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Parallel synthesis of cell-penetrating peptide conjugates of PMO toward exon skipping enhancement in Duchenne muscular dystrophy.

ABSTRACT: We describe two new methods of parallel chemical synthesis of libraries of peptide conjugates of phosphorodiamidate morpholino oligonucleotide (PMO) cargoes on a scale suitable for cell screening prior to in vivo analysis for therapeutic development. The methods represent an extension of the SELection of PEPtide CONjugates (SELPEPCON) approach previously developed for parallel peptide-peptide nucleic acid (PNA) synthesis. However, these new methods allow for the utilization of commercial PMO as cargo with both C- and N-termini unfunctionalized. The synthetic methods involve conjugation in solution phase, followed by rapid purification via biotin-streptavidin immobilization and subsequent reductive release into solution, avoiding the need for painstaking high-performance liquid chromatography purifications. The synthesis methods were applied for screening of PMO conjugates of a 16-member library of variants of a 10-residue ApoE peptide, which was suggested for blood-brain barrier crossing. In this work the conjugate library was tested in an exon skipping assay using skeletal mouse mdx cells, a model of Duchene's muscular dystrophy where higher activity peptide-PMO conjugates were identified compared with the starting peptide-PMO. The results demonstrate the power of the parallel synthesis methods for increasing the speed of optimization of peptide sequences in conjugates of PMO for therapeutic screening.

SUBMITTER: O'Donovan L

PROVIDER: S-EPMC4296749 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature

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Parallel synthesis of cell-penetrating peptide conjugates of PMO toward exon skipping enhancement in Duchenne muscular dystrophy.

O'Donovan Liz L Okamoto Itaru I Arzumanov Andrey A AA Williams Donna L DL Deuss Peter P Gait Michael J MJ

Nucleic acid therapeutics 20141120 1

We describe two new methods of parallel chemical synthesis of libraries of peptide conjugates of phosphorodiamidate morpholino oligonucleotide (PMO) cargoes on a scale suitable for cell screening prior to in vivo analysis for therapeutic development. The methods represent an extension of the SELection of PEPtide CONjugates (SELPEPCON) approach previously developed for parallel peptide-peptide nucleic acid (PNA) synthesis. However, these new methods allow for the utilization of commercial PMO as ...[more]

PMID: 25412073

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Project description:BACKGROUND:Exon skipping has been considered a promising therapeutic approach for Duchenne muscular dystrophy (DMD). Eteplirsen received conditional approval in the United States in 2016. To date, no systematic reviews or meta-analyses of randomized controlled trials (RCTs) of exon skipping drugs have been published to determine the pooled estimates for the effect of exon skipping in treating DMD. METHODS:A systematic review and meta-analysis of double-blind RCTs comparing exon-skipping drugs with placebo in DMD was performed. Trials were identified by searching published and unpublished studies from electronically available databases and clinical trial registries through October 2017. The primary outcomes were changes in the 6-min walk test (6MWT) distance, North Star Ambulatory Assessment (NSAA) scores, and adverse events. Random-effects meta-analysis and assessment of risk of bias were performed. This systematic review was registered at PROSPERO (CRD42016037504). RESULTS:Five studies involving 322 participants were included, investigating eteplirsen in one and drisapersen in four studies. There were no changes in 6MWT distance (mean difference [MD] -?9.16, 95% confidence interval [CI] -?21.94 to 3.62) or NSAA scores (MD 1.20, 95% CI -?2.35 to 4.75) after 24 weeks of treatment in the exon-skipping group compared with placebo. Subgroup analysis for a 6 mg/kg weekly injection of drisapersen showed significant changes in the 6MWT, favoring drisapersen after 24 weeks (MD -?20.24; 95% CI -?39.59 to -?0.89). However, drisapersen resulted in a significant increase in injection site reactions (risk ratio [RR] 3.67, 95% CI 1.96 to 6.89, p?<?0.0001) and renal toxicity (RR 1.81, 95% CI 1.11 to 2.94, p?=?0.02). Risk of bias was high in two of the five studies, including the eteplirsen and one drisapersen study. CONCLUSIONS:Current available data do not show evidence that exon-skipping drugs are effective in DMD. Despite potential effectiveness when used at a specific dose, significant side effects were reported with drisapersen. The small number of RCTs with relatively small numbers of participants indicate the difficulty in conducting sufficiently powered studies of DMD. Prospectively planned meta-analysis and utilization of the real-world data may provide a more precise estimate of the effect of exon skipping in this disease.

Dataset Information

Parallel synthesis of cell-penetrating peptide conjugates of PMO toward exon skipping enhancement in Duchenne muscular dystrophy.

Publications

Parallel synthesis of cell-penetrating peptide conjugates of PMO toward exon skipping enhancement in Duchenne muscular dystrophy.

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