Exploring the structure-activity relationship of brevibacillin: Discovery of synthetic analogues with an extended spectrum of activity

Brevibacillin is an antimicrobial lipopeptide produced by Brevibacillus laterosporus with promising activity against several clinically relevant Gram-positive pathogens, including antibiotic-resistant strains such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE). Herein, we describe the optimization of the synthetic approach to produce brevibacillin and new analogues as well as a structure-activity relationship investigation to increase the activity, modify the spectrum of action and reduce the cytotoxicity. The alanine-scan of the lipopeptide revealed that cationic amino acids are essential for its activity against Gram-negative strains while the overall hydrophobicity plays an important role in the inhibition of Gram-positive bacteria and hemolysis. Substitution of the N-terminal acyl chain with fatty acids of different length yielded equipotent analogues but enhanced the hemolytic activity compared to brevibacillin. In another series of analogues, the introduction of a cationic amino acid at position 1 increased the inhibitory activity toward Gram-negative bacteria and significantly reduced the hemolytic activity, but at the cost of less potency against Gram-positive strains. Finally, the combination of both N-acyl and position 1 substitutions resulted in analogues exhibiting an extended spectrum of activity with minimal inhibitory concentration values in the low micromolar range against Gram-positive S. aureus, MRSA, Bacillus subtilis, Enterococcus faecium and Gram-negative Escherichia coli, Salmonella enterica, Klebsiella pneumoniae and Acinetobacter baumannii as well as enhanced killing kinetics compared to brevibacillin. With their enhanced synthetic accessibility, strong inhibitory activity, shifted or extended spectrum of action and improved selectivity index towards Gram-negative bacteria, these brevibacillin analogues are promising candidates for the development of new antimicrobials.