Human monoclonal antibodies specific for glypican-3 and use thereof
Inventors
Ho, Mitchell • Kim, Heungnam • Gao, Wei
Assignees
US Department of Health and Human Services
Publication Number
US-9394364-B2
Publication Date
2016-07-19
Expiration Date
2032-04-19
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Abstract
Described herein is the identification of human monoclonal antibodies that bind GPC3 or heparan sulfate (HS) chains on GPC3 with high affinity. The antibodies described herein are capable of inhibiting HCC cell growth and migration. Provided are human monoclonal antibodies specific for GPC3 or HS chains on GPC3, including immunoglobulin molecules, such as IgG antibodies, as well as antibody fragments, such as single-domain VH antibodies or single chain variable fragments (scFv). Further provided are compositions including the antibodies that bind GPC3 or HS chains on GPC3, nucleic acid molecules encoding these antibodies, expression vectors comprising the nucleic acids, and isolated host cells that express the nucleic acids. Methods of treating cancer and/or inhibiting tumor growth or metastasis are also provided. Further provided are methods of detecting cancer in a subject and confirming a diagnosis of cancer in a subject.
Core Innovation
The invention describes the identification and characterization of isolated human monoclonal antibodies that specifically bind glypican-3 (GPC3) or the heparan sulfate (HS) chains on GPC3 with high affinity. These antibodies include immunoglobulin molecules (such as IgG antibodies) and antibody fragments like single-domain VH antibodies and single-chain variable fragments (scFv). The antibodies bind specifically to GPC3 or its HS chains and have the capability to inhibit hepatocellular carcinoma (HCC) cell growth and migration. The disclosure also covers compositions containing these antibodies, nucleic acid molecules encoding them, expression vectors, isolated host cells expressing these antibodies, immunoconjugates comprising the antibodies linked to effector molecules such as toxins, and methods for treating cancer as well as detecting or confirming cancer diagnoses in subjects.
The disclosed antibodies provide means to target GPC3, which is highly expressed in hepatocellular carcinoma and some other human cancers but not in normal tissues. The problem addressed is the lack of good tumor-specific targets for effective immunotherapy of liver cancer, particularly because liver cancer often shows resistance to chemotherapy and is usually detected late. Glypican-3 is a cell-surface heparan sulfate proteoglycan with roles in cell proliferation and growth signaling pathways like Wnt and Hedgehog. The antibodies target either the core protein of GPC3 or the HS chains attached thereto, offering new mechanisms to inhibit tumor growth and metastasis by interfering with these signaling processes and tumor cell functions.
The invention includes specific human monoclonal antibodies such as the single-domain VH antibody named HN3 that binds a conformational epitope on GPC3 core protein, and the HS20 antibody that binds the HS chains on GPC3 and inhibits HCC cell migration by disturbing GPC3 interactions with Wnt3a. The antibodies bind with subnanomolar affinity and can induce cell cycle arrest, apoptosis, and inhibit tumor progression in vitro and in vivo. The immunoconjugates comprising these antibodies linked to toxins like Pseudomonas exotoxin (PE) or its derivatives demonstrate potent tumor cell killing and tumor growth inhibition in xenograft models. The disclosed antibodies and their derivatives solve the problem of lacking effective, tumor-specific immunotherapeutic agents for cancers expressing GPC3, such as hepatocellular carcinoma.
Claims Coverage
The patent claims cover isolated human monoclonal antibodies specific for heparan sulfate (HS) on glypican-3 (GPC3), compositions containing these antibodies, immunoconjugates with effector molecules, methods of treatment of cancers expressing GPC3, diagnostic methods to detect GPC3, nucleic acid molecules, vectors, host cells encoding the antibodies, and bispecific antibodies comprising these monoclonal antibodies.
Human monoclonal antibody binding heparan sulfate on GPC3
An isolated human monoclonal antibody that binds HS on GPC3, wherein the heavy chain comprises amino acid residues 26-33, 51-58 and 97-105 of SEQ ID NO: 14, and the light chain comprises amino acid residues 27-32, 50-52 and 89-97 of SEQ ID NO: 16 or SEQ ID NO: 31.
Antibody fragment and isotype variations
The antibody can be a Fab fragment, Fab' fragment, F(ab)'2 fragment, single chain variable fragment (scFv), disulfide stabilized variable fragment (dsFv), or an IgG molecule.
Labeled antibodies
The antibody can be labeled with fluorescence, enzymatic, or radioactive labels for detection purposes.
Therapeutic compositions
Compositions comprising therapeutically effective amounts of the antibody in pharmaceutically acceptable carriers.
Immunoconjugates with effector molecules
Isolated immunoconjugates comprising the antibody linked to effector molecules, such as toxins including Pseudomonas exotoxin (PE) or its variants like PE38.
Methods of treating cancer using compositions
Methods of treating subjects with cancers expressing GPC3 by administering therapeutically effective amounts of the antibody-containing compositions to inhibit tumor growth or metastasis.
Diagnostic methods for GPC3 detection
Methods of detecting GPC3 in tissue samples by contacting with the antibody and detecting increased binding relative to control samples to diagnose presence of GPC3-positive cancers.
Genetic constructs and host cells
Isolated nucleic acid molecules encoding the antibodies, expression vectors comprising these nucleic acids operably linked to promoters, and isolated host cells transformed with said vectors.
Bispecific antibodies
Bispecific antibodies comprising the human monoclonal antibody specific for HS on GPC3, potentially combined with antibodies targeting T cell receptor components such as CD3.
The claims comprehensively cover isolated human monoclonal antibodies specific for the HS chains on GPC3, including their variants, labeled forms, immunoconjugates, compositions, methods for treatment and detection of GPC3-positive cancers, nucleic acid sequences, vectors, host cells, and bispecific antibodies, highlighting the therapeutic and diagnostic applications of these antibodies.
Stated Advantages
HN3 is a fully human single-domain antibody with high affinity, expected to have low immunogenicity and better tumor penetration due to its small size.
The antibodies specifically inhibit HCC cell proliferation (HN3) or migration (HS20) and tumor growth in vivo.
HS20 inhibits the interaction of GPC3 and Wnt3a, disturbing Wnt signaling important for tumor cell migration and metastasis.
Immunoconjugates comprising the antibodies and toxins show potent cytotoxic activity against GPC3-expressing tumor cells with minimal off-target toxicity.
Documented Applications
Treatment of cancers expressing GPC3, including hepatocellular carcinoma (HCC), melanoma, lung cancer, and ovarian cancer.
Inhibition of tumor growth and metastasis in subjects with GPC3-positive cancers using antibodies or immunoconjugates thereof.
Diagnostic applications including detection and confirmation of GPC3-positive cancer in subject tissue or biological samples via antibody binding assays such as immunohistochemistry, ELISA, flow cytometry, or immunoassays.
Generation of engineered cytotoxic T lymphocytes (CTLs) expressing chimeric antigen receptors (CARs) that include antigen-binding fragments of the GPC3-specific monoclonal antibodies for adoptive immunotherapy.
Use in bispecific antibodies targeting both GPC3 and T cell receptors (e.g., CD3) for cancer immunotherapy.
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