Abstract
Purpose :
In glaucoma, although reasonable IOP lowering therapy exists, it is not always effective and is not without the side effects making this disease a leading cause of blindness in the US and throughout the world. Dysfunctional mitochondria often associate with glaucomatous RGCs which may contribute to or be part of the disease pathology. Thus, therapeutic agent that would restore mitochondrial health in RGCs could provide glaucoma neuroprotection.
Methods :
We have used well characterized stem cell differentiated hRGCs and small molecule screening to identify drugs that restore mitochondrial health. We tested identified hit compound for its direct effect on pure hRGCs with glaucomatous OPTN-E50K mutation and measured mitochondrial membrane polarity (n= 30-40 cells), cell viability, and excitotoxicity (n= 4-6 biological repeats). Mitochondrial mass was measured by confocal immunofluorescence (Tom20) and western blot (Tom70). Activation of mitochondrial biogenesis master regulator PGC1α was measured by its nuclear localization. For in-vivo efficacy against direct RGC injury, mice were treated intraperitoneally daily 5 days before optic nerve crush (ONC) until the measurements were done. We measured RGC numbers in the retina, axons in the optic nerve, and vision by optokinetic response (OKR) between day 6 to 21 post ONC. Naïve, ONC and ONC treated groups (n = 4) were compared. Two-way ANOVA with Tukey’s post-hoc test was used for statistics.
Results :
From screening, we identified serotonin receptor 5-HT1A antagonist ADC1 that restored mitochondrial membrane polarity, reduced excitotoxicity and increased cell viability for E50K hRGCs. 5-HT1A is an inhibitory GPCR and ADC1 treatment enhanced cAMP level, PGC1α activation and mitochondrial mass following classic GPCR negative feedback signalling. Remarkably, ADC1 treatment protected RGC bodies in the 10 days post ONC retina (3771.6±242.5/mm2) to the level of naïve (4205.5±110.1/mm2) mouse, while untreated ONC mice suffered severe RGC loss (885.62±50.8/mm2). We observed similar protection for RGC axons, visual acuity, and contrast sensitivity between day 6 to 21 post ONC.
Conclusions :
Here we have discovered a pharmacological agent that provides direct neuroprotection to glaucomatous pure human RGCs and against optic nerve injury in mouse. Thus, likely to provide glaucoma neuroprotection independent of risk factors such as high IOP.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.