Synthesis of High-Quality Antisense Drugs. Addition of Acrylonitrile to Phosphorothioate Oligonucleotides: Adduct Characterization and Avoidance

It is demonstrated that the acrylonitrile (AN) generated during the ammonolysis step of oligonucleotide manufacture selectively adds to thymine residue present in ISIS 2302 to give a full-length oligonucleotide in which thymine is replaced by an N 3 -cyanoethylthymine residue. Treatment of support-bound ISIS 2302 with a solution of triethylamine in CH 3 CN before ammonolysis is sufficient to prevent formation of this class of impurity.