Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides
Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disease caused by frameshift or nonsense mutations in the <i>DMD</i> gene, resulting in the loss of dystrophin from muscle membranes. Exon skipping using splice-switching oligonucleotides (SSOs) restores the reading frame of &l...
| Main Authors: | , , , , , , , , , , , , , , |
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| Format: | Journal article |
| Language: | English |
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Mary Ann Liebert
2021
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| _version_ | 1797050263672979456 |
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| author | Tone, Y Mamchaoui, K Tsoumpra, MK Hashimoto, Y Terada, R Maruyama, R Gait, MJ Arzumanov, AA McClorey, G Imamura, M Takeda, S Yokota, T Wood, MJA Mouly, V Aoki, Y |
| author_facet | Tone, Y Mamchaoui, K Tsoumpra, MK Hashimoto, Y Terada, R Maruyama, R Gait, MJ Arzumanov, AA McClorey, G Imamura, M Takeda, S Yokota, T Wood, MJA Mouly, V Aoki, Y |
| author_sort | Tone, Y |
| collection | OXFORD |
| description | Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disease caused by frameshift or nonsense mutations in the <i>DMD</i> gene, resulting in the loss of dystrophin from muscle membranes. Exon skipping using splice-switching oligonucleotides (SSOs) restores the reading frame of <i>DMD</i> pre-mRNA by generating internally truncated but functional dystrophin protein. To potentiate effective tissue-specific targeting by functional SSOs, it is essential to perform accelerated and reliable <i>in vitro</i> screening-based assessment of novel oligonucleotides and drug delivery technologies, such as cell-penetrating peptides, before their <i>in vivo</i> pharmacokinetic and toxicity evaluation. We have established novel canine immortalized myoblast lines by transducing murine cyclin-dependent kinase-4 and human telomerase reverse transcriptase genes into myoblasts isolated from beagle-based wild-type or canine X-linked muscular dystrophy in Japan (CXMD<sub>J</sub>) dogs. These myoblast lines exhibited improved myogenic differentiation and increased proliferation rates compared with passage-15 primary parental myoblasts, and their potential to differentiate into myotubes was maintained in later passages. Using these dystrophin-deficient immortalized myoblast lines, we demonstrate that a novel cell-penetrating peptide (Pip8b2)-conjugated SSO markedly improved multiexon skipping activity compared with the respective naked phosphorodiamidate morpholino oligomers. <i>In vitro</i> screening using immortalized canine cell lines will provide a basis for further pharmacological studies on drug delivery tools. |
| first_indexed | 2024-03-06T18:02:35Z |
| format | Journal article |
| id | oxford-uuid:0052aad1-fe0c-421d-ac26-9d0990db3db6 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-06T18:02:35Z |
| publishDate | 2021 |
| publisher | Mary Ann Liebert |
| record_format | dspace |
| spelling | oxford-uuid:0052aad1-fe0c-421d-ac26-9d0990db3db62022-03-26T08:28:56ZImmortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotidesJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:0052aad1-fe0c-421d-ac26-9d0990db3db6EnglishSymplectic ElementsMary Ann Liebert2021Tone, YMamchaoui, KTsoumpra, MKHashimoto, YTerada, RMaruyama, RGait, MJArzumanov, AAMcClorey, GImamura, MTakeda, SYokota, TWood, MJAMouly, VAoki, YDuchenne muscular dystrophy (DMD) is a severe muscle-wasting disease caused by frameshift or nonsense mutations in the <i>DMD</i> gene, resulting in the loss of dystrophin from muscle membranes. Exon skipping using splice-switching oligonucleotides (SSOs) restores the reading frame of <i>DMD</i> pre-mRNA by generating internally truncated but functional dystrophin protein. To potentiate effective tissue-specific targeting by functional SSOs, it is essential to perform accelerated and reliable <i>in vitro</i> screening-based assessment of novel oligonucleotides and drug delivery technologies, such as cell-penetrating peptides, before their <i>in vivo</i> pharmacokinetic and toxicity evaluation. We have established novel canine immortalized myoblast lines by transducing murine cyclin-dependent kinase-4 and human telomerase reverse transcriptase genes into myoblasts isolated from beagle-based wild-type or canine X-linked muscular dystrophy in Japan (CXMD<sub>J</sub>) dogs. These myoblast lines exhibited improved myogenic differentiation and increased proliferation rates compared with passage-15 primary parental myoblasts, and their potential to differentiate into myotubes was maintained in later passages. Using these dystrophin-deficient immortalized myoblast lines, we demonstrate that a novel cell-penetrating peptide (Pip8b2)-conjugated SSO markedly improved multiexon skipping activity compared with the respective naked phosphorodiamidate morpholino oligomers. <i>In vitro</i> screening using immortalized canine cell lines will provide a basis for further pharmacological studies on drug delivery tools. |
| spellingShingle | Tone, Y Mamchaoui, K Tsoumpra, MK Hashimoto, Y Terada, R Maruyama, R Gait, MJ Arzumanov, AA McClorey, G Imamura, M Takeda, S Yokota, T Wood, MJA Mouly, V Aoki, Y Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides |
| title | Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides |
| title_full | Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides |
| title_fullStr | Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides |
| title_full_unstemmed | Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides |
| title_short | Immortalized canine dystrophic myoblast cell lines for development of peptide-conjugated splice-switching oligonucleotides |
| title_sort | immortalized canine dystrophic myoblast cell lines for development of peptide conjugated splice switching oligonucleotides |
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