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Kevin Schneider, Marilyn Chwa, Adam BenMohamed, Cristina Kenney; Effects of Bevacizumab and Risuteganib on ARPE19 AMD Cybrid cells. Invest. Ophthalmol. Vis. Sci. 2019;60(9):1934.
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The primary clinical treatment for exudative (wet) age-related macular degeneration (AMD) is intravitreal injections of ani-VEGF drugs such as Avastin (bevacizumab). Such treatments have been effective, however long-term side effects are a concern. Risuteganib holds promise to control angiogenesis and VEGF expression through targeting of integrin receptor signaling. This study investigates the effects of bevacizumab and risuteganib on RPE cybrid cells generated using the mitochondria of AMD patients. The transmitochondrial cybrid model allows a “personalized” approach in examining drug effects. We hypothesize that treatment with these drugs will produce differential effects on cell health and gene expression.
We utilize a transmitochondrial cybrid model in which platelets, containing mitochondria but no nuclei, from clinically well-characterized patients, are isolated and fused with human retinal pigmented epithelial cells (ARPE-19) lacking mitochondria (Rho0). Cybrid cell lines were generated from age matched AMD patients (n=7). Cells were treated with 1x clinical dose drug for 48 hours. Cell health and apoptosis assay performed using Incucyte Live Cell Imager. RNA isolation with PureLink, CDNA creation with Vilo IV and QPCR with PowerUp Syber Green. Statistical analyses were by paired t-test.
AMD cybrids treated with risuteganib exhibited no statistically significant change in Nuclight or activated caspase 3/7 staining compared to untreated cells (high intensity indicate dying cells). However, bevacizumab had significant increases in Nuclight and activated caspase 3/7 (1.7 fold and 1.95 fold, p=<0.005). Risuteganib decreased gene expression of VEGFA (0.8 fold, p=0.04) while bevacizumab increased expression (1.272 fold, p=0.05). Risuteganib decreased expression of ITGB1 (0.87 fold p=0.01) an integrin gene associated with angiogenesis. Finally, risuteganib decreased expression of the pro-apoptotic genes BAX and BCL2L13 (0.72 fold, p=0.048 and 0.81 fold, p=0.05). Treatment with bevacizumab had significant decrease in only BAX gene expression. (0.84 fold, p=0.009)
Our results with bevacizumab and risuteganib support our hypothesis that the drugs utilized for the treatment of macular degeneration can influence cellular health and gene expression. Data suggests that risuteganib is well tolerated when compared with existing treatments, and potentially could serve as a powerful alternative treatment.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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