AAV9-mCherry

Systemic AAV9 delivery offers the potential for widespread and efficient retinal gene delivery and may therefore be a useful approach for treating diseases that are not amenable to intraocular injections, syndromes that affect multiple organs, or diseases that require early intervention. Expression produced by intravenous AAV9 is more effective in neonates than in adults, and here researchers describe the effects of age on AAV9 retinal transduction in the mouse retina. They found that the pattern of expression in neonatal mice correlated with retinal vascular development and that the areas of retinal transduction, as well as the cell types infected, varied depending on age at injection. Furthermore, sequential injections of AAV9 vectors carrying two different transgenes infected adjacent areas of the retina, providing greater coverage. Finally, these studies show that the spatiotemporal expression pattern of endogenous retinal Mfsd2a, a protein associated with maturation of a functional blood-brain barrier, is consistent with the inhibition of retinal transduction by intravenous AAV9, suggesting that AAV9 crosses the blood-brain barrier by endocytosis.

Systemic delivery of AAV to the mouse retina holds great promise for proof-of-concept gene therapy and basic studies of retinal development. However, results showed that a single injection transduced only a portion of the retina (determined by the age at the time of injection), limiting the effectiveness of this approach. Therefore, the researchers tested whether serial injections at two time points could be used to target a larger portion of the retina (Figure 1). Newborn mice (n = 5) were injected with AAV9-mCherry at P1 and then again with AAV9-GFP at P5. Using this approach, up to 2/3 of the retinal area could be routinely transduced. High-resolution images of the retina (Figure 1b, c) showed that P1 and P5 injections transduced non-overlapping populations of cells. Comparison of retinas injected at P1 alone, at P5 alone, or at both P1 and P5 showed that immune responses (including the production of neutralizing antibodies against the vector capsid) did not appear to interfere with expression of the second vector, likely due to the short time interval between injections (Figure 1d).

Figure 1. Sequential injections of AAV9-mCherry and AAV9-GFP cover an increased area of the retina. (Byrne L C, et al., 2015)