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Abstract #123212 Published in IGR 25-1
Comparison of SNCG and NEFH Promoter-Driven Expression of Human SIRT1 Expression in a Mouse Model of Glaucoma
O'Neill N; Meng M; Chaqour B; Dine K; Sarabu N; Pham JC; Shindler KS; Ross AGTranslational vision science & technology 2024; 13: 37
PURPOSE: Adeno-associated virus (AAV) demonstrates promise in delivering therapeutic genes to retinal ganglion cells (RGCs). Delivery of neuroprotective genes is constrained by packaging size and/or cell selectivity. This study compares the ability of the RGC-selective gamma-synuclein (SNCG) promoter and the smaller RGC-selective neurofilament heavy chain (NEFH) promoter, as well as portions of the RGC-selective atonal bHLH transcription factor 7 (ATOH7) enhancer, to drive gene expression in RGCs. METHODS: AAV2 constructs with green fluorescent protein (GFP) or human sirtuin 1 (hSIRT1) driven by cytomegalovirus (CMV) enhancer and NEFH promoter (AAV2-eCMV-NEFH) or distal active sequences of the ATOH7 enhancer (DiATOH7) with the SNCG promoter (AAV2-DiATOH7-SNCG) were intravitreally injected into C57BL/6J mice. RGCs were immunolabeled with Brn3a antibodies and counted. AAV constructs with the utmost transduction efficiency were used to test the therapeutic efficacy of the hSIRT1 gene in 12-week-old C57BL/6J mice subjected to microbead (MB)-induced intraocular pressure (IOP) elevation. Visual function was measured using optokinetic responses (OKRs). RESULTS: The eGFP transduction efficiency of AAV2-eCMV-NEFH was similar to that of AAV2-eCMV-SNCG and AAV2-DiATOH7-SNCG. When combined with the SNCG promoter, a larger ATOH7 enhancer was less efficient than the shorter DiATOH7 enhancer. Similarly, the hSIRT1 efficiency of AAV2-eCMV-NEFH was similar to that of AAV2-eCMV-SNCG. The latter two vectors were equally efficient in increasing RGC survival and improving visual function in the mouse model of MB-induced IOP elevation. CONCLUSIONS: SNCG and NEFH promoters represent two equally efficient and comparable RGC selective promoter sequences; however, the NEFH promoter offers a smaller packaging size. TRANSLATIONAL RELEVANCE: Smaller enhancer-promoter combinations can be used to deliver larger genes in human cells and for treatment in optic neuropathies including glaucoma.
Department of Ophthalmology, University of Pennsylvania, Philadelphia, PA, USA.
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