WO2020216383A1 - Anti-lag-3 antibodies - Google Patents

Anti-lag-3 antibodies Download PDF

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Publication number
WO2020216383A1
WO2020216383A1 PCT/CN2020/087141 CN2020087141W WO2020216383A1 WO 2020216383 A1 WO2020216383 A1 WO 2020216383A1 CN 2020087141 W CN2020087141 W CN 2020087141W WO 2020216383 A1 WO2020216383 A1 WO 2020216383A1
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WO
WIPO (PCT)
Prior art keywords
seq
amino acid
acid sequence
variable domain
chain variable
Prior art date
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PCT/CN2020/087141
Other languages
French (fr)
Inventor
Leyan TANG
Allison SCHULKINS
Kimberly THAN
Chun-Nan Chen
Jingyun MIAO
Xiaohui Zhang
Xiaojuan Shi
Lin Zhang
Pingju GE
Original Assignee
Single Cell Technology, Inc.
Acrobiosystems, Inc.
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Application filed by Single Cell Technology, Inc., Acrobiosystems, Inc. filed Critical Single Cell Technology, Inc.
Publication of WO2020216383A1 publication Critical patent/WO2020216383A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/70—Immunoglobulins specific features characterized by effect upon binding to a cell or to an antigen
    • C07K2317/76—Antagonist effect on antigen, e.g. neutralization or inhibition of binding
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
    • C07K2317/92—Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value

Definitions

  • Lymphocyte Activation Gene-3 is a member of the immunoglobulin supergene family, and is expressed on activated T cells (Huard et al., 1994a) , NK cells (Triebel et al., 1990) , regulatory T cells (Huang et al., 2004; Camisaschi et al., 2010; Gagliani et al., 2013) , and plasmacytoid dendritic cells (DCs) (Workman et al., 2009) .
  • LAG-3 is a membrane protein encoded by a gene located on chromosome 12 and is structurally and genetically related to CD4.
  • LAG-3 can interact with MHC class II molecules on the cell surface (Baixeras et al., 1992; Huard et al., 1996) . It has been suggested that the direct binding of LAG-3 to MHC class II plays a role in down-regulating antigen-dependent stimulation of CD4+T lymphocytes (Huard et al., 1994b) and LAG-3 blockade has also been shown to reinvigorate CD8+ lymphocytes in both tumor or self-antigen (Grosso et al., 2007; Blackburn et al., 2009) .
  • LAG-3 intra-cytoplasmic region of LAG-3 can interact with LAP (LAG-3-associated protein) , which is a signal transduction molecule involved in the downregulation of the CD3/TCR activation pathway (Iouzalen et al., 2001) .
  • LAP LAG-3-associated protein
  • CD4+CD25+ regulatory T cells have been shown to express LAG-3 upon activation, which contributes to the suppressor activity of Treg cells (Huang et al., 2004) .
  • LAG-3 can also negatively regulate T cell homeostasis by Treg cells in both T cell-dependent and independent mechanisms (Workman and Vignali, 2005) .
  • the anti-LAG-3 antibody molecules disclosed herein can be used to treat, prevent and/or diagnose cancerous or malignant disorders (e.g., cancers such melanoma, e.g., advanced stage melanoma; pancreatic cancer, e.g., advanced pancreatic cancer; solid tumors; breast cancer, e.g., metastatic breast carcinoma; renal cell carcinoma, e.g., advanced or metastatic renal cell carcinoma (MRCC) or clear cell renal cell carcinoma) , as well as potentially infectious diseases (e.g., hepatitis, e.g., hepatitis B; influenza) .
  • cancers such melanoma, e.g., advanced stage melanoma
  • pancreatic cancer e.g., advanced pancreatic cancer
  • solid tumors breast cancer, e.g., metastatic breast carcinoma
  • renal cell carcinoma e.g., advanced or metastatic renal cell carcinoma (MRCC) or clear cell renal cell carcinoma
  • infectious diseases e.g.,
  • SCT-Ca20 mature heavy chain variable domain protein sequence 20 SCT-Ca21 mature heavy chain variable domain protein sequence 21 SCT-Ca22 mature heavy chain variable domain protein sequence 22 SCT-Ca23 mature heavy chain variable domain protein sequence 23 SCT-Ca24 mature heavy chain variable domain protein sequence 23 SCT-Ca25 mature heavy chain variable domain protein sequence 24 SCT-Ca26 mature heavy chain variable domain protein sequence 25 SCT-Ca27 mature heavy chain variable domain protein sequence 26 SCT-Ca28 mature heavy chain variable domain protein sequence 27 SCT-Ca29 mature heavy chain variable domain protein sequence 28 SCT-Ca30 mature heavy chain variable domain protein sequence 29 SCT-Ca31 mature heavy chain variable domain protein sequence 29 SCT-Ca32 mature heavy chain variable domain protein sequence 30 SCT-Ca33 mature heavy chain variable domain protein sequence 31 SCT-Ca34 mature heavy chain variable domain protein sequence 32 SCT-Ca35 mature heavy chain variable domain protein sequence 33 SCT-Ca36 mature heavy chain variable domain protein sequence 34 SCT-Ca37 mature heavy chain variable domain protein sequence 35 SCT
  • SCT-Ca61 mature heavy chain variable domain protein sequence 59 SCT-Ca62 mature heavy chain variable domain protein sequence 59 SCT-Ca63 mature heavy chain variable domain protein sequence 60 SCT-Ca64 mature heavy chain variable domain protein sequence 61 SCT-Ca65 mature heavy chain variable domain protein sequence 62 SCT-Ca66 mature heavy chain variable domain protein sequence 63 SCT-Ca67 mature heavy chain variable domain protein sequence 64 SCT-Ca68 mature heavy chain variable domain protein sequence 65 SCT-Ca69 mature heavy chain variable domain protein sequence 66 SCT-Ca70 mature heavy chain variable domain protein sequence 67 SCT-Ca01 mature light chain variable domain protein sequence 68 SCT-Ca02 mature light chain variable domain protein sequence 69 SCT-Ca03 mature light chain variable domain protein sequence 70 SCT-Ca04 mature light chain variable domain protein sequence 71 SCT-Ca05 mature light chain variable domain protein sequence 72 SCT-Ca06 mature light chain variable domain protein sequence 73 SCT-Ca07 mature light chain variable domain protein sequence 74 SCT-C
  • SCT-Ca32 mature light chain variable domain protein sequence 93 SCT-Ca33 mature light chain variable domain protein sequence 94 SCT-Ca34 mature light chain variable domain protein sequence 95 SCT-Ca35 mature light chain variable domain protein sequence 96 SCT-Ca36 mature light chain variable domain protein sequence 97 SCT-Ca37 mature light chain variable domain protein sequence 98 SCT-Ca38 mature light chain variable domain protein sequence 99 SCT-Ca39 mature light chain variable domain protein sequence 100 SCT-Ca40 mature light chain variable domain protein sequence 101 SCT-Ca41 mature light chain variable domain protein sequence 102 SCT-Ca42 mature light chain variable domain protein sequence 103 SCT-Ca43 mature light chain variable domain protein sequence 103 SCT-Ca44 mature light chain variable domain protein sequence 103 SCT-Ca45 mature light chain variable domain protein sequence 104 SCT-Ca46 mature light chain variable domain protein sequence 105 SCT-Ca47 mature light chain variable domain protein sequence 106 SCT-Ca48 mature light chain variable domain protein sequence 107 SCT
  • FIG. 1. shows dose-dependent blockade of anti-human LAG-3 antibody interfering the interaction between LAG-3 and its ligand FGL1 with LAG-3 coated on ELISA plates.
  • FIG. 2. shows dose-dependent blockade of anti-human LAG-3 antibody interfering the interaction between LAG-3 and its ligand FGL1 with FGL1 coated on ELISA plates.
  • FIG. 3. shows interaction analysis of human LAG-3 with anti-LAG-3 antibody (SCT-Ca01) in a surface plasmon resonance assay on Biacore T200.
  • the disclosure provides antibodies that bind to LAG-3.
  • the disclosure provides an isolated antibody that binds to SEQ ID NO: 1.
  • the disclosure provides antibodies that bind specifically to SEQ ID NO: 1.
  • the term “antibody” as used herein includes both full-length immunoglobulins and antibody fragments that bind to LAG-3.
  • the antibodies can be, e.g., a monoclonal, polyclonal, chimeric, humanized, or single chain antibody.
  • the terms “antigen binding fragment, ” “fragment, ” and “antibody fragment” are used interchangeably to refer to any fragment that comprises a portion of a full-length antibody, generally at least the antigen binding portion or the variable region thereof.
  • Examples of antibody fragments include, but are not limited to, diabodies, single-chain antibody molecules, multi-specific antibodies, Fab, Fab’, F (ab') 2, Fv or scFv.
  • terapéuticaally effective is intended to qualify the amount of active ingredients used in the treatment of a disease or disorder or on the effecting of a clinical endpoint.
  • terapéuticaally acceptable refers to those compounds (or salts, prodrugs, tautomers, zwitterionic forms, etc. ) which are suitable for use in contact with the tissues of patients without undue toxicity, irritation, and allergic response, are commensurate with a reasonable benefit/risk ratio, and are effective for their intended use.
  • treating, ” “treatment, ” and the like, as used herein, mean ameliorating a disease, so as to reduce, ameliorate, or eliminate its cause, its progression, its severity, or one or more of its symptoms, or otherwise beneficially alter the disease in a subject.
  • Reference to “treating, ” or “treatment” of a patient is intended to include prophylaxis.
  • Treatment may also be preemptive in nature, i.e., it may include prevention of disease in a subject exposed to or at risk for the disease. Prevention of a disease may involve complete protection from disease, for example as in the case of prevention of infection with a pathogen or may involve prevention of disease progression.
  • prevention of a disease may not mean complete foreclosure of any effect related to the diseases at any level, but instead may mean prevention of the symptoms of a disease to a clinically significant or detectable level. Prevention of diseases may also mean prevention of progression of a disease to a later stage of the disease.
  • subject and “patient” are used interchangeably herein to mean all mammals including humans. Examples of subjects include, but are not limited to, humans, monkeys, dogs, cats, horses, cows, goats, sheep, pigs, and rabbits. In one embodiment, the subject or patient is a human.
  • the cancer treated with the anti-LAG-3 antibody molecule includes but is not limited to, a solid tumor, a hematological cancer (e.g., leukemia, lymphoma, myeloma) , and a metastatic lesion thereof.
  • the cancer is a solid tumor.
  • solid tumors include malignancies, e.g., sarcomas and carcinomas (e.g., adenocarcinomas) , of the various organ systems, such as those affecting lung, breast, lymphoid, gastrointestinal or colorectal, genitals and genitourinary tract (e.g., renal, urothelial, bladder cells) , pharynx, CNS (e.g., brain, neural or glial cells) , skin (e.g., melanoma) , head and neck (e.g., head and neck squamous cell carcinoma (HNCC) ) , and pancreas.
  • malignancies e.g., sarcomas and carcinomas (e.g., adenocarcinomas)
  • carcinomas e.g., adenocarcinomas
  • the various organ systems such as those affecting lung, breast, lymphoid, gastrointestinal or colorectal, genital
  • melanoma colon cancers, gastric cancer, rectal cancer, renal-cell carcinoma, breast cancer (e.g., a breast cancer that does not express one, two or all of estrogen receptor, progesterone receptor, or Her2/neu, e.g., a triple negative breast cancer) , liver cancer, a lung cancer (e.g., a non-small cell lung cancer (NSCLC) (e.g., a NSCLC with squamous and/or non-squamous histology) or small cell lung cancer) , prostate cancer, cancer of head or neck (e.g., HPV+ squamous cell carcinoma) , cancer of the small intestine and cancer of the esophagus.
  • NSCLC non-small cell lung cancer
  • head or neck e.g., HPV+ squamous cell carcinoma
  • hematological cancer examples include, but is not limited to, leukemia (e.g., a myeloid leukemia, lymphoid leukemia, or chronic lymphocytic leukemia (CLL) ) , lymphoma (e.g., Hogdkin lymphoma (HL) , non-Hogdkin lymphoma (NHL) , Diffuse large B-cell lymphoma (DLBCL) , T-cell lymphoma, or mantle cell lymphoma (MCL) ) , and myeloma, e.g., multiple myeloma.
  • the cancer may be at an early, intermediate, late stage or metastatic cancer.
  • LAG-3 generally known in the art, please see U.S. Pat. No. 9,207,238, which is incorporated in its entirety by reference.
  • the provided antibodies may be used to diagnose, treat, or monitor LAG-3-expressing cancer and its progression, regression, or stability; to determine whether or not a patient should be treated for cancer; or to determine whether or not a subject is afflicted with LAG-3-expressing cancer and thus may be amenable to treatment with a LAG-3-specific anti-cancer therapeutic.
  • the antibodies or fragments thereof described herein may be used for various in vitro molecular-biology applications such as, for example, enzyme-linked immunosorbent assays (ELISA) , Western blots, immunohistochemistry, immunocytochemistry, flow cytometry and fluorescence-activated cell sorting (FACS) , immunoprecipitation, and/or enzyme-linked immunospot assays (ELISPOT) .
  • ELISA enzyme-linked immunosorbent assays
  • FACS fluorescence-activated cell sorting
  • ELISPOT enzyme-linked immunospot assays
  • the antibodies or fragments thereof may be packaged in kits with or without additional reagents known to those of skill in the art for practicing any of the molecular biology techniques disclosed above.
  • the disclosure provides the antibodies SCT-Ca01, SCT-Ca02, SCT-Ca03, SCT-Ca04, SCT-Ca05, SCT-Ca06, SCT-Ca07, SCT-Ca08, SCT-Ca09, SCT-Ca10, SCT-Ca11, SCT-Ca12, SCT-Ca13, SCT-Ca14, SCT-Ca15, SCT-Ca16, SCT-Ca17, SCT-Ca18, SCT-Ca19, SCT-Ca20, SCT-Ca21, SCT-Ca22, SCT-Ca23, SCT-Ca24, SCT-Ca25, SCT-Ca26, SCT-Ca27, SCT-Ca28, SCT-Ca29, SCT-Ca30, SCT-Ca31, SCT-Ca32, SCT-Ca33, SCT-Ca34, SCT-Ca35, SCT-C
  • recombinant anti-LAG-3 antibodies such as chimeric and humanized monoclonal antibodies, comprising both human and non-human portions, which can be made using standard recombinant DNA techniques, are within the scope of the disclosure.
  • Such chimeric and humanized monoclonal antibodies can be produced by recombinant DNA techniques such as, for example, the methods described in U.S. Pat. No. 7,112,421; Better et al. (1988) Science 240: 1041-1043; or Liu et al. (1987) Proc. Natl. Acad. Sci. USA 84: 3439-3443.
  • Antibody Variable Domain Sequence such as, for example, the methods described in U.S. Pat. No. 7,112,421; Better et al. (1988) Science 240: 1041-1043; or Liu et al. (1987) Proc. Natl. Acad. Sci. USA 84: 3439-3443.
  • the antibodies of the disclosure may comprise the heavy chain variable domain sequences of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, S
  • the heavy chain variable domain sequences may consist essentially of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO
  • the antibodies of the disclosure may comprise the light chain variable domain sequences of SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO:
  • the light chain variable domain sequences may consist essentially of SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, S
  • variable domain sequence comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to a sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 31, S
  • variable domain sequence comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to a sequence selected from NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO:
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 4 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 67.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 4 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 68.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 5 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 69.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 6 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 70.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 7 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 71.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 8 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 72.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 9 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 73.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 9 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 74.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 10 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 75.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 11 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 76.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 12 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 77.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 12 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 78.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 13 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 79.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 14 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 80.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 15 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 81.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 16 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 79.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 17 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 18 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 18 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 83.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 19 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 84.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 20 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 85.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 21 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 22 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 23 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 23 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 84.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 24 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 86.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 25 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 87.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 26 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 88.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 27 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 89.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 28 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 90.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 29 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 91.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 29 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 92.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 30 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 93.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 31 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 94.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 32 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 95.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 33 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 96.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 34 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 97.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 35 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 98.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 36 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 99.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 37 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 100.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 38 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 101.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 39 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 102.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 40 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 103.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 41 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 103.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 42 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 103.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 43 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 104.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 44 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 105.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 45 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 106.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 46 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 107.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 47 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 108.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 48 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 109.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 49 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 110.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 50 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 111.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 51 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 112.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 52 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 113.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 53 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 114.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 54 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 115.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 55 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 115.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 56 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 116.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 57 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 117.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 58 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 118.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 59 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 119.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 59 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 120.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 60 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 121.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 61 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 122.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 62 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 123.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 63 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 124.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 64 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 125.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 65 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 117.
  • the disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 66 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 113.
  • Table 1 provides a summary of the LAG-3-specific antibodies described herein:
  • compositions comprising the antibodies or antibody fragments of the present disclosure are also contemplated and can be used in the methods disclosed herein.
  • Pharmaceutical compositions can comprise one or more of the antibodies or antibody fragments described herein and a pharmaceutically acceptable carrier or excipient.
  • the carrier or excipient may facilitate administration, it should not itself induce the production of antibodies harmful to the subject or individual receiving the composition; nor should it be toxic.
  • Suitable carriers may be large, slowly metabolized macromolecules such as proteins, polypeptides, liposomes, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers and inactive virus particles, and are known to one of skill in the art.
  • the antibodies or an antigen binding fragments described herein, or the pharmaceutical compositions disclosed herein may be administered by any number of routes including, but not limited to, oral, intravenous, intramuscular, intra-arterial, intramedullary, intraperitoneal, intrathecal, intraventricular, transdermal, transcutaneous, topical, subcutaneous, intranasal, enteral, sublingual, intravaginal or rectal routes.
  • the therapeutic compositions may be prepared as injectables, either as liquid solutions or suspensions. Solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection may also be prepared.
  • the antibody, or an antigen binding fragment thereof, or pharmaceutical composition is administered intravenously. In another embodiment, the antibody, or an antigen binding fragment thereof, or pharmaceutical composition is administered by intravenous infusion.
  • Direct delivery of the compositions will generally be accomplished by injection, subcutaneously, intraperitoneally, intravenously or intramuscularly, or delivered to the interstitial space of a tissue.
  • the compositions can also be administered into a lesion.
  • Dosage treatment may be a single dose schedule or a multiple dose schedule.
  • Known antibody-based pharmaceuticals provide guidance relating to frequency of administration e.g., whether a pharmaceutical should be delivered daily, weekly, monthly, etc. Frequency and dosage may also depend on the severity of symptoms.
  • the active ingredient in the composition will be an antibody molecule, an antibody fragment or variants and derivatives thereof. As such, it will be susceptible to degradation in the gastrointestinal tract. Thus, if the composition is to be administered by a route using the gastrointestinal tract, the composition will need to contain agents which protect the antibody from degradation, but which release the antibody once it has been absorbed from the gastrointestinal tract.
  • the methods of the present invention can use an antibody, or an antigen binding fragment thereof, as described above, alone or in combination with other pharmaceutically active compounds, to treat conditions such as those disclosed hereinabove.
  • the additional pharmaceutically active compound (s) can be administered simultaneously (either in the same dosage form or in separate dosage forms) or sequentially.
  • the present invention comprises methods for treating a condition by administering to the subject a therapeutically effective amount of an antibody, or an antigen binding fragment thereof, of the present invention and one or more additional pharmaceutically active compounds.
  • the antibody, or an antigen binding fragment thereof, of the present invention is used in combination with existing LAG-3-related disease therapies.
  • Example 1 Isolation of murine anti-human LAG-3 antibodies immunization, &single cell suspension generation
  • Recombinant human LAG-3 (huLAG-3-His tagged, catalog no: LA3-H5222, ACROBiosystems, Beijing, China, SEQ ID NO: 2) was used to immunize young C57/BL6 mice each with 80 ⁇ g of the protein in Sigma Adjuvant (Sigma-Aldrich, St. Louis, MO) over a period of 35 days using a rapid immunization protocol of Antibody Solutions (Sunnyvale, CA) .
  • the lymph nodes were harvested on day 35. Single cell suspension of the lymph node was generated, and the suspension was filtered through a 70 ⁇ m mesh (BD Bioscience) to remove clumps.
  • the filtered lymphocyte suspension was enriched for plasma cells actively secreting IgGs using a kit based on cell surface expression of CD138 (Miltenyi, Auburn, CA) .
  • a kit based on cell surface expression of CD138 Molyzed Cell Sorting Kit *Ultra Convenient*.
  • freshly enriched plasma cells were deposited on a PDMS device to allow a single cell settled in the microwells on the device.
  • Antibody secreted from each plasma cell was captured on a derivatized microscope slide.
  • Antigen-specific antibody secreting cells were identified by interrogating the antibody capture slide with varying concentrations of fluorescently labeled LAG-3 protein tagged with human Fc (ACROBiosystems, Beijing, China, catalog no: LA3-H5255, SEQ ID NO: 3) . Labeling was done using a kit (AnaSpec, Fremont, CA, AS-72046, AnaTag TM HiLyte TM Fluor 555 Microscale Protein Labeling Kit *Ultra Conveni
  • oligonucleotide microarray (Agilent, Santa Clara, CA) . This procedure was previously described in U.S. Patent No. 9,328,172.
  • the custom oligonucleotide microarray is prepared such that each feature contains not only a unique tag specifying its coordinate but also capture probes for all subclasses (1, 2a, 2b, and 3) of murine IgG heavy chain, murine Ig kappa light chain.
  • Captured mRNA on the custom microarray was further processed to synthesize cDNA of each mRNA incorporating the unique tag originally on each feature.
  • the cDNA is then amplify using a Taq polymerase (Promega, Madison, WI) and appropriate set of primers to allow amplification of the following genes: variable domain of IgG heavy chain subclasses and variable domain of Ig kappa light chain. Though now released from cells, these fragments of each gene are now labeled with the unique tag from the custom oligonucleotide microarray manifesting their originating locations.
  • the amplicons were further manipulated to have appropriate sequence attached at both ends to enable sequencing on an Illumina MiSeq instrument using 2 x 250 bp chemistry at SeqMatic LLC (Fremont, CA) .
  • V H or V L sequence containing the identified CDR3s was identified and the associated sequencing reads were assembled into full-length cDNA sequences for V H and V L .
  • the pair of full-length cDNA was correlated with the affinity measurements associated with each of the antigen-specific antibody spot.
  • a select subset of paired V H and V L anti-LAG-3 antibody sequences was used to synthesize corresponding gene fragments by a service provider according to the known art.
  • the resulting gene fragments were cloned into an appropriate plasmid vector and transfected into an appropriate mammalian host, such as HEK293, for recombinant expression to produce an antibody preparation in full-IgG format.
  • the antibody preparations were characterized by measurements at OD280 to assess the amount produced and by gel electrophoresis on PAGE to assess the size of the antibody chains produced.
  • the dose-dependent blockade activity of each of the recombinant antibodies to interfere with the interaction between LAG-3 and its recently discovered ligand fibrinogen-like protein 1 (FGL1) was assessed using ELISA in two different configurations as explained below.
  • First set of experiment involved coating the recombinant human LAG-3 (His tag) onto ELISA plates, followed by adding serially diluted concentrations of each anti-LAG-3 antibody preparation to bind to the coated LAG-3. Then a fixed amount of biotinylated FGL1 is added to be detected by HRP-conjugated streptavidin.
  • LAG-3 antibody upon binding to LAG-3, to block FGL1 interaction with LAG-3 is assessed by the amount of FGL1 bound to LAG-3 (FIG. 1. with IC50 shown in Table 2) .
  • Second set of experiments involved flipping the two analytes, LAG-3 and FGL1, where FGL1 is now coated onto the ELISA plates and assess how much of the LAG-3 bound by anti-LAG-3 antibody can bind to the immobilized FGL1. Analogously, serially diluted concentrations of each anti-LAG-3 antibody preparation was first bound to biotinylated LAG-3 and the complex was allowed to interact with the immobilized FGL1 on the ELISA plates.
  • the amount of LAG-3 was detected by HRP-conjugated streptavidin (FIG. 2. With IC50 shown in Table 2) .
  • the binding affinity of one of the anti-LAG-3 antibodies (SCT-Ca01) showing significant blockade activity in both configurations was measured on a surface plasmon resonance (SPR) instrument, such as a Biacore T200 TM , against human LAG-3.
  • SPR surface plasmon resonance
  • CDR complementarity-determining region
  • Baixeras E., Huard, B., Miossec, C., Jitsukawa, S., Martin, M., Hercend, T., Auffray, C., Triebel, F., and Piatier-Tonneau, D. (1992) . Characterization of the lymphocyte activation gene 3-encoded protein. A new ligand for human leukocyte antigen class II antigens. J. Exp. Med. 176, 327–337.
  • LAG-3 expression defines a subset of CD4 (+) CD25 (high) Foxp3 (+) regulatory T cells that are expanded at tumor sites. J. Immunol. Baltim. Md 1950 184, 6545–6551.
  • Lymphocyte-activation gene 3/major histocompatibility complex class II interaction modulates the antigenic response of CD4+ T lymphocytes. Eur. J. Immunol. 24, 3216–3221.
  • LAP lymphocyte activation gene-3 (LAG-3) -associated protein that binds to a repeated EP motif in the intracellular region of LAG-3, may participate in the down-regulation of the CD3/TCR activation pathway. Eur. J. Immunol. 31, 2885–2891.
  • Fibrinogen-like Protein 1 Is a Major Immune Inhibitory Ligand of LAG-3. Cell 176, 334-347. e12.

Abstract

Provided are antibodies that recognize lymphocyte activation gene-3 protein (LAG-3).

Description

ANTI-LAG-3 ANTIBODIES
CROSS-REFERENCE TO RELATED APPLICATIONS
This application claims priority to U.S. Provisional Application No. 62/839,134 filed 26 April 2019, which is incorporated herein by reference in its entirety for all purposes.
INCORPORATION OF SEQUENCE LISTING
The instant application contains a Sequence Listing which is hereby incorporated by reference in its entirety. The ASCII copy, created on April 26, 2020, is named SCT0006-401-PC_Sequence_Listing_ST. txt, and is 165 KB in size.
BACKGROUND
Lymphocyte Activation Gene-3, or LAG-3 (also known as CD223) , is a member of the immunoglobulin supergene family, and is expressed on activated T cells (Huard et al., 1994a) , NK cells (Triebel et al., 1990) , regulatory T cells (Huang et al., 2004; Camisaschi et al., 2010; Gagliani et al., 2013) , and plasmacytoid dendritic cells (DCs) (Workman et al., 2009) . LAG-3 is a membrane protein encoded by a gene located on chromosome 12 and is structurally and genetically related to CD4.
Similar to CD4, LAG-3 can interact with MHC class II molecules on the cell surface (Baixeras et al., 1992; Huard et al., 1996) . It has been suggested that the direct binding of LAG-3 to MHC class II plays a role in down-regulating antigen-dependent stimulation of CD4+T lymphocytes (Huard et al., 1994b) and LAG-3 blockade has also been shown to reinvigorate CD8+ lymphocytes in both tumor or self-antigen (Grosso et al., 2007; Blackburn et al., 2009) . Further, the intra-cytoplasmic region of LAG-3 can interact with LAP (LAG-3-associated protein) , which is a signal transduction molecule involved in the downregulation of the CD3/TCR activation pathway (Iouzalen et al., 2001) . Moreover, CD4+CD25+ regulatory T cells (Treg) have been shown to express LAG-3 upon activation, which contributes to the suppressor activity of Treg cells (Huang et al., 2004) . LAG-3 can also negatively regulate T cell homeostasis by Treg cells in both T cell-dependent and independent mechanisms (Workman and Vignali, 2005) .
Given the importance of LAG-3 in downregulating an immune response, the need exists for developing novel agents that modulate its activity to activate the immune system. The anti-LAG-3 antibody molecules disclosed herein can be used to treat, prevent and/or diagnose cancerous or malignant disorders (e.g., cancers such melanoma, e.g., advanced stage melanoma; pancreatic cancer, e.g., advanced pancreatic cancer; solid tumors; breast cancer, e.g., metastatic breast carcinoma; renal cell carcinoma, e.g., advanced or metastatic renal cell carcinoma (MRCC) or clear cell renal cell carcinoma) , as well as potentially infectious diseases (e.g., hepatitis, e.g., hepatitis B; influenza) . Thus, methods for detecting LAG-3, as well as methods for treating various disorders, including cancer and potentially infectious diseases using the anti-LAG-3 antibody molecules, alone or in combination, are disclosed herein.
BRIEF DESCRIPTION OF THE SEQUENCES
TABLE 1
SEQ ID NO. Description of sequence
1 Human LAG-3 protein sequence
2 Human LAG-3 His tagged protein sequence
3 Human LAG-3 Human Fc tagged protein sequence
4 SCT-Ca01 mature heavy chain variable domain protein sequence
4 SCT-Ca02 mature heavy chain variable domain protein sequence
5 SCT-Ca03 mature heavy chain variable domain protein sequence
6 SCT-Ca04 mature heavy chain variable domain protein sequence
7 SCT-Ca05 mature heavy chain variable domain protein sequence
8 SCT-Ca06 mature heavy chain variable domain protein sequence
9 SCT-Ca07 mature heavy chain variable domain protein sequence
9 SCT-Ca08 mature heavy chain variable domain protein sequence
10 SCT-Ca09 mature heavy chain variable domain protein sequence
11 SCT-Ca10 mature heavy chain variable domain protein sequence
12 SCT-Ca11 mature heavy chain variable domain protein sequence
12 SCT-Ca12 mature heavy chain variable domain protein sequence
13 SCT-Ca13 mature heavy chain variable domain protein sequence
14 SCT-Ca14 mature heavy chain variable domain protein sequence
15 SCT-Ca15 mature heavy chain variable domain protein sequence
16 SCT-Ca16 mature heavy chain variable domain protein sequence
17 SCT-Ca17 mature heavy chain variable domain protein sequence
4 SCT-Ca01 mature heavy chain variable domain protein sequence
18 SCT-Ca18 mature heavy chain variable domain protein sequence
18 SCT-Ca19 mature heavy chain variable domain protein sequence
19 SCT-Ca20 mature heavy chain variable domain protein sequence
20 SCT-Ca21 mature heavy chain variable domain protein sequence
21 SCT-Ca22 mature heavy chain variable domain protein sequence
22 SCT-Ca23 mature heavy chain variable domain protein sequence
23 SCT-Ca24 mature heavy chain variable domain protein sequence
23 SCT-Ca25 mature heavy chain variable domain protein sequence
24 SCT-Ca26 mature heavy chain variable domain protein sequence
25 SCT-Ca27 mature heavy chain variable domain protein sequence
26 SCT-Ca28 mature heavy chain variable domain protein sequence
27 SCT-Ca29 mature heavy chain variable domain protein sequence
28 SCT-Ca30 mature heavy chain variable domain protein sequence
29 SCT-Ca31 mature heavy chain variable domain protein sequence
29 SCT-Ca32 mature heavy chain variable domain protein sequence
30 SCT-Ca33 mature heavy chain variable domain protein sequence
31 SCT-Ca34 mature heavy chain variable domain protein sequence
32 SCT-Ca35 mature heavy chain variable domain protein sequence
33 SCT-Ca36 mature heavy chain variable domain protein sequence
34 SCT-Ca37 mature heavy chain variable domain protein sequence
35 SCT-Ca38 mature heavy chain variable domain protein sequence
36 SCT-Ca39 mature heavy chain variable domain protein sequence
37 SCT-Ca40 mature heavy chain variable domain protein sequence
38 SCT-Ca41 mature heavy chain variable domain protein sequence
39 SCT-Ca42 mature heavy chain variable domain protein sequence
40 SCT-Ca43 mature heavy chain variable domain protein sequence
41 SCT-Ca44 mature heavy chain variable domain protein sequence
42 SCT-Ca45 mature heavy chain variable domain protein sequence
43 SCT-Ca46 mature heavy chain variable domain protein sequence
44 SCT-Ca47 mature heavy chain variable domain protein sequence
45 SCT-Ca48 mature heavy chain variable domain protein sequence
46 SCT-Ca49 mature heavy chain variable domain protein sequence
47 SCT-Ca50 mature heavy chain variable domain protein sequence
48 SCT-Ca51 mature heavy chain variable domain protein sequence
49 SCT-Ca52 mature heavy chain variable domain protein sequence
50 SCT-Ca53 mature heavy chain variable domain protein sequence
51 SCT-Ca54 mature heavy chain variable domain protein sequence
52 SCT-Ca55 mature heavy chain variable domain protein sequence
53 SCT-Ca56 mature heavy chain variable domain protein sequence
54 SCT-Ca57 mature heavy chain variable domain protein sequence
55 SCT-Ca58 mature heavy chain variable domain protein sequence
56 SCT-Ca59 mature heavy chain variable domain protein sequence
57 SCT-Ca60 mature heavy chain variable domain protein sequence
58 SCT-Ca61 mature heavy chain variable domain protein sequence
59 SCT-Ca62 mature heavy chain variable domain protein sequence
59 SCT-Ca63 mature heavy chain variable domain protein sequence
60 SCT-Ca64 mature heavy chain variable domain protein sequence
61 SCT-Ca65 mature heavy chain variable domain protein sequence
62 SCT-Ca66 mature heavy chain variable domain protein sequence
63 SCT-Ca67 mature heavy chain variable domain protein sequence
64 SCT-Ca68 mature heavy chain variable domain protein sequence
65 SCT-Ca69 mature heavy chain variable domain protein sequence
66 SCT-Ca70 mature heavy chain variable domain protein sequence
67 SCT-Ca01 mature light chain variable domain protein sequence
68 SCT-Ca02 mature light chain variable domain protein sequence
69 SCT-Ca03 mature light chain variable domain protein sequence
70 SCT-Ca04 mature light chain variable domain protein sequence
71 SCT-Ca05 mature light chain variable domain protein sequence
72 SCT-Ca06 mature light chain variable domain protein sequence
73 SCT-Ca07 mature light chain variable domain protein sequence
74 SCT-Ca08 mature light chain variable domain protein sequence
75 SCT-Ca09 mature light chain variable domain protein sequence
76 SCT-Ca10 mature light chain variable domain protein sequence
77 SCT-Ca11 mature light chain variable domain protein sequence
78 SCT-Ca12 mature light chain variable domain protein sequence
79 SCT-Ca13 mature light chain variable domain protein sequence
79 SCT-Ca16 mature light chain variable domain protein sequence
80 SCT-Ca14 mature light chain variable domain protein sequence
81 SCT-Ca15 mature light chain variable domain protein sequence
82 SCT-Ca17 mature light chain variable domain protein sequence
82 SCT-Ca18 mature light chain variable domain protein sequence
82 SCT-Ca22 mature light chain variable domain protein sequence
82 SCT-Ca23 mature light chain variable domain protein sequence
82 SCT-Ca24 mature light chain variable domain protein sequence
83 SCT-Ca19 mature light chain variable domain protein sequence
84 SCT-Ca20 mature light chain variable domain protein sequence
84 SCT-Ca25 mature light chain variable domain protein sequence
85 SCT-Ca21 mature light chain variable domain protein sequence
86 SCT-Ca26 mature light chain variable domain protein sequence
87 SCT-Ca27 mature light chain variable domain protein sequence
88 SCT-Ca28 mature light chain variable domain protein sequence
89 SCT-Ca29 mature light chain variable domain protein sequence
90 SCT-Ca30 mature light chain variable domain protein sequence
91 SCT-Ca31 mature light chain variable domain protein sequence
92 SCT-Ca32 mature light chain variable domain protein sequence
93 SCT-Ca33 mature light chain variable domain protein sequence
94 SCT-Ca34 mature light chain variable domain protein sequence
95 SCT-Ca35 mature light chain variable domain protein sequence
96 SCT-Ca36 mature light chain variable domain protein sequence
97 SCT-Ca37 mature light chain variable domain protein sequence
98 SCT-Ca38 mature light chain variable domain protein sequence
99 SCT-Ca39 mature light chain variable domain protein sequence
100 SCT-Ca40 mature light chain variable domain protein sequence
101 SCT-Ca41 mature light chain variable domain protein sequence
102 SCT-Ca42 mature light chain variable domain protein sequence
103 SCT-Ca43 mature light chain variable domain protein sequence
103 SCT-Ca44 mature light chain variable domain protein sequence
103 SCT-Ca45 mature light chain variable domain protein sequence
104 SCT-Ca46 mature light chain variable domain protein sequence
105 SCT-Ca47 mature light chain variable domain protein sequence
106 SCT-Ca48 mature light chain variable domain protein sequence
107 SCT-Ca49 mature light chain variable domain protein sequence
108 SCT-Ca50 mature light chain variable domain protein sequence
109 SCT-Ca51 mature light chain variable domain protein sequence
110 SCT-Ca52 mature light chain variable domain protein sequence
111 SCT-Ca53 mature light chain variable domain protein sequence
112 SCT-Ca54 mature light chain variable domain protein sequence
113 SCT-Ca55 mature light chain variable domain protein sequence
113 SCT-Ca70 mature light chain variable domain protein sequence
114 SCT-Ca56 mature light chain variable domain protein sequence
115 SCT-Ca57 mature light chain variable domain protein sequence
115 SCT-Ca58 mature light chain variable domain protein sequence
116 SCT-Ca59 mature light chain variable domain protein sequence
117 SCT-Ca60 mature light chain variable domain protein sequence
117 SCT-Ca69 mature light chain variable domain protein sequence
118 SCT-Ca61 mature light chain variable domain protein sequence
119 SCT-Ca62 mature light chain variable domain protein sequence
120 SCT-Ca63 mature light chain variable domain protein sequence
121 SCT-Ca64 mature light chain variable domain protein sequence
122 SCT-Ca65 mature light chain variable domain protein sequence
123 SCT-Ca66 mature light chain variable domain protein sequence
124 SCT-Ca67 mature light chain variable domain protein sequence
125 SCT-Ca68 mature light chain variable domain protein sequence
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1. shows dose-dependent blockade of anti-human LAG-3 antibody interfering the interaction between LAG-3 and its ligand FGL1 with LAG-3 coated on ELISA plates.
FIG. 2. shows dose-dependent blockade of anti-human LAG-3 antibody interfering the interaction between LAG-3 and its ligand FGL1 with FGL1 coated on ELISA plates.
FIG. 3. shows interaction analysis of human LAG-3 with anti-LAG-3 antibody (SCT-Ca01) in a surface plasmon resonance assay on Biacore T200.
DETAILED DESCRIPTION
The disclosure provides antibodies that bind to LAG-3. In one embodiment, the disclosure provides an isolated antibody that binds to SEQ ID NO: 1.
Antibodies
The disclosure provides antibodies that bind specifically to SEQ ID NO: 1. The term “antibody” as used herein, includes both full-length immunoglobulins and antibody fragments that bind to LAG-3. The antibodies can be, e.g., a monoclonal, polyclonal, chimeric, humanized, or single chain antibody. As used herein, the terms “antigen binding fragment, ” “fragment, ” and “antibody fragment” are used interchangeably to refer to any fragment that comprises a portion of a full-length antibody, generally at least the antigen binding portion or the variable region thereof. Examples of antibody fragments include, but are not limited to, diabodies, single-chain antibody molecules, multi-specific antibodies, Fab, Fab’, F (ab') 2, Fv or scFv.
The phrase “therapeutically effective” is intended to qualify the amount of active ingredients used in the treatment of a disease or disorder or on the effecting of a clinical endpoint.
The term “therapeutically acceptable” refers to those compounds (or salts, prodrugs, tautomers, zwitterionic forms, etc. ) which are suitable for use in contact with the tissues of patients without undue toxicity, irritation, and allergic response, are commensurate with a reasonable benefit/risk ratio, and are effective for their intended use.
The terms “treating, ” “treatment, ” and the like, as used herein, mean ameliorating a disease, so as to reduce, ameliorate, or eliminate its cause, its progression, its severity, or one  or more of its symptoms, or otherwise beneficially alter the disease in a subject. Reference to “treating, ” or “treatment” of a patient is intended to include prophylaxis. Treatment may also be preemptive in nature, i.e., it may include prevention of disease in a subject exposed to or at risk for the disease. Prevention of a disease may involve complete protection from disease, for example as in the case of prevention of infection with a pathogen or may involve prevention of disease progression. For example, prevention of a disease may not mean complete foreclosure of any effect related to the diseases at any level, but instead may mean prevention of the symptoms of a disease to a clinically significant or detectable level. Prevention of diseases may also mean prevention of progression of a disease to a later stage of the disease.
The terms “subject” and “patient” are used interchangeably herein to mean all mammals including humans. Examples of subjects include, but are not limited to, humans, monkeys, dogs, cats, horses, cows, goats, sheep, pigs, and rabbits. In one embodiment, the subject or patient is a human.
In certain embodiments, the cancer treated with the anti-LAG-3 antibody molecule, alone or in combination, includes but is not limited to, a solid tumor, a hematological cancer (e.g., leukemia, lymphoma, myeloma) , and a metastatic lesion thereof. In one embodiment, the cancer is a solid tumor. Examples of solid tumors include malignancies, e.g., sarcomas and carcinomas (e.g., adenocarcinomas) , of the various organ systems, such as those affecting lung, breast, lymphoid, gastrointestinal or colorectal, genitals and genitourinary tract (e.g., renal, urothelial, bladder cells) , pharynx, CNS (e.g., brain, neural or glial cells) , skin (e.g., melanoma) , head and neck (e.g., head and neck squamous cell carcinoma (HNCC) ) , and pancreas. For example, melanoma, colon cancers, gastric cancer, rectal cancer, renal-cell carcinoma, breast cancer (e.g., a breast cancer that does not express one, two or all of estrogen receptor, progesterone receptor, or Her2/neu, e.g., a triple negative breast cancer) , liver cancer, a lung cancer (e.g., a non-small cell lung cancer (NSCLC) (e.g., a NSCLC with squamous and/or non-squamous histology) or small cell lung cancer) , prostate cancer, cancer of head or neck (e.g., HPV+ squamous cell carcinoma) , cancer of the small intestine and cancer of the esophagus. Examples of hematological cancer include, but is not limited to, leukemia (e.g., a myeloid leukemia, lymphoid leukemia, or chronic lymphocytic leukemia (CLL) ) , lymphoma (e.g., Hogdkin lymphoma (HL) , non-Hogdkin lymphoma (NHL) , Diffuse large B-cell lymphoma (DLBCL) , T-cell lymphoma, or mantle cell lymphoma (MCL) ) , and myeloma, e.g., multiple  myeloma. The cancer may be at an early, intermediate, late stage or metastatic cancer. For discovering and developing antibody therapeutics to LAG-3 generally known in the art, please see U.S. Pat. No. 9,207,238, which is incorporated in its entirety by reference.
The provided antibodies may be used to diagnose, treat, or monitor LAG-3-expressing cancer and its progression, regression, or stability; to determine whether or not a patient should be treated for cancer; or to determine whether or not a subject is afflicted with LAG-3-expressing cancer and thus may be amenable to treatment with a LAG-3-specific anti-cancer therapeutic. In some embodiments, the antibodies or fragments thereof described herein may be used for various in vitro molecular-biology applications such as, for example, enzyme-linked immunosorbent assays (ELISA) , Western blots, immunohistochemistry, immunocytochemistry, flow cytometry and fluorescence-activated cell sorting (FACS) , immunoprecipitation, and/or enzyme-linked immunospot assays (ELISPOT) . In some embodiments, the antibodies or fragments thereof may be packaged in kits with or without additional reagents known to those of skill in the art for practicing any of the molecular biology techniques disclosed above.
The disclosure provides the antibodies SCT-Ca01, SCT-Ca02, SCT-Ca03, SCT-Ca04, SCT-Ca05, SCT-Ca06, SCT-Ca07, SCT-Ca08, SCT-Ca09, SCT-Ca10, SCT-Ca11, SCT-Ca12, SCT-Ca13, SCT-Ca14, SCT-Ca15, SCT-Ca16, SCT-Ca17, SCT-Ca18, SCT-Ca19, SCT-Ca20, SCT-Ca21, SCT-Ca22, SCT-Ca23, SCT-Ca24, SCT-Ca25, SCT-Ca26, SCT-Ca27, SCT-Ca28, SCT-Ca29, SCT-Ca30, SCT-Ca31, SCT-Ca32, SCT-Ca33, SCT-Ca34, SCT-Ca35, SCT-Ca36, SCT-Ca37, SCT-Ca38, SCT-Ca39, SCT-Ca40, SCT-Ca41, SCT-Ca42, SCT-Ca43, SCT-Ca44, SCT-Ca45, SCT-Ca46, SCT-Ca47, SCT-Ca48, SCT-Ca49, SCT-Ca50, SCT-Ca51, SCT-Ca52, SCT-Ca53, SCT-Ca54, SCT-Ca55, SCT-Ca56, SCT-Ca57, SCT-Ca58, SCT-Ca59, SCT-Ca60, SCT-Ca61, SCT-Ca62, SCT-Ca63, SCT-Ca64, SCT-Ca65, SCT-Ca66, SCT-Ca67, SCT-Ca68, SCT-Ca69, and SCT-Ca70. Each of these is a murine monoclonal antibody.
Additionally, recombinant anti-LAG-3 antibodies, such as chimeric and humanized monoclonal antibodies, comprising both human and non-human portions, which can be made using standard recombinant DNA techniques, are within the scope of the disclosure. Such chimeric and humanized monoclonal antibodies can be produced by recombinant DNA techniques such as, for example, the methods described in U.S. Pat. No. 7,112,421; Better et al. (1988) Science 240: 1041-1043; or Liu et al. (1987) Proc. Natl. Acad. Sci. USA 84: 3439-3443.  Antibody Variable Domain Sequence
The antibodies of the disclosure may comprise the heavy chain variable domain sequences of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, or SEQ ID NO: 66. The heavy chain variable domain sequences may consist essentially of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, or SEQ ID NO: 66.
The antibodies of the disclosure may comprise the light chain variable domain sequences of SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88,  SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, or SEQ ID NO: 125. The light chain variable domain sequences may consist essentially of SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, or SEQ ID NO: 125.
The disclosure also provides a variable domain sequence comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to a sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60,  SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, or SEQ ID NO: 66. The disclosure also provides a variable domain sequence comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to a sequence selected from NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, or SEQ ID NO: 125. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 4 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 67. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 4 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 68. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 5 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 69. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 6 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 70. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 7 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 71. The disclosure also  provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 8 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 72. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 9 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 73. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 9 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 74. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 10 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 75. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 11 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 76. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 12 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 77. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 12 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 78. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 13 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 79. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 14 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 80. The disclosure also provides antibodies comprising a heavy chain variable domain  sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 15 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 81. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 16 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 79. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 17 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 18 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 18 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 83. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 19 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 84. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 20 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 85. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 21 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 22 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%,  at least 90%, or at least 95%identical to SEQ ID NO: 23 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 82. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 23 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 84. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 24 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 86. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 25 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 87. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 26 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 88. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 27 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 89. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 28 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 90. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 29 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 91. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 29 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 92. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 30 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 93. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 31 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 94. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 32 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 95. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 33 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 96. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 34 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 97. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 35 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 98. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 36 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 99. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 37 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 100. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 38 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 101. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 39 and a light  chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 102. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 40 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 103. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 41 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 103. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 42 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 103. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 43 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 104. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 44 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 105. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 45 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 106. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 46 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 107. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 47 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 108. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 48 and a light chain variable domain sequence that  is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 109. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 49 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 110. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 50 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 111. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 51 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 112. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 52 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 113. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 53 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 114. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 54 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 115. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 55 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 115. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 56 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 116. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 57 and a light chain variable domain sequence that is at least 80%, at least 85%, at least  90%, or at least 95%identical to SEQ ID NO: 117. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 58 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 118. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 59 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 119. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 59 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 120. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 60 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 121. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 61 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 122. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 62 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 123. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 63 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 124. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 64 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 125. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 65 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ  ID NO: 117. The disclosure also provides antibodies comprising a heavy chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 66 and a light chain variable domain sequence that is at least 80%, at least 85%, at least 90%, or at least 95%identical to SEQ ID NO: 113.
Table 1 provides a summary of the LAG-3-specific antibodies described herein:
Pharmaceutical Compositions
Pharmaceutical compositions comprising the antibodies or antibody fragments of the present disclosure are also contemplated and can be used in the methods disclosed herein. Pharmaceutical compositions can comprise one or more of the antibodies or antibody fragments described herein and a pharmaceutically acceptable carrier or excipient. Although the carrier or excipient may facilitate administration, it should not itself induce the production of antibodies harmful to the subject or individual receiving the composition; nor should it be toxic. Suitable carriers may be large, slowly metabolized macromolecules such as proteins, polypeptides, liposomes, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers and inactive virus particles, and are known to one of skill in the art.
The antibodies or an antigen binding fragments described herein, or the pharmaceutical compositions disclosed herein, may be administered by any number of routes including, but not limited to, oral, intravenous, intramuscular, intra-arterial, intramedullary, intraperitoneal, intrathecal, intraventricular, transdermal, transcutaneous, topical, subcutaneous, intranasal, enteral, sublingual, intravaginal or rectal routes. Typically, the therapeutic compositions may be prepared as injectables, either as liquid solutions or suspensions. Solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection may also be prepared.
In one embodiment, the antibody, or an antigen binding fragment thereof, or pharmaceutical composition is administered intravenously. In another embodiment, the antibody, or an antigen binding fragment thereof, or pharmaceutical composition is administered by intravenous infusion.
Direct delivery of the compositions will generally be accomplished by injection, subcutaneously, intraperitoneally, intravenously or intramuscularly, or delivered to the interstitial space of a tissue. The compositions can also be administered into a lesion. Dosage treatment  may be a single dose schedule or a multiple dose schedule. Known antibody-based pharmaceuticals provide guidance relating to frequency of administration e.g., whether a pharmaceutical should be delivered daily, weekly, monthly, etc. Frequency and dosage may also depend on the severity of symptoms.
It will be appreciated that the active ingredient in the composition will be an antibody molecule, an antibody fragment or variants and derivatives thereof. As such, it will be susceptible to degradation in the gastrointestinal tract. Thus, if the composition is to be administered by a route using the gastrointestinal tract, the composition will need to contain agents which protect the antibody from degradation, but which release the antibody once it has been absorbed from the gastrointestinal tract.
The methods of the present invention can use an antibody, or an antigen binding fragment thereof, as described above, alone or in combination with other pharmaceutically active compounds, to treat conditions such as those disclosed hereinabove. The additional pharmaceutically active compound (s) can be administered simultaneously (either in the same dosage form or in separate dosage forms) or sequentially. Accordingly, in one embodiment, the present invention comprises methods for treating a condition by administering to the subject a therapeutically effective amount of an antibody, or an antigen binding fragment thereof, of the present invention and one or more additional pharmaceutically active compounds.
In some embodiments, the antibody, or an antigen binding fragment thereof, of the present invention is used in combination with existing LAG-3-related disease therapies.
EXAMPLES
The following example is put forth so as to provide those of ordinary skill in the art with a complete description of how to make and use the present disclosure, and is not intended to limit the scope of what the inventors regard as their disclosure nor is it intended to represent that the experiment below is all or the only experiment that could be performed. Efforts have been made to ensure accuracy with respect to numbers used (e.g. amounts, temperature, etc. ) but some experimental errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, molecular weight is weight average molecular weight, temperature is in degrees Centigrade, and pressure is at or near atmospheric.
Example 1: Isolation of murine anti-human LAG-3 antibodies immunization, &single cell suspension generation
Recombinant human LAG-3 (huLAG-3-His tagged, catalog no: LA3-H5222, ACROBiosystems, Beijing, China, SEQ ID NO: 2) was used to immunize young C57/BL6 mice each with 80 μg of the protein in Sigma Adjuvant
Figure PCTCN2020087141-appb-000001
 (Sigma-Aldrich, St. Louis, MO) over a period of 35 days using a rapid immunization protocol of Antibody Solutions (Sunnyvale, CA) . The lymph nodes were harvested on day 35. Single cell suspension of the lymph node was generated, and the suspension was filtered through a 70 μm mesh (BD Bioscience) to remove clumps.
Plasma cell isolation, antibody capture, and antigen interrogation
The filtered lymphocyte suspension was enriched for plasma cells actively secreting IgGs using a kit based on cell surface expression of CD138 (Miltenyi, Auburn, CA) . Using a method described in U.S. Patent No. 9,328,172, freshly enriched plasma cells were deposited on a PDMS device to allow a single cell settled in the microwells on the device. Antibody secreted from each plasma cell was captured on a derivatized microscope slide. Antigen-specific antibody secreting cells were identified by interrogating the antibody capture slide with varying concentrations of fluorescently labeled LAG-3 protein tagged with human Fc (ACROBiosystems, Beijing, China, catalog no: LA3-H5255, SEQ ID NO: 3) . Labeling was done using a kit (AnaSpec, Fremont, CA, AS-72046, AnaTag TM HiLyte TM Fluor 555 Microscale Protein Labeling Kit *Ultra Convenient*) .
mRNA capture
After antibody capture, the medium was removed, and replaced with lysis buffer followed by prompt closure of the top of the microwells with a custom oligonucleotide microarray (Agilent, Santa Clara, CA) . This procedure was previously described in U.S. Patent No. 9,328,172. The custom oligonucleotide microarray is prepared such that each feature contains not only a unique tag specifying its coordinate but also capture probes for all subclasses (1, 2a, 2b, and 3) of murine IgG heavy chain, murine Ig kappa light chain.
cDNA synthesis, PCR amplification, and Next Generation Sequencing
Captured mRNA on the custom microarray was further processed to synthesize cDNA of each mRNA incorporating the unique tag originally on each feature. The cDNA is then amplify using a Taq polymerase (Promega, Madison, WI) and appropriate set of primers to  allow amplification of the following genes: variable domain of IgG heavy chain subclasses and variable domain of Ig kappa light chain. Though now released from cells, these fragments of each gene are now labeled with the unique tag from the custom oligonucleotide microarray manifesting their originating locations. The amplicons were further manipulated to have appropriate sequence attached at both ends to enable sequencing on an Illumina MiSeq instrument using 2 x 250 bp chemistry at SeqMatic LLC (Fremont, CA) .
Bioinformatic analysis of images and DNA sequences
Sequencing reads from MiSeq were processed and the embedded tag in each read was identified and converted into coordinates. The coordinates were plotted to yield a synthetic map of the mRNA recovered. Most of the coordinates form clusters that designate the location of the originating cell for the recovered mRNA sequences. Next, CDR3 motif present in each read with the coordinates was identified and collated according to the clusters that matched the location of an antibody spot visualized by an appropriate fluorescently labeled secondary antibody. Identical or nearly identical CDR3s for a given antibody spot were organized and form consistent pair of V H and V L sequences. The remaining part of V H or V L sequence containing the identified CDR3s was identified and the associated sequencing reads were assembled into full-length cDNA sequences for V H and V L. The pair of full-length cDNA was correlated with the affinity measurements associated with each of the antigen-specific antibody spot.
Example 2: Molecular reconstruction and recombinant expression of anti-LAG-3 antibodies
A select subset of paired V H and V L anti-LAG-3 antibody sequences was used to synthesize corresponding gene fragments by a service provider according to the known art. The resulting gene fragments were cloned into an appropriate plasmid vector and transfected into an appropriate mammalian host, such as HEK293, for recombinant expression to produce an antibody preparation in full-IgG format. The antibody preparations were characterized by measurements at OD280 to assess the amount produced and by gel electrophoresis on PAGE to assess the size of the antibody chains produced.
Example 3: Characterization of recombinant anti-LAG-3 antibodies
The dose-dependent blockade activity of each of the recombinant antibodies to interfere with the interaction between LAG-3 and its recently discovered ligand fibrinogen-like protein 1 (FGL1) (Wang et al., 2019) was assessed using ELISA in two different configurations  as explained below. First set of experiment involved coating the recombinant human LAG-3 (His tag) onto ELISA plates, followed by adding serially diluted concentrations of each anti-LAG-3 antibody preparation to bind to the coated LAG-3. Then a fixed amount of biotinylated FGL1 is added to be detected by HRP-conjugated streptavidin. The ability of a LAG-3 antibody, upon binding to LAG-3, to block FGL1 interaction with LAG-3 is assessed by the amount of FGL1 bound to LAG-3 (FIG. 1. with IC50 shown in Table 2) . Second set of experiments involved flipping the two analytes, LAG-3 and FGL1, where FGL1 is now coated onto the ELISA plates and assess how much of the LAG-3 bound by anti-LAG-3 antibody can bind to the immobilized FGL1. Analogously, serially diluted concentrations of each anti-LAG-3 antibody preparation was first bound to biotinylated LAG-3 and the complex was allowed to interact with the immobilized FGL1 on the ELISA plates. The amount of LAG-3 was detected by HRP-conjugated streptavidin (FIG. 2. With IC50 shown in Table 2) . The binding affinity of one of the anti-LAG-3 antibodies (SCT-Ca01) showing significant blockade activity in both configurations was measured on a surface plasmon resonance (SPR) instrument, such as a Biacore T200 TM, against human LAG-3.
Some of the antibodies recovered from the antibody campaign described above are listed herein. The complementarity-determining region (CDR) sequences for the anti-LAG-3 antibodies heavy-chains (e.g., HCDR1) and light-chains (e.g., LCDR1) described herein are depicted in Table 3. The heavy-chain variable domain (V H) and light-chain variable domain (V L) sequences for the anti-LAG-3 antibodies described herein are depicted in Table 4. TABLE 2
Figure PCTCN2020087141-appb-000002
TABLE 3
Figure PCTCN2020087141-appb-000003
Figure PCTCN2020087141-appb-000004
Figure PCTCN2020087141-appb-000005
TABLE 4
Figure PCTCN2020087141-appb-000006
Figure PCTCN2020087141-appb-000007
Figure PCTCN2020087141-appb-000008
Figure PCTCN2020087141-appb-000009
Figure PCTCN2020087141-appb-000010
Figure PCTCN2020087141-appb-000011
The preceding merely illustrates the principles of the disclosure. It will be appreciated that those skilled in the art will be able to devise various arrangements which, although not explicitly described or shown herein, embody the principles of the disclosure and are included within its spirit and scope. Furthermore, all examples and conditional language recited herein are principally intended to aid the reader in understanding the principles of the disclosure and the concepts contributed by the inventors to furthering the art and are to be construed as being without limitation to such specifically recited examples and conditions. Moreover, all statements herein reciting principles, aspects, and embodiments of the disclosure as well as specific examples thereof, are intended to encompass both structural and functional equivalents thereof. Additionally, it is intended that such equivalents include both currently known equivalents and equivalents developed in the future, i.e., any elements developed that perform the same function, regardless of structure. The scope of the present disclosure, therefore, is not intended to be limited to the exemplary embodiments shown and described herein. Rather, the scope and spirit of present disclosure is embodied by the appended claims.
References Cited
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Claims (35)

  1. An antibody that binds human LAG-3 (SEQ ID NO: 1) comprising:
    a) a HCDR1 having the amino acid sequence of SEQ ID NO: 126, a HCDR2 having the amino acid sequence of SEQ ID NO: 127, a HCDR3 having the amino acid sequence of SEQ ID NO: 128, a LCDR1 having the amino acid sequence of SEQ ID NO: 129, a LCDR2 having the amino acid sequence of SEQ ID NO: 130, and a LCDR3 having the amino acid sequence of SEQ ID NO: 131; or
    b) a HCDR1 having the amino acid sequence of SEQ ID NO: 146, a HCDR2 having the amino acid sequence of SEQ ID NO: 147, a HCDR3 having the amino acid sequence of SEQ ID NO: 148, a LCDR1 having the amino acid sequence of SEQ ID NO: 149, a LCDR2 having the amino acid sequence of SEQ ID NO: 150, and a LCDR3 having the amino acid sequence of SEQ ID NO: 151; or
    c) a HCDR1 having the amino acid sequence of SEQ ID NO: 180, a HCDR2 having the amino acid sequence of SEQ ID NO: 156, a HCDR3 having the amino acid sequence of SEQ ID NO: 157, a LCDR1 having the amino acid sequence of SEQ ID NO: 178, a LCDR2 having the amino acid sequence of SEQ ID NO: 130, and a LCDR3 having the amino acid sequence of SEQ ID NO: 174; or
    d) a HCDR1 having the amino acid sequence of SEQ ID NO: 140, a HCDR2 having the amino acid sequence of SEQ ID NO: 182, a HCDR3 having the amino acid sequence of SEQ ID NO: 183, a LCDR1 having the amino acid sequence of SEQ ID NO: 184, a LCDR2 having the amino acid sequence of SEQ ID NO: 185, and a LCDR3 having the amino acid sequence of SEQ ID NO: 186; or
    e) a HCDR1 having the amino acid sequence of SEQ ID NO: 190, a HCDR2 having the amino acid sequence of SEQ ID NO: 141, a HCDR3 having the amino acid sequence of SEQ ID NO: 220, a LCDR1 having the amino acid sequence of SEQ ID NO: 223, a LCDR2 having the amino acid sequence of SEQ ID NO: 150, and a LCDR3 having the amino acid sequence of SEQ ID NO: 222; or
    f) a HCDR1 having the amino acid sequence of SEQ ID NO: 140, a HCDR2 having the amino acid sequence of SEQ ID NO: 232, a HCDR3 having the amino acid sequence of SEQ ID NO: 233, a LCDR1 having the amino acid sequence of SEQ ID NO: 234, a  LCDR2 having the amino acid sequence of SEQ ID NO: 144, and a LCDR3 having the amino acid sequence of SEQ ID NO: 235; or
    g) a HCDR1 having the amino acid sequence of SEQ ID NO: 265, a HCDR2 having the amino acid sequence of SEQ ID NO: 266, a HCDR3 having the amino acid sequence of SEQ ID NO: 267, a LCDR1 having the amino acid sequence of SEQ ID NO: 268, a LCDR2 having the amino acid sequence of SEQ ID NO: 269, and a LCDR3 having the amino acid sequence of SEQ ID NO: 270; or
    h) a HCDR1 having the amino acid sequence of SEQ ID NO: 265, a HCDR2 having the amino acid sequence of SEQ ID NO: 274, a HCDR3 having the amino acid sequence of SEQ ID NO: 267, a LCDR1 having the amino acid sequence of SEQ ID NO: 275, a LCDR2 having the amino acid sequence of SEQ ID NO: 276, and a LCDR3 having the amino acid sequence of SEQ ID NO: 277; or
    i) a HCDR1 having the amino acid sequence of SEQ ID NO: 265, a HCDR2 having the amino acid sequence of SEQ ID NO: 274, a HCDR3 having the amino acid sequence of SEQ ID NO: 267, a LCDR1 having the amino acid sequence of SEQ ID NO: 268, a LCDR2 having the amino acid sequence of SEQ ID NO: 276, and a LCDR3 having the amino acid sequence of SEQ ID NO: 277; or
    j) a HCDR1 having the amino acid sequence of SEQ ID NO: 251, a HCDR2 having the amino acid sequence of SEQ ID NO: 252, a HCDR3 having the amino acid sequence of SEQ ID NO: 292, a LCDR1 having the amino acid sequence of SEQ ID NO: 254, a LCDR2 having the amino acid sequence of SEQ ID NO: 255, and a LCDR3 having the amino acid sequence of SEQ ID NO: 256.
  2. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 126, the HCDR2 has the amino acid sequence of SEQ ID NO: 127, the HCDR3 has the amino acid sequence of SEQ ID NO: 128, the LCDR1 has the amino acid sequence of SEQ ID NO: 129, the LCDR2 has the amino acid sequence of SEQ ID NO: 130, and the LCDR3 has the amino acid sequence of SEQ ID NO: 131.
  3. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 146, the HCDR2 has the amino acid sequence of SEQ ID NO: 147, the HCDR3 has  the amino acid sequence of SEQ ID NO: 148, the LCDR1 has the amino acid sequence of SEQ ID NO: 149, the LCDR2 has the amino acid sequence of SEQ ID NO: 150, and the LCDR3 has the amino acid sequence of SEQ ID NO: 151.
  4. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 180, the HCDR2 has the amino acid sequence of SEQ ID NO: 156, the HCDR3 has the amino acid sequence of SEQ ID NO: 157, the LCDR1 has the amino acid sequence of SEQ ID NO: 178, the LCDR2 has the amino acid sequence of SEQ ID NO: 130, and the LCDR3 has the amino acid sequence of SEQ ID NO: 174.
  5. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 140, the HCDR2 has the amino acid sequence of SEQ ID NO: 182, the HCDR3 has the amino acid sequence of SEQ ID NO: 183, the LCDR1 has the amino acid sequence of SEQ ID NO: 184, the LCDR2 has the amino acid sequence of SEQ ID NO: 185, and the LCDR3 has the amino acid sequence of SEQ ID NO: 186.
  6. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 190, the HCDR2 has the amino acid sequence of SEQ ID NO: 141, the HCDR3 has the amino acid sequence of SEQ ID NO: 220, the LCDR1 has the amino acid sequence of SEQ ID NO: 223, the LCDR2 has the amino acid sequence of SEQ ID NO: 150, and the LCDR3 has the amino acid sequence of SEQ ID NO: 222.
  7. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 140, the HCDR2 has the amino acid sequence of SEQ ID NO: 232, the HCDR3 has the amino acid sequence of SEQ ID NO: 233, the LCDR1 has the amino acid sequence of SEQ ID NO: 234, the LCDR2 has the amino acid sequence of SEQ ID NO: 144, and the LCDR3 has the amino acid sequence of SEQ ID NO: 235.
  8. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 265, the HCDR2 has the amino acid sequence of SEQ ID NO: 266, the HCDR3 has the amino acid sequence of SEQ ID NO: 267, the LCDR1 has the amino acid sequence of  SEQ ID NO: 268, the LCDR2 has the amino acid sequence of SEQ ID NO: 269, and the LCDR3 has the amino acid sequence of SEQ ID NO: 270.
  9. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 265, the HCDR2 has the amino acid sequence of SEQ ID NO: 274, the HCDR3 has the amino acid sequence of SEQ ID NO: 267, the LCDR1 has the amino acid sequence of SEQ ID NO: 275, the LCDR2 has the amino acid sequence of SEQ ID NO: 276, and the LCDR3 has the amino acid sequence of SEQ ID NO: 277.
  10. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 265, the HCDR2 has the amino acid sequence of SEQ ID NO: 274, the HCDR3 has the amino acid sequence of SEQ ID NO: 267, the LCDR1 has the amino acid sequence of SEQ ID NO: 268, the LCDR2 has the amino acid sequence of SEQ ID NO: 276, and the LCDR3 has the amino acid sequence of SEQ ID NO: 277.
  11. The antibody of claim 1, wherein the HCDR1 has the amino acid sequence of SEQ ID NO: 251, the HCDR2 has the amino acid sequence of SEQ ID NO: 252, the HCDR3 has the amino acid sequence of SEQ ID NO: 292, the LCDR1 has the amino acid sequence of SEQ ID NO: 254, the LCDR2 has the amino acid sequence of SEQ ID NO: 255, and the LCDR3 has the amino acid sequence of SEQ ID NO: 256.
  12. An antibody that binds human LAG-3 (SEQ ID NO: 1) comprising:
    a) a V H having the amino acid sequence of SEQ ID NO: 4, and a V L having the amino acid sequence of SEQ ID NO: 67; or
    b) a V H having the amino acid sequence of SEQ ID NO: 8, and a V L having the amino acid sequence of SEQ ID NO: 72; or
    c) a V H having the amino acid sequence of SEQ ID NO: 19, and a V L having the amino acid sequence of SEQ ID NO: 84; or
    d) a V H having the amino acid sequence of SEQ ID NO: 24, and a V L having the amino acid sequence of SEQ ID NO: 86; or
    e) a V H having the amino acid sequence of SEQ ID NO: 35, and a V L having the  amino acid sequence of SEQ ID NO: 98; or
    f) a V H having the amino acid sequence of SEQ ID NO: 46, and a V L having the amino acid sequence of SEQ ID NO: 107; or
    g) a V H having the amino acid sequence of SEQ ID NO: 56, and a V L having the amino acid sequence of SEQ ID NO: 116; or
    h) a V H having the amino acid sequence of SEQ ID NO: 58, and a V L having the amino acid sequence of SEQ ID NO: 118; or
    i) a V H having the amino acid sequence of SEQ ID NO: 59, and a V L having the amino acid sequence of SEQ ID NO: 120; or
    j) a V H having the amino acid sequence of SEQ ID NO: 65, and a V L having the amino acid sequence of SEQ ID NO: 117.
  13. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 4, and the V L has the amino acid sequence of SEQ ID NO: 67.
  14. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 8, and the V L has the amino acid sequence of SEQ ID NO: 72.
  15. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 19, and the V L has the amino acid sequence of SEQ ID NO: 84.
  16. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 24, and the V L has the amino acid sequence of SEQ ID NO: 86.
  17. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 35, and the V L has the amino acid sequence of SEQ ID NO: 98.
  18. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 46, and the V L has the amino acid sequence of SEQ ID NO: 107.
  19. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 56, and the V L has the amino acid sequence of SEQ ID NO: 116.
  20. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 58, and the V L has the amino acid sequence of SEQ ID NO: 118.
  21. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 59, and the V L has the amino acid sequence of SEQ ID NO: 120.
  22. The antibody of claim 12, wherein the V H has the amino acid sequence of SEQ ID NO: 65, and the V L has the amino acid sequence of SEQ ID NO: 117.
  23. An antibody that binds human LAG-3 (SEQ ID NO: 1) comprising:
    a) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 4 and a light-chain comprising the amino acid sequence of SEQ ID NO: 67; or
    b) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 8 and a light-chain comprising the amino acid sequence of SEQ ID NO: 72; or
    c) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 19 and a light-chain comprising the amino acid sequence of SEQ ID NO: 84; or
    d) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 24 and a light-chain comprising the amino acid sequence of SEQ ID NO: 86; or
    e) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 35 and a light-chain comprising the amino acid sequence of SEQ ID NO: 98; or
    f) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 46 and a light-chain comprising the amino acid sequence of SEQ ID NO: 107; or
    g) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 56 and a light-chain comprising the amino acid sequence of SEQ ID NO: 116; or
    h) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 58 and a light-chain comprising the amino acid sequence of SEQ ID NO: 118; or
    i) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 59 and a light-chain comprising the amino acid sequence of SEQ ID NO: 120; or
    j) a heavy-chain comprising the amino acid sequence of SEQ ID NO: 65 and a light-chain comprising the amino acid sequence of SEQ ID NO: 117.
  24. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 4 and the light-chain comprises the amino acid sequence of SEQ ID NO: 67.
  25. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 8 and the light-chain comprises the amino acid sequence of SEQ ID NO: 72.
  26. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 19 and the light-chain comprises the amino acid sequence of SEQ ID NO: 84.
  27. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 24 and the light-chain comprises the amino acid sequence of SEQ ID NO: 86.
  28. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 35 and the light-chain comprises the amino acid sequence of SEQ ID NO: 98.
  29. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 46 and the light-chain comprises the amino acid sequence of SEQ ID NO: 107.
  30. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 56 and the light-chain comprises the amino acid sequence of SEQ ID NO: 116.
  31. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 58 and the light-chain comprises the amino acid sequence of SEQ ID  NO: 118.
  32. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 59 and the light-chain comprises the amino acid sequence of SEQ ID NO: 120.
  33. The antibody of claim 23, wherein the heavy-chain comprises the amino acid sequence of SEQ ID NO: 65 and the light-chain comprises the amino acid sequence of SEQ ID NO: 117.
  34. A pharmaceutical composition comprising the antibody of any of claims 1-33.
  35. A kit comprising the antibody of any of claims 1-34.
PCT/CN2020/087141 2019-04-26 2020-04-27 Anti-lag-3 antibodies WO2020216383A1 (en)

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WO2016028672A1 (en) * 2014-08-19 2016-02-25 Merck Sharp & Dohme Corp. Anti-lag3 antibodies and antigen-binding fragments
WO2017015560A2 (en) * 2015-07-22 2017-01-26 Sorrento Therapeutics, Inc. Antibody therapeutics that bind lag3
WO2017062888A1 (en) * 2015-10-09 2017-04-13 Regeneron Pharmaceuticals, Inc. Anti-lag3 antibodies and uses thereof
WO2017087901A2 (en) * 2015-11-19 2017-05-26 Sutro Biopharma, Inc. Anti-lag3 antibodies, compositions comprising anti-lag3 antibodies and methods of making and using anti-lag3 antibodies
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WO2016028672A1 (en) * 2014-08-19 2016-02-25 Merck Sharp & Dohme Corp. Anti-lag3 antibodies and antigen-binding fragments
WO2017015560A2 (en) * 2015-07-22 2017-01-26 Sorrento Therapeutics, Inc. Antibody therapeutics that bind lag3
WO2017062888A1 (en) * 2015-10-09 2017-04-13 Regeneron Pharmaceuticals, Inc. Anti-lag3 antibodies and uses thereof
WO2017087901A2 (en) * 2015-11-19 2017-05-26 Sutro Biopharma, Inc. Anti-lag3 antibodies, compositions comprising anti-lag3 antibodies and methods of making and using anti-lag3 antibodies
WO2018208868A1 (en) * 2017-05-10 2018-11-15 Smet Pharmaceutical Inc Human monoclonal antibodies against lag3 and uses thereof

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