Mechanisms of carbon dot-guided rAAV gene vector formulation and release forimproved, safe rAAV-mediated therapeutic gene transfer in human cartilage lesions.

局灶性软骨创伤损伤是骨科的关键问题，因为成人关节软骨由于缺乏血管形成而不能完全自行愈合。鉴于软骨完全自我修复的能力有限，已经开发了各种方案来改善软骨修复，但没有一种方案能完全修复软骨损伤。将有效的、临床适用的重组腺相关病毒(RAAV)载体应用于局灶性软骨缺损是一种在损伤部位暂时和空间转移软骨修复基因以延长基因产物表达的有效途径。然而，rAAV的安全、临床应用仍然受到体内各种障碍的阻碍，特别是在人类群体中自然存在的针对AAV衣壳蛋白的中和抗体。为了解决这个问题，本提案的目的是测试这样一种假设，即编码软骨修复(增殖的，促合成代谢的)基因(软骨特异性性别决定区域Y型高迁移率盒9-sox9转录因子和转化生长因子-β-转化生长因子)的治疗性raav载体可以通过碳点(Cds)作为保护性的、受控的递送系统来安全、有效和持久地增强体外自然3D环境中原代人骨髓间充质基质细胞的软骨修复相关的过程和机制，在人的实验性骨软骨缺损的翻译前模型中，以及在临床上相关的体内的翻译性局灶性软骨缺损与直接应用其自由形式的载体有关。该项目可能会在不久的将来提供新的、有效的治疗方法来加强患者的软骨修复。

Focal cartilage traumatic lesions are critical issues in orthopaedics since the adult articular cartilage does not completely heal on itself as a result of the lack of vascularization in this tissue. In light of the restricted ability of the cartilage for full self-healing, various options have been developed to improve cartilage repair, but none can fully heal cartilage lesions. Administration of the potent, clinically adapted recombinant adeno-associated virus (rAAV) vectors in focal cartilage defects is a strong approach to temporarily and spatially transfer chondroreparative genes in sites of injury for an extended expression of the gene products. Yet, the safe, clinical use of rAAV is still hindered by various obstacles in vivo, especially by the natural presence of neutralizing antibodies against the AAV capsid proteins in the human population. To address this issue, the goal of the present proposal is to test the hypothesis that therapeutic rAAV vectors coding for chondroreparative (proliferative, pro-anabolic) genes (the cartilage-specific sex-determining region Y-type high mobility box 9 - SOX9 - transcription factor and the transforming growth factor beta - TGF-ß) may be delivered via carbon dots (CDs) as protective, controlled delivery systems to safely, effectively, and durably enhance the processes and mechanisms relevant of cartilage repair in primary human bone marrow-derived mesenchymal stromal cells in a natural 3D environment in vitro, in a pre-translational model of human experimental osteochondral defect in situ, and in clinically relevant, translational focal cartilage defects in vivo relative to direct application of the vectors in their free form. This project may offer new, effective therapies to enhance cartilage repair in patients in a close future.