Identification of a novel cancer therapeutic antibody against human epidermal growth factor receptor 2 (Her2) and antibody engineering for development of cancer therapeutics

by Chao Chen

Institution: University of Hong Kong
Degree: PhD
Year: 2013
Keywords: Cancer - Treatment; Monoclonal antibodies - Therapeutic use
Record ID: 1177591
Full text PDF: http://hdl.handle.net/10722/196461


Cancer is one of the leading causes of death worldwide. Monoclonal antibodies (mAbs) have been proved effective for cancer therapy. MAbs possess advantages over chemical drugs and small molecular drugs in cancer treatment, such as high specificity, low toxicity, effector function, long half-life in circulation system and less side effects. There are eight FDA-approved anti-cancer antibody drugs now, and many more are under development. Antibodies have two functional domains, the Fab region that is responsible for antigen recognition, and the Fc region that couples the antibody to immune effector pathways. Fab-mediated interference with cancer cell signalling may lead to growth inhibition and direct cell death, while Fc-mediated effector function through interactions with Fc-gamma receptors (FcrRs) expressed in immune cells or through complement cascades may lead to target cell cytotoxicity. Antibody engineering to increase the binding affinity and effector function may improve antibody in vivo efficacy. Anti-Her2 mAb herceptin (trastuzumab) is effective in treatment of Her2-overexpressing breast cancer patients. However, only 25–30% of patients with Her2-overexpressing tumors respond to single agent trastuzumab, and drug resistance develops even in responding patients. Accumulating evidence showed that cross-talk between Her2 and the insulin-like growth factor receptor type I (IGF-IR), including receptor heterodimerization and transactivation, and elevated IGF-IR signalling have been associated with trastuzumab resistance. Therefore, we hypothesized that dual specific antibodies co-targeting both IGF-IR and Her2 may prevent or delay the emergence of resistance to mono-specific antibodies. Mouse monoclonal antibody, M590 showed very good binding activity to IGF-IR. By engineering the CH3 domain of human Fc in pDR12 plasmid, we developed a “knobs-into-holes” hybrid IgG expression system, and successfully produced M590-Herceptin bi-specific IgG, which showed high binding avidity for both antigens and preserved antibody-dependent cell-mediated cytotoxicity (ADCC), a main route of immune protections conferred by therapeutic antibodies in vivo. M590-Herceptin dual specific antibody inhibited breast cancer and ovarian cancer cell proliferation in vitro, and inhibited cancer growth in a SKOV-3 Her2- and IGF-IR-overexpressing ovarian cancer xenograft mouse model. M590-Herceptin hybrid showed better anti-cancer activity compared with M590 and Herceptin alone, or in combination. Meantime, I also constructed a phage display antibody Fab library using the mRNA of rabbits immunized by membrane proteins of SKOV-3 cells, and isolated a novel anti-Her2 mAb, designated as 1C6. Results from in vitro assays showed that 1C6 had anti-cancer activity which was comparable to that of herceptin. M590-1C6 hybrid IgG was also constructed, and the results from in vitro assays and mouse study showed that M590-1C6 hybrid IgG also possess better inhibitory activity of Her2 positive tumours compared with m590 or 1C6 alone. In summary, this…