Kinome Reprogramming in Response to Targeted Kinase Inhibitors

响应靶向激酶抑制剂的激酶组重编程

• 批准号: 8457042
• 负责人: GARY L. JOHNSON
• 金额: $ 26.78万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-15 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8457042
• 关键词: Affect; Affinity; Animals; Apoptosis; Automobile Driving; Behavior; Binding; Biological Assay; Breast Cancer Cell; Bypass; Cancer cell line; Cell Line; Cells; Chemicals; Clinical Trials; Combined Modality Therapy; DDR1 gene; Disease; Dose; Drug resistance; Feedback; Genetically Engineered Mouse; Goals; Human; Knowledge; Lead; MAP2K1 gene; MAPK3 gene; MEK inhibition; MEKs; Mass Spectrum Analysis; Measures; Mediating; Methods; Modeling; Mutation; Pathway interactions; Patients; Phase; Phase I Clinical Trials; Phase I/II Trial; Phosphorylation; Phosphotransferases; Pre-Clinical Model; Protein Kinase; Proteomics; Reagent; Receptor Protein-Tyrosine Kinases; Regulation; Resistance; Resistance development; Role; Running; Sepharose; Signal Transduction; Techniques; Technology; Testing; Therapeutic; Time; Tumor Cell Line; Up-Regulation; base; c-myc Genes; cancer therapy; design; disorder subtype; inhibitor/antagonist; innovation; kinase inhibitor; malignant breast neoplasm; meetings; mouse model; neoplastic cell; novel; novel strategies; pre-clinical; prevent; research clinical testing; response; small molecule; therapeutic target; triple-negative invasive breast carcinoma; tumor; tumor progression

DESCRIPTION (provided by applicant): A novel approach has been developed to study the reprogramming of protein kinases "en masse" allowing ~40-60% of the expressed kinome assayed in a single mass spectroscopy run. Our methods utilize Multiplexed Inhibitor Beads (MIBs), consisting of Sepharose beads with covalently immobilized, linker adapted, kinase inhibitors. The technique allows interrogation of kinases known by sequence but which have been understudied due to lack of biologic or phenotypic knowledge or the availability of reagents. MIB/MS identified a kinome response signature to a select MEK1/2 inhibitor AZD6244 that is currently in clinical testing for triple negative breast cancer. The only defined substrate for MEK1/2 are ERK1 and 2, yet we observed changes in activity of kinases in every subfamily of the kinome in response to MEK inhibition. Kinome assessment showed a time-dependent reprogramming that involved an early loss of ERK feedback regulation of RAF and MEK that allowed upstream reactivation of the MEK-ERK pathway. The time dependent change in MIB binding of specific receptor tyrosine kinases (RTKs) such as PDGFR¿ and DDR1 was readily detected and provided the critical experimental observation that MEK inhibition was driving the expression and activation of multiple RTKs. c-Myc degradation was a key mechanism mediating kinome reprogramming. The fact that multiple RTKs are activated in response to MEK inhibition demonstrates the difficulty in using single kinase inhibitors to arrest tumor progression. The aims of this proposal include: 1. Define kinome activation state in the human basal/claudin-low like SUM159 cell line and the pre-clinical C3Tag GEMM for basal/claudin-low breast cancer. Kinase signatures will be defined in response to MEK1/2 and PI3K inhibitors alone and in combination, which are currently in clinical testing. 2. Define mechanisms of kinome reprogramming in MEK and PI3K inhibitor resistant tumors and cell lines. 3. Develop rational predictions of kinase inhibitor combinations based on kinome signatures of sensitive and resistant tumors for testing in the C3Tag GEMM for basal/claudin-low breast cancer. The goal is to define combination therapies that overcome resistance and cause apoptosis and tumor regression.

描述（由申请人提供）：已经开发出一种新的方法来研究蛋白质激酶的“整体”重编程，允许在单次质谱运行中检测约40-60%的表达激酶。我们的方法利用多路抑制剂珠（MIBs），由磷酸酶珠与共价固定，连接适应，激酶抑制剂组成。该技术允许对已知序列的激酶进行询问，但由于缺乏生物学或表型知识或试剂的可用性而未得到充分研究。MIB/MS鉴定了一种MEK1/2抑制剂AZD6244的kinome应答特征，该抑制剂目前正在进行三阴性乳腺癌的临床试验。MEK1/2唯一确定的底物是ERK1和2，但我们观察到，在MEK抑制作用下，每个激酶亚家族的激酶活性都发生了变化。Kinome评估显示了一种时间依赖性重编程，涉及RAF和MEK的ERK反馈调节的早期丢失，从而允许MEK-ERK途径的上游重新激活。特异性受体酪氨酸激酶（rtk）如PDGFR¿和DDR1的MIB结合的时间依赖性变化很容易被检测到，并提供了关键的实验观察，即MEK抑制驱动多种rtk的表达和激活。c-Myc降解是介导激酶重编程的关键机制。多个rtk在MEK抑制下被激活的事实表明，使用单一激酶抑制剂来阻止肿瘤进展是困难的。这一建议的目的包括：1。确定人基底/低claudin样SUM159细胞系的kinome激活状态和基底/低claudin乳腺癌的临床前C3Tag GEMM。激酶特征将被定义为对MEK1/2和PI3K抑制剂单独或联合的反应，目前正在临床试验中。2. 明确MEK和PI3K抑制剂耐药肿瘤和细胞系中kinome重编程的机制。3. 基于敏感和耐药肿瘤的激酶组特征，开发合理的激酶抑制剂组合预测，用于基础/低cludin乳腺癌的C3Tag GEMM测试。目标是确定克服耐药性并引起细胞凋亡和肿瘤消退的联合疗法。