# Acanthamoeba castellanii: Non-Steroidal Anti-Inflammatory Drugs Affect Adhesion, Motility, and Encystment, Suggesting a Link with a gp63-like Protein Candidate

## TL;DR

This study shows that common anti-inflammatory drugs can reduce the harmful behavior of Acanthamoeba castellanii, a parasite that causes serious eye infections.

## Contribution

The study identifies a potential role for a gp63-like protein in NSAID-sensitive pathways affecting Acanthamoeba pathogenicity.

## Key findings

- NSAIDs significantly reduced adhesion, migration, and encystment of Acanthamoeba castellanii.
- Drug-treated amoebae preserved actin fluorescence in co-cultures with epithelial cells.
- gp63-like protein showed increased colocalization with F-actin in cysts and migrating forms.

## Abstract

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.