Combination therapy with a CC chemokine receptor 4 (CCR4) antagonist and one or more checkpoint inhibitors

Abstract

The present invention relates to combination therapies of C-C chemokine receptor 4 (CCR4) antagonists and one or more immune checkpoint inhibitors in the treatment of cancer.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority under 35 U.S.C. §119(e) to U.S. Provisional Applications Serial No. 62 / 724,412, filed on August 29, 2018, and Serial No. 62 / 771,853, filed on November 27, 2018, the disclosures of each of which are incorporated herein by reference in their entirety.

[0003] STATEMENT REGARDING RIGHTS TO INventions Made Under Federally Sponsored Research and Development

[0004] not applicable.

[0005] Reference to a Sequence Listing, Table, or Computer Program Listing Appendix Submitted on CD-ROM

[0006] not applicable. Background Art

[0007] Cancer is a class of diseases in which cells exhibit dysregulated replication and growth. Recent cancer models have implicated the immune system, including cellular homing and immune checkpoints, in the development and progression of cancer. While significant progress has been made in understanding the biological basis of cancer, the disease remains a leading cause of death.

[0008] CC chemokine receptor 4 (CCR(4)), first identified by Power et al. (Power et al. (1995) J. Biol. Chem. 270:19495-19500), is a G protein-coupled receptor that binds to a group of chemokines, including CCL22, also known as macrophage-derived chemokine (MDC; CC chemokine is reported to be a chemoattractant for the Th2 subset of peripheral blood T cells, dendritic cells, and natural killer (NK) cells), and CCL17, also known as TARC (thymus and activation-regulated chemokine), which is also produced by monocytes and dendritic cells.

[0009] CCR (4) is involved in immune regulatory processes such as cell homing to specific tissues, including homing of T lymphocytes to the skin and lungs (see, for example, Campbell et al. (1999) Nature 400:776-780, Gonzalo et al. (1999) J. Immunol 163:403-5411, Lloyd et al. (2000) J. Exp. Med. 191:265-273, Kawasaki et al. (2001) J. Immunol 166:2055-2062). Modulators of CCR4 activity have been described, for example, in WO 2013 / 082490.

[0010] Cytotoxic T-lymphocyte antigen-4 (CTLA-4) is considered a key regulator of adaptive immune responses. In particular, CTLA-4 is understood to play a central role in maintaining and maintaining the components of acute T-cell responses. Therefore, CTLA-4, as an immune checkpoint inhibitor, is considered a potential therapeutic target for the treatment of cancer and inflammation. CTLA-4 modulators have been described, for example, in WO2018 / 035710.

[0011] Programmed death-1 (PD-1) is a transmembrane receptor protein that negatively regulates T cell function through interaction with its two natural ligands, PD-L1 and PD-L2. Like CTLA4, PD-1 is a major regulator of the immune system and is also considered an immune checkpoint inhibitor. PD-1 / PD-L1 modulators have been described, for example, in WO2018 / 005374.

[0012] Given the roles of cell homing and immune checkpoint pathways in cancer development and progression, there is a need to develop combination therapies that can improve cancer treatment. Summary of the Invention

[0013] The present invention relates to combination therapies of CC chemokine receptor 4 (CCR4) antagonists and one or more checkpoint inhibitors in the treatment of cancer.

[0014] In some embodiments, the CCR4 receptor antagonist has the following formula I

[0015]

[0016] Each variable is described below.

[0017] In some embodiments, the CCR4 antagonist has the formula

[0018]

[0019] or a pharmaceutically acceptable salt thereof.

[0020] In some embodiments, the CCR4 antagonist has the formula

[0021]

[0022] or a pharmaceutically acceptable salt thereof.

[0023] In some embodiments, the CCR4 antagonist has the formula

[0024]

[0025] or a pharmaceutically acceptable salt thereof.

[0026] In some embodiments, the CCR4 antagonist has the formula

[0027]

[0028] or a pharmaceutically acceptable salt thereof.

[0029] BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 The study design used for the KCM orthotopic pancreatic tumor model is described.

[0031] Figure 2A -C shows the primary tumor + pancreas weight (Group A), total tumor + pancreas weight (Group B), and spleen weight (Group C) of each test group. Combination treatment with anti-CTLA-4 antibody and Compound 1 reduced tumor weight compared to antibody or Compound 1 alone.

[0032] Figure 3A Panels A to C report the relative number of suppressive immune cell populations in each test group. Panels A to C report cells per gram of tissue, while panels D to F report the percentage of CD45+ cells. Panels A and D report T-reg cells; panels B and E report G-MDSC cells; and panels C and F report M-MDSC cells.

[0033] Figure 4 The study design used for the CT26 colon cancer tumor model is described.

[0034] Figure 5 The survival rate of mice from each test group in the CT26 tumor model during the study period is plotted.

[0035] Figure 6 The mean tumor volume of mice from each test group is plotted.

[0036] Figure 7A-D plots the individual tumor volumes of mice from each test group. Panel A reports the antibody isotype and vehicle group; Panel B reports the antibody isotype and compound 1 group; Panel C reports the anti-CTLA-4 antibody and vehicle group; Panel D reports the anti-CTLA-4 antibody and compound 1 group.

[0037] Figure 8A -F plots the effect of re-challenged and naive mice with CT26 colon cancer cells and 4T1 breast cancer cells. The tumor volume of mice was measured. Re-challenged mice were those that had complete tumor regression and had been previously treated with anti-CTLA-4 (6 mice) or anti-CTLA-4 and compound 1 (8 mice). Blank mice were mice that had been previously challenged with CT26 colon cancer cells. Panel A showed that 5 / 6 mice from the anti-CTLA-4 treatment group were resistant to reintroduction of CT26 tumors; Panel B showed that none of the mice from the anti-CTLA-4 treatment group were resistant to 4T1 tumor cells; Panel C showed that 8 / 8 mice from the anti-CTLA-4 / compound 1 treatment group were resistant to reintroduction of CT26 tumors; Panel D showed that 2 of the 8 mice from the anti-CTLA-4 / compound 1 treatment group were resistant to 4T1 tumor cells; Panel E showed that all blank mice had tumor growth when first challenged with CT26 cells; Panel F showed that no blank mice were resistant to 4T1 tumor cells.

[0038] Figure 9A -C plots flow cytometry results of blood samples obtained from mice tested in Figure 8. Panel A shows that mice previously exposed to CT26 cells (and treated with CTLA-4 or anti-CTLA-4 and compound) had cytotoxic T cells against CT26 tumor cells before rechallenge, whereas naive mice did not. Panels B and C show the number of cytotoxic T cells responding to CT26 tumor cells before and one week after rechallenge. DETAILED DESCRIPTION

[0039] I.General

[0040] The present invention relates to the surprising and unexpected discovery that combination therapy using a CCR4 antagonist and one or more immune checkpoint inhibitors significantly improves cancer treatment compared to one or more checkpoint inhibitors by themselves.

[0041] II. Abbreviations and Definitions

[0042] Unless otherwise indicated, the term "alkyl", by itself or as part of another substituent, refers to a radical having the specified number of carbon atoms (i.e., C 1-8refers to a straight or branched chain hydrocarbon group of 1 to 8 carbons. Examples of alkyl groups include: methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, and the like. The term "alkenyl" refers to an unsaturated alkyl group having one or more double bonds. Similarly, the term "alkynyl" refers to an unsaturated alkyl group having one or more triple bonds. Examples of such unsaturated alkyl groups include: vinyl, 2-propenyl, crotyl, 2-isopentenyl, 1,2-(butadienyl), 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl, 3-butynyl, and higher homologs and isomers. The term "cycloalkyl" refers to a cycloalkyl group having a specified number of ring atoms (e.g., C 3-6 "Cycloalkyl" refers to a hydrocarbon ring that is fully saturated or has no more than one double bond between ring vertices. "Cycloalkyl" may also refer to bicyclic and polycyclic hydrocarbon rings, for example, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, etc. The term "heterocycloalkyl" refers to a cycloalkyl group containing one to five heteroatoms selected from N, O, and S, wherein the nitrogen and sulfur atoms are optionally oxidized and the nitrogen atom is optionally quaternized. The heterocycloalkyl group may be a monocyclic, bicyclic, or polycyclic ring system. The non-limiting example of heterocycloalkyl comprises: pyrrolidine, imidazolidine, pyrazolidine, butyrolactam, valerolactam, imidazolinone, hydantoin, dioxolane, phthalimide, piperidine, 1,4-dioxane, morpholine, thiomorpholine, thiomorpholine-S-oxide, thiomorpholine-S, S-oxide, piperazine, pyrans, pyridone, 3-pyrroline, thiopyrans, pyrone, tetrahydrofuran, tetrahydrothiophene, quinuclidine, and similar groups.Heterocycloalkyl can be connected to other parts of molecule by ring carbon or heteroatom.For such as cycloalkylalkyl and heterocycloalkylalkyl, refer to that cycloalkyl or heterocycloalkyl are connected to the rest of molecule by alkyl or alkylene linking group.For example, cyclobutylmethyl-is the cyclobutyl ring of the methylene linking group connected to the rest of molecule.

[0043] The term "alkylene" by itself or as part of another substituent refers to a divalent group derived from an alkane, as shown in -CH2CH2CH2CH2-. Typically, an alkyl (or alkylene) group will have from 1 to 24 carbon atoms, with those groups having 10 or fewer carbon atoms being preferred in the present disclosure. A "lower alkyl" or "lower alkylene" group is a short-chain alkyl or alkylene group typically having four or fewer carbon atoms. Similarly, "alkenylene" and "alkynylene" refer to unsaturated forms of "alkylene" having double or triple bonds, respectively.

[0044] Unless otherwise indicated, the term "heteroalkyl" by itself or in combination with another term refers to a stable straight or branched chain or cyclic hydrocarbon group, or a combination thereof, consisting of the stated number of carbon atoms and one to three heteroatoms selected from O, N, Si and S, and wherein the nitrogen and sulfur atoms may be optionally oxidized, and the nitrogen heteroatom may be optionally quaternized. The heteroatoms O, N and S may be located at any position within the heteroalkyl group. The heteroatom Si may be located at any position within the heteroalkyl group, including the position at which the alkyl group is attached to the rest of the molecule. Examples include: -CH2-CH2-O-CH3, -CH2-CH2-NH-CH3, -CH2-CH2-N(CH3)-CH3, -CH2-S-CH2-CH3, -CH2-CH2, -S(O)-CH3, -CH2-CH2-S(O)2-CH3, -CH=CH-O-CH3, -Si(CH3)3, -CH2-CH=N-OCH3, and -CH=CH-N(CH3)-CH3. Up to two heteroatoms may be consecutive, for example -CH2-NH-OCH3 and -CH2-O-Si(CH3)3. Similarly, unless otherwise indicated, the terms "heteroalkenyl" and "heteroalkynyl" by themselves or in combination with another term refer to an alkenyl or alkynyl group, respectively, containing the stated number of carbon atoms and having from one to three heteroatoms selected from O, N, Si, and S, and wherein the nitrogen and sulfur atoms may optionally be oxidized, and the nitrogen heteroatom may optionally be quaternized. The heteroatoms O, N, and S may be located at any position within the heteroalkyl group.

[0045] The term "heteroalkylene," by itself or as part of another substituent, refers to a saturated or unsaturated or polyunsaturated divalent radical derived from heteroalkyl, for example, -CH2-CH2-S-CH2CH2- and -CH2-S-CH2-CH2-NH-CH2-, -O-CH2-CH=CH-, -CH2-CH=C(H)CH2-O-CH2-, and -S-CH2-C≡C-. For heteroalkylene, heteroatoms can also occupy one or both of the chain termini (e.g., alkyleneoxy, alkylenedioxy, alkyleneamino, alkylenediamino, etc.).

[0046] The terms "alkoxy," "alkylamino," and "alkylthio" (or thioalkoxy) are used in their conventional sense to refer to those alkyl groups attached to the rest of the molecule through an oxygen atom, an amino group, or a sulfur atom, respectively. Additionally, for dialkylamino groups, the alkyl moieties may be the same or different and may be combined with the nitrogen atom to which they are attached to form a 3-7 membered ring. Thus, with -NR a R b Representative groups include piperidinyl, pyrrolidinyl, morpholinyl, azetidinyl, and the like.

[0047] The terms "halo" or "halogen," by themselves or as part of another substituent, mean, unless otherwise stated, a fluorine, chlorine, bromine, or iodine atom. Additionally, terms such as "haloalkyl" are intended to include monohaloalkyl and polyhaloalkyl. For example, the term "C 1-4 "Haloalkyl" is intended to include trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.

[0048] Unless otherwise indicated, the term "aryl" refers to a polyunsaturated, usually aromatic, hydrocarbon radical which may be a monocyclic ring or polycyclic rings (up to three rings) fused together or covalently linked. The term "heteroaryl" refers to an aryl group (or ring) containing from one to five heteroatoms selected from N, O, and S, wherein the nitrogen and sulfur atoms are optionally oxidized and the nitrogen atom is optionally quaternized. A heteroaryl group may be attached to the rest of the molecule via a heteroatom. Non-limiting examples of aryl groups include: phenyl, naphthyl, and biphenyl; and non-limiting examples of heteroaryl groups include: pyridyl, pyridazinyl, pyrazinyl, pyrimidinyl, triazinyl, quinolinyl, quinoxalinyl, quinazolinyl, cinnolinyl, phthalaziniyl, benzotriazinyl, purinyl, benzimidazolyl, benzopyrazolyl, benzotriazolyl, benzisoxazolyl, isobenzofuranyl, isoindolyl, indolinazinyl ( and the like. Substituents for each of the above-recited aryl and heteroaryl ring systems are selected from the group consisting of acceptable substituents described hereinafter.

[0049] For the sake of brevity, when the term "aryl" is used in combination with other terms (e.g., aryloxy, arylthioxy, arylalkyl), aryl includes both aryl and heteroaryl rings as defined above. Thus, the term "arylalkyl" is intended to include those groups in which an aromatic group is attached to an alkyl group that is attached to the rest of the molecule (e.g., benzyl, phenethyl, pyridylmethyl, etc.).

[0050] In some embodiments, the above terms (e.g., "alkyl," "aryl," and "heteroaryl") will include both substituted and unsubstituted forms of the indicated groups. Preferred substituents for various types of groups are provided below. For simplicity, the terms aryl and heteroaryl will refer to substituted or unsubstituted forms as provided below, while the term "alkyl" and related aliphatic groups refer to the unsubstituted form unless indicated as substituted.

[0051] Substituents for alkyl groups (including those groups commonly referred to as alkylene, alkenyl, alkynyl, and cycloalkyl) can be various groups selected from the group consisting of: -halogen, -OR', -NR'R", -SR', -SiR'R"R"', -OC(O)R', -C(O)R', -C02R', -CONR'R", -OC(O)NR'R", -NR"C(O)R', -NR'-C(O)NR"R"', -NR" C(O)2R', -NH-C(NH2)=NH, -NR'C(NH2)=NH, -NH-C(NH2)=NR', -S(O)R', -S(O)2R', -S(O)2NR'R", -NR'S(O)2R", -CN and -NO2, in numbers from 0 to (2m'+1), where m' is the total number of carbon atoms in these groups. R', R" and R'" each independently refer to hydrogen, unsubstituted C 1-8 Alkyl, unsubstituted heteroalkyl, unsubstituted aryl, aryl substituted with 1-3 halogens, unsubstituted C 1-8 Alkyl, C 1-8 Alkoxy or C 1-8 Thioalkoxy, or unsubstituted aryl-C 1-4 Alkyl. When R' and R" are attached to the same nitrogen atom, they may be combined with the nitrogen atom to form a 3-, 4-, 5-, 6-, or 7-membered ring. For example, -NR'R" is intended to include: 1-pyrrolidinyl and 4-morpholinyl. The term "acyl" (used alone or as part of another group) refers to an alkyl group in which both substituents on the carbon closest to the point of attachment of the group are replaced with a substituent =0 (e.g., C(O)CH3, -C(O)CH2CH2OR', etc.).

[0052] Similarly, substituents for aryl and heteroaryl groups vary and are typically selected from: -halogen, -OR', -OC(O)R', -NR'R", -SR', -R', -CN, -NO2, -CO2R', -CONR'R", -C(O)R', -OC(O)NR'R", -NR"C(O)R', -NR"C(O)2R', -NR'-C(O)NR"R"', -NH-C (NH2) = NH, -NR'C(NH2) = NH, -NH-C(NH2) = NR', -S(O)R', -S(O)2R', -S(O)2NR'R", -NR'S(O)2R", -N3, perfluoro(C1-C4)alkoxy and perfluoro(C1-C4)alkyl, in a number ranging from zero to the total number of open valences on the aromatic ring system; and R', R" and R'"' are independently selected from: hydrogen, C 1-8 Alkyl, C 3-6 Cycloalkyl, C 2-8 Alkenyl, C 2-8Alkynyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl)-C 1-4 Alkyl and unsubstituted aryloxy-C 1-4 Other suitable substituents include the above-mentioned aryl substituents attached to the ring atoms via an alkylene chain of 1 to 4 carbon atoms.

[0053] Two of the substituents on adjacent atoms of an aryl or heteroaryl ring may optionally be replaced by a substituent of the formula -TC(O)-(CH2) q -U- substituent, wherein T and U are independently -NH-, -O-, -CH2- or a single bond, and q is an integer from 0 to 2. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may be optionally replaced by a -U- substituent of the formula -A-(CH2) r wherein A and B are independently -CH2-, -O-, -NH-, -S-, -S(O)-, -S(O)2-, -S(O)2NR'-, or a single bond, and r is an integer from 1 to 3. One of the single bonds of the new ring thus formed may optionally be replaced by a double bond. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may optionally be replaced by a -(CH2)-substituted aryl or heteroaryl ring. s -X-(CH2) t -substituted by a substituent, wherein s and t are independently integers from 0 to 3, and X is -O-, -NR'-, -S-, -S(O)-, -S(O)2- or -S(O)2NR'-. The substituent R' in NR'- and -S(O)2NR'- is selected from hydrogen or unsubstituted C 1-6 alkyl.

[0054] As used herein, the term "heteroatom" is intended to include oxygen (O), nitrogen (N), sulfur (S), and silicon (Si).

[0055] For the compounds provided herein, a bond drawn from a substituent (typically an R group) to the center of an aromatic ring (e.g., benzene, pyridine, etc.) will be understood to mean a bond providing attachment at any available vertex of the aromatic ring. In some embodiments, the description will also include attachment to a ring fused to the aromatic ring. For example, a bond drawn to the center of the benzene portion of an indole will represent a bond to any available vertex of the six-membered or five-membered ring portion of the indole.

[0056] The term "pharmaceutically acceptable salts" is intended to include salts of the active compounds prepared with relatively nontoxic acids or bases, depending on the specific substituents present on the compounds described herein. When the compounds of the present invention contain relatively acidic functional groups, base addition salts can be obtained by contacting the neutral form of these compounds with a sufficient amount of the desired base (either neat or in a suitable inert solvent). Examples of salts derived from pharmaceutically acceptable inorganic bases include aluminum salts, ammonium salts, calcium salts, copper salts, iron salts, ferrous salts, lithium salts, magnesium salts, manganic salts, manganous salts, potassium salts, sodium salts, zinc salts, and the like. Salts derived from pharmaceutically acceptable organic bases include salts of primary, secondary, and tertiary amines, including substituted amines, cyclic amines, naturally occurring amines, and the like, such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucosamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, and the like. When the compounds of the present invention contain relatively basic functional groups, acid addition salts can be obtained by contacting the neutral forms of these compounds with a sufficient amount of the desired acid (either neat or in a suitable inert solvent). Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, monohydrogencarbonic acid, phosphoric acid, monohydrogenphosphoric acid, dihydrogenphosphoric acid, sulfuric acid, monohydrogensulfuric acid, hydroiodic acid or phosphorous acid, and salts derived from relatively non-toxic organic acids such as acetic acid, propionic acid, isobutyric acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid, etc. Also included are salts of amino acids such as arginine, and organic acids such as glucuronic acid or galacturonic acid (see, for example, Berge, SM et al., "Pharmaceutical Salts", Journal of Pharmaceutical Science, 1977, 66, 1-19). Certain specific compounds of the present invention contain both basic and acidic functional groups, allowing the compounds to be converted into base or acid addition salts.

[0057] The neutral forms of the compounds can be regenerated by contacting the salt with a base or acid and isolating the parent compound in the conventional manner. The parent form of the compound differs from its various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of this invention.

[0058] In addition to salt forms, the present invention provides compounds that exist in prodrug form. Prodrugs of the compounds described herein are those compounds that readily undergo chemical changes under physiological conditions to provide the compounds of the present invention. Additionally, prodrugs can be converted into the compounds of the present invention by chemical or biochemical methods in an ex vivo environment. For example, when the prodrug is placed in a transdermal patch reservoir with a suitable enzyme or chemical reagent, the prodrug will slowly convert into the compounds of the present invention.

[0059] Certain compounds of the present invention may exist in unsolvated forms as well as solvated forms, including hydrated forms. Typically, solvated forms are equivalent to unsolvated forms and are intended to be within the scope of the present invention. Certain compounds of the present invention may exist in polycrystalline or amorphous forms. Typically, all physical forms are equivalent for the purposes contemplated by the present invention and are intended to be within the scope of the present invention.

[0060] Certain compounds of the present invention have asymmetric carbon atoms (optical centers) or double bonds; racemates, diastereomers, geometric isomers, regioisomers and individual isomers (e.g., isolated enantiomers) are all intended to be included within the scope of the present invention. When compounds provided herein have defined stereochemistry (represented by R or S, or indicated by a dotted line or wedge-shaped bond), those skilled in the art will understand that those compounds are substantially free of other isomers (e.g., at least 80%, 90%, 95%, 98%, 99%, and up to 100% free of other isomers).

[0061] The compounds of the present invention may also contain unnatural proportions of atomic isotopes at one or more of the atoms comprising these compounds. An unnatural proportion of an isotope can be defined as the amount from that found in nature to 100% being composed of the atom in question. For example, the compounds may incorporate radioactive isotopes, such as tritium ( 3 H), iodine-125 ( 125 I) or carbon-14 ( 14 C) or non-radioactive isotopes such as deuterium ( 2 H) or carbon-13 ( 13 C). Such isotopic variations may provide additional utility to those described elsewhere in this application. For example, isotopic variants of the compounds of the present invention may find other uses, including but not limited to, as diagnostic and / or imaging agents, or as cytotoxic / radiotoxic therapeutic agents. In addition, isotopic variants of the compounds of the present invention may have altered pharmacokinetic and pharmacodynamic characteristics, which may help improve safety, tolerability, or efficacy during treatment. All isotopic variants of the compounds of the present invention, whether radioactive or not, are intended to be encompassed within the scope of this invention.

[0062] As used herein, the term "selective CCR4 antagonist" refers to a highly selective compound that inhibits CCR4 activity with little or no cross-reactivity to non-target proteins (e.g., CCR1, CCR2, CCR3, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, CCR12, CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6 and / or CXCR7). In some embodiments, a "selective CCR4 antagonist" has an IC50 value that is at least 10; 100; 500; 1,000; 2,000; 5,000; or more times lower than the IC50 value of a protein such as CCR1, CCR2, CCR3, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, CCR12, CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6 and / or CXCR7 when tested in the assay used in Example 3 of the present application. In some embodiments, a "selective CCR4 antagonist" does not inhibit CCR1, CCR2, CCR3, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, CCR12, CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6, and / or CXCR7 at a concentration of 1 μM or less in the assay used in Example 2 of this application. These proteins are considered "not inhibited" when they maintain 100%, 99%, 95%, 90%, or 85% of their activity under reference conditions using the selective CCR4 antagonist.

[0063] III. Combination therapy with a CCR4 antagonist and one or more immune checkpoint inhibitors

[0064] Provided herein are methods, compositions, and kits for treating cancer using the synergistic effects of CCR4 antagonists and immune checkpoint inhibitors. Combination therapy including a CCR4 antagonist and one or more immune checkpoint inhibitors is more effective in treating cancer than using each alone.

[0065] Cancer generally includes any of a variety of malignant tumors characterized by the proliferation of undifferentiated cells that tend to invade surrounding tissues and metastasize to new body sites. Non-limiting examples of different types of cancer that may be treated using the compositions of the present disclosure include ovarian cancer, breast cancer, lung cancer (e.g., non-small cell lung cancer), bladder cancer, thyroid cancer, liver cancer, pleural cancer, pancreatic cancer, cervical cancer, prostate cancer, testicular cancer, colon cancer, anal cancer, colorectal cancer, bile duct cancer, gastrointestinal carcinoid tumors, esophageal cancer, gallbladder cancer, rectal cancer, appendix cancer, small intestine cancer, stomach (gastric) cancer, kidney cancer (i.e., renal cell carcinoma), central nervous system cancer, skin cancer, choriocarcinoma, head and neck cancer, bone cancer, osteosarcoma, fibrosarcoma, neuroblastoma, glioma, melanoma, leukemia (e.g., acute lymphocytic leukemia, chronic lymphocytic leukemia, acute myeloid leukemia, chronic myeloid leukemia, or hairy cell leukemia), lymphoma (e.g., non-Hodgkin's lymphoma, Hodgkin's lymphoma, B-cell lymphoma, or Burkitt's lymphoma), lymphoma) and multiple myeloma.

[0066] In some embodiments, the cancer is lung cancer (such as non-small cell lung cancer), melanoma, epithelial cancer (such as prostate cancer, ovarian cancer, breast cancer), or blood cancer (such as leukemia, lymphoma, multiple myeloma).

[0067] In some embodiments, the cancer is a solid cancer. In some embodiments, the cancer is pancreatic cancer. In some embodiments, the cancer is colon cancer.

[0068] A. CCR4 antagonists,

[0069] CCR4 antagonists are compounds that reduce or inhibit CCR4 activity. Many such compounds are known in the art. In some embodiments, the CCR4 antagonists of the present invention are selective CCR4 antagonists.

[0070] In some embodiments, the CCR4 antagonist is a small molecule inhibitor of CCR4 having the following formula (I):

[0071]

[0072] and pharmaceutically acceptable salts thereof, wherein

[0073] R 1 Selected from: hydrogen, C 1-8 Alkyl, C 1-8 Halogenated alkyl, C 1-8 Hydroxyalkyl, C 3-8 Cycloalkyl, halogen, -CN, -SO2Me and -C(O)NH2;

[0074] Each R 2Selected from: C 1-8 Alkyl, C 1-8 Haloalkyl, halogen, -CN and C 1-8 Alkoxy; or two R attached to adjacent carbon atoms 2 The groups are optionally linked to form a 5- or 6-membered ring (aliphatic or aromatic, cycloalkyl or heterocycloalkyl);

[0075] R 3 Selected from: hydrogen, methyl and C 1-4 alkyl halide;

[0076] R 4 Selected from: hydrogen, C 1-8 Alkyl, C 1-8 Haloalkyl and C 1-8 hydroxyalkyl;

[0077] Each subscript n is independently an integer from 0 to 3;

[0078] B is a bond or C(O);

[0079] Q is selected from the group consisting of: C, CH, N, O, S, S(O), and SO2:

[0080] W, X, Y and Z are independently selected from: C, CH and N, but Q and W are not both N;

[0081] R 5 and R 6 Not present or independently selected from: H, -OH, C 1-8 Alkyl, C 1-8 Hydroxyalkyl, C 1-4 Alkoxy-C 1-4 Alkyl, -C(O)NR a R b 、C 1-8 Alkylene-C(O)NR a R b 、-NH-C 1-4 Alkylene-C(O)NR a R b 、-C(O)-C 1-4 Alkylene-NR a R b , -CO2H and acid isosteres, C 1-8 Alkylene -CO2H and acid isosteres, -N(R a )C(O)NR a R b 、C 1-8 Alkylene-N(R a )C(O)NR a R b 、-NR a R b、C 1-8 Alkylene-NR a R b 、C 1-8 Alkoxy, -C(O)OR a 、C 1-8 Alkylene-C(O)OR a 、-CN、-C(O)R a 、-SO2R a and -N(R a )C(O)R b ;

[0082] in,

[0083] Each R a and R b Independently selected from: hydrogen, C 1-8 Alkyl, C 1-8 Hydroxyalkyl, C 1-8 Haloalkyl and C 1-8 alkoxy; and

[0084] R 7 Not present or selected from: hydrogen, C 1-8 Alkyl and C 1-8 Halogenated alkyl.

[0085] In one group of embodiments, the compounds provided herein are those wherein X and Y are not both N. In another group of embodiments, R 3 is H, and each R 2 Independently selected from: C 1-8 Alkyl, C 1-8 Haloalkyl, halogen, and -CN.

[0086] In another group of embodiments, the compounds provided herein have the following Formula (Ia):

[0087]

[0088] Among them, each R 1 、R 2 、R 4 、R 5 、R 6 、R 7 , X, Y, Z, W, Q, B and subscript n are as described for Formula I. In selected embodiments, X is C or CH.

[0089] In another group of embodiments, the compounds provided herein have the following Formula (Ib):

[0090]

[0091] Among them, each R 2Selected from: C 1-8 Alkyl, C 1-8 Haloalkyl, halogen, and –CN, as well as each R 1 、R 4 、R 5 、R 6 、R 7 , X, Z, W, Q, B and subscript n are as described for Formula I.

[0092] In another group of embodiments, the compounds provided herein have the following Formula (Ic):

[0093]

[0094] Among them, each R 2 Selected from: C 1-8 Alkyl, C 1-8 haloalkyl, halogen, and –CN; subscript n is 0 or 1; and each R 1 、R 4 、R 6 、R 7 , X, Y, Z and Q are as described for Formula I. In selected embodiments, n is 1, and R 4 is hydrogen or methyl.

[0095] In another group of embodiments, the compounds provided herein have the following Formula (Id):

[0096]

[0097] Among them, each R 2 is a member selected from the group consisting of: C 1-8 Alkyl, C 1-8 haloalkyl, halogen, and –CN; subscript n is 0 or 1; and each R 1 、R 4 、R 6 、R 7 , X, Z and Q are as described in Formula I. In selected embodiments, n is 1, and R 4 is hydrogen or methyl.

[0098] In another group of embodiments, the compounds provided herein have the following Formula (Ie):

[0099]

[0100] Among them, each R 2 Selected from: C 1-8 Alkyl, C 1-8 haloalkyl, halogen, and –CN; subscript n is 0 or 1; and each R 1 、R 4 、R 6 、R7 , Y, Z and Q are as described for Formula I. In selected embodiments, n is 1, and R 4 is hydrogen or methyl.

[0101] In another group of embodiments, the compounds provided herein have the following formula (If):

[0102]

[0103] Among them, each R 2 Selected from: C 1-8 Alkyl, C 1-8 haloalkyl, halogen, and –CN; and each R 1 、R 4 、R 6 、R 7 and Q are as described for Formula I. In selected embodiments, R 4 is hydrogen or methyl.

[0104] In yet other embodiments, compounds provided herein have the following formulae (I), (Ia), and (Ib), including the specific embodiments provided above, wherein B is C(O). Still further, compounds are provided wherein the ring having Z as a ring vertex is selected from pyrrolidine and piperidine. In selected embodiments, compounds are provided wherein the ring having Z as a ring vertex is selected from pyrrolidin-2-yl and piperidin-2-yl, and R 5 、R 6 and R 7 At least one of them is not hydrogen.

[0105] In yet other embodiments, provided herein are compounds having the following formulae (I), (Ia) and (Ib), including the specific embodiments provided above, wherein B is a bond. In a related embodiment, B is a bond and the ring having Z as a ring vertex is selected from pyrrolidine, piperidine and cyclohexane. In a specific embodiment, B is a bond and the ring having Z as a ring vertex is selected from pyrrolidin-1-yl, pyrrolidin-2-yl, piperidin-1-yl, piperidin-2-yl, piperidin-3-yl and cyclohexane. In yet other embodiments, B is a bond and the ring having Z as a ring vertex is selected from the group consisting of pyrrolidin-1-yl, pyrrolidin-2-yl, piperidin-1-yl, piperidin-2-yl, piperidin-3-yl and cyclohexane; and R 5 、R 6 and R 7 At least one of them is not hydrogen.

[0106] In one group of embodiments, Z is CH or N.

[0107] In some embodiments, the CCR4 antagonist has the formula

[0108]

[0109] or a pharmaceutically acceptable salt thereof.

[0110] In some embodiments, the CCR4 antagonist has the formula

[0111]

[0112] or a pharmaceutically acceptable salt thereof.

[0113] In some embodiments, the CCR4 antagonist has the formula

[0114]

[0115] or a pharmaceutically acceptable salt thereof.

[0116] In some embodiments, the CCR4 antagonist has the formula

[0117]

[0118] or a pharmaceutically acceptable salt thereof.

[0119] In some embodiments, the CCR4 antagonist is selected from the compounds or pharmaceutical compositions disclosed in WO 2013 / 082490 filed by ChemoCentryx, the contents of which are incorporated herein for all purposes.

[0120] B. Immune checkpoint inhibitors

[0121] Immune checkpoints are signaling proteins that stimulate or inhibit immune responses. Compositions targeting immune checkpoints can modulate these proteins, thereby altering an individual's natural immune response. This targeted approach is useful because certain cancer cells can bypass these checkpoints to escape otherwise innate immune responses. Two specific immune checkpoints are programmed cell death protein 1 (PD-1) and cytotoxic T lymphocyte-associated protein 4 (CTLA-4).

[0122] i. PD-1 inhibitors

[0123] Programmed cell death protein-1 (PD-1) is an immune checkpoint protein that is most commonly found on T cells. Normally, PD-1 binds to its natural ligands, PD-L1 and PD-L2, which are expressed on the surface of different cells. When bound to its natural ligands, T cells are considered to be in the "off" position. Notably, some cancer cells express abnormally high levels of PD-L1, which means that the activity of T cells and the associated anti-cancer immune response are abnormally suppressed. Most importantly, the use of PD-1 inhibitors that block the interaction with its natural ligands can stimulate the immune response to help fight cancer.

[0124] The PD-1 inhibitors of the present invention include small molecules and antibodies.

[0125] In some embodiments, the PD-1 inhibitor is a small molecule PD-1 / PD-L1 inhibitor.

[0126] In some embodiments, the small molecule PD-1 / PD-L1 inhibitor has the following formula:

[0127]

[0128] In some embodiments, the small molecule PD-1 / PD-L1 inhibitor is a compound having formula (II)

[0129]

[0130] or a pharmaceutically acceptable salt thereof; wherein,

[0131] R 1 Selected from the group consisting of halogen, C 5-8 Cycloalkyl, C 6-10 Aryl and thienyl, where C 6-10 The aryl and thienyl groups are optionally substituted with 1 to 5 R x substituted by a substituent;

[0132] Each R x Independently selected from the group consisting of halogen, -CN, -R c 、-CO2R a 、-CONR a R b 、-C(O)R a 、-OC(O)NR a R b 、-NR b C(O)R a 、-NR b C(O)2R c 、-NR a -C(O)NR a R b 、-NRa R b 、-OR a 、-OX 1 -OR a 、-OX 1 -CO2R a 、-OX 1 -CONR a R b 、-X 1 -OR a 、-X 1 -NR a R b 、-X 1 -CO2R a 、-X 1 -CONR a R b , -SF5 and -S(O)2NR a R b , where each X 1 C 1-4 Alkylene; wherein each R a and R b Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 haloalkyl, or when attached to the same nitrogen atom, may be combined with the nitrogen atom to form a five-membered or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members; wherein the five-membered or six-membered ring is optionally substituted with oxo; each R c Independently selected from the group consisting of: C 1-8 Alkyl, C 2-8 Alkenyl, C 2-8 Alkynyl and C 1-8 Haloalkyl; and optionally when two R x When the substituents are on adjacent atoms, they combine to form a fused five, six or seven membered carbocyclic or heterocyclic ring, which is optionally substituted with 1 to 3 substituents independently selected from the group consisting of halogen, oxo, C 1-8 Haloalkyl and C 1-8 alkyl;

[0133] Each R 2a 、R 2b and R 2c Independently selected from the group consisting of H, halogen, -CN, -R d 、-CO2R e 、-CONR e R f 、-C(O)R e 、-OC(O)NR e R f 、-NRf C(O)R e 、-NR f C(O)2R d 、-NR e -C(O)NR e R f 、-NR e R f 、-OR e 、-OX 2 -OR e 、-OX 2 -NR e R f 、-OX 2 -CO2R e 、-OX 2 -CONR e R f 、-X 2 -OR e 、-X 2 -NR e R f 、-X 2 -CO2R e 、-X 2 -CONR e R f 、-SF5、-S(O)2NR e R f 、C 6-10 Aryl and C 5-10 Heteroaryl, wherein each X 2 C 1-4 Alkylene; each R e and R f Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 haloalkyl, or when attached to the same nitrogen atom, may be combined with the nitrogen atom to form a five-membered or six-membered ring having 0 to 2 additional heteroatoms selected from N, O and S as ring members, and optionally substituted with oxo; each R d Independently selected from the group consisting of: C 1-8 Alkyl, C 2-8 Alkenyl and C 1-8 alkyl halide;

[0134] R 3 Selected from the group consisting of: -NR g R h and C 4-12 Heterocyclic group, wherein the C 4-12 The heterocyclic group is optionally substituted by 1 to 6 R y replaced by;

[0135] Each Ry Independently selected from the group consisting of halogen, -CN, -R i 、-CO2R j 、-CONR j R k 、-CONHC 1-6 Alkyl-OH, -C(O)R j 、-OC(O)NR j R k 、-NR j C(O)R k 、-NR j C(O)2R k 、CONOH、PO3H2、-NR j -C 1-6 Alkyl-C(O)2R k 、-NR j C(O)NR j R k 、-NR j R k 、-OR j 、-S(O)2NR j R k 、-OC 1-6 Alkyl-OR j 、-OC 1-6 Alkyl-NR j R k 、-OC 1-6 Alkyl-CO2R j 、-OC 1-6 Alkyl-CONR j R k 、-C 1-6 Alkyl-OR j 、-C 1-6 Alkyl-NR j R k 、-C 1-6 Alkyl-CO2R j 、-C 1-6 Alkyl-CONR j R k and SF5,

[0136] Among them, R y C 1-6 The alkyl portion is optionally further substituted with OH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 Alkyl or CO2H substituted, wherein each R j and R kindependently selected from: hydrogen, optionally substituted by 1 to 2 selected from OH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 C substituted by alkyl or CO2H 1-8 Alkyl, and optionally 1 to 2 selected from OH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 C substituted by alkyl or CO2H 1-8 haloalkyl, or when attached to the same nitrogen atom, R j and R k can be combined with the nitrogen atom to form a five-membered or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with an oxo group; each R i Independently selected from the group consisting of: -OH, C 1-8 Alkyl, C 2-8 Alkenyl and C 1-8 haloalkyl, each of which is optionally substituted with OH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 Alkyl or CO2H substituted;

[0137] R g Selected from the following group: H, C 1-8 Haloalkyl and C 1-8 alkyl;

[0138] R h Selected from: -C optionally substituted with CO2H 1-8 Alkyl, C 1-8 Halogenated alkyl, C 1-8 Alkyl-COOH, C 1-8 Alkyl-OH, C 1-8 Alkyl-CONH2, C 1-8 Alkyl-SO2NH2, C 1-8 Alkyl-PO3H2, C 1-8 Alkyl-CONOH, C 1-8 Alkyl-NR h1 R h2 、-C(O)-C 1-8 Alkyl, -C(O)-C 1-8 Alkyl-OH, -C(O)-C 1-8 Alkyl-COOH, C 3-10 Cycloalkyl, -C 3-10 Cycloalkyl-COOH, -C 3-10 Cycloalkyl-OH, C 4-8 Heterocyclic group, -C 4-8 Heterocyclic -COOH, -C 4-8 Heterocyclic-OH, -C1-8 Alkyl-C 4-8 Heterocyclic group, -C 1-8 Alkyl-C 3-10 Cycloalkyl, C 5-10 Heteroaryl, -C 1-8 Alkyl-C 5-10 Heteroaryl, C 10 Carbocyclic group, -C 1-8 Alkyl-C 6-10 Aryl, -C 1-8 Alkyl-(C=O)-C 6-10 Aryl, -C 1-8 Alkyl-NH(C=O)-C 1-8 Alkenyl, -C 1-8 Alkyl-NH(C=O)-C 1-8 Alkyl, -C 1-8 Alkyl-NH(C=O)-C 1-8 Alkynyl, -C 1-8 Alkyl-(C=O)-NH-C 1-8 Alkyl-COOH and -C 1-8 Alkyl-(C=O)-NH-C 1-8 Alkyl-OH;

[0139] R h The N-linked peptide is a mono-, di- or tripeptide comprising 1-3 natural amino acids and 0-2 unnatural amino acids, wherein

[0140] The unnatural amino acid has an alpha carbon substituent selected from the group consisting of: C 2-4 Hydroxyalkyl, C 1-3 Alkyl-guanidino and C 1-4 Alkyl-heteroaryl,

[0141] The alpha carbon of each natural or unnatural amino acid is optionally further substituted with a methyl group, and

[0142] The terminal moiety of the mono-, di- or tripeptide is selected from the group consisting of C(O)OH, C(O)OC 1-6 Alkyl and PO3H2, where

[0143] R h1 and R h2 Each independently selected from the following group: H, C 1-6 Alkyl and C 1-4 hydroxyalkyl;

[0144] R h C 1-8 The alkyl moiety is optionally further substituted with 1 to 3 substituents independently selected from the group consisting of OH, COOH, SO2NH2, CONH2, CONOH, COO-C 1-8alkyl, PO3H2 and optionally 1 to 2 C 1-3 Alkyl substituents substituted C 5-6 heteroaryl,

[0145] R h C 10 Carbocyclic group, C 5-10 Heteroaryl and C 6-10 The aryl moiety is optionally substituted with 1 to 3 substituents independently selected from the group consisting of OH, B(OH)2, COOH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 Alkyl, C 1-4 Alkyl, C 1-4 Alkyl-OH, C 1-4 Alkyl-SO2NH2, C 1-4 AlkylCONH2, C 1-4 Alkyl-CONOH, C 1-4 Alkyl-PO3H2, C 1-4 Alkyl-COOH and phenyl and

[0146] R h C 4-8 Heterocyclic and C 3-10 The cycloalkyl moiety is optionally substituted with 1 to 4 R w Substituent substitution;

[0147] Each R w The substituents are independently selected from: C 1-4 Alkyl, C 1-4 Alkyl-OH, C 1-4 Alkyl-COOH, C 1-4 Alkyl-SO2NH2, C 1-4 AlkylCONH2, C 1-4 Alkyl-CONOH, C 1-4 Alkyl-PO3H, OH, COO-C 1-8 Alkyl, COOH, SO2NH2, CONH2, CONOH, PO3H2 and oxo;

[0148] R 4 Selected from the following group: OC 1-8 Alkyl, OC 1-8 Halogenated alkyl, OC 1-8 Alkyl-R z 、C 6-10 Aryl, C 5-10 Heteroaryl, -OC 1-4 Alkyl-C 6-10 Aryl and -OC 1-4 Alkyl-C 5-10 Heteroaryl, wherein C 6-10Aryl and C 5-10 Heteroaryl is optionally substituted by 1 to 5 R z replace;

[0149] Each R z Independently selected from the group consisting of halogen, -CN, -R m 、-CO2R n 、-CONR n R p 、-C(O)R n 、-OC(O)NR n R p 、-NR n C(O)R p 、-NR n C(O)2R m 、-NR n -C(O)NR n R p 、-NR n R p 、-OR n 、-OX 3 -OR n 、-OX 3 -NR n R p 、-OX 3 -CO2R n 、-OX 3 -CONR n R p 、-X 3 -OR n 、-X 3 -NR n R p 、-X 3 -CO2R n 、-X 3 -CONR n R p 、-SF5、-S(O)2R n R p 、-S(O)2NR n R p , and a three- to seven-membered carbocyclic ring or a four- to seven-membered heterocyclic ring, wherein the three- to seven-membered carbocyclic ring or the four- to seven-membered heterocyclic ring is optionally substituted by 1 to 5 R t substituted, wherein each R t Independently selected from the group consisting of: C 1-8 Alkyl, C 1-8 Haloalkyl, -CO2R n 、-CONR n R p 、-C(O)R n 、-OC(O)NRn R p 、-NR n C(O)R p 、-NR n C(O)2R m 、-NR n -C(O)NR n R p 、-NR n R p 、-OR n 、-OX 3 -OR n 、-OX 3 -NR n R p 、-OX 3 -CO2R n 、-OX 3 -CONR n R p 、-X 3 -OR n 、-X 3 -NR n R p 、-X 3 -CO2R n 、-X 3 -CONR n R p , -SF5 and -S(O)2NR n R p ;

[0150] Among them, each X 3 C 1-4 Alkylene; each R n and R p Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 haloalkyl, or when attached to the same nitrogen atom, may be combined with the nitrogen atom to form a five-membered or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with oxo; each R m Independently selected from the following groups: C 1-8 Alkyl, C 2-8 Alkenyl and C 1-8 Haloalkyl; and optionally when two R z When the substituents are on adjacent atoms, they combine to form a fused five- or six-membered carbocyclic or heterocyclic ring optionally substituted with oxo;

[0151] n is 0, 1, 2, or 3;

[0152] Each R 5Independently selected from the group consisting of halogen, -CN, -R q 、-CO2R r 、-CONR r R s 、-C(O)R r 、-OC(O)NR r R s 、-NR r C(O)R s 、-NR r C(O)2R q 、-NR r -C(O)NR r R s 、-NR r R s 、-OR r 、-OX 4 -OR r 、-OX 4 -NR r R s 、-OX 4 -CO2R r 、-OX 4 -CONR r R s 、-X 4 -OR r 、-X 4 -NR r R s 、-X 4 -CO2R r 、-X 4 -CONR r R s 、-SF5、-S(O)2NR r R s , where each X 4 C 1-4 Alkylene; each R r and R s are independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 haloalkyl, or when attached to the same nitrogen atom, may be combined with the nitrogen atom to form a five-membered or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with oxo; each R q Independently selected from the group consisting of: C 1-8 Alkyl and C 1-8 alkyl halide;

[0153] R 6a Selected from the following group: H, C 1-4 Alkyl and C 1-4alkyl halide;

[0154] Each R 6b Independently selected from the group consisting of F, C 1-4 Alkyl, OR u 、C 1-4 Halogenated alkyl, NR u R v , where each R u and R v Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 Haloalkyl, or when attached to the same nitrogen atom, may be combined with the nitrogen atom to form a five- or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with oxo; and

[0155] m is 0, 1, 2, 3 or 4.

[0156] In some embodiments, the compound, or a pharmaceutically acceptable salt thereof, has formula (IIa)

[0157]

[0158] In some embodiments, the compound, or a pharmaceutically acceptable salt thereof, has formula (IIb)

[0159]

[0160] In some embodiments, R 1 Selected from the group consisting of phenyl and thienyl, wherein the phenyl and thienyl groups are optionally substituted by 1 to 5 R x In some embodiments, R 1 is optionally replaced by 1 or 2 R x substituted phenyl, wherein each R x Independently selected from: halogen, C 1-8 Alkyl, OC 1-8 Alkyl, OC 1-8 Haloalkyl, -NR a R b and CN, and optionally when both R x When the substituents are on adjacent atoms, they combine to form a fused six-membered heterocyclic ring optionally substituted with 1 to 3 substituents independently selected from the group consisting of oxo, C 1-8 Haloalkyl and C 1-8 In some embodiments, R 1 is phenyl optionally substituted with F. In some embodiments, R 1 Select from the following groups:

[0161]

[0162] In some embodiments, each R 2a 、R 2b and R 2c Independently selected from the group consisting of H, halogen, -CN, -R d 、-NR e R f 、-OR e 、-X 2 -OR e 、-X 2 -NR e R f , where X 2 C 1-4 Alkylene; each R e and R f Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 haloalkyl, or when attached to the same nitrogen atom, may be combined with the nitrogen atom to form a five-membered or six-membered ring having 0 to 2 additional heteroatoms selected from N, O or S as ring members, and optionally substituted with oxo; each R d Independently selected from the group consisting of: C 1-8 Alkyl, C 2-8 Alkenyl and C 1-8 haloalkyl; in some embodiments, R 2b and R 2c Both H and R 2a Selected from the group consisting of halogen, C 1-4 Alkyl, C 2-4 Alkenyl, C 1-3 In some embodiments, R 2b and R 2c Both H and R 2a In some embodiments, R 2b and R 2c Both H and R 2a For Cl.

[0163] In some embodiments, n is 0, 1, or 2 and each R 5 Independently selected from the group consisting of halogen, -CN, -R q 、-NR r R s AND-OR r , where each R r and R s Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 haloalkyl and each R q Independently selected from the group consisting of: C1-8 Alkyl and C 1-8 In some embodiments, n is 0.

[0164] In some embodiments, R 6a is H. In some embodiments, m is 0. In some embodiments, m is 1 and R 6b Select from the following groups: F, C 1-4 Alkyl, OR u 、C 1-4 Halogenated alkyl and NR u R v , where each R u and R v Independently selected from: hydrogen, C 1-8 Alkyl and C 1-8 In some embodiments, m is 1 and R 6b For F.

[0165] In some embodiments, for

[0166] In some embodiments, for

[0167] In some embodiments, R 4 Selected from the following group: OC 1-4 Alkyl, OC 1-6 Alkyl-R z 、C 6-10 Aryl, C 5-10 Heteroaryl, -OC 1-4 Alkyl-C 6-10 Aryl and -OC 1-4 Alkyl-C 5-10 Heteroaryl, wherein the C 6-10 Aryl and the C 5-10 Heteroaryl is optionally substituted by 1 to 2 R z Substituted, wherein each R z Independently selected from the group consisting of halogen, -CN, -R m 、-CO2R n 、-CONR n R p 、-C(O)R n 、-OC(O)NR n R p 、-NR n C(O)R p 、-NR n C(O)2R m 、-NR n -C(O)NRn R p 、-NR n R p 、-OR n 、-S(O)2NR n R p , three to seven membered carbocyclic rings and four to seven membered heterocyclic rings, wherein the three to seven membered carbocyclic rings or four to seven membered heterocyclic rings are optionally substituted by 1 to 2 R t Substituted, wherein each R t Independently selected from the group consisting of: C 1-8 Alkyl, C 1-8 Haloalkyl, -CO2R n 、-CONR n R p 、-C(O)R n 、-OC(O)NR n R p 、-NR n C(O)R p 、-NR n C(O)2R m 、-NR n -C(O)NR n R p 、-NR n R p 、-OR n and -S(O)2NR n R p In some embodiments, R 4 Selected from the following group: OC 1-4 Alkyl, OC 1-6 Alkyl-CN, phenyl, pyridyl, -OC 1-2 Alkyl-pyridyl, -OC 1-2 Alkyl-pyrimidinyl, -OC 1-2 Alkyl-pyridazinyl and -OC 1-2 Alkyl-phenyl, wherein the pyridyl, phenyl, pyrimidinyl and pyridazinyl are optionally substituted by 1 to 2 R z Substituted, wherein each R z Independently selected from the group consisting of halogen, -CN, -CO2R n 、-NR n R p 、-OR n and piperidinyl optionally substituted with OH.

[0168] In some embodiments, R 4 Select from the following groups:

[0169]

[0170] In some embodiments, R 4 for

[0171] In some embodiments, R 3 Selected from the following group: NR g R h and C 4-6 Heterocyclic group, wherein the C 4-6 The heterocyclic group is optionally substituted by 1 to 3 R y Substitute, where R g Selected from the following group: H, C 1-8 Haloalkyl and C 1-8 alkyl, and wherein R h is -C substituted by 1 to 3 substituents independently selected from the following 1-8 Alkyl: OH, COOH, SO2NH2, CONH2, CONOH, COO-C 1-8 Alkyl, C 5-6 Heteroaryl, C 5-6 Heterocyclic group and PO3H2, wherein C 5-6 Heteroaryl and C 5-6 The heterocyclic group is optionally substituted with 1 to 3 substituents independently selected from the group consisting of OH, B(OH)2, COOH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 Alkyl, C 1-4 Alkyl, C 1-4 Alkyl-OH, C 1-4 Alkyl-SO2NH2, C 1-4 AlkylCONH2, C 1-4 Alkyl-CONOH, C 1-4 Alkyl-PO3H2 and C 1-4 Alkyl-COOH and wherein the C 5-6 The heterocyclic group is optionally substituted with an oxo group. 3 is selected from the group consisting of azetidinyl, pyrrolidinyl and piperidinyl, wherein the azetidinyl, pyrrolidinyl or piperidinyl is attached via a nitrogen atom, and wherein the azetidinyl, pyrrolidinyl or piperidinyl is optionally substituted by 1 to 3 R y Substituted, wherein each R y Independently selected from the group consisting of -CO2H, CONOH, PO3H2, OH, SO2NH2, CONH2 and COO-C 1-8 In some embodiments, R 3 NHR h , where R h is -C substituted by 1 to 2 substituents independently selected from the following 1-8Alkyl: OH, COOH, CONH2, PO3H2, tetrazolyl, tetrazolonyl and pyrazolyl. 3 Select from the following groups:

[0172]

[0173] In some embodiments, R 3 -NR g R h In some embodiments, R h The N-linked peptide is a mono-, di- or tripeptide comprising 1-3 natural amino acids and 0-2 unnatural amino acids, wherein:

[0174] The non-natural amino acid has an α-carbon substituent selected from the group consisting of: C 2-4 Hydroxyalkyl, C 1-3 Alkyl-guanidino and C 1-4 Alkyl-heteroaryl,

[0175] The alpha carbon of each natural or unnatural amino acid is optionally further substituted with a methyl group, and

[0176] The terminal moiety of the mono-, di- or tripeptide is selected from the group consisting of C(O)OH, C(O)OC 1-6 Alkyl and PO3H2.

[0177] In some embodiments, R h Each natural amino acid is independently selected from the group consisting of serine, alanine, glycine, lysine, arginine, threonine, phenylalanine, tyrosine, aspartic acid, asparagine, histidine and leucine.

[0178] In some embodiments, R 1 is optionally replaced by 1 to 3 R x Substituted phenyl, R 6b H, R 4 Selected from the following group: OC 1-4 Alkyl, OC 1-6 Alkyl-CN, phenyl, pyridyl, -OC 1-2 Alkyl-pyridyl, -OC 1-2 Alkyl-pyrimidinyl, -OC 1-2 Alkyl-pyridazinyl and -OC 1-2 Alkyl-phenyl, wherein the pyridyl, phenyl, pyrimidinyl and pyridazinyl are optionally substituted by 1 to 2 R z Substituted, wherein each R z Independently selected from the group consisting of halogen, -CN, -CO2R n 、-NR n Rp 、-OR n and piperidinyl optionally substituted with OH, and R 3 Selected from the following group: NR g R h and C 4-6 Heterocyclic group, wherein the C 4-6 The heterocyclic group is optionally substituted by 1 to 3 R y Substitute, where R g Selected from the following group: H, C 1-8 Haloalkyl and C 1-8 alkyl, and wherein R h is -C substituted by 1 to 3 substituents independently selected from the following 1-8 Alkyl: OH, COOH, SO2NH2, CONH2, CONOH, COO-C 1-8 Alkyl, C 5-6 Heteroaryl, C 5-6 Heterocyclic group and PO3H2, wherein C 5-6 Heteroaryl and C 5-6 The heterocyclic group is optionally substituted with 1 to 3 substituents independently selected from the group consisting of OH, B(OH)2, COOH, SO2NH2, CONH2, CONOH, PO3H2, COO-C 1-8 Alkyl, C 1-4 Alkyl, C 1-4 Alkyl-OH, C 1-4 Alkyl-SO2NH2, C 1-4 AlkylCONH2, C 1-4 Alkyl-CONOH, C 1-4 Alkyl-PO3H2 and C 1-4 Alkyl-COOH, and wherein the C 5-6 The heterocyclyl group is also optionally substituted with oxo.

[0179] In some embodiments, R 1 is optionally replaced by 1 or 2 R x substituted phenyl, wherein each R x Independently selected from: halogen, C 1-8 Alkyl, OC 1-8 Alkyl, OC 1-8 Halogenated alkyl, -NR a R b and CN, where R 2b and R 2c Both H, R 2a Selected from the group consisting of halogen, C 1-4 Alkyl, C 1-3 Haloalkyl, -CN, -OMe and OEt, R 6a is H, m is 0, n is 0, R4 for and R 3 Selected from the following group: NHR h , azetidinyl, pyrrolidinyl and piperidinyl, wherein the azetidinyl, pyrrolidinyl or piperidinyl is connected through a nitrogen atom, and wherein the azetidinyl, pyrrolidinyl or piperidinyl is optionally substituted by 1 to 3 R y Substituted, wherein each R y Independently selected from the group consisting of CO2H, CONOH, PO3H2, OH, SO2NH2, CONH2 and COO-C 1-8 Alkyl, and wherein R h is C substituted by 1 to 2 substituents independently selected from the following 1-8 Alkyl: OH, COOH, CONH2, PO3H2, tetrazolyl, tetrazolonyl and pyrazolyl. 2a It is a halogen.

[0180] In some embodiments, the small molecule PD-1 / PD-L1 inhibitor is selected from the compounds or pharmaceutical compositions disclosed in WO2018 / 005374 filed by ChemoCentryx on June 26, 2017, the contents of which are incorporated herein for all purposes.

[0181] In some embodiments, the PD-1 inhibitor is an antibody. In some embodiments, the PD-1 inhibitor antibody is selected from the group consisting of: nivolumab, pembrolizumab, and pidilizumab. In some embodiments, the PD-1 inhibitor antibody is nivolumab. In some embodiments, the PD-1 inhibitor antibody is pembrolizumab. In some embodiments, the PD-1 inhibitor antibody is pidilizumab.

[0182] The PD-1 inhibitors of the present invention can be prepared using methods known in the art. For example, SCID mice can be used to prepare the human monoclonal antibodies of the present disclosure, into which human immune cells have been recombined so that a human antibody response can be produced upon immunization. Such mice are described, for example, in US Patent Serial Nos. 5,476,996 and 5,698,767 to Wilson et al. The PD-1 inhibitors of the present disclosure can be formulated to delay the degradation of the compound or antibody or to minimize the immunogenicity of the antibody. A variety of techniques are known in the art to achieve this goal.

[0183] ii. CTLA-4 inhibitors

[0184] Cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) is an immune checkpoint protein most commonly expressed by activated T cells. Similar to PD-1, CTLA-4 acts as a negative regulator of T cell immune function. Providing CTLA-4 inhibitors reduces negative regulation and enhances T cell anti-tumor activity.

[0185] In some embodiments, the CTLA-4 inhibitor is an antibody. In some embodiments, the CTLA-4 inhibitor antibody has about 10 8 M -1 In some embodiments, the CLTA-4 inhibitor antibody has a binding affinity of about 10 9 M -1 In some embodiments, the CTLA-4 inhibitor antibody can inhibit the binding of human CTLA-4 to B7-1 or B7-2.

[0186] In some embodiments, the CTLA-4 inhibitor antibody is selected from the group consisting of: ipilimumab, tremelimumab, AGEN1884, and AGEN2041. In some embodiments, the CTLA-4 inhibitor antibody is ipilimumab. In some embodiments, the CTLA-4 inhibitor antibody is tremelimumab. In some embodiments, the CTLA-4 inhibitor antibody is AGEN1884. In some embodiments, the CTLA-4 inhibitor antibody is AGEN2041.

[0187] In some embodiments, the CTLA-4 inhibitor is selected from the compounds or pharmaceutical compositions disclosed in WO / 2009 / 100140 or WO2017 / 194265, each of which is incorporated herein for all purposes.

[0188] The CTLA-4 inhibitors of the present invention can be prepared using methods known in the art. For example, human monoclonal antibodies of the present disclosure can be prepared using SCID mice into which human immune cells have been recombined so that a human antibody response can be generated upon immunization. Such mice are described, for example, in U.S. Patent Nos. 5,476,996 and 5,698,767 to Wilson et al. The CTLA-4 inhibitors of the present disclosure can also be formulated to delay the degradation of a compound or antibody or to minimize the immunogenicity of the antibody. Various techniques are known in the art to achieve this goal.

[0189] IV. Methods of Administration of Combination Therapies

[0190] In another aspect, the present disclosure provides a combination therapy for treating cancer. The combination therapy includes a therapeutically effective amount of a CCR4 antagonist and a therapeutically effective amount of one or more immune checkpoint inhibitors. In some embodiments, the one or more immune checkpoint inhibitors are PD-1 inhibitors. In some embodiments, the one or more immune checkpoint inhibitors are CTLA-4 inhibitors. In some embodiments, the one or more immune checkpoint inhibitors are PD-1 inhibitors and CTLA-4 inhibitors. The combined synergistic effect of the therapeutic drugs affects the treatment or prevention of cancer.

[0191] The term "therapeutically effective amount" refers to the amount of a compound of interest that will elicit the biological or medical response in a cell, tissue, system, or animal (eg, human) that is being sought by a researcher, veterinarian, medical doctor, or other treatment provider.

[0192] Depending on the disease state and the condition of the subject, the compounds, antibodies and formulations of the present disclosure can be administered orally, parenterally (e.g., intramuscularly, intraperitoneally, intravenously, ICV, intracisternal injection or infusion, subcutaneous injection or implant), inhaled, nasally, vaginally, rectally, sublingually or topically. In addition, the compounds and antibodies can be formulated, alone or together, into suitable dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants and vehicles suitable for each route of administration. The present disclosure also contemplates the administration of the compounds and antibodies of the present disclosure as long-acting formulations.

[0193] It is understood that the specific dosage level and frequency of dosage for any particular patient may vary and will depend on a variety of factors including the activity of the specific compound employed, the metabolic stability and duration of action of the compound, the age, weight, genetic characteristics, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, the severity of the particular disease state, and the host being treated.

[0194] Combination therapy includes co-administration of a CCR4 antagonist and one or more immune checkpoint inhibitors, sequential administration of a CCR4 antagonist and one or more checkpoint inhibitors, administration of a composition comprising a CCR4 antagonist and one or more checkpoint inhibitors, or simultaneous administration of a separate composition (one composition containing a CCR4 antagonist and one or more compositions containing one or more checkpoint inhibitors). In embodiments where two immune checkpoint inhibitors are administered, it will be understood that each immune checkpoint inhibitor can be formulated separately or in a single dosage unit.

[0195] Co-administration is included in administering the CCR4 antagonist of the present disclosure within 0.5, 1, 2, 4, 6, 8, 10, 12, 16, 20 or 24 hours of administering one or more immune checkpoint inhibitors of the present disclosure.Co-administration also includes simultaneous administration, approximately simultaneous administration (for example, within about 1, 5, 10, 15, 20 or 30 minutes between each other), or in any order of sequential administration.In addition, CCR4 antagonist and one or more checkpoint inhibitors can be each administered once a day, or twice, three times or multiple times a day, to provide a preferred dosage level every day.

[0196] V. Kit

[0197] In some aspects, provided herein are kits containing CCR4 antagonists and one or more immune checkpoint inhibitors, which can be used to treat cancer. A kit can include a pharmaceutical composition containing a CCR4 antagonist compound (e.g., a small molecule inhibitor of CCR4) and one or more pharmaceutical compositions containing immune checkpoint inhibitors (e.g., anti-PD-1 inhibitors and / or anti-CTLA-4 inhibitors). In some cases, the kit includes written materials, for example, instructions for use of a compound, an antibody, or its pharmaceutical composition. Without limitation, the kit may include a buffer, a diluent, a filter, a needle, a syringe, and an instruction package insert for performing any method disclosed herein.

[0198] VI. Examples

[0199] Example 1: CC Chemokine Receptor 4 (CCR4) Antagonists Enhance the Effects of Checkpoint Inhibition in a Mouse Tumor Model (Summary)

[0200] Chemokines and their receptors influence many hallmark processes of cancer: they act not only on infiltrating leukocytes but also on fibroblasts, endothelial cells, and directly on certain tumor cell types. C-C chemokine receptor 4 (CCR4) and its ligands are found to be highly expressed in multiple human tumor types and are associated with poor prognosis. CCR4 antagonists have been shown to reduce tumor growth in various mouse tumor models. Here, we evaluated small-molecule inhibition of CCR4 as a therapeutic agent to enhance the effects of checkpoint inhibitors in the CT26 and KCM tumor models.

[0201] method

[0202] The effects of a CCR4 inhibitor (Compound 1) in combination with an anti-CTLA-4 antibody (BioXcell: anti-mouse CTLA-4 (Clone: ​​9H10), raised in Syrian Hamsters and purified with Protein G) were evaluated in a subcutaneous CT26 colon cancer model and an orthotopic KCM pancreatic cancer model. CT26 cells were implanted into the flanks of 9-week-old female Balb / c mice. Mice were randomized into study groups based on tumor size on day 6, and administration of Compound 1 and anti-CTLA-4 began on day 7. For the orthotopic pancreatic cancer model, KCM cells were implanted directly into the pancreas, and administration of Compound 1 and anti-CTLA-4 also began on day 7. Compound 1 was administered orally twice daily at a dose of 30 mg / kg, and anti-CTLA-4 was administered intraperitoneally (IP) on days 7, 11, and 15 at a dose of 100 μg / mouse.

[0203] result

[0204] Blockade of CCR4 significantly enhanced the therapeutic efficacy of anti-CTLA-4 in both models. Combined anti-CTLA-4 / CCR4 inhibition significantly reduced tumor size and increased the proportion of long-term survivors in the CT26 model. The anti-tumor response was specific to CT26; long-term survivors were resistant to reinoculation with CT26 cells (without the need for further administration of either drug), but 4T1 mammary tumors grew well after challenge of CT26 survivors. Mice with tumor regression exhibited a high proportion of CD8 T cells that recognized CT26-specific neoantigens, as indicated by AH1 peptide-MHC tetramer staining.

[0205] Although long-term survival in the KCM model has not been studied, compound 1 alone significantly reduced tumor burden in three independent studies. Anti-CTLA-4 alone provided substantial inhibition of tumor growth in this model, an effect further enhanced by compound 1.

[0206] Example 2: Compound 1 is a highly potent and selective CCR4 inhibitor

[0207] By determining the IC relative to different groups of receptors, enzymes, and ion channels 50 The selectivity of compound 1 was tested using the 5-mercaptoethanol (5-mercaptoethanol) value. Compound 1 was found to be highly specific for CCR4. See Table 1.

[0208] Table 1: Selectivity of Compound 1

[0209]

[0210] The binding activity of compound 1 in human and mouse is reported in Table 2.

[0211] Table 2: Cross-species activity

[0212]

[0213] Example 3: Combination therapy using the KCM orthotopic pancreatic tumor model.

[0214] As briefly described in Example 1, the KCM pancreatic cancer model was used to evaluate the efficacy of CCR4 inhibitors in combination with anti-CTLA-4 antibodies. Figure 1 As shown in . Mice were divided into 4 groups (n=12 per group): antibody isotype + vehicle; antibody isotype + compound 1; anti-CTLA-4 + vehicle; anti-CTLA-4 + compound 1. The mice used were 7-8 week old C57BL / 6 female mice. On days 7, 11 and 16, anti-CTLA-4 (BioXcell: anti-mouse CTLA-4 clone (Clone): 9H10), Syrian Hamster culture, purified with protein G) and isotype-matched control antibodies were injected intraperitoneally (ip) at a dose of 100 μg / mouse. Compound 1 (40 mg / kg) and vehicle were administered twice daily (bid) pi. KCM cells in matrigel were injected into the pancreas.

[0215] After the study was completed, tumor / pancreas weights were measured for each group. The results are shown in Figures AC. As shown in the figure, compound 1, alone or in combination with anti-CTLA-4, reduced tumor burden in the KCM tumor model. In this model, most mice develop secondary tumors on the abdominal wall near the incision site. These secondary tumors are dissected and their weights are included in the total tumor plus pancreas weight. Because the primary tumor cannot be separated from the pancreas by surgery, they are weighed together. Spleen weight was also reduced by compound 1 treatment. Adequate compound levels were confirmed by measuring plasma compound concentrations at the trough on day 12.

[0216] At the completion of the study, changes in suppressive immune cell populations were also measured. On day 25 after KCM cell inoculation, primary tumors (along with the pancreas) and secondary tumors were dissected, minced, and digested with collagenase D for immune cell analysis by flow cytometry. Pancreata from sham-operated mice were processed and analyzed. Figure 3A-F shows the results of the changes in T regulatory cells, G-MDSC and M-MDSC from the processed materials. These cell populations are shown as the number of cells per gram of tissue (groups AC) or as the percentage of CD45+ cells (groups DF). Although no changes in T regulatory (CD4+ / FoxP3+) cells were observed, monocytes and granulocyte bone marrow-derived suppressor cells (mMDSC-CD11b+ / Ly6G- / Ly6C-high, gMDSC-CD11b+ / Ly6G+ / Ly6C-low) were shown to be decreasing when treated with a-CTLA-4 / compound 1 compared to a-CTLA-4 alone.

[0217] Example 4: Combination therapy using the CT26 colon cancer model.

[0218] As briefly described in Example 1, the CT26 colon cancer model was used to evaluate the efficacy of CCR4 inhibitors in combination with anti-CTLA-4 antibodies. Figure 4 As shown in . Mice were divided into 4 groups (n=12 per group): antibody isotype + vehicle; antibody isotype + compound 1; anti-CTLA-4 + vehicle; anti-CTLA-4 + compound 1. The mice used were 7-8 week old C57BL / 6 female mice. On days 7, 11 and 16, anti-CTLA-4 (BioXcell: anti-mouse CTLA-4 clone (Clone): 9H10), Syrian Hamster culture, purified with protein G) and isotype-matched control antibodies were injected intraperitoneally (ip) at a dose of 100 μg / mouse. Compound 1 (40 mg / kg) and vehicle were administered pi twice daily (bid). At the beginning of the study, CT25 cells were injected subcutaneously (sc).

[0219] Tumor volume and survival percentage were monitored throughout the study. Survival was plotted on Figure 5 , and the mean tumor volume is plotted on Figure 6 In. From Figure 6 As can be seen in Figure 2, the combination of Compound 1 and anti-CTLA-4 reduced tumor growth and improved survival. Figure 7A Note how remarkable it is that the tumors of mice in the aCTLA-4 + vehicle group (Panel C) did not respond at all to aCTLA-4 treatment, whereas the combination of aCTLA + Compound 1 showed that all tumors responded to the treatment (Panel D).

[0220] Three months after the last dose, mice that had previously been treated with α-CTLA-4 (6 mice) or α-CTLA-4 / Compound 1 (8 mice) and had completely regressed tumors were re-challenged with CT26 tumor cells injected subcutaneously (sc) in the left flank. These mice were also injected subcutaneously (sc) in the right flank with 4T1 mouse breast cancer cells. Figure 8A -F. As can be seen from this figure, one of the six mice previously treated with α-CTLA-4 developed a CT26 tumor (Group A), and none of the eight mice previously treated with α-CTLA-4 / Compound 1 developed a CT26 tumor (Group B). None of the six mice previously treated with α-CTLA-4 developed a 4T1 tumor (Group D), and none of the eight mice previously treated with α-CTLA-4 / Compound 1 developed a 4T1 tumor (Group E). All five blank mice developed both CT26 and 4T1 tumors (Groups C and F).

[0221] Before and 7 days after CT26 rechallenge, blood cells from the mice in Figure 8 were stained with fluorescently labeled AH1 peptide. AH1 is an immunodominant peptide in the cytotoxic T cell response to CT26 tumor cells and can label CT26 antigen-reactive CD8+ T cells. Flow cytometric analysis of these samples was performed as shown in Figure 8. Figure 9A These CT26 antigen-reactive CD8+ T cells were largely absent from the peripheral blood of naive mice, but 3 months after the last dose, abundant CD8+ T cells were present in most surviving CT26 mice (Panels A and B). Rechallenge with CT26 cells further increased the number of these cells in previously treated surviving mice, but had little effect in naive mice (Panel C).

[0222] Although the foregoing invention has been described in detail by way of illustration and example for purposes of clarity of understanding, it will be appreciated by those skilled in the art that certain changes and modifications may be implemented within the scope of the appended claims. In addition, each reference provided herein is incorporated by reference in its entirety to the same extent as if each reference were individually incorporated by reference. In the event of a conflict between the present application and the references provided herein, the present application shall prevail.

Claims

1. Use of a small molecule CCR4 antagonist and an anti-CTLA-4 antibody in the preparation of a drug or kit for treating colon cancer or pancreatic cancer; in, The small molecule CCR4 antagonist is compound 1 shown below; or a pharmaceutically acceptable salt thereof.

2. The use according to claim 1, characterized in that The anti-CTLA-4 antibody is selected from the group consisting of ipilimumab, tremelimumab, AGEN1884, and AGEN2041.

3. The use according to claim 2, characterized in that The anti-CTLA-4 antibody is ipilimumab.

4. The use according to claim 2, characterized in that The anti-CTLA-4 antibody is tremelimumab.

5. The use according to claim 2, characterized in that The anti-CTLA-4 antibody is AGEN1884.

6. The use according to claim 2, characterized in that The anti-CTLA-4 antibody is AGEN2041.

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