Assignment of the absolute configuration at stereogenic phosphorus atoms in P-diastereomers of dithymidyl-(N3′→P5′)-phosphoramidothioate

P-diastereomers of dithymidyl-(N3′→P5′)-phosphoramidothioate (TNPST) were synthesized by condensation of chromatographically separated P-diastereomers of 5′-O-DMT-2′,3′-dideoxythymidine-3′-N-(2-thio-4,4-dimethyl-1,3,2-oxathiaphospholanes) (OTP derivatives, assigned as fast and slow-eluting diastereoisomers) with 3′-O-acetylthymidine. The resulting (TNPST) dimers were transformed into S-methyl derivatives by a standard treatment with methyl iodide in a pyridine/acetonitrile solution. The absolute configuration at phosphorus atoms of each S-methylated diastereoisomer was assigned by the analysis of the 2D NMR ROESY cross-peaks, analogously to the approach already published for 2′-deoxyribonucleoside methylphosphonamidates NNPMeN. To the TNPST dimers originating from the fast- and slow-OTP substrate the SP and RP absolute configurations were assigned, respectively. The obtained NMR results are well supported by molecular modeling calculations.