title: Unveiling benznidazole's mechanism of action through overexpression of DNA repair proteins in Trypanosoma cruzi.
abstract: Benznidazole (BZ) is the most commonly used drug for the treatment of Chagas disease. Although BZ is known to induce the formation of free radicals and electrophilic metabolites within the parasite Trypanosoma cruzi, its precise mechanisms of action are still elusive. Here, we analyzed the survival of T. cruzi exposed to BZ using genetically modified parasites overexpressing different DNA repair proteins. Our results indicate that BZ induces oxidation mainly in the nucleotide pool, as heterologous expression of the nucleotide pyrophosphohydrolase MutT (but not overexpression of the glycosylase TcOgg1) increased drug resistance in the parasite. In addition, electron microscopy indicated that BZ catalyzes the formation of double-stranded breaks in the parasite, as its genomic DNA undergoes extensive heterochromatin unpacking following exposure to the drug. Furthermore, the overexpression of proteins involved in the recombination-mediated DNA repair increased resistance to BZ, reinforcing the idea that the drug causes double-stranded breaks. Our results also show that the overexpression of mitochondrial DNA repair proteins increase parasite survival upon BZ exposure, indicating that the drug induces lesions in the mitochondrial DNA as well. These findings suggest that BZ preferentially oxidizes the nucleotide pool, and the extensive incorporation of oxidized nucleotides during DNA replication leads to potentially lethal double-stranded DNA breaks in T. cruzi DNA.
authors.affiliation: Departamento de Bioquímica e Imunologia, Instituto de Ciências Biológicas, UFMG, Belo Horizonte, Minas Gerais.
authors.forename: Matheus Andrade
authors.initials: MA
authors.lastname: Rajão
authors.forename: Carolina
authors.initials: C
authors.lastname: Furtado
authors.forename: Ceres Luciana
authors.initials: CL
authors.lastname: Alves
authors.forename: Danielle Gomes
authors.initials: DG
authors.lastname: Passos-Silva
authors.forename: Michelle Barbi
authors.initials: MB
authors.lastname: de Moura
authors.forename: Bruno Luiz
authors.initials: BL
authors.lastname: Schamber-Reis
authors.forename: Marianna
authors.initials: M
authors.lastname: Kunrath-Lima
authors.forename: Aline Araújo
authors.initials: AA
authors.lastname: Zuma
authors.forename: João Pedro
authors.initials: JP
authors.lastname: Vieira-da-Rocha
authors.forename: Juliana Borio Ferreira
authors.initials: JB
authors.lastname: Garcia
authors.forename: Isabela Cecília
authors.initials: IC
authors.lastname: Mendes
authors.forename: Sérgio Danilo Junho
authors.initials: SD
authors.lastname: Pena
authors.forename: Andrea Mara
authors.initials: AM
authors.lastname: Macedo
authors.forename: Glória Regina
authors.initials: GR
authors.lastname: Franco
authors.forename: Nadja Cristhina
authors.initials: NC
authors.lastname: de Souza-Pinto
authors.forename: Marisa Helena Gennari
authors.initials: MH
authors.lastname: de Medeiros
authors.forename: Angela Kaysel
authors.initials: AK
authors.lastname: Cruz
authors.forename: Maria Cristina Machado
authors.initials: MC
authors.lastname: Motta
authors.forename: Santuza Maria Ribeiro
authors.initials: SM
authors.lastname: Teixeira
authors.forename: Carlos Renato
authors.initials: CR
authors.lastname: Machado
mesh_terms: D000818:Animals; D002470:Cell Survival; D014355:Chagas Disease / Q000188:drug therapy / Q000235:genetics / Q000469:parasitology; D045647:DNA Glycosylases / Q000235:genetics; D004260:DNA Repair / Q000187:drug effects; D045643:DNA Repair Enzymes / Q000235:genetics*; D016054:DNA, Protozoan / Q000187:drug effects; D004351:Drug Resistance / Q000235:genetics*; D006147:Guanine / Q000031:analogs & derivatives / Q000378:metabolism; D009593:Nitroimidazoles / Q000494:pharmacology*; D015800:Protozoan Proteins / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D014344:Trypanocidal Agents / Q000494:pharmacology*; D014349:Trypanosoma cruzi / Q000187:drug effects* / Q000235:genetics
chemical_list: D016054:DNA, Protozoan; D009593:Nitroimidazoles; D015800:Protozoan Proteins; D014344:Trypanocidal Agents; C024829:8-hydroxyguanine; D006147:Guanine; D045647:DNA Glycosylases; C061738:mutY adenine glycosylase; D045643:DNA Repair Enzymes; C009999:benzonidazole
keywords: Benznidazole; DNA repair; Trypanosoma cruzi; drug resistance; oxidative stress