Novel approaches for the treatment of multidrug-resistant Gram-negative 
bacterial infections are urgently required. One approach is to potentiate the 
efficacy of existing antibiotics whose spectrum of activity is limited by the 
permeability barrier presented by the Gram-negative outer membrane. Cationic 
peptides derived from polymyxin B have been used to permeabilize the outer 
membrane, granting antibiotics that would otherwise be excluded access to their 
targets. We assessed the in vitro efficacies of combinations of SPR741 with 
conventional antibiotics against Escherichia coli, Klebsiella pneumoniae, and 
Acinetobacter baumannii Of 35 antibiotics tested, the MICs of 8 of them were 
reduced 32- to 8,000-fold against E. coli and K. pneumoniae in the presence of 
SPR741. The eight antibiotics, azithromycin, clarithromycin, erythromycin, 
fusidic acid, mupirocin, retapamulin, rifampin, and telithromycin, had diverse 
targets and mechanisms of action. Against A. baumannii, similar potentiation was 
achieved with clarithromycin, erythromycin, fusidic acid, retapamulin, and 
rifampin. Susceptibility testing of the most effective antibiotic-SPR741 
combinations was extended to 25 additional multidrug-resistant or clinical 
isolates of E. coli and K. pneumoniae and 17 additional A. baumannii isolates in 
order to rank the potentiated antibiotics. SPR741 was also able to potentiate 
antibiotics that are substrates of the AcrAB-TolC efflux pump in E. coli, 
effectively circumventing the contribution of this pump to intrinsic antibiotic 
resistance. These studies support the further development of SPR741 in 
combination with conventional antibiotics for the treatment of Gram-negative 
bacterial infections.