Effective local delivery of bone anabolic agent to accelerate the healing of delayed fracture union

摘要 骨折是主要的肌肉骨骼损伤，每年影响超过940万美国人。大致 约10%的骨折合并有愈合延迟或骨不连，导致慢性疼痛、活动障碍和 生活质量差。病理生理和习惯性危险因素，包括骨折部位不稳定，同时存在 疾病(如糖尿病)，长期接触某些药物(如糖皮质激素或GC)和 吸烟/酗酒也是主要诱因，这可能会干扰骨折愈合的生物学。 并导致延迟愈合或骨不连。GCS被广泛用于治疗住院的COVID患者。vt.给出 目前大流行的规模和美国人口中显著的COVID感染率，一篇文章 大流行的骨折风险激增和延迟的骨折愈合病例是可以预见的。目前，骨科 手术和自体骨移植仍然是临床治疗延迟骨折愈合的金标准。 FDA批准的非侵入性治疗措施很少用于治疗GC诱导的延迟 骨折愈合。为了解决这一重大的未得到满足的临床需求，我们开发了N-(2- 羟丙基甲基丙烯酰胺(HPMA)为基础的水溶性温敏聚合物前药(P-TAN) 丹参酮IIA(丹参酮IIA，一种有效的骨合成代谢剂)。P-Tan的水溶液在室内是自由流动的 温度和在≥27°C时转变为水凝胶(ProGel-Tan)，这为 持续在当地交付Tan。当在泼尼松诱导的延迟骨折愈合的小鼠模型中进行测试时， 连续两个月的ProGel-Tan治疗可以加速骨折愈合5周 未观察到任何副作用。骨的生物力学性能也完全恢复。而当 每月两次的ProGel-Tan治疗似乎很有希望，目前的骨科实践青睐单次 在骨折稳定过程中的剂量治疗是出于实用的原因以及较少的感染 风险。因此，我们假设，通过进一步的结构和配方优化，我们将能够确定 一种安全高效的单剂普罗格列坦治疗GC的优化处方 延迟骨折愈合。为了验证这一假设，我们在这个项目中提出了三个具体目标：1.表征 不同结构和处方参数对ProGel-Tan的影响；2.鉴别单剂ProGel-Tan 提供最佳骨折愈合疗效的Tan配方；3.了解ProGel-Tan的独特之处 药理学和假定的作用机制。在成功完成建议的研究后，我们 预期确定至少一种最优的ProGel-Tan配方，以满足对 单次给药加速GC诱导的延迟骨折愈合约5周 使小鼠损伤骨的生物力学特性正常化。然后，它将接受进一步的临床前研究 为临床翻译做准备的大动物模型的研究。

ABSTRACT Fracture is the leading musculoskeletal injury affecting over 9.4 million Americans annually. Approximately ~10% of fractures are complicated by delayed healing or non-union, resulting chronic pain, impaired mobility and poor quality of life. Pathophysiological and habitual risk factors, including fracture site instability, concurrent diseases (e.g., diabetes), chronic exposure to certain medications (e.g., glucocorticoids or GCs) and tobacco/alcohol use are also major contributing causes, which may interfere with the fracture healing biology and lead to delayed healing or non-union. GCs are widely used in treating hospitalized COVID patients. Given the scale of the current pandemic and the significant COVID infection rate among the US population, a post pandemic surge of fracture risk and delayed fracture healing cases may be anticipated. Currently, orthopaedic surgery and autologous bone grafts are still the gold standard in clinical management of delayed fracture healing. There are few FDA approved non-invasive therapeutic interventions for the treatment of GC-induced delayed fracture healing. To address this significant unmet clinical need, we have developed a N-(2- hydroxypropyl)methacrylamide (HPMA)-based water-soluble thermoresponsive polymeric prodrug (P-TAN) of Tanshinone IIA (TAN, a potent bone anabolic agent). The aqueous solution of P-TAN is free-flowing at room temperature and transitions into a hydrogel (ProGel-TAN) at ≥ 27°C, which provide a unique mechanism for sustained local delivery of TAN. When tested in a prednisone-induced delayed fracture healing mouse model, two consecutive monthly ProGel-TAN treatments were found to accelerate the fracture healing by 5 weeks without any side effects observed. The biomechanical properties of the bone were also fully restored. While two monthly ProGel-TAN treatment seems to be very promising, the current orthopaedic practice favors a single dose treatment at the time of the fracture stabilization procedure for pragmatic reasons as well as less infection risk. Therefore, we hypothesize that with further structural and formulation optimization, we will be able to identify an optimized single dose ProGel-TAN formulation that is highly effective and safe in treating the GC-induced delayed fracture union. To test this hypothesis, we propose three Specific Aims in this project: 1. To characterize the impact of different structural and formulation parameters of ProGel-TAN; 2. To identify single dose ProGel- TAN formulations that provide optimal fracture healing efficacy; 3. To understand ProGel-TAN’s unique pharmacology and putative mechanisms of action. At the successful completion of the proposed research, we anticipate the identification of at least one optimal ProGel-TAN formulation that would satisfy the demand for a single dose treatment to accelerate the healing of the GC-induced delayed fracture union by ~ 5 weeks and normalize biomechanical properties of the injured bone in mice. It will then be subjected to further pre-clinical studies on large animal models in preparation for clinical translation.