GRHL3和c-Myc在食管鳞状细胞癌组织中的表达及其临床意义

目的:研究食管鳞状细胞癌(esophageal squamous cell carcinoma,ESCC)患者肿瘤组织及癌旁组织中GRHL3和c-Myc的表达情况及其临床意义.方法:收集2015年4月至2016年7月在河北医科大学第四医院胸外科行肿瘤切除并经病理证实的64例ESCC患者的肿瘤组织、癌旁组织,采用Real-time PCR法和免疫组化法检测GRHL3和c-Myc基因的mRNA和蛋白表达情况,分析其与患者临床特征的关系.结果:与癌旁组织相比,ESCC组织中GRHL3 mRNA表达水平和蛋白阳性表达水平均显著升高[(2.85±2.83) vs(2.06±2.02),P＜0.01;81.30％ vs 25.00％,P＜0.01],ESCC组织中c-Myc mRNA表达水平和蛋白阳性表达水平均显著升高[5.13±5.11)vs (2.03±2.00),P＜0.01;42.20％ vs.20.30％,P＜0.01].ESCC组织中GRHL3 mRNA的表达与c-Myc mRNA的表达呈显著正相关关系(P＜0.05),GRHL3蛋白表达和c-Myc蛋白表达也呈显著正相关(P＜0.01).GRHL3蛋白表达和c-Myc蛋白表达与患者肿瘤浸润程度、淋巴结转移、临床分期、分化程度相关.结论:ESCC患者肿瘤组织中GRHL3与c-Myc表达水平显著提高,两者表达呈正相关,且两者与患者临床病理特征密切相关,可能是影响ESCC病理进程的重要因素.     

c-Myc 和 Bin1 在人食管鳞状细胞癌组织中的表达及其临床意义

目的： 研究人食管鳞状细胞癌（esophageal squamous cell cancer, ESCC）组织及其癌旁组织中 c-Myc和 Bin1 基因的表达情况,并分析其临床意义。 方法： 收集2013年4月至2014年3月在河北医科大学第四医院胸外科行肿瘤切除并经病理证实的ESCC患者肿瘤组织和相应癌旁组织标本各54例,使用RT-PCR和免疫组织化学法检测ESCC组织和癌旁组织中c-Myc和 Bin1 基因的mRNA和蛋白的表达。 结果： 与癌旁组织相比,ESCC组织中的 c-Myc mRNA表达水平和蛋白阳性表达率均显著升高\[（0.34±0.29） vs （0.17±0.16）, P <0.001;55.56% vs 33.33%, P =0.033\], Bin1 mRNA表达水平和蛋白阳性表达率均显著降低\[（0.25±0.19） vs （0.33±0.20）, P <0.001;57.41% vs 84.48%, P =0.007\]。ESCC组织中 c-Myc mRNA的表达与 Bin1 mRNA的表达呈显著负相关关系（ r =0.790, P <0.001）,c-Myc蛋白表达与Bin1蛋白表达也呈显著负相关关系（ P =0.019）。c-Myc和 Bin1蛋白表达均与患者TNM分期、肿瘤侵犯深度、分化程度、淋巴结转移相关。 结论： 人ESCC患者肿瘤组织中c-Myc呈高表达,而Bin1呈低表达,两者表达呈负相关,均与ESCC进展和淋巴结转移有关联。     

食管鳞状细胞癌组织Bin1基因启动子甲基化状态及其临床意义

目的:探讨食管鳞状细胞癌(esophageal squamous cell cancer,ESCC)患者的食管鳞癌组织、癌旁组织中Bin1基因启动子甲基化状态及其mRNA的表达及其临床意义.方法:采用实时荧光定量PCR(qRT-PCR)分别检测58例经病理证实的ESCC患者的食管鳞癌组织、癌旁组织中Bin1基因mRNA的表达情况;用甲基化特异性PCR(MSP)检测上述食管鳞癌组织中Bin1基因启动子甲基化状态,比较ESCC患者Bin1甲基化状态与临床病理分期的关系.结果:ESCC组织中Bin1基因启动子甲基化率明显高于癌旁组织(58.62％ vs 25.86％,x2=12.76,P＜0.01),Bin1甲基化状态与患者TNM分期、肿瘤侵润深度、分化程度、淋巴结转移相关(均P ＜0.05).ESCC组织中Bin1 mRNA的表达水平明显低于癌旁组织[(0.78 ±0.05) vs (1.03±0.03),t=9.643,P＜0.01)];发生Bin1甲基化的组织中Bin1 mRNA表达水平明显低于未发生甲基化的组织[(0.68±0.04) vs (0.85±0.07),t=2.476,P＜0.05].结论:Bin1基因启动子区甲基化状态可能与ESCC的发生密切相关,它是ESCC中Bin1 mRNA低表达或缺失的机制之一,且与ESCC进展和淋巴结转移有关.     

SATB1基因在结直肠癌中的表达及其临床意义

目的 检测结直肠癌组织中特异AT序列结合蛋白1(special AT-rich sequence binding protein,SATB1)的表达情况与结直肠癌生物学行为的相关性.方法 采用反转录多聚酶链式反应(RT-PCR)法检测39例结直肠癌及相应癌旁对照组织中SATB1 mRNA的表达,采用免疫组织化学方法检测39例结直肠癌及相应癌旁对照组织中SATB1的表达,分析SATB1基因表达与患者临床病理因素的相关性.结果 癌组织中SATB1蛋白表达阳性率为69.23％(27/39),SATB1 mRNA表达量为(1.178±0 644),均高于癌旁组织,差异有统计学意义(P＜0.05).而SATB1蛋白在结直肠癌组织中的阳性表达率与有无淋巴结转移显著相关(P＜0.01),有淋巴结转移其SATB1蛋白阳性表达率为100％,与肿瘤浸润深度和临床TNM分期密切相关(P＜0.05).SATB1 mRNA的表达与肿瘤浸润深度、是否淋巴结转移显著相关(P＜0.01),与肿瘤临床分期密切相关(P＜0.05).结论 SATB1基因在癌组织中的表达较癌旁组织显著增高表明其可能与结直肠癌的发生有关,而SATB1高表达预示结直肠癌恶性程度高临床分期较晚,可能存在淋巴结转移,预后不良,因此检测SATB1表达可作为临床上筛选结直肠癌高危转移患者、制定治疗方案和判断预后的一个新指标,并有望成为新的治疗靶点.     

ADAMTS1、ADAMTS18基因在食管鳞癌中的表达及其临床意义

目的:分析食管鳞状细胞癌(esophageal squamous cell carcinoma,ESCC)组织中含凝血酶敏感素1型基序的解聚素样金属蛋白酶(a disintegrin and metalloproteinase with thrombospondin motifs 1,ADAMTS1)及ADAMTS18基因的表达,探讨ADAMTS1和ADAMTS18基因在ESCC发生、发展中的作用。方法:收集2008年6月至2011年8月河北医科大学第四医院手术切除的72例ESCC组织及癌旁组织,应用RT-PCR及免疫组化S-P法分别检测ESCC组织及癌旁组织中ADAMTS1、ADAMTS18 mRNA及其蛋白的表达。结果:1 ESCC组织中ADAMTS1 mRNA的表达显著低于相应的癌旁组织[(0.394±0.123)vs(0.895±0.276),P<0.01],有淋巴结转移组ADAMTS1 mRNA表达显著高于无淋巴结转移组[(0.482±0.157)vs(0.298±0.102),P<0.05];2ESCC组织中ADAMTS18 mRNA表达显著低于相应的癌旁组织[(0.361±0.115)vs(0.879±0.265),P<0.01),高、中分化鳞癌组ADAMTS18 mRNA表达显著高于低分化鳞癌组[(0.496±0.153)vs(0.232±0.088),P<0.05];3ESCC组织中ADAMTS1和ADAMTS18蛋白阳性表达率显著低于癌旁组织[38.9%(28/72)、34.7%(25/72)vs94.4%(68/72)、93.1%(67/72),均P<0.01],高、中分化组ADAMTS18蛋白阳性表达率显著高于低分化鳞癌组[45.4%(20/44)vs 17.8%(5/28),P<0.05];4ESCC组织中ADAMTS1与ADAMTS18的蛋白表达无明显相关性(χ2=1.338,P>0.05)。结论:ADAMTS1基因在ESCC中的异常表达可能与ESCC的发生及有无淋巴结转移有关;ADAMTS18基因在ESCC中的异常表达可能与ESCC的发生及组织分化程度有关;两种基因在ESCC组织中蛋白的表达不存在明显的相关性。     

IL-6与VEGF在食管鳞状细胞癌中的表达及其临床意义

目的：检测食管鳞状细胞癌（esophageal squamous cell cancer,ESCC）患者血清中白细胞介素-6（interleukin-6,IL-6）和ESCC组织中血管内皮生长因子（vascular endothelial growth factor,VEGF）的表达及其两者关系,并研究两者表达对ESCC的临床意义。方法：收集河北医科大学第四医院胸外科于2014年1月至2015年1月期间行ESCC切除术的患者52例,每例患者均取原发灶组织和癌旁组织标本;同时在术前抽取患者外周血5 ml,再取52例健康体检者外周血5 ml为血清对照。应用ELISA法测定ESCC患者血清中IL-6的水平,免疫组化技术检测VEGF在ESCC组织中的表达,RT-PCR法检测肿瘤组织中IL-6和VEGF mRNA的表达情况。结果：ESCC患者血清IL-6表达水平为（116.71±25.98）pg/ml,明显高于健康对照组的\[（78.43±9.36）pg/ml\]（P＜0.05）,血清中IL-6的表达水平与患者肿瘤分化程度、TNM分期、肿瘤浸润深度和淋巴结转移相关（P＜0.05）。肿瘤组织VEGF蛋白阳性表达率明显高于癌旁组织（67.31% vs 32.69%,P<0.01),且与患者肿瘤分化程度、TNM分期、肿瘤浸润深度和淋巴结转移相关（P＜0.05）。肿瘤组织中IL-6 mRNA的表达与VEGF mRNA的表达呈显著正相关（r=7.113,P＜0.05）。结论：ESCC患者肿瘤组织中IL-6和VEGF均呈高表达且两者呈正相关,两者可能在ESCC侵袭和转移中发挥重要作用。     

半乳糖凝集素-9在食管鳞状细胞癌中的表达及其与临床病理特征的关系

目的:探讨半乳糖凝集素-9(galectin-9)在食管鳞状细胞癌(esophageal squamous cell cancer,ESCC)组织及癌旁组织中的表达及对ESCC细胞迁移和侵袭能力的影响.方法:收集山东大学附属济南市中心医院胸外科于2012年1月至2013年1月期间手术切除的ESCC患者组织标本89例及距癌组织边缘5 cm以上的癌旁组织标本20例,采用免疫组化法检测ES-CC及癌旁组织中galectin-9蛋白的表达,并分析其与患者性别、年龄、浸润深度、肿瘤大小、分化程度、病理分期(pTNM)及淋巴结转移等临床病理特征之间的关系.采用脂质体介导法将pcDNA3.1-galectin-9转染食管癌EC9706细胞系,运用细胞划痕愈合实验和Transwell侵袭实验,检测galectin-9表达上调对食管癌EC9706细胞迁移、侵袭能力的影响.结果:ESCC组织中galectin-9蛋白的阳性表达率明显低于癌旁组织[25.8％ (23/89) vs 50％ (10/20),P＜0.05],galectin-9蛋白的表达与ESCC患者肿瘤分化程度及淋巴结转移相关(均P＜0.05),而与患者性别、年龄、肿瘤大小、浸润深度及病理分期无关(均P＞0.05).在食管癌EC9706细胞中过表达galectin-9后,与阴性对照组相比,肿瘤细胞的侵袭[(45.0±8.0) vs (160.0±12.0),P＜0.01]和迁移能力[(20.6±2.1) vs(87.6±4.9),P＜0.05]明显下降.结论:galectin-9的表达缺失参与了食管癌的发生、发展,ga-lectin-9可以抑制ESCC细胞的侵袭及迁移.     

食管鳞癌组织MTA2表达与淋巴结转移相关性的探讨

目的:探讨食管鳞状细胞癌(ESCC)组织中转移相关基因2(MTA2)的表达与淋巴结转移的关系及其临床意义.方法:免疫组织化学(SP)方法及流式细胞技术检测102例ESCC标本反12例癌旁正常黏膜组织中MTA2蛋白的表达,统计分析其表达与ESCC淋巴结转移的关系.结果:MTA2在癌旁正常食管黏膜组织中无表达,在部分ESCC组织中呈阳性表达,阳性率为68.6％(70/102),其中强阳性28例;ESCC组织中淋巴结转移组与未转移组MTA2蛋白表达的强阳性率分别为68.2％(15/22)、27.1％(13/48),差异有统计学意义,P=0.003; MTA2的阳性表达与ESCC的淋巴结转移数目及淋巴结转移率均呈正相关,P值分别为0.006、0.001;流式细胞技术结果显示,淋巴结转移组与非转移组MTA2蛋白相对含量分别为252.11±30.22、161.42±22.89,差异有统计学意义,t=15.11,P=0.000.结论:MTA2蛋白的阳性表达与SCC的淋巴结转移密切相关,MTA2可能是ESCC一种新的标志及治疗靶点.     

lncRNARP11-259P1.1在食管鳞状细胞癌中的表达及其与临床预后的关系

目的:探讨lncRNA RP 11-259P 1.1在食管鳞状细胞癌(esophageal squamous cell cancer,ESCC)中的表达水平及其与患者临床病理特征和预后的关系.方法:收集2012年1月1日至2016年12月31日西南医科大学附属医院行手术切除的130例ESCC原发病灶及对应癌旁组织标本、人ESCC细胞株KYSE510、Eca109及食管上皮细胞株Het-1A,采用实时荧光定量PCR法检测lncRNA RP11-259P 1.1在ESCC组织及细胞中的表达水平,分析其在组织中表达水平与患者临床病理特征及预后的关系.结果:lncRNA RP11-259P1.1在ESCC细胞及组织中表达水平显著高于正常管上皮细胞及癌旁组织(均P＜0.01).lncRNA RP11-259P1.1高表达与肿瘤大小、肿瘤分期、淋巴结转移及术前放化疗(CRT)后肿瘤缓解明显相关(均P＜0.05),与患者年龄、性别、肿瘤部位、吸烟状况等无关(均P＞0.05).在63例术前接受新辅助CRT治疗患者中,低表达lncRNA RP11-259P1.1患者与高表达患者比较:(1)病理完全缓解率明显升高(60.00％ vs 21.21％,P＜0.01);(2)术前CRT的疗效更佳(P＜0.05);(3)中位无进展生存时间和生存时间均明显延长[(28.00±2.47) vs(17.00±1.90)个月,(41.57±2.45) vs (30.00±2.55)个月;均P＜0.01].lncRNA RP11-259P1.1为ESCC患者独立的预后因素(P＜0.05).结论:lncRNA RP11-259P1.1可能是ESCC潜在的预后和术前CRT疗效评价的分子标志物,低表达lncRNA RP 11-259P 1.1的患者术前给予CRT疗效更佳.     

Ets-1、VEGF在乳腺浸润性导管癌组织的表达及其临床意义

目的:探讨Ets-1和VEGF在乳腺浸润性导管癌( invasive ductal carcinoma,IDC)组织中的表达规律,分析其与临床病理特征的关系.方法:选择临床确诊的40例乳腺浸润性导管癌患者手术切除癌组织和癌旁组织标本,通过免疫组织化学和RT-PCR方法检测Ets-1和VEGF蛋白和mRNA的表达.结果:(1)Ets-1和VEGF蛋白在乳腺IDC组织中高表达,明显高于癌旁乳腺组织(P＜0.01);癌组织中Ets-1蛋白表达高于VEGF,且两者呈正相关(r为0.8827,P＜0.05);Ets-1和VEGF蛋白表达与年龄、肿瘤大小无关,与临床分期、淋巴结转移密切相关(P<0.05).(2)Ets-1 mRNA、VEGF mRNA在乳腺IDC组织的表达明显高于癌旁乳腺组织(P＜0.01);Ets-1mRNA水平明显高于VEGF mRNA (P＜0.01),且两者正相关(r＝0.984,P＜0.01);Ets-1 mRNA、VEGF mRNA表达与年龄、肿瘤大小无关,与临床分期、淋巴结转移密切相关(P<0.05).结论:乳腺浸润性导管癌组织中Ets-1和VEGF在mRNA和蛋白水平均高表达,两者表达间均呈正相关;两者表达均与肿瘤临床分期和淋巴结转移相关.     

Ⅱ相代谢酶基因多态性与广西肝癌发生的关系

目的　探讨Ⅱ相代谢酶谷胱甘肽硫转移酶M 1、T 1(GSTM 1、GSTT1)及微粒体环氧化物水解酶 (mEH )基因多态性与广西肝癌易感性的关系 ,以及基因与基因间的相互作用。方法　采用多重PCR、PCR RFLP技术 ,对广西地区 10 5例肝癌患者及 15 1例健康对照的GSTM 1、GSTT 1、mEH基因型进行检测。结果　GSTM 1基因缺失率在病例组与对照组中分别为 64 .76%和5 0 .99% ,两者比较有显著性差异 (P <0 .0 5 ,OR =1.77) ;病例组GSTT 1基因缺失率 (4 0 .95 % )高于对照组 (3 3 .11% ) ,mEH 3种基因型频率在病例组分别为 2 7.62 %、2 1.90 %、5 0 .48% ,对照组则分别为 2 1.19%、3 4.44 %、44 .3 7% ,两组比较无显著性差异 (P >0 .0 5 ) ;GSTM 1、T 1基因同时缺失的个体患肝癌的危险性增大了 1.2 2倍。结论　GSTM 1、T1基因同时缺失是肝癌的易感因素 ,可作为肝癌高危人群筛选的标记物。     

Expression of seven-in-absentia homologue 1 and hypoxia-inducible factor 1 alpha: Novel prognostic factors of nasopharyngeal carcinoma

Nasopharyngeal carcinoma (NPC) is an EBV-associated cancer. We analysed Siah1 expression as well as LMP1 and HIF1α expression by immuno-histochemical staining in 74 NPC biopsy specimens and found that the expression of Siah1 was significantly correlated with advanced tumour status and stage. Moreover, Siah1-positive and HIF1α-positive cases had significantly worse prognoses. The expression score for LMP1 was remarkably correlated with that of Siah1, whereas there was little correlation between LMP1 expression and the other markers evaluated. This is the first study to evaluate the pattern and clinical significance of Siah1 and HIF1α expression in NPC, and such an evaluation is valuable for identifying those patients at a high risk for a poor prognosis.     

Upregulated long non-coding RNA AFAP1-AS1 expression is associated with progression and poor prognosis of nasopharyngeal carcinoma

Altered expression of long noncoding RNAs (lncRNAs) associated with human carcinogenesis. We performed a cDNA microarray analysis of lncRNA expression in 12 cases of nasopharyngeal carcinoma (NPC) and 4 non-tumor nasopharyngeal epitheliums. One lncRNA, actin filament associated protein 1 antisense RNA1 (AFAP1-AS1), was identified and selected for further study. AFAP1-AS1 expression was upregulated in NPC and associated with NPC metastasis and poor prognosis. In vitro experiments demonstrated that AFAP1-AS1 knockdown significantly inhibited the NPC cell migration and invasive capability. AFAP1-AS1 knockdown also increased AFAP1 protein expression. Proteomic and bioinformatics analyses suggested that AFAP1-AS1 affected the expression of several small GTPase family members and molecules in the actin cytokeratin signaling pathway. AFAP1-AS1 promoted cancer cell metastasis via regulation of actin filament integrity. AFAP1-AS1 might be a potential novel marker that can predict cancer patient prognosis and as a potential therapeutic target for NPC.     

Methylation Profiles of BRCA1, RASSF1A and GSTP1 in Vietnamese Women with Breast Cancer

Objective: This study investigated the DNA promoter methylation profiles of BRCA1, RASSF1A and GSTP1 genes,both individually and in an integrative manner in order to clarify their correlation with clinicopathological parameters ofbreast cancer from Vietnamese patients, and establish new potential integrative methylation biomarkers for breast cancerdetection. Material and methods: The methylation frequencies of BRCA1, RASSF1A and GSTP1 were analyzed bymethylation specific polymerase chain reaction (MSP) in 70 specimens of breast carcinomas and 79 pairs of tumor andmatched adjacent normal tissues from breast cancer patients. Results: All the three analyzed genes showed a concordanceconcerning their promoter methylation in tumor and adjacent normal tissue. The methylation of BRCA1, RASSF1Aand GSTP1 was found in 58.23 %, 74.68 % and 59.49 % of tumor tissues and 51.90 %, 63.29 % and 35.44 % ofcorresponding adjacent tissues, respectively. When each gene was assessed individually, only the methylation ofGSTP1 was significantly associated with tumor tissues (p=0.003). However, the methylation frequency of at least one ofthe three genes and the methylation frequency of all the three genes both showed significant association with tumor(p=0.008 and p=0.04, respectively). The methylation of BRCA1 was found to be significantly associated with tumorgrade (p=0.01). Conclusion: This study emphasized that the panel of the three genes BRCA1, RASSF1A and GSTP1can be further developed as potential biomarkers in diagnosis and classification of breast cancer in Vietnamese women.     

大、小鼠摄入AFB_1和羧乙基锗倍半氧化物体内超氧化物歧化酶变化研究

本实验通过观察摄入ＡＦＢ１及Ｇｅ－１３２后Ｗｉｓｔａｒ大鼠和昆明小鼠体内ＳＯＤ活力的变化，进一步了解ＡＦＢ１代谢途径和Ｇｅ－１３２与自由基的关系。实验表明，摄入ＡＦＢ１的ＳＯＤ活力受到抑制，而摄入Ｇｅ－１３２后ＳＯＤ活力得到提高，且一次性摄入ＡＦＢ１后ＳＯＤ活力最终得到恢复，其中大鼠对ＡＦＢ１敏感性相对较大。     

PDK-1 regulates lactate production in hypoxia and is associated with poor prognosis in head and neck squamous cancer

Here we describe the expression and function of a HIF-1-regulated protein pyruvate dehydrogenase kinase-1 (PDK-1) in head and neck squamous cancer (HNSCC). Using RNAi to downregulate hypoxia-inducible PDK-1, we found that lactate and pyruvate excretion after 16-48 h of hypoxia was suppressed to normoxic levels. This indicates that PDK-1 plays an important role in maintaining glycolysis. Knockdown had no effect on proliferation or survival under hypoxia. The immunohistochemical expression of PDK-1 was assessed in 140 cases of HNSCC. PDK-1 expression was not expressed in normal tissues but was upregulated in HNSCC and found to be predominantly cytoplasmic with occasional strong focal nuclear expression. It was strongly related to poor outcome (P=0.005 split by median). These results indicate that HIF regulation of PDK-1 has a key role in maintaining lactate production in human cancer and that the investigation of PDK-1 inhibitors should be investigated for antitumour effects.     

新基因NBEAL1的克隆、表达、纯化及其与胶质瘤病理分级的相关性

目的：构建新基因neurobeachin like 1（NBEAL1）的表达载体,研究NBEAL1 的表达与胶质瘤病理分级的相关性。方法：提取人脑胶质瘤细胞U251的总RNA,构建pGEX-KG/NBEAL1表达载体,转化大肠杆菌BL21,IPTG诱导NBEAL1重组蛋白的表达,GST亲和纯化,Western blotting鉴定重组蛋白的纯度;以免疫组化法用纯化蛋白制备的单克隆抗体分析NBEAL1表达与胶质瘤病理分级的相关性。结果：NBEAL1基因片段被成功地克隆入pGEXKG表达载体,测序证实克隆片段序列正确。NBEAL1融合蛋白在大肠杆菌BL21包涵体中表达,表达量占菌体总蛋白的30%以上,纯化的重组蛋白纯度95%以上。Western blotting证明所纯化的蛋白含有GST标签与NBEAL1肽段。免疫组化法检测显示NBEAL1蛋白的表达在正常脑组织中较低,而在低级别的脑胶质瘤组织中表达明显上调,但随着恶性程度增加NBEAL1的表达程度降低,两者呈负相关。结论：成功地克隆、表达及纯化NBEAL1蛋白,NBEAL1蛋白在人脑胶质瘤组织中的表达与其恶性程度呈负相关。     

Triplex Polymerase Chain Reaction with Confronting Two-Pair Primers (PCR-CTPP) for NQO1 C609T,GSTM1 and GSTT1 Polymorphisms: the Most Convenient Genotyping Method

The polymerase chain reaction with confronting two-pair primers (PCR-CTPP) is a time-saving and inexpensive ‍genotyping method, which is applicable for most single nucleotide polymorphisms (SNPs). To date, we have established ‍PCR-CTPP conditions for tens of SNPs, including duplex genotyping. This paper introduces triplex PCR-CTPP to ‍simultaneously genotype three functional polymorphisms of carcinogen-detoxifying enzymes, NQO1 C609T, GSTM1 ‍null, and GSTT1 null, all of which are reported to have a significant association with smoking-related cancers. We ‍applied this method for 241 non-cancer patients to demonstrate the performance. Among the subjects, the genotype ‍frequency of NQO1 C609T was 35.7% for CC, 44.4% for CT and 19.9% for TT. The null type frequencies of GSTM1 ‍and GSTT1 were 53.4% and 44.0%, respectively. Their distributions were similar to those reported for Japanese by ‍other studies. This is the first paper reporting the success of triplex PCR-CTPP. The polymorphisms applied are ‍useful examples, which could be adopted not only for research purposes, but also for risk assessment of individuals ‍exposed to carcinogenic substances, such as smokers. This convenient genotyping approach has advantages for ‍application in cancer prevention, especially in the Asian Pacific region.     

Multiplex PCR with Confronting Two-pair Primers for CYP1A1 Ile462Val,GSTM1, GSTT1, and NQO1 C609T

Polymerase chain reaction with confronting two-pair primers (PCR-CTPP) is an effective genotyping method ‍for single nucleotide polymorphisms (SNPs) in aspects of reducing time and costs for analysis. So far we have ‍established PCR-CTPP conditions for tens of SNPs, including a triplex genotyping (Kawase et al., 2003). In the ‍present study we report a quadruplex PCR-CTPP to genotype simultaneously four functional polymorphisms of ‍carcinogen-metabolizing enzymes, CYP1A1 Ile462Val, GSTM1 null, GSTT1 null and NQO1 C609T, which were ‍reported that they have significant associations with smoking-related cancers. We applied this method for 475 health ‍check-up examinees to demonstrate the performance. Among the subjects, the genotype frequency of CYP1A1 ‍Ile462Val was 56.8% for Ile/Ile, 38.1% for Ile/Val and 5.1% for Val/Val. The null type frequencies of GSTM1 and ‍GSTT1 were 52.8% and 49.9%, respectively. And the genotype frequency of NQO1 C609T was 41.9% for C/C, ‍41.3% for C/T and 16.8% for T/T. Their distributions were similar to those reported for Japanese by other studies. ‍To the best of our awareness, this is the first paper that reports the success in quadruplex PCR-CTPP. The applied ‍polymorphisms are useful ones, which would be adopted not only for research purposes, but also for risk assessment ‍of individuals exposed to carcinogenic substances. This convenient genotyping would be applied for cancer prevention ‍especially in Asian Pacific regions, where expensive genotyping methods are hardly available.