@article{981fdd8dadc749c9b8f0e11487218f8a,
title = "Efficient electroporation of neuronal cells using synthetic oligonucleotides: Identifying duplex RNA and antisense oligonucleotide activators of human frataxin expression",
abstract = "Oligonucleotide drugs are experiencing greater success in the clinic, encouraging the initiation of new projects. Resources are insufficient to develop every potentially important project, and persuasive experimental data using cell lines close to disease target tissue is needed to prioritize candidates. Friedreich{\textquoteright}s ataxia (FRDA) is a devastating and currently incurable disease caused by insufficient expression of the enzyme frataxin (FXN). We have previously shown that synthetic nucleic acids can activate FXN expression in human patient-derived fibroblast cells. We chose to further test these compounds in induced pluripotent stem cell-derived neuronal progenitor cells (iPSC-NPCs). Here we describe methods to deliver oligonucleotides and duplex RNAs into iPSC-NPCs using electroporation. Activation of FXN expression is potent, easily reproducible, and potencies parallel those determined using patient-derived fibroblast cells. A duplex RNA and several antisense oligonucleotides (ASOs) with different combinations of 2′′-methoxyethyl (2′′-MOE), 2′′-fluoro (2′′-F), and constrained ethyl (cEt) were active, providing multiple starting points for further development and highlighting improved potency as an important goal for preclinical development. Our data support the conclusion that ASO-mediated activation of FXN is a feasible approach for treating FRDA and that electroporation is a robust method for introducing ASOs to modulate gene expressions in neuronal cells.",
keywords = "Antisense oligonucleotide, Electroporation, Frataxin, Gene activation, Therapeutic development",
author = "Xiulong Shen and Sharon Beasley and Putman, {Jennifer N.} and Yanjie Li and Prakash, {Thahza P.} and Frank Rigo and Marek Napierala and Corey, {David R.}",
note = "Funding Information: This study was supported R35GM118103 (D.R.C.) from the National Institutes of Health, the Robert A. Welch Foundation I-1244 (D.R.C.), the Friedreich{\textquoteright}s Ataxia Research Alliance, and the Paul D. Wellstone MDCRC Trainee Fellowship Award (X.S.) from UT Southwestern Medical Center. D.R.C. is the Rusty Kelley Professor of Biomedical Science. Work in the Napierala laboratory was supported by National Institutes of Health (R01NS081366) and Muscular Dystrophy Association (MDA418838). The research was also supported by a generous gift from the Doremus family. We thank MaxCyte for providing the MaxCyte STX system and related supplies. Funding Information: This study was supported R35GM118103 (D.R.C.) from the National Institutes of Health, the Robert A. Welch Foundation I-1244 (D.R.C.), the Friedreich{\textquoteright}s Ataxia Research Alliance, and the Paul D. Wellstone MDCRC Trainee Fellowship Award (X.S.) from UT Southwestern Medical Center. D.R.C. is the Rusty Kelley Professor of Biomedical Science. Work in the Napierala laboratory was supported by National Institutes of Health (R01NS081366) and Muscular Dystrophy Association (MDA418838). The research was also supported by a generous gift from the Doremus family. We thank MaxCyte for providing the MaxCyte STX system and related supplies. Author contributions: X.S. helped write the manuscript and executed most of the experiments. J.N.P. assisted with assays. Y.L. and M.N. were responsible for providing neuronal progenitor cells. T.P.P. and F.R. provided most of the compounds tested in this study. S.B. helped develop successful electroporation protocols. D.R.C. supervised experiments and wrote the manuscript. T.P.P. and F.R. are employees of Ionis Pharmaceuticals. S.B. is an employee of MaxCyte Inc. D.R.C., T.P.P., and F.R. have filed patents related to this research. Publisher Copyright: {\textcopyright} 2019 Shen et al.",
year = "2019",
doi = "10.1261/rna.071290.119",
language = "English (US)",
volume = "25",
pages = "1118--1129",
journal = "RNA",
issn = "1355-8382",
publisher = "Cold Spring Harbor Laboratory Press",
number = "9",
}