Alternative NF-kB in Bone Microenvironment

描述(申请人提供)：破骨细胞(OCs)是唯一能够骨吸收的细胞，这是正常骨稳态和病理性骨丢失所必需的过程。在肿瘤转移到骨的背景下，招募的OCs不仅吸收骨，造成疼痛和骨折，而且改变微环境，促进肿瘤生长。OCs的吸收活性释放骨储存因子，为肿瘤细胞提供燃料；反过来，肿瘤细胞分泌增加OCs数量和活性的因子，为肿瘤介导的骨溶解创造一个正反馈循环。了解OC激活的机制及其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。核因子？B家族转录因子对OC的分化和细胞存活至关重要。在第一个资助阶段，我们确定经典的由p65/relA亚单位介导的NF-B通路对于OC前体的存活是重要的，但对分化不是重要的。相反，我们发现由NIK和RelB介导的另一种NF-B途径控制OC的分化。此外，在肿瘤介导的骨溶解的背景下，NIK或RelB的缺失减少了体内的病理性骨丢失。在缺乏RelB的情况下，OC分化的关键转录因子NFATc1和c-fos的表达减少，几个NFATc1靶基因的表达也减少，提示RelB可能通过NFATc1和c-fos调节破骨细胞的形成。最近，CIAP和TRAF3被认为是NIK和RelB的关键负调控因子。药物阻断cIAPs和阻断Nik/TRAF3的相互作用都会导致选择性的NF-B途径的结构性激活。我们的中心假设是，通过NIK和RelB激活替代的NF-B途径，控制NFATc1和c-fos的诱导，促进破骨细胞的形成，促进有利于骨质疏松和骨转移的骨微环境。这项研究利用遗传学和药理学的方法，探讨了在骨转移的背景下，Nik通路的结构性激活的影响，并将解决Nik及其下游效应因子RelB控制破骨细胞形成的转录机制。因此，我们的具体目标如下：目的1：确定CIAP拮抗剂治疗对骨转移患者骨微环境的影响。目的2：确定破骨细胞中NIK的结构性基因激活影响基础骨稳态和肿瘤介导的骨溶解的机制。目的：探讨NIK和RelB在卵巢癌分化过程中对NFATc1和c-fos转录调控中的作用。总体而言，该建议建立在最初的R01中的发现的基础上，即NIK下游的核因子-βB亚单位RelB对OC分化至关重要，并扩展了研究以了解这一效应的转录机制，为治疗抑制Nik/RelB途径以防止骨丢失提供了更强有力的知识基础。由于一类新开发的抗癌药物-CIAP拮抗剂-激活Nik/RelB途径，我们还将探讨该途径的激活对骨的影响，重点是骨转移模型。这些研究将对了解和预防这些NIK激活药物对骨完整性和骨转移的发生率/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.