Somatic Cell Transfer to Model Medulloblastoma in Mice

描述（申请人提供）：髓母细胞瘤（MBs）是由儿童小脑神经祖细胞转化而产生的恶性脑肿瘤。积极的治疗方法结合手术、颅脊髓放射和化疗，5年生存率超过70%。治疗相关的神经毒性产生了一个关键的需求，即识别信号分子，可以靶向治疗，以最大限度地抑制肿瘤生长和最小化附带脑损伤。本研究的总体目标是利用RCAS/tv-a基因转移系统建立MB小鼠模型，作为分子靶向治疗MB的临床前测试平台。该实验系统使用来自禽白血病病毒的逆转录病毒载体（RCAS）和一种转基因小鼠系，该小鼠系在Nestin基因启动子的控制下表达逆转录病毒受体，该基因启动子在正常小脑发育期间在神经祖细胞中活跃。利用该系统，我们发现在小鼠出生后小脑中异位表达Sonic Hedgehog （Shh）可诱导MBs。此外，我们鉴定了属于不同功能类别的蛋白质，这些蛋白质与Shh合作以增强MB的形成。这些增强因子是(a) Myc癌蛋白，在正常发育过程中刺激神经祖细胞的增殖，(b) Bcl-2，有效抑制细胞凋亡，(c)胰岛素样生长因子- ii，通过激活磷脂酰肌醇3-激酶（PI3K）信号转导途径同时刺激细胞增殖并阻断细胞凋亡，以及(d)肝细胞生长因子（HGF），一种对肿瘤生长具有多效作用的生长因子。所有这些蛋白在人MBs中高表达的事实表明，它们在小鼠中的促肿瘤活性准确地反映了人类疾病的发病机制。此外，这些蛋白及其下游信号分子可以被认为是治疗靶点。具体目的1是确定抑制Shh信号传导是否与阻断HGF信号传导共同增强携带Shh？诱导MBs。特异性目的2是确定HGF单克隆抗体治疗对MB生长的抑制是否可以通过药物抑制HGF受体c- Met或下游PI3K信号传导而增强。具体目的3是使用RCAS/tv-a系统鉴定导致sh诱导的MBs转移到脊柱的基因。脊柱转移是MBs患者的一个非常不利的预后因素。这些目标的实现将有可能将机制发现转化为针对MB儿童的分子靶向治疗。

DESCRIPTION (provided by applicant): Medulloblastomas (MBs) are malignant brain tumors that arise by transformation of neural progenitor cells in the cerebellum in children. Aggressive treatment approaches combining surgery, craniospinal radiation, and chemotherapy result in 5-year survival rates exceeding 70%. Treatment- related neurotoxicity has created a critical need to identify signaling molecules that can be targeted therapeutically to maximize tumor growth suppression and minimize collateral brain damage. The overall objective of this research is to use a mouse model of MB, which we developed using the RCAS/tv-a gene transfer system, as a preclinical testing platform for molecular targeted MB therapy. This experimental system uses a retroviral vector (RCAS) derived from avian leukosis virus and a transgenic mouse line that expresses the retrovirus receptor under control of the Nestin gene promoter, which is active in neural progenitor cells during normal cerebellar development. Using this system, we showed that ectopic expression of Sonic Hedgehog (Shh) in the postnatal cerebellum induces MBs in mice. Furthermore, we identified proteins belonging to different functional classes that cooperate with Shh to enhance MB formation. These enhancing factors are (a) Myc oncoproteins, which stimulate proliferation of neural progenitors during normal development, (b) Bcl-2, which potently inhibits apoptosis, (c) insulin-like growth factor-II, which concomitantly stimulates proliferation and blocks apoptosis by activating the phosphatidylinositol 3-kinase (PI3K) signal transduction pathway, and (d) hepatocyte growth factor (HGF), a growth factor with pleiotropic effects on tumor growth. The fact that all of these proteins are highly expressed in human MBs indicates that their tumor-promoting activity in mice accurately reflects the pathogenesis of the human disease. Moreover, these proteins and their downstream signaling molecules can be considered therapeutic targets. Specific aim 1 is to determine whether inhibiting Shh signaling cooperates with blockade of HGF signaling to enhance treatment response in mice bearing Shh?induced MBs. Specific aim 2 is to determine whether MB growth suppression by HGF monoclonal antibody therapy can be enhanced by pharmacologic inhibition of the HGF receptor c- Met or downstream PI3K signaling. Specific aim 3 is to use the RCAS/tv-a system to identify genes that cause Shh-induced MBs to metastasize to the spine. Spinal metastasis is a highly unfavorable prognostic factor for patients with MBs. Achievement of these aims will make it possible to translate the mechanistic discoveries to molecular targeted therapies for children with MB.