Sequential intra-articular HCAd-NFκB-IL-1Ra delivery improves therapeutic efficacy in post-traumatic osteoarthritis.

Osteoarthritis is the most common joint disease for which disease-modifying therapies remain unavailable. Intra-articular gene delivery of interleukin-1 receptor antagonist (IL-1Ra) using high-capacity adenovirus (HCAd) has shown therapeutic promise; however, the duration of therapeutic benefit and the feasibility of repeat dosing under anti-adenoviral immunity remain unresolved. Using the murine anterior cruciate ligament transection model of osteoarthritis, we show that a single intraarticular injection of HCAd5-NFκB-IL-1Ra provides structural preservation and functional improvement in early-stage osteoarthritis but fails to sustain cartilage protection as disease progresses. However, HCAd5 transduction following repeated treatment is limited due to pre-existing immunity against this serotype. Notably, exchanging serotypes for repeated treatments effectively restores vector transduction and transgene expression in both healthy and osteoarthritic joints. Leveraging this strategy, we demonstrate that sequential intra-articular HCAd-NFκB-IL-1Ra administration does not further improve pain or motor function compared to the initial treatment, but preserves cartilage as assessed by histopathology and phase-contrast μCT, irrespective of serotype. These findings establish HCAd serotype switching as a feasible approach to overcome immune barriers to repeat intra-articular gene therapy. Importantly, sequential HCAd-NFκB-IL-1Ra administration enhances therapeutic durability in post-traumatic osteoarthritis, providing a translational framework for repeat intra-articular gene delivery strategies aimed at long-term disease modification in osteoarthritis.