AN ACTIVE ROLE OF ADAPTOR PROTEINS IN TYROSINE KINASE INHIBITOR RESISTANCE

衔接蛋白在酪氨酸激酶抑制剂抗性中的积极作用

• 批准号: 8756983
• 负责人: ERIC B. HAURA
• 金额: $ 21.99万
• 依托单位: H. LEE MOFFITT CANCER CTR & RES INST
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8756983
• 关键词: Adaptor Signaling Protein; Address; Affinity Chromatography; Binding; Biological Assay; Cancer cell line; Cells; Complex; Dimerization; Drug resistance; Epidermal Growth Factor Receptor; Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor; Erlotinib; Future; Goals; Growth Factor; Hand; In Situ; Laboratories; Ligands; Ligation; Malignant neoplasm of lung; Mass Spectrum Analysis; Measurement; Measures; Mediating; Medicine; Modeling; Mutate; Oncogenes; Outcome; Patients; Peptides; Pharmaceutical Preparations; Phosphorylation; Phosphotyrosine; Play; Process; RNA Interference; Receptor Protein-Tyrosine Kinases; Reporting; Resistance; Role; Shapes; Signal Transduction; Testing; Therapeutic Uses; Tyrosine; Tyrosine Kinase Inhibitor; Tyrosine Phosphorylation; advanced disease; base; cancer cell; cell type; improved; liquid chromatography mass spectrometry; mutant; neoplastic cell; novel; novel therapeutic intervention; protein complex; public health relevance; receptor; response; success; tumor; tumor microenvironment

DESCRIPTION (provided by applicant): The goal of this project is to define an active role of adaptor proteins in mediating resistance to receptor tyrosine kinase (RTK) inhibitors. In the field of lung cancer therapeutics, use of tyrosine kinase inhibitors (TKI) directed against mutant forms of EGFR or EML4-ALK rearranged lung cancers has made major improvements in patient outcome with advanced disease. However, intrinsic or acquired drug resistance remains a persistent obstacle towards a cure. Growth factor ligands secreted in the tumor microenvironment can negate the effects of TKI across multiple tumor cell types driven by mutant oncogenes. We propose that adaptor proteins involved in RTK signaling are playing an active role in this process. We hypothesize that adaptors, displaced from driver RTK by TKI, shape co-expressed RTK signaling and influence the effects of microenvironmental ligands that drive resistance. Our model, to be tested in this project, consists of adaptor proteins being relocalized from the targeted driver RTK to other co-expressed RTK. This switching of adaptors to the co- expressed RTK enables priming phosphorylation mediated by direct binding of the adaptor and dimerization of two RTK molecules. This allows for full RTK activation in the presence of minute amounts of growth factor ligands. In our laboratory, using various phosphoproteomic approaches, including direct measurements of phosphotyrosine peptide abundance using mass spectrometry, we have consistently identified increased abundance of RTK tyrosine phosphorylation in co-expressed RTK following treatment of cells with TKI directed against the driver oncogene. Our results suggest that priming phosphorylation following TKI treatment of co- expressed RTK can be related to ability of some growth factors to drive TKI resistance. We will test this model using mass spectrometry and proximity ligation assays to quantitatively and qualitatively examine adaptor and RTK complexes in cells following TKI treatment. Specific aim 1 will characterize the fate of adaptor proteins and changes in protein complexes in co-expressed RTK following TKI. Specific aim 2 will characterize how RTK:adaptor pairing relates to growth factor ligand rescue. Specific aim 3 will characterize the role of adaptor proteins in priming tyrosine phosphorylation of co-expressed RTK. Success in this project will identify an active role in adaptor proteins in shaping TKI responses and can in the future yield predictive markers identifying tumors rapidly adapting to TKI through growth factor ligand rescue. The hypothesis is novel and therefore important to be tested. Complex approaches to measuring protein complexes are in hand further increasing the chances of success.

描述（由申请人提供）：该项目的目标是确定衔接蛋白在介导受体酪氨酸激酶（RTK）抑制剂耐药性中的积极作用。在现场 在肺癌治疗中，使用针对 EGFR 突变形式或 EML4-ALK 重排肺癌的酪氨酸激酶抑制剂 (TKI) 已显着改善晚期疾病患者的预后。然而，内在或获得性耐药性仍然是治愈的持续障碍。肿瘤微环境中分泌的生长因子配体可以抵消 TKI 对突变癌基因驱动的多种肿瘤细胞类型的影响。我们认为参与 RTK 信号转导的接头蛋白在此过程中发挥着积极作用。我们假设由 TKI 取代驱动 RTK 的接头形成共表达 RTK 信号并影响驱动耐药性的微环境配体的作用。我们的模型将在该项目中进行测试，由从目标驱动 RTK 重新定位到其他共表达 RTK 的接头蛋白组成。这种将接头转换为共表达 RTK 能够引发由接头直接结合和两个 RTK 分子二聚化介导的磷酸化。这允许在微量生长因子配体存在的情况下完全激活 RTK。在我们的实验室中，使用各种磷酸蛋白质组学方法，包括使用质谱法直接测量磷酸酪氨酸肽丰度，我们一致地发现，在用针对驱动癌基因的 TKI 处理细胞后，共表达 RTK 中 RTK 酪氨酸磷酸化的丰度增加。我们的结果表明，TKI 治疗共表达 RTK 后引发的磷酸化可能与某些生长因子驱动 TKI 耐药的能力有关。我们将使用质谱和邻近连接测定来测试该模型，以定量和定性检查 TKI 治疗后细胞中的接头和 RTK 复合物。具体目标 1 将描述 TKI 后共表达 RTK 中接头蛋白的命运和蛋白复合物的变化。具体目标 2 将描述 RTK：适配器配对如何与生长因子配体救援相关。具体目标 3 将描述衔接蛋白在引发共表达 RTK 酪氨酸磷酸化中的作用。该项目的成功将确定衔接蛋白在塑造 TKI 反应中的积极作用，并在未来产生预测标记，通过生长因子配体拯救来识别肿瘤快速适应 TKI。该假设是新颖的，因此值得检验。测量蛋白质复合物的复杂方法正在进一步增加成功的机会。