Typing the Transcriptome in Cancer Using Splicing Array

描述(由申请人提供)： 选择性剪接是高等真核细胞的永久特征，理解选择性剪接如何改变蛋白质组的组成和功能是后基因组时代的一大挑战。对于癌症研究，独特的信使核糖核酸异构体可能为诊断和预后提供一套强有力的生物标志物，而癌症特异性信使核糖核酸异构体可能成为有效治疗干预的区分靶点。此外，理解剪接选择是如何在发育和疾病中做出和调节的，是癌细胞生物学中的一个基本问题。信使核糖核酸异构体敏感的微阵列技术将非常适合和及时地解决与选择性剪接有关的广泛的临床和机制问题。 在过去的IMAT资助期，我们开发了一个独特而新颖的技术平台来解决拼接问题。经过系统和大量的数据库建设和实验开发，目前该技术已经成熟，并在各种条件下可重复性测定mRNA异构体方面显示了其优越性。剪接阵列的独特之处在于需要逐步扩大用于准确注释的mRNA异构体的数据库，并准备用于测量的相应寡核苷酸集。因此，我们正在寻求IMAT的支持，以便将技术付诸实践，并让研究界利用技术发展。 我们在下一阶段将这项新兴技术应用于癌症分子分析方面有三个具体目标。(1)我们计划使用这项技术来识别与前列腺癌相关的独特的mRNA亚型。我们将调查现有的未经处理或用雄激素和雌激素处理的前列腺癌细胞系，以及处于不同恶性阶段的癌症组织，以确定肿瘤特异性和激素调节的替代剪接。(2)我们建议应用这项技术来研究剪接调控机制，方法是在击倒和敲除细胞中识别大量剪接调控因子的直接靶点。(3)随着拟议的技术应用，我们将随着技术的发展逐步扩大高质量的数据库，并通过增加新的特性和功能来继续完善和扩大数据库，并开发用于拼接阵列数据分析的链接软件。

DESCRIPTION (provided by applicant): Alternative splicing is a permanent feature in higher eukaryotic cells and understanding of how alternative splicing alters the composition and function of the proteome represents a major challenge in the post-genome era. For cancer research, unique mRNA isoforms may provide a robust set of biomarkers for diagnosis and prognosis, and cancer-specific mRNA isoforms may serve as discriminating targets for effective therapeutic interventions. Furthermore, understanding of how splice choice is made and regulated in development and disease is a fundamental issue in cancer cell biology. An mRNA isoform-sensitive microarray technology would be ideally and timely suited for addressing a wide range of clinical and mechanistic questions regarding alternative splicing. In the past IMAT funding period, we have developed a unique and novel technology platform to attack the splicing problem. After a systematic and substantial effort in database construction and experimental development, the technology is now matured, and its superiority in reproducible measurement of mRNA isoforms under a variety of conditions has been demonstrated. Unique to the splicing array is the need to progressively enlarge the database for accurately annotated mRNA isoforms and preparation of corresponding oligo sets for measurement. We are therefore seeking IMAT support to put the technology in practical use and let the research community to take advantage of the technology development. We have three specific goals for the next phase in applying the emerging technology for molecular analysis of cancer. (1) We plan to use the technology to identify unique mRNA isoforms associated with prostate cancer. We will survey existing prostate cancer cell lines untreated or treated with androgen and estrogen as well as cancer tissues at different malignant stages to identify tumor-specific and hormonal regulated alternative splicing. (2) We propose to apply the technology to address mechanisms of splicing regulation by identifying direct targets for a large number of splicing regulators in knockdown and knockout cells. (3) Along with the proposed technology applications, we will progressively enlarge the high quantity databases coupled with the technology development and continue to improve and enlarge the database by adding new features and functions and develop linked software for splicing array data analysis.