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Ahmed M Alkaliby, Jaime Toledo Corral, Paula Sakemi Fukuhara, Noor-Ul-Ain Shekoh, Cristina M Kenney, Baruch D Kuppermann; Effect of bevacizumab and aflibercept on wet AMD cybrid cells. Invest. Ophthalmol. Vis. Sci. 2020;61(7):5398.
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© ARVO (1962-2015); The Authors (2016-present)
To evaluate the effect of bevacizumab and aflibercept on reactive oxygen species (ROS) related to angiogenesis pathways in wet AMD cybrid cells
Cybrids were created by fusing ARPE-19 Rho0 cells that were depleted of their mitochondrial DNA (mtDNA), with platelets isolated from wet AMD patients (n=7). These cybrids have the same nuclear genome but different mtDNA. The cybrid cell lines were: 14-136, 14-145, 14-146, 15-147, 15-151, 15-157 and 15-159. Cybrids were cultured in 24-wells plates for 48 hours, and then treated with bevacizumab and aflibercept at 1X, 2X, 4X or 10X concentrations of the clinical intravitreal dose (equivalent to 0.05 ml of drug distributed in 4 ml of vitreous volume). After 24 hours, ROS production levels were measured using the fluorescent dye 2’7’-dichlodihydrofluorescein diacetate (H2DCFDA) reagent and fluorescent plate reader. Results were normalized to untreated cells. All the experiments were repeated at least three times and p value<0.005 was considered statistically significant.
Cybrid 14-136 showed a progressive reduction in the ROS levels at all concentrations of bevacizumab (p<0.0001) and aflibercept (1X,p=0.0005; 2X,p=0.0011; 4X,p<0.0001; 10X,p<0.0001). In contrast, Cybrid 14-145 showed an increase in ROS levels with bevacizumab (1X,p=0.0012; 2X,p=0.0065; 4X,p<0.0001; 10X,p<0.0001) and a reduction with aflibercept 4X (p=0.0023) and 10X (p=0.0117) concentrations. Cybrids 15-151 also showed decreased of ROS levels in all concentrations when treated with aflibercept(1X,4X,10X, p<0.0001; 2X,p=0.0245), but no significant results with bevacizumab. Cybrids 15-157 had a significant reduction with aflibercept in 4X (p=0.0096) and 10X (p=0.027) but were not significant in none of bevacizumab groups. No significant change in ROS levels was found at any of the rest of drug concentrations.
The AMD cybrids, which had identical nuclei but mitochondria from different AMD patients, produced differential ROS levels after treatment with bevacizumab and aflibercept. This supports the hypothesis that (a) mitochondria from AMD patients can have unique responses to drugs and (b) AMD cybrids are an excellent model to screen responses to drugs. Further investigation is needed to better understand the interaction between anti-VEGF drugs and mitochondria as well as its clinical significance in wet AMD patients.
This is a 2020 ARVO Annual Meeting abstract.
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