Improved Impurity Profile of Phosphorothioate Oligonucleotides Through the Use of Dimeric Phosphoramidite Synthons

Abstract

Abstract Phosphorothioate oligonucleotides synthesized through assembly of dimeric phosphoramidite synthons show a significantly improved impurity profile compared to oligomers synthesized through coupling of standard monomer phosphoramidites. A greater than 70% reduction of the (n-1)-mer population and a ca. 50% reduction of phosphodiester linkages has been achieved. Notes Nomenclature: T19 denotes a nonadecathymidylate phosphorothioate, (T2)9T and (TdC)9T indicate monomer assembly, (T 2)9T, (TdC)9T indicate dimer assembly, TdC indicates S-CE protection Cruachem, Inc., Glasgow, Scotland Columns: dT-CPG (1μmol) from Glen Research. Standard detritylation, capping and 1H-tetrazole solutions (Applied Biosystems). Amidites (0.1 M in CH3CN), coupling time 200 s. Sulfurization: 3H-1, 2-benzodithiol-3-one-1, 1-dioxide (0.2 M in CH3CN, R. I. Chemical, Orange, CA), 900 s (not optimized) Precise quantitation below 1% is difficult due to lack of baseline resolution. The actual value may be significantly lower.