title: Design of a cyclotide antagonist of neuropilin-1 and -2 that potently inhibits endothelial cell migration.
abstract: Neuropilin-1 and -2 are critical regulators of angiogenesis, lymphangiogenesis, and cell survival as receptors for multiple growth factors. Disulfide-rich peptides that antagonize the growth factor receptors neuropilin-1 and neuropilin-2 were developed using bacterial display libraries. Peptide ligands specific for the VEGFA binding site on neuropilin-1 were identified by screening a library of disulfide-rich peptides derived from the thermostable, protease-resistant cyclotide kalata B1. First generation ligands were subjected to one cycle of affinity maturation to yield acyclic peptides with affinities of 40-60 nM and slow dissociation rate constants (∼1 × 10(-3) s(-1)). Peptides exhibited equivalent affinities for human and mouse neuropilin-1 and cross-reacted with human neuropilin-2 with lower affinity. A C-to-N cyclized variant (cyclotide) of one neuropilin ligand retained high affinity, exhibited increased protease resistance, and conferred improved potency for inhibiting endothelial cell migration in vitro (EC50 ≈ 100 nM). These results demonstrate that potent, target-specific cyclotides can be created by evolutionary design and that backbone cyclization can confer improved pharmacological properties.
authors.affiliation: Department of Chemical Engineering, University of California , Santa Barbara, California 93106, United States.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Getz
authors.forename: Olivier
authors.initials: O
authors.lastname: Cheneval
authors.forename: David J
authors.initials: DJ
authors.lastname: Craik
authors.forename: Patrick S
authors.initials: PS
authors.lastname: Daugherty
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002465:Cell Movement / Q000187:drug effects*; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects*; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D039942:Neuropilin-1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D039943:Neuropilin-2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry* / Q000494:pharmacology*
chemical_list: D039943:Neuropilin-2; D010455:Peptides; D039942:Neuropilin-1