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在人肠微粒体孵育体系中研究抗艾滋病候选药3-氰甲基-4-甲基-DCK(CMDCK)的肠道代谢转化、CYP同工酶表型、酶动力学特点以及CYP抑制剂对其代谢的影响。将CMDCK(4μmol·L-1)与一组重组人源CYP同工酶(CYP1A2、2C9、2C19、2D6和3A4)孵育并在人肠微粒体中与CYP同工酶特异性化学抑制剂共孵育后,应用LC-MS法测定孵育液中原形化合物的剩余量,确定CYP3A4是主导CMDCK肠代谢的同工酶,并观察到其主要代谢途径是氧化反应,应用LC-MS筛查到多个单氧化产物。CMDCK在体外肠微粒体中的代谢消除半衰期(T1/2)为25.7min,酶动力学参数Km和Vmax分别为45.6μmol·L-1和0.33μmol·L-1·min-1。应用WellStirred模型对肠微粒体的动力学参数进行外推,预测得到其体内肠道清除率为3.3mL·min-1·kg-1,接近于人肠道的平均血流量(4.6mL·min-1·kg-1)。由此推测,CMDCK的肠道代谢可能对其口服首过效应有显著影响。应用CYP450酶特异性化学抑制剂及底物与CMDCK共同孵育均能对CMDCK的肠微粒体代谢产生抑制作用。HIV蛋白酶抑制剂利托那韦能有效抑制CMDCK的肠道代谢,提高其代谢稳定性;同时发现利托那韦对CMDCK的抑制作用与是否预孵育无关,将CMDCK与利托那韦合用可降低其肠道首过代谢。
Study on Intestinal Metabolic Transformation, CYP Isozyme Phenotype, Enzyme Kinetic Characteristics and CYP Inhibitor in Anti-AIDS Candidate 3-Cyanomethyl-4-methyl-DCK (CMDCK) in Human Enteric Microsomal Incubation System Its metabolic effects. CMDCK (4 μmol·L-1) was incubated with a panel of recombinant human CYP isozymes (CYP1A2, 2C9, 2C19, 2D6 and 3A4) and co-incubated with CYP isoenzyme specific chemical inhibitors in human gut microsomes LC-MS method was used to determine the residual amount of the prototype compound in the incubation solution. It was confirmed that CYP3A4 is the isoenzyme leading to the intestinal metabolism of CMDCK, and the main metabolic pathway was observed to be oxidation reaction. Oxidation products. The half-life time (T1 / 2) of CMDCK in vitro microsomes was 25.7 min, and the kinetic parameters Km and Vmax were 45.6 μmol·L-1 and 0.33 μmol·L-1 · min-1, respectively. The kinetic parameters of enteric microsome were extrapolated using WellStirred model. The intestinal clearance rate was estimated as 3.3 mL · min-1 · kg-1, which was close to the average blood flow of human intestine (4.6 mL · min- 1 · kg-1). From this, we speculated that the intestinal metabolism of CMDCK may have a significant impact on its oral first pass effect. Application CYP450 enzyme-specific chemical inhibitors and substrates and CMDCK incubated with CMDCK intestinal microsomal metabolism can be inhibited. The inhibitory effect of ritonavir, an HIV protease inhibitor, on the intestinal metabolism of CMDCK and its metabolic stability. At the same time, it was found that the inhibitory effect of ritonavir on CMDCK was not related to the preincubation. The combination of CMDCK with ritonavir could be reduced The first intestinal metabolism.