Cytokine Signaling in Glioblastoma Cells

胶质母细胞瘤细胞中的细胞因子信号传导

• 批准号: 6574428
• 负责人: SAIKH JAHARUL HAQUE
• 金额: $ 33.15万
• 依托单位: CLEVELAND CLINIC FOUNDATION
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-01 至 2004-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6574428
• 关键词: apoptosis; athymic mouse; autocrine; biological signal transduction; cell line; cell proliferation; cellular pathology; clinical research; ecdysone; gene expression; glioblastoma multiforme; hormone regulation /control mechanism; human tissue; interleukin 13; interleukin 4; interleukin 6; molecular pathology; neoplasm /cancer genetics; polymerase chain reaction; tissue /cell culture; transcription factor; xenotransplantation

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to understand cytokine-mediated regulation of proliferation and apoptosis of malignant glioblastoma cells. Glioblastoma multiforme (GBM) is the most common and malignant form of primary brain tumors, with an average survival of less than a year. GBM arises from a complex series of molecular events that include inactivation of tumor suppressor genes as well as overexpression and activation of proto-oncogenes. Consequently GBM cells become highly proliferative and resistant to apoptosis. The latent transcription factor Stat3, which is activated by IL-6-family of cytokines and other growth factors induces the expression of genes that are responsible for the suppression of apoptosis in a variety of human cancer cells. Although GBM cells secrete IL-6 and respond to it, little is known about the role of Stat3 activation in the regulation of apoptosis in GBM cells. We found that Stat3 is constitutively activated by an autocrine action of IL-6 in GBM tumor tissues and GBM cell lines. Inhibition of Stat3 activation by the Jak-specific tyrosine kinase inhibitor AG490 reduces steady state levels of anti-apoptotic proteins Bcl-2, BcI-XL and Mcl-1, and induces apoptosis in GBM cells. In contrast, AG490 does not induce apoptosis in normal human astrocytes. Interestingly, Stat3 is activated by IL-4 in GBM cells that is in part, attributable to the expression of IL-13Ra2, a decoy receptor for IL-13. IL-4 normally activates Stat6 but not Stat3 by signaling through the classical Jak-Stat pathway, and produces growth arrest in normal human astrocytes and low-grade gliomas that do not express IL-13Ra2. In consideration of these observations, we hypothesize that (i) constitutive activation of Stat3 via an autocrine action of IL-6, provides survival signal in GBM cells by inducing the expression of anti-apoptotic genes, and (ii) IL-4 induces further activation of Stat3 in these cells via an IL-13Ra2-dependent novel mechanism. To test these hypotheses we will pursue the following specific aims. 1. To define the role of activated Stat3 in the survival of GBM cells we will: (a) Express a dominant negative mutant Stat3 (DNStat3) and the suppressor of cytokine signaling (SOCS)-I in GBM cells via an ecdysone-inducible system, and determine their apoptotic response in vitro, and (b) Determine the effects of Stat3 inactivation by DNStat3, SOCS-1 and AG490 on the growth of intracranial and subcutaneous transplants of GBM cell lines in rodent brain tumor models. 2. To identify cellular and molecular mechanisms underlying the IL-4-mediated activation of Stat3 in IL-13Ra2-expressing glioma cells we will: (a) Determine if IL-13Ra2 expression level parallels the malignancy grade of glioma and is associated with aberrant Stat activation by IL-4, and (b) Define the role of IL-13Ra2 in the regulation of IL-4-dependent signal transduction in GBM cells. This investigation will define cellular and molecular mechanisms underlying Stat3-mediated survival of GBM cells, and thus significantly advance our current understanding of the molecular pathobiology of GBM, and importantly will lead to the development of novel therapies for this deadly disease.

描述（由申请人提供）：该提案的长期目标是了解细胞因子介导的恶性胶质母细胞瘤细胞增殖和凋亡的调节。多形性胶质母细胞瘤（GBM）是原发性脑肿瘤中最常见和最恶性的形式，平均生存期不到一年。 GBM 源于一系列复杂的分子事件，包括肿瘤抑制基因的失活以及原癌基因的过度表达和激活。因此，GBM 细胞变得高度增殖并抵抗细胞凋亡。潜在转录因子 Stat3 被细胞因子 IL-6 家族和其他生长因子激活，诱导多种人类癌细胞中负责抑制细胞凋亡的基因表达。尽管 GBM 细胞分泌 IL-6 并对其做出反应，但人们对 Stat3 激活在 GBM 细胞凋亡调节中的作用知之甚少。我们发现 Stat3 在 GBM 肿瘤组织和 GBM 细胞系中被 IL-6 的自分泌作用组成型激活。 Jak 特异性酪氨酸激酶抑制剂 AG490 对 Stat3 激活的抑制会降低抗凋亡蛋白 Bcl-2、Bcl-XL 和 Mcl-1 的稳态水平，并诱导 GBM 细胞凋亡。相反，AG490 不会诱导正常人星形胶质细胞凋亡。有趣的是，Stat3 在 GBM 细胞中被 IL-4 激活，部分归因于 IL-13Ra2（IL-13 的诱饵受体）的表达。 IL-4 通常通过经典 Jak-Stat 途径发出信号激活 Stat6，但不激活 Stat3，并在正常人星形胶质细胞和不表达 IL-13Ra2 的低级别神经胶质瘤中产生生长停滞。考虑到这些观察结果，我们假设（i）通过 IL-6 的自分泌作用组成型激活 Stat3，通过诱导抗凋亡基因的表达在 GBM 细胞中提供生存信号，以及（ii）IL-4 进一步诱导通过 IL-13Ra2 依赖性新机制激活这些细胞中的 Stat3。为了检验这些假设，我们将追求以下具体目标。 1. 为了定义激活的 Stat3 在 GBM 细胞存活中的作用，我们将： (a) 通过蜕皮激素诱导系统在 GBM 细胞中表达显性失活突变体 Stat3 (DNStat3) 和细胞因子信号传导抑制因子 (SOCS)-I ，并确定它们的体外凋亡反应，以及（b）确定 DNStat3、SOCS-1 和 AG490 灭活 Stat3 对颅内和在啮齿动物脑肿瘤模型中进行 GBM 细胞系的皮下移植。 2. 为了确定表达 IL-13Ra2 的神经胶质瘤细胞中 IL-4 介导的 Stat3 激活的细胞和分子机制，我们将： (a) 确定 IL-13Ra2 表达水平是否与神经胶质瘤的恶性程度平行并与异常相关。 IL-4 的 Stat 激活，以及 (b) 定义 IL-13Ra2 在 GBM 细胞中 IL-4 依赖性信号转导调节中的作用。这项研究将定义 Stat3 介导的 GBM 细胞存活的细胞和分子机制，从而显着推进我们目前对 GBM 分子病理学的理解，重要的是，将导致针对这种致命疾病的新疗法的开发。