干扰素基因刺激剂（STING）已成为癌症免疫治疗的一个有前景的靶标。 2'3'-cGAMP 是 STING 的天然激动剂，通过刺激免疫细胞显示出抗癌活性，但它在体内容易被水解酶降解。因此，在临床上作为免疫疗法进行评估的环状二核苷酸类似物含有水解稳定的硫代磷酸酯部分，其中硫部分位于含磷酸盐的环外。然而，这些硫代磷酸酯的合成会产生非对映体，这给分离带来了挑战。另一种硫代磷酸酯（称为内-S-硫代磷酸酯），其中硫原子位于环状磷酸环的内部（即5'-S-硫代磷酸酯键），在磷上不具有手性，因此不会造成非对映异构体分离问题。在此，我们报告了新型 5'-内切硫代磷酸酯取代的 2'3'cGAMP 类似物的设计和合成，该类似物对外核苷酸磷酸二酯酶 I（ENPP1，一种哺乳动物磷酸二酯酶）和痘病毒免疫核酸酶（痘病毒，痘病毒中的一种磷酸二酯酶）均具有水解稳定性。 ）但保留了 STING-TBK1-IRF 激活，与 THP1 单核细胞中的临床候选药物 ADU-S100 相当。本期刊版权所有 © 英国皇家化学学会。

The stimulator of interferon genes (STING) has emerged as a promising target for cancer immunotherapy. 2'3'-cGAMP, a natural agonist of STING, shows anticancer activity via stimulation of immune cells but it is susceptible to degradation in vivo by hydrolytic enzymes. Consequently, the cyclic dinucleotide analogues that are being evaluated in the clinic as immunotherapies contain the hydrolytically stable phosphorothioate moiety, whereby the sulfur moiety is exo to the phosphate containing ring. The synthesis of these phosphorothioates however produces diastereomers, which presents separation challenges. An alternative phosphorothioate (referred to as endo-S-phosphorothioate) whereby the sulfur atom is endo to the cyclic phosphate ring (i.e. 5'-S-phosphorothioester linkage) would not have chirality at phosphorus and hence not pose diastereomer separation problems. Herein, we report the design and synthesis of novel 5'-endo-phosphorothioate substituted 2'3'cGAMP analogues that are hydrolytically stable towards both ectonucleotide phosphodiesterase I (ENPP1, a mammalian phosphodiesterase) and poxvirus immune nucleases (poxin, a phosphodiesterase in Poxvirus) but retains STING-TBK1-IRF activation, comparable to clinical candidate, ADU-S100 in THP1 monocytes.This journal is © The Royal Society of Chemistry.