The Role of SRC-1 in Breast Cancer

描述（由申请人提供）：转移（Meta）是导致几乎所有乳腺癌（BC）死亡的原因；然而，其病理生理机制尚不清楚。本项目的长期目标是了解BC Meta的分子机制，并开发新的思路/策略来控制BC Meta。类固醇受体共激活因子1 （SRC-1）是一个新发现的Meta基因，它可以促进Twist的表达，从而增强BC细胞上皮-间质转化（EMT）和肺Meta。Twist是侵袭性BC中过表达的dna结合转录因子，是EMT和BC Meta的主要调节因子。Twist在BC中的表达也与化疗耐药的获得有关。此外，EMT和间充质上皮转化（mesenchymal epithelial transition， MET）之间的BC细胞可塑性被认为是可转换的，并在Meta中发挥关键作用，但这种顺序事件及其与Meta的直接联系的直接体内证据仍然缺乏。在目前的项目中，研究人员假设BC细胞EMT-MET可塑性存在于局部和转移性肿瘤中。Twist具有促进EMT和抑制MET的作用。Twist的失活将提高BC化疗的疗效，而不会对成人造成严重的健康问题。Twist蛋白的稳定性受磷酸化控制，抑制负责这种磷酸化的激酶并增加相应的磷酸酶活性会降低BC细胞中Twist蛋白的水平及其功能。本文提出了三个具体目标（SAs）来检验这些假设。第一个SA是明确Twist在MG肿瘤细胞可塑性中的作用，以及MG肿瘤细胞可塑性在BC Meta中的作用。EMT依赖性启动子报告传感器系统将用于在BC细胞EMT- met可塑性和肺Meta过程中追踪EMT细胞谱系。我们将利用慢病毒诱导小鼠乳腺肿瘤内源性表达、过表达或敲除Twist (KO)，以确定Twist在肿瘤细胞可塑性和Meta中的作用。第二个SA是评估Twist失活对一般健康的副作用，并测试靶向Twist是否能提高BC化疗疗效。Twist将通过诱导KO系统在成年小鼠中被敲除。研究人员将研究Twist KO是否会在成年小鼠中引起任何严重的健康问题，以及Twist KO是否会增强紫杉醇化疗的疗效。这将为Twist是否可以作为一种副作用可容忍的癌症治疗靶点提供信息。通过诱导型KO模型，确定Twist失活对BC化疗的影响。第三个SA是明确Twist磷酸化在调节其蛋白稳定性中的作用和分子机制，并测试通过控制Twist磷酸化程度来破坏Twist的不稳定性是否会抑制BC细胞侵袭和Meta。这些研究的成功结果不仅将为了解Twist如何调节BC细胞可塑性和癌症Meta提供见解，而且还将验证Twist作为控制BC Meta的靶点，并提供直接的体内证据来证明或反驳BC Meta过程中EMT和MET之间的顺序联系。

DESCRIPTION (provided by applicant): Metastasis (Meta) is responsible for almost all breast cancer (BC) mortality; however, its pathophysiological mechanism is poorly understood. The long-term goal of this project is to understand the molecular mechanisms of BC Meta and develop new ideas/strategies to control BC Meta. The steroid receptor coactivator 1 (SRC-1) is a newly identified Meta gene and it promotes Twist expression to enhance BC cell epithelial-mesenchymal transition (EMT) and lung Meta. Twist is a DNA-binding transcription factor overexpressed in invasive BCs and a master regulator of EMT and BC Meta. Twist expression in BC is also associated with acquisition of chemotherapy resistance. In addition, the BC cell plasticity between EMT and mesenchymal epithelial transition (MET) has been proposed to be convertible and play a crucial role in Meta, but direct in vivo evidences for such sequential events and their direct link to Meta are still absent. In the current project, the investigators hypothesize that BC cell EMT-MET plasticity exists in both local and metastatic tumors. Twist plays a role in promoting EMT and suppressing MET. Inactivation of Twist will enhance the efficacy of BC chemotherapy without causing severe health problems in adults. Twist protein stability is controlled by phosphorylation, and inhibition of kinase responsible for such phosphorylation combined with an increase in corresponding phosphotase activity will reduce Twist protein level and its function in BC cells. Three specific aims (SAs) are proposed to test these hypotheses. The first SA is to define the role of Twist in MG tumor cell plasticity and the role of MG tumor cell plasticity in BC Meta. An EMT- dependent promoter-reporter senor system will be used to trace the EMT cell lineage during the processes of BC cell EMT-MET plasticity and lung Meta. Lentivirus will be used to induce mouse mammary tumors with endogenous expression, overexpression or knockout (KO) of Twist to determine the role of Twist in tumor cell plasticity and Meta. The second SA is to evaluate the side effects of Twist inactivation on general health and test whether targeting Twist enhances BC chemotherapy efficacy. Twist will be knocked out in adult mice using an inducible KO system. Investigators will examine whether Twist KO in adult mice causes any severe health problems and whether Twist KO will enhance the efficacy of Paclitaxel chemotherapy. This will provide information about whether Twist can be used as a cancer therapy target with tolerable side effects. With the inducible KO model, the effect of Twist inactivation on BC chemotherapy will be determined. The third SA is to define the role and molecular mechanisms of Twist phosphorylation in regulation of its protein stability and test whether destabilization of Twist by controlling its phosphorylation degree will inhibit BC cel invasion and Meta. Successful results from these studies will not only provide insights into understanding how Twist regulates BC cell plasticity and cancer Meta, but also validate Twist as a target for controlling BC Meta and provide direct in vivo evidence to prove or disprove the sequential connection between EMT and MET during BC Meta.