Mesyl Phosphoramidate Oligonucleotides as Potential Splice-Switching Agents: Impact of Backbone Structure on Activity and Intracellular Localization
Suzan M. Hammond, Olga V. Sergeeva, Pavel Melnikov, Larissa Goli, Jessica Stoodley, Timofei S. Zatsepin, Dmitry A. Stetsenko, Matthew J. A. Wood
Abstract
A series of 2′-deoxy and novel 2′- O -methyl and 2′- O -(2-methoxyethyl) (2′-MOE) oligonucleotides with internucleotide methanesulfonyl (mesyl, μ) or 1-butanesulfonyl (busyl, β) phosphoramidate groups has been synthesized for evaluation as potential splice-switching oligonucleotides. Evaluation of their splice-switching activity in spinal muscular atrophy patient-derived fibroblasts revealed no significant difference in splice-switching efficacy between 2′-MOE mesyl oligonucleotide and the corresponding phosphorothioate (nusinersen). Yet, a survival study with model neonatal mice has shown the antisense 2′-MOE mesyl oligonucleotide to be inferior to nusinersen at the highest dose of 40 mg/kg. A reason for their lower activity in vivo as ascertained by cellular uptake study by fluorescent confocal microscopy in HEK293 cell line could possibly be ascribed to compromised endosomal release and/or nuclear uptake of the 2′-OMe or 2′-MOE μ- and β-oligonucleotides compared to their phosphorothioate analog.