Generation of VacA-targeting guide peptides to increase specific antimicrobial peptide toxicity against Helicobacter...

The H. pylori virulence factors VacA and CagA are the primary determinants of gastric cancer globally. In this study we increased the activity of antimicrobial peptides (AMPs) against H. pylori by using phage display against VacA to rapidly generate peptides targeting VacA, subsequently fusing these peptides to the AMP N-terminus to confer specificity.

After four rounds of phage display, 36 phage clones were ranked for whole cell H. pylori binding by ELISA. The displayed 12-mer peptides of the top nine candidate clones were fused to GFP as guide peptides and analyzed for binding to wild type H. pylori 60190 and a ∆vacA strain. The three guides with the best differential binding were fused to the AMP pexiganan using two different linker peptides. All guide/linker combinations led to increased toxicity against H. pylori and most also decreased off-target toxicity. Guide P5 linked to pexiganan was the top configuration, delivering 64- to > 256-fold differential toxicity against H. pylori compared to off-target bacteria and a therapeutic window exceeding 128-fold when tested against cultured gastric cells.

Guide peptide biopanning provides an effective, scalable method to increase specific activity of antimicrobial peptides based on attraction to a key virulence factor.