STING agonist combination therapy with cytokines

Abstract

The present disclosure provides, inter alia, methods and uses for treating a disease or disorder, particularly a tumor in a cancer patient, comprising co-administering to the patient effective amounts of a STING agonist, a cytokine, and optionally an immune checkpoint inhibitor, wherein the STING agonist or the cytokine is administered intratumorally to the patient.

Description

[Technical field]

[0001] 1. Sequence Listing This application is filed through the USPTO Patent Center and includes a sequence listing containing three sequences, which is incorporated herein by reference in its entirety. The XML copy, created on July 18, 2022, is named "39143-52882 008WO_Sequence Listing.xml" and is 14 kilobytes in size.

[0002] 2. Field The present disclosure relates, inter alia, to the use of agonists of STimulator of Interferon Genes (STING) in combination with cytokines to activate the immune system to treat certain diseases or disorders, including cancer. The present disclosure also relates to the use of STING agonists (such as cyclic dinucleotides), cytokines (such as interleukins), and immune checkpoint inhibitors to treat certain diseases or disorders, including cancer. [Background technology]

[0003] 3.Background Treatment of advanced solid tumor malignancies and many hematological malignancies continues to be defined by a high unmet medical need. In most situations, treatment with cytotoxic chemotherapy and targeted kinase inhibitors leads to the emergence of drug-resistant tumor clones and subsequent tumor progression and metastasis.

[0004] In recent years, notable successes have been achieved through alternative approaches centered on the activation of immune-mediated tumor destruction. The immune system plays a pivotal role in defending humans and animals against cancer. Antitumor efficacy is controlled by positive factors that activate antitumor immunity and negative factors that inhibit the immune system. Negative factors that inhibit antitumor immunity include immune checkpoint proteins such as cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), programmed cell death 1 (PD-1), and programmed death ligand 1 (PD-L1). Immuno-oncology (IO) approaches involving antibodies against these checkpoint proteins have shown remarkable efficacy in several types of human cancers.

[0005] However, existing cancer immunotherapies via immune checkpoint blockade are effective in only a small proportion of cancer patients (on average 20–30%). Patients who are refractory to immune checkpoint blockade often have non-inflamed tumors, or so-called "cold" tumor cells, i.e., they lack tumor-infiltrating leukocytes (TILs), e.g., cluster of differentiation 8 (CD8) T cells, or the tumor microenvironment suppresses the function of TILs. The main thrust of ongoing cancer drug development research remains focused on converting "cold" tumor cells into "hot" tumor cells to achieve better tumor control across a broader range of patients.

[0006] The innate immune system, the first line of defense against pathogens and cancer cells, is critical for transforming a non-inflammatory tumor ("cold") into an inflammatory ("hot") microenvironment. A recently discovered innate immune pathway, the cGAS-STING pathway, involving the protein cyclic GMP-AMP synthase (cGAS), plays a key role in anti-tumor immunity. cGAS is a DNA-sensing enzyme that activates the type I interferon pathway. Upon binding to DNA, cGAS is activated to synthesize the cyclic dinucleotide (CDN) 2'3'-cyclic GMP-AMP (2'3'-cGAMP), which then functions as a secondary messenger that binds to and activates the adaptor protein STING. STING then activates a signaling cascade that leads to the production of type I interferons, cytokines and other immune mediators.

[0007] Although cytokine production is essential to generate antitumor immunity, high cytokine levels pose safety concerns. Specifically, high cytokine levels can induce dangerous inflammatory responses in cancer patients undergoing immunotherapy, thereby hindering the use of cytokines in IO applications. Selection of the appropriate cytokine type and amount to exploit its antitumor effects while reducing or limiting its systemic toxicity remains a challenging unmet need. As a result, there remains a significant unmet medical need that requires the development of therapies that can trigger specific and systemic immune responses against tumors throughout the body, including tumors that are not or cannot be directly treated (i.e., via the abscopal effect), for example, due to the location or size of the tumor. Summary of the Invention

[0008] 4. Overview The present disclosure provides methods of administering a STING agonist to a patient, e.g., a human cancer patient, in combination with a cytokine, optionally further in combination with one or more immune checkpoint inhibitors, e.g., inhibitors of CTLA-4, PD-1 and / or PD-L1, particularly antibody inhibitors of these proteins. The present disclosure also provides combination therapies that can be used in such methods and treatments.

[0009] In one aspect, the present disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, wherein the STING agonist or the cytokine is administered intratumorally to the patient. In certain embodiments, both the STING agonist and the cytokine are administered intratumorally to the patient. In a particular aspect, the present disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, wherein the STING agonist or the cytokine is administered intratumorally to the patient, wherein the patient shows reduced tumor recurrence after treatment, including in the absence of further treatment. In certain of these embodiments, both the STING agonist and the cytokine are administered intratumorally to the patient.

[0010] In certain embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising systemically co-administering to the cancer patient an effective amount of an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody).

[0011] In other specific embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist (e.g., a CDN), a cytokine (e.g., an interleukin), and an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody), wherein both the STING agonist and the cytokine are administered intratumorally to the patient and the immune checkpoint inhibitor is administered systemically to the cancer patient.

[0012] In another aspect, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein the STING agonist or the cytokine is administered intratumorally to the patient, hi certain embodiments, both the STING agonist and the cytokine are administered intratumorally to the patient.

[0013] In certain embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising systemically co-administering to the cancer patient an effective amount of an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody).

[0014] In other specific embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, the method comprising co-administering to the patient effective amounts of a STING agonist (e.g., a CDN), a cytokine (e.g., an interleukin), and an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody), wherein both the STING agonist and the cytokine are administered intratumorally to the patient and the immune checkpoint inhibitor is administered systemically to the cancer patient.

[0015] In yet another aspect, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells or macrophages, or a combination thereof) in a cancer patient, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, wherein the STING agonist or cytokine is administered intratumorally to the patient. In certain embodiments, both the STING agonist and the cytokine are administered intratumorally to the patient.

[0016] In certain embodiments, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells or macrophages, or a combination thereof) in a cancer patient, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering an effective amount of an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody) globally to the cancer patient.

[0017] In other specific embodiments, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof) in a cancer patient, comprising co-administering to the patient effective amounts of a STING agonist (e.g., CDN) and a cytokine (e.g., an interleukin), and an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody), wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered systemically to the cancer patient.

[0018] In yet another aspect, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein the STING agonist or the cytokine is administered intratumorally to the patient, hi certain embodiments, both the STING agonist and the cytokine are administered intratumorally to the patient.

[0019] In certain embodiments, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising systemically co-administering to the cancer patient an effective amount of an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody).

[0020] In other specific embodiments, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist (e.g., a CDN), a cytokine (e.g., an interleukin), and an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody), wherein both the STING agonist and the cytokine are administered intratumorally to the patient and the immune checkpoint inhibitor is administered systemically to the cancer patient.

[0021] In another aspect, the present disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising having a STING agonist (e.g., CDN), a cytokine (e.g., interleukin), and an immune checkpoint inhibitor (e.g., anti-PD-1 antibody, anti-PD-L1 antibody, or anti-CTLA-4 antibody) present in the patient's body simultaneously. In a particular embodiment, the method comprises administering an effective amount of a STING agonist to the patient, where the patient has already been administered the cytokine and the immune checkpoint inhibitor. In another particular embodiment, the method comprises administering an effective amount of a cytokine to the patient, where the patient has already been administered the STING agonist and the immune checkpoint inhibitor. In yet another particular embodiment, the method comprises administering an effective amount of an immune checkpoint inhibitor to the patient, where the patient has already been administered the STING agonist and the cytokine.

[0022] In another aspect, the present disclosure provides a method of reducing tumor recurrence in a patient, comprising simultaneously administering to the patient a STING agonist (e.g., CDN), a cytokine (e.g., interleukin), and an immune checkpoint inhibitor (e.g., anti-PD-1 antibody, anti-PD-L1 antibody, or anti-CTLA-4 antibody). In a particular embodiment, the method comprises administering an effective amount of a STING agonist to the patient, where the patient has already been administered a cytokine and an immune checkpoint inhibitor. In another particular embodiment, the method comprises administering an effective amount of a cytokine to the patient, where the patient has already been administered a STING agonist and an immune checkpoint inhibitor. In yet another particular embodiment, the method comprises administering an effective amount of an immune checkpoint inhibitor to the patient, where the patient has already been administered a STING agonist and a cytokine.

[0023] In another aspect, the present disclosure provides a method of preventing tumor recurrence in a patient, comprising simultaneously administering to the patient a STING agonist (e.g., CDN), a cytokine (e.g., an interleukin), and an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody). In a particular embodiment, the method comprises administering an effective amount of a STING agonist to the patient, where the patient has already been administered a cytokine and an immune checkpoint inhibitor. In another particular embodiment, the method comprises administering an effective amount of a cytokine to the patient, where the patient has already been administered a STING agonist and an immune checkpoint inhibitor. In yet another particular embodiment, the method comprises administering an effective amount of an immune checkpoint inhibitor to the patient, where the patient has already been administered a STING agonist and a cytokine.

[0024] In a further aspect, the disclosure provides a combination therapy for treating a tumor, e.g., in a cancer patient in need thereof, comprising a STING agonist and a cytokine, wherein the STING agonist or the cytokine is formulated for intratumoral administration to the patient. In certain embodiments, both the STING agonist and the cytokine are formulated for intratumoral administration to the patient.

[0025] In certain embodiments, the disclosure provides combination therapies for treating a tumor, e.g., in a cancer patient in need thereof, where both the STING agonist and the cytokine are formulated for intratumoral administration to the patient, and the combination therapy further includes an immune checkpoint inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody) formulated for systemic administration to the cancer patient.

[0026] In some embodiments, the present disclosure provides a mixture comprising a STING agonist, a cytokine and an immune checkpoint inhibitor. In some embodiments, the mixture further comprises human plasma.

[0027] In certain embodiments, the disclosure provides a mixture comprising a STING agonist that is a CDN, a cytokine that is an interleukin, an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, and human plasma.

[0028] In certain embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein the STING agonist or the cytokine is administered intratumorally to the patient, and the patient exhibits reduced tumor recurrence following treatment. In certain embodiments, both a STING agonist and a cytokine are administered intratumorally to the patient.

[0029] In certain embodiments, the STING agonist used in the methods, uses, and combination therapies disclosed herein is a CDN, such as a compound having the following structure ("Compound A"), or a pharma- ceutically acceptable salt thereof: [ka]

[0030] Compound A is a cyclic dinucleotide that can activate STING and is described in US Patent Application Publication No. 2018 / 0230177, which is incorporated herein by reference. Compound A in various salt forms can be administered to cancer patients. For example, in one embodiment, an effective amount of the sodium salt of Compound A is administered to cancer patients. It will be understood that any reference to Compound A in this disclosure also includes its pharma- ceutically acceptable salts. In certain embodiments, Compound A is used in the methods, uses, and combination therapies disclosed herein in combination with the cytokine IL-12.

[0031] In certain embodiments, the cytokines used in the methods, uses, and combination therapies disclosed herein are interleukins, such as human interleukins IL-2, IL-7, IL-10, IL-12, IL-15, or combinations thereof. In certain embodiments, the interleukins are IL-2, IL-7, IL-10, IL-12, or combinations thereof. In some embodiments, the interleukins are IL-2, IL-12, IL-15, or combinations thereof. In one embodiment, the interleukin is IL-2. In another embodiment, the interleukin is IL-7. In another embodiment, the interleukin is IL-10. In another embodiment, the interleukin is IL-15. In certain embodiments, the interleukin is IL-12. In certain other embodiments, the cytokines used in the methods, uses, and combination therapies disclosed herein are interleukins that are fused to a protein to form a fusion protein, such as IL-12 fused to collagen-binding lumican. In other specific embodiments, the cytokine used in the methods, uses and combination therapies disclosed herein is an interleukin fused to a protein to form a fusion protein (e.g., IL-2 fused to collagen-binding lumican). IL-12 or IL-2 fused to lumican is described in PCT Publication WO 2020 / 068261, which is incorporated herein by reference. In certain embodiments, compound A is used in the methods, uses and combination therapies disclosed herein in combination with the cytokine IL-12 fused to lumican. [Brief description of the drawings]

[0032] 5. Brief description of the drawings [Figure 1] Figure 1 shows the antitumor efficacy of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and cytokines (IL-2, IL-12, or IL-15) in a mouse model. Panel A of Figure 1 shows primary and distant tumor growth over time. Data are presented as mean ± SEM. Panel B of Figure 1 shows mouse survival over time.

[0033] [Diagram 2] Figure 2 shows the antitumor efficacy of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and a cytokine (IL-7 or IL-10) in a mouse model. Panel A of Figure 2 shows primary and distant tumor growth over time. Data are presented as mean ± SEM. Panel B of Figure 2 shows mouse survival over time.

[0034] [Figure 3A] Figure 1 shows the antitumor efficacy of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and various doses of IL-12 (50 ng) in a mouse model. Panel A shows primary and distant tumor growth over time. Panel B shows mouse survival over time. Panel C shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM. [Figure 3B] Figure 1 shows the antitumor effect of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and various doses of IL-12 (200 ng) in a mouse model. Panel A shows primary and distant tumor growth over time. Panel B shows mouse survival over time. Panel C shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM. [Figure 3C] Figure 1 shows the antitumor efficacy of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and various doses of IL-12 (1 μg) in a mouse model. Panel A shows primary and distant tumor growth over time. Panel B shows mouse survival over time. Panel C shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM.

[0035] [Figure 4]Figure 4 shows the antitumor effect of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and various doses of IL-12 (3ng, 10ng, or 30ng) in a mouse model. Panel A of Figure 4 shows primary and distant tumor growth over time. Panel B of Figure 4 shows mouse survival over time. Panel C of Figure 4 shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM.

[0036] [Diagram 5] Figure 5 shows the antitumor effect of a combination of STING agonist (compound A), immune checkpoint inhibitor (anti-PD-L1 antibody) and IL-12-Fc in a mouse model. Panel A of Figure 5 shows primary and distant tumor growth over time. Panel B of Figure 5 shows mouse survival over time. Panel C of Figure 5 shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM.

[0037] [Figure 6] Figure 6 shows the antitumor effect of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and various doses of IL-12 (5ng, 17ng, or 50ng) in a mouse model. Panel A of Figure 6 shows primary and distant tumor growth over time. Panel B of Figure 6 shows mouse survival over time. Panel C of Figure 6 shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM.

[0038] [Figure 7]Figure 7 shows the antitumor efficacy of a triple combination of a STING agonist (Compound A) and an immune checkpoint inhibitor (anti-PD-L1 antibody) with interleukin IL-12-Fc (30 ng) or IL12-MSA-lumican (20 ng, 60 ng or 200 ng) in a mouse model. Panel A of Figure 7 shows primary and distant tumor growth over time. Panel B of Figure 7 shows mouse survival over time. Panel C of Figure 7 shows mouse weight change over time. Data for panels A and C are shown as mean ± SEM.

[0039] [Figure 8] Figure 8 shows the antitumor effects of various combinations of STING agonist (compound A), immune checkpoint inhibitor (anti-PD-L1 antibody) and interleukin IL12-MSA-lumican in a mouse model. Panel A of Figure 8 shows primary and distant tumor growth over time. Panel B of Figure 8 shows mouse survival over time. Data in Panel A are shown as mean ± SEM.

[0040] [Figure 9] Figure 2 shows tumor growth in naive mice or mice previously treated with a STING agonist (Compound A) and a triple combination of immune checkpoint inhibitor (anti-PD-L1 antibody) and interleukin IL12-MSA-lumican (20 ng, 60 ng or 200 ng). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0041] 6. Detailed Description 6.1.Definition As used in this specification and the appended claims, unless specified to the contrary, the following terms and abbreviations have the meanings indicated.

[0042] "Combination therapy" as used herein refers to the administration regimen of the listed substances for the specific listed administration route to treat the listed disease.For example, a combination therapy for treating tumors in cancer patients disclosed herein that includes a STING agonist and a cytokine, where both the STING agonist and the cytokine are formulated for intratumoral administration to patients, will include an intratumoral administration regimen for each of the STING agonist and the cytokine at a dosage and frequency sufficient to treat tumors in cancer patients.

[0043] "Co-administered" herein refers to any administration form of two or more different therapeutic compounds, such that the second administered compound is administered while the first administered therapeutic compound is still effective in the body (e.g., the two compounds are effective in the patient simultaneously, which may include additive or synergistic effects of the two compounds). For example, the STING agonist and cytokine disclosed herein can be administered simultaneously or sequentially, either in the same formulation or in separate formulations. In certain embodiments, the STING agonist and cytokine disclosed herein can be administered within 1 hour, 2 hours, 12 hours, 24 hours, 36 hours, 48 ​​hours, 72 hours, or within 1 week of each other. In some embodiments, the STING agonist is administered first, and in other embodiments, the cytokine is administered first. Thus, an individual undergoing such treatment can benefit from the combined effects of the different therapeutic compounds.

[0044] "Effective amount," as used herein, refers to an amount of a substance disclosed herein (e.g., a STING agonist, cytokine, or immune checkpoint inhibitor disclosed herein) sufficient to treat a described disease, disorder, or condition, or to have a desired described effect on a disease, disorder, or condition, or on one or more mechanisms underlying a disease, disorder, or condition, or to have a desired described biological effect (e.g., enhancing an anti-tumor response, increasing immune cell population or function, or increasing tumor-infiltrating leukocyte proliferation or function) in a human subject, such as a cancer patient. In certain embodiments, when a STING agonist is co-administered with a cytokine (preferably, but optionally an immune checkpoint inhibitor) for the treatment of a tumor, an effective amount refers to both the amount of STING agonist and the amount of cytokine (and amount of checkpoint inhibitor) that, upon co-administration to a human, treats or ameliorate a tumor in a human, or shows a detectable therapeutic or biological effect in a human. The therapeutic effect can be detected, for example, by the reduction of the size of one or more tumors, the reduction of tumor growth, and the increase of survival time.The biological effect can be evaluated by measuring the number of tumor-infiltrating leukocytes using surface markers such as CD45, determining the population of specific immune cells, including but not limited to T cells, NK cells, B cells, dendritic cells or macrophages, in tumor biopsy and blood, and measuring gene expression in single cells and bulk cells in tumors.The biological effect and safety of the treatment can also be examined by measuring various inflammatory cytokines in tumor and blood, by weight and temperature measurement, and by standard clinical and anatomical evaluations that are deemed necessary and appropriate by a licensed clinician.

[0045] "Reducing tumor recurrence" or "preventing tumor recurrence" in cancer patients, as used herein, refers to reducing or preventing tumor recurrence in cancer patients who have been administered a specified agent (e.g., STING agonist, cytokine, preferably optional immune checkpoint inhibitor) compared to similarly affected cancer patients or patient types who have not been administered the specified agent. In certain preferred embodiments, reducing or preventing tumor recurrence occurs even if the patient does not receive further treatment with the specified agent. Without wishing to be bound by theory, in some cases, in addition to treating existing cancer / tumor, treatment with the specified agent enhances the anti-tumor response of the patient's immune system so as to reduce or prevent future tumor recurrence after treatment with the specified agent has ended.

[0046] "Treatment" or "treating" as used herein refers to a therapeutic application associated with the co-administration of a STING agonist and a cytokine (preferably, but optionally with an immune checkpoint inhibitor) disclosed herein to improve the indicated disease, disorder, or condition, or one or more underlying mechanisms of said disease, disorder, or condition, including slowing or stopping the progression of the disease, disorder, or condition, or one or more underlying mechanisms in a human subject, such as a cancer patient. In certain embodiments, when a STING agonist and a cytokine (preferably, but optionally with an immune checkpoint inhibitor) disclosed herein are co-administered for the treatment of a tumor to be treated (e.g., in treating cancer), treatment refers to a therapeutic application for slowing or stopping the progression of a tumor or cancer and / or reversal of a tumor or cancer. Tumor or cancer reversal differs from a therapeutic application that slows or stops a tumor or cancer in that the method of not only stopping but also reversing the progression of the tumor or cancer moves cellular behavior to some extent toward the normal state observed in the absence of the tumor or cancer.

[0047] 6.2. Administration of STING agonists in combination with cytokines and related combination therapies The present disclosure provides a method for treating a disease or disorder, particularly cancer, in a patient in need thereof, for example, a method for treating a tumor in a cancer patient in need thereof, comprising administering to the patient an effective amount of a STING agonist in combination with a cytokine (e.g., conjointly), wherein the STING agonist or cytokine is administered intratumorally to the patient. In certain embodiments, the patient is currently receiving an immune checkpoint inhibitor as part of an antitumor therapy. Conjoint administration contemplates that the STING agonist can be administered simultaneously with, before, or after the administration of the cytokine.

[0048] In certain aspects, the present disclosure provides a method of treating tumors in cancer patients in need thereof, comprising administering to the patient an effective amount of a STING agonist in combination with a cytokine (e.g., conjointly), wherein the STING agonist or cytokine is administered intratumorally to the patient, and the patient shows reduced tumor recurrence after treatment. In certain embodiments, the patient is currently receiving an immune checkpoint inhibitor as part of antitumor therapy. Conjoint administration contemplates that the STING agonist can be administered simultaneously with, before, or after the administration of the cytokine.

[0049] In a further aspect, the present disclosure provides a method for reducing tumor recurrence in a cancer patient in need thereof, comprising administering to the patient an effective amount of a STING agonist in combination with a cytokine (e.g., conjointly), wherein the STING agonist or cytokine is administered intratumorally to the patient. In certain embodiments, the patient is currently receiving an immune checkpoint inhibitor as part of an antitumor therapy. Conjoint administration contemplates that the STING agonist can be administered simultaneously with, before, or after the administration of the cytokine.

[0050] In another aspect, the present disclosure provides a method for enhancing anti-tumor response in a cancer patient, comprising administering to the patient an effective amount of a STING agonist in combination with a cytokine (e.g., synergistically), wherein the STING agonist or cytokine is administered intratumorally to the patient.In certain embodiments, the patient is currently receiving an immune checkpoint inhibitor as part of anti-tumor therapy.As discussed herein, the enhanced anti-tumor response can be shown, for example, by shrinkage of one or more tumors or by prolonged survival.

[0051] In yet another aspect, the disclosure provides a method of increasing immune cell (e.g., T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof) population or function in a cancer patient, comprising administering to the patient an effective amount of a STING agonist in combination (e.g., conjunctively) with a cytokine, where the STING agonist or cytokine is administered intratumorally to the patient. In certain embodiments, the patient is currently receiving an immune checkpoint inhibitor as part of an antitumor therapy. In certain embodiments, such methods increase T cell population or function. In other embodiments, such methods increase NK cell population or function. In other embodiments, such methods increase B cell population or function. In other embodiments, such methods increase dendritic cell population or function. In other embodiments, such methods increase macrophage population or function. As discussed herein, an increase in immune cell population or function can be demonstrated by, for example, determining a population of specific immune cells, including but not limited to T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof, in tumor biopsies and blood.

[0052] In yet another aspect, the present disclosure provides a method for increasing the proliferation or function of tumor-infiltrating leukocytes in cancer patients, comprising administering to the patient an effective amount of a STING agonist in combination with a cytokine (e.g., synergistically), wherein the STING agonist or cytokine is administered intratumorally to the patient.In certain embodiments, the patient is currently receiving an immune checkpoint inhibitor as part of antitumor therapy.The increase in proliferation or function of tumor-infiltrating leukocytes can be shown, for example, by measuring the number of tumor-infiltrating leukocytes using a surface marker such as CD45.

[0053] In some embodiments of the methods and uses of the present disclosure, the STING agonist and the cytokine can both be administered intratumorally to the patient. In these embodiments, the STING agonist and the cytokine can be administered together in the same pharmaceutical composition or in separate pharmaceutical compositions. In other embodiments, the cytokine can be administered intratumorally to the patient and the STING agonist can be administered systemically to the patient (e.g., intravenously, intramuscularly, subcutaneously, or orally). In certain embodiments, the cytokine can be administered intratumorally to the patient and the STING agonist can be administered intravenously to the patient. In certain embodiments, the cytokine can be administered intratumorally to the patient and the STING agonist can be administered intramuscularly to the patient. In other embodiments, the cytokine can be administered intratumorally to the patient and the STING agonist can be administered orally to the patient. In other embodiments, the STING agonist can be administered intratumorally to the patient and the cytokine can be administered systemically to the patient (e.g., intravenously, intramuscularly, or subcutaneously). In certain embodiments, the STING agonist can be administered intratumorally to the patient and the cytokine can be administered intravenously to the patient. In certain embodiments, the STING agonist can be administered intratumorally to the patient, and the cytokine can be administered intramuscularly to the patient. In certain embodiments, the STING agonist can be administered intratumorally to the patient, and the cytokine can be administered subcutaneously to the patient. In certain embodiments, the method is a method for treating tumors in a cancer patient in need thereof. In certain embodiments, the method is a method for reducing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method for preventing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method for enhancing anti-tumor response in a cancer patient. In certain embodiments, the method is a method for increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells or macrophages, or a combination thereof) in a cancer patient.

[0054] In embodiments in which the STING agonist and the cytokine are administered in separate compositions, the two compositions can be administered simultaneously or sequentially. In certain embodiments in which the cytokine and the STING agonist are administered sequentially, the STING agonist can be administered before the administration of the cytokine. Alternatively, the STING agonist can be administered after the administration of the cytokine.

[0055] In some embodiments, the STING agonist and cytokine can be combined, e.g., co-administered, without an additional therapeutic agent. Surprisingly, in some tumors, such as those exemplified herein, the combination of the STING agonist and cytokine provides sufficient tumor inhibition such that additional chemotherapeutic or immunotherapeutic agents do not provide further tumor inhibition.

[0056] However, in other embodiments, the STING agonist and cytokine are administered in combination with one or more additional anti-cancer agents, for example, in combination with an immune checkpoint inhibitor, for example, a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor, including an anti-PD-1 antibody, an anti-PD-L1 antibody, and an anti-CTLA-4 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-PD-1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-PD-L1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-CTLA-4 antibody. Thus, in some embodiments, the present disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising administering to the patient an effective amount of a STING agonist and a cytokine in combination (e.g., conjointly), wherein the STING agonist or cytokine is administered intratumorally to the patient, and further comprising administering to the patient an effective amount of an immune checkpoint inhibitor in combination (e.g., conjointly). In certain embodiments, the immune checkpoint inhibitor is administered to the patient intratumorally. In other embodiments, the immune checkpoint inhibitor is administered to the patient systemically (e.g., intravenously, intramuscularly, or subcutaneously). In certain embodiments, the immune checkpoint inhibitor is administered intravenously. In certain embodiments, the immune checkpoint inhibitor is administered intramuscularly. In certain embodiments, the immune checkpoint inhibitor is administered subcutaneously. In certain embodiments, the patient is receiving the immune checkpoint inhibitor as part of an anti-tumor therapy.

[0057] In certain aspects, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising administering to the patient an effective amount of a STING agonist and a cytokine in combination (e.g., conjointly), wherein the STING agonist or cytokine is administered intratumorally to the patient, and further comprising administering to the patient an effective amount of an immune checkpoint inhibitor in combination (e.g., conjointly), wherein the patient exhibits reduced tumor recurrence after treatment. In certain embodiments, the immune checkpoint inhibitor is administered intratumorally to the patient. In other embodiments, the immune checkpoint inhibitor is administered systemically (e.g., intravenously, intramuscularly, or subcutaneously) to the patient. In certain embodiments, the immune checkpoint inhibitor is administered intravenously. In certain embodiments, the immune checkpoint inhibitor is administered intramuscularly. In certain embodiments, the immune checkpoint inhibitor is administered subcutaneously. In certain embodiments, the patient is receiving an immune checkpoint inhibitor as part of an antitumor therapy.

[0058] In some embodiments, the methods and uses described herein include co-administering an effective amount of a STING agonist, a cytokine and an immune checkpoint inhibitor to a patient, wherein the STING agonist and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered systemically to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody or an anti-CTLA-4 antibody. In certain embodiments, the method is a method of treating a tumor in a cancer patient in need thereof. In certain embodiments, the method is a method of reducing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method of preventing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method of enhancing anti-tumor response in a cancer patient. In certain embodiments, the method is a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells or macrophages, or a combination thereof) in a cancer patient.

[0059] In some embodiments, the methods and uses described herein include co-administering to a patient an effective amount of a STING agonist, a cytokine and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are administered intratumorally to the patient, the immune checkpoint inhibitor is administered systemically to the patient, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody or an anti-CTLA-4 antibody, and the STING agonist is a cyclic dinucleotide (CDN). In certain embodiments, the immune checkpoint inhibitor is an anti-PD-1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-PD-L1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-CTLA-4 antibody. In certain embodiments, the method is a method of treating a tumor in a cancer patient in need thereof. In certain embodiments, the method is a method of reducing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method of preventing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method of enhancing an anti-tumor response in a cancer patient. In certain embodiments, the method is a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells or macrophages, or a combination thereof) in a cancer patient.

[0060] In some embodiments, the methods and uses described herein include co-administering to a patient an effective amount of a STING agonist, a cytokine and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered systemically to the patient, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody or an anti-CTLA-4 antibody, the cytokine is an interleukin, and the STING agonist is a cyclic dinucleotide (CDN). In certain embodiments, the immune checkpoint inhibitor is an anti-PD-1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-PD-L1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-CTLA-4 antibody. In certain embodiments, the method is a method of treating a tumor in a cancer patient in need thereof. In certain such embodiments, the interleukin is IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain embodiments, the method is a method of reducing tumor recurrence in a cancer patient in need thereof. In certain embodiments, the method is a method for preventing tumor recurrence in cancer patients who need it.In certain embodiments, the method is a method for enhancing the anti-tumor response of cancer patients.In certain embodiments, the method is a method for increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells or macrophages, or combinations thereof) in cancer patients.

[0061] In some embodiments, the present disclosure provides a method for treating tumors in a cancer patient in need thereof, comprising having a STING agonist, a cytokine and an immune checkpoint inhibitor present in the patient's body at the same time.In some embodiments, the method comprises administering an effective amount of a STING agonist to the patient, and the patient has already been administered a cytokine and an immune checkpoint inhibitor.In some embodiments, the method comprises administering an effective amount of a cytokine to the patient, and the patient has already been administered a STING agonist and an immune checkpoint inhibitor.In some embodiments, the method comprises administering an effective amount of an immune checkpoint inhibitor to the patient, and the patient has already been administered a STING agonist and a cytokine.

[0062] In some embodiments, the present disclosure provides a method for reducing tumor recurrence in a patient, comprising simultaneously presenting a STING agonist, a cytokine and an immune checkpoint inhibitor in the patient's body. In some embodiments, the method comprises administering an effective amount of a STING agonist to the patient, and the patient has already been administered a cytokine and an immune checkpoint inhibitor. In some embodiments, the method comprises administering an effective amount of a cytokine to the patient, and the patient has already been administered a STING agonist and an immune checkpoint inhibitor. In some embodiments, the method comprises administering an effective amount of an immune checkpoint inhibitor to the patient, and the patient has already been administered a STING agonist and a cytokine.

[0063] In some embodiments, the present disclosure provides a method for preventing tumor recurrence in a patient, comprising simultaneously presenting a STING agonist, a cytokine and an immune checkpoint inhibitor in the patient's body. In some embodiments, the method comprises administering an effective amount of a STING agonist to the patient, and the patient has already been administered a cytokine and an immune checkpoint inhibitor. In some embodiments, the method comprises administering an effective amount of a cytokine to the patient, and the patient has already been administered a STING agonist and an immune checkpoint inhibitor. In some embodiments, the method comprises administering an effective amount of an immune checkpoint inhibitor to the patient, and the patient has already been administered a STING agonist and a cytokine.

[0064] In certain embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering systemically (e.g., intravenously) to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. In certain such embodiments, the STING agonist is a CDN and / or the cytokine is an interleukin.

[0065] In further specific embodiments, the present disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering an effective amount of an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, systemically (e.g., intravenously) to the cancer patient, wherein the patient shows reduced tumor recurrence after treatment. In certain such specific embodiments, the STING agonist is CDN. In certain such specific embodiments, the cytokine is an interleukin, such as IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain such specific embodiments, the STING agonist is CDN and the cytokine is an interleukin, such as IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain embodiments, patients exhibit a reduction in tumor recurrence following treatment.

[0066] In certain embodiments, the present disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering an effective amount of an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, systemically (e.g., intravenously) to the cancer patient. In certain such specific embodiments, the STING agonist is a CDN. In certain such specific embodiments, the cytokine is an interleukin, such as IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain such specific embodiments, the STING agonist is a CDN and the cytokine is an interleukin, such as IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain embodiments, the patient shows reduced tumor recurrence after treatment.

[0067] In certain embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist that is a CDN and a cytokine that is IL-12, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering systemically (e.g., intravenously) to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, anti-PD-L1 antibody, or anti-CTLA-4 antibody.

[0068] In certain embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist, a cytokine, and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are administered intratumorally to the cancer patient, and the immune checkpoint inhibitor is administered systemically to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

[0069] In certain embodiments, the present disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist that is a CDN, a cytokine that is IL-12, and an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both the CDN and IL-12 are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such embodiments, the IL-12 is a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain embodiments, the patient shows reduced tumor recurrence after treatment.

[0070] In further particular embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the cancer patient an effective amount of a STING agonist, which is compound A, a cytokine, and an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both compound A and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a collagen binding protein, such as lumican. In certain such particular embodiments, the cytokine is fused to an immunoglobulin Fc domain. In certain embodiments, the patient shows reduced tumor recurrence after treatment.

[0071] In further particular embodiments, the disclosure provides a method of treating a tumor in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist that is a CDN and a cytokine that is IL-12, wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody systemically (e.g., intravenously), wherein the patient exhibits reduced tumor recurrence following treatment. In certain such particular embodiments, the IL-12 is a fusion protein of IL-12, e.g., IL-12-Fc or IL-12-MSA-lumican.

[0072] In other embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, comprising administering to the patient an effective amount of a STING agonist and a cytokine in combination (e.g., conjointly), wherein the STING agonist or the cytokine is administered intratumorally to the patient, and further comprising administering to the patient an effective amount of an immune checkpoint inhibitor in combination (e.g., conjointly). In certain embodiments, the immune checkpoint inhibitor is administered intratumorally to the patient. In other embodiments, the immune checkpoint inhibitor is administered systemically (e.g., intravenously, intramuscularly, or subcutaneously) to the patient. In certain embodiments, the immune checkpoint inhibitor is administered intravenously. In certain embodiments, the immune checkpoint inhibitor is administered intramuscularly. In certain embodiments, the immune checkpoint inhibitor is administered subcutaneously. In certain embodiments, the patient is receiving an immune checkpoint inhibitor as part of an anti-tumor therapy.

[0073] In certain embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, wherein both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering systemically (e.g., intravenously) to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. In certain such embodiments, the STING agonist is a CDN and / or the cytokine is an interleukin.

[0074] In certain embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, comprising co-administering to the patient effective amounts of a STING agonist that is a CDN and a cytokine that is IL-12, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering systemically (e.g., intravenously) to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

[0075] In certain embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, the method comprising co-administering to the patient effective amounts of a STING agonist, a cytokine, and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered systemically to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

[0076] In certain embodiments, the disclosure provides a method of enhancing an anti-tumor response in a cancer patient, comprising co-administering to the patient effective amounts of a STING agonist that is a CDN, a cytokine that is IL-12, and an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both the CDN and IL-12 are administered intratumorally to the patient and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such embodiments, the IL-12 is a fusion protein of IL-12, e.g., IL-12-Fc or IL-12-MSA-lumican.

[0077] In further particular embodiments, the present disclosure provides a method of enhancing anti-tumor response in a cancer patient, comprising co-administering to the cancer patient an effective amount of a STING agonist, which is compound A, a cytokine, and an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both compound A and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a collagen-binding protein, such as lumican. In certain such particular embodiments, the cytokine is fused to an immunoglobulin Fc domain.

[0078] In other embodiments, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof) in a cancer patient, comprising administering to the patient an effective amount of a STING agonist and a cytokine in combination (e.g., conjunctively), wherein the STING agonist or cytokine is administered intratumorally to the patient, and further comprising administering to the patient an effective amount of an immune checkpoint inhibitor in combination (e.g., conjunctively). In certain embodiments, the immune checkpoint inhibitor is administered intratumorally to the patient. In other embodiments, the immune checkpoint inhibitor is administered systemically (e.g., intravenously, intramuscularly, or subcutaneously) to the patient. In certain embodiments, the immune checkpoint inhibitor is administered intravenously. In certain embodiments, the immune checkpoint inhibitor is administered intramuscularly. In certain embodiments, the immune checkpoint inhibitor is administered subcutaneously. In certain embodiments, the patient is receiving an immune checkpoint inhibitor as part of an antitumor therapy. In certain embodiments, such methods increase the population or function of T cells. In other embodiments, such methods increase the population or function of NK cells. In other embodiments, such methods increase B cell population or function. In other embodiments, such methods increase dendritic cell population or function. In other embodiments, such methods increase macrophage population or function.

[0079] In certain embodiments, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof) in a cancer patient, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody systemically (e.g., intravenously) to the cancer patient. In certain such embodiments, the STING agonist is a CDN and / or the cytokine is an interleukin. In certain embodiments, such methods increase the population or function of T cells. In other embodiments, such methods increase the population or function of NK cells. In other embodiments, such methods increase the population or function of B cells. In other embodiments, such methods increase the population or function of dendritic cells. In other embodiments, such methods increase the population or function of macrophages.

[0080] In certain embodiments, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof) in a cancer patient, comprising co-administering to the patient an effective amount of a STING agonist that is a CDN and a cytokine that is IL-12, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering systemically (e.g., intravenously) to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. In certain embodiments, such methods increase the population or function of T cells. In other embodiments, such methods increase the population or function of NK cells. In other embodiments, such methods increase the population or function of B cells. In other embodiments, such methods increase the population or function of dendritic cells. In other embodiments, such methods increase the population or function of macrophages.

[0081] In certain embodiments, the disclosure provides a method of increasing the population or function of immune cells (e.g., T cells, NK cells, B cells, dendritic cells, or macrophages, or a combination thereof) in a cancer patient, comprising co-administering to the patient effective amounts of a STING agonist, a cytokine, and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered systemically to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. In certain embodiments, such methods increase the population or function of T cells. In other embodiments, such methods increase the population or function of NK cells. In other embodiments, such methods increase the population or function of B cells. In other embodiments, such methods increase the population or function of dendritic cells. In other embodiments, such methods increase the population or function of macrophages.

[0082] In certain embodiments, the present disclosure provides a method of enhancing anti-tumor responses in a cancer patient, comprising co-administering to the patient an effective amount of a STING agonist that is a CDN, a cytokine that is IL-12, and an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both the CDN and IL-12 are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered intravenously to the cancer patient. In certain such embodiments, the IL-12 is a fusion protein of IL-12, e.g., IL-12-Fc or IL-12-MSA-lumican. In certain embodiments, such methods increase the population or function of T cells. In other embodiments, such methods increase the population or function of NK cells. In other embodiments, such methods increase the population or function of B cells. In other embodiments, such methods increase the population or function of dendritic cells. In other embodiments, such methods increase the population or function of macrophages.

[0083] In further particular embodiments, the present disclosure provides a method of enhancing anti-tumor response in a cancer patient, comprising co-administering to the cancer patient an effective amount of a STING agonist, which is compound A, a cytokine, and an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both compound A and the cytokine are administered intratumorally to the cancer patient, and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a collagen-binding protein, such as lumican. In certain such particular embodiments, the cytokine is fused to an immunoglobulin Fc domain.

[0084] In certain embodiments, such methods increase T cell population or function. In other embodiments, such methods increase NK cell population or function. In other embodiments, such methods increase B cell population or function. In other embodiments, such methods increase dendritic cell population or function. In other embodiments, such methods increase macrophage population or function.

[0085] In other embodiments, the present disclosure provides a method for increasing tumor-infiltrating leukocyte proliferation or function in a cancer patient, comprising co-administering a STING agonist and a cytokine in combination (e.g., synergistically) to the patient, wherein the STING agonist or cytokine is administered intratumorally to the patient, and further comprising administering an effective amount of an immune checkpoint inhibitor in combination (e.g., synergistically) to the patient. In certain embodiments, the immune checkpoint inhibitor is administered intratumorally to the patient. In other embodiments, the immune checkpoint inhibitor is administered systemically (e.g., intravenously, intramuscularly, or subcutaneously) to the patient. In certain embodiments, the immune checkpoint inhibitor is administered intravenously.

[0086] In certain embodiments, the present disclosure provides a method of increasing tumor-infiltrating leukocyte proliferation or function in a cancer patient, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering an effective amount of an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, systemically (e.g., intravenously) to the cancer patient. In certain such embodiments, the STING agonist is a CDN and / or the cytokine is an interleukin. In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, where the protein is an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, where the protein is not an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a collagen-binding protein, such as lumican. In certain such embodiments, the cytokine is fused to an immunoglobulin Fc domain.

[0087] In certain embodiments, the present disclosure provides a method of increasing tumor-infiltrating leukocyte proliferation or function in a cancer patient, comprising co-administering to the patient an effective amount of a STING agonist that is a CDN and a cytokine that is IL-12, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody systemically (e.g., intravenously). In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, where the protein is an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, where the protein is not an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a collagen-binding protein, such as lumican. In certain such embodiments, the cytokine is fused to an immunoglobulin Fc domain.

[0088] In one aspect, the disclosure provides a method of treating or preventing metastasis in a human cancer patient, comprising administering to the patient an effective amount of a STING agonist and a cytokine in combination (e.g., conjointly), wherein the STING agonist or cytokine is administered intratumorally to the patient, and further comprising administering to the patient an effective amount of an immune checkpoint inhibitor in combination (e.g., conjointly), if necessary. For example, the method can be used to treat primary or metastatic tumors that are resistant to immune checkpoint therapy. In some such embodiments, the STING agonist and cytokine are co-administered with a PD-1, PD-L1 or CTLA-4 inhibitor, or the cancer patient is currently receiving an immune checkpoint inhibitor as part of an anti-tumor therapy. In certain such specific embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain such specific embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain such specific embodiments, the cytokine is fused to a collagen-binding protein, such as lumican. In certain such embodiments, the cytokine is fused to an immunoglobulin Fc domain.

[0089] In other embodiments, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising administering to the patient an effective amount of a STING agonist and a cytokine in combination (e.g., conjunctively), wherein the STING agonist or the cytokine is administered intratumorally to the patient, and further comprising administering to the patient an effective amount of an immune checkpoint inhibitor in combination (e.g., conjunctively). In certain embodiments, the immune checkpoint inhibitor is administered intratumorally to the patient. In other embodiments, the immune checkpoint inhibitor is administered systemically (e.g., intravenously, intramuscularly, or subcutaneously) to the patient. In certain embodiments, the immune checkpoint inhibitor is administered intravenously. In certain embodiments, the immune checkpoint inhibitor is administered intramuscularly. In certain embodiments, the immune checkpoint inhibitor is administered subcutaneously. In certain embodiments, the patient is receiving an immune checkpoint inhibitor as part of an anti-tumor therapy.

[0090] In certain embodiments, the present disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering an effective amount of an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, systemically (e.g., intravenously) to the cancer patient. In certain such specific embodiments, the STING agonist is CDN. In certain such specific embodiments, the cytokine is an interleukin, such as IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican. In certain such specific embodiments, the STING agonist is CDN and the cytokine is an interleukin, such as IL-12, such as a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican.

[0091] In certain embodiments, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist that is a CDN and a cytokine that is IL-12, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering systemically (e.g., intravenously) to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

[0092] In certain embodiments, the present disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient an effective amount of a STING agonist that is a CDN and a cytokine, where both the STING agonist and the cytokine are administered intratumorally to the patient, and further comprising co-administering to the cancer patient an effective amount of an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody systemically (e.g., intravenously). In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, where the protein is an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, where the protein is not an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a collagen binding protein, such as lumican. In certain such embodiments, the cytokine is fused to an immunoglobulin Fc domain.

[0093] In certain embodiments, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist that is a CDN, a cytokine that is IL-12, and an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both the CDN and IL-12 are administered intratumorally to the patient and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such embodiments, the IL-12 is a fusion protein of IL-12, e.g., IL-12-Fc or IL-12-MSA-lumican.

[0094] In certain embodiments, the disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the patient effective amounts of a STING agonist, a cytokine, and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are administered intratumorally to the cancer patient, and the immune checkpoint inhibitor is administered systemically to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

[0095] In further particular embodiments, the present disclosure provides a method of reducing tumor recurrence in a cancer patient in need thereof, comprising co-administering to the cancer patient an effective amount of a STING agonist, which is compound A, a cytokine, and an immune checkpoint inhibitor, which is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, wherein both compound A and the cytokine are administered intratumorally to the patient, and the immune checkpoint inhibitor is administered intravenously to the patient. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain such particular embodiments, the cytokine is fused to a collagen-binding protein, such as lumican.

[0096] In some embodiments, the present disclosure provides a mixture comprising a STING agonist, a cytokine and an immune checkpoint inhibitor. In some embodiments, the mixture further comprises human plasma.

[0097] In certain embodiments, the present disclosure provides a mixture comprising a STING agonist that is a CDN, a cytokine, an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, and human plasma. In further specific embodiments, the present disclosure provides a mixture comprising a STING agonist that is a CDN, a cytokine that is an interleukin, an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, and human plasma. In yet further specific embodiments, the present disclosure provides a mixture comprising a STING agonist that is compound A, a cytokine that is an interleukin (e.g., IL-12), an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, and human plasma. In certain such specific embodiments, the IL-12 is a fusion protein of IL-12, such as IL-12-Fc or IL-12-MSA-lumican.

[0098] Thus, in some embodiments, a STING agonist and a cytokine can be administered to a cancer patient in combination with, e.g., synergistically, a PD-1, PD-L1, or CTLA-4 inhibitor, such as those described herein. In such cases, the PD-1, PD-L1, or CTLA-4 inhibitor can be administered simultaneously, before, or after administration of the STING agonist and / or cytokine. In some embodiments, the PD-1, PD-L1, or CTLA-4 inhibitor can be administered intratumorally. In other embodiments, the PD-1, PD-L1, or CTLA-4 inhibitor can be administered systemically, e.g., intravenously, subcutaneously, or intramuscularly. In certain embodiments, both the STING agonist and the cytokine are administered intratumorally to a cancer patient, and the PD-1, PD-L1, or CTLA-4 inhibitor is administered systemically, e.g., intravenously, subcutaneously, or intramuscularly. In other embodiments, the cytokine is administered intratumorally to the cancer patient, and both the STING agonist and the PD-1, PD-L1, or CTLA-4 inhibitor are administered systemically, for example, intravenously, subcutaneously, intramuscularly, or orally. In other embodiments, the STING agonist is administered intratumorally to the cancer patient, and both the cytokine and the PD-1, PD-L1, or CTLA-4 inhibitor are administered systemically, for example, intravenously, subcutaneously, or intramuscularly. In certain embodiments, both the cytokine and the PD-1, PD-L1, or CTLA-4 inhibitor are administered intratumorally to the cancer patient, and the STING agonist is administered systemically, for example, intravenously, subcutaneously, intramuscularly, or orally. In some embodiments, both the STING agonist and the PD-1, PD-L1, or CTLA-4 inhibitor are administered intratumorally to the cancer patient, and the cytokine is administered systemically, for example, intravenously, subcutaneously, or intramuscularly. In other embodiments, the STING agonist, cytokine, and PD-1, PD-L1, or CTLA-4 inhibitor are all administered intratumorally to cancer patients.

[0099] In certain embodiments of the disclosed methods, the STING agonist and cytokine are combined, e.g., co-administered, with a CTLA-4 inhibitor and either a PD-1 inhibitor or a PD-L1 inhibitor. In certain such embodiments, the CTLA-4 inhibitor is an anti-CTLA-4 antibody that is administered intratumorally or systemically, particularly intratumorally.

[0100] In certain embodiments, the methods and uses described herein provide that administration of a STING agonist and a cytokine and optionally an immune checkpoint inhibitor produces an abscopal effect in tumors distal to the intratumoral administration site of the STING agonist or cytokine.For example, in some embodiments, the methods and uses described herein treat tumors distal to the intratumoral administration site of the STING agonist and / or cytokine.In some embodiments, the methods and uses described herein treat tumors distal to the intratumoral administration site of the STING agonist.In some embodiments, the methods and uses described herein treat tumors distal to the intratumoral administration site of the STING agonist.

[0101] In one embodiment, STING agonist and cytokine are administered to cancer patients who have already undergone immune checkpoint inhibitor therapy, for example, cancer patients whose tumor or cancer has stabilized.In certain embodiments, cancer patients undergo at least 1 or 2 cycles of immune checkpoint inhibitor therapy before administering STING agonist and cytokine.For example, cancer patients may undergo 2, 3, 4, 5, 6, 7 or 8 cycles of immune checkpoint inhibitor therapy before administering STING agonist and cytokine.In certain of these embodiments, cancer patients continue to undergo immune checkpoint inhibitor therapy with continuous cycles of STING agonist and cytokine.

[0102] In some embodiments, the present disclosure provides a combination therapy for treating a tumor, for example, in a cancer patient in need thereof, comprising a STING agonist and a cytokine, wherein the STING agonist or the cytokine is formulated for intratumoral administration to the patient. In certain embodiments, both the STING agonist and the cytokine are formulated for intratumoral administration to the patient. In some embodiments, the cytokine is formulated for intratumoral administration, and the STING agonist is formulated for systemic administration, for example, intravenous, subcutaneous, intramuscular or oral administration, to the patient. In certain embodiments, the STING agonist is formulated for intravenous administration. In certain embodiments, the STING agonist is formulated for subcutaneous administration. In certain embodiments, the STING agonist is formulated for intramuscular administration. In certain embodiments, the STING agonist is formulated for oral administration. In other embodiments, the STING agonist is formulated for intratumoral administration, and the cytokine is formulated for systemic administration, for example, intravenous, subcutaneous or intramuscular administration, to the patient. In certain embodiments, the cytokine is formulated for intravenous administration. In certain embodiments, the cytokine is formulated for subcutaneous administration. In certain embodiments, the cytokine is formulated for intramuscular administration.

[0103] In certain embodiments, the disclosure provides combination therapies for treating tumors, e.g., in a cancer patient in need thereof, where both the STING agonist and the cytokine are formulated for intratumoral administration to the patient, and the combination therapy further comprises an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, formulated for systemic administration to the cancer patient. In certain such embodiments, the STING agonist is a CDN and / or the cytokine is an interleukin.

[0104] In certain embodiments, the disclosure provides a combination therapy for treating a tumor in a cancer patient in need thereof, where the STING agonist is CDN and the cytokine is IL-12, and where both the STING agonist and the cytokine are formulated for intratumoral administration to the patient, and the combination therapy further comprises an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, formulated for systemic administration to the cancer patient.

[0105] In certain embodiments, the combination therapy disclosed herein further comprises an immune checkpoint inhibitor. In certain embodiments, the combination therapy disclosed herein further comprises an immune checkpoint inhibitor, such as a PD-1, PD-L1 or CTLA-4 inhibitor (e.g., an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody). In certain embodiments, the immune checkpoint inhibitor is an anti-PD-1 antibody. In certain embodiments, the immune checkpoint inhibitor is an anti-PD-L1 antibody. In certain embodiments, the immune checkpoint inhibitor is a CTLA-4 antibody. In some embodiments, the immune checkpoint inhibitor is formulated for intratumoral administration to the cancer patient. In other embodiments, the immune checkpoint inhibitor is formulated for systemic administration, such as intravenous, subcutaneous or intramuscular administration, to the cancer patient. In certain embodiments, the immune checkpoint inhibitor is formulated for intravenous administration. In certain embodiments, the immune checkpoint inhibitor is formulated for subcutaneous administration. In certain embodiments, the immune checkpoint inhibitor is formulated for intramuscular administration.

[0106] In certain embodiments, the disclosure provides combination therapies for treating tumors, e.g., in a cancer patient in need thereof, where a STING agonist or cytokine is formulated for intratumoral administration to the patient, and the combination therapy further comprises an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, formulated for systemic administration to the cancer patient. In certain embodiments, the disclosure provides combination therapies for treating tumors, e.g., in a cancer patient in need thereof, where a STING agonist and a cytokine are both formulated for intratumoral administration to the patient, and the combination therapy further comprises an immune checkpoint inhibitor that is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, formulated for systemic administration to the cancer patient.

[0107] In certain embodiments, the disclosure provides combination therapies for treating tumors, e.g., in a cancer patient in need thereof, comprising a STING agonist, a cytokine and an immune checkpoint inhibitor, wherein the STING agonist and the cytokine are formulated for intratumoral administration to the patient, and the immune checkpoint inhibitor is formulated for systemic administration to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, and an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

[0108] In certain embodiments, the present disclosure provides a combination therapy for treating a tumor in a cancer patient in need thereof, wherein the STING agonist is a CDN, the cytokine is an interleukin, both the STING agonist and the cytokine are formulated for intratumoral administration to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, and is formulated for systemic administration to the cancer patient. In certain such specific embodiments, the STING agonist is Compound A. In certain such specific embodiments, the cytokine is an interleukin that is IL-12. In certain embodiments, the present disclosure provides a combination therapy for treating a tumor in a cancer patient in need thereof, wherein the STING agonist is Compound A, the cytokine is IL-12, both the STING agonist and the cytokine are formulated for intratumoral administration to the patient, and the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody, and is formulated for systemic administration to the cancer patient. In certain such specific embodiments, the cytokine is fused to a protein to form a fusion protein. In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain such embodiments, the cytokine is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment.

[0109] 6.3.STING agonists, cytokines and immune checkpoint inhibitors In certain embodiments, the STING agonist used in the methods, uses, combination therapies and mixtures described herein is a cyclic dinucleotide (CDN) compound. For example, the STING agonist can be 2'3'-CDN, such as 2'3'-cGAMP, compound A, compound B, or compound C, particularly compound A, as shown below. In other embodiments, the STING agonist is 3'3'-CDN, 2'2'-CDN, or 3'2'-CDN, such as 3'3'-cGAMP, 2'2'-cGAMP, or 3'2'-cGAMP. In some embodiments, the STING agonist is a CDN that is an analog of 2'3'-cGAMP (i.e., 2'3'-CDN that contains guanine nucleobase and adenine nucleobase), such as compound A and compound B, particularly compound A.

[0110] In some embodiments, the STING agonist is a benzophenone analogue. In further embodiments, the STING agonist is a dimeric amide benzimidazole.

[0111] Examples of STING agonists that can be used in accordance with the present disclosure include ADU-S100 (MIW815), BMS-986301, CRD5500, CMA (10-carboxymethyl-9-acridanone), diABZI STING agonist-1 (e.g., CAS number: 2138299-34-8), DMXAA (ASA404 / vadimezan), E7766, GSK-532, GSK-3745417, MK-1454, MK-2118, SB-11285, SRCB-0074, TAK-676, TTI-10001, SR-717, and MSA-2.

[0112] In one embodiment, the CDN used in the methods and combination therapies according to the present disclosure is the following compound ("Compound A"), or a pharma- ceutically acceptable salt thereof: [ka]

[0113] Compound A can exert a strong antitumor effect by acting both locally and systemically. Compound A can substantially reduce or prevent the spread of metastasis when administered at a certain dosage to a cancer patient in need thereof. According to the present disclosure, the ability of Compound A to reduce or prevent the onset and / or progression of metastasis can be enhanced when administered in combination, e.g., co-administered, with a cytokine. Furthermore, according to the present disclosure, Compound A has been discovered to exert a strong abscopal effect when administered in combination with a cytokine.

[0114] In some embodiments where compound A acts as a STING agonist administered in combination with a cytokine, compound A can be administered over multiple cycles. For example, in one embodiment, the first cycle includes administering compound A on days 1, 8, and 15 of a four week period, and subsequent cycles include administering compound A on days 1 and 15 of a four week period (i.e., every other week). Compound A can be administered intratumorally or systemically, including subcutaneously, intramuscularly, or intravenously. In some embodiments, compound A can be administered at a dosage ranging from 50 μg to 6,500 μg on the days of the cycle designated for administration. In some embodiments, compound A can be administered at a dosage ranging from 100 μg to 3,000 μg on the days of the cycle designated for administration. In some embodiments, compound A can be administered at a dosage ranging from 100 μg to 1,200 μg on the days of the cycle designated for administration.

[0115] In one embodiment, the CDN used in the methods and combination therapies according to the present disclosure is the following compound ("Compound B"), or a pharma- ceutically acceptable salt thereof: [ka]

[0116] In another embodiment, the CDN used in the methods and combination therapies according to the present disclosure is the following compound ("Compound C"), or a pharma- ceutically acceptable salt thereof: [ka]

[0117] In another embodiment, the STING agonist used in the methods and combination therapies according to the present disclosure is a compound disclosed in International Publication No. WO 2019 / 165032, which is incorporated herein by reference. Such STING agonists can be administered to patients orally, systemically, or intratumorally. One such example of a STING agonist that can be used according to the present disclosure is SR-717 ("Compound D"), which has the following structure, or a pharma- ceutical acceptable salt thereof: [ka]

[0118] In another embodiment, the STING agonist used in the methods and combination therapies according to the present disclosure is MSA-2 (“Compound E”), or a pharma- ceutically acceptable salt thereof, which has the following structure: [ka] MSA-2 can be administered to patients orally, systemically, or intratumorally.

[0119] Further examples of CDNs that can be used as STING agonists in the present methods and combination therapies can be found in the following publications: WO 2014 / 144666, WO 2014 / 179335, WO 2014 / 189806, WO 2015 / 161762, WO 2016 / 096174, WO 2017 / 027646 No. 2017 / 027645, No. 2017 / 161349, No. 2018 / 118664, No. 2018 / 118665, No. 2018 / 208667, No. 2019 / 165032 and No. 2019 / 046511, the contents of each of which are incorporated herein by reference.

[0120] In other embodiments, the STING agonists used in the methods and combination therapies according to the present disclosure can be conjugated to an antibody or antigen-binding fragment, thus generating an antibody-drug conjugate (ADC).

[0121] In one embodiment, the ADCs administered in accordance with the methods and combination therapies disclosed herein have a structure as described in U.S. Patent Application Publication No. 2017 / 0298139, WO 2017 / 100305, WO 2018 / 200812, or WO 2018 / 140831, the contents of each of which are incorporated herein by reference.

[0122] In certain embodiments, an ADC that can be used in the methods and combination therapies according to the disclosure has the structure of formula IA: (IA)Ab-[-LD] n [In the formula, "D" is a CDN having the structure of formula IIa: [ka] (In the formula, W, X, Y and Z are independently CH or N; R 1 is a thiol group, an amino group, or C 1~6 C substituted with alkylamino groups 2~4 is alkyl, R p is independently for each occurrence a hydroxyl, a thiol, or a C 1~6 Alkyl, Borano (-BH3 - ), or -NR'R", where R' and R" are independently selected at each occurrence from hydrogen, halogen, thiol, hydroxyl, carboxyl, C 1~6 Alkoxy, C 1~6 Hydroxyalkoxy, -OC(O)C 1~6 Alkyl, -N(H)C(O)C 1~6 Alkyl, -N(C 1~3Alkyl)C(O)C 1~6 Alkyl, Amino, C 1~6 Alkylamino, di(C 1~6 C optionally substituted with one or more groups selected from amino, oxo, azido, and cyano 1~6 alkyl or R' and R" on the same nitrogen together form C 3~5 (forming a heterocyclic ring) or a pharma- ceutically acceptable salt thereof, "Ab" refers to an antibody or binding fragment thereof that binds to a target antigen; "L" represents, independently for each occurrence, a linker linking one or more occurrences of D to Ab; "n" represents the number of occurrences of D linked to Ab via a linker (L); CDN(D) is the R of CDN 1 Thiol group, amino group or C 1~6 covalently attached to a linker (L) at the alkylamino group.

[0123] In some embodiments where the STING agonist is part of an ADC of formula IA, the CDN of the ADC has the structure of formula IIb: [ka] or a pharma- ceutically acceptable salt thereof.

[0124] In some embodiments where the STING agonist is part of an ADC of formula IA, the CDN of the ADC has the structure of formula IIc: [ka] or a pharma- ceutically acceptable salt thereof.

[0125] In some embodiments where the STING agonist is part of an ADC of formula IA, the ADC has the structure of formula III: [ka] has.

[0126] In some embodiments where the STING agonist is part of an ADC of formula IA, the ADC has the structure of formula IV: [ka] has.

[0127] In some embodiments where the STING agonist is part of an ADC of formula IA, the ADC ("Compound F") has the following structure: [ka] has.

[0128] In some embodiments where the STING agonist is part of an ADC of formula IA, the ADC ("compound G") has the following structure: [ka] has.

[0129] Examples of cytokines that can be used in the methods, uses, combination therapies and mixtures disclosed herein include various interleukins, such as human interleukins IL-2, IL-7, IL-10, IL-12, IL-15, or combinations thereof. In certain embodiments, the interleukin is IL-2, IL-7, IL-10, IL-12, or combinations thereof. In some embodiments, the interleukin is IL-2, IL-12, IL-15, or combinations thereof. In one embodiment, the interleukin is IL-2. In another embodiment, the interleukin is IL-7. In another embodiment, the interleukin is IL-10. In another embodiment, the interleukin is IL-15. In a particular embodiment, the interleukin is IL-12.

[0130] In certain embodiments, an interleukin is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain embodiments, an interleukin is fused to an antibody to form a fusion protein. In certain embodiments, an interleukin is fused to an antibody fragment to form a fusion protein. In certain embodiments, an interleukin is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment.

[0131] Examples of interleukin fusion proteins with lumican that can be used in the present methods, uses and combination therapies are disclosed in PCT Publication WO 2020 / 068261, the contents of each of which are incorporated by reference.

[0132] In certain embodiments, the interleukin is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain embodiments, the interleukin is fused to a collagen binding protein. In certain embodiments, the collagen binding protein is lumican. In certain embodiments, the interleukin of the fusion protein is IL-12. In certain embodiments, the interleukin of the fusion protein is IL-2.

[0133] In certain embodiments, the interleukin is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain embodiments, the interleukin is fused to an IL-2 receptor alpha chain, a prostate specific antigen cleavage sequence, a matrix metalloproteinase cleavage sequence, or an alum binding peptide. In certain embodiments, the interleukin of the fusion protein is IL-12. In certain embodiments, the interleukin of the fusion protein is IL-2.

[0134] In certain embodiments, the interleukin is IL-12 and is fused to a protein to form a fusion protein, where the protein is an antibody or an antibody fragment. In certain embodiments, the interleukin is fused to an antibody to form a fusion protein. In certain embodiments, the interleukin is fused to an antibody fragment to form a fusion protein. In certain embodiments, the interleukin is fused to a protein to form a fusion protein, where the protein is not an antibody or an antibody fragment. In certain embodiments, the interleukin is fused to an antibody that recognizes a DNA / histone complex. In certain embodiments, the interleukin is fused to the human monoclonal IgG1 antibody NHS76. An example of IL-12 fused to the IgG1 antibody NHS76 is disclosed in Greiner et al., 2021, Immunotargets Ther. May 27; 10: 155-169. In certain embodiments, the interleukin can be fused to the IL-2 receptor alpha chain, a prostate specific antigen cleavage sequence, a matrix metalloproteinase cleavage sequence, or an antibody fragment scFv. Examples of such interleukin fusion proteins are disclosed in Puskas et al., 2011, Immunology, Jun; 133(2): 206-20. In certain embodiments, interleukins can be fused to alum-binding peptides (ABPs). Examples of interleukins bound to ABPs are disclosed in Agarwal et al., 2022, Nat Biomed Eng 6, 129-143; and Puskas et al., 2011, Immunology, Jun; 133(2): 206-20.

[0135] In certain embodiments, the interleukin is IL-2 and is fused to a protein to form a fusion protein, and the protein is an antibody or an antibody fragment. In certain embodiments, the interleukin is fused to an antibody to form a fusion protein. In certain embodiments, the interleukin is fused to an antibody fragment to form a fusion protein. In certain embodiments, the interleukin is fused to a protein to form a fusion protein, and the protein is not an antibody or an antibody fragment. In certain embodiments, the interleukin is a fusion protein.

[0136] In certain embodiments, the interleukin is a fusion protein, such as an Fc-fusion interleukin, such as an Fc-fusion IL-2, IL-7, IL-10, IL-12, IL-15, or a combination thereof. In certain embodiments, the interleukin is an Fc-fusion IL-2, IL-7, IL-10, IL-12, or a combination thereof. In some embodiments, the interleukin is an Fc-fusion IL-2, IL-12, IL-15, or a combination thereof. In one embodiment, the interleukin is an Fc-fusion IL-2. In another embodiment, the interleukin is an Fc-fusion IL-7. In another embodiment, the interleukin is an Fc-fusion IL-10. In another embodiment, the interleukin is an Fc-fusion IL-15. In certain embodiments, the interleukin is an Fc-fusion IL-12. In other embodiments, the interleukin is not a fusion protein.

[0137] As discussed above, the immune checkpoint inhibitor, when used in the methods and combination therapies disclosed herein, can be a PD-1 inhibitor, including anti-PD-1 antibodies, anti-PD-L1 antibodies, and anti-CTLA-4 antibodies, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

[0138] Examples of CTLA-4 inhibitors that can be used according to the present disclosure include, but are not limited to, ipilimumab (Yervoy®) and tremelimumab (ticilimumab), CBT-509, CS1002, BMS-986249, AGEN1181, AGEN1194, AGN2041, BA3071, ATOR-1015, ATOR-1144, ADV-1604, and BCD-145. In certain embodiments, the CTLA-4 inhibitor is an anti-CTLA-4 antibody selected from ipilimumab (Yervoy®) and tremelimumab. CTLA-4 inhibitors can generally be administered systemically or intratumorally, and in certain embodiments, the CTLA-4 inhibitor is an anti-CTLA-4 antibody administered intratumorally.

[0139] In some embodiments, when the immune checkpoint inhibitor is a CTLA-4 inhibitor, e.g., an anti-CTLA-4 antibody, the CTLA-4 inhibitor inhibits the interaction between CTLA-4 on T cells and CD80 (B7.1) or CD86 (B7.2) on antigen-presenting cells, such as dendritic cells or macrophages, in the tumor microenvironment.

[0140] Examples of PD-1 inhibitors that can be used in accordance with the present disclosure include, but are not limited to, pembrolizumab (Keytruda®), nivolumab (Opdivo®), cemiplimab (Libtayo®), AMP-224, AMP-514, or PDR001. PD-1 inhibitors can generally be administered systemically or intratumorally.

[0141] Examples of PD-L1 inhibitors that can be used in accordance with the present disclosure include, but are not limited to, atezolizumab (Tecentriq®), avelumab (Bavencio®), durvalumab (Imfinzi®), BMS-936559, or CK-301. PD-L1 inhibitors can generally be administered systemically or intratumorally.

[0142] 6.4. Further Treatment Methods In some embodiments, both the STING agonist and the cytokine are administered intratumorally into the patient's primary tumor. It has been found that when certain STING agonists (e.g., Compound A) are administered intratumorally into the primary tumor, tumor growth is suppressed not only at the site of the primary tumor, but also at the site of a distant tumor. Thus, such STING agonists exhibit an abscopal effect. Furthermore, STING agonists can enhance T cell priming and inflammation in the tumor microenvironment, both at the injection site and in distant regions. Cytokines, such as interleukins, can expand T cells. Thus, the present disclosure contemplates that the combination of STING agonist and cytokines results in increased and even synergistic proliferation and / or function of T cells, resulting in an even greater abscopal effect than administering the STING agonist in the absence of cytokines. However, the present disclosure provides such combination therapy including STING agonist and cytokines while reducing or limiting systemic toxicity from cytokines.

[0143] Thus, the present disclosure provides methods for treating both primary and distant tumors (including accessible and inaccessible cancers) by administering the combination therapies disclosed herein. In certain embodiments, the methods described herein treat tumors distal to the site of intratumoral administration of a STING agonist and / or cytokine.

[0144] The present disclosure also provides a method of treating a patient who is being treated systemically (e.g., intravenously, intramuscularly, subcutaneously, orally) or intratumorally with a STING agonist as described herein, comprising administering to the patient a cytokine as described herein. In certain embodiments, the cytokine is administered intravenously. In other embodiments, the cytokine is administered systemically (e.g., intravenously, intramuscularly, or subcutaneously). In some embodiments, the method further comprises administering to the patient a PD-1 inhibitor (e.g., an anti-PD-1 antibody), a PD-L1 inhibitor (e.g., an anti-PD-L1 antibody), or a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody) as described herein. In certain of these embodiments, the patient is afflicted with a cancer, such as those described herein. In some embodiments, the method of treating a patient treats the patient for cancer.

[0145] In certain embodiments, the combination therapy of the present disclosure can be used to treat cancer of the lung, bone, pancreas, skin, head, neck, uterus, ovaries, stomach, colon, breast, esophagus, small intestine, bowel, endocrine system, thyroid, parathyroid, adrenal gland, urethra, prostate, penis, testes, ureters, bladder, kidney, or liver. Additional cancers treatable by the combination therapy of the present disclosure include rectal cancer; cancer of the anal region; fallopian tube, endometrial, cervical, vaginal, vulvar, renal pelvis, and renal cell cancer; soft tissue sarcoma; myxoma; rhabdomyoma; fibroma; lipoma; teratoma; cholangiocarcinoma; hepatoblastoma; angiosarcoma; hemangioma; hepatoma; fibrosarcoma; chondrosarcoma; myeloma; chronic or acute leukemia; lymphocytic lymphoma; primary CNS lymphoma; CNS neoplasms; spinal axis tumors; squamous cell carcinoma; synovial sarcoma; malignant pleural mesothelioma; brain stem glioma; pituitary adenoma; bronchial adenoma; chondromatous hamartoma; dermoid; Hodgkin's disease; or a combination of one or more of the foregoing cancers.

[0146] In certain embodiments, the combination therapy of the present disclosure can be used to treat cancer that is refractory or non-responsive to immune checkpoint inhibitor therapy.In some cases, such cancers show low immunogenicity tumors.Such cancers can include, but are not limited to, prostate cancer, pancreatic cancer, lymphoma, head and neck cancer, kidney cancer, melanoma, colon cancer, breast cancer, and lung cancer.In certain embodiments, the cancer is selected from prostate cancer, pancreatic cancer, lymphoma, head and neck cancer, and kidney cancer.In some embodiments, the cancer is selected from melanoma, colon cancer, breast cancer, and lung cancer.

[0147] In certain embodiments, the combination therapy and methods of the present disclosure are useful for treating solid tumors, such as melanoma or tumors associated with cancers of the kidney, lung, liver, colon, pancreas, brain, head and neck, bladder, prostate, breast, ovary, cervix and thyroid. In some cases, the combination therapy and methods are useful for treating such tumors when the tumor is a primary tumor.

[0148] In other embodiments, the combination therapies and methods of the present disclosure are useful in treating metastatic cancers that may spread or have already spread to multiple organs.

[0149] In certain embodiments, the combination therapy of the present disclosure can be used to reduce tumor recurrence after initial treatment.

[0150] It will be appreciated by those skilled in the art that the methods disclosed herein are also disclosed as the corresponding "Swiss-type" or "EPC2000" equivalents. Thus, a method of treating a tumor in a cancer patient in need thereof, comprising co-administering an effective amount of a STING agonist and a cytokine to the patient, is understood to also disclose the use of a STING agonist in the manufacture of a medicament for treating a tumor in a cancer patient in need thereof, wherein treating comprises co-administering an effective amount of a STING agonist and a cytokine to the patient, or the use of a cytokine in the manufacture of a medicament for treating a tumor in a cancer patient in need thereof, wherein treating comprises co-administering an effective amount of a cytokine and a STING agonist to the patient. Similarly, the disclosure of the above method is understood to disclose a combination of a STING agonist and a cytokine for treating a tumor in a cancer patient in need thereof.

[0151] 6.5. Pharmaceutical Compositions and Kits The present disclosure further provides a pharmaceutical composition comprising a STING agonist, a cytokine and a pharma- ceutically acceptable carrier. In certain embodiments, the pharmaceutical composition is an injectable pharmaceutical composition, for example for intratumoral injection. In some embodiments, the pharma- ceutically acceptable carrier may comprise saline or phosphate-buffered saline (PBS). A particular advantage provided by the present disclosure is that the STING agonist and the cytokine can be administered intratumorally in a single composition. The administration of a single composition reduces the number of injections required and reduces the incidence of side effects associated with the administration of multiple doses of individual therapeutic agents. Furthermore, due to the synergistic effect observed when the cytokine is administered together with the STING agonist, the dose of either of the agents to achieve efficacy is less than the dose to achieve efficacy when either of the agents is administered as a monotherapy. This synergistic effect therefore results in a lower incidence of side effects such as irritation.

[0152] In other embodiments, the present disclosure provides a kit for treating a disease or disorder, including cancer, comprising a STING agonist and a cytokine. In certain embodiments, the kit provides a cytokine formulated for intratumoral administration and a STING agonist formulated for intratumoral or systemic (e.g., intravenous, intramuscular, subcutaneous, or oral) administration. In other embodiments, the kit provides a STING agonist formulated for intratumoral administration and a cytokine formulated for intratumoral or systemic (e.g., intravenous, intramuscular, or subcutaneous) administration. In some embodiments, both the STING agonist and the cytokine are formulated for intratumoral administration.

[0153] In certain embodiments, the kit further comprises a PD-1 inhibitor (e.g., an anti-PD-1 antibody), a PD-L1 inhibitor (e.g., an anti-PD-L1 antibody), or a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody). In some such embodiments, the PD-1 inhibitor, PD-L1 inhibitor, or CTLA-4 inhibitor is formulated for intratumoral or systemic (e.g., intravenous, intramuscular, or subcutaneous) administration. In certain embodiments, both the cytokine and the STING agonist are formulated for intratumoral administration, and the PD-1 inhibitor, PD-L1 inhibitor, or CTLA-4 inhibitor is formulated for systemic (e.g., intravenous, intramuscular, or subcutaneous) administration. In other embodiments, the cytokine is formulated for intratumoral administration, and both the STING agonist and the PD-1 inhibitor, PD-L1 inhibitor, or CTLA-4 inhibitor are formulated for systemic (e.g., intravenous, intramuscular, subcutaneous, or oral) administration. In certain embodiments, both the cytokine and the PD-1 inhibitor, PD-L1 inhibitor or CTLA-4 inhibitor are formulated for intratumoral administration, and the STING agonist is formulated for systemic (e.g., intravenous, intramuscular, subcutaneous, or oral) administration. In some embodiments, both the STING agonist and the PD-1 inhibitor, PD-L1 inhibitor or CTLA-4 inhibitor are formulated for intratumoral administration, and the cytokine is formulated for systemic (e.g., intravenous, intramuscular, or subcutaneous) administration. In other embodiments, the STING agonist, the cytokine and the PD-1 inhibitor, PD-L1 inhibitor or CTLA-4 inhibitor are all formulated for intratumoral administration.

[0154] 6.6. Dosing regimen The dosage of STING agonists varies depending on the particular STING agonist and the route of administration. In general, for systemic or intratumoral administration, STING agonists can be administered at a dose ranging from 1 to 1000 μg / kg. For oral administration, STING agonists can be administered at a dose ranging from 5 to 5000 μg / kg.

[0155] In certain embodiments, when the STING agonist is a 2'3'-cGAMP analog such as Compound A, the STING agonist can be administered intratumorally or systemically in the range of 1-100 μg / kg. For example, 2'3'-cGAMP analogs such as Compound A can be administered in the range of 1-10 μg / kg, 5-10 μg / kg, 5-20 μg / kg, 5-30 μg / kg, 5-40 μg / kg, 5-50 μg / kg, 10-20 μg / kg, 10-30 μg / kg, 10-40 μg / kg, 10-50 μg / kg, 15-20 μg / kg, 15-40 μg / kg, 20-30 μg / kg, 20-40 μg / kg, 20-50 μg / kg, 30- It can be administered to cancer patients in the ranges of 40μg / kg, 30-50μg / kg, 5-75μg / kg, 10-75μg / kg, 15-75μg / kg, 20-75μg / kg, 25-75μg / kg, 35-75μg / kg, 5-100μg / kg, 10-100μg / kg, 15-100μg / kg, 20-100μg / kg, 25-100μg / kg, 35-100μg / kg, or 50-100μg / kg.

[0156] In some embodiments, the 2'3'-cGAMP analog, such as Compound A, can be administered to the cancer patient at a dose ranging from 10 to 6,500 μg, such as 50 to 6,500 μg, for example, in a single or divided dose. In certain embodiments, the 2'3'-cGAMP analog, such as Compound A, can be administered to the cancer patient at a dose ranging from 100 to 3,000 μg, for example, in a single or divided dose. In other embodiments, the 2'3'-cGAMP analog, such as Compound A, can be administered to the cancer patient at a dose ranging from 100 to 1,200 μg, for example, in a single or divided dose. For example, 2'3'-cGAMP analogs such as compound A may be administered in the following doses: 10-50 μg, 10-100 μg, 10-200 μg, 50-200 μg, 100-200 μg, 100-400 μg, 100-500 μg, 100-800 μg, 200-400 μg, 400-600 μg, 400-800 μg, 100-1,000 μg, 250-1,000 μg, It can be administered to cancer patients in the following ranges: 500-1,000 μg, 500-3,000 μg, 1,000-3,000 μg, 500-4,500 μg, 1,000-4,500 μg, 500-6,500 μg, 1,000-6,500 μg, 2,000-6,500 μg, 3,000-6,500 μg, or 4,500-6,500 μg.

[0157] In embodiments involving administration of a priming dose and a maintenance dose of a 2'3'-cGAMP analog, such as Compound A, the priming dose can be administered to a cancer patient at a dosage ranging from 10 to 1,000 μg. For example, the priming dose of a 2'3'-cGAMP analog, such as Compound A, can be administered to a cancer patient at a dosage ranging from 10 to 20 μg, 10 to 40 μg, 10 to 50 μg, 10 to 80 μg, 20 to 40 μg, 40 to 60 μg, 40 to 80 μg, 50 to 100 μg, 100 to 200 μg, 100 to 300 μg, 100 to 500 μg, 200 to 500 μg, 200 to 800 μg, 200 to 1,000 μg, 500 to 800 μg, or 500 to 1,000 μg. In certain embodiments, a priming dose of a 2'3'-cGAMP analog, such as Compound A, can be administered to a cancer patient at a dosage ranging from 0.15-20 μg / kg, e.g., 0.15-1 μg / kg, 0.25-1 μg / kg, 0.5-1 μg / kg, 0.5-2 μg / kg, 1-3 μg / kg, 1-5 μg / kg, 2-5 μg / kg, 2-7 μg / kg, 1-10 μg / kg, 2-10 μg / kg, 3-10 μg / kg, 5-10 μg / kg, 5-15 μg / kg, 10-20 μg / kg, or 15-20 μg / kg.

[0158] In embodiments involving administration of a priming dose and a maintenance dose of a 2'3'-cGAMP analog, such as Compound A, the maintenance dose can be administered to a cancer patient at a dosage ranging from 100 to 3,000 μg. In other embodiments, the maintenance dose of a 2'3'-cGAMP analog, such as Compound A, can be administered to a cancer patient at a dosage ranging from 100 to 1,200 μg. For example, the maintenance dose of a 2'3'-cGAMP analog such as Compound A may be 50-200μg, 100-200μg, 100-400μg, 100-500μg, 100-800μg, 100-1,000μg, 200-400μg, 200-800μg, 200-1,200μg, 250-1,000μg, 400-600μg, 400-800μg, 400-1,200μg, 500-1,000μg, 500-1,200μg, 500-1,500μg, 500-1,500μg, 500-1,600μg, 500-1,700μg, 500-1,800μg, 500-1,900μg, 500-2,000μg, 500-2,000μg, 500-2,000μg, 500-3,000μg, 500-3,000μg, 500-4,000μg, 500-5,000μg, 500-6,000μg, 500-8,000μg, 500-1,900μg, 500-1,800μg, 500-1,900μg, 500-2,000μg, 500-3,000μg, 500-4,000μg, 500-5,000μg, 500-6,000μg, 500-1,900μg In certain embodiments, a maintenance dose of 2'3'-cGAMP analog such as Compound A can be administered to a cancer patient at a dose ranging from 1 to 100 μg / kg, for example 1 to 50 μg / kg.For example, the maintenance dose of a 2'3'-cGAMP analog such as Compound A may be 1-10μg / kg, 5-10μg / kg, 5-20μg / kg, 5-30μg / kg, 5-40μg / kg, 5-50μg / kg, 10-20μg / kg, 10-30μg / kg, 10-40μg / kg, 10-50μg / kg, 15-20μg / kg, 15-40μg / kg, 20-30μg / kg, 20-40μg / kg, 20-50μg / kg, 30-40μg / kg, 40-50μg / kg, 50-60μg / kg, 60-70μg / kg, 70-80μg / kg, 80-90μg / kg, 90-100μg / kg, 100-120μg / kg, 120-140μg / kg, 140-160μg / kg, 160-180μg / kg, 180-200μg / kg, 180-220μg / kg, 180-320μg / kg, 180-40μg / kg, 180-50μg / kg, 180-260μg / kg, 180-360μg / kg, 180-40μg / kg, 180-50μg / kg, 180-28 ... It can be administered to cancer patients in the ranges of 0-40μg / kg, 30-50μg / kg, 5-75μg / kg, 10-75μg / kg, 15-75μg / kg, 20-75μg / kg, 25-75μg / kg, 35-75μg / kg, 5-100μg / kg, 10-100μg / kg, 15-100μg / kg, 20-100μg / kg, 25-100μg / kg, 35-100μg / kg, or 50-100μg / kg.

[0159] In another embodiment, the dosing cycle includes administering a priming dose of a 2'3'-cGAMP analog such as Compound A on day 1 of the treatment cycle, followed by administration of a 2'3'-cGAMP analog such as Compound A under two maintenance dosing regimens. The first maintenance dosing regimen includes administering a maintenance dose of a 2'3'-cGAMP analog such as Compound A on days 8, 15 and 22 of the treatment cycle (i.e., the first days of weeks 2, 3 and 4), followed by a one week period (i.e., week 5) during which the patient is not administered the 2'3'-cGAMP analog. The second maintenance dosing regimen includes administering a 2'3'-cGAMP analog such as Compound A in a biweekly dosing regimen. For example, a 2'3'-cGAMP analog such as Compound A can be administered at the beginning of weeks 6 and 8 of the dosing cycle. In some embodiments, an additional biweekly dose of a 2'3'-cGAMP analog such as Compound A can be administered to the patient. For example, a 2'3'-cGAMP analog such as Compound A can be administered at week 10 of a dosing cycle, at weeks 10 and 12 of a dosing cycle, at weeks 10, 12 and 14 of a dosing cycle, at weeks 10, 12, 14 and 16 of a dosing cycle, etc.

[0160] Generally, for systemic or intratumoral administration, the amount of cytokine, such as interleukin, administered to a cancer patient may range from 0.001 μg / kg to 2 mg / kg, particularly 0.01 μg / kg to 1 mg / kg, depending on the cytokine or interleukin used.

[0161] For example, for IL-12, the amount of cytokine administered intratumorally to a cancer patient may range from 0.01 to 100 μg / kg, such as 0.01 to 0.1 μg / kg, 0.01 to 1 μg / kg, 0.05 to 0.5 μg / kg, 0.05 to 1 μg / kg, 0.1 to 0.5 μg / kg, 0.1 to 1 μg / kg, 0.5 to 5 μg / kg, 1 to 10 μg / kg, 5 to 50 μg / kg, or 10 to 100 μg / kg. In certain embodiments, the amount of IL-12 administered intratumorally to a cancer patient may range from 0.01, 0.05, 0.1, 0.5 or 1 μg / kg to 1.5, 5, 10, 25, 50 or 100 μg / kg. In some embodiments, the amount of IL-12 administered intratumorally to a cancer patient may range from 0.01, 0.05 or 0.1 μg / kg to 0.5, 1, 1.5 or 2 μg / kg.

[0162] In another embodiment, for IL-2, IL-7, IL-10 or IL-15, the amount of cytokine administered intratumorally to a cancer patient may range from 0.1 μg / kg to 1 mg / kg, for example, 0.1-1 μg / kg, 0.1-10 μg / kg, 0.5-5 μg / kg, 0.5-50 μg / kg, 1-10 μg / kg, 1-50 μg / kg, 1-100 μg / kg, 5-50 μg / kg, 5-100 μg / kg, 10-100 μg / kg, 50-500 μg / kg or 100 μg / kg to 1 mg / kg. In certain embodiments, the amount of IL-12 administered intratumorally to a cancer patient may range from 0.1, 0.5, 1, 1.5 μg / kg to 5, 10, 25, 50, 100, 500 or 1,000 μg / kg.

[0163] In methods described herein involving combination therapy and administration (e.g., co-administration) of a STING agonist and cytokine with an immune checkpoint inhibitor, where the immune checkpoint inhibitor is administered systemically, the immune checkpoint inhibitor can be administered to the cancer patient in an amount approved by a relevant regulatory agency, such as the U.S. Food and Drug Administration. For example, in some embodiments, the immune checkpoint inhibitor is a PD-1 inhibitor, such as pembrolizumab (Keytruda®) or nivolumab (Opdivo®), administered intravenously at a dose of 100-400 mg, such as 200 mg for pembrolizumab or 240 mg for nivolumab. In another example, the immune checkpoint inhibitor is a PD-L1 inhibitor, such as atezolizumab (Tecentriq®), avelumab (Bavencio®), or durvalumab (Imfinzi®), administered intravenously at a dose of 400-2,000 mg, such as 840-1680 mg for atezolizumab, 800 mg for avelumab, or 1500 mg or 10 mg / kg for durvalumab. In another example, the immune checkpoint inhibitor is a CTLA-4 inhibitor, such as ipilimumab (Yervoy®), administered intravenously at a dose of 2-5 mg / kg, such as 3 mg / kg.

[0164] 6.7. STING agonist dosing regimens with improved safety profiles In some embodiments, the STING agonist is administered in a dosing schedule that includes a priming dose followed by multiple maintenance doses. The priming dose refers to a dose that is administered at a lower dose than the maintenance dose to increase the body's tolerance to a particular active agent (e.g., STING agonist). It has been found that administering a priming dose of the STING agonist improves the safety profile of the STING agonist and allows the compound to be delivered at a higher maintenance dose level than would otherwise be tolerated. In general, the priming dose is less than the maintenance dose over the course of a given dosing cycle.

[0165] Thus, the present disclosure provides a novel dosing schedule for STING agonists based on a specific dosing schedule that requires administration of a priming dose followed by administration of a maintenance dose. In certain embodiments, the novel STING agonist dosing schedule described herein also includes co-administration with a cytokine as disclosed herein, and optionally one or more immune checkpoint inhibitors, particularly PD-1 inhibitors, PD-L1 inhibitors, or CTLA-4 inhibitors. The combination of the priming / maintenance dosing regimen of STING agonists with cytokines is expected to provide an improved therapeutic index.

[0166] Specific STING agonists that can be administered using the priming / maintenance dosing schedule of the present disclosure are described above. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound A. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is not Compound A. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound B. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound C. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound D. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound E. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound F. In some embodiments, the STING agonist administered with the priming / maintenance dosing schedule of the present disclosure is Compound G. In certain embodiments, the STING agonist administered in the priming / maintenance dosing schedule of the present disclosure is administered as part of an ADC, e.g., as described herein.

[0167] In some embodiments, the priming dose of the STING agonist can be administered at a 2-100th fold lower (by weight) than the respective maintenance dose in a given administration cycle. For example, the priming dose can be administered at a 2-70th fold lower, a 2-50th fold lower, a 2-30th fold lower, a 2-20th fold lower, a 2-10th fold lower, a 10th fold lower, a 10th fold lower, a 20th fold lower, or a 20th fold lower. In some embodiments, the priming dose can be administered at a 2-4th fold lower than the maintenance dose in a given cycle. In some embodiments, the priming dose can be administered at a 2-5th fold lower than the maintenance dose in a given cycle. In some embodiments, the priming dose can be administered at a half to eighth times less than the maintenance dose in a given cycle. In some embodiments, the priming dose can be administered at a third to fifth times less than the maintenance dose in a given cycle. In some embodiments, the priming dose can be administered at a third to eighth times less than the maintenance dose in a given cycle. In some embodiments, the priming dose can be administered at a quarter to eighth times less than the maintenance dose in a given cycle.

[0168] In some embodiments, the priming dose may be delivered at about 2 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 3 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 4 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 5 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 10 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 15 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 20 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at about 50 times less than the maintenance dose over the course of the administration cycle. In some embodiments, the priming dose may be delivered at a dose that is about 100-fold less than the maintenance dose over the course of an administration cycle.

[0169] It should be understood that the relative amounts of the above priming doses to the individual maintenance doses can be expressed as a ratio. For example, in an embodiment in which the priming dose is administered at a dose about half-times less than the maintenance dose, a dosing regimen is described that includes a 1:2 ratio of the priming dose to the individual maintenance dose. Thus, in certain embodiments, the disclosure provides a method of treating cancer comprising administering a combination of a STING agonist and a cytokine to a patient in need thereof, wherein the STING agonist is administered in a ratio of 1:2 to 1:100 of a priming dose to an individual maintenance dose, e.g., 1:2, 2:5, 3:8, 1:3, 2:7, 1:4, 1:5, 1:6, 1:8, 1:9, 1:10, 1:11, 1:12, 1:15, 1:20, 1:30, 1:50, 1:75 or 1:100 (ranges provided by these ratios, e.g., 1:2 to 1:3, 1:2 to 1:4, 1:2 to 1:5, 1:10 to 1:20, 1:20 to 1:30, 1:50, 1:75 or 1:100). In one embodiment, the method comprises administering the medicament according to a dosing regimen comprising administering the medicament according to any one of claims 1 to 10, including 1:2 to 1:8, 1:2 to 1:10, 1:4 to 1:8, 1:4 to 1:10, 1:4 to 1:15, 1:4 to 1:20, 1:8 to 1:10, 1:8 to 1:15, 1:8 to 1:20, 1:8 to 1:30, 1:10 to 1:15, 1:10 to 1:20, 1:10 to 1:30, 1:10 to 1:50, 1:20 to 1:30, 1:20 to 1:50, 1:20 to 1:75, 1:20 to 1:100, 1:30 to 1:50, 1:30 to 1:75, 1:30 to 1:100, 1:50 to 1:75, 1:50 to 1:100, or 1:75 to 1:100.

[0170] In some embodiments, the disclosure provides a method of treating cancer comprising administering a combination of a STING agonist and a cytokine to a patient in need thereof, wherein the STING agonist is administered according to a dosing regimen comprising a priming dose to individual maintenance dose ratio of 1:4 or 1:5, or a ratio within the range of 1:3 to 1:6, e.g., 1:3 to 1:5, 1:4 to 1:6, or 1:4 to 1:5. In other embodiments, the ratio is 1:8 or 1:10, or a ratio within the range of 1:5 to 1:15, e.g., 1:6 to 1:12, 1:8 to 1:12, 1:8 to 1:10, or 1:9 to 1:10.

[0171] In some embodiments, the priming dose can be administered on the first day of the treatment cycle, and the maintenance dose can be administered thereafter according to the above administration schedule.The first maintenance dose can be administered at least 2 days after the administration of the priming dose, i.e., on the third day.For example, the first maintenance dose can be administered 2, 3, 4, 5, 6, 7, 8, 9 or 10 days after the administration of the priming dose.

[0172] In one embodiment, the dosing cycle comprises administering a priming dose of a STING agonist on day 1 of the treatment cycle, followed by administration of maintenance doses of a STING agonist on days 8, 15 and 22 of the treatment cycle (i.e., the first days of weeks 2, 3 and 4), followed by a one week period (i.e., week 5) during which no STING agonist is administered to the patient. The maintenance dosing cycle may be repeated, or an altered maintenance dosing schedule may be used.

[0173] In another embodiment, the dosing cycle comprises administering a priming dose of a priming dose on day 1 of the treatment cycle, followed by administration of a maintenance dose of a STING agonist on days 8 and 22 of the dosing schedule (i.e., biweekly administration). The maintenance dosing cycle may be repeated or an altered maintenance dosing schedule may be used.

[0174] 7. Example Example 1. Combination study involving STING agonists, immune checkpoint inhibitors and cytokines The antitumor effects of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody) and the cytokines IL-2, IL-12 or IL-15 were investigated.

[0175] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously on the right flank (primary tumor) on day 0 and on the left flank (distal tumor) on day 2. Tumors were measured on day 7 and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Mice were subsequently mock-treated with PBS or treated intraperitoneally with 200 μg of anti-PD-L1 antibody or treated intratumorally on the right (primary) side with 1 μg of Compound A alone or in combination with IL-2 (5 μg), IL-12 (2 μg) or IL-15 (5 μg) on ​​days 7, 11, and 15. Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.2 mg / mL bovine serum albumin. Tumor volumes were measured every 2-3 days and survival was monitored daily. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH), and IL-2 (212-12), IL-12 (210-12) and IL-15 (210-15) were purchased from PeproTech (Rocky Hill, NJ).

[0176] Increased antitumor efficacy against primary tumors was observed with triple combinations of Compound A and anti-PD-L1 antibody with cytokines IL-2, IL-12, or IL-15 compared to treatment with dual combinations of Compound A and anti-PD-L1 antibody (Figure 1, Panel A).

[0177] A more dramatic increase in anti-tumor efficacy (i.e., abscopal effect) against distant untreated tumors was observed when using the triple combination of Compound A and anti-PD-L1 antibody with cytokines IL-2, IL-12, or IL-15, when compared to treatment with the dual combination of Compound A and anti-PD-L1 antibody (Figure 1, Panel A). This dramatically increased anti-tumor efficacy was also reflected in increased mouse survival with the triple combination (Figure 1, Panel B).

[0178] Example 2. Further combination studies involving STING agonists, immune checkpoint inhibitors and cytokines The antitumor effects of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and cytokines IL-7 or IL-10 were investigated.

[0179] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously on day 0 in the right flank (primary tumor) and on day 2 in the left flank (distal tumor). Tumors were measured on day 7, and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Mice were subsequently mock-treated with PBS, or treated intraperitoneally with 200 μg of anti-PD-L1 antibody, or treated intratumorally on the right side (primary) with 1 μg of Compound A alone or in combination with IL-7 (5 μg) or IL-10 (5 μg) on ​​days 7, 11, and 15. Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.2 mg / mL bovine serum albumin. Tumor volumes were measured every 2-3 days, and survival was monitored daily. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH), and IL-7 (217-17) and IL-10 (210-10) were purchased from PeproTech (Rocky Hill, NJ).

[0180] A significantly increased antitumor effect against primary tumors was observed with the triple combination of Compound A, anti-PD-L1 antibody and cytokines IL-7 or IL-10 compared to treatment with the dual combination of Compound A and anti-PD-L1 antibody (Figure 2, Panel A).

[0181] A modest increase in anti-tumor efficacy (i.e., abscopal effect) against distant untreated tumors was also observed when using the triple combination of Compound A with an anti-PD-L1 antibody and the cytokines IL-7 or IL-10, when compared to treatment with the dual combination of Compound A and an anti-PD-L1 antibody (Figure 2, Panel A). This increased anti-tumor efficacy was also reflected in increased mouse survival with the triple combination (Figure 2, Panel B).

[0182] Example 3. Further combination studies involving STING agonists, immune checkpoint inhibitors and IL-12 The antitumor efficacy of a triple combination of a STING agonist (compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody) and various doses of IL-12 (50 ng, 200 ng and 1 μg) was investigated in comparison with the dual combination of compound A and anti-PD-L1 antibody or IL-12 and anti-PD-L1 antibody.

[0183] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously on the right flank (primary tumor) on day 0 and on the left flank (distal tumor) on day 2. Tumors were measured on day 6 and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Then, on days 6 and 9, 12 and 15, mice were mock-treated with PBS or treated intraperitoneally with 200 μg of anti-PD-L1 antibody, or treated intratumorally at the right site (primary) with 1 μg of Compound A alone; 50 ng, 200 ng or 1 μg of IL-12 alone; or a combination of 1 μg of Compound A and 50 ng, 200 ng or 1 μg of IL-12. Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.2 mg / mL bovine serum albumin. Tumor volumes were measured every 2-3 days, and survival was monitored daily. Body weight change (%) was measured over time, starting 6 days before the first treatment. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH), and IL-12 (210-12) was purchased from PeproTech (Rocky Hill, NJ).

[0184] Increased anti-tumor effects against primary tumors were observed with the triple combination of Compound A, anti-PD-L1 antibody and IL-12 when compared with treatment with the dual combination of Compound A and anti-PD-L1 antibody or IL-12 and anti-PD-L1 antibody (Panel A of Figures 3A, 3B, and 3C, respectively).

[0185] When the triple combination of Compound A, anti-PD-L1 antibody, and IL-12 (50 ng or 200 ng) was used, a significant increase in anti-tumor efficacy (i.e., abscopal effect) was observed against distant untreated tumors when compared to treatment with the dual combination of Compound A, anti-PD-L1 antibody, or IL-12, and anti-PD-L1 antibody (Figure 3A and Figure 3B, Panel A, respectively). These enhanced anti-tumor efficacy was also reflected in increased mouse survival for the triple combination (Figure 3A and Figure 3B, Panel B, respectively). A significant abscopal effect was also observed for the triple combination of Compound A, anti-PD-L1 antibody, and IL-12 (1 μg) when compared to treatment with the dual combination of Compound A and anti-PD-L1 antibody (Figure 3C, Panel A). However, when compared with the double combination of IL-12 (1 μg) and anti-PD-L1 antibody, no additional abscopal effect was observed with this triple combination, likely due to the saturation effect of IL-12 at this higher dose. The triple combination of Compound A, anti-PD-L1 antibody, and IL-12 (1 μg) resulted in increased survival for certain mice, but also early death in other mice, likely due to toxicity associated with the higher dose of IL-12 in the triple combination (Figure 3C, Panel B).

[0186] All treatment groups except those receiving the combination with 1 μg IL-12 showed initial, transient and minimal (<5%) weight loss followed by rapid recovery, indicating low toxicity of the various combinations (Panel C of Figures 3A and 3B, respectively). However, combination treatment with 1 μg IL-12 caused significant weight loss (up to 15%), indicating increased mouse toxicity for this dose (Panel C of Figure 3C).

[0187] Example 4. Dose-finding study of STING agonists, immune checkpoint inhibitors and IL-12 The antitumor efficacy of a triple combination of a STING agonist (Compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody) and various doses of IL-12 (3 ng, 10 ng and 30 ng) was investigated.

[0188] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously on day 0 in the right flank (primary tumor) and on day 2 in the left flank (distal tumor). Tumors were measured on day 9 and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Mice were subsequently mock-treated with PBS or treated intraperitoneally with 200 μg of anti-PD-L1 antibody or treated intratumorally on the right side (primary) with 1 μg of Compound A alone or in combination with 3 ng, 10 ng or 30 ng of IL-12 on days 9 and 12, 15 and 18. Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.2 mg / mL bovine serum albumin. Tumor volumes were measured every 2-3 days and survival was monitored daily. Body weight change (%) was measured over time, starting 9 days before the first treatment. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH), and IL-12 (210-12) was purchased from PeproTech (Rocky Hill, NJ).

[0189] Compared to the dual combination of Compound A and anti-PD-L1 antibody, increased anti-tumor effects against primary tumors were observed when the triple combination of Compound A and anti-PD-L1 antibody with 3 ng, 10 ng or 30 ng of IL-12 was used (Figure 4, Panel A).

[0190] A dose-responsive anti-tumor effect against distant untreated tumors (i.e., the abscopal effect) was observed when using triple combinations of Compound A, anti-PD-L1 antibody, and 3 ng, 10 ng, or 30 ng of IL-12 (Figure 4, Panel A). This dose-responsive anti-tumor effect was also reflected in increased mouse survival for the various triple combinations (Figure 4, Panel B).

[0191] The initial transient and minimal (<5%) weight loss followed by rapid recovery indicated low toxicity of the various combinations (Figure 4, Panel C).

[0192] Example 5. Combination study involving STING agonists, immune checkpoint inhibitors and IL-12-Fc The antitumor effects of a combination of a STING agonist (compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and IL-12-Fc were investigated.

[0193] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously on the right flank (primary tumor) on day 0 and on the left flank (distal tumor) on day 2. Tumors were measured on day 9 and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Mice were then mock-treated or treated intratumorally on the right flank (primary) with 1 μg Compound A and 50 ng IL-12-Fc on day 9 and days 12 and 15, or treated intratumorally on the right flank (primary) with 200 μg anti-PD-L1 antibody intraperitoneally and a combination of 1 μg Compound A alone, 50 ng IL-12-Fc alone, or 1 μg Compound A and 50 ng IL-12-Fc. Cytokines and Compound A were administered in a vehicle of 50 μL PBS containing 0.2 mg / mL bovine serum albumin. Tumor volumes were measured every 2-3 days and survival was monitored daily. Body weight change (%) was measured over time starting 9 days before the first treatment. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH).

[0194] The IL-12-Fc protein contains two subunits of mouse interleukin-12 (p35 and p40), each fused to the Fc domain of human IgG1 with the following amino acid sequence: (SEQ ID NO:1) Mu IL-12 p35-linker-huIgG1 Fc(Whole):

number

number

[0195] The cDNA encoding each subunit was cloned into the pcDNA3.4-TOPO vector (Invitrogen, A14697). To express IL-12-Fc, both plasmids were transfected into CHO (ATCC (CCL-61) cells using ExpiFectamine™ CHO Transfection Kit (Gibco, A29129) according to the manufacturer's protocol. Seven days after transfection, the culture medium was collected and the IL-12-Fc protein was loaded onto a 5 ml HiTrap Protein A column (GE, 17-0403-01) on a Bio-Rad NGC chromatography system (Bio-Rad, NGC Quest 10, 7880001). The column was washed with 50 ml PBS and eluted with 25 ml 0.1 M glycine (pH 2.5). The eluate was concentrated and further purified on a gel filtration column (Bio-Rad, ENrich 650, 7801650) equilibrated with PBS.

[0196] Improved antitumor efficacy was observed in primary tumors for the triple combination of compound A, anti-PD-L1 antibody, and IL-12-Fc, compared with the double combination of compound A and anti-PD-L1 antibody, anti-PD-L1 antibody, and IL-12-Fc. The increase in antitumor efficacy (i.e., abscopal effect) against distant untreated tumors was also improved in the triple combination as well as the double combination of anti-PDL1 antibody and IL-12-Fc (Panel A of Figure 5). However, the improved antitumor efficacy for the triple combination is reflected in increased mouse survival (Panel B of Figure 5).

[0197] The initial transient and minimal (<5%) weight loss followed by rapid recovery indicated low toxicity of the various combinations (Figure 5, Panel C).

[0198] Example 6. Dose-finding study involving a STING agonist, immune checkpoint inhibitor and IL-12-Fc The antitumor effects of various doses of a STING agonist (Compound A) and a triple combination of an immune checkpoint inhibitor (anti-PD-L1 antibody) and IL-12-Fc were investigated.

[0199] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously on day 0 in the right flank (primary tumor) and on day 2 in the left flank (distal tumor). Tumors were measured on day 9 and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Mice were subsequently mock-treated with PBS or treated intraperitoneally with 200 μg of anti-PD-L1 antibody or treated intratumorally on the right side (primary) with 1 μg of Compound A alone or in combination with 5 ng, 17 ng or 50 ng of IL-12-Fc on days 9 and 12, 15 and 18. Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.2 mg / mL bovine serum albumin. Tumor volumes were measured every 2-3 days and survival was monitored daily. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH). IL-12-Fc protein was obtained as described above.

[0200] Comparable increased anti-tumor efficacy against primary and distant untreated tumors was observed for various triple combinations of Compound A, anti-PD-L1 antibody, and IL-12-Fc compared to the dual combination of Compound A and anti-PD-L1 antibody (Figure 6, Panel A). The improved anti-tumor efficacy for the triple combinations was also reflected in increased mouse survival compared to the dual combination of Compound A and anti-PD-L1 antibody (Figure 6, Panel B).

[0201] The initial transient and minimal (<5%) weight loss followed by rapid recovery indicated low toxicity of the various combinations (Figure 6, Panel C).

[0202] Example 7. Combination studies involving STING agonists, immune checkpoint inhibitors and interleukins IL-12-Fc or mIL-12-MSA-lumican The antitumor effects of a STING agonist (Compound A) and a combination of an immune checkpoint inhibitor (anti-PD-L1 antibody) with IL-12-Fc or mIL-12-MSA-lumican were investigated.

[0203] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously in the right flank (primary tumor) on day 0 and in the left flank (distal tumor) on day 2. Tumors were measured on day 9 and mice were re-grouped so that each group had a similar mean tumor volume. Each group contained 5 mice. Subsequently, on days 9 and 12 and 15, mice were mock-treated with PBS or treated intraperitoneally with 200 μg of anti-PD-L1 antibody or treated intratumorally in the right flank (primary) with 1 μg of Compound A alone or in combination with IL-12-Fc (30 ng) or mIL-12-MSA-lumican (20 ng, 60 ng or 200 ng). Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.5% mouse serum. Tumor volumes were measured every 2-3 days and survival was monitored daily. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH).

[0204] The fusion protein, called mIL-12-MSA-lumican, contains (from N-terminus to C-terminus) mouse interleukin-12, mouse serum albumin, and lumican, a collagen-binding moiety that anchors the molecule to tumors. To express this protein, the encoding cDNA was cloned into the pcDNA3.4-TOPO vector (Invitrogen, A14697) and transfected into CHO cells using the ExpiFectamine™ CHO Transfection Kit (Gibco, A29129) according to the manufacturer's protocol. Seven days after transfection, the culture medium was collected and loaded onto a 5 ml HisTrap Excel column (Cytiva, 17371206) on a Bio-Rad NGC chromatography system (Bio-Rad, NGC Quest 10, 7880001). The column was washed with 50 ml of PBS and eluted with 25 ml of 0.5 M imidazole in 50 mM Tris-HCl solution (pH 8.0). The eluate was concentrated and further purified on a gel filtration column (Bio-Rad, ENrich 650, 7801650) equilibrated with PBS. (SEQ ID NO:3) Amino acid sequence of mIL12-MSA-lumican:

number

[0205] The double combination of compound A and anti-PD-L1 antibody showed anti-tumor effect in primary tumors, but only limited anti-tumor effect in distant tumors. In contrast, the triple combination of compound A and anti-PD-L1 antibody and IL-12-Fc or mIL-12-MSA-lumican showed anti-tumor effect in primary tumors and distant tumors (Panel A of Figure 7). The survival of mice was shown to be dose-dependent in the group treated with mIL-12-MSA-lumican (Panel B of Figure 7).

[0206] All triple combination treatment groups showed comparable body weight changes, similar to the patterns observed in the mock-treated groups or the dual combination of Compound A and anti-PD-L1 antibody, indicating low toxicity of the various combinations (Figure 7, Panel C).

[0207] Example 8. Further combination studies involving STING agonists, immune checkpoint inhibitors and interleukin mIL-12-MSA-lumican We investigated the antitumor effects of various combinations of a STING agonist (compound A), an immune checkpoint inhibitor (anti-PD-L1 antibody), and interleukin mIL-12-MSA-lumican.

[0208] Female C57BL6 mice (Jackson Laboratory) aged 7–8 weeks were incubated for 10 6 B16F10 (ATCC CRL-6475) melanoma cells were implanted subcutaneously into the right flank (primary tumor) on day 0 and into the left flank (distal tumor) on day 2. Tumors were measured on day 9, and mice were reassigned so that each group had a similar mean tumor volume. Each group contained 5 mice. Subsequently, on days 9 and 12 and 15, mice were mock treated with PBS or 1) treated intratumorally in the right flank (primary) with 60 ng mIL-12-MSA-lumican and intraperitoneally with 200 μg anti-PD-L1 antibody; 2) treated intratumorally in the right flank (primary) with 1 μg Compound A and intraperitoneally with 200 μg anti-PD-L1 antibody; 3) treated intratumorally in the right flank (primary) with 1 μg Compound A and intratumorally in the right flank (primary) with 60 ng mIL-12-MSA-lumican; or 4) treated intratumorally in the right flank (primary) with 1 μg Compound A and 60 ng mIL-12-MSA-lumican and intraperitoneally with 200 μg anti-PD-L1 antibody. Cytokines and Compound A were administered in a vehicle of 50 μL of PBS containing 0.5% mouse serum. Tumor volumes were measured every 2-3 days and survival was monitored daily. Anti-PD-L1 antibody (BE0101) was purchased from BioXcell (Lebanon, NH).

[0209] The double combination of mIL-12-MSA-lumican and anti-PD-L1 antibody showed partial anti-tumor effect in primary tumors, but no anti-tumor effect in distant tumors. The double combination of compound A and anti-PD-L1 antibody showed anti-tumor effect in primary tumors, but only limited anti-tumor effect in distant tumors. The double combination of compound A and mIL-12-MSA-lumican showed anti-tumor effect in primary tumors, but reduced anti-tumor effect in distant tumors. The triple combination of compound A, anti-PD-L1 antibody and mIL-12-MSA-lumican showed anti-tumor effect in primary tumors and distant tumors (Panel A of Figure 8). Similarly, mouse survival was greatest in mice treated with the triple combination, relative to all three double combinations (Panel B of Figure 8).

[0210] Example 9. Tumor growth in naive and pretreated mice. Naïve female C57BL6 mice (Jackson Laboratory) (n=3) or mice that were tumor-free for 35 days after a complete treatment cycle with Compound A described in Example 7 and the triple combination of anti-PD-L1 antibody and mIL-12-MSA-lumican (20 ng, 60 ng, 200 ng) were administered 10 6 B16F10 (ATCC CRL-6475) melanoma cells were inoculated subcutaneously and tumor growth was evaluated over time. Naive mice showed tumor progression over time. In contrast, mice pre-treated with the triple combination of compound A, anti-PD-L1 antibody, and mIL-12-MSA-lumican (20 ng) showed slower tumor growth than naive mice. Mice pre-treated with the triple combination of compound A, anti-PD-L1 antibody, and mIL-12-MSA-lumican (60 ng) and compound A, anti-PD-L1 antibody, and mIL-12-MSA-lumican (200 ng) showed complete tumor suppression (Figure 9).

Claims

1. A method for treating a tumor in a cancer patient in need thereof, or (ii) a method for enhancing the anti-tumor response of a cancer patient, or (iii) a method for increasing the population or function of immune cells (such as T cells, NK cells, B cells, dendritic cells or macrophages, or combinations thereof) in a cancer patient, or (iv) a method for reducing tumor recurrence in a cancer patient in need thereof A STING agonist for use in The method comprising co-administering to the patient an effective amount of a STING agonist and a cytokine, The STING agonist or the cytokine being administered intratumorally to the patient, The STING agonist being compound A 【Chemical 1】 or a pharmaceutically acceptable salt thereof, STING agonist.

2. The STING agonist according to claim 1, further comprising co-administering to the patient an effective amount of an immune checkpoint inhibitor together with the STING agonist and the cytokine.

3. (i) The STING agonist is administered intratumorally to the patient, or the STING agonist is administered systemically to the patient, preferably, the STING agonist is administered intravenously, intramuscularly, subcutaneously, or orally to the patient, (ii) The cytokine is administered intratumorally to the patient, or the cytokine is administered systemically to the patient, preferably, the cytokine is administered intravenously, intramuscularly, or subcutaneously to the patient, or (iii) Both the STING agonist and the cytokine are administered intratumorally to the patient, The STING agonist according to claim 1 or 2.

4. The STING agonist according to claim 2 or 3, wherein the immune checkpoint inhibitor is administered intratumorally to the patient, or the immune checkpoint inhibitor is administered systemically to the patient, preferably, the immune checkpoint inhibitor is administered intravenously, intramuscularly, or subcutaneously to the patient.

5. The method according to any one of claims 2 to 4, wherein the immune checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor or a CTLA-4 inhibitor, preferably, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody.

6. A combination therapy for treating tumors in cancer patients, preferably those in need thereof, comprising a STING agonist and a cytokine, wherein the STING agonist or the cytokine is formulated for intratumoral administration to the patient, and the STING agonist is compound A 【Chemical 2】 or a pharmaceutically acceptable salt thereof, the combination therapy. **Claim 7**: (i) The STING agonist is formulated for intratumoral administration to the patient, or the STING agonist is formulated for systemic administration to the patient, preferably, the STING agonist is formulated for intravenous, intramuscular, subcutaneous, or oral administration to the patient, (ii) The cytokine is formulated for intratumoral administration to the patient, or the cytokine is formulated for systemic administration to the patient, preferably, the cytokine is formulated for intravenous, intramuscular, or subcutaneous administration to the patient, or (iii) Both the STING agonist and the cytokine are formulated for intratumoral administration to the patient. The combination therapy according to claim 6. **Claim 8** The combination therapy according to claim 6 or 7, further comprising an immune checkpoint inhibitor. **Claim 9** The immune checkpoint inhibitor is formulated for intratumoral administration to the patient, or the immune checkpoint inhibitor is formulated for systemic administration to the patient, preferably, the immune checkpoint inhibitor is formulated for intravenous, intramuscular, or subcutaneous administration to the patient. The combination therapy according to claim 8. **Claim 10** The immune checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor, preferably, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. The combination therapy according to claim 8 or 9. **Claim 11** A mixture comprising a STING agonist, a cytokine, and an immune checkpoint inhibitor, wherein the STING agonist is compound A 【Chemical Formula 3】 or a pharmaceutically acceptable salt thereof, the mixture. **Claim 12** The cytokine is an interleukin, preferably, (i) The interleukin is IL-2, IL-7, IL-10, IL-12, IL-15, or a combination thereof, more preferably IL-2, IL-7, IL-10, IL-12, or a combination thereof, more preferably IL-2, IL-12, IL-15, or a combination thereof, more preferably IL-12 or IL-2. (ii) The interleukin is fused to a protein to form a fusion protein, preferably the protein is an antibody, preferably the antibody is an antibody that recognizes a DNA / histone complex, or the protein is an antibody fragment, preferably the antibody fragment is an immunoglobulin scFv domain or an immunoglobulin Fc domain, or (iii) The interleukin is fused to a protein that is not an antibody or an antibody fragment, preferably the interleukin is fused to an IL-2 receptor alpha chain, a prostate-specific antigen cleavage sequence, a matrix metalloproteinase cleavage sequence, or a heparin-binding peptide, or the interleukin is fused to a collagen-binding protein, preferably the collagen-binding protein is lumican. The STING agonist, combination therapy, or mixture according to any one of claims 1 to 11.

13. The interleukin is a fusion protein, such as an Fc-fusion interleukin, preferably the interleukin is an Fc-fusion IL-12, or the interleukin is not a fusion protein. The STING agonist, combination therapy, or mixture according to claim 12.

14. The STING agonist is conjugated to an antibody, thus forming an antibody-drug conjugate (ADC). The STING agonist, combination therapy, or mixture according to any one of claims 1 to 13.

15. The mixture according to claim 11, further comprising human plasma.

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