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Jaime Toledo Corral, Mohamed Mohamed, Marilyn Chwa, Shari Atilano, Cristina M Kenney, Baruch D Kuppermann; Effect of aflibercept and ranibizumab on reactive oxygen species (ROS) production in wet AMD cybrid cells. Invest. Ophthalmol. Vis. Sci. 2018;59(9):1462.
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To evaluate the effect of aflibercept and ranibizumab on the oxidative stress species related to angiogenesis pathways in cybrid cells.
Cybrids were created by fusing Rho0 cells, ARPE-19 cells depleted of mitochondrial DNA (mtDNA), with platelets isolated from wet AMD patients n=7. These cybrids have the same nuclear genome but different mtDNA. Cybrids were cultured for 48 hours, then treated with aflibercept or ranibizumab at 1X, 2X, 4X or 10X concentrations of the clinical intravitreal dose (equivalent to 0.05 ml of drug in 4 ml of vitreous volume). After 24 hours, ROS production levels were measured using the H2DCFDA reagent. Results were normalized to untreated cells. All the experiments were repeated three times and unpaired t test was used to analyze data.
Comparing different concentrations of the same drug: cybrids 14-145 showed an increase in ROS levels with ranibizumab (2X 20.3%, 4X 17.6%, 10X 21.2% p values <0.04) and a reduction with aflibercept (4X 28.6%, 10X 28%, p values <0.02). Cybrids 14-136 (ranibizumab 1X 4.5%, 2X 9.7%, 4X 8.9% and 10X 9.9%, p values <0.02; aflibercept 1X 16.5%, 2X 18.4%, 4X 27.1% and 10X 24.4%, p values <0.01) and 15-151 (ranibizumab 1X 8.4%, 2X 9%, 4X 13.4% and 10X 12.5%, p values <0.03; aflibercept 1x 14.5%, 2X 12.7%, 4X 26.2% and 10X 17.4%, p values <0.03) showed a progressive reduction of ROS levels with increasing concentrations of both drugs; Cybrid 15-157 showed a reduction with aflibercept (4X 13.51%, 10X 12.61% p values <0.03) while cybrids 14-146, 15-147 and 15-159 ROS levels were not significantly different between concentrations.Comparing ranibizumab to aflibercept at each concentration: Cybrid 14-136 showed lower ROS levels at 1X, 4X and 10X (p<0.01); cybrid 14-147 showed lower levels at 2X (p<0.05); cybrid 15-151 showed lower levels at 4X (p<0.04), cybrid 14-145 showed lower levels at all aflibercept concentrations (p<0.04). Cybrid 14-146 showed lower levels at 2X (p<0.03) ranibizumab concentration. No significant change in ROS levels was found at any other drug concentrations.
Since all cybrids had identical nuclei but different mitochondria, we propose that the difference in ROS levels after anti-VEGF treatment is related to this variable. 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 an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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