Treating Cancer with RAF Inhibitors

Abstract

Provided herein is a composition and method for treating cancer.The composition comprises a RAF inhibitor.Some embodiments include combination therapy using the RAF inhibitor with at least one tumor therapeutic agent.

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims the benefit of U.S. Patent Application No. 63 / 178,922, filed April 23, 2021, which is incorporated by reference in its entirety herein. [Background technology]

[0002] RAF kinases function in the Ras-RAF-MEK-ERK mitogen-activated protein kinase (MAPK) pathway (also known as the MAPK / ERK pathway) by phosphorylating and activating MEK, and ultimately, MAPK-dependent activation of transcriptional programs promotes cell proliferation and survival. Deregulation of MAPK activity occurs frequently in tumors. Thus, therapies targeting RAF kinase activity are desirable for use in the treatment of cancer and other disorders characterized by aberrant MAPK / ERK pathway signaling. Summary of the Invention

[0003] One embodiment provides a method of treating cancer in a patient in need thereof, comprising administering to the patient (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof.

[0004] One embodiment provides a method of treating cancer in a patient in need thereof, comprising administering to the patient a pharmaceutical composition comprising (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, and at least one pharma- ceutically acceptable excipient.

[0005] One embodiment is a method of treating cancer in a patient in need of such treatment, comprising: (a) a pharmaceutical composition comprising (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof; (b) at least one oncology therapeutic agent selected from a MEK inhibitor, an immune checkpoint inhibitor, a CDK inhibitor, an EGFR kinase inhibitor, an EGFR antibody, an EGFR PROTAC therapeutic agent, an FGFR inhibitor, an SOS1 inhibitor, an SHP2 inhibitor, a KRAS inhibitor, a taxane, a topoisomerase inhibitor, an ERK inhibitor, a platinum-based chemotherapy agent, folinic acid, 5-fluorouracil, or an anti-VEGF therapeutic agent; to a patient. [Brief description of the drawings]

[0006] [Figure 1A] 1A-1C illustrate A-375 (class I BRAF mutant melanoma) xenograft tumor volume (FIG. 1A), mouse body weight (FIG. 1B), and percent change in xenograft tumor volume at the end of the study relative to baseline (FIG. 1C) following treatment with Compound 1. [Figure 1B] 1A-1C illustrate A-375 (class I BRAF mutant melanoma) xenograft tumor volume (FIG. 1A), mouse body weight (FIG. 1B), and percent change in xenograft tumor volume at the end of the study relative to baseline (FIG. 1C) following treatment with Compound 1. [Figure 1C] 1A-1C illustrate A-375 (class I BRAF mutant melanoma) xenograft tumor volume (FIG. 1A), mouse body weight (FIG. 1B), and percent change in xenograft tumor volume at the end of the study relative to baseline (FIG. 1C) following treatment with Compound 1. [Figure 2A]2A-2D illustrate BxPC-3 (class II BRAF-mutated PDAC) and WM3629 (class II IBRAF-mutated melanoma) xenograft tumor volumes (FIG. 2A, FIG. 2C) and mouse body weights (FIG. 2B, FIG. 2D) during treatment with Compound 1. [Figure 2B] 2A-2D illustrate BxPC-3 (class II BRAF-mutated PDAC) and WM3629 (class II IBRAF-mutated melanoma) xenograft tumor volumes (FIG. 2A, FIG. 2C) and mouse body weights (FIG. 2B, FIG. 2D) during treatment with Compound 1. [Figure 2C] 2A-2D illustrate BxPC-3 (class II BRAF-mutated PDAC) and WM3629 (class II IBRAF-mutated melanoma) xenograft tumor volumes (FIG. 2A, FIG. 2C) and mouse body weights (FIG. 2B, FIG. 2D) during treatment with Compound 1. [Figure 2D] 2A-2D illustrate BxPC-3 (class II BRAF-mutated PDAC) and WM3629 (class II IBRAF-mutated melanoma) xenograft tumor volumes (FIG. 2A, FIG. 2C) and mouse body weights (FIG. 2B, FIG. 2D) during treatment with Compound 1. [Figure 3A] 3A and 3B illustrate waterfall plots of the individual BxPC-3 and WM3629 tumor xenograft responses shown in FIG. 3A and FIG. 3B, respectively. [Figure 3B] 3A and 3B illustrate waterfall plots of the individual BxPC-3 and WM3629 tumor xenograft responses shown in FIG. 3A and FIG. 3B, respectively. [Figure 4A] 4A-4C illustrate WM3629 tumor xenograft responses to combination therapy of Compound 1 and binimetinib as described in Example 3C. [Figure 4B] 4A-4C illustrate WM3629 tumor xenograft responses to combination therapy of Compound 1 and binimetinib as described in Example 3C. [Figure 4C]4A-4C illustrate WM3629 tumor xenograft responses to combination therapy of Compound 1 and binimetinib as described in Example 3C. [Diagram 5] FIG. 5 illustrates the cellular activity of Compound 1 in combination with binimetinib in NRAS mutant melanoma as described in Example 4. [Figure 6] FIG. 6 illustrates the antitumor activity of Compound 1 in combination with binimetinib in the human SK-MEL-2 (NRAS Q61R mutant) melanoma xenograft model described in Example 5. [Figure 7] FIG. 7 illustrates the antitumor activity of Compound 1 in combination with binimetinib in the human SK-MEL-2 (NRAS Q61R mutant) melanoma xenograft model described in Example 5. [Figure 8A] 8A-8C illustrate the antitumor activity of Compound 1 in combination with binimetinib in the WM3629 melanoma xenograft model described in Example 6. [Figure 8B] 8A-8C illustrate the antitumor activity of Compound 1 in combination with binimetinib in the WM3629 melanoma xenograft model described in Example 6. [Figure 8C] 8A-8C illustrate the antitumor activity of Compound 1 in combination with binimetinib in the WM3629 melanoma xenograft model described in Example 6. [Figure 9A] 9A-9C illustrate the antitumor activity of Compound 1 in the NSCLC xenograft model described in Example 7. [Figure 9B] 9A-9C illustrate the antitumor activity of Compound 1 in the NSCLC xenograft model described in Example 7. [Figure 9C] 9A-9C illustrate the antitumor activity of Compound 1 in the NSCLC xenograft model described in Example 7.

[0007] (Incorporated by reference) All publications, patents, and patent applications mentioned herein are hereby incorporated by reference for the specific purposes identified herein.

[0008] Detailed Description of the Invention Specific Terms As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, a reference to an "agent" includes a plurality of such agents, a reference to a "cell" includes a reference to one or more cells (or cells) and equivalents thereof known to those of skill in the art, and so forth. When ranges for physical properties, such as molecular weight, or chemical properties, such as chemical formulas, are used herein, it is intended to include all combinations and subcombinations of the ranges and specific embodiments therein. When referring to a number or numerical range, the term "about" means that the number or numerical range referred to is an approximation within experimental variability (or within statistical experimental error), and thus the number or numerical range will vary between 1% and 15% of the stated number or numerical range, as the case may be. The term "comprising" (and related terms such as "comprise" or "comprises" or "having" or "including") is not intended to exclude other specific embodiments, such as, for example, any composition, composition, method, or process described herein, from "consisting of" or "consisting essentially of" the recited features.

[0009] As used in this specification and the appended claims, unless stated otherwise, the following terms have the meanings indicated below.

[0010] "Pharmaceutically acceptable salt" includes both acid addition salt and base addition salt.The pharmaceutically acceptable salt of the heterocyclic RAF inhibitor described herein is intended to include any and all pharmaceutically suitable salt forms.Preferred pharmaceutically acceptable salt of the compound described herein is pharmaceutically acceptable acid addition salt and pharmaceutically acceptable base addition salt.

[0011] "Pharmaceutically acceptable acid addition salts" refers to salts which retain the biological effectiveness and properties of the free base and which are biologically or otherwise undesirable and are formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, hydroiodic acid, hydrofluoric acid, phosphorous acid, etc. Also included are salts formed with organic acids such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxyalkanoic acids, alkanedioic acids, aromatic acids, aliphatic and aromatic sulfonic acids, etc., such as acetic acid, trifluoroacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc. Thus, exemplary salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, trifluoroacetate, propionate, caprylate, isobutyrate, oxalate, malonate, succinate suberate, sebacate, fumarate, maleate, mandelate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, phthalate, benzenesulfonate, toluenesulfonate, phenylacetate, citrate, lactate, maleate, tartrate, methanesulfonate, and the like. Salts of amino acids such as arginates, gluconates, and galacturonates (see, e.g., Berge SM et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science, 66:1-19 (1997)) are also contemplated. Acid addition salts of basic compounds are prepared in some embodiments by contacting the free base form with a sufficient amount of the desired acid to produce the salt according to methods and techniques familiar to those of ordinary skill in the art.

[0012] "Pharmaceutically acceptable base addition salt" refers to a salt that retains the biological effectiveness and properties of the free acid and is not biologically or otherwise undesirable. These salts are prepared from the addition of an inorganic or organic base to the free acid. Pharmaceutically acceptable base addition salts are formed in some embodiments with metals or amines, such as alkali and alkaline earth metals or organic amines. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum salts, and the like. Salts derived from organic bases include, but are not limited to, salts of primary, secondary and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, N,N-dibenzylethylenediamine, chloroprocaine, hydrabamine, choline, betaine, ethylenediamine, ethylenedianiline, N-methylglucamine, glucosamine, methylglucamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine, polyamine resins, etc. See Berge et al., supra.

[0013] "Pharmaceutically acceptable solvate" refers to a composition of matter that is a solvent addition form. In some embodiments, the solvate contains either stoichiometric or non-stoichiometric amount of solvent and is formed during the process of making with pharma-ceutically acceptable solvents such as water, ethanol, etc. Hydrates are formed when the solvent is water, or alcoholates are formed when the solvent is alcohol. Solvates of the compounds described herein are conveniently prepared or formed during the process described herein. The compounds provided herein optionally exist in either unsolvated or solvated form.

[0014] The term "subject" or "patient" includes mammals. Examples of mammals include, but are not limited to, any member of the class Mammalia, including humans, non-human primates such as chimpanzees, and other ape and monkey species, domestic animals such as cows, horses, sheep, goats, pigs, domestic animals such as rabbits, dogs and cats, and laboratory animals including rodents such as rats, mice and guinea pigs. In one aspect, the mammal is a human.

[0015] As used herein, "treatment" or "treating" or "alleviating" or "ameliorating" are used interchangeably. These terms refer to an approach to obtain a beneficial or desired result, including, but not limited to, therapeutic benefit and / or prophylactic benefit. "Therapeutic benefit" refers to the eradication or amelioration of the underlying disease being treated. Also, therapeutic benefit is achieved by eradication or amelioration of one or more of the physiological symptoms associated with the underlying disease, such that an improvement is observed in the patient, even though the patient is still suffering from the underlying disease. For prophylactic benefit, the composition is administered in some embodiments to a patient at risk of developing a particular disease, or to a patient who reports one or more of the physiological symptoms of the disease, even if a diagnosis of the disease has not been made. As used herein, the term "treating" means, unless otherwise indicated, to reverse, alleviate, inhibit progression, or prevent the disorder or disease to which such term applies, or one or more symptoms of such disorder or disease. In some embodiments, the term "treating" includes slowing or slowing the progression of the disease or disorder to which the term applies. Furthermore, in some embodiments, the term "treat" applies to one or more of the complications resulting from the disease or disorder to which the term applies. The term "treatment", as used herein, unless otherwise indicated, refers to the act of treating as "treatment" defined above.

[0016] The term "tumor" or "cancer" as used herein refers to neoplastic cell growth, unless otherwise specified, and includes precancerous and cancerous cells and tissues. Tumors usually exist as a lesion or mass. As used herein, "treating" a tumor means that one or more symptoms of the disease, such as the tumor itself, the tumor's vascularization, or other parameters characteristic of the disease, are reduced, ameliorated, inhibited, placed in remission, or maintained in remission. "Treatment" of a tumor also means that one or more characteristics of the tumor may be eliminated, reduced, or prevented by treatment. Non-limiting examples of such characteristics include uncontrolled degradation of the basement membrane and proximal extracellular matrix, migration, division, and organization of endothelial cells into new functional capillaries, and persistence of such functional capillaries.

[0017] The terms "resistance" or "refractory to treatment" indicate that a patient has failed to respond to treatment altogether.

[0018] The term "relapsed" or "relapsed after treatment" indicates that a patient, after an initial response to a previous treatment, has progressive disease due to acquired resistance and / or intolerance.

[0019] The term "resistance to therapy" or "acquired resistance to therapy" refers to a patient who, after an initial response to a previous therapy, has progressive disease due to clinical or molecular resistance to the therapy. Acquired resistance can result from the emergence of resistance mutations in the molecular target of the therapy or in the development of physiological functions such as efflux pumps.

[0020] As used herein, the phrase "therapeutically effective amount" refers to that amount of a drug or pharmaceutical agent that elicits the biological or medical response in a tissue, system, animal or human that is desired by a researcher, veterinarian, physician or other.

[0021] Other aspects, advantages and features of the present invention will become apparent from the following detailed description.

[0022] RAF family kinases RAF family kinases (ARAF, BRAF, and RAF1) act downstream of receptor tyrosine kinases (RTKs) to regulate important cellular functions through regulation of the MAPK signaling pathway. In normal cells, external ligand stimulation of RTKs activates RAS GTPases that promote RAF dimer formation, protomer transactivation, and ultimately MAPK-dependent activation of the transcriptional program that drives cell proliferation and survival (Zaman et al., 2019). Mutational activation of the RTK-RAS-RAF-MAPK pathway is frequently observed in human cancers and represents an established target for therapeutic intervention (Roberts & Der, 2007, Imperial et al., 2017).

[0023] Oncogenic BRAF alterations occur in approximately 4–8% of all human cancers and can be functionally classified into three classes based on their unique structural and signaling properties (Davies, 2002; Owsley, 2020). Class I BRAF V600 mutations result in active BRAF monomers that signal independent of RAS recruitment and RAF dimerization / protomer transactivation (Yaeger & Corcoran, 2019). Class II BRAF mutations result in RAS-independent BRAF dimerization and protomer transactivation. Class III BRAF mutations impair BRAF kinase activity but promote its association with RAS. In this case, amplified MAPK signaling remains dependent on upstream RAS activation and heterodimerization with wild-type RAF (ARAF or RAF1). In contrast to constitutively active class I and II BRAF mutations, which rarely co-occur with other MAPK pathway alterations, class III BRAF alterations are frequently observed in association with aberrant RAS activation via RTK dysregulation, RAS mutations, or NF1 loss of function ( Yeager & Corcoran, 2019 ).

[0024] First-generation BRAF inhibitors (e.g., vemurafenib, dabrafenib) target class I mutations and have conferred significant clinical benefit to patients with BRAF V600 mutation-driven metastatic melanoma (~50% response rate, median progression-free survival of 5–6 months) as monotherapy, as well as other cancer types (Chapman, 2011; Hauschild, 2012; Subbiah, 2020) as part of combination therapy regimens. However, currently approved BRAF inhibitors have not proven effective in patients with class II and III BRAF mutations, which account for a significant proportion of BRAF mutations (e.g., 65% in NSCLC and 21% in melanoma patients; Owsley, 2021). Although RAF inhibitors designed to be effective in all classes of BRAF mutations would be desirable, class II and III BRAF-altered tumors exhibit resistance to current FDA-approved therapies and represent a patient population with unmet clinical need.

[0025] Heterocyclic RAF inhibitors The heterocyclic RAF inhibitors described herein refer to compound 1 having the following structure and chemical name: (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide.

[0026] [ka]

[0027] Compound 1 is a reversible small molecule RAF inhibitor. Throughout this disclosure, when referring to a heterocyclic RAF inhibitor, or a pharma- ceutically acceptable salt or solvate thereof, reference is made to Compound 1, or a pharma-ceutically acceptable salt or solvate thereof.

[0028] Cancer and methods of treatment In one embodiment, a method for inhibiting RAF enzyme is provided, comprising contacting the enzyme with compound 1 disclosed herein or a pharma- ceutically acceptable salt or solvate thereof.In a particular aspect, a method for treating cancer in an individual in need of cancer treatment is disclosed herein, comprising administering to the individual an effective amount of a heterocyclic RAF inhibitor as described herein.In a particular aspect, a heterocyclic RAF inhibitor as described herein is disclosed herein for use in treating cancer.In a particular aspect, a heterocyclic RAF inhibitor as described herein is disclosed herein for use in preparing a medicament for treating cancer.

[0029] One embodiment provides a method of treating cancer in a patient in need thereof, comprising administering to the patient (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof. Another embodiment provides a method, wherein the cancer is characterized as having an oncogenic BRAF alteration. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class I BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class II BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class III BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a BRAF V600 mutation. Another embodiment provides a method, wherein the BRAF V600 mutation is a V600E or V600K mutation. Another embodiment provides a method in which the cancer is characterized as having wild-type BRAF. Another embodiment provides a method in which the cancer is a solid tumor. Another embodiment provides a method in which the cancer is an NRAS mutant tumor. Another embodiment provides a method in which the cancer is selected from melanoma, pancreatic cancer, pancreatic adenocarcinoma, ovarian cancer, colon cancer, glioma, Langerhans cell histiocytosis, leukemia, hairy cell leukemia, thyroid cancer, anaplastic thyroid cancer, papillary thyroid cancer, follicular thyroid cancer, or medullary thyroid cancer. Another embodiment provides a method in which the cancer is selected from small cell lung cancer, non-small cell lung cancer, prostate cancer, gastric cancer, and esophageal cancer. Another embodiment provides a method in which the cancer is melanoma. Another embodiment provides a method in which the cancer is an NRAS mutant melanoma. Another embodiment provides a method in which the cancer is pancreatic cancer. Another embodiment provides a method in which the cancer is pancreatic adenocarcinoma. Another embodiment provides a method in which the cancer is ovarian cancer. Another embodiment provides a method in which the cancer is colon cancer. Another embodiment provides a method, wherein the cancer is glioma. Another embodiment provides a method, wherein the cancer is Langerhans cell histiocytosis. Another embodiment provides a method, wherein the cancer is leukemia. Another embodiment provides a method, wherein the cancer is hairy cell leukemia. Another embodiment provides a method, wherein the cancer is thyroid cancer.Another embodiment provides a method wherein the cancer is anaplastic thyroid cancer. Another embodiment provides a method wherein the cancer is papillary thyroid cancer. Another embodiment provides a method wherein the cancer is follicular thyroid cancer. Another embodiment provides a method wherein the cancer is medullary thyroid cancer. Another embodiment provides a method wherein the cancer is small cell lung cancer. Another embodiment provides a method wherein the cancer is non-small cell lung cancer. Another embodiment provides a method wherein the cancer is NRAS mutant non-small cell lung cancer. Another embodiment provides a method wherein the cancer is NRAS mutant lung cancer. Another embodiment provides a method wherein the cancer is prostate cancer. Another embodiment provides a method wherein the cancer is gastric cancer. Another embodiment provides a method wherein the cancer is cholangiocarcinoma. Another embodiment provides a method wherein the cancer is esophageal cancer. Another embodiment provides a method wherein the cancer is metastatic. Another embodiment provides a method wherein the cancer is adjuvant chemotherapy following surgical resection. Another embodiment provides a method wherein the patient has relapsed following previous treatment. Another embodiment provides a method wherein the patient has acquired resistance to previous treatment. Another embodiment provides a method wherein the patient exhibits resistance to treatment. Another embodiment provides a method in which (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, is orally administered. Another embodiment provides a method in which the oral administration is every other day, once a day, twice a day, or three times a day.

[0030] One embodiment provides a method of treating cancer in a patient in need thereof, comprising administering to the patient a pharmaceutical composition comprising (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, and at least one pharma- ceutically acceptable excipient. Another embodiment provides a method, wherein the cancer is characterized as having an oncogenic BRAF alteration. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class I BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class II BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class III BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a BRAF V600 mutation. Another embodiment provides a method in which the BRAF V600 mutation is a V600E or V600K mutation. Another embodiment provides a method in which the cancer is characterized as having wild-type BRAF. Another embodiment provides a method in which the cancer is a solid tumor. Another embodiment provides a method in which the cancer is an NRAS mutant tumor. Another embodiment provides a method in which the cancer is selected from melanoma, pancreatic cancer, pancreatic adenocarcinoma, ovarian cancer, colorectal cancer, glioma, Langerhans cell histiocytosis, leukemia, hairy cell leukemia, thyroid cancer, anaplastic thyroid cancer, papillary thyroid cancer, follicular thyroid cancer, or medullary thyroid cancer. Another embodiment provides a method in which the cancer is selected from small cell lung cancer, non-small cell lung cancer, prostate cancer, gastric cancer, and esophageal cancer. Another embodiment provides a method in which the cancer is selected from melanoma. Another embodiment provides a method in which the cancer is an NRAS mutant melanoma. Another embodiment provides a method in which the cancer is pancreatic cancer. Another embodiment provides a method in which the cancer is pancreatic adenocarcinoma. Another embodiment provides a method, wherein the cancer is ovarian cancer. Another embodiment provides a method, wherein the cancer is colon cancer. Another embodiment provides a method, wherein the cancer is glioma. Another embodiment provides a method, wherein the cancer is Langerhans cell histiocytosis. Another embodiment provides a method, wherein the cancer is leukemia. Another embodiment provides a method, wherein the cancer is hairy cell leukemia.Another embodiment provides a method in which the cancer is thyroid cancer. Another embodiment provides a method in which the cancer is anaplastic thyroid cancer. Another embodiment provides a method in which the cancer is papillary thyroid cancer. Another embodiment provides a method in which the cancer is follicular thyroid cancer. Another embodiment provides a method in which the cancer is medullary thyroid cancer. Another embodiment provides a method in which the cancer is small cell lung cancer. Another embodiment provides a method in which the cancer is non-small cell lung cancer. Another embodiment provides a method in which the cancer is NRAS mutant non-small cell lung cancer. Another embodiment provides a method in which the cancer is NRAS mutant lung cancer. Another embodiment provides a method in which the cancer is prostate cancer. Another embodiment provides a method in which the cancer is gastric cancer. Another embodiment provides a method in which the cancer is cholangiocarcinoma. Another embodiment provides a method in which the cancer is esophageal cancer. Another embodiment provides a method in which the cancer is metastatic. Another embodiment provides a method in which the cancer is adjuvant chemotherapy following surgical resection. Another embodiment provides a method in which the patient has relapsed after previous treatment. Another embodiment provides a method in which the patient has acquired resistance to previous treatment. Another embodiment provides a method in which a patient exhibits resistance to treatment. Another embodiment provides a method in which (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, is orally administered. Another embodiment provides a method in which the oral administration is every other day, once a day, twice a day, or three times a day.

[0031] One embodiment is a method of treating cancer in a patient in need of such treatment, comprising: (a) a pharmaceutical composition comprising (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof; (b) at least one oncology therapeutic agent selected from a MEK inhibitor, an immune checkpoint inhibitor, a CDK inhibitor, an EGFR kinase inhibitor, an EGFR antibody, an EGFR PROTAC therapeutic agent, an FGFR inhibitor, an SOS1 inhibitor, an SHP2 inhibitor, a KRAS inhibitor, a taxane, a topoisomerase inhibitor, an ERK inhibitor, a platinum-based chemotherapy agent, folinic acid, 5-fluorouracil, or an anti-VEGF therapeutic agent; to a patient. Another embodiment provides a method in which the at least one tumor therapeutic is a MEK inhibitor. Another embodiment provides a method in which the MEK inhibitor is selected from binimetinib, trametinib, cobimetinib, selumetinib, pimasertib, or mirdametinib. Another embodiment provides a method in which the at least one tumor therapeutic is an immune checkpoint inhibitor. Another embodiment provides a method in which the immune checkpoint inhibitor is a CTLA-4 inhibitor, a PD-1 inhibitor, or a PD-L1 inhibitor. Another embodiment provides a method in which the CTLA-4 inhibitor is ipilimumab. Another embodiment provides a method in which the PD-1 inhibitor is spartalizumab, nivolumab, pembrolizumab, or cemiplimab. Another embodiment provides a method in which the PD-L1 inhibitor is atezolizumab, avelumab, or durvalumab. Another embodiment provides a method in which the at least one tumor therapeutic is a CDK inhibitor. Another embodiment provides a method in which the CDK inhibitor is a CDK4 / 6 inhibitor. Another embodiment provides a method in which the CDK4 / 6 inhibitor is palbociclib, abemaciclib, or ribociclib. Another embodiment provides a method in which the at least one tumor therapeutic is an EGFR kinase inhibitor or antibody. Another embodiment provides a method in which the EGFR kinase inhibitor is nazartinib, gefitinib, erlotinib, afatinib, brigutinib, icotinib, neratinib, osimertinib, dacomitinib, or lapatinib. Another embodiment provides a method in which the EGFR antibody is cetuximab, panitumumab, zalutumumab, nimotuzumab, or matuzumab. Another embodiment provides a method in which the at least one tumor therapeutic is a SHP2 inhibitor. Another embodiment provides a method in which at least one tumor therapeutic is an SOS1 inhibitor. Another embodiment provides a method in which at least one tumor therapeutic is an FGFR inhibitor. Another embodiment provides a method in which at least one tumor therapeutic is a KRAS inhibitor. Another embodiment provides a method in which the KRAS inhibitor is sotorasib, adagrasib, or BI-1701963. Another embodiment provides a method in which at least one tumor therapeutic is a taxane.Another embodiment provides a method in which at least one tumor therapeutic is a topoisomerase inhibitor. Another embodiment provides a method in which the topoisomerase inhibitor is irinotecan, topotecan, or belotecan. Another embodiment provides a method in which at least one tumor therapeutic is an ERK inhibitor. Another embodiment provides a method in which at least one tumor therapeutic is a platinum-based chemotherapeutic agent. Another embodiment provides a method in which the platinum-based chemotherapeutic agent is oxaliplatin, cisplatin, or carboplatin. Another embodiment provides a method in which at least one tumor therapeutic is folinic acid. Another embodiment provides a method in which at least one tumor therapeutic is 5-fluorouracil. Another embodiment provides a method in which at least one tumor therapeutic comprises folinic acid and folinic acid. Another embodiment provides a method further comprising administration of a topoisomerase inhibitor or a platinum-based chemotherapeutic agent, or a combination thereof. Another embodiment provides a method in which at least one tumor therapeutic is an anti-VEGF therapeutic agent. Another embodiment provides a method, wherein the anti-VEGF therapeutic is bevacizumab or aflibercept. Another embodiment provides a method, wherein the cancer is characterized as having an oncogenic BRAF alteration. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class I BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class II BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a class III BRAF mutation. Another embodiment provides a method, wherein the oncogenic BRAF alteration is a BRAF V600 mutation. Another embodiment provides a method, wherein the BRAF V600 mutation is a V600E or V600K mutation. Another embodiment provides a method, wherein the cancer is characterized as having wild-type BRAF. Another embodiment provides a method, wherein the cancer is a solid tumor. Another embodiment provides a method, wherein the cancer is an NRAS mutant tumor. Another embodiment provides a method, wherein the cancer is selected from melanoma, pancreatic cancer, pancreatic adenocarcinoma, ovarian cancer, colorectal cancer, glioma, Langerhans cell histiocytosis, leukemia, hairy cell leukemia, thyroid cancer, anaplastic thyroid cancer, papillary thyroid cancer, follicular thyroid cancer, or medullary thyroid cancer.Another embodiment provides a method wherein the cancer is selected from small cell lung cancer, non-small cell lung cancer, prostate cancer, gastric cancer, and esophageal cancer. Another embodiment provides a method wherein the cancer is selected from melanoma. Another embodiment provides a method wherein the cancer is an NRAS mutant tumor. Another embodiment provides a method wherein the cancer is pancreatic cancer. Another embodiment provides a method wherein the cancer is pancreatic adenocarcinoma. Another embodiment provides a method wherein the cancer is ovarian cancer. Another embodiment provides a method wherein the cancer is colorectal cancer. Another embodiment provides a method wherein the cancer is glioma. Another embodiment provides a method wherein the cancer is Langerhans cell histiocytosis. Another embodiment provides a method wherein the cancer is leukemia. Another embodiment provides a method wherein the cancer is hairy cell leukemia. Another embodiment provides a method wherein the cancer is thyroid cancer. Another embodiment provides a method wherein the cancer is anaplastic thyroid cancer. Another embodiment provides a method wherein the cancer is papillary thyroid cancer. Another embodiment provides a method wherein the cancer is follicular thyroid cancer. Another embodiment provides a method wherein the cancer is medullary thyroid cancer. Another embodiment provides a method, wherein the cancer is small cell lung cancer. Another embodiment provides a method, wherein the cancer is non-small cell lung cancer. Another embodiment provides a method, wherein the cancer is NRAS mutant non-small cell lung cancer. Another embodiment provides a method, wherein the cancer is NRAS mutant lung cancer. Another embodiment provides a method, wherein the cancer is prostate cancer. Another embodiment provides a method, wherein the cancer is gastric cancer. Another embodiment provides a method, wherein the cancer is cholangiocarcinoma. Another embodiment provides a method, wherein the cancer is esophageal cancer. Another embodiment provides a method, wherein the cancer is metastatic. Another embodiment provides a method, wherein the cancer is adjuvant chemotherapy following surgical resection. Another embodiment provides a method, wherein the patient has relapsed after previous treatment. Another embodiment provides a method, wherein the patient has acquired resistance to previous treatment. Another embodiment provides a method, wherein the patient exhibits resistance to treatment. Another embodiment provides a method wherein (S)—N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, is administered orally.Another embodiment provides a method wherein oral administration occurs every other day, once a day, twice a day, or three times a day.

[0032] In certain embodiments, the heterocyclic RAF inhibitors described herein are administered as pure chemicals. In other embodiments, the heterocyclic RAF inhibitors described herein are combined with a pharma- ceutically suitable or acceptable carrier (also referred to herein as a pharma- ceutically suitable or acceptable excipient, a physiologically suitable or acceptable excipient, or a physiologically suitable or acceptable carrier) selected based on the selected route of administration and standard pharmaceutical practice.

[0033] Provided herein are pharmaceutical compositions comprising a heterocyclic RAF inhibitor as described herein, or a stereoisomer, pharma- ceutically acceptable salt, hydrate, or solvate thereof, together with one or more pharma- ceutically acceptable carriers. A carrier (or excipient) is acceptable or suitable if it is compatible with the other components of the composition and not deleterious to the recipient of the composition (i.e., the subject or patient).

[0034] One embodiment provides a method for preparing a pharmaceutical composition comprising mixing a heterocyclic RAF inhibitor described herein, or a stereoisomer, pharma- ceutically acceptable salt, hydrate, or solvate thereof, with a pharma- ceutically acceptable carrier.

[0035] Provided herein is a method in which pharmaceutical compositions are orally administered. Suitable oral dosage forms include, for example, tablets, pills, sachets, or capsules of hard or soft gelatin, methylcellulose, or other suitable material that dissolves easily in the digestive tract. In some embodiments, suitable non-toxic solid carriers are used, including, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, talc, cellulose, glucose, sucrose, magnesium carbonate, etc. (see, for example, Remington: The Science and Practice of Pharmacy (Gennaro, 21st Ed. Mack Pub. Co., Easton, PA (2005)).

[0036] Provided herein is a method in which pharmaceutical compositions are administered by injection.In some embodiments, the heterocyclic RAF inhibitor described herein or its pharma-ceutically acceptable salt or solvate is formulated for administration by injection.In some examples, the injection formulation is an aqueous formulation.In some examples, the injection formulation is a non-aqueous formulation.In some examples, the injection formulation is an oil-based formulation, such as sesame oil.

[0037] The dosage of the composition comprising the heterocyclic RAF inhibitor described herein, or its stereoisomer, pharma- ceutically acceptable salt, hydrate or solvate, varies depending on the condition of the subject or patient (e.g., human). In some embodiments, such factors include general health, age, and other factors. The pharmaceutical composition is administered in a manner appropriate to the disease to be treated (or prevented). The appropriate dosage and the appropriate duration and frequency of administration are determined by factors such as the patient's condition, the type and severity of the patient's disease, the specific form of the active ingredient, and the method of administration. In general, the appropriate dosage and treatment regimen provides the composition in an amount sufficient to provide therapeutic and / or prophylactic benefits (e.g., improved clinical outcomes, e.g., more frequent complete or partial remissions, or longer disease-free and / or overall survival, or reduced severity of symptoms). The optimal dosage depends on the patient's body weight, body weight, or blood volume. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0038] These examples are provided for illustrative purposes only and are not intended to limit the scope of the claims provided herein.

[0039] Example 1. RAF Inhibitory Activity of Compound 1 method Inhibition of BRAF and RAPT (CRAF) kinases by compound 1 was measured by ADP-Glo ​​assay in 17 and 18 independent experiments for BRAF and CRAF, respectively, as shown in Table 1. In the assay, ATP is converted to ADP by the test kinase in the presence of substrate. The assay reagent then converts the remaining ADP to ATP, resulting in a luciferase reaction and luminescent readout that is proportional to the relative kinase activity. Compound 1 was diluted in DMSO and used in a 10-point, 3-fold dose curve for both assays. A high dose of 1 μM and a low dose of 0.05 nM were tested. Final concentrations of 6 nM BRAF (CarnaBio, Cat. No. 09-122) or 3 nM RAF1 (CarnaBio, Cat. No. 09-125) and 30 nM MEK1 substrate (Millipore, Cat. No. 14-420) were incubated for 90 min at room temperature with 3 μM ATP, 10 mM MgCl2, 0.003% Brij-35, 2 mM DTT, 0.05% BSA, 1 mM EGTA, and 50 mM HEPES before the addition of ADP-Glo ​​reagent (Promega, Cat. No. V9102) for 40 min. Luminescence was read on an Envision plate reader (PerkinElmer) and percent remaining activity was used to calculate IC with a four-parameter fitting model using Dotmatics Knowledge Solutions Studies curve fitting software (Dotmatics, Bishops Stortford, UK, CM23). 50 was calculated.

[0040] [Table 1]

[0041] Reaction Biology's (Malvern, PA) Kinase Screening Service measures the effect of test compounds on kinase activity via its Kinase Hot Spot Assay, using the transfer of radioactive 33P-labeled phosphate from ATP to the kinase substrate (Anastas Siadis T et al 2011). Reactions were performed with Km ATP of 15 μM, 20 μM, 20 μM, and 10 μM for ARAF, BRAF, BRAF-V600E, and RAF1 (CRAF), respectively. Appropriate substrates were diluted in reaction buffer containing 20 mM HEPES (pH 7.5), 10 mM MgCl2, 1 mM EGTA, 0.01% Brij35, 0.02 mg / ml BSA, 0.1 mM Na3V04, 2 mM DTT, 1% DMSO. After incorporating the kinase into the solution, compound 1 was added in a 10-point, 3-fold dose curve starting at a top dose of 3 μM and a low dose of 0.152 μM for a 20 minute pre-incubation at room temperature. 33 The reaction was initiated by adding P-ATP to the reaction mixture and incubated at room temperature for 2 h. Kinase activity was detected by P81 filter binding method. IC 50 Values ​​were calculated using a four-parameter fitting model using Dotmatics Knowledge Solutions Studies curve fitting software (Dotmatics, Bishops Stortford, UK, CM23) and are provided in Table 2.

[0042] [Table 2]

[0043] Compound 1 was tested with 1 μM ATP (when available) against the human kinome in the Cama Biosciences Kinase Panel of 299 kinases by Cama Biosciences (Kobe, Japan) using the Caliper LabChip3000 assay (Caliper Life Sciences, Hopkinton, MA), a mobility shift assay that combines the basic principles of capillary electrophoresis in a microfluidic environment (Perrin et al., 2010). 4x substrate / ATP / metal solutions were prepared in kit buffer (20 mM HEPES, 0.01% Triton X-100, 5 mM DTT, pH 7.5) and 2x kinase solutions were prepared in assay buffer (20 mM HEPES, 0.01% Triton X-100, 1 mM DTT, pH 7.5). 5 μL of 4x compound solution, 5 mL of 4x substrate / ATP / metal solution, and 10 mL of 2x kinase solution were mixed and incubated at room temperature for 1 or 5 hours in wells of a polypropylene 384-well microplate. (*; kinase dependent). 70 mL of termination buffer (QuickScout Screening Assist MSA, Carna Biosciences) was added to the wells. The reaction mixture was applied to a LabChip™ system (PerkinElmer) and product and substrate peptide peaks were separated and quantified. Kinase reactions were assessed by product ratios (P / (P+S)) calculated from the peak heights of product (P) and substrate (S) peptides. Kinases outside the RAF family that were inhibited by >10% by 1 μM compound 1 are listed in Table 3.

[0044] [Table 3]

[0045] conclusion The biochemical potency and kinase selectivity of Compound 1 were determined using a suite of biochemical assays. The inhibitory potency of Compound 1 against rapidly accelerated fibrosarcoma (RAF) kinase was profiled in both an adenosine diphosphate (ADP)-Glo screening assay as well as in a radiometric kinase assay. Compound 1 showed an IC 50 Compound 1 was demonstrated to be a potent inhibitor of RAF1, BRAF, and ARAF kinases with activity <5 nM. Non-RAF kinome selectivity was assessed in a panel of kinases as measured by mobility shift assays, revealing that only DDR1 and DDR2 were inhibited by ≥40% at 1 μM. In summary, compound 1 is a potent, selective RAF inhibitor with minimal activity against the remainder of the kinome.

[0046] Example 2. Compound 1 inhibits cell proliferation in cell culture Compound 1 is a selective and potent inhibitor of RAF kinase designed to target class II and III BRAF mutations in addition to class I BRAF mutations. Here, we evaluated the in vitro pharmacodynamic biomarker activity of compound 1 across multiple cell models representing four categories of BRAF mutation status: class I (A-375), class II (BxPC-3, OV-90, NCI-H2405), class III (WM3629 and CAL12-T), and wild-type (MIA PaCa-2, CHL, NCI-H358).

[0047] method A-375, BxPC-3, OV-90, NCI-H2405, CAL-12T, MIA PaCa-2, NCI-H358, and CHL-1 cells were seeded at 8000 cells / well in 24 μL growth medium in 384-well plates and allowed to adhere overnight at 37 °C with 5% CO2. WM3629 cells were seeded at 1500 cells / well in 24 μL growth medium in 384-well plates and allowed to adhere overnight at 37 °C with 5% CO2. The next day, compounds were serially diluted in a 10-point, 3-fold dilution curve in 384-well plates. Compound 1 was transferred to the cell plates to a final concentration range of 0.508 nM to 10 μM in 0.1% DMSO, and 0.1% DMSO was used as a negative control. Cells were incubated with compound 1 for 1 h at 37 °C with 5% CO2. Cells were lysed by the addition of 8 μL of 4X lysis buffer provided with the HTRF kit plus 1X protease / phosphatase inhibitor cocktail. 20 μL of lysate was transferred to the HTRF plate and 2.5 μL each of anti-ERK1 / 2-europium / terbium cryptate and anti-phospho-ERK1 / 2 antibody solutions were transferred according to the manufacturer's instructions. The HTRF plate was incubated overnight at room temperature and then read in HTRF mode at 665 nm / 620 nm (donor / acceptor). EC was calculated using dose-response regression curve fitting utilizing a four-parameter analysis method. 50 values ​​were calculated.

[0048] Compound 1 demonstrated variable cellular activity across BRAF mutant and wild-type models as determined by pharmacodynamic modulation of the pERK biomarker 1 hour after treatment, as shown in Table 4A. In tumor cells harboring class III BRAF alterations, models WM3629 and CAL-12T, compound 1 demonstrated mean EC 50 In the class II BRAF mutant models, BxPC-3, OV-90, and NCI-H2405, compound 1 had mean EC values ​​of 50.7, 26.0, and 10.1 nM, respectively. 50 Cells harboring class I BRAF mutations, A-375, were the least responsive mutant class, with an average EC 50 In three wild-type models, MIA PaCa-2, CHL-1, and NCI-H358, Compound 1 had mean EC 50 Further cellular activities are provided in Table 4B.

[0049] [Table 4A]

[0050] [Table 4B]

[0051] conclusion In summary, compound 1 demonstrated variable cellular activity across BRAF mutant and wild-type models as determined by modulation of the pERK biomarker. Class II and class III mutant models demonstrated the strongest cellular responses when treated with compound 1, whereas wild-type BRAF models were the least responsive.

[0052] Example 3. Determination of antiproliferative activity in a xenograft model method

[0053] Groups and treatments were assigned to patients with mean tumor volumes between 200 and 335 mm as shown in the results section. 3 Treatment was started when tumor volume reached 100 mg / kg / day. Mice were assigned to respective groups based on their starting tumor volume and body weight, such that the mean values ​​were the same for each treatment group. The test groups and number of animals per group are shown in Tables 5-7.

[0054] A. Evaluation of the antitumor activity of compound 1 in class I BRAF mutant human cancers The antitumor activity of Compound 1 was initially evaluated in a human A-375 melanoma xenograft model harboring a class I BRAF V600E mutation. Treatment with Compound 1 (15, 30, or 60 mg / kg, free base form) was administered to tumors with tumor volumes of approximately 200 mm 3 Treatment with Compound 1 was initiated when tumor volumes reached an average of 205 mm and continued once daily (QD) for 3 weeks. Mean A-375 xenograft tumor volume (FIG. 1A), mouse body weight (FIG. 1B), and percent change in xenograft tumor volume at the end of the study relative to baseline (FIG. 1C) following treatment with Compound 1 are shown by dose cohort. Compound 1 treatment was initiated when tumor volumes reached an average of 205 mm 3 Treatment was started when tumor volume reached 0.01 mg / kg / day and continued for 21 days at the indicated doses (n=9 mice / group). Mean tumor volume and body weight are plotted, respectively, with error bars indicating standard error of the mean. Percentage change in tumor volume was calculated using the formula (TVf-TV0) / TV0x100%, where TVf=final tumor volume (at the end of treatment) and TV0=initial tumor volume (at the start of treatment).

[0055] [Table 5]

[0056] Compound 1 treatment resulted in a systemic plasma exposure (AUClast) of 11,900-156,000 h*ng / mL when administered orally for 24 hours after the last dose in mice bearing mutant A-375 BRAF class I melanoma tumors. Dose-dependent inhibition of xenograft tumor growth relative to control (vehicle-treated) tumors was observed with compound 1 treatment (Figure 1A). All of the doses tested were well tolerated, with some weight loss observed in animals treated at 60 mg / kg (5.6% mean weight loss, Figure 1B). One animal in the 60 mg / kg cohort had a >10% weight loss over the course of treatment but recovered by the end of the study.

[0057] Waterfall plots of individual tumor response, defined as change in tumor volume from baseline, are shown in Figure 1C. Mean TGI achieved at daily doses of 30 mg / kg and 60 mg / kg was 57% and 106%, respectively (p ≤ 0.0001, Figure 1C), with 8 of 9 mice (89%) in the latter group exhibiting tumor regression. The lowest tested dose, 15 mg / kg, did not result in a significant TGI.

[0058] B. Evaluation of the antitumor activity of compound 1 in class II / III BRAF mutant human cancers We next evaluated the antitumor activity of compound 1 in human xenograft models displaying class II / III BRAF mutations. Xenografts derived from BxPC-3 pancreatic cancer and WM3629 melanoma cell lines harboring class II BRAF insertion deletions (indels, V487_P492delinsA) and class III BRAF D594G mutations, respectively, were treated daily for 2 weeks with a total of 3–20 mg / kg of compound 1 (sulfate salt form). 3 Treatment began when tumor volume reached 100 mg / kg, and animals were administered one (QD, 3–20 mg / kg) or two (BID, 1.5–10 mg / kg) equivalent total doses daily. Mean BxPC-3 (class II BRAF mutant PD-AC) and WM3629 (class III BRAF mutant melanoma) xenograft tumor volumes (Figure 2A, 2C) and mouse weights (Figure 2B, 2D) during treatment with compound 1 are presented by dose cohort. Compound 1 treatment was administered once daily, with tumor volumes reaching an average of 240–283 mm. 3 Treatment was started when the tumor volume reached 100 mg / kg and continued for 14 days at the indicated dose and frequency (n=9 mice / group). Mean tumor volumes and body weights are plotted with error bars indicating standard error of the mean.

[0059] [Table 6]

[0060] Compound 1 treatment resulted in systemic plasma exposures (AUC ) of 4,030–42,100 h*ng / mL and 5,200–53,700 h*ng / mL when administered orally BID or QD, respectively, for 24 hours after the last dose in mice bearing mutant BRAF class II BxPC-3 pancreatic and class III WM3629 melanoma xenografts. last ) Dose-dependent inhibition of BxPC-3 and WM3629 xenograft tumor growth relative to control (vehicle-treated) tumors was observed with compound 1 treatment (Figure 2A, Figure 2C). A trend toward greater TGI was achieved with BID compared to QD administration of an equivalent total daily dose of compound 1 in both models. All tested doses and schedules were well tolerated as measured by the change in animal body weight during treatment (Figure 2B, Figure 2D).

[0061] Waterfall plots of individual BxPC-3 tumor and WM3629 tumor xenograft responses are shown in Figure 3A and Figure 3B, respectively. Statistically significant reductions in BxPC-3 tumor growth were achieved at all tested doses up to 10 mg / kg total compound 1 / day (51-88% inhibition, p ≤ 0.0001, Figure 3A). Additionally, tumor regressions were observed among animals in the 20 mg / kg QD and 3-10 mg / kg BID treatment groups (TGI of 109% and 118% at 20 mg / kg QD and 10 mg / kg BID, respectively, p ≤ 0.0001). Similarly, TGI was observed at all tested doses and schedules in the WM3629 model (47-101% inhibition, p ≤ 0.04-0.0001), with regression observed at the highest daily dose in some cases (Figure 3B). Consistent with the BxPC-3 model, regression was also achieved in one animal at a lower dose (5 mg / kg) when administered BID.

[0062] C. Evaluation of the antitumor activity of compound 1 in combination with MEK inhibition in class III BRAF mutant human cancers The antitumor activity of compound 1 in combination with binimetinib, a MEK inhibitor approved for use in combination with encorafenib for the treatment of advanced melanoma harboring the BRAF V600E / K mutation, was evaluated in a human WM3629 (class III BRAF mutant) melanoma xenograft model. Treatment with compound 1 (15 or 30 mg / kg QD, free base form), binimetinib (10 mg / kg QD), and / or the combination of the two therapies was administered to patients with tumor volumes of approximately 335 mm. 3 The treatment started when the patient reached 100 mg / kg / day and continued once daily for 2 weeks.

[0063] [Table 7]

[0064] Systemic plasma exposure (AUC last ) were achieved with compound 1 and / or binimetinib treatment, respectively, at the doses tested. A dose-dependent response to compound 1 monotherapy was observed (Figure 4A,C). The mean TGI was 86% and 99% at 15 mg / kg and 30 mg / kg, respectively. Treatment with binimetinib (10 mg / kg QD) as a single agent did not result in significant inhibition of tumor growth. However, the addition of compound 1 to binimetinib resulted in improved tumor xenograft growth inhibition with evidence of tumor regression (TGI of 101% and 108% with binimetinib / compound 1 at 10 / 15 mg / kg and 10 / 30 mg / kg, respectively, p ≤ 0.0001, Figure 4C). Importantly, treatment with compound 1 plus binimetinib was well tolerated, with only one animal at the highest combination therapy dose showing a 10% weight loss at the start of a single treatment (Figure 4B).

[0065] conclusion Compound 1 was well tolerated and effective at doses up to 60 mg / kg / day in athymic nude mouse xenograft models of BRAF mutant human cancer. Dose-dependent inhibition of A-375 (BRAF class I mutated), BxPC-3 (BRAF class II mutated), or WM3629 (BRAF class III mutated) xenograft tumor growth was observed with daily compound 1 treatment. Although a trend toward more robust tumor responses was observed with twice-daily compared with once-daily administration of compound 1, the two dosing regimens resulted in similar tumor growth inhibition (TGI) and regression (mean TGI of 101-118%, p ≤ 0.0001) at comparable total daily doses. Compound 1 was also well tolerated and effective when combined with a MEK inhibitor resulting in WM3629 xenograft tumor regression (mean TGI of 101-106%, p ≤ 0.0001). These data indicate that compound 1 has potent antitumor activity as a single agent and can also be safely combined with other targeted therapies for the treatment of BRAF-driven cancers.

[0066] Example 4. Evaluation of the cellular activity of Compound 1 in combination with binimetinib in NRAS-mutated melanoma Objective: To evaluate the cellular potency and activity of compound 1, a selective and potent inhibitor of rapidly accelerated fibrosarcoma (RAF) kinase, in combination with binimetinib, a mitogen-activated protein kinase kinase (MEK) inhibitor.

[0067] Methods. The combinatorial activity of the RAF inhibitor compound 1 plus binimetinib was evaluated in human neuroblastoma rat sarcoma viral oncogene homolog (NRAS) mutant melanoma cells in a 7-day proliferation assay. SK-MEL-2 human melanoma cells were obtained from the American Type Culture Collection and maintained in DMEM 10% FBS 1% penicillin / streptomycin. All cells were maintained in a humidified incubator at 37°C with 5% carbon dioxide (CO2). SK-MEL-2 cells were seeded at 1,000 cells / well in 96 μL growth medium in 96-well plates (Corning 3904) and allowed to adhere overnight at 37°C in 5% CO2. The next day, compound 1 was serially diluted in dimethyl sulfoxide (DMSO) into a 9-point, 3-fold dilution curve in 96-well plates starting at 1 mM. Binimetinib was serially diluted in DMSO into a 5-point, 4-fold dilution curve in a 96-well plate starting at 500 mM. Then, 2 μL of each compound was transferred into 196 μL of medium to make a 10X working dilution. 10 μL of the working dilution was dispensed into the corresponding wells of the 96-well plate to a total volume of 100 μL. A 0.2% solution of DMSO was used as a negative control. Cells were incubated with compounds at 37° C. with 5% CO2 for 7 days. The plate was removed from the incubator and equilibrated at room temperature for 15 minutes. Then, 100 μL of CellTiter-Glo (CTG) reagent was added to each well to be detected (1:1 to medium). The plate was kept at room temperature for 30 minutes and then read on a plate reader. EC 50Values ​​were calculated using dose-response regression curve fitting utilizing a four-parameter analysis method. Synergy analysis of the data was performed to evaluate the potential synergistic combination benefit of compound 1 and binimetinib (Meyer et al. 2019, Ianevski et al. 2020). Single agent compound 1 (including DMSO control) had a proliferation inhibitory EC of 164 nM. 50 When combined with increasing concentrations of binimetinib, the potency of compound 1 was enhanced, with EC values ​​of 2, 7.8, 31.3, 125, and 500 nM for binimetinib concentrations, respectively, as shown in Table 8 and Figure 5. 50 The values ​​shifted to 108 nM, 106 nM, 65 nM, 39 nM, and 12 nM. This cellular study demonstrates the combination benefit of adding the MEK inhibitor binimetinib to Compound 1 treatment in human NRAS mutant melanoma cell lines.

[0068] [Table 8]

[0069] Potential synergy between compound 1 and binimetinib in SK-MEL-2 cells was evaluated using multiple available synergy models. The multidimensional synergy of combination (MuSyC) model takes into account both potency (α) and efficacy (β) in the model. A score >0 indicates synergy between the two drugs at the indicated parameters. The highest single agent (HSA), Bliss, and Loewe synergy models were evaluated and utilized to find the most synergistic area score, with a score >10 indicating that the interaction between the two drugs is likely to be synergistic. Results from the 7-day proliferation assay were analyzed using multiple synergy models (see Table 9). The MuSyC model resulted in a beta score of 0.44 and an alpha score of 10.61, indicating both synergistic combination efficacy and synergistic potency in the SK-MEL-2 study. The HSA, Bliss, and Loewe models calculated the most synergistic area scores corresponding to 21.6, 16.2, and 11.6, respectively, indicating synergy within the ranges shown in Table 9. Taken together, this cellular study and subsequent synergy analysis demonstrates the benefit of a synergistic combination between compound 1 and binimetinib in the SK-MEL-2 human NRAS mutant melanoma cell line.

[0070] [Table 9]

[0071] Summary of Results: Compound 1 demonstrated potent inhibition of NRAS mutant human melanoma tumor cell models as evidenced by inhibition of cell proliferation in a 7-day proliferation assay. The growth inhibitory potency of Compound 1 was enhanced by combination with the MEK inhibitor binimetinib, as evidenced by a decrease in half-maximal effective concentration (EC50) values ​​with increasing doses of binimetinib.

[0072] Example 5. Evaluation of the antitumor activity of Compound 1 in combination with MEK inhibition in NRAS mutant human melanoma The antitumor activity of Compound 1 in combination with binimetinib was evaluated in a human SK-MEL-2 (NRAS Q61R mutant) melanoma xenograft model. Treatment with Compound 1 (10 or 30 mg / kg BID), binimetinib (3 mg / kg BID), or the combination of the two therapies was compared in patients with tumors of approximately 307 mm 3 Treatment was started when the patient reached 18.5 mg / kg / day and continued twice daily for 4 weeks.

[0073] [Table 10]

[0074] Systemic plasma exposure (AUC 0-24 ) was achieved with compound 1 and / or binimetinib treatment, respectively, at the doses tested. Tumor growth inhibition was observed in the monotherapy and combination therapy groups (Figure 6). The mean TGI was 13% and 135% at 10 mg / kg and 30 mg / kg, respectively. Treatment with binimetinib (3 mg / kg BID) as a single agent resulted in 23% tumor growth inhibition. The addition of 10 mg / kg compound 1 to binimetinib resulted in improved tumor xenograft growth inhibition with evidence of tumor regression (122%, 10 / 3 mg / kg binimetinib / compound 1, respectively, Figure 7).

[0075] Example 6. Evaluation of the antitumor activity of Compound 1 in combination with binimetinib in NRAS mutant BRAF mutant melanoma xenograft models Objective: To evaluate the tolerability and antitumor activity of compound 1, a selective and potent inhibitor of fibrosarcoma (RAF) kinase, and binimetinib, a mitogen-activated protein kinase (MEK) inhibitor, in neuroblastoma rat sarcoma viral oncogene homolog (NRAS) mutant and rapidly accelerated fibrosarcoma kinase homolog B (BRAF) mutant human cancer cell line-derived xenograft models.

[0076] Methods. Athymic BALB / c nude mice were inoculated subcutaneously with 1x107 human cancer cells in the right flank. Compound 1 and / or binimetinib were orally (PO) administered twice daily (BID) until tumor volume (TV) of approximately 300 mm 3 The treatment period began when the tumor volume reached 10 μg / kg and continued for 2 weeks. Animals were monitored for tumor growth, weight change, and general health / behavior.

[0077] The average TV is about 300mm 3 Groups and treatments were initiated when the mice reached a TV of 0.01 mg / kg / day. Mice were assigned to each group based on their starting TV and body weight so that the mean values ​​were the same for each treatment group. The test groups and number of animals per group are shown in Table 11.

[0078] [Table 11]

[0079] Mice were implanted with 1x10 cells in 0.1 mL of 1:1 medium / Matrigel on the right flank for tumor development. 7 WM3629 cells were inoculated subcutaneously. Tumor-bearing mice were treated according to the study design shown in Table 12. For routine monitoring, all study animals were monitored for tumor growth as well as any abnormalities in behavior and appearance, such as mobility, food and water consumption, body weight, eye appearance, and any other abnormal observations. Any mortality and / or abnormal clinical signs were recorded. All animals were weighed and recorded every other day throughout the study. Tumor size measurements were made with calipers and recorded three times in the first week, and twice weekly in the second and third weeks. TV (mm 3 ) was estimated using the formula: TV=axb 2 / 2, where "a" and "b" are the long and short diameters of the tumor, respectively.

[0080]

number

[0081] Patients were excluded if they showed overt signs of severe distress and / or pain, showed significant weight loss (weight loss >20%), were unable to reach adequate food or water, were otherwise observed to be in a state of continued deterioration, or had tumors measuring ≥2,500 mm 3 Animals were euthanized if the group mean TV was >2,500 mm 3 When the GI tract infection was reached, all animals in a given test group were euthanized.

[0082] Results. The antitumor activity of compound 1 in combination with binimetinib was evaluated in human WM3629 (NRAS mutant, class III BRAF mutant) melanoma xenograft models. Treatment with compound 1 (1.5 or 3 mg / kg twice daily [BID]), binimetinib (3 mg / kg BID), and / or the combination of the two therapies resulted in tumor progression with a TV of approximately 300 mm 3 (The actual mean TV for all groups was 283 mm. 3 ) and continued BID for 2 weeks. AUC ranges of 2,530-6,750hr*ng / mL and 5,400-5,730hr*ng / mL 0-last Values ​​were achieved with Compound 1 and / or binimetinib treatment at the doses tested, respectively. Neither Compound 1 nor binimetinib exposure was substantially different when administered as monotherapy or in combination.

[0083] A dose-dependent response to compound 1 monotherapy was observed (Figures 8A and 8C). The mean TGI was 66% and 79% with 1.5 mg / kg BID and 3 mg / kg BID compound 1, respectively (Table 13). Treatment with binimetinib (3 mg / kg BID) as a single agent resulted in a mean TGI of 37% (Figures 8A and 8C, Table 13). The addition of compound 1 to binimetinib resulted in improved xenograft growth inhibition, with TGIs of 90% and 96% with binimetinib at 3 mg / kg / 1.5 mg / kg compound 1 BID and binimetinib at 3 mg / kg / 3 mg / kg compound 1 BID, respectively, Figures 8A and 8C, Table 13.

[0084] TGI was statistically significantly greater than vehicle-treated animals after all treatments (p=0.0041 for binimetinib monotherapy, p<0.0001 for all other treatments, Table 12), and TGI after combination therapy with Compound 1 and binimetinib was significantly greater than that seen with binimetinib monotherapy (p<0.0001 for both dose levels of the combination therapy). Importantly, treatment with Compound 1 plus binimetinib was well tolerated, with only two animals at the 3 mg / kg binimetinib / 1.5 mg / kg Compound 1 BID dose level and no animals at the 3 mg / kg binimetinib / 3 mg / kg Compound 1 BID dose level experiencing weight loss of more than 10% (see FIG. 8B).

[0085] [Table 12]

[0086] Summary of Results. Low-dose compound 1 (1.5 or 3 mg / kg BID) was well tolerated and effective when administered orally BID for 14 days in the NRAS mutant and BRAF mutant melanoma xenograft model, WM3629, either as monotherapy or in combination with the MEK inhibitor, binimetinib (3 mg / kg BID). Low-dose compound 1 monotherapy resulted in modest tumor growth inhibition (TGI, mean 66-79%), whereas the combination of compound 1 with binimetinib resulted in significantly enhanced TGI (mean 90-96%, p<0.0001) compared to binimetinib alone (mean TGI, 37%). These data indicate that compound 1 can be safely and effectively combined with MEK inhibitor therapy for combined antitumor benefit in NRAS mutant melanoma.

[0087] Conclusions: Combination therapy of compound 1 with the MEK inhibitor binimetinib was well tolerated and resulted in significant tumor inhibition in xenograft models of NRAS mutant, class III BRAF mutant melanoma, WM3629 (mean TGI of 90% and 96% for 3 / 1.5 mg / kg BID and 3 / 3 mg / kg BID binimetinib / compound 1, respectively). Tumor inhibition with compound 1 / binimetinib combination therapy was significantly greater than binimetinib monotherapy (p<0.0001). This study demonstrates the antitumor combination benefit of adding a MEK inhibitor to compound 1 treatment in a human melanoma model harboring an NRAS mutation.

[0088] Example 7. Evaluation of the antitumor activity of Compound 1 in a BRAF-mutated NSCLC xenograft model

[0089] Objective: To evaluate the tolerability and antitumor activity of compound 1, a selective and potent inhibitor of rapidly accelerated fibrosarcoma (RAF) kinase, in RAF kinase homolog B (BRAF) mutant human non-small cell lung cancer (NSCLC) cell line-derived xenograft models.

[0090] Method: Athymic mice were injected with 1x10 7Human cancer cells were inoculated subcutaneously. Compound 1 was administered orally (PO) twice daily (BID) to mice with tumor volumes (TV) of approximately 300 mm 3 The treatment period began when the tumor volume reached 10 μg / kg and continued for 3 weeks. Animals were monitored for tumor growth, weight change, and general health / behavior.

[0091] The average TV is about 300mm 3 Groups and treatments were initiated when the mice reached a starting TV and body weight, with the average values ​​being the same for each treatment group. The test groups and number of animals per group are shown in Table 13.

[0092] [Table 13]

[0093] Mice were implanted with 1x10 cells in 0.1 mL of 1:1 medium / Matrigel on the right flank for tumor development. 7 NCI-H2405 cells were inoculated subcutaneously. Tumor-bearing mice were treated according to the study design shown in Table 13. For routine monitoring, all study animals were monitored for tumor growth as well as any abnormalities in behavior and appearance, such as mobility, food and water consumption, body weight, eye appearance, and any other abnormal observations. Any mortality and / or abnormal clinical signs were recorded. All animals were weighed and recorded every other day throughout the study. Tumor size measurements were made with calipers and recorded three times in the first week, and twice weekly in the second and third weeks. TV (mm 3 ) into the formula, TV=axb 2 / 2, where "a" and "b" are the long and short diameters of the tumor, respectively. The TGI was calculated using the following formula:

[0094]

number

[0095] Patients were excluded if they showed overt signs of severe distress and / or pain, showed significant weight loss (weight loss >20%), were unable to reach adequate food or water, were otherwise observed to be in a state of continued deterioration, or had tumors measuring ≥2,500 mm 3 Animals were euthanized if the group mean TV was >2,500 mm 3 All animals in a given study group were euthanized when 0.01 mg / kg / day was reached. Plasma and tumor tissue samples were collected for pharmacokinetic and pharmacodynamic analyses.

[0096] Results. The antitumor activity of Compound 1 was evaluated in human NCI-H2405 (class II BRAF mutant) NSCLC xenograft models. Treatment with Compound 1 (1.5, 3, 10, or 15 mg / kg twice daily [BID]) reduced tumor size by approximately 300 mm 3 (The actual mean TV for all groups was 304 mm. 3 ) and continued BID for 3 weeks.

[0097] Dose-dependent exposure and antitumor responses to Compound 1 treatment were observed. AUC ranged from 2878 to 50799 hr*ng / mL. 0-24 Values ​​were achieved at the doses tested, and tumor growth inhibition (TGI) was observed compared to controls (vehicle-treated animals) (Figure 9A, Figure 9C). The mean TGI with increasing doses of 1.5, 3, 10, or 15 mg / kg BID was 42%, 58%, 85%, and 107%, respectively (Table 14). Statistically significant reductions in tumor growth were achieved with Compound 1 at doses of 1.5, 3, 10, and 15 mg / kg BID (p<0.0001 for all comparisons, Table 14). Importantly, treatment with Compound 1 was well tolerated at all doses tested, and no group showed a weight loss of >10% during the study (Figure 9B).

[0098] [Table 14]

[0099] Summary of Results. Compound 1 (1.5-15 mg / kg BID) was well tolerated and efficacious when administered orally for 21 days in a BRAF mutant NSCLC xenograft model, NCI-112405. Dose-dependent tumor growth inhibition (TGI) was observed with compound 1 treatment twice daily (BID), with low doses of compound 1 (1.5-3.0 mg / kg BID) resulting in moderate TGI (average 42-58%) and higher compound 1 doses (10-15 mg / kg BID) resulting in strong TGI (average 85-107%). TGI in all compound 1 treatment groups was statistically significantly different from vehicle-treated animals (p<0.0001). These data indicate that compound 1 can be used safely and effectively for potent antitumor benefit in BRAF mutant NSCLC.

[0100] Example 8. Use of Compound 1 in Human Clinical Trials Name: Phase 1 / 1b open-label, multicenter study to investigate safety, Tolerability, Pharmacokinetics, and Antitumor Activity of Compound 1 in Participants with BRAF and / or NRAS Mutation-Positive Solid Tumors

[0101] Trial Phase: Phase 1 / 1b

[0102] Indications: BRAF mutation-positive solid tumors and / or NRAS mutation-positive solid tumors

[0103] Introduction: This is a two-part, open-label, multicenter, dose-escalation and dose-expansion study designed to evaluate the safety, tolerability, and pharmacokinetics (PK) of Compound 1, a pan-rapidly accelerated fibrosarcoma (RAF) small molecule kinase inhibitor, in participants with rapidly accelerated fibrosarcoma, homolog B (BRAF) mutation-positive and / or neuroblastoma RAS (NRAS) mutation-positive tumors, to determine a recommended phase 2 dose (RP2D) of Compound 1 for further clinical development, and to evaluate objective responses to Compound 1 treatment alone and in combination with binimetinib, a mitogen-activated protein kinase (MEK) inhibitor.

[0104] Test Purpose

[0105] Part A. The primary objective of the Part A dose-escalation Compound 1 monotherapy component of the study (hereafter referred to as Part A1) is to determine the safety and tolerability of oral administration of Compound 1, including dose-limiting toxicities (DLTs), in participants with BRAF mutation-positive advanced or metastatic solid tumors or melanoma with NRAS mutations. Also, to identify the maximum tolerated dose (MTD) and / or appropriate dose for further clinical investigation in Part B dose expansion. Secondary objectives include characterization of PK properties and the effect of food on the PK of Compound 1. The primary objective of the Part A dose-escalation Compound 1 + binimetinib combination component of the study (hereafter referred to as Part A2) is to determine the safety and tolerability of oral administration of Compound 1 + binimetinib, including DLTs, in participants with BRAF class II or III mutation-positive advanced or metastatic solid tumors or melanoma with NRAS mutations, as well as to identify the MTD and / or appropriate dose for further clinical trials. Secondary objectives include characterization of the PK properties of the combined Compound 1 and binimetinib.

[0106] Part B. The primary objective of the Part B dose expansion portion of the study is to evaluate preliminary evidence of anticancer activity of Compound 1 in participants with advanced or metastatic solid tumors harboring either class II or class III BRAF genomic alterations. Secondary objectives include further evaluation of the safety, tolerability, and PK of Compound 1 at RP2D.

[0107] Study design

[0108] Part A (dose escalation)

[0109] Part A will include participants with solid tumors with any BRAF class I, class II, or class III mutation and / or NRAS mutation-positive melanoma who meet all protocol-defined eligibility requirements. The number of participants with tumors harboring BRAF class I mutations enrolled in Part A of the study will be monitored on an ongoing basis.

[0110] Part A1 will evaluate the safety, tolerability, PK and pharmacodynamics (PD) of Compound 1 in participants with BRAF mutation-positive advanced or metastatic solid tumors and / or melanoma with NRAS mutations. This component combines an accelerated titration design (single participant dose level) followed by a traditional 3+3 dose escalation schema to identify the MTD and / or RP2D of Compound 1. The MTD is defined as the highest daily oral dose at which less than 33% of participants experience a DLT during the 28-day DLT evaluation period. Compound 1 will be administered as an oral dose twice daily (BID) for 28 days in a 28-day treatment cycle.

[0111] The starting dose, dose level 1 (DL1), is 50 mg / day (administered as 25 mg BID), with dose escalation increments following a modified Fibonacci sequence. Planned dose levels of Compound 1 are indicated in the protocol.

[0112] An accelerated dose escalation design principle is used in which one participant per cohort is evaluated until evidence of biological activity is observed, at which point the cohort is expanded by two participants. This, and all subsequent dose level cohorts, will progress using a 3+3 dose escalation design. For purposes of the above discussion, "evidence of biological activity" is defined as either (1) a DLT or (2) at least one grade ≥2 adverse event (AE) not clearly attributable to underlying disease or to exogenous causes (excluding grade 2 laboratory investigations where the AE is deemed non-clinically significant by the investigator).

[0113] Starting from DL3, the dose of Compound 1 is escalated using a conventional 3+3 study design, continuing until the stopping rule is met or the dose at which the MTD or maximum expected pharmacological activity (as informed by PK and PD biomarkers) is achieved. In this 3+3 design, at least three participants are enrolled at each dose level from DL3 onwards. If none of the three participants at a dose level experience a DLT within the DLT period (28 days from the first dose of Compound 1 for all dose levels), another three participants are treated at the next higher dose level. If two or more participants at that dose level develop a DLT in cycle 1, no further participants are treated at a given dose level.

[0114] For the first two participants to receive Compound 1 at a particular dose level, there will be at least a 24-hour interval between the first dose of Compound 1 in the first and second participants. Dose escalation will continue until the highest planned dose level is determined to be safe and tolerable in a minimum of six DLT-evaluable participants at that dose level (i.e., the dose level is considered to be the RP2D) or until the MTD is reached. A maximum of approximately 26 participants are required to estimate the MTD and / or RP2D.

[0115] A Dose Review Committee (DRC), consisting of Investigators and Sponsor representatives, will review all available safety, PK and PD data before initiating enrollment at the next dose level. The DRC will review the safety and tolerability of each dose level of Compound 1 monotherapy after participants at that particular dose level have completed at least one full cycle. Part A will continue until stopping rules are met or until the MTD or dose at which the expected maximum pharmacological activity (e.g., RP2D) is achieved. The DRC will also evaluate all available safety and PK / PD data to determine the RP2D for Part B.

[0116] If a participant receiving Compound 1 monotherapy experiences disease progression during treatment, the participant may be given the option to receive the combination of Compound 1 + binimetinib based on the participant's eligibility for combination therapy, experience with Compound 1 treatment, response characteristics, or other relevant considerations. This decision is at the investigator's discretion but should be made in consultation with the sponsor.

[0117] Part A2 will not begin until preliminary proof of mechanism (PPOM) clinical criteria are met in the monotherapy dose escalation cohort. Eligibility will be limited to participants with class II / III BRAF and / or NRAS mutated tumors. Parallel escalation will begin with compound 1 and binimetinib (binimetinib at a dose of 45 mg [PO] BID). The starting dose of combined compound 1 (designated combination dose level 1) will be one dose level below the dose level at which the PPOM criteria were met or one dose level below the highest dose level deemed safe, at the discretion of the DRC. Dose escalation will follow the 3+3 design described in the protocol, with binimetinib administered together as a daily oral dose BID in 28-day treatment cycles in participants with class II / III BRAF and NRAS mutated tumors.

[0118] At the end of each dosing cohort, the DRC is responsible for determining whether the criteria for PPOM have been met and for recommending whether combination dose escalation should be initiated. When combination dose escalation is initiated, the specific Compound 1 dose is increased according to a modified Fibonacci design, and an intermediate dose may be selected in practice.

[0119] At the investigator's discretion and after consultation with the sponsor, participant intravenous dose escalation and backfills are permitted in Part A1. Participant intravenous dose escalation may only be considered if the participant meets all protocol-specified criteria.

[0120] The effect of food on the PK of Compound 1 will be evaluated using a randomized crossover design in at least 6 participants during Part A1 in participants in DL3, DL4, and DL5.

[0121] Part B (dose expansion)

[0122] The dose expansion portion of the study (Part B) may begin once the MTD and / or biologically active dose (i.e., RP2D) has been determined from the Compound 1 monotherapy arm in Part A. Part B will evaluate the antitumor activity of Compound 1 at the RP2D determined from Part A in the following cohorts: Cohort 1: Participants with unresectable locally advanced (American Joint Committee on Cancer [AJCC] stage III) or metastatic (AJCC stage IV) non-small cell lung cancer (NSCLC) with BRAF class II or class III mutations Cohort 2: Participants with unresectable and locally advanced (AJCC stage III) or metastatic (AJCC stage IV) melanoma with any BRAF class II or class III mutation Cohort 3: Participants with any unresectable locally advanced (AJCC stage III, or equivalent stage according to other staging systems) or metastatic (AJCC stage IV) solid tumor (other than NSCLC or melanoma) with any BRAF class II or class III mutation Enrollment of participants into the dose expansion cohorts will occur simultaneously. Enrollment of participants with various tumor types, including BRAF class II or class III mutations, will be monitored over the course of the study in Cohort 3. Enrollment of specific tumor types may be limited to ensure broad representation of various solid tumors in this cohort.

[0123] Study endpoints

[0124] primary

[0125] Safety endpoints included the following: Incidence of DLT Incidence of AEs, including treatment-emergent and treatment-related AEs Clinically significant changes in vital signs, physical examination, electrocardiogram, and laboratory tests

[0126] Efficacy (as assessed by the investigator) will be measured by: Objective response rate, defined as the proportion of complete responses plus partial responses per Response Evaluation Criteria in Solid Tumors (RECIST) v1.1 as assessed by the investigator Disease control rate Duration of overall response Duration of stable disease

[0127] secondary Pharmacokinetic parameters for Compound 1 and Compound 1 + binimetinib include, but are not limited to, maximum observed plasma concentration (Cmax), time to reach Cmax (tmax), and area under the plasma concentration-time curve (AUC), including fed and fasted states.

[0128] search Compound 1 and Compound 1 + Binimetinib Exposure / Safety and Exposure / Efficacy Relationships Characterization of potential metabolites of compound 1 in plasma and urine Progression-free survival and overall survival Quantification of biomarkers by biochemical and / or genetic analysis of blood and / or tumor samples, including, but not limited to, pharmacodynamic effects on phospho-extracellular signal-regulated kinase (ERK), DUSP6 ribonucleic acid (RNA) and the proliferation marker Ki-67. Changes in circulating tumor-derived (blood) nucleic acid (ctDNA) concentrations and mutation profiles Potential biomarkers by biochemical and / or genetic analysis of blood and / or tumor samples ·Collective PK Analysis

[0129] Sample size Approximately 155 participants are expected to be enrolled in the study.

[0130] Part A (dose escalation)

[0131] In Part A1, up to 41 participants will be enrolled in the monotherapy dose escalation phase. Approximately 26 participants will be enrolled to estimate the MTD and / or RP2D, from which 6 participants will be evaluated for the effect of food on the PK of Compound 1.

[0132] For Part A2, up to 36 participants will be enrolled in the combination dose escalation phase. Approximately 24 participants may be required to estimate the MTD of the combination of Compound 1 + binimetinib.

[0133] Approximately 30 additional participants may be enrolled for backfill across Parts A1 and A2.

[0134] Part B (dose expansion)

[0135] Approximately 75 participants will be enrolled in Part B, which will include three cohorts of participants with BRAF class II or III mutations, including participants with NSCLC, melanoma, and other solid tumors. A Simon two-stage optimal design is planned for cohorts 1 and 2. 12 participants will be enrolled in stage 1. If ≥3 participants respond, it will proceed to stage 2, where the cohort will enroll an additional 13 participants for a total sample size of 25. At the end of stage 2, if ≥8 of the 25 participants respond, compound 1 will be considered promising for further evaluation. Within each cohort, the design will have 80% power and a maximum type-1 error rate of 0.1 to test the null hypothesis that the response rate is ≤20% against the alternative hypothesis of ≥40%. For futility assessment in stage 1, it may not be necessary to confirm the response. Additional considerations beyond the response rate may also be included at the discretion of the DRC and sponsor.

[0136] Key inclusion and exclusion criteria

[0137] Key inclusion criteria included:

[0138] Adult participants (≥18 years of age) with a histologically or cytologically confirmed diagnosis of metastatic or advanced malignancy are eligible for this study. Part A, dose escalation, will enroll participants with any type of advanced or metastatic solid tumor. Part B, dose expansion, will enroll participants with unresectable and locally advanced (AJCC stage III) or metastatic (AJCC stage IV) NSCLC, unresectable and locally advanced (AJCC stage III) or metastatic (AJCC stage IV) melanoma, or any unresectable and locally advanced (AJCC stage III, or equivalent stage according to other staging systems) or metastatic (AJCC stage IV) solid tumor other than NSCLC or melanoma. In addition, genetic abnormalities of BRAF and / or NRAS are required, as specified in the inclusion criteria. Participants must have received previous standard of care appropriate for their tumor type and stage of disease, or in the opinion of the investigator, and be resistant to or unlikely to derive clinically meaningful benefit from appropriate standard therapy. In Part A1, participants with certain cancers driven by BRAF class I mutations (NSCLC, melanoma, colorectal cancer [CRC], and anaplastic thyroid cancer) must have previously received an approved BRAF inhibitor (with or without an approved MEK inhibitor, if approved and available).

[0139] Participants' tumors must harbor a BRAF mutation or be melanomas with an NRAS mutation identified by prior genomic analysis of tumor tissue or ctDNA performed in a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory (in the United States [US]) or in accordance with local regulatory requirements (in other countries). Participants will provide archived tumor tissue specimens (formalin-fixed paraffin-embedded [FFPE] specimens) obtained within the last 5 years (if available) and undergo mandatory pre-treatment tumor biopsies, if medically feasible. A complete list of inclusion / exclusion criteria will be included in the protocol. Key exclusion criteria include:

[0140] Participants with known active brain metastases from non-brain tumors are not eligible. Participants in Part A with other solid tumors driven by BRAF class I mutations (non-NSCLC, melanoma, CRC, and anaplastic thyroid cancer) may not have had prior treatment with any approved or investigational small molecule BRAF-, MEK-, or mitogen-activated protein kinase (MAPK)-directed inhibitor therapy. This criterion will not be implemented until a decision to do so is made and communicated by the DRC. In Part B, prior treatment with any approved or investigational small molecule BRAF-, MEK-, or MAPK-directed inhibitor therapy is excluded. Participants may not have any unresolved toxicity from prior antitumor therapy. Participants may not have any unresolved toxicity from prior antitumor therapy. Exclusionary concomitant medications, contraceptive requirements, and diseases and other conditions that would exclude participants from the study will be detailed in the protocol.

[0141] Treatment Duration Participants will receive Compound 1 or Compound 1 + binimetinib in 28-day cycles until evidence of disease progression, unacceptable toxicity, intolerance to study drug, initiation of new systemic cancer therapy, withdrawal of consent, Investigator's decision, Sponsor's decision, or death.

[0142] Statistical Consideration Descriptive statistics are provided for selected demographic, safety, PK, efficacy and biomarker data by dose, dose schedule and time as appropriate. Descriptive statistics for continuous data include mean, median, standard deviation and range, whereas categorical data are summarized using frequency counts and percentages. Objective response rates are presented with 95% exact confidence intervals. Graphical summaries of the data may also be presented.

Claims

1. A composition for treating cancer in a patient in need of such treatment, comprising (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof.

2. A pharmaceutical composition for treating cancer in a patient in need of such treatment, comprising (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, and at least one pharma- ceutically acceptable excipient.

3. The composition of claim 1 or 2, wherein the cancer is characterized as having an oncogenic BRAF alteration.

4. The composition of claim 3, wherein the oncogenic BRAF alteration is a class I BRAF mutation, a class II BRAF mutation, or a class III BRAF mutation.

5. The composition of claim 3, wherein the oncogenic BRAF alteration is a BRAF V600 mutation.

6. 6. The composition of claim 5, wherein the BRAF V600 mutation is a V600E or V600K mutation.

7. The composition of claim 1 or 2, wherein the cancer is characterized as having wild-type BRAF.

8. The composition of claim 1 or 2, wherein the cancer is a solid tumor.

9. 3. The composition of claim 1 or 2, wherein the cancer is selected from melanoma, pancreatic cancer, pancreatic adenocarcinoma, ovarian cancer, colorectal cancer, glioma, Langerhans cell histiocytosis, leukemia, hairy cell leukemia, thyroid cancer, anaplastic thyroid cancer, papillary thyroid cancer, follicular thyroid cancer, or medullary thyroid cancer.

10. The composition of claim 1 or 2, wherein the cancer is selected from small cell lung cancer, non-small cell lung cancer, prostate cancer, gastric cancer, and esophageal cancer.

11. The composition of claim 1 or 2, wherein the cancer is NRAS mutant melanoma.

12. The composition of claim 1 or 2, wherein the cancer is cholangiocarcinoma.

13. The composition of claim 1 or 2, wherein the cancer is metastatic.

14. The composition described in claim 1 or 2, wherein the treatment is adjuvant chemotherapy followed by surgical resection.

15. The composition of claim 1 or 2, wherein the patient is relapsing after a previous treatment.

16. The composition of claim 1 or 2, wherein the patient has acquired resistance to a previous treatment.

17. The composition of claim 1 or 2, wherein the patient is resistant to treatment. (a) (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof; (b) at least one tumor therapeutic selected from a MEK inhibitor, an immune checkpoint inhibitor, a CDK inhibitor, an EGFR kinase inhibitor, an EGFR antibody, an EGFR PROTAC therapeutic, an FGFR inhibitor, an SOS1 inhibitor, an SHP2 inhibitor, a KRAS inhibitor, a taxane, a topoisomerase inhibitor, an ERK inhibitor, a platinum-based chemotherapy drug, folinic acid, 5-fluorouracil, or an anti-VEGF therapeutic; 13. A composition for treating cancer in a patient in need of such treatment comprising:

19. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a MEK inhibitor.

20. 20. The composition of claim 18 or 19, wherein the MEK inhibitor is selected from binimetinib, trametinib, cobimetinib, selumetinib, pimasertib, or mirdametinib.

21. The composition described in claim 18 or 19, wherein the MEK inhibitor is binimetinib.

22. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is an immune checkpoint inhibitor.

23. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a CDK inhibitor.

24. 19. The composition of claim 18, wherein the at least one tumor therapeutic agent is an EGFR kinase inhibitor or an antibody.

25. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is an SHP2 inhibitor.

26. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a SOS1 inhibitor.

27. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is an FGFR inhibitor.

28. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a KRAS inhibitor.

29. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a taxane.

30. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a topoisomerase inhibitor.

31. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is an ERK inhibitor.

32. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is a platinum-based chemotherapy agent.

33. 20. The composition of claim 18, wherein the at least one tumor therapeutic agent is folinic acid.

34. The composition of claim 18, wherein the at least one tumor therapeutic agent is 5-fluorouracil.

35. 20. The composition of claim 18, wherein the at least one tumor therapeutic is an anti-VEGF therapeutic.

36. 20. The composition of claim 18, wherein the cancer is characterized as having an oncogenic BRAF alteration.

37. The composition of claim 18, wherein the cancer is characterized as having wild-type BRAF.

38. The composition of claim 18 , wherein the cancer is a solid tumor.

39. 20. The composition of any one of claims 1, 2 or 18, wherein the (S)-N-(3-(2-(((R)-1-hydroxypropan-2-yl)amino)-6-morpholinopyridin-4-yl)-4-methylphenyl)-3-(2,2,2-trifluoroethyl)pyrrolidine-1-carboxamide, or a pharma- ceutically acceptable salt or solvate thereof, is administered orally.

40. 40. The composition of claim 38, wherein the oral administration occurs every other day, once a day, twice a day, or three times a day.

Images