Efficient exon skipping of SGCG mutations mediated by phosphorodiamidate morpholino oligomers

Eugene Jeffrey Wyatt, Alexis R Demonbreun, Ellis Y. Kim, Megan J Roy-Puckelwartz, Andy H. Vo, Lisa Marie Castillo, Quan Q. Gao, Mariz Vainzof, Rita C.M. Pavanello, Mayana Zatz, Elizabeth M McNally

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Exon skipping uses chemically modified antisense oligonucleotides to modulate RNA splicing. Therapeutically, exon skipping can bypass mutations and restore reading frame disruption by generating internally truncated, functional proteins to rescue the loss of native gene expression. Limb-girdle muscular dystrophy type 2C is caused by autosomal recessive mutations in the SGCG gene, which encodes the dystrophin-associated protein γ-sarcoglycan. The most common SGCG mutations disrupt the transcript reading frame abrogating γ-sarcoglycan protein expression. In order to treat most SGCG gene mutations, it is necessary to skip 4 exons in order to restore the SGCG transcript reading frame, creating an internally truncated protein referred to as Mini-Gamma. Using direct reprogramming of human cells with MyoD, myogenic cells were tested with 2 antisense oligonucleotide chemistries, 2'-O-methyl phosphorothioate oligonucleotides and vivo-phosphorodiamidate morpholino oligomers, to induce exon skipping. Treatment with vivo-phosphorodiamidate morpholino oligomers demonstrated efficient skipping of the targeted exons and corrected the mutant reading frame, resulting in the expression of a functional Mini-Gamma protein. Antisense-induced exon skipping of SGCG occurred in normal cells and those with multiple distinct SGCG mutations, including the most common 521ΔT mutation. These findings demonstrate a multiexon-skipping strategy applicable to the majority of limb-girdle muscular dystrophy 2C patients.

Original languageEnglish (US)
JournalJCI insight
Volume3
Issue number9
DOIs
StatePublished - May 3 2018

Fingerprint

Morpholinos
Exons
Reading Frames
Mutation
Sarcoglycans
Antisense Oligonucleotides
Dystrophin-Associated Proteins
Proteins
Phosphorothioate Oligonucleotides
Limb-Girdle Muscular Dystrophies
RNA Splicing
Genes
Gene Expression

Keywords

  • Genetics
  • Monogenic diseases
  • Muscle Biology
  • Neuromuscular disease
  • Skeletal muscle

Cite this

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title = "Efficient exon skipping of SGCG mutations mediated by phosphorodiamidate morpholino oligomers",
abstract = "Exon skipping uses chemically modified antisense oligonucleotides to modulate RNA splicing. Therapeutically, exon skipping can bypass mutations and restore reading frame disruption by generating internally truncated, functional proteins to rescue the loss of native gene expression. Limb-girdle muscular dystrophy type 2C is caused by autosomal recessive mutations in the SGCG gene, which encodes the dystrophin-associated protein γ-sarcoglycan. The most common SGCG mutations disrupt the transcript reading frame abrogating γ-sarcoglycan protein expression. In order to treat most SGCG gene mutations, it is necessary to skip 4 exons in order to restore the SGCG transcript reading frame, creating an internally truncated protein referred to as Mini-Gamma. Using direct reprogramming of human cells with MyoD, myogenic cells were tested with 2 antisense oligonucleotide chemistries, 2'-O-methyl phosphorothioate oligonucleotides and vivo-phosphorodiamidate morpholino oligomers, to induce exon skipping. Treatment with vivo-phosphorodiamidate morpholino oligomers demonstrated efficient skipping of the targeted exons and corrected the mutant reading frame, resulting in the expression of a functional Mini-Gamma protein. Antisense-induced exon skipping of SGCG occurred in normal cells and those with multiple distinct SGCG mutations, including the most common 521ΔT mutation. These findings demonstrate a multiexon-skipping strategy applicable to the majority of limb-girdle muscular dystrophy 2C patients.",
keywords = "Genetics, Monogenic diseases, Muscle Biology, Neuromuscular disease, Skeletal muscle",
author = "Wyatt, {Eugene Jeffrey} and Demonbreun, {Alexis R} and Kim, {Ellis Y.} and Roy-Puckelwartz, {Megan J} and Vo, {Andy H.} and Castillo, {Lisa Marie} and Gao, {Quan Q.} and Mariz Vainzof and Pavanello, {Rita C.M.} and Mayana Zatz and McNally, {Elizabeth M}",
year = "2018",
month = "5",
day = "3",
doi = "10.1172/jci.insight.99357",
language = "English (US)",
volume = "3",
journal = "JCI insight",
issn = "2379-3708",
publisher = "The American Society for Clinical Investigation",
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Efficient exon skipping of SGCG mutations mediated by phosphorodiamidate morpholino oligomers. / Wyatt, Eugene Jeffrey; Demonbreun, Alexis R; Kim, Ellis Y.; Roy-Puckelwartz, Megan J; Vo, Andy H.; Castillo, Lisa Marie; Gao, Quan Q.; Vainzof, Mariz; Pavanello, Rita C.M.; Zatz, Mayana; McNally, Elizabeth M.

In: JCI insight, Vol. 3, No. 9, 03.05.2018.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Efficient exon skipping of SGCG mutations mediated by phosphorodiamidate morpholino oligomers

AU - Wyatt, Eugene Jeffrey

AU - Demonbreun, Alexis R

AU - Kim, Ellis Y.

AU - Roy-Puckelwartz, Megan J

AU - Vo, Andy H.

AU - Castillo, Lisa Marie

AU - Gao, Quan Q.

AU - Vainzof, Mariz

AU - Pavanello, Rita C.M.

AU - Zatz, Mayana

AU - McNally, Elizabeth M

PY - 2018/5/3

Y1 - 2018/5/3

N2 - Exon skipping uses chemically modified antisense oligonucleotides to modulate RNA splicing. Therapeutically, exon skipping can bypass mutations and restore reading frame disruption by generating internally truncated, functional proteins to rescue the loss of native gene expression. Limb-girdle muscular dystrophy type 2C is caused by autosomal recessive mutations in the SGCG gene, which encodes the dystrophin-associated protein γ-sarcoglycan. The most common SGCG mutations disrupt the transcript reading frame abrogating γ-sarcoglycan protein expression. In order to treat most SGCG gene mutations, it is necessary to skip 4 exons in order to restore the SGCG transcript reading frame, creating an internally truncated protein referred to as Mini-Gamma. Using direct reprogramming of human cells with MyoD, myogenic cells were tested with 2 antisense oligonucleotide chemistries, 2'-O-methyl phosphorothioate oligonucleotides and vivo-phosphorodiamidate morpholino oligomers, to induce exon skipping. Treatment with vivo-phosphorodiamidate morpholino oligomers demonstrated efficient skipping of the targeted exons and corrected the mutant reading frame, resulting in the expression of a functional Mini-Gamma protein. Antisense-induced exon skipping of SGCG occurred in normal cells and those with multiple distinct SGCG mutations, including the most common 521ΔT mutation. These findings demonstrate a multiexon-skipping strategy applicable to the majority of limb-girdle muscular dystrophy 2C patients.

AB - Exon skipping uses chemically modified antisense oligonucleotides to modulate RNA splicing. Therapeutically, exon skipping can bypass mutations and restore reading frame disruption by generating internally truncated, functional proteins to rescue the loss of native gene expression. Limb-girdle muscular dystrophy type 2C is caused by autosomal recessive mutations in the SGCG gene, which encodes the dystrophin-associated protein γ-sarcoglycan. The most common SGCG mutations disrupt the transcript reading frame abrogating γ-sarcoglycan protein expression. In order to treat most SGCG gene mutations, it is necessary to skip 4 exons in order to restore the SGCG transcript reading frame, creating an internally truncated protein referred to as Mini-Gamma. Using direct reprogramming of human cells with MyoD, myogenic cells were tested with 2 antisense oligonucleotide chemistries, 2'-O-methyl phosphorothioate oligonucleotides and vivo-phosphorodiamidate morpholino oligomers, to induce exon skipping. Treatment with vivo-phosphorodiamidate morpholino oligomers demonstrated efficient skipping of the targeted exons and corrected the mutant reading frame, resulting in the expression of a functional Mini-Gamma protein. Antisense-induced exon skipping of SGCG occurred in normal cells and those with multiple distinct SGCG mutations, including the most common 521ΔT mutation. These findings demonstrate a multiexon-skipping strategy applicable to the majority of limb-girdle muscular dystrophy 2C patients.

KW - Genetics

KW - Monogenic diseases

KW - Muscle Biology

KW - Neuromuscular disease

KW - Skeletal muscle

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U2 - 10.1172/jci.insight.99357

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