Lamba Jatinder K, Crews Kristine, Pounds Stanley, Schuetz Erin G, Gresham Jessica, Gandhi Varsha, Plunkett William, Rubnitz Jeffrey, Ribeiro Raul
Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, 332 North Lauderdale St., Memphis, TN 38105, USA.
J Pharmacol Exp Ther. 2007 Dec;323(3):935-45. doi: 10.1124/jpet.107.128595. Epub 2007 Sep 12.
Deoxycytidine kinase (DCK) is a rate-limiting enzyme in the activation of nucleoside analogs such as cytarabine (ara-C), gemcitabine, clofarabine, and others. The present study was undertaken to identify and to determine the functional consequences of genetic variants in DCK. We sequenced 1.5 kilobases of the DCK proximal promoter and all seven coding exons in International HapMap Project panels (n = 90 each) with European (Centre d' Etude du Polymorphisme Humain; CEPH) or African (Yoruba people in Ibadan, Nigeria; YRI) ancestry. Sixty-four genetic polymorphisms, including three nonsynonymous coding changes (I24V, A119G, and P122S) were identified. Compared with DCK-wild-type (WT) protein, the activity of the recombinant DCK24Val, DCK119Gly, and DCK122Ser proteins was 85 +/- 5, 66 +/- 3, and 43 +/- 4%, respectively. DCK119Gly and DCK122Ser mutants had lower Km (p < 0.01) and Vmax (p < 0.001) compared with DCK-WT protein. Lymphoblast cell lines from subjects heterozygous for the coding changes had significantly lower DCK activity compared with homozygous WT subjects. Ethnic differences were observed, with African ancestry subjects demonstrating significantly higher DCK mRNA expression compared with subjects with European ancestry. In both CEPH and YRI subjects, the C allele of a 3'-untranslated region single-nucleotide polymorphism (SNP) (35708 C>T) was significantly associated with lower DCK mRNA expression. This SNP was strongly linked with other intronic SNPs, forming a major haplotype block in both ethnic groups. In an exploratory analysis, the 35708C allele was also associated with lower blast ara-C-5'-triphosphate (ara-CTP) levels in acute myeloid leukemia patients receiving ara-C as continuous infusion. These results suggest that genetic variation in DCK influences its activity and expression and may predict the variability observed in intracellular levels of the ara-C active metabolite ara-CTP.
脱氧胞苷激酶（DCK）是阿糖胞苷（ara-C）、吉西他滨、氯法拉滨等核苷类似物激活过程中的限速酶。本研究旨在鉴定并确定DCK基因变异的功能后果。我们对国际人类基因组单体型图计划中欧洲（法国人类多态性研究中心；CEPH）或非洲（尼日利亚伊巴丹的约鲁巴人；YRI）血统的样本（每组n = 90）的DCK近端启动子的1.5千碱基以及所有7个编码外显子进行了测序。共鉴定出64个基因多态性，包括3个非同义编码变化（I24V、A119G和P122S）。与DCK野生型（WT）蛋白相比，重组DCK24Val、DCK119Gly和DCK122Ser蛋白的活性分别为85±5%、66±3%和43±4%。与DCK-WT蛋白相比，DCK119Gly和DCK122Ser突变体的Km（p < 0.01）和Vmax（p < 0.001）较低。编码变化杂合子受试者的淋巴母细胞系与纯合WT受试者相比，DCK活性显著降低。观察到种族差异，非洲血统受试者的DCK mRNA表达明显高于欧洲血统受试者。在CEPH和YRI受试者中，3'非翻译区单核苷酸多态性（SNP）（35708 C>T）的C等位基因与较低的DCK mRNA表达显著相关。该SNP与其他内含子SNP紧密连锁，在两个种族群体中形成一个主要的单倍型块。在一项探索性分析中，35708C等位基因还与接受持续静脉滴注阿糖胞苷的急性髓系白血病患者较低的阿糖胞苷-5'-三磷酸（ara-CTP）水平相关。这些结果表明，DCK的基因变异会影响其活性和表达，并可能预测阿糖胞苷活性代谢物ara-CTP细胞内水平的变异性。
