Novel RAS inhibitor

Abstract

The present invention relates to the use of the compound of formula (I) as a RAS inhibitor and as a medicament, in particular for the treatment of proliferative diseases, inflammatory diseases and / or genetic diseases. The present invention further relates to a pharmaceutical composition comprising the compound of formula (I). Furthermore, the present invention relates to a method for inhibiting the growth, proliferation or metastasis of cancer cells in a subject in need thereof, in particular a subject who may include some patients defined by the mutation status of the RAS oncogene, or patients who have developed resistance to treatment with standard therapy or RAS mutation-specific inhibitors. The present invention also relates to a method for inhibiting RAS molecules in the treatment of genetic diseases such as RAS syndrome or inflammatory diseases such as adenomyosis in which the KRAS gene is activated by mutation. In addition, the present invention relates to a method for inhibiting the growth and / or secretion of factors from a cell population sensitive to inhibiting the activation of RAS in vitro, in particular a cell population sensitive to inhibiting the activation of KRAS, HRAS and NRAS in vitro. Furthermore, the present invention relates to a kit comprising a formulation of a pharmaceutical composition containing the compound of formula (I).

Description

Technical Field

[0001] The present invention relates to the use of the compound of formula (I) as a RAS inhibitor and as a medicament, in particular for the treatment of proliferative diseases, inflammatory diseases and / or genetic diseases. The present invention further relates to a pharmaceutical composition comprising the compound of formula (I). Furthermore, the present invention relates to a method for inhibiting the growth, proliferation or metastasis of cancer cells in a subject in need thereof, in particular a subject that may include some patients defined by the mutational status of the RAS oncogene, or patients who have developed resistance to treatment with standard therapy or RAS mutation-specific inhibitors. The present invention also relates to a method for inhibiting RAS molecules in the treatment of genetic diseases such as RAS syndrome or inflammatory diseases such as adenomyosis in which the KRAS gene is activated by mutation. The present invention also relates to a method for inhibiting the growth and / or secretion of factors from a cell population sensitive to inhibiting the activation of RAS in vitro, in particular a cell population sensitive to inhibiting the activation of KRAS, HRAS and NRAS in vitro. Furthermore, the present invention relates to a kit comprising a formulation of a pharmaceutical composition containing the compound of formula (I).

Background Art

[0002] RAS proteins represent a group of closely related monomeric globular proteins that are associated with the plasma membrane and can bind to either GDP or GTP. RAS containing bound GDP represents an "inactive" state, but the binding of RAS to GTP instead of GDP represents an "active" state, as a result of which the protein can interact with other "effector" proteins of downstream targets. RAS proteins can be regarded as small GTPases that function as molecular switches controlling the transmission of extracellular signals from the extracellular to the nucleus by various effector proteins.

[0003] There are three types of RAS isoforms (KRAS, HRAS, and NRAS), and their activation cycles are regulated by the binding of GDP or GTP, which is controlled by GAP or GEF. Their GTP-bound forms bind to their effector proteins, inducing multiple signaling pathways that control various basic cellular processes.

[0004] Normally, RAS mutations lead to defects in GTP hydrolysis via GAP, resulting in the accumulation of the GTP-bound active state of RAS. This leads to uncontrolled growth, which is characteristic of cancer cells. Uncontrolled RAS activation is also detected in hereditary diseases such as RAS syndrome. Activation of RAS mutations has been detected in inflammatory diseases such as adenomyosis / endometriosis, which contributes to the growth and invasion of endometrial cells and resistance to progesterone (S. Inoue, Nature Comm. 2019, 10, 5785; PMID: 31857578).

[0005] Recent research has led to the development of mutant-specific KRAS inhibitors targeting the KRASG12C variant, which have been approved for clinical use. Further inhibitors targeting other mutant-specific versions of KRAS are currently under development.

[0006] Furthermore, patients are known to frequently develop resistance to KRAS oncogene inhibitors, such as KRAS G12C inhibitors (Tanaka et al., Cancer Discov, 2021, PMID33824136). Also, patients treated with KRAS G12C inhibitors often develop secondary mutations in other RAS isoforms (Awad MM et al., New England J. Med., 2021, PMID34161704).

[0007] Quinacrine is a drug with several uses. The main uses of quinacrine are as an antiprotozoal agent, an anti-rheumatic agent, and a sclerosing agent for the pleura.

[0008] Oien et al., Seminar in Cancer Biology, 68(25), 2019, p21 discloses the use of quinacrine as an anticancer agent. This is selective for cancer cells and shows synergistic effects when used in combination with other anticancer agents.

[0009] KR1020200114680 discloses a composition containing 5-fluorouracil and quinacrine for the treatment of cancer.

[0010] Kalogera et al., Gyn.Onco.146(1), 2017, p.187 relates to the use of quinacrine for inhibiting tumor formation in endometrial cancer.

[0011] Guo et al., Ongonene 28(8), 2009, p.1151 relates to the use of 9-aminoacridine and its derivative quinacrine as anticancer agents targeting the PI3K / AKT / mTOR, NF-kB and p35 pathways.

[0012] Bonse et al., J.Med.Chem.42(26), 1999, p.5488 relates to the kinetic study of 9-amino- and 9-thioacridine for inhibiting trypanothione reductase.

[0013] Irvin et al., J.Am.Chem.Soc., 72(6), 1950, p.2763 and Hammick et al., J.of Chem.So., 1950, p.346 relate to the physicochemical property test of quinacrine.

[0014] In contrast, the compounds according to the present invention function by a mechanism other than those described in the above literature. The data in this specification suggest that the compound may uncouple active RAS from its effector within the plane of the plasma membrane in the cell. This event is much closer to the activation of downstream kinases in the signal transduction cascade. It cannot be predicted that the activation of the three isoforms will also be inhibited when the effector molecule is inhibited by a compound, such as quinacrine dihydrochloride.

[0015] None of the above documents disclose or suggest that quinacrine dihydrochloride is suitable for use as an inhibitor of RAS protein activation involving KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96C, KRAS G12C / Y96DS, KRAS G13C, KRAS G13D, KRASG13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K.

[0016] To date, it has not been known that quinacrine dihydrochloride hydrate inhibits the expression and / or activation of RAS. Activation is defined as the ability of RAS to bind GTP-dependently to its effector molecules (RAF kinase, PI3K kinase, etc.) via the RAS-binding domain (RBD) or RAS-related domain (RA domain, etc.) present within the effector protein (RASSF, etc.). Furthermore, targeting activation rather than stability may be advantageous for certain medical conditions, but targeting both may be useful in combating specific subtypes of RAS mutant cancers.

[0017] In addition, the mechanisms that promote the activation of HRAS, NRAS, and KRAS are different, and each RAS isoform exhibits different functions and biological specificities. Therefore, efforts have been made to target HRAS and NRAS in specific tumor subtypes in which HRAS and NRAS function as cancer drivers.

[0018] Finally, targeting other RAS isoforms such as HRAS and NRAS is necessary to combat secondary and acquired resistance to standard treatments including some cancer therapeutics.

[0019] KRASG12-specific C inhibitors have been developed, but targeting HRAS and NRAS in cancers in which these isoforms are mutated remains a challenge.

[0020] However, effective targeting, particularly inhibition of RAS oncogene activation by small molecules with limited toxicity, remains a challenge.

[0021] Accordingly, an object of the present invention is to provide a pharmaceutically active compound having the ability to inhibit RAS oncogene activation, particularly with high specificity at low concentrations, especially in tumor cells. This object is achieved by the use of a compound of formula (I). Surprisingly, it has been found that the compounds of formula (I), particularly the compounds of formula (A), inhibit the activity of RAS oncogenes, particularly KRAS, HRAS and NRAS, regardless of their mutant states. Even more surprisingly, the compounds of formula (I), particularly the compounds of formula (A), uncouple GTP-dependent active RAS-effector interactions (unknown mutations). This could not be predicted from the prior art. Without being bound by any theory, it is assumed that the interaction between RAS activated by mutation and its effector molecule is disrupted. SUMMARY OF THE INVENTION

[0022] The present invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of diseases involving RAS signaling, JPEG2025524343000002.jpg6180wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0023] In particular, the present invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases involving RAS signaling, JPEG2025524343000003.jpg6180wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0024] In particular, the present invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of genetic diseases involving RAS signaling, in particular for use as an inhibitor of RAS protein activation for the treatment and / or prevention of RAS syndrome, JPEG2025524343000004.jpg6180wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0025] In particular, the present invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of inflammatory diseases involving RAS signaling, in particular for use as an inhibitor of RAS protein activation for the treatment and / or prevention of endometriosis and / or adenomyosis when KRAS is mutated, JPEG2025524343000005.jpg6180wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0026] In particular, the present invention relates to a compound of formula (I) as defined above and below or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative and / or inflammatory diseases involving KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K. JPEG2025524343000006.jpg6180 wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0027] In particular, the present invention relates to a compound of formula (I) as defined above and below or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative and / or inflammatory diseases that are resistant to treatment with RAS mutation-specific inhibitors different from the compound of formula (I), preferably resistant due to secondary mutations, particularly secondary mutations involving KRAS G12C / Y96D, KRAS G12C / Y96C, and / or KRAS G12C / Y96S. JPEG2025524343000007.jpg6180 wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0028] The present invention particularly relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation. JPEG2025524343000008.jpg6180 wherein, R 1 is H, C1-C2 alkyl; R 2 is H, C1-C2 alkyl; n is 1, 2, 3, or 4.

[0029] The present invention further relates to a composition (1) comprising at least one compound of formula (I) as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt, and a RAS mutation-specific inhibitor. The RAS mutation-specific inhibitor is different from the compound of formula (I).

[0030] The present invention further relates to a compound (I) as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), for use as a RAS-RAF inhibitor, or a composition (1) as defined above and below.

[0031] The present invention further relates to a compound (I) as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), for use as a medicament, or a composition (1) as defined above and below.

[0032] The present invention further relates to a compound (I) according to the present invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), for treating and / or preventing a disease, or a composition (1) as defined above and below.

[0033] The present invention further relates to a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), or a composition (1) as defined above and below, for use in the treatment of proliferative diseases.

[0034] The present invention further relates to a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), or a composition (1) as defined above and below, for use in the treatment of genetic diseases.

[0035] The present invention further relates to a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), or a composition (1) as defined above and below, for use in the treatment of RAS syndromes.

[0036] The present invention further relates to a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), or a composition (1) as defined above and below, for use in the treatment of inflammatory diseases.

[0037] The present invention further relates to the use of the compound (I) according to the present invention as defined above and below, in particular the compound of formula (A), or a pharmaceutically acceptable salt thereof, or the composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), for the treatment of endometriosis and / or adenomyosis, more specifically in the case where KRAS is mutated.

[0038] The present invention further relates to the use of the compound (I) according to the present invention as defined above and below, in particular the compound of formula (A), or a pharmaceutically acceptable salt thereof, or the composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), for the treatment of cancer.

[0039] The present invention further relates to the use of the compound (I) according to the present invention as defined above and below, in particular the compound of formula (A), or a pharmaceutically acceptable salt thereof, or the composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), as an inhibitor of RAS protein (KRAS, HRAS or NRAS oncogene) activation.

[0040] The present invention further relates to the use of the compound (I) according to the present invention as defined above and below, in particular the compound of formula (A), or a pharmaceutically acceptable salt thereof, or the composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular one selected from compound (A), for the treatment or prevention of any disease or condition associated with the activity of RAS protein (RAS oncogene).

[0041] The present invention further relates to the use of a compound (I) according to the present invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), for the treatment of proliferative diseases, inflammatory diseases and / or genetic diseases in which RAS signaling is involved, preferably KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K is involved.

[0042] The present invention further relates to a pharmaceutical composition comprising at least one compound (I) according to the present invention as defined above and below, in particular a compound of formula (A), or a composition (1) as defined above and below, or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, and a pharmaceutically acceptable carrier.

[0043] The present invention further relates to the use of a RAS protein activation inhibitor for the treatment and / or prevention of proliferative diseases and / or inflammatory diseases that are resistant to treatment with a RAS mutation-specific inhibitor different from the compound of formula (I), preferably resistant due to secondary mutations, in particular secondary mutations in which KRAS G12C / Y96D, KRAS G12C / Y96C, and / or KRAS G12C / Y96S are involved, for the pharmaceutical composition as defined above and below.

[0044] The present invention further relates to a pharmaceutical composition comprising at least one compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a pharmaceutically acceptable salt thereof, or a composition (1) as defined above and below, said pharmaceutical composition also comprising a further active substance, preferably selected from chemotherapeutic agents, radiotherapy agents, immuno-oncology agents, and combinations thereof.

[0045] The present invention further relates to a method of inhibiting the growth, proliferation, or metastasis of cancer cells in a subject in need thereof, said method comprising administering to said subject a therapeutically effective amount of a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a therapeutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), or a composition (1) as defined above and below.

[0046] The present invention further relates to a method of inhibiting the proliferation of a cell population sensitive to inhibiting RAS activation in vitro or ex vivo, said method comprising contacting said cell population with a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a therapeutically acceptable salt thereof, or a composition (1) as defined above and below, or a pharmaceutical composition comprising at least one compound of formula (I) as defined above and below, in particular selected from compound (A), or a composition (1) as defined above and below.

[0047] [[ID=|11]] The present invention further relates to a kit comprising: a) a pharmaceutical composition comprising a compound (I) according to the invention as defined above and below, in particular a compound of formula (A), or a composition (1) as defined above and below, or a pharmaceutical composition comprising a compound (I) according to the invention as defined above or below, in particular a compound of formula (A), or a therapeutically acceptable salt thereof, or a composition (1) as defined above and below, and a pharmaceutically acceptable carrier; and b) instructions for administering a pharmaceutical composition for the treatment of a disease for which inhibition of RAS activation is therapeutically effective. Description of the Invention

[0048] The present invention has the following advantages: - The compounds according to the present invention exhibit advantageous RAS inhibitory properties. In other words, the compounds according to the present invention are eligible as inhibitors of RAS oncogene activation. The interaction between RAS and effectors is assumed to be disrupted, especially when RAS is activated by oncogenic somatic mutations. - The said compounds inhibit KRAS, regardless of mutations, at pharmacologically achievable concentrations in human patients. The resistance of these molecules has been well studied in other diseases. - The said compounds inhibit NRAS and HRAS by functionally decoupling their binding to their effectors in cells.

[0049] Compound of formula (I) In the context of the present invention, inactivation means inhibiting the activity of a protein, in particular an RAS protein, specifically an NRAS, KRAS or HRAS protein, based on a direct or indirect interaction between at least one compound of formula (I) and the protein. This inhibition includes inhibition involved in the complex with the RAS protein. The said interaction is not part of the translation process or the translation process.

[0050] Furthermore, activation means the ability of RAS to bind GTP-dependently to its effector molecule (RAF kinase, PI3K kinase, etc.) via the RAS binding domain (RBD) or RAS-related domain (RA domain, etc.) present in the effector protein (RASSF, etc.).

[0051] As used herein, the term "synergistic" or "synergy" refers to a combination of treatments that is more effective than the additive effect of two or more single agents. That is, the term "synergistic effect" refers to the effect of a given combination of two compounds in which the combined activity exceeds the sum of the individual activities of the compounds when administered separately. For this reason, the said combination can be used at a lower dosage rate based on the individual components and achieve a therapeutic effect comparable to that of the individual components.

[0052] The following equation is applied to measure whether a combination of a compound of formula (I) and at least one additional compound different from the compound of formula (I) exhibits a synergistic effect: E = X + Y - (X·Y / 100), wherein X = the effect (%) when the compound of formula (I) is used at a dosage ratio a; Y = the effect (%) when the compound of formula (II) is used at a dosage ratio b; E = the predicted effect (%) when the compound of formula (I) and the compound of formula (II) are used at a dosage ratio a + b. In the above equation, the value of E corresponds to the effect (inhibition) predicted when the activities of the individual compounds are additive. If the observed effect is higher than the value of E calculated according to the equation, a synergistic effect exists.

[0053] The term "proliferative disease" refers to a disease associated with some degree of abnormal cell proliferation. Preferably, the proliferative disease is cancer. Specifically, the cancer is selected from prostate, colon, rectum, pancreas, cervix, stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck, skin (including melanoma and basal cell carcinoma), mesothelium, white blood cells (including lymphoma and leukemia), esophagus, breast, muscle, connective tissue, lung (including small cell lung cancer and non-small cell cancer), adrenal gland, thyroid, kidney, or bone cancer; or glioblastoma, mesothelioma, renal cell carcinoma, gastric cancer, sarcoma (including Kaposi's sarcoma), choriocarcinoma, skin basal cell carcinoma, hematological malignancies (including blood, bone marrow, and lymph nodes), or testicular seminoma.

[0054] Unless otherwise specified herein, references in the singular may also include the plural. For example, "a" and "an" may refer to one, or one or more.

[0055] In the context of the present invention, the prefix C n ~C m indicates the number of carbon atoms that the molecule or residue thereby specified may contain.

[0056] In the context of the present invention, the expression "C1-C4 alkyl" refers to an unbranched or branched saturated hydrocarbon group having 1 to 4 carbon atoms. C1-C4 alkyl is, for example, methyl, ethyl, propyl, 1-methylethyl, butyl, 1-methylpropyl, 2-methylpropyl, 1,1-dimethylethyl.

[0057] The compounds of formula (I) and (A) form salts which are also within the scope of the present invention. Pharmaceutically acceptable (i.e., non-toxic and physiologically acceptable) salts are preferred, but other salts may also be useful, for example, in the isolation or purification steps used during the preparation. The salts of the compounds of formula (I) and (A) can be formed, for example, by reacting the compounds of formula (I) and (A) with at least one acid or base. The acid or base is added in an amount suitable for partial or complete neutralization, for example, an equivalent amount.

[0058] As used herein, the term "pharmaceutically acceptable salt" refers to salts containing pharmacologically acceptable anions or cations, unless otherwise indicated, for example, hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate (hydrogen sulfate), phosphate, hydrogen phosphate, dihydrogen phosphate, isonicotinate, acetate, lactate, salicylate, citrate, acid citrate, tartrate, pantothenate, hydrogen tartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, and pamoate [i.e., 4,4'-methylene-bis-(3-hydroxy-2-naphthoate)].

[0059] Preferred is a compound of formula (I) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation, wherein R 1 is C1-C2 alkyl; R 2 is C1-C2 alkyl; n is 3.

[0060] In particular, R 1 and R 2 have the same meaning.

[0061] In the first embodiment, the compound of formula (I) is compound (A), which is compound (A) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation. JPEG2025524343000009.jpg52116

[0062] Preferably, the present invention relates to compound (A) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of diseases involving RAS signaling.

[0063] In particular, the present invention relates to compound (A) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases, inflammatory diseases and / or genetic diseases involving RAS signaling.

[0064] Specifically, the present invention relates to compound (A) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases involving KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRASG13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K.

[0065] Specifically, the present invention relates to compound (A) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of inflammatory diseases involving KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K.

[0066] In another preferred embodiment, the present invention relates to compound (A) or a pharmaceutically acceptable salt thereof for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative and / or inflammatory diseases that are resistant to treatment with a RAS mutation-specific inhibitor different from the compound of formula (I), preferably resistant due to secondary mutations, particularly secondary mutations involving KRAS G12C / Y96D, KRAS G12C / Y96C, and / or KRAS G12C / Y96S.

[0067] A further aspect of the present invention is a composition (1) comprising at least one compound of formula (I) as defined above and below, or a pharmaceutically acceptable salt thereof, and at least one RAS mutation-specific inhibitor, wherein the RAS mutation-specific inhibitor is different from the compound of formula (I).

[0068] In one embodiment, the composition (1) comprises a compound (A) or a pharmaceutically acceptable salt thereof, and at least one RAS mutation-specific inhibitor, particularly a KRAS mutation-specific inhibitor, specifically sotorasib (also known as AMG510 or (1M)-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-4-[(2S)-2-methyl-4-(prop-2-enoyl)piperazin-1-yl]pyrido[2,3-d]pyrimidin-2(1H)-one) and / or adagrasib (also known as MRRTX848 or (2S)-4-[7-(8-chloro-1-naphthyl)-5,6,7,8-tetrahydro-2-[[(2"S")-1-methyl-2-pyrrolidinyl]methoxy]pyrido[3,4-"d"]pyrimidin-4-yl]-1-(2-fluoro-1-oxo-2-propen-1-yl)-2-piperazineacetonitrile).

[0069] Pharmaceutical composition The phrase "pharmaceutically acceptable" is used to refer to compounds, materials, compositions, and / or dosage forms that, within the scope of sound medical judgment, are suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0070] The phrase "therapeutically effective" is intended to define the amount of each agent that, while avoiding the deleterious side effects typically associated with alternative therapies, achieves the goal of improving the severity and frequency of occurrence of a disease. For example, an effective anti-cancer agent extends the survival rate of a patient, improves the quality of life, inhibits the growth of rapidly proliferating cells associated with tumor formation, or results in tumor regression.

[0071] As used herein, the terms "treat," "treating," and "treatment" refer to any type of intervention or process that is performed on a subject or that administers an active agent to a subject for the purpose of reversing, alleviating, improving, suppressing, delaying, or preventing the progression, onset, severity, or recurrence of a symptom, complication, condition, or biochemical indicator associated with a disease. In contrast, "prevent" or "prevention" refers to administration to a subject without the disease for the purpose of preventing the occurrence of the disease.

[0072] As used herein, the term "cell" is meant to refer to cells in vitro, ex vivo, or in vivo. In the context of the present invention, an ex vivo cell can be part of a tissue sample excised from an organism such as a mammal. In the context of the present invention, an in vitro cell can be a cell in a cell culture. In the context of the present invention, an in vivo B cell is a cell living in an organism such as a mammal.

[0073] The term "patient" includes humans and animals that receive either treatment or prophylactic treatment.

[0074] The term "subject" includes any human or animal. For example, the methods and compositions disclosed herein can be used to treat subjects suffering from cancer.

[0075] Animals (non-human) include all vertebrates, such as mammals and non-mammals including cows, sheep, pigs, goats, horses, poultry, dogs, cats, non-human primates, rodents, etc. In one embodiment, the subject is a human subject.

[0076] As used herein, the phrase "pharmaceutically acceptable carrier" means a pharmaceutically acceptable material, composition, or medium involved in transporting or delivering a subject compound from one organ or part of the body to another, such as a liquid or solid diluent, solvent, excipient, manufacturing aid (e.g., lubricant), or encapsulating material. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation.

[0077] Suitable other components are the aforementioned carriers and further additives including adjuvants, preservatives, fillers, fluidity regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, bittering agents, fragrances, antibacterial agents, antifungal agents, lubricants, dispensing agents and the like. Suitable additives are selected according to the dosage form and the nature of the pharmaceutical form and so as not to cause harm to the patient.

[0078] The term "pharmaceutical composition" refers to a composition comprising the compound of the present invention a) at least one further pharmaceutically active substance, and b) at least one additional pharmaceutically acceptable carrier and / or additive in combination with at least one further compound selected therefrom.

[0079] The term "RAS inhibitor" refers to an agent that can reduce the level of RAS protein, reduce the level of RAS activity, and / or inhibit the level of RAS expression intracellularly. The RAS inhibitor may be a reversible inhibitor or an irreversible inhibitor. As used herein, the "RAS" protein refers to a protein that is a member of a related protein family expressed in all cell lineages and organs of humans and animals. All members of the RAS protein family belong to a class of proteins called small GTPases (also called small G proteins, a family of hydrolases that can bind to and hydrolyze GTP), and are involved in signal transduction (cell signaling) within the cell. RAS is a typical member of the RAS superfamily of proteins, all of which are related in three-dimensional structure and regulate diverse cell behaviors. When RAS is "switched on" by an incoming signal, subsequently other proteins are switched on, and ultimately genes involved in cell growth, differentiation, and survival are switched on. Mutations in the RAS gene produce permanently activated RAS proteins, which can cause unintended excessive signal transduction within the cell even in the absence of an incoming signal. Since these signals lead to cell growth and division, excessive activation of RAS signaling can ultimately lead to cancer. The three RAS genes in humans (HRAS, KRAS, and NRAS) are the most common oncogenes in human cancers. As described above, the clinically most notable members of the RAS subfamily are HRAS, KRAS, and NRAS. However, there are other members of this subfamily, such as those selected from DIRAS1, DIRAS2, DIRAS3, ERAS, GEM, MRAS, NKIRAS1, NKIRAS2, NRAS, RALA, RALB, RAP1A, RAP1B, RAP2A, RAP2B, RAP2C, RASD1, RASD2, RASL10A, RASL10B, RASL11A, RASL11B, RASL12, REM1, REM2, RERG, RERGL, RRAD, RRAS, RRAS2.In other words, the most common changes in NRAS are NRAS mutation (2.87%), NRAS exon 3 mutation (1.90%), NRAS exon 3 missense (1.88%), NRAS codon 61 missense (1.72%), and NRAS exon 2 mutation (0.95%). The most common changes in HRAS are HRAS mutation (0.77%), HRAS missense (0.75%), HRAS exon 2 mutation (0.30%), HRAS codon 61 missense (0.26%), and HRAS Q61R (0.14%). The most common changes in HRAS are HRAS mutation (0.77%), HRAS missense (0.75%), HRAS exon 2 mutation (0.30%), HRAS codon 61 missense (0.26%), and HRAS Q61R (0.14%) (Source: Mycancer genome portal).

[0080] The compounds of formulas (I) and (A) as defined above, the composition (1) as defined above, and the pharmaceutical composition comprising at least one of the compounds of formulas (I) and (A) as defined above, or the composition (1) as defined above, may be administered to humans and animals, preferably humans.

[0081] In principle, any administration method may be used to deliver the compound or pharmaceutical composition according to the present invention to a subject. Suitable administration methods are oral, enteral, parenteral, intravenous, topical, intramuscular, and subcutaneous route administrations.

[0082] The compounds of formulas (I) and (A) as defined above can selectively reduce the intracellular RAS protein level, reduce the RAS activity level, and in particular reduce the activity levels of HRAS and NRAS in cells, specifically KRAS4A and KRAS4B. For example, the compounds of formulas (I) and (A) as defined above can be used to selectively reduce the RAS activity level in cells or in an individual in need of a reduction in the RAS protein level, and also to reduce the RAS activity level by administering an inhibitory amount of the compounds of formulas (I) and (A) as defined above or a salt thereof.

[0083] In one embodiment, the present invention provides a formulation comprising a compound of formula (I) and (A) as defined above and / or a pharmaceutically acceptable salt thereof, and an additional therapeutic agent for use simultaneously, separately or sequentially in the treatment and / or prevention of (a) disease(s), preferably a proliferative disease (e.g., cancer), particularly a disease associated with the activity of the RAS protein.

[0084] The additional therapeutic agent is selected from chemotherapeutic agents, radiotherapy agents, immuno-oncology agents, and combinations thereof.

[0085] In one aspect, the compounds of formula (I) and (A) as defined above are sequentially administered prior to the administration of the immuno-oncology agent. In another aspect, the compounds of formula (I) and (A) as defined above are administered simultaneously with the immuno-oncology agent. In yet another aspect, the compounds of formula (I) and (A) as defined above are sequentially administered after the administration of the immuno-oncology agent.

[0086] In another aspect, the compounds of formula (I) and (A) as defined above may be formulated together with the immuno-oncology agent.

[0087] Immuno-oncology agents include, for example, small molecule drugs, antibodies or other biological molecules or small molecules. Examples of biological immuno-oncology agents include, but are not limited to, cancer vaccines, antibodies and cytokines. In one aspect, the antibody is a monoclonal antibody. In another aspect, the monoclonal antibody is humanized or human.

[0088] In one aspect, the immuno-oncology agent is (i) an agonist of a stimulatory (including co-stimulatory) receptor or (ii) an antagonist of an inhibitory (including co-inhibitory) signal on T cells, both of which result in an amplification of the antigen-specific T cell response (often referred to as immune checkpoint regulators).

[0089] Suitable stimulatory and inhibitory molecules are members of the immunoglobulin superfamily (IgSF). One important family of membrane-bound ligands that bind to co-stimulatory or co-inhibitory receptors is the B7 family, which includes B7-1, B7-2, B7-H1 (PD-L1), B7-DC (PD-L2), B7-H2 (ICOS-L), B7-H3, B7-H4, B7-H5 (VISTA), and B7-H6. Another family of membrane-bound ligands that bind to co-stimulatory or co-inhibitory receptors is the TNF family of molecules that bind to cognate TNF receptor family members, which includes CD40 and CD40L, OX-40, OX-40L, CD70, CD27L, CD30, CD30L, 4-1BBL, CD137 (4-1BB), TRAIL / Apo2-L, TRAILR1 / DR4, TRAILR2 / DR5, TRAILR3, TRAILR4, OPG, RANK, RANKL, TWEAKR / FnI4, TWEAK, BAFFR, EDAR, EXDAR, TACI, APRIL, BCMA, LTpR, LIGHT, DcR3, HVEM, VEGETL1A, TRAMP / DR3, EDAR, EDA1, XEDAR, EDA2, TNFR1, lymphotoxin a / TNFp, TNFR2, TNFa, LTpR, lymphotoxin a 1b2, FAS, FASL, RELT, DR6, TROY, NGFR.

[0090] In one aspect, the T cell response can be stimulated by a combination of a compound of formula (I) and (A) as defined above with one or more of the following. (i) antagonists of proteins that inhibit T cell activation (e.g., immune checkpoint inhibitors), such as CTLA-4, PD-1, PD-L1, PD-L2, LAG-3, TIM-3, galectin 9, CEACAM-1, BTLA, CD69, galectin-1, TIGIT, CD113, GPR56, VISTA, 2B4, CD48, GARP, PD1H, LAIR1, TIM-1, and TIM-4, and (ii) Agents that stimulate the activation of T cells, such as B7-1, B7-2, CD28, 4-1BB (CD137), 4-1BBL, ICOS, ICOS-L, OX40, OX40L, GITR, GITRL, CD70, CD27, CD40, DR3, and CD28H.

[0091] Other agents that can be used in combination with the compounds of formulas (I) and (A) defined above for cancer treatment include antagonists of inhibitory receptors on NK cells or stimulants of activating receptors on NK cells. For example, the compounds of formulas (I) and (A) defined above can be used in combination with an antagonist of KIR, such as lirilumab.

[0092] Still other agents for combination therapy include agents that inhibit or deplete macrophages or monocytes, including, but not limited to, CSF-1R antagonists such as the CSF-1R antagonist antibody RG7155.

[0093] The combination therapy is intended to include administering these therapeutic agents sequentially, i.e., administering each therapeutic agent at different times, and administering these therapeutic agents or at least two of the therapeutic agents substantially simultaneously.

[0094] Substantial simultaneous administration can be achieved, for example, by administering to a subject a single dosage form having a defined ratio of each therapeutic agent, or multiple single dosage forms of each therapeutic agent. Sequential or substantial simultaneous administration of each therapeutic agent can be effected by any suitable route of administration including, but not limited to, oral, intravenous, intramuscular, or direct absorption through mucosal tissue. The therapeutic agents can be administered by the same route or by different routes. For example, the first therapeutic agent of a selected combination can be administered by intravenous injection while another therapeutic agent of the combination can be administered orally. Alternatively, for example, all therapeutic agents can be administered orally or all therapeutic agents can be administered by intravenous injection. The combination therapy also includes further combining the administration of the therapeutic agents with other biologically active components and non-pharmacological treatments (e.g., surgery or radiation therapy). Where the combination therapy further includes non-pharmacological treatment, the non-pharmacological treatment can be effected at an appropriate time as long as a beneficial effect is obtained from the co-action of the combination of the therapeutic agent and the non-pharmacological treatment. For example, where appropriate, the non-pharmacological treatment can be temporarily, perhaps for several days or weeks, separated from the administration of the therapeutic agent and a beneficial effect can still be achieved.

[0095] The types of cancer that can be treated with the compounds of formula (I) and (A) as defined above include, but are not limited to, cancer of the prostate, colon, rectum, pancreas, cervix, stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck, skin (including melanoma and basal cell carcinoma), mesothelium, leukocytes (including lymphoma and leukemia), esophagus, breast, muscle, connective tissue, lung (including small cell lung cancer and non-small cell cancer), adrenal gland, thyroid, kidney, or bone; or glioblastoma, mesothelioma, renal cell carcinoma, gastric cancer, sarcoma (including Kaposi's sarcoma), choriocarcinoma, skin basal cell carcinoma, hematological malignancies (including blood, bone marrow and lymph nodes), or testicular seminoma.

[0096] In one embodiment, the present invention relates to the inhibition of HRAS mutations detected in bladder urothelial carcinoma, breast invasive ductal carcinoma, lung adenocarcinoma, prostate cancer, and colon adenocarcinoma. This accounts for approximately 0.94% of all human cancers and approximately 1.02% of solid tumors. In another embodiment, the present invention also relates to the inhibition of HRAS mutations detected in other cancers selected from chronic myelomonocytic leukemia, non-Hodgkin lymphoma, thyroid cancer, head and neck squamous cell carcinoma, lung squamous cell carcinoma, ovarian cancer, poorly differentiated thyroid cancer, squamous cell carcinoma, small cell lung cancer, glioma, low-grade glioma, pancreatic cancer, acute lymphoblastic leukemia, histiocytic and dendritic cell tumors, multiple myeloma, neurofibromatosis type, pancreatic ductal adenocarcinoma, thyroid follicular carcinoma, fetal rhabdomyosarcoma, malignant thyroid tumor, undifferentiated thyroid (anaplastic) carcinoma, thymic carcinoma, urothelial carcinoma, papillary thyroid carcinoma, cutaneous melanoma, mucosal melanoma, endometrial carcinoma, malignant peripheral nerve sheath tumor, neuroblastoma, prostate cancer, soft tissue sarcoma, breast cancer, colorectal adenocarcinoma, gastric cancer, diffuse large B-cell lymphoma, diffuse glioma, myelodysplastic syndrome, renal cell carcinoma, astrocytic tumor, hepatocellular carcinoma, and schwannoma.

[0097] NRAS mutations are detected in approximately 3.03% of all human cancers (cutaneous melanoma, melanoma, colorectal adenocarcinoma, acute myeloid leukemia, thyroid cancer, and lung adenocarcinoma) in which the mutations frequently occur. Accordingly, another embodiment of the invention relates to the inhibition of NRAS mutations detected in cutaneous melanoma, melanoma, colorectal adenocarcinoma, acute myeloid leukemia, thyroid cancer, and lung adenocarcinoma. This accounts for approximately 2.83% of patients with solid malignancies. In another embodiment, the invention also relates to the inhibition of NRAS mutations detected in colorectal cancer, non-small cell lung cancer, acute myeloid leukemia, myelodysplastic syndrome, chronic myelomonocytic leukemia, colorectal adenocarcinoma, multiple myeloma, non-Hodgkin lymphoma, pancreatic cancer, cutaneous melanoma, ovarian cancer, pancreatic ductal adenocarcinoma, acute lymphoblastic leukemia, thyroid cancer, glioma, neurofibromatosis type 1, poorly differentiated thyroid cancer, secondary acute myeloid leukemia, therapy-related acute myeloid leukemia, blast excess myelodysplastic syndrome 2, juvenile myelomonocytic leukemia, histiocytic and dendritic cell tumors, head and neck squamous cell carcinoma, small cell lung cancer, low-grade glioma, squamous cell lung cancer, breast cancer, chronic myelomonocytic leukemia-2, chronic myelomonocytic leukemia, anaplastic thyroid cancer (undifferentiated carcinoma), fetal rhabdomyosarcoma, follicular thyroid cancer, T cell acute lymphoblastic leukemia, mucosal melanoma, chronic myelomonocytic leukemia-1, low-grade ovarian serous adenocarcinoma, papillary thyroid cancer, refractory anemia with excess blasts, myeloid neoplasm myelodysplasia / myeloproliferative neoplasm, unclassifiable rectal cancer, colorectal cancer, malignant peripheral nerve sheath tumor, cholangiocarcinoma, endometrial cancer mantle cell lymphoma, secondary myelodysplastic syndrome, therapy-related myelodysplastic syndrome, lymphoma neuronal and neuronal / glial cell mixed tumor, ganglioglioma, soft tissue sarcoma, bladder cancer, esophageal cancer, sarcoma, thymic carcinoma, lung adenocarcinoma, lung cancer, uveal melanoma head and neck cancer, diffuse glioblastoma, squamous cell carcinoma, chronic myelogenous leukemia, adenocarcinoma gastroesophageal junction, glioblastoma multiforme, neuroblastoma, astrocytoma, hepatocellular carcinoma, pancreatic adenocarcinoma, diffuse large B cell lymphoma, anaplastic astrocytoma, gastric adenocarcinoma, gastric cancer, prostate cancer, renal cell carcinoma, acute myeloid leukemia arising from previous myelodysplastic syndrome, B cell acute lymphoblastic leukemia, double hit lymphoma, dysplastic neuroepithelial tumor, gangliocytoma, low-grade neuroepithelial tumor, peripheral T cell lymphoma, pilocytic astrocytoma, pilomyxoid astrocytoma, rhabdoid tumor, and schwannoma.

[0098] One or more additional pharmaceuticals or therapies, such as antiviral agents, chemotherapeutic agents or other anti-cancer agents, immunopotentiators, immunosuppressants, radiation, anti-tumor and anti-viral vaccines, cytokine therapies (e.g., IL2 and GM-CSF), and / or tyrosine kinase inhibitors, can optionally be used in combination with the compounds of formula (I) and (A) as defined above for treating RAS protein-related diseases, disorders or conditions. The agents can be administered in combination with the compound in a single dosage form, or the agents can be administered simultaneously or sequentially as separate dosage forms.

[0099] Suitable chemotherapeutic agents or other anti-cancer agents include, for example, alkylating agents (including but not limited to nitrogen mustards, ethyleneimine derivatives, alkyl sulfonates, nitrosoureas, and triazenes) such as uracil mustard, chloromethine, cyclophosphamide (CYTOXAN®), ifosfamide, melphalan, chlorambucil, pipobroman, triethylenemelamine, triethylenethiophosphoramide, busulfan, carmustine, lomustine, streptozocin, dacarbazine, and temozolomide.

[0100] In the treatment of melanoma, suitable agents for use in combination with the compounds of formula (I) and (A) as defined above include dacarbazine (DTIC), optionally in combination with other chemotherapeutic agents such as carmustine (BCNU) or cisplatin; the "Dartmouth regimen" consisting of DTIC, BCNU, cisplatin, and tamoxifen; combinations of cisplatin, vinblastine, DTIC, temozolomide or YERVOY (trademark). The compounds of formula (I) and (A) as defined above may also be used in combination with immunotherapeutic agents including cytokines such as interferon α, interleukin 2, and tumor necrosis factor (TNF) in the treatment of melanoma. The compounds of formula (I) and (A) as defined above may also be used in combination with vaccine therapy in the treatment of melanoma. Anti-melanoma vaccines are, in a sense, similar to anti-viral vaccines used to prevent diseases caused by viruses such as polio, measles, and mumps. To stimulate the body's immune system to destroy melanoma cells, weakened melanoma cells or parts of melanoma cells, called antigens, may be injected into the patient.

[0101] Melanomas localized to the arms or legs may be treated using a combination of agents containing one or more of the compounds of formula (I) and (A) as defined, using the method of isolated limb perfusion with hyperthermia. In this treatment protocol, the blood circulation of the affected limb is temporarily separated from the rest of the body, and a high dose of chemotherapeutic agent is injected into the artery supplying blood to that limb. This allows a high dose to be administered to the tumor area without these agents, which can cause severe side effects, being exposed to the internal organs. Usually, the fluid is warmed to 38.9 - 40 °C. Melphalan is the drug most commonly used in this chemotherapy. This can be administered together with another agent called tumor necrosis factor (TNF).

[0102] Suitable chemotherapeutic agents or other anti-cancer agents include, for example, antimetabolites such as methotrexate, 5-fluorouracil, floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabine phosphate, pentostatin, and gemcitabine (including, but not limited to, folic acid antagonists, pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors).

[0103] Suitable chemotherapeutic agents or other anti-cancer agents further include specific natural products and their derivatives such as vinblastine, vincristine, vindesine, bleomycin, dactinomycin, daunorubicin, doxorubicin, epirubicin, idarubicin, Ara-C, paclitaxel (Taxol), mitramycin, deoxycoformycin, mitomycin-C, L-asparaginase, interferon (especially IFN-α), etoposide, and teniposide (for example, vinca alkaloids, antitumor antibiotics, enzymes, lymphokines, and epipodophyllotoxins).

[0104] Other cytotoxic agents include navelbine, CPT-11, anastrozole, letrozole, capecitabine, raloxifene, and droloxifene.

[0105] Also, suitable cytotoxic agents are, for example, epipodophyllotoxins, antitumor enzymes, topoisomerase inhibitors, procarbazine, quinacrine, platinum complexes such as cisplatin and carboplatin, biological response modifiers, growth inhibitors, antihormonal therapeutic agents, leucovorin, tegafur, and hematopoietic growth factors.

[0106] Other anti-cancer agents include antibody therapeutic agents such as trastuzumab (Herceptin®), antibodies against costimulatory molecules such as CTLA-4, 4-1BB, and PD-1, or antibodies against cytokines (IL-10 or TGF-β).

[0107] Other anti-cancer agents also include those that block the migration of immune cells, such as antagonists against chemokine receptors such as CCR2 and CCR4.

[0108] Other anti-cancer agents also include adjuvants or those that enhance the immune system such as adoptive T cell transfer.

[0109] Anti-cancer vaccines include dendritic cells, synthetic peptides, DNA vaccines, and recombinant viruses.

[0110] In certain embodiments of the present invention, at least one compound of formula (I) and (A) as defined above and at least one chemotherapeutic agent are administered to a patient simultaneously or sequentially. In other words, at least one compound of formula (I) and (A) as defined above may be administered first, at least one chemotherapeutic agent may be administered first, or at least one compound of formula (I) and (A) as defined above may be administered simultaneously. Also, when two or more compounds of formula (I) and (A) as defined above and / or chemotherapeutic agents are used, these compounds may be administered in any order.

[0111] The present invention also provides a pharmaceutical composition comprising a therapeutically effective amount of one or more compounds of formula (I) and (A) as defined above, one or more pharmaceutically acceptable carriers (additives) and / or diluents, and optionally one or more additional therapeutic agents as described above formulated together.

[0112] The compounds of formula (I) and (A) as defined above may be administered in any suitable route, preferably in the form of a pharmaceutical composition adapted to such a route, in a therapeutically effective dose for the intended treatment. The compounds of formula (I) and (A) as defined above and compositions of the compounds can be administered by any suitable means, for example, oral administration, such as tablets, capsules (including sustained release or extended release formulations), pills, powders, granules, elixirs, tinctures, suspensions (including nano-suspensions, micro-suspensions, spray-dried dispersions), syrups, emulsions; sublingual administration; buccal mucosal administration; parenteral administration, such as subcutaneous, intravenous, intramuscular, intra-sternal injection, or infusion (e.g., by sterile aqueous or non-aqueous solutions or suspensions); nasal administration, such as administration to the nasal mucosa by inhalation spray; topical administration, such as in the form of creams or ointments; or rectal administration, such as in the form of suppositories. They can be administered alone, but are usually administered with a pharmaceutical carrier selected based on the chosen route of administration and standard pharmaceutical practice.

[0113] In the case of oral administration, the pharmaceutical composition may be, for example, in the form of tablets, capsules, liquid capsules, suspensions, or liquids. The pharmaceutical composition is preferably manufactured in the form of dosage units containing a specific amount of the active ingredient. For example, the pharmaceutical composition may be provided as tablets or capsules containing an amount of the active ingredient in the range of about 0.1 to 1000 mg, preferably about 0.25 to 250 mg, more preferably about 0.5 to 100 mg. The daily dose suitable for humans or animals may vary greatly depending on the condition of the patient and other factors, but can be determined using conventional methods.

[0114] Any pharmaceutical composition contemplated herein can be administered orally, for example, via any acceptable and suitable oral formulation. Oral formulations include, for example, tablets, troches, lozenges, aqueous and oily suspensions, dispersible powders or granules, emulsions, hard and soft capsules, liquid capsules, syrups, elixirs, but are not limited thereto. The pharmaceutical composition for oral administration can be prepared according to any method known in the art for manufacturing pharmaceutical compositions for oral administration. To provide a pharmaceutically palatable formulation, the pharmaceutical composition according to the present invention can contain at least one agent selected from sweetening agents, flavoring agents, bittering agents, coloring agents, lubricating agents, antioxidants, and preservatives.

[0115] Tablets can be prepared, for example, by mixing at least one compound of formula (I) and (A) as defined above, and / or at least one pharmaceutically acceptable salt, with at least one non-toxic pharmaceutically acceptable excipient suitable for the manufacture of tablets. Examples of excipients include, for example, inert diluents such as calcium carbonate, sodium carbonate, lactose, calcium phosphate, and sodium phosphate; granulating and disintegrating agents such as microcrystalline cellulose, croscarmellose sodium, corn starch, and alginic acid; binding agents such as starch, gelatin, polyvinylpyrrolidone, and acacia; and lubricants such as magnesium stearate, stearic acid, and talc, but are not limited thereto. Also, tablets can be uncoated or coated by known techniques to hide the unpleasant taste of unpalatable drugs or to delay the decomposition and absorption of the active ingredient in the gastrointestinal tract, thereby prolonging the effect of the active ingredient for a longer period. Examples of water-soluble taste masking materials include, but are not limited to, hydroxypropylmethylcellulose and hydroxypropylcellulose. Examples of time delay materials include, but are not limited to, ethylcellulose and cellulose acetate butyrate.

[0116] Hard gelatin capsules can be prepared, for example, by mixing at least one compound of formula (I) and (A) as defined above, and / or at least one pharmaceutically acceptable salt thereof, with at least one inert solid diluent such as calcium carbonate, calcium phosphate and kaolin, etc.

[0117] Soft gelatin capsules can be prepared, for example, by mixing at least one compound of formula (I) and (A) as defined above, and / or at least one pharmaceutically acceptable salt thereof, with at least one water-soluble carrier (e.g., polyethylene glycol) and at least one oily medium (e.g., peanut oil, liquid paraffin, and olive oil).

[0118] An aqueous suspension can be prepared, for example, by mixing at least one compound of formula (I) and (A) as defined above and / or at least one pharmaceutically acceptable salt thereof with at least one excipient suitable for the manufacture of an aqueous suspension. Examples of excipients suitable for the manufacture of an aqueous suspension include suspending agents such as sodium carboxymethylcellulose, hydroxypropylmethylcellulose and hydroxypropylcellulose, sodium alginate, alginic acid, polyvinylpyrrolidone, tragacanth gum, and acacia gum; dispersing or wetting agents such as naturally occurring phospholipids such as lecithin; condensation products of alkylene oxides and fatty acids such as polyoxyethylene stearate; condensation products of ethylene oxide and long-chain aliphatic alcohols such as heptadecaethyleneoxycetanol; condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol such as polyoxyethylene sorbitol monooleate; and condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides such as polyethylene sorbitan monooleate, but are not limited thereto. The aqueous suspension can also contain at least one preservative (e.g., ethyl and n-propyl p-hydroxybenzoate), at least one coloring agent, at least one flavoring agent, and / or at least one sweetening agent (including but not limited to sucrose, saccharin, and aspartame).

[0119] An oily suspension can be prepared, for example, by suspending at least one compound of formula (I) and (A) as defined above and / or at least one pharmaceutically acceptable salt thereof in a vegetable oil such as peanut oil, olive oil, sesame oil, and coconut oil, or in a mineral oil such as liquid paraffin. The oily suspension can also contain at least one thickening agent such as beeswax, hard paraffin, and cetyl alcohol. To provide a palatable oily suspension, at least one sweetening agent and / or at least one flavoring agent as already described above can be added to the oily suspension. The oily suspension can further contain at least one preservative including, but not limited to, antioxidants such as butylated hydroxyanisole and α-tocopherol.

[0120] Dispersible powders and granules can be prepared, for example, by mixing at least one compound of formula (I) and (A) as defined above and / or at least one pharmaceutically acceptable salt thereof with at least one dispersing agent and / or wetting agent; at least one suspending agent; and / or at least one preservative. Suitable dispersing agents, wetting agents, and suspending agents are as already described above. Examples of preservatives include, but are not limited to, antioxidants such as ascorbic acid. Also, the dispersible powders and granules can contain at least one excipient including, but not limited to, sweetening agents, flavoring agents, and coloring agents.

[0121] An emulsion of at least one compound of formula (I) and (A) as defined above and / or at least one pharmaceutically acceptable salt thereof can be prepared, for example, as an oil-in-water emulsion. The oil phase of the emulsion containing the compounds of formula (I) and (A) as defined above can be composed of known components in a known manner. The oil phase can be provided by, for example, vegetable oils such as olive oil and peanut oil, mineral oils such as liquid paraffin, and mixtures thereof, but is not limited thereto. This phase may be composed of only an emulsifier, or may be composed of a mixture of at least one emulsifier and a fat or an oil, or a mixture of both a fat and an oil. Suitable emulsifiers include, for example, naturally occurring phospholipids such as soy lecithin; esters or partial esters derived from fatty acids and hexitol anhydrides such as sorbitan monooleate; and condensation products of partial esters and ethylene oxide such as polyoxyethylene sorbitan monooleate, but are not limited thereto. Preferably, the hydrophilic emulsifier is included together with a lipophilic emulsifier that acts as a stabilizer. It is also preferred to include both an oil and a fat. The emulsifier (with or without a stabilizer) constitutes a so-called emulsifying wax, and this wax, together with the oil and the fat, constitutes a so-called emulsifying ointment base that forms the oily dispersed phase of the cream preparation. The emulsion can also contain a sweetening agent, a flavoring agent, a preservative, and / or an antioxidant. Emulsifiers and emulsion stabilizers applicable to the formulations of the present invention include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl monostearate, sodium lauryl sulfate, glyceryl distearate (alone or in combination with a wax), or other materials well known in the art.

[0122] The compounds of formula (I) and (A) as defined above and / or at least one pharmaceutically acceptable salt thereof can be administered, for example, intravenously, subcutaneously and / or intramuscularly via any pharmaceutically acceptable, suitable, injectable form. Examples of injectable forms include, for example, sterile aqueous solutions containing acceptable media and solvents such as water, Ringer's solution and isotonic sodium chloride solution; sterile water-in-oil microemulsions and aqueous or oily suspensions, but are not limited thereto.

[0123] Formulations for parenteral administration may be in the form of isotonic sterile injection solutions or suspensions, aqueous or non-aqueous. These solutions and suspensions can be prepared from sterile powders or granules using one or more of the carriers or diluents mentioned for use in formulations for oral administration, or using other suitable dispersing agents, wetting agents, or suspending agents. The present compounds can be dissolved in water, polyethylene glycol, propylene glycol, ethanol, corn oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, tragacanth gum, and / or various buffer solutions. Other adjuvants and methods of administration are well known and widely known in the pharmaceutical art. The active ingredient may be administered by injection as a composition with a suitable carrier including physiological saline, dextrose and water, or as a composition with a cyclodextrin solubilizing agent (i.e., Captisol), a co-solvent solubilizing agent (i.e., propylene glycol), or a micelle solubilizing agent (i.e., Tween80).

[0124] Sterile injectable formulations may also be sterile injectable solutions or suspensions in a non-toxic parenterally acceptable diluent or solvent, for example as a solution in 1,3-butanediol. Among the acceptable media and solvents that can be used are water, Ringer's solution and isotonic sodium chloride solution. Also, sterile solid oils have been conventionally used as solvents or suspending media. For this purpose, any odorless solid oil containing synthetic mono- or diglycerides can be used. Also, fatty acids such as oleic acid are used in the preparation of injectables.

[0125] An oil-in-water type microemulsion in sterile water for injection can be prepared, for example, by: 1) dissolving at least one compound of formula (I) and (A) as defined above in an oil phase such as a mixture of soybean oil and lecithin; 2) combining the compound of formula (I) and (A) containing the oil phase with a mixture of water and glycerol; and 3) treating the mixture to form a microemulsion.

[0126] A sterile aqueous suspension or an oily suspension can be prepared according to methods already known in the art. For example, a sterile aqueous solution or suspension such as 1,3-butanediol can be prepared with a non-toxic parenterally acceptable diluent or solvent, and a sterile oily suspension can be prepared with a sterile non-toxic acceptable solvent or suspending medium such as a sterile solid oil like synthetic monoglycerides or diglycerides, and a fatty acid such as oleic acid.

[0127] Pharmaceutically acceptable carriers are formulated according to many factors within the understanding of those skilled in the art. These include, but are not limited to, the type and nature of the active agent to be formulated, the subject to whom the drug containing the composition is administered, the intended route of administration of the composition, and the targeted therapeutic indication. Pharmaceutically acceptable carriers include both aqueous and non-aqueous liquid media, as well as various solid and semi-solid dosage forms. Such carriers can contain many different components and additives in addition to the active agent. Such additional components are included in the formulation for various reasons well known to those skilled in the art, such as stabilization of the active agent and binders. Information on suitable pharmaceutically acceptable carriers and the factors for their selection can be found in various readily available sources, for example, Allen, L.V. Jr. et al., Remington: The Science and Practice of Pharmacy (2 volumes), 22nd edition (2012), Pharmaceutical Press, etc.

[0128] Pharmaceutically acceptable carriers, adjuvants and vehicles that can be used in the pharmaceutical compositions of the present invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, self-emulsifying drug delivery systems (SEDD), such as d-α-tocopherol polyethylene glycol 1000 succinate, surfactants used in pharmaceutical dosage forms such as Tweens, polyethoxylated castor oils such as CREMOPHOR surfactants (BASF), or other similar polymeric delivery matrices, serum proteins such as human serum albumin, buffering substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose substances, polyethylene glycol, sodium carboxymethyl cellulose, polyacrylate, wax, polyethylene-polyoxypropylene block polymers, polyethylene glycol, and lanolin. Cyclodextrins, such as α-, β-, γ-cyclodextrins, or chemically modified derivatives, such as hydroxyalkyl cyclodextrins (including 2- and 3-hydroxypropyl-cyclodextrins), or other solubilizing derivatives can also be advantageously used to enhance the delivery of the compounds of the formula described herein.)

[0129] The pharmaceutically active compounds of the present invention can be processed according to conventional pharmaceutical methods to produce pharmaceuticals for administration to patients including humans and other mammals. The pharmaceutical compositions are subjected to conventional pharmaceutical operations such as sterilization and / or may contain conventional adjuvants such as preservatives, stabilizers, wetting agents, emulsifying agents, buffering agents, etc. Tablets and pills can be further prepared by applying enteric coatings. Such compositions may also contain adjuvants such as wetting agents, sweeteners, flavoring agents, and fragrances.

[0130] Said For therapeutic purposes, the active compounds of the invention are usually combined with one or more adjuvants suitable for the indicated route of administration. For oral administration, the compounds are mixed with lactose, sucrose, starch powder, cellulose esters of alkanoic acids, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric and sulfuric acids, gelatin, gum arabic, sodium alginate, polyvinylpyrrolidone, and / or polyvinyl alcohol, and may be tableted or encapsulated for ease of administration. Such capsules or tablets may contain controlled-release formulations that can be provided in a dispersion of the active compound in hydroxypropylmethylcellulose.

[0131] The amount of the compound administered and the dosing schedule for treating a disease state using the compounds and / or compositions of the invention depend on various factors including age, body weight, sex, the health status of the subject, the type of disease, the severity of the disease, the route and frequency of administration, and the particular compound used. Accordingly, the dosing schedule can vary widely but can be routinely determined using standard methods. The daily dosage can suitably be about 0.001 to 100 mg / kg body weight, preferably about 0.0025 to about 50 mg / kg body weight, and most preferably about 0.005 to 10 mg / kg body weight. The daily dosage can be administered 1 to 4 times a day. Other dosing schedules include once a week and once every two days.

[0132] The pharmaceutical compositions of the invention comprise at least one compound of formula (I) and (A) as defined above, and / or at least one pharmaceutically acceptable salt thereof, or at least one composition (1) as defined above, and optionally an additional agent selected from any pharmaceutically acceptable carrier, adjuvant, and vehicle. Alternative compositions of the invention comprise a compound of formula (I) and (A) as defined above, or a prodrug thereof, and a pharmaceutically acceptable carrier, adjuvant, or vehicle.

[0133] The present invention also includes a pharmaceutical kit useful, for example, for the treatment or prevention of RAS protein-related diseases. Accordingly, the present invention also relates to a formulation comprising: a) a compound of formula (I) and (A) as defined above, or a pharmaceutically acceptable salt thereof, or at least one composition (1) as defined above, and a pharmaceutical composition comprising a pharmaceutically acceptable carrier; and b) a kit comprising instructions for administering a pharmaceutical composition for the treatment of a disease in which inhibition of RAS activation is effective for the treatment of the disease.

[0134] Such a kit can further include, if necessary, one or more various components of conventional pharmaceutical kits that are readily understandable to those skilled in the art, such as containers containing one or more pharmaceutically acceptable carriers, additional containers, and the like. Instructions (either as an insert or a label) indicating the amount of the component to be administered, administration guidelines, and / or guidelines for mixing the components can also be included in the kit.

[0135] The dosing regimen of the compounds of the present invention will of course vary depending on known factors such as, for example, the pharmacodynamic properties of a particular drug, the method and route of administration, the species, age, sex, health status, medical condition and body weight of the patient, the nature and degree of the symptoms, the type of combination therapy, the frequency of treatment, the route of administration, the renal and hepatic functions of the patient, and the expected effect.

[0136] As a general guideline, the daily oral dosage of each active ingredient, when used for the indicated effect, is about 0.001 to about 5000 mg / day, preferably about 0.01 to about 1000 mg / day, and most preferably about 0.1 to about 250 mg / day. In the case of intravenous administration, the most preferred dosage ranges from about 0.01 to about 10 mg / kg / min during a constant rate infusion. The compounds of formula (I) and (A) may be administered once a day or the total daily dosage may be divided and administered 2, 3, or 4 times a day.

[0137] The compound is typically appropriately selected according to the intended dosage form, for example, tablets, capsules, elixirs, and syrups for oral administration, etc., and is administered after being mixed with a suitable pharmaceutical diluent, excipient, or carrier (collectively referred to as a pharmaceutical carrier herein) in accordance with conventional pharmaceutical practices.

[0138] Dosage forms (pharmaceutical compositions) suitable for administration may contain from about 1 mg to about 200 mg of the active ingredient per dosage unit. In these pharmaceutical compositions, the active ingredient is usually present in an amount of about 0.1 to 95% by weight based on the total weight of the composition.

[0139] A typical capsule for oral administration contains at least one compound of formula (I) and (A) (250 mg), lactose (75 mg), and magnesium stearate (15 mg). The mixture is passed through a 60-mesh sieve and filled into No. 1 gelatin capsules.

[0140] A typical injectable preparation is manufactured by aseptically placing at least one compound of formula (I) and (A) (250 mg) into a vial, aseptically freeze-drying, and sealing it. At the time of use, the contents of the vial are mixed with 2 mL of physiological saline to prepare an injection.

[0141] The present invention includes, within its scope, pharmaceutical compositions containing, as an active ingredient, a therapeutically effective amount of at least one compound of formula (I) and (A) as defined above, alone or in combination with a pharmaceutical carrier. Optionally, the compounds of formula (I) and (A) as defined above can be used alone, in combination with other compounds of formula (I) and (A) as defined above, or in combination with one or more other therapeutic agents, such as anticancer agents or other pharmaceutically active substances.

[0142] Regardless of the selected route of administration, the compounds of formula (I) and (A) as defined above, and / or the pharmaceutical compositions of the present invention, which can be used in a suitable hydrated form, are formulated into pharmaceutically acceptable dosage forms by conventional methods known to those skilled in the art.

[0143] The actual dosage level of the active ingredient in the pharmaceutical composition or composition (1) of the present invention can be varied so as to obtain an amount of the active ingredient effective to achieve the desired therapeutic response for a particular patient, composition, and method of administration without causing toxicity to the patient.

[0144] The selected dosage level depends on a variety of factors including the activity of the specific compounds of formulas (I) and (A) as defined above, or their esters, salts, or amides, used, the route of administration, the time of administration, the rate of excretion or metabolism of the specific compounds used, the rate and extent of absorption, the duration of treatment, other drugs, compounds and / or substances used in combination with the specific compounds used, the age, sex, weight, condition, general health, and medical history of the patient being treated, and similar factors well known in the medical arts.

[0145] A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the required pharmaceutical composition. For example, the physician or veterinarian can start with a dosage of the compounds of formulas (I) and (A) as defined above used in the pharmaceutical composition that is lower than the required amount and gradually increase the dosage until the desired effect is achieved to achieve the desired therapeutic effect.

[0146] Generally, the appropriate daily dosage of the compounds of formulas (I) and (A) as defined above is the amount of the compound that is the lowest dosage effective to produce a therapeutic effect. Such effective dosages generally depend on the above factors. Generally, the oral, intravenous, intracerebroventricular, and subcutaneous dosages of the compounds of formulas (I) and (A) as defined above for a patient are in the range of about 0.01 to about 50 mg per kilogram of body weight per day.

[0147] Optionally, the effective daily dosage of the active compound may be administered in unit dosage form as one, two, three, four, five, six, or more sub-dosages administered separately at appropriate intervals throughout the day. In some embodiments of the present invention, the administration is once a day.

[0148] Although the compounds of formula (I) and (A) defined above can be administered alone, they are preferably administered as a pharmaceutical preparation (composition).

[0149] When the other therapeutic agents described above are used in combination with the compounds of formula (I) and (A) defined above, they may be used, for example, in the amounts indicated in the Physician's Desk Reference (PDR) or in amounts determined by those skilled in the art. In the methods of the present invention, such other therapeutic agents may be administered before, simultaneously with, or after the administration of the compounds of the present invention.

[0150] The present invention will be further described with reference to the following examples, but its scope is not limited to the specific embodiments described. The present invention includes any combination of the described features, particularly preferred features, which are not mutually exclusive.

[0151] The present invention will be further described with reference to the following examples, but its scope is not limited to the specific embodiments described. The present invention includes any combination of the described features, particularly preferred features, which are not mutually exclusive.

Brief Description of the Drawings

[0152] Figures 1a) and 1b): NanoBit assay for RAS activation (KRAS G12V, NRAS G12V, HRAS G12V) Constructs of N-terminal LgBit and C-terminal SmBit were purchased from Promega. KRAS G12V,NRAS G12V and HRAS G12V (full length) were cloned into LgBit with Xhol and Bgl II, and CRAF Ras-binding domain (1-149) was cloned into SmBit with EcoRI and Bgl II. For transfection, 1 μg or 2 μg (12-well / 6-well) of plasmid was transfected into cells with 0.5 mM PEI reagent in PBS solution (100 μl or 200 μl). One day after transfection, cells were harvested and seeded on half of the area of a 96-well white plate (Greiner). After another day, the medium was changed to serum-free DMEM and cells were cultured with inhibitors for 2 hours. The NanoGIo assay was performed according to the manufacturer's instructions. Luminescence was measured using a Tecan infinite (Tecan). Cells treated with DMSO were set as 1. The results shown are the mean ± SEM of 3 independent experiments.

[0153] Quinacrine dihydrochloride hydrate inhibits K-RAS, N-RAS, and H-RAS.

[0154] Figure 2a) and 2b): MTT assay for cell proliferation Cell viability was quantified using the Cell Proliferation Kit I (Roche, Basel, Switzerland). For NCI-H358 cells, 5000 cells per well were used and seeded into 96-well cell culture plates. After treatment, 10 μl of MTT solution was added and incubated in a CO2 incubator for 2 hours. Then, 100 μl of solubilization buffer was added to each well and incubated overnight in a CO2 incubator. Absorbance at 570 nm was measured to quantify cell viability, which was evaluated by the amount of metabolized MTT.

[0155] MTT assay for growing different tumor cell lines with RAS mutations in soft agar containing quinacrine dihydrochloride NCI-H358, H358, H2122, ASPC-1, HCT-116, T24, and HT1080 cells were used in the soft agar colony formation assay. After treatment with quinacrine dihydrochloride hydrate, colonies were stained with MTT and quantified by measuring the absorbance at 570 nm after lysis. The results represent the mean ± SEM of 3 independent experiments. JPEG2025524343000010.jpg17135

[0156] Quinacrine dihydrochloride inhibits the growth of different RAS-isoform-dependent cell lines and arrests cancer growth.

[0157] Figure 3a) and 3b): MTT assay for cell viability in 96-well cell culture plates Cells were treated with quinacrine dihydrochloride hydrate for 48 hours and cell viability was evaluated by MTT assay. Cells treated with DMSO were set as 1. The results shown are the mean ± SEM of 3 independent experiments.

[0158] Figure 3a): Quinacrine dihydrochloride inhibits KRAS-dependent cells regardless of the presence or absence of mutations. The G12C / Y96D mutation is usually seen in patients who develop resistance to G12C inhibitors. Therefore, patients resistant to G12C inhibitors can also be treated with quinacrine dihydrochloride.

[0159] Figure 3b): Comparison of single compounds of quinacrine dihydrochloride and MRTX849 (Adagrasib) and a mixture of quinacrine dihydrochloride (Q) and MRTX849 (M) in the MTT assay in RAS mutant cells. Additionally, the calculated (expected) and measured (true) values of the mixture are shown. The following equations were used to obtain the expected values. Expected value = Q + M - (P·M / 100) Expected value = [100 - [Q + M - (Q·M / 100)]] / 100 (normalized)

[0160] Figure 4: RAS syndrome model of quinacrine dihydrochloride; MTT assay for cell viability in 96-well cell culture plates Cells were treated with quinacrine dihydrochloride hydrate for 48 hours, and cell viability was evaluated by MTT assay. Cells treated with DMSO were set as 1. The results shown are the mean ± SEM of 3 independent experiments. JPEG2025524343000011.jpg2098

[0161] Quinacrine dihydrochloride inhibits the RAS protein in the RAS syndrome.

[0162] Cell culture HeLa S3 (DSMZ) was authenticated by Eurofin genomics and cultured in DMEM (10% heat-inactivated FBS). NCI-H358 cells were cultured in RPMI-1640 (10% heat-inactivated FBS).

Example

[0163] Quinacrine dihydrochloride hydrate (4-N-(6-chloro-2-methoxyacridin-9-yl)-1-N,1-N-diethylpentane-1,4-diamine, hydrate; dihydrochloride) JPEG2025524343000012.jpg3685 is commercially available from Sigma-Aldrich.

Claims

1. A compound of formula (I): for use as an inhibitor of RAS protein activation for the treatment and / or prevention of diseases involving RAS signaling: Wherein: R １ is H, C １ ~C ２ alkyl; R ２ is H, C １ ~C ２ alkyl; n is 1, 2, 3, or 4, or a pharmaceutically acceptable salt thereof.

2. A compound of formula (I): for use as an inhibitor of RAS protein activation: Wherein: R １ is H, C １ ~C ２ alkyl; R ２ is H, C １ ~C ２ alkyl; n is 1, 2, 3, or 4, or a pharmaceutically acceptable salt thereof.

3. The compound of formula (I) according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases, inflammatory diseases and / or genetic diseases involving RAS signaling.

4. The compound of formula (I) according to claims 1 to 3, or a pharmaceutically acceptable salt thereof, for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases and / or inflammatory diseases involving KRAS G12V, KRAS G12A,NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K.

5. The compound of formula (I) according to claims 1 to 4, or a pharmaceutically acceptable salt thereof, for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases and / or inflammatory diseases that are resistant to treatment with RAS mutation-specific inhibitors different from the compound of formula (I).

6. The compound of formula (I) according to claim 5, or a pharmaceutically acceptable salt thereof, wherein said resistance is due to secondary mutations, specifically secondary mutations involving KRAS G12C / Y96D, KRAS G12C / Y96C, and / or KRAS G12C / Y96S.

7. For use as an inhibitor of RAS protein activation, preferably for use as an inhibitor of RAS protein activation for the treatment and / or prevention of diseases involving RAS signaling, in particular for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases, inflammatory diseases, and / or genetic diseases involving RAS signaling, specifically KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K, the formula (I) according to claims 1 to 6: Wherein, R １ is C １ to C ２ alkyl; R ２ is C １ to C ２ alkyl; n is 3, a compound or a pharmaceutically acceptable salt thereof.

8. Compound (A) for use as an inhibitor of RAS protein activation: The compound of formula (I) according to any one of the preceding claims or a pharmaceutically acceptable salt thereof, which is

9. Compound (A) for use as an inhibitor of RAS protein activation for the treatment and / or prevention of diseases involving RAS signaling, the compound of formula (I) according to any one of claims 1 to 8 or a pharmaceutically acceptable salt thereof.

10. Compound (A) for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases, inflammatory diseases, and / or genetic diseases involving RAS signaling, specifically KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K, the compound of formula (I) according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof.

11. A compound (A) for use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases and / or inflammatory diseases that are resistant to treatment with a RAS mutation-specific inhibitor different from the compound of formula (I), preferably, said resistance is due to a secondary mutation, specifically a secondary mutation involving KRAS G12C / Y96D, KRAS G12C / Y96C, and / or KRAS G12C / Y96S, the compound of formula (I) according to any one of claims 1 to 8 or a pharmaceutically acceptable salt thereof.

12. A composition (1) comprising at least one compound of formula (I) according to any one of claims 1 to 11, in particular a compound (A) selected therefrom, or a pharmaceutically acceptable salt, and a RAS mutation-specific inhibitor, in particular a KRAS mutation-specific inhibitor, specifically sotorasib and / or adagrasib.

13. The compound of formula (I) according to any one of claims 1 to 11, in particular compound (A), or a pharmaceutically acceptable salt thereof, or the composition (1) according to claim 12, for the treatment of proliferative diseases, inflammatory diseases and / or genetic diseases involving RAS signaling.

14. The compound of formula (I) according to any one of claims 1 to 11, in particular compound (A), or a pharmaceutically acceptable salt thereof, or the composition (1) according to claim 12, for the treatment of proliferative diseases and / or inflammatory diseases involving KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K.

15. For the treatment of proliferative diseases that are cancers, specifically, cancers of the prostate, colon, rectum, pancreas, cervix, stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck, skin (including melanoma and basal cell carcinoma), mesothelium, white blood cells (including lymphoma and leukemia), esophagus, breast, muscle, connective tissue, lung (including small cell lung cancer and non-small cell cancer), adrenal gland, thyroid gland, kidney, or bone; or for the treatment of cancers selected from glioblastoma, mesothelioma, renal cell carcinoma, gastric cancer, sarcoma (including Kaposi's sarcoma), choriocarcinoma, skin basal cell carcinoma, hematological malignancies (including blood, bone marrow, and lymph nodes), or testicular seminoma, the compound of formula (I) according to any one of claims 1 to 11, particularly compound (A), or a pharmaceutically acceptable salt thereof, or the composition (1) according to claim 12.

16. A pharmaceutical composition for the prevention and / or treatment of proliferative diseases and / or genetic diseases involving RAS signaling, comprising at least one compound of formula (I) according to any one of claims 1 to 11, particularly selected from compound (A), or a pharmaceutically acceptable salt thereof, or the composition (1) according to claim 12.

17. A pharmaceutical composition for the prevention and / or treatment of proliferative diseases and / or inflammatory diseases involving KRAS G12V, KRAS G12A, NRAS G12V, HRAS G12V, KRAS G12C, KRAS G12D, KRAS G12C / Y96D, KRAS G12C / Y96C, KRAS G12C / Y96S, KRAS G13C, KRAS G13D, KRAS G13S, KRAS Q61H, KRAS Q61R, or KRAS Q61K, comprising at least one compound of formula (I) according to any one of claims 1 to 11, particularly selected from compound (A), or a pharmaceutically acceptable salt thereof, or the composition (1) according to claim 12.

18. For use as an inhibitor of RAS protein activation for the treatment and / or prevention of proliferative diseases and / or inflammatory diseases that are resistant to treatment with a RAS mutation-specific inhibitor different from the compound of formula (I), preferably, said resistance is due to secondary mutations, specifically secondary mutations involving KRAS G12C / Y96D, KRAS G12C / Y96C, and / or KRAS G12C / Y96S, the pharmaceutical composition according to claim 16 or 17.

19. At least one compound of formula (I), especially those selected from compound (A), or a pharmaceutically acceptable salt thereof, or the composition (1) according to claim 12, and a pharmaceutically acceptable carrier, the pharmaceutical composition according to claims 16 - 18.

20. The pharmaceutical composition according to claims 16 - 19, further comprising an additional active substance, preferably selected from chemotherapeutic agents, radiotherapy agents, immuno - oncology agents, and combinations thereof.

21. A method of inhibiting the growth, proliferation, or metastasis of cancer cells in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of at least one compound (I) according to any one of claims 1 - 11, especially those selected from the compound of formula (A), or a therapeutically acceptable salt thereof, or the composition (1) according to claim 12.

22. The cancer is selected from prostate, colon, rectum, pancreas, cervix, stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck, skin (including melanoma and basal cell carcinoma), mesothelium, white blood cells (including lymphoma and leukemia), esophagus, breast, muscle, connective tissue, lung (including small - cell lung cancer and non - small - cell cancer), adrenal gland, thyroid, kidney, or bone cancer; or glioblastoma, mesothelioma, renal cell carcinoma, gastric cancer, sarcoma (including Kaposi's sarcoma), choriocarcinoma, skin basal cell carcinoma, hematological malignancies (including blood, bone marrow, and lymph nodes), or testicular seminoma, the method according to claim 21.

23. A method of inhibiting the proliferation of a cell population sensitive to inhibiting RAS activation in vitro or ex vivo, comprising contacting the cell population with at least one compound of formula (I) according to any one of claims 1 - 11, especially those selected from compound (A), or a therapeutically acceptable salt thereof, or the composition (1) according to claim 12.

24. a) At least one compound of formula (I) according to any one of claims 1 to 11, especially those selected from compound (A), or composition (1) according to claim 12, or at least one compound of formula (I) according to any one of claims 1 to 11, especially those selected from compound (A), or a pharmaceutically acceptable salt thereof, a pharmaceutical composition, or a formulation comprising composition (1) according to claim 12 and a pharmaceutically acceptable carrier; and b) A kit comprising instructions for administering a pharmaceutical composition for the treatment of a disease for which inhibition of RAS activation is effective in the treatment of the disease.

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