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

项目概要/摘要 每日注射间歇性甲状旁腺激素（PTH）可增加骨形成，而连续性 甲状旁腺素导致骨吸收，限制了其治疗价值。了解分子机制， 促进有益的合成代谢PTH作用和有问题的副作用对于改善 基于PTH的治疗的疗效。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）在短暂的 暴露于PTH和用PTH持续治疗下调成骨细胞中的PTHR。尚不清楚 持续PTH引起的骨丢失是否通过泛素-蛋白酶体途径介导。研究中 在NIAMS R 03基金的支持下，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和促进破骨细胞再吸收活性。在 此外，PTH激活多种信号通路，但并非所有通路都是合成代谢通路。其他人的最新数据 我们的研究小组已经证明β-连环蛋白与PTHR羧基末端区域相互作用， 将PTHR信号传导从Gs/cAMP转换为Gq/PLC激活。此外，我们的初步数据显示， Ixazomib是一种新批准的口服蛋白酶体抑制剂，毒性较低，能够阻断持续性PTH诱导的 PTHR蛋白酶体降解和通过从PTHR解离β-连环蛋白来逆转PTHR信号转导开关。 基于这些发现，我们假设ixazomib能够将持续性代谢的分解代谢作用 PTH通过阻断PTHR降解和解离β-连环蛋白来发挥合成代谢作用。我们的目标 提出的研究是：1）建立原理证明，抑制PTHR下调和抑制PTHR下调， 与β-连环蛋白的相互作用使得持续的PTHR激活在促进骨形成中更有效; 和2）生成重要的临床前数据，评估ixazomib在我们研究中的疗效、安全性和副作用。 持续PTH介导的骨丢失的鼠模型。三个具体的目标是开发来测试这一假设 并实现这些目标，采用独立和互补的战略。目标1将详细说明ixazomib 拯救持续的PTH诱导的PTHR分解代谢信号。在目标2中，我们将确定ixazomib是否 在体内将持续的PTH分解代谢作用转化为骨增强作用。目标3将通过以下方面来描述机制： Ixazomib在体外逆转持续PTH的骨分解作用。成功完成 拟议的研究将极大地促进我们对促进/限制PTH作用的机制的理解， 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 骨质疏松症的治疗

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.