Abstract
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.
Footnotes
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This work is supported in part by the National Institutes of Health/National Institute of General Medical Sciences Pharmacogenetics Research Network and Database (U01GM61374; http://www.pharmgkb.org) under Grant U01 GM61393; in part by the Cancer Center Support Grant CA-21765; and by the American Lebanese Syrian Associated Charities. J.G. was supported in part by Grant 5 R25 CA23944 from the National Cancer Institute.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.107.128595.
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ABBREVIATIONS: ara-C, 1-β-d-arabinofuranosyl-cytosine (cytarabine); AML, acute myeloid leukemia; ara-CTP, ara-C-5′-triphosphate; DCK, deoxycytidine kinase; hENT1, human equilibrative nucleoside transporter; LD, linage disequilibrium; mt, mutant; SNP, single nucleotide polymorphism; HapMap, Haplotype Map (of human genome); CdA, cladribine; PCR, polymerase chain reaction; CEPH, Centre d' Etude du Polymorphisme Humain; YRI, Yoruba people in Ibadan, Nigeria; UTR, untranslated region; WT, wild-type; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; CT, cycle threshold; ERS, Glu-Arg-Ser; SR, serine/arginine; IVS, intervening sequence (intron).
- Received July 22, 2007.
- Accepted September 6, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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