Therapeutic Application of Painless Nerve Growth Factor to Accelerate Endochondral Fracture Repair

摘要 该项目的长期目标是开发和验证一种可注射、可生物降解的纳米线输送方式 用于局部和持续释放“无痛”神经生长因子(NGF)亚型以加速的平台 骨折愈合在临床情况下的延迟愈合。约有1500万人骨折受伤 在美国，每年估计有10%-15%的健康人群中的骨折导致 延迟愈合或非愈合。7，8然而，在患有血管疾病的患者中，延迟愈合率增加到近50% 损害或高共病负担，如糖尿病、年龄增加、吸烟和肥胖。9，10目前 对于延迟愈合或骨不连的护理标准是手术干预，以增加稳定性或促进 通过应用骨移植进行愈合。骨形态发生蛋白(BMP)是FDA唯一的生物 批准用于骨折修复，只有在狭窄的适应症窗口内才能在标签上使用。然而，BMP 需要手术植入，通常仅限于风险最高的骨折，因为成本高，有限 临床疗效的证据，以及严重非目标效应的风险。11-14因此，存在未满足的临床 需要能够在非手术输送平台中刺激骨再生的生物制剂。这 应用建立在强有力的初步数据基础上，表明NGF在注射时加速骨折修复 进入长骨愈合的软骨期。重要的是，我们的初步数据首次表明NGF 作用于软骨细胞，促进与软骨内骨化(EO)相关的程序。这样做的目的是 格兰特将在这些初步数据的基础上，将NGF开发成一个适合临床翻译的平台。在……里面 第一个目标，我们优化了NGF突变形式(NGFR100W)刺激内软骨的剂量和时机 骨折修复。NGFR100W是一种新型的无痛NGF，它能有效地与TrkA受体结合，提供同样的功能 与野生型NGF一样具有营养作用，但不能与p75NTR受体结合，从而显著降低发病风险 在第二个目的中，我们探讨了NGF/NGFR100W刺激骨折修复的机制 有条件地删除TrkA受体。治疗性递送刺激的分子通路研究进展 神经生长因子在长骨骨折愈合中的作用尚未得到严格的研究。最后，在第三个目标中，我们修改了 先前开发的可注射肝素包裹的聚己内酯(PCL)纳米线17，用于包裹和 持续输送无痛神经生长因子。在这里，我们还结合了建立的糖尿病临床前模型(LEPOB) 在畸形愈合的临床相关场景中，展示延迟愈合以挑战我们的治疗。这些目标 允许我们测试中心假设，即无痛NGF疗法可以通过作用促进骨折愈合 通过TrkA信号刺激软骨细胞向成骨细胞转化。我们的跨学科团队 骨折愈合、生物材料和NGF/TrkA信号方面的专家使我们能够成功 完成建议的研究。重要的是，我们的方法基于创建与翻译相关的 有可能显著改善骨折后患者预后的治疗平台。

ABSTRACT The long-term goal of this project is to develop and validate an injectable, biodegradable nanowire delivery platform for local and sustained release of a “painless” nerve growth factor (NGF) isoform to accelerate fracture healing in clinical scenarios of delayed healing. Approximately 15 million fracture injuries occur each year in the United States (US).6 An estimated 10-15% of fractures within a healthy population result in delayed- or non-union.7,8 However, delayed healing rates increase to almost 50% in patients with vascular damage or high co-morbidity burdens such as diabetes, increased age, smoking, and obesity.9,10 The current standard of care for delayed healing or non-union is surgical intervention to increase stability or to promote healing through application of bone grafts. Bone morphogenetic protein (BMP) is the only biologic with FDA approval for use in fracture repair, with “on-label” use only within a narrow indication window. However, BMP requires surgical implantation and is typically limited to only the most at-risk fractures due to the high cost, limited evidence of clinical efficacy, and risk of severe off-target effects.11-14 As such, there exists an unmet clinical need for biologics that could stimulate bone regeneration in a non-surgical delivery platform. This application builds on strong preliminary data demonstrating that NGF accelerates fracture repair when injected into the cartilaginous phase of long bone healing. Importantly, our preliminary data is the first to show that NGF acts on chondrocytes to promote programs associated with endochondral ossification (EO). The goal of this grant is to build upon these preliminary data to develop NGF into a platform suitable for clinical translation. In the first Aim, we optimize the dose and timing of a mutant form of NGF (NGFR100W) to stimulate endochondral fracture repair. NGFR100W is a novel “painless” NGF that efficiently binds to the TrkA receptor to provide the same trophic effect as wild type NGF, but fails to bind to the p75NTR receptor to significantly reduce risk of nociception.15,16 In the second Aim, we probe the mechanism by which NGF/NGFR100W stimulates fracture repair by conditionally deleting the TrkA receptor. To date the molecular pathways stimulated by therapeutic delivery of NGF have not been rigorously studied in long bone fracture healing. Lastly, in the third Aim, we modify our previously developed injectable heparin coated polycaprolactone (PCL) nanowires17 for encapsulation and sustained delivery of painless NGF. Here we also incorporate a pre-clinical model of diabetes (Lepob) established to demonstrate delayed healing to challenge our therapy in a clinically relevant scenario of malunion. These aims allow us to test the central hypothesis that a painless NGF therapy can improve fracture healing by acting through TrkA signaling to stimulate chondrocyte-to-osteoblast transformation. Our interdisciplinary team of experts in fracture healing, biomaterials, and NGF/TrkA signaling uniquely positions us to successfully accomplish the proposed study. Importantly, our approach is grounded in creating a translationally relevant therapeutic platform that has the potential to significantly improve patient outcomes following a fracture.