Investigating the role of Eya3 in the regulation of innate immune signaling cascades in Triple Negative Breast Cancer

项目摘要/摘要： 三阴性乳腺癌(TNBC)是乳腺癌的一种亚型，以低度或缺失为特征 雌激素受体和HER2生长因子受体的表达。TNBC的转移率很高 总体预后较差，很大程度上是由于缺乏治疗局部和 播散性疾病。 眼睛缺失(EYA)蛋白家族是转录辅助因子，既具有内在的 酪氨酸磷酸酶和相关的丝氨酸/苏氨酸磷酸酶活性，这个蛋白家族有 已被证明在小鼠和人类的正常发育过程中发挥着重要作用。除了他们的 发育作用，Eya蛋白也被牵连到促进几乎所有已知的 癌症。我的导师海德·福特博士的实验室最近发表的研究表明，Eya3 在TNBC肿瘤中通过下调表达抑制CD8 T细胞功能起重要作用 主要通过其苏氨酸磷酸酶活性，初步数据表明，它还可能 在此背景下，在调节各种细胞因子/趋化因子的表达方面发挥作用。先天的激活 免疫信号在肿瘤细胞中的级联，如核因子-kB，以及下游的细胞因子/趋化因子信号，已经 已被证明在多种肿瘤中促进肿瘤进展、免疫逃逸和远处转移 类型，包括乳腺癌。在初步实验中，我们观察到shRNA介导的Eya3 基因敲除(KD)使小鼠TNBC细胞多种细胞因子和趋化因子的表达减少 核因子-kB在体外的激活，表明Eya3在这些信号级联中具有很强的调节作用。另外， 体内实验表明，小鼠TNBC细胞中的Eya3KD极大地抑制了原发肿瘤的生长和 免疫活性小鼠模型中的自发转移，这种变化与显著的 与对照相比，Eya3KD肿瘤内先天免疫群体的改变. 考虑到这些初步发现，我们假设Eya3的表达调节先天的 TNBC细胞中的免疫信号轴改变肿瘤中的趋化因子/细胞因子信号 微环境，促进原发肿瘤的生长和转移。我们的目标是 1.确定Eya3介导的天然免疫信号在TNBC细胞中的作用 对肿瘤启动的免疫抑制、肿瘤生长和转移的影响；2.确定其作用机制 Eya3在TNBC中调节天然免疫信号通路的作用。这些研究提出了一个 不仅有机会研究控制肿瘤细胞串扰的重要生物学因素和 免疫颠覆，但还将确定潜在的预防或治疗新的治疗靶点 播散性的TNBC。

PROJECT SUMMARY/ABSTRACT: Triple Negative Breast Cancer (TNBC) is a subtype of breast cancer characterized by low or absent expression of the estrogen receptor and the HER2 growth factor receptor. TNBC has high rates of metastasis and an overall poor prognosis in large part due to a lack of targeted therapeutics for treating both localized and disseminated disease. The Eyes absent (Eya) family of proteins are transcriptional cofactors that possess both intrinsic tyrosine phosphatase and associated serine/threonine phosphatase activity, and this family of proteins has been shown to play important roles in normal development in mice and humans. In addition to their developmental roles, Eya proteins have also been implicated in promoting nearly all known hallmarks of cancer. Recent published work from the laboratory of my mentor, Dr. Heide Ford, demonstrated that Eya3 plays an important role in suppressing CD8+ T-cell function within TNBC tumors by downregulating expression of PD-L1, primarily through its threonine phosphatase activity, and preliminary data suggests that it may also play a role in regulating the expression of various cytokines/chemokines in this context. Activation of innate immune signaling cascades in tumor cells, such as NF-kB, and downstream cytokine/chemokine signaling, has been shown to promote tumor progression, immune evasion, and distant metastasis in a variety of tumor types, including breast cancer. In preliminary experiments, we observed that shRNA-mediated Eya3 knockdown (KD) in murine TNBC cells reduced expression of multiple cytokines and chemokines and reduced NF-kB activation in vitro, suggesting a strong regulatory role of Eya3 in these signaling cascades. Additionally, in vivo experiments suggest that Eya3 KD in murine TNBC cells greatly reduces primary tumor growth and spontaneous metastasis in immune-competent mouse models, and this change correlates with significant alterations in innate immune populations within the Eya3 KD tumors compared to their control counterparts. With these preliminary findings in mind, we hypothesize that Eya3 expression regulates an innate immune signaling axis in TNBC cells which alters chemokine/cytokine signaling in the tumor microenvironment and facilitates enhanced primary tumor growth and metastasis. Our aims are as follows: 1. To determine the contribution of Eya3-mediated induction of innate immune signaling in TNBC cells to tumor-initiated immune suppression, tumor growth, and metastasis and 2. To identify the mechanism of action by which Eya3 regulates innate immune signaling pathways in TNBC. These studies present an opportunity not only to investigate the important biology governing tumor cell crosstalk and mechanisms of immune subversion, but additionally will identify potential novel therapeutic targets for prevention or treatment of disseminated TNBC.