Potent inhibitors of human matriptase-1 based on the scaffold of sunflower trypsin inhibitor

Sunflower trypsin inhibitor-1 (SFTI-1), a bicyclic tetradecapeptide, has become a versatile tool as a scaffold for the development of the inhibitors of therapeutically relevant serine proteases, among them matriptase and kallikreins. Herein, we report the rational design of potent monocyclic and bicyclic inhibitors of human matriptase-1. We found that the presence of positive charge and lack of bulky residues at the peptide N-terminus is required for the maintenance of inhibitory activity. Replacement of the N-terminal glycine residue by lysine allowed for the chemical conjugation with a fluorophor via the ε-amino group without significant loss of inhibitory activity. Head-to-tail and side-chain-to-tail cyclization resulted in potent inhibitors with comparable activities against matriptase-1. The most potent synthetic bicyclic inhibitor found in this study (K_i = 2.6 nM at pH 7.6) is a truncated version of SFTI-1 (cyclo-KRCTKSIPPRCH) lacking a C-terminal proline and aspartate residue. It combines an internal disulfide bond with a peptide macrocycle that is formed through side-chain-to-tail cyclization of the ε-amino group of an N-terminal lysine and a C-terminal proline. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.

We report the rational design, synthesis, and biological activity of potent monocyclic and bicyclic peptidic inhibitors of human matriptase-1 based on an improved variant of the sunflower trypsin inhibitor-1. The activity of the resulted synthetic variants against cancer-related human matriptase-1 was examined in order to evaluate the influence of structural elements, e.g. cyclic backbone, chain length, and bulky substituents. The most potent inhibitors showed activity against matriptase-1 in single-digit nanomolar range (K_i = 2.1 nM).