PROTEIN KINASE C SIGNALING PATHWAYS IN BRAIN TUMORS

这项研究的总体目标是确定 利用蛋白激酶C(PKC)作为分子治疗靶点 抑制多形性胶质母细胞瘤(GM)的生长 成人脑瘤。转基因对所有的治疗方法都是高度难治的 与正常脑组织不同的是，GM AS的高比例 随着GM来源的细胞系表现出高水平和活性的 PKCalpha。我们发现反义PKCalpha基因的表达 抑制血清依赖的体外生长和体内致瘤性 提示PKCalpha在GM细胞株U-87中起到积极的调节作用 在这些细胞的增殖中起着重要作用。许多初级和成熟的通用汽车 细胞系过表达生长因子受体，如PDGF和EGF，并 表现出增长的自分泌机制，这似乎与 高水平的PKC。因此，这项提案将解决监管 PKCalpha和其他PKC亚型在生长因子依赖中的作用 GM的自分泌生长，并使用PKC作为分子靶点 心理治疗。该提案的具体目标将确定：1) 特定的PKC亚型是转录调控的还是转录后调控的 通过生长因子在GM细胞中转录，2)肿瘤是否生长 可以被PKC异构体特异的反义c DNA或 体内外反义寡核苷酸(ODN)，3) 生长因子信号通路的组成部分受以下因素的影响 抑制特定的PKC亚型，反之，抑制哪些PKC亚型 受生长因子受体酪氨酸磷酸化的调节 酪氨酸激酶或受体相关的酪氨酸激酶，以及4)顺式 PKCalpha和其他PKC亚型中的调节元件被发现 在GM细胞中被生长因子转录上调。 初步研究表明，PKCalpha表达上调 在U-87细胞中通过PDGF-BB转录，但不通过PDGF-AA转录。5‘- 未翻译的PKCalpha基因组序列已从一名人类 基因组文库，并将用于确定转录起始 Site(S)和鉴定启动子中的调控元件 区域。这些研究将确定PKC作为一种选择性的 GM的治疗靶点及其在生长因子依赖性肿瘤中的作用 成长。

The broad objective of this study is to determine the efficacy of utilizing protein kinase C (PKC) as a molecular therapeutic target for inhibiting the growth of glioblastoma multiforme (GM), the most common brain tumor in adults. GM is highly refractory to all therapeutic modalities, and unlike normal brain tissue, a high percentage of GM as well as GM-derived cell lines demonstrate high levels and activity of PKCalpha. We have found that expression of the antisense PKCalpha cDNA attenuates serum-dependent growth in vitro and the tumorigenicity in vivo of GM cell line U-87, suggesting that PKCalpha plays a positive regulatory role in the proliferation of these cells. Many primary and established GM cell lines overexpress growth factor receptors such as PDGF and EGF and exhibit an autocrine mechanism of growth, which appears to be related to high levels of PKC. Therefore, this proposal will address the regulatory role of PKCalpha and other PKC isoforms in the growth factor-dependent autocrine growth of GM, and the use of PKC as a molecular target for therapy. The Specific Aims of this proposal will be to determine: 1) whether specific PKC isoforms are regulated transcriptionally or post- transcriptionally in GM cells by growth factors, 2) whether tumor growth can be selectively blocked by a PKC isoformspecific antisense cDNA or an antisense oligodeoxynucleotide (ODN) in vitro and in vivo, 3) which components of the growth factor signaling pathway are affected by inhibition of specific PKC isoforms, and conversely, which PKC isoforms are modulated by tyrosine phosphorylation by growth factor receptor tyrosine kinases or receptor-associated tyrosine kinases, and 4) the cis regulatory elements in PKCalpha and other PKC isoforms which are found to be transcriptionally upregulated in GM cells by growth factors. Preliminary studies have shown that PKCalpha is upregulated transcriptionally in U-87 cells by PDGF-BB, but not by PDGF-AA. The 5'- untranslated genomic sequence of PKCalpha has been cloned from a human genomic library and will be used to determine the transcription start site(s) and to characterize the regulatory elements in the promoter region. These studies will determine the importance of PKC as a selective therapeutic target in GM and its role in growth factor-dependent tumor growth.