TUMOR SUPPRESSOR RB FAMILY/PRB2 IN MEDULLOBLASTOMA

髓母细胞瘤中的肿瘤抑制因子 RB 家族/PRB2

• 批准号: 6825071
• 负责人: Antonio Giordano
• 金额: $ 25.98万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6825071
• 关键词: Papovaviridae; cell cycle proteins; cellular oncology; disease /disorder model; gene mutation; genetically modified animals; human tissue; immunocytochemistry; immunoprecipitation; laboratory mouse; medulloblastoma; molecular oncology; neoplastic growth; pathologic process; protein structure function; retinoblastoma protein; tissue /cell culture; tumor antigens; tumor suppressor genes; virus antigen; virus related neoplasm /cancer; western blottings

Project #2: Tumor Suppressor Rb Family/pRb2 in Medulloblastoma. The human neurotropic virus, JCV, is widespread in the human population and is the established etiologic agent of the fatal demyelinating disease of the central nervous system (CNS), Progressive Multifocal Leukoencephalopathy (PML). Like other papovaviruses, the JCV genome consists of circular double-stranded DNA that is separated into early and late coding sequences by the viral cell type-specific regulatory region. The viral early gene, T-antigen has important regulatory functions in orchestrating the viral lytic cycle and possesses the ability to interact with several important cellular proteins including the tumor suppressor protein, pRb pointing to the oncogenic potential of this virus. In support of this notion, earlier studies have revealed that JCV has the ability to induce neural origin tumors in several animal models. More recently, in collaboration with Dr. Khalili (Leader Project #1), a transgenic animal which developed tumor from external granular layer of cerebellun_ mimicking human medulloblastoma was created. The transgene which express T-antigen exhibited the ability to interact with pRb as well as p53 and that may de-regulate cell cycle pathway leading to evolution of tumor However, the underlying mechanism of transactivation of these two tumor suppressors, particularly pRb remains unknown. To better understand the molecular events involved in the genesis of T-antigen inducec medulloblastomas, we have developed cell lines from mouse tumors. Interestingly, similar to tumor tissue, not all cells expressed T-antigen, leading to speculation that as tumor cells develop, expression of T-antigen may no longer be needed and expression of T-antigen becomes silent. Loss of T-antigen expression which is concomitant with the lack of the hypophosphorylated form (active form) of pRb2/pl30 suggests that at the early stage of tumorigenesis, expression of T-antigen and its association with pRb2/pl30 functionally inactivates this protein, while at the latter stage of tumor development, the active form of pRb2/pl30 may be lost due to an increase in the level of phosphorylation of pRb2/pl30 and/or enhancement in its proteolytic degradation. The observation on murine medulloblastoma tumors is in accord with our earlier observation on human medulloblastoma that demonstrated detection of active pRb2/p 130 in JCV-associated medulloblastomas expressing T-antigen, but not in tumor cells lacking JCV and its early protein. As the biological function ot pRb2/pl30 is dictated at several stage, most notably through its partnership with p27 Kip1 and the E2F family, in this research proposal we will utilize the mouse model of medulloblastoma as well as a collection of well-characterized human medulloblastomas to decipher the molecular events involved in dysregulation o1 pRb2/pl30 during various stages of the cell cycle in vitro and during the course of tumor formation in the cerebellum of experimental animals. The outcome should provide useful information for therapeutic intervention.

项目#2：髓母细胞瘤中的肿瘤抑制因子 Rb 家族/pRb2。 人类嗜神经病毒 JCV 在人群中广泛传播，是中枢神经系统 (CNS) 致命性脱髓鞘疾病、进行性多灶性白质脑病 (PML) 的既定病原体。与其他乳多空病毒一样，JCV 基因组由环状双链 DNA 组成，该双链 DNA 被病毒细胞类型特异性调控区分为早期和晚期编码序列。病毒早期基因 T 抗原在协调病毒裂解周期中具有重要的调节功能，并具有与多种重要细胞蛋白相互作用的能力，包括肿瘤抑制蛋白 pRb 指向 该病毒的致癌潜力。为了支持这一观点，早期的研究表明 JCV 能够在多种动物模型中诱导神经源性肿瘤。最近，与 Khalili 博士（项目#1 领导者）合作，创造了一种转基因动物，它从小脑外部颗粒层发展出肿瘤，模仿人类髓母细胞瘤。表达T抗原的转基因表现出与pRb以及p53相互作用的能力，并且可能解除细胞周期途径的调节，从而导致肿瘤的进化。然而，这两种肿瘤抑制因子（特别是pRb）反式激活的潜在机制仍然未知。为了更好地了解 T 抗原诱导的髓母细胞瘤发生过程中涉及的分子事件，我们开发了来自小鼠肿瘤的细胞系。有趣的是，与肿瘤组织类似，并非所有细胞都表达T抗原，这导致人们推测，随着肿瘤细胞的发展，可能不再需要T抗原的表达，并且T抗原的表达变得沉默。 T抗原表达的丧失伴随着pRb2/pl30低磷酸化形式（活性形式）的缺乏，这表明在肿瘤发生的早期阶段，T抗原的表达及其与pRb2/pl30功能性的关联 使该蛋白质失活，而在肿瘤发展的后期，由于pRb2/p130磷酸化水平的增加和/或其蛋白水解降解的增强，pRb2/p130的活性形式可能会丢失。对小鼠髓母细胞瘤的观察结果与我们早期对人髓母细胞瘤的观察结果一致，即在表达 T 抗原的 JCV 相关髓母细胞瘤中检测到活性 pRb2/p 130，但在缺乏 JCV 及其早期蛋白的肿瘤细胞中未检测到活性 pRb2/p 130。由于 pRb2/pl30 的生物学功能是在几个阶段决定的，最显着的是通过其与 p27 Kip1 和 E2F 家族的伙伴关系，在本研究提案中，我们将利用髓母细胞瘤的小鼠模型以及一系列特征良好的人类髓母细胞瘤破译在体外细胞周期的各个阶段以及肿瘤形成过程中与 o1 pRb2/pl30 失调相关的分子事件实验动物的小脑。结果应该为治疗干预提供有用的信息。