Nanobody-based gene therapy targeting complement component C3 reduces choroidal neovascularization in mice.

Wet age-related macular degeneration (wAMD) is a leading cause of vision loss and is characterized by choroidal neovascularization (CNV). Current CNV management requires multiple treatments and lacks long-term efficiency, creating a need for better therapeutics. wAMD pathogenesis is associated with excessive activation of the complement system, contributing to retinal damage. Therefore, we generated a vector expressing the small alternative pathway-targeting nanobody, hC3Nb1, to treat wAMD. We demonstrate that hC3Nb1 is efficiently expressed and secreted by mammalian cells and shows full alternative pathway and partial classical pathway inhibition in vitro. A dual-promoter approach was used to generate a lentiviral-based vector for co-expression of hC3Nb1 and marker protein eGFP. Profound and safe hC3Nb1-expression, along with its secretion from the retinal pigment epithelium (RPE), was confirmed following subretinal injection of nanobody expressing-vector in mice. The therapeutic potential of vector-encoded hC3Nb1 was demonstrated in vitro by protecting RPE from complement-mediated stress, and in vivo by reducing laser-induced CNV sizes in a mouse model consistent with complement inhibition. For the first time, nanobodies expressed in the eye are used therapeutically, and our findings suggest that hC3Nb1-based gene therapy may be a safe and long-acting treatment for wAMD and other chorioretinal diseases with dysregulated complement activation.