Integrating molecular thermodynamics and systems biology to improve the cellular effectiveness of antisense oligonucleotides

Antisense oligonucleotides (AS ONs) are a powerful means to inhibit the expression of specific genes, but their effectiveness is limited by factors including cellular delivery, biochemical attack, and poor binding to target. We have developed a systems model of the processes required for an antisense oligonucleotide to be successfully delivered and to exert activity in a cell. The model demonstrates a sensitive dependence of the extent of gene expression inhibition on the binding rate for the ON-mRNA interaction. We have focused on the binding event and developed a molecular thermodynamic model based on ON and mRNA folding that predicts accurately the affinity of ON-mRNA binding, and we have observed experimentally that affinity correlates directly with binding rate for these species. Most importantly, we have found that the model successfully selects those ONs that are most successful in cell culture. Taken together, these tools may help accelerate the development of this promising technology.