Abstract：
Ovarian cancer, a significant contributor to cancer-related mortality, exhibits limited responsiveness to hormonal therapies targeting the estrogen receptor (ERα). This study aimed to elucidate the mechanisms behind ERα resistance to the therapeutic drug Fulvestrant (ICI182780 or ICI). Notably, compared to the cytoplasmic version, nuclear ERα was minimally degraded by ICI, suggesting a mechanism for drug resistance via the protective confines of the nuclear substructures. Of these substructures, we identified a 1.3MDa Megacomplex comprising transcription factors ERα, FOXA1, and PITX1 using size exclusion chromatography (SEC) in the ovarian cancer cell line, PEO4. ChIP-seq revealed these factors colocalized at 6,775 genomic positions representing sites of Megacomplex formation. Megacomplex ERα exhibited increased resistance to degradation by ICI compared to cytoplasmic and nuclear ERα. A small molecule inhibitor of active chromatin and super-enhancers, JQ1, in combination with ICI significantly enhanced ERα degradation from Megacomplex as revealed by SEC and ChIP-seq. Interestingly, this combination degraded both the cytoplasmic as well as nuclear ERa. Pathway enrichment analysis showed parallel results for RNA-seq gene sets following Estradiol, ICI, or ICI plus JQ1 treatments as those defined by Megacomplex binding identified through ChIP-seq. Furthermore, similar pathway enrichments were confirmed in mass-spec analysis of the Megacomplex macromolecule fractions after modulation by Estradiol or ICI. These findings implicate Megacomplex in ERα-driven ovarian cancer chromatin regulation. This combined treatment strategy exhibited superior inhibition of cell proliferation and viability. Therefore, by uncovering ERα\'s resistance within the Megacomplex, the combined ICI plus JQ1 treatment elucidates a novel drug treatment vulnerability.
摘要：
卵巢癌,癌症相关死亡率的重要贡献者，对靶向雌激素受体（ERα）的激素疗法表现出有限的反应。这项研究旨在阐明ERα对治疗药物Fulvestrant（ICI182780或ICI）的耐药机制。值得注意的是,与细胞质版本相比，核ERα被ICI最低限度地降解，表明通过核子结构的保护范围产生抗药性的机制。在这些子结构中，我们鉴定了一个1.3MDaMegacomplex，包含转录因子ERα，FOXA1和PITX1在卵巢癌细胞系中使用尺寸排阻色谱（SEC），PEO4.ChIP-seq揭示了这些因子在6,775个基因组位置共同定位，代表Megacomplex形成位点。与细胞质和核ERα相比，MegacomplexERα对ICI降解的抗性增加。一种活性染色质和超级增强剂的小分子抑制剂，如SEC和ChIP-seq所揭示的,JQ1与ICI组合显著增强了来自Megacomplex的ERα降解。有趣的是,这种组合降解了细胞质和核ERa。通路富集分析显示雌二醇后RNA-seq基因集的平行结果，ICI或ICI加JQ1处理如通过ChIP-seq鉴定的Megacomplex结合所定义的那些。此外，在雌二醇或ICI调节后,在Megacomcomplex大分子级分的质谱分析中证实了相似的途径富集.这些发现暗示Megacomplex参与ERα驱动的卵巢癌染色质调节。这种组合的治疗策略表现出对细胞增殖和活力的优异抑制。因此,通过在Megacomplex中发现ERα的抗性，ICI+JQ1联合治疗阐明了一种新的药物治疗脆弱性.
