Methods of treating or inhibiting onset of huntington's disease

Abstract

The disclosure herein generally relates to methods of treating or inhibiting onset of Huntington's disease. Such methods involve selecting a subject having or at risk of having Huntington's disease, and administering to the subject one or more modulators of one or more genes as described herein or proteins encoded thereby under conditions effective to treat or inhibit onset of Huntington's disease in the subject.

Description

[0001] This application is a divisional application of a Chinese patent application with the application number 201980054882.0, the filing date of June 19, 2019, and the invention title of "Methods for Treating or Suppressing the Onset of Huntington's Disease", and the original application is a national stage application with the international application number PCT / US2019 / 037987. This application claims the benefit of U.S. Provisional Patent Application Serial No. 62 / 688,174, filed June 21, 2018, which is hereby incorporated by reference in its entirety. Field of the Invention

[0002] This application relates to methods for treating or suppressing the onset of Huntington's disease. Background of the Invention

[0004] Huntington's disease (HD) is a fatal autosomal dominant neurodegenerative disorder characterized by progressive behavioral, cognitive, and motor dysfunction. HD is caused by a CAG trinucleotide repeat in the first exon of the huntingtin (HTT) gene, which encodes an expanded polyglutamine. The age of onset and severity are proportional to the length of this repeat expansion, where a CAG length of more than 35 inevitably leads to clinical disease. This is associated with the intracellular accumulation and aggregation of mutant HTT (mHTT), which results in neuronal loss.Although the pathological changes in HD are mainly reflected by the progressive loss of medium spiny neurons (MSNs) in the striatum and the consequent striatal atrophy, MRI studies have shown that HD is also characterized by early demyelination and white matter loss, which may occur before the onset of symptoms (Tabrizi et al., “Potential Endpoints for Clinical Trials in Premanifest and Early Huntington's Disease in the TRACK-HD Study: Analysis of 24 Month Observational Data,” The Lancet Neurology 11:42-53 (2012)). Similarly, studies in mouse models of HD have revealed early myelination defects (Teo et al., “Structural and Molecular Myelination Deficits Occur Prior to Neuronal loss in the YAC128 and BACHD Models of Huntington Disease,” Human Molecular Genetics 25:2621-2632 (2016)), associated with defects in the key myelinogenic gene MYRF (Huang et al., “Mutant Huntingtin Downregulates Myelin Regulatory Factor-Mediated Myelin Gene Expression and Affects Mature Oligodendrocytes,” Neuron 85:1212-1226 (2015); Jin et al., “Early White Matter Abnormalities, Progressive Brain Pathology and Motor Deficits in a Novel Knock-In Mouse Model of Huntington's Disease,” Human Molecular Genetics 24:2508-2527 (2015)). Collectively, these observations suggest that the pathological changes in HD are associated with white matter loss, which in turn may reflect dysfunction of myelin-producing oligodendrocytes.

[0005] However, although these data suggest the presence of white matter abnormalities and myelination disorders in HD, and parallel studies have shown that glial replacement can improve the symptoms of HD transgenic mice (Benraiss et al., “Human Glia can Both Induce and Rescue Aspects of Phenotype in Huntington Disease,” Nature Communications 7:11758 (2016)), the cellular and molecular bases of glial pathology in human HD have not been well explored.

[0006] The present disclosure aims to overcome these and other deficiencies in the prior art. SUMMARY OF THE INVENTION

[0007] A first aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such method involves selecting a subject having or at risk of having Huntington's disease, and administering to the selected subject one or more modulators of a glial cell differentiation regulatory gene, the glial cell differentiation regulatory gene selected from the group consisting of: BMP2, LINGO1, MAG, NKX2-2, NR2E1, NTRK3, OLIG2, SERPINE2, SIRT2, and TCF7L2, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0008] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such method involves selecting a subject having or at risk of having Huntington's disease, and administering to the selected subject one or more modulators of a gene associated with myelination, the gene associated with myelination selected from the group consisting of: FA2H, GAL3ST1, MAG, MBP, MYRF, NFASC, OLIG2, OMG, PLLP, POU3F2, SIRT2, SLC8A3, TCF7L2, TF, and UGT8, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0009] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such method involves selecting a subject having or at risk of having Huntington's disease, and administering to the selected subject one or more modulators of an oligodendrocyte differentiation gene, the oligodendrocyte differentiation gene selected from the group consisting of: FA2H, GLI3, LINGO1, MYRF, NKX2-2, OLIG1, OLIG2, OMG, SIRT2, SLC8A3, SOX10, and TCF7L2, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0010] Another aspect of the present disclosure relates to a method for treating or suppressing the onset of Huntington's disease. This method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a gliogenesis-regulating gene, which gliogenesis-regulating gene is selected from the group consisting of: BMP2, LINGO1, MAG, MYC, NKX2-2, NR2E1, NTRK3, OLIG2, SERPINE2, SIRT2, SOX10, TCF7L2, TF, and ZCCHC24, or a protein encoded thereby.

[0011] Another aspect of the present disclosure relates to a method for treating or suppressing the onset of Huntington's disease. This method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a neuron ensheathment gene, which neuron ensheathment gene is selected from the group consisting of: FA2H, GAL3ST1, MAG, MBP, MYRF, NFASC, OLIG2, OMG, PLLP, POU3F2, SIRT2, SLC8A3, TCF7L2, TF, and UGT8, or a protein encoded thereby.

[0012] Another aspect of the present disclosure relates to a method for treating or suppressing the onset of Huntington's disease. This method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of an axon guidance gene, which axon guidance gene is selected from the group consisting of: ALCAM, BCL11B, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, MNX1, NFASC, PLXNC1, PRKCQ, PTPRO, ROBO2, SEMA6B, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0013] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a neuronal projection guidance gene, the neuronal projection guidance gene selected from the group consisting of: ALCAM, BCL11B, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, MNX1, NFASC, PLXNC1, PRKCQ, PTPRO, ROBO2, SEMA6B, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0014] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of an axonogenesis gene, the axonogenesis gene selected from the group consisting of: ADGRB1, ALCAM, BCL11B, CACNA1A, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, LINGO1, LRRC4C, MAG, MBP, MNX1, NFASC, NR2E1, NTNG1, NTRK3, OMG, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, SEMA6B, SLITRK2, SLITRK3, SNAP91, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0015] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of an axon development gene, the axon development gene selected from the group consisting of: ADGRB1, ALCAM, BCL11B, CACNA1A, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, LINGO1, LRRC4C, MAG, MBP, MNX1, NEFM, NFASC, NR2E1, NTNG1, NTRK3, OMG, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, RTN4RL2, SEMA6B, SLITRK2, SLITRK3, SNAP91, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0016] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a cell projection morphogenesis gene, the cell projection morphogenesis gene being selected from the group consisting of: ADGRB1, ALCAM, BCL11B, CACNA1A, CAMK2A, DSCAM, EHD3, FOXD1, GAS1, GLI3, HOXA1, HOXA2, KANK1, LINGO1, LRRC4C, MAG, MBP, MNX1, NEDD4L, NEURL1, NFASC, NR2E1, NTNG1, NTRK3, OMG, PCDH15, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, SEMA6B, SGK1, SLITRK2, SLITRK3, SNAP91, SNX10, UGT8, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0017] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a synapse structure or activity regulatory gene, the synapse structure or activity regulatory gene being selected from the group consisting of: ADGRB1, ADGRL3, BCAN, CALB1, CAMK2A, FGF14, LRRTIM1, NCDN, NETO1, NEURL1, NR2E1, NTRK3, PPFIA3, ROBO2, SERPINE2, SHISA7, SIX4, SLC8A3, SLITRK2, SLITRK3, and SYNDIG1, or a protein encoded thereby.

[0018] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a synaptic signaling pathway gene, the synaptic signaling pathway gene being selected from the group consisting of: BCAN, CACNA1A, CACNA1G, CALB1, CAMK2A, CHRNA4, FGF12, FGF14, GRIA2, GRIA4, GRID2, GRIK4, KCND2, LRRTM1, MBP, MPZ, NCDN, NETO1, NEURL1, NOVA1, NR2E1, P2RX7, PDE7B, PLCL1, PPFIA3, RAPGEF4, RGS8, RIT2, S1PR2, SERPINE2, SHISA7, SLC18A1, SLC1A1, SLC1A2, SLC8A3, SNAP91, SNPH, and SYT6 or a protein encoded thereby.

[0019] Another aspect of the present disclosure relates to a method of treating or suppressing the onset of Huntington's disease. Such a method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a synaptic gene, the synaptic gene being selected from the group consisting of: ADGRB1, BCAN, BCAS1, CACNA1A, CALB1, CAMK2A, CHRNA4, CTTNBP2, DSCAM, GRIA2, GRID1, GRID2, GRIK4, HCN2, KCND2, LGI3, LRRC4C, LRRTM1, NETO1, NEURL1, NTM, P2RX7, PCDH15, PDE4B, PPFIA3, PRIMA1, PRKCQ, PTPRO, RAPGEF4, SERPINE2, SHISA7, SLC17A8, SLC18A1, SLC1A1, SLC1A2, SLC8A3, SNAP91, SNPH, SYNDIG1, and SYT6, or a protein encoded thereby.

[0020] Another aspect of the present disclosure relates to methods of treating or inhibiting the onset of Huntington's disease. Such methods involve selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a monovalent inorganic cation transporter gene, the monovalent inorganic cation transporter gene selected from the group consisting of: ABCC9, ASIC4, CACNA1A, CHRNA4, CNGB1, CNTN1, DPP10, DPP6, FGF12, FGF14, HCN2, KCND2, KCNJ9, KCNQ1, KCNS3, NALCN, NEDD4L, NKAIN4, P2RX7, PTGER3, SERPINE2, SGK1, SLC10A4, SLC17A8, SLC18A1, SLC22A3, SLC2A13, SLC5A9, SLC8A3, and SLC9A7, or a protein encoded thereby.

[0021] Another aspect of the present disclosure relates to methods of treating or inhibiting the onset of Huntington's disease. Such methods involve selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a neuronal projection gene, the neuronal projection gene selected from the group consisting of: ADGRL3, ALCAM, BCAN, BCL11B, CACNA1A, CACNA1G, CALB1, CAMK2A, CHRNA4, CTTNBP2, DSCAM, GRIA2, GRIA4, GRID2, GRIK4, HCN2, KCND2, LGI3, LRRTM1, MAG, MBP, MYC, NCAM2, NCDN, NEFM, NEURL1, NFASC, NTM, PDE4B, PIK3R1, PTGER3, PTPRO, RAPGEF4, RGS8, ROBO2, SGK1, SIRT2, SLC17A8, SLC1A2, SLC8A3, SNAP91, SNPH, SYNDIG1, and UNC5A, or a protein encoded thereby.

[0022] Another aspect of the present disclosure relates to a method for treating or suppressing the onset of Huntington's disease. This method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a TCF7L2 target gene, the TCF7L2 target gene consisting of the group consisting of: BMP4, CCND1, CCND2, DOCK10, DOCK9, DUSP15, ENPP4, EPAS1, EPHB1, ERBB3, EVI2A, EVI2B, FA2H, GJB1, HAPLN2, HSPA2, ID3, LGI3, MBP, MOG, MYC, MYRF, NFASC, NKAIN1, NKX6-2, OLIG2, PLEKHB1, PLP1, PPP1R16B, RAB33A, RASGEF1B, RTKN, SIRT2, SLC1A2, SOX10, ST18, TMEM125, TMEM2, TPPP, TSPAN15, UGT8 and AATK, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0023] Another aspect of the present disclosure relates to a method for treating or suppressing the onset of Huntington's disease. This method involves selecting a subject having Huntington's disease or at risk of developing Huntington's disease, and administering to the selected subject one or more modulators of a gene or protein encoded thereby involved in the NKX2.2→OLIG2→SOX10→MYRF regulatory cascade under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0024] This disclosure examined whether the gene expression patterns of human glial progenitor cells (hGPCs) expressing mHTT could reflect cell-autonomous molecular pathological changes, and if so, whether they could predict white matter disease in HD. Bipotent oligodendrocyte-astrocyte hGPCs were initially generated from human embryonic stem cells (hESCs) derived from Huntington protein mutant embryos or their sibling controls. These cells were then isolated using fluorescence-activated cell sorting (FACS) based on the expression of the GPC-selective CD140a (Sim et al., “CD140a Identifies a Population of Highly Myelinogenic, Migration-Competent and Efficiently Engrafting Human Oligodendrocyte Progenitor Cells,” Nat. Biotechnol. 29:934-941 (2011); Wang et al., “Human iPSC-Derived Oligodendrocyte Progenitor Cells Can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12:252-264 (2013), which are hereby incorporated by reference in their entirety), followed by whole-transcriptome RNA sequencing (RNA-seq) analysis to assess mHTT-dependent changes in their gene expression. We found that in hGPCs generated from hESCs derived from three different HD embryos, a coherent set of key transcription factors associated with astroglial and oligodendroglial differentiation and with downstream myelin biosynthesis was significantly downregulated as a function of mHTT expression relative to controls. Thus, when HD hESC-derived hGPCs were transplanted into neonatal myelin-deficient immunodeficient shiverer mice (MBP shi / shi ), myelination of the resulting glial chimeras was slower and less complete than that of littermate controls transplanted with hGPCs derived from normal control hESCs. In addition, chimeras established with HD hGPCs exhibited a significant delay and disruption of astrocyte morphogenesis relative to mice chimerized with normal sibling hGPCs. Collectively, these data suggest that white matter failure and reduced myelination in human HD may be the result of cell-autonomous defects in the terminal glial differentiation of hGPCs expressing mHTT, rather than secondary to neuronal loss, and that the occurrence of such defects may be central to the pathogenesis and neurological manifestations of HD.

[0025] The present invention also includes at least the following embodiments:

[0026] Embodiment 1. A method for treating or suppressing the onset of Huntington's disease, the method comprising:

[0027] selecting a subject having Huntington's disease or at risk of having Huntington's disease; and

[0028] administering to the selected subject one or more modulators of a glial cell differentiation regulatory gene, the glial cell differentiation regulatory gene selected from the group consisting of: BMP2, LINGO1, MAG, NKX2-2, NR2E1, NTRK3, OLIG2, SERPINE2, SIRT2, and TCF7L2, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0029] Embodiment 2. The method according to Embodiment 1, wherein the one or more modulators are selected from the group consisting of: Hh-Ag 1.1, Hh-Ag 1.2, Hh-Ag 1.3, Hh-Ag1.4, Hh-Ag 1.5, 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, opicinumab, GSK-249320, sodium dodecyl sulfate, repaglinide, atelatinib, chembl2007421, PLX-3397, radicicol, thyroxine, entrectinib, LOXO-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, midostaurin, and combinations thereof.

[0030] Embodiment 3. A method for treating or suppressing the onset of Huntington's disease, the method comprising:

[0031] selecting a subject having Huntington's disease or at risk of having Huntington's disease; and

[0032] administering to the selected subject one or more modulators of a gene associated with myelination, the gene associated with myelination selected from the group consisting of: FA2H, GAL3ST1, MAG, MBP, MYRF, NFASC, OLIG2, OMG, PLLP, POU3F2, SIRT2, SLC8A3, TCF7L2, TF, and UGT8, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0033] Embodiment 4. The method according to Embodiment 3, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, GSK-249320, sodium dodecyl sulfate, repaglinide, cyclosporine, interferon β-1A, prednisone, quercetin, rutin, and combinations thereof.

[0034] Embodiment 5. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0035] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0036] administering to the selected subject one or more modulators of an oligodendrocyte differentiation gene, the oligodendrocyte differentiation gene being selected from the group consisting of: FA2H, GLI3, LINGO1, MYRF, NKX2-2, OLIG1, OLIG2, OMG, SIRT2, SLC8A3, SOX10, and TCF7L2, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0037] Embodiment 6. The method according to Embodiment 5, wherein the one or more modulators are selected from the group consisting of: Hh-Ag 1.1, Hh-Ag 1.2, Hh-Ag 1.3, Hh-Ag1.4, Hh-Ag 1.5, 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, opalizumab, sodium dodecyl sulfate, repaglinide, vemurafenib, and combinations thereof.

[0038] Embodiment 7. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0039] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0040] administering to the selected subject one or more modulators of a gliogenesis regulatory gene, the gliogenesis regulatory gene being selected from the group consisting of: BMP2, LINGO1, MAG, MYC, NKX2-2, NR2E1, NTRK3, OLIG2, SERPINE2, SIRT2, SOX10, TCF7L2, TF, and ZCCHC24, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0041] Embodiment 8. The method according to Embodiment 7, wherein the one or more modulators are selected from the group consisting of: Hh-Ag 1.1, Hh-Ag 1.2, Hh-Ag 1.3, Hh-Ag1.4, Hh Ag 1.5, 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, opizumab, GSK-249320, sodium dodecyl sulfate, vemurafenib, repaglinide, nadroparin calcium, 4'-hydroxytamoxifen, azacitidine, thioguanine, activin, adozelesin, amifostine, aminopterin, antibiotics, bizelesin, bromocriptine, bryostatin, calcitriol, diethylstilbestrol, esafosfamide, estrone, folic acid, glutamine, hypoxanthine, imatinib, cilastatin, melatonin, methylprednisolone, N-methyl-n-nitrosourea, novobiocin, Chembl35482, phorbol 12-myristate 13-acetate, prednisone, quinapril, vorinostat, sulindac, thrombin, thyrotropin, β-nicotinamide adenine dinucleotide phosphate sodium, troglitazone, verapamil, Chembl100014, Chembl1213492, chorionic gonadotropin, perillyl alcohol, AMG-900, Alisertib, Dinaciclib, Roniciclib, temozolomide, plerixafor, atelatinib, chembl2007421, PLX-3397, radicicol, thyroxine, entrectinib, LOXO-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, midostaurin, and combinations thereof.

[0042] Embodiment 9. A method of treating or suppressing the onset of Huntington's disease, the method comprising:

[0043] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0044] administering to the selected subject one or more modulators of a neuron ensheathing gene, the neuron ensheathing gene being selected from the group consisting of FA2H, GAL3ST1, MAG, MBP, MYRF, NFASC, OLIG2, OMG, PLLP, POU3F2, SIRT2, SLC8A3, TCF7L2, TF, and UGT8, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0045] Embodiment 10. The method according to embodiment 9, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, GSK-249320, cyclosporine, interferon beta-1A, prednisone, quercetin, rutin, sodium dodecyl sulfate, repaglinide, and combinations thereof.

[0046] Embodiment 11. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0047] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0048] administering to the selected subject one or more modulators of an axon guidance gene, the axon guidance gene being selected from the group consisting of: ALCAM, BCL11B, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, MNX1, NFASC, PLXNC1, PRKCQ, PTPRO, ROBO2, SEMA6B, UNC5A, VAX1, and WNT7B or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0049] Embodiment 12. The method according to embodiment 11, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, fluorouracil, CEP-2563, staurosporine, Chembl369507, D-phosphoserine, ticlopidine, GSK-690693, sotrastaurin, (7S)-hydroxy-staurosporine, midostaurin, quercetin, bryostatin, acetate sotrastaurin, ingenol mebutate, carboplatin, paclitaxel, and combinations thereof.

[0050] Embodiment 13. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0051] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0052] Administering to a selected subject one or more modulators of a neuronal projection guidance gene, selected from the group consisting of ALCAM, BCL11B, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, MNX1, NFASC, PLXNC1, PRKCQ, PTPRO, ROBO2, SEMA6B, UNC5A, VAX1, and WNT7B, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0053] Embodiment 14. The method of embodiment 13, wherein the one or more modulators are selected from the group consisting of 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, fluorouracil, CEP-2563, staurosporine, Chembl369507, D-3-phosphoserine, ticlopidine, GSK-690693, sertraline, (7S)-hydroxy-staurosporine, midostaurin, quercetin, bryostatin, sertraline acetate, ingenol mebutate, carboplatin, paclitaxel, and combinations thereof.

[0054] Embodiment 15. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0055] Selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0056] Administering to the selected subject one or more modulators of an axonogenesis gene, selected from the group consisting of ADGRB1, ALCAM, BCL11B, CACNA1A, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, LINGO1, LRRC4C, MAG, MBP, MNX1, NFASC, NR2E1, NTNG1, NTRK3, OMG, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, SEMA6B, SLITRK2, SLITRK3, SNAP91, UNC5A, VAX1, and WNT7B, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0057] Embodiment 16. The method according to embodiment 15, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, opium monoclonal antibody, GSK-249320, cyclosporin, interferon β-1A, prednisone, quercetin, rutin, fluorouracil, CEP-2563, staurosporine, Chembl369507, right phosphoserine, ticlopidine, GSK-690693, sotrastaurin, (7S)-hydroxy-staurosporine, midostaurin, bryostatin, acetate sotrastaurin, ingenol mebutate, carboplatin, paclitaxel, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, hydrochloric acid bepridil, amecarbazone, atalatinib, chembl2007421, PLX-3397, radicicola, thyroxine, entrectinib, Loxo-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, and combinations thereof.

[0058] Embodiment 17. A method for treating or suppressing the onset of Huntington's disease, the method comprising:

[0059] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0060] administering to the selected subject one or more modulators of an axon development gene, the axon development gene being selected from the group consisting of ADGRB1, ALCAM, BCL11B, CACNA1A, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, LINGO1, LRRC4C, MAG, MBP, MNX1, NEFM, NFASC, NR2E1, NTNG1, NTRK3, OMG, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, RTN4RL2, SEMA6B, SLITRK2, SLITRK3, SNAP91, UNC5A, VAX1, and WNT7B, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0061] Embodiment 18. The method according to Embodiment 17, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, opitzumab, D-phosphoserine, fluorouracil, CEP-2563, staurosporine, Chembl369507, GSK-249320, ticlopidine, GSK-690693, sertraline, (7S)-hydroxy-staurosporine, midostaurin, quercetin, bryostatin, sertraline acetate, ingenol mebutate, carboplatin, paclitaxel, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, bepridil hydrochloride, amocarbose, atalatinib, chembl2007421, PLX-3397, radicicola, thyroxine, entrectinib, Loxo-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, cyclosporine, interferon beta-1A, prednisone, rutin, and combinations thereof.

[0062] Embodiment 19. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0063] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0064] administering to the selected subject one or more modulators of a cell projection morphogenesis gene under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the cell projection morphogenesis gene being selected from the group consisting of ADGRB1, ALCAM, BCL11B, CACNA1A, CAMK2A, DSCAM, EHD3, FOXD1, GAS1, GLI3, HOXA1, HOXA2, KANK1, LINGO1, LRRC4C, MAG, MBP, MNX1, NEDD4L, NEURL1, NFASC, NR2E1, NTNG1, NTRK3, OMG, PCDH15, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, SEMA6B, SGK1, SLITRK2, SLITRK3, SNAP91, SNX10, UGT8, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0065] Embodiment 20. The method according to embodiment 19, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, opizumab, GSK-249320, cyclosporine, interferon beta-1A, prednisone, quercetin, rutin, D-phosphoserine, fluorouracil, CEP-2563, staurosporine, Chembl369507, ticlopidine, GSK-690693, sertraline, (7S)-hydroxy-staurosporine, midostaurin, bryostatin, sertraline acetate, ingenol mebutate, carboplatin, paclitaxel, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, bepridil hydrochloride, amocarbazone, atalatinib, Chembl2007421, PLX-3397, radicicola, thyroxine, entrectinib, Loxo-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, hydrochlorothiazide, chembl549906, chembl550795, sodium chloride, GSK-650394, and combinations thereof.

[0066] Embodiment 21. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0067] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0068] administering to the selected subject one or more modulators of a synapse structure or activity regulatory gene, the synapse structure or activity regulatory gene being selected from the group consisting of ADGRB1, ADGRL3, BCAN, CALB1, CAMK2A, FGF14, LRRTIM1, NCDN, NETO1, NEURL1, NR2E1, NTRK3, PPFIA3, ROBO2, SERPINE2, SHISA7, SIX4, SLC8A3, SLITRK2, SLITRK3, and SYNDIG1 or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0069] Embodiment 22. The method according to embodiment 21, wherein the one or more modulators are selected from the group consisting of: L-serine phosphate, atalatinib, chembl2007421, PLX-3397, radicicola, thyroxine, entrectinib, Loxo-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, midostaurin, and combinations thereof.

[0070] Embodiment 23. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0071] selecting a subject having or at risk of having Huntington's disease; and

[0072] administering to the selected subject one or more modulators of a synaptic signaling pathway gene, the synaptic signaling pathway gene being selected from the group consisting of: BCAN, CACNA1A, CACNA1G, CALB1, CAMK2A, CHRNA4, FGF12, FGF14, GRIA2, GRIA4, GRID2, GRIK4, KCND2, LRRTM1, MBP, MPZ, NCDN, NETO1, NEURL1, NOVA1, NR2E1, P2RX7, PDE7B, PLCL1, PPFIA3, RAPGEF4, RGS8, RIT2, S1PR2, SERPINE2, SHISA7, SLC18A1, SLC1A1, SLC1A2, SLC8A3, SNAP91, SNPH, and SYT6 or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0073] Embodiment 24. The method according to Embodiment 23, wherein the one or more modulators are selected from the group consisting of: pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, bepridil hydrochloride, ameixin, cyclosporine, interferon β-1A, prednisone, quercetin, rutin, polacrilex nicotine, tabarbital, secbutabarbital, butabarbital, secobarbital, metharbital, thiopental, primidone, tolbutabarbital, phenobarbital, varenicline, amobarbital, aprobarbital, butessa, heptabarbital, hexobarbital, barbital, prazacrine, cytisine, rivanicline, epibatidine, chembl1876219, chembl3103988, atracurium, chembl490153, hexamethonium, chembl407217, TC-2216, ABT-560, isopropylcran, solfenicline, TC-6499, AZD1446, CP-601927, dexmedetomidine, nicotine, varenicline tartrate, benzatropine mesylate, pentolonium, azd0328, bradanicline, pentobarbital, chembl1201135, dexfenfluramine, mecamylamine (chembl267936), dianicline, aticline, mivacurium, oleic acid, tibalcrine tosylate, mibampator, butessa, (r,s)-AMPA, chembl123132, aniracetam, chembl136800, chembl1255648, cyclothiazide, chembl77862, chembl334920, chembl1097939, piracetam, chembl320642, chembl265301, gyki-52466, nbqx, chembl222418, teixampanel, (s)-AMPA, chembl594840, chembl121915, quisqualic acid, chembl337577, chemb l27130, dnqx, chembl333964, (s)-willardiine, chembl28472, talampanel, perampanel, elempanel, CX1739, darampanel, becampanel, farapato, mk-8777, zonampanel, pentobarbital, pf-04958242, seropanel, dalfampridine, guanidine hydrochloride, tedisamil, neripridine, evt401, adenosine triphosphate, chembl335550, chelerythrine, acebutolol, moclobemide, ivermectin, chemb377219, chembl255787, methylclothiazide 、chembl550637、sodium orthovanadate、chembl2338352、benzonatate、GSK1482160、AZD9056、CE224535、xanthin、chembl484928、dipyridamole、flavonoid hydrochloride、pentoxifylline、quinacrine、chembl2313646、chembl570352、ozanimod、chembl225155、chembl1368758、fingolimod hydrochloride、amisimod hydrochloride、reserpine、norepinephrine、chembl126506、methamphetamine、ketanserin、tetrabenazine、L -glutamate, dihydrocainate, 2s,4r-4-methylglutamate, o-benzyl-l-serine, chembl1628669, teixampanel, domoic acid, dysiherbaine, kainic acid, mesalamine, topiramate, aspartic acid, clozapine, alcohol, haloperidol, wortmannin, olanzapine, phorbol myristate acetate, risperidone, lidocaine, mibefradil dihydrochloride, trimethadione, cinnarizine, ethosuximide, zonisamide, cannabinoids, mibefradil, chembl1684954, flunarizine, methsuximide, phensuximide, ethadione, celecoxib, and combinations thereof. ,

[0074] Embodiment 25. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0075] Selecting subjects who have Huntington's disease or are at risk for Huntington's disease; and

[0076] Administer to a selected subject one or more modulators of a synaptic gene, selected from the group consisting of ADGRB1, BCAN, BCAS1, CACNA1A, CALB1, CAMK2A, CHRNA4, CTTNBP2, DSCAM, GRIA2, GRID1, GRID2, GRIK4, HCN2, KCND2, LGI3, LRRC4C, LRRTM1, NETO1, NEURL1, NTM, P2RX7, PCDH15, PDE4B, PPFIA3, PRIMA1, PRKCQ, PTPRO, RAPGEF4, SERPINE2, SHISA7, SLC17A8, SLC18A1, SLC1A1, SLC1A2, SLC8A3, SNAP91, SNPH, SYNDIG1, and SYT6, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0077] Embodiment 26. The method according to embodiment 25, wherein the one or more modulators are selected from the group consisting of: L-phosphoserine, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, bepridil hydrochloride, aminocaproic acid, mibampator, butesa, secbutabarbital, butalbital, talbutal, secobarbital, metharbital, thiopental, primidone, mephobarbital, phenobarbital, (R,s)-AMPA, CHEMBL123132, aniracetam, CHEMBL136800, CHEMBL1255648, cyclothiazide, CHEMBL77862, CHEMBL334920, CHEMBL1097939, piracetam, CHEMBL320642, CHEMBL265301, GYKI-52466, NBQX, CHEMBL222418, tezampanel, amobarbital, aprobarbital, heptabarbital, hexobarbital, barbital, (s)-AMPA, CHEMBL594840, CHEMBL121915, quisqualic acid, CHEMBL337577, CHEMBL27130, DNQX, CHEMBL333964, (s)-willardiine, CHEMBL28472, talampanel, perampanel, elampanel, CX1739, darampanel, beclamapanel, farampator, MK-8777, zonampanel, topiramate, pentobarbital, PF-04958242, selurampanel, polacrilex nicotine, varenicline, barbital, pazacrine, cytisine, rivanicline, epibatidine, CHEMBL1876219, CHEMBL3103988, atracurium, CHEMBL490153, hexamethonium, CHEMBL407217, TC-2216, ABT-560, isopropylcranaline, sophinicline, TC-6499, AZD1446, CP-601927, dexmedetomidine, nicotine, varenicline tartrate, benzatropine mesylate, pentolonium, AZD0328, bradanicline, pentobarbital, CHEMBL1201135, dexfenfluramine, mecamylamine (CHEMBL267936), dianicline, aticline, miamine, oleic acid, tibacrine tosylate, polacrilex nicotine, carboplatin, paclitaxel, L-glutamate, dalfampridine, guanidine hydrochloride, tedisamil, niliridine, EVT401, adenosine triphosphate, CHEMBL335550, chelerythrine, acebutolol, moclobemide, ivermectin, CHEMB377219, CHEMBL255787, methyclothiazide, CHEMBL550637, sodium orthovanadate, CHEMBL2338352, benzonatate, GSK1482160, AZD9056, CE224535, reserpine, norepinephrine, CHEMBL126506, methamphetamine, ketanserin, tetrabenazine, dihydrocinchoninate, 2S,4R-4-methylglutamate, O-benzyl-L-serine, chembl1628669, tezampanel, domoic acid, dysiherbaine, kainic acid, mesalamine, topiramate, CEP-2563, staurosporine, Chembl369507, D-phosphoserine, ticlopidine, GSK-690693, surotomycin, (7S)-hydroxy-staurosporine, midostaurin, quercetin, bryostatin, surotomycin acetate, ingenol mebutate, adenosine phosphate, theophylline, dyphylline, pentoxifylline, enprofylline, iloprost, papaverine, theobromine, amrinone, [r]-mexiropramine, roflumilast, piclamilast, rolipram, fevipiprant, chembl1230617, chembl519827, cilomilast, (-)-rolipram, crisaborole, ibudilast, apremilast, chembl521203, chembl74078, propoxyphene, cdp840, sodium phenylbutyrate, chembl1232082, dipyridamole, sodium glycinate theophylline, flavoxate hydrochloride, aminophylline, resveratrol, caffeine, choline theophylline, amlexanox, edazolam, cilobradine, zaltidine, chembl2052019, chembl395336, cyclic adenosine monophosphate, aspartic acid, clozapine, alcohol, haloperidol, wortmannin, olanzapine, phorbol 12-myristate 13-acetate, risperidone, lidocaine, and combinations thereof.,

[0078] Embodiment 27. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0079] selecting a subject having or at risk of having Huntington's disease; and

[0080] administering to the selected subject one or more modulators of a monovalent inorganic cation transport gene, said monovalent inorganic cation transport gene selected from the group consisting of: ABCC9, ASIC4, CACNA1A, CHRNA4, CNGB1, CNTN1, DPP10, DPP6, FGF12, FGF14, HCN2, KCND2, KCNJ9, KCNQ1, KCNS3, NALCN, NEDD4L, NKAIN4, P2RX7, PTGER3, SERPINE2, SGK1, SLC10A4, SLC17A8, SLC18A1, SLC22A3, SLC2A13, SLC5A9, SLC8A3 and SLC9A7, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in said subject.

[0081] Embodiment 28. The method according to embodiment 27, wherein the one or more modulators are selected from the group consisting of: nanonidil, adenosine triphosphate, glibenclamide, saralasin, pinacidil hydrate, minoxidil, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, bepridil hydrochloride, ameixin, chembl549906, chembl550795, sodium chloride, GSK-650394, dalfampridine, guanidine hydrochloride, tedisamil, nilipridin, evt401, adenosine triphosphate, chembl335550, sanguinarine, acebutolol, moclobemide, ivermectin, chemb377219, chembl255787, methyclothiazide, chembl550637, sodium orthovanadate, chembl2338352, benzonatate, GSK1482160, AZD9056, CE224535, hydrochlorothiazide, chembl1229875, polacrilex phosphate nicotine, heptabarbital, secbutabarbital, butabarbital, secobarbital, metharbital, thiopental, primidone, methylphenobarbital, phenobarbital, varenicline, amobarbital, aprobarbital, butessa, heptabarbital, hexobarbital, barbital, pazaracrine, cytisine, rivanicline, epibatidine, chembl1876219, chembl3103988, atracurium, chembl490153, hexamethonium, chembl407217, tc-2216, abt-560, isopropylclan, sophinicline, tc-6499, cilopride, zaltidine, chembl2052019, chembl395336, cyclic adenosine monophosphate, chembl99951, flupirtine, indapamide, azimilide, chembl2070953, mefenamic acid, chembl1907717, niflumic acid, chembl298475, chembl342375, chembl332826, dolasetron, celecoxib, nilipridin, ezogabine, indometacin, tacrolimus, guanidine hydrochloride, tedisamil, dalfampridine, pyrimethamine, cobalt (ii) ion verapamil pyrimethamine cobalt (ii) ion, dihydrocinchonine salt, bimatoprost, dinoprostone, misoprostol, beraprost, chembl1628262, carbacyclin, cicaprost, cloprostenol (chembl2220404), enprostil, fluprostenol, iloprost, dinoprost, sulprostone, treprostinil, chembl357834, chembl1317823, chembl565591, chembl358653, sarcnu, and combinations thereof

[0082] Embodiment 29. A method of treating or suppressing the onset of Huntington's disease, the method comprising:

[0083] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0084] administering to the selected subject one or more modulators of a neuronal projection gene, the neuronal projection gene selected from the group consisting of: ADGRL3, ALCAM, BCAN, BCL11B, CACNA1A, CACNA1G, CALB1, CAMK2A, CHRNA4, CTTNBP2, DSCAM, GRIA2, GRIA4, GRID2, GRIK4, HCN2, KCND2, LGI3, LRRTM1, MAG, MBP, MYC, NCAM2, NCDN, NEFM, NEURL1, NFASC, NTM, PDE4B, PIK3R1, PTGER3, PTPRO, RAPGEF4, RGS8, ROBO2, SGK1, SIRT2, SLC17A8, SLC1A2, SLC8A3, SNAP91, SNPH, SYNDIG1 and UNC5A, or a protein encoded thereby, under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0085] Embodiment 30. The method of embodiment 29, wherein the one or more modulators are selected from the group consisting of adenosine phosphate, theophylline, theophylline, pentoxifylline, enprophylline, iloprost, papaverine, theobromine, aminopyrazone, [r]-methopram, roflumilast, pyramilast, rolipram, filminlast, chemb11230617, chemb1519827, cilomilast, (-)-rolipram, criborol, ibudilast, apremilast, chemb1521203, chemb174 078, propoxyphene, cdp840, sodium phenylbutyrate, chembl1232082, dipyridamole, theophylline sodium glycinate, flavonoid hydrochloride, aminophylline, resveratrol, caffeine, theophylline, amlexanox, edizalate, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, epezetrine, atagabalin, bepridil hydrochloride, amine octanoic acid, carboplatin, paclitaxel, chembl549906, chembl550795, sodium chloride, GSK-650394 , dalfampridine, guanidine hydrochloride, tedisamil, neripridine, L-glutamate, dihydrocainate, 2S,4R-4-methylglutamate, O-benzyl-L-serine, chembl1628669, mesalamine, fluorouracil, mibefradil dihydrochloride, trimethadione, cinnarizine, ethosuximide, zonisamide, cannabinoids, mibefradil, chembl1684954, flunarizine, methsuximide, phensuximide, methylethadione, polacrifos, nicotine, talbutal, butalbital, secobarbital, metharbital, sulfur Pentobarbital, primidone, mebendal, phenobarbital, varenicline, amobarbital, aprobarbital, butesa, enanthate, hexobarbital, barbital, pazaklan, cytisine, rivanicline, epibatidine, chembl1876219, chembl3103988, atracurium, chembl490153, hexamethonium, chembl407217, tc-2216, abt-560, isopyruvate, sofenacline, tc-6499, mibampator, (r,s)-AMPA, CHEMBL123132, aniracetam, CHEMBL136800, CHEMBL1255648, cyclothiazide, CHEMBL77862, CHEMBL334920, CHEMBL1097939, piracetam, CHEMBL320642, CHEMBL265301, GYKI-52466, NBQX, CHEMBL222418, tezampanel, (s)-AMPA, CHEMBL594840, CHEMBL121915, quisqualic acid, CHEMBL337577, CHEMBL27130, DNQX, CHEMBL333964, (s)-willardiine, CHEMBL28472, talampanel, perampanel, elampanel, CX1739, darampanel, beclamipanel, farampator, MK-8777, zonampanel, topiramate, pentobarbital, PF-04958242, serupipanel, cyclothiazide, CHEMBL334920, CHEMBL1097939, Joro spider toxin, domoic acid, dysherbaine, kainic acid, 2S,4R-4-Methylglutamate, chembl2313646, cyclosporine, interferon beta-1A, prednisone, quercetin, rutin, GSK-249320, cilostazol, zaltoprofen, chembl2052019, chembl395336, cyclic adenosine monophosphate, sodium dodecyl sulfate, bimatoprost, dinoprostone, misoprostol, beraprost, chembl1628262, carprocyclin, cicaprost, cloprostenol (chembl2220404), enprostil, fluprostenol, iloprost, dinoprost, sulprostone, treprostinil, chembl357834, chembl1317823, chembl565591, chembl358653, nadroparin calcium, 4'-hydroxytamoxifen, azacitidine, thioguanine, activin, adozelesin, amifostine, aminopterin, antibiotics, bizelesin, bromocriptine, bryostatin, calcitriol, diethylstilbestrol, elsamitrucin, estrone, folic acid, glutamine, hypoxanthine, imatinib, cilastatin, melatonin, methylprednisolone, N-methyl-N-nitrosourea, novobiocin, Chembl35482, phorbol 12-myristate 13-acetate, quinapril, vorinostat, sulindac, thrombin, thyrotropin, beta-nicotinamide adenine dinucleotide phosphate sodium salt, troglitazone,, Chembl100014, Chembl1213492, chorionic gonadotropin, perillyl alcohol, AMG-900, Alisertib, Dinaciclib, Roniciclib, temozolomide, prexasertib, PF-04691502, pquitinib, PA-799, isoproterenol, sf-1126, wortmannin, gsk-2636771, ds-7423, omipalisib, recilisib, pwt-33587, rg-7666, vs-5584, copanlisib, gedatolisib, sonidegib, apitolisib, taselisib, pillarsib (chembl3360203), votarisib, zstk-474, alpesib, pi-103, pillarsib (chembl3218575), wx-037, datolisib, bgt-226 (chembl3545096), pictilisib, buparlisib, panolisib, gsk-1059615, azd-6482, buparlisib hydrochloride, LY-3023414, and combinations thereof.,

[0086] Embodiment 31. A method for treating or suppressing the onset of Huntington's disease, the method comprising:

[0087] Select subjects having Huntington's disease or at risk of having Huntington's disease; and

[0088] Administer to the selected subjects one or more modulators of a TCF7L2 target gene, the TCF7L2 target gene consisting of the group consisting of: BMP4, CCND1, CCND2, DOCK10, DOCK9, DUSP15, ENPP4, EPAS1, EPHB1, ERBB3, EVI2A, EVI2B, FA2H, GJB1, HAPLN2, HSPA2, ID3, LGI3, MBP, MOG, MYC, MYRF, NFASC, NKAIN1, NKX6-2, OLIG2, PLEKHB1, PLP1, PPP1R16B, RAB33A, RASGEF1B, RTKN, SIRT2, SLC1A2, SOX10, ST18, TMEM125, TMEM2, TPPP, TSPAN15, UGT8, and AATK, or a protein encoded thereby, under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0089] Embodiment 32. The method according to embodiment 31, wherein the one or more modulators are selected from the group consisting of: 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, simvastatin, arsenic trioxide, paracetamol, vitamin E, cytarabine, gossypol, roniciclib, ribociclib, palbociclib, methotrexate, mycophenolic acid, nifedipine, tamoxifen, troglitazone, uracil, abemaciclib, briciclib, abemaciclib, decitabine, palbociclib, perifosine, cyclosporine, interferon beta-1a, prednisone, quercetin, rutin, vemurafenib, nadroparin calcium, 4'-hydroxytamoxifen, azacitidine, thioguanine, asivicin, adozelesin, amifostine, aminopterin, antibiotics, bizelesin, bromocriptine, bryostatin, calcitriol, diethylstilbestrol, elsamitrucin, estrone, folic acid, glutamine, hypoxanthine, imatinib, indomethacin, lithium, semustine, melatonin, methylprednisolone, N-methyl-N-nitrosourea, novobiocin, chembl35482, phorbol 12-myristate 13-acetate,, quinapril, vorinostat, sulindac, thrombin, thyrotropin, beta-nicotinamide adenine dinucleotide phosphate sodium, troglitazone, verapamil, chembl100014, chembl1213492, gonadotropin, chorionic gonadotropin, perillyl alcohol, amg-900, alisertib, dinaciclib, temozolomide, prexasertib, sodium dodecyl sulfate, L-glutamate, dihydrocaine, 2S,4R-4-methylglutamate, O-benzyl-L-serine, chembl1628669, mesalamine, perifosine, and combinations thereof.

[0090] Embodiment 33. A method of treating or inhibiting the onset of Huntington's disease, the method comprising:

[0091] selecting a subject having Huntington's disease or at risk of developing Huntington's disease; and

[0092] administering to the selected subject one or more modulators of a gene involved in the NKX2.2→OLIG2→SOX10→MYRF regulatory cascade or a protein encoded thereby under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0093] Embodiment 34. The method according to embodiment 33, wherein the gene is the NKX2.2 gene or a protein encoded thereby.

[0094] Embodiment 35. The method according to embodiment 33, wherein the gene is the OLIG2 gene or the protein encoded thereby.

[0095] Embodiment 36. The method according to embodiment 33, wherein the gene is the SOX10 gene or the protein encoded thereby.

[0096] Embodiment 37. The method according to embodiment 33, wherein the gene is the MYRF gene or the protein encoded thereby.

[0097] Embodiment 38. The method according to embodiment 33, wherein the one or more modulators are selected from the group consisting of vemurafenib.

[0098] Embodiment 39. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, wherein the administration is carried out using intracerebral delivery, intrathecal delivery, intranasal delivery or by direct infusion into the cerebral ventricle.

[0099] Embodiment 40. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, the method further comprising:

[0100] Administering a preparation of human glial progenitor cells to a selected subject.

[0101] Embodiment 41. The method according to embodiment 40, wherein the preparation of glial progenitor cells is astrocyte-biased glial progenitor cells.

[0102] Embodiment 42. The method according to embodiment 40, wherein the glial progenitor cells of the preparation are A2B5 + , CD140a + and / or CD44 + .

[0103] Embodiment 43. The method according to embodiment 40, wherein the preparation of glial progenitor cells is administered to the striatum, forebrain, brainstem and / or cerebellum of the subject.

[0104] Embodiment 44. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, wherein the subject is a human.

[0105] Embodiment 45. The method according to embodiment 40, wherein the glial progenitor cells are derived from fetal tissue.

[0106] Embodiment 46. The method according to embodiment 40, wherein the glial progenitor cells are derived from embryonic stem cells.

[0107] Embodiment 47. The method according to embodiment 40, wherein the glial progenitor cells are derived from induced pluripotent stem cells.

[0108] Embodiment 48. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, wherein Huntington's disease is treated.

[0109] Embodiment 49. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, wherein the onset of Huntington's disease is inhibited.

[0110] Embodiment 50. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, wherein the one or more modulators are agonists.

[0111] Embodiment 51. The method according to embodiment 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31 or 33, wherein the one or more modulators are antagonists. Brief Description of the Drawings

[0113] Figure 1A-1G Shows that HD hESC-derived hGPCs exhibit large mHTT-dependent gene expression changes. Figure 1A Shows a principal component analysis (PCA) based on the expression of rv26,000 transcripts. The expression data are shown as transcripts per million (TPM), with post-normalization to account for variance (Risso et al., “Normalization of RNA-seq Data Using Factor Analysis of Control Genes or Samples,” Nat. Biotechnol. 32:896-902 (2014), which is hereby incorporated by reference in its entirety). The PCA plot shows the unique transcriptomic range of expression signatures of HD-derived human glial progenitor cells (hGPCs). Figure 1BIt is a Venn diagram that shows the intersection (green, downregulated; red, upregulated; fold change [FC] > 2.0, FDR 1%) of a list of differentially expressed genes (DEGs) obtained by comparing hGPC from 3 different HD patients with pooled control hGPC from 2 donors. Then, the list of DEGs shared by the 3 HD patients was filtered by intersecting with those DEGs (FC > 2.0, FDR 1%) found in patient HD20 (from GENEA20) compared to normal sibling CTR19 (GENEA19); this filtering step further increased the specificity of the DEGs associated with mHTT. The intersections highlighted in gray together constitute the entire gene set that is differentially expressed in all HD lines relative to their pooled control. Figure 1C Shows an expression heatmap based on Figure 1B the TPM values of the 429 DEGs highlighted in Figure 1C , which shows the clustering of hGPC according to disease status. The dendrogram shows hierarchical clustering based on Euclidean distance, which was calculated from log2 - TPM values from three HD - ESC lines (HD - 17, HD - 18, and HD - 20) and two matched control lines (CTR19 and CTR02). Figure 1D Shows Figure 1B a network representation of the functional annotation of the 429 intersecting DEGs highlighted in Figure 1C (Gene Ontology: Biological Process and Cellular Component, Bonferroni - corrected p < 0.01). Genes are circular nodes, and the edge color represents their dysregulation direction (green, downregulated; red, upregulated). Rounded - rectangle nodes represent annotation terms. Nodes are sized by degree and colored by tightly interconnected modules (M1 - M3) identified by community detection. For each module, the top 3 annotations are listed by importance and enrichment fold. Selected gene nodes are marked, which include genes encoding key hGPC lineage transcription factors and stage - regulating proteins. Figure 1E Is an expression heatmap of the 63 conserved DEGs identified in M1 (( Figure 1D ) purple in Figure 1E ), which has annotations related to glial cell differentiation and myelination. Figure 1F Is an expression heatmap of the 56 conserved DEGs identified in M2 (( Figure 1D ) light purple in Figure 1F ), which has annotations related to axon guidance and axonogenesis. Figure 1G Is an expression heatmap of the 68 conserved DEGs identified in M3 ( Figure 1D ) yellow in Figure 1G ), which has annotations related to the regulation of synaptic structure and synaptic signaling. All differentially expressed (DE) results are 1% FDR and FC > 2; Gene Ontology (GO) annotation results are Bonferroni - corrected to p < 0.01.

[0114] Figure 2A - Figure 2D Genes differentially expressed between hGPCs derived from different HD hESCs and pooled controls are shown. Figure 2A - Figure 2B Genes differentially expressed ( Figure 2A , upregulated genes; Figure 2B , downregulated genes) obtained by comparing each CD140a-sorted HD-derived GPC line (HD17, HD18, and HD20) with pooled control-derived GPCs are shown. Differentially expressed genes in HD GPCs were significant at 1% FDR and FC > 2.00. Figure 2C - Figure 2D Comparison of CD44-sorted HD-derived APC lines (HD17, HD18, and HD20) with control-derived APCs is shown ( Figure 2C , upregulated; Figure 2D , downregulated). Differentially expressed genes in HD APCs were significant at 5% FDR. In both cases, 20 vs 19 represents the comparison of HD line HD20 (Genea20) with its sibling control line CTR19 (Genea19). The horizontal bars represent the overall size of the gene sets, and the vertical bars represent the size of the gene set intersections. The vertical bars are sorted first by the number of gene sets in the intersection and then by the size of the intersection. These points correspond to the gene sets that make up each intersection.

[0115] Figure 3A - Figure 3B Shown Functional annotation revealed impairments in the transcription of genes associated with gliogenesis, myelination, and synaptic transmission associated with HD. Gene Ontology (GO) functional annotation was performed on 429 differentially expressed genes (DEGs) in 3 mHTT hGPC lines relative to pooled control hGPCs (see Figure 1B - 1C ). Fifty significantly associated GO annotation terms (biological processes and cellular components, Bonferroni-corrected p < 0.01) were identified using the ToppCluster annotation tool (Kaimal et al., “ToppCluster: a Multiple Gene List Feature Analyzer for Comparative Enrichment Clustering and Network-based Dissection of Biological Systems,” Nucleic Acids Res. 38:W96-W102 (2010), which is hereby incorporated by reference in its entirety). Through network analysis, these GO terms and their associated DEGs were grouped together into three functionally related modules (M1 to M3, see Figure 1D)。For each GO term, given the number of DEGs with annotation and the total number of human protein-coding genes found in the term, the expected value is assumed to be a constant ratio. The enrichment fold is the ratio of the observed number of DEGs found in the term to the expected number. In each functional module, GO terms are first sorted by p-value and then by enrichment fold. Three GO terms, GO:0007268 (chemical synaptic transmission), GO:0098916 (anterograde trans-synaptic signaling), and GO:0099537 (trans-synaptic signaling), ranked 3rd to 5th respectively in the M3 module. They contain a set of 37 associated DEGs, which are included within the 38 DEGs associated with GO:0099536 (synaptic signaling) that ranked second in M3. To reduce redundancy, these three GO terms are omitted from the figure. Figure 3A is a bar graph showing the top 5 GO terms for each functional module. Figure 3B is a table that lists the calculated values and associated DEGs for each top-ranked term. The associated DEGs are colored according to their direction of dysregulation in the hGPCs from the HD source compared to the control source (green, downregulated; red, upregulated).

[0116] Figure 4A - Figure 4C Shows that increasing CAG length is associated with decreased oligodendrocyte gene expression. Figure 4A Shows an expression heatmap based on TPM values, which were calculated from the raw counts of 429 DEGs (1% FDR, FC>2.0) found in the intersection of DEGs by comparing hGPCs from each of three different HD patients to pooled control hGPCs from two different donors. The color on the side of the row shows the Pearson's correlation coefficient (R) between the fold change of each of the 429 DEGs shown in the heatmap for the corresponding hGPC line and the CAG repeat number in that HD line (HD17 = 40x CAG, HD18 = 46xCAG, HD20 = 48x CAG). Selected genes encoding transcription factors and stage-regulating proteins involved in glial differentiation and myelination are listed. Figure 4B Shows a combined scatter plot with a linear fit line that was obtained by Figure 4A regressing the fold change of each of the 429 DEGs shown in the heatmap against the CAG repeat number in the corresponding hGPC line. Figure 4CThis is a histogram showing the distribution of Pearson's correlation coefficient (R) for the correlation between the fold change (FC) of DEGs in 3 HD lines and the corresponding CAG lengths. For 255 out of 429 genes (|Pearson R| > 0.75), correlation analysis showed that the absolute magnitude of FC increased with increasing CAG repeat number; 228 of these genes showed a negative correlation between gene expression level and CAG repeat number, with longer repeats associated with decreased glial gene expression.

[0117] Figure 5A - Figure 5D Show that human and mouse glia exhibit overlap in genes dysregulated as a function of CAG repeat length. There is a high degree of overlap between these hGPC genes and ontologies that were found to be progressively dysregulated with increasing CAG repeat length in hGPCs and were noted to be dysregulated with CAG repeat length in mouse brain tissue (Langfelder et al., “Integrated Genomics and Proteomics Define Huntingtin CAG Length-Dependent Networks in Mice,” Nat. Neurosci. 19:623-633 (2016), which is hereby incorporated by reference in its entirety). Figure 5A Shows a representative list obtained by comparing the differentially expressed genes (DEGs) obtained from CD140-sorted GPCs from HD sources and CD44-sorted APCs from HD sources, with the mouse mHtt allele series ( Figure 5A and Figure 5B ) and the differentially expressed results of 6-month Q175 profiled tissues ( Figure 5C and Figure 5D ) from (Langfelder et al., “Integrated Genomics and Proteomics Define Huntingtin CAG Length-Dependent Networks in Mice,” Nat. Neurosci. 19:623-633 (2016), which is hereby incorporated by reference in its entirety herein). Figure 5A and Figure 5CThe network diagram in [document] shows the significant pairwise set intersections (Fisher's exact test, p < 0.05) between the DEG sets (yellow nodes) from CD140 and CD44 HD Genea and the DEG sets (analyzed (gray nodes)) from Langfelder et al., "Integrated Genomics and Proteomics Define Huntingtin CAG Length-Dependent Networks in Mice," Nat. Neurosci. 19: 623-633 (2016) (which is hereby incorporated by reference in its entirety). The size of the nodes is determined by the total number of DEGs indicated in parentheses for each node. The number of DEGs in the HD Genea set was obtained after ID conversion to mouse orthologous genes. The edge thickness represents the significance of the gene set intersection, calculated as -log10(Fisher's exact test p-value). The edge color and label show the number of genes in the pairwise set intersection. Only the DEG sets from Langfelder et al., "Integrated Genomics and Proteomics Define Huntingtin CAG Length-Dependent Networks in Mice," Nat. Neurosci. 19: 623-633 (2016) (which is hereby incorporated by reference in its entirety) that have significant overlap with either of the two HD Genea sets are shown. Figure 5B and Figure 5D The dot plots in [document] show, respectively, for Figure 5A and Figure 5CComparison of Gene Ontology (GO): Biological Process annotations of the DEG sets in []. The size of the dots is determined according to the proportion of the genes relative to the DEG sets. The color of the dots represents the significance of the associated GO terms. All DEG sets with significant annotations (BH-corrected p < 0.01) are shown. The most significant intersections were observed between the CD140 DEG set and the DEGs in the 6-month striatum Q175 samples (p = 1.10E-06; 150 genes) among the allelic series DEGs, and between the CD140 DEG set and the 6-month Q175 cerebellum DEGs of the Q175 tissue (p = 9.86E-13; 85 genes). These intersections included the glial regulators Nkx2-2, Olig1, and Olig2, as well as genes encoding proteins involved in myelination, ion channel activity, and synaptic transmission. Overall, many similar significant annotations were observed for the HDGenea CD140 DEGs and brain-derived DEGs from Langfelder et al., “Integrated Genomics and Proteomics Define Huntingtin CAG Length-Dependent Networks in Mice,” Nat. Neurosci. 19:623-633 (2016) (which is hereby incorporated by reference in its entirety) involved in functions including gliogenesis, myelination, axon development, and ion channel activity.

[0118] Figure 6A - Figure 6BGenes associated with glial cell differentiation were shown to be dysregulated in mHTT-expressing GPCs. Expression of selected genes dysregulated in HD-derived GPCs identified by RNA-seq analysis was evaluated by TaqMan low density array (TLDA) RT-qPCR and compared to control GPCs. Expression data were normalized against 18S and GAPDH endogenous controls. Mean ddCt values and ranges of standard errors are shown, which were calculated by comparing 3 pooled HD GPC lines (n = 3 for GENEA17 and GENEA20 lines, n = 5 for GENEA18, total n = 11) to 2 pooled control GPC lines (n = 6 for GENEA02, n = 3 for GENEA19 line, total n = 9). Differences in expression between HD and control GPCs were evaluated by paired t-test followed by Benjamini-Hochberg (BH) multiple test correction (***p < 0.01, **p < 0.05, *p < 0.1). Genes assayed on both arrays are highlighted in bold. Analysis of TLDA data was performed in ExpressionSuite software 1.1 (Applied Biosciences). Most genes identified by RNA-seq as dysregulated in HD-derived GPCs were confirmed as such by TLDA. Figure 6A Genes encoding key GPC lineage transcription factors and myelin-related proteins regulating stages are shown. Forty-four genes are shown, excluding MOBP and MOG, which were found to have a high proportion of unreliable reactions. Figure 6B Transcriptional targets of TCF7L2 predicted by upstream regulator analysis in IPA are shown. A total of 42 genes are shown, excluding genes with a high proportion of unreliable reactions.

[0119] Figure 7 HD-derived hGPCs were shown to exhibit significant dysregulation of potassium channel genes. Differential gene expression comparison of each HD-derived hGPC line to pooled control hGPCs (FDR of 5%, no fold change threshold) revealed that in 3 HD-derived lines, 25 potassium channel genes were dysregulated in at least 2 of the lines. NS = not significant.

[0120] Figures 8A - 8N show impaired myelination in mice chimeric for human GPC expressing mHTT. Human glial chimeric mice were established by neonatal injection of hGPC into quaking x rag2 hosts, and the mice were sacrificed at 8, 13, and 18 weeks. Figures 8A and 8D show that by 8 weeks post - neonatal implantation, myelin basic protein (MBP) expression of control hGPC (GENEA19) was evident (Figure 8A), while mice implanted with HD - derived hGPC expressing mHTT (GENEA20) showed little or no MBP immunolabeling at this point (Figure 8D). Figures 8B and 8E show that by 13 weeks, mice implanted with control hGPC showed robust myelination (Figure 8B), while only scattered islands of MBP expression were found in recipients of matched HD - derived GPC (Figure 8E). Figures 8C and 8F show that relative to mHTT GPC - chimeric mice (Figure 8F), myelin formation from control GPC became increasingly robust by 18 weeks (Figure 8C). Figure 8G - Figure 8I showed that the density of implanted human GPCs did not differ between the control group and mHTT hGPCs at any time point evaluated ( Figure 8G ), but in hGPCs expressing mHTT, the fraction of hGPCs that differentiated into transferrin (TF)+ oligodendrocytes was significantly lower ( Figure 8H ), resulting in fewer TF - defined oligodendrocytes in chimeras implanted with MHTT hGPCs ( Figure 8I ). Figure 8J –8L showed that the proportion of myelinating donor - derived oligodendrocytes, as defined by co - expression of human TF and MBP of MBP, was significantly lower in chimeric brains implanted with mHTT - hGPCs compared to chimeric brains implanted with control hGPCs ( Figure 8J ). Similarly, the proportion of MBP expression among all donor cells was significantly higher in mice implanted with control hGPCs compared to mice implanted with HD - derived hGPCs ( Figure 8K ). Thus, the myelin brightness, as evaluated on MBP immunostained sections, was significantly higher in the corpus callosum implanted with control compared to the white matter implanted with mHTT GPCs ( Figure 8L ). Figure 8M and Figure 8N showed that neither the density of implanted human GPCs ( Figure 8G ) nor the distribution ( Figure 8M and Figure 8N, dot plots) showed no significant differences between control and HD-derived hGPCs, indicating that the myelination defects in the brains implanted with mHTT hGPCs were due to impaired oligodendrocyte differentiation and myelination, rather than differential implantation. Scale bar, 50 mm. Values are represented as mean ± SEM according to two-way ANOVA and Bonferroni post hoc test. **p < 0.01 and ***p < 0.001.

[0121] Figures 9A - 9H show that the brains implanted with mHTT GPCs exhibited reduced and delayed axonal myelination. Figures 9A - 9F are confocal images of the tremor corpus callosum implanted with hGPCs, which show higher MBP expression and a higher proportion of ensheathed axons in mice implanted with GENEA19 control hGPCs (Figures 9A–9C) compared to mice implanted with hGPCs expressing mHTT from the GENEA20 source (Figures 9D–9F). Figures 9D’ and 9E’ show confocal z-stacks with orthogonal views of donor-derived MBP+ oligodendrocytes.

[0122] Figure 9F’ shows a higher magnification of Figure 9F, which shows MBP immunoreactivity around the ensheathed axons. Figures 9G and 9H show the proportion of total NF+ host axons myelinated by MBP (Figure 9G), and the proportion of NF+ host axons myelinated by MBP per MBP+ donor-derived oligodendrocyte (Figure 9H). Scale bars represent 20 mm (Figures 9A–9F) and 5 mm (Figures 9A’–9C’). Values are represented as mean ± SEM according to two-way ANOVA and Bonferroni post hoc test. **p < 0.01 and ***p < 0.001.

[0123] Figure 10 Shown is that SOX10-MYRF transduction restored myelination gene expression in mHTT GPCs. The figure shows a graphical representation of the qPCR data outlined in Table 1 below.

[0124] Table 1. SOX10-MYRF transduction restores myelination gene expression in mHTT GPCs

[0125]

[0126] These qPCR data show the ddCT values of selected oligoneogenic and myelination genes in normal hGPCs and hGPCs expressing mHTT after transfection with a bicistronic plasmid expressing SOX10 and MYRF, normalized to 18S and then to cells transfected with a control plasmid, which reflect the relative mRNA levels. Welch’s t-test. Primers located on the coding sequence; Primers located in the 3’UTR. Endo: Endogenous gene; Virus: Viral transgenic product. *p<0.05.

[0127] Following transfection with a bicistronic plasmid expressing SOX10 and MYRF, the expression values of selected oligogenic and myelination genes, normalized to cells transfected with 18S and a control plasmid, in normal hGPC (Genea19, black bars) and hGPC expressing mHTT (Genea 20, red). Welch's t-test comparisons: 1) SOX10-MYRF-transfected vs EGFP-transfected, performed independently for each line, significance indicated by asterisks; or 2) SOX10-MYRF-transfected Genea 20 vs EGFP-control-transfected Genea19 (significance indicated by hash marks). * / #p<0.05. ** / ##p<0.01.; *** / p<0.001.; **** / #p<0.0001. Primers located on the coding sequence; Primers located in the 3’UTR.

[0128] Figure 11A - Figure 11M shows that SOX10 and MYRF rescued oligodendrocyte differentiation and myelination via mHTT GPC. Figure 11A Shows the doxycycline-regulated dual-vector lentiviral (LV) transduction strategy that allows doxycycline (DOX) to trigger the interdependent overexpression of SOX10 and MYRF and the simultaneous expression of CD4 to permit FACS-based immunoisolation of SOX10-MYRF-transduced hGPC. Figure 11B - Figure 11D Illustrates the effect of SOX10 and MYRF overexpression in hGPC expressing mHTT, which was evaluated by transducing a matched pool of 180DIV GENEA20-derived hGPC with a DOX-regulated lentiviral SOX10 / MYRF and exposing some cultures to DOX while leaving matched control cultures untreated. After an additional week in vitro, cells were immunostained with mAb O4, which recognizes oligodendrocytic sulfatide. In the absence of DOX, mHTT hGPC were maintained stably and did not express detectable O4 ( Figure 11B ). In contrast, those mHTT hGPC fed DOX ( Figure 11C ) had upregulated SOX10 and MYRF expression and showed a sharp and significant increase in oligodendrocyte differentiation ( Figure 11D)。This schematic outlines the experimental design used to assess the in vivo myelination capacity of HD-derived hGPCs in the presence and absence of SOX10 and MYRF expression rescue. All cells were transiently exposed to DOX in vitro to initiate CD4 expression and allow FACS sorting prior to transplantation into neonatal immunodeficient shiverer mice. At 9 weeks of age, DOX was administered to implanted mice for an additional 4 weeks to initiate SOX10 and MYRF expression (+DOX), or not (-DOX, control). Neonatally implanted shiverer mice with hGPCs (GENEA19) derived from normal HTT-expressing hESCs produced abundant MBP expression and oligodendrocyte morphology in vivo by 13 weeks. In contrast, mice implanted with mHTT-expressing hGPCs (GENEA20[G20]) generated from HD hESCs produced little detectable MBP at this time point. Figures 11H and 11I show that at 9 weeks of age, DOX was orally administered to some mice implanted with GENEA20 mHTT hGPCs to trigger SOX10 and MYRF expression (Figure 11H), while dox was not administered to matched controls (Figure 11I). DOX(+) mice showed a significant number of MBP+ myelinated oligodendrocytes in the implanted white matter (Figure 11H). Figures 11J and 11K show that by the same time point, no donor cells in DOX(-) control mice produced MBP expression (Figure 11J), despite similar donor cell implantation (Figure 11K). Figures 11L and 11M show that in DOX(+) mice implanted with SOX10 / MYRF-transduced GENEA20 hGPCs, donor-derived oligodendrocytes induced robust formation of nodes of Ranvier (Figure 11L), demonstrated by clustering of βIV-spectrin flanked by Caspr protein characteristic of the nodal architecture, which was not present in untreated shiverer brains. Scale bars represent 50 mm ( Figure 11B , 11C and 11F–11I), 1 mm (Figure 11L), and 0.5 mm (Figure 11M). Values represent mean ± SEM. ***p < 0.001 (t test).

[0129] Figure 12A- Figure 12PDelayed astrocyte differentiation was shown in mHTT GPCs. Figures 12A - 12C show significantly delayed astrocyte differentiation in mHTT glial chimeras. Neonatal mice transplanted with hGPCs from normal HTT GENEA19 began to develop a large number of donor-derived GFAP+ astrocytes by 8 weeks (Figure 12A), began to robustly develop donor-derived GFAP+ astrocytes by 13 weeks (Figure 12B), and had dense astrocyte colonization in the corpus callosum white matter by 18 weeks (Figure 12C). Figures 12D - 12F show that, in contrast, hGPCs expressing mHTT derived from GENEA20 sibling hESCs developed an astrocyte phenotype more slowly at 8 weeks (Figure 12D) and 13 weeks (Figure 12E), with barely detectable GFAP expression, and only moderately matured GFAP+ astrocytes at 18 weeks (Figure 12F). Figures 12G and 12H show the differences in the mature astrocyte morphology between control (Figure 12G) and mHTT-expressing (Figure 12H) astrocytes, where mHTT astrocytes generally did not exhibit the degree of radial symmetry of their control-derived counterparts. Figure 12I The proportion of GFAP-expressing cells among all donor cells was consistently lower in mice implanted with mHTT hGPCs compared to mice implanted with controls. Figure 12J - Figure 12M Sholl analysis of cells traced in 3D mode in NeuroLucida, which were shown to be flat in Figure 12O and Figure 12P revealed that normal donor astrocytes had greater fiber complexity ( Figure 12J ) and more primary processes ( Figure 12K ) compared to astroglia expressing mHTT, but had shorter average fiber length and maximum fiber length ( Figure 12L and Figure 12M ). Figure 12N – Figure 12P Fan-in radial analysis of volume occupancy rate (Dang et al., “Formoterol, a Long-Acting β2 Adrenergic Agonist, Improves Cogntive Function and Promotes Dendritic Complexity in a Mouse Model of Down Syndrome,” Biol. Psychiatry 75:179 - 188 (2014), which is hereby incorporated by reference in its entirety) was shown, which revealed that mHTT astrocytes had significantly more regions not occupied by glial processes compared to control astrocytes ( Figure 12N ). Figure 12O andFigure 12P The illustrations therein indicate their discontinuous domain structures. Values represent mean ± SEM. *p < 0.05; **p < 0.01; ***p < 0.001, using two-way ANOVA and Bonferroni post hoc test ( Figure 12I ), comparison of non-linear regression (p < 0.0001)( Figure 12J ), and unpaired t-test comparing the mean per mouse among all scored cells ( Figure 12K - Figure 12N )(n = 4 controls, 7 mHTT mice). Scale bars represent 25 mm (Figure 12A–Figure 12F) and 10 mm (Figure 12G, Figure 12H, Figure 12O and Figure 12P ).

[0130] Figure 13A - Figure 13E It is shown that astrocytes expressing mHTT exhibit reduced complexity and an incomplete domain structure. Figure 13A Sholl analysis of human cells immunostained for GFAP in human glial chimeras 18 weeks after neonatal implantation is shown. Non-linear regression curves (Lorentzian curve fitting) of the radial intersections of each cell line as a function of branch order are shown. Comparison of control group (N = 7) versus mHTT mice (N = 10); p < 0.0001. Figure 13B It is shown that both the normal HTT control line GENEA19 and the unrelated normal HTT hiPS cell line C27 have more primary processes than the GENEA lines expressing mHTT (GENEA18 and GENEA20). Control GENEA19 and C27 are not different from each other, but both GENEA18 and GENEA20 are significantly different from the controls (one-way ANOVA and Dunnett's post hoc t-test; p < 0.0001). Figure 13C It is shown that the fiber distribution of astrocytes derived from the two control lines C27 and GENEA19 is more radially symmetric than that of either of the two mHTT lines. Using one-way ANOVA and Dunnett's post hoc test, with C27 as the control, p < 0.0001. Both GENEA18 and GENEA20 are significantly different from C27, p < 0.0001. Figure 13A - 13C , control: C27, gray; and GENEA 19, black. HD-derived: GENEA 18, orange; GENEA 20, red. Figure 13D A flattened three-dimensional coronal tracing of astrocytes in the corpus callosum of a mouse transplanted with hGPCs derived from GENEA 18 is shown ( Figure 13E ), compared to a flattened three-dimensional coronal tracing of astrocytes in the corpus callosum of a mouse transplanted with control hGPCs derived from C27. Scale bar: Figure 13D , 25 μm.

[0131] Figure 14A - Figure 14H The display of CD44+ astrocytes derived from HD hESCs shows mHTT-dependent gene expression changes. Figure 14A As shown Figure 1A PCA performed as in Figure 14B but using CD44-sorted astrocytes and their precursors verified the distinct expression signatures of HD-derived cells and normal cells. Figure 14C A Venn diagram highlighting the intersection of the DEG lists (green, downregulated; red, upregulated; FDR 5%) is shown, which was obtained by comparing astrocytes from 3 HD patients with pooled control cells and using the same cell lines and analysis pipeline as in Figure 1. The list of DEGs shared by the 3 HD patients was filtered using those genes that were differentially expressed in patient HD20 (GENEA20) relative to its sibling donor CTR19 (GENEA19). Figure 14B A heatmap of log2-transformed TPM values calculated based on the raw counts of the 114 DEGs highlighted in ( Figure 14D ) is shown, which depicts clustering according to disease status. Figure 14B ) is shown, which depicts clustering according to disease status. Figure 14E A heatmap of the expression of the 14 conserved DEGs identified in M1 ( Figure 14D yellow in Figure 14F ) with annotations related to postsynaptic and receptor complex components is shown. Figure 14D A heatmap of the 9 conserved DEGs identified in M2 and annotated for perinuclear and early endosome components ( Figure 14G gray in Figure 14D ) is shown. Figure 14H A heatmap of the 11 conserved DEGs identified in M3 ( Figure 14D ) is shown, which has annotations related to plasma membrane, cell-cell junction, and desmosome components. Detailed Description

[0132] The disclosure herein generally relates to methods of treating or suppressing the onset of Huntington's disease. Such methods involve selecting a subject having or at risk of having Huntington's disease and administering to the subject one or more modulators of one or more genes or proteins encoded thereby as described in Table 2 or Table 3 under conditions effective to treat or suppress the onset of Huntington's disease in the subject.

[0133]

[0134]

[0135]

[0136]

[0137]

[0138]

[0139]

[0140]

[0141]

[0142]

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[0144]

[0145] Huntington's disease is an autosomal dominant neurodegenerative disorder characterized by persistent progressive motor disturbances accompanied by devastating mental and cognitive deterioration. Huntington's disease is associated with a persistent severe atrophy of the neostriatum, which is related to a significant loss of GABAergic medium spiny projection neurons, the principal output neurons of the striatum. Huntington's disease is characterized by an abnormally long CAG repeat expansion in the first exon of the huntingtin gene ("HTT"). Encoded polyglutamine expansions in the mutant huntingtin disrupt its normal function and protein-protein interactions, ultimately resulting in widespread neuropathological changes, most prominent in the neostriatum.

[0146] As used herein, the term "glial cell" refers to a population of non-neuronal cells that provide support and nutrition, maintain homeostasis, form myelin or promote myelin formation, and participate in signal transmission in the nervous system. "Glial cells" as used herein include fully differentiated cells in the glial lineage, such as oligodendrocytes or astrocytes, as well as glial progenitor cells. Glial progenitor cells are cells that have the potential to differentiate into cells in the glial lineage, such as oligodendrocytes and astrocytes.

[0147] As used herein, "treating" or "treatment" refers to any indication of success in ameliorating an injury, pathological change or disorder, including any objective or subjective parameter, such as alleviation; remission; reduction of symptoms or making the injury, pathological change or disorder more tolerable to the patient; slowing the rate of degeneration or decline; making the end point of degeneration less debilitating; or improving the physical or mental health of the subject. Treatment or amelioration of symptoms can be based on objective or subjective parameters; including the results of physical examination, neurological examination and / or psychiatric evaluation. "Treatment" includes administering glial progenitor cells to prevent or delay, to slow down, or to arrest or inhibit the development of symptoms or conditions associated with a disease, disorder or condition. "Treatment effect" refers to alleviating, eliminating or preventing a disease, disease symptoms or side effects of a disease, disorder or condition in a subject. Treatment can be prophylactic (to prevent or delay the onset or progression of a disease, disorder or condition, or to prevent the manifestation of its clinical or subclinical symptoms), or therapeutic suppression or slowing of symptoms after the manifestation of the disease, disorder or condition.

[0148] Suitable subjects for treatment according to the methods described herein include any mammalian subject having or at risk of having Huntington's disease. Exemplary mammalian subjects include humans, mice, rats, guinea pigs and other small rodents, dogs, cats, sheep, goats and monkeys. In one embodiment, the subject is a human.

[0149] One or more modulators for use in the methods described herein can be, but are not limited to, peptides, nucleic acid molecules or small molecule compounds. The modulators can be, for example, naturally occurring, semi-synthetic or synthetic agents. For example, a modulator can be a drug that targets a specific function of one or more genes. In certain embodiments, the one or more modulators can be antagonists or agonists.

[0150] The modulators of the present invention can be administered orally, parenterally, for example subcutaneously, intravenously, intramuscularly, intraperitoneally, by intranasal instillation, or by application to mucous membranes, such as the mucous membranes of the nose, larynx and bronchi. They can be administered alone or in combination with a suitable pharmaceutical carrier, and can be in solid or liquid form, such as tablets, capsules, powders, solutions, suspensions or emulsions.

[0151] The regulators of the present invention can be administered orally, for example, with an inert diluent or with an assimilable edible carrier, or they can be encapsulated in hard or soft shell capsules, or they can be compressed into tablets, or they can be directly incorporated into the diet. For oral therapeutic administration, these regulators can be combined with excipients and used in the form of tablets, capsules, elixirs, suspensions, syrups, etc. Such compositions and articles should contain at least 0.1% of the active compound. The percentage of the compound in these compositions can of course vary and can conveniently be between about 2% and about 60% by weight of the unit. The amount of the active compound in such therapeutically useful compositions is such that a suitable dosage can be obtained. Preferred compositions according to the present invention are prepared such that the oral dosage unit contains between about 1 mg and 250 mg of the active compound.

[0152] Tablets, capsules, etc. may also contain binders such as gum tragacanth, gum arabic, corn starch or gelatin; excipients such as dibasic calcium phosphate; disintegrants such as corn starch, potato starch, alginic acid; lubricants such as magnesium stearate; and sweeteners such as sucrose, lactose or saccharin. When the dosage unit form is a capsule, it may also contain a liquid carrier such as a fatty oil in addition to the materials of the above types.

[0153] Various other materials may be present as coatings or for modifying the physical form of the dosage unit. For example, tablets may be coated with shellac, sugar or both. Syrups may contain, in addition to the active ingredient, sucrose as a sweetener, methyl p-hydroxybenzoate and propyl p-hydroxybenzoate as preservatives, dyes and flavoring agents such as cherry or orange flavor.

[0154] These regulators can also be administered parenterally. Solutions or suspensions of these regulators can also be prepared in water, suitably mixed with a surfactant such as hydroxypropylcellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols and mixtures thereof in oils. Exemplary oils are oils of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil or mineral oil. Generally, water, saline, dextrose aqueous solutions and related sugar solutions, and diols such as propylene glycol or polyethylene glycol are preferred liquid carriers, especially for injectable solutions. Under conventional storage and use conditions, these articles contain preservatives to prevent the growth of microorganisms.

[0155] Pharmaceutical forms suitable for injection include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the form must be sterile and must be fluid to the extent that easy injection is possible. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyols (such as glycerol, propylene glycol and liquid polyethylene glycol), suitable mixtures thereof and vegetable oils.

[0156] The modulators of the present invention can also be administered directly to the airways in the form of an aerosol. For use as an aerosol, the compounds of the present invention in solution or suspension form can be packaged in a pressurized aerosol container together with a suitable propellant, such as a hydrocarbon propellant with conventional adjuvants, such as propane, butane or isobutane. The materials of the present invention can also be administered in a non-pressurized form, such as in a nebulizer or atomizer.

[0157] If modulation is to be achieved at the DNA level, this can be accomplished by using gene therapy to knock out or disrupt the target gene. As used herein, "knockout" can be gene knockdown or the gene can be knocked out by techniques known in the art including, but not limited to, retroviral gene transfer, by mutations such as point mutations, insertions, deletions, frameshifts or missense mutations.

[0158] In one embodiment, the one or more modulators can repress the expression of one or more genes described herein by zinc finger nucleases. Zinc finger nucleases (ZFNs) are artificial restriction enzymes generated by fusing a zinc finger DNA-binding domain to a DNA cleavage domain. The zinc finger domain can be engineered to target a desired DNA sequence, which enables zinc finger nucleases to target unique sequences in complex genomes (Urnov et al., "Genome Editing with Engineered Zinc Finger Nucleases," Nat. Rev. Genet. 11:636-646 (2010), which is hereby incorporated by reference in its entirety). By exploiting the endogenous DNA repair mechanism, these reagents can be used to precisely alter the genomes of higher organisms.

[0159] The one or more regulators can also be meganucleases and TAL effector nucleases (TALENs, Cellectis Bioresearch) (Joung & Sander, “TALENs: A Widely Applicable Technology for Targeted Genome Editing,” Nat. Rev. Mol. Cell Biol. 14:49 - 55 (2013), which is hereby incorporated by reference in its entirety). Composed of a TALE DNA - binding domain for sequence - specific recognition fused to the catalytic domain of an endonuclease that introduces double - strand breaks (DSBs). The DNA - binding domain is capable of targeting large recognition sites (e.g., 17 bp) with high precision. Meganucleases are sequence - specific endonucleases, naturally occurring “DNA scissors,” derived from various single - cell organisms such as bacteria, yeast, algae, and some plant organelles. Meganucleases have long recognition sites of between 12 and 30 base pairs. The recognition sites of meganucleases can be modified to target native genomic DNA sequences (such as endogenous genes).

[0160] In another embodiment, the one or more regulators are CRISPR - Cas9 - guided nucleases (Wiedenheft et al., “RNA - Guided Genetic Silencing Systems in Bacteria and Archaea,” Nature 482:331 - 338 (2012); Zhang et al., “Multiplex Genome Engineering Using CRISPR / Cas Systems,” Science 339(6121):819 - 23 (2013); and Gaj et al., “ZFN, TALEN, and CRISPR / Cas - based Methods for Genome Engineering,” Cell 31(7):397 - 405 (2013), which are hereby incorporated by reference in their entirety). Like TALEN and ZFN, CRISPR - Cas9 interference is a genetic technology that allows sequence - specific control of gene expression in prokaryotic and eukaryotic cells through double - strand DNA cleavage by a guided nuclease. It is based on the CRISPR (clustered regularly interspaced short palindromic repeats) pathway derived from the bacterial immune system.

[0161] The regulation of one or more genes described herein can also be effected using antisense oligonucleotides (ASOs). Suitable therapeutic ASOs for inhibiting one or more genes described herein include, but are not limited to, antisense RNAs, DNAs, RNA / DNA hybrids (e.g., gapmers), and their chemical analogs such as morpholinos, peptide nucleic acid oligomers, ASOs composed of locked nucleic acids. All antisense oligomers other than RNA oligomers, PNAs, and morpholinos act in eukaryotic cells via an RNase H-mediated target cleavage mechanism. PNAs and morpholinos bind complementary DNA and RNA targets with high affinity and specificity and thus act by simple steric blockage of the RNA translation machinery and appear to be completely resistant to nuclease attack.

[0162] "Antisense oligomer" refers to an antisense molecule or antigene agent that comprises an oligomer of at least about 10 nucleotides in length. In embodiments, the antisense oligomer comprises at least 15, 18, 20, 25, 30, 35, 40, or 50 nucleotides. The antisense approach involves the design of oligonucleotides (DNA, RNA, DNA / RNA, or chemically modified derivatives thereof) that are complementary to the RNA encoded by the polynucleotide sequence of the genes identified herein. Antisense RNAs can be introduced into cells to inhibit the translation or activity of a complementary mRNA by base pairing with the complementary mRNA and physically blocking either the translation of the complementary mRNA or the activity of the complementary mRNA. Thus, this effect is stoichiometric. Absolute complementarity, although preferred, is not required. A sequence "complementary" to a portion of RNA as referred to herein means a sequence having sufficient complementarity to be able to hybridize with the RNA, forming a stable duplex. In the case of a double-stranded antisense polynucleotide sequence, thus, one strand of the double-stranded DNA can be tested, or the formation of a triple helix can be assayed. The ability to hybridize will depend on the degree of complementarity and the length of the antisense polynucleotide sequence. Generally, the longer the hybridizing polynucleotide sequence, the more base mismatches it may have with the RNA it may contain and still form a stable duplex (or triple helix, as the case may be). Those skilled in the art can ascertain the tolerated degree of mismatch by using standard procedures to determine the melting point of the hybrid complex.

[0163] In one embodiment, the one or more modulators are antisense oligonucleotides that specifically bind to one or more of the genes described herein and inhibit the functional expression of one or more of the genes described herein. For example, common modifications of ASOs for increasing duplex stability include incorporation of 5-methyl-dC, 2-amino-dA, locked nucleic acids, and / or peptide nucleic acid bases. Common modifications for enhancing nuclease resistance include conversion of normal phosphodiester linkages to phosphorothioate or dithiophosphate linkages, or use of propargyl analogue bases, 2'-O-methyl or 2'-O-methoxyethyl RNA bases.

[0164] RNA interference (RNAi) using small interfering RNA (siRNA) is another form of post-transcriptional gene silencing that can be used to modulate one or more genes in a subject as described herein.

[0165] Accordingly, in one embodiment, the one or more modulators are siRNAs. An siRNA is a double-stranded synthetic RNA molecule approximately 20-25 nucleotides in length with short 2-3 nucleotide 3' overhangs at both ends. The double-stranded siRNA molecule represents the sense and antisense strands of a portion of the target mRNA molecule. siRNA molecules are typically designed to target a region of the mRNA target approximately 50-100 nucleotides downstream of the start codon. The siRNAs of the present invention can comprise partially purified RNA, substantially pure RNA, synthetic RNA, or recombinantly produced RNA, as well as altered RNAs that differ from naturally occurring RNAs by addition, deletion, substitution, and / or alteration of one or more nucleotides. Such alterations can include addition of non-nucleotide moieties to, for example, one or more termini of the siRNA or to one or more internal nucleotides of the siRNA, including modifications that render the siRNA resistant to nuclease digestion. After introduction into cells, the siRNA complex triggers the endogenous RNAi pathway, resulting in cleavage and degradation of the target mRNA molecule. Various improvements to siRNA compositions, such as incorporation of modified nucleosides or motifs into one or both strands of the siRNA molecule to enhance stability, specificity, and efficacy, have been described and are suitable for use in accordance with this aspect of the invention (see, for example, WO2004 / 015107 by Giese et al.; WO2003 / 070918 by McSwiggen et al.; WO1998 / 39352 by Imanishi et al.; U.S. Patent Application Publication No. 2002 / 0068708 by Jesper et al.; U.S. Patent Application Publication No. 2002 / 0147332 by Kaneko et al.; U.S. Patent Application Publication No. 2008 / 0119427 by Bhat et al., which are hereby incorporated by reference in their entirety).

[0166] In another embodiment, the one or more modulators comprise siRNA prepared by an endoribonuclease (esiRNA), which comprises a mixture of siRNA oligonucleotides formed by cleavage of long double-stranded RNA with an endoribonuclease (e.g., RNase III or dicer). Digestion of the synthetic long double-stranded RNA yields short overlapping siRNA fragments between 18-25 bases in length, all of which target the same mRNA sequence. A complex mixture of many different siRNAs all targeting the same mRNA sequence results in increased silencing efficacy. Targeting of long non-coding RNAs using esiRNA technology has been described in the art (Theis et al., “Targeting Human Long Noncoding Transcripts by Endoribonuclease-Prepared siRNAs,” J. Biomol. Screen 20(8):1018-1026 (2015), which is hereby incorporated by reference in its entirety).

[0167] The one or more modulators can also be short or small hairpin RNAs. Short or small hairpin RNA molecules are functionally similar to siRNA molecules but contain longer RNA sequences that form a tight hairpin loop. The shRNA is cleaved by the cellular machinery into siRNA, and gene expression is silenced via the cellular RNA interference pathway.

[0168] Nucleic acid aptamers that specifically bind to one or more of the genes described herein can also be used in the methods of the invention. A nucleic acid aptamer is a single-stranded, partially single-stranded, partially double-stranded or double-stranded nucleotide sequence, advantageously a replicable nucleotide sequence, capable of specifically recognizing a selected non-oligonucleotide molecule or group of molecules by a mechanism other than Watson-Crick base pairing or triplex formation. Aptamers include, but are not limited to, defined sequence segments and sequences that contain nucleotides, ribonucleotides, deoxyribonucleotides, nucleotide analogs, modified nucleotides, and nucleotides containing backbone modifications, branch points, and non-nucleotide residues, groups or bridges. Nucleic acid aptamers include partially and fully single-stranded and double-stranded nucleotide molecules and sequences; synthetic RNAs, DNAs and chimeric nucleotides; hybrids; duplexes; heteroduplexes; and any ribonucleotide, deoxyribonucleotide or their chimeric counterparts and / or the corresponding complementary sequences, promoters or primer annealing sequences required to amplify, transcribe or replicate all or part of the aptamer molecule or sequence.

[0169] In the above-described embodiments, the one or more modulators may be packaged in a suitable delivery vehicle or carrier for delivery to a subject. Suitable delivery vehicles include, but are not limited to, viruses, virus-like particles, bacteria, bacteriophages, biodegradable microspheres, microparticles, nanoparticles, exosomes, liposomes, collagen minipellets, and cochleates. These and other biological gene delivery vehicles are well known to those of skill in the art (see, e.g., Seow and Wood, “Biological Gene Delivery Vehicles: Beyond Viral Vectors,” Mol. Therapy 17(5):767-777 (2009), which is hereby incorporated by reference in its entirety).

[0170] In one embodiment, the modulator is packaged into a therapeutic expression vector to facilitate delivery. Suitable expression vectors are well known in the art and include, but are not limited to, viral vectors such as adenoviral vectors, adeno-associated viral vectors, retroviral vectors, lentiviral vectors, or herpes viral vectors. The viral vector or other suitable expression vector contains a sequence encoding the inhibitory nucleic acid molecule of the invention (e.g., siRNA, ASO, etc.) and any suitable promoter for expressing the inhibitory sequence. Suitable promoters include, for example but not limited to, U6 or HI RNA pol III promoter sequences and cytomegalovirus promoters. The selection of other suitable promoters is within the scope of those of skill in the art. The expression vector may also contain an inducible or regulatable promoter for expressing the inhibitory nucleic acid molecule in a tissue- or cell-specific manner.

[0171] Gene therapy vectors carrying therapeutic inhibitory nucleic acid molecules are administered to a subject by, for example, intravenous injection, local administration (U.S. Patent No. 5,328,470 to Nabel et al., which is hereby incorporated by reference in its entirety), or by stereotactic injection (see, e.g., Chen et al., “Gene Therapy for Brain Tumors: Regression of Experimental Gliomas by Adenovirus Mediated Gene Transfer In Vivo,” Proc. Nat’l. Acad. Sci. USA 91:3054 - 3057 (1994), which is hereby incorporated by reference in its entirety). The pharmaceutical composition of the therapeutic vector can comprise the therapeutic vector in an acceptable diluent, or can comprise a sustained release matrix in which the therapeutic delivery vehicle is encapsulated. Alternatively, when the intact therapeutic delivery vector can be produced intact from recombinant cells (e.g., retroviral vectors), the pharmaceutical composition can comprise one or more cells that produce the therapeutic delivery system. Gene therapy vectors typically utilize constitutive regulatory elements that are responsive to endogenous transcription factors.

[0172] Another suitable route for delivering the modulators of the present disclosure involves the use of liposomal delivery vehicles or nanoparticle delivery vehicles.

[0173] In one embodiment, a pharmaceutical composition or formulation containing an inhibitory nucleic acid molecule (e.g., an siRNA molecule) is encapsulated in a lipid formulation to form a nucleic acid - lipid particle, as described in Semple et al., “Rational Design of Cationic Lipids for siRNA Delivery,” Nature Biotech. 28:172–176 (2010), and WO2011 / 034798, WO2009 / 111658, and WO2010 / 105209 to Bumcrot et al. (said documents are hereby incorporated by reference in their entirety). Other cationic lipid carriers suitable for delivering ASOs include, but are not limited to: N-[1-(2,3 - dioleyloxy)propyl]-N,N,N - trimethylammonium chloride (DOTMA) and N-[1-(2,3 - dioleyloxy)propyl]-N,N,N - trimethylammonium methyl sulfate (DOTAP) (see Chan et al., “Antisense Oligonucleotides: From Design to Therapeutic Application,” Clin. Exp. Pharm. Physiol. 33:533 - 540 (2006), which is hereby incorporated by reference in its entirety).

[0174] In another embodiment of the present invention, the delivery vehicle is a nanoparticle. A variety of nanoparticle delivery vehicles are known in the art and are suitable for delivering the modulators of the present invention (see, for example, van Vlerken et al., “Multi-functional Polymeric Nanoparticles for Tumour-Targeted Drug Delivery,” Expert Opin. Drug Deliv. 3(2):205–216 (2006), which is hereby incorporated by reference in its entirety).Suitable nanoparticles include, but are not limited to, poly(β - amino esters) (Sawicki et al., “Nanoparticle Delivery of Suicide DNA for Epithelial Ovarian Cancer Cell Therapy,” Adv. Exp. Med. Biol. 622:209–219 (2008), which is hereby incorporated by reference in its entirety), polyethyleneimine - alt - poly(ethylene glycol) copolymers (Park et al., “Degradable Polyethylenimine - alt - Poly(ethylene glycol) Copolymers As Novel Gene Carriers,” J. Control Release 105(3):367–80 (2005) and Park et al., “Intratumoral Administration of Anti - KITENIN shRNA - Loaded PEI - alt - PEG Nanoparticles Suppressed Colon Carcinoma Established Subcutaneously in Mice,” J. Nanosci. Nanotechnology 10(5):3280–3 (2010), which are hereby incorporated by reference in their entirety), poly(d,l - lactide - co - glycolide) (Chan et al., “Antisense Oligonucleotides: From Design to Therapeutic Application,” Clin. Exp. Pharm. Physiol. 33:533 - 540 (2006), which is hereby incorporated by reference in its entirety), and liposome - encapsulated siRNA nanoparticles (Kenny et al., “Novel Multifunctional Nanoparticle Mediates siRNA Tumor Delivery, Visualization and Therapeutic Tumor Reduction In Vivo,” J. Control Release 149(2):111–116 (2011), which is hereby incorporated by reference in its entirety). Other nanoparticle delivery mediators suitable for the present invention include the microcapsule nanotube device disclosed in U.S. Patent Publication No. 2010 / 0215724 to Prakash et al. (which is hereby incorporated by reference in its entirety).

[0175] In another embodiment, the pharmaceutical composition is contained within a liposome delivery vehicle. The term "liposome" means a vesicle composed of amphiphilic lipids arranged in one or more spherical bilayers. Liposomes are single- or multi-layered vesicles that have a membrane formed from lipophilic substances and an aqueous interior. The aqueous portion contains the composition to be delivered. Cationic liposomes have the advantage of being able to fuse with cell walls. Non-cationic liposomes, while not able to fuse efficiently with cell walls, are taken up by macrophages in the body.

[0176] Several advantages of liposomes include: they are biocompatible and biodegradable, can incorporate a wide range of water-soluble and lipid-soluble drugs; and they can protect the encapsulated drugs from metabolism and degradation. Important considerations in preparing liposome formulations are the lipid surface charge, vesicle size, and the aqueous volume of the liposome.

[0177] Liposomes can be used to transfer and deliver active ingredients to the site of action. Since the liposome membrane is structurally similar to biological membranes, when liposomes are applied to tissues, the liposomes begin to fuse with cell membranes, and as the fusion of liposomes and cells progresses, the liposome contents are emptied into the cells where the active agent can act.

[0178] Methods for preparing liposomes for use in the present invention include those disclosed in the following: Bangham et al., "Diffusion of Univalent Ions Across the Lamellae of Swollen Phospholipids," J. Mol. Biol. 13:238–52 (1965); U.S. Patent No. 5,653,996 to Hsu; U.S. Patent No. 5,643,599 to Lee et al.; U.S. Patent No. 5,885,613 to Holland et al.; U.S. Patent No. 5,631,237 to Dzau et al.; and U.S. Patent No. 5,059,421 to Loughrey et al., which are incorporated herein by reference in their entirety.

[0179] In one aspect, one or more modulators of a glial cell differentiation regulatory gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the glial cell differentiation regulatory gene selected from the group consisting of BMP2, LINGO1, MAG, NKX2-2, NR2E1, NTRK3, OLIG2, SERPINE2, SIRT2, and TCF7L2, or a protein encoded thereby.

[0180] Exemplary modulators of these genes include, but are not limited to: the synthetic non-peptidyl small molecules Hh-Ag 1.1 and related molecules Hh-Ag 1.2, Hh-Ag 1.3, Hh-Ag 1.4, and Hh Ag1.5, which affect the Hedgehog signaling pathway (Frank-Kamenetsky et al., “Small-molecule Modulators of Hedgehog Signaling: Identification and Characterization of Smoothened Agonists and Antagonists,” J. Biol. 1(2):10(2002), which is hereby incorporated by reference in its entirety) and agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and Simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28(2014) (which is hereby incorporated by reference in its entirety); Opicinumab; GSK-249320; sodium dodecyl sulfate; repaglinide; altiratinib; chembl2007421; PLX-3397; radicicol; thyroxine; entrectinib; LOXO-101; CEP-2563; lestaurtinib; PLX-7486; AZD-6918; AZD-7451; midostaurin; and combinations thereof.

[0181] In another aspect, one or more modulators of a gene associated with myelination are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the gene associated with myelination being selected from the group consisting of: FA2H, GAL3ST1, MAG, MBP, MYRF, NFASC, OLIG2, OMG, PLLP, POU3F2, SIRT2, SLC8A3, TCF7L2, TF, and UGT8, or a protein encoded thereby.

[0182] Exemplary modulators for these genes include, but are not limited to: agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); GSK-249320; sodium dodecyl sulfate, repaglinide; cyclosporine; interferon β-1A; prednisone; quercetin and rutin, and combinations thereof.

[0183] In yet another aspect, one or more modulators of oligodendrocyte differentiation genes are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the oligodendrocyte differentiation genes selected from the group consisting of: FA2H, GLI3, LINGO1, MYRF, NKX2-2, OLIG1, OLIG2, OMG, SIRT2, SLC8A3, SOX10, and TCF7L2, or the proteins encoded thereby.

[0184] Exemplary modulators for these genes include, but are not limited to: the synthetic non-peptidyl small molecules Hh-Ag 1.1 and related molecules Hh-Ag 1.2, Hh-Ag 1.3, Hh-Ag 1.4, and Hh Ag1.5, which affect the Hedgehog signaling pathway (Frank-Kamenetsky et al., “Small-molecule Modulators of Hedgehog Signaling: Identification and Characterization of Smoothened Agonists and Antagonists,” J. Biol. 1(2):10 (2002), which is hereby incorporated by reference in its entirety), and agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); opicinumab; sodium dodecyl sulfate; repaglinide; vemurafenib; and combinations thereof.

[0185] In a further aspect, one or more modulators of glia-generating regulatory genes are administered to a selected subject under conditions effective to treat or inhibit onset of Huntington's disease in the subject, the glia-generating regulatory genes being selected from the group consisting of: BMP2, LINGO1, MAG, MYC, NKX2-2, NR2E1, NTRK3, OLIG2, SERPINE2, SIRT2, SOX10, TCF7L2, TF, and ZCCHC24, or proteins encoded thereby.

[0186] Exemplary modulators of these genes include, but are not limited to: the synthetic non-peptidyl small molecules Hh-Ag 1.1 and related molecules Hh-Ag 1.2, Hh-Ag 1.3; Hh-Ag 1.4 and Hh Ag1.5, which affect the Hedgehog signaling pathway (Frank-Kamenetsky et al., “Small-molecule Modulators of Hedgehog Signaling: Identification and Characterization of Smoothened Agonists and Antagonists,” J. Biol. 1(2):10(2002) (which is hereby incorporated by reference in its entirety)), and agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28(2014) (which is hereby incorporated by reference in its entirety); opitzumab; GSK-249320; sodium dodecyl sulfate; vemurafenib; repaglinide; nadroparin calcium; 4'-hydroxytamoxifen; azacitidine; thioguanine; activin; adozelesin; amifostine; aminopterin; antibiotics; bizelesin; bromocriptine; bryostatin; calcitriol; diethylstilbestrol; elsamitrucin; estrone; folic acid; glutamine; hypoxanthine; imatinib; Cilmostin; melatonin; methylprednisolone; N-methyl-n-nitrosourea; novobiocin; Chembl35482; phorbol myristate acetate; prednisone; quinapril; Vorinostat; sulindac; thrombin; thyrotropin; β-nicotinamide adenine dinucleotide phosphate sodium; troglitazone; verapamil; Chembl100014; Chembl1213492; chorionic gonadotropin; perillyl alcohol; AMG-900; Alisertib; Dinaciclib; Roniciclib; temozolomide; Prexasertib; atelatinib; chembl2007421; PLX-3397; radicicol; thyroxine; entrectinib; LOXO-101; CEP-2563; lestaurtinib; PLX-7486; AZD-6918; AZD-7451; midostaurin; and combinations thereof.

[0187] In another aspect of the present disclosure, one or more modulators of neuronal ensheathment genes are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the neuronal ensheathment genes selected from the group consisting of: FA2H, GAL3ST1, MAG, MBP, MYRF, NFASC, OLIG2, OMG, PLLP, POU3F2, SIRT2, SLC8A3, TCF7L2, TF, and UGT8, or the proteins encoded thereby.

[0188] Exemplary modulators of these genes include, but are not limited to: agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); GSK-249320; cyclosporine; interferon β-1A; prednisone; quercetin; rutin; sodium dodecyl sulfate; repaglinide; and combinations thereof.

[0189] In another aspect, one or more modulators of axon guidance genes are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the axon guidance genes selected from the group consisting of: ALCAM, BCL11B, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, MNX1, NFASC, PLXNC1, PRKCQ, PTPRO, ROBO2, SEMA6B, UNC5A, VAX1, and WNT7B, or the proteins encoded thereby.

[0190] Exemplary modulators for these genes include, but are not limited to, agonists of the Wnt signaling pathway, including, but not limited to, 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); fluorouracil; CEP-2563; staurosporine; Chembl369507; dexfosfomycin; ticlopidine; GSK-690693; sertraline; (7S)-hydroxy-staurosporine; midostaurin; quercetin; bryostatin; sertraline acetate; ingenol mebutate; carboplatin; paclitaxel; and combinations thereof.

[0191] In yet another aspect, one or more modulators of a neuronal projection guidance gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the neuronal projection guidance gene selected from the group consisting of: ALCAM, BCL11B, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, MNX1, NFASC, PLXNC1, PRKCQ, PTPRO, ROBO2, SEMA6B, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0192] Exemplary modulators for these genes include, but are not limited to: agonists of the Wnt signaling pathway, including, but not limited to, 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); fluorouracil; CEP-2563; staurosporine; Chembl369507; dexfosfomycin; ticlopidine; GSK-690693; sertraline; (7S)-hydroxy-staurosporine; midostaurin; quercetin; bryostatin; sertraline acetate; ingenol mebutate; carboplatin; paclitaxel; and combinations thereof.

[0193] In another aspect, one or more modulators of axonogenic genes are administered to a selected subject under conditions effective to treat or inhibit onset of Huntington's disease in the subject, the axonogenic genes selected from the group consisting of: ADGRB1, ALCAM, BCL11B, CACNA1A, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, LINGO1, LRRC4C, MAG, MBP, MNX1, NFASC, NR2E1, NTNG1, NTRK3, OMG, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, SEMA6B, SLITRK2, SLITRK3, SNAP91, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0194] Exemplary modulators of these genes include, but are not limited to: agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); opicinumab; GSK-249320; cyclosporine; interferon beta-1A; prednisone; quercetin; rutin; fluorouracil; CEP-2563; staurosporine; Chembl369507; D-serine; ticlopidine; GSK-690693; sotrastaurin; (7S)-hydroxy-staurosporine; midostaurin; bryostatin; sotrastaurin acetate; ingenol mebutate; carboplatin; paclitaxel; pregabalin; verapamil; bepridil; celecoxib; nisoldipine; gabapentin; gabapentin enacarbil; eptinezumab; atogepant; bepridil hydrochloride; amocarbose; ataluren; chembl2007421; PLX-3397; radicicola; thyroxine; entrectinib; loxo-101; CEP-2563; lestaurtinib; PLX-7486; AZD-6918; AZD-7451; and combinations thereof.

[0195] In another aspect, one or more modulators of an axon development gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the axon development gene selected from the group consisting of: ADGRB1, ALCAM, BCL11B, CACNA1A, DSCAM, FOXD1, GAS1, GLI3, HOXA1, HOXA2, LINGO1, LRRC4C, MAG, MBP, MNX1, NEFM, NFASC, NR2E1, NTNG1, NTRK3, OMG, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, RTN4RL2, SEMA6B, SLITRK2, SLITRK3, SNAP91, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0196] Exemplary modulators of these genes include, but are not limited to: agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); opicinumab; dexofosfoserine; fluorouracil; CEP-2563; staurosporine; Chembl369507; GSK-249320; ticlopidine; GSK-690693; sotrastaurin; (7S)-hydroxy-staurosporine; midostaurin; quercetin; bryostatin; acetate sotrastaurin; and ingenol mebutate; carboplatin; paclitaxel; pregabalin; verapamil; bepridil; celecoxib; nisoldipine; gabapentin; gabapentin enacarbil; eptinezumab; atogabalin; hydrochloride bepridil; amocarbose; ataluren; chembl2007421; PLX-3397; radicicola; thyroxine; entrectinib; loxo-101; CEP-2563; lestaurtinib; PLX-7486; AZD-6918; AZD-7451; cyclosporine; interferon beta-1A; prednisone; rutin; and combinations thereof.

[0197] In a further aspect of the present disclosure, one or more modulators of a cell projection morphogenesis gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the cell projection morphogenesis gene being selected from the group consisting of: ADGRB1, ALCAM, BCL11B, CACNA1A, CAMK2A, DSCAM, EHD3, FOXD1, GAS1, GLI3, HOXA1, HOXA2, KANK1, LINGO1, LRRC4C, MAG, MBP, MNX1, NEDD4L, NEURL1, NFASC, NR2E1, NTNG1, NTRK3, OMG, PCDH15, PLXNC1, POU3F2, PRKCQ, PTPRO, ROBO2, SEMA6B, SGK1, SLITRK2, SLITRK3, SNAP91, SNX10, UGT8, UNC5A, VAX1, and WNT7B, or a protein encoded thereby.

[0198] Exemplary modulators of these genes include, but are not limited to: agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); opicinumab; GSK-249320; cyclosporine; interferon β-1A; prednisone; quercetin; rutin; D-serine; fluorouracil; CEP-2563; staurosporine; Chembl369507; ticlopidine; GSK-690693; sortilin; (7S)-hydroxy-staurosporine; midostaurin; bryostatin; sortilin acetate; and ingenol mebutate; carboplatin; paclitaxel; pregabalin; verapamil; bepridil; celecoxib; nisoldipine; gabapentin; gabapentin enacarbil; eptinezumab; ataluren; benpridil hydrochloride; amocarbose; atelestratide; Chembl2007421; PLX-3397; radicicola; thyroxine; entrectinib; loxo-101; CEP-2563; lestaurtinib; PLX-7486; AZD-6918; AZD-7451; hydrochlorothiazide; chembl549906; chembl550795; sodium chloride; GSK-650394; and combinations thereof.

[0199] In another aspect of the present disclosure, one or more modulators of a synaptic structure or activity-regulating gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the synaptic structure or activity-regulating gene being selected from the group consisting of: ADGRB1, ADGRL3, BCAN, CALB1, CAMK2A, FGF14, LRRTIM1, NCDN, NETO1, NEURL1, NR2E1, NTRK3, PPFIA3, ROBO2, SERPINE2, SHISA7, SIX4, SLC8A3, SLITRK2, SLITRK3, and SYNDIG 1 or a protein encoded thereby.

[0200] Exemplary modulators of these genes include, but are not limited to: D-serine, entrectinib, chembl2007421, PLX-3397, radicicola, thyroxine, entrectinib, loxo-101, CEP-2563, lestaurtinib, PLX-7486, AZD-6918, AZD-7451, midostaurin, and combinations thereof.

[0201] In a further aspect, one or more modulators of a synaptic signaling pathway gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the synaptic signaling pathway gene being selected from the group consisting of: BCAN, CACNA1A, CACNA1G, CALB1, CAMK2A, CHRNA4, FGF12, FGF14, GRIA2, GRIA4, GRID2, GRIK4, KCND2, LRRTM1, MBP, MPZ, NCDN, NETO1, NEURL1, NOVA1, NR2E1, P2RX7, PDE7B, PLCL1, PPFIA3, RAPGEF4, RGS8, RIT2, S1PR2, SERPINE2, SHISA7, SLC18A1, SLC1A1, SLC1A2, SLC8A3, SNAP91, SNPH, and SYT6 or a protein encoded thereby.

[0202] Exemplary modulators for these genes include, but are not limited to: pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, elpetrigine, atagabalin, bepridil hydrochloride, imagabalin, cyclosporine, interferon beta-1A, prednisone, quercetin, rutin, nicotine polacrilex, talbutal, butabarbital, butalbital, secobarbital, metharbital, thiopental, primidone, mephobarbital, phenobarbital, varenicline, amobarbital, aprobarbital, butethal, heptabarbital, hexobarbital, barbital, pozanicline, cytisine, rivanicline, epibatidine, chembl1876219, chembl3103988, atracurium, chembl490153, hexamethonium, chembl407217, TC-2216, ABT-560, ispronicline, sofinicline, TC-6499, AZD1446, CP-601927, dexmecamylamine, nicotine, varenicline tartrate, benztropine mesylate, pentolinium, azd0328, bradanicline, pentobarbital,chembl1201135, dexefaroxan, mecamylamine (chembl267936), dianicline, altinicline, trimethaphan, oleic acid, tebanicline tosylate, mibampator, Butesa, (r,s)-ampa, chembl123132, aniracetam, chembl136800, chembl1255648, cyclothiazide, chembl77862, chembl334920, chembl1097939, piracetam, chembl320642, chembl265301, gyki-52466, NBQX, chembl222418, tezampanel, (s)-ampa, chembl594840, chembl121915, quisqualate, chembl337577, chembl27130, DNQX, chembl333964, (s)-willardiine, chembl28472, talampanel, perampanel, irampanel, CX1739, dasolampanel, becampanel, farampator, mk-8777, zonampanel, pentobarbital, pf-04958242, Selurampanel, dalfampridine, guanidine hydrochloride, tedisamil, nerispirdine, evt401, adenosine triphosphate, chembl335550, chelerythrine, acebutolol, moclobemide, ivermectin, chemb377219, chembl255787, methylclothiazide, chembl550637, sodium orthovanadate, chembl2338352, benzonatate, GSK1482160, AZD9056, CE224535,Dyphylline, chembl484928, Dipyridamole, Flavoxate Hydrochloride, Pentoxifylline, Quinacrine, chembl2313646, chembl570352, Ozanimod, chembl225155, chembl1368758, Fingolimod Hydrochloride, Amiselimod Hydrochloride, Reserpine, Norepinephrine, chembl126506, Methamphetamine, Ketanserin, Tetrabenazine, L - Glutamate, Dihydrokainate, 2s,4r - 4 - Methylglutamate, o - Benzyl - L - Serine, chembl1628669, and Mesalamine, Tezampanel, Domoic Acid, Dysiherbaine, Kainic Acid, Mesalamine, Topiramate, Aspartic Acid, Clozapine, Alcohol, Haloperidol, Wortmannin, Olanzapine, Phorbol Myristate Acetate, Risperidone, Lidocaine, Pregabalin, Gabapentin Enacarbil, Mibefradil Dihydrochloride, Trimethadione, Cinnarizine, Ethosuximide, Zonisamide, Anandamide, Mibefradil, chembl1684954, Flunarizine, Methsuximide, Bepridil Hydrochloride,Gabapentin, phensuximide, paramethadione, atagabalin, celecoxib, imagabalin; and combinations thereof.

[0203] In another aspect, administering to a selected subject one or more modulators of a synaptic gene, said synaptic gene selected from the group consisting of: ADGRB1, BCAN, BCAS1, CACNA1A, CALB1, CAMK2A, CHRNA4, CTTNBP2, DSCAM, GRIA2, GRID1, GRID2, GRIK4, HCN2, KCND2, LGI3, LRRC4C, LRRTM1, NETO1, NEURL1, NTM, P2RX7, PCDH15, PDE4B, PPFIA3, PRIMA1, PRKCQ, PTPRO, RAPGEF4, SERPINE2, SHISA7, SLC17A8, SLC18A1, SLC1A1, SLC1A2, SLC8A3, SNAP91, SNPH, SYNDIG1 and SYT6, or a protein encoded thereby, under conditions effective to treat or inhibit onset of Huntington's disease in said subject.

[0204] Exemplary modulators for these genes include, but are not limited to: L-2-Aminophosphonobutyric acid, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptinezumab, atalampretig, bepridil hydrochloride, octanoic acid, mibampator, bursatol, secbutabarbital, butalbital, talbutal, secobarbital, metharbital, thiopental, primidone, mephobarbital, phenobarbital, (R,s)-AMPA, CHEMBL123132, aniracetam, CHEMBL136800, CHEMBL1255648, cyclothiazide, CHEMBL77862, CHEMBL334920, CHEMBL1097939, piracetam, CHEMBL320642, CHEMBL265301, GYKI-52466, NBQX, CHEMBL222418, tezampanel, amobarbital, aprobarbital, heptabarbital, hexobarbital, barbital, (s)-AMPA, CHEMBL594840, CHEMBL121915, quisqualic acid, CHEMBL337577, CHEMBL27130, DNQX, CHEMBL333964, (s)-willardiine, CHEMBL28472, talampanel, perampanel, elampanel, CX1739, darampanel, becampanel, farampator, MK-8777, zonampanel, topiramate, pentobarbital, PF-04958242, selurampanel, polacrilex nicotine, varenicline, butex, pazacrine, cytisine, rivanicline, epibatidine, CHEMBL1876219, CHEMBL3103988, atracurium, CHEMBL490153, hexamethonium, CHEMBL407217, TC-2216, ABT-560, isopropylcran, sophinicline, TC-6499, AZD1446, CP-601927, dexmecamylamine, nicotine, varenicline tartrate, benzatropine mesylate, pentolonium, AZD0328, bradanicline, pentobarbital, CHEMBL1201135, dexefaroxan, mecamylamine (CHEMBL267936), dianicline, aticline, trimethaphan, oleic acid, tibacrine tosylate, polacrilex nicotine, carboplatin, paclitaxel, L-glutamate, dalfampridine, guanidine hydrochloride, tedisamil, niliridine, EVT401, adenosine triphosphate, CHEMBL335550, chelerythrine, acebutolol, moclobemide, ivermectin, CHEMB377219, CHEMBL255787, methyclothiazide, CHEMBL550637, sodium orthovanadate, CHEMBL2338352, benzonatate, GSK1482160, AZD9056, CE224535, reserpine, norepinephrine, CHEMBL126506, methamphetamine, ketanserin, tetrabenazine, L-glutamate, dihydrocinchoninate, 2S,4R-4-methylglutamate, O-benzyl-L-serine, chembl1628669, mesalamine, tezampanel, domoic acid, dysiherbaine, kainic acid, mesalamine, topiramate, CEP-2563, staurosporine, Chembl369507, ticlopidine, GSK-690693, surotomaline, (7S)-hydroxy-staurosporine, midostaurin, quercetin, bryostatin, surotomaline acetate, ingenol mebutate, adenosine phosphate, theophylline, aminophylline, pentoxifylline, enprofylline, iloprost, papaverine, theobromine, inamrinone, [r]-mexepramine, roflumilast, piclamilast, rolipram, fevipiprant, chembl1230617, chembl519827, cilomilast, (-)-rolipram, crisaborole, ibudilast, apremilast, chembl521203, chembl74078, propoxyphene, cdp840, sodium phenylbutyrate, chembl1232082, dipyridamole, sodium theophyllinate, flavoxate hydrochloride, aminophylline, resveratrol, caffeine, choline theophyllinate, amlexanox, edizole, ciloridine, zaltidine, chembl2052019, chembl395336, cyclic adenosine monophosphate, aspartic acid, clozapine, alcohol, haloperidol, wortmannin, olanzapine, phorbol 12-myristate 13-acetate, risperidone, lidocaine, and combinations thereof.,

[0205] In yet another aspect, one or more modulators of a monovalent inorganic cation transporter gene are administered to a selected subject under conditions effective to treat or inhibit onset of Huntington's disease in the subject, the monovalent inorganic cation transporter gene selected from the group consisting of ABCC9, ASIC4, CACNA1A, CHRNA4, CNGB1, CNTN1, DPP10, DPP6, FGF12, FGF14, HCN2, KCND2, KCNJ9, KCNQ1, KCNS3, NALCN, NEDD4L, NKAIN4, P2RX7, PTGER3, SERPINE2, SGK1, SLC10A4, SLC17A8, SLC18A1, SLC22A3, SLC2A13, SLC5A9, SLC8A3, and SLC9A7, or a protein encoded thereby.

[0206] Exemplary modulators for these genes include, but are not limited to: nimodipine, adenosine triphosphate, glibenclamide, saralasin, pinacidil hydrate, minoxidil, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, eptazocine, atagabalin, bepridil hydrochloride, amelexanox, chembl549906, chembl550795, sodium chloride, GSK-650394, dalfampridine, guanidine hydrochloride, tedisamil, nilipride, evt401, adenosine triphosphate, chembl335550, sanguinarine, acebutolol, moclobemide, ivermectin, chemb377219, chembl255787, methyclothiazide, chembl550637, sodium orthovanadate, chembl2338352, benzonatate, GSK1482160, AZD9056, CE224535, hydrochlorothiazide, chembl1229875, varenicline, secbutabarbital, amobarbital, butabarbital, secobarbital, metharbital, thiopental, primidone, tolbutamide, phenobarbital, varenicline, amobarbital, aprobarbital, butethal, heptabarbital, hexobarbital, barbital, prazacrine, cytisine, rivanicline, epibatidine, chembl1876219, chembl3103988, atracurium, chembl490153, hexamethonium, chembl407217, tc-2216, abt-560, isopropylcranil, solfenicline, tc-6499, ciladrine, zatebradine, chembl2052019, chembl395336, cyclic adenosine monophosphate, chembl99951, flupirtine, indapamide, bepridil, azimilide, chembl2070953, mefenamic acid, chembl1907717, niflumic acid, chembl298475, chembl342375, chembl332826, dolasetron, celecoxib, nilipride, ezogabine, indomethacin, tacrolimus, guanidine hydrochloride, tedisamil, dalfampridine, pyrimethamine, cobalt(ii) ion verapamil pyrimethamine cobalt(ii) ion, dihydrocinchonine, bimatoprost, dinoprostone, misoprostol, beraprost, chembl1628262, carbacyclin, cicaprost, cloprostenol (chembl2220404), enprostil, fluprostenol, iloprost, dinoprost, sulprostone, treprostinil, chembl357834, chembl1317823, chembl565591, chembl358653, sarcnu, and combinations thereof.

[0207] In a further aspect of the present disclosure, one or more modulators of a neuronal projection gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the neuronal projection gene being selected from the group consisting of ADGRL3, ALCAM, BCAN, BCL11B, CACNA1A, CACNA1G, CALB1, CAMK2A, CHRNA4, CTTNBP2, DSCAM, GRIA2, GRIA4, GRID2, GRIK4, HCN2, KCND2, LGI3, LRRTM1, MAG, MBP, MYC, NCAM2, NCDN, NEFM, NEURL1, NFASC, NTM, PDE4B, PIK3R1, PTGER3, PTPRO, RAPGEF4, RGS8, ROBO2, SGK1, SIRT2, SLC17A8, SLC1A2, SLC8A3, SNAP91, SNPH, SYNDIG1, and UNC5A, or a protein encoded thereby.

[0208] Exemplary modulators of these genes include, but are not limited to, adenosine phosphate, theophylline, theophylline, pentoxifylline, enprophylline, iloprost, papaverine, theobromine, aminopyrazone, [R]-methopram, roflumilast, pyramilast, rolipram, filminlast, chemb11230617, chemb1519827, cilomilast, (-)-rolipram, crisaborole, ibudilast, apremilast, chemb1521203, chemb174078, propoxyphene, cdp840, sodium phenylbutyrate, c hembl1232082, dipyridamole, theophylline sodium glycinate, flavoxate hydrochloride, aminophylline, resveratrol, caffeine, theophylline, amlexanox, edizalate, pregabalin, verapamil, bepridil, celecoxib, nisoldipine, gabapentin, gabapentin enacarbil, elpetrigine, atagabalin, bepridil hydrochloride, amine octanoic acid, carboplatin, paclitaxel, chembl549906, chembl550795, sodium chloride, GSK-650394, dalfampridine, guanidine hydrochloride, tedisamil, neripridine, L-glutamate, dihydrocainate, 2S,4R-4-methylglutamate, O-benzyl-L-serine, chembl1628669, mesalamine, fluorouracil, pregabalin, gabapentin enacarb, mibefradil dihydrochloride, trimethadione, cinnarizine, ethosuximide, zonisamide, cannabinoids, mibefradil, chembl1684954, flunarizine, ethosuximide, bepridil hydrochloride, gabapentin, phensuximide, ethadione, atagabalin, celecoxib, and aminomethacrylic acid, polacrifo nicotine, talbutal, butalbital, butalbital, butalbital, butalbital cobarbital, metharbital, thiopental, primidone, mebendal, phenobarbital, varenicline, amobarbital, aprobarbital, butesa, enanthate, hexobarbital, barbital, pazakal, cytisine, rivanicline, epibatidine, chembl1876219, chembl3103988, atracurium, chembl490153, hexamethonium, chembl407217, tc-2216, abt-560, isopyruvate, sofenicline, tc-6499, mibampator, (r,s)-AMPA, CHEMBL123132, aniracetam, CHEMBL136800, CHEMBL1255648, cyclothiazide, CHEMBL77862, CHEMBL334920, CHEMBL1097939, piracetam, CHEMBL320642, CHEMBL265301, GYKI-52466, NBQX, CHEMBL222418, tezampanel, (s)-AMPA, CHEMBL594840, CHEMBL121915, quisqualic acid, CHEMBL337577, CHEMBL27130, DNQX, CHEMBL333964, (s)-willardiine, CHEMBL28472, talampanel, perampanel, elampanel, CX1739, darampanel, beclamipanel, farampator, MK-8777, zonampanel, topiramate, pentobarbital, PF-04958242, selurampanel, cyclothiazide, CHEMBL334920, CHEMBL1097939, joro spider toxin, domoic acid, dysherbacine, kainic acid, mesalamine, 2S,4R-4-Methylglutamate, Chembl2313646, Cyclosporine, Interferon beta-1A, Prednisone, Quercetin, Rutin, GSK-249320, Cilostazol, Zaltoprofen, Chembl2052019, Chembl395336, Cyclic adenosine monophosphate, Sodium dodecyl sulfate, Bimatoprost, Dinoprostone, Misoprostol, Beraprost, Chembl1628262, Carbacyclin, Cicaprost, Cloprostenol (Chembl2220404), Enprostil, Fluprostenol, Iloprost, Dinoprost, Sulprostone, Treprostinil, Chembl357834, Chembl1317823, Chembl565591, Chembl358653, Nadroparin calcium, 4'-Hydroxytamoxifen, Azacitidine, Thioguanine, Activin, Adozelesin, Amifostine, Aminopterin, Antibiotics, Bizelesin, Bromocriptine, Bryostatin, Calcitriol, Diethylstilbestrol, Esafosfamide, Estrone, Folic acid, Glutamine, Hypoxanthine, Imatinib, Cilastatin, Melatonin, Methylprednisolone, N-Methyl-N-nitrosourea, Novobiocin, Chembl35482, Phorbol 12-myristate 13-acetate, Prednisone, Quinapril, Vorinostat, Sulindac, Thrombin, Thyrotropin, Beta-nicotinamide adenine dinucleotide phosphate sodium salt, Troglitazone, Verapamil, Chembl100014, Chembl1213492, Chorionic gonadotropin, Perillyl alcohol, AMG-900, Alisertib, Dinaciclib, Roniciclib, Temozolomide, Prexasertib, PF-04691502, Pracinostat, PA-799, Isoproterenol, sf-1126, Wortmannin, GSK-2636771, DS-7423, Omipalisib, Recilisib, PWT-33587, RG-7666, VS-5584, Copanlisib, Gedatolisib, Sonidegib, Apitolisib, Taselisib, Pilaralisib (Chembl3360203), Votarisib, ZSTK-474, Alpelisib, PI-103, Pilaralisib (Chembl3218575), WX-037, Dactolisib, BGT-226 (Chembl3545096), Pictilisib, Buparlisib, Panolisib, GSK-1059615, AZD-6482, Buparlisib hydrochloride, LY-3023414, and combinations thereof.,

[0209] In another aspect, one or more modulators of a TCF7L2 target gene are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject, the TCF7L2 target gene consisting of the group consisting of: BMP4, CCND1, CCND2, DOCK10, DOCK9, DUSP15, ENPP4, EPAS1, EPHB1, ERBB3, EVI2A, EVI2B, FA2H, GJB1, HAPLN2, HSPA2, ID3, LGI3, MBP, MOG, MYC, MYRF, NFASC, NKAIN1, NKX6-2, OLIG2, PLEKHB1, PLP1, PPP1R16B, RAB33A, RASGEF1B, RTKN, SIRT2, SLC1A2, SOX10, ST18, TMEM125, TMEM2, TPPP, TSPAN15, UGT8 and AATK, or a protein encoded thereby.

[0210] Exemplary modulators of these genes include, but are not limited to: agonists of the Wnt signaling pathway, including but not limited to 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (2-AMBMP), curcumin, and simvastatin, as described in Blagodatski et al., “Targeting the Wnt Pathways for Therapies,” Mol. Cell Ther. 2:28 (2014) (which is hereby incorporated by reference in its entirety); arsenic trioxide; acetaminophen; vitamin E; cytarabine; gossypol; roniciclib; ribociclib; palbociclib; methotrexate; mycophenolic acid; nifedipine; tamoxifen; troglitazone; uracil; abemaciclib; briciclib; abemaciclib; decitabine; palbociclib; pelareorep; cyclosporine; interferon beta-1a; prednisone; quercetin; rutin; vemurafenib; nadroparin calcium; 4'-hydroxytamoxifen; azacitidine; thioguanine; asivicin; adozelesin; amifostine; aminopterin; antibiotics; bizelesin; bromocriptine; bryostatin; calcitriol; diethylstilbestrol; elsamitrucin; estrone; folic acid; glutamine; hypoxanthine; imatinib; indomethacin; lithium; semustine; melatonin; methylprednisolone; N-methyl-N-nitrosourea; novobiocin, chembl35482; phorbol 12-myristate 13-acetate; prednisone; quinapril; vorinostat; sulindac; thrombin; thyrotropin; beta-nicotinamide adenine dinucleotide phosphate sodium; troglitazone; verapamil; chembl100014; chembl1213492; gonadotropins; chorionic gonadotropin; perillyl alcohol; AMG-900; alisertib; dinaciclib; temozolomide; prexasertib; sodium dodecyl sulfate; L-glutamate; dyclonine hydrochloride; 2S,4R-4-methylglutamate; O-benzyl-L-serine; chembl1628669; mesalamine; pelareorep; and combinations thereof.

[0211] In a last aspect of the disclosure, one or more modulators of a gene involved in the NKX2.2→OLIG2→SOX10→MYRF regulatory cascade or a protein encoded thereby are administered to a selected subject under conditions effective to treat or inhibit the onset of Huntington's disease in the subject.

[0212] Exemplary modulators of genes in this pathway include, but are not limited to, vemurafenib.

[0213] Exemplary regulators of the present invention and their corresponding gene targets are given in Table 4 below.

[0214] Table 4.

[0215]

[0216]

[0217]

[0218]

[0219]

[0220]

[0221]

[0222]

[0223]

[0224]

[0225]

[0226]

[0227]

[0228]

[0229] All references listed in Table 4 are hereby incorporated by reference in their entirety.

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[0261] In one embodiment, the methods described herein further comprise administering to a selected subject a preparation of human glial progenitor cells.

[0262] The human glial progenitor cells can be derived from any suitable glial cell source, such as but not limited to human induced pluripotent stem cells (iPSCs), embryonic stem cells, fetal tissue, and / or astrocytes, as described in more detail below.

[0263] iPSCs are pluripotent cells derived from non-pluripotent cells such as somatic cells. For example, but not limited to, iPSCs can be derived from tissues, peripheral blood, cord blood, and bone marrow (see, e.g., Cai et al., “Generation of Human Induced Pluripotent Stem Cells from Umbilical Cord Matrix and Amniotic Membrane Mesenchymal Cells,” J. Biol. Chem. 285(15):112227-11234 (2110); Giorgetti et al., “Generation of Induced Pluripotent Stem Cells from Human Cord Blood Cells with only Two Factors: Oct4 and Sox2,” Nat. Protocol. 5(4):811-820 (2010); Streckfuss-Bomeke et al., “Comparative Study of Human-Induced Pluripotent Stem Cells Derived from Bone Marrow Cells, Hair Keratinocytes, and Skin Fibroblasts,” Eur. Heart J. doi:10.1093 / eurheartj / ehs203 (July 12, 2012); Hu et al., “Efficient Generation of Transgene-Free Induced Pluripotent Stem Cells from Normal and Neoplastic Bone Marrow and Cord Blood Mononuclear Cells,” Blood doi:10.1182 / blood-2010-07-298331 (February 4, 2011); Sommer et al., “Generation of Human Induced Pluripotent Stem Cells from Peripheral Blood using the STEMCCA Lentiviral Vector,” J. Vis. Exp. 68:e4327 doi:10.3791 / 4327 (2012), which are hereby incorporated by reference in their entirety). Somatic cells are reprogrammed to an embryonic stem cell-like state through genetic manipulation.Exemplary somatic cells suitable for forming iPSCs include fibroblasts (see, e.g., Streckfuss-Bomeke et al., “Comparative Study of Human-Induced Pluripotent Stem Cells Derived from Bone Marrow Cells, Hair Keratinocytes, and Skin Fibroblasts,” Eur. Heart J. doi:10.1093 / eurheartj / ehs203 (2012), which is hereby incorporated by reference in its entirety), such as dermal fibroblasts obtained through a skin sample or biopsy, synoviocytes from synovial tissue, keratinocytes, mature B cells, mature T cells, pancreatic β cells, melanocytes, hepatocytes, foreskin cells, cheek cells, or lung fibroblasts.

[0264] Methods for generating induced pluripotent stem cells are known in the art and generally involve expressing a combination of reprogramming factors in somatic cells. Suitable reprogramming factors that promote and induce iPSC generation include one or more of Oct4, Klf4, Sox2, c-Myc, Nanog, C / EBPα, Esrrb, Lin28, and Nr5a2. In certain embodiments, at least two reprogramming factors are expressed in the somatic cells to successfully reprogram the somatic cells. In other embodiments, at least three reprogramming factors are expressed in the somatic cells to successfully reprogram the somatic cells.

[0265] iPSCs can be obtained by methods known in the art, including using integrating viral vectors (e.g., lentiviral vectors, inducible lentiviral vectors, and retroviral vectors), excisable vectors (e.g., transposons and floxed lentiviral vectors), and non-integrating vectors (e.g., adenoviruses and plasmid vectors) to deliver genes that promote cellular reprogramming (see, e.g., Takahashi and Yamanaka, Cell 126:663-676 (2006); Okita, et al., Nature 448:313-317 (2007); Nakagawa et al., Nat. Biotechnol. 26:101-106 (2007); Takahashi et al., Cell 131:1-12 (2007); Meissner et al., Nat. Biotech. 25:1177-1181 (2007); Yu et al., Science 318:1917-1920 (2007); Park et al., Nature 451:141-146 (2008); and U.S. Patent Application Publication No. 2008 / 0233610, which is hereby incorporated by reference in its entirety). Other methods for generating iPS cells include those disclosed in the following: WO2007 / 069666; WO2009 / 006930; WO2009 / 006997; WO2009 / 007852; WO2008 / 118820; U.S. Patent Application Publication No. 2011 / 0200568 to Ikeda et al.; U.S. Patent Application Publication No. 2010 / 0156778 to Egusa et al.; U.S. Patent Application Publication No. 2012 / 0276070 to Musick; and U.S. Patent Application Publication No. 2012 / 0276636 to Nakagawa et al.; Shi et al., Cell Stem Cell 3(5):568-574 (2008); Kim et al., Nature 454:646-650 (2008); Kim et al., Cell 136(3):411-419 (2009); Huangfu et al., Nat. Biotechnol. 26:1269-1275 (2008); Zhao et al., Cell Stem Cell 3:475-479 (2008); Feng et al., Nat. Cell Biol. 11:197-203 (2009); and Hanna et al., Cell 133(2):250-264 (2008), which are hereby incorporated by reference in their entirety).

[0266] The above-described iPSC generation method can be modified to include small molecules that enhance reprogramming efficiency or even replace reprogramming factors. These small molecules include, but are not limited to, epigenetic regulators such as the DNA methyltransferase inhibitor 5'-azacytidine, the histone deacetylase inhibitor VPA, the G9a histone methyltransferase inhibitor BIX-01294, and BayK8644 (an L-type calcium channel agonist). Other small molecule reprogramming factors include those that target signal transduction pathways such as TGF-β inhibitors and kinase inhibitors (e.g., kenpaullone) (see review by Sommer and Mostoslavsky, “Experimental Approaches for the Generation of Induced Pluripotent StemCells,” Stem Cell Res. Ther. 1:26 doi:10.1186 / scrt26 (August 10, 2010), which is hereby incorporated by reference in its entirety).

[0267] Methods for obtaining highly enriched glial progenitor cell products from iPSCs suitable for use in the methods described herein are disclosed in Goldman and Wang's WO2014 / 124087 and Wang et al., “Human iPSC-Derived Oligodendrocyte Progenitors Can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12(2):252-264 (2013), which are hereby incorporated by reference in their entirety.

[0268] In another embodiment, human glial progenitors are derived from embryonic stem cells. Human embryonic stem cells provide an almost limitless source of clonally / genetically modified cells that can potentially be used in tissue replacement therapies. Methods for obtaining highly enriched glial progenitor cell products from embryonic cells suitable for use in the methods of the present disclosure are described in Wang et al., “Human iPSC-derived oligodendrocyte progenitor cells can myelinate and rescue a mouse model of congenital hypomyelination,” Cell Stem Cell 12:252-264 (2013) (which is hereby incorporated by reference in its entirety).

[0269] In another embodiment, the human glial progenitor cells are derived from human fetal tissue. The glial progenitor cells can be directly isolated from fetal brain tissue containing a mixed cell population by using a promoter-specific isolation technique as described in U.S. Patent Application Publication Nos. 20040029269 and 20030223972 to Goldman, which are hereby incorporated by reference in their entirety. This method involves selecting a promoter that functions specifically in glial progenitor cells and introducing a nucleic acid encoding a marker protein under the control of the promoter into the mixed cell population. The mixed cell population is made to express the marker protein, and the cells expressing the marker protein are isolated from the cell population. The isolated cells are glial progenitor cells. The human glial progenitor cells can be isolated from the ventricles or subventricular zone of the brain, or from the subcortical white matter.

[0270] Glial-specific promoters that can be used to isolate glial progenitor cells from a mixed cell population include the CNP promoter (Scherer et al., Neuron 12:1363-75 (1994), which is hereby incorporated by reference in its entirety), the NCAM promoter (Holst et al., J. Biol. Chem. 269:22245-52 (1994), which is hereby incorporated by reference in its entirety), the myelin basic protein promoter (Wrabetz et al., J. Neurosci. Res. 36:455-71 (1993), which is hereby incorporated by reference in its entirety), the JC virus minimal core promoter (Krebs et al., J. Virol. 69:2434-42 (1995), which is hereby incorporated by reference in its entirety), the myelin-associated glycoprotein promoter (Laszkiewicz et al., “Structural Characterization of Myelin-associated Glycoprotein Gene Core Promoter,” J. Res. 50(6):928-36 (1997), which is hereby incorporated by reference in its entirety), or the proteolipid protein promoter (Cook et al., “Regulation of Rodent Myelin Proteolipid Protein Gene Expression,” Neurosci. Lett. 137(1):56-60 (1992); Wight et al., “Regulation of Murine Myelin Proteolipid Protein Gene Expression,” J. Neurosci. Res. 50(6):917-27 (1997); and Cambi et al., Neurochem. Res. 19:1055-60 (1994), which is hereby incorporated by reference in its entirety). See also, U.S. Patent No. 6,245,564 to Goldman et al., which is hereby incorporated by reference in its entirety.

[0271] Glial progenitor cell populations derived from fetal tissue can be enriched by first removing neurons or neural progenitor cells from a mixed cell population. When separating neuronal progenitor cells from a mixed cell population, they can be removed based on their surface expression of NCAM, PSA-NCAM, or any other surface moiety specific for neurons or neural progenitor cells. Neurons or neural progenitor cells can also be separated from a mixed cell population using promoter-based separation techniques. Neuronal or neural progenitor cell-specific promoters useful for separating nerve cells from a mixed cell population include the Tα1 tubulin promoter (Gloster et al., J. Neurosci. 14:7319-30 (1994), which is hereby incorporated by reference in its entirety), the Hu promoter (Park et al., “Analysis of Upstream Elements in the HuC Promoter Leads to the Establishment of Transgenic Zebrafish with Fluorescent Neurons,” Dev. Biol. 227(2):279-93 (2000), which is hereby incorporated by reference in its entirety), the ELAV promoter (Yao et al., “Neural Specificity of ELAV Expression: Defining a Drosophila Promoter for Directing Expression to the Nervous System,” J. Neurochem. 63(1):41-51 (1994), which is hereby incorporated by reference in its entirety), the MAP-1B promoter (Liu et al., Gene 171:307-08 (1996), which is hereby incorporated by reference in its entirety), or the GAP-43 promoter. Techniques for introducing nucleic acid molecules of a construct into a plurality of cells and then sorting the cells are described in U.S. Patent No. 6,245,564 to Goldman et al. and U.S. Patent Application Publication No. 20040029269 to Goldman et al., the patents and patent application publications being hereby incorporated by reference.

[0272] As an alternative to using promoter-based cell sorting to recover glial progenitor cells from a mixed cell population, an immunoisolation procedure can be used. In positive immunoisolation techniques, the desired cells (i.e., glial progenitor cells) are isolated based on surface markers that are properties of proteins that naturally occur on the progenitor cells. For example, the surface marker A2B5 is an early marker of glial progenitor cells that are initially expressed (Nunes et al., “Identification and Isolation of Multipotential Neural Progenitor Cells from the Adult Human White Matter,” Soc. Neurosci. Abstr. (2001), which is hereby incorporated by reference in its entirety). Using an antibody specific for A2B5, glial progenitor cells can be isolated from a mixed population of cell types. Similarly, the surface marker CD44 identifies astrocyte-biased glial progenitor cells (Liu et al., “CD44 Expression Identifies Astrocyte-Restricted Precursor Cells,” Dev. Biol. 276:31-46 (2004), which is hereby incorporated by reference in its entirety). Using a bead technology conjugated with CD44, astrocyte-biased glial progenitor cells can be isolated from a mixed population of cell types. Oligodendrocyte-biased glial progenitor cells can be isolated from a mixed population of cell types based on the expression of PDGFαR, the extracellular domain of PDGFαR CD140a, or CD9. Cells expressing markers of non-glial cell types (e.g., neurons, inflammatory cells, etc.) can be removed from the glial cell preparation to further enrich the preparation for the desired glial cell type using immunoisolation techniques. For example, a glial progenitor cell population is preferably negative for the PSA-NCAM marker and / or other markers of neuronal lineage cells, negative for one or more inflammatory cell markers, e.g., negative for the CD11 marker, negative for the CD32 marker, and / or negative for the CD36 marker, which is a microglial cell marker. Exemplary bead technologies include beads, columns, and Separator. Additional examples of immune isolation are described in the following references: Wang et al., "Prospective Identification, Direct Isolation, and Expression Profiling of a Telomerase Expressing Subpopulation of Human Neural Stem Cells, Using Sox2 Enhancer-Directed FACS," J. Neurosci. 30:14635-14648 (2010); Keyoung et al., "High-Yield Selection and Extraction of Two Promoter-Defined Phenotypes of Neural Stem Cells from the Fetal Human Brain," Nat. Biotechnol. 19:843-850 (2001); and Windrem et al., "Neonatal Chimerization with Human Glial Progenitor Cells can both Remyelinate and Rescue the Otherwise Lethally Hypomyelinated Shiverer Mouse," Cell Stem Cell 2:553-565 (2008), which are hereby incorporated by reference in their entirety.

[0273] According to the methods described herein, a selected administered product of human glial progenitor cells comprises at least about 80% glial progenitor cells, including, for example, about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 100% glial progenitor cells. The selected product of glial progenitor cells can be relatively devoid (e.g., contain less than 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1%) of other cell types, such as neurons or neuronal lineage cells, fibrous astrocytes and fibrous astrocyte lineage cells, and pluripotent stem cells (such as ES cells). Optionally, an exemplary cell population is a substantially pure population of glial progenitor cells.

[0274] The glial progenitor cells of the administered product can optionally be genetically modified to express other target proteins. For example, the glial progenitor cells can be modified to express a therapeutic biomolecule, an exogenous targeting moiety, an exogenous marker (e.g., for imaging purposes), etc. The glial progenitor cells of the product can optionally be modified to overexpress an endogenous biomolecule, targeting moiety, and / or marker.

[0275] The glial progenitor cells of the administered composition can be astrocyte-biased glial progenitor cells, oligodendrocyte-biased glial progenitor cells, unbiased glial progenitor cells, or a combination thereof. The glial progenitor cells of the administered composition express one or more markers of the glial cell lineage. For example, in one embodiment, the glial progenitor cells of the administered composition can express A2B5 + . In another embodiment, the glial progenitor cells of the administered composition are positive for the PDGFαR marker. The PDGFαR marker is optionally the extracellular domain of PDGFαR, such as CD140a. PDGFαR and CD140a are markers of oligodendrocyte-biased glial progenitor cells. In another embodiment, the glial progenitor cells of the administered composition are CD44 + . CD44 is a marker of astrocyte-biased glial progenitor cells. In another embodiment, the glial progenitor cells of the administered composition are positive for the CD9 marker. The CD9 marker is optionally the extracellular domain of CD9. In one embodiment, the glial progenitor cells of the composition are A2B5 + , D140a + and / or CD44 + . The foregoing glial progenitor cell surface markers can be used to identify, isolate, and / or enrich compositions of glial progenitor cells prior to administration.

[0276] Optionally, the administered composition of glial progenitor cells is negative for the PSA-NCAM marker and / or other neuronal lineage markers, and / or negative for one or more inflammatory cell markers, e.g., negative for the CD11 marker, negative for the CD32 marker, and / or negative for the CD36 marker (the CD36 marker is a marker of microglia). Optionally, the composition of glial progenitor cells is negative for any combination or subset of these additional markers. Thus, for example, the composition of glial progenitor cells is negative for any one, two, three, or four of these additional markers.

[0277] Suitable methods for introducing cells into the striatum, forebrain, brainstem, and / or cerebellum of a subject are well known to those of skill in the art and include, but are not limited to, injection, deposition, and implantation as described herein.

[0278] In one embodiment, glial progenitor cells are bilaterally transplanted into multiple sites of a subject as described in Goldman, Windrem et al., U.S. Patent No. 7,524,491, “Neonatal Chimerization With Human Glial Progenitor Cells Can Both Remyelinate and Rescue the Otherwise Lethally Hypomyelinated Shiverer Mouse,” Cell Stem Cell 2:553-565 (2008), Han et al., “Forebrain Engraftment by Human Glial Progenitor Cells Enhances Synaptic Plasticity and Learning Adult Mice,” Cell Stem Cell 12:342-353 (2013), and Wang et al., “Human iPSCs-Derived Oligodendrocyte Progenitor Cells Can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12:252-264 (2013), which are hereby incorporated by reference in their entireties). Methods for transplanting neural tissue and cells into the host brain are described in Bjorklund and Stenevi (eds.), Neural Grafting in the Mammalian CNS, Ch. 3-8, Elsevier, Amsterdam (1985); U.S. Patent No. 5,082,670 to Gage et al.; and U.S. Patent No. 6,497,872 to Weiss et al. (the aforementioned documents are hereby incorporated by reference in their entireties into the present text). Typical procedures include intracerebral, intra-corpus callosum, intraventricular, intrathecal, and intravenous transplantation.

[0279] Intraparenchymal transplantation is achieved by injecting or depositing tissue in the host brain so that it fits (apposed) with the brain parenchyma when transplanted. The two main procedures for intraparenchymal transplantation are: 1) injecting donor cells into the host brain parenchyma, or 2) preparing a cavity by surgical means to expose the host brain parenchyma, and then depositing the implant into the cavity ((Bjorklund and Stenevi (eds.), Neural Grafting in the Mammalian CNS, Ch.3, Elsevier, Amsterdam (1985), which is hereby incorporated by reference in its entirety). Both methods allow for substantial fit between donor cells and host brain tissue when implanted, and both promote anatomical integration between implants and host brain tissue. This is very important if the donor cells are required to become an integral part of the host brain and survive throughout the host's life.

[0280] Glial progenitor cells can also be delivered in the corpus callosum as described in U.S. Patent Application Publication No. 20030223972 of Goldman (which is hereby incorporated by reference in its entirety). Glial progenitor cells can also be delivered directly to the lower forebrain cortex, particularly in the anterior and posterior anlagen of the corpus callosum. Glial progenitor cells can also be delivered to the cerebellar peduncle white matter to enter the main cerebellum and brainstem bundles. Glial progenitor cells can also be delivered to the spinal cord.

[0281] Alternatively, the cells can be placed in a chamber, such as a cerebral ventricle. Implantation of cells into the chamber can be accomplished by injecting donor cells, or growing the cells in a substrate such as 30% collagen to form a solid tissue plug, which can then be implanted into the chamber to prevent dislocation of the implanted cells. For subdural implantation, cells can be injected around the brain surface after a slit is made in the dura mater.

[0282] Suitable techniques for glial cell delivery are described above.In one embodiment, the glial progenitor cell preparation is administered to the striatum, forebrain, brainstem and / or cerebellum of a subject.

[0283] Delivery of cells to a subject can include single-step or multi-step injection directly into the nervous system. Although adult and fetal oligodendrocyte progenitor cells are widely distributed in the brains of transplant recipients, for a wide range of disorders, multiple injection sites can be injected to optimize treatment. Optionally, direct injection into regions of the central nervous system, such as white matter tracts such as the corpus callosum (e.g., injection into the anterior and posterior primordia), dorsal column, cerebellar peduncle, cerebral peduncle. Such injections can be performed unilaterally or bilaterally using precise localization methods (such as stereotactic surgery), optionally utilizing accompanying imaging methods (e.g., high-resolution MRI imaging). Those skilled in the art recognize that brain regions vary by species; however, those skilled in the art also recognize that brain regions are comparable across mammalian species.

[0284] The cell graft is optionally injected as dissociated cells, but can also be provided by local placement of non-dissociated cells. In either case, the cell graft optionally contains an acceptable solution. Such acceptable solutions include solutions that avoid unwanted bioactivity and contamination. Suitable solutions include pharmaceutically acceptable salts in appropriate amounts to render the formulation isotonic. Examples of pharmaceutically acceptable solutions include, but are not limited to, saline, Ringer's solution, dextrose solution, and culture medium. The pH of the solution is preferably from about 5 to about 8, more preferably from about 7 to about 7.5.

[0285] Injection of dissociated cell grafts can be by streaming injection through the entry path, exit path, or both the entry and exit paths of an injection device (e.g., cannula, needle, or tube). Automation can be utilized to provide uniform entry and exit speeds as well as injection speed and volume.

[0286] Depending on the size and species of the recipient and the volume of tissue requiring cell replacement, the number of glial progenitor cells administered to the subject can be in the range of about 10 2 -10 8 per administration (e.g., injection site). A single administration (e.g., injection) dose can span from 10 3 -10 5 cells, 10 4 -10 7 cells, and 10 5 -10 8 cells, or any total amount for a graft recipient patient.

[0287] Because the CNS is an immunologically privileged site, administered cells, including xenogeneic cells, can survive and, optionally, immunosuppressive drugs or typical immunosuppressive regimens are not used in the treatment method. Optionally, however, immunosuppressants can also be administered to the subject. Immunosuppressants and their dosing regimens are known to those skilled in the art and include agents such as Azathioprine, Sodium Azathioprine, Cyclosporine, Daltroban, Gusperimus Trihydrochloride, Sirolimus, and Tacrolimus. The dosage range and duration of the regimens can vary depending on: the condition being treated; the degree of rejection; the activity of the particular immunosuppressant employed; the age, weight, general health, sex, and diet of the subject; the time of administration; the route of administration; the excretion rate of the particular immunosuppressant employed; the duration and frequency of treatment; and drugs used in combination. Those skilled in the art can determine the acceptable dosage and duration of immunosuppression. In the event of any contraindications or changes in the subject's condition, the individual physician can adjust the dosage regimen.

[0288] Examples

[0289] The following examples are intended to illustrate the practice of the embodiments of the present disclosure but are in no way intended to limit its scope.

[0290] Materials and Methods for Examples

[0291] Production of GPCs from human embryonic stem cells (hESCs). GPCs were generated from human embryonic stem cells (ESCs) using a previously described protocol (Wang et al., “Human iPSC-derived Oligodendrocyte Progenitor Cells can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12:252-264 (2013); Windrem et al., “Human iPSC Glial Mouse Chimeras Reveal Glial Contributions to Schizophrenia,” Cell Stem Cell 21:195-208 (2017), which are hereby incorporated by reference in their entirety), which was outlined in great detail in Wang et al., “Human iPSC-derived Oligodendrocyte Progenitor Cells can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12:252-264 (2013) (which is hereby incorporated by reference in its entirety). Cells were harvested between 160 and 240, at which point most cells typically expressed the bipotential GPC marker CD140a, while the remaining cells were mainly composed of A2B5 + / CD140a - immature astrocytes. Cells expressing SSEA4 were not detected. Human ES cells were obtained from GENEA, Inc. (Sydney, Australia) in the form of: GENEA02 line and GENEA19 line (normal HTT: 15 / 18 CAG) and GENEA 17, 18 and 20 (mHTT: 40 / 12, 46 / 17 and 48 / 17 CAG, respectively) (Bradley et al., “Derivation of Huntington's Disease-Affected Human Embryonic Stem Cell Lines,” Stem Cells Dev 20:495-502 (2011), which are hereby incorporated by reference in their entirety). GENEA02 and 17 are male, while GENEA18, 19 and 20 are female. Notably, GENEA 19 and 20 were donated and derived as a pair of female siblings (one normal and one with HD). The C27 control line is male.

[0292] Host. Homozygous shiverer mice (The Jackson Laboratory, Bar Harbor, ME) were crossed with homozygous rag2 gene knockout (null) immunodeficient mice (Shinkai et al., “RAG-2-deficient Mice Lack Mature Lymphocytes Owing to Inability to Inititate V(D)J Rearrangement,” Cell 68:855-867 (1992), which is hereby incorporated by reference in its entirety) on a C3h background (Taconic, Germantown, NY, USA) to generate shi / shi x rag2 - / - Myelin-deficient immunodeficient mice (Windrem et al., “Neonatal Chimerization with Human Glial Progenitor Cells Can Both Remyelinate and Rescue the Otherwise Lethally Hypomyelinated Shiverer Mouse,” Cell Stem Cell 2:553-565 (2008), which is hereby incorporated by reference in its entirety). Mice were maintained on a 12:12 hour light cycle in a temperature- and humidity-controlled chamber (64-79°F; 30%–70% humidity) placed in a pathogen-free housing room. They had free access to Mod Lab Diet 5P00 containing 0.025% trimethoprim / 0.124% sulfamethoxyzole and autoclaved acidic water (pH 2.5-3.0).

[0293] Centrifuge the suspension of single cells or small clusters of hESC-derived GPCs to a density of 100,000 cells / ml. Anesthetize neonatal animals by cooling and transplant a total of 200,000 cells bilaterally into the corpus callosum of the neonatal animals as described in (Windrem et al., “Fetal and Adult Human Oligodendrocyte Progenitor Cell Isolates Myelinate the Congenitally Dysmyelinated Brain,” Nat. Med. 10:93-97 (2004), which is hereby incorporated by reference in its entirety). At 8 weeks of age, 12-13 weeks of age, or 18 weeks of age, anesthetize the transplanted mice with pentobarbital, then perfuse-fix with cold HBSS followed by perfusion-fixation with 4% paraformaldehyde. Remove the brain and post-fix in cold paraformaldehyde for 2 hours.

[0294] All procedures were approved by the University of Rochester's University Committee on Animal Resources (UCAR) under protocol 2004-129.

[0295] Cell products for transplantation. Prior to injection, perform flow cytometry to confirm the predominance of CD140a in each culture. Then collect the suspended cell clusters from the wells, centrifuge, and resuspend them in a small volume of Ca 2+ / Mg 2+ -free HBSS. Transfer the resuspended cell clusters to a 100 mm cell culture dish and then cut them with a No. 11 scalpel to obtain fragments with a diameter of 100-200 mm. Then collect these fragments, centrifuge, wash them with Ca 2+ / Mg 2+ -free HBSS, and resuspend them in Ca2+ / Mg2+-free HBSS to a concentration of approximately 10 5 cells / ml.

[0296] Transplantation. Transplantation was performed on neonatal Trembler x Rag2 knockout mice on the 1st or 2nd day after birth. Half of a litter of pups was removed from the dam and placed in a humidified heated chamber. For this, a sterile plastic box lined with sterile gauze moistened with Hank's balanced salt solution and heated on a heating block was used. The pups to be injected were then wiped with povidone iodine and wrapped in sterile gauze to prevent direct contact with ice, and then cryoanesthetized for 2 to 6 minutes according to size. The pups were then removed from the ice, cleaned with an alcohol prep pad, and then placed in a custom-made neonatal mouse holder made of baked molding clay. The pups were injected directly into the rostral (AP +1.0 mm; ML ±1.0 mm, ventral 1.0 mm) and caudal (AP -1.0, ML ±1.0 mm, ventral 0.9 mm) corpus callosum through the skin and skull-like bone. After injection, the pups were cleaned with an alcohol prep pad and returned to the warming chamber for recovery. After recovery, the first half of a litter of pups was returned to the dam, while the second half was placed in the humidified chamber. The pups were weaned between 21 and 28 days and then housed in groups.

[0297] Immunolabeling of tissue sections. Brains were cryopreserved, embedded in OCT (Tissue-Tek OCT, Sakura Finetek, Torrance, CA), and sagittal or coronal sections were made at 20 μm on a cryostat. Human cells were identified with mouse anti-human nucleus (clone 235-1, 1:800) (MAB1281; EMD Millipore, Billerica, MA). Oligodendrocytes were labeled with MBP rat anti-MBP (Ab7349; Abcam, Cambridge, MA) at 1:25, astrocytes were labeled with anti-human specific GFAPF (SMI 21 at 1:1000, Covance, Princeton, NJ), and axons were labeled with mouse anti-neurofilament at 1:5000 (SMI-311) or 1:1000 (SMI-312; Covance, Princeton, NJ). Alexa Fluor secondary antibodies, goat anti-mouse and anti-rat 488, 568, 594, and 647 (Life Technologies, Carlsbad, CA) were used at 1:400.

[0298] Antibodies and dilutions used.

[0299] Table 5 - Key sources

[0300]

[0301]

[0302]

[0303]

[0304]

[0305]

[0306] RNA-seq. First, human glial progenitor cells (hGPCs) that had been evaluated for gene expression were sorted by fluorescence-activated cell sorting according to the cell surface marker CD140a (BD PharMingen) using a FACS Aria IIIu (Becton Dickinson) as described in (Sim et al., “CD140a Identifies a Population of Highly Myelinogenic, Migration-competent and Efficiently Engrafting Human Oligodendrocyte progenitor Cells,” Nat. Biotechnol. 29:934-941 (2011), which is hereby incorporated by reference in its entirety). mRNA was isolated from FACS-sorted PDGFRα-positive GPC lines generated from human embryonic stem cells (ES) from 3 Huntington's disease (HD) patients (designated HD lines 17 [N = 5 independent cell collection products], 18 [N = 5], and 20 [N = 6]) and 2 healthy controls (designated CTR lines 02 [N = 6] and 19 [N = 6], siblings of HD20) by a polyA-selection protocol. Sequencing libraries were prepared using the Illumina TruSeq RNA v2 kit and sequenced on an Illumina HiSeq 2500 sequencer, generating approximately 45 million 100-bp single-end reads per sample for all cell lines except the control CTR02 line, which had a similar sequencing depth but was sequenced in a paired-end read mode of 125 bp. The sequencing data were then preprocessed by trimming off adapters and low-quality sequences using Trimmomatic (Bolger et al., “Trimmomatic: a Flexible Trimmer for Illumina Sequence Data,” Bioinformatics 30:2114-2120 (2014), which is hereby incorporated by reference in its entirety). The read quality before and after preprocessing was evaluated using FastQC.The preprocessed reads are then aligned to the RefSeq NCBI reference human genome version GRCh38 (Pruitt et al., “NCBI Reference Sequences (RefSeq): a Curated Non-Redundant Sequence Database of Genomes, Transcripts and Proteins,” Nucleic Acids Res. 35:D61-D65 (2007), which is hereby incorporated by reference in its entirety) using the Subread read aligner (Liao et al., “The Subread Aligner: Fast, Accurate and Scalable Read Mapping by Seed-and-Vote,” Nucleic Acids Res. 41:e108 (2013), which is hereby incorporated by reference in its entirety). Raw gene counts are obtained from the BAM alignment file using featureCounts (Liao et al., “featureCounts: an Efficient General Purpose Program for Assigning Sequence Reads to Genomic Features,” Bioinformatics 30:923-930 (2014), which is hereby incorporated by reference in its entirety).

[0307] SOX10 / MYRF rescue of myelination. For this set of experiments, the SOX10 and MYRF transcripts were cloned in two separate lentiviral vectors: pTANK-TRE-MYRF-CAG-rtTA3G-WPRE and pTANK-TRE-Sox10-P2A-DC4-WPRE. In this Tet-On system, cell surface expression of the selectable marker CD4 requires expression from both viruses, ensuring co-expression of the MYRF and SOX10 transgenes. Viral particles pseudotyped with vesicular stomatitis virus G glycoprotein were generated, concentrated by ultracentrifugation, and titrated on 293HEK cells. The G20hGPC cultures were infected at 1.0 MOI in glial medium. Cells were washed with HBSS and maintained in glial medium supplemented with 1 mg / ml DOX (Millipore-Sigma, St. Louis, MO) for 4 days. Then, hGPCs were selected for membrane expression of CD4 as described by MACS (Miltenyi, Germany) as in (Windrem et al., “Neonatal Chimerization with Human Glial Progenitor Cells Can Both Remyelinate and Rescue the Otherwise Lethally Hypomyelinated Shiverer Mouse,” Cell Stem Cell 2:553-565 (2008), which is hereby incorporated by reference in its entirety).

[0308] Rescue of oligodendrocyte differentiation in vitro. MACS-isolated CD4+ cells were allowed to adhere overnight in glial medium (Wang et al., “Human iPSC-derived Oligodendrocyte Progenitor Cells can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12:252-264 (2013), which is hereby incorporated by reference in its entirety). DOX was maintained in the overexpressed state throughout the differentiation induction. The next day, the cells were washed once in HBSS and switched to differentiation medium (Neurobasal medium (GIBCO), 1x N2 (Thermo Fisher), 1x B27 (Thermo Fisher), 1x GlutaMAX (Thermo Fisher), 20 ng / ml BDNF (R&D Systems), 0.2 mM L-ascorbic acid (Sigma), 60 ng / ml T3 (Sigma), 0.2 mM dibutyryl cAMP (Sigma), 100 ng / ml biotin (Sigma), 1x insulin-transferrin-selenium (Thermo Fisher), 10 ng / ml NT3 (R&D), and 100 ng / ml IGF1 (R&D)). The medium was changed every other day before fixation for 2 weeks. Oligodendrocyte differentiation was quantified by O4 immunostaining.

[0309] Rescue of oligodendrocyte differentiation in vitro. Cells for transplantation were prepared and then injected into the corpus callosum of neonatal jimpy mice at two sites unilaterally. Starting at 9 weeks of age, half of the transplanted mice were administered DOX (2 mg / ml, in water containing 5% sucrose (Chow et al., “A Doxycycline-Inducible, Tissue-Specific Aromatase-Expressing Transgenic Mouse,” Transgenic Res. 21:415-428 (2012), which is hereby incorporated by reference in its entirety)) or normal drinking water in their water bottles for 5 weeks. Then, at 13 weeks of age, control and experimental mice were sacrificed and prepared for immunolabeling against MBP and human nuclear antigen, and axonal ensheathment by MBP-expressing oligodendrocytes was evaluated by confocal imaging as described above.

[0310] Imaging and quantitative histology. To map the distribution of human cell nuclei, sections were imaged on a Nikon Instruments Ni-E equipped with a Nikon Digital Sight camera DS-Fi1 and counted and scored in Nikon NIS Elements v4.5. To image the distribution of myelin at low power, whole brain sections were imaged on a Leica LMD 6500. Higher power confocal images of myelin sheaths were obtained using a Nikon C2+ confocal and images were acquired using a 100x objective with a 0.2 mm step size. Cell type-specific markers were imaged on an Olympus BX51 using a Hammamatsu camera driven by Stereo Investigator software (MBF, Williston, VT). Higher magnification confocal stacks of astrocytes subjected to Sholl analysis were obtained using a Leica SP8 confocal.

[0311] Cell counting. Quantification of donor cell density in the corpus callosum was based on counts 1 mm lateral to the midline. Randomly initiated, evenly sampled brain coronal sections were labeled for human cell nuclei, DAPI, and other phenotype-specific markers (Olig2, hGFAP, TF, and MBP). For Olig2 and hGFAP quantification, the target area of each section was imaged at 40x on an Olympus BX51 equipped with a Hamamatsu camera. Z-stacks were obtained at 1 mm steps. For TF and MBP quantification, the target area was imaged at 20x on a Nikon Ni-E Eclipse microscope equipped with a DS-Fi1 camera. Z-stacks were obtained at 0.7 - 1 mm steps. In Nikon NIS Elements v.4.5, immunolabeled cells were counted using high-intensity projections of z-stack images on three evenly spaced coronal sections per mouse.

[0312] Morphometric analysis of astrocytes. Trembler x rag2 knockout mice were sacrificed at 18 weeks of age, and their white matter astrocyte morphology was evaluated. Coronal sections 150 μm thick were obtained from the bregma -1.0 mm of mice implanted with control (GENEA19) or HD (GENEA20) hGPC using a Vibratome, and the sections were incubated in mouse anti - hGFAP at 4 °C for 1 week, then incubated in Alexa 568 goat anti - mouse antiserum for 4 hours. The sections were mounted on glass slides and imaged at 100× magnification by confocal microscopy (Leica SP8). Images were traced using Neurolucida 360 (MicroBright Field, Inc.); all tracings were done by an experimenter blinded to the treatment conditions.

[0313] Individual astrocytes were selected at mid - depth from the middle of the corpus callosum to completely capture the cell and its processes. Three cells / section and three sections / brain were taken 500 μm, 1000 μm, and 1500 μm lateral to the midline, and the cells were analyzed by Sholl analysis using Neurolucida. A total of 14 brains that had undergone neonatal implantation were evaluated (GENEA18, n = 21 cells / 3 brains; GENEA19, 32 cells / 4 brains; GENEA20, 42 cells / 7 brains), yielding 63 traced mHTT astrocytes (from GENEA18 - and GENEA20 - derived) and 32 control (GENEA19) astrocytes. For Sholl analysis, concentric shells with diameters increasing by 5 μm in succession were placed at the center of the cell body, and the number of intersections between the cell processes and the shells was counted (Sholl, “Dendritic Organization in the Neurons of the Visual and Motor Cortices of the Cat,” J. Anat. 87:387 - 406 (1953), which is hereby incorporated by reference in its entirety). For the evaluation and quantitative description of the 3D structure of astrocytic fibers, fan - in analysis (MBF Biosciences) was used as previously described for dendritic topology studies (Dang et al., “Formoterol, a Long - Acting β2 Adrenergic Agonist, Improves Cogntive Function and Promotes Dendritic Complexity in a Mouse Model of Down Syndrome,” Biol. Psychiatry 75:179 - 188 (2014), which is hereby incorporated by reference in its entirety).

[0314] Myelin brightness analysis. To measure forebrain myelination, a brightness analysis based on MBP immunofluorescence measurement was used. MBP staining was performed on evenly spaced and evenly sampled coronal sections as described, and images were taken at 10x magnification using a Nikon Ni-E and a Nikon DS-Fi1 camera. The corpus callosum was selected as the target area, and the average intensity value was obtained using NIS Elements v.4.5.

[0315] Statistical analysis of histological data. All analyses were performed using v.7 (GraphPad software) with two-way ANOVA and Bonferroni post hoc t-tests. Statistical significance was considered when the P-value was less than 0.05. Significance is expressed as *p < 0.05, **p < 0.01, and ***p < 0.001. Graphs and figures were made and assembled using Prism 7, and all data are shown as the mean (SEM) ± standard error of the mean.

[0316] Bioinformatics. After examining the principal component and hierarchical clustering plots generated using native R functions (R Core Team, “R: a Language and Environment for Statistical Computing,” R Foundation for Statistical Computing (2014), which is hereby incorporated by reference in its entirety), one mis-clustered outlier sample was removed from the analysis of the HD17 line (GENEA17), and 2 outliers were also removed from the HD20 line (GENEA20) and the CTR19 line (GENEA19). After eliminating low-expressed transcripts, those transcripts with at least 5 read counts in more than 3 samples were retained, and the count data was normalized using RUVSeq (Risso et al., “Normalization of RNA-seq Data Using Factor Analysis of Control Genes or Samples,” Nat. Biotechnol. 32:896-902 (2014), which is hereby incorporated by reference in its entirety). The R Bioconductor package (Gentleman et al., “Bioconductor: Open Software Development for Computational Biology and Bioinformatics,” Genome Biol. 5:R80 (2004), which is hereby incorporated by reference in its entirety)) was used to illustrate the differences.As described in the RUVSeq documentation, normalization is accomplished by the following three-step procedure: 1) Determine negative in silico control genes by performing first-pass differential expression analysis using the edgeR (Robinson et al., “edgeR: a Bioconductor Package for Differential Expression Analysis of Digital Gene Expression Data,” Bioinformatics 26:139-140 (2010), which is hereby incorporated by reference in its entirety) and DESeq2 (Love et al., “Moderated Estimation of Fold Change and Dispersion for RNA-seq Data with DESeq2,” Genome Biol. 15:550 (2014), which is hereby incorporated by reference in its entirety) R Bioconductor packages, including those genes with an FDR-adjusted p-value > 0.75 as calculated by both methods; 2) Then use the negative in silico control genes in the RUVg function of RUVSeq to calculate the variance factor; and 3) Use the original raw counts and adjust the variance factor calculated by RUVg using the multi-factor GLM model implemented in the edgeR and DESeq2 packages to perform second-pass differential expression analysis (1% FDR and log2 fold change > 1) to identify disease dysregulated genes.

[0317] This three-step analysis, which filters out low and non-expressed transcripts, was used to compare each HD-derived hGPC cell line to pooled CTR-derived HGPC, as well as for the sibling pair comparison of HD20 vs HD19. In all comparisons, one variance factor calculated by RUVg was used. The intersection of the four resulting differentially expressed gene lists was considered a conservative representative list of HD-dysregulated genes. To obtain the average FC and p-values of dysregulated genes in all three HD-derived GPC lines, differential expression comparisons were performed on the pooled HD lines and the pooled CTR lines using the same workflow and the same number of variance factors.

[0318] For all differential expression comparisons, only significant results that were concordant between edgeR and DESeq2 were used in downstream analyses. Fold changes and FDR-adjusted p-values reported in the results were calculated by edgeR. Functional annotation of a set of conserved HD dysregulated genes was performed using TopCLuster (Kaimal et al., “ToppCluster: a Multiple Gene List Feature Analyzer for Comparative Enrichment Clustering and Network-based Dissection of Biological Systems,” Nucleic Acids Res. 38:W96-W102 (2010), which is hereby incorporated by reference in its entirety) and Ingenuity Pathway Analysis (IPA) (QIAGEN) (Kramer et al., “Causal Analysis Approaches in Ingenuity Pathway Analysis,” Bioinformatics 30:523-530 (2014), which is hereby incorporated by reference in its entirety).

[0319] TaqMan RT-qPCR arrays were used for gene expression validation. Total RNA extracted was amplified using ribo-SPIA-based whole transcriptome amplification (NuGen). Expression of cell type markers and pathway-specific genes was evaluated by real-time polymerase chain reaction (RT-PCR) using a 48-gene Taqman low density array (TLDA) (Applied Biosystems). Relative abundance of transcript expression was calculated by ΔΔCt analysis, and expression data were normalized against the mean of 18S and GAPDH used as endogenous controls. Differences in expression in HD and control GPC were evaluated by paired t-test and subsequently by multiple testing correction using the Benjamini-Hochberg (Bh) procedure (Benjamini and Hochberg, “Controlling the False Discovery Rate: a Practical and Powerful Approach to Multiple Testing,” J. R. Stat. Soc. Series B Stat. Methodol. 57:289-300 (1995), which is hereby incorporated by reference in its entirety). Analysis of TLDA data was performed in ExpressionSuite software version 1.1 provided by Applied Biosciences.

[0320] SOX10 / MYRF rescue of myelination gene expression. hGPCs derived from mHTT and normal sibling hESCs were transfected with a plasmid expressing tandem SOX10 and MYRF under the regulatory control of the constitutive promoter EF1α (pTANK-EF1α-Sox10-P2A-Myrf–T2A-EGFP-WPRE), or a control plasmid expressing only EGFP (pTANK-EF1α-EGFP-WPRE). Transfection was performed in P3 buffer using the CA205 transfection program with a nucleofector (Lonza, Germany) according to the manufacturer's protocol. Cells were harvested 72 hours after transfection for RT-qPCR of potential SOX10 and MYRF target genes. RNA was extracted using the Qiagen RNeasy Micro kit (Qiagen, Germany). First-strand cDNA was synthesized using TaqMan reverse transcription reagents (Applied Biosystems). 5 ng RNA input was used per reaction; these were performed using the FastStart Universal SybrGreen premix (Roche Diagnostics, Germany) on a real-time PCR instrument (CFX Connect Real-Time System thermocycler; Bio-Rad, USA). Samples of hGPCs from G19- and G20- were assayed in triplicate for each target gene assayed (primers are available in Table 6 below).

[0321] Table 6 - Primers for real-time PCR

[0322]

[0323]

[0324] Melting curve analysis was performed after each PCR to confirm the specificity of the reaction and identify the target peaks in all samples. Results were normalized against the expression level of 18S from the same sample.

[0325] Data and software availability. All raw RNA-seq data have been deposited in GEO under accession number GEO:GSE105041. The complete reproducible workflow, including R scripts and count matrices, has also been deposited. All differential expression data have been uploaded to a publicly accessible interactive laboratory website where interested users can perform further evaluation and query of the differentially expressed gene sets. All data have also been uploaded to Mendeley Data.

[0326] Network visualization and analysis. The TopCLuster annotation tool was used because it is capable of representing term-to-gene associations as networks (Kaimal et al., “ToppCluster: a Multiple GeneList Feature Analyzer for Comparative Enrichment Clustering and Network-Based Dissection of Biological Systems,” Nucleic Acids Res. 38:W96-W102 (2010), which is hereby incorporated by reference in its entirety). The annotation results were exported via ToppCluster's network generator as a list of term-to-gene associations representing the network edges. For all subsequent network visualization and analysis, the term-to-gene association network was input into Gephi graph visualization software (Jacomy et al., “ForceAtlas2, a Continuous Graph Layout Algorithm for Handy Network Visualization Designed for the Gephi Software,” PLoS ONE 9:e98679 (2014), which is hereby incorporated by reference in its entirety). Basic node centrality metrics and node degrees were calculated, and the network was arranged using the Force Atlas layout with default parameters. For the CD140a-derived annotation network, randomization and resolution parameters of 1.3 were used, and for the CD44-derived annotation network, randomization and resolution parameters of 2.0 were used. Tightly interconnected node modules were identified using the built-in community detection algorithm (Blondel et al., “Fast Unfolding of Communities in Large Networks,” arXiv arXiv:0803.0476 (2008), which is hereby incorporated by reference in its entirety) to optimize the grouping and number of communities.

[0327] Example 1 - Transcriptional Determinants of mHTT OPC Downregulation of Glial Lineage Progression

[0328] To address the role of glial transcriptional dysregulation in the pathogenesis of HD, the differential gene expression of bipotent hGPCs derived from mutant huntingtin hESCs was evaluated for the first time. To this end, GPCs were prepared and purified from three different hESC lines (GENEA17, GENEA18, and GENEA20; GENEA Biocells) obtained from blastocysts expressing mHTT, as well as two control lines (GENEA02 and GENEA19) (Bradley et al., “Derivation of Huntington's Disease-Affected Human Embryonic Stem Cell Lines,” Stem Cells Dev 20:495-502 (2011), which is hereby incorporated by reference in its entirety). GPCs were prepared from hESCs using a previously described method (Wang et al., “CD133 / CD140a-Based Isolation of Distinct Human Multipotent Neural Progenitor Cells and Oligodendrocyte Progenitor Cells,” Stem Cells and Development 22:2121-2131 (2013), which is hereby incorporated by reference in its entirety), and the resulting GPC fraction was then isolated by CD140a-based FACS (>99% Cd140a + )(Sim et al., “CD140a Identifies a Population of Highly Myelinogenic, Migration-Competent and Efficiently Engrafting Human Oligodendrocyte Progenitor Cells,” Nature Biotechnology 29:934-941 (2011), which is hereby incorporated by reference in its entirety). Importantly, one of the controls (GENEA19; 18CAG) is a sibling of one of the mHTT-expressing lines (GENEA20; 48CAG); these lines, donated by the same parents, are dizygotic female twins.

[0329] After average propagation times of 190 ± 16 days and 174 ± 14 days, respectively, mHTT and control hGPCs were harvested as stably expanded hGPCs. Flow cytometry revealed that 54% ± 3.4% of normal cells (GENEA02 and GENEA19; n = 12 culture runs) and 44% ± 3.3% of Huntington's protein mutant cells (GENEA17, 18, and 20; n = 16) expressed CD140a at these time points (mean ± SEM). Each culture's CD140a fraction was then separated by FACS to near purity and RNA-seq was performed using an Illumina HiSeq 2500 sequencer, which showed severe transcriptional dysregulation in hGPCs derived from three HD lines relative to pooled control hESCGPCs. Principal component analysis (PCA) showed clear separation of hGPCs expressing mHTT and control hGPCs ( Figure 1A ). As a group, using a 2-fold change (FC) cutoff and a 1% false discovery rate (FDR), 239 genes were upregulated and 530 genes were downregulated in mHTT hGPCs relative to controls ( Figure 1B ). To further refine the list of genes with differential expression, the differential expression of GENEA20 (mHTT)-derived hGPCs was then compared to its sibling GENEA19-derived control, and this sibling comparison was added to the overall comparison; this served as an additional filter and yielded a more tightly defined list of genes with differential expression, which consisted of 64 upregulated genes and 365 downregulated genes in hGPCs from all HD-derived hGPC cell lines relative to pooled control hGPCs ( Figure 1B and 1C ).

[0330] Using this gene set, functional analysis was performed using annotations from Gene Ontology (GO), by which we identified 50 significantly associated GO annotation terms (Bonferroni-corrected p < 0.01, in terms of the Biological Process and Cellular Component GO domains), which represented 187 out of 429 differentially regulated genes (Figure 2 and Figure 3A - 3B ). By network analysis, these annotation terms, along with their associated genes, were further grouped into three functionally related modules, each of which was characterized by its most important annotation term ( Figure 1D ). These three modules represent genes and functions related to (1) glial cell differentiation and myelination, (2) axon guidance and axonogenesis, and the regulation of synaptic structure and synaptic signaling ( Figure 1D)。The first and second modules are tightly interconnected and contain a series of key oligodendrocyte lineage transcription factors (including SOX10, SIRT2, MYRF, NKX2.2, TCF7L2, OLIG1, and OLIG2), as well as stage-regulated myelin-associated proteins (including TF, MBP, MAG, OMG, UGT8, and FA2H); all of these are significantly downregulated in HD hGPCs. The third module contains genes related to the regulation of synaptic transmission components, most notably SYNDIG1, BCAN, NETO1, and SNPH, as well as genes encoding glutamate receptor signaling proteins GRIA2, GRIA4, GRID1, GRID2, and GRIK4 and potassium channels encoded by KCND2, KCNJ9, KCNQ1, and KCNS3; all of these are significantly downregulated( Figure 1E - Figure 1G )。Collectively, these HD dysregulated genes and their associated functions reveal that the differentiation of hGPCs into mature oligodendrocytes is suppressed in an HD-dependent manner.

[0331] Example 2 - mHTT HGPCs Downregulate Transcriptional Determinants of Myelination

[0332] As revealed by differential expression analysis, a set of key transcription factors associated with oligodendrocyte differentiation and myelin biosynthesis are significantly and substantially downregulated as a function of mHTT expression. These include the early oligodendrocyte regulators NKX2.2, OLIG2, and SOX10, each of which is acutely downregulated in hGPCs expressing mHTT( Figure 1E)。In addition, downstream of the oligodendrocyte lineage transcription factors repressed by mHTT, the levels of MYRF (myelin regulatory factor) expressed by mHTT hGPCs are drastically reduced. MYRF co-activates many genes essential for myelination (Bujalka et al., “MYRF is a Membrane-Associated Transcription Factor that Autoproteolytically Cleaves to Directly Activate Myelin Genes,” PLoS Biology 11:e1001625 (2013), which is incorporated herein by reference in its entirety), and the production of MYRF has been found to be lacking in mouse mHTT-transgenic oligodendrocytes (Huang et al., “Mutant Huntingtin Downregulates Myelin Regulatory Factor-Mediated Myelin Gene Expression and Affects Mature Oligodendrocytes,” Neuron 85:1212-1226 (2015), which is incorporated herein by reference in its entirety). In hGPCs derived from human ESCs, the myelinogenesis transcripts MBP, MAG, OMG, PLP1, and MOG regulated by MYRF are all significantly downregulated ( Figure 1E )。In addition, when directly comparing the expression patterns of hGPCs derived from sibling pairs with minimal background genetic variation (GENEA20 for mHTT and GENEA19 for normal HTT), differential downregulation of those genes associated with myelination was again noted in mHTT hGPCs. These include MYRF (-4.04 (fold decrease in mHTT hGPCs); log2 scale), MAG (-6.78), MBP (5.14), MOG (-10.35), OMG (-5.15), and PLP1 (-2.22), indicating widespread downregulation of transcripts associated with myelination in HD hGPCs. Importantly, when comparing the RNA expression patterns of hGPCs derived from three different mHTT hESC lines (GENEA17, GENEA18, and GENEA20, which have 40, 46, and 48 CAG repeats in their HTT genes, respectively), a progressive downregulation of these same genes associated with differentiation and myelination was noted to be closely associated with longer CAG repeat lengths ( Figure 4A - Figure 4C)。Importantly, there is a high degree of overlap between these genes and ontologies, and it was found that as the CAG repeat length in hGPC increases, the degree of dysregulation of these genes and ontologies becomes increasingly high, while in HD transgenic mice, the dysregulation of these genes and ontologies increases with the increase in CAG repeat length (Langfelder et al., “Integrated Genomics and Proteomics Define Huntingtin CAG Length-Dependent Networks in Mice,” Nat. Neurosci. 19:623-633 (2016), which is hereby incorporated by reference in its entirety)( Figure 5A - Figure 5D )。

[0333] To validate these RNA-seq-based expression data, qRT-PCR with TaqMan low density arrays (TLDA) was then used to compare the expression levels of these genes associated with differentiation between mHTT and control hGPC. Most of those genes identified as differentially dysregulated in mHTT hGPC in the RNA-seq analysis were confirmed to be so( Figure 6A - Figure 6B )。These genes include the key oligodendrocyte lineage transcription factors MYRF, SOX10, and OLIG2, as well as their downstream targets associated with myelination, including PLP1, MOG, and MBP. Based on the downregulation of this broad set of genes associated with myelination, it was predicted that mHTT hGPC has significant disruption to both myelin biogenesis and maintenance.

[0334] Example 3 - Suppressed potassium channel expression associated with mHTT results in containment of differentiation

[0335] Among the genes that are functionally related, those encoding ion channels and transporters, particularly potassium channels, are the most differentially dysregulated by mHtt expression. This large group of genes includes 117 known members in the human genome (Pruitt et al., “NCBI Reference Sequences (RefSeq): A Curated Non-Redundant Sequence Database of Genomes, Transcripts and Proteins,” Nucleic Acids Research 35:D61-D65 (2006), which is hereby incorporated by reference in its entirety), of which 93 are detectably expressed by hGPC (raw count > 5 in at least 3 samples within the dataset). Among these, of the 93 identified K +Among the channel and transporter genes, 25 were dysregulated in HD hGPCs relative to their pooled hESC GPC controls (using an FC cutoff of >2.0 and a 5% FDR threshold); 23 of these genes remained significantly dysregulated even at 1% FDR( Figure 7 ). These genes include many inwardly rectifying K + channels, the co-suppression of which revealed a basis for disrupted potassium buffering in the HD brain (Tong et al., “Astrocyte Kir4.1 Ion Channel Deficits Contribute to Neuronal Dysfunction in Huntington's Disease Model Mice,” Nat Neurosci 17:694 - 703 (2014), which is hereby incorporated by reference in its entirety). To further refine and prioritize within this large group of co-dysregulated K + channel genes, the comparison of GENEA20 versus GENEA19 sibling pairs was included as an additional filter. By this most stringent analysis, 4 genes (KCND2, KCNJ9, KCNQ1, and KCNS3) remained strongly and significantly downregulated in all HD hGPC lines relative to the pooled controls and in the sibling sets of mHTT- and normal hESC-derived hGPCs. Collectively, the dysregulated expression of these K + channel genes is of particular significance because they play important roles in maintaining stable interstitial K + levels and determining action potential thresholds. Thus, the suppression of hGPC K + channels associated with mHTT, among other effects, mediates synaptic K +The reuptake of glutamate may be the cause of the neuronal hyperexcitability observed in striatal neurons in HD (Benraiss et al., “Human Glia can Both Induce and Rescue Aspects of Phenotype in Huntington Disease. Nature Communications 7:11758 (2016); Shin et al., “Expression of Mutant Huntingtin in Glial Cells Contributes to Neuronal Excitotoxicity,” J Cell Biol 171:1001-1012 (2005); Tong et al., “Astrocyte Kir4.1 Ion Channel Deficits Contribute to Neuronal Dysfunction in Huntington's Disease Model Mice,” Nat Neurosci 17:694-703 (2014), which are hereby incorporated by reference in their entirety).

[0336] Given the concurrent dysregulation of glial differentiation and K + channel expression and the dependence of the latter on the former, the question to be asked is whether there might be a common upstream regulator that is itself dysregulated as a function of mHTT expression. Using Ingenuity Pathway Analysis (IPA), TCF7L2 was found to be predicted as a positive regulator of a variety of genes associated with glial differentiation, said genes associated with glial differentiation including several genes that have been reported to regulate K + channel gene expression, such as KCNB1 regulated by SOX10 (Liu et al., “Chromatin Landscape Defined by Repressive Histone Methylation During Oligodendrocyte Differentiation,” J Neurosci 35:352-365 (2015), which is hereby incorporated by reference in its entirety), which is downregulated in hGPCs derived from all three HD lines tested. Among these genes associated with glial differentiation, some are significantly lacking in expression in mHTT glia relative to their controls ( Figure 6A ). On this basis, the RNA-seq dataset was queried for TCF7L2 and TCF7L2-regulated transcripts, and it was found that TCF7L2 was indeed differentially downregulated in HD relative to normal hGPCs, and genes regulated by TCF7L2 were concomitantly downregulated (Figure 6B )。Since TCF7L2 is closely implicated in glial differentiation, especially oligodendrocyte differentiation, these results further emphasize the cell-intrinsic nature of the block in glial differentiation in HD.

[0337] Example 4–HD hGPCs exhibit impaired myelination in vivo

[0338] Since mHTT hGPCs appear to be deficient in acquiring gene expression patterns representative of oligodendrocyte maturation and myelination, the question was posed whether hypomyelinated mice implanted with HD GPCs lack the ability to myelinate relative to mice implanted with GPCs from normal siblings. To this end, using the multi-site injection protocol utilizing bilateral hemisphere injections, neonatal hGPCs expressing mHTT and control hGPCs, derived from sibling female GENEA20 and GENEA19 lines in paired cultures respectively, were transplanted into immunodeficient shiverer mice. This protocol produces a stereotyped pattern and time course of donor-derived myelination in the host brain when using normal pluripotent stem cell-derived or tissue-derived hGPCs (Wang et al., “Human iPSC-Derived Oligodendrocyte Progenitor Cells can Myelinate and Rescue a Mouse Model of Congenital Hypomyelination,” Cell Stem Cell 12:252-264 (2013b); Windrem et al., “Neonatal Chimerization with Human Glial Progenitor Cells can Both Remyelinate and Rescue the Otherwise Lethally Hypomyelinated Shiverer Mouse,” Cell Stem Cell 2:553-565 (2008), which are hereby incorporated by reference in their entirety). In this case, although non-isogenic (true isogenic lines of normal and mutant huntingtin have not been reported), the use of sibling lines in this experiment minimized genetic variation as much as possible. Using these paired lines and this in vivo model, the pattern of oligodendrocyte differentiation and myelination was evaluated in implanted mice at 8, 13, and 18 weeks of age (n = 3-5 mice per time point, a total of 12 mice implanted with HD hGPCs and 10 mice implanted with control hGPCs). Brains of these mice were cryosectioned, immunolabeled for oligodendrocyte and myelin antigens, and confocal imaging was performed to compare the differentiation and myelination efficiency of HD- and control-derived hESC hGPCs in vivo.

[0339] It was found that, relative to animals implanted with HD hGPCs, the appearance time of oligodendrocyte phenotype markers and myelin protein production indices was significantly advanced in mice implanted with control hGPCs. However, 8 weeks after neonatal implantation, the expression of myelin basic protein in axonal junctions was significant when using control hGPCs, but mice implanted with HD hGPCs did not show significant MBP immunolabeling at this time point (Figures 8A and 8D). By 12 - 13 weeks of age (by which time mice implanted with control hGPCs showed robust myelin production), only scattered islands of MBP expressed by immature oligodendrocytes appeared in the matched HD GPC recipients (Figures 8B and 8E). The relatively delayed myelination of the white matter implanted with HD GPCs continued for at least 4 months; however, by the 18th week, mice implanted with control GPCs showed dense corpus callosum and capsular myelination, while confluent areas of MBP - defined myelination only appeared in the brains implanted with mHTT (Figures 8C and 8F). Thus, in mice implanted with GENEA19 control GPCs, the differentiation into transferrin + oligodendrocytes( Figure 8H and Figure 8I ) and their derivatives, MBP + myelinated oligodendrocytes( Figure 8J and Figure 8K ) of human donor cells was significantly higher than the fraction of such human donor cells in mice implanted with GENEA20 mHTT GPCs. Similarly, the myelin brightness, as evaluated on MBP immunostained sections, was significantly higher in the corpus callosum of mice implanted with control GPCs at both time points than the myelin brightness in their counterparts implanted with mHTT GPCs( Figure 8L ). Nevertheless, there were no significant differences in the implantation density and distribution of human GPCs between control cells and HD - derived cells( Figure 8G , Figure 8M , Figure 8N ), indicating that the myelination defect in the brains implanted with HD hGPCs was due to a disorder in oligodendrocyte differentiation and myelin production of donor cells associated with mHTT, rather than due to differential implantation.

[0340] The myelin formation delay associated with mHTT had a significant impact on the rate and efficiency of axonal myelination. When analyzing corpus callosum myelination by high - resolution confocal imaging of individual corpus callosum axons, it was evident that axonal ensheathment was impaired in the brains implanted with MHTT - HgPc (Figures 9A - 9F). At the 13 - week and 18 - week time points, mHTT hGPC - chimeric brains showed fewer myelinated axons (Figure 9G); along the length of the visualized axons, a larger proportion of those myelinated axons were indeed so incomplete, and for each identified MBP+ Oligodendrocytes with fewer axons myelinated (Figure 9H). Collectively, these data indicate that chimeric tremor mice engineered to express mHTT in hGPCs failed to myelinate as rapidly or as well as mice implanted with normal hESC hGPCs, generating relatively hypomyelinated animals with insufficient axonal myelination. Thus, the differentiation arrest associated with mHTT, as revealed by the mHTT hGPC expression profile, appears to be reflected in their relative lack of ability to differentiate into oligodendrocytes, resulting in reduced myelination in vivo.

[0341] Example 5 - Myelin Gene Expression and Myelination in Vivo Can Be Rescued by SOX10 and MYRF

[0342] Given the primacy of SOX10 and MYRF in regulating myelination (Bujalka et al., “MYRF is a Membrane-Associated Transcription Factor that Autoproteolytically Cleaves to Directly Activate Myelin Genes,” PLoS Biology 11:e1001625 (2013); Emery et al., “Myelin Gene Regulatory Factor is a Critical Transcriptional Regulator Required for CNS Myelination,” Cell 138:172-185 (2009); Lopez-Anido et al., “Differential Sox10 Genomic Occupancy in Myelinating Glia,” Glia 63:1897-1914 (2015), which are hereby incorporated by reference in their entirety) and their role as terminal effectors of myelin gene expression, the data suggest that transcriptional activation of SOX10 and MYRF may be sufficient to rescue the myelination defect in HD. On this basis, the next question to ask is whether forced expression of SOX10 and MYRF in hGPCs expressing mHTT can rescue the expression of MAG, MBP, and other key genes involved in myelin biosynthesis. To this end, the expression of SOX10 and MYRF was induced in mHTT- and normal control hESC-derived hGPCs (GENEA20 and GENEA19, respectively) by plasmid transfection using a bicistronic plasmid in which both genes were placed under the control of the constitutive EF1a promoter. Their expression was then compared to the expression of downstream myelination genes (including MAG, MBP, MOG, PDGFRA, PLP1, TF, and LINGO1) in SOX10-MYRF- and control plasmid-transfected cells using qPCR. Overexpression of SOX10-MYRF was found to indeed rescue the expression of most myelin-associated genes in transfected mHTT hGPCs (Table 1; Figure 10 ).

[0343] Based on these data, the next question to ask is whether overexpression of SOX10 and MYRF is sufficient to rescue downstream oligodendrocyte differentiation and myelination. To this end, a doxycycline-regulated dual-vector lentiviral transduction strategy was developed that allows doxycycline (DOX)-triggered, interdependent overexpression of SOX10 and MYRF as well as simultaneous expression of CD4 to allow FACS-based immunoisolation of SOX10-MYRF-transduced hGPCs (Figure 11A )。First, the effects of SOX10 and MYRF overexpression in mHTT-expressing hGPCs were evaluated by transducing a collection of matched 180-day in vitro (DIV) GENEA20-derived hGPCs with lentiviral SOX10 / MYRF regulated by DOX, and then exposing some of the cultures to DOX while leaving the matched control cultures untreated. It has been confirmed that in cells grown in the absence of DOX, SOX10 and MYRF expression is indistinguishable from that of untransduced GENEA20-derived hGPCs. After an additional week in vitro, the cells were then immunostained for oligodendrocyte sulfatide recognized by O4, which is lineage-restricted and expressed primarily by postmitotic human oligodendrocytes. In the absence of DOX, mHTT hGPCs remained as such and did not express detectable O4. In contrast, those mHTT hGPCs with upregulated SOX10 and MYRF expression grown in DOX showed a sharp and significant increase in oligodendrocyte differentiation, with over 15% expressing O4 immunoreactivity ( Figure 11B - Figure 11D ).

[0344] Since induction of SOX10 and MYRF expression appears to be sufficient to rescue oligodendrocyte differentiation from mHTT hGPCs in vitro, the next question was whether SOX10 and MYRF expression is similarly sufficient to rescue myelination in vivo. To this end, a vector system expressing SOX10 and MYRF simultaneously reported by CD4 expression was used. GENEA20-derived HD hGPCs were transduced with lentiviral SOX10 / MYRF regulated by DOX as described above, cells were sorted on CD4, and SOX10 / MYRF-transduced mHTT hGPCs were transplanted into neonatal shiverer mice. At 9 weeks of age, some of the transplanted mice were given DOX (orally, introduced into the water they drank ad libitum) to trigger SOX10 and MYRF expression, while other mice were not given DOX and thus served as matched controls (Figure 11E). At 13 weeks of age (the time point when normal hGPCs typically begin myelination, while untreated mHTT hGPCs have not yet begun myelination; Figures 11F and 11G), the mice were sacrificed, their brains were sectioned, and immunostaining for MBP was performed. It was found that in DOX(+) mice in which donor-derived hGPC SOX10 and MYRF were induced, a large amount of MBP was shown in the host white matter subjected to implantation +Myelinating oligodendrocytes. Quantitatively, DOX(+) mice implanted with SOX10 / MYRF-transduced, DOX-regulated GENEA20 GPCs exhibited robust myelination: by 13 weeks, 28.6% ± 0.8% (n = 3 mice; mean ± SEM) of donor cells expressed MBP, whereas no detectable MBP expression was observed in donor cells of DOX(-) mice (n = 6 mice) that underwent the same implantation (p < 0.0001). In contrast, 18.1% ± 2.1% (n = 5) of normal GENEA19-derived GPCs showed MBP expression at the same time point, indicating that SOX10 / MYRF-transduced HD hGPCs are at least as efficient as normal hGPCs in MBP-defined myelination in vivo.

[0345] In DOX(+) mice implanted with SOX10 / MYRF-transduced GENEA20 hGPCs, the resulting oligodendrocytes were shown to be sufficient to induce the formation of nodes of Ranvier on resident trembler axons, which exhibited typical clustering of βIV-spectrin flanked by CASPR1 characteristic of the nodal architecture (Figures 11L and 11M). In contrast, at the same time point, no MBP expression was observed in donor cells in DOX(-) control mice (Figures 11H - 11J), and no distinct nodes were observed despite similar donor cell implantation (Figure 11K). These data suggest that forced expression of SOX10 and MYRF is sufficient to rescue oligodendrocyte differentiation and myelination of hGPCs expressing mHTT.

[0346] Example 7 – mHTT impairs human astrocyte differentiation in vivo

[0347] Since hGPCs give rise to astrocytes and oligodendrocytes, defects associated with mHTT in oligodendrocyte lineage progression, along with RNA expression data suggesting a transcriptional block in glial differentiation upstream of the astrocyte-oligodendrocyte fate choice, reveal a similar block in astrocyte differentiation. On this basis, the next question was whether mice neonatally injected with hGPCs expressing mHTT (GENEA20-derived) would exhibit any differences in astrocyte differentiation in vivo compared to mice injected with normal HTT sibling control hGPCs (GENEA19). To this end, the same mice previously used to examine the effect of the HD genotype on myelination were used to assess its effect on the maturation of glial fibrillary acidic protein (GFAP)-defined white matter astrocytes. At 8, 13, and 18 weeks post-neonatal implantation, the trembler brains implanted with control and HD hGPCs were immunostained with a species-specific anti-human GFAP antibody.

[0348] It has been found that astrocyte maturation from the implanted hGPCs was significantly lacking in the brains (n = 12, across 3 time points) implanted with HD (GENEA20) hGPCs as compared to the counterparts implanted with control (GENEA19) hGPCs (n = 10). Focusing on the white matter compartments of the corpus callosum and internal capsule that underwent the most rapid massive implantation, it was found that astrocyte differentiation defined by GFAP produced by HD hGPCs was significantly reduced relative to control GPCs and remained so up to the 18-week observation point (Figures 12A - 12F). To quantitatively verify this observation, those brains sacrificed at 13 and 18 weeks were scored. At 13 weeks, mice implanted with control hGPCs showed significant GFAP + astrocyte maturation such that 5.9% ± 0.5% of the human donor cells in the corpus callosum expressed GFAP (n = 4 mice; 170 GFAP + cells out of a total of 2,669 scored donor cells); in contrast, in the corpus callosum of mice implanted with mHTT GPCs, only 3.3% ± 0.3% of the human cells were GFAP + cells (n = 5 mice; 60 GFAP + ) among 2,153 scored donor cells (p = 0.026)( Figure 12I ). By 18 weeks, the mHTT-dependent suppression of astrocyte maturation remained significant; by this time point, 8.5% ± 1.0% of the control-derived cells had developed the GFAP + astrocyte phenotype (n = 3 mice; 209 GFAP + among 2,452 scored donor cells), while only 4.9% ± 0.8% of the human donor cells expressing mHTT were so (n = 4 mice; 147 GFAP + ) among 3,522 scored donor cells (p < 0.005)( Figure 12I ). In summary, these data indicate that astrocyte differentiation of hESC GPCs expressing mHTT was significantly delayed relative to normal hESC GPCs (F = 16.31 [1, 16 degrees of freedom (df)], two-way ANOVA; overall p = 0.0009). Thus, one might expect that HD may impair developing circuit integration as well as adult astrocyte function.

[0349] Example 8 - Abnormal Fiber Distribution and Domains in mHTT GPC White Matter Astrogliogenesis

[0350] Given the decreased and delayed astrocyte differentiation noted in mice implanted with mHTT-hGPC, the next question was whether the morphology of those HD astrocytes that did mature was normal, or whether their mature architecture was ultimately different from that of their more rapidly developing control HGPC-derived counterparts. Gross assessment revealed that the mature astrocyte morphology of mHTT-expressing astrocytes differed from that of control astrocytes in that mHTT-expressing HD-derived astrocytes generally did not exhibit the degree of radial symmetry of their control-derived counterparts (Figs. 12G and 12H). To investigate this observation, Sholl analysis was used to assess the complexity of individual astroglial morphologies; Sholl analysis is based on the number of intersections of cellular processes with concentric circles placed successively at greater radii (Sholl, D. A., “Dendritic Organization in the Neurons of the Visual and Motor Cortices of the Cat,” J Anat 87:387-406 (1953), which is hereby incorporated by reference in its entirety). By imaging human GFAP-immunostained cells in a z-stack of 150-mm sections and reconstructing these cells in Neurolucida (MBF Biosciences), the fiber architecture of donor-derived astrocytes in the white matter of mice implanted with two different lines of mHTT hESC hGPC (GENEA18 and GENEA20) was compared to that of donor-derived astrocytes in the white matter of mice implanted with hGPC derived from two control hESC lines (C27iPSC and GENEA19 hESC, the latter being a sibling of GENEA20). Sholl analysis revealed that the fiber complexity of mHTT-expressing astrocytes was significantly reduced relative to that of astrocytes derived from their sibling control hGPC ( Figure 13A - Figure 13D ). This effect was particularly evident in the comparison of mHTT astrocytes derived from GENEA20 hESC with normal astrocytes from their matched GENEA19 siblings ( Figure 12J - Figure 12P ). Human astrocytes in chimeras implanted with mHTT-hGPC were significantly different from those in mice implanted with normal GPC, with lower fiber network complexity ( Figure 12J ) and characterized by fewer but longer processes ( Figure 12K - Figure 12M ). When 3D Neurolucida tracings were additionally evaluated by fan-in radial analysis ( Figure 12O and Figure 12P)When assessing the extent to which the fibrous domain of each cell occupies its immediate volume environment (Dang et al., “Formoterol, A Long-Acting Beta2 Adrenergic Agonist, Improves Cognitive Function and Promotes Dendritic Complexity in a Mouse Model of Down syndrome,” Biol Psychiatry 75:179-188 (2014), which is hereby incorporated by reference in its entirety), it was found that mHTT astrocytes showed significantly more areas not occupied by glial processes compared to control-derived astrocytes( Figure 12N - Figure 12P ), indicating that the domain structure is discontinuous and incomplete.

[0351] To better understand the transcriptional concomitants of these HD-associated morphological abnormalities in astrocyte morphology, the gene expression patterns of HD versus control-derived astrocytes were next evaluated. To this end, CD140a-defined hGPCs were prepared according to a standard protocol and then induced to differentiate into astrocytes by switching the cells to serum-containing medium supplemented with 20 ng / mL BMP4. The cells were then sorted based on CD44, which is differentially expressed in brain cells by astrocytes and their committed progenitor cells (Cai et al., “CD44-Positive Cells are Candidates for Astrocyte Precursor Cells in Developing Mouse Cerebellum,” Cerebellum 11:181-193 (2012); Liu et al., “Chromatin Landscape Defined by Repressive Histone Methylation During Oligodendrocyte Differentiation,” J Neurosci 35:352-365 (2015), which are hereby incorporated by reference in their entirety). RNA-seq analysis was then performed on the extracted RNA of HD and control-derived CD44-defined astrocytes, which was thus confirmed by their virtually identical GFAP expression. This analysis revealed significant differences in gene expression between astrocytes expressing mHTT and control-derived CD44 + astrocytes( Figure 14A - Figure 14C)。Network analysis revealed differential expression of four discrete modules, which included functional ontologies related to (1) synaptic genes, postsynaptic genes, and genes associated with receptors, (2) endosomal transcripts, (3) desmosomes and cell-cell junction genes; and extracellular matrix components( Figure 14D - Figure 14H )。Among them, the largest set of differentially expressed genes was the set of those genes related to synaptic and receptor regulation; these gene sets included many genes that regulate fiber outgrowth and motility, including MYL7 and MYLK2, myosin light chain-7 and myosin light chain kinase-2 (which were both sharply downregulated in astrocytes expressing mHTT relative to controls)( Figure 14E )。Importantly, glial myosin and its kinase are not only involved in the elaboration of glial fibers but also in astrocytic calcium signaling (Cotrina et al., “Cytoskeletal Assembly and ATP Release Regulate Astrocytic Calcium Signaling,” J. Neurosco. 18:8794-8804 (1998), which is hereby incorporated by reference in its entirety). The lack of their expression in HD astrocytes may then lead to abnormal morphological development of HD astrocytes (Khakh et al., “Unravelling and Exploiting Astrocyte Dysfunction in Huntington’s Disease,” Trends Neurosci. 40:422-437 (2017); Octeau et al., “An Optical Neuron-Astrocyte Proximity Assay at Synaptic Distance Scales,” Neuron 98:49-66 (2018), which are hereby incorporated by reference in their entirety), while predicting abnormal signaling within the glial syncytium of the HD brain (Jiang et al., “Dysfunctional Calcium and Glutamate Signaling in Striatal Astrocytes from Huntington’s Disease Model Mice,” J. Neurosci. 36:3453-3470 (2016), which is hereby incorporated by reference in its entirety). Collectively, these data emphasize the association of HD with deficiencies in astrocyte differentiation and functional development and impaired oligodendrocyte maturation and myelination.

[0352] Example Discussion

[0353] These experiments demonstrate that white matter failure in HD is a product of mHTT-dependent blockade in the differentiation of affected hGPCs, such that mRNAs encoding a set of key glial lineage transcription factors are differentially downregulated in hGPCs expressing mHTT. In particular, the mHTT-associated inhibition of oligodendrocyte differentiation, as manifested by the downregulation of the expression of NKX2.2, OLIG2, and SOX10, is accompanied by a decrease in the expression of MYRF, a myelin regulatory factor regulated by SOX10. This results in the suppression of myelination, which requires MYRF-dependent transcription of key mRNAs associated with myelin biogenesis, such as MAG and MBP (Bujalka et al., “MYRF is a Membrane-Associated Transcription Factor that Autoproteolytically Cleaves to Directly Activate Myelin Genes,” PLoS Biology 11:e1001625 (2013); Emery et al., “Myelin Gene Regulatory Factor is a Critical Transcriptional Regulator Required for CNS Myelination,” Cell 138:172-185 (2009), which are hereby incorporated by reference in their entirety). Interestingly, a similar downregulation of MYRF was also noted in mature oligodendrocytes of HD transgenic mice expressing particularly long CAG repeat sequences (150Q and 250Q) (Jin et al., “Early White Matter Abnormalities, Progressive Brain Pathology and Motor Deficits in a Novel Knock-in Mouse Model of Huntington’s Disease,” Hum. Mol. Genet. 24:2508-2527 (2015), which is hereby incorporated by reference). These data suggest that in humans, the mHTT-associated glial cell differentiation blockade occurs at an earlier stage than previously thought and is evident in bipotential hGPCs that give rise to both astrocytes and oligodendrocytes. Thus, mHTT has been found to significantly impede the development of these two glial lineages in HD, and more importantly, this developmental arrest occurs in human GPCs expressing CAG repeat expansion lengths of 40-48Q, which is characteristic of human HD.

[0354] These expression data suggest that the mHTT-dependent repression of NKX2.2, OLIG2, and SOX10 in the white matter deficiency of HD indicates that overexpressing SOX10 and MYRF or otherwise activating the transcription of SOX10 and MYRF may be sufficient to alleviate the myelin formation defects of the disease. It was found that the forced expression of SOX10 and MYRF in hGPCs expressing mHTT rescued the expression of key genes involved in myelin biogenesis and restored myelin formation in HD-derived glial cells in vivo. Thus, targeted activation or upregulation of SOX10 and MYRF can be used as a means to restore the myelin-forming capacity of oligodendrocytes expressing mHTT in HD.

[0355] In addition to the defects in oligodendrocyte maturation and myelin formation associated with mHTT, it was also noted that astrocyte differentiation was impaired, which might be expected since glial transcription was dysregulated as early as the NKX2.2 and OLIG2 stages approaching the astrocyte-oligodendrocyte fate choice. This defective astrocyte maturation in HD hGPCs suggests that the HD phenotype may have an important developmental component, as any delay in astrocyte differentiation of hGPCs expressing mHTT may impair developmental synaptogenesis and circuit formation, each of which depends on astrocyte guidance (Clarke et al., “Glia Keep Synapse Distribution Under Wraps,” Cell 154:267-268 (2013); Ullian et al., “Control of Synapse Number by Glia,” Science (New York, NY) 291:657-661 (2001), which are hereby incorporated by reference in their entirety). Additionally, any such disease-dependent delay in astrocyte maturation is expected to contribute to the delay in HD (and ultimately to the hypomyelination in HD) since oligodendrocytes have a metabolic dependence on local astrocytes) (Amaral et al., “Metabolic aspects of neuron-oligodendrocyte-astrocyte interactions,” Front Endocrinol (Lausanne) 4:54 (2013), which is hereby incorporated by reference). Whether the rescue of astrocyte maturation from HD-derived hGPCs may alleviate these effects on synapse development and organization remains to be seen; if it can, we can predict that in a manner similar to how the rescue of oligodendrocyte differentiation seems sufficient to alleviate the myelin formation defects in HD, astrocyte replacement may be sufficient to rescue the synaptic pathology in HD.

[0356] In addition to their contributions to neural network formation and synaptic architecture, both hGPCs and astrocytes are intimately involved in maintaining adult interstitial ion homeostasis and in regulating neuronal excitability. Thus, it is of interest that the arrested terminal differentiation of mHTT-expressing hGPCs is associated with the widespread repression of several glial potassium channel families. These include inwardly rectifying K + channels, including KCNJ8 and KCNJ9, among others. This repression of inwardly rectifying K + channels (which are responsible for importing potassium ions into cells) may contribute to the hyperexcitability of HD neurons by inhibiting the glial reuptake of synaptically released K + (Shin et al., “Expression of Mutant Huntingtin in Glial Cells Contributes to Neuronal Excitotoxicity,” J Cell Biol 171:1001-1012 (2005), which is hereby incorporated by reference in its entirety). In this regard, Khakh and colleagues have reported reduced astrocytic expression of the inward rectifier channel Kir4.1 (KCNJ10) in an HD mouse model (Tong et al., “Astrocyte Kir4.1 Ion Channel Deficits Contribute to Neuronal Dysfunction in Huntington's Disease Model Mice,” Nat Neurosci 17:694-703 (2014), which is hereby incorporated by reference in its entirety), which may similarly reflect the impact of disrupted glial maturation on potassium channel expression and glial K + uptake. In human GPCs expressing mHTT that are arrested at a stage prior to astrocytic terminal maturation, it appears that a large number of K + channel transcripts are coordinately repressed, suggesting inhibition of a shared upstream activator of K + channel gene expression. Although the upstream regulators of these potassium channel genes have not been identified, it is reasonable to hypothesize that mHTT-dependent repression of terminal glial differentiation may result in the failure of glial potassium homeostatic mechanisms to develop, mechanisms that would otherwise regulate and protect neuronal activity.

[0357] In summary, these observations suggest that any disruption of HD hGPCs to astrocyte maturation is likely to significantly impact the development and adult manifestation of neural networks in HD. Importantly, a corollary of these findings is that replacement of mHTT-expressing hGPCs with wild-type or genetically corrected counterparts may be sufficient to restore functional astrocytes and oligodendrocytes in the affected HD brain. This possibility was initially suggested by the ability of neonatally delivered wild-type hGPCs to outcompete diseased hGPCs in a model of congenital hypomyelination (Windrem et al., “Neonatal Chimerization with Human Glial Progenitor Cells can Both Remyelinate and Rescue the Otherwise Lethally Hypomyelinated Shiverer Mouse,” Cell Stem Cell 2:553-565 (2008), which is hereby incorporated by reference in its entirety), and equally notable is that neonatal glial replacement is also sufficient to correct deficits in potassium homeostasis in HD transgenic mice (Benraiss et al., “Human Glia can Both Induce and Rescue Aspects of Phenotype in Huntington Disease. Nature Communications 7:11758 (2016), which is hereby incorporated by reference in its entirety). Whether this competitive advantage of healthy cells over diseased cells will emerge in adult HD remains to be determined, but if this proves to be feasible, such glial replacement strategies may prove to be a realistic therapeutic avenue for HD disease amelioration.

[0358] It should be understood that the above-described variations and other features and functions, or alternatives thereof, can be combined into many other different systems or applications. Various presently unforeseen or unexpected substitutions, modifications, variations or improvements may subsequently be made by those skilled in the art, which are also intended to be encompassed by the appended claims.

Claims

1. Use of one or more modulators of at least one gene or protein encoded thereby that participate in the NKX2.2→OLIG2→SOX10→MYRF regulatory cascade in the central nervous system or the brain in the preparation of a medicament for treating or inhibiting the onset of Huntington's disease in a human subject, wherein the at least one gene or protein encoded thereby is the SOX10 or MYRF gene or protein or a combination thereof, wherein the human subject has Huntington's disease or is at risk of developing Huntington's disease, and wherein the medicament is administered to the human subject under conditions effective to treat or inhibit the onset of Huntington's disease in the human subject.

2. The use according to claim 1, wherein the at least one gene or protein is SOX10.

3. The use according to claim 1, wherein the at least one gene or protein is MYRF.

4. The use according to claim 1, wherein the at least one gene or protein is SOX10 and MYRF.

5. The use according to any one of claims 1-4, wherein the administration is carried out using intracerebral delivery, intrathecal delivery, intranasal delivery, or by direct infusion into the cerebral ventricle.

6. The use according to any one of claims 1-5, wherein the treatment or inhibition further comprises: administering to the human subject a preparation of human glial progenitor cells.

7. The use according to claim 6, wherein the human glial progenitor cells are astrocyte-biased glial progenitor cells.

8. The use according to claim 6, wherein the glial progenitor cells of the article are A2B5 + , CD140a + and / or CD44 + .

9. The use according to claim 6, wherein the glial progenitor cells are derived from fetal tissue.

10. The use according to claim 6, wherein the glial progenitor cells are derived from embryonic stem cells.

11. The use according to claim 6, wherein the glial progenitor cells are derived from induced pluripotent stem cells.

12. The use according to any one of claims 6-11, wherein the preparation of human glial progenitor cells is administered to the striatum, forebrain, brainstem, and / or cerebellum of the human subject.

13. The use according to any one of claims 1-12, wherein Huntington's disease is treated.

14. The use according to any one of claims 1-12, wherein the onset of Huntington's disease is inhibited.

15. The use according to any one of claims 1-12, wherein the one or more modulators are agonists.

16. The use according to any one of claims 1-15, wherein the one or more modulators are selected from the group consisting of: a) nucleic acid molecules; b) peptides; and c) small molecules.

17. The use according to claim 16, wherein the one or more modulators are nucleic acid molecules that encode: a) SOX10 protein; b) MYRF protein; c) a combination of SOX10 protein and MYRF protein.

18. The use according to claim 17, wherein the nucleic acid molecule comprises an expression vector, optionally, wherein the expression vector is (1) a viral vector, such as an adenovirus, adeno-associated virus vector, retrovirus vector, lentivirus vector, or herpesvirus vector; or (2) a non-viral vector, such as a plasmid.

19. The use according to any one of claims 17-18, wherein the nucleic acid molecule is operably linked to a promoter, optionally, wherein the promoter is an inducible promoter.

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