Modifiers of Tumor Susceptibility in Murine Neuroblastoma

神经母细胞瘤是一种起源于周围神经嵴的肿瘤，是儿童期常见的致死性肿瘤，其转录因子MYCN扩增频繁发生，并与肿瘤的进展有关。 疾病我们通过在酪氨酸羟化酶（TH）启动子的控制下，将MYCN转基因定向表达到外周神经嵴，产生了一种高危神经母细胞瘤的转基因小鼠模型。 遗传分析确定了人类和小鼠肿瘤之间的保守遗传变化，并认为TH-MYCN转基因小鼠代表了儿童神经母细胞瘤的重要遗传模型。我们推测，在小鼠中导致神经母细胞瘤形成的额外遗传和表观遗传病变将与儿童神经母细胞瘤相关的基因中。本申请的长期目标是鉴定鼠和人神经母细胞瘤中的遗传和表观遗传变化。在这项研究中鉴定的基因可能揭示与人类MYCN扩增的神经母细胞瘤相关的新机制和途径，最终导致新的治疗靶点。不同品系的小鼠对肿瘤的易感性不同。FVB/N品系中的TH-MYCN转基因小鼠不发生肿瘤，129/SvJ品系中的几乎所有转基因小鼠在4月龄时死于肿瘤，129/SvJ FVB/N F1小鼠显示4%的肿瘤发生率。这些观察结果表明，种系修饰基因的结构或表观遗传变化在菌株之间不同，与Mycn相互作用，并构成菌株之间易感性差异的基础。这些菌株特异性差异提供了一个关键的资源，以确定在小鼠和人类神经母细胞瘤重要的二级遗传和表观遗传事件。我们建议动员一个强大的插入诱变剂，脊椎动物睡美人（SB）转座子，并结合联合收割机使用体细胞插入诱变与比较基因组杂交和修饰遗传学，以确定影响小鼠神经母细胞瘤肿瘤易感性的基因。目的1利用SB转座子插入突变技术加速F1代小鼠肿瘤基因突变，提高肿瘤的转移率。Aim 2应用基于数组的比较 基因组杂交来表征F1小鼠中自发和转座子诱导的肿瘤中的拷贝数异常，表征品系特异性甲基化模式，并鉴定肿瘤中的表观遗传变化。目的3在体外验证和表征候选基因 以及体内，基于人神经母细胞瘤中特定候选基因的参与进行优先级排序。

Neuroblastoma, a tumor of peripheral neural crest origin, is a common and lethal tumor of childhood.Amplification of the transcription factor MYCN occurs frequently in this tumor, and correlates with advanced disease. We generated a transgenic mouse model for high-risk neuroblastoma by directing expression of a MYCN transgene to the peripheral neural crest, under control of the Tyrosine Hydroxylase (TH) promoter. Genetic analyses identified conserved genetic changes between human and murine tumors, and argue that mice transgenic for TH-MYCN represent an important genetic model for childhood neuroblastoma. We hypothesize that the additional genetic and epigenetic lesions which contribute to neuroblastoma formation in the mouse will be in genes relevant to neuroblastoma in children. The long term objective of this application is to identify genetic and epigenetic changes in murine and human neuroblastoma. Genes identified in this study may reveal novel mechanisms and pathways relevant to human MYCN-amplified neuroblastoma, ultimately leading to novel therapeutic targets. Strains of mice differ in susceptibility to tumors. Mice transgenic for TH-MYCN in strain FVB/N do not develop tumors, nearly all transgenic mice in strain 129/SvJ die of tumors by 4 months of age, and 129/SvJ FVB/N F1 mice show 4% penetrance. These observations suggest that structural or epigenetic changes in germ line modifier genes differ between strains, interact with Mycn, and underlie the differences in susceptibility between strains. These strain-specific differences provide a critical resource to identify secondary genetic and epigenetic events important in both murine and human neuroblastoma. We propose to mobilize a powerful insertional mutagen, the vertebrate Sleeping Beauty (SB) transposon, and to combine use of somatic insertional mutagenesis with comparative genomic hybridization and modifier genetics to identify genes that influence susceptibility to tumors in murine neuroblastoma. Aim 1 uses SB transposon-based insertional mutagenesis to accelerate oncogenic mutations and to increase penetrance of tumors in F1 mice. Aim 2 applies array-based comparative genomic hybridization to characterize copy-number abnormalities in both spontaneous and transposon-induced tumors in F1 mice, to characterize strain-specific methylation patterns, and to identify epigenetic changes in tumors. Aim 3 validates and characterizes candidate genes in-vitro and in-vivo, prioritizing based on the involvement of specific candidate genes in human neuroblastoma.