Physicochemical properties of (R)- vs (S)-methylphosphonate substitution on antisense DNA hybridization determined by free energy perturbation and molecular dynamics

Methylphosphonate (MP)-substituted antisense DNA oligomers have significant potential for genome targeted therapy. The physicochemical basis of the difference in R- versus S-MP diastereomers incorporated into DNA oligomers with regard to complementary DNA target hybridization has been poorly understood. State of the art advanced molecular computational methods and supercomputer technology were applied to identify key physicochemical determinants involved in stabilizing and destabilizing target hybridization. MP-oligomer:DNA target hybridization is more stable with R-MP substitution and is due to favorable hydrophobic interactions between the equatorial projecting methyl group and water molecules. S-MP destabilizes the double strand helix formation by less favorable local hydrophobic interactions with waters, which result in DNA helix unwinding, and by promoting local changes from C2′ endo to C3′ endo in the 5′ furanose ring. In the oligomer studied, R-MP thymine is more stable than R-MP uracil, resulting from hydrophobic interaction to locally stabilize the helix due to the presence of the pyrimidine C5 methyl group. The presence of the C5 methyl group in thymine also appears to influence helical winding and the local conformation of the furanose ring. These numerical simulations are supported by experimental data, enabling us to propose several mechanisms which underlie some of the experimental observations. These results support further development of diastereomerically pure MP analogues as diagnostic/therapeutic agents involving sequence-specific DNA interactions.