Elucidating the Role of Sulfatase-2 in RA Pathogenesis

项目总结 在类风湿关节炎(RA)中，细胞因子肿瘤坏死因子-α(肿瘤坏死因子-α)在驱动滑膜方面起主导作用 软骨和骨骼的增生和进行性破坏。我们的初步数据显示细胞外 硫酸酯酶-2是类风湿关节炎滑膜成纤维细胞肿瘤坏死因子-α信号转导的潜在介质(或促进剂) (RASF)。在细胞表面，Sulf-2从硫酸乙酰肝素蛋白多糖(HSPGs)上裂解硫酸盐基团，这是 进而影响受体/配体结合和随后的一系列趋化因子、细胞因子和 增长因素。肿瘤坏死因子-α与热休克蛋白G之间的相互作用以及SURF-2在肿瘤坏死因子-α中的作用尚不清楚 信号仍未被探索。我们发现siRNA或小分子对人RASF中Sulf-2的抑制作用 拮抗剂OKN-007显著降低肿瘤坏死因子-α诱导的黏附分子ICAM1、VCAM1的表达 和粘附素-11，以及产生炎症介质IL-6和IL-8。用RNA进行转录组分析 测序(RNAseq)显示，与阴性对照siRNA相比，Sulf-2的基因敲除显著 减少肿瘤坏死因子-α对与黏附、细胞增殖和趋化因子相关基因的诱导，同时伴随 增加骨保护和抗炎基因的表达。RNAseq结果的通径分析 结果显示，在肿瘤坏死因子-α刺激的RASF中，Sulf-2基因敲除显著改变了48个基因的表达 巨噬细胞、成纤维细胞和内皮细胞在类风湿性关节炎中的顶级典型通路作用。这 Proposal利用了这些新的观察结果，我们研究的中心假设是Sulf-2介导了 肿瘤坏死因子-α信号促进类风湿关节炎滑膜增生、侵袭和组织破坏。作为一个细胞外的 酶，Sulf-2是一个有吸引力的药物抑制靶点，而Sulf-2的小分子抑制剂 在人体临床试验中显示出很高的生物安全性。我们的目的是阐明Sulf-2参与肿瘤坏死因子-2的机制。 α信号转导机制，并探讨靶向SURF-2依赖通路对卵巢癌的潜在治疗价值。 类风湿关节炎的治疗。目标1中提出的研究将探讨Sulf-2调节血管生成的分子机制。 肿瘤坏死因子-α信号通路及其对RASF发病下游介质的影响。在《目标2》中，我们 将在人肿瘤坏死因子-α转基因小鼠模型中测试SURF-2的体内药理抑制作用 拉。这些研究的成功将揭示一种新的调控肿瘤坏死因子-α信号的分子机制。 通过调节Sulf-2调节RASF，并验证一种潜在的新药物治疗的安全性和有效性 治疗这种常见的、使人衰弱的疾病。

PROJECT SUMMARY In rheumatoid arthritis (RA), the cytokine tumor necrosis factor-α (TNF-α) plays a dominant role in driving synovial hyperplasia and progressive destruction of cartilage and bone. Our preliminary data suggests the extracellular enzyme Sulfatase-2 (Sulf-2) to be a potential mediator (or facilitator) of TNF-α signaling in RA synovial fibroblasts (RASFs). At the cell surface, Sulf-2 cleaves sulfate groups from heparan sulfate proteoglycans (HSPGs), which in turn affects the receptor/ligand binding and subsequent signaling of an array of chemokines, cytokines and growth factors. Interactions between TNF-α and HSPGs are not well defined, and the role of Sulf-2 in TNF-α signaling remains unexplored. We found that inhibition of Sulf-2 in human RASFs with siRNA or a small molecule inhibitor (OKN-007) significantly reduced TNF-α-induced expression of adhesion molecules ICAM-1, VCAM-1 and Cadherin-11, and production of inflammatory mediators IL-6 and IL-8. Transcriptome analysis by RNA sequencing (RNAseq) showed that, compared to a negative control siRNA, knockdown of Sulf-2 significantly reduced TNF-α induction of genes related to adhesion, cell proliferation, and chemokines, while concomitantly increasing expression of bone-protective and anti-inflammatory genes. Pathway analysis of RNASeq results showed that Sulf-2 knockdown in TNF-α-stimulated RASFs significantly altered expression of 48 genes in the top canonical pathway Role of Macrophages, Fibroblasts and Endothelial Cells in Rheumatoid Arthritis. This proposal capitalizes on these novel observations, and the central hypothesis of our study is that Sulf-2 mediates TNF-α signaling to promote synovial hyperplasia, invasion and tissue destruction in RA. As an extracellular enzyme, Sulf-2 is an attractive target for pharmacologic inhibition, and a small molecule inhibitor of Sulf-2 has shown high biosafety in human clinical trials. We aim to elucidate the mechanism of Sulf-2 participation in TNF- α signaling and investigate the potential therapeutic value of targeting Sulf-2-dependent pathways for the treatment of RA. Studies proposed in Aim 1 will investigate the molecular mechanisms of Sulf-2 modulation of TNF-α signaling pathways and effects on downstream mediators of pathogenesis in human RASFs. In Aim 2 we will test the in vivo effects of pharmacologic inhibition of Sulf-2 in a human TNF-α transgenic mouse model of RA. The success of these studies will reveal a novel molecular mechanism of modulating TNF-α signaling in RASFs by regulating Sulf-2, and validate the safety and effectiveness of a potential new pharmacologic treatment for this common and debilitating disease.