Role of NF-kB signaling in supporting ovarian cancer tumor-initiating cells responsible for cancer recurrence

 描述(由申请人提供)：本提案的总体目标是确定核因子-kB通路在卵巢癌肿瘤启动细胞(TICS)中的作用，并确定化疗后肿瘤成功再生长所需的机制。卵巢癌是美国最致命的妇科恶性肿瘤，尽管大多数患者最初对以铂为基础的化疗有反应，但超过70%的晚期肿瘤复发导致高发病率和死亡率。因此，迫切需要阐明支持治疗抵抗和肿瘤再生长的过程，本研究的目的是确定可以靶向预防肿瘤复发的机制。核因子-kB活性与卵巢癌侵袭性增强、转移潜能增强和预后不良有关。核因子-kB是一种普遍存在的信号通路，其靶基因编码的蛋白质调节免疫反应、细胞存活、增殖、黏附以及与微环境的相互作用。这项建议调查了一种假设，即核因子-kB通路支持卵巢癌细胞亚群，这些亚群可以抵抗化疗并推动肿瘤复发。拟议的项目将解决与卵巢癌复发的生物学相关的几个问题。首先，核因子-kB的哪些功能支持占肿瘤细胞总数一小部分的TICS，以及构成肿瘤大部分的非TICS？非抽搐对于肿瘤的有效再生长是必要的吗？核因子-kB的激活是否与MAPK/ERK整合以支持非抽搐？维持干细胞表型的转录调控因子是否控制着核因子-kB的表达和活性？这项建议旨在了解卵巢癌复发所必需的抽动和非抽动的多个通路的协调。为了开始回答这些问题，我首先开发了一种培养系统来丰富TIC，并将这些细胞中的NF-kB和MAPK/ERK活性与在非TIC丰富条件下生长的细胞(贴壁单层)进行了比较。我的初步研究表明，经典的核因子-kB和MAPK/ERK活性支持非痉挛，而替代的核因子-kB活性支持痉挛。我将从基因上阻断已建立的细胞系和原发患者细胞系中导致经典和替代核因子-kB活性的成分，并使用异种移植小鼠模型测量体内的化疗耐药性和肿瘤再繁殖。这些研究将在分类的TIC和非TIC人群中扩展。分选将使用一种对干细胞转录因子的活性做出反应的新型报告来完成，而不是使用依赖于上下文的传统细胞表面标记。对痉挛或非痉挛中特异性激活的通路的研究将阐明这些不同类型的细胞在肿瘤成功再繁殖中的潜在合作。最后，我将研究导致抽搐中替代核因子-kB激活的新机制，以提供对预防继发性疾病的新靶点的见解。在K99阶段，我将接受广泛的训练，使用小鼠模型来测量化疗耐药和肿瘤复发，我将开发一种可靠的报告，以识别和分离卵巢癌TIC。在K99阶段结束时，我将拥有必要的工具，成为一名拥有异种移植小鼠模型和卵巢TIC生物学专业知识的独立研究员。在这个奖项的R00阶段，这项新的培训将应用于我的整体研究设计，旨在解决我们目前对卵巢癌复发的理解中的重大差距。在K99阶段获得的专业知识，加上我的分子生物学和基因组学背景，将使我能够开发一个综合研究计划，旨在调查卵巢癌持续和复发的新机制，最终导致新的治疗方法，以延长患有这种疾病的女性的生命。这一建议的结果将通过提供对卵巢癌复发和特定细胞亚型的作用的机制洞察来推动卵巢癌领域的发展。拟议目标的完成将指导卵巢癌患者替代治疗策略的发展，并可能突出与其他癌症类型相关的机制。

 DESCRIPTION (provided by applicant): The overall objective of this proposal is to define the role of the NF-kB pathway in ovarian cancer tumor-initiating cells (TICs) and to ascertain mechanisms required for successful repopulation of tumors following chemotherapy. Ovarian cancer is the most lethal gynecological malignancy in the United States and although most patients initially respond to platinum-based chemotherapy, over 70% of advanced stage tumors relapse leading to high morbidity and mortality. Thus, there is an urgent need to clarify processes that support therapy resistance and tumor regrowth, and the purpose of this research is to identify mechanisms that can be targeted to prevent tumor recurrence. NF-kB activity is associated with increased aggressiveness, enhanced metastatic potential and poor outcome in ovarian cancer. NF-kB is a ubiquitous signaling pathway whose target genes encode proteins that regulate immune response, cell survival, proliferation, adhesion, and interaction with the microenvironment. This proposal investigates the hypothesis that NF-kB pathways support subpopulations of ovarian cancer cells that can resist chemotherapy and drive tumor recurrence. The proposed project will address several questions relevant to the biology of ovarian cancer relapse. First, which functions of NF-kB support TICs that comprise a small percentage of the total cells in the tumor, and non-TICs that make up the bulk of the tumor? Are non- TICs necessary for efficient repopulation of the tumor? Does NF-kB activation integrate with MAPK/ERK to support non-TICs? Do transcriptional regulators that maintain stem cell phenotypes control NF-kB expression and activity? This proposal seeks to understand the coordination of multiple pathways in TICs and non-TICs necessary for ovarian cancer relapse. To begin answering these questions I first developed a culture system to enrich for TICs and compared NF-kB and MAPK/ERK activity in these cells with cells grown in non-TIC enriching conditions (adherent monolayer). My preliminary studies suggest classical NF-kB and MAPK/ERK activity support non-TICs whereas alternative NF-kB activity supports TICs. I will genetically interrupt the components leading to classical and alternative NF-kB activity in established cell lines and primary patient cell lines, and measure chemoresistance and tumor repopulation in vivo using xenograft mouse models. These studies will be extended in sorted TIC and non-TIC populations. Sorting will be accomplished using a novel reporter that responds to activity of stem cell transcription factors rather than traditional cell surface markers that ar context dependent. Investigation of pathways specifically activated in TICs or non-TICs will elucidate the potential cooperation of these different cell types in successful repopulation of tumors. Lastly, I will investigate novel mechanisms leading to alternative NF-kB activation in TICs to provide insights into new targets for prevention of secondary disease. During the K99 phase I will receive extensive training using mouse models to measure chemoresistance and tumor relapse and I will develop a reliable reporter for identifying and isolating ovarian cancer TICs. At the end of the K99 phase I will possess the tools necessary to be an independent investigator with expertise in xenograft mouse models and ovarian TIC biology. This new training will be applied to my overall study design during the R00 phase of this award, which aims to address significant gaps in our current understanding of ovarian cancer relapse. The expertise gained during the K99 phase, together with my molecular biology and genomics background, will allow me to develop an integrated research program designed to investigate novel mechanisms of ovarian cancer persistence and relapse, leading ultimately to new therapies to prolong the lives of women with this disease. Results of this proposal will advance the ovarian cancer field by providing mechanistic insight into ovarian cancer recurrence and the role of specific cell subtypes. Completion of the proposed aims will guide the development of alternative therapeutic strategies for patients with ovarian cancer and may highlight mechanisms relevant to other cancer types.