Effect of Bevacizumab, Ranibizumab and Aflibercept on retinal pigment epithelium phagocytosis

Abstract

Purpose : Controversy exists regarding the safety of vascular endothelial growth factor (VEGF) inhibitors in the retinal pigment epithelium (RPE) and it is currently unknown whether the development of geographic atrophy in patients with age-related macular degeneration (AMD) treated with these drugs is a toxic effect, or otherwise responds to the natural course of the disease. The aim of this work was to evaluate if phagocytosis, one of the most important functions of the RPE, is affected by Bevacizumab, Ranibizumab and Aflibercept and their effect on the main molecular regulators of that function.

Methods : Assays were performed in vitro using the human ARPE-19 cell line. Polarized cell monolayers were treated with clinical concentrations of Bevacizumab, Ranibizumab and Aflibercept during 24 hours and phagocytosis of fluorescent microspheres was assessed at 48 hours by confocal microscopy. We also evaluated the effect of anti-VEGFs in MerTK, phospho-MerTK and a5bv integrin by western blot, as mediators of the phagocytosis process. Moreover, the structure of the cytoskeleton labelled with fluorescent phalloydin (F-actin) was also analyzed by confocal microscopy.

Results : Compared to untreated control cells, exposure to VEGF produced a statistically significant decrease in ARPE-19 cells phagocytosis of fluorescent microspheres (p=0.036). On the contrary, exposure to Bevacizumab and Aflibercept (p=0.016) but not for Ranibizumab (p=0.286) increased the phagocytosis. The expression of MerTK, phospho-MerTK or a5bv integrin was not modified by VEGF treatment and also by any of the anti-VEGFs drugs compared to untreated cells. However, exposure to VEGF produced a disruption of the normal distribution of actin F-actin cytoskeleton fibers that was not observed in cells treated with anti-VEGFs or untreated controls.

Conclusions : Our results suggest that inhibition of VEGF with antiangiogenics treatments increases phagocytosis in RPE cells, while exposure to VEGF has the opposite effect. These results were not explained by differences in the expression of the main molecular regulators of phagocytosis; however, our experiments suggest that they could be related to changes in the normal architecture of the cytoskeleton that was found to be disrupted by exposure to VEGF.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.