Hydroxyalkylated phosphoramidate, phosphoramidothioate and phosphorodiamidothioate derivatives as thiophosphate protecting groups in the development of thermolytic DNA prodrugs

Article abstract of DOI:10.1039/b9nj00692c

The hydroxyalkylated phosphoramidate 4a, phosphoramidothioates 4b, 4e-j, and phosphorodiamidothioates 4c and 4d have been identified as a new class of heat-sensitive thiophosphate protecting groups in the development of thermolytic immunomodulatory DNA prodrugs. These alcohols are converted to their deoxyribonucleoside phosphoramidite derivatives 6a-j, which are then used in the preparation of the thermosensitive dinucleoside phosphorothioates 7a-j. The negatively charged thiophosphate protecting groups of 7a-b and 7e-j presumably undergo thermolytic cyclodeesterification at elevated temperature under essentially neutral conditions. The thiophosphate protecting groups of 7e and 7f show relatively rapid deprotection kinetics at 37 °C (t_1/2 = 20 and 42 h, respectively) and are therefore suitable for the protection of phosphodiester functions flanking the CpG motifs of immunomodulatory DNA sequences, whereas the thiophosphate protecting groups of 7g-j with thermolytic deprotection half-lives in the range of 94-265 h at 37 °C are more appropriate for the thiophosphate protection of CpG motifs. Furthermore, the thermostability of the group protecting the thiophosphate function of 7a (t _1/2 = 82 min at 90 °C) should offer adequate protection of the 5'- and/or 3'-terminal phosphodiester functions of DNA prodrugs against ubiquitous extracellular and intracellular exonucleases.