Development of a novel molecular targeting therapy and probe, using antibodies against pathogenic epitopes in TDP-43

• Project/Area Number: 26290023
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Partial Multi-year Fund
• Section: 一般
• Research Field: Neurochemistry/Neuropharmacology
• Research Institution: Shiga University of Medical Science (2016-2017); Kyoto University (2014-2015)
• Principal Investigator: Makoto Urushitani 滋賀医科大学, 医学部, 教授 (60332326)
• Co-Investigator(Kenkyū-buntansha): 守村 敏史 滋賀医科大学, 神経難病研究センター, 助教 (20333338)
• Project Period (FY): 2014-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥16,510,000 (Direct Cost: ¥12,700,000、Indirect Cost: ¥3,810,000); Fiscal Year 2017: ¥3,770,000 (Direct Cost: ¥2,900,000、Indirect Cost: ¥870,000); Fiscal Year 2016: ¥4,030,000 (Direct Cost: ¥3,100,000、Indirect Cost: ¥930,000); Fiscal Year 2015: ¥4,030,000 (Direct Cost: ¥3,100,000、Indirect Cost: ¥930,000); Fiscal Year 2014: ¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000)
• Keywords: 筋萎縮性側索硬化症（ALS) / ミスフォールドプロテイン / 神経変性疾患 / 抗体医療 / 蛋白質分解 / 脳神経疾患 / 蛋白質 / シグナル伝達 / 筋萎縮性側索硬化症 / TDP-43 / 抗体 / エピトープ / 先制医療 / プローブ / モデルマウス / 治療

Outline of Final Research Achievements

Using our original antibodies against TDP-43, a pathogenic protein for amyotrophic lateral sclerosis, we tested their utility as a tool for high throughput drug screening, and as a specific scavenger of pathogenic TDP-43 aggregates in cells.
For high throughput screening, a polyclonal antibody against dimer interface residues in RRM1 domain, was used in the competitive ELISA, in which molecules which prevents antibody binding with TDP-43 were screening as a hit molecules.Screening worked and several hits were picked up, which interacted dimer interface and efficiently prevents aggregate formation in the transfected culture cells.
To eliminate intracellular TDP-43 aggregates, several types of single chain fragment of variant (scFV) containing proteolysis signals, were generated. These scFv were transfected with normal or aggregate-prone TDP-43. It was shown that our scFv was a promising tool to eliminate misfolded and pathogenic TDP-43 in ALS.

Research Products

• [Journal Article] Amyotrophic Lateral Sclerosis after Receiving the Human Papilloma Virus Vaccine: A Case Report of a 15-year-old Girl (2018)
  • Author(s): Hikiami Ryota、Yamakado Hodaka、Tatsumi Shinsui、Ayaki Takashi、Hashi Yuichiro、Yamashita Hirofumi、Sawamoto Nobukatsu、Tsuji Teruyuki、Urushitani Makoto、Takahashi Ryosuke
  • Journal Title: Internal Medicine Volume: 57 Issue: 13 Pages: 1917-1919
  • DOI: 10.2169/internalmedicine.0285-17
  • NAID: 130007387154
  • ISSN: 0918-2918, 1349-7235
  • Year and Date: 2018-07-01
  • Peer Reviewed / Open Access
• [Journal Article] Elimination of TDP-43 inclusions linked to amyotrophic lateral sclerosis by a misfolding-specific intrabody with dual proteolytic signals. (2018)
  • Author(s): Tamaki Y, Shodai A, Morimura T, Hikiami R, Minamiyama S, Ayaki T, Tooyama I, Furukawa Y, Takahashi R, Urushitani M.
  • Journal Title: Scientific Reports Volume: 印刷中 Issue: 1 Pages: 6030-6030
  • DOI: 10.1038/s41598-018-24463-3
  • NAID: 120006482849
  • Peer Reviewed / Open Access
• [Journal Article] Pathological Endogenous α-Synuclein Accumulation in Oligodendrocyte Precursor Cells Potentially Induces Inclusions in Multiple System Atrophy (2018)
  • Author(s): Kaji Seiji、Maki Takakuni、Kinoshita Hisanori、Uemura Norihito、Ayaki Takashi、Kawamoto Yasuhiro、Furuta Takahiro、Urushitani Makoto、Hasegawa Masato、Kinoshita Yusuke、Ono Yuichi、Mao Xiaobo、Quach Tran H.、Iwai Kazuhiro、Dawson Valina L.、Dawson Ted M.、Takahashi Ryosuke
  • Journal Title: Stem Cell Rep Volume: 10 Issue: 2 Pages: 356-365
  • DOI: 10.1016/j.stemcr.2017.12.001
  • NAID: 120006414715
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Multiple proteinopathies in familial ALS cases with optineurin mutation. (2017)
  • Author(s): Ayaki T, Ito H, Komure O, Kamada M, Nakamura M, Wate R, Kusaka H, Yamaguchi Y, Li F, Kawakami H, Urushitani M, Takahashi R
  • Journal Title: J Neuropathol Exp Neurol Volume: 77 Issue: 2 Pages: 128-138
  • DOI: 10.1093/jnen/nlx109
  • Peer Reviewed
• [Journal Article] Charcot-Marie-Tooth disease type 2A with an autosomal recessive inheritance; the first report of an adult-onset disease. (2017)
  • Author(s): Hikiami R, Yamashita H, Koita N, Jingami N, Sawamoto N, Furukawa K, Kawai H, Terashima T, Oka N, Hashiguchi A, Takashima H, Urushitani M, and Takahashi
  • Journal Title: J Hum Genet Volume: 63 Issue: 1 Pages: 89-92
  • DOI: 10.1038/s10038-017-0353-3
  • Peer Reviewed
• [Journal Article] Velocity of intraneural blood flow is increased in inflammatory neuropathies: sonographic observation. (2016)
  • Author(s): Carandang A, Takamatsu N, Nodera H, Mori A, Mimura N, Okada N, Kinoshita H, Kuzuya A, Urushitani M, Takahashi R, Izumi Y, Kaji R.
  • Journal Title: J Neurol Neurosurg Psychiatr Volume: - Issue: 5 Pages: 455-457
  • DOI: 10.1136/jnnp-2016-314547
  • Peer Reviewed
• [Journal Article] Localization of a mutant SOD1 protein in E40K-heterozygous dogs: implications for non-cell-autonomous pathogenesis of degenerative myelopathy. (2016)
  • Author(s): Kobatake Y, Sakai H, Tsukui T, Yamato O, Kohyama M, Sasaki J, Kato S, Urushitani M, Maeda S, Kamishina H.
  • Journal Title: J Neurol Sci Volume: 372 Pages: 369-378
  • DOI: 10.1016/j.jns.2016.10.034
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Activated caspase-9 immunoreactivity in glial and neuronal cytoplasmic inclusions in multiple system atrophy. (2016)
  • Author(s): Kawamoto Y, Ayaki T, Urushitani M, Ito H, Takahashi R.
  • Journal Title: Neurosci Lett. Volume: 628 Pages: 207-212
  • DOI: 10.1016/j.neulet.2016.06.036
  • Peer Reviewed / Open Access
• [Journal Article] CUL2-mediated clearance of misfolded TDP-43 is paradoxically affected by VHL in oligodendrocytes in ALS. (2016)
  • Author(s): Uchida T, Tamaki Y, Ayaki T, Shodai A, Kaji S, Morimura T, Banno Y, Nishitsuji K, Sakashita N, Maki T, Yamashita H, Ito H, Takahashi R, Urushitani M.
  • Journal Title: Scientific Reports Volume: 6 Issue: 1 Pages: 19118-19136
  • DOI: 10.1038/srep19118
  • NAID: 120005689210
  • Peer Reviewed / Open Access
• [Journal Article] Gradual cerebral hypoperfusion in spontaneously hypertensive rats induces slowly evolving white matter abnormalities and impairs working memory (2016)
  • Author(s): Kitamura A, Saito S, Maki A, Ayaki T, Oishi N, Kalaria RN, Hattori Y, Yamamoto Y, Urushitani M, Horsburgh K, Fukuyama H, Takahashi R, Ihara M
  • Journal Title: J Cereb Blood Flow Metab Volume: - Issue: 9 Pages: 1592-1602
  • DOI: 10.1177/0271678x15606717
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Accumulation and aggregate formation of mutant superoxide dismutase 1 in canine degenerative myelopathy. (2015)
  • Author(s): Nakamae S, Kobatake Y, Suzuki R, Tsukui T, Kato S, Yamato O, Sakai H, Urushitani M, Maeda S, Kamishina H.
  • Journal Title: Neuroscience Volume: 303 Pages: 229-240
  • DOI: 10.1016/j.neuroscience.2015.06.066
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] ALSとhnRNPA2B1 (2015)
  • Author(s): 玉木良高、漆谷 真
  • Journal Title: 神経内科 Volume: 82 Pages: 394-397
• [Journal Article] A cysteine residue affects the conformational state and neuronal toxicity of mutant SOD1 in mice: relevance to the pathogenesis of ALS (2015)
  • Author(s): Nagano S, Takahashi Y, Yamamoto K, Masutani H, Fujiwara N, Urushitani M, Araki T
  • Journal Title: Hum Mol Genet. Volume: 0n line Issue: 12 Pages: 1-13
  • DOI: 10.1093/hmg/ddv093
  • Peer Reviewed
• [Journal Article] 遺伝子改変によるALSモデルマウス (2015)
  • Author(s): 山下博史, 漆谷 真
  • Journal Title: 医学のあゆみ Volume: 252 Pages: 309-309
• [Journal Article] Immunoreactivity of valosin-containing protein in sporadic amyotrophic lateral sclerosis and in a case of its novel mutant (2014)
  • Author(s): Ayaki T, Ito H, Fukushima H, Inoue T, Kondo T, Ikemoto A, Asano T, Shodai A, Fujita T, Fukui S, Morino H, Nakano S, Kusaka H, Yamashita H, Ihara M, Matsumoto R, Kawamata J, Urushitani M, Kawakami H, Takahashi R.
  • Journal Title: Acta Neuropathol Commun. Volume: 2 Issue: 1 Pages: 172-185
  • DOI: 10.1186/s40478-014-0172-0
  • Peer Reviewed / Open Access
• [Journal Article] Proteasome dysfunction induces muscle growth defects and protein aggregation (2014)
  • Author(s): Kitajima Y, Tashiro Y, Suzuki N, Warita H, Kato M, Tateyama M, Ando R, Izumi R, Yamazaki M, Abe M, Sakimura K, Ito H, Urushitani M, Nagatomi R, Takahashi R, and Aoki M
  • Journal Title: J Cell Sci Volume: 127 Pages: 5204-5217
  • DOI: 10.1242/jcs.150961
  • Peer Reviewed / Open Access
• [Presentation] 分解型細胞内抗体を用いたALSの分子標的治療 (2017)
  • Author(s): 漆谷 真.
  • Organizer: 第35回日本神経治療学会総会
  • Invited
• [Presentation] 孤発性筋萎縮性側索硬化症における細胞内抗体を用いた異常TDP-43除去の試み (2017)
  • Author(s): 漆谷 真.
  • Organizer: 第29回日本神経免疫学会学術集会
  • Invited
• [Presentation] 病原蛋白質特異的エピトープに対するALSの分子標的治療開発戦略. (2017)
  • Author(s): 漆谷 真.
  • Organizer: 日本薬学会, 次世代を担う創薬・医療薬理シンポジウム2017
  • Invited
• [Presentation] Cilostazol reduced gliovascular damage and working memory impairment via endotelial protection in a mouse model of vascular dementia. (2017)
  • Author(s): itamura A, Sanz Y, Duncombe J, Koudelka J, Binnie M, Lennen R, Webster S, Jansen M, Marshall I, Ogawa N, Urushitani M, Kalaria R, Ihara M, Horsburgh K.
  • Organizer: World Congress of Neurology. 2017
  • Int'l Joint Research
• [Presentation] Myositis and muscular inclusions in Nakajo-Nishimura syndrome. (2017)
  • Author(s): Ayaki T, Murata K, Kanazawa N, Uruha A, Nishino I, Omura K, Sugie K, Mori M, Ueno S, Furukawa F, Ito H, Urushitani M, Takahashi R
  • Organizer: World Congress of Neurology. 2017
  • Int'l Joint Research
• [Presentation] Degradative intrabody for selective elimination of pathogenic TDP-43 aggregates in vitro and in murine embryos’ cerebrum (2017)
  • Author(s): Tamaki Y, Shodai A, Hikiami R, Minamiyama S, Morimura T, Ayaki T, Takahashi R, Urushitani M.
  • Organizer: World Congress of Neurology. 2017
  • Int'l Joint Research
• [Presentation] Pathological analysis of a novel conformational specific anti-SOD1 antibody using human and mice tissues in amyotrophic lateral sclerosis (2017)
  • Author(s): Yamaguchi Y, Ayaki T, Urushitani M, Takahashi R.
  • Organizer: World Congress of Neurology. 2017,
  • Int'l Joint Research
• [Presentation] Development of an effective technology for inductive differentiation from bone marrow-derived mononuclear cells to neuroprotective microglia. (2017)
  • Author(s): Kobashi S, Terashima T, Nakae Y, Katagi M, Morimura T, Kojima H, Urushitani M
  • Organizer: World Congress of Neurology. 2017,
  • Int'l Joint Research
• [Presentation] The development of therapeutic intrabody against misfolded TDP-43 for ALS (2016)
  • Author(s): Tamaki Y, Shodai A, Hikiami R, Minamiyama S, Ayaki T, Takahashi R, Urushitani M
  • Organizer: Annual Meeting of Society for Neuroscience. 2016
  • Place of Presentation: San Diego （U.S.A)
  • Year and Date: 2016-11-12
• [Presentation] 上位運動ニューロン徴候を主徴とし、急速に進行したTDP-43変異筋萎縮性側索硬化症の剖検例 (2016)
  • Author(s): 北川尚之、内野彰子、山下博史、漆谷 真、櫻井道雄、村山繁雄
  • Organizer: 第57回日本神経病理学会総会学術研究会.
  • Place of Presentation: 弘前
  • Year and Date: 2016-06-01
• [Presentation] Impaired VHL/CUL2-mediated clearance of misfolded TDP-43 underlies glial cytoplasmic inclusions (GCIs) in ALS (2016)
  • Author(s): 漆谷 真
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸市
  • Year and Date: 2016-05-20
  • Invited
• [Presentation] VHL/CUL2によるミスフォールド型TDP-43の分解障害とオリゴデンドロサイト封入体形成 (2016)
  • Author(s): 漆谷 真
  • Organizer: 第57回日本神経学会学術大会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-18
  • Invited
• [Presentation] 再発性多発軟骨炎に伴う脳炎2症例の検討 (2016)
  • Author(s): 岡田信久、和田隆史、樽野陽亮、眞木崇州、吉藤元、丹治正大、三橋賢大、山門穂高、上田紗希帆、前川嵩太、引網亮太、木下久徳、綾木孝、山下博史、澤本伸克、漆谷真、池田昭夫、高橋良輔.
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-18
• [Presentation] Activation of caspase-9 in the neuronal and glial elements in dementia with Lewy bodies (2016)
  • Author(s): 河本恭裕、綾木孝、漆谷 真、伊東秀文、高橋良輔
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-18
• [Presentation] The mechanism of Motor Neuron Degeneration in Ubiquitin Proteasome System Dysfunction (2016)
  • Author(s): 星野友則、山下博史、田代善祟、三澤日出己、漆谷 真、高橋良輔
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-18
• [Presentation] 自己免疫性てんかんにおける末梢血リンパ球の表面抗原解析 (2016)
  • Author(s): 端祐一郎、坂本光弘、十川純平、岡田洋一郎、藤井ちひろ、越智博文、中川正法、水野俊樹、下竹昭寛、松本理器、漆谷 真、池田昭夫、高橋良輔
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-18
• [Presentation] Measurement of TDP-43 in peripheral blood cells of ALS. (2016)
  • Author(s): 菱澤美貴、山下博史、漆谷真、高橋良輔
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-18
• [Presentation] The development of therapeutic intrabody against misfolded/unfolded TDP-43 for ALS (2016)
  • Author(s): 玉木良高、小代明美、綾木孝、高橋良輔、漆谷 真.
  • Organizer: 第57回日本神経学会 学術集会
  • Place of Presentation: 神戸
  • Year and Date: 2016-05-13
• [Presentation] TDP-43の病原構造の同定と抗体を用いた分子標的治療 (2015)
  • Author(s): 漆谷 真
  • Organizer: 第56回 日本神経学会学術集会
  • Place of Presentation: 新潟市
  • Year and Date: 2015-05-22
  • Invited
• [Presentation] FUS遺伝子変異を伴う筋萎縮性側索硬化症の臨床像：若年発症例を含む4症例の検討 (2015)
  • Author(s): 引網亮太,, 端祐一郎, 山門穂高, 辰己 新水, 山下博史, 澤本伸克, 辻輝之, 川本未知, 幸原伸夫, 井上治久, 漆谷真, 高橋良輔
  • Organizer: 第56回日本神経学会 学術集会
  • Place of Presentation: 新潟市
  • Year and Date: 2015-05-22
• [Presentation] VHL とCUL2; TDP-43 の凝集を促進するVHL と分解を促進するCUL2. TDP-43 proteinopathy におけるVHL の2 面性 (2015)
  • Author(s): 内田 司, 玉木良高, 小代明美, 綾木 孝, 守村敏史, 辰巳新水, 伊東秀文, 坂下 直実, 高橋良輔, 漆谷真
  • Organizer: 第56回日本神経学会 学術集会
  • Place of Presentation: 新潟市
  • Year and Date: 2015-05-22
• [Presentation] Clinicopathologic study of autopsied familial ALS cases with optineurin mutation (2015)
  • Author(s): 綾木 孝, 伊東秀文、小牟禮修、辰己新水、鎌田正紀、漆谷 真、川上秀史、髙橋良輔
  • Organizer: 第56回日本神経学会 学術集会
  • Place of Presentation: 新潟市
  • Year and Date: 2015-05-21
• [Presentation] イヌの筋萎縮性側索硬化症モデルの臨床像、病理像および変異型SOD1蛋白の生化学的特性 (2014)
  • Author(s): 小畠 結，小宅 香苗，津久井 利広，酒井 洋樹，大和 修，齋藤 弥代子，漆谷 真，加藤 信介，佐々木 淳，柴田 早苗，前田 貞俊，神志那 弘明
  • Organizer: 第３7回日本神経科学大会
  • Place of Presentation: パシフィコ横浜
  • Year and Date: 2014-09-13
• [Presentation] The role of conformation of RNA recognition motifs of TDP-43 in the protein quality control system in ALS (2014)
  • Author(s): Makoto Urushitani
  • Organizer: 第３7回日本神経科学大会 第6回日本カナダ合同シンポジウム
  • Place of Presentation: パシフィコ横浜
  • Year and Date: 2014-09-12
  • Invited
• [Presentation] ALSのTDP-43のinclusion形成におけるVon Hippel-Lindau proteinの役割 (2014)
  • Author(s): 内田 司，小代 明美，守村 敏史，辰巳 新水，高橋 良輔，漆谷 真
  • Organizer: 第３7回日本神経科学大会
  • Place of Presentation: パシフィコ横浜
  • Year and Date: 2014-09-11
• [Presentation] TDP-43のCullin-2型E3リガーゼの同定及び機能解析 (2014)
  • Author(s): 内田 司, 小代明美, 大野美樹, 辰巳新水, 守村敏史, 高橋良輔, 漆谷 真
  • Organizer: 第55回日本神経学会学術集会
  • Place of Presentation: 福岡国際会議場
  • Year and Date: 2014-05-23
• [Book] 脳内環境辞典 (2017)
  • Author(s): 漆谷 真.
  • Total Pages: 156
  • Publisher: メディカルドゥ社
  • ISBN: 9784944157648
• [Book] 脳内環境辞典 (2017)
  • Author(s): 高橋良輔、山中宏二、樋口真人、漆谷 真
  • Total Pages: 156
  • Publisher: メディカルドゥ
• [Book] 分子脳科学 (2015)
  • Author(s): 山下博史, 漆谷 真., 髙橋良輔
  • Publisher: 化学同人
• [Book] レドックスUPDATE (2015)
  • Author(s): 玉木良高、漆谷 真
  • Publisher: 医歯薬出版株式会社
• [Remarks] 滋賀医科大学 内科学講座神経内科ホームページ 「研究について」
  • URL: http://shiga-neurology.com/research/
• [Patent(Industrial Property Rights)] 異常TDP-43を分解除去する抗体断片 (2017)
  • Inventor(s): 漆谷真、玉木良高
  • Industrial Property Rights Holder: 漆谷真、玉木良高
  • Industrial Property Rights Type: 特許
  • Industrial Property Number: 2018-049752
  • Filing Date: 2017