New Class of Drug Modalities: Directed Evolution of a De Novo Designed Helix–Loop–Helix Peptide to Bind VEGF for Tumor Growth Inhibition

Abstract

As a small affinity molecule to serve as an alternative to antibodies, we have developed a conformationally constrained peptide with a de novo designed helix−loop−helix (HLH) scaffold. To evaluate its potential for biomedical applications, we performed directed evolution of HLH peptides to obtain an inhibitor for vascular endothelial growth factor-A (VEGF). A phage-displayed library of HLH peptides was constructed and screened against VEGF, giving the peptide VS42 that inhibits the VEGF/VEGF receptor-2 interaction (IC50 = 210 nM), which was further improved by in vitro affinity maturation using a yeast-displayed library. An identified HLH peptide, VS42-LR3, exhibited improved inhibitory activity (IC50 = 37 nM), high thermal stability, and excellent resistance against chemical denaturation. In biological activity tests, the HLH peptide was found to block VEGF-induced proliferation of human umbilical vein endothelial cells and suppress tumor growth in a murine xenograft model of human colorectal cancer.