The greatest challenge in targeted cancer therapy is the lack of good therapeutic targets. Our current and future research is focused on investigating tumor-specific targets and their roles in tumorigenesis and metastases in addition to developing new therapeutic antibodies. We are studying three tumor antigens: (1) mesothelin, (2) CA125 (also known as MUC16) and (3) a new liver cancer antigen. As a promising new target for cancer immunotherapy, mesothelin is a cell surface biomarker that is highly expressed in mesothelioma, ovarian cancer and other malignant tumors. A mouse monoclonal antibody (mAb) variable fragment (Fv) specific for mesothelin, called SS1, was identified by Dr. Ira Pastans lab. It has been developed as an immunotoxin or a chimeric antibody and is currently in clinical trials for cancer therapy. Due to their low immunogenicity in patients, fully human monoclonal antibodies are the most desirable antibody format for clinical applications. We have screened phage display libraries and isolated several lead Fv molecules. In FY09, we (1) identified HN1 as a high affinity Fv specific for mesothelin, (2) developed a fully human IgG molecule based on the HN1 Fv, (3) showed that HN1 binds specifically to cell surface-associated mesothelin on mesothelioma and ovarian cancer cells, and (4) demonstrated its specific cytotoxic activity on cancer cells. We are currently evaluating HN1 as a candidate therapeutic agent to treat mesothelin-expressing tumors. Our work on HN1 was presented in several international meetings and yielded an international patent application in FY09. CA125 represents another important biomarker found on a majority of ovarian cancer and mesothelioma cells. Ovarian cancer and mesothelioma cells commonly spread within the peritoneal cavity via seeding to mesothelium-lined structures. The interaction between CA125 and mesothelin has been suggested to facilitate implantation and metastasis of tumors. We hypothesize that blocking the mesothelin-CA125 interaction may prevent or reverse metastasis and lead to an overall improved survival in cancer patients. A better understanding of how these two proteins interact may eventually aid in developing such therapy. To this end, my laboratory has vigorously examined the molecular interaction of mesothelin and CA125. In FY08, we (1) identified the binding site on mesothelin for CA125 and (2) revealed specific amino acids within the CA125-binding site of mesothelin that are involved in the interaction with CA125. In FY09, we developed novel anti-cancer therapeutics based on our work on CA125. In one approach, we have (1) developed a new fully human molecule specific for CA125-expressing cancer cells with high affinity and excellent specificity, and (2) found that this molecule can effectively block the binding of CA125 to mesothelin, thereby suggesting it could be used as an antagonist to inhibit tumor progression and spread in the peritoneal cavity. We are now studying its potential cytotoxic activity on cancer cells. In another approach, we have developed a novel hybrid molecule containing a pro-apoptotic peptide to induce cell death in CA125-expressing cancer cells. Our work targeting CA125 yielded one publication and an international patent application in FY09. In FY09, we became interested in a recently-identified liver cancer-specific antigen. Highly expressed in liver tumors but not in normal liver tissues, we were able to express and purify the liver cancer antigen as a recombinant Fc fusion protein. We are currently investigating its molecular interaction with a possible partner protein. This work may provide mechanistic insights on the origin of liver cancer and how liver cancer is associated with hepatitis viral infection. We plan to isolate high affinity monoclonal antibodies using hybridoma and phage display technology. In order to evaluate the efficacy of antibody therapeutics in a clinically relevant tumor model, my laboratory recently developed a novel malignant mesothelioma xenograft in mice. This model may be very useful for investigating antibodies and other therapeutics pertaining to the treatment of malignant mesothelioma. Previously, our work led to the development of mammalian cell display, a new method for antibody engineering. Based on this approach, functional single chain Fv antibodies (scFv) may then be expressed on human cells and high affinity scFv binders are rapidly identified and isolated from a combinatory Fv library. We are improving this method in order to make a large and permanent Fv library with the purpose of isolating high affinity antibodies for cancer therapy. In FY09, our work on antibody engineering yielded three publications and co-authored two publications.