Developing Cyclic Peptide Inhibitors to the von Willebrand Factor-Collagen Interaction: Implications in Thrombosis Therapeutics

Cyclic peptides are favorable in the development of new therapeutics as they use functional groups recognized by biological targets and the physical constraint of a ring to prevent degradation. Targeting the interaction of von Willebrand Factor (vWF) to collagen is significant to inhibiting the initiation stage of thrombosis, which is linked to cardiovascular disease. We developed several increasingly potent cyclic peptide inhibitors using two approaches for their design. The first, ongoing method, is the synthesis of head-to-tail cyclized peptides with the inclusion of non-natural, aromatic amino acid substitutions naphthylalanine or biophenylalanine in one to two positions in rings of seven amino acids. By creating peptides that are rigid and small we have observed successful sub-micromolar inhibition of vWF binding to collagen using our in vitro fluorescently-linked immunosorbent assay. The second method was in collaboration, as we expressed lasso peptides with a threaded, macrocyclic, structure that bears an isopeptide bond between the N-terminus and a key glutamate. Using site-directed mutagenesis we substituted four residues in the loop region of a well-established scaffold, obtaining single micromolar inhibition in the same in vitro assay. All of our peptides were tested against a panel of proteases, displaying an average low level of degradation over time. These studies provide a novel direction in the development of peptides as anti-thrombosis agents.