(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)
`(19) World Intellectual Property
`Organization
`International Bureau
`
`(43) International Publication Date
`19 November 2015 (19.11.2015)
`
`WIPOI PCT
`
`\9
`
`(10) International Publication Number
`
`WO 2015/175599 A2
`
`Designated States (unless otherwise indicated, for every
`kind ofnational protection available): AE, AG, AL, AM,
`AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY,
`BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM,
`DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT,
`HN, HR, HU, ID, IL, IN, IR, IS, JP, KE, KG, KN, KP, KR,
`KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME, MG,
`MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM,
`PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, SC,
`SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN,
`TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW.
`
`(51)
`
`International Patent Classification:
`A613 5/024 (2006.01)
`H04L 29/10 (2006.01)
`
`(81)
`
`(21)
`
`International Application Number:
`
`PCT/US2015/030472
`
`(22)
`
`International Filing Date:
`
`(25)
`
`(26)
`
`(30)
`
`(71)
`
`(72)
`
`Filing Language:
`
`Publication Language:
`
`13 May 2015 (13.05.2015)
`
`English
`
`English
`
`Priority Data:
`61/992,456
`
`13 May 2014 (13.05.2014)
`
`US
`
`Applicant: ONCOMED PHARMACEUTICALS INC.
`[US/US]; 800 Chesapeake Drive, Redwood City, CA
`94063-4748 (US).
`
`Inventors: GURNEY, Austin, L.; 949 Diamond Street,
`San Francisco, CA 94114 (US). XIE, Ming-hong; 416
`Biscayne Avenue, Foster City, CA 94404 (US). PORTER,
`James, Gordon; 7261 Dumas Place, Newark, CA 94560
`(US).
`
`(74)
`
`Agents: CALVO, Paul, A. et al.; Sterne, Kessler, Gold—
`slein & Fox, PLLC, 1100 New York Ave., N W, Washing-
`ton, DC 20005 (US).
`
`(84)
`
`Designated States (unless otherwise indicated, for every
`kind of regional protection available): ARIPO (BW, GH,
`GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ,
`TZ, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU,
`TJ, TM), European (AL, AT, BE, BG, CH, CY, CZ, DE,
`DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU,
`LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK,
`SM, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ,
`GW, KM, ML, MR, NE, SN, TD, TG).
`Published:
`
`without international search report and to be republished
`upon receipt ofthat report (Rule 48.2(g))
`
`(54)
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`Title: IMMUNOTHERAPY WITH BINDING AGENTS
`
`(57) Abstract: Binding agents that modulate the immune response are disclosed. The binding agents may include antibodies, soluble
`receptors, and/or polypeptides. Also disclosed are methods of using the binding agents for the treatment of diseases such as cancer.
`
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`PCT/US2015/030472
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`IMMUNOTHERAPY WITH BINDING AGENTS
`
`CROSS-REFERENCE TO RELATED APPLICATONS
`
`[001]
`
`This application claims priority benefit of US. Provisional Application No. 61/992,456, filed
`
`May 13, 2014, which is hereby incorporated by reference herein in its entirety.
`
`FIELD OF THE INVENTION
`
`[002]
`
`This invention generally relates to agents that modulate the immune response, such as antibodies,
`
`soluble receptors, and small molecules, as well as to methods of using the agents for the treatment of
`
`diseases such as cancer.
`
`BACKGROUND OF THE INVENTION
`
`[003]
`
`The basis for immunotherapy is the manipulation and/or modulation of the immune system,
`
`including both innate immune responses and adaptive immune responses. The general aim of
`
`immunotherapy is to treat diseases by controlling the immune response to a “foreign agent”, for example a
`
`pathogen or a tumor cell. However, in some instances immunothcrapy is used to treat autoimmune
`
`diseases which may arise from an abnormal immune response against proteins, molecules, and/or tissues
`
`normally present in the body. Immunotherapy may include methods to induce or enhance specific
`
`immune responses or to inhibit or reduce specific immune responses. The immune system is a highly
`
`complex system made up of a great number of cell types, including but not limited to, T—cells, B—cells,
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`natural killer cells, antigen-presenting cells, dendritic cells, monocytes, and macrophages. These cells
`
`possess complex and subtle systems for controlling their interactions and responses. The cells utilize both
`
`activating and inhibitory mechanisms and feedback loops to keep responses in check and not allow
`
`negative consequences of an uncontrolled immune response (e.g., autoimmune diseases).
`
`[004] Generally, an immune response is initiated through antigen recognition by the T-cell receptor
`
`(TCR) and is regulated by a balance between stimulatory and inhibitory signals (i.e., immune
`
`checkpoints). Under normal conditions, immune checkpoints are necessary to maintain a balance between
`
`activating and inhibitory signals and to ensure the development of an effective immune response while
`
`safeguarding against the development of autoimmunity or damage to tissues when the immune system is
`
`responding to a foreign or pathogenic agent. An important immune checkpoint receptor is CTLA4 which
`
`is expressed on T-cells and is highly expressed on regulatory T-cells (Tregs). CTLA4 is considered to act
`
`as an inhibitory molecule or an immune response “brake” and primarily regulates the amplitude ofT—cell
`
`activation. CTLA4 counteracts the activity of the co—stimulatory receptor, CD28, which acts in concert
`
`with the TCR to activate T-cells. CTLA4 and CD28 share identical ligands or counter-receptors, B7—l
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`(CD80) and B7-2 (CD86) and the balance of the immune response probably involves competition of
`
`CTLA4 and CD28 for binding to the ligands. Another important immune checkpoint receptor is PD]
`
`which is expressed on T—cells after activation, highly expressed on Tregs, and expressed on other
`
`activated cells including B-cells and natural killer (NK) cells. Similar to CTLA4, PDl is considered to act
`
`as an inhibitory molecule and brake on the immune response. There are two ligands/counter—receptors for
`
`PD], PDLl (also known as B7-H1 and CD247) and PDL2 (also known as B7—DC and CD273). (See,
`
`Pardoll, 2012, Nature Reviews Cancer, 12:252—264).
`
`[005]
`
`The concept of cancer immunosurveillance is based on the theory that the immune system can
`
`recognize tumor cells, mount an immune response, and suppress the development and/or progression of a
`
`tumor. However, it is clear that many cancerous cells have developed mechanisms to evade the immune
`
`system which can allow for uninhibited growth of tumors. Immune checkpoints can be dysregulated by
`
`tumors and may be manipulated by tumors to be used as an immune resistance mechanism. Cancer
`
`immunotherapy focuses on the development of agents that can activate and/or boost the immune system to
`
`achieve a more effective response to killing tumor cells and inhibiting tumor growth.
`
`BRIEF SUMMARY OF THE INVENTION
`
`[006]
`
`The present invention provides agents, such as antibodies, soluble receptors, and small molecules
`
`that modulate the immune response. In some embodiments, the agents activate or increase the immune
`
`response to cancer and/or a tumor. The invention also provides compositions, such as pharmaceutical
`
`compositions, comprising the agents. The invention further provides methods of administering the agents
`
`to a subject in need thereof. In some embodiments, the invention provides methods of using the agents for
`
`cancer immunotherapy.
`
`[007]
`
`In one aspect, the present invention provides agents that bind V—set and transmembrane domain-
`
`containing protein 4 (VSTM4). In some embodiments, the invention provides an agent that specifically
`
`binds the extracellular domain, or a fragment thereof, of VSTM4. In some embodiments, the agent
`
`specifically binds the Ig—like domain of VSTM4. In some embodiments, the agent specifically binds the
`
`IgV domain of VSTM4. In some embodiments, the agent specifically binds human VSTM4. In some
`
`embodiments, the agent specifically binds mouse VSTM4. In some embodiments, the agent specifically
`
`binds human VSTM4 and mouse VSTM4. In some embodiments, the agent binds within amino acids 24—
`
`180 of human VSTM4 and/or mouse VSTM4. In some embodiments, the agent binds within amino acids
`
`24-155 of human VSTM4 and/or mouse VSTM4. In some embodiments, the agent binds with amino
`
`acids 24—130 of human VSTM4 and/or mouse VSTM4. In some embodiments, the agent binds within
`
`SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID N028, SEQ ID N029, SEQ ID NO:10, or a
`
`fragment thereof.
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`[008] As used herein, an “agent” or “binding agent” includes but is not limited to, an antibody, a soluble
`
`receptor, a secreted (e. g., soluble) protein, a polypeptide, and a small molecule.
`
`[009]
`
`In some embodiments, the agent is an antibody. In some embodiments, the antibody is a
`
`monoclonal antibody, a chimeric antibody, a humanized antibody, a human antibody, a bispecific
`
`antibody, or an antibody fragment.
`
`[010]
`
`In some embodiments, the agent is a soluble receptor or a soluble protein. In some embodiments,
`
`the soluble receptor comprises the extracellular domain, or a fragment thereof, of VSTM4. In some
`
`embodiments, the soluble receptor comprises the extracellular domain, or a fragment thereof, of human
`
`VSTM4. In some embodiments, the soluble receptor comprises the extracellular domain, or a fragment
`
`thereof, of mouse VSTM4. In some embodiments, the soluble receptor comprises the Ig—like domain of
`
`VSTM4. In some embodiments, the soluble receptor comprises the IgV domain of VSTM4. In some
`
`embodiments, the soluble receptor comprises amino acids 24-180 of human VSTM4 or amino acids 24-
`
`179 of mouse VSTM4. In some embodiments, the soluble receptor comprises amino acids 24—155 of
`
`human VSTM4 or amino acids 24-154 of mouse VSTM4. In some embodiments, the soluble receptor
`
`comprises amino acids 24—130 of human VSTM4 or amino acids 24—129 of mouse VSTM4. In some
`
`embodiments, the soluble receptor comprises SEQ ID NO:3, SEQ ID NO:4, SEQ ID N025, SEQ ID
`
`NO:8, SEQ ID NO:9, SEQ ID NO:10, or a fragment thereof. In some embodiments, the soluble receptor
`
`or soluble protein is a fusion protein. In some embodiments, the fusion protein comprises a non-VSTM4
`
`polypeptide (i.e., a heterologous protein). In some embodiments, the filsion protein comprises a Fc
`
`region. In some embodiments, the non-VSTM4 polypeptide comprises a Fc region. In some
`
`embodiments, the Fc region is selected from the group consisting of SEQ ID NO:21, SEQ ID NO:22, SEQ
`
`ID NO:23, SEQ ID NO:24, SEQ ID N025, and SEQ ID NO:26.
`
`[011]
`
`In some embodiments of each of the aforementioned aspects and embodiments, as well as other
`
`aspects and embodiments described herein, the agent is monovalent. In some embodiments, the agent is
`
`bivalent. In some embodiments, the agent is monospecific. In some embodiments, the agent is bispecific.
`
`[012]
`
`In some embodiments of each of the aforementioned aspects and embodiments, as well as other
`
`aspects and embodiments described herein, the agent is a heteromultimeric protein. In some
`
`embodiments, the agent is a heterodimeric protein. In some embodiments, the heterodimeric protein
`
`comprises a first polypeptide which binds VSTM4 and a second polypeptide which binds a second target.
`
`In some embodiments, the heterodimeric protein comprises a first polypeptide which binds VSTM4 and a
`
`second polypeptide which is an immune response stimulating agent. In some embodiments, the
`
`heterodimeric protein comprises a first polypeptide which binds B7—H4 and a second polypeptide which
`
`binds a second target. In some embodiments, the heterodimeric protein comprises a first polypeptide
`
`which binds B7-H4 and a second polypeptide which is an immune response stimulating agent. In some
`
`embodiments, the heterodimeric protein comprises a first polypeptide comprising an agent described
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`herein and a second polypeptide comprising an immune response stimulating agent. In some
`
`embodiments, the immune response stimulating agent is selected from the group consisting of
`
`granulocyte-macrophage colony stimulating factor (GM-CSF), macrophage colony stimulating factor (M-
`
`CSF), granulocyte colony stimulating factor (G-CSF), interleukin 3 (IL-3), interleukin 12 (IL-l2),
`
`interleukin 1 (IL—1), interleukin 2 (IL-2), B7-l (CD80), B7-2 (CD86), anti-CD3 antibody, anti-CTLA4
`
`antibody, anti-PD-Ll antibody, and anti-PD] antibody. In some embodiments, the heterodimeric protein
`
`comprises two polypeptides, wherein each polypeptide comprises a human IgG2 CH3 domain, and
`
`wherein the amino acids at positions corresponding to positions 249 and 288 of SEQ ID NO:28 of one
`
`IgG2 CH3 domain are replaced with glutamate or aspartate, and wherein the amino acids at positions
`
`corresponding to positions 236 and 278 of SEQ ID NO:28 of the other IgG2 CH3 domain are replaced
`
`with lysine.
`
`[013]
`
`In some embodiments of each of the aforementioned aspects and embodiments, as well as other
`
`aspects and embodiments described herein, the binding agent induces, activates, promotes, increases,
`
`enhances, and/or prolongs an immune response. In some embodiments, the binding agent increases cell—
`
`mediated immunity.
`
`In some embodiments, the binding agent increases T-cell activity.
`
`In some
`
`embodiments, the agent increases cytolytic T—cell (CTL) activity. In some embodiments, the agent
`
`increases natural killer (NK) cell activity. In some embodiments, the agent decreases Treg activity. In
`
`some embodiments, the agent is an antagonist of B7-H4-mediated signaling. In some embodiments, the
`
`agent is an antagonist of VSTM4 signaling. In some embodiments, the agent inhibits VSTM4 signaling.
`
`In some embodiments, the agent inhibits or blocks the interaction between B7-H4 and VSTM4.
`
`[014]
`
`In some embodiments of each of the aforementioned aspects and embodiments, as well as other
`
`aspects and embodiments described herein, the agent specifically binds VSTM4 and disrupts signaling of
`
`a B7—H4/VSTM4 pathway. In some embodiments, the agent specifically binds B7-H4 and disrupts
`
`signaling of a B7-H4 pathway. In some embodiments, the agent specifically binds VSTM4 and the agent
`
`disrupts binding of VSTM4 to B7—H4, and/or disrupts B7-H4 activation of VSTM4 signaling. In some
`
`embodiments, the agent specifically binds B7—H4 and the agent disrupts binding of B7—H4 to VSTM4,
`
`and/or disrupts B7—H4 activation of VSTM4 signaling. In some embodiments, the agent disrupts binding
`
`of B7-H4 to VSTM4. In some embodiments, the agent disrupts binding of VSTM4 to B7-H4. In some
`
`embodiments, the agent disrupts B7—H4 activation of VSTM4 signaling. In some embodiments, the agent
`
`inhibits activity of VSTM4. In some embodiments, the agent inhibits the inhibitory activity of VSTM4.
`
`In some embodiments, the agent induces, activates, promotes, increases, enhances, and/or prolongs an
`
`immune response.
`
`[015]
`
`In some embodiments of each of the aforementioned aspects and embodiments, as well as other
`
`aspects and embodiments described herein, the agent specifically binds B7-H4 and the agent disrupts the
`
`interaction between B7-H4 and VSTM4, and/or disrupts B7—H4 activation of VSTM4 signaling. In some
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`embodiments, the agent disrupts the interaction between B7-H4 and VSTM4. In some embodiments, the
`
`agent disrupts the interaction between VSTM4 and B7—H4. In some embodiments, the agent disrupts B7-
`
`H4 activation of VSTM4 signaling.
`
`ln some embodiments, the agent induces, activates, promotes,
`
`increases, enhances, or prolongs an immune response.
`
`[016]
`
`In another aspect, the invention provides compositions comprising an antibody, a soluble
`
`receptor, a soluble protein, a polypeptide, or a binding agent described herein. Methods of using a
`
`composition comprising an agent described herein are also provided.
`
`[017]
`
`In another aspect, the invention provides pharmaceutical compositions comprising an antibody, a
`
`soluble receptor, a soluble protein, a polypeptide, or a binding agent described herein and a
`
`pharmaceutically acceptable carrier. Methods of treating cancer and/or inhibiting tumor growth in a
`
`subject (e. g., a human) comprising administering to the subject an effective amount of a composition
`
`comprising an agent described herein are also provided. Methods of treating a viral infection in a subject
`
`(e. g., a human) comprising administering to the subject an effective amount of a composition comprising
`
`an agent described herein are also provided.
`
`[018]
`
`In certain embodiments of each of the aforementioned aspects, as well as other aspects and/or
`
`embodiments described elsewhere herein, the antibody, the soluble receptor, the soluble protein, the
`
`polypeptide, or the binding agent is isolated. In certain embodiments, the antibody, the soluble receptor,
`
`the soluble protein, the polypeptide, or the binding agent is substantially pure.
`
`[019]
`
`In another aspect, the invention provides polynuclcotidcs comprising a polynucleotide that
`
`encodes an antibody, a soluble receptor, a soluble protein, a polypeptide, or a binding agent described
`
`herein. In some embodiments, the polynucleotide is isolated. In some embodiments, the invention
`
`provides vectors that comprise the polynuclcotidcs, as well as cells that comprise the vectors and/or the
`
`polynucleotides. In some embodiments, the invention also provides cells comprising or producing an
`
`antibody, a soluble receptor, a soluble protein, a polypeptide, or a binding agent described herein. In
`
`some embodiments, the cell is a monoclonal cell line.
`
`[020]
`
`In another aspect, the invention provides methods of modulating the immune response of a
`
`subject. In some embodiments, the invention provides a method of inducing an immune response in a
`
`subject comprising using an agent described herein. In some embodiments, the invention provides a
`
`method of activating an immune response in a subject comprising using an agent described herein. In
`
`some embodiments, the invention provides a method of promoting an immune response in a subject
`
`comprising using an agent described herein. In some embodiments, the invention provides a method of
`
`increasing an immune response in a subject comprising using an agent described herein. In some
`
`embodiments, the invention provides a method of enhancing an immune response in a subject comprising
`
`using an agent described herein. In some embodiments, the invention provides a method of prolonging an
`
`immune response in a subject comprising using an agent described herein. In some embodiments, the
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`immune response is to an antigenic stimulation. In some embodiments, the antigenic stimulation is a
`
`tumor or a tumor cell. In some embodiments, the antigenic stimulation is a pathogen. In some
`
`embodiments, the antigenic stimulation is a virus. In some embodiments, the antigenic stimulation is a
`
`virally-infected cell. In some embodiments, the invention provides a method of increasing the activity of
`
`immune cells. In some embodiments, the invention provides a method of increasing the activity of
`
`immune cells comprising contacting the cells with an effective amount of an agent described herein. In
`
`some embodiments, the immune cells are T—cells, NK cells, monocytes, macrophages, and/or B—cells. In
`
`some embodiments, the invention provides a method of increasing the activity of NK cells in a subject
`
`comprising administering to the subject a therapeutically effective amount of an agent described herein.
`
`In some embodiments, the invention provides a method of increasing the activity of T—cells in a subject
`
`comprising administering to the subject a therapeutically effective amount of an agent described herein.
`
`In some embodiments, the invention provides a method of increasing the activation of T-cells and/or NK
`
`cells in a subject comprising administering to the subject a therapeutically effective amount of an agent
`
`described herein. In some embodiments, the invention provides a method of increasing the T-cell
`
`response in a subject comprising administering to the subject a therapeutically effective amount of an
`
`agent described herein. In some embodiments, the invention provides a method of increasing the activity
`
`of CTLs in a subject comprising administering to the subject a therapeutically effective amount of an
`
`agent described herein.
`
`[021]
`
`In another aspect, the invention provides methods of inducing, activating, promoting, increasing,
`
`enhancing, or prolonging an immune response in a subject, comprising administering to the subject a
`
`therapeutically effective amount of an agent described herein. In some embodiments, the invention
`
`provides methods of inducing, activating, promoting, increasing, enhancing, or prolonging an immune
`
`response in a subject, comprising administering to the subject a therapeutically effective amount of an
`
`agent that disrupts the signaling of a B7-H4/VSTM4 pathway. In some embodiments, the disruption of
`
`the signaling of a B7-H4/VSTM4 pathway inhibits the B7-H4-mediated suppression of immune
`
`responses. In some embodiments, the immune response is against a tumor or cancer. In some
`
`embodiments, the immune response is against a Viral infection, a Viral antigen, or a virally—infected cell.
`
`[022]
`
`In another aspect, the invention provides methods of inhibiting tumor growth comprising
`
`contacting a tumor or tumor cell with an effective amount of an agent described herein. In some
`
`embodiments, a method of inhibiting growth of a tumor comprises contacting a tumor or tumor cell with
`
`an effective amount of an agent that disrupts the interaction between B7—H4 and VSTM4.
`
`[023]
`
`In another aspect, the invention provides methods of inhibiting tumor growth in a subject
`
`comprising administering to the subject a therapeutically effective amount of an agent described herein.
`
`In some embodiments, a method of inhibiting growth of a tumor in a subject comprises administering to
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`the subject a therapeutically effective amount of an agent that disrupts the interaction between B7-H4 and
`
`VSTM4.
`
`[024]
`
`In another aspect, the invention provides methods of treating cancer in a subject comprising
`
`administering to the subject a therapeutically effective amount of an agent described herein. In some
`
`embodiments, a method of treating cancer in a subject comprises administering to the subject a
`
`therapeutically effective amount of an agent that disrupts the interaction between B7-H4 and VSTM4.
`
`[025]
`
`In another aspect, the invention provides methods of stimulating a protective response in a subject
`
`comprising administering to the subject a therapeutically effective amount of an agent described herein in
`
`combination with an antigen of interest. In some embodiments, the antigen of interest is a tumor antigen.
`
`In some embodiments, the antigen of interest is a cancer stem cell marker.
`
`[026]
`
`In some embodiments of each of the aforementioned aspects and embodiments, as well as other
`
`aspects and embodiments described herein, the methods comprise administering to the subject an immune
`
`response stimulating agent. In some embodiments, the immune response stimulating agent is selected
`
`from the group consisting of GM-CSF, M-CSF, G-CSF, IL-3, IL-12, IL—1, IL—2, B7-l (CD80), B7—2
`
`(CD86), anti—CD3 antibodies, anti-CTLA—4 antibodies, anti-CD28 antibodies, anti-PD-Ll antibodies, and
`
`anti-PD] antibodies.
`
`[027] Where aspects or embodiments of the invention are described in terms of a Markush group or
`
`other grouping of alternatives, the present invention encompasses not only the entire group listed as a
`
`whole, but also each member of the group individually and all possible subgroups of the main group, and
`
`also the main group absent one or more of the group members. The present invention also envisages the
`
`explicit exclusion of one or more of any of the group members in the claimed invention.
`
`BRIEF DESCRIPTION OF THE FIGURES
`
`[028]
`
`Figure 1. Family tree of B7 family members.
`
`[029]
`
`Figure 2. FACS analysis of binding interactions between B7-H4 and VSTM4. HEK-293T cells
`
`were transiently transfected with a cDNA expression vector encoding membrane—bound VSTM4—
`
`CD4TM-GFP and then subsequently mixed with soluble B7—H4—COMP—FLAG. Specific binding is
`
`indicated by the presence of signal within the dark square overlay on each FACS plot.
`
`[030]
`
`Figure 3. Sequence of human VSTM4 (SEQ ID NO:1). The predicted signal sequence is
`
`underlined (solid line); the predicted Ig—like domain is underlined (dotted line); and the predicted
`
`transmembrane domain is boxed. The ITIM and ITSM sequences are marked.
`
`[031]
`
`Figure 4. Diagrammatic representation of immune checkpoint signaling systems.
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`DETAILED DESCRIPTION OF THE INVENTION
`
`[032]
`
`The present invention provides novel agents, including, but not limited to, antibodies,
`
`polypeptides, soluble receptors, and soluble proteins that modulate the immune response. The agents
`
`include agonists and antagonists of receptors and counter-receptors (or ligands) that are members of the
`
`immunoglobulin superfamily involved in cell interactions and immune response signaling. Related
`
`polypeptides and polynucleotides, compositions comprising the agents, pharmaceutical compositions
`
`comprising the agents, and methods of making the agents are also provided. Methods of screening for
`
`agents that modulate the immune response are provided. Methods of using the novel agents, such as
`
`methods of activating an immune response, methods of inducing an immune response, methods of
`
`promoting an immune response, methods of increasing an immune response, methods of prolonging an
`
`immune response, methods of activating natural killer (NK) cells and/or T-eells, methods of increasing the
`
`activity of NK cells and/or T-cells, methods of promoting the activity of NK cells and/or T—cells, methods
`
`of decreasing the activity of Treg cells, methods of inhibiting tumor growth, and/or methods of treating
`
`cancer are further provided.
`
`[033]
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`The CD28/CTLA4 signaling system is recognized as containing two ligands or counter-receptors,
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`B7—l (CD80) and B7-2 (CD86) which each bind to CTLA4 and CD28. Within this signaling axis, CD28
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`serves as an activating receptor, whereas CTLA4 serves as an inhibitory receptor. The intracellular
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`domain of CD28 contains an immunoreceptor tyrosine—based activation motif or ITAM characterized by
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`the consensus sequence YXX(L/l)(X)(6_g)YXX(L/I) (SEQ ID NO:37), that is, at least in part, responsible
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`for the stimulatory activity of CD28. In comparison, the intracellular domain of CTLA4 contains an
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`immunoreceptor tyrosine-based inhibitory motif or ITIM characterized by the consensus sequence
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`(S/IN/L)XYXX(I/V/L) (SEQ ID NO:38), that is, at least in part, responsible for the inhibitory activity of
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`CTLA4 (Barrow A et al., 2006, Eur Jlmmunol., 36: 1 646-53). In each motif consensus sequence the “X”
`
`represents any amino acid.
`
`[034]
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`The PD-l signaling system is recognized as containing two ligands or counter-receptors, PD—Ll
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`(B7—H2) and PD—L2 (B7—DC), which each bind to the PD—l receptor. Similar to CTLA4, PD—l contains
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`an ITIM which is responsible for providing an inhibitory signal to T—cells. It is noteworthy that there has
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`been no activating receptor identified for PD-Ll or PD—L2 that would con‘espond in an analogous fashion
`
`to the CD28 receptor utilized by B7—l and B7—2. However, there has been speculation that such a receptor
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`or receptors exist (see, e.g., Ishiwata et al., 2010, J. Immunol., 184:2086—2094; Shin et al., 2003, J. Exp.
`
`Med, 198:31-38; Shin et al., 2005, J. Exp. Med, 201:1531—1541; Wang et al., 2003, J. Exp. Med,
`
`197:1083-1091).
`
`[035]
`
`B7—H4 is a member of the B7 family, which includes, but may not be limited to, B7-l, B7-2, PD-
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`Ll, PD—L2, B7-H2, B7-H3, B7-H4, B7—H5, B7—H6, Gi24, and the butyrophilin and butyrophilin-like
`
`proteins. A family tree depicting members of the B7 family is shown in Figure l. A number of B7
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`proteins are considered to be “orphan molecules” as their interacting receptors, either stimulatory or
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`inhibitory, have not yet been identified and/or reported. This group includes PD-Ll (stimulatory
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`receptor), PD-L2 (stimulatory receptor), B7-l—l3, B7-l—l4, B7—l-15, Gi24, and the BTN family of proteins.
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`B7—H4 has been shown to not bind known CD28 family members such as CD28, CTLA-4, ICOS, and PD-
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`] (Sica et al., 2003, Immunity, 18:849—861). Functional studies using B7-H4 transfectants and B7-H4-Ig
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`fusion proteins demonstrate that B7—H4 delivers a signal that inhibits T-cell proliferation, cell-cycle
`
`progression, and lL—2 production. It is believed that B7—H4 may be part of another important immune
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`checkpoint signaling pathway. Thus it would be highly advantageous to identify the receptor for B7—H4
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`and to develop agents that inhibit the B7-H4 signaling pathway.
`
`1. Definitions
`
`[036]
`
`To facilitate an understanding of the present invention, a number of tenns and phrases are defined
`
`below.
`
`[037]
`
`The terms “agonist” and “agonistic” as used herein refer to or describe an agent that is capable of,
`
`directly or indirectly, substantially inducing, activating, promoting, increasing, or enhancing the biological
`
`activity of a target and/or a pathway. The term “agonist” is used herein to include any agent that partially
`
`or fully induces, activates, promotes, increases, or enhances the activity of a protein. Suitable agonists
`
`specifically include, but are not limited to, agonist antibodies or fragments thereof, soluble receptors,
`
`other fusion proteins, polypeptides, and small molecules.
`
`[038]
`
`The terms “antagonist” and “antagonistic” as used herein refer to or describe an agent that is
`
`capable of, directly or indirectly, partially or fully blocking, inhibiting, reducing, or neutralizing a
`
`biological activity of a target and/or pathway. The term “antagonist” is used herein to include any agent
`
`that partially or fully blocks, inhibits, reduces, or neutralizes the activity of a protein. Suitable antagonist
`
`agents specifically include, but are not limited to, antagonist antibodies or fragments thereof, soluble
`
`receptors, other fusion proteins, polypeptides, and small molecules.
`
`[039]
`
`The terms “modulation” and “modulate” as used herein refer to a change or an alteration in a
`
`biological activity. Modulation includes, but is not limited to, stimulating or inhibiting an activity.
`
`Modulation may be an increase or a decrease in activity, a change in binding characteristics, or any other
`
`change in the biological, filnctional, or immunological properties associated with the activity of a protein,
`
`a pathway, a system, or other biological targets of interest.
`
`[040]
`
`The term “antibody” as used herein refers to an immunoglobulin molecule that recognizes and
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`specifically binds a target, such as a protein, polypeptide, peptide, carbohydrate, polynucleotide, lipid, or
`
`combinations of the foregoing, through at least one antigen recognition site within the variable region of
`
`the immunoglobulin molecule. As used herein, the term encompasses intact polyclonal antibodies, intact
`
`monoclonal antibodies, antibody fragments (such as Fab, Fab', F (ab')2, and Fv fragments), single chain Fv
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`(scFv) antibodies, multispecific antibodies such as bispecific antibodies generated from at least two intact
`
`antibodies, bispecific antibodies, monospecific antibodies, monovalent antibodies, chimeric antibodies,
`
`humanized antibodies, human antibodies, fusion proteins comprising an antigen-binding site of an
`
`antibody, and any other modified immunoglobulin molecule comprising an antigen—binding site as long as
`
`the antibodies exhibit the desired biological activity. An antibody can be any of the five major classes of
`
`immunoglobulins: lgA, IgD, lgE, lgG, and IgM, or subclasses (isotypes) thereof (e.g., IgGl, IgG2, IgG3,
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`IgG4, lgAl and IgA2), based on the identity of their heavy—chain constant domains referred to as alpha,
`
`delta, epsilon, gamma, and mu, respectively. The different classes of immunoglobulins have different and
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`well-known subunit structures and three-dimensional configurations. Antibodies can be naked or
`
`conjugated to other molecules, including but not limited to, toxins and radioisotopes.
`
`[041]
`
`The term “antibody fragment” refers to a portion of an intact antibody and refers to the antigenic
`
`detemiining variable regions of an intact antibody. Examples of antibody fi‘agments include, but are not
`
`limited to, Fab, Fab', F(ab')2, and FV fragments, linear antibodies, single chain antibodies, and
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`multispecific antibodies for