A Validated Resource of Thyroid Cancer Cell Lines for Pathway Discovery

描述（由申请人提供）：该申请涉及广泛的挑战领域（15）转化科学，15- ca -103：甲状腺癌细胞系项目，该项目直接响应我们最近的发现，显示40个“甲状腺”癌细胞系中有17个是多余的或被错误识别的（即不同的肿瘤谱系，包括黑色素瘤和结肠癌）。其余23个系是独特的，基于有限数量的甲状腺特异性基因的表达，可能起源于甲状腺。然而，这些细胞系在遗传上与肿瘤的起源没有联系，也没有在遗传或分子水平上仔细地表征。在最近的NCI会议上，主要研究人员参加了对研究界这个问题的讨论。研究人员强调，需要经过充分验证的甲状腺癌细胞系来研究癌症的发生和进展机制以及发现新的治疗靶点。在本提案中，我们将使用三种互补的策略来提供大量全面表征的甲状腺癌细胞系，代表不同组织学类型和突变亚型（BRAF, RET/PTC， RAS, PIK3CA, AKT1, CTNNB1）的原发性和转移性肿瘤。我们将专注于从低分化甲状腺癌和转移性病变中培养细胞系，这些肿瘤更有可能携带突变，而这些突变在目前的甲状腺癌细胞系中没有表现出来。为了建立新的细胞系，我们将使用体内和体外并行的方法来最大化我们的成功率。新的细胞系将被仔细表征，并通过短串联重复序列（STR）分析与相应的肿瘤组织进行比较，以确认其来源。STR分析和甲状腺特异性基因的表达将用于研究文献中报道的另外36个甲状腺细胞系的起源，以便组装一个完整的甲状腺癌细胞系。我们将使用真实甲状腺癌细胞系的表达阵列来识别一个签名，该签名可用于验证失去甲状腺特异性标记表达或无法获得患者DNA的细胞系。最后，我们将应用全球分子和基因组方法，对新的和现有的甲状腺癌细胞系进行新颖的计算分析，以揭示甲状腺癌发生和进展的重要途径。所有细胞系，以及基因组和基因表达数据，将在NCI的协助下通过中央存储库提供给研究界，数据将通过NCBI GEO数据库和UCSC基因组浏览器访问。可行性和影响：从实体肿瘤中产生新的细胞系是一项具有挑战性的任务。我们建议在12-15个月的时间内使用平行的体内和体外方法产生至少10个新细胞系。科罗拉多大学癌症中心（University of Colorado Cancer Center， UCCC）拥有丰富的专业知识，在从实体癌中培养细胞系方面取得了成功。我们相信这一雄心勃勃的提议在两年的时间内是可行的，因为加州大学丹佛分校和MSKCC之间的独特合作目前拥有所有这些先进技术（开发细胞系的新程序，基因表达平台，质谱基因分型，CGH阵列分析，关键癌基因和肿瘤抑制基因的突变筛选以及通过Western blotting进行高通量途径激活分析）。阵列和突变筛选的大部分样品制备和研究将由经验丰富的人员在设备齐全的中心进行，周转时间短。此外，我们正在与国家认可的生物信息学和计算专家合作，应用新的计算生物学策略来识别重要的信号通路，从而更好地定义疾病的发病机制。这些目标的成功完成将提供大约40个具有良好特征的甲状腺癌细胞系，供研究界用于揭示甲状腺癌的发生、进展、转移潜力的机制，并作为识别晚期疾病的新治疗靶点的平台。摘要：甲状腺癌是最常见的内分泌恶性肿瘤，年发病率约3.4万例。在美国，至少有30万人被诊断患有甲状腺癌，其中每年有1500多人死于这种疾病。来自患者癌症的永久性甲状腺癌细胞系对于帮助我们了解甲状腺癌的病因，肿瘤如何生长和扩散，以及我们如何找到治疗目前无法治愈的晚期疾病的新方法至关重要。我们的研究小组最近的研究表明，目前使用的近一半的“甲状腺”癌细胞系并不是唯一的，许多甚至不是起源于甲状腺癌。为了提供具有良好特征的甲状腺癌细胞系供研究界研究，我们将产生新的甲状腺癌细胞系，这些癌细胞系可以与原始肿瘤组织联系起来。我们还将开发一些方法，通过这些方法我们可以确定一些现有的甲状腺癌细胞系是否真实。我们将开发所有癌细胞系的遗传图谱，从中我们将能够表征可能解释这些癌症如何发展的异常信号网络。一旦我们知道是什么导致了这些癌症的发展，我们就能更好地了解如何治疗它们。在国家癌症研究所的帮助下，我们将把这些细胞系广泛地提供给研究界。

DESCRIPTION (provided by applicant): This application addresses the broad Challenge Area (15) Translational Science, 15-CA-103: Thyroid Cancer Cell Line Project which is in direct response to our recent discovery showing that 17 out of 40 "thyroid" cancer cell lines are either redundant or misidentified (i.e. of a different tumor lineage including melanoma and colon cancer). The remaining 23 lines are unique and are likely of thyroid origin based on the expression of a limited number of thyroid-specific genes. These cell lines, however, have not been genetically linked to tumors of origin or carefully characterized at a genetic or molecular level. This problem for the research community was reviewed at a recent NCI conference in which the Principal Investigators participated. The need for well- validated thyroid cancer cell lines to study mechanisms of cancer development and progression as well as discovery of novel therapeutic targets was emphasized. In this proposal, we will use three complementary strategies to provide a large panel of comprehensively characterized thyroid cancer cell lines representing primary and metastatic tumors of different histologic types and mutational subtypes (BRAF, RET/PTC, RAS, PIK3CA, AKT1, CTNNB1). We will focus on developing cell lines from poorly differentiated thyroid cancer as well as metastatic lesions that are more likely to harbor mutations that are not represented in the current panel of thyroid cancer cell lines. To establish new cell lines, we will use a parallel in vivo and in vitro approach to maximize our success rate. The new cell lines will be carefully characterized and compared to the corresponding tumor tissue of origin by short tandem repeat (STR) profiling to confirm their derivation. STR profiling and expression of thyroid-specific genes will be used investigate the origins of an additional 36 thyroid cell lines reported in the literature in order to assemble a complete panel of thyroid cancer cell lines. We will use expression arrays of authentic thyroid cancer cell lines to identify a signature that can be used to validate cell lines that have lost expression of thyroid-specific markers or from which patient DNA is unavailable. Finally, we will apply global molecular and genomic approaches with novel computational analyses to the new and existing thyroid cancer cell lines to uncover pathways important in thyroid cancer development and progression. All cell lines, as well as the genomic and gene expression data, will be made available to the research community through a central repository with the assistance of the NCI, and data will be accessible through the NCBI GEO database and UCSC genome browser. Feasibility and Impact: The generation of new cell lines from solid tumors is a challenging task. We propose to generate at least 10 new cell lines in a 12-15 month time period using parallel in vivo and in vitro approaches. There is significant expertise at the University of Colorado Cancer Center (UCCC) with a proven record of success in generating cell lines from solid cancers. We believe that this ambitious proposal is feasible within the two-year time frame because the unique collaboration between the UC Denver and MSKCC has all of these advanced technologies currently in place (new procedures to develop cell lines, gene expression platforms, mass spectrometry genotyping, CGH array analysis, mutational screen of key oncogenes and tumor suppressor genes and high throughput pathway activation analysis by Western blotting). A majority of the sample preparation and studies for the arrays and mutation screening will be performed by experienced personnel in well-equipped cores with rapid turn-around times. Furthermore, we are collaborating with nationally recognized bioinformatics and computational experts to apply novel computational biology strategies to identify important signaling pathways that will better define disease pathogenesis. Successful completion of these aims will provide approximately 40 well-characterized thyroid cancer cell lines for use by the research community to uncover mechanisms of thyroid cancer development, progression, metastatic potential and to serve as a platform for identification of novel therapeutic targets for advanced disease. Lay Summary: Thyroid cancer is the most common endocrine malignancy with an annual incidence of ~34,000. At least 300,000 people are living with a diagnosis of thyroid cancer in the United States, and more than 1,500 of them die each year from this disease. Permanent thyroid cancer cell lines derived from patient's cancers are critical to help us understand the causes of thyroid cancer, how the tumor grows and spreads, and how we can find new ways to treat advanced disease for which there is currently no cure. Recent studies by our research groups have shown that nearly half of the "thyroid" cancer cell lines in current use are not unique and many did not even originate from a thyroid cancer. In order to provide well characterized thyroid cancer cell lines for study by the research community, we will generate new thyroid cancer cell lines that can be linked back to the original tumor tissue. We will also develop approaches by which we can determine whether some of the existing thyroid cancer cell lines are authentic or not. We will develop genetic profiles of all the cancer cell lines, from which we will be able to characterize the abnormal signaling networks that may explain how these cancers develop. Once we know what makes these cancers grow, we will have a better understanding of how to treat them. With the help of the National Cancer Institute, we will make these cell lines widely available to the research community. PUBLIC HEALTH RELEVANCE: The generation of a well-characterized panel of thyroid cancer cell lines has significant relevance for public health as validated tools for the broader research community. Successful completion of the proposed aims will provide approximately 40 well-characterized thyroid cancer cell lines in a central repository for use by the research community to uncover mechanisms of thyroid cancer development, progression, metastatic potential and to serve as a platform for identification of novel therapeutic targets for many types of advanced cancer.