Peptidomimetic inhibitors of the VEGF-A165/NRP-1 complex obtained by modification of the C-terminal arginine.

Inhibitors of the interaction between Neuropilin-1 (NRP-1) and Vascular Endothelial Growth Factor-A₁₆₅ (VEGF-A₁₆₅) hold significant promise as therapeutic and diagnostic agents directed against cancers overexpressing NRP-1. In our efforts in this field, a few series of strong and fairly stable peptide-like inhibitors of the general formula Lys(Har)¹-Xaa²-Xaa³-Arg⁴ have been previously discovered. In the current work, we focused on Lys(Har)-Dap/Dab-Pro-Arg sequence. The aim was to examine whether replacing C-terminal Arg with its homologs and mimetics would yield more stable yet still potent inhibitors. Upon considering the results of modelling and other factors, ten novel analogues with Xaa⁴ = homoarginine (Har), 2-amino-4-guanidino-butyric acid (Agb), 2-amino-3-guanidino-propionic acid (Agp), citrulline (Cit), 4-aminomethyl-phenylalanine [Phe(4-CH₂-NH₂)] were designed, synthesized and evaluated. Two of the proposed modifications resulted in inhibitors with activity slightly lower [e.g. IC₅₀ = 14.3 μM for Lys(Har)-Dab-Pro-Har and IC₅₀ = 19.8 μM for Lys(Har)-Dab-Pro-Phe(4-CH₂-NH₂)] than the parent compounds [e.g. IC₅₀ = 4.7 μM for Lys(Har)-Dab-Pro-Arg]. What was a surprise to us, the proteolytic stability depended more on position two of the sequence than on position four. The Dab²-analogues exhibited half-life times beyond 60 h. Our results build up the knowledge on the structural requirements that effective VEGF-A₁₆₅/NRP-1 inhibitors should fulfil.