A phase 1 multiple-ascending dose study of tirzepatide in Japanese participants with type 2 diabetes.

AIM: To investigate the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of tirzepatide in Japanese participants with type 2 diabetes (T2D). METHODS: This phase 1, double-blind, placebo-controlled, parallel-dose, multiple-ascending dose study randomized participants to once-weekly subcutaneous tirzepatide or placebo. The tirzepatide treatment groups were: 5 mg (5 mg, weeks 1-8), 10 mg (2.5 mg, weeks 1-2; 5 mg, weeks 3-4; 10 mg, weeks 5-8), and 15 mg (5 mg, weeks 1-2; 10 mg, weeks 3-6; 15 mg, weeks 7-8). The primary outcome was tirzepatide safety and tolerability. RESULTS: Forty-eight participants were randomized. The most frequently reported treatment-emergent adverse events (AEs) were decreased appetite and gastrointestinal AEs, which were generally dose-dependent and mild in severity. The plasma tirzepatide concentration half-life was approximately 5 days. After 8 weeks of treatment, fasting plasma glucose decreased from baseline with tirzepatide versus placebo; the least squares (LS) mean decrease compared with placebo (95% confidence interval [CI]) was 52.7 (35.9-69.6), 69.1 (52.3-85.9), and 68.9 (53.2-84.6) mg/dL in the 5-, 10-, and 15-mg treatment groups, respectively (P < .0001 for all treatment groups). Tirzepatide also resulted in LS mean decreases from baseline versus placebo at 8 weeks in HbA1c up to 1.6% (95% CI 1.2%-1.9%; P < .0001 for all treatment groups) and body weight up to 6.6 kg (95% CI 5.3-7.9; P < .0001 for all treatment groups). CONCLUSIONS: All tirzepatide doses were well tolerated. The safety, tolerability, PK, and PD profiles of tirzepatide support further evaluation of once-weekly dosing in Japanese people with T2D.

Effects of cytokines on CYP3A4 expression and reversal of the effects by anti-cytokine agents in the three-dimensionally cultured human hepatoma cell line FLC-4.

The expression of hepatic cytochrome P450 (CYP) enzymes is altered under pathological conditions with increased levels of cytokines. In this study, we analyzed the effects of cytokines (interleukin [IL]-1ß, IL-6 and tumor necrosis factor α) on the expression of CYP3A4 using newly introduced three-dimensionally cultured human hepatocarcinoma FLC-4 cells. The mRNA level of CYP3A4 was significantly decreased by IL-1ß, IL-6 and tumor necrosis factor-α. Formation of α-hydroxytriazolam catalyzed by CYP3A was decreased by IL-1ß and IL-6. Pre-treatment with IL-6 enhanced the cytotoxic effects of gefitinib and paclitaxel. In addition, tocilizumab and IL-1 receptor antagonist restored the decreased expression of CYP3A4 mRNA by IL-6 and IL-1ß, respectively. These results obtained by using three-dimensionally cultured FLC-4 cells are consistent with results obtained by using primary human hepatocytes and results of clinical studies. Therefore, three-dimensionally cultured FLC-4 cell system may be a promising cellular tool to assess the effects of cytokines on CYP3A4 expression.

Cytotoxic effects of benzbromarone and its 1'-hydroxy metabolite in human hepatocarcinoma FLC4 cells cultured on micro-space cell culture plates.

Treatment with benzbromarone (BBR), a potent uricosuric drug, can be associated with liver injury. Recently, we reported that culture of human hepatocellular carcinoma FLC-4 cells on micro-space cell culture plates could increase the functional expression of drug-metabolizing enzymes including CYP3A4 and CYP2C9, which are involved in 1'-hydroxylation and 6-hydroxylation of BBR, respectively. Therefore, we examined whether BBR and its two metabolites (1'-hydroxy BBR and 6-hydroxy BBR) have cytotoxic effects in FLC4 cells cultured on micro-space cell culture plates. The present study showed that BBR and 1'-hydroxy BBR, but not 6-hydroxy BBR, have cytotoxic effects in cells cultured on micro-space cell culture plates. BBR-induced cytotoxicity was decreased by CYP3A inhibitors (itraconazole and ketoconazole), an Nrf2 activator (tert-butylhydroquinone) and a GSH precursor (N-acetyl-L-cystein). In contrast, BBR-induced cytotoxicity was increased by a GSH biosynthesis inhibitor (buthionine sulfoximine) and an inhibitor of NAD(P)H quinone oxidoreductase 1 (dicoumarol). These results suggested that metabolic activation of 1'-hydroxy BBR via CYP3A, formation of quinone metabolites and the decrease in GSH levels were involved in the BBR-induced cytotoxicity observed in FLC4 cells cultured on micro-space cell culture plates.

Increased expression of drug-metabolizing enzymes in human hepatocarcinoma FLC-4 cells cultured on micro-space cell culture plates.

Human hepatocellular carcinoma cell lines cultured in a monolayer show negligible activities of drug-metabolizing enzymes such as cytochrome P450s (CYPs) and UDP-glucuronosyltransferases (UGTs). Here, we show that culture of human hepatocellular carcinoma FLC-4 cells on 24-well plates arrayed with uniform micro-sized compartments on the bottom of the plates (micro-space cell culture plates) resulted in increased expression of drug-metabolizing enzymes (CYP1A2, CYP2C9, CYP3A4, UGT1A1, etc.) and nuclear receptors (pregnane X receptor, constitutive androstane receptor, etc.). When cells were treated with a typical CYP3A substrate (triazolam), CYP2C9 substrate (diclofenac) or UGT1A1 substrate (SN-38), large amounts of their metabolites were detected in the medium of cells cultured on micro-space cell culture plates. The formation of metabolites from triazolam, diclofenac and SN-38 was strongly inhibited by co-treatment with a CYP3A inhibitor (ketoconazole), CYP2C9 inhibitor (sulfaphenazole) and UGT1A1 inhibitor (ketoconazole), respectively. On the other hand, formation of metabolites was not observed in the medium of cells cultured in a monolayer. Finally, the cytotoxic effect of aflatoxin B1 was more potent in cells cultured on micro-space cell culture plates than in cells cultured in a monolayer. The results suggest that FLC-4 cells cultured on micro-space cell culture plates are useful for studying drug metabolism and drug-induced hepatotoxicity.