Short (16-mer) locked nucleic acid splice-switching oligonucleotides restore dystrophin production in Duchenne Muscular Dystrophy myotubes

Pires, Vanessa Borges; Simões, Ricardo G.; Mamchaoui, Kamel; Carvalho, Célia; Carmo‐Fonseca, Maria

doi:10.1371/journal.pone.0181065

Cited by 8 publications

(4 citation statements)

References 49 publications

(54 reference statements)

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“…Our previous report revealed that the different length of SSOs change the off-target effects. Additionally, Pires et al mentioned that the appropriate length of LNA-modified SSO enables the prevention of off-target effects [ 28 ]. Although further study is necessary to determine the off-target effects of BNA-modified SSOs, we at least suggest that both AmNA and GuNA are candidates of chemical modifications for better splicing modulation.…”

Section: Discussionmentioning

confidence: 99%

Design and In Vitro Evaluation of Splice-Switching Oligonucleotides Bearing Locked Nucleic Acids, Amido-Bridged Nucleic Acids, and Guanidine-Bridged Nucleic Acids

Shimo

Nakatsuji

Tachibana

et al. 2021

IJMS

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Our group previously developed a series of bridged nucleic acids (BNAs), including locked nucleic acids (LNAs), amido-bridged nucleic acids (AmNAs), and guanidine-bridged nucleic acids (GuNAs), to impart specific characteristics to oligonucleotides such as high-affinity binding and enhanced enzymatic resistance. In this study, we designed a series of LNA-, AmNA-, and GuNA-modified splice-switching oligonucleotides (SSOs) with different lengths and content modifications. We measured the melting temperature (Tm) of each designed SSO to investigate its binding affinity for RNA strands. We also investigated whether the single-stranded SSOs formed secondary structures using UV melting analysis without complementary RNA. As a result, the AmNA-modified SSOs showed almost the same Tm values as the LNA-modified SSOs, with decreased secondary structure formation in the former. In contrast, the GuNA-modified SSOs showed slightly lower Tm values than the LNA-modified SSOs, with no inhibition of secondary structures. We also evaluated the exon skipping activities of the BNAs in vitro at both the mRNA and protein expression levels. We found that both AmNA-modified SSOs and GuNA-modified SSOs showed higher exon skipping activities than LNA-modified SSOs but each class must be appropriately designed in terms of length and modification content.

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Section: Discussionmentioning

confidence: 99%

Design and In Vitro Evaluation of Splice-Switching Oligonucleotides Bearing Locked Nucleic Acids, Amido-Bridged Nucleic Acids, and Guanidine-Bridged Nucleic Acids

Shimo

Nakatsuji

Tachibana

et al. 2021

IJMS

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“… 29 Determining the optimal SSOs for exon skipping has drawn specific attention due to the insufficient outcomes in clinical trials. 30 , 31 , 32 By using in silico prediction tools and in vitro screening approaches, Echigoya et al. have defined 30-mer PMOs with better exon-skipping efficacy than eteplirsen; these PMOs targets the initial 5′ site of the exon 51 instead of the internal region targeted by eteplirsen.…”

Section: Discussionmentioning

confidence: 99%

Shorter Phosphorodiamidate Morpholino Splice-Switching Oligonucleotides May Increase Exon-Skipping Efficacy in DMD

Akpulat

Wang

Becker

et al. 2018

Molecular Therapy - Nucleic Acids

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Duchenne muscular dystrophy is a fatal muscle disease, caused by mutations in DMD, leading to loss of dystrophin expression. Phosphorodiamidate morpholino splice-switching oligonucleotides (PMO-SSOs) have been used to elicit the restoration of a partially functional truncated dystrophin by excluding disruptive exons from the DMD messenger. The 30-mer PMO eteplirsen (EXONDYS51) developed for exon 51 skipping is the first dystrophin-restoring, conditionally FDA-approved drug in history. Clinical trials had shown a dose-dependent variable and patchy dystrophin restoration. The main obstacle for efficient dystrophin restoration is the inadequate uptake of PMOs into skeletal muscle fibers at low doses. The excessive cost of longer PMOs has limited the utilization of higher dosing. We designed shorter 25-mer PMOs directed to the same eteplirsen-targeted region of exon 51 and compared their efficacies in vitro and in vivo in the mdx52 murine model. Our results showed that skipped-dystrophin induction was comparable between the 30-mer PMO sequence of eteplirsen and one of the shorter PMOs, while the other 25-mer PMOs showed lower exon-skipping efficacies. Shorter PMOs would make higher doses economically feasible, and high dosing would result in better drug uptake into muscle, induce higher levels of dystrophin restoration in DMD muscle, and, ultimately, increase the clinical efficacy.

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“…So far, ASOs at lower doses seem to be well-tolerated in humans overall. For example, two ASO therapeutics were recently approved for treating familial hypercholesterolemia, and nusinersen, a splice-switching ASO, was approved for use in patients with spinal muscular atrophy, and multiple ASOs were approved for Duchenne's muscular dystrophy [168,169]. A small subset of mutations within the CFTR gene result in aberrant pre-mRNA splicing.…”

Section: Nucleic Acid-based Therapiesmentioning

confidence: 99%

Immunomodulation in Cystic Fibrosis: Why and How?

Giacalone

Dobosh

Gaggar

et al. 2020

IJMS

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Cystic fibrosis (CF) lung disease is characterized by unconventional mechanisms of inflammation, implicating a chronic immune response dominated by innate immune cells. Historically, therapeutic development has focused on the mutated cystic fibrosis transmembrane conductance regulator (CFTR), leading to the discovery of small molecules aiming at modulating and potentiating the presence and activity of CFTR at the plasma membrane. However, treatment burden sustained by CF patients, side effects of current medications, and recent advances in other therapeutic areas have highlighted the need to develop novel disease targeting of the inflammatory component driving CF lung damage. Furthermore, current issues with standard treatment emphasize the need for directed lung therapies that could minimize systemic side effects. Here, we summarize current treatment used to target immune cells in the lungs, and highlight potential benefits and caveats of novel therapeutic strategies.

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Short (16-mer) locked nucleic acid splice-switching oligonucleotides restore dystrophin production in Duchenne Muscular Dystrophy myotubes

Cited by 8 publications

References 49 publications

Design and In Vitro Evaluation of Splice-Switching Oligonucleotides Bearing Locked Nucleic Acids, Amido-Bridged Nucleic Acids, and Guanidine-Bridged Nucleic Acids

Design and In Vitro Evaluation of Splice-Switching Oligonucleotides Bearing Locked Nucleic Acids, Amido-Bridged Nucleic Acids, and Guanidine-Bridged Nucleic Acids

Shorter Phosphorodiamidate Morpholino Splice-Switching Oligonucleotides May Increase Exon-Skipping Efficacy in DMD

Immunomodulation in Cystic Fibrosis: Why and How?

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