Acanthamoeba castellanii, an opportunistic free-living amoeba, causes severe infections including Acanthamoeba keratitis. This exploratory study evaluated whether three non-steroidal anti-inflammatory drugs (NSAIDs)-acetylsalicylic acid, ibuprofen, and diclofenac (100 µM)-modulate pathogenicity-related processes in A. castellanii and explored the involvement of a gp63-like protein during encystment and adhesion. Trophozoites were continuously exposed to each drug and analyzed for adhesion, migration on host-derived discontinuous brain micropatterns, encystment efficiency, and parasite-induced cytoskeletal remodeling in MDCK epithelial cells. In silico docking was performed to assess potential drug-protein interactions. Drug exposure reduced adhesion with maximal inhibition at 60 min. After 1 h, migration decreased by 49%, 64%, and 38%, and encystment was reduced by 50%, 85%, and up to 90%, respectively, in cultures treated with acetylsalicylic acid, ibuprofen, and diclofenac. Co-incubation with untreated trophozoites lowered actin fluorescence to approximately 50%, whereas drug-treated co-cultures preserved fluorescence near control levels. Colocalization analysis showed increased spatial overlap between gp63-like protein and F-actin in cysts (~40%) and migrating trophozoites (~20%) compared with non-stimulated forms (~3.8%). Collectively, these findings suggest that NSAID-sensitive pathways influence host interaction, migration, and encystment in A. castellanii and allow for the proposal of gp63-like protein as a putative molecular component of the NSAIDs sensitive pathways.
Keywords: Acanthamoeba castellanii; NSAIDs; cyclooxygenase inhibitors; gp63-like; leishmanolysin.