Functionalization of the 3′-Ends of DNA and RNA Strands with N-ethyl-N-coupled Nucleosides: A Promising Approach To Avoid 3′-Exonuclease-Catalyzed Hydrolysis of Therapeutic Oligonucleotides

Authors

  • Dr. Montserrat Terrazas,

    Corresponding author
    1. Institute for Research in Biomedicine (IRB Barcelona) and Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Cluster Building, Baldiri i Reixac 10, 08028 Barcelona (Spain)
    • Institute for Research in Biomedicine (IRB Barcelona) and Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Cluster Building, Baldiri i Reixac 10, 08028 Barcelona (Spain)
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  • Adele Alagia,

    1. Institute for Research in Biomedicine (IRB Barcelona) and Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Cluster Building, Baldiri i Reixac 10, 08028 Barcelona (Spain)
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  • Ignacio Faustino,

    1. Joint IRB–BSC Program on Computational Biology, Institute for Research in Biomedicine (IRB Barcelona), Baldiri i Reixac 10, 08028 Barcelona (Spain)
    2. Department of Biochemistry, University of Barcelona, Diagonal 647, 08028 Barcelona (Spain)
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  • Prof. Dr. Modesto Orozco,

    1. Joint IRB–BSC Program on Computational Biology, Institute for Research in Biomedicine (IRB Barcelona), Baldiri i Reixac 10, 08028 Barcelona (Spain)
    2. Department of Biochemistry, University of Barcelona, Diagonal 647, 08028 Barcelona (Spain)
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  • Prof. Dr. Ramon Eritja

    Corresponding author
    1. Institute for Research in Biomedicine (IRB Barcelona) and Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Cluster Building, Baldiri i Reixac 10, 08028 Barcelona (Spain)
    • Institute for Research in Biomedicine (IRB Barcelona) and Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Cluster Building, Baldiri i Reixac 10, 08028 Barcelona (Spain)
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Abstract

The development of nucleic acid derivatives to generate novel medical treatments has become increasingly popular, but the high vulnerability of oligonucleotides to nucleases limits their practical use. We explored the possibility of increasing the stability against 3′-exonucleases by replacing the two 3′-terminal nucleotides by N-ethyl-N-coupled nucleosides. Molecular dynamics simulations of 3′-N-ethyl-N-modified DNA:Klenow fragment complexes suggested that this kind of alteration has negative effects on the correct positioning of the adjacent scissile phosphodiester bond at the active site of the enzyme, and accordingly was expected to protect the oligonucleotide from degradation. We verified that these modifications conferred complete resistance to 3′-exonucleases. Furthermore, cellular RNAi experiments with 3′-N-ethyl-N-modified siRNAs showed that these modifications were compatible with the RNAi machinery. Overall, our experimental and theoretical studies strongly suggest that these modified oligonucleotides could be valuable for therapeutic applications.

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