Within the last decade, the Bromodomain and Extra-Terminal domain family (BET) of proteins have emerged as promising drug targets in diverse clinical indications including oncology, auto-immune disease, heart failure, and male contraception. The BET family consists of four isoforms (BRD2, BRD3, BRD4, and BRDT/BRDT6) which are distinguished by the presence of two tandem bromodomains (BD1 and BD2) that independently recognize acetylated-lysine (KAc) residues and appear to have distinct biological roles. BET BD1 and BD2 bromodomains differ at five positions near the substrate binding pocket: the variation in the ZA channel induces different water networks nearby. We designed a set of congeneric 2- and 3-heteroaryl substituted tetrahydroquinolines (THQ) to differentially engage bound waters in the ZA channel with the goal of achieving bromodomain selectivity. SJ830599 (9) showed modest, but consistent, selectivity for BRD2-BD2. Using isothermal titration calorimetry, we showed that the binding of all THQ analogs in our study to either of the two bromodomains was enthalpy driven. Remarkably, the binding of 9 to BRD2-BD2 was marked by negative entropy and was entirely driven by enthalpy, consistent with significant restriction of conformational flexibility and/or engagement with bound waters. Co-crystallography studies confirmed that 9 did indeed stabilize a water-mediated hydrogen bond network. Finally, we report that 9 retained cytotoxicity against several pediatric cancer cell lines with EC₅₀ values comparable to BET inhibitor (BETi) clinical candidates.

在过去的十年中，蛋白质的溴结构域和末端结构域家族 (BET) 已成为肿瘤、自身免疫性疾病、心力衰竭和男性避孕等多种临床适应症的有前景的药物靶点。 BET 家族由四种同工型（BRD2、BRD3、BRD4 和 BRDT/BRDT6）组成，其特征在于存在两个串联溴结构域（BD1 和 BD2），它们独立识别乙酰化赖氨酸 (KAc) 残基，并且似乎具有不同的生物学特性。角色。 BET BD1 和 BD2 溴结构域在底物结合口袋附近的五个位置上有所不同：ZA 通道的变化导致附近不同的水网络。我们设计了一组同源的 2- 和 3- 杂芳基取代的四氢喹啉 (THQ)，以差异化地结合 ZA 通道中的结合水，以实现溴结构域选择性。 SJ830599 ( 9 ) 对 BRD2-BD2 显示出适度但一致的选择性。使用等温滴定量热法，我们表明我们研究中的所有 THQ 类似物与两个溴结构域中的任何一个的结合都是焓驱动的。值得注意的是， 9与 BRD2-BD2 的结合以负熵为标志，并且完全由焓驱动，这与构象灵活性和/或与结合水的结合的显着限制一致。共晶研究证实9确实稳定了水介导的氢键网络。最后，我们报告9对几种儿科癌细胞系保留了细胞毒性，其 EC ₅₀值与 BET 抑制剂 (BETi) 临床候选药物相当。