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Chemically modified antisense oligonucleotides (ASOs) are widely used as a tool to functionalize microRNAs (miRNAs). Reduction of miRNA level after ASO inhibition is commonly reported to show efficacy. Whether this is the most relevant endpoint for measuring miRNA inhibition has not been adequately addressed in the field although it has important implications for evaluating miRNA targeting studies. Using a novel approach to quantitate miRNA levels in the presence of excess ASO, we have discovered that the outcome of miRNA inhibition can vary depending on the chemical modification of the ASO. Although some miRNA inhibitors cause a decrease in mature miRNA levels, we have identified a novel 2'-fluoro/2'-methoxyethyl modified ASO motif with dramatically improved in vivo potency which does not. These studies show there are multiple mechanisms of miRNA inhibition by ASOs and that evaluation of secondary endpoints is crucial for interpreting miRNA inhibition studies.

Potent inhibition of microRNA in vivo without degradation