Small-molecule targeting of brachyury transcription factor addiction in chordoma [rnaseq_compound]

Chordoma is a primary bone cancer with no approved therapy. The identification of therapeutic targets in this disease has been challenging due to the infrequent occurrence of clinically actionable somatic mutations in chordoma tumors. Here we describe the discovery of therapeutically targetable chordoma dependencies via genome-scale CRISPR-Cas9 screening and focused small-molecule sensitivity profiling. These systematic approaches reveal that the developmental transcription factor T (brachyury; TBXT) is the top selectively essential gene in chordoma, and that transcriptional cyclin-dependent kinase (CDK) inhibitors targeting CDK7/12/13 and CDK9 potently suppress chordoma cell proliferation. In other cancer types, transcriptional CDK inhibitors have been observed to downregulate highly expressed, enhancer-associated oncogenic transcription factors (TFs). In chordoma, we find that T is associated with a 1.5-Mb region containing “super-enhancers” and is the most highly expressed super-enhancer-associated TF. Strikingly, transcriptional CDK inhibition leads to preferential and concentration-dependent downregulation of cellular brachyury protein levels in all models tested. Together, these data demonstrate small-molecule targeting of brachyury TF addiction in chordoma, identify a mechanism of T gene regulation that underlies this therapeutic strategy and provide a blueprint for applying systematic genetic and chemical screening approaches to discover vulnerabilities in genomically quiet cancers. Overall design: RNA-sequencing of THZ1-, actinomycin D-, or DMSO-treated UM-Chor1 chordoma cells. Each compound was tested at two concentrations. Two biological replicates were generated for each condition.

脊索瘤（Chordoma）是一种原发骨恶性肿瘤，目前尚无获批的治疗方案。由于脊索瘤肿瘤中临床可操作的体细胞突变发生率极低，该疾病治疗靶点的鉴定颇具挑战。本研究通过全基因组CRISPR-Cas9筛选与聚焦型小分子敏感性分析，发掘了可靶向治疗的脊索瘤依赖基因。这些系统性研究方法揭示，发育转录因子T（brachyury；TBXT）是脊索瘤中选择性必需性排名第一的基因；同时靶向CDK7/12/13的转录周期蛋白依赖性激酶（CDK）抑制剂与CDK9抑制剂，可强效抑制脊索瘤细胞增殖。在其他癌种中，转录CDK抑制剂已被证实可下调高表达、增强子相关的致癌转录因子（TFs）。在脊索瘤中，我们发现T基因与一段包含“超级增强子（super-enhancers）”的1.5 Mb区域相关，且是表达量最高的超级增强子相关转录因子。值得注意的是，在所有受试模型中，转录CDK抑制剂可呈浓度依赖性优先下调细胞内brachyury蛋白水平。综上，本研究证实了可通过小分子靶向脊索瘤中brachyury转录因子的成瘾性，阐明了支撑该治疗策略的T基因调控机制，并为应用系统性遗传与化学筛选方法发掘基因组静默型癌症的脆弱位点提供了研究范式。整体实验设计：经THZ1、放线菌素D或二甲基亚砜（DMSO）处理的UM-Chor1脊索瘤细胞的RNA测序（RNA-sequencing）分析。每种化合物均设置两个浓度梯度，每个处理条件设置两个生物学重复。