Bredschneider Monika, Klein Kathrin, Mürdter Thomas E, Marx Claudia, Eichelbaum Michel, Nüssler Andreas K, Neuhaus Peter, Zanger Ulrich M, Schwab Matthias
Dr Margarete Fischer-Bosch Institute of Clinical Pharmacology, Stuttgart, Germany.
Clin Pharmacol Ther. 2002 Jun;71(6):479-87. doi: 10.1067/mcp.2002.124518.
High-dose busulfan is widely used as part of conditioning regimens for patients who are undergoing hematopoietic stem cell or bone marrow transplantation. High plasma concentrations of busulfan have been linked to the occurrence of hepatic venoocclusive disease (VOD), a severe complication associated with a high mortality. Because conjugation with glutathione, the major route of biotransformation of busulfan, is predominantly catalyzed by the isozyme glutathione S-transferase A1 (GSTA1), we hypothesized that low expression or function of GSTA1 in liver caused by genetic polymorphisms may be the mechanism underlying VOD.
Immunoblot analysis of GSTA and measurement of busulfan-glutathione conjugation by liquid chromatography-mass spectrometry were performed in 48 normal human liver samples. To search for polymorphisms, the complete GSTA1 coding regions and the promoter fragment were sequenced. All results were compared by multivariate analysis.
Absolute levels of GSTA protein and formation rates of busulfan-glutathione conjugate displayed a 7- and 8-fold range, from 240 to 1600 pmol/mg and 25 to 205 pmol/min per milligram of total cytosolic protein, respectively, and correlate (r2 = 0.49, P <.0001). A total of 8 single nucleotide polymorphisms (SNPs) of GSTA1 were identified, 1 of which was a silent mutation in exon 5 (A375G); all others were found in the promoter region. Haplotype analysis revealed the existence of 5 defined alleles. There was no significant relationship between any of the GSTA1 SNPs or haplotypes and either hepatic glutathione S-transferase A (GSTA) expression or GSTA1 function.
The identified GSTA1 polymorphisms are not likely to be related to the VOD because they do not appear to be associated with changes in GSTA expression or function. Compared with other members of the GST family, GSTA1 displays surprisingly little variation.
大剂量白消安广泛用于造血干细胞或骨髓移植患者的预处理方案。白消安的高血浆浓度与肝静脉闭塞病（VOD）的发生有关，VOD是一种严重并发症，死亡率很高。由于与谷胱甘肽结合是白消安生物转化的主要途径，主要由同工酶谷胱甘肽S-转移酶A1（GSTA1）催化，我们推测基因多态性导致肝脏中GSTA1低表达或功能异常可能是VOD的潜在机制。
对48例正常人肝脏样本进行GSTA免疫印迹分析，并通过液相色谱-质谱法测定白消安-谷胱甘肽结合情况。为寻找多态性，对GSTA1完整编码区和启动子片段进行测序。所有结果通过多变量分析进行比较。
GSTA蛋白的绝对水平和白消安-谷胱甘肽结合物的形成速率分别在240至1600 pmol/mg和25至205 pmol/min每毫克总胞质蛋白范围内变化7倍和8倍，且二者相关（r2 = 0.49，P <.0001）。共鉴定出8个GSTA1单核苷酸多态性（SNP），其中1个是外显子5的沉默突变（A375G）；其他均位于启动子区域。单倍型分析揭示了5个已定义等位基因的存在。任何GSTA1 SNP或单倍型与肝谷胱甘肽S-转移酶A（GSTA）表达或GSTA1功能之间均无显著关系。
已鉴定出的GSTA1多态性不太可能与VOD相关，因为它们似乎与GSTA表达或功能的变化无关。与GST家族的其他成员相比，GSTA1的变异惊人地少。
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