Allele-selective inhibition of mutant atrophin-1 expression by duplex and single-stranded RNAs.

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Dentatorubral-pallidoluysian atrophy (DRPLA ) is a progressive neurodegenerative disorder that currently has no curative treatments . DRPLA is caused by an expansion of a CAG trinucleotide repeat region within the protein-encoding sequence of the atrophin-1 (ATN-1 ) gene . Inhibition of mutant ATN-1 protein expression is one strategy for treating DRPLA , and allele-selective gene silencing agents that block mutant expression over wild-type expression would be lead compounds for therapeutic development . Here we develop an assay for distinguishing mutant from wild-type ATN-1 protein by gel electrophoresis . We use this assay to evaluate duplex RNAs and single -stranded silencing RNAs (ss-si RNAs ) for allele-selective inhibition of ATN-1 protein expression . We observed potent and allele-selective inhibition by RNA duplexes that contain mismatched bases relative to the CAG target and have the potential to form miRNA-like complexes . ss-si RNAs that contained mismatches were as selective as mismatch-containing duplexes . We also report allele-selective inhibition by duplex RNAs containing unlocked nucleic acids or abasic substitutions , although selectivities are not as high . Five compounds that showed >8 -fold allele selectivity for mutant ATN-1 were also selective for inhibiting the expression of two other trinucleotide repeat disease genes , ataxin-3 (ATXN-3 ) and huntingtin (HTT ). These data demonstrate that the expanded trinucleotide repeat within ATN-1 mRNA is a potential target for compounds designed to achieve allele-selective inhibition of ATN-1 protein , and one agent may allow the targeting of multiple disease genes .