The blockade of A_2A adenosine receptor (A_2AAR) activates immunostimulatory response through regulating signaling in tumor microenvironment. Thus, A_2AAR has been proposed as a promising target for cancer immunotherapy. In this work, we designed a new series of benzo[4,5]imidazo[1,2-a]pyrazin-1-amine derivatives bearing an amide substitution at 3-position to obtain potent antitumor antagonist in vivo. The structure–activity relationship studies were performed by molecular modeling and radioactive assay. The in vitro anticancer activities were evaluated by 3′,5′-cyclic adenosine monophosphate (cAMP) functional and T cell activation assay. The most potent compound 12o·2HCl showed much higher affinity toward A_2AAR (K_i = 0.08 nM) and exhibited more significant in vitro immunostimulatory anticancer activity than clinical antagonist AZD4635. More importantly, 12o·2HCl significantly inhibited the growth of triple-negative breast cancer by reversing immunosuppressive tumor microenvironment in the xenograft mouse model without severe toxicity at the testing dose. These results make 12o·2HCl a promising immunotherapy anticancer drug candidate.

中文翻译：

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发现新型苯并[4,5]咪唑并[1,2-a]吡嗪-1-胺-3-酰胺-酮衍生物作为抗癌人A2A腺苷受体拮抗剂

A _2A腺苷受体 (A _2A AR) 的阻断通过调节肿瘤微环境中的信号传导来激活免疫刺激反应。因此，A _2A AR 已被提议作为癌症免疫治疗的有希望的靶标。在这项工作中，我们设计了一系列新的苯并[4,5]咪唑并[1,2 - a ]吡嗪-1-胺衍生物，在3位具有酰胺取代，以在体内获得有效的抗肿瘤拮抗剂。通过分子建模和放射性测定进行构效关系研究。通过3',5'-环磷酸腺苷 (cAMP) 功能和 T 细胞活化测定评估体外抗癌活性。最强的化合物12o·2HCl与临床拮抗剂 AZD4635 相比，对 A _2A AR ( K _i = 0.08 nM) 显示出更高的亲和力，并表现出更显着的体外免疫刺激抗癌活性。更重要的是，12o·2HCl通过逆转异种移植小鼠模型中的免疫抑制性肿瘤微环境，显着抑制三阴性乳腺癌的生长，且在试验剂量下无严重毒性。这些结果使12o·2HCl成为一种很有前途的免疫治疗抗癌药物候选者。