Background: Loperamide is an anti-diarrheal drug prescribed for non-infectious
diarrhea. The drug is an opioid receptor agonist, blocker of voltage-dependent calcium channel
(Cav) and calmodulin (CaM) inhibitor on human cells. Loperamide has been reported to
exert anti-amoebic effects against pathogenic strains of Acanthamoeba castellanii.
Objectives: The precise mode of antibiotic action, cellular target homology with human
counterparts and the pattern of cell death induced by loperamide in Acanthamoeba castellanii
remain to be established. Additionally, we attempt to establish the presence a primitive
Cav in Acanthamoeba castellanii.
Methods: Bioinformatics, 3D structural modelling, ligand binding predictions and apoptotic/
amoebicidal assays were used in this study to answer the above queries. Amino acid
sequences and structural models were compared between human and A. castellanii proteins
that are involved in the regulation of calcium (Ca+2) homeostasis.
Results: Our results show that A. castellanii expresses similar, to near identical types of
primitive calcium channels Cav Ac and CaM that are well known targets of loperamide in
humans. The growth assays showed anti-amoebic effects of loperamide at different doses,
both alone and in combinations with other Ca+2- CaM inhibitors. The synergistic actions of
loperamide with haloperidol showed to be more amoebicidal than when either of them used
alone. Imaging with Annexin V, Acridine orange and Propidium iodide showed apoptosis in
A. castellanii at a dose of 100 µg/ml and necrosis at higher doses of 250 µg/ml.
Conclusion: Though, Acanthamoeba does not express a homolog of the human mu-opioid
receptor, but does shows evidence of the homologs for other known human targets of loperamide
that are involved in Ca+2 uptake and Ca+2 signal transduction pathways. This suggests
optimization of similar drug interactions with these targets may be useful in developing
new approaches to control the growth of this parasite and possibly the diseases caused by it.