Regulation, integration and impact of NFkB-signaling within the oncogenic signaling network in multiple myeloma

多发性骨髓瘤致癌信号网络中 NFkB 信号的调节、整合和影响

• 批准号: 144836417
• 负责人: Dr. Daniela Siegmund
• 金额: --
• 依托单位: Medizinische Klinik und Poliklinik II
• 依托单位国家: 德国
• 项目类别: Clinical Research Units
• 财政年份: 2009
• 资助国家: 德国
• 起止时间: 2008-12-31 至 2017-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/144836417?language=en
• 关键词: Regulation; integration; impact; NFkB; signaling

In the first funding period we found that the NFκB system is far from being maximally activated in myeloma cells and its activity could consequently be boosted exogenously by stimulation of TNF receptors or treatment with SMAC mimetics. Noteworthy, such exogenous stimulation of the NFκB system showed no major protective effect against various drugs targeting oncogenic pathways and even enhanced sensitivity for the endogenous apoptosis inducer CD95L by upregulation of its receptor CD95. We also found that individual as well as dual inhibition of the classical and alternative NFκB pathways using the IKK2-specifc inhibitors TPCA1, the NEDD8-activating enzyme inhibitor MLN4924 or IKK1 and IKK2 knockdown elicited no or a rather mild cytotoxic effect in short term assays (12-48 h). The myeloma-supporting activity of stromal cells, however, was strongly reduced by inhibitors of NFκB signaling. Thus, the NFκB system appears less important for myeloma cell survival as initially anticipated but could be of crucial relevance for myeloma cells beyond sole apoptosis protection and might further be required to maintain a myeloma compatible microenvironment.Based on these results, we now want to address the following four issues: First, in view of our own findings on the complex regulatory role of the two TNF receptors TNFR1 and TNFR2 on extrinsic apoptosis of myeloma cells and clinical evidence arguing for relevance of the TNF-TNF receptor system in MM, we want to evaluate in more detail the therapeutic potential of TNF and TNF receptor targeting reagents in MM. For this, we will analyze in cooperation with project Z1 treatment regimes in syngeneic MM models that allow selective exogenous stimulation of TNFR1 or TNFR2 with and without concomitant blockade of endogenous TNF. Most NFκB-related mutations affect molecules that act upstream of NIK and the IKKs. Because these molecules also act outside the NFκB system, we will secondly address the possibility that such NFκB-independent functions contribute to the MM phenotype. Specifically, we will investigate whether in myeloma cells crosstalk between NIK and/or IKK1 with Wnt, Notch and STAT3 signaling exists. Thirdly, many of the NFκB-related mutations found in MM affect components or inducers of the TRAF2-cIAP1/2 complex, which not only inhibits alternative NFκB signaling but also stimulates classical NFκB signaling and moreover inhibits the caspase-8 activating ripoptosome. Therefore, we will evaluate in vitro and in vivo whether MM cells are particularly prone to the therapeutic effects of the ripoptosome activating SMAC mimetic BV6. Finally, to identify in an unbiased fashion NFκB-dependent and -independent IKK/NIK-regulated targets, SILAC/mass spectrometry screens will be performed in co-operation with project Z4N using various inhibitors targeting distinct stages of NFκB signaling.

在第一个资助期，我们发现NFκB系统在骨髓瘤细胞中远未被最大限度地激活，因此可以通过刺激TNF受体或用SMAC模拟物处理来外源性地增强其活性。值得注意的是，NFκB系统的这种外源性刺激对靶向致癌途径的各种药物没有明显的保护作用，甚至通过上调其受体CD 95来增强内源性凋亡诱导剂CD 95 L的敏感性。我们还发现，使用IKK 2特异性抑制剂TPCA 1、NEDD 8激活酶抑制剂MLN 4924或IKK 1和IKK 2敲低对经典和替代NFκB通路进行单独抑制以及双重抑制，在短期试验（12-48 h）中未引起或引起相当轻微的细胞毒性效应。然而，NFκB信号传导抑制剂强烈降低了基质细胞的骨髓瘤支持活性。因此，NFκB系统对骨髓瘤细胞存活的重要性似乎不如最初预期的那么重要，但可能与骨髓瘤细胞的重要相关性超出了单独的凋亡保护，并且可能进一步需要维持骨髓瘤相容的微环境。基于这些结果，我们现在想要解决以下四个问题：首先，鉴于我们自己对两种TNF受体TNFR 1和TNFR 2对骨髓瘤细胞外源性凋亡的复杂调节作用的发现以及支持TNF-TNF受体系统与MM相关性的临床证据，我们希望更详细地评估TNF和TNF受体靶向试剂在MM中的治疗潜力。为此，我们将与项目Z1合作分析同基因MM模型中的治疗方案，该治疗方案允许选择性外源性刺激TNFR 1或TNFR 2，同时阻断或不阻断内源性TNF。大多数NFκ B相关突变影响NIK和IKK上游的分子。由于这些分子也在NFκB B系统外发挥作用，我们将在第二部分讨论这种NFκ B非依赖性功能对MM表型的作用。具体而言，我们将研究在骨髓瘤细胞中NIK和/或IKK 1与Wnt、Notch和STAT 3信号传导之间是否存在串扰。第三，MM中发现的许多NFκ B相关突变影响TRAF 2-cIAP 1/2复合物的组分或诱导物，其不仅抑制替代NFκB信号传导，而且刺激经典NFκB信号传导，并且还抑制caspase-8活化核糖体。因此，我们将在体外和体内评估MM细胞是否特别倾向于ripoptosome激活SMAC模拟物BV 6的治疗效果。最后，为了以无偏倚的方式鉴定NFκ B依赖性和非依赖性IKK/NIK调节靶点，将与项目Z4 N合作，使用靶向NFκB信号传导不同阶段的各种抑制剂进行SILAC/质谱筛选。