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Identification of Genetic Polymorphisms of CYP2W1 in the Three Main Chinese Ethnicities: Han, Tibetan, and Uighur

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Identification of Genetic Polymorphisms of CYP2W1 in the Three Main Chinese Ethnicities: Han, Tibetan, and Uighur Supplemental material to this article can be found at: http://dmd.aspetjournals.org/content/suppl/2016/06/15/dmd.115.069153.DC1 1521-009X/44/9/1510–1515$25.00 http://dx.doi.org/10.1124/dmd.115.069153 DRUG METABOLISM AND DISPOSITION Drug Metab Dispos 44:1510–1515, September 2016 Copyright ª 2016 by The American Society for Pharmacology and Experimental Therapeutics Identification of Genetic Polymorphisms of CYP2W1 in the Three Main Chinese Ethnicities: Han, Tibetan, and Uighur Yanwei Li,1 Xing Kang,1 Ge Yang, Penggao Dai, Chao Chen, and Huijuan Wang National Engineering Research Center for Miniaturized Detection Systems, School of Life Sciences, Northwest University, Xi’an, China Received December 21, 2015; accepted June 14, 2016 ABSTRACT CYP2W1 is an orphan member of the cytochrome P450 superfamily. data of nine exonic variants found in this study. CYP2W1*1A, *1B, *2, Recently, CYP2W1 has gained great research interest because of its *4,and*6 were detected as the main alleles/haplotypes. Moreover, unknown enzymatic function and tumor-specific expression property. one, three, and two ethnically specific haplotypes were observed in Downloaded from This study aims to investigate the genetic polymorphisms of the the Han, Tibetan, and Uighur samples, respectively. Then, the effects CYP2W1 gene in Chinese populations and explore the functions of the of four detected missense mutations (Ala181Thr, Gly376Ser, Val432Ile, detected variants. All of the nine exons and exon-intron junction and Pro488Leu) on the CYP2W1 protein function were predicted using regions of the CYP2W1 gene were sequenced in 150 Chinese three in silico tools: Polymorphism Phenotyping v2, Sorts Intolerant subjects, including 50 Han Chinese, 50 Tibetans, and 50 Uighurs. from Tolerant, and MutationTaster. The results showed that A total of 26 genetic variants were identified in this study, and Gly376Ser and Pro488Leu may have deleterious effects. In summary, dmd.aspetjournals.org 19 polymorphisms were detected in each population. Frequency this study showed that the genetic pattern of CYP2W1 is interethni- comparison between populations showed that nine variants exhibited cally different among the three Chinese populations, and this finding significantly different allelic distributions. A total of 12 different can extend our understanding of population genetics of CYP2W1 in haplotypes were inferred from 150 samples by using the genotype the Chinese population. at ASPET Journals on September 24, 2021 Introduction Guengerich, 2012; Xiao and Guengerich, 2012). Thus, human CYP2W1 CYP2W1 is an extrahepatic cytochrome P450 (P450) enzyme, and is is a potential drug target for cancer therapy using antibodies and/or specific considered as an “orphan” P450 because of its unknown physiologic CYP2W1 activated prodrugs (Nishida et al., 2010; Travica et al., 2013). functions (Karlgren and Ingelman-Sundberg, 2007). Recent studies have Many of the P450 isoforms are highly polymorphic. The genetic reported that CYP2W1 is not expressed in adult human liver or in human polymorphisms in P450 genes can alter enzymatic activity and change the primary hepatocytes. Only low amounts of CYP2W1 mRNA were found metabolic rate of the drugs or procarcinogens catalyzed by the enzymes, in human adult extrahepatic and fetal tissues (Karlgren et al., 2006). resulting in varied drug responses or disease susceptibilities among However, CYP2W1 is highly expressed in several human tumor tissues, individuals (Wijnen et al., 2007). In a study that investigated the distribution especially colon tissues. Therefore, the expression of CYP2W1 seems of the genetic polymorphisms in a Japanese population, only six alleles tumor-specific (Karlgren et al., 2006; Choong et al., 2015). Further and five nonsynonymous single-nucleotide polymorphisms (nsSNPs) studies have demonstrated that the expression of CYP2W1 is regulated in CYP2W1 were reported (http://www.cypalleles.ki.se/cyp2w1.htm) by DNA methylation, and that the CYP2W1 gene is silenced by DNA (Hanzawa et al., 2008). Of the nsSNPs identified, Pro488Leu (CYP2W1*6) methylation after development and then reactivated in tumors by was found to be more common in the Japanese population than in hypomethylation (Karlgren et al., 2006; Gomez et al., 2007; Choong Caucasians (allele frequency: 0.368 vs. 0.06). In contrast, Ala181Thr et al., 2015). The in vitro expression of CYP2W1 in a heterologous (CYP2W1*2) is prevalent in Caucasians but rare in the Japanese (allele system revealed a broad range of catalytic activities toward several frequency: 0.21 vs. 0.014) (Hanzawa et al., 2008; Gervasini et al., 2010; chemicals, including arachidonic acid, benzphetamine, and several Stenstedt et al., 2013). Therefore, the distribution of CYP2W1 genetic procarcinogens (polycyclic aromatic hydrocarbon dihydrodiols, afla- polymorphisms is interethnically different. Still, further studies with toxin B1, and sterigmatocystin) (Wu et al., 2006; Yoshioka et al., 2006; different ethnic groups are needed for a more comprehensive understand- Eun et al., 2010; Nishida et al., 2010; Tan et al., 2011; Wang and ing of the distribution of CYP2W1 genetic polymorphisms in humans. Several studies have investigated the function of CYP2W1 genetic polymorphisms, and their conclusions are inconsistent with respect to the effect of CYP2W1 genetic variants on tumor susceptibility (Gervasini This work was supported by the National Natural Science Foundation of China [Grant J1210063] and the China Postdoctoral Science Foundation [Grant et al., 2010; Cardenas-Rodriguez et al., 2012; Stenstedt et al., 2013). One 2015M582695]. study showed that CYP2W1*2 is significantly associated with increased 1Y.L. and X.K. contributed equally to this work. colorectal cancer risk in a Caucasian population (Gervasini et al., 2010); dx.doi.org/10.1124/dmd.115.069153. however, another study debunked this relationship in another sample s This article has supplemental material available at dmd.aspetjournals.org. (Stenstedt et al., 2013). By using a human colon cancer cell expression ABBREVIATIONS: LD, linkage disequilibrium; nsSNP, nonsynonymous SNP; P450, cytochrome P450; PolyPhen-2, Polymorphism Phenotyping v2; SIFT, Sorts Intolerant from Tolerant; SNP, single-nucleotide polymorphism; 39UTR, 39 untranslated region. 1510 Population Genetic Study of the CYP2W1 Gene 1511 system, the latter study also demonstrated that CYP2W1*2 and Results CYP2W1*6 do not alter the metabolic kinetics of CYP2W1 substrates CYP2W1 Genetic Polymorphisms and Distribution. A total of in vitro. Further studies are needed to characterize the function of 26 different genetic variations, including three novel ones in the CYP2W1 genetic polymorphisms and address their roles in disease CYP2W1 gene, were detected in 150 individuals (Table 1). The three susceptibility and drug metabolism. novel variants were all SNPs, located in the promoter (v01), intron7 (v21), The current study screened the genetic variants of CYP2W1 in three and 39UTR (v25), respectively. A total of nine variations were identified Chinese populations (Han, Tibetan, and Uighur) by sequencing the in the coding region, whereas the remaining variants were located in exonic sequences of the CYP2W1 gene in 150 samples. This process was the promoter (two variations), introns (13 variations), and 39UTR (two carried out to understand CYP2W1 genetic polymorphisms. The geno- variations). Four of the nine exonic SNPs were nsSNPs, located in type and haplotype distributions of CYP2W1 in Chinese populations exons 4, 7, and 9, giving rise to the amino acid exchanges Ala181Thr were investigated, and the effects of identified nsSNPs in these samples (v11), Gly376Ser (v20), Val432Ile (v23), and Pro488Leu (v24). One were predicted by using in silico tools. insertion variation (v09) in intron1 was also identified. The frequency of detected variations in specific populations was Materials and Methods calculated. First, 19 polymorphic variations were found in each Samples. Blood samples were collected from 150 unrelated healthy volunteers population (Table 2). They consist of 14 identical variations and five from three different nationalities: 50 Han Chinese from Shaanxi Province, ethnically specific variations. Second, statistical analysis showed that 50 Tibetans from Tibet, and 50 Uighurs from Xinjiang. All of the volunteers were nine variants (v3, v4, v05, v08, v9, v10, v13, v24, and v26) exhibited Downloaded from sophomores at Xizang Minzu University, aged about 19 years. The percentage of significantly different frequencies among three studied populations females in the Han, Tibetan, and Uighur samples is 52, 55, and 49%, respectively. (P , 0.05). Three variants (v04, v08, and v10) remained significantly This study was approved by the Ethics Committee of Northwest University, and different after Bonferroni correction. Third, of the four nsSNPs all participants signed an informed consent form before participating in the study. identified in this study, Gly376Ser (v20) was found to be heterozy- Genomic DNA Extraction. The genomic DNA samples were extracted from gous in one of the 50 Uighur subjects (Supplemental Table 2); m – 200 l of EDTA anticoagulated peripheral blood by using the QIAamp DNA Val432Ile (v23) presented with relatively low and comparable frequency blood mini kit (Qiagen, Shanghai, China). The extracted genomic DNA was dmd.aspetjournals.org (0.02–0.05, P = 1) among three populations; Ala181Thr (v11) exhibited dissolved in an elution buffer (10 mM Tris-HCl, 0.5 mM EDTA, pH 9.0) and stored
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