Discovery of repurposing drug candidates for the treatment of diseases caused by pathogenic free-living amoebae.

Christopher A Rice,Beatrice L Colon,Emily Chen,Dennis E Kyle,Mitchell V Hull,Abhay R Satoskar

doi:10.1371/journal.pntd.0008353

Abstract

Diseases caused by pathogenic free-living amoebae include primary amoebic meningoencephalitis (Naegleria fowleri), granulomatous amoebic encephalitis (Acanthamoeba spp.), Acanthamoeba keratitis, and Balamuthia amoebic encephalitis (Balamuthia mandrillaris). Each of these are difficult to treat and have high morbidity and mortality rates due to lack of effective therapeutics. Since repurposing drugs is an ideal strategy for orphan diseases, we conducted a high throughput phenotypic screen of 12,000 compounds from the Calibr ReFRAME library. We discovered a total of 58 potent inhibitors (IC50 <1 μM) against N. fowleri (n = 19), A. castellanii (n = 12), and B. mandrillaris (n = 27) plus an additional 90 micromolar inhibitors. Of these, 113 inhibitors have never been reported to have activity against Naegleria, Acanthamoeba or Balamuthia. Rapid onset of action is important for new anti-amoeba drugs and we identified 19 compounds that inhibit N. fowleri in vitro within 24 hours (halofuginone, NVP-HSP990, fumagillin, bardoxolone, belaronib, and BPH-942, solithromycin, nitracrine, quisinostat, pabinostat, pracinostat, dacinostat, fimepinostat, sanguinarium, radicicol, acriflavine, REP3132, BC-3205 and PF-4287881). These compounds inhibit N. fowleri in vitro faster than any of the drugs currently used for chemotherapy. The results of these studies demonstrate the utility of phenotypic screens for discovery of new drugs for pathogenic free-living amoebae, including Acanthamoeba for the first time. Given that many of the repurposed drugs have known mechanisms of action, these compounds can be used to validate new targets for structure-based drug design.

Highlights

Naegleria fowleri, the causative agent of primary amoebic meningoencephalitis (PAM), was first discovered to be a human pathogen by Fowler and Carter after four fatal infections from 1961–1965 in Adelaide, Australia [1]
Due to poor detection and inadequate treatment options available for pathogenic Free-living amoebae (FLA), the fatality rates are still > 90% for the diseases caused by Balamuthia mandrillaris, Naegleria fowleri, and Acanthamoeba spp
This is extremely helpful for neglected diseases including pathogenic FLA where there is a need for new active therapies with limited budgets

Summary

Introduction

The causative agent of primary amoebic meningoencephalitis (PAM), was first discovered to be a human pathogen by Fowler and Carter after four fatal infections from 1961–1965 in Adelaide, Australia [1]. Once N. fowleri enters the nasal cavity, the amoebae traverse the cribriform plate the olfactory nerve to reach the frontal cerebral cortex of the brain where they cause hemorrhagic meningoencephalitis with the classical PAM symptoms of severe headache, stiff neck, hallucinations, seizures, coma, and, in 97% of cases, death [12,13]. Several recent PAM survival cases included inducement of therapeutic hypothermia (cooling of the core body temperature to 32–34 ̊C) to better manage intracranial pressure; this technique reduces reactive oxygen and nitrogen species, neural apoptosis, and proinflammatory cytokine levels, all of which increase brain injury through hyperinflammation [16,17]

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