Methods of treating cancer

Abstract

Provided herein are methods for predicting the efficacy of a cancer treatment in a subject. Methods for treating a cancer subject are also disclosed. Provided herein are methods for treating a cancerous subject comprising administering a therapeutically effective amount of an immune checkpoint inhibitor to subjects previously identified or diagnosed as having cancer and previously identified as a reduced M2 macrophage level compared to a reference level.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 324,943, filed on March 29, 2022, the entire contents of which are incorporated herein by reference. Summary of the invention

[0003] Provided herein are methods for treating a subject having cancer, comprising administering a therapeutically effective amount of an immune checkpoint inhibitor to a subject previously identified or diagnosed as having cancer and previously identified as having reduced levels of M2 macrophages compared to a reference level.

[0004] Also provided herein are methods of selecting a pharmaceutical composition comprising an immune checkpoint inhibitor for a subject previously identified or diagnosed as having cancer and previously identified as having reduced levels of M2 macrophages compared to a reference level.

[0005] Also provided herein are methods of selecting a subject previously identified or diagnosed as having cancer and previously identified as having reduced levels of M2 macrophages compared to a reference level for treatment with a pharmaceutical composition comprising an immune checkpoint inhibitor.

[0006] Also provided herein is a method for predicting the therapeutic efficacy of a pharmaceutical composition comprising an immune checkpoint inhibitor on a subject in need thereof, the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having a reduced level of M2 macrophages compared to a reference level has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or determining that a subject diagnosed or identified as having cancer and identified as having an increased level of M2 macrophages compared to a reference level has a reduced likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition.

[0007] In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

[0008] In some embodiments of any of the methods described herein, the method further comprises: determining the level of M2 macrophages in a sample obtained from the subject; and identifying that the subject has an increased (or elevated) level of M2 macrophages compared to a reference level.

[0009] In some embodiments of any of the methods described herein, the method further comprises: determining the level of M2 macrophages in a sample obtained from the subject; and identifying that the subject has a reduced level of M2 macrophages compared to a reference level.

[0010] Also provided herein are methods of treating a subject having cancer by administering a therapeutically effective amount of an anti-ILT4 binding agent to a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of M2 macrophages compared to a reference level.

[0011] Also provided herein are methods of selecting a pharmaceutical composition comprising an anti-ILT4 binding agent for a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of M2 macrophages compared to a reference level.

[0012] Also provided herein are methods of selecting a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of M2 macrophages compared to a reference level for treatment with a pharmaceutical composition comprising an anti-ILT4 binding agent.

[0013] In some embodiments of any of the methods described herein, the anti-ILT4 binding agent is an anti-ILT4 antibody or an antigen-binding fragment thereof (eg, a blocking antibody that blocks the interaction of ILT4 / LILRB2 with an HLA-G molecule).

[0014] Some embodiments of any of the methods described herein further comprise administering an immune checkpoint inhibitor to the subject. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

[0015] Some embodiments of any of the methods described herein further comprise: determining the level of M2 macrophages in a sample obtained from the subject; and identifying that the subject has an elevated level of M2 macrophages compared to a reference level.

[0016] In some embodiments of any of the methods described herein, the M2 macrophages express elevated levels of ILT4 compared to a reference level.

[0017] In some embodiments of any of the methods described herein, the reference level of M2 macrophages is the level of M2 macrophages in a population of healthy subjects.

[0018] In some embodiments of any method described herein, the reference level of M2 macrophages is 75% of the median level of the number of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 80% of the median level of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 85% of the median level of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 90% of the median level of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 95% of the median level of M2 macrophages in a healthy patient population.

[0019] In some embodiments of any of the methods described herein, the cancer is a solid tumor. In some embodiments of any of the methods described herein, wherein the solid tumor is non-small cell lung cancer (NSCLC), hepatocellular carcinoma (HCC), small cell lung cancer (SCLC), head and neck cancer, or esophageal cancer.

[0020] In some embodiments of any of the methods described herein, the cancer is a hematological cancer.

[0021] Also provided herein are methods of treating a subject having cancer comprising administering a therapeutically effective amount of an anti-CCR8 binding agent to a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level.

[0022] Also provided herein are methods of selecting a pharmaceutical composition comprising an anti-CCR8 binding agent for a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level.

[0023] Also provided herein are methods of selecting a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level for treatment with a pharmaceutical composition comprising an anti-CCR8 binding agent.

[0024] Also provided herein is a method for predicting the therapeutic efficacy of a pharmaceutical composition comprising an anti-CCR8 binding agent for a subject in need thereof, the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells and decreased levels of NK cells compared to a reference level has a decreased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition.

[0025] In some embodiments of any of the methods described herein, the anti-CCR8 binding agent is an anti-CCR8 antibody or antigen-binding fragment thereof (eg, a depleting antibody or antigen-binding fragment thereof that has the ability to block ligand binding to CCR8).

[0026] In some embodiments of any of the methods described herein, the method further comprises administering an immune checkpoint inhibitor to the subject. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

[0027] In some embodiments of any of the methods described herein, the method further comprises: determining the level of Treg cells and NK cells in a sample obtained from the subject; and identifying that the subject has elevated levels of Treg cells and NK cells compared to a reference level.

[0028] In some embodiments of any of the methods described herein, the reference level of Treg cells is the level of Treg cells in a population of healthy subjects.

[0029] In some embodiments of any method described herein, the reference level of Treg cells is 75% of the median level of Treg cell number in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 80% of the median level of Treg cells in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 85% of the median level of Treg cells in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 90% of the median level of Treg cells in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 95% of the median level of Treg cells in a healthy patient population.

[0030] In some embodiments of any of the methods described herein, the reference level of NK cells is the level of NK cells in a population of healthy subjects.

[0031] In some embodiments of any method described herein, the reference level of NK cells is 75% of the median level of NK cell number in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 80% of the median level of NK cells in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 85% of the median level of NK cells in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 90% of the median level of NK cells in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 95% of the median level of NK cells in a healthy patient population.

[0032] In some embodiments of any of the methods described herein, the cancer is a solid tumor. In some embodiments of any of the methods described herein, wherein the solid tumor is non-small cell lung cancer (NSCLC), hepatocellular carcinoma (HCC), small cell lung cancer (SCLC), head and neck cancer, or esophageal cancer.

[0033] In some embodiments of any of the methods described herein, the cancer is a hematological cancer.

[0034] Also provided herein are methods for treating a subject having cancer, comprising administering a therapeutically effective amount of an anti-CCR8 binding agent and a therapeutically effective amount of an anti-ILT4 binding agent to a subject previously identified or diagnosed as having cancer and previously identified as having increased levels of Treg cells, increased levels of NK cells, and increased numbers of M2 macrophages compared to a reference level.

[0035] Also provided herein are methods of selecting an anti-CCR8 binding agent and an anti-ILT4 binding agent for a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of NK cells compared to a reference level.

[0036] Also provided herein are methods of selecting a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to a reference level for treatment with an anti-CCR8 binding agent and an anti-ILT4 binding agent.

[0037] Also provided herein are methods for predicting the therapeutic efficacy of an anti-CCR8 binder and an anti-ILT4 binder in a subject in need thereof, the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to reference levels has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or determining that a subject diagnosed or identified as having cancer and not identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to reference levels has a reduced likelihood of having a positive therapeutic response to treatment with the anti-CCR8 binder and an anti-ILT4 binder.

[0038] In some embodiments of any of the methods described herein, the anti-CCR8 binding agent is an anti-CCR8 antibody or antigen-binding fragment thereof (eg, a depleting antibody or antigen-binding fragment thereof that has the ability to block ligand binding to CCR8).

[0039] In some embodiments of any of the methods described herein, the anti-ILT4 binding agent is an anti-ILT4 antibody or an antigen-binding fragment thereof (eg, a blocking antibody that blocks the interaction of ILT4 / LILRB2 with an HLA-G molecule).

[0040] In some embodiments of any of the methods described herein, the method further comprises administering an immune checkpoint inhibitor to the subject. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof. In some embodiments of any of the methods described herein, the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

[0041] In some embodiments of any of the methods described herein, the method further comprises: determining the level of Treg cells, NK cells, and M2 macrophages in a sample obtained from the subject; and identifying that the subject has elevated levels of Treg cells, NK cells, and M2 macrophages compared to a reference level.

[0042] In some embodiments of any of the methods described herein, the reference level of Treg cells is the level of Treg cells in a population of healthy subjects.

[0043] In some embodiments of any method described herein, the reference level of Treg cells is 75% of the median level of Treg cell number in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 80% of the median level of Treg cells in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 85% of the median level of Treg cells in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 90% of the median level of Treg cells in a healthy patient population. In some embodiments of any method described herein, the reference level of Treg cells is 95% of the median level of Treg cells in a healthy patient population.

[0044] In some embodiments of any of the methods described herein, the reference level of NK cells is the level of NK cells in a population of healthy subjects.

[0045] In some embodiments of any method described herein, the reference level of NK cells is 75% of the median level of NK cell number in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 80% of the median level of NK cells in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 85% of the median level of NK cells in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 90% of the median level of NK cells in a healthy patient population. In some embodiments of any method described herein, the reference level of NK cells is 95% of the median level of NK cells in a healthy patient population.

[0046] In some embodiments of any of the methods described herein, the reference level of M2 macrophages is the level of M2 macrophages in a population of healthy subjects.

[0047] In some embodiments of any method described herein, the reference level of M2 macrophages is 75% of the median level of the number of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 80% of the median level of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 85% of the median level of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 90% of the median level of M2 macrophages in a healthy patient population. In some embodiments of any method described herein, the reference level of M2 macrophages is 95% of the median level of M2 macrophages in a healthy patient population.

[0048] In some embodiments of any of the methods described herein, the cancer is a solid tumor. In some embodiments of any of the methods described herein, wherein the solid tumor is non-small cell lung cancer, hepatocellular carcinoma, small cell lung cancer, head and neck cancer, or esophageal cancer.

[0049] In some embodiments of any of the methods described herein, the cancer is a hematological cancer.

[0050] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention belongs. Methods and materials used in the present invention are described herein; other suitable methods and materials known in the art may also be used. The materials, methods, and examples are illustrative only and not limiting. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated herein by reference in their entirety. In the event of a conflict, the present specification, including definitions, shall prevail.

[0051] Other features and advantages of the present invention will become readily apparent from the following detailed description, the accompanying drawings and the appended claims. BRIEF DESCRIPTION OF THE DRAWINGS

[0052] Figure 1A-1B Figure provided shows the association between high levels of immunosuppressive M2 macrophages and limited PD(L)-1 response in patients with small cell lung cancer (SCLC). Figure 1A The graph shows a reanalysis of the patient survival probability data. Figure 1B The chart shows a reanalysis of the patient heat map data.

[0053] Figure 2A-2B The graph provided shows higher ILT4 expression and lower immune T cells in SCLC patients with higher levels of M2 macrophages. Figure 2A The graph shows ILT4 expression in M2 samples versus non-M2 samples. Figure 2B The graph shows CD8 T cells, CD4 naive T cells, CD4 memory resting T cells, and CD4 memory activated T cells in M2-enriched samples. ****p<0.00001

[0054] Figure 3 The graph shows that approximately 30% of non-small cell lung cancer (NSCLC) patients have high expression of immunosuppressive M2 macrophages.

[0055] Figure 4 Schematic diagram showing that anti-ILT4 antibodies repolarize M2 (suppressive) macrophages to M1 (inflammatory) macrophages.

[0056] Figure 5The chart shows a re-analysis of the NSCLC patient heat map data. About 30% of the patients had a high Treg signature, with cluster 1 having high Treg and high NK cells (Treg 高 NK 高 ), group 2 has high Treg and low NK cells (Treg 高 NK 低 ).

[0057] Figure 6 Schematic diagram showing Treg cell interference with anti-PD(L)1 efficacy (left panel) and effective tumor killing using CCR8 antibody in the presence of NK cells (right panel).

[0058] Figure 7 The graph shows the patient populations that will benefit most from the new combination therapy of ILT4 and CCR8 with toripalimab. The graph shows the percentage of patients in the two identified populations, the "Treg Hi, Effector Hi" population and the "M2-enriched" population, that may benefit from the combination therapy. Data are shown for hepatocellular carcinoma (HCC), NSCLC adenocarcinoma, SCLC, head and neck cancer, and esophageal cancer. SCLC M1 and M2 values ​​are based on ADAPTS:LM22 profiles. Treg patterns are derived from GSVA, which uses the ranked LM22 profiles. DETAILED DESCRIPTION

[0059] The present disclosure provides a method for predicting the efficacy of cancer treatment in an object (e.g., a cancer patient). For example, the present disclosure provides a method for predicting immune checkpoint inhibitors (ICIs), anti-CCR8 binding agents (e.g., anti-CCR8 antibodies, such as depleting antibodies) and / or anti-ILT4 binding agents (e.g., anti-ILT4 antibodies, such as blocking antibodies) in cancer patients. A method for treating an object (e.g., a patient) with one or more cancers with cancer treatment is also disclosed. For example, the present disclosure provides a method for treating an object with one or more cancers with ICI, anti-CCR8 binding agents, anti-ILT4 binding agents and / or any combination thereof.

[0060] Predicting the effectiveness of cancer treatment

[0061] Provided herein are methods for predicting the therapeutic efficacy of a pharmaceutical composition comprising an immune checkpoint inhibitor for a subject in need thereof (e.g., a subject suffering from any of the cancers described herein), the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having a decreased level of M2 macrophages compared to a reference level has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or determining that a subject diagnosed or identified as having cancer and identified as having an increased level of M2 macrophages compared to a reference level has a decreased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition.

[0062] Also provided herein is a method for predicting the therapeutic efficacy of a pharmaceutical composition comprising an anti-CCR8 binding agent for a subject in need thereof (e.g., a subject suffering from any of the exemplary cancers described herein), the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells and elevated levels of NK cells compared to reference levels has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells and decreased levels of NK cells compared to reference levels has a decreased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition.

[0063] Also provided herein are methods for predicting the therapeutic efficacy of an anti-CCR8 binding agent and an anti-ILT4 binding agent in a subject in need thereof (e.g., a subject suffering from any of the exemplary cancers described herein), the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to reference levels has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or determining that a subject diagnosed or identified as having cancer and not identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to reference levels has a reduced likelihood of having a positive therapeutic response to treatment with the anti-CCR8 binding agent and an anti-ILT4 binding agent.

[0064] M2 macrophages

[0065] Unlike classically activated macrophages (also known as M1 macrophages), M2 macrophages are alternatively activated. M2 macrophages are immunosuppressive in nature. Unlike inflammatory M1 macrophages, M2 macrophages tend to promote angiogenesis and neovascularization, as well as matrix activation and remodeling, which has a positive impact on cancer progression and a negative impact on patient prognosis. M1 macrophages are characterized by the expression of HLA-DR and CD197, while M2 types highly express CD163, CD209, CD206, CCL2, etc. M2 macrophages are described in more detail in the art. See, for example, Liu et al., Cancer Cell Int, 21:389, 2021; Tiainen et al., Histopathology, 66:873–83, 2015.

[0066] As used herein, the "reference level" of a parameter may refer to the level of the parameter in a healthy patient (e.g., a patient not suffering from an autoimmune disorder or an inflammatory disorder or an allergy) or the level of the parameter in a healthy subject population. For example, the "reference" level of M2 macrophages may refer to the level of M2 macrophages in a healthy patient (e.g., a patient not suffering from an autoimmune disorder or an inflammatory disorder or an allergy) or the level of M2 macrophages in a healthy subject population. In some cases, the reference level of M2 macrophages is the 55% percentile, 60% percentile, 65% percentile, 70% percentile, 75% percentile, 80% percentile, 85% percentile, 90% percentile, or 95% percentile of the median level of M2 macrophage quantity in a healthy patient population.

[0067] Treg cells

[0068] Regulatory T cells (Treg or Treg cells) are an important mechanism for regulating immune system homeostasis and immune tolerance, and play a vital role in tumor immunoregulation. Treg cells can inhibit CD4 + Helper T cells and CD8 + Activation and differentiation of cytotoxic T cells, thereby inducing responses against self- and tumor-expressed antigens. In the tumor microenvironment (TME), Treg cells can be induced to differentiate by conventional T cells and have strong immunosuppressive functions, inhibiting anti-tumor immunity and promoting the occurrence and development of tumors. Treg cells can also inhibit the function of immune effector cells through a variety of mechanisms and are a key factor in tumor immune escape. Forkhead / winged helix transcription factor (Foxp3) is specifically expressed in Treg cells, CD4 + CD25 + Foxp3 +It is currently considered to be a classic combination marker of Treg cells. In fact, in addition to its ability to mark Treg cells, Foxp3 also mainly controls the function of Treg cells. Only its continuous expression can ensure the maintenance of complete Treg cell inhibitory ability. Treg cells are described in more detail in the art. See, for example, Li et al., Mol Cancer, 19:116, 2020; Nishikawa and Sakaguchi, Curr Opin Immunol, 27:1–7, 2014.

[0069] As used herein, the "reference level" of Treg cells can refer to the level of Treg cells in healthy patients (e.g., patients not suffering from autoimmune disorders or inflammatory disorders or allergies) or the level of Treg cells in healthy subject populations. In some cases, the reference level of Treg cells is the 55% percentile, 60% percentile, 65% percentile, 70% percentile, 75% percentile, 80% percentile, 85% percentile, 90% percentile, or 95% percentile of the median level of Treg cell number in healthy patient populations.

[0070] NK cells

[0071] Natural killer (NK) cells are lymphocytes that have the innate ability to lyse tumor cells without prior sensitization. NK cells can trigger target cell death by releasing cytotoxic granules containing granzymes and perforins and through death receptor-mediated pathways (e.g., FasL / Fas). NK cells also play an immunomodulatory function by secreting chemokines and cytokines (such as RANTES and IFN-γ). In humans, the traditional identification method of NK cells is to determine the absence of CD3 on their surface and the presence of CD56 by flow cytometry. NK cells are described in more detail in the art. See, for example, Liu et al., J Hematol Oncol, 14:7, 2021; Freud et al., Immunity, 47(5):820–833, 2017; Spits et al., Nat Rev Immunol, 13(2):145–9, 2013. NK cells also have Fc receptors, which are very important for binding the Fc region of therapeutic antibodies, leading to NK cell activation and degranulation for effective target cell killing.

[0072] As used herein, the "reference level" of NK cells can refer to the level of NK cells in healthy patients (e.g., patients not suffering from autoimmune disorders or inflammatory disorders or allergies) or the level of NK cells in healthy subject populations. In some cases, the reference level of NK cells is the 55% percentile, 60% percentile, 65% percentile, 70% percentile, 75% percentile, 80% percentile, 85% percentile, 90% percentile, or 95% percentile of the median level of NK cell count in healthy patient populations.

[0073] Immune checkpoint inhibitors

[0074] ICI is a new type of immunotherapy drug that improves the treatment of various cancers (such as metastatic melanoma, non-small lung cancer or renal cell carcinoma). ICI may include humanized antibodies and human monoclonal antibodies targeting inhibitory receptors (e.g., CTLA-4, PD-1, LAG-3, TIM-3) and ligands (e.g., PD-L1) expressed on T lymphocytes, antigen presenting cells and tumor cells, and induce anti-tumor responses by stimulating the immune system. For example, the ICI described herein can be an anti-PD-1 antibody or an antigen binding fragment thereof, an anti-PD-L1 antibody or an antigen binding fragment thereof, and / or an anti-CTLA-4 antibody or an antigen binding fragment thereof.

[0075] Table 1 provides an overview of different ICIs, including corresponding immune checkpoints, targeted indications (e.g., tumor types targeted by these ICIs), and clinical trials (if any). Additional details about ICIs can be found in the art. See, for example, Franzin et al., Front Immunol, 11:574-271, 2020; Johnson et al., Nat Rev Clin Oncol, 19:254-267, 2022.

[0076] Table 1. Overview of major ICIs, targeted tumors, and clinical trials

[0077]

[0078]

[0079]

[0080]

[0081]

[0082]

[0083] Anti-CCR8 binders

[0084] The anti-CCR8 binding agent can be an agent that specifically binds to chemokine (CC motif) receptor 8 (CCR8). For example, the anti-CCR8 binding agent can be an anti-CCR8 antibody or a fragment thereof. As used herein, the term "antibody" is referred to in the broadest sense and specifically covers various embodiments, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies formed by at least two complete antibodies) and antibody fragments (such as, diabodies), as long as they exhibit the desired biological activity. Antibodies are mainly amino acid-based molecules, but can also contain one or more modifications (including but not limited to the addition of one or more detectable labels). In some embodiments, the anti-CCR8 binding agent (e.g., anti-CCR8 antibody) can be a depletion antibody. In certain embodiments, the depletion antibody can have the ability to block ligand binding to CCR8. In some cases, non-fucosylated anti-CCR8 antibodies can be used, which have increased antibody-dependent cellular cytotoxicity (ADCC) activity and are therefore good depletion products for Tregs expressing CCR8.

[0085] In some cases, BMS-986340 can be used as an anti-CCR8 binding agent (e.g., an anti-CCR8 antibody) in the methods of the present disclosure. BMS-986340 is a newly developed anti-CCR8 mAb that can reduce the abundance of CCR8 in human tumor explants. + Treg. Other anti-CCR8 antibodies for use in the methods of the present disclosure are described in the art. See, e.g., Campbell et al., Cancer Res, 81(11):2983–2994, 2021.

[0086] Anti-ILT4 binders

[0087] An anti-ILT4 binder can be an agent that specifically binds to the inhibitory immune checkpoint receptor immunoglobulin-like transcript 4 (ILT4; leukocyte immunoglobulin-like receptor subfamily B member 2; LILRB2; lymphocyte immunoglobulin-like receptor 2; LIR2; monocyte / macrophage immunoglobulin-like receptor 10; MIR-10; CD85d). For example, an anti-ILT4 binder can be an anti-ILT4 antibody or a fragment thereof. In some embodiments, an anti-ILT4 binder (e.g., an anti-ILT4 antibody) can be a blocking antibody. In certain embodiments, a blocking antibody can inhibit the interaction of ILT4 (LILRB2) with its ligands, including HLA-G.

[0088] In some cases, MK-4830 can be used as an anti-ILT4 binder (e.g., an anti-ILT4 antibody) in the methods of the present disclosure. MK-4830 is a human monoclonal antibody against ILT4 with potential immunomodulatory and anti-tumor activity. Other anti-ILT4 antibodies for use in the methods of the present disclosure are described in the art. See, e.g., Chen et al., Theranostics 11(7):3392–3416, 2021.

[0089] Treatment

[0090] Provided herein are methods for treating a subject having cancer (e.g., any exemplary cancer described herein), the methods comprising administering a therapeutically effective amount of an immune checkpoint inhibitor to a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having increased levels of M2 macrophages as compared to a reference level (e.g., any exemplary reference level described herein).

[0091] Also provided herein is a method of treating a subject having cancer (e.g., any exemplary cancer described herein), the method comprising administering a therapeutically effective amount of an anti-ILT4 binding agent to a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having elevated levels of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein).

[0092] Also provided herein is a method for treating a subject having cancer (e.g., any of the exemplary cancers described herein), the method comprising administering a therapeutically effective amount of an anti-CCR8 binding agent to a subject previously identified or diagnosed as having cancer (e.g., any of the exemplary cancers described herein) and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level (e.g., any of the exemplary reference levels described herein).

[0093] Also provided herein is a method of treating a subject having cancer (e.g., any exemplary cancer described herein), the method comprising administering a therapeutically effective amount of an anti-CCR8 binding agent and a therapeutically effective amount of an anti-ILT4 binding agent to a subject previously identified or diagnosed as having cancer and previously identified as having increased levels of Treg cells, increased levels of NK cells, and increased numbers of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein).

[0094] As used herein, "treat" or "treating" refers to alleviating, ameliorating, improving, relieving, delaying the onset of, inhibiting the progression of, reducing the severity of, and / or reducing the incidence of one or more symptoms or characteristics of a particular disease, disorder, and / or condition. For example, "treating" cancer may refer to prolonging the survival of a subject diagnosed or identified as having cancer. Additionally or alternatively, "treating" cancer may refer to inhibiting the growth and / or spread of a cancer or tumor.

[0095] As used herein, the term "subject" refers to any organism to which the disclosed methods can be applied, such as those used for experimental and / or therapeutic purposes. Typical subjects include animals, such as mammals (e.g., mice, rats, rabbits, dogs, pigs, non-human primates, and humans). In some cases, a "subject" may refer to a cancer patient, such as an individual diagnosed and / or identified as suffering from cancer.

[0096] The cancer useful for the present treatment method may include any malignant cell type, such as those found in solid tumors or hematological tumors. Exemplary solid tumors may include, but are not limited to, tumors of organs selected from the following groups: pancreas, colon, cecum, stomach, brain, head, neck, ovary, kidney, larynx, sarcoma, lung (including small cell lung cancer, non-small cell lung cancer, lung adenocarcinoma, lung squamous cell carcinoma), bladder, melanoma, prostate and breast. Exemplary hematological tumors include myeloid tumors, T or B cell malignancies, leukemia, lymphoma, blastoma, myeloma, etc. Other examples of cancers that can be treated using the methods provided herein include, but are not limited to, lung cancer (including small cell lung cancer, non-small cell lung cancer, lung adenocarcinoma and lung squamous cell carcinoma), peritoneal cancer, gastric cancer (including gastrointestinal cancer and gastrointestinal stromal cancer), pancreatic cancer, cervical cancer, ovarian cancer, liver cancer, bladder cancer, breast cancer, colon cancer, colorectal cancer, endometrial cancer or uterine cancer, salivary gland cancer, kidney cancer, prostate cancer, vulvar cancer, thyroid cancer, various head and neck cancers and melanoma.

[0097] In certain embodiments, the cancer treated according to the methods described herein includes, but is not limited to, prostate cancer, breast cancer, lung cancer, colorectal cancer, melanoma, bronchial cancer, bladder cancer, brain cancer or central nervous system cancer, peripheral nervous system cancer, uterine or endometrial cancer, oral or pharyngeal cancer, non-Hodgkin's lymphoma, thyroid cancer, kidney cancer, biliary tract cancer, small intestine or appendix cancer, salivary gland cancer, thyroid cancer, adrenal cancer, squamous cell carcinoma, mesothelioma, bone cancer, thymoma / thymic cancer, glioblastoma, myelodysplastic syndrome, soft tissue sarcoma, DIPG, adenocarcinoma, osteosarcoma, chondrosarcoma, leukemia, or pancreatic cancer. In some embodiments, the cancer treated according to the methods described herein includes carcinoma (e.g., adenocarcinoma), lymphoma, blastoma, melanoma, sarcoma, or leukemia. In certain embodiments, the cancer treated according to the methods described herein comprises squamous cell carcinoma, small cell lung cancer, non-small cell lung cancer, gastrointestinal cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, pancreatic cancer, glioblastoma, glioma, cervical cancer, ovarian cancer, liver cancer (e.g., hepatic carcinoma and hepatocellular carcinoma), bladder cancer, breast cancer, inflammatory breast cancer, Merkel cell carcinoma, colon cancer, colorectal cancer, gastric cancer, bladder cancer, endometrial cancer, myeloma (e.g., multiple myeloma), salivary gland cancer, kidney cancer (e.g., renal cell carcinoma and Wilms tumor), basal cell carcinoma, melanoma, prostate cancer, vulvar cancer, thyroid cancer, testicular cancer, esophageal cancer, serous adenocarcinoma, or various types of head and neck cancer. In certain embodiments, the cancer treated according to the methods described herein comprises desmoplastic melanoma, inflammatory breast cancer, thymoma, rectal cancer, anal cancer, or operable or inoperable brain stem glioma. In some instances, the cancer treated according to the methods described herein comprises adrenocortical carcinoma, bladder urothelial carcinoma, glioblastoma multiforme, low-grade glioma, cervical and endocervical cancer, colon adenocarcinoma, colorectal adenocarcinoma, esophageal cancer, head and neck squamous cell carcinoma, renal clear cell carcinoma, renal papillary cell carcinoma, liver hepatocellular carcinoma, lung adenocarcinoma, lung adenocarcinoma, lung squamous cell carcinoma, small cell lung cancer, ovarian serous adenocarcinoma, pancreatic cancer, prostate cancer, skin melanoma, gastric adenocarcinoma, thyroid cancer, and / or uterine corpus endometrial carcinoma.

[0098] Methods for selecting treatment for a subject

[0099] Also provided herein are methods of selecting a pharmaceutical composition comprising an immune checkpoint inhibitor for a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having reduced levels of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein).

[0100] Also provided herein are methods of selecting a pharmaceutical composition comprising an anti-ILT4 binding agent for a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having elevated levels of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein).

[0101] Also provided herein are methods for selecting a pharmaceutical composition comprising an anti-CCR8 binding agent for a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level (e.g., any exemplary reference level described herein).

[0102] Also provided herein are methods for selecting an anti-CCR8 binding agent and an anti-ILT4 binding agent for a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having increased levels of Treg cells, increased levels of NK cells, and increased levels of NK cells compared to a reference level (e.g., any exemplary reference level described herein).

[0103] Methods for selecting patients for treatment

[0104] Also provided herein are methods of selecting a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having reduced levels of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein) for treatment with a pharmaceutical composition comprising an immune checkpoint inhibitor.

[0105] Also provided herein are methods of selecting a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having elevated levels of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein) for treatment with a pharmaceutical composition comprising an anti-ILT4 binding agent.

[0106] Also provided herein are methods for selecting a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level (e.g., any exemplary reference level described herein) for treatment with a pharmaceutical composition comprising an anti-CCR8 binding agent.

[0107] Also provided herein are methods for selecting a subject previously identified or diagnosed as having cancer (e.g., any exemplary cancer described herein) and previously identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to a reference level (e.g., any exemplary reference level described herein) for treatment with an anti-CCR8 binding agent and an anti-ILT4 binding agent.

[0108] Example

[0109] The present invention is further illustrated by the following examples, which do not limit the scope of the present invention described in the claims.

[0110] Example 1. Conversion of transcriptome data into immune cells

[0111] We wanted to mine bioinformatics insights from existing databases so that data related to past failures could inform future successes in immuno-oncology. To this end, we evaluated survival curves to identify patterns. We analyze cancer omics data and transform genomic data into actionable insights. Existing omics data from large and small public and private databases were analyzed and gene expression was converted into immune cell subtypes. Heatmaps were also evaluated to understand whether there was a response. Specifically, the CIBERSORT algorithm used a leukocyte gene signature matrix (LM22) containing 547 genes to generate gene weights for various cell types. It was then tested on 3k human transcriptomes. The LM22 matrix was used to calculate immune cell enrichment using the open source ADAPTS (Automated Deconvolution Amplification of Profiles for Tissue-Specific Cells) software. See, for example, cran.r-project.org / web / packages / ADAPTS / index.html. In addition, the ranked genes in LM22 were also used for gene set variation analysis (GSVA). The method was validated using sorted cells from patients with early-stage non-small cell lung cancer (NSCLC) and follicular lymphoma. We are able to improve the efficiency of our analytical methods and use these insights to prioritize our portfolio.

[0112] Example 2. The tumor microenvironment limits the benefits of PD-1 inhibitors to patients

[0113] Publicly available data from The Cancer Genome Atlas (TCGA) and additional sources were analyzed to identify patterns associated with patient nonresponse to PD-1 blockade. For example, we analyzed data from Ayers et al., 2017 (J Clin Invest, 127(8):2930-2940) showing progression-free survival (PFS) time and T-cell inflammatory gene expression profile (GEP) score in 244 patients treated with the PD-1 inhibitor KEYTRUDA. A general pattern of lack of objective response was observed in patients with lower tumor gene expression levels, which is thought to represent tumors without a T-cell inflammatory phenotype. Patients with lower tumor T-cell inflammatory GEP scores (e.g., less than -0.3) generally had more rapid disease progression, while patients with higher T-cell inflammatory GEP scores had a longer time to progression. A large proportion of patients were identified to have additional immunosuppressive mechanisms that may interfere with the killing of T cells against tumors. In addition, adverse immunosuppressive mechanisms in the tumor microenvironment (TME) may also contribute to the poor prognosis of these patients. In summary, the analysis revealed novel immunosuppressive mechanisms in the TME.

[0114] Example 3. Converting an unfavorable TME into a TME favorable for anti-PD-1 therapeutic response

[0115] As described in Example 2, our analysis of cancer omics data revealed novel immunosuppressive mechanisms in the TME. Given the role of the TME in limiting the response to PD-1 inhibitor treatment, our goal is to transform an unfavorable TME into a TME that is favorable for anti-PD-1 treatment response. To this end, we used an improved algorithm containing 30-45 genes to convert transcriptomic data from cancer patients into immune cell types. Next, a group of cancer patients with addressable dominant MOA (NSCLC / SCLC, etc.) was identified. Next, first-in-class candidate drugs are developed based on biologically identified targets. This approach will address the main immunosuppressive mechanisms that may solve the problem of "immune escape". In addition, drug combinations will address unfavorable TMEs, which will bring better outcomes for patients.

[0116] Example 4. High levels of immunosuppressive macrophages may be responsible for limited PD(L)-1 responses in SCLC patients

[0117] Canceromics data from patients with small cell lung cancer (SCLC) were reanalyzed to identify patterns associated with limited patient response to PD(L)-1 therapy. Specifically, the survival plot and heat map data from George et al., 2015 (Nature, 524(7563):47-53(2015)) were reanalyzed using the methods described in the previous example. The results are described in Figure 1A and 1B .like Figure 1A and1B As shown, patients with enriched M1 macrophages (i.e., patients with higher levels of M1 macrophages; high M1 (M1 enriched)) showed a higher probability of survival compared with patients with higher levels of immunosuppressive M2 macrophages (high M2 (M2 enriched)). Profile analysis of resting dendritic cells (DCs) and elevated levels of M2 macrophages showed that approximately 30% of SCLC patients had a high M2 (i.e., immunosuppressive macrophage) signature and a poor prognosis. Therefore, our analysis suggests that high levels of immunosuppressive macrophages within SCLC may be responsible for the limited PD(L)-1 response in SCLC patients.

[0118] Example 5. ILT4 expression is higher and immune T cells are lower in SCLC patients with high M2

[0119] Furthermore, a reanalysis of the SCLC patient data showed that ILT4 expression was significantly higher and immune T cells were lower in SCLC patients with high M2 (i.e., high levels of immunosuppressive M2 macrophages). The results are described in Figure 2A and 2B Specifically, it was found that ILT4 expression in M2 samples was significantly higher than that in non-M2 samples ( Figure 2A ). In addition, it was found that there were fewer immune T cells in the M2-enriched samples ( Figure 2B ).

[0120] Example 6. NSCLC patients with high levels of M2 macrophages may not respond to PD(L)-1 therapy

[0121] Next, we reanalyzed the published data on patients with non-small cell lung cancer (NSCLC) to identify patterns associated with limited response to PD(L)-1 therapy in NSCLC patients. Specifically, we reanalyzed the published data from The Cancer Genome Atlas using the methods described in the previous examples. The results are described in Figure 3 .like Figure 3 As shown in Figure 2, approximately 30% of patients have high expression of immunosuppressive M2 macrophages. Therefore, our analysis suggests that NSCLC patients with high levels of M2 macrophages may not respond to PD(L)-1 therapy.

[0122] The lessons learned from the above examples allowed us to rationally develop anti-ILT4 antibodies with the goal of repolarizing M2 (immunosuppressive) macrophages to M1 (inflammatory) macrophages. Figure 4 Depicted is a schematic diagram showing that anti-ILT4 antibodies repolarize M2 (suppressive) macrophages to M1 (inflammatory) macrophages.

[0123] Example 7. NSCLC patients with Treg characteristics may benefit from the combination therapy of depleting anti-CCR8 antibody and toripalimab

[0124] Reanalysis of the NSCLC patient data also showed that approximately 30% of NSCLC patients had a high Treg signature and that these patients could benefit from the combination of a depleting anti-CCR8 antibody with toripalimab. The results are described in Figure 5 .like Figure 5 As shown in Figure 1, high levels of regulatory T cells (Treg) were found in patients who were unlikely to benefit from PD-1 therapy. Among them, some patients had both high Treg and high NK cells (group 1; Treg 高 NK 高 ), while other patients had high Treg and low NK cells (group 2; Treg 高 NK 低 In general, the high Treg population appears to be distinct from the high M2 population (described in the previous example). Since there are fewer NK cells, anti-CCR-8 is less likely to make Treg 高 NK 低 Patients with high levels of Tregs and NK cells (i.e., CCRR-8 therapy can benefit these patients, but only when NK cells are present). 高 NK 高 Patients with leukemia and renal cell carcinoma may benefit greatly from the combination of toripalimab and anti-CCR8 or similar combination treatment regimens.

[0125] This analysis suggests that depletion of immunosuppressive Tregs allows cytotoxic T cells to effectively kill tumors. Specifically, the presence of Treg cells may affect the efficacy of anti-PD(L)1, so anti-CCR8 and effector NK cells are required to effectively eliminate Tregs. Therefore, CCR8 antibodies can effectively kill tumors in the presence of NK cells. Figure 6 A schematic diagram of the concept is shown.

[0126] Example 8. Patient Populations Who Would Benefit from Combination Therapy

[0127] Re-analysis of the patient data (as described in the previous examples) identified patient populations that may benefit from the above combination therapy. Specifically, our analysis identified the Treg Hi effector Hi population and the M2 enriched population as the patient populations that would benefit most from a therapy comprising ILT4 and CCR8 in combination with toripalimab. The results are described in Figure 7 and Table 2 below. Figure 7The graphs in show the percentage of patients in two defined populations, the “Treg Hi, Effector Hi” population and the “M2-enriched” population, that may benefit from combination therapy. Data are shown for hepatocellular carcinoma (HCC), NSCLC adenocarcinoma, SCLC, head and neck cancer, and esophageal cancer. SCLC M1 and M2 values ​​are based on ADAPTS:LM22 profiles. Treg patterns are derived from GSVA, which uses the ranked LM22 profiles. Given the limited overlap between these two patient classes, CCR8 and anti-ILT4 antibody therapy may have the potential to benefit a large proportion of patients with tumors.

[0128] Table 2. Patient groups that may benefit from combination therapy

[0129]

[0130]

[0131] Other Implementations

[0132] It should be understood that although the invention has been described in conjunction with specific embodiments, the foregoing description is intended to illustrate rather than limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages and improvements are within the scope of the claims.

Claims

1. A method for treating a subject suffering from cancer, wherein the method comprises: A therapeutically effective amount of an immune checkpoint inhibitor is administered to a subject previously identified or diagnosed as having cancer and previously identified as having reduced levels of M2 macrophages compared to a reference level.

2. A method of selecting a pharmaceutical composition comprising an immune checkpoint inhibitor for a subject previously identified or diagnosed as having cancer and previously identified as having reduced levels of M2 macrophages compared to a reference level.

3. A method of selecting a subject previously identified or diagnosed as having cancer and previously identified as having decreased levels of M2 macrophages compared to a reference level for treatment with a pharmaceutical composition comprising an immune checkpoint inhibitor.

4. A method for predicting the therapeutic efficacy of a pharmaceutical composition comprising an immune checkpoint inhibitor on a subject in need thereof, the method comprising: determining that a subject diagnosed or identified as having cancer and identified as having a reduced level of M2 macrophages compared to a reference level has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or It is determined that a subject diagnosed or identified as having cancer and identified as having an elevated level of M2 macrophages compared to a reference level has a reduced likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition.

5. The method according to any one of claims 1-4, wherein the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof.

6. The method of any one of claims 1-4, wherein the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

7. The method according to any one of claims 1 to 6, wherein the method further comprises: determining the level of M2 macrophages in a sample obtained from the subject; as well as The identified subjects had reduced levels of M2 macrophages compared to reference levels.

8. A method of treating a subject suffering from cancer, wherein the method comprises: A therapeutically effective amount of an anti-ILT4 binding agent is administered to a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of M2 macrophages compared to a reference level.

9. A method of selecting a pharmaceutical composition comprising an anti-ILT4 binding agent for a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of M2 macrophages compared to a reference level.

10. A method of selecting a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of M2 macrophages compared to a reference level for treatment with a pharmaceutical composition comprising an anti-ILT4 binding agent.

11. The method of any one of claims 8-10, wherein the anti-ILT4 binding agent is an anti-ILT4 antibody or an antigen-binding fragment thereof.

12. The method of any one of claims 8-11, wherein the method further comprises administering an immune checkpoint inhibitor to the subject.

13. The method of claim 12, wherein the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof.

14. The method of claim 12, wherein the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

15. The method according to any one of claims 8 to 14, wherein the method further comprises: determining the level of M2 macrophages in a sample obtained from the subject; as well as The identified subjects had increased levels of M2 macrophages compared to reference levels.

16. The method according to any one of claims 1-15, wherein the M2 macrophages have an increased ILT4 expression level compared to a reference level.

17. The method according to any one of claims 1-16, wherein the reference level of M2 macrophages is the level of M2 macrophages in a population of healthy subjects.

18. The method according to any one of claims 1-16, wherein the reference level of M2 macrophages is 75% of the median level of the number of M2 macrophages in a healthy patient population.

19. The method of any one of claims 1-16, wherein the reference level of M2 macrophages is 80% of the median level of M2 macrophages in a healthy patient population.

20. The method of any one of claims 1-16, wherein the reference level of M2 macrophages is 85% of the median level of M2 macrophages in a healthy patient population.

21. The method of any one of claims 1-16, wherein the reference level of M2 macrophages is 90% of the median level of M2 macrophages in a healthy patient population.

22. The method of any one of claims 1-16, wherein the reference level of M2 macrophages is 95% of the median level of M2 macrophages in a healthy patient population.

23. The method of any one of claims 1-22, wherein the cancer is a solid tumor.

24. The method of claim 23, wherein the solid tumor is non-small cell lung cancer, hepatocellular carcinoma, small cell lung cancer, head and neck cancer, or esophageal cancer.

25. The method of any one of claims 1-22, wherein the cancer is a hematological cancer.

26. A method of treating a subject suffering from cancer, wherein the method comprises: A therapeutically effective amount of an anti-CCR8 binding agent is administered to a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level.

27. A method of selecting a pharmaceutical composition comprising an anti-CCR8 binding agent for a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level.

28. A method of selecting a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level for treatment with a pharmaceutical composition comprising an anti-CCR8 binding agent.

29. A method for predicting the therapeutic efficacy of a pharmaceutical composition comprising an anti-CCR8 binding agent in a subject in need thereof, the method comprising: Determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells and elevated levels of NK cells compared to a reference level has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or It is determined that a subject diagnosed or identified as having cancer and identified as having increased levels of Treg cells and decreased levels of NK cells compared to a reference level has a reduced likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition.

30. The method of any one of claims 26-29, wherein the anti-CCR8 binding agent is an anti-CCR8 antibody or an antigen-binding fragment thereof.

31. The method of any one of claims 26-30, wherein the method further comprises administering an immune checkpoint inhibitor to the subject.

32. The method of claim 31, wherein the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof.

33. The method of claim 31, wherein the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

34. The method according to any one of claims 26-33, wherein the method further comprises: determining the levels of Treg cells and NK cells in a sample obtained from the subject; as well as The identified subjects had increased levels of Treg cells and NK cells compared to reference levels.

35. The method according to any one of claims 26-34, wherein the reference level of Treg cells is the level of Treg cells in a population of healthy subjects.

36. The method according to any one of claims 26-34, wherein the reference level of Treg cells is 75% of the median level of Treg cell number in a healthy patient population.

37. The method of any one of claims 26-34, wherein the reference level of Treg cells is 80% of the median level of Treg cells in a healthy patient population.

38. The method of any one of claims 26-34, wherein the reference level of Treg cells is 85% of the median level of Treg cells in a healthy patient population.

39. The method of any one of claims 26-34, wherein the reference level of Treg cells is 90% of the median level of Treg cells in a healthy patient population.

40. The method of any one of claims 26-34, wherein the reference level of Treg cells is 95% of the median level of Treg cells in a healthy patient population.

41. The method of any one of claims 26-40, wherein the reference level of NK cells is the level of NK cells in a population of healthy subjects.

42. The method of any one of claims 26-40, wherein the reference level of NK cells is 75% of the median level of NK cell number in a healthy patient population.

43. The method of any one of claims 26-40, wherein the reference level of NK cells is 80% of the median level of NK cells in a healthy patient population.

44. The method of any one of claims 26-40, wherein the reference level of NK cells is 85% of the median level of NK cells in a healthy patient population.

45. The method of any one of claims 26-40, wherein the reference level of NK cells is 90% of the median level of NK cells in a healthy patient population.

46. ​​The method of any one of claims 26-40, wherein the reference level of NK cells is 95% of the median level of NK cells in a healthy patient population.

47. The method of any one of claims 26-46, wherein the cancer is a solid tumor.

48. The method of claim 47, wherein the solid tumor is non-small cell lung cancer, hepatocellular carcinoma, small cell lung cancer, head and neck cancer, or esophageal cancer.

49. The method of any one of claims 26-46, wherein the cancer is a hematological cancer.

50. A method of treating a subject suffering from cancer, wherein the method comprises: A therapeutically effective amount of an anti-CCR8 binding agent and a therapeutically effective amount of an anti-ILT4 binding agent are administered to a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated numbers of M2 macrophages compared to a reference level.

51. A method of selecting an anti-CCR8 binding agent and an anti-ILT4 binding agent for a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of NK cells compared to a reference level.

52. A method of selecting a subject previously identified or diagnosed as having cancer and previously identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to a reference level for treatment with an anti-CCR8 binding agent and an anti-ILT4 binding agent.

53. A method for predicting the therapeutic efficacy of an anti-CCR8 binding agent and an anti-ILT4 binding agent in a subject in need thereof, the method comprising: Determining that a subject diagnosed or identified as having cancer and identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to a reference level has an increased likelihood of having a positive therapeutic response to treatment with the pharmaceutical composition; or It is determined that subjects diagnosed or identified as having cancer and not identified as having elevated levels of Treg cells, elevated levels of NK cells, and elevated levels of M2 macrophages compared to reference levels have a reduced likelihood of having a positive therapeutic response to treatment with an anti-CCR8 binder and an anti-ITL4 binder.

54. The method of any one of claims 50-53, wherein the anti-CCR8 binding agent is an anti-CCR8 antibody or an antigen-binding fragment thereof.

55. The method of any one of claims 50-54, wherein the anti-ILT4 binding agent is an anti-ILT4 antibody or an antigen-binding fragment thereof.

56. The method of any one of claims 50-55, wherein the method further comprises administering an immune checkpoint inhibitor to the subject.

57. The method of claim 56, wherein the immune checkpoint inhibitor is an anti-PD-1 antibody or an antigen-binding fragment thereof.

58. The method of claim 56, wherein the immune checkpoint inhibitor is an anti-PD-L1 antibody or an antigen-binding fragment thereof.

59. The method according to any one of claims 50-58, wherein the method further comprises: determining the levels of Treg cells, NK cells, and M2 macrophages in a sample obtained from the subject; as well as The subject is identified as having increased levels of Treg cells, increased levels of NK cells, and increased levels of M2 macrophages compared to a reference level.

60. The method of any one of claims 50-59, wherein the reference level of Treg cells is the level of Treg cells in a population of healthy subjects.

61. The method of any one of claims 50-59, wherein the reference level of Treg cells is 75% of the median level of Treg cell number in a healthy patient population.

62. The method of any one of claims 50-59, wherein the reference level of Treg cells is 80% of the median level of Treg cells in a healthy patient population.

63. The method of any one of claims 50-59, wherein the reference level of Treg cells is 85% of the median level of Treg cells in a healthy patient population.

64. The method of any one of claims 50-59, wherein the reference level of Treg cells is 90% of the median level of Treg cells in a healthy patient population.

65. The method of any one of claims 50-59, wherein the reference level of Treg cells is 95% of the median level of Treg cells in a healthy patient population.

66. The method of any one of claims 50-65, wherein the reference level of NK cells is the level of NK cells in a population of healthy subjects.

67. The method of any one of claims 50-65, wherein the reference level of NK cells is 75% of the median level of NK cell number in a healthy patient population.

68. The method of any one of claims 50-65, wherein the reference level of NK cells is 80% of the median level of NK cells in a healthy patient population.

69. The method of any one of claims 50-65, wherein the reference level of NK cells is 85% of the median level of NK cells in a healthy patient population.

70. The method of any one of claims 50-65, wherein the reference level of NK cells is 90% of the median level of NK cells in a healthy patient population.

71. The method of any one of claims 50-65, wherein the reference level of NK cells is 95% of the median level of NK cells in a healthy patient population.

72. The method of any one of claims 50-71, wherein the reference level of M2 macrophages is the level of M2 macrophages in a population of healthy subjects.

73. The method of any one of claims 50-71, wherein the reference level of M2 macrophages is 75% of the median level of the number of M2 macrophages in a healthy patient population.

74. The method of any one of claims 50-71, wherein the reference level of M2 macrophages is 80% of the median level of M2 macrophages in a healthy patient population.

75. The method of any one of claims 50-71, wherein the reference level of M2 macrophages is 85% of the median level of M2 macrophages in a healthy patient population.

76. The method of any one of claims 50-71, wherein the reference level of M2 macrophages is 90% of the median level of M2 macrophages in a healthy patient population.

77. The method of any one of claims 50-71, wherein the reference level of M2 macrophages is 95% of the median level of M2 macrophages in a healthy patient population.

78. The method of any one of claims 50-77, wherein the cancer is a solid tumor.

79. The method of claim 78, wherein the solid tumor is non-small cell lung cancer, hepatocellular carcinoma, small cell lung cancer, head and neck cancer, or esophageal cancer.

80. The method of any one of claims 50-77, wherein the cancer is a hematological cancer.

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