WO2021086889A1 - Methods and compositions for treatment of cancer and infectious diseases - Google Patents

Abstract

The present invention relates to methods and compositions for the treatment of diseases with STING agonists. As described hereinafter, STING activation can be modulated to induce local versus systemic immune activation. At lower doses, intratumoral dosing of STING agonists can produce a robust tumor-specific T-cell response capable of ablating a target tumor at a site distal to the STING agonist administration. At higher doses, the distal anti-tumor response is reduced, and ablation of tumor-draining lymph nodes is observed. It is possible to inhibit this ablation of the tumor-draining lymph nodes by co-administration of a TNF antagonist. This co-administration of STING agonists and TNF antagonists can permit the administration of either increased concentrations of STING agonist or STING agonists having greater activity, while maintaining effective tumor control and generation of an anti-tumor adaptive immune response.

Description

METHODS AND COMPOSITIONS FOR TREATMENT OF CANCER AND

INFECTIOUS DISEASES RELATED APPLICATION

[0001] This application claims priority to United States Provisional Application Serial No. 62/927,639, filed October 29, 2019, which is hereby incorporated by reference in its entirety.

FIELD OF THE INVENTION

[0002] The present invention relates to the use of STING agonists in combination with TNF antagonists in the treatment of diseases such as cancer and infectious diseases.

BACKGROUND OF THE INVENTION

[0003] The following discussion of the background of the invention is merely provided to aid the reader in understanding the invention and is not admitted to describe or constitute prior art to the present invention.

[0004] Tumor necrosis factor (TNF, tumor necrosis factor alpha, TNFa, TNF-a, TNF- alpha, cachexin, or cachectin) is a cell signaling protein (cytokine) involved in systemic inflammation and is one of the cytokines that make up the acute phase reaction. It is produced chiefly by activated macrophages, although it can be produced by many other cell types such as CD4+ lymphocytes, NK cells, neutrophils, mast cells, eosinophils, and neurons.

[0005] TNF was originally identified in the 1970s as the serum mediator of innate immunity capable of inducing hemorrhagic necrosis in tumors and is involved in immune system homeostasis and the inflammatory response. Approximately a quarter of cancers are reportedly susceptible to direct cell killing mediated by soluble TNF. The anti-tumor activity of TNF-alpha is now well established and can be mediated through a variety of mechanisms including: (1) Cellular apoptosis by binding to tumor cell surface receptors; (2) T-effector cell activation (macrophage and NK cells) by blocking T-Reg cells that are immune suppressors; (3) Inducing tumor microvasculature collapse through endothelial cell modulation and disruption of neoangiogenesis including disruption of tumor vasculature; (4) Promoting TAM (tumor associated macrophages) to Ml anti-tumor stage; (5) Attraction and stimulation of neutrophils and monocytes to sites of activation for anti tumor immune responses; and (6) Downregulation of IL-13 expression by eosinophilic- like cells and inhibition of tumor induced monocyte differentiation to immunosuppressive phenotypes.

[0006] Stimulator of Interferon Genes (STING) (12), also known as MPYS and TMEM173, acts as a receptor for cyclic dinucleotides (CDNs), including cyclic-di- adenosine-monophosphate (CDA), cyclic-di-guanosine-monophosphate (CDG), and cyclic-GMP-AMP (cGAMP). Endogenously, STING activation by cyclic dinucleotides induces type 1 interferon (IFN) production through the cGAS-STING-TBKl-IRF3-IFN-I signaling axis. It has been reported in the literature that type I IFN signaling plays an important role in CDN-mediated cross-presentation. Firussi et ah, EBioMedicine 22: 100- 111, 2017 (doi: 10.1016/j.ebiom.2017.07.016). It has also been reported in the literature that CDNs activate STING-dependent, IFN-I-independent TNF-a production. Blaauboer et ah, J. Immunol. 192: 492-502, 2014. Development of compounds that induce STING signaling (referred to herein as “STING agonists”) has recently been the focus of intense research for the treatment of cancer and infectious diseases and as vaccine adjuvants. Sivick et ah, Cell Rep. 25: 3074-85, 2018.

[0007] When TNF-related pathways are aberrantly regulated, TNF becomes involved in autoimmune disorders such as rheumatoid arthritis, ankylosing spondylitis, inflammatory bowel disease, psoriasis, hidradenitis suppurativa and refractory asthma.

The pathological effects of TNF can be alleviated by administration of TNF antagonists, such as soluble TNF receptor fragments or anti-TNF antibodies. These agents bind circulating TNF, thus preventing the binding of TNF to its cognate receptors, and thus reducing TNF-a signaling. A number of TNF antagonists have been approved by the U.S. Food and Drug Administration for the treatment of these pathological disorders.

SUMMARY OF THE INVENTION

[0008] The present invention relates to methods and compositions for the treatment of diseases with STING agonists. As described hereinafter, STING activation can be modulated to induce local versus systemic immune activation. At lower doses, intratumoral dosing of STING agonists can produce a robust tumor- specific T-cell response capable of ablating a target tumor at a site distal to the STING agonist administration. At higher doses, the distal anti-tumor response is reduced, and ablation of tumor-draining lymph nodes is observed. It is possible to inhibit this ablation of the tumor-draining lymph nodes by co-administration of a TNF antagonist. This co administration of STING agonists and TNF antagonists can permit the administration of either increased concentrations of STING agonist or STING agonists having greater activity, while maintaining effective tumor control and generation of an anti-tumor adaptive immune response.

[0009] In a first aspect, the present invention provides methods for potentiating a T- cell-dependent immune response to one or more tumors in a mammal. These methods comprise: administering a STING agonist to the mammal in need thereof in an amount sufficient to activate a STING-dependent increase in cytokine production; and administering a TNF antagonist to the mammal in need thereof in an amount sufficient to inhibit TNF-dependent damage to one or more tumor-draining lymph nodes resulting from administration of the STING agonist, wherein T-cells are detectable in at least one of the one or more tumor-draining lymph nodes following administration of the STING agonist.

[0010] In some embodiments of the first aspect, the STING agonist is administered directly into one of the one or more tumors. In some embodiments, the STING agonist is administered directly into one of the one or more tumors in an amount sufficient to induce TNF-dependent damage to the associated tumor draining lymph node. In some embodiments, the amount is sufficient to induce TNF-dependent damage to the associated tumor-draining lymph node that would result in undetectable levels of T-cells in the associated tumor-draining lymph node.

[0011] In some embodiments of the first aspect and above embodiments thereof, the TNF antagonist is administered subcutaneously. In some embodiments wherein the STING agonist is injected directly into one of the one or more tumors, the TNF antagonist is administered subcutaneously between the STING agonist injected tumor and the associated tumor-draining lymph node, wherein T-cells are detectable in the associated tumor-draining lymph node. In some embodiments, T-cells specific to an antigen associated with the one or more tumors are detectable in the at least one tumor-draining lymph node. In some embodiments wherein the STING agonist is injected directly into one of the one or more tumors, T-cells specific to an antigen associated with the one or more tumors are detectable in one or more tumor-draining lymph nodes not associated with the STING agonist injected tumor.

[0012] In a second aspect, the present invention provides methods for treating one or more tumors in a mammal. These methods comprise: administering a STING agonist to the mammal in need thereof in an amount sufficient to activate a STING-dependent increase in cytokine production; and administering a TNF antagonist to the mammal in need thereof in an amount sufficient to increase the number of CD8⁺ T-cells present in one or more tumor draining lymph nodes, wherein the increase is relative to the number of CD8⁺ T-cells that would be present in the one or more tumor-draining lymph nodes following administration of the STING agonist in the absence of the TNF antagonist administration.

[0013] In some embodiments of the second aspect, the STING agonist is administered directly into one of the one or more tumors. In some embodiments, the STING agonist is administered directly into one of the one or more tumors in an amount sufficient to decrease the number of CD8+ T-cells present in the associated tumor-draining lymph node. In some embodiments, the amount is sufficient to decrease the number of CD8+ T- cells present in the associated tumor-draining lymph node to undetectable levels.

[0014] In some embodiments of the second aspect and above embodiments thereof, the TNF antagonist is administered subcutaneously. In some embodiments wherein the STING agonist is injected directly into one of the one or more tumors, the TNF antagonist is administered subcutaneously between the STING agonist injected tumor and the associated tumor-draining lymph node, wherein CD8⁺ T-cells are increased in the associated tumor-draining lymph node. In some embodiments, CD8⁺ T-cells are specific to an antigen associated with the one or more tumors. In some embodiments wherein the STING agonist is injected directly into one of the one or more tumors, CD8⁺ T-cells specific to an antigen associated with the one or more tumors are detectable in one or more tumor-draining lymph nodes not associated with the STING agonist injected tumor.

[0015] STING agonists are described in, for example, W02005030186,

W02007070598, WO2011003025, US20060040887, US20080286296, US20120041057, US20140205653, WO2014179335, WO2014179760, US20150056224, WO2015074145, WO2015185565, WO2016096174, W02016145102, W02017004499, W02017011444, W02017011622, W02017011920, WO2017027645, WO2017027646, WO2017075477, WO2017093933, WO2017123657, WO2017123669, WO2017161349, WO2017175147, WO2017175156, W02018009466, W02018045204, W02018009648, W02018009652, WO2018013887, W02018013908, W02018065360, W02018098203, W02018100558, WO2018138684, WO2018138685, WO2018156625, WO2018198076, WO2018198084, US20180002369, US20180092937, US20180093964, US2018237468, US2018230177, US2018273578, WO2018208667, WO2019023459, WO2019027858, US20190055277, US2019183917, US2019185509, US2019185510, US2019248828, WO2019046496, WO2019046498, W02019046500, W02019046511, WO2019051488, WO2019051489, WO2019074887, WO2019069275, WO2019069269, WO2019079261, W02019092660, WO2019118839, WO2019125974, WO2019165032, WO2019175776, WO2019180683, WO2019185477, WO2019185476, WO2019193533, WO2019193542, WO2019193543, WO2019211799, WO2019195063, WO2019195124, W02019100061, WO2019219820, WO20 19227007 , WO2019243823 W02020006432, WO2019238786, W02020010155, W02020010092, W02020010451, W02020014127, W02020016782, W02020028565, W02020028566, W02020072492, W02020057546, W02020074004, US20200131209 (hybrid), W02020124059, W02020117623, W02020117624, W02020117625, WO2020132549, WO2020132566, WO2020132582, WO2020146237, WO2020178768, WO2020178769, W02020178770, W02020194160, and W0202020532, the disclosures of which are hereby incorporated by reference as it relates to STING agonists.

[0016] In certain embodiments, the STING agonist is a cyclic dinucleotide (CDN). Preferred CDNs are described in, for example, W02005030186, WO2011003025, US20060040887, US20080286296, US20120041057, US20140205653, WO2014179335, WO2014179760, US20150056224, WO2015074145, WO2015185565, WO2016096174, W02016145102, W02017011444, WO2017027645, WO2017027646, WO2017075477, WO2017093933, WO2017123657, WO2017123669, WO2017161349, W02018009466, W02018045204, W02018009648, W02018009652, W02018065360, W02018098203, W02018100558, WO2018138684, WO2018138685, WO2018156625, WO2018198076, WO2018198084, US20180002369, US20180092937, US2018237468, US2018230177, US2018273578, WO2018208667, WO2019023459, US20190055277, US2019183917, US2019185509, US2019185510, US2019248828, WO2019046496, WO2019046498, W02019046500, W02019046511, WO2019051488, WO2019051489, WO2019074887, WO20 19069275, WO2019069269, WO2019079261, W02019092660, WO2019118839, WO2019125974, WO2019165032, WO2019175776, WO2019180683, WO2019185477, WO2019185476, WO2019193533, WO2019193542, WO2019193543, WO2019211799, W02020014127, W02020016782, W02020057546, W02020074004, US 20200131209, W02020117623, W02020117624, W02020117625, WO2020178768, WO2020178769, W02020178770, and W02020205323. Preferred STING agonists include ADU-S100 (also referred to as 2’3’-RR-(A)(A); 2’,5’-3’,5’-R/?Rp-bisphosphorothioate-CDA; or dithio-(i?p, i?p)-cyclic-[A(2’,5’)pA(3’,5’)p]); or 3’3’-RR-(2’F-A)(2’F-A) (also referred to as 3’,5’-3’,5’-RpR/?-bisphosphorothioate-di-F-CDA; or dithio-(R/?,R/?)-cyclic-[2’F- A(3 ’ ,5 ’ )p2 ’ F- A(3 ’ ,5 ’ )p] ) .

[0017] In certain other embodiments, the STING agonist is an amidobenzimidazole, such as a dimeric amidobenzimidazole STING agonist described in Ramanjulu et ah, Nature (doi: 10.1038/s41586-018-0705-y) and WO2017175147 and WO2017175156. In certain other embodiments, the STING agonist is a benzothiophene, for example as described in US20180093964, WO2019027858, WO2019195063, WO2019195124, and WO20 19219820.

[0018] Preferred STING agonists 2’3’-RR-(A)(A) (See US20150056224) and 3’3’- RR-(2’F-A)(2’F-A) (See W02016145102) are represented by the structures

respectively.

[0019] In certain embodiments, the TNF antagonist is an anti-TNF antibody or a soluble TNF receptor. Preferred TNF antagonists include, but are not limited to, Adalimumab (Humira®), Adalimumab-atto (Amjevita™), Certolizumab pegol (Cimzia®), Etanercept (Enbrel®), Etanercept-szzs (Erelzi®), Golimumab (Simponi®, Simponi Aria®), Infliximab (Remicade®), Infliximab-abda (Renflexis®) and Infliximab- dyyb (Inflectra®).

[0020] In certain embodiments, a TNF antagonist includes an inhibitor or antagonist of a pathway that leads to production of TNF, such as an inhibitor of Receptor-interacting serine/threonine -protein kinase 1 (RIPK1). In certain embodiments, the RIPK1 inhibitor is GSK2982772, GSK3145095, DNL747, DNL758 (SAR443122), RIPA 56, necrostatin- 1, or a compound as described in WO2017136727, W02017096301, WO2016027253, W02016101885, W02016101887, US20170152268, W02018109097, WO2018154520, WO2019072942, WO2019123219, WO2019204537, WO2019213445, WO2019224774, W02020020119, W02020043173, W02020044206, W02020088194, W02020103859, or W02020103884, the disclosures of which are hereby incorporated by reference as it relates to RIPK1 inhibition.

[0021] In preferred embodiments, the STING agonist is administered parenterally, and most preferably intratumorally or peritumorally. In preferred embodiments, the TNF antagonist is administered parenterally, and most preferably subcutaneously, intravenously, intraperitoneally, intratumorally or peritumorally, more preferably subcutaneously. In one embodiment, the TNF antagonist is administered subcutaneously between the STING agonist injected tumor and the associated tumor draining lymph node, preferably wherein the TNF antagonist is administered prior to the administration of the STING agonist. In one embodiment, the TNF antagonist is administered directly into the STING agonist injected tumor associated tumor draining lymph node, preferably wherein the TNF antagonist is administered prior to the administration of the STING agonist.

[0022] In certain embodiments, the method further comprises administration of a checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor is selected from the group consisting of a CTLA-4 pathway antagonist, a PD-1 pathway antagonist, a TIM-3 pathway antagonist, a VISTA pathway antagonist, a BTLA pathway antagonist, a LAG-3 pathway antagonist, and a TIGIT pathway antagonist. In some embodiments, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-Ll antibody, an anti- CTLA-4 antibody, an anti-TIM-3 antibody, an anti-BTLA antibody, an anti-B7-H3 antibody, an anti-CD70 antibody, an anti-CD40 antibody, an anti-CD 137 antibody, an anti-GITR antibody, an anti-OX40 antibody, an anti-KIR antibody or an anti-LAG-3 antibody. By way of example only, the immune checkpoint inhibitor is selected from the group consisting of nivolumab, pembrolizumab, pidilizumab, PDR001, MED 10680, cemiplimab, AMP-224, ipilimumab, BMS-936559, atezolizumab, durvalumab, and avelumab, or an antibody as described in W02018020476 or WO2018025178, the disclosures of which are hereby incorporated by reference as it relates to anti-PD-1 or anti-CTLA-4 antibodies, respectively. This list is not meant to be limiting. In a preferred embodiment, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-Ll antibody or an anti-CTLA-4 antibody, preferably wherein the antibody is administered intravenously.

[0023] Administration of the STING agonist may be by a single administration, or by multiple administrations over a period of days, weeks, or months. By way of example, the administration may be weekly, biweekly, or daily for two or more cycles. In certain embodiments, the STING agonist is administered weekly for two or more weeks. Preferably, the TNF antagonist is administered proximate in time to one or more STING agonist administration. Proximate in time as used herein refers to within 2 days of a STING agonist administration (i.e. any time between 2 days prior to or 2 days after the STING agonist administration, e.g. within about 1 minute, 2 minutes, 5 minutes, 10 minutes 1 hour, 2 hours, 5 hours, 12 hours, 24 hours 36 hours or 48 hours prior to or after the STING agonist administration, including administration of the TNF antagonist and STING agonist simultaneously). Most preferably, the first administration of the TNF antagonist is administered within 2 days prior to the first STING agonist administration, e.g. within 1 minute, 2 minutes, 5 minutes, 10 minutes 1 hour, 2 hours, 5 hours, 12 hours, 24 hours 36 hours or 48 hours prior to the first STING agonist administration.

[0024] In certain embodiments, the STING agonist is administered intratumorally on day 1, day 8, and day 15 of each 21 -day cycle, with the TNF antagonist administered subcutaneously every 2 weeks starting on day 1. In certain embodiments, the STING agonist is administered intratumorally on day 1 and day 8 of each 21 -day cycle, with the TNF antagonist administered subcutaneously every 2 weeks starting on day 1. In certain embodiments, the STING agonist is administered intratumorally on day 1, day 8, day 15, and day 22 of each 28-day cycle, with the TNF antagonist administered subcutaneously every 2 weeks starting on day 1. In certain embodiments, the STING agonist is administered intratumorally on day 1, day 8, and day 15 of each 28-day cycle, with the TNF antagonist administered subcutaneously every 2 weeks starting on day 1. In certain embodiments, the STING agonist is administered intratumorally on day 1 and day 8 of each 28-day cycle, with the TNF antagonist administered subcutaneously every 2 weeks starting on day 1. The scheduled dosing as described in this paragraph may vary, for example, each day of dosing can be plus or minus 1 or 2 days, and every two weeks can be every 14 day plus or minus 1 or 2 days. In a preferred embodiment, if the scheduled dosing of the STING agonist and the TNF antagonist falls on the same day, or is within 1 or 2 days of each other, the dosing of the TNF antagonist will be adjusted to occur prior to the dosing of the STING agonist. In certain embodiments, the STING agonist is injected into a tumor in an amount of 200-3600 pg, 400-3200 pg or 400-2400 pg, for example 200 pg, 400 pg, 600 pg, 800 pg, 1000 pg, 1200 pg, 1400 pg, 1600 pg, 1800 pg, 2000 pg, 2200 pg, 2400 pg, 2600 pg, 2800 pg, 3000 pg, 3200 pg, 3400 pg, or 3600 pg, and the TNF antagonist is administered subcutaneously in an amount of 10-200 mg, for example 10 mg, 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 110 mg. 120 mg, 130 mg, 140 mg, 150 mg, 160 mg, 170 mg, 180 mg, 190 mg, or 200 mg, preferably wherein the TNF antagonist is an antibody or an antibody fragment. In certain embodiments, wherein the method comprises dosing of an anti-PD-1 antibody, an anti- PD-L1 antibody or an anti-CTLA-4 antibody, the antibody is administered intravenously in an amount of 50-500 mg, for example 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, or 500 mg, preferably wherein the antibody is administered on day 1 of each cycle, e.g. 21-day cycle or 28-day cycle.

[0025] In preferred embodiments, following administration of the STING agonist into a tumor, CD8⁺ T-cells are detectable in the tumor-draining lymph node of the injected tumor, preferably wherein the CD8+ T-cells are specific to an antigen of the injected tumor. In most preferred embodiments, following administration of the STING agonist into a tumor, CD8⁺ T-cells are detectable in the tumor-draining lymph node of a distal tumor preferably wherein the CD8+ T-cells are specific to an antigen of the injected tumor.

[0026] It is to be understood that the invention is not limited in its application to the details of construction and to the arrangements of the components set forth in the following description or illustrated in the drawings. The invention is capable of embodiments in addition to those described and of being practiced and carried out in various ways. Also, it is to be understood that the phraseology and terminology employed herein, as well as the abstract, are for the purpose of description and should not be regarded as limiting.

[0027] As such, those skilled in the art will appreciate that the conception upon which this disclosure is based may readily be utilized as a basis for the designing of other structures, methods and systems for carrying out the several purposes of the present invention. It is important, therefore, that the claims be regarded as including such equivalent constructions insofar as they do not depart from the spirit and scope of the present invention.

BRIEF DESCRIPTION OF THE FIGURES

[0028] Fig. 1A-1B depict the CT26 tumor volume in B ALB/c mice following treatment with intratumoral injection of ADU-S100 at 10 pg (1A) or 500 pg (IB) with isotype control antibody or anti-CD8a antibody.

[0029] Fig. 2 depicts the percentage of tumor specific CD8+ T-cells in peripheral blood following intratumoral injection of CT26 tumors in BAFB/c mice with 1, 10, 100 or 500 pg of ADU-S100. [0030] Fig. 3 depicts the tumor specific CD8+ T-cells in the spleen, distal tumor draining lymph node, and tumor-draining lymph node associated with the injected tumor in BALB/c mice implanted with CT26 tumor cells in both rear flanks.

[0031] Fig. 4 shows the tumor-draining lymph node following injection of CT26 tumor bearing BALB/c mice with PBS or with 10 pg ADU-S100, 500 pg ADU-S100, or 500 pg ADU-S100 plus 200 pg of anti-TNF antibody.

[0032] Fig. 5 depicts the tumor specific IFNy+ CD8 T-cells in splenocytes of CT26 tumor implanted BALB/c mice treated with 20 pg or 500 pg of ADU-S100 and isotype control antibody or anti-TNF antibody.

[0033] Fig. 6 depicts the tumor specific IFNy+ CD8 T-cells in splenocytes of CT26 tumor implanted BALB/c mice treated with 500 pg of ADU-S100, 100 pg of Comp. 1, 100 pg of Comp. 2, or 100 pg of Comp. 3 and isotype control antibody or anti-TNF antibody.

DETAILED DESCRIPTION OF THE INVENTION [0034] Definitions

[0035] “Administration” as it applies to a mammal (e.g., human), mammalian subject, animal, veterinary subject, placebo subject, research subject, experimental subject, cell, tissue, organ, or biological fluid, refers without limitation to contact of an exogenous ligand, reagent, placebo, small molecule, pharmaceutical agent, therapeutic agent, diagnostic agent, or composition to the subject, cell, tissue, organ, or biological fluid, and the like. “Administration” can refer, e.g., to therapeutic, pharmacokinetic, diagnostic, research, placebo, and experimental methods. Treatment of a cell encompasses contact of a reagent to the cell, as well as contact of a reagent to a fluid, where the fluid is in contact with the cell. "Administration" also encompasses in vitro and ex vivo treatments, e.g., of a cell, by a reagent, diagnostic, binding composition, or by another cell.

[0036] The term “subject” or “individual” as used herein refers to a human or non human organism. Thus, the methods and compositions described herein are applicable to both human and veterinary disease. In certain embodiments, subjects are “patients,” i.e., living humans that are receiving medical care for a disease or condition. This includes persons with no defined illness who are being investigated for signs of pathology. Preferred are subjects who have an existing diagnosis of a particular cancer which is being targeted by the compositions and methods of the present invention. Preferred cancers for treatment with the methods described herein include, but are not limited to, a colorectal cancer, an aero-digestive squamous cancer, a lung cancer, a brain cancer, a liver cancer, a stomach cancer, a bladder cancer, a thyroid cancer, an adrenal cancer, a gastrointestinal cancer, an oropharyngeal cancer, an esophageal cancer, a head and neck cancer (e.g. head and neck squamous cell carcinoma), an ovarian cancer, a uterine cancer, a cervical cancer, an endometrial cancer, a breast cancer (e.g., a “triple negative” breast cancer), a non-Hodgkin’s lymphoma, a Merkle cell carcinoma, a melanoma, a prostate cancer, a pancreatic carcinoma, a renal carcinoma, a sarcoma (e.g. soft tissue sarcoma), a leukemia, a lymphoma and a multiple myeloma.

[0037] An “agonist,” as it relates to a ligand and receptor, comprises a molecule, combination of molecules, a complex, or a combination of reagents, that stimulates the receptor. For example, in the case of a receptor protein such as STING, an “agonist” includes a molecule that binds to the protein in a conformation that activates downstream signaling by the protein.

[0038] An “antagonist,” as it relates to a ligand and/or receptor, comprises a molecule, combination of molecules, or a complex, that inhibits, counteracts, downregulates, and/or desensitizes the receptor. “Antagonist” encompasses any reagent that inhibits a constitutive activity of the receptor. A constitutive activity is one that is manifest in the absence of a ligand/receptor interaction. “Antagonist” also encompasses any reagent that inhibits or prevents a stimulated (or regulated) activity of a receptor. By way of example, an antagonist of GM-CSF receptor includes, without implying any limitation, an antibody that binds to the ligand (GM-CSF) and prevents it from binding to the receptor, or an antibody that binds to the receptor and prevents the ligand from binding to the receptor, or where the antibody locks the receptor in an inactive conformation. “Antagonist” also includes any reagent that inhibits the production of a ligand, e.g. a molecule that inhibits in the pathway leading to production of the ligand, such as an inhibitor of an enzyme in the pathway. “Antagonist” as used herein may also refer to an inhibitor, e.g a TNF antagonist may also be referred to as a TNF inhibitor. [0039] “Specifically” or “selectively” binds, when referring to a ligand/receptor, nucleic acid/complementary nucleic acid, antibody/antigen, or other binding pair (e.g., a cytokine to a cytokine receptor) indicates a binding reaction that is determinative of the presence of the protein in a heterogeneous population of proteins and other biologies. Thus, under designated conditions, a specified ligand binds to a particular receptor and does not bind in a significant amount to other proteins present in the sample. Specific binding can also mean, e.g., that the binding compound, nucleic acid ligand, antibody, or binding composition derived from the antigen-binding site of an antibody, of the contemplated method binds to its target with an affinity that is often at least 25% greater, more often at least 50% greater, most often at least 100% (2-fold) greater, normally at least ten times greater, more normally at least 20-times greater, and most normally at least 100-times greater than the affinity with any other binding compound.

[0040] In a typical embodiment an antibody will have an affinity that is greater than about 10⁹ liters/mol, as determined, e.g., by Scatchard analysis (Munsen, et al. (1980) Analyt. Biochem. 107:220-239). It is recognized by the skilled artisan that some binding compounds can specifically bind to more than one target, e.g., an antibody specifically binds to its antigen, to lectins by way of the antibody’s oligosaccharide, and/or to an Fc receptor by way of the antibody’s Fc region.

[0041] “Treatment” or “treating” (with respect to a condition or a disease) is an approach for obtaining beneficial or desired results including and preferably clinical results. For purposes of this invention, beneficial or desired results with respect to a disease include, but are not limited to, one or more of the following: improving a condition associated with a disease, curing a disease, lessening severity of a disease, delaying progression of a disease, delaying relapse of a disease, alleviating one or more symptoms associated with a disease, increasing the quality of life of one suffering from a disease, and/or prolonging survival. Likewise, for purposes of this invention, beneficial or desired results with respect to a condition include, but are not limited to, one or more of the following: improving a condition, curing a condition, lessening severity of a condition, delaying progression of a condition, delaying relapse of a condition, alleviating one or more symptoms associated with a condition, increasing the quality of life of one suffering from a condition, and/or prolonging survival.

IB [0042] “Vaccine” encompasses preventative vaccines, including vaccines for prevention of the relapse of a disease. Vaccine also encompasses therapeutic vaccines, e.g., a vaccine administered to a mammal that comprises a condition or disorder associated with the antigen or epitope provided by the vaccine.

[0043] STING Agonists

[0044] STING (Stimulator of INterferon Genes), also known as TMEM173, MITA, MPYS, and ERIS, is an intracellular receptor that acts as a sensor of cytosolic nucleic acids. Activation of the STING pathway results in production of Type I interferons (mainly IFN-a and IFN-b) induced through the IRF3 (interferon regulatory factor 3) pathway. Activation of IRF3 is thought to be mediated by TBK1 that recruits and phosphorylates IRF3 thus forming an IRF3 homodimer capable of entering the nucleus to transcribe type I interferon and other genes (Liu S, et al. "Phosphorylation of innate immune adaptor proteins MAVS, STING, and TRIF induces IRF3 activation" Science. 2015: 2630-2637). TBK1 also activates the nuclear factor kappa-light-chain-enhancer of activated B cells pathway which leads to production of pro-inflammatory cytokines (IL- la, IL-Ib, IL-2, IL- 6, TNF-a, etc.), via the oncogenic transcription factor NF-KB. In addition, STING activates STAT6 (signal transducer and activator of transcription 6) to induce (Th2-type), increase (IL-12) or decrease (IL-10) production of various cytokines, including the chemokines CCL2, CCL20, and CCL26. Direct phosphorylation of STING on Ser366 upon activation has also been reported to occur through TBK1 or ULK1 (Corrales, L. et al "Direct activation of STING in the tumor microenvironment leads to potent and systemic tumor regression and immunity" Cell Reports, 2015, vol.ll : 1-13; Konno, H. et al. "Cyclic dinucleotides trigger ULK1 (ATG1) phosphorylation of STING to prevent sustained innate immune signaling" Cell, 2013, vol. 155: 688-698).

[0045] Cyclic-di-nucleotides (CDNs) such as cyclic-di-AMP (produced by Listeria monocytogenes and other bacteria) and its analogs cyclic-di-GMP and cyclic-GMP-AMP are recognized by the host cell as a pathogen associated molecular pattern (PAMP), which bind STING. The mammalian ligand that binds to and activates STING is (2’,5’-3’,5’) cyclic guanosine monophosphate-adenosine monophosphate (2’,3’-cGAMP) which is produced by the mammalian enzyme cGAS (also known as C6orfl50 or MB21D1). Activation of the STING pathway triggers an immune response that results in generation of both antigen- specific CD4⁺ and CD8⁺ T cells as well as pathogen- specific antibodies. [0046] Examples of cyclic purine dinucleotides are described in some detail in, for example: W02005030186, WO2011003025, US20060040887, US20080286296, US20120041057, US20140205653, WO2014179335, WO2014179760, US20150056224, WO2015074145, WO2015185565, WO2016096174, W02016145102, W02017011444, WO2017027645, WO2017027646, WO2017075477, WO2017093933, WO2017123657, WO2017123669, WO2017161349, W02018009466, W02018045204, W02018009648, W02018009652, W02018065360, W02018098203, W02018100558, WO2018138684, WO2018138685, WO2018156625, WO2018198076, WO2018198084, US20180002369, US20180092937, US2018237468, US2018230177, US2018273578, WO2018208667, WO2019023459, US20190055277, US2019183917, US2019185509, US2019185510, US2019248828, WO2019046496, WO2019046498, W02019046500, W02019046511, WO2019051488, WO2019051489, WO2019074887, WO2019069275, WO2019069269, WO20 19079261, W02019092660, WO2019118839, WO2019125974, WO2019165032, WO2019175776, WO2019180683, WO2019185477, WO2019185476, WO2019193533, WO2019193542, WO2019193543, WO2019211799, W02020014127, W02020016782, W02020057546, W02020074004, US 20200131209, W02020117623, W02020117624, W02020117625, WO2020178768, WO2020178769, W02020178770, and W02020205323.

[0047] Recently a class of small molecule (non-CDN-based) STING agonists have been described that are based on an amidobenzimidazole structure. See, e.g., Ramanjulu et ah, Nature (doi: 10.1038/s41586-018-0705-y) and WO2017175147, and WO2017175156. Another class of small molecule STING agonists has been described based on a benzothiophene structure, described in US20180093964, WO2019027858, WO2019195063, WO2019195124, and WO2019219820. Additional non-CDN small molecule STING agonists are described in W02007070598, W02017004499, W02017011622, W02017011920, WO2018013887, W02018013908, W02019100061, WO20 19227007 , WO2019243823 W02020006432, WO2019238786, W02020010155, W02020010092, W02020010451, W02020028565, W02020028566, W02020072492, W 02020124059, WO2020132549, WO2020132566, WO2020132582, and WO2020146237.

[0048] TNF Antagonists [0049] TNF antagonists typically function in one of two ways. First, antagonists will bind to or sequester the TNF molecule itself with sufficient affinity and specificity to substantially neutralize the TNF epitopes responsible for TNF receptor binding (hereinafter termed sequestering antagonists). Alternatively, TNF antagonists will compete with native TNF for the cell surface receptor or intracellular site to which TNF binds in the course of cell lysis (i.e., competitive antagonists). Both groups of antagonists are useful, either alone or together, in the methods described herein. Also included are compounds that inhibit a pathway involved in the production of TNF, for example reducing TNF production by inhibiting RIP1 kinase (also referred to as RIPK1. RIPK1 inhibitors that can be used as a TNF antagonist including, for example, GSK2982772, GSK3145095, or DNL747, DNL758 (SAR443122), RIPA 56, necrostatin-1, or a compound as described in WO2017136727, W02017096301, WO2016027253, W02016101885, W02016101887, US20170152268, W02018109097, WO2018154520, WO2019072942, WO2019123219, WO2019204537, WO2019213445, WO2019224774, W02020020119, W02020043173, W02020044206, W02020088194, W02020103859, or W02020103884.

[0050] Sequestering antagonists include TNF cell surface receptors and neutralizing antibodies to TNF, TNF-a neutralizing antibodies are readily raised in animals such as rabbits or mice by immunization with TNF-a in Freund's adjuvant followed by boosters as required. Immunized mice are particularly useful for providing sources of B cells for the manufacture of hybridomas, which in turn are cultured to produce large quantities of inexpensive anti-TNF-a monoclonal antibodies. Monoclonal antibodies have been obtained that exhibit high anti-TNF-a activity, on the order of 10¹⁰ liters/mole.

[0051] Competitive TNF antagonists preferably are TNF variants. Such variants include substitutions, deletions or insertions of residues (amino acid sequence variants) as well as other covalent modifications, e.g. alkylated TNF. However, amino acid sequence variants are most preferred as they are readily manufactured in recombinant cell culture and recovered as chemically uniform, homogeneous compositions. Further, covalent modifications are more likely to immunize the patient against TNF, an event that is undesirable from the standpoint of long term patient recovery.

[0052] Antagonistic TNF amino acid sequence variants are variants of the mature TNF amino acid sequence that are capable of inhibiting TNF cytotoxic activity, but which have substantially no cytotoxic activity of their own. Antagonistic TNF sequence variants will competitively bind to cell surface receptors or intracellular TNF recognition sites without exerting any substantial cytotoxic effect, thereby displacing TNF or preventing TNF from binding to or interacting with the cells.

[0053] Antagonistic TNF sequence variants are characterized by amino acid deletions, substitutions and/or insertions which lead to the substantial inactivation of TNF cytotoxic activity without substantially interfering with the ability of the antagonist to inhibit TNF cytotoxic activity. Typically, deletional mutations are preferred as they are less likely to induce an active anti-TNF immune response in patients to whom the antagonists are administered.

[0054] The sites within the human TNF-a molecule that are selected for sequence variation generally are located within about residues 10 to 66, 113 to 134 and 150 to 157 (numbered following Pennica et al., Proc. Natl. Aced. Sci. 88: 6060, 1985.). Residues within these regions are believed to be important for TNF-a cytotoxic activity. Since an amino terminal domain within about residues 1 to 40 is believed to be externalized and therefore likely to be involved in receptor binding, non-cytotoxic analogues that are capable of competing with endogenous TNF-a generally will be mutated at points in the C-terminal direction from this region. Deletions, substitutions or insertions of disparate residues in these regions (40-66, 110-134 and 150-157) are anticipated to yield TNF variants having reduced or no cytotoxic activity combined with varying degrees of antagonist activity. Those mutants having the optimum combination of antagonist activity and reduced cytotoxicity are identified by the screening assays described above.

[0055] Preferred TNF antagonists include, but are not limited to, Adalimumab, Adalimumab-atto, Certolizumab pegol, Etanercept, Etanercept-szzs, Golimumab, Infliximab, Infliximab-abda, and Infliximab-dyyb. TNF antagonists include RIPK1 inhibitors GSK2982772, GSK3145095, or DNL747, DNL758 (SAR443122), RIPA 56, necrostatin-1, or a compound as described in WO2017136727, W02017096301,

WO20 16027253, W02016101885, W02016101887, US20170152268, W02018109097, WO2018154520, WO2019072942, WO2019123219, WO2019204537, WO2019213445, WO20 19224774, W02020020119, W02020043173, W02020044206, W02020088194, W02020103859, or W02020103884. [0056] Additional Therapeutic Agents

[0057] Methods for co-administration with an additional therapeutic agent are well known in the art (Hardman, et al. (eds.) (2001) Goodman and Gilman’s The Pharmacological Basis of Therapeutics, 10th ed., McGraw-Hill, New York, NY; Poole and Peterson (eds.) (2001) Pharmacotherapeutics for Advanced Practice:A Practical Approach, Lippincott, Williams & Wilkins, Phila., PA; Chabner and Longo (eds.) (2001) Cancer Chemotherapy and Biotherapy, Lippincott, Williams & Wilkins, Phila., PA).

[0058] Additional agents that are beneficial to raising a T cell response may be used in combination with the STING agonist/TNF antagonist combination of the present invention. These include, without limitation, B7 costimulatory molecule, interleukin-2, interferon- y , GM-CSF, CTLA-4 antagonists, OX-40/OX-40 ligand, CD40/CD40 ligand, sargramostim, levamisol, vaccinia vims, Bacille Calmette-Guerin (BCG), liposomes, alum, Freund's complete or incomplete adjuvant, detoxified endotoxins, mineral oils, surface active substances such as lipolecithin, pluronic polyols, polyanions, peptides, and oil or hydrocarbon emulsions. Carriers for inducing a T cell immune response that preferentially stimulate a cytolytic T cell response versus an antibody response are preferred, although those that stimulate both types of response can be used as well. In cases where the agent is a polypeptide, the polypeptide itself or a polynucleotide encoding the polypeptide can be administered. The carrier can be a cell, such as an antigen presenting cell (APC) or a dendritic cell. Antigen presenting cells include such cell types as macrophages, dendritic cells and B cells. Other professional antigen-presenting cells include monocytes, marginal zone Kupffer cells, microglia, Langerhans' cells, interdigitating dendritic cells, follicular dendritic cells, and T cells. Facultative antigen- presenting cells can also be used. Examples of facultative antigen-presenting cells include astrocytes, follicular cells, endothelium and fibroblasts. The carrier can be a bacterial cell that is transformed to express the polypeptide or to deliver a polynucleoteide that is subsequently expressed in cells of the vaccinated individual. Adjuvants, such as aluminum hydroxide or aluminum phosphate, can be added to increase the ability of the vaccine to trigger, enhance, or prolong an immune response. Additional materials, such as cytokines, chemokines, and bacterial nucleic acid sequences, like CpG, a toll-like receptor (TLR) 9 agonist as well as additional agonists for TLR 2, TLR 4, TLR 5, TLR 7, TLR 8, TLR9, including lipoprotein, LPS, monophosphoryl lipid A, lipoteichoic acid, imiquimod, resiquimod, and other like immune modulators used separately or in combination with the described compositions are also potential adjuvants. Other representative examples of adjuvants include the synthetic adjuvant QS-21 comprising a homogeneous saponin purified from the bark of Quillaja saponaria and Corynebacterium parvum (McCune et ak, Cancer, 1979; 43:1619). It will be understood that the adjuvant is subject to optimization. In other words, the skilled artisan can engage in routine experimentation to determine the best adjuvant to use.

[0059] In an embodiment of the invention, the STING agonist/TNF antagonist combination is further combined with one or more additional pharmaceutically active components selected from the group consisting of an immune checkpoint inhibitor (e.g. CTLA-4, PD-1, TIM-3, VISTA, BTLA, LAG-3 and TIGIT pathway antagonists; PD-1 pathway blocking agents; PD-L1 inhibitors; including without limitation anti-PD-1 antibodies nivolumab, pembrolizumab, pidilizumab, cemiplimab, PDR001, or MEDI0680; PD-1 inhibitor AMP-224; anti-CTLA-4 antibody ipilimumab; and anti-PD- L1 antibodies BMS-936559, atezolizumab, durvalumab, or avelumab); a TLR agonist (e.g. CpG or monophosphoryl lipid A); an inactivated or attenuated bacteria that induce innate immunity (e.g., inactivated or attenuated Listeria monocytogenes ); a composition that mediates innate immune activation via Toll-like Receptors (TLRs), via (NOD)-like receptors (NLRs), via Retinoic acid inducible gene-based (RIG)-Tlike receptors (RLRs), via C-type lectin receptors (CLRs), or via pathogen-associated molecular patterns (PAMPs); and a chemotherapeutic agent. In some embodiments, the immune checkpoint inhibitor is selected from the group consisting of a CTLA-4 pathway antagonist, a PD-1 pathway antagonist, a TIM-3 pathway antagonist, a VISTA pathway antagonist, a BTLA pathway antagonist, a LAG-3 pathway antagonist, and a TIGIT pathway antagonist. In some embodiments, the immune checkpoint inhibitor is an anti-PD-1 antibody, an anti-PD-Ll antibody, an anti-CTLA-4 antibody, an anti-TIM-3 antibody, an anti-BTLA antibody, an anti-B7-H3 antibody, an anti-CD70 antibody, an anti-CD40 antibody, an anti-CD137 antibody, an anti-GITR antibody, an anti-OX40 antibody, an anti-KIR antibody or an anti-LAG-3 antibody. In some embodiments, the immune checkpoint inhibitor is selected from the group consisting of nivolumab, pembrolizumab, pidilizumab, PDR001, MEDI0680, cemiplimab, AMP-224, ipilimumab, BMS-936559, atezolizumab, durvalumab, and avelumab. In some embodiments, the TLR agonist is CpG or monophosphoryl lipid A. [0060] Pharmaceutical Compositions

[0061] The term “pharmaceutical” as used herein refers to a chemical substance intended for use in the cure, treatment, or prevention of disease and which is subject to an approval process by the U.S. Food and Drug Administration (or a non-U. S. equivalent thereof) as a prescription or over-the-counter drug product. Details on techniques for formulation and administration of such compositions may be found in Remington, The Science and Practice of Pharmacy 21^st Edition (Mack Publishing Co., Easton, PA) and Nielloud and Marti-Mestres, Pharmaceutical Emulsions and Suspensions: 2^nd Edition (Marcel Dekker, Inc, New York).

[0062] For the purposes of this disclosure, the pharmaceutical compositions may be administered by a variety of means including non-parenterally, parenterally, by inhalation spray, topically, or rectally in formulations containing pharmaceutically acceptable carriers, adjuvants and vehicles. "Non-parenteral administration" encompasses oral, buccal, sublingual, topical, transdermal, ophthalmic, otic, nasal, rectal, cervical, pulmonary, mucosal, and vaginal routes. The term parenteral as used here includes but is not limited to subcutaneous, intravenous, intramuscular, intratumoral, peritumoral, intraarterial, intradermal, intrathecal and epidural injections with a variety of infusion techniques. Intraarterial and intravenous injection as used herein includes administration through catheters. Administration via intracoronary stents and intracoronary reservoirs is also contemplated.

[0063] Intratumoral (directly into the tumor mass) or peritumoral (around the tumor mass) administration of a pharmaceutical may directly activate locally infiltrating dendritic cells, directly promote tumor cell apoptosis or sensitize tumor cells to cytotoxic agents. The term oral as used herein includes, but is not limited to oral ingestion, or delivery by a sublingual or buccal route. Oral administration includes fluid drinks, energy bars, as well as pill formulations.

[0064] Pharmaceutical compositions may be in any form suitable for the intended method of administration. When used for oral use for example, tablets, troches, lozenges, aqueous or oil suspensions, dispersible powders or granules, emulsions, hard or soft capsules, syrups or elixirs may be prepared. Compositions intended for oral use may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions may contain one or more agents including sweetening agents, flavoring agents, coloring agents and preserving agents, in order to provide a palatable preparation. Tablets containing a drug compound in admixture with non-toxic pharmaceutically acceptable excipient which are suitable for manufacture of tablets are acceptable. These excipients may be, for example, inert diluents, such as calcium or sodium carbonate, lactose, calcium or sodium phosphate; granulating and disintegrating agents, such as maize starch, or alginic acid; binding agents, such as starch, gelatin or acacia; and lubricating agents; such as magnesium stearate, stearic acid or talc. Tablets may be uncoated, or may be coated by known techniques including enteric coating, colonic coating, or microencapsulation to delay disintegration and adsorption in the gastrointestinal tract and/or provide a sustained action over a longer period. For example, a time delay material such as glyceryl monostearate or glyceryl distearate alone or with a wax may be employed.

[0065] Formulations for oral use may be also presented as hard gelatin capsules where the drug compound is mixed with an inert solid diluent, for example calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, such as peanut oil, liquid paraffin or olive oil.

[0066] Pharmaceutical compositions may be formulated as aqueous suspensions in admixture with excipients suitable for the manufacture of aqueous-suspensions. Such excipients include a suspending agent, such as sodium carboxymethylcellulose, methylcellulose, hydroxypropyl methylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia, and dispersing or wetting agents such as a naturally occurring phosphatide (e.g., lecithin), a condensation product of an alkylene oxide with a fatty acid (e.g., polyoxyethylene stearate), a condensation product of ethylene oxide with a long chain aliphatic alcohol (e.g., heptadecaethyleneoxycetanol), a condensation product of ethylene oxide with a partial ester derived from a fatty acid and a hexitol anhydride (e.g., polyoxyethylene sorbitan monooleate). The aqueous suspension may also contain one or more preservatives such as ethyl or n-propyl p-hydroxy-benzoate, one or more coloring agents, one or more flavoring agents and one or more sweetening agents, such as sucrose or saccharin.

[0067] Oil suspensions may be formulated by suspending the active ingredient in a vegetable oil, such as arachis oil, olive oil, sesame oil or coconut oil, or a mineral oil such as liquid paraffin. The oral suspensions may contain a thickening agent, such as beeswax, hard paraffin or cetyl alcohol. Sweetening agents, such as those set forth above, and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an antioxidant such as ascorbic acid.

[0068] Dispersible powders and granules of the disclosure suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, a suspending agent, and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those disclosed above. Additional excipients, for example sweetening, flavoring and coloring agents, may also be present.

[0069] The pharmaceutical compositions of the disclosure may also be in the form of oil-in- water emulsions. The oily phase may be a vegetable oil, such as olive oil or arachis oil, a mineral oil, such as liquid paraffin, or a mixture of these. Suitable emulsifying agents include naturally-occurring gums, such as gum acacia and gum tragacanth, naturally occurring phosphatides, such as soybean lecithin, esters or partial esters derived from fatty acids and hexitol anhydrides, such as sorbitan monooleate, and condensation products of these partial esters with ethylene oxide, such as polyoxyethylene sorbitan monooleate. The emulsion may also contain sweetening and flavoring agents.

[0070] The pharmaceutical compositions of the disclosure may be in the form of a sterile injectable preparation, such as a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent such as a solution in 1,3-butane-diol or prepared as a lyophilized powder. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile fixed oils may conventionally be employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid may likewise be used in the preparation of injectables. [0071] The amount of active ingredient that may be combined with the carrier material to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. For example, a time-release formulation intended for oral administration to humans may contain approximately 20 to 500 mg of active material compounded with an appropriate and convenient amount of carrier material which may vary from about 5 to about 95% of the total compositions. It is preferred that the pharmaceutical composition be prepared which provides easily measurable amounts for administration. Typically, an effective amount to be administered systemically is about 0.1 mg/kg to about 100 mg/kg and depends upon a number of factors including, for example, the age and weight of the subject (e.g., a mammal such as a human), the precise condition requiring treatment and its severity, the route of administration, and will ultimately be at the discretion of the attendant physician or veterinarian. It will be understood, however, that the specific dose level for any particular patient will depend on a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex and diet of the individual being treated; the time and route of administration; the rate of excretion; other drugs which have previously been administered; and the severity of the particular condition undergoing therapy, as is well understood by those skilled in the art.

[0072] As noted above, formulations of the disclosure suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient, as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid, or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The pharmaceutical compositions may also be administered as a bolus, electuary or paste.

[0073] A tablet may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free flowing form such as a powder or granules, optionally mixed with a binder (e.g., povidone, gelatin, hydroxypropyl ethyl cellulose), lubricant, inert diluent, preservative, disintegrant (e.g., sodium starch glycolate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose) surface active or dispersing agent. Molded tablets may be made in a suitable machine using a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropyl methylcellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with an enteric or colonic coating to provide release in parts of the gut other than the stomach.

[0074] Formulations suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavored base, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert base such as gelatin and glycerin, or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier.

[0075] Formulations for rectal administration may be presented as a suppository with a suitable base comprising for example cocoa butter or a salicylate.

[0076] Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.

[0077] Formulations suitable for parenteral administration include aqueous and non- aqueous isotonic sterile injection solutions which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described. A STING agonist compound for use herein, including those depicted by structure, includes any pharmaceutically acceptable salt thereof. It is to be understood that the compound or salt, includes solvates (particularly, hydrates) thereof, and may exist in crystalline forms, non-crystalline forms or a mixture thereof. The compound or salt, or solvates (particularly, hydrates) thereof, may also exhibit polymorphism (i.e., the capacity to occur in different crystalline forms). These different crystalline forms are typically known as “polymorphs.” It is to be understood that when named or depicted by structure, the disclosed compound, or solvates (particularly, hydrates) thereof, also include all polymorphs thereof. Polymorphs have the same chemical composition but differ in packing, geometrical arrangement, and other descriptive properties of the crystalline solid state. Polymorphs may have different physical properties such as density, shape, hardness, stability, and dissolution properties. Polymorphs typically exhibit different melting points, IR spectra, and X-ray powder diffraction patterns, which may be used for identification. One of ordinary skill in the art will appreciate that different polymorphs may be produced, for example, by changing or adjust the conditions used during the crystallization or recrystallization of the compound.

[0078] For solvates of compounds for use in this invention, or salts thereof, that are in crystalline form, the skilled artisan will appreciate that pharmaceutically acceptable solvates may be formed wherein solvent molecules are incorporated into the crystalline lattice during crystallization. Solvates may involve nonaqueous solvents such as ethanol, isopropanol, dimethyl sulfoxide, acetic acid, ethanolamine, and ethyl acetate, or they may involve water as the solvent that is incorporated into the crystalline lattice. Solvates wherein water is the solvent that is incorporated into the crystalline lattice are typically referred to as “hydrates.” Hydrates include stoichiometric hydrates as well as compositions containing variable amounts of water. The invention includes all such solvates.

[0079] Suitable pharmaceutically acceptable salts include those described by P. Heinrich Stahl and Camille G. Wermuth in Handbook of Pharmaceutical Salts:

Properties, Selection, and Use, 2^nd ed. (Wiley-VCH: 2011) and also Remington’s Pharmaceutical Sciences, 18^th ed. (Mack Publishing, Easton PA: 1990) and also Remington: The Science and Practice of Pharmacy, 19^th ed. (Mack Publishing, Easton PA: 1995).

[0080] Salts of a compound containing a basic amine or other basic functional group may be prepared by any suitable method known in the art, including treatment of the free bases with an inorganic acid, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like, or with an organic acid, such as acetic acid, trifluoroacetic acid, maleic acid, succinic acid, mandelic acid, fumaric acid, malonic acid, formic acid, alginic acid, pyruvic acid, oxalic acid, glycolic acid, salicylic acid, pyranosildyl acid, such as glucuronic acid or galacturonic acid, alphahydroxy acid, such as citric acid or tartaric acid, amino acid, such as aspartic acid or glutamic acid, aromatic acid, such as benzoic acid or cinnamic acid, sulfonic acid, such as p-toluenesulfonic acid, methanesulfonic acid, ethanesulfonic acid or the like. Examples of pharmaceutically acceptable salts include sulfates, pyrosulfates, bisulfates, sulfites, bisulfites, phosphates, chlorides, bromides, iodides, acetates, propionates, decanoates, caprylates, acrylates, formates, isobutyrates, caproates, heptanoates, propiolates, oxalates, malonates succinates, suberates, sebacates, fumarates, maleates, butyne-l,4-dioates, hexyne-1,6- dioates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, hydroxybenzoates, methoxybenzoates, phthalates, phenylacetates, phenylpropionates, phenylbutrates, citrates, lactates, glycolate, resinate, lactates, camsylates, tartrates, mandelates, and sulfonates, such as xylenesulfonates, methanesulfonates, propanesulfonates, naphthalene- 1 -sulfonates and naphthalene-2- sulfonates.

[0081] Salts of a compound containing a phosphate diester, phosphorothioate diester or other acidic functional group can be prepared by reacting with a suitable base. Pharmaceutically acceptable salts include, but are not limited to: pyridine, ammonium, piperazine, diethylamine, nicotinamide, formic, urea, sodium, potassium, calcium, magnesium, zinc, lithium, cinnamic, methylamino, methanesulfonic, picric, tartaric, triethylamino, dimethylamino, and tris(hydoxymethyl)aminomethane. Additional pharmaceutically acceptable salts are known to those skilled in the art.

[0082] Such a pharmaceutically acceptable salt may be made with a base which affords a pharmaceutically acceptable cation, which includes alkali metal salts (especially sodium and potassium), alkaline earth metal salts (especially calcium and magnesium), aluminum salts and ammonium salts, zinc, as well as salts made from physiologically acceptable organic bases such as diethylamine, isopropylamine, olamine, benzathine, benethamine, tromethamine (2-amino-2-(hydroxymethyl)propane-l,3-diol), morpholine, epolamine, piperidine, piperazine, picoline, dicyclohexylamine, N,N’- dibenzylethylenediamine, 2-hydroxyethylamine, tri-(2-hydroxyethyl)amine, chloroprocaine, choline, deanol, imidazole, diethanolamine, ethylenediamine, meglumine (N-methylglucamine), procaine, dibenzylpiperidine, dehydroabietylamine, glucamine, collidine, quinine, quinolone, erbumine and basic amino acids such as lysine and arginine.

[0083] If a compound containing a basic amine or other basic functional group is isolated as a salt, the corresponding free base form of that compound may be prepared by any suitable method known to the art, including treatment of the salt with an inorganic or organic base, suitably an inorganic or organic base having a higher pK_a than the free base form of the compound. Similarly, if a compound containing a phosphate diester, phosphorothioate diester or other acidic functional group is isolated as a salt, the corresponding free acid form of that compound may be prepared by any suitable method known to the art, including treatment of the salt with an inorganic or organic acid, suitably an inorganic or organic acid having a lower pK_a than the free acid form of the compound.

[0084] CDN STING agonists that include salts thereof can be described by structures wherein the -SH of the thiophosphate bonds are represented as -S with a corresponding cation to form salts of the compounds as described herein. For example, salts of 2’3’-RR- (A)(A) can be represented by the following structures:

wherein A^y+ represents a mono or polyvalent salt cation, and n and m are the lowest possible whole number for a given y. For example when A^y+ is monovalent, i.e., when y is 1, such as Na⁺, K⁺, NH4⁺, TEAH⁺ or the like, n is 1 and m is 2; when y is 2, such as Ca²⁺, Mg²⁺ and the like, n is 1 and m is 1; when y is 3, e.g., Al³⁺ or the like, n is 3 and m is 2. For example, salts of a monovalent or divalent salt cation can be represented as

, respectively, or in cases where n = 1, these can be represented without brackets, e.g., as

Alternatively, monovalent salts can be depicted with A⁺ adjacent each -S . For example, the sodium salt of 2’3’-RR-(A)(A) can be depicted as

[0085] The invention includes within its scope all possible stoichiometric and non-stoichiometric forms of the salts of STING agonists for use herein.

[0086] An effective amount of a pharmaceutical for a particular patient may vary depending on factors such as the condition being treated, the overall health of the patient, the route and dose of administration and the severity of side effects. Guidance for methods of treatment and diagnosis is available (see, e.g., Maynard, et al. (1996) A Handbook of SOPs for Good Clinical Practice, Interpharm Press, Boca Raton, FL; Dent (2001) Good Laboratory and Good Clinical Practice, Urch Publ., London, UK).

[0087] An effective amount may be given in one dose, but is not restricted to one dose. Thus, the administration can be two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, or more, administrations of a pharmaceutical composition. Where there is more than one administration of a pharmaceutical composition in the present methods, the administrations can be spaced by time intervals of one minute, two minutes, three, four, five, six, seven, eight, nine, ten, or more minutes, by intervals of about one hour, two hours, three, four, five, six, seven, eight, nine, ten, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 hours, and so on. In the context of hours, the term “about” means plus or minus any time interval within 30 minutes. The administrations can also be spaced by time intervals of one day, two days, three days, four days, five days, six days, seven days, eight days, nine days, ten days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, 21 days, and combinations thereof. The invention is not limited to dosing intervals that are spaced equally in time, but encompass doses at non-equal intervals.

[0088] A dosing schedule of, for example, once/week, twice/week, three times/week, four times/week, five times/week, six times/week, seven times/week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, and the like, is available for the invention. The dosing schedules encompass dosing for a total period of time of, for example, one week, two weeks, three weeks, four weeks, five weeks, six weeks, two months, three months, four months, five months, six months, seven months, eight months, nine months, ten months, eleven months, and twelve months. [0089] Provided are cycles of the above dosing schedules. The cycle can be repeated about, e.g., every seven days; every 14 days; every 21 days; every 28 days; every 35 days; 42 days; every 49 days; every 56 days; every 63 days; every 70 days; and the like. An interval of non dosing can occur between a cycle, where the interval can be about, e.g., seven days; 14 days; 21 days; 28 days; 35 days; 42 days; 49 days; 56 days; 63 days; 70 days; and the like. In this context, the term “about” means plus or minus one day, plus or minus two days, plus or minus three days, plus or minus four days, plus or minus five days, plus or minus six days, or plus or minus seven days.

[0090] Methods for co-administration with an additional therapeutic agent are well known in the art (Hardman, et al. (eds.) (2001) Goodman and Gilman’s The Pharmacological Basis of Therapeutics, 10th ed., McGraw-Hill, New York, NY; Poole and Peterson (eds.) (2001) Pharmacotherapeutics for Advanced Practice:A Practical Approach, Lippincott, Williams & Wilkins, Phila., PA; Chabner and Longo (eds.) (2001) Cancer Chemotherapy and Biotherapy, Lippincott, Williams & Wilkins, Phila., PA). Generally, co-administration or administration together indicates treating a subject with two or more agents, where the agents can be administered simultaneously or at different times. For example, such agents may be delivered to a single subject as separate administrations, which may be at essentially the same time or different times, and which may be by the same route or different routes of administration. Such agents may be delivered to a single subject in the same administration (e.g., same formulation) such that they are administered at the same time by the same route of administration.

[0091] As noted, the compositions of the present invention are preferably formulated as pharmaceutical compositions for parenteral or enteral delivery. A typical pharmaceutical composition for administration to an animal subject comprises a pharmaceutically acceptable vehicle such as aqueous solutions, non-toxic excipients, including salts, preservatives, buffers and the like. See, e.g., Remington's Pharmaceutical Sciences, 15th Ed., Easton ed. , Mack Publishing Co., pp 1405-1412 and 1461- 1487 (1975); The National Formulary XIV, 14th Ed., American Pharmaceutical Association, Washington, DC (1975) . Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oil and injectable organic esters such as ethyloleate. Aqueous carriers include water, alcoholic/aqueous solutions, saline solutions, parenteral vehicles such as sodium chloride, Ringer's dextrose, etc. Intravenous vehicles include

BO fluid and nutrient replenishers. Preservatives include antimicrobial agents, anti-oxidants, chelating agents and inert gases. The pH and exact concentration of the various components the pharmaceutical composition are adjusted according to routine skills in the art.

EXAMPLES

[0092] The following examples serve to illustrate the present invention. These examples are in no way intended to limit the scope of the invention.

[0093] Example 1: The CD8 T cell mediated anti-tumor activity of ADU-S100 is dependent on dose of compound.

[0094] Female BALB/c mice were purchased from Charles River Laboratories and acclimated a minimum of 72 hours prior to the initiation of studies. Mice were implanted with 2xl0⁵ CT26 tumor line, cultured at 37°C with 5% CO2 in RPMI-1640 containing 2 mM glutaMAX (GIBCO), 10% heat-inactivated FCS, 1% penicillin-streptomycin,

1% sodium pyruvate (GIBCO), 1 mM HEPES buffer (HyClone), 1% NEAA (Corning),

55 mM b-Mercaptoethanol (GIBCO). Mice were depleted of CD8 T cells by treatment with an anti-CD8 antibody twice per week, six days after tumor implantation. At day eight after implantation, tumors were injected with the indicated amount of ADU-S100 (also referred to as S100 in Figures).

[0095] As shown in Fig. 1, statistical analysis demonstrated that tumor control was reduced by 45.7% (P=0.0009) upon depletion of CD8 T cells where mice were treated with 10 pg of ADU-S100. In contrast, no difference in tumor control was observed in mice depleted of CD8 T cells, following treatment with 500 pg of ADU-S100.

[0096] Example 2: Intratumoral injection of ADU-S 100 is immunogenic but not at high doses

[0097] Six to eight-week-old BALB/c mice were implanted with CT26 tumor cells, subcutaneously in a single flank. Tumors were injected eight days after implantation with a dose range of ADU-S 100. Seven days after IT injection peripheral blood cells were stained with a tumor-peptide-MHC I tetramer (H-2Ld-AHl). As shown, while lower doses of ADU-S100: 1,10 and 100 mg, induced significantly increased tumor specific CD8 T cells, the 500 pg dose failed to do so.

[0098] Example 3: The injected tumor, draining lymph node is the site of CD8

T cell activation when tumors are injected with ADU-S100.

[0099] Six to eight-week-old BALB/c mice were implanted with CT26 tumor cells, subcutaneously in both rear flanks, generating two tumors per mouse. Nine days post implantation tumors were injected with 10 pg of ADU-S100. Organs from the indicated anatomical locations and times post IT injection were assessed for their frequencies of activated antigen specific T cells by staining cells with AHl-H-2Ld peptide-MHC tetramer and the activation markers Ki-67 and CD25.

[00100] As shown in Fig. 3, the greatest frequency of activated tetramer+ CD8 T cells was found in the injected side tumor draining lymph node (TDLN). This demonstrates that the reactivation site and location of T-cell expansion is in the tumor draining lymph. This demonstrates that the ADU-S100 injected tumor draining lymph node is critical for the generation of tumor specific CD8 T cells. ****= p value <0.0001 as determined by one-way Anova and Tukey-Kramer HSD test.

[00101] Example 4: High doses of STING agonist cause TNF-dependent disruption of the tumor draining lymph node

[00102] CT26 tumor bearing mice, implanted as above, were injected with anti-TNF antibody (Bioxcell, West Lebanon, NH; clone XT3.11, 200 ug dose) the night prior to IT injection of the indicated amounts of ADU-S100. Three days after IT injection, tumor draining lymph nodes were harvested, embedded in FFPE, sectioned and stained by hematoxylin and eosin. Arrows indicate areas of blood and dotted line demarks a region with low cellularity (see Fig. 4). These images suggest that the lack of anti-tumor CD8 T cell induction induced by the 500 pg dose of ADU-S100 is due to disruption of the tumor draining lymph. As shown in Fig 3., this is the location of CD8 T cell activation following ADU-S100 injection. Further, when 500 pg of ADU-S100 was injected in combination with TNF-a neutralizing antibody, the disruption of the tumor draining lymph node did not occur (see Fig. 4). [00103] Example 5: Anti-TNF treatment restores the immunogenicity of high dose ADU-S100.

[00104] Mice were implanted with CT26 tumor cells as above. Tumor bearing mice were treated with anti-TNF antibody or its isotype control (Bioxcell, West Febanon, NH; clone XT3.11, HRPN respectively, 500 ug dose) the evening prior to and the day of IT injection of ADU-S100 (20 pg or 500 pg). Seven days after IT injection, splenocytes were stimulated with the tumor-peptide AHlin the presence of brefeldin A, a Golgi inhibitor, and stained for Interferon gamma (IFNg).

[00105] As shown in Fig. 5, a significant increase in tumor- specific CD8 T cells was produced when mice injected intratumorally with 20 pg ADU-S100 and an isotype control antibody. Injection of 500 pg of ADU-S100 and the isotype control antibody did not induce many tumor specific CD8 T cells showing similar levels compared to the control injection of PBS. In contrast, injection of 500 pg of ADU-S100 and anti-TNF showed considerable increase in the number the tumor specific CD8 T cells (*= p value of 0.0492) compared to the isotype control as determined by a one-way ANOVA analysis and Bonferroni-Sidak’s multiple comparison test. These results indicate that TNF is detrimental to the induction of tumor specific CD8 T cells in the context of high doses of ADU-S100.

[00106] Example 6: Blocking TNF restores the immunogenicity of additional

CDN- STING agonists

[00107] BAFB/c mice were implanted with CT26hmeso tumor cells. Ten days later mice were injected intraperitoneally with anti-TNF antibody or its isotype control (200 pg XT3.11, HRPN respectively), followed by IT injection of the indicated CDN molecules six hours later. The CDNs used were ADU-S100; 3’3’-RR-(G)(G) (Comp. 1, see US20140205653); 2’3’-RR-(G)(2’F-A) (Comp. 2, see WO2017075477); and 2’3’-RR- (3’F-G)(LNA-A) (Comp. 3, see W02018009466). Antibodies were administered on the next day as well. On day 17, splenocytes were stimulated with the tumor-peptide AHlin the presence of brefeldin A, a Golgi inhibitor, and stained for Interferon gamma (IFNg). As shown, 500 pg of ADU-S100 or 100 pg doses of highly potent STING-agonists Comp. 1, 2 or 3, failed to induce tumor- specific CD 8 T cells much above the levels present in PBS injected mice for isotype control treated mice. In contrast the combination of anti-TNF and the indicated molecules showed an increase of tumor specific CD8 T cells above that present in PBS injected mice or the isotype control mice. While these results were not statistically significant, the anti-TNF- samples showed higher levels of tumor specific CD8 T cells for all four compounds.

[00108] Example 7. Treatment of patients with TNF inhibitor plus STING agonist and checkpoint inhibitor

[00109] Patients with non-Hodgkin’s lymphoma, melanoma, head and neck squamous cell carcinoma, Merkle cell carcinoma, soft tissue sarcoma, or breast cancer are administered ADU-S100, pembrolizumab, and adalimumab according to the following administration schedule: between 400-2400 pg (1.0 mL) ADU-S100 is administered intratumorally on Day 1 and 8 of each 21-day cycle. Dosing solution is an aqueous solution, e.g. a PBS solution, which can be diluted with sterile water to a desired concentration prior to injection;

40 mg (0.8 mL) adalimumab is delivered subcutaneously every 2 weeks for 12 weeks, provided that on Day 1 and subsequent days when ADU-S100 delivery will coincide with delivery of adalimumab, adalimumab will be administered prior to the administration of ADU-S100; and

200 mg (8.0 mL) pembrolizumab is administered intravenously every 3 weeks starting on Day 1.

[00110] It is to be understood that the invention is not limited in its application to the details of construction and to the arrangements of the components set forth in the following description or illustrated in the drawings. The invention is capable of embodiments in addition to those described and of being practiced and carried out in various ways. Also, it is to be understood that the phraseology and terminology employed herein, as well as the abstract, are for the purpose of description and should not be regarded as limiting.

[00111] As such, those skilled in the art will appreciate that the conception upon which this disclosure is based may readily be utilized as a basis for the designing of other structures, methods and systems for carrying out the several purposes of the present invention. It is important, therefore, that the claims be regarded as including such equivalent constructions insofar as they do not depart from the spirit and scope of the present invention.

[00112] While the invention has been described and exemplified in sufficient detail for those skilled in this art to make and use it, various alternatives, modifications, and improvements should be apparent without departing from the spirit and scope of the invention. The examples provided herein are representative of preferred embodiments, are exemplary, and are not intended as limitations on the scope of the invention.

Modifications therein and other uses will occur to those skilled in the art. These modifications are encompassed within the spirit of the invention and are defined by the scope of the claims.

[00113] It will be readily apparent to a person skilled in the art that varying substitutions and modifications may be made to the invention disclosed herein without departing from the scope and spirit of the invention.

[00114] All patent applications, patents, publications and other references mentioned in the specification are indicative of the levels of those of ordinary skill in the art to which the invention pertains and are each incorporated herein by reference. The references cited herein are not admitted to be prior art to the claimed invention.

[00115] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In the case of conflict, the present specification, including definitions, will control.

[00116] The use of the articles “a”, “an”, and “the” in both the description and claims are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. The terms “comprising”, “having”, “being of’ as in “being of a chemical formula”, “including”, and “containing” are to be construed as open terms (i.e., meaning “including but not limited to”) unless otherwise noted. Additionally whenever “comprising” or another open-ended term is used in an embodiment, it is to be understood that the same embodiment can be more narrowly claimed using the intermediate term “consisting essentially of’ or the closed term “consisting of’. [00117] The term “about”, “approximately”, or “approximate”, when used in connection with a numerical value, means that a collection or range of values is included. For example, “about X” includes a range of values that are ±20%, ±10%, ±5%, ±2%,

±1%, ±0.5%, ±0.2%, or ±0.1% of X, where X is a numerical value. In one embodiment, the term “about” refers to a range of values which are 10% more or less than the specified value. In another embodiment, the term “about” refers to a range of values which are 5% more or less than the specified value. In another embodiment, the term “about” refers to a range of values which are 1% more or less than the specified value.

[00118] Recitation of ranges of values are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. A range used herein, unless otherwise specified, includes the two limits of the range. For example, the terms “between X and Y” and “range from X to Y, are inclusive of X and Y and the integers there between. On the other hand, when a series of individual values are referred to in the disclosure, any range including any of the two individual values as the two end points is also conceived in this disclosure. For example, the expression “a dose of about 100 mg, 200 mg, or 400 mg” can also mean “a dose ranging from 100 to 200 mg”, “a dose ranging from 200 to 400 mg”, or “a dose ranging from 100 to 400 mg”.

[00119] The invention illustratively described herein suitably may be practiced in the absence of any element or elements, limitation or limitations which is not specifically disclosed herein. Thus, for example, in each instance herein any of the terms “comprising”, “consisting essentially of’ and “consisting of’ may be replaced with either of the other two terms. The terms and expressions which have been employed are used as terms of description and not of limitation, and there is no intention that in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically disclosed by preferred embodiments and optional features, modification and variation of the concepts herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this invention as defined by the appended claims. [00120] Other embodiments are set forth within the following claims.

Claims

We claim:

1. A method of potentiating a T-cell-dependent immune response to one or more tumors in a mammal, comprising: administering a STING agonist to the mammal in need thereof in an amount sufficient to activate a STING-dependent increase in cytokine production; and administering a TNF antagonist to the mammal in need thereof in an amount sufficient to inhibit TNF-dependent damage to one or more tumor-draining lymph nodes resulting from administration of the STING agonist, wherein T-cells are detectable in at least one of the one or more tumor-draining lymph nodes following administration of the STING agonist.

2. The method according to claim 1, wherein the STING agonist is a cyclic dinucleotide.

3. The method according to claim 1 or 2, wherein the STING agonist is administered intratumorally or peritumorally.

4. The method according to claim 3, wherein T-cells are detectable in the STING agonist treated tumor-draining lymph node.

5. The method according to one of claims 1-4, wherein the TNF antagonist is administered parenterally.

6. The method according to claim 5, wherein the TNF antagonist is administered intravenously, intraperitoneally, or subcutaneously.

7. A method of treating one or more tumors in a mammal, comprising: administering a STING agonist to the mammal in need thereof in an amount sufficient to activate a STING-dependent increase in cytokine production; and administering a TNF antagonist to the mammal in need thereof in an amount sufficient to increase the number of CD8⁺ T-cells present in at least one tumor-draining lymph node, wherein the increase is relative to the number of CD8⁺ T-cells that would be present in the at least one tumor-draining lymph node following administration of the STING agonist in the absence of the TNF antagonist administration.

8. The method according to claim 7, wherein the STING agonist is a cyclic dinucleotide.

9. The method according to claim 7, wherein the STING agonist is administered intratumorally or peritumorally.

10. The method according to claim 9, wherein the at least one tumor-draining lymph node is a STING agonist treated tumor-draining lymph node.

11. The method according to one of claims 7-10, wherein the TNF antagonist is administered parenterally.

12. The method according to claim 11, wherein the TNF antagonist is administered intravenously, intraperitoneally, or subcutaneously.

13. The method according to one of claims 1-12, wherein the TNF antagonist is selected from the group consisting of Adalimumab, Certolizumab, Etanercept, Golimumab, Infliximab, and biosimilars thereof.

14. The method according to one of claims 1-13, wherein the STING agonist is ADU-

S 100.

15. The method according to one of claims 1-14, wherein the method further comprises administration of a checkpoint inhibitor.

16. The method according to one of claims 1-15, wherein the TNF antagonist is administered prior to the STING agonist.

17. The method according to one of claims 1-15, wherein the STING agonist is administered approximately weekly for two or more weeks, and the TNF antagonist is administered within 2 days prior to each STING agonist administration.

18. A method according to one of claims 1-15, wherein the STING agonist is administered approximately weekly for two or more weeks, and the TNF antagonist is administered within 2 days prior to the first STING agonist administration and within 2 days prior to every other subsequent STING agonist administration.

19. The method according to one of claims 1-18, wherein tumor antigen specific CD8⁺T-cells are detectable in one or more of the tumor-draining lymph nodes following administration of the STING agonist.