CYP2C9新变异体A149V的发现及药物代谢活性研究  

Identification of a New CYP2C9 Variant A149V and Assessment of Its Drug Metabolic Activity

作　　者：周姗 赵桉煦 赵芳玲 洪芸 陈浩[2] 刘健 罗庆锋[3] 杨杰孚[2] 蔡剑平[1] 戴大鹏[1] ZHOU Shan;ZHAO Anxu;ZHAO Fangling(The Key Laboratory of Geriatrics,Beijing Institute of Geriatrics,Beijing Hospital,National Center of Gerontology,National Health Commission,Institute of Geriatric Medicine,Chinese Academy of Medical Sciences,Beijing 100730,China)

机构地区：[1]北京医院、国家老年医学中心、国家卫生健康委北京老年医学研究所、国家卫生健康委北京老年医学重点实验室、中国医学科学院老年医学研究院,100730 [2]北京医院心内科,100730 [3]北京医院消化科,100730

出　　处：《医学研究杂志》2022年第6期28-31,共4页Journal of Medical Research

基　　金：国家重点研发计划项目(2020YFC2008301);国家自然科学基金资助项目(81971323,81570307)。

摘　　要：目的研究中国汉族人群中发现的CYP2C9新变异体A149V的药物代谢活性。方法随机采集2018年北京医院心内科住院患者血样并采用磁珠法抽提基因组DNA,PCR扩增CYP2C9基因启动子区及9个外显子区用于基因分型。体外构建双表达杆状病毒载体pFastBac-OR-CYP2C9,转染DH10Bac大肠杆菌获得杆粒,使用昆虫系统表达CYP2C9及CYPOR,并进行Western blot法验证。采用差速离心法制备CYP2C9微粒体,使用3种探针药物进行体外酶活性研究。结果513例样本中检测到一种新的CYP2C9突变446C>T,可导致CYP2C9第149位氨基酸由丙氨酸被缬氨酸替换(A149V)。利用昆虫表达系统成功获得了新变异体A149V及典型CYP2C9变异体的微粒体,Western blot法检测结果显示,A149V的氨基酸替换并不会影响CYP2C9酶的表达水平。体外药物代谢活性数据显示,新变异体A149V对3种探针药物甲苯磺丁脲、双氯芬酸和氯沙坦的清除率分别是野生型的17.90%、24.73%和5.54%。结论中国汉族人群中发现的CYP2C9新变异体A149V为慢代谢类型。Objective To study the metabolic activity of a newly detected CYP2C9 variant A149V in Chinese Han population.Methods Blood samples were collected from inpatients in the Department of Cardiology and genomic DNAs were extracted with magnetic beads.The promoter region and 9 exons of CYP2C9 gene were amplified by PCR and used for the sequencing and genotyping.The baculovirus vector pFASTBac-OR-CYP2C9 was constructed in vitro and transfected into Escherichia coli DH10Bac to obtain the bacmid plasmid.CYP2C9 and CYPOR were then expressed with insect cell expression system and verified by Western blot.The CYP2C9 microsomes were prepared by differential centrifugation and their metabolic activities were investigated with three probe drugs in vitro.Results A new CYP2C9 mutation 446C>T was detected in 513 individuals,and this genetic mutation can cause the amino acid substitution of alanine with valine at site 149(named A149V).Microsomes of A149V and typical CYP2C9 variants were successfully obtained by the insect expression system.Western blot results showed that amino acid substitution of A149V did not affect the expression level of CYP2C9 enzyme.In vitro data of metabolic activity analysis indicated that the clearance rates of novel variant A149V for tolbutamide,diclofenac and losartan were only 17.90%,24.73%and 5.54%of the wild type,respectively.Conclusion Newly detected CYP2C9 variant A149V in Chinese Han population belongs to the poor metabolic mutation.

关 键 词：CYP2C9 变异体 A149V 代谢活性 

分 类 号：R394.6[医药卫生—医学遗传学]