Shimada T, Tsumura F, Yamazaki H, Guengerich F P, Inoue K
Osaka Prefectural Institute of Public Health, Japan.
Pharmacogenetics. 2001 Mar;11(2):143-56. doi: 10.1097/00008571-200103000-00005.
Twenty-four genetic polymorphisms in the CYP2D6 gene were analysed in liver DNA samples of 39 Japanese and 44 Caucasians and compared with CYP2D6 protein levels and bufuralol 1'- and 6-hydroxylation activities in liver microsomes of these human samples. We detected 13 types of CYP2D6 genetic polymorphisms and classified these into 20 genotypes; nine types were found in Japanese and 14 types in Caucasian samples. CYP2D610B, but not CYP2D610A, was the most frequent (34.6%) in Japanese. In Caucasians, several CYP2D6 polymorphisms including CYP2D6*4, *4D, *4E, *4L, *3, *9, *5 and 2E (frequencies of 6.8, 3.4, 4.5, 9.1, 1.1, 2.3, 2.3 and 4.5%, respectively) were detected. A Caucasian having CYP2D63/5 had a protein with slower gel mobility (immunoblotting with anti-CYP2D6 antibody) and very low activity for bufuralol 1'-hydroxylation. Five Caucasian samples (CYP2D64/*4, *4/*4L, or *4D/4L) had no measurable CYP2D6 protein and very low bufuralol 1'-hydroxylation activities. Seven Japanese subjects with CYP2D610B/10B had CYP2D6 protein at levels of approximately 20% of those present in humans with CYP2D61 and 2 and catalysed bufuralol 1'-hydroxylation at low rates. Kinetic analysis of bufuralol 1'- and 6-hydroxylation indicates that (i) the Km values for 1'-hydroxylation were lower in individuals with CYP2D61/*1, *1/*2, *1/*2X2, and *2/2 than those with CYP2D64/*4, *4/*4L, *4D/*4L, or *10B/*10B and Vmax values tended to be higher in the former groups (*1, 2), and (ii) individuals with heterozygous CYP2D61/*4D, *1/*4L, and *1/5 had relatively high Vmax/Km ratios, whereas individuals with heterozygous CYP2D61/*9, *2/4D, *2/*5, *2/*10B, *2E/*4E, *3/*5, *4L/*9, and 10B/39 had lower Vmax/Km ratios for bufuralol 1'-hydroxylation. Quinidine inhibited bufuralol 1'-hydroxylation in liver microsomes, particularly at low substrate concentrations, in individuals with CYP2D61/1, and 1/12, but not those with CYP2D64/4 and very slightly in individuals with CYP2D610B/10B. The latter two groups were found to be more sensitive to alpha-naphthoflavone than the former groups, indicative of the contribution of CYP1A2. These results support the view that CYP2D63, *4, *4D, and 4L are major genotypes producing poor metabolizer phenotypes in CYP2D6 in Caucasians, whereas CYP2D610B is a major factor in decreased CYP2D6 protein expression and catalytic activities in Japanese.
对39名日本人及44名高加索人的肝脏DNA样本中的CYP2D6基因的24个基因多态性进行了分析，并与这些人类样本肝脏微粒体中的CYP2D6蛋白水平及丁呋洛尔1'-和6-羟基化活性进行了比较。我们检测到13种CYP2D6基因多态性，并将其分为20种基因型；在日本人中发现了9种，在高加索人样本中发现了14种。CYP2D610B在日本人中最为常见（34.6%），而CYP2D610A并非如此。在高加索人中，检测到了几种CYP2D6多态性，包括CYP2D6*4、*4D、*4E、4L、3、9、5和2E（频率分别为6.8%、3.4%、4.5%、9.1%、1.1%、2.3%、2.3%和4.5%）。一名具有CYP2D63/5的高加索人其蛋白质具有较慢的凝胶迁移率（用抗CYP2D6抗体进行免疫印迹），且丁呋洛尔1'-羟基化活性非常低。五个高加索人样本（CYP2D64/4、4/4L或4D/4L）未检测到可测量的CYP2D6蛋白，且丁呋洛尔1'-羟基化活性非常低。七名具有CYP2D610B/'10B的日本人的CYP2D6蛋白水平约为具有CYP2D61及2的人的20%，且丁呋洛尔'1'-羟基化催化速率较低。丁呋洛尔1'-和6-羟基化的动力学分析表明：（i）具有CYP2D61/*1、*1/*2、*1/2X2及2/2的个体中1'-羟基化的Km值低于具有CYP2D64/*4、*4/*4L、*4D/4L或10B/*10B的个体，且前几组（*1、2）的Vmax值倾向于更高；（ii）具有杂合子CYP2D61/*4D、*1/4L及1/5的个体具有相对较高的Vmax/Km比值，而具有杂合子CYP2D61/*9、*2/4D、*2/*5、*2/*10B、*2E/*4E、*3/*5、4L/9及10B/39的个体丁呋洛尔1'-羟基化的Vmax/Km比值较低。奎尼丁抑制具有CYP2D61/1及1/12的个体肝脏微粒体中的丁呋洛尔1'-羟基化，尤其是在低底物浓度时，但不抑制具有CYP2D64/4的个体，对具有CYP2D610B/10B的个体抑制作用非常轻微。发现后两组比前两组对α-萘黄酮更敏感，表明CYP1A2的作用。这些结果支持以下观点：CYP2D63、4、4D及4L是导致高加索人CYP2D6中代谢不良者表型的主要基因型，而CYP2D610B是导致日本人CYP2D6蛋白表达及催化活性降低的主要因素。
