Exon 44 skipping in Duchenne muscular dystrophy: NS-089/NCNP-02, a dual-targeting antisense oligonucleotide

Naoki Watanabe; Yuichiro Tone; Tetsuya Nagata; Satoru Masuda; Takashi Saito; Norio Motohashi; Kazuchika Takagaki; Yoshitsugu Aoki; Shin'ichi Takeda

doi:10.1016/j.omtn.2023.102034

Exon 44 skipping in Duchenne muscular dystrophy: NS-089/NCNP-02, a dual-targeting antisense oligonucleotide

Mol Ther Nucleic Acids. 2023 Sep 20:34:102034. doi: 10.1016/j.omtn.2023.102034. eCollection 2023 Dec 12.

Authors

Naoki Watanabe¹, Yuichiro Tone¹, Tetsuya Nagata^{2

3}, Satoru Masuda², Takashi Saito², Norio Motohashi², Kazuchika Takagaki¹, Yoshitsugu Aoki², Shin'ichi Takeda²

Affiliations

¹ Discovery Research Laboratories in Tsukuba, Nippon Shinyaku Co., Ltd, Tsukuba, Ibaraki, Japan.
² Department of Molecular Therapy, National Institute of Neuroscience, National Center of Neurology and Psychiatry (NCNP), Kodaira, Tokyo, Japan.
³ Department of Neurology and Neurological Science, Graduate School of Medicine, Tokyo Medical and Dental University, Tokyo, Japan.

Abstract

Exon-skipping therapy mediated by antisense oligonucleotides is expected to provide a therapeutic option for Duchenne muscular dystrophy. Antisense oligonucleotides for exon skipping reported so far target a single continuous sequence in or around the target exon. In the present study, we investigated antisense oligonucleotides for exon 44 skipping (applicable to approximately 6% of all Duchenne muscular dystrophy patients) to improve activity by using a novel antisense oligonucleotide design incorporating two connected sequences. Phosphorodiamidate morpholino oligomers targeting two separate sequences in exon 44 were created to target two splicing regulators in exon 44 simultaneously, and their exon 44 skipping was measured. NS-089/NCNP-02 showed the highest skipping activity among the oligomers. NS-089/NCNP-02 also induced exon 44 skipping and dystrophin protein expression in cells from a Duchenne muscular dystrophy patient to whom exon 44 skipping is applicable. We also assessed the in vivo activity of NS-089/NCNP-02 by intravenous administration to cynomolgus monkeys. NS-089/NCNP-02 induced exon 44 skipping in skeletal and cardiac muscle of cynomolgus monkeys. In conclusion, NS-089/NCNP-02, an antisense oligonucleotide with a novel connected-sequence design, showed highly efficient exon skipping both in vitro and in vivo.

Keywords: Duchenne muscular dystrophy; MT: Oligonucleotides: Therapies and Applications; NS-089/NCNP-02; antisense oligonucleotide; antisense therapeutics; dual-targeting; dystrophin; exon 44; exon skipping; morpholino; splice modulation.