Repurposing Azithromycin and Rifampicin Against Gram-Negative Pathogens by Combination With Peptidomimetics

Abstract

Synthetic peptidomimetics may be designed to mimic functions of antimicrobial peptides, including potentiation of antibiotics, yet possessing improved pharmacological properties. Pairwise screening of 42 synthetic peptidomimetics combined with the antibiotics azithromycin and rifampicin in multidrug-resistant (MDR) Escherichia coli ST131 and Klebsiella pneumoniae ST258 led to identification of two subclasses of α-peptide/β-peptoid hybrids that display synergy with azithromycin and rifampicin (fractional inhibitory concentration indexes of 0.03-0.38). Further screening of the best three peptidomimetics in combination with a panel of 21 additional antibiotics led to identification of peptidomimetics that potentiated ticarcillin/clavulanate and erythromycin against E. coli, and clindamycin against K. pneumoniae. The study of six peptidomimetics was extended to Pseudomonas aeruginosa, confirming synergy with antibiotics for five of them. The most promising compound, H-(Lys-βNPhe)₈-NH₂, exerted only a minor effect on the viability of mammalian cells (EC₅₀ ≥ 124-210 μM), and thus exhibited the highest selectivity toward bacteria. This compound also synergized with rifampicin and azithromycin at sub-micromolar concentrations (0.25-0.5 μM), thereby inducing susceptibility to these antibiotics at clinically relevant concentrations in clinical MDR isolates. This peptidomimetic lead and its analogs constitute promising candidates for efficient repurposing of rifampicin and azithromycin against Gram-negative pathogens.

Keywords: Gram-negative; antibiotic adjuvant; antibiotic potentiation; multidrug resistance; peptidomimetic; synergy.

Figure 1

Structures of the α-peptide/β-peptoid hybrids listed in Table 2 and similar compounds that exhibit activity in vivo (Zaknoon et al., ; Czyzewski et al., , respectively).

Figure 2

Peptidomimetic 1 potentiates activity of azithromycin and rifampicin in growth-curve assays (A–F). Growth of E. coli ST131 (A,D), K. pneumoniae ST258 (B,E) and P. aeruginosa ATCC 27853 (C,F) in absence or presence of antibiotic, peptidomimetic (PM) 1 or their combinations were recorded at regular intervals by measuring the optical density (OD) of each culture at 600 nm, and then these data were graphed over time (h). Initial bacterial concentration was ~5 ×10⁵ CFU/mL. The MIC values of each compound are listed in Table 6.

Figure 3

Peptidomimetic 1-combinations enhance bactericidal activity (A–F). Time-kill kinetics for E. coli ST131 (A,D), K. pneumoniae ST258 (B,E) and P. aeruginosa ATTC 27853 (C,F) exposed to peptidomimetic (PM) 1, azithromycin (AZM), rifampicin (RIF), peptidomimetic-antibiotic combinations or without treatment (controls); CFU/mL data are graphed at time points 0, 1, 2, 4, 8, and 24 h of exposure. Initial bacterial concentration was ~10⁶ CFU/mL. The MBC values of each compound are listed in Table 6.