Acanthamoeba spp. is known to cause a sight-threatening Acanthamoeba keratitis (AK) and a usually fatal encephalitis called granulomatous amebic encephalitis (GAE). Though a successful prognosis of AK depends upon an early diagnosis, the availability of safer drugs to successfully treat AK remains a challenge. The rarity of AK has made the pharmaceutical industry reluctant to invest in the development of drugs against this vision threating keratitis. After AK was recognized as an orphan disease, a funded project, the Orphan Drug for AK (ODAK), the drug polyhexamethylene biguanide (PHMB) has been studied in Acanthamoeba polyphaga keratitis model [1]. There are reports that Acanthamoeba can persist causing an active infection despite treatment with chlorhexidine or PHMB [2]. Recent discoveries of cell surface receptors and druggable proteins in pathogenic variants of Acanthamoeba castellanii have revived the interest of repurposing of some anticholinergic drugs like atropine that are already used in non-infectious ocular disorders. The evidence of the presence of human-like muscarinic receptors in Acanthamoeba spp. [3], that are known targets of atropine, supports the possible forte of repurposing atropine and its structural analogs in AK. Furthermore, very convincing indications have come from retrospective studies where atropine was reported to treat  cases of AK without the use of any concurrent anti-amebic agents [4, 5]. Since atropine is already given as an adjuvant drug in cases of AK, this novel action of atropine has the potential to make it the drug of choice in AK if large-scale human trials provide definitive results.

Atropine used in AK could have a clear superiority over the newer drugs as it is already known for its safety in the eye and is in use for diverse ocular indications. Atropine and analogs like dicyclomine and pirenzepine that have proven to be amoebicidal in Acanthamoeba castellanii in vitro [3] (Fig. 1) could also be investigated for their use in AK in an animal model of this disease. Dicyclomine and pirenzepine, of which the latter is a more specific muscarinic M1 receptor (mAChR1) antagonist, have proven to be more effective amoebicidal agents in doses lower than atropine in vitro studies [5] (Fig. 1d), but these drugs need to be evaluated for their ocular safety and efficacy in animal model and in human trials. Use of eye drops containing polyethylene glycol 400 0.4% and propylene glycol 0.3% have also been reported to be amoebicidal and cysticidal. A decrease in the mitochondrial membrane potential and ATP levels  were shown to induce chromatin condensation in Acanthamoeba spp. in a study showing the toxic effects of selected proprietary dry eye drops [6].

Fig. 1
figure 1

A Histogram showing the results of muscarinic receptor antagonist drugs (pirenzepine 100 µg/ml and dicyclomine 90 µg/ml) on 0.5 × 106 of the trophozoites of Acanthamoeba castellanii. Experiments were performed in triplicates. One-way Anova showed a P-value of 0.0003. Dunnet’s comparison test revealed a P-value < 0.01 for –ve control vs dicyclomine 90 µg/ml and pirenzepine 100 µg/ml. Images show Acanthamoeba trophozoites in growth medium (a), solvent control (b), compared with 100 µg/ml of pirenzepine (c), dicyclomine (d), and 200 µg/ml of atropine (e), where the counts appear to be reduced, reflecting the anti-proliferative effects of these drugs in Acanthamoeba trophozoites

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AK, wound infection and GAE are notorious illnesses that are caused by Acanthamoeba spp., which are resistant to many anti-amebic drugs used at present. The prognosis of infections caused by this protist pathogen is very poor, which almost always results in blindness in AK (Fig. 2) and death in GAE. Development of new drugs is challenging and difficult as it takes a very long period to evaluate them for their safety and efficacies. Acetylcholine is known to stimulated the growth and proliferation of Acanthamoeba castellanii (Fig. 2a) [3] and anticholinergics like atropine, dicyclomine and pirenzepine antagonize strong proliferative signals generated from mAChR1 receptors (Fig. 2a) to exert an amoebicidal and cysticidal effects [3, 4]. These drugs offer a unique opportunity to repurpose them for their utility in AK [5] which is known to be common in contact lens users (Fig. 2b). Anticholinergic approved by FDA and their synthetic structural analogs could be speedily brought to human trials and clinical utility in infections like AK and GAE caused by Acanthamoeba spp.

Fig. 2
figure 2

a  Schematic figure showing the effects of  cholinergic  agonist and antagonists in Acanthamoeba trophozoites. Muscarinic receptor stimulation has a growth promoting effect that results in proliferation in Acanthamoeba via calmodulin-AKT and calcium influx via voltage- gated calcium channels. Antagonism of muscarinic like-homolog, the Acanthamoeba muscarine binding (AMB) receptor that has been recently reported in this protist pathogen by anticholinergic drugs (a) results in reduced proliferation and growth. b Shows sequence of events that follow the use of infected contact lens. Contaminated lens cleaning liquids are the most common source of infection that leads to Acanthamoeba keratitis (AK). Atropine used in the past has been reported to treat AK cases possibly by antagonizing the AMB receptor and its downstream pathways (a)

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