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Modeling sporadic ALS in iPSC-derived motor neurons identifies a potential therapeutic agent

Nature Medicine volume 24, pages 1579–1589 (2018)Cite this article

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Abstract

Amyotrophic lateral sclerosis (ALS) is a heterogeneous motor neuron disease for which no effective treatment is available, despite decades of research into SOD1-mutant familial ALS (FALS). The majority of ALS patients have no familial history, making the modeling of sporadic ALS (SALS) essential to the development of ALS therapeutics. However, as mutations underlying ALS pathogenesis have not yet been identified, it remains difficult to establish useful models of SALS. Using induced pluripotent stem cell (iPSC) technology to generate stem and differentiated cells retaining the patients’ full genetic information, we have established a large number of in vitro cellular models of SALS. These models showed phenotypic differences in their pattern of neuronal degeneration, types of abnormal protein aggregates, cell death mechanisms, and onset and progression of these phenotypes in vitro among cases. We therefore developed a system for case clustering capable of subdividing these heterogeneous SALS models by their in vitro characteristics. We further evaluated multiple-phenotype rescue of these subclassified SALS models using agents selected from non-SOD1 FALS models, and identified ropinirole as a potential therapeutic candidate. Integration of the datasets acquired in this study permitted the visualization of molecular pathologies shared across a wide range of SALS models.

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Fig. 1: Time-course analysis of in vitro ALS pathology with aging using iPSC-derived motor neurons.
Fig. 2: Pathological analysis of FALS models focusing on the transitional stage of in vitro pathology.
Fig. 3: Multi-phenotypic screening for ALS therapeutics.
Fig. 4: Pathological analysis and phenotype-based clustering of multiple SALS models.
Fig. 5: Evaluation of drug efficacy using multiple SALS models.
Fig. 6: Transcriptional characterization of SALS models and identification of ROPI target pathways using in vitro multi-phenotypic case classification.

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Acknowledgements

The authors would like to thank W. Akamatsu (Juntendo University), D. Sipp, S. Morimoto, and Y. Nishimoto (Keio University) for providing invaluable comments on the project and all the members of the H.O. laboratory for their encouragement and kind support. We would also like to thank H. Inoue, S. Yamanaka, and M. Nakagawa (Kyoto University) for donating hiPSC clones (A21412, A21428, A3411, A3416, and 201B7) and M. Ishikawa (Keio University) for constructing and providing the existing drug library. This work was supported by funding from the Research Project for Practical Applications of Regenerative Medicine from the Japan Agency for Medical Research and Development (AMED) (grant nos. 15bk0104027h0003, 16bk0104016h0004, and 17bk0104016h0005 to H.O.), the Research Center Network for Realization Research Centers/Projects of Regenerative Medicine (the Program for Intractable Disease Research Utilizing Disease-specific iPS Cells and the Acceleration Program for Intractable Diseases Research Utilizing Disease-specific iPS Cells) from AMED (grant nos. 15bm0609003h0004, 16bm0609003h0005, 17bm0609003h0006 and 17bm0804003h0001 to H.O.), the research grants for the sporadic ALS patients registry study (JaCALS) from AMED (grant no. 17lk0201057h0002 to G.S.), a research grant on “Development of therapy for sporadic ALS from omics analyses using a large-scale clinical database, genomic DNA and immortalized cell repository of Japanese ALS patients from AMED” (grant no. 18ek0109284s0302 to G.S. and H.O.), an IBC grant from the Japan ALS Association (to H.O.), the Translational Research Network Program from AMED (to H.O.), the Practical Research Project for Rare/Intractable Diseases from AMED (grant nos. 18ek0109284s0302 to G.S. and H.O. and 18ek0109329h0001 to H.O.), the grant-in-aid project Scientific Research on Innovation Area (Brain Protein Aging and Dementia Control) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (to G.S.and H.O.), Research Fellowships of the Japan Society for the Promotion of Science for Young Scientists (grant no. JP16J06437 to K.F.), the Keio University Grant-in-Aid for the Encouragement of Young Medical Scientists (to K.F.), and the Keio University Doctorate Student Grant-in-Aid Program (to K.F.).

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Authors and Affiliations

  1. Department of Physiology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan

    Koki Fujimori, Mitsuru Ishikawa & Hideyuki Okano

  2. The Institute of Medical Sciences, Tokai University, Isehara, Kanagawa, Japan

    Asako Otomo & Shinji Hadano

  3. Department of Molecular Life Sciences, Tokai University School of Medicine, Isehara, Kanagawa, Japan

    Asako Otomo & Shinji Hadano

  4. Micro/Nano Technology Center, Tokai University, Hiratsuka, Kanagawa, Japan

    Asako Otomo

  5. Department of Neurology, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan

    Naoki Atsuta, Ryoichi Nakamura & Gen Sobue

  6. Department of Neurology, Tohoku University Graduate School of Medicine, Sendai, Miyagi, Japan

    Tetsuya Akiyama & Masashi Aoki

  7. Research Center for Brain and Nervous Diseases, Tokai University Graduate School of Medicine, Isehara, Kanagawa, Japan

    Shinji Hadano

  8. Division of Gene Regulation, Institute for Advanced Medical Research, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan

    Hideyuki Saya

  9. Research Division of Dementia and Neurodegenerative Disease, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan

    Gen Sobue

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  1. Koki Fujimori
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Contributions

K.F. and H.O. designed the study. K.F., A.O., and S.H. established motor neuron differentiation protocols. K.F. generated iPSCs from 32 SALS patients, analyzed the in vitro pathology of FALS and SALS models, designed the phenotype-based clustering system, performed drug screening, conducted in vitro pharmacology, performed transcriptome analysis, and analyzed the data. K.F. and H.S. designed the drug screening system. M.I. established iPSCs from ALS patients carrying SOD1 mutations. N.A., R.N., T.A., M.A., and G.S. provided ALS patient samples and analyzed the data from clinical observations. H.S. provided the existing drug library for screening. Project management was conducted by S.H., M.A., H.S., G.S., and H.O. The manuscript was prepared by K.F. and H.O. All authors contributed to the final editing and approval of the manuscript.

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Correspondence to Hideyuki Okano.

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H.O. is a paid Scientific Advisory Board Member at SanBio Co., Ltd. and K Pharma Inc.

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Supplementary Figures 1–16 and Supplementary Tables 1–4

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Supplementary Dataset 1

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Fujimori, K., Ishikawa, M., Otomo, A. et al. Modeling sporadic ALS in iPSC-derived motor neurons identifies a potential therapeutic agent. Nat Med 24, 1579–1589 (2018). https://doi.org/10.1038/s41591-018-0140-5

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