von Moltke L L, Greenblatt D J, Grassi J M, Granda B W, Venkatakrishnan K, Schmider J, Harmatz J S, Shader R I
Department of Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, New England Medical Center Hospital, Boston, MA 02111, USA.
J Pharm Pharmacol. 1998 Sep;50(9):997-1004. doi: 10.1111/j.2042-7158.1998.tb06914.x.
Cytochromes mediating the biotransformation of dextromethorphan to dextrorphan and 3-methoxymorphinan, its principal metabolites in man, have been studied by use of liver microsomes and microsomes containing individual cytochromes expressed by cDNA-transfected human lymphoblastoid cells. In-vitro formation of dextrorphan from dextromethorphan by liver microsomes was mediated principally by a high-affinity enzyme (Km (substrate concentration producing maximum reaction velocity) 3-13 microM). Formation of dextrorphan from 25 microM dextromethorphan was strongly inhibited by quinidine (IC50 (concentration resulting in 50% inhibition) = 0.37 microM); inhibition by sulphaphenazole was approximately 18% and omeprazole and ketoconazole had minimal effect. Dextrorphan was formed from dextromethorphan by microsomes from cDNA-transfected lymphoblastoid cells expressing CYP2C9, -2C19, and -2D6 but not by those expressing CYP1A2, -2E1 or -3A4. Despite the low in-vivo abundance of CYP2D6, this cytochrome was identified as the dominant enzyme mediating dextrorphan formation at substrate concentrations below 10 microM. Formation of 3-methoxy-morphinan from dextromethorphan in liver microsomes proceeded with a mean Km of 259 microM. For formation of 3-methoxymorphinan from 25 microM dextromethorphan the IC50 for ketoconazole was 1.15 microM; sulphaphenazole, omeprazole and quinidine had little effect. 3-Methoxymorphinan was formed by microsomes from cDNA-transfected lymphoblastoid cells expressing CYP2C9, -2C19, -2D6, and -3A4, but not by those expressing CYP1A2 or -2E1. CYP2C19 had the highest affinity (Km = 49 microM) whereas CYP3A4 had the lowest (Km = 1155 microM). Relative abundances of the four cytochromes were determined in liver microsomes by use of the relative activity factor approach. After adjustment for relative abundance, CYP3A4 was identified as the dominant enzyme mediating 3-methoxymorphinan formation from dextromethorphan, although CYP2C9 and -2C19 were estimated to contribute to 3-methoxymorphinan formation, particularly at low substrate concentrations. Although formation of dextrorphan from dextromethorphan appears to be sufficiently specific to be used as an in-vitro or in-vivo index reaction for profiling of CYP2D6 activity, the findings raise questions about the specificity of 3-methoxymorphinan formation as an index of CYP3A activity.
细胞色素介导右美沙芬在人体内转化为其主要代谢产物右啡烷和3-甲氧基吗啡喃的过程，已通过使用肝微粒体以及含有由cDNA转染的人淋巴母细胞系所表达的单个细胞色素的微粒体进行了研究。肝微粒体将右美沙芬体外转化为右啡烷主要由一种高亲和力酶介导（Km（产生最大反应速度的底物浓度）为3 - 13微摩尔）。25微摩尔右美沙芬生成右啡烷的过程受到奎尼丁的强烈抑制（IC50（导致50%抑制的浓度）= 0.37微摩尔）；磺胺苯吡唑的抑制率约为18%，奥美拉唑和酮康唑的影响极小。表达CYP2C9、-2C19和-2D6的cDNA转染淋巴母细胞系的微粒体能将右美沙芬转化为右啡烷，而表达CYP1A2、-2E1或-3A4的微粒体则不能。尽管CYP2D6在体内丰度较低，但在底物浓度低于10微摩尔时，该细胞色素被确定为介导右啡烷形成的主要酶。肝微粒体中右美沙芬生成3-甲氧基吗啡喃的平均Km为259微摩尔。对于25微摩尔右美沙芬生成3-甲氧基吗啡喃，酮康唑的IC50为1.15微摩尔；磺胺苯吡唑、奥美拉唑和奎尼丁的影响较小。表达CYP2C9、-2C19、-2D6和-3A4的cDNA转染淋巴母细胞系的微粒体能生成3-甲氧基吗啡喃，而表达CYP1A2或-2E1的微粒体则不能生成。CYP2C19具有最高亲和力（Km = 49微摩尔），而CYP3A4具有最低亲和力（Km = 1155微摩尔）。通过相对活性因子法测定了肝微粒体中这四种细胞色素的相对丰度。在对相对丰度进行调整后，CYP3A4被确定为介导右美沙芬生成3-甲氧基吗啡喃的主要酶，尽管据估计CYP2C9和-2C19也参与3-甲氧基吗啡喃的生成，尤其是在低底物浓度时。尽管右美沙芬生成右啡烷的过程似乎具有足够的特异性，可作为体外或体内评估CYP2D6活性的指标反应，但这些发现引发了关于3-甲氧基吗啡喃生成作为CYP3A活性指标的特异性的疑问。
