Molecular Genetic Epidemiology of Leading U.S. Cancers

美国主要癌症的分子遗传学流行病学

• 批准号: 8157731
• 负责人: Kenneth Buetow
• 金额: $ 4.55万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8157731
• 关键词: Malignant Neoplasms; Molecular Genetics; genetic epidemiology

Genotyping assays were successfully carried out for 35 candidate loci. DNA was isolated from samples of whole blood. Single Necleotide polymorphism (SNP)genotypes were determined using homogeneous MassEXTENDTM assays with primer extension (Sequenom, San Diego, CA; www.sequenom.com). Sequenom is a high-throughput, multiplex SNP genotyping chemistry, which employs matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF)distinguish between the allele-specific extension products. Each SNP locus could be assayed with 2-4 ng of DNA. PCR amplification of short tandem repeat polymorphic fragments (STRPs) was performed using custom primers (Applied Biosystems, Foster City, CA) PCR products were detected and separated with an ABI 3730 DNA Analyzer (Applied Biosystems, Foster City, CA) and allele sizing and calling was carried out using GeneMapper 3.0 software (Applied Biosystems, Foster City, CA). We have evaluated correlations between variants of multiple genes simultaneously, viewing them as a composite system, and culminating in a pathway based on the pre-existing biological literature. We have shown that by treating the entire system as a marker of drug response, the limitations inherent in evaluating individual genetic variants, each conferring a minor effect on clinical outcome, can be overcome. Critical to this process is the underlying candidate gene approach, with variants known from pre-existing biological literature offering a qualitative framework on which to quantitatively model actual data. Our application of pathway analysis to the genetic network underlying tamoxifen metabolism has shown the ability to discern differences between two drug exposures.

已成功对 35 个候选基因座进行基因分型测定。从全血样本中分离出 DNA。使用具有引物延伸的同质 MassEXTENDTM 测定法（Sequenom，圣地亚哥，加利福尼亚州；www.sequenom.com）确定单核苷酸多态性（SNP）基因型。 Sequenom 是一种高通量、多重 SNP 基因分型化学，采用基质辅助激光解吸/电离飞行时间质谱 (MALDI-TOF) 区分等位基因特异性延伸产物。 每个 SNP 位点可用 2-4 ng DNA 进行分析。使用定制引物（Applied Biosystems，福斯特城，加利福尼亚州）进行短串联重复多态性片段（STRP）的 PCR 扩增，使用 ABI 3730 DNA 分析仪（Applied Biosystems，福斯特城，加利福尼亚州）检测和分离 PCR 产物，并使用 GeneMapper 3.0 软件（Applied Biosystems，福斯特城，加利福尼亚州）进行等位基因大小和识别。 我们同时评估了多个基因变异之间的相关性，将它们视为一个复合系统，并最终形成了一条基于现有生物学文献的途径。我们已经证明，通过将整个系统视为药物反应的标志，可以克服评估个体遗传变异所固有的局限性，每个遗传变异对临床结果影响较小。这一过程的关键是潜在的候选基因方法，从现有的生物学文献中已知的变体提供了一个定性框架，可以在该框架上对实际数据进行定量建模。我们对他莫昔芬代谢的遗传网络进行路径分析的应用表明能够辨别两种药物暴露之间的差异。