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可注射的、可生物降解的纳米线递送 用于局部和持续释放“无痛”神经生长因子（NGF）同种型的平台， 骨折愈合延迟的临床情况。大约有1500万例骨折 6据估计，健康人群中10-15%的骨折会导致 延迟愈合或骨不连。7，8然而，在血管性骨折患者中，延迟愈合率增加至近50%。 损害或高并发症负担，如糖尿病、年龄增加、吸烟和肥胖。 延迟愈合或骨不连的标准护理是手术干预以增加稳定性或促进 通过应用骨移植来愈合。骨形态发生蛋白（BMP）是唯一获得FDA批准的生物制品。 批准用于骨折修复，仅在狭窄的适应症窗口内按标签使用。然而，BMP 需要外科植入，并且由于高成本、有限的 临床疗效的证据和严重脱靶效应的风险。11 -14因此，存在未满足的临床 需要能够在非手术递送平台中刺激骨再生的生物制剂。这 该应用建立在强有力的初步数据的基础上，这些数据表明，当注射NGF时， 进入长骨愈合的软骨阶段重要的是，我们的初步数据首次表明， 作用于软骨细胞以促进与软骨内骨化（EO）相关的程序。这个目标 格兰特是建立在这些初步的数据，发展神经生长因子成为一个平台，适合临床翻译。在 第一个目标是优化NGF突变体（NGFR 100 W）刺激软骨内分泌的剂量和时间 骨折修复NGFR 100 W是一种新型的“无痛”NGF，其有效地结合TrkA受体以提供相同的功能。 与野生型NGF一样具有营养作用，但不能与p75 NTR受体结合以显著降低 在第二个目标中，我们探索了NGF/NGFR 100 W刺激骨折修复的机制。 通过有条件地删除TrkA受体。确定治疗传递刺激的分子途径的日期 NGF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，我们修改了我们的 先前开发的用于封装的可注射肝素涂覆的聚己内酯（PCL）微囊17， 持续输送无痛神经生长因子在这里，我们还纳入了糖尿病的临床前模型（Lepob）建立 证明延迟愈合，以挑战我们在临床相关的畸形愈合情况下的治疗。这些目标 这使我们能够测试中心假设，即无痛的NGF治疗可以通过作用于骨折愈合， 通过TrkA信号传导来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信号传导方面的专家，使我们能够成功地 完成拟议的研究。重要的是，我们的方法立足于创建一个与实践相关的 这是一种治疗平台，有可能显著改善骨折后的患者结局。

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.