目的 研究新型他克林衍生物ST09、ST10的体外代谢稳定性并分别考察了其对人肝微粒CYP450各亚型酶的抑制作用。方法 采用人肝微粒体模型分别孵育ST09、ST10,液相色谱-质谱联用法测定不同孵育时间体系中剩余底物浓度,通过底物消除法计算其消除半衰期。利用LC-MS/MS测定6种CYP450主要代谢产物生成量,计算得到 IC50,分别评价ST09、ST10对主要人CYP450酶的抑制活性。结果 ST09在0.1 mg·mL-1人肝微粒体中消除半衰期小于1 min;其主要代谢产物ST10为NADPH非依赖性代谢,体外消除半衰期为32 min,体外固有清除率为0.043 mL·min-1·mg(protein)-1。ST09与ST10抑制人肝微粒体CYP3A4(咪达唑仑为底物)、CYP3A4(睾酮为底物)、CYP1A2、CYP2C9、CYP2C19、CYP2D6的IC50 分别为0.42/0.25、1.27/0.81、24.92/18.21、36.53/54.34、67.64/144.9、6.43/5.30 μmol·L-1。结论 ST09与ST10在人肝微粒体系统代谢消除较快,两者均对人肝微粒体中CYP3A4及CYP2D6有显著抑制作用。
Abstract
OBJECTIVE To determin the in vitro metabolic stability of ST09 and ST10 in human liver microsomes (HLMs), and to evaluate their potential inhibitions on five HLM cytochrome P450 isoforms.METHODS A liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to assess remaining concentration of ST09 and ST10 at designed time points during the HLM incubation. Six major metabolites of cytochrome P450 were simultaneously measured with LC-MS/MS, and the inhibitory effects of ST09 and ST10 were respectively evaluated with IC50 values.RESULTS ST09 was extremely unstable in vitro, and t1/2 was less than 1 min. However, ST10, the major metabolite of ST09, was NADPH-independent metabolized in HLMs, while its t1/2 and microsomal intrinsic clearance (CLint) were 32 min and 0.043 mL·min-1·mg(protein)-1, respectively. IC50 values of ST09 and ST10 on CYP3A4 (midazolam as substrate), CYP3A4 (testosterone as substrate), CYP1A2, CYP2C9, CYP2C19 and CYP2D6 were 0.42/0.25, 1.27/0.81, 24.92/18.21, 36.53/54.34, 67.64/144.90, 6.43/5.30 μmol·L-1, respectively.CONCLUSION ST09 and ST10 are extensively metabolized in vitro and both compounds had significant inhibition on CYP3A4 and CYP2D6.
关键词
ST09 /
ST10 /
代谢稳定性 /
酶抑制 /
液质联用
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Key words
ST09 /
ST10 /
metabolic stability /
enzyme inhibition /
LC-MS/MS
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中图分类号：
R969.1
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参考文献
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脚注
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基金
国家新药创制重大专项资助项目(2012ZX09103-101-045)
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