論文種別	原著(症例報告除く)
言語種別	英語
査読の有無	査読あり
表題	Novel EGFP reporter cell and mouse models for sensitive imaging and quantification of exon skipping.
掲載誌名	正式名：Scientific reports
掲載区分	国外
巻・号・頁	10(1),pp.10110-10110
著者・共著者	Yuko Hara,Yoshitaka Mizobe,Yukiko U Inoue,Yasumasa Hashimoto,Norio Motohashi,Yoshiaki Masaki,Kohji Seio,Shin'ichi Takeda,Tetsuya Nagata,Matthew J A Wood,Takayoshi Inoue,Yoshitsugu Aoki
発行年月	2020/06/22
概要	Duchenne muscular dystrophy (DMD) is a fatal X-linked disorder caused by nonsense or frameshift mutations in the DMD gene. Among various treatments available for DMD, antisense oligonucleotides (ASOs) mediated exon skipping is a promising therapeutic approach. For successful treatments, however, it is requisite to rigorously optimise oligonucleotide chemistries as well as chemical modifications of ASOs. To achieve this, here, we aim to develop a novel enhanced green fluorescence protein (EGFP)-based reporter assay system that allows us to perform efficient and high-throughput screenings for ASOs. We design a new expression vector with a CAG promoter to detect the EGFP fluorescence only when skipping of mdx-type exon 23 is induced by ASOs. Then, an accurate screening was successfully conducted in C57BL/6 primary myotubes using phosphorodiamidate morpholino oligomer or locked nucleic acids (LNA)/2'-OMe mixmers with different extent of LNA inclusion. We accordingly generated a novel transgenic mouse model with this EGFP expression vector (EGFP-mdx23 Tg). Finally, we confirmed that the EGFP-mdx23 Tg provided a highly sensitive platform to check the effectiveness as well as the biodistribution of ASOs for exon skipping therapy. Thus, the assay system provides a simple yet highly sensitive platform to optimise oligonucleotide chemistries as well as chemical modifications of ASOs.
DOI	10.1038/s41598-020-67077-4
PMID	32572084