HER2-specific monoclonal antibodies and conjugates thereof
Inventors
Dimitrov, Dimiter S. • Zhu, Zhongyu • Qasba, Pradman K. • Ramakrishnan, Boopathy
Assignees
US Department of Health and Human Services
Publication Number
US-9738726-B2
Publication Date
2017-08-22
Expiration Date
2034-06-09
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Abstract
The identification of Her2-specific monoclonal antibody m860 is described. The m860 antibody was identified from a human naïve phage display Fab library by panning against the extracellular domain of human Her2. M860 binds to cell surface-associated Her2 with an affinity comparable to that of trastuzumab (Herceptin®), but binds to a different epitope. Using site-specific glycan engineering, m860 was conjugated to the small molecule drug auristatin F. The antibody-drug conjugate was stable, bound cell-surface expressed Her2 and exhibited potent cell killing of Her2-positive cancer cells, including trastuzumab-resistant breast cancer cells.
Core Innovation
The invention discloses a monoclonal antibody m860 that specifically binds to the extracellular domain of human Her2 at an epitope distinct from that of trastuzumab. The antibody was identified from a human naive phage display Fab library by panning against the extracellular domain of Her2. M860 binds cell surface-associated Her2 with an affinity comparable to trastuzumab.
Further, the invention provides an antibody-drug conjugate (ADC) where the m860 antibody is site-specifically conjugated, via glycan engineering of the N-linked oligosaccharide in the CH2 domain, to a small molecule anti-microtubule drug, auristatin F. This ADC is stable, retains binding to Her2 on cancer cells, and exhibits potent killing of Her2-positive cancer cells, including trastuzumab-resistant breast cancer cells.
The problem solved by this invention arises from the clinical challenge that many patients treated with trastuzumab do not respond or stop responding, and ultimately progress, highlighting the need for new therapies targeting Her2-positive cancers. ADCs that can provide tumor antigen-specific targeting of cytotoxic agents offer a promising approach to improve efficacy while reducing off-target toxicity. The site-specific drug conjugation via glycan engineering aims to enhance stability, reduce immunogenicity, and improve the therapeutic index of ADCs targeting Her2.
Claims Coverage
The patent contains multiple independent claims covering antibodies binding Her2, conjugates thereof, antibody-drug conjugates with specific drug conjugation sites, nucleic acids encoding the antibodies, and methods of detection and treatment.
Isolated monoclonal antibody binding specific Her2 epitopes
An isolated monoclonal antibody that binds Her2, wherein the VH domain comprises amino acid residues 26-33, 50-58 and 97-110 of SEQ ID NO: 2 and the VL domain comprises amino acid residues 23-34, 52-54 and 91-100 of SEQ ID NO: 4.
Monoclonal antibody or fragment with defined VH and VL sequences
Antibodies or fragments comprising VH and VL domain sequences substantially identical or comprising the exact sequences of SEQ ID NO: 2 and SEQ ID NO: 4 that specifically bind Her2.
Conjugates comprising the monoclonal antibody and effector molecules
Isolated conjugates made of the monoclonal antibody and an effector molecule such as a small molecule drug (e.g., auristatin F), toxin (e.g., Pseudomonas exotoxin or variant), or detectable label (fluorescent, enzymatic, radioactive).
Antibody-drug conjugate with site-specific drug conjugation via N-glycan
An ADC comprising a drug conjugated to the Her2-binding monoclonal antibody, wherein the drug is conjugated to an N-glycan of the CH2 domain of the antibody, with antibody to drug ratios of 1:4 or 1:2. The drug can be an anti-microtubule agent such as auristatin F with a non-cleavable linker at the C-terminus.
In vitro and in vivo methods using the antibody or ADC
Methods of killing Her2-positive cells using the ADC and methods for detecting Her2-expressing cells or cancers by contacting a biological sample with the monoclonal antibody, with or without labeling or secondary antibodies.
Nucleic acids and expression vectors for the monoclonal antibody
Isolated nucleic acid molecules encoding the monoclonal antibody with sequences comprising SEQ ID NO: 1 and SEQ ID NO: 3, operably linked to promoters, expression vectors, and host cells transformed therewith.
The independent claims cover the isolated monoclonal antibody that binds specific Her2 epitopes, its conjugates including ADCs with drugs conjugated site-specifically via the N-glycan, nucleic acids encoding these antibodies, and methods for detection and treatment of Her2-positive cancers employing these compositions.
Stated Advantages
The site-specific glycan engineering conjugation minimizes undesirable modification of the peptide backbone, improving efficacy and safety with lower immunogenicity.
This conjugation method increases stability and half-life of the ADC in vivo.
The ADC binds to a different epitope than trastuzumab allowing potential combination therapy.
The ADC exhibits potent and specific killing of Her2-positive cancer cells, including trastuzumab-resistant cells.
Documented Applications
Therapeutic treatment of Her2-positive cancers such as breast cancer, gastric cancer, esophageal cancer, ovarian cancer, endometrial cancer, stomach cancer, uterine cancer, pancreatic cancer, prostate cancer, bladder cancer, colon cancer, salivary gland carcinoma, renal adenocarcinoma, mammary gland carcinoma, non-small cell lung carcinoma, and head and neck carcinoma using the antibody or ADC compositions.
In vitro killing of Her2-positive cells using the antibody-drug conjugate.
Detection and confirmation of Her2-expressing cells or cancers in biological samples, including tissue or blood samples, by immunoassays using the monoclonal antibody, optionally labeled or detected via a secondary antibody.
Production of expression vectors and transformed host cells for expression of the monoclonal antibodies.
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