Genetic Correction of Huntington's Disease Phenotypes in Induced Pluripotent Stem Cells

The Buck Institute for Research on Aging, Novato, CA 94945, USA.
Cell stem cell (Impact Factor: 22.27). 06/2012; 11(2):253-63. DOI: 10.1016/j.stem.2012.04.026
Source: PubMed


Huntington's disease (HD) is caused by a CAG expansion in the huntingtin gene. Expansion of the polyglutamine tract in the huntingtin protein results in massive cell death in the striatum of HD patients. We report that human induced pluripotent stem cells (iPSCs) derived from HD patient fibroblasts can be corrected by the replacement of the expanded CAG repeat with a normal repeat using homologous recombination, and that the correction persists in iPSC differentiation into DARPP-32-positive neurons in vitro and in vivo. Further, correction of the HD-iPSCs normalized pathogenic HD signaling pathways (cadherin, TGF-β, BDNF, and caspase activation) and reversed disease phenotypes such as susceptibility to cell death and altered mitochondrial bioenergetics in neural stem cells. The ability to make patient-specific, genetically corrected iPSCs from HD patients will provide relevant disease models in identical genetic backgrounds and is a critical step for the eventual use of these cells in cell replacement therapy.

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    • "Symptoms of early and late HD have been demonstrated in the YAC128 HD mouse model, including early susceptibility and later resistance to excitotoxicity induced by quinolinic acid as well as early hyperactivity and later decreases in activity (Slow et al., 2003). Mutant huntingtin is known to affect the activation of the MAPK signaling pathway (Bodai and Marsh, 2012), and the dysregulation pattern is biphasic, depending on the stage of HD. Young YAC128 mice show decreased phosphorylation of ERK, whereas 1-year-old YAC128 mice show an increased level of p-ERK (Gladding et al., 2014). "
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    ABSTRACT: Huntington disease (HD) is an incurable brain disorder characterized by the late onset of motor and cognitive symptoms, even though the neurons in the brain begin to suffer dysfunction and degeneration long before symptoms appear. Several molecular and developmental effects of HD have been identified using neural stem cells (NSCs) and differentiated cells, such as neurons and astrocytes. Still, little is known regarding the molecular pathogenesis of HD in pluripotent cells, such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). Therefore, we examined putative signaling pathways and processes involved in HD pathogenesis in pluripotent cells. We tested naïve mouse HD YAC128 iPSCs and two types of human HD iPSCs that were generated from HD and juvenile HD patients. Surprisingly, we found that a number of changes affecting cellular processes in HD were also present in undifferentiated pluripotent HD iPSCs, including the deregulation of the MAPK and Wnt signaling pathways and the deregulation of the expression of genes related to oxidative stress, such as Sod1. Interestingly, a common protein interactor of the huntingtin protein and the proteins in the above pathways is p53, and the expression of the p53 gene was deregulated in HD YAC128 iPSCs and human HD iPSCs. In summary, our findings demonstrate that multiple molecular pathways that are characteristically deregulated in HD are already altered in undifferentiated pluripotent cells and that the pathogenesis of HD may begin during the early stages of life. © 2015. Published by The Company of Biologists Ltd.
    Disease Models and Mechanisms 06/2015; 8(9). DOI:10.1242/dmm.019406 · 4.97 Impact Factor
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    • "It also allows designing of new treatment strategies for HD. Moreover, if to be used in cell replacement therapy, it is essential to correct mutation of the HTT in HD iPSCs as reported recently using the replacement of the expanded CAG repeat with a normal repeat via homologous recombination [22] "
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    ABSTRACT: Huntington's disease is the most common inherited neurodegenerative disorder among polyglutamine diseases caused by CAG repeat expansion in exon 1 of the huntingtin gene whose translation results in polyQ stretch in the N-terminus of the huntingtin protein. This mutation significantly affects huntingtin conformation, proteolysis, post-translational modifications, as well as its ability to bind interacting proteins. As a consequence, a variety of cellular mechanisms such as transcription, mitochondrial energy metabolism, axonal transport, neuronal vulnerability to oxidative stress, neurotransmission, and immune response are altered and involved in the pathogenesis of Huntington's disease. Promising candidate molecular biomarkers of HD have emerged from proteomic studies. Recent analyses focused on huntingtin protein itself, its post-translational modification and interacting proteins, which are of great importance for disease course. Furthermore, brain, body fluids and immune system are intensively studied in order to search for additional proteins with a view to their use as a biomarker(s) or set of biomarkers in clinical trials in HD translational research. This article is protected by copyright. All rights reserved.
    Proteomics. Clinical applications 02/2015; DOI:10.1002/prca.201400073 · 2.96 Impact Factor
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    • "Moreover, the cytosine adenine guanine (CAG) repeat iPSC lines were established from HD followed by successful targeted gene correction of HD by homologous recombination. The genetically corrected iPSCs gave rise to specific neural subtypes, which reversed disease phenotype of the neural stem cells and could potentially be used for cell replacement therapy in the future [37]. "
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    ABSTRACT: Incurable neurological disorders such as Parkinson’s disease (PD), Huntington’s disease (HD), and Alzheimer’s disease (AD) are very common and can be life-threatening because of their progressive disease symptoms with limited treatment options. To provide an alternative renewable cell source for cell-based transplantation and as study models for neurological diseases, we generated induced pluripotent stem cells (iPSCs) from human dermal fibroblasts (HDFs) and then differentiated them into neural progenitor cells (NPCs) and mature neurons by dual SMAD signaling inhibitors. Reprogramming efficiency was improved by supplementing the histone deacethylase inhibitor, valproic acid (VPA), and inhibitor of p160-Rho associated coiled-coil kinase (ROCK), Y-27632, after retroviral transduction. We obtained a number of iPS colonies that shared similar characteristics with human embryonic stem cells in terms of their morphology, cell surface antigens, pluripotency-associated gene and protein expressions as well as their in vitro and in vivo differentiation potentials. After treatment with Noggin and SB431542, inhibitors of the SMAD signaling pathway, HDF-iPSCs demonstrated rapid and efficient differentiation into neural lineages. Six days after neural induction, neuroepithelial cells (NEPCs) were observed in the adherent monolayer culture, which had the ability to differentiate further into NPCs and neurons, as characterized by their morphology and the expression of neuron-specific transcripts and proteins. We propose that our study may be applied to generate neurological disease patient-specific iPSCs allowing better understanding of disease pathogenesis and drug sensitivity assays.
    PLoS ONE 09/2014; 9(9):e106952. DOI:10.1371/journal.pone.0106952 · 3.23 Impact Factor
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