Cephalosporin-3'-Diazeniumdiolate NO Donor Prodrug PYRRO-C3D Enhances Azithromycin Susceptibility of Nontypeable Haemophilus influenzae Biofilms.

Abstract

PYRRO-C3D is a cephalosporin-3-diazeniumdiolate nitric oxide (NO) donor prodrug designed to selectively deliver NO to bacterial infection sites. The objective of this study was to assess the activity of PYRRO-C3D against nontypeable Haemophilus influenzae (NTHi) biofilms and examine the role of NO in reducing biofilm-associated antibiotic tolerance. The activity of PYRRO-C3D on in vitro NTHi biofilms was assessed through CFU enumeration and confocal microscopy. NO release measurements were performed using an ISO-NO probe. NTHi biofilms grown on primary ciliated respiratory epithelia at an air-liquid interface were used to investigate the effects of PYRRO-C3D in the presence of host tissue. Label-free liquid chromatography-mass spectrometry (LC/MS) proteomic analyses were performed to identify differentially expressed proteins following NO treatment. PYRRO-C3D specifically released NO in the presence of NTHi, while no evidence of spontaneous NO release was observed when the compound was exposed to primary epithelial cells. NTHi lacking β-lactamase activity failed to trigger NO release. Treatment significantly increased the susceptibility of in vitro NTHi biofilms to azithromycin, causing a log fold reduction (10-fold reduction or 1-log-unit reduction) in viability (P < 0.05) relative to azithromycin alone. The response was more pronounced for biofilms grown on primary respiratory epithelia, where a 2-log-unit reduction was observed (P < 0.01). Label-free proteomics showed that NO increased expression of 16 proteins involved in metabolic and transcriptional/translational functions. NO release from PYRRO-C3D enhances the efficacy of azithromycin against NTHi biofilms, putatively via modulation of NTHi metabolic activity. Adjunctive therapy with NO mediated through PYRRO-C3D represents a promising approach for reducing biofilm-associated antibiotic tolerance.

• Authors: Samuel A Collins, Michael J Kelso, Ardeshir Rineh, Nageshwar R Yepuri, Janice Coles, Claire L Jackson, Georgia D Halladay, Woolf T Walker, Jeremy S Webb, Luanne Hall-Stoodley, Gary J Connett, Martin Feelisch, Saul N Faust, Jane S A Lucas, Raymond N Allan
• Journal: Antimicrobial agents and chemotherapy (Antimicrob Agents Chemother) Vol. 61 Issue 2 (02 2017) ISSN: 1098-6596 [Electronic] United States
• PMID: 27919896 (Publication Type: Journal Article)
• Copyright: Copyright © 2017 American Society for Microbiology.

Chemical References

• Anti-Bacterial Agents
• Azo Compounds
• Cephalosporins
• Nitric Oxide Donors
• Nitrogen Oxides
• Prodrugs
• diazeniumdiolate
• Azithromycin
• beta-Lactamases

Topics

• Anti-Bacterial Agents (pharmacology)
• Azithromycin (pharmacology)
• Azo Compounds (pharmacology)
• Biofilms (drug effects)
• Cephalosporins (pharmacology)
• Chromatography, Liquid
• Drug Resistance, Bacterial
• Haemophilus influenzae (drug effects)
• Mass Spectrometry
• Microbial Sensitivity Tests
• Nitric Oxide Donors (pharmacology)
• Nitrogen Oxides (metabolism)
• Prodrugs (pharmacology)
• Proteomics
• beta-Lactamases (metabolism)