The long-term objective of this research program is to develop novel therapeutic strategies targeting the bidirectional interactions between tumor cells and the stromal cells that are found in tumors. The central hypothesis of this proposal is that inhibition of fibroblast activation protein (FAP) proteolytic activity will abrogate the invasive and metastatic capabilities of tumors, leading to attenuated tumor growth and diminished metastatic potential. This hypothesis is based on the highly selective expression of human FAP by tumor stromal fibroblasts in epithelial carcinomas, but not by epithelial carcinoma cells, normal fibroblasts, or other normal tissue. Pathways by which tumor fibroblasts participate in tumor growth can be targeted for disruption by monoclonal antibodies, which can be developed for clinical use. This proposal aims to generate monoclonal antibodies targeting murine FAP, and characterize their binding properties. This has been initiated by immunizing mice with murine FAP, and screening of hybridoma supernatants by ELISA for FAP binding. Antibodies that target FAP have been identified, and will be further characterized by surface plasmon resonance, live cell binding assays, and immunohistochemistry studies. FAP antibodies are also being assessed for inhibition of FAP proteolytic activity by dipeptidyl peptidase assays, enzyme immunocapture assays, and quantitative zymography. FAP inhibitory antibodies will be administered in an animal model to test the hypothesis that inhibition of FAP proteolytic activity inhibits tumor growth and metastases. If FAP inhibitory monoclonal antibodies cannot be produced by conventional immunization and fusion techniques, a human single-chain Fv (scFv) phage display library will be panned for FAP binding and enzymatic function inhibition. Inhibitory scFv fragments will be converted into IgG formats. Disruption of murine FAP proteolytic pathways by inhibitory antibodies resulting in inhibition of tumor growth and metastases will be considered sufficiently significant to warrant the clinical development of a novel therapeutic strategy targeting human cancers that induce FAP proteolytic activity. ? ?
