Triple-negative breast cancers contain a spectrum of epithelial and mesenchymal phenotypes. SUM-229PE cells represent a model for this heterogeneity, maintaining both epithelial and mesenchymal subpopulations that are genomically similar but distinct in gene expression profiles. We identified differential regions of open chromatin in epithelial and mesenchymal cells that were strongly correlated with regions of H3K27ac. Motif analysis of these regions identified consensus sequences for transcription factors that regulate cell identity. Treatment with the MEK inhibitor trametinib induced enhancer remodeling that is associated with transcriptional regulation of genes in epithelial and mesenchymal cells. Motif analysis of enhancer peaks downregulated in response to chronic treatment with trametinib identified AP-1 motif enrichment in both epithelial and mesenchymal subpopulations. Chromatin immunoprecipitation sequencing (ChIP-seq) of JUNB identified subpopulation-specific localization, which was significantly enriched at regions of open chromatin. These results indicate that cell identity controls localization of transcription factors and chromatin-modifying enzymes to enhancers for differential control of gene expression. We identified increased H3K27ac at an enhancer region proximal to CXCR7, a G-protein–coupled receptor that increased 15-fold in expression in the epithelial subpopulation during chronic treatment. RNAi knockdown of CXCR7 inhibited proliferation in trametinib-resistant cells. Thus, adaptive resistance to chronic trametinib treatment contributes to proliferation in the presence of the drug. Acquired amplification of KRAS following trametinib dose escalation further contributed to POS cell proliferation. Adaptive followed by acquired gene expression changes contributed to proliferation in trametinib-resistant cells, suggesting inhibition of early transcriptional reprogramming could prevent resistance and the bypass of targeted therapy. Implications: We defined the differential responses to trametinib in subpopulations of a clinically relevant in vitro model of TNBC, and identified both adaptive and acquired elements that contribute to the emergence of drug resistance mediated by increased expression of CXCR7 and amplification of KRAS.

中文翻译：

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三阴性乳腺癌亚群对 MEK 抑制剂的离散适应性反应

三阴性乳腺癌包含一系列上皮和间充质表型。SUM-229PE 细胞代表了这种异质性的模型，维持基因组相似但基因表达谱不同的上皮和间充质亚群。我们确定了与 H3K27ac 区域密切相关的上皮和间充质细胞中开放染色质的差异区域。这些区域的基序分析确定了调节细胞身份的转录因子的共有序列。用 MEK 抑制剂曲美替尼治疗诱导增强子重塑，这与上皮细胞和间充质细胞中基因的转录调控有关。响应于曲美替尼的慢性治疗而下调的增强子峰的基序分析确定了上皮和间充质亚群中的 AP-1 基序富集。JUNB 的染色质免疫沉淀测序 (ChIP-seq) 确定了亚群特异性定位，该定位在开放染色质区域显着富集。这些结果表明，细胞身份控制转录因子和染色质修饰酶对增强子的定位，以对基因表达进行差异控制。我们发现在靠近 CXCR7 的增强子区域增加了 H3K27ac，CXCR7 是一种 G 蛋白偶联受体，在慢性治疗期间上皮亚群中的表达增加了 15 倍。CXCR7 的 RNAi 敲低抑制了曲美替尼耐药细胞的增殖。因此，对慢性曲美替尼治疗的适应性耐药有助于药物存在时的增殖。曲美替尼剂量增加后获得的 KRAS 扩增进一步促进了 POS 细胞增殖。适应性随后获得性基因表达变化有助于曲美替尼耐药细胞的增殖，这表明抑制早期转录重编程可以防止耐药性和靶向治疗的绕过。启示：我们在临床相关的 TNBC 体外模型的亚群中定义了对曲美替尼的不同反应，并确定了适应性和获得性因素，这些因素有助于由 CXCR7 表达增加和 KRAS 扩增介导的耐药性的出现。曲美替尼剂量增加后获得的 KRAS 扩增进一步促进了 POS 细胞增殖。适应性随后获得性基因表达变化有助于曲美替尼耐药细胞的增殖，这表明抑制早期转录重编程可以防止耐药性和靶向治疗的绕过。启示：我们在临床相关的 TNBC 体外模型的亚群中定义了对曲美替尼的不同反应，并确定了适应性和获得性因素，这些因素有助于由 CXCR7 表达增加和 KRAS 扩增介导的耐药性的出现。曲美替尼剂量增加后获得的 KRAS 扩增进一步促进了 POS 细胞增殖。适应性随后获得性基因表达变化有助于曲美替尼耐药细胞的增殖，这表明抑制早期转录重编程可以防止耐药性和靶向治疗的绕过。启示：我们在临床相关的 TNBC 体外模型的亚群中定义了对曲美替尼的不同反应，并确定了适应性和获得性因素，这些因素有助于由 CXCR7 表达增加和 KRAS 扩增介导的耐药性的出现。表明抑制早期转录重编程可以防止耐药性和靶向治疗的绕过。启示：我们在临床相关的 TNBC 体外模型的亚群中定义了对曲美替尼的不同反应，并确定了适应性和获得性因素，这些因素有助于由 CXCR7 表达增加和 KRAS 扩增介导的耐药性的出现。表明抑制早期转录重编程可以防止耐药性和靶向治疗的绕过。启示：我们在临床相关的 TNBC 体外模型的亚群中定义了对曲美替尼的不同反应，并确定了适应性和获得性因素，这些因素有助于由 CXCR7 表达增加和 KRAS 扩增介导的耐药性的出现。