Allele-Specific Knockdown of ALS-Associated Mutant TDP-43 in Neural Stem Cells Derived from Induced Pluripotent Stem Cells

作者：Nishimura Agnes L; Shum Carole; Scotter Emma L; Abdelgany Amr; Sardone Valentina; Wright Jamie; Lee Youn Bok; Chen Han Jou; Bilican Bilada; Carrasco Monica; Maniatis Tom; Chandran Siddharthan; Rogelj Boris; Gallo Jean Marc; Shaw Christopher E^*

来源：PLos One, 2014, 9(3): e91269.

DOI：10.1371/journal.pone.0091269

TDP-43 is found in cytoplasmic inclusions in 95% of amyotrophic lateral sclerosis (ALS) and 60% of frontotemporal lobar degeneration (FTLD). Approximately 4% of familial ALS is caused by mutations in TDP-43. The majority of these mutations are found in the glycine-rich domain, including the variant M337V, which is one of the most common mutations in TDP-43. In order to investigate the use of allele-specific RNA interference (RNAi) as a potential therapeutic tool, we designed and screened a set of siRNAs that specifically target TDP-43(M337V) mutation. Two siRNA specifically silenced the M337V mutation in HEK293T cells transfected with GFP-TDP-43(wt) or GFP-TDP-43(M337V) or TDP-43 C-terminal fragments counterparts. C-terminal TDP-43 transfected cells show an increase of cytosolic inclusions, which are decreased after allele-specific siRNA in M337V cells. We then investigated the effects of one of these allele-specific siRNAs in induced pluripotent stem cells (iPSCs) derived from an ALS patient carrying the M337V mutation. These lines showed a two-fold increase in cytosolic TDP-43 compared to the control. Following transfection with the allele-specific siRNA, cytosolic TDP-43 was reduced by 30% compared to cells transfected with a scrambled siRNA. We conclude that RNA interference can be used to selectively target the TDP-43(M337V) allele in mammalian and patient cells, thus demonstrating the potential for using RNA interference as a therapeutic tool for ALS.

• 出版日期2014-3-20