Ganglioside Regulation of NFkbeta and Apoptosis

神经节苷脂对 NFkbeta 和细胞凋亡的调节

• 批准号: 6321322
• 负责人: JAMES H FINKE
• 金额: $ 27.64万
• 依托单位: CLEVELAND CLINIC FOUNDATION
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-01 至 2006-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6321322
• 关键词: BCL2 gene /protein; CD95 molecule; T lymphocyte; apoptosis; clone cells; cysteine endopeptidases; cytokine receptors; enzyme activity; gangliosides; gene expression; human tissue; immunogenetics; mitochondria; nuclear factor kappa beta; transfection; tumor necrosis factor alpha

DESCRIPTION: The generation of T cell mediated immunity is often impaired in cancer patients. Our studies have shown that T cells from cancer patients display increased susceptibility to activation induced cell death (AICD) and also exhibit a defect in NFKB activation. These alterations may contribute to the immune dysfunction observed in T cells from these patients. Gangliosides, which are known to be overexpressed in certain tumor types, may be responsible for the suppression of NFKB and for the increased sensitivity of patient T cells to apoptosis. We and others have found that select gangliosides, including GD3, can sensitize normal human T cells to activation induced cell death (AICD). Although GD3 alone is not sufficient to induce apoptosis, it does inhibit expression of the anti-apoptotic genes clAP-1, cIAP-2 and TRAF-2. GD3 also inhibits NFkB activation, which appears to be due to stimulus-dependent degradation of Re1A . The induction of apoptosis and suppression of NFKB activity are caspase dependent, since both events are blocked by the caspase inhibitors Z-VAD-fmk and DEVD-frnk. Based on our findings, we hypothesize that immunity to tumors is diminished in cancer patients due to an imbalance between the intracellular levels of pro- versus anti-apoptotic proteins in T cells. Gangliosides including GD3 contribute to this imbalance by suppressing anti-apoptotic gene expression (cIAP/TRAF). The reduced expression of cIAPs/TRAFs allows increased caspase activity, leading to stimulus dependent RelA degradation, further depressing anti-apoptotic gene expression and inducing apoptosis. Aim 1 will define how GD3 regulates anti-apoptotic gene expression in T cells, and will assess the role of those molecules in protecting cells from NFKB inhibition and apoptosis. Initial experiments will determine the extent to which GD3 inhibits the anti-apoptotic gene expression that normally protects T cells from apoptosis. Experiments will also determine the mechanism by which GD3 alters the steady state levels of antiapoptotic genes, including clAPs, and TRAFs. Studies will also test the impact that overexpression of the antiapoptotic genes (cIAPs/TRAFs) has on RelA degradation, caspase activation, and sensitivity to AICD in GD3-treated T cells. Aim 2 will determine the mechanism by which GD3 inhibits NFKB in T cells, and the relationship of that inhibition to anti-apoptotic gene expression and to AICD. The role of GD3 in modulating caspase activity in T cells will be assessed by identifying which caspases are activated in response to GD3, T cell activation or both. The role these activated caspases play in degradation of ReIA will then be assessed by blocking select caspase function. In a final set of experiments, ReIA will be overexpressed to determine its protective effect on both GD3-inhibited antiapoptotic gene expression and in rendering cells resistant to AICD. These studies will provide insight into how gangliosides inhibit NFKB and survival gene expression, leading to increased T cell sensitivity to apoptosis.

描述：T细胞介导的免疫力的产生通常在以下情况下受损： 癌症患者。我们的研究表明癌症患者的T细胞 显示对活化诱导的细胞死亡（AICD）的敏感性增加， 也表现出NF κ B活化缺陷。这些变化可能有助于 在这些患者的T细胞中观察到的免疫功能障碍。神经节苷脂， 在某些肿瘤类型中过度表达， 对于NF κ B的抑制和对于患者T细胞的敏感性增加， 细胞凋亡。我们和其他人已经发现，选择神经节苷脂， 包括GD 3，可以使正常人T细胞对活化诱导的细胞 死亡（AICD）。虽然单独的GD 3不足以诱导细胞凋亡，但它可以诱导细胞凋亡。 抑制抗凋亡基因cIAP-1、cIAP-2和TRAF-2的表达。GD3 也抑制NFkB活化，这似乎是由于刺激依赖性 Re 1A的降解。细胞凋亡的诱导和NF κ B的抑制 活性是半胱天冬酶依赖性的，因为这两个事件都被半胱天冬酶阻断。 抑制剂Z-VAD-frnk和DEVD-frnk。根据我们的发现，我们假设 癌症患者对肿瘤的免疫力下降，这是由于 T细胞中促凋亡蛋白与抗凋亡蛋白的细胞内水平。 包括GD 3在内的神经节苷脂通过抑制 抗凋亡基因表达（cIAP/TRAF）。表达减少 cIAPs/TRAFs允许增加caspase活性，导致刺激依赖性 RelA降解，进一步抑制抗凋亡基因表达， 诱导细胞凋亡。目的1明确GD 3如何调控抗凋亡基因 在T细胞中的表达，并将评估这些分子的作用， 保护细胞免受NF κ B抑制和凋亡。初步实验将 确定GD 3抑制抗凋亡基因表达的程度 保护T细胞免于凋亡实验也将决定 GD 3改变抗凋亡蛋白稳态水平的机制 基因，包括clAP和TRAF。研究还将测试 抗凋亡基因（cIAPs/TRAFs）的过度表达对RelA 在GD 3处理的T细胞中降解，半胱天冬酶激活和对AICD的敏感性 细胞目的2将确定GD 3抑制T细胞中NF κ B的机制。 细胞，以及这种抑制与抗凋亡基因的关系 表达和AICD。GD 3对T细胞caspase活性的调节作用 细胞将通过鉴定哪些半胱天冬酶在应答中被激活来评估 GD 3、T细胞活化或两者。这些激活的半胱天冬酶在 然后通过阻断选择的半胱天冬酶功能来评估ReIA的降解。 在最后一组实验中，ReIA将被过表达以确定其 对GD 3抑制的抗凋亡基因表达和细胞凋亡的保护作用。 使细胞对AICD产生抗性。这些研究将提供洞察如何 神经节苷脂抑制NF-κ B和存活基因表达，导致T 细胞对凋亡的敏感性