CN115813930A - Use of macrocyclic compound in combination with MEK inhibitor in medicine for treating diseases - Google Patents

Abstract

The present invention provides the use of a compound of the formula (I) or a pharmaceutically acceptable salt, stereoisomer thereof, in combination with a MEK inhibitor in the preparation of a drug for treating diseases.

Description

Use of macrocyclic compound in combination with MEK inhibitor in medicine for treating diseases

technical field

The present invention relates to the use of a combination of a macrocyclic compound, especially a fluorine-containing macrocyclic compound, and a MEK inhibitor for treating KRAS mutation cancer.

technical background

Kirsten rat sarcoma viral oncogene homolog (KRAS) is one of the most frequently mutated oncogenes in human cancers (COSMIC database), implying poor prognosis of the disease. In the past many years, due to the lack of therapeutic methods targeting this gene, it was regarded as an undruggable target. With the continuous progress of research in recent years, the targeted drug Sotorasib (AMG510) for the KRAS G12C mutation was approved by the FDA on May 29, 2021 for the treatment of KRAS G12C mutation non-small cell lung cancer ( NSCLC).

KRAS protein is a cell membrane-bound guanylate triphosphatase (GTPase), which is activated when it binds to GTP, and the GTPase activity of KRAS converts GTP into inactive GDP, guanylate exchange factor (GEF), For example, SOS1 protein can promote the transformation of RAS protein into the GTP-bound active state. The KRAS protein cycles between the GTP and GDP states, with a half-life of approximately 24 hours for resynthesis. KRAS is like a "cell switch". When it is "turned on" by extracellular stimuli, it will activate a variety of downstream signaling pathways, including cancer-related RAF-MEK-ERK, PI3K-AKT-mTOR, and Ral-GEF pathways, which are involved in cell proliferation , cell cycle regulation, metabolic changes, cell survival and cell differentiation, etc. ).

Point mutations are a common form of KRAS mutations, which lead to KRAS in a persistent GTP-bound activation state, activating downstream oncogenic signaling pathways. The three tumors with the highest frequency of KRAS mutations were pancreatic cancer (88%), colorectal cancer (45%-50%) and lung cancer (31%-35%). KRAS mutations are more common in lung adenocarcinoma (20%-40%), and relatively rare in lung squamous cell carcinoma (5%) (Jemal A, Siegel R, Ward E, et al: Cancer statistics, 2008. CA Cancer JClin58 :71-96, 2008). KRAS mutations, mainly at codons 12 and 13, are seen in up to 30% of NSCLC patients. A large number of studies have found that the most common mutation of KRAS is G12C, accounting for 39%; followed by G12V (18-21%) and G12D (17-18%) (Clin.Cancer Res.18(2012) 6169-6177; J.Clin.Oncol .35(2017) 9021).

History of KRAS-targeted therapy

Nearly 40 years of drug development has been carried out on KRAS targets, including targeting KRAS protein itself, post-translational modification, membrane localization, protein-protein interaction, and downstream signaling pathways, but most of them have failed in clinical research, the reason may be It is that the KRAS protein lacks small molecule drug-binding domains in addition to the GTP/GDP binding pocket.

1) Directly targeting KRAS

Directly targeting KRAS is challenging due to the biochemical complexity of KRAS proteins, the high affinity of GTP for KRAS, and its limited active binding sites. Although recent advances in computer modeling and structural crystallization studies have identified small molecules that directly bind specific RAS conformations, the binding affinities of these early compounds still need to be improved. Although Sotorasib (AMG510) received accelerated approval from the FDA on May 29, 2021, it only targets the G12C mutation.

2) Indirect targeting of KRAS

These include targeting post-translational modifications, membrane localization, protein-protein interactions, and inhibition of downstream signaling pathways. Tumor microenvironment reprogramming is the main feature of tumors. Tumor cells can reprogram tumor microenvironment through various pathways (such as immunosuppression, induction of angiogenesis, and alteration of metabolism) (hanahan et al. Robert A Weinberg, Hallmarks of cancer: the next generation. 2011). Many studies have shown that the KRAS signaling pathway can induce the expression of a large number of immune regulatory factors, such as TGFβ, GM-CSF, CXCL8, IL-6 and IL-10, which interact with the tumor microenvironment, resulting in immunosuppression (Cavalho et al. al. KRASOncogenic Signaling Extends beyond Cancer Cells to Orchestrate the Microenvironment. 2018 ; cullis et al. Jane Cullis, Kras and Tumor Immunity: Friendor Foe 2018 ; Maldegem et al. Mutant KRASattheHeart ofTumor ImmuneEvasion. 2020). Tumor cells driven by KRAS mutations can reprogram stromal cells to a state conducive to tumorigenesis (Cavalho et al. Targeting the Tumor Microenvironment: An Unexplored Strategy for Mutant KRASTumors.2019 ), inhibiting tumorigenesis by blocking autocrine cytokine signaling pathways (Zhu et al. Inhibition of KRAS-DrivenTumorigenicity by Interruption of an Autocrine Cytokine Circuit. 2014 ). One example is the secretion of IL-6 by tumor cells with KRAS mutations. IL-6 is upregulated in lung cancer and mediates signaling pathways that promote KRAS-driven lung tumorigenesis (Brooks et al. IL6 Trans-signaling Promotes KRAS-Driven Lung Carcinogenesis.2016). KRAS mutant tumor cells promote angiogenesis by secreting vascular endothelial growth factor (VEGF) and other angiogenic factors such as CXC chemokines, a paracrine process (Matsuo et al. K-Ras Promotes Angiogenesis Mediated by Immortalized Human Pancreatic Epithelial Cells through Mitogen-Activated Protein Kinase Signaling Pathways.2009 ). In addition to affecting immune evasion and angiogenesis, paracrine processes can also alter tumor cell processes by remodeling the stroma. For example, KRAS mutant cells can secrete insulin-like growth factor-1 (IGF1R), which can increase the mitochondrial capacity of tumor cells through the IGF1R signaling pathway (Tape et al. Oncogenic KRASRegulates Tumor Cell Signaling via Stromal Reciprocation.2016). In summary, for tumor patients with KRAS mutations, an effective treatment plan not only needs to target tumor cells, but also needs to target the tumor microenvironment.

Farnesyltransferase Inhibition: Farnesylation of RAS proteins is required for both their normal physiological function and their oncogenic mutational function. Farnesyltransferase inhibitors (FTIs) Tipifarnib and Salirasib have been studied clinically, but they have not shown efficacy in KRAS-mutated NSCLC.

MEK Inhibition: MEK inhibitors have not been effective as monotherapy in clinical studies. Selumetinib and Trametinib are allosteric selective inhibitors of MEK1/2. Selumetinib was active against KRAS-mutant tumors in preclinical studies. In a phase II clinical study, for 87 patients with previously treated KRAS-mutated advanced NSCLC, there was no significant difference in OS between Selumetinib combined with Docetaxel and Docetaxel alone (9.4 months vs 5.2 months, p=0.21). In another study, for 510 patients with KRAS-mutated NSCLC, selumetinib combined with Docetaxel did not improve PFS compared with Docetaxel alone (HR=0.93). In a phase II clinical trial comparing Trametinib and Docetaxel in the treatment of KRAS-mutated NSCLC patients, the trial was terminated early because the efficacy of Trametinib exceeded the invalidity boundary in the interim analysis. (Blumenschein et al 2015) Therefore, currently known single agents of MEK inhibitors and combination chemotherapy of MEK inhibitors are not effective for KRAS-mutated NSCLC patients.

Intrinsic and adaptive resistance are the limiting factors for the clinical use of KRAS inhibitors as single agents, and the resistance mechanisms include KRAS nucleotide cycling, negative feedback reactivation, and tumor cells bypassing the dependence on KRAS (Hallin et al, cancer discovery, 2020, 10(1), 54-71). Combining drugs that reactivate the MAPK feedback pathway, activate the RTK-induced PI3K pathway, increase apoptosis, and inhibit the pro-inflammatory tumor microenvironment may be a way to significantly improve clinical benefits. KRAS covalent inhibitors combined with immune checkpoint antibodies (such as PD-1 antibodies) showed better efficacy (Canon et al, nature 2019, 575, 217-223). Mechanisms of resistance to MEK inhibitors in KRAS mutant tumors have been found to include compensatory upregulation of PI3K/AKT survival signaling, intrinsic or treatment-induced epithelial-mesenchymal transition (EMT), etc. (Mohanty A J, Sishc B J, Falls K C, et al.GC4419 enhances the response of non-small cell lungcarcinoma cell lines to cisplatin and cisplatin plus radiation through a ROS-mediated pathway[J].2018.). The SRC/FAK signaling pathway regulates the PI3K/AKT survival signaling pathway and is involved in integrin-mediated EMT signal transduction. Activation of the JAK2/STAT3 pathway is a mechanism of resistance to MEK inhibitors.

SRC Inhibition and FAK Inhibition: Src Kinases Are Associated with Drug Resistance Issues in Various Cancer Therapies, Including Radiotherapy, Chemotherapy and Targeted Therapy (Siyuan Zhang.Targeting Src family kinases in anti-cancer therapies:turning promise into triumph.2012) . The Src kinase family promotes mitotic signaling from growth factor receptors through multiple pathways, including signaling pathways required to initiate DNA synthesis, regulation of receptor expression, actin cytoskeleton rearrangement, migration, and survival (Bromann et al. The interplay between Srcfamily kinases and receptor tyrosine kinases. 2004). It has been reported that KRAS induces Src/PEAK1/ErbB2 to form a positive feedback amplification loop, thereby driving pancreatic cancer cell metastasis and drug resistance. Kelber et al. (Kelber et al.KRas Induces a Src/PEAK1/ErbB2Kinase Amplification Loop ThatDrives Metastatic Growth and Therapy Resistance in Pancreatic Cancer.2012) found that the Src inhibitor Dasatinib enhanced the activity of TAZ (Transcriptional coactivator with PDZ-binding motif) Antitumor activity of MEK inhibitors, the combination of Dasatinib and Trametinib is a potential therapeutic strategy for the treatment of KRAS-driven tumors (Rao et al.Dasatinib sensitises KRAS-mutantcancer cells to mitogen-activated protein kinase inhibitor via inhibition of TAZ activity.2012) .

FAK may be involved in inhibiting p53 expression to promote cell survival (Golubovskaya et al. Simultaneous Inhibition of Focal Adhesion Kinase and Src Enhances Detachment and Apoptosis in Colon Cancer Cell Lines. 2007). In preclinical studies, KRAS-mutant cell lines and xenografts with concurrent TP53 or CDKN2A mutations were sensitive to FAK inhibitors. However, in a phase II clinical study, the FAK inhibitor Defactinb monotherapy in patients with KRAS-mutated NSCLC who had undergone multiple lines of therapy had weak clinical efficacy, with a PFS of only 45 days, and the efficacy was not related to TP53 and CDKN2A mutations.

FAK plays an important role in signaling pathways mediated by integrins, RTKs, RAS and TGFβ (Kanteti et al. FAK and paxillin, two potential targets in pancreatic cancer. 2012). Recent studies have found that integrins are involved in the biological regulation of cancer stem cells, and participate in tumor development, metastasis and drug resistance through the SRC/FAK signaling pathway (Seguin et al. Integrins and cancer: regulators of cancer stemness, metastasis, and drug resistance.2015 ). Src has been identified as a key mediator in the process of thyroid proto-neoplasia and is a potential therapeutic target for thyroid cancer. However, inhibition of Src alone leads to activation of the IL-1β>FAK>p130Cas>c-jun>MMP signaling axis, which promotes the development of tumors towards an aggressive phenotype, whereas the combination of FAK and Src inhibitors may block Src inhibitors Induced phenotype switch and drug resistance problem (Kessler et al. Resistance to Src inhibitors the BRAF-mutant tumor secretome to promote an invasive phenotype and therapeutic escape through a FAK>p130Cas>c-Jun signaling axis.2019).

Compensatory upregulation of PI3K/AKT signaling is one of the resistance mechanisms targeting KRAS mutations. FAK directly interacts with the SH2 domain of the PI3K regulatory subunit p85 by phosphorylating Y397, activates the PI3K pathway, and inhibits Doxorubicin-induced apoptosis (van Nimwegen et al.Focal Adhesion Kinase and Protein Kinase BCooperate to Suppress Doxorubicin-Induced Apoptosis of Breast Tumor Cells. 2006). RhoA-FAK is a signaling axis required for the maintenance of KRAS-driven lung adenocarcinoma. Inhibition of FAK in vivo downregulates p-AKT but does not trigger PI3K/AKT-dependent compensatory mechanisms (Konstantinidou et al. RHOA-FAK Is a Required Signaling Axis for the Maintenance of KRAS-Driven LungAdenocarcinomas. 2013).

Phosphorylation of FAK on Y925 constitutes a linking site for GRB2, which can activate the small GTP protein RAS and downstream ERK2 (MAPK) (Kanteti et al. FAK and paxillin, two potential targets inpancreatic cancer. 2016). Paxillin is the main component of focal adhesions, connecting the extracellular matrix and actin cytoskeleton. In tumor cells, Scr and FAK-mediated phosphorylation regulates the function of paxillin. Evidence of increased cell detachment, inhibition of AKT/ERK1/2 and Src, and increased apoptosis showed that dual inhibition of FAK and Src was more effective than inhibition of FAK alone (Golubovskaya et al. Simultaneous Inhibition of Focal Adhesion Kinase and Src Enhances Detachment and Apoptosis in Colon Cancer Cell Lines. 2003). Interferon and inflammation-related genes are enriched in KRAS-mutant colorectal cancer cell lines that exhibit intrinsic and acquired resistance to MEK inhibition (Wagner et al. Suppression of interferon gene expression overcomes resistance to MEK inhibition in KRAS-mutant colorectal cancer.2019 ). In addition, Src and FAK can regulate STAT3, thereby controlling the expression of angiogenic factors such as VEGF and other cytokines (Niu et al oncogene Constitutive Stat3 activity up-regulates VEGF expression and tumor angiogenesis.2002; cavalho et al cancers Targeting the Tumor Microenvironment: An Unexplored Strategy for Mutant KRAS Tumors. 2019 ).

JAK2 inhibition: JAK2 provides signal transduction for inflammatory cytokines, and inhibition of JAK2 may reduce the secretion of interferon and inflammation-related genes and sensitize KRAS mutant cell lines to MEK inhibitors. Studies have reported that KRAS mutations can activate p-STAT3 (Tyr705) in the absence of IL-6 secretion, and STAT3-mediated upregulation of BCL-XL contributes to KRAS mutation-mediated apoptosis resistance in colon cancer cells (Zaanan et al. The Mutant KRAS Gene Up-regulates BCL-XL Protein via STAT3 to Confer Apoptosis Resistance That Is Reversed by BIM Protein Induction and BCL-XL Antagonism. 2015 ). Therefore, inhibition of JAK2 can regulate the phosphorylation of STAT3, resulting in a synergistic apoptotic effect in KRAS-mutant tumors, including colorectal cancer. In preclinical studies, inhibition of MEK leads to autocrine activation of STAT3 through JAK and FGFR kinase activity, leading to drug resistance. The MEK inhibitor Cobimetinib combined with the JAK1/2 inhibitor Ruxolitinib and the multi-targeted kinase inhibitor Ponatinib (including FGFR inhibitors) showed enhanced efficacy in a mouse xenograft tumor model (Lee et al.Drug Resistance via Feedback Activation of Stat3 in Oncogene-Addicted Cancer Cells. 2019).

In summary, Src, FAK, and JAK2 play an important role in KRAS mutant tumors by regulating angiogenesis in tumors, constructing tumor growth-promoting immune responses in the tumor microenvironment, and signaling intracellularly and extracellularly. However, the development of drugs targeting the central downstream signaling effectors of mutant activated RAS proteins has always been challenging. Combination of SRC/FAK/JAK2 inhibitors with MEK inhibitors, especially Trametinib, represents a new treatment modality that maximizes antitumor activity and response time of MEK inhibitors, especially Trametinib against KRAS mutations patient benefit from treatment.

Compound 1, a new generation of multi-target drug under investigation, can effectively inhibit the activity of SRC/FAK/JAK2 kinase at therapeutic concentrations. In a series of lung cancer, pancreatic cancer and colorectal cancer cells with different types of KRAS mutations, compared with single drug, compound 1 combined with MEK inhibitor Trametinib significantly increased the anti-tumor proliferation activity, and the proportion of tumor cell apoptosis increased. In a subset of tumor subtypes with KRAS mutations, Trametinib combined with compound 1 can inhibit the rebound of AKP phosphorylation (pAKT) level induced by Trametinib. However, the SRC inhibitor Dasatinib, the JAK1/2 inhibitor Ruxolitinib, or the FAK inhibitor Defactinib combined with Trametinib did not inhibit Trametinib-induced AKT phosphorylation, suggesting that simultaneous inhibition of SRC/FAK/JAK2 is required for the increased efficacy of the combination. In the KRAS mutant mouse xenograft experiment, the in vivo anti-tumor effect of the combination was stronger than that of Trametinib alone. The combination of Trametinib and compound 1 can play a stronger and longer-lasting anti-tumor effect by continuously inhibiting the mutant KRAS signaling pathway. Combining MEK inhibitors with drugs that simultaneously inhibit SRC, FAK, and JAK2 may be a promising therapeutic approach to effectively target KRAS-mutated cancers. Furthermore, combined inhibition of SRC, FAK, and JAK2 may have potential roles in other inflammation-related diseases, such as asthma, inflammatory bowel disease, ulcerative colitis, Crohn's disease, and fibrosis.

Contents of the invention

The present invention relates to the use of a compound of formula (I) or its pharmaceutically acceptable salt, stereoisomers, combined with MEK inhibitors in the preparation of medicines for treating diseases,

Wherein said R is hydrogen, halogen, methyl, ethyl.

Further, the use of the present invention, wherein R in the compound of formula (I) is hydrogen or F; more preferably R is F.

Further, the use of the present invention, wherein the compound of formula (I) is an inhibitor of FAK, Src or JAK2.

In some embodiments, the use of the present invention, wherein the disease is cancer or an inflammation-related disease, wherein the cancer is that at least one genetically altered oncogene has previously been identified in a patient, wherein the at least one genetically altered The oncogene is genetically altered Kras, genetically altered MEK; wherein the inflammation-related diseases are asthma, inflammatory bowel disease, ulcerative colitis, Crohn's disease and fibrosis.

Further described purposes, wherein said genetically altered Kras comprises a group selected from G12C, G12V, G12D, G12A, G13C, G12S, D12R, D12F, G13D, G13V, G13R, G13E, Q61H, Q61E, Q61L and Q61R at least one mutation; or selected from G12D, G13D, and Q61H; or KRAS includes at least one mutation other than G12A, G12C, G12S, G12V, and Q61K.

Further, the use of the present invention, wherein the disease is colorectal cancer, pancreatic cancer, lung cancer or gastric cancer.

Further, the use of the present invention, wherein the dose of the compound of formula (I) is about 1 mg to about 1000 mg.

Further, the use of the present invention, wherein the dose of the MEK inhibitor is about 0.5 mg to about 100 mg.

Further, the use of the present invention, wherein the MEK inhibitor is trametinib, slumetinib, LY3214996, R05126766, TNO155 (SHP099) or midametinib or its pharmaceutically acceptable salt or its solvent compounds. Trametinib or a pharmaceutically acceptable salt or solvate thereof is preferred.

Further, the use of the present invention, wherein the compound of formula (I) is administered once a day or twice a day in an amount of about 1-500 mg.

Further, the use of the present invention, wherein the MEK inhibitor is administered in an amount of about 1-100 mg.

In some embodiments the use of the present invention, wherein the compound of formula (I) is administered simultaneously with the MEK inhibitor.

In some embodiments the use of the invention wherein the compound of formula (I) is administered prior to the MEK inhibitor.

In some embodiments the use of the invention wherein the compound of formula (I) is administered after the MEK inhibitor.

In some embodiments the use of the invention wherein said patient has not received previous treatment.

In some embodiments the use of the invention wherein said patient has received at least one prior treatment with one or more chemotherapeutic agents or immunotherapy.

In some embodiments the use of the invention wherein said patient has received at least one prior treatment with one or more chemotherapeutic agents or immunotherapy and has developed acquired resistance to said treatment, and/ or develop bypass resistance to the treatment.

In another aspect the invention provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in a method of treating cancer in a patient in need of such treatment, in combination with a therapeutically effective amount of a MEK inhibitor.

On the other hand, a compound of formula (I) or a pharmaceutically acceptable salt thereof, stereoisomers of the present invention, and its use in the preparation of medicines for treating FAK, SRC or JAK2-related diseases

Wherein said R is hydrogen, halogen, methyl, ethyl; preferably R is H or fluorine, more preferably R is F.

Further said use is cancer or autoimmune disease.

Further the use, wherein the cancer is: lung cancer, pancreatic cancer, rectal cancer.

Further the use, wherein the autoimmune diseases are: asthma, inflammatory bowel disease, ulcerative colitis, Crohn's disease and fibrosis.

The term "cancer" as used herein; includes, but is not limited to, lung cancer, such as non-small cell lung cancer (NSCLC), adenocarcinoma, squamous cell carcinoma of the lung, large cell carcinoma, and large cell neuroendocrine tumors, small cell lung cancer (SCLC), neuroblastoma Cytoma, inflammatory myofibroblastic tumor, adult renal cell carcinoma, pediatric renal cell carcinoma, breast cancer, e.g. triple negative breast cancer, triple positive breast cancer, colon adenocarcinoma, glioblastoma, glia multiforme Blastoma, thyroid cancer, such as anaplastic thyroid cancer, cholangiocarcinoma, ovarian cancer, gastric cancer, such as gastric adenocarcinoma, colorectal cancer (CRC), inflammatory myofibroblastoma, angiosarcoma, epithelioid hemangioendothelioma, liver Internal cholangiocarcinoma, thyroid papillary carcinoma, spittoon tumor, sarcoma, astrocytoma, subcerebral glioma, secretory breast cancer, breast mimic carcinoma, acute myeloid leukemia, congenital mesodermal nephroma, congenital fibrosarcoma, pH-like acute lymphoblastic leukemia, thyroid cancer, skin cancers such as cutaneous melanoma, head and neck squamous cell carcinoma (HNSC), pediatric glioma CML, prostate cancer, ovarian serous bladder cancer, cutaneous melanoma, Castration-resistant prostate cancer, Hodgkin lymphoma, serous and clear cell endometrial carcinoma. It will be understood that the term "cancer"; includes primary cancer or primary tumor and metastatic cancer or metastatic tumor. For example, metastatic NSCLC, metastatic CRC, metastatic pancreatic cancer, metastatic colorectal cancer, metastatic HNSCC, etc. It will be understood that the term "cancer" includes cancers that involve the upregulation of certain genes or inherited mutations in certain genes that can lead to disease progression, such as upregulation of the epidermal growth factor receptor.

In some embodiments of the various aspects described herein, the cancer is mediated by at least one genetically altered oncogene selected from the group consisting of genetically altered KRAS, genetically altered NRAS, genetically altered HRAS, genetically altered BRAF, genetically altered MEK or genetically altered PI3K, or such genetically altered oncogenes. In some embodiments of the various aspects described herein, the cancer is non-small cell lung cancer mediated by a genetically altered KRAS comprising a group selected from G12C, G12V, G12D, G12A, G13C, G12S, D12R, D12F, G13D , at least one mutation among G13V, G13R, G13E, Q61H, Q61E, Q61L, and Q61R. In some embodiments of the aspects described herein, the cancer is non-small cell lung cancer mediated by a genetically altered KRAS comprising at least one mutation selected from the group consisting of G12D, G13D, and Q61H; as described herein In some embodiments of the various aspects described above, the cancer is non-small cell lung cancer mediated by a genetically altered KRAS comprising at least one mutation that is not G12A, G12C, G12S, G12V, or Q61K.

In some embodiments of the various aspects described herein, the cancer is colorectal cancer mediated by a genetically altered KRAS comprising a group selected from G12D, G12V, G13D, A146T, G12C, G12A, G12S, K117N , at least one mutation among Q61K, G12R, M72V, S17G, K5R, D69G, G13C, G13R, Q61H, K117E, Q61L, Q61R, K117R, A146V, A146P, K147N, and R97I. In some embodiments of the various aspects described herein, the cancer is pancreatic cancer mediated by a genetically altered KRAS comprising at least one selected from the group consisting of G12D, G12V, G12R, Q61H, G12C, and G12S mutation.

"KRAS" in the present invention refers to the KRAS gene, the corresponding mRNA produced by the transcription of the KRAS gene, or the protein encoded by the KRAS gene, called K-ras, which involves the Ras/MAPK signaling pathway. The terms KRAS gene, K-ras and Ras/MAPK signaling pathway will be known and understood by those skilled in the art. It is understood that KRAS mutations occur in approximately one-seventh of all metastatic cancers, and that these mutations can occur at various locations within the coding sequence of the KRAS gene. KRAS mutations mainly occur in KRAS codons 12 and 13, but also in low-frequency codons 18, 61, 117, and 146, and have pronounced effects on tumor cell signaling based on codon and missense mutations. Examples of KRAS mutations include, but are not limited to, KRAS G12D, KRAS G12V, KRAS G12R, KRAS G12S, KRAS G13C, KRAS G13D, KRAS A18D, KRASQ61H, KRAS K117N, and the like.

The "MEK inhibitor" used in the present invention; including but not limited to those known in the art to inhibit MAPK/ERK kinase-1 and -2 genes or to inhibit proteins encoded by MAPK/ERK kinase-1 and -2 genes (MEK1 and MEK2 ; MAP2K1 and MAP2K2) any compound or agent. Exemplary MEK inhibitors for use in the methods and compositions described herein include, but are not limited to trametinib, pimasatinib (AST03026); selemetinib (AZD6244); cobimetinib; midametinib Rifatinib (PD-0325901); Rifatinib (RDEA119); TAK733; MEK162; R05126766; WX-554; R04987655; GDC-0973; AZD8330; AZD6244; ; HL-085.

The "trametinib" of the present invention relates to a compound of the following formula or a pharmaceutically acceptable salt or a solvate thereof, also known as GSK1120212 or N-(3-{3-cyclopropyl-5-[( 2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d] Pyrimidin-1(2H)-yl}phenyl)acetamide. Trametinib is an orally bioavailable inhibitor of mitogen-activated protein kinase kinase (MEKMAPK/ERK kinase) with potential antineoplastic activity. Trametinib specifically binds and inhibits MEK1 and 2, resulting in inhibition of growth factor-mediated cell signaling and cell proliferation in various cancers.

The pharmaceutically acceptable salt of trametinib is obtained by salt formation of trametinib with acid, including but not limited to mesylate, maleate, tartrate, succinate, acetate, difluoroacetic acid Salt, fumarate, citrate, citrate, besylate, benzoate, naphthalenesulfonate, lactate, malate, hydrochloride, hydrobromide, sulfate , and phosphates.

"Pharmaceutically acceptable salt" is meant to include salts formed by alkali ions and free acids or salts formed by acid ions and free bases, such as hydrochloride, hydrobromide, sulfate, phosphate, formic acid salt, acetate, trifluoroacetate, fumarate, oxalate, etc.

An "effective amount" or "therapeutically effective amount" includes an amount sufficient to ameliorate or prevent a symptom or condition of a medical condition. An effective amount also means an amount sufficient to permit or facilitate diagnosis. Effective amounts for a particular patient or veterinary subject may vary depending on factors such as the condition being treated, the general health of the patient, the method, route and dosage of administration and the severity of side effects. An effective amount may be the maximum dose or dosing regimen that avoids significant or toxic side effects.

在专利WO2021115401中报道。The structure of compound 1 is:

Reported in patent WO2021115401.

参考专利WO2019210835实施例6制备。The structure of compound 2 is:

Refer to Example 6 of patent WO2019210835 for preparation.

Trametinib was purchased from Shanghai Haoyuan Biomedical Technology Co., Ltd., batch number 30987.

Test Example 1: In vitro Proliferation Inhibition Experiment of Combination Drugs

The purpose of the experiment: By detecting the viability of tumor cells after the effect of the compound, the IC ₅₀ value is used as the index to evaluate the in vitro proliferation inhibitory effect of the combined drug and the single drug on HCT116 and other KRAS mutant cells.

Experimental materials: HCT116, KP-4, SW480 and NCI-H23 were all purchased from Nanjing Kebai, CellCounting-LiteTM2.0

experimental method:

(1) Take the logarithmic growth phase cells HCT116, KP-4, SW480 and NCI-H23, digest with trypsin, centrifuge at 1000rpm for 3 minutes, discard the supernatant, resuspend the medium and count, according to 2-5* ¹⁰³ / Each well was inoculated into a 96-well plate and cultured for 24 hours at 37°C in a 5% CO ₂ incubator.

(2) The compound to be tested was dissolved in 100% DMSO, prepared as a 10 mM stock solution, and stored at 4° C. in the dark. Set single drug wells, combined wells, control wells and blank wells. Single drug wells were compound 1 or compound 2 or Trametinib (initial concentration 3 μM) with gradient concentration, 3-fold dilution, a total of 9 concentrations; combined drug wells were gradient concentration Trametinib and 1 μM compound 1 or compound 2; control wells contained Cells and media; blank wells contain media only. All wells contained 0.5% DMSO. After adding the drug, the cells were placed in a 37°C, 5% CO ₂ incubator to continue culturing for 72h.

(3) Take out the cell culture plate to be tested, equilibrate at room temperature for 30 minutes, add CellCounting-LiteTM2.0 equal to the volume of the cell culture to be tested, shake and mix for 10 minutes to fully lyse the cells, and use the multifunctional enzyme label after standing for 5 minutes The instrument detects the luminescent signal.

data analysis:

Calculation formula: cell viability=(Ls-Lb)/(Lc-Lb)*100%

Among them: Ls represents the luminescence value of the experimental well, Lb represents the luminescence value of the blank well, and Lc represents the luminescence value of the control well.

Fitting the dose-response curve

Taking the log value of the concentration as the X-axis and the cell viability as the Y-axis, the dose-effect curve was fitted using the log (inhibitor) vs. response-variable slope (log (inhibitor) vs. response–Variable slope) of the analysis software GraphPad Prism 5 , so as to obtain the _IC50 value of each compound.

Experimental conclusion: Trametinib combined with compound 1 or compound 2 has significantly stronger inhibitory effects on the in vitro proliferation of tumor cells HCT116, NCI-H23, KP-4, and SW480 than each of them alone.

Experimental example 2: JAK2 kinase activity detection

The purpose of the experiment: to detect the inhibitory activity of the compound on JAK2 kinase

Experimental Materials:

experimental method:

1) Prepare 1X kinase buffer solution (prepared by ddH ₂ O, containing 40mM Tris, 20mM MgCl ₂ , 0.10% BSA, 0.5mMDTT)

2) The compound was diluted 3-fold with DMSO in the dilution plate, and the initial concentration of the compound was 10 μM

3) Dilute the compound 50-fold into 1X kinase reaction buffer and shake on the shaker for 20 minutes

4) Prepare 2X Kinase with 1X Enzyme Reaction Buffer

5) Add 2 μL of kinase (prepared in step 4) to each well of the reaction plate

6) Add 1 μL of the compound diluted in buffer to each well, seal the plate with a sealing film, centrifuge at 1000g for 30 seconds, and place at room temperature for 10 minutes

7) Prepare 4x MBP Protein and ATP (ATP final concentration 10 μM) mixture with 1X enzyme reaction buffer, add 1 μL 4x MBP Protein/ATP mixture to the reaction plate

8) Seal the plate with a sealing film, centrifuge at 1000g for 30 seconds, and react at room temperature for 60 minutes

9) Transfer 4 μL of ADP-Glo to the 384 reaction plate at 1000 rpm, centrifuge for 1 min, and incubate at 25°C for 40 min

10) Transfer 8 μL of Detection solution to the 384 reaction plate at 1000 rpm, centrifuge for 1 min, and incubate at 25°C for 40 min

11) Read the RLU (Relative luminescence unit) signal using a Biotek multifunctional plate reader. Signal intensity is used to characterize the degree of kinase activity

12) Data processing:

a) Compound inhibition rate%=(1-(RLU analyte-RLU positive control)/(RLU negative control-RLU positive control))*100%

b) Data analysis was performed with Graphpad7.0 software. The IC50 (half maximal inhibitory concentration) of the compound was obtained using the following nonlinear fitting formula: Y=Bottom+(Top-Bottom)/(1+10^((LogIC50-X)*Hill Slope)). Wherein, X is the log value of the compound concentration, and Y is the inhibition rate (%inhibition).

13) Experimental results:

Experimental conclusion: compound 1 and compound 2 have significant inhibitory effect on JAK2 kinase.

Experimental example 3: FAK kinase activity detection

The purpose of the experiment: to detect the inhibitory activity of the compound on FAK kinase

Experimental Materials:

experimental method:

1) Prepare 1X kinase buffer solution (prepared by ddH ₂ O, containing 5mM MgCl ₂ ; 1mM DTT; 3.9nM SEB)

2) Dilute the compound to be tested 5 times with 100% DMSO, perform 3-fold equal dilution in a 96-well dilution plate, take 1 μL of the compound and add it to 39 μL of kinase reaction buffer, and shake it on a microplate shaker for 20 minutes

3) Transfer 2 μL of 2.5X kinase to the 384 reaction plate, add 1 μL of the compound to be tested (prepared in step 2) to the 384 reaction plate (Greiner, 784075), centrifuge at 1000 rpm for 1 min, and incubate at 25°C for 10 min

4) Transfer 2 μL of 2.5X substrate mixture to the 384 reaction plate, centrifuge at 1000 rpm for 1 min, and incubate at 25°C for 40 min

5) Prepare 2X Sa-XL 665/TK-antibody-Cryptate mixture with HTRF detection buffer

6) Add 5 μL Sa-XL 665/TK-antibody-Cryptate to each well, centrifuge at 1000 g for 30 seconds, and react at room temperature for 1 hour

7) Read the fluorescent signals at 615nm (Cryptate) and 665nm (XL665) with Biotek

8) Data analysis

a) Calculate the ratio of each well (Ratio_665/615nm)

b) Substituting the ratio of each well into the following formula to calculate the corresponding inhibition rate: compound inhibition rate%=(1-(compound-positive control)/(negative control-positive control))*100%

c) Taking the log value of the compound concentration on the X axis and the percentage inhibition rate on the Y axis, use the analysis software GraphPadPrism7.0 to fit the dose-effect curve to obtain the IC ₅₀ of the compound. The fitting formula is Y=Bottom+(Top-Bottom)/ (1+10^((LogIC50-X)*Hill Slope)).

Experimental results:

Experimental conclusion: compound 1 and compound 2 have significant inhibitory effect on FAK kinase.

Experimental example 4: Src kinase activity detection

The purpose of the experiment: to detect the inhibitory activity of the compound on Src kinase

Experimental Materials:

Materials and Reagents factory Item No. SRC Carna 08-173 Kinase substrate 4 GL 112395 DMSO Sigma D8418-1L 384-well plate Corning 3573 SRC Carna 08-173 Instruments and Equipment factory Item No. or Model centrifuge Eppendorf 5430 Echo 550 Labcyte Echo 550 Microplate reader Perkin Elmer Caliper EZ Reader

experimental method:

1) Prepare 1X kinase buffer solution

2) Preparation of the compound concentration gradient: the test compound test concentration is 10 μM starting, 3-fold dilution, a total of 10 concentrations, single well. Dilute to 100% final concentration of 100% DMSO solution in 384source plate. Use a dispenser Echo 550 to transfer 250 nL of 100-fold final concentration of the compound to the 384-well plate of the destination plate. Positive and negative control wells were added with 250nL DMSO.

3) Prepare a 2.5-fold final concentration kinase solution with 1X kinase buffer solution.

4) Add 10 μL of 2.5-fold final concentration kinase solution to compound wells and positive control wells; add 10 μL 1× kinase buffer solution to negative control wells.

5) Centrifuge at 1000rpm for 30 seconds, shake the reaction plate and incubate at room temperature for 10 minutes.

6) Use 1x kinase buffer solution to prepare a mixed solution of ATP and kinase substrate 4 with a final concentration of 25/15 times.

7) Add 15 μL of a mixed solution of ATP and kinase substrate 4 at 25/15 times the final concentration to start the reaction.

8) Centrifuge the 384-well plate at 1000 rpm for 30 seconds, shake and mix well, and incubate at room temperature for 30 minutes.

9) Add 30 μL of stop detection solution to stop the kinase reaction, centrifuge at 1000 rpm for 30 seconds, shake and mix well.

10) Read the conversion rate with Caliper EZ Reader.

11) Data analysis:

a) Calculation of inhibition rate: compound inhibition rate %=(Conversion%max-Conversion%sample)/(Conversion%max-Conversion%min)*100%, where Conversion%sample is the conversion rate reading of the sample; Conversion%min is negative The average value of the control wells represents the conversion rate readings of the wells without enzyme activity; Conversion%max is the average value of the positive control wells, representing the conversion rate readings of the wells without compound inhibition.

b) Taking the log value of the compound concentration on the X axis and the percent inhibition rate on the Y axis, use the analysis software GraphPadPrism5.0 to fit the dose-effect curve to obtain the IC ₅₀ of the compound. The fitting formula is Y=Bottom+(Top-Bottom)/ (1+10^((LogIC ₅₀ -X)*Hill Slope)).

Experimental results:

Experimental conclusion: both compound 1 and compound 2 have inhibitory effect on Src kinase.

Test Example 5: Antitumor Pharmacodynamics Evaluation of Combination Drugs on the MiniPDX Model of KRAS ^G12V Mutant Lung Cancer Experiment Objective: By establishing the MiniPDX model of KRAS ^G12V mutant lung cancer and detecting the antitumor efficacy of the compound, the relative proliferation rate of tumor cells was used as an index , to evaluate the in vivo proliferation-inhibitory effects of combination therapy and single therapy on KRAS ^G12V mutant lung cancer.

Experimental Materials:

1) In vivo xenograft model information:

model number patient sex patient age mutation type Pathological description LD1-0025-200616 female 75 KRAS^G12V poorly-moderately differentiated adenocarcinoma

2) Experimental animals: 18 BALB/c-Nu mice, female, 6-8 weeks old, weighing 18-22 g, purchased from Jiangsu Jicui Yaokang Biotechnology Co., Ltd.

3) Other reagents and instruments:

experimental method:

1) MiniPDX preparation:

a) For the tumor model required for the recovery experiment, when the tumor grows to 500-800 mm ³ , the tumor tissue is aseptically removed by surgery, and the non-tumor tissue and necrotic tissue are removed in a biological safety cabinet;

b) Cut the tumor tissue into small pieces of 1-3 mm3, digest the tumor piece with digestion solution at 37°C for 1-2 hours, then collect the cell suspension through a 70μm sieve, centrifuge at 1200rpm for 3 minutes to remove the supernatant, and use 10mL containing 1 Cells were resuspended in PBS with %FBS and counted on a hemocytometer;

c) After removing the mouse-derived cells, centrifuge at 1200rpm for 3 minutes to remove the supernatant, resuspend the cells with BIO-MPM-1 cell culture medium (serum-free), count on a hemocytometer, and adjust the cell density. Fill the cell suspension into the MiniPDX device;

d) The mice were randomly grouped according to body weight; the MiniPDX device was inoculated subcutaneously on the back of the mice, and each mouse was inoculated with 2 MiniPDX devices. The day of inoculation was Day 0, and the whole experiment was carried out for 7 days.

2) Grouping and administration of mice:

On day 0, grouping and administration were performed according to the table below.

Note: N: number of repetitions; administration volume is 10 μl/g; QD×7: once a day, 7 times in total; p.o.: oral administration.

Throughout the experiment, the use and observation of experimental animals were carried out in accordance with the relevant regulations of AAALAC animal use and management. After the experimental animals were inoculated with the MiniPDX tumor device, they were observed every day, and the behavior, food intake, water intake and hair of all experimental animals were monitored, and the body weight of the mice was recorded.

Twenty-four hours after the administration on the sixth day (ie, the seventh administration), the experiment ended, all mice were euthanized, and the MiniPDX device was taken out. Cell viability was detected by CellTiter-Glo method.

data processing:

1) Calculation formula:

a) Relative tumor cell proliferation rate (%)=(V drug treatment group d ₇ -V control group d ₀ )/(V control group d ₇ -V control group d ₀ )×100%. (V drug treatment group d7: fluorescence value of the administration group on the 7th day; V control group d ₇ : fluorescence value of the control group on the 7th day; V control group d ₀ : fluorescence value of the control group on the 0th day).

b) The body weight of all tumor-bearing mice was measured once a day. Simultaneously calculate the change ratio of mouse body weight after administration: RCBW (%)=(BW _i -BW ₀ )/BW ₀ ×100, BWi is the average body weight after the start of administration, and BW ₀ is the average body weight at the time of the first administration .

2) Data analysis:

All the data are represented by Mean ± SEM, and the Student's t-test is used to compare whether there is a significant difference in the fluorescence value after CTG detection between the treatment group and the control group. All data were analyzed with Graphpad. P<0.05 was considered to have a significant difference.

Experimental results:

In the LD1-0025-200616KRAS ^G12V lung cancer MiniPDX pharmacodynamic model, compound 1, 10mg/kg, TPX-0005, 10mg/kg, trametinib, 1mg/kg, trametinib, 1mg/kg+TPX-0005 , 10mg/kg and trametinib, 1mg/kg+compound 1, 10mg/kg administration group, the relative proliferation rates of tumor cells were 57%, 37%, 46%, 55% and 28% respectively on the 7th day after group administration. %. Compared with the control group, the fluorescence value (RLU) of trametinib, 1mg/kg + compound 1, 10mg/kg combined administration group was significantly reduced (p<0.05), showing a significant tumor inhibitory effect. In addition, compared with the corresponding monotherapy group, trametinib, 1 mg/kg + compound 1, 10 mg/kg combined administration group had a synergistic or additive effect. During the administration period, the mice in all the administration groups did not show a significant decrease in body weight (RCBW%<-15%), and no other abnormal behaviors and performances were observed. The following drugs were well tolerated.

Experimental conclusion: trametinib combined with compound 1 has a significantly stronger inhibitory effect on the proliferation of tumor cells in the miniPDX model of KRAS ^G12V mutant lung cancer than trametinib or compound 1 alone. The combination of compound 1 and trametinib was significantly better than the combination of TPX-0005 and trametinib.

Test ^{Example 6: Evaluation of antitumor pharmacodynamics of combined drugs on the MiniPDX model of KRASG12D mutant} lung cancer , to evaluate the in vivo proliferation-inhibitory effects of combination therapy and monotherapy on KRAS ^G12D -mutant lung cancer.

Experimental Materials:

1) In vivo xenograft model information:

model number patient sex patient age mutation type Pathological description LD1-0025-370740 male 70 KRAS^G12D poorly differentiated adenocarcinoma

2) Experimental animals: 18 BALB/c-Nu mice, female, 6-8 weeks old, weighing 18-22 g, purchased from Jiangsu Jicui Yaokang Biotechnology Co., Ltd.

3) Other reagents and instruments:

experimental method:

1) MiniPDX preparation:

a) For the tumor model required for the recovery experiment, when the tumor grows to 500-800 mm ³ , the tumor tissue is aseptically removed by surgery, and the non-tumor tissue and necrotic tissue are removed in a biological safety cabinet;

b) Cut the tumor tissue into small pieces of 1-3 mm3, digest the tumor piece with digestion solution at 37°C for 1-2 hours, then collect the cell suspension through a 70μm sieve, centrifuge at 1200rpm for 3 minutes to remove the supernatant, and use 10mL containing 1 Cells were resuspended in PBS with %FBS and counted on a hemocytometer;

c) After removing the mouse-derived cells, centrifuge at 1200rpm for 3 minutes to remove the supernatant, resuspend the cells with BIO-MPM-1 cell culture medium (serum-free), count on a hemocytometer, and adjust the cell density. Fill the cell suspension into the MiniPDX device;

d) The mice were randomly grouped according to body weight; the MiniPDX device was inoculated subcutaneously on the back of the mice, and each mouse was inoculated with 2 MiniPDX devices. The day of inoculation was Day 0, and the whole experiment was carried out for 7 days.

2) Grouping and administration of mice:

On day 0, grouping and administration were performed according to the table below.

Note: N: number of repetitions; administration volume is 10 μl/g; QD×7: once a day, 7 times in total; p.o.: oral administration.

Throughout the experiment, the use and observation of experimental animals were carried out in accordance with the relevant regulations of AAALAC animal use and management. After the experimental animals were inoculated with the MiniPDX tumor device, they were observed every day, and the behavior, food intake, water intake and hair of all experimental animals were monitored, and the body weight of the mice was recorded.

Twenty-four hours after the administration on the sixth day (ie, the seventh administration), the experiment ended, all mice were euthanized, and the MiniPDX device was taken out. Cell viability was detected by CellTiter-Glo method.

data processing:

1) Calculation formula:

a) Relative tumor cell proliferation rate (%)=(V drug treatment group d ₇ -V control group d ₀ )/(V control group d ₇ -V control group d ₀ )×100%. (V drug treatment group d7: fluorescence value of the administration group on the 7th day; V control group d ₇ : fluorescence value of the control group on the 7th day; V control group d ₀ : fluorescence value of the control group on the 0th day).

b) The body weight of all tumor-bearing mice was measured once a day. Simultaneously calculate the change ratio of mouse body weight after administration: RCBW (%)=(BW _i -BW ₀ )/BW ₀ ×100, BWi is the average body weight after the start of administration, and BW ₀ is the average body weight at the time of the first administration .

2) Data analysis:

All the data are represented by Mean ± SEM, and the Student's t-test is used to compare whether there is a significant difference in the fluorescence value after CTG detection between the treatment group and the control group. All data were analyzed with Graphpad. P<0.05 was considered to have a significant difference.

Experimental results:

In the LD1-0025-370740KRAS ^G12D lung cancer MiniPDX pharmacodynamic model, compound 1, 10mg/kg, TPX-0005, 10mg/kg, trametinib, 1mg/kg, trametinib, 1mg/kg+TPX-0005 , 10mg/kg and trametinib, 1mg/kg+ compound 1, 10mg/kg administration group, the relative proliferation rate of tumor cells was 97%, 60%, 30%, 54% and 19% respectively on the 7th day after group administration. %. Compared with the control group, the fluorescence values (RLU) of all the administration groups showed different degrees of reduction, among which trametinib, 1mg/kg administration group and trametinib, 1mg/kg+compound 1, 10mg/kg The fluorescence value of the administration group was significantly reduced (p<0.05), showing a significant tumor-inhibiting effect. In addition, compared with the corresponding monotherapy group, trametinib, 1 mg/kg + compound 1, 10 mg/kg combined administration showed synergistic or additive effects. During the administration period, in the Trametinib, 1mg/kg+Compound 1, 10mg/kg administration group, the body weight of one mouse decreased by 15.14% on the 6th day, and the body weight of the other two mice in this group changed normally. At the same time, the mice in all other administration groups did not show a significant decrease in body weight (RCBW%<-15%), and no other abnormal behaviors and performances were observed, indicating that all tumor-bearing mice have no effect on the model. Each administration group under the reagent dose can tolerate it, and the weight loss of individual mice may be caused by individual differences.

Experimental conclusion: trametinib combined with compound 1 has a significantly stronger inhibitory effect on the proliferation of tumor cells in the KRAS ^G12D mutant lung cancer miniPDX model than trametinib or compound 1 alone. The combination of compound 1 and trametinib was significantly better than the combination of TPX-0005 and trametinib.

Claims (22)

1. A compound of formula (I) or a pharmaceutically acceptable salt thereof, stereoisomers, combined with MEK inhibitors in the preparation of medicines for treating diseases

Wherein said R is hydrogen, halogen, methyl, ethyl. 2. The use according to claim 1, wherein R in the compound of formula (I) is hydrogen or F; more preferably R is F. 3. The use according to claim 1 or 2, wherein the compound of formula (I) is an inhibitor of FAK, SRC or JAK2. 4. The use according to claim 1, wherein the disease is cancer or an inflammation-related disease, wherein the cancer is at least one genetically altered oncogene previously identified in a patient, wherein the at least one genetically altered The oncogene is genetically altered KRAS, genetically altered MEK; wherein the inflammation-associated disease is asthma, inflammatory bowel disease, ulcerative colitis, Crohn's disease and fibrosis. 5. The use according to claim 4, wherein said genetically altered KRAS comprises a group selected from G12C, G12V, G12D, G12A, G13C, G12S, D12R, D12F, G13D, G13V, G13R, G13E, Q61H, Q61E, Q61L and at least one mutation in Q61R; or selected from G12D, G13D, and Q61H; or KRAS includes at least one mutation other than G12A, G12C, G12S, G12V, and Q61K. 6. The use according to any one of claims 1-5, wherein the disease is colorectal cancer, pancreatic cancer, lung cancer or gastric cancer. 7. The use according to any one of claims 1-6, wherein the compound of formula (I) is administered in an amount of about 1 mg to about 1000 mg. 8. The use according to any one of claims 1-7, wherein the MEK inhibitor is administered in an amount of about 0.5 mg to about 100 mg. 9. The use according to any one of claims 1-8, wherein the MEK inhibitor is Trametinib, Slumetinib, LY3214996, R05126766, TNO155 (SHP099) or Midametinib or its pharmaceutical An acceptable salt or a solvate thereof; preferably Trametinib or a pharmaceutically acceptable salt or a solvate thereof. 10. The use according to any one of claims 1-9, wherein the compound of formula (I) is administered once a day or twice a day in an amount of about 1-500 mg. 11. The use according to any one of claims 1-9, wherein the MEK inhibitor is administered in an amount of about 1-100 mg. 12. The use according to any one of claims 1-11, wherein the compound of formula (I) is administered simultaneously with the MEK inhibitor. 13. The use according to any one of claims 1-11, wherein the compound of formula (I) is administered before the MEK inhibitor. 14. The use according to any one of claims 1-11, wherein the compound of formula (I) is administered after the MEK inhibitor. 15. The use according to any one of claims 4-11, wherein the patient has not received previous treatment. 16. The use according to any one of claims 4-11, wherein the patient has received at least one prior treatment with one or more chemotherapeutic agents or immunotherapy. 17. Use according to any one of claims 4-11, wherein the patient has received at least one prior treatment with one or more chemotherapeutic agents or immunotherapy and has developed acquired resistance to said treatment resistance, and/or develop bypass resistance to the treatment. 18. A method of treating cancer in a patient in need of such treatment, in combination with a compound of formula (I) or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a MEK inhibitor. 19. A compound of the formula (I) or its pharmaceutically acceptable salt, stereoisomer, in the preparation of medicines for the treatment of FAK, SRC or JAK2-related diseases

Wherein said R is hydrogen, halogen, methyl, ethyl; preferably R is H or F. 20. The use according to claim 19, said disease is cancer or autoimmune disease. 21. Use according to claim 20, wherein said cancer is: lung cancer, pancreatic cancer, rectal cancer. 22. Use according to claim 21, wherein said autoimmune diseases are: asthma, inflammatory bowel disease, ulcerative colitis, Crohn's disease and fibrosis.