Mechanisms of self-assembly temperature-sensitive polymers forenhanced rAAV-mediated gene transfer into human mesenchymalstem cells

关节软骨损伤是包括创伤和骨性关节炎(OA)在内的各种关节疾病中普遍存在的、尚未解决的主要问题。目前修复创伤性或骨性关节炎软骨的治疗方案多种多样，但没有一种方法能完全恢复软骨功能。基因治疗允许在损伤部位直接应用编码治疗因子的基因，导致临时和空间限定的候选试剂的递送。重组腺相关病毒载体(RAAV)是实现这一目标的特别适合临床的工具。使用它们的重要限制仍然包括相对较低的穿透效率(通过硫酸乙酰肝素蛋白多糖细胞受体在载体-细胞接触时有效内化约13%)，个体中存在针对病毒衣壳的中和抗体，为了克服这些问题，一个潜在的途径是通过聚合物生物材料来传递rAAV，以通过不同的细胞进入机制来提高载体转移的速度。本项目的目的是验证rAAV能够有效地从聚环氧乙烷(PEO)和聚环氧丙烷(PPO)三嵌段共聚物的胶束和水凝胶中释放到hMSCs和人关节软骨细胞作为参与软骨修复过程的关键细胞群的假设。这些共聚物的不同特征(组成和结构)使得rAAV传递能够得到可调节的rAAV实现高水平和持续的基因转移的系统。这一策略可能构成一种新的安全、在软骨损伤的局部部位提供治疗候选的有效工具，作为改善该组织修复过程的手段。

Damage to the articular cartilage is a prevalent, major unresolved problem in a variety of joint disorders including trauma and osteoarthritis (OA). Current therapeutic options to repair traumatic or OA cartilage are diverse, but none can fully restore the cartilage function. Gene therapy allows for a direct application of genes coding for therapeutic factors in sites of injury, resulting in a temporarily and spatially defined delivery of the candidate agent.Recombinant adeno-assciated viral vectors (rAAV) are particularly clinically adapted tools to achieve this goal.Important limitations to their use still include a relatively low efficacy of penetration (~ 13% of effective internalization upon vector-cell contact via the heparan sulphate proteoglycan cellular receptor), the presence of neutralizing antibodies against the viral capsid in individuals, and the possible presence of inhibitory anticoagulants (heparin) in samples.A potential approach to overcome these issues is to deliver rAAV via polymeric biomaterials in order to enhance the rate of vector transfer via different cell entry mechanisms.The goal of this project is to test the hypothesis that rAAV can be effectively encapsulated and released from micelles and hydrogels of poly-ethylene oxide (PEO) and poly-propylene oxide (PPO) triblock copolymers to target hMSCs and human articular chondrocytes as key cell populations involved in the processes of cartilage repair.The differential features (composition vs. structure) of these copolymers enables the obtaining of tunable rAAV delivery systems for achieving high and sustained levels of gene transfer.This strategy could constitute a new safe, effective tool for delivering of therapeutic candidates in localized sites of cartilage lesions as a means to improve the repair processes of this tissue.