Proteins of the bromodomain and extra-terminal (BET) domain family are epigenetic readers that bind acetylated histones through their bromodomains to regulate gene transcription. Dual-bromodomain BET inhibitors (DbBi) that bind with similar affinities to the first (BD1) and second (BD2) bromodomains of BRD2, BRD3, BRD4 and BRDt have displayed modest clinical activity in monotherapy cancer trials. A reduced number of thrombocytes in the blood (thrombocytopenia) as well as symptoms of gastrointestinal toxicity are dose-limiting adverse events for some types of DbBi^1,2,3,4,5. Given that similar haematological and gastrointestinal defects were observed after genetic silencing of Brd4 in mice⁶, the platelet and gastrointestinal toxicities may represent on-target activities associated with BET inhibition. The two individual bromodomains in BET family proteins may have distinct functions^7,8,9 and different cellular phenotypes after pharmacological inhibition of one or both bromodomains have been reported^10,11, suggesting that selectively targeting one of the bromodomains may result in a different efficacy and tolerability profile compared with DbBi. Available compounds that are selective to individual domains lack sufficient potency and the pharmacokinetics properties that are required for in vivo efficacy and tolerability assessment^10,11,12,13. Here we carried out a medicinal chemistry campaign that led to the discovery of ABBV-744, a highly potent and selective inhibitor of the BD2 domain of BET family proteins with drug-like properties. In contrast to the broad range of cell growth inhibition induced by DbBi, the antiproliferative activity of ABBV-744 was largely, but not exclusively, restricted to cell lines of acute myeloid leukaemia and prostate cancer that expressed the full-length androgen receptor (AR). ABBV-744 retained robust activity in prostate cancer xenografts, and showed fewer platelet and gastrointestinal toxicities than the DbBi ABBV-075¹⁴. Analyses of RNA expression and chromatin immunoprecipitation followed by sequencing revealed that ABBV-744 displaced BRD4 from AR-containing super-enhancers and inhibited AR-dependent transcription, with less impact on global transcription compared with ABBV-075. These results underscore the potential value of selectively targeting the BD2 domain of BET family proteins for cancer therapy.

溴结构域和末端外 (BET) 结构域家族的蛋白质是表观遗传阅读器，它们通过其溴结构域结合乙酰化组蛋白以调节基因转录。双溴结构域 BET 抑制剂 (DbBi) 与 BRD2、BRD3、BRD4 和 BRDt 的第一个 (BD1) 和第二个 (BD2) 溴结构域具有相似的亲和力，在单一疗法癌症试验中显示出适度的临床活性。血液中血小板数量减少（血小板减少症）以及胃肠道毒性症状是某些类型的 DbBi ^1,2,3,4,5的剂量限制性不良事件。鉴于在小鼠中Brd4基因沉默后观察到类似的血液学和胃肠道缺陷⁶，血小板和胃肠道毒性可能代表与 BET 抑制相关的靶向活性。BET 家族蛋白中的两个单独的溴结构域可能具有不同的功能^{7,8,9并且在一个或两个溴结构域的药理学抑制作用已被报道10,11}后具有不同的细胞表型，这表明选择性靶向其中一个溴结构域可能导致不同的功效和与 DbBi 相比的耐受性概况。对单个域具有选择性的可用化合物缺乏足够的效力和体内功效和耐受性评估所需的药代动力学特性^10,11,12,13. 在这里，我们开展了一项药物化学活动，从而发现了 ABBV-744，这是一种具有药物样特性的 BET 家族蛋白 BD2 结构域的高效选择性抑制剂。与 DbBi 诱导的广泛细胞生长抑制相比，ABBV-744 的抗增殖活性在很大程度上但不仅限于表达全长雄激素受体 (AR) 的急性髓性白血病和前列腺癌的细胞系。 . ABBV-744 在前列腺癌异种移植物中保留了强大的活性，并且显示出比 DbBi ABBV-075 更少的血小板和胃肠道毒性¹⁴. 对 RNA 表达和染色质免疫沉淀进行测序后的分析表明，与 ABBV-075 相比，ABBV-744 将 BRD4 从含有 AR 的超级增强子中置换出来并抑制 AR 依赖性转录，对整体转录的影响较小。这些结果强调了选择性靶向 BET 家族蛋白的 BD2 结构域用于癌症治疗的潜在价值。