Treatment of osteoarthritis by inhibition of aggrecanases using targeted delivery of engineered TIMP-3

研究背景骨关节炎(OA)是英国最常见的关节炎形式，影响着近900万人，但目前还没有有效的治疗方法。因此，临床上对减缓疾病进展和改善患者生活质量的新的、更好的治疗方法的需求仍未得到满足。几十年来，使用TIMP-3等负责软骨破坏的酶的内源性抑制剂的可能性一直是一个耐人寻味的可能性。然而，尽管进行了多年的研究，但许多挑战阻碍了TIMP-3作为治疗OA的方法的开发，包括不可接受的副作用和生产足够数量的蛋白质的问题。我们现在已经克服了这些困难，通过改造TIMP-3来选择性地针对仅用于抑制的聚集聚糖酶，并开发了一种新的蛋白质生产方法，从而使重组TIMP-3的产量增加了10倍。这个项目的目的是开发一种新的治疗策略，使用这种工程的TIMP-3来抑制侵袭酶来治疗骨性关节炎。这里提出的策略是使用一种新的方法(LAP技术)将这种分子传递到体内的关节炎关节。这种方法能够在疾病部位(S)传递高浓度的治疗分子，从而消除副作用，提高疗效。目的与目的本方案的目的是开发一种治疗骨性关节炎的新治疗策略。我们的假设是，局部注射工程TIMP-3将通过获得高浓度的TIMP-3来抑制侵袭酶的活性，从而改善受OA影响的关节软骨的破坏。这些目标将通过完成以下目标来实现：1.利用LAP融合蛋白技术表达和纯化大量潜在的工程化TIMP-3。2.检测潜伏工程化TIMP-3对小鼠骨性关节炎发生发展的影响。描述工程TIMP-3对参与骨性关节炎发病机制的ADAM和ADAMTS家族成员的抑制作用。潜在的应用和益处目前还没有治疗骨性关节炎的有效方法，目前的治疗方法旨在减轻骨性关节炎患者的疼痛。然而，这些疗法在阻止疾病进展方面没有效果，结果是终末期骨性关节炎目前通过关节置换手术进行治疗。这项工作的应用可能会转化为一种治疗骨性关节炎的新疗法，或许可以与现有疗法相结合，创建一种治疗方案，减少关节炎关节中软骨退化和随后的软骨下骨增厚。这可以显著减少关节损伤，从而减缓疾病的进展，从而为患者提供更高的生活质量，并可能推迟或避免关节置换手术。这里提出的方法有许多优点。首先，通过使用一种不被宿主免疫系统识别的自然产生的蛋白质的重组形式，将这种治疗的副作用的风险降至最低。其次，治疗分子在疾病部位(S)的局部激活将导致局部浓度较高，并进一步将非靶向效应的风险降至最低。最后，我们设计了TIMP-3分子以提高其特异性，从而避免了以前使用合成酶抑制剂时出现的副作用。此外，这一战略具有很大的翻译潜力。这项建议中概述的研究可能导致临床试验，以开发中期(7-10年)治疗骨性关节炎的新战略。

Context of researchOsteoarthritis (OA) is the most common form of arthritis affecting nearly 9 million people in the UK, yet there are no effective treatments. As such, there is a significant unmet clinical need for new and better therapies to slow progression of disease and improve patient quality of life. The possibility of using endogenous inhibitors of the enzymes responsible for cartilage destruction, such as TIMP-3, has been an intriguing possibility for decades. However, despite many years of research a number of challenges have prevented development of TIMP-3 as a therapy for OA, including unacceptable side-effects and problems in producing sufficient quantities of protein. We have now overcome these difficulties by engineering TIMP-3 to selectively target aggrecanases only for inhibition and developing a novel method of protein production that results in a 10-fold increase in production of recombinant TIMP-3. The aim of this project is to develop a new therapeutic strategy for the treatment of OA using this engineered TIMP-3 to inhibit aggrecanases. The strategy proposed here is to employ a novel method (the LAP technology) of delivering this molecule to arthritic joints in vivo. This approach enables the delivery of high local concentrations of therapeutic molecules at site(s) of disease, thus eliminating side-effects and increasing therapeutic efficacy.Aims and ObjectivesThe aim of this proposal is to develop a new therapeutic strategy for treatment of OA. Our hypothesis is that local delivery of engineered TIMP-3 will ameliorate cartilage destruction in joints affected by OA by achieving high local concentrations of TIMP-3 that will inhibit aggrecanase activity. These aims will be achieved by the completion of the following objectives:1. Express and purify substantial quantities of latent engineered TIMP-3 using the LAP fusion protein technology. 2. Determine the effects of latent engineered TIMP-3 on the development and progression of OA in a mouse model of disease.3. Characterise the inhibitory effects of engineered TIMP-3 on members of the ADAM and ADAMTS families of enzymes involved in the pathogenesis of OA. Potential applications and benefitsThere are no effective treatments for OA and current therapies aim to alleviate the pain experienced by people with OA. However, these therapies have no effect on halting disease progression with the result that end-stage OA is currently treated by joint-replacement surgery. The applications of this work may translate into a new therapy for OA, perhaps in combination with existing therapies, to create a treatment regimen that reduces cartilage degradation and subsequent subchondral bone thickening in arthritic joints. This could significantly reduce joint damage so that disease progression is curtailed, thus providing patients with improved quality of life and possibly delaying or avoiding joint replacement surgery. There are a number of advantages to the approach proposed here. Firstly, the risks of side effects of this treatment are minimized by using a recombinant form of a naturally occurring protein that is not recognised by the host immune system. Secondly, local activation of therapeutic molecules at the site(s) of disease will result in high local concentrations and further minimise the risks of off-target effects. Finally, we have engineered the TIMP-3 molecule to increase its specificity thus avoiding the side-effects previously seen with synthetic enzyme inhibitors. Furthermore, this strategy has a great deal of translational potential. The research outlined in this proposal could lead to clinical trials for the development of a new strategy for treating OA in the medium term (7-10 years).