The Coalescence of a "Happy Accident" and an In Vitro Structure-Activity Panel: Increasing the Potency of a Cyclic...

Cyclic peptides for novel drug discovery utilize the functional groups recognized by most biological targets but impose physical constraints as a preventative to degradation. Substituting non-natural, aromatic amino acids in cyclic peptide motifs can improve the ability of such peptides to bind to their protein targets and can further prevent instability. Here, we appended our previously described synthetic head-to-tail cyclized peptide with naphthylalanine (Nap) or biphenylalanine (BiPhe) substitutions in one or two positions. Our first-generation inhibitor targeted the protein-protein interaction between von Willebrand factor and collagen, implicated in thrombosis linked to cardiovascular diseases. Through an unplanned modification, we omitted a glycine that usually aids the flexibility of ring closing. Using our previously developed fluorescently linked immunosorbent assay, we saw modest improvements for most of the Nap- or BiPhe-substituted peptides, with the largest changes occurring with the no-glycine compound, yielding an inhibitory concentration of 6.47 μM. An array of stability was obtained for all of the peptides using protease degradation assays; however, every BiPhe- or Nap-substituted peptide showed limited degradation over time. Through a "happy accident," we discovered a new cyclic peptide scaffold, decreasing flexibility in our head-to-tail ring structure, paving the way for novel, potent, cyclic peptides with therapeutic value against thrombosis.