Anti-CD112R Compositions and Methods

Abstract

To provide anti-CD112R antibody compositions and their use in treating cancer.SOLUTION: In some embodiments, an isolated anti-CD112R antibody is provided. Such isolated anti-CD112R antibody binds to human CD112R, wherein the antibody blocks the binding interaction between human CD112 and human CD112R and does not block the binding interaction between mouse CD112 and mouse CD112R, wherein the antibody is optionally fully human or humanized.SELECTED DRAWING: None

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 62 / 701,065, filed July 20, 2018, and U.S. Provisional Application No. 62 / 844,958, filed May 8, 2019, the contents of which are incorporated herein by reference in their entireties.

[0002] Anti-CD112R antibodies are provided, as are their uses in enhancing, increasing, and maintaining anti-tumor immune responses, treating cancer, and enhancing CD226 interaction with CD112. [Background technology]

[0003] Both the innate and adaptive arms of the immune system utilize highly specialized immune cells to patrol the body, searching for signs of malignancy. Innate immunity provides a first line of defense and rapid response, using mechanisms such as nonspecific, naturally occurring barrier and destructive peptides. Natural killer (NK) cells are a type of lymphocyte that are part of the innate immune system and can recognize and destroy virus-infected cells and tumor cells using granzymes stored in their cytoplasm.

[0004] Adaptive immunity develops over time in response to an antigen and provides long-lasting immunity. + Cytotoxic lymphocytes (CTLs), also known as T cells, are part of the adaptive immune response because they recognize viral and tumor-derived antigens presented by antigen-presenting cells (APCs). CTLs are activated by interaction with APCs, such as dendritic cells or macrophages. The APCs present tumor antigens in the context of MHC molecules to the T cell receptor (TCR) on the T cell surface. During this cognate interaction, the APCs provide costimulatory signals that lead to T cell activation, T cell proliferation, and the reduction or elimination of antigen-expressing cells via cytotoxic mechanisms.

[0005] Administration of anti-CD112R immunotherapy offers an opportunity to increase, enhance, and sustain immune responses. CD112R is an inhibitory receptor expressed primarily by T cells and NK cells and competes with the activating receptor CD226 for CD112 binding. The interaction of CD112 with CD112R is of higher affinity than with CD226, thereby effectively controlling CD226-mediated cell activation. Anti-CD112R antibodies that block the interaction with CD112 limit inhibitory signaling immediately downstream of CD112R while simultaneously promoting greater immune cell activation by increasing CD226 interaction with CD112. In vitro studies have shown that anti-CD112R antibodies increase the proliferation, activation, and cytotoxicity of immune effector cells.

[0006] CD112R mRNA expression is detected in many cancer tissues, based on predictive analysis using the TCGA (The Cancer Genome Atlas) dataset. Its expression is strongest in tumors enriched for T cells and NK cells. In addition to being expressed on myeloid cells, expression of the CD112R ligand, CD112, is routinely elevated on tumor cells of various cellular origins. Given these circumstances, engagement of CD112R on tumor-infiltrating immune cells has the potential to negatively regulate local immune responses within the tumor microenvironment.

[0007] Therapeutic treatment with anti-CD112R antibodies may thereby presumptively target CD112R-expressing immune cells when they engage CD112 on tumor cells and / or myeloid cells within the tumor microenvironment. It offers an opportunity to down-regulate the inhibitory signaling that occurs, with the potential to enhance, increase, and sustain anti-tumor immune responses. Summary of the Invention [Means for solving the problem]

[0008] In some embodiments, an isolated anti-CD112R antibody is provided, wherein the antibody binds to human CD112R, the antibody blocks the binding interaction between human CD112 and human CD112R, and does not block the binding interaction between mouse CD112 and mouse CD112R, and the antibody is optionally fully human or humanized.

[0009] In some embodiments, the present disclosure provides: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 4; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 6; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 101; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 102; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 103; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 104; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 105; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 106; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 201; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 202; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 203; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 204; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 205; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 206; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 301; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 302; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 303; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 304; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 305; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 306; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 401; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 402; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 403; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 404; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 405; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 406; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 501; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 502; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 503; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 504; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 505; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 506; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 601; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 602; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 603; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 604; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 605; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 606; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 701; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 702; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 703; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 704; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 705; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 706; teeth (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 801; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 802; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 803; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 804; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 805; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 806; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 901; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 902; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 903; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 904; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 905; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 906; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 1003; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 1004; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 1005; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 1006; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 2001; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2002; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 2003; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 2004; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 2005; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 2006; or (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 3001; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 3002; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3003; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 3004; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 3005; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 3006; or An isolated antibody is provided, comprising: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 4001; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 4002; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 4003; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 4004; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 4005; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 4006.

[0010] In some embodiments, the antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL): the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 12 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 18; or The VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 112, and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 118, or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 212, and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 218, or wherein the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 312 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 318; or The VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 412 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 418, or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 512 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 518. or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 612 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 618, or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 712. or 100% identical to the amino acid sequence of SEQ ID NO: 812, and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 718, or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 812, and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 818. or 100% identical to the amino acid sequence of SEQ ID NO: 912; and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 918; or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 1012.94%, 95%, 96%, 97%, 98%, 99% or 100% identical, and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 1018; or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 2018; or the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 3012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 3018; or The VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 4012, and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 4018.

[0011] In some embodiments, the antibody comprises six CDRs (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3) described herein and a VH and / or VL sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH and / or VL amino acid sequences described herein. In some embodiments, the VH and / or VL sequences are not 100% identical to the amino acid sequences described herein. In some embodiments, the antibody comprises the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 amino acid sequences described herein and sequence diversity within the VH and / or VL sequences outside of the CDR sequences. In such embodiments, the sequence diversity of the VH and / or VL sequences is within one or more framework regions of the VH and / or VL.

[0012] In some embodiments, the antibodies comprise VH and / or VL sequences that are at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequences set forth herein. In some embodiments, the VH and / or VL sequences are not 100% identical to the amino acid sequences set forth herein. In such embodiments, sequence diversity in the VH and / or VL sequences is within and / or outside of the CDR sequences, unless otherwise specified.

[0013] In some embodiments, the antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL): VH comprises the amino acid sequence of SEQ ID NO: 12 and VL comprises the amino acid sequence of SEQ ID NO: 18; or VH comprises the amino acid sequence of SEQ ID NO: 112 and VL comprises the amino acid sequence of SEQ ID NO: 118; or VH comprises the amino acid sequence of SEQ ID NO: 212 and VL comprises the amino acid sequence of SEQ ID NO: 218; or VH comprises the amino acid sequence of SEQ ID NO: 312 and VL comprises the amino acid sequence of SEQ ID NO: 318; or VH comprises the amino acid sequence of SEQ ID NO: 412 and VL comprises the amino acid sequence of SEQ ID NO: 418; or VH comprises the amino acid sequence of SEQ ID NO: 512 and VL comprises the amino acid sequence of SEQ ID NO: 518; or VH comprises the amino acid sequence of SEQ ID NO: 612 and VL comprises the amino acid sequence of SEQ ID NO: 618; or VH comprises the amino acid sequence of SEQ ID NO: 712 and VL comprises the amino acid sequence of SEQ ID NO: 718; or VH comprises the amino acid sequence of SEQ ID NO: 812 and VL comprises the amino acid sequence of SEQ ID NO: 818; or VH comprises the amino acid sequence of SEQ ID NO: 912 and VL comprises the amino acid sequence of SEQ ID NO: 918; or VH comprises the amino acid sequence of SEQ ID NO: 1012 and VL comprises the amino acid sequence of SEQ ID NO: 1018; or VH comprises the amino acid sequence of SEQ ID NO: 2012 and VL comprises the amino acid sequence of SEQ ID NO: 2018; or VH comprises the amino acid sequence of SEQ ID NO: 3012 and VL comprises the amino acid sequence of SEQ ID NO: 3018; or The VH comprises the amino acid sequence of SEQ ID NO:4012, and the VL comprises the amino acid sequence of SEQ ID NO:4018.

[0014] In some embodiments, anti-CD112R antibodies that activate NK cells are provided. In some embodiments, anti-CD112R antibodies that upregulate CD137 on NK cells are provided. In some embodiments, the anti-CD112R antibody that activates NK cells and / or upregulates CD137 on NK cells is an antibody that is any of the antibodies set forth in the Sequence Listing, such as antibodies 32, 33, 34, 35, and 36. In some embodiments, the anti-CD112R antibody that activates NK cells and / or upregulates CD137 on NK cells is any of the antibodies set forth in the Sequence Listing, such as antibodies 32, 33, 34, 35, and 36, respectively. It contains four CDRs (see sequence listing).

[0015] In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a humanized antibody. In some embodiments, the antibody is a fully human antibody. In some embodiments, the antibody is an antibody fragment selected from a Fab, Fab', Fv, scFv, or (Fab')2. In some embodiments, the antibody is a full-length antibody. In some embodiments, the isolated antibody comprises an IgG1, IgG2, IgG3, or IgG4 Fc region. In some embodiments, the present disclosure provides a composition comprising an antibody disclosed herein and a pharmaceutically acceptable carrier.

[0016] In some embodiments, the antibody increases NK cell degranulation, increases NK cell activation, increases intratumor NK cell activation when presented in combination with an anti-TIGIT antibody, inhibits tumor growth in vivo, and / or prevents tumor engraftment upon rechallenge with a tumor. In some such embodiments, the antibody comprises a human IgG1 heavy chain constant region, and the antibody increases NK cell degranulation, increases NK cell activation, increases intratumor NK cell activation when presented in combination with an anti-TIGIT antibody, inhibits tumor growth in vivo, and / or prevents tumor engraftment upon rechallenge with a tumor relative to an otherwise identical antibody comprising a human IgG heavy chain constant region of a different isotype.

[0017] In some embodiments, the present disclosure provides methods of enhancing, increasing, and / or maintaining an anti-tumor immune response in a subject, comprising administering to the subject having a tumor an antibody or composition described herein.

[0018] In some embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering to a subject having cancer an antibody or composition described herein. In some embodiments, the cancer is carcinoma, lymphoma, blastoma, sarcoma, or leukemia. In some embodiments, the cancer is squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer (including squamous cell non-small cell lung cancer), lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, renal cell carcinoma, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, hepatocellular carcinoma, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma, or various types of head and neck cancer (including head and neck squamous cell carcinoma).

[0019] In some embodiments, the present disclosure provides methods of enhancing CD226 interaction with CD112 in a subject, comprising administering to the subject an antibody or composition described herein.

[0020] In some embodiments, the present disclosure provides methods of enhancing CD8 T cell activation in a subject, comprising administering to a subject in need thereof an antibody or composition described herein.

[0021] In some embodiments, the present disclosure provides methods of enhancing CD8 T cell interferon gamma production in a subject, comprising administering to a subject in need thereof an antibody or composition described herein.

[0022] In some embodiments, the present disclosure provides a method of enhancing NK cell activation in a subject comprising administering to a subject in need thereof an antibody or composition described herein.

[0023] In some embodiments, the present disclosure provides a method of enhancing NK cell-mediated cytotoxicity in a subject comprising administering to a subject in need thereof an antibody or composition described herein.

[0024] In some embodiments, the methods described herein further comprise administering a second therapy. In some embodiments, the second therapy is radiation therapy, surgery, or administration of a second agent. In some embodiments, the second therapy is a second agent. In some such embodiments, the second agent is an antagonist of PD-1, PD-L1, CTLA-4, Lag-3, or TIM-3. In some embodiments, the second agent is an antagonist of TIGIT or CD96. In some embodiments, the second agent is an antagonist of PVRL1, PVRL2, PVRL3, PVRL4, or CD155. In some embodiments, the second agent is an antagonist of CD47, CD39, or IL-27. In some embodiments, the second agent is a STING agonist.

[0025] In some embodiments, the present disclosure provides uses of antibodies or compositions described herein to enhance and / or increase and / or maintain anti-tumor immunity, and / or to treat cancer, and / or to enhance CD226 interaction with CD112.

[0026] In some embodiments, the present disclosure provides for the use of a composition described herein in the preparation of an antibody or a medicament for enhancing and / or increasing and / or maintaining an anti-tumor immune response, and / or for treating cancer, and / or for enhancing CD226 interaction with CD112.

[0027] In some embodiments, the present disclosure provides nucleic acids encoding the antibodies disclosed herein.

[0028] In some embodiments, the present disclosure provides a host cell comprising nucleic acid encoding an antibody disclosed herein.

[0029] In some embodiments, the present disclosure provides a method of producing an antibody disclosed herein, comprising culturing a host cell comprising nucleic acid encoding an antibody disclosed herein, wherein the host cell is cultured under conditions such that the antibody is expressed. In some embodiments, the method further comprises purifying the antibody.

[0030] Exemplary embodiments of the present disclosure include the following. Embodiment 1. An isolated anti-CD112R antibody that binds to human CD112R, wherein said antibody blocks the binding interaction between human CD112 and human CD112R and does not block the binding interaction between mouse CD112 and mouse CD112R, and wherein the antibody is optionally fully human or humanized. Embodiment 2. The isolated antibody comprises: i) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 701, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 702, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 703, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 704, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 705, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 706, or ii) (a) HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001, (b) HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002, (c) HCDR3 comprising the amino acid sequence of SEQ ID NO: 1003, (d) LCDR1 comprising the amino acid sequence of SEQ ID NO: 1004, (e) LCDR1 comprising the amino acid sequence of SEQ ID NO: 10 (f) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 1005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 1006, or iii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 2001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 2003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 2004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 2005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 2006, or iv) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 3001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 3002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 3004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 3005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 3006, or v) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 4001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 4002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 4003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 4004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 4005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 4006, or vi) An isolated antibody according to embodiment 1, comprising: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001 with one, two, or three amino acid changes at positions 4, 5, and / or 6 of SEQ ID NO: 1001; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002 with one, two, three, four, or five amino acid changes at positions 1, 3, 5, 6, and / or 8 of SEQ ID NO: 1002; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 1003; (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 1004; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 1005; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 1006. Embodiment 3. The antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), i)V His at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 712 and VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 718; or ii) V H is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 1012 and VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 1018; or iii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 2018; or iv) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 3012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 3018; or v) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 4012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 4018; 3. The isolated antibody of embodiment 1 or 2, wherein, optionally, if any sequence diversity is present in the CDRs, such sequence diversity is within HCDR1 or HCDR2, with no more than three amino acid changes, such as no more than two amino acid changes to positions 4, 5, and / or 6 of HCDR1, and no more than five amino acid changes, such as no more than two amino acid changes to positions 1, 3, 5, 6, and / or 8 of HCDR2, and optionally, no diversity is present within HCDR3, LCDR1, LCDR2, and LCDR3. Embodiment 4. The antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), i) the VH comprises the amino acid sequence of SEQ ID NO: 712 and the VL comprises the amino acid sequence of SEQ ID NO: 718, or ii) the VH comprises the amino acid sequence of SEQ ID NO: 1012 and the VL comprises the amino acid sequence of SEQ ID NO: 1018, or iii) the VH comprises the amino acid sequence of SEQ ID NO: 2012 and the VL comprises the amino acid sequence of SEQ ID NO: 2018; or iv) the VH comprises the amino acid sequence of SEQ ID NO: 3012 and the VL comprises the amino acid sequence of SEQ ID NO: 3018; or v) An isolated antibody according to any one of embodiments 1 to 3, wherein VH comprises the amino acid sequence of SEQ ID NO: 4012 and VL comprises the amino acid sequence of SEQ ID NO: 4018. Embodiment 5. The isolated antibody of any one of the preceding embodiments, wherein the antibody is a monoclonal antibody. Embodiment 6. The isolated antibody of any one of the preceding embodiments, wherein the antibody is a full-length antibody or an antibody fragment, optionally a Fab, Fab', Fv, scFv, or (Fab')2. Embodiment 7. The isolated antibody of any one of embodiments 1 to 5, wherein the antibody comprises an IgG1, IgG2, IgG3, or IgG4 Fc region, and wherein the antibody optionally comprises a human IgG1 heavy chain constant region, a human IgG4 heavy chain constant region, or a variant human IgG4 heavy chain constant region, wherein the variant human IgG4 heavy chain constant region optionally comprises a mutation selected from a substitution at Ser228, a substitution at Leu235, a substitution at Asn297, or a combination thereof, when numbered according to EU numbering, or a S228P substitution and a L235E substitution, when numbered according to EU numbering. Embodiment 8. The antibody is i) increase NK cell degranulation, and / or ii) increase NK cell activation, and / or iii) increase the activation of intratumoral NK cells when presented in combination with an anti-TIGIT antibody; and / or iv) inhibiting tumor growth in vivo, and / or v) The isolated antibody of any one of embodiments 1 to 7, which prevents tumor engraftment upon rechallenge with a tumor, and optionally, the antibody is IgG1 or IgG4. Embodiment 9. A pharmaceutical composition comprising the antibody of any one of embodiments 1 to 8, wherein the composition optionally comprises an opsonizing agent, a regulatory T cell depleting agent, chemotherapy, and / or an antagonist of PD-1, PD-L1, CTLA-4, Lag-3, or TIM-3, and a pharmaceutically acceptable carrier. Embodiment 10. The isolated antibody of any one of embodiments 1 to 8 or the pharmaceutical composition of embodiment 9 for use in enhancing, increasing, and / or maintaining an anti-tumor immune response in a subject, wherein optionally CD8 T cell activation is enhanced or CD8 T cell interferon gamma production is enhanced in the subject, or optionally NK cell activation is enhanced in the subject or NK cell-mediated cytotoxicity is enhanced in the subject, or optionally CD226 interaction with CD112 is enhanced in the subject. Embodiment 11. The cancer is optionally carcinoma, lymphoma, blastoma, sarcoma, or leukemia, or the cancer is optionally squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, or leukemia. 10. The isolated antibody of any one of embodiments 1 to 8 or the pharmaceutical composition of embodiment 9 for use in treating cancer in a subject that is glioma, soft tissue sarcoma, non-small cell lung cancer (including squamous cell non-small cell lung cancer), lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, renal cell carcinoma, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, hepatocellular carcinoma, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma, or various types of head and neck cancer (including head and neck squamous cell carcinoma). Embodiment 12. The isolated antibody or pharmaceutical composition for use of embodiment 10 or 11, wherein the use further comprises administering a second therapy, which is optionally radiation therapy, surgery, or administration of a second agent, which is optionally an antagonist of PD-1, PD-L1, CTLA-4, Lag-3, or TIM-3, or an antagonist of TIGIT or CD96, or an antagonist of PVRL1, PVRL2, PVRL3, PVRL4, and CD155, or an antagonist of CD47, or an antagonist of CD39, or an antagonist of IL-27, or a STING agonist, and wherein the second agent is optionally an antagonist antibody. Embodiment 13. A nucleic acid encoding the antibody of any one of embodiments 1 to 8. Embodiment 14. A host cell comprising the nucleic acid of embodiment 13. Embodiment 15. A method for producing an antibody according to any one of embodiments 1 to 8, comprising culturing a host cell according to embodiment 14 under conditions in which the antibody is expressed, and optionally purifying the antibody. The present invention provides, for example, the following items. (Item 1) 1. An isolated antibody, comprising: i) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 4, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 5, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 6, or ii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 101, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 102, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 103, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 104, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 105, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 106, or iii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 201, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 202, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 203, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 204, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 205, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 206, or iv) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 301, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 302, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 303, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 304, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 305, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 306, or v) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 401, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 402, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 403, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 404, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 405, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 406, or vi) (a) HCDR1 comprising the amino acid sequence of SEQ ID NO: 501, (b) HCDR2 comprising the amino acid sequence of SEQ ID NO: 502, (c) HCDR comprising the amino acid sequence of SEQ ID NO: 503 3, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 504, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 505, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 506, or vii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 601, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 602, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 603, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 604, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 605, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 606, or viii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 701, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 702, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 703, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 704, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 705, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 706, or ix) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 801, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 802, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 803, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 804, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 805, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 806, or x) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 901, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 902, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 903, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 904, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 905, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 906, or xi) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 1003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 1004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 1005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 1006, or xii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 2001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 2003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 2004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 2005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 2006, or xiii) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 3001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 3002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 3004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 3005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 3006, or xiv) The isolated antibody, comprising (a) an HCDR1 having the amino acid sequence of SEQ ID NO: 4001, (b) an HCDR2 having the amino acid sequence of SEQ ID NO: 4002, (c) an HCDR3 having the amino acid sequence of SEQ ID NO: 4003, (d) an LCDR1 having the amino acid sequence of SEQ ID NO: 4004, (e) an LCDR2 having the amino acid sequence of SEQ ID NO: 4005, and (f) an LCDR3 having the amino acid sequence of SEQ ID NO: 4006. (Item 2) the antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL); i) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 12 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 18; or ii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 112 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 118; or iii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 212 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 218; or iv) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 312 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 318; or v) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 412 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 418; or vi) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 512 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 518; or vii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 612 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 618; or viii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 712 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 718; or ix) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 812 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 818; or x) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 912 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 918; or xi) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 1012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 1018; or xii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2018; or xiii) the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 3012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 3018; or xiv) The isolated antibody of item 1, wherein the VH is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 4012 and the VL is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 4018. (Item 3) the antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL); i) the VH comprises the amino acid sequence of SEQ ID NO: 12 and the VL comprises the amino acid sequence of SEQ ID NO: 18, or ii) the VH comprises the amino acid sequence of SEQ ID NO: 112 and the VL comprises the amino acid sequence of SEQ ID NO: 118; or iii) the VH comprises the amino acid sequence of SEQ ID NO: 212 and the VL comprises the amino acid sequence of SEQ ID NO: 218; or iv) the VH comprises the amino acid sequence of SEQ ID NO: 312 and the VL comprises the amino acid sequence of SEQ ID NO: 318; or v) the VH comprises the amino acid sequence of SEQ ID NO: 412 and the VL comprises the amino acid sequence of SEQ ID NO: 418; or vi) the VH comprises the amino acid sequence of SEQ ID NO: 512 and the VL comprises the amino acid sequence of SEQ ID NO: 518; or vii) the VH comprises the amino acid sequence of SEQ ID NO: 612 and the VL comprises the amino acid sequence of SEQ ID NO: 618; or viii) the VH comprises the amino acid sequence of SEQ ID NO: 712 and the VL comprises the amino acid sequence of SEQ ID NO: 718; or ix) the VH comprises the amino acid sequence of SEQ ID NO: 812 and the VL comprises the amino acid sequence of SEQ ID NO: 818; or x) the VH comprises the amino acid sequence of SEQ ID NO: 912 and the VL comprises the amino acid sequence of SEQ ID NO: 918, or xi) the VH comprises the amino acid sequence of SEQ ID NO: 1012 and the VL comprises the amino acid sequence of SEQ ID NO: 1018; or xii) the VH comprises the amino acid sequence of SEQ ID NO: 2012 and the VL comprises the amino acid sequence of SEQ ID NO: 2018; or xiii) the VH comprises the amino acid sequence of SEQ ID NO: 3012 and the VL comprises the amino acid sequence of SEQ ID NO: 3018; or xiv) The isolated antibody of item 1 or 2, wherein the VH comprises the amino acid sequence of SEQ ID NO: 4012 and the VL comprises the amino acid sequence of SEQ ID NO: 4018. (Item 4) Item 10. The isolated antibody of any one of the preceding items, wherein the antibody is a monoclonal antibody. (Item 5) Item 10. The isolated antibody of any one of the preceding items, wherein the antibody is an antibody fragment. (Item 6) 6. The isolated antibody of item 5, wherein the fragment is a Fab, a Fab', an Fv, an scFv, or an (Fab')2. (Item 7) 5. The isolated antibody of any one of items 1 to 4, wherein the antibody is a full-length antibody. (Item 8) 8. The isolated antibody of item 7, wherein the Fc region of the antibody comprises IgG1, IgG2, IgG3, or IgG4. (Item 9) 9. The isolated antibody of any one of items 1 to 8, wherein the antibody comprises a human IgG1 heavy chain constant region. (Item 10) 9. The isolated antibody of any one of items 1 to 8, wherein the antibody comprises a human IgG4 heavy chain constant region. (Item 11) 11. The isolated antibody of claim 10, wherein the antibody comprises a variant human IgG4 heavy chain constant region. (Item 12) 12. The isolated antibody of item 11, wherein the mutant IgG4 heavy chain constant region comprises a mutation selected from a substitution at Ser228, a substitution at Leu235, a substitution at Asn297, or a combination thereof, numbered according to EU numbering. (Item 13) 12. The isolated antibody of item 11, wherein the mutant IgG4 heavy chain constant region comprises an S228P substitution and an L235E substitution, numbered according to EU numbering. (Item 14) The antibody comprises a human IgG1 heavy chain constant region, and the antibody comprises: i) increase NK cell degranulation, and / or ii) increase NK cell activation, and / or iii) increase the activation of intratumoral NK cells when presented in combination with an anti-TIGIT antibody; and / or iv) inhibiting tumor growth in vivo, and / or v) The isolated antibody of any one of items 1 to 9, which prevents tumor engraftment upon rechallenge with a tumor. (Item 15) A humanized or fully human version of the antibody of any one of the preceding items. (Item 16) A composition comprising the antibody according to any one of items 1 to 15 and a pharmaceutically acceptable carrier. (Item 17) 17. The composition of item 16, further comprising an antagonist of PD-1, PD-L1, CTLA-4, Lag-3, or TIM-3. (Item 18) 18. A method for enhancing, increasing, and / or maintaining an anti-tumor immune response in a subject, the method comprising administering to a subject having a tumor the antibody of any one of items 1 to 15 or the composition of item 16 or 17. (Item 19) A method for enhancing CD8 T cell activation in a subject, the method comprising administering the antibody of any one of items 1 to 15 or the composition of item 16 or 17 to a subject in need of CD8 T cell activation. (Item 20) A method for enhancing CD8 T cell interferon gamma production in a subject, the method comprising administering the antibody of any one of items 1 to 15 or the composition of item 16 or 17 to a subject in need of CD8 T cell interferon gamma production. (Item 21) A method for enhancing NK cell activation in a subject, the method comprising administering the antibody of any one of items 1 to 15 or the composition of item 16 or 17 to a subject in need of NK cell activation. (Item 22) A method for enhancing NK cell-mediated cytotoxicity in a subject, the method comprising administering the antibody of any one of items 1 to 15 or the composition of item 16 or 17 to a subject in need of increased NK cell-mediated cytotoxicity. (Item 23) 18. A method for treating cancer in a subject, comprising administering the antibody of any one of items 1 to 15 or the composition of item 16 or 17 to a subject having cancer. (Item 24) 24. The method of claim 23, wherein the cancer is carcinoma, lymphoma, blastoma, sarcoma, or leukemia. (Item 25) 24. The method of claim 23, wherein the cancer is squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer (including squamous cell non-small cell lung cancer), lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, renal cell carcinoma, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, hepatocellular carcinoma, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma, or various types of head and neck cancer (including head and neck squamous cell carcinoma). (Item 26) A method for enhancing CD226 interaction with CD112 in a subject, the method comprising administering to the subject the antibody of any one of items 1 to 15 or the composition of item 16 or 17. (Item 27) 27. The method of any one of items 18 to 26, wherein the method further comprises administering a second therapy. (Item 28) 28. The method of item 27, wherein the second therapy is radiation therapy or surgery. (Item 29) 28. The method of item 27, wherein the second therapy is administration of chemotherapy, an opsonizing agent, or a regulatory T cell depleting agent. (Item 30) 28. The method of item 27, wherein the second therapy is administration of an antagonist of PD-1, PD-L1, CTLA-4, Lag-3, or TIM-3. (Item 31) 28. The method of item 27, wherein the second therapy is administration of an antagonist of TIGIT or CD96. (Item 32) 28. The method of item 27, wherein the second therapy is administration of an antagonist of PVRL1, PVRL2, PVRL3, PVRL4, and CD155. (Item 33) 28. The method of item 27, wherein the second therapy is administration of a CD47 antagonist. (Item 34) Item 35. The method of Item 27, wherein the second therapy is administration of a CD39 antagonist. 28. The method of item 27, wherein the second therapy is administration of an antagonist of IL-27. (Item 36) 28. The method of item 27, wherein the second therapy is administration of a STING agonist. (Item 37) 36. The method of any one of items 30 to 35, wherein the antagonist is an antibody. (Item 38) A nucleic acid encoding the antibody according to any one of items 1 to 15. (Item 39) A host cell comprising the nucleic acid of item 38. (Item 40) 16. A method for producing the antibody according to any one of items 1 to 15, comprising culturing the host cell according to item 39 under conditions in which the antibody is expressed. (Item 41) 41. The method of claim 40, further comprising purifying the antibody. [Brief explanation of the drawings]

[0031] [Figure 1] 1 shows the ability of anti-CD112R antibodies to bind to Jurkat cells engineered to overexpress human CD112R, compared to an IgG1 isotype control antibody. Binding strength was assessed by the geometric mean fluorescence intensity (gMFI) of the Alexa Fluor® 647 antibody label. [Figure 2A]Figure 1 shows the ability of anti-CD112R antibodies to block the interaction of CD112R with CD112 on Jurkat cells engineered to overexpress human CD112R, compared with an IgG1 isotype control antibody. Cells were preincubated with either the IgG1 isotype control or the anti-CD112R antibody. After washing, cells were simultaneously stained with biotinylated his-tagged human CD112 and streptavidin-PE. The ability of anti-CD112R antibodies to block CD112 binding to CD112R was assessed by the geometric mean fluorescence intensity (gMFI) of the PE label and expressed as percent inhibition. Percent inhibition was calculated as [100 - ((test sample MFI / maximum MFI) * 100%)]. [Figure 2B] 1 shows the ability of anti-CD112R antibodies to block the interaction of CD112R with CD112 on Jurkat cells engineered to overexpress human CD112R, compared to an IgG1 isotype control antibody. Percent inhibition of the interaction between human CD112R and CD112 by anti-CD112R antibodies described herein, as measured by ELISA, is shown. [Figure 3] Figure 1 shows enhanced human NK cell-mediated cytotoxicity against REH cells (human leukemia cell line) in the presence of anti-CD112R antibody compared with IgG1 isotype antibody. Activated NK cells and cellTrace Violet-labeled REH cells were co-cultured for 4 hours. After co-culture, REH cell viability was assessed by staining with 7-AAD. Cytotoxicity (percent over isotype) was calculated as ((test mortality - isotype mortality) / isotype mortality) x 100. [Figure 4]Figure 1 shows enhanced antigen-driven activation of CD8+ T cells, as measured by IFNγ secretion, in the presence of anti-CD112R antibodies compared to an IgG1 isotype control antibody. Colo205 cells were pulsed with pp65 peptide and cocultured with human CMV-specific T cells in the presence of anti-CD112R antibodies or an IgG1 isotype control. IFNγ levels in the supernatants of cultured cells were measured by Luminex. CD8+ T cells treated with anti-CD112R antibodies 2 and 5 resulted in greater IFNγ secretion than observed with the isotype control. [Figure 5] This graph shows that the combination of mouse anti-CD112R and mouse anti-TIGIT antibodies has therapeutic efficacy in a mouse syngeneic CT-26 tumor model. Tumor-bearing mice were randomized into four groups and treated twice weekly by IP injection for two weeks with 1) an isotype control antibody, 2) an anti-TIGIT antibody, 3) an anti-CD112R antibody, or 4) an anti-TIGIT antibody combined with an anti-CD112R antibody. The mean tumor volume for each treatment group is shown as a function of time. The results demonstrate that the combination of anti-CD112R with anti-TIGIT was effective in reducing tumor growth compared to isotype-treated animals, while anti-CD112R or anti-TIGIT monotherapy either showed no activity or only a minimal effect on tumor growth reduction. [Figure 6] Graph showing increased expression of CD112R in PBMCs after anti-CD3 activation. Human PBMCs were stimulated in vitro with anti-CD3 antibody, and CD112R expression was assessed by flow cytometry. Quantitation of CD112R antibody binding was assessed by the geometric mean fluorescence intensity (gMFI) of Alexa Fluor® 647 antibody labeling for the indicated cell types. CD112R is shown as the fold change over the negative control (FON, (CD112R gMFI divided by isotype gMFI)). [Figure 7]Enhanced NK cell-mediated degranulation in response to tumor cells was observed in the presence of a CD112R antibody with an IgG1 isotype. Human NK cells and Raji.CD112 cells were co-cultured with a CD107a PE antibody in the presence of a CD112R antibody for 4 hours. After co-culture, NK cell degranulation was determined by the frequency of CD107a-positive NK cells. [Figure 8A] Figure 8 shows enhanced NK cell activation in the presence of a CD112R antibody with an IgG1 isotype. Human PBMCs from two different donors and K562 cells were co-cultured for 16 hours in the presence of a CD112R antibody. After co-culture, NK cell activation was determined by the frequency of CD137-positive NK cells. The results for donor 1 and donor 2 in two independent assays are shown in Figures 8A-8D, respectively. [Figure 8B] Figure 8 shows enhanced NK cell activation in the presence of a CD112R antibody with an IgG1 isotype. Human PBMCs from two different donors and K562 cells were co-cultured for 16 hours in the presence of a CD112R antibody. After co-culture, NK cell activation was determined by the frequency of CD137-positive NK cells. The results for donor 1 and donor 2 in two independent assays are shown in Figures 8A-8D, respectively. [Figure 8C] Figure 8 shows enhanced NK cell activation in the presence of a CD112R antibody with an IgG1 isotype. Human PBMCs from two different donors and K562 cells were co-cultured for 16 hours in the presence of a CD112R antibody. After co-culture, NK cell activation was determined by the frequency of CD137-positive NK cells. The results for donor 1 and donor 2 in two independent assays are shown in Figures 8A-8D, respectively. [Figure 8D] Figure 8 shows enhanced NK cell activation in the presence of a CD112R antibody with an IgG1 isotype. Human PBMCs from two different donors and K562 cells were co-cultured for 16 hours in the presence of a CD112R antibody. After co-culture, NK cell activation was determined by the frequency of CD137-positive NK cells. The results for donor 1 and donor 2 in two independent assays are shown in Figures 8A-8D, respectively. [Figure 9] Figure 1 shows tumor growth inhibition in mice treated with anti-CD112R antibody. The figure shows a summary of three experiments, with N = 44-45 per group. Statistical analysis was performed by Mann-Whitney test on day 24 after implantation. [Figure 10A]

[0033] Figure 1 shows that treatment with anti-CD112R antibody increases overall survival of mice inoculated with CT-26 tumors and protects them from tumor rechallenge. Survival rates of mice after primary tumor challenge with anti-CD112R treatment are shown. Surviving mice did not exhibit palpable tumors 50 days after inoculation and were considered complete responders. [Figure 10B] Figure 10 shows that treatment with anti-CD112R antibody increases the overall survival of mice inoculated with CT-26 tumors and protects them from tumor rechallenge. Tumor growth inhibition in surviving mice upon tumor rechallenge is shown compared to naive control mice. Statistical analysis was performed by Mantel-Cox test at 50 days post-implantation (Figure 10A) and Mann-Whitney test at 15 days post-implantation (Figure 10B). [Figure 11] Figure 1 shows that the in vivo efficacy of CD112R blockade in the CT26 mouse tumor model is dependent on NK cells and CD8 T cells. The figure shows tumor growth inhibition in anti-CD112R-treated mice that were simultaneously depleted of either NK cells or CD8 T cells. [Figure 12] 12A and 12B show the expression of CD69 (FIG. 12A) and granzyme B (FIG. 12B) on intratumoral NK cells in a CT-26 tumor model after treatment with anti-CD112R antibody. [Figure 13A] Shown are mean (FIG. 13A) and individual (FIGS. 13B-E) tumor volume measurements as a function of time in the CT-26 tumor model following administration of anti-CD112R antibody alone and in combination with anti-PD1 antibody. [Figure 13B] Shown are mean (FIG. 13A) and individual (FIGS. 13B-E) tumor volume measurements as a function of time in the CT-26 tumor model following administration of anti-CD112R antibody alone and in combination with anti-PD1 antibody. [Figure 13C]Shown are mean (FIG. 13A) and individual (FIGS. 13B-E) tumor volume measurements as a function of time in the CT-26 tumor model following administration of anti-CD112R antibody alone and in combination with anti-PD1 antibody. [Figure 13D] Shown are mean (FIG. 13A) and individual (FIGS. 13B-E) tumor volume measurements as a function of time in the CT-26 tumor model following administration of anti-CD112R antibody alone and in combination with anti-PD1 antibody. [Figure 13E] Shown are mean (FIG. 13A) and individual (FIGS. 13B-E) tumor volume measurements as a function of time in the CT-26 tumor model following administration of anti-CD112R antibody alone and in combination with anti-PD1 antibody. [Figure 13F] Figure 13A shows mean (Figure 13A) and individual (Figures 13B-E) tumor volume measurements as a function of time in the CT-26 tumor model after administration of anti-CD112R antibody alone and in combination with anti-PD1 antibody. Overall tumor-free survival at 50 days post-implantation is shown as the percentage of tumor-free survivors per group. [Figure 14A] Figures 14A-14C show alignments of the CDR sequences of the antibodies described herein. Figures 14A-14C show the CDR sequences of the heavy chain variable regions. In each of Figures 14A-14F, the first column shows HCDR1 (Figure 14A), HCDR2 (Figure 14B), and HCDR3 (Figure 14C), LCDR1 (Figure 14D), LCDR2 (Figure 14E), and LCDR3 (Figure 14F). The antibody clone numbers are provided before H1 (for HCDR1), H2 (for HCDR2), H3 (for HCDR3), L1 (for LCDR1), L2 (for LCDR2), and L3 (for LCDR3). The second column shows the sequence, the third column shows the number of amino acids in the sequence, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 14B]Figures 14A-14C show alignments of the CDR sequences of the antibodies described herein. Figures 14A-14C show the CDR sequences of the heavy chain variable regions. In each of Figures 14A-14F, the first column shows HCDR1 (Figure 14A), HCDR2 (Figure 14B), and HCDR3 (Figure 14C), LCDR1 (Figure 14D), LCDR2 (Figure 14E), and LCDR3 (Figure 14F). The antibody clone numbers are provided before H1 (for HCDR1), H2 (for HCDR2), H3 (for HCDR3), L1 (for LCDR1), L2 (for LCDR2), and L3 (for LCDR3). The second column shows the sequence, the third column shows the number of amino acids in the sequence, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 14C] Figures 14A-14C show alignments of the CDR sequences of the antibodies described herein. Figures 14A-14C show the CDR sequences of the heavy chain variable regions. In each of Figures 14A-14F, the first column shows HCDR1 (Figure 14A), HCDR2 (Figure 14B), and HCDR3 (Figure 14C), LCDR1 (Figure 14D), LCDR2 (Figure 14E), and LCDR3 (Figure 14F). The antibody clone numbers are provided before H1 (for HCDR1), H2 (for HCDR2), H3 (for HCDR3), L1 (for LCDR1), L2 (for LCDR2), and L3 (for LCDR3). The second column shows the sequence, the third column shows the number of amino acids in the sequence, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 14D]Alignment of CDR sequences of antibodies described herein is shown. Figures 14D-14F show the CDR sequences of the light chain variable region. In each of Figures 14A-14F, the first column shows HCDR1 (Figure 14A), HCDR2 (Figure 14B), and HCDR3 (Figure 14C), LCDR1 (Figure 14D), LCDR2 (Figure 14E), and LCDR3 (Figure 14F), with antibody clone numbers provided before H1 (for HCDR1), H2 (for HCDR2), H3 (for HCDR3), L1 (for LCDR1), L2 (for LCDR2), and L3 (for LCDR3). The second column shows the sequence, the third column shows the number of amino acids in the sequence, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 14E] Alignment of CDR sequences of antibodies described herein is shown. Figures 14D-14F show the CDR sequences of the light chain variable region. In each of Figures 14A-14F, the first column shows HCDR1 (Figure 14A), HCDR2 (Figure 14B), and HCDR3 (Figure 14C), LCDR1 (Figure 14D), LCDR2 (Figure 14E), and LCDR3 (Figure 14F), with antibody clone numbers provided before H1 (for HCDR1), H2 (for HCDR2), H3 (for HCDR3), L1 (for LCDR1), L2 (for LCDR2), and L3 (for LCDR3). The second column shows the sequence, the third column shows the number of amino acids in the sequence, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 14F]Alignment of CDR sequences of antibodies described herein is shown. Figures 14D-14F show the CDR sequences of the light chain variable region. In each of Figures 14A-14F, the first column shows HCDR1 (Figure 14A), HCDR2 (Figure 14B), and HCDR3 (Figure 14C), LCDR1 (Figure 14D), LCDR2 (Figure 14E), and LCDR3 (Figure 14F), with antibody clone numbers provided before H1 (for HCDR1), H2 (for HCDR2), H3 (for HCDR3), L1 (for LCDR1), L2 (for LCDR2), and L3 (for LCDR3). The second column shows the sequence, the third column shows the number of amino acids in the sequence, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15A] Alignment of framework region sequences of antibodies described herein is shown. Figures 15A-15D show the framework region sequences of the heavy chain variable regions. In each of Figures 15A-15F, the first column shows the heavy chain FR1 (Figure 15A), FR2 (Figure 15B), FR3 (Figure 15C), and FR4 (Figure 15D) and the light chain FR1 (Figure 15E), FR2 (Figure 15F), and FR3 (Figure 15G), and FR4 (Figure 15H). The antibody clone numbers are provided before the VH (for HFR1, HFR3, HFR3, and HFR4) and VL (for LFR1, LFR2, LFR3, and LFR4). The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15B]Alignment of framework region sequences of antibodies described herein is shown. Figures 15A-15D show the framework region sequences of the heavy chain variable regions. In each of Figures 15A-15F, the first column shows the heavy chain FR1 (Figure 15A), FR2 (Figure 15B), FR3 (Figure 15C), and FR4 (Figure 15D) and the light chain FR1 (Figure 15E), FR2 (Figure 15F), and FR3 (Figure 15G), and FR4 (Figure 15H). The antibody clone numbers are provided before the VH (for HFR1, HFR3, HFR3, and HFR4) and VL (for LFR1, LFR2, LFR3, and LFR4). The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15C] Alignment of framework region sequences of antibodies described herein is shown. Figures 15A-15D show the framework region sequences of the heavy chain variable regions. In each of Figures 15A-15F, the first column shows the heavy chain FR1 (Figure 15A), FR2 (Figure 15B), FR3 (Figure 15C), and FR4 (Figure 15D) and the light chain FR1 (Figure 15E), FR2 (Figure 15F), and FR3 (Figure 15G), and FR4 (Figure 15H). The antibody clone numbers are provided before the VH (for HFR1, HFR3, HFR3, and HFR4) and VL (for LFR1, LFR2, LFR3, and LFR4). The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15D]Alignment of framework region sequences of antibodies described herein is shown. Figures 15A-15D show the framework region sequences of the heavy chain variable regions. In each of Figures 15A-15F, the first column shows the heavy chain FR1 (Figure 15A), FR2 (Figure 15B), FR3 (Figure 15C), and FR4 (Figure 15D) and the light chain FR1 (Figure 15E), FR2 (Figure 15F), and FR3 (Figure 15G), and FR4 (Figure 15H). The antibody clone numbers are provided before the VH (for HFR1, HFR3, HFR3, and HFR4) and VL (for LFR1, LFR2, LFR3, and LFR4). The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15E] Alignment of framework region sequences of antibodies described herein is shown. Figures 15E-15H show the framework region sequences of the light chain variable region. In each of Figures 15A-15F, the first column shows the heavy chain FR1 (Figure 15A), FR2 (Figure 15B), FR3 (Figure 15C), and FR4 (Figure 15D) and the light chain FR1 (Figure 15E), FR2 (Figure 15F), and FR3 (Figure 15G), and FR4 (Figure 15H). The antibody clone numbers are provided before the VH (for HFR1, HFR3, HFR3, and HFR4) and VL (for LFR1, LFR2, LFR3, and LFR4). The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15F]Alignment of framework region sequences of antibodies described herein is shown. Figures 15E-15H show the framework region sequences of the light chain variable region. In each of Figures 15A-15F, the first column shows the heavy chain FR1 (Figure 15A), FR2 (Figure 15B), FR3 (Figure 15C), and FR4 (Figure 15D) and the light chain FR1 (Figure 15E), FR2 (Figure 15F), and FR3 (Figure 15G), and FR4 (Figure 15H). The antibody clone numbers are provided before the VH (for HFR1, HFR3, HFR3, and HFR4) and VL (for LFR1, LFR2, LFR3, and LFR4). The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15G] Figures 15E-15H show an alignment of the framework region sequences of the antibodies described herein. Figures 15E-15H show the framework region sequences of the light chain variable region. The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 15H] Figures 15E-15H show an alignment of the framework region sequences of the antibodies described herein. Figures 15E-15H show the framework region sequences of the light chain variable region. The second column shows the sequences, and the last column shows the percent identity of each sequence relative to the sequence from parent antibody clone 32. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 16A]

[0023] Figure 1 shows an alignment of the variable region sequences of the antibodies described herein. The heavy chain variable region sequences are shown. Each sequence is labeled with its corresponding clone number. The percent identity of each sequence to the sequence from antibody clone 32 is shown. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 16B]

[0023] Figure 1 shows an alignment of the variable region sequences of the antibodies described herein. The light chain variable region sequences are shown. Each sequence is labeled with its corresponding clone number. The percent identity of each sequence to the sequence from antibody clone 32 is shown. Family member clones 32, 33, 34, 35, and 36 are shown in bold. [Figure 17] 1 shows the extent of binding of anti-CD112R antibodies described herein and additional anti-CD112R antibodies (antibodies A, B, and C that bind to human CD112R) to cells expressing mouse CD112R. [Figure 18] 1 shows the extent of binding to soluble mouse CD112R of anti-CD112R antibodies described herein and additional anti-CD112R antibodies (antibodies A, B, and C that bind to human CD112R). [Figure 19] 1 shows the percent inhibition of the interaction between mouse CD11R and mouse CD112 by anti-CD112R antibodies described herein and additional anti-CD112R antibodies (antibodies A, B, and C that bind to human CD112R). DETAILED DESCRIPTION OF THE INVENTION

[0032] I. Definition In this application, the use of "or" means "and / or" unless stated otherwise. In the context of multiple dependent claims, the use of "or" refers back to two or more preceding independent or dependent claims, in the alternative only. The terms "comprising," "including," and "having" may be used interchangeably herein.

[0033] The terms "CD112R," "PVR-related immunoglobulin domain-containing," "CD112 receptor," "poliovirus receptor-related immunoglobulin domain-containing protein," "poliovirus receptor-related immunoglobulin domain-containing," "nectin-2 receptor," "C7orf15," and "transmembrane protein PVRIG" are all used interchangeably and refer to native human CD112R unless specifically indicated otherwise (e.g., mouse CD112R, cynomolgus monkey CD112R, etc.). The term includes full-length, unprocessed CD112R, as well as any form of CD112R that results from processing in cells. The term encompasses naturally occurring variants of human CD112R (e.g., splice variants or allelic variants). External IDs for the CD112R gene include Entrez Gene: 79037, Ensembl: ENSG00000213413, OMIM: 617012, and UniProtKB: Q6DKI7.

[0034] "Affinity" refers to the strength of the sum total of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless otherwise indicated, as used herein, "binding affinity" refers to the intrinsic binding affinity, which reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a molecule X for its partner Y is generally determined by the dissociation constant (K D ) Affinity can be measured by common methods known in the art, including those described herein. Specific illustrations and exemplary embodiments for measuring binding affinity are described below.

[0035] An "affinity matured" antibody refers to an antibody that has one or more alterations in one or more hypervariable regions (HVRs) compared to a parent antibody that does not possess such alterations, which optionally result in an improvement in the affinity of the antibody for antigen.

[0036] The term "antibody" as used herein is used in the broadest sense and encompasses a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, so long as they exhibit the desired antigen-binding activity.

[0037] "Antibody fragment" refers to a molecule other than an intact antibody that contains a portion of an intact antibody that binds to the antigen to which the intact antibody binds. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2; diabodies; linear antibodies; single-chain antibody molecules (e.g., scFv); and multispecific antibodies formed from antibody fragments.

[0038] The term "blocking" in the context of an interaction between two or more molecules is used herein to refer to the inhibition or prevention of the interaction between two or more molecules, where the inhibition or prevention of the interaction between two or more molecules is complete or nearly complete under at least one condition. "Near-complete" inhibition is about 70-99.9% inhibition, and "complete" inhibition is 100%. For example, a molecule is said to "inhibit" an interaction between two or more other molecules if it completely or nearly completely inhibits such interaction at a certain concentration in a dose-dependent manner.

[0039] The term "cancer" is used herein to refer to a group of cells that exhibit abnormally high levels of proliferation and growth. Cancers can be benign (also referred to as benign tumors), pre-malignant, or malignant. Cancer cells can be solid cancer cells or leukemic cancer cells. The term "tumor" is used herein to refer to a cell or cells that comprise cancer. The term "tumor growth" is used herein to refer to proliferation or growth by a cell or cells that comprise cancer, resulting in a corresponding increase in the size or extent of the cancer.

[0040] The term "chimeric" antibody refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while the remainder of the heavy and / or light chain is derived from a different source or species.

[0041] The "class" of an antibody refers to the type of constant domain or constant region carried by its heavy chain. There are five major antibody classes: IgA, IgD, IgE, IgG, and IgM, and some of these can be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy-chain constant domains that correspond to the different classes of immunoglobulins are called α, δ, ε, γ, and μ, respectively.

[0042] Administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive (sequential) administration in any order.

[0043] The term "cytotoxic agent" as used herein refers to a substance that inhibits or prevents cell function and / or causes cell death or destruction. Cytotoxic agents include radioisotopes (e.g., At 211 , I 131 , I 125 , Y 90 ,Re 186 ,Re 188 , Sm 153 , Bi 212 , P 32 , Pb 212 , and radioactive isotopes of Lu); chemotherapeutic agents or drugs (e.g., methotrexate, adriamycin, vinca alkaloids (vincristine, vinblastine, etoposide), doxorubicin, melphalan, mitomycin C, chlorambucil, daunorubicin, or other intercalating agents); growth inhibitory agents; enzymes and fragments thereof, such as nucleases; antibiotics; toxins, such as small molecule toxins or enzymatically active toxins (including fragments and / or variants thereof) of bacterial, fungal, plant, or animal origin; and various anti-tumor or anti-cancer agents disclosed below, including, but not limited to, Not limited to:

[0044] "Effector functions" refer to those biological activities attributable to the Fc region of an antibody, which vary depending on the antibody isotype. Examples of antibody effector functions include C1q binding and complement-dependent cytotoxicity (CDC), Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, down-regulation of cell surface receptors (e.g., B cell receptors), and B cell activation.

[0045] An "effective amount" of an agent, eg, a pharmaceutical formulation, refers to an amount effective to achieve a desired therapeutic or prophylactic result at the dosages required and for a period of time.

[0046] The term "Fc region" is used herein to define the C-terminal region of an immunoglobulin heavy chain containing at least a portion of the constant region. The term includes native sequence Fc regions and variant Fc regions. In some embodiments, a human IgG heavy chain Fc region extends from Cys226 or Pro230 to the carboxyl terminus of the heavy chain. However, the C-terminal lysine (Lys447) of the Fc region is not included in the Fc region as described by Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed. Follow the EU numbering system, also known as the EU index, as described in Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991).

[0047] "Framework" or "framework region" or "FR" refers to variable domain residues other than hypervariable region (HVR) residues. The FR of a variable domain generally consists of four FR domains, FR1, FR2, FR3, and FR4. Thus, the HVR and FR sequences generally appear in VH (or VL) in the following order: FR1-H1(L1)-FR2-H2(L2)-FR3-H3(L3)-FR4.

[0048] The terms "full length antibody," "intact antibody," and "whole antibody" are used interchangeably herein to refer to an antibody having a structure substantially similar to a native antibody structure or having a heavy chain containing an Fc region as defined herein.

[0049] The terms "host cell," "host cell line," and "host cell culture" are used interchangeably and refer to cells into which exogenous nucleic acid has been introduced, including the progeny of such cells. Host cells include "transformants" and "transformed cells," including the primary transformed cell and its progeny without regard to the number of transfers. Progeny may not be completely identical in nucleic acid content to the parent cell, but may contain mutations. Mutant progeny that have the same function or biological activity as screened or selected for in the originally transformed cell are included herein.

[0050] A "human antibody" is one that possesses an amino acid sequence that corresponds to the amino acid sequence of an antibody produced by a human or human cell, or derived from a non-human source that utilizes the human antibody repertoire, or other human antibody coding sequence. This definition of human antibody specifically excludes humanized antibodies that comprise non-human antigen-binding residues.

[0051] The term "variable region" or "variable domain" refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to an antigen. The variable domains of the heavy and light chains of a natural antibody (VH and VL, respectively) generally have similar structures, with each domain containing four conserved framework regions (FR) and three hypervariable regions (HVR). (See, e.g., Kindt et al., Kuby Immunology, 6 th (See, e.g., W.H. Freeman and Co., page 91 (2007).) A single VH or VL domain may be sufficient to confer antigen-binding specificity. Antibodies that bind to an antigen can be isolated using a VH or VL domain from an antibody that binds to the antigen to screen a library of complementary VL or VH domains, respectively. See, e.g., Portolano et al., J. Immunol. 150:880-887 (1993); Clarkson et al., Nature 352:624-628 (1991).

[0052] A "human consensus framework" is a framework that represents the most commonly occurring amino acid residues in a selection of human immunoglobulin VL or VH framework sequences. Generally, the selection of human immunoglobulin VL or VH sequences is from a subgroup of variable domain sequences. Generally, the subgroup of sequences is a subgroup in Kabat et al., Sequences of Proteins of Immunological Interest, Fifth Edition, NIH Publication 91-3242, Bethesda, MD (1991), vols. 1-3. In some embodiments, for VL, the subgroup is subgroup kappa I as in Kabat et al., supra. In some embodiments, for VH, the subgroup is subgroup III as in Kabat et al., supra.

[0053] As used herein, the term "hypervariable region" or "HVR" refers to each of the regions of an antibody variable domain that are hypervariable in sequence ("complementarity determining regions" or "CDRs") and / or form structurally defined loops ("hypervariable loops") and / or contain antigen-contacting residues ("antigen contactors"). Generally, antibodies contain six HVRs: three in the VH (H1, H2, H3) and three in the VL (L1, L2, L3).

[0054] An "immunoconjugate" is an antibody conjugated to one or more heterologous molecule(s), including, but not limited to, a cytotoxic agent.

[0055] An "individual" or "subject" is a mammal. Mammals include, but are not limited to, domestic animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain embodiments, the individual or subject is human.

[0056] An "isolated" antibody is one that has been separated from components of its natural environment. In some embodiments, the antibody is purified to greater than 95% or greater than 99% purity, for example, as determined by electrophoretic methods (e.g., SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatographic methods (e.g., ion exchange or reverse-phase HPLC). For a review of methods for assessing antibody purity, see, for example, Flatman et al., J. Chromatogr. B848:79-87 (2007).

[0057] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and / or bind to the same epitope, with the exception of possible variant antibodies that contain, for example, naturally occurring mutations or that arise during production of the monoclonal antibody preparation; such variants are generally present in minor amounts. In contrast to polyclonal antibody preparations that typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. Thus, the modifier "monoclonal" indicates the character of the antibody as being obtained from a population of substantially homogeneous antibodies and is not to be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present invention may be produced using techniques such as hybridoma technology, recombinant DNA technology, phage display technology, and human immunoglobulin G (HVIG) technology. Monoclonal antibodies can be produced by a variety of techniques, including, but not limited to, methods utilizing transgenic animals containing all or part of the phosphoryl locus; such methods and other exemplary methods for producing monoclonal antibodies are described herein.

[0058] A "naked antibody" refers to an antibody that is not conjugated to a heterologous moiety (e.g., a cytotoxic moiety) or radiolabel. The naked antibody may be present in a pharmaceutical formulation.

[0059] "Native antibodies" refer to naturally occurring immunoglobulin molecules with diverse structures. For example, native IgG antibodies are heterotetrameric glycoproteins of approximately 150,000 daltons, composed of two identical light chains and two identical heavy chains that are disulfide-bonded. From the N-terminus to the C-terminus, each heavy chain has a variable region (VH), also called a variable heavy domain or heavy chain variable domain, followed by three constant domains (CH1, CH2, and CH3). Similarly, from the N-terminus to the C-terminus, each light chain has a variable region (VL), also called a variable light domain or light chain variable domain, followed by a constant light (CL) domain. The light chain of an antibody can be assigned to one of two types, called kappa (κ) or lambda (λ), based on the amino acid sequence of its constant domain.

[0060] "Percent (%) amino acid sequence identity" to a reference polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical to those in the reference polypeptide sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity; any conservative substitutions are not considered as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in a variety of ways within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms required to achieve maximum alignment across the full length of the sequences being compared. However, for purposes herein, percent amino acid sequence identity values ​​are generated using the sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program is written by Genentech, Inc., and the source code, together with user documentation, is registered under the U.S. Copyright The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, California, and is registered under U.S. Copyright No. TXU510087. The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, California, or can be compiled from source code. The ALIGN-2 program should be compiled for use on UNIX operating systems, including Digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and cannot be varied.

[0061] In situations where ALIGN-2 is used for amino acid sequence comparison, the % amino acid sequence identity of a given amino acid sequence A to, with, or relative to a given amino acid sequence B (alternatively, it can be expressed as a given amino acid sequence A having or comprising a certain % amino acid sequence identity to, with, or relative to a given amino acid sequence B) is calculated as follows: 100 x fraction X / Y where X is the number of amino acid residues scored as perfect matches by the sequence alignment program ALIGN-2 in the program's alignment of A and B, and Y is the total number of amino acid residues in B. If the length of amino acid sequence A is not equal to the length of amino acid sequence B, the % amino acid sequence identity of A to B is calculated by multiplying the length of amino acid sequence A by the length of amino acid sequence B by the % amino acid sequence identity of B to A. It will be understood that the percent amino acid sequence identity does not equate to the percent identity. Unless specifically stated otherwise, all percent amino acid sequence identity values ​​used herein are obtained using the ALIGN-2 computer program as described in the immediately preceding paragraph.

[0062] The term "pharmaceutical formulation" or "pharmaceutical composition" refers to a preparation that is in a form that allows for the biological activity of the active ingredient contained therein and that does not contain additional components that are unacceptably toxic to the subject to which the formulation is administered.

[0063] A "pharmaceutically acceptable carrier" refers to an ingredient in a pharmaceutical formulation or composition, other than an active ingredient, that is non-toxic to a subject. Pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, or preservatives.

[0064] As used herein, "treatment" (and grammatical variations thereof, such as "treat" or "treating") refers to a clinical intervention aimed at altering the natural course of the individual being treated and may be performed for prophylaxis or during the course of clinical pathology. Desirable effects of treatment include, but are not limited to, prevention of disease onset or recurrence, alleviation of symptoms, diminishment of any direct or indirect pathological consequences of the disease, prevention of metastasis, reduction in the rate of disease progression, amelioration or palliation of the disease state, and remission or improved prognosis. In some embodiments, the antibodies of the invention are used to delay disease onset or slow the progression of the disease.

[0065] The term "vector," as used herein, refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is linked. The term includes vectors as self-replicating nucleic acid structures, as well as vectors that integrate into the genome of a host cell into which they have been introduced. Certain vectors are capable of directing the expression of nucleic acids to which they are operatively linked. Such vectors are referred to herein as "expression vectors."

[0066] II. Compositions and Methods Anti-CD112R antibodies, compositions comprising the described antibodies, and methods of their use are provided.

[0067] A. Exemplary Anti-CD112R Antibodies The sequence listing below provides sequences for certain embodiments of the antibodies disclosed and claimed herein.

[0068] In certain embodiments, antibodies are provided that bind to CD112R and / or block binding of CD112R to CD112 and / or enhance T cell and NK cell activation. In some embodiments, antibodies are provided that bind to CD112R. In some embodiments, antibodies are provided that block CD112R binding to CD112. In some embodiments, antibodies are provided that enhance CD226, T cell, and / or NK cell activation.

[0069] Inhibition of binding between CD112R and CD112, such as on T cells and NK cells, can be determined by measuring inhibition of binding of cells to which CD112R binds in the presence and absence of the antibody.

[0070] Provided herein are antibodies that specifically bind to CD112R.

[0071] In some embodiments, the antibody binds to human CD112R.

[0072] In some embodiments, the antibodies bind to human CD112R and block the interaction of human CD112R with human CD112. In some embodiments, the antibodies bind to human CD112R and block the interaction of human CD112R with human CD112, but do not block the interaction of mouse CD112R with mouse CD112. In some embodiments, antibodies that bind to human CD112R and block the interaction of human CD112R with human CD112, but do not block the interaction of mouse CD112R with mouse CD112, include antibodies 32, 33, 34, 35, and 36.

[0073] In certain embodiments, the CD112R antibody comprises a heavy chain variable region ("VH") comprising the VH CDR1, CDR2, and / or CDR3 of any of the CD112R antibodies provided herein (i.e., antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36).

[0074] In certain embodiments, the CD112R antibody comprises a VH comprising the VH CDR1, CDR2, and / or CDR3 of any of the CD112R antibodies provided herein, and a VL comprising the CDR1, CDR2, and / or CDR3 of any of the CD112R antibodies provided herein. In certain embodiments, a CD112R antibody comprises a VH comprising the VH CDR1, CDR2, and / or CDR3 of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36, and a VL comprising the VL CDR1, CDR2, and / or CDR3 of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36, optionally wherein the VH and VL CDRs are from the same antibody clone.

[0075] In some embodiments, an antibody is provided that comprises: (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1; (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2; (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 4; (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 6; or (b) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 101, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 102, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 103, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 104, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 105, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 106, or (c) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 201, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 202, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 203, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 204, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 205, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 206, or (d) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 301, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 302, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 303, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 304, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 305, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 306, or (e) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 401, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 402, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 403, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 404, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 405, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 406, or (f) HCDR1 comprising the amino acid sequence of SEQ ID NO: 501; (b) HCDR2 comprising the amino acid sequence of SEQ ID NO: 502; (c) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 503, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 504, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 505, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 506, or (g) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 601, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 602, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 603, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 604, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 605, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 606, or (h) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 701, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 702, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 703, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 704, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 705, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 706, or (i) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 801, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 802, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 803, with or without (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 804, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 805, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 806, or (j) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 901, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 902, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 903, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 904, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 905, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 906, or (k) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 1003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 1004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 1005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 1006, or (l) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 2001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 2003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 2004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 2005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 2006, or (m) (a) an HCDR1 comprising the amino acid sequence of SEQ ID NO: 3001, (b) an HCDR2 comprising the amino acid sequence of SEQ ID NO: 3002, (c) an HCDR3 comprising the amino acid sequence of SEQ ID NO: 3003, (d) an LCDR1 comprising the amino acid sequence of SEQ ID NO: 3004, (e) an LCDR2 comprising the amino acid sequence of SEQ ID NO: 3005, and (f) an LCDR3 comprising the amino acid sequence of SEQ ID NO: 3006, or (n) (a) HCDR1 comprising the amino acid sequence of SEQ ID NO: 4001, (b) HCDR2 comprising the amino acid sequence of SEQ ID NO: 4002, (c) HCDR3 comprising the amino acid sequence of SEQ ID NO: 4003, (d) LCDR1 comprising the amino acid sequence of SEQ ID NO: 4004, (e) LCDR2 comprising the amino acid sequence of SEQ ID NO: 4005, and (f) LCDR3 comprising the amino acid sequence of SEQ ID NO: 4006.

[0076] In certain embodiments, a CD112R antibody comprises a VL that includes the VL CDR1, CDR2, and CDR3 of any of the CD112R antibodies provided herein. In certain embodiments, a CD112R antibody comprises a VL that includes the VL CDR1, CDR2, and CDR3 of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0077] In some embodiments, the CD112R antibody (a) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 2, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 2; or (b) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 5, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 5; or (c) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 44, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 44; or (d) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 58, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 58; or (e) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 10, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 10; or (f) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 38, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 38; or (g) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 15, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 15; or (h) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 35, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 35; or (i) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 47, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 47; or (j) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 46, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 46; or (k) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 32, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 32; or (l) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 33, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 33; or (m) a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 34, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 34; or (n) It may comprise a VH comprising the amino acid sequences of VH CDR1, CDR2, and CDR3 of antibody clone No. 36, and a VL comprising VL CDR1, CDR2, and CDR3 of antibody clone No. 36.

[0078] The sequence listing below provides the heavy and light chain variable region sequences of certain disclosed antibodies.

[0079] In certain embodiments, the CD112R antibody comprises a VH comprising the amino acid sequence of the VH of any of the CD112R antibodies provided herein. In certain embodiments, the CD112R antibody comprises antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, and a VH comprising the amino acid sequence of any one of VH of 46, 47, 32, 33, 34, or 36.

[0080] In some embodiments, the CD112R antibody has 1, 2, 3, 4, or 5 amino acid substitutions outside the complementarity determining regions (CDRs), such as 1, 2, 3, 4, or 5 conservative substitutions outside the CDRs, but comprises a VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36. In some embodiments, the CD112R antibody has 1, 2, 3, 4, or 5 revert substitutions outside the complementarity determining regions (CDRs), but comprises a VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0081] In some embodiments, the CD112R antibody has one, two, three, four, or five amino acid substitutions in the framework regions of the VH sequence, such as one, two, three, four, or five conservative substitutions, but comprises a VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36. In some embodiments, the CD112R antibody has one, two, three, four, or five back substitutions in the framework regions of the VH sequence but comprises a VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0082] In some embodiments, the CD112R antibody comprises the VH and VL CDRs of any of the CD112R antibodies described herein, wherein each CDR comprises 0, 1, 2, or 3 amino acid additions, substitutions (e.g., conservative substitutions), or deletions.

[0083] In certain embodiments, a CD112R antibody comprises a VH CDR1, CDR2, and CDR3 comprising the amino acid sequence of the VH CDRs of any of the CD112R antibodies provided herein, and comprises a VH that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH of any of the CD112R antibodies provided herein. In certain embodiments, a CD112R antibody comprises a VH that comprises an amino acid sequence that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of the VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36. In certain embodiments, the VH of the antibody differs from that of the VH sequence shown in the sequence listing by 1, 2, 3, 4, or 5 amino acid substitutions in the framework regions of the VH sequence, such as 1, 2, 3, 4, or 5 conservative substitutions. In certain embodiments, the VH of the antibody differs from that of the VH sequence shown in the sequence listing by 1, 2, 3, 4, or 5 back substitutions in the framework regions of the VH sequence.

[0084] In certain embodiments, a CD112R antibody comprises a VH consisting of the amino acid sequence of the VH of any of the CD112R antibodies provided herein, hi certain embodiments, a CD112R antibody comprises a VH consisting of the amino acid sequence of the VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0085] In certain embodiments, a CD112R antibody comprises a VL comprising the amino acid sequence of the VL of any of the CD112R antibodies provided herein. The CD112R antibody comprises a VL comprising the amino acid sequence of the VL of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36. In certain embodiments, the CD112R antibody comprises a VL CDR1, CDR2, and CDR3 comprising the amino acid sequences of the VL CDRs of any of the CD112R antibodies provided herein, and which is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VL of any of the CD112R antibodies provided herein. In certain embodiments, a CD112R antibody comprises a VL comprising an amino acid sequence that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of the VL of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36. In certain embodiments, the VL of the antibody differs from that of the VL sequence set forth in the sequence listing by one, two, three, four, or five amino acid substitutions in the framework regions of the VL sequence, such as one, two, three, four, or five conservative substitutions. In certain embodiments, the VL of the antibody differs from that of the VL sequence set forth in the sequence listing by one, two, three, four, or five back substitutions.

[0086] In certain embodiments, a CD112R antibody comprises a VL consisting of the amino acid sequence of the VL of any of the CD112R antibodies provided herein, hi certain embodiments, a CD112R antibody comprises a VL consisting of the amino acid sequence of the VL of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0087] In certain embodiments, a CD112R antibody comprises a VH comprising the amino acid sequence of the VH of any one of the CD112R antibodies provided herein, and a VL comprising the amino acid sequence of the VL of any of the same CD112R antibodies provided herein. In certain of these embodiments, the CD112R antibody comprises a VH comprising the amino acid sequence of the VH of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36, and a VL comprising the amino acid sequence of the VL of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36, wherein optionally, the VH and VL are from the same antibody clone number.

[0088] In some embodiments, the VH of the antibody has 1, 2, 3, 4, or 5 amino acid substitutions, such as 1, 2, 3, 4, or 5 conservative substitutions, in the framework regions of the VH sequence of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36, and the VL is of any one of the same antibodies from the list above. In certain embodiments, the VH of the antibody has 1, 2, 3, 4, or 5 conservative substitutions in the framework regions of the VH sequence of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0089] In certain embodiments, the CD112R antibody comprises a VH and a VL comprising the amino acid sequences of the VH and VL of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0090] In certain embodiments, the CD112R antibody comprises a VH CDR1, CDR2, and VH CDR3 comprising the amino acid sequences of the VH CDRs of any of the CD112R antibodies provided herein. and CDR1, CDR2, and CDR3, as well as VL CDRs comprising the amino acid sequences of any of the CD112R antibodies provided herein, and comprising a VH and VL that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical, respectively, to the corresponding VH and VL of any of the CD112R antibodies provided herein. In certain embodiments, the VH and VL of the antibody differ from the VH and VL sequences set forth in the sequence listing by one, two, three, four, or five amino acid substitutions in the framework regions of the sequences, such as one, two, three, four, or five conservative substitutions, or one, two, three, four, or five back substitutions.

[0091] In certain embodiments, a CD112R antibody comprises a VH and a VL consisting of the amino acid sequences of the VH and VL of any of the CD112R antibodies provided herein. In certain embodiments, a CD112R antibody comprises a VH and a VL consisting of the amino acid sequences of the VH and VL, respectively, of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, and 36.

[0092] The CD112R antibody (a) a VH comprising the amino acid sequence of the VH of antibody clone No. 2, and a VL comprising the amino acid sequence of the VL of antibody clone No. 2; or (b) a VH comprising the amino acid sequence of the VH of antibody clone No. 5, and a VL comprising the amino acid sequence of the VL of antibody clone No. 5; or (c) a VH comprising the amino acid sequence of the VH of antibody clone No. 44, and a VL comprising the amino acid sequence of the VL of antibody clone No. 44, or (d) a VH comprising the amino acid sequence of the VH of antibody clone No. 58, and a VL comprising the amino acid sequence of the VL of antibody clone No. 58; or (e) a VH comprising the amino acid sequence of the VH of antibody clone No. 10, and a VL comprising the amino acid sequence of the VL of antibody clone No. 10; or (f) a VH comprising the amino acid sequence of the VH of antibody clone No. 38, and a VL comprising the amino acid sequence of the VL of antibody clone No. 38, or (g) a VH comprising the amino acid sequence of the VH of antibody clone No. 15, and a VL comprising the amino acid sequence of the VL of antibody clone No. 15, or (h) a VH comprising the amino acid sequence of the VH of antibody clone No. 35, and a VL comprising the amino acid sequence of the VL of antibody clone No. 35, or (i) a VH comprising the amino acid sequence of the VH of antibody clone No. 47, and a VL comprising the amino acid sequence of the VL of antibody clone No. 47; or (j) a VH comprising the amino acid sequence of the VH of antibody clone No. 46, and a VL comprising the amino acid sequence of the VL of antibody clone No. 46, or (k) a VH comprising the amino acid sequence of the VH of antibody clone No. 32, and a VL comprising the amino acid sequence of the VL of antibody clone No. 32; or (l) a VH comprising the amino acid sequence of the VH of antibody clone No. 33, and a VL comprising the amino acid sequence of the VL of antibody clone No. 33, or (m) a VH comprising the amino acid sequence of the VH of antibody clone No. 34, and a VL comprising the amino acid sequence of the VL of antibody clone No. 34, or (n) It may comprise a VH comprising the amino acid sequence of the VH of antibody clone No. 36, and a VL comprising the amino acid sequence of the VL of antibody clone No. 36.

[0093] The CD112R antibody (a) a VH comprising the VH CDR of the VH of antibody clone No. 2, and a VL comprising the VL CDR of antibody clone No. 2, and a VH and VL of antibody clone No. 2 that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, or at least VH and VL amino acid sequences that are at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical; or (b) a VH comprising the VH CDRs of the VH of antibody clone No. 5, and a VL comprising the VL CDRs of antibody clone No. 5, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 5; or (c) a VH comprising the VH CDRs of the VH of antibody clone No. 44, and a VL comprising the VL CDRs of antibody clone No. 44, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 44; or (d) a VH comprising the VH CDRs of the VH of antibody clone No. 58, and a VL comprising the VL CDRs of antibody clone No. 58, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 58; or (e) a VH comprising the VH CDRs of the VH of antibody clone No. 10, and a VL comprising the VL CDRs of antibody clone No. 10, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 10; or (f) a VH comprising the VH CDRs of the VH of antibody clone No. 38, and a VL comprising the VL CDRs of antibody clone No. 38, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 38; or (g) a VH comprising the VH CDRs of the VH of antibody clone No. 15, and a VL comprising the VL CDRs of antibody clone No. 15, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 15; or (h) a VH comprising the VH CDRs of the VH of antibody clone No. 35, and a VL comprising the VL CDRs of antibody clone No. 35, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 35; or (i) a VH comprising the VH CDRs of the VH of antibody clone No. 47, and a VL comprising the VL CDRs of antibody clone No. 47, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 47; or (j) a VH comprising the VH CDRs of the VH of antibody clone No. 46, and a VL comprising the VL CDRs of antibody clone No. 46, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 46; or (k) A VH comprising the VH CDR of the VH of antibody clone No. 32, and a VL comprising the VL CDR of antibody clone No. 32, and a VH and VL of antibody clone No. 32 that are at least 90%, at least 91%, at least 92%, at least 93%, or at least 94% identical to the VH and VL of antibody clone No. 32 , VH and VL amino acid sequences that are at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical, or (l) a VH comprising the VH CDRs of the VH of antibody clone No. 33, and a VL comprising the VL CDRs of antibody clone No. 33, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 33; or (m) a VH comprising the VH CDRs of the VH of antibody clone No. 34, and a VL comprising the VL CDRs of antibody clone No. 34, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 34; or (n) A VH comprising the VH CDRs of the VH of antibody clone No. 36, and a VL comprising the VL CDRs of antibody clone No. 36, and VH and VL amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the VH and VL of antibody clone No. 36.

[0094] In some of the above embodiments, VH and / or VL may differ from the respective species sequences by the presence of 1, 2, 3, 4, or 5 amino acid substitutions, such as 1, 2, 3, 4, or 5 conservative substitutions, hi some embodiments, VH may contain 1, 2, 3, 4, or 5 back substitutions.

[0095] The CD112R antibody (a) a VH consisting of the amino acid sequence of the VH of antibody clone No. 2 and a VL consisting of the VL of antibody clone No. 2, or (b) a VH consisting of the amino acid sequence of the VH of antibody clone No. 5 and a VL consisting of the VL of antibody clone No. 5, or (c) a VH consisting of the amino acid sequence of the VH of antibody clone No. 44 and a VL consisting of the VL of antibody clone No. 44, or (d) a VH consisting of the amino acid sequence of the VH of antibody clone No. 58 and a VL consisting of the VL of antibody clone No. 58, or (e) a VH consisting of the amino acid sequence of the VH of antibody clone No. 10 and a VL consisting of the VL of antibody clone No. 10, or (f) a VH consisting of the amino acid sequence of the VH of antibody clone No. 38 and a VL consisting of the VL of antibody clone No. 38, or (g) a VH consisting of the amino acid sequence of the VH of antibody clone No. 15 and a VL consisting of the VL of antibody clone No. 15, or (h) a VH consisting of the amino acid sequence of the VH of antibody clone No. 35 and a VL consisting of the VL of antibody clone No. 35, or (i) a VH consisting of the amino acid sequence of the VH of antibody clone No. 47 and a VL consisting of the VL of antibody clone No. 47; or (j) a VH consisting of the amino acid sequence of the VH of antibody clone No. 46 and a VL consisting of the VL of antibody clone No. 46, or (k) a VH consisting of the amino acid sequence of the VH of antibody clone No. 32 and a VL consisting of the VL of antibody clone No. 32, or (l) a VH consisting of the amino acid sequence of the VH of antibody clone No. 33 and a VL consisting of the VL of antibody clone No. 33, or (m) a VH consisting of the amino acid sequence of the VH of antibody clone No. 34 and a VL consisting of the VL of antibody clone No. 34, or (n) It may comprise a VH consisting of the amino acid sequence of the VH of antibody clone No. 36, and a VL consisting of the VL of antibody clone No. 36.

[0096] In certain embodiments, the CD112R antibody comprises the variable region and / or any of the variable region CDRs 1-3 of the antibodies described above and elsewhere herein, such as in the sequence listing.

[0097] In some embodiments, the CD112R antibody is an IgG antibody, such as an IgG1, IgG2, IgG3, or IgG4 antibody, or a modified form thereof as described in the following sections. In some embodiments, the constant region has an effector function, and in some embodiments, the constant region does not comprise an effector.

[0098] In certain embodiments, a CD112R antibody comprises a heavy chain comprising the amino acid sequence of the heavy chain (HC) of any of the CD112R antibodies provided herein. In certain embodiments, a CD112R antibody comprises a heavy chain comprising the amino acid sequence of the heavy chain of any one of antibody clone numbers 2, 5, 44, 58, 10, 38, 15, 35, 46, 47, 32, 33, 34, or 36.

[0099] In some embodiments, the CD112R antibody (a) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 2 and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 2, or (b) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 5 and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 5, or (c) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 44, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 44, or (d) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 58, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 58, or (e) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 10, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 10; or (f) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 38 and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 38, or (g) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 15, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 15, or (h) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 35, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 35, or (i) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 47, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 47, or (j) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 46, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 46, or (k) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 32, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 32, or (l) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 33, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 33, or (m) a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 34, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 34, or (n) It may comprise a heavy chain comprising the amino acid sequence of the heavy chain of antibody clone No. 36, and a light chain comprising the amino acid sequence of the light chain of antibody clone No. 36.

[0100] The CD112R antibody (a) a heavy chain (HC) containing the HC CDRs of antibody clone No. 2, and a light chain (LC) containing the LC CDRs of antibody clone No. 2, and the HC and LC of antibody clone No. 2 or HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical, respectively, to (b) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 5, and a light chain (LC) comprising the LC CDRs of antibody clone No. 5, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 5, respectively; or (c) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 44, and a light chain (LC) comprising the LC CDRs of antibody clone No. 44, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 44, respectively; or (d) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 58, and a light chain (LC) comprising the LC CDRs of antibody clone No. 58, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 58, respectively; or (e) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 10, and a light chain (LC) comprising the LC CDRs of antibody clone No. 10, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 10, respectively; or (f) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 38, and a light chain (LC) comprising the LC CDRs of antibody clone No. 38, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 38, respectively; or (g) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 15, and a light chain (LC) comprising the LC CDRs of antibody clone No. 15, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 15, respectively; or (h) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 35, and a light chain (LC) comprising the LC CDRs of antibody clone No. 35, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 35, respectively; or (i) a HC comprising the HC CDRs of the HC of antibody clone No. 47, and a light chain (LC) comprising the LC CDRs of antibody clone No. 47, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 47, respectively; or (j) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 46, and a light chain (LC) comprising the LC CDRs of antibody clone No. 46, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 46, respectively; or (k) a light chain (LC) comprising the HC CDR of the HC of antibody clone No. 32 and the LC CDR of antibody clone No. 32, and a light chain (LC) comprising the HC and LC of antibody clone No. 32 HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical, respectively; or (l) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 33, and a light chain (LC) comprising the LC CDRs of antibody clone No. 33, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 33, respectively; or (m) a light chain (LC) comprising the HC CDRs of the HC of antibody clone No. 34, and a light chain (LC) comprising the LC CDRs of antibody clone No. 34, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 34, respectively; or (n) a HC comprising the HC CDRs of the HC of antibody clone No. 36, and a light chain (LC) comprising the LC CDRs of antibody clone No. 36, and HC and LC amino acid sequences that are at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the HC and LC of antibody clone No. 36, respectively.

[0101] In some of the above embodiments, the HC and / or LC may differ from the sequence of each of the species by the presence of 1, 2, 3, 4, or 5 amino acid substitutions, such as 1, 2, 3, 4, or 5 conservative substitutions. In some of the above embodiments, the HC and / or LC may differ from the sequence of each of the species by the presence of 1, 2, 3, 4, or 5 amino acid substitutions, such as 1, 2, 3, 4, or 5 back substitutions.

[0102] 1. Exemplary Classes of Antibodies with Shared Structural and Functional Characteristics In some embodiments, anti-CD112R antibodies are provided. In certain embodiments, anti-CD112 antibodies share certain structural and / or functional characteristics. In some embodiments, a class of antibodies includes a parent antibody and its affinity matured variants. One exemplary class of antibodies includes, but is not limited to, parent clone 32 and its affinity matured variants. In some embodiments, affinity matured variants include antibodies 33, 34, 35, and 36. In some embodiments, affinity matured variants include antibodies with conservative substitutions compared to antibodies 32, 33, 34, 35, and 36.

[0103] a. Structural Features of Exemplary Classes of Antibodies In some embodiments, anti-CD112R antibodies share structural features such as, for example, those shown in Figures 14 and 15. It should be understood that when a "class" or "member of a class" of antibodies is described herein, embodiments describing a single anti-CD112R antibody or multiple anti-CD112R antibodies are encompassed / contemplated. In some embodiments, anti-CD112R antibodies comprise identical HCDR3s. In some embodiments, anti-CD112R antibodies comprise identical LCDR1s. In some embodiments, anti-CD112R antibodies comprise identical LCDR2s. In some embodiments, anti-CD112R antibodies comprise identical LCDR3s. In some embodiments, anti-CD112R antibodies comprise identical HCDR3s and identical LCDR1s, LCDR2s, and / or LCDR3s. In some embodiments, the anti-CD112R antibody comprises an HCDR3 comprising the amino acid sequence of SEQ ID NO: 1003, and / or an LCDR1 comprising the amino acid sequence of SEQ ID NO: 1004, and / or an LCDR2 comprising the amino acid sequence of SEQ ID NO: 1005, and / or an LCDR3 comprising the amino acid sequence of SEQ ID NO: 1006.

[0104] In some embodiments, each member of the class of antibodies is selected from the group consisting of SEQ ID NOs: 1007, 1008, 1009, 1010, 1011, 1012, 1013, 1014, 1015, 1016, 1017, 1018, 1019, 1020, 1021, 1022, 1023, , 1009, and 1010. In some embodiments, each member of the class of antibodies comprises a heavy chain variable region amino acid sequence that is at least 90%, at least 91%, at least 92%, or at least 93% identical to the amino acid sequence of SEQ ID NO: 1012, and any and all of the sequence diversity relative to SEQ ID NO: 1012 is in HCDR1 and / or HCDR2. In some embodiments, each member of the class of antibodies comprises a light chain framework region comprising the amino acid sequence of SEQ ID NO: 1013, 1014, 1015, and 1016. In some embodiments, each member of the class of antibodies comprises a light chain variable region amino acid sequence of SEQ ID NO: 1018.

[0105] Members of this exemplary class of antibodies may contain some diversity in the amino acid sequences of HCDR1 and HCDR2. In some embodiments, each member of the class of antibodies comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001, or the amino acid sequence of SEQ ID NO: 1001 with one, two, or three amino acid changes at positions 4, 5, and / or 6 of SEQ ID NO: 1001. In some embodiments, the class of antibodies comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 1001 with one, two, or three amino acid changes at positions that differ among the amino acids of SEQ ID NOs: 1001, 2001, 3001, 4001, and 701 shown in Figure 14. In some embodiments, one or more of the one, two, or three amino acid changes are not conservative substitutions. In some embodiments, one or more of the one, two, or three amino acid changes are conservative substitutions. In certain embodiments, members of the class of antibodies comprise an HCDR1 comprising the amino acid sequence of SEQ ID NO: 2001, 3001, 701, or 4001.

[0106] In some embodiments, each member of the class of antibodies comprises an HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002, or the amino acid sequence of SEQ ID NO: 1002 with 1, 2, 3, 4, or 5 amino acid changes at positions 1, 3, 5, 6, and / or 8 of SEQ ID NO: 1002. In some embodiments, the class of antibodies comprises an HCDR2 comprising the amino acid sequence of SEQ ID NO: 1002 with 1, 2, 3, 4, or 5 amino acid changes at positions that differ among the amino acids of SEQ ID NOs: 1002, 2002, 3002, 4002, and 702 shown in Figure 14. In some embodiments, one or more of the 1, 2, 3, 4, or 5 amino acid changes are not conservative substitutions. In some embodiments, one or more of the 1, 2, 3, 4, or 5 amino acid changes are conservative substitutions. In certain embodiments, members of the class of antibodies comprise an HCDR2 comprising the amino acid sequence of SEQ ID NO: 2002, 3002, 702, or 4002.

[0107] In some embodiments, each member of a class of antibodies comprises identical heavy and light chain framework regions.

[0108] b. Functional Characteristics of Exemplary Classes of Antibodies In some embodiments, anti-CD112R antibodies are provided, which share the special technical effects of binding to human CD112R, blocking the interaction of human CD112R with CD112, and failing to block the interaction of mouse CD112R with CD112. In some embodiments, each member of the antibody class binds to human CD112R and blocks the binding interaction between human CD112 and human CD112R. In some embodiments, each member of the antibody class does not block the binding interaction between mouse CD112 and mouse CD112R. While members of the antibody class do not block the interaction between mouse CD112 and mouse CD112R, members of the antibody class partially inhibit the binding interaction between mouse CD112 and mouse CD112R or do not inhibit the binding interaction between mouse CD112 and mouse CD112R. In some such embodiments, members of the antibody class do not inhibit the interaction between mouse CD112R and mouse CD112 by more than 50%. In some embodiments, each member of a class of antibodies is a soluble mouse In some embodiments, the antibodies exhibit at least some binding to CD112R. In some embodiments, the antibodies are fully human or humanized. In some embodiments, each member of a class of antibodies binds to the same epitope on human CD112R.

[0109] 2. Antibody fragment In certain embodiments, the antibodies provided herein are antibody fragments. Antibody fragments include, but are not limited to, Fab, Fab', Fab'-SH, F(ab')2, Fv, and scFv fragments, as well as other fragments described below. For a review of certain antibody fragments, see Hudson et al. Nat. Med. 9:129-134 (2003). For a review of scFv fragments, see, e.g., Pluckthun, in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., (Springer-Verlag, New York), pp. 269-315 (1994); WO 93 / 16185; and U.S. Patent Nos. 5,571,894 and 5,587,458. See US Pat. No. 5,869,046 for a discussion of Fab and F(ab')2 fragments that contain salvage receptor binding epitope residues and have increased in vivo half-lives.

[0110] Diabodies are antibody fragments with two antigen-binding sites, which may be bivalent or bispecific. See, e.g., EP 404,097, WO 1993 / 01161, Hudson See Hudson et al., Nat. Med. 9:129-134 (2003), and Hollinger et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993). Triabodies and tetrabodies are also described in Hudson et al., Nat. Med. 9:129-134 (2003).

[0111] Single-domain antibodies are antibody fragments that contain all or part of the heavy chain variable domain or all or part of the light chain variable domain of an antibody. In certain embodiments, single-domain antibodies are human single-domain antibodies (Domantis, Inc., Waltham, MA; see, e.g., U.S. Patent No. 6,248,516 B1).

[0112] Antibody fragments can be produced by a variety of techniques, including, but not limited to, proteolytic digestion of intact antibodies, as well as production by recombinant host cells (e.g., E. coli or phage), as described herein.

[0113] 3. Multispecific antibodies In certain embodiments, the antibodies provided herein are multispecific antibodies, e.g., bispecific antibodies. Multispecific antibodies are monoclonal antibodies that have binding specificities for at least two different sites. In certain embodiments, one of the binding specificities is for CD112R and the other is for any other antigen. In certain embodiments, one of the binding specificities is for CD112R and the other is for an antigen independently selected from one (in the bispecific case) or more (in the multispecific case) of PD-1, PD-L1, CTLA-4, Lag-3, TIM-3, TIGIT, CD96, PVRL1, PVRL2, PVRL3, PVRL4, CD155, STING, CD47, CD39, and IL-27. In certain embodiments, bispecific antibodies can bind to two different epitopes of CD112R. Bispecific antibodies can also be used to localize cytotoxic agents to cells expressing CD112R. Bispecific antibodies can be prepared as full-length antibodies or antibody fragments.

[0114] A technique for generating multispecific antibodies is the recombinant co-expression of two immunoglobulin heavy-chain-light pairs with different specificities (Milstein and Cuello, Nature 305:537 (1983)), WO 93 / 08829, and Traunecker et al., EMBO J. 10:3655 (1991)), and "knob-in-hole" operation (see, e.g., U.S. Pat. No. 5,731,168). Multispecific antibodies can be made by manipulating electrostatic steering effects to create antibody Fc-heterodimeric molecules (WO 2009 / 089004 A1), cross-linking two or more antibodies or fragments (see, e.g., U.S. Pat. No. 4,676,980, and Brennan et al., Science, 229:81 (1985)), using leucine zippers to produce bispecific antibodies (see, e.g., Kostelny et al., J. Immunol., 148(5):1547-1553 (1992)), using "diabody" technology to create bispecific antibody fragments (see, e.g., Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993)), and using single-chain Fv (sFv) dimers (see, e.g., Gruber et al., J. Immunol., 90:6444-6448 (1993)). al., J. Immunol., 152:5368 (1994)), and by preparing trispecific antibodies as described, for example, in Tutt et al. J. Immunol. 147:60 (1991).

[0115] Engineered antibodies with three or more functional antigen binding sites, including "octopus antibodies," are also included herein (see, eg, US2006 / 0025576A1).

[0116] The antibodies or fragments herein also include "Dual Acting F antibodies" or "DAFs" that contain an antigen binding site that binds to CD112R as well as another, different antigen (see, e.g., US2008 / 0069820).

[0117] 4. Antibody Variants In certain embodiments, amino acid sequence variants of the antibodies provided herein are contemplated. For example, it may be desirable to improve the binding affinity and / or other biological properties of the antibody. Amino acid sequence variants of antibodies can be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody or by peptide synthesis. Such modifications include, for example, deletions from, and / or insertions into, and / or substitutions of residues within the amino acid sequence of the antibody. Any combination of deletions, insertions, and substitutions can be made to arrive at the final construct, provided that the final construct possesses the desired properties, e.g., antigen binding.

[0118] 5. Substitution, insertion, and deletion variants In certain embodiments, antibody variants with one or more amino acid substitutions are provided. Sites of interest for substitutional mutagenesis include HVRs and FRs. Conservative substitutions are shown in Table 1 as being "exemplary substitutions." Amino acid substitutions can be introduced into an antibody of interest, and the products can be screened for a desired activity, such as retained / improved antigen binding, reduced immunogenicity, or improved ADCC or CDC. [Table 1] Amino acids can be grouped according to the following general side chain properties: (1) Hydrophobic: Norleucine, Met, Ala, Val, Leu, Ile, (2) Neutral hydrophilicity: Cys, Ser, Thr, Asn, Gln, (3) Acidic: Asp, Glu, (4) Basic: His, Lys, Arg, (5) Residues that affect chain orientation: Gly, Pro, (6) Aromatic: Trp, Tyr, Phe.

[0119] Non-conservative substitutions would involve exchanging a member of one of these classes for another class.

[0120] Modifications (e.g., substitutions) can be made in HVRs, for example, to improve antibody affinity. Such modifications can be made in HVR "hot spots," i.e., residues encoded by codons that undergo frequent mutation during the somatic maturation process (see, e.g., Chowdhury, Methods Mol. Biol. 207:179-196 (2008)), and / or residues that contact the antigen, and the resulting variants VH or VL are tested for binding affinity. Affinity maturation by construction and reselection from secondary libraries can be performed, for example, as described by Hoogenboom et al. in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, ( (2001). In some embodiments of affinity maturation, diversity is introduced into the variable genes selected for maturation by any of a variety of methods (e.g., error-prone PCR, chain shuffling, or oligonucleotide-directed mutagenesis). A secondary library is then created. The library is then screened to identify any antibody variants with the desired affinity. Another method for introducing diversity involves an HVR-directed approach, in which several HVR residues (e.g., 4-6 residues at a time) are randomized. HVR residues involved in antigen binding can be specifically identified using, for example, alanine scanning mutagenesis or modeling. CDR-H3 and CDR-L3 in particular are often targeted.

[0121] In certain embodiments, substitutions, insertions, or deletions can occur within one or more HVRs as long as such modifications do not substantially reduce the ability of the antibody to bind to the antigen. For example, conservative modifications (e.g., conservative substitutions provided herein) that do not substantially reduce binding affinity can be made in HVRs. Such modifications can, for example, be outside the antigen contact residues in the HVRs. In certain embodiments of the variant VH and VL sequences provided above, each HVR is either unaltered or contains no more than one, two, or three amino acid substitutions.

[0122] A useful method for identifying antibody residues or regions that can be targeted for mutagenesis is called "alanine scanning mutagenesis," described by Cunningham and Wells (1989) Science, 244:1081-1085. In this method, certain residues or groups among target residues (e.g., charged residues such as arg, asp, his, lys, and glu) are identified and replaced with neutral or negatively charged amino acids (e.g., alanine or polyalanine) to determine whether the antibody's interaction with the antigen is affected. Further substitutions can be introduced at amino acid locations that demonstrate functional sensitivity to the initial substitution. Alternatively, or in addition, a crystal structure of an antigen-antibody complex can be used to identify contact points between the antibody and antigen. Such contact residues and neighboring residues can be targeted or eliminated as candidates for substitution. Variants can be screened to determine whether they contain the desired properties.

[0123] Amino acid sequence insertions include amino- and / or carboxyl-terminal fusions, ranging in length from one residue to polypeptides containing 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. An example of a terminal insertion includes an antibody with an N-terminal methionyl residue. Other insertional variants of the antibody molecule include the fusion to the N- or C-terminus of the antibody to an enzyme (e.g., for ADEPT) or a polypeptide which increases the serum half-life of the antibody.

[0124] 6. Glycosylation variants In certain embodiments, the antibodies provided herein are modified to increase or decrease the extent to which the antibody is glycosylated. Addition or deletion of glycosylation sites to an antibody can be conveniently accomplished by altering the amino acid sequence such that one or more glycosylation sites are created or removed.

[0125] If the antibody comprises an Fc region, the carbohydrate attached thereto may be modified. Natural antibodies produced by mammalian cells typically contain branched, biantennary oligosaccharides, generally linked by an N-linkage to Asn297 in the CH2 domain of the Fc region. See, e.g., Wright et al. TIBTECH 15:26-32 (1997). The oligosaccharides may contain various carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose attached to GlcNAc in the "stem" of the biantennary oligosaccharide structure. In some embodiments, the antibodies of the present invention may contain a biantennary oligosaccharide. Modifications of the oligosaccharides in the antibody can be made to generate antibody variants with certain improved properties.

[0126] In some embodiments, antibody variants are provided that have carbohydrate structures lacking fucose attached (directly or indirectly) to the Fc region. For example, the amount of fucose in such antibodies can be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is determined by calculating the average amount of fucose in the glycan at Asn297 relative to the sum of all glycan structures (e.g., complex, hybrid, and high-mannose structures) attached to Asn297 as measured by MALDI-TOF mass spectrometry, e.g., as described in WO 2008 / 077546. Asn297 refers to an asparagine residue located at approximately position 297 (Eu numbering of Fc region residues) in the Fc region; however, due to minor sequence diversity in antibodies, Asn297 can also be located approximately ±3 amino acids upstream or downstream from position 297, i.e., between positions 294 and 300. Such fucosylated variants may have improved ADCC function. See, e.g., U.S. Patent Publication Nos. US2003 / 0157108 (Presta, L.) and US2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd.). Examples of publications related to "defucosylated" or "fucose-deficient" antibody variants include US2003 / 0157108, WO2000 / 61739, WO2001 / 29246, US2003 / 0115614, US2002 / 0164328, US2004 / 0093621, and US2004 / 0132140. , US2004 / 0110704, US2004 / 0110282, US2004 / 0109865, WO2003 / 085119, WO2003 / 0 84570, WO2005 / 035586, WO2005 / 035778, WO2005 / 053742, WO2002 / 031140, Okazaki et al.J.Mol.Biol.336:1239-1249(2004), Yamane-Ohnuki et al.Biotech.Bioeng.87:614(2004).An example of a cell line capable of producing defucosylated antibodies is Lec13 CHO cells, which are deficient in protein fucosylation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); U.S. Patent Application No. US2003 / 0157108 A1; Presta, L; and WO2004 / 056312 A1, Adams). et al., especially Example 11), and knockout cell lines such as alpha-1,6-fucosyltransferase gene, FUT8, knockout CHO cells (see, e.g., Yamane-Ohnuki et al. Biotech. Bioeng. 87:614 (2004), Kanda, Y. et al., Biotechnol. Bioeng., 94(4):680-688 (2006), and WO2003 / 085107).

[0127] Further provided are antibody variants having bisected oligosaccharides, for example, where a biantennary oligosaccharide attached to the Fc region of the antibody is bisected by GlcNAc. Such antibody variants may have reduced fucosylation and / or improved ADCC function. Examples of such antibody variants are described, for example, in WO 2003 / 011878 (Jean-Mairet et al.), U.S. Patent No. 6,602,684 (Umana et al.), and US 2005 / 0123546 (Umana et al.). Antibody variants having at least one galactose residue in the oligosaccharide attached to the Fc region are also provided. Such antibody variants may have improved CDC function. Such antibody variants are described, for example, in WO1997 / 30087 (Patel et al.), WO1998 / 58964 (Raju, S.), and WO1999 / 22764 (Raju, S.).

[0128] 7. Fc region variants In certain embodiments, one or more amino acid modifications can be introduced into the Fc region of an antibody provided herein, thereby generating an Fc region variant. The antibody can comprise a human Fc region sequence (eg, a human IgG1, IgG2, IgG3, or IgG4 Fc region) that comprises an amino acid modification (eg, a substitution) at one or more amino acid positions.

[0129] In certain embodiments, the present invention contemplates antibody variants that retain some, but not all, effector functions, making them desirable candidates for applications in which in vivo antibody half-life is important but certain effector functions (such as complement and ADCC) are unnecessary or deleterious. In vitro and / or in vivo cytotoxicity assays can be performed to confirm reduced / depleted CDC and / or ADCC activity. For example, Fc receptor (FcR) binding assays can be performed to ensure that the antibody lacks FcγR binding (and thus likely lacks ADCC activity) but retains FcRn binding ability. NK cells, the primary cells for mediating ADCC, express only FcγRIII, whereas monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol. 9:457-492 (1991). Non-limiting examples of in vitro assays to assess ADCC activity of a molecule of interest are described in U.S. Patent Nos. 5,500,362 (see, e.g., Hellstrom, I. et al., Proc. Nat'l Acad. Sci. USA 83:7059-7063 (1986) and Hellstrom, I. et al., Proc. Nat'l Acad. Sci. USA 82:1499-1502 (1985)), and 5,821,337 (see, Bruggemann, M. et al., J. Exp. Med. 166:1351-1361 (1987)). Alternatively, non-radioactive assay methods can be used (see, e.g., ACTI™ Non-Radioactive Cytotoxicity Assay for Flow Cytometry (CellTechnology, Inc. Mountain View, CA), and CytoTox 96® Non-Radioactive Cytotoxicity Assay (Promega, Madison, WI)). Useful effector cells for such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells.Alternatively, or additionally, ADCC activity of the molecule of interest can be assessed in vivo, e.g., in an animal model such as that disclosed in Clynes et al. Proc. Nat'l Acad. Sci. USA 95:652-656 (1998). C1q binding assays can also be performed to confirm that the antibody is unable to bind C1q and thus lacks CDC activity. See, e.g., C1q and C3c binding ELISAs in WO2006 / 029879 and WO2005 / 100402. To assess complement activation, a CDC assay can be performed (see, e.g., Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996); Cragg, MS et al., Blood 101:1045-1052 (2003); and Cragg, MS and MJ Glennie, Blood 103:2738-2743 (2004)). FcRn binding and in vivo clearance / half-life determinations can also be performed using methods known in the art (see, e.g., Petkova, SB et al., Int'l. Immunol. 18(12):1759-1769 (2006)).

[0130] Antibodies with reduced effector function include those with substitutions of one or more of Fc region residues 238, 265, 269, 270, 297, 327, and 329 (U.S. Patent No. 6,737,056). Such Fc variants include Fc variants with substitutions at two or more of amino acid positions 265, 269, 270, 297, and 327, including the so-called "DANA" Fc variant with substitutions of residues 265 and 297 to alanine (U.S. Patent No. 7,332,581).

[0131] Certain antibody variants with improved or diminished binding to FcRs have been described. (See, e.g., U.S. Pat. No. 6,737,056, WO 2004 / 056312, and Shields et al., J. Biol. Chem. 9(2):6591-6604 (2001).)

[0132] In certain embodiments, the antibody variant comprises an Fc region with one or more amino acid substitutions that improve ADCC, e.g., substitutions at positions 298, 333, and / or 334 (EU numbering of residues) of the Fc region.

[0133] In some embodiments, modifications that result in altered (i.e., either improved or decreased) C1q binding and / or complement dependent cytotoxicity (CDC) are made in the Fc region, e.g., as described in U.S. Pat. No. 6,194,551, WO 99 / 51642, and Idusogie et al. J. Immunol. 164:4178-4184 (2000).

[0134] Antibodies with increased half-lives and improved binding to neonatal Fc receptors (FcRn), which account for the transfer of maternal IgG to the fetus (Guyer et al., J. Immunol. 117:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)), are described in US2005 / 0014934A1 (Hinton et al.). These antibodies comprise an Fc region with one or more substitutions therein that improve binding of the Fc region to FcRn. Such Fc variants include those with substitutions at one or more of Fc region residues 238, 252, 254, 256, 265, 272, 286, 303, 305, 307, 311, 312, 317, 340, 356, 360, 362, 376, 378, 380, 382, ​​413, 424, or 434, e.g., substitution of Fc region residue 434 (U.S. Patent No. 7,371,826).

[0135] See also Duncan & Winter, Nature 322:738-40 (1988), U.S. Patent No. 5,648,260, U.S. Patent No. 5,624,821, and WO 94 / 29351 for other examples of Fc region variants.

[0136] In some embodiments, the antibody is provided according to the sequence listing and is of human IgG1 isotype. In some embodiments, the antibody is provided according to the sequence listing and is of human IgG4 isotype. In some embodiments, the antibody is provided according to the sequence listing and is of human IgG4 isotype with a single mutation at serine 228 to proline (S228P).

[0137] 8. Cysteine-engineered antibody variants In certain embodiments, it may be desirable to generate cysteine-engineered antibodies, e.g., "ThioM antibodies," in which one or more residues of an antibody are substituted with a cysteine ​​residue. In certain embodiments, the substituted residues occur at accessible sites on the antibody. By substituting those residues with cysteine, reactive thiol groups are thereby positioned at accessible sites on the antibody, which can be used to conjugate the antibody to other moieties, such as drug moieties or linker-drug moieties, to generate immunoconjugates, as further described herein. In certain embodiments, any one or more of the following residues can be substituted with cysteine: V205 (Kabat numbering) of the light chain, A118 (EU numbering) of the heavy chain, and S400 (EU numbering) of the heavy chain Fc region. Cysteine-engineered antibodies can be generated, for example, as described in U.S. Pat. No. 7,521,541.

[0138] 9. Antibody derivatives In certain embodiments, the antibodies provided herein may be further modified to contain additional nonproteinaceous moieties that are known in the art and readily available. Suitable moieties for derivatizing antibodies include, but are not limited to, water-soluble polymers. Non-limiting examples of water-soluble polymers include polyethylene glycol (PEG), ethylene glycol / propylene glycol copolymers, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymers, polyamino acids (either homopolymers or random copolymers), and dextran or poly(n-vinylpyrrolidone), polyethylene glycol, propylene glycol homopolymer, propylene oxide / ethylene oxide copolymer, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may have manufacturing advantages due to its stability in water. Polymers may be of any molecular weight and may be branched or unbranched. The number of polymers attached to the antibody may vary, and if two or more polymers are attached, they may be the same or different molecules. In general, the number and / or type of polymers used for derivatization can be determined based on considerations including, but not limited to, the particular property or function of the antibody to be improved, whether the antibody derivative will be used in therapy under defined conditions, etc.

[0139] In another embodiment, a conjugate of an antibody and a nonproteinaceous moiety is provided that can be selectively heated by exposure to radiation. In some embodiments, the nonproteinaceous moiety is a carbon nanotube (Kam et al., Proc. Natl. Acad. Sci. USA 102:11600-11605 (2005)). The radiation can be of any wavelength, including, but not limited to, wavelengths that do not harm normal cells but heat the nonproteinaceous moiety to temperatures that kill cells adjacent to the antibody-nonproteinaceous moiety.

[0140] B. Recombinant Methods Antibodies may be produced using recombinant methods and compositions, such as those described in U.S. Patent No. 4,816,567. In some embodiments, isolated nucleic acids encoding the anti-CD112R antibodies described herein are provided. Such nucleic acids may encode an amino acid sequence comprising the VL and / or an amino acid sequence comprising the VH of the antibody (e.g., the light and / or heavy chains of the antibody). In further embodiments, one or more vectors (e.g., expression vectors) comprising such nucleic acids are provided. In further embodiments, host cells comprising such nucleic acids are provided. In one such embodiment, the host cell comprises (e.g., is transformed with) (1) a vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and an amino acid sequence comprising the VH of the antibody, or (2) a first vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and a second vector comprising a nucleic acid encoding an amino acid sequence comprising the VH of the antibody. In some embodiments, the host cell is eukaryotic, such as a Chinese hamster ovary (CHO) cell or a lymphoid cell (e.g., a Y0, NS0, or Sp20 cell). In some embodiments, methods of making an anti-CD112R antibody are provided, the methods comprising culturing a host cell comprising nucleic acid encoding the antibody, as provided above, under conditions suitable for expression of the antibody, and optionally recovering the antibody from the host cell (or host cell culture medium).

[0141] For recombinant production of an anti-CD112R antibody, nucleic acid encoding the antibody is isolated, e.g., as described above, and inserted into one or more vectors for further cloning and / or expression in host cells. Such nucleic acid can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes capable of binding specifically to genes encoding the antibody heavy and light chains).

[0142] Suitable host cells for cloning or expressing antibody-encoding vectors are described herein. These include prokaryotic and eukaryotic cells. For example, antibodies can be produced in bacteria, particularly when glycosylation and Fc effector functions are not required. For expression of antibody fragments and polypeptides in bacteria, see, e.g., U.S. Patent Nos. 5,648,237, 5,789,199, and 5,840,523. (See also Charlton, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ, 2003), pp. 245-254, which describes the expression of antibody fragments in E. coli.) After expression, the antibody can be isolated from the bacterial cell paste in a soluble fraction and further purified.

[0143] In addition to prokaryotes, eukaryotic microbes such as filamentous fungi or yeast are suitable cloning or expression hosts for antibody-encoding vectors, including fungal and yeast strains that have been "humanized" in their glycosylation pathways, resulting in the production of antibodies with partially or fully human glycosylation patterns. See Gerngross, Nat. Biotech. 22:1409-1414 (2004), and Li et al., Nat. Biotech. 24:210-215 (2006).

[0144] Suitable host cells for the expression of glycosylated antibodies are also derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. A number of baculovirus strains have been identified that can be used in conjunction with insect cells, particularly for transfection of Spodoptera frugiperda cells.

[0145] Plant cell cultures may also be utilized as hosts. See, e.g., U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (which describe PLANTIBODIES™ technology for producing antibodies in transgenic plants).

[0146] Vertebrate cells can also be used as hosts. For example, mammalian cell lines adapted to grow in suspension can be useful. Other examples of useful mammalian host cell lines include the monkey kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney lines (e.g., 293 or 293 cells described in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (e.g., TM4 cells described in Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); canine kidney cells (MDCK); buffalo rat hepatocytes (BRL 3A); human lung cells (W138); human liver cells (Hep G2); mouse mammary tumor (MMT 060562); TRI cells, described in Mather et al., Annals NYAcad. Sci. 383:44-68 (1982); MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines are DHFR - These include Chinese hamster ovary (CHO) cells, including CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)), as well as myeloma cell lines such as Y0, NS0, and Sp2 / 0. For a review of specific mammalian host cell lines suitable for antibody production, see, e.g., Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0147] C. Immune complex The present invention also provides immunoconjugates comprising an anti-CD112R antibody herein conjugated to one or more other therapeutic agents or radioisotopes.

[0148] In another embodiment, the immunoconjugate comprises an antibody described herein conjugated to a radioactive atom to form a radioconjugate. A variety of radioisotopes are available for the production of radioconjugates. Examples include At 211 , I 131 , I 125 , Y 90 ,Re 186 ,Re 188 , Sm 153 , Bi 212 , P 32 , Pb 212 When radioconjugates are used for detection, they may contain radioactive atoms, such as tc99m or I123, for scintigraphic studies, or spin labels for nuclear magnetic resonance (NMR) imaging (also known as magnetic resonance imaging, mri), again such as iodine-123, iodine-131, indium-111, fluorine-19, carbon-13, nitrogen-15, oxygen-17, gadolinium, manganese, or iron.

[0149] Antibody conjugates can be made using a variety of bifunctional protein coupling agents, such as N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP), succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC), iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCl), active esters (such as disuccinimidyl suberate), aldehydes (such as glutaraldehyde), bis-azido compounds (such as bis(p-azidobenzoyl)hexanediamine), bis-diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (such as toluene 2,6-diisocyanate), and bis-active fluorine compounds (such as 1,5-difluoro-2,4-dinitrobenzene). For example, ricin immunotoxins are described in Vitetta et al., Science 238:1098 (1987). Carbon-14 labeled 1-isothiocyanatobenzyl-3-methyldiethylenetriaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugating radionucleotides to antibodies. See WO94 / 11026. The linker may be a "cleavable linker" that facilitates the release of the cytotoxic drug in cells. For example, an acid-labile linker, peptidase-sensitive linker, photolabile linker, dimethyl linker, or disulfide-containing linker (Chari et al., Cancer Res. 52:127-131 (1992); U.S. Pat. No. 5,208,020) may be used.

[0150] The immunoconjugates or ADCs herein expressly contemplate, but are not limited to, such conjugates prepared with crosslinker reagents, including, but not limited to, BMPS, EMCS, GMBS, HBVS, LC-SMCC, MBS, MPBH, SBAP, SIA, SIAB, SMCC, SMPB, SMPH, sulfo-EMCS, sulfo-GMBS, sulfo-KMUS, sulfo-MBS, sulfo-SIAB, sulfo-SMCC, and sulfo-SMPB, and commercially available SVSB (succinimidyl-(4-vinylsulfone)benzoate) (e.g., from Pierce Biotechnology, Inc., Rockford, IL, USA).

[0151] D. Pharmaceutical Formulations and Compositions Pharmaceutical formulations or compositions of the anti-CD112R antibodies described herein are prepared in the form of lyophilized formulations or aqueous solutions by mixing such antibodies having the desired degree of purity with one or more optional pharmaceutically acceptable carriers, diluents, and / or excipients (Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)). Pharmaceutically acceptable carriers, diluents, and excipients are generally non-toxic to recipients at the dosages and concentrations employed, and include buffers such as sterile water, phosphates, citrates, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzylammonium chloride, hexamethonium chloride, benzalkonium chloride, benzethonium chloride, phenol, butyl, or benzyl alcohol, methyl or propyl paraben, etc.). Examples of surfactants include, but are not limited to: alkylparabens, catechol, resorcinol, cyclohexanol, 3-pentanol, and m-cresol; low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes), and / or non-ionic surfactants such as polyethylene glycol (PEG). Exemplary pharmaceutically acceptable carriers herein further include interstitial drug dispersing agents, such as soluble neutral-active hyaluronidase glycoproteins (sHASEGPs), e.g., human soluble PH-20 hyaluronidase glycoproteins, such as rHuPH20 (HYLENEX®, Baxter International, Inc.). Certain exemplary sHASEGPs and methods of use, including rHuPH20, are described in U.S. Patent Publication Nos. 2005 / 0260186 and 2006 / 0104968. In one embodiment, a sHASEGP is combined with one or more additional glycosaminoglycanases, such as chondroitinases.

[0152] Exemplary lyophilized antibody formulations are described in U.S. Patent No. 6,267,958. Aqueous antibody formulations include those described in U.S. Patent No. 6,171,586 and WO2006 / 044908, the latter formulations including a histidine-acetate buffer.

[0153] The formulations herein may also contain more than one active ingredient as necessary for the particular indication being treated, preferably those with complementary activities that do not adversely affect each other. Such active ingredients are preferably present in combination in amounts that are effective for the purpose intended.

[0154] The active ingredient can be encapsulated, for example, in microcapsules prepared by coacervation techniques or by interfacial polymerization, e.g., hydroxymethylcellulose or gelatin-microcapsules and poly-(methyl methacrylate) microcapsules, in colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules) or in macroemulsions, respectively. Such techniques are described in Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980).

[0155] Sustained-release preparations can be prepared. Suitable examples of sustained-release preparations include semipermeable matrices of solid hydrophobic polymers containing the antibody, which matrices are in the form of shaped articles, e.g., films, or microcapsules.

[0156] Formulations or compositions to be used for in vivo administration are generally sterile, which may be readily accomplished, for example, by filtration through sterile filtration membranes.

[0157] E. Treatment method Any of the anti-CD112R antibodies provided herein can be used in therapeutic methods. It should also be understood that throughout, when "antibody" is discussed, compositions comprising the antibody are also encompassed.

[0158] In one aspect, an anti-CD112R antibody is provided for use as a pharmaceutical. In some embodiments, an anti-CD112R antibody is provided for use in enhancing, increasing, and / or maintaining an anti-tumor immune response in a subject having a tumor. In some embodiments, the tumor is cancerous. In some embodiments, the anti-CD112R antibody is provided for use in treating cancer. In some embodiments, anti-CD112R antibodies for use in enhancing CD226 interaction with CD112 are provided.

[0159] In a further aspect, the invention provides use of an anti-CD112R antibody in the manufacture or preparation of a medicament. In some embodiments, the medicament is for use in enhancing, increasing, and / or maintaining an anti-tumor immune response in a subject having a tumor. In some embodiments, the tumor is cancerous. In some embodiments, the medicament is for treating cancer. In some embodiments, the medicament is for enhancing CD226 interaction with CD112.

[0160] In a further aspect, the present invention provides methods for treating diseases and / or disorders in which blocking CD112R is desirable. In some embodiments, methods are provided for enhancing, increasing, and / or maintaining an anti-tumor immune response in a subject with a tumor, comprising administering an anti-CD112R antibody described herein. In some embodiments, the tumor is cancerous. In some embodiments, methods are provided for treating cancer in a subject with cancer, comprising administering an anti-CD112R antibody described herein. In some embodiments, methods are provided for enhancing CD226 interaction with CD112, optionally in a subject with cancer, comprising administering an anti-CD112R antibody described herein.

[0161] In some aspects, the present invention provides methods for alleviating one or more symptoms of a CD112R protein-associated disease or disorder, or medicaments comprising an anti-CD112R antibody for alleviating one or more symptoms of a CD112R protein-associated disease or disorder (such as any of the diseases or disorders described herein, e.g., cancer). In some aspects, the present invention provides methods for reducing the number of symptoms or the severity of one or more symptoms of a CD112R protein-associated disease or disorder, or medicaments comprising an anti-CD112R antibody for reducing the number of symptoms or the severity of one or more symptoms of a CD112R protein-associated disease or disorder (such as any of the diseases or disorders described herein, e.g., cancer). In certain embodiments, the symptom of a CD112R protein-associated disease or disorder is a tumor, and the reduction is a decrease in tumor size, failure of tumor growth, or elimination of the tumor.

[0162] The antibodies described herein can be used, for example, to treat cancer. In some embodiments, a method for treating cancer is provided, comprising administering an effective amount of an antibody described herein to a subject. In some embodiments, the antibody can induce or enhance an immune response in a subject, such as an antigen-specific immune response. In some embodiments, the antibody can stimulate T cell activity. In some embodiments, the antibody can inhibit the growth of at least one tumor in a subject.

[0163] Provided herein is a method for treating a subject with cancer, comprising administering to the subject a therapeutically effective amount of a CD112R antibody described herein, so that the subject is treated.The CD112R antibody can be used alone.Alternatively, the CD112R antibody can be used in conjunction with another agent, as further described below.

[0164] The cancer can be a cancer involving a solid tumor or a hematological malignancy (e.g., a liquid tumor).

[0165] Non-limiting examples of cancers for treatment include squamous cell carcinoma, small cell lung cancer, non-small cell lung cancer, squamous non-small cell lung cancer (NSCLC), non-squamous NSCLC, glioma, gastrointestinal cancer, renal cancer (e.g., clear cell carcinoma), ovarian cancer, liver cancer, colon cancer, endometrial cancer, renal cancer (e.g., renal clear cell carcinoma), and ovarian cancer. Cancer (RCC)), prostate cancer (e.g., hormone-refractory prostate cancer), thyroid cancer, neuroblastoma, pancreatic cancer, glioblastoma (e.g., glioblastoma multiforme), cervical cancer, stomach cancer, bladder cancer, hepatocellular carcinoma, breast cancer, colon carcinoma, and head and neck cancer (or carcinoma), gastric cancer, germ cell tumors, pediatric sarcoma, sinonasal natural killer, melanoma (e.g., metastatic melanoma such as cutaneous melanoma or intraocular melanoma), bone cancer, skin cancer, uterine cancer, anal cancer, testicular cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, esophageal cancer, small intestine cancer, endocrine system cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, pediatric solid tumors, ureteral cancer, renal pelvis cancer, central nervous system ( CNS neoplasms, primary CNS lymphomas, tumor angiogenesis, spinal axis tumors, brain cancers, brain stem gliomas, pituitary adenomas, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, environmentally induced cancers including those induced by asbestos, virus-associated cancers or cancers of viral origin (e.g., tumors related to or derived from human papillomavirus (HPV)), and hematological malignancies derived from the two major blood cell lineages, i.e., myeloid cell lineages (producing granulocytes, erythrocytes, platelets, macrophages, and mast cells) or lymphoid cell lineages (producing B, T, NK, and plasma cells), such as all types of leukemia, lymphoma, and myeloma, e.g., acute myeloid leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML), anaplastic AML (AML), acute, chronic, lymphocytic, and / or myeloid leukemias, such as MO), myeloblastic leukemia (M1), myeloblastic leukemia (M2, with cellular maturation), promyelocytic leukemia (M3 or M3 variant [M3V]), myelomonocytic leukemia (M4 or M4 variant [M4E] with eosinophilia), monocytic leukemia (M5), erythroleukemia (M6), megakaryoblastic leukemia (M7), isolated granulocytic sarcoma, and chloroma;Hodgkin's lymphoma (HL), non-Hodgkin's lymphoma (NHL), B-cell hematologic malignancies, e.g., B-cell lymphoma, T-cell lymphoma, lymphoplasmacytic lymphoma, monocytic B-cell lymphoma, mucosa-associated lymphoid tissue (MALT) lymphoma, anaplastic (e.g., Ki 1+) large cell lymphoma, adult T-cell lymphoma / leukemia, mantle cell lymphoma, hematoimmunoblastic T-cell lymphoma, angiocentric lymphoma, intestinal T-cell lymphoma, primary mediastinal B-cell lymphoma, precursor T-lymphoblastic lymphoma, T-lymphoblastic lymphoma; as well as lymphoma / leukemia (T-Lbly / T-ALL), peripheral T-cell lymphoma, lymphoblastic lymphoma, post-transplant lymphoproliferative disorder, true lymphoma Histocytic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, B-cell lymphoma, lymphoblastic lymphoma (LBL), lymphoid hematopoietic tumors, acute lymphoblastic leukemia, diffuse large B-cell lymphoma, Burkitt lymphoma, follicular lymphoma, diffuse histiocytic lymphoma (DHL), immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma, cutaneous T-cell lymphoma (CTLC) ( mycosis fungoides or Sézary syndrome), and lymphoplasmacytic lymphoma (LPL) with Waldenstrom's macroglobulinemia; myelomas such as IgG myeloma, light chain myeloma, non-secretory myeloma, smoldering myeloma (also called indolent myeloma), solitary plasmacytoma, and multiple myeloma, chronic lymphocytic leukemia (CLL), hairy cell lymphoma; hematopoietic tumors of myeloid lineage, tumors of mesenchymal origin including fibrosarcoma and rhabdomyosarcoma; and other tumors including melanoma, xeroderma pigmentosum, keratoacanthoma, seminoma, thyroid follicular carcinoma and teratoma, hematopoietic tumors of lymphoid lineage, e.g., T-cell and B-cell tumors including, but not limited to, T-cell cytotoxicity such as T-prolymphocytic leukemia (T-PLL), including those of the small cell and gyrus cell types; large granular lymphocytic leukemia (LGL) of the T-cell type; T-NHL hepatosplenic lymphoma; peripheral / mature T-cell lymphoma (pleomorphic and immunoblastic subtypes); angiocentric (nasal) T-cell lymphoma;head and neck cancer, acute myeloid lymphoma, as well as any combination of the foregoing cancers. The methods described herein may also be used to treat metastatic cancer, unresectable, refractory cancer (e.g., cancer refractory to previous immunotherapy, e.g., with blocking CTLA-4 or PD-1 antibodies), and / or recurrent cancer;

[0166] In certain embodiments, the antibodies described herein are administered to subjects with cancer that has had an inadequate response to prior treatment, e.g., prior treatment with an immuno-oncology or immunotherapeutic agent, or that has progressed during prior treatment with an immuno-oncology or immunotherapeutic agent. In some embodiments, the cancer is refractory or resistant to previous treatment, either inherently refractory or resistant (e.g., refractory to a PD-1 pathway antagonist), or the resistant or refractory state is acquired. For example, the antibodies described herein can be administered to subjects who do not respond or do not adequately respond to a first therapy, or who have disease progression after treatment, e.g., anti-PD-1 pathway antagonist treatment, alone or in combination with another therapy (e.g., with anti-PD-1 pathway antagonist therapy). In other embodiments, the antibodies described herein are administered to subjects who have not previously received (i.e., been treated with) an immuno-oncology agent, e.g., a PD-1 pathway antagonist.

[0167] F. Combination The antibodies of the invention can be used alone or in combination with other agents in a therapy, for example, the antibodies of the invention can be co-administered with at least one additional therapeutic agent (e.g., further comprising administering a second therapy).

[0168] In some embodiments, targeting additional independent inhibitory pathways or combinations thereof has the potential to result in further enhanced immune cell activation beyond monotherapy.

[0169] In some embodiments, the additional therapeutic agent or second agent is a chemotherapeutic agent, an opsonizing agent, a regulatory T cell ("Treg") depleting agent, an antagonist of a target other than CD112R, or an agonist of a target other than CD112R. In certain embodiments, the second agent is a chemotherapeutic agent described herein or any known chemotherapeutic agent. In some embodiments, the second agent is an opsonizing agent, and the opsonizing agent is an antibody other than an anti-CD112R antibody that targets cancer or tumor cells. In some embodiments, the second agent is a Treg depleting agent described herein or any known Treg depleting agent. In some embodiments, the second agent is an antagonist of a target other than CD112R. In some embodiments, the second agent is an agonist of a target other than CD112R.

[0170] In some cases, the second agent targets an independent inhibitory pathway, such as, for example, a pathway involving PD-1, PD-L1, CTLA-4, Lag-3, or TIM-3. In some embodiments, the second agent antagonizes one or more of PD-1, PD-L1, CTLA-4, Lag-3, and TIM-3. Antagonists suitable for use in the combination therapies described herein include, but are not limited to, ligands, antibodies (e.g., monoclonal antibodies and bispecific antibodies), and multivalent agents. In one embodiment, the antagonist is a fusion protein, e.g., an Fc fusion protein such as AMP-244. In some embodiments, the PD-1 antagonist is an anti-PD-1 or anti-PD-L1 antibody.

[0171] An exemplary anti-PD-1 antibody is nivolumab (BMS-936558) or an antibody comprising the CDRs or variable regions of one of antibodies 17D8, 2D3, 4H1, 5C4, 7D3, 5F4, and 4A11 described in WO2006 / 121168. In certain embodiments, the anti-PD-1 antibody is MK-3475 (lambrolizumab), described in WO2012 / 145493, AMP-514, described in WO2012 / 145493, or PDR001. Additional known PD-1 antibodies and other PD-1 inhibitors include those described in WO2009 / 014708, WO03 / 099196, WO2009 / 114335, WO2011 / 066389, WO2011 / 161699, WO2012 / 145493, U.S. Patent Nos. 7,635,757 and 8,217,149, and U.S. Patent Publication No. 2009 / 0317368. Antibodies disclosed in WO2013 / 173223 Any of the PD-1 antibodies can also be used, as can anti-PD-1 antibodies that compete for binding to the same epitope on PD-1 and / or bind to the same epitope on PD-1, such that one of these antibodies can also be used in combination therapy.

[0172] In some embodiments, the anti-PD-L1 antibody useful in combination therapy is BMS-936559 (referred to as 12A4 in WO2007 / 005874 and US Patent No. 7,943,743), or an antibody that comprises the CDRs or variable regions of 3G10, 12A4, 10A5, 5F8, 10H10, 1B12, 7H1, 11E6, 12B7, and 13G4, which are described in PCT Publication No. WO07 / 005874 and US Patent No. 7,943,743. In certain embodiments, the anti-PD-L1 antibody is MEDI4736 (also known as durvalumab and anti-B7-H1), MPDL3280A (also known as atezolizumab and RG7446), MSB0010718C (also known as avelumab, WO2013 / 79174), or rHigM12B7. Any of the anti-PD-L1 antibodies disclosed in WO2013 / 173223, WO2011 / 066389, WO2012 / 145493, U.S. Patent Nos. 7,635,757 and 8,217,149, and U.S. Publication No. 2009 / 145493 may also be used. Anti-PD-L1 antibodies that compete with and / or bind to the same epitope as any of these antibodies may also be used in combination therapy.

[0173] In certain embodiments, the CD112R antibodies of the present disclosure may be used in conjunction with a CTLA-4 antagonist, for example, an anti-CTLA-4 antibody. In one embodiment, the anti-CTLA-4 antibody is an antibody selected from the group of Yervoy® (ipilimumab or antibody 10D1, described in PCT Publication No. WO 01 / 14424), tremelimumab (formerly ticilimumab, CP-675,206), monoclonal or anti-CTLA-4 antibodies described in any of the following publications: WO 98 / 42752, WO 00 / 37504, U.S. Patent No. 6,207,156, Hurwitz et al. (1998) Pro. Natl. Acad. Sci. USA 95(17):10067-10071, Camacho et al. (2004) J. Clin. Oncology 22(145):antibodiestract No. 2505 (antibody CP-675206), and Mokyr et al. (1998) Cancer Res. 58:5301-5304. Any of the anti-CTLA-4 antibodies disclosed in WO2013 / 173223 can also be used.

[0174] In some embodiments, the CD112R antibodies of the present disclosure are used in combination with a LAG-3 (also referred to herein and elsewhere as LAG3) antagonist. Examples of anti-LAG3 antibodies include antibodies comprising the CDRs or variable regions of antibodies 25F7, 26H10, 25E3, 8B7, 11F2, or 17E5, and are described in U.S. Patent Publication Nos. US2011 / 0150892, WO10 / 19570, and WO2014 / 008218. In one embodiment, the anti-LAG-3 antibody is BMS-986016. Other art-recognized anti-LAG-3 antibodies that can be used include IMP731 and IMP-321, described in US2011 / 007023, WO08 / 132601, and WO09 / 44273. Anti-LAG-3 antibodies that compete with and / or bind to the same epitope as any of these antibodies may also be used in combination therapy.

[0175] In some embodiments, targeting two or more of the TIGIT, CD96, and CD112R receptors simultaneously increases CD226-mediated signaling beyond anti-CD112R monotherapy. Thus, in some embodiments, the second agent is an antagonist of TIGIT and / or CD96. Antagonists suitable for use in the combination therapies described herein include ligands, antibodies (e.g., monoclonal antibodies and bispecific antibodies), and the like. These include, but are not limited to, monovalent antibodies, and polyvalent agents.

[0176] In some embodiments, members of the PVR gene family are upregulated on tumor cells and can exhibit intrinsic tumor-promoting properties. Targeting additional members of the PVR gene family in combination with an anti-CD112R antibody results in enhanced tumor susceptibility beyond monotherapy. Thus, in some embodiments, the second agent is selected from one or more of antagonists of PVRL1, PVRL2, PVRL3, PVRL4, and CD155. Antagonists suitable for use in the combination therapies described herein include, but are not limited to, ligands, antibodies (e.g., monoclonal antibodies and bispecific antibodies), and multivalent agents.

[0177] STING agonists induce innate immune cell activation, increasing T cell priming and immune cell recruitment to the tumor microenvironment. Targeting STING agonists in combination with CD112R has the potential to further increase T cell and NK cell recruitment and activation.

[0178] Increased phagocytosis via anti-CD47 antibodies can lead to increased presentation of cancer-derived antigens by macrophages to T cells. Combination treatment with anti-CD47 and anti-CD112R antibodies, such as the anti-CD112R antibodies provided herein, offers the opportunity to enhance cancer antigen-specific T cell responses and is fully encompassed herein.

[0179] Adenosine inhibits the activation of T cells and NK cells via adenosine receptors expressed on immune cells. Anti-CD39 antibodies inhibit the production of adenosine by preventing the hydrolysis of adenosine triphosphate (ATP). Combination treatment with anti-CD39 and anti-CD112R antibodies, such as the anti-CD112R antibodies provided herein, provides an opportunity to further enhance CD112R therapy by inhibiting adenosine-mediated cell signaling in immune cells.

[0180] Cytokines can effectively regulate the activation of T cells and NK cells. IL-27 is an immunosuppressive cytokine that inhibits T cell- and NK cell-mediated responses. Anti-IL-27 antibodies provide an opportunity to enhance CD112R therapy by limiting the signaling of immunosuppressive cytokines in immune cells. Therefore, combination treatment with an anti-IL-27 antibody and an anti-CD112R antibody, such as the anti-CD112R antibody provided herein, is provided.

[0181] The antibodies herein can also be provided prior to, substantially simultaneously with, or following the administration of other therapeutic modalities, e.g., surgery, chemotherapy, radiation therapy, or a biologic such as another therapeutic antibody. In some embodiments, the cancer has recurred or progressed after therapy selected from surgery, chemotherapy, and radiation therapy, or a combination thereof. For example, the CD112R antibodies described herein can be administered as adjuvant therapy when there is a risk that micrometastases may be present and / or to reduce the risk of recurrence.

[0182] For the treatment of cancer, the combination may be administered in conjunction with one or more additional anti-cancer agents, such as chemotherapeutic agents, growth inhibitory agents, anti-cancer vaccines such as gene therapy vaccines, anti-angiogenic agents, and / or anti-neoplastic compositions.

[0183] In some embodiments, the anti-inflammatory agent may be administered in combination with a steroid or a nonsteroidal anti-inflammatory drug (NSAID), such as 17a-ethynylestradiol, diethylstilbestrol, testosterone, or steroids, in cases where it is desirable to render the abnormally proliferative cells quiescent, in conjunction with or prior to treatment with a CD112R antibody described herein. Hormones and steroids (including synthetic analogs) such as prednisone, fluoxymesterone, dromostanolone propionate, testolactone, megestrol acetate, methylprednisolone, methyl-testosterone, prednisolone, triamcinolone, chlorotrianisene, hydroxyprogesterone, aminoglutethimide, estramustine, medroxyprogesterone acetate, leuprolide, flutamide, toremifene, ZOLADEX®, and the like, may also be administered to the subject. When using the methods or compositions described herein, other agents used to regulate tumor growth or metastasis in clinical settings, such as antimimetic drugs, may also be administered as needed.

[0184] Such combination therapy, as described above, encompasses combined administration (where two or more therapeutic agents are included in the same or separate formulations or compositions) and separate administration, where administration of an antibody of the invention can occur prior to, concurrently with, and / or after administration of the additional therapeutic agent or agents. In some embodiments, administration of the anti-CD112R antibody and administration of the additional therapeutic agent occur within about one month, or within about one, two, or three weeks, or within about one, two, three, four, five, or six days of each other.

[0185] The antibodies of the invention (and any additional therapeutic agents) can be administered by any suitable means, including parenteral, intrapulmonary, and intranasal, and, if desired for localized treatment, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Dosing can be by any suitable route, for example, injections such as intravenous or subcutaneous injections, depending in part on whether the administration is brief or chronic. Various dosing schedules are contemplated herein, including, but not limited to, single or multiple administrations over various time points, bolus administration, and pulse infusions.

[0186] The antibodies of the present invention can be formulated, dosed, and administered in a manner consistent with good medical practice. Factors to consider in this context include the particular disorder being treated, the particular mammal being treated, the clinical condition of the individual subject, the cause of the disorder, the site of drug delivery, the method of administration, the schedule of administration, and other factors known to medical professionals. As used herein, a "split dose" refers to the division of a single unit dose or total daily dose into two or more doses, for example, two or more administrations of a single unit dose. The antibody can be administered as a "split dose."

[0187] The antibody is optionally, but need not be, formulated with one or more agents currently used to prevent or treat the disorder in question. Effective amounts of such other agents depend on the amount of antibody present in the formulation or composition, the type of disorder or treatment, and other factors discussed above. These are generally used in the same dosages and by the routes of administration described herein, or about 1-99% of the dosages described herein, or any dosage and by any route empirically / clinically determined to be appropriate. In some embodiments, the antibody is provided in a formulation for immediate release and the other agent is formulated for sustained release, or vice versa.

[0188] G.Manufactured products In another aspect of the present invention, an article of manufacture is provided containing materials useful for the treatment, prevention, and / or diagnosis of the above-mentioned disorders. The article of manufacture comprises a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, IV solution bags, and the like. The container can be formed from a variety of materials, such as glass or plastic. The container holds a composition, by itself or in combination with another composition, that is effective in the treatment, prevention, and / or diagnosis of a condition, and can have a sterile access port (e.g., the container can be an intravenous solution bag or a vial with a stopper pierceable by a hypodermic needle). At least one active agent in the composition is an antibody of the present invention. The label or package insert indicates that the composition is used to treat a selected condition. Furthermore, the article of manufacture may include (a) a first container containing a composition comprising an antibody of the invention, and (b) a second container containing a composition comprising an additional cytotoxic or otherwise therapeutic agent. The article of manufacture of this embodiment of the invention may further include a package insert indicating that the composition can be used to treat a particular condition. Alternatively, or additionally, the article of manufacture may further include a second (or third) container containing a pharmaceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer's solution, and dextrose solution. It may further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles, and syringes.

[0189] It will be understood that any of the above articles of manufacture may comprise an immunoconjugate of the present invention in place of, or in addition to, an anti-CD112R antibody. [Example]

[0190] III. Working Examples Example 1. Anti-CD112R antibody generation Antibodies were biotinylated using the EZ-Link sulfo-NHS-biotinylation kit from Pierce. Goat F(ab')2 anti-human kappa-FITC (LC-FITC), ExtrAvidin-PE (EA-PE), and streptavidin-AF633 (SA-633) were obtained from Southern Biotech, Sigma, and Molecular Probes, respectively. Streptavidin MicroBeads and MACS LC separation columns were purchased from Miltenyi Biotec. Goat anti-human IgG-PE (human-PE) was obtained from Southern Biotech.

[0191] Major findings. Eight naive human synthetic yeast libraries with a diversity of approximately 10 were each grown as previously described (see, e.g., Y. Xu et al., Addressing polyspecificity of antibodies selected from an in vitro yeast presentation system: a FACS-based, high-throughput selection and analytical tool. PEDS 26.10, 663-70 (2013); WO2009036379; WO2010105256; and WO2012009568). For the first two rounds of selection, magnetic bead sorting was performed using the Miltenyi MACS system as previously described (see, e.g., Siegel et al., High efficiency recovery and epitope-specific sorting of an scFv yeast display library. J Immunol Methods 286(1-2), 141-153 (2004). Briefly, yeast cells (approximately 10 cells / library) were incubated with 1.5 ml of 10 nM biotinylated Fc fusion antigen in wash buffer (phosphate-buffered saline (PBS) / 0.1% bovine serum albumin (BSA)) for 15 minutes at 30°C. After washing once with 40 ml of ice-cold wash buffer, the cell pellet was resuspended in 20 ml of wash buffer, and streptavidin MicroBeads (500 μl) were added to the yeast and incubated for 15 minutes at 4°C. The yeast were then pelleted, resuspended in 20 ml of wash buffer, and loaded onto a Miltenyi LS column. After loading 20 ml, the column was washed three times with 3 ml of wash buffer. The column was then removed from the magnetic field and the yeast were eluted with 5 mL of growth medium and then grown overnight. The following rounds of selection were performed using flow cytometry: approximately 2 x 10 7 Yeast cells are pelleted, washed three times with wash buffer, and incubated under equilibration conditions with decreasing concentrations of biotinylated antigen (100 to 1 nM) or with nonspecific antibodies to remove nonspecific antibodies from the selection. The libraries were incubated at 30°C with either a polyspecific depletion reagent (PSR) or a biotinylated PSR reagent. For PSR depletion, the libraries were incubated with a 1:10 dilution of biotinylated PSR reagent as previously described (see, e.g., Y. Xu et al., Addressing polyspecificity of antibodies selected from an in vitro yeast presentation system: a FACS-based, high-throughput selection and (See analytical tool. PEDS 26.10, 663-70 (2013).) The yeast were then washed twice with wash buffer and stained with LC-FITC (diluted 1:100) and either SA-633 (diluted 1:500) or EAPE (diluted 1:50) as a secondary reagent for 15 minutes at 4°C. After washing twice with wash buffer, the cell pellet was resuspended in 0.3 mL wash buffer and transferred to a sort tube with a strainer cap. Selection using affinity pressure to select and isolate higher affinity antibodies was performed by competing with cold (i.e., unlabeled) antigen.

[0192] Sorting was performed using a FACS ARIA sorter (BD Biosciences), and sort gates were determined to select for antibodies with the desired characteristics. Selection rounds were repeated until a population with all of the desired characteristics was obtained. After the final round of sorting, yeast were plated, and individual colonies were selected for characterization.

[0193] Light Chain Batch Shuffle. A light chain diversification protocol was used during the primary discovery phase for further discovery and improvement of the antibody.

[0194] Light chain batch diversification protocol: Heavy chains from the naive selection output were extracted from yeast via PCR and 5 x 10 6The resulting light chain library was transformed with a diversity of 10 ...

[0195] Antibody Optimization Antibody optimization was carried out by introducing diversity into the heavy chain variable region as described below.

[0196] CDRH1 and CDRH2 selection: CDRH3 of a single antibody was selected from 1 × 10 8 The CDRH1 and CDRH2 variants were recombined into a ready-made library with a diversity of 1000, ...

[0197] Antibody production and purification Yeast clones were grown to saturation and then induced for 48 hours at 30°C with shaking. After induction, yeast cells were pelleted and the supernatant was collected for purification. IgG was purified using a Protein A column and eluted with acetic acid, pH 2.0. Fab fragments were generated by papain digestion and purified on KappaSelect (GE Healthcare LifeSciences).

[0198] ForteBio KD Measurement ForteBio affinity measurements were generally performed on Octet RED384 as previously described (see, e.g., Estep et al., High throughput solution-based measurement of antibody-antigen affinity and epitope binning. Mabs 5(2), 270-278 (2013)). Briefly, ForteBio affinity measurements were performed by loading IgG onto the AHQ sensor online. The sensor was equilibrated offline in assay buffer for 30 minutes and then monitored online for 60 seconds to establish a baseline. The IgG-loaded sensor was exposed to 100 nM antigen for 3 minutes, then transferred to assay buffer for 3 minutes for off-rate measurements. All kinetics were analyzed using a 1:1 binding model.

[0199] ForteBio epitope binning / ligand blocking Epitope binning / ligand blocking was performed using a standard sandwich-format cross-blocking assay. Control anti-target IgG or receptor was loaded onto the AHQ sensor, and unoccupied Fc binding sites on the sensor were blocked with an irrelevant (non-target) human IgG1 antibody. The sensor was then exposed to 100 nM of target antigen followed by a second anti-target antibody. Additional binding by the secondary antibody or ligand after antigen association indicates an unoccupied epitope (non-competitor), while a lack of binding indicates epitope blocking (competitor or ligand blocking).

[0200] Biacore kinetic assay For Biacore-based measurements, antigen was covalently coupled to an anti-mouse-Fc capture C1 chip using an amine coupling kit (GE Healthcare Bio-Sciences). The association of antigen with a five-point, three-fold titration of antibody starting at 27 nM was measured for 300 seconds. The dissociation between antigen and antibody was then measured for 3600 seconds. Kinetic data were analyzed and globally fitted using a 1:1 binding model.

[0201] Example 2. Anti-CD112R antibodies bind to CD112R. About cell binding assays The ability of anti-CD112R antibodies to bind to CD112R expressed on cells was assessed. 1 x 10 wild-type or engineered to overexpress human CD112R (Jurkat-CD112R OE) cells were used. 5 Jurkat cells (acute T-cell leukemia cell line, ATCC No. TIB-152) were added to each well of a 96-well V-bottom plate and stained with either anti-CD112R antibody or IgG1 isotype control (0.63 μg / mL) for 1 hour at 4°C. Cells were washed twice with PBS + 2% FCS, resuspended in Alexa Fluor® 647 anti-human IgG Fc antibody (Biolegend, catalog no. 409320) diluted 1:100 in PBS + 2% FCS, and incubated for 30 minutes at 4°C. Subsequently, cells were washed twice and resuspended in PBS + 2% FCS. Cell data were acquired using an LSRFortessa X-20 (BD Biosciences) and analyzed with FlowJo software (Tree Star).

[0202] The results are shown in Figure 1. Quantitation of antibody binding to Jurkat-CD112R OE cells was assessed by the geometric mean fluorescence intensity (gMFI) of the Alexa Fluor® 647 signal. These results demonstrate that anti-CD112R antibodies bind to cells expressing CD112R. A summary of antibody binding is shown in Table 2.

[0203] Example 3. Anti-CD112R antibodies block CD112 binding to CD112R-expressing cells. About the cell blocking assay The ability of anti-CD112R antibodies to block CD112 binding to CD112R-expressing cells was evaluated. 1 × 10 cells engineered to overexpress human CD112R (Jurkat-CD112R OE) were used. 5Jurkat cells (acute T-cell leukemia cell line, ATCC No. TIB-152) were added to each well of a 96-well V-bottom plate and stained with serial dilutions of either anti-CD112R antibody or an IgG1 isotype control (highest concentration, 10 μg / mL) for 1 hour at 4°C. The cells were washed twice with PBS + 2% FCS and resuspended in PBS + 2% FCS with biotinylated his-tagged human CD112 (3 μg / mL) (BD Bioscience, No. 71234) and PE-streptavidin (5 μg / mL) (Biolegend, No. 405204) for 2 hours at 4°C. The cells were then washed twice and resuspended in PBS + 2% FCS. Cell data were acquired using an LSRFortessa X-20 (BD Biosciences) and analyzed with FlowJo software (Tree Star).

[0204] The results are shown in Figure 2A and Table 2. Quantitation of CD112 binding to cells was assessed by the geometric mean fluorescence intensity (gMFI) of the PE signal and expressed as percent inhibition. Percent inhibition was calculated as [100-((test sample MFI / maximum MFI)*100%)]. These results demonstrate that anti-CD112R antibodies inhibit the ability of CD112 to bind to CD112R-expressing cells in a dose-dependent manner. A summary of antibody binding and blocking is shown in Table 2.

[0205] The ability of anti-CD112R antibodies to block human CD112R was also evaluated by ELISA. Briefly, 96-well Nunc Maxisorp plates were coated overnight at 4°C with 1 μg / mL of CD112R-hIgG4 fusion protein in PBS. The plates were then washed six times with PBS + 0.01% Tween-20 (PBST) and subsequently blocked with 200 μl of PBS + 1% BSA at room temperature for 1.5 hours. After blocking, the plates were washed six times with PBST. Next, 100 μl of anti-CD112R antibody in PBS + 1% BSA was added using a four-fold serial dilution at a final starting concentration of 10 μg / mL. The plates were incubated at room temperature for 1.5 hours. The plate was then washed six times with PBST and then incubated with 1 μg / ml CD112 Fc protein (R&D Systems, Cat. No. 9317-N2-050) biotinylated with a sulfo-NHS biotinylation kit (Thermo Fisher, Cat. No. 21925) in 100 μl of PBS + 1% BSA for 1 hour at room temperature. The plate was then washed six times with PBST, followed by incubation with streptavidin-HRP (Biolegend, Cat. No. 405210) diluted in PBS + 1% BSA according to the manufacturer's recommendations for 1 hour at room temperature. After this incubation, the plate was then washed six times with PBST and developed with TMB substrate (Life Technologies, Cat. No. 002023). The reaction was stopped with an equal volume of stop solution (Life Technologies, Cat. No. SS04). The absorbance at 450 nm (OD450) was measured on a SpectraMax plate reader.

[0206] The results are shown in Figure 2B and Table 2. These results demonstrate that anti-CD112R antibodies block human CD112R from binding to human CD112. Percent inhibition was calculated as [100-((test sample OD450 / maximum OD450)*100%)]. Maximum OD450 was defined as the absorbance at 450 nm in the absence of antibody. [Table 2]

[0207] Example 4. Anti-CD112R antibodies enhance NK cell-mediated killing. NK cytotoxicity assay To determine the effect of anti-human CD112R antibodies on NK cell-mediated cytotoxicity, human NK cells were cocultured with REH target cells (a non-T / B cell acute lymphocytic leukemia cell line, ATCC number CRL-8286) in the presence of anti-CD112R antibodies or an isotype control.

[0208] Briefly, NK cells were isolated from PBMCs of three healthy donors via negative selection (Easysep™ NK Cell Isolation Kit, Stem Cell #17955) and activated with IL-2 (10 units / mL) (Peprotech #200-02) and IL-12 (20 ng / mL (Peprotech #200-12)) in RPMI + 10% FBS + 1% penicillin-streptomycin (R10) (ThermoFisher) for 16 hours. REH cells were washed, resuspended in PBS (ThermoFisher), and labeled with CellTrace™ Violet (CTV) (ThermoFisher #C34557) for 12 minutes at 37°C. Subsequently, REH cells were washed with PBS + 10% FBS and then resuspended in R10. After activation, NK cells were washed and resuspended in R10. 2.5 x 10 5 NK cells and 5 x 10 4 REH cells were added to each well of a 96-well flat-bottom plate at an effector-target cell ratio of 5:1. Anti-CD112R and IgG1 isotype antibodies were diluted in R10 and added to each well at a final concentration of 10 μg / mL. Each condition was performed in duplicate. The plates were then incubated at 37°C for 4 hours. The cells were then washed and incubated in the dark at room temperature for 30 minutes with 7-AAD viability dye (1 μg / mL) (Biolegend #420404) to specifically label dead cells. Cell viability data were analyzed using an LSRFortessa X-ray microscope. The images were acquired using a microscope (BD Biosciences) and analyzed with FlowJo software (Tree Star). Dead REH cells were defined as CTV and 7-AAD double-positive cells.

[0209] The results are presented in Figure 3. Cytotoxicity (percent above isotype) was calculated as ((test mortality - isotype mortality) / isotype mortality) x 100. Treatment of NK cells with each of the anti-CD112R antibodies described herein resulted in increased cell-mediated cytotoxicity against REH cells compared to the isotype control. These results demonstrate that anti-CD112R antibodies promote NK cell-mediated killing.

[0210] Example 5. Anti-CD112R antibodies enhance antigen-driven activation of CD8+ T cells. Antigen-specific CD8+ T cell assay. The effect of anti-CD112R antibody on antigen-driven activation of CD8+ T cells was evaluated. A primary HLA-A*0201-restricted cytomegalovirus (CMV)-specific CD8+ T cell line (Astarte Biologics #1049, Lot #3782DE17) was incubated with peptide-pulsed Colo205 cells (colon adenocarcinoma cell line, ATCC #CCL-222) in the presence of anti-CD112R antibody or isotype control.

[0211] Briefly, CMV-specific T cells were thawed, washed, and resuspended in X-VIVO 10 (ThermoFisher no. BW04380Q). 4 CMV T cells were added to each well of a 96-well round-bottom plate and incubated for 4 hours at 37°C. After the initial incubation period, 5 x 10 4Colo205 cells and CMV pp65 peptide (1 ng / mL) (Anaspec #AS-63937) were added to each well. Anti-CD112R and isotype antibodies were then diluted in X-VIVO 10 and added to each well at a final concentration of 10 μg / mL. Each condition was run in duplicate. Plates were then incubated at 37°C for 16 hours. Supernatants from each well were harvested and subjected to one freeze / thaw cycle before cytokine evaluation. After thawing, assay supernatants were diluted 1:5 in X-VIVO 10, and interferon gamma (IFNg) was then measured in the assay supernatants using a Luminex Human CD8+ T Cell Magnetic Bead Panel Multiplex Assay (Millipore Sigma #HCD8MAG-15K). It was run on FlexMap 3D.

[0212] The results are presented in Figure 4. IFNg levels under the test conditions were quantified based on a standard curve generated at defined IFNg concentrations. CD8+ T cells treated with anti-CD112R antibodies 2 and 5 resulted in greater IFNg secretion than observed with the isotype control. These results demonstrate that anti-CD112R antibodies enhance antigen-driven CD8+ T cell activation.

[0213] Example 6. The combination of anti-mouse CD112R and anti-mouse TIGIT antibodies has therapeutic effect in the mouse CT-26 tumor model. In vivo efficacy of combined CD112R and Tigit blockade The efficacy of CD112R and TIGIT blockade as single agents and in combination was tested in a CT26 colon adenocarcinoma syngeneic mouse tumor model. Seven-week-old Balb / c female mice (Charles River Laboratories, #028) were inoculated with 0.1 × 10 6 CT26.WT cells (ATCC No. CRL-2638) were implanted subcutaneously in 0.1 mL of 50% Matrigel seeding matrix. Mice were placed between 80 and 120 mm 3The mice were randomized into groups of 10 mice each (40 mice total) in a stratified manner by tumor volume range and treated by intraperitoneal injection twice a week for 2 weeks as shown in Table 3. [Table 3]

[0214] Tumors were within the IACUC limit size (<2000 mm 3 Tumor volumes were measured every 2-3 days until tumor volume reached 1000 mg / kg. [Table 4]

[0215] The results are presented in Figure 5. The graph shows the mean tumor volume for each treatment group as a function of time. The results demonstrate that the combination of anti-CD112R with anti-TIGIT was effective in reducing tumor growth compared to isotype-treated animals, while anti-CD112R or anti-TIGIT monotherapy either showed no activity or only a slight effect on tumor growth reduction. While anti-CD112R alone showed no activity in this assay, other experiments presented herein demonstrate the benefit of anti-CD112R monotherapy. See, for example, Figure 9.

[0216] Example 7. Increased expression of CD112R on PBMCs after anti-CD3 activation. CD112R is upregulated in in vitro activated PBMCs To determine the effect of cell activation on CD112R expression, human PBMCs were stimulated in vitro with anti-CD3 antibodies. Peripheral blood mononuclear cells (PBMCs) from healthy donors were isolated from buffy coats (Research Blood Components). Individual buffy coats were processed separately. 15 mL of buffy coat was added to each 50 mL conical tube (Corning #430290) and diluted with 15 mL of PBS (Thermofisher #14190144) + 2 mM EDTA (Fisher Scientific #BP2482-500) to a total volume of 30 mL per tube. The diluted buffy coat was placed under 14 mL of Ficolpaque (GE Healthcare Life Science #17-544203) and buffy coats were then added to each 50 mL conical tube (Corning #430290). The cells were centrifuged at 2000 RPM for 20 minutes at room temperature with the rake off. The gradient interphase was collected and washed twice with PBS + 2 mM EDTA. Isolated cells were counted and diluted to 2.5-5 x 10 in 10% DMSO (Sigma-Aldrich #472301) + 90% heat-inactivated FBS (ThermoFisher #16140-071). 7 The cells were resuspended at 1000 cells / mL.

[0217] Frozen PBMCs were quickly thawed and resuspended in supplemented RPMI medium containing RPMI + GlutaMax (1x) (ThermoFisher #61870-036), 10% heat-inactivated FBS, 1x MEM non-essential amino acid solution (ThermoFisher #15140-122), 1 mM sodium pyruvate (ThermoFisher #11360070), 100 U / mL Pen / Strep (ThermoFisher #15140-122), 1x 2-mercaptoethanol (ThermoFisher #21985023), and 10 mM Hepes (ThermoFisher #15630-080). Isolated PBMCs were washed, counted, and resuspended at a concentration of 0.5x10. 6 Cells / mL were resuspended at 1 x 10 per well. 6Cells were plated in 24-well flat-bottom plates (Corning #3526) and stimulated with 0.25 μg / mL of anti-CD3 antibody (clone UCHT1, Biolegend #300414). Cells were harvested at the indicated time points, washed in FACS buffer containing 1x PBS, 2% FBS, and 2 mM EDTA, and transferred to 96-well V-bottom plates (Costar #3894) for antibody staining.

[0218] The cells were spun down at 1500 RPM for 3 minutes and the supernatant was removed by gentle shaking. The cells were resuspended in FACS buffer and incubated with the primary antibodies in Table 5 for 1 hour at 4°C as follows: [Table 5]

[0219] The cells were washed twice and incubated with Alexa 647-conjugated anti-human IgG antibody (clone HP6017, Biolegend no. 409320) diluted 1:100 for 30 minutes at 4°C. The cells were washed once with FACS buffer and washed with 1x PBS ( Cells were stained with Live / Dead Aqua viability dye diluted 1:500 in PBS (Thermofisher #L34966) for 10 min at 4°C. Cells were washed once and acquired directly on a flow cytometer X-20 Fortessa (BD Biosciences). Data were analyzed using Flowjo (TreeStar) and Graphpad Prism (Graphpad Software).

[0220] The results are shown in Figure 6. Quantitation of CD112R antibody binding was assessed by the geometric mean fluorescence intensity (gMFI) of the Alexa Fluor® 647 signal for the indicated cell types. Anti-CD112R binding is shown as fold over negative (FON, (CD112R gMFI divided by isotype gMFI)). These results demonstrate that CD112R expression is increased on NK cells and T cells after anti-CD3 activation.

[0221] Example 8. Anti-CD112R antibodies enhance NK cell degranulation in tumor cell co-cultures. To determine the effect of anti-CD112R antibodies on NK cell-mediated degranulation, human NK cells were cocultured with Raji target cells (Burkitt lymphoma cell line, ATCC no. CCL-86) previously transduced with lentivirus to express CD112 (Origene, no. RC213693L2) in the presence of antibodies 35, 38, 44, and an isotype control.

[0222] Briefly, NK cells were isolated and pooled from PBMCs of three healthy donors via negative selection (Easysep™ NK Cell Isolation Kit, Stemcell #17955) and cultured for 16 hours in DMEM + 10% FBS + 1% penicillin-streptomycin (D10) (ThermoFisher). After overnight incubation at 37°C, NK cells were washed and resuspended in D10. Raji.CD112 cells were harvested, washed, and then resuspended in D10. 1 x 10 5 NK cells and 5 x 10 4Raji.CD112 cells were added to each well of a 96-well flat-bottom plate at a 2:1 effector-to-target cell ratio. Anti-CD112R antibody and IgG1 isotype control antibody were diluted in D10 and added to each well at a starting concentration of 10 μg / mL using 10-fold serial dilutions. Each condition was performed in duplicate. PE anti-CD107a antibody (Biolegend, no. 328608) and monensin (Biolegend, no. 420701) were also added to each well at the manufacturer's specified concentrations. The final volume for each well was 200 μl. The plate was then incubated at 37°C for 4 hours. After 4 hours, anti-CD3 FITC (Biolegend, no. 300306) and anti-NKp46 APC (Biolegend no. 331914) antibodies were diluted in D10, and 50 μl was added to each well. The plates were then incubated for an additional 30 minutes at 4°C for staining. Cells were then transferred to V-bottom plates, washed twice, and resuspended in PBS + 2% FBS. Data were acquired using an LSRFortessa X-20 (BD Biosciences) flow cytometer and analyzed with FlowJo software (Tree Star). NK degranulation was measured by CD3 - NKp46 + CD107a in the lymphocyte gate + It was defined as the frequency of cells.

[0223] The results are presented in Figure 7. Treatment of NK cells with the anti-CD112R antibodies described herein resulted in increased NK degranulation, as measured by CD107a staining, compared to isotype control.

[0224] Example 9. Anti-CD112R antibodies increase NK cell activation in PBMC-tumor cell co-cultures. To evaluate the effect of CD112R antibodies on NK cell activation, several antibodies were tested. NK cell activation was assessed in PBMC-tumor cell cocultures. Upregulation of CD137 (4-1BB), previously established as a marker of NK cell activation (Baessler et al. (2010) Blood 115(15); Andre et al. (2018) Cell 175, 1731-1743), was measured on NK cells from PBMCs cocultured with K562 target cells (chronic myeloid leukemia cell line, ATCC number CCL-243) with anti-CD112R or isotype control antibodies.

[0225] Briefly, frozen PBMCs isolated from buffy coats of healthy donors were thawed, washed, and resuspended in DMEM + 10% FBS + 1% penicillin-streptomycin (D10) at 5 × 10 per well. 5 K562 cells were plated into a 96-well flat-bottom plate at a concentration of 5 × 10 cells / well and incubated at 37°C for 4 hours before adding target cells and antibodies. In the first experiment (Figures 8A-8B), CD112R antibody and IgG1 isotype control antibody were then diluted in D10 and added to each well at a starting concentration of 10 μg / mL using a 10-fold serial dilution. In the second experiment (Figures 8C-8D), a single concentration (1 μg / mL) of anti-CD112R or IgG1 isotype control antibody was added to each well. For both experiments, each condition was performed in duplicate. K562 cells were then harvested, washed, resuspended in D10, and added at 5 × 10 cells / well. 4Cells were added to each well at a concentration of 100 μl. The final volume for each well was 200 μl. The plate was then incubated at 37°C for 16 hours. After 16 hours, the cells were transferred to a V-bottom plate and washed twice in PBS + 2% FBS. The cells were stained with anti-CD3 FITC (Biolegend, no. 300306), anti-NKp46 BV421 (Biolegend no. 331914), and anti-CD137 APC (Biolegend, no. 309810) in PBS + 2% FBS for 30 minutes at 4°C. Subsequently, the cells were washed twice and resuspended in PBS + 2% FBS. Data were acquired using an LSRFortessa X-20 (BD Biosciences) flow cytometer and analyzed with FlowJo software (Tree Star). NK cell activation was defined as the frequency of CD137+ cells within the CD3-NKp46+ lymphocyte gate.

[0226] Results from two individual donors from two independent experiments are presented in Figures 8A-8D. Addition of anti-CD112R antibody to PBMC-K562 cell cocultures resulted in significant activation of NK cells compared to isotype control, as measured by CD137 upregulation on NK cells.

[0227] Example 10. Anti-CD112R reduces tumor growth in the CT-26 model. The in vivo efficacy of CD112R blockade was evaluated in a CT26.WT colon adenocarcinoma syngeneic mouse tumor model. Seven-week-old BALB / cAnNTac female mice (Taconic Biosciences, Cat. No. BALB-F) were inoculated with 0.2 × 10 CD112R cells in 0.1 mL of 50% Geltrex (GIBCO, Cat. No. A1432-02) and 50% RPMI-1640 serum-free medium (GIBCO, Cat. No. A10491-01). 6 CT26.WT (ATCC, Catalog No. CRL-2638) tumors were implanted subcutaneously into the right flank. Mice with palpable tumors were randomized 4 days after implantation and treated intraperitoneally twice weekly for 3 weeks, starting on the day of randomization, as shown in Table 6 below.

[0228] [Table 6]

[0229] Tumor volume was determined by the tumor size limit set by the IACUC (<2000 mm 3 The tumor volume (mm) was measured every 2-3 days with a vernier caliper. 3 ) was calculated as follows: width (mm) × [length (mm)] × 0.5.

[0230] The results are presented in Figure 9. The graph shows pooled data from three independent experiments showing the mean tumor volume for each treatment group as a function of time. These results demonstrate that treatment of tumor-bearing mice with CD112R antibody resulted in a significant inhibition of tumor growth as measured 24 days after inoculation.

[0231] Example 11. CD112R blockade results in anti-tumor immunity in mice with complete tumor rejection in the CT26 model Anti-tumor immunity was evaluated in anti-CD112R-treated mice that underwent complete response from primary CT26.WT tumor challenge. For primary challenge, 7-week-old BALB / cAnNTac female mice (Taconic Biosciences, catalog no. BALB-F) were injected with 0.2 × 10 IgG in 0.1 mL of 50% Geltrex (GIBCO, catalog no. A1432-02) and 50% RPMI-1640 serum-free medium (GIBCO, catalog no. A10491-01). 6 CT26.WT (ATCC, Catalog No. CRL-2638) tumors were implanted subcutaneously into the right flank. Mice with palpable tumors were randomized 4 days after implantation and treated intraperitoneally twice weekly for 3 weeks, starting on the day of randomization, as shown in Table 7 below. [Table 7]

[0232] Tumor volume was determined by the tumor size limit set by the IACUC (<2000 mm 3The tumor volume (mm) was measured every 2-3 days with a vernier caliper. 3 ) was calculated as follows: width (mm) × [length (mm)] × 0.5.

[0233] All surviving mice at day 50 post-implantation that lacked any discernible tumor were considered survivors / complete responders. Complete responder mice (n=8) from the anti-CD112R treatment group were treated with 1x10 6 cells / mL of CD112R in 0.1 mL of 50% Geltrex (GIBCO, Cat. No. A1432-02) and 50% RPMI-1640 serum-free medium (GIBCO, Cat. No. A10491-01), a 5-fold increase from the primary inoculation dose. 6 As controls, age-matched naive Balb / c female mice (n=5) were also inoculated with 1 x 10 CT26.WT cells (ATCC, Cat. No. CRL-2638) via inoculation in the left flank. 6 CT26.WT cells were inoculated into the left flank. Mice did not receive any additional treatment. Tumor volume was determined based on tumor size within the IACUC limit (<2000 mm). 3 The tumor volume (mm3) was calculated as follows: width (mm) × [length (mm)] × 0.5.

[0234] The results are presented in Figures 10A-10B. Figure 10A shows pooled data from three independent experiments showing the survival frequency of mice implanted with anti-CD112R-treated CT26 primary tumors. Figure 11B shows the mean tumor volume for anti-CD112R-treated complete responders after tumor rechallenge and naive-challenged controls as a function of time. Statistical analysis was performed by Mantel-Cox test at 50 days post-implantation (Figure 10A) and Mann-Whitney test at 15 days post-implantation (Figure 10B). These results demonstrate that anti-CD112R-treated mice exhibit complete responses after primary tumor challenge, and the subsequent rapid rejection upon tumor rechallenge in these mice also demonstrates that treatment with anti-CD112R antibodies leads to the development of immunological memory and protective immunity.

[0235] Example 12. Both NK cells and CD8 T cells contribute to the therapeutic activity of anti-CD112R in CT26 tumor burden. The in vivo efficacy of CD112R blockade was evaluated in a CT26 syngeneic mouse tumor model after depletion of NK cells or CD8 T cells. To deplete NK and CD8 T cells, mice were treated twice weekly for 3 weeks starting at randomization with an Asialo-GM1 antibody ("asGM1" in FIG. 11, Biolegend, catalog number 146002, dose 100 μL / mouse, intraperitoneally) and an anti-CD8a antibody (Bioxcell, catalog number BE0085, 200 μg / mouse, intraperitoneally), respectively.

[0236] Seven-week-old BALB / cAnNTac female mice (Taconic Biosciences, catalog no. BALB-F) were inoculated with 0.2 × 10 cells in 0.1 mL of 50% Geltrex (GIBCO, catalog no. A1432-02) and 50% RPMI-1640 serum-free medium (GIBCO, catalog no. A10491-01). 6CT26.WT (ATCC, Catalog No. CRL-2638) tumors were implanted subcutaneously in the right flank. Mice with palpable tumors were randomized 4 days after implantation and treated with antibody 46 (anti-CD112R mouse IgG2a, 12.5 mg / kg, i.p.) intraperitoneally twice weekly for 3 weeks, starting on the day of randomization.

[0237] Tumor volume was determined by the tumor size limit set by the IACUC (<2000 mm 3 The tumor volume (mm) was measured every 2-3 days until the tumor reached a volume of 1000 mm. 3 ) was calculated as follows: width (mm) × [length (mm)] 2 ×0.5.

[0238] The results are presented in Figure 11. The graph shows the mean tumor volume for each treatment group as a function of time. These results demonstrate that the therapeutic effect of anti-CD112R is significantly reduced after depletion of NK cells or CD8 T cells. These results indicate that both CD8 T cells and NK cells are required for effective tumor growth inhibition mediated by anti-CD112R.

[0239] Example 13. CD112R blockade activates tumor NK cells in vivo. Ex vivo assessment of NK activation markers after dosing with anti-CD112R monotherapy.

[0240] To determine the effect of anti-CD112R antibodies on NK cell activation in vivo, 7-week-old BALB / cAnNTac female mice (Taconic Biosciences, catalog number BALB-F) were inoculated with 0.2 × 10 NK cells in 0.1 mL of 50% Geltrex (GIBCO, catalog number A1432-02). 6 CT26.WT (ATCC, Cat. No. CRL-2638) tumors were implanted subcutaneously in the right flank. Mice with palpable tumors were , were randomized on day 4 post-transplant and administered either 500 μg of isotype control antibody (clone C1.18.4, BioXcell, catalog number 0085) or antibody 46 (anti-CD112R mouse IgG2a). Both groups also received 500 μg of isotype control (clone MOPC-21, BioXcell, catalog number 0083). Treatments were prepared in sterile 1x PBS (GIBCO catalog number 14190-136) and injected intraperitoneally in a total volume of 100 μL.

[0241] Tumor Processing: Mice were euthanized, and tumors were excised 24 hours after treatment. Tumors were processed into single-cell suspensions by disrupting the tissue with a 440-micron mesh filter (Costar #3480) placed on a 50-mL centrifuge tube (Falcon #352350) using the rough end of a 3-mL syringe plunger (BD 301077) in FACS buffer (1x PBS, 2% heat-inactivated FBS, GIBCO Cat. No. 16140-071, 2 mM EDTA, Fisher Bioreagents, Cat. No. BP2482-500). The removed tumor was filtered again over a 70-micron strainer (Falcon Cat. No. 352350), and the remaining tissue was further disrupted using a 3-mL syringe plunger. Cells were spun down at 800 g for 10 minutes. The cell pellet was resuspended in FACS buffer.

[0242] Ex vivo restimulation: Approximately half of the single cell suspension was transferred to a 96-well U-bottom polypropylene 2 mL deep plate (Thermofisher #AB-0932) and centrifuged at 1000 g for 5 minutes at 4°C. Cells were resuspended in 20 ng / mL PMA (Abcam #ab120297), 500 ng / mL ionomycin, Ca2+, and 1000 mg / mL ionomycin. +The cells were resuspended in prewarmed 1x RPMI + Glutamax (GIBCO #61870-035) medium with 10% heat-inactivated FBS containing salts (Abcam #ab120116), 5 μg / mL brefeldin A (Biolegend #420601), and 2 μM monensin (Biolegend #420701). The cells were incubated at 37°C, 5% CO for 3.5 hours. The cells were centrifuged again as above.

[0243] Surface antigen antibody staining for FACS: The cell pellet was washed once with 500 μL of cold FACS buffer. The cells were resuspended in 100 μL of FACS buffer with TruStain fcX™ (anti-mouse CD16 / 32) at the dilution shown in Table 8. The cells were incubated on ice for 15 minutes. A surface antibody cocktail was prepared (see Table 8 for details) and added directly to the pre-blocked cells. The cells were incubated on ice for 1 hour. The cells were then washed twice with 500 μL of FACS buffer.

[0244] Viability dye staining: Cells were stained with Live / Dead Aqua viability dye diluted 1:500 in 1x PBS (Thermofisher #L34966) for 10 minutes at 4°C.

[0245] Fixation: Cells were washed once and fixed overnight at 4°C with 200 μL of eBioscience Foxp3 Fixation / Permeabilization Buffer (Thermofisher number 00-5523-00, use manufacturer's protocol for dilution guidelines).

[0246] Intracellular antigen-antibody staining for FACS: Cells were permeabilized by directly adding 200 μL of 1x eBioscience permeabilization buffer (see manufacturer's protocol for dilution guidelines). Cells were centrifuged at 1000 g for 5 minutes and stained with an intracellular panel of antibodies at a final dilution in 1x eBioscience permeabilization buffer. Cells were incubated for 1 hour at room temperature. Cells were washed twice with 500 μL of permeabilization buffer and resuspended in 150 μL of FACS buffer.

[0247] Cells were acquired on an X-20 Fortessa flow cytometer (BD Biosciences). Data were analyzed using Flowjo (Flowjo, LLC) and Graphpad Prism (Graphpad Software). [Table 8]

[0248] The results are shown in Figures 12A-B. Figure 12A shows the frequency of tumor infiltrating NK cells expressing CD69. Figure 12B shows the frequency of tumor infiltrating NK cells expressing granzyme B after ex vivo restimulation. NK cells were gated as follows: CD45 + , CD45 + SSC-A 低 , survival, singlet, NKp46 + TCRb - Population. Positive gates were set based on the fluorescence minus 1 (FMO) of the negative control. p-values ​​were derived using an unpaired t-test (*, p<0.05; **, p<0.01; ***, p<0.001). These results demonstrate that CD112R blockade significantly increases the expression of the early activation marker CD69 and the cytotoxic granule protein granzyme B in intratumoral NK cells in the CT-26 tumor model.

[0249] Example 14. The combination of anti-CD112R antibody and anti-PD1 antibody has therapeutic efficacy and increased tumor-free survival in the murine CT-26 tumor model. In vivo efficacy of combined CD112R and PD-1 blockade The efficacy of CD112R and PD-1 blockade as monotherapy and combination therapy was tested in the CT-26 colon adenocarcinoma syngeneic tumor model. Six-week-old BALB / cAnNTac female mice (Taconic Biosciences, Balb-F) were inoculated with 0.2 × 10 IgG1-positive cells in 0.1 mL of 50% Geltrex (GIBCO, Cat. No. A1432-02) and 50% RPMI-1640 serum-free medium (GIBCO, Cat. No. A10491-01). 6 CT26.WT (ATCC, Cat. No. CRL-2638) were implanted subcutaneously in the right flank. Mice were randomized to 90 mm 3 The mice were randomized into groups of 10 mice each with a mean tumor volume of 10. The mice were treated by intraperitoneal injection twice a week for 2 weeks as in Table 9. Details on the treatment agents are included in Table 10. [Table 9] [Table 10]

[0250] Tumor volumes were measured twice weekly until tumors reached the IACUC limit size (<2000 mm). Tumor volume (mm) was calculated as follows: width (mm) × [length (mm)] × 0.52.

[0251] The results are presented in Figures 13A-F. Figures 13A-E show the mean and individual tumor volume measurements for each treatment group as a function of time, respectively. The results shown in Figure 13A demonstrate that the combination of anti-CD112R with anti-PD-1 was effective and statistically significant (as measured by an unpaired t-test at day 21) in reducing tumor growth compared to isotype-treated animals. Anti-CD112R or anti-PD-1 monotherapy also demonstrated activity in reducing tumor growth. Figure 13F shows the overall tumor-free survival at day 50 post-implantation as the percentage of tumor-free survivors per group following the above treatments. These results demonstrate that the combination of anti-CD112R with anti-PD-1 confers higher tumor-free survival than the isotype control or either monotherapy agent.

[0252] Example 15. Binding of anti-CD112R antibodies to cells expressing mouse CD112R The ability of anti-CD112R antibodies to bind to mouse CD112R was evaluated on cells overexpressing mouse CD112R. 0.8 x 10 cells engineered to overexpress mouse CD112R (293T.mCD112R) were used. 5 293T cells (ATCC CRL-3216) were added to each well of a 96-well V-bottom plate and stained with either anti-CD112R antibody or IgG1 isotype control at a starting concentration of 10 μg / mL using 3-fold serial dilutions for 30 minutes at 4° C. Cells were washed twice with PBS+2% FCS and stained with Alexa Fluor® 647 anti-human IgG Fc antibody (Biolegend, catalog no. 4 Cells were resuspended in PBS (09320), diluted 1:100 in PBS + 2% FCS, and incubated for 20 minutes at 4°C. Cells were then washed twice and resuspended in PBS + 2% FCS. Cell data were acquired using an LSRFortessa X-20 (BD Biosciences) and analyzed with FlowJo software (Tree Star).

[0253] The results are shown in Figure 17 and Table 11. Quantitation of antibody binding to 293T.mCD112R cells was assessed by the geometric mean fluorescence intensity (gMFI) of the Alexa Fluor® 647 signal. These results demonstrate that several anti-CD112R antibodies bound to cells expressing mouse CD112R.

[0254] Example 16. Binding of anti-CD112R antibodies to soluble mouse CD112R The ability of anti-CD112R antibodies to bind to soluble mouse CD112R was evaluated by ELISA. Briefly, 96-well Nunc Maxisorp plates were coated overnight at 4°C with 1 μg / mL of anti-CD112R antibody or isotype control (Biolegend, Cat. No. 403502) in PBS. The plates were then washed six times with PBS + 0.01% Tween-20 (PBST) and subsequently blocked with 200 μL of PBS + 1% BSA at room temperature for 1.5 hours. After blocking, the plates were washed six times with PBST. Next, 100 μL of mouse CD112R-hIgG4 fusion protein in PBS + 1% BSA was added using a four-fold serial dilution to a final starting concentration of 10 μg / mL. The plates were incubated at room temperature for 1.5 hours. The plate was then washed six times with PBST and then incubated with 100 μL of anti-IgG4 HRP (Thermo Fisher, Cat. No. MA1-33437) at room temperature for 1 hour. The plate was then washed six times with PBST and developed with TMB substrate (Life Technologies, Cat. No. 002023). The reaction was stopped with an equal volume of stop solution (Life Technologies, Cat. No. SS04). The absorbance at 450 nm (OD450) was measured on a SpectraMax plate reader.

[0255] The results are shown in Figure 18 and Table 11. These results demonstrate that the anti-CD112R antibodies bound to soluble murine CD112R.

[0256] Example 17. Inhibition or blocking of murine CD112R binding to CD112 The ability of species-cross-reactive anti-CD112R antibodies to block mouse CD112R binding to mouse CD112 was evaluated by ELISA. Briefly, 96-well Nunc Maxisorp plates were coated overnight at 4°C with 1 μg / mL mouse CD112 (Sino Biological, Cat. No. 50318-M08H) in PBS. The plates were then washed six times with PBS + 0.01% Tween-20 (PBST) and subsequently blocked with 200 μL of PBS + 1% BSA at room temperature for 1.5 hours. After blocking, the plates were washed six times with PBST. Next, 50 μL of anti-CD112R antibody or isotype control (Biolegend, Cat. No. 403502) in PBS + 1% BSA was added using two-fold serial dilutions to a final starting concentration of 40 μg / mL. 50 μL of mouse CD112R-hIgG4 fusion protein was also added to each well at a final concentration of 2 μg / mL. The plate was incubated at room temperature for 1.5 hours. The plate was then washed six times with PBST, followed by addition of 100 μL of anti-IgG4 HRP (Thermo The plates were incubated with 100% PBST for 1 hour at room temperature with 100% TMB substrate (Life Technologies, Cat. No. MA1-33437). The plates were then washed six times with PBST and developed with TMB substrate (Life Technologies, Cat. No. 002023). The reaction was stopped with an equal volume of stop solution (Life Technologies, Cat. No. SS04). The absorbance at 450 nm (OD450) was measured on a SpectraMax plate reader.

[0257] The results are shown in Figure 19 and Table 11. These results demonstrate that the anti-CD112R antibodies inhibited mouse CD112R binding to mouse CD112 to different degrees. The percent inhibition was calculated as [100-((test sample OD450 / maximum OD450)*100%)]. The maximum OD450 was defined as the absorbance at 450 nm in the absence of antibody.

[0258] As shown in Table 11 and Figures 17-19, antibodies 32, 33, 34, 35, and 36 are able to block the interaction of human CD112 with human CD112R, but are unable to block the binding interaction between mouse CD112R and mouse CD112, according to the definition of blocking described herein. See, for example, the last two columns of Table 11, which show % inhibition of the mouse interaction at 0, 28.3, 20.3, 24.2, and 41.6, respectively, for antibodies 32, 33, 34, 35, and 36, compared to % inhibition of the human interaction at 75.1, 78.8, 80, 81.7, and 86.2, respectively. Antibodies that do not belong to the exemplary class of antibodies related to antibody 32 do not exhibit such differential blocking. [Table 11]

[0259] The foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, but the descriptions and examples should not be construed as limiting the scope of the invention. The disclosures of all patent and scientific literature cited herein are expressly incorporated by reference in their entirety. [Table 12-1] [Table 12-2] [Table 12-3] [Table 12-4] [Table 12-5] [Table 12-6]

Table 12-7

Table 12-8

Table 12-9

Table 12-10

Table 12-11

Table 12-12

Table 12-13

Table 12-14

Table 12-15

Table 12-16

Table 12-17

Table 12-18

Table 12-19

Table 12-20

Claims

1. An isolated antibody or antigen-binding fragment thereof that binds to CD112R, comprising: a. a heavy chain variable region CDR1 comprising the amino acid sequence of SEQ ID NO: 701; b. a heavy chain variable region CDR2 comprising the amino acid sequence of SEQ ID NO: 702; c. a heavy chain variable region CDR3 comprising the amino acid sequence of SEQ ID NO: 703; d. a light chain variable region CDR1 comprising the amino acid sequence of SEQ ID NO: 704; e. a light chain variable region CDR2 comprising the amino acid sequence of SEQ ID NO: 705, and f. A light chain variable region CDR3 comprising the amino acid sequence of SEQ ID NO: 706 a nucleic acid encoding the isolated antibody or antigen-binding fragment thereof, comprising:

2. The nucleic acid of claim 1, wherein the isolated antibody or antigen-binding fragment thereof comprises a heavy chain variable region having a sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:712 and a light chain variable region having a sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:

718.

3. A nucleic acid encoding an isolated antibody or antigen-binding fragment thereof that binds to CD112R and comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 712 and a light chain variable region having the amino acid sequence of SEQ ID NO:

718.

4. A nucleic acid described in any one of claims 1 to 3, wherein the isolated antibody is a monoclonal antibody.

5. A nucleic acid described in any one of claims 1 to 4, wherein the isolated antibody is a fully human antibody.

6. The nucleic acid of any one of claims 1 to 5, wherein the isolated antibody comprises an IgG1, IgG2, IgG3, or IgG4 Fc region.

7. The nucleic acid described in claim 6, wherein the isolated antibody comprises a human IgG1 heavy chain constant region.

8. The nucleic acid described in claim 6, wherein the isolated antibody comprises a human IgG4 heavy chain constant region.

9. The nucleic acid of claim 8, wherein the isolated antibody comprises a mutant human IgG4 heavy chain constant region.

10. The nucleic acid of claim 9, wherein the mutant human IgG4 heavy chain constant region comprises a mutation selected from a substitution at Ser228, a substitution at Leu235, a substitution at Asn297, or a combination thereof, when numbered according to EU numbering.

11. The nucleic acid of claim 10, wherein the mutant human IgG4 heavy chain constant region comprises an S228P substitution and an L235E substitution when numbered according to EU numbering.

12. A nucleic acid described in any one of claims 1 to 11, wherein the CD112R is human CD112R.

13. A nucleic acid described in any one of claims 1 to 12, wherein the nucleic acid comprises the sequence of SEQ ID NO:

711.

14. A nucleic acid described in any one of claims 1 to 13, wherein the nucleic acid comprises the sequence of SEQ ID NO:

717.

15. A host cell comprising a nucleic acid described in any one of claims 1 to 14.

16. A method for producing an antibody or antigen-binding fragment thereof, comprising culturing a host cell of claim 15 under conditions in which the antibody or antigen-binding fragment thereof is expressed.

17. The method of claim 16, further comprising purifying the antibody or antigen-binding fragment thereof.

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