Dilipid ultrashort cationic lipopeptides as adjuvants for chloramphenicol and other conventional antibiotics against Gram-negative bacteria

The necessity to develop therapeutic agents and strategies to abate the spread of antibiotic-resistant pathogens is prominent. Antimicrobial peptides (AMPs) provide scaffolds and inspiration for antibiotic development. As an AMP of shorter scaffold, eight dilipid ultrashort cationic lipopeptides (dUSCLs) were prepared consisting of only four amino acids and varying dilipids. Lipids were acylated at the peptide N-terminus and the epsilon-amine side chain of the N-terminal l-lysine. Compounds that possess aliphatic dilipids of 11 carbons-long showed significant hemolysis and therefore limited therapeutic application. Several non-hemolytic dUSCLs were identified to enhance the activity of chloramphenicol and other conventional antibiotics against Gram-negative bacteria. Compounds 2 and 6 have a short peptide sequence of KKKK and KKGK, respectively, and are both acylated with an aliphatic dilipid of nine carbons-long potentiated chloramphenicol against MDR clinical isolates of Pseudomonas aeruginosa, Acinetobacter baumannii and Enterobacteriaceae. Both dUSCLs showed comparable adjuvant potency in combination with chloramphenicol. However, dUSCL 2 synergized with a wider span of antibiotic classes against P. aeruginosa relative to dUSCL 6 that included rifampicin, trimethoprim, minocycline, fosfomycin, piperacillin, ciprofloxacin, levofloxacin, moxifloxacin, linezolid and vancomycin. Our data revealed that dUSCLs can indirectly disrupt active efflux of chloramphenicol in P. aeruginosa. This along with their membrane-permeabilizing properties may explain the dUSCLs synergistic combination with conventional antibiotics against Gram-negative bacteria.