Application of ADAM9 inhibitors in the preparation of drugs for preparing immunomodulators

Abstract

The use of ADAM9 inhibitors in the preparation of drugs for the preparation of immunomodulators, which can change the distribution of immune cells in tumors and increase the infiltration of immune cells, thereby enhancing the effectiveness of cancer immunotherapy.

Description

Technical Field

[0001] The present invention relates to a use of an ADAM9 inhibitor, in particular to the use of the ADAM9 inhibitor as an immunomodulator. Background Art

[0002] Cancer, also known as malignant tumor, is an abnormal proliferation of cells, and these proliferating cells may invade other parts of the body. It is a disease caused by the abnormal control of cell division and proliferation. The number of people suffering from cancer in the world is increasing. Cancer is one of the top ten causes of death in China, and has been the top of the list for 27 consecutive years. The occurrence of cancer metastasis is the main cause of high cancer mortality. For example, the prognosis of brain metastasis of lung cancer is extremely poor. Even if early lung cancer patients can undergo surgery to remove the tumor, 25% will still develop remote cancer metastasis. Although common anticancer drugs can be used to inhibit the growth of cancer cells, they are still limited in preventing cancer recurrence. Targeting the molecular effects that promote metastasis can be regarded as a forward-looking approach.

[0003] Disintegrin and metalloproteinase domain-containing protein 9 (ADAM9) is overexpressed in many tumors, such as pancreatic cancer, breast cancer, prostate cancer, and lung cancer, and high expression of ADAM9 is associated with poor prognosis in cancer patients and can be used as a predictive marker. Because ADAM9 expression can help tumor cells adapt to adverse environments, ADAM9 is believed to promote tumor development and is considered a better therapeutic target than other cancer-related metalloproteinases. ADAM9 participates in tumorigenesis due to its ability to cut and release many molecules involved in cancer progression, and ADAM9 secreted by stromal cells around tumors can promote tumor development through neovascularization.

[0004] Previous studies have shown that inhibiting ADAM9 expression and its downstream signaling can significantly prolong the survival of mice with lung tumors. Clinical samples of lung cancer and breast cancer clearly show that patients with low ADAM9 expression levels in tumor specimens survive longer than those with high ADAM9 expression levels. In addition, since ADAM9-deficient mice have no obvious phenotype, it means that ADAM9 as a target protein may have good drug tolerance. Summary of the invention

[0005] One aspect of the present invention is to provide a use of an ADAM9 inhibitor in the preparation of a drug for preparing an immunomodulator, wherein the ADAM9 inhibitor is selected from:

[0006]

[0007] and

[0008]

[0009] According to the aforementioned application, the immunomodulator can be used to enhance the effectiveness of cancer immunotherapy. Preferably, the subject of the cancer immunotherapy can be a patient with low immune function.

[0010] According to the aforementioned application, the immunomodulator can be used to stimulate immune cells to infiltrate into tumors. Preferably, the immune cells can be T cells, natural killer cells, macrophages, neutrophils, dendritic cells or bone marrow-derived suppressor cells.

[0011] According to the aforementioned application, the immunomodulator can be used to change the types of chemokines in the tumor microenvironment.

[0012] According to the aforementioned application, the immunomodulator can be used to treat tumors with resistance to immunotherapy. Preferably, the tumor with resistance to immunotherapy may be resistant to checkpoint inhibitors, adoptive cell transfer, therapeutic antibodies, therapeutic vaccines, cytokines, immune cell therapy or a combination thereof. The checkpoint inhibitor may be an anti-CTLA-4 antibody, an anti-PDL1 antibody or an anti-PD-1 antibody.

[0013] According to the aforementioned application, the immunomodulator can be co-formulated with a checkpoint inhibitor. Preferably, the checkpoint inhibitor can be an anti-CTLA-4 antibody, an anti-PDL1 antibody or an anti-PD-1 antibody.

[0014] Thus, the ADAM9 inhibitor of the embodiment of the present invention can have an immunomodulatory function by changing the distribution of immune cells in the tumor and increasing the infiltration of immune cells, and is suitable for preparing an immunomodulator. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] In order to make the above and other objects, features, advantages and embodiments of the present invention more clearly understood, the accompanying drawings are described as follows:

[0016] Figure 1A and Figure 1B This is a diagram showing the analysis results of the inhibition of ADAM9 activity by the compound of formula (I);

[0017] Figure 2A , Figure 2B , Figure 2C , Figure 2D , Figure 2E , Figure 2F , Figure 2G and Figure 2H The figure is a characteristic analysis result diagram of the compound of formula (I) in in vitro test;

[0018] Figure 3 This is a diagram of the analysis results after eliminating the effect of ADAM9 on tumor growth;

[0019] Figure 4A , Figure 4B , Figure 4C and Figure 4D This is the result of RNA analysis of ADAM9, CD4, CD8 and IFN-γ in tumor tissues after ADAM9 was deleted;

[0020] Figure 5A and Figure 5B This is a diagram of the analysis results after eliminating the effects of ADAM9 on lung metastatic tumor growth and immune cell infiltration;

[0021] Figure 6 This is a graph showing the results of analyzing the apoptosis rate of target cells and effector cells after 24 hours of co-culture;

[0022] Fig. 7A This is a diagram showing the analysis results of the anti-tumor effect of ADAM9 inhibitors;

[0023] Figure 7B Flow chart of the use of ADAM9 inhibitors to treat syngeneic orthotopic breast tumor animal models;

[0024] Fig. 8A and Figure 8B This is a diagram showing the analysis results of the effect of ADAM9 inhibitors on immune cell infiltration;

[0025] Fig. 9 Figure 1 shows the results of the analysis for co-treatment of ADAM9 inhibitors and checkpoint inhibitors; and

[0026] Fig. 10A , Fig. 10B and Fig. 10C This is a diagram showing the analysis results of ADAM9 inhibitors stimulating tumor cells to secrete cytokines. DETAILED DESCRIPTION

[0027] The following description of specific embodiments and examples is for illustration only and not for limitation, and those skilled in the art may make various modifications or change various non-critical parameters to produce substantially similar results. The present invention provides a variety of embodiments.

[0028] The present invention provides a use of an ADAM9 inhibitor, which is used to prepare an immunomodulator. The immunomodulator can be used to stimulate immune cells to infiltrate into tumors. Preferably, the immune cells can be T cells, natural killer cells, macrophages, neutrophils, dendritic cells or suppressor cells derived from bone marrow. In addition, the immunomodulator can be used to change the type of chemokines in the tumor microenvironment or to enhance the effectiveness of cancer immunotherapy. In particular, the subject of administration of the cancer immunotherapy can be a patient with low immune function.

[0029] The present invention further provides a use of an ADAM9 inhibitor, which is used to prepare an immunomodulator. The immunomodulator can be used to treat tumors with resistance to immunotherapy. Preferably, the tumor with resistance to immunotherapy may be resistant to checkpoint inhibitors, adoptive cell transfer, therapeutic antibodies, therapeutic vaccines, cytokines, immune cell therapy or a combination thereof. Wherein the checkpoint inhibitor may be a molecule used to inhibit immune checkpoint proteins in immune cell activation, preferably, an anti-CTLA-4 antibody, an anti-PDL1 antibody or an anti-PD-1 antibody.

[0030] The aforementioned ADAM9 inhibitor can also be used in combination with a checkpoint inhibitor to enhance the effectiveness of cancer immunotherapy. In addition, the aforementioned ADAM9 inhibitor can also be used in combination with a chemotherapeutic agent and / or a checkpoint inhibitor as a pharmaceutical composition for treating cancer.

[0031] The ADAM9 inhibitor may be a compound of formula (I), or a pharmaceutically acceptable salt or stereoisomer thereof,

[0032]

[0033] wherein R is a C1-C ... 18 or a C1-C ... 18 A hydrocarbon group, and R contains C5-C 10 An organic ring which is an aryl group containing 0 to 3 heteroatoms independently selected from nitrogen, sulfur and oxygen.

[0034] According to the aforementioned compound of formula (I), R can be a secondary amine, an amide, NHR1, NHCOR2 or a structure as shown in formula (i):

[0035]

[0036] Wherein R1 may be an aryl group containing 0 to 3 heteroatoms independently selected from nitrogen, sulfur and oxygen. Preferably, R1 may be 2-thiazol-4-yl-ethyl-isoindoline-1,3-dione. R2 may be a phenyl group or a benzoheterocyclic group containing 0 to 3 heteroatoms independently selected from nitrogen, sulfur and oxygen. Preferably, R2 may be benzothiophene. R3 may be an optionally substituted phenyl group, a benzoheterocyclic group or a heterocyclic group containing 0 to 3 heteroatoms independently selected from nitrogen, sulfur and oxygen.

[0037] Unless otherwise specified, "a" and "an" as used herein and in this specification refer to the grammatical object of the article as one or more (i.e., at least one). In addition, a class is an abbreviation for all members included in the class, for example, C1-C3 alkyl is an abbreviation for all C1-C3 alkyl groups, for example, C1-C3 alkyl includes methyl, ethyl, propyl and isomers thereof.

[0038] The following terms, phrases and symbols are generally intended to have the meanings set forth below, unless the context in which they are used indicates otherwise. The following abbreviations and terms have the indicated meanings.

[0039] The "alkenyl" described in the present specification refers to a straight chain and branched hydrocarbon group containing at least one C=C double bond and 2-18, 2-12 or 2-6 carbon atoms. Examples of alkenyl groups may be selected from, but are not limited to, vinyl or vinyl, prop-1-enyl, prop-2-enyl, 2-methylprop-1-enyl, but-1-enyl, but-2-enyl, but-3-enyl, but-1,3-dienyl, 2-methylbut-1,3-diene, hex-1-enyl, hex-2-enyl, hex-3-enyl, hex-4-enyl and hexa-1,3-diene groups.

[0040] The "aryl" described in this specification refers to a group selected from the following: 5- and 6-membered carbocyclic aromatic rings, such as phenyl, bicyclic systems and tricyclic systems. The bicyclic system may be a 7-12-membered bicyclic system, wherein at least one ring is a carbocyclic ring and an aromatic ring, which is selected from, but not limited to, naphthalene, indane and 1,2,3,4-tetrahydroquinoline. The tricyclic system may be a 10-15-membered tricyclic system, wherein at least one ring is a carbocyclic ring and an aromatic ring, such as fluorene. For example, the aryl group is selected from a 5- and 6-membered carbocyclic aromatic ring fused with a 5- to 7-membered cycloalkyl or heterocyclic ring optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur. When the carbocyclic aromatic ring is fused with a heterocyclic ring, the connection point is on the carbocyclic aromatic ring, and when the carbocyclic aromatic ring is fused with a cycloalkyl, the connection point may be on the carbocyclic aromatic ring or the cycloalkyl. A divalent group formed by a substituted benzene derivative and having a free valence on a ring atom is referred to as a substituted phenylene group. Divalent radicals derived from monovalent polycyclic hydrocarbon radicals whose names end in "-idene" by removing one hydrogen atom from the carbon atom with the free valence are named by adding "-yl" to the name of the corresponding monovalent radical, e.g. naphthyl with two points of attachment is called naphthylene. However, aryl does not include or overlap with heteroaryl, which are defined separately below in this specification. Thus, if one or more carbocyclic aromatic rings are fused to a heterocyclic aromatic ring, the resulting ring system is a heteroaryl, not an aryl as defined in this specification.

[0041] As used herein, "heteroaryl" refers to a group selected from the following: a 5- to 7-membered aromatic monocyclic ring containing 1, 2, 3 or 4 heteroatoms selected from nitrogen, oxygen and sulfur, and the remaining ring atoms are carbon; an 8- to 12-membered bicyclic ring containing 1, 2, 3 or 4 heteroatoms selected from nitrogen, oxygen and sulfur, the remaining ring atoms are carbon, and at least one ring is aromatic and at least one heteroatom is present in the aromatic ring. For example, the heteroaryl group comprises a 5- to 7-membered heterocyclic aromatic ring fused to a 5- to 7-membered cycloalkyl ring. The aforementioned fused bicyclic heteroaryl ring system, in which only one ring contains at least one heteroatom, the connection point can be on the heteroaryl ring or the cycloalkyl ring. When the total number of sulfur and oxygen atoms in the heteroaryl group exceeds 1, the heteroatoms are not adjacent to each other. In some embodiments, the total number of sulfur and oxygen atoms in the heteroaryl group is not greater than 2, and in other embodiments, the total number of sulfur and oxygen atoms in the aromatic heterocyclic ring is not greater than 1. Examples of heteroaryl groups include, but are not limited to (numbering from the link position assigned a priority of 1), pyridyl (e.g., 2-pyridyl, 3-pyridyl, or 4-pyridyl), quinolyl, pyrazinyl, 2,4-pyrimidinyl, 3,5-pyrimidinyl, 2,4-imidazolyl, imidazopyridinyl, isoxazolyl, oxazolyl, thiazolyl, isothiazolyl, thiadiazolyl, tetrazolyl, thienyl, triazinyl, benzothienyl, furanyl, benzofuranyl, benzimidazolyl, indolyl, isoindolyl, indolinyl, phthalazinyl, pyrazinyl, pyridazinyl, pyrrolyl, triazolyl, quinolyl, isoquinolyl, pyrazolyl, pyrrolopyridinyl (e.g., 1H-pyrrolo[2,3-b]pyridine-5-yl) 1H-pyrazolo[3,4-b]pyridin-5-yl), benzoxazolyl (e.g., benzo[d]oxazol-6-yl), piperidinyl, purinyl, 1-oxa-2,3-oxadiazolyl, 1-oxa-2,4-oxadiazolyl, 1-oxa-2,5-oxadiazolyl, 1-oxa-3,4-oxadiazolyl, 1-thiophene-3,4-oxadiazolyl, furanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, furopyridinyl, benzothiazolyl (e.g., benzo[d]thiazol-6-yl), indazolyl (e.g., 1H-indazol)-5-yl), and 5,6,7,8-tetrahydroisoquinoline.

[0042] Compounds may contain asymmetric centers and therefore may exist as mirror image isomers. When a compound has two or more asymmetric centers, it may additionally exist as non-mirror image isomers. Mirror image isomers and non-mirror image isomers belong to a wider class of stereoisomers. All possible stereoisomers include substantially pure separable mirror image isomers, racemic mixtures thereof, and mixtures of non-mirror image isomers. All stereoisomers of a compound and / or its pharmaceutically acceptable salts are included where applicable. Unless otherwise indicated, the isomers mentioned are applicable to any possible isomers. And when an isomeric composition is not specified, all possible isomers are included.

[0043] The "pharmaceutically acceptable salts" described in this specification include, but are not limited to, salts formed with inorganic acids and salts formed with organic acids. Salts formed with inorganic acids can be selected from, for example, hydrochlorides, phosphates, diphosphates, hydrobromides, sulfates, sulfinates and nitrates. Salts formed with organic acids can be selected from, for example, maleates, fumarates, tartrates, succinates, citrates, lactates, methanesulfonates, p-toluenesulfonates, 2-hydroxyethylsulfonates, benzoates, salicylates, stearates, alkanoates (e.g., acetates) and salts formed with HOOC-(CH2) n -COOH, wherein n is 0 to 4. Examples of pharmaceutically acceptable cations include, but are not limited to, sodium, potassium, calcium, aluminum, lithium, and ammonium.

[0044] In addition, if the compound is an acid addition salt, the free base can be obtained by basifying a solution of the acid salt. In contrast, if the product is a free base, an addition salt (e.g., a pharmaceutically acceptable addition salt) can be prepared by dissolving the free base in a suitable organic solvent and treating the solution with an acid, and then following conventional methods for preparing acid addition salts from base compounds. Various synthetic methods that can be used by those skilled in the art without undue experimentation can prepare non-toxic pharmaceutically acceptable addition salts.

[0045] The term "treatment" as used herein refers to administering at least one compound and / or at least one stereoisomer and / or at least one pharmaceutically acceptable salt thereof to a subject in need thereof, such as a cancer patient.

[0046] The "effective amount" described in this specification refers to the amount of at least one compound and / or at least one stereoisomer thereof and / or at least one pharmaceutically acceptable salt thereof that is effective in "treating" a disease or condition of a subject. The effective amount is related to the biological or medical response of the tissue, system, animal or human to which it is administered, for example, when administered, it is sufficient to prevent the development of one or more diseases or conditions or alleviate the symptoms of one or more conditions or conditions being treated to a certain extent. The therapeutically effective amount will vary depending on the compound, the disease and its severity, and the age and weight of the mammal to be treated.

[0047] As used herein, "cancer" refers to or describes a physiological condition in mammals that is typically characterized by unregulated cell growth. "Tumor" includes one or more cancer cells. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia or lymphoid malignancies. More specific examples of such cancers include squamous cell carcinoma (e.g., epithelial squamous cell carcinoma), lung cancer (including small cell lung cancer, non-small cell lung cancer (NSCLC), lung adenoma, and lung squamous cell carcinoma), peritoneal cancer, hepatocellular carcinoma, gastric cancer (including gastrointestinal cancer), pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, rectal cancer, colorectal cancer, endometrial cancer or uterine cancer, salivary gland cancer, kidney cancer, prostate cancer, vulvar cancer, thyroid cancer, liver cancer, anal cancer, penile cancer, and head and neck cancer.

[0048] The compounds of the present invention, their stereoisomers and pharmaceutically acceptable salts thereof can be used alone or in combination with at least one other therapeutic agent for treatment. In some embodiments, the compounds, their stereoisomers and pharmaceutically acceptable salts thereof can be used in combination with at least one other therapeutic agent. The at least one other therapeutic agent can be selected from anti-hyperproliferative agents, anticancer agents and chemotherapeutic agents. The compounds and / or a pharmaceutically acceptable salt disclosed in this specification can be administered together with at least one other therapeutic agent in a single dosage form, or as a separate dosage form. When administered as a separate dosage form, at least one other therapeutic agent can be administered before, simultaneously or after the administration of the compounds and / or a pharmaceutically acceptable salt disclosed in this specification.

[0049] The "chemotherapeutic agent" described in this specification refers to a chemical compound used to treat cancer regardless of its mechanism of action. Chemotherapeutic agents include compounds used for "targeted therapy" and conventional chemotherapy. Suitable chemotherapeutic agents can be selected from: apoptosis-inducing agents, polynucleotides (such as ribozymes), polypeptides (such as enzymes), drugs, biomimetic substances, alkaloids, alkylating agents, antitumor antibiotics, antimetabolites, hormones, platinum compounds, monoclonal antibodies bound to anticancer drugs, toxins and / or radionuclides, biological response modifiers (such as interferons such as IFN-α and interleukins such as IL-2), adoptive immunotherapy agents, hematopoietic growth factors, agents that induce tumor cell differentiation (such as all-trans retinoic acid), gene therapy agents, antisense therapy agents and nucleotides, tumor vaccines, and angiogenesis inhibitors.

[0050] Examples of chemotherapeutic agents include, but are not limited to, erlotinib. Genentech / OSIPharm; Bortezomib, Millennium Pharm; Fulvestrant, AstraZeneca; Sunitinib Pfizer); Letrozole, Novartis); Imatinib mesylate, Novartis); PTK787 / ZK 222584 (Novartis); Oxaliplatin ( Sanofi); 5-FU (5-fluorouracil); leucovorin; rapamycin (Rapamycin, sirolimus, Wyeth; Lapatinib GSK572016, Glaxo SmithKline; Lonafarnib (SCH 66336); Sorafenib (Sorafenib, Bayer); Irinotecan (Irinotecan, Pfizer) and gefitinib (Gefitinib, AstraZeneca); AG1478, AG1571 (SU 5271, Sugen); alkylating agents such as thiotepa and cyclophosphamide; alkyl sulfonates such as busulfan, improsulfan and piposulfan; azepanes such as benzodepa, carboquone, meturedopa and uredopa; ethyleneimines and methylmelamines such as altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide and trimethylolomelamine; acetylsalicylic acids such as acetylsalicylic acids; etogenins, such as bullatacin and bullatacinone; camptothecins, such as the synthetic analog topotecan; bryostatin; callystatin, CC-1065 and its synthetic analogs adozelesin, carzelesin and bizelesin; cryptophycins, such as cryptophycin I and cryptophycin 8; dolastatin; duocarmycin and its synthetic analogs (such as KW-2189 and CB1-TM1); eleutherobin; pancratistatin; sarcodictyin; spongistatin;Nitrogen mustards, for example, chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard, mustard; nitrosoureas, such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine and ranimustine; antibiotics, such as the enediynes (e.g., calicheamicins, especially calicheamicin gamma 1I and calicheamicin omega 1I); anthracyclines (dynemicins), including dynemicins A; bisphosphonates, such as clodronate; esperamicin; and neocarzinostatin chromophores and related chromoprotein enediyne antibiotic chromophores, aclacinomycin, actinomycin, anthramycin, azaserine, bleomycin, cactinomycin, carabicin, carminomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin, detorubicin, 6-diaza-5-oxo-L-norleucine; Doxorubicin, including morpholinodoxorubicin, cyanomorpholinodoxorubicin, 2-pyrrolodoxorubicin and deoxydoxorubicin; epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins such as mitomycin C, mycophenolic acid acid, nogalamycin, olivomycin, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites, such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs, such as denopterin, methotrexate, pteropterin, terin), trimetrexate; purine analogs, such as fludarabine, mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs, such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine; androgens, such as calusterone, dromostanolone propionate, propionate, epitiostanol, mepitiostane, testolactone; antiadrenal agents, such as aminoglutethimide, mitotane, trilostane; folic acid supplements, such as folinic acid; aceglatone;aldophosphamideglycoside; aminolevulinic acid; eniluracil; amsacrine; bestrabucil; bisantrene; edatraxate; defosfamide; demecolcine; diaziquone; elfornithine; elliptinium acetate acetate; epothilone; etoglucid; gallium nitrate; hydroxyurea; lentinan; lonidamine; maytansinoids, such as maytansine and ansamitocin; mitoguazone; mitoxantrone; mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; losoxantrone; podophyllinic acid; 2-ethylhydrazide; procarbazine; polysaccharide complex (JHS Natural Products, Eugene, OR; razoxane; rhizoxin; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2,2',2"-trichlorotriethylamine; trichothecenes, especially T-2 toxin, verrucarin A, roridin A, and anguidin; urethan; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C");Cyclophosphamide; thiotepa; taxoids, for example; Paclitaxel (Bristol-Myers Squibb Oncology, Princeton, NJ), Cremophor, albumin-modified nanoparticle formulation of paclitaxel (American Pharmaceutical Partners, Schaumberg, Illinois), and doxetaxel (Rhone-Poulenc Rorer, Antony, France); chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs, such as cisplatin, oxaliplatin, and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; Vinorelbine; novantrone; teniposide; edatrexate; daunomycin; aminopterin; xeloda; ibandronate; irinotecan (Camptosar, CPT-11); topoisomerase inhibitor RFS2000; difluoromethylornithine (DMFO); retinoids (e.g., retinoic acid); and any pharmaceutically acceptable salts, acids, and derivatives thereof.

[0051] The "chemotherapeutic agent" may also be selected from: (i) anti-hormonal drugs used to regulate or inhibit the effects of hormones on tumors, such as anti-estrogens and selective estrogen receptor modulators (SERMs), including tamoxifen (including tamoxifen citrate), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and (ii) aromatase inhibitors that inhibit aromatase, which regulates estrogen production in the adrenal glands, such as 4(5)-imidazole, aminoglutethimide, (Megestrol acetate), (exemestane; Pfizer), formestanie, fadrozole, (vorozole), (letrozole; Novartis) and (anarozole; AstraZeneca); (iii) antiandrogens, such as flutamide, nilutamide, bicalutamide, leuprolide and goserelin; and troxacitabine (1,3-dioxolane cytidine nucleoside analog); (iv) protein kinase inhibitors, such as MEK inhibitors (WO2007 / 044515); (v) lipid kinase inhibitors; (vi) antisense oligonucleotides, particularly those that inhibit genes in signal transduction pathways associated with abnormal cell proliferation, such as PKC-α, Raf and H-Ras; (vii) ribozymes, such as VEGF expression inhibitors (e.g. ) and HER2 expression inhibitors; (viii) vaccines, such as gene therapy vaccines, e.g. and rlL-2; topoisomerase I inhibitors, such as and rmRH; (ix) anti-angiogenic agents, such as bevacizumab ( Genentech); and pharmaceutically acceptable salts, acids and derivatives of any of the foregoing.

[0052] The "chemotherapeutic agent" may also be selected from therapeutic antibodies, such as alemtuzumab (Campath), bevacizumab ( Genentech); Cetuximab monoclonal antibody ( Imclone); Panmu monoclonal antibody ( Amgen), rituximab monoclonal antibody ( Genentech / Biogen Idee), Pertuzumab monoclonal antibody (OMNITARG TM ,2C4,Genentech), trostizumab monoclonal antibody ( Genentech), tosimo monoclonal antibody (Bexxar, Corixia), and antibody-drug conjugates, gemtuzumab monoclonal antibody ogamicin ( Wyeth).

[0053] The humanized monoclonal antibody with therapeutic potential as a chemotherapeutic agent, and its target compound, its stereoisomer and its pharmaceutically acceptable salt can be selected from: alemtuzumab, apolizumab, aselizumab, atlizumab, bapineuzumab, bevacizumab, bivatuzumabmertansine, cantuzumab mertansine, cedelizumab, certolizumab pegol), cidfusituzumab, cidtuzumab, daclizumab, eculizumab, efalizumab, epratuzumab, erlizumab, felvizumab, fontolizumab, gemtuzumab ozogamicin, inotuzumabozogamicin), ipilimumab, labetuzumab, lintuzumab, matuzumab, mepolizumab, motavizumab, motovizumab, natalizumab, nimotuzumab, nolovizumab, numavizumab, ocrelizumab, omalizumab, palivizumab, pascol izumab), pecfusituzumab, pectuzumab, pertuzumab, pexelizumab, ralivizumab, ranibizumab, reslivizumab, reslizumab, resyvizumab, rovelizumab, ruplizumab, sibrotuzumab, siplizumab, sontuzumab, tacatuzumab tetraxetan), tadocizumab, talizumab, tefibazumab, tocilizumab, toralizumab, trastuzumab, tucotuzumab celmoleukin, tucusituzumab, umavizumab, urtoxazumab, and visilizumab.

[0054] The present invention is further illustrated by the following specific test examples, which are used to facilitate those skilled in the art to which the present invention belongs, so that the present invention can be fully utilized and practiced without excessive interpretation. These test examples should not be regarded as limiting the scope of the present invention, but are used to illustrate how to implement the materials and methods of the present invention.

[0055] The present invention proposes a novel use of an ADAM9 inhibitor for preparing an immunomodulator. In the specification of the present invention, the compound of formula (I) is used as an example of an ADAM9 inhibitor to illustrate that the ADAM9 inhibitor has the efficacy claimed in the scope of the present invention.

[0056] 1. Chemical synthesis and structural identification of the compound of formula (I)

[0057] The derivatives having the structure shown in formula (I) of the embodiments of the present invention are designed and synthesized with a view to inhibiting the activity of ADAM9, and a molecular docking method is used to actually screen the compound structure that can fit the catalytic site of the metalloproteinase domain of ADAM9.

[0058] Please refer to Table 1 below, which shows the structural formulas of Examples 1 to 32 of the compounds of formula (I).

[0059] Table 1

[0060]

[0061]

[0062]

[0063]

[0064]

[0065]

[0066]

[0067]

[0068]

[0069] In the experiment, derivatives of compound 6X-0310 were selected from Examples 1 to 32 for testing. The selected compounds were compound 4X-0296, compound SW1, compound 2X-0295, compound SW2, compound 12W-0264 and compound 6X-0310. Compound 9R-0655, compound MS-1176, compound 5W-0369 and compound 161 that were not based on compound 6X-0310 as the core structure were also included in the experiment.

[0070] Please refer to Figure 1A , Figure 1B Table 2 is a graph showing the analysis results of the inhibition of ADAM9 activity by the compound of formula (I). Figure 1ABm7 lung cancer cells were treated with the derivatives of the aforementioned compound 6X-0310 and then cultured under non-adhesive culture conditions. The expression of ADAM9 and CDCP1 in the Bm7 lung cancer cells was detected by Western blotting, wherein EF1α was used as an internal reference, and the negative control group was Bm7 lung cancer cells without drug treatment. Figure 1A The results showed that compound 4X-0296, compound SW1, compound 2X-0295, compound SW2, compound 12W-0264 and compound 6X-0310 can all reduce the expression of CDCP1. The compounds of formula (I) of Examples 1 to 32 were further subjected to cytotoxicity assays to measure the IC of the compounds of formula (I) of the present invention in lung cancer cell lines. 50 For test results, please refer to Figure 1B and Table 2, Figure 1B 1 is a graph showing the cytotoxicity assay results of compound 9R-0655, compound 4X-0296, compound SW1, compound 2X-0295, compound SW2, compound 12W-0264, compound MS-1176 and compound 6X-0310 on Bm7 lung cancer cells. Table 2 shows the cytotoxicity assay results of Examples 1 to 32 on different tumor cell lines, where "+" indicates an effect of inhibiting tumor cell growth, and the number is IC 50 Value (unit: μM).

[0071] Table 2

[0072]

[0073]

[0074]

[0075] Figure 1B The results in Table 2 show that Examples 1 to 32 have cytotoxic effects on renal cancer cells (293), breast cancer cells (231-brm) and different lung cancer cells (A549, H1299, TC1, Bm7brmx2, Bm7brmx2-1st, Bm7brmx2-2nd and PE10 WBC), and compared with the core structure - compound 6X-0310, compound SW1, compound SW2, compound 12W-0264, compound 2X-0295 and compound 4X-0296 have stronger tumor cell cytotoxic effects, especially compound SW1 and compound SW2.

[0076] In the experiment, Autodock v.4.2 was further used to calculate the Ki (inhibition constant) of the matrix metalloproteinase (MMP) inhibitor CGS27023A (Novartis) and compound SW1 and compound SW2 to evaluate whether they are potential ADAM9 inhibitors. Please refer to Table 3 for the evaluation results.

[0077] Table 3

[0078] Compound Binding energy (kcal) Ki (inhibition constant) CGS27023A -6.64 13470nM SW1 -9.49 100nM SW2 -9.49 100nM

[0079] The results in Table 3 show that the Ki values ​​of compound SW1 and compound SW2 are lower than that of the MMP inhibitor-CGS27023A, and it is predicted that they can achieve the same inhibitory effect of ADAM9 activity as CGS27023A at a higher concentration at a lower concentration.

[0080] II. Effect of the compound of formula (I) on inhibiting ADAM9 activity

[0081] In order to measure the efficacy of the compound of formula (I) in inhibiting ADAM9 activity, an ELISA system was established using recombinant human ADAM9 and a fluorescent peptide substrate (R&D system), which included a broad-spectrum MMP inhibitor-BB-94 as a positive control group. The compounds of formula (I) used in this experiment were compound SW1 and compound SW2.

[0082] Figure 2A , Figure 2B , Figure 2C , Figure 2D , Figure 2E , Figure 2F , Figure 2G and Figure 2H , is a graph showing the characteristic analysis results of compound SW1 and compound SW2 of the examples of the present invention in in vitro tests.

[0083] Figure 2A To analyze the inhibition of ADAM9 activity with different concentrations of compound SW1 and compound SW2, Figure 2B To analyze the inhibition of ADAM17 activity with different concentrations of compound SW1 and compound SW2, Figure 2A and Figure 2B The results showed that compound SW1 and compound SW2 had similar inhibitory effects on ADAM9 activity and were dose-dependent, but compound SW1 and compound SW2 had no inhibitory effect on ADAM17.

[0084] To further evaluate the specificity of compound SW1 and compound SW2, control Bm7 lung cancer cells (treated with shGFP) and ADAM9 knockout Bm7 lung cancer cells (treated with shADAM9) were treated with 25 μM of compound SW1 or compound SW2, respectively, and then cell migration inhibition assay was performed. Figure 2C The results showed that treatment with compound SW1 or compound SW2 could significantly reduce the migration ability of control Bm7 lung cancer cells.

[0085] In order to verify that compounds SW1 and SW2 can block the anti-anoikis ability of cancer cells, the IC values ​​of compounds SW1 and SW2 on anoikis (non-adhesion-induced apoptosis) in lung cancer brain metastasis cell line Bm7brmx2, breast cancer brain metastasis cell line MDA-231brm and leukocytes under non-adhesion culture conditions were determined. 50 Please refer to Table 4 below. The results show that compound SW1 and compound SW2 can induce cell death under non-attached culture conditions and provide good therapeutic effects in the breast cancer brain metastasis cell line - MDA-231brm.

[0086] Table 4

[0087]

[0088] To further test the implantation efficiency of tumor cells treated with compound SW1 and compound SW2, control Bm7 lung cancer cells (treated with shGFP) and ADAM9 knockout Bm7 lung cancer cells (treated with shADAM9) were treated with 20μM compound SW1 or compound SW2, and wild-type TC1 lung cancer cells and ADAM9 knockout TC1 lung cancer cells were treated with 10μM compound SW1 or compound SW2, and then colony formation assay was used to confirm the implantation efficiency of each test group. Please refer to Figure 2D and Figure 2E , where the negative control group was a tumor cell line that was not treated with compound SW1 or compound SW2. Representative images of colony formation assays are shown at the bottom, and the statistical data are the mean ± SD of 6 independent wells. The results showed that treatment with compound SW1 or compound SW2 could significantly reduce the implantation efficiency of control Bm7 lung cancer cells and wild-type TC1 lung cancer cells, indicating that it could reduce the growth of control Bm7 lung cancer cells and wild-type TC1 lung cancer cells.

[0089] In the experiment, wild-type TC1 lung cancer cells and ADAM9 knockout TC1 lung cancer cells were further treated with compound SW1 or compound SW2 for 24 hours, and then stained with Annexin V and PI to detect cell apoptosis. Please refer to Figure 2F , of which Annexin V + The percentage of cells (apoptotic cells) is the statistical result of 3 independent experiments, with mean ± SD. Figure 2F The results showed that compared with ADAM9 knockout TC1 lung cancer cells, compounds SW1 and SW2 could significantly reduce the growth of control lung cancer cells and induce cell apoptosis.

[0090] In addition, the IC values ​​of compound SW1 and compound SW2 for pancreatic cancer cells were also tested. 50 Value (unit: μM), please refer to Figure 2G and the following Table 5. The results show that the compounds SW1 and SW2 of the present invention have toxic effects on different pancreatic cancer cells.

[0091] Table 5

[0092] Pancreatic cancer cells SW2 SW1 Panc-1 34.2 424 MiaPaCa 8.5 - PC-080 89.6 446 Pan18-GFP-Luc 112 0.3 Su86-86-GFP-Luc 66.8 59.2 HPAC-GFP-Luc 29.3 126.5

[0093] In the experiment, Bm7 lung cancer cells were treated with 25 μM of compound SW1 or compound SW2 for 12 hours, and the number of cancer cell spheres formed was measured. Figure 2H The negative control group consisted of Bm7 lung cancer cells that were not treated with drugs. The results showed that treatment with compound SW1 and compound SW2 could significantly reduce the number of cancer cell spheres formed (* indicates p<0.05, ** indicates p<0.01).

[0094] Effects of ADAM9 inhibitors in cancer immunotherapy

[0095] To verify the effect of ADAM9 on the immune response to tumor progression, wild-type TC1 lung cancer cells (n=10) and ADAM9-knocked-out TC1 lung cancer cells (n=10) were subcutaneously injected into C57BL / 6 mice with normal immunity to establish a subcutaneous lung tumor mouse model. The mice in the TC1-wild-type group and TC1-ADAM9-knocked-out group were sacrificed on the 22nd day after cell injection, and their tumor tissues were removed, the volume of the tumor tissue was measured, and the RNA expression levels of ADAM9, CD4, CD8 and IFN-γ in the tumor tissues of three separate tumor samples in each group were analyzed by RT-qPCR.

[0096] Please refer to Figure 3 This is a diagram of the analysis results of the effect of eliminating ADAM9 on tumor growth, which includes photos of tumor tissue size and statistical results of mice in the TC1-wild-type group and the TC1-ADAM9 elimination group. Figure 3 Results showed that ADAM9 ablation in TC1 lung cancer cells reduced tumor size and metastatic nodules in a subcutaneous lung tumor mouse model.

[0097] Please refer to Figure 4A , Figure 4B , Figure 4C and Figure 4D , is a graph showing the RNA analysis results of ADAM9, CD4, CD8, and IFN-γ in tumor tissues after ADAM9 was knocked out, where wild type represents mice in the TC1-wild type group, and knockout represents mice in the TC1-ADAM9 knockout group. The results showed that CD8 and IFN-γ RNA were overexpressed in the tumors of mice in the TC1-ADAM9 knockout group. Since neither CD8 nor IFN-γ was detected in the cultured ADAM9 knockout TC1 lung cancer cells, but overexpression of CD8 and IFN-γ was detected in the tumors of mice in the TC1-ADAM9 knockout group, the above results showed that ADAM9 gene knockout increased CD8 in tumor tissues. + The infiltration of T cells and the expression of IFN-γ can induce immune responses that inhibit TC1 tumor progression.

[0098] To further confirm whether ADAM9 gene knockout would also affect the immune response of lung metastatic tumors, wild-type TC1 lung cancer cells (n=5) and ADAM9 knockout TC1 lung cancer cells (n=5) were injected into C57BL / 6 mice with normal immunity via the tail vein to establish a lung metastasis model. The mice in the TC1-wild-type group and TC1-ADAM9 knockout group were sacrificed on the 28th day after cell injection, and their lung tissues were removed. The immune cell profile was analyzed by immunohistochemical staining of lung tissue sections, and the number of immune cells in each range of the immunohistochemical staining analysis result graph was counted.

[0099] Please refer to Figure 5A and Figure 5B , which is the analysis result diagram of eliminating the effect of ADAM9 on lung metastatic tumor growth and immune cell infiltration, Figure 5B Immunohistochemical staining of inflammatory cells in lung tissues with antibodies against T cells (CD3), macrophages (F4 / 80), and neutrophils (MPO) was performed to detect the expression of T cells, macrophages, and neutrophils in tumor tissues of the TC1-wild-type group and the TC1-ADAM9 knockout group. Figure 5A and Figure 5B The results showed that ADAM9 knockout in TC1 lung cancer cells reduced the size of lung metastatic tumors. In addition, increased CD8 + T cell infiltration was increased, while the number of neutrophils was decreased.

[0100] From the above results, it can be seen that the ADAM9 gene has an impact on tumor growth and immune cell infiltration. This experiment further conducted a comprehensive analysis of genes involved in ADAM9-mediated immunosuppression through lung tumor RNA sequencing, and found that ADAM9 in cancer cells may affect and interact with immune cells in the microenvironment, thereby contributing to the occurrence, development and metastasis of tumors. In order to more accurately identify the genes regulated by ADAM9, we identified genes whose expression profiles were consistent in the subcutaneous tumors of cancer cell lines and mice in the TC1-wild-type group and TC1-ADAM9 knockout group as ADAM9-mediated tumor genes (329 genes). Please refer to Table 6 for the functional analysis of 329 ADAM9-mediated tumor genes affected in cultured cells and tumors.

[0101] Table 6

[0102] Function Gene number -log(p-value) Regulates cytokine production 18 6.1 Positive regulation of multicellular biological processes 20 5.3 Negative regulation of multicellular biological processes 15 5.2 Response to interferon beta 5 4.8 Response to interferon-gamma 6 4.6 The connection or rolling of white blood cells 4 4.6 Negative regulation of the immune system 11 4.5 Defense response to viruses 9 4.4

[0103] From the above analysis, it can be seen that ADAM9-mediated tumor genes act on the regulation of IFN-γ and the production and response of IFN-γ cytokines. Therefore, the target cells (wild-type TC1 lung cancer cells or ADAM9-deficient TC1 lung cancer cells) and effector cells (splenocytes of C57BL / 6 mice) of C57BL / 6 mice were co-cultured at a ratio of 1:40 for 24 hours, and the cell apoptosis rate was measured to verify the above analysis results.

[0104] Please refer to Figure 6 , is a graph showing the results of apoptosis analysis after 24 hours of co-culturing target cells and effector cells, where wild type represents wild-type TC1 lung cancer cells, and ADAM9 knockout represents ADAM9 knockout TC1 lung cancer cells. The results showed that wild-type TC1 lung cancer cells were more resistant to immune cell attack than ADAM9 knockout TC1 lung cancer cells. These results indicate that ADAM9 knockout in tumors can increase the infiltration of immune cells and make them more sensitive to immune cell responses.

[0105] To further evaluate whether treatment with ADAM9 inhibitors would yield the same results as ADAM9 gene knockout, wild-type TC1 lung cancer cells were subcutaneously injected into C57BL / 6 mice with normal immunity to establish a subcutaneous lung tumor mouse model, and ADAM9 inhibitors (compound SW1, 10 mg / kg) were treated by subcutaneous injection. From day 15 to day 24, compound SW1 was injected once a day for 9 days. Tumor volume was observed on days 12, 20, and 27, respectively.

[0106] Please refer to Fig. 7A , is the analysis result of the anti-tumor effect of ADAM9 inhibitor. The control group is injected with DMSO as a control for the treatment of ADAM9 inhibitor. Fig. 7A The results showed that treatment with compound SW1 reduced tumor growth in mice bearing subcutaneous lung tumors compared with the DMSO-treated control group (N=5 per group).

[0107] Please refer to Figure 7B , is a flow chart of the use of ADAM9 inhibitors to treat isogenic orthotopic breast tumor animal models. 4 ) was transplanted into the mammary fat pad of BALB / c mice to establish a syngeneic orthotopic breast tumor animal model, which was detected using the IVIS imaging system, and the tumor size was expressed as the light flux of the IVIS image. After the tumor image was detected on the 15th day, ADAM9 inhibitors (compound SW1, 10 mg / kg) were pretreated on the 16th, 18th and 20th days by subcutaneous injection. After surgical resection of the tumor on the 21st day, ADAM9 inhibitors (compound SW1, 10 mg / kg) were continued to be treated on the 23rd, 25th, 27th, 29th and 31st days. The surgically resected syngeneic orthotopic breast tumor animal model tumors were subjected to immunohistochemical staining. In the experiment, a control group was injected with DMSO as a control for the treatment of ADAM9 inhibitors.

[0108] Please refer to Fig. 8A and Figure 8B , is the analysis result diagram of the effect of ADAM9 inhibitor on immune cell infiltration. Fig. 8A Results of an analysis of primary 4T1 breast tumors to count infiltrating T cells and neutrophils. Figure 8B Analysis of infiltrating T cells and neutrophils enumerated for lung metastatic breast tumors. Fig. 8A and Figure 8B The results showed that after 4T1 breast tumors were treated with ADAM9 inhibitors (compound SW1), the number of infiltrating T cells increased, especially in lung metastatic breast tumors. In both 4T1 primary breast tumors and lung metastatic breast tumors, the number of infiltrating neutrophils decreased significantly. This shows that ADAM9 inhibitors can increase infiltrating T cells in 4T1 breast tumors and reduce infiltrating neutrophils.

[0109] We further wanted to analyze whether ADAM9 inhibitors and checkpoint inhibitors have synergistic effects. In the experiment, TC1 subcutaneous lung tumor mice were treated with ADAM9 inhibitors (compound SW1), checkpoint inhibitors (anti-PDL1 antibodies) alone or co-treated with ADAM9 inhibitors (compound SW1) and checkpoint inhibitors (anti-PDL1 antibodies), and the expression of CD8 + T cells and regulatory T cells (T RegThe experiment also included a control group treated with vehicle (DMSO).

[0110] Please refer to Fig. 9 , is a graph showing the analysis results of co-treatment with ADAM9 inhibitors and checkpoint inhibitors. The results showed that compared with the groups treated with vehicle, compound SW1 alone, and anti-PDL1 antibody, CD8 + T cells and regulatory T cells (T Reg ) was significantly increased, indicating that ADAM9 inhibitors and checkpoint inhibitors have synergistic effects.

[0111] We further analyzed whether treatment with ADAM9 inhibitors would stimulate tumor cells to secrete cytokines to activate the anti-tumor immunity of ADAM9 inhibitors. In the experiment, human colon cancer cells SW620 and human pancreatic cancer cells PANC-1 were treated with 12.5μM of compound SW1, and human esophageal cancer cells CE146T were treated with 5μM of compound SW1. After 24 hours, the cells were collected and RT-qPCR was used to detect the RNA expression of CXCL10, CXCL11, CXC3CL1 and IFNB. Among them, CXCL10, CXCL11 and CXCL1 are CD8 + IFNB is a cytokine of T cell immune response and has anti-cancer effects.

[0112] Please refer to Fig. 10A , Fig. 10B and Fig. 10C , which is the analysis result of ADAM9 inhibitor stimulating tumor cells to secrete cytokines. FIG. 10A to FIG. 10C The results showed that treatment with compound SW1 increased the expression of CXCL10, CXCL11, CXC3CL1 and IFNB in ​​human colon cancer cells, human pancreatic cancer cells and human esophageal cancer cells, indicating that ADAM9 inhibitors can stimulate cancer cells to secrete cytokines related to immune activation.

[0113] In summary, the present invention proposes a new use of an ADAM9 inhibitor, which can be used as an immunomodulator to enhance the effectiveness of cancer immunotherapy. It has been confirmed by experimental data that it can reduce tumor growth, change the microenvironment of tumors, stimulate immune cells to infiltrate tumors, and stimulate cancer cells to secrete cytokines related to immune activation, and can be used to treat tumors with resistance to immunotherapy. ADAM9 inhibitors can be used for cancer immunotherapy and can treat cancer, and they can have a synergistic effect when co-formulated with test inhibitors, and can also increase the effectiveness of cancer immunotherapy, and have the potential to be used in the biomedical health care market.

[0114] Although the present invention has been disclosed in the above embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention shall be determined by the scope of the attached patent application.

Claims

1. Use of an ADAM9 inhibitor in the preparation of a drug for preparing an immunomodulator, characterized in that: The ADAM9 inhibitor is selected from: and 2. The use according to claim 1, characterized in that: The immunomodulator is used to enhance the effectiveness of a cancer immunotherapy.

3. The use according to claim 2, characterized in that: The cancer immunotherapy is administered to an immunocompromised patient.

4. The use according to claim 2, characterized in that: The immunomodulator is used to stimulate an immune cell to infiltrate a tumor.

5. The use according to claim 4, characterized in that: The immune cell is a T cell, a natural killer cell, a macrophage, a neutrophil, a dendritic cell or a myeloid-derived suppressor cell.

6. The use according to claim 2, characterized in that: The immunomodulator is used to alter the chemokine species of the tumor microenvironment.

7. The use according to claim 2, characterized in that: The immunomodulator is used to treat a tumor that is resistant to immunotherapy.

8. The use according to claim 7, characterized in that: The immunotherapy-resistant tumor is resistant to a checkpoint inhibitor, an adoptive cell transfer, a therapeutic antibody, a therapeutic vaccine, a cytokine, an immune cell therapy, or a combination thereof.

9. The use according to claim 8, characterized in that: The checkpoint inhibitor is an anti-CTLA-4 antibody, an anti-PDL1 antibody or an anti-PD-1 antibody.

10. The use according to claim 1, characterized in that: The immunomodulator is co-formulated with a checkpoint inhibitor.

11. The use according to claim 10, characterized in that The checkpoint inhibitor is an anti-CTLA-4 antibody, an anti-PDL1 antibody or an anti-PD-1 antibody.

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