posted on 2021-08-17, 12:06authored byFadi Soukarieh, Alaa Mashabi, William Richardson, Eduard Vico Oton, Manuel Romero, Shaun N. Roberston, Scott Grossman, Tomas Sou, Ruiling Liu, Nigel Halliday, Irena Kukavica-Ibrulj, Roger C. Levesque, Christel A. S. Bergstrom, Barrie Kellam, Jonas Emsley, Stephan Heeb, Paul Williams, Michael J. Stocks, Miguel Cámara
P.
aeruginosa (PA) continues to
pose a threat to global public health due to its high levels of antimicrobial
resistance (AMR). The ongoing AMR crisis has led to an alarming shortage
of effective treatments for resistant microbes, and hence there is
a pressing demand for the development of novel antimicrobial interventions.
The potential use of antivirulence therapeutics to tackle bacterial
infections has attracted considerable attention over the past decades
as they hamper the pathogenicity of target microbes with reduced selective
pressure, minimizing the emergence of resistance. One such approach
is to interfere with the PA pqs quorum sensing system
which upon the interaction of PqsR, a Lys-R type transcriptional regulator,
with its cognate signal molecules 4-hydroxy-2-heptylquinoline (HHQ)
and 2-heptyl-3-hydroxy-4-quinolone (PQS), governs multiple virulence
traits and host–microbe interactions. In this study, we report
the hit identification and optimization of PqsR antagonists using
virtual screening coupled with whole cell assay validation. The optimized
hit compound 61 ((R)-2-(4-(3-(6-chloro-4-oxoquinazolin-3(4H)-yl)-2-hydroxypropoxy)phenyl)acetonitrile)
was found to inhibit the expression of the PA PpqsA promoter controlled by PqsR with an IC50 of 1 μM.
Using isothermal titration calorimetry, a Kd of 10 nM for the PqsR ligand binding domain (PqsRLBD)
was determined for 61. Furthermore, the crystal structure
of 61 with PqsRLBD was attained with a resolution
of 2.65 Å. Compound 61 significantly reduced levels
of pyocyanin, PQS, and HHQ in PAO1-L, PA14 lab strains and PAK6085
clinical isolate. Furthermore, this compound potentiated the effect
of ciprofloxacin in early stages of biofilm treatment and in Galleria mellonella infected with PA. Altogether,
this data shows 61 as a potent PqsR inhibitor with potential
for hit to lead optimization toward the identification of a PA QS
inhibitor which can be advanced into preclinical development.