A flow cytometer-based method to simultaneously assess activity and selectivity of compounds against the intracellular forms of Trypanosoma cruzi.
Chagas disease is a major unsolved health issue in Latin America and an emerging threat worldwide. New drugs are urgently needed for chemotherapy as those available (benznidazole and nifurtimox) have variable efficacy and elevated toxicity. Efforts are actually oriented to improve tools and technologies (e.g. transgenic parasites, flow cytometry or image-based systems) for the screening of large numbers of candidate compounds for their activity against Trypanosoma cruzi (T. cruzi). Methods that test drug efficacy and selectivity in the same assay are suitable to accelerate the process of drug discovery. Here, we developed a GFP expressing T. cruzi from a moderate virulence stock and confirmed that the transgenic parasite retained the biological characteristics of the parental strain. With this tool, we established a flow cytometer-based method to simultaneously test drug activity against intracellular amastigotes and toxicity to the host cell. This one-step procedure allows determining the selectivity index of the tested compound in a sensitive and accurate manner even with low infection rates. This method can provide additional information on the interactions between drug, parasites and host cell and could be adapted to other trypanosomatids and protozoa with intracellular multiplication.
Facultad de Medicina, Instituto de Investigaciones en Microbiología y Parasitología Médicas (IMPaM),UBA-CONICET, Universidad de Buenos, Paraguay 2155, Ciudad Autónoma de Buenos Aires CC1121ABG, Argentina. Electronic address: cristiangam_@hotmail.com.
Cristian Gabriel
CG
Miranda
Facultad de Medicina, Instituto de Investigaciones en Microbiología y Parasitología Médicas (IMPaM),UBA-CONICET, Universidad de Buenos, Paraguay 2155, Ciudad Autónoma de Buenos Aires CC1121ABG, Argentina. Electronic address: melisolana@yahoo.com.ar.
Maria Elisa
ME
Solana
Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Investigaciones en Ingeniería Genética y Biología Molecular (INGEBI-CONICET), Vuelta de Obligado 2490 Piso 2, Ciudad Autónoma de Buenos Aires C1428ADN, Argentina. Electronic address: amycurto@dna.uba.ar.
Maria de Los Angeles
MLA
Curto
Facultad de Medicina, Instituto de Investigaciones en Microbiología y Parasitología Médicas (IMPaM),UBA-CONICET, Universidad de Buenos, Paraguay 2155, Ciudad Autónoma de Buenos Aires CC1121ABG, Argentina. Electronic address: paradife@fmed.uba.ar.
Estela Maria
EM
Lammel
Laboratorio de Biología Molecular de la Enfermedad de Chagas, Instituto de Investigaciones en Ingeniería Genética y Biología Molecular (INGEBI-CONICET), Vuelta de Obligado 2490 Piso 2, Ciudad Autónoma de Buenos Aires C1428ADN, Argentina. Electronic address: schijman@dna.uba.ar.
Alejandro Gabriel
AG
Schijman
Facultad de Medicina, Instituto de Investigaciones en Microbiología y Parasitología Médicas (IMPaM),UBA-CONICET, Universidad de Buenos, Paraguay 2155, Ciudad Autónoma de Buenos Aires CC1121ABG, Argentina. Electronic address: catalina.alba@gmail.com.
Catalina Dirney
CD
Alba Soto
D014355:Chagas Disease / Q000188:drug therapy* / Q000469:parasitology*; D055808:Drug Discovery / Q000379:methods*; D005434:Flow Cytometry; D006801:Humans; D009547:Nifurtimox / Q000627:therapeutic use*; D009593:Nitroimidazoles / Q000627:therapeutic use*; D014344:Trypanocidal Agents / Q000627:therapeutic use*; D014349:Trypanosoma cruzi / Q000187:drug effects*
D009593:Nitroimidazoles; D014344:Trypanocidal Agents; D009547:Nifurtimox; C009999:benzonidazole
Compound screening; Cytotoxicity; Drug efficacy; Flow cytometry; Fluorescent parasites; Propidium iodide; Selectivity index; Trypanosoma cruzi