Targeting the ubiquitin-proteasome pathway to reverse catabolic action of PTH in bone

项目摘要/摘要 每天间歇性注射甲状旁腺激素(PTH)可促进骨形成，而持续注射 甲状旁腺激素引起骨吸收，限制了其治疗价值。了解分子机制， 促进甲状旁腺激素有益的合成代谢作用和有问题的不良反应是改善 以甲状旁腺素为基础的治疗效果。泛素-蛋白酶体途径在 调控骨代谢。1型甲状旁腺激素受体(PTHR)对短暂的 甲状旁腺素暴露和甲状旁腺素持续治疗可下调成骨细胞的甲状旁腺素受体。这是未知的 甲状旁腺激素引起的持续性骨丢失是否通过泛素-蛋白酶体途径介导。在研究中 在NIAMS R03拨款的支持下，我们确定持续的PTH治疗诱导PTHR泛素化 和降解，从而抑制成骨细胞分化，促进破骨细胞吸收活性。在……里面 此外，PTH激活了多条信号通路，但并非所有信号通路都是合成代谢的。来自其他公司的最新数据 我们的团队已经证明了β-连环蛋白与PTHR羧基末端区域相互作用，并 将PTHR信号从Gs/cAMP切换到GQ/PLC激活。此外，我们的初步数据显示， Ixazomib是新近批准的一种毒性较小的口服蛋白酶体抑制剂，能够阻断甲状旁腺素诱导的持续性 PTHR蛋白酶体降解和反向PTHR信号开关通过将β-连环蛋白从PTHR中解离出来。 基于这些发现，我们假设ixazomib能够转换持续的分解代谢效应 PTH通过阻止PTHR的降解和解离β-连环蛋白而发挥合成代谢的作用。我们的目标是 建议的研究是：1)建立抑制PTHR下调和PTHR的证据原则 与β-连环蛋白的相互作用使持续的PTHR激活在促进骨形成方面更有效； 和2)产生重要的临床前数据，评估ixazomib在我们的 甲状旁腺素介导的持续性骨丢失的小鼠模型。为了检验这一假说，我们制定了三个具体目标 并采用独立和互补的战略，实现这些目标。Aim 1将详细说明ixazomib如何 挽救持续的甲状旁腺素诱导的甲状旁腺素分解代谢信号。在目标2中，我们将确定ixazomib 在体内将持续的甲状旁腺素分解代谢作用转化为骨合成代谢。目标3将通过以下方式描述机制 其中Ixazomib在体外可逆转持续甲状旁腺激素的骨代谢作用。圆满完成 拟议的研究将极大地促进我们对促进/限制甲状旁腺激素影响的机制的理解。 骨形成，并提供强有力的基础和临床前数据，将阐明更有效的途径 骨质疏松症治疗。

Project Summary/Abstract Intermittent parathyroid hormone (PTH) by daily injection increases bone formation, whereas continuous PTH causes bone resorption and limits its therapeutic value. Understanding the molecular mechanisms that promote both the beneficial anabolic PTH actions and the problematic adverse effects is critical to improving the therapeutic efficacy of PTH-based treatments. The ubiquitin-proteasome pathway plays an important role in regulating and controlling bone metabolism. The type 1 PTH receptor (PTHR) desensitizes in response to brief exposure to PTH and sustained treatment with PTH downregulates the PTHR in osteoblasts. It is unknown whether continuous PTH-caused bone loss is mediated through the ubiquitin-proteasome pathway. In studies supported by an NIAMS R03 grant, we determined that continuous PTH treatment induces PTHR ubiquitination and degradation, thereby inhibiting osteoblast differentiation and promoting osteoclast resorptive activity. In addition, PTH activates multiple signaling pathways but not all of them are anabolic. Recent data from others and our group have demonstrated that beta-catenin interacts with the PTHR carboxyl-terminal region and switches PTHR signaling from Gs/cAMP to Gq/PLC activation. Furthermore, our preliminary data show that ixazomib, a newly approved oral proteasome inhibitor with less toxicity, is able to block continuous PTH-induced PTHR proteasomal degradation and reverse PTHR signaling switch by dissociating beta-catenin from the PTHR. Based on these findings, we hypothesize that ixazomib is capable of converting the catabolic effect of continuous PTH to an anabolic effect by blocking PTHR degradation and dissociating beta-catenin. The goals of our proposed studies are to: 1) establish the proof-of-principle that inhibition of PTHR downregulation and PTHR interaction with beta-catenin renders continuous PTHR activation more effective in promoting bone formation; and 2) generate important pre-clinical data assessing the efficacy, safety, and side effects of ixazomib in our murine model of continuous PTH-mediated bone loss. Three specific aims are developed to test this hypothesis and achieve these goals, employing independent and complementary strategies. Aim 1 will detail how ixazomib rescues continuous PTH-induced PTHR catabolic signaling. In Aim 2, we will establish whether ixazomib converts continuous PTH catabolic effect to bone anabolism in vivo. Aim 3 will characterize mechanisms by which ixazomib reverses the osteocatabolic effect of continuous PTH in vitro. Successful completion of the proposed research will greatly advance our understanding of the mechanisms that promote/limit PTH effects on bone formation, and provide strong basic and preclinical data that will clarify the path to a more effective osteoporosis treatment.