von Ahsen Nicolas, Oellerich Michael, Armstrong Victor W
Department of Clinical Chemistry, University of Göttingen, Germany.
Ther Drug Monit. 2008 Feb;30(1):16-22. doi: 10.1097/FTD.0b013e318161a21a.
Inosine triphosphatase (ITPA) cleaves phosphate residues from inosine triphosphate (ITP) and deoxy ITP (dITP), thereby recovering inosine monophosphate, which is a substrate for further purine nucleotide pathways. Deficient ITPA activity leads to intracellular accumulation of ITP/dITP and would, under thiopurine therapy, lead to accumulation of unusual thio-inosine metabolites (thio-ITP) with the potential for adverse metabolic effects. ITPA is a promising candidate for a more comprehensive understanding of thiopurine pharmacogenetics. We therefore studied the haplotype structure, haplotype-phenotype association, and promoter function of ITPA in a Western European population.ITPA haplotyping was performed based on haplotype tagging SNPs (selected from HapMap data) in healthy 130 controls. Haplotypes were reconstructed, and the haplotype-phenotype association was assessed by haplotype trend regression. A 1.5 kb upstream region and stepwise deletions thereof were tested for promoter activity in reporter gene assays in HepG2 and CCRF-CEM cells. Transcription factor binding (Sp1, Sp3) to the proximal promoter region was studied by chromatin immunoprecipitation. Among haplotypes with a frequency greater than 0.01, we did not find any new low-activity haplotypes besides those carrying 94C>A or IVS2 + 21A>C variants. Two promoter SNPs had no influence on promoter activity. An approximately 200 bp sized GC-rich proximal promoter region was sufficient to fully drive reporter gene activity. Chromatin immunoprecipitation showed binding of Sp1 and Sp3 transcription factors to this region. Only the two haplotypes carrying 94C>A or IVS2 + 21A>C were associated with reduced enzyme activity. The gene promoter is associated with a CpG island and driven by Sp-family transcription factors. There was no evidence for functional promoter SNPs, and it is suggested that only SNPs within the very proximal promoter region (approximately 200 bp) have the potential to be functionally significant.
肌苷三磷酸酶（ITPA）可从肌苷三磷酸（ITP）和脱氧肌苷三磷酸（dITP）上切割磷酸残基，从而生成肌苷单磷酸，后者是进一步嘌呤核苷酸代谢途径的底物。ITPA活性不足会导致ITP/dITP在细胞内蓄积，在硫唑嘌呤治疗时，会导致异常硫代肌苷代谢产物（硫代ITP）蓄积，可能产生不良代谢效应。ITPA是更全面了解硫唑嘌呤药物遗传学的一个很有前景的候选对象。因此，我们在一个西欧人群中研究了ITPA的单倍型结构、单倍型-表型关联及启动子功能。在130名健康对照中，基于单倍型标签单核苷酸多态性（从HapMap数据中选取）进行ITPA单倍型分型。重建单倍型，并通过单倍型趋势回归评估单倍型-表型关联。在HepG2和CCRF-CEM细胞的报告基因检测中，检测了一个1.5 kb的上游区域及其逐步缺失片段的启动子活性。通过染色质免疫沉淀研究转录因子（Sp1、Sp3）与近端启动子区域的结合。在频率大于0.01的单倍型中，除了携带94C>A或IVS2 + 21A>C变异的单倍型外，我们未发现任何新的低活性单倍型。两个启动子单核苷酸多态性对启动子活性无影响。一个约200 bp大小的富含GC的近端启动子区域足以完全驱动报告基因活性。染色质免疫沉淀显示Sp1和Sp3转录因子与该区域结合。只有携带94C>A或IVS2 + 21A>C的两个单倍型与酶活性降低有关。该基因启动子与一个CpG岛相关，并由Sp家族转录因子驱动。没有证据表明启动子单核苷酸多态性具有功能意义，提示只有近端启动子区域（约200 bp）内的单核苷酸多态性可能具有功能重要性。
