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bioRxiv · 10.1101/2020.06.18.159533

A Novel Therapeutic Approach to Corneal Alkaline Burn Model by Targeting Fidgetin-like 2, a Microtubule Regulator

Wang, J.·Dey, A.·Kramer, A.·Miao, Y.·Liu, J.·Baker, L.·Friedman, J.·Nacharaju, P.·Chuck, R. S.·Zhang, C.·Sharp, D.
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Abstract

Purpose To determine the efficacy of nanoparticle-encapsulated FL2 siRNA (FL2-NPsi), a novel therapeutic agent targeting the Fidgetin-like 2 (FL2) gene, for the treatment of corneal alkaline chemical injury.Methods Eighty 12-week-old, male Sprague-Dawley rats were divided evenly into 8 treatment groups: prednisolone, empty nanoparticles, control-NPsi (1 μM, 10 μM, 20 μM) and FL2-NPsi (1 μM, 10 μM, 20 μM). An alkaline burn was induced onto the cornea of each rat, which was then treated for 14 days according to group assignment. Clinical (N=10 per group), histopathologic (N=6 per group), and immunohistochemical (N=4 per group) analyses were conducted to assess for wound healing. FL2-NPsi-mediated knockdown of FL2 was confirmed by in vitro qPCR. Toxicity assays were performed to assess for apoptosis (TUNEL assay, N=3 per group) and nerve damage (whole mount immunochemical staining, N=2 per group). Statistical analyses were performed using student’s t-test and ANOVA.Results Compared to controls, FL2-NPsi-treated groups demonstrated enhanced corneal wound healing, with the 10 and 20 μM FL2-NPsi-treated groups demonstrating maximum rates of corneal re-epithelialization (p=0.0003 at Day 4 and p<0.0001 at Day 8) as assessed by ImageJ software, enhanced corneal transparency, and improved stromal organization on histology. Immunohistochemical analysis of vascular endothelial cells, macrophages, and neutrophils did not show significant differences between treatment groups. FL2-NPsi was not found to be toxic to nerves or induce apoptosis (p=0.917).Conclusion Dose-response studies found both 10 and 20 μM FL2-NPsi to be efficacious in this rat model. FL2-NPsi may offer a novel treatment for corneal alkaline chemical injuries.Competing Interest StatementDr. Wang, Dr. Kramer, Dr. Baker, and Dr. Dey are employees of MicroCures, Inc. Dr. Sharp is co-founder and Chief Scientific Officer for MicroCures, Inc and is the inventor of U.S. Patent #20130022667 entitled "Fidgetin-like 2 as a target to enhance wound healing,” which has been licensed by MicroCures. Dr. Chuck is a scientific advisor for MicroCures, Inc. Dr. Friedman holds stock in MicroCures. No other author has a conflict of interest.View Full Text

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Wang, J., Dey, A., Kramer, A., Miao, Y., Liu, J., Baker, L., Friedman, J., Nacharaju, P., Chuck, R. S., Zhang, C., Sharp, D.. 2020-06-19. A Novel Therapeutic Approach to Corneal Alkaline Burn Model by Targeting Fidgetin-like 2, a Microtubule Regulator. https://doi.org/10.1101/2020.06.18.159533

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