Functional Analyses of Native Structure Specific Monoclonal Antibody against Autoactivation Loop of Membrane Protease PRSS14

초록

PRSS14 is a type II transmembrane serine protease and overexpressed in various epithelial cancer types. Previously, we showed that it is involved in angiogenesis, TGFβ induced EMT, transendothelial migration, and metastasis of epithelial cancers. Bioinformatics analysis of The Cancer Genome Atlas (TCGA) revealed its values as a prognosis marker for basal type breast cancer and as better prognosis marker than HER2. Autoactivation cleavage at aa615 is required for enzymatic activity that processes the protein substrates. In order to control the protease activation of PRSS14 with monoclonal antibodies (mAbs), we designed antigenic peptide of human PRSS14 sequences containing the autoactivation site. Peptide sequences were optimized to maintain the native structural conformation by forming loop structure and used for the production of antibody. Using hybridoma technology, we obtained the loop structure specific monoclonal antibody, 3F3 that showed strong specificity to native form of antigen tested by immunoprecipitation and flow cytometry. It shows cross reactivity to ortholog mouse protein. This mAb detects inactive form of PRSS14 and increases shedding of protein from the membrane, unexpectedly, activation of protein induced by low pH. 3F3 antibody can enhance motility and invasiveness of breast and prostate cancer cell in wound healing and transwell invasion assay. Mode of action is currently under investigation.