Mechanisms of pNaSS-grafted poly(epsilon-caprolactone) scaffold-guided rAAV-mediated gene transfer for enhanced, safe human anterior cruciate ligament repair

膝关节前交叉韧带（ACL）损伤是骨科中普遍存在的严重问题和社会经济负担，可能导致骨关节炎，因为ACL具有有限的内在自我修复能力。由于目前的临床（保守、手术）干预措施都不支持ACL病变的持久、全面修复，基于基因治疗的创新选择可能为增强ACL修复过程提供强有力的工具。在ACL病变中使用强效的、临床适应的重组腺相关病毒（rAAV）载体是一种有效的方法，可以在损伤部位暂时和空间上转移修复基因，从而延长基因产物的表达。尽管如此，rAAV的安全、临床应用仍然受到体内各种障碍的阻碍，特别是存在针对人类AAV衣壳蛋白的天然中和抗体。为了解决这一具有挑战性的问题，本研究的目标是验证一种假设，即编码修复（增殖，促合成）基因（基本成纤维细胞生长因子- FGF-2 -和转化生长因子- TGF-ß）的治疗性rAAV载体可以通过功能化聚苯乙烯磺酸钠-接枝聚（epsilon-己内酯）（pnas -接枝PCL）膜作为保护性的、受控的递送平台，安全、有效地递送。在人类实验性ACL病变的体外翻译前模型中，包括在实验性体液中和条件下，以及在临床相关的体内翻译实验性ACL病变中，相对于直接应用无膜形式的载体，可以持久地增强与ACL修复相关的过程和机制。该项目可能在不久的将来提供新的、有效的治疗方法来增强患者的ACL修复。

Injuries of the knee anterior cruciate ligament (ACL) are prevalent, critical issues and a socioeconomical burden in orthopaedics, potentially leading to osteoarthritis, since the ACL has a limited intrinsic ability for self healing. As none of the current clinical (conservative, surgical) interventions support the durable, full repair of ACL lesions, innovative options based on gene therapy may provide strong tools to enhance the processes of ACL repair. Administration of the potent, clinically adapted recombinant adeno-associated virus (rAAV) vectors in ACL lesions is a potent approach to temporarily and spatially transfer reparative genes in sites of injury for a prolonged expression of the gene products. Still, the safe, clinical use of rAAV remains impeded by diverse obstacles in vivo, especially by the presence of natural neutralizing antibodies against the AAV capsid proteins in humans. To address this challenging issue, the goal of the present proposal is to test the hypothesis that therapeutic rAAV vectors coding for reparative (proliferative, pro-anabolic) genes (the basic fibroblast growth factor - FGF-2 - and the transforming growth factor beta - TGF-ß) may be delivered via functionalized poly(sodium styrene sulfonate)-grafted poly(epsilon-caprolactone) (pNaSS-grafted PCL) films as protective, controlled delivery platforms to safely, effectively, and durably enhance the processes and mechanisms relevant of ACL repair in a pre-translational model of human experimental ACL lesions ex vivo, including in conditions of experimental humoral neutralization, and in clinically relevant, translational experimental ACL lesions in vivo relative to direct application of the vectors in their film-free form. This project may offer new, effective therapies to enhance ACL repair in patients in a close future.