siRNA Targeting Caspase 2 Protects Retinal Ganglion Cells in the Rat Models of Optic Nerve Injury

Abstract

Purpose: : To assess the efficacy of short interfering RNA (siRNA)-mediated knockdown of pro-apoptotic gene Caspase 2 in the survival of adult rat retinal ganglion cells (RGCs) after optic nerve injury. Therapeutic neuroprotective agents have the potential in the treatment of ocular diseases that were heretofore untreatable or difficult to treat, including disorders primarily involving death of photoreceptors (such as in AMD, retinitis pigmentosa and retinal detachment) or RGC (such as in acute or chronic optic neuropathies). Caspase2 was identified as a primary target for inhibition to prevent RGC apoptosis in optic nerve crush and axotomy models in rats. Whereas Caspase2 was predominantly expressed and activated in RGC, the primary localization of Caspase 3 was in horizontal and bipolar cells of the inner nuclear layer.

Methods: : siRNA delivery to the retina was confirmed by quantitation and in situ hybridization. For efficacy models, RGCs were labeled by application of the retrograde tracer, Fluorogold. Optic nerve injury was performed one week later. Intraocular injections of proprietary chemically stabilized anti-Caspase 2 or control siRNAs were performed at the time of injury by infusion into the vitreous chamber. The density of surviving RGCs was quantified at different time points after optic nerve injury by counting Fluorogold-labeled neurons in retinal wholemounts. For longer time points, the animals received 2^nd siRNA injection.

Results: : siRNA was detectable in the retina for up to one week after injection to injured eyes and was localized to RGC. siRNA targeting Caspase 2 significantly enhanced RGC survival compared to controls. There was no difference in RGC survival in injured eyes treated with either control siRNAs or with PBS intravitreal injections.

Conclusions: : These results demonstrate that nuclease-stable, modified siRNA targeting Caspase 2 has sustained neuroprotective effects on RGC survival following optic nerve injury.

Keywords: ganglion cells • neuroprotection • RNAi