Novel Role of ThPOK in Mammary Carcinoma

项目摘要/摘要 综述：目前迫切需要了解HER2乳腺癌的分子基础，因为 大多数患者最终变得对包括抗HER2治疗在内的治疗无效。在此，我们 提供初步数据，将人类HER2乳腺癌的不良临床结果与 细胞质中高水平的转录因子ThPOK(细胞ThPOK)。此外，使用一种新的小鼠模型 细胞质限制性ThPOK(ThPOKΔ非小鼠)，我们建立了细胞ThPOK之间的因果关系 和高渗透性Her2乳腺癌的发展。小鼠乳腺癌细胞的蛋白质组学分析 提示细胞ThPOK与多种参与Her2信号转导的胞浆蛋白相互作用，包括 几个SH3蛋白可能与ThPOK中一个保守的富含脯氨酸的基序结合。鉴于的核心作用 HER2信号在HER2乳腺癌生物学中的作用，我们假设细胞ThPOK与这些因子相互作用 以某种方式增强HER2介导的信号转导，这是乳腺癌的重要驱动因素 发展/进步，这一点到目前为止一直被忽视。 研究热点：POK转录因子与多种人类癌症有关 被认为反映了对转录的直接影响。相反，我们现在认为ThPOK的胞浆定位 在人和小鼠的乳腺癌中，展示了一种新的POK介导的肿瘤发生模式 关于核功能。这些发现为拟议的研究提供了一个创新和令人信服的前提。 具体目标：我们将根据3个目标阐明细胞ThPOK在乳腺癌中的作用：SA-1： ThokNLS介导的Δ致癌的分子基础--检测细胞ThPOK对肿瘤细胞的影响 HER2/EGFR在细胞中的表达和信号转导，并测试ThPOK SH3-相互作用的要求 细胞ThPOK介导的小鼠乳腺癌的结构域。SA-2：解剖预后价值及分子生物学意义 人外周血中ThPOK胞浆定位的基础--评价两者的相关性 细胞ThPOK与人不同HER2乳腺癌亚型生存率的关系及其分子基础的确定 ThPOK在人乳腺癌细胞中的胞质定位。SA-3：阐明细胞ThPOK的作用 关于BC肿瘤的维持-阐明细胞ThPOK是否是肿瘤维持/进展所必需的 并鉴定了一种新的人源化Δ小鼠模型，该模型 表达在一些人类乳腺癌患者中发现的hThPOKp.H22pTfs*6等位基因。 影响：鉴于在细胞质ThPOK强制定位的小鼠中观察到癌症的高发病率 以及HER2乳腺癌患者细胞质ThPOK的高频率定位 细胞ThPOK促进乳腺癌的分子基础可能对人类产生很大影响 健康。在这里，我们结合新的动物模型和分子方法来阐明这些机制。 这些研究有能力改变推定POK因素的基本概念框架 促进肿瘤发生。阐明这些机制可能会导致新的治疗方法。 针对HER2乳腺癌和其他可能与细胞ThPOK有关的人类恶性肿瘤。

PROJECT SUMMARY/ABSTRACT Overview: There is an urgent need to understand the molecular basis of HER2+ breast cancer, given that the majority of patients eventually become refractory to treatment including with anti-HER2 therapy. Herein, we provide preliminary data linking poor clinical outcome of human HER2+ breast cancers with the presence of high cytoplasmic levels of the transcription factor ThPOK (cytoThPOK). Further, using a novel mouse model of cytoplasmically restricted ThPOK (ThPOKΔNLS mice), we establish a causal relationship between cytoThPOK and development of highly penetrant Her2+ breast cancer. Proteomic analysis of mouse breast cancer cells indicates that cytoThPOK interacts with multiple cytosolic proteins implicated in Her2 signaling, including several SH3 proteins that probably bind to a conserved proline-rich motif in ThPOK. Given the central role of HER2 signaling in HER2+ breast cancer biology, we hypothesize that cytoThPOK interaction with these factors enhances HER2-mediated signaling in some way, and that this represents an important driver of breast cancer development/progression, that has so far been overlooked. Research Focus: POK transcription factors have been implicated in diverse human cancers, which was presumed to reflect direct effects on transcription. In contrast, we now implicate cytosolic localization of ThPOK in breast cancer in humans and mice, demonstrating a novel mode of POK-mediated oncogenesis not based on nuclear function. These findings provide an innovative and compelling premise for the proposed studies. Specific Aims: We will elucidate the role of cytoThPOK in breast cancer according to 3 aims: SA-1: Elucidating molecular basis of ThPOKΔNLS–mediated oncogenesis - to test effect of cytoThPOK on HER2/EGFR expression and signaling in cell lines, and test the requirement for the ThPOK SH3-interaction domain for cytoThPOK-mediated breast cancer in mice. SA-2: Dissecting prognostic value and molecular basis for cytoplasmic localization of ThPOK in human BC - to evaluate correlation between high cytoThPOK and survival in different human HER2+ breast cancer subtypes, and determine the molecular basis for cytoplasmic localization of ThPOK in human breast cancer cells. SA-3: Elucidating effect of cytoThPOK on BC tumor maintenance - to elucidate whether cytoThPOK is required for tumor maintenance/progression of established cancers from ThPOKΔNLS mice, and characterize a new humanized ThPOK mouse model that expresses a variant hThPOKp.H22pTfs*6 allele found in some human breast cancer patients. Impact: Given the high incidence of cancer observed in mice with enforced cytoplasmic ThPOK localization and the high frequency of cytoplasmic ThPOK localization in human HER2+ breast cancer patients, elucidating the molecular basis by which cytoThPOK promotes breast cancer is likely to have high impact on human health. Here we combine novel animal models and molecular approaches to elucidate these mechanisms. These studies have the capacity to shift the basic conceptual framework by which POK factors are presumed to promote oncogenesis. Potentially, elucidating these mechanisms may lead to novel therapeutic approaches to target HER2+ breast cancer and other human malignancies in which cytoThPOK may be implicated.