Proteases are directly involved in cancer pathogenesis. Expression of fibroblast activation protein (FAP) is upregulated in stromal fibroblasts in more than 90% of epithelial cancers and is associated with tumor progression. FAP expression is minimal or absent in most normal adult tissues, suggesting its promise as a target for the diagnosis or treatment of various cancers. Here, we report preparation of a polymer conjugate (an iBody) containing a FAP-specific inhibitor as the targeting ligand. The iBody inhibits both human and mouse FAP with low nanomolar inhibition constants but does not inhibit close FAP homologues dipeptidyl peptidase IV, dipeptidyl peptidase 9, and prolyl oligopeptidase. We demonstrate the applicability of this iBody for the isolation of FAP from cell lysates and blood serum as well as for its detection by ELISA, Western blot, flow cytometry, and confocal microscopy. Our results show the iBody is a useful tool for FAP targeting in vitro and potentially also for specific anticancer drug delivery.

Institute of Biochemistry and Experimental Oncology 1st Faculty of Medicine Charles University U Nemocnice 5 12853 Prague 2 Czech Republic

Institute of Macromolecular Chemistry The Czech Academy of Sciences Heyrovského nám 2 16206 Prague 6 Czech Republic

Institute of Organic Chemistry and Biochemistry of The Czech Academy of Sciences Flemingovo nám 2 16610 Prague 6 Czech Republic Department of Biochemistry Faculty of Science Charles University Hlavova 8 12843 Prague 2 Czech Republic

Institute of Organic Chemistry and Biochemistry of The Czech Academy of Sciences Flemingovo nám 2 16610 Prague 6 Czech Republic Department of Cell Biology Faculty of Science Charles University Viničná 7 12843 Prague 2 Czech Republic

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