Aberrant activation of multiple complex signaling pathways underlies the pathogenesis of rhabdomyosarcoma (RMS), which remains a cause of mortality in approximately 30% of children with RMS. Bromodomain and extraterminal (BET) domain chromatin remodeling regulates several of these pathways. Here, we targeted bromodomain 4 (BRD4) in combination with another molecular metabolic tumor driver, the Akt/mTOR signaling pathway, to provide a highly effective treatment for this neoplasm. We demonstrated that a nexus of these two molecular pathways underlies RMS pathogenesis. Our data show that the combined inhibition of the BET bromodomain and mTORC1/2 signaling abrogates aggressive RMS growth. Thus, the bromodomain inhibitor RVX-208 significantly augmented the therapeutic effects of the dual mTORC1/2 inhibitors, OSI-027 and PP242, both in vitro and in a human xenograft murine model. Drug-treated residual tumors showed a decrease in the activation of underlying signaling mechanisms characterized by a reduction in the expression of p-AKT, p-mTOR, p-p70S6K, cyclin D1, and proliferation. Our ChIP-seq data demonstrated that RVX-208 effectively blocked BRD4 occupancy on its target promoters. ChIP-qPCR assays further confirmed that RVX-208 treatment resulted in a significant decrease in H3K27ac and H4K8ac signals at their target loci. While single RVX-208 treatment induces apoptosis and a single mTORC1/2 inhibitor induces macropinocytosis, their combined treatment led to necroptosis-mediated cell death. These data suggest that combined treatment with drugs targeting BRD4 and mTORC1/2 may be an effective therapeutic intervention for drug-resistant RMS.

中文翻译：

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BET 溴结构域和 mTORC1/2 的联合抑制通过转换细胞死亡机制为横纹肌肉瘤提供治疗优势

多个复杂信号通路的异常激活是横纹肌肉瘤 (RMS) 发病机制的基础，它仍然是约 30% RMS 儿童死亡的原因。溴结构域和末端外 (BET) 结构域染色质重塑调节其中的几个途径。在这里，我们将溴结构域 4 (BRD4) 与另一种分子代谢肿瘤驱动因素 Akt/mTOR 信号通路结合起来，为这种肿瘤提供高效的治疗方法。我们证明这两种分子途径的联系是 RMS 发病机制的基础。我们的数据表明，BET 溴结构域和 mTORC1/2 信号传导的联合抑制可消除 RMS 的侵袭性生长。因此，无论是在体外还是在人异种移植小鼠模型中，溴结构域抑制剂 RVX-208 均显着增强了双重 mTORC1/2 抑制剂 OSI-027 和 PP242 的治疗效果。药物治疗后的残留肿瘤显示出潜在信号机制的激活减少，其特征是 p-AKT、p-mTOR、p-p70S6K、细胞周期蛋白 D1 的表达和增殖减少。我们的 ChIP-seq 数据表明 RVX-208 有效阻止 BRD4 占据其目标启动子。ChIP-qPCR 测定进一步证实 RVX-208 处理导致目标位点的 H3K27ac 和 H4K8ac 信号显着减少。虽然单一 RVX-208 治疗会诱导细胞凋亡，而单一 mTORC1/2 抑制剂会诱导巨胞饮作用，但它们的联合治疗会导致坏死性凋亡介导的细胞死亡。这些数据表明，与靶向 BRD4 和 mTORC1/2 的药物联合治疗可能是治疗耐药 RMS 的有效治疗干预措施。