Identification of 1-((2,4-Dichlorophenethyl)Amino)-3-Phenoxypropan-2-ol, a Novel Antibacterial Compound Active against Persisters of Pseudomonas aeruginosa

Abstract

Antibiotics typically fail to completely eradicate a bacterial population, leaving a small fraction of transiently antibiotic-tolerant persister cells intact. Persisters are therefore seen to be a major cause of treatment failure and greatly contribute to the recalcitrant nature of chronic infections. The current study focused on Pseudomonas aeruginosa, a Gram-negative pathogen belonging to the notorious ESKAPE group of pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) and, due to increasing resistance against most conventional antibiotics, posing a serious threat to human health. Greatly contributing to the difficult treatment of P. aeruginosa infections is the presence of persister cells, and elimination of these cells would therefore significantly improve patient outcomes. In this study, a smallmolecule library was screened for compounds that, in combination with the fluoroquinolone antibiotic ofloxacin, reduced the number of P. aeruginosa persisters compared to the number achieved with treatment with the antibiotic alone. Based on the early structure-activity relationship, 1-((2,4-dichlorophenethyl)amino)-3-phenoxypropan-2-ol (SPI009) was selected for further characterization. Combination of SPI009 with mechanistically distinct classes of antibiotics reduced the number of persisters up to 10 6 -fold in both lab strains and clinical isolates of P. aeruginosa. Further characterization of the compound revealed a direct and efficient killing of persister cells. SPI009 caused no erythrocyte damage and demonstrated minor cytotoxicity. In conclusion, we identified a novel antipersister compound active against P. aeruginosa with promising applications for the design of novel, case-specific combination therapies in the fight against chronic infections.

Antibacterial Activity of 1-[(2,4-Dichlorophenethyl)amino]-3-Phenoxypropan-2-ol against Antibiotic-Resistant Strains of Diverse Bacterial Pathogens, Biofilms and in Pre-clinical Infection Models

“…The activity of SPI009 was previously assessed in P. aeruginosa PA14 and several clinical isolates where combination with ofloxacin significantly decreased the persister fraction in all strains tested ( Liebens et al, 2017 ). In the present study, we challenged a panel of clinically relevant species, including the ESKAPE pathogens ( Figure 1A ), B. cenocepacia and E. coli ( Figure 1B ).…”

“…Antibacterial assays were performed on different clinically relevant pathogens as previously described ( Liebens et al, 2017 ). Briefly, stationary phase cultures were treated for 5 h with 17 or 34 μg/mL of SPI009 alone or in combination with an appropriate antibiotic to assess anti-bacterial and anti-persister effects, respectively.…”

“…Infection of human THP-1 cells was performed as described previously, with minor modifications ( Buyck et al, 2013 ). Since a newly synthesized batch of SPI009 was used for this experiment, cytotoxicity assessment via an LDH enzyme assay was repeated for the THP-1 cell line, as previously described ( Liebens et al, 2017 ). After THP-1 infection with P. aeruginosa PAO1 and subsequent removal of any non-phagocytozed or adherent bacteria, ciprofloxacin and SPI009 were added in final concentrations of, respectively, 0–20 μg/mL and 6.8 or 10.2 μg/mL.…”

“…We recently reported the identification of a novel anti-persister molecule capable of directly killing persister cells of P. aeruginosa ( Liebens et al, 2017 ). SPI009 was identified in a screening of 23,909 small molecules for compounds that decrease the persister fraction of P. aeruginosa in combination with the conventional antibiotic ofloxacin.…”

Unravelling mechanisms of meropenem induced persistence facilitates identification of GRAS compounds with anti-persister activity against Acinetobacter baumannii

“…The kinetics of inhibition of A. baumannii cells in the presence of meropenem (at 100X MIC) upon addition of thymol at different time intervals was studied, as described previously (80). SP cells of A. baumannii AYE were treated with meropenem (100X MIC) or combination of meropenem and 1X MIC of thymol.…”