Alternative NF-kB in Bone Microenvironment

描述（由申请人提供）：破骨细胞（OC）是唯一能够进行骨吸收的细胞，骨吸收是正常骨稳态和病理性骨丢失所需的过程。在肿瘤向骨转移的背景下，募集的OC不仅吸收骨，引起疼痛和骨折，而且改变微环境以促进肿瘤生长。OC的再吸收活性释放了为肿瘤细胞提供燃料的骨储存因子;反过来，肿瘤细胞分泌增加OC数量和活性的因子，为肿瘤介导的骨质溶解创造了正反馈回路。了解OC激活的机制以及它们如何将微环境改变为更有利于肿瘤生长的微环境，对于开发骨转移的治疗方法至关重要。 NF-？B家族转录因子对OC分化和细胞存活至关重要。在第一个资助期，我们确定，经典的NF-？p65/RelA亚基介导的B通路对OC前体细胞的存活很重要，但对分化不重要。相反，我们发现，替代NF-？B途径由NIK和RelB介导，控制OC的分化。此外，在肿瘤介导的骨质溶解的情况下，NIK或RelB的缺乏减少了体内病理性骨丢失。在不存在RelB的情况下，OC分化的关键转录因子NFATc 1和c-fos的表达减少，几种NFATc 1靶基因的表达也是如此，这表明RelB可能通过NFATc 1和c-fos调节破骨细胞生成。最近，cIAP和TRAF 3被认为是NIK的关键负调节因子，因此也是RelB的关键负调节因子。阻断cIAP的药物和NIK/TRAF 3相互作用的破坏都导致替代NF-？B途径。我们的中心假设是，激活的替代NF-？B通路通过NIK和RelB控制NFATc 1和c-fos的诱导，以增强破骨细胞生成并促进有利于骨质疏松和骨转移的骨微环境。该提案探讨了在骨转移的背景下，使用遗传学和药理学方法，这NIK通路的组成性激活的影响，并将解决NIK及其下游效应器RelB控制破骨细胞生成的转录机制。因此，我们的具体目标如下：目标1：确定cIAP拮抗剂治疗对基线和骨转移背景下骨微环境的药理学NIK激活作用。目标二：确定破骨细胞中NIK的组成性遗传激活影响基础骨稳态和肿瘤介导的骨溶解的机制。目的3：研究NIK和RelB在OC分化过程中对NFATc 1和c-fos转录调控的作用。总体而言，这一建议的基础上，在原来的R 01，RelB，NF-？NIK下游的B亚基对OC分化至关重要，并扩展了研究以了解这种作用的转录机制，为治疗性抑制NIK/RelB途径以防止骨丢失提供了更强的知识基础。由于一类新开发的抗癌药物-cIAP拮抗剂-激活NIK/RelB通路，我们还将探索激活该通路对骨的影响，重点是骨转移模型。这些研究对于了解和预防这些NIK激活药物对骨完整性和骨转移发生率/严重程度的潜在不良影响至关重要。

DESCRIPTION (provided by applicant): Osteoclasts (OCs) are the only cells capable of bone resorption, a process required for both normal bone homeostasis and pathological bone loss. In the context of tumor metastasis to bone, recruited OCs not only resorb bone, causing pain and fractures, but also change the microenvironment to encourage tumor growth. The resorptive activity of OCs releases bone-stored factors that fuel tumor cells; in turn, tumor cells secrete factors that increase the number and activity of OCs, creating a positive feedback loop for tumor-mediated osteolysis. Understanding the mechanisms of OC activation and how they change the microenvironment to one more favorable for tumor growth is critical for developing treatments for bone metastasis. The NF-?B family of transcription factors is critical for OC differentiation and cell survival. In the first funding period, we determined that the classical NF-?B pathway, mediated by the p65/RelA subunit, is important for OC precursor survival, but not differentiation. In contrast, we found that the alternative NF-?B pathway, mediated by NIK and RelB, controls OC differentiation. Furthermore, absence of either NIK or RelB reduces pathological bone loss in vivo in the context of tumor-mediated osteolysis. Expression of NFATc1 and c-fos, key transcription factors for OC differentiation, is diminished in the absence of RelB, as is the expression of several NFATc1 target genes, suggesting that RelB may modulate osteoclastogenesis via NFATc1 and c- fos. Recently, cIAP and TRAF3 have been implicated as critical negative regulators of NIK, and thus RelB. Drugs blocking cIAPs and disruption of NIK/TRAF3 interaction both result in constitutive activation of the alternative NF-?B pathway. Our central hypothesis is that activation of the alternative NF-?B pathway, via NIK and RelB, controls induction of NFATc1 and c-fos, to enhance osteoclastogenesis and promote a bone microenvironment conducive to osteoporosis and bone metastasis. This proposal explores the effects of constitutive activation of this NIK pathway in the context of bone metastasis, using genetic and pharmacologic approaches, and will address the transcriptional mechanism by which NIK and its downstream effector RelB control osteoclastogenesis. Thus, our specific aims are as follows: Aim 1: Determine the effect of pharmacologic NIK activation by cIAP antagonist treatment on the bone microenvironment at baseline and in the context of bone metastasis. Aim 2: Determine the mechanism by which constitutive genetic activation of NIK, in osteoclasts, affects basal bone homeostasis and tumor- mediated osteolysis. Aim 3: Determine the role of NIK and RelB in the transcriptional control of NFATc1 and c-fos during OC differentiation. Overall, this proposal builds on the findings in the original R01 that RelB, the NF-?B subunit downstream of NIK, is critical for OC differentiation, and extends the studies to understand the transcriptional mechanism for this effect, providing a stronger knowledge base for therapeutic inhibition of the NIK/RelB pathway to prevent bone loss. Because a newly developed class of drugs being developed as anti-cancer agents - the cIAP antagonists -- activates the NIK/RelB pathway, we will also probe the effects of activation of this pathway on bone, focusing on models of bone metastasis. These studies will be critical for understanding and preventing potential adverse effects of these NIK-activating drugs on bone integrity and the incidence/severity of bone metastasis.