A prospective study in the rational design of efficient antisense oligonucleotides for exon skipping in the DMD gene.

Abstract

Antisense oligonucleotide (AON)-mediated exon skipping to restore dystrophin expression in Duchenne muscular dystrophy (DMD) therapy shown promise in a number of human clinical trials. Current AON design methods are semi-empirical, involving either trial-and-error and/or preliminary experimentations. Therefore, a rational approach to design efficient AONs to address the wide spectrum of patients' mutations is desirable. Retrospective studies have extracted many AON design variables, but they were not tested prospectively to design AONs for skipping DMD exons. Not only did the variables differ among the various studies, no numerical cutoff for each variable was inferred, which makes their use in AON design difficult. The challenge is to thus select a minimal set of key independent variables that can consistently design efficient AONs. In this prospective study, a novel set of design variables with respective cutoff values was used to design 23 novel AONs, each to skip one of nine DMD exons. Nineteen AONs were found to be efficacious in inducing specific exon skipping (83% of total), of which 14 were considered efficient (61% of total), i.e., they induced exon skipping in >25% of total transcripts. Notably, the satisfactory success rates were achieved by using only three design variables; namely, co-transcriptional binding accessibility of target site, presence of exonic splicing enhancers, and target length. Retrospective analyses revealed that the most efficient AON in every exon targeted has the lowest average cumulative position (ACP) score. Taking the prospective and retrospective studies together, we propose that design guidelines recommend using the ACP score to select the most efficient AON for each exon.

• Authors: Zacharias Aloysius Dwi Pramono, Keng Boon Wee, Jian Li Wang, Yi Jun Chen, Qian Bin Xiong, Poh San Lai, Woon Chee Yee
• Journal: Human gene therapy (Hum Gene Ther) Vol. 23 Issue 7 Pg. 781-90 (Jul 2012) ISSN: 1557-7422 [Electronic] United States
• PMID: 22486275 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• DMD protein, human
• Dystrophin
• Oligonucleotides, Antisense
• Protein Isoforms
• RNA Precursors

Topics

• Algorithms
• Alternative Splicing
• Base Sequence
• Cells, Cultured
• Computer Simulation
• Dystrophin (genetics, metabolism)
• Exons
• Genetic Therapy
• Humans
• Models, Molecular
• Muscular Dystrophy, Duchenne (therapy)
• Myoblasts (metabolism)
• Nucleic Acid Conformation
• Oligonucleotides, Antisense (genetics)
• Prospective Studies
• Protein Isoforms (genetics, metabolism)
• RNA Precursors (genetics)