Structure-activity study of oncocins modified with cationic polar moieties at the termini

The critical emergence of antimicrobial resistance is aggravated by a pipeline deficient in novel antibiotic classes against Gram-negative pathogens. Proline-rich antimicrobial peptides (PrAMPs) are attractive starting points for novel antibiotics, capable of targeting Gram-negative pathogens. Here, we explored a series of oncocin analogs, which were modified with bacteria-penetrating moieties to promote transporter-independent antibacterial activity. Elongation with arginine-rich motifs conferred 2- to 8-fold improved potency against the Gram-negative Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii. The highest degree of enhancement was seen for K. pneumoniae, in which the most active analogs had minimal inhibitory concentrations of 0.25 μg/mL (∼0.05 μM). Notably, antibacterial activity proved considerably less dependent on the inner-membrane peptide transporter SbmA, being the main transporter involved in internalization of PrAMPs. Permeabilization experiments showed that these cationic modifications did not alter the original intracellular mode of action of oncocin itself towards a membrane-disruptive mechanism for these analogs. Importantly, their effect on human cell viability remained low for the best analogs (i.e., below 5 % hemolysis at 400 μg/mL and IC₅₀ values ≥ 1280 μg/mL in HepG2 cells). Moreover, the frequencies of existing resistance of best analogs in E. coli and K. pneumoniae were low (i.e., 1-5 × 10⁻¹⁰). Finally, two analogs were tested in a mouse peritonitis model with E. coli, while one analog was tested in a model with K. pneumoniae. This showed efficacy comparable to that of colistin in the E. coli model, while a lower efficacy than that of colistin was seen in the K. pneumoniae model.