Evidence of human‐like Ca²⁺ channels and effects of Ca²⁺ channel blockers in Acanthamoeba castellanii

Abstract

The evolution of voltage-gated calcium channel (Cav) in eukaryotes is an area of interest for biologists worldwide. The CLAN CL0030 and its family Ion_Trans 2 PF 07885 have been known to be present in prokaryotes, but the origin of these ion channels in Acanthamoeba spp. is yet to be determined. We inferred the origin of primitive forms of two-pore channels like proteins, human-like Cav 1.1 of L-type, and Cav subunit alpha-2/delta-1 in Acanthamoeba spp. early during evolution. By in-depth investigation into genomics, transcriptomics, use of bioinformatics tools and experimentations done with drugs like amlodipine and gabapentin on Acanthamoeba spp., we show the evidence of primitive forms of these channels in this protist pathogen.Genomics and transcriptomics of proteins ACA1_167020, 092610, and 270170 reflected their cellular expression in Acanthamoeba spp. We performed amino acid sequence homology, 3D structural modeling, ligand binding predictions, and dockings. Bioinformatics and 3D structural models show similarities between ACA1_167020, 092610, 270170, and different types of known human Cav. We show amoebicidal effects of amlodipine and gabapentin on Acanthamoeba spp., which can help design their structural analogs to target pathogenic genotypes of Acanthamoeba in diseases like Acanthamoeba keratitis and granulomatous amoebic encephalitis.

Repurposing drugs: Ca2+ ion dependency that can be exploited to treat keratitis caused by Acanthamoeba castellanii

“…Mitochondrial calcium accumulation has been studied in Acanthamoeba, but a channel system that allows Ca 2+ to get through the outer mitochondrial membrane in A. castellanii has not been explored previously [2] and the same is the case with our knowledge regarding the movement of this ion across the cell membrane. With the rationale that Ca 2+ ions play a vital role in Acanthamoeba spp., and the recently reported expression of human-like Ca 2+ ion-channels and the effects of Ca 2+ -channel blockers [3] in trophozoite forms of keratitis isolate of Acanthamoeba castellanii, we tested chelators of Ca 2+ ions in trophozoite forms of this protist pathogen. Our intention was to elucidate the role of extracellular Ca 2+ in the biology of Acanthamoeba spp.…”

“…The eukaryotic cells have intracellular storage of Ca 2+ ions but these storage sites get depleted if Ca 2+ ions are not replenished from extracellular resources. Recent studies have reported that the depletion of cytosolic Ca 2+ by blocking human-like voltage-gated Ca 2+ channels and calmodulin in Acanthamoeba castellanii with the drugs loperamide [3], amlodipine and gabapentin [4] exert amoebistatic and amoebicidal effect in low and high doses respectively. However, the effects of Ca 2+ chelation in extracellular space in A. castellanii have not been tested yet.…”

“…Note, pilocarpine 75 µg/ml was not effective in inducing Ca-influx when given with 5 and 10 µg/ml of EDTA (EDTA-1 + Pilocarpine 75 and EDTA-2 + Pilocarpine 75: last row), respectively Exploiting the voltage-gated Ca 2+ channels and calmodulin in unicellular microbes like Acanthamoeba spp. has proven to be effective in killing this microbe [3,4]. Ca 2+ ion chelation with EDTA and structural analogues can repurpose them for the treatment of AK and possibly granulomatous amoebic encephalitis.…”

“…In Acanthamoeba spp., Ca 2+ influx, its passage through voltage-gated channels [3], and cardinal cellular events that occur subsequent to the binding of this divalent cation to calmodulin [4] are novel molecular targets that can be exploited to gain therapeutic benefits in AK. By inhibition of the influx of Ca 2+ in the trophozoites of this protist pathogen, we show the vulnerability of Acanthamoeba spp.…”

Neuroleptic Drug Targets a Brain-Eating Amoeba: Effects of Promethazine on Neurotropic Acanthamoeba castellanii

“…As promethazine easily penetrates the blood–brain barrier and has a high margin of safety, it could be tested in animal models of AK and GAE. On the basis of the rationale of repurposing drugs that are already approved by drug-regulating authorities that target calcium channels, proteins like CaM and muscarinic receptor subtype mAChR1 in Acanthamoeba have been suggested to be tested in human trials in AK. , Successful results in AK and possibly GAE with promethazine and its structural analogs would encourage the repurposing of drugs that can speedily reach from bench to bedside in AK and possibly GAE.…”