Validating the efficacy of SITREX in preventing heterotopic ossification

在冲击波诱导截肢等创伤后，会发生一系列事件，包括炎症、血管生成、纤维增殖和软骨生成，这些事件共同促进了损伤部位的成骨和异位骨的形成。这种异位骨化被称为异位骨化，可能会非常痛苦和虚弱-手术切除是治疗这种疾病的唯一方法。目前还没有有效的治疗方法来预防异位骨化，我们认为这在一定程度上是因为在级联反应开始后，针对创伤反应级联反应的一个方面进行了治疗。当这个项目开始时，我们开始开发一个创伤反应指数(T-REX)，它将由改变的基因组成，促进异位骨化的发展。我们假设，针对这些基因，这些基因的变化发生在这个级联的顶端，将能够识别防止异位骨化的治疗方法。我们使用了一系列下一代测序、分子和细胞生物学方法来识别组成T-REX第一次迭代的6个基因(见方法部分)。在进一步研究后，我们发现，用沉默的RNA抑制一个特定的基因，在暴露于创伤和骨化刺激的细胞中，导致体外骨形成减少70%。为了使其适用于临床翻译，我们将这种siRNA包装成纳米颗粒，可以通过水凝胶输送到损伤部位。这种纳米颗粒水凝胶是我们的产品SITREX，也是当前提案的主题。在这项资助中，我们的目标是在体外3D器官模型和体内创伤反应性异位骨化模型中验证SITREX在预防异位骨化方面的有效性。这一目标弥合了已经开展的基础科学研究和转化性研究之间的差距。在体外和体内验证SITREX在这些异位骨化模型中的有效性将提高技术准备水平，并为未来的资助者降低SITREX的风险。

After trauma such as a blast wave-induced amputation, there is a cascade of events including inflammation, angiogenesis, fibroproliferation, and chondrogenesis that collectively contribute to osteogenesis and formation of ectopic bone at the injury site. This ectopic bone, known as heterotopic ossification, can be extremely painful and debilitating - surgical resection is the only way to treat the disease. There are no effective therapeutic treatments to prevent heterotopic ossification and we believe that is in part due to the therapeutics targeting one aspect of this trauma-response cascade, after the cascade has started. When this project started we set out to develop a Trauma-Response indEX (T-REX), that would comprise of genes that changed, facilitating the development of heterotopic ossification. We hypothesised that targeting these genes, changes to which occur at the apex of this cascade, would enable identification of a therapeutic which prevented heterotopic ossification. We used a range of next generation sequencing, molecular and cell biology approaches to identify 6 genes that would comprise the first iteration of T-REX (see approach section). After delving into this further, we found that inhibiting one gene in particular, with a silencing RNA, resulted in a 70% reduction of bone formation in vitro, in cells exposed to both trauma and ossification stimuli. To make this applicable for clinical translation, we packaged this siRNA into nanoparticles that could be delivered via a hydrogel to the site of injury. This nanoparticle laden hydrogel is our product, SITREX, and the subject of this current proposal. In this grant, our objective is to validate the effectiveness of SITREX in preventing heterotopic ossification in a 3D organotypic model in vitro, and a trauma responsive heterotopic ossification model in vivo. This objective bridges the gap between the basic science research already conducted, and translational studies. Validating the efficacy of SITREX in these heterotopic ossification models in vitro and in vivo will increase the technology readiness level, and de-risk SITREX for future funders.