Amodiaquine-induced oxidative stress in a hepatocyte inflammation model

Abstract

Amodiaquine is an antimalarial drug that causes life-threatening agranulocytosis and hepatotoxicity in about 1 in 2000 patients, which is usually associated with an inflammatory response. It was found that the LC 50 (2 h) of amodiaquine towards isolated rat hepatocytes was 1 mM. The cytotoxic mechanism involved protein carbonylation as well as P450 activation to a reactive metabolite. The cytotoxicity, however, was not reactive oxygen species (ROS)-mediated, as ROS scavengers did not prevent cytotoxicity or protein carbonylation, and it was not accompanied by glutathione (GSH) oxidation or intracellular H 2O 2 formation. On the other hand, the cytotoxicity could be attributed to a quinoneimine metabolite formation which formed GSH conjugates and GSH-depleted hepatocytes were much more susceptible to amodiaquine. Furthermore, when a non-toxic H 2O 2 generating system and peroxidase was used to mimic the products formed by inflammatory immune cells, only 15 μM amodiaquine was required to cause 50% cell death. In the absence of amodiaquine, hepatocyte viability and glutathione levels were not affected by the H 2O 2 generating system with or without peroxidase. The toxicity mechanism of amodiaquine in this hepatocyte H 2O 2/peroxidase model involved oxidative stress, as cytotoxicity was accompanied by GSH oxidation, decreased mitochondrial membrane potential and protein carbonyl formation which were inhibited by ROS scavengers, 4-hydroxy-2,2,6,6-tetramethylpiperidene-1-oxyl (TEMPOL) or mannitol suggesting a role for a semiquinoneimine radical and ROS in the amodiaquine-H 2O 2-mediated cytotoxic mechanism.