Cell Heterogeneity and Emergent Trastuzumab Resistance in Breast Cancer: Concept

描述(由申请人提供)：理解曲妥珠单抗的紧急耐药受阻于不能量化异种乳腺癌细胞群体中的异常细胞信号通路。因此，迫切需要多学科的方法来评估和解释乳腺癌肿瘤中细胞异质性的临床重要性。我们的长期目标是通过为一种预后技术建立科学基础来改善癌症的临床管理，该技术将识别对分子靶向治疗产生抵抗力的个体。因此，这项拟议的研究与NIH的使命相关，因为它发展了基础知识，可能有助于减轻人类癌症的负担。这项应用的总体目标是确定与药物敏感性相关的信号蛋白的独特模式，并应用反应途径分析的计算工具来解释这些蛋白表达模式的意义。我们的中心假设是，过度表达ErbB2的乳腺癌细胞对曲妥珠单抗的反应表现出异质性。此外，这种异质性是由于影响ErbB2信号通路的蛋白质表达的差异。先前的研究发现，这些蛋白质分别与曲妥珠单抗耐药相关。目前的挑战是推断这些蛋白质如何协同作用影响曲妥珠单抗的耐药性。我们计划测试我们的中心假设，并通过追求以下具体目标来实现这一应用的总体目标：1)建立对曲妥珠单抗反应的细胞异质性在已建立的细胞系中；以及2)建立如何使用反应通路分析来解释细胞信号网络中蛋白质表达的差异模式。在第一个目标下，我们希望测试我们的工作假设，即肿瘤群体中的细胞，如现有的乳腺癌细胞株，表现出各种蛋白质表达的变化，使其对曲妥珠单抗治疗具有不同的敏感性。为了解决这一假设，我们将从现有的ErbB2过表达的乳腺癌细胞系中选择对曲妥珠单抗具有不同敏感性的细胞群。这些细胞群体将用2D-Gel电泳法对蛋白质表达的变化进行实验测试。在第二个目标下，我们的工作假设是，可以使用计算机辅助组装反应机制的算法来构建ErbB2信号网络中早期信号事件的无偏模型。这项拟议研究的基本原理是，识别与曲妥珠单抗敏感性相关的信号蛋白模式，将使在肿瘤活检样本的单细胞水平上测量这些蛋白模式成为可能。这项拟议的研究具有创新性，因为它提供了一种新的方法，将计算系统生物学和蛋白质组学的尖端技术结合起来，以解决乳腺癌患者对曲妥珠单抗紧急耐药的紧迫问题。公共卫生相关性：拟议的研究是了解ErbB2信号网络中的细胞异质性对乳腺癌患者曲妥珠单抗紧急耐药的影响的重要一步。这项拟议的研究预计将对公共健康产生重要的积极影响，因为拟议的方法将使人们能够超越目前限制根除乳腺癌这一危及生命的疾病的进展的遗传范式。这些研究还将为反应途径分析、癌细胞生物学和生物分析化学之间的研究生和本科生提供培训机会。这个跨学科的项目将使学生接触到生物学研究的各个方面，并接触到解决生物医学问题的尖端技术。在西弗吉尼亚大学的这种独特的培训经历将帮助学生在未来的研究职业生涯中取得成功。

DESCRIPTION (provided by applicant): Understanding emergent resistance to trastuzumab is inhibited by the inability to quantify aberrant cell signaling pathways among heterogeneous populations of breast cancer cells. Thus there is urgent need for multidisciplinary approaches to assess and interpret the clinical importance of cellular heterogeneity within breast cancer tumors. Our long-term goal is to improve the clinical management of cancer by establishing the scientific foundation for a prognostic technology that will identify individuals who will develop resistance to molecularly targeted therapies. Thus, the proposed research is relevant to NIH's mission by developing fundamental knowledge that will potentially help to reduce the burdens of human cancer. The overall objective of this application is to identify unique patterns of signaling proteins associated with drug sensitivity and apply the computational tools of reaction pathway analysis to interpret the significance of these patterns of protein expression. Our central hypothesis is that breast cancer cells that overexpress ErbB2 exhibit heterogeneity in response to trastuzumab. Furthermore, this heterogeneity is due to variations in expression of proteins that influence the ErbB2 signaling pathway. Prior studies identify such proteins that individually correlate with trastuzumab resistance. The challenge is inferring how these proteins act in concert to influence trastuzumab resistance. We plan to test our central hypothesis and accomplish the overall objective of this application by pursuing the following specific aims: 1) Establish that cellular heterogeneity in response to trastuzumab exists within established cell lines; and 2) Establish how reaction pathway analysis can be used to interpret differential patterns of protein expression within cellular signaling networks. Under the first aim, we expect to test our working hypothesis that cells within a tumor population, as represented by existing breast cancer cell lines, exhibit a variety of changes in protein expression that confer differential sensitivity to trastuzumab treatment. To address this hypothesis, we will select cell populations with varying sensitivity to trastuzumab from existing ErbB2-overexpressing breast cancer cell lines. These cell populations will be experimentally tested for variations in protein expression using 2D-gel electrophoresis. Under the second aim, our working hypothesis is that an unbiased model of the early signaling events in the ErbB2 signaling network can be constructed using an algorithm for the computer-assisted assembly of reaction mechanisms. The rationale that underlies the proposed research is that identifying patterns of signaling proteins that are correlated with sensitivity to trastuzumab will enable measuring these protein patterns at the single-cell level in tumor biopsy samples. The proposed research is innovative as it provides a novel approach that combines cutting-edge techniques in computational systems biology and proteomics to address the pressing issue of emergent resistance to trastuzumab in breast cancer patients. PUBLIC HEALTH RELEVANCE: The proposed studies are an important step towards understanding the implications of cellular heterogeneity within the ErbB2 signaling network on emergent resistance to trastuzumab in breast cancer patients. The proposed research is expected to have an important positive impact on public health, because the approach proposed will enable transcending the current genetic paradigm that limits progress toward eradicating breast cancer as a life-threatening disease. These studies will also provide training opportuni- ties for graduate and undergraduate students at the interface between reaction pathway analysis, cancer cell biology, and bioanalytical chemistry. This interdisciplinary project will expose the students to various facets of biological research and to cutting-edge techniques for solving biomedical problems. This unique training experience at WVU will help students to succeed in their future research careers.