DNA Repair Mechanisms in Huntington’s Disease

Ida Jonson & Rune Ougland & Elisabeth Larsen

Received: 8 November 2012 /Accepted: 13 January 2013# Springer Science+Business Media New York 2013

Abstract The human genome is under continuous attack by aplethora of harmful agents. Without the development of sev-eral dedicated DNA repair pathways, the genome would havebeen destroyed and cell death, inevitable. However, whileDNA repair enzymes generally maintain the integrity of thewhole genome by properly repairing mutagenic and cytotoxicintermediates, there are cases in which the DNA repair ma-chinery is implicated in causing disease rather than protectingagainst it. One case is the instability of gene-specific trinu-cleotides, the causative mutations of numerous disorders in-cluding Huntington’s disease. The DNA repair proteinsinduce mutations that are different from the genome-widemutations that arise in the absence of repair enzymes; theyoccur at definite loci, they occur in specific tissues duringdevelopment, and they are age-dependent. These latter char-acteristics make pluripotent stem cells a suitable model systemfor triplet repeat expansion disorders. Pluripotent stem cellscan be kept in culture for a prolonged period of time and caneasily be differentiated into any tissue, e.g., cells along theneural lineage. Here, we review the role of DNA repairproteins in the process of triplet repeat instability in Hunting-ton’s disease and also the potential use of pluripotent stemcells to investigate neurodegenerative disorders.

Keywords Huntington’s disease . Dementia . DNA repair .

Induced pluripotent stem cells

Introduction

Huntington’s disease (HD) was first identified by theNorwegian doctor Johan C. Lund and the American doctor

Georg Huntington during the late nineteenth century. Theydescribed a series of symptoms including involuntarymovements and progressive dementia. Later, it has becomeevident that HD is a late-onset incurable neurodegenerativedisorder with a prevalence of three to ten affected subjectsper 100,000 individuals in Western Europe and NorthAmerica [1]. HD is characterized by cognitive and memoryimpairment, dementia, changed personality and behavior,weight loss, and prominent choreic motor abnormalities.Once manifest disease occurs, the duration of HD is ap-proximately 15–20 years, with the disease symptoms be-coming increasingly disabling prior to death [2].

Despite years of research, the molecular pathogenesis andcause of neuronal death in HD remains unknown. A majorbreakthrough in the understanding of HD came with theidentification of the Huntingtin gene (HTT), located onchromosome 4. HD is a purely genetic disease and in HDpatients, the HTT gene contains an expanded and unstabletrinucleotide repeat (TNR) sequence lying within exon 1[3–5] (Fig. 1). The disease severity and onset depends uponthe length of this glutamine-encoding cytosine–adenine–guanine (CAG) repeat sequence [6]. The Huntingtin protein(HTT) plays a role in protein trafficking, vesicle transport,postsynaptic signaling, transcription, and apoptosis [7]. Theglutamine tract leads to protein aggregation and abnormaldegradation of the HTT protein into smaller fragments thatsubsequently undergo ubiquitination. These fragmentsmove from the cytosol to the nucleus and aggregate to causedamage and induced apoptosis [8, 9]. The HTT protein isubiquitously expressed during embryonic development andat high levels in the testis and in mature postmitotic neuronsin adult human brain. Loss of function of the normal HTTprotein and a toxic gain of the function of mutant HTTcontribute to the disruption of multiple intracellular path-ways, ultimately leading to neurodegeneration. HD isinherited in an autosomal dominant trait; thus, disease phe-notype depends upon mutation on either of an individual'stwo copies of the HTT gene, and it is almost 100 %

I. Jonson : R. Ougland : E. Larsen (*)Department of Microbiology, University of Oslo, Oslo UniversityHospital, Rikshospitalet, P. O. Box 4950 Nydalen,0424 Oslo, Norwaye-mail: elisabeth.larsen@rr-research.no

Mol NeurobiolDOI 10.1007/s12035-013-8409-7

penetrant. When the number of repeats reaches a certainthreshold, the repeat sequences become unstable and multi-ply as generations pass, hence leading to a more severedisease with an earlier onset in successive generations—aphenomenon called “genetic anticipation” [10]. The repeatsequence does expand in germ cells during differentiation,and it is less stable during spermatogenesis than oogenesis,meaning that paternally inherited alleles are more prone toexpansions. Normal individuals are within the range of 6–35CAG repeats in the HTT gene, whereas HD-affected indi-viduals have CAG repeat numbers of 36–250. CAG sizesabove 65 leads to the rather rare juvenile form. In addition,expansion is tissue-specific, with the greatest variabilityseen in certain regions of the brain, particularly in thestriatum and cortex, the areas of the brain most vulnerableto neurodegeneration in HD. The exact molecular process ofsomatic expansion has yet to be elucidated. However, thereis evidence for dependence upon DNA repair pathways, assomatic expansion is abrogated in mouse models of HD bythe deletion of mismatch repair genes Msh2 and Msh3 [11,12]. In addition, the modification of age-dependent somaticinstability by the base excision repair (BER) enzyme OGG1links oxidative DNA damage to CAG repeat instability [13].

The brain is our most complex organ, and it uses 20 % oftotal inspired oxygen, making neurons particularly vulnera-ble to oxidative stress. An increasing body of evidencesuggests that accumulation of oxidative DNA damage with-in specific types of neurons is a critical contributor tonormal aging as well as neurodegenerative disease [14].Substantial biological changes related to a condition ofoxidative stress have been found in brain tissue as well asin peripheral tissues in HD individuals, and in HD miceoxidative DNA damage has been shown to preferentiallyaccumulate at CAG repeats in a length-dependent manner.Although the mutation responsible for HD was discoveredalmost two decades ago, the molecular mechanisms under-lying the disease are still poorly understood. To date, thereare no effective treatments to cure the disease or slow itsprogression. Also, investigation of HD development at the

molecular level is encumbered by the lack of an appropriatemodel system. Initiation of the disease occurs long beforethe patient develops any symptoms, and relevant patienttissue (brain and neural tissue in the case of HD) is limitedand difficult to obtain. However, recent discoveries withinthe field of stem cell research, e.g. the induction of pluri-potency, may have solved this problem.

Pluripotent stem cells (PSCs) are characterized by theirability to self-renew indefinitely and to differentiate into anyof the 220 different cell types of the human body [15]. Arecent research has demonstrated the possible conversion ofterminally differentiated cells into cells with an embryonic-like phenotype. These cells are called induced pluripotentstem cells (iPSCs) [16–19]. iPSCs are the product of somat-ic cell reprogramming achieved by the introduction of adefined and limited set of transcription factors and by cul-turing these cells under embryonic stem cell conditions(Fig. 2). The reprogramming procedure is straightforward,robust, and has been independently replicated by multiplegroups. To date, numerous iPSCs with disease phenotypeshave been generated to serve as a tool for human diseasemodeling [20–25].

In this review, we will focus on the relationship betweenHD and the DNA repair pathways that might be involved indisease initiation and progression, with a special emphasison oxidative DNA damage and its repair. In the end, we willdiscuss how iPSCs, and differentiated derivatives of these,may provide a proper model system for detailed molecularstudies of HD pathogenesis. Ultimately, iPSCs may even beused for cell therapy and represent a new treatment optionfor HD patients.

DNA Damage in Mammalian Cells

It is estimated that, every day, each single cell of the humanbody is subjected to an average of approximately one mil-lion DNA lesions, including apurinic/apyrimidinic (AP)sites (abasic sides), DNA adducts, single-strand breaks,

Fig. 1 Graphic showing the excessive repetitions of the cytosine–ade-nine–guanine nucleotide sequence in a gene from a Huntington's diseasepatient (bottom) compared to a gene from a person without the

neurodegenerative disorder (top). The CAG encoding the glutamine tractleads to protein aggregation and abnormal degradation of the HTT proteininto smaller fragments that subsequently undergo ubiquitination

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double-strand breaks, insertion/deletion mismatches, andDNA–proteins cross-links [26]. DNA-damaging agents are ofboth exogenous origin, like sunlight and tobacco smoke, andof endogenous origin, such as water and reactive oxygenspecies [27]. Lesions can affect either the nitrogenous basesor the backbone of the nucleic acids, i.e., sugar and phosphate.Purins are lost more easily than pyrimidines, and the brain isthe most affected organ, followed by the colon and heart. Thecell cannot tolerate damage to the DNA, and various cellularresponses have evolved to deal with such attacks. At thecellular level, DNA lesions may hamper processes such astranscription and replication. To allow time for repair, the cellcycle may be arrested before replication and cell division inresponse to DNA damage [28, 29]. If the damage load is toolarge for the cell to handle, it may initiate apoptosis to preventmultiplication of highly defective cells [30]. Based on theircellular consequences, it is convenient to arrange harmfullesions into two major classes, referred to as either mutagenicor cytotoxic [31]. A mutagenic lesion has altered base pairingproperties and may give rise to mutations during replicationand possibly aberrant proteins. Among the main mutageniclesions are the O-methylations like O6-methylguanine and O4-methylthymine, oxidations like 8-hydroxyguanine, deamina-tion due to hydrolytic attacks and etheno lesions like 1,N6-ethenoadenine. Cytotoxic lesions will block the DNA andRNA polymerases as well as translating ribosomes, thus givingrise to truncated mRNAs and peptides with harmful effects.This interferes with normal cellular processes, e.g., DNA rep-lication and protein synthesis, and is evidently deleterious tothe cell. The most frequent cytotoxic lesions include loss ofbases, thus generating AP sites, DNA inter/intra strand cross-links, protein-DNA/RNA cross-links, and the N-methylationslike 1-methyladenine, 3-methyladenine, and 3-methylcytosine.If not properly repaired, DNA damage can lead to genomicinstability, senescence, apoptosis at a cellular level, and even-tually development of human disease [32]. In fact, numerousstudies provide evidence for increased levels of oxidativeDNA damage in both HD patients and in experimental models

of HD [33, 34]. Collectively, these results support a role foroxidative stress in neuronal degeneration. Furthermore, a cor-relation between oxidative stress and disease stage in HDpatients is established [35], and there is a close link betweenincreased DNA fragmentation in HD patients and the length ofthe CAG triplet repeat, thus underscoring the implication ofDNA repair pathways in the development of HD [36].

DNA Repair Pathways in Mammalian Cells

DNA was once considered so genuine and pure that thepossibility of the genes to be subject to insults and repairevents was practically unthinkable. Later, however, the exis-tence of several DNA repair pathways became evident, andFrancis Crick wrote: “We totally missed the possible role ofenzymes in DNA repair…I later came to realize that DNA isso precious that probably many distinct repair mechanismswould exist. Nowadays one could hardly discuss mutationwithout considering repair at the same time” [37]. In fact, allorganisms have evolved DNA repair mechanisms to counter-act different types of DNA damage. In the human genome,more than 130 genes have been found to be involved in repairmechanisms [38]. As soon as the damage has been located,specific repairing molecules are recruited and bound to or nearthe damaged site, inducing other molecules to bind and makea complex that is able to repair the damage. Mammals possessfive major repair pathways and a battery of repair enzymes tocounteract the daily amount of DNA damage. These arenonhomologous end joining, homologous recombination,mismatch repair (MMR), nucleotide excision repair, andBER. In addition, there are at least two enzymes removinglesions from DNA/RNA by a direct repair mechanism(ALKBH2 and ALKBH3). Usually, however, DNA repairpathways are multi-step reactions involving large proteincomplexes. Some have very narrow substrate specificities,dealing with only one or a few different lesions, whereasothers have broader specificity and take care of all the various

Fig. 2 Reprogramming patient-specific cells for disease modeling. Bytaking a skin biopsy of the patient and reprogramming the cells via theintroduction of certain genes such as OCT4, MYC, KLF4, and SOX2,patient-specific iPSCs can be generated. These genes in turn induce the

expression of the endogenous stem cell master regulators, OCT4 andNANOG. The iPSCs can then be recovered and differentiated thera-peutically into cells and tissue

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lesions making up a particular class of damage. DNA repairdeficiencies have been linked to numerous neurodegenerativedisorders. In the following, MMR and BER will be discussedin further detail with respect to HD development and tripletrepeat expansion.

Base Excision Repair

BER is one of the most active DNA repair processes thatallows the specific recognition and excision of a damagedDNA base (Fig. 3). The majority of the DNA damagesrepaired by BER are caused by hydrolysis, oxygen radicals,and alkylating agents. Mechanisms for BER are highlyconserved and apparently represent an ancient mechanismof defense that counteracts spontaneous decay of DNA [27,39–41]. Compared with other repair pathways, BER appears

to be the simplest and most thorough of all repair processesand consist of five basic steps catalyzed by differentenzymes. The specificity of the repair pathway is deter-mined by the DNA glycosylase that initiates BER. EachDNA glycosylase is specific for a limited number of dam-aged bases [42, 43]. The DNA glycosylase cleaves the N-C1′ glycosylic bond between the base and deoxyribose, thusreleasing the damaged base and leaving an AP site that iscytotoxic and mutagenic and must be further processed.DNA glycosylases are either monofunctional and only re-move the base or bifunctional with an additional lyaseactivity which cleaves DNA 3′ of the AP site. All knownDNA glycosylases directed against bases damaged by oxi-dation are bifunctional. The discovery of a 5′-deoxyribo-phosphatase activity of DNA polymerase β (Pol β) allowedreconstitution of the mammalian BER pathway with justfour purified enzymes [44, 45]. A DNA glycosylase initiates

Fig. 3 A simplified view of thebranched pathways of BER inhumans. In the left pathway, asingle nucleotide is replaced,whereas several nucleotides arereplaced in the right pathway.Regular AP sites, resulting fromspontaneous or glycosylase-initiated removal of the base,can be repaired via the single-nucleotide pathway, although aminor fraction is repaired viathe right pathway. Oxidized orreduced AP sites are removedvia the right pathway. BERinitiated by a DNA glycosylasewith associated AP lyaseactivity, OGG1 or NTH1, isrepaired slightly different fromthe pathways illustrated. NEIL,Nei-like glycosylase; OGG1, 8-oxoguanine glycosylase 1;UDG, uracil-DNA glycosylase;APEX1, AP endonuclease 1;Pol, DNA polymerase; FEN1,flap endonuclease 1; Lig, DNAligase

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the BER pathway by removal of the damaged base, the APendonuclease APEX1 nick the DNA strand at the AP site,Pol β fills the gap and removes the 5′-phosphate sugarmoiety, and the repair is completed by the action of aDNA ligase. This repair pathway is named short patchBER, and it was recently demonstrated that translocationof APE1 to the mitochondria is disrupted in HD cells [46].Shortly thereafter, the FEN1-dependent long patch BER wasreconstituted with purified human enzymes. In vitro, onlythe FEN1 enzyme is required in addition to those describedfor the short patch pathway [47, 48]. The choice of sub-pathway in BER depends on whether the 5′-deoxyribosephosphate (dRP) intermediate can be efficiently removedby the Pol β lyase activity to yield a 5′-phosporylatedDNA strand capable of serving as a substrate for DNAligase [49, 50]. When such processing is inefficient, longpatch BER can occur, and more extensive strand displace-ment that involves incorporation of up to six nucleotides bypolymerase δ, ε, or β takes place. FEN1, together withproliferating cell nuclear antigen (PCNA) and replicationfactor A, removes the overhang formed by repair synthesis,thereby displacing the dRP-containing strand. A DNA li-gase seals the nick [47]. The first direct evidence linkingBER to HD was provided by Kovtun et al. demonstratingthat somatic expansion of the CAG tract was dependent onthe DNA glycosylase OGG1 [13]. Expansion of CAGrepeats in the HTT gene is the underlying cause of HD,where an ongoing mutational process enhances toxicity ofthe resulting mutant HTT protein in specific tissues. Recentstudies have shown that the stoichiometry of BER enzymes iscorrelated to CAG expansion in HD brain and specifically thatsuboptimal LP-BER promotes CAG instability [33, 51].Derevyanko et al. analyzed the kinetics of excision of 8-oxoGby OGG1 from substrates containing a CAG run and foundthat long runs expand easier than short ones, not only becauseof more probable appearance of the damage but also due tomore effective repair within the run [52]. In line with this,Jarem et al. found that 8-oxoG accumulates within in the stem-loop hairpin generated during BER as a result of a reducedaffinity and activity of human OGG1 at the hairpin loop [53].Evidence supporting that oxygen radicals and BER are in-volved in the expansion of the CAG repeat comes from studiesconnecting mitochondria dysfunction to HD [54]. Mitochon-dria are responsible for the production of most of the energy incells. Neurons have intense energy demands and limited re-generative capacity, making them vulnerable for mitochondriadysfunction [55]. Individuals with HD have impaired mito-chondria dynamics (shape, size, distribution, etc.), and there isincreasing evidence of altered mitochondria trafficking anddysregulation as a cause rather than consequence of HD [56].Mitochondria dysfunction leads to an increased load of oxygenradicals followed by activation of the BER enzymes, connect-ing DNA repair to the pathology of HD.

Mismatch Repair

MMR is a highly conserved DNA repair system that greatlycontributes to maintain genome stability through the correc-tion of mismatched base pairs [57, 58]. The major source ofmismatched base pairs is replication errors, such as incor-porations of mismatched nucleotides or insertions and dele-tion loops, resulting from DNA polymerase slippage. Inaddition, mismatches can occur in DNA because of damageto the nucleotide precursors in the cellular nucleotide pool orby spontaneous deamination of 5-methylcytosin to thymine(G-T mismatch) or cytosine to uracil (G-U) mismatch. If leftunrepaired, these errors may result in permanent mutationsthat could change the behavior of a cell and give rise totumorigenesis. In brief, this pathway involves four keyprocesses: recognition of the erroneous bases or insertion–deletion loops, excision of these lesions, substitution of thelesion with the correct sequence, and religation of the DNA(Fig. 4). The MMR system includes the two key compo-nents, MutS and MutL, which are highly conserved acrossspecies. The initial recognition of mismatches is performedby hMutS, which is found in two major forms, as hMutSα (ahMSH2/hMSH6 dimer) or hMutSβ (a hMSH2/hMSH3 di-mer). hMutSα recognizes base–base mismatches and shortinsertion–deletion loops, whereas hMutSβ detects largermismatches up to 13 nucleotides [59–61]. When hMutSbinds to a DNA lesion, it initiates the recruitment of hMutL,a second protein complex normally composed of MLH1 andPMS2. hMutL coordinates the recruitment of additionalproteins such as exonuclease I, DNA polymerases, andreplication factors to complete the repair process. In prokar-yotes and eukaryotes, deletion of the highly conservedMMR genes mutS and mutL results in several hundred-fold increase in mutation frequency [62]. In humans, germ-line MMR defects predispose people to several differentcancers [63], underscoring the importance of MMR in main-taining genetic stability. MMR is intimately associated withsomatic CAG expansion in HD. The deletion of MSH2 inHD mice abolished somatic expansion and loss of MSH3abrogated somatic expansion in transgenic mouse models ofHD. Loss of MSH6, however, does not prevent somaticexpansion (reviewed by Slean et al. [64]). Seriola et al.recently demonstrated a correlation between repeat instabil-ity and MMR in myotonic dystrophy hESCs, where a nat-ural MMR downregulation upon differentiation leads to astabilization of the CTG repeat expansion [65].

Neural Sensitivity to DNA Damage

Research over the past few decades has revealed the wide-spread involvement of oxidative stress in a number of dis-eases, most notably those that have increased incidence with

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age. A common pathological feature of neurological dis-eases is the loss of a subset of neurons in the central nervoussystem [66–68]. The central nervous system is particularlyvulnerable to oxidative insults on account of the high rate ofO2 utilization and relatively low concentrations of classicalantioxidants and related enzymes [69]. A large body ofevidence suggests that oxidative stress may play a signifi-cant role in the pathogenesis of HD. Recently, Kovtun et al.suggested a role for an oxidative damage repair protein inTNR instability by identifying a molecular link between theBER enzyme OGG1 and somatic CAG/CTG repeat expan-sion [13]. OGG1 creates single-stranded DNA breaks duringthe repair process, which may cause long CAG repeats toexpand. Dramatic somatic CAG expansions have beendetected in diseased human brains and in transgenic HDmice [70]. Yet, the mechanism of expansion remains poorlyunderstood. There is also evidence that MMR proteins arerequired for the active mutagenesis of expanded CAGrepeats, but the biological mechanism is still unclear. Stud-ies in transgenic mice containing CAG repeats have shownthat certain MMR proteins are required for the somaticincrease in repeat length. Particularly, in mice deficient forMSH2, MSH3, or PMS2, repeats are somatically stabilized,whereas deficiencies in MSH6 either lead to no change insomatic instability or else an increase in expansion [64].

Moreover, deficiency of MSH2 or MSH3 resulted in CAGrepeat stability in all tissues tested. In contrast, an OGG1deficiency only affected instability in a subset of tissue andto a lesser degree than MMR deficiencies. Therefore, Sleanet al. [64] suggested that there may be at least two distinctpathways for CAG repeat instability: one which is OGG1-dependent and independent of MSH2-MSH3 and the otheris MSH2-MSH3-dependent, but independent of OGG1. Itremains to be seen if both OGG1 and MMR cooperatephysically in the process of CAG repeat.

Somatic Instability of CAG Repeats in Different Tissueand Its Role in Disease Pathogenesis

The CAG repeat in HD patients is somatic, unstable, andcontinues to expand throughout the lifetime of the individ-ual [70, 71]. Analysis of postmortem brain tissue from HDpatients has revealed high levels of variation in the numberof CAG repeats in somatic tissue and very large expansionin the striatum and cortex [72]. The somatic expansionoccurs in postmitotic neurons, which are the brain regionsthat are targets of neurodegeneration [73]. How the somaticinstability of CAG repeats is related to the differences inrepeat instability in different tissues and at different ages are

Fig. 4 A simplified view ofmismatch repair. Mispaired orunpaired bases are recognizedby the MSH2-MSH6 (MutSα)dimer, and insertions/deletionsare recognized by the MSH2-MSH3 dimer. Binding ofMSH2-MSH6/MSH2-MSH3stimulates binding of theMLH1–PMS2 (MutLα)complex. Excision and repairsynthesis is performed by ExoI,PCNA, and Polδ. MSH, MutShomologues; PMS, postmeioticsegregation protein; EXO1,exonuclease 1; PCNA,proliferating cell nuclearantigen; Pol, DNA polymerase;Lig, DNA ligase

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puzzling open questions. The age of onset of the HD diseaseis strongly correlated to the number of CAG repeats, butseveral studies have demonstrated that hereditary factors arealso involved [74–76]. Since these modifiers directly alteronset age in HD patients, identifying these factors is animportant step towards developing therapeutic targets inHD patients. How significant is somatic instability in theHD pathogenic process? It has recently been proposed thatsomatic expansion over a certain “pathological threshold” of115 CAG repeats is required before HD symptoms ensue[77]. Therefore, somatic expansion is necessary for diseaseonset. That would predict that, starting at the same consti-tutive repeat length, individuals with more somatic expan-sion would reach this threshold earlier with less somaticexpansion and therefore earlier disease onset.

Pluripotent Stem Cells as a Model Systemfor Huntington’s Disease

Despite the known genetic basis for HD, insight into thedisease mechanisms and identification of effective therapiesremain elusive due to the lack of an appropriate modelsystem. Until now, research on Huntington’s disease andmany other neurological disorders relay on rodent modelsthat may not fully recapitulate human diseases. Postmortemtissue can provide a window into alterations in human brainstructure at a cellular and molecular level, and efforts havealso been made to establish primary cultures from postmor-tem brain tissue, but this has proven very difficult. Analternative source of human HD tissue is fibroblasts; how-ever, seeing HD as a neurodegenerative disease, these cellsare not particularly informative. Stem cells have the capacityto proliferate and differentiate into multiple lineages. Re-search on human embryonic stem cells (hESCs) started inthe late nineties when Thomson and colleagues derivedhESCs from the inner cell mass of a developing blastocyst[78]. The opportunity to generate neuron in a dish in thelaboratory created enthusiasm in the field. ESCs affordoptions for disease modeling and research on molecularmechanism behind HD. In addition, ESCs also have thepotential to restore functional loss of medium spiny neuronswhich is lost in HD patients. Injections of neural progenitorstem cells into HD rodents demonstrated incorporation aswell as migration to secondary sites associated with thedisease [79].

More recently, the development of iPSCs has provided anadditional source for autologous stem cells for modeling andtreating diseases. iPSCs are generated from somatic tissuesuch as fibroblasts derived from skin biopsies. Selectedtranscription factors reprogram these fibroblasts into ES-like cells, and multiple research groups have now success-fully reprogrammed fibroblasts using vector, virus, protein,

or RNA-mediated approaches to deliver the transcriptionfactors (Fig. 4) [80–86]. iPSCs afford options for diseasemodeling and provide novel sources for autologous cellulartherapies. In 2012, several papers published on humaniPSCs were related to HD [24, 87–91]. One of the papers,a study by An and colleagues, represents a major improve-ment because they have demonstrated that iPSCs from a HDpatient can be corrected to have two normal HTT alleles byhomologous recombination [24]. The authors furtherreported that the correction persisted during differentiationto neurons and reversed the disease phenotype in a HDmouse model when transplanted into the striatum. Thisstudy is exciting, because the ability to make patient-specific, genetically corrected iPSCs from HD patients willgive relevant disease models in identical genetic back-grounds, which brings us a step closer to clinical applica-tions. Several experiments have shown that fetal tissuetransplanted into the brain of rodents and primate modelsof HD survive and integrate well into the damaged area ofthe brain and reverse motor and cognitive dysfunctions [92,93]. A clinical trial where fetal tissue was transplanted intoHD patients conferred a certain degree of motor and cogni-tive improvement for 2–3 years, but not permanently [94].Currently ongoing clinical trials with fetal tissue (e.g., StudyNCT0019045 on www.clinicaltrials.gov) would providevaluable information on the efficacy of the fetal tissue-mediated therapeutic approach.

However, iPSC technology is still new, and geneticprogramming may create differences that may be harmful.Therefore, careful characterization of patient iPS cells mustbe performed. With the continuous advancement of iPStechnology, however, directed differentiation of patient iPScells may be utilized to model HD processes for mechanisticand therapeutic discovery.

Conclusions and Future Directions

Since the discovery of the HD gene in 1993, tremendousprogress has been made in understanding the molecularmechanisms underlying the pathology of HD. However, stillno effective cure exists for the disease. Recent research hasfocused on selectively blocking, or reducing, the expressionof HTT protien in HD patients. Also, inhibitors of certainmembers of either the BER or MMR DNA repair pathwaysmay prove to slow down the development of HD. Furtherresearch is required to investigate these possible treatments.The latest addition to therapeutic approaches for HD is theiPSC technology and in vitro differentiation of cells towardsthe neural lineage. Such cells have been used with success totreat animal models of HD, and future studies on humansubjects may establish this as the definitive treatment to cureHD.

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Conflict of Interest The authors declare no financial or other conflictof interests.

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