Efficient drug screening and gene correction for treating liver disease using patient-specific stem cells

Authors

  • Su Mi Choi,

    1. Department of Oncology, The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD
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  • Yonghak Kim,

    1. Department of Oncology, The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD
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  • Joong Sup Shim,

    1. Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, MD
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  • Joon Tae Park,

    1. Department of Surgery and the McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University, Baltimore, MD
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  • Rui-Hong Wang,

    1. Genetics of Development and Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD
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  • Steven D. Leach,

    1. Department of Surgery and the McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University, Baltimore, MD
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  • Jun O. Liu,

    1. Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, MD
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  • Chuxia Deng,

    1. Genetics of Development and Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD
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  • Zhaohui Ye,

    Corresponding author
    1. Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD
    • Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD, USA
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  • Yoon-Young Jang

    Corresponding author
    1. Department of Oncology, The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD
    2. Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD
    • Stem Cell Biology Program, Department of Oncology, Johns Hopkins University School of Medicine, 1550 Orleans Street, CRB2 Room 552, Baltimore, MD 21231
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    • fax: 410-502-5742


  • Potential conflict of interest: Nothing to report.

  • This work was supported by the National Institutes of Health (NIH; grant no.: AA020020) and the Maryland Stem Cell Research Fund (MSCRF; grant nos.: 2010-MSCRFII-0101-00, 2011-MSCRFE-0087, and 2012-MSCRFF-0152-00). This work was also supported by the NIH Center for Regenerative Medicine. S.D.L. is supported by the NIH (grant no.: DK61215) and the Paul K. Neumann Professorship at Johns Hopkins University. The maintenance of Johns Hopkins Drug Library was supported, in part, by the National Cancer Institute (R01CA122814), the Flight Attendant Medical Research Institute, and by grant number UL1 RR 025005 from the National Center for Research Resources, a component of the NIH and the NIH Roadmap for Medical Research (to Johns Hopkins School of Medicine).

Abstract

Patient-specific induced pluripotent stem cells (iPSCs) represent a potential source for developing novel drug and cell therapies. Although increasing numbers of disease-specific iPSCs have been generated, there has been limited progress in iPSC-based drug screening/discovery for liver diseases, and the low gene-targeting efficiency in human iPSCs warrants further improvement. Using iPSC lines from patients with alpha-1 antitrypsin (AAT) deficiency, for which there is currently no drug or gene therapy available, we established a platform to discover new drug candidates and correct disease-causing mutation with a high efficiency. A high-throughput format screening assay, based on our hepatic differentiation protocol, was implemented to facilitate automated quantification of cellular AAT accumulation using a 96-well immunofluorescence reader. To expedite the eventual application of lead compounds to patients, we conducted drug screening utilizing our established library of clinical compounds (the Johns Hopkins Drug Library) with extensive safety profiles. Through a blind large-scale drug screening, five clinical drugs were identified to reduce AAT accumulation in diverse patient iPSC-derived hepatocyte-like cells. In addition, using the recently developed transcription activator-like effector nuclease technology, we achieved high gene-targeting efficiency in AAT-deficiency patient iPSCs with 25%-33% of the clones demonstrating simultaneous targeting at both diseased alleles. The hepatocyte-like cells derived from the gene-corrected iPSCs were functional without the mutant AAT accumulation. This highly efficient and cost-effective targeting technology will broadly benefit both basic and translational applications. Conclusions: Our results demonstrated the feasibility of effective large-scale drug screening using an iPSC-based disease model and highly robust gene targeting in human iPSCs, both of which are critical for translating the iPSC technology into novel therapies for untreatable diseases. (HEPATOLOGY 2013;57:2458–2468)

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