Pharmacological inhibition of Trypanosoma cruzi aldo-keto reductase (TcAKR) and its effect on benznidazole resistance.

Chagas disease, caused by the protozoan Trypanosoma cruzi, has become a global health concern due to increased globalization. Several studies suggest that the aldo-keto reductase from T. cruzi (TcAKR) is involved in resistance to benznidazole, the drug commonly used to treat this infection. To further support the role of TcAKR in drug resistance, we evaluated its interaction with four compounds -quercetin, flufenamic acid, phenolphthalein, and menadione-previously reported as inhibitors of other AKRs. Molecular docking was performed to assess affinity and molecular specific interactions, and the inhibitory effects of these compounds on both TcAKR activities -aldo-keto reductase and quinone-oxidoreductase- were experimentally determined. Binding affinities, in decreasing order, were: quercetin > flufenamic acid > phenolphthalein > menadione. Both quercetin and flufenamic acid interacted with amino acid residues located outside the enzyme's active site. Quercetin completely inhibited both TcAKR activities, while flufenamic acid inhibited approximately 50 %. Phenolphthalein and menadione showed low levels of inhibition. The inhibition profiles of quercetin and flufenamic acid were consistent with a noncompetitive mechanism. The effect of quercetin on benznidazole resistance was evaluated in transfected parasites overexpressing TcAKR, which are 1.8-fold more resistant to this drug. Quercetin treatment restored benznidazole sensitivity in these parasites, reducing the IC₅₀ to levels comparable to those of control parasites. These results provide further evidence of TcAKR's involvement in benznidazole resistance and suggest that its inhibition can enhance treatment efficacy.