A Strategy for the Stereoselective Preparation of Thymidine Phosphorothioates with the (<i>R</i>) or the (<i>S</i>) Configuration at the Stereogenic Phosphorus Atom and Their Application to the Synthesis of Oligodeoxyribonucleotides with Stereochemically Pure Phosphate/Phosphorothioate Chimeric Backbones

Abstract

Abstract This paper describes a new method for the highly stereoselective preparation of dithymidine phosphorothioates (TpsT) with the ( R ) or the ( S ) configuration at the stereogenic phosphorus atom [( R p)‐ or ( S p)‐TpsT, respectively], together with the synthesis of oligodeoxyribonucleotides with stereochemically pure phosphate/phosphorothioate mixed backbones through the use of ( R p)‐ or ( S p)‐TpsT as building blocks.Stereochemically pure ( R p)‐ or ( S p)‐TpsT was produced through a five‐step process: 1) 1 H ‐tetrazole‐promoted thermodynamic equilibration of a diastereomeric mixture of allyl ( R p)‐ and ( S p)‐thymidine 3′,5′‐cyclic phosphate to give the ( S p) isomer as the major component, 2) stereospecific sulfurization of the allyl ( S p)‐thymidine 3′,5′‐cyclic phosphate to afford an allyl ( R p)‐thymidine 3′,5′‐cyclic phosphorothioate, 3) regioselective and stereoselective methanolysis of the allyl ( R p)‐thymidine 3′,5′‐cyclic phosphorothioate to provide an allyl methyl ( R p)‐thymidine 3′‐phosphorothioate as the main product, 4) stereospecific removal of the methyl group from the phosphorothioate moiety to give an allyl ( R p)‐5′‐ O ‐dimethoxytritylthymidine 3′‐phosphorothioate diester, or stereospecific removal of the allyl group from the phosphate moiety to form a methyl ( S p)‐5′‐ O ‐dimethoxytritylthymidine 3′‐phosphorothioate diester, and 5) stereospecific condensation of these ( R p)‐ and ( S p)‐diesters with a 5′‐ O ‐free thymidine in a Mitsunobu reaction to produce the allyl (3′–5′)‐linked ( S p)‐dithymidine phosphorothioate and the methyl (3′–5′)‐linked ( R p)‐dithymidine phosphorothioate, respectively. The resulting ( R p)‐ and ( S p)‐dithymidine phosphorothioates were converted into their 3′‐phosphoramidites. Oligodeoxyribonucleotides with stereochemically pure phosphate/phosphorothioate‐mixed backbones were then synthesized by use of these phosphoramidites as building blocks. (© Wiley‐VCH Verlag GmbH &amp; Co. KGaA, 69451 Weinheim, Germany, 2006)