunol. 2010 Feb 15;184(4):1918-30), high mobility group protein 1 (HMGB1, also known as HMG1, HMG3, SBP-1, HMG-1, and high mobility group box 1) (Chiba et al., Tumor-infiltrating DCs suppress nucleic acid-mediated innate immune responses through interactions between the receptor TIM-3 and the alarmin HMGB1, Nat Immunol. 2012 Sep;13(9):832-42), and carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1, also known as BGP, BGP1, BGPI, carcinoembryonic antigen-related cell adhesion molecule 1) (Huang et al., CEACAM1 regulates TIM-3-mediated tolerance and exhaustion, Nature. 2015 Jan 15;517(7534):386-90).

[0257] BTLA (B and T lymphocyte attenuator, BTLA1, CD272, and also known as B and T lymphocyte associated) is a single-pass type I membrane protein of approximately 27.3 kDa that is involved in lymphocyte inhibition during the immune response. BTLA is constitutively expressed in both B and T cells. BTLA interacts with HVEM (herpesvirus entry mediator), a member of the tumor necrosis factor receptor (TNFR) family (Gonzalez et al., Proc. Natl. Acad. Sci. USA, 2005, 102:1116 - 21). The interaction between BTLA, which belongs to the CD28 family of the immunoglobulin superfamily, and HVEM, a co-stimulatory tumor necrosis factor (TNF) receptor (TNFR), is unique in that they define cross-talk between these two receptor families. BTLA contains a membrane-proximal immunoreceptor tyrosine-based inhibitory motif (ITIM) and a membrane-distal immunoreceptor tyrosine-based switch motif (ITSM). Disruption of either the ITIM or the ITSM eliminates the ability of BTLA to recruit either SHP1 or SHP2, suggesting that BTLA recruits SHP1 and SHP2 by a mechanism distinct from PD-1 and that both tyrosine motifs are required to block T cell activation. The BTLA cytoplasmic tail also contains a third conserved tyrosine-containing motif within the cytoplasmic domain whose sequence is similar to the Grb-2 recruitment site (YXN). Also, a phosphorylated peptide containing this BTLA N-terminal tyrosine motif can interact in vitro with the p85 subunit of GRB2 and PI3K, although the functional effect of this interaction has not yet been investigated in vivo (Gavrieli et al., Biochem. Biophysi Res Commun, 2003, 312, 1236 - 43). BTLA is a receptor for the ligands PTPN6 / SHP-1, PTPN11 / SHP-2, TNFRSF14 / HVEM, and B7H4.

[0258] VISTA (V-domain Ig suppressor of T cell activation VSIR, B7-H5, B 7H5, GI24, PP2135, SISP1, DD1 alpha, VISTA, C10orf54 (also known as chromosome 10 open reading frame 54), PD-1H, and known as a V-set immunoregulatory receptor) is a single-pass type I membrane protein of approximately 33.9 kDa related to T cell inhibitory responses, embryonic stem cell differentiation via BMP4 signaling inhibition, and MMP14-mediated MMP2 activation (Yoon et al., Control of signaling-mediated clearance of apoptotic cells by the tumor suppressor p53, Science. 2015 Jul 31;349(6247):1261669). VISTA interacts with the ligand VSIG-3 (Wang et al., VSIG-3 as a ligand of VISTA inhibits human T-cell function, Immunology. 2019 Jan;156(1):74-85).

[0259] LAG-3 (also known as lymphocyte activation gene 3, LAG3, CD223, and lymphocyte activation 3) is a single-pass type I membrane protein of approximately 57.4 kDa related to lymphocyte activation that also binds to HLA class-II antigens. LAG-3 is a member of the immunoglobulin supergene family and is expressed on activated T cells (Huard et al., 1994, Immunogenetics 39:213), NK cells (Triebel et al., 1990, J. Exp. Med. 171: 1393 - 1405), regulatory T cells (Huang et al., 2004, Immunity 21: 503 - 513, Camisaschi et al., 2010, J Immunol. 184: 6545 - 6551, Gagliani et al., 2013, Nat Med 19: 739 - 746), and plasmacytoid dendritic cells (DC) (Workman et al., 2009, J Immunol 182: 1885 - 1891). LAG - 3 is a membrane protein encoded by a gene located on chromosome 12 and is structurally and genetically related to CD4. Similar to CD4, LAG - 3 can interact with MHC class II molecules on the cell surface (Baixeras et al., 1992, J. Exp. Med. 176: 327 - 337, Huard et al., 1996, Eur. J. Immunol. 26: 1180 - 1186). Direct binding of LAG - 3 to MHC class II has been suggested to play a role in the down - regulation of antigen - dependent stimulation of CD4+ T lymphocytes (Huard et al., 1994, Eur. J. Immunol. 24: 3216 - 3221), and LAG - 3 blockade has been shown to re - activate CD8+ lymphocytes in both tumor or self - antigen (Gross et al., 2007, J Clin Invest. 117: 3383 - 3392) and viral models (Blackburn et al., 2009, Nat. Immunol. 10: 29 - 37). Furthermore, the intracellular region of LAG - 3 can interact with LAP (LAG - 3 - associated protein), a signaling molecule involved in the down - regulation of the CD3 / TCR activation pathway (Iouzalen et al., 2001, Eur. J. Immunol. 31: 2885 - 2891). Additionally, CD4+CD25+ regulatory T cells (Treg) have been shown to express LAG - 3 upon activation, which contributes to the suppressor activity of Treg cells (Huang, C. et al., 2004, Immunity 21: 503 - 513).LAG-3 can also negatively regulate T cell homeostasis by Treg cells in both T cell-dependent and -independent mechanisms (Workman, C.J. and Vignali, D.A., 2005, J. Immunol. 174:688-695).

[0260] LAG-3 has been shown to interact with MHC class II molecules (Huard et al., CD4 / major histocompatibility complex class II interaction analyzed with CD4- and lymphocyte activation gene-3 (LA G-3)-Ig fusion proteins, Eur J Immunol. 1995 Sep;25(9):2718-21).

[0261] In addition, several kinases are known to be checkpoint inhibitors. For example, CHEK-1, CHEK-2, and A2aR.

[0262] CHEK-1 (also known as CHK1 kinase, CHK1, and checkpoint kinase 1) is a serine / threonine-protein kinase of approximately 54.4 kDa that is involved in checkpoint-mediated cell cycle arrest and the activation of DNA repair in response to DNA damage and / or unreplicated DNA.

[0263] CHEK-2 (also known as CHK2 kinase, CDS1, CHK2, HuCds1, LFS2, PP1425, RAD53, hCds1, and checkpoint kinase 2) is a serine / threonine-protein kinase of approximately 60.9 kDa that is involved in checkpoint-mediated cell cycle arrest, DNA repair activation, and double-strand break-mediated apoptosis.

[0264] A2aR (also known as adenosine A2A receptor, ADORA2A, adenosine A2a receptor, A2aR, ADORA2, and RDC8) is a multi-pass membrane receptor of approximately 44.7 kDa for adenosine and other ligands.

[0265] In various embodiments, the immunotherapeutic agent can include an antibody or an antigen-binding fragment thereof. Within this definition, immune checkpoint inhibitors include bispecific antibodies and immune cell-engaging multivalent antibodies / fusion proteins / constructs known in the art. In some embodiments, an immunotherapeutic agent that includes a bispecific antibody is bivalent and can include a bispecific antibody that binds to either the same epitope of an immune checkpoint molecule, two different epitopes of the same immune checkpoint molecule, or different epitopes of two different immune checkpoints.

[0266] One of ordinary skill in the art can realize several bispecific antibody formats known in the art that target one or more of CTLA4, PD1, PD-L1 TIM-3, LAG-3, various B-7 ligands, B7H3, B7H4, CHK1 and CHK2 kinases, BTLA, A2aR, OX40, 41BB, LIGHT, CD40, GITR, TGF-beta, SIRP-alpha, TIGIT, VSIG8, SIGLEC7, SIGLEC9, ICOS, FAS, BTNL2, and others for use in the combinations described herein.

[0267] In various embodiments, the immunotherapeutic agent can include an immune cell-engaging multivalent antibody / fusion protein / construct.

[0268] In one embodiment of the present disclosure, a checkpoint inhibitor is combined with a compound of Formula I' to reduce, or inhibit, the formation or establishment of a primary tumor or cancer metastasis to other sites, or metastatic tumors or cancers at other sites distal to the primary tumor or cancer, thereby inhibiting, or reducing, tumor or cancer recurrence or tumor or cancer progression.

[0269] In a further embodiment of the present disclosure, provided is a combination therapy for treating cancer, which includes enhancing the therapeutic effect and having more manageable toxicity, and blocking a checkpoint inhibitor having the ability to induce a strong and persistent immune response, in combination with a compound of formula I'.

[0270] In a further embodiment of the present disclosure, provided is a combination therapy for treating cancer, which includes a compound of formula I' and an immune checkpoint inhibitor. In one embodiment of the present disclosure, formula I' Provided is a method for treating cancer and / or preventing the establishment of metastasis by using a checkpoint inhibitor that acts synergistically with a compound of formula I'.

[0271] In a further embodiment, the method of the present disclosure includes one or more of the following: 1) reducing or inhibiting the growth, proliferation, motility or invasiveness of tumors or cancer cells that may cause or are causing metastasis; 2) reducing or inhibiting the formation or establishment of metastases that occur from a primary tumor or cancer to one or more other sites, locations or regions separate from the primary tumor or cancer; 3) reducing or inhibiting the growth or proliferation of metastases at one or more other sites, locations or regions separate from the primary tumor or cancer after the metastases have formed or been established; 4) reducing or inhibiting the formation or establishment of additional metastases after the metastases have formed or been established; 5) prolonging the overall survival period; 6) prolonging the progression-free survival, or 7) disease stabilization.

[0272] In one embodiment of the present disclosure, the administration of an immunotherapeutic agent in combination with a compound of formula I' results in a detectable or measurable improvement in the condition of a given subject, such as a remission or alleviation of one or more adverse (physical) symptoms or outcomes associated with cell proliferativity or hyperproliferative disorders, neoplasms, tumors or cancers, or the presence of metastases, i.e., a therapeutic or beneficial effect.

[0273] A therapeutic or beneficial effect is any objective or subjective transient, temporary, or long-term improvement in a condition or pathology, or a decrease in the onset, severity, duration, or frequency of adverse symptoms associated with or resulting from a cell proliferation or cell over-proliferation disorder such as a neoplasm, tumor, or cancer, or metastasis. These can lead to an improvement in survival rate. A satisfactory clinical endpoint of a treatment method according to the present disclosure is, for example, a gradual or partial decrease in the severity, duration, or frequency of one or more associated pathologies, adverse symptoms, or complications, or an inhibition or reversal of one or more physiological, biochemical, or cellular symptom manifestations or characteristics of a cell proliferation or cell over-proliferation disorder such as a neoplasm, tumor, or cancer, or metastasis. Thus, a therapeutic effect or improvement can be, but is not limited to, the destruction of target proliferating cells (e.g., a neoplasm, tumor, or cancer, or metastasis), or the elimination of one or more, most, or all of the pathologies, adverse symptoms, or complications associated with or resulting from a cell proliferative or cell over-proliferative disorder such as a neoplasm, tumor, or cancer, or metastasis. However, a therapeutic effect or improvement does not necessarily require the cure or complete destruction of all target proliferating cells (e.g., a neoplasm, tumor, or cancer, or metastasis) or the elimination of all pathologies, adverse symptoms, or complications associated with or resulting from a cell proliferation or cell over-proliferative disorder such as a neoplasm, tumor, or cancer, or metastasis. For example, partial destruction of the cell mass of a tumor or cancer by inhibition of the progression or worsening of the tumor or cancer, or stabilization of the amount, size, or cell number of the tumor or cancer, can reduce mortality and extend lifespan even if a portion or most of the amount, size, or cells of the tumor or cancer remain.

[0274] Specific non-limiting examples of a therapeutic effect include a decrease in the volume (size or cell mass) or cell number of a neoplasm, tumor, or cancer, or metastasis, inhibition or prevention (e.g., stabilization) of an increase in the volume of a neoplasm, tumor, or cancer, delay or inhibition of the progression, worsening, or metastasis of a neoplasm, tumor, or cancer, or inhibition of the proliferation, growth, or metastasis of a neoplasm, tumor, or cancer.

[0275] In one embodiment of the present disclosure, administration of an immunotherapeutic agent in combination therapy with a compound of Formula I' provides (i) complete disappearance of all lesions and the absence of new lesions (confirmed by repeated serial evaluations four weeks or more after the initially recorded date), regardless of whether irCR is measurable or not, and (ii) an irPR = a reduction in tumor burden of 50% or more compared to baseline detectable or measurable improvement or complete response according to irRC (obtained from time point response assessment and based on tumor burden), including one or more of (confirmed by serial evaluations at least four weeks after the first record), providing.

[0276] Optionally, any of the methods described herein may not produce immediate efficacy. For example, after treatment, an increase in the number or amount of neoplastic, tumor or cancer cells may continue, but over time, eventual stabilization or decrease in the amount, size or number of tumor cells in a given subject may subsequently occur.

[0277] Additional adverse symptoms and complications associated with neoplasms, tumors, cancers and metastases that can be inhibited, reduced, decreased, delayed or prevented include, for example, nausea, anorexia, lethargy, pain and discomfort. Thus, partial or complete reduction or decrease in the severity, duration or frequency of adverse symptoms or complications associated with or resulting from a cell proliferative disorder, and improvement in the quality of life and / or health of a subject such as increased energy, appetite, mental health are all specific non-limiting examples of a therapeutic effect.

[0278] Accordingly, a therapeutic effect or improvement may also include a subjective improvement in the quality of life of the subject being treated. In additional embodiments, the method extends or prolongs the lifespan (survival) of the subject. In a further embodiment, the method improves the quality of life of the subject.

[0279] In one embodiment, the administration of an immunotherapeutic agent in combination therapy with a compound of formula I’ results in a clinically relevant improvement in one or more markers of disease state and progression selected from one or more of (i) overall survival, (ii) progression-free survival, (iii) overall response rate, (iv) reduction of metastatic disease, (v) circulating levels of tumor antigens such as carbohydrate antigen 19.9 (CA19.9) and carcinoembryonic antigen (CEA) or others according to the tumor, (vii) nutritional status (weight, appetite, serum albumin), (viii) pain management or use of analgesics, (ix) CRP / albumin ratio.

[0280] Treatment with a compound of formula I’ in combination with an immunotherapeutic agent results in a more complex immunity that includes not only the development of innate and type 1 immunity but also immunomodulation that more efficiently restores appropriate immune function.

[0281] In various exemplary ways, checkpoint inhibitory antibodies (monoclonal or polyclonal, bispecific, trispecific, or immunocyte-engaging multivalent antibodies / fusion proteins / constructs) directed to a checkpoint molecule of interest (e.g., PD-1) can be sequenced, and then the polynucleotide sequence can be cloned into a vector for expression or propagation. The sequence encoding the antibody of interest or its antigen-binding fragment can be maintained in a vector in a host cell, and then the host cell can be expanded and frozen for future use. The production of recombinant monoclonal antibodies in cell culture can be carried out by cloning antibody genes from B cells by means known in the art. See, for example, Tiller et al., 2008, J. Immunol. Methods 329, 112, U.S. Patent No. 7,314,622.

[0282] In some embodiments, a method of generating a recombinant antibody can include culturing a host cell containing an isolated nucleic acid(s) encoding an antibody of the present disclosure. Methods for culturing a host cell containing an isolated nucleic acid(s) encoding an antibody of the present disclosure can be performed in a variety of ways depending on the nature of the antibody. In some embodiments, when the antibody of the present disclosure is a conventional full-length antibody, for example, the heavy chain variable region and the light chain variable region can be isolated under conditions such that the antibody is produced.

[0283] Generally, a nucleic acid encoding an antibody or an antigen-binding fragment thereof of the present disclosure is obtained. Such polynucleotides encode both the variable and constant regions of the heavy and light chains, although other combinations are contemplated in the present disclosure. The present disclosure also contemplates the disclosed polynucleotides and oligonucleotide fragments derived from nucleic acid sequences complementary to these polynucleotides.

[0284] The polynucleotide can be in the form of RNA, DNA, cDNA, genomic DNA, nucleic acid analogs, and synthetic DNA. The DNA can be double-stranded or single-stranded, and if single-stranded, can be the coding (sense) strand or the non-coding (antisense) strand. The coding sequence encoding the polypeptide can be identical to the coding sequence or can be a different coding sequence encoding the same polypeptide as a result of redundancy or degeneracy of the genetic code.

[0285] In some embodiments, the nucleic acid(s) encoding the antibody of the present disclosure is incorporated into an expression vector, which may be episomal or may be designed to integrate into the genome of the introduced host cell. The expression vector may contain any number of suitable control sequences (including, but not limited to, transcriptional and translational control sequences, promoters, ribosome binding sites, enhancers, origins of replication, etc.) or other components (such as selectable genes), all of which are operably linked as is well known in the art. In some cases, two nucleic acids are used and each is placed into a different expression vector (e.g., the heavy chain in a first expression vector and the light chain in a second expression vector), or alternatively, they can be placed into the same expression vector. Those skilled in the art will appreciate that the design of the expression vector(s), including the selection of control sequences, can depend on factors such as the choice of host cell and the desired level of protein expression.

[0286] Generally, the nucleic acid molecule(s) is selected such that it is operably linked to one or more expression control elements (e.g., in a vector, in a construct made by a process in a cell, integrated into the host cell genome), and any method suitable for the selected host cell (e.g., transformation, transfection, electroporation, infection) is used to introduce the nucleic acid and / or expression into the appropriate host cell to generate a recombinant host cell. The resulting recombinant host cell can be maintained under conditions suitable for expression (e.g., in the presence of an inducer, in a suitable non-human animal, in a suitable culture medium supplemented with appropriate salts, growth factors, antibiotics, nutrient supplements, etc.), thereby producing the encoded polypeptide(s). In some cases, the heavy chain is produced in one cell and the light chain is produced in another cell.

[0287] Mammalian cell lines available as hosts for expression are known in the art and include, but are not limited to, Chinese hamster ovary (CHO) cells, HEK293 cells, NSO cells, HeLa cells, baby hamster kidney (BHK) cells, simian kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), and many immortalized cell lines available from the American Type Culture Collection (ATCC), Manassas, VA USA, including several other cell lines. Non-mammalian cells, including but not limited to bacteria, yeast, insects, and plants, can also be used to express recombinant antibodies. In some embodiments, antibodies can be generated in transgenic animals such as cows or chickens.

[0288] Exemplary and illustrative recombinant methods for antibody molecular biology, expression, purification, and screening are described, for example, in Antibody Engineering, edited by Kontermann & Dubel, Springer, Heidelberg, 2001 and 2010 Hayhurst & Georgiou, 2001, Curr. Opin. Ch em. Biol. 5: 683 - 689, Maynard & Georgiou, 2000, Annu. Rev. Biomed. Eng. 2: 339 - 76, and Morrison, S. (1985) Science 229: 1202, the entire disclosures of which are incorporated herein by reference.

[0289] In various embodiments, polynucleotide sequences encoding selected variable heavy and light chains can be used for genetic manipulation to humanize an antibody or to improve the affinity or other characteristics of the antibody. Antibodies can also be customized for use, for example, in dogs, cats, primates, horses, and cows.

[0290] In some embodiments, fully human antibodies can be obtained by using commercially available mice engineered to express specific human immunoglobulin proteins. Transgenic animals designed to yield more desirable (e.g., fully human antibodies) or stronger immune responses can also be used for humanization or the production of human antibodies. Examples of such technologies are Xenomouse™ from Abgenix, Inc. (Fremont, Calif.), as well as HuMAb-Mouse® and TC Mouse™ from Medarex, Inc. (Princeton, N.J.).

[0291] The immune checkpoint regulatory antibodies of the present disclosure can be recombinantly produced by first isolating antibodies and antibody-producing cells from a host animal, obtaining the gene sequences, and using those gene sequences to recombinantly express the antibodies in a host cell (e.g., CHO cells). Another method that can be used is to express the antibody sequences in plants (e.g., tobacco) or in yeast cells (e.g., Pichia pastoris or Sacchromyces cerevisiae). Methods for recombinantly expressing antibodies in plants or yeast are disclosed. See, for example, Peeters, et al. Vaccine 19:2756, 2001, Lonberg, N. and D. Huszar Int. Rev. Immunol 13:65, 1995, and Horwitz, A. H. et al., Proc. Natl. Acad. Sci. 85:8678-8682, the entire disclosures of which are incorporated herein by reference. Methods for making derivatives of antibodies, such as domains, single chains, etc., are known in the art.

[0292] Flow cytometry sorting techniques such as immunoassays and fluorescence-activated cell sorting (FACS) can also be used to isolate antibodies specific for checkpoint molecules.

[0293] In some embodiments, the polynucleotide comprises a sequence encoding the heavy and / or light chain variable regions of a checkpoint inhibitory antibody of the present disclosure or an antigen-binding fragment thereof. The sequence encoding the desired antibody or an antigen-binding fragment thereof can be maintained in a host cell with a vector and then the host cell can be expanded and frozen for future use. Vectors (including expression vectors) and host cells are further described herein.

[0294] The present disclosure includes affinity matured checkpoint regulatory antibodies. For example, affinity matured antibodies can be generated by procedures known in the art (Marks et al., 1992, Bio / Technology, 10:779-783, Barbas et al., 1994, Proc Nat. Acad. Sci. USA 91:3809-3813). One method of characterizing the CDRs of an antibody and / or altering (e.g., improving) the binding affinity of a polypeptide such as an antibody is termed "library scanning mutagenesis." Exemplary methods for obtaining affinity matured antibodies and antigen-binding fragments are to use methods recognized in the art to mutate one or more amino acid positions in the CDR It may involve replacement with two or more (such as 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) amino acids. A clone library is generated, each having the complexity of two or more members (when two or more amino acids are replaced at each position). Generally, the library also includes clones containing natural (unsubstituted) amino acids. A small number of clones from each library, for example, about 20 to 80 clones (depending on the complexity of the library), are screened for binding affinity to the target polypeptide (or other binding target), and candidates with increased, the same, decreased, or no binding are identified. Methods for determining binding affinity are well known in the art. Binding affinity can be determined, for example, using Biacore™ surface plasmon resonance analysis, Kinexa® biosensor, scintillation proximity assay, ELISA, ORIGEN® immunoassay, fluorescence quenching, fluorescence shift, and / or yeast display, which can detect a difference in binding affinity of about twofold or more. Binding affinity can also be screened using an appropriate bioassay. Biacore™ is particularly useful when the starting antibody already binds with a relatively high affinity, for example, with a KD of about 10 nM or less. Then, the clone library can be recombinantly introduced into the selection construct using any method known in the art for selection, including phage display, yeast display, and ribosome display.

[0295] For example, in order to modify the binding properties of an antibody, the antibody can be modified, for example, in the variable domains of the heavy and / or light chains. Changes in the variable region may alter the binding affinity and / or specificity. In some embodiments, 1 to 5 conservative amino acid substitutions are made within the CDR domain. In other embodiments, 1 to 3 conservative amino acid substitutions are made within the CDR domain. For example, mutations can be made in one or more of the CDR regions to increase or decrease the KD of an antibody directed to a checkpoint molecule, to increase or decrease kon, or to alter the binding specificity of the antibody. Techniques for site-directed mutagenesis are well known in the art. See, for example, Sambrook et al. and Ausubel et al.

[0296] The pharmaceutical composition containing the compound of formula I' according to the present disclosure typically comprises an effective amount of the compound of formula I', an immunotherapeutic agent, and / or both, dispersed in a pharmaceutically acceptable carrier. The phrase "pharmaceutically or pharmacologically acceptable" refers to molecular entities and compositions that do not produce adverse, allergic, or other untoward reactions when administered to animals, such as humans, as appropriate. The preparation of pharmaceutical compositions containing the compound of formula I' will be apparent to those skilled in the art in view of the present disclosure, as exemplified by Remington’s Pharmaceutical Sciences, 18th Ed. Mack Printing Company, 1990. Further, it will be understood that in the administration to animals (e.g., humans), the formulation should meet sterility, pyrogenicity, general safety, and purity standards. Specific examples of pharmaceutically acceptable carriers for combination compositions containing the compound of formula I' mixed with an immunotherapeutic agent as described herein are boric acid buffer or sterile physiological saline (0.9% NaCl).

[0297] Formulations of immunotherapeutic agents, such as immune checkpoint regulatory antibodies, used in accordance with the present disclosure can be prepared in the form of lyophilized formulations or aqueous solutions and / or suspensions for storage by mixing an antibody having a desired purity with any optional pharmaceutically acceptable carrier, additive, or stabilizer as detailed and exemplified in Remington’s Pharmaceutical Sciences 16th edition, Osol, A. Ed.

[1980] . Acceptable carriers, additives, buffers, or stabilizers are non-toxic to the recipient at the dosages and concentrations employed, and they include suitable aqueous and / or non-aqueous additives that can be used in the pharmaceutical compositions of the present disclosure, such as water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, etc.), And suitable mixtures thereof, vegetable oils such as olive oil, and organic esters for injection such as ethyl oleate are included. Appropriate fluidity can be maintained, for example, by the use of coating materials such as lecithin, in the case of dispersants by maintaining the required particle size, and by the use of buffers such as surfactants, phosphoric acid, citric acid, and other organic acids. Antioxidants include, for example, (1) water-soluble antioxidants such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite; (2) oil-soluble antioxidants such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol; and (3) metal chelating agents such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, preservatives (octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol, etc.), and low molecular weight (less than about 10 residues) may be included. Other exemplary pharmaceutically acceptable additives may include polypeptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrin; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants such as TWEEN (registered trademark), PLURONICS (registered trademark) or polyethylene glycol (PEG).

[0298] In one exemplary embodiment, the pharmaceutical composition may optionally contain pharmaceutically acceptable auxiliary substances such as pH regulators and buffers and toxicity regulators, such as sodium acetate, sodium chloride, potassium chloride, calcium chloride, and sodium lactate, if necessary to approximate physiological conditions. In some embodiments, the checkpoint inhibitory antibody or antigen-binding fragment thereof of the present disclosure can be formulated, lyophilized for storage, and reconstituted in a suitable additive prior to use according to lyophilization and reconstitution techniques known in the art. In one exemplary pharmaceutical composition containing one or more checkpoint inhibitory antibodies or antigen-binding fragments thereof, the composition is formulated as a preservative-free sterile solution of one or more checkpoint inhibitory antibodies or antigen-binding fragments thereof for intravenous or subcutaneous administration. The formulation can be supplied as a prefilled pen for single use, such as a prefilled glass syringe containing about 1 mL for single use, or as a vial for in-facility single use. Preferably, the pharmaceutical composition containing the checkpoint inhibitory antibody or antigen-binding fragment thereof is clear and colorless and has a pH in the range of about 6.9 to 5.0, preferably 6.5 to 5.0, and even more preferably about 6.0 to about 5.0. In various embodiments, when the formulation containing the pharmaceutical composition is reconstituted and administered to a subject, it can contain from about 500 mg to about 10 mg, or from about 400 mg to about 20 mg, or from about 300 mg to about 30 mg, or from about 200 mg to about 50 mg of the checkpoint inhibitory antibody or antigen-binding fragment thereof per mL of solution. Exemplary injectable or infusion additives can include mannitol, citric acid monohydrate, sodium hydrogen phosphate dihydrate, sodium dihydrogen phosphate dihydrate, polysorbate 80, sodium chloride, sodium citrate, and water for parenteral administration, such as intravenous, intramuscular, intraperitoneal, or subcutaneous administration.

[0299] In another exemplary embodiment, one or more immunotherapeutic agents, or antigen-binding fragments thereof, are formulated for intravenous or subcutaneous administration as a sterile aqueous solution containing the antibody at 1 to 75 mg / mL, or more preferably, about 5 to 60 mg / mL, or even more preferably, about 10 to 50 mg / mL, or even more preferably, about 10 to 40 mg / mL, together with sodium acetate, polysorbate 80, and sodium chloride at a pH in the range of about 5 to 6. Preferably, the intravenous or subcutaneous formulation is a sterile aqueous solution containing 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 mg / mL of an immunotherapeutic agent, such as an immune checkpoint inhibitory antibody or an antigen-binding fragment thereof, at pH 5.5 together with 20 mM sodium acetate, 0.2 mg / mL polysorbate 80, and 140 mM sodium chloride. Further, the solution containing the checkpoint inhibitory antibody or an antigen-binding fragment thereof can contain, among many other compounds, histidine, mannitol, sucrose, trehalose, glycine, poly(ethylene) glycol, EDTA, methionine, and any combination thereof, as well as many other compounds known in the relevant art.

[0300] In one embodiment, the pharmaceutical composition of the present disclosure contains the following components: 5 to 500 mg of the immunotherapeutic agent or an antigen-binding fragment thereof of the present disclosure, 10 mM histidine, 5% sucrose, and 0.01% polysorbate 80 at pH 5.8, with or without the compound of formula I'. This composition can also be provided as a lyophilized powder. When the powder is reconstituted to its total volume, the composition maintains the same formulation. Alternatively, the powder can be reconstituted to half its volume, in which case the composition contains 10 to 500 mg of the immunotherapeutic agent or an antigen-binding fragment thereof of the present disclosure, 20 mM histidine, 10% sucrose, and 0.02% polysorbate 80 at pH 5.8.

[0301] In one embodiment, a portion of the dose of the immunotherapeutic agent formulation is administered by intravenous bolus and the remainder by infusion. For example, an intravenous injection of an immunotherapeutic agent, or an antigen-binding fragment thereof, in an amount of about 0.001 to about 200 mg / kg, such as about 0.001 mg / kg to about 100 mg / kg, or about 0.001 mg / kg to about 50 mg / kg, or about 0.001 mg / kg to about 10 mg / kg can be given as a bolus, and the remainder of the antibody dose can be administered by intravenous infusion. A predetermined dose of the immunotherapeutic agent, or an antigen-binding fragment thereof, may be administered over a period of, for example, 1 to 2 hours, 5 hours.

[0302] In a further embodiment, a portion of the dose of the immunotherapeutic agent formulation is administered subcutaneously and / or by infusion in the form of a bolus, and the remainder by infusion. In some exemplary doses, the immunotherapeutic agent formulation can be administered subcutaneously at a dose in the range of about 0.001 to about 200 mg / kg, such as about 0.001 mg / kg to about 100 mg / kg, or about 0.001 mg / kg to about 50 mg / kg, or an intravenous injection of an immunotherapeutic agent, or an antigen-binding fragment thereof, in an amount of about 0.001 mg / kg to about 10 mg / kg. In some embodiments, the dose can be given as a bolus and the remainder of the immunotherapeutic agent dose can be administered subcutaneously or by intravenous infusion. A predetermined dose of the immunotherapeutic agent, or an antigen-binding fragment thereof, may be administered over a period of, for example, 1 to 2 hours, 5 hours.

[0303] The formulations herein may contain more than one active compound, preferably compounds having complementary activities that do not adversely affect each other, if necessary to treat a particular indication. For example, it may be desirable to provide one or more immunotherapeutic agents having other specificities. Alternatively or in addition, the composition may contain an anti-inflammatory agent, a chemotherapeutic agent, a cytotoxic agent, a cytokine, a growth inhibitor and / or a small molecule antagonist. Such molecules are present in a suitable combination in an amount effective for the intended purpose.

[0304] The formulations used for in vivo administration should be sterile or nearly sterile. This can be readily achieved by filtration through a sterile filtration membrane.

[0305] In various embodiments, exemplary formulations of the pharmaceutical compositions described herein can be prepared using methods well known in the art of pharmaceutical formulations. Generally, such preparation methods can include the step of mixing the active ingredient with a carrier or one or more other accessory ingredients, and then, optionally, packaging the product into the desired single or multiple dose units.

[0306] In some embodiments, the compositions containing the compounds of formula I' can also be delivered in vesicles, and immunotherapeutic agents can be delivered in the same liposomal formulation or in another formulation compatible with the liposomal formulation containing the compounds of formula I'. In some examples, liposomes containing one or more liposomal surface moieties, such as polyethylene glycol, and antibodies and antibody fragments targeting the desired tumor surface antigen, receptor, growth factor, glycoprotein, glycolipid or neoantigen, and these surface moieties, antibodies and antibody fragments are selectively transported to specific cells or organs, thus enhancing targeted drug delivery.

[0307] In another embodiment, the compounds of formula I' can be delivered in vesicles, particularly liposomes (see Langer, Science 249:1527-1533 (1990), Treat et al., LIPOSOMES IN THE THERAPY OF INFECTIOUS DISEASE AND CANCER, Lopez-Berestein and Fidler (eds.), Liss, N.Y., pp. 353-365 (1989), Lopez-Berestein, ibid., pp. 317-327, generally see ibid.).

[0308] In yet another embodiment, a composition containing a compound of formula I', or a combination, or a composition containing an immunotherapeutic agent can be delivered by a controlled release system. In one embodiment, a pump can be used (see Langer, supra, Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987), Buchwald et al., Surgery 88:507 (1980), Saudek et al., N. Engl. J. Med. 321:574 (1989)). In another embodiment, the controlled release of the compound of formula I' can include a polymeric material for providing sustained, intermediate, pulsatile, or alternating release (see MEDICAL APPLICATIONS OF CONTROLLED RELEASE, Langer and Wise (eds.), CRC Pres., Boca Raton, Fla. (1974), CONTROLLED DRUG BIOAVAILABILITY, DRUG PRODUCT DESIGN AND PERFORMANCE, Smolen and Ball (eds.), Wiley, New York (1984), Ranger and Peppas, J. Macromol. Sci. Rev. Macromol. Chem. 23:61 (1983). See also Levy et al., Science 228:190 (1985), During et al., Ann. Neurol. 25:351 (1989), Howard et al., J. Neurosurg. 71:105 (1989)). Other controlled release systems discussed in the review by Langer (Science 249:1527-1533 (1990)) can also be used.

[0309] The optimal concentration of the active ingredient(s) in a selected medium can be determined empirically according to procedures well known to those skilled in the art and should depend on the desired final pharmaceutical formulation and the use employed.

[0310] The present disclosure also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the components of the pharmaceutical compositions of the present disclosure, including at least a compound of formula I' as described herein and one or more checkpoint inhibitory antibodies or antigen-binding fragments thereof. In other embodiments, the kit may include one or more additional containers providing pharmaceutically acceptable additives, such as diluents. In one embodiment, the kit may include at least one container that can contain a compound of formula I' of the present disclosure, a checkpoint inhibitory antibody or an antigen-binding fragment thereof. The kit may include a series of instructions for preparing the final pharmaceutical composition for treating a checkpoint molecule-mediated disease or disorder and administering it to a subject in need thereof. It may include a series of instructions for preparing the final pharmaceutical composition for treating a checkpoint molecule-mediated disease or disorder and administering it to a subject in need thereof.

[0311] In some embodiments of the present disclosure, the immunotherapeutic agent is a population of immune cells that can be administered in combination with a compound of formula I' to treat a subject having cancer. In some embodiments, the immunotherapeutic agent is a population of immune cells, such as leukocytes (nucleated white blood cells) that contain (e.g., express) receptors that bind to an antigen of interest. The leukocytes of the present disclosure can be, for example, neutrophils, eosinophils, basophils, lymphocytes or monocytes. In some embodiments, the leukocytes are lymphocytes. Examples of lymphocytes include T cells, B cells, natural killer (NK) cells or NKT cells. In some embodiments, the T cells are CD4+ Th (T helper) cells, CD8+ cytotoxic T cells, γδ T cells or regulatory (suppressor) T cells. In some embodiments, the immune cells are dendritic cells.

[0312] In some embodiments, the immune cells of the present disclosure are genetically engineered to express an antigen-binding receptor. A cell is considered "engineered" when it contains the engineered (exogenous) nucleic acid. The engineered nucleic acid of the present disclosure can be introduced into the cell by any known (e.g., conventional) method. For example, the engineered nucleic acid can be introduced by electroporation (e.g., Heiser W.C. Transcription Factor Protocols: Methods (see, e.g., in Molecular Biology.TM.2000;130:117-134), chemicals (e.g., calcium phosphate or lipids), transfection (see, e.g., Lewis W.H., et al., Somatic Cell Genet. 1980 May;6(3):333-47, Chen C., et al., Mol Cell Biol. 1987 August;7(8):2745-2752), fusion with bacterial protoplasts containing recombinant plasmids (see, e.g., Schaffner W. Proc Natl Acad Sci USA. 1980 April;77(4):2163-7), direct microinjection of purified DNA into the nucleus of the cell (see, e.g., Capecchi M.R. Cell. 1980 November;22(2 Pt 2):479-88), or transduction with retroviruses.

[0313] Some aspects of the present disclosure provide an "adoptive cell" approach, which involves isolating immune cells (e.g., T cells) from a subject having cancer, genetically engineering the immune cells (e.g., to express an antigen-binding receptor such as a chimeric antigen receptor), expanding the cells ex vivo, and then reintroducing the immune cells into the subject. This method results in a greater number of engineered immune cells in the subject compared to the number that could be achieved by conventional gene delivery and vaccination methods. In some embodiments, immune cells are isolated from the subject, expanded ex vivo without genetic modification, and then reintroduced into the subject.

[0314] The immune cells of the present disclosure include receptors that bind to antigens, such as antigens encoded by exogenously delivered nucleic acids, as provided herein. In some embodiments, leukocytes are modified (e.g., genetically modified) to express a receptor that binds to an antigen. The receptor may, in some embodiments, be a naturally occurring antigen receptor (normally expressed on immune cells), a recombinant antigen receptor (normally not expressed on immune cells), or a chimeric antigen receptor (CAR). Naturally occurring antigen receptors and recombinant antigen receptors encompassed by the present disclosure include T cell receptors, B cell receptors, NK cell receptors, NKT cell receptors, and dendritic cell receptors. A "chimeric antigen receptor" refers to an artificial immune cell receptor that has been engineered to recognize and bind to an antigen expressed by a tumor cell. Generally, a CAR is a chimera of a signaling domain of a T cell receptor (TcR) complex and an antigen recognition domain (e.g., a single-chain fragment (scFv) of an antibody) (Enblad et al., Human Gene Therapy. 2015; 26(8): 498 - 505), the entire disclosure of which is incorporated herein by reference.

[0315] In some embodiments, the antigen-binding receptor is a chimeric antigen receptor (CAR). T cells that express a CAR are referred to as "CAR T cells." A "CAR T cell receptor" includes, in some embodiments, a signaling domain of a T cell receptor (TcR) complex and an antigen recognition domain (e.g., a single-chain fragment (scFv) of an antibody) (Enblad et al., Human Gene Therapy. 2015; 26(8): 498 - 505), the entire disclosure of which is incorporated herein by reference.

[0316] There are four generations of CARs, each containing different components. First-generation CARs link an antibody-derived scFv to the CD3 zeta (zeta or z) intracellular signaling domain of the T cell receptor via a hinge and transmembrane domain. Second-generation CARs incorporate additional domains, such as CD28, 4-1BB (41BB), or ICOS, to provide co-stimulatory signals. Third-generation CARs contain two co-stimulatory domains fused to the TcR CD3-zeta chain. Third-generation co-stimulatory domains can include, for example, combinations of CD3z, CD27, CD28, 4-1BB, ICOS, or OX40. In some embodiments, CARs generally contain an ectodomain (e.g., CD3) derived commonly from a single-chain variable fragment (scFv), a hinge, a transmembrane domain, and an endodomain having one (first generation), two (second generation), or three (third generation) signaling domains derived from CD3Z and / or co-stimulatory molecules (Maude et al., Blood. 2015;125(26):4017-4023, Kakarla and Gottschalk, Cancer J. 2014;20(2):151-155, the entire disclosures of which are incorporated herein by reference).

[0317] In some embodiments, the chimeric antigen receptor (CAR) is a T-cell redirected for universal cytokine killing (TRUCK), also known as a fourth-generation CAR. TRUCKs are CAR redirected T cells used as vehicles to produce and secrete a transgenic cytokine that accumulates in a target tissue, e.g., a target tumor tissue. The transgenic cytokine is released when the CAR engages the target. TRUCK cells can deposit various therapeutic cytokines within the target. This results in concentration of the therapeutic agent at the target site and can avoid systemic toxicity.

[0318] CARs typically differ in their functional properties. The CD3 zeta signaling domain of the T cell receptor, when engaged, activates T cells and induces their proliferation, but can also result in anergy (lack of response by the body's defense mechanisms that leads to the direct induction of peripheral lymphocyte tolerance). Lymphocytes are considered anergic when they are unable to respond to specific antigens. The addition of costimulatory domains in second-generation CARs improved the replicative ability and survival of the modified T cells. In vitro, similar antitumor effects have been observed for CD28 CAR or 4-1BB CAR, but preclinical in vivo studies suggest that 4-1BB CAR can result in superior proliferation and / or survival. Clinical trials suggest that both of these second-generation CARs can induce substantial T cell proliferation in vivo, but CARs containing the 4-1BB costimulatory domain appear to survive longer. Third-generation CARs combine multiple signaling domains (costimulatory) to enhance efficacy. Fourth-generation CARs are additionally modified with a constitutive or inducible expression cassette for a transgenic cytokine released by the CAR T cells to regulate the T cell response. See, for example, Enblad et al., Human Gene Therapy. 2015;26(8):498-505, Chmielewski and Hi nrich, Expert Opinion on Biological Therapy. 2015;15(8):1145-1154, which are hereby incorporated by reference in their entireties.

[0319] In some embodiments, the exemplary immunotherapeutic agent is a first-generation chimeric antigen receptor (CAR). In some embodiments, the chimeric antigen receptor is a third-generation CAR. In some embodiments, the chimeric antigen receptor is a second-generation CAR. In some embodiments, the chimeric antigen receptor is a third-generation CAR. In some embodiments, the chimeric antigen receptor is a fourth-generation CAR or a T cell redirected for universal cytokine-mediated killing (TRUCK).

[0320] In some embodiments, the chimeric antigen receptor (CAR) comprises an extracellular domain comprising an antigen-binding domain, a transmembrane domain, and a cytoplasmic domain. In some embodiments, the CAR is fully human. In some embodiments, the antigen-binding domain of the CAR is specific for one or more antigens. In some embodiments, a "spacer" domain or "hinge" domain is disposed between the extracellular domain (including the antigen-binding domain) of the CAR and the transmembrane domain, or between the cytoplasmic domain of the CAR and the transmembrane domain. A "spacer domain" refers to any oligopeptide or polypeptide that functions to link the transmembrane domain to the extracellular domain and / or cytoplasmic domain within the polypeptide chain. A "hinge domain" refers to any oligopeptide or polypeptide that functions to confer flexibility to the CAR or its domain, or to prevent steric hindrance of the CAR or its domain. In some embodiments, the spacer domain or hinge domain can comprise up to 300 amino acids (e.g., 10 to 100 amino acids, or 5 to 20 amino acids). In some embodiments, one or more spacer domains can also be included within other regions of the CAR.

[0321] In some embodiments, the CARs of the present disclosure comprise an antigen-binding domain such as a single-chain Fv (scFv) specific for a tumor antigen. The choice of binding domain depends on the type and number of ligands that define the surface of the target cell. For example, the antigen-binding domain can be selected to recognize a ligand that acts as a cell surface marker on the target cell associated with a particular disease state such as cancer or an autoimmune disease. Thus, examples of cell surface markers that can act as ligands for the antigen-binding domain within the CARs of the present disclosure include cell surface markers associated with cancer cells and / or other forms of diseased cells. In some embodiments, as provided herein, the CARs are engineered to target a desired tumor antigen by engineering the desired antigen-binding domain that specifically binds to an antigen on the tumor cell, encoded by the engineered nucleic acid.

[0322] An antigen-binding domain (e.g., scFv) that "specifically binds" to a target or epitope is a term understood in the art, and methods for determining such specific binding are also known in the art. A molecule "specifically binds" when it reacts or associates more frequently, rapidly, for a longer duration, and / or with greater affinity with a particular target antigen than with alternative targets. An antigen-binding domain (e.g., scFv) that specifically binds to a first target antigen may or may not specifically bind to a second target antigen. Thus, "specific binding" does not necessarily require exclusive binding (although it may include this).

[0323] In some embodiments, immune cells expressing a CAR are genetically modified to recognize multiple targets or antigens, thereby enabling recognition of unique target or antigen expression patterns on tumor cells. Examples of CARs capable of binding multiple targets include "split-signal CARs" that limit full immune cell activation to tumors expressing multiple antigens, "tandem CARs" (TanCARs) containing an ectodomain with two scFvs, and "universal external domain CARs" that incorporate avidin or fluorescein isothiocyanate (FITC)-specific scFvs to recognize tumor cells incubated with a tagged monoclonal antibody (Mab). Included are those that incorporate avidin or fluorescein isothiocyanate (FITC)-specific scFvs to recognize tumor cells incubated with a tagged monoclonal antibody (Mab).

[0324] A CAR is considered "bispecific" when it recognizes two distinct antigens (has two distinct antigen recognition domains). In some embodiments, the bispecific CAR is composed of two distinct antigen recognition domains that are tandemly present on a single transgenic receptor (referred to as TanCAR. See, for example, Grada Z et al. Molecular Therapy Nucleic Acids 2013;2:e105, which is hereby incorporated by reference in its entirety). Thus, in some embodiments, the method comprises delivering to a tumor a combination comprising a compound of formula I' and an immunotherapeutic agent that is an engineered nucleic acid encoding an antigen, or delivering to a tumor an engineered nucleic acid that induces the expression of a self-antigen, and delivering to the tumor immune cells that express a bispecific CAR that binds to two antigens, one of which is encoded by the engineered nucleic acid.

[0325] In some embodiments, the CAR is, for example, an antigen-specific inhibitory CAR (iCAR) that can be used to avoid toxicity outside the tumor (Fedorov, V D et al. Sci. Transl. Med., published online on December 11, 2013, which is hereby incorporated by reference in its entirety). The iCAR contains, for example, an antigen-specific inhibitory receptor to block non-specific immunosuppression that can arise from off-tumor target expression. The iCAR can be based, for example, on the inhibitory molecules CTLA-4 or PD-1. In some embodiments, these iCARs block the T cell response from T cells activated by either their endogenous T cell receptor or an activating CAR. In some embodiments, this inhibitory effect is transient.

[0326] In some embodiments, the CAR can be used in adoptive cell transfer, where immune cells are removed from a subject and modified to express receptors specific for an antigen, such as a tumor-specific antigen. The modified immune cells, which can then recognize and kill cancer cells, are then reintroduced into the subject (Pule, et al., Cytotherapy. 2003;5(3):211-226, Maude et al., Blood. 2015;125(26):4017-4023, each of which is hereby incorporated by reference in its entirety).

[0327] In other aspects of the disclosure, the tumor antigen component in the vaccine of the invention can be any natural or synthetic tumor-related protein or peptide or combination of tumor-related proteins and / or peptides or glycoproteins or glycopeptides. In yet other aspects, the antigen component can be patient-specific or common to many or most patients having a particular type of cancer. In one aspect, the antigen component consists of a cell lysate derived from tumor tissue removed from the patient to be treated. In another aspect, the lysate can be engineered or synthesized from exosomes derived from tumor tissue. In yet another aspect, the antigen component consists of a cell lysate derived from tumor tissue extracted from one or more unrelated individuals or from a tumor cell line.

[0328] In various embodiments, exemplary immunotherapeutic agents include one or more cancer vaccines for use in combination with a compound of Formula I’. The tumor-associated antigen components of the vaccine may be made by any of a variety of well-known techniques. For individual protein components, the antigenic protein may be isolated from tumor tissue or tumor cell lines by standard chromatographic means such as high performance liquid chromatography or affinity chromatography, or alternatively, synthesized in a suitable expression system such as E. coli, yeast or plants by standard recombinant DNA techniques. The tumor-associated antigen protein is then purified from the expression system by standard chromatographic means. In the case of peptide antigen components, these are generally prepared by standard automated synthesis. Proteins and peptides can be modified by the addition of amino acids, lipids and other agents to improve incorporation into the vaccine delivery system (such as multilamellar liposomes). For tumor-associated antigen components derived from the patient's own tumor, or tumors from other individuals, or cell lines, the tumor tissue, or a single cell suspension derived from the tumor tissue, is typically homogenized in a suitable buffer. The homogenate can also be fractionated by centrifugation or the like to isolate specific cell components such as cell membranes or soluble substances. The tumor material can be used as is, or alternatively, extracted using a buffer containing a suitable agent such as a low concentration of surfactant to incorporate the tumor-associated antigen into the vaccine. An example of a suitable surfactant for extracting antigenic proteins from tumor tissue, tumor cells, and tumor cell membranes is diheptanoyl phosphatidylcholine. Exosomes derived from tumor tissue or tumor cells can be used as starting materials for antigen components for incorporation into the vaccine or for extracting tumor-associated antigens, whether autologous or heterologous to the patient.

[0329] In some embodiments of the present disclosure, the cancer vaccine comprises at least one tumor-associated antigen, at least one immunostimulatory agent, and optionally, at least one cell-based immunotherapeutic agent. In some embodiments, the immunostimulatory agent component in the cancer vaccine of the present disclosure is any biological response modifier (BRM) having the ability to enhance the effectiveness of the therapeutic cancer vaccine and induce humoral and cellular immune responses against cancer cells in a patient. According to one aspect, the immunostimulatory agent is a cytokine or a combination of cytokines. Examples of such cytokines include interferons such as IFN-gamma, interleukins such as IL-2, IL-15 and IL-23, colony-stimulating factors such as M-CSF and GM-CSF, and tumor necrosis factor. According to another aspect, the immunostimulatory agent component of the disclosed cancer vaccine includes one or more adjuvant-type immunostimulatory agents, with or without immunostimulatory cytokines, such as APC Toll-like receptor agonists or co-stimulatory / cell adhesion membrane proteins. Examples of Toll-like receptor agonists include lipid A and CpG, and co-stimulatory / adhesion proteins such as CD80, CD86, and ICAM-1.

[0330] In some embodiments, the immunostimulant is selected from the group consisting of IFN-gamma (IFN-γ), IL-2, IL-15, IL-23, M-CSF, GM-CSF, tumor necrosis factor, lipid A, CpG, CD80, CD86, and ICAM-1, or combinations thereof. According to other aspects, the cell-based immunotherapeutic agent is selected from the group consisting of dendritic cells, tumor-infiltrating T lymphocytes, chimeric antigen receptor-modified T effector cells directed to the patient's tumor type, B lymphocytes, natural killer cells, myeloid cells, and any other cells of the patient's immune system, or combinations thereof. In one aspect, the cancer vaccine immunostimulant comprises one or more cytokines, such as interleukin 2 (IL-2), GM-CSF, M-CSF, and interferon-gamma (IFN-γ), one or more Toll-like receptor agonists and / or adjuvants, such as monophosphoryl lipid A, lipid A, muramyl dipeptide (MDP) lipid conjugate, and double-stranded RNA, or one or more costimulatory membrane proteins and / or cell adhesion proteins, such as CD80, CD86, and ICAM-1, or any combination of the above. In one aspect, the cancer vaccine comprises an immunostimulant that is a cytokine selected from the group consisting of interleukin 2 (IL-2), GM-CSF, M-CSF, and interferon-gamma (IFN-γ). In another aspect, the cancer vaccine comprises an immunostimulant that is a Toll-like receptor agonist and / or adjuvant selected from the group consisting of monophosphoryl lipid A, lipid A, and muramyl dipeptide (MDP) lipid conjugate and double-stranded RNA. In yet another aspect, the cancer vaccine comprises an immunostimulant that is a costimulatory membrane protein and / or cell adhesion protein selected from the group consisting of CD80, CD86, and ICAM-1.

[0331] In various embodiments, the immunotherapeutic agent can potentially be used to construct the fusion protein according to the present invention with any tumor antigen, particularly the following: (a) Testicular cancer antigens including, for example, NY-ESO-1, SSX2, SCP1, and further including RAGE, BAGE, GAGE, and MAGE family polypeptides such as GAGE-1, GAGE-2, MAGE-1, MAGE-2, MAGE-3, MAGE-4, MAGE-5, MAGE-6, and MAGE-12, which can be used to address, for example, melanoma, lung, head and neck, NSCLC, breast, gastrointestinal, and bladder tumors; (b) Mutant antigens including p53 associated with various solid tumors such as colorectal, lung, and head and neck cancers; p21 / Ras associated with, for example, melanoma, pancreatic cancer, and colorectal cancer; CDK4 associated with, for example, melanoma; MUM1 associated with, for example, melanoma; caspase-8 associated with, for example, head and neck cancer; CIA0205 associated with, for example, bladder cancer; HLA-A2-R1701, beta-catenin associated with, for example, melanoma; TCR associated with, for example, T-cell non-Hodgkin lymphoma; BCR-abl associated with, for example, chronic myelogenous leukemia; triosephosphate isomerase; KIA0205; CDC-27, and LDLR-FUT; (c) Overexpressed antigens including galectin 4 associated with, for example, colorectal cancer; galectin 9 associated with, for example, Hodgkin disease; proteinase 3 associated with, for example, chronic myelogenous leukemia; WT1 associated with, for example, various leukemias; carbonic anhydrase associated with, for example, renal cell carcinoma; aldolase A associated with, for example, lung cancer; PRAME associated with, for example, melanoma; HER-2 / neu associated with, for example, breast, colon, lung, and ovarian cancers; mammaglobin, alpha-fetoprotein associated with, for example, hepatocellular carcinoma; KSA associated with, for example, colorectal cancer; gastrin associated with, for example, pancreatic and gastric cancers; telomerase catalytic protein, MUC-1 associated with, for example, breast and ovarian cancers; G-250 associated with, for example, renal cell carcinoma; p53 associated with, for example, breast and colon cancers; and carcinoembryonic antigen associated with cancers of the digestive tract such as, for example, breast cancer, lung cancer, and colorectal cancer; (d) Melanoma-melanocyte differentiation antigens such as MART-1 / Melan A; gp100; MC1R; melanocyte-stimulating hormone receptor; tyrosinase;For example, common antigens including tyrosinase-related protein-1 / TRP1 and tyrosinase-related protein-2 / TRP2 associated with melanoma, (e) prostate-related antigens including PAP, PSA, PSMA, PSH-P1, PSM-P1, PSM-P2 associated with prostate cancer, for example, (f) cancer vaccines incorporating immunoglobulin idiotypes associated with multiple myeloma and B-cell lymphoma may be included. In certain embodiments, one or more TAAs are pi5, Hom / Mel-40, H-Ras, E2A-PRL, H4-RET, IGH-IGK, MYL-RAR, Epstein-Barr virus antigen, EBNA, human papillomavirus (HPV) antigens including E6 and E7, hepatitis B and C virus antigens, human T-cell lymphotropic virus antigen, TSP-180, p185erbB2, p180erbB-3, c-met, mn-23H1, TAG-72-4, CA19-9, CA72-4, CAM17.1, NuMa, K-ras, pi6, TAGE, PSCA, CT7, 43-9F, 5T4, 791Tgp72, beta-HCG, BCA225, BTAA, CA125, CA15-3 (CA27.29\BCAA), CA195, CA242, CA-50, CAM43, CD68\KP1, CO-029, FGF-5, Ga733 (EpCAM), HTgp-175, M344, MA-50, MG7-Ag, MOV18, NB / 70K, NY-CO-1, RCAS1, SDCCAG16, TA-90 (Mac-2 binding protein / cyclophilin C-related protein), TAAL6, TAG72, TLP, TPS or any combination thereof.;

[0332] In some embodiments, the cancer vaccine of the present disclosure for use in combination with a compound of Formula I' is the following human proteins: TCTN1 (Gene ID: ENSG00000204852), TCTN2 (Gene ID: ENSG00000168778), TCTN3 (Gene ID: ENSG00000119977), HIGD2A (Gene ID: ENSG00000146066), HIGD2B (Gene ID: ENSG00000175202), C4ORF32 (Gene ID: ENSG00000174749), FAM62A(E -SYT1, gene ID: ENSG00000139641), COLEC11 (gene ID: ENSG00000118004), FSTL5 (gene ID: ENSG00000168843), FAM82A2 (gene ID: ENSG00000137824), SCARA5 (gene ID: ENSG00000168079), VSTM1 (gene ID: ENSG00000189068), RNF5 (gene ID: ENSG00000183574), UNQ6126 (gene ID: gi|169216088), DPY19L3 (gene ID: ENSG00000178904), SLC39A10 (gene ID: ENSG00000196950), GPR107 (gene ID: ENSG00000148358), COL20A1 (gene ID: ENSG00000101203), GLT25D2 (gene ID: ENSG00000198756), SYTL3 (gene ID: ENSG00000164674), DENND1B (gene ID: ENSG00000162701), C6orf98 (gene ID: EG:387079), FAM69B (gene ID: ENSG00000165716), EMID1 (gene ID: OTTHUMG00000030824), KLRG2 (gene ID: ENSG00000188883), ERMP1 (gene ID: ENSG00000099219), VMO1 (gene ID: ENSG00000182853), C9orf46 (gene ID: ENSG00000107020), F1137107 (gene ID: ENSG00000177990), YIPF2 (gene ID: ENSG00000130733), TRYX3 (PRSS58, ENSG00000258223.2), C14orf135 (gene ID: ENSG00000126773), ANGPTL7 (gene ID: ENSG00000171819), TPCN2 (gene ID: ENSG00000162341), C18orf19 (gene ID: ENSG00000177150), OLFML1 (gene ID: ENSG00000183801), LYPD4 (gene ID: ENSG00000101203), MEGF8 (gene ID: ENSG00000105429), F1142986 (gene ID: ENSG00000196460), SLC46A1 (gene ID: ENSG00000076351), FAM180A (gene ID: ENSG00000189320), CRISP-3 (gene ID: ENSG00000096006), or a tumor antigen comprising the entire amino acid sequence, a part thereof, or a specific immunogenic epitope of any one of these combinations. These tumor antigens are disclosed in WO2010 / 086162, WO2010 / 086163, WO2011 / 051278, WO2011 / 051276, WO2011 / 051277, WO2011 / 051280, WO2011 / 051271, WO2011 / 135068, WO2014 / 198919, the entire contents of which are incorporated herein by reference.

[0333] In various embodiments, an exemplary immunotherapeutic agent may comprise an mRNA that is operable to encode any one or more of the above-described cancer antigens useful for synthesizing a cancer vaccine. In some exemplary embodiments, the mRNA-based cancer vaccine may have one or more of the following characteristics: a) the mRNAs encoding each cancer antigen are interspersed by cleavage-susceptible sites; b) the mRNAs encoding each cancer antigen are directly bound to each other without a linker; c) the mRNAs encoding each cancer antigen are bound to each other by a single nucleotide linker; d) each cancer antigen comprises 20 to 40 amino acids and contains a centrally located SNP mutation; e) at least 40% of the cancer antigens have a very high affinity for class I MHC molecules from the subject; f) at least 40% of the cancer antigens have a very high affinity for class II MHC molecules from the subject; g) at least 40% of the cancer antigens have a predicted binding affinity with IC>500 nM for HLA-A, HLA-B, and / or DRB1; h) the mRNA encodes 1 to 15 cancer antigens; i) 10 to 60% of the cancer antigens have a binding affinity for class I MHC and 10 to 60% of the cancer antigens have a binding affinity for class II MHC; and / or j) the mRNAs encoding the cancer antigens are arranged such that the cancer antigens are ordered to minimize mimotopes.

[0334] In various embodiments, a combination comprising a compound of Formula I’ as disclosed herein and a cancer vaccine immunotherapeutic can be used to induce an immune response against a cancer antigen in a subject. This method involves administering to the subject, in combination with administration of the compound of Formula I’ either in the same composition or in a separate composition administered simultaneously or sequentially, an RNA vaccine comprising at least one RNA polynucleotide having an open reading frame encoding at least one antigen polypeptide or an immunogenic fragment thereof, thereby inducing in the subject an immune response specific to the antigen polypeptide or an immunogenic fragment thereof, wherein the anti-antigen polypeptide antibody titer in the subject increases after vaccination as compared to the anti-antigen polypeptide antibody titer in a subject vaccinated with a prophylactically effective dose of a conventional vaccine against cancer. An “anti-antigen polypeptide antibody” is a serum antibody that specifically binds to the antigen polypeptide.

[0335] A prophylactically effective dose is a therapeutically effective dose that prevents progression of cancer at a clinically acceptable level. In some embodiments, the therapeutically effective dose is the dose described in the package insert for the vaccine. As used herein, a conventional vaccine refers to a vaccine other than the mRNA vaccine of the present invention. For example, conventional vaccines include, but are not limited to, live microbial vaccines, killed microbial vaccines, subunit vaccines, protein antigen vaccines, DNA vaccines, and the like. In an exemplary embodiment, the conventional vaccine is a vaccine that has received regulatory approval and / or is registered by a national drug regulatory agency, such as the Food and Drug Administration (FDA) in the United States or the European Medicines Agency (EMA) in Europe.

[0336] In some embodiments, the anti-antigen polypeptide antibody titer in a subject increases by 1 log to 10 logs after vaccination compared to the anti-antigen polypeptide antibody titer in a subject vaccinated with a prophylactically effective dose of a conventional vaccine against cancer. In some embodiments, the anti-antigen polypeptide antibody titer in a subject increases by 1 log after vaccination compared to the anti-antigen polypeptide antibody titer in a subject vaccinated with a prophylactically effective dose of a conventional vaccine against cancer. In some embodiments, the anti-antigen polypeptide antibody titer in a subject increases by 2 logs after vaccination compared to the anti-antigen polypeptide antibody titer in a subject vaccinated with a prophylactically effective dose of a conventional vaccine against cancer.

[0337] Aspects of the invention provide a nucleic acid vaccine comprising one or more RNA polynucleotides having an open reading frame encoding a first antigen polypeptide, wherein the RNA polynucleotide is present in a formulation for in vivo administration to a recipient and provides an antibody titer that is superior to the criteria for determining antibody retention rate for the first antigen in an acceptable percentage of human subjects. In some embodiments, the antibody titer generated by the mRNA vaccine of the invention is a neutralizing antibody titer. In some embodiments, the neutralizing antibody titer is higher than that of a protein vaccine. In other embodiments, the neutralizing antibody titer generated by the mRNA vaccine of the invention is higher than that of an adjuvant-added protein vaccine. In still other embodiments, the neutralizing antibody titer generated by the mRNA vaccine of the invention is 1,000 to 10,000, 1,200 to 10,000, 1,400 to 10,000, 1,500 to 10,000, 1,000 to 5,000, 1,000 to 4,000, 1,800 to 10,000, 2000 to 10,000, 2,000 to 5,000, 2,000 to 3,000, 2,000 to 4,000, 3,000 to 5,000, 3,000 to 4,000, or 2,000 to 2,500. The neutralization titer is typically expressed as the maximum serum dilution required to achieve a 50% reduction in the number of plaques.

[0338] In a preferred embodiment, the RNA vaccine immunotherapeutic agent (e.g., mRNA vaccine) of the present disclosure results in antigen-specific antibodies at prophylactically and / or therapeutically effective levels, concentrations, and / or titers in the blood or serum of the vaccinated subject. As defined herein, the term antibody titer refers to the amount of antigen-specific antibodies produced in a subject, e.g., a human subject. In an exemplary embodiment, the antibody titer is expressed as the reciprocal of the highest dilution (in a serial dilution) that still shows a positive result. In an exemplary embodiment, the antibody titer is determined or measured by an enzyme-linked immunosorbent assay (ELISA). In an exemplary embodiment, the antibody titer is determined or measured by a neutralization assay, e.g., a micro-neutralization assay. In certain embodiments, the antibody titer measurement is expressed as a ratio such as 1:40, 1:100, etc.

[0339] In exemplary embodiments of the present invention, an effective vaccine results in an antibody titer of greater than 1:40, greater than 1:100, greater than 1:400, greater than 1:1000, greater than 1:2000, greater than 1:3000, greater than 1:4000, greater than 1:500, greater than 1:6000, greater than 1:7500, greater than 1:10000. In exemplary embodiments, the antibody titer occurs, or is achieved, by 10 days after vaccination, by 20 days after vaccination, by 30 days after vaccination, by 40 days after vaccination, or at 50 days or more after vaccination. In exemplary embodiments, the titer occurs, or is achieved, after a single dose of the vaccine has been administered to the subject. In other embodiments, the titer occurs, or is achieved, after multiple administrations, for example, after the first and second administrations (e.g., booster doses). In an exemplary aspect of the present invention, antigen-specific antibodies are measured in units of g / ml or in units of IU / L (international units per liter) or mIU / ml (milli-international units per ml). In exemplary embodiments of the present invention, an effective vaccine results in >0.5 μg / mL, >0.1 μg / mL, >0.2 μg / mL, >0.35 μg / mL, >0.5 μg / mL, >1 μg / mL, >2 μg / mL, >5 μg / mL or >10 μg / mL. In exemplary embodiments of the present invention, an effective vaccine results in >10 mIU / mL, >20 mIU / mL, >50 mIU / mL, >100 mIU / mL, >200 mIU / mL, >500 mIU / ml or >1000 mIU / ml. In exemplary embodiments, the antibody level or concentration occurs, or is achieved, by 10 days after vaccination, by 20 days after vaccination, by 30 days after vaccination, by 40 days after vaccination, or at 50 days or more after vaccination. In exemplary embodiments, the level or concentration occurs, or is achieved, after a single dose of the vaccine has been administered to the subject. In other embodiments, the level or concentration occurs, or is achieved, after multiple administrations, for example, after the first and second administrations (e.g., booster doses). In exemplary embodiments, the antibody level or concentration is determined or measured by an enzyme-linked immunosorbent assay (ELISA). In exemplary embodiments, the antibody level or concentration is determined or measured by a neutralization assay, for example, by a micro-neutralization assay.A nucleic acid vaccine comprising one or more RNA polynucleotides having an open reading frame encoding a first antigen polypeptide or a chain polypeptide, wherein the RNA polynucleotide is present in a formulation for in vivo administration to a recipient, has a stabilizing element or is formulated with an adjuvant and induces a high antibody titer that persists longer than the antibody titer induced by an mRNA vaccine encoding the first antigen polypeptide. Also provided is the nucleic acid vaccine. In some embodiments, the RNA polynucleotide is formulated to generate neutralizing antibodies within one week of a single administration. In some embodiments, the adjuvant is selected from cationic peptides and immunostimulatory nucleic acids. In some embodiments, the cationic peptide is protamine.

[0340] An immunotherapeutic agent comprising a nucleic acid vaccine comprising one or more RNA polynucleotides having an open reading frame encoding a first antigen polypeptide or a chain polypeptide, the RNA polynucleotide being present in a formulation for in vivo administration to a recipient, and the level of antigen expression in the recipient being such that it has a stabilizing element or is formulated with an adjuvant and significantly exceeds the level of antigen expression resulting from an mRNA vaccine encoding the first antigen polypeptide.

[0341] Another aspect provides a nucleic acid vaccine comprising one or more RNA polynucleotides having an open reading frame encoding a first antigen polypeptide or a chain polypeptide, the vaccine having at least 10-fold less RNA polynucleotide than is required for an unmodified mRNA vaccine to provide an equivalent antibody titer, the vaccine comprising at least one chemical modification or optionally no nucleotide modification. In some embodiments, the RNA polynucleotide is present at a dosage of 25 to 100 micrograms.

[0342] Aspects of the invention also provide a vaccine dosage unit formulated for delivery to a human subject, comprising from 10 μg to 400 μg of one or more RNA polynucleotides having an open reading frame encoding a first antigen polypeptide or a chain polypeptide, which may or may not contain at least one chemical modification, or optionally nucleotide modifications, and a pharmaceutically acceptable carrier or additive. In some embodiments, the vaccine further comprises cationic lipid nanoparticles.

[0343] Aspects of the invention provide a method of creating, maintaining or restoring antigenic memory against a tumor in an individual or population of individuals, the method comprising administering to the individual or population of individuals: (a) at least one RNA polynucleotide comprising at least one chemical modification or optionally no nucleotide modifications and comprising two or more codon-optimized open reading frames encoding a series of reference antigen polypeptides; and (b) optionally, an antigenic memory booster nucleic acid vaccine comprising a pharmaceutically acceptable carrier or additive. In some embodiments, the vaccine is administered to the individual via a route selected from the group consisting of intramuscular, intradermal, and subcutaneous administration. In some embodiments, the administering step comprises contacting the muscle tissue of the subject with an apparatus suitable for injection of the composition. In some embodiments, the administering step comprises contacting the muscle tissue of the subject with an apparatus suitable for injection of the composition in combination with electroporation.

[0344] Aspects of the invention provide a method of vaccinating a subject, the method comprising administering to the subject a single dose of from 25 μg / kg to 400 μg / kg of a nucleic acid vaccine comprising one or more RNA polynucleotides having an open reading frame encoding a first antigen polypeptide or a chain polypeptide in an effective amount to vaccinate the subject.

[0345] Another aspect provides a nucleic acid vaccine having an open reading frame comprising at least one chemical modification, wherein the open reading frame comprises one or more RNA polynucleotides encoding a first antigen polypeptide or a chain polypeptide, and the nucleic acid vaccine has at least 10-fold less RNA polynucleotides than are required for an unmodified mRNA vaccine to provide an equivalent antibody titer. In some embodiments, the RNA polynucleotides are present at a dosage of 25 to 100 micrograms.

[0346] In some embodiments, an exemplary immunotherapeutic agent can comprise one or more interfering RNAs that can be administered in combination with a compound of Formula I'. As used herein, "RNA interfering agent" is defined as any agent that interferes with or inhibits the expression of a target biomarker gene by RNA interference (RNAi). Such RNA interfering agents include, but are not limited to, nucleic acid molecules comprising RNA molecules homologous to the target biomarker gene of the invention, or fragments thereof, short interfering RNAs (siRNAs), and small molecules that interfere with or inhibit the expression of the target biomarker nucleic acid by RNA interference (RNAi). "Short interfering RNAs" (siRNAs), also referred to herein as "small interfering RNAs", are, for example It is defined as an agent that functions to inhibit the expression of a target biomarker nucleic acid by RNAi. siRNA can be chemically synthesized, generated by in vitro transcription, or generated within a host cell. In one embodiment, the siRNA is a double-stranded RNA (dsRNA) molecule that is about 15 to about 40 nucleotides in length, preferably about 15 to about 28 nucleotides, more preferably about 19 to about 25 nucleotides in length, and more preferably about 19, 20, 21, or 22 nucleotides in length, and contains 3' and / or 5' overhangs having a length of about 0, 1, 2, 3, 4, or 5 nucleotides on each strand. The lengths of the overhangs are independent between the two strands, i.e., the length of the overhang on one strand does not depend on the length of the overhang on the second strand. Preferably, the siRNA can promote RNA interference by degradation of the target messenger RNA (mRNA) or specific post-transcriptional gene silencing (PTGS).

[0347] Antisense oligonucleotides can be, for example, about 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 or more nucleotides in length. Antisense nucleic acids can be constructed using chemical synthesis and enzymatic ligation reactions using procedures known in the art. For example, antisense nucleic acids (e.g., antisense oligonucleotides) can be chemically synthesized using naturally occurring nucleotides or various modified nucleotides designed to increase the biological stability of the molecule or to increase the physical stability of the duplex formed between the antisense and sense nucleic acids, for example, phosphorothioate derivatives and acridine-substituted nucleotides can be used. Examples of modified nucleotides that can be used to generate antisense nucleic acids include 5-fluorouracil, 5-bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xanthine, 4-acetylcytosine, 5-(carboxyhydroxymethyl)uracil, 5-carboxymethylaminomethyl-2-thiouridine, 5-carboxymethylaminomethyluracil, dihydrouracil, beta-D-galactosylqueosine, inosine, N6-isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methyladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-adenine, 7-methylguanine, 5-methylaminomethyluracil, 5-methoxyaminomethyl-2-thiouracil, beta-D-mannosylqueosine, 5'-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N6-isopentenyladenine, uracil-5-oxyacetic acid (v), wybutoxosine, pseudouracil, queuosine, 2-thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, methyl ester of uracil-5-oxyacetic acid, uracil-5-oxyacetic acid (v), 5-methyl-2-thiouracil, 3-(3-amino-3-N-2-carboxypropyl)uracil, (acp3)w, and 2,6-diaminopurine.Alternatively, the antisense nucleic acid can be biologically produced using an expression vector in which the nucleic acid is subcloned in an antisense orientation (i.e., the RNA transcribed from this inserted nucleic acid is in an antisense orientation to the target nucleic acid of interest, as further described in the following subsections).

[0348] The antisense nucleic acid molecules of the invention are typically administered to a subject or generated in situ, and as a result, hybridize or bind to the cellular mRNA and / or genomic DNA encoding the polypeptide corresponding to the selected marker of the invention, thereby inhibiting the expression of the marker, for example, by inhibiting transcription and / or translation. This hybridization may be due to conventional nucleotide complementarity to form a stable double strand, or, for example, in the case of an antisense nucleic acid molecule that binds to a DNA double strand, may be through specific interactions in the major groove of the double helix. Examples of routes of administration of the antisense nucleic acid molecules of the invention include direct injection at the tissue site or injection of the antisense nucleic acid into the blood or bone marrow-related body fluids. Alternatively, the antisense nucleic acid molecule can be modified to target selected cells and then systemically administered. For example, in systemic administration, the antisense molecule can be modified to specifically bind to a receptor or antigen expressed on the surface of selected cells by, for example, binding the antisense nucleic acid molecule to a peptide or antibody that binds to a cell surface receptor or antigen. The antisense nucleic acid molecule can also be delivered to cells using the vectors described herein. To achieve a sufficient intracellular concentration of the antisense molecule, a vector construct in which the antisense nucleic acid molecule is placed under the control of a strong pol II or pol III promoter is preferred. Antigens that can be targeted to synthesize the corresponding antisense RNA molecule include any antigen specific to one or more tumors, for example, the antigens exemplified above with respect to cancer vaccines.

[0349]

[0350] In some embodiments, the combination of the immunotherapeutic agent and the compound of Formula I’ may include a bispecific antibody immunotherapeutic agent. The bispecific antibody may include a protein construct having a first antigen-binding portion and a second antigen-binding site that binds to a cytotoxic immune cell. The first antigen-binding site may bind to a tumor antigen specifically treated with the combination of the present invention. For example, the first antigen-binding portion may bind, inter alia, to non-limiting examples of tumor antigens selected from EGFR, HGFR, Her2, Ep-CAM, CD20, CD30, CD33, CD47, CD52, CD133, CEA, gpA33, mucin, TAG-72, CIX, PSMA, folate-binding protein, GD2, GD3, GM2, VEGF, VEGFR, integrin αVβ3, integrin α5β1, MUC1, ERBB2, ERBB3, MET, IGF1R, EPHA3, TRAILR1, TRAILR2, RANKL, FAP and tenascin. In some embodiments, the first antigen-binding portion has specificity for a protein or peptide that is overexpressed on tumor cells as compared to corresponding non-tumor cells. In some embodiments, the first antigen-binding portion has specificity for a protein that is overexpressed on tumor cells as compared to corresponding non-tumor cells. As used herein, “corresponding non-tumor cells” refers to non-tumor cells of the same cell type as the origin of the tumor cells. Note that such proteins are not necessarily different from tumor antigens. Non-limiting examples include carcinoembryonic antigen (CEA) overexpressed in many colon, rectal, breast, lung, pancreatic and gastrointestinal cancers; heregulin receptor (HER-2, neu or c-erbB-2) frequently overexpressed in breast, ovarian, colon, lung, prostate and cervical cancers; epidermal growth factor receptor (EGFR) highly expressed in a series of solid tumors including breast, head and neck, non-small cell lung and prostate solid tumors; asialoglycoprotein receptor; transferrin receptor; serpin enzyme complex receptor expressed on hepatocytes; fibroblast growth factor receptor (FGFR) overexpressed on pancreatic ductal adenocarcinoma cells; vascular endothelial growth factor receptor (VEGFR) in anti-angiogenic gene therapy; folate receptor selectively overexpressed in 90% of non-mucinous ovarian cancers; cell surface glycocalyx; carbohydrate receptor; and multimeric immunoglobulin receptor.

[0351] The second antigen-binding portion is any molecule that specifically binds to an antigen or protein or polypeptide expressed on the surface of cytotoxic immune cells (CIK cells). Exemplary non-limiting antigens expressed on the surface of cytotoxic immune cells suitable for use in the present disclosure include CD2, CD3, CD4, CD5, CD8, CD11a, CD11b, CD14, CD16a, CD27, CD28, CD45, CD45RA, CD56, CD62L, Fc receptor, LFA, LFA-1, TCRαβ, CCR7, macrophage inflammatory protein 1a, perforin, PD-1, PD-L1, PD-L2, or CTLA-4, LAG-3, OX40, 41BB, LIGHT, CD40, GITR, TGF-beta, TIM-3, SIRP-alpha, TIGIT, VSIG8, BTLA, SIGLEC7, SIGLEC9, ICOS, B7H3, B7H4, FAS, BTNL2, CD27 and Fas A ligand may be included. In some embodiments, the second antigen-binding portion binds to CD3 of a cytotoxic immune cell, such as a CIK cell. In some embodiments, the second antigen-binding portion binds to CD56 of a cytotoxic immune cell. In some embodiments, the second antigen-binding portion binds to an Fc receptor of a cytotoxic immune cell. In some embodiments, the Fc region of the bispecific antibody binds to an Fc receptor of a cytotoxic immune cell. In some embodiments, the second antigen-binding portion is any molecule that specifically binds to an antigen expressed on the surface of a cytotoxic immune cell (e.g., a CIK cell). The second antigen-binding portion is specific for an antigen on the cytotoxic immune cell. Exemplary cytotoxic immune cells include, but are not limited to, CIK cells, T cells, CD8+ T cells, activated T cells, monocytes, natural killer (NK) cells, NK T cells, lymphokine-activated killer (LAK) cells, macrophages, and dendritic cells. The second antigen-binding portion specifically binds to an antigen expressed on the surface of a cytotoxic immune cell. Exemplary non-limiting antigens expressed on the surface of a cytotoxic immune cell suitable for modulation in the present disclosure include CD2, CD3, CD4, CD5, CD8, CD11a, CD11b, CD14, CD16a, CD27, CD28, CD45, CD45RA, CD56, CD62L, Fc receptors, LFA, LFA-1, TCRαβ, CCR7, macrophage inflammatory protein 1a, perforin, PD-1, PD-L1, PD-L2, or CTLA-4, LAG-3, OX40, 41BB, LIGHT, CD40, GITR, TGF-beta, TIM-3, SIRP-alpha, TIGIT, VSIG8, BTLA, SIGLEC7, SIGLEC9, ICOS, B7H3, B7H4, FAS, BTNL2, CD27 and Fas ligand may be included. In other embodiments, the bispecific antibody modulator is an activator of a costimulatory molecule (e.g., an OX40 agonist). In one embodiment, the OX40 agonist is a bispecific antibody molecule against OX40 and another tumor antigen or costimulatory antigen.An OX40 agonist can be administered alone or in combination with other immunomodulatory factors, for example, in combination with an inhibitor (e.g., an antibody construct) of PD-1, PD-L1, CTLA-4, CEACAM (e.g., CEACAM-1, -3 and / or -5), TIM-3 or LAG-3. In some embodiments, the anti-OX40 antibody molecule is a bispecific antibody that binds to GITR and PD-1, PD-L1, CTLA-4, CEACAM (e.g., CEACAM-1, -3 and / or -5), TIM-3 or LAG-3. In one exemplary embodiment, the OX40 antibody molecule is administered in combination with an anti-PD-1 antibody molecule (e.g., an anti-PD-1 molecule as described herein). The OX40 antibody molecule and the anti-PD-1 antibody molecule may be in the form of separate antibody compositions or as a bispecific antibody molecule. In other embodiments, the OX40 agonist can be administered in combination with an agonist of other costimulatory molecules, for example, GITR, CD2, CD27, CD28, CDS, ICAM-1, LFA-1 (CD11a / CD18), ICOS (CD278), 4-1BB (CD137), CD30, CD40, BAFFR, HVEM, CD7, LIGHT, NKG2C, SLAMF7, NKp80, CD160, B7-H3, or CD83 ligand. In some embodiments, the second antigen-binding portion binds to an Fc receptor on a cytotoxic immune cell, for example, a CIK cell.

[0352] In some embodiments, the bispecific antibody immunotherapeutic has specificity for a tumor antigen and CIK cells, thus bringing tumor antigen-expressing tumor cells into proximity with CIK cells and resulting in the elimination of tumor cells through the anti-tumor cell cytotoxicity of the CIK cells. In some embodiments, the bispecific antibody has specificity for a tumor antigen but not for CIK cells; however, since the Fc region of the bispecific antibody can bind to the Fc receptor of CIK cells, it then brings tumor cells into proximity with CIK cells and results in the elimination of tumor cells through the anti-tumor cell cytotoxicity of the CIK cells. In some embodiments, the bispecific antibody has specificity for CIK cells but not for tumor cells; however, since the Fc region of the bispecific antibody can bind to the Fc receptor of tumor cells, it then brings tumor cells into proximity with CIK cells and results in the elimination of tumor cells through the anti-tumor cell cytotoxicity of the CIK cells.

[0353] In some embodiments, the combination of the immunotherapeutic and the compound of Formula I’ may include an immune cell-engaging multivalent antibody / fusion protein / construct immunotherapeutic. In various embodiments, exemplary immunotherapeutics may include recombinant constructs, e.g., immune cell-engaging multivalent antibodies / fusion proteins / constructs that may include any engineered antibody that does not mimic the native IgG structure. In this case, various strategies for multimerizing antibody fragments are utilized. For example, by shortening the peptide linker between V domains, scFvs can be self-associated to form dimers (diabodies, 55 kDa). Bispecific diabodies are formed by non-covalent association of two VHA-VLB and VHB-VLA fragments expressed in the same cell. This results in the formation of a heterodimer with two different binding sites. Single-chain diabodies (sc-diabodies) are bispecific molecules in which the VHA-VLB and VHB-VLA fragments are linked by an additional third linker. Tandem-diabodies (Tandab) are tetravalent bispecific antibodies made from two sc diabodies.

[0354] Also included are diabodies known in the art. This 130 kDa molecule is formed by fusing a diabody to the N-terminus of the CH3 domain of IgG, resulting in an IgG-like structure. Further diabody derivatives are triabodies and tetra-bodies that are folded by shortening the linker to <5 or to 0-2 residues, resulting in trimeric and tetrameric fragments. Also exemplified is a (scFv) 2 construct known as a "bispecific T cell engager" (BITE). A BITE is a bispecific single-chain antibody consisting of two scFv antibody fragments linked via a flexible linker and directed against a surface antigen on a target cell and CD3 on a T cell. Also exemplified are bivalent (Fab)2 and trivalent (Fab)3 antibody formats. Also exemplified are minibodies and trimerbodies generated from scFv. Exemplary constructs useful for targeting tumor antigens can include one or more of diabodies, single-chain (sc)-diabodies (scFv)2, miniantibodies, minibodies, barnase-barstar, scFv-Fc, sc(Fab)2, trimeric antibody constructs, triabody antibody constructs, trimerbody antibody constructs, tribody antibody constructs, Collabody antibody constructs, (scFv-TNFa)3, F(ab)3 / DNL. In each of these exemplary constructs, at least one binding moiety can bind to an antigen or protein or polypeptide expressed on the surface of a cytotoxic immune cell, and at least one binding moiety specifically binds to an antigen on a cytotoxic immune cell. Exemplary cytotoxic immune cells include, but are not limited to, CIK cells, T cells, CD8+ T cells, activated T cells, monocytes, natural killer (NK) cells, NK T cells, lymphokine-activated killer (LAK) cells, macrophages, and dendritic cells.

[0355] In some embodiments, the combination of the immunotherapeutic agent and the compound of Formula I' can include a radiolabeled conjugate immunotherapeutic agent.

[0356] In various embodiments, a radioactive conjugate is coupled or otherwise attached to one or more radionuclides, and the binding of the radioactive conjugate to its target (a protein or molecule on or in a cancer cell) is such that it causes death or morbidity of the cancer cell, a small molecule or macromolecule (also referred to herein as a "cell targeting agent"), such as a polypeptide, antibody or antibody fragment thereof. In various embodiments, the radioactive conjugate may be a cell targeting agent labeled with a radionuclide, or the cell targeting agent may be coupled or otherwise attached to a particle, micro-particle or nano-particle containing a plurality of radionuclides, where the radionuclides are the same or different. Methods for synthesizing radioactive conjugates are known in the art and may include conjugates of immunoglobulin or antigen-binding portions thereof to toxic radionuclides.

[0357] In some embodiments, the molecule that binds to the cancer cell may also be known as a "cell targeting drug." As used herein, exemplary cell targeting drugs may enable a drug-containing nanoparticle or a radionuclide to target a specific cell type of interest. Examples of cell targeting drugs include, but are not limited to, small molecules (e.g., folic acid, adenosine, purine) and macromolecules (e.g., peptides or antibodies) that bind to or target tumor-associated antigens. Examples of tumor-associated antigens include, but are not limited to, adenosine receptor, alpha v beta 3, aminopeptidase P, alpha-fetoprotein, cancer antigen 125, carcinoembryonic antigen, caveolin-1, chemokine receptor, clusterin, carcinoembryonic antigen, CD20, epithelial tumor antigen, melanoma-associated antigen, Ras, p53, Her2 / Neu, ErbB2, ErbB3, ErbB4, folate receptor, prostate-specific membrane antigen, prostate-specific antigen, purine receptor, radiation-inducible cell surface receptor, serpin B3, serpin B4, squamous cell carcinoma antigen, thrombospondin, tumor antigen 4, tumor-associated glycoprotein 72, thioredoxin, and tyrosine kinase. In some embodiments, the cell targeting drug is folic acid or a folic acid derivative that specifically binds to the folate receptor (FR). In some embodiments, the cell targeting drug is an antibody, bispecific antibody, trispecific antibody, or antigen-binding construct thereof that specifically binds to a cancer antigen selected from, inter alia, EGFR, HGFR, Her2, Ep-CAM, CD20, CD30, CD33, CD47, CD52, CD133, CEA, gpA33, mucin, TAG-72, CIX, PSMA, folate-binding protein, GD2, GD3, GM2, VEGF.VEGFR, integrin alphaVbeta3, integrin alpha5beta1, MUC1, ERBB2, ERBB3, MET, IGF1R, EPHA3, TRAILR1, TRAILR2, RANKL, FAP, and tenascin.

[0358] The use of folic acid as a target drug in a radioactive conjugate also allows both tumor cells and regulatory T (Treg) cells to be targeted for degradation. It is well recognized that a large number of Treg cells suppress tumor immunity. Specifically, Treg cells suppress (foreign and self) reactive T cells without killing them through contact dependence or cytokine (e.g., IL-10, TGF-beta, etc.) secretion. FR4 is selectively upregulated in Treg cells. Antibody blockade of FR4 has been shown to deplete Treg cells in tumor-bearing mice and induce tumor immunity. Therefore, folic acid-coated PBM nanoparticles carrying a cytotoxic agent utilize FR-expressing cells to destroy them, and that destruction will inhibit tumor progression both directly (i.e., BrCa cells) and indirectly (i.e., breast tumor-associated and peripheral Treg cells).

[0359] In another further embodiment, the target drug can be, but is not limited to: an antibody or peptide that binds to a tumor-associated antigen consisting of, but not limited to: adenosine receptor, alpha v beta 3, aminopeptidase P, alpha-fetoprotein, cancer antigen 125, carcinoembryonic antigen, caveolin-1, chemokine receptor, clusterin, carcinoembryonic antigen, CD20, human growth factor receptor (HGFR), epithelial tumor antigen, melanoma-associated antigen, MUC1, Ras, p53, Her2 / Neu, ErbB2, ErbB3, ErbB4, folate receptor, prostate-specific membrane antigen, prostate-specific antigen, purine receptor, radiation-induced cell surface receptor, serpin B3, serpin B4, squamous cell carcinoma antigen, thrombospondin, tumor antigen 4, tumor-associated glycoprotein 72, tyrosinase, and tyrosine kinase, or an immunocyte-engaging multivalent antibody / fusion protein / construct.

[0360] In one embodiment, the treatment method includes co - administration of a compound as disclosed herein or a pharmaceutically acceptable salt thereof and at least one cytotoxic agent. As used herein, the term "cytotoxic agent" refers to a substance that inhibits or blocks cell function and / or results in cell death or destruction. Cytotoxic agents include, but are not limited to, radioisotopes (e.g., At 211 、I 131 、I 125 、Y 90 、Re 186 、Re 188 、Sm 153 、Bi 212 、P 32 、Pb 212 and radioisotopes of Lu); chemotherapeutic agents; proliferation inhibitors; enzymes such as nuclease and fragments thereof; and toxins such as small molecule toxins or enzyme - active toxins of bacterial, fungal, plant or animal origin (including fragments and / or variants thereof).

[0361] Exemplary cytotoxic agents can be selected from antimicrotubule agents, platinum coordination complexes, alkylating agents, antibiotics, topoisomerase II inhibitors, antimetabolites, topoisomerase I inhibitors, hormones and hormone analogs, signal transduction pathway inhibitors, non - receptor tyrosine kinase angiogenesis inhibitors, immunotherapeutic agents, apoptosis - promoting agents, inhibitors of LDH - A; inhibitors of fatty acid biosynthesis; cell cycle signal transduction inhibitors; HDAC inhibitors, proteasome inhibitors; and inhibitors of cancer metabolism.

[0362] "Chemotherapeutic agents" include chemical compounds useful in the treatment of cancer. Examples of chemotherapeutic agents include erlotinib (TARCEVA®, Genentech / OSI Pharm.), bortezomib (VELCADE®, Millennium Pharm.), disulfiram, epigallocatechin gallate, salinosporamide A, carfilzomib, 17-AAG (geldanamycin), radicicol, lactate dehydrogenase A (LDH-A), fulvestrant (FASLODEX®, AstraZeneca), sunitinib (SUTENT®, Pfizer / Sugen), letrozole (FEMARA®, Novartis), imatinib mesylate (GLEEVEC®, Novartis), finasunate (VATALANIB®, Novartis), oxaliplatin (ELOXATIN®, Sanofi), 5-FU (5-fluorouracil), leucovorin, rapamycin (sirolimus, RAPAMUNE®, Wyeth), lapatinib (TYKERB®, GSK572016, Glaxo Smith Kline), lonafarnib (SCH66336), sorafenib (NEXAVAR®, Bayer Labs), gefitinib (IRESSA®, AstraZeneca), AG1478; alkylating agents such as thiotepa and CYTOXAN®; cyclophosphamide; alkyl sulfonates such as busulfan, improsulfan and piposulfan; aziridines such as benzodopa, carboquone, meturedopa, and uredopa; ethyleneimines and methylamelamines including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, and trimethylomelamine; acetogenins (especially, bullatacin and bullatacinone); camptothecin (including topotecan and irinotecan); bryostatin; calistatin; CC-1065 (including its adozelesin, carzelesin and bizelesin synthetic analogs); cryptophycins (especially, cryptophycin 1 and cryptophycin 8); corticosteroids (including prednisone and prednisolone);Cyproterone acetate; 5 alpha-reductase including finasteride and dutasteride); vorinostat, romidepsin, panobinostat, valproic acid, mocetinostat, dacostatins; aldosterone, talactomycin (synthetic analogs including KW-2189 and CB1-TM1)); erythrovidin; pancratistatin; sarcodictyin; spongistatin; nitrogen mustards such as chlorambucil, chlomaphazine, chlorophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trophosphamide, uracil mustard; nitrosoureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimnustine; enediyne antibiotics (e.g., calicheamicin, especially antibiotics such as calicheamicin gamma 1I and calicheamicin omega 1I (Angew Chem.Intl.Ed.Engl. 1994 33:183-186)); dynemicin A including dynemicin; bisphosphonates such as clodronate; esperamicin; ; furthermore, neocarzinostatin chromophore and related chromophore of pigment protein-engineered antibiotics, aclacinomysin, actinomycin, authramycin, azaserine, bleomycin, cactinomycin, carabicin, caminomycin, carzinophilin, chromomycinis, daunorubicin, doxorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, ADRIAMYCIN (registered trademark) (doxorubicin), morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin, epirubicin, esorubicin, idarubicin, marcellomycin, mitomycin such as mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, porfiromycin, puromycin, keramycin, rhodomycin, streptozocin, streptozotocin, tubercidin, ubenimex, dinostatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogs such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, didoxyridine, doxifluridine, enocitabine, floxuridine; androgens such as calusterone, drostanolone propionate, epitioestanol, mepitiostane, testolactone; anti-adrenal drugs such as aminoglutethimide, mitotane, trilostane; folic acid supplements such as folinic acid; aceglatone; aldophosphamide glycoside; aminolevulinic acid;Eniluracil; amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demeclocycline; diaziquone; elfomithine; elliptinium acetate; epothilone; etoglucid; gallium nitrate; hydroxyurea; lentinan; lonidainine; maytansinoids such as maytansine and ansamitocin; mitoguazone; mitoxantrone; mopidamnol; nitraerine; pentostatin; phenamet; pirarubicin; losoxantrone; podophyllinic acid; 2-ethylhydrazide; procarbazine; PSK (registered trademark) polysaccharide complex (JHS Natural; Products, Eugene, Ore.); razoxane; rhizoxin; sizofuran; spirogermanium; tenuazonic acid; triazicon; 2,2’,2’’-trichloroethylamine; trichothecene (particularly, T-2 toxin, verracurin A, roridin A, and anguidine); urethane; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside (“Ara-C”); cyclophosphamide; thiotepa; taxoids, for example, taxol (paclitaxel; Bristol-Myers Squibb Oncology, Princeton, N.J.), ABRAXANE® (cremophor-free), albumin-engineered nanoparticle formulation of paclitaxel (American Pharmaceutical Partners, Schaumberg, Ill.), and TAXOTERE® (docetaxel, doxetaxel, Sanofi-Aventis); chlorambucil; GEMZAR® (gemcitabine); 6-thioguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin and carboplatin; vinblastine; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; NAVE LBINE® (vinorelbine); novantrone; teniposide; edatrexate; daunomycin; aminopterin; capecitabine (XELODA®); ibandronate; CPT-11; topoisomerase inhibitor RFS2000; difluoromethylornithine (DMFO); retinoids such as retinoic acid; and pharmaceutically acceptable salts, acids, and derivatives of any of the foregoing are included.

[0363] Chemotherapeutic agents include: (i) antihormonal agents that act to regulate or inhibit the hormonal action on tumors, such as antiestrogens and selective estrogen receptor modulators (SERMs), for example, tamoxifen (including NOLVADEX (registered trademark), tamoxifen citrate), raloxifene, droloxifene, iodoxyfene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and FARESTON (registered trademark) (toremifine citrate); (ii) aromatase inhibitors that inhibit the enzyme aromatase, which regulates estrogen production in the adrenal glands, for example, 4(5)-imidazole, aminoglutethimide, MEGASE (registered trademark) (megestrol acetate), AROMASIN (registered trademark) (exemestane, Pfizer), formestanie, fadrozole, RIVISOR (registered trademark) (vorozole), FEMARA (registered trademark) (letrozole, Novartis), and ARIMIDEX (registered trademark) (anastrozole, AstraZeneca); (iii) antiandrogens such as flutamide, nilutamide, bicalutamide, leuprorelin, and goserelin; buserelin, tripterelin, medroxyprogesterone acetate, diethylstilbestrol, Premarin, fluoxymesterone, all-trans retinoic acid, fenretinide, and further troxacitabine (a 1,3-dioxolane nucleoside cytosine analog); (iv) protein kinase inhibitors; (v) lipid kinase inhibitors; (vi) antisense oligonucleotides, for example, those that inhibit the expression of genes in signal transduction pathways involved in abnormal cell growth, such as PKC-alpha, Ralf, and H-Ras; (vii) ribozymes such as VEGF expression inhibitors (e.g., ANGIOZYME (registered trademark)) and HER2 expression inhibitors; (viii) gene therapy vaccines, for example, vaccines such as ALLOVECTIN (registered trademark), LEUVECTIN (registered trademark), and VAXID (registered trademark);PROLEUKIN (registered trademark), rIL-2; topoisomerase 1 inhibitors such as LURTOTECAN (registered trademark); ABARELIX (registered trademark), and (ix) also include any pharmaceutically acceptable salts, acids and derivatives of the above.

[0364] Chemotherapeutic agents include alemtuzumab (Campath), bevacizumab (AVASTIN (registered trademark), Genentech); cetuximab (ERBITUX (registered trademark), Imclone); panitumumab (VECTIBIX (registered trademark), Amgen), rituximab (RITUXAN (registered trademark), Genentech / Biogen Idec), pertuzumab (OMNITARG (registered trademark), 2C4, Genentech), trastuzumab (HERCEPTIN (registered trademark), Genentech), tositumomab (Bexxar, Corixia) and the antibodies as described above, and antibody-drug conjugates, gemtuzumab ozogamicin (MYLOTARG (registered trademark), Wyeth) are also included. Additional humanized monoclonal antibodies having therapeutic ability as an active substance in combination with the compounds of the present invention include apolizumab, aselizumab, atorizumab, bavituximab, bivatuzumab mertansine, canertuzumab mertansine, cedelizumab, certolizumab pegol, cidfusituzumab, cidtuzumab, daclizumab, eclizumab, efalizumab, epratuzumab, erlizumab, felvizu mab), fontrizumab, gemtuzumab ozogamicin, inotuzumab ozogamicin, ipilimumab, labelizumab, lintuzumab, matuzumab, mepolizumab, motavizumab (motovizumab), natalizumab, nimotuzumab, nivolumab, nolovizumab (nolovizumab), numavizumab (numavizumab), ocrelizumab, omalizumab, palivizumab, pascolizumab (pascolizumab), pecfusituzumab, pectuzumab, pexelizumab, ralivizumab (ralivizumab), ranibizumab, reslivizumab (reslivizumab), reslizumab, resyvizumab (resyvizumab), rovelizumab, ruplizumab, sibrotuzumab, ciplizumab, sontuzumab (sontuzumab), tacatuzumab tetraxetan, tadocizumab, talizumab, tefibazumab, tocilizumab, toralizumab (toralizumab), tucotuzumab celmoleukin, tucusituzumab, umavizumab, ultoxizumab, ustekinumab, besilesizumab, and a recombinant, exclusively human sequence full-length IgG genetically modified to recognize the interleukin 12p40 protein 1 including the anti-interleukin 12 (ABT-8744695, Wyeth Research and Abbott Laboratories), which is a lambda antibody

[0365] Chemotherapeutic agents include EGFR inhibitors; small molecule HER2 tyrosine kinase inhibitors such as mubritinib (TAK165, Takeda); CP-724.714 (Axon Medchem BV, an oral selective inhibitor of the ErbB2 receptor tyrosine kinase); dual HER inhibitors such as EKB-569 (available from Wyeth) that preferentially binds to EGFR but inhibits both HER2 and EGFR overexpressing cells; lapatinib (GSK572016, available from Glaxo-SmithKline), an oral HER2 and EGFR tyrosine kinase inhibitor; PKI-166 (available from Novartis); pan-HER inhibitors such as canertinib (CI-1033, Pharmacia); Raf-1 inhibitors such as the antisense drug ISIS-5132 available from ISIS Pharmaceuticals that inhibits Raf-1 signal transduction; non-HER targeted TK inhibitors such as imatinib mesylate (GLEEVEC®, available from Glaxo SmithKline); multi-targeted tyrosine kinase inhibitors such as sunitinib (SUTENT®, available from Pfizer); VEGF receptor tyrosine kinase inhibitors such as vatalanib (PTK787 / ZK222584, available from Novartis / Schering AG); MAPK extracellular regulated kinase 1 inhibitor CI-1040 (available from Pharmacia); quinazolines such as PD153035, 4-(3-chloroanilino)quinazoline; pyridopyrimidines; pyrimidopyrimidines; pyrrolopyrimidines such as CGP59326, CGP60261 and CGP62706; pyrazolopyrimidines, 4-(phenylamino)-7H-pyrrolo[2,3-d]pyrimidine; curcumin (diferuloylmethane, 4,5-bis(4-fluoroanilino)phthalimide); tyrphostine containing a nitrothiophene moiety; antisense molecules (e.g., those that bind to HER coding nucleic acids); quinoxaline (U.S. Patent No. 5,804,396); tryphostins (U.S. Patent No. 5,804,396); Affinitac (ISIS 3521, Isis / Lilly); PKI166 (Novartis); semaxinib (Pfizer);INC-1C11 (Imclone), rapamycin (sirolimus, RAPAMUNE®); or any of the following patent publications: U.S. Patent No. 5,804,396, WO1999 / 09016 (American Cyanamid), WO1998 / 43960 (American; Cyanamid), WO1997 / 38983 (Warner Lambert), WO1999 / 06378 (Warner Lambert), WO1999 / 06396 (Warner Lambert), WO1996 / 30347 (Pfizer, Inc), WO1996 / 33978 (Zeneca), WO1996 / 3397 (Zeneca), and WO1996 / 33980 (Zeneca), also includes "tyrosine kinase inhibitors" described in any of them. Tyrosine kinase inhibitors include erlotinib (Tarceva®), gefitinib (Iressa®), dasatinib (Sprycel®), nilotinib (Tasigna®), crizotinib (Xalkori®), ruxolitinib (Jakafi®), vemurafenib (Zelboraf®), vandetanib (Caprelsa®), pazopanib (Votrient®), afatinib, alisertib, amuvatinib, axitinib, bosutinib, brivanib, canertinib, cabozantinib, cediranib, clenolanib, dabrafenib, dacomitinib, danusertib, dovitinib, foretinib, ganetespib, ibrutinib, iniparib, lenvatinib, linifanib, linsitinib, masitinib, momelotinib, motesanib, neratinib, niraparib, oprozomib, olaparib, pictilisib, ponatinib, quizartinib, regorafenib, rigosertib, rucaparib, saracatinib, saredutant, tandutinib, tasocitinib, teratinib, tibantinib, tiboxanib, tofacitinib, trametinib, veriparib, bisindolylmaleimide, borosertib, cobimetinib (Cotellic®), etc.

[0366] Chemotherapeutic agents include dexamethasone, interferon, colchicine, metoprine, cyclosporine, amphotericin, metronidazole, alemtuzumab, alitretinoin, allopurinol, amifostine, arsenic trioxide, asparaginase, live BCG, bevacuzimab, bexarotene, cladribine, clofarabine, darbepoetin alfa, denileukin, dexrazoxane, epoetin alfa, erlotinib, filgrastim, histrelin acetate, ibritumomab, interferon alpha-2a, interferon alpha-2b, lenalidomide, levamisole, mesna, methoxsalen, nandrolone, nelarabine, nolfetumomab, oprelvekin, palifermin, pamidronate, pegademase, pegaspargase, pegfilgrastim, pemetrexed disodium, plicamycin, porfimer sodium, quinacrine, rasburicase, sargramostim, temozolomide, VM-26, 6-TG, tamoxifen, tretinoin, ATRA, valrubicin, zoledronate, and zoledronic acid, as well as pharmaceutically acceptable salts thereof.

[0367] Chemotherapeutic agents include hydrocortisone, hydrocortisone acetate, cortisone acetate, tixocortol pivalate, triamcinolone acetonide, triamcinolone alcohol, mometasone, amcinonide, budesonide, desonide, fluocinide, fluocinolone acetonide, betamethasone, betamethasone sodium phosphate, dexamethasone, dexamethasone sodium phosphate, fludrocortisone, hydrocortisone-17-butyrate, hydrocortisone-17-valerate, aclometasone dipropionate, betamethasone valerate, betamethasone dipropionate, prednicarbate, clobetasone-17-butyrate, clobetasol-17-propionate, fludrocortisone caproate, fludrocortisone pivalate and flucloronide acetate; immune-selective anti-inflammatory peptides (ImSAIDs) such as phenylalanine-glutamine-glycine (FEG) and its ...

Claims

1. A compound of formula B-1: 【Chemical 1】 or a pharmaceutically acceptable salt thereof, wherein R ａ１ is optionally substituted with (C １ to C ６ ) alkyl, R １ is -CN, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted phenyl, optionally substituted 4- to 6-membered heterocycloalkyl, optionally substituted 5- to 6-membered heteroaryl, -SO ２ -(C １ ~C ６ )alkyl, -SONH ２ NH ２ or -SO ２ -NH(C １ ~C ６ )alkyl, or P(O)((C １ ~C ６ )alkyl) ２ and is, or R １ is 【Chemical 203】 wherein 【Chemical 204】 represents a bond, wherein Y １ does not exist Y ２ is absent or is -O-, -NH-, -NHO-, -NH-NH-, or -N-(C １ ~C ６ )alkyl- or Y ２ is optionally replaced 【Chemical 205】 wherein ring A is a 4-membered ring 【Chemical 206】 represents a bond Z １ is O, NH, N-(C １ ~C ６ )alkyl, NHO, or NO-(C １ ~C ６ )alkyl and R ａ is -H, -(C １ ~C ６ )alkyl, 4-6 membered heterocycloalkyl, 3-6 membered cycloalkyl, -(C ２ ~C ６ )alkylene-OH, -CH ２ CHOH-(C ２ ~C ６ )alkylene-OH, -(C ２ ~C ６ )alkylene-NH ２ , -(C ２ ~C ６ )alkylene-NH(C １ ~C ６ )alkyl, -(C ２ ~C ６ )alkylene-N((C １ ~C ６ )alkyl) ２ , or -(C ２ ~C ６ )alkylene-N-(4-6 membered heterocycloalkyl), where the heterocycloalkyl is optionally substituted, R ３ is -H or halo, R ４ is -H, halo, (C １ ~C ６ -C) alkyl, (C １ ~C ６ -C) alkoxy, or (C １ ~C ６ -C) haloalkyl, and R １４ is -H, Y is -O- subscript n is an integer of 1, 2, 3, or 4 subscript m is an integer of 1, 2, 3, 4, or 5, and subscript p is an integer of 0, 1, 2, 3, or 4 Here, in each occurrence, "optionally substituted" means unsubstituted or substituted with a substituent selected from halogen, -CN, -OH, -OR ａａ , -COOR ａａ , (C １ ~C １０ )alkyl, (C ３ ~C １０ )carbocycloalkyl, 3- to 14-membered heterocycloalkyl, (C ６ ~C １４ )aryl, and 5- to 14-membered heteroaryl, where R ａａ is (C １ ~C １０ )alkyl, provided that the compound is not 1-N'-(4-fluorophenyl)-1-N-[4-[7-methoxy-6-(methylcarbamoyl)quinolin-4-yl]oxyphenyl]cyclopropane-1,1-dicarboxamide or a pharmaceutically acceptable salt thereof the compound or a pharmaceutically acceptable salt thereof.

2. R ａ１ is (C １ to C ６ ) alkyl, R １ is -CN, cyclopropyl optionally substituted, phenyl optionally substituted, 4- to 6-membered azetidinyl optionally substituted, pyrrolidinyl optionally substituted, piperidinyl optionally substituted, oxetanyl optionally substituted, oxazolyl optionally substituted, pyridinyl optionally substituted, imidazolyl optionally substituted, pyrrolyl optionally substituted, furanyl optionally substituted, pyrazolyl optionally substituted, oxadiazolyl optionally substituted, -SO ２ -(C １ to C ６ )alkyl, -SONH ２ NH ２ is -SO ２ -NH(C １ to C ６ )alkyl, or P(O)((C １ to C ６ )alkyl) ２ or R １ is 【Chemical 207】 wherein Y ２ is O, NH, NHO, NH-NH, or N-(C １ ~C ６ ) alkyl, or Y ２ is an azetidinyl optionally substituted with Z is O, NH, or N-(C １ ~C ６ ) alkyl, and R ａ is H, (C １ -C ６ )alkyl, -(C ２ -C ６ )alkylene-OH, -CH ２ CHOH-(C ２ -C ６ )alkylene-OH, -(C ２ -C ６ )alkylene-NH ２ , -(C ２ -C ６ )alkylene-NH(C １ -C ６ )alkyl, -(C ２ -C ６ )alkylene-N((C １ -C ６ )alkyl) ２ , -(C ２ -C ６ )alkylene-N-(4-6 membered heterocycloalkyl), and 4-6 membered heterocycloalkyl, wherein the heterocycloalkyl is optionally substituted, the compound according to claim 1 or a pharmaceutically acceptable salt thereof.

3. R ａ１ The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 or 2, wherein R is methoxy.

4. A compound of formula C: 【Chemical Formula 4】 or a pharmaceutically acceptable salt thereof [wherein Y ２ is O, NH, NHO, NH-NH, or N-(C １ ~C ６ ) alkyl, or Y ２ is an azetidinyl which is optionally substituted, Z １ is O, NH, NO-(C １ ~C ６ )alkyl, or N-(C １ ~C ６ )alkyl, and R ａ is H, (C １ ~C ６ )alkyl, -(C ２ ~C ６ )alkylene-OH, -CH ２ CHOH-(C ２ ~C ６ )alkylene-OH, -(C ２ ~C ６ )alkylene-NH ２ , -(C ２ ~C ６ )alkylene-NH(C １ ~C ６ )alkyl, -(C ２ ~C ６ )alkylene-N((C １ ~C ６ )alkyl) ２ , -(C ２ ~C ６ )alkylene - optionally substituted 4- to 6-membered heterocycloalkyl), or optionally substituted 4- to 6-membered heterocycloalkyl, and R ２ is (C １ to C ６ ) alkoxy], the compound according to claim 1 or a pharmaceutically acceptable salt thereof.

5. R ａ is H, methyl, ethyl, -(C ２ ~C ６ ) alkylene-OH, -CH ２ CHOH -(C ２ ~C ６ )-alkylene-OH, -(C ２ ~C ６ )-alkylene-NH ２ , -(C ２ ~C ６ )-alkylene-NHMe, -(C ２ ~C ６ )-alkylene-N(Me) ２ , -(C １ ~C ６ )-alkylene-morpholinyl), -(C １ ~C ６ )-alkylene-piperidinyl), (C １ ~C ６ )-alkylene-(optionally substituted pyrrolidinyl), optionally substituted azetidinyl, or optionally substituted oxetanyl, the compound according to claim 4 or a pharmaceutically acceptable salt thereof.

6. A compound of formula C-1 【Chemical Formula 6】 or a pharmaceutically acceptable salt thereof, the compound according to claim 4 or claim 5.

7. A compound of formula E: [Chemical Formula 7] or a pharmaceutically acceptable salt thereof [wherein Ring A is optionally substituted (C ６ ~C １０ ) aryl, optionally substituted (C ３ ~C １０ ) cycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 4- to 10-membered heterocycloalkyl, and R ２ is methoxy], the compound according to claim 1.

8. A compound of formula F: 【Chemical 8】 or a pharmaceutically acceptable salt thereof, the compound according to claim 1.

9. R ａ１ is methyl, and R １ is -CN, optionally substituted cyclopropyl, optionally substituted phenyl, optionally substituted 4-6 membered azetidinyl, optionally substituted pyrrolidinyl, optionally substituted piperidinyl, optionally substituted oxetanyl, optionally substituted oxazolyl, optionally substituted pyridinyl, optionally substituted imidazolyl, optionally substituted pyrrolyl, optionally substituted furanyl, optionally substituted pyrazolyl, optionally substituted oxadiazolyl, -SO ２ -(C １ -C ６ )alkyl, -SONH ２ NH ２ -SO ２ -NH(C １ -C ６ )alkyl, or P(O)((C １ -C ６ )alkyl) ２ or R １ is 【Chemical 224】 wherein: Y ２ is O, NH, NHO, NH-NH, or N-(C １ ~C ６ )alkyl, or Y ２ is an azetidinyl optionally substituted with Z １ is O, NH, or N-(C １ ~C ６ ) alkyl, and R ａ is H, (C １ ~C ６ )alkyl, -(C ２ ~C ６ )alkylene-OH, -CH ２ CHOH-(C ２ ~C ６ )alkylene-OH, -(C ２ ~C ６ )alkylene-NH ２ , -(C ２ ~C ６ )alkylene-NH(C １ ~C ６ )alkyl, -(C ２ ~C ６ )alkylene-N((C １ ~C ６ )alkyl) ２ , -(C ２ ~C ６ )alkylene-N-(4- to 6-membered)heterocycloalkyl), and 4- to 6-membered heterocycloalkyl, wherein the heterocycloalkyl is optionally substituted, the compound according to claim 8 or a pharmaceutically acceptable salt thereof.

10. A compound of formula H: 【Chemical 10】 or a pharmaceutically acceptable salt thereof [wherein Y is -O- R １ is selected from -CN, -CO-NR ５ R ６ , -CO ２ R ７ , phenyl, 5- to 6-membered heteroaryl, C ３ -C ８ cycloalkyl, 4- to 6-membered heterocycloalkyl, -SO ２ NR ８ R ９ , and -(SO ２ )-C １ -C ６ alkyl, R ２ is C １ to C ６ alkoxy, R ３ is -H or halo, R ４ is -H or halo, R ５ and R ６ are each independently, -H, C １ ~C ６ alkyl, or C １ ~C ６ alkoxy, R ７ is -H or C １ ~C ６ alkyl, and R ８ and R ９ are each independently, -H or C １ ~C ６ alkyl]], and provided that the compound is not 1-N'-(4-fluorophenyl)-1-N-[4-[7-methoxy-6-(methylcarbamoyl)quinolin-4-yl]oxyphenyl]cyclopropane-1,1-dicarboxamide or a pharmaceutically acceptable salt thereof the compound or a pharmaceutically acceptable salt thereof.

11. R ３ The compound according to claim 10, or a pharmaceutically acceptable salt thereof, wherein R is H.

12. R ４ The compound according to claim 10, or a pharmaceutically acceptable salt thereof, wherein R is halo.

13. R ４ The compound according to claim 12, or a pharmaceutically acceptable salt thereof, wherein R is perfluoro.

14. R １ is -CN, -CO ２ H, -CO ２ Me, -CO-NHR ６ , -CO-NH ２ , or -CO-NMeR ６ The compound according to claim 10 or a pharmaceutically acceptable salt thereof, wherein it is

15. R １ is -CN, -(SO ２ )NH ２ -(SO ２ )CH ３ , 【Chemical 234-1】 【Chemical 234-2】 selected from, the compound according to claim 1 or a pharmaceutically acceptable salt thereof.

16. R １ is 【Chemical 238】 selected from, the compound according to claim 10 or a pharmaceutically acceptable salt thereof.

17. A compound of formula I-A: 【Chemical 17】 or a pharmaceutically acceptable salt thereof [wherein, R ６ is (C １ to C ６ ) alkyl, R ２ is (C １ to C ６ ) alkoxy, R ３ is -H or halo, and R ４ is halo] and provided that the compound is not 1-N'-(4-fluorophenyl)-1-N-[4-[7-methoxy-6-(methylcarbamoyl)quinolin-4-yl]oxyphenyl]cyclopropane-1,1-dicarboxamide or a pharmaceutically acceptable salt thereof the compound or a pharmaceutically acceptable salt thereof.

18. The compound is 【Table 6-1】 【Table 6-2】 【Table 6-3】 【Table 6-4】 【Table 6-5】 【Table 6-6】 【Table 6-7】 【Table 6-8】 The compound according to claim 1, or a pharmaceutically acceptable salt thereof, selected from

19. The compound is 【Table 7-1】 【Table 7-2】 【Table 7-3】 The compound according to claim 1, or a pharmaceutically acceptable salt thereof, selected from

20. A pharmaceutical composition comprising the compound according to any one of claims 1 to 19, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

21. A composition for use in a method for treating cancer, comprising the compound according to any one of claims 1 to 19 or the pharmaceutical composition according to claim 20.

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