Molecular mechanisms of chemoresistance in breast cancer

摘要： 细胞凋亡是细胞程序性死亡的一种形式，在正常组织的动态平衡和 发展。在对各种遗传毒性或细胞毒性应激源的反应中，细胞凋亡是通过释放 来自线粒体的细胞色素c，它促进一种被称为凋亡体的结构的成核 激活半胱氨酸蛋白酶，称为半胱氨酸酶。与未转化的细胞相比，癌细胞表现出明显的 通过阻断线粒体细胞色素c的释放和/或通过抑制凋亡体来抵抗细胞凋亡 细胞色素c释放后形成。 已知受体酪氨酸激酶ErbB2的过度表达与预后不良有关 乳腺癌患者。尽管有ErbB2特异的分子抑制剂可用，但大多数ErbB2- 过度表达的乳腺癌最终会在一段时间内对这些抑制剂产生耐药性(即， 获得性耐药)，表明存在目前尚不清楚的化疗耐药机制。 在这项提案中，我研究了乳腺癌细胞凋亡抑制的分子机制。 对FDA批准的ErbB2抑制剂拉帕替尼具有获得性耐药性。我的中心假设是 抑制凋亡体的形成在乳腺癌获得性拉帕替尼耐药中起关键作用。这 假设是基于我们实验室以及其他证明 端粒体抑制可通过两条关键的独立途径发生：HSP90低磷酸化和CAS 降低监管。这些机制对细胞凋亡体的抑制可能会覆盖由 各种促细胞凋亡的刺激。为此，我将追求以下两个具体目标：1)确定 HSP90低磷酸化在获得性拉帕替尼耐药中的作用；2)阐明分子机制 中科院下调机制。我的主要职业目标是获得一个终身教职的职位。 背景设在美国一所主要大学。我的长期目标是建立一个独立的分子研究事业 癌症生物学家。为了实现这些目标，我希望既发展我的智力知识基础，也 在整个拟议的研究过程中，增加我的技术技能。这一培训应该是 在杜克大学医学中心的科恩布鲁斯实验室很容易获得；我们的实验室有相当多的 在生化分析和复杂信号通路的分子解剖方面的专业知识，在 细胞凋亡与细胞周期进程。此外，我还获得了两位杰出的合作者尼尔博士 Spector和Mark DeWhirst，他们是乳腺癌和老鼠领域的知名研究人员 分别建立肿瘤异种移植模型。此外，为了促进和确保我在 在职业发展奖期间，我组织了一个由久负盛名的咨询委员会组成的 在与我的研究和职业目标相关的不同领域拥有专业知识的资深科学家。总体而言， 拟议的研究将对化疗耐药的机制和潜在的策略提供重要的见解 使细胞对促凋亡剂敏感。此外，这项工作将为我提供建立 我作为一名独立调查员。

Abstract: Apoptosis is a form of programmed cell death that plays a crucial role in normal tissue homeostasis and development. In response to various genotoxic or cytotoxic stressors, apoptosis occurs by release of cytochrome c from the mitochondria, which promotes nucleation of a structure known as the apoptosome to activate cysteine proteases called caspases. In contrast to untransformed cells, cancer cells exhibit marked resistance to apoptosis by blocking mitochondrial cytochrome c release and/or by inhibiting apoptosome formation following cytochrome c release. Overexpression of the receptor tyrosine kinase ErbB2 is known to be associated with poor prognosis in breast cancer patients. Despite the availability of ErbB2-specific molecular inhibitors, the majority of ErbB2- overexpressing breast cancers eventually develop resistance to these inhibitors over a period of time (i.e., acquired resistance), suggesting the presence of currently uncharacterized mechanisms of chemoresistance. In this proposal, I investigate the molecular mechanisms contributing to apoptotic inhibition in breast cancers with acquired resistance to lapatinib, an FDA-approved ErbB2 inhibitor. It is my central hypothesis that inhibition of apoptosome formation plays a critical role in acquired lapatinib resistance in breast cancer. This hypothesis was formulated based on evidence from our laboratory as well as from others demonstrating that apoptosome inhibition can occur by two key independent pathways: HSP90¿ hypophosphorylation and CAS downregulation. Apoptosome inhibition by these mechanisms may override cell death signals caused by various pro-apoptotic stimuli. Towards this end, I will pursue the following two specific aims: 1) to determine the role of HSP90¿ hypophosphorylation in acquired lapatinib resistance, and 2) to elucidate the molecular mechanism of CAS downregulation. My primary career goal is to obtain a tenure-track position in an academic setting at a major U.S. university. My long-term goal is to build an independent research career as a molecular cancer biologist. To achieve these goals, I hope to both develop my intellectual knowledge base as well as increase my technical skill repertoire throughout the duration of the proposed study. This training should be readily attainable in the Kornbluth laboratory at Duke University Medical Center; our lab has considerable expertise in the biochemical analysis and molecular dissection of complex signaling pathways, both during apoptosis and cell cycle progression. In addition, I have obtained two outstanding collaborators, Drs. Neil Spector and Mark Dewhirst, who are established investigators in the fields of breast cancer and mouse xenograft models of cancer, respectively. Moreover, in order to promote and assure my progress during the Career Development Award period, I have organized an advisory committee consisting of well-established senior scientists with expertise in different areas relevant to my research and career goals. Collectively, the proposed studies will provide significant insight into mechanisms of chemoresistance and potential strategies for sensitizing cells to pro-apoptotic agents. Moreover, this work will provide me with the means to establish myself as an independent investigator.