In vitro susceptibility of Trypanosoma cruzi discrete typing units (DTUs) to benznidazole: A systematic review and meta-analysis

Abstract

Background: Chagas disease, a neglected tropical disease endemic to Latin America caused by the parasite Trypanosoma cruzi, currently affects 6-7 million people and is responsible for 12,500 deaths each year. No vaccine exists at present and the only two drugs currently approved for the treatment (benznidazole and nifurtimox), possess serious limitations, including long treatment regimes, undesirable side effects, and frequent clinical failures. A link between parasite genetic variability and drug sensibility/efficacy has been suggested, but remains unclear. Therefore, we investigated associations between T. cruzi genetic variability and in vitro benznidazole susceptibility via a systematic article review and meta-analysis.

Methodology/principal findings: In vitro normalized benznidazole susceptibility indices (LC50 and IC50) for epimastigote, trypomastigote and amastigote stages of different T. cruzi strains were recorded from articles in the scientific literature. A total of 60 articles, which include 189 assays, met the selection criteria for the meta-analysis. Mean values for each discrete typing unit (DTU) were estimated using the meta and metaphor packages through R software, and presented in a rainforest plot. Subsequently, a meta-regression analysis was performed to determine differences between estimated mean values by DTU/parasite stage/drug incubation times. For each parasite stage, some DTU mean values were significantly different, e.g. at 24h of drug incubation, a lower sensitivity to benznidazole of TcI vs. TcII trypomastigotes was noteworthy. Nevertheless, funnel plots detected high heterogeneity of the data within each DTU and even for a single strain.

Conclusions/significance: Several limitations of the study prevent assigning DTUs to different in vitro benznidazole sensitivity groups; however, ignoring the parasite's genetic variability during drug development and evaluation would not be advisable. Our findings highlight the need for establishment of uniform experimental conditions as well as a screening of different DTUs during the optimization of new drug candidates for Chagas disease treatment.

Fig 1. PRISMA flow diagram process for selection of eligible articles and assays.

The flow diagram shows the literature search and assays selection process employed for the meta-analysis of benznidazole LC50/IC50 mean values for different T. cruzi DTUs. *If a single assay was available for a given “parasite stage/drug incubation time/DTU” combination, it was deleted from the meta-analysis.

Fig 2. Meta-analysis of benznidazole LC50 and IC50 mean values.

Rainforest plot diplaying mean LC50 and IC50 values for benznidazole (Bz) and their 95% confidence intervals (x-axis) estimated by meta-analysis. Combined assays per parasite stage [“E” for epimastigote (blue), “T” for trypomastigote (black) and “A” for amastigote (red)], drug incubation times and DTUs are shown (y-axis); the number of assays per group is displayed in brackets. Mean values and confidence intervals (CI) are shown at the right side of the figure. For comparison of results between DTUs, statistics were applied only between groups belonging to the same parasite stage-incubation time; only significant p-values (< 0.05) are reported.