(499e) Epitope-Targeted Peptide Immunostimulants to Combat Antibiotic-Resistant Bacteria

Antibiotic resistance in bacterial pathogens poses a rapidly emerging health threat. Today, many bacteria have acquired resistance to all available antibiotics, while the discovery of new antimicrobials has slowed. Without a reliable and perhaps general method to generate antibiotics against newly emerging resistant bacterial strains, antibiotic resistance will grow as a health threat. We envision a synthetically versatile molecular platform, called epitope-targeted peptide immunostimulants (EPIs), as a sustainable therapeutic countermeasure against drug-resistant pathogens. EPI molecules comprise of two chemical moieties: (1) a polypeptide ligand that binds a protein at a bacterial surface and (2) an immunogenic group that recruits endogenous antibodies. EPIs adsorb to the surfaces of the targeted bacteria, promote opsonization, and ultimately enhance phagocytic killing by innate immune cells. To test this, we used a powerful combinatorial chemistry technique, the protein-catalyzed capture agent (PCC) approach, to identify macrocyclic peptide ligands that bind epitopes at the surface of carbapenem-resistant Klebsiella pneumoniae, which are of high clinical significance. Guided by multi-omic and bioinformatics analyses to select target epitopes, the PCC approach was employed to develop EPI ligands against four 14-residue epitopes on the highly-abundant fimbiral subunit protein, MrkA, on K. pneumoniae. These screens yielded several EPI ligands, and the top-performing ligand exhibits high-affinity (EC₅₀~30 nM) to the full-length MrkA protein and selectively binds the surfaces of MrkA-expressing K. pneumoniae, versus other pathogenic bacterial species. EPIs that are conjugated with an immunogenic 2,4-dinitrophenyl (DNP) moiety enabled the recruitment of anti-DNP antibodies to K. pneumoniae surfaces and, further, that K. pneumoniae opsonized in this manner were more susceptible to phagocytosis by macrophages than non-treated cells. While K. pneumoniae was targeted in this study, the PCC approach enables EPIs to be developed against virtually any epitope and so EPIs could easily be generated for other resistant pathogens. Our work demonstrates EPIs as a promising all-synthetic solution to antibiotic resistance.