Functional genomics of breast cancer

同时应用了许多技术来确定特定生物旋转剂的分子轮廓。这些技术中的大多数目前使用基于微阵列的方法，但它们正在迅速得到补充，甚至被不断发展的下一代DNA测序技术所取代。几种不同种类的微阵列用于不同的目的，但目前主要的技术方法使用与固体载体结合的合成寡核苷酸，并与从感兴趣的生物样品制备的标记核酸进行询问。在这项技术的当前实施例中，这种方法的力量很大程度上基于已知基因组序列与完全由计算手段控制的微阵列设计之间的直接联系。这使得研究人员可以构建专门针对所需分析量身定做的任意设计的阵列，并将阵列的分辨率调整到非常精细的水平。因此，例如，现在可以通过外显子确定mRNAs外显子的表达，并在比单基因分辨率更好的情况下观察基因拷贝数的变化(扩增或缺失)。从任何感兴趣的细胞或组织来源制备的荧光探针然后与这些阵列杂交，提供大规模的高分辨率基因组图像。目前，我们正致力于将尽可能多的分析方法过渡到微量样本(例如在常规临床护理过程中通常可能收集的样本)和福尔马林固定石蜡包埋(FFPE)样本。当人们考虑到将在这项工作过程中所做的发现转化为临床护理的可能性时，使用FFPE样本的能力尤其重要，在临床实验室中，基于FFPE的方法是稳定生物菌群的标准方法。重要的是，我们已经证明，有可能在数百个样本上并行确定400,000多个CPGS的甲基化状态，结果与从冷冻样本获得的结果相匹配。这打开了大量现有的FFPE样本档案供调查。我们现在也经常从FFPE样本中获得优秀的拷贝数数据。我们对特定转录因子在确定乳腺癌表型中的作用特别感兴趣，并一直在通过染色质免疫沉淀结合微阵列或测序分析来研究这些作用。我们最近使用DNA甲基化图谱研究了乳腺癌的表观遗传学特征。我们证明，表达雌激素受体α的肿瘤有两个亚组，根据它们的甲基化特征可以明显区分它们。其中一种与正常乳腺上皮的甲基化模式不同，其预后明显更差。我们有兴趣更好地描述这一现象，并将其与基因表达及其潜在的生物学机制联系起来。

A number of technologies are applied in parallel to determine the molecular profile of a given biospecimen. The majority of these technologies currently use microarray based methods, but they are rapidly being supplemented and even supplanted by evolving next generation DNA sequencing technologies. Several varieties of microarray are used for various purposes, but the predominant current technical approaches use synthetic oligonucleotides bound to a solid support and interrogated with labeled nucleic acids prepared from the biospecimen of interest. The power of this approach in the current embodiment of this technology is based largely on the direct connection between known genome sequence and the design of microarrays completely controlled by computational means. This allows the investigator to construct arrays of arbitrary design tailored specifically to the desired analysis and to adjust the resolution of the arrays to a remarkably fine level. Thus, for example, it is now possible to determine the expression of mRNAs exon by exon and to observe changes in gene copy number (amplification or deletion) at better than single gene resolution. Fluorescent probes prepared from any cell or tissue source of interest are then hybridized to these arrays providing a large scale high resolution view of the genome. Currently we are focused on transitioning as many assays as possible to minute samples (such as may typically be collected in the course of routine clinical care) and formalin fixed paraffin embedded (FFPE) specimens. The ability to work with FFPE samples is particularly important when one considers the potential to transition discoveries made in the course of this work to clinical care where FFPE based methods are the standard method of stabilizing biospecimens in the clinical laboratory. Of importance we have demonstrated that it is possible to determine the methylation status of more than 400,000 CpGs in parallel on hundreds of samples with results which match those obtained from frozen specimens. This opens vast existing archives of FFPE samples to investigation. We now routinely obtain excellent copy number data from FFPE samples as well. We are particularly interested in the role of specific transcription factors in determining breast cancer phenotypes and have been investigating these through chromatin immunoprecipitation combined with microarray or sequencing analysis. We have recently investigated the epigenetic profiles of breast cancers using DNA methylation profiling. We demonstrated that there are two subsets of estrogren receptor alpha expressing tumors which can be sharply distinguished based on their methylation profiles. One of these which deviates from the methylation pattern of normal mammary epithelium has a markedly worse prognosis. We are interested in better characterizing this phenomenon and linking it to gene expression and its underlying biological mechanisms.