Osteoarthritis (OA) is a prevalent degenerative joint disorder and the leading cause of disability. Presently, there are no disease-modifying osteoarthritis drugs (DMOADs). Diagnosis and pharmacological intervention occur mostly at a late stage, and current treatments offer only temporary, palliative relief before disease progression necessitates joint replacement. OA prevalence is high, at roughly 27% of those over 40 years old, and occurrence of post-traumatic OA (PTOA) is even higher (over 50%) following injury of large joints such as the knee. Given its high incidence and predictability, PTOA has potential to be treated prophylactically, a strategy that is both conducive to achieving disease-modifying outcomes and commercially/clinically feasible, provided treatment offers long-term protection. We aim to achieve persistent joint protection by permanently converting cells in situ into “on-demand” TIMP-3 “factories”, harnessing TIMP-3 as a pan-MMP inhibitor that blocks multiple aspects of OA pathology, including cartilage degradation, angiogenesis, and inflammation. “On-demand” expression of TIMP-3 will be achieved via a targeted and permanent gene insertion that hijacks the Mmp13 promoter. This approach is based on a nonviral CRISPR-based nanoparticle and activates TIMP-3 expression only when pathological (OA) stimuli are present, minimizing potential side-effects. We propose to optimize a nanoparticle formulation for non-viral gene knock-in and quantify the therapeutic efficacy of TIMP-3 knock-in in vitro and in vivo. This therapy has potential to avoid significant loss in quality of life for patients who experience a large joint injury and is uniquely enabled by our team with expertise in intracellular delivery (Duvall), polymer and nanoparticle chemistry (D’Arcy), genome editing and synthetic biology (Brunger), and PTOA biology (Hasty).