a novel mutation was found and we want to confirm the sequence of it and explore the funtional change due to it.. A NOVEL MUTATION IN CYP2C9

During a retrospective study, we analyzed the cytochrome P450 2C9 (CYP2C9) and vitamin K epoxide reductase complex subunit 1 genes among 120 patients taking warfarin. A new coding variant was identified by sequencing CYP2C9. a patient taking high-dose warfarin in order to achieve the target international normalized ratio was found to have a mutation in the CYP2C9 gene. We confirmed the novel mutation by resequencing and performed the functional experiment of the variant in vitro. cDNA of the novel variant was constructed by site-directed mutagenesis and the recombinant protein was expressed in vitro using a baculovirus–expression system. Its enzymatic activities toward probe drugs for CYP2C9 were then assessed.

本研究为一项回顾性研究，我们对120名服用华法林（warfarin）的患者的细胞色素P450 2C9（cytochrome P450 2C9，CYP2C9）及维生素K环氧还原酶复合物亚基1基因（vitamin K epoxide reductase complex subunit 1）进行了分析。通过对CYP2C9基因进行测序，我们发现了一种新的编码变异。一名为达到目标国际标准化比值（international normalized ratio）而服用高剂量华法林的患者，被检出携带CYP2C9基因的突变。我们通过重测序验证了该新型突变，并针对该变异开展了体外功能实验：通过定点诱变（site-directed mutagenesis）技术构建该新型变异的cDNA，利用杆状病毒表达系统（baculovirus-expression system）在体外表达重组蛋白，随后检测其针对CYP2C9探针药物的酶活性。