Inflammation elicited by rAAV vectors continues to present a critical challenge for long-term efficacy and safety of gene therapy in the eye. Preclinical models of gene therapy-associated uveitis (GTAU) show that despite resolution of early acute inflammatory response, persistent subclinical inflammation remains. Here, we employ the GTAU model in Cx3cr1CreER:R26-tdTomato+/- mice to reveal that intravitreal (IVT) rAAV2 administration elicits sustained microglial dysregulation and retention of CD3+ T cells extending to 50 days post-injection (dpi). Deploying pharmacological and genetic approaches, we define the absolute requirement for microglia and T cells to mediate rAAV2-induced inflammation. Targeted depletion confirmed microglia-independent mechanisms initiate GTAU, whilst elimination of lymphocytes prevented both inflammation and microglial activation. Systematic evaluation of therapeutic strategies reveals identified inhibition of T cell recruitment via sphingosine-1-phosphate receptor modulation, but not B cell depletion, as an effective steroid-sparing strategy to prevent both acute and long-term subclinical inflammation. Collectively, our findings challenge the paradigm of microglia-driven ocular inflammation and support the utility of targeted T cell immunomodulation strategies to control GTAU and maintain long-term ocular homeostasis.
Langer et al. (Sun,) studied this question.