Giardia duodenalis flavohemoglobin is a target of 5-nitroheterocycle and benzimidazole compounds acting as enzymatic inhibitors or subversive substrates

Abstract

Giardia duodenalis causes giardiasis in humans, companion, livestock and wild animals. Control of infection involves drugs as benzimidazoles (e.g., albendazole, ABZ) and 5-nitroheterocyclics [5-NHs: metronidazole (MTZ), furazolidone (FZD), nitazoxanide (NTZ)] as first-line agents. During infection, Giardia is exposed to immune and pro-oxidant host responses involving nitric oxide (NO). In Giardia, NO is detoxified by a flavohemoglobin (gFlHb), a heme-containing enzyme which is absent in mammals. gFlHb has NO dioxygenase and NADH oxidase activities converting NO into nitrate and producing a superoxide anion (O₂^•−) that causes oxidative stress and parasite death. The modulation of gFlHb activities may provide novel approaches for treatment of giardiasis. We investigated the capacity of selected benzimidazole-2-carbamates (BZCs: ABZ, oxibendazole, nocodazole), non-BZCs (thiabendazole), an ehtylphenylcarbamate (LQM-996) and 5-NHs (MTZ, NTZ, FZD and some derivatives) to bind to recombinant gFlHb at the heme group, modifying NADH consumption activity and/or inducing ROS production. Of these, BZCs and NTZ bind to heme and increased O₂^•− production (i.e. caused enzyme subversion), whereas MTZ binds to heme but inhibited NADH consumption. LQM-996 decreased NADH consumption and two out of four NTZ derivatives altered NADH oxidase activity. In silico docking and molecular dynamics studies suggested the interaction of distinct drug moieties in ABZ and NTZ with gFlHb sites involved in NADH and NO catalysis. These findings provide new insights on gFlHb as a novel target of BZCs, MTZ and NTZ, and provides a useful platform to assess the compounds binding capacity to gFlHb prior to experimental and clinical trials in giardiasis.