Dioctanoyl Ultrashort Tetrabasic β-Peptides Sensitize Multidrug-Resistant Gram-Negative Bacteria to Novobiocin and Rifampicin

Abstract

Recently reported peptidomimetics with increased resistance to trypsin were shown to sensitize priority multidrug-resistant (MDR) Gram-negative bacteria to novobiocin and rifampicin. To further optimize proteolytic stability, β-amino acid-containing derivatives of these compounds were prepared, resulting in three dioctanoyl ultrashort tetrabasic β-peptides (dUSTBβPs). The nonhemolytic dUSTBβP 3, comprised of three β3-homoarginine residues and two fatty acyl tails eight carbons long, enhanced the antibacterial activity of various antibiotics from different classes. Notably, compound 3 retained the ability to potentiate novobiocin and rifampicin in wild-type Gram-negative bacteria against MDR clinical isolates of Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae. dUSTBβP 3 reduced the minimum inhibitory concentration of novobiocin and rifampicin below their interpretative susceptibility breakpoints. Furthermore, compound 3 exhibited improved in vitro stability (86.8 ± 3.7% remaining) relative to its α-amino acid-based counterpart (39.5 ± 7.4% remaining) after a 2 h incubation in human plasma.

Antimicrobial Peptide Mimics for Clinical Use: Does Size Matter?

“…These compounds can be cheaply produced in large amounts and with three diverse leads in clinical trials it is expected that these types of small synthetic mimics will play an important role in the future management of microbial infections. Recent studies on synergistic effects in combination with traditional antibiotics, further highlight the imminent role these compounds may have (103,106,178,179). Collectively, it is shown that a high antimicrobial activity can be obtained using diverse low molecular weight compounds and it is clear that shape matters more than size for optimal activity.…”

Effects of Lysine N-ζ-Methylation in Ultrashort Tetrabasic Lipopeptides (UTBLPs) on the Potentiation of Rifampicin, Novobiocin, and Niclosamide in Gram-Negative Bacteria

“…All UTBLPs were prepared by Fmoc SPPS [ 27 ] and as previously described [ 15 , 33 ]. The N-terminus of the amino acids was protected with Fmoc.…”

“…The in vitro antibacterial activity of the UTBLPs was assessed against wild-type and clinically isolated bacterial strains. To obtain the MIC of the compounds, the microbroth dilution susceptibility assay was performed according to the Clinical and Laboratory Standards Institute (CLSI, Wayne, PA, USA) guidelines [ 36 ] and as previously described [ 15 , 33 ]. Briefly, the compounds at varying concentrations were incubated with bacterial inoculum (5 × 10 5 CFU/mL final concentration) at 37 °C for 18 h. Growth in the form of turbidity was confirmed using an EMax Plus microplate reader (Molecular Devices, San Jose, CA, USA) at 590 nm.…”

“…The synergy of the UTBLPs with various antibiotics was assessed using checkerboard assay as previously described [ 15 , 33 ]. Briefly, the combination of antibiotic and OMP at varying concentrations were incubated with bacterial inoculum (5 × 10 5 CFU/mL final concentration) at 37 °C for 18 h. Growth in the form of turbidity was confirmed using an EMax Plus microplate reader (Molecular Devices, Union City, CA, USA) at 590 nm.…”

“…The ability of UTBLPs to permeabilize the OM was assessed using NPN as previously described with minor modifications [ 13 , 15 , 33 , 37 ]. In brief, NPN (10 μ M final concentration) was incubated with the cell suspension (OD 600 = 0.4–0.6) in buffer (5 mM HEPES, 5 mM glucose, 5 μM carbonyl cyanide 3-chlorophenylhydrazone, pH 7.2) at room temperature for 30 min in darkness.…”