GENE THERAPY FOR PROSTHETIC LOOSENING

假体松动的基因治疗

• 批准号: 6196509
• 负责人: Regis J O'Keefe
• 金额: $ 33.96万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2004-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6196509
• 关键词: RNase protection assay; adeno associated virus group; biomaterial interface interaction; cytokine; enzyme linked immunosorbent assay; flow cytometry; gel mobility shift assay; gene therapy; human tissue; iatrogenic disease; inflammation; laboratory mouse; macrophage; mechanical stress; nuclear factor kappa beta; osteoclasts; paracrine; pathologic bone resorption; postoperative complications; prosthesis; southern blotting; tissue /cell culture; transfection /expression vector; western blottings

DESCRIPTION (Adapted from the Applicant's Abstract): More than 400,000 arthroplasties are performed annually in the US and up to 20 percent of these require revision surgery due to aseptic loosening. This complication constitutes a major source of morbidity and represents billions of dollars in health care costs. This proposal is designed to (1) define the role of TNFalpha/NFkB/RANK signal transduction in the disease process; and (2) develop a gene therapy protocol targeted to block the TNFalpha/NFkB/RANK mediated inflammatory bone resorption as a potential treatment for aseptic loosening. The hypothesis to be studied is that (i) particles stimulate NFkB activation leading to induction of TNFalpha; (ii) TNFalpha amplifies the inflammatory response as a paracrine mediator of NFkB stimulation; (iii) macrophage-secreted inflammatory mediators stimulate RANK ligand secretion in stromal cells; and (iv) RANK ligand stimulates osteoclastogenesis and bone resorption. It is the applicant group's hypothesis that TNFR-Ig primarily inhibits inflammation and directly inhibits osteoclastogenesis and bone resorption, while RANK-Ig/OPG are direct regulators of osteoclast formation, activation and apoptosis. These hypotheses will be investigated in a series of in vitro and in vivo experiments that include examination of the effects of particles on NFkB activation and cytokine secretion in cell lines genetically engineered to have deficient NFkB signaling, as well as in cells treated with TNFR-Ig and RANK-Ig. The effect of these proteins on macrophage induced (i) stromal cell expression of RANK ligand and OPG; (ii) osteoclastogenesis; and (iii) bone resorption will be assessed. Finally, they will investigate adeno-associated viral (AAV) gene transfer as an effective delivery system for TNFR-Ig and OPG. The applicants hypothesize that particles will stimulate AAV second strand synthesis and lead to target protein expression that will be sustained and occur after a period of dormancy. The hypothesis is that the AAV-transduced target proteins, TNFR-Ig and OPG, inhibit particle-induced inflammatory bone resorption in vivo, expecting that the effects will be greatest when used in combination, due to their actions at different stages in the process of inflammatory bone resorption.

描述（改编自申请人的摘要）：超过 400,000 美国每年都会进行关节置换术，其中高达 20% 由于无菌性松动需要进行修复手术。这种并发症 是发病率的一个主要来源，并且涉及数十亿美元 医疗保健费用。该提案旨在 (1) 定义以下角色： 疾病过程中TNFα/NFkB/RANK信号转导； (2) 开发 旨在阻断 TNFα/NFkB/RANK 介导的基因治疗方案 炎症性骨吸收作为无菌性松动的潜在治疗方法。 要研究的假设是 (i) 粒子刺激 NFkB 激活 导致 TNFα 的诱导； (ii) TNFα 放大炎症 作为 NFkB 刺激的旁分泌介质的反应； (iii) 巨噬细胞分泌 炎症介质刺激基质细胞中的 RANK 配体分泌；和 (iv) RANK配体刺激破骨细胞生成和骨吸收。它是 申请人小组的假设是 TNFR-Ig 主要抑制炎症 直接抑制破骨细胞生成和骨吸收，而 RANK-Ig/OPG 则 破骨细胞形成、活化和凋亡的直接调节剂。这些 假设将通过一系列体外和体内实验进行研究 包括检查粒子对 NFkB 激活的影响以及 经基因改造缺乏 NFkB 的细胞系中的细胞因子分泌 信号传导，以及用 TNFR-Ig 和 RANK-Ig 处理的细胞。的效果 巨噬细胞上的这些蛋白质诱导 (i) 基质细胞表达 RANK 配体 和OPG； (ii) 破骨细胞生成； (iii) 将评估骨吸收。 最后，他们将研究腺相关病毒（AAV）基因转移作为 TNFR-Ig 和 OPG 的有效递送系统。申请人假设 颗粒将刺激 AAV 第二链合成并产生目标蛋白 休眠一段时间后将持续并发生的表达。这 假设 AAV 转导的靶蛋白 TNFR-Ig 和 OPG 抑制 颗粒诱导的体内炎症性骨吸收，预计 由于它们的作用，组合使用时效果会最大 炎症性骨吸收过程的不同阶段。