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1. The thiazolidinedione (TZD) ring present in drugs available for type II diabetes may contribute to hepatic injury. Another TZD ring-containing compound, 3-(3,5dichlorophenyl)-2,4-thiazolidinedione (DCPT), produces liver damage in rats. Accordingly, the effects of gender, dose and time on DCPT hepatotoxicity were therefore evaluated. 2. Male rats were more sensitive to DCPT (0.4-1.0 mmol/kg by i.p. administration) as shown by increased serum alanine aminotransferase (ALT) levels and altered hepatic morphology 24 h post-dosing. Effects in both genders were dose-dependent. In males, DCPT (0.6 mmol/kg) produced elevations in ALTs and changes in liver sections 3 h after dosing that progressively worsened up to 12 h. DCPT-induced renal effects were mild. 3. We conclude that male rats are more susceptible to DCPT hepatotoxicity and that damage occurs rapidly. DCPT primarily affects the liver and may be a useful compound to investigate the role of the TZD ring in hepatic injury. However, the gender dependency and rapid onset of DCPT hepatotoxicity require further investigation.

Cytochrome P450 (CYP)-mediated metabolism in the thiazolidinedione (TZD) ring may contribute to the hepatotoxicity of the insulin-sensitizing agents such as troglitazone. We were interested in determining if biotransformation could also be a factor in the liver damage associated with another TZD ring containing compound, 3-(3,5-dichlorophenyl)-2,4-thiazolidinedione (DCPT). Therefore, hepatotoxic doses of DCPT (0.6 or 1.0 mmol/kg, i.p.) were administered to male Fischer 344 rats after pretreatment with vehicle, 1-aminobenzotriazole (ABT, non-selective CYP inhibitor) and troleandomycin (TAO, CYP3A inhibitor). Alternatively, rats were pretreated with vehicle or the CYP3A inducer dexamethasone (DEX) prior to a non-toxic DCPT dose (0.2 mmol/kg, i.p.). Vehicle-, ABT-, TAO-and DEX-only control groups were also run. Toxicity was assessed 24 hours after DCPT administration. Both hepatotoxic doses of DCPT induced elevations in serum alanine aminotransferase (ALT) levels that were attenuated by ABT or TAO pretreatment. Liver sections from rats that received vehicle + DCPT revealed areas of gross necrosis and neutrophil invasion, whereas sections from ABT + DCPT and TAO + DCPT rats showed minor changes compared to controls. DEX pretreatment potentiated ALT levels associated with the non-toxic DCPT dose. Furthermore, DEX + DCPT rat liver sections exhibited hepatic injury when compared against rats that received vehicle + DCPT. Blood urea nitrogen levels, urinalysis and kidney morphology were not markedly altered by any combination of pretreatments or treatments. Enzyme activity and Western blotting experiments with rat liver microsomes confirmed the effects of the various pretreatments. Our results suggest that hepatic CYP3A isozymes may be involved in DCPT-induced liver damage in male rats. We believe this is the first report demonstrating that modulation of the biotransformation of a TZD ring-containing compound can alter hepatotoxicity in a common animal model.

Abbreviations: λz, terminal elimination constant; ADI, acceptable daily intake; AUC0-last, area under the curve from time zero until the last sampling time point; AE, adverse events; AUC, area under the plasma concentration-time curve; AUC-0.75-inf, AUC from time zero to infinity; AUClast, AUC to the last quantifiable observation; BLQ, below the limit of quantification; BMI, body mass index; bw, body weight; Cmax, concentration maximum; CSAF, chemical-specific adjustment factor; EE, efficacy evaluable population; EKG, electrocardiogram; HBsAg, hepatitis B surface antigen; IQR, interquartile range; IRB, Institutional Review Board; JECFA, Joint FAO/WHO Expert Committee on Food Additives; LC-MS/MS, liquid chromatography-tandem mass spectrometry; LLOQ, lower limit of quantification, LOQ, limit of quantification; NOAEL, noobserved-adverse-effect level; PP, per protocol population; SD, standard deviation; T1/2, half-life; Tmax, time to maximum concentration; WHO, World Health Organization * Corresponding author: Tel: +1-905-542-2900 E-mail address: ashley.roberts@intertek.com AbstractThe acceptable daily intake (ADI) of commercially available steviol glycosides is currently 4 mg/kg body weight (bw)/day, based on application of a 100-fold uncertainty factor to a noobserved-adverse-effect-level value from a chronic rat study. Within the 100-fold uncertainty factor is a 10-fold uncertainty factor to account for inter-species differences in toxicokinetics (4-fold) and toxicodynamics (2.5-fold). Single dose pharmacokinetics of stevioside were studied in rats (40 and 1,000 mg/kg bw) and in male human subjects (40 mg/kg bw) to generate a chemical-specific, inter-species toxicokinetic adjustment factor. Tmax values for steviol were at ~8 and ~20 hours after administration in rats and humans, respectively. Peak concentrations of steviol were similar in rats and humans, while steviol glucuronide concentrations were significantly higher in humans. Glucuronidation in rats was not saturated over the dose range 40-1,000 mg/kg bw. The AUC0-last for steviol was approximately 2.8-fold greater in humans compared to rats. Chemical-specific adjustment factors for extrapolating toxicokinetics from rat to human of 1 and 2.8 were established based on Cmax and AUC0-last data respectively. Because these factors are lower than the default value of 4.0, a higher ADI for steviol glycosides of between 6 to 16 mg/kg bw/d is justified.

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