Phosphorodiamidate morpholino oligomers suppress mutant huntingtin expression and attenuate neurotoxicity

Xin Sun, Leonard O. Marque, Zachary Cordner, Jennifer L. Pruitt, Manik Bhat, Pan Li, Geetha Kannan, Ellen Elizabeth Ladenheim, Timothy H Moran, Russell Louis Margolis, Dobrila D. Rudnicki

Research output: Contribution to journalArticle

Abstract

Huntington's disease (HD) is a neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the huntingtin (HTT) gene. Disease pathogenesis derives, at least in part, from the long polyglutamine tract encoded by mutant HTT. Therefore, considerable effort has been dedicated to the development of therapeutic strategies that significantly reduce the expression of the mutant HTT protein. Antisense oligonucleotides (ASOs) targeted to the CAG repeat region of HTT transcripts have been of particular interest due to their potential capacity to discriminate between normal and mutant HTT transcripts. Here, we focus on phosphorodiamidate morpholino oligomers (PMOs), ASOs that are especially stable, highly soluble and non-toxic. We designed three PMOs to selectively target expanded CAG repeat tracts (CTG22, CTG25 and CTG28), and two PMOs to selectively target sequences flanking the HTT CAG repeat (HTTex1a and HTTex1b). In HD patient-derived fibroblasts with expanded alleles containing 44, 77 or 109 CAG repeats, HTTex1a and HTTex1b were effective in suppressing the expression of mutant and non-mutant transcripts. CTGn PMOs also suppressed HTT expression, with the extent of suppression and the specificity for mutant transcripts dependent on the length of the targeted CAG repeat and on the CTG repeat length and concentration of the PMO. PMO CTG25 reduced HTT-induced cytotoxicity in vitro and suppressed mutant HTT expression in vivo in the N171-82Q transgenic mouse model. Finally, CTG28 reduced mutant HTT expression and improved the phenotype of Hdh(Q7/Q150) knock-in HD mice. These data demonstrate the potential of PMOs as an approach to suppressing the expression of mutant HTT.

Original languageEnglish (US)
Pages (from-to)6302-6317
Number of pages16
JournalHuman Molecular Genetics
Volume23
Issue number23
DOIs
StatePublished - Dec 1 2014

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Morpholinos
Huntington Disease
Antisense Oligonucleotides
Trinucleotide Repeat Expansion
Mutant Proteins
Neurodegenerative Diseases
Transgenic Mice
Fibroblasts
Alleles
Phenotype
Genes

ASJC Scopus subject areas

  • Medicine(all)

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Phosphorodiamidate morpholino oligomers suppress mutant huntingtin expression and attenuate neurotoxicity. / Sun, Xin; Marque, Leonard O.; Cordner, Zachary; Pruitt, Jennifer L.; Bhat, Manik; Li, Pan; Kannan, Geetha; Ladenheim, Ellen Elizabeth; Moran, Timothy H; Margolis, Russell Louis; Rudnicki, Dobrila D.

In: Human Molecular Genetics, Vol. 23, No. 23, 01.12.2014, p. 6302-6317.

Research output: Contribution to journalArticle

Sun, Xin ; Marque, Leonard O. ; Cordner, Zachary ; Pruitt, Jennifer L. ; Bhat, Manik ; Li, Pan ; Kannan, Geetha ; Ladenheim, Ellen Elizabeth ; Moran, Timothy H ; Margolis, Russell Louis ; Rudnicki, Dobrila D. / Phosphorodiamidate morpholino oligomers suppress mutant huntingtin expression and attenuate neurotoxicity. In: Human Molecular Genetics. 2014 ; Vol. 23, No. 23. pp. 6302-6317.
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AU - Sun, Xin

AU - Marque, Leonard O.

AU - Cordner, Zachary

AU - Pruitt, Jennifer L.

AU - Bhat, Manik

AU - Li, Pan

AU - Kannan, Geetha

AU - Ladenheim, Ellen Elizabeth

AU - Moran, Timothy H

AU - Margolis, Russell Louis

AU - Rudnicki, Dobrila D.

PY - 2014/12/1

Y1 - 2014/12/1

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AB - Huntington's disease (HD) is a neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the huntingtin (HTT) gene. Disease pathogenesis derives, at least in part, from the long polyglutamine tract encoded by mutant HTT. Therefore, considerable effort has been dedicated to the development of therapeutic strategies that significantly reduce the expression of the mutant HTT protein. Antisense oligonucleotides (ASOs) targeted to the CAG repeat region of HTT transcripts have been of particular interest due to their potential capacity to discriminate between normal and mutant HTT transcripts. Here, we focus on phosphorodiamidate morpholino oligomers (PMOs), ASOs that are especially stable, highly soluble and non-toxic. We designed three PMOs to selectively target expanded CAG repeat tracts (CTG22, CTG25 and CTG28), and two PMOs to selectively target sequences flanking the HTT CAG repeat (HTTex1a and HTTex1b). In HD patient-derived fibroblasts with expanded alleles containing 44, 77 or 109 CAG repeats, HTTex1a and HTTex1b were effective in suppressing the expression of mutant and non-mutant transcripts. CTGn PMOs also suppressed HTT expression, with the extent of suppression and the specificity for mutant transcripts dependent on the length of the targeted CAG repeat and on the CTG repeat length and concentration of the PMO. PMO CTG25 reduced HTT-induced cytotoxicity in vitro and suppressed mutant HTT expression in vivo in the N171-82Q transgenic mouse model. Finally, CTG28 reduced mutant HTT expression and improved the phenotype of Hdh(Q7/Q150) knock-in HD mice. These data demonstrate the potential of PMOs as an approach to suppressing the expression of mutant HTT.

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