Exploring an animal model of amodiaquine-induced liver injury in rats and mice

Abstract

Amodiaquine (AQ) is associated with a relatively high incidence of idiosyncratic drug-induced liver injury (IDILI) and agranulocytosis. A previous study reported that a combination of high dose AQ and glutathione (GSH) depletion led to liver injury. However, the characteristics of this toxicity were very different from AQ-induced liver injury in humans. We developed a model of AQ-induced liver injury with characteristics similar to the injury in humans by treating mice with lower doses of AQ for several weeks. In this study we found that not only did GSH depletion not increase AQ covalent binding to hepatic proteins at this lower dose, but also it paradoxically prevented the liver injury. We extended the model to rats and found AQ treatment led to a mild delayed onset liver injury that resolved despite continued treatment with AQ. Immunohistochemistry indicated the presence of Kupffer cell activation, apoptosis and hepatocyte proliferation in the liver. There was also an increase in serum IL-2, IL-5, IL-9, IL-12, MCP-1 and TGFβ, but a decrease in leptin. Coincident with the elevated serum ALT, the number of liver CD4(+) T-cells, IL-17 secreting cells and TH17/Treg cells increased at Week 3 and decreased during continued treatment. Increases in NK1.1+ cells and activated M2 macrophages were also observed during liver injury. These results suggest that the outcome of the liver injury was determined by the balance between effector and regulatory cells. Co-treatment with cyclosporin prevented AQ-induced liver injury, which supports an immune mechanism. Retinoic acid (RA), which has been reported to enhance natural killer (NK) cell activity, exacerbated AQ-induced liver injury. These results suggest that AQ-induced IDILI is immune mediated and the subsequent adaptation appears to represent immune tolerance.

The Emerging Role of the Innate Immune Response in Idiosyncratic Drug Reactions

“…In one case, BSO (700 mg/kg intraperitoneally) was administered 1 hour before amodiaquine (180 mg/kg orally), and liver injury was greatly exacerbated in 6-48 hours compared with amodiaquine treatment alone (Shimizu et al, 2009). In contrast, BSO (4.4 g/l in drinking water), administered 1 week before amodiaquine (;200 mg/kg per day in rodent meal), in addition to diethyl maleate (4 mmol/kg, intraperitoneally), administered 1 day before amodiaquine, prevented liver injury (Liu et al, 2016). It is important to note that the liver injury occurred acutely in the former model, which was likely due to the higher exposure of the mice to amodiaquine by bolus administration.…”

“…Several groups have investigated the impact of amodiaquine on hepatic structure and function. In general, studies characterizing the effects of amodiaquine monotherapy in the absence of a pre-existing disease have consistently demonstrated elevated ALT levels in the first few weeks of treatment, which then resolves (Clarke et al, 1990;Shimizu et al, 2009;Uetrecht, 2015a, 2019;Metushi et al, 2015;Liu et al, 2016).…”

“…Few studies have sought to characterize changes in inflammatory mediators with amodiaquine treatment. Amodiaquine monotherapy in female mice and male rats caused significant increases in numerous proinflammatory cytokines and chemokines beginning after 1 week of treatment (Metushi et al, 2015;Liu et al, 2016). Interestingly, the addition of amodiaquine was reported to attenuate increases in some inflammatory cytokines in models of acute tissue injury such as hepatitis or intracerebral hemorrhage (Yokoyama et al, 2007;Kinoshita et al, 2019).…”

“…Although the patterns observed are organ-specific with respect to timing and specific cell types, studies that investigated the effect of amodiaquine treatment on immune cells have consistently reported a decrease in leukocytes in the first several days to weeks of treatment, followed by an increase around a month of treatment (Clarke et al, 1990;Ajani et al, 2008;Uetrecht, 2015a, 2019;Metushi et al, 2015;Liu et al, 2016). In studies that undertook phenotyping of specific populations, NK cells were demonstrated to be the most important effector cell in response to amodiaquine treatment, with increased populations observed in the lymph node, spleen, and liver beginning after 1 week of treatment (Metushi et al, 2015;Liu et al, 2016;Mak and Uetrecht, 2019).…”

In-Vitro Approaches to Predict and Study T-Cell Mediated Hypersensitivity to Drugs

“…Clinical utility of amodiaquine has been somewhat vitiated by its capacity to elicit idiosyncratic adverse drug reactions; particularly hepatotoxicity and agranulocytosis (138,139). Amodiaquine sports an aminophenol structural alert which undergoes enzyme-mediated oxidation to form a reactive quinoneimine species which covalently binds proteins and elicits immunological responses (129,(140)(141)(142)(143)(144)(145). To circumvent this deleterious bioactivation, several routes of structural redesign were pursued (Figure 7, bottom); including the addition of two electron accepting groups at 3′ and 5′ positions to enhance potency (146) and isomerization of the 4′-hydroxyl group with the 3′-diethylamino side chain or fluorination of the 4′position to prevent quinonoid bioactivation (147,148).…”