Epigallocathechin-O-3-Gallate Inhibits Trypanothione Reductase of Leishmania infantum, Causing Alterations in Redox Balance and Leading to Parasite Death.

Job D F Inacio,Ana M Tomas,Helena Castro,Elmo E Almeida-Amaral,Myslene S Fonseca,Gabriel Limaverde-Sousa

doi:10.3389/fcimb.2021.640561

Abstract

Leishmania infantum is a protozoan parasite that causes a vector borne infectious disease in humans known as visceral leishmaniasis (VL). This pathology, also caused by L. donovani, presently impacts the health of 500,000 people worldwide, and is treated with outdated anti-parasitic drugs that suffer from poor treatment regimens, severe side effects, high cost and/or emergence of resistant parasites. In previous works we have disclosed the anti-Leishmania activity of (-)-Epigallocatechin 3-O-gallate (EGCG), a flavonoid compound present in green tea leaves. To date, the mechanism of action of EGCG against Leishmania remains unknown. This work aims to shed new light into the leishmanicidal mode of action of EGCG. Towards this goal, we first confirmed that EGCG inhibits L. infantum promastigote proliferation in a concentration-dependent manner. Second, we established that the leishmanicidal effect of EGCG was associated with i) mitochondria depolarization and ii) decreased concentration of intracellular ATP, and iii) increased concentration of intracellular H2O2. Third, we found that the leishmanicidal effect and the elevated H2O2 levels induced by of EGCG can be abolished by PEG-catalase, strongly suggesting that this flavonoid kills L. infantum promastigotes by disturbing their intracellular redox balance. Finally, we gathered in silico and in vitro evidence that EGCG binds to trypanothione reductase (TR), a central enzyme of the redox homeostasis of Leishmania, acting as a competitive inhibitor of its trypanothione substrate.

Highlights

Visceral leishmaniasis (VL) is a neglected tropical disease caused by Leishmania infantum and L. donovani that affects 500,000 people and is fatal in over 95% of cases if left untreated
Based on the observation that in mice Epigallocatechin 3-O-gallate (EGCG) can modulate two important antioxidant proteins, hepatic glutathione peroxidase and glutathione reductase (Dong et al, 2016), we considered here the possibility that EGCG activity in Leishmania interferes with the parasite antioxidant mechanisms In Leishmania spp. and related trypanosomatid organisms, redox balance is largely accomplished by the enzyme trypanothione reductase (Leroux and Krauth-Siegel, 2016)
To investigate whether synthesis of this oxidant occurred in promastigotes, we monitored its levels of this oxidant in EGCG-treated parasites (125 – 500 μM) during 72 h resorting to Amplex Red

Summary

Introduction

Visceral leishmaniasis (VL) is a neglected tropical disease caused by Leishmania infantum and L. donovani that affects 500,000 people and is fatal in over 95% of cases if left untreated. The disease has a high prevalence in the Americas, in Brazil where 96% of the cases are reported (Pan American Health Organization, 2018). In the absence of a vaccine, VL treatment is largely based on chemotherapy, pentavalent antimonials and amphotericin B being the most used drugs. Even though these treatments have saved thousands of lives, they can only be administered parenterally, present severe side effects and are expensive (Barral et al, 1991; Grimaldi and Tesh, 1993; Croft and Coombs, 2003; Amato et al, 2008). An increasing number of clinical relapses has been reported (Rijal et al, 2013) rendering the development of new, more effective, safer, and accessible drugs extremely urgent (Chappuis et al, 2007; Capela et al, 2019)

Objectives

Methods

Results

Conclusion