Autophagy modulation by OmpA-targeted lead compounds restores autophagic flux, reduces biofilm, and enhances immune response in <i>Acinetobacter baumannii</i> -infected pulmonary cells

ABSTRACT Acinetobacter baumannii is an opportunistic pathogen known for causing severe nosocomial infections. Autophagy, a key host defense mechanism against invading pathogens, is modulated by the virulence factor OmpA. OmpA is also responsible for bacterial adherence to host cells. Targeting OmpA provides a potential strategy to counteract these autophagic disruptions. In silico analysis (molecular mechanics and molecular dynamics simulations) and experimental validation of identified small molecule lead against OmpA were performed. These leads were tested in A549 cells infected with A. baumannii , revealing significant autophagic modulations. mRNA expression analysis showed accumulation of the autophagy marker LC3B and suppression of elongation-associated genes upon infection. Confocal microscopy of mCherry-GFP-LC3-transfected cells demonstrated a reduced mCherry/GFP ratio, indicating downregulated autophagic flux during infection. Treatment with the leads reversed these effects, restoring autophagic activity to levels seen in uninfected cells. Additionally, SEM analysis confirmed that the leads inhibited OmpA-mediated biofilm formation. They also reduced the internalization of A. baumannii in A549 cells. The leads also enhanced IFN-γ and IL-13 expression while reducing IL-10, promoting immune clearance of A. baumannii via autophagy modulation and suppression of Treg functions. These findings suggest that the identified leads hold promise as autophagy modulators, potentially regressing A. baumannii’s ability to escape host defense. IMPORTANCE Autophagic escape of Acinetobacter baumannii is mediated by OmpA; therefore, targeting OmpA represents a promising strategy to counteract autophagy disruption. The identified lead effectively reverses autophagy modulation, inhibits OmpA-mediated biofilm formation and bacterial internalization, and promotes a pro-clearance immune response.