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Sonali Nashine, Marilyn Chwa, Shari Atilano, Sudhakar R. Subramaniam, Howard Federoff, Anthony B Nesburn, Baruch D Kuppermann, Cristina M Kenney; PU-91, a mitochondria-stabilizing drug, rescues AMD ARPE-19 cybrid cells; implications for macular degeneration therapeutics. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3268.
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The dry form of age-related macular degeneration (AMD) characterized by loss of retinal pigment epithelial (RPE) cells and subsequent geographic atrophy, accounts for approximately 80-90% of AMD cases, and currently has no available FDA-approved treatments. Herein, we tested the hypothesis that PU-91, an FDA-approved mitochondria-stabilizing drug, protects RPE cells in an in vitro macular degeneration model.
The in vitro transmitochondrial cybrid model was created by fusing blood platelets obtained from AMD patients with human ARPE-19 cells lacking mtDNA (Rho0). All cybrids had identical nuclei but differed in mtDNA content. Cybrids (n=4-5) were treated with 50 µM PU-91 drug for 72 hr, DNA and RNA were extracted, and RT-qPCR, cell viability assay (MTT) and mtDNA copy number assay were performed. Statistical differences were measured using Student’s t-test, and significance was determined at P<0.05. Data were normalized to the untreated AMD group, which served as control.
Treatment with PU-91 significantly increased the gene expression of: 1) PGC-1α, a master regulator of mitochondrial biogenesis, by 208% (P=0.0018) and 2) Mitochondrial biogenesis markers, i.e., NRF-1 by 46% (P=0.04), NRF-2 by 38% (P=0.04), PPAR-α by 19% (P=0.02), and PPAR-γ by 32% (P=0.03). The MT-RNR2 (mt 16S rRNA gene) gene which codes for the cyto-protective mitochondrial-derived peptides (MDPs) such as Humanin, was increased by 104% (P=0.039) in PU-91-treated AMD cybrids. PU-91 down-regulated the apoptosis genes i.e., Caspase-3 by 34% (P=0.02) and BAX by 20% (P=0.01). Furthermore, PU-91-treated cybrids had a higher number of viable cells and higher mtDNA copy number compared to their untreated counterparts (P<0.05).
In summary, the PU-91 drug rescues AMD cybrids by: 1) inducing mitochondrial biogenesis (PGC-1α, NRFs, PPARs), 2) decreasing apoptosis (Caspase-3, BAX), and by 3) inducing transcription of the MDP-coding gene (MT-RNR2). It is likely that PU-91 improves cell viability by triggering production of MDPs which are known to be cyto-protective in AMD. To conclude, our results present novel findings that identify PU-91 to be a promising prospect as therapeutics for AMD, a devastating retinal degenerative disease that currently has no viable treatment options. PU-91 being an FDA-approved drug, it will be a smoother bridge from lab bench to clinic.
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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