Cell Stem Cell
Genetic Correction of Huntington’s Disease
Phenotypes in Induced Pluripotent Stem Cells
Mahru C. An,1,2Ningzhe Zhang,1,2Gary Scott,1Daniel Montoro,1Tobias Wittkop,1Sean Mooney,1Simon Melov,1
and Lisa M. Ellerby1,*
1The Buck Institute for Research on Aging, Novato, CA 94945, USA
2These authors contributed equally to this work
Huntington’s disease (HD) is caused by a CAG ex-
pansion 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 re-
combination, 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 path-
ways (cadherin, TGF-b, BDNF, and caspase activa-
tion) and reversed disease phenotypes such as
susceptibility to cell death and altered mitochondrial
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
Huntington’s disease (HD) is a progressive autosomal dominant
neurodegenerative disorder caused by an expansion of poly-
glutamine (CAG) repeats in the huntingtin (HTT) protein.
HD patients have progressive motor dysfunction, cognitive
decline, and psychological problems. The age of onset for these
symptoms is directly correlated with the number of repeats,
where more than 36 repeats is considered a pathological
threshold. With no effective disease modifying therapies yet
developed for HD, regenerative medicine may offer new thera-
peutic approaches for treating the disease. While preliminary
clinical trials using fetal neural grafts showed modest success
(Bachoud-Le ´vi et al., 2006; Dunnett and Rosser, 2004), recent
studies have looked to build upon progress in the human
stem cell field to explore a potentially more expandable and
adaptable source of cells for transplantation therapy (Dey
et al., 2010; Schwarz and Schwarz, 2010; Snyder et al., 2010;
Vazey et al., 2010).
The recent landmark discovery that somatic cells can be re-
programmed into induced pluripotent stem cells (iPSCs) (Okita
et al., 2007) sparked a confluence of studies to understand their
growth and manipulation, as well as their potential use in
regenerative medicine (Brennand et al., 2011; Park et al.,
2008; Wu and Hochedlinger, 2011). In devastating neurological
diseases such as HD, Parkinson’s disease (PD), and amyotro-
phic lateral sclerosis (ALS), and in stroke and spinal cord injury,
it is the loss of neurons and the inability to efficiently mobilize
inherent regenerative mechanisms to recover from the acute
or progressive damage that underlies the pathology and prog-
nosis. As such, stem-cell-based therapies hold promise for the
future treatment of these diseases where currently no disease-
modifying therapy exists. Complimentary to stem cell replace-
ment therapies is the study of human stem cells as disease
models to identify novel drugs by providing a platform for a
better understanding of these diseases at the molecular and
cellular level (Ku et al., 2010; Ruby and Zheng, 2009). In
accord with this goal, we recently described an HD-iPSC-
derived neural stem cell (NSC) model to understand disease
pathogenesis and screen for drugs in HD research (Zhang
et al., 2010).
The ability to genetically modify human stem cells is an invalu-
able approach for both modeling diseases and providing poten-
tial therapies. Gene targeting by homologous recombination is
one feasible and surgically targeted modification that holds
promise for providing normal cells that can be used to treat
genetic diseases such as HD. Such methods have been well
developed into highly optimized protocols for the genetic
manipulation of mouse embryonic stem cells (ESCs). Yet due
to a variety of technical challenges posed by human stem cell
manipulation, gene targeting in human ESCs and iPSCs has
not been as well established (Costa et al., 2007; Davis et al.,
2009; Ruby and Zheng, 2009; Song et al., 2010; Urbach et al.,
2004; Yusa et al., 2011; Zou et al., 2009; Zwaka and Thomson,
2003). Thus, gene targeting protocols in human pluripotent
stem cells are still in their infancy, and require further develop-
ment to establish their feasibility as tools for regenerative
As a critical step in advancing gene targeting strategies for the
correction of disease mutations, we established methods to
perform genetic correction in iPSCs derived from an HD patient.
We obtained a line of HD-patient-derived iPSCs (HD-iPSC)
established and characterized by G.Q. Daley’s group (Park
et al., 2008) and us (Zhang et al., 2010). The results of our study
are described below.
Cell Stem Cell 11, 253–263, August 3, 2012 ª2012 Elsevier Inc. 253
Cell Stem Cell
Stem Cells and Huntington’s Disease
254 Cell Stem Cell 11, 253–263, August 3, 2012 ª2012 Elsevier Inc.
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