Ausín Cristina, Grajkowski Andrzej, Cieślak Jacek, Gapeev Alexei, Beaucage Serge L
Food and Drug Administration, Bethesda, Maryland, USA.
Curr Protoc Nucleic Acid Chem. 2010 Dec;Chapter 4:Unit 4.42.. doi: 10.1002/0471142700.nc0442s43.
This unit describes the preparation of alkylthioalkylated and formamidoalkylated alcohols, an amidoalkylated alcohol, a hydroxylalkylated phosphoramidate, and their phosphoramidothioate derivatives, all of which have been identified as heat-sensitive thiophosphate-protecting groups in the development of thermolytic immunostimulatory DNA prodrugs. The alcohols are converted to their deoxyribonucleoside phosphoramidite derivatives, which are then used in the preparation of thermosensitive dinucleoside phosphorothioates. The thiophosphate-protecting groups of these dinucleoside phosphorothioates presumably undergo thermolytic cyclodeesterification at elevated temperature under essentially neutral conditions to release the desired phosphorothioate diester function. On the basis of their thermolytic deprotection kinetics, one can identify those thiophosphate-protecting groups that (i) may be useful for thiophosphate protection of CpG motifs of immunostimulatory DNA oligonucleotides (CpG ODNs); (ii) are suitable for protection of phosphodiester functions flanking the CpG motifs; and (iii) offer adequate protection of terminal phosphodiester functions against ubiquitous extracellular and intracellular exonucleases that may be found in biological environments.
本单元描述了烷硫基烷基化醇、甲酰胺基烷基化醇、酰胺基烷基化醇、羟烷基化亚磷酰胺及其硫代磷酸酯衍生物的制备方法。在热解免疫刺激DNA前药的开发过程中，所有这些化合物都被确定为热敏性硫代磷酸酯保护基团。这些醇被转化为其脱氧核糖核苷亚磷酰胺衍生物，然后用于制备热敏性二核苷硫代磷酸酯。这些二核苷硫代磷酸酯的硫代磷酸酯保护基团可能在基本中性的条件下，于高温下发生热解环酯交换反应，以释放所需的硫代磷酸二酯官能团。基于它们的热解脱保护动力学，可以确定那些硫代磷酸酯保护基团：（i）可能对免疫刺激DNA寡核苷酸（CpG ODNs）的CpG基序的硫代磷酸酯保护有用；（ii）适用于保护CpG基序侧翼的磷酸二酯官能团；（iii）能为末端磷酸二酯官能团提供充分保护，使其免受生物环境中可能存在的普遍存在的细胞外和细胞内核酸外切酶的作用。
