FIBROBLAST ACTIVATION PROTEIN - PURIFICATION, EPITOPE MAPPING AND INDUCTION BY GROWTH-FACTORS

The human fibroblast activation (FAP) defined by monoclonal antibody ( MAb) F19 is a cell surface antigen expressed in reactive stromal fibro blasts of breast, colorectal, lung and other epithelial cancers. In co ntrast to its stroma-specific localization in epithelial neoplasms, FA P is expressed in the malignant mesenchymal cells of bone and soft tis sue sarcomas. FAP is transiently expressed in some fetal mesenchymal t issues but is absent or expressed at low levels in most adult tissues. FAP is induced in cultured fibroblasts and, in these cells, consists of a M(r) 95,000 subunit (FAP alpha) carrying the F19 epitope and a no n-covalently bound M(r) 105,000 subunit (FAP beta) lacking the F19 epi topes. Using MAb F19 and 5 newly derived MAbs, we identify 3 distinct epitopes on FAP alpha and tentatively assign one epitope to FAP beta. Analysis of detergent extracts of a FAP alpha(high)beta-sarcoma cell l ine by size exclusion-high performance liquid chromatography (HPLC) re vealed that FAP alpha does not elute as a M(r) 95,000 species but as p art of a high-molecular weight complex (M(r) > 400,000) that dissociat es into M(r) 95,000 subunits in SDS gels. Immunoaffinity purification of FAP alpha followed by tryptic digestion, reversed-phase HPLC and mi crosequencing identified 3 unique FAP alpha peptides, with 2 showing s equence similarity (23/38 identical amino acids) to segments of CD26, a T-cell activation antigen. CD26 is a membrane-bound enzyme (dipeptid yl aminopeptidase IV), but immunopurified FAP alpha has little if any dipeptidase activity with typical CD26 substrates. Finally, studies wi th FAP(low) leptomeningeal fibroblasts revealed that transforming grow th factor-beta, 12-O-tetradecanoyl phorbol-13-acetate and retinoids ca n upregulate FAP expression, whereas serum and several other factors h ad no or little effect on FAP levels. FAP and CD26 may belong to a fam ily of structurally related but functionally distinct activation prote ins that are expressed on different cell types and show unique modes o f regulation in normal and malignant cells. (C) 1994 Wiley-Liss, Inc.