WO2025062392A1 - Combinaisons de blocage anti-ilt3 - Google Patents

Combinaisons de blocage anti-ilt3 Download PDF

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WO2025062392A1
WO2025062392A1 PCT/IL2024/050926 IL2024050926W WO2025062392A1 WO 2025062392 A1 WO2025062392 A1 WO 2025062392A1 IL 2024050926 W IL2024050926 W IL 2024050926W WO 2025062392 A1 WO2025062392 A1 WO 2025062392A1
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seq
antibody
amino acid
acid sequence
cancer
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Tsuri PERETZ
Yoav PIZEM
Liat IANCOVICI
Motti HAKIM
Ilana MANDEL
Yair SAPIR
Avidor Shulman
Tehila Ben-Moshe
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Biond Biologics Ltd
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Biond Biologics Ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IG], 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IG], 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
    • C07K16/2818—Immunoglobulins [IG], 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 against CD28 or CD152
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IG], 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
    • C07K16/2827—Immunoglobulins [IG], 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 against B7 molecules, e.g. CD80, CD86
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2863—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against receptors for growth factors, growth regulators
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2887—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against CD20
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/32—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from animals or humans against translation products of oncogenes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • A61K2039/507—Comprising a combination of two or more separate antibodies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
    • 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
    • C07K2317/73—Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
    • 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
    • C07K2317/73—Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
    • C07K2317/732—Antibody-dependent cellular cytotoxicity [ADCC]
    • 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

Definitions

  • the present invention is in the field of immune checkpoint inhibition.
  • cancer cells can evade immune surveillance through various mechanisms. Understanding these mechanisms of escape is critical for developing novel cancer therapy.
  • Cancer immunotherapy aims to re-activate the patients’ immune system for the elimination of cancer cells and is currently a highly active area in cancer research with unprecedented results in the clinic.
  • new therapies that will allow for a more substantial increase in patients' mortality and quality of life as a standalone treatment or in combination with other agents.
  • ILT3 Leukocyte immunoglobulin-like receptor subfamily B member 4
  • TAM tumor associated macrophages
  • MDSCs myeloid derived suppressor cells
  • DCtol tolerogenic dendritic cells
  • ILT3 contains 2 extracellular immunoglobulin-like domains and 3 intracellular tyrosine-based inhibitory motifs (ITIMs). The expression of ILT3 in myeloid cells and in a soluble form was reported in various malignancies in association with tumor immune escape, metastasis and poor prognosis.
  • ILT3 signaling on myeloid cells leads to reduced expression of co -stimulatory receptors and differentiation to an immunosuppressive phenotype.
  • ILT3 signaling induces, through unknown ligands, T cell anergy, and differentiation to T regulatory and T suppressor phenotype.
  • ILT3 ligands remained undefined, but recently there is accumulating evidence that ILT3 binds proteins in the tumor microenvironment (TME) and tumor extracellular matrix (ECM). Proteins such as Apolipoprotein E (APOE) and Fibronectin (FN1) are present in the TME and can bind ILT3.
  • APOE Apolipoprotein E
  • FN1 Fibronectin
  • ILT3 blocking antibodies increase tumor cell elimination. This may occur by inhibition of the immunosuppressive effect of tumor resident myeloid cells, thereby inducing proinflammatory phenotype of myeloid cells which support tumor infiltrated T cells’ (TILs’) activation and thereby remodeling the TME from immunosuppressive to proinflammatory TME.
  • TILs tumor infiltrated T cells
  • ILT3 blocking increases the effector activity of various immune cells against the malignant cells.
  • Anti-ILT3 antibodies have been disclosed in International Patent Application WO2006/138739, WO2018/089300, WO2019/099597, and W02020/056077A.
  • the combination of anti-ILT3 antibodies and anti-PD-1 antibodies is disclosed in International Patent Application PCT/IL2023/050426. New superior blockade combinations of anti-ILT3 antibodies with tumor targeting therapeutics are still greatly needed.
  • the present invention provides methods of enhancing immune cell activation against a cancer cell comprising contacting the immune cell with an inhibitor of leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) and contacting the cancer cell with an inhibitor of epidermal growth factor receptor (EGFR), CD47 or Programmed death-ligand 1 (PD-L1).
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • EGFR epidermal growth factor receptor
  • CD47 CD47
  • P-L1 Programmed death-ligand 1
  • Methods of treating cancer by administering an inhibitor of ILT3 and an inhibitor of any one of EGFR, CD47 and PD-L1 or an anticancer agent comprising an IgGl or IgG3 scaffold are also provided.
  • Methods of increasing antibody dependent cell cytotoxicity (ADCC) or antibody dependent cell phagocytosis (ADCP) against a cancer cell and treating cancer comprising blocking binding of ILT3 to at least one of its ligands are also provided and are methods of treating cancer in a subject in need thereof, comprising administering an ILT3 antibody and at least one of a CD20 antibody, a epidermal growth factor receptor (EGFR) antibody, a Receptor tyro sine-protein kinase erbB-2 (HER2) antibody, a CD47 antibody, a Programmed death-ligand 1 (PD-L1) antibody and a Claudin-18 (CLDN18) antibody.
  • ADCC antibody dependent cell cytotoxicity
  • ADCP antibody dependent cell phagocytosis
  • an antibody or antigen binding fragment thereof that binds to leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) for use in treating cancer in a subject in need thereof in combination with an antibody or antigen binding fragment thereof that binds to a factor expressed on the cancer selected from: epidermal growth factor receptor (EGFR), receptor tyrosine-protein kinase erbB-2 (HER2), CD47, and claudin-18 (CLDN18) or with the anti-programmed death-ligand 1 (PD-L1) antibody selected from: Avelumab and an antibody with the CDRs of Atezolizumab or Durvalumab and which induces antibody dependent cellular cytotoxicity (ADCC) or antibody dependent cellular phagocytosis (ADCP).
  • ADCC antibody dependent cellular cytotoxicity
  • ADCP antibody dependent cellular phagocytosis
  • the antibody that binds to ILT3 is an ILT3 blocking or inhibitory antibody.
  • the blocking or inhibitory antibody blocks interaction between ILT3 and an ILT3 ligand.
  • the ligand is selected from Apolipoprotein E (APOE), fibronectin (FN 1) or both.
  • APOE Apolipoprotein E
  • FN 1 fibronectin
  • the ligand is fibronectin.
  • the cancer is a EGFR positive cancer and the ILT3 antibody or antigen binding fragment thereof is for use in combination with the EGFR antibody or antigen binding fragment thereof
  • the cancer is a HER2 positive cancer and the ILT3 antibody or antigen binding fragment thereof is for use in combination with the HER2 antibody or antigen binding fragment thereof
  • the cancer is a CD47 positive cancer and the ILT3 antibody or antigen binding fragment thereof is for use in combination with the CD47 antibody
  • antigen binding fragment thereof or the cancer is a CLDN18 positive cancer and the ILT3 antibody or antigen binding fragment thereof is for use in combination with the CLDN18 antibody or antigen binding fragment thereof or the cancer is a PD-L1 positive cancer and the ILT3 antibody or antigen binding fragment thereof is for use in combination with Avelumab.
  • the antibody or antigen binding fragment thereof that binds to ILT3 comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein at least one of: a.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 1 (GYSFXiGF) wherein Xi is S or T
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 2 (FPSX2GE) wherein X2 is S or N
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 3 (QAFYYFDX3) wherein X3 is S or Y
  • CDR-L 1 comprises the amino acid sequence as set forth in SEQ ID NO: 4 (KSSQSLLSSSNQKNYLA)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6 (
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS)
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 50 (AITFGGGNTYYPDSVKG)
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 51 (HGDGNYDFYAMDY)
  • CDR-L 1 comprises the amino acid sequence as set forth in SEQ ID NO: 52 (KSSQSLLNSGNQKNYLT)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 53 (QNDYSYPLT); c.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS), CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 56 (TISSDGGNTYYTDSVKG), CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 57 (HDGRGALDY), CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 58 (RASQDISNYLN), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 59 (YTSRLHS), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 60 (QQGNTLPWT); and d.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 63 (NSAVH)
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 64 (VIWAGGNTNYNSTLMS)
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 65 (HETYGDSFDY)
  • CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 66 (RSSQSLLDSDGKTYLN)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 67 (LVSKLDS)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 68 (WQGTHFPFT).
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE)
  • SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY)
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE)
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS)
  • c is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 16 FPSNGE
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS)
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); d.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); e.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); or f.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYF
  • the antibody or antigen binding fragment thereof comprises at least one of a. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 34 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 35; b. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 36 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 37; c. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 38 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; d.
  • a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 40 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 41; e. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 42 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 41; f. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 42 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; g. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 43 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; h.
  • a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 45; i. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; j. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 46 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; k. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; and l.
  • the ILT3 antibody or antigen binding fragment thereof is an antibody or antigen binding fragment thereof wherein CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 20, CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 19, CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 17, CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4, CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5, and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6.
  • the antibody is antibody 5E5 or an antibody that competes with antibody 5E5 for binding to ILT3.
  • the ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55.
  • the ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 61 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 62.
  • the ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 69 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 70.
  • the antibody to EGFR is selected from cetuximab, panitumumab, nimotuzumab and necitumumab.
  • the antibody to HER2 is selected from trastuzumab, pertuzumab, and margetuximab.
  • the antibody to CD47 is selected from magrolimab and letaplimab.
  • antibody to CLDN18 is zolbetuximab.
  • the cancer is a solid cancer or the cancer cell is a cell of a solid cancer.
  • the cancer is selected from breast cancer, kidney cancer, head and neck cancer, lung cancer, sarcoma, gastric cancer, colorectal cancer and ovarian cancer.
  • the cancer is characterized by the presence of tumor infiltrating immune cells expressing ILT3.
  • the cancer’s tumor microenvironment is characterized by expression of fibronectin (FN1), Apolipoprotein E (APOE) or both.
  • the ILT3 antibody or antigen binding fragment comprises an IgG4 constant region, optionally wherein the IgG4 constant region comprises a sequence with at least 80% sequence identity to SEQ ID NO: 21 (ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV LQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAP EFEGGPSVFLFSPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNA KTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQ PREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL DSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK),
  • the ILT3 antibody or antigen binding fragment comprises a light chain comprising a kappa constant region, optionally wherein the kappa constant region comprises a sequence with at least 80% sequence identity to SEQ ID NO: 22 (RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQES VTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC).
  • the ILT3 antibody or antigen binding fragment thereof is selected from the group consisting of a Fv, Fab, F(ab')2, scFV, a scFV2 fragment and a single domain antibody.
  • a method of increasing antibody dependent cellular cytotoxicity (ADCC) or antibody dependent cellular phagocytosis (ADCP) by a therapeutic antibody in a subject comprising blocking binding of leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) to at least one of its ligands in the subject; thereby increasing ADCC or ADCP by a therapeutic antibody.
  • the method comprises administering the therapeutic antibody to the subject.
  • the subject suffers from cancer.
  • the therapeutic antibody is an IgGl or IgG3 antibody.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an IgGl or IgG3 antibody and blocking binding of leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) to at least one of its ligands in the subject; thereby treating cancer.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • the at least one ILT3 ligand is fibronectin (FN1), Apolipoprotein E (APOE) or both.
  • the ILT3 ligand is FN1.
  • the blocking binding comprises administering to the subject an antibody or antigen binding fragment thereof that binds to ILT3, and comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein at least one of: a.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 1 (GYSFXiGF) wherein Xi is S or T
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 2 (FPSX2GE) wherein X2 is S or N
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 3 (QAFYYFDX3) wherein X3 is S or Y
  • CDR-L 1 comprises the amino acid sequence as set forth in SEQ ID NO: 4 (KSSQSLLSSSNQKNYLA)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6 (QQYYSYPLT);
  • GYSFXiGF GYSFXiGF
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 2 (FPSX2GE) wherein
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS)
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 50 (AITFGGGNTYYPDSVKG)
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 51 (HGDGNYDFYAMDY)
  • CDR-L 1 comprises the amino acid sequence as set forth in SEQ ID NO: 52 (KSSQSLLNSGNQKNYLT)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 53 (QNDYSYPLT); c.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS), CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 56 (TISSDGGNTYYTDSVKG), CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 57 (HDGRGALDY), CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 58 (RASQDISNYLN), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 59 (YTSRLHS), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 60 (QQGNTLPWT); and d.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 63 (NSAVH)
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 64 (VIWAGGNTNYNSTLMS)
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 65 (HETYGDSFDY)
  • CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 66 (RSSQSLLDSDGKTYLN)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 67 (LVSKLDS)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 68 (WQGTHFPFT).
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE)
  • SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY)
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE)
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS)
  • c is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 16 FPSNGE
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS)
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); d.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); e.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); or f.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYF
  • the antibody or antigen binding fragment thereof comprises at least one of a. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 34 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 35; b. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 36 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 37; c. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 38 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; d.
  • a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 40 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 41; e. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 42 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 41; f. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 42 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; g. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 43 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; h.
  • a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 45; i. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; j. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 46 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; k. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; and l. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 48 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39.
  • the ILT3 antibody or antigen binding fragment thereof is an antibody or antigen binding fragment thereof wherein CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 20, CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 19, CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 17, CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4, CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5, and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6.
  • the ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55.
  • the ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 61 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 62.
  • the ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 69 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 70.
  • the cancer is a CD20-positive cancer and the IgGl or IgG3 antibody is an anti-CD20 antibody selected from rituximab, ibritumomab, ofatumumab, obinutuzumab, ocrelizumab, veltuzumab, ublituximab and ocaratuzumab;
  • the cancer is an epidermal growth factor receptor (EGFR) -positive cancer and the IgGl or IgG3 antibody is an anti-EGFR antibody selected from: cetuximab, panitumumab, nimotuzumab and necitumumab; c.
  • EGFR epidermal growth factor receptor
  • the cancer is a Receptor tyrosine-protein kinase erbB-2 (HER2)-positive cancer and the IgGl or IgG3 antibody is an anti-HER2 antibody selected from: trastuzumab, pertuzumab, and margetuximab; d.
  • the cancer is a CD47 -positive cancer and the IgGl or IgG3 antibody is an anti-CD47 antibody selected from: magrolimab and letaplimab; e.
  • the cancer is a Programmed death-ligand 1 (PD-Ll)-positive cancer and the IgGl or IgG3 antibody is an anti-PD-Ll antibody selected from avelumab, atezolizumab, durvalumab and an antibody comprising the CDRs of atezolizumab or durvalumab; and f.
  • the cancer is a Claudin-18 (CLDN18)-positive cancer and the IgGl or IgG3 antibody is the anti-CLDN18 antibody zolbetuximab.
  • the cancer is a solid cancer or the cancer cell is a cell of a solid cancer.
  • the cancer is selected from breast cancer, kidney cancer, head and neck cancer, lung cancer, sarcoma, gastric cancer, colorectal cancer and ovarian cancer.
  • the cancer is characterized by the presence of tumor infiltrating immune cells expressing ILT3.
  • the cancer’s tumor microenvironment is characterized by expression of fibronectin (FN1), Apolipoprotein E (APOE) or both.
  • the ILT3 antibody or antigen binding fragment comprises an IgG4 constant region, optionally wherein the IgG4 constant region comprises a sequence with at least 80% sequence identity to SEQ ID NO: 21 (ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV LQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAP EFEGGPSVFLFSPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNA KTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQ PREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL DSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK),
  • the ILT3 antibody or antigen binding fragment comprises a light chain comprising a kappa constant region, optionally wherein the kappa constant region comprises a sequence with at least 80% sequence identity to SEQ ID NO: 22
  • the ILT3 antibody or antigen binding fragment thereof is selected from the group consisting of a Fv, Fab, F(ab')2, scFV, and a scFV2 fragment.
  • a method of enhancing immune cell activation against a cancer cell comprising: a. contacting the immune cell with an inhibitor of leukocyte immunoglobulin- like receptor subfamily B member 4 (LILRB4/ILT3); and b. contacting the cancer cell with an inhibitor of epidermal growth factor receptor (EGFR); thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin- like receptor subfamily B member 4
  • EGFR epidermal growth factor receptor
  • a method of enhancing immune cell activation against a cancer cell comprising: a. contacting the immune cell with an inhibitor of ILT3; and b. contacting the cancer cell with an inhibitor of CD47; and thereby enhancing immune cell activation against a cancer cell.
  • a method of enhancing immune cell activation against a cancer cell comprising: a. contacting the immune cell with an inhibitor of ILT3; and b. contacting the cancer cell with an inhibitor of Programmed death-ligand 1 (PD-L1); thereby enhancing immune cell activation against a cancer cell.
  • the immune cell is selected from a monocyte, a macrophage and dendritic cell.
  • the method is performed in a subject in need thereof.
  • the method further comprises contacting the immune cell with the cancer cell.
  • a method of treating cancer in a subject in need thereof comprising enhancing immune cell activation in the subject by a method of the invention, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of EGFR, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of CD47, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of PD-L1, thereby treating cancer.
  • the cancer is an EGFR positive cancer.
  • the cancer is a CD47 positive cancer.
  • the cancer is a PD-L1 positive cancer.
  • the inhibitor is an inhibitory antibody or antigen binding fragment.
  • an inhibitory antibody or antigen binding fragment of ILT3 comprising three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR- L), wherein at least one of: a.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 1 (GYSFXiGF) wherein Xi is S or T
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 2 (FPSX2GE) wherein X2 is S or N
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 3 (QAFYYFDX3) wherein X3 is S or Y
  • CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4 (KSSQSLLSSSNQKNYLA)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6 (QQQ
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS)
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 50 (AITFGGGNTYYPDSVKG)
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 51 (HGDGNYDFYAMDY)
  • CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 52 (KSSQSLLNSGNQKNYLT)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 53 (QNDYSYPLT); c.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS), CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 56 (TISSDGGNTYYTDSVKG), CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 57 (HDGRGALDY), CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 58 (RASQDISNYLN), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 59 (YTSRLHS), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 60 (QQGNTLPWT); and d.
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 63 (NSAVH)
  • CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 64 (VIWAGGNTNYNSTLMS)
  • CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 65 (HETYGDSFDY)
  • CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 66 (RSSQSLLDSDGKTYLN)
  • CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 67 (LVSKLDS)
  • CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 68 (WQGTHFPFT).
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE)
  • SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY)
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE)
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS)
  • c is SEQ ID NO: 15 (GYSFTGF)
  • SEQ ID NO: 16 FPSNGE
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS)
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); d.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); e.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY); or f.
  • SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF), SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE) and SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYF
  • the antibody or antigen binding fragment thereof comprises at least one of a. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 34 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 35; b. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 36 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 37; c. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 38 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; d.
  • a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 40 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 41; e. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 42 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 41; f. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 42 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; g. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 43 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; h.
  • a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 45; i. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 44 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; j. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 46 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; k. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39; and l. a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 48 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 39.
  • the inhibitory antibody of ILT3 is an antibody wherein CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 20, CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 19, CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 17, CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4, CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5, and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6.
  • the antibody is antibody 5E5 or an antibody that competes with antibody 5E5 for binding to ILT3.
  • the inhibitory antibody or antigen binding fragment of ILT3 comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55.
  • the inhibitory antibody or antigen binding fragment of ILT3 comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 61 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 62.
  • the inhibitory antibody or antigen binding fragment of ILT3 comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 69 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 70.
  • the inhibitor of EGFR, CD47, and/or PD-L1 is a blocking antibody comprising an IgGl or IgG3 scaffold.
  • an inhibitory antibody of EGFR is selected from cetuximab, panitumumab, nimotuzumab, and necitumumab.
  • an inhibitory antibody of CD47 is selected from magrolimab and letaplimab.
  • an inhibitory antibody of PD-L1 is selected from avelumab, atezolizumab and durvalumab.
  • the cancer is a solid cancer or the cancer cell is a cell of a solid cancer.
  • the cancer is selected from breast cancer, kidney cancer, head and neck cancer, lung cancer, sarcoma, gastric cancer, colorectal cancer and ovarian cancer.
  • the cancer is characterized by the presence of tumor infiltrating immune cells expressing ILT3.
  • the cancer’s tumor microenvironment is characterized by expression of fibronectin (FN1).
  • the inhibitory antibody or antigen binding fragment comprises an IgG4 constant region, optionally wherein the IgG4 constant region comprises a sequence with at least 80% sequence identity to SEQ ID NO: 21 (ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV EQSSGEYSESSVVTVPSSSEGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAP EFEGGPSVFEFSPKPKDTEMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNA KTKPREEQFNSTYRVVSVETVEHQDWENGKEYKCKVSNKGEPSSIEKTISKAKGQ PREPQVYTEPPSQEEMTKNQVSETCEVKGFYPSDIAVEWESNGQPENNYKTTPPVE
  • the inhibitory antibody or antigen binding fragment comprises a light chain comprising a kappa constant region, optionally wherein the kappa constant region comprises a sequence with at least 80% sequence identity to SEQ ID NO: 22
  • an ILT3 inhibitor inhibits binding of ILT3 to an ILT3 ligand.
  • the inhibitor inhibits binding of ILT3 to Apolipoprotein E (APOE), fibronectin (FN1) or both.
  • APOE Apolipoprotein E
  • FN1 fibronectin
  • the inhibitor of ILT3 is a blocking antibody or antigen binding fragment thereof selected from the group consisting of a Fv, Fab, F(ab')2, scFV, a scFV2 fragment and a single domain antibody.
  • Figures 1A-1D Bar graphs of (1A) IFNg secretion from CD8 T cells in the presence of autologous MDSCs, (IB) TNFa secretion from DCs and (1C) IL-8 secretion from THP- 1 cells in the presence of fibronectin and Rituximab treated with 5E5.
  • 1A includes an inset of the 11 donor samples tested showing levels after administration of the isotype control and 5E5 at a concentration of 16 pg/ml.
  • ID Bar graph of IL-8 secretion from THP-1 cells contacted with Rituximab in the presence of fibronectin or APOE.
  • Figures 2A-2B Bar graph of TNFa secretion from monocytes differentiated into Ml activated macrophages (MF) in the presence of (2A) gastric cancer cells or (2B) colon cancer cells with and without the h5E5 antibody. * P ⁇ 0.05; un-paired Student’s T-test.
  • FIG. 3 Bar graphs of TNFa secretion from immature DCs stimulated with IL-10 and LPS to generate DCtols when cultured with or without anti-ILT3 antibodies or isotype control. Samples from 2 donors are shown.
  • Figures 4A-4B Bar graphs of (4A) IFNg secretion from CD8 T cells incubated with autologous MDSC, and (4B) IFNg secretion from CD4 T cells co-cultured with the indicated dendritic cells treated with anti-PDl antibody, 5E5 or a combination of the two.
  • FIG. 5A-5E Bar graphs of (5A) IFNg production from sarcoma tumoroid and (5B) CXCL1 production from breast cancer tumoroid activated in the presence of 5E5 or isotype control.
  • 5C Bar graphs of CCL3, CCL4 and IL-8 production from breast cancer tumoroids activated in the presence of 5E5 or isotype control.
  • 5D Bar graphs of CCL3, M-CSF, IL-lb, CCL4, CXCL1, IFNg, IL-8, IL-6, CCL2 and TNFa production from renal cell carcinoma tumoroids activated in the presence of 5E5 or other known anti-ILT3 antibodies or isotype control.
  • FIGS 6A-6B Line graphs of MC38 tumor size over time in syngeneic transgenic hILT3 mice treated with (6A) 5E5 or an isotype control or (6B) anti-PD-1 alone or in combination with 5E5.
  • X-axis lists the first, second, third and fourth injections of the therapeutics.
  • TGI tumor growth inhibition
  • *P ⁇ 0.05 2-way repeated measures ANOVA.
  • n 10 mice per group.
  • Figures 7A-7F (7A-7B) Line graphs of EO771 tumor size over time in syngeneic transgenic hILT3 mice treated with anti-PDl antibody, or combinations of anti-PDl with anti-ILT3 antibodies 5E5 and h52B8.
  • the vertical dotted lines indicate the group’s randomization point.
  • the horizontal dotted lines indicate the 400 mm A 3 cutoff.
  • TGI tumor growth inhibition; P ⁇ 0.001, 2-way repeated measures ANOVA vs. control IgG and vs. anti- PD-1.
  • N 10-11 mice for each group.
  • TGI tumor growth inhibition
  • FIGS 8A-8E Bar graphs of IL-8 secretion from THP-1 cells activated by (8A) Erbitux, (8B) Herceptin, (8C) Avelumab or (8D) Zolbetuximab in the presence or absence of fibronectin and antibody 5E5.
  • Figures 9A-9K (9A-9I) Bar graphs of levels of (9A) CCL2, (9B) CCL3, (9C) CCL4, (9D) IL-6, (9E, 91) IL-8, (9F) TNFa, (9G) CXCL1, and (9H) M-CSF in media after culture of macrophages with A549 cells.
  • A549 cells are preincubated either with Erbitux, Magrolimab or nothing and A357 cells are preincubated with Avelumab or nothing. Combinations of the variously treated macrophages and cancer cells are tested for the recited molecules in the media after 24 hours. Results using macrophages from (9A-9H and 9J-9K) a first healthy donor and (91) a second healthy donor are presented.
  • Figures 10A-10B Bar graphs of NK cytotoxicity against (10A) A549 and (10B) H441 cancer cells. NK cells were either cultured alone or cocultured with M2c macrophages. Macrophages were generated with or without addition of the 5E5 antibody. Cancer cells were preincubated with (10A) Erbitux or (10B) Avelumab. hlgGl and hIgG4 were used as negative controls for 5E5 and Erbitux or Avelumab. Cytotoxicity was assessed in triplicates and evaluated by CytoTox-Glo assay kit (Promega, cat No G9291), according to the manufacturer's instructions. Percent of specific cytotoxicity was calculated as: lOOx est sam pl e- Low control (target cells only)-Effector cells
  • the present invention provides methods of enhancing immune cell activation against a cancer cell comprising contacting the immune cell with an inhibitor of leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) and contacting the cancer cell with an inhibitor of epidermal growth factor receptor (EGFR), CD47 or Programmed death-ligand 1 (PD-L1).
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • EGFR epidermal growth factor receptor
  • CD47 CD47
  • P-L1 Programmed death-ligand 1
  • Methods of treating cancer by administering an inhibitor of ILT3 and an inhibitor of any one of EGFR, CD47 and PD-L1 or an anticancer agent comprising an IgGl or IgG3 scaffold are also provided.
  • Methods of increasing antibody dependent cell cytotoxicity (ADCC) or antibody dependent cell phagocytosis (ADCP) against a cancer cell and treating cancer comprising blocking binding of fibronectin to ILT3 are also provided and are methods of treating cancer in a subject in need thereof, comprising administering an ILT3 antibody and at least one of a CD20 antibody, a epidermal growth factor receptor (EGFR) antibody, a Receptor tyro sine-protein kinase erbB-2 (HER2) antibody, a CD47 antibody, a Programmed death-ligand 1 (PD-L1) antibody and a Claudin-18 (CLDN18) antibody.
  • ADCC antibody dependent cell cytotoxicity
  • ADCP antibody dependent cell phagocytosis
  • the invention is based on the surprising finding that combining ILT3 blockade of immune cells with agents that target cancer surface proteins, and in specific CD20, EGFR, HER2, CD47, CLDN18 and PD-L1, produces a synergistic effect that greatly enhances immune cell activation, ADCC, ADCP and cancer cell killing.
  • This synergistic effect is far more than merely an addition of the effect of the ILT3 blockade with the effect of the anticancer agent and instead causes a wholly unexpected anticancer response.
  • ADCC antibody dependent cellular cytotoxicity
  • ADCP antibody dependent cellular phagocytosis
  • a method of treating cancer in a subject comprising administering to the subject an antibody and blocking binding of leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) to at least one of its ligands in the subject; thereby treating cancer.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting CD20 on the cancer cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting epidermal growth factor receptor (EGFR) on the cancer cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • EGFR epidermal growth factor receptor
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting Receptor tyrosine-protein kinase erbB-2 (HER2) on the cancer cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • HER2 Receptor tyrosine-protein kinase erbB-2
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting CD47 on the cancer cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting Programmed death-ligand 1 (PD-L1) on the cancer cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • PD-L1 Programmed death-ligand 1
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting Claudin- 18 (CLDN18) on the cancer cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • CLDN18 Claudin- 18
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and inhibiting Programmed cell death protein 1 (PD-1) on the immune cell, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • PD-1 Programmed cell death protein 1
  • a method of enhancing immune cell activation against a cancer cell comprising inhibiting leukocyte immunoglobulin-like receptor subfamily B member 4 (LILRB4/ILT3) on an immune cell and contacting the cancer cell with an anticancer agent comprising an IgGl or IgG3 scaffold, thereby enhancing immune cell activation against a cancer cell.
  • LILRB4/ILT3 leukocyte immunoglobulin-like receptor subfamily B member 4
  • the method is an in vitro method. In some embodiments, the method is an in vivo method. In some embodiments, the method is an ex vivo method. In some embodiments, the method is in a subject in need thereof. In some embodiments, the method is performed in a subject in need of the method. In some embodiments, the method is performed in culture. In some embodiments, the method is a therapeutic method. In some embodiments, the subject suffers from cancer.
  • the method further comprises contacting the immune cell with the cancer cell. In some embodiments, contacting is coculturing. In some embodiments, the contacted immune cell is contacted with the contacted cancer cell. In some embodiments, the method is an ex vivo method and the cells are contacted. In some embodiments, the method further comprises administering an antibody to the subject. In some embodiments, the method further comprises administering an anti-ILT3 antibody to the subject.
  • a method of treating cancer in a subject in need thereof comprising enhancing immune cell activation in the subject by a method of the invention, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of CD20, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of EGFR, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of HER2, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of CD47, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of PD-L1, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of CLDN18, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an inhibitor of PD-1, thereby treating cancer.
  • a method of treating cancer in a subject in need thereof comprising administering to the subject an inhibitor of LILRB4/ILT3 and an anticancer agent comprising an IgGl or IgG3 scaffold, thereby treating cancer.
  • enhancing immune cell activation is enhancing immune cell activation against the cancer.
  • the subject suffers from cancer.
  • the cancer is a CD20 positive cancer.
  • the cancer is an EGFR positive cancer.
  • the cancer is a HER2 positive cancer.
  • the cancer is a CD47 positive cancer.
  • the cancer is a PD-L1 positive cancer.
  • the cancer is a CLDN18 positive cancer.
  • the cancer is treatable by the anticancer agent.
  • the cancer is treatable by the antibody.
  • the cancer expresses a surface protein that is the target of the anticancer agent.
  • the cancer expresses a surface protein that is the target of the antibody.
  • the surface protein is a receptor.
  • the surface protein is a cancer antigen.
  • the antibody is a therapeutic antibody.
  • the antibody binds to a factor expressed on the cancer.
  • expressed on the cancer is expressed on the surface of the cancer.
  • the factor is selected from CD20, EGFR, HER2, CD47, CLDN18 and PD-L1. In some embodiments, the factor is selected from HER2, CD47, and CLDN18.
  • inhibiting comprises administering an agent.
  • the agent is an inhibitor.
  • the agent is a therapeutic molecule.
  • the agent is a composition comprising the inhibitor.
  • agent is an antibody or antigen binding fragment thereof.
  • the antibody or antigen binding fragment thereof is a therapeutic antibody or antigen binding fragment thereof.
  • the antibody or antigen binding fragment thereof is a therapeutic antibody is an IgGl antibody.
  • the antibody or antigen binding fragment thereof is a therapeutic antibody is an IgGl or IgG3 antibody.
  • the antibody or antigen binding fragment thereof is an opsonizing antibody or antigen binding fragment thereof.
  • an “opsonizing agent” is any agent that can bind to a target cell (e.g., a cancer cell, a cell harboring an intracellular pathogen, etc.) and opsonize the target cell.
  • a target cell e.g., a cancer cell, a cell harboring an intracellular pathogen, etc.
  • an opsonizing agent is an antibody that induces antibody dependent cellular phagocytosis (ADCP).
  • the antibody is an antibody that induces antibody dependent cellular cytotoxicity (ADCC) and/or antibody dependent cellular phagocytosis (ADCP).
  • inhibiting ILT3 comprises administering an agent that binds to ILT3.
  • the agent is specific to ILT3.
  • the agent is specific to ILT3 and not to other ILT proteins.
  • the agent inhibits ILT3.
  • the agent blocks ILT3.
  • the agent is an ILT3 inhibitor.
  • the ILT3 inhibitor is an ILT3 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • the agent is an ILT3 antibody or antigen binding fragment thereof.
  • inhibiting CD20 comprises administering an agent that binds to CD20.
  • the agent is specific to CD20.
  • the agent inhibits CD20.
  • the agent blocks CD20.
  • the agent is a CD20 inhibitor.
  • the CD20 inhibitor is a CD20 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • an agent that binds CD20 is a CD20 antibody.
  • X antibody and “anti-X antibody” shall be used interchangeably and have the same meaning.
  • inhibiting EGFR comprises administering an agent that binds to EGFR.
  • the agent is specific to EGFR.
  • the agent inhibits EGFR.
  • the agent blocks EGFR.
  • the agent is an EGFR inhibitor.
  • the EGFR inhibitor is an EGFR inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • an agent that binds EGFR is an EGFR antibody.
  • inhibiting HER2 comprises administering an agent that binds to HER2.
  • the agent is specific to HER2.
  • the agent inhibits HER2.
  • the agent blocks HER2.
  • the agent is a HER2 inhibitor.
  • the HER2 inhibitor is a HER2 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • an agent that binds HER2 is a HER2 antibody.
  • inhibiting CD47 comprises administering an agent that binds to CD47.
  • the agent is specific to CD47.
  • the agent inhibits CD47.
  • the agent blocks CD47.
  • the agent is a CD47 inhibitor.
  • the CD47 inhibitor is a CD47 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • inhibiting PD-L1 comprises administering an agent that binds to PD-L1.
  • the agent is specific to PD-L1.
  • the agent inhibits PD-L1.
  • the agent blocks PD-L1.
  • the agent is a PD-L1 inhibitor.
  • the PD-L1 inhibitor is a PD-L1 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • inhibiting CLDN18 comprises administering an agent that binds to CLDN18.
  • the agent is specific to CLDN18.
  • the agent inhibits CLDN18.
  • the agent blocks CLDN18.
  • the agent is a CLDN18 inhibitor.
  • the CLDN18 inhibitor is a CLDN18 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • an agent that binds CLDN18 is a CLDN18 antibody.
  • inhibiting PD-1 comprises administering an agent that binds to PD-1.
  • the agent is specific to PD-1.
  • the agent inhibits PD-1.
  • the agent blocks PD-1.
  • the agent is a PD-1 inhibitor.
  • the PD-1 inhibitor is a PD-1 inhibitory antibody or antigen binding fragment thereof.
  • an inhibitory antibody is a blocking antibody.
  • an inhibitor is an antagonist.
  • the agent is an inhibitor. In some embodiments, the agent is an antibody. In some embodiments, the agent is a blocking antibody. In some embodiments, the agent is an antagonist. In some embodiments, the agent binds to and blocks/inhibits a target protein on the surface of the cancer cell. In some embodiments, the agent is specific to the surface protein. In some embodiments, the agent binds the surface protein. In some embodiments, the agent inhibits the surface protein. In some embodiments, the agent blocks the surface protein. In some embodiments, the agent comprises an IgGl scaffold. In some embodiments, the agent comprises an IgG3 scaffold.
  • the terms “IgGl or IgG3 scaffold” refer to an agent comprising the constant region or Fc domain of IgGl or IgG3.
  • the agent is an IgGl antibody.
  • the agent is an IgG3 antibody.
  • the agent promotes antibody-dependent cellular cytotoxicity (ADCC).
  • the agent promotes antibody-dependent cellular phagocytosis (ADCP).
  • ADCC antibody-dependent cellular cytotoxicity
  • ADCP antibody-dependent cellular phagocytosis
  • promotes is induces. It will be understood by a skilled artisan that unmodified IgGl and IgG3 antibodies induce both ADCC and ADCP. However, there are modifications/mutations that can diminish or abolish this function.
  • mutation of a glycosylation site in the IgG backbone can abolish glycosylation and ADCC/ADCP.
  • IgGl and IgG3 it is directed to forms that have not been mutated to abolish/diminish ADCC/ADCP.
  • Leukocyte immunoglobulin-like receptor subfamily B member 4 is known as LILRB4, ILT3 and CD85K among many other names.
  • the Entrez gene identifier for human ILT3 is 11006 and the mouse identifier is 14727.
  • the protein sequence for human ILT3 can be found under the Uniprot identifier Q8NHJ6 and the mouse protein sequence under identifier Q61450.
  • the human mRNA sequence can be found in RefSeq sequences NM_001081438, NM_001278426, NM_001278427, NM_001278428 and NM_001278429.
  • the mouse mRNA sequence can be found in RefSeq sequences NM_001291892, NM_001291893 and NM_008147.
  • the human protein sequence can be found in RefSeq sequences NP_001265355, NP_001265356, NP_001265357, NP_001265358 and NP_001265359.
  • the mouse protein sequence can be found in RefSeq sequences NP001278821, NP_00127882 and NP_032173.
  • the amino acid sequence of ILT3 comprises
  • the amino acid sequence of ILT3 consists of SEQ ID NO: 71.
  • SEQ ID NO: 71 is human ILT3.
  • B-lymphocyte antigen CD20 is known as CD20, MS4A1, CVID5 among other names.
  • the Entrez gene identifier for human CD20 is 931 and the mouse identifier is 12482.
  • the protein sequence for human CD20 can be found under the Uniprot identifier Pl 1836.
  • the human mRNA sequence can be found in RefSeq sequences NM_152866, NM_021950 and NM_152867.
  • the human protein sequence can be found in RefSeq sequences NP_068769, NP_690605, and NP_690606.
  • Epidermal growth factor receptor is known as EGFR, IAP and OA3 among other names.
  • the Entrez gene identifier for human EGFR is 1956 and the mouse identifier is 13649.
  • the protein sequence for human EGFR can be found under the Uniprot identifier P00533.
  • the human mRNA sequence can be found in RefSeq sequences NM_001346897, NM_01346898, NM_001346899, NM_001346900 and NM_001346941.
  • the human protein sequence can be found in RefSeq sequences NP_001333826, NP_001333827, NP_001333828, NP_001333829 and NP_001333870.
  • Receptor tyro sine-protein kinase erbB-2 is known as HER2, ERBB2 and CD340 among many other names.
  • the Entrez gene identifier for human ERBB2 is 2064 and the mouse identifier is 13866.
  • the protein sequence for human ERBB2 can be found under the Uniprot identifier P04626.
  • the human mRNA sequence can be found in RefSeq sequences NM_004448, NM_001005862, NM_001289936, NM_001289937 and NM_001289938.
  • the human protein sequence can be found in RefSeq sequences NP_004439, NP_001005862, NP_001276865, NP_001276866 and NP_001276867.
  • Cluster of Differentiation 47 is known as CD47, ErbB-1 and HER1 among many other names.
  • the Entrez gene identifier for human CD47 is 961 and the mouse identifier is 16423.
  • the protein sequence for human CD47 can be found under the Uniprot identifier Q08722.
  • the human mRNA sequence can be found in RefSeq sequences NM_001025079, NM_001025082, NM_001777, NM_198793 and NM_001382306.
  • the human protein sequence can be found in RefSeq sequences NP_001768, NP_942088, and NP_001369235.
  • Programmed death-ligand 1 is known as PD-L1, CD274 and B7-H1 among other names.
  • the Entrez gene identifier for human PD-L1 is 29126 and the mouse identifier is 60533.
  • the protein sequence for human PD-L1 can be found under the Uniprot identifier Q9NZQ7.
  • the human mRNA sequence can be found in RefSeq sequences NM_001314029, NM_001267706, and NM_014143.
  • the human protein sequence can be found in RefSeq sequences NP_001254635, NP_001300958, and NP_054862.
  • Claudin-18 is known as CLDN18, SFTA5 and SFTPJ among many other names.
  • the Entrez gene identifier for human CLDN18 is 51208 and the mouse identifier is 56492.
  • the protein sequence for human CLDN18 can be found under the Uniprot identifier P56856.
  • the human mRNA sequence can be found in RefSeq sequences NM_016369, and NM_001002026.
  • the human protein sequence can be found in RefSeq sequences NP_057453 and NP_001002026.
  • Programmed cell death protein 1 is known as PD-1, CD279 and PDCD1 among other names.
  • the Entrez gene identifier for human PD-1 is 5133 and the mouse identifier is 18566.
  • the protein sequence for human PD-1 can be found under the Uniprot identifier Q15116.
  • the human mRNA sequence can be found in RefSeq sequence NM_005018.
  • the human protein sequence can be found in RefSeq sequences NP_005009.
  • an antibody refers to a polypeptide or group of polypeptides that include at least one binding domain that is formed from the folding of polypeptide chains having three-dimensional binding spaces with internal surface shapes and charge distributions complementary to the features of an antigenic determinant of an antigen.
  • An antibody typically has a tetrameric form, comprising two identical pairs of polypeptide chains, each pair having one "light” and one "heavy” chain. The variable regions of each light/heavy chain pair form an antibody binding site.
  • An antibody may be oligoclonal, polyclonal, monoclonal, chimeric, camelised, CDR-grafted, multi- specific, bi-specific, catalytic, humanized, fully human, anti- idiotypic and antibodies that can be labeled in soluble or bound form as well as fragments, including epitope-binding fragments, variants or derivatives thereof, either alone or in combination with other amino acid sequences.
  • An antibody may be from any species.
  • the term antibody also includes binding fragments, including, but not limited to Fv, Fab, Fab', F(ab')2 single stranded antibody (svFC), dimeric variable region (Diabody) and disulphide-linked variable region (dsFv).
  • antibodies include immunoglobulin molecules and immunologically active fragments of immunoglobulin molecules, i.e., molecules that contain an antigen binding site.
  • Antibody fragments may or may not be fused to another immunoglobulin domain including but not limited to, an Fc region or fragment thereof.
  • Other fusion products may be generated including but not limited to, scFv- Fc fusions, variable region (e.g., VL and VH) ⁇ Fc fusions and scFv-scFv-Fc fusions.
  • the antibody is a full antibody.
  • the antibody comprises an IgGl or IgG3 scaffold.
  • the antibody is an IgGl antibody.
  • the antibody is an IgG3 antibody.
  • Immunoglobulin molecules can be of any type (e.g., IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl and IgA2) or subclass.
  • the antibody is a full antibody.
  • the antibody comprises an IgGl or IgG3 scaffold.
  • the antibody is an IgGl antibody.
  • the antibody is an IgG3 antibody.
  • the antibody is a murine antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the antibody is a humanized antibody. In some embodiments, the antibody is a human antibody. In some embodiments, the antibody is a full IgG. In some embodiments, the antibody is an IgGl. In some embodiments, the antibody is an IgG2. In some embodiments, the antibody is an IgG3. In some embodiments, the antibody is an IgG4. In some embodiments, the antibody or antigen binding domain is an antigen binding domain that lacks an Fc domain.
  • Examples of further antibodies that may be used as part of the methods of the invention are provided in Table 1. These include IgGl antibodies as well as antibodies that are known to induce ADCC and/or ADCP.
  • the peptide is selected from an antibody, an antigen binding fragment of an antibody, a Fab fragment, a single chain antibody, a single-domain antibody, a nanobody, a VHH antibody and an antibody mimetic.
  • antibody mimetic refers to an organic compound that can specifically bind to a target antigen.
  • an antibody mimetic is not structurally related to an antibody. Examples of antibody mimetics include, but are not limited to, affilins, affimers, affitins, alphabodies, anticalins, avimers, DARPins, fynomers, Kunitz domain peptides, monobodies, and nanoCLAMPS.
  • the antibody mimetic is a DARPin. All of these agents are well known in the art and are known to be useful in blocking interactions between receptors and their ligands.
  • the protein is an antibody mimetic.
  • DARPin refers to a designed ankyrin repeat protein. DARPins are genetically engineered antibody mimetic proteins that are generally highly specific for their protein target.
  • the basic unit of the naturally occurring antibody structure is a heterotetrameric glycoprotein complex of about 150,000 Daltons, composed of two identical light (L) chains and two identical heavy (H) chains, linked together by both noncovalent associations and by disulfide bonds. Each heavy and light chain also has regularly spaced intra-chain disulfide bridges.
  • Five human antibody classes (IgG, IgA, IgM, IgD and IgE) exist, and within these classes, various subclasses, are recognized based on structural differences, such as the number of immunoglobulin units in a single antibody molecule, the disulfide bridge structure of the individual units, and differences in chain length and sequence.
  • the class and subclass of an antibody is its isotype.
  • variable domains The amino terminal regions of the heavy and light chains are more diverse in sequence than the carboxy terminal regions, and hence are termed the variable domains.
  • This part of the antibody structure confers the antigen-binding specificity of the antibody.
  • a heavy variable (VH) domain and a light variable (VL) domain together form a single antigenbinding site, thus, the basic immunoglobulin unit has two antigen-binding sites.
  • Particular amino acid residues are believed to form an interface between the light and heavy chain variable domains (Chothia et al., J. Mol. Biol. 186, 651-63 (1985); Novotny and Haber, (1985) Proc. Natl. Acad. Sci. USA 82 4592-4596).
  • the carboxy terminal portion of the heavy and light chains form the constant domains i.e., CHI, CH2, CH3, CL. While there is much less diversity in these domains, there are differences from one animal species to another, and further, within the same individual there are several different isotypes of antibody, each having a different function.
  • FR frame region
  • hypervariable region refers to the amino acid residues in the variable domain of an antibody, which are other than the hypervariable region amino acid residues as herein defined.
  • hypervariable region refers to the amino acid residues in the variable domain of an antibody, which are responsible for antigen binding.
  • the hypervariable region comprises amino acid residues from a “complementarity determining region” or “CDR”.
  • CDRs are primarily responsible for binding to an epitope of an antigen.
  • the extent of FRs and CDRs has been precisely defined (see, Kabat et al.).
  • Immunoglobulin variable domains can also be analyzed using the IMGT information system (//imgt. cines.fr/) (IMGT/V-Quest) to identify variable region segments, including CDRs. See, e.g., Brochet, X. et al, Nucl. Acids Res. J6:W503-508 (2008).
  • Chothia et al. also defined a numbering system for variable domain sequences that is applicable to any antibody.
  • One of ordinary skill in the art can unambiguously assign this system of "Chothia numbering" to any variable domain sequence, without reliance on any experimental data beyond the sequence itself.
  • “Chothia numbering” refers to the numbering system set forth by Chothia et al., Journal of Molecular Biology, “Canonical Structures for the Hypervariable regions of immunoglobulins” (1987) and Chothia et al., Nature, “Conformations of Immunoglobulin Hypervariable Regions” (1989).
  • humanized antibody refers to an antibody from a nonhuman species whose protein sequences have been modified to increase similarity to human antibodies.
  • a humanized antibody may be produced by production of recombinant DNA coding for the CDRs of the non-human antibody surrounded by sequences that resemble a human antibody.
  • the humanized antibody is a chimeric antibody.
  • humanizing comprises insertion of the CDRs of the invention into a human antibody scaffold or backbone. Humanized antibodies are well known in the art and any method of producing them that retains the CDRs of the invention may be employed.
  • the antibody is a monoclonal antibody.
  • monoclonal antibody or “mAb” as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and/or bind the same epitope, except for possible variants that may arise during production of the monoclonal antibody, such variants generally being present in minor amounts.
  • polyclonal antibody preparations that typically include different antibodies directed against different determinants (epitopes)
  • each monoclonal antibody is directed against a single determinant on the antigen.
  • the monoclonal antibodies are advantageous in that they are uncontaminated by other immunoglobulins.
  • the modifier "monoclonal” indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies and is not to be construed as produced by any specific preparation method.
  • Monoclonal antibodies to be used in accordance with the methods provided herein may be made by the hybridoma method first described by Kohler et al, Nature 256:495 (1975), or may be made by recombinant DNA methods (see, e.g., U.S. Patent No. 4,816,567).
  • the “monoclonal antibodies” may also be isolated from phage antibody libraries using the techniques described in Clackson et al, Nature 352:624-628 (1991) and Marks et al, J. Mol. Biol. 222:581-597 (1991), for example.
  • the mAb of the present invention may be of any immunoglobulin class including IgG, IgM, IgD, IgE or IgA.
  • a hybridoma producing a mAb may be cultivated in vitro or in vivo. High titers of mAbs can be obtained in vivo production where cells from the individual hybridomas are injected intraperitoneally into pristine -primed Balb/c mice to produce ascites fluid containing high concentrations of the desired mAbs.
  • mAbs of isotype IgM or IgG may be purified from such ascites fluids, or from culture supernatants, using column chromatography methods well known to those of skill in the art.
  • Antibody fragments comprise a portion of an intact antibody, preferably comprising the antigen binding region thereof.
  • antibody fragments include Fab, Fab', F(ab')2, and Fv fragments; diabodies; tandem diabodies (taDb), linear antibodies (e.g., U.S. Patent No. 5,641,870, Example 2; Zapata et al, Protein Eng.
  • Papain digestion of antibodies produces two identical antigen-binding fragments, called “Fab” fragments, each with a single antigen-binding site, and a residual "Fc” fragment, whose name reflects its ability to crystallize readily.
  • Pepsin treatment yields an F(ab')2 fragment that has two antigen-binding sites and is still capable of cross-linking antigen.
  • Fv is the minimum antibody fragment that contains a complete antigen-recognition and antigen-binding site. This region consists of a dimer of one heavy chain and one light chain variable domain in tight, non-covalent association. It is in this configuration that the three surfaces of the VH-VE dimer. Collectively, the six hypervariable regions confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv comprising only three hypervariable regions specific for an antigen) has the ability to recognize and bind antigen, although at a lower affinity than the entire binding site.
  • the Fab fragment also contains the constant domain of the light chain and the first constant domain (CHI) of the heavy chain.
  • Fab' fragments differ from Fab fragments by the addition of a few residues at the carboxy terminus of the heavy chain CHI domain including one or more cysteines from the antibody hinge region.
  • Fab'-SH is the designation herein for Fab' in which the cysteine residue(s) of the constant domains bear at least one free thiol group.
  • F(ab')2 antibody fragments originally were produced as pairs of Fab' fragments that have hinge cysteines between them. Other chemical couplings of antibody fragments are also known.
  • the "light chains" of antibodies (immunoglobulins) from any vertebrate species can be assigned to one of two clearly distinct types, called kappa and lambda, based on the amino acid sequences of their constant domains.
  • antibodies can be assigned to different classes. There are five major classes of intact antibodies: IgA, IgD, IgE, IgG, and IgM, and several of these may be further divided into subclasses (isotypes), e.g., IgGl, IgG2, IgG3, IgG4, IgA, and IgA2.
  • the heavy chain constant domains that correspond to the different classes of antibodies are called a, delta, e, gamma, and micro, respectively.
  • the subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known.
  • Single-chain Fv or “scFv” antibody fragments comprise the VH and VE domains of antibody, wherein these domains are present in a single polypeptide chain.
  • the Fv polypeptide further comprises a polypeptide linker between the VH and VL domains that enables the scFv to form the desired structure for antigen binding.
  • diabodies refers to small antibody fragments with two antigen-binding sites, which fragments comprise a heavy chain variable domain (VH) connected to a light chain variable domain (VL) in the same polypeptide chain (VH - VL).
  • VH heavy chain variable domain
  • VL light chain variable domain
  • multispecific antibody is used in the broadest sense and specifically covers an antibody that has polyepitopic specificity.
  • Such multispecific antibodies include, but are not limited to, an antibody comprising a heavy chain variable domain (VH) and a light chain variable domain (VL), where the VHVL unit has polyepitopic specificity, antibodies having two or more VL and VH domains with each VHVL unit binding to a different epitope, antibodies having two or more single variable domains with each single variable domain binding to a different epitope, full length antibodies, antibody fragments such as Fab, Fv, dsFv, scFv, diabodies, bispecific diabodies, triabodies, tri-functional antibodies, antibody fragments that have been linked covalently or non-covalently.
  • Polyepitopic specificity refers to the ability to specifically bind to two or more different epitopes on the same or different target(s).
  • the monoclonal antibodies of the invention may be prepared using methods well known in the art. Examples include various techniques, such as those in Kohler, G. and Milstein, C, Nature 256: 495-497 (1975); Kozbor et al, Immunology Today 4: 72 (1983); Cole et al, pg. 77-96 in MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc. (1985).
  • antibodies can be generated in vitro using phage display technology. Such a production of recombinant antibodies is much faster compared to conventional antibody production, and they can be generated against an enormous number of antigens. Furthermore, when using the conventional method, many antigens prove to be non-immunogenic or extremely toxic, and therefore cannot be used to generate antibodies in animals. Moreover, affinity maturation (i.e., increasing the affinity and specificity) of recombinant antibodies is very simple and relatively fast. Finally, large numbers of different antibodies against a specific antigen can be generated in one selection procedure.
  • a library can be made in several ways: One can generate a synthetic repertoire by cloning synthetic CDR3 regions in a pool of heavy chain germline genes and thus generating a large antibody repertoire, from which recombinant antibody fragments with various specificities can be selected.
  • Non-human antibodies may be humanized by any methods known in the art.
  • the non-human complementarity determining regions (CDRs) are inserted into a human antibody or consensus antibody framework sequence. Further changes can then be introduced into the antibody framework to modulate affinity or immunogenicity.
  • antibodies and portions thereof include: antibodies, fragments of antibodies, Fab and F(ab')2, single-domain antigen -binding recombinant fragments and natural nanobodies.
  • the antigen binding fragment is selected from the group consisting of a Fv, Fab, F(ab')2, scFV or a SCFV2 fragment.
  • each framework region present in the encoded polypeptide may comprise at least one amino acid substitution relative to the corresponding human acceptor framework.
  • the framework regions may comprise, in total, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, or fifteen amino acid substitutions relative to the acceptor framework regions.
  • amino acid substitutions i.e. "conservative substitutions” may be made, for instance, on the basis of similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, and/or the amphipathic nature of the residues involved.
  • the antibody is an anti-ILT3 antibody. In some embodiments, the antibody binds to ILT3. In some embodiments, the antibody is specific to ILT3. In some embodiments, binds to is specifically binds to. In some embodiments, the anti-ILT3 antibody is one disclosed in International Patent Application PCT/IL2023/050426, the contents of which are hereby incorporated herein by reference. In some embodiments, the antibody is an ILT3 blocking antibody. In some embodiments, the antibody is an ILT3 inhibitory antibody. In some embodiments, the antibody is an ILT3 antagonist.
  • the anti-ILT3 antibody or antigen-binding portion thereof comprising three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein: CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 1 (GYSFXiGF) wherein Xi is S or T, CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 2 (FPSX2GE) wherein X2 is S or N, CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 3 (QAFYYFDX3) wherein X3 is S or Y, CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4 (KSSQSLLSSSNQKNYLA), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6 (QQYYSYPL
  • the anti-ILT3 antibody or antigen-binding portion thereof comprising three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein: CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 13 (GFYID), CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 14 (YIFPSX2GETSYNQKFKG) wherein X2 is S or N, CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 3 (QAFYYFDX3) wherein X3 is S or Y, CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4 (KSSQSLLSSSNQKNYLA), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6 (QQYYSYPLT).
  • CDR-H1 comprises the amino acid sequence
  • the CDRs are numbered by the Kabat numbering system and the CDRs comprise SEQ ID NO: 13-14 and 3-6. It will be understood by a skilled artisan that the Chothia and Kabat systems are interchangeable and that CDRs given for either system can be converted into the CDRs of the other system.
  • SEQ ID NO: 1 is SEQ ID NO: 15 (GYSFTGF). In some embodiments, SEQ ID NO: 1 is SEQ ID NO: 20 (GYSFSGF). In some embodiments, SEQ ID NO: 2 is SEQ ID NO: 16 (FPSNGE). In some embodiments, SEQ ID NO: 2 is SEQ ID NO: 19 (FPSSGE). In some embodiments, SEQ ID NO: 3 is SEQ ID NO: 17 (QAFYYFDY). In some embodiments, SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS).
  • SEQ ID NO: 1 is SEQ ID NO: 15
  • SEQ ID NO: 2 is SEQ ID NO: 16
  • SEQ ID NO: 3 is SEQ ID NO: 17.
  • SEQ ID NO: 3 is SEQ ID NO: 18 (QAFYYFDS).
  • SEQ ID NO: 14 is SEQ ID NO: 32 (YIFPSNGETSYNQKFKG).
  • SEQ ID NO: 14 is SEQ ID NO: 33 (YIFPSSGETSYNQKFKG).
  • SEQ ID NO: 1 is SEQ ID NO: 15, SEQ ID NO: 2 is SEQ ID NO: 16 and SEQ ID NO: 3 is SEQ ID NO: 18. In some embodiments, SEQ ID NO: 1 is SEQ ID NO: 15, SEQ ID NO: 2 is SEQ ID NO: 19 and SEQ ID NO: 3 is SEQ ID NO: 17. In some embodiments, SEQ ID NO: 1 is SEQ ID NO: 15, SEQ ID NO: 2 is SEQ ID NO: 19 and SEQ ID NO: 3 is SEQ ID NO: 18. In some embodiments, SEQ ID NO: 1 is SEQ ID NO: 20, SEQ ID NO: 2 is SEQ ID NO: 16 and SEQ ID NO: 3 is SEQ ID NO: 17.
  • SEQ ID NO: 1 is SEQ ID NO: 20
  • SEQ ID NO: 2 is SEQ ID NO: 16
  • SEQ ID NO: 3 is SEQ ID NO: 18.
  • SEQ ID NO: 1 is SEQ ID NO: 20
  • SEQ ID NO: 19 is SEQ ID NO: 17
  • the antibody or antigen binding fragment is 5E5.
  • SEQ ID NO: 1 is SEQ ID NO: 20
  • SEQ ID NO: 2 is SEQ ID NO: 19 and SEQ ID NO: 3 is SEQ ID NO: 18.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain.
  • the heavy chain is an IgG heavy chain.
  • the IgG is IgG4.
  • the heavy chain comprises an IgG4 constant region.
  • the IgG4 constant region comprises a sequence with at least 70, 75, 80, 85, 90, 95, 93, 95, 97, 99 or 100% identity to ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAV LQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAP EFEGGPSVFLFSPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNA KTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQ PREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL DSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK (SEQ ID NO: 21).
  • the IgG4 constant region comprises a sequence with at least 80% identity to SEQ ID NO: 21. In some embodiments, the IgG4 constant region comprises a sequence with at least 90% identity to SEQ ID NO: 21. In some embodiments, the IgG4 constant region does not induce ADCC or CDC. In some embodiments, the IgG4 constant region comprises a mutation of serine 124 to proline (S124P). In some embodiments, the numbering is with respect to SEQ ID NO: 21. In some embodiments, S124P is S241P, and the numbering is with respect to a full heavy chain.
  • the IgG4 constant region comprises a mutation of leucine 131 to glutamic acid (L131E).
  • the numbering is with respect to SEQ ID NO: 21.
  • L131E is L248E, and the numbering is with respect to a full heavy chain. It will be understood that a depending on deletions or insertions into the full heavy chain the numbering provided for these two mutations may be shifted slightly.
  • the antibody or antigen binding fragment comprises a light chain.
  • the light chain is kappa light chain.
  • the light chain is lambda light chain.
  • the antibody or antigen binding fragment comprises a kappa constant region.
  • the kappa constant region comprises a sequence with at least 70, 75, 80, 85, 90, 95, 93, 95, 97, 99 or 100% identity to
  • the kappa constant region comprises a sequence with at least 80% identity to SEQ ID NO: 22. In some embodiments, the kappa constant region comprises a sequence with at least 90% identity to SEQ ID NO: 22.
  • the antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of EVQLQQSGPELVKPGASVKISCKASGYSFTGFYIDWVKQSPGKSLEWIGYIFPSNGE TSYNQKFKGKATLTVDKSSSTVNMQLNSLTSEDSAVYYCARQAFYYFDYWGQGT TLTVSS (SEQ ID NO: 34).
  • the antibody or antigen binding fragment comprises a light chain variable region comprising or consisting of the amino acid sequence of
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 34 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 35.
  • the anti-ILT3 antibody or antigen binding fragment is a mouse antibody. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is mouse 5E5.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of EVQLQQSGPELVKPGASVKISCKASGYSFTGFYIDWVKQSPGKSLEWIGYIFPSNGE TSYNQKFKGKATLTVDKSSSTVNMQLNSLTSEDSAVYYCARQAFYYFDYWGQGT TLTVSS (SEQ ID NO: 36).
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain variable region comprising or consisting of the amino acid sequence of
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 36 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 37.
  • the anti-ILT3 antibody or antigen binding fragment is a chimeric antibody. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is chimeric 5E5.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSS GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSS (SEQ ID NO: 38).
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain variable region comprising or consisting of the amino acid sequence of
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 38 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is humanized 5E5.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFTGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSS (SEQ ID NO: 40).
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain variable region comprising or consisting of the amino acid sequence of
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 40 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 41.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSS (SEQ ID NO: 42).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 42 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 41.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 42 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFTGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDSWGQ GTTVTVSS (SEQ ID NO: 43).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 43 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSS (SEQ ID NO: 44).
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain variable region comprising or consisting of the amino acid sequence of
  • the anti-IET3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 44 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 44 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 45.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFTGFYIDWVKQPPGKGLEWIGYIFPSS GETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSS (SEQ ID NO: 46).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 46 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSSG ETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQG TTVTVSS (SEQ ID NO: 47).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 47 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain variable region comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSS GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDSWGQ GTTVTVSS (SEQ ID NO: 48).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 48 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 39.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain constant region comprising or consisting of the amino acid sequence of EVQLQQSGPELVKPGASVKISCKASGYSFTGFYIDWVKQSPGKSLEWIGYIFPSNGE TSYNQKFKGKATLTVDKSSSTVNMQLNSLTSEDSAVYYCARQAFYYFDYWGQGT TLTVSSAKTTPPSVYPLAPGSAAQTNSMVTLGCLVKGYFPEPVTVTWNSGSLSSGV HTFPAVLQSDLYTLSSSVTVPSSTWPSETVTCNVAHPASSTKVDKKIVPRDCGCKP CICTVPEVSSVFIFPPKPKDVLTITLTPKVTCVVVDISKDDPEVQFSWFVDDVEVHT AQTQPREEQFNSTFRSVSELPIMHQDWLNGKEFKCRVNSAAFPAPIEKTISKTKGRP KAPQVYTIPPPKEQMAKDKVSL
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain comprising or consisting of the amino acid sequence of DIVMSQSPSSLAVSVGEKVTMSCKSSQSLLSSSNQKNYLAWYQQKPGQSPKLLIY WASTRESGVPDRFAGSGSGTDFTLTISSVKAEDLAVYYCQQYYSYPLTFGAGTKL ELKRAD AAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVL NSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC (SEQ ID NO: 8).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of SEQ ID NO: 7 and a light chain comprising or consisting of SEQ ID NO: 8. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a mouse antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of EVQLQQSGPELVKPGASVKISCKASGYSFTGFYIDWVKQSPGKSLEWIGYIFPSNGE TSYNQKFKGKATLTVDKSSSTVNMQLNSLTSEDSAVYYCARQAFYYFDYWGQGT TLTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGV HTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGV EVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTIS KAKGQPREPQVYTLPPS
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain comprising or consisting of the amino acid sequence of DIVMSQSPSSLAVSVGEKVTMSCKSSQSLLSSSNQKNYLAWYQQKPGQSPKLLIY WASTRESGVPDRFAGSGSGTDFTLTISSVKAEDLAVYYCQQYYSYPLTFGAGTKL ELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNS QESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 10).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of SEQ ID NO: 9 and a light chain comprising or consisting of SEQ ID NO: 10. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a chimeric antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSS GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEE
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain comprising or consisting of the amino acid sequence of DIVMTQSPDSLAVSLGERATINCKSSQSLLSSSNQKNYLAWYQQKPGQAPRLLIYW ASTRESGVPDRFAGSGSGTDFTLTISSLQAEDVAVYYCQQYYSYPLTFGQGTKLEI KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQE SVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 12).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of SEQ ID NO: 11 and a light chain comprising or consisting of SEQ ID NO: 12. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFTGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRATMTVDKSTSTVYMEESSERSEDTAVYYCARQAFYYFDYWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEEMTK
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain comprising or consisting of the amino acid sequence of DIVMTQSPDSLAVSLGERATINCKSSQSLLSSSNQKNYLAWYQQKPGQPPKLLIYW ASTRESGVPDRFAGSGSGTDFTLTISSLQAEDVAVYYCQQYYSYPLTFGQGTKLEI KRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQE SVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 24).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of SEQ ID NO: 23 and a light chain comprising or consisting of SEQ ID NO: 24. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEE
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of SEQ ID NO: 25 and a light chain comprising or consisting of SEQ ID NO: 24. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 25 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 12.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFTGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDSWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEE
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 26 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 12. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSN GETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEEMT
  • the anti-ILT3 antibody or antigen binding fragment comprises a light chain comprising or consisting of the amino acid sequence of DIVMTQSPDSLAVSLGERATINCKSSQSLLSSSNQKNYLAWYQQKPGQPPKLLIYW ASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYSYPLTFGQGTKLEIK RTVAAPSVFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWKVDNAEQSGNSQES VTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 28).
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 27 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 12. In some embodiments, the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 27 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 28. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFTGFYIDWVKQPPGKGLEWIGYIFPSS GETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEEMT
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 29 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 12.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of EVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSSG ETSYNQKFKGRATMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDYWGQG TTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG VHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPP CPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDG VEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTI SKAKGQPREPQVYTLPPSQEEMT
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 30 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 12.
  • the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising or consisting of the amino acid sequence of QVQLVQSGAEVKKPGASVKVSCKASGYSFSGFYIDWVKQPPGKGLEWIGYIFPSS GETSYNQKFKGRVTMTVDKSTSTVYMELSSLRSEDTAVYYCARQAFYYFDSWGQ GTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTS GVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEE
  • the anti-ILT3 antibody or antigen binding fragment comprises a heavy chain comprising a variable region comprising or consisting of SEQ ID NO: 31 and a light chain comprising a variable region comprising or consisting of SEQ ID NO: 12. In some embodiments, the anti-ILT3 antibody or antigen binding fragment is a humanized antibody.
  • the antibody is the 5E5 antibody. In some embodiments, the 5E5 antibody is the humanized 5E5 antibody. In some embodiments, the 5E5 antibody is an antibody wherein CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 20, CDR-H2 comprises the amino acid sequence as set forth in SEQ ID NO: 19, CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 17, CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 4, CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5, and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 6. In some embodiments, the antibody is an antibody that competed with the 5E5 antibody for binding to ILT3.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein: CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS), CDR- H2 comprises the amino acid sequence as set forth in SEQ ID NO: 50 (AITFGGGNTYYPDSVKG), CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 51 (HGDGNYDFYAMDY), CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 52 (KSSQSLLNSGNQKNYLT), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 5 (WASTRES), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 53 (QNDYSYPLT).
  • the anti-ILT3 antibody comprising CDRS: of SEQ ID NO: 49-53 and 5 is 3F6
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region comprising the amino acid sequence EVKLVESGGGLVKPGGSLKLSCAASGFTFSSYAMSWVRQTPAKRLEWVAAITFGG GNTYYPDSVKGRFTISRDNARNTLYLQMSSLRSEDTAMYYCARHGDGNYDFYAM DYWGQGTSVTVSS (SEQ ID NO: 54).
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region consisting of SEQ ID NO: 54.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a light chain variable region comprising the amino acid sequence DIVMTQSPSSLTVTAGEKVTMSCKSSQSLLNSGNQKNYLTWYQQKPGQPPKLLIY WASTRESGVPDRFTGSGSRTDFTLTISRVQAEDLAVYYCQNDYSYPLTFGAGTKLE LK (SEQ ID NO: 55).
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a light chain variable region consisting of SEQ ID NO: 55.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region comprising or consisting of SEQ ID NO: 54 and a light chain variable region comprising or consisting of SEQ ID NO: 55.
  • anti- ILT3 antibody 3F6 comprises a heavy chain variable region comprising or consisting of SEQ ID NO: 54 and a light chain variable region comprising or consisting of SEQ ID NO: 55.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein: CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 49 (SYAMS), CDR- H2 comprises the amino acid sequence as set forth in SEQ ID NO: 56 (TISSDGGNTYYTDSVKG), CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 57 (HDGRGALDY), CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 58 (RASQDISNYLN), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 59 (YTSRLHS), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 60 (QQGNTLPWT).
  • the anti-ILT3 antibody comprising CDRS: of SEQ ID NO: 56-60 and 49 is 7A5.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region comprising the amino acid sequence EVKLVESGGGLVKPGGSLKLSCAASGFTFSSYAMSWVRQTPAKRLEWVATISSDG GNTYYTDSVKGRFTISRDNARNTLDLQMSSLRSEDTAMYYCARHDGRGALDYW GQGTSVTVSS (SEQ ID NO: 61).
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region consisting of SEQ ID NO: 61.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a light chain variable region comprising the amino acid sequence DIQMTQTTSSLSASLGDRVTISCRASQDISNYLNWYQQKPDGTVKLLIYYTSRLHS GVPSRFSGSGSGTDYSLTISNLEQEDIATYFCQQGNTLPWTFGGGTKLEIK (SEQ ID NO: 62).
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a light chain variable region consisting of SEQ ID NO: 62.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region comprising or consisting of SEQ ID NO: 61 and a light chain variable region comprising or consisting of SEQ ID NO: 62.
  • anti-ILT3 antibody 7A5 comprises a heavy chain variable region comprising or consisting of SEQ ID NO: 61 and a light chain variable region comprising or consisting of SEQ ID NO: 62.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises three heavy chain CDRs (CDR-H) and three light chain CDRs (CDR-L), wherein: CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 63 (NSAVH), CDR- H2 comprises the amino acid sequence as set forth in SEQ ID NO: 64 (VIWAGGNTNYNSTLMS), CDR-H3 comprises the amino acid sequence as set forth in SEQ ID NO: 65 (HETYGDSFDY), CDR-L1 comprises the amino acid sequence as set forth in SEQ ID NO: 66 (RSSQSLLDSDGKTYLN), CDR-L2 comprises the amino acid sequence as set forth in SEQ ID NO: 67 (LVSKLDS), and CDR-L3 comprises the amino acid sequence as set forth in SEQ ID NO: 68 (WQGTHFPFT).
  • CDR-H1 comprises the amino acid sequence set forth in SEQ ID NO: 63 (NSAVH)
  • CDR- H2 comprises the amino acid sequence
  • the anti-ILT3 antibody comprising CDRS: of SEQ ID NO: 63-68 is 10G12.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region comprising the amino acid sequence QVQLKESGPVLVAPSQSLSITCTVSGFSLTNSAVHWVRQPPGKGLEWLGVIWAGG NTNYNSTLMSRLTINKDNSKSQVFLRMNSLQTDDTAIYYCAKHETYGDSFDYWG QGTTLTVSS (SEQ ID NO: 69).
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region consisting of SEQ ID NO:
  • the antibody or antigen binding fragment thereof comprises a light chain variable region comprising the amino acid sequence DVVMTQTPLTLSVTIGQPASISCRSSQSLLDSDGKTYLNWLFQRPGQSPKRLIYLVS KLDSGVPDRFTGSGSGTDFTLKISRVEAEDLGVYYCWQGTHFPFTFGSGTKLEIK (SEQ ID NO: 70).
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a light chain variable region consisting of SEQ ID NO: 70.
  • the anti-ILT3 antibody or antigen binding fragment thereof comprises a heavy chain variable region comprising or consisting of SEQ ID NO: 69 and a light chain variable region comprising or consisting of SEQ ID NO: 70.
  • anti- ILT3 antibody 10G12 comprises a heavy chain variable region comprising or consisting of SEQ ID NO: 69 and a light chain variable region comprising or consisting of SEQ ID NO:
  • the antibody or antigen binding fragment thereof is capable of binding ILT3.
  • the antibody or antigen binding fragment thereof is an anti-ILT3 antibody.
  • the target antigen of the antibody is ILT3.
  • ILT3 is membrane ILT3 (mILT3).
  • ILT3 is soluble ILT3 (sILT3).
  • An “anti-ILT3 antibody”, “an antibody which recognizes ILT3”, or “an antibody against ILT3” is an antibody that binds ILT3, with sufficient affinity and specificity.
  • the antibody has increased binding to ILT3.
  • the antibody has increased binding to ILT3 as compared to a commercially available ILT3 antibody.
  • the commercially available ILT3 antibody is selected from ZM4.1, h52B8 and 10202. In some embodiments, the ILT3 antibody is selected from ZM4.1, h52B8, 10202 and NGM831. In some embodiments, the anti-ILT3 antibody is h52B8. H52B8 is an anti-ILT3 antibody developed by Merck. In some embodiments, the anti-ILT3 antibody is 10202. 10202 is an anti-LILRB4 antibody developed by Immune-One Therapeutics. In some embodiments, the anti-ILT3 antibody is NGM83 1. NGM831 is an anti-LILRB4 antibody developed by NGM. In some embodiments, the antibody or antigen-binding fragment thereof does not significantly bind to any other target.
  • any other target is any other ILT protein.
  • ILT3 is mammalian ILT3.
  • ILT3 is human ILT3.
  • mammalian is monkey.
  • monkey is selected from rhesus and cynomolgus.
  • monkey is cynomolgus.
  • mammalian is human.
  • mammalian is not murine.
  • the antibody does not bind murine ILT3.
  • the antibody binds human and monkey ILT3.
  • membranal ILT3 is on the cell surface.
  • the membrane is the plasma membrane.
  • the membrane is the membrane of an immune cell.
  • the immune cell is selected from myeloid cells, dendritic cells, and macrophages.
  • the immune cell is a myeloid cell.
  • the immune cell is a dendritic cell.
  • the immune cell is a macrophage.
  • the immune cell is a monocyte.
  • the myeloid cell is a myeloid derived suppressor cell (MDSC).
  • the dendritic cell is a tolerogenic dendritic cell.
  • the macrophage is a suppressive macrophage.
  • the macrophage is an M2 macrophage.
  • an M2 macrophage is a suppressive macrophage.
  • the immune cell is a tumor associated immune cell.
  • the immune cell is a tumor infiltrating immune cell.
  • the macrophage is a tumor associated macrophage (TAM).
  • TAM tumor associated macrophage
  • the immune cell is selected from a monocyte and macrophage.
  • the immune cell is selected from a monocyte, a macrophage and a dendritic cell.
  • the contacted immune cell enhances immune cell activation of a cytotoxic immune cell.
  • enhancing activation comprises enhancing cytotoxicity.
  • cytotoxicity is toxicity against the cancer or cancer cells.
  • a cytotoxic immune cell is selected from a T cell and a natural killer (NK) cell.
  • the T cell is a CD8 T cell.
  • the cytotoxic immune cell is an NK cell.
  • the cytotoxic immune cell is a CD8 T cell.
  • the method further comprises contacting the immune cell and cancer cell with a cytotoxic immune cell.
  • a cytotoxic immune cell is a cell capable of directly killing cancer cells.
  • a cytotoxic immune cell is a macrophage.
  • a cytotoxic immune cell is not a macrophage.
  • an anti-ILT3 antibody is an antibody that binds to the ILT3, with sufficient affinity and specificity.
  • an anti-ILT3 antibody has ILT3 as the antigen to which it binds.
  • An anti-CD20 antibody is an antibody that binds to the CD20, with sufficient affinity and specificity.
  • an anti-CD20 antibody has CD20 as the antigen to which it binds.
  • an “anti-EGFR antibody”, “an EGFR antibody”, “an antibody which recognizes EGFR”, or “an antibody against EGFR” is an antibody that binds to the EGFR, with sufficient affinity and specificity.
  • an anti-EGFR antibody has EGFR as the antigen to which it binds.
  • An “anti-HER2 antibody”, “a HER2 antibody”, “an antibody which recognizes HER2”, or “an antibody against HER2” is an antibody that binds to the HER2, with sufficient affinity and specificity.
  • an anti-HER2 antibody has HER2 as the antigen to which it binds.
  • an “anti- CD47 antibody”, “a CD47 antibody”, “an antibody which recognizes CD47”, or “an antibody against CD47” is an antibody that binds to the CD47, with sufficient affinity and specificity.
  • an anti-CD47 antibody has CD47 as the antigen to which it binds.
  • An “anti-PD-Ll antibody”, “an antibody which recognizes PD-L1”, or “an antibody against PD- Ll” is an antibody that binds to the PD-L1, with sufficient affinity and specificity.
  • an anti-PD-Ll antibody has PD-L1 as the antigen to which it binds.
  • an “anti- CLDN18 antibody”, “a CLDN18 antibody”, “an antibody which recognizes CLDN18”, or “an antibody against CLDN18” is an antibody that binds to the CLDN18, with sufficient affinity and specificity.
  • an anti-CLDN18 antibody has CLDN18 as the antigen to which it binds.
  • An “anti-PD-1 antibody”, “an antibody which recognizes PD- 1", or “an antibody against PD-1” is an antibody that binds to the PD-1, with sufficient affinity and specificity.
  • an anti-PD-1 antibody has PD-1 as the antigen to which it binds.
  • an "antigen” is a molecule or a portion of a molecule capable of eliciting antibody formation and being bound by an antibody.
  • An antigen may have one or more than one epitope. The specific reaction referred to above is meant to indicate that the antigen will react, in a highly selective manner, with its corresponding antibody and not with the multitude of other antibodies which may be evoked by other antigens.
  • the term "antigenic determinant” or “epitope” according to the invention refers to the region of an antigen molecule that specifically reacts with particular antibody.
  • Peptide sequences derived from an epitope can be used, alone or in conjunction with a carrier moiety, applying methods known in the art, to immunize animals and to produce additional polyclonal or monoclonal antibodies.
  • Immunoglobulin variable domains can also be analyzed using the IMGT information system (www://imgt. cines.fr/) (IMGT®/V-Quest) to identify variable region segments, including CDRs. See, e.g., Brochet, X. et al, Nucl. Acids Res. J6:W503-508 (2008).
  • Kabat et al. also defined a numbering system for variable domain sequences that is applicable to any antibody.
  • One of ordinary skill in the art can unambiguously assign this system of "Kabat numbering" to any variable domain sequence, without reliance on any experimental data beyond the sequence itself.
  • Kabat numbering refers to the numbering system set forth by Kabat et al, U.S. Dept, of Health and Human Services, "Sequence of Proteins of Immunological Interest" (1983).
  • the agent is an antibody or antigen binding fragment thereof. In some embodiments, the antigen binding fragment is a Fab fragment. In some embodiments, the antibody is a single domain antibody. In some embodiments, the anti-ILT3 antibody is a single domain antibody. In some embodiments, the single domain antibody is a camelid or shark antibody. In some embodiments, the single domain antibody is a camelid antibody. In some embodiments, the single domain antibody is a VHH. In some embodiments, the antibody lacks a Fc domain. In some embodiments, the agent is an antigen binding domain that lacks an Fc domain. In some embodiments, the agent is a single-domain antibody. In some embodiments, the agent is a camelid, shark or nanobody.
  • the antibody or fragment is fused to another protein or fragment of a protein.
  • the second protein or fragment increases half-life, particularly in serum.
  • the half-life extending protein is human serum albumin.
  • the agent is modified by a chemical that produces a modification that enhances half-life.
  • the modification is PEGylation and the chemical is polyethylene glycol.
  • the binding of the ILT3 antibody or antigen binding domain to a cell does not kill the cell. In some embodiments, the binding of the ILT3 antibody or antigen binding domain to a cell does not lead to death of the cell. In some embodiments the ILT3 antibody or antigen binding domain does not induce antibody dependent cell-mediated cytotoxicity (ADCC). In some embodiments, the ILT3 antibody or antigen binding domain does not induce complement-dependent cytotoxicity (CDC). In some embodiments, the ILT3 antibody or antigen binding domain does not induce ADCC and/or CDC. In some embodiments, the ILT3 antibody or antigen binding domain comprises an IgG2 or IgG4 domain.
  • the ILT3 antibody or antigen binding domain comprises an IgG2 domain. In some embodiments, the ILT3 antibody or antigen binding domain comprises an IgG4 domain. In some embodiments, the ILT3 antibody or antigen binding domain comprises an IgGl or IgG3 mutated to reduce cell death mediated by binding of the antibody. In some embodiments, the mutation mutates an Fc receptor binding domain. In some embodiments, a Fc domain of the ILT3 antibody is engineered or mutated to decrease CDC, ADCC or both. Fc engineering is well known in the art, and any mutation or amino acid change that is known to decrease antibody mediated cell killing may be used.
  • the ILT3 antibody or antigen binding domain does not comprise IgGl and/or IgG3. In some embodiments, the ILT3 antibody or antigen binding domain does not induce antibody-dependent cell-mediated cytotoxicity (ADCC). In some embodiments the ILT3 antibody or antigen binding domain does not induce complementdependent cytotoxicity (CDC). In some embodiments, the ILT3 antibody or antigen binding domain comprises an IgGl or IgG3 comprising a mutation that reduces ADCC, CDC or both induced by the antibody’s binding. In some embodiments, the mutation reduces the ADCC, CDC or both to nothing. ADCC and CDC are well characterized and antibody sequences that allow for these cytotoxic pathways to be induced are well known. Mutations, such as for non-limiting examples, mutation of IgGl or IgG3 to IgG2 or IgG4 are well known. Any such mutation may be used in the backbone of the antibodies of the invention.
  • binding ILT3 inhibits binding of ILT3 to a ligand.
  • an ILT3 inhibitor inhibits binding of ILT3 to a ligand.
  • binding ILT3 inhibits binding of a ligand to ILT3.
  • the ligand is an ILT3 ligand.
  • the ligand is Apolipoprotein E (APOE).
  • the ligand is fibronectin (FN1).
  • the ligand is APOE and FN 1.
  • the ligand inhibits binding of ILT3 to both APOE and FN 1.
  • the inhibitor inhibits binding of ILT3 to FN1.
  • the inhibitor inhibits binding of ILT3 to APOE. In some embodiments, the inhibitor inhibits binding of ILT3 to both FN 1 and APOE. In some embodiments, the APOE is present in the tumor microenvironment (TME). In some embodiments, the FN1 is present in the TME. In some embodiments, the APOE is present in extracellular matrix. In some embodiments, the FN1 is present in extracellular matrix. In some embodiments, the extracellular matrix is tumor extracellular matrix. In some embodiments, the tumor’s microenvironment comprises FN1. In some embodiments, the tumor’s microenvironment comprises APOE. In some embodiments, the tumor’s microenvironment comprises FN1 and/or APOE.
  • the method comprises blocking binding of ILT3 to at least one of its ligands.
  • the ligand is FN 1.
  • the ligand is FN1 and APOE.
  • the method comprises blocking binding of ILT3 to FN1.
  • the method comprises blocking binding of ILT3 to FN1 and APOE.
  • the method comprises administering the therapeutic antibody.
  • the method comprises administering the IgGl or IgG3 antibody.
  • blocking binding comprises administering to the subject an antibody or antigen binding fragment thereof that binds to ILT3.
  • the antibody or antigen binding fragment thereof is an immune checkpoint inhibitor (ICI).
  • the antibody or antigen binding fragment is an ILT3 blocking antibody.
  • binding to ILT3 releases immune cells from ILT3-mediated inhibition.
  • inhibition is immunosuppression.
  • the immune cell is a dendritic cell.
  • the immune cell is T cells.
  • the T cell is a CD8 T cell.
  • the T cell is a CD4 T cell.
  • binding of ILT3 releases T cells from ILT3-mediated inhibition.
  • binding of ILT3 releases monocytes from ILT3-mediated inhibition. In some embodiments, binding of ILT3 releases macrophages from ILT3-mediated inhibition. In some embodiments, binding of ILT3 releases dendritic cells from ILT3 -mediated inhibition.
  • the immune cell is a macrophage. In some embodiments, the macrophages are MO macrophages. In some embodiments, a MO macrophage is a non-activated macrophage. In some embodiments, a MO macrophage is a naive macrophage. In some embodiments, the macrophages are Ml macrophages. In some embodiments, a Ml macrophage is an inflammatory macrophage.
  • a Ml macrophage is a pro -inflammatory macrophage.
  • the macrophages are M2 macrophages.
  • a M2 macrophage is an anti-inflammatory macrophage.
  • a M2 macrophage is a suppressive macrophage.
  • a M2 macrophage is a tolerogenic macrophage.
  • the monocytes are macrophages.
  • the macrophages are immature macrophages.
  • the macrophages are inflammatory macrophages.
  • the macrophages are tolerogenic macrophages.
  • release is decrease. In some embodiments, decrease is a decrease of at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 97, 99 or 100%. Each possibility represents a separate embodiment of the invention.
  • release comprises immune cell activation. In some embodiments, activation is increased activation. In some embodiments, increased is an increase of at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 97, 100, 125, 150, 175, 200, 250, 300, 350, 400, 450 or 500%. Each possibility represents a separate embodiment of the invention.
  • dendritic cell activation is increased.
  • T cell activation is increased.
  • monocyte activation in increased.
  • macrophage activation is increased.
  • increasing T cell activation comprises restoring T cell activation suppressed by MDSCs.
  • increasing T cell activation comprises restoring T cell activation suppressed by tolerogenic DCs (DCtols).
  • DCtols tolerogenic DCs
  • activation is Fc receptor mediated activation.
  • increasing DC activation comprises restoring DC activation suppressed by fibronectin or APOE.
  • increasing monocyte activation comprises restoring monocyte activation suppressed by fibronectin or APOE.
  • release comprises increased proliferation. In some embodiments, activation comprises increased proliferation. In some embodiments, release comprises increased pro -inflammatory cytokine secretion. In some embodiments, release comprises increased pro-inflammatory chemokine secretion. In some embodiments, activation comprises increased pro -inflammatory cytokine secretion. In some embodiments, activation comprises increased pro-inflammatory chemokine secretion. In some embodiments, the pro-inflammatory cytokine is selected from TNFa, IFNg, IL- IB, M-CSF, IL-6 and IL-8. In some embodiments, the pro-inflammatory cytokine is tumor necrosis factor alpha (TNFa).
  • TNFa tumor necrosis factor alpha
  • the pro-inflammatory cytokine is interferon gamma (IFNg). In some embodiments, the pro -inflammatory cytokine is interleukin 1 beta (IL- IB). In some embodiments, the pro-inflammatory cytokine is colony stimulating factor 1 (CSF1/M-CSF). In some embodiments, the pro -inflammatory cytokine is IL-6. In some embodiments, the pro -inflammatory cytokine is IL-8. In some embodiments, the pro- inflammatory chemokine is selected from CCL2, CCL3, CXCL1 and CCL4. In some embodiments, the pro-inflammatory chemokine is chemokine (C-C motif) 2 (CCL2).
  • C-C motif CCL2
  • the pro-inflammatory chemokine is CCL3. In some embodiments, the pro-inflammatory chemokine is chemokine (C-X-C motif) ligand 1 (CXCL1). In some embodiments, the pro-inflammatory chemokine is CCL4.
  • the inhibitor is an antibody or antigen binding fragment thereof that competes with an anti-ILT3 antibody or antigen binding fragment described herein for binding to ILT3.
  • Antibody competition assays are well known in the art and any such method may be used to determine antibody competition.
  • competing for binding comprises binding the same epitope.
  • Methods of determining the epitope to which an antibody binds are well known in the art and any such method can be used for determining the epitope to which the antibody of the invention binds.
  • the epitope is within a ligand binding domain.
  • the epitope is sufficiently close to a ligand binding domain so that binding of the antibody or antigen binding domain occludes, blocks or alters the ligand binding domain.
  • the epitope comprises low homology to the corresponding mouse sequence of ILT3.
  • the antibody is an anti-CD20 antibody.
  • the anticancer agent is an anti-CD20 antibody.
  • the antibody binds to CD20.
  • the antibody is specific to CD20.
  • binds to is specifically binds to.
  • the antibody is a CD20 blocking antibody.
  • the antibody is a CD20 inhibitory antibody.
  • the antibody is a CD20 antagonist.
  • Therapeutic anti-CD20 antibodies are well known in the art and any such antibodies may be used in the method of the invention.
  • the anti-CD20 antibody is selected from rituximab, ibritumomab, ofatumumab, obinutuzumab, ocrelizumab, veltuzumab, ublituximab and ocaratuzumab.
  • the anti-CD20 antibody is rituximab.
  • the anti- CD20 antibody comprises an IgGl or IgG3 domain.
  • the antibody or antigen binding domain comprises an IgGl domain.
  • the antibody or antigen binding domain comprises an IgG3 domain.
  • the anti-CD20 antibody is an IgGl antibody.
  • the anti-CD20 antibody is an IgG3 antibody.
  • the anti-CD20 antibody induces ADCC.
  • the anti-CD20 antibody induces ADCP.
  • the antibody is an anti-EGFR antibody.
  • the anticancer agent is an anti-EGFR antibody.
  • the antibody binds to EGFR.
  • the antibody is specific to EGFR.
  • binds to is specifically binds to.
  • the antibody is an EGFR blocking antibody.
  • the antibody is an EGFR inhibitory antibody.
  • the antibody is an EGFR antagonist. Therapeutic anti- EGFR antibodies are well known in the art and any such antibodies may be used in the method of the invention.
  • the anti-EGFR antibody is selected from cetuximab, panitumumab, nimotuzumab, and necitumumab. In some embodiments, the anti- EGFR antibody is cetuximab. In some embodiments, the anti-EGFR antibody comprises an IgGl or IgG3 domain. In some embodiments, the antibody or antigen binding domain comprises an IgGl domain. In some embodiments, the antibody or antigen binding domain comprises an IgG3 domain. In some embodiments, the anti-EGFR antibody is an IgGl antibody. In some embodiments, the anti-EGFR antibody is an IgG3 antibody. In some embodiments, the anti-EGFR antibody induces ADCC. In some embodiments, the anti- EGFR antibody induces ADCP.
  • the antibody is an anti-HER2 antibody.
  • the anticancer agent is an anti-HER2 antibody.
  • the antibody binds to HER2.
  • the antibody is specific to HER2.
  • binds to is specifically binds to.
  • the antibody is an HER2 blocking antibody.
  • the antibody is a HER2 inhibitory antibody.
  • the antibody is a HER2 antagonist.
  • Therapeutic anti-HER2 antibodies are well known in the art and any such antibodies may be used in the method of the invention.
  • the anti-HER2 antibody is selected from trastuzumab, pertuzumab, and margetuximab.
  • the anti-HER2 antibody is trastuzumab. In some embodiments, the anti-HER2 antibody comprises an IgGl or IgG3 domain. In some embodiments, the antibody or antigen binding domain comprises an IgGl domain. In some embodiments, the antibody or antigen binding domain comprises an IgG3 domain. In some embodiments, the anti-HER2 antibody is an IgGl antibody. In some embodiments, the anti- HER2 antibody is an IgG3 antibody. In some embodiments, the anti-HER2 antibody induces ADCC. In some embodiments, the anti-HER2 antibody induces ADCP. [0230] In some embodiments, the antibody is an anti-CD47 antibody. In some embodiments, the anticancer agent is an anti-CD47 antibody.
  • the antibody binds to CD47. In some embodiments, the antibody is specific to CD47. In some embodiments, binds to is specifically binds to. In some embodiments, the antibody is a CD47 blocking antibody. In some embodiments, the antibody is a CD47 inhibitory antibody. In some embodiments, the antibody is a CD47 antagonist. Therapeutic anti-CD47 antibodies are well known in the art and any such antibodies may be used in the method of the invention. In some embodiments, the anti-CD47 antibody is selected from magrolimab, and letaplimab. In some embodiments, the anti-CD47 antibody is magrolimab. In some embodiments, the anti-CD47 antibody comprises an IgGl or IgG3 domain.
  • the antibody or antigen binding domain comprises an IgGl domain. In some embodiments, the antibody or antigen binding domain comprises an IgG3 domain. In some embodiments, the anti-CD47 antibody is an IgGl antibody. In some embodiments, the anti-CD47 antibody is an IgG3 antibody. In some embodiments, the anti-CD47 antibody induces ADCC. In some embodiments, the anti- CD47 antibody induces ADCP.
  • the antibody is an anti-PD-Ll antibody.
  • the anticancer agent is an anti-PD-Ll antibody.
  • the antibody binds to PD-L1.
  • the antibody is specific to PD-L1.
  • binds to is specifically binds to.
  • the antibody is a PD- L1 blocking antibody.
  • the antibody is a PD-L1 inhibitory antibody.
  • the antibody is a PD-L1 antagonist.
  • Therapeutic anti-PD-Ll antibodies are well known in the art and any such antibodies may be used in the method of the invention.
  • the anti-PD-Ll antibody is selected from avelumab, atezolizumab and durvalumab. In some embodiments, the anti-PD-Ll antibody is selected from avelumab and an antibody comprising the CDRs of atezolizumab or durvalumab and which induces ADCC, ADCP or both. It is well known in the art that atezolizumab comprises a mutation that results in a lack of glycosylation. This absent/reduced glycosylation results in decreased ADCC/ADCP. Similarly, durvalumab comprises a triple mutation that abrogates binging to FcyR and thus blocks ADCC/ADCP.
  • Avelumab is known to induce ADCC/ADCP and thus is superior to other known anti-PD-Ll antibodies for combination with ILT3 blocking.
  • anti-PD-Ll antibodies have been generated with the CDRs of atezolizumab and durvalumab but which lack the mutations and thus do induce ADCC/ADCP.
  • the anti-PD-Ll antibody is avelumab.
  • the anti-PD-Ll antibody is an antibody with the CDRs of atezolizumab or durvalumab.
  • Other anti-PD-Ll inhibitors include KN035, cosibelimab, AUNP12, CA-170 and BMS-986189.
  • the anti- PD-L1 antibody comprises an IgGl or IgG3 domain. In some embodiments, the antibody or antigen binding domain comprises an IgGl domain. In some embodiments, the antibody or antigen binding domain comprises an IgG3 domain. In some embodiments, the anti-PD-Ll antibody is an IgGl antibody. In some embodiments, the anti-PD-Ll antibody is an IgG3 antibody. In some embodiments, the anti- PD-Ll antibody induces ADCC. In some embodiments, the anti-PD-Ll antibody induces ADCP.
  • the antibody is an anti-CLDN18 antibody.
  • the anticancer agent is an anti-CLDN18 antibody.
  • the antibody binds to CLDN18.
  • the antibody is specific to CLDN18.
  • binds to is specifically binds to.
  • the antibody is an CLDN18 blocking antibody.
  • the antibody is an CLDN18 inhibitory antibody.
  • the antibody is an CLDN18 antagonist.
  • Therapeutic anti-CLDN18 antibodies are well known in the art and any such antibodies may be used in the method of the invention.
  • the anti-CLDN18 antibody is zolbetuximab.
  • the anti-CLDN18 antibody comprises an IgGl or IgG3 domain. In some embodiments, the antibody or antigen binding domain comprises an IgGl domain. In some embodiments, the antibody or antigen binding domain comprises an IgG3 domain. In some embodiments, the anti-CLDN18 antibody is an IgGl antibody. In some embodiments, the anti-CLDN18 antibody is an IgG3 antibody. In some embodiments, the anti-CLDN18 antibody induces ADCC. In some embodiments, the anti-CLDN18 antibody induces ADCP.
  • the antibody is an anti-PD-1 antibody. In some embodiments, the antibody binds to PD-1. In some embodiments, the antibody is specific to PD-1. In some embodiments, binds to is specifically binds to. In some embodiments, the antibody is a PD- 1 blocking antibody. In some embodiments, the antibody is a PD-1 inhibitory antibody. In some embodiments, the antibody is a PD-1 antagonist. Therapeutic anti-PD-1 antibodies are well known in the art and any such antibodies may be used in the method of the invention. In some embodiments, the anti-PD-1 antibody is selected from pembrolizumab, nivolumab, cemiplimab, dostarlimab and retifanlimab.
  • the anti-PD-1 antibody is pembrolizumab.
  • Other anti-PD-1 inhibitors include vopratelimab, spartalizumab, camrelizumab, sintilimab, toripalimab, INCMGA00012, AMP-224, AMP-514 and acrixolimab.
  • the binding of the antibody or antigen binding domain to a cancer cell kills the cancer cell. In some embodiments, the binding of the antibody or antigen binding domain to a cancer cell opsonizes the cancer cell. In some embodiments, the binding of the anticancer agent to a cancer cell kills the cancer cell. In some embodiments, the binding of the antibody or antigen binding domain to a cancer cell leads to death of the cancer cell. In some embodiments, the binding of the anticancer agent to a cancer cell leads to death of the cancer cell. In some embodiments the antibody or antigen binding domain induces antibody dependent cell-mediated cytotoxicity (ADCC). In some embodiments, the antibody or antigen binding domain induces complement-dependent cytotoxicity (CDC).
  • ADCC antibody dependent cell-mediated cytotoxicity
  • CDC complement-dependent cytotoxicity
  • the antibody or antigen binding domain induces ADCC and/or CDC.
  • the antibody or antigen binding domain comprises an IgGl or IgG3 domain. In some embodiments, the antibody or antigen binding domain comprises an IgGl domain. In some embodiments, the antibody or antigen binding domain comprises an IgG3 domain.
  • the anticancer agent comprises an IgGl or IgG3 domain. In some embodiments, the anticancer agent comprises an IgGl domain. In some embodiments, the anticancer agent comprises an IgG3 domain.
  • anti-ILT3 antibody and anti-PD-1 antibody should not induce killing as it is desirous for the immune cells to remain alive and be activated; the anti- CD20/EGFR/HER2/CD47/PD-L1/CLDN18 antibodies all may (but do not necessarily have to) induce killing via their binding.
  • these antibodies and not the anti-ILT3 antibody may be IgGl or IgG3 and induce ADCC and/or CDC.
  • administering the agent comprises administering a composition comprising the agent.
  • administering an inhibitor comprises administering a composition comprising the inhibitor.
  • the composition comprises a therapeutically effective amount of the agent.
  • the composition comprises a therapeutically effective amount of the inhibitor.
  • the composition is a pharmaceutical composition. In some embodiments, the composition is a therapeutic composition. In some embodiments, the composition is a preventative composition. In some embodiments, the composition comprises a therapeutically effective amount of the antibody or antigen binding domain. In some embodiments, the composition comprises a therapeutically acceptable carrier, excipient or adjuvant.
  • terapéuticaally effective amount refers to an amount of a drug effective to treat a disease or disorder in a mammal in combination with the other agent as recited herein.
  • a therapeutically effective amount refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result. The exact dosage form and regimen would be determined by the physician according to the patient's condition.
  • the therapeutically effective amount is the amount effective as a monotherapy.
  • the therapeutically effective amount is the amount effective as a combination therapy as described herein.
  • carrier refers to any component of a pharmaceutical composition that is not the active agent.
  • pharmaceutically acceptable carrier refers to non-toxic, inert solid, semi-solid liquid filler, diluent, encapsulating material, formulation auxiliary of any type, or simply a sterile aqueous medium, such as saline.
  • sugars such as lactose, glucose and sucrose, starches such as corn starch and potato starch, cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt, gelatin, talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, com oil and soybean oil; glycols, such as propylene glycol, polyols such as glycerin, sorbitol, mannitol and polyethylene glycol; esters such as ethyl oleate and ethyl laurate, agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline, Ringer's solution; ethy
  • substances which can serve as a carrier herein include sugar, starch, cellulose and its derivatives, powered tragacanth, malt, gelatin, talc, stearic acid, magnesium stearate, calcium sulfate, vegetable oils, polyols, alginic acid, pyrogen-free water, isotonic saline, phosphate buffer solutions, cocoa butter (suppository base), emulsifier as well as other non-toxic pharmaceutically compatible substances used in other pharmaceutical formulations.
  • Wetting agents and lubricants such as sodium lauryl sulfate, as well as coloring agents, flavoring agents, excipients, stabilizers, antioxidants, and preservatives may also be present.
  • any non- toxic, inert, and effective carrier may be used to formulate the compositions contemplated herein.
  • Suitable pharmaceutically acceptable carriers, excipients, and diluents in this regard are well known to those of skill in the art, such as those described in The Merck Index, Thirteenth Edition, Budavari et al., Eds., Merck & Co., Inc., Rahway, N.J. (2001); the CTFA (Cosmetic, Toiletry, and Fragrance Association) International Cosmetic Ingredient Dictionary and Handbook, Tenth Edition (2004); and the “Inactive Ingredient Guide,” U.S. Food and Drug Administration (FDA) Center for Drug Evaluation and Research (CDER) Office of Management, the contents of all of which are hereby incorporated by reference in their entirety.
  • CTFA Cosmetic, Toiletry, and Fragrance Association
  • Examples of pharmaceutically acceptable excipients, carriers and diluents useful in the present compositions include distilled water, physiological saline, Ringer's solution, dextrose solution, Hank's solution, and DMSO. These additional inactive components, as well as effective formulations and administration procedures, are well known in the art and are described in standard textbooks, such as Goodman and Gillman’s: The Pharmacological Bases of Therapeutics, 8th Ed., Gilman et al. Eds. Pergamon Press (1990); Remington’s Pharmaceutical Sciences, 18th Ed., Mack Publishing Co., Easton, Pa.
  • compositions may also be contained in artificially created structures such as liposomes, ISCOMS, slow-releasing particles, and other vehicles which increase the half-life of the peptides or polypeptides in serum.
  • liposomes include emulsions, foams, micelles, insoluble monolayers, liquid crystals, phospholipid dispersions, lamellar layers and the like.
  • Liposomes for use with the presently described peptides are formed from standard vesicle-forming lipids which generally include neutral and negatively charged phospholipids and a sterol, such as cholesterol.
  • the selection of lipids is generally determined by considerations such as liposome size and stability in the blood.
  • a variety of methods are available for preparing liposomes as reviewed, for example, by Coligan, J. E. et al, Current Protocols in Protein Science, 1999, John Wiley & Sons, Inc., New York, and see also U.S. Pat. Nos. 4,235,871, 4,501,728, 4,837,028, and 5,019,369.
  • the carrier may comprise, in total, from about 0.1% to about 99.99999% by weight of the pharmaceutical compositions presented herein.
  • the composition is for use in a method of the invention.
  • the agent is for use in a method of the invention.
  • the inhibitor is for use in a method of the invention.
  • the ILT3 antibody or antigen binding fragment thereof is for use in a method of the invention.
  • an ILT3 inhibitor is for use in combination with an CD20 inhibitor.
  • an CD20 inhibitor is for use in combination with an ILT3 inhibitor.
  • an ILT3 inhibitor is for use in combination with an EGFR inhibitor.
  • an EGFR inhibitor is for use in combination with an ILT3 inhibitor.
  • an ILT3 inhibitor is for use in combination with an HER2 inhibitor. In some embodiments, an HER2 inhibitor is for use in combination with an ILT3 inhibitor. In some embodiments, an ILT3 inhibitor is for use in combination with a CD47 inhibitor. In some embodiments, a CD47 inhibitor is for use in combination with an ILT3 inhibitor. In some embodiments, an ILT3 inhibitor is for use in combination with a PD-L1 inhibitor. In some embodiments, a PD-L1 inhibitor is for use in combination with an ILT3 inhibitor. In some embodiments, an ILT3 inhibitor is for use in combination with an CLDN18 inhibitor. In some embodiments, an CLDN18 inhibitor is for use in combination with an ILT3 inhibitor.
  • an ILT3 inhibitor is for use in combination with a PD-1 inhibitor.
  • a PD-1 inhibitor is for use in combination with an ILT3 inhibitor.
  • the ILT3 antibody or antigen binding fragment thereof is for use in combination with at least one of: a HER2 antibody or antigen binding fragment, a CD47 antibody or antigen binding fragment and a CLDN18 antibody or antigen binding fragment.
  • the HER2, CD47 and/or CLDN18 antibody is a therapeutic, blocking or inhibitory antibody.
  • an ILT3 inhibitor is for use in combination with an anticancer agent comprising an IgGl or IgG3 scaffold. In some embodiments, an ILT3 inhibitor is for use in combination with an IgGl or IgG3 antibody. In some embodiments, an ILT3 inhibitor is for use in combination with an IgGl or IgG3 therapeutic antibody. In some embodiments, a scaffold is a domain. In some embodiments, the use is treating cancer. In some embodiments, the use is enhancing immune cell activation against a cancer. In some embodiments, the use is increasing ADCC. In some embodiments, the use is increasing ADCP. In some embodiments, increasing is enhancing.
  • the antibody or antigen binding domain is for use in a method of the invention. In some embodiments, the composition is for use in enhancing immune function. In some embodiments, the antibody or antigen binding domain is for use in enhancing immune function. In some embodiments, enhancing immune function is alleviating immune suppression. In some embodiments, immune function is immune activation. In some embodiments, the composition is for use in treating cancer. In some embodiments, the antibody or antigen binding domain is for use in treating cancer.
  • the composition is formulated for administration to a subject.
  • the composition is formulated for systemic administration.
  • the composition is formulated for intratumoral administration.
  • the terms “administering,” “administration,” and like terms refer to any method which, in sound medical practice, delivers a composition containing an active agent to a subject in such a manner as to provide a therapeutic effect.
  • One aspect of the present subject matter provides for intravenous administration of a therapeutically effective amount of a composition of the present subject matter to a patient in need thereof.
  • Other suitable routes of administration can include parenteral, subcutaneous, oral, intramuscular, or intraperitoneal.
  • the dosage administered will be dependent upon the age, health, and weight of the recipient, kind of concurrent treatment, if any, frequency of treatment, and the nature of the effect desired.
  • immune activation is immune function.
  • immune function is immune activation.
  • increasing immune function comprises increasing immune surveillance.
  • increasing immune function comprises increasing an immune response.
  • immune activation comprises ADCC.
  • immune activation comprises ADCP.
  • increasing immune function comprises decreasing immunosuppression.
  • immune function comprises immune mediated cell killing.
  • the killing is killing of a disease cell.
  • the immune response is against a disease cell.
  • the cell is a target cell.
  • the disease is a disease treatable by killing a target cell. In some embodiments, the disease is a disease treatable by inducing an immune response against a target cell. In some embodiments, the target cell is a diseased cell. In some embodiments, the disease is characterized by the presence of immune cells expressing ILT3.
  • the disease is cancer.
  • target cell is a cancer cell.
  • a cancer cell is a tumoroid.
  • the cancer is a solid cancer.
  • the cancer is a tumor.
  • the target cell is a tumor cell.
  • the cancer is characterized by the presence of immune cells expressing ILT3.
  • the cancer is characterized by the presence of tumor infiltrating immune cell expressing ILTs.
  • the cancer is characterized by the expression of APOE, FN 1 or both.
  • the cancer is characterized by a TME comprising APOE, FN1 or both.
  • the cancer is characterized by the expression of FN1. In some embodiments, the cancer is the cancer extracellular matrix. In some embodiments, the cancer is the cancer TME. In some embodiments, expression is overexpression. In some embodiments, overexpression is as compared to non-disease tissue. In some embodiments, non-disease tissue is non-cancerous tissue. In some embodiments, the tissue is of the same type as the disease tissue. In some embodiments, the tissue is of the same type as the tumor. In some embodiments, of the same tissue type is derived from the same tissue type.
  • cancer or "pre-malignancy” are diseases associated with cell proliferation.
  • Non-limiting types of cancer include carcinoma, sarcoma, lymphoma, leukemia, blastoma and germ cells tumors.
  • the cancer is solid cancer. In some embodiments, the cancer is a tumor. In some embodiments, the cancer is selected from hepato-biliary cancer, cervical cancer, urogenital cancer (e.g., urothelial cancer), testicular cancer, prostate cancer, thyroid cancer, ovarian cancer, nervous system cancer, ocular cancer, lung cancer, soft tissue cancer, bone cancer, pancreatic cancer, bladder cancer, skin cancer, intestinal cancer, hepatic cancer, rectal cancer, colorectal cancer, esophageal cancer, gastric cancer, gastroesophageal cancer, breast cancer (e.g., triple negative breast cancer), renal cancer (e.g., renal carcinoma), skin cancer, head and neck cancer, leukemia and lymphoma.
  • urogenital cancer e.g., urothelial cancer
  • testicular cancer prostate cancer
  • thyroid cancer ovarian cancer
  • nervous system cancer ocular cancer
  • lung cancer soft tissue cancer
  • bone cancer pancreatic cancer
  • bladder cancer skin cancer, intestinal cancer,
  • the cancer is selected from breast cancer, kidney cancer, head and neck cancer, lung cancer, sarcoma, gastric cancer and ovarian cancer.
  • the cancer is gastric cancer.
  • the cancer is colon cancer.
  • the cancer is lung cancer.
  • the lung cancer is non-small cell lung cancer (NSCLC).
  • the cancer is breast cancer. In some embodiments, the cancer is cholangiocarcinoma. In some embodiments, the cancer is colorectal cancer. In some embodiments, the colorectal cancer is K-Ras wild-type colorectal cancer. In some embodiments, the colorectal cancer is N-Ras wild-type colorectal cancer. In some embodiments, the colorectal cancer is BRAF mutant colorectal cancer. In some embodiments, the cancer is esophageal cancer. In some embodiments, the esophageal cancer is an adenocarcinoma or squamous cell carcinoma of the esophagus. In some embodiments, the cancer is gastric cancer.
  • the cancer is gastric or gastroesophageal junction adenocarcinoma.
  • the cancer is head and neck cancer.
  • the head and neck cancer is squamous cell carcinoma of the head and neck.
  • the cancer is lung cancer.
  • the lung cancer is non-small cell lung cancer.
  • the cancer is skin cancer.
  • the skin cancer is melanoma.
  • the cancer is ovarian cancer.
  • the cancer is pancreatic cancer.
  • the pancreatic cancer is pancreatic adenocarcinoma.
  • the cancer is renal cancer.
  • the renal cancer is renal cell carcinoma.
  • the cancer is sarcoma.
  • the sarcoma is soft tissue sarcoma.
  • the cancer is CD20 positive cancer. In some embodiments, the cancer is EGFR positive cancer. In some embodiments, the cancer is HER2 positive cancer. In some embodiments, the cancer is CD47 positive cancer. In some embodiments, the cancer is PD-L1 positive cancer. In some embodiments, the cancer is CLDN18 positive cancer. In some embodiments, positive is comprising expression above a predetermined threshold. In some embodiments, the threshold is the expression level of healthy cells. In some embodiments, the healthy cells are cells adjacent to the cancer. In some embodiments, the healthy cells are cells of the same cell type as the cancer. In some embodiments, the healthy cells are cells from the same tissue as the cancer.
  • a length of about 1000 nanometers (nm) refers to a length of 1000 nm+- 100 nm.
  • Example 1 The 5E5 anti-ILT3 antibody
  • a competition assay was performed to examine if the 5E5 antibody binds the same epitope as the commercial antibody ZM4.1. ZM4.1 and 5E5 did not compete with either indicating that 5E5 binds a unique epitope.
  • 5E5 was also examined functionally. CD8 T cell activation after MDSC suppression was examined. 5E5 increased IFNg secretion from the T cells and produced a statistically significant increase as compared to the IgG control (Fig. 1A). DC activation in the presence of Rituximab and fibronectin was examined as well and 5E5 again increased activation (Fig. IB). Similar results were also observed for restoring THP1 activation (Fig. 1C). In order to determine if other ILT3 ligands beyond fibronectin could inhibit activation, THP-1 cells were incubated with Rituximab in the presence of fibronectin or APOE. APOE is another known ligand of ILT3, but its addition to the media did not inhibit Rituximab mediated activation (Fig. ID). This indicates that it is specifically ILT3 -fibronectin blocking that restores activation.
  • h5E5 function monocytes were stimulated with M-CSF for 5 days in the presence or absence of gastric tumor cells (GA-04) or primary colon cancer cells (CO- 10) and the h5E5 antibody to produce MO macrophages.
  • the MO macrophages then underwent the process of differentiation into Ml macrophages still in the presence or absence of the cancerous cells and h5E5 antibody and were activated by LPS stimulation.
  • the Ml macrophages secreted high levels of TNFa, but this was abrogated when the cells were cocultured with autologous gastric cancer cells (GA-04) (Fig. 2A) or colon cells (CO- 10) (Fig. 2B).
  • iDCs immature DCs
  • DCtol were generated by culturing iDCs with IE-10 and LPS for 2 days.
  • the iDC stimulation was done in presence of the 5E5 antibody, humanized h52B8 antibody, isotype control or without any additions (medium alone).
  • TNFa secretion levels were determined by ELISA.
  • DCtols produced very low levels of TNFa and the same was true when the isotype control was used.
  • stimulation in the presence of the 5E5 antibody produced significantly increased levels of the cytokine secretion, which were superior to those produced when the h52B8 antibody was present (Fig. 3).
  • ILT3 blockade is known to be combinable with anti-PD-1 immune checkpoint inhibitors (ICIs).
  • ICIs anti-PD-1 immune checkpoint inhibitors
  • T cell activation in the presence of MDSCs was used to monitor the effect of 5E5, anti-PDl and combination h5E5+PD-l treatment. Indeed, it was found that the two blockades have a combined effect on T cell activation and produced a more robust effect than either antibody on their own (Fig. 4A). Indeed, the combined treatment was able to elevate IFNg secretion levels to above those observed in control activated CD8 T cells. A similar combined effect was observed when IFNg secretion from CD4 cells cocultured with DCtol was examined (Fig. 4B). Similar results were observed with a DCtols and CD4 T cell coincubation assay.
  • Example 2 Ex vivo and in vivo evaluation of therapeutic effect of the 5E5 antibody
  • a renal cell carcinoma (RCC) tumor sample was also used for this assay and again a statistically significant increase in proinflammatory cytokines and chemokines was observed (Fig. 5D).
  • Results in a colorectal cancer (CRC) tumoroid were similar though the effect of the 5E5 antibody was less pronounced (Fig. 5E).
  • Pembrolizumab did not have a significant effect on cytokine/chemokine secretion, however, when Pembrolizumab was combined with h5E5 the increase was highly significant and indicates a synergistic increase over each agent alone (Fig. 5E).
  • hILT3 transgenic mice were used to evaluate the therapeutic efficacy of the 5E5 antibody.
  • the mice were subcutaneously inoculated with MC-38 cancer cells (a syngeneic murine colon cancer cell line).
  • MC-38 engrafted mice were then treated with 5E5 or hIgG4 control (20mg/kg/dose) twice a week starting from day 0 (treatments on days 0,3,7,10, and 14). Tumor volume was recorded 3 times a week.
  • the anti- ILT3 antibody was highly effective, reducing the rate of tumor growth and the overall size of the tumor.
  • mice inoculated with MC-38 colorectal cancer cells were also treated with anti-PD- 1 antibody (2 mg/kg) (with a control IgG) or PD-1 (2 mg/kg) in combination with 5E5 (10 mg/kg). Mice were treated twice a week by intravenous injection and were monitored for tumor growth. Anti-PD-1 alone produced a modest not statistically significant effect, which was greatly enhanced by the combination with the 5E5 antibody (Fig. 6B).
  • hILT3 transgenic mice were inoculated orthotopically (mammary fat pad) with cells of the E0771 murine breast cancer cell line. Once tumors reached a volume of 50 mm A 3 mice were randomized into 4 treatment groups: 1) IgG control, 2) anti PD-1 (2mg/kg), 3) anti PD-1 (2mg/kg) + 5E5 (5mg/kg) and 4) anti PD-1 (2mg/kg) + h52B8 (5mg/kg). Mice were treated twice a week by i.v, starting from the randomization day, and monitored for tumor growth. As can be seen in Figure 7A, anti-PD-1 had a modest effect on its own that was greatly enhanced by 5E5.
  • the combination produced a tumor growth inhibition of 61% as compared to the control IgG group.
  • all animals in the control group had tumors that exceed a volume of 400 mm A 3.
  • 7 of 11 animal that received anti- PD-1 monotherapy had tumors of this volume on day 22, 5 of 11 from the combination therapy with h528B and PD- 1 reached this volume and only 1 of the 11 animals that received combination 5E5 and PD-1 reached this volume (Fig. 7B).
  • Another commercially available anti-ILT3 antibody, 10202 was also tested and was found to be comparable to h52B8 (data not shown).
  • mice were randomized into 5 different treatment groups: 1) IgG control, 2) anti PD-L1 (2 mg/kg) + IgG control, 3) anti PD-L1 (2 mg/kg) + 5E5 (10 mg/kg), 4) anti-PD-Ll (2 mg/kg) + h52B8 (10 mg/kg) and 5) anti-PD-Ll (2 mg/kg) + 10202 (10 mg/kg).
  • Anti-PD-Ll had a modest effect on its own.
  • combination with the 5E5 antibody greatly enhanced the anti-PD-Ll effect (Fig. 7C). Both antibodies tested produced effects similar to 5E5, indicated that the combined synergistic effect is universal to ILT3 blocking.
  • THP-1 monocyte cells were activated with immobilized Cetuximab (Erbitux).
  • Erbitux is an epidermal growth factor receptor (EGFR) inhibitory antibody. Activation was monitored by measuring proinflammatory cytokine IL-8 secretion. When fibronectin was added to the culture it inhibited IL- 8 secretion, essentially completely abolishing secretion (Fig. 8A). 5E5 was not only able to reverse the fibronectin mediated inhibition, but at its highest concentration was even able to produce an increase in IL- 8 secretion over what Erbitux alone produced (Fig. 8A).
  • Avelumab (Bavencio) an anti-PD-Ll antibody and Zolbetuximab (Vyloy) an anti-claudin-18 antibody.
  • Avelumab is a human IgGi anti-PD-Ll blocking mAb that may also trigger ADCC and ADCP against cancer cells as an additional antitumor activity.
  • Trastuzumab and Zolbetuximab also both can trigger ADCC and ADCP.
  • ILT3 blockade not only reversed the inhibitory effect of fibronectin, but even at fairly low concentrations was able to produce an increase in IL- 8 secretion over that produced by Herceptin alone (Fig. 8B), Avelumab alone (Fig. 8C) or Zolbetuximab alone (Fig. 8D).
  • M2 macrophages were generated by stimulating monocytes from a healthy donor with M-CSF for 5 days to generate M-0 macrophages. This was followed by incubation with IL- 10 for an additional 48hr with or without (medium control) the addition of 5E5 or hIgG4.
  • A549 non-small cell lung cancer cells which are positive for EGFR, CD47 and fibronectin were cultured overnight either alone or with the addition of Erbitux (0.2pg/ml) or Magrolimab (anti-CD47 Ab; 5pg/ml).
  • hlgGl was used as a negative control for these antibodies.
  • E:T effector cell to target cell
  • the molecule itself did not produce a significant increase in secretion of the molecules, but combination with 5E5 produced a highly significant increase (even as compared to 5E5 alone) which was more than additive and was in fact synergistic.
  • ILT3 blockade of macrophages in combination with EGFR or CD47 inhibition of cancer cells is highly synergistic and greatly increases the levels of proinflammatory cytokines and chemokines produced against the cancer.
  • the effect of Erbitux on IL- 8 was also confirmed with macrophages from a second healthy donor (Fig. 91).
  • Avelumab The tumor cell targeting antibody Avelumab was also tested. This antibody binds to PD-L1 on tumor cells and inhibits the proteins’ ability to bind PD-1. As A375 tumor cells do not naturally express high levels of PD-L1 the cells were transfected with PD-L1 and then cultured overnight with or without Avelumab. Although the effect of anti-PD-1 blockade had been tested, this antibody targets the immune cells themselves whereas anti-PD-Ll targets the cancer cells. As before, 5E5 incubation during macrophage maturation significantly increased IL- lb levels in the media (Fig. 9 J) as well as IL-6 levels (Fig. 9K). Avelumab alone produced a similar increase in IL- lb, but an even greater increase in IL-6. When Avelumab was combined with ILT3 blockade an even greater increase was observed. This increase, however, appeared to be additive.
  • NK cells were isolated from the blood of healthy donors and stored in liquid nitrogen.
  • M2 macrophages were generated by stimulating monocytes (isolated from autologous healthy donors) with M-CSF for 5 days to generate M-0 macrophages followed by incubation with IL- 10 for an additional 48 hours. As before, the IL- 10 incubation was performed with or without the 5E5 antibody. hIgG4 was again used as a negative control.
  • NK cells were thawed and incubated with the autologous M2c macrophages in complete RPMI medium supplemented with IL-2 (200 U/ml). NK cells and macrophages were incubated at a ratio of 1:1 for 24 hours.
  • A549 cancer cells were pretreated with Erbitux or IgGl or were left untreated. The A549 cells were then incubated with only NK cells or NK cells co-cultured with M2c suppressive cells (E:T ratio of 5:1). After 5 hours cytotoxicity was evaluated.
  • Avelumab a non-small cell lung cancer cells line that is positive for PD-L1
  • Avelumab in the absence of macrophages produced a modest, though significant, increase in cytotoxicity. This increase was abrogated by the M2c macrophages.
  • Addition of 5E5 not only returned Avelumab ’s effectiveness in inducing cytotoxicity but greatly increased it. Avelumab produced less than 10% cytotoxicity whereas the combination of Avelumab and 5E5 produced greater than 60% cytotoxicity. This result demonstrates the synergy of ILT3 blockade combined with blocking PD-L1.

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Abstract

L'invention concerne des procédés d'augmentation de la cytotoxicité cellulaire dépendante des anticorps (ADCC) ou de la phagocytose cellulaire dépendante des anticorps (ADCP) contre une cellule cancéreuse et des méthodes de traitement du cancer, comprenant le blocage de la liaison de la fibronectine au récepteur de type immunoglobulines de la sous-famille B, membre 4 (LILRB4/ILT3). L'invention concerne en outre des méthodes de traitement du cancer chez un sujet dont l'état le nécessite, comprenant l'administration d'un anticorps anti-ILT3 et d'au moins l'un d'un anticorps anti-CD20, d'un anticorps dirigé contre le récepteur du facteur de croissance épidermique (EGFR), d'un anticorps dirigé contre le récepteur tyrosine-protéine kinase ErbB-2 (HER2), d'un anticorps anti-CD47, d'un anticorps anti-ligand de mort programmée 1 (PD-L1) et d'un anticorps anti-claudine-18 (CLDN18).
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