Abstract Targeting bromodomain-containing protein 4 (BRD4) has been proved to be an effective strategy for cancer therapy. To date, numerous BRD4 inhibitors and degraders have been identified, some of which have advanced into clinical trials. In this work, a focused library of new [1,2,4]triazolo[1,5-a]pyrimidine derivatives were discovered to be able to inhibit BRD4. WS-722 inactivated BRD4 (BD1/BD2), BRD2 (BD1/BD2) and BRD3 (BD1/BD2) broadly with the IC50 values less than 5 μmol/L. Besides, WS-722 inhibited growth of THP-1 cells with an IC50 value of 3.86 μmol/L. Like (+)-JQ1, WS-722 inhibited BRD4 in a reversible manner and enhanced protein stability. Docking studies showed that WS-722 occupied the central acetyl-lysine (Kac) binding cavity and formed a hydrogen bond with Asn140. In THP-1 cells, WS-722 showed target engagement to BRD4. Cellular effects of WS-722 on THP-1 cells were also examined, showing that WS-722 could block c-MYC expression, induce G0/G1 phase arrest and p21 up-regulation, and promote differentiation of THP-1 cells. BRD4 inhibition by WS-722 resulted in cell apoptosis and up-regulated expression of cleaved caspased-3/7 and PARP in THP-1 cell lines. The [1,2,4]triazolo[1,5-a]pyrimidine is a new template for the development of new BRD4 inhibitors.

中文翻译：

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发现[1,2,4]三唑并[1,5-a]嘧啶衍生物作为新型含溴结构域的蛋白4（BRD4）抑制剂

摘要靶向含溴结构域的蛋白质4（BRD4）已被证明是一种有效的癌症治疗策略。迄今为止，已鉴定出许多BRD4抑制剂和降解剂，其中一些已进入临床试验。在这项工作中，发现了新的[1,2,4]三唑并[1,5-a]嘧啶衍生物的聚焦文库能够抑制BRD4。WS-722使BRD4（BD1 / BD2），BRD2（BD1 / BD2）和BRD3（BD1 / BD2）广泛失活，IC50值小于5μmol/ L。此外，WS-722抑制THP-1细胞的生长，IC50值为3.86μmol/ L。像（+）-JQ1一样，WS-722以可逆方式抑制BRD4，并增强了蛋白质稳定性。对接研究表明，WS-722占据了中央乙酰赖氨酸（Kac）的结合腔，并与Asn140形成了氢键。在THP-1细胞中，WS-722显示出对BRD4的靶标参与。还检查了WS-722对THP-1细胞的细胞作用，表明WS-722可以阻断c-MYC表达，诱导G0 / G1期阻滞和p21上调，并促进THP-1细胞的分化。WS-722抑制BRD4导致细胞凋亡，并在THP-1细胞系中上调了caspased-3 / 7和PARP的表达。[1,2,4]三唑并[1,5-a]嘧啶是开发新的BRD4抑制剂的新模板。