Oral Presentation Australian Society for Microbiology Annual Scientific Meeting 2018

Identification of small molecules as novel therapeutic options that suppress virulence in Neisseria gonorrhoeae and increase susceptibility to CAMPs (#45)

Christopher A Mullally 1 , Keith Stubbs 1 , Anandhi Anandan 1 , Genevieve Evans 1 , Martin Scanlon 2 , Gary A Jarvis 3 , Constance John 3 , Kathy Fuller 1 , Alice Vrielink 1 , Charlene M Kahler 1
  1. University of Western Australia, Perth, W.A., Australia
  2. Monash University, Melbourne, Victoria, Australia
  3. University of California San Francisco, San Francisco, California, United States of America

Neisseria gonorrhoeae is an exclusively human pathogen that most commonly infects the urogenital tract resulting in gonorrhoea. Empirical treatment of gonorrhea requires antibiotics, but multi-drug resistance has occurred to all first-line treatments thus resulting in the imperative to find new treatment options. The enzyme lipooligosaccharide phosphoethanolamine transferase A (EptA) is responsible for the addition of phosphoethanolamine (PEA) to lipid A as it is transported through the periplasm to the outer membrane. The addition of PEA to lipid A is essential for bacterial resistance to cationic antimicrobial peptides (CAMPs) and for attachment to human epithelial cells. We hypothesised that small molecules that inhibit EptA will result in increased sensitivity to CAMPs and enhance natural clearance of gonococci via the human innate immune response.

A library of 250 compounds has been synthesised and tested using an in vitro microbroth dilution assay against the reference strain N. gonorrhoeae FA1090. Seventy-four of these compounds have enhanced the sensitivity of strain FA1090 to polymyxin B (PxB), a CAMP. Compound 2B7 increased the sensitivity of strain FA1090 to PxB by 4-fold. MALDI-TOF MS analysis of lipid A extracted from FA1090 cells treated with the compound revealed a ~20% decrease in PEA decoration of the lipid A when compared to untreated bacteria. This was validated by a cytokine assay using THP-1 cells exposed to wild-type LOS and LOS from bacteria treated with the compound. The LOS from the compound treated cells showed a 55% decrease in TNF╬▒ induction consistent with reduced cytotoxicity resulting from the inhibition of PEA decoration of lipid A. To further confirm these observations a fluorescent assay using dansyl-polymyxin is being trailed.

In conclusion, small molecules can be designed to bind and inhibit EptA and can sensitise N. gonorrhoeae to killing by cationic antimicrobial peptides.