Combination therapy and cancer treatment

Abstract

Combination therapies comprising selective protein kinase C inhibitors and cMET inhibitors are described, which are useful for treating metastatic uveal melanoma and other proliferative diseases (e.g., tumors with GNAQ or GNA11 mutations). The combination therapies can be provided by pharmaceuticals, methods of treatment, or kits. The combination therapies can include the use of protein kinase C inhibitors and cMET inhibitors. Methods useful for selecting patients or directing a course of treatment are also provided. In various examples, the protein kinase C can have a structure according to Formula II:

Description

[Background technology]

[0001] Uveolar melanoma is the most common primary intraocular tumor in adults. Although relatively rare, this cancer can be very deadly. Radiation, resection, and enucleation are widely used as first-line treatments, but these treatments are only moderately effective. Even after enucleation, the cancer can recur and spread in up to half of patients. Common sites of metastasis include the liver, as well as the lungs, bones, subcutaneous tissue, and lymph nodes. The overall 5-year survival rate is approximately 15%. For patients with metastatic disease, the median survival time is approximately 10 months. Currently, there are no approved drug therapies for the treatment of metastatic uveal melanoma. Metastatic uveal melanoma remains an incurable disease. See Non-Patent Literature 1.

[0002] Protein kinase C is being studied as a target for treating metastatic uveal melanoma. Approximately 90% of uveal melanoma tumors have mutations in the guanine nucleotide-binding proteins GNAQ or GNA11, which can function to activate PKC family proteins and downstream MAPK pathways involved in tumor growth. Non-uveal tumors can also exhibit such mutations. In humans, the PKC family comprises numerous different isoforms, each protein isoform conferring distinct regulatory and intracellular localization properties to the others. Human PKC isoforms include the "classical" calcium-dependent PKC alpha (α), beta-1 (βI), beta-2 (βII), and gamma (γ); the "novel" calcium-independent PKC delta (δ), epsilon (ε), eta (η), and theta (θ); and the "atypical" PKC zeta (ζ) and iota (ι).

[0003] Sotrastaurin is a maleimide-type PKC inhibitor that exhibits non-selective or "pan-selective" activity across the PKC isoform family and activity against several kinases outside the PKC family. See Patent Document 1. Another maleimide-type PKC inhibitor, Enzastaurin, is a PKC inhibitor that targets the classical calcium-dependent PKC isoform, specifically PKC isoform beta-1. See Patent Document 2. Neither compound has been approved for therapeutic use in humans.

[0004] IDE196 represents a new class of PKC inhibitor, exhibiting high efficacy against novel and classical PKC isoforms, being more active against novel calcium-independent PKC isoforms than against classical PKC isoforms, and delivering significant therapeutic effects against primary uveal melanoma. See Patent Document 3. However, metastatic uveal melanoma appears to be resistant to the development of therapeutic agents.

[0005] Therefore, there is an unmet need for therapies that can treat uveal melanoma, particularly metastatic uveal melanoma, as well as cancers characterized by tumors with GNAQ and GNA11 mutations. [Prior art documents] [Patent Documents]

[0006] [Patent Document 1] International Publication No. 2014 / 174478 [Patent Document 2] U.S. Patent Application Publication No. 2010 / 0267742 [Patent Document 3] International Publication No. 2019 / 053595 [Non-patent literature]

[0007] [Non-Patent Document 1] Croce et al.,Targeted Therapy of Uveal Melanoma:Recent Failures and New Perspectives,Cancers 2019,11(6),846 [Overview of the project]

[0008] These and other needs are met by the present invention relating to pharmaceuticals, compositions, methods, and kits useful for treating metastatic uveal melanoma, tumors with GNAQ mutations or GNA11 mutations, and other proliferative disorders. In various embodiments, the present invention includes combination therapies comprising protein kinase C inhibitors and cMET inhibitors.

[0009] For example, one aspect of the present invention relates to a method for treating cancer. In various embodiments, patients with metastatic uveal melanoma or tumors having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor") are selected, and a cMET inhibitor and a protein kinase C inhibitor are co-administered to the selected patients.

[0010] Protein kinase C inhibitors are expressed by formula II:

[0011] [ka]

[0012] or can be represented by a pharmaceutically acceptable salt thereof, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3Alkyl may optionally be substituted with one, two, three, or more substituents each independently selected from the group consisting of hydroxyl, halogen, and C 1~3 alkoxy (optionally substituted with one or more halogens); R 5 is H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 alkyl, and C 2~3 alkyl may optionally be substituted with one, two, three, or more substituents each independently selected from the group consisting of fluorine, hydroxyl, and C 1~3 alkoxy (optionally substituted with one or more halogens); R 5a and R 5b are each independently H, 2 H, and C 1~3 alkyl, and C 1~3 alkyl may optionally be substituted with one, two, three, or more substituents each independently selected from the group consisting of fluorine, hydroxyl, and C 1~3 alkoxy, or R 5a and R 5b together form a methylene or ethylene bridging group; R 5c and R 5d are each independently H, 2 H, fluorine, hydroxyl, C 1~3 alkoxy, and C 1~3 alkyl, and C 1~3 alkyl may optionally be substituted with one, two, three, or more substituents each independently selected from the group consisting of fluorine, hydroxyl, and C 1~3 alkoxy, or R 5c and R 5d together form a methylene or ethylene or -CH2-O- bridging group; R6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The alkoxy may be optionally substituted with one, two, three, or more halogens.

[0013] In another embodiment, the method may include treating cancer in patients having metastatic uveal melanoma showing elevated cMET, which can be determined by evaluating a biopsy of the metastatic uveal melanoma. Patients having such a form of metastatic uveal melanoma can be treated by co-administration of a cMET inhibitor and a protein kinase C inhibitor that can be represented by formula II above.

[0014] In yet another embodiment, the method may include treating cancer in patients having tumors with GNAQ mutations or GNA11 mutations ("GNAQ / 11 tumors") and exhibiting elevated cMET. GNAQ / 11 tumors may be primary GNAQ / 11 tumors or metastatic GNAQ / 11 tumors. The presence of elevated cMET in GNAQ / 11 tumors can be determined by evaluating a biopsy. Such patients can be treated with co-administration of a cMET inhibitor and a protein kinase C inhibitor that can be represented by formula II above.

[0015] The present invention also provides pharmaceuticals and kits useful for selecting and treating patients with proliferative disorders such as metastatic uveal melanoma or GNAQ / 11 tumors. The various products, kits, and compositions described herein incorporate combinations of agents comprising PKC inhibitors and cMET inhibitors.

[0016] A preferred example of a PKC inhibitor is 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide (compound A). Preferred examples of cMET inhibitors include crizotinib, capmatinib, cabozantinib, tivantinib, and any combination thereof. [Brief explanation of the drawing]

[0017] [Figure 1A-1]This chart shows the antagonistic effect of HGF on IDE196 (compound A, as defined below) in MEL-202 primary uveal melanoma cells. [Figure 1A-2] Same as above. [Figure 1A-3] Same as above. [Figure 1A-4] Same as above. [Figure 1B] This paper provides Western blots demonstrating the pharmacodynamic effects (individually and in combination) of IDE196 (compound A) and HGF in the MEL-202 primary uveal melanoma cell line. [Figure 2A-1] 92.1 This section provides a chart showing the antagonistic effect of HGF on IDE196 (compound A) in primary uveal melanoma cells. [Figure 2A-2] Same as above. [Figure 2A-3] Same as above. [Figure 2A-4] Same as above. [Figure 2B] 92.1 This paper provides Western blots showing the pharmacodynamic effects (individually and in combination) of IDE196 (compound A) and HGF in primary uveal melanoma cell lines. [Figure 3A-1] This chart shows the antagonistic effect of HGF on IDE196 (compound A) in MM28 metastatic uveal melanoma cells. [Figure 3A-2] Same as above. [Figure 3A-3] Same as above. [Figure 3A-4] Same as above. [Figure 3B] This provides Western blots demonstrating the pharmacodynamic effects (individually and in combination) of IDE196 (compound A) and HGF in the MM28 metastatic uveal melanoma cell line. [Figure 4] Western blots comparing the relative presence of cMET in MEL-202, 92.1, and MM28 uveal melanoma cell lines are provided. T-MET corresponds to total cMET concentration, and P-MET corresponds to phosphorylated cMET concentration. GAPDH is the reference standard. [Figure 5A-1]This chart shows the effects of PKC inhibitors, such as IDE196 (compound A), and cMET inhibitors (crizotinib, also abbreviated as "crizo" in the figure) on the survival rate of MEL-20 primary uveal melanoma cells in the presence of HGF. [Figure 5A-2] Same as above. [Figure 5A-3] Same as above. [Figure 5A-4] Same as above. [Figure 5B-1] This chart shows the effects of PKC inhibitors, such as IDE196 (compound A), and cMET inhibitors (capmatinib, also abbreviated as "cap" in the figure) on the survival rate of MEL-20 primary uveal melanoma cells in the presence of HGF. [Figure 5B-2] Same as above. [Figure 5B-3] Same as above. [Figure 5B-4] Same as above. [Figure 5C] This provides Western blots demonstrating the pharmacodynamic effects of drug therapies on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways. [Figure 6A-1] This chart shows the effects of PKC inhibitors, such as IDE196 (compound A), and cMET inhibitors (crizotinib) on the survival rate of 92.1 primary uveal melanoma cells in the presence of HGF. [Figure 6A-2] Same as above. [Figure 6A-3] Same as above. [Figure 6A-4] Same as above. [Figure 6B-1] This chart shows the effects of PKC inhibitors, such as IDE196 (compound A), and cMET inhibitors (capmatinib) on the survival rate of 92.1 primary uveal melanoma cells in the presence of HGF. [Figure 6B-2] Same as above. [Figure 6B-3] Same as above. [Figure 6B-4] Same as above. [Figure 6C]This provides Western blots demonstrating the pharmacodynamic effects of drug therapies on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways. [Figure 7A-1] This chart shows the effects of PKC inhibitors, such as IDE196 (compound A), and cMET inhibitors (crizotinib) on the survival rate of MM28 metastatic uveal melanoma cells in the presence of HGF. [Figure 7A-2] Same as above. [Figure 7A-3] Same as above. [Figure 7A-4] Same as above. [Figure 7B-1] This chart shows the effects of PKC inhibitors, such as IDE196 (compound A), and cMET inhibitors (capmatinib) on the survival rate of MM28 metastatic uveal melanoma cells in the presence of HGF. [Figure 7B-2] Same as above. [Figure 7B-3] Same as above. [Figure 7B-4] Same as above. [Figure 7C] This provides Western blots demonstrating the pharmacodynamic effects of drug therapies on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways. [Figure 8A] This report provides bar graphs showing cMET expression / MET activation (MET signature) across patients with metastatic uveal melanoma enrolled in the IDE196 (compound A) monotherapy trial (NCT02601378), grouped by response type. The patient group includes those with a clinical outcome determined to be progressive disease (metastatic uveal melanoma), those determined to have stable metastatic uveal melanoma for less than 6 months, those determined to have stable metastatic uveal melanoma for more than 6 months, and those determined to have a partial response to treatment (i.e., a reduction of 30% or more in metastatic uveal melanoma tumor size according to RESIST criteria). [Figure 8B] Same as above. [Modes for carrying out the invention]

[0018] definition This definition section is an integral part of the detailed description of the invention. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art.

[0019] The use of the articles “a,” “an,” and “the” in both this specification and the claims should be interpreted as encompassing both singular and plural forms unless otherwise indicated herein or unless the context clearly contradicts this. The terms “comprising,” “having,” “including,” and “containing” should be interpreted as open terms (e.g., “including but not limited to”) unless otherwise indicated. The term “or” is used to refer to a non-exclusive “or” unless otherwise indicated or unless the context clearly contradicts this. The statement “at least one of A and B” or “at least one of A or B” is equivalent to “A or B, or A and B.” Furthermore, it should be understood that any expressions or terms used herein, unless otherwise defined, are for illustrative purposes only and not for limitation. Any use of section headings is intended to aid in the reading of this document and should not be interpreted as limiting, and the information related to a section heading may occur inside or outside that particular section.

[0020] In the context of this application, the term “may” means “permitted” or “possible” and is a synonym for the term “can.” As used herein, the term “may” does not imply possibility or opportunity.

[0021] In the methods described herein, the actions may be performed in any order without departing from the principles of the invention, unless a temporal or operational order is explicitly stated. Furthermore, unless the language of an explicit claim states that the specified actions are performed separately, the specified actions may be performed simultaneously. For example, the action of performing claimed X and the action of performing claimed Y may be performed simultaneously within a single action, or sequentially in any order, and the resulting process falls within the literal scope of the claimed process.

[0022] The terms “approximately,” “approximately,” or “approximately,” when used in relation to a number, mean that a set or range of values ​​is included. As used herein, “approximately X” includes a range of values ​​that are ±25%, ±20%, ±10%, ±5%, ±2%, ±1%, ±0.5%, ±0.2%, or ±0.1% of X, where X is a number. In one embodiment, the term “approximately” refers to a range of values ​​that is 25% greater than or less than a given value. In another embodiment, the term “approximately” refers to a range of values ​​that is 20% greater than or less than a given value. In yet another embodiment, the term “approximately” refers to a range of values ​​that is 10% greater than or less than a given value. Preferably, the term “approximately” refers to a range of values ​​that is 5% greater than or less than a given value. Unless otherwise indicated, all numbers used herein and in the claims to represent quantities of ingredients, reaction conditions, etc., should be understood in all cases to be modified by the term “approximately.” Therefore, unless otherwise indicated, the numerical parameters described herein and in the appended claims are approximations that may be modified by the desired characteristics to be obtained by this disclosure.

[0023] The term "alkyl" refers to alkyl groups that do not contain heteroatoms. Therefore, this term includes linear alkyl groups (e.g., methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, etc.). This term also includes, but is not limited to, branched isomers of linear alkyl groups, including: -CH(CH3)2, -CH(CH3)(CH2CH3), -CH(CH2CH3)2, -C(CH3)3, -C(CH2CH3)3, -CH2CH(CH3)2, -CH2CH(CH3)(CH2CH3), -CH2CH(CH2CH3)2, -CH2C(CH3)3, -CH2C(CH2CH3)3, -CH (CH3)-CH(CH3)(CH2CH3), -CH2CH2-CH(CH3)2, -CH2CH2CH(CH3)(CH2CH3), -CH2CH2CH(CH2CH3)2, -CH2CH2C(CH3)3, -CH2CH2C(CH2CH3)3, -CH(CH3)CH2-CH(CH3)2, -CH(CH3)CH(CH3)CH(CH3)2, -CH(CH2CH3)CH(CH3)CH(CH3)(CH2CH3), etc. This phrase also includes cyclic alkyl groups (e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl), as well as such rings substituted with linear and branched alkyl groups as defined above. Thus, "C 1~12 The term "alkyl group" includes primary alkyl groups, secondary alkyl groups, and tertiary alkyl groups. Alkyl groups include linear and branched alkyl groups having 1 to 12 carbon atoms, as well as cyclic alkyl groups, where cyclic alkyl groups have at least 3 carbon atoms.

[0024] When used herein, "C 1~6 "Alkyl" includes linear or branched alkyl groups, both substituted and unsubstituted, having 1 to 6 carbon atoms. Typical C 1~6 Examples of alkyl groups include C 1~3 Alkyl, C 2~3Examples include alkyl, methyl, ethyl, propyl, isopropyl, n-butyl, TERT-butyl, neopentyl, trifluoromethyl, and pentafluoroethyl. Unless otherwise specified, C 1~6 Alkyl groups may be substituted with, for example, halo groups, hydroxyl groups, amino groups, nitro groups, and / or cyano groups. Typical C 1~3 Haloalkyl and C 1~3 Examples of hydroxyalkyl compounds include chloromethyl, trichloromethyl, trifluoromethyl, fluoromethyl, fluoroethyl, chloroethyl, hydroxymethyl, and hydroxyethyl. Other suitable substitutions include C. 1~3 Unless otherwise specified, the alkyl portion includes, for example, arylalkyl, aminoalkyl, aminoaralkyl, carbonylaminoalkyl, alkylcarbonylaminoalkyl, arylcarbonylaminoalkyl, arylalkylcarbonylaminoalkyl, aminoalkoxyalkyl, and arylaminoalkyl.

[0025] Where used herein, "C 1~6 "Axoxy" is a radical RO- (where R is C) 1~6 C refers to an alkyl group. 1~6 Typical examples of alkoxy groups include C 1~3 Examples include alkoxy, methoxy, ethoxy, t-butoxy, and trifluoromethoxy compounds.

[0026] As used herein, the terms "halogen" or "halo" refer to chloro, bromo, fluoro, and iodo groups. "Haloalkyl" refers to a C atom substituted with one or more halogen atoms. 1~3 This refers to alkyl radicals. The term "haloalkoxy" refers to a C atom substituted with one or more halogen atoms. 1~3 This refers to alkoxy radicals.

[0027] "Hydroxy" or "hydroxyl" refers to the group -OH. In this specification, "amino" refers to the group -NH2. The term "aryl" herein refers to a fully unsaturated conjugated carbocyclic ring system of 6 to 10 members (e.g., phenyl or naphthyl). The aryl ring(s) may be unsubstituted or contain one or more halos, C 2~3 Alkinyl, C 2~3 Alkenil, CN, C 1~3 Alkyl, C 1~3 Alkoxy, C 1~3 Haloalkyl, C 1~3 Haloalkoxy, C 3~7 Cycloalkyl, CONH2, CONHC 1~3 Alkyl, CONHC 6~10 Aryl, SO2NH2, SO2NHC 1~3 Alkyl, SO2NHC 6~10 The heterocyclyl ring may be substituted with an aryl, heteroaryl group, and / or a 4-7 membered heterocyclyl ring having 1-3 heteroatoms selected from N, O, and S, wherein the heterocyclyl ring is optionally H, 2 H, Haro, CN, C 1~3 Alkyl, C 1~3 Alkoxy, C 1~3 Haloalkyl and C 1~3 They are substituted with one or two substituents independently selected from the group consisting of haloalkoxyalkyl groups.

[0028] The terms "carbocycloalkyl, carbon bicyclic, and carbobicyryl" refer to a compound where all ring atoms are carbon atoms. 3~16 This refers to cycloalkyl and bicycloalkyl groups. When used in relation to cycloalkyl substituents, the term “polycyclic” refers herein to condensed and uncondensed cyclic alkyl structures. The term “bicyclic or carbobicyryl” refers to a saturated or partially unsaturated carbocyclic ring condensed to another carbocyclic ring, aryl ring, heterocyclic ring, or heteroaryl ring. Cycloalkyl groups are either unsubstituted or substituted.

[0029] The term “C 1~3"Alkylamino" as used herein refers to the group -NRR', where R and R' are each independently hydrogen or C 1~3 alkyl, provided that at least one of R and R' is C 1~3 alkyl.

[0030] The term "arylamino" as used herein refers to the group -NRR', where R is C 6~10 aryl containing phenyl and R' is hydrogen, C 1~3 alkyl, or C 6~10 aryl containing phenyl.

[0031] The terms "heterocycloalkyl," "heterocyclyl," "heterocycle," and "heterocyclic" are synonymous and refer herein to carbocyclic and heterocyclic rings that may be partially unsaturated or fully saturated and that may have 1 to 5, more typically 1 to 4, heteroatoms in the ring structure. Suitable heteroatoms for such rings are nitrogen, oxygen, and sulfur. Representative heterocycloalkyl / heterocyclyl moieties include, for example, morpholino, piperazinyl, piperidinyl, 1,2-oxazinane, 2-oxopiperazinyl, 2-oxopiperidinyl, N-methylpiperazinyl, and morpholinyl, each optionally substituted.

[0032] The term "heterobicyclic" refers to a bicyclic group in which a heterocyclic ring is fused to a benzene ring or another 5- or 6-membered heterocyclic ring. The heterocyclic moiety is unsubstituted or substituted with hydroxy, halo, oxo (C=O), alkylimino (RN=, where R is C 1~3 alkyl or C 1~3 alkoxy group), amino, C 1~3 alkylamino, C 1~3 dialkylamino, acylaminoalkyl, C 1~3 alkoxy, C 1~3 alkyl, cycloalkyl, or C 1~3It may be mono- or di-substituted with various substituents independently selected from haloalkyl. The heterocyclic group (heterocyclyl) may be attached at various positions as will be apparent to those skilled in organic chemistry and pharmaceutical chemistry in conjunction with this specification.

[0033] The term "heteroaryl" refers to a 5- to 10-member unsaturated conjugated heterocyclic ring system including a fused ring system having 1 to 4 heteroatoms independently selected from the group consisting of O, N, and S. The heteroaryl group may optionally be substituted with one or two substituents. A subset of the "heteroaryl" system is an aromatic C having 1 to 4 heteroatoms as ring atoms in the aromatic ring and the remainder of the ring atoms being carbon atoms. 6~10 It is a heteroaryl group. Exemplary substituents include halo, CN, C 1~3 alkyl, C 1~3 alkoxy, C 1~3 haloalkyl, C 1~3 haloalkoxy, C 3~7 cycloalkyl, and 4- to 7-member heterocyclyl having one or two heteroatoms selected from N, O, and S, but are not limited thereto, and the heterocyclyl may optionally be halo, CN, C 1~3 alkyl, C 1~3 alkoxy, C 1~3 haloalkyl, and C 1~3They are substituted with 1 to 3 substituents independently selected from the group consisting of haloalkoxys. Typical heteroaryl groups include, for example, the following: Representative heteroaryl compounds include, for example, imidazolyl, pyridinyl (also called aspyridyl), pyrazinyl, azetidinyl, thiazolyl, triazolyl, benzimidazolyl, benzothiazolyl, thiazolyl, thiazolidinyl, isothiazolyl, isothiazolidinyl, indolyl, quinolinyl, isoquinolinyl, azetidinyl, N-methylazetidinyl, pyrimidinyl, pyridadinyl, oxazolyl, oxazolidinyl, isoxazolyl, isoazolidinyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, furyl, thienyl, triazolyl, benzothienyldiazapinyl, pyril, pyrrolinil, pyrrolidinyl, pyrazolyl, pyrazolinil, pyrazolidinyl, imidazoyl, imidazolinyl, imidazolidinyl, and benzoxazolyl. Heteroaryl compounds are either unsubstituted or H, 2 H, Halo, C 2~3 Alkinyl, C 2~3 Alkenil, CN, C 1~3 Alkyl, C 1~3 Alkoxy, C 1~3 Haloalkyl, C 1~3 Haloalkoxy, C 3~7 Cycloalkyl, CONH2, CONHC 1~3 Alkyl, CONHC 6~10 Aryl, SO2NH2, SO2NHC 1~3 Alkyl, SO2NHC 6~10 The molecule is substituted with 1 to 3 substituents independently selected from the group consisting of aryls and 4 to 7-membered heterocyclines having 1 to 3 heteroatoms selected from N, O, and S, wherein the heterocycline is optionally substituted with H, 2 H, Haro, CN, C 1~3 Alkyl, C 1~3 Alkoxy, C 1~3 Heteroalkyl, and C 1~3 They are substituted with one or two substituents independently selected from the group consisting of heteroalkoxy compounds.

[0034] " 2 The term "H" refers to the heavy isotope of hydrogen, also known as deuterium (D). It should be understood that the above definition of an organic group is not intended to include unacceptable substitution patterns (e.g., a methyl group substituted with five fluoro groups, or a halogen atom substituted with another halogen atom).

[0035] The term "ligand" refers to a substance that can form covalent, complex, electrostatic, hydrophobic, hydrophilic, lipophilic, polar, steric, and / or similar molecular interactions with another substance. Ligands may be small molecules, charged organic or inorganic moieties, peptides, oligopeptides, DNA or RNA fragments, immunoproteins, antibody fragments, polyclonal antibodies, monoclonal antibodies, humanized monoclonal antibodies, and / or similar chemical substances. Ligands may, though not necessarily, have signaling groups (e.g., radioactive groups, fluorescent groups, or phosphorescent groups) so that when they form molecular interactions with another substance, the combination can be detected. Ligands are preferably peptides, any type of antibody, or any type of antibody fragment. A subset of ligands may consist of two ligands, the first ligand which interacts with another substance, and the second ligand which has a signaling group and also interacts with the first ligand.

[0036] As used herein, the term "PKC" refers to protein kinase C. The PKC family of serine / threonine kinases consists of at least 10 isoforms and is critical to various cell differentiation processes through characteristic regulatory mechanisms. Depending on the context, the term PKC may refer to the entire family of isoforms or to a specific isoform. The PKC family includes the conventional (classical) isoforms alpha, beta-1, beta-2, and gamma; the novel isoforms delta, epsilon, eta, and theta; and the atypical isoforms zeta and iota. As used herein, the terms "protein kinase C inhibitor" or "PKC inhibitor" refer to a protein kinase C inhibitor that may be pan-subtype or selective for one or more PKC isozymes.

[0037] The term "inhibitor" refers to a regulatory molecule or compound that partially or completely blocks, reduces, interferes with, delays, inactivates, desensitizes, or downregulates the biological activity or expression of, for example, a protein, kinase, or other biological structure (e.g., PKC or cMET). For example, inhibitors can bind to enzymes reversibly or irreversibly, competitively, non-competitively, non-competitively, or in combination thereof.

[0038] The term "pharmaceutical composition" refers to an active agent formulated with one or more pharmaceutically acceptable carriers. Examples of pharmaceutical compositions include parenteral solutions, and tablets are used herein to refer to compounds, materials, compositions, and / or dosage forms suitable for use in contact with human tissue without excessive toxicity, irritation, allergic reactions, or other problems or complications commensurate with a reasonable benefit / risk ratio, within the bounds of sound medical judgment.

[0039] The term "pharmaceutically acceptable" is used herein to mean a compound, material, composition, excipient, and / or dosage form that is suitable for use in contact with human tissue without excessive toxicity, irritation, allergic reaction, or other problems or complications commensurate with a reasonable benefit / risk ratio, within the bounds of sound medical judgment.

[0040] The terms “to treat,” “to treat,” or “treatment,” as used herein, mean a method of alleviating, reducing, or improving a disease (e.g., a tumor), condition, or symptom; a method of preventing the onset of further symptoms; a method of improving the underlying cause of symptoms; a method of inhibiting a disease or condition; a method of stopping or reducing the onset or progression of a disease or condition; a method of alleviating a disease or condition; a method of causing a disease or condition to regress; a method of alleviating a condition caused by a disease or condition; or a method of stopping the symptoms of a disease or condition. In various embodiments, the treatments described herein can be used prophylactically to prevent a disease (e.g., metastatic disease), condition, or symptom.

[0041] The term “patient” refers to a person, in particular a person who is receiving, but not necessarily, medical care. For example, a patient may be a person who has cancer or has been diagnosed with cancer. Another example is a patient who has been diagnosed with a high risk of developing cancer, a tumor, or a metastatic disease. For example, a patient may have been previously diagnosed with and treated for uveal melanoma and is now diagnosed with a high risk of recurrence or progression of a metastatic disease.

[0042] The term “patient population” refers to a group of patients who share one or more common disease characteristics. For example, a patient population may correspond to patients with the same histological type of tumor. Another example is a patient population that may be a group of patients with the same histological type of tumor having mutations in GNAQ or GNA11. A patient population may be patients with uveal melanoma. A patient population may be patients with metastatic uveal melanoma. A patient population may be two or more patients, for example, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 50, 100, 500, 1000, 2000, 5000, 7000 or more patients. In various embodiments, a patient population may be defined to provide a representative group suitable for assessing the relative severity of the disease among patients. In some cases, a patient population may be further defined to include characteristics other than disease characteristics, such as demographic, phenotypic, or genotypic characteristics, for the purpose of providing a more accurate assessment and evaluation of patients. In some embodiments, the patient population may alternatively refer to the “healthy population,” or the general population that includes both healthy individuals and patients suffering from the disease.

[0043] The terms “administer” or “to administer” refer to bringing a patient into contact with a particular compound or composition in order to treat or prevent a disease or its symptoms. In various examples, administering includes bringing a patient’s tumor into contact with a particular compound or composition, or bringing the affected tissue in which the tumor is located into contact with a particular compound or composition.

[0044] Where used herein, “disease” is intended to be synonymous with and interchangeable with the terms “disorder,” “syndrome,” and “condition” (as in a medical condition), all in that they reflect an abnormal condition of the body or a part of the body of a human or animal that impairs normal function, typically manifests by identifying signs and symptoms, and reduces the lifespan and / or quality of life of the human. “Proliferative disorders” are diseases characterized by malignant proliferation of cells. For example, a disease may be cancer in which protein kinase C or other substances play a role in the pathogenesis of cancer. For another example, a disease may be a tumor, such as a solid tumor, a metastatic tumor, or both. Examples of proliferative disorders in general include pancreatic cancer, gastric cancer, colorectal cancer, cervical cancer, lung cancer, bladder cancer, liver cancer, breast cancer, head and neck cancer, eye cancer, and brain cancer, or tumors thereof.

[0045] When used herein, "a patient in need of treatment" means a judgment made by a physician or other caregiver that the patient needs treatment or would benefit from treatment. "A patient diagnosed with a specific disease" means a judgment made by a physician or other caregiver that the patient has a disease. "A patient suffering from a specific disease" means a patient suffering from a disease that can be demonstrated based on the progression of the disease or a judgment made by a physician or other caregiver that the patient has a disease. Such a judgment may be made based on a variety of factors within the scope of the physician's or caregiver's expertise. In various embodiments, the treatments described herein are offered to patients who need treatment for, have been diagnosed with, or suffer from one of the diseases described herein.

[0046] As used herein, the term “tumor” includes solid tumors, humoral tumors, or combinations thereof. For example, a tumor may be a primary uveal melanoma, a metastatic uveal melanoma, a tumor with one or more mutations in GNAQ or GNA11, a tumor other than uveal melanoma, or a metastatic tumor in the liver. Further examples include a metastatic uveal melanoma in the liver, or a primary uveal melanoma with one or more mutations in GNAQ or GNA11. In various embodiments, a tumor is a malignant (cancerous) solid tumor. In various embodiments, a tumor is a humoral tumor.

[0047] As used herein, “mutation” may refer to one or more changes in a polynucleotide sequence. Mutations may occur in tumor cells, or they may occur in cells prior to the appearance or diagnosis of a tumor. Mutations may be acquired or germline (hereditary) mutations. Mutations may represent deviations from the healthy function of a polynucleotide sequence and its derived protein. Mutations may be nucleotide substitutions, insertions, or deletions, such as single nucleotide substitutions. For example, GNAQ and GNA11 mutations are typically activating mutations that lead to constitutive activation of the α subunit. While not theoretically bound, constitutive activity is thought to result from a lack of GTP-hydrolase activity in the mutant GNAQ or GNA11 protein. Activating mutations may also refer to mutations that result in the loss or reduction of GTP hydrolysis activity of the Gα subunit. Mutations in GNAQ and GNA11 may include substitutions of arginine R183 or glutamine Q209 in the codon, or other mutations. In some embodiments, mutations in GNAQ and / or GNA11 may be selected from the group consisting of Q209P, Q209L, Q209H, Q209K, Q209Y, Q209R, Q209H, R183Q, and R183. For example, GNAQ Q209 may be mutated to either P or L and either R or H; GNAQ R183 may be mutated to Q; GNA11 Q209 may be mutated to L and either P or K; and GNAQ R183 may be mutated to either C or H. Gene mutations may include any of the following: insertion mutations, substitution mutations, deletion mutations, gain-of-function mutations, loss-of-function mutations, and non-synonymous mutations. Gain-of-function mutations result in altered gene products with new molecular function or new gene expression patterns. In contrast, loss-of-function mutations produce altered gene products that lack the molecular function of the equivalent wild-type gene.

[0048] As used herein, the term “GNAQ” refers to the alpha-Q gene, a guanine nucleotide-binding protein encoding the Gq alpha subunit (Gαq), and the term “GNA11” refers to the alpha-11 gene, a guanine nucleotide-binding protein encoding the G11 alpha subunit (Gα11). The term “GNAQ / 11” refers to GNAQ and / or GNA11. This term encompasses the nucleic acids and polymorphic variants, alleles, mutants, and fragments of GNAQ and GNA11. GNAQ and GNA11 sequences are well known in the art. An example of a human GNAQ sequence is available in the NCBI nucleotide database under the reference sequence NM_002072 (nucleotide sequence) and accession number NP_002063.2 (polypeptide sequence). Human GNAQ is localized to chromosome 9q21. Examples of human GNA11 sequences are available in the NCBI nucleotide database under the reference sequence NM_002067 (nucleotide sequence) and accession number NP_002058.2 (polypeptide sequence). Human GNA11 is localized to chromosomal region 19p13.3.

[0049] As used herein, the term “mutation load” refers to the level, e.g., number, of changes (e.g., one or more changes, e.g., one or more somatic changes) per pre-selected unit (e.g., per megabase) in a given set of genes or all analyzed genes (e.g., the coding region of a given set of genes). Mutation load can be measured, for example, on the whole genome or exome, or on a subset of the genome or exome. In certain embodiments, the whole genome or exome mutation load can be determined by extrapolating a mutation load measured on a subset of the genome or exome. The terms “mutation load,” “mutational load,” “mutation burden,” and “mutational burden” are used interchangeably herein. In relation to tumors, mutation load is also referred herein to as “tumor mutational burden,” “tumor mutation burden,” or “TMB.”

[0050] BAP1 refers to the BRCA1-related protein-1 gene (ubiquitin carboxy-terminal hydrolase; BAP1). The nucleic acid and amino acid sequence of BAP1 are publicly known and publicly available (Genbank NM_004656.2, Genbank NP_004647.1). BAP1 is functionally involved in the DNA damage response, as well as in the regulation of apoptosis, senescence, and the cell cycle. Deletion and inactivation mutations in BAP1 have previously been associated with breast and lung tumors, and, consistent with BAP1's role as a tumor suppressor, restoration of BAP1 function has been shown to suppress cell growth and tumorigenicity in BAP1-mutated lung cancer cell lines.

[0051] SF3B1 refers to the gene encoding splicing factor 3b subunit 1. The nucleic acid and amino acid sequence of SF3B1 are publicly known and publicly available (NM_012433.3, Genbank NP_036565.2). Subunit 1 of the splicing factor 3b protein complex plays several important roles in the cellular splicing mechanism. Mutations in SF3B1 affect the cell's ability to convert pre-mRNA containing intronic sequences into mature mRNA.

[0052] E1F1AX refers to the gene encoding protein X-binding eukaryotic translation initiation factor 1A, which plays a role in protein synthesis. The nucleic acid and amino acid sequence of E1F1AX are publicly known and publicly available (NM_001412.4, Genbank NP_001403.1). E1F1AX is commonly mutated in uveal melanoma.

[0053] TERT refers to either the gene encoding the enzyme telomerase reverse transcriptase (TERT), or the enzyme (i.e., the protein) itself. TERT refers to the nucleoprotein, or the portion of the enzyme, telomerase. The TERT gene is also known as the "Ever Shorter Telomeres" or "EST" gene. Mutations in the promoter region of TERT have been associated with cancers including, but not limited to, thyroid cancer, bladder cancer, and glioblastoma. The nucleic acid and amino acid sequences of TERT are publicly known and publicly available (NM_198253.2, Genbank NP_937983.2).

[0054] NRAS, or "neuroblastoma RAS virus oncogene homolog," refers to a small GTPase Ras family protein encoded on chromosome 1. The nucleic acid and amino acid sequences of NRAS are publicly known and publicly available (NM_002524, Genbank NP_002515).

[0055] BRAF, or "v-Raf mouse sarcoma virus oncogene homolog B," refers to the Raf kinase family serine / threonine-specific protein kinases that interact with AKT1, CRaf, HRAS, and YWHAB. The BRAF sequence is well known in the art for many species, e.g., human BRAF (NM_004333, Genbank NP_004324). NRAS and / or BRAF sequence mutations can be point mutations. In some embodiments, an NRAS point mutation can result in one of the following amino acid residue changes: G12D, G12S, G13A, G13C, G13D, G12R, G13V, Q61H1, Q61K, Q61L, Q61R1, and Q61R2. In some embodiments, a BRAF point mutation may be a point mutation resulting in one of the following amino acid residue changes: V600D TG / AT, V600E T / A, V600E TG / AA, and V600K GT / AA.

[0056] Detailed explanation Embodiments of the present invention provide combination therapies useful for treating, among other things, metastatic uveal melanoma, tumors having GNAQ mutations or GNA11 mutations, and other proliferative disorders (hereinafter referred to as target tumors). GNAQ / 11 tumors may be primary GNAQ / 11 tumors or metastatic GNAQ / 11 tumors. Examples of other proliferative disorders include diseases mediated by tissue abnormalities in tissues, which have one or more mutations in GNAQ or GNA11 that activate the PKC signaling pathway in tissues. Combination therapies may be provided as pharmaceuticals, methods of treatment, or kits and may involve the use of protein kinase C inhibitors and cMET inhibitors. Embodiments of the present invention also provide methods and kits useful for selecting patients or directing the course of treatment. For example, embodiments of the present invention provide a method for identifying whether or not to administer a combination of protein kinase C inhibitors and cMET inhibitors to a patient.

[0057] This invention is at least in part based on the finding that the antiproliferative activity of the PKC inhibitors of this embodiment, acting on PKC isoforms present in target tumors, is regulated by the presence of elevated cMET. The functions and downstream signaling of various PKC isoforms are complex and can vary depending on the cellular environment. The PKC inhibitors of this embodiment are highly potent against both novel PKC isoforms delta (δ), epsilon (ε), eta (η), and theta (θ), as well as classical PKC isoforms alpha (α), beta-1 (βI), beta-2 (βII), and gamma (γ). Within these classes of PKC isoforms, the PKC inhibitors exhibit greater activity against novel PKC isoforms than against classical PKC isoforms. The PKC inhibitors of this embodiment are selective against PKC isoforms compared to off-target kinases and other off-target phosphotransferases, and exhibit favorable therapeutic index and PKC activity compared to other known pan-PKC inhibitors and / or PKC inhibitors acting on classical PKC isoforms.

[0058] The antiproliferative activity of the PKC inhibitor in this embodiment was found to be antagonized in target tumors exhibiting elevated cMET levels. Furthermore, the elevation of cMET in target tumors can be altered, for example, when exposed to an equal amount of exogenous hepatocyte growth factor (HGF), an endogenous ligand for the extracellular cMET receptor. In particular, the antiproliferative effect of IDE196 on uveal melanoma cell lines 92.1, MM28, and MEL-202 is antagonized to different degrees, even when each of these cell lines is experimentally stimulated with the same concentration of HGF. This cMET antagonism was observed most strongly in the MEL-202 and MM28 cell lines and weaker in the 92.1 cell line. This correlates with the elevated cMET expression levels in the MEL-202 and MM28 cell lines compared to the 92.1 cell line.

[0059] Furthermore, data from record-holding metastatic uveal melanoma tissue from human clinical trials evaluating IDE196 as monotherapy demonstrate that cMET expression and MET activation can also be altered in human tumors, as indicated by the MET signature, and that elevated cMET expression and MET activation generally correlate with a poorer clinical response. These data are consistent with observations from in vitro cell experiments that the presence of elevated cMET antagonizes the antiproliferative activity of IDE196. Referring to Example 5 and Figures 8A and 8B, in this study, some patients showed little to no improvement in proliferative target tumors when administered IDE196 as monotherapy, while others showed at least a partial response. In all patients, the metastatic uveal melanoma site of the tumor was in the liver. Patients with little to no remission response to IDE196 showed higher tumor cMET expression or MET activation, while patients who were more responsive to IDE196 showed lower levels of tumor cMET expression or MET activation. These in vitro cell data and in vivo human clinical data demonstrate that the level of cMET expression or MET activation, rather than the presence of HGF, determines the antagonistic effect of IDE196 on the antiproliferative effect of target tumors.

[0060] These findings indicate that the combination of PKC inhibitors and cMET inhibitors in this embodiment provides advantageous treatment for patients with metastatic uveal melanoma and patients with tumors containing GNAQ or GNA11 mutations, including mutations that activate the PKC signaling pathway. It is known that cMET inhibitors control cMET expression or cMET activation. Importantly, however, cMET inhibitors alone do not affect uveal melanoma cell viability in any of the cell lines tested. Notably, the magnitude of the synergistic effect of cMET inhibitor / PKC inhibitor combinations correlates with the cMET expression level of target tumor cells. The greatest synergy was observed in MEL-202 and MM28 cell lines treated with a combination of crizotinib or capmatinib (both cMET inhibitors) and IDE196, while the combination of crizotinib or capmatinib and IDE196 showed a relative lack of synergy when administered to the 92.1 cell line.

[0061] The use of such combinations may further include methods for patient selection and criteria for determining whether to administer PKC inhibitors as combination therapy with cMET inhibitors, or whether to administer PKC inhibitors without cMET inhibitors, for example, as monotherapy or in combination with other therapeutic agents. Thus, in various embodiments, the present invention solves problems related to the treatment of difficult-to-treat proliferative disorders such as metastatic uveal melanoma and / or tumors having GNAQ mutations or GNA11 mutations.

[0062] Generally speaking, each additional therapeutic agent administered to a patient carries a risk of side effects and may reduce the therapeutic index of the prescribed treatment. However, it is preferable that the treatment improves therapeutic efficacy while reducing risks. In various embodiments, the present invention solves the problem of identifying which patients may and may not benefit from the combination, and thus provides the further advantage of providing a treatment with an improved therapeutic index.

[0063] Reference now will be made in detail to specific embodiments of the disclosed subject matter. The disclosed subject matter is described in conjunction with the recited claims, but it will be understood that the exemplified subject matter is not intended to limit the disclosed subject matter to the recited claims.

[0064] Combination therapy The present invention provides, inter alia, a combination therapy useful for treating metastatic uveal melanoma, tumors having a GNAQ mutation or a GNA11 mutation, preferably a GNAQ mutation or a GNA11 mutation that activates the PKC signaling pathway (a "GNAQ / 11 tumor"), and other proliferative diseases such as diseases mediated by tissue abnormalities in a tissue having one or more mutations in GNAQ or GNA11 that activate the PKC signaling pathway in such a tissue, the combination therapy comprising a protein kinase C inhibitor and a cMET inhibitor. The GNAQ / 11 tumor can be a primary GNAQ / 11 tumor or a metastatic GNAQ / 11 tumor. The combination therapy can be provided by a pharmaceutical, a pharmaceutical composition, a method of treatment, or a kit, each of which can comprise a protein kinase C inhibitor and a cMET inhibitor. For example, the combination therapy can include treating a patient having metastatic uveal melanoma or having a GNAQ / 11 tumor by co-administering to the patient a cMET inhibitor and a protein kinase C inhibitor either simultaneously or sequentially.

[0065] In various embodiments, the protein kinase C inhibitor may have a structure represented by Formula II:

[0066]

Chemical formula

[0067] or a pharmaceutically acceptable salt thereof, wherein X is N or CR; R, R 2 、R 3 、and R 4 are each independently H, 2H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The alkoxy may be optionally substituted with one, two, three, or more halogens.

[0068] In various embodiments, the compound of formula II may have one or more of the following, or each of them: X is N; R 2 , R 3 , and R 4 Each is independently H or halo; R 5 is H or CH3; or R 6 and R 7 These are H, Halo, and C, respectively, and are independent of each other. 1~3 Haloalkyl, C 1~3 Haloalkoxy, C 3~7 Selected from cycloalkyl, morpholino, piperidinyl, and piperazinyl. For example, R 2 , R 3 , and R 4 Each of these can be H. As another example, R 6 and R 7 C 1~3 Haloalkyl, C 1~3 Haloalkoxy, C 3~7 These are cycloalkyl, morpholino, piperidinyl, and piperazinyl, and R 6 and R 7 One of them is H. In various further forms, R 6 and R 7 One of them is trifluoromethyl or trifluoromethoxy, R 6 and R 7 One of them is H. In a further embodiment, R 5a , R 5b , R 5c , and R 5d These are H.

[0069] For example, protein kinase C inhibitors have the following structure:

[0070] [ka]

[0071] It may also have a pharmaceutically acceptable salt thereof. As further examples, protein kinase C inhibitors include 3-amino-N-(3-(4-aminopiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-aminopiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(methoxymethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-( (Lifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(hydroxymethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(hydroxymethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methyl Lupiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(4-methoxy-3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide , 3-amino-N-(3-(4-amino-4-ethylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-chloropyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-fluoropyridine-2-yl)pyrazine-2-carboxamide,3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-fluoro-4-methoxypyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(methoxymethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-fluoropyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-cyanopyridine-2-yl)pyrazine-2-carboxamide Boxamide, 3-amino-N-(3-(4-amino-4-ethylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-cyano-4-methoxypyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(methoxymethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl Xy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(2-hydroxyethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(2-hydroxyethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-fluoropyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl) (L)pyridine-2-yl)-6-(4-cyano-3-fluoropyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-(2-methoxyethyl)piperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, (+)3-amino-N-(3-((cis)-4-amino-3-fluoropiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide,3-amino-N-(3-(4-aminopiperidine-1-yl)pyridine-2-yl)-6-(3-fluoropyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-((3S,4R)-4-amino-3-fluoropiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-ethylpiperidine-1-yl)pyridine-2-yl)-6-(3-fluoropyridine-2-yl)pyrazine-2 -Carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(4-cyano-3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(4-ethoxy-3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)- 6-(4-chloro-3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-cyano-4-methoxypyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-aminopiperidine-1-yl)-6-methylpyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-aminopiperidine) (Zin-1-yl)-6-methylpyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-3-methoxypiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethoxy)pyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-ethylpiperidine-1-yl)pyridine-2-yl)-6-(3-fluoro-4-methylpyridine-2-yl)pyrazine-2-carboxamide,3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(4-ethoxy-3-fluoropyridine-2-yl)pyrazine-2-carboxamide, 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(4-(hydroxymethyl)-3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, and 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(4-(methoxymethyl)-3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, or pharmaceutically acceptable salts thereof.

[0072] In various preferred embodiments, the protein kinase C inhibitor may be 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide (compound A), or a pharmaceutically acceptable salt thereof. The protein kinase C inhibitor has the following structure:

[0073] [ka]

[0074] It may also have a pharmaceutically acceptable salt thereof. The protein kinase C inhibitors described herein represent only a subset of available protein kinase C inhibitors. The combination therapies described herein involve the use of protein kinase C inhibitors that have significant potency against PKC isoforms and selectivity for the PKC family compared to other kinases. Many existing kinase inhibitors are not particularly selective for their target kinomes, but in some cases, the same cross-reactivity with other non-PKC kinases can be the source of biological benefits. For example, sotrastaurin is a PKC inhibitor, but it also inhibits GSK-3 beta, which is involved in a wide variety of diseases and may play a role in the survival of malignant cells. Sotrastaurin is also a panselective PKC inhibitor, exhibiting high potency across the entire family of PKC isoforms and showing greater inhibitory activity against classical PKC isoforms than against novel PKC isoforms. While it has become recognized that different inhibitory differences for specific PKC isoforms offer different biological effects, the diverse properties of each isoform are not fully understood in all biological systems. As an example of an isoform-specific PKC inhibitor, enzastaurin is selective for the classical PKC isoform, i.e., the beta isoform, and has been studied as a cancer treatment based on this selectivity. In contrast, the PKC inhibitors of this embodiment are highly potent against novel and classical PKC isoforms, exhibiting greater activity against the novel isoform compared to their activity against the classical isoform. Further protein kinase C inhibitors are described in U.S. Patent No. 9,845,309, which is incorporated herein by reference in its entirety.

[0075] While not intended to limit to any particular theory of function, various carcinogenic conditions arising from mutations in metastatic uveal melanoma, or in GNAQ or GNA11, are related to intracellular DAG and / or Ca +This can lead to various changes. For example, activated GNAQ or GNA11 can further activate PLC-B, releasing various PKC signaling molecules by hydrolyzing the plasma membrane. The PKC family plays a role in regulating the activation of various other enzymatic pathways, including the MAPK pathway, which has an activation cascade mediated by RAS, RAF, MEK, and then ERK. PKC inhibition can lead to decreased ERK phosphorylation by reducing downstream activation of MAPK kinases. The antiproliferative activity of the PKC inhibitors described herein may result from this reduced inhibition of MAPK signaling and ERK activity. PKC activation also phosphorylates MARCKS, a more proximal pharmacodynamic biomarker for PKC activation.

[0076] The various PKC inhibitors described herein are highly selective for inhibiting PKC family proteins without significant cross-reactivity with other non-PKC kinases. In various cases, PKC inhibitors are more active against novel PKC isoforms than against classical PKC isoforms. While the various selective PKC inhibitors described herein are not thought to act on the cMET pathway, it has been found that hepatocyte growth factor (an endogenous ligand for cMET) can induce a high degree of antagonism against the PKC compounds described herein in certain tumor cells. This antagonism has been found to be a result of downstream signaling competition arising from elevated cMET concentrations and the use of the PKC inhibitors described herein. The nature of this downstream competition appears to be complex and does not simply correlate with HGF levels. This complexity may be partly due to the differing actions of various PKC isoforms on interquinome signaling and cMET transport. For example, see Kang, Protein Kinase C (PKC) Isozymes and Cancer, New Journal of Science, vol. 2014; Garg et al., Protein Kinase C And Cancer: What We Know And What We Do Not, Oncogene, vol. 33, 2014, 5225-5237; and Kermorgant et al., PKC Controls HGF-Dependent C-Met Traffic, Signaling And Cell Migration, EMBO J. 2004 Sep 29;23(19):3721-3734, each of which is incorporated herein by reference in its entirety.

[0077] The combination therapies described herein may involve the use of a protein kinase C inhibitor described herein that is highly active against one or more novel PKC isoforms, e.g., delta, epsilon, eta, or theta. In various embodiments, the protein kinase C inhibitor is effective against one or more of the novel PKC isoforms delta, epsilon, eta, or theta, or each of them, with an IC of approximately 5, 10, 15, 20, 25, 30, 35, 40, 45, 40, 50, 60, 70, 80, 90, or 100 nM or less. 50 It may have.

[0078] In various embodiments, the protein kinase C inhibitors of this embodiment are more active against one or more novel PKC isoforms, delta, epsilon, eta, or theta, compared to other PKC isoforms, for example, non-novel PKC isoforms alpha, beta 1, beta 2, gamma, zeta, and iota. In particular, the protein kinase C inhibitors of this embodiment have greater inhibitory activity against each of the protein kinase C isoforms delta, epsilon, eta, and theta than against classical protein kinase C isoforms alpha, beta 1, beta 2, or gamma. As an example, the protein kinase C inhibitors of this embodiment may exhibit at least 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, or 100-fold lower inhibitory activity against protein kinase C isoforms alpha, beta 1, beta 2, or gamma than against protein kinase C isoforms delta, epsilon, eta, or theta. In a further example, the protein kinase C inhibitors of this embodiment exhibit at least 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, or 100-fold lower inhibitory activity against protein kinase C isoform beta-1 than against protein kinase C isoform theta. In yet another example, the protein kinase C inhibitors of this embodiment may have an IC50 of about 5, 10, 15, 20, 25, 30, 35, 40, 45, 40, or 50 nM or less against one or more novel PKC isoforms delta, epsilon, eta, or theta, and an IC50 of about 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, or 100 nM or more against one or more classical PKC isoforms alpha, beta-1, beta-2, or gamma.

[0079] In a further aspect, the protein kinase C inhibitor of the present embodiment is selective for all PKC isoforms as compared to other kinases outside the protein kinase C kinome. For example, the protein kinase C inhibitor of the present embodiment may have an inhibitory activity that is at least 100-fold, 200-fold, 300-fold, 400-fold, 500-fold, 1000-fold, 2000-fold, 2500-fold, or 5000-fold lower against non-PKC kinases than against one or more or each of the novel or classical protein kinase C isoforms. As another example, the PKC inhibitor of the present embodiment may have an inhibitory activity that is at least 100-fold, 200-fold, 300-fold, 400-fold, 500-fold, 1000-fold, 2000-fold, 2500-fold, or 5000-fold lower against PIM2, GSK3β, or both than against one or more or each of the novel or classical protein kinase C isoforms. In a further aspect, the protein kinase C inhibitor of the present embodiment may have an IC50 of about 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, or 10,000 nM or more against PIM2, GSK3β, PKC zeta, PKZ iota, or a combination thereof.

[0080] The cMET inhibitor may be a small molecule inhibitor of cMET. The cMET inhibitor may be an inhibitor of intracellular ATP-cMET ligand-receptor binding. Examples of cMET inhibitors include crizotinib, capmatinib, cabozantinib, tivantinib, or a combination thereof.

[0081] In various aspects, the cMET inhibitor may also have activity as an ALK inhibitor, a RAS1 inhibitor, or both. For example, the cMET inhibitor may be crizotinib. In various further aspects, the cMET inhibitor has the following structure:

[0082]

Chemical formula

[0083] It may have one of the following or a pharmaceutically acceptable salt thereof. For example, cMET inhibitors are

[0084] [ka]

[0085] Alternatively, a pharmaceutically acceptable salt thereof may be used. As another example, cMET inhibitors are

[0086] [ka]

[0087] Alternatively, a pharmaceutically acceptable salt thereof may be used. Treatment method The present invention provides a method for treating patients with metastatic uveal melanoma or tumors with GNAQ mutations or GNA11 mutations (in particular, GNAQ mutations or GNA11 mutations that activate the PKC signaling pathway) ("GNAQ / 11 tumors") by providing a combination therapy comprising a cMET inhibitor and a PKC inhibitor, in each case. For example, the method may include co-administration of the protein kinase C inhibitor and cMET inhibitor of this embodiment (in particular, one or more of crizotinib, capmatinib, cabozantinib, or tivantinib), for example, simultaneously or sequentially. The PKC inhibitor may also be a compound according to Formula II above. The term "co-administration" is intended to encompass the administration of two or more compounds described herein, such as a protein kinase C inhibitor and a cMET inhibitor, to a single patient such that the therapeutic benefits of each of the two or more compounds overlap at least partially.

[0088] PKC inhibitors and cMET inhibitors can be administered to a patient simultaneously as a single composition, simultaneously as separate compositions, or sequentially as separate compositions. For example, PKC inhibitors and cMET inhibitors may be formulated as a single combined composition, such as a tablet, according to the prescription labeling rules for each compound, or according to pharmacokinetic and pharmacodynamic studies related to the simultaneous oral administration of the two compounds. Alternatively, a healthcare provider may instruct a patient to take the two separate compositions according to the labeling instructions associated with each of the protein kinase C inhibitors and cMET inhibitors. In one approach, a healthcare provider may instruct a patient to take the two separate compositions, a PKC inhibitor and a cMET inhibitor, simultaneously or nearly simultaneously, for example, during the same patient care visit, or within 0.5, 1, 2, 3, 4, 5, 10, 15, 20, 30, 60, 120, or 180 minutes, or otherwise, within a period indicated by the pharmacodynamic / pharmacokinetic profiles, such as the half-lives of the two compositions. The use of the pharmacodynamic / pharmacokinetic profiles of the two inhibitors allows healthcare providers to ensure that patients are exposed to detectable, preferably clinically significant, amounts of the active pharmaceutical ingredients of the PKC inhibitor and the cMET inhibitor. In some cases, the PKC inhibitor may be administered before the cMET inhibitor, and in other cases, the cMET inhibitor may be administered before the PKC inhibitor. Preferably, co-administration, whether simultaneous or sequential, is such that the combination of two or more compounds provides an additive, additive or more additive, or preferably synergistic, benefit to the patient.

[0089] The simultaneous or sequential administration of separate doses of PKC inhibitors and cMET inhibitors may, but preferably, follow the same route of administration. For example, co-administration may include the simultaneous or sequential administration of a protein kinase C inhibitor via oral or intravenous route and a cMET inhibitor via suppositories or intravenous or intravenous route. The attending physician can determine the most appropriate route of administration for separate doses of the two inhibitors. The choice may depend on several factors, including the patient's overall health and condition, the patient's swallowing ability, the patient's walking ability, and other physical and mental conditions of the patient, which may be assessed according to the attending physician's knowledge.

[0090] The administration of separate doses of PKC inhibitors and cMET inhibitors is performed when the compound administered first is its C in the patient. maxThe subsequent compound may be administered to the patient in a sequential procedure, having an in vivo concentration of at least or about 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, 0.01, 0.001, 0.0001, or 0.00001. The sequential administration of the subsequent compound may be performed before the Tmax of the previously administered compound, or at approximately the in vivo Tmax of the previously administered compound, or within approximately 5, 10, 20, 50, 100, or approximately 1000 times the in vivo Tmax of the previously administered compound. The sequential administration of the subsequent compound may be performed within the half-life of the previously administered compound, or at approximately 1 half-life of the previously administered compound, or within approximately 2, 3, 4, 5, 6, 7, 8, 9, or 10 half-lives of the previously administered compound. As a further example, a protein kinase C inhibitor may be administered to the patient approximately 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 21, 24, 36, 42, or 48 hours after administration of crizotinib, which has a half-life of approximately 42 hours, or within that time. As yet another example, a protein kinase C inhibitor may be administered to the patient approximately 0.5, 1, 2, 3, 4, 5, 6, 6.5, 7, 8, 9, 10, 11, 12, or 13 hours after administration of capmatinib, which has a half-life of approximately 6.5 hours, or within that time. In any case, sequential administration preferably follows a protocol in which the administered PKC inhibitor and cMET inhibitor provide additive, additive or synergistic effects, such as antiproliferative effects.

[0091] Patient selection The methods for treating patients described herein may further include a patient selection step or a step of determining which treatment process to offer a patient. In various embodiments, the method involves selecting patients having metastatic uveal melanoma or tumors having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"), preferably GNAQ / 11 tumors having mutations that activate the PKC signaling pathway. GNAQ / 11 tumors may be primary GNAQ / 11 tumors or metastatic GNAQ / 11 tumors. Patient selection may represent selection from a population of patients or selection of patients at a certain point in time or disease state throughout the treatment process or disease progression of individual patients. Thus, patient selection can identify not only which patients should receive the combination therapy described herein, but also when patients should receive it. In various embodiments, patient selection provides the benefit of an increased treatment index, for example, by reducing risk or increasing benefit to a given patient or patient population.

[0092] For example, the present invention provides a method for selecting and treating patients who have metastatic uveal melanoma or tumors having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"), preferably GNAQ / 11 tumors having mutations that activate the PKC signaling pathway, and co-administering a cMET inhibitor and a protein kinase C inhibitor represented by formula II to the patients as described above. The GNAQ / 11 tumor may be a primary GNAQ / 11 tumor or a metastatic GNAQ / 11 tumor.

[0093] As an example, the method may include selecting patients with metastatic uveal melanoma who have elevated cMET as determined by biopsy of uveal melanoma, and co-administering a cMET inhibitor and a protein kinase C inhibitor represented by formula II to the patients. In a further example, the method may include selecting patients with tumors having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor") who have elevated cMET as determined by biopsy of GNAQ / 11, and co-administering a cMET inhibitor and a protein kinase C inhibitor represented by formula II to the patients. A GNAQ / 11 tumor may be a primary GNAQ / 11 tumor or a metastatic GNAQ / 11 tumor. Therefore, patient selection may include (i) selecting patients with metastatic uveal melanoma or patients with a GNAQ / 11 tumor, and (ii) selecting patients who have elevated cMET as determined by biopsy of metastatic uveal melanoma or a GNAQ / 11 tumor, respectively.

[0094] The selection step can also help identify which patients may benefit from combination therapy and which may not. For example, the selection step can determine which patients should receive a protein kinase C inhibitor as combination therapy with a cMET inhibitor, and which patients should receive a protein kinase C inhibitor without a cMET inhibitor (e.g., a PKC inhibitor alone or in combination with another drug). The selection step can also determine when a given patient should receive combination therapy, or when he or she should receive monotherapy. Thus, the selection step can also help identify the patient's treatment course.

[0095] As a further example, the present invention also relates to a method for treating a patient having metastatic uveal melanoma or a tumor having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"). In biopsies of metastatic uveal melanoma or GNAQ / 11 tumors, the presence of cMETs should be evaluated. If the presence of cMET is elevated in metastatic uveal melanoma or GNAQ / 11 tumors, co-administration of a cMET inhibitor and a protein kinase C inhibitor to the patient is recommended. If the presence of cMET is not elevated in metastatic uveal melanoma or GNAQ / 11 tumors, the patient may be administered a protein kinase C inhibitor without a cMET inhibitor, including: Protein kinase C inhibitors provide a method represented by the above formula II.

[0096] Evaluation of the existence of cmet cMET is a receptor tyrosine kinase involved in several signaling pathways, including the MAPK and PI3K pathways. Its native endogenous ligand is hepatocyte growth factor (HGF). However, when cMET expression is stimulated by experimentally adding HGF to uveal melanoma cell lines, only some cell lines were found to exhibit antagonistic effects, which are the antiproliferative effects of PKC inhibitors of formula II. Despite maintaining the same HGF concentration for all cell lines in such experiments, it was discovered that some cell lines had elevated cMET presence while others did not. Therefore, without limiting the scope of this invention, it can be theorized that the complex mechanisms governing cMET and the PKC pathway, as well as ligand / messenger cross-conversion rather than HGF, are key factors in determining whether elevated cMET presence occurs. This invention is at least partially based on this finding that the presence of elevated cMET corresponds to tumor subtypes that can be favorably treated with the combination of protein kinase C inhibitors and cMET inhibitors described herein.

[0097] In various embodiments, the present invention provides a method for treating patients with metastatic uveal melanoma or tumors having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor") by co-administering a cMET inhibitor and a PKC inhibitor to patients who have elevated cMET, as determined by biopsy of the metastatic uveal melanoma or GNAQ / 11 tumor. The method may include, for example, determining which patients should receive combination therapy or confirming that combination therapy is appropriate for a patient by determining the presence of elevated cMET in the patient's biopsy.

[0098] For example, the present invention provides a method for treating a patient with metastatic uveal melanoma. The method comprises determining the presence of elevated cMET in a biopsy of the metastatic uveal melanoma and co-administering a cMET inhibitor and a protein kinase C inhibitor represented by formula II to the patient, as described above.

[0099] As another example, the present invention provides a method for treating patients having a tumor having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"), preferably a GNAQ / 11 tumor having a mutation that activates the PKC signaling pathway. The method comprises determining the presence of elevated cMET in a biopsy of the GNAQ / 11 tumor and co-administering a cMET inhibitor and a protein kinase C inhibitor represented by formula II to the patient, as described above. The GNAQ / 11 tumor may be a primary GNAQ / 11 tumor or a metastatic GNAQ / 11 tumor.

[0100] As used herein, the term “presence” refers qualitatively or quantitatively to the amount, e.g., concentration or number, of cMET in a given tumor, tissue, whole cells, cell lysates, cell fractions, or cell homogenates. Presence can be determined directly, for example, by measuring the cMET protein itself via IHC, immunoassay, or flow cytometry, or indirectly, for example, by measuring the expression level of cMET via RNA-seq or other transcriptome profiling. In some examples, presence refers to the cMET mRNA expression level, or presence refers to the cMET protein level, or in further examples, the cMET mRNA expression level may be used as a substitute for the cMET protein level due to the correlation between mRNA expression and the presence of the protein. Presence may be described in an absolute sense, for example, in terms of molar concentration, ng / ml, number, density per cell, or expression number, RNA expression level, or presence may be described in a relative sense compared to a baseline such as a calibration standard. For example, the “presence” determined by RNA expression levels may be compared to the RNA expression of stable “housekeeping genes” that are also present in normal cells and / or melanoma cells from a biopsy, as well as in similar types of normal (non-melanoma) cells taken from another location within the patient. For example, the “presence” determined by cMET protein levels may be compared to baseline cMET protein levels taken from healthy tissue of the same type taken from the patient or a similar patient, or from primary uveal melanoma cell cultures showing low cMET protein levels, or to standard baseline cMET protein levels established from a patient population.

[0101] More specific embodiments for evaluating the presence of cMET can be carried out by techniques such as ELISA, Western blotting, IHC-F, IHC-P, immunocytochemistry, immunofluorescence, flow cytometry, mass cytometry, and immunoprecipitation. Such techniques may include the use of anti-cMET antibodies. Anti-cMET antibodies suitable for use in ELISA, IHC, immunoprecipitation, Western blotting, flow cytometry, immunofluorescence, and other techniques are available from Thermo Fisher Scientific (Waltham, Massachusetts). The expression level of cMET RNA can be evaluated using cMET RNA-seq transcriptome techniques, Nanostring®, quantitative PCR, or other similar methods that measure the relative level of cMET mRNA. Such techniques are described in Example 5 below.

[0102] Where not specifically identified but described in relative terms, the presence of cMET can be understood as relative to a typical baseline level determined by a pathologist, physician, or other healthcare provider familiar with or trained in understanding cMET levels in a patient population with the same histological type of tumor. In various further examples, baseline may be defined by a reference sample, reference value, previous patient status, or standard. Furthermore, the presence of cMET can be determined based on tissue, whole cells, or their lysates, fractions, or homogenates. In each case, where relative cMET is described, it is a comparison between cMET assessments of the same type of sample. For example, cMET may typically be located in a transmembrane state having both extracellular and intracellular portions, or may be entirely intracellular, or extracellular, as in the case of cell lysates. In some samples, cMET may exist as a mixture of such states, but one state is typically evident as the most relevant to the measurement. In various embodiments, the presence of cMET may involve determining transmembrane cMET, intracellular cMET, extracellular cMET, or a combination thereof. Unless otherwise specified, the presence of cMETs in tissue or whole cells refers to transmembrane cMETs, i.e., transmembrane cMETs that function as cell surface receptors, while the presence of cMETs in lysates refers to extracellular cMETs.

[0103] The term “elevated” in relation to the presence of cMET refers to the presence of a cMET that is greater than baseline. For example, the presence of cMET may be elevated compared to a healthy state, a previous patient state, a reference sample, a reference value, or a standard, or any of these. In some cases, an elevated cMET may be determined by the opinion or diagnosis of a pathologist, physician, or other healthcare provider who is familiar with or trained to understand cMET levels in patient populations with the same histological type of tumor.

[0104] The assessment of the presence of cMET in a patient's metastatic uveal melanoma or GNAQ / 11 tumor is typically performed by evaluating the presence of cMET in biopsies obtained from such patient's metastatic uveal melanoma or GNAQ / 11 tumor, respectively. The cMET level in the biopsy can be assessed by performing measurements on the biopsy itself, or simply by referring to information or reports indicating the level of cMET present in the biopsy, such as cMET expression level, RNA copy number, or cMET concentration.

[0105] The level of cMET presence can be determined by any means, including the application of conventional biochemical techniques. While the presence of cMET as used herein may correspond to total cMET, the various techniques used to determine total cMET do not necessarily require direct measurement of total cMET concentration. For example, certain techniques allow for the determination of total cMET concentration by measuring RNA expression levels, modified forms of cMET, or biomarkers.

[0106] In various embodiments, the method further includes taking and obtaining a biopsy from a patient, for example, by extraction or collection. For example, a biopsy may be obtained by surgical excision, by needle biopsy (e.g., core needle biopsy), or by other methods or approaches known in the art. The biopsy may typically be a solid tumor biopsy. The cells of the biopsy may be isolated, cultured, lysed, fractionated, homogenized, or a combination thereof. The method may include taking a single biopsy or taking multiple biopsies. For example, the method may include obtaining multiple biopsies or obtaining information regarding the presence of cMETs in multiple biopsies taken at different time points. Taking multiple biopsies over time may be useful in determining how the patient's disease has progressed and may help in identifying when combination therapy is appropriate for a given patient.

[0107] In various embodiments, assessing the presence of cMET as a protein or RNA transcriptome involves contacting the biopsy material with a combination of a cMET-specific ligand and a reporter ligand (labeled ligand) that interacts with the cMET-specific ligand. As used herein, a ligand refers to a compound or biological molecule that has a binding interaction with cMET. For example, a ligand may be a cMET-specific antibody or an RNA sequence that selectively binds to cMET RNA. Also, in the context of this section, the term "presence of cMET" means cMET protein and / or cMET RNA. The content of the discussion regarding the assessment of cMET presence indicates whether a protein or RNA is being assessed. The ligand may also be a small molecule that binds to cMET. The ligand or antibody may be conjugated with a label, or a secondary ligand that can associate with a primary ligand may possess a label. Examples of labels include chromogenic labels, fluorescent labels, radioactive labels, or isotopic labels. Anti-cMET antibodies are commercially available from Thermo Fisher Scientific (Waltham, Massachusetts) in conjugate-labeled and unconjugate forms and may be labeled using conventional labeling techniques before use, or detected using commercially available reporter ligands. Antibodies can be detected by, for example, radiolabeling, fluorescent labeling, hapten labeling such as biotin, or by directly labeling the antibody itself with enzymes such as horseradish peroxidase or alkaline phosphatase, or by using a secondary antibody against a primary antibody. Alternatively, an unlabeled primary antibody specific to cMET can be used in conjunction with a labeled secondary antibody specific to the primary antibody. Alternatively, cMET RNA expression can be measured in biopsy using RNA-seq, nanostringing, quantitative PCR, or other similar methods that measure the relative level of cMET mRNA. RNA expression thresholds and cutoffs are obtained from low-expression and high-expression cMET tissues to determine whether a particular uveal melanoma tissue has high or low cMET RNA expression.

[0108] The anti-cMET antibody may be specific to the phosphorylated form of cMET, specific to the unphosphorylated form of cMET, or nonspecific with respect to phosphorylated or unphosphorylated cMET. The antibody may also be specific to human cMET. Since phosphorylated cMET protein, unphosphorylated cMET protein, and total cMet protein are correlated, each corresponds to the presence of an appropriate elevated concentration of cMET.

[0109] In various embodiments, the presence of cMET can be determined from a biopsy prepared in the form of a tissue section, which may be fixed and paraffin-embedded or frozen. The presence of cMET in the tissue can be assessed by contacting such a tissue section with a labeled antibody. When determining the relative presence of cMET, the tissue section can be compared to a tissue section from the same type of healthy tissue from which the biopsy originated, either from the same patient or a representative patient. The tissue section can also be compared to a tissue section from a previous biopsy obtained from the same tumor. For example, the presence of cMET can be assessed by obtaining an initial biopsy from a patient's metastatic uveal melanoma or GNAQ / 11 tumor and preparing a first tissue section therefrom; obtaining a subsequent biopsy from the same site of the patient's metastatic uveal melanoma or GNAQ / 11 tumor and preparing a second tissue section therefrom; contacting the first tissue section with a labeled cMET antibody; contacting the second tissue section with a labeled cMET antibody; and determining the presence of elevated cMET in the second tissue section compared to the first tissue section.

[0110] Biopsy preparations can be compared or scored according to the presence of cMET. Such scoring systems can be devised using values ​​that represent the range of cMET levels expected in a given patient population, or against previous levels of cMET in the same tumor in the same patient, or against cMET levels based on a reference sample. In this way, the presence of elevated cMET can be determined. The scoring system may be based on a scale of 0 to 4, for example, where a score of 0 corresponds to the absence of cMET expression in the tissue, a score of 1 may correspond to cMET expression in healthy tissue, and tissues showing elevated cMET correspond to a score of 2 or higher, or 3 or 4. For example, tissue sections can be evaluated using IHC (immunohistochemistry) and scored on a scale of 0 to 3 or 4, where a score of 0 corresponds to the absence of cMET expression in the tissue, while a score of 1 may correspond to the presence of cMET in healthy tissue of the type from which the tumor originates, and a score of 2, 3 or 4 corresponds to a reference sample with increased cMET presence. In some cases, a score of 1 may refer to a standard cMET level, i.e., a low cMET score, derived from uveal melanoma tumors in which the combination therapy described herein did not exhibit a synergistic antiproliferative effect. In contrast, a score of 2, 3, or 4 may refer to a standard cMET level derived from uveal melanoma tumors in which the combination exhibited a synergistic antiproliferative effect, as determined by the Bliss, HSA, or Loewe synergy / antagonism models. Other immunoassay techniques are similarly applicable and can utilize similar scoring systems. Such immunoassay techniques are described in Example 6 of the experimental section herein and are generally and particularly preferred approaches for determining the presence of elevated cMET according to each of the various aspects, embodiments, and sub-embodiments of the Invention.See also the references cited therein, such as Wikipedia at en.wikipedia.org / wiki / immunohistochemistry(2020) and J.A.Ramos-Vara, “Technical Aspects of Immunohistochemistry,” Veterinary Pathology, 42(4);405-426(2005). Alternatively, the presence of cMET can be determined using RNA transcriptome techniques. Such techniques are described in Example 5 of the Experimental Section of this Specification and are generally and in particular preferred approaches for determining the presence of elevated cMET according to each of the various aspects, embodiments, and sub-embodiments of the Invention.

[0111] As another example, the presence of cMET can be determined from tumor cells obtained by biopsy. Tumor cells from a biopsy may be optionally isolated, cultured, lysed, fractionated, homogenized, or a combination thereof. The presence of cMET in cells can be assessed by contacting the cells, their lysates, fractions, or homogenates with a labeled antibody. When determining the relative presence of cMET, the cells, lysates, fractions, or homogenates tested may be compared to cell samples from healthy cells of the same type from which the biopsy originated, either from the same patient or a representative patient. Cells may also be compared to cells from previous biopsies obtained from the same tumor. In other examples, cells may be compared to reference cells, which may be healthy cells or primary uveal melanoma cells (e.g., cell lines that do not contain elevated cMET). For example, evaluating the presence of cMET in reference cells may involve first obtaining a biopsy from the patient's metastatic uveal melanoma or GNAQ / 11 tumor, and then preparing a cell preparation therefrom in the form of cell isolates, lysates, homogenates, fractions, or a combination thereof. Next, reference cells (e.g., uveal melanoma cells or healthy cells of the same histological type as the biopsy, but not limited to these) are obtained and prepared in the same form as the biopsy cell preparation. The level of cMET in the biopsy cell preparation and the reference cell preparation was evaluated using a labeled cMET antibody. The levels of cMET were compared to determine whether the biopsy cell preparation contains a higher or significantly higher level of cMET than the cMET level in the reference cell preparation. This evaluation can be determined using IHC techniques as described above.

[0112] In various cases, the presence of cMETs in the biopsy is elevated by at least or approximately 25%, 50%, 75%, 100%, 125%, 150%, 200%, 300%, 400%, 500%, 1000%, 2500%, or at least or approximately 5000% compared to a given level. The given level may correspond to the presence of cMETs in healthy uveal cells or healthy cells of the type from which the biopsy originates in the same patient population relative to the average presence of cMETs in healthy uveal cells or healthy cells of the type from which the biopsy originates in the same patient or patient population relative to the average presence of cMETs in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the type from which the biopsy originates in the same patient population relative to a previously assessed level of cMET presence in a previous biopsy of the same cell type from the same patient relative to the presence of cMETs in healthy uveal cells or healthy cells of the type from which the biopsy originates in the same patient relative to a previously assessed level of cMET presence in a previous biopsy of the same cell type from the same patient.

[0113] tumor Methods described herein may include treating or selecting patients with proliferative disorders such as metastatic uveal melanoma or tumors having a gene mutation in GNAQ or GNA11, preferably a GNAQ mutation or GNA11 mutation that activates the PKC signaling pathway ("GNAQ / 11 tumor"). A GNAQ / 11 tumor may be a primary GNAQ / 11 tumor or a metastatic GNAQ / 11 tumor. For example, a patient may have a metastatic uveal melanoma determined or diagnosed by a physician or other healthcare provider who has experience or is trained in identifying metastatic uveal melanoma. As an alternative example, a patient may have a GNAQ / 11 tumor, as determined based on evaluation of a biopsy from the patient. A GNAQ / 11 tumor may be a primary GNAQ / 11 tumor or a metastatic GNAQ / 11 tumor. Evaluation of patient biopsies may include DNA sequencing (e.g., next-generation sequencing (NGS)) analysis of solid tumor or liquid biopsies (e.g., blood biopsies), detection of cell-free circulating tumor DNA (cfDNA), or detection of DNA in circulating tumor cells (CTCs).

[0114] GNAQ / 11 tumors may contain one or more mutations, including substitution mutations, insertion mutations, and / or deletion mutations. In some embodiments, GNAQ mutations or GNA11 mutations are gain-of-function mutations. In some embodiments, GNAQ mutations or GNA11 mutations activate the PKC signaling pathway. In various embodiments, GNAQ mutations or GNA11 mutations may be substitutions of glutamine (Q209) at codon 209 and / or arginine (R183) at codon 183. GNAQ mutations or GNA11 mutations may be substitutions other than glutamine (Q209) at codon 209, substitutions other than arginine (R183) at codon 183, or neither. In some embodiments, the GNAQ mutation is one of Q209P, Q209L, Q209H, Q209K, or Q209Y, or the GNA11 mutation is one of Q209P, Q209L, Q209K, or Q209H. In further embodiments, the GNAQ mutation may be R183Q, or the GNA11 mutation may be R183C or R183H. In further examples, the GNAQ mutation or GNA11 mutation is found in one or more of the following: R256, L279, R166, A168, R210, R213, R166, A231, A342, D333, G171, R147, R73, T47, E191, E221, R149, T175, T379, T85, A86, E163, D195, E319, E191, E280, E49, P293, R300, R338, R60, D155, D205, D321, I226, R37, or V240. In further examples, a GNAQ / 11 tumor may contain one or more of the Q209P, Q209L, Q209H, Q209K, Q209Y, or R183Q mutations in GNAQ, or a GNAQ / 11 tumor may contain one or more of the Q209P, Q209L, Q209H, or Q209K mutations in GNA11. Further examples of mutations in GNAQ or GNA11 are described in International Publication 2020 / 146355, which is incorporated herein by reference in its entirety.

[0115] In various aspects, GNAQ / 11 tumors are either primary uveal melanoma or metastatic uveal melanoma. In various aspects, GNAQ / 11 tumors may be non-uveal tumors, such as pancreatic tumors, gastric tumors, colorectal tumors, uterine tumors, cervical tumors, bladder tumors, hepatocellular carcinomas, head and neck tumors, prostate tumors, breast tumors, lung adenocarcinomas, or cutaneous melanomas. In other aspects, the tumors may be central nervous system tumors, such as glioblastomas. In further aspects, GNAQ / 11 tumors may be non-uveal tumors, or non-melanocyte tumors, or non-uveal non-melanocyte tumors. In some aspects, the tumors may be carcinomas. For example, the tumors may be pancreatic adenocarcinomas, gastric adenocarcinomas, cervical cancers, and lung adenocarcinomas. In various aspects, the aforementioned tumors may be metastatic or they may be non-metastatic. In some aspects, the tumors may be non-metastatic non-uveal tumors. For example, the tumors may be other than metastatic uveal melanomas. In even further examples, the tumors may be metastatic tumors.

[0116] For example, a GNAQ / 11 tumor may be a solid tumor that has metastasized to a secondary site such as the liver. Another example is a GNAQ / 11 tumor that may be a metastatic uveal melanoma that has metastasized to the liver. In a further embodiment, GNAQ / 11 tumors have a high risk of recurrence or metastasis, as determined by a gene expression profile such as DecisionDx™, available from Castle Biosciences (Friendswood, Texas). In some embodiments, the method includes selecting patients at high risk of uveal melanoma recurrence or metastasis, as determined by a pathologist, physician, or other healthcare provider who is familiar with or trained in understanding the risk of recurrence or metastasis of tumor types such as uveal melanoma or GNAQ / 11 tumors.

[0117] In various embodiments, GNAQ / 11 tumors are metastatic, metastasizing to secondary sites with HGF concentrations ranging from approximately 0.1 ng / ml to approximately 1,000 ng / ml. In another example, a patient may have metastatic uveal melanoma that has metastasized to, or is at risk of metastasizing to, secondary sites with HGF concentrations ranging from approximately 0.1 ng / ml to approximately 1,000 ng / ml. Such HGF concentrations include, for example, 0.1 ng / ml to approximately 500 ng / ml, 1 ng / ml to approximately 500 ng / ml, 1 ng / ml to approximately 100 ng / ml, 1.5 ng / ml to approximately 100 ng / ml, 3 ng / ml to approximately 100 ng / ml, 5 ng / ml to approximately 100 ng / ml, 7 ng / ml to approximately 100 ng / ml, 10 ng / ml to approximately 100 ng / ml, 15 ng / ml to approximately 100 ng / ml, 20 ng / ml to approximately 100 ng / ml, 1 ng / ml to approximately 30 ng / ml, and 1.5 ng / ml. 1 ng / ml to approximately 30 ng / ml, 3 ng / ml to approximately 30 ng / ml, 5 ng / ml to approximately 30 ng / ml, 7 ng / ml to approximately 30 ng / ml, 10 ng / ml to approximately 30 ng / ml, 15 ng / ml to approximately 30 ng / ml, 20 ng / ml to approximately 30 ng / ml, 1 ng / ml to approximately 16 ng / ml, 1.5 ng / ml to approximately 16 ng / ml, 3 ng / ml to approximately 16 ng / ml, 5 ng / ml to approximately 16 ng / ml, 7 ng / ml to approximately 16 ng / ml, or 10 ng / ml to approximately 16 ng / ml. For example, the HGF concentration may be approximately or at least 0.5, 1.0, 1.5, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0, 11.0, 12.0, 13.0, 14.0, 15.0, 16.0, 17.0, 18.0, 19.0, 20.0, 25.0, 30.0, 35.0, 40.0, 50.0, 60.0, 70.0, 80.0, 90.0, or approximately or at least 100.0 ng / ml. In some embodiments, metastatic uveal melanoma or GNAQ / 11 tumor may be a metastatic tumor located in the liver, i.e., a liver metastasis of a previous primary tumor, for example, originating histologically from another tissue.

[0118] In another embodiment, a GNAQ / 11 tumor may lack one or more activating mutations in BRAF, KRAS, or EGRF, or a GNAQ / 11 tumor may have one or more low-loading mutations in BAP1, SF3B1, EIF1AX, TERT, BRAF, CDKN2A, NRAS, KRAS, or EGRF.

[0119] Evaluation of gene mutations Aspects of the present invention may include evaluating whether a patient, tumor, or biopsy exhibits a genetic mutation in GNAQ or GNA11. Such evaluation may include identifying a GNAQ mutation or a GNA11 mutation in a patient, for example, in a solid tumor biopsy or a liquid biopsy (e.g., a blood biopsy), by identifying circulating cell-free tumor DNA (cfDNA) or DNA in circulating tumor cells (CTCs) in the patient, which can be obtained, for example, by taking a liquid sample from the patient. The liquid sample may be a blood sample. Genetic mutations may also be evaluated from the patient's solid tumor, which typically involves taking a tissue biopsy from the tumor.

[0120] Determining whether a patient, tumor, or biopsy has a specific mutation, such as a GNAQ or GNA11 gene mutation, may involve collecting and / or analyzing patient samples. Examples of patient samples of interest include carcinoma tissue, cancerous tissue, solid tumor tissue, tumor tissue, body tissue, blood, serum, plasma, or bodily fluids (e.g., circulating blood containing tumor DNA) obtained from a patient. Patient samples used in the methods described herein may also include, but are not limited to, tissue samples such as gastrointestinal, mucosa, submucosa, intestine, esophagus, ileum, rectum, cervix, colon, epidermis, lung, thymus, pancreas, stomach, rectum, skin, subcutaneous, or lymph samples. Samples may also include cell samples, such as skin cell samples. The presence of a target gene mutation in patient-derived samples can be determined using a variety of analyses. For example, gene mutations can be determined by various methods, such as gene sequencing, Southern blotting, FISH, high-throughput sequencing, phage display, shotgun sequencing, PCR, or nucleotide analysis such as RT-PCR. Gene mutations can also be determined by analyzing mRNA, protein, or nucleotide levels, or by performing dynamic allele-specific hybridization techniques. Appropriate techniques for evaluating gene mutations are described in International Publication No. 2020 / 146355, which is incorporated herein by reference in its entirety.

[0121] HGF evaluation In a further aspect of the present invention, the method may include evaluating hepatocyte growth factor (HGF) levels in a patient's tumor, a tumor biopsy, or non-tumor tissue in which the tumor is located. Some types of tumor cells can secrete HGF, which can induce cMET antagonistic activity to the protein kinase C inhibitors described herein in certain types of tumors. In some embodiments, HGF is secreted by tumor cells, while in other embodiments, HGF is secreted by tumor tissue or non-tumor tissue in which the tumor cells are located. Such evaluation of HGF can be performed, for example, by an enzyme-linked immunosorbent assay (ELISA). For example, an HGF ELISA kit is available from Thermo Fisher Scientific (Waltham, Massachusetts).

[0122] In various embodiments, patients have metastatic uveal melanoma or GNAQ / 11 tumors with HGF concentrations ranging from approximately 0.1 ng / ml to approximately 1,000 ng / ml during biopsy. The biopsy may be a tissue biopsy, surgical excision, or a combination of both. In some embodiments, the method further comprises selecting patients having metastatic uveal melanoma or GNAQ / 11 tumors with HGF concentrations ranging from approximately 0.1 ng / ml to approximately 1,000 ng / ml, as determined by evaluating the biopsy of the metastatic uveal melanoma or GNAQ / 11 tumors. Such HGF concentrations include, for example, 0.1 ng / ml to approximately 500 ng / ml, 1 ng / ml to approximately 500 ng / ml, 1 ng / ml to approximately 100 ng / ml, 1.5 ng / ml to approximately 100 ng / ml, 3 ng / ml to approximately 100 ng / ml, 5 ng / ml to approximately 100 ng / ml, 7 ng / ml to approximately 100 ng / ml, 10 ng / ml to approximately 100 ng / ml, 15 ng / ml to approximately 100 ng / ml, 20 ng / ml to approximately 100 ng / ml, 1 ng / ml to approximately 30 ng / ml, and 1.5 ng / ml. 1 ng / ml to approximately 30 ng / ml, 3 ng / ml to approximately 30 ng / ml, 5 ng / ml to approximately 30 ng / ml, 7 ng / ml to approximately 30 ng / ml, 10 ng / ml to approximately 30 ng / ml, 15 ng / ml to approximately 30 ng / ml, 20 ng / ml to approximately 30 ng / ml, 1 ng / ml to approximately 16 ng / ml, 1.5 ng / ml to approximately 16 ng / ml, 3 ng / ml to approximately 16 ng / ml, 5 ng / ml to approximately 16 ng / ml, 7 ng / ml to approximately 16 ng / ml, or 10 ng / ml to approximately 16 ng / ml. For example, the HGF concentration may be approximately or at least 0.5, 1.0, 1.5, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0, 11.0, 12.0, 13.0, 14.0, 15.0, 16.0, 17.0, 18.0, 19.0, 20.0, 25.0, 30.0, 35.0, 40.0, 50.0, 60.0, 70.0, 80.0, 90.0, or approximately or at least 100.0 ng / ml.

[0123] Pharmaceuticals Aspects of the present invention further provide pharmaceuticals comprising a protein kinase C inhibitor, a cMET inhibitor, and a pharmaceutically acceptable carrier.

[0124] A pharmaceutical product may comprise one or more pharmaceutical compositions in which a protein kinase C inhibitor and a cMET inhibitor may be formulated together or separately. For example, a pharmaceutical product may comprise a single pharmaceutical composition comprising a protein kinase C inhibitor, a cMET inhibitor, and a pharmaceutically acceptable carrier. In such an example, the pharmaceutical product may be represented by a single dosage form comprising a protein kinase C inhibitor, a cMET inhibitor, and a pharmaceutically acceptable carrier.

[0125] In further examples, the pharmaceutical may contain a first pharmaceutical composition comprising a protein kinase C inhibitor and a second pharmaceutical composition comprising a cMET inhibitor, each independently having a pharmaceutically acceptable carrier.

[0126] A pharmaceutical product may include “fixed combinations” and “unfixed combinations” of two or more active ingredients, such as a protein kinase C inhibitor and a cMET inhibitor. The term “fixed combination” means that the active ingredients, e.g., a protein kinase C inhibitor and a cMET inhibitor, are formulated as a single pharmaceutical composition. For example, a fixed combination may have a protein kinase C inhibitor and a cMET inhibitor in the same unit dosage form (e.g., a capsule, tablet, or sachet). The term “unfixed combination” refers to two or more active ingredients, e.g., a protein kinase C inhibitor and a cMET inhibitor, that are formulated independently as separate pharmaceutical compositions and can be administered to a patient separately. In various embodiments, each active ingredient in an unfixed combination is administered within a period that allows the protein kinase C inhibitor and the cMET inhibitor to exhibit a synergistic effect, e.g., additive, multi-additive, or synergistic effect. An unfixed combination may also include the use of a single drug together with one or more fixed combination products, where each independent formulation contains different amounts of the active ingredients. Therefore, it should be understood that pharmaceuticals include cases where the active ingredients, such as protein kinase C inhibitors and cMET inhibitors, are administered as completely separate pharmaceutical dosage forms. In various examples, protein kinase C inhibitors and cMET inhibitors may each be present in separate pharmaceutical compositions that are marketed independently of each other.

[0127] Independent components of a drug can be administered simultaneously or at staggered intervals; that is, each individual component of a drug can be administered at the same or different time points in time, and at the same or different time intervals between the administration of any given component. For example, the time interval between administrations can be selected such that the effect on the disease treated by the combination of a protein kinase C inhibitor and a cMET inhibitor is greater / higher than the effect obtained by using either a protein kinase C inhibitor or a cMET inhibitor alone.

[0128] In various embodiments, the pharmaceutical comprises a protein kinase C inhibitor, a cMET inhibitor, and a pharmaceutically acceptable carrier, the protein kinase C inhibitor being represented by formula II as described herein.

[0129] In various embodiments, the pharmaceutical product comprises a protein kinase C inhibitor and a cMET inhibitor formulated together in a single unit dosage form containing one or more pharmaceutically acceptable carriers. In a further embodiment, the pharmaceutical comprises a protein kinase C inhibitor and a cMET inhibitor formulated into two distinct unit dosage forms, each having an independently selected pharmaceutically acceptable carrier.

[0130] A pharmaceutical product may contain one or more pharmaceutically acceptable carriers. A pharmaceutically acceptable carrier may be useful for preparing a pharmaceutical composition that is generally safe, non-toxic, and not biologically or otherwise undesirable. For example, a carrier may be acceptable for human pharmaceutical use. A product or composition containing a pharmaceutically acceptable carrier may contain one or more such carriers. Examples of pharmaceutically acceptable carriers include solid carriers, solvents, non-solvent liquid carriers, dispersions, coatings, surfactants, antioxidants, preservatives (e.g., antimicrobials, antifungals), isotonic agents, absorption retarders, salts, preservatives, drugs, drug stabilizers, binders, excipients, disintegrants, lubricants, sweeteners, flavorings, dyes, combinations thereof, or other carriers. (See Remington's Pharmaceutical Sciences, 18th Ed., Mack Printing Company, 1990, pp. 1289–1329). Some examples of substances that can serve as pharmaceutically acceptable carriers include: (1) sugars (e.g., lactose, glucose, and sucrose), (2) starches (e.g., corn starch, potato starch, and substituted or unsubstituted 3-cyclodextrins), (3) cellulose and its derivatives (e.g., sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate), (4) tragacanth powder, (5) malt, (6) gelatin, (7) talc, (8) excipients (e.g., cocoa butter and suppository waxes), (9) oils (e.g., peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil) Examples include (10) oat oil and soybean oil), (11) glycols (e.g., propylene glycol), (12) polyols (e.g., glycerin, sorbitol, mannitol, and polyethylene glycol), (13) esters (e.g., ethyl oleate and ethyl laurate), (14) agar, (15) buffers (e.g., magnesium hydroxide and aluminum hydroxide), (16) alginic acid, (17) water free of pyrogens, (18) isotonic saline, (19) Ringer's solution, (20) ethyl alcohol, (21) phosphate buffer, and (22) other non-toxic, suitable substances used in pharmaceutical formulations.

[0131] It can be understood that various methods of disclosure, pharmaceuticals, and kits may include PKC inhibitors and cMET inhibitors as part of a pharmaceutical composition. Such compositions may be administered, for example, in volumetric unit form. Pharmaceutical compositions are preferably formulated to suit their intended route of administration. It will be understood that the compounds and compositions described herein may be administered by various routes of administration, e.g., orally, parenterally, rectally, or vaginally. Administration may be by injection, for example, subcutaneous, intrapancreatic, intravenous, intramuscular, intraperitoneal, intrathecal, intraperitoneal, intraocular, or intrasternal injection. Further examples include infusion techniques, inhalation sprays, sublingual, cutaneous, rectal, or ocular administration, for example, in the form of eye drops.

[0132] Preparations for injection, such as sterile aqueous or oily suspensions for injection, can be formulated according to known techniques using appropriate dispersants or wetting and suspending agents. Sterile preparations for injection may also be sterile solutions or suspensions for injection in non-toxic, parenterally acceptable diluents or solvents (e.g., as a solution in 1,3-propanediol). Acceptable vehicles and solvents that can be used include water, Ringer's solution, and isotonic sodium chloride solutions. Furthermore, sterile fixatives have traditionally been used as solvents or suspension media. For this purpose, any low-irritation fixative, including synthetic monoglycerides or diglycerides, can be used. In addition, fatty acids such as oleic acid are used in the preparation of injections.

[0133] Formulations suitable for oral administration may be in the form of capsules, cachets, pills, tablets, lozenges (using a flavored base, usually sucrose and acacia or tragacanth), powders, granules, or liquids or suspensions in aqueous or non-aqueous liquids, or as oil-in-water or water-in-oil liquid emulsions, or as elixirs or syrups, or as lozenges (using an inert matrix such as gelatin and glycerin, or sucrose and acacia), and / or mouthwashes, each containing a predetermined amount of the compound of the present invention as the active ingredient. The composition may also be administered as a bolus, lick, or paste.

[0134] Solid dosage forms for oral administration may include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active agent may be mixed with at least one inert diluent (e.g., sucrose, lactose, or starch). Such dosage forms may also contain further substances other than inert diluents (e.g., lubricants such as magnesium stearate), as is common practice. In the case of capsules, tablets, and pills, the dosage form may contain a buffer. Tablets and pills may further be prepared using enteric coating. In solid dosage forms for oral administration (capsules, tablets, pills, sugar-coated tablets, powders, granules, etc.), the compounds of the present invention are mixed with one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate, and / or any of the following:

[0135] (1) Fillers or bulking agents (e.g., starch, cyclodextrin, lactose, sucrose, glucose, mannitol, and / or silicic acid); (2) Binders (e.g., carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and / or acacia); (3) Moisturizers (e.g., glycerol); (4) Disintegrants (e.g., agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate); (5) Dissolution retarders (e.g., paraffin); (6) Absorption enhancers (e.g., quaternary ammonium compounds) (7) Humectants (e.g., acetyl alcohol and glycerol monostearate); (8) Absorbents (e.g., kaolin and bentonite clay); (9) Lubricants (e.g., talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof); and (10) Coloring agents. In the case of capsules, tablets, and pills, the pharmaceutical composition may also contain buffering agents. Similar types of solid compositions may also be used as fillers in soft and hard-filled gelatin capsules, using excipients such as lactose or milk sugar and high molecular weight polyethylene glycol.

[0136] Tablets may be prepared by compression or molding with one or more auxiliary components as optional. Compressed tablets may be prepared using a binder (e.g., gelatin or hydroxypropyl methylcellulose), a lubricant, an inert diluent, a preservative, a disintegrant (e.g., sodium starch glycolate or cross-linked carboxymethylcellulose sodium), a surfactant, or a dispersant. Molded tablets may be prepared by molding a mixture of powdered inhibitors moistened with an inert liquid diluent using appropriate machinery.

[0137] Tablets, as well as other solid dosage forms such as sugar-coated tablets, capsules, pills, and granules, can be optionally notched or prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulation field. They may also be formulated to provide sustained or controlled release of the active ingredient therein, for example, using hydroxypropyl methylcellulose, other polymer matrices, liposomes, and / or microspheres in varying proportions to provide a desired release profile. They may be sterilized, for example, by filtration through a bacterial-retaining filter or by incorporating a sterilizer in the form of a sterile solid composition that can be dissolved in sterile water or some other sterile injection medium immediately before use. These compositions may also optionally contain an opacifying agent and may be compositions that release only the active ingredient, or optionally in a delayed manner, preferentially in a particular part of the gastrointestinal tract.

[0138] Examples of embedding compositions that can be used include polymeric substances and waxes. The compounds of the present invention may also be in microencapsulated form, together with one or more of the above excipients, if appropriate.

[0139] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups, and elixirs. In addition to the active ingredient, the liquid dosage form may contain inert diluents and solubilizers commonly used in the art, such as water or other solvents, as well as emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerol, tetrahydrofuryl alcohol, polyethylene glycol, and fatty acid esters of sorbitan, and mixtures thereof. In addition to inert diluents, the oral composition may also contain adjuvants such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents, colorants, fragrances, and preservatives. In addition to the activity inhibitor, the suspension may contain suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methhydroxyl, bentonite, agar and tragacanth, and mixtures thereof.

[0140] The activator can also be administered in the form of liposomes. As is known in the art, liposomes are generally derived from phospholipids or other lipid substances. Liposomes are formed by monolayers or multilayers of hydrated liquid crystals dispersed in an aqueous medium. Any non-toxic, physiologically acceptable, and metabolizable lipid capable of forming liposomes can be used. The composition in liposome form may contain stabilizers, preservatives, excipients, etc., in addition to the compounds of the present invention. Examples of lipids are both natural and synthetic phospholipids and phosphatidylcholine (lecithin). Methods for forming liposomes are known in the art. See, for example, Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, NW, p.33 et seq. (1976).

[0141] Formulations for rectal or vaginal administration may be provided as suppositories, which may be prepared by mixing one or more inhibitors with one or more suitable non-irritating excipients or carriers (e.g., cocoa butter, polyethylene glycol, suppository wax, or salicylate), which are solid at room temperature but liquid at body temperature, and thus melt in the rectum or vaginal cavity to release the active agent. Formulations suitable for vaginal administration also include pessaries, tampons, creams, gels, pastes, foams, or spray formulations containing carriers known to be suitable in the art.

[0142] Dosage forms for topical or transdermal administration of inhibitors include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches, and inhalants. The active ingredient may be mixed under sterile conditions with a pharmaceutically acceptable carrier and any preservatives, buffers, or propellants as needed. Ointments, pastes, creams, and gels may contain, in addition to the compounds of the present invention, excipients such as animal and vegetable fats, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silicic acid, talc, and zinc oxide, or mixtures thereof. Powders and sprays may contain, in addition to the compounds of the present invention, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate, and polyamide powder, or mixtures thereof. Sprays may further contain conventional propellants such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons (such as butane and propane). Alternatively, compounds useful for application of the methods of the present invention may be administered by aerosol. This is achieved by preparing aqueous aerosols, liposome preparations, or solid particles containing the composition. Non-aqueous (e.g., fluorocarbon propellant) suspensions can be used. Ultrasonic nebulizers are preferred because they minimize the exposure of the agent to shear, which can lead to the degradation of the compound. Typically, aqueous aerosols are prepared by formulating an aqueous solution or suspension of the compound of the present invention together with conventional pharmaceutically acceptable carriers and stabilizers. Carriers and stabilizers vary depending on the requirements of the particular composition but typically include nonionic surfactants (Tween®, Pluronic®, sorbitan esters, lecithin, Cremophor®), pharmaceutically acceptable cosolvents such as polyethylene glycol, harmless proteins such as serum albumin, amino acids such as oleic acid and glycine, buffers, salts, sugars, or sugar alcohols. Aerosols are generally prepared from isotonic solutions.

[0143] Transdermal patches offer the further advantage of providing controlled delivery of the compounds of the present invention to the body. Such dosage forms can be prepared by dissolving or dispersing the drug in a suitable medium. The flow of the inhibitor through the skin can also be increased by using absorption enhancers. The rate of such flow can be controlled by providing a rate-controlled membrane or by dispersing the inhibitor in a polymer matrix or gel.

[0144] Depot formulations for injection are prepared by forming a microcapsule matrix of the inhibitor within a biodegradable polymer such as polylactide-polyglycolide. The drug release rate can be controlled depending on the drug-to-polymer ratio and the properties of the specific polymer used. Other examples of biodegradable polymers include poly(orthoester) and poly(anhydride). Depot injection formulations are also prepared by encapsulating the drug in liposomes or microemulsions that are compatible with body tissues.

[0145] Pharmaceutical compounds and compositions may be administered “systemically” or “peripherally,” meaning an administration that enters the patient’s system in a manner that allows for metabolism and other similar processes (e.g., subcutaneous administration). As used herein, the term “administered parenterally” means a mode of administration other than intestinal and topical administration, usually by injection, and includes, but is not limited to, intravenous, intramuscular, intra-arterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subepidermal, intra-articular, subcapsular, subarachnoid, intraspinal, and intrasternal injections and infusions.

[0146] A pharmaceutical composition of the present invention suitable for parenteral administration comprises one or more compounds of the present invention in combination with one or more pharmaceutically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions, or emulsions, or sterile powders that can be reconstituted into a sterile injectable solution or dispersion immediately before becoming isotonic with the blood of the target recipient, or a suspending agent or thickener. Examples of suitable aqueous and non-aqueous carriers that may be used in the pharmaceutical composition of the present invention include water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.), and suitable mixtures thereof, vegetable oils (e.g., olive oil), and injectable organic esters (e.g., ethyl oleate). Appropriate fluidity can be maintained, for example, by the use of a coating material such as lecithin, by maintaining the required particle size in the case of a dispersion, and by the use of a surfactant.

[0147] These compositions may also contain adjuvants such as preservatives, humectants, emulsifiers, and dispersants. Prevention of microbial action can be ensured by including various antimicrobial and antifungal agents, such as parabens, chlorobutanol, and phenolsorbic acid. It may also be desirable to include tonic modifiers such as sugars and sodium chloride in the composition. Furthermore, sustained absorption of injectable pharmaceutical forms can be achieved by including absorption-delaying agents such as aluminum monostearate and gelatin.

[0148] In some cases, it is desirable to delay the absorption of compounds from subcutaneous or intramuscular injection in order to extend the effect of compounds useful in carrying out the methods of the present invention. For example, delayed absorption of parenterally administered drug forms can be achieved by dissolving or suspending the drug in an oil vehicle.

[0149] In various embodiments, the compounds described herein may be in the form of pharmaceutically acceptable salts. Such salts may be acidic salts or metallic salts, such as alkali or alkaline earth salts. The salts can be prepared in situ during the final isolation and purification of the compound by separately reacting the base or acidic functional group in the compound with a suitable organic or inorganic acid or base, respectively. Representative salts include, but are not limited to, acetate, adipine, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphor sulfonate, digluconate, cyclopentanepropionate, dodecyl sulfate, ethanesulfonate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, and undecanoate. Furthermore, the basic nitrogen-containing group can be quaternized with agents such as chlorides, bromides, and alkyl halides such as methyl iodide, ethyl, propyl, and butyl; dialkyl sulfates such as dimethyl sulfate, diethyl, dibutyl, and diamyl; long-chain halides such as chlorides, bromides, and decyl iodide, lauryl, myristyl, and stearyl; and aralkyl halides such as benzyl bromide and phenethyl.

[0150] Examples of acids that can be used to form pharmaceutically acceptable acid addition salts include inorganic acids such as hydrochloric acid, sulfuric acid, and phosphoric acid, as well as organic acids such as oxalic acid, maleic acid, methanesulfonic acid, succinic acid, and citric acid. Base addition salts can be prepared in situ during the final isolation and purification of the compounds of this disclosure by reacting the carboxylic acid moiety with a suitable base such as a pharmaceutically acceptable metal cation hydroxide, carbonate, or bicarbonate, or with ammonia or an organic primary, secondary, or tertiary amine. Examples of pharmaceutically acceptable salts include cations based on alkali metals and alkaline earth metals (e.g., sodium, lithium, potassium, calcium, magnesium, aluminum salts), as well as non-toxic ammonium, quaternary ammonium, and amine cations (such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, and ethylamine, but not limited to these). Other representative organic amines useful for base addition salt formation include diethylamine, ethylenediamine, ethanolamine, diethanolamine, and piperazine.

[0151] This disclosure also provides a kit comprising a protein kinase C inhibitor, a cMET inhibitor, and instructions for the combined use of the protein kinase C inhibitor and the cMET inhibitor in combination therapy. In other embodiments, the kit may comprise a pharmaceutical composition comprising the protein kinase C inhibitor and a pharmaceutically acceptable carrier, along with a probe or reference standard for assessing the presence of cMET. Such a kit may further comprise a cMET inhibitor. For example, the kit may comprise the protein kinase C inhibitor and the cMET inhibitor, which are pharmaceuticals described herein, together with instructions for their use in combination therapy or together with instructions for assessing the presence of cMET.

[0152] For example, the kit may include a protein kinase C inhibitor and a cMET inhibitor, which are formulated together or separately into one or more pharmaceutical compositions, each containing a pharmaceutically acceptable carrier. The kit may further include instructions on the use of the protein kinase C inhibitor and the cMET inhibitor together in combination therapy. The protein kinase C inhibitor is represented by formula II above.

[0153] This disclosure also provides a kit having a probe or reference standard for evaluating the presence of cMET. The probe may be a labeled antibody against cMET. The reference standard for evaluating the presence of cMET may include, for example, the amount of cMET derived from normal uveal cells.

[0154] For example, the kit may include a pharmaceutical composition comprising a protein kinase C inhibitor and a pharmaceutically acceptable carrier, and a probe or reference standard for evaluating the presence of cMET. The protein kinase C inhibitor can be represented by formula II as described herein.

[0155] This disclosure also provides various uses of the pharmaceuticals, kits, or compositions described herein for the preparation of pharmaceuticals for treating, preventing, or otherwise alleviating symptoms of proliferative disorders described herein, such as metastatic uveal melanoma or tumors having mutations in GNAQ or GNA11. The present invention also provides the use of pharmaceuticals, kits, or various compositions described herein for treating, preventing, or alleviating symptoms of proliferative disorders described herein, such as metastatic uveal melanoma or tumors having mutations in GNAQ or GNA11. For example, the use of pharmaceuticals, kits, or compositions comprising protein kinase C inhibitors and cMET inhibitors to reduce the proliferation, reduce growth, or reduce disease progression of metastatic uveal melanoma or tumors having mutations in GNAQ or GNA11.

[0156] The present invention also provides combinations of protein kinase C inhibitors and cMET inhibitors, for example as pharmaceuticals, kits, or compositions, for treating proliferative disorders described herein, such as metastatic uveal melanoma or tumors having mutations in GNAQ or GNA11.

[0157] dosage The compounds and their pharmaceutical compositions described herein may be administered in various forms according to the methods described herein, as is well known in the art, depending on the disorder being treated, as well as the patient's age, condition, and weight. As consistently recommended and required by medical authorities and government registration authorities for medicinal products, administration should ultimately be provided under the guidance and prescription of a primary physician with the insight, experience, and knowledge to control the patient's treatment.

[0158] The compounds can be formulated in a form suitable for the intended mode of administration. For example, if the compounds are to be administered orally, they may be formulated as tablets, capsules, granules, powders, or syrups; or, if administered parenterally, they may be formulated as injections (intravenous, intramuscular, or subcutaneous), infusions, or suppositories. For application via the ocular mucosa or other similar transmucosal routes, they may be formulated as drops or ointments.

[0159] The dosage may vary depending on the patient's symptoms, age, and weight, the patient's sex, the nature and severity of the disorder being treated or prevented, the route of administration, and the form of the drug. However, generally, for adult patients, a daily dose of 0.0001 to 2000 mg, preferably 0.001 to 1000 mg, more preferably 0.001 to 500 mg, particularly more preferably 0.001 to 250 mg, and most preferably 0.001 to 150 mg of the compound is recommended, which may be administered as a single dose or in divided doses. Alternatively, the daily dose can be given according to body weight, for example, 1 nanogram / kg (ng / kg) to 200 mg / kg, preferably 10 ng / kg to 100 mg / kg, more preferably 10 ng / kg to 10 mg / kg, and most preferably 10 ng / kg to 1 mg / kg. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form is generally the amount of the compound that produces the therapeutic effect.

[0160] The precise timing and / or amount of administration that yields the most effective results in terms of therapeutic efficacy in a given patient depends on the activity, pharmacokinetics, and bioavailability of the particular compound, the patient's physiological state (including age, sex, disease type and stage, overall physical condition, response to a given dose, and type of drug), and the route of administration. However, the above guidelines can be used as a basis for fine-tuning treatment, such as determining the optimal timing and / or amount of administration, and do not require anything more than routine experimentation consisting of monitoring the patient and adjusting the dose and / or timing.

[0161] The actual dose levels of compounds useful for applying the method of the present invention in the pharmaceutical compositions of the present invention can be varied to be effective in achieving the desired therapeutic response for a particular patient, composition, and mode of administration, and to obtain an amount of the active ingredient that is not toxic to the patient.

[0162] The concentration of compounds useful for applying the method of the present invention in a pharmaceutically acceptable mixture varies depending on several factors, including the dose of the compound administered, the pharmacokinetic properties of the compound used, and the route of administration.

[0163] In general, compositions useful for the application of the methods of the present invention may be provided for parenteral administration in aqueous solutions containing about 0.1 to 10% w / v of the compounds disclosed herein in other substances. Typical dose ranges are those given above, preferably about 0.001 to about 500 mg / kg body weight per day, and are given in 1 to 4 divided doses. Each divided dose may contain the same or different compounds of the present invention. The dose is an effective amount depending on several factors, including the patient's overall health, as well as the dosage form and route of administration of the selected compound.

[0164] Protein kinase C inhibitors may be administered in doses of approximately 1, 25, 50, 100, 150, 200, 250, 300, 400, 450, 500, 600, 700, or 800 mg, or may be present in pharmaceutical compositions, products, or kits. Protein kinase C inhibitors may be administered once, twice, or three times daily, or every other day. For example, protein kinase C inhibitors may be administered twice daily ("BID").

[0165] In various embodiments, protein kinase C inhibitors are administered according to a dosing regimen comprising a dosing cycle including a first dosing series, followed by a second dosing series, where (a) the first dosing series comprises compound (I) or a pharmaceutically acceptable salt thereof in a dose of approximately 200 mg BID, and (b) the second dosing series comprises compound (I) or a pharmaceutically acceptable salt thereof in a dose of approximately 400 mg BID. In various examples, the length of the first dosing series is 5 to 10 days, the length of the second dosing series is 18 to 23 days, and the length of the first dosing cycle comprising the first and second dosing series is 28 days. Further exemplary doses and dosing regimens are described in [reference].

[0166] In various embodiments, the protein kinase C inhibitor may be 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide administered in doses of approximately 50 mg BID, 100 mg BID, 150 mg BID, 200 mg BID, 250 mg BID, 300 mg BID, 350 mg BID, or 400 mg BID.

[0167] cMET inhibitors may be administered in doses of approximately 1, 10, 50, 100, 200, 300, 400, 500, 600, 700, or 800 mg, or may be present in pharmaceutical compositions, products, or kits. cMET inhibitors may be administered once, twice, or three times daily, or every two days. For example, a cMET inhibitor may be administered at 200, 250, 300, 350, or 400 mg twice daily. As a further example, a cMET inhibitor may be capmatinib administered orally at 400 mg twice daily. As another example, a cMET inhibitor may be crizotinib administered orally at 200 mg or 250 mg twice daily.

[0168] Unless otherwise specified, the weight or dose referred to herein for any particular compound in this disclosure (e.g., compound A) is the weight or dose of the compound itself, and not the weight or dose of its salt, and may vary to achieve the intended therapeutic effect. In various embodiments of the pharmaceuticals and compositions described herein, protein kinase C inhibitors and cMET inhibitors may exist in relative weight ratios of about 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, or about 1:10.

[0169] In various embodiments, patients have metastatic uveal melanoma or GNAQ / 11 tumors, and a protein kinase C inhibitor is administered in a dose that, when taken together with a cMET inhibitor, provides a greater therapeutic benefit than that achieved by either compound alone in the same amount. Such therapeutic benefit may be additive or greater than that achieved by either compound alone in the same amount. In some embodiments, therapeutic benefit is achieved by the use of the combination, while the use of either compound alone does not provide therapeutic benefit.

[0170] The methods, pharmaceuticals, and kits described herein can offer various therapeutic benefits in patients with metastatic uveal melanoma or GNAQ / 11 tumors. For example, in various embodiments, co-administration of a PKC inhibitor and a cMET inhibitor may be effective in reducing cell proliferation or growth of GNAQ / 11 tumors in patients. In various further embodiments, co-administration of a PKC inhibitor and a cMET inhibitor may be effective in reducing cell proliferation, growth, or progression of metastatic uveal melanoma. In further examples, the combination therapies described herein may be effective in treating metastatic disease or preventing the progression of metastatic disease.

[0171] For example, if a patient has a GNAQ / 11 tumor, a protein kinase C inhibitor, when taken together with a cMET inhibitor, may reduce the proliferation, growth, or metathesis of the GNAQ / 11 tumor, but the same amount of protein kinase C inhibitor administered alone may not reduce the proliferation, growth, or metathesis of the GNAQ / 11 tumor in the patient. Furthermore, if a patient has metastatic uveal melanoma, a protein kinase C inhibitor, when taken together with a cMET inhibitor, may reduce the proliferation, growth, or metathesis of the metastatic uveal melanoma, but the same amount of protein kinase C inhibitor administered alone may not reduce the proliferation, growth, or metathesis of the metastatic uveal melanoma in the patient. In another example, a patient has a GNAQ / 11 tumor, and a cMET inhibitor reduces the proliferation, growth, or metathesis of the GNAQ / 11 tumor when taken together with a protein kinase C inhibitor, but the same amount of cMET inhibitor administered alone does not reduce the proliferation, growth, or metathesis of the GNAQ / 11 tumor in the patient. In yet another example, a patient has metastatic uveal melanoma, and a cMET inhibitor reduces the proliferation, growth, or metathesis of the metastatic uveal melanoma when taken together with a protein kinase C inhibitor, but the same amount of cMET inhibitor administered alone does not reduce the proliferation, growth, or metathesis of the metastatic uveal melanoma in the patient.

[0172] In various embodiments, protein kinase C inhibitors, cMET inhibitors, or both are administered to patients without activating BRAF, KRAS, ERK, GSK3-beta, PIM2, or EGRF.

[0173] In various further embodiments, the co-administered PKC inhibitors and cMET inhibitors described herein are provided to a patient in a therapeutically effective dose to treat, prevent, or improve the symptoms of a disease. As used herein, the terms “effective dose” or “therapeutic dose” refer to the amount of a compound described herein (e.g., compound (A)) or a pharmaceutical composition containing a compound described herein that is administered in an amount sufficient to treat a disease. The appropriate “effective” dose in any individual case can be determined using techniques such as dose escalation studies. In relation to the administration of a drug, “effective dose” refers to the amount that produces a beneficial effect to the patient, e.g., improvement of symptoms, cure, reduction of disease burden, reduction of tumor burden or cell number, extension of lifespan, improvement of quality of life, or other effects that are generally perceived as positive by a physician familiar with the treatment of a particular type of disease or condition. A combination product, e.g., a fixed combination dosage form containing two or more compounds, has an effective dose corresponding to the total amount of the combination formulation, and each individual component also has an effective dose corresponding to the individual component used alone or in combination. [Examples]

[0174] The following embodiments are illustrative and do not limit the scope or content of this disclosure in any way. Active drug Crizotinib is available from Sigma-Aldrich (St. Louis, Missouri). Capmatinib is available from Novartis International AG (Basel, Switzerland). Compound A, corresponding to 3-amino-N-(3-(4-amino-4-methylpiperidine-1-yl)pyridine-2-yl)-6-(3-(trifluoromethyl)pyridine-2-yl)pyrazine-2-carboxamide, is available from IDEAYA Biosciences (South San Francisco, California). Compound A is shown in paragraph 0073 of the English specification and has the following structure.

[0175] [ka]

[0176] cell line MEL-202 and 92.1 cells can be purchased from Sigma-Aldrich (St. Louis, Missouri). MM28 cells can be obtained from the American Type Culture Collection (ATCC) (Manassas, Virginia) (accession number CRL-3295).

[0177] The 92.1 cell line originates from primary uveal melanoma, the MEL-202 cell line originates from uveal melanoma that recurred in the eye after prior irradiation, and MM28 originates from metastatic uveal melanoma cells. Each cell line tested corresponds to a GNAQ / 11 tumor, meaning each has one or more mutations in GNAQ or GNA11.

[0178] Primary uveal melanoma cell lines MEL-202 and 92.1 cells were maintained in 10% FBS / RPMI medium, and MM28 cells derived from metastatic uveal melanoma hepatoma were maintained in 20% FBS / RPMI medium in a humidified incubator at 37°C and 5% CO2.

[0179] Cell viability measurement: For all experiments, cell viability was measured using the Cell Titer-Glo Luminescent Cell Viability Assay kit (Promega) on day 3 for MEL-202 and 92.1 cells, and on day 5 for MM28 cells. Luminescence was read using a plate reader (TECAN). The average luminescence of cells treated with DMSO was set to 100%, and the viable cell percentage was calculated accordingly. Synergistic / antagonistic effects were analyzed using Combenefit software. Graphs for monotherapy with HGF or crizotinib were analyzed using GraphPad. Interactions between various combinations were evaluated using Bliss, HSA, and Loewe synergistic / antagonistic models. Uveal melanoma cells (92.1, MEL-202, MM28) were treated with hepatocyte growth factor (HGF) and compound A matrix in four different ways, each at varying titers. The synergistic, additive, or antagonistic effects of these treatments were determined using Bliss, HSA, and Loewe models. Cell viability was measured by CTG 3 / 5 days after treatment.

[0180] Examples 1 and 2 IDE196-HGF combination and IDE196-cMET inhibitor combination Treatment conditions for the IDE196-HGF combination: MEL-202 cells, MM28 cells, and 92.1 cells were seeded at 2500-5000 cells / well in 10% RPMI medium in 96 white wells with clear bottom plates 24 hours prior to treatment. After 24 hours, the medium was replaced with 2% FBS / RPMI medium. Furthermore, the cell lines were screened for sensitivity to PKC inhibitors, IDE196 alone, or in combination with hepatocyte growth factor (HGF) (Peprotech) using a TECAN® digital dispenser, increasing the dose according to Table 1.

[0181] [Table 1]

[0182] Treatment conditions for the IDE196-MET inhibitor combination: MEL-202 cells, MM28 cells, and 92.1 cells were seeded at 2500-5000 cells / well in 96 white wells with clear bottom plates 24 hours prior to treatment. After 24 hours, the medium was replaced with 2% FBS / RPMI medium containing 1.23 ng / ml HGF. Crizotinib or capmatinib and IDE196 were immediately dispensed using a TECAN digital dispenser, increasing the dose according to Table 2.

[0183] [Table 2]

[0184] Pharmacodynamic evaluation For all experiments, cells were placed in 10% RPMI medium in 10 cm³ 24 hours prior to treatment. 2 2.5 × 10⁶ 6 Cells were seeded. For the IDE196-HGF combination, cells were treated with a combination of IDE196 and HGF in 2% RPMI medium for 2 hours, according to Table 3. For the IDE196-MET inhibitor experiment, 24 hours after seeding, the medium was replaced with an appropriate inhibitor combination in 2% RPMI medium for 2 hours in the presence of 1.23–3.7 ng / ml of HGF, according to Table 4.

[0185] For all experiments, protein lysates were prepared by washing cells with ice-cold PBS and lysing the cells in RIPA buffer containing protease inhibitors and phosphatase inhibitors. Protein samples (15 ug / well) were separated on 4-20% Bis-Tris SDS gels and electroblotted onto nitrocellulose membranes using iBlot® (Invitrogen). The membranes were incubated in Superblock buffer at room temperature for 1 hour, followed by incubation overnight at 4°C with the primary antibody. The primary antibodies Phopho-Met(Tyr1234 / 1235)(3077S), cMET(3148S), Phopho-Akt(Ser473)(4060S), Akt(pan)(2920S), Phopho-MARCKS(Ser159 / 163)(11992S), MARCKS(5607S), PKCδ(9616S), Phopho-p44 / 42 MAPK(ERK1 / 2)(Thr202 / Tyr204)(4377S), and pRAS40 were obtained from Cell Signaling. The total ERK(ab184699) and PKC delta(phospho S299)(ab133456) antibodies were manufactured by Abcam. After washing three times with PBST, the membrane was incubated with HRP-conjugated secondary antibody at room temperature for 1 hour, washed three times with PBST, and detected using enhanced chemiluminescence (Biorad Chemidoc XRS).

[0186] [Table 3]

[0187] [Table 4]

[0188] Example 3 Increased presence of cMETs in uveal melanoma cells Place the cells in 10% RPMI medium for 10 cm 24 hours before treatment. 2 2.5 × 10⁶ 6Cells were seeded. For all experiments, protein lysates were prepared by washing the cells with ice-cold PBS and lysing the cells in RIPA buffer containing protease and phosphatase inhibitors. Protein samples (15 ug / well) were separated on 4-20% Bis-Tris SDS gel and electroblotted onto nitrocellulose membranes using iBlot (Invitrogen). The membranes were incubated in Superblock buffer at room temperature for 1 hour, followed by incubation overnight at 4°C with the primary antibody. The primary antibodies Phopho-Met(Tyr1234 / 1235)(3077S) and Met(3148S) were used. After washing three times with PBST, the membranes were incubated with HRP-conjugated secondary antibody at room temperature for 1 hour, washed three times with PBST, and detected using enhanced chemiluminescence (Biorad Chemidoc XRS). Cell lines MM28 and MEL-202 produced high levels of cMET, while cell line 92.1 produced low levels of cMET. These results are shown as a Western blot graph in Figure 4.

[0189] Example 4 Results of the IDE196-cMET inhibitor combination. The cell preparations described above were cultured as described to evaluate cell viability according to the combination of IDE196-cMET inhibitors after exposure to HGF cell culture.

[0190] Results of IDE196 sensitivity to exogenous HGF administration. Cell viability: To evaluate IDE196 sensitivity in the presence of HGF, we assessed changes in cell viability in MEL-202, 92.1, and MM28 cells with different combinations of IDE196 and HGF. MEL-202, 92.1, and MM28 cells showed dose-dependent inhibition of cell viability in response to IDE196 at EC50 = 250 nM, 113 nM, and 342 nM, respectively (N=3, Figures 1A, 2A, and 3A). Exposure of these cell lines to exogenous HGF significantly increased cell viability in MEL-202 and MM28 cells, but not in 92.1 cells. This is attributed to higher cMET expression in MEL-202 and MM28 cells compared to 92.1 cells (Figure 4). The combined results showed that in MEL-202 and MM28 cells, high antagonistic effects of IDE196-induced growth inhibition were observed at all IDE196 concentrations, with concentrations of IDE196 above 3.7 ng / mL HGF (Figures 1A and 3A). In 92.1 cells, only mild antagonistic effects were observed at HGF concentrations of 1.23–33.33 ng / mL (Figure 2A). The results were consistent using the HSA, Loewe, and Bliss algorithms included in the combined analysis.

[0191] PD analysis: cMET was activated by high concentrations of HGF in all three cell lines and was not reduced by IDE196 in the presence of HGF (Figures 1B, 2B, and 3B). Dose-dependent reductions of pMARCKS and pPKC-delta were observed after IDE196 monotherapy. The addition of HGF did not significantly alter pMARCKS and pPKC-delta, either alone or in combination with IDE196. In the absence of HGF, pERK was inhibited by IDE196 only at doses greater than 400 nM in all three cell lines. The addition of HGF induced pERK (MAPK signaling) as well as pAKT and pPRAS40 (PI3K signaling) in a dose-dependent manner in all three cell lines, and this was not inhibited by the addition of IDE196 in combination therapy. These results demonstrate that HGF activates cMET and promotes MAPK and PI3K signaling in these cell lines, and that this activation reduces the sensitivity of these cells to IDE196.

[0192] As shown in Figures 1A and 1B, HGF strongly antagonizes the effect of IDE196 on the viability of MEL-202 uveal melanoma cells. Figure 1A shows the cytotoxic effects of IDE196 and HGF individually and in combination on the viability of MEL-202 cells (N=3). Figure 1B shows PD analysis showing the effects of drug and HGF treatment on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways. These data indicate that experimentally induced cMET by exogenous HGF activates cMET as well as the MAPK and P13K signaling pathways, strongly antagonizing the antiproliferative activity of IDE196 in MEL-202 cells.

[0193] As shown in Figures 2A and 2B, HGF mildly antagonizes the effect of IDE196 on the viability of 92.1 uveal melanoma cells. Figure 2A shows the individual and combined cytotoxic effects of IDE196 and HGF on the viability of 92.1 cells (N=3). Figure 2B shows PD analysis showing the effects of drug and HGF treatment on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways. These data indicate that exogenous HGF does not activate cMET or the MAPK and PI3K signaling pathways, and only mildly antagonizes the antiproliferative activity of IDE196 in 92.1 uveal melanoma cells. The milder antagonism observed in this cell line is attributed to the relatively lower expression of cMET in the 92.1 cell line compared to the MEL-202 and MM28 cell lines (Figure 4).

[0194] As shown in Figures 3A and 3B, HGF strongly antagonizes the effect of IDE196 on the viability of MM28 uveal melanoma cells. Figure 3A shows the cytotoxic effects of IDE196 and HGF individually and in combination on the viability of MM28 cells (N=3). Figure 3B shows PD analysis showing the effects of drug and HGF treatment on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways. These data indicate that experimentally induced cMET by exogenous HGF activates the cMET, MAPK, and PI3K signaling pathways and strongly antagonizes the antiproliferative activity of IDE196 in MM28 cells.

[0195] As shown in Figure 4, the expression of total MET and phospho-MET in uveal melanoma cell lines is shown on Western blots. The faint lines for P-MET and T-MET for cell line 92.1 indicate that cell line 92.1 has much lower cMET expression compared to other cell lines. Compared to 92.1 cells, an increased presence of cMET is shown in MEL-202 and MM28 cells. These MEL-202 and MM28 cell lines with elevated cMET correlate with the data from Example 1, which showed greater antagonism against IDE196 (compound A) in the presence of HGF. Both MEL-202 and MM28 cells show increased total cMET concentration (T-MET). Since p-MET or activated cMET correlates with total cMET concentration, determining total cMET provides an assessment of the extent of the presence of elevated cMET involved in the PKC pathway.

[0196] In summary, these synergistic / antagonistic models show that MEL-202 and MM28 cells exhibit unexpectedly greater antagonism to IDE196 (compound A) compared to 92.1 cells, even when treated with the same concentration of HGF. Furthermore, this greater antagonism persists in MEL-202 and MM28 cells, maintaining their viability even as the dose of IDE196 (compound A) increases. Thus, the results indicate that HGF activates cMET, MAPK, and PI3K signaling in MM28 or MEL-202 cells that are not inhibited by IDE196 (compound A). MAPK is downstream of both PKC and cMET. Therefore, cMET counteracts the effect of IDE196 (compound A) on MAPK signaling. Both MM28 and MEL-202 cells exhibit similar signaling profiles in response to IDE196 (compound A) and HGF, despite having different cell viability profiles in response to the same drug.

[0197] In particular, HGF showed a relatively flat dose-response curve in 92.1 cells compared to MEL-202 and MM28 cells. The response of IDE196 (compound A) to the viability of 92.1 cells was relatively flat over HGF titration, whereas in MEL-202 and MM28 cells, the greater antagonism of HGF increased sharply at HGF concentrations that may be physiologically relevant to certain tumors. These results indicate that compound A can provide a potent antiproliferative effect in 92.1 cells, even in the presence of HGF, with or without dose shifts. Furthermore, for MEL-202 and MM28, these results indicate that the potent antagonism of HGF-stimulated cMETs cannot be significantly overcome by dose shifts to higher doses of PKC inhibitors, particularly at higher concentrations of HGF.

[0198] Physiological concentrations of HGF in the liver can range from 0.3 ng / ml (normal liver) to 1-16 ng / ml or higher in liver disease. Furthermore, certain uveal melanoma cells can mutate to secrete HGF. Thus, the stimulating effect of HGF represents the relevant effects in HGF-rich tissues, particularly in tumors that have metastasized to the liver, and in other tumors, such as tumors that can secrete HGF. In addition, different antagonistic effects against IDE196 (compound A) in different cell lines indicate the need for different therapeutic approaches due to the relative presence of cMETs, which may vary depending on the type of tumor cell being treated.

[0199] IDE196 (compound A) - cMET inhibitor combination Cell viability: To evaluate the sensitivity of uveal melanoma cell lines to IDE196-MET inhibitor combinations in the presence of HGF, changes in cell viability in MEL-202, 92.1, and MM28 cells were assessed with different combinations of IDE196 and the cMET inhibitors crizotinib and capmatinib. The combination results showed that the highest synergistic effects were observed in MEL-202 cells at crizotinib concentrations of 41.15–370.37 nM and IDE196 concentrations of 80–400 nM (Figure 5A). The capmatinib-IDE196 combination showed high synergistic effects across all capmatinib doses, starting from a dose of 80 nM IDE196 in MEL-202 cells (Figure 5B). In 92.1 cells, mild synergistic effects were observed at crizotinib concentrations of 1–3 μM and IDE196 concentrations of 0.8–10 μM (Figure 6A). However, capmatinib and IDE196 did not show synergistic effects in any dose combination in 92.1 cells (Figure 6B). In MM28 cells, high synergistic effects were observed at crizotinib concentrations of 0.123–10 μM and IDE196 concentrations of 0.8–10 μM (Figure 7A). The capmatinib-IDE196 combination showed high synergistic effects across all doses in MM28 cells (Figure 7B). Synergistic effects were observed across all algorithms used in the combine analysis (HSA, Loewe, and Bliss).

[0200] PD analysis: HGF induced pMET, pERK levels (MAPK pathway), and pAKT and pPRAS40 levels (PI3K pathway) in MEL-202 (Figure 5C) and MM28 (Figure 6C). Crizotinib, alone or in combination with IDE196, strongly inhibited the activation of all these analytes in these cell lines. In 92.1 cells (Figure 7C), basal pMET levels were lower compared to MEL-202 and MM28, but milder decreases in pERK, pAKT, and pPRAS40 were observed with higher concentrations of crizotinib alone or in combination with IDE196. In the presence of HGF, IDE196 alone did not inhibit pERK even at the highest concentration, and had no effect on pAKT or pPRAS40 in any cell line. Dose-dependent reductions in pMARCKS or pPKCδ were observed in the presence of HGF with IDE196 alone and in combination with crizotinib. These results demonstrate that the combination of crizotinib and IDE196 inhibits HGF-induced cMET activation and signaling via the MAPK and PI3K pathways. This data corresponds well to the high synergy observed in MEL-202 and MM28. In 92.1, the reduction in signaling via the MAPK and PI3K pathways was moderate, which correlated with the moderate synergy observed between crizotinib and IDE196.

[0201] Figures 5A, 5B, and 5C show the synergistic effects of crizotinib / capmatinib and IDE196 on MEL-202 cell viability, and relevant PD analyses for the crizotinib-IDE196 combination. Figure 5A shows the cytotoxic effects of IDE196 and crizotinib individually and in combination on MEL-202 cell viability (N=3). Figure 5B shows the cytotoxic effects of IDE196 and capmatinib individually and in combination on MEL-202 cell viability (N=3). Figure 5C shows PD analyses showing the effects of drug treatment on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways.

[0202] Figures 6A, 6B, and 6C show the extent of any synergistic effect (if any) between crizotinib / capmatinib and IDE196 on 92.1 cell viability, and relevant PD analyses for the crizotinib-IDE196 combination. Figure 6A shows the cytotoxic effects of IDE196 and crizotinib individually and in combination on 92.1 cell viability (N=3). Figure 6B shows the cytotoxic effects of IDE196 and capmatinib individually and in combination on 92.1 cell viability (N=3). Figure 6C shows PD analyses showing the effects of drug treatment on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways.

[0203] Figures 7A, 7B, and 7C show the synergistic effects of crizotinib / capmatinib and IDE196 on MM28 cell viability, and relevant PD analyses for the crizotinib-IDE196 combination. Figure 7A shows the cytotoxic effects of IDE196 and crizotinib individually and in combination on MM28 cell viability (N=3). Figure 7B shows the cytotoxic effects of IDE196 and capmatinib individually and in combination on MM28 cell viability (N=3). Figure 7C shows PD analyses showing the effects of drug treatment on analytes of the cMET, MAPK, PI3K, and PKC signaling pathways.

[0204] In summary, these data and various referenced figures show the following: a. Results of synergistic / antagonistic effects for each plate in dose matrices of compound A and crizotinib in MEL-202 cells containing HGF (HSA, Bliss, and Loewe); a summary of cell viability data across the same matrix, as well as dose-response curves for compound A and crizotinib, respectively.

[0205] b. Synergistic / antagonistic effects for each plate in dose matrices of compound A and capmatinib in MEL-202 cells containing HGF (HSA, Bliss, and Loewe); a summary of cell viability data across the same matrix, as well as dose-response curves for compound A and capmatinib, respectively.

[0206] c. Synergistic / antagonistic effects for each plate in a dose matrix of compound A and crizotinib in 92.1 cells containing HGF (HSA, Bliss, and Loewe); a summary of cell viability data across the same matrix, as well as dose-response curves for compound A and crizotinib, respectively.

[0207] d. Synergistic / antagonistic effects for each plate in dose matrices of compound A and capmatinib in 92.1 cells containing HGF (HSA, Bliss, and Loewe); a summary of cell viability data across the same matrix, as well as dose-response curves for compound A and capmatinib, respectively.

[0208] e. Results of synergistic / antagonistic effects for each plate in dose matrices of compound A and crizotinib in MM28 cells containing HGF (HSA, Bliss, and Loewe); a summary of cell viability data across the same matrix, as well as dose-response curves for compound A and crizotinib, respectively.

[0209] f. Synergistic / antagonistic effects for each plate in dose matrices of compound A and capmatinib in MM28 cells containing HGF (HSA, Bliss, and Loewe); a summary of cell viability data across the same matrix, as well as dose-response curves for compound A and capmatinib, respectively.

[0210] This paper provides pharmacodynamic results for various concentrations of compound A and crizotinib combinations in g.92.1 cells (containing exogenous HGF), MEL-202 cells (containing exogenous HGF), and MM28 cells (containing exogenous HGF).

[0211] Therefore, these results indicate that IDE196 (compound A) and the crizotinib / capmatinib combination work together in the presence of exogenous HGF to block MAPK signaling activation, and further block cMET, MARCKS, and PI3K signaling. Thus, the combination of a cMET inhibitor and a PKC inhibitor provides a synergistic antiproliferative effect against tumor cells with elevated cMET levels.

[0212] Example 5 Evaluation of the presence of cMETs in uveal melanoma tissue using RNA-Seq. IDE196 was clinically tested in humans in a monotherapy trial (NCT02601378). Pre- and post-administration biopsies were obtained from patients in this trial, and whole-transcriptome RNA-seq was performed on these biopsies to determine transcriptional changes at treatment. To assess MET expression / activity in baseline patient samples, RNA-seq data were aligned to GrCh38 using STAR, and then transcripts per million copies (TPM) were quantified using RSEM. cMET expression was then plotted across response categories, including patients with progressive disease (PD); stable disease (SD) for less than 6 months or more than 6 months; or partial response (PR). See Figure 8A.

[0213] In addition to MET gene expression, MET signature scores were also applied to these samples, as shown in Figure 8B. The scores were calculated from the RNA expression of genes correlated with cMET. First, the correlation between the gene-level TPM and cMET for each gene was calculated. After examining the distribution of correlations, genes with the highest subset of correlations: genes correlated with cMET >0.7 or <-0.7 were characterized as “module” genes. All intercorrelations of these module genes were calculated, and then the module genes were split into two modules: 1) an “up” module containing all module genes positively correlated with cMET, and 2) a “down” module containing all genes negatively correlated with cMET. The mean TPM across genes in each module was calculated for each of the up and down modules for each patient, providing two means per patient. These two means were then subtracted (up module - down module) to create a single final cMET signature score. Here, a high positive number indicates relatively higher cMET signaling activity in the patient compared to others in the cohort. To statistically test whether this score differed between response groups, ANOVA and ordinal regression were calculated (linear regression, where for each patient, the response was assigned a number representing where it falls in the appropriate order on the response scale; PD=4, SD<6 months=3, SD≧6 months=2, PR=1). The ANOVA was near significant (p=0.052), and the ordinal regression was very significant (p=0.0079). The box plot shows that cMET expression was highest in MUM (metastatic uveal melanoma) patients with progressive disease who did not respond to PKC monotherapy, and lowest in patients with a partial response to monotherapy, i.e., PR patients. Based on this analysis, since cMET activation was highest in patients with progressive disease and lowest in patients with a partial response in this human clinical trial, it supports the finding that elevated cMET activation / expression is a prognostic indicator of a less favorable response to IDE196 monotherapy.For example, patients who did not respond to PKC monotherapy generally had MET signature scores of 2 or higher, with most scores clustered around 3–5 relative to the study patient population, e.g., approximately 3, 3.5, 4, and 4.5. In contrast, patients who did respond to PKC monotherapy generally had scores of less than 2, with most scores clustered around 0–1.5 relative to the study patient population, e.g., approximately 0, 0.5, 1, and 1.5. The specific RNA transcripts that provided the MET signature score data shown in Figure 8B correspond to the genes shown in Table 5 below.

[0214] [Table 5-1]

[0215] [Table 5-2]

[0216] [Table 5-3]

[0217] Example 6 Evaluation of the presence of cMETs in uveal melanoma tissue using immunohistochemistry (IHC). cMET expression levels were evaluated in record-keeping tissue specimens using CONFIRM anti-total MET (SP44) rabbit monoclonal primary antibody (Ventana Medical Systems, Inc.; catalog number 790-4430) according to the manufacturer's instructions, which may include preparing tissue specimens by fixing in 10% buffered formalin, embedding in paraffin, and then dividing into serial 4 μm tissue sections for IHC examination. Staining was performed using a Ventana Benchmark XT instrument with CC1 standard antigen retrieval from Ventana Medical Systems (Oro Valley, Arizona). Incubation with the primary antibody was performed at 37°C for 16 minutes using a primary antibody concentration of 9.75 μg / mL. Specifically bound primary antibody was detected using the ultraView methodology (Ventana Medical Systems) with diaminobenzidine, and sections were counterstained with hematoxylin. Stain intensity was evaluated by two pathologists who were not informed of the individual patient's diagnosis. c-MET is primarily localized in the cytoplasm and membrane. Intensity is scored according to a four-level system: 0: no staining, 1+: weak, 2+: moderate, 3+: strong. The scoring system represents a composite scoring designed to evaluate both staining intensity (negative, weak, moderate, strong) and the percentage of tumor cells exhibiting each staining intensity. Tumors in which more than 50% of tumor cells exhibit moderate to strong staining intensity are predefined as cMET-positive before treatment assignment is deblinded.See Koeppen H, Yu W, Zha J, et al. Biomarker analyses from a placebo-controlled phase II study evaluating erlotinib±onartuzumab in advanced non-small cell lung cancer: MET expression levels are predictive of patient benefit. Clin Cancer Res. 2014;20(17):4488-4498.doi:10.1158 / 1078-0432.CCR-13-1836, Xu et al., BMC Cancer. 2015;15:6, and Spigel et al., Clin Lung Cancer. 2012;13(6):500, each of which is incorporated herein by reference in its entirety.

[0218] In the first scoring system, the H score evaluation is determined by the protocol corresponding to Xu et al., BMC Cancer. 2015;15:6 (which is incorporated herein by reference in its entirety). This scoring system uses staining intensity (0-3) and percentage of positive cells (0-100%). Each individual intensity level is multiplied by the percentage of cells, and all values ​​are added together to obtain a final IHC score ranging from 0 to 300. The final score is calculated from the scores of evaluations in membrane and cytoplasmic expression. Scores were evaluated for a panel of tissue from a population of patients with uveal melanoma, and the median was determined. This defines the cutoff values ​​for elevated and non-elevated cMETs. For example, in Xu et al., an H score of 20 is determined as the cutoff for tumors with elevated cMETs. Here, the cutoff is determined by analysis of tissue samples from a representative patient population. Subsequently, the scoring can be determined by comparing representative stained tissue samples for scores 0, 1, 2, or 3.

[0219] The second scoring system is based on scoring criteria adapted from Spigel et al., Clin Lung Cancer. 2012;13(6):500 (which is incorporated herein by reference in its entirety), which is clearly defined as having 50% or more positive tumor cells with moderate or strong intensity, i.e., 2+ or higher, for membrane or cytoplasmic and / or c-MET immunostaining.

[0220] The third scoring system is based on the table below, where "weak" staining is scaled to correspond to previous biopsies from the same tumor site in the same patient, and elevated cMET corresponds to a score of 2 or higher. In the fourth scoring system, based on the table below, "weak" staining corresponds to the level of staining seen in 92.1 cells, "moderate" staining corresponds to the intensity level of staining seen in MEL-202 and MM28 cells, and elevated cMET corresponds to a score of 2 or higher.

[0221] [Table 6]

[0222] Example 7 In vivo evaluation of the IDE196 (compound A)-cMET inhibitor combination. The combination of IDE196 and crizotinib will be evaluated in vivo using two potentially different humanized HGF orthotopic MUM models of the liver (see details below). These models allow for the direct transplantation of uveal melanoma tumor cells into the liver microenvironment, where they form tumors that can be measured in response to drug treatment. This approach has been used for both uveal melanoma PDX models and cell lines. See Sugase et al, Kageyama et al, and Cheng. In this study, we will use an orthotopic human uveal melanoma liver metastasis mouse model with tdTomato fluorescent proteins (tdUM001 and tdUM004) and the MM28 metastatic uveal melanoma cell line. We will quantify the in vivo efficacy of treatment with IDE196 and crizotinib as monotherapy and in combination, and evaluate in vivo tumor PD by observing the logarithmic fluorescence signal intensity (IVIS) and tumor size (CT scan) of the tumors. Kageyama K, Ohara M, Saito K, et al.Establishment of an orthotopic patient-derived xenograft mouse model using uveal melanoma hepatic metastasis.J Transl Med.2017;15(1):145.Published 2017 Jun 23.doi:10.1186 / s12967-017-1247-z, Cheng H, Terai Please refer to M, Kageyama K, et al.Paracrine Effect of NRG1 and HGF Drives Resistance to MEK Inhibitors in Metastatic Uveal Melanoma.Cancer Res.2015;75(13):2737-2748.doi:10.1158 / 0008-5472.CAN-15-0370.

[0223] Mouse model Since mouse HGF cannot activate human MET present in tumor cells, both mouse models show elevated hHGF levels driven by either a transgenic promoter (first model) or the introduction of an hHGF knock-in allele (second model). The first model exhibits a severe combined immunodeficiency spontaneous mutation (Prkdc scid We use hHGF-Tg SCID mice, which are homozygous for the hHGF-Tg transgene (commonly known as SCID) and hemizygous for the hHGF-Tg transgene (Tg(Mt1-HGF)#Gvw; the mouse metallothionein I promoter that drives human HGF expression). These mice have elevated serum hHGF titers, which are known to enhance the growth of MET-expressing human tumor xenografts derived from the lung, mammary gland, kidney, colon, stomach, and pancreas. See Zhang YW, Su Y, Lanning N, et al. Enhanced growth of human MET-expressing xenografts in a new strain of immunocompromised mice transgenic for human hepatocyte growth factor / scatter factor. Oncogene. 2005;24(1):101-106. doi:10.1038 / sj.onc.1208181.

[0224] The second model is Hgf tm1.1(HGF)Aveo NOD.scid.Il2Rγc possesses the "humanization" knock-in allele (hHGFki). nullThe study utilizes the NSG-hHGFki mouse, an animal species referred to as "NSG". Homozygous mice for hHGFki express only human-type HGF. These mice express human HGF instead of endogenous mouse HGF because the endogenous mouse promoter drives the expression of human hepatocyte growth factor (HGF). In homozygous hHGFki mice, HGF expression from the knock-in allele is observed in developing embryos, as well as in the liver, kidneys, and lungs of adults. Serreze DV,Chapman HD,Post CM,Johnson EA,Suarez-Pinzon WL,Rabinovitch A.Th1 to Th2 cytokine shifts in nonobese diabetic mice:sometimes an outcome,rather than the cause,of diabetes resistance elicited by immunostimulation.J Immunol.2001;166(2):1352-1359,Jangphattanont N,Sato H,Imamura See R,et al.Distinct Localization of Mature HGF from its Precursor Form in Developing and Repairing the Stomach.Int J Mol Sci.2019;20(12):2955.Published 2019 Jun 17.doi:10.3390 / ijms20122955.

[0225] Exemplary Embodiments The following exemplary embodiments are provided, and their numbering should not be construed as indicating a level of importance. These exemplary embodiments provide further details and descriptions of aspects of the invention, including methods of treatment, pharmaceutical composition products, combination compositions, and kits.

[0226] Embodiment 1 is a method of treatment, The selection of patients who have metastatic uveal melanoma or tumors with GNAQ mutations or GNA11 mutations ("GNAQ / 11 tumors"), This includes co-administering a cMET inhibitor and a protein kinase C inhibitor to the patient, Protein kinase C inhibitors, Formula II:

[0227] [ka]

[0228] or represented by a pharmaceutically acceptable salt thereof, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5bThese are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The method provides a method in which the alkoxy may be optionally substituted with one, two, three, or more halogens.

[0229] Embodiment 2 is a method for treating a patient with metastatic uveal melanoma, Select patients with metastatic uveal melanoma who have elevated cMETs determined by evaluating biopsy of the metastatic uveal melanoma, This includes co-administering a cMET inhibitor and a protein kinase C inhibitor to the patient, Protein kinase C inhibitors are expressed by formula II:

[0230] [ka]

[0231] or represented by a pharmaceutically acceptable salt thereof, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The method provides a method in which the alkoxy may be optionally substituted with one, two, three, or more halogens.

[0232] Embodiment 3 is a method for treating a patient having a tumor with a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"), Select patients with GNAQ / 11 tumors who have elevated cMETs, as determined by evaluating biopsy of the GNAQ / 11 tumor. This includes co-administering a cMET inhibitor and a protein kinase C inhibitor to the patient, Protein kinase C inhibitors, Formula II:

[0233] [ka]

[0234] or represented by a pharmaceutically acceptable salt thereof, in the formula X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The method provides a method in which the alkoxy may be optionally substituted with one, two, three, or more halogens.

[0235] Embodiment 4 provides the method of Embodiment 1, further comprising obtaining a biopsy of metastatic uveal melanoma or obtaining information related to the biopsy. Embodiment 5 provides the method of Embodiment 1, further comprising obtaining a biopsy of a GNAQ / 11 tumor or obtaining information related to the biopsy.

[0236] Embodiment 6 provides the method of Embodiment 4 or 5, further comprising evaluating the presence of cMET during biopsy. Embodiment 7 provides the method of Embodiment 6, further comprising selecting patients having elevated cMETs determined by evaluating a biopsy.

[0237] Embodiment 8 provides a method for any one of Embodiments 2 to 7, further comprising obtaining a biopsy from a patient. Embodiment 9 further includes obtaining multiple biopsies or obtaining information about the presence of cMETs in multiple biopsies, and in each case, the multiple biopsies are taken at separate time points, providing one of the methods of Embodiments 2 to 8.

[0238] Embodiment 10 provides one of the methods of Embodiments 2 to 9, wherein the biopsy is a tumor biopsy. Embodiment 11 provides one of the methods of Embodiments 2 to 10, wherein the biopsy is a tissue biopsy, a surgical excision, or both.

[0239] Embodiment 12 provides one of the methods from Embodiments 2 to 11, wherein the biopsy is obtained by core needle biopsy. Embodiment 13 provides one of the methods from Embodiments 2 to 12, which includes qualitatively or quantitatively measuring total cMET, phosphorylated cMET, non-phosphorylated cMET, or a combination thereof in a biopsy.

[0240] Embodiment 14 provides a method according to any one of Embodiments 2 to 13, which includes contacting an anti-cMET antibody with a biopsy sample. Embodiment 15 provides the method of Embodiment 14, wherein the anti-cMET antibody is phosphate-specific.

[0241] Embodiment 16 provides the method of Embodiment 14, wherein the anti-cMET antibody is not phosphate-specific. Embodiment 17 provides one of the methods described in Embodiments 14 to 16, wherein an anti-cMET antibody detects total cMET (T-MET).

[0242] Embodiment 18 provides one of the methods of Embodiments 14 to 17, wherein the anti-cMET antibody is optionally labeled or comprises a second reporter antibody capable of detecting the anti-cMET antibody.

[0243] Embodiment 19 provides one of the methods from Embodiments 2 to 13, which includes contacting a ligand for cMET with a biopsy sample. Embodiment 20 provides the method of Embodiment 19, wherein the ligand is a ligand for the extracellular domain of cMET.

[0244] Embodiment 21 provides the method of Embodiment 19, wherein the ligand is a ligand for the intracellular domain of cMET. Embodiment 22 provides one of the methods from Embodiments 19 to 21, wherein the ligand is an antibody.

[0245] Embodiment 23 provides one of the methods of Embodiments 18 to 22, wherein the ligand can optionally support or detect a separate colorimetric label, fluorescent label, radioactive label, or isotopic label.

[0246] Embodiment 24 provides a method according to any one of Embodiments 2 to 23, which includes determining the presence of elevated cMET in a biopsy by performing ELISA, Western blotting, IHC-F, IHC-P, immunocytochemistry, immunofluorescence, flow cytometry, mass cytometry, or immunoprecipitation.

[0247] Embodiment 25 provides one of the methods of Embodiments 2 to 24, wherein the biopsy is prepared in the form of a tissue section. Embodiment 26 provides the method of Embodiment 25, wherein tissue sections are fixed and embedded in paraffin.

[0248] Embodiment 27 provides one of the methods described in Embodiments 2 to 24 for freezing a biopsy. Embodiment 28 provides one of the methods of Embodiments 2 to 24, wherein the biopsy is prepared in the form of isolated cells, lysed cells, homogenates, cell fractions, or a combination thereof.

[0249] Embodiment 29 provides the method of Embodiment 28, wherein the biopsy is of cultured cells. Embodiment 30 provides one of the methods of Embodiments 2 to 29, comprising monitoring a patient over a period of time during which the presence of cMET is periodically evaluated in biopsies in multiple biopsies over that period.

[0250] Embodiment 31 is, Obtaining the aforementioned biopsy from the patient, To determine the concentration level of cMET during biopsy, The present invention provides any one of the embodiments 2 to 30, further comprising evaluating whether the determined level of cMET in the biopsy is equal to or greater than a predetermined level of cMET.

[0251] Embodiment 32 is, Obtain an initial biopsy from the patient's metastatic uveal melanoma or GNAQ / 11 tumor, and prepare a first tissue section therefrom. Obtain a second biopsy from healthy tissue of the patient that is the same type as the tissue containing metastatic uveal melanoma or GNAQ / 11 tumor, and prepare a second tissue section therefrom. Selectively, the labeled cMET antibody is brought into contact with the first tissue section, Selectively, the labeled cMET antibody is brought into contact with the second tissue section, The present invention provides any one of embodiments 2 to 31, further comprising determining the presence of elevated cMET in a second tissue section compared to a first tissue section.

[0252] Embodiment 33 is, Obtain a biopsy from a patient's metastatic uveal melanoma or GNAQ / 11 tumor and prepare cell preparations therefrom in the form of cell isolates, lysates, homogenates, fractions, or combinations thereof. To obtain healthy cells of the same type and morphology as the biopsy cell preparation, Selectively, the labeled cMET antibody is brought into contact with the biopsy cell preparation, Selectively contacting a labeled cMET antibody with a preparation of healthy cells, The present invention provides any one of the methods described in Embodiments 2 to 31, which includes determining the presence of elevated cMET in a biopsy cell preparation compared to a preparation of healthy cells.

[0253] Embodiment 34 provides a method according to any one of Embodiments 2 to 33, which includes evaluating the HGF concentration during a biopsy. Embodiment 35 optionally provides the method of Embodiment 31 or 32, wherein the labeled cMET antibody comprises a primary cMET antibody and a secondary labeled reporter antibody that binds to the primary cMET antibody.

[0254] Embodiment 36 provides one of the methods from Embodiments 2 to 34, wherein the biopsy has an HGF concentration of approximately 0.1 ng / ml to approximately 1,000 ng / ml. Embodiment 37 provides one of the methods from Embodiments 2 to 35, wherein the biopsy has an HGF concentration of approximately 1.0 ng / ml or higher.

[0255] Embodiment 38 provides a method according to any one of Embodiments 1 to 36, which includes evaluating gene mutations in a patient. Embodiment 39 provides a method according to any one of Embodiments 1 to 37, which includes evaluating gene mutations in patient cfDNA.

[0256] Embodiment 40 provides a method from any one of Embodiments 1 to 38, which includes obtaining a sample containing cfDNA. Embodiment 41 provides one of the methods from Embodiments 1 to 39, wherein the sample is a liquid sample.

[0257] Embodiment 42 provides one of the methods from Embodiments 1 to 40, wherein the liquid sample is a blood sample. Embodiment 43 provides one of the methods from Embodiments 2 to 36, which includes evaluating gene mutations in a biopsy.

[0258] Embodiment 44 provides the methods of Embodiments 37-42, in which gene mutations are evaluated by gene sequencing, Southern blotting, FISH, high-throughput sequencing, phage display, shotgun sequencing, PCR, or RT-PCR.

[0259] Embodiment 45 provides one of Embodiments 1 or 3-43, wherein the GNAQ mutation or GNA11 mutation is a gain-of-function mutation. Embodiment 46 provides one of Embodiments 1 or 3-44, wherein the GNAQ mutation or GNA11 mutation is a substitution of glutamine (Q209) at codon 209, a substitution of arginine (R183) at codon 183, or both.

[0260] Embodiment 47 provides one of Embodiments 1 or 3-45, wherein the GNAQ mutation or GNA11 mutation is other than a substitution of glutamine (Q209) at codon 209 or an arginine (R183) at codon 183, respectively.

[0261] Embodiment 48 provides a method according to any one of Embodiments 1 or 3 to 46, wherein the GNAQ / 11 tumor contains one or more mutations in GNAQ, namely Q209P, Q209L, Q209H, Q209K, Q209Y, or R183Q.

[0262] Embodiment 49 provides a method according to Embodiment 1 or any one of Embodiments 3 to 47, wherein the GNAQ / 11 tumor contains one or more Q209P, Q209L, Q209H, or Q209K mutations in GNA11.

[0263] Embodiment 50 provides one of Embodiments 1 or 3-48, wherein the GNAQ / 11 tumor is a solid tumor. Embodiment 51 provides one of Embodiments 1 or 3-49, wherein the GNAQ / 11 tumor is pancreatic, gastric, colorectal, uterine, cervical, bladder, hepatocellular carcinoma, head and neck, prostate, breast, lung adenocarcinoma, or cutaneous melanoma.

[0264] Embodiment 52 provides one of Embodiments 1 or 3-50, wherein the GNAQ / 11 tumor is pancreatic, gastric, colorectal, cervical, bladder, lung adenocarcinoma, or cutaneous melanoma. Embodiment 53 provides one of Embodiments 1 or 3-51, wherein the GNAQ / 11 tumor is a cutaneous melanoma.

[0265] Embodiment 54 provides one of Embodiments 1 or 3-52, wherein the GNAQ / 11 tumor is in the colon. Embodiment 55 provides one of Embodiments 1 or 3-53, wherein the GNAQ / 11 tumor is pancreatic.

[0266] Embodiment 56 provides a method for a GNAQ / 11 tumor located in the liver, according to any one of Embodiments 1 or 3-54. Embodiment 57 provides one of Embodiments 1 or 3-55, wherein the GNAQ / 11 tumor is a metastatic tumor.

[0267] Embodiment 58 provides one of Embodiments 1 or 3 to 56, wherein the GNAQ / 11 tumor is a metastatic tumor other than metastatic uveal melanoma. Embodiment 59 provides one of Embodiments 1 or 3 to 57, wherein the GNAQ / 11 tumor is metastatic and has metastasized to secondary sites having HGF concentrations of approximately 0.1 ng / ml to approximately 1,000 ng / ml.

[0268] Embodiment 60 provides the method of Embodiment 58, wherein the secondary site has an HGF concentration of approximately 1.0 ng / ml or higher. Embodiment 61 provides the method of Embodiment 58 or 59, wherein the secondary site is the liver.

[0269] Embodiment 62 provides one of Embodiments 1 or 3-49, wherein the GNAQ / 11 tumor is metastatic uveal melanoma. Embodiment 63 provides one of Embodiments 1 to 49 or 61, wherein the patient has uveal melanoma that has metastasized to or is at risk of metastasizing to a secondary site having an HGF concentration of approximately 0.1 ng / ml to approximately 1,000 ng / ml.

[0270] Embodiment 64 provides the method of Embodiment 62, wherein the secondary site has an HGF concentration of approximately 1.0 ng / ml or higher. Embodiment 65 provides the method of Embodiment 62 or 63, wherein the secondary site is the liver.

[0271] Embodiment 66 provides a method for which the GNAQ / 11 tumor is non-metastatic, either in one of Embodiments 1 or 3-55. Embodiment 67 provides a method in which the GNAQ / 11 tumor is a non-uveal tumor, either in one of Embodiments 1 or 3-57.

[0272] Embodiment 68 provides one of Embodiments 1, or any one of Embodiments 3-51, 53-57, wherein the GNAQ / 11 tumor is a non-melanocyte tumor. Embodiment 69 provides one of Embodiments 1 or 3-49, wherein GNAQ / 11 is a primary uveal melanoma.

[0273] Embodiment 70 provides a method in which GNAQ / 11 is a recurrent tumor, one of Embodiments 1 or 3-68. Embodiment 71 provides one of Embodiments 1 or 3-69, wherein the GNAQ / 11 tumor lacks one or more activating mutations in BRAF, KRAS, or EGRF.

[0274] Embodiment 72 provides one of Embodiments 1 or 3-70, wherein the GNAQ / 11 tumor has a low-load mutation in one or more of the following: BAP1, SF3B1, EIF1AX, TERT, BRAF, CDKN2A, NRAS, KRAS, or EGRF.

[0275] Embodiment 73 provides one of the methods of Embodiments 1 to 71, wherein a protein kinase C inhibitor, a cMET inhibitor, or both are administered to a patient without activating BRAF, KRAS, ERK, GSK3-beta, PIM2, or EGRF.

[0276] Embodiment 74 provides one of the methods of Embodiments 2 to 72, wherein the increase in the presence of cMET is relative to a predetermined level. Embodiment 75 provides one of the methods of Embodiments 2 to 73, wherein the increase in the presence of cMET is relative to the presence of cMET in healthy uveal cells in the same patient, or in healthy cells of the type from which the biopsy originates.

[0277] Embodiment 76 provides any one of Embodiments 2 to 73, wherein the increase in the presence of cMETs is relative to the average presence of cMETs in healthy uveal cells or healthy cells of the type from which the biopsy originates, in a patient population of patients with the same histological type of tumor.

[0278] Embodiment 77 provides one of the methods of Embodiments 2 to 73, wherein the increased presence of cMETs is relative to the presence of cMETs in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the biopsy origin in the same patient.

[0279] Embodiment 78 provides any one of Embodiments 2 to 73, wherein the increase in the presence of cMETs is relative to the average presence of cMETs in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the type from which the biopsy originates in a patient population of patients with the same histological type of tumor.

[0280] Embodiment 79 provides any one of Embodiments 2 to 77, wherein the increase in the presence of cMET is relative to a previously assessed level of cMET presence in a previous biopsy of the same cell type from the same patient.

[0281] Embodiment 80 provides one of the methods of Embodiments 2 to 78, wherein the increase in the presence of cMET is relative to the presence of cMET in standard primary uveal melanoma cells in a primary uveal melanoma cell culture.

[0282] Embodiment 81 provides any one of Embodiments 2 to 79 in which the presence of cMET is determined by a histological score of at least 2 on the IHC scale, or by the measured presence of cMET being at least 25% higher than the previously assessed level of the presence of cMET in a previous biopsy of the same cell type from the same patient, compared to the presence of cMET in healthy uveal cells or healthy cells of the type from which the biopsy originates in a patient population of patients with the same histological type of tumor, compared to the presence of cMET in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the type from which the biopsy originates in the same patient, compared to the average level of the presence of cMET in a previous biopsy of the same cell type from the same patient.

[0283] Embodiment 82 provides any one of Embodiments 2 to 80 in which the presence of cMET is determined by a histological score of at least 2 on the IHC scale, or by the measured presence of cMET being at least 50% higher than the previously assessed level of cMET presence in a previous biopsy of the same cell type from the same patient, compared to the presence of cMET in healthy uveal cells or healthy cells of the type from which the biopsy originates in a patient population of patients with the same histological type of tumor, compared to the presence of cMET in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the type from which the biopsy originates in the same patient, compared to the average level of cMET presence in a previous biopsy of the same cell type from the same patient.

[0284] Embodiment 83 provides one of Embodiments 2 to 81 in which the presence of cMET is determined by a histological score of at least 3 on the IHC scale, or by the measured presence of cMET being at least 100% higher than the presence of cMET in healthy uveal cells or healthy cells of the type from which the biopsy originates in the same patient, compared to the average presence of cMET in healthy uveal cells or healthy cells of the type from which the biopsy originates in a patient population of patients with the same histological type of tumor, compared to the presence of cMET in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the type from which the biopsy originates in the same patient, compared to the average presence of cMET in non-metastatic uveal melanoma cells or non-metastatic GNAQ / 11 tumor cells of the type from which the biopsy originates in a patient population of patients with the same histological type of tumor, compared to a previously assessed level of the presence of cMET in a previous biopsy of the same cell type from the same patient.

[0285] Embodiment 84 provides one of Embodiments 2 to 83, which includes determining the presence of elevated cMET during a biopsy by performing an evaluation of cMET expression using next-generation sequencing (NGS).

[0286] Embodiment 85 provides a method according to any one of Embodiments 2 to 84, which includes determining the presence of elevated cMET in a biopsy by performing RNA-seq.

[0287] Embodiment 86 provides one of Embodiments 2 to 85, wherein the presence of elevated cMET is determined by measuring the cMET mRNA expression level relative to the expression level of healthy uveal cells or healthy cells of the type derived from a biopsy in the same patient, relative to the average presence of cMET in healthy uveal cells or healthy cells of the type derived from a biopsy in a patient population of patients with the same histological type of tumor.

[0288] Embodiment 87 provides one of the methods from Embodiments 2 to 86, in which the presence of elevated cMET is determined by measuring cMET mRNA expression levels normalized by an internal expression reference.

[0289] Embodiment 88 provides the method of Embodiment 87, wherein the internal expression reference is a housekeeping gene. Embodiment 89 provides a method for reducing cell proliferation in metastatic uveal melanoma, according to any one of Embodiments 1, 2, or 4-88.

[0290] Embodiment 90 provides a method for reducing cell proliferation in GNAQ / 11 tumors, which is one of Embodiments 1 or 3-89. Embodiment 91 provides a method for reducing the growth of metastatic uveal melanoma, which is one of Embodiments 1, 2, or 4-90.

[0291] Embodiment 92 provides a method for reducing the growth of GNAQ / 11 tumors, either in Embodiment 1 or any one of Embodiments 3-91. Embodiment 93 provides one of the methods from Embodiments 1 to 92 for treating a metastatic disease or preventing the progression of a metastatic disease.

[0292] Embodiment 94 provides one of Embodiments 1 or 3-93, wherein a patient has a GNAQ / 11 tumor and a protein kinase C inhibitor, when administered together with a cMET inhibitor, reduces the proliferation, growth, or metathesis of the GNAQ / 11 tumor, but the same amount of protein kinase C inhibitor administered alone does not reduce the proliferation, growth, or metathesis of the GNAQ / 11 tumor in the patient.

[0293] Embodiment 95 provides a method according to any one of Embodiments 1, 2, or 4-94, wherein a patient has metastatic uveal melanoma, and a protein kinase C inhibitor, when administered together with a cMET inhibitor, reduces the proliferation, growth, or metathesis of the metastatic uveal melanoma, but the same amount of protein kinase C inhibitor administered alone does not reduce the proliferation, growth, or metathesis of the metastatic uveal melanoma in the patient.

[0294] Embodiment 96 provides one of Embodiments 1 or 3-95, wherein the patient has a GNAQ / 11 tumor and the cMET inhibitor, when administered together with a protein kinase C inhibitor, reduces the proliferation, growth, or metathesis of the GNAQ / 11 tumor, but the same amount of the cMET inhibitor administered alone does not reduce the proliferation, growth, or metathesis of the GNAQ / 11 tumor in the patient.

[0295] Embodiment 97 provides a method according to any one of Embodiments 1, 2, or 4-96, wherein the patient has metastatic uveal melanoma and the cMET inhibitor, when administered together with a protein kinase C inhibitor, reduces the proliferation, growth, or metathesis of the metastatic uveal melanoma, but the same amount of the cMET inhibitor administered alone does not reduce the proliferation, growth, or metathesis of the metastatic uveal melanoma in the patient.

[0296] Embodiment 98 provides one of the methods of Embodiments 2 to 97, wherein the biopsy shows the presence of elevated cMET, and the protein kinase C inhibitor and the cMET inhibitor are administered to the patient in amounts that provide therapeutic utility when taken together, but do not provide therapeutic utility when administered individually in the same amount.

[0297] Embodiment 99 provides one of the methods described in Embodiments 1 to 98, wherein the PKC inhibitor and the cMET inhibitor are administered simultaneously to the patient as a single composition or as separate compositions. Embodiment 100 provides one of the methods described in Embodiments 1 to 99, wherein a PKC inhibitor and a cMET inhibitor are administered to a patient as a single composition.

[0298] Embodiment 101 provides one of the methods of Embodiments 1 to 100, wherein the PKC inhibitor and the cMET inhibitor are administered to the patient as separate compositions at different times. Embodiment 102 provides the method of Embodiment 101, wherein the PKC inhibitor and the cMET inhibitor are administered sequentially in any order such that at least some portion of the first half-lives of each inhibitor overlap.

[0299] Embodiment 103 provides the method of Embodiment 102, wherein a PKC inhibitor is administered first, and a cMET inhibitor is administered second, within the half-life period of the PKC inhibitor. Embodiment 104 provides the method of Embodiment 102, wherein a cMET inhibitor is administered first, and a PKC inhibitor is administered second, within the half-life period of the cMET inhibitor.

[0300] Embodiment 105 is a method for treating a patient having metastatic uveal melanoma or a tumor having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"), To evaluate the presence or activity of cMET in biopsies of metastatic uveal melanoma or GNAQ / 11 tumors, If the presence or activity of cMET is elevated in metastatic uveal melanoma or GNAQ / 11 tumors, co-administration of a cMET inhibitor and a protein kinase C inhibitor to the patient is recommended. If the presence of cMET is not elevated in metastatic uveal melanoma or GNAQ / 11 tumors, the patient may be administered a protein kinase C inhibitor without a cMET inhibitor, including: Protein kinase C inhibitors are expressed by formula II:

[0301] [ka]

[0302] or represented by a pharmaceutically acceptable salt thereof, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The method provides a method in which the alkoxy may be optionally substituted with one, two, three, or more halogens.

[0303] Embodiment 106 is one in which X is N and R 2 , R 3 , and R 4 The present invention provides one of the embodiments 1 to 105, wherein each of the elements is independently either H or a halo. Embodiment 107 is R 2 , R 3 , and R 4 The present invention provides one of the methods from Embodiments 1 to 106, wherein each of these is H.

[0304] Embodiment 108 is R 5 The present invention provides one of the methods from Embodiments 1 to 107, wherein is H or CH3. Embodiment 109 is R 5 The present invention provides one of the embodiments 1 to 108, wherein CH3 is provided.

[0305] Embodiment 110 is R 6 and R 7 However, each is independent of H, Haro, and C. 1~3 Haloalkyl, C 1~3 Haloalkoxy, C 3~7 The present invention provides one of the methods from Embodiments 1 to 109, selected from cycloalkyl, morpholino, piperidinyl, and piperazinyl.

[0306] Embodiment 111 is R 6 and R 7 One of them is C 1~3 Haloalkyl, C 1~3 Haloalkoxy, C 3~7 These are cycloalkyl, morpholino, piperidinyl, and piperazinyl, and R 6 and R 7 The present invention provides one of the methods of Embodiments 1 to 110, wherein one of the is H.

[0307] Embodiment 112 is R 6 and R 7 One of them is trifluoromethyl or trifluoromethoxy, R 6 and R 7 The present invention provides one of the methods from Embodiments 1 to 111, wherein one of the is H.

[0308] Embodiment 113 is R 5a , R 5b , R 5c , and R 5d The present invention provides one of the methods from Embodiments 1 to 112, wherein each of these is H. Embodiment 114 describes a protein kinase C inhibitor,

[0309] [ka]

[0310] The present invention provides one of the methods described in Embodiments 1 to 113, which is either a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable salt thereof. Embodiment 115 describes a protein kinase C inhibitor,

[0311] [ka]

[0312] The present invention provides one of the methods described in Embodiments 1 to 114, which is either a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable salt thereof. Embodiment 116 provides one of the methods of Embodiments 1 to 115, wherein the cMET inhibitor is an inhibitor of intracellular ATP-cMET ligand-receptor binding.

[0313] Embodiment 117 provides one of the methods described in Embodiments 1 to 116, wherein the cMET inhibitor is an ATP-competitive small molecule cMET inhibitor. Embodiment 118 provides one of the methods of Embodiments 1 to 117, wherein the cMET inhibitor is selected from the group consisting of crizotinib, capmatinib, cabozantinib, tivantinib, and any combination thereof.

[0314] Embodiment 119 provides one of the methods from Embodiments 1 to 118, wherein the cMET inhibitor is capmatinib. Embodiment 120 provides one of the methods of Embodiments 1 to 118, wherein the cMET inhibitor also has activity as an ALK inhibitor, a RAS1 inhibitor, or both.

[0315] Embodiment 121 provides one of the methods of Embodiments 1 to 118, wherein the cMET inhibitor is crizotinib. Embodiment 122 provides one of the methods described in Embodiments 1 to 121, wherein the protein kinase C inhibitor is administered at a dose of approximately 100 mg to approximately 1000 mg / day.

[0316] Embodiment 123 provides a method according to any one of Embodiments 1 to 122, wherein the protein kinase C inhibitor is administered in a dose of approximately 10 mg to approximately 400 mg BID. Embodiment 124 provides one of Embodiments 1 to 123, wherein the protein kinase C inhibitor is administered in doses of approximately 50 mg BID, 100 mg BID, 150 mg BID, 200 mg BID, 250 mg BID, 300 mg BID, 350 mg BID, or 400 mg BID.

[0317] Embodiment 125 describes a drug regimen in which a protein kinase C inhibitor is administered according to a drug regimen that includes a drug cycle comprising a first drug series and a subsequent second drug series. (a) The first series of medications contains compound (I) or a pharmaceutically acceptable salt thereof in a dose of approximately 200 mg BID. (b) A second dosing series provides any one of Embodiments 1 to 124, comprising a dose of compound (I) or a pharmaceutically acceptable salt thereof in a dose of approximately 400 mg BID.

[0318] Embodiment 126 provides the method of Embodiment 125, wherein the length of the first drug series is 5 to 10 days, the length of the second drug series is 18 to 23 days, and the length of the first drug cycle, including the first and second drug series, is 28 days.

[0319] Embodiment 127 is a pharmaceutical product comprising a protein kinase C inhibitor, a cMET inhibitor, and a pharmaceutically acceptable carrier, wherein the protein kinase C inhibitor is of formula II:

[0320] [ka]

[0321] or represented by a pharmaceutically acceptable salt thereof, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5dThese are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The present invention provides a pharmaceutical product in which the alkoxy may be optionally substituted with one, two, three, or more halogens.

[0322] Embodiment 128 provides the pharmaceutical product of Embodiment 127, wherein a protein kinase C inhibitor and a cMET inhibitor are formulated together in a single unit dosage form containing a pharmaceutically acceptable carrier.

[0323] Embodiment 129 provides the pharmaceutical product of Embodiment 128, wherein the protein kinase C inhibitor and the cMET inhibitor are formulated into two distinct unit dosage forms, each having a pharmaceutically acceptable carrier.

[0324] Embodiment 130 is, One or more pharmaceutical compositions, each containing a pharmaceutically acceptable carrier, contain, together or separately, formulated protein kinase C inhibitors and cMET inhibitors. A kit including instructions on the use of a protein kinase C inhibitor and a cMET inhibitor together in combination therapy, Protein kinase C inhibitors are expressed by formula II:

[0325] [ka]

[0326] or represented by its pharmaceutically acceptable salt, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5a and R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The kit provides an alkoxy that may be optionally substituted with one, two, three, or more halogens.

[0327] Embodiment 131 is, A pharmaceutical composition comprising a protein kinase C inhibitor and a pharmaceutically acceptable carrier, A kit comprising a probe or reference standard for evaluating the presence of cMET, Protein kinase C inhibitors are expressed by formula II:

[0328] [ka]

[0329] or represented by its pharmaceutically acceptable salt, in the formula, X is either N or CR; R, R 2 , R 3 , and R 4 These are H, independently of each other. 2 H, halogen, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5 H, 2 H, -CH3, -CH2F, -CHF2, -CF3, -CH2OH, and C 2~3 Selected from the group consisting of alkyl groups, C 2~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (optionally substituted with one or more halogens); R 5aand R 5b These are H, independently of each other. 2 H and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5a and R 5b They combine to form methylene or ethylene crosslinking groups; R 5c and R 5d These are H, independently of each other. 2 H, fluorine, hydroxyl, C 1~3 Alkoxy, and C 1~3 Selected from the group consisting of alkyl groups, C 1~3 Alkyls can optionally contain fluorine, hydroxyl, and C. 1~3 They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys; or R 5c and R 5d They combine to form methylene, ethylene, or -CH2-O-crosslinking groups; R 6 , R 7 , and R 8 These are H, independently of each other. 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 Selected from the group consisting of cycloalkyls and 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S, C 1~3 The alkoxy may optionally be substituted with one, two, three, or more halogens, C 1~3 Alkyls can optionally be hydroxyl, halogen, and C. 1~3They may be substituted with one, two, three, or more substituents independently selected from the group consisting of alkoxys (which are optionally substituted with one or more halogens); or R 6 and R 8 They optionally form a partially unsaturated carbon bicyclic or heterobicyclic ring with the heteroaryl ring to which they are bonded, and the carbon bicyclic or heterobicyclic ring optionally forms 2 H, halogen, C 1~3 Alkyl, C 1~3 Alkoxy, C 3~7 They may also be substituted with cycloalkyl groups and one, two, or three groups independently selected from the group consisting of 4- to 7-membered heterocyclines having one, two, or three heteroatoms independently selected from the group consisting of N, O, and S; C 1~3 Alkyl and C 1~3 The kit provides an alkoxy that may be optionally substituted with one, two, three, or more halogens.

[0330] Embodiment 132 provides a method, pharmaceutical, or kit according to any one of Embodiments 1 to 131, wherein the protein kinase C inhibitor has at least 1000 times greater inhibitory activity against protein kinase C isoforms delta, epsilon, eta, or theta than against kinases other than protein kinase C quinome.

[0331] Embodiment 133 provides a method, pharmaceutical, or kit according to any one of Embodiments 1 to 131, wherein the protein kinase C inhibitor has at least 1000 times lower inhibitory activity against GSK3-beta than against the protein kinase C isoform theta.

[0332] Embodiment 134 provides one of the methods, pharmaceuticals, or kits from Embodiments 1 to 131, wherein the protein kinase C inhibitor has greater inhibitory activity against protein kinase C isoforms delta, epsilon, eta, and theta, respectively, than against protein kinase C isoforms alpha, beta 1, beta 2, or gamma.

[0333] Embodiment 135 provides one of the methods, pharmaceuticals, or kits from Embodiments 1 to 131, wherein the protein kinase C inhibitor exhibits at least 5-fold lower inhibitory activity against protein kinase C isoforms alpha, beta 1, beta 2, or gamma than against protein kinase C isoforms delta, epsilon, eta, or theta.

[0334] Embodiment 136 provides one of the methods, pharmaceuticals, or kits from Embodiments 1 to 131, wherein the protein kinase C inhibitor exhibits at least 20 times lower inhibitory activity against protein kinase C isoform beta-1 than against protein kinase C isoform theta.

[0335] Embodiment 137 provides a method, pharmaceutical, or kit according to any one of Embodiments 1 to 131, wherein the protein kinase C inhibitor has an IC50 less than 50 nM for one or more protein kinase C isoforms delta, epsilon, eta, and theta.

[0336] Embodiment 138 provides any one of Embodiments 1 to 131, a method, pharmaceutical, or kit, wherein the protein kinase C inhibitor has an IC50 less than 5 nM with respect to one or more protein kinase C isoforms delta, epsilon, eta, and theta.

[0337] Embodiment 139 provides a method, pharmaceutical, or kit according to any one of Embodiments 1 to 131, wherein the protein kinase C inhibitor has an IC50 greater than 50 nM with respect to protein kinase C isoform beta-1, beta-2, or both.

[0338] Embodiment 140 provides any one of Embodiments 1 to 131, a method, pharmaceutical, or kit, wherein the protein kinase C inhibitor has an IC50 greater than 5,000 nM with respect to PIM2, GSK3β, or both.

[0339] Embodiment 141 provides the method of Embodiment 85, which includes performing RNA-Seq to evaluate the MET signature score based on the calculation of RNA expression from one or more genes that correlate with cMET.

[0340] Embodiment 142 provides the method of Embodiment 141, in which the elevation of cMET presence is determined by measuring the patient's MET signature score relative to the patient's MET signature score in a patient population.

[0341] Embodiment 143 provides the method of Embodiment 142, wherein the MET signature score in the patient population is ranked according to the response group, preferably 4 = progressive disease, 3 = stable disease for less than 6 months, 2 = stable disease for 6 months or more, and 1 = partial response to monotherapy, and the patient's MET signature score is 2 or greater, 3 or greater, or about 3, 3.5, 4, or 4.5.

[0342] Embodiment 144 provides one of the pharmaceutical products or kits from Embodiments 127 to 140 for use in the treatment of patients with metastatic uveal melanoma, or for use in the treatment of tumors having a GNAQ mutation or a GNA11 mutation ("GNAQ / 11 tumor"), or both.

[0343] Embodiment 145 provides one of the pharmaceuticals or kits from Embodiments 127 to 140 for use in any one of Embodiments 1 to 126 or 141 to 143.

[0344] Embodiment 146 provides the use of cMET inhibitors and protein kinase C inhibitors, or the pharmaceuticals of Embodiments 127-129, for the manufacture of a pharmaceutical for treating a patient according to any one of Embodiments 1-126 or 141-143.

[0345] Embodiment 147 provides a method, pharmaceutical product, or kit of any one or a combination of the embodiments described herein, and is optionally configured such that any or all of the elements or options described herein are available for use or selection.

[0346] Overview The present invention has been described in conjunction with its detailed description, but the foregoing description is intended to be illustrative and does not limit the scope of the invention as defined by the appended claims. Accordingly, it will be understood from the foregoing that certain non-limiting embodiments of the invention are described herein for illustrative purposes, but various modifications can be made without departing from the spirit and scope of the invention. Other aspects, advantages, and modifications are within the following claims, and the invention is not limited except as provided for by the appended claims.

[0347] The present invention has been described broadly and generally in this specification. Each of the narrower species and subgenera classifications included in the general disclosure also forms part of the present invention. This includes the general description of the present invention having conditions or negative limitations that remove any patient substance from a genus, whether or not the deleted substance is specifically described herein.

[0348] The terms and expressions used are for illustrative purposes only, not limiting ones, and in using such terms and expressions there is no intention to exclude any equivalents of the exhibited and described features or any part thereof, but it is recognized that various modifications are possible within the scope of the claimed invention. Accordingly, although the present invention is specifically disclosed by various non-limiting embodiments and / or preferred non-limiting embodiments and any features, it will be understood that any modifications and variations of the concepts disclosed herein, which may be reclassified by those skilled in the art, are considered to be within the scope of the invention as defined by the appended claims.

[0349] All patents, publications, scientific papers, websites, and other documents, as well as any references or references herein, demonstrate the level of skill of those skilled in the art relating to the present invention, and each such referenced document and material is incorporated herein by reference to the same extent as it is incorporated verbatim and presented herein in its entirety. The right to physically incorporate any and all material and information from any such patent, publication, scientific paper, website, electronically available information, textbook, or other referenced material or document is reserved herein.

[0350] All references cited herein are incorporated herein by reference as if they were contained in their entirety.

Claims

1. A pharmaceutical composition for use in a method of treating metastatic uveal melanoma in a patient, wherein the pharmaceutical composition comprises a protein kinase C (PKC) inhibitor. The protein kinase C inhibitor of the pharmaceutical composition is administered to the patient in combination with a cMET inhibitor. The protein kinase C inhibitor 【Chemistry 1】 or a pharmaceutically acceptable salt thereof, A pharmaceutical composition wherein the cMET inhibitor is (i) crizotinib or a pharmaceutically acceptable salt thereof, or (ii) capmatinib or a pharmaceutically acceptable salt thereof.

2. The pharmaceutical composition according to claim 1, wherein the method of treatment includes obtaining a biopsy of the metastatic uveal melanoma or obtaining information regarding the biopsy.

3. The pharmaceutical composition according to claim 2, wherein the method of treatment includes evaluating the presence of cMET in the biopsy.

4. The pharmaceutical composition according to claim 3, wherein the method of treatment comprises selecting a patient having elevated cMET, as determined by evaluating the biopsy.

5. The pharmaceutical composition according to any one of claims 2 to 4, wherein the method of treatment comprises qualitatively or quantitatively measuring total cMET, phosphorylated cMET, non-phosphorylated cMET, or a combination thereof in the biopsy.

6. The pharmaceutical composition according to any one of claims 2 to 5, wherein the method of treatment comprises determining the presence of elevated cMET in the biopsy by performing ELISA, Western blotting, IHC-F, IHC-P, immunocytochemistry, immunofluorescence, flow cytometry, mass cytometry, immunoprecipitation, or cMET RNA transcriptome analysis.

7. The aforementioned treatment method is Obtaining the biopsy from the aforementioned patient, To determine the concentration level of cMET in the biopsy, To evaluate whether the determined cMET level in the biopsy is above a predetermined level of cMET, A pharmaceutical composition according to any one of claims 2 to 6, comprising:

8. The pharmaceutical composition according to any one of claims 1 to 7, wherein the method of treatment includes evaluating gene mutations in the patient.

9. The protein kinase C inhibitor described above is 【Chemistry 2】 The pharmaceutical composition according to any one of claims 1 to 8.

10. The pharmaceutical composition according to any one of claims 1 to 9, wherein the cMET inhibitor is crizotinib.

11. The pharmaceutical composition according to any one of claims 1 to 9, wherein the cMET inhibitor is capmatinib.

12. The pharmaceutical composition according to any one of claims 1 to 11, wherein the protein kinase C inhibitor and the cMET inhibitor are administered simultaneously as separate compositions.

13. The pharmaceutical composition according to any one of claims 1 to 11, wherein the protein kinase C inhibitor and the cMET inhibitor are administered sequentially as separate compositions.

14. The pharmaceutical composition according to any one of claims 1 to 13, wherein the protein kinase C inhibitor and the cMET inhibitor are for oral administration.

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