Two amino-substituted Diphenyl Fumaramide Derivatives inhibit the virulence regulated by quorum sensing system of Pseudomonas aeruginosa

Abstract Aim Pseudomonas aeruginosa employs the quorum sensing (QS) system, a sophisticated cell-to-cell communication mechanism, to modulate the synthesis and secretion of a range of virulence factors, which contribute to the establishment of acute or chronic infections in hosts. This study seeks to attenuate the virulence of P. aeruginosa by inhibiting the QS system, thereby reducing its pathogenicity as a promising alternative to traditional antibiotics. Methods and Results Two compounds with an amino-substituted diphenyl fumaramide core, N1-(4-bromophenyl)-N4-(4'-oxo-3',4'-dihydro-1'H-spiro [cyclopentane-1,2'-quinazolin]-6'-yl) fumaramide (10D) and N1-(3-chloro-4-fluorophenyl)-N4-(4-oxo-3,4,4',5'-tetrahydro-1H,2'H-spiro [quinazoline-2,3'-thiophen]-6-yl) fumaramide (12A), were identified through in-silico screening. The QS inhibitory potential of both compounds was explored in vitro and in vivo. In in vitro experiments, neither compound exhibited bactericidal effects but significantly inhibited the production of QS-regulated extracellular protease and pyocyanin. Quantitative PCR analysis revealed that QS-activated genes and downstream virulence genes were transcriptionally suppressed by 10D or 12A. Molecular docking and molecular dynamics simulations predicted stable interactions between these compounds and the key QS regulators LasR and PqsR. When combined with polymyxin B, kanamycin, and levofloxacin, 10D and 12A exhibited synergistic antibacterial activity. Furthermore, compounds 10D and 12A significantly improved the survival of mice challenged with P. aeruginosa and effectively reduced the bacterial load in the lungs. Conclusion This study indicates that 10D and 12A possess considerable QS inhibitory potential, effectively attenuating the pathogenicity of P. aeruginosa. Moreover, the study offers structural insights and methodological guidance for the advancement of anti-virulence drug development.