EphA5-binding antibodies and uses thereof and selection of phage-displayed accessible recombinant targeted antibodies
Inventors
Pasqualini, Renata • Arap, Wadih • Staquicini, Fernanda Iamassaki • Ferrara, Fortunato • D'Angelo, Sara • Bradbury, Andrew R. M.
Assignees
US Department of Energy • Rutgers State University of New Jersey
Publication Number
US-11254751-B2
Publication Date
2022-02-22
Expiration Date
2037-09-21
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Abstract
Isolated or recombinant Eph A5 or GRP78 targeting antibodies are provided. In some cases, antibodies of the embodiments can be used for the detection, diagnosis and/or therapeutic treatment of human diseases, such as cancer. A method of rapidly identifying antibodies or antibody fragments for the treatment of cancer using a combination of in vitro and in vivo methodologies is also provided.
Core Innovation
The invention provides isolated or recombinant monoclonal antibodies that specifically bind to ephrin type-A receptor 5 (EphA5) or the 78 kDa glucose-regulated protein (GRP78). These antibodies potently block or reduce EphA5 or GRP78 signaling and inhibit cancer cell proliferation. Antibody fragments comprising complementarity determining regions (CDRs) from specific clones (E31, F31, TW3, T22 for EphA5; B4, D1, F6 for GRP78) and recombinant polypeptides comprising VH and VL domains related to these antibodies are also provided. The antibodies may be human, humanized, or de-immunized, and can be in various isotypes and fragment forms, potentially conjugated to imaging agents, chemotherapeutic agents, toxins, or radionuclides. The invention also discloses a method termed Selection of Phage-Display Accessible Recombinant Targeted Antibodies (SPARTA) that combines in vitro screening of a naïve human antibody phage library against immobilized tumor-associated recombinant antigens with in vivo selection based on tumor-homing of antibody displaying phage particles in tumor-bearing animals.
The problem addressed arises from the challenge in generating effective targeted cancer therapies due to the time-consuming and often unsuccessful processes of developing monoclonal antibodies against therapeutically relevant targets. Lung cancer, for instance, remains a leading cause of cancer death, and novel targets such as EphA5 and GRP78 have recently been implicated in cancer pathogenesis and chemoresistance but lacked effective therapeutic antibodies. Traditional antibody generation methods are limited in speed and success, and identifying antibodies that bind to targets accessible in the tumor microenvironment is difficult due to complex tumor biology and immune system interactions. SPARTA overcomes these limitations by integrating in vitro selection with functional in vivo tumor-homing screening, allowing rapid identification of human antibodies that not only bind the target proteins but also selectively home to tumors and exhibit therapeutic potential as antibody-drug conjugates.
Claims Coverage
The patent contains a set of independent claims focusing on isolated monoclonal antibodies specifically binding ephrin type-A receptor 5 (EphA5) with defined complementarity determining regions (CDRs).
Antibody specificity and composition
An isolated monoclonal antibody specifically binding EphA5 comprising: (a) a first VH CDR at least 80% identical to VH CDR1 of E31, F31, TW3, or T22 antibodies; (b) a second VH CDR at least 80% identical to VH CDR2 of said antibodies; (c) a third VH CDR at least 80% identical to VH CDR3 of said antibodies; (d) a first VL CDR at least 80% identical to VL CDR1 of said antibodies; (e) a second VL CDR at least 80% identical to VL CDR2 of said antibodies; and (f) a third VL CDR at least 80% identical to VL CDR3 of said antibodies.
Antibody formats and origins
The antibody can be recombinant, of isotypes IgG, IgM, IgA, or antigen binding fragments such as Fab′, F(ab′)2, F(ab′)3, monovalent or bivalent scFv. It may be human, humanized, or de-immunized.
Antibody conjugates
The antibody may be conjugated to an imaging agent, chemotherapeutic agent, toxin (including auristatin or monomethyl auristatin E MMAE), or radionuclide.
Antibody amino acid sequence specifics
The antibody comprises CDR sequences as set forth in SEQ ID NOs corresponding to E31, F31, TW3, or T22 clones and may have VH and VL domains with at least 80% sequence identity to the VH and VL domains of these clones.
Treatment methods
A method of treating or ameliorating lung cancer by administering an effective amount of the antibody specific for EphA5 described above.
Diagnostic methods
A method for detecting lung cancer by testing for elevated EphA5 levels in a sample using the isolated monoclonal antibody described above.
The claims cover isolated monoclonal antibodies specifically binding EphA5 with defined CDRs and variable regions related to four named antibodies (E31, F31, TW3, T22), their recombinant forms, formats, conjugates, compositions, and methods of using these antibodies for treating or detecting lung cancer.
Stated Advantages
The method SPARTA provides rapid and cost-effective identification of human recombinant monoclonal antibodies with high specificity and tumor-targeting capabilities.
Selected antibodies demonstrate enhanced tumor homing in vivo and functional binding to target antigens in their native conformations.
Antibodies and antibody-drug conjugates (ADCs) exhibit potent cytotoxicity against cancer cell lines expressing targets EphA5 or GRP78, with potential therapeutic uses.
Combination of phage- and yeast-display technologies enables selection of diverse and functional antibody populations.
Use of multivalent display phage with helper plasmids reduces non-specific binding and increases specific tumor targeting in vivo.
Documented Applications
Detection and diagnosis of human cancers expressing EphA5 or GRP78 by using antibodies specific to these targets.
Therapeutic treatment of cancers including lung cancer, breast cancer, and related epithelial cancers by administering the isolated antibodies or antibody conjugates.
Development and manufacturing of antibody-drug conjugates for targeted cancer therapies using antibodies specific for EphA5 or GRP78.
Use of the SPARTA method for selecting tumor-specific human monoclonal antibodies for cancer therapy and imaging.
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