Estrogen receptor fusions genes as drivers of endocrine resistance in breast cancer

抗雌激素治疗是防治雌激素受体（ER，ESR 1）阳性乳腺癌的关键 癌症，但治疗耐药性发生在多达一半的情况下，导致无法治愈的晚期疾病。我们 最近报道了在晚期内分泌抵抗性乳腺癌中复发的ESR 1融合，配体丢失 结合代表了典型的耐药性模型。有趣的是，ESR 1融合的病例同时发生 ESR 1碱基对突变，可能指向在长期抗雌激素治疗下的趋同进化。 疗法来自我们小组和其他人的证据表明，在约10-20%的晚期乳腺癌中发现了ESR 1融合。 然而，在乳腺癌中，许多ESR 1融合体具有不同的3'配偶体基因，使得检测具有挑战性。 为了确定患病率，我们将全面重新分析转移性乳腺癌患者数据中的ESR 1基因座。 乳腺癌此外，我们已经开发并优化了能够进行以下操作的靶捕获测序测定： 全面检测肿瘤活检和ctDNA中的ESR 1融合。我们已经确定了内源性的 在两种患者来源的异种移植物（PDX）（ESR 1-LPP和ESR 1-NFkB）中以及在两种患者来源的异种移植物（PDX）（ESR 1-LPP和ESR 1-NFkB）中表达的ESR 1融合物。 ER阳性乳腺癌细胞系MDA-MB-435（ESR 1-SYNE 1）。我们将表达ESR 1融合体， 产生的患者源性类器官（PDO）显示ER表达和对雌激素的反应， 或大于传统细胞系。有趣的是，不同的ESR 1融合体显示出明显的活性差异， 暗示3 ′基因配偶体在调节ESR 1 N末端结构域（NTD）活性中的作用。中立旅行证件 是一种内在无序的蛋白质内的反式激活结构域，并使用一种新的整合生物物理学 方法，我们最近报道说，虽然它主要是无序的，它可以采取一个意想不到的紧凑， 构象，其反过来指导与辅调节子的相互作用。在这个建议中，我们试图检验假设， ESR 1融合是内分泌抵抗和乳腺癌死亡率的关键介质。我们会委聘 EnRich确定国家和国际临床研究中ESR 1融合（和点突变）的患病率 晚期内分泌抵抗性乳腺癌的队列，并在治疗期间使用 纵向收集血浆和ctDNA分析。我们将使用患者来源的乳腺类器官模型， 癌症，以比较和对比ESR 1融合和点突变相对于野生型ESR 1融合和点突变的活性和功能。 ER型，并将其转化为体内患者来源的类器官异种移植物（PDOX）和（PDX）模型，以评估 内分泌抵抗我们将研究作用机制，包括对DNA结合的影响， 转录活性，以及3'配偶体基因在任何功能活性获得中的作用。最后我们将 研究ESR 1融合蛋白的结构-功能，重点是NTD，并确定NTD中改变的结构。 在评估特定NTD氨基酸的功能之前融合配偶体的背景以及它们如何 改变结构和随后的靶基因表达。我们相信我们的多学科研究将提供 机制的洞察力，以帮助开发新的治疗乳腺癌与ESR 1融合。

Anti-estrogen therapy is key to the prevention and treatment of estrogen receptor (ER, ESR1) positive breast cancer, but therapy resistance occurs in up to a half of cases resulting in incurable advanced disease. We recently reported recurrent ESR1 fusions in advanced endocrine-resistant breast cancers, with loss of ligand binding representing a canonical model of drug resistance. Intriguingly, cases with ESR1 fusions co-occurred with ESR1 base-pair mutations, likely pointing towards convergent evolution under prolonged anti-estrogen therapy. Evidence from our group and others suggest that ESR1 fusions are found in ~10-20% of advanced breast cancers, however, many ESR1 fusions harbor different 3’ partner genes, making detection challenging. To determine prevalence, we will comprehensively re-analyze the ESR1 genomic locus in data from metastatic breast cancers. Further, we have developed and optimized a target capture sequencing assay capable of comprehensively detecting ESR1 fusions in both tumor biopsies and ctDNA. We have identified endogenous ESR1 fusions that are expressed in two patient derived xenografts (PDX) (ESR1-LPP and ESR1-NFkB) and in an ER-positive breast cancer cell line MDA-MB-435 (ESR1-SYNE1). We will express ESR1 fusions in recently generated patient-derived organoids (PDO) which show ER expression and response to estrogen that is equal or greater than traditional cell lines. Intriguingly, different ESR1 fusions show stark differences in activity, implicating a role for the 3’ gene partner in modulating activity of the ESR1 N terminal domain (NTD). The NTD is an intrinsically disordered protein within the transactivation domain, and using a novel integrative-biophysics approach, we recently reported that while it is mostly disordered, it can adopt an unexpectedly compact conformation which in turn directs interaction with coregulators. In this proposal we seek to test the hypothesis that ESR1 fusions are key mediators of endocrine-resistance and breast cancer mortality. We will employ EnRich to determine the prevalence of ESR1 fusions (and point mutations) in national and international clinical cohorts of advanced endocrine resistant breast cancer and monitor when they arise during therapy using longitudinal collection of plasma and ctDNA analyses. We will use patient-derived organoid models of breast cancer to compare and contrast the activity and function of ESR1 fusions and point mutations relative to wild- type ER and translate this into in vivo patient derived organoid xenograft (PDOX) and (PDX) models to assess endocrine resistance. We will study mechanism of action, including the effects on DNA binding and transcriptional activity, and the role of the 3’ partner gene in any gain of function activities. Finally, we will examine the structure-function of ESR1 fusions focusing on the NTD and determining the altered structure in the context of the fusion partner before assessing the function of specific NTD amino acids and how they change structure and subsequent target gene expression. We believe our multi-disciplinary studies will provide mechanistic insight to aid in the development of new therapies against breast cancers with ESR1 fusions.