Monoclonal Antibodies Recognizing Unique Native Structure of PRSS14/Epithin/Matriptase/ST14 can Interfere Cancer Cell Migration

초록

PRSS14/Epithin, also known as Matriptase and ST14, is a type II transmembrane serine protease and upregulated widely in epithelial type cancers. In previous studies, we showed that it is involved in angiogenesis, epithelial mesenchymal transition, transendothelial migration, and metastasis. It cleaves extracellular matrix proteins (collagen, fibronectin) and activates many well known tumor associated proteins (PAR-2, pro-HGF, pro-uPA, pro-MMP3). To generate antibody reacting to the unique structural motifs of serine protease domain in PRSS14/epithin protein, we designed peptide antigen based on the antigenicity, evolutionary conservation, and structural modeling. The chosen amino acid sequence of PRSS14/Epithin is not present any other proteins. Eleven independent monoclonal antibodies were generated using the designed peptide antigen and hybridoma technology. To capture native structure, we performed immunoprecipitation and flow cytometry with various cells expressing protein. Several mAbs show cross-reactivity to mouse PRSS14/Epithin, indicating that they are useful for studies in vitro with well characterized mouse and human cell lines. Immunocytochemistry showed mAbs bind to protein present in the cell contacts. Many mAbs inhibited cancer cell migration. Therefore, these mAbs will be useful for diagnosis and therapeutic application as well as for the studies of protein function in mouse models.