MicroRNA-based therapy for rheumatoid arthritis

项目摘要 microRNA（miRNAs）是一类单链非编码RNA，是近年来出现的一种新型的转录后调控分子， 类风湿性关节炎（RA）的发病机制。然而，最近的研究表明， 是在疾病状态如癌症和RA中miRNA表达的总体减少。虽然， 开发的miRNA疗法在概念上类似于其他抑制方法，恢复了细胞的功能， miRNA替代miRNA是一种特征较少的，但从未在RA中测试过的潜在治疗选择。有趣的是， 在RA中破坏miRNA生物合成和降解过程的根本原因还不完全清楚。 我们最近的发现提供了证据表明，miR-17在RA血清、滑膜成纤维细胞中的表达显著降低， (SFs)和滑膜组织（ST），以及在血清和关节炎诱导（AIA）大鼠。rna测序 分析显示，在人类中通过使用前体（pre）-miR-17恢复miR-17水平调节了664个基因。 RASF RNA测序数据的免疫途径分析鉴定了TNF-α中的泛素蛋白酶体系统 信号通路作为miR-17的主要靶点。此外，使用miR-17抑制剂的miR-17水平的恢复， 17（pre-miR-17）降低了泛素E3连接酶TRAF 2与其信号传导伴侣cIAP 2结合的能力，从而 抑制TNF-α诱导的下游信号转导通路，抑制IL-6、IL-8的产生， MMP-1和MMP-13。虽然这些新的发现为miR-17的影响提供了证据， 在RASFs中，对TNF-α信号转导至关重要的翻译后过程的替代，对miR- 17生物发生和周转仍然没有答案，包括RA中严重低表达的原因， 促炎性细胞因子对miR-17生物合成和周转的影响，以及miR-17的相关性和功效 RA的替代疗法基于这些新的观察结果，我们提出miR-17替代疗法可以 改善RA。因此，在具体目标1中，我们将确定miR-17生物发生的失调机制， 在RA中的周转。在目标2中，我们将评估miR-17恢复对TNF-α- 诱导人RASFs的分子和表型变化。最后，目标3将测试miR-17的体内功效。 在人RA的TNF-α转基因（hTNF-tg）小鼠模型和大鼠AIA模型中的递送。这些研究的成功 将导致两个临床上不同的发现：1）阐明了改变的miRNA生物发生和周转机制， RA发病机制，和2）验证基于miRNA的治疗方法治疗RA。

PROJECT SUMMARY MicroRNAs (miRNAs) are single stranded non-coding RNAs that have emerged as a novel posttranscriptional regulators in rheumatoid arthritis (RA) pathogenesis. However, recent studies provide evidence that in general there is an overall reduction in miRNA expression in disease states such as cancer and RA. While the rationale for developing miRNA therapeutics are conceptually similar to other inhibitory approaches, restoring the function of a miRNA by miRNA replacement is a less characterized, yet potential therapeutic option never tested in RA. Intriguingly, the underlying reasons for disrupted miRNA biogenesis and degradation processes in RA are not fully understood. Our recent finding provides evidence that miR-17 expression was significantly low in RA serum, synovial fibroblasts (SFs), and synovial tissues (STs), as well as in the serum and joints of adjuvant-induced (AIA) rats. RNA-sequencing analysis showed modulation of 664 genes by the restoration of miR-17 levels using precursor (pre)-miR-17 in human RASFs. Ingenuity pathway analysis of RNA-sequencing data identified the ubiquitin proteasome system in the TNF-α signaling pathway as a primary target of miR-17. Furthermore, the restoration of miR-17 levels using precursor-miR- 17 (pre-miR-17) reduced the ability of ubiquitin E3 ligase TRAF2 to associate with its signaling partner, cIAP2, thereby inhibiting TNF-α-induced downstream signal transduction pathways and suppressing the production of IL-6, IL-8, MMP-1, and MMP-13 in human RASFs. While these novel findings provide evidence for the impact of miR-17 replacement on posttranslational processes critical in TNF-α signaling in RASFs, several questions important to miR- 17 biogenesis and turnover remains unanswered, including the reasons for severely low expression in RA, the impact of proinflammatory cytokines on miR-17 biogenesis and turnover, and the relevance and efficacy of miR-17 replacement therapy in RA. Based on these novel observations, we propose that miR-17 replacement therapy could ameliorate RA. Thus, in specific aim 1, we will determine the deregulated mechanism of miR-17 biogenesis and turnover in RA. In aim 2, we will evaluate the relevance and molecular mechanisms of miR-17 restoration on TNF-α- induced molecular and phenotypic changes in human RASFs. Finally, aim 3 will test the in vivo efficacy of miR-17 delivery in TNF-α transgenic (hTNF-tg) mouse model and rat AIA model of human RA. The success of these studies will lead to two clinically distinct findings: 1) Elucidation of the altered miRNA biogenesis and turnover mechanism in RA pathogenesis, and 2) the validation of miRNA-based therapeutic approaches for the treatment of RA.