Abstract

Acanthamoeba castellanii is a free-living amoeba that exists in two different forms, the trophozoite, and the cyst. A. castellanii is known to cause two rare infections: Acanthamoeba keratitis (AK), a disease where the parasites infect the cornea, and a central nervous system (CNS) infection, known as granulomatous amoebic encephalitis (GAE). Patients with AK often lose significant vision in the infected eye and, if both eyes are infected, may lead to blindness. The other infection caused by this parasite, GAE, has a mortality rate of around 95%. The current chemotherapeutic drugs have not been able to reduce the morbidity and mortality associated with AK and GAE and the cost of de novo drug development for such rare diseases has led to the development of alternative approaches for the identification of drug candidates. This is the driving force for the work presented in this thesis and is the overarching theme for the papers included in this work. The publications contributing to this thesis describe work on the discovery of novel drug targets in Acanthamoeba and the use of this information to identify drugs that could be re-purposed for the treatment of AK and GAE. Acanthamoeba castellanii was shown to express human-like calcium channels, calcium regulating adapter proteins, G-protein coupled receptors and muscarinic receptor-like proteins that are needed for growth and proliferation. Bioinformatic analysis based around amino acid sequence homology, modeling, drug docking studies and transcriptomic profiles of Acanthamoeba revealed these proteins as possible drug targets. There are currently a range of drugs that are antagonists for these targets in humans and these are used for a variety of non-infectious disease presentations. Our research studied loperamide, amlodipine, digoxin, amiodarone, anticholinergic agents like procyclidine, dicyclomine and atropine. Studies were able to show that these drugs have activity against trophozoite and cystic forms of Acanthamoeba spp. and that many work via disrupting calcium homeostasis. These drugs are fully characterised and their profile of adverse effects and the margin of safety and toxicity is well known. These drugs could now be evaluated for their clinical utility for the treatment of AK and GAE. This is much faster than for traditional novel drug discovery. Thus, the major conclusion from this body of work is that the repurposing of drugs, already in clinical use for the treatment of non-infectious diseases in humans, will provide an alternative and viable option in drug development against AK and GAE.