Abstract
Purpose :
Purpose: Hydroxychloroquine (HCQ) was originally developed as antimalarial drug and is now, due to its anti-inflammatory properties, primarily used for treating autoimmune disorders. One side effect of long term/high dosage HCQ use is retinopathy, specifically manifesting as “bull’s eye” maculopathy at advanced disease stages. HCQ is known to exhibit high melanin binding affinity, leading to its accumulation in pigmented tissues such as skin and retinal pigment epithelium (RPE), likely leading to RPE and retinal toxicity. HCQ’s ability to increase lysosomal pH is thought to be the main subcellular reason for its toxicity. The purpose of this study was to develop an in vitro platform to discover HCQ genetic sensitivity and develop potential therapies.
Methods :
Methods: iRPE were treated with HCQ (10uM-640uM) and characterized for viability assessed by ATP assay; lysosomal pH measured using the pH-sensitive LysoSensor™ dye, cell layer integrity using transepithelial resistance (TER) measurements, and photoreceptor outer segment (POS) phagocytosis and degradation in a flow cytometry-based assay.
Results :
Results: HCQ (20μM) significantly elevated iRPE lysosomal pH and led to a marked decrease in POS degradation while RPE monolayer integrity was compromised at high concentrations only (>200μM). iRPE viability was significantly reduced at concentrations higher than 320μM.
Conclusions :
Conclusions: Low HCQ treatment concentrations were sufficient to significantly increase iRPE lysosomal pH and abolish POS degradative capacity almost completely. HCQ-effects on cytotoxicity and cell layer integrity were only observed at high concentrations which is why we hypothesize that HCQ induced retinopathy is directly related to loss of RPE degradative function caused by lysosomal impairment and subsequent death of photoreceptors due to RPE inability to support photoreceptor survival. As a next step, we will attempt to understand genetic risk associated with HCQ sensitivity in iRPE cells.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.