Seneca Valley Virus Combination Therapy for Treating Checkpoint Inhibitor-Resistant Cancers

Abstract

Provided herein are compositions and methods using Seneca Valley Virus (SVV) or SVV derivatives in combination with checkpoint inhibitors to treat cancers that are refractory to checkpoint inhibitor treatment. Also provided herein are kits including Seneca Valley Virus (SVV) or SVV derivatives and checkpoint inhibitors to treat cancers that are refractory to checkpoint inhibitor treatment.

Description

[Technical field]

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 135,914, filed January 11, 2021, which is incorporated by reference in its entirety.

[0002] The disclosed invention relates to compositions and methods for treating cancer. More specifically, the disclosed invention relates to the field of treating cancer in a subject using oncolytic viruses, particularly Seneca Valley Virus (SVV) or SVV derivatives, in combination with checkpoint inhibitors. [Background technology]

[0003] Cancer is the second leading cause of death in the United States. Every year, one in four people die from cancer and more than one million new cases of cancer are diagnosed. The disease begins with the uncontrolled proliferation and growth of abnormally transformed cells. However, the definition does not end with the description of one disease, but includes the description of hundreds of different diseases. No two cancers are the same, no clones. The mutations that occur during cellular transformation can be similar, but are never identical. This challenge adds complexity and heterogeneity to the symptoms presented to subjects. Current cancer treatments, including chemotherapy and radiation therapy, work best when combined with immunomodulatory agents to create and strengthen an anti-tumor microenvironment. Many malignant tumors can be resistant to treatment with these traditional methods.

[0004] Oncolytic viruses have shown great potential as anticancer agents. The picornavirus Seneca Valley Virus (SVV) is a single-stranded (+)RNA virus that has been investigated as an oncolytic therapy. SVV has been shown to be able to target and promote regression of many intractable malignancies, including small cell and non-small cell lung cancer, as well as pediatric solid tumors.

[0005] A frequent problem associated with cancer treatment is that cancers are refractory to certain cancer therapies. The term "refractory" generally means stubborn or intractable, and in medicine it is specifically applied to diseases that do not respond to treatment. Refractory cancers refer to cancers that are resistant at the start of treatment or that become resistant during treatment. For example, checkpoint inhibitors (a type of anticancer drug) are known to be refractory to neuroendocrine and small cell lung cancer (SCLC) tumors.

[0006] Therefore, what is needed are improved therapeutic approaches using oncolytic viruses, particularly SVV, to treat cancers that have become refractory or are refractory to cancer treatment with monotherapy, such as checkpoint inhibitors. Summary of the Invention

[0007] Provided herein are improved methods, compositions, kits, and pharmaceutical compositions for treating cancers refractory to checkpoint inhibitor monotherapy using Seneca Valley Virus (SVV) or SVV derivatives in combination with a checkpoint inhibitor. In particular, the cancers include triple-negative breast cancer, small cell lung cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer. Also, the cancers include neuroblastoma, melanoma, neuroendocrine carcinoma, or small cell lung cancer (SCLC) tumors. In certain embodiments, the SVV derivative encodes a checkpoint inhibitor. In other embodiments, the SVV derivative encodes the checkpoint inhibitor.

[0008] One embodiment of the present invention is a method of treating cancer in a subject in need thereof comprising administering to the subject an effective amount of Seneca Valley Virus (SVV) or an SVV derivative, wherein the subject is also administered an effective amount of at least one checkpoint inhibitor, wherein the cancer is refractory to monotherapy with a checkpoint inhibitor.

[0009] Another embodiment of the invention is a method for improving the success rate of oncolytic cancer virus therapy, comprising administering to a subject having cancer an effective amount of Seneca Valley Virus (SVV) or an SVV derivative, wherein the subject is also administered an effective amount of at least one checkpoint inhibitor, wherein the cancer treatment is improved when compared to a subject who only received the effective amount of the Seneca Valley Virus or SVV derivative, and wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0010] In the methods, the checkpoint inhibitor is administered prior to, concurrently with, or following administration of the Seneca Valley Virus (SVV) or SVV derivative.

[0011] Also provided herein is a pharmaceutical composition for treating cancer in a subject, the pharmaceutical composition comprising a checkpoint inhibitor, SVV or an SVV derivative, and a pharma- ceutical acceptable carrier, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0012] Further provided herein is a kit for treating cancer in a subject, comprising Seneca Valley Virus (SVV) or an SVV derivative in combination with at least one checkpoint inhibitor, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0013] Additionally, provided herein is a Seneca Valley Virus (SVV) or SVV derivative in combination with at least one checkpoint inhibitor for use in the manufacture of a medicament for the treatment of cancer, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0014] In certain embodiments, the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor. In other embodiments, the checkpoint inhibitor is an anti-PD1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. In certain embodiments, the checkpoint inhibitor inhibits one or more of the following checkpoint proteins on cancer cells: PD-1, PD-L1, CTLA-4, B7-1, B7-2. The checkpoint inhibitor may be an antibody (e.g., a monoclonal antibody) or a nanobody. In certain embodiments, the SVV derivative encodes a checkpoint inhibitor. In other embodiments, the SVV derivative encodes the checkpoint inhibitor.

[0015] Other features and advantages of the present invention will become apparent from the following detailed description and examples. [Brief description of the drawings]

[0016] For the purpose of illustrating the invention, there is shown in the drawings specific embodiments of the invention. However, the invention is not limited to the precise arrangements and equipment of the embodiments depicted in the drawings.

[0017] [Figure 1] 1 shows the treatment schedule for the Pan02 tumor model. [Figure 2A] FIG. 1 shows the effect of treatment with SVV, checkpoint inhibitors, and SVV and checkpoint inhibitors on primary tumors based on tumor volume (mm3) as a function of days post-inoculation. [Figure 2B]FIG. 1 shows the effect of treatment with SVV, checkpoint inhibitors, and SVV and checkpoint inhibitors on contralateral tumors based on tumor volume (mm3) as a function of days post-inoculation. [Figure 3A] 1 shows Kaplan-Meier plots depicting cell viability as a function of time following treatment with SVV, checkpoint inhibitors, or a combination thereof. [Figure 3B] 1 shows Kaplan-Meier plots depicting cell viability as a function of time following treatment with SVV, checkpoint inhibitors, or a combination thereof. [Figure 3C] 1 shows Kaplan-Meier plots depicting cell viability as a function of time following treatment with SVV, checkpoint inhibitors, or a combination thereof. [Figure 4] Tumor volume after treatment with SVV, checkpoint inhibitors, or their combination based on tumor volume (mm3) as a function of days post-inoculation (study days). SVV-001 treatment was stopped on day 12. Anti-PD1 / anti-CTLA4 treatment was stopped on day 23. [Figure 5A] Tumor volume after treatment with SVV, checkpoint inhibitors, or their combination based on tumor volume (mm3) as a function of days post-inoculation (study days). [Figure 5B] Tumor volume after treatment with SVV, checkpoint inhibitors, or their combination based on tumor volume (mm3) as a function of days post-inoculation (study days). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0018] The general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention, as defined in the appended claims. Other aspects of the invention will be apparent to those skilled in the art from consideration of the detailed description of the invention provided herein.

[0019] The present invention relates to compositions and methods of using Seneca Valley Virus (SVV) or derivatives thereof to treat cancer in a subject. SVV or SVV derivatives are useful for a variety of applications, such as treating, reducing, or inhibiting the growth of cancer cells, and increasing the survival rate of subjects suffering from cancer. In particular, the present invention relates to compositions and methods of using SVV or SVV derivatives in combination with checkpoint inhibitors in patients with cancers that are refractory to checkpoint inhibitor monotherapy.

[0020] The present invention is based on the surprising discovery that the use of SVV or SVV derivatives in combination with checkpoint inhibitors to treat cancers refractory to treatment with checkpoint inhibitors improves the success rate of cancer treatment. The present invention is also based on the surprising discovery that the treatment of cancer with SVV and checkpoint inhibitors alone does not result in a high cure rate or a good abscopal effect (systemic anti-tumor immune response) in cancer model animals (mice), but the combination of the two shows both a high cure rate and a systemic anti-tumor effect.

[0021] definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in carrying out the testing of the present invention, the preferred materials and methods are described herein. In describing and claiming the present invention, the following terms are used.

[0022] It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.

[0023] As used herein, the articles "a" and "an" are used to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0024] As used herein, the term "about," when referring to a measurable value, such as an amount, duration, and the like, may be intended to encompass variations of ±20% or ±10%, more preferably ±5%, even more preferably ±1%, and even more preferably ±0.1% from the specified value, such variations being deemed appropriate for carrying out the disclosed methods.

[0025] The terms "biological" or "biological sample" refer to a sample obtained from an organism or components of an organism (e.g., cells, etc.). A biological sample may be obtained from tumor cells or tumor tissue. The sample may be any biological tissue or fluid. Often, the sample is a "clinical sample," which is a sample derived from a patient. Such samples include, but are not limited to, bone marrow, cardiac tissue, sputum, blood, lymph, blood cells (e.g., white blood cells, etc.), tissue or fine needle biopsy samples, urine, peritoneal fluid, and pleural fluid, or cells thereof. Biological samples also include tissue sections, such as frozen sections taken for histological purposes.

[0026] As used herein, the terms "comprising," "including," "containing," and "characterized by" are interchangeable, inclusive, and open-ended and do not exclude additional, unrecited elements or method steps. In particular, any recitation herein of the term "comprising," in a description of components of a composition or a description of elements of a device, is understood to encompass compositions and methods that consist essentially of and consist of the recited components or elements.

[0027] As used herein, the term "consisting of" excludes any element, step, or ingredient not specified in the claim element.

[0028] As used herein, the term "Seneca Valley Virus" or "SVV" encompasses wild-type SVV or SVV derivatives. Exemplary suitable SSV strains include SVV-001, NTX-010, and SVV strains having ATCC Patent Deposit No. PTA-5343. As used herein, the term "derivative" indicates that a derivative of a virus has a difference in nucleic acid or amino acid sequence relative to the nucleic acid or amino acid sequence of a template virus. For example, an SVV derivative can refer to an SVV that has a different nucleic acid or amino acid sequence relative to the wild-type SVV nucleic acid or amino acid sequence of ATCC Patent Deposit No. PTA-5343. In some embodiments, an SVV derivative encompasses an SVV mutant, SVV variant, or modified SVV (e.g., genetically engineered SVV). In one embodiment, an SVV derivative is an SVV virus that has been modified to express a therapeutic protein. Examples of such SVV derivatives can be found in U.S. Provisional Patent Application No. 63 / 138,999, filed January 19, 2021, the disclosure of which is incorporated herein insofar as it relates to armed SVV constructs. Exemplary suitable SVV derivatives are the following SVV armed constructs: (1) SVV-001 / Enhanced IL-2 (SVV-024), (2) SVV-001 / Anti-PD-L1 (SVV-012), (3) SVV-001 / CXCL9 (SVV-037), (4) SVV-001 / TGF beta decoy (SVV-044), (5) SVV-001 / Nitroreductase (SVV-058), (6) SVV-001 / IL2-IL15 fusion protein (SVV-069), and (6) SVV-001 / Ovalbumin epitope (SVV-077). These armed SVV constructs were designed by Seneca Therapeutics, Inc. In another embodiment, the SVV derivative is ONCR-788. In some embodiments, the modified SVV derivatives are modified so that they are capable of recognizing different cellular receptors or evading the immune system while still being able to enter, replicate and kill cells of interest (i.e., cancer cells).In other embodiments, the SVV is modified to express a chemical compound useful for treating cancer. In general, the SVV or SVV derivative can be derived from an existing virus stock that has been passaged to produce more virus. The SVV or SVV derivative can also be derived from a plasmid. The SVV derivative can also code for a checkpoint inhibitor, such as a nanobody.

[0029] As used herein, "higher" refers to an expression level that is at least 10% higher than the control standard, for example, 20%, 30%, 40%, or 50%, 60%, 70%, 80%, 90% or more higher, and / or 1.1-fold, 1.2-fold, 1.4-fold, 1.6-fold, 1.8-fold, 2.0-fold or more higher, and any and all whole or partial increments therebetween. A higher expression level than the reference value disclosed herein refers to an expression level (mRNA or protein) that is higher than the normal or control level from expression (mRNA or protein) measured in a healthy subject or as defined or used in the art.

[0030] As used herein, "lower" refers to an expression level that is at least 10% or more lower than a control standard, for example, 20%, 30%, 40%, or 50%, 60%, 70%, 80%, 90% or more lower, and / or 1.1-fold, 1.2-fold, 1.4-fold, 1.6-fold, 1.8-fold, 2.0-fold or more lower, and any and all total or partial decrements therebetween. An expression level lower than a reference value disclosed herein refers to an expression level (mRNA or protein) that is lower than a normal or control level from expression (mRNA or protein) measured in a healthy subject or as defined or used in the art.

[0031] As used herein, the terms "control" or "reference" can be used interchangeably and refer to a value used as a standard for comparison.

[0032] As used herein, "combination therapy" means that a first agent is administered in combination with another agent. "In combination with" or "In conjunction with" refers to adding one therapeutic modality in addition to another therapeutic modality. Thus, "in combination with" refers to administering one therapeutic modality before, during, or after delivery of another therapeutic modality to an individual. Such a combination is considered to be part of a single treatment regimen or regime. For purposes of this specification, combination therapy can include treatment regimes that each include administration of an oncolytic virus and another anticancer agent to treat the same hyperproliferative disease or condition, such as the same tumor or cancer. In a preferred embodiment, combination therapy includes administration of SVV or an SVV derivative in combination with one or more checkpoint inhibitors in a patient.

[0033] As used herein, the terms "peptide", "polypeptide" and "protein" are used interchangeably and refer to a compound composed of amino acid residues covalently linked by peptide bonds. A protein or peptide must contain at least two amino acids, with no limit on the maximum number of amino acids that may make up a protein or peptide sequence. A polypeptide includes any peptide or protein that contains two or more amino acids linked together by peptide bonds. As used herein, the term refers to both short chains, commonly referred to in the art as peptides, oligopeptides and oligomers, and longer chains, commonly referred to in the art as proteins, of which there are many varieties. A "polypeptide" includes, for example, biologically active fragments, substantially homologous polypeptides, oligopeptides, homodimers, heterodimers, variants of polypeptides, modified polypeptides, derivatives, analogs, fusion proteins, and the like. A polypeptide includes natural peptides, recombinant peptides, synthetic peptides, or combinations thereof.

[0034] The term "RNA" as used herein is defined as ribonucleic acid.

[0035] The term "treatment" as used within the context of the present invention is meant to include not only therapeutic treatment of a disease or disorder, but also preventive or suppressive treatment. As used herein, "treatment" and related terms such as "treat" and "treating" refer to reducing the progression, severity, and / or duration of a disease condition or at least one symptom thereof. Thus, the term "treatment" refers to any regimen that can benefit a patient. Treatment may be for an existing condition or may be preventative (prophylactic treatment). Treatment may include a curative, palliative, or preventive effect. "Therapeutic" and "prophylactic" treatment herein are to be considered in their broadest context. The term "therapeutic" does not necessarily mean that a patient is treated until complete recovery. Similarly, the term "prophylactic" does not necessarily mean that a patient will not eventually develop a disease condition. Thus, for example, the term treatment includes the administration of an agent prior to or after the onset of a disease or disorder, thereby preventing or eliminating all signs of a disease or disorder. As another example, the administration of an agent after clinical symptoms of a disease are present to address symptoms of a disease is included in the "treatment" of a disease.

[0036] As used herein, the term "nucleic acid" refers to polynucleotides such as deoxyribonucleic acid (DNA), and, where appropriate, ribonucleic acid (RNA). The term should also be understood to include, as equivalents, analogs of either RNA or DNA made from nucleotide analogs, and, as applied to the described embodiments, single-stranded (sense or antisense) and double-stranded polynucleotides. ESTs, chromosomes, cDNA, mRNA, and rRNA are representative examples of molecules that may be referred to as nucleic acids.

[0037] As used herein, the term "pharmaceutical composition" refers to a mixture of at least one compound useful within the present invention with other chemical components, such as carriers, stabilizers, diluents, adjuvants, dispersing agents, suspending agents, thickening agents, and / or excipients. A pharmaceutical composition facilitates administration of a compound to an organism. In the art, multiple techniques of administering a compound include, but are not limited to, intratumoral, intravenous, intrapleural, oral, aerosol, parenteral, ophthalmic, pulmonary, and topical administration.

[0038] The phrase "pharmaceutically acceptable carrier" includes pharma- ceutically acceptable salts, pharma- ceutically acceptable materials, compositions, or carriers, such as liquid or solid fillers, diluents, excipients, solvents, or encapsulating materials that are involved in carrying or transporting a compound of the invention in or to a subject so that it can perform its intended function. Typically, such compounds are carried or transported from one organ or part of the body to another. Each salt or carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not harmful to the patient. Some examples of materials which may function as pharma- ceutically acceptable carriers include sugars such as lactose, glucose and sucrose, starches such as corn starch and potato starch, cellulose and its derivatives such as sodium carboxymethylcellulose, ethylcellulose and cellulose acetate, powdered tragacanth, malt, gelatin, talc, excipients such as cocoa butter and suppository wax, oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil, glycols such as propylene glycol, polyols such as glycerin, sorbitol, mannitol, sorb ... Examples of suitable non-toxic compatible materials include nitrile and polyethylene glycol, esters such as ethyl oleate and ethyl laurate, agar, buffers such as magnesium hydroxide and aluminum hydroxide, alginic acid, pyrogen-free water, isotonic saline, Ringer's solution, ethyl alcohol, phosphate buffer, diluents, granules, lubricants, binders, disintegrants, wetting agents, emulsifiers, colorants, release agents, coating agents, sweeteners, flavorings, preservatives, antioxidants, plasticizers, gelling agents, thickeners, hardeners, solidifying agents, suspending agents, surfactants, humectants, carriers, stabilizers, and other non-toxic compatible materials used in pharmaceutical formulations, or any combination thereof. As used herein, "pharmaceutically acceptable carrier" also includes any and all coating agents, antibacterial and antifungal agents, and absorption retardants that are compatible with the activity of the compound and physiologically acceptable to the patient. Supplementary active ingredients can also be included in the composition.

[0039] As used herein, the term "effective amount" or "therapeutically effective amount" refers to the amount of viral particles or infectious units produced from a vector of the invention that is necessary to prevent a particular disease state or that reduces and / or ameliorates the severity of the disease state or at least one symptom thereof, or a condition associated therewith.

[0040] As used herein, the phrase "cancer refractory to monotherapy" with checkpoint inhibitors refers to any cancer that is resistant to checkpoint inhibitor or SVV monotherapy at the start of treatment or acquires resistance to checkpoint inhibitor or SVV monotherapy during treatment. This phrase includes cancers that have been treated with checkpoint inhibitors but have not responded (i.e., are resistant to cancer treatment). This phrase also includes cancers that have been treated with checkpoint inhibitors and initially responded to treatment, but then the tumors re-grow (relapsed / resistant). For purposes of this definition, the term checkpoint inhibitor monotherapy refers to cancers that have been treated with a checkpoint inhibitor as the only anti-cancer agent. Examples of such cancers include cold tumors, which are cancers that are not recognized by the immune system or that do not provoke a strong immune system response. Cold tumors are resistant to checkpoint inhibitors and / or checkpoint blockade.

[0041] As used herein, a "subject" or "patient" may be a human or a non-human mammal. Non-human mammals include, for example, livestock and pets, such as cows, pigs, dogs, cats, and mice. Preferably, the subject is a human.

[0042] Ranges: Throughout this disclosure, various aspects of the invention may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Thus, the description of a range should be considered to specifically disclose each and every numerical value within that range, along with all possible subranges. For example, 1-6 should be considered to specifically disclose subranges such as 1-3, 1-4, 1-5, 2-4, 2-6, 3-6, etc., and each and every numerical value within that range, such as 1, 2, 2.7, 3, 4, 5, 5.3, and 6. This applies regardless of the breadth of the range.

[0043] SVV or a SVV derivative and a checkpoint inhibitor for treating cancers refractory to checkpoint inhibitor monotherapy The present disclosure provides methods, compositions, kits, and pharmaceutical compositions for treating cancers refractory to checkpoint inhibitor monotherapy utilizing Seneca Valley Virus (SVV) or SVV derivatives in combination with a checkpoint inhibitor.

[0044] In one embodiment, the present disclosure provides methods, compositions, kits, and pharmaceutical compositions for treating cancers refractory to checkpoint inhibitor monotherapy utilizing SVV derivatives encoding checkpoint inhibitors. In certain embodiments, SVV derivatives encoding checkpoint inhibitors are used alone or in combination with other checkpoint inhibitors.

[0045] In certain embodiments of the present disclosure, the cancer is refractory to both SVV and checkpoint inhibitors.

[0046] The checkpoint inhibitor may be a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, or a combination thereof. In certain embodiments, the checkpoint inhibitor is an anti-PD1 antibody, an anti-PD-L1 antibody, or an anti-CTLA-4 antibody. In certain embodiments, the checkpoint inhibitor inhibits one or more of the following checkpoint proteins in the cancer cells: PD-1, PD-L1, CTLA-4, B7-1, B7-2. In other embodiments, the checkpoint inhibitor inhibits one or more of the following checkpoint proteins: LAG-3, TIM-3, TIGIT, VISTA, B7-H3, BTLA, and Siglec-15. See Qin, S. et al. Mol Cancer 18, 155 (2019); Gaynor et al. Semin Cancer Biol. 2020 Jul 2; S1044-579X (20) 30152-8. The checkpoint inhibitor may be an antibody, such as, for example, a monoclonal antibody.

[0047] In certain embodiments, the SVV derivative encoding a checkpoint inhibitor encodes more than one checkpoint inhibitor. In certain alternative embodiments, the SVV derivative encoding a checkpoint inhibitor encodes a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, or a combination thereof. In certain embodiments, the SVV derivative encoding a checkpoint inhibitor comprises a nucleic acid encoding an anti-CTLA4 antibody, a nucleic acid encoding an anti-PDL1 antibody, or both. In certain embodiments, the SVV derivative encoding a checkpoint inhibitor comprises a nucleic acid encoding an anti-CTLA4 nanobody, a nucleic acid encoding an anti-PDL1 nanobody, or both. In certain embodiments, the SVV derivative encoding a checkpoint inhibitor is an SVV derivative disclosed in U.S. Provisional Patent Application No. 63 / 138,999, filed January 19, 2021.

[0048] Additional exemplary suitable checkpoint inhibitors include, but are not limited to, ipilimumab (Yervoy®), pembrolizumab (Keytruda®), nivolumab (Opdivo®), and atezolizumab (Tecentriq®). In one embodiment, the checkpoint inhibitor is an anti-PD-1 antibody or nanobody. In other embodiments, the checkpoint inhibitor is an anti-CTLA4 antibody or nanobody.

[0049] The cancer treated with the combination of SVV or an SVV derivative and a checkpoint inhibitor is refractory to monotherapy with the checkpoint inhibitor. If more than one checkpoint inhibitor is used, the cancer is refractory to monotherapy with at least one of the checkpoint inhibitors.

[0050] The cancer treatment provided herein includes the treatment of solid tumors or the treatment of metastatic cancer. Metastatic cancer is a form of cancer in which transformed or malignant cells migrate and spread the cancer from one site to another. Such cancers include, but are not limited to, cancers of the skin, breast, brain, cervix, testis, and the like. More specifically, the cancers include, but are not limited to, organs or systems such as the heart, lung, gastrointestinal, urinary system, liver, bone, nervous system, gynecological system, blood, skin, and adrenal gland. More specifically, the methods herein may be used to treat glioma (schwannoma, glioblastoma, astrocytoma), neuroblastoma, pheochromocytoma, paraganglioma, meningioma, adrenal cortical carcinoma, kidney cancer, various vascular cancers, osteoblastic bone cancer, prostate cancer, ovarian cancer, uterine leiomyoma, salivary gland cancer, choroid plexus cancer, breast cancer, pancreatic cancer, colon cancer, and megakaryoblastic leukemia, and the like. Skin cancers include malignant melanoma, basal cell carcinoma, squamous cell carcinoma, Kaposi's sarcoma, dysplastic nevi, lipoma, hemangioma, dermatofibroma, keloids, and psoriasis.

[0051] In some embodiments, cancers treated by the presently disclosed methods include triple-negative breast cancer, small cell lung cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer.

[0052] In other embodiments, the cancer is neuroblastoma or melanoma. In yet other embodiments, the cancer is a neuroendocrine cancer or a small cell lung cancer (SCLC) tumor.

[0053] In certain embodiments, the combination of SVV or an SVV derivative and a checkpoint inhibitor provides improved cancer treatment compared to using SVV alone. In other embodiments, the combination of SVV or an SVV derivative and a checkpoint inhibitor provides improved cancer treatment compared to using SVV or an SVV derivative or a checkpoint inhibitor alone.

[0054] In certain embodiments, SVV derivatives encoding checkpoint inhibitors provide improved cancer treatment compared to using SVV alone, hi other embodiments, SVV derivatives encoding checkpoint inhibitors provide improved cancer treatment compared to using SVV or checkpoint inhibitors alone.

[0055] Combination therapy The compositions and methods for treating cancer in a subject using SVV or SVV derivatives and checkpoint inhibitors described herein may be combined with at least one additional compound useful for treating cancer. The compositions and methods for treating cancer in a subject using SVV derivatives encoding checkpoint inhibitors described herein may be combined with at least one additional compound useful for treating cancer. Additional compounds include commercially available compounds known to treat, prevent, or alleviate symptoms of cancer and / or metastasis. In certain embodiments, the additional compound is another checkpoint inhibitor.

[0056] In one embodiment, the pharmaceutical composition disclosed herein comprises an mTOR inhibitor, SVV or an SVV derivative, and a pharmaceutically acceptable carrier.The pharmaceutical composition can be used in combination with a therapeutic agent, such as an antitumor agent, including but not limited to a chemotherapeutic agent, an anti-cell proliferation agent, or any combination thereof.For example, any conventional chemotherapeutic agent of the following non-limiting exemplary classes is included in the present invention: alkylating agents, nitrosoureas, antimetabolites, antitumor antibiotics, plant alkaloids, taxanes, hormones, and miscellaneous agents.In another embodiment, the pharmaceutical composition disclosed herein is used in combination with radiation therapy.

[0057] Most alkylating agents are cell cycle non-specific. In certain embodiments, alkylating agents crosslink the guanine bases of DNA double helix strands, thereby stopping tumor growth. Non-limiting examples include busulfan, carboplatin, chlorambucil, cisplatin, cyclophosphamide, dacarbazine, ifosfamide, mechlorethamine hydrochloride, melphalan, procarbazine, thiotepa, and uracil mustard.

[0058] Antimetabolites block the incorporation of bases into DNA during the synthesis (S) phase of the cell cycle, inhibiting normal development and division. Non-limiting examples of antimetabolites include 5-fluorouracil, 6-mercaptopurine, capecitabine, cytosine arabinoside, floxuridine, fludarabine, gemcitabine, methotrexate, and thioguanine.

[0059] Antitumor antibiotics generally work by blocking enzymes necessary for cell division or by altering the membranes that encase the cells. This class includes anthracyclines, such as doxorubicin, which work by disrupting the structure of DNA and halting its function. These drugs are non-cell-phase specific. Non-limiting examples of antitumor antibiotics include aclacinomycin, actinomycin, anthramycin, azaserine, bleomycin, cactinomycin, calicheamicin, carubicin, caminomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin, esorubicin, idarubicin, marcellomycin, mitomycin, mitoxantrone, mycophenolic acid, nogalamycin, olivomycin, peplomycin, porfiromycin, puromycin, chelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, and zorubicin.

[0060] Plant alkaloids inhibit or stop mitosis or inhibit enzymes that prevent cells from making proteins necessary for cell growth. Frequently used plant alkaloids include vinblastine, vincristine, vindesine, and vinorelbine. However, the present invention should not be construed as being limited to only these plant alkaloids.

[0061] Taxanes affect cellular structures called microtubules, which are important for cellular function. In normal cell growth, microtubules form when cells begin to divide, but when cells stop dividing, they break down or are destroyed. Taxanes prevent microtubules from breaking down, so that cancer cells become clogged with microtubules and are unable to grow or divide. Non-limiting exemplary taxanes include paclitaxel and docetaxel.

[0062] Hormonal and hormone-like drugs are used for certain types of cancer, including, for example, leukemia, lymphoma, and multiple myeloma. They are often employed with other types of chemotherapy drugs to enhance their effectiveness. Sex hormones alter the action or production of female and male hormones and are used to slow the growth of breast, prostate, and endometrial cancers. Blocking the production (aromatase inhibitors) or action (tamoxifen) of these hormones is often used as an adjunct to treatment. Some other tumors are also hormone-dependent. Tamoxifen is a non-limiting example of a hormonal drug that blocks the activity of estrogen, which promotes the growth of breast cancer cells.

[0063] Miscellaneous agents include chemotherapeutic agents such as bleomycin, hydroxyurea, L-asparaginase, and procarbazine.

[0064] Other examples of chemotherapeutic agents include, but are not limited to, the following, and pharma- ceutically acceptable salts, acids, and derivatives thereof: MEK inhibitors, such as, but not limited to, refametinib, selumetinib, trametinib, or cobimetinib; nitrogen mustards, such as chlorambucil, chromafazine, chlorophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, nobembitine, phenesterine, prednimustine, trofosfamide. , uracil mustard; nitrosoureas, for example, carmustine, chlorozotocin, fotemustine, lomustine, nimustine, ranimustine; purine analogues, for example, fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogues, for example, ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enokitabine, floxuridine, 5-FU; androgens, for example, calsterone, dromostanolone propionate, epithiostanol, methymustine, Pitiostane, testolactone; antiadrenergics, such as aminoglutethimide, mitotane, trilostane; folic acid supplements, such as floric acid; aceglatone; aldophosphamide glycosides; aminolevulinic acid; amsacrine; bestravcil; bisantrene; edatrexate; defofamine; demecolcine; diaziquone; eflornithine; elliptinium acetate; etoglucide; gallium nitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone; mopidamol; nitracrine; pentostatin; Phenamet; pirarubicin; podophyllic acid; 2-ethylhydrazide; procarbazine; polysaccharide-K (PSK); razoxane; schizofuran; spirogermanium; tenuazonic acid; triaziquone; 2,2',2"-trichlorotriethylamine; urethane; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids, e.g., paclitaxel (TAXOLO, Bristol-Myers Squibb Oncology, Princeton, NJ) and docetaxel (TAXOTERE, Rhone-Poulenc Rorer, Antony, France); chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogues, such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine; navelbine; novantrone; teniposide; daunomycin; aminopterin; xeloda; ibandronate; CPT-11; topoisomerase inhibitors RFS2000; difluoromethylornithine (DMFO); retinoic acid; esparamycin; and capecitabine.

[0065] The anti-cell proliferation agent can be further defined as an apoptosis inducer or a cytotoxic agent. The apoptosis inducer can be a granzyme, a Bcl-2 family member, cytochrome C, a caspase, or a combination thereof. Exemplary granzymes include granzyme A, granzyme B, granzyme C, granzyme D, granzyme E, granzyme F, granzyme G, granzyme H, granzyme I, granzyme J, granzyme K, granzyme L, granzyme M, granzyme N, or a combination thereof. In other specific embodiments, the Bcl-2 family member is, for example, Bax, Bak, Bcl-Xs, Bad, Bid, Bik, Hrk, Bok, or a combination thereof.

[0066] In additional embodiments, the caspase is caspase-1, caspase-2, caspase-3, caspase-4, caspase-5, caspase-6, caspase-7, caspase-8, caspase-9, caspase-10, caspase-11, caspase-12, caspase-13, caspase-14, or a combination thereof. In certain embodiments, the cytotoxic agent is TNF-α, gelonin, prodigiosin, ribosome inhibitor protein (RIP), Pseudomonas exotoxin, Clostridium difficile Toxin B, Helicobacter pylori VacA, Yersinia Enterocolitica YopT, violacein, diethylenetriaminepentaacetic acid, irofulven, diphtheria toxin, mitogillin, ricin, botulinum toxin, cholera toxin, saporin 6, or a combination thereof.

[0067] Immunotherapeutic agents include, but are not limited to, interleukin 2 or other cytokines, inhibitors of programmed cell death protein 1 (PD-1) signaling such as ipilimumab, a monoclonal antibody that binds to PD-1. Immunotherapeutic agents can also interfere with cytotoxic T-lymphocyte-associated antigen A-4 (CTLA-4) signaling and can be associated with cancer vaccines and dendritic cell-based therapies.

[0068] In one embodiment, the subject suffering from cancer is also administered at least one anti-cancer therapeutic agent selected from the group consisting of: checkpoint inhibitors, PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, cytokines, growth factors, photosensitizers, toxins, siRNA molecules, signal transduction modulators, anti-cancer antibiotics, anti-cancer antibodies, angiogenesis inhibitors, chemotherapeutic compounds, anti-metastatic compounds, immunotherapeutic compounds, CAR therapy, dendritic cell-based therapy, cancer vaccines, oncolytic viruses, engineered anti-cancer viruses or virus derivatives, and any combination thereof. In one embodiment, the at least one anti-cancer therapeutic agent is administered before, simultaneously with, or after administration of the SVV or SVV derivative.

[0069] In one embodiment, the subject is also administered an IFN-I inhibitor.An IFN-I inhibitor as used herein includes any agent known in the art for inhibiting, suppressing or reducing the IFN-I pathway, partially or completely, and temporarily or permanently.In some embodiments, the inhibitory effect of the IFN-I inhibitor can be reversible.In other embodiments, the inhibition of IFN-I is reversed.

[0070] Inhibitors include siRNAs, ribozymes, antisense molecules, aptamers, peptidomimetics, small molecules, mTOR inhibitors, histone deacetylase (HDAC) inhibitors, Janus kinase (JAK) inhibitors, IFN inhibitors, IFN antibodies, IFN-α receptor 1 antibodies, IFN-α receptor 2 antibodies, and viral peptides, and any combination thereof. The viral peptide can be, but is not limited to, the NS1 protein from influenza virus or the NS2B3 protease complex from dengue virus.

[0071] The mTOR pathway and its inhibition are known to be involved in various diseases, including cancer. Rapamycin is a natural product produced by the bacterium Streptomyces hygroscopicus, which can inhibit mTOR through its association with the intracellular receptor FK-506-binding protein 12 (FKBP12). The FKBP12-rapamycin complex directly binds to the FKBP12-rapamycin-binding domain of mTOR. mTOR functions as a catalytic subunit of two distinct molecular complexes, mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). mTORC1 is composed of mTOR regulatory associated protein (Raptor) and mammalian LST8 / G protein β subunit-like protein (mLST8 / GβL). This complex functions as a nutrient / energy / redox sensor and is responsible for controlling protein synthesis. mTORC1 activity is stimulated by insulin, growth factors, serum, phosphatidic acid, amino acids (especially leucine), and oxidative stress (Hay and Sonenberg, Genes Dev. 18(16):1926-1945, 2004; Wullschleger et al., Cell 124(3):471-484). On the other hand, mTORC1 is known to be inhibited by low nutrient levels, growth factor deprivation, reductive stress, caffeine, rapamycin, farnesylthiosalicylic acid, and curcumin (Beevers et al., Int. J. Cancer 119(4):757-764, 2006; McMahon et al., Mol. Endocrinol. 19(1):175-183). Components of mTORC2 are rapamycin-insensitive companion of mTOR (Rictor), GβL, mammalian stress-activated protein kinase-interacting protein 1, and mTOR.mTORC2 has been shown to function as a regulator of the cytoskeleton through stimulation of F-actin stress fibers, paxillin, RhoA, Rac1, Cdc42, and protein kinase Cα (Sarbassov et al., Curr. Biol. 14(14):1296-302, 2004; Sarbassov et al., Science 307(5712):1098-101, 2005). Unlike mTORC1, mTORC2 is insensitive to rapamycin.

[0072] Many mTOR inhibitors are known in the art and have strong immunosuppressive and antitumor activity. mTOR inhibitors, such as rapamycin, rapamycin analogs, or rapamycin derivatives, are known to exhibit immunosuppressive and antiproliferative properties. Other mTOR inhibitors include everolimus, tacrolimus, zotarolimus (ABT-578), pimecrolimus, biolimus, FK-506, PP242 (2-(4-amino-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-1H-indol-5-ol), KuPP242 (2-(4-amino-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-1H-indol-5-ol), Ku-00637 94 (rel-5-[2-[(2R,6S)-2,6-dimethyl-4-morpholinyl]-4-(4-morpholinyl)pyrido[2,3-d]pyrimidin-7-yl]-2-methoxybenzenemethanol), PI-103 (3-(4-(4-morpholinyl)pyrido[3',2":4,5]furo[3,2-d]pyrimidin-2-yl)phenol), PKI-179 (N-[4-[4-morpholinyl)-6-(3-oxa-8-azabicyclo[3.2.1]octo-8 -yl)-1,3,5-triazin-2-yl]phenyl]-N'-4 pyridinylurea hydrochloride), AZD8055 (5-[2,4-bis[(3S)-3-methyl-4-morpholinyl]pyrido[2,3d]pyrimidin-7-yl]-2-methoxybenzenemethanol), WYE-132 / WYE-125132 (1-{4-[1-(1,4-dioxa-spiro[4.5]dec-8-yl)-4-(8-oxa-3-aza-bicyclo[3.2.1]octo-3-yl)-1,3,5-triazin-2-yl]phenyl)-N'-4 pyridinylurea hydrochloride), (4-{6-[4-(3-cyclopropyl-ureido)-phenyl]-4-morpholin-4-yl-pyrazolo[3,4-d]pyrimidin-1-yl}-piperidine-1-carboxylic acid methyl ester), WYE-28 (4-(6-{4-[3(4-hydroxymethyl-phenyl)-ureido]-phenyl}-4-morpholin-4-yl-pyrazolo[3,4-d]pyrimidin-1-yl)piperidine-1-carboxylic acid methyl ester), WYE-354 (4-[6-[4[(methoxycarbonyl)amino]phenyl]-4-(4-morpholinyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]-1 piperidine carboxylic acid methyl ester), C20-methallylrapamycin, and C16-(S)-butylsulfonylrapamycin, C16-(S)-3-methylindole Rapamycin (C16-iRap), C16-(S)-7-methylindolerapamycin (AP21967 / C16-AiRap), CCI-779 (temsirolimus), RAD001 (40-O-(2-hydroxyethyl)-rapamycin), AP-23575, AP-23675, AP-23573, 20-thiarapamycin, 15-deoxo-19-sulfoxylrapamycin, WYE-592, ILS-920, (3S ,6R,7E,9R,10R,12R,14S,15E,17E,19E,21S,23S,26R,27R,34aS)-9,10,12,13,14,2-1,22,23,24,25,26,27,32,33,34,34a-Hexadecahydro-9,27-dihydroxy-3-[(1R)-2-[(-1S,3R,4R)-3-methoxy-4-tetrazol-1-yl]cyclohexyl-1-methylethyl]-10,2 1-Dimethoxy-6,8,12,14,20,26-hexamethyl-23,27-epoxy-3H-pyrido[2,1-c][1,4]oxaazacyclohentriacontin-1,5,11,28,29(4H,6H,31H)-pentone) 23,27-epoxy-3H-pyrido[2,1-c][1,4]oxaazacyclohentriacontin-1,5,11,28,29(4H,6H,31H)-pentone (U.S. Patent No. 6,015,No. 815), A-94507, deforolimus, AP-23675, AP-23841, zotarolimus, CC1779 / temsirolimus, RAD-001 / iberolimus, 7-epi-rapamycin, 7-thiomethyl-rapamycin, 7-epi-trimethoxy-rapamycin, 2-desmethyl-rapamycin, and 42-O-(2-hydroxy)ethyl-rapamycin, AP-23841, 7-epi-rapamycin, 7-thiomethyl-rapamycin, 7-epi-trimethoxyphenyl-rapamycin, 7-epi-thiomethyl-rapamycin, 7-demethoxy-rapamycin, 32-demethoxy-rapamycin, 2-desmethyl-rapamycin, and 42-O-(2-hydroxy)ethyl-rapamycin. Isin, 42-O-(2-hydroxy)ethylrapamycin, ridaforolimus, ABI-009, MK8669, TOP216, TAFA93, TORISEL™ (prodrug), CERTICAN™, Ku-0063794, PP30, Torin 1, Torin 2, ECO371, AP23102, AP23573, AP23464, AP23841; 40-(2-hydroxyethyl)rapamycin, 40-[3-hydroxy(hydroxymethyl)methylpropanoate]-rapamycin (also known as CC1779), 32-deoxorapamycin, and 16-pentynyloxy-32(S)-dihydrorapamycin. Non-rapamycin analogue mTOR inhibitors include, but are not limited to, LY294002, wortmannin, quercetin, myricetin, staurosporine, and ATP-competitive inhibitors. Other examples of suitable mTOR inhibitors are described in U.S. Patent No. 6,329,386, U.S. Publication No. 2003 / 129215, and U.S. Publication No. 2002 / 123505.

[0073] In some embodiments, the disclosed mTOR inhibitors inhibit at least one of mTORC1 and mTORC2. In other embodiments, the disclosed mTOR inhibitor is Torin1 or Torin2.

[0074] A large number of HDAC inhibitors are known and used in the art. The most common HDAC inhibitors bind to the zinc-containing catalytic domain of HDAC. These HDAC inhibitors can be classified into several groups named according to their chemical structure and the chemical moiety that binds to the zinc ion. Some examples include, but are not limited to, hydroxamic acid or hydroxamate (such as trichostatin A, or vorinostat / SAHA (FDA approved)), aminobenzamide entinostat (MS-275), takecynarin (C1994), mocetinostat (MGCD0103), cyclic peptides (apicidin, romidepsin (FDA approved)), cyclic tetrapeptides or epoxyketones (such as trapoxin B), depsipeptides, benzamides, electrophilic ketones, carboxylic fatty acid compounds (such as butyric acid, phenylbutyrate, valproic acid, valproic acid, etc.). Other HDAC inhibitors include, but are not limited to, belinostat (PXD101), LAQ824, and panobinostat (LBH589). Examples of HDCA inhibitors in clinical trials include panobinostat (LBH-589), belinostat (PXD101), entinostat (MS275), mocetinostat (MGCD01030), divinostat (ITF2357), practinostat (SB939), chidamide (CS055 / HBI-8000), xinostat (JNJ-26481585), abexinostat (PCI-24781), CHR-3996, and AR-Z2. In one embodiment, the HDAC inhibitor is trichostatin A.

[0075] JAK inhibitors (also known as JAK / SAT inhibitors) inhibit the activity of one or more of the Janus kinase family of enzymes (e.g., JAK1, JAK2, JAK3 and / or TYK2, etc.), thereby disrupting the JAK-STAT signaling pathway. Various JAK inhibitors are known in the art and are used to treat inflammatory diseases or cancer. Non-limiting examples of JAK inhibitors include FDA-approved compounds such as ruxolitinib (Jakafi / Jakavi), tofacitinib (Jakvinus, formerly known as tasocitinib and CP-690550), oclacitinib (Apoquel), baricitinib (Olumient, LY3009104), and decernotinib (VX-509). Other JAK inhibitors are in clinical trials and / or are used as experimental drugs. Examples include filgotinib (G-146034, GLPG-0634), celduratinib (PRT062070), gandotinib (LY-2784544), lestaurtinib (CEP-701), momelotinib (GS-0387, CYT-387), pacurtinib (SB1518), PF-04965842, upadacitinib (ABT494), peficitinib (ASPO15K, JNJ-54781532), fedratinib (SAR302503), cucurbitacin I, CHZ868, ABT-494, dimethyl fumarate (DMF, Tecfidera), GLPG0634, and CEP-33779. In one embodiment, the JAK / STAT inhibitor is staurosporine (STS; antibiotic AM-2282), an inhibitor of protein kinase C (PKC).

[0076] In one embodiment, the subject is further administered at least one IFN-I inhibitor selected from the group consisting of: HDAC inhibitors, JAK / STAT inhibitors, IFN inhibitors, IFN antibodies, IFN-α receptor 1 antibodies, IFN-α receptor 2 antibodies, and viral peptides, and any combination thereof. In another embodiment, the at least one IFN-I inhibitor is administered before, during, or after administration of SVV or an SVV derivative. In some embodiments, after SVV or an SVV derivative replicates in tumor cells, the at least one IFN-I inhibitor is removed before adding an anti-cancer drug (such as, for example, a checkpoint inhibitor).

[0077] In one embodiment, the anti-cancer agent is administered before, simultaneously with, or after the administration of at least one IFN-I inhibitor. In one embodiment, the anti-cancer agent is administered subsequent to the administration of at least one IFN-I inhibitor. In another embodiment, the anti-cancer agent is administered subsequent to the administration of at least one IFN-I inhibitor and SVV or an SVV derivative.

[0078] In one embodiment, treatment with SVV or SVV derivatives and checkpoint inhibitors is performed prior to administration of IFN-I inhibitors. In one embodiment, once replication of SVV or SVV derivatives and death of cancer cells are confirmed, administration of IFN-I inhibitors is terminated. For example, cancer cells can be administered IFN-I inhibitors (e.g., (5-(tetradecyloxy)-2-furoic acid), acetyl-CoA carboxylase inhibitor: TOFA) 24 hours prior to SVV or SVV derivative treatment, and then both treatments can be continued for several weeks until robust replication of SVV or SVV derivatives is observed and markers of cell death are detected. Then, treatment with IFN-I inhibitors can be terminated and treatment with anti-cancer drugs can be initiated. As SVV or SVV derivatives replicate, various nucleic acids and cell debris are produced, which triggers activation of the influx of immune cells (T cells, NK cells, APCs, etc.) to proceed with suppression, reduction, and / or elimination / killing of cancer cells. This process of immune response is further enhanced by the end of IFN-I inhibition.

[0079] Pharmaceutical Compositions In a particular embodiment, the present invention is directed to a pharmaceutical composition comprising SVV or an SVV derivative and a checkpoint inhibitor. In another embodiment, the present invention is directed to a pharmaceutical composition comprising SVV or an SVV derivative and another pharmaceutical composition comprising a checkpoint inhibitor. In yet another embodiment, the present invention is directed to a pharmaceutical composition comprising an SVV derivative encoding a checkpoint inhibitor, optionally comprising a further checkpoint inhibitor.

[0080] Provided herein is a pharmaceutical composition for treating cancer in a subject in need thereof.The pharmaceutical composition comprises a checkpoint inhibitor, SVV or an SVV derivative, and a pharma- ceutical acceptable carrier, and the cancer is refractory to monotherapy with checkpoint inhibitors.

[0081] Also provided herein is a pharmaceutical composition for treating cancer in a subject in need thereof.The pharmaceutical composition comprises a checkpoint inhibitor, SVV or an SVV derivative, and a pharma- ceutical acceptable carrier, and the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0082] Such pharmaceutical compositions are in a form suitable for administration to a subject, or the pharmaceutical compositions may further comprise one or more pharma- ceutically acceptable carriers, one or more additional ingredients, or some combination thereof.Various components of the pharmaceutical compositions may be present in the form of physiologically acceptable salts, such as in combination with physiologically acceptable cations or anions, as is well known in the art.

[0083] In one embodiment provided herein, pharmaceutical compositions useful for practicing the methods of the invention are administered to deliver a dose between 1 ng / kg / day and 100 mg / kg / day. In another embodiment, pharmaceutical compositions useful for practicing the invention are administered to deliver a dose between 1 ng / kg / day and 500 mg / kg / day. The relative amounts of active ingredient, pharma- ceutically acceptable carrier, and any additional components in the pharmaceutical compositions of the invention will vary depending on the identity, size, and condition of the subject being treated, as well as the route by which the composition is administered. By way of example, the compositions contain 0.1% to 100% (w / w) active ingredient.

[0084] Pharmaceutical compositions useful in the methods of the present invention can be developed for inhalation, oral, rectal, vaginal, parenteral, topical, transdermal, pulmonary, intranasal, buccal, ocular, intrathecal, intravenous or another route of administration. Other contemplated formulations include projected nanoparticles, liposomal preparations, resealed erythrocytes containing the active ingredient, and immunologically-based formulations. The route of administration is readily apparent to one skilled in the art and depends on any number of factors, including the type and severity of the disease being treated, the type and age of the animal or human patient being treated, and the like.

[0085] The formulation of pharmaceutical compositions described herein can be prepared by any method known or developed in the art of pharmacy.In general, such preparation method includes combining active ingredient with carrier or one or more other additional ingredients, and then, if necessary or desired, shaping or packaging the product into desired single or multiple dose units.In some embodiments, SVV or SVV derivative can be formulated as natural capsid, modified capsid, naked RNA, or encapsulated in protective coat.

[0086] The amount of active ingredient is generally equal to the dosage of the active ingredient administered to a subject, or a convenient fraction of such a dosage, such as, for example, 1 / 2 or 1 / 3 of such a dosage. The unit dosage form may be administered once per day, or may be one of multiple daily doses (e.g., about 1 to 4 or more times per day). In the case of multiple daily doses, the unit dosage form may be the same or different for each dose.

[0087] The description of pharmaceutical compositions provided herein is directed in principle to pharmaceutical compositions suitable for ethical administration to humans, but it will be understood by those skilled in the art that such compositions are generally suitable for administration to any type of animal. Modifications of pharmaceutical compositions suitable for administration to humans to make them suitable for administration to various animals are well understood, and an ordinarily skilled veterinary pharmacologist can design and implement such modifications, if any, with no more than routine experimentation. Subjects contemplated for administration of pharmaceutical compositions in the present invention include, but are not limited to, humans and other primates, as well as commercially relevant mammals, such as cows, pigs, horses, sheep, cats, and dogs. In one embodiment, the subject is a human or a non-human mammal, such as, but not limited to, horses, sheep, cows, pigs, dogs, cats, and mice. In one embodiment, the subject is a human.

[0088] In one embodiment, the composition is formulated with one or more pharma- ceutically acceptable excipients or carriers. In one aspect, a pharmaceutical composition for treating cancer in a subject is disclosed. The pharmaceutical composition comprises a checkpoint inhibitor, SVV or an SVV derivative, and a pharma- ceutically acceptable carrier. In one embodiment, the pharmaceutical composition comprises an SVV derivative encoding a checkpoint inhibitor and a pharma- ceutically acceptable carrier. Useful pharma- ceutically acceptable carriers include, but are not limited to, glycerol, water, saline, ethanol, and pharma- ceutically acceptable salt solutions, such as phosphates and organic acid salts. The carrier may be a solvent or dispersion medium, and includes, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. Proper fluidity is maintained, for example, by the use of a coating agent, such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, etc. In many cases, it is preferable to include isotonic agents, such as sugars, sodium chloride, polyalcohols (such as mannitol and sorbitol) in the composition. Prolonged absorption of injectable compositions can be brought about by including in the composition an agent which delays absorption, such as aluminum monostearate or gelatin.

[0089] The preparations may be used in admixture with conventional excipients, i.e., pharma- ceutically acceptable organic or inorganic carrier substances suitable for oral, parenteral, nasal, intravenous, subcutaneous, enteral, or any other suitable mode of administration known in the art. The pharmaceutical preparations may be sterilized and, if desired, may be mixed with auxiliary substances, such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure buffers, coloring agents, flavorings and / or aromatic substances, etc. Also, if desired, they may be combined with other active agents, such as other analgesics.

[0090] The composition may contain a preservative at about 0.005% to 2.0% of the total weight of the composition. Preservatives are used to prevent spoilage when exposed to environmental contaminants. Examples of preservatives useful in accordance with the present invention include, but are not limited to, those selected from the group consisting of benzyl alcohol, sorbic acid, parabens, imidurea, and combinations thereof. A particularly preferred preservative is a combination of about 0.5% to 2.0% benzyl alcohol and 0.05% to 0.5% sorbic acid.

[0091] The composition may include antioxidants and chelating agents that inhibit degradation of the compound. Preferred antioxidants for some compounds are BHT, BHA, α-tocopherol, and ascorbic acid in the preferred range of about 0.01%-0.3% by weight based on the total weight of the composition, and more preferably BHT in the range of 0.03%-0.1%. Preferably, the chelating agent is present in an amount of 0.01%-0.5% by weight based on the total weight of the composition. Particularly preferred chelating agents include edetate (e.g., edetate disodium) and citric acid in the range of about 0.01%-0.20% by weight based on the total weight of the composition, and more preferably 0.02%-0.10% by weight based on the total weight of the composition. Chelating agents are useful for chelating metal ions in the composition that may adversely affect the shelf life of the formulation. While BHT and disodium edetate are particularly preferred antioxidants and chelating agents, respectively, for some compounds, other suitable and equivalent antioxidants and chelating agents may be substituted, as known to those of skill in the art.

[0092] The pharmaceutical compositions disclosed herein can be used in combination with additional therapeutic agents, such as antitumor agents, including, but not limited to, chemotherapeutic agents, anti-cell proliferation agents, or any combination thereof.For example, any conventional chemotherapeutic agent of the following non-limiting exemplary classes is included in the present invention: alkylating agents, nitrosoureas, antimetabolites, antitumor antibiotics, plant alkyloids, taxanes, hormonal agents, and miscellaneous agents.In another embodiment, the pharmaceutical compositions disclosed herein are used in combination with radiation therapy.

[0093] Administration / Dosage In certain embodiments of the present invention, SVV or SVV derivative and checkpoint inhibitor are administered simultaneously. In other embodiments, checkpoint inhibitor is administered before SVV or SVV derivative is administered. In another embodiment, checkpoint inhibitor is administered after SVV or SVV derivative is administered. In alternative embodiments, SVV derivative encoding checkpoint inhibitor is administered without another checkpoint inhibitor. In other embodiments, SVV derivative encoding checkpoint inhibitor is administered with another checkpoint inhibitor that is administered before, after, or simultaneously with SVV derivative encoding checkpoint inhibitor.

[0094] The administration regimen may affect the effective amount. For example, the therapeutic formulation may be administered to the patient subject either before or after a cancer-related surgical procedure, or immediately after the patient is diagnosed with cancer. Furthermore, several divided doses or different doses may be administered daily or sequentially, and the dose may be continuously infused or bolus injected. Furthermore, the dosage of the therapeutic formulation may be increased or decreased in proportion to the exigencies of the therapeutic or prophylactic situation.

[0095] Generally, SVV or SVV derivatives are 7 ~1×10 11 It is administered in a dose of vp / kg. The dosage depends on various factors, such as the age, weight, and sex of the patient, and the size and severity of the tumor being treated.

[0096] SVV or SVV derivatives are usually administered in a therapeutically effective amount. A therapeutically effective amount refers to an amount of virus that results in an improvement in the patient's condition or a prolongation of survival. The toxicity and therapeutic effect of the virus can be determined by standard methods in cell cultures or experimental animals. For example, LD 50 (the dose lethal to 50% of the animals or cell population; for viruses, the dose is in vp / kg), and ED 50(therapeutically effective dose in 50% of the animals or cell population, vp / kg), or TC 10 (therapeutic concentration or dose that can inhibit 50% of tumor cells, which can be related to PFU), or EC 50 (the concentration effective in 50% of animals or a cell population, vp / cell). The dose ratio between toxic and therapeutic effects is the therapeutic index, and the LD 50 and ED 50 or EC 50 The dose of virus is preferably at least 50% of the ED that produces little or no toxicity. 50 or EC 50 The dosage varies within this range depending upon the dosage form used and the route of administration.

[0097] The SVV or SVV derivative may be present in the composition in multiple and single dose amounts, including but not limited to about 1×10 5 ~1×10 12 p.f.u., 1 × 10 6 ~1×10 10 pfu, or 1 × 10 7 ~1×10 10 pfu, e.g., at least or about at least 1 x 10 5 , 1×10 6 , 1×10 7 , 1×10 8 , 1×10 9 , 2×10 9 , 3×10 9 , 4×10 9 , 5×10 9 , 6×10 9 , 7×10 9 , 8×10 9 , 9×10 9 , 1×10 10 , 1×10 11 , or 1×10 12 Each of these inclusively includes pfu.

[0098] The volume of the composition can be any volume and can be administered in a single or multiple doses, including but not limited to about 0.01 mL to 100 mL, about 0.1 mL to 100 mL, 1 mL to 100 mL, 10 mL to 100 mL, 0.01 mL to 10 mL, 0.1 mL to 10 mL, 1 mL to 10 mL, 0.02 mL to 20 mL, 0.05 mL to 5 mL, 0.5 mL to 50 mL, or 0.5 mL to 5 mL.

[0099] The infectivity of the SVV or SVV derivative can be manifested, for example, by an increase in the titer or half-life of the oncolytic virus upon exposure to a bodily fluid such as blood or serum. The infectivity can be increased by any amount, including, but not limited to, at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 1.6-fold, 1.7-fold, 1.8-fold, 1.9-fold, 2.0-fold, 2.5-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold.

[0100] Administration of the compositions of the present invention to a patient subject, preferably a mammal, more preferably a human, can be carried out using known procedures at dosages and for periods of time effective to treat cancer in the subject. The effective amount of the therapeutic compound required to obtain a therapeutic effect can vary depending on factors such as the activity of the particular compound employed; the timing of administration; the rate of excretion of the compound; the length of treatment; other drugs, compounds or substances used in combination with the compound; the state of the disease or disorder, age, sex, weight, condition, general health and past medical history of the patient being treated, and similar factors well known in the medical arts. The dosage regimen may be adjusted to provide an optimal therapeutic response. For example, the compound may be administered in several doses per day, or the dosage may be reduced in proportion to the exigencies of the therapeutic situation. A non-limiting example of an effective dosage range for the therapeutic compounds of the present invention is about 0.01 to 50 mg / kg body weight / day.

[0101] The SVV or SVV derivative and checkpoint inhibitor may be administered to a subject as frequently as several times a day, or less frequently, for example, once a day, once a week, once every two weeks, once a month, or even less frequently, for example, once every few months or once a year or less. It is understood that the amount of compound administered daily may be administered, in non-limiting examples, every day, every other day, every third day, every fourth day, or every fifth day. For example, alternate day administration may begin with a dose of 5 mg per day on Monday, a first subsequent dose of 5 mg per day on Wednesday, a second subsequent dose of 5 mg per day on Friday, etc. The frequency of administration will be readily apparent to one of skill in the art and will depend on any number of factors, including, but not limited to, the type and severity of the disease being treated, and the type and age of the animal. The actual dosage of the active ingredient in the pharmaceutical composition of the present invention can be varied for a specific patient, composition, and mode of administration, to obtain the amount of active ingredient that is not toxic to the patient and is effective to achieve the desired therapeutic response.A medical doctor, such as a physician or veterinarian, who has ordinary skill in the art, can easily determine and prescribe the effective amount of the pharmaceutical composition required.For example, a physician or veterinarian can start the dosage of the compound of the present invention used in the pharmaceutical composition at a level lower than the level required to achieve the desired therapeutic effect, and gradually increase the dosage until the desired effect is achieved.

[0102] In certain embodiments, it is particularly advantageous to formulate the compound in dosage unit form for ease of administration and uniformity of dosage.Dosage unit form as used herein refers to a physically separate unit suitable as a unitary dosage for the patient to be treated, each unit containing a predetermined amount of therapeutic compound calculated to produce a desired therapeutic effect in association with required pharmaceutical vehicle.The dosage unit form of the present invention is determined by and directly depends on (a) the unique characteristics of therapeutic compound and the specific therapeutic effect to be achieved, and (b) the inherent limitations of the technology of compounding / formulating such therapeutic compound to treat cancer in patients.

[0103] Route of administration Those skilled in the art will recognize that, although multiple routes of administration can be used, certain routes can provide more immediate and effective responses than other routes. In certain embodiments of the present invention, SVV or SVV derivatives and checkpoint inhibitors are administered via the same administration route. In other embodiments, SVV or SVV derivatives and checkpoint inhibitors are administered via different administration routes.

[0104] In one embodiment, the SVV or SVV derivative is administered intratumorally. In another embodiment, the checkpoint inhibitor is administered systemically. In an alternative embodiment, the SVV or SVV derivative is administered intratumorally and the checkpoint inhibitor is administered systemically.

[0105] Routes of administration of the disclosed compositions (including SVV or SVV derivatives and / or checkpoint inhibitors) include inhalation, oral, nasal, rectal, parenteral, sublingual, transdermal, transmucosal (e.g., sublingual, lingual, (trans)buccal, (trans)urethral, ​​vaginal (e.g., vaginal and perivaginal), (trans)nasal, and (trans)rectal), intravesical, intrapulmonary, intraduodenal, intragastric, intrathecal, subcutaneous, intramuscular, intradermal, intraarterial, intravenous, intrabronchial, inhalation, and topical administration. Suitable compositions and dosage forms include, for example, tablets, capsules, caplets, pills, gel caps, troches, dispersions, suspensions, solutions, syrups, granules, beads, transdermal patches, gels, powders, pellets, magmas, lozenges, creams, pastes, patches, lotions, discs, suppositories, liquid sprays for nasal or oral administration, dry powder or aerosol formulations for inhalation, compositions and formulations for intravenous administration, and the like. It should be understood that the formulations and compositions contemplated as useful in the present invention are not limited to the specific formulations and compositions described herein. In one embodiment, SVV or SVV derivative therapy and / or checkpoint inhibitor therapy comprises a route of administration selected from the group consisting of inhalation, oral, rectal, vaginal, parenteral, topical, transdermal, pulmonary, intranasal, buccal, intraocular, hepatic arterial, intrapleural, intrathecal, intratumoral, intravenous, and any combination thereof.

[0106] kit The invention also includes a kit comprising SVV or an SVV derivative and a checkpoint inhibitor, which is used to treat cancers refractory to checkpoint inhibitor monotherapy.

[0107] In further embodiments, a kit for treating or ameliorating cancer is provided as described elsewhere herein, the kit comprising a) SVV or SVV derivative or a composition comprising SVV or SVV derivative, b) a checkpoint inhibitor, and optionally c) an additional agent or therapy as described herein. The kit may further comprise instructions or a label for using the kit to treat or ameliorate cancer. The kit may also comprise an assay for confirming that the cancer is indeed refractory to the checkpoint inhibitor. In yet other embodiments, the invention extends to the assay of the kit for a given cancer (such as, by way of non-limiting example, small cell lung cancer or triple negative breast cancer) as described herein. Such a kit may, for example, comprise reagents for PCR or other nucleic acid hybridization techniques (microarrays), or reagents for immunologically-based detection techniques (e.g., ELISpot, ELISA).

[0108] Working Example The present invention will now be described with reference to the following examples, which are provided for illustrative purposes only, and the present invention should in no way be construed as being limited to these examples, but rather as embracing any and all variations that become evident as a result of the teachings provided herein.

[0109] Without further description, one of ordinary skill in the art will be able, using the preceding description and the following illustrative examples, to make and utilize the compounds of the present invention and practice the claimed methods. Thus, the following examples specifically point out preferred embodiments of the present invention, and are not to be construed as limiting in any way the remainder of the disclosure. EXAMPLES

[0110] Example 1: Combination therapy with SVV and checkpoint inhibitors in cancers refractory to checkpoint inhibitors. Seneca Valley Virus (SVV-001) is an oncolytic virus of the Picornaviridae family that, to date, has only been tested as a single-dose intravenous monotherapy in early clinical trials in patients with cancers with neuroendocrine characteristics (e.g., NETs, ​​NECs). Recent data in the field of oncolytic therapy and immunotherapy have shown that intratumoral injection of oncolytic viral agents in combination with systemic administration of checkpoint inhibitors (CPIs) enhances efficacy in preclinical models and clinical trials. Two mouse synthetic tumor models derived from neuroblastoma and melanoma that are resistant to CPI therapy were tested. When injected intratumorally in these immunocompetent animal models, SVV-001 reversed resistance to CPIs and enhanced the efficacy of checkpoint inhibitors in both tumor models.

[0111] SVV-001 has been identified as a potent oncolytic virus with extremely high tumor selectivity against human tumors. Recently, it was discovered that the receptor for SVV is TEM8, a protein highly and specifically expressed in multiple cell types within solid tumors. Interestingly, TEM8 is expressed in malignant tumor cells and cancer stem cells, as well as in “normal” cells in the tumor microenvironment, such as angiogenic endothelial cells, cancer-associated fibroblasts, and pericytes. Although SVV infects cancer cells and cancer stem cells, it is unclear whether it infects or inhibits the growth of these important cell types in the tumor microenvironment. SVV-001 has many characteristics that make it an exemplary oncolytic virus: its ability to target and enter solid tumors due to its very small size (27 nM), its specificity for tumor cells due to its expression of TEM8, its ease of manufacture, and the ability to arm SVV-001 with antitumor genes, as insertional mutagenesis is not possible with this virus. SVV-001 has been studied in early phase trials in both pediatric and adult patients reporting safety (one DLT reported in 76 patients due to tumor pain) and showing promising signs of efficacy in patients.

[0112] The addition of immune checkpoint inhibitor (CPI) therapy can significantly enhance antitumor efficacy in a variety of cancers, but neuroendocrine and small cell lung cancer (SCLC) tumors have proven refractory to CPI monotherapy. We therefore evaluated the efficacy of combining SVV-001 with a checkpoint inhibitor (anti-PD-1 antibody) CPI therapy in mouse N1E-115 neuroblastoma and B16F10 metastatic melanoma models engineered to express the TEM8 receptor. Both mouse strains are resistant to anti-PD-1 treatment. N1E-115 cells, but not B16F10, endogenously express high levels of TEM8, making them resistant to SVV. Transfection of B16F10 with TEM8 renders these cells susceptible to SVV infection.

[0113] SVV-001 was injected twice weekly as monotherapy or in combination with murine anti-PD1 administered twice weekly. The infected cell type, immune infiltration, SVV replication, and immune responses were investigated along with efficacy. SVV increased response rates 3-6-fold (p<0.01) compared to CPI monotherapy and 6-fold (p<0.01) compared to SVV-001 monotherapy, with improved survival in animals receiving combination therapy. Thus, two syngeneic mouse models for SVV-001 immunotherapy were developed.

[0114] This example also shows that the combination of SVV-001 and anti-PD-1 leads to a significant improvement in anti-tumor response. This study may lay the foundation for the practical application of SVV-001 oncolytic virus therapy in combination with anti-PD-1 antibodies in patients with neuroendocrine tumors. Thus, this example demonstrates that SVV overcomes resistance to checkpoint inhibitor treatment in mouse models of neuroendocrine tumors and melanoma. EXAMPLES

[0115] Example 2: Combination therapy with SVV and checkpoint inhibitors Further studies were conducted investigating combination therapy of SVV and various checkpoint inhibitors. The Pan02 tumor model treatment schedule for this study is shown in Figure 1.

[0116] Research Summary: Pan02 pancreatic syngeneic tumor cells were implanted into the right flank of syngeneic C57BL / 6 mice. Tumors were 100 mm 3 When mice reached 100% CI, treatment with SVV+ / -anti-PD-1;anti-CTLA4 was started with four injections (days 1–12). On day 26, Pan02 tumor cells were injected into the left flank, as well as into naive control mice. Tumor growth and survival were monitored. In this study, murine antibodies anti-PD-1 (RMP1-14) and anti-CTLA-4 (9D9), analogs of nivolumab and ipilimumab (BMS), were used.

[0117] The results of this study are shown in Figures 2A, 2B, 3A, 3B, 3C and 4. The effect of administering SVV, checkpoint inhibitors, and SVV and checkpoint inhibitors to primary tumors is shown in Figure 2. 5 / 6 cures were observed in the SVV+aPD1+aCTLA4 group.

[0118] The effect of administering SVV, checkpoint inhibitors, and SVV and checkpoint inhibitors in the contralateral tumor is shown in Figure 2B. Again, 5 / 6 cures were observed in the SVV+aPD1+aCTLA4 group. As shown in Figure 2B, the combination of SVV, anti-PD-1 and anti-CTLA4 in the contralateral tumor results in 83% cures. There were no cures when the control or checkpoint inhibitors were administered alone.

[0119] Kaplan-Meier plots depicting cell viability as a function of time following treatment with SVV, checkpoint inhibitors, or their combination are shown in Figures 3A and 3B. Figure 3A shows results up to day 115. Figures 3B and 3C show results up to day 165. All remaining animals were sacrificed on day 165. As shown in Figures 3A-3C, SVV+aPD1+aCTLA4 results in long-term cure.

[0120] As can be seen from the plot, the combination of SVV plus anti-PD-1 and anti-CTL4 resulted in the longest survival. Tumor volumes following treatment with SVV, checkpoint inhibitors, or their combination as a function of time are shown in Figure 4. SVV-001 treatment was stopped on day 12. Anti-PD1 / anti-CTLA4 treatment was stopped on day 23.

[0121] Consideration: SVV monotherapy and SVV+anti-PD1 are effective for the duration of treatment (up to day 13); after this time, these tumors grow similarly to PBS-treated mice (i.e., tumors become unresponsive to viral treatment).

[0122] The anti-PD1 + anti-CTLA4 combination shows tumor growth control for approximately 16-20 days, after which the tumor grows, but slower than the PBS control. The SVV + anti-PD1 + anti-CTLA4 (S+P+C) combination shows tumor regression and cure by day 20. Four mice show tumor growth by day 33, after which three mice have regression leading to cure. Overall, 5-6 mice show sustained cure through day 120. In the anti-PD1 + anti-CTLA4 (P+C) group, tumor growth was delayed, but no long-term cure was observed. In the contralateral tumor, 5-6 mice in the S+P+C group showed complete regression, whereas only 1 of 6 mice in the P+C group showed regression. Mice in the S+P+C group are alive and tumor-free beyond day 145. EXAMPLES

[0123] Example 3: Combination therapy with SVV and checkpoint inhibitors Further studies were performed using the protocol of Example 2. Figures 5A and 5B show tumor volumes after treatment with SVV, checkpoint inhibitors, or a combination thereof. Figure 5A shows data up to day 17. Figure 5B shows data up to day 34.

[0124] The efficacy of this study was similar to that shown in Example 2. S+P+C causes tumor regression / cure, while P+C only shows tumor reduction. SVV+anti-CTLA4 does not result in a cure and has an effect similar to anti-CTLA4 alone. Anti-PD1+anti-CTLA4 results in 2 cures out of 8 mice. S+P+C results in 5-7 cures out of 8 mice (depending on treatment group).

[0125] Exemplary embodiments Provided herein are exemplary embodiments of the disclosed technology. These embodiments are merely exemplary and are not intended to limit the scope of the disclosure or the appended claims.

[0126] Embodiment 1. A method of treating cancer in a subject in need thereof, comprising administering to the subject an effective amount of Seneca Valley Virus (SVV) or an SVV derivative and an effective amount of a checkpoint inhibitor, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0127] Embodiment 2. The method of embodiment 1, wherein the cancer is also refractory to SVV monotherapy.

[0128] Embodiment 3. The method of embodiment 1, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, or a combination thereof.

[0129] Embodiment 4. The method of embodiment 2, wherein the method comprises administering a PDL-1 inhibitor and a CTLA-4 inhibitor.

[0130] Embodiment 5. The method of embodiment 1, wherein the method comprises administering an SVV derivative encoding the checkpoint inhibitor.

[0131] Embodiment 6. The method of embodiment 4, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

[0132] Embodiment 7. The method of embodiment 4, further comprising administering an additional checkpoint inhibitor.

[0133] Embodiment 8. The method of embodiment 3, wherein the checkpoint inhibitor is an antibody or nanobody.

[0134] Embodiment 9. The method of embodiment 1, wherein the checkpoint inhibitor is an anti-PD-1 antibody.

[0135] Embodiment 10. The method of embodiment 1, wherein the method comprises administering an anti-PD-1 antibody and an anti-CTLA-4 antibody.

[0136] Embodiment 11. The method according to any one of embodiments 1 to 10, wherein the checkpoint inhibitor is administered before, simultaneously with, or after administration of the Seneca Valley Virus (SVV) or SVV derivative.

[0137] Embodiment 12. The method according to any one of embodiments 1 to 11, wherein the Seneca Valley Virus (SVV) or SVV derivative is administered intratumorally.

[0138] Embodiment 13. The method of any one of embodiments 1 to 12, wherein the checkpoint inhibitor is administered systemically.

[0139] Embodiment 14. The method according to any one of embodiments 1 to 12, wherein the Seneca Valley Virus (SVV) or SVV derivative is administered intratumorally and the checkpoint inhibitor is administered systemically.

[0140] Embodiment 15. The method according to any one of embodiments 1 to 14, wherein the treatment is improved compared to monotherapy with Seneca Valley Virus (SVV) or an SVV derivative or said checkpoint inhibitor.

[0141] Embodiment 16. The method according to any one of embodiments 1 to 15, wherein the Seneca Valley Virus (SVV) or SVV derivative and the checkpoint inhibitor are administered at the same dosing interval.

[0142] Embodiment 17. The method of embodiment 11, wherein the Seneca Valley Virus (SVV) or SVV derivative and checkpoint inhibitor are administered weekly.

[0143] Embodiment 18. The method according to any one of embodiments 1 to 18, wherein the cancer is neuroblastoma or melanoma.

[0144] Embodiment 19. The method according to any one of embodiments 1 to 18, wherein the cancer is a neuroendocrine cancer or a small cell lung cancer (SCLC) tumor.

[0145] Embodiment 20. The method of any one of embodiments 1 to 18, wherein the cancer comprises triple-negative breast cancer, small cell lung cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer.

[0146] Embodiment 21. A pharmaceutical composition for treating cancer in a subject, the pharmaceutical composition comprising a checkpoint inhibitor, SVV or an SVV derivative, and a pharma- ceutical acceptable carrier, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0147] Embodiment 22. The pharmaceutical composition of embodiment 21, wherein the cancer is also refractory to monotherapy with SVV.

[0148] Embodiment 23. The pharmaceutical composition of embodiment 21 or 22, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

[0149] Embodiment 24. The pharmaceutical composition of embodiment 23, wherein the checkpoint inhibitor is an antibody or a nanobody.

[0150] Embodiment 25. The pharmaceutical composition of embodiment 21, wherein the checkpoint inhibitor is an anti-PD-1 antibody.

[0151] Embodiment 26. A pharmaceutical composition according to any one of embodiments 21 to 23, wherein the composition comprises an SVV derivative encoding the checkpoint inhibitor.

[0152] Embodiment 27. The pharmaceutical composition of embodiment 26, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

[0153] Embodiment 28. The pharmaceutical composition according to any one of embodiments 21 to 26, wherein the composition comprises an anti-PD-1 antibody and an anti-CTLA-4 antibody.

[0154] Embodiment 29. The pharmaceutical composition according to any one of embodiments 21 to 28, wherein the cancer is neuroblastoma or melanoma.

[0155] Embodiment 30. The pharmaceutical composition according to any one of embodiments 21 to 21, wherein the cancer is a neuroendocrine cancer or a small cell lung cancer (SCLC) tumor.

[0156] Embodiment 31. The pharmaceutical composition according to any one of embodiments 21 to 28, wherein the cancer comprises triple-negative breast cancer, small cell lung cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer.

[0157] Embodiment 32. A kit for treating cancer in a subject, comprising a Seneca Valley Virus (SVV) or an SVV derivative in combination with a checkpoint inhibitor, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0158] Embodiment 33. The kit of embodiment 32, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

[0159] Embodiment 34. The kit of embodiment 32, wherein the checkpoint inhibitor is an antibody or nanobody.

[0160] Embodiment 35. The kit of embodiment 32, wherein the kit comprises an SVV derivative encoding the checkpoint inhibitor.

[0161] Embodiment 36 The kit of embodiment 32, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

[0162] Embodiment 37. The kit according to any one of embodiments 32 to 36, wherein the cancer is a neuroblastoma, melanoma, neuroendocrine carcinoma, or small cell lung cancer (SCLC) tumor.

[0163] Embodiment 38. A kit according to any one of embodiments 32 to 36, wherein the cancer comprises triple-negative breast cancer, small cell lung cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer.

[0164] Embodiment 39. A Seneca Valley Virus (SVV) or an SVV derivative (SVV) in combination with a checkpoint inhibitor for use in the manufacture of a medicament for the treatment of cancer, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

[0165] Embodiment 40. The combination according to embodiment 39, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

[0166] Embodiment 41. The combination according to embodiment 39, wherein the checkpoint inhibitor is an antibody or a nanobody.

[0167] Embodiment 42. The combination according to embodiment 39, wherein the SVV derivative encodes the checkpoint inhibitor.

[0168] Embodiment 43. The combination according to embodiment 42, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

[0169] Embodiment 44. The combination according to any one of embodiments 39 to 43, wherein the cancer is a neuroblastoma, melanoma, neuroendocrine carcinoma, or small cell lung cancer (SCLC) tumor.

[0170] Embodiment 45. The combination of any one of embodiments 39 to 43, wherein the cancer comprises triple-negative breast cancer, small cell lung cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer.

[0171] Although the present disclosure has been described in conjunction with its preferred specific embodiments, it will be understood that the foregoing description and the following examples are intended to illustrate, but not to limit, the scope of the present disclosure. It will be understood by those skilled in the art that various modifications can be made and equivalents can be substituted without departing from the scope of the present disclosure, and that other aspects, advantages, and modifications will be apparent to those skilled in the art to which the present disclosure pertains. In addition to the embodiments described herein, the present disclosure contemplates and claims inventions resulting from combinations of features of the disclosure cited herein and features of the cited prior art documents that complement the features of the present disclosure. Similarly, it will be understood that any described material, feature, or article can be used in combination with any other material, feature, or article, and such combinations are considered to be within the scope of the present disclosure.

[0172] The disclosures of each patent, patent application, and publication cited or described in this specification are hereby incorporated by reference in their entirety for all purposes.

Claims

1. A pharmaceutical composition comprising a checkpoint inhibitor, SVV or an SVV derivative, and a pharma- ceutically acceptable carrier.

2. 2. The pharmaceutical composition of claim 1, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

3. 3. The pharmaceutical composition of claim 2, wherein the checkpoint inhibitor is an antibody or a nanobody.

4. 2. The pharmaceutical composition of claim 1, wherein the checkpoint inhibitor is an anti-PD-1 antibody.

5. 2. The pharmaceutical composition of claim 1, wherein the composition comprises an SVV derivative encoding the checkpoint inhibitor.

6. The pharmaceutical composition of claim 5, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

7. 2. The pharmaceutical composition of claim 1, wherein the composition comprises an anti-PD-1 antibody and an anti-CTLA-4 antibody.

8. A kit comprising Seneca Valley Virus (SVV) or an SVV derivative and a checkpoint inhibitor, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

9. 9. The kit of claim 8, wherein the checkpoint inhibitor is an antibody or a nanobody.

10. 10. The kit of claim 8 or 9, wherein the kit comprises an SVV derivative encoding the checkpoint inhibitor.

11. The kit of claim 10, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

12. A Seneca Valley Virus (SVV) or SVV derivative for use in treating cancer in a subject in need thereof, said use comprising administering to said subject an effective amount of SVV or an SVV derivative and an effective amount of a checkpoint inhibitor, wherein said cancer is refractory to monotherapy with said checkpoint inhibitor or is refractory to monotherapy with SVV.

13. 13. The SVV or SVV derivative for use according to claim 12, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, or a combination thereof.

14. 14. An SVV or SVV derivative for use according to claim 12 or 13, said use comprising administering a PDL-1 inhibitor and a CTLA-4 inhibitor, or an SVV derivative encoding said checkpoint inhibitor.

15. 15. The SVV or SVV derivative for use according to claim 14, wherein the SVV derivative encodes a PD-1 inhibitor, a CTLA-4 inhibitor, or both.

16. SVV or an SVV derivative for use according to any one of claims 12 to 15, wherein the checkpoint inhibitor is an antibody or a nanobody.

17. SVV or an SVV derivative for use according to any one of claims 12 to 16, wherein the checkpoint inhibitor is an anti-PD-1 antibody, or an anti-PD-1 antibody and an anti-CTLA-4 antibody.

18. 17. The SVV or SVV derivative for use according to any one of claims 12 to 16, wherein the checkpoint inhibitor is administered before, simultaneously with or after administration of the Seneca Valley Virus (SVV) or SVV derivative, the Seneca Valley Virus (SVV) or SVV derivative is administered intratumorally and / or the checkpoint inhibitor is administered systemically.

19. 19. The Seneca Valley Virus (SVV) or SVV derivative for use according to any one of claims 12 to 18, wherein said treatment is improved compared to monotherapy with SVV or an SVV derivative or said checkpoint inhibitor.

20. 20. The Seneca Valley Virus (SVV) or SVV derivative for use according to any one of claims 12 to 19, wherein the Seneca Valley Virus (SVV) or SVV derivative and the checkpoint inhibitor are administered at the same dosing interval, or the Seneca Valley Virus (SVV) or SVV derivative and the checkpoint inhibitor are administered weekly.

21. 21. The SVV or SVV derivative for use according to any one of claims 12 to 20, wherein the cancer is neuroblastoma, melanoma, neuroendocrine cancer, small cell lung cancer (SCLC), triple negative breast cancer, non-small cell lung cancer, non-small cell squamous cell carcinoma, adenocarcinoma, glioblastoma, skin cancer, hepatocellular carcinoma, colorectal cancer, cervical cancer, ovarian cancer, endometrial cancer, neuroendocrine cancer, pancreatic cancer, thyroid cancer, renal cancer, bone cancer, esophageal cancer, or soft tissue cancer.

22. 1. A Seneca Valley Virus (SVV) or an SVV derivative (SVV) in combination with a checkpoint inhibitor for use in the manufacture of a medicament for the treatment of cancer, wherein the cancer is refractory to monotherapy with the checkpoint inhibitor.

23. 23. The combination of claim 22, wherein the checkpoint inhibitor is a PD-1 inhibitor, a PD-L1 inhibitor, or a CTLA-4 inhibitor.

24. 23. The combination of claim 22, wherein the checkpoint inhibitor is an antibody or a nanobody.

25. 23. The combination of claim 22, wherein the SVV derivative encodes the checkpoint inhibitor.

Images