Dissecting the contribution of the transcriptional regulators of SNS fate to neuroblastoma oncogenesis

项目摘要/摘要： 神经母细胞瘤(NB)是最常见的婴儿期癌症，起源于交感神经未成熟细胞。 从神经脊干细胞(NCC)衍生而来的神经系统(SNS)。NB负责15%的儿科 癌症死亡，大多数高危NB患者最终因耐药疾病复发。查找 对于高危神经母细胞瘤，迫切需要新的治疗方法，特别是对复发疾病的治疗。这个 神经母细胞瘤起始的标准模型假定分化受阻。我们认为，SNS因素 对SNS的生长和建立很重要的是神经母细胞瘤突变所选择的 支持肿瘤的生长和建立。此外，根据我们的初步调查结果，我们建议 一类被称为BET抑制剂的药物，被发现对神经母细胞瘤非常有效，部分 通过抑制重要的社交网络因子发挥作用。在这里，我们假设SNS细胞命运调节器，是 Nb对肿瘤的发生至关重要，并与Nb对BET抑制剂的敏感性有关。这两个目标是我们 在这项提案中将追求：目标1：细胞命运的调节者是NB中BET抑制的关键靶点吗？目标2： 细胞特性/发育阶段是否有助于原发NCC的恶变？对于第一次 目的我们将在神经母细胞瘤细胞系中确定BET抑制剂是否通过关闭关键的SNS起作用 各种因素。因此，我们将在这里：1)探讨BET抑制剂对SNS谱系因子表达的影响。2) 研究BET抑制剂JQ1对人NB细胞系BET蛋白BRD4结合的影响。 3)检测是否单独或联合沉默SNS谱系因子对人NB细胞生长的影响 LINES和NB肿瘤。4)确定谱系因子是否在JQ1不敏感的MSCV中反式表达 启动子在人NB细胞系和患者来源的异种移植瘤中传递对BET抑制剂的耐药性。 目标1将确定SNS血统因素是否对NB的维持是必要的。在目标2中，我们将询问是否 这些因子在神经母细胞瘤的发生中是必需的。为了测试第二个目标，我们将使用一个独特的系统 我们已经开发出将正常未成熟的SNS小鼠细胞NCCs转化为神经母细胞瘤的方法，以确定 重要的SNS因子是否参与了神经母细胞瘤的发生。对于第二个目标，我们将：1) 确定SNS核心因子是否阻止NCC分化为交感神经元。2)我们将确定是否 SNS核心转录因子在体外促进肿瘤的发生。3)确定SNS核心是否 转录因子单独或联合N-Myc促进NCC来源的NB肿瘤的形成 或1p36杂合性丢失，这是NB中的两个常见突变。总体而言，我们的工作将告诉我们是否 对SNS发展很重要的因素也会导致神经母细胞瘤。进一步提升我们的工程设计能力 来自未成熟SNS小鼠细胞的神经母细胞瘤独特地使我们能够阐明 神经母细胞瘤对BET抑制的敏感性，并发现导致这种疾病的新的基因因素。

Project Summary/Abstract: Neuroblastoma (NB) is the most common cancer of infancy and arises from immature cells of the sympathetic nervous system (SNS) that derive from neural crest stem cells (NCCs). NB is responsible for 15% of pediatric cancer deaths with the majority of high-risk NB patients eventually relapsing with resistant disease. Finding novel treatments, especially for relapse disease, is desperately needed for high-risk neuroblastoma. The standard model of neuroblastoma initiation assumes a block in differentiation. We propose that SNS factors important for the growth and establishment of the SNS are co-opted by neuroblastoma mutations to instead support the growth and establishment of the tumor. Further based on our preliminary findings we propose a class of drugs called BET inhibitors, which were found to be very effective against neuroblastoma, partially function by suppressing important SNS factors. Here, we hypothesize that SNS cell fate regulators, are critical for NB oncogenesis and responsible for NB sensitivity to BET inhibitors. The two aims we will pursue in this proposal: Aim 1: Are regulators of cell fate critical targets of BET inhibition in NB? Aim 2: Does cell identity/developmental stage contribute to malignant transformation of primary NCCs? For the first aim we will determine in neuroblastoma cell lines, if BET inhibitors work through turning off critical SNS factors. Thus, here we will: 1) Interrogate the impact of BET inhibitors on SNS lineage factors expression. 2) Investigate loss of the BET protein BRD4 binding in response to the BET inhibitor JQ1, in human NB cell lines. 3) Test if silencing SNS lineage factors individually or in combination interferes with growth of human NB cell lines and NB tumors. 4) Determine if lineage factors expressed in trans from the JQ1-insensitive MSCV promoter conveys resistance to BET inhibitors in human NB cell lines and patient derived xenograft tumors. Aim 1 will establish if SNS lineage factors are necessary for NB maintenance. In Aim 2, we will interrogate if these factors are functionally required for NB oncogenesis. To test the second aim we will use a unique system we have developed for turning normal immature SNS mouse cells, the NCCs, into neuroblastoma to determine if the important SNS factors can contribute to initiating neuroblastoma. For the second aim we will: 1) Determine if the SNS core factors block NCC differentiation into sympathetic neurons. 2) We will determine if the SNS core transcription factors promote tumorigenic behavior in vitro. 3) Determine if the SNS core transcription factors promotes the formation of NCC-derived NB tumors alone or in combination with N-Myc or 1p36 loss of heterozygosity, two common mutations in NB. Overall, our work will inform us of whether factors important for SNS development also contribute to neuroblastoma. Further our ability to engineer neuroblastoma tumors from immature SNS mouse cells uniquely enables us to illuminate novel mechanisms of neuroblastoma sensitivity to BET inhibition and discover new genetic contributors to this disease.