一群新的基因组不稳定颗粒神经元前体细胞在Ptc1杂合性缺失致髓母细胞瘤中的作用研究

杂合性缺失（LOH）与Ptc1基因缺陷所致小脑髓母细胞瘤（MB）密切相关，但LOH的易感细胞及产生机制不明。申请人在小脑外颗粒层（EGL）中新发现一群存在DNA损伤修复障碍的前体细胞（NEPs），与EGL中其他细胞被大量杀伤不同，放射辐照能刺激NEPs增殖，而胞内SHH通路的异常活化会使NEPs具有更高的成瘤性。放射辐照促进Ptc1基因缺陷杂合小鼠（与人MB发病相似度最高）MB发生率显著升高符合NEPs的表现，我们推测NEPs是自发MB的主要肿瘤来源细胞，而DNA损伤修复障碍是其LOH发生及成瘤的原因。本项目拟以上述小鼠为对象，追踪成瘤过程中NEPs及LOH的状态，比较辐照诱发与自发MB成瘤中LOH发生的异同，筛选并探讨DNA损伤修复基因Rad51ap1、p53等对LOH及MB发生的影响。本项目的实施将加深MB发生机制的认识，将为以LOH为靶点的早期MB治疗策略研究提供新的思路。

Medulloblastoma (MB) is the most common malignant brain tumor in children. Approximately 30% of MB cases have mutations in components of the sonic hedgehog (Shh) pathway. As a receptor for Shh, patched (Ptc) serves as an antagonist for this pathway. Mice with homozygous Ptc mutation are embryonically lethal, but Ptc heterozygous mice (Ptc+/- mice) survive and ~15% of them develop MB after 3 months of age. Interestingly, pre-neoplastic lesions are found in the cerebella of 60% of Ptc+/- mice at 6 weeks. Majority of pre-neoplastic cells finally differentiate, and a proportion of them proceed to give rise to MB. It is well established that expression of wild-type Ptc allele is repressed in both preneoplastic lesions and end-stage tumors in Ptc+/- mice. However, the mechanism underlying the silencing of wild-type Ptc allele in pre-neoplastic cells is still unknown. We have recently identified a rare population of neuronal progenitors in mouse developing cerebellum that expresses Nestin. Although Nestin is widely considered a marker for multipotent stem cells, these Nestin-expressing progenitors (NEPs) are committed to the granule neuron lineage. Unlike conventional GNPs, which reside in the outer EGL and proliferate extensively, NEPs reside in the deep part of the EGL and are quiescent. Expression profiling revealed that NEPs are distinct from GNPs and, in particular, express markedly reduced levels of genes associated with DNA repair. Consistent with this, upon aberrant activation of Sonic hedgehog (Shh) signaling, NEPs exhibited more severe genomic instability and gave rise to tumors more efficiently than GNPs.In this study, we propose to determine the important role of NEPs in the tumorigenesis of Ptc+/- MB.Upon completion of this project,we will elucidate the important role of LOH-induced Ptc deficiency in NEPs during MB development of, which will shed light on the molecular basis of MB tumorigenesis. Our studies will demonstrate that NEPs is a important source of MB cells and LOH represent the essential property of pre-neoplastic cells, and could be used to design early stage approaches for tumor treatment and prevention.