Apparent mechanism-based inhibition of human CYP2D6 in vitro by paroxetine: Comparison with fluoxetine and quinidine
Apparent mechanism-based inhibition of human CYP2D6 in vitro by paroxetine: Comparison with fluoxetine and quinidine
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DOI：
10.1124/dmd.31.3.289
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发表时间：
2003-03-01
影响因子：
3.9
通讯作者：
Greenblatt, DJ
中科院分区：
文献类型：
--
作者：
Bertelsen, KM;Venkatakrishnan, K;Greenblatt, DJ
帕罗西汀是一种选择性血清素再摄取抑制剂，是细胞色素 P450 2D6 (CYP2D6) 活性的有效抑制剂，但抑制机制尚未确定。为了确定预孵育是否影响帕罗西汀对人肝微粒体右美沙芬去甲基化活性的抑制，我们使用了两步孵育方案，其中所有酶测定组分（减去底物）均与帕罗西汀一起预孵育。还通过改变预孵育时间以及抑制剂浓度来估计抑制的动力学参数。根据这些数据，构建了 Kitz-Wilson 图，从而可以估计半最大灭活 (K-I) 所需的表观灭活剂浓度和此相互作用的最大灭活速率常数 (k(INACT)) 值。帕罗西汀与人肝微粒体预孵育导致 IC50 值降低约 8 倍（0.34 与 2.54 muM）。时间依赖性抑制被证明具有 4.85 muM 的表观 K-I 和 0.17 min(-1) 的表观 k(INACT) 值。用帕罗西汀对 CYP2D6 进行光谱扫描，456 nm 处的吸光度增加，表明帕罗西汀通过形成代谢物中间复合物使 CYP2D6 失活。该模式与帕罗西汀中亚甲二氧基取代基的代谢一致；此类取代基可能会产生基于机制的细胞色素 P450 酶失活。相比之下，奎尼丁和氟西汀（两者都是 CYP2D6 活性的抑制剂）并未表现出预孵育依赖性的抑制效力增加。这些数据与帕罗西汀（而非奎尼丁或氟西汀）对 CYP2D6 的基于机制的抑制一致。
Paroxetine, a selective serotonin reuptake inhibitor, is a potent inhibitor of cytochrome P450 2D6 (CYP2D6) activity, but the mechanism of inhibition is not established. To determine whether preincubation affects the inhibition of human liver microsomal dextromethorphan demethylation activity by paroxetine, we used a two-step incubation scheme in which all of the enzyme assay components, minus substrate, are preincubated with paroxetine. The kinetic parameters of inhibition were also estimated by varying the time of preincubation as well as the concentration of inhibitor. From these data, a Kitz-Wilson plot was constructed, allowing the estimation of both an apparent inactivator concentration required for half-maximal inactivation (K-I) and the maximal rate constant of inactivation (k(INACT)) value for this interaction. Preincubation of paroxetine with human liver microsomes caused an approximately 8-fold reduction in the IC50 value (0.34 versus 2.54 muM). Time-dependent inhibition was demonstrated with an apparent K-I of 4.85 muM and an apparent k(INACT) value of 0.17 min(-1). Spectral scanning of CYP2D6 with paroxetine yielded an increase in absorbance at 456 nm suggesting paroxetine inactivation of CYP2D6 via the formation of a metabolite intermediate complex. This pattern is consistent with the metabolism of the methylenedioxy substituent in paroxetine; such substituents may produce mechanism-based inactivation of cytochrome P450 enzymes. In contrast, quinidine and fluoxetine, both of which are inhibitors of CYP2D6 activity, did not exhibit a preincubation-dependent increase in inhibitory potency. These data are consistent with mechanism-based inhibition of CYP2D6 by paroxetine but not by quinidine or fluoxetine.