Molecular mechanisms of chemoresistance in breast cancer

描述(申请人提供)：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育中起着至关重要的作用。在对各种遗传毒性或细胞毒性应激源的反应中，细胞凋亡通过从线粒体释放细胞色素c来发生，这种细胞色素c促进了被称为凋亡体的结构的成核，从而激活了被称为半胱氨酸蛋白酶的半胱氨酸蛋白酶。与未转化的细胞相比，癌细胞通过阻止线粒体细胞色素c的释放和/或通过抑制细胞色素c释放后的凋亡体的形成而表现出显著的抗凋亡能力。 受体酪氨酸激酶ErbB2的过度表达被认为与乳腺癌患者的预后不良有关。尽管存在ErbB2特异性的分子抑制剂，但大多数ErbB2高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(即获得性耐药)，这表明目前尚不清楚的化疗耐药机制的存在。在这项提案中，我研究了对FDA批准的ErbB2抑制剂拉帕替尼产生获得性耐药的乳腺癌细胞凋亡抑制的分子机制。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着关键作用。这一假说是基于我们实验室和其他实验室的证据提出的，这些证据表明，凋亡体抑制可以通过两个关键的独立途径发生：HSP902低磷酸化和CAS下调。这些机制对细胞凋亡体的抑制可能会覆盖由各种促凋亡刺激引起的细胞死亡信号。为此，我将追求以下两个具体目标：1)确定HSP902低磷酸化在获得性拉帕替尼耐药中的作用；2)阐明CAS下调的分子机制。我的主要职业目标是在美国一所主要大学获得终身教职。我的长期目标是建立一个独立的研究生涯，成为一名分子癌症生物学家。为了实现这些目标，我希望在整个拟议的研究期间，既发展我的智力知识基础，又增加我的技术技能。这种培训应该很容易在杜克大学医学中心的KornBluth实验室获得；我们的实验室在复杂信号通路的生化分析和分子解剖方面拥有丰富的专业知识，包括在细胞凋亡和细胞周期进展过程中。此外，我还获得了两位杰出的合作者Neil Spector博士和Mark DeWhirst博士，他们分别是乳腺癌和小鼠异种移植癌症模型领域的知名研究人员。此外，为了在职业发展奖期间促进和确保我的进步，我组织了一个咨询委员会，由具有与我的研究和职业目标相关的不同领域专业知识的知名资深科学家组成。总之，拟议的研究将为化疗耐药的机制和潜在的使细胞对促凋亡剂增敏的策略提供重要的见解。此外，这项工作将为我提供作为一名独立调查员的手段。 公共卫生相关性：癌细胞对细胞凋亡性死亡具有明显的抵抗力。在这项建议中，我们将重点放在乳腺癌细胞死亡失调的机制上，特别是在对化疗药物产生获得性耐药性的情况下。这项研究应该有助于阐明乳腺化疗耐药的机制，并有助于开发新的治疗策略来避免这种耐药。

DESCRIPTION (provided by applicant): 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: HSP902 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 HSP902 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. PUBLIC HEALTH RELEVANCE: Cancer cells are markedly resistant to apoptotic cell death. In this proposal, we focus on mechanisms of cell death dysregulation in breast cancers, particularly under conditions of acquired resistance to chemotherapeutic agents. This study should help to elucidate mechanisms of breast chemoresistance and aid in the development of novel therapeutic strategies to avoid such resistance.