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: 8607578
• 负责人: GARY L. JOHNSON
• 金额: $ 27.72万
• 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/8607578
• 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; PDGFRB gene; 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; SUM-159 Breast Cancer Cell Line; 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）：已经开发出一种新方法来研究“整体”蛋白激酶的重编程，允许在单次质谱运行中分析约 40-60% 的表达激酶组。我们的方法利用多重抑制剂珠子 (MIB)，该珠子由带有共价固定、接头适应的激酶抑制剂的琼脂糖珠子组成。该技术允许对序列已知的激酶进行询问，但由于缺乏生物学或表型知识或试剂的可用性，这些激酶尚未得到充分研究。 MIB/MS 确定了针对特定 MEK1/2 抑制剂 AZD6244 的激酶组反应特征，该抑制剂目前正在进行三阴性乳腺癌的临床测试。 MEK1/2 唯一确定的底物是 ERK1 和 2，但我们观察到激酶组每个亚家族中激酶活性因 MEK 抑制而发生变化。激酶组评估显示，时间依赖性重编程涉及 RAF 和 MEK 的 ERK 反馈调节的早期丧失，从而允许上游 MEK-ERK 通路重新激活。特定受体酪氨酸激酶 (RTK)（例如 PDGFR¿和 DDR1）的 MIB 结合的时间依赖性变化很容易检测到，并提供了关键的实验观察结果，即 MEK 抑制正在驱动多个 RTK 的表达和激活。 c-Myc 降解是介导激酶组重编程的关键机制。多个 RTK 因 MEK 抑制而被激活的事实表明，使用单一激酶抑制剂来阻止肿瘤进展是困难的。该提案的目标包括： 1. 定义人类 basal/claudin-low 样 SUM159 细胞系和用于 basal/claudin-low 乳腺癌的临床前 C3Tag GEMM 中的激酶组激活状态。激酶特征将根据单独和组合的 MEK1/2 和 PI3K 抑制剂来定义，目前正在进行临床测试。 2. 定义 MEK 和 PI3K 抑制剂抗性肿瘤和细胞系中激酶组重编程的机制。 3. 根据敏感和耐药肿瘤的激酶组特征对激酶抑制剂组合进行合理预测，以在 C3Tag GEMM 中测试基底/密蛋白低乳腺癌。目标是确定克服耐药性并导致细胞凋亡和肿瘤消退的联合疗法。