Identification of proteins associated with ATXN8OS-related amyotrophic lateral sclerosis and treatment of iPS cell models

• Project/Area Number: 19K07984
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 52020:Neurology-related
• Research Institution: Kindai University
• Principal Investigator: Hirano Makito 近畿大学, 医学部, 教授 (50347548)
• Co-Investigator(Kenkyū-buntansha): 竹原 俊幸 近畿大学, 大学病院, 助教 (60580561)
• Project Period (FY): 2019-04-01 – 2022-03-31
• Project Status: Completed (Fiscal Year 2021)
• Budget Amount: ¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000); Fiscal Year 2021: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2020: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2019: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
• Keywords: 核酸医薬品 / 筋萎縮性側索硬化症 / iPS細胞 / 非翻訳領域反復配列延長 / 運動ニューロン / ATXN8OS / SCA8 / ATXN8 / 非翻訳領域反復配列 / iPS

Outline of Research at the Start

筋萎縮性側索硬化症(ALS)は、運動ニューロンの変性により筋肉が徐々に萎縮する予後不良の神経難病であり、原因・根治療法は確立されていない。欧米で多いnon-coding RNA(ncRNA)に関連するC9ORF72遺伝子の反復配列延長は、日本では、まれである。私たちは、ncRNAに関連する脊髄小脳萎縮症８型の原因遺伝子ATXN8OSの反復配列がALSの約３％で延長していることを発見した(Neurol Genet 2018;4:e252)。本研究では、ATXN8OS関連ALSの病態関連蛋白の同定とiPS細胞由来の運動ニューロンモデルの構築と治療法開発を目指し、病理組織も用いて結果を検証する。

Outline of Final Research Achievements

This study was aimed to elucidate the pathogenesis of ATXN8OS-related amyotrophic lateral sclerosis (ALS), to establish an iPS cell-derived motor neuron model, and to develop a therapeutic approach. We received tissue samples of pathologically confirmed ALS from the Brain Bank and analyzed their DNA. We identified an ATXN8OS mutation in one patient. In this patient, TDP43 aggregation was found in the anterior horn motoneurons of the spinal cord. In addition, induced pluripotent stem (iPS) cells from another ATXN8OS-related ALS patient were induced into motor neurons. A substantial decrease in the number of viable cells was observed, as compared to normal controls. In situ hybridization using fluorescent probes to detect repetitive sequences revealed intracellular aggregates, which were co-localized with TDP43. For therapeutic intervention, strand-specific siRNAs were expressed, and two siRNAs significantly increased the number of viable cells in patient-derived motor neurons.

Academic Significance and Societal Importance of the Research Achievements

今回の研究成果により、ATXN8OS関連ALSは、古典的ALSと共有するTDP43が介在する病態機序が考えられた。また、患者iPS由来運動ニューロンでは、細胞生存率が低下しており、細胞内凝集も再現されたことから、この細胞はALSのモデル細胞として使用できると考えられた。また、遺伝子発現低下を目指したsiRNA導入にて細胞死が減少することから、ATXN8OS変異による毒性が病態に関与していることが示唆された。さらに、モデル細胞をsiRNAにより治療させることができる可能性を示した点で、今後の治療開発に対して、重要な知見であると考えられる。

Research Products

• [Journal Article] Identification of a novel mutation and genotype-phenotype relationship in MEGF10 myopathy (2022)
  • Author(s): Fujii Kanako、Hirano Makito、Terayama Atsushi、Inada Rino、Saito Yoshihiko、Nishino Ichizo、Nagai Yoshitaka
  • Journal Title: Neuromuscular Disorders Volume: - Issue: 5 Pages: 436-440
  • DOI: 10.1016/j.nmd.2022.01.009
  • Peer Reviewed
• [Journal Article] Phenotypic and molecular diversities of spinocerebellar ataxia type 2 in Japan (2021)
  • Author(s): Inada R, Hirano M, Oka N, Samukawa M, Saigoh K, Suzuki H, Udaka F, Hashiguchi A, Takashima H, Hamada Y, Nakamura Y, Kusunoki S.
  • Journal Title: J Neurol. Volume: Feb 24 Issue: 8 Pages: 2933-2942
  • DOI: 10.1007/s00415-021-10467-z
  • Peer Reviewed / Open Access
• [Journal Article] Rasagiline monotherapy improves swallowing in patients with Parkinson's disease (2020)
  • Author(s): Hirano Makito、Isono Chiharu、Samukawa Makoto、Fukuda Kanji、Kusunoki Susumu
  • Journal Title: Parkinsonism & Related Disorders Volume: 78 Pages: 98-99
  • DOI: 10.1016/j.parkreldis.2020.07.010
  • Peer Reviewed
• [Journal Article] Effects of the rotigotine transdermal patch versus oral levodopa on swallowing in patients with Parkinson's disease (2019)
  • Author(s): Hirano M, Isono C, Fukuda K, Ueno S, Nakamura Y, Kusunoki S
  • Journal Title: J Neurol Sci Volume: - Pages: 5-10
  • DOI: 10.1016/j.jns.2019.07.003
  • Peer Reviewed
• [Journal Article] Pathological Findings in Male Patients With Anti-N-methyl-d-Aspartate Receptor Encephalitis (2019)
  • Author(s): Hirano M1,2, Itoh T3, Fujimura H4, Inoue K4, Samukawa M1,2, Nose K5, Sakamoto H2, Maekura S6, Ueno S1,2, Satou T3, Nishioka T4, Kusunoki S, Nakamura Y
  • Journal Title: J Neuropathol Exp Neurol Volume: - Issue: 8 Pages: 735-741
  • DOI: 10.1093/jnen/nlz052
  • Peer Reviewed / Open Access
• [Journal Article] 脊髄小脳変性症の遺伝子診断と近畿圏における病型 (2019)
  • Author(s): 仲間美奈、西郷和真、平野牧人、濱田征宏、金城ちなつ、寒川 真、長谷川隆典、北口正孝、三井良之、巽 順子、田村和朗、楠 進
  • Journal Title: 日本カウンセリング学会誌 Volume: 40 Pages: 95-100
  • Peer Reviewed
• [Presentation] Gene analyses for spinocerebellar ataxia type 2 identify neuropathy phenotype in Japan. (2022)
  • Author(s): Rino Inada, Makito Hirano, Nobuyuki Oka, Makoto Samukawa, Kazumasa Saigoh, Hidekazu Suzuki, Fukashi Udaka, Akihiro Hashiguchi, Hiroshi Takashima, Yukihiro Hamada, Yusaku Nakamura, Susumu Kusunoki
  • Organizer: 第63回 日本神経学会総会
• [Presentation] Phenotypic variation and therapeutic strategy of SCA8-associated amyotrophic lateral sclerosis (2022)
  • Author(s): Makito Hirano, Toshiyuki Takehara, Shigeo Murayama, Yuishin Izumi, Makoto Samukawa, Tomoyasu Matsubara, Yuko Saito, Kazumasa Saigoh, Yusaku Nakamura, Kanji Fukuda, Susumu Kusunoki, Yoshitaka Nagai
  • Organizer: 第63回 日本神経学会総会
• [Presentation] 脊髄小脳失調症8型遺伝子異常を有する筋萎縮性側索硬化症患者の嚥下機能 (2021)
  • Author(s): 磯野千春、平野牧人、福田寛二、寒川 真、西郷和真、中村雄作、楠 進
  • Organizer: 第62回 日本神経学会総会
• [Presentation] パーキンソン病と関連疾患における新しい分子病態と嚥下障害～治療と対策～ (2021)
  • Author(s): 平野牧人
  • Organizer: 第62回 日本神経学会総会
  • Invited
• [Presentation] 神経筋疾患の嚥下障害 ～基本から最新知見まで～ (2021)
  • Author(s): 平野牧人
  • Organizer: 第61回日本神経学会
  • Invited
• [Presentation] The first patient with pathologically-definite ATXN8OS-associated amyotrophic lateral sclerosis (2020)
  • Author(s): Makito Hirano, Shigeo Murayama, Yuishin Izumi, Makoto Samukawa, Tomoyasu Matsubara, Kazumasa Saigoh, Yusaku Nakamura, Susumu Kusunoki
  • Organizer: 第61回日本神経学会
• [Presentation] NON-CODING REPEAT EXPANSIONS IN THE ATXN8OS GENE ARE IDENTIFIED IN JAPANESE PATIENTS WITH AMYOTROPHIC LATERAL SCLEROSIS. (2019)
  • Author(s): Makito Hirano, Makoto Samukawa, Chiharu Isono, Kazumasa Saigoh, Yusaku Nakamura, Susumu Kusunoki
  • Organizer: World Congress of Neurology 2019
  • Int'l Joint Research
• [Presentation] Clinical features and progression of dysphagia in bulbar-onset or limb-onset amyotrophic lateral sclerosis between patients with genetic mutations and those without mutations (2019)
  • Author(s): Chiharu Isono, Makito Hirano, Kanji Fukuda, Makoto Samukawa, Kazumasa Saigoh, Yusaku Nakamura, Susumu Kusunoki
  • Organizer: World Congress of Neurology 2019
  • Int'l Joint Research
• [Presentation] Non-coding repeats in the ATXN8OS gene are expanded in Japanese patients with amyotrophic lateral sclerosis (2019)
  • Author(s): Makito Hirano, Makoto Samukawa, Chiharu Isono, Kazumasa Saigoh, Yusaku Nakamura, Susumu Kusunoki
  • Organizer: 2019 Annual Meeting - American Academy of Neurology
  • Int'l Joint Research