PRODUCTION OF ALTERNATIVE FGF RECEPTOR FORMS IN TUMOR

肿瘤中替代性 FGF 受体形式的产生

基本信息

批准号：
2895316
负责人：
GILBERT J. COTE
金额：
$ 19.33万
依托单位：
UNIVERSITY OF TX MD ANDERSON CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
1995
资助国家：
美国
起止时间：
1995-07-01 至 2000-04-30
项目状态：
已结题

项目摘要

Malignant astrocytomas comprise the majority of central nervous system tumors in humans and are associated with a dismal prognosis despite the application of multimodality therapy. It is hypothesized that the genesis of these tumors occurs as a multistep progression from benign astrocytoma to anaplastic astrocytoma (AA), and finally, to glioblastoma multiforme (GBM). In this transformation from benign to malignant, numerous, and as of now, poorly understood cytogenetic and biochemical changes take place. Recently it was demonstrated that the progression of astrocytes from a benign to a malignant phenotype is accompanied by a change in the RNA processing of fibroblast growth factor receptor 1 (FGFR-1) gene. The level of a high affinity form of the FGFR-1 is dramatically elevated as a result altered expression and RNA splicing. The strong correlation between altered FGFR gene expression and astrocyte malignancy underscores the importance of understanding the role of FGFR in normal and malignant astrocyte cell growth. This proposal will test the hypothesis that alterations in FGFR RNA splicing contribute to abnormal growth of malignant astrocytes. The specific objectives of this proposal are 1) to develop a model system which mimics the FGFR-1 RNA processing pathways observed in normal brain glial cells and glioblastoma, 2) to use this model to identify sequence elements within the FGFR-1 precursor RNA which act to regulate splicing, 3) to use the regulatory sequence information to identify regulatory factors acting in trans, 4) to determine the relationship between trans-acting factor expression and transformation. Aim 1 will be accomplished by examining available astrocyte cell lines for the RNA splicing patterns of FGFR-1 transcripts. We will also express a chimeric FGFR-1 minigene in these cell lines to determine if its RNA transcripts are processed in a manner analogous to the endogenous gene. Aim 2 will be accomplished by introducing sequence alterations, deletions, substitutions and mutations, into the FGFR-1 minigene and determining their effect on RNA processing decisions. In Aim 3 we will develop an assay systems to be used to purify the factor(s) involved in regulation of FGFR-1 RNA splicing. Two approaches will be used: identification of RNA- binding proteins through sequence-specific interaction and use of expression cloning which functionally detects RNA splicing. Finally, in Aim 4 we will use cDNA sequence information derived from Aim 3 to measure the level of trans-acting factor expression in graded tumor samples. A further understanding of the mechanisms underlying the changes in FGFR-1 RNA processing in malignant astrocytomas may shed light on the transformation process in astrocytes and provide new targets for suppressing their growth.

恶性星形细胞瘤占中枢神经系统的大部分尽管人类肿瘤与不良预后相关多学科综合治疗的应用。据推测，起源这些肿瘤的发生是良性星形细胞瘤的多步进展间变性星形细胞瘤 (AA)，最后是多形性胶质母细胞瘤（GBM）。在这种从良性到恶性的转变中，数量众多，而且正如目前，人们对细胞遗传学和生化变化的了解还知之甚少。最近的研究表明，星形胶质细胞的进展是从良性到恶性表型伴随着 RNA 的变化成纤维细胞生长因子受体 1 (FGFR-1) 基因的加工。水平结果，高亲和力形式的 FGFR-1 显着升高表达和 RNA 剪接改变。之间的强相关性 FGFR 基因表达的改变和星形胶质细胞恶性肿瘤强调了了解 FGFR 在正常和恶性疾病中的作用的重要性星形胶质细胞生长。该提案将检验以下假设： FGFR RNA 剪接的改变导致异常生长恶性星形胶质细胞。该提案的具体目标是 1) 开发模拟 FGFR-1 RNA 加工途径的模型系统在正常脑胶质细胞和胶质母细胞瘤中观察到，2）使用此识别 FGFR-1 前体 RNA 内序列元件的模型调节剪接，3) 使用调节序列信息确定反式中作用的调节因素，4) 确定反式作用因子表达与转化之间的关系。目标 1 将通过检查可用的星形胶质细胞系来实现 FGFR-1 转录本的 RNA 剪接模式。我们也会表达一个这些细胞系中的嵌合 FGFR-1 小基因以确定其 RNA 是否转录本的处理方式与内源基因类似。目标 2 将通过引入序列改变、删除、取代和突变，进入 FGFR-1 小基因并确定它们对 RNA 加工决策的影响。在目标 3 中，我们将开发一个用于纯化参与调节的因子的测定系统 FGFR-1 RNA 剪接。将使用两种方法：鉴定 RNA- 通过序列特异性相互作用和使用来结合蛋白质表达克隆，可功能性检测 RNA 剪接。最后，在目标 4 我们将使用源自目标 3 的 cDNA 序列信息来测量分级肿瘤样本中反式作用因子的表达水平。一个进一步了解FGFR-1变化的机制恶性星形细胞瘤中的 RNA 加工可能有助于揭示星形胶质细胞的转化过程并为抑制它们的生长。