Anti-PVRIG antibody preparations and their use

Abstract

We provide a stable liquid pharmaceutical formulation of an antibody containing the anti-human poliovirus receptor-associated immunoglobulin domain (PVRIG). [Solution] A stable liquid pharmaceutical formulation of an anti-PVRIG antibody, wherein (a) the anti-PVRIG antibody comprises i) a heavy chain variable domain including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain of CHA.7.518.1.H4(S241P), and ii) vlCDR1, vlCDR2, and A liquid pharmaceutical formulation is provided, comprising (b) 10 mM to 100 mM histidine, (c) 30 mM to 100 mM NaCl, (d) 20 mM to 150 mM L-arginine, and (e) 0.005% to 0.1 w / v% polysorbate 80, wherein the composition has a pH of 5.5 to 7.0.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority to U.S. Provisional Patent Applications No. 62 / 880,021 filed on 29 July 2019, No. 62 / 893,051 filed on 28 August 2019, No. 62 / 930,206 filed on 4 November 2019, No. 62 / 968,660 filed on 31 January 2020, No. 62 / 985,702 filed on 5 March 2020, and No. 63 / 009,367 filed on 13 April 2020, all of which are incorporated herein by reference in their entirety. [Background technology]

[0002] Naive T cells must receive two independent signals from antigen-presenting cells (APCs) to be productively activated. The first signal, signal 1, is antigen-specific and occurs when the T cell antigen receptor encounters the appropriate antigen-MHC complex on the APC. The fate of the immune response is determined by a second antigen-independent signal (signal 2), delivered via T cell costimulatory molecules that bind to ligands expressed on that APC. This second signal can be either stimulating (positive co-stimulation) or inhibitory (negative co-stimulation or co-inhibition). In the absence of a co-stimulatory signal or in the presence of a co-inhibitory signal, T cell activation is impaired or interrupted, which can lead to a state of antigen-specific unresponsiveness (known as T cell anergy) or T cell apoptosis.

[0003] Costimulatory molecule pairs typically consist of a ligand expressed on APCs and its homogeneous receptor expressed on T cells. Prototype ligand / receptor pairs of costimulatory molecules are B7 / CD28 and CD40 / CD40L. The B7 family consists of structurally related cell surface protein ligands that can provide stimulatory or inhibitory input to the immune response. Members of the B7 family are structurally related, and their extracellular domains contain at least one variable or constant immunoglobulin domain.

[0004] Both positive and negative co-stimulatory signals play essential roles in regulating cell-mediated immune responses, and the molecules that mediate these signals have proven to be effective targets for immunomodulation. Based on this knowledge, several therapeutic approaches involving targeting co-stimulatory molecules have been developed, proving useful in cancer prevention and treatment by turning on or preventing the immune response from being turned off in cancer patients. They have also been shown to be useful in the prevention and treatment of autoimmune and inflammatory diseases, as well as in allograft rejection, by turning off uncontrolled immune responses in subjects with these pathological conditions, or by inducing "off-signals" through negative co-stimulation (or co-inhibition).

[0005] Manipulating signals delivered by B7 ligands has shown promise in the treatment of autoimmune diseases, inflammatory diseases, and graft rejection. Therapeutic strategies include blocking co-stimulation using monoclonal antibodies against the ligand or receptor of the co-stimulatory pair, or using soluble fusion proteins composed of a co-stimulatory receptor that can bind to the appropriate ligand and block it. Another approach is inducing co-inhibition using soluble fusion proteins of the inhibitory ligand. These approaches rely at least in part on the final elimination of autoreactive or alloreactive T cells (which are involved in the pathological processes in autoimmune diseases or transplantation, respectively), probably because T cells become highly susceptible to apoptosis induction in the absence of co-stimulation (which induces cell survival genes). Therefore, disease Novel drugs that can modulate co-stimulatory signals without impairing the immune system's ability to defend against pathogens would be highly advantageous for the treatment and prevention of such pathological conditions.

[0006] Co-stimulatory pathways play a crucial role in tumor development. Interestingly, tumors have been shown to evade immune destruction by inhibiting T cell activation through the inhibition of costimulatory factors in the B7-CD28 and TNF family, as well as by inhibiting the antitumor T cell response by inducing regulatory T cells (Wang (2006), “Immune Suppression by Tumor Specific CD4 +Regulatory T cells in Cancer”,Semin.Cancer.Biol.16:73-79,Greenwald,et al.(2005),“The B7 Family Revisited”,Ann.Rev.Immunol.23:515-48;Watts(2005),“TNF / TNFR Family Members in Co-stimulation of T Cell Responses”, Ann.Rev.Immunol.23:23-68; Sadum, et al., (2007) “Immune Signatures of Murine and Human Cancers Reveal Unique Mechanisms of Tumor Escape and New Targets for Cancer See “Immunotherapy”, Clin.Canc.Res.13(13):4016-4025. Costimulatory molecules expressed in such tumors have become attractive cancer biomarkers and may serve as tumor-associated antigens (TAAs). Furthermore, costimulatory pathways have been identified as immune checkpoints that attenuate T-cell-dependent immune responses at both the initiation and effector function levels within tumor metastases. As engineered cancer vaccines continue to improve, it is becoming clear that such immune checkpoints are major barriers to the ability of vaccines to induce therapeutic antitumor responses. In this regard, costimulatory molecules can serve as immunostimulants for active (vaccination) and passive (antibody-mediated) cancer immunotherapies, providing strategies to inhibit immune tolerance and stimulate the immune system.

[0007] Over the past decade, various co-stimulatory protein activators and / or antagonists have been developed for the treatment of autoimmune diseases, graft rejection, allergies, and cancer. For example, CTLA4-Ig (Abatacept, Orencia®) is approved for the treatment of rheumatoid arthritis (RA), mutant CTLA4-Ig (Belatacept, Nulojix®) is approved for the prevention of acute kidney transplant rejection, and an anti-CTLA4 antibody (Ipilimumab, Yervoy®) has recently been approved for the treatment of melanoma. Other co-stimulatory modulogenators, such as Merck (Keytruda®) and BMS (Opdivo®) anti-PD-1 antibodies, have been approved for cancer treatment and are also in the testing phase for viral infections.

[0008] The specific area of ​​interest is PVRIG. PVRIG is a 326-amino acid transmembrane domain protein that contains a signal peptide (ranging from amino acids 1 to 40), an extracellular domain (ranging from amino acids 41 to 171), a transmembrane domain (ranging from amino acids 172 to 190), and a cytoplasmic domain (ranging from amino acids 191 to 326). The full-length human PVRIG protein is shown in Figure 1. There are two methionine molecules that could be start codons, but the mature protein is identical.

[0009] The PVRIG protein contains an immunoglobulin (Ig) domain within its extracellular domain, which is a PVR-like Ig folding domain. Due to its similarity to other B7 family members, the PVR-like Ig folding domain may be involved in functional counterpart binding. The PVR-like Ig folding domain in the extracellular domain has a single disulfide formed between intradomain cysteine ​​residues. It contains phytobonds, which are typical of this folding and may be important to its structure and function. These cysteines are located at residues 22 and 93 (or 94). In one embodiment, a PVRIG soluble fragment is provided that can be used for testing PVRIG antibodies. The definition of the PVRIG protein includes PVRIG ECD fragments, including known ECD fragments such as those described in U.S. Patent No. 9,714,289.

[0010] PVRIG has also been identified as an inhibitory receptor that recognizes CD112 rather than CD155, and may be involved in the negative regulation of DNAM-1-mediated antitumor function. PVRL2 has been identified as a ligand for PVRIG, aligning PVRIG with the DNAM / TIGIT immune receptor axis (Liang et al., Journal of (See Clinical Oncology 2017 35:15_suppl,3074-3074).

[0011] Anti-PVRIG antibodies (including antigen-binding fragments) that both bind to PVRIG and prevent its activation by PVLR2 (for example, most commonly by blocking the interaction between PVRIG and PVLR2) are used to enhance T cell and / or NK cell activation and to treat diseases such as cancer and pathogen infections. Therefore, formulations for administering such antibodies are necessary.

[0012] Therefore, an object of the present invention is to provide a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody, or to use it for the treatment of a disease (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3). [Overview of the Initiative]

[0013] Therefore, the object of the present invention is to provide a stable liquid pharmaceutical formulation of the anti-PVRIG antibody described herein.

[0014] The present invention (a) An anti-PVRIG antibody, wherein the anti-PVRIG antibody i) a heavy chain variable domain comprising vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), and ii) a light chain variable domain comprising vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody, (b) 10 mM to 100 mM histidine, and (c) 30 mM to 100 mM NaCl, and (d) 20 mM to 150 mM L-arginine, and (e) 0.005% to 0.1 w / v% polysorbate 80, and provide a stable liquid pharmaceutical formulation of the anti-PVRIG antibody, The composition has a pH of 5.5 to 7.0.

[0015] In some embodiments of the stable liquid pharmaceutical formulation, the anti-PVRIG antibody comprises the CH1-hinge-CH2-CH3 sequence (SEQ ID NO: 17 or SEQ ID NO: 50) of IgG4, and the hinge region optionally contains mutations.

[0016] In some embodiments of the stable liquid pharmaceutical formulation, the anti-PVRIG antibody comprises the CH1-hinge-CH2-CH3 region from IgG1, IgG2, IgG3, or IgG4, and the hinge region optionally contains mutations.

[0017] In some embodiments of the stable liquid pharmaceutical formulation, the heavy chain variable domain is from the heavy chain (SEQ ID NO: 4) of CHA .7.518.1.H4(S241P), and the light chain variable domain is from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P).

[0018] In some embodiments of the stable liquid pharmaceutical formulation, the anti-PVRIG antibody comprises the CL region of the human kappa 2 light chain.

[0019] In some embodiments of the stable liquid pharmaceutical preparation, the pharmaceutical preparation comprises histidine at 10 mM to 80 mM, 15 mM to 70 mM, 20 mM to 60 mM, 20 mM to 50 mM, or 20 mM to 30 mM.

[0020] In some embodiments of the stable liquid pharmaceutical preparation, the pharmaceutical preparation comprises about 25 mM of histidine.

[0021] In some embodiments of the stable liquid pharmaceutical preparation, the pharmaceutical preparation comprises NaCl at 30 mM to 100 mM, 30 mM to 90 mM, 40 mM to 80 mM, 30 mM to 70 mM, or 45 mM to 70 mM.

[0022] In some embodiments of the stable liquid pharmaceutical preparation, the pharmaceutical preparation comprises about 60 mM of NaCl.

[0023] In some embodiments of the stable liquid pharmaceutical preparation, the pharmaceutical preparation comprises L-arginine at 20 mM to 140 mM, 30 mM to 140 mM, 40 mM to 130 mM, 50 mM to 120 mM, 60 mM to 110 mM, 70 mM to 110 mM, 80 mM to 110 mM, or 90 mM to 110 mM.

[0024] In some embodiments of the stable liquid pharmaceutical preparation, the pharmaceutical preparation comprises about 100 mM of L-arginine.

[0025] In some embodiments of a stable liquid pharmaceutical formulation, the pharmaceutical formulation comprises 0.006% to 0.1 w / v% polysorbate 80, 0.007% to 0.09 w / v% polysorbate 80, 0.008% to 0.08 w / v% polysorbate 80, 0.009% to 0.09 w / v% polysorbate 80, 0.01% to 0.08 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, or 0.01% to 0.06 w / v% polysorbate 80, or 0.009% to 0.05% w / v polysorbate 80.

[0026] In some embodiments of the stable liquid pharmaceutical formulation, the pharmaceutical formulation contains about 0.01% polysorbate 80.

[0027] In some embodiments of stable liquid pharmaceutical formulations, the pH is 6 to 7.0.

[0028] In some embodiments of stable liquid pharmaceutical formulations, the pH is 6.3 to 6.8.

[0029] In some embodiments of stable liquid pharmaceutical formulations, the pH is 6.5 ± 0.2.

[0030] In some embodiments of a stable liquid pharmaceutical formulation, the anti-PVRIG antibody is 10m The concentrations are in the ranges of g / mL to 40 mg / mL, 15 mg / mL to 40 mg / mL, 15 mg / mL to 30 mg / mL, 10 mg / mL to 25 mg / mL, or 15 mg / mL to 25 mg / mL.

[0031] In some embodiments of a stable liquid pharmaceutical formulation, the formulation is stable at 2°C to 8°C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 weeks.

[0032] In some embodiments of the stable liquid pharmaceutical formulation, the formulation is stable at approximately 20°C to 25°C for at least one, two, three, four, five, or six weeks.

[0033] In some embodiments of the stable liquid pharmaceutical formulation, the formulation is stable at 35°C to 40°C for at least one, two, three, four, or five weeks.

[0034] In some embodiments of the stable liquid pharmaceutical formulation, the anti-PVRIG antibody is present at a concentration of approximately 20 mg / mL.

[0035] In some embodiments of a stable liquid pharmaceutical formulation, the anti-PVRIG antibody formulation is: a) A heavy chain, i) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, b) Light chain, i) A VL-CL comprising a light chain, wherein the VL is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is from the human kappa 2 light chain.

[0036] In some embodiments of the stable liquid pharmaceutical formulation, the hinge region optionally includes mutations.

[0037] In some embodiments of the stable liquid pharmaceutical formulation, the hinge region optionally includes mutations.

[0038] In some embodiments of a stable liquid pharmaceutical formulation, the anti-PVRIG antibody formulation is: i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P).

[0039] In some embodiments of a stable liquid pharmaceutical formulation, the anti-PVRIG antibody formulation is: (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0040] In some embodiments of a stable liquid pharmaceutical formulation, the anti-PVRIG antibody formulation is: (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0041] In some embodiments of the stable liquid pharmaceutical formulation, the formulation is administered at a dose of anti-PVRIG antibody ranging from approximately 0.01 mg / kg to approximately 20 mg / kg. In some embodiments of the stable liquid pharmaceutical formulation, the formulation is administered at a dose of anti-PVRIG antibody ranging from approximately 0.01 mg / kg to approximately 10 mg / kg.

[0042] In some embodiments of the stable liquid pharmaceutical formulation, the formulation is administered at doses of anti-PVRIG antibody of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg.

[0043] In some embodiments of the stable liquid pharmaceutical formulation, the anti-PVRIG antibody is administered at a dose of 20 mg / kg every four weeks. In some embodiments of the stable liquid pharmaceutical formulation, the anti-PVRIG antibody is administered intravenously at a dose of 20 mg / kg every four weeks.

[0044] In some embodiments of the treatment method, the formulation is administered intravenously at a dose of 20 mg / kg every four weeks for up to 24 months until disease progression, unacceptable toxicity, initiation of a new anticancer therapy, withdrawal of consent from the subject, or death. In some embodiments, administration is for up to 6 months, 12 months, 18 months, or 24 months until disease progression, unacceptable toxicity, initiation of a new anticancer therapy, withdrawal of consent from the subject, and / or death.

[0045] In some embodiments of a stable liquid pharmaceutical formulation, the stable liquid pharmaceutical formulation is administered for the treatment of cancer.

[0046] In some embodiments of a stable liquid pharmaceutical formulation, the stable liquid pharmaceutical formulation is intended for use in a method of treating cancer.

[0047] In some embodiments of the stable liquid pharmaceutical formulation, the cancers include prostate cancer, liver cancer (HCC), colorectal cancer (CRC), colorectal cancer MSS (MSS-CRC; including refractory MSS colorectal cancer), CRC (MSS of unknown origin), ovarian cancer (including ovarian carcinoma), endometrial cancer (including endometrial carcinoma), breast cancer, pancreatic cancer, gastric cancer, cervical cancer, head and neck cancer, thyroid cancer, testicular cancer, urothelial carcinoma, lung cancer, melanoma, and non-melanoma skin cancer (squamous). Epithelial and basal cell carcinomas), gliomas, renal cell carcinoma (RCC), renal cell carcinoma (RCC), lymphomas (non-Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HD)), acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia (T-ALL), diffuse large B-cell lymphoma, testicular germ cell tumors, mesothelioma, esophageal cancer, trinegative breast cancer, Merkel cell carcinoma, MSI high-grade cancers, KRAS variant tumors, adult T-cell leukemia / lymphoma, pleural mesothelioma The group consists of anal SCC, neuroendocrine lung cancer (including neuroendocrine lung carcinoma), NSCLC, NSCL (large cell), NSCLC large cell, NSCLC squamous cell, cervical SCC, malignant melanoma, pancreatic cancer, pancreatic adenocarcinoma, adenoid cystic carcinoma (including adenoid cystic carcinoma), primary peritoneal cancer, microsatellite-stabilized primary peritoneal cancer, platinum-resistant microsatellite-stabilized primary peritoneal cancer, and / or myelodysplastic syndrome (MDS).

[0048] This invention provides the use of a stable liquid pharmaceutical formulation of anti-PVRIG antibodies in a method for treating cancer, wherein the anti-PVRIG antibody is (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0049] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraph

[0048] , the anti-PVRIG antibody comprises a CH1-hinge-CH2-CH3 sequence of IgG4 (SEQ ID NO: 17 or SEQ ID NO: 50), wherein the hinge region optionally contains mutations.

[0050] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0049] , the anti-PVRIG antibody comprises a CH1-hinge-CH2-CH3 region from IgG1, IgG2, IgG3, or IgG4, wherein the hinge region optionally contains mutations.

[0051] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0050] , the heavy chain variable domain is derived from the heavy chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 4), and the light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9).

[0052] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0051] , the anti-PVRIG antibody comprises the CL region of the human kappa 2 light chain.

[0053] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0052] , the pharmaceutical formulation comprises 10 mM to 80 mM histidine, 15 mM to 70 mM histidine, 20 mM to 60 mM histidine, 20 mM to 50 mM histidine, or 20 mM to 30 mM histidine.

[0054] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0053] , the pharmaceutical formulation contains about 25 mM histidine.

[0055] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0054] , the pharmaceutical formulation comprises 30 mM to 100 mM NaCl, 30 mM to 90 mM NaCl, 40 mM to 80 mM NaCl, 30 mM to 70 mM histidine, or 45 mM to 70 mM NaCl.

[0056] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0055] , the pharmaceutical formulation contains about 60 mM NaCl.

[0057] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0056] , the pharmaceutical formulation comprises 20 mM to 140 mM L-arginine, 30 mM to 140 mM L-arginine, 40 mM to 130 mM L-arginine, 50 mM to 120 mM L-arginine, 60 mM to 110 mM L-arginine, 70 mM to 110 mM L-arginine, 80 mM to 110 mM L-arginine, or 90 mM to 110 mM L-arginine.

[0058] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0057] , the pharmaceutical formulation contains about 100 mM L-arginine.

[0059] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in a method for treating cancer described in paragraphs

[0048] to

[0058] , the pharmaceutical formulation comprises 0.006% to 0.1 w / v% polysorbate 80, 0.007% to 0.09 w / v% polysorbate 80, 0.008% to 0.08 w / v% polysorbate 80, 0.009% to 0.09 w / v% polysorbate 80, 0.01% to 0.08 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, or 0.01% to 0.06 w / v% polysorbate 80, or 0.009% to 0.05% w / v polysorbate 80.

[0060] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0059] , the pharmaceutical formulation contains about 0.01% polysorbate 80.

[0061] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0060] , the pH is 6 to 7.0.

[0062] In some embodiments of the methods for treating cancer described in paragraphs

[0048] to

[0061] , where a stable liquid pharmaceutical formulation of an anti-PVRIG antibody is used, the pH is 6.3 to 6.8.

[0063] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0062] , the pH is 6.5+ / -0.2.

[0064] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0063] , the anti-PVRIG antibody is present in concentrations of 10 mg / mL to 40 mg / mL, 15 mg / mL to 40 mg / mL, and 15 mg / mL. The concentrations are between 10 mg / mL and 30 mg / mL, 10 mg / mL and 25 mg / mL, or 15 mg / mL and 25 mg / mL.

[0065] In some embodiments using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0064] , the formulation is stable at 2°C to 8°C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 weeks.

[0066] In some embodiments using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0065] , the formulation is stable at about 20°C to 25°C for at least 1, 2, 3, 4, 5, or 6 weeks.

[0067] In some embodiments using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0066] , the formulation is stable at 35°C to 40°C for at least one, two, three, four, or five weeks.

[0068] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0067] , the anti-PVRIG antibody is at a concentration of about 20 mg / mL.

[0069] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in a method for treating cancer described in paragraphs

[0048] to

[0068] , the anti-PVRIG antibody formulation is: a) A heavy chain, i) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, b) Light chain, i) A VL-CL comprising a light chain, wherein the VL is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is from the human kappa 2 light chain.

[0070] In some embodiments of the methods for treating cancer described in paragraphs

[0048] to

[0069] , which use a stable liquid pharmaceutical formulation of an anti-PVRIG antibody, the hinge region optionally includes mutations.

[0071] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0070] , the hinge region optionally includes mutations.

[0072] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in a method for treating cancer described in paragraphs

[0048] to

[0071] , the anti-PVRIG antibody formulation is: i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P).

[0073] In the method for treating cancer described in paragraphs

[0048] to

[0072] , anti-PVRIG anti In some embodiments using a stable liquid pharmaceutical formulation, the anti-PVRIG antibody formulation is, (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0074] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in a method for treating cancer described in paragraphs

[0048] to

[0073] , the anti-PVRIG antibody formulation is: (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0075] In some embodiments of the method for treating cancer described in paragraphs

[0048] to

[0074] , which uses a stable liquid pharmaceutical formulation of anti-PVRIG antibody, the formulation is administered in doses of anti-PVRIG antibody ranging from about 0.01 mg / kg to about 20 mg / kg. In some embodiments of the stable liquid pharmaceutical formulation, the formulation is administered in doses of anti-PVRIG antibody ranging from about 0.01 mg / kg to about 10 mg / kg.

[0076] In some embodiments of using a stable liquid pharmaceutical formulation of anti-PVRIG antibody in the method for treating cancer described in paragraphs

[0048] to

[0075] , the formulation is administered in doses of anti-PVRIG antibody of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg.

[0077] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in a method of treating cancer, the anti-PVRIG antibody is administered at a dose of 20 mg / kg every four weeks. In some embodiments of use in the methods for treatment described in paragraphs

[0048] to

[0076] , the formulation is administered intravenously at a dose of 20 mg / kg every four weeks.

[0078] In some embodiments of the treatment methods described in paragraphs

[0048] to

[0077] , the formulation is administered intravenously at a dose of 20 mg / kg every four weeks for up to 24 months until disease progression, unacceptable toxicity, initiation of a new anticancer therapy, withdrawal of consent from the subject, or death. In some embodiments, administration is for up to 6 months, 12 months, or 18 months until disease progression, unacceptable toxicity, initiation of a new anticancer therapy, withdrawal of consent from the subject, and / or death. , or 24 months.

[0079] In some embodiments of using a stable liquid pharmaceutical formulation of an anti-PVRIG antibody in a method for treating cancer described in paragraphs

[0048] to

[0078] , the cancer is prostate cancer, liver cancer (HCC), colorectal cancer (CRC), colorectal cancer MSS (MSS-CRC; including refractory MSS colorectal cancer), CRC (MSS unknown), ovarian cancer (including ovarian carcinoma), endometrial cancer (including endometrial carcinoma), breast cancer, pancreatic cancer, gastric cancer, cervical cancer, head and neck cancer, thyroid cancer, testicular cancer, urothelial carcinoma, lung cancer, melanoma, non-melanoma skin cancer (squamous and basal cell carcinoma), glioma, renal cell carcinoma (RCC), renal cell carcinoma (RCC), lymphoma (non-Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HD)), The group is selected from acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia (T-ALL), diffuse large B-cell lymphoma, testicular germ cell tumor, mesothelioma, esophageal cancer, trinegative breast cancer, Merkel cell carcinoma, MSI hypercarcinoma, KRAS variant tumor, adult T-cell leukemia / lymphoma, pleural mesothelioma, anal SCC, neuroendocrine lung cancer (including neuroendocrine lung carcinoma), NSCLC, NSCL (large cell), NSCLC large cell, NSCLC squamous cell, cervical SCC, malignant melanoma, pancreatic cancer, pancreatic adenocarcinoma, adenoid cystic carcinoma (including adenoid cystic carcinoma), primary peritoneal cancer, microsatellite-stable primary peritoneal cancer, platinum-resistant microsatellite-stable primary peritoneal cancer, and / or myelodysplastic syndrome (MDS). [Brief explanation of the drawing]

[0080] [Figure 1] The full-length sequence of human PVRIG is shown. [Figure 2] The sequence of the human poliovirus receptor-associated protein 2 (PVLR2, also known as nectin-2, CD112, or herpesvirus entry mediator B (HVEB)), which is the binding partner of PVRIG, is illustrated. PVLR2 is a human plasma membrane glycoprotein. [Figure 3]The variable heavy and light chains of CHA.7.518.1.H4(S241P) of the present invention, as well as the sequences of vhCDR1, vhCDR2, vhCDR3, vlCDR1, vlCDR2, and vlCDR3, are shown in the diagram. [Figure 4] The sequences of human IgG1, IgG2, IgG3, and IgG4 are shown in the diagram. [Figures 5A-5D] The sequences of other PVRIG antibodies that can be formulated using the stable liquid formulation of the anti-PVRIG antibody of the present invention are shown. [Figure 6] The formulation parameters, including the buffer and excipients, of the CHA.7.518.1.H4(S241P) antibody preparation are illustrated. [Figures 7A-7B] The data for CHA.7.518.1.H4(S241P) is shown in the following diagrams: A) Stress and storage conditions are shown in the diagram. B) Samples and test schedule are shown. [Figure 8] This provides the visual appearance results for the CHA.7.518.1.H4(S241P) formulation at T=0. [Figure 9] Provides results for A280 at T=0: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 10] Provides LabChip results for T=0: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 11] Provides SEC results at T=0: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 12] Provides cIEF results at T=0: Determination of isoelectric point using imaging cIEF analysis of CHA.7.518.1.H4(S241P) formulation. [Figure 13] Provides MFI results at T=0: Particle analysis of formulation CHA.7.518.1.H4(S241P) by microfluidic imaging (MFI). [Figure 14]This provides a binding assay with T=0: a potency ELISA for evaluating the CHA.7.518.1.H4(S241P) formulation. [Figure 15] This provides the results of the visual appearance of the CHA.7.518.1.H4(S241P) formulation after freezing / thawing. [Figure 16] Provides results for freeze / thaw A280: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 17] Provides LabChip results for freeze / thaw: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 18] Provides SEC results for freeze / thaw: CHA.7.518.1.H4(S241P) size exclusion chromatography for determining protein aggregation of formulations. [Figure 19] Provides freeze / thaw cIEF results: CHA.7.518.1.H4(S241P) Determination of isoelectric point using imaging cIEF analysis of formulations. [Figure 20] Provides MFI results for freeze / thaw: Particle analysis of formulations by microfluidic imaging (MFI) of CHA.7.518.1.H4(S241P). [Figure 21] Provides a freeze / thaw binding assay: Efficacy ELISA for evaluating CHA.7.518.1.H4(S241P) formulations. [Figure 22] This provides the visual appearance results of the stirring of the CHA.7.518.1.H4(S241P) formulation. [Figure 23] A280 results of agitation are provided: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 24] Provides LabChip results for agitation: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 25]Provides the results of SEC with agitation: CHA.7.518.1.H4(S241P) Size exclusion chromatography for determining protein aggregation of formulations. [Figure 26] Provides cIEF results for agitation: CHA.7.518.1.H4(S241P) Determination of isoelectric point using imaging cIEF analysis of formulations. [Figure 27] Provides results of MFI with agitation: Particle analysis of formulations by microfluidic imaging CHA.7.518.1.H4(S241P). [Figure 28] Provides a stirring-bound assay: Efficacy ELISA for evaluating CHA.7.518.1.H4(S241P) formulations. [Figure 29] This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation after 1 week at 40°C. [Figure 30] Results of A280 at 40°C for 1 week: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 31] Provides LabChip results for 1 week at 40°C: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 32] Provides results of SEC at 40°C for one week: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 33] Provides results for cIEF at 40°C for one week: Determination of isoelectric point using imaging cIEF analysis of CHA.7.518.1.H4(S241P) formulation. [Figure 34] We provide results of microfluidic imaging of the CHA.7.518.1.H4(S241P) formulation at 40°C for one week: Particle analysis by microfluidic imaging. [Figure 35] Provides a binding assay at 40°C for one week: potency ELISA for evaluating CHA.7.518.1.H4(S241P). [Figure 36]This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation after 2 weeks at 40°C. [Figure 37] Results for A280 at 40°C for 2 weeks: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 38] Provides LabChip results for 2 weeks at 40°C: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 39] SEC results at 40°C for 2 weeks: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 40] cIEF results at 40°C for 2 weeks: Determination of the isoelectric point of CHA.7.518.1.H4(S241P) formulation using imaging cIEF analysis. [Figure 41] We provide MFI results from 2 weeks at 40°C: Particle analysis of CHA.7.518.1.H4(S241P) formulation by microfluidic imaging. [Figure 42] This provides a binding assay at 40°C for two weeks: an efficacy ELISA for evaluating CHA.7.518.1.H4(S241P) formulations. [Figure 43] This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation at ambient temperature for two weeks. [Figure 44] The results for A280 over two weeks at ambient temperature are provided: Determination of protein concentration using SoloVPE in the CHA.7.518.1.H4(S241P) formulation. [Figure 45] Provides LabChip results for 2 weeks at ambient temperature: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 46]Provides SEC results for 2 weeks at ambient temperature: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 47] Provides cIEF results for 2 weeks at ambient temperature: Determination of the isoelectric point of CHA.7.518.1.H4(S241P) formulation using imaging cIEF analysis. [Figure 48] Provides MFI results for 2 weeks at ambient temperature: Particle analysis of CHA.7.518.1.H4(S241P) formulation by microfluidic imaging. [Figure 49] Provides a 2-week binding assay at ambient temperature: Efficacy ELISA for evaluating CHA.7.518.1.H4(S241P) formulations. [Figure 50] This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation at ambient temperature for 4 weeks. [Figure 51] This provides results for A280 over 4 weeks at ambient temperature: determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 52] Provides LabChip results for 4 weeks at ambient temperature: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 53] Provides SEC results for 4 weeks at ambient temperature: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 54] Provides cIEF results for 4 weeks at ambient temperature: Determination of isoelectric point using imaging cIEF analysis of CHA.7.518.1.H4(S241P) formulation. [Figure 55] Provides MFI results for 4 weeks at ambient temperature: Particle analysis of CHA.7.518.1.H4(S241P) formulation by microfluidic imaging. [Figure 56]Provides a 4-week binding assay at ambient temperature: Efficacy ELISA for evaluating the CHA.7.518.1.H4(S241P) formulation. [Figure 57] This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation after 2 weeks at 2-8°C. [Figure 58] This provides results for A280 at 2-8°C for 2 weeks: determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 59] Provides LabChip results for 2 weeks at 2-8°C: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 60] SEC results are provided for 2 weeks at 2-8°C: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 61] Provides cIEF results for 2 weeks at 2-8°C: Determination of CHA.7.518.1.H4(S241P) isoelectric point using imaging cIEF analysis. [Figure 62] This provides MFI results for 2 weeks at 2-8°C: Particle analysis of CHA.7.518.1.H4(S241P) formulation by microfluidic imaging. [Figure 63] This provides a binding assay for 2 weeks at 2-8°C: Efficacy ELISA for evaluating CHA.7.518.1.H4(S241P) formulations. [Figure 64] This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation at 2-8°C for 4 weeks. [Figure 65] This provides results for A280 over 4 weeks at 2-8°C: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 66] Provides LabChip results for 4 weeks at 2-8°C: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 67]SEC results for 4 weeks at 2-8°C: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 68] Provides cIEF results for 4 weeks at 2-8°C: Determination of isoelectric point using imaging cIEF analysis of CHA.7.518.1.H4(S241P) formulation. [Figure 69] This provides MFI results for 4 weeks at 2-8°C: Particle analysis of CHA.7.518.1.H4(S241P) formulation by microfluidic imaging. [Figure 70] This provides a binding assay for 4 weeks at 2-8°C: an efficacy ELISA for evaluating the CHA.7.518.1.H4(S241P) formulation. [Figure 71] This provides the visual appearance results of the CHA.7.518.1.H4(S241P) formulation at 2-8°C for 8 weeks. [Figure 72] Results for A280 at 2-8°C for 8 weeks: Determination of protein concentration using SoloVPE in CHA.7.518.1.H4(S241P) formulation. [Figure 73] Provides LabChip results for 8 weeks at 2-8°C: Determination of IgG purity of CHA.7.518.1.H4(S241P) formulation by LabChip (reduced and unreduced). [Figure 74] SEC results for 8 weeks at 2-8°C: Size exclusion chromatography for determining protein aggregation of CHA.7.518.1.H4(S241P) formulation. [Figure 75] Provides cIEF results for 8 weeks at 2-8°C: Isoelectric point determination using imaging cIEF analysis of CHA.7.518.1.H4(S241P) formulation. [Figure 76] This provides MFI results for 8 weeks at 2-8°C: Particle analysis of CHA.7.518.1.H4(S241P) formulation by microfluidic imaging. [Figure 77] This provides an 8-week binding assay at 2-8°C: an efficacy ELISA for evaluating the CHA.7.518.1.H4(S241P) formulation. [Figure 78]This provides data showing receptor occupancy at various doses of CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8, light chain: SEQ ID NO: 13). [Figure 79] This provides data showing receptor occupancy at various doses of CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8, light chain: SEQ ID NO: 13). [Figure 80] This provides data indicating that PVRIG is a novel checkpoint for TIGIT / DNAM-1 AXIS. [Figure 81] This provides data demonstrating that PVRIG inhibition reduces tumor growth in a mouse cancer model. [Figure 82] A schematic diagram of the research design is provided. [Figure 83] Provides information about the patient's baseline characteristics. [Figure 84] We provide information regarding the breakdown of treatments received by patients. [Figure 85] Provides information on the treatment of urgent adverse events. [Figure 86] Provides information on the treatment of urgent serious adverse events. [Figure 87] Provides a swimmer plot of patient data. [Figure 88] Provides a waterfall plot of patient data. [Figure 89] This provides information on patients with stable conditions and the relationship between dose and response. [Figure 90] This provides information on the optimal timing of treatment response for patients with treatment-resistant diseases. [Figure 91] A graph of therapeutic dosage data is provided. [Figure 92] This paper provides data on PVRIG binding by anti-PVRIG using a receptor occupancy assay. [Figure 93] Provides baseline characteristic data for patients. [Figure 94] Provides a summary of the patient breakdown. [Figure 95] This shows a dose escalation scheme. [Figure 96] A safety analysis set that provides an overview of adverse events. [Figure 97] Provide an overview of serious adverse events that may lead to discontinuation of the research treatment (Group A). [Figure 98] Provides the incidence of treatment-related adverse events (TEAEs) in three or more patients receiving monotherapy. [Figure 99] Provides information on the incidence of TEAEs in 3 or more patients - combination therapy. [Figure 100] Provides the incidence of serious TEAEs in all patients - monotherapy (n=18). [Figure 101] Provides the incidence of serious TEAEs in all patients - combination therapy (n=13). [Figure 102] Provides CHA.7.518.1.H4(S241P)PK profiles after IV infusion in Cycle 1 on day 1 - Group A and Group B. [Figure 103] This report provides summaries of responses assessed by the principal investigators in Group A and Group B (according to recist v1.1 dlt - evaluable population). [Figure 104A-104B] Swimmer plots of data from Group A and Group B are provided. A summary plot is provided in Figure 105C. [Figure 105] Provides waterfall plots of data from Group A and Group B. [Figure 106] Data on CHA.7.518.1.H4(S241P) + nivolumab is provided - a partial response (PR) was observed in a patient with MSS (microsatellite stable state) colorectal cancer (44-week study treatment is ongoing). [Figure 107] CHA.7.518.1.H4(S241P) provides data on monotherapy - PR was observed in a 25-week study treatment of an advanced platinum-resistant primary peritoneal cancer patient in a microsatellite-stable state (MSS). [Modes for carrying out the invention]

[0081] I. Introduction Cancer can be thought of as a condition where the patient lacks the ability to recognize and eliminate cancerous cells. Often, these transformed (e.g., cancerous) cells counteract immune surveillance. Natural regulatory mechanisms exist in the body that limit T cell activation and prevent uncontrolled T cell activity, which cancerous cells may utilize to evade or suppress the immune response. The goal of immunotherapy is to restore the ability of immune effector cells, particularly T cells, to recognize and eliminate cancer. The field of immuno-oncology, sometimes referred to as "immunotherapy," is rapidly evolving, and includes T cell checkpoint inhibitors such as Yervoy, Keytruda, and Opdivo. These are some of the recently approved antibodies. These antibodies are generally referred to as "checkpoint inhibitors" because they block normally negative regulatory factors of T-cell immunity. It is generally understood that an optimal antigen-specific immune response can be integrated using a variety of immunomodulatory signals, both co-stimulatory and co-inhibitory. Generally, these antibodies bind to checkpoint inhibitor proteins such as CTLA-4 and PD-1, which under normal circumstances prevent or suppress the activation of cytotoxic T cells (CTLs). By inhibiting checkpoint proteins, an increased T-cell response against tumors can be achieved, for example, through the use of antibodies that bind to these proteins. In other words, these cancer checkpoint proteins suppress the immune response, and when these proteins are blocked, for example using antibodies against checkpoint proteins, it leads to immune stimulation by activating the immune system, resulting in the treatment of conditions such as cancer and infectious diseases.

[0082] The present invention relates to formulations comprising antibodies against a human poliovirus receptor-associated immunoglobulin domain-containing protein or "PVRIG," which may also be referred to herein as "PV protein." PVRIG is expressed on the cell surface of NK and T cells and shares several similarities with other known immune checkpoints.

[0083] Accordingly, the present invention provides a formulation comprising an antibody comprising an antigen-binding domain that binds to human PVRIG and its peptide, and a method for activating T cells and / or NK cells to treat diseases such as cancer and infectious diseases, as well as other conditions in which increased immune activity leads to treatment. In particular, the present invention provides a formulation comprising an antibody comprising heavy and light chains, as well as vhCDR1, vhCDR2, vhCDR3, vlCDR1, vlCDR2, and vlCDR3 sequences from CHA.7.518.1.H4(S241P). In some embodiments, the anti-PVRIG antibody comprises having the same CDR as shown in Figure 3. In some embodiments, the anti-PVRIG antibody comprises having the same CDR as shown in Figures 5A-5D, as well as an anti-PVRIG antibody comprising the heavy and light chains provided in Figures 5A-5D. II. PVRIG protein The present invention provides a formulation comprising an antibody that specifically binds to the PVRIG protein. In this context, "protein" is used interchangeably with "polypeptide" and also includes peptides. The present invention provides an antibody that specifically binds to the PVRIG protein. PVRIG is a 326-amino acid transmembrane domain protein having a signal peptide (ranging from amino acids 1 to 40), an extracellular domain (ranging from amino acids 41 to 171), a transmembrane domain (ranging from amino acids 172 to 190), and a cytoplasmic domain (ranging from amino acids 191 to 326). The full-length human PVRIG protein is shown in Figure 1. There are two methionines that could be start codons, but the mature protein is identical.

[0084] Therefore, as used herein, the terms “PVRIG” or “PVRIG protein” or “PVRIG polypeptide” may optionally include any such protein, or its variants, conjugates, or fragments, including, but not limited to, known or wild-type PVRIG as described herein, as well as any naturally occurring splice variants, amino acid variants, or isoforms, and in particular, ECD fragments of PVRIG. The term “soluble” form of PVRIG may also be used interchangeably with the terms “soluble external domain (ECD)” or “external domain” or “extracellular domain (ECD)” and “fragment of PVRIG polypeptide,” which may broadly refer to one or more of the following optionally selected polypeptides: The PVRIG protein contains an immunoglobulin (Ig) domain within its extracellular domain, which is a PVR-like Ig folding domain. The PVR-like Ig folding domain is also present in other B7 proteins. Due to its similarity to Millie members, it may be involved in functional counterpart binding. The PVR-like Ig folding domain of the extracellular domain contains a single disulfide bond formed between intradomain cysteine ​​residues, which is typical of this folding and may be important to its structure and function. These cysteines are located at residues 22 and 93 (or 94). In one embodiment, a PVRIG soluble fragment is provided that can be used for testing a PVRIG antibody. The definition of the PVRIG protein includes PVRIG ECD fragments, including known ECD fragments such as those described in U.S. Patent No. 9,714,289 (which is incorporated herein by reference in its entirety).

[0085] As noted herein and more fully described below, anti-PVRIG antibodies (including antigen-binding fragments) that both bind to PVRIG and prevent activation by PVRL2 (e.g., most commonly by blocking the interaction between PVRIG and PVRL2) are used to enhance T cell and / or NK cell activation and are also used to treat diseases such as cancer and pathogen infections. III. Antibodies Accordingly, the present invention can be formulated by the formulations described herein and provides an anti-PVRIG antibody as shown in Figure 3 (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3). PVRIG, also known as the poliovirus receptor-associated immunoglobulin domain-containing protein, Q6DKI7, or C7orf15, relates to the amino acid and nucleic acid sequence shown in the RefSeq entrusted identifier NP_076975 shown in Figure 1. The antibody of the present invention is specific to the extracellular domain of PVRIG.

[0086] As will be discussed below, the term “antibody” is used in general terms. The antibodies found in use in the present invention can take several forms as described herein, including conventional antibodies, as well as antibody derivatives, fragments, and mimetic forms, as described below. Generally, the term “antibody” includes any polypeptide comprising at least one antigen-binding domain, as will be more fully described below. Antibodies may be polyclonal, monoclonal, heterogeneous, homogeneous, syngeneic, or modified forms thereof as described herein, with monoclonal antibodies finding particular use in many embodiments. In some embodiments, the antibodies of the present invention bind specifically or substantially specifically to the PVRIG molecule. The terms “monoclonal antibody” and “monoclonal antibody composition,” as used herein, refer to a group of antibody molecules comprising only one type of antigen-binding site capable of immune reaction with a particular epitope of an antigen, while “polyclonal antibody” and “polyclonal antibody composition” refer to a group of antibody molecules comprising multiple types of antigen-binding sites capable of interacting with a particular antigen. Monoclonal antibody compositions typically exhibit a single binding affinity to the particular antigen with which they immune reaction.

[0087] Conventional full-length antibody structural units are typically composed of tetramers. Each tetramer typically consists of two identical polypeptide chain pairs, each pair having one “light” chain (typically with a molecular weight of about 25 kDa) and one “heavy” chain (typically with a molecular weight of about 50–70 kDa). Human light chains are classified as kappa light chains and lambda light chains. The present invention focuses on the IgG class, which includes, but is not limited to, IgG1, IgG2, IgG3, and IgG4, and several subclasses. Thus, as used herein, “isotype” means any subclass of immunoglobulin defined by the chemical and antigenic characteristics of their constant regions. An exemplary antibody referred to herein as “CPA” is based on the IgG1 heavy constant region as shown in Figure 4, but anti-PVRIG antibodies of the present invention include those using IgG2, IgG3, and IgG4 sequences, or combinations thereof. For example, as is known in the art, different IgG isotypes have different effector functions, which may or may not be desirable. Therefore, the CPA antibody of the present invention may also have its IgG1 constant domain replaced with an IgG2, IgG3, or IgG4 constant domain (as shown in Figure 4), and IgG2 and IgG4 may find special use in some situations, for example, for ease of manufacture, or the latter may be preferred in some situations where reduced effector function is desired.

[0088] The amino-terminal portion of each chain contains a variable region of approximately 100-110 or more amino acids, commonly referred to in the art and herein as the “Fv domain” or “Fv region,” which is primarily involved in antigen recognition. Within the variable region, three loops converge for each of the V domains of the heavy and light chains to form an antigen-binding site. Each of the loops is called a complementarity-determining region (hereinafter referred to as “CDR”), where the variation in the amino acid sequence is most pronounced. “Variable” refers to the fact that certain segments of the variable region differ significantly in sequence between antibodies. The variability within the variable region is not uniformly distributed. Instead, the V region consists of a relatively invariant stretch called a framework region (FR) of 15-30 amino acids, separated by a shorter, highly variable region called a “hypervariable region.”

[0089] Each VH and VL consists of three hypervariable regions ("complementarity-determining regions," or "CDRs") and four FRs, arranged in the order FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4 from the amino terminus to the carboxyl terminus.

[0090] The hypervariable region generally includes amino acid residues from approximately 24-34 (LCDR1; "L" represents the light chain), 50-56 (LCDR2), and 89-97 (LCDR3) in the light chain variable region, and approximately 31-35B (HCDR1; "H" represents the heavy chain), 50-65 (HCDR2), and 95-102 (HCDR3) in the heavy chain variable region (in some cases, the numbering shifts slightly, as will be understood by those skilled in the art; Kabat et al., SEQUENCES OF PROTEINS OF IMMUNOLOGICAL INTEREST, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991) and / or those residues that form the hypervariable loop (e.g., residues 26-32 (LCDR1), 50-52 (LCDR2), and 91-96 (LCDR3) in the light chain variable region and 26-32 (HCDR1), 53-55 (HCDR2), and 96-101 (HCDR3) in the heavy chain variable region); Chothia and Lesk (1987) J.Mol.Biol.196:901-917. The specific CDR of the present invention is described below and is also shown in Figures 6A-6D.

[0091] The carboxyl terminus of each chain defines a constant region primarily involved in effector function. Kabat et al. collected numerous primary sequences of the variable regions of the heavy and light chains. Based on the degree of sequence conservation, Kabat et al. classified the individual primary sequences into CDRs and frameworks and compiled a list (see Sequences of Immunological Interest, 5th edition, NIH publication, No. 91-3242, E. Kabat et al., for the complete list).

[0092] The IgG subclass of immunoglobulins contains several immunoglobulin domains in the heavy chain. In this specification, “immunoglobulin (Ig) domain” refers to a region of immunoglobulin having a distinctly different tertiary structure. Heavy chain domains, including constant heavy (CH) domains and hinge domains, are of interest in this invention. In relation to IgG antibodies, each IgG isotype has three CH domains. Therefore, the “CH” domains in relation to IgG are as follows: “CH1” is found in Kabat. "CH2" refers to positions 118-220 according to the EU index, as shown in Kabat, and "CH3" refers to positions 341-447 according to the EU index, as shown in Kabat.

[0093] Accordingly, the present invention provides variable heavy domains, variable light domains, heavy constant domains, light constant domains, and Fc domains used as outlined herein. As used herein, “variable region” means a region of immunoglobulin comprising one or more Ig domains substantially encoded by either Vκ or Vλ, and / or VH genes constituting the kappa, lambda, and heavy chain immunoglobulin loci, respectively. Thus, the variable heavy domains comprise vhFR1-vhCDR1-vhFR2-vhCDR2-vhFR3-vhCDR3-vhFR4, and the variable light domains comprise vlFR1-vlCDR1-vlFR2-vlCDR2-vlFR3-vlCDR3-vlFR4. As used herein, “heavy chain constant region” means the CH1-hinge-CH2-CH3 portion of an antibody. As used herein, “Fc,” “Fc region,” or “Fc domain” refers to the polypeptide comprising the constant region of an antibody, excluding the first constant region immunoglobulin domain, and, in some cases, a portion of the hinge. Thus, Fc refers to the last two constant region immunoglobulin domains of IgA, IgD, and IgG, the last three constant region immunoglobulin domains of IgE and IgM, and the mobile hinge N-terminus to these domains. In the case of IgA and IgM, Fc may include the J chain. In the case of IgG, the Fc domain includes the immunoglobulin domains Cγ2 and Cγ3 (Cγ2 and Cγ3), as well as the lower hinge region between Cγ1 (Cγ1) and Cγ2 (Cγ2). While the boundaries of the Fc region may differ, the human IgG heavy chain Fc region is typically defined as containing residue C226 or P230 at its carboxyl terminus, numbered here according to the EU index similar to that of Kabat. In some embodiments, amino acid modifications are made to the Fc region to alter the binding to one or more FcγR receptors or FcRn receptors, as will be described in more detail below.

[0094] Therefore, as used herein, “Fc variant” or “variant Fc” means a protein containing amino acid modifications in the Fc domain. The Fc variants of the present invention are defined according to the amino acid modifications that constitute them. Thus, for example, N434S or 434S is an Fc variant having a substituted serine at position 434 relative to the parent Fc polypeptide, where the numbering follows the EU index. Similarly, M428L / N434S defines an Fc variant having substitutions M428L and N434S relative to the parent Fc polypeptide. The identity of the WT amino acids does not have to be specified, in which case the aforementioned variant is called 428L / 434S. The order in which the substitutions are provided is arbitrary, i.e., for example, 428L / 434S is the same Fc variant as M428L / N434S. For all positions discussed in the present invention relating to antibodies, unless otherwise specified, the numbering of amino acid positions follows the EU index.

[0095] As used herein, “Fab” or “Fab region” means a polypeptide containing the VH, CH1, VL, and CL immunoglobulin domains. Fab may refer to this region in isolation, or to this region in relation to a full-length antibody, antibody fragment, or Fab fusion protein. As used herein, “Fv” or “Fv fragment” or “Fv region” means a polypeptide containing the VL and VH domains of a single antibody. As will be understood by those skilled in the art, they generally consist of two strands.

[0096] Throughout this specification, either the IMTG numbering system or the Kabat numbering system is generally used to refer to residues within the variable domain (approximately residues 1-107 in the light chain variable region and residues 1-113 in the heavy chain variable region) (e.g., Kabat et al. (see above) (1991)). EU numbering, as seen in Kabat, is generally fixed. Used for regular domains and / or Fc domains.

[0097] CDRs contribute to the formation of antigen binding sites, or more specifically, the formation of antibody epitope binding sites. An "epitope" refers to a determinant that interacts with a specific antigen-binding site within the variable region of an antibody molecule, also known as a paratope. Epitopes are groups of molecules, such as amino acids or sugar side chains, and typically possess specific structural and charge properties. A single antigen may have two or more epitopes.

[0098] An epitope may include amino acid residues directly involved in binding (also called the immunodominant components of the epitope) and other amino acid residues not directly involved in binding, such as amino acid residues that are effectively blocked by specific antigen-binding peptides, in other words, amino acid residues that lie within the footprint of the specific antigen-binding peptide.

[0099] Epitopes can be either conformational or linear. Conformational epitopes are generated by the spatial juxtaposition of amino acids from different segments of a linear polypeptide chain. Linear epitopes are generated by adjacent amino acid residues within a polypeptide chain. Conformational and non-conformational epitopes can be distinguished in that binding to the former is lost in the presence of a denaturing solvent, while binding to the latter is not.

[0100] An epitope typically contains at least three, more commonly five, or eight to ten amino acids within its unique spatial structure. Antibodies that recognize the same epitope can be validated in simple immunoassays that demonstrate one antibody's ability to block the binding of another antibody to its target antigen, for example, "binning." Specific binning is described below.

[0101] The "antigen-binding portion" of an antibody (which can also be used interchangeably as "antigen-binding fragment," "antibody fragment," and "antibody derivative") is included within the definition of "antibody." In other words, for the purposes of the present invention, the antibody of the present invention has the minimum functional requirement that it binds to the PVRIG antigen. As will be understood by those skilled in the art, these include: (i) Fab fragments consisting of VL, VH, CL, and CH1 domains; (ii) Fd fragments consisting of VH and CH1 domains; (iii) F(ab')2 fragments, bivalent fragments containing two linked Fab fragments; (vii) single-chain Fv molecules (scFv) (the VH and VL domains are linked by a peptide linker that allows the two domains to bind and form an antigen-binding site) (Bird et al., 1988, Science 242:423-426, Huston et al., 1988, Proc. Natl. Acad. Sci. USA 85:5879-5883, the whole is incorporated by reference); (iv) "diabodies" or "triabodies," multivalent or multispecific fragments constructed by gene fusion (Tomlinson et al., 2000, Methods (v) "domain antibodies" or "dAbs" (sometimes called "immunoglobulin monovariate domains," including monoantibody variable domains from other species such as rodents (e.g., disclosed in WO00 / 29004), nurse shark and camel V-HH dAbs), (vi) numerous antigen fragments and derivatives with alternative structures, including but not limited to SMIPs (small molecule immunopharmaceuticals), camel bodies, nanobodies, and IgNARs.

[0102] Furthermore, an antibody or its antigen-binding portion (antigen-binding fragment, antibody fragment, antibody portion) may be covalently or noncovalently bonded to one or more other proteins or peptides. They may be part of larger immunoadhesion molecules (sometimes referred to as "fusion proteins") formed by binding. Examples of immunoadhesion molecules include the use of streptavidin core regions to construct tetramer scFv molecules, as well as the use of cysteine ​​residues, marker peptides, and C-terminal polyhistidine tags to construct divalent and biotinylated scFv molecules. Antibody moieties, e.g., Fab and F(ab')2 fragments, can be prepared from whole antibodies using conventional techniques such as papain digestion or pepsin digestion of the whole antibody, respectively. Furthermore, antibodies, antibody moieties, and immunoadhesion molecules can be obtained using standard recombinant DNA techniques described herein.

[0103] Generally speaking, the anti-PVRIG antibodies of the present invention are recombinant. As used herein, “recombinant” broadly refers to the product, for example, cells, or nucleic acids, proteins, or vectors, and indicates that the cells, nucleic acids, proteins, or vectors are modified by the introduction of heterologous nucleic acids or proteins or by modification of native nucleic acids or proteins, or that the cells are derived from such modified cells. Therefore, for example, recombinant cells express genes not found in the cell’s native (non-recombinant) form, or express native genes that are otherwise abnormally expressed, underexpressed, or not expressed at all.

[0104] As used herein, the term “recombinant antibody” includes all antibodies prepared, expressed, produced, or isolated by recombinant means, for example, (a) antibodies isolated from animals (e.g., mice) that are transgenic or transchromosomal to human immunoglobulin genes, or hybridomas prepared therefrom (as further described below), (b) antibodies isolated from host cells transformed to express human antibodies, for example, transfectomas, (c) antibodies isolated from recombinant combinatorial human antibody libraries, and (d) antibodies prepared, expressed, produced, or isolated by any other means involving splicing of human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies have variable regions in which the framework and CDR region are derived from human germline immunoglobulin sequences. However, in certain embodiments, such recombinant human antibodies may be subjected to in vitro mutagenesis (or, if transgenic animals for human Ig sequences are used, in vivo somatic mutagenesis), and thus the V of recombinant antibodies H and V L The amino acid sequence of the region is human germline V H and V L These sequences are derived from and related to other sequences, but may not naturally exist within the human antibody germline repertoire in vivo. A. Selective antibody manipulation The anti-PVRIG antibody of the present invention (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3) can be modified or manipulated to alter its amino acid sequence by amino acid substitution.

[0105] In this specification, “amino acid substitution” or “substitution” means replacing an amino acid at a specific position in a parent polypeptide sequence with a different amino acid. In particular, in some embodiments, the substitution is for an amino acid that does not naturally exist at a particular position (either not naturally present in an organism or not present in any organism). For example, substitution E272Y refers to a variant polypeptide, in this case the Fc variant, in which glutamic acid at position 272 is substituted with tyrosine. To clarify, a protein that has been manipulated to alter the nucleic acid coding sequence but not the starting amino acid (e.g., replacing CGG (coding arginine) with CGA (still encoding arginine) to increase the host organism’s expression level) is not an “amino acid substitution.” That is, if, despite the creation of a new gene encoding the same protein, that protein has the same amino acid at the specific position where it starts, it is not an amino acid substitution.

[0106] As discussed herein, amino acid substitutions may be made to alter the affinity of the CDR to the PVRIG protein (including both increased and decreased binding, as fully outlined below) and to modify additional functional properties of the antibody. For example, an antibody may be manipulated to modify one or more functional properties of the antibody, such as serum half-life, complement binding, Fc receptor binding, and / or antigen-dependent cytotoxicity, typically including modifications within the Fc region. Furthermore, antibodies according to at least some embodiments of the present invention may also be chemically modified (e.g., one or more chemical moieties may bind to the antibody) or modified to alter their glycosylation, also to modify one or more functional properties of the antibody. Such embodiments are further described below. The numbering of residues within the Fc region is the numbering of the Kabat EU index.

[0107] In one embodiment, C H1The hinge region of CH1 is modified by altering the number of cysteine ​​residues within the hinge region, for example, by increasing or decreasing it. This approach is further described in U.S. Patent No. 5,677,425 (Bodmer et al.). The number of cysteine ​​residues within the hinge region of CH1 is modified, for example, to facilitate the assembly of light and heavy chains, or to increase or decrease the stability of the antibody.

[0108] In another embodiment, the Fc hinge region of the antibody is mutated to reduce the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc-hinge fragment such that the antibody has a Staphylococcyl protein A (SpA) bond that is impaired in function compared to the native Fc-hinge domain SpA bond. This approach is described in more detail in U.S. Patent No. 6,165,745 by Ward et al.

[0109] In some embodiments, amino acid substitutions may be made within the Fc region to generally modify binding to the FcγR receptor. As used herein, “Fc gamma receptor,” “FcγR,” or “Fc gamma R” means any member of the protein family that binds to the Fc region of an IgG antibody and is encoded by the FcγR gene. In humans, this family includes FcγRI(CD64) (including isoforms FcγRIa, FcγRIb, and FcγRIc); FcγRII(CD32) (isoforms FcγRIIa (including allotypes H131 and R131), FcγRIIb (including FcγRIIb-1 and FcγRIIb-2), and FcγRIIc); and FcγRIII(CD16) (isoforms FcγRIIIa (including allotypes V158 and F158) and FcγRIIIb (including allotypes FcγRIIIb-NA1 and FcγRIIIb-NA2)) (Jefferis et al., 2002, Immunol Lett). This includes, but is not limited to, 82:57-65 (the whole is incorporated by reference), as well as any undiscovered human FcγR or FcγR isoform or allotype. FcγR can originate from any organism, including, but not limited to, humans, mice, rats, rabbits, and monkeys. Mouse FcγR includes, but is not limited to, FcγRI(CD64), FcγRII(CD32), FcγRIII-1(CD16), and FcγRIII-2(CD16-2), as well as any undiscovered mouse FcγR or FcγR isoform or allotype.

[0110] Several useful Fc substitutions exist that can be performed to modify binding to one or more FcγR receptors. Substitutions that result in increased or decreased binding may be useful. For example, increased binding to FcγRIIIa generally leads to an increase in ADCC (antibody-dependent cell-mediated cytotoxicity), i.e., a cell-mediated response in which nonspecific cytotoxic cells expressing FcγR recognize a bound antibody on target cells, and subsequently cause lysis of the target cells. It is known that, similarly, under certain circumstances, reduced binding to FcγRIIb (an inhibitory receptor) may also be beneficial. Amino acid substitutions useful in the present invention include those described in U.S. Patent Application No. 11 / 124,620 (particularly Figure 41) and U.S. Patent No. 6,737,056, both of which are expressly incorporated herein by reference in their entirety, particularly the variants disclosed therein. Specific variants that find use include, but are not limited to, 236A, 239D, 239E, 332E, 332D, 239D / 332E, 267D, 267E, 328F, 267E / 328F, 236A / 332E, 239D / 332E / 330Y, 239D, 332E / 330L, 299T, and 297N.

[0111] In addition, the antibodies of the present invention are modified to increase their biological half-life. Various approaches are possible. For example, one or more of the following mutations: T252L, T254S, T256F can be introduced, as described in U.S. Patent No. 6,277,375 (Ward). Alternatively, to increase the biological half-life, the antibody may contain a salvage receptor-binding epitope obtained from two loops of the CH2 domain of the Fc region of IgG, as described in U.S. Patents No. 5,869,046 and No. 6,121,022 (Presta et al.). H1 or C L It may be modified within the region. Further mutations for increasing the serum half-life are disclosed in U.S. Patents 8,883,973, 6,737,056, and 7,371,826, which include 428L, 434A, 434S, and 428L / 434S.

[0112] In yet another embodiment, the Fc region is modified by replacing at least one amino acid residue with a different amino acid residue to alter the effector function of the antibody. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320, and 322 can be replaced with a different amino acid residue so that the antibody has a modified affinity for the effector ligand, while the antigen-binding ability of the parent antibody is retained. The effector ligand whose affinity is modified may be, for example, the Fc receptor or the C1 component of complement. This approach is described in further detail in both U.S. Patents 5,624,821 and 5,648,260 by Winter et al.

[0113] One or more amino acids selected from amino acid residues 329, 331, and 322 can be replaced with different amino acid residues so that the antibody has modified C1q binding and / or reduced or absent complement-dependent cytotoxicity (CDC). This approach is described in more detail in U.S. Patent No. 6,194,551 by Idusogie et al.

[0114] In another example, one or more amino acid residues within positions 231 and 239 are modified, thereby altering the antibody's ability to fix complement. This approach is further described in PCT Publication WO94 / 29351 by Bodmer et al.

[0115] In yet another example, the Fc region is located at the following positions: 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 309, 312, 315, 320, 322, 32 Modifying one or more amino acids of 4, 326, 327, 329, 330, 331, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, ​​388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438, or 439 increases the ability of antibodies to mediate antibody-dependent cytotoxicity (ADCC), and / or

[0116]

number

[0117] The antibody is modified to increase its affinity for the receptor. This approach is further described in Presta's PCT Publication WO00 / 42072.

[0118]

number

[0119] The binding sites on human IgG1 for FcRn are mapped, and variants with improved binding are described (see Shields, R. Let al. (2001) J. Biol. Chem. 276:6591-6604). Specific mutations at positions 256, 290, 298, 333, 334, and 339 are shown to improve binding to FcyRIII. Additionally, the following variant combinations: T256A / S298A, S298A / E333A, S298A / K224A, and S298A / E333A / K334A,

[0120]

number

[0121] This has been shown to improve binding. Furthermore, mutations such as M252Y / S254T / T256E or M428L / N434S improve binding to FcRn and increase the circulating half-life of the antibody (see Chan CA and Carter PJ (2010) Nature Rev Immunol 10:301-316).

[0122] In another embodiment, the antibody may be modified to inhibit in vivo Fab arm exchange. Specifically, this process involves the exchange of IgG4 halves (one heavy chain + one light chain) between other IgG4 antibodies, effectively resulting in a functionally monovalent bispecific antibody. Mutations in the hinge region and constant domain of the heavy chain inhibit this exchange (see Aalberse, RC, Schuurman J., 2002, Immunology 105:9-19).

[0123] In yet another embodiment, the glycosylation of the antibody is modified. For example, an aglycosylated antibody can be produced (i.e., the antibody lacks glycosylation). Glycosylation can be modified, for example, to increase the affinity of the antibody to an antigen or to reduce effector functions such as ADCC. Such carbohydrate modification can be achieved, for example, by modifying one or more glycosylation sites in the antibody sequence, e.g., N297. For example, glycosylation can be eliminated at one or more amino acid substitutions that result in the exclusion of glycosylation sites in one or more variable region frameworks.

[0124] Additionally or alternatively, antibodies with modified types of glycosylation can be produced, for example, low-fucosylated antibodies with reduced amounts of fucosyl residues or antibodies with increased bipartite GlcNac structures. Such modifications of glycosylation patterns have been shown to increase the ADCC capacity of antibodies. Such carbohydrate modifications can be achieved, for example, by expressing antibodies in host cells with modified glycosylation mechanisms. Cells with modified glycosylation mechanisms have been described in the art and can be used as host cells to express recombinant antibodies according to at least some embodiments of the present invention, thereby producing antibodies with modified glycosylation. For example, the cell lines Ms704, Ms705, and Ms709 lack the fucosyltransferase gene FUT8 (α(1,6) fucosyltransferase) so that antibodies expressed in the Ms704, Ms705, and Ms709 cell lines lack the carbohydrate fucose. The Ms704, Ms705, and Ms709 FUT8 cell lines are created by targeted disruption of the FUT8 gene in CHO / DG44 cells using two substitution vectors (US Patent Publication No. 2004 / 0110704 by Yamane et al.). , and Yamane-Ohnuki et al. (2004) Biotechnol (Bioeng 87:614-22). As another example, Hanai et al. (EP1,176,195) describe cell lines with a functionally disrupted FUT8 gene, which encodes fucosyltransferase, and therefore antibodies expressed in such cell lines exhibit hypofucosylation by reducing or removing α1,6 binding-related enzymes. Hanai et al. further describe cell lines that have low enzymatic activity, adding fucose to N-acetylglucosamine that binds to the Fc region of the antibody, or, for example, rat myeloma cell line YB2 / 0 (ATCC CRL 1662) has no enzymatic activity at all. Presta's PCT publication WO03 / 035835 describes the Lec13 cell line, a variant CHO cell line that exhibits reduced ability to bind Asn(297) linked carbohydrates to fucose, and also results in reduced fucosylation of antibodies expressed in its host cells (see also Shields, R. Let al. (2002) J. Biol. Chem. 277:26733-26740). Umana et al., PCT Publication No. WO99 / 54342, describes cell lines engineered to express glycoprotein-modifying glycosyltransferases (e.g., β(1,4)-N-acetylglucosaminyltransferase III (GnTIII)), in which antibodies expressed in such engineered cell lines show an increase in bifid GlcNac structures, resulting in increased ADCC activity of the antibodies (see also Umana et al. (1999) Nat. Biotech. 17:176-180). Alternatively, fucosidase enzymes can be used to cleave fucose residues in antibodies. For example, α-L-fucosidase removes fucosyl residues from antibodies (Tarentino, A et al. (1975) Biochem. 14:5516-23).

[0125] Another modification of the antibodies contemplated herein by the present invention is, for example, pegylation, i.e., the addition of other water-soluble moieties (typically polymers) to increase the half-life. Antibodies can be pegylated, for example, to increase the biological (e.g., serum) half-life of the antibody. To pegylate an antibody, the antibody or a fragment thereof is typically reacted with polyethylene glycol (PEG), e.g., a reactive ester or aldehyde derivative of PEG, under conditions such that one or more PEG groups bind to the antibody or antibody fragment. Preferably, pegylation is carried out via an acylation or alkylation reaction with a reactive PEG molecule (or similar reactive water-soluble polymer). As used herein, the term "polyethylene glycol" encompasses any of the forms of PEG that have been used to derivatize other proteins, such as mono(C1-C 10 ) alkoxy, or aryloxy-polyethylene glycol, or polyethylene glycol-maleimide. In certain embodiments, the antibody to be pegylated is an aglycosylated antibody. Methods for pegylating proteins are known in the art and can be applied to antibodies according to at least some embodiments of the present invention. See, for example, EP0154316 by Nishimura et al., and EP0401384 by Ishikawa et al.

[0126] In addition to substitutions made to modify the binding affinity for FcγR and / or FcRn and / or increase the in vivo serum half-life, additional antibody modifications can be made, as described in more detail below.

[0127] In some cases, affinity maturation is carried out. Amino acid modifications in the CDRs are sometimes referred to as "affinity maturation." An "affinity matured" antibody has one or more modifications in one or more CDRs, which result in an improvement in the affinity of the antibody for the antigen compared to the parental antibody that does not have those modifications. In some cases, although rare, it may be desirable to reduce the affinity of the antibody for its antigen, but this is generally not preferred.

[0128] In some embodiments, one or more amino acid modifications are performed in one or more CDRs of the PVRIG antibody of the present invention. Generally, only one, two, or three amino acids are substituted in any single CDR, and generally, one, two, three, or fewer. Four, five, six, seven, eight, nine, or ten changes are performed within a set of CDRs. However, it should be understood that any combination of no substitution, one, two, or three substitutions in any CDR can be independently and arbitrarily combined with any other substitution.

[0129] Affinity maturation may be performed to increase the binding affinity of the antibody to the PVRIG antigen by at least approximately 10% to 50-100-150%, or 1 to 5 times, compared to the “parent” antibody. Preferred affinity-matured antibodies have nanomolar affinity or even picomolar affinity to the PVRIG antigen. Affinity-matured antibodies are produced by known procedures. For example, see Marks et al., 1992, Biotechnology 10:779-783, where affinity maturation by variable heavy chain (VH) and variable light chain (VL) domain shuffling is described. Random mutagenesis of CDRs and / or framework residues is described, for example, in Barbas, et al. 1994, Proc. Nat. Acad. Sci, USA 91:3809-3813, Shier et al., 1995, Gene 169:147-155, Yelton et al., 1995, J. Immunol. 155:1994-2004, Jackson et al., 1995, J. Immunol. 154(7):3310-9, and Hawkins et al., 1992, J. Mol. Biol. 226:889-896.

[0130] Alternatively, for example, "silent" amino acid modifications that do not significantly alter the antibody's affinity for the antigen may be performed on one or more of the CDRs of the antibody of the present invention. These may be performed for several reasons, including optimization of expression (as may be performed on the nucleic acid encoding the antibody of the present invention).

[0131] Therefore, variant CDRs and antibodies are included in the definitions of CDRs and antibodies of the present invention, i.e., antibodies of the present invention may have amino acid modifications in one or more of the CDRs of the enumerated antibodies of the present invention. In addition, as outlined below, amino acid modifications may be carried out independently and optionally in any region outside the CDR, including the framework and constant region. IV.PVRIG antibody The present invention provides an anti-PVRIG antibody. (For convenience, "anti-PVRIG antibody" and "PVRIG antibody" are used interchangeably.) The anti-PVRIG antibody of the present invention includes, for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3, and specifically binds to the ECD of human PVRIG, preferably human PVRIG1, as illustrated in Figure 3.

[0132] Specific binding of PVRIG or PVRIG to an epitope is, for example, at least about 10 -4 M, at least about 10 -5 M, at least about 10 -6 M, at least about 10 -7 M, at least about 10 -8 M, at least about 10 -9 M, alternatively at least 10 -10 M, at least about 10 -11 M, at least about 10 -12 This can be indicated by antibodies with a KD of M or greater, where KD refers to the rate of dissociation of a particular antibody-antigen interaction. Typically, antibodies that specifically bind to an antigen will have a KD 20-fold, 50-fold, 100-fold, 500-fold, 1000-fold, 5,000-fold, 10,000-fold, or more greater than that of a control molecule compared to the PVRIG antigen or epitope.

[0133] However, as shown in the examples, for optimal binding to PVRIG expressed on the surface of NK and T cells, the antibody is preferably less than 50 nM, most preferably 1 nM. Having a KD of less than 0.1 nM, and 1 pM and less than 0.1 pM are found to be useful in the methods of the present invention.

[0134] Furthermore, specific binding to a particular antigen or epitope may be demonstrated by an antibody whose KA or Ka for a PVRIG antigen or epitope is at least 20 times, 50 times, 100 times, 500 times, 1000 times, 5,000 times, 10,000 times, or more than 20 times, 50 times, 10000 times, or greater than that of a control, where KA or Ka refers to the association rate of a particular antibody-antigen interaction.

[0135] In some embodiments, the anti-PVRIG antibody of the present invention has a K content of 100 nM or less, 50 nM or less, 10 nM or less, or 1 nM or less (i.e., higher binding affinity), or 1 pM or less. D It binds to human PVRIG, and K D This is determined by known methods, such as surface plasmon resonance (SPR, e.g., Biacore assay), ELISA, and KINEXA, most typically by SPR at 25°C or 37°C.

[0136] The present invention provides an antigen-binding domain containing a full-length antibody, comprising several specific enumerated sets of six CDRs, as provided in Figure 3.

[0137] The present invention further provides variable weight and light domains, as well as full-length weight and light chains.

[0138] As discussed herein, the present invention further provides variants of the above components, including variants in the CDR outlined above. In addition, a variable heavy chain may be at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% identical to the “VH” sequence herein, and / or may contain 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more amino acid changes when using an Fc variant. A variable light chain is provided which may be at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% identical to the “VL” sequence herein, and / or may contain 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more amino acid changes when using an Fc variant. Similarly, heavy and light chains are provided that are at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% identical to the “HC” and “LC” sequences herein, and / or, when using an Fc variant, contain 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acid changes.

[0139] Therefore, the present invention provides antibodies comprising the following CHA set of CDRs, typically full-length or scFv domains, the sequences of which are shown in Figure 3: CHA.7.518.1.H4(S241P)vhCDR1, CHA.7.518.1.H4(S241P)vhCDR2, CHA.7.518.1.H4(S241P)vhCDR3, C HA.7.518.1.H4(S241P)vlCDR1, CHA.7.518.1.H4(S241P)vlCDR2, and CHA.7.518.1.H4(S241P)vlCDR3.

[0140] In addition, the variable heavy and variable light chain framework regions can be humanized as known in the art (occasionally, variants are generated in the CDR as needed), thus generating humanized variants of the VH and VL chains shown in Figure 3. Furthermore, the humanized variable heavy and light domains can subsequently be fused with human constant regions such as constant regions from IgG1, IgG2, IgG3, and IgG4.

[0141] In addition, the range includes sequences that may have the same CDR but may have variations within the variable domain (or the entire heavy or light chain). For example, PVRIG antibodies may include those with the same CDR as shown in Figure 3 or Figures 5A-5D, but with lower identity across the variable region, for example, 95% or 98% identity percentage.

[0142] The percentage of identity between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput.Appl.Biosci.,4:11-17(1988)) incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. In addition, the percentage of identity between two amino acid sequences can be determined using either a Blossum 62 matrix or a PAM250 matrix, along with gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6, incorporating the algorithm of Needleman and Wunsch (J.Mol.Biol.48:444-453(1970)) incorporated into the GAP program in the GCG software package (commercially available), using either a Blossum 62 matrix or a PAM250 matrix, as well as gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6.

[0143] Additionally or alternatively, the protein sequences of the present invention can be used as further "query sequences" to perform searches against public databases, for example, to identify relevant sequences. Such searches can be performed using the XBLAST program (version 2.0) described in Altschul, et al. (1990) J Mol. Biol. 215:403-10. A BLAST protein search can be performed using the XBLAST program with a score of 50 and a word length of 3 to obtain amino acid sequences homologous to antibody molecules according to at least some embodiments of the present invention. To obtain gapped alignments for comparative purposes, Gapped BLAST can be used as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402. When using the BLAST and Gapped BLAST programs, the default settings parameters for each program (e.g., XBLAST and NBLAST) can be used.

[0144] Generally, the percentage of identity for comparison between PVRIG antibodies is at least 75%, at least 80%, and at least 90%, with a preferred percentage of identity of at least about 95%, 96%, 97%, 98%, or 99%. The percentage of identity may relate to the entire amino acid sequence, e.g., the entire heavy or light chain, or to a portion of the chain. For example, antibodies sharing identity across the entire variable region (e.g., 95% or 98% identity across the variable region, or in some embodiments, at least 95% or at least 98% identity across the constant region), or across the entire constant region, or only across the Fc domain, are included in the definition of anti-PVRIG antibodies of the present invention. V. Formulation of anti-PVRIG antibodies A composition of an anti-PVRIG antibody and / or its antigen-binding moiety (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3) can be formulated into a pharmaceutical composition comprising a carrier suitable for the desired delivery method. Suitable carriers include any substance that, when combined with the therapeutic composition, preserves the antitumor function of the therapeutic composition and is generally not reactive with the patient's immune system. Examples include, but are not limited to, any of several standard pharmaceutical carriers, such as sterile phosphate-buffered saline or bacteriostatic water. (Generally, Remington's Pharmaceutical Sciences 16) th (See Edition, A. Osal., Ed., 1980). Acceptable carriers, excipients, or stabilizers are non-toxic to the recipient at the dosage and concentration used and include buffers such as phosphates, citrates, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl, or benzyl alcohol; alkylparabens such as methyl or propylparaben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol, etc.); low molecular weight (less than approximately 10 residues) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulin; poly This includes hydrophilic polymers such as vinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrin; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; sweeteners and other flavoring agents; fillers such as microcrystalline cellulose, lactose, corn, and other starches; binding factors; additives; colorants; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants such as TWEEN®, PLURONICS®, or polyethylene glycol (PEG).

[0145] In a preferred embodiment, the pharmaceutical composition comprising the anti-PVRIG antibody of the present invention (having the same CDR as that shown in Figure 3) may be in a water-soluble form, such as existing as a pharmaceutically acceptable salt, intended to contain both an acid-addition salt and a base-addition salt. A "pharmaceutically acceptable acid-addition salt" refers to a salt formed by an inorganic acid, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc., and an organic acid, such as acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc., that retains the biological efficacy of the free base and is not biologically or otherwise inappropriate. "Pharmacologically acceptable base addition salts" include those derived from inorganic bases, such as sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Ammonium, potassium, sodium, calcium, and magnesium salts are particularly preferred. Salts derived from pharmaceutically acceptable organic non-toxic bases include primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and salts of basic ion exchange resins such as isopropylamine, trimethylamine, diethylamine, and ethanolamine. Formulations used for in vivo administration are preferably sterilized. This can be easily achieved by filtration through a sterile filtration membrane or by other methods.

[0146] As used herein, the term “activity” refers to the functional activity (plural) of an anti-PVRIG antibody and / or its antigen-binding moiety. Functional activity includes, but is not limited to, biological activity and / or binding affinity.

[0147] As used herein, the term “stability” is used in a structural context, for example, in relation to the structural integrity of the anti-PVRIG antibody and / or its antigen-binding moiety, or in a functional context, for example, in relation to the ability of the anti-PVRIG antibody and / or its antigen-binding moiety to maintain its function and / or activity over time (including, for example, the stability of the anti-PVRIG antibody and / or its antigen-binding moiety or the formulation stability of the anti-PVRIG antibody and / or its antigen-binding moiety, including anti-PVRIG antibodies having the same CDR as shown in Figure 3). To be understood, the anti-PVRIG antibody and / or its antigen-binding moiety under discussion may be contained in formulations by the methods and compositions described herein, and the stability of the protein refers to its stability in those formulations. Several implementations In this context, the stability of the anti-PVRIG antibody and / or its antigen-binding moiety composition is determined by measuring the binding activity of the composition, including, for example, using the assay described in this application and the figures provided therewith, as well as other applicable assays known in the art. In some embodiments, the stability of the anti-PVRIG antibody and / or its antigen-binding moiety composition is compared with compositions of anti-PVRIG antibody and / or its antigen-binding moiety formulated with sugars, sugar alcohols, and / or nonionic surfactants, as described herein, and without at least one amino acid, salt, and / or nonionic surfactant, and / or with different combinations of components. In some embodiments, the formulation does not contain sugars and / or sugar alcohols.

[0148] As used herein, “storage-stable” aqueous anti-PVRIG antibody and / or its antigen-binding moiety composition refers to a solution comprising, for example, an anti-PVRIG antibody and / or its antigen-binding moiety formulated to enhance the stability of the protein in at least 10% solution over a given storage period. In the context of this disclosure, an anti-PVRIG antibody and / or its antigen-binding moiety can be made “storage-stable” by adding at least one amino acid, salt, or nonionic surfactant as a stabilizer. In some embodiments, the stability of an anti-PVRIG antibody and / or its antigen-binding moiety in any given formulation can be measured, for example, by monitoring the formation of aggregates, loss of bulk binding activity, or formation of degradation products over a period of time. The absolute stability of the formulation, and the stabilizing effect of sugars, sugar alcohols, or nonionic surfactants, will vary depending on the specific composition being stabilized. In one embodiment, the stability of an anti-PVRIG antibody and / or its antigen-binding moiety composition is determined by measuring the binding activity of the anti-PVRIG antibody and / or its antigen-binding moiety in the composition, for example, by using ELISA or other binding activity assays. In one embodiment, the stability of a composition of anti-PVRIG antibody and / or its antigen-binding moiety formulated with sugars, sugar alcohols, and / or nonionic surfactants as described herein is compared to a composition of anti-PVRIG antibody and / or its antigen-binding moiety formulated without at least one amino acid, salt, and / or nonionic surfactant, and / or with different combinations of components. In some embodiments, the formulation does not contain sugars and / or sugar alcohols.

[0149] As used herein, “shelf life” refers to the period during which a formulation maintains a predetermined level of stability at a predetermined temperature. In certain embodiments, the predetermined temperature refers to storage at freezing temperatures (e.g., -80°C, -25°C, 0°C), refrigeration (e.g., 0°C to 10°C), or room temperature (e.g., 18°C ​​to 32°C).

[0150] As used herein, the term “stability time” refers to the length of time during which a formulation is considered stable. For example, the stability time of a formulation may refer to the length of time during which the level of protein aggregation and / or degradation in the formulation remains below a certain threshold (e.g., 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, etc.), and / or the length of time during which the formulation maintains biological activity above a certain threshold (e.g., 100%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, etc.) of the amount of activity (including binding activity) present in the formulation at the start of the storage period.

[0151] In the context of this disclosure, a storage-stable aqueous composition of an anti-PVRIG antibody and / or its antigen-binding moiety formulated with sugars, sugar alcohols, and / or nonionic surfactants is longer stable than a composition of the same anti-PVRIG antibody and / or its antigen-binding moiety formulated without at least one amino acid, salt, and / or nonionic surfactant. It will have a fixed time. In some embodiments, the storage-stable aqueous composition of anti-PVRIG antibody and / or its antigen-binding moiety will have a stability time that is, for example, at least 10% longer than that of the composition of anti-PVRIG antibody and / or its antigen-binding moiety formulated in the absence of at least one amino acid, salt, and / or nonionic surfactant, or at least 15% to 20% longer than that of the composition of anti-PVRIG antibody and / or its antigen-binding moiety formulated in the absence of at least one amino acid, salt, and / or nonionic surfactant. It will have a stabilization time that is 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190% longer, or at least twice as long, or at least 2.5 times, 3.0 times, 3.5 times, 4.0 times, 4.5 times, 5.0 times, 5.5 times, 6.0 times, 6.5 times, 7.0 times, 7.5 times, 8.0 times, 8.5 times, 9.0 times, 9.5 times, 10 times, or longer.

[0152] As used herein, "BDS" refers to "bulk active pharmaceutical ingredient." A. Stabilized liquid formulation In some embodiments, the disclosure provides stabilized aqueous formulations of anti-PVRIG antibodies and / or their antigen-binding moieties (e.g., anti-PVRIG antibodies having the same CDR as shown in Figure 3). The following embodiments are, in part, based on the finding that the inclusion of at least one amino acid, salt, and / or nonionic surfactant stabilizes the liquid anti-PVRIG antibody and / or its antigen-binding moiety composition compared to compositions lacking at least one amino acid, salt, and / or nonionic surfactant. In some embodiments, the formulations do not contain sugars and / or sugar alcohols.

[0153] As will be recognized by those skilled in the art, anti-PVRIG antibodies and / or their antigen-binding moieties formulated by embodiments provided herein may, in addition to the explicitly disclosed components, include counterions contributed by the inclusion of solution components or pH modifiers, such as sodium or potassium from acetate, sodium hydroxide, or potassium hydroxide, or chlorides contributed by calcium chloride or hydrochloric acid. In the context of this disclosure, a storage-stable anti-PVRIG antibody and / or its antigen-binding moiety composition consisting of, or essentially consisting of, a given formulation may further include one or more counterions required by the formulation process at a specific pH.

[0154] In one embodiment, the storage-stable anti-PVRIG antibody and / or antigen-binding moiety provided herein is stabilized at a refrigerated temperature (i.e., 2°C to 10°C) for a certain period of time. For example, in one embodiment, a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment is stable when stored at a refrigerated temperature for at least 4 days. In other embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable at a refrigerated temperature for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 21, 28, or more days. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 1 month. In some embodiments, the composition is stable for at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48 or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable at a temperature of 2°C to 8°C. It remains stable for a long period of time when stored.

[0155] In one embodiment, a stable liquid pharmaceutical formulation comprising an anti-PVRIG antibody or its antigen-binding fragment provided herein is stabilized at room temperature (i.e., 18°C ​​to 32°C) for a certain period of time. For example, in one embodiment, a stable liquid pharmaceutical formulation comprising an anti-PVRIG antibody or its antigen-binding fragment is stable when stored at room temperature for at least 4 days. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable at room temperature for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 21, 28, or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 1 month. In yet another embodiment, the composition is stable for at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48 or more. In some embodiments, room temperature refers to 20°C–30°C, 21°C–29°C, 22°C–28°C, 23°C–27°C, 24°C–26°C, or about 25°C. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for a long period of time when stored at a temperature of 20°C–25°C. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for extended periods when stored at a temperature of approximately 25°C.

[0156] In one embodiment, the storage-stable anti-PVRIG antibody and / or antigen-binding moiety provided herein is stabilized at a high temperature (i.e., 32°C to 42°C) for a certain period of time. For example, in one embodiment, a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment is stable when stored at a high temperature for at least 4 days. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable at a high temperature for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 21, 28, or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 1 month. In yet another embodiment, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48 or more. In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is stable for a long period of time when stored at a temperature of 35°C to 40°C.

[0157] In one embodiment, a stored anti-PVRIG antibody and / or antigen-binding moiety composition is considered storage stable as long as the composition maintains at least 40% of the antibody-binding activity present at the start of the storage period (e.g., time = 0). In another embodiment, a stored composition is considered stable as long as the composition maintains at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or more of the antibody-binding activity present at the start of the storage period (e.g., time = 0). In one embodiment, the antibody-binding activity is measured using any assay known in the art.

[0158] In some embodiments, a composition of an anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (e.g., at least one amino acid, salt, and / or nonionic surfactant) if the composition of the anti-PVRIG antibody and / or antigen-binding moiety contains at least 10% more antibody-binding activity after storage for a certain period compared to a composition of an anti-PVRIG antibody and / or antigen-binding moiety that does not contain a stabilizer or contains a small amount of a stabilizer. In other embodiments, a composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (e.g., at least one amino acid, salt, and / or nonionic surfactant) if the composition contains at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, or more than00%, of anti-PVRIG antibody and / or antigen-binding moiety activity after storage for a certain period of time, compared to a composition of anti-PVRIG antibody and / or antigen-binding moiety that does not contain a stabilizer or contains a small amount of a stabilizer.

[0159] In one embodiment, a stored composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stable as long as the proportion of anti-PVRIG antibody and / or antigen-binding moiety present in an aggregated state remains 50% or less. In several embodiments, a stored composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stable as long as the proportion of anti-PVRIG antibody and / or antigen-binding moiety present in an aggregated state remains 45%, 40%, 35%, 30%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less.

[0160] In some embodiments, a composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (composition of anti-PVRIG antibody and / or antigen-binding moiety, at least one amino acid, salt, and / or nonionic surfactant) if the composition contains anti-PVRIG antibody and / or antigen-binding moiety that is present in at least 10% less aggregated state after storage for a certain period compared to a composition of anti-PVRIG antibody and / or antigen-binding moiety that does not contain a stabilizer or contains a small amount of a stabilizer. In other embodiments, a composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (e.g., at least one amino acid, salt, and / or nonionic surfactant) if the composition contains anti-PVRIG antibody and / or antigen-binding moiety that, after storage for a certain period, is present in an aggregated state by at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, or more, compared to a composition of anti-PVRIG antibody and / or antigen-binding moiety that does not contain a stabilizer or contains a small amount of a stabilizer.

[0161] In some embodiments, a stored anti-PVRIG antibody and / or antigen-binding moiety composition is considered stable as long as it maintains at least 40% of its initiation binding activity (e.g., at time = 0) after exposure to mechanical stress. In another embodiment, a stored composition is considered stable as long as it maintains 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or more of its initiation binding activity (e.g., at time = 0) after exposure to mechanical stress. In some embodiments, the mechanical stress is agitation (e.g., shaking).

[0162] In some embodiments, the anti-PVRIG antibody and / or antigen-binding moiety composition is compared to the anti-PVRIG antibody and / or antigen-binding moiety composition which contains no stabilizer or a small amount of stabilizer. A composition is considered stabilized by the addition of a stabilizer (e.g., at least one amino acid, salt, or nonionic surfactant) if it contains at least 10% more binding activity after exposure to mechanical stress. In other embodiments, a composition of an anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (e.g., sugar, sugar alcohol, or nonionic surfactant) if, after exposure to mechanical stress, the composition contains at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, or more than

[0163] In some embodiments, a stored anti-PVRIG antibody and / or antigen-binding moiety composition is considered stable as long as the proportion of anti-PVRIG antibody and / or antigen-binding moiety present in an aggregated state after exposure to mechanical stress remains 50% or less. In other embodiments, a stored anti-PVRIG antibody and / or antigen-binding moiety composition is considered stable as long as the proportion of anti-PVRIG antibody and / or antigen-binding moiety present in an aggregated state after exposure to mechanical stress remains 45%, 40%, 35%, 30%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less. In some embodiments, the mechanical stress is agitation (e.g., shaking).

[0164] In some embodiments, a composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (e.g., at least one amino acid, salt, or nonionic surfactant) if the composition contains an anti-PVRIG antibody and / or antigen-binding moiety that is present in at least 10% less aggregated state after exposure to mechanical stress compared to a composition of anti-PVRIG antibody and / or antigen-binding moiety that does not contain a stabilizer or contains a small amount of a stabilizer. In some embodiments, a composition of anti-PVRIG antibody and / or antigen-binding moiety is considered stabilized by the addition of a stabilizer (e.g., at least one amino acid, salt, or nonionic surfactant) if the composition contains anti-PVRIG antibody and / or antigen-binding moiety that, after exposure to mechanical stress, is present in an aggregated state by at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, or more, compared to a composition of anti-PVRIG antibody and / or antigen-binding moiety that contains no stabilizer or a small amount of stabilizer. In certain embodiments, the mechanical stress is agitation (e.g., shaking).

[0165] In some embodiments, the highly stabilized formulations of the present invention have a shelf life of at least 6 months. As understood, this shelf life may be in liquid or lyophilized form at freezing temperatures (i.e., -80°C, -25°C, 0°C), refrigeration (0°C to 10°C), or room temperature (20°C to 32°C). In further embodiments, the highly stabilized formulations of the present invention have a shelf life of at least 12, 18, 24, 30, 36, 42, 48, 54, or 60 months.

[0166] In some embodiments, shelf life is determined by the percentage of activity remaining after storage at any of the above temperatures for any of the above periods. In some embodiments, shelf life is determined by any of the assays described herein or known in the art, compared to the activity before the formulation is stored at any of the above temperatures for any of the above time periods, as measured by any of the assays described herein or known in the art, at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85% This means that it retains %, 90%, 95%, 96%, 97%, 98%, 99%, and 100% of the furin activity.

[0167] In some embodiments, the present invention is (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody contains an antibody having the same CDR as shown in Figure 3, (b) 25 mM histidine and (c) 60 mM NaCl and (d) 100 mM L-arginine and (e) To provide a stable liquid pharmaceutical formulation of an anti-PVRIG antibody containing 0.01 w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0168] In some embodiments, the anti-PVRIG antibody is at a concentration of 10 mg / mL to 40 mg / mL, 15 mg / mL to 40 mg / mL, 15 mg / mL to 30 mg / mL, 10 mg / mL to 25 mg / mL, or 15 mg / mL to 25 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 10 mg / mL to 40 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 15 mg / mL to 40 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 15 mg / mL to 30 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 10 mg / mL to 25 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 15 mg / mL to 25 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 10 mg / mL to 25 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 15 mg / mL to 25 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of 20 mg / mL to 25 mg / mL. In some embodiments, the anti-PVRIG antibody is at a concentration of approximately 20 mg / mL.

[0169] In some embodiments, the present invention is (a) an anti-PVRIG antibody comprising an antibody having the same CDR as shown in Figure 3, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Provide a stable liquid pharmaceutical formulation of an anti-PVRIG antibody containing 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0. B. Amino Acids In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or its antigen-binding fragment containing at least one amino acid (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3). In some embodiments, the at least one amino acid is histidine. In some embodiments, the at least one amino acid is arginine. In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or its antigen-binding fragment containing at least two amino acids. In some embodiments, the at least two amino acids are histidine and arginine.

[0170] In some embodiments, the pharmaceutical formulation contains 10 mM to 80 mM histidine, 15 mM to 70 mM histidine, 20 mM to 60 mM histidine, and 20 mM to 50 mM histidine. The formulation contains histidine, or 20 mM to 30 mM histidine. In some embodiments, the pharmaceutical formulation contains 10 mM to 80 mM histidine. In some embodiments, the pharmaceutical formulation contains 15 mM to 70 mM histidine. In some embodiments, the pharmaceutical formulation contains 20 mM to 60 mM histidine. In some embodiments, the pharmaceutical formulation contains 20 mM to 50 mM histidine. In some embodiments, the pharmaceutical formulation contains 20 mM to 30 mM histidine. In some embodiments, the pharmaceutical formulation contains about 25 mM histidine.

[0171] In some embodiments, the pharmaceutical formulation contains 10 mM to 80 mM histidine. In some embodiments, the pharmaceutical formulation contains 15 mM to 70 mM histidine. In some embodiments, the pharmaceutical formulation contains 20 mM to 60 mM histidine. In some embodiments, the pharmaceutical formulation contains 20 mM to 50 mM histidine. In some embodiments, the pharmaceutical formulation contains 20 mM to 30 mM histidine. In some embodiments, the pharmaceutical formulation contains about 25 mM histidine.

[0172] In some embodiments, the pharmaceutical formulation contains L-arginine in concentrations of 20 mM to 140 mM, 30 mM to 140 mM, 40 mM to 130 mM, 50 mM to 120 mM, 60 mM to 110 mM, 70 mM to 110 mM, 80 mM to 110 mM, or 90 mM to 110 mM. In some embodiments, the pharmaceutical formulation contains L-arginine in concentrations of 20 mM to 140 mM, 30 mM to 140 mM, 40 mM to 130 mM, 50 mM to 120 mM, 60 mM to 110 mM, 70 mM to 110 mM, 80 mM to 110 mM, or 90 mM to 110 mM.

[0173] In some embodiments, the pharmaceutical formulation contains 20 mM to 140 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 30 mM to 140 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 40 mM to 130 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 50 mM to 120 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 60 mM to 110 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 70 mM to 110 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 80 mM to 110 mM L-arginine. In some embodiments, the pharmaceutical formulation contains 90 mM to 110 mM L-arginine. In some embodiments, the pharmaceutical formulation contains about 100 mM L-arginine. C. Sugars and / or sugar alcohols In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or an antigen-binding fragment thereof (e.g., an anti-PVRIG antibody having the same CDRs as those shown in FIG. 3) that does not contain sugars and / or sugar alcohols. In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or an antigen-binding fragment thereof (e.g., an anti-PVRIG antibody having the same CDRs as those shown in FIG. 3) that does not contain sugars. In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or an antigen-binding fragment thereof (e.g., an anti-PVRIG antibody having the same CDRs as those shown in FIG. 3) that does not contain sugar alcohols.

[0174] In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or an antigen-binding fragment thereof that contains sugars and / or sugar alcohols. In some embodiments, the sugar is trehalose or sucrose. In some embodiments, the sugar is trehalose. In some embodiments, the sugar is sucrose. In some embodiments, the sugar is only one of trehalose or sucrose, not both.

[0175] In some embodiments, the sugar is present in an amount of about 0.5% to 10%, 1% to 9.5%, 1.5% to 9%, 2.0% to 8.5%, 2.5% to 8%, 3.0% to 7.5%, 3.5% to 7%, 4.0% to 6.5%, 4.5% to 6%, and / or 4.5% to 5.5%. In some embodiments, the sugar is present in an amount of about 0.5% to 10%. In some embodiments, the sugar is present in an amount of about 1% to 9.5%. In some embodiments, the sugar is present in an amount of about 1.5% to 9%. In some embodiments, the sugar is present in an amount of about 2.0% to 8.5%. In some embodiments, the sugar is present in an amount of about 2.5% to 8%. In some embodiments, the sugar is present in an amount of about 3.0% to 7.5%. In some embodiments, the sugar is present in an amount of about 3.5% to 7%. In some embodiments, the sugar is present in an amount of about 4.0% to 6.5%. In some embodiments, the sugar is present in an amount of about 4.5% to 6%. In some embodiments, the sugar is present in an amount of about 4.5% to 5.5%. In some embodiments, the sugar is present in an amount of about 5%. D. Nonionic Surfactant In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or an antigen-binding fragment thereof (e.g., an anti-PVRIG antibody having the same CDRs as those shown in FIG. 3) containing a nonionic surfactant. In some embodiments, the storage-stable composition of the anti-PVRIG antibody or antigen-binding fragment contains a nonionic surfactant selected from nonionic water-soluble monoglycerides, nonionic water-soluble diglycerides, nonionic water-soluble triglycerides, nonionic water-soluble monofatty acid esters of polyethylene glycol, nonionic water-soluble difatty acid esters of polyethylene glycol, nonionic water-soluble sorbitan fatty acid esters, nonionic polyglycolated glycerides, nonionic water-soluble triblock copolymers, and combinations thereof. In some embodiments, the nonionic surfactant is polysorbate 80 (polyoxyethylene (20) sorbitan monooleate).

[0176] In some embodiments, the stable liquid pharmaceutical formulation comprises 0.006% to 0.1 w / v% polysorbate 80, 0.007% to 0.09 w / v% polysorbate 80, 0.008% to 0.08 w / v% polysorbate 80, 0.009% to 0.09 w / v% polysorbate 80, 0.01% to 0.08 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, or 0.01% to 0.06 w / v% polysorbate 80, or 0.009% to 0.05% w / v polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.006% to 0.1 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.007% to 0.09 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.008% to 0.08 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.009% to 0.09 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.01% to 0.08 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.01% to 0.07 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.01% to 0.07 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.01% to 0.06 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains 0.009% to 0.05 w / v% polysorbate 80. In some embodiments, the stable liquid pharmaceutical formulation contains about 0.01% polysorbate 80. E. Pharmaceutically acceptable salts In some embodiments, the present invention provides stable liquid pharmaceutical formulations of anti-PVRIG antibodies or their antigen-binding fragments (e.g., anti-PVRIG antibodies having the same CDR as shown in Figure 3) that include a salt, for example, a pharmaceutically acceptable salt.

[0177] In some embodiments, a stable liquid pharmaceutical formulation comprising an anti-PVRIG antibody or its antigen-binding fragment provided herein contains a pharmaceutically acceptable salt at a concentration acceptable by the anti-PVRIG antibody or its antigen-binding fragment during storage. In some embodiments, the pharmaceutically acceptable salt is a chloride salt. In some embodiments, the pharmaceutically acceptable salt is a monovalent chloride salt. In more specific embodiments, the pharmaceutically acceptable salt is sodium chloride, potassium chloride, or a combination thereof.

[0178] In some embodiments, the stable liquid pharmaceutical formulation contains 30 mM to 100 mM NaCl, 30 mM to 90 mM NaCl, 40 mM to 80 mM NaCl, 30 mM to 70 mM histidine, or 45 mM to 70 mM NaCl.

[0179] In some embodiments, the stable liquid pharmaceutical formulation contains 30 mM to 100 mM NaCl. In some embodiments, the stable liquid pharmaceutical formulation contains 30 mM to 90 mM NaCl. In some embodiments, the stable liquid pharmaceutical formulation contains 40 mM to 80 mM NaCl. In some embodiments, the stable liquid pharmaceutical formulation contains 30 mM to 70 mM histidine. In some embodiments, the stable liquid pharmaceutical formulation contains 45 mM to 70 mM NaCl. In some embodiments, the pharmaceutical formulation contains about 60 mM NaCl. F. Buffering agent In some embodiments, the present invention provides a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or its antigen-binding fragment (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3) buffered at a physiologically acceptable pH. In some embodiments, the physiologically acceptable pH is about 6.0 to about 7.0.

[0180] In some embodiments, a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or its antigen-binding fragment has a pH of 6 to 7.0. In some embodiments, a stable liquid pharmaceutical formulation of an anti-PVRIG antibody or its antigen-binding fragment has a pH of 6, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, or 7.0. In some embodiments, the pH is 6.1 to 6.9. In some embodiments, the pH is 6.2 to 6.9. In some embodiments, the pH is 6.3 to 6.8. In some embodiments, the pH is 6.3 to 6.7. In some embodiments, the pH is 6.4 to 6.8. In some embodiments, the pH is 6.5 to 6.8. In some embodiments, the pH is 6.6 to 6.8. In some embodiments, the pH is 6.3, 6.4, 6.5, 6.6, or 6.7. In some embodiments, the pH is 6.5 + / - 0.2. G. Method for diluting aqueous compositions [In some embodiments, the method includes adding a dilution buffer to form a diluted, stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment (for example, an anti-PVRIG antibody having the same CDR as shown in Figure 3). In some embodiments, the dilution buffer is added in a ratio of 1:1 (dilution buffer: formulation) to 1000:1 (dilution buffer: formulation). In other embodiments, the dilution buffer is added in a ratio of 1:1 (dilution buffer: formulation) to 500:1 (dilution buffer: formulation). In other embodiments, the dilution buffer is added in a ratio of 1:1 (dilution buffer: formulation) to 250:1 (dilution buffer: formulation). The dilution buffer is added in the ratio of (dilution buffer:formulation). In another embodiment, the dilution buffer is added in a ratio of 1:1 (dilution buffer:formulation) to 200:1 (dilution buffer:formulation). In another embodiment, the dilution buffer is added in a ratio of 1:1 (dilution buffer:formulation) to 100:1 (dilution buffer:formulation). In another embodiment, the dilution buffer is added in a ratio of 1:1 (dilution buffer:formulation) to 50:1 (dilution buffer:formulation).

[0181] In some embodiments, a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment is available in dilutions of 1x to 1000x, 1x to 500x, 1x to 250x, 1x to 200x, 1x to 100x, 1x to 50x, 1x to 10x, 10x to 1000x, 10x to 500x, 10x to 250x, 10x to 200x, and 10x. It is diluted to ~100 times, 10 to 50 times, 50 to 1000 times, 50 to 500 times, 50 to 250 times, 50 to 200 times, 50 to 100 times, 100 to 1000 times, 100 to 500 times, 100 to 250 times, 100 to 200 times, 200 to 1,000 times, 200 to 500 times, or 200 to 250 times. H. Stability assay As discussed herein, stable liquid pharmaceutical formulations containing an anti-PVRIG antibody or its antigen-binding fragment (e.g., an anti-PVRIG antibody having the same CDR as shown in Figure 3) exhibit improved stability compared to a control formulation. In one embodiment, the improved stability includes retention of a higher percentage of binding activity and / or no decrease in binding activity compared to a control formulation in various stability assays. Such assays can be used to determine whether a formulation is a highly stabilized formulation. In some embodiments, a highly stabilized formulation has at least 5%, 10%, 20%, 30%, 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or greater activity than a control formulation when evaluated by any of the stability assays discussed herein or known in the art.

[0182] In some embodiments, liquid pharmaceutical formulations containing an anti-PVRIG antibody or its antigen-binding fragment are tested under stressor conditions such as storage at high temperatures, agitation, freeze / thaw cycles, or some combination thereof. After such stressors, the formulations are assayed using any of the methods described herein or known in the art to determine their stability under these conditions. A280 and visual analysis In some embodiments, the appearance of a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment can be examined using A280 by the SoloVPE assay.

[0183] In some embodiments, the SoloVPE assay can be used to determine the concentration of a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment.

[0184] A280: Amino acids containing aromatic side chains exhibit strong ultraviolet absorption at a wavelength of 280 nm. Once the extinction coefficient of a given protein is established, the concentration of the protein in solution can be calculated from its absorbance. This method is designed to determine the protein concentration by measuring the absorbance at 280 nm using a SoloVPE instrument without dilution (https: / / www.ctechnologiesinc.com / products / solovpe).

[0185] Appearance: The appearance of the sample is determined by holding the sample in a controlled light source and observing the material's appearance. The solution is evaluated by gently stirring it and determining whether its appearance changes when viewed against a black and white background. Clarity is described using adjectives such as "transparent," "turbid," or "slightly turbid." The color of the material is specifically described. If the material is colorless, this is stated as a result (i.e., a transparent, colorless solution). The physical state of the sample is clearly described (i.e., liquid or frozen liquid). Binding assay analysis In some embodiments, a binding assay can be performed to investigate the activity of a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment. LabChip analysis In some embodiments, LabChip analysis is used to examine the purity of a stable liquid pharmaceutical formulation containing, for example, an anti-PVRIG antibody or its antigen-binding fragment, including IgG purity and HC+LC ratio. In some embodiments, the stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment exhibits an IgG purity of over 94%, over 95%, over 96%, over 97%, or over 98%. In some embodiments, the stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment exhibits an IgG purity of approximately 95% to 98%. In some embodiments, the stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment exhibits an IgG purity of approximately 96% to 97%. In some embodiments, the stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment exhibits an HC+LC ratio of approximately 96% to 100%. In some embodiments, the stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment exhibits an HC+LC ratio of approximately 97% to 100%. In some embodiments, a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment exhibits an HC+LC ratio of approximately 98% to 100%. cIEF analysis In some embodiments, capillary isoelectric focusing (cIEF) is used to analyze a stable liquid pharmaceutical formulation containing an anti-PVRIG antibody or its antigen-binding fragment for the presence of additional species, such as minor acidic species. MFI analysis Antibodies can form invisible particles in response to stress conditions such as heat, freeze / thaw cycles, and agitation. In some embodiments, microfluidic imaging (MFI) analysis can be used to analyze stable liquid pharmaceutical formulations containing anti-PVRIG antibodies or their antigen-binding fragments for particle formation in response to stress conditions. In some embodiments, the stable liquid pharmaceutical formulation of anti-PVRIG antibodies or their antigen-binding fragments provides a formulation in which the anti-PVRIG antibody or its antigen-binding fragment is stabilized against these stress conditions, preventing particle formation. Using MFI, particle counts in different size ranges (<2 μm, <5 μm, <10 μm, and <25 μm) can be evaluated in different formulations under stress conditions. Typically, MFI data can be evaluated to select a suitable formulation based on the minimum particle / mL generation for all particle sizes across all time points, conditions, and formulations. SEC analysis In some embodiments, size exclusion chromatography (SEC) can be used to analyze stable liquid pharmaceutical formulations containing anti-PVRIG antibodies or their antigen-binding fragments. SEC data showed high molecular weight (HMW) throughout all time points and conditions. However, it remained stable at approximately 1%. Low molecular weight (LMW) was present at 8 weeks under accelerated conditions and at 2–8°C. Under conditions below 40°C, LMW was approximately 1–3% from week 1 to week 2. It increased to [a certain value]. I. Embodiments of the selected formulation In some embodiments, the present invention is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM of NaCl, and (d) 20 mM to 150 mM of L - arginine, and (e) 0.005% to 0.1 w / v% of polysorbate 80, to provide a stable liquid pharmaceutical formulation of an anti - PVRIG antibody, The composition has a pH of 5.5 to 7.0.

[0186] In some embodiments, the present invention (a) an anti - PVRIG antibody, i) a heavy - chain variable domain comprising vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), and ii) a light - chain variable domain comprising vlCDRⅠ, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), an anti - PVRIG antibody comprising, (a) an anti - PVRIG antibody, and (b) about 25 mM of histidine, and (c) about 60 mM of NaCl, and (d) about 100 mM of L - arginine, and (e) about 0.01 w / v% of polysorbate 80, to provide a stable liquid pharmaceutical formulation of an anti - PVRIG antibody, The composition has a pH of 6.5 + / - 0.2.

[0187] In some embodiments, the present invention (a) an anti - PVRIG antibody, i) the heavy - chain variable domain is from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) the light - chain variable domain is from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), an anti - PVRIG antibody, and (b) 10 mM to 100 mM of histidine, and (c) 30 mM to 100 mM of NaCl, and (d) 20 mM to 150 mM of L - arginine, and (e) Provide a stable liquid pharmaceutical formulation containing 0.005% to 0.1 w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0188] In some embodiments, the present invention is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) The light chain variable domain is the light chain of CHA.7.518.1.H4(S241P) From Sequence ID No. 9), the anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) Provide a stable liquid pharmaceutical formulation containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0189] In some embodiments, the present invention is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Provide a stable liquid pharmaceutical formulation containing 0.005% to 0.1 w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0190] In some embodiments, the present invention is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) Provide a stable liquid pharmaceutical formulation containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0191] In some embodiments, the present invention is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Provide a stable liquid pharmaceutical formulation containing 0.005% to 0.1 w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0192] In some embodiments, the present invention is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) Provide a stable liquid pharmaceutical formulation containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2. VI. Administration of anti-PVRIG antibody preparations The pharmaceutical composition containing the anti-PVRIG antibody of the present invention (e.g., an anti-PVRIG antibody having the same CDR as shown in Figure 3), preferably in the form of a sterile aqueous solution, can be administered by various methods. As is known in the art, protein-based therapeutics are often delivered by IV infusion. The antibody of the present invention can also be delivered using such a method. For example, administration may be by intravenous infusion using 0.9% sodium chloride as the infusion vehicle. Such a technique is disclosed in Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed., 1980.

[0193] The dosage and frequency of administration are selected in some embodiments to be therapeutically or prophylactically effective. As is readily known in the art, adjustments may be necessary for the rate of proteolysis, systemic versus local delivery, and novel protease synthesis, as well as for age, weight, overall health, sex, diet, administration time, drug interactions, and severity of condition, which can be verified by those skilled in the art using conventional experiments. To treat a patient, a therapeutically effective dose of the Fc variant of the present invention may be administered. As used herein, “therapeutic dose” means the dose that produces the effect for which it was administered. VII. Dosage In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety of the present invention may be formulated for administration, for example, as a unit dose formulation. In some embodiments, the formulation of the anti-PVRIG antibody and / or its antigen-binding moiety is administered at a dose of 0.01 mg / kg of the anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the formulation of the anti-PVRIG antibody and / or its antigen-binding moiety is administered at a dose of 0.02 mg / kg of the anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the formulation of the anti-PVRIG antibody and / or its antigen-binding moiety is administered at a dose of 0.03 mg / kg of the anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the formulation of the anti-PVRIG antibody and / or its antigen-binding moiety is administered at a dose of 0.04 mg / kg of the anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.05 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, anti-PV The RIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.06 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.07 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.08 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.09 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.1 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.2 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.3 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.5 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 0.8 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 1 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 2 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 3 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety.In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 4 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 5 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 6 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 7 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 8 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 9 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 10 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of 20 mg / kg of anti-PVRIG antibody and / or its antigen-binding moiety.

[0194] In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of approximately 0.01 mg / kg to approximately 20 mg / kg of anti-PVRIG antibody. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of approximately 0.01 mg / kg to approximately 10 mg / kg of anti-PVRIG antibody. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of approximately 20 mg / kg. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered at a dose of approximately 20 mg / kg every four weeks. The formulation is administered in the following doses. In some embodiments, the anti-PVRIG antibody and / or its antigen-binding moiety formulation is administered intravenously at a dose of approximately 20 mg / kg every four weeks. In some embodiments, the formulation is administered in doses of anti-PVRIG antibody ranging from approximately 0.1 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered in doses of anti-PVRIG antibody ranging from approximately 1 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered in doses of anti-PVRIG antibody ranging from approximately 2 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered in doses of anti-PVRIG antibody ranging from approximately 3 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered in doses of anti-PVRIG antibody ranging from approximately 4 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered in doses of anti-PVRIG antibody ranging from approximately 5 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered at a dose of anti-PVRIG antibody of approximately 5 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered at a dose of anti-PVRIG antibody of approximately 7 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered at a dose of anti-PVRIG antibody of approximately 8 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered at a dose of anti-PVRIG antibody of approximately 9 mg / kg to approximately 10 mg / kg. In some embodiments, the formulation is administered at a dose of anti-PVRIG antibody of approximately 20 mg / kg. In some embodiments, the formulation is administered at a dose of anti-PVRIG antibody of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg. A. Selected medication according to the formulation embodiment In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0195] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (a) Anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0196] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0197] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0198] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0199] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0200] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0201] In some embodiments, the present invention provides the administration of a stable liquid pharmaceutical formulation of anti-PVRIG antibody, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0202] In some embodiments of stable liquid pharmaceutical formulations, the formulation is administered together with an anti-PD-1 antibody.

[0203] In some embodiments of stable liquid pharmaceutical formulations, the anti-PD-1 antibody is an antibody selected from the group consisting of pembrolizumab and nivolumab.

[0204] In some embodiments of the stable liquid pharmaceutical formulation, the anti-PD-1 antibody is nivolumab. In some embodiments of the stable liquid pharmaceutical formulation, the anti-PD-1 antibody is nivolumab and is administered in doses of approximately 360 mg or 480 mg. In some embodiments of the stable liquid pharmaceutical formulation, the anti-PD-1 antibody is nivolumab and is administered in doses of approximately 360 mg. In some embodiments of the stable liquid pharmaceutical formulation, the anti-PD-1 antibody is nivolumab and is administered in doses of approximately 480 mg.

[0205] In some embodiments of stable liquid pharmaceutical formulations, the anti-PD-1 antibody is pembrolizumab. VIII. How to use anti-PVRIG antibody preparations A. Therapeutic use Anti-PVRIG antibodies (for example, anti-PVRIG antibodies having the same CDR as shown in Figure 3) are generally used to treat patients, such as human subjects, who have a PVRIG-related condition. As used herein, the term “treatment” refers in this example to both therapeutic treatments and preventive or preventive measures for cancer, but as also described below, the use of antibodies and pharmaceutical compositions also provides treatments for infectious diseases, sepsis, and / or autoimmune conditions, as well as inhibition of undesirable immune activation following gene therapy. Those requiring treatment include those who already have cancer and those for whom cancer should be prevented. Thus, the mammals being treated herein may be diagnosed with cancer, or may be prone to or susceptible to cancer. As used herein, the term “to treat” means to prevent adverse effects, delay their onset, cure, reverse, reduce, mitigate, minimize, suppress, stop, or stabilize the recognizable symptoms of the cancerous disease, disorder, or condition described above. This also includes managing cancer as described above. "To manage" means reducing the severity of a disease, reducing the frequency of disease attacks, reducing the duration of such attacks, reducing the severity of such attacks, delaying / reducing cancer cell growth or proliferation, delaying the progression of at least one symptom, or alleviating at least one measurable physical parameter. For example, an immunostimulatory anti-PVRIG immune molecule is supposed to treat cancer or infectious diseases by promoting T cell or NK or cytokine immunity against target cells, e.g., cancer, infected, or pathogenic cells, thereby depleting the cells involved in the disease state. Conversely, an immunosuppressive anti-PVRIG immune molecule is supposed to reduce T cell or NK activity and / or the secretion of inflammatory cytokines involved in the disease pathology of certain immune disorders, such as autoimmune, inflammatory, or allergic conditions, thereby treating or alleviating disease pathology and tissue destruction that may be associated with such conditions (e.g., joint destruction associated with rheumatoid arthritis).

[0206] The PVRIG antibody of the present invention is provided in a therapeutically effective dose. The "therapeutically effective dose" of the anti-PVRIG immune molecule according to at least some embodiments of the present invention preferably reduces the severity of disease symptoms, increases the frequency and duration of disease-free periods, increases survival time, and induces disease remission. or prevent or reduce functional or physical impairment resulting from the suffering of the disease. For example, in the treatment of PVRIG-positive tumors, the “therapeutic effective dose” preferably inhibits cell growth or tumor growth by at least about 20%, more preferably at least about 40%, even more preferably at least about 60%, and still more preferably at least about 80% compared to an untreated subject. The ability of a compound to inhibit tumor growth can be evaluated in animal model systems to predict efficacy in human tumors. Alternatively, this property of a composition can be evaluated by examining the ability of the compound to inhibit, such in vitro inhibition, by assays known to those skilled in the art. A therapeutic effective dose of the therapeutic compound can reduce tumor size or, in other ways, alleviate symptoms in a subject.

[0207] Those skilled in the art will be able to determine the effective therapeutic dose based on factors such as the size of the subject, the severity of the subject's symptoms, and the specific composition or route of administration selected. 1. Cancer treatment The PVRIG antibody formulations of the present invention find outstanding use in the treatment of cancer. Generally, the antibodies of the present invention are immunomodulatory in that, rather than directly attacking cancer cells, the anti-PVRIG antibodies of the present invention stimulate the immune system by generally inhibiting the action of PVRIG. Thus, unlike tumor-targeted therapies that aim to inhibit molecular pathways essential for tumor growth and development, and / or deplete tumor cells, cancer immunotherapy aims to stimulate the patient's own immune system to eliminate cancer cells and provide sustained tumor destruction. Various approaches can be used in cancer immunotherapy, including therapeutic cancer vaccines to induce tumor-specific T cell responses, and immunostimulatory antibodies (i.e., antagonists of inhibitory receptors = immune checkpoints) to eliminate immunosuppressive pathways.

[0208] Clinical responses to targeted therapy or conventional anticancer therapy tend to be transient because cancer cells develop resistance and tumor recurrence occurs. However, clinical use of cancer immunotherapy in recent years has shown that this type of therapy can have a lasting clinical response and a dramatic impact on long-term survival. However, although the response is long-term, it is only experienced by a small number of patients (relative to conventional or targeted therapies, where many patients respond, but the response is transient).

[0209] By the time a tumor is clinically detected, it has already evaded the immune defense system by acquiring immune resistance and immunosuppressive properties, as well as by creating an immunosuppressive tumor microenvironment through various mechanisms and different types of immune cells.

[0210] Therefore, the anti-PVRIG antibody of the present invention is useful in the treatment of cancer. Due to the nature of its immuno-oncological mechanism of action, PVRIG does not necessarily need to be overexpressed on or correlated with a particular type of cancer; in other words, the goal is for the anti-PVRIG antibody to release the suppression of T cell and NK cell activation, allowing the immune system to target cancer.

[0211] As used herein, “cancer” broadly refers to any neoplastic disease (whether invasive or metastatic) characterized by abnormal and uncontrolled cell division that causes malignant growth or tumor (e.g., uncontrolled cell growth). The terms “cancer” or “cancerous” as used herein should be understood to encompass any neoplastic disease (whether invasive, non-invasive, or metastatic) characterized by abnormal and uncontrolled cell division that causes malignant growth or tumor, and these non-limiting examples are described herein. This includes any physiological condition in mammals that typically features uncontrolled cell proliferation.

[0212] In some embodiments, the anti-PVRIG formulations of the present invention can be used to treat solid tumors (e.g., including cancers of the lung, liver, breast, brain, and GI ducts) and hematological malignancies (e.g., leukemia and preleukemic disorders, lymphoma, plasmacytotoxicity), carcinomas, lymphomas, blastomas, sarcomas, and leukemia or lymphoid malignancies. In some embodiments, the cancer is in its early stages. In some embodiments, the cancer is advanced (including metastasis). In some embodiments, cancers suitable for treatment of the present invention include cancers that express or do not express PVRIG, and further include non-metastatic or non-invasive cancers as well as invasive or metastatic cancers, including cancers in which PVRIG expression by the immune system, stroma, or lesion cells suppresses the antitumor response and anti-invasive immune response. In some embodiments, the anti-PVRIG formulations can be used to treat angiogenic tumors. In some embodiments, cancers suitable for treatment using the anti-PVRIG formulations of the present invention include carcinomas, lymphomas, sarcomas, and / or leukemias. In some embodiments, cancers suitable for treatment using the anti-PVRIG formulations of the present invention include melanoma, non-melanoma skin cancer (squamous and basal cell carcinoma), mesothelioma, squamous cell carcinoma, lung cancer (small cell lung cancer, non-small cell lung cancer, soft tissue sarcoma, Kaposi's sarcoma, adenocarcinoma of the lung, squamous cell carcinoma of the lung, peritoneal cancer, esophageal cancer, hepatocellular carcinoma, liver cancer (including HCC), gastric cancer, stomach cancer (including gastrointestinal cancer), pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, urothelial carcinoma, bladder cancer, hepatocellular carcinoma, glioma, brain cancer (and edema such as that associated with brain tumors), breast cancer (e.g., tertiary thrombocytopenia negativitis), testicular cancer, testicular germ cell tumor, colon cancer, colorectal cancer (CRC), colorectal cancer MSS (MSS-CRC; including refractory MSS colorectal cancer);MSS (microsatellite stable state), primary peritoneal cancer, microsatellite-stable primary peritoneal cancer, platinum-resistant microsatellite-stable primary peritoneal cancer, CRC (MSS unknown), rectal cancer, endometrial cancer (including endometrial carcinoma), uterine carcinoma, salivary gland carcinoma, kidney cancer, renal cell carcinoma (RCC), renal cell carcinoma (RCC), prostate cancer, vulvar cancer, thyroid cancer, liver carcinoma, carcinoid carcinoma, head and neck cancer, B-cell lymphoma (non-Hodgkin lymphoma) This includes tumors, as well as low-grade / follicular non-Hodgkin lymphoma (NHL), small lymphocytic (SL) NHL, moderate / follicular NHL, moderate diffuse NHL, diffuse large B-cell lymphoma, high-grade immunoblastic NHL, high-grade lymphoblastic NHL, high-grade small non-incisional nuclear cell NHL, giant lesion NHL, mantle cell lymphoma, AIDS-associated lymphoma, and Waldenström macroglobulinemia, Hodgkin lymphoma (HD), chronic lymphocytes. Cholecystic leukemia (CLL), acute lymphoblastic leukemia (ALL), T-cell acute lymphoblastic leukemia (T-ALL), acute myeloid leukemia (AML), hairy cell leukemia, chronic myeloblastic leukemia, multiple myeloma, post-transplant lymphoproliferative disorder (PTLD), abnormal angiogenesis associated with nevus, Meigs syndrome, Merkel cell carcinoma, MSI high cancer, KRAS variant tumors, adult T-cell leukemia / lymphoma, adenoid cystic carcinoma (including adenoid cystic carcinoma), malignant melanoma, pancreatic cancer, pancreatic This includes adenocarcinoma, ovarian cancer (including ovarian carcinoma), pleural mesothelioma, neuroendocrine lung cancer (including pleural mesothelioma and neuroendocrine lung carcinoma), NSCL (large cell), NSCLC large cell adenocarcinoma, non-small cell lung cancer (NSCLC), NSCLC squamous cell carcinoma, cervical squamous cell carcinoma (cervical SCC), anal squamous cell carcinoma (anal SCC), neuroendocrine lung cancer, carcinoma of unknown primary origin, gallbladder cancer, malignant melanoma, pleural mesothelioma, and / or myelodysplastic syndrome (MDS).

[0213] In some embodiments, cancers suitable for treatment using the anti-PVRIG formulations of the present invention include prostate cancer, liver cancer (HCC), colorectal cancer (CRC), colorectal cancer MSS (MSS-CRC; including refractory MSS colorectal cancer), CRC (MSS of unknown origin), ovarian cancer (including ovarian carcinoma), endometrial cancer (including endometrial carcinoma), breast cancer, pancreatic cancer, gastric cancer, cervical cancer, head and neck cancer, thyroid cancer, testicular cancer, urothelial carcinoma, lung cancer, melanoma, non-melanoma skin cancer (squamous and basal cell carcinoma), glioma, renal cell carcinoma (RCC), renal cell carcinoma (RCC), lymphoma (non-Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HD)), acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia (T- This includes cancers selected from the group consisting of ALL), diffuse large B-cell lymphoma, testicular germ cell tumor, mesothelioma, esophageal cancer, trinegative breast cancer, Merkel cell carcinoma, MSI hypercarcinoma, KRAS variant tumor, adult T-cell leukemia / lymphoma, pleural mesothelioma, anal SCC, neuroendocrine lung cancer (including neuroendocrine lung carcinoma), NSCLC, NSCL (large cell), NSCLC large cell, NSCLC squamous cell, cervical SCC, malignant melanoma, pancreatic cancer, pancreatic adenocarcinoma, adenoid cystic carcinoma (including adenoid cystic carcinoma), primary peritoneal cancer, microsatellite-stable primary peritoneal cancer, platinum-resistant microsatellite-stable primary peritoneal cancer, and / or myelodysplastic syndrome (MDS).

[0214] In this specification, “cancer therapy” means any method for preventing or treating cancer or alleviating one or more of the symptoms of cancer. Typically, such therapy includes administering an immunostimulatory anti-PVRIG antibody (including an antigen-binding fragment) alone or in combination with chemotherapy, radiotherapy, or other biological agents, in order to enhance its activity, i.e., in an individual in whom PVRIG expression suppresses the antitumor response and the efficacy of chemotherapy, radiotherapy, or other biological agents.

[0215] In some embodiments, the anti-PVRIG antibody is used in combination with a PD-1 targeting antagonist antibody (e.g., an anti-PD-1 antibody), including but not limited to nivolumab and / or pembrolizumab. In some embodiments, the anti-PD-1 antibody is an antibody selected from the group consisting of nivolumab and pembrolizumab. In some embodiments, the anti-PD-1 antibody is nivolumab. In some embodiments, the anti-PD-1 antibody is pembrolizumab. In some embodiments, the anti-PD-1 antibody is nivolumab and is administered at 360 mg. In some embodiments, the anti-PD-1 antibody is nivolumab and is administered intravenously at 360 mg. In some embodiments, the anti-PD-1 antibody is nivolumab and is administered intravenously at 360 mg for 3 weeks (e.g., 360 mg IV for 3 weeks). In some embodiments, the anti-PD-1 antibody is nivolumab and is administered at 480 mg. In some embodiments, the anti-PD-1 antibody is nivolumab, administered intravenously at 480 mg. In some embodiments, the anti-PD-1 antibody is nivolumab, administered intravenously at 480 mg for 3 weeks (e.g., 480 mg IV for 3 weeks). In some embodiments, subjects administered anti-PVRIG antibody in combination with anti-PD-1 antibody have exhausted all available standard therapies, including but not limited to ECOG 0-1, previous anti-PD-1, previous anti-PD-L1, previous anti-CTLA-4, previous OX-40, and / or previous CD137 therapy. 2. Selected monotherapy treatments according to the formulation embodiments. In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0216] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (a) Anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0217] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0218] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0219] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0220] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0221] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain containing the heavy chain from CHA.7.518.1.H4(S241P) (Sequence Number) 8) and, ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0222] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of anti-PVRIG antibody to subjects in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2. 3. Selected combination therapies according to the formulation embodiment In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0223] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) vh from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P) Heavy chain variable domains including CDR1, vhCDR2, and vhCDR3, ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (a) Anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0224] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0225] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0226] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0227] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0228] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of nivolumab and an anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of the anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0229] In some embodiments, the present invention provides nivolumab and anti- The treatment of cancer is provided by administering a stable liquid pharmaceutical formulation of PVRIG antibody. The anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg. The stable liquid formulation of anti-PVRIG antibody is... (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0230] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0231] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0232] In some embodiments, the present invention involves administering 360 mg of Nivol to a subject in need of treatment. The treatment of cancer is provided by administering a stable liquid pharmaceutical formulation of mab and anti-PVRIG antibody. The anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg. The stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0233] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0234] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0235] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0236] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0237] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 360 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0238] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0239] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody comprising a light chain variable domain containing vlCDR1, vlCDR2, and vlCDR3 from the light chain (SEQ ID NO: 9) of CHA.7.518.1.H4(S241P), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0240] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0241] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) The heavy chain variable domain is derived from the heavy chain (sequence number 4) of CHA.7.518.1.H4(S241P), ii) An anti-PVRIG antibody whose light chain variable domain is derived from the light chain of CHA.7.518.1.H4(S241P) (SEQ ID NO: 9), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0242] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0243] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain, a) A heavy chain containing VH-CH1-hinge-CH2-CH3, where VH is from CHA.7.518.1.H4(S241P) (SEQ ID NO: 4) and the CH1-hinge-CH2-CH3 region is from IgG4, ii) A light chain, a) A light chain containing VL-CL, wherein VL is derived from CHA.7.518.1.H4(S241P) (SEQ ID NO: 9) and the CL region is derived from the human kappa-2 light chain, and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2.

[0244] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) Contains 0.005%~0.1w / v% polysorbate 80, The composition has a pH of 5.5 to 7.0.

[0245] In some embodiments, the present invention provides cancer treatment by administering a stable liquid pharmaceutical formulation of 480 mg of nivolumab and anti-PVRIG antibody to a subject in need of treatment, wherein the anti-PVRIG antibody is administered in doses of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg, and the stable liquid formulation of anti-PVRIG antibody is (a) an anti-PVRIG antibody, i) A heavy chain (sequence number 8) containing a heavy chain from CHA.7.518.1.H4(S241P), ii) A light chain (SEQ ID NO: 13) containing a light chain from CHA.7.518.1.H4(S241P), and an anti-PVRIG antibody containing, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl, (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The composition has a pH of 6.5 ± 0.2. [Examples]

[0246] Example 1: PVRIG antibody preparation test The 20 conditions shown in Figure 6 were prepared by adding the corresponding excipient to each buffer (A and B) containing the formulated PVRIG antibody. Each formulation was identified by its formulation ID.

[0247] The formulated materials were exposed to the stress and storage conditions shown in Figures 7A-7B. Pharmaceutical Research Procedure Steps Sampling requirements For each condition, one spare vial was added, requiring a total of two vials for the tests. One vial was needed for LabChip (reducing and non-reducing), cIEF, concentration (A280nm), efficacy assay, and SEC-HPLC analysis on CPS. The other vial was needed for visual appearance and MFI analysis. Except for the samples for MFI and appearance testing, appropriate samples were removed at the specified time and frozen below -60°C until analysis was started. MFI and appearance testing were performed immediately after removal.

[0248] Time zero (T0) samples of 20 formulated samples were taken from a stock of vials stored at 2–8°C and frozen below -60°C until analysis (or immediately for visual and MFI testing). Freezing / thawing research storage at temperatures below -60°C In each cycle, three vials per formulation (60 vials in total) were placed in a storage area below -60°C. After a minimum of 16 hours, all three vials per formulation (60 vials in total) were removed from the storage conditions below -60°C and warmed to room temperature for 3-5 hours until thawed.

[0249] After being placed in the freezer for cycle 3, the three vials were not allowed to thaw at room temperature until they were ready to begin the test. The samples were then allowed to return to room temperature before analysis.

[0250] The sample was assayed as described in this example. stirring Three vials from each of the 20 formulations (60 vials in total) were placed in a shaker rotating at approximately 200 rpm at room temperature and fixed in place. The vials were agitated for between 24 and 48 hours. Except for immediate MFI and visual inspection, all vials were stored frozen at below -60°C until analysis. Samples were equilibrated to room temperature before analysis.

[0251] The sample was assayed as described in this example. Store at 2-8°C for 8 weeks (tested at 0, 2, 4, and 8 weeks). Twelve vials were taken from each of the 20 formulations (a total of 240 vials, including T0 vials) and stored at 2–8°C. The two T0 vials were immediately analyzed for MFI and appearance. All other samples were labeled with temperature and time and stored frozen at below -60°C until analysis. Samples were returned to room temperature before analysis.

[0252] At each subsequent point in time, three vials were removed from each formulation from storage conditions of 2–8°C, labeled with temperature and time, and stored frozen at below -60°C until analysis, except for immediate MFI and visual inspection. The samples were returned to room temperature before analysis.

[0253] The sample was assayed as described in this example. Stored at ambient temperature for 4 weeks (25°C) (tested at 2 and 4 weeks). Six vials were taken from each of the 20 formulations (a total of 120 vials), and stored at ambient temperature (25°C).

[0254] At each point in time, three vials were taken from the ambient temperature storage area for each formulation, labeled with temperature and time, and frozen below -60°C until analysis, except for immediate MFI and visual inspection. Samples were allowed to return to room temperature before analysis.

[0255] The sample was assayed as described in this example. Store at 40°C for 2 weeks (tested at 1 and 2 weeks). Six vials were taken from each of the 20 formulations (a total of 120 vials), and stored at 40°C.

[0256] At each point in time, three vials were taken from the 40°C storage area for each formulation, labeled with temperature and time, and stored frozen at below -60°C until analysis, except for an immediate MFI visual test. The samples were returned to room temperature before analysis.

[0257] The sample was assayed as described in this example. Exam planning and schedule Sampling and testing were performed for each formulation condition. result All data from each formulation and time point are provided throughout the study and are shown in Figures 8–78). The results of this example are discussed. Each formulation was evaluated and compared to the conditions studied.

[0258] These figures graphically represent the key assay results that were edited and analyzed. The key assays analyzed to determine the appropriate formulation were SEC, cIEF, and MFI. High and low molecular weight SEC species were monitored throughout the study (Figures 11, 18, 25, 32, 39, 46, 53, 60, 67, and 74). cIEF results were obtained throughout the study (Figures 12, 19, 26, 33, 40, 47, 54, 61, 68, and 75). Finally, MFI particle / mL of various sizes were monitored throughout (Figures 13, 20, 27, 34, 41, 48, 55, 62, 69, and 76). Consideration A280 and visual analysis SoloVPE and visual inspection of A280 did not show significant changes over time and formulation and were not used to determine the final formulation.

[0259] During freeze / thaw analysis, SoloVPE yielded varying concentrations across all different formulations, exceeding the instrument's specifications. A spare vial was removed and the analysis repeated. However, the analysis still showed varying results. Therefore, the same sample was repeated using a 320nm correction for light scattering. The A280 result shows significantly less variation (Figure 34). The freeze / thaw data suggests that a 320nm correction may be necessary for this product after repeated freeze / thawing. This variation did not occur under other conditions. Since this product is being studied for long-term stability, a 320nm correction should be used when SoloVPE is used for concentration determination. Binding assay analysis The binding assay was evaluated, but it did not show significant changes in activity across conditions or formulations. The observed changes were within the range of method variability. Therefore, this method indicates that the molecule is stable with respect to binding activity. This method was not a critical assay for determining the formulation. LabChip analysis Evaluation of LabChip data showed that IgG purity and HC+LC percentages were fairly stable across time and conditions. IgG purity percentages ranged from 96% to 97%. The HC+LC ratio was in the range of 98-100%. Since there was no significant change in the results over time, this method was not used to determine the formulation. cIEF analysis cIEF analysis, yielding results at 40°C for one week, revealed the presence of additional minor acidic species in formulations A, B9, and B10. This led to their removal from the recommendations at this point in the study. However, the presence of this minor species was subsequently demonstrated at various levels under all accelerated conditions and from 2–8°C for four weeks onward. Therefore, the minor species' stability was monitored across all accelerated conditions when determining the appropriate formulation. MFI analysis Proteins can form invisible particles in response to stress conditions such as heat, freeze / thaw cycles, and agitation. An optimal formulation can stabilize the protein against these stress conditions and prevent particle formation. MFI was used to evaluate particle counts in different size ranges (<2 μm, <5 μm, <10 μm, and <25 μm) in different formulations under stress conditions. Evaluating MFI data allows for selection of the appropriate formulation based on the minimum particle / mL generated for all particle sizes across all time points, conditions, and formulations. SEC analysis SEC data showed HMW throughout all time points and conditions. However, it remained stable at approximately 1%. LMW was present at 8 weeks under accelerated conditions and at 2–8°C. Under conditions below 40°C, LMW increased from approximately 1%–3% from week 1 to week 2. This type should be monitored throughout the program and identified later by further characterization analysis. In determining the appropriate formulation, LMW and HMW were evaluated for stability across time points and conditions. conclusion Based on this data, it was determined that the buffer designated as B4 (25 mM histidine, 60 mM NaCl, 100 mM L-arginine, 0.01% PS 80, pH 6.5) was likely the final formulation. This formulation showed consistent SEC results and low HMW and LMW. In addition, MFI data indicated low particle volume / mL for all particle sizes. LabChip data showed that IgG purity and HC+LC ratio were stable in formulation B4 compared to T0. Therefore, the toxicology batch was formulated with this buffer. Example 2: PVRIG antibody preparation Description and composition of the injection solution The formulation is supplied as a sterile, preservative-free liquid dosage form at a concentration of 20 mg / mL in a clear borosilicate glass vial of type 10R, equipped with a gray bromobutyl rubber stopper and an aluminum flip-cap crimp. The vial is filled to a target volume of 10 mL. The formulation is stored and shipped frozen at -20°C. Before use, the vial is thawed at ambient temperature and mixed by gently rotating. For administration to patients, the formulation is diluted with 0.9% sodium chloride. Container closure system A single container closure system exists for the formulation, consisting of a 10R Type I clear borosilicate glass vial, a 20 mm bromobutyl rubber stopper, and a 20 mm aluminum flip cap crimp.

[0260] The formulation was prepared by thawing and pooling the active pharmaceutical ingredient, followed by sterile filtration through a 0.22 μm filter, and then filling it into sterile 10R glass vials using a Vetter filter.

[0261] The nominal formulation components and quantitative composition of the formulation per vial (10 mL) are shown in Table 1 below (see also B4 in Figures 6-77).

[0262] [Table 1]

[0263] A sufficient volume is filled into the vial based on the net filling weight, ensuring a recoverable volume of 10 mL. Example 3: Phase 1 study to evaluate anti-PVRIG antibodies in patients with progressive solid tumors. background: There is a high unmet medical need for the treatment (tx) of patients (pt) who are resistant to or subsequently relapse with checkpoint inhibitors. There is an urgent need for novel checkpoint therapies with new mechanisms of action that can activate T cells and demonstrate antitumor activity in this pre-treatment patient population. CHA.7.518.1.H4(S241P)(heavy chain: SEQ ID NO: 8; light chain: SEQ ID NO: 13) is a novel first-in-class humanized IgG4 monoclonal antibody that binds with high affinity to PVRIG (containing the poliovirus receptor-associated immunoglobulin domain) and blocks its interaction with its ligand, PVRL2. Both PVRIG and PVRL2 are part of the DNAM axis, as are TIGIT and PD1. Inhibition of PVRIG enhances T cell and NK cell activation, and PVRIG inhibits tumor growth in mouse tumor models. We assume that CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8; light chain: SEQ ID NO: 13) exhibits antitumor activity at pt, which is the pre-treatment stage for checkpoint inhibitors. method: This example describes an ongoing, open-label, first-in-human phase 1 study in patients with advanced solid tumors. The first part of this study (Group A) evaluates CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8; light chain: SEQ ID NO: 13) monotherapy IV Q3 weekly escalating dose in a single-pt cohort, initially 4, then 3+3 design. Key inclusion criteria: 18 years of age or older, histologically confirmed locally advanced / metastatic solid malignancy, and exhaustion of acceptable available standard treatments, ECOG 0-1, prior anti-PD-1, anti-PD-L1, anti-CTLA-4, OX-40, and CD137.

[0264] The primary exclusion criteria are active autoimmune disease requiring systemic therapy in the past two years, symptomatic interstitial or inflammatory lung disease, and untx or symptomatic central nervous system metastases. The primary objective is to identify the safety and tolerability of CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8; light chain: SEQ ID NO: 13) as measured by the incidence of adverse events (AEs) and dose-limiting toxicity (21-day DLT window), the pharmacokinetics of CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8; light chain: SEQ ID NO: 13), and the maximum tolerated dose and / or recommended expanded dose. A secondary objective is to characterize the immunogenicity and preliminary antitumor activity of CHA.7.518.1.H4(S241P) (heavy chain: SEQ ID NO: 8; light chain: SEQ ID NO: 13).

[0265] Statistical considerations: AEs graded according to CTCAE v4.03, responses according to RECIST v1.1. All analyses of research objectives are descriptive and hypothesis-generating. DLTs were not observed in single-pt cohorts. Assessment of pts enrolled in Cohort 5 is ongoing at the time of this submission. Example 4: Phase 1 study evaluating anti-PVRIG monotherapy and its combination with nivolumab in patients with advanced solid tumors. background: CHA.7.518.1.H4(S241P) is a novel first-in-class checkpoint inhibitor of the poliovirus receptor-associated immunoglobulin domain (PVRIG). It inhibits the binding of PVRIG to its ligand, PVRL2. The anti-PD-1 drug nivolumab is approved for pt with advanced malignancies (Nivolumab prescribing information: http: / / packageinserts.bms.com / pi / pi_opdivo.pdf, accessed 07 / 22 / 2019). The DNAM signaling axis, consisting of PVRL2, TIGIT, and DNAM, has been shown to play a role in regulating T / NK cell activity. PD-1 inhibitors also play an important role in this axis by regulating DNAM activation. Preclinical studies have shown that blocking PVRIG alone and in combination with PD-1 inhibitors activates T cells in the tumor microenvironment, thereby resulting in an anti-tumor immune response and inhibition of tumor growth. There is a high unmet medical need for novel ICIs as monotherapy for relapsed pt (premature tumor) following treatment with approved immune checkpoint inhibitors (ICIs), and for enhancing clinical response in combination with approved ICIs. While not theoretically bound, CHA.7.518.1.H4(S241P) is hypothesized to be safe, tolerable, and to show preliminary antitumor activity as monotherapy in pt with R / R solid tumors, and in combination with nivolumab. Previous reports indicate no DLTs (disruptive dose thromboembolic events) up to dose level 6 with CHA.7.518.1.H4(S241P) monotherapy (Phase I study evaluating CHA.7.518.1.H4(S241P) in patients with advanced solid tumors, J Clin Oncol). 37,2019(suppl;abstr TPS2657)). method: An ongoing Phase 1 dose-escalation study in a single-pt cohort and 3+3 study design of CHA.7.518.1.H4(S241P) as monotherapy IV Q3 weeks and in combination with nivolumab 360 mg IV Q3 weeks. Primary inclusion criteria: 18 years of age or older, histologically confirmed progressive solid tumor, acceptable available standard treatment, ECOG 0-1, prior exhaustion of anti-PD-1, anti-PD-L1, anti-CTLA-4, OX-40, and CD137. Primary exclusion criteria: active autoimmune disease requiring systemic therapy in the past two years, symptomatic interstitial or inflammatory lung disease, untx or symptomatic CNS metastases. Primary objective: To identify the safety and tolerability of CHA.7.518.1.H4(S241P) monotherapy and in combination with nivolumab, as measured by the incidence of adverse events (AEs) and dose-limiting toxicity (21-day DLT window), the pharmacokinetics of CHA.7.518.1.H4(S241P), and the maximum tolerated dose and / or recommended expansion dose as monotherapy / in combination with nivolumab. Secondary objective: To characterize the immunogenicity and preliminary antitumor activity of CHA.7.518.1.H4(S241P) in combination with nivolumab. Statistical considerations: AEs according to CTCAE v4.03, responses according to RECIST v1.1. The analysis of the study objectives is descriptive and hypothesis-generating. result: As of the date of this submission, no dose-limiting trials (DLTs) have been observed up to dose level 7 for CHA.7.518.1.H4(S241P) monotherapy and dose level 1 for CHA.7.518.1.H4(S241P) in combination with nivolumab. Conclusion: Safety and tolerability assessments are ongoing for all phases. Updated results will be analyzed throughout the entire clinical trial. Example 5: Phase 1 study of the safety, tolerability, and preliminary antitumor activity of CHA.7.518.1.H4(S241P) monotherapy in patients with advanced solid tumors. background: CHA.7.518.1.H4(S241P) is a novel first-in-class checkpoint inhibitor (ICI) of the poliovirus receptor-associated immunoglobulin domain (PVRIG) [1]. It inhibits the binding of PVRIG to its ligand, PVRL2. PVRIG is a member of the DNAM / TIGIT signaling axis that modulates T / NK cell activity. Preclinical studies have shown that PVRIG inhibition alone, as well as in combination with anti-PD-1 and / or TIGIT blockers, activates T cells in the tumor microenvironment, leading to an antitumor immune response and inhibition of tumor growth [1]. While ICIs have revolutionized cancer treatment, there is an urgent need to develop treatments for patients who are resistant to ICI therapy or who subsequently relapse. This study was designed to demonstrate that CHA.7.518.1.H4(S241P) is safe, tolerable, and exhibits preliminary antitumor activity. method: Dose escalation of CHA.7.518.1.H4(S241P) monotherapy utilizing a Phase 1a hybrid acceleration and 3+3 study design was performed to determine safety and tolerability, evaluate pharmacokinetics (PK) and pharmacodynamics, determine the recommended Phase 2 dose, and assess the preliminary antitumor activity of CHA.7.518.1.H4(S241P). Patients with advanced solid tumors that had failed standard treatment and a performance status of ECOG 0-1 were included. Prior ICIs were acceptable. CHA.7.518.1.H4(S241P) 0.01, 0.03, 0.1, 0.3, 1, 3, and 10 mg / kg IV every 3 weeks was administered until progression, to unacceptable toxicity, or until determined by the investigator or patient. Adverse events were reported according to CTCAE v4.03, and antitumor activity was reported according to REC. Evaluation was performed using IST v1.1. Dose-limiting toxicity (DLT) was assessed within a 21-day window. result: During dose escalation with CHA.7.518.1.H4(S241P), a total of 13 patients were enrolled and treated, of which 6 had metastatic colorectal cancer (CRC), 5 had microsatellite stable disease (MSS), and 1 was of unknown status. Patients were frequently prior to 7 previous anticancer therapies (range 2–15). No DLTs were reported up to a CHA.7.518.1.H4(S241P) dose level of 10 mg / kg. The most frequent toxicities were fatigue (8%) and abdominal pain (6%). Potential immune-related adverse events: elevated TSH and rash were observed in 2 patients. Overall, 7 out of 13 patients (54%), including 5 out of 6 patients (83%) with CRC, maintained the best response with stable disease (SD) for more than 12 weeks (13.6–43 weeks). Five patients are continuing study treatment. Peripheral PVRIG receptor occupancy (≥90%) was observed at CHA.7.518.1.H4(S241P) doses of 1 mg / kg or higher, and the PK profile supports IV Q3 weekly dosing. Conclusion: CHA.7.518.1.H4(S241P) monotherapy demonstrated an acceptable safety and tolerability profile with preliminary antitumor activity in a patient population that had previously received multiple anticancer therapies. References: 1.Spencer L,Ofer L et al,Discovery of COM701,a therapeutic antibody targeting the novel immune checkpoint PVRIG,for the treatment of cancer.J Clin Oncol.2017;(suppl;abstr 3074). Example 6: Data from an ongoing Phase 1 trial of CHA.7.518.1.H4(S241P) in patients with progressive solid tumors. CHA.7.518.1.H4(S241P) was well tolerated, and no dose-limiting toxicity was observed.

[0266] In the dose-escalation group of the study, initial signals of antitumor activity were frequently observed in the patient population that had received prior treatment.

[0267] Preliminary results from an ongoing Phase 1 dose-escalation study of CHA.7.518.1.H4(S241P), a first-in-class anti-PVRIG antibody, in patients with progressive solid tumors. CHA.7.518.1.H4(S241P) was well-tolerated and showed no dose-limiting toxicity. Furthermore, CHA.7.518.1.H4(S241P) showed an initial signal of antitumor activity in the frequently prior-treated patient population enrolled in the study.

[0268] The novel safety profile and initial antitumor activity of CHA.7.518.1.H4(S241P) were promising. The primary objective of this portion of the trial was to examine the safety and tolerability of CHA.7.518.1.H4(S241P) in the entire patient population, as well as the initial signaling of antitumor activity in patients with difficult-to-treat conditions, including those with microsatellite-stable colorectal cancer (MSS-CRC). We anticipate initiating an expanded cohort of biomarker-driven CHA.7.518.1.H4(S241P) monotherapy in patients with ovarian cancer, endometrial cancer, breast cancer, and lung cancer. HA.7.518.1.H4(S241P) can expand the prospects for checkpoint inhibitors in these indications, based on an understanding of the biological pathway of PVRIG.

[0269] Given the number of patients with advanced cancer who are unresponsive to or resistant to currently available therapies, there is an urgent need to expand the range of cancer immunotherapy agents to a broader patient population. The initial signal of antitumor activity of CHA.7.518.1.H4(S241P) is promising, especially considering the all-visiting patient population, many of whom are frequently prior-treated and resistant to previous therapies. A trend in dose-response relationships was observed in this difficult-to-treat patient population, and furthermore, a promising signal of antitumor activity was observed in 5 out of 6 patients with MSS colorectal cancer, a difficult indication that is typically unresponsive to current immune checkpoint blockers.

[0270] The reported data are from the monotherapy group of an ongoing Phase 1, open-label, dose-escalation study, including the first six cohorts (n=13) at dose levels of 0.01, 0.03, 0.1, 0.3, 1, 3, and 10 mg / kg IV, every three weeks. Main findings: CHA.7.518.1.H4(S241P) was well tolerated up to 10 mg / kg, and no dose-limiting toxicity was observed.

[0271] The best time-point response for stable disease (SD) / disease control rates was reported in 9 out of 13 patients (69%), based on a median of 7 prior anticancer therapies (ranging from 2 to 15).

[0272] All patients with CRC (N=6) were in a microsatellite stable state, and 5 out of 6 patients (83%) showed the best time-point response for a stable disease.

[0273] The pharmacokinetic profile supports weekly medication in IV Q3.

[0274] A peripheral PVRIG receptor occupancy rate of over 90% was observed with CHA.7.518.1.H4(S241P) at doses of 1 mg / kg or higher.

[0275] Three patients are continuing research treatment with CHA.7.518.1.H4(S241P) monotherapy.

[0276] Enrollment in monotherapy with CHA.7.518.1.H4(S241P) at 20 mg / kg per week (Q4) is ongoing. Regarding the Phase 1 study CHA.7.518.1.H4(S241P) A Phase 1 open-label clinical trial of CHA.7.518.1.H4(S241P) is designed to evaluate the safety and tolerability of escalating doses of CHA.7.518.1.H4(S241P) monotherapy and combination therapy with Bristol-Myers Squibb's Opdivo (registered commercial) in patients with advanced solid tumors. Additional secondary endpoints include preliminary antitumor activity, pharmacokinetics, and pharmacodynamics of CHA.7.518.1.H4(S241P) monotherapy and combination therapy with Opdivo in patients with selected tumor types, including non-small cell lung cancer, ovarian cancer, breast cancer, and endometrial cancer. The Phase 1 study, expected to enroll approximately 140 patients, is currently recruiting in the United States. Additional information can be found at www.clinicaltrials.gov(NTC03667716). Example 7: Phase 1 study of the safety, tolerability, and preliminary antitumor activity of CHA.7.518.1.H4(S241P) monotherapy in patients with advanced solid tumors. background CHA.7.518.1.H4(S241P) is a novel first-in-class checkpoint inhibitor (ICI) of the poliovirus receptor-associated immunoglobulin domain (PVRIG) discovered by Compugen's computer discovery program.[1] It inhibits the binding of PVRIG to its ligand, PVRL2.

[0277] PVRIG is a member of the DNAM / TIGIT signaling axis that regulates TMK cell activity. Preclinical studies have shown that inhibiting PVRIG activates T cells in the tumor microenvironment, leading to an antitumor immune response and inhibition of tumor growth.[2] There is an urgent need to develop treatments for patients who are resistant to current ICI therapy or who subsequently experience relapses.

[0278] We hypothesized that CHA.7.518.1.H4(S241P) is safe, tolerable, and exhibits preliminary antitumor activity as monotherapy in patients with advanced solid tumors. Main eligibility criteria inclusion ● Age 18 or older ● Locally advanced or metastatic solid malignant tumors have been histologically or cytologically confirmed, and all available standard treatments have been exhausted or are not candidates for available standard treatments. ●ECOG Performance Status 0-1 ● Previous immune checkpoint inhibitors are acceptable. ● Appropriate blood, liver, and kidney function exclusion ● Symptomatic interstitial lung disease or inflammatory interstitial pneumonia ● Untreated or symptomatic central nervous system metastases ● History of immune-related events that led to discontinuation of immunotherapy treatment result No dose-limiting toxicities were reported within the evaluated dose range of CHA.7.518.1.H4(S241P) (0.01–10 mg / kg).

[0279] No discontinuation of treatment due to adverse events was reported. Most of the TEAEs were G1-2. ● Frequent TEAEs were fatigue (46%), nausea (31%), and anxiety (23%), all of which were Grade 1-2; disease progression was Grade 5 (23%). ●Possible immune-related adverse events were rash (G1) and elevated TSH levels (G1). Serious adverse events were reported in 5 out of 13 patients. ●In the three patients, SAE was due to disease progression. All patients had stage IV disease at the start of the study, and 8 out of 13 (62%) showed the best response to PD to their last prior treatment (i.e., refractory disease) before enrolling in this study.

[0280] ● 5 out of 8 patients (63%) showed the best time-to-time response with SD, SD was confirmed in 1 out of 5 patients, and 2 patients are currently undergoing ongoing research and treatment. The best time-point response for SD / disease control rate was reported in 9 out of 13 patients (69%). Colorectal cancer was the most common tumor type registered, presenting in 6 out of 13 patients, and all 6 patients were in a microsatellite-stable state (MSS-CRC).

[0281] ● Disease control rate (SD) in 5 out of 6 patients (83%) with CRC ●SD (Stable Disease) (week 12) was confirmed in 4 out of 6 patients (67%) with CRC (Clinical Research Cognitive Respiratory Cognitive) status. ●Best time-point response to 12-week SD with pembrolizumab in patients with 11% DCR and MSS-CRC in medical history data [3] All three registered patients with CRC-kras mutations showed the best time-point response to SD, and SD was confirmed in 2 out of 3 patients. CHA.7.518.1.H4(S241P) Exposure levels are proportional to repeated dosing. Peripheral CHA7.518.1.H4(S241P) receptor occupancy mg / kg IV Q3 week conclusion CHA.7.518.1.H4(S241P) is well-acceptable as monotherapy. Disease control rate -9 / 13 patients (69%) Signaling of antitumor activity in pt with difficult-to-treat MSS-CRC and CRC with KRAS mutations. ●SD confirmed in 4 out of 6 people (67%) - MSS-CRC ●SD was confirmed in 2 out of 3 patients with CRC-kras mutations. Signals of the antitumor response: ● In patients with previously resistant diseases ● In patients previously treated with ICI Trends in dose-response relationships CHA.7.518.1.H4(S241P) Exposure dose proportional to permissible IV Q3 week medication mg / kg CHA.7.518.1.H4(S241P)IV Q3: 90% peripheral receptor occupancy in 3 weeks. Two patients are continuing with research treatment. Study enrollment is ongoing in Group A (CHA.7.518.1.H4(S241P) monotherapy) and Group B (CHA.7.518.1.H4(S241P) in combination with nivolumab).

[0282] Study NCT03667716 was conducted in collaboration with Bristol-Myers Squibb. That is References 1. Whelan S, Ophir E, et al.PVRIG and PVRL2 Are Induced in Cancer and Inhibit CD8+ T-cell Function.Cancer Immunol Res.2019 Feb;7(2):257-268. 2.Murter B,Pan X,et al.Mouse PVRIG Has CD8+ T Cell-Specific Contributory Functions and Dampens Antitumor Immunity.Cancer Immunol Res.2019 Feb;7(2):244-256. 3.Le DT,Uram JN,Wang H,et al.N Engl J Med.PD-1 Blockade in Tumors with Mismatch-Repair Deficiency.2015 Jun 25;372(26):2509-20. Example 8: Phase 1 study of CHA.7.518.1.H4(S241P) monotherapy and combination therapy with nivolumab in patients with advanced solid tumors. background CHA.7.518.1.H4(S241P) is a novel first-in-class humanized IgG4 monoclonal antibody that binds with high affinity to poliovirus receptor-associated immunoglobulin domain-containing (PVRIG) and blocks its interaction with its ligand, PVRL2 [1].

[0283] Nivolumab is an anti-PD-1 antibody approved for use in patients with several malignancies.[2]

[0284] PD-1 inhibitors play a key role in this axis by regulating DNAM activation [3].

[0285] Preclinical studies have shown that PVRIG inhibition alone, as well as in combination with anti-PD-1, activates T cells in the tumor microenvironment, leading to an antitumor immune response and inhibition of tumor growth.[1] While ICIs have revolutionized cancer treatment, there is an urgent need to develop treatments for patients who are resistant to ICI therapy or who subsequently experience relapses.

[0286] We assume that CHA.7.518.1.H4(S241P) is safe, tolerable, and exhibits antitumor activity in patients with R / R solid tumors. method NCT03667716 is an ongoing, open-label, first-in-human phase 1 study in patients with R / R solid tumors.

[0287] This report presents the first part of a study evaluating the safety and tolerability of CHA.7.518.1.H4(S241P) monotherapy IV Q3 weeks and dose-escalating combination therapy with nivolumab 360 mg IV Q3 weeks. Main Outcome Measures As monotherapy in patients with advanced solid tumors, and in combination with nivolumab Evaluate the safety profile of CHA.7.518.1.H4(S241P). Incidence of adverse events and dose-limiting toxicities (21-day DLT window) graded according to CTCAE v4.03 Identify the maximum tolerated dose and / or recommended dose expansion. To characterize the pharmacokinetic profile of CHA.7.518.1.H4(S241P) as monotherapy and in combination with nivolumab. Secondary outcome scale To characterize the immunogenicity of CHA.7.518.1.H4(S241P) alone and in combination with nivolumab. To evaluate the preliminary antitumor activity of CHA.7.518.1.H4(S241P) in combination with nivolumab (phase 1b only) in accordance with RECIST v1.1. Exploratory outcome measures To evaluate the preliminary antitumor activity of CHA.7.518.1.H4(S241P) as monotherapy. To evaluate all associations between DNAM axis members and clinical outcomes. To investigate evidence of PD effects mediated by CHA.7.518.1.H4(S241P) in the blood, both as monotherapy and in combination with nivolumab. Main selection criteria Age 18 or older A locally advanced or metastatic solid malignant tumor has been histologically or cytologically confirmed, and all available standard treatments have been exhausted or are not candidates for available standard treatments. ECOG Performance Status 0-1 Previous anti-PD-1, anti-PD-L1, anti-CTLA-4, OX-40, and CD137 formulations are acceptable. Appropriate blood, liver, and kidney function Main exclusion criteria Active autoimmune disease requiring systemic therapy in the two years prior to the initial administration of CHA.7.518.1.H4(S241P) Symptomatic interstitial lung disease or inflammatory interstitial pneumonia Untreated or symptomatic central nervous system metastases History of immune-related events leading to discontinuation of immunotherapy treatment Outbreak information No dose-limiting toxicity was observed at the seventh CHA.7.518.1.H4(S241P) monotherapy dose level or at previous dose levels (red box).

[0288] No dose-limiting toxicity was observed at the third CHA.7.518.1.H4(S241P) + nivolumab dosage level or at previous dosage levels (green box).

[0289] As of the date of this presentation, the eighth CHA.7.518.1.H4(S241P) monotherapy and the fourth CHA.7.518.1.H4(S241P) + nivolumab dosage levels are accepting enrollment on an IV Q4 week schedule.

[0290] Study NCT03667716 was conducted in collaboration with Bristol-Myers Squibb. References Spencer L,Ofer L et al,Discovery of COM701,a therapeutic antibody targeting the novel immune checkpoint PVRIG,for the treatment of cancer.J Clin Oncol.2017;(suppl;abstr 3074) Nivolumab package insert.http: / / packageinserts.bms.com / pi / pi_opdivo.pdf.Accessed 07 / 22 / 2019. Wang B, Zhang W et al.,Combination cancer immunotherapy targeting PD-1 and GITR can rescue CD8+ T cell dysfunction and maintain memory phenotype.Sci.Immunol.2018;Nov 2:3(29). Example 9: CHA.7.518.1.H4(S241P) demonstrates antitumor activity as monotherapy and in combination with nivolumab in patients with advanced malignant tumors. background Introduction: CHA.7.518.1.H4(S241P) is a novel first-in-class immune checkpoint inhibitor (ICI) that binds with high affinity to poliovirus receptor-associated immunoglobulin domain-containing (PVRIG), blocks its interaction with its ligand PVRL2, and modulates T / NK cell activity via the DNAM / TIGIT axis. In preclinical studies, inhibition of PVRIG alone, as well as in combination with anti-PD1 and / or TIGIT, inhibits tumor growth in the microenvironment, activates T cells, and elicits an antitumor response. method: A total of 28 patients with various cancer types (16 / 12 in groups A / B) were enrolled (including patients with various tumor types who had failed all available standard treatments). Sixteen patients (dose escalation with CHA.7.518.1.H4(S241P) monotherapy) were in group A, and 12 patients (dose escalation with CHA.7.518.1.H4(S241P) with nivolumab) were in group B. Hybrid acceleration (first 4 dose cohorts in group A) and 3+3 study design (cohorts 5-8 in group A and all cohorts in group B). Patients with advanced or metastatic solid tumors who had failed standard treatment and had a performance status of ECOG 0-1 were eligible. Prior ICIs were acceptable. In group A, patients with CHA.7.51 8.1.H4(S241P) monotherapy was administered at doses of 0.01, 0.03, 0.1, 0.3, 1, 3, and 10 mg / kg (all IV Q3 weeks) and 20 mg / kg (IV Q4 weeks). In group B, pt received 0.3, 1, or 3 mg / kg of CHA.7.518.1.H4(S241P) and nivolumab 360 mg IV q3 weeks (3 pt / dose cohort), and 3 pt received 10 mg / kg and nivolumab 480 mg IV q4 weeks. Treatment-related adverse events (TEAEs) were reported according to CTCAE v4.03, and responses were reported according to RECIST v1.1. Dose-limiting toxicity (DLT) was evaluated within a 21-day or 28-day window (for 3 or 4-week dosing schedules, respectively). The data cutoff date was January 23, 2020. result: The median number of previous anticancer therapies was 7 (range 2-15) in group A and 5 (range 2-9) in group B. No dose-limiting trials (DLTs) were reported in any dose cohort. Treatment was well tolerated, and no patients discontinued treatment due to toxicity. The most frequent TEAEs in group A were fatigue (46%), nausea (31%), and anxiety (23%), all of grade 1-2. In group B, patients with 4 or more pts experienced anemia, lower extremity edema, rash, and fatigue, mostly grade 1-2 (88%). In groups A and B, the partial response (PR) + stable disease (SD) rate was 57% (16 / 28). Note: In group A (CHA.7.518.1.H4(S241P) 20 mg / kg IV q 4 weeks): PR was confirmed in pts with primary peritoneal cancer that continued treatment for more than 15 weeks. In Group B, no PRs have been confirmed in patients with MSS-CRC receiving CHA.7.518.1.H4(S241P) 0.3 mg / kg and nivolumab. In Group A, a partial response was observed in a patient with microsatellite-stable primary peritoneal cancer enrolled in the 8th and last dose cohort, and the patient is continuing research treatment (over 15 weeks).

[0291] I am currently continuing treatment with 360mg IV for 3 weeks, and have been receiving treatment for over 34 weeks.

[0292] A total of 11 out of 28 patients are continuing with the study treatment, including 3 patients who have not yet reached the initial imaging assessment. In both treatment groups, time-point response / disease control rates of partial response and stable disease were reported in 16 out of 28 patients (57%). Conclusion: CHA.7.518.1.H4(S241P) is well tolerated as monotherapy and in combination with nivolumab in frequently various previously treated pts with advanced or metastatic solid tumors. CHA.7.518.1.H4(S241P) shows preliminary enhancement of antitumor activity with an objective response as monotherapy and in combination with nivolumab in difficult-to-treat tumor types (primary peritoneal, microsatellite-stable primary peritoneal cancer (MSS-primary peritoneal cancer or MSS-PPC), and microsatellite-stable colorectal cancer (MSS-CRC)). Example 10: CHA.7.518.1.H4(S241P) exhibits antitumor activity in patients with advanced malignant tumors, both as monotherapy and in combination with nivolumab. Introduction: There is a high unmet medical need for treating patients who are resistant to or subsequently relapse with checkpoint inhibitor therapy.

[0293] In a mouse tumor model, inhibition of poliovirus receptor-associated immunoglobulin domain-containing (PVRIG) promotes the activation of T cells and NK cells, thereby inhibiting tumor growth (Spencer L, Ofer L et al, Discovery of COM701, a therapeutic antibody targeting the novel.immune checkpoint PVRIG,for the treatment of cancer.J Clin Oncol.2017;(suppl;abstr 3074)).

[0294] CHA.7.518.1.H4(S241P) is a novel first-in-class humanized IgG4 monoclonal antibody that binds to PVRIG with high affinity and blocks its interaction with its ligand, PVRL2.

[0295] Previous data supported preliminary antitumor activity of CHA.7.518.1.H4(S241P) monotherapy (Dumbrava E, Fleming G, Hamilton E et al. Journal for ImmunoTherapy of Cancer 2019, 7(Suppl 1):P421. SITC Nov 2019.) The current data provides preliminary safety and antitumor activity data for CHA.7.518.1.H4(S241P) in combination with nivolumab (Group B) and provides updated data for the CHA.7.518.1.H4(S241P) monotherapy dose cohort (Group A).

[0296] CHA.7.518.1.H4(S241P) is well-tolerated and has a manageable safety profile as monotherapy and in combination with nivolumab.

[0297] a. There is no increase in toxicity when used in combination with nivolumab.

[0298] b. No subjects discontinued research treatment due to toxicity from the research drug.

[0299] Monotherapy with MTD CHA.7.518.1.H4(S241P) 20 mg / kg IV for 4 weeks; dose escalation in combination is continued.

[0300] Partial responses were observed in two patients.

[0301] CHA.7.518.1.H4(S241P) monotherapy 20mg / kg IV Q4 week - Primary peritoneal cancer (25 weeks of research treatment ongoing).

[0302] CHA.7.518.1.H4(S241P), (CHA.7.518.1.H4(S241P) 0.3 mg / kg IV Q3 weeks) + Nivolumab (480 mg IV Q3 weeks) - MSS - CRC (44 weeks of ongoing research treatment).

[0303] The disease control rate with CHA.7.518.1.H4(S241P) monotherapy was 11 / 16 [69%] across various tumor types.

[0304] The disease control rate for CHA.7.518.1.H4(S241P) + nivolumab was 9 / 12 [75%] across various tumor types.

[0305] 6 out of 28 patients with ptosis and various tumor types had persistent, stable disease (SD > 6 months).

[0306] Group A (CHA.7.518.1.H4(S241P) monotherapy): Adenoid cystic cyst CA, CRC-MSS.

[0307] Group B (CHA.7.518.1.H4(S241P) + nivolumab): anal SCC, CRC-MSS, endometrium, NSCLC (squamous epithelium).

[0308] The preliminary CHA.7.518.1.H4(S241P)PK profile is for Q4 week. I support the administration of this medication.

[0309] Dose expansion of CHA.7.518.1.H4(S241P) monotherapy is planned for RDFE (NSCLC, OVCA, breast, endometrial, MSS-CRC).

[0310] All headings and section titles are used for clarity and reference purposes only and should not be considered limiting. For example, those skilled in the art will recognize the usefulness of combining various aspects from different headings and sections as needed, in accordance with the spirit and scope of the invention as described herein.

[0311] All references cited herein are incorporated herein by reference in their entirety for the same degree as each individual publication or patent or patent application is specifically and individually indicated by reference as if it were incorporated in its entirety for the sole purpose.

[0312] As will be apparent to those skilled in the art, many modifications and variations of this application may be made without departing from the spirit and scope of this application. The specific embodiments and examples described herein are provided only as examples, and this application should be limited only by the terminology of the appended claims, in addition to the entire scope of the equivalent for which the claims are granted.

Claims

1. A stable liquid pharmaceutical formulation of an anti-PVRIG antibody, (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA. 7.518.

1. H4 (S241P), ii) CHA. 7.518.

1. An anti-PVRIG antibody comprising a light chain variable domain including vlcDr1, vlcDr2, and vlcDr3 from the light chain of H4 (S241P) (SEQ ID NO: 9), (b) Histidine in concentrations of 10 mM to 100 mM, (c) 30 mM to 100 mM NaCl, (d) L-arginine in concentrations of 20 mM to 150 mM, (e) containing 0.005% to 0.1 w / v% polysorbate 80, The composition is a liquid pharmaceutical preparation having a pH of 5.5 to 7.

0.

2. The stable liquid pharmaceutical formulation according to claim 1, wherein the anti-PVRIG antibody comprises an IgG4 CH1-hinge-CH2-CH3 sequence (SEQ ID NO: 17 or SEQ ID NO: 50), and the hinge region optionally contains a mutation.

3. The stable liquid pharmaceutical formulation according to claim 1 or 2, wherein the anti-PVRIG antibody comprises the CH1-hinge-CH2-CH3 region from IgG1, IgG2, IgG3, or IgG4, and the hinge region optionally contains a mutation.

4. A stable liquid pharmaceutical formulation according to any one of claims 1 to 3, wherein the heavy chain variable domain is derived from the heavy chain (SEQ ID NO: 4) of CHA. 7.518.

1. H4 (S241P), and the light chain variable domain is derived from the light chain (SEQ ID NO: 9) of CHA. 7.518.

1. H4 (S241P).

5. The stable liquid pharmaceutical formulation according to any one of claims 1 to 4, wherein the anti-PVRIG antibody comprises the CL region of the human kappa 2 light chain.

6. A stable liquid pharmaceutical formulation according to any one of claims 1 to 5, wherein the pharmaceutical formulation comprises 10 mM to 80 mM histidine, 15 mM to 70 mM histidine, 20 mM to 60 mM histidine, 20 mM to 50 mM histidine, or 20 mM to 30 mM histidine.

7. A stable liquid pharmaceutical preparation according to any one of claims 1 to 6, wherein the pharmaceutical preparation contains approximately 25 mM histidine.

8. A stable liquid pharmaceutical formulation according to any one of claims 1 to 7, wherein the pharmaceutical formulation comprises 30 mM to 100 mM NaCl, 30 mM to 90 mM NaCl, 40 mM to 80 mM NaCl, 30 mM to 70 mM histidine, or 45 mM to 70 mM NaCl.

9. The stable liquid pharmaceutical formulation according to any one of claims 1 to 8, wherein the pharmaceutical formulation contains approximately 60 mM NaCl.

10. The aforementioned pharmaceutical preparation contains 20 mM to 140 mM L-arginine, 30 mM to 140 mM L-arginine, 40 mM to 130 mM L-arginine, 50 mM to 120 mM L-arginine, 60 mM to 110 mM L-arginine, 70 mM to 110 mM L-arginine, 80 mM to 110 mM L-arginine, or 90 mM to 110 mM L-arginine. A stable liquid pharmaceutical preparation according to any one of claims 1 to 9, comprising ginine.

11. The stable liquid pharmaceutical formulation according to any one of claims 1 to 10, wherein the pharmaceutical formulation contains about 100 mM L-arginine.

12. The stable liquid pharmaceutical formulation according to any one of claims 1 to 11, wherein the pharmaceutical formulation comprises 0.006% to 0.1 w / v% polysorbate 80, 0.007% to 0.09 w / v% polysorbate 80, 0.008% to 0.08 w / v% polysorbate 80, 0.009% to 0.09 w / v% polysorbate 80, 0.01% to 0.08 w / v% polysorbate 80, 0.01% to 0.07 w / v% polysorbate 80, or 0.01% to 0.06 w / v% polysorbate 80, or 0.009% to 0.05% w / v polysorbate 80.

13. The stable liquid pharmaceutical formulation according to any one of claims 1 to 12, wherein the pharmaceutical formulation contains about 0.01% polysorbate 80.

14. A stable liquid pharmaceutical preparation according to any one of claims 1 to 13, wherein the pH is 6 to 7.

0.

15. A stable liquid pharmaceutical preparation according to any one of claims 1 to 14, wherein the pH is 6.3 to 6.

8.

16. A stable liquid pharmaceutical preparation according to any one of claims 1 to 15, wherein the pH is 6.5 ± 0.

2.

17. A stable liquid pharmaceutical formulation according to any one of claims 1 to 16, wherein the anti-PVRIG antibody is at a concentration of 10 mg / mL to 40 mg / mL, 15 mg / mL to 40 mg / mL, 15 mg / mL to 30 mg / mL, 10 mg / mL to 25 mg / mL, or 15 mg / mL to 25 mg / mL.

18. A stable liquid pharmaceutical formulation according to any one of claims 1 to 17, wherein the formulation is stable at 2°C to 8°C for at least 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, or 10 weeks.

19. A stable liquid pharmaceutical formulation according to any one of claims 1 to 18, wherein the formulation is stable at approximately 20°C to 25°C for at least one week, two weeks, three weeks, four weeks, five weeks, or six weeks.

20. A stable liquid pharmaceutical formulation according to any one of claims 1 to 19, wherein the formulation is stable at 35°C to 40°C for at least one week, two weeks, three weeks, four weeks, or five weeks.

21. The stable liquid pharmaceutical formulation according to any one of claims 1 to 20, wherein the anti-PVRIG antibody is at a concentration of about 20 mg / mL.

22. The aforementioned anti-PVRIG antibody preparation is a) A heavy chain, i) A heavy chain comprising VH-CH1-hinge-CH2-CH3, wherein VH is derived from CHA. 7.518.

1. H4 (S241P) (SEQ ID NO: 4), and the CH1-hinge-CH2-CH3 region is derived from IgG4, b) A light chain, i) A stable liquid pharmaceutical formulation according to any one of claims 1 to 21, comprising a light chain, the VL-CL comprising the VL-CL, wherein the VL is derived from CHA. 7.518.

1. H4 (S241P) (SEQ ID NO: 9), and the CL region is derived from a human kappa 2 light chain.

23. The stable liquid pharmaceutical formulation according to claim 24, wherein the hinge region optionally includes a mutation.

24. The stable liquid pharmaceutical formulation according to claim 23, wherein the hinge region optionally includes a mutation.

25. The aforementioned anti-PVRIG antibody preparation is i) A heavy chain (Sequence ID 8) containing the heavy chain from CHA. 7.518.

1. H4 (S241P), ii) A stable liquid pharmaceutical preparation according to any one of claims 1 to 24, comprising a light chain (SEQ ID NO: 13) containing the light chain from CHA. 7.518.

1. H4 (S241P).

26. The aforementioned anti-PVRIG antibody preparation is (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) Heavy chain variable domains including vhCDR1, vhCDR2, and vhCDR3 from the heavy chain (SEQ ID NO: 4) of CHA. 7.518.

1. H4 (S241P), ii) CHA. 7.518.

1. An anti-PVRIG antibody comprising a light chain variable domain including vlcDr1, vlcDr2, and vlcDr3 from the light chain of H4 (S241P) (SEQ ID NO: 9), (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl and (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The stable liquid pharmaceutical formulation according to any one of claims 1 to 25, wherein the composition has a pH of 6.5 ± 0.

2.

27. The aforementioned anti-PVRIG antibody preparation is (a) an anti-PVRIG antibody, wherein the anti-PVRIG antibody is i) A heavy chain containing a heavy chain from CHA. 7.518.

1. H4 (S241P) (Sequence No. 8), ii) A light chain (SEQ ID NO: 13) containing the light chain from CHA. 7.518.

1. H4 (S241P), and an anti-PVRIG antibody, (b) Approximately 25 mM histidine, (c) Approximately 60 mM NaCl and (d) Approximately 100 mM L-arginine and (e) containing approximately 0.01 w / v%% polysorbate 80, The stable liquid pharmaceutical formulation according to any one of claims 1 to 26, wherein the composition has a pH of 6.5 ± 0.

2.

28. A stable liquid pharmaceutical preparation according to any one of claims 1 to 27, wherein the anti-PVRIG antibody is administered in a dose of approximately 0.01 mg / kg to approximately 20 mg / kg or approximately 0.01 mg / kg to approximately 10 mg / kg.

29. A stable liquid pharmaceutical preparation according to any one of claims 1 to 27, wherein the anti-PVRIG antibody is administered in a dose of approximately 0.01 mg / kg, 0.03 mg / kg, 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg.

30. The stable liquid pharmaceutical preparation according to any one of claims 1 to 29, wherein the anti-PVRIG antibody is administered at a dose of 20 mg / kg every four weeks.

31. The stable liquid pharmaceutical preparation according to any one of claims 1 to 30, wherein the stable liquid pharmaceutical preparation is administered for the treatment of cancer.

32. A stable liquid pharmaceutical preparation according to any one of claims 1 to 31, for use in a method of treating cancer.

33. The aforementioned cancers include prostate cancer, liver cancer (HCC), colorectal cancer (CRC), colorectal cancer MSS (MSS-CRC; including refractory MSS colorectal cancer), CRC (MSS of unknown origin), ovarian cancer (including ovarian carcinoma), endometrial cancer (including endometrial carcinoma), breast cancer, pancreatic cancer, stomach cancer, cervical cancer, head and neck cancer, thyroid cancer, testicular cancer, urothelial carcinoma, lung cancer, melanoma, non-melanoma skin cancer (squamous and basal cell carcinoma), glioma, renal cell carcinoma (RCC), renal cell carcinoma (RCC), lymphoma (non-Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HD)), acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia (T-ALL), and diffuse large B cell leukemia. A stable liquid pharmaceutical formulation according to claim 31 or 32, selected from the group consisting of lymphoma, testicular germ cell tumor, mesothelioma, esophageal cancer, trinegative breast cancer, Merkel cell carcinoma, MSI hypercarcinoma, KRAS variant tumor, adult T-cell leukemia / lymphoma, pleural mesothelioma, anal SCC, neuroendocrine lung cancer (including neuroendocrine lung carcinoma), NSCLC, NSCLC (large cell), NSCLC large cell, NSCLC squamous cell, cervical SCC, malignant melanoma, pancreatic cancer, pancreatic adenocarcinoma, adenoid cystic carcinoma (including adenoid cystic carcinoma), primary peritoneal cancer, microsatellite-stable primary peritoneal cancer, platinum-resistant microsatellite-stable primary peritoneal cancer, and / or myelodysplastic syndrome (MDS).

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