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Cephalosporin-3’ -diazeniumdiolate NO-donor prodrug PYRRO-C3D enhances azithromycin susceptibility of Non-typeable Haemophilus influenzae biofilms

Cephalosporin-3’ -diazeniumdiolate NO-donor prodrug PYRRO-C3D enhances azithromycin susceptibility of Non-typeable Haemophilus influenzae biofilms
Cephalosporin-3’ -diazeniumdiolate NO-donor prodrug PYRRO-C3D enhances azithromycin susceptibility of Non-typeable Haemophilus influenzae biofilms
Objectives: 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 non-typeable Haemophilus influenzae (NTHi) biofilms and examine the role of NO in reducing biofilm-associated antibiotic tolerance.

Methods: 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 LC/MS proteomic analyses were performed to identify differentially expressed proteins following NO treatment.

Results: 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 ?-lactamase activity failed to trigger NO release. Treatment significantly increased the susceptibility of in vitro NTHi biofilms to azithromycin, causing a log-fold 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 reduction was observed (p<0.01). Label-free proteomics showed that NO increased expression of sixteen proteins involved in metabolic and transcriptional/translational functions.

Conclusions: 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.
0066-4804
e02086-16
Collins, Samuel A.
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Kelso, Michael J.
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Rineh, Ardeshir
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Yepuri, Nageshwar R.
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Coles, Janice
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Jackson, Claire L.
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Halladay, Georgia D.
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Walker, Woolf T.
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Webb, Jeremy S.
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Hall-Stoodley, Luanne
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Connett, Gary J.
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Feelisch, Martin
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Faust, Saul N.
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Lucas, Jane S.A.
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Allan, Raymond N.
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Collins, Samuel A.
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Kelso, Michael J.
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Rineh, Ardeshir
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Yepuri, Nageshwar R.
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Coles, Janice
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Jackson, Claire L.
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Halladay, Georgia D.
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Walker, Woolf T.
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Webb, Jeremy S.
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Hall-Stoodley, Luanne
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Connett, Gary J.
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Feelisch, Martin
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Faust, Saul N.
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Lucas, Jane S.A.
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Allan, Raymond N.
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Collins, Samuel A., Kelso, Michael J., Rineh, Ardeshir, Yepuri, Nageshwar R., Coles, Janice, Jackson, Claire L., Halladay, Georgia D., Walker, Woolf T., Webb, Jeremy S., Hall-Stoodley, Luanne, Connett, Gary J., Feelisch, Martin, Faust, Saul N., Lucas, Jane S.A. and Allan, Raymond N. (2017) Cephalosporin-3’ -diazeniumdiolate NO-donor prodrug PYRRO-C3D enhances azithromycin susceptibility of Non-typeable Haemophilus influenzae biofilms. Antimicrobial Agents and Chemotherapy, 61 (2), e02086-16. (doi:10.1128/AAC.02086-16). (PMID:27919896)

Record type: Article

Abstract

Objectives: 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 non-typeable Haemophilus influenzae (NTHi) biofilms and examine the role of NO in reducing biofilm-associated antibiotic tolerance.

Methods: 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 LC/MS proteomic analyses were performed to identify differentially expressed proteins following NO treatment.

Results: 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 ?-lactamase activity failed to trigger NO release. Treatment significantly increased the susceptibility of in vitro NTHi biofilms to azithromycin, causing a log-fold 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 reduction was observed (p<0.01). Label-free proteomics showed that NO increased expression of sixteen proteins involved in metabolic and transcriptional/translational functions.

Conclusions: 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.

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Accepted/In Press date: 26 November 2016
e-pub ahead of print date: 5 December 2016
Published date: February 2017
Organisations: Clinical & Experimental Sciences

Identifiers

Local EPrints ID: 403594
URI: http://eprints.soton.ac.uk/id/eprint/403594
ISSN: 0066-4804
PURE UUID: 6eb2bda0-0626-4fde-94db-faf8e4d5f051
ORCID for Claire L. Jackson: ORCID iD orcid.org/0000-0002-1200-0935
ORCID for Jeremy S. Webb: ORCID iD orcid.org/0000-0003-2068-8589
ORCID for Gary J. Connett: ORCID iD orcid.org/0000-0003-1310-3239
ORCID for Martin Feelisch: ORCID iD orcid.org/0000-0003-2320-1158
ORCID for Saul N. Faust: ORCID iD orcid.org/0000-0003-3410-7642
ORCID for Jane S.A. Lucas: ORCID iD orcid.org/0000-0001-8701-9975

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Date deposited: 06 Dec 2016 16:57
Last modified: 16 Mar 2024 04:09

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Contributors

Author: Samuel A. Collins
Author: Michael J. Kelso
Author: Ardeshir Rineh
Author: Nageshwar R. Yepuri
Author: Janice Coles
Author: Claire L. Jackson ORCID iD
Author: Georgia D. Halladay
Author: Woolf T. Walker
Author: Jeremy S. Webb ORCID iD
Author: Luanne Hall-Stoodley
Author: Gary J. Connett ORCID iD
Author: Martin Feelisch ORCID iD
Author: Saul N. Faust ORCID iD
Author: Jane S.A. Lucas ORCID iD
Author: Raymond N. Allan

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