Repurposing in vitro approaches for screening anti-parasitic drugs against the brain-eating amoeba Naegleria fowleri

Abstract

Naegleria fowleri is both a pathogenic and a free-living microbial eukaryote, responsible for the development of primary amoebic meningoencephalitis (PAM) in humans. PAM is a rapid, severe and fatal underestimated infectious disease, which has been reported in countries with warmer climates. The major drawbacks with PAM are the lack of effective therapies and delay in diagnosis. The current frontline treatment presents a low rate of recovery (5%) and severe adverse effects. For example, many drug candidates lack efficacy, since they do not effectively cross the blood-brain-barrier. Consequently, more effective drugs are urgently needed. Herein, we report a new in vitro method suitable for medium- and high-throughput drug discovery assays, using the closely related Naegleria gruberi as a model. We have subsequently used this method to screen a library of 1175 Food and Drug Administration-approved drugs. As a result, we present three drugs (camptothecin, pyrimethamine, and terbinafine) that can be repurposed, and are anticipated to readily cross the blood-brain-barrier with activity against Naegleria species in therapeutically achievable concentrations. Successively, we integrated several in vitro assays that resulted in identifying fast-acting and high amoebicidal drugs. In conclusion, we present a new approach for the identification of anti- Naegleria drugs along with three potential drug candidates for further development for the treatment of PAM.

A Fluorometric Assay for the In Vitro Evaluation of Activity against Naegleria fowleri Cysts

“…Moreover, this compound was the only one in which the anticyst activity is lower than the antitrophozoite one. The biochemical mechanism of the amphotericin B in N. fowleri is based on the lysis of the cell membrane and pore formation by binding to ergosterol and, hence, altering the membrane permeability ( 19 , 20 ). In the cystSt of N. fowleri , the cytoplasm is protected by a cyst wall whose composition is not clear ( 21 ); however, Chávez-Munguía et al reported the presence of enolase in the cyst wall ( 22 ).…”

Identification of new drug candidates against Trichomonas gallinae using high-throughput screening

“…Recently, a widely used process called drug repositioning that involves re-screening compounds with known biological activity and toxicity profiles has been found to be an effective strategy to accelerate drug discovery ( Xue et al, 2018 ; Martín-Escolano et al, 2021 ). However, the lack of standardized screening methods has stymied drug-repositioning efforts, with numerous studies relying on either the microscopy method or metabolically activated dyes (the resazurin microtiter method, the MTT colorimetric method) ( Duarte et al, 2009a ; Pothineni et al, 2016 ; Youssefi et al, 2017 ; Bailén et al, 2022 ).…”