Combination of checkpoint inhibitors and oncolytic viruses for treatment of cancer

Abstract

The present disclosure relates to methods for inhibiting tumor growth in a patient in need thereof. In some embodiments, the methods comprise administering to the patient an oncolytic virus in combination with a therapeutically effective amount of a PD-1 inhibitor (e.g., samipril monoclonal antibody or a biological equivalent thereof) and optionally a therapeutically effective amount of a CTLA-4 inhibitor (e.g., ipimumab or a biological equivalent thereof). In some embodiments, administration of the combination results in enhanced tumor inhibition as compared to administration of the antibody or virus alone. In some embodiments, the oncolytic virus is a recombinant vesicular stomatitis virus (VSV), such as VV1.

Description

Technical Field

[0001] The present disclosure generally relates to combination therapy for cancer treatment with oncolytic viruses and checkpoint inhibitors (e.g., programmed death 1 (PD-1) pathway inhibitors and / or cytotoxic T-lymphocyte antigen-4 (CTLA-4) inhibitors). Background Art

[0002] VV1 (also known as VSV-IFNβ-NIS) is derived from vesicular stomatitis virus (VSV), a bullet-shaped, negative-sense ribonucleic acid (RNA) virus with low human seropositivity; it is engineered to selectively replicate in and kill human cancer cells. The virus encodes human interferon β (IFNβ) to enhance antitumor immune responses and improve tumor specificity, plus the thyroid sodium-iodine symporter (NIS) to allow imaging of viral spread. VSV-IFNβ-NIS synergizes with different anti-PD-(L)1 antibodies in several tumor models.

[0003] VV1 is a live RNA virus modified to express both human IFNβ genes and NIS genes. The virus is constructed by inserting the IFNβ gene downstream of the M gene and the NIS gene (cDNA) downstream of the G protein gene into the full-length infectious molecular clone of the Indiana VSV strain [Naik 2012]. The virus is not a vaccine. VSV-IFNβ-NIS propagates on BHK cells with similar kinetics to the parental virus strain and can grow to high titers. It selectively propagates in human cancer cells because many cancer cells cannot enhance effective antiviral responses mediated by the IFNβ pathway. However, IFNβ production from infected cells will be used to protect non-cancerous cells from the effects of the virus. As a result, the virus is directly cytopathic to tumor cells, resulting in rapid lysis as the virus amplifies [Barber 2004; Lichty 2004)]. Preclinical studies have shown that IV treatment with oncolytic VSV selectively targets tumor cells, inducing an acute inflammatory response and IT immune infiltration [Obuchi 2003; Willmon 2009; Saloura 2010; Naik 2012; Kurisetty 2014; Patel 2015].

[0004] VSV-IFNβ-NIS-infected tumor cells also express NIS, a membrane ion channel that actively transports iodide into the cell. The uptake of radioiodine by NIS-expressing cells provides the basis for in vivo imaging using 99mTc pertechnetate or radioiodine I-123, which can reveal the time-dependent profile of VSV-IFNβ-NIS gene expression and the location of VSV-IFNβ-NIS-infected cells during viral spread and clearance [Naik 2012]. F18-tetrafluoroborate (F18-TFB) PET imaging can also be used to track viral replication and spread [Miller 2016]. Recently, the safety and feasibility of using F18-TFB PET imaging to locate tissues that physiologically express NIS was demonstrated in healthy human subjects [Jiang 2017].

[0005] Programmed death 1 (PD-1) plays an important role in autoimmunity, tumor immunity and infection immunity, and is therefore an ideal target for immunotherapy. Blocking PD-1 with antagonists (including monoclonal antibodies) has been studied in the treatment of cancer and chronic viral infections. Blocking of PD-1 is also an effective and well-tolerated method of stimulating immune responses, and has achieved therapeutic advantages for a variety of human cancers, including melanoma, renal cell carcinoma and non-small cell lung cancer (NSCLC).

[0006] (Sheridan 2012, Nat. Biotechnol., 30: 729-730; Postow et al., 2015, J ClinOncol, 33: 1974-1982; Chen et al. al., 2013, Nat. Rev. Immunol., 13: 227-242; Riley, 2009, Immunol. Rev., 229: 114-125; Dong et al., 1999, Nature Med., 5(12): 1365-1369; Zou, 2008, Nat.Rev.Immunol., 8: 467-77; Ribas 2012, NEJM 366: 2517-2519; Watanabe et al., 2012, Clin.Dev.Immunol.Vol.2012, Article ID: 269756; Wang et al., 2013, J. ViralHep., 20: 27-39; Flies et al., 2011, Yale J. Biol. Med., 84: 409-421; Pardoll, 2012, Nature, 12: 252-264; Freeman, 2008, PNAS, 105: 10275-10276; Francisco et al., 2010, Immunol. Rev, 236: 219-242).

[0007] Monoclonal antibodies against PD-1 are known in the art and have been used in, for example,

[0008] US 9987500, US 8008449, US 8168757, US 20110008369, US 20130017199, US20130022595, WO 2006121168, WO 20091154335, WO 2012145493, WO 2013014668, WO2009101611. EP 2262837 and EP 2504028. Cemiplimab (also known as REGN2810; ) For example, it is a high-affinity, fully human, hinge-stabilized IgG4P antibody against the PD-1 receptor, which can effectively block the interaction between PD-1 and its ligands PD-L1 and PD-L2.

[0009] Blockade of cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) has been shown to increase the number of activated T cells, thereby allowing the expansion of tumor-specific cytotoxic T cells. CTLA-4 is a type I transmembrane T cell inhibitory checkpoint receptor expressed on conventional and regulatory T cells. CTLA-4 negatively regulates T cell activation by outcompeting the binding of the stimulatory receptor CD28 to its shared natural ligands B7-1 (CD80) and B7-2 (CD86). Ipilimumab is a recombinant humanized monoclonal antibody that binds to CTLA-4.

[0010] Lung cancer is the leading cause of cancer death worldwide [Bray 2018]. In the United States, it is estimated that approximately 228,000 patients will be diagnosed with lung cancer in 2019, and 80% to 85% of these cases will be of NSCLC histology [ACS 2019d]. The etiological association of NSCLC with exposure to tobacco carcinogens is particularly relevant to cancer immunotherapy, as current approaches targeting immune checkpoints show improved responses in tumors with a high number of somatic mutations, as is the case in smoking-induced NSCLC. In contrast, immune checkpoint blockade has been significantly less successful in never-smokers, including the majority of NSCLC with anaplastic lymphoma kinase ALK rearrangements or epidermal growth factor receptor (EGFR) mutations [Garassino 2017]. Most patients with NSCLC present with an advanced stage of disease and are symptomatic at diagnosis, with an associated poor prognosis and no curative options. In stage IV, systemic palliative therapy is recommended, including cytotoxic chemotherapy, immune checkpoint inhibitors (CPIs), and a range of molecularly selected targeted agents in defined subgroups of NSCLC patients. Despite significant advances in the treatment of NSCLC with anti-PD-(L)1 therapy, advanced disease remains largely incurable.

[0011] Melanoma remains quite common, with over 96,000 new cases diagnosed in the United States in 2019, but overall mortality remains low [Siegel 2019]. However, despite recent advances in CPI therapy, metastatic melanoma continues to maintain a relatively high incidence. Overall survival for patients with advanced melanoma has improved from approximately 9 months before 2011 to at least 2 years and may be longer for specific mutations, such as BRAF V600 mutant melanoma [Luke 2017], with a subset of patients experiencing long-term tumor control. Melanoma is amenable to immunotherapy for a number of reasons, including extensive tumor infiltration of T cells, high mutational load, and crosstalk between oncogenic signaling pathways and immunobiology. The use of ipilimumab, nivolumab, and pembrolizumab has become the standard of care for patients with advanced metastatic melanoma. However, despite long-term benefit in several patients, durable responses remain elusive. Resistance mechanisms to immunotherapy, such as loss of tumor antigen presentation and T-cell trafficking, have been observed.

[0012] Approximately 65,000 patients are diagnosed with head and neck squamous cell carcinoma (HNSCC) each year in the United States and approximately 14,000 deaths occur annually secondary to HNSCC [Siegel 2019]. Globally, more than 600,000 patients are diagnosed with the disease each year [Bray 2018]. Head and neck cancers consist of a spectrum of tumors that arise from the squamous cells lining the mucosal surfaces of the head and neck, referred to as HNSCC, and are classified according to the region of the head or neck in which they arise: oral cavity, pharynx (including nasopharynx, oropharynx, and hypopharynx), paranasal sinuses, nasal cavity, and salivary glands. Infection with human papillomavirus (HPV), particularly HPV-16 and HPV-18, is a risk factor for developing HNSCC, particularly oropharyngeal cancer involving the tonsils or base of the tongue [Chaturvedi 2011]. In the United States, the incidence of oropharyngeal cancer caused by HPV infection is increasing, whereas the incidence of oropharyngeal cancer associated with other causes is decreasing [Chaturvedi 2011].

[0013] When HNSCC is detected and treated in its earliest stages, it is curable with excellent long-term OS. However, once patients develop local recurrence or metastatic disease, HNSCC remains essentially incurable, with 5-year survival less than 50% [ACS2020]. A monoclonal antibody targeting the T-cell immune CPIPD-(L)1 has recently shown significant antitumor activity in patients with advanced HNSCC. Pembrolizumab received accelerated approval based on response rate and durability of response in patients with recurrent or metastatic HNSCC whose disease has progressed after platinum-containing chemotherapy (2nd line), regardless of HPV status. In this population, the ORR following pembrolizumab was approximately 16% [Chow 2016]. However, although recent advances in the treatment of advanced HNSCC have improved patients' OS, the disease remains incurable, and pembrolizumab has not improved response rates beyond the EXTREME, only the duration of response.

[0014] Colorectal cancer (CRC) is one of the most diagnosed cancers in the world, with an incidence rate that ranks second in women and third in men worldwide [Sung 2021]. In the United States, approximately 149,500 new cases of colorectal cancer are diagnosed each year, and approximately 52,980 Americans die from CRC, making it the third leading cause of cancer death in both men and women [Siegel 2021]. Approximately 50% to 60% of patients with colorectal cancer have metastatic disease at the time of diagnosis, with common sites of involvement including the liver, lymph nodes, lungs, peritoneum, and soft tissues [Van Cutsem 2006, Lee 2007, Yoo 2006], and approximately half of early-stage patients eventually develop metastatic or advanced disease [Martini 2017]. Survival outcomes for patients with metastatic CRC are poor, with a 5-year survival rate of approximately 14% [Bray 2018].

[0015] Compared with HNSCC, survival of patients with CRC treated with novel therapies has been only modestly improved, and this is largely due to the lack of efficacy of anti-PD-(L)1 therapy, except in a small group of patients with microsatellite instability high (MSI-H) CRC. The exact means by which microsatellite stable (MSS) CRCs evade immune detection and destruction is not fully understood. MSS CRCs can be classified into 2 patterns that represent non-inflammatory tumors. The first is through immunological ignorance, in which the tumor is rarely infiltrated by lymphocytes, and the second is through an immunosuppressive microenvironment that is created to evade immune surveillance by recruiting myeloid-derived suppressor cells or by secreting factors including TGFβ, thereby inhibiting the chemokines and cytokines required to promote T cell infiltration into the tumor [Hegde 2016].

[0016] Improvements in conventional treatments (e.g., chemotherapy, radiation therapy, surgery, and targeted therapy) and recent advances in immunotherapy have improved outcomes for patients with advanced solid tumors. Over the past few years, the FDA and the European Medicines Agency have approved six CPIs: ipilimumab, a monoclonal antibody that targets the CTLA-4 pathway, and six antibodies targeting PD-L / PD-L1, including atezolizumab, avelumab, durvalumab, nivolumab, cemiplimab, and pembrolizumab, for the treatment of patients with a variety of cancer types, primarily solid tumors [Gentzler 2016; Ribas 2017]. These approvals have dramatically changed the landscape of cancer treatment. However, the majority of cancer patients, including the majority with tumors that are considered “sensitive” to these agents, such as melanoma, NSCLC, urothelial carcinoma, renal cancer, etc., do not respond to or become resistant to these agents [Arora 2019]. Additionally, some of the most prevalent tumors, colorectal, breast, and prostate cancers, have been shown to be largely refractory to checkpoint inhibition [Borcherding 2018].

[0017] Many immunotherapeutic treatments have shown efficacy only in selected cancer subgroups, such as those expressing PD-L1[ 2019], cancer subgroups with microsatellite instability (MSI) / mismatch repair (MMR) deficiency or high tumor mutational burden [Luchini 2019], and even new targeted agents address only very small subsets of tumor types [ 2019]. In fact, despite considerable early success and relatively few side effects compared with other systemic therapies, the majority of cancer patients do not respond to CPIs or new targeted therapies [Arora 2019].

[0018] As cure rates remain low in patients with advanced solid tumors, there remains an unmet medical need for more effective and less toxic treatments, such as combination therapies with synergistic mechanisms of action with the immuno-CPIs disclosed herein. Summary of the invention

[0019] In one aspect, the disclosed technology relates to a method for treating a tumor or inhibiting tumor growth, comprising: (a) selecting a patient suffering from cancer; and (b) administering a combination therapy to a patient in need thereof, wherein the combination therapy comprises: (i) a therapeutically effective amount of an oncolytic virus; and (ii) a therapeutically effective amount of an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the anti-PD-1 antibody or the antigen-binding fragment thereof comprises three heavy chain complementary determining regions (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1 and three light chain complementary determining regions (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO: 2.

[0020] In some embodiments, the oncolytic vesicular virus includes an oncolytic vesicular stomatitis virus (VSV). In some embodiments, VSV includes recombinant VSV. In some embodiments, the recombinant VSV expresses cytokines. In some embodiments, the cytokine includes interferon-β (IFNb). In some embodiments, the nucleic acid sequence encoding IFNb is located between the M and G viral genes. In some embodiments, the recombinant VSV also expresses a sodium / iodine symporter (NIS). In some embodiments, the nucleic acid sequence encoding NIS is located between the G and L viral genes. In some embodiments, the oncolytic virus is Voyager V1. In some embodiments, the anti-PD-1 antibody or its antigen-binding fragment comprises HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 containing the amino acid sequences of SEQ ID NO:3, 4, 5, 6, 7 and 8, respectively. In some embodiments, the anti-PD-1 antibody or its antigen-binding fragment comprises an HCVR containing the amino acid sequence of SEQ ID NO:1; and an LCVR containing the amino acid sequence of SEQ ID NO:2. In some embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 9 and a light chain comprising the amino acid sequence of SEQ ID NO: 10. In some embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof comprises cemiplimab or a bioequivalent thereof.

[0021] In some embodiments, the oncolytic virus is present in one or more 10 to 10 14 TCID 50 In some embodiments, the oncolytic virus is administered to a patient at a dose of 10 9 TCID 50 or 10 11 TCID 50 In some embodiments, a 10 9 TCID 50 dose of oncolytic virus was administered to the patient and another 10 11 TCID 50 In some embodiments, the anti-PD-1 antibody or its antigen-binding fragment is administered to the patient at a dose of one or more about 0.1 mg / kg to about 20 mg / kg of the patient's body weight. In some embodiments, the anti-PD-1 antibody or its antigen-binding fragment is administered to the patient at a dose of one or more about 1 mg to about 1000 mg. In some embodiments, the anti-PD-1 antibody or its antigen-binding fragment is administered to the patient at a dose of one or more 350 mg.

[0022] In some embodiments, the oncolytic virus is administered to the patient intratumorally or intravenously. In some embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient intravenously, subcutaneously, or intraperitoneally.

[0023] In some embodiments, the method further comprises administering a therapeutically effective amount of a CTLA-4 inhibitor, the CTLA-4 inhibitor comprising ipilimumab or a bioequivalent thereof. In some embodiments, wherein the CTLA-4 inhibitor is ipilimumab.

[0024] In some embodiments, the method includes administering one or more doses of an oncolytic virus and one or more doses of a CTLA-4 inhibitor simultaneously. In some embodiments, the method includes administering one or more doses of an oncolytic virus and one or more doses of a CTLA-4 inhibitor prior to administering one or more doses of an anti-PD-1 antibody or an antigen-binding fragment thereof. In some embodiments, the method includes administering one or more doses of an oncolytic virus and one or more doses of a CTLA-4 inhibitor 7 days prior to administering one or more doses of an anti-PD-1 antibody or an antigen-binding fragment thereof.

[0025] In some embodiments, the CTLA-4 inhibitor is administered to a patient at one or more doses of about 0.1 mg / kg to about 15 mg / kg of patient body weight. In some embodiments, the CTLA-4 inhibitor is administered to a patient at one or more doses of about 1 mg to about 600 mg. In some embodiments, the CTLA-4 inhibitor is administered to a patient at one or more doses of 50 mg. In some embodiments, the CTLA-4 inhibitor is administered to a patient intravenously, subcutaneously, or intraperitoneally.

[0026] In some embodiments, the cancer is selected from adrenal tumors, bile duct cancer, bladder cancer, brain cancer, breast cancer, epithelial cancer, cancer of central or peripheral nervous system tissue, cervical cancer, colon cancer, endocrine or neuroendocrine cancer or hematopoietic cancer, esophageal cancer, fibroma, gastrointestinal cancer, glioma, head and neck cancer, Ley-Fraumeni tumor, liver cancer, lung cancer, lymphoma, melanoma, meningioma, multiple neuroendocrine neoplasms type I and II, nasopharyngeal cancer, oral cancer, oropharyngeal cancer, osteosarcoma tumors, ovarian cancer, pancreatic cancer, islet cell carcinoma, parathyroid cancer, pheochromocytoma, pituitary tumor, prostate cancer, rectal cancer, kidney cancer, respiratory cancer, sarcoma, skin cancer, stomach cancer, testicular cancer, thyroid cancer, tracheal cancer, genitourinary cancer, and uterine cancer.

[0027] In some embodiments, the administration of combined treatment produces at least one selected from the following therapeutic effects: reduced number of tumor cells, tumor regression, partial response and complete response. In some embodiments, compared with patients treated with a single treatment selected from an oncolytic virus and an anti-PD-1 antibody or its antigen-binding fragment, the administration of combined treatment produces at least one selected from the following improvements: abscopal effect, tumor growth delay, overall survival improvement, progression-free survival improvement, total response rate improvement, complete response improvement, partial response improvement and disease stabilization improvement. In some embodiments, compared with patients treated with an oncolytic virus monotherapy, the administration of combined treatment produces at least one selected from the following improvements: abscopal effect, tumor growth delay, overall survival improvement, progression-free survival improvement, total response rate improvement, complete response improvement, partial response improvement and disease stabilization improvement. In some embodiments, the tumor growth is inhibited by at least 50% compared with untreated patients.

[0028] In some embodiments, compared with patients treated with a single treatment selected from an oncolytic virus, an anti-PD-1 antibody or its antigen-binding fragment and a CTLA-4 inhibitor, the administration of a combination therapy produces at least one improvement selected from the following: a distant effect, tumor growth delay, overall survival improvement, progression-free survival improvement, overall response rate improvement, complete response improvement, partial response improvement and disease stabilization improvement. In some embodiments, compared with patients who are treated with an oncolytic virus, an anti-PD-1 antibody or its antigen-binding fragment or a CTLA-4 inhibitor as a single treatment, tumor growth is inhibited by at least 50%. In some embodiments, compared with patients who are treated with any two of an oncolytic virus, an anti-PD-1 antibody or its antigen-binding fragment and a CTLA-4 inhibitor, tumor growth is inhibited by at least 50%. In some embodiments, the method further includes administering an additional therapeutic agent or treatment to the patient.In some embodiments, the additional therapeutic agent or treatment is selected from radiation, surgery, chemotherapeutic agents, cancer vaccines, B7-H3 inhibitors, B7-H4 inhibitors, lymphocyte activation gene 3 (LAG3) inhibitors, T cell immunoglobulin and mucin-domain containing-3 (TIM3) inhibitors, galectin 9 (GAL9) inhibitors, V-domain immunoglobulin-containing suppressor of T-cell activation (VISTA) inhibitors, Killer-Cell Immunoglobulin-Like Receptor (KIR) inhibitors, B and T lymphocyte weakening (B and T lymphocyte attenuator (BTLA) inhibitors, T cell immunoreceptor with Ig and ITIM domains (TIGIT) inhibitors, CD47 inhibitors, indoleamine-2,3-dioxygenase (IDO) inhibitors, vascular endothelial growth factor (VEGF) antagonists, angiopoietin-2 (Ang2) inhibitors, transforming growth factor beta (TGFβ) inhibitors, epidermal growth factor receptor (EGFR) inhibitors, antibodies against tumor-specific antigens, Bacillus Calmette-Guerin vaccine, granulocyte-macrophage colony-stimulating factor (GM-CSF), cytotoxins, interleukin 6 receptor (IL-6R) inhibitors, interleukin 4 receptor (IL-4R) inhibitors. 4receptor, IL-4R) inhibitors, IL-10 inhibitors, IL-2, IL-7, IL-12, IL-21, IL-15, antibody-drug conjugates, anti-inflammatory drugs and combinations thereof.

[0029] In another aspect, the disclosed technology relates to a combination of an oncolytic virus, an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), and a CTLA-4 inhibitor including ipilimumab or a bioequivalent thereof, wherein the combination is used in a method for treating a tumor or inhibiting tumor growth, the method comprising: (a) selecting a patient suffering from cancer; and (b) administering a combination therapy to a patient in need thereof, the combination therapy comprising: (i) a therapeutically effective amount of an oncolytic virus; and (ii) a therapeutically effective amount of an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises three heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1 and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO: 2.

[0030] In another aspect, the disclosed technology relates to a combination of an oncolytic virus, an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), and a CTLA-4 inhibitor including ipilimumab or a bioequivalent thereof, wherein the combination is used in a method for treating a tumor or inhibiting tumor growth, the method comprising: (a) selecting a patient suffering from cancer; and (b) administering a combination therapy to a patient in need thereof, the combination therapy comprising: (i) a therapeutically effective amount of an oncolytic virus; (ii) a therapeutically effective amount of an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises three heavy chain complementary determining regions (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1 and three light chain complementary determining regions (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO: 2; and (iii) a therapeutically effective amount of a CTLA-4 inhibitor including ipilimumab or a bioequivalent thereof.

[0031] In another aspect, the disclosed technology relates to a kit comprising a combination of an oncolytic virus and an antibody or antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1) and the following: written instructions for using a therapeutically effective amount of the combination of the oncolytic virus and the anti-PD-1 antibody or antigen-binding fragment thereof to treat a patient's tumor or inhibit the growth of a patient's tumor.

[0032] In another aspect, the disclosed technology relates to a kit comprising an oncolytic virus, an antibody or antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), and a CTLA-4 inhibitor including ipilimumab or a bioequivalent thereof, in combination with the following: written instructions for using a therapeutically effective amount of the combination of the oncolytic virus, the anti-PD-1 antibody or antigen-binding fragment thereof, and the CTLA-4 inhibitor to treat a patient's tumor or inhibit the growth of a patient's tumor.

[0033] The foregoing overview is not intended to limit each aspect of the present disclosure, and other aspects are described in other parts such as the following specific embodiments. The entire document is intended to be associated as a unified disclosure, and it should be understood that the combination of all features described herein is expected, even if the combination of features does not appear together in the same sentence, or paragraph, or part of this document. From the following specific embodiments, other features and advantages of the present invention will become apparent. However, it should be understood that although the specific embodiments and specific examples show some specific embodiments of the present disclosure, they are only provided by way of example, because according to the specific embodiments, various changes and modifications in the spirit and scope of the present disclosure will become apparent to those skilled in the art. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 Schematic representation of the study design described in Example 1 herein. Abbreviations: 1L = first line; 2L - second line; CPI = checkpoint inhibitor; DRC (data review committee) = data review committee; IV = intravenous; NSCLC = non-small cell lung cancer; ORR (overall response rate) = overall response rate; PD-(L)1 = programmed death-(ligand) 1; TCID 50 =median tissue culture infectious dose; VV1=Voyager V1.

[0035] Figure 2 is a schematic diagram of the study design described in Example 2 herein. DETAILED DESCRIPTION

[0036] It should be understood that the present disclosure is not limited to the specific methods and experimental conditions described, because such methods and conditions can vary. It should also be understood that the terms used herein are only for the purpose of describing specific embodiments, and are not intended to be limiting, and the scope of the present disclosure will be limited only by the appended claims. Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those of ordinary skill in the art to which the present disclosure belongs. Although any methods and materials similar to or equivalent to those described herein and materials can be used in the practice or testing of the present disclosure, preferred methods and materials are now described. Unless otherwise stated, all publications mentioned herein are hereby incorporated by reference as a whole.

[0037] The present disclosure generally relates to novel dual or triple combination therapies of oncolytic viruses, PD-1 pathway inhibitors, and CTLA-4 inhibitors, which dual or triple combination therapies exhibit superior activity in inhibiting tumor growth compared to any single therapy, or the triple combination exhibits superior activity in inhibiting tumor growth compared to any dual combination therapy of oncolytic viruses, PD-1 pathway inhibitors, and CTLA-4 inhibitors.

[0038] In one aspect, the present disclosure relates to the use of dual or triple combination therapy in a method for treating a tumor or inhibiting tumor growth, the method comprising: selecting a patient suffering from cancer; and administering to the patient in need thereof: (i) a therapeutically effective amount of an oncolytic virus in combination with (ii) a therapeutically effective amount of a PD-1 pathway inhibitor (e.g., cemiplimab or a bioequivalent thereof) and / or (iii) a therapeutically effective amount of a CTLA-4 inhibitor (e.g., ipilimumab or a bioequivalent thereof).

[0039] The term "patient" as used herein can be used interchangeably with the term "subject". The expression "subject in need thereof" means such a human or non-human mammal that exhibits one or more symptoms or indications of cancer and / or has been diagnosed with cancer. In some embodiments, a human subject may be diagnosed with a primary or metastatic tumor and / or has one or more symptoms or indications including, but not limited to, enlarged lymph nodes, swollen abdomen, chest pain / pressure, unexplained weight loss, fever, night sweats, persistent fatigue, loss of appetite, enlarged spleen, pruritus. This expression includes patients who have received one or more cycles of chemotherapy with toxic side effects. In some embodiments, the expression "subject in need thereof" includes patients who have cancer, have been treated for it, but have subsequently relapsed or metastasized. For example, such a patient is treated with the methods of the present disclosure: the patient may have received treatment with one or more anticancer agents, resulting in tumor regression, but then relapsed with cancer resistant to one or more anticancer agents (e.g., chemotherapy-resistant cancer).

[0040] As used herein, the terms "treat," "treat," "treat," and variations thereof mean to alleviate or reduce the severity of at least one symptom or indication, eliminate the cause of a symptom on a temporary or permanent basis, delay or inhibit tumor growth, reduce tumor cell load or tumor burden, promote tumor regression, cause tumor shrinkage, necrosis and / or disappearance, prevent tumor recurrence, prevent or inhibit metastasis, inhibit the growth of metastatic tumors, eliminate the need for radiation or surgery, and / or increase the duration of patient survival.

[0041] In many embodiments, the terms "tumor", "lesion", "neoplastic lesion", "cancer", and "malignancy" are used interchangeably and refer to one or more cancerous growths. In some embodiments, the cancer is selected from adrenal tumors, bile duct cancer, bladder cancer, brain cancer, breast cancer, epithelial cancer, cancer of central or peripheral nervous system tissue, cervical cancer, colon cancer, endocrine or neuroendocrine cancer or hematopoietic cancer, esophageal cancer, fibroma, gastrointestinal cancer, glioma, head and neck cancer, Ley-Fraumeni tumor, liver cancer, lung cancer, lymphoma, melanoma, meningioma, multiple neuroendocrine type I and II neoplasms, nasopharyngeal cancer, oral cancer, oropharyngeal cancer, osteosarcoma tumors, ovarian cancer, pancreatic cancer, islet cell carcinoma, parathyroid cancer, pheochromocytoma, pituitary tumor, prostate cancer, rectal cancer, kidney cancer, respiratory cancer, sarcoma, skin cancer, stomach cancer, testicular cancer, thyroid cancer, tracheal cancer, genitourinary cancer, and uterine cancer.

[0042] According to some embodiments, the present disclosure includes methods for treating tumors, delaying or inhibiting tumor growth. In some embodiments, the present disclosure includes methods for promoting tumor regression. In some embodiments, the present disclosure includes methods for reducing tumor cell load or reducing tumor burden. In some embodiments, the present disclosure includes methods for preventing tumor recurrence.

[0043] In some embodiments, the method includes administering one or more doses of an oncolytic virus to a patient before, after, or simultaneously with administering one or more doses of a PD-1 pathway inhibitor and / or one or more doses of a CTLA-4 inhibitor to the patient. In some embodiments, one or more doses of a PD-1 pathway inhibitor may be administered in combination with one or more doses of a CTLA-4 inhibitor.

[0044] The term "combination" as used herein includes the sequential or simultaneous administration of two or more of an oncolytic virus, a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or an antigen-binding fragment thereof), and a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or an antigen-binding fragment thereof). In some embodiments, one treatment disclosed herein may be administered to a patient before or after the simultaneous administration of two other treatments disclosed herein.

[0045] For example, when a first treatment is administered "before" a second and / or third treatment, one or more doses of the first treatment can be administered about 12 weeks, about 11 weeks, about 10 weeks, about 9 weeks, about 8 weeks, about 7 weeks, about 6 weeks, about 5 weeks, about 4 weeks, about 3 weeks, about 2 weeks, about 7 days, about 150 hours, about 150 hours, about 100 hours, about 72 hours, about 60 hours, about 48 hours, about 36 hours, about 24 hours, about 12 hours, about 10 hours, about 8 hours, about 6 hours, about 4 hours, about 2 hours, about 1 hour, about 30 minutes, about 15 minutes, or about 10 minutes prior to administration of one or more doses of the second and / or third treatment.

[0046] For example, when a first treatment is administered "after" a second and / or third treatment, one or more doses of the first treatment can be administered about 12 weeks, about 11 weeks, about 10 weeks, about 9 weeks, about 8 weeks, about 7 weeks, about 6 weeks, about 5 weeks, about 4 weeks, about 3 weeks, about 2 weeks, about 7 days, about 150 hours, about 150 hours, about 100 hours, about 72 hours, about 60 hours, about 48 hours, about 36 hours, about 24 hours, about 12 hours, about 10 hours, about 8 hours, about 6 hours, about 4 hours, about 2 hours, about 1 hour, about 30 minutes, about 15 minutes, or about 10 minutes after administration of one or more doses of the second and / or third treatment.

[0047] As used herein, "simultaneous" administration means that two or more of an oncolytic virus, a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or an antigen-binding fragment thereof), and a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or an antigen-binding fragment thereof) are administered to a patient in separate dosage forms on the same day (e.g., within a 10-minute period), or a single combined dose formulation comprising two or more of an oncolytic virus, a PD-1 pathway inhibitor, and a CTLA-4 inhibitor is administered to a patient.

[0048] In some embodiments, the disclosed methods may further comprise administering an anti-tumor therapy. Anti-tumor therapy includes, but is not limited to, conventional anti-tumor therapy, such as chemotherapy, radiation, surgery, or as described elsewhere herein.

[0049] In some embodiments, the treatment produces a therapeutic effect selected from one or more of the following: tumor growth delay, tumor cell number reduction, tumor regression, survival improvement, partial response and complete response. In some embodiments, compared with tumor growth in untreated patients, tumor growth in patients is delayed by at least 10 days. In some embodiments, compared with untreated patients, tumor growth is inhibited by at least 20% (for example, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%). In some embodiments, compared with patients who are administered oncolytic viruses, PD-1 pathway inhibitors or CTLA-4 inhibitors as monotherapy, tumor growth is inhibited by at least 20% (for example, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%). In some embodiments, tumor growth is inhibited by at least 20% (e.g., at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 200%, at least 300%) compared to patients administered an oncolytic virus alone, or only two of an oncolytic virus, a PD-1 pathway inhibitor, and a CTLA-4 inhibitor.

[0050] In some embodiments, the administration of combination therapy (oncolytic virus with PD-1 pathway inhibitor and / or CTLA-4 inhibitor) produces at least one selected from the following therapeutic effects: reduction in tumor cell number, tumor regression, partial response and complete response. In some embodiments, compared with patients treated with oncolytic virus as a monotherapy, or compared with patients treated with PD-1 pathway inhibitor as a monotherapy, and / or compared with patients treated with CTLA-4 inhibitor as a monotherapy, the administration of combination therapy (oncolytic virus with PD-1 pathway inhibitor and / or CTLA-4 inhibitor) produces at least one selected from the following improvements: abscopal effect, tumor growth delay, overall survival improvement, progression-free survival improvement, overall response rate improvement, complete response improvement, partial response improvement and disease stabilization improvement.

[0051] In some embodiments, administering the combination therapy (an oncolytic virus in combination with a PD-1 pathway inhibitor and / or a CTLA-4 inhibitor) comprises selecting a subject having a first tumor lesion and a second tumor lesion, and administering one or more doses of the combination therapy to the first tumor lesion but not the second tumor lesion, wherein the administration results in tumor regression in both the first and second (i.e., administered and non-administered) tumor lesions, thereby achieving an abscopal effect.

[0052] Oncolytic viruses

[0053] Oncolytic viruses are cancer treatments that use modified or naturally evolved cancer-tropic viruses to stimulate tumor cell death in treated patients. In general, when a replicating oncolytic virus is administered, the infected tumor cells have the potential to produce progeny viruses, allowing the destructive infection to spread to adjacent tumor cells. The potential for viral replication is determined by the ability of the cell to sense and respond to viral infection.

[0054] In some embodiments, the oncolytic virus is a recombinant VSV (eg, VV1). In some embodiments, the oncolytic virus is a replication-competent oncolytic rhabdovirus. Such oncolytic rhabdoviruses include, but are not limited to, wild-type or genetically modified Arajas virus, Chandipura virus, Cocalvirus, Isfahan virus, Maraba virus, Piryvirus, Vesicular stomatitis Alagoas virus, Vesicular stomatitis virus (VSV), BeAn 157575 virus, Boteke virus, Calchaqui virus, Eel virus American, Gray Lodge virus, Jurona virus, Klamath virus, Kwattavirus, La Joya virus, Malpais Spring virus, Mount Elgon bat virus, Perinet virus, virus, Tupaia virus, Farmington, Bahia Grande virus, Muir Springs virus, Reed Ranch virus, Hart Park virus, Flanders virus, Kamese virus, Mosqueiro virus, Mossurilvirus, Barur virus, Fukuoka virus, Kern Canyonvirus, Nkolbisson virus, Le Dantec virus, Keuraliba virus, Connecticut virus, New Minto virus, Sawgrass virus, Chaco virus, Senamadureira virus,Madureiravirus, Timbo virus, Almpiwar virus, Aruac virus, Bangoran virus, Bimbo virus, Bivens Arm virus, Blue crab virus, Charleville virus, Coastal Plains virus, DakArK 7292virus, Entamoeba virus, Garba virus, Gossas virus, Humpty Doo virus, Joinjakaka virus, Kannamangalam virus, Kolongo virus, Koolpinyahvirus, Kotonkon virus, Landjia virus vims, Manitoba vims, Marco vims, Nasoule vims, Navarro vims, Ngaingan vims, Oak-Vale vims, Obodhiang vims, Oita vims, Ouango vims, Parry Creek vims, Rio Grande ci chlid vims, Sandjimba vims, Sigma vims, Sripur vims, Sweetwater Branch vims, Tibrogargan vims, Xiburema vims, Yata vims, Rhode Island, Adelaide River vims, Berrimah vims, vims, Kimberley vims, or Bovine transient fever virusephemeral fever vims).

[0055] As mentioned above, vesicular stomatitis virus (VSV) is a member of the Rhabdoviridae family. The VSV genome is a single molecule of negative-sense RNA that encodes five major polypeptides: nucleocapsid (N) polypeptide, phosphoprotein (P) polypeptide, matrix (M) polypeptide, glycoprotein (G) polypeptide, and viral polymerase (L) polypeptide.

[0056] In some embodiments, the oncolytic virus is a wild-type or recombinant VSV. In some embodiments, the recombinant VSV comprises one or more mutations, such as an M51R substitution (also referred to herein as VSV-M51R).

[0057] In some embodiments, the oncolytic virus may be modified to express one or more cytokines, such as interferon-β (IFNb). In some embodiments, IFNb (e.g., interferon β-1a) may be human or mouse IFNb or a variant thereof. In some embodiments, IFNb comprises an amino acid sequence having at least 90% (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity with the amino acid sequence of SEQ ID NO: 23 or SEQ ID NO: 24, or comprises an amino acid sequence of SEQ ID NO: 23 or SEQ ID NO: 24. In some embodiments, the nucleic acid sequence encoding IFNb is located between the M and G viral genes. Such a position allows the virus to express such an amount of IFNb polypeptide: the amount can effectively activate the antiviral immune response in non-cancerous tissues, and thereby reduce potential viral toxicity without hindering effective viral replication in cancer cells.

[0058] In some embodiments, the recombinant VSV also expresses a sodium / iodine symporter (NIS) or a variant thereof. In some embodiments, NIS comprises an amino acid sequence having at least 90% (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to the amino acid sequence of SEQ ID NO: 25, or comprises the amino acid sequence of SEQ ID NO: 25. In some embodiments, the nucleic acid sequence encoding NIS is located between the G and L viral genes, which allows for appropriate expression levels of the NIS polypeptide.

[0059] In certain embodiments, the oncolytic virus is a recombinant VSV known in the art, such as Voyager V1 described, for example, in US 9428736, which is hereby incorporated by reference in its entirety.

[0060] PD-1 pathway inhibitors

[0061] In some embodiments, the methods disclosed herein include administering a therapeutically effective amount of a PD-1 pathway inhibitor. As used herein, "PD-1 pathway inhibitor" refers to any molecule that can inhibit, block, eliminate or interfere with the activity or expression of PD-1. In some embodiments, the PD-1 pathway inhibitor can be an antibody, a small molecule compound, a nucleic acid, a polypeptide, or a functional fragment or variant thereof. Some non-limiting examples of suitable PD-1 pathway inhibitors include anti-PD-1 antibodies and antigen-binding fragments thereof, anti-PD-L1 antibodies and antigen-binding fragments thereof, and anti-PD-L2 antibodies and antigen-binding fragments thereof.

[0062] Other non-limiting examples of suitable PD-1 pathway inhibitors include RNAi molecules such as anti-PD-1 RNAi molecules, anti-PD-L1 RNAi and anti-PD-L2 RNAi, antisense molecules such as anti-PD-1 antisense RNA, anti-PD-L1 antisense RNA and anti-PD-L2 antisense RNA, and dominant negative proteins such as dominant negative PD-1 protein, dominant negative PD-L1 protein and dominant negative PD-L2 protein. Some examples of the aforementioned PD-1 pathway inhibitors are described in, for example, US 9308236, US10011656 and US20170290808, the parts of which identifying PD-1 pathway inhibitors are incorporated herein by reference.

[0063] The term "antibody" as used herein is intended to refer to an immunoglobulin molecule (i.e., a "complete antibody molecule") composed of four polypeptide chains (two heavy (H) chains and two light (L) chains interconnected by disulfide bonds), as well as multimers thereof (e.g., IgM) or antigen-binding fragments thereof. Each heavy chain comprises a heavy chain variable region ("HCVR" or "VH") and a heavy chain constant region (composed of domains CH1, CH2, and CH3). Each light chain comprises a light chain variable region ("LCVR" or "VL") and a light chain constant region (light chain constant region, CL). The VH and VL regions can be further subdivided into hypervariable regions (called complementarity determining regions (CDRs)), interspersed with more conserved regions (called framework regions (FRs)). Each VH and VL consists of three CDRs and four FRs arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. In some embodiments, the FR of an antibody (or its antigen-binding fragment) may be identical to a human germline sequence, or may be natural or artificially modified. The amino acid consensus sequence may be defined based on a side-by-side analysis of two or more CDRs. The term "antibody" as used herein also includes an antigen-binding fragment of a complete antibody molecule.

[0064] As used herein, the terms "antigen binding fragment" of an antibody, "antigen binding portion" of an antibody, and the like include any naturally occurring, enzymatically obtainable, synthetic, or genetically engineered polypeptide or glycoprotein that specifically binds to an antigen to form a complex. Antigen binding fragments of an antibody can be derived from, for example, a complete antibody molecule using any suitable standard techniques involving manipulation and expression of DNA encoding antibody variable domains and optional constant domains, such as proteolytic digestion or recombinant genetic engineering techniques. Such DNA is known and / or can be readily obtained from, for example, commercial sources, DNA libraries (including, for example, phage-antibody libraries), or can be synthesized. The DNA can be sequenced and manipulated chemically or by using molecular biology techniques, for example, to arrange one or more variable and / or constant domains into a suitable configuration, or to introduce codons, generate cysteine ​​residues, modify, add or delete amino acids, and the like.

[0065] Some non-limiting examples of antigen-binding fragments include: (i) Fab fragments; (ii) F(ab')2 fragments; (iii) Fd fragments; (iv) Fv fragments; (v) single-chain Fv (scFv) molecules; (vi) dAb fragments; and (vii) the smallest recognition unit consisting of amino acid residues that mimic the hypervariable region of an antibody (e.g., an isolated complementary determining region (CDR), such as a CDR3 peptide), or a constrained FR3-CDR3-FR4 peptide. Other engineered molecules such as domain-specific antibodies, single domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetrabodies, minibodies, nanobodies (e.g., monovalent nanobodies, bivalent nanobodies, etc.), small modular immunopharmaceuticals (SMIPs) and shark variable IgNAR domains are also encompassed in the expression "antigen-binding fragment" used herein.

[0066] The antigen-binding fragment of an antibody generally comprises at least one variable domain. A variable domain may have any size or amino acid composition and generally comprises at least one CDR that is adjacent to or in frame with one or more framework sequences. L Domain-associated V H In the antigen-binding fragment of the structural domain, V H and V L The domains may be positioned relative to each other in any suitable arrangement. For example, the variable region may be a dimer and include V H -V H 、V H -V L or V L -V L Alternatively, the antigen-binding fragment of an antibody may comprise a monomer V H or V L Structural domain.

[0067] In some embodiments, the antigen-binding fragment of an antibody may comprise at least one variable domain covalently linked to at least one constant domain. Non-limiting exemplary configurations of variable and constant domains that may be present in the antigen-binding fragment of an antibody of the present disclosure include:

[0068] (i)V H -C H 1; (ii) V H -C H 2; (iii) V H -C H 3; (iv) V H -C H1-C H 2; (v) V H -C H 1-C H 2-C H 3; (vi) V H -C H 2-C H 3; (vii) V H -C L ; (viii) V L -C H 1; (ix) V L -C H 2; (x) V L -C H 3; (xi) V L -C H 1-C H 2; (xii) V L -C H 1-C H 2-C H 3; (xiii) V L -C H 2-C H 3; and (xiv) V L -C L .

[0069] In any configuration of variable domains and constant domains (including any of the exemplary configurations listed above), the variable domains and constant domains may be directly connected to each other, or may be connected by all or part of a hinge or linker region. The hinge region may consist of at least 2 (e.g., 5, 10, 15, 20, 40, 60 or more) amino acids that cause a flexible or semi-flexible connection between adjacent variable domains and / or constant domains in a single polypeptide molecule. In addition, the antigen-binding fragments of the antibodies of the present disclosure may comprise one or more monomeric V domains that are bound to each other and / or to one or more monomeric V domains. H or V L Homodimers or heterodimers (or other multimers) of any of the variable and constant domain configurations listed above in which the domains are non-covalently associated (e.g., by disulfide bonds).

[0070] The antibody used in the disclosed method herein can be a human antibody. The term "human antibody" used herein refers to an antibody with a variable region and a constant region derived from a human germline immunoglobulin sequence. Nevertheless, the human antibody of the present disclosure may include amino acid residues that are not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutations in vivo), e.g., in CDR, and particularly in CDR3. However, the term "human antibody" used herein is not intended to include antibodies in which a CDR sequence derived from another mammalian species (e.g., mouse) germline has been transplanted to a human framework sequence.

[0071] The antibody used in the method disclosed herein can be a recombinant human antibody. The term "recombinant human antibody" used herein includes all human antibodies prepared, expressed, produced or separated by recombinant means, such as antibodies expressed using recombinant expression vectors transfected into host cells (further described below), antibodies separated from recombinant, combinatorial human antibody libraries (further described below), antibodies separated from animals (e.g., mice) that are transgenic for human immunoglobulin genes (see, e.g., Taylor et al. (1992) Nucl. Acids Res. 20: 6287-6295) or antibodies prepared, expressed, produced or separated by any other means involving splicing human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies have variable and constant regions derived from human germline immunoglobulin sequences. However, in some embodiments, such recombinant human antibodies are subjected to in vitro mutagenesis (or, when using animals transgenic for human Ig sequences, in vivo somatic mutagenesis), and thus the V H and V L The amino acid sequence of the region is such that although it is derived from human germline V H and V L Sequences that are related to these but may not naturally occur in the human antibody germline repertoire in vivo.

[0072] Anti-PD-1 antibodies and antigen-binding fragments thereof

[0073] In some embodiments, the PD-1 pathway inhibitor used in the methods disclosed herein is an antibody or antigen-binding fragment thereof (e.g., an anti-PD-1 antibody) that specifically binds to PD-1. The term "specific binding" and the like means that the antibody or its antigen-binding fragment forms a complex with an antigen that is relatively stable under physiological conditions. Methods for determining whether an antibody specifically binds to an antigen are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. For example, an antibody that "specifically binds" to PD-1 as used in the context of the present disclosure includes an antibody that has the following K DBinding to PD-1 or a portion thereof: less than about 500 nM, less than about 300 nM, less than about 200 nM, less than about 100 nM, less than about 90 nM, less than about 80 nM, less than about 70 nM, less than about 60 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM, less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM, or less than about 0.5 nM, as measured in a surface plasmon resonance assay. However, an isolated antibody that specifically binds to human PD-1 may have cross-reactivity with other antigens, such as PD-1 molecules from other (non-human) species.

[0074] According to certain exemplary embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof comprises a heavy chain variable region (HCVR), a light chain variable region (LCVR), and / or a complementarity determining region (CDR) comprising the amino acid sequence of any anti-PD-1 antibody set forth in US 9987500, which is hereby incorporated by reference in its entirety.

[0075] In certain exemplary embodiments, an anti-PD-1 antibody or antigen-binding fragment thereof useful in the context of the present disclosure comprises a heavy chain complementarity determining region (HCDR) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1, and a light chain complementarity determining region (LCDR) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO: 2.

[0076] According to some embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof comprises three HCDRs (HCDR1, HCDR2, and HCDR3) and three LCDRs (LCDR1, LCDR2, and LCDR3), wherein HCDR 1 comprises the amino acid sequence of SEQ ID NO: 3; HCDR2 comprises the amino acid sequence of SEQ ID NO: 4; HCDR3 comprises the amino acid sequence of SEQ ID NO: 5; LCDR1 comprises the amino acid sequence of SEQ ID NO: 6; LCDR2 comprises the amino acid sequence of SEQ ID NO: 7; and LCDR3 comprises the amino acid sequence of SEQ ID NO: 8.

[0077] In other embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof comprises a HCVR comprising SEQ ID NO: 1 and a LCVR comprising SEQ ID NO: 2. In some embodiments, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 9. In some embodiments, the anti-PD-1 antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 10.

[0078] An exemplary antibody comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 1 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 2 is known as cemiplimab (also known as REGN2810; )'s fully human anti-PD-1 antibody.

[0079] According to certain exemplary embodiments, the methods of the present disclosure include the use of cemipril monoclonal antibody or its bioequivalent. As used herein, the term "bioequivalent" with respect to PD-1 pathway inhibitors refers to such anti-PD-1 antibodies or PD-1 binding proteins or fragments thereof, which are drug equivalents or drug substitutes, and when administered at the same molar dose (single dose or multiple doses) under similar experimental conditions, their absorption rate and / or absorption extent do not show significant differences from the absorption rate and / or absorption extent of the reference antibody (e.g., cemipril monoclonal antibody). In the context of the present disclosure, the term "bioequivalent" includes antigen binding proteins that bind to PD-1 and have no clinically significant differences from cemipril monoclonal antibody in terms of safety, purity and / or efficacy.

[0080] According to some embodiments of the present disclosure, the anti-human PD-1 or antigen-binding fragment thereof comprises a HCVR having at least 90% (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to SEQ ID NO: 1.

[0081] According to some embodiments of the present disclosure, the anti-human PD-1 or antigen-binding fragment thereof comprises a LCVR having (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to SEQ ID NO: 2. Sequence identity can be measured by methods known in the art (e.g., GAP, BESTFIT, and BLAST).

[0082] According to some embodiments of the present disclosure, the anti-human PD-1 or antigen-binding fragment thereof comprises a HCVR comprising an amino acid sequence of SEQ ID NO: 1 with no more than 10 amino acid substitutions. According to some embodiments of the present disclosure, the anti-human PD-1 or antigen-binding fragment thereof comprises a LCVR comprising an amino acid sequence of SEQ ID NO: 2 with no more than 10 amino acid substitutions.

[0083] Variants of any HCVR, LCVR and / or CDR amino acid sequences disclosed herein having one or more conservative amino acid substitutions are also within the scope of the present disclosure. For example, the present disclosure includes the use of anti-PD-L1 antibodies having HCVR, LCVR and / or CDR amino acid sequences having, for example, 10 or fewer, 8 or fewer, 6 or fewer, 4 or fewer, etc., conservative amino acid substitutions relative to any HCVR, LCVR and / or CDR amino acid sequences disclosed herein.

[0084] Other anti-PD-1 antibodies or antigen-binding fragments thereof that can be used in the context of the methods of the present disclosure include, for example, those referred to and known in the art as nivolumab, pembrolizumab, MEDI0608, pidilizumab, BI 754091, spartalizumab (also known as PDR001), camrelizumab (also known as SHR-1210), JNJ-63723283, MCLA-134, or any of the anti-PD-1 antibodies set forth in U.S. Patent Nos. 6,808,710, 7,488,802, 8,008,449, 8,168,757, 8,354,509, 8,609,089, 8,686,119, 8,779,105, 8,900,587, and 9,987,500, and in patent publications WO 2006 / 121168, WO 2009 / 114335. All of the above disclosures identifying anti-PD-1 antibodies are hereby incorporated by reference.

[0085] The anti-PD-1 antibodies used in the context of the methods of the present disclosure may have pH-dependent binding properties. For example, the anti-PD-1 antibodies used in the methods of the present disclosure may exhibit reduced binding to PD-1 at acidic pH compared to neutral pH. Alternatively, the anti-PD-1 antibodies of the present disclosure may exhibit enhanced binding to their antigens at acidic pH compared to neutral pH. The expression "acidic pH" includes pH values ​​less than about 6.2, such as about 6.0, 5.95, 5.9, 5.85, 5.8, 5.75, 5.7, 5.65, 5.6, 5.55, 5.5, 5.45, 5.4, 5.35, 5.3, 5.25, 5.2, 5.15, 5.1, 5.05, 5.0 or less. As used herein, the expression "neutral pH" means a pH of about 7.0 to about 7.4. The expression "neutral pH" includes pH values ​​of about 7.0, 7.05, 7.1, 7.15, 7.2, 7.25, 7.3, 7.35 and 7.4.

[0086] In certain instances, “reduced binding to PD-1 at acidic pH compared to neutral pH” is defined as the K of the antibody that binds to PD-1 at acidic pH. D The K values ​​of antibodies binding to PD-1 at neutral pHD For example, if an antibody or antigen-binding fragment thereof exhibits an acidic / neutral K of about 3.0 or greater, D For the purposes of the present disclosure, an antibody or antigen-binding fragment thereof may be considered to exhibit "reduced binding to PD-1 at acidic pH compared to neutral pH". In certain exemplary embodiments, the acidic / neutral K of an antibody or antigen-binding fragment of the present disclosure is D The ratio may be about 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, 10.0, 10.5, 11.0, 11.5, 12.0, 12.5, 13.0, 13.5, 14.0, 14.5, 15.0, 20.0, 25.0, 30.0, 40.0, 50.0, 60.0, 70.0, 100.0 or more.

[0087] Antibodies with pH-dependent binding characteristics can be obtained, for example, by screening a population of antibodies that have reduced (or enhanced) binding to a specific antigen at an acidic pH compared to a neutral pH. In addition, modifications to the antigen-binding domain at the amino acid level can produce antibodies with pH-dependent characteristics. For example, by replacing one or more amino acids in the antigen-binding domain (e.g., within a CDR) with histidine residues, antibodies with reduced antigen binding at an acidic pH relative to a neutral pH can be obtained. As used herein, the expression "acidic pH" means a pH of 6.0 or less. Anti-PD-L1 antibodies and antigen-binding fragments thereof

[0088] In some embodiments, the PD-1 pathway inhibitor used in the methods disclosed herein is an antibody or antigen-binding fragment thereof that specifically binds to PD-L1 (e.g., an anti-PD-L1 antibody). For example, an antibody that "specifically binds" PD-L1 as used in the context of the present disclosure includes an antibody that specifically binds to PD-L1 at a concentration of about 1×10 -8 M or smaller K D (For example, a smaller K D An antibody that binds to PD-L1 or a portion thereof (expressed as K ) is an antibody that has a binding affinity for PD-L1 (expressed as K ) and has a binding affinity for PD-L1 (expressed as K ). D ) of those mAbs, the K D For at least 10 -8 M (e.g. 10 -9 M, 10 -10 M, 10 -11 M or 10 -12 M), such as by surface plasmon resonance (e.g. BIACORE TMHowever, an isolated antibody that specifically binds to human PD-L1 may have cross-reactivity with other antigens, such as PD-L1 molecules from other (non-human) species.

[0089] According to certain exemplary embodiments, the anti-PD-L1 antibody or antigen-binding fragment thereof comprises a heavy chain variable region (HCVR), a light chain variable region (LCVR), and / or a complementarity determining region (CDR) comprising the amino acid sequence of any anti-PD-L1 antibody set forth in US 9938345, which is hereby incorporated by reference in its entirety.

[0090] In certain exemplary embodiments, an anti-PD-L1 antibody or antigen-binding fragment thereof that can be used in the context of the present disclosure comprises a heavy chain complementarity determining region (HCDR) comprising a heavy chain variable region (HCVR) of SEQ ID NO: 11 and a light chain complementarity determining region (LCDR) comprising a light chain variable region (LCVR) of SEQ ID NO: 12. An exemplary anti-PD-L1 antibody comprising a HCVR of SEQ ID NO: 11 and a LCVR of SEQ ID NO: 12 is REGN3504.

[0091] According to some embodiments of the present disclosure, the anti-human PD-L1 antibody or antigen-binding fragment thereof comprises a HCVR having at least 90% (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to SEQ ID NO: 11. According to some embodiments of the present disclosure, the anti-human PD-L1 antibody or antigen-binding fragment thereof comprises a LCVR having at least 90% (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to SEQ ID NO: 12.

[0092] According to some embodiments of the present disclosure, the anti-human PD-L1 antibody or antigen-binding fragment thereof comprises a HCVR comprising the amino acid sequence of SEQ ID NO: 11 with no more than 10 amino acid substitutions. According to some embodiments of the present disclosure, the anti-human PD-L1 antibody or antigen-binding fragment thereof comprises a LCVR comprising the amino acid sequence of SEQ ID NO: 12 with no more than 10 amino acid substitutions.

[0093] Variants of any HCVR, LCVR and / or CDR amino acid sequences disclosed herein having one or more conservative amino acid substitutions are also within the scope of the present disclosure. For example, the present disclosure includes the use of anti-PD-L1 antibodies having HCVR, LCVR and / or CDR amino acid sequences having, for example, 10 or fewer, 8 or fewer, 6 or fewer, 4 or fewer, etc., conservative amino acid substitutions relative to any HCVR, LCVR and / or CDR amino acid sequences disclosed herein.

[0094] Other anti-PD-L1 antibodies that can be used in the context of the methods of the present disclosure include, for example, antibodies referred to and known in the art as: MDX-1105, atezolizumab (TECENTRIQ TM ), Imfinzi TM ), Avelumab (BAVENCIO TM ), LY3300054, FAZ053, STI-1014, CX-072, KN035 (Zhang et al., Cell Discovery, 3, 170004 (March 2017)), CK-301 (Gorelik et al., American Association for Cancer Research Annual Meeting (AACR), 2016-04-04 Abstract 4606), or in US Patent Nos. 7943743, 8217149, 9402899, 9624298, and 9938345 and patent publications WO 2007 / 005874, WO 2010 / 077634, WO 2013 / 181452, WO 2013 / 181634, WO 2016 / 149201, WO Any other anti-PD-L1 antibody shown in 2017 / 034916 or EP3177649. All of the above disclosures identifying anti-PD-L1 antibodies are hereby incorporated by reference.

[0095] Anti-PD-L2 antibodies and antigen-binding fragments thereof

[0096] In some embodiments, the PD-1 pathway inhibitor used in the methods disclosed herein is an antibody or antigen-binding fragment thereof that specifically binds to PD-L2 (e.g., an anti-PD-L2 antibody). For example, an antibody that "specifically binds" to PD-L2 as used in the context of the present disclosure includes an antibody that specifically binds to PD-L2 at a concentration of about 1×10 -8An antibody that binds to PD-L2 or a portion thereof with a KD of at least 10 M or less (e.g., a smaller KD indicates tighter binding). A "high affinity" anti-PD-L2 antibody refers to those mAbs that have a binding affinity (expressed as a KD) for PD-L2 that is at least 10 -8 M (e.g. 10 -9 M, 10 -10 M, 10 -11 M or 10 -12 M), as measured by surface plasmon resonance (e.g., BIACORE™ or solution affinity ELISA). However, an isolated antibody that specifically binds to human PD-L2 may have cross-reactivity with other antigens, such as PD-L2 molecules from other (non-human) species.

[0097] Anti-PD-L2 antibodies that can be used in the context of the methods of the present disclosure include, for example, those set forth in US Patent Nos. 8,552,154 and 10,647,771. All of the above disclosures identifying anti-PD-L2 antibodies are hereby incorporated by reference.

[0098] CTLA-4 inhibitors

[0099] In some embodiments, the methods disclosed herein include administering a therapeutically effective amount of a CTLA-4 inhibitor. As used herein, "CTLA-4 inhibitor" refers to any molecule that can inhibit, block, eliminate or interfere with the activity or expression of CTLA-4. In some embodiments, the CTLA-4 inhibitor can be an antibody, a small molecule compound, a nucleic acid, a polypeptide, or a functional fragment or variant thereof. Some non-limiting examples of suitable CTLA-4 inhibitors include anti-CTLA-4 antibodies and antigen-binding fragments thereof. Other non-limiting examples of suitable CTLA-4 inhibitors include RNAi molecules (e.g., anti-CTLA-4 RNAi molecules) and dominant negative proteins (e.g., dominant negative CTLA-4 proteins).

[0100] Anti-CTLA-4 antibodies and antigen-binding fragments thereof

[0101] In some embodiments, the CTLA-4 inhibitor used in the methods disclosed herein is an antibody or antigen-binding fragment thereof that specifically binds to CTLA-4 (e.g., an anti-CTLA-4 antibody). The term "specific binding" and the like means that the antibody or its antigen-binding fragment forms a complex with an antigen that is relatively stable under physiological conditions. Methods for determining whether an antibody specifically binds to an antigen are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. For example, antibodies that "specifically bind" to CTLA-4 as used in the context of the present disclosure include antibodies that have the following K DBinding to CTLA-4 antibody or portion thereof: less than about 500 nM, less than about 300 nM, less than about 200 nM, less than about 100 nM, less than about 90 nM, less than about 80 nM, less than about 70 nM, less than about 60 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM, less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM, or less than about 0.5 nM as measured in a surface plasmon resonance assay. However, an isolated antibody that specifically binds human CTLA-4 may have cross-reactivity with other antigens, such as CTLA-4 molecules from other (non-human) species.

[0102] In certain exemplary embodiments, an anti-CTLA-4 antibody or antigen-binding fragment thereof that can be used in the context of the present disclosure comprises a heavy chain complementarity determining region (HCDR) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO:13, and a light chain complementarity determining region (LCDR) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO:14.

[0103] According to some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment thereof comprises three HCDRs (HCDR1, HCDR2, and HCDR3) and three LCDRs (LCDR1, LCDR2, and LCDR3), wherein HCDR1 comprises the amino acid sequence of SEQ ID NO: 15; HCDR2 comprises the amino acid sequence of SEQ ID NO: 16; HCDR3 comprises the amino acid sequence of SEQ ID NO: 17; LCDR1 comprises the amino acid sequence of SEQ ID NO: 18; LCDR2 comprises the amino acid sequence of SEQ ID NO: 19; and LCDR3 comprises the amino acid sequence of SEQ ID NO: 20.

[0104] In other embodiments, the anti-CTLA-4 antibody or antigen-binding fragment thereof comprises a HCVR comprising the amino acid sequence of SEQ ID NO: 13 and a LCVR comprising the amino acid sequence of SEQ ID NO: 14. In some embodiments, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 21. In some embodiments, the anti-CTLA-4 antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 22.

[0105] An exemplary antibody comprising a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 13 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 14 is the fully human anti-CTLA-4 antibody designated REGN4659.

[0106] According to certain exemplary embodiments, the methods of the present disclosure include the use of REGN4659 or a bioequivalent thereof. As used herein, the term "bioequivalent" with respect to CTLA-4 inhibitors refers to an anti-CTLA-4 antibody or CTLA-4 binding protein or fragment thereof that is a pharmaceutical equivalent or pharmaceutical substitute, and whose absorption rate and / or extent of absorption do not show significant differences from the absorption rate and / or extent of absorption of a reference antibody (e.g., REGN4659) when administered at the same molar dose (single or multiple doses) under similar experimental conditions. In the context of the present disclosure, the term "bioequivalent" includes antigen binding proteins that bind to CTLA-4 and have no clinically significant differences from REGN4659 in terms of safety, purity, and / or potency.

[0107] According to some embodiments of the present disclosure, the anti-human CTLA-4 or antigen-binding fragment thereof comprises a HCVR having at least 90% (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to the amino acid sequence of SEQ ID NO:13.

[0108] According to some embodiments of the present disclosure, the anti-human CTLA-4 or antigen-binding fragment thereof comprises a LCVR having (e.g., 90%, 95%, 96%, 97%, 98%, 99%) sequence identity to the amino acid sequence of SEQ ID NO:14.

[0109] According to some embodiments of the present disclosure, the anti-human CTLA-4 or antigen-binding fragment thereof comprises a HCVR comprising the amino acid sequence of SEQ ID NO: 13 with no more than 10 amino acid substitutions. According to some embodiments of the present disclosure, the anti-human CTLA-4 or antigen-binding fragment thereof comprises a LCVR comprising the amino acid sequence of SEQ ID NO: 14 with no more than 10 amino acid substitutions.

[0110] Variants of any HCVR, LCVR and / or CDR amino acid sequences disclosed herein having one or more conservative amino acid substitutions are also within the scope of the present disclosure. For example, the present disclosure includes the use of anti-PD-L1 antibodies having HCVR, LCVR and / or CDR amino acid sequences having, for example, 10 or fewer, 8 or fewer, 6 or fewer, 4 or fewer, etc., conservative amino acid substitutions relative to any HCVR, LCVR and / or CDR amino acid sequences disclosed herein.

[0111] Other anti-CTLA-4 antibodies or antigen-binding fragments thereof that can be used in the context of the methods of the present disclosure include, for example, antibodies referred to and known in the art as ipilimumab, tremelimumab, or any of the anti-CTLA-4 antibodies set forth in US Patent Nos. 7,527,969, 8,779,098, 7,666,424, 7,737,258, 7,740,845, 8,148,154, 8,414,892, 8,501,471, and 9,062,110, and patent publications US 2013 / 0078234, US 2010 / 0143245, WO 2017062615A2, WO 2004 / 001381, and WO 2012 / 147713. All of the above disclosures identifying anti-CTLA-4 antibodies are hereby incorporated by reference.

[0112] Pharmaceutical compositions and administration

[0113] The present disclosure includes methods comprising administering oncolytic viruses, PD-1 pathway inhibitors and / or CTLA-4 inhibitors to a subject, wherein the antibody is contained in a separate or combined (single) pharmaceutical composition. The pharmaceutical composition of the present disclosure can be formulated with suitable carriers, excipients and other agents providing suitable transfer, delivery, tolerance, etc. A variety of suitable preparations can be found in formularies known to all pharmaceutical chemists: Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa. These preparations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, vesicles (e.g., LIPOFECTIN) containing lipids (cationic or anionic), DNA conjugates, anhydrous absorption pastes, oil-in-water and water-in-oil emulsions, emulsions carbon wax (polyethylene glycol of various molecular weights), semisolid gels and semisolid mixtures containing carbon wax. See also Powell et al. PDA (1998) J Pharm Sci Technol 52: 238-311.

[0114] A variety of delivery systems are known and can be used to administer the pharmaceutical compositions of the present disclosure, e.g., encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing mutant viruses, receptor-mediated endocytosis (see, e.g., Wu et al. (1987) J. Biol. Chem. 262:4429-4432). Methods of administration include, but are not limited to, intradermal, intramuscular, intratumoral, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The composition can be administered by any convenient route, e.g., by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.), and can be administered with other biologically active agents.

[0115] The pharmaceutical composition comprising an oncolytic virus, a PD-1 pathway inhibitor or a CTLA-4 inhibitor can be delivered intratumorally, subcutaneously or intravenously with a standard needle and syringe. In some embodiments, one or more of an oncolytic virus, a PD-1 pathway inhibitor and / or a CTLA-4 inhibitor can be delivered by more than one route of administration, such as one or more doses administered intratumorally and one or more doses administered intravenously, or delivered by some other route of administration. In addition, with respect to subcutaneous delivery, a pen delivery device is easily applicable to the delivery of the pharmaceutical composition of the present disclosure. Such a pen delivery device can be reusable or disposable. A reusable pen delivery device typically uses a replaceable cartridge containing a pharmaceutical composition. Once all the pharmaceutical compositions in the cartridge have been applied and the cartridge is empty, the empty cartridge can be easily discarded and replaced with a new cartridge containing the pharmaceutical composition. The pen delivery device can then be reused. In a disposable pen delivery device, there is no replaceable cartridge. In contrast, a disposable pen delivery device is pre-filled with a pharmaceutical composition contained in a reservoir in the device. Once the reservoir has no pharmaceutical composition, the entire device is discarded.

[0116] In some cases, the pharmaceutical composition can be delivered in a controlled release system. In one embodiment, a pump can be used. In another embodiment, a polymeric material can be used; see, for example, Medical Applications of Controlled Release, Langer and Wise (eds.), 1974, CRC Pres., Boca Raton, Fla. In another embodiment, the controlled release system can be placed near the target of the composition, so only a portion of the systemic dose is required (see, for example, Goodson, 1984, in Medical Applications of Controlled Release (supra), Vol. 2, pp. 115 to 138). Other controlled release systems are discussed in the review of Langer, 1990, Science 249: 1527-1533.

[0117] Injectable preparations can include dosage forms for intravenous, subcutaneous, intradermal and intramuscular injections, instillations, etc. These injectable preparations can be prepared by known methods. For example, injectable preparations can be prepared by, for example, dissolving, suspending or emulsifying the above-mentioned antibody or its salt in a conventional sterile aqueous medium or an oily medium for injection. As an aqueous medium for injection, there are, for example, physiological saline, isotonic solutions containing glucose and other adjuvants, etc., which can be used in combination with suitable solubilizers such as alcohol (such as ethanol), polyols (such as propylene glycol, polyethylene glycol), nonionic surfactants [such as polysorbate 80, HCO-50 (polyoxyethylene (50mol) adducts of hydrogenated castor oil)], etc. As an oily medium, use, for example, sesame oil, soybean oil, etc., which can be used in combination with solubilizers (such as benzyl benzoate, benzyl alcohol, etc.). The injection thus prepared is preferably filled in a suitable ampoule.

[0118] Advantageously, the pharmaceutical composition for oral or parenteral use is prepared into a unit dosage form suitable for the dosage of the active ingredient. Such a unit dosage form includes, for example, tablets, pills, capsules, injections (ampoules), suppositories, etc.

[0119] The present disclosure also provides a kit comprising an oncolytic virus and a PD-1 pathway inhibitor in combination with: written instructions for using a therapeutically effective amount of the combination of the oncolytic virus and the PD-1 pathway inhibitor to treat a patient's tumor or inhibit the growth of a patient's tumor.

[0120] The present disclosure also provides a kit comprising an oncolytic virus and a CTLA-4 inhibitor in combination with written instructions for using a therapeutically effective amount of the combination of the oncolytic virus and the CTLA-4 inhibitor to treat a patient's tumor or inhibit the growth of a patient's tumor.

[0121] The present disclosure also provides a kit comprising an oncolytic virus, a PD-1 pathway inhibitor, and a CTLA-4 inhibitor in combination with: written instructions for using a therapeutically effective amount of the combination of the oncolytic virus, the PD-1 pathway inhibitor, and the CTLA-4 inhibitor to treat a patient's tumor or inhibit the growth of a patient's tumor.

[0122] Administration regimen

[0123] The method of the present disclosure may include administering an oncolytic virus, a PD-1 pathway inhibitor (e.g., an anti-PD-1, anti-PD-L1, or anti-PD-L2 antibody, or an antigen-binding fragment thereof), or a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or an antigen-binding fragment thereof) to a subject at the following dosing frequency: about four times a week, twice a week, once a week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, once every six weeks, once every eight weeks, once every twelve weeks, or less frequently, as long as a therapeutic response is achieved. The method of the present disclosure may also include administering a single dose of an oncolytic virus, a PD-1 pathway inhibitor, or a CTLA-4 inhibitor.

[0124] In some embodiments, at least one of an oncolytic virus, a PD-1 pathway inhibitor, or a CTLA-4 inhibitor is administered to the patient once a day, once every two days, once every three days, once every four days, once every five days, once a week, once every two weeks, or once every three weeks.

[0125] In some embodiments, an oncolytic virus, a PD-1 pathway inhibitor, and a CTLA-4 inhibitor are administered to a patient simultaneously.

[0126] In some embodiments, the method may include sequentially administering two or more of an oncolytic virus, a PD-1 pathway inhibitor, and a CTLA-4 inhibitor to a subject. In some embodiments, an oncolytic virus is administered to a patient before or after a PD-1 pathway inhibitor and a CTLA-4 inhibitor. In some embodiments, a PD-1 pathway inhibitor is administered to a patient before or after an oncolytic virus and a CTLA-4 inhibitor. In some embodiments, a CTLA-4 inhibitor is administered to a patient before or after an oncolytic virus and a PD-1 pathway inhibitor.

[0127] In some embodiments, the method may include sequentially administering an oncolytic virus and a PD-1 pathway inhibitor (or CTLA-4 inhibitor) to a subject. In some embodiments, an oncolytic virus is administered to a patient before or after the PD-1 pathway inhibitor. In some embodiments, a PD-1 pathway inhibitor is administered to a patient before or after oncolysis.

[0128] As used herein, "sequential administration" means that each dose of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor is administered to a subject at different time points (e.g., on different days) at predetermined intervals (e.g., hours, days, weeks or months). The present disclosure includes such a method, which includes sequentially administering a single initial dose of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor to a patient, followed by one or more second doses of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor, and optionally followed by one or more third doses of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor. In some embodiments, the method also includes sequentially administering a single initial dose of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor to a patient, followed by one or more second doses of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor, and optionally followed by one or more third doses of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor.

[0129] The terms "initial dose", "second dose" and "tertiary dose" refer to the time sequence of administration. Therefore, the "initial dose" is the dose administered at the beginning of the treatment regimen (also referred to as the "baseline dose"); the "second dose" is the dose administered after the initial dose; and the "third dose" is the dose administered after the second dose. The initial, second and third doses may all contain the same amount of oncolytic virus, PD-1 pathway inhibitor or CTLA-4 inhibitor. However, in some embodiments, during the course of treatment, the amounts contained in the initial, second and / or third doses are different from each other (e.g., adjusted upward or downward as appropriate). In some embodiments, one or more (e.g., 1, 2, 3, 4 or 5) doses are administered as a "loading dose" at the beginning of the treatment regimen, followed by subsequent doses (e.g., "maintenance doses") based on less frequent administration.

[0130] In an exemplary embodiment of the present disclosure, each second and / or third dose is 1 / 2 to 14 weeks (e.g., 1 / 2, 1, 1 1 / 2,2,2 1 / 2,3,3 1 / 2,4,4 1 / 2,5,5 1 / 2,6,6 1 / 2,7,7 1 / 2,8,8 1 / 2,9,9 1 / 2, 10, 10 1 / 2, 11, 11 1 / 2, 12, 121 / 2, 13, 13 1 / 2, 14, 14 1 / 2 or more weeks) administration. As used herein, the phrase "immediately preceding dose" means the dose of an oncolytic virus, PD-1 pathway inhibitor, or CTLA-4 inhibitor administered to a patient prior to the administration of the next dose in the order of multiple administrations without an intervening dose.

[0131] In some embodiments, the method may include administering to the patient any number of second and / or third doses of oncolytic virus, PD-1 pathway inhibitor (e.g., anti-PD-1 antibody or antigen binding fragment thereof) and / or CTLA-4 inhibitor (e.g., anti-CTLA-4 antibody or antigen binding fragment thereof). For example, in some embodiments, only a single second dose is administered to the patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7, 8 or more) second doses are administered to the patient. Similarly, in some embodiments, only a single third dose is administered to the patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7, 8 or more) third doses are administered to the patient.

[0132] In embodiments involving multiple second doses, each second dose can be administered at the same frequency as other second doses. For example, each second dose can be administered to the patient 1 to 2 weeks immediately after the previous dose. Similarly, in embodiments involving multiple third doses, each third dose can be administered to the patient at the same frequency as other third doses. For example, each third dose can be administered to the patient 2 to 4 weeks immediately after the previous dose. Alternatively, the frequency at which the second and / or third dose is administered to the patient can vary during the course of the treatment regimen. During the course of treatment, the physician can also adjust the frequency of administration according to the needs of individual patients after clinical examination.

[0133] In some embodiments, one or more doses of oncolytic virus, PD-1 pathway inhibitor and / or CTLA-4 inhibitor are administered in one or more cycles. "Initial treatment cycle" is a treatment cycle administered at the beginning of the treatment regimen; "Second treatment cycle" refers to a treatment cycle administered after the initial treatment cycle; and "Third treatment cycle" is a treatment cycle administered after the second treatment cycle. In the context of the present disclosure, the treatment cycles may be the same or different from each other. In some embodiments, the duration of any of the aforementioned treatment cycles may be 1 week (7 days), 2 weeks (14 days), 3 weeks (21 days), 4 weeks (28 days), 5 weeks (35 days), 6 weeks (42 days), 7 weeks (49 days), 8 weeks (56 days) or longer.

[0134] In some embodiments, one or more doses of an oncolytic virus, PD-1 pathway inhibitor, and / or CTLA-4 inhibitor are administered at the beginning of the treatment regimen as an "induction dose" based on a higher frequency (twice a week, once a week, or once every 2 weeks), followed by subsequent doses ("consolidation doses" or "maintenance doses") administered at a lower frequency (e.g., once every 4 to 12 weeks).

[0135] In some embodiments, the combined administration regimen includes simultaneous administration of an oncolytic virus (e.g., a recombinant VSV, such as VV1) and an optional CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or an antigen-binding fragment thereof, such as ipilimumab or a bioequivalent thereof), and subsequent administration of a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or an antigen-binding fragment thereof, such as cemiplimab or a bioequivalent thereof). For example, the combined administration regimen may include administration of an oncolytic virus and an optional CTLA-4 inhibitor on day 1, and subsequent administration of a PD-1 pathway inhibitor 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 days later.

[0136] In some embodiments, the administration regimen of the combination includes one or more of the following: intratumoral administration of an oncolytic virus (e.g., a recombinant VSV, such as VV1), intravenous administration of an oncolytic virus (e.g., a recombinant VSV, such as VV1), intravenous administration of a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or an antigen-binding fragment thereof, such as ipilimumab or a bioequivalent thereof), and intravenous administration of a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or an antigen-binding fragment thereof, such as cemiplimab or a bioequivalent thereof).

[0137] For example, the administration regimen may include intratumoral administration of a therapeutically effective amount of an oncolytic virus on the first day of a 28-day initial treatment cycle, intravenous administration of a therapeutically effective amount of an oncolytic virus on the first day of the initial treatment cycle, intravenous administration of a therapeutically effective amount of a CTLA-4 inhibitor on the first day of the initial treatment cycle, intravenous administration of a therapeutically effective amount of a PD-1 pathway inhibitor on the eighth day of the initial treatment cycle, and intravenous administration of a therapeutically effective amount of a PD-1 pathway inhibitor on the first day of the second and each subsequent treatment cycle.

[0138] In another example, the administration regimen may include intravenous administration of a therapeutically effective amount of a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or an antigen-binding fragment thereof, such as cemiplimab or a bioequivalent thereof) on the first day of a 21-day initial treatment cycle and intratumoral administration of a therapeutically effective amount of an oncolytic virus on the first day of the initial treatment cycle.

[0139] dose

[0140] The amount of oncolytic virus, PD-1 pathway inhibitor (e.g., anti-PD-1 antibody or antigen-binding fragment thereof), or CTLA-4 inhibitor (e.g., anti-CTLA-4 antibody or antigen-binding fragment thereof) administered to a subject according to the methods disclosed herein is generally a therapeutically effective amount. As used herein, the term "therapeutically effective amount" means an amount of an oncolytic virus, a PD-1 pathway inhibitor, and / or a CTLA-4 inhibitor that results in one or more of the following: (a) reducing the severity or duration of a symptom or indication of cancer (e.g., tumor lesions); (b) inhibiting tumor growth, or increasing tumor necrosis, tumor shrinkage, and / or tumor disappearance; (c) delaying tumor growth and occurrence; (d) inhibiting tumor metastasis; (e) preventing tumor growth recurrence; (f) improving the survival of subjects with cancer; and / or (g) reducing the use or need for conventional anti-cancer treatments (e.g., eliminating the need for surgery or reducing or eliminating the use of chemotherapeutic or cytotoxic agents), compared to untreated subjects, subjects treated with a single therapy, or subjects treated with any two of the three therapeutic agents disclosed herein (PD-1 pathway inhibitor, CTLA-4 inhibitor, and oncolytic virus).

[0141] In some embodiments, the combined oncolytic virus can be used as one or more of 10, 100, 10 3 , 10 4 , 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , 10 11 , 10 12 , 10 13 , 10 14 Or more viral particles (viral particles, vp) or plaque forming units (plaque-forming unit, pfu) unit dose administration. In some embodiments, the oncolytic virus is an oncolytic rhabdovirus (e.g., wild-type or genetically modified VSV) and is administered as one or more 10 4 Up to 10 14 pfu, 10 6 Up to 10 14 pfu, 10 8 Up to 10 14 pfu, 10 4 Up to 10 12 pfu, 10 6 Up to 10 12 pfu, 10 8 Up to 10 12 pfu, 10 9 Up to 1011 pfu or 10 10 Up to 10 12 pfu, or any range therebetween, is administered to a human suffering from cancer.

[0142] In some embodiments, the combined oncolytic virus can be used as one or more of 10, 100, 10 3 , 10 4 , 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , 10 11 , 10 12 , 10 13 , 10 14 or more 50% tissue culture infectious dose (TCID 50 ) in a unit dose. In some embodiments, the oncolytic virus is an oncolytic rhabdovirus (e.g., wild-type or genetically modified VSV) and is administered in one or more 10 4 Up to 10 14 TCID 50 , 10 6 Up to 10 14 TCID 50 , 10 8 Up to 10 14 TCID 50 , 10 4 Up to 10 12 TCID 50 , 10 6 Up to 10 12 TCID 50 or 10 8 Up to 10 12 TCID 50 , 10 9 Up to 10 11 TCID 50 or 10 10 Up to 10 12 TCID 50 , or any range therebetween, is administered to a human with cancer. In some embodiments, the oncolytic virus is a recombinant VSV, such as VV1, and is administered in one or more 10 4 Up to 10 14 TCID 50 , 10 6 Up to 10 14 TCID 50 , 10 8 Up to 10 14 TCID 50, 10 4 Up to 10 12 TCID 50 , 10 6 Up to 10 12 TCID 50 , 10 8 Up to 10 12 TCID 50 , 10 9 Up to 10 11 TCID 50 , 10 10 Up to 10 12 TCID 50 , or any range therebetween is administered to a person suffering from cancer. In some embodiments, the therapeutically effective amount of an oncolytic virus (e.g., a recombinant VSV, such as VV1) can be 10, 100, 10 3 , 10 4 , 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , 10 11 , 10 12 , 10 13 , 10 14 or more TCID 50 .

[0143] In some embodiments, a therapeutically effective amount of a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or antigen-binding fragment thereof, such as cemiplimab or a bioequivalent thereof) can be about 0.05 mg to about 1500 mg, about 1 mg to about 800 mg, about 5 mg to about 600 mg, about 10 mg to about 550 mg, about 50 mg to about 400 mg, about 75 mg to about 350 mg, or about 100 mg to about 300 mg of the antibody. For example, in various embodiments, the amount of the PD-1 pathway inhibitor is about 0.05 mg, about 0.1 mg, about 1.0 mg, about 1.5 mg, about 2.0 mg, about 5 mg, about 10 mg, about 15 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 110 mg, about 120 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg, about 180 mg, about 190 mg, about 200 mg, about 210 mg. , about 220mg, about 230mg, about 240mg, about 250mg, about 260mg, about 270mg, about 280mg, about 290mg, about 300mg, about 310mg, about 320mg, about 330mg, about 340mg, about 350mg, about 360mg, about 370mg, about 380mg, about 390mg, about 400mg, about 410mg, about 420mg, about 430mg, about 440mg, about 450mg, about 460mg, about 470mg, about 480mg, about 490mg, about 500mg, about 510mg g, about 520mg, about 530mg, about 540mg, about 550mg, about 560mg, about 570mg, about 580mg, about 590mg, about 600mg, about 610mg, about 620mg, about 630mg, about 640mg, about 650mg, about 660mg, about 670mg, about 680mg, about 690mg, about 700mg, about 710mg, about 720mg, about 730mg, about 740mg, about 750mg, about 760mg, about 770mg, about 780mg, about 790mg, about 800mg, about 810 mg, about 820mg, about 830mg, about 840mg, about 850mg, about 860mg, about 870mg, about 880mg, about 890mg, about 900mg, about 910mg, about 920mg, about 930mg, about 940mg, about 950mg, about 960mg, about 970mg, about 980mg, about 990mg, about 1000mg, about 1010mg, about 1020mg, about 1030mg, about 1040mg, about 1050mg, about 1060mg, about 1070mg, about 1080mg, about 1090mg,about 1200 mg, about 1210 mg, about 1220 mg, about 1230 mg, about 1240 mg, about 1250 mg, about 1260 mg, about 1270 mg, about 1280 mg, about 1290 mg, about 1300 mg, about 1310 mg, about 1320 mg, about 1330 mg, about 1340 mg, about 1350 mg, about 1360 mg, about 1370 mg, about 1380 mg, about 1390 mg, about 1400 mg, about 1410 mg, about 1420 mg, about 1430 mg, about 1440 mg, about 1450 mg, about 1460 mg, about 1470 mg, about 1480 mg, about 1490 mg, or about 1500 mg.

[0144] The amount of PD-1 pathway inhibitor (e.g., anti-PD-1 antibody or its antigen-binding fragment) contained in a single dose can be expressed in terms of mg of antibody per kilogram of subject body weight (i.e., mg / kg). In some embodiments, the PD-1 pathway inhibitor used in the methods disclosed herein can be administered to the subject at a dose of about 0.0001 to about 100 mg / kg of subject body weight. In some embodiments, the anti-PD-1 antibody can be administered at a dose of about 0.1 mg / kg to about 20 mg / kg of patient body weight. In some embodiments, the methods of the present disclosure include administering a PD-1 pathway inhibitor (e.g., anti-PD-1 antibody or its antigen-binding fragment) at a dose of about 1 mg / kg to 3 mg / kg, 1 mg / kg to 5 mg / kg, 1 mg / kg to 10 mg / kg, 3 mg / kg to 10 mg / kg, 5 mg / kg to 10 mg / kg, 1 mg / kg, 3 mg / kg, 5 mg / kg or 10 mg / kg of patient body weight.

[0145] In some embodiments, each dose of the PD-1 pathway inhibitor comprises 0.1 to 10 mg / kg (e.g., 0.3 mg / kg, 1 mg / kg, 3 mg / kg, or 10 mg / kg) of subject body weight. In certain additional embodiments, each dose contains 5 to 1500 mg of a PD-1 pathway inhibitor (e.g., an anti-PD-1 antibody or antigen-binding fragment thereof), for example, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 40 mg, 45 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg, 850 mg, 900 mg, 950 mg, 1000 mg, 1050 mg, 1100 mg, 1150 mg, 1200 mg, 1550 mg, 1300 mg, 1350 mg, 1400 mg, 1450 mg, or 1500 mg of a PD-1 pathway inhibitor.

[0146] In some embodiments, a therapeutically effective amount of a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or antigen-binding fragment thereof, such as ipilimumab or a bioequivalent thereof) can be about 0.05 mg to about 1000 mg, about 1 mg to about 800 mg, about 5 mg to about 600 mg, about 10 mg to about 550 mg, about 50 mg to about 400 mg, about 75 mg to about 350 mg, or about 100 mg to about 300 mg of the antibody. For example, in various embodiments, the amount of the CTLA-4 inhibitor is about 0.05 mg, about 0.1 mg, about 1.0 mg, about 1.5 mg, about 2.0 mg, about 5 mg, about 10 mg, about 15 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 110 mg, about 120 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg, about 180 mg, about 190 mg, about 200 mg, about 210 mg, about 220 mg, about 230 mg, about 240 mg, about 250 mg, about 260 mg, about 270 mg, about 280 mg, about 290 mg, about 300 mg, about 310 mg, about 320 mg, about 330 mg, about 340 mg, about 350 mg, about 360 mg, about 370 mg, about 380 mg, about 390 mg, about 400 mg, about 410 mg, about 420 mg, about 430 mg, about 440 mg, about 450 mg, about 460 mg, about 470 mg, about 480 mg, about 490 mg, about 500 mg, about 510 mg, about 520 mg, about 530 mg, about 540 mg 90mg, about 200mg, about 210mg, about 220mg, about 230mg, about 240mg, about 250mg, about 260mg, about 270mg, about 280mg, about 290mg, about 300mg, about 310mg, about 320mg, about 330mg, about 340mg, about 350mg, about 360mg, about 370mg, about 380mg, about 390mg, about 400mg, about 410mg, about 420mg, about 430mg, about 440mg, about 450mg, about 460 mg, about 470mg, about 480mg, about 490mg, about 500mg, about 510mg, about 520mg, about 530mg, about 540mg, about 550mg, about 560mg, about 570mg, about 580mg, about 590mg, about 600mg, about 610mg, about 620mg, about 630mg, about 640mg, about 650mg, about 660mg, about 670mg, about 680mg, about 690mg, about 700mg, about 710mg, about 720mg, about 730mg , about 740 mg, about 750 mg, about 760 mg, about 770 mg, about 780 mg, about 790 mg, about 800 mg, about 810 mg, about 820 mg, about 830 mg, about 840 mg, about 850 mg, about 860 mg, about 870 mg, about 880 mg, about 890 mg, about 900 mg, about 910 mg, about 920 mg, about 930 mg, about 940 mg, about 950 mg, about 960 mg, about 970 mg, about 980 mg, about 990 mg or about 1000 mg.

[0147] The amount of a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or antigen-binding fragment thereof) contained in a single dose can be expressed in terms of milligrams of antibody per kilogram of subject body weight (i.e., mg / kg). In some embodiments, the anti-CTLA-4 antibody can be administered at a dose of about 0.1 mg / kg to about 20 mg / kg of patient body weight. In some embodiments, the methods of the present disclosure comprise administering a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or antigen-binding fragment thereof) at a dose of about 1 mg / kg to 3 mg / kg, 1 mg / kg to 5 mg / kg, 1 mg / kg to 10 mg / kg, 1 mg / kg, 3 mg / kg, 5 mg / kg, 10 mg / kg, or 15 mg / kg of patient body weight.

[0148] In some embodiments, each dose of the CTLA-4 inhibitor comprises 0.1 to 10 mg / kg (e.g., 0.3 mg / kg, 1 mg / kg, 3 mg / kg, or 10 mg / kg) of the subject's body weight. In certain other embodiments, each dose comprises 5 to 1000 mg of a CTLA-4 inhibitor (e.g., an anti-CTLA-4 antibody or antigen-binding fragment thereof), for example, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 40 mg, 45 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg, 850 mg, 900 mg, 950 mg, or 1000 mg of a CTLA-4 inhibitor.

[0149] In some embodiments, the methods of the present disclosure further comprise administering to the subject an additional therapeutic agent or treatment. Additional therapeutic agents or treatments may be administered to improve anti-tumor efficacy, to reduce toxic effects of one or more treatments, and / or to reduce the dose of one or more treatments. In various embodiments, additional therapeutic agents or treatments may include one or more of the following: radiation, surgery, cancer vaccines, imiquimod, antiviral agents (e.g., cidofovir), photodynamic therapy, lymphocyte activation gene 3 (LAG3) inhibitors (e.g., anti-LAG3 antibodies, glucocorticoid-induced tumor necrosis factor receptor (GITR) agonists (e.g., anti-GITR antibodies), inhibitors containing T cell immunoglobulin and mucin-3 (TIM3), inhibitors of B and T lymphocyte attenuators (BTLA), inhibitors of T cell immune receptors with Ig and ITIM domains (TIGIT), CD38 inhibitors, CD47 inhibitors, indoleamine-2,3-dioxygenase (IDO) inhibitors, CD28 activators, vascular endothelial growth factor (VEGF) antagonists (e.g., "VEGF-Trap" (e.g., aflibercept), or anti-VEGF antibodies or antigen-binding fragments thereof (e.g., bevacizumab or ranibizumab) or small molecule kinase inhibitors of VEGF receptors (e.g., sunitinib, sorafenib or pazopanib)), angiopoietin-2 (Ang2) inhibitors, transforming growth factor β (TGFβ) inhibitors, epidermal growth factor receptor (EGFR) inhibitors, antibodies against tumor-specific antigens (e.g., CA9, CA125, melanoma-associated antigen 3 (MAGE3), carcinoembryonic antigen (CEA), vimentin, tumor-M2-PK, prostate-specific antigen ( antigen (PSA), mucin-1, MART-1, and CA19-9), vaccines (e.g., BCG), granulocyte-macrophage colony-stimulating factor (GM-CSF), second oncolytic viruses, cytotoxins, chemotherapeutic agents (e.g., pemetrexed, dacarbazine, temozolomide, cyclophosphamide, docetaxel, doxorubicin, daunorubicin, cisplatin, carboplatin, gemcitabine, methotrexate, pterin, mitoxantrone, oxaliplatin, paclitaxel, topotecan, irinotecan, vinorelbine and vincristine), IL-6R inhibitors, IL-4R inhibitors, IL-10 inhibitors, cytokines (e.g., IL-2, IL-7, IL-12, IL-21 and IL-15), antibody drug conjugates, anti-inflammatory drugs (e.g., corticosteroids, non-steroidal anti-inflammatory drugs (NSAIDs)), cryotherapy, anti-HPV therapy, laser therapy, electrosurgical removal of cells containing HPV, and combinations thereof.

[0150] In some embodiments, the method further comprises administering an additional therapeutic agent, such as an anticancer drug. As used herein, "anticancer drug" means any agent that can be used to treat cancer, including but not limited to cytotoxins and agents, such as antimetabolites, alkylating agents, anthracyclines, antibiotics, antimitotic agents, procarbazine, hydroxyurea, asparaginase, corticosteroids, mitotane (O, P'-(DDD)), biological agents (e.g., antibodies and interferons) and radiopharmaceuticals. "Cytotoxin or cytotoxic agent" as used herein also refers to chemotherapeutic agents and means any agent that is harmful to cells. Examples include TAXOL (paclitaxel), temozolomide, cytochalasin B, gramicidin D, ethidium bromide, emetine, cisplatin, mitomycin, etoposide, teniposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxy anthracin dione, mitoxantrone, mithramycin, actinomycin D, 1-dihydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol and puromycin, and analogs or homologs thereof.

[0151] Additional Definitions

[0152] In order to help understand the specific embodiments of the compositions and methods according to the present disclosure, some clear definitions are provided to promote the clear disclosure of various aspects of the present disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by ordinary technicians in the field to which the present disclosure belongs.

[0153] As used herein, the term "agent" refers to a chemical compound, a mixture of chemical compounds, a biomacromolecule (e.g., a nucleic acid, an antibody, a protein or a portion thereof (e.g., a peptide)), or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues. The activity of such an agent may make it suitable as a "therapeutic agent," which is a biologically, physiologically, or pharmacologically active substance (or substances) that acts locally or systemically in a subject. In the context of the present disclosure, the term "therapeutic agent" refers to any PD-1 pathway inhibitor, CTLA-4 inhibitor, or oncolytic virus disclosed herein.

[0154] As used herein, the terms "therapeutic agent," "agent with therapeutic ability," or "therapeutic agent" are used interchangeably and refer to a molecule or compound that confers some beneficial effect upon administration to a subject. Beneficial effects include achieving a diagnostic determination; amelioration of a disease, symptom, disorder, or pathological condition; reducing or preventing the onset of a disease, symptom, disorder, or condition; and generally combating a disease, symptom, disorder, or pathological condition.

[0155] As used herein, the term "pharmaceutically acceptable" refers to a material, such as a carrier or diluent, that does not destroy the biological activity or properties of the composition and is relatively non-toxic, i.e., the material can be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any component of the composition in which it is contained.

[0156] As used herein, the term "pharmaceutically acceptable carrier" includes pharmaceutically acceptable salts, pharmaceutically acceptable materials, compositions or carriers, such as liquid or solid fillers, diluents, excipients, solvents or encapsulating materials, which participate in carrying or transporting the compounds of the present disclosure in or to a subject so that they can perform their intended function. Generally speaking, such compounds are carried or transported from one organ or body part to another organ or body part. 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 subject. Some examples of materials that can be used as pharmaceutically 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 waxes; 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 glycerol, sorbitol, mannitol, 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; solution); ethanol; phosphate buffer solution; diluent; granulating agent; lubricant; binder; disintegrant; wetting agent; emulsifier; colorant; release agent; coating agent; sweetener; flavoring agent; aromatic agent; preservative; antioxidant; plasticizer; gelling agent; thickener; hardener; fixative; suspending agent; surfactant; humectant; carrier; stabilizer; and other non-toxic compatible substances for pharmaceutical preparations, or any combination thereof. As used herein, "pharmaceutically acceptable carrier" also includes any and all coatings, antibacterial and antifungal agents, and absorption delaying agents, etc., that are compatible with the activity of one or more components of the present disclosure and are physiologically acceptable to the subject. Supplementary active compounds can also be incorporated into the composition.

[0157] Doses are usually expressed with respect to body weight. Thus, a dose expressed as [g, mg or other unit] / kg (or g, mg, etc.) usually refers to [g, mg or other unit] " / kg (or g, mg, etc.) body weight", even if the term "body weight" is not explicitly mentioned. Treatment may include a variety of "unit doses". A unit dose is defined as a therapeutic composition containing a predetermined amount. A unit dose need not be administered as a single injection, but may include continuous infusion over a set period of time. For oncolytic viruses, unit doses may be described in terms of plaque forming units (pfu) or viral particles for viral constructs. Unit doses range from 10 3 , 10 4 , 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , 10 11 , 10 12 , 10 13 pfu or vp and higher. Alternatively, 1 to 100, 10 to 50, 100 to 1000, or up to about 1×10 4 , 1×10 5 , 1×10 6 , 1×10 7 , 1×10 8 , 1×10 9 , 1×10 10 , 1×10 11 , 1×10 12 , 1×10 13 , 1×10 14 or 1×10 15 or more infectious virus particles (vp) to the patient or patient cells. Alternatively, the unit dose of oncolytic virus is determined by TCID 50 Indicates. 50 ” refers to “tissue culture infectious dose” and is defined as the virus dilution required to infect 50% of a given batch of inoculated cell cultures. A variety of methods known to those skilled in the art can be used to calculate TCID 50 , the method includes the Spearman-Karber method used throughout this specification. For a description of the Spearman-Karber method, see BW Mahy & H.O. Kangro, Virology Methods Manual 25-46 (1996). In some embodiments, the unit dose range is 10 3 , 10 4 , 10 5 , 10 6, 10 7 , 10 8 , 10 9 , 10 10 , 10 11 , 10 12 , 10 13 TCID 50 and above or any range therebetween.

[0158] As used herein, the term "disease" is intended to be generally synonymous and used interchangeably with the terms "disorder" and "condition" (as in medical condition) because they all reflect an abnormal condition of the human or animal body or one of its parts that impairs normal function (e.g., cancer), usually manifests itself by distinguishing signs and symptoms, and results in a decrease in the duration or quality of life of the human or animal.

[0159] As used herein, the term "in vitro" refers to events that occur in an artificial environment (eg, in a test tube or reaction vessel, in cell culture, etc.), rather than in a multicellular organism.

[0160] As used herein, the term "in vivo" refers to events that occur within a multicellular organism (eg, a non-human animal).

[0161] As used herein, unmodified nouns mean one or more, unless the context clearly indicates otherwise.

[0162] Unless otherwise stated, the terms "including," "comprising," "containing," or "having" and variations thereof used herein are meant to encompass the items listed thereafter and equivalents thereof as well as additional subject matter.

[0163] As used herein, the phrases "in one embodiment," "in embodiments," "in some embodiments," etc. are used repeatedly. Such phrases are not necessarily referring to the same embodiment, but they may unless the context indicates otherwise.

[0164] As used herein, the term "and / or" or " / " means any one item, any combination of items, or all items related to the term.

[0165] As used herein, the word "substantially" does not exclude "completely", for example, a composition "substantially free of" Y may be completely free of Y. If necessary, the word "substantially" may be omitted from the definition of the present disclosure.

[0166] As used herein, the term "each," when used in relation to a collection of items, is intended to identify a single item in the collection, but does not necessarily refer to every item in the collection. Exceptions may occur if the explicit disclosure or context clearly dictates otherwise.

[0167] As used herein, when applied to one or more target values, the term "approximately" or "approximately" refers to a value similar to the reference value. In some embodiments, unless otherwise specified or other situations are obvious from the context, the term "approximately" or "approximately" refers to a value range (except for such a number that will exceed 100% of the possible value) within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or less in either direction of the reference value (greater than or less than) (except for such a number that will exceed 100% of the possible value). Unless otherwise specified herein, the term "approximately" is intended to include values ​​close to the range, such as weight percentages, which are equivalent in terms of the functions of a single component, composition or embodiment.

[0168] As disclosed herein, multiple value ranges are provided.It should be understood that, unless the context clearly states otherwise, each intermediate value between the upper limit value and the lower limit value of the scope is also specifically disclosed, up to one-tenth of the lower limit unit.Each smaller range between any specified value or intermediate value in the stated range and any other specified value or intermediate value in the stated range is included in the present disclosure.The upper limit value and lower limit value of these smaller ranges can be included or excluded outside the scope independently, and one of them, both limits do not include or both limits are included in each scope of the smaller range is also included in the present disclosure, subject to any clear exclusion in the stated range.When the stated range includes one or two limits, the scope of excluding any one or two of those included limits is also included in the present disclosure.

[0169] The use of any and all examples or exemplary language (e.g., "such as") provided herein is intended only to better illustrate the present disclosure and does not limit the scope of the present disclosure unless otherwise stated. No language in this specification should be construed to indicate any non-claimed element is essential to the practice of the present disclosure.

[0170] Unless otherwise noted herein or clearly contradictory to the context, all methods described herein are performed in any suitable order. With respect to any method provided, the steps of the method may occur simultaneously or sequentially. Unless otherwise noted, when the sequence of steps of the method occurs, these steps may occur in any order. Where the method includes a combination of steps, unless otherwise noted herein, each and every combination or sub-combination of steps is encompassed within the scope of the present disclosure.

[0171] Each publication, patent application, patent and other reference cited herein is incorporated by reference in its entirety to the extent that it does not conflict with the present disclosure. The publications disclosed herein are provided only for their disclosures prior to the date of submission of the present disclosure. Nothing herein should be interpreted as admitting that the present disclosure is not entitled to such publications due to prior disclosures. In addition, the publication date provided may be different from the actual publication date, which may require independent confirmation.

[0172] It should be understood that the examples and embodiments described herein are for illustrative purposes only and that a person skilled in the art will identify themselves with various modifications or changes thereto, and that these modifications or changes are intended to be included within the spirit and scope of this application and the scope of the appended claims.

[0173] Example

[0174] The following examples are presented to provide a complete disclosure and description of how to prepare and use the methods and compositions of the present disclosure to those of ordinary skill in the art, and are not intended to limit the scope of the invention that the inventors regard as their invention. Likewise, the present disclosure is not limited to any particular preferred embodiment described herein. In fact, when reading this specification, modifications and variations of the present embodiment may be obvious to those skilled in the art, and may be modified and varied without departing from its spirit and scope. Efforts have been made to ensure accuracy with respect to the numerals used (e.g., amount, temperature, etc.), but some experimental errors and deviations should be considered. Unless otherwise stated, parts are parts by weight, molecular weight is average molecular weight, temperature is in degrees Celsius, room temperature is about 25°C, and pressure is atmospheric pressure or near atmospheric pressure.

[0175] Embodiment 1: A phase 2 trial of Voyager V1 (vesicular stomatitis virus expressing NIS and human interferon beta, VV1) in combination with cemiplimab in patients with hepatocellular carcinoma, non-small cell lung cancer, melanoma, or endometrial cancer

[0176] This example relates to an open-label, Simon 2 Phase 2 clinical trial to determine the anti-tumor activity of Voyager V1 (VV1) in combination with cemiplimab alone and with ipilimumab in patients with hepatocellular carcinoma, non-small cell lung cancer (NSCLC), melanoma, or endometrial cancer.

[0177] Background: VV1 is an oncolytic vesicular stomatitis virus that has been engineered to express human interferon beta (IFNβ) to enhance cellular antitumor immune responses and tumor selectivity. New immunotherapies are needed to address patient populations that have never responded or no longer respond to immune checkpoint inhibitors (CPIs). Phase 1 studies have demonstrated VV1 antitumor activity in several malignancies with or without CPIs. This example involves optimizing the efficacy of VV1 in combination with cemiplimab (an anti-PD1 antibody approved for lung, basal and squamous cell skin cancer) in two ways: using higher doses and triple combinations in representative patient populations.

[0178] Patients with advanced melanoma (after progression on anti-PD1) are enrolled in the study, which will eventually include first-line NSCLC patients whose disease is amenable to immunotherapy. The study first explored the preliminary antitumor activity, safety, and immunogenicity of the following combination: 1.0×10 11 The dose is intravenous (IV) once on VV1 followed by Q3W IV cemiplimab combination starting on D8, or the same regimen concurrently once on D1 1.0 × 10 9 TCID 50 Additional intratumoral injection of VV1. Patients receive IV simiprilimab Q3W until disease progression or intolerable toxicity is determined. Once at least six patients in the two melanoma double-arm groups have been safely treated, the 3rd (triple) group will begin to increase a single dose of 50mg ipilimumab at D1 in the same patient population. Once at least six melanoma patients have safely received triple therapy, the first-line NSCLC group will start triple therapy. Based on the Simon 2 stage design, the group will be expanded using a type I error rate of 0.05 and a power of 85%. The ineffective overall response rate (ORR) is 10%, the target for second-line melanoma is 35%, and the ineffective ORR in first-line NSCLC is 40%, with a true target of 70%. Melanoma cohort will need ≥2 responses in 10 patients in the first stage to add 11 more in the second stage, while NSCLC will first need 5 / 9 evaluable patients to respond, and then add another 13 to complete the design. According to RECIST v1.1, response is assessed at week 7, then Q9W. This study includes serial biopsies in ≥3 / 10 patients in the first stage of each IV melanoma cohort (double and triple therapy), all patients in the second stage of these IV melanoma cohorts, and all patients in both the first stage and the second stage of intravenous / intratumoral (IV / IT) melanoma cohorts to allow a thorough investigation of the impact of the three immunotherapies in the study.

[0179] Research Rationale: Cancer immunotherapy has shown significant clinical success, producing durable responses in patients with advanced, previously refractory cancers. Specifically, immune CPI agents that block tumor signals that co-opt the suppression of immune responses, thereby activating T cell-mediated anti-tumor immunity, have shown clinical efficacy and are approved for the treatment of a variety of indications, including melanoma, NSCLC, urothelial carcinoma, head and neck cancer, gastric cancer, hepatocellular carcinoma (HCC), and other tumors. Most patients remain resistant to CPI treatment, with response rates ranging from 10% to 40%, depending on the indication. Patient populations whose diseases are resistant to or refractory to CPIs are now becoming a significant unmet medical need. Not all NSCLC or melanoma patients, for whom anti-PD-(L)1 therapy has become the standard of care, receive long-term, durable benefits from such treatment. Once resistant, these patients have limited treatment options. In addition, in these indications, patients who do not respond to and are refractory to CPI treatment continue to have unmet medical needs.

[0180] It is predicted that the combination therapy of oncolytic VSV-IFNβ-NIS with CPI will enhance the infiltration and activation of intratumoral CD8+ T cells to enhance the therapeutic response of cancer patients who were previously resistant to CPI monotherapy. This study, conducted in a population of melanoma patients who were refractory to single-agent CPI, was designed to provide insights into the putative mechanism of resensitization to CPI by infusion of a single dose of oncolytic virus, assuming that cell infection will expand the available endogenous anti-tumor T cell pool, increase tumor cell antigenicity recognized by T cells, enhance antigen presentation, and ultimately convert a non-inflammatory or immune-excluded phenotype to a highly inflammatory phenotype.

[0181] Study Objectives: The primary objective of the study was to determine preliminary antitumor activity in each study cohort, as measured by overall response rate (ORR) according to standard criteria (RECIST 1.1) [Eisenhauer 2009]. Secondary objectives of the study included: determining the safety and tolerability of VV1 in combination with cemiplimab; evaluating the systemic pharmacokinetics (PK) of VV1 and cemiplimab; studying the pharmacodynamics of VV1 in the blood by measuring serum IFNβ expression: determining preliminary clinical activity in each cohort [e.g., progression-free survival (PFS), duration of response (DOR), disease control rate (DCR), and overall survival (OS)]; and evaluating the safety and efficacy of administering VV1, cemiplimab, and ipilimumab. Additional objectives include assessment of: immune cell infiltration in selected pre- and post-treatment tumor samples; time course of viremia, viral shedding, and viral persistence; effects on host peripheral blood immune cell expansion / suppression; immunogenicity as measured by anti-drug antibodies to VV1+cemiplimab; and the ability of ruxolitinib to mitigate the risk of IFNβ toxicity.

[0182] Overall study design: This is a Phase 2 study designed to determine preliminary antitumor activity and to determine the safety of VV1 in combination with cemiplimab alone and subsequently with ipilimumab. The hepatocellular carcinoma and endometrial cancer cohorts were closed, and the melanoma and NSCLC cohorts were modified in the following manner: the VV1 dose was escalated to 1.0 × 10 11 TCID 50 The NSCLC cohort will enroll patients who have not received CPI treatment. Figure 1 Shown in . Patients with advanced melanoma will be recruited into two parallel double cohorts; in one cohort (IV melanoma cohort), patients will receive IV VV1, and patients in the other cohort (IV / IT melanoma cohort) will receive both IV VV1 and IT VV1; on day 8 after receiving VV1 treatment, both cohorts will receive IV simiprilimab. After completing the high-dose double melanoma cohort, the subsequent triple melanoma cohort will receive IV VV1, IV simiprilimab and ipilimumab (1 dose) combination treatment. Finally, the cohort of patients with NSCLC will receive the triple combination.

[0183] Found as many as 1.7×10 11 TCID 50The dose of VSV-IFNβ-NIS (IV monotherapy) was safe and may be more effective than lower doses.

[0184] All patients enrolled in this study will be required to stay in the hospital overnight on Day 1 and, if clinically stable, may be discharged for outpatient follow-up on Day 2. Patients will then be required to undergo outpatient safety evaluations on Days 3, 4, and 5. Laboratory assessments performed on Days 2, 3, 4, and 5 must be evaluated and possible IFNβ toxicity assessed on these days before the patient is discharged home during these periods.

[0185] Patients will be recruited in each melanoma double group at the same time. The 3 patients of the safety lead-in in the IV group will be dosed and observed for 28 days, and their safety data will be reviewed before the IT / IV melanoma group will be recruited. A total of six (6) patients in the two groups will be dosed and observed for one cycle. Once the combination is considered safe, recruitment in the IV melanoma triple group will be carried out.

[0186] For all cohorts in the study, the Simon two-stage design [Simon 1989] will be used. In the first stage, 10 evaluable patients will be added in each melanoma tumor cohort; based on a combination of safety and efficacy, the cohort will progress to the second stage. If there are less than 2 objective responses in each melanoma cohort of 10 patients, the cohort will be stopped; otherwise, 11 additional patients will be added, for a total of 21 patients in each melanoma cohort. A similar design will be used for NSCLC, where 9 evaluable patients will be enrolled in the first stage. In this NSCLC cohort, for a total of 22 evaluable patients, a higher efficacy bar of at least 5 of the 9 patients with an objective response will be required to trigger the second stage.

[0187] Patients in the IV melanoma cohort will receive IV VV1, and patients in the IV / IT melanoma cohort will receive both IV VV1 and IT VV1 on Day 1 of Cycle 1; both cohorts will receive IV cemiplimab on Day 8 after receiving VV1. US package insert, Cemiplimab treatment will continue Q3W, with efficacy assessments after cycle 2, and then every 9 weeks until progressive disease (PD). After completion of the high-dose melanoma cohort, the subsequent triple melanoma cohort will receive a combination of IV VV1, ipilimumab (1 dose), and IV Cemiplimab. Finally, a group of patients with NSCLC will receive the triple combination.

[0188] NCI CTCAE, v5.0 will be used to grade overall toxicity and ASTCT consensus grading will be used to grade CRS and neurotoxicity [Lee 2019]. Safety assessments will include AEs, SAEs, PEs, vital sign measurements, ECOG status, clinical safety laboratory evaluations (hematology, serum chemistry and hepatic panel, coagulation, and urinalysis), and ECG.

[0189] Study Population: Only patients who met all the inclusion criteria and did not meet any of the exclusion criteria were eligible to receive the drug.

[0190] Inclusion Criteria: Patients who meet all of the following inclusion criteria on Screening / First Dosing Day 1 will be eligible to participate in this study: (1) Age ≥18 years on the day of signing the informed consent form. (2) Specific according to tumor cohort: (a) For the NSCLC cohort, a diagnosis of histologically confirmed advanced and / or metastatic NSCLC suitable for first-line immunotherapy. NSCLC with activating EGFR mutations or anaplastic lymphoma kinase (ALK) rearrangements must also have progressed after receiving available EGFR or ALK targeted therapy in addition to treatment with platinum-based chemotherapy (unless ineligible for platinum treatment). (i) Ability to provide archived (or fresh) formalin-fixed, paraffin-embedded tumor tissue collected within 6 months prior to enrollment for determination of programmed death ligand 1 (PD-L1) status. (ii) PD-L1 status ≥50% according to local standard testing. Samples should be provided to a central laboratory for post-hoc centralized testing. (b) For the melanoma cohort, a diagnosis of histologically confirmed advanced and / or cutaneous metastatic melanoma in which radiological progression has been demonstrated during treatment with a PD-(L)1 immune checkpoint inhibitor and for whom no existing options are considered to provide clinical benefit (first-line PD-(L)1 therapy only allowed). Progression on ipilimumab is not required. Patients with BRAF V600 mutations must have progressed on, or been intolerant of, BRAF + / - MEK inhibitor therapy. i. Patients with BRAF V600-positive tumors should have received prior treatment with a BRAF inhibitor (alone or in combination with a MEK inhibitor) in addition to treatment with pembrolizumab or nivolumab with or without ipilimumab, or in addition to having refused targeted therapy. NOTE: Based on the investigator's decision, patients with BRAF V600-positive tumors and no clinically significant tumor-related symptoms or evidence of rapidly progressive disease do not need to be treated with a BRAF inhibitor (alone or in combination with a MEK inhibitor). For the IV / IT melanoma cohort: (i) At least one tumor lesion was receptive to IT injection by palpation or ultrasound. Injections of deep visceral lesions were not permitted; (ii) Consent to provide a newly obtained biopsy of the injected lesion prior to the start of study treatment and to repeat the biopsy twice during study treatment, and to provide tissue harvested for biomarker analysis. The tissue obtained for biopsy must not have been previously irradiated, but new or progressing lesions in the irradiated area are acceptable. (3) For patients previously treated with anti-PD-(L)1 therapy: (a) The last dose of anti-PD-(L)1 must be within 16 weeks of starting study treatment. (b) Patients must have received at least 4 doses on a q2w regimen, 3 doses on a q3w regimen, or 2 doses on a q4w regimen of prior anti-PD-(L)1 therapy.(c) Progression on prior anti-PD-(L)1 therapy must be defined by: (i) Radiographic progression on a single radiographic scan if treatment with anti-PD-(L)1 was ≥16 weeks; (ii) Radiographic progression on two consecutive radiographic scans at least 4 weeks apart if treatment with anti-PD-(L)1 therapy was between 8 and 16 weeks; if radiographic progression is associated with clinical progression, a single scan may be used for assessment; (iii) if progression is only in lymph nodes, a biopsy is required to provide histological confirmation of progression in lymph nodes. (4) Measurable disease based on RECIST 1.1. (5) Performance status of 0 or 1 on the ECOG performance scale; (6) Life expectancy >3 months; (7) Willingness to provide biological samples required for the duration of the study, including fresh tumor biopsies. At least 3 of the 10 patients in stage 1 of each IV melanoma cohort (double and triple therapy) must provide fresh tumor biopsies before and on-treatment. In Phase 2 of these cohorts, all patients must provide fresh tumor biopsy samples before and during treatment. All patients in both Phase 1 and Phase 2 of the IT / IV melanoma cohort must provide fresh tumor biopsy samples before and during treatment. (8) Adequate organ function is defined as laboratory values ​​obtained ≤14 days before enrollment as shown in Table 1; (9) Negative pregnancy test for female patients of childbearing potential; (10) Lack of active CNS involvement. Note: Pre-enrollment imaging is not mandatory for asymptomatic patients; (11) Ability to provide written informed consent.

[0191] Table 1: Adequate organ function

[0192]

[0193]

[0194] Exclusion Criteria: Patients who meet any of the following exclusion criteria on Screening / First Dose Day -1 will not participate in this study: (1) Alternative curative treatment options are available and accepted by the patient; (2) Recent or ongoing serious infection, including any active grade 3 or higher viral, bacterial, or fungal infection within 2 weeks of enrollment according to NCI CTCAE, v5.0; (3) Patients diagnosed with ocular, mucosal, or acral melanoma; (4) Known HIV seropositivity and active infection with HIV. a. Patients who are HIV seropositive but are receiving antiviral therapy and do not show detectable viral load and have a normal CD4 T cell count for at least 6 months are eligible; (5) HBV seropositive and have evidence of active HBV viral infection. Patients who are HBsAg negative and HBV viral DNA negative are eligible. (a) Patients with HBV but have received antiviral therapy and have not shown detectable viral DNA for 6 months are eligible; (b) Patients who are seropositive due to HBV vaccine are eligible; (6) HCV seropositivity and active HCV viral infection.a. Patients with HCV who have received antiviral therapy and have not shown detectable HCV viral RNA for 6 months are eligible; (7) known history of active or latent TB; (8) any concomitant serious health condition that the investigator believes would place the patient at excessive risk from this study, including uncontrolled hypertension and / or diabetes, clinically significant pulmonary disease (e.g., chronic obstructive pulmonary disease requiring hospitalization within 3 months), or neurological disorder (e.g., active seizure disorder within 3 months); (9) prior treatment within the following time ranges before the planned start of study treatment as follows: (a) small molecule inhibitors and / or other investigational agents: ≤ 2 weeks or 5 half-lives, whichever is shorter; (b) chemotherapy, other monoclonal antibodies, antibody-drug conjugates, or other similar experimental treatments: ≤ 3 weeks or 5 half-lives, whichever is shorter; (c) radioimmunoconjugates (10) NYHA class III or IV, known symptomatic coronary artery disease, or symptoms of coronary artery disease on review of systems, or known arrhythmias (atrial fibrillation or SVT); (11) Any known or suspected active organ-threatening autoimmune disease, such as inflammatory bowel disease, autoimmune hepatitis, lupus, or pneumonitis, except for hypothyroidism and type 1 diabetes controlled with therapy; (12) Immunodeficiency or immunosuppression, including systemic corticosteroids >10 mg / day of prednisone or equivalent within 1 week prior to the planned start of study treatment; (13) History of grade 3 or 4 immune-mediated adverse reactions to immuno-CPIs; (14) Toxicity from previous therapy that has not resolved to grade 1 or less; (15) Current or historical non-infectious pneumonitis requiring steroids, or current pneumonitis; (16) High burden disease (high volume disease), as clinically assessed by a medical monitor using parameters such as radiographic signatures and tumor markers or LDH; (17) portal vein thrombosis involving more than the intrahepatic portal vein branches: thrombosis of the right or left portal vein branches or bifurcations, partial or complete occlusion of the main portal vein; (18) known concurrent malignancy. Exceptions: basal cell carcinoma of the skin, squamous cell carcinoma of the skin, or in situ cervical cancer treated with curative intent, prostate cancer confined to the prostate with a Gleason score <6 or a PSA <1, and any stage 1 cancer treated with curative intent or any previous cancer with a disease-free interval of ≥3 years; (19) any other concurrent anticancer treatment (chemotherapy, immunotherapy, radiation therapy, or any adjuvant therapy considered investigational [for non-FDA-approved indications and research investigations]); (20) receipt of a live vaccine within 30 days of the planned start of study treatment. Injectable seasonal influenza vaccines are generally inactivated influenza vaccines and are permitted. COVID-19 mRNA vaccines are also permitted; intranasal influenza vaccines (e.g. ) is a live attenuated vaccine and is not permitted 4 weeks before the first dose of VV1 and 12 weeks after the last dose of VV1; (21) Because the genotoxic, mutagenic, and teratogenic effects of the investigational agents involved in this study on the developing fetus and neonate are unknown, any of the following conditions are excluded: (a) Sexually active men or WOCBP who are unwilling to practice highly effective contraception before the first dose / first treatment, during the study, and for at least 4 weeks after the last dose*. Highly effective contraceptive measures include: stable use of combined (estrogen and progestin) hormonal contraception (oral, vaginal, transdermal) or progestin-only hormonal contraception (oral, injectable, implantable) associated with ovulation suppression for 2 or more menstrual cycles before screening; intrauterine device (IUD); intrauterine hormone-releasing system (IUS); bilateral tubal ligation; vasectomized partners; and / or sexual abstinence. (i) stable use of combined (estrogen and progestin) hormonal contraception (oral, intravaginal, transdermal) or progestin-only hormonal contraception (oral, injectable, implantable) associated with ovulation suppression for the initiation of 2 or more menstrual cycles prior to screening; (ii) IUD; IUS; (iii) bilateral tubal ligation; (iv) vasectomized partner (provided that the vasectomized male partner is the only sexual partner of the WOCBP study participant and the vasectomized partner has been medically evaluated as a successful surgical procedure); and / or sexual abstinence *WOCBP is defined as a woman who is fertile after menarche until she becomes postmenopausal, unless permanently sterilized. Permanent sterilization methods include hysterectomy, bilateral salpingectomy, and bilateral oophorectomy. Postmenopausal women are defined as having no menstruation for 12 months without an alternative medical reason. High follicle stimulating hormone (FSH) levels in the postmenopausal range can be used to determine the postmenopausal state in women who are not using hormonal contraception or hormone replacement therapy. However, in the absence of 12 months of amenorrhea, FSH measurements alone are insufficient to determine the onset of postmenopausal status. Pregnancy testing and contraception are not required for women with a documented hysterectomy or tubal ligation. Sexual abstinence is considered a highly effective approach only if it is defined as abstention from heterosexual intercourse during the entire risk period associated with the study drug. The reliability of sexual abstinence needs to be evaluated based on the duration of the clinical trial and the patient's preferences and usual lifestyle. Periodic abstinence (calendar method, symptom-temperature method, postovulation method), withdrawal method (excretion), spermicide-only method and lactational amenorrhoea method (LAM) are unacceptable contraceptive methods. Female and male condoms should not be used together; (22) pregnant or lactating women; (23) men who are unwilling to use condoms during vaginal intercourse, while taking medication and 4 weeks after stopping treatment (even if they have undergone a previous vasectomy); (24) such patients: due to receiving ipilimumab: a. Hypersensitivity to the excipients of ipilimumab: hypersensitivity to the excipients of the product (Tris hydrochloride (2-amino-2-hydroxymethyl-1,3-propanediol hydrochloride), sodium chloride, mannitol (E421), pentetic acid (diethylenetriaminepentaacetic acid), polysorbate 80, sodium hydroxide (for pH adjustment), hydrochloric acid (for pH adjustment)

[0195] Study treatment: VSV-IFNβ-NIS was prepared (ie, undiluted) in a buffer consisting of 5% sucrose, 50 mM Tris (pH 7.4), 2 mM MgCl2. After preparation, the virus was stored frozen at ≤ -65°C until ready for use immediately prior to administration. Cimiprilimab Follow the package insert for preparation, storage, and handling. Follow its package insert for preparation, storage, and handling.

[0196] Dosage regimen and administration: The first cycle is 28 days. Subsequent cycles are 21 days. Both IV and IT VV1 will be administered on day 1. In the triple group, Ipilimumab should also be given on day 1. Cimiprili monoclonal antibody is given on day 8 of cycle 1 and day 1 of cycle 2 and thereafter (21-day cycles). The treatment regimen for combined therapy is shown in Table 2. Cimiprili monoclonal antibody should be given on day 8 of cycle 1 (28-day cycle) and then on day 1 of each subsequent 21-day cycle for up to 2 years. In the triple group, Ipilimumab should be given on day 1.

[0197] Table 2: Combination therapy regimens

[0198]

[0199] *IT dose and schedule for IV / IT melanoma cohort only.

[0200] **Ipilimumab only in the triplet cohort.

[0201] Cimiprizumab Administration: Cimiprizumab is administered as an IV infusion over 30 minutes under close observation according to institutional standard of care. Cimiprizumab should be given on Day 8 of Cycle 1 (28-day cycle) and on Day 1 of each subsequent 21-day cycle for up to 2 years.

[0202] Intravenous VVi administration: The dose of VV1 infused IV on day 1 will be 1.0 × 10 11 TCID 50 The virus was diluted in 100 mL of normal saline containing 1% HSA and administered by slow IV infusion (30 minutes) under close observation.

[0203] IV / IT Melanoma Cohort Dosing: For this cohort, all patients will receive both IV VV1 and IT VV1. For convenience, the order of administration should be adjusted. On Day 1, VV1 will be administered by injection at 1×10 9 TCID 50 A total dose of VV1 is administered once intratumorally (IT) to one or more tumor lesions. The needle type can also be at the discretion of the investigator to best distribute the virus within the lesion. VV1 can be administered IT to multiple accessible lesions using the appropriate syringe and needle type for the lesion location. The rationale for dosing multiple lesions is to generate an immune response against a greater range of genetic mutations and antigenic diversity than is possible in a single lesion, which in turn is expected to reduce the chance of tumor immune escape.

[0204] Only direct administration into the tumor or accessible lymph nodes is allowed. Injections of skin, soft tissue, or deep visceral lesions (e.g., liver or lung metastases) that require radiation control by CT or MRI, or referral to an interventional radiologist are not allowed. If multiple lesions are specified for injection, the lesions should be located in different parts of the body as much as possible. Once the lesions are identified, the maximum volume to be injected is 4 mL, distributed over different injectable tumors. The size of the lesion must be greater than 0.5 cm to qualify for ITVV1 administration, with a total of up to 8 lesions. The allocation of the infusion volume will be determined based on the size of the lesion to be injected (Table 3):

[0205] Table 3: IT VV1 administration

[0206]

[0207] *Only lesions ≤ 0.5 cm may be injected if clinically responsive to prior IT therapy.

[0208] IV VV1 administration will be performed in a similar manner to all other cohorts. Patients will receive a single IV dose of 1.0 × 1011 TCID 50 All of the above concomitant measures to mitigate infusion-related reactions (IRRs) should be initiated prior to IT administration in patients in this cohort. If a patient experiences an IRR as a result of the first administration of VV1, the second form of VV1 may be postponed until Days 2 to 4.

[0209] Ipilimumab Administration for Triple Melanoma and NSCLC Cohorts: All patients in the triple cohort will receive both IV VV1 and ipilimumab on Day 1 of Cycle 1 and cemiplimab on Day 8. Ipilimumab will be administered as a single 50 mg IV infusion over 90 minutes. Ipilimumab will be given only on Day 1 of Cycle 1. The order of administration on Day 1 should be: ipilimumab followed by VV1.

[0210] Duration of treatment: In the absence of treatment delays due to AEs, treatment may continue for up to a maximum of 2 years or until one of the following criteria applies: (i) disease progression; (ii) intercurrent illness that prevents further administration of treatment; (iii) AE requiring treatment cessation: Any dosing interruption lasting >12 weeks, with the following exceptions: (a) Dosing interruptions of >12 weeks for non-drug-related reasons may be permitted if approved by the PI. The PI must be consulted before restarting treatment in patients with dosing interruptions lasting >12 weeks; (b) Tumor assessments should continue according to the protocol even if dosing is interrupted; (iv) The patient decides to withdraw from the study; (v) The overall or specific changes in the patient's condition make the patient unacceptable to further treatment, in the investigator's judgment. Clinical progression was defined as progression occurring when the patient's condition worsened and, in the investigator's judgment, the patient could not continue to receive study treatment, regardless of imaging findings; (vi) patient noncompliance; (vii) pregnancy; (viii) study termination by the sponsor; (ix) study agent no longer available from the drug manufacturer; (x) a discussion between the attending physician and the investigator regarding discontinuation of study treatment could be considered if the patient had a confirmed complete CR, had been treated with cemiplimab for at least 24 weeks, and had been in CR for at least two cycles.

[0211] Pre-existing and concomitant illnesses and medications: Patients should receive comprehensive supportive care during the study, including blood products, antibiotics, and treatment for concurrent medical conditions and other newly diagnosed illnesses.

[0212] All pre-existing major illnesses relevant to patient safety or that the patient has experienced prior to screening should be recorded. Additional illnesses present at the time of informed consent should be considered concomitant illnesses. Illnesses that first occurred or were detected during the study and / or worsening of concomitant illnesses during the study were recorded as AEs. If a patient enrolled in this study is infected with COVID-19, any difficulties in performing protocol-specific procedures (including administration or use of investigational products or compliance with protocol-authorized visits and laboratory / diagnostic tests) should be recorded. The reason for failure to obtain an assessment or delayed assessment should be recorded (e.g., identification of specific limitations imposed by COVID-19 that prevented protocol-specific assessments from being performed). COVID-19 positive test results should be collected as AEs. Imaging of patients with early COVID-19 pneumonia may be confused with pneumonia, which has been described in the context of immune checkpoint inhibitors. If a pneumonia AE is diagnosed, COVID-19 should be included in the differential. Once the patient has clinically recovered from the actual COVID-19 infection, or sufficient time has passed since exposure to a high-risk scenario (i.e., considered low risk), and the patient is clinically stable, protocol-specific procedures and planned treatments may continue.

[0213] Prohibited treatments: Patients should not receive other investigational agents or participate in device studies within 3 weeks prior to study entry, or should not receive systemic anticancer therapy according to eligibility criteria and during the study, and should make every effort not to start any other investigational product or device study within 28 days after the last drug administration. Participation in non-therapeutic studies is allowed. The following are not allowed 1 week before and during study drug administration: systemic steroids >10 mg / day of prednisone or equivalent unless required to treat an AE; systemic immunosuppressants; other systemic anticancer treatments; live vaccines (which should also be avoided for 3 months after the last dose of VV1 administration unless the risks outweigh the benefits).

[0214] Description of Evaluation: A medical history will be taken during screening to ensure patient eligibility and will focus on relevant current or past abnormalities or diseases of the following systems: gynecological, cardiovascular, respiratory, gastrointestinal, hepatic, biliary, renal, endocrine / metabolic, musculoskeletal, hematologic / lymphatic, neurologic / psychiatric, cutaneous, immune, infectious diseases, bleeding tendencies, and allergies / drug sensitivities.

[0215] Demographics: Information on date of birth, sex, race / ethnicity, detailed smoking history, and alcohol consumption history will be recorded during screening.

[0216] Physical Examination: A complete physical examination will include examination of the general appearance, skin, head, ears, eyes, nose, throat, heart, lungs, abdomen, lymph nodes, extremities, reproductive system (if indicated based on symptomatology and medical history), and neurologic system, as well as measurement of weight. A brief (focused) physical examination should include a cardiopulmonary assessment and abdominal examination, but will focus on new symptoms and will include examination of relevant systems as determined by the investigator. Height and weight will be assessed at Screening, and only weight will be assessed thereafter.

[0217] ECOG performance status and vital signs: ECOG performance status will be recorded based on the investigator's assessment of the patient's performance status. ECOGPS: 0 = fully active, able to perform all pre-disease manifestations without limitation; 1 = limited physical activity, but ambulatory and able to perform light or sedentary work, such as light housework, office work; 2 = ambulatory and fully self-care, but unable to perform any work activities; up to 50% and approximately more than 50% of waking hour; 3 = limited self-care; confined to bed or chair for more than 50% of waking hour; 4 = completely incapacitated; unable to perform any self-care; completely confined to bed or chair; 5 = death. Vital signs will include temperature, respiratory rate, heart rate, and systolic and diastolic blood pressure. Blood pressure and heart rate will be measured oscillometrically using a blood pressure measuring device after 5 minutes in the sitting or semi-recumbent position. The measurement will be repeated if the systolic blood pressure is below 100 mmHg or above 150 mmHg, and / or the diastolic blood pressure is below 50 mmHg or above 90 mmHg. The heart rate measurement will be repeated when it is below 50 beats per minute (bpm) or above 100 bpm. If the measurement is still outside the normal range, it will be at the discretion of the investigator whether the measurement should be repeated. Vital sign measurements outside the normal range will be assessed by the investigator as "abnormal, not clinically significant" or "abnormal, clinically significant". In the latter case, abnormal vital sign measurements will be reported as AEs and further investigation will be performed as clinically indicated.

[0218] Electrocardiogram: A single standard 12-lead ECG will be performed at Screening and at one time point after VV1 administration. If the ECG is abnormal, it must be repeated. Thereafter, an ECG should be performed as clinically indicated and in accordance with standard practice. The ECG parameters to be recorded are as follows: rhythm, PR interval, heart rate, QRS interval, QT interval, QTcF interval, and QRS axis. The ECG will be recorded while the patient is resting in the supine or semi-recumbent position. The ECG will be read at site and classified as "normal," "abnormal, not clinically significant," or "abnormal, clinically significant."

[0219] Pharmacokinetic assessments: Viral PK will be assessed by RT-PCR of VSV-IFNβ-NIS. The first whole blood sample collection (baseline) will be performed at any time within the first 24 hours before VV1 administration. Cimiprilimab PK in serum will be assessed by ELISA. Blood samples will be drawn before dosing (usually within 2 hours before study treatment) and at the end of the first cycle infusion, and the frequency will be reduced thereafter.

[0220] Tumor biopsy: At screening, all patients will have baseline FFPE tissue identified for related studies. In addition, for at least 3 of 10 patients in the first stage of each IV melanoma group (double and triple treatment), all patients in the second stage of these IV melanoma groups, and all patients in the first and second stages of the IV / IT melanoma groups, a mandatory fresh pre-treatment core biopsy (fine needle aspiration is not enough) is required. Pre-treatment biopsies must be obtained within 28 days before recruitment and sent to the central laboratory for related retrospective studies. All patients who need to provide fresh pre-treatment biopsy samples also need to undergo treatment biopsies of the same lesions on days 3 to 8 of the first cycle and day 1 of the second cycle. If the patient agrees, additional optional biopsies can be performed on any other lesions. In the case where the lesions of the initial biopsy become too small to repeat the biopsy, additional lesions can be biopsied as an alternative. For all groups in both stages 1 and 2, at the time of EOT disease progression, if the patient agrees, additional optional biopsies can be performed on the lesions in progress. IHC of current FFPE biopsies is used to assess markers including PD-L1, CD3+ T cells, and CD8+ T cells. Other markers may be evaluated, including TIM3, LAG3, and IDO1 expression, as well as cytotoxic cells, NK cells, Th-1 cells, or other immune cell markers in the injected tumor microenvironment. IHC results are compared to baseline.

[0221] Efficacy assessment: throughout the study, RECIST 1.1 is used to assess tumor response. According to RECIST 1.1, although CT or MRI can be used, CT is the preferred imaging technique in this study. During the entire study duration, the same imaging mode must be used for each patient. According to RECIST1.1, digital photos are used to assess the skin melanoma lesions of the melanoma group. Imaging assessment will be performed until PD. The radiological assessment of CR or PR needs to be performed at least 4 weeks after the initial response assessment is observed to be definitive imaging. The assessment of DCR, DOR and PFS will be based on RECIST measurement results. The first study treatment is applied to the time of the first recorded disease progression or death and PFS will be determined. The first observed response to the time of the first recorded disease progression or death will determine DOR. OS will be estimated from the time of the first study treatment application. After the end of the study treatment, the patient will be followed up every 3 months.

[0222] Table 4 summarizes the requirements for tumor assessment at baseline.

[0223] Table 4: Baseline Eligibility Tumor Assessments

[0224]

[0225]

[0226] In the study tumor assessment, target lesions will be evaluated as follows:

[0227] Complete Response (CR): All target lesions disappear. The short axis of any pathological lymph node (whether target or non-target) must be reduced to <10mm. Tumor marker results must normalize.

[0228] Partial Response (PR): The total diameter of the target lesion is reduced by at least 30% based on the baseline total diameter.

[0229] Stable Disease (SD): There was neither sufficient shrinkage to qualify as a PR nor sufficient increase to qualify as a PD, with the smallest (nadir) sum of diameters since the start of treatment as reference.

[0230] Progressive Disease (PD): The sum of the diameters of target lesions increases by at least 20% with the minimum sum (nadir) since the start of treatment as a reference, or one or more new lesions appear. Not only is the increase of 20%, but the absolute increase is also required to exceed the sum by at least 5 mm.

[0231] In the study tumor assessment, non-target lesions were evaluated as follows:

[0232] Complete response (CR): disappearance of all non-target lesions and normalization of tumor markers. The size of all lymph nodes must be non-pathological (<10 mm short axis).

[0233] Stable disease (SD): persistence of one or more non-target lesions and / or persistence of tumor marker levels above normal limits.

[0234] Progressive disease (PD): The appearance of one or more new lesions and / or unequivocal progression of existing non-target lesions. When the patient also has measurable disease, the overall level of non-target disease must have significantly worsened such that the overall tumor burden has increased sufficiently to warrant discontinuation of treatment, even in the presence of SD or PR in target disease.

[0235] Table 5 summarizes the method used to evaluate the best overall response. When nodular disease is included in the sum of target lesions and the nodules are reduced to a "normal" size (<10 mm), they may still have measurements reported in the scan. Even if the nodules are normal, this measurement should be recorded so as not to exaggerate the progression because it is based on the improvement of the nodule size. As mentioned above, this means that the sum of patients with CR may not be "zero". If disease progression is suspected based on clinical or laboratory findings before the planned assessment, an unplanned assessment should be performed. If the patient's health condition deteriorates overall, treatment needs to be stopped, but there is no objective evidence of disease progression at the time, it should be reported as "symptom deterioration". Even after stopping treatment, every effort should be made to record objective progression.

[0236] Table 5: Evaluation of the best overall response

[0237]

[0238] Adverse Event: An AE is any symptom, sign, syndrome, or disease that occurs during the study or, if present at Screening, worsens during the study, regardless of the suspected cause of the event. All medical and psychiatric conditions present at Screening (except those related to the indications under study) will be recorded. Changes in these conditions and new symptoms, signs, syndromes, or diseases should be noted during the remainder of the study. Laboratory, vital sign, and ECG abnormalities should also be recorded as AEs when they are considered clinically significant. AEs may occur spontaneously by the patient, be discovered as a result of general questioning by the study staff, or be determined by physical examination. During each visit to the study clinic, patients will be asked, "Have you had any problems since your last visit?" All AEs will be recorded.

[0239] Main outcome measurement / analysis, tumor response ratio: The primary efficacy endpoint is the ratio of tumor responses (ORR) obtained by RECIST 1.1 (investigator assessment). Unless otherwise specified, tumor response is considered synonymous with success. A confirmed tumor response is defined as CR or PR, recorded as an objective status of two consecutive evaluations at least 4 weeks apart. Tumor response will be evaluated using all treatment cycles. ORR will be calculated by group / subgroup and overall using RECIST 1.1. The proportion of success will be estimated by dividing the number of successes by the total number of evaluable patients in each group. The confidence interval of the true success ratio in each group will be calculated according to the method of Duffy and Santner (Duffy, 1987). If more than the target number of patients are added to the expansion group, the additional patients in that group will not be used for any decision making; however, they will be included in the final endpoint estimates and confidence intervals.

[0240] Rationale: In this study, combination treatment with oncolytic VV1, anti-PD-1 cemiplimab, and anti-CTLA4 monoclonal antibody ipilimumab is expected to enhance infiltration and activation of IT CD8+ T cells to improve treatment response in cancer patients resistant to anti-PD-1 / PD-L1 alone. In this study, conducted in a population of melanoma patients who were refractory to single-agent CPIs, it is expected that cell infection will expand the pool of available endogenous anti-tumor T cells, increase tumor cell antigenicity recognized by T cells, enhance antigen presentation, and ultimately convert a non-inflammatory or immune-rejecting phenotype to a highly inflammatory phenotype.

[0241] Example 2 : A phase 2 trial of Voyager V1 (vesicular stomatitis virus expressing NIS and human interferon beta, VV1) in combination with cemiplimab in patients with selected solid tumors

[0242] This example relates to an open-label, non-randomized Phase 2 study designed to determine the antitumor activity of VV1 in combination with cemiplimab (350 mg IV) and to determine its safety. This study is a variation of the study described in Example 1, in which the hepatocellular, endometrial, and NSCLC cohorts were closed, the melanoma cohort was modified, and the head and neck squamous cell carcinoma (HNSCC) and colorectal cancer (CRC) cohorts were added. Thus, the study described in Example 2 includes melanoma, HNSCC, and CRC cohorts; and the study design is Figure 2 Shown in.

[0243] Prepared (i.e. undiluted) VSV-IFNβ-NIS in a buffer consisting of 5% sucrose, 50 mM Tris (pH 7.4), 2 mM MgCl2. After preparation, the virus was stored frozen at ≤-65°C until ready for use immediately prior to administration. The virus was thawed and diluted in 100 mL of saline containing 1% HSA and administered under close observation.

[0244] On day 1 (D1), VV1 was administered at a dose of 1.0 × 10 9 TCID 50 / mL (total up to 5 mL; up to 5 lesions) in combination with IV cemiplimab (also given on D1) by intratumoral (IT) injection. Data from ongoing trials (VYR-VSV2-101) and investigator-initiated studies suggest that up to 1.7 × 10 11 TCID 50 (IV monotherapy and combination) and 3.0×10 9 TCID 50 The dose (IT in combination) is safe and potentially more effective than lower doses of VV1. Based on experience to date with VV1 and other oncolytic viruses in combination with CPIs, no increase in toxicity of cemiplimab or VV1 is expected.

[0245] Compared with IV VV1, the IT route of VV1 administration offers the advantage of significantly reduced systemic toxicity, with a more favorable safety profile without compromising antitumor activity. In the clinical development program, no sustained high circulating concentrations of virally encoded IFNβ were observed after IT VV1 administration. IT administration of VV1 is better tolerated than IV administration, with fewer systemic symptoms and fewer requirements for pre-medication and post-medication.

[0246] It is expected that the combination therapy of oncolytic VSV-IFNβ-NIS and cemiplimab will enhance the infiltration and activation of intratumoral CD8+T cells to enhance the therapeutic response in cancer patients previously resistant to CPI monotherapy. The trial will include relevant studies monitoring viral PK and pharmacodynamics (PD), including local and systemic immune responses, especially monitoring CD8+T cell responses. These studies will help establish the safety of VSV combined with checkpoint blockade; provide preliminary evidence of clinical efficacy in patients with advanced cancer; and clarify the clinical mechanism of action and biomarkers of response to combined treatment.

[0247] The present study, conducted in a population of melanoma patients who were refractory to single-agent CPIs, was designed to provide insight into the putative mechanisms of resensitization to CPIs by infusion of a single dose of oncolytic virus, hypothesizing that infection of cells would expand the pool of available endogenous antitumor T cells, increase tumor cell antigenicity recognized by T cells, enhance antigen presentation, and ultimately convert a noninflammatory or immune-rejecting phenotype to a highly inflammatory phenotype. Preliminary studies of immune responses in patients treated with VSV-IFNβ-NIS have shown that some patients mount T-cell responses to both viral antigens and known tumor antigens.

[0248] Study Objectives: The primary objective of the study was to determine preliminary antitumor activity in each study cohort, as measured by ORR both in injected lesions and according to standard criteria (RECIST 1.1) [Eisenhauer 2009]. Secondary objectives of the study included: determining the safety and tolerability of IT VV1 in combination with cemiplimab; evaluating the systemic PK of IT VV1 and cemiplimab; studying the PD of IT VV1 in blood by measuring the expression of serum IFNβ; and determining preliminary clinical activity [e.g., PFS, DOR, DCR, and OS] in each cohort.

[0249] Overall study design: Approximately 30 to 87 patients will participate in this study. For the purpose of group expansion, a 30% shrinkage of the injected lesion STL is considered a response in the absence of disease progression of non-injected lesions defined by RECIST 1.1. Patients will be recruited into three parallel double-arm groups, each with an optimal Simon two-stage design; in all groups (melanoma, HNSCC, and CRC), patients will receive the same IT VV1 / IV cimiprilimab dual. After the first cycle, cimiprilimab will be administered on D1 of every 3-week cycle. A second injection of IT VV1 will be performed on D1 of the second cycle, and if feasible, IT VV1 can also be repeated every 3 weeks (every 3week, Q3W) until there is a lack of clinical benefit or limited toxicity. Efficacy evaluation will be performed every 6 weeks (every 6week, Q6W) (at the end of every other cycle).

[0250] Data from an ongoing trial (VYR-VSV2-101) and investigator-initiated studies suggest that up to 1.7 × 10 11 TCID 50 (IV monotherapy and combination) and 3.0×10 9 TCID 50The dose (IT in combination) is safe and potentially more effective than lower doses of VV1 (see Section 1.1.4). The IT route of VV1 administration offers the advantage of significantly reduced systemic toxicity compared to IV VV1, with a more favorable safety profile without compromising anti-tumor activity. In the clinical development program, no sustained high circulating concentrations of virally encoded IFNβ were observed following IT VV1 administration.

[0251] All patients enrolled in this study will receive IT VV1 only and therefore will not require an overnight hospital stay on Day 1 and can be discharged to outpatient follow-up after 6 hours of observation if clinically stable. Patients will then be required to undergo outpatient safety evaluations on Days 2, 3, 4, and 5. Laboratory assessments performed on Days 2, 3, 4, and 5 must be evaluated and possible IFNβ toxicity will be assessed on these days.

[0252] For all cohorts in this study, the optimal Simon two-stage design [Simon 1989] will be used. In the first stage, 10 evaluable patients will be added in each tumor cohort; based on the combination of safety and efficacy, the cohort will advance to the second stage. If there are less than 3 objective responses in the HNSCC cohort, less than 2 responses in the melanoma cohort, or less than 1 response in the CRC cohort among the first 10 patients, the cohort will be stopped; otherwise, in the second stage, 11 to 19 additional patients will be added, for a total of 21 to 29 evaluable patients in each cohort.

[0253] Patients must have evaluable disease according to RECIST 1.1. Patients must consent to provide a fresh biopsy prior to starting study treatment and to repeat biopsies during study treatment, as well as to provide tissue obtained and other blood samples and cheek swabs for biomarker analysis.

[0254] NCI CTCAE, v5.0 will be used to grade overall toxicity, and ASTCT consensus grading will be used to grade CRS and neurotoxicity [Lee 2019]. Safety assessments will include AEs, SAEs, PEs, vital sign measurements, ECOG status, clinical safety laboratory evaluations (hematology, serum chemistry and liver function panels, coagulation, and urinalysis), and ECG.

[0255] For a given patient, the end of their participation generally corresponded to the safety visit conducted 90 days after the last dose of cemiplimab. However, if a patient discontinued treatment for reasons other than PD and continued to have efficacy assessments after the 90-day safety visit, the end of their participation was the date of the last study assessment.

[0256] Inclusion Criteria: Patients who meet all of the following inclusion criteria on Screening / First Dosing Day 1 will be eligible to participate in this study: (1) Age ≥18 years on the day of signing the informed consent. (2) Specificity according to tumor cohort: (a) For the HNSCC cohort, a diagnosis of histologically confirmed advanced and / or metastatic HNSCC suitable for first-line immunotherapy: (i) HPV+ and HPV- patients are allowed; (ii) Primary tumor location in the oropharynx, oral cavity, hypopharynx, or larynx. Participants may not have a primary tumor site in the nasopharynx (any histology); (iii) PD-L1 status ≥10% based on local CPS score. Samples should be provided to a central laboratory for post hoc testing; (iv) at least 12 months have passed between the last dose of prior adjuvant therapy and the date of diagnosis of relapse (if given); (v) no prior anti-PD-(L)1 therapy for HNSCC; (b) for the melanoma cohort, a diagnosis of histologically confirmed advanced and / or metastatic cutaneous melanoma for which no options are considered to provide clinical benefit: (i) best response to uPR, SD, or PD with an anti-PD-(L)1-containing regimen; (ii) prior anti-PD-(L)1 therapy must have lasted ≥12 weeks; (iii) radiological progression was shown during or after treatment with a PD-(L)1 immuno-CPI (only prior first-line PD-(L)1 therapy was allowed); (iv) if the patient received anti-PD-1 as prior adjuvant therapy, the patient should have relapsed during treatment or within the subsequent 6 months after the last dose. Note: Progression on ipilimumab is not required; (v) Patients with BRAF V600-positive tumors should have received prior treatment with a BRAF inhibitor (alone or in combination with a MEK inhibitor) in addition to anti-PD-1 therapy or in addition to having refused targeted therapy. Note: Patients with BRAF V600-positive tumors and no clinically significant tumor-related symptoms or evidence of rapidly progressive disease do not require treatment with a BRAF inhibitor (alone or in combination with a MEK inhibitor) based on the investigator's decision; (c) For the CRC cohort, histologically confirmed diagnosis of advanced and / or metastatic CRC: (i) receiving or ineligible for standard of care fluoropyrimidine, oxaliplatin, irinotecan, anti-VEGF, and EGFR targeted therapy; (ii) non-microsatellite instability-high (non-MSI-high); (iii) progression on prior systemic therapy.

[0257] (3) At least one previously unirradiated tumor lesion eligible for IT injection and biopsy. (a) If the lesion to be injected has not been previously biopsied and confirmed to be malignant, the clinical and / or radiographic features of the lesion must be clearly consistent with malignancy. (4) Measurable disease based on RECIST 1.1, including ≥1 measurable lesion to be injected. (5) Performance status of 0 or 1 on the ECOG performance scale. (6) Life expectancy >3 months. (7) Willingness to provide biological samples required for the duration of the study, including fresh tumor biopsy samples while on study. Note: At baseline, samples may be fresh or archived with no invasive systemic therapy prior to enrollment. Tissue obtained for biopsy must not have been previously irradiated, but new or progressive lesions in the irradiated area are acceptable. (8) Adequate organ function is defined as laboratory values ​​obtained ≤14 days prior to enrollment, as shown in Table 6.

[0258] Table 6

[0259]

[0260] (9) Female patients of childbearing potential with negative pregnancy test. (10) Lack of active CNS involvement. Note: Preregistration imaging is not mandatory for asymptomatic patients. (11) Able to provide written informed consent.

[0261] Exclusion Criteria: Patients who meet any of the following exclusion criteria on Screening / First Dose Day -1 will not be enrolled in this study: (1) Alternative curative treatment options are available and accepted by the patient. (2) Recent or ongoing serious infection, including any active grade 3 or higher viral, bacterial, or fungal infection within 2 weeks of enrollment according to NCI CTCAE, v5.0. (3) Patients diagnosed with ocular, mucosal, or acral melanoma. (4) Known HIV seropositivity and active infection with HIV; (a) Patients who are HIV seropositive but are receiving antiviral therapy and do not show detectable viral load and have a normal CD4 T cell count for at least 6 months are eligible.

[0262] (5) HBV seropositive and with evidence of active HBV viral infection. Patients who are HBsAg negative and HBV viral DNA negative are eligible; (a) Patients who have HBV but have received antiviral treatment and have not shown detectable viral DNA for 6 months are eligible. (b) Patients who are seropositive due to HBV vaccination are eligible. (6) HCV seropositive and HCV active viral infection; (a) Patients who have HCV but have received antiviral treatment and have not shown detectable HCV viral RNA for 6 months are eligible. (7) Known history of active or latent TB. (8) Any concomitant serious health condition that the investigator believes would place the patient at excessive risk from this study, including uncontrolled hypertension and / or diabetes, clinically significant pulmonary disease (e.g., chronic obstructive pulmonary disease requiring hospitalization within 3 months), or neurological disorders (e.g., active seizure disorder within 3 months).

[0263] (9) Prior treatment within the following timeframes prior to the planned start of study treatment: (a) small molecule inhibitors and / or other investigational agents: ≤ 2 weeks or 5 half-lives, whichever is shorter; (b) chemotherapy, other monoclonal antibodies, antibody-drug conjugates, or other similar experimental treatments: ≤ 3 weeks or 5 half-lives, whichever is shorter; (c) radioimmunoconjugates or other similar experimental treatments: ≤ 6 weeks or 5 half-lives, whichever is shorter. (10) NYHA class III or IV, known symptomatic coronary artery disease, or symptoms of coronary artery disease on review of systems, or known arrhythmias (atrial fibrillation or SVT). (11) Any known or suspected active organ-threatening autoimmune disease, such as inflammatory bowel disease, autoimmune hepatitis, lupus, or pneumonia, with the exception of hypothyroidism and type 1 diabetes controlled with therapy. (12) Immunodeficiency or immunosuppression, including systemic corticosteroids >10 mg / day of prednisone or equivalent within 1 week prior to the planned start of study treatment.

[0264] (13) History of grade 3 or 4 immune-mediated adverse reactions to immuno-CPIs. (14) Toxicity of previous therapy that has not resolved to grade 1 or less. (15) History of non-infectious pneumonitis requiring steroids, or current pneumonitis. (16) High disease burden, as assessed clinically by the medical monitor using parameters such as radiographic imprint and tumor markers or LDH. (17) Portal vein thrombosis involving more than the intrahepatic portal vein branches: thrombosis of the right or left portal vein branches or bifurcations, partial or complete occlusion of the main portal vein. (18) Known concurrent malignancy. Exceptions: basal cell carcinoma of the skin, squamous cell carcinoma of the skin, or in situ cervical cancer treated with curative intent, prostate cancer confined to the prostate with a Gleason score <6 or a PSA <1, and any stage I cancer treated with curative intent or any previous cancer with a disease-free interval of ≥3 years. (19) Any other concurrent anticancer treatment (chemotherapy, immunotherapy, radiation therapy, or any adjuvant therapy considered investigational [for non-FDA-approved indications and investigational conditions]).

[0265] (20) Received a live vaccine within 30 days of the planned start of study treatment. Injectable seasonal influenza vaccines are generally inactivated influenza vaccines and are allowed. COVID-19 mRNA vaccines are also allowed according to the following guidelines: Patients who are being screened but have not yet started study treatment should be scheduled for the vaccine to receive the second dose at least 2 weeks before Day 1 (assuming the vaccine is a 2-dose vaccine). If a single-dose vaccine is approved, the single dose should be at least 2 weeks before Day 1. Vaccination during Cycle 1 between the first and second doses of simiprilimab is not encouraged. Starting from Cycle 2, there must be a minimum interval of 7 days from the last treatment to the CoVID-19 vaccination, and patients must have a 7-day interval from vaccination to the next treatment in the study. Efforts should be made to avoid treatment delays, but if necessary, the next treatment with the study drug will be allowed to be delayed by up to 7 days. Intranasal influenza vaccines (e.g., FLU-MIST) are live attenuated vaccines and are not allowed 4 weeks before the first dose of VV1 and 12 weeks after the last dose of VV1.

[0266] (21) Because the genotoxic, mutagenic, and teratogenic effects of the investigational agents involved in this study on the developing fetus and neonate are unknown, any of the following conditions are excluded: (a) Sexually active men or WOCBP who are unwilling to use highly effective contraception before the first dose / first treatment, during the study, and for at least 4 weeks after the last dose*. Highly effective contraceptive measures include: stable use of combined (estrogen and progestin) hormonal contraception (oral, intravaginal, transdermal) or progestin-only hormonal contraception (oral, injectable, implantable) associated with ovulation suppression for 2 or more menstrual cycles prior to screening; intrauterine device (IUD); intrauterine hormone-releasing system (IUS); bilateral tubal ligation; vasectomized partner; and / or sexual abstinence. (i) stable use of combined (estrogen and progestin) hormonal contraception (oral, intravaginal, transdermal) or progestin-only hormonal contraception (oral, injectable, implantable) associated with ovulation suppression for the initiation of 2 or more menstrual cycles prior to screening; (ii) IUD; IUS; (iii) bilateral tubal ligation; (iv) vasectomized partner (provided that the vasectomized male partner is the only sexual partner of the WOCBP study participant and the vasectomized partner has been medically evaluated as a successful surgical procedure); and / or sexual abstinence

[0267] *WOCBP is defined as a woman who is fertile after menarche until she becomes postmenopausal, unless permanently sterilized. Permanent sterilization methods include hysterectomy, bilateral salpingectomy, and bilateral oophorectomy. Postmenopausal women are defined as those who have not had menstruation for 12 months without an alternative medical reason. High follicle-stimulating hormone (FSH) levels in the postmenopausal range can be used to determine postmenopausal status in women who are not using hormonal contraception or hormone replacement therapy. However, in the absence of 12 months of amenorrhea, FSH measurements alone are insufficient to determine the onset of postmenopausal status. Pregnancy testing and contraception are not required for women with a documented hysterectomy or tubal ligation.

[0268] Sexual abstinence is considered a highly effective approach only if it is defined as abstention from heterosexual intercourse during the entire risk period associated with the study drug. The reliability of sexual abstinence needs to be evaluated based on the duration of the clinical trial and the patient's preferences and usual lifestyle.

[0269] Periodic abstinence (calendar method, symptothermal method, postovulation method), withdrawal method (withdrawal), spermicide-only method, and lactational amenorrhea method (LAM) are unacceptable methods of contraception. Female and male condoms should not be used together.

[0270] (22) Pregnant or lactating women. (23) Men who are unwilling to use condoms during vaginal intercourse, while taking the drug, and for 4 weeks after stopping treatment (even if they have undergone a previous vasectomy). (24) Toxicity ≥ Grade 3 from previous anti-PD-1 therapy. (25) Intended injection of lesions involving major blood vessels (e.g., carotid artery, large hepatic vessels). (26) Disease progression within three (3) months of completing systemic treatment with curative intent for locally advanced head and neck cancer.

[0271] Dosage regimen and administration: Cycles are defined as 3 weeks (21 days). Both IT VV1 and cemiplimab are administered on Day 1 of Cycle 1 and Day 1 of Cycle 2 and beyond. VV1 IT injections should continue Q3W, if feasible, and cemiplimab infusions should continue Q3W until limiting toxicity or lack of clinical benefit. End of treatment is defined as the last dose of either study drug.

[0272] The treatment regimen for the combination therapy is shown in Table 7. VV1 dose is based on previous clinical experience and data (see Investigator's Brochure). Cemiprilimab dose is as described in the package insert [ 2020]. Cimiprilimab should be given on day 1 of each 21-day cycle for up to 2 years.

[0273] Table 7: Treatment regimens for VV1 / Cimiprilimab combination

[0274]

[0275] If feasible, VV1 IT injections should continue Q3W, and cemiplimab infusions should continue Q3W until limiting toxicity or lack of clinical benefit. End of treatment was defined as the last dose of either study drug.

[0276] Cimiprilimab administration: Should be administered according to the package insert [ 2020] Prepare Cimiprilimab. Cimiprilimab is administered as an IV infusion over 30 minutes prior to IT VV1 under close observation according to institutional standards of care. Patients should then be observed for 60 minutes for IRR prior to receiving VV1. Cimiprilimab should be given on day 1 of each subsequent 21-day cycle for up to 2 years.

[0277] IT VVI Dosing: VV1 is administered as described above to one or more tumor lesions using a TB syringe with a 20 to 23G needle (or equivalent) on Day 1. The needle type may also be determined by the investigator to allow for optimal distribution of the virus within the lesion.

[0278] VV1 can be administered IT to multiple accessible lesions using the appropriate syringe and needle type for the lesion location. For VV1 treatment, 1 to 5 lesions can be injected; each tumor is treated with at least 1.0 × 10 9 TCID 50 / mL for IT injection, starting from the largest to the smallest, until a maximum volume of 5 mL is reached (a total of 5.0 × 10 9 TCID 50 ), or until all injectable lesions have been treated. If the patient meets retreatment criteria, newly formed lesions should be treated first in the next cycle. In cases where only one or two large lesions are present, more than 1 mL may be administered per tumor at the physician's discretion and with the consent of the medical supervisor.

[0279] Only direct administration into the tumor or accessible lymph nodes is permitted. If injection of multiple lesions is indicated, the lesions should be located in as different locations in the body as possible.

[0280] Once the lesion is identified, the maximum volume to be injected is 5 mL, distributed over the different injectable tumors. Lesions must be greater than 1.0 cm in size at baseline to qualify for IT VV1 administration. The distribution of injection volumes will be determined based on the size of the lesion to be injected, as shown in Table 8.

[0281] Table 8: General guidelines for IT VV1 administration

[0282]

[0283] *Lesions ≤ 1.0 cm may only be injected if they have clinically responded to a previous VV1 injection. The treating physician has the authority to identify and treat the lesion, but this must be documented on the injection worksheet. VV1 is injected at 1.0 × 10 9 TCID 50 A total of 5.0 × 10 9 TCID 50 Each tumor was treated with at least 1.0 × 10 9 TCID 50 / mL for IT injection, injecting 3 to 5 or more lesions (where applicable), starting from the largest to the smallest, until a maximum volume of 5 mL is reached (a total of 5.0 × 10 9 TCID 50 ), or until all injectable lesions are treated. Newly formed lesions should be treated first in the next cycle. In cases where only one or two large lesions are present, more than 1 mL per tumor may be given at the physician's discretion. All concomitant measures to alleviate virus-related systemic symptoms should be initiated prior to IT VV1 administration.

[0284] If the requirements shown in Table 9 are met, patients can continue to receive IT VV1 treatment.

[0285] Table 9. Requirements for continued study drug

[0286]

[0287] *Weeks 3 to 9 = Week 6; Week 12 = Week 10+

[0288] Description of Evaluation: A medical history will be taken during Screening to ensure patient eligibility and will focus on relevant current or past abnormalities or diseases of the following systems: gynecological, cardiovascular, respiratory, gastrointestinal, hepatic, biliary, renal, endocrine / metabolic, musculoskeletal, hematologic / lymphatic, neurologic / psychiatric, cutaneous, immune, infectious diseases, bleeding tendencies, and allergies / drug sensitivities.

[0289] Demographics: Information on date of birth, sex, race / ethnicity, and detailed smoking and alcohol use history will be recorded during screening.

[0290] Physical Examination: A complete physical examination will include examination of the general appearance, skin, head, ears, eyes, nose, throat, heart, lungs, abdomen, lymph nodes, extremities, reproductive system (if specified based on symptomatology and medical history), and neurological system, as well as measurement of weight. A brief (targeted) physical examination should include a cardiopulmonary assessment and abdominal examination, but will focus on new symptoms and will include examination of relevant systems as determined by the investigator. Reporting of all AEs identified as clinically significant changes should be completed. Height and weight will be assessed at Screening, and only weight will be assessed thereafter.

[0291] ECOG performance status and vital signs: ECOG performance status will be recorded based on the investigator's assessment of the patient's performance status. ECOGPS: 0 = fully active, able to perform all pre-disease manifestations without limitation; 1 = limited in vigorous physical activity, but ambulatory and able to perform light or sedentary work, such as light housework, office work; 2 = ambulatory and fully self-care, but unable to perform any work activities; up to 50% and approximately more than 50% of waking time; 3 = limited self-care; confined to bed or chair for more than 50% of waking time; 4 = completely incapacitated; unable to perform any self-care; completely confined to bed or chair; 5 = death. Vital signs will include temperature, respiratory rate, heart rate, and systolic and diastolic blood pressure. Blood pressure and heart rate will be measured oscillometrically using a blood pressure measuring device after 5 minutes in the sitting or semi-recumbent position. The measurement will be repeated if the systolic blood pressure is less than 100 mmHg or greater than 150 mmHg, and / or the diastolic blood pressure is less than 50 mmHg or greater than 90 mmHg. The heart rate measurement will be repeated when it is less than 50 beats per minute (bpm) or greater than 100 bpm. If the measurement is still outside the normal range, the investigator will judge whether the measurement should be repeated. Vital sign measurements outside the normal range will be assessed by the investigator as "abnormal, not clinically significant" or "abnormal, clinically significant". In the latter case, the abnormal vital sign measurement will be reported as an AE and further investigation will be performed as clinically indicated.

[0292] Laboratory Parameters: The following laboratory tests will be performed: serology / virology, urinalysis, electrocardiogram (ECG), drug administration, pharmacokinetic assessment, viral pharmacokinetics, cemiplimab pharmacokinetics, PD assessment, anti-VSV IgG assay, cemiplimab immunogenicity, serum IFN-β and cytokine profiles, peripheral blood mononuclear cells, tumor biopsy, efficacy assessment, and baseline assessments.

[0293] Serologic and virologic measurements should be performed at screening, however, at the discretion of the Medical Monitor, the most recent previously reported results may be used for eligibility purposes. HIV (1 / 2) testing is required if the patient has never been tested for HIV, or if testing occurred before primary cancer diagnosis, or if testing was performed ≥12 months prior to the study. Virologic tests at screening are: HBsAg; HCV RNA; HIV-1 / HIV-2 antibodies. Patients with HIV, HBV, or HCV infection and receiving antiviral therapy must demonstrate no detectable viral RNA / load for 6 months prior to enrollment in this study.

[0294] Standard dipstick urinalysis is adequate for research purposes; however, sites may perform urinalysis according to their typical site practices. Any clinically significant abnormalities should be investigated according to standard medical practice, and any diagnosed AEs should be recorded.

[0295] A single standard 12-lead ECG will be performed at Screening and at one time point after VV1 administration. If the ECG is abnormal, it must be repeated. Thereafter, an ECG should be performed as clinically indicated and in accordance with standard practice. The ECG parameters to be recorded are as follows: rhythm, PR interval, heart rate, QRS interval, QT interval, QTcF interval, and QRS axis. The ECG will be recorded while the patient is resting in the supine or semi-recumbent position. The ECG will be read on-site and classified as "normal," "abnormal, not clinically significant," or "abnormal, clinically significant." If the ECG after study drug administration is determined to be abnormal, clinically significant, the appropriate AE report for the clinically significant finding must be completed.

[0296] Virus PK will be assessed by RT-PCR of VSV-IFNβ-NIS. The first whole blood sample collection (baseline) will be performed at any time within the first 24 hours before VV1 administration. Cimiprili monoclonal antibody PK in serum will be assessed by ELISA. Blood samples will be drawn before dosing (usually within 2 hours before study treatment) and at the end of the first cycle infusion, and the frequency will be reduced thereafter.

[0297] Anti-VSV IgG assays are required to determine the patient's immune response to oncolytic viruses. IFNβ will be evaluated in combination with PK data. Other proinflammatory cytokines and those related to safety can be determined on the same sample, as determined by the safety profile and ongoing findings in the study. Peripheral blood mononuclear cells (PBMCs) will be collected at baseline and at designated time points after treatment for T cell assays, including sequencing.

[0298] Tumor biopsy: At screening, all patients will have a baseline formalin-fixed, paraffin-embedded (FFPE) tissue identified for relevant studies. In addition, if the patient has received systemic therapy since the previous biopsy, a new pre-treatment core biopsy is required (fine needle aspiration is not sufficient). Pre-treatment biopsy tissue cannot be pre-irradiated, but new or progressing lesions in the radiation field are acceptable. This pre-treatment biopsy is optional in Phase 1 of the study, but mandatory in Phase 2. All patients are also required to undergo an on-treatment biopsy of the same lesion according to the assessment schedule. If the patient agrees, an additional optional biopsy can be performed on any other lesion. In the event that the lesion initially biopsied becomes too small to repeat the biopsy, an additional lesion can be biopsied as an alternative. If the previously injected lesion is not suitable for biopsy, this information is recorded. For all cohorts in both Phase 1 and Phase 2, at EOT disease progression, if the patient agrees, an additional optional biopsy can be performed on the progressing lesion. IHC of current FFPE biopsies is used to assess markers including PD-L1, CD3+ T cells, and CD8+ T cells. Additional markers may be evaluated, including TIM3, LAG3, and IDO1 expression, as well as markers of cytotoxic cells, NK cells, Th-1 cells, or other immune cells in the injected tumor microenvironment. IHC results are compared to baseline.

[0299] Efficacy assessment: throughout this study, both the individual measurement results of the shrinkage of the injected lesions and RECIST1.1 will be used to assess tumor response. According to RECIST 1.1, although CT or MRI can be used, CT is the preferred imaging technique in this study. During the entire study duration, the same imaging mode must be used for each patient. According to RECIST 1.1, digital photos are used to assess skin melanoma lesions. Imaging assessment will be performed until PD. The radiological assessment of CR or PR needs to be performed for at least 4 weeks after observing the initial response assessment to perform definitive imaging. RECIST 1.1 will be followed through injection lesion measurement, but only the injected lesions of the target lesion sum will be included. The assessment of DCR, DOR and PFS will be based on RECIST measurement results. The first study treatment is applied to the time of the first recorded disease progression or death will determine PFS. The first observed response to the time of the first recorded disease progression or death will determine DOR. OS will be estimated from the time of the first study treatment application. After the end of the study treatment, the patient will be followed up every 3 months.

[0300] Baseline eligibility tumor assessments will be performed according to Table 10.

[0301] Table 10

[0302]

[0303]

[0304] Tumor assessments in studies including evaluation of target lesions and non-target lesions will be the same as shown in Example 1 and Table 5.

[0305] Concomitant Medication Assessment: Any medications or blood products used within 4 weeks prior to the first dose of study drug and up to 28 days (all concomitant medications) or 90 days (concomitant medications used to treat study drug-related AEs) after the last dose will be recorded, along with the primary reason for their prescription. Any new anticancer therapy started within 90 days of the last dose of study drug should be recorded.

[0306] Primary outcome measurement / analysis, tumor response ratio: The primary efficacy endpoint is the tumor response ratio (ORR) in the injected lesions and overall by RECIST 1.1 (investigator assessment). Unless otherwise specified, tumor response is considered synonymous with success. A confirmed tumor response is defined as CR or PR, recorded as the objective status of two consecutive evaluations at least 4 weeks apart. Tumor response will be evaluated using all treatment cycles. ORR will be calculated by cohort / subgroup and overall using RECIST 1.1. The proportion of success will be estimated by dividing the number of successes by the total number of evaluable patients in each cohort.

[0307] Efficacy parameters will include the following: DCR is defined as the percentage of CR, PR or SD patients for at least 2 consecutive tumor assessments (i.e., CR / PR or SD determined for ≥12 weeks). For all evaluable patients reaching objective response, DOR is defined as the date when the patient's earliest best objective state is first recorded as CR or PR to the earliest date of recording progress. DOR will be estimated using the Kaplan-Meier method. PFS is limited to the time from the first dose of study drug to the earliest date of recording disease progression or death due to any reason. In general, the Kaplan-Meier method [Kaplan and Meier 1958] will be used to estimate the distribution of PFS and for appropriate subgroups and cohorts, RECIST 1.1 will be used, as described above. The percentage of no progression at time points such as 6, 12, 18, 24 months can also be reported. Survival time is defined as the time from the first dose of study drug to death due to any reason. The distribution of survival times will be estimated generally using the method of Kaplan-Meier [Kaplan and Meier 1958] and for appropriate subgroups and cohorts RECIST 1.1 will be used as described above. Survival percentages at 6, 12, 18, 24 months, etc., may also be reported.

[0308] *****

[0309] Sequence Listing: The sequence listing of this application is submitted electronically as an xml file in ST.26 format, with the file name "11163seqlist_ST26", the creation date is November 23, 2022, and the size is 30.8. The submitted sequence listing is part of this specification and is incorporated by reference in its entirety. The length of the following sequences is less than the minimum length allowed under ST.26 format: SEQ ID NO: 7 (A1a Ala Ser) and SEQ ID NO: 19 (AlaAla Ser).

[0310] The present disclosure is not limited in scope by the specific embodiments described herein. In fact, according to the aforementioned description and accompanying drawings, in addition to those described herein, multiple modifications of the present invention will become apparent to those skilled in the art. Such modifications are intended to fall within the scope of the appended claims.

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Claims

1. A method for treating a tumor or inhibiting tumor growth, comprising: (a) Selecting patients with cancer; as well as (b) administering to said patient in need thereof a combination therapy comprising: (i) a therapeutically effective amount of an oncolytic virus; and (ii) a therapeutically effective amount of an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the anti-PD-1 antibody or the antigen-binding fragment thereof comprises three heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1 and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO:

2.

2. The method of claim 1, wherein the oncolytic vesicular virus comprises an oncolytic vesicular stomatitis virus (VSV).

3. The method of claim 2, wherein the VSV comprises a recombinant VSV.

4. The method of claim 3, wherein the recombinant VSV expresses a cytokine.

5. The method of claim 4, wherein the cytokine comprises interferon-β (IFNb).

6. The method of claim 5, wherein the nucleic acid sequence encoding IFNb is located between the M and G viral genes.

7. The method of any one of claims 3 to 6, wherein the recombinant VSV further expresses a sodium / iodide symporter (NIS).

8. The method of claim 7, wherein the nucleic acid sequence encoding NIS is located between the G and L viral genes.

9. The method of any one of claims 1 to 8, wherein the oncolytic virus is Voyager V1.

10. The method of any one of claims 1 to 9, wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NOs: 3, 4, 5, 6, 7, and 8, respectively.

11. The method of any one of claims 1 to 11, wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises a HCVR comprising the amino acid sequence of SEQ ID NO: 1 and a LCVR comprising the amino acid sequence of SEQ ID NO:

2.

12. The method of any one of claims 1 to 11, wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 9 and a light chain comprising the amino acid sequence of SEQ ID NO:

10.

13. The method of any one of claims 1 to 12, wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises cemiplimab or a bioequivalent thereof.

14. The method of any one of claims 1 to 13, wherein the oncolytic virus is administered in the form of one or more 10 to 10 14 TCID 50 A dose of is administered to the patient.

15. The method of claim 14, wherein the oncolytic virus is administered in the form of one or more 10 9 TCID 50 or 10 11 TCID 50 A dose of is administered to the patient.

16. The method of claim 13 or 14, wherein a 10 9 TCID 50 The patient was given an oncolytic virus dose of 10 11 TCID 50 A dose of oncolytic virus is administered to the patient.

17. The method of any one of claims 1 to 16, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at one or more doses of about 0.1 mg / kg to about 20 mg / kg of patient body weight.

18. The method of any one of claims 1 to 17, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at one or more doses of about 1 mg to about 1000 mg.

19. The method of claim 18, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at one or more doses of 350 mg.

20. The method of any one of claims 1 to 19, wherein the oncolytic virus is administered to the patient intratumorally or intravenously.

21. The method of any one of claims 1 to 20, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient intravenously, subcutaneously, or intraperitoneally.

22. The method of any one of claims 1 to 21, wherein the method further comprises administering a therapeutically effective amount of a CTLA-4 inhibitor comprising ipilimumab or a bioequivalent thereof.

23. The method of claim 22, wherein the CTLA-4 inhibitor is ipilimumab.

24. The method of claim 22 or 23, comprising administering one or more doses of the oncolytic virus and one or more doses of the CTLA-4 inhibitor simultaneously.

25. The method of any one of claims 22 to 24, comprising administering one or more doses of the oncolytic virus and one or more doses of the CTLA-4 inhibitor prior to administering one or more doses of the anti-PD-1 antibody or antigen-binding fragment thereof.

26. The method of any one of claims 22 to 25, comprising administering one or more doses of the oncolytic virus and one or more doses of the CTLA-4 inhibitor 7 days prior to administering one or more doses of the anti-PD-1 antibody or antigen-binding fragment thereof.

27.

28. The method of any one of claims 22 to 27, wherein the CTLA-4 inhibitor is administered to the patient at one or more doses of about 0.1 mg / kg to about 15 mg / kg of patient body weight.

29. The method of any one of claims 22 to 28, wherein the CTLA-4 inhibitor is administered to the patient at one or more doses of about 1 mg to about 600 mg.

30. The method of claim 29, wherein the CTLA-4 inhibitor is administered to the patient in one or more doses of 50 mg.

31. The method of any one of claims 22 to 30, wherein the CTLA-4 inhibitor is administered to the patient intravenously, subcutaneously, or intraperitoneally.

32. The method of any one of claims 1 to 29, wherein the cancer is selected from the group consisting of adrenal tumors, bile duct cancer, bladder cancer, brain cancer, breast cancer, epithelial cancer, cancer of central or peripheral nervous system tissue, cervical cancer, colon cancer, endocrine or neuroendocrine cancer or hematopoietic cancer, esophageal cancer, fibroma, gastrointestinal cancer, glioma, head and neck cancer, Ley-Fraumeni tumor, liver cancer, lung cancer, lymphoma, melanoma, meningioma, multiple neuroendocrine neoplasms type I and II, nasopharyngeal cancer, oral cancer, oropharyngeal cancer, osteogenic sarcoma tumors, ovarian cancer, pancreatic cancer, islet cell carcinoma, parathyroid cancer, pheochromocytoma, pituitary tumor, prostate cancer, rectal cancer, kidney cancer, respiratory cancer, sarcoma, skin cancer, stomach cancer, testicular cancer, thyroid cancer, tracheal cancer, genitourinary cancer, and uterine cancer.

33. The method of any one of claims 1 to 30, wherein administration of the combination therapy produces at least one therapeutic effect selected from the group consisting of: a decrease in tumor cell number, tumor regression, a partial response, and a complete response.

34. The method of any one of claims 1 to 33, wherein administration of the combination therapy results in at least one improvement selected from the group consisting of abscopal effect, tumor growth delay, improved overall survival, improved progression-free survival, improved overall response rate, improved complete response, improved partial response, and improved stable disease, compared to a patient treated with a monotherapy selected from the oncolytic virus and the anti-PD-1 antibody, or antigen-binding fragment thereof.

35. The method of claim 34, wherein administration of the combination therapy results in at least one improvement selected from the group consisting of abscopal effects, tumor growth delay, improved overall survival, improved progression-free survival, improved overall response rate, improved complete response, improved partial response, and improved disease stabilization, compared to patients treated with oncolytic virus monotherapy.

36. The method of any one of claims 1 to 35, wherein the tumor growth is inhibited by at least 50% compared to untreated patients.

37. The method of any one of claims 22 to 33, wherein administration of the combination therapy results in at least one improvement selected from the group consisting of abscopal effect, tumor growth delay, improved overall survival, improved progression-free survival, improved overall response rate, improved complete response, improved partial response, and improved stable disease, compared to patients treated with a monotherapy selected from the oncolytic virus, the anti-PD-1 antibody or antigen-binding fragment thereof, and the CTLA-4 inhibitor.

38. The method of claim 37, wherein the tumor growth is inhibited by at least 50% compared to a patient administered the oncolytic virus, the anti-PD-1 antibody or antigen-binding fragment thereof, or the CTLA-4 inhibitor as a monotherapy.

39. The method of claim 37 or 38, wherein the tumor growth is inhibited by at least 50% compared to a patient administered any two of the oncolytic virus, the anti-PD-1 antibody or antigen-binding fragment thereof, and the CTLA-4 inhibitor.

40. The method of any one of claims 1 to 39, further comprising administering to the patient an additional therapeutic agent or treatment.

41. The method of claim 40, wherein the additional therapeutic agent or treatment is selected from the group consisting of radiation, surgery, chemotherapeutic agents, cancer vaccines, B7-H3 inhibitors, B7-H4 inhibitors, lymphocyte activation gene 3 (LAG3) inhibitors, inhibitors of T cell immunoglobulin and mucin domain-3 (TIM3), inhibitors of galectin 9 (GAL9), inhibitors of V-domain immunoglobulin (Ig) inhibitor of T cell activation (VISTA), inhibitors of killer cell immunoglobulin-like receptor (KIR), inhibitors of B and T lymphocyte attenuators (BTLA), inhibitors of T cell immunoreceptors with Ig and ITIM domains (TIGIT) , CD47 inhibitors, indoleamine-2,3-dioxygenase (IDO) inhibitors, vascular endothelial growth factor (VEGF) antagonists, angiopoietin-2 (Ang2) inhibitors, transforming growth factor β (TGFβ) inhibitors, epidermal growth factor receptor (EGFR) inhibitors, antibodies against tumor-specific antigens, BCG, granulocyte-macrophage colony-stimulating factor (GM-CSF), cytotoxins, interleukin 6 receptor (IL-6R) inhibitors, interleukin 4 receptor (IL-4R) inhibitors, IL-10 inhibitors, IL-2, IL-7, IL-12, IL-21, IL-15, antibody-drug conjugates, anti-inflammatory drugs, and combinations thereof.

42. A combination of an oncolytic virus and an antibody or antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the combination is used in a method for treating a tumor or inhibiting tumor growth, the method comprising: (a) Selecting patients with cancer; as well as (b) administering to said patient in need thereof a combination therapy comprising: (i) a therapeutically effective amount of an oncolytic virus; and (ii) a therapeutically effective amount of an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the anti-PD-1 antibody or the antigen-binding fragment thereof comprises three heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1 and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO:

2.

43. A combination of an oncolytic virus, an antibody or antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), and a CTLA-4 inhibitor comprising ipilimumab or a bioequivalent thereof, wherein the combination is used in a method for treating a tumor or inhibiting tumor growth, the method comprising: (a) Selecting patients with cancer; as well as (c) administering to said patient in need thereof a combination therapy comprising: (i) a therapeutically effective amount of an oncolytic virus; (ii) a therapeutically effective amount of an antibody or an antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), wherein the anti-PD-1 antibody or the antigen-binding fragment thereof comprises three heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO: 1 and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO: 2, and (iii) a therapeutically effective amount of a CTLA-4 inhibitor comprising ipilimumab or a bioequivalent thereof.

44. A kit comprising a combination of an oncolytic virus and an antibody or antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1) and the following: written instructions for using a therapeutically effective amount of the combination of the oncolytic virus and the anti-PD-1 antibody or antigen-binding fragment thereof to treat a patient's tumor or inhibit the growth of a patient's tumor.

45. A kit comprising an oncolytic virus, an antibody or antigen-binding fragment thereof that specifically binds to programmed death 1 (PD-1), and a CTLA-4 inhibitor comprising ipilimumab or a bioequivalent thereof, in combination with: written instructions for using a therapeutically effective amount of the oncolytic virus, the anti-PD-1 antibody or antigen-binding fragment thereof, and the CTLA-4 inhibitor in combination to treat a patient's tumor or inhibit the growth of a patient's tumor.

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