Targeting the MEKK2-ERK5 Signaling Node in Triple Negative Breast Cancer

靶向三阴性乳腺癌中的 MEKK2-ERK5 信号节点

• 批准号: 8332123
• 负责人: John Scott
• 金额: $ 11.16万
• 依托单位: NORTH CAROLINA CENTRAL UNIVERSITY
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8332123
• 关键词: Adenocarcinoma Cell; Alanine; Apoptosis; Back; Binding; Biochemical; Biological Assay; Biological Markers; Bioluminescence; Biopsy; Breast Adenocarcinoma; Breast Cancer Cell; Cell Cycle; Cell Line; Cells; Co-Immunoprecipitations; Complex; Disease Progression; Drug usage; Epidermal Growth Factor Receptor; Fatty acid glycerol esters; Female; Fluorescence Resonance Energy Transfer; Future; Gene Expression; Genes; Genetically Engineered Mouse; Goals; Growth; Human; Image; In Vitro; Lead; Luciferases; MAP Kinase Gene; MAP Kinase Signaling Pathways; MAPK11 gene; MAPK14 gene; MAPK7 gene; MAPK8 gene; Malignant Neoplasms; Malignant neoplasm of prostate; Mammary Neoplasms; Mammary gland; Measures; Mediating; Monitor; Mutate; Mutation; Neoplasm Metastasis; Pathway interactions; Peptide Hydrolases; Phenotype; Phosphorylation; Phosphotransferases; Protein Analysis; Protein Tyrosine Kinase; Regulation; Research; Role; SCID Mice; Signal Transduction; Specificity; System; Testing; Time; Tumor Angiogenesis; Tyrosine; Tyrosine Phosphorylation; Ultrasonography; Vascularization; Xenograft Model; Xenograft procedure; abstracting; anticancer research; base; cytokine; erbB-2 Receptor; in vivo; inhibitor/antagonist; kinase inhibitor; knock-down; malignant breast neoplasm; matrigel; mouse model; mutant; neoplastic cell; new therapeutic target; outcome forecast; overexpression; response; small hairpin RNA; small molecule; small molecule libraries; tool; triple-negative invasive breast carcinoma; tumor; tumor growth; tumorigenesis; tumorigenic

Abstract: Approximately 15-20% of breast cancers have a triple negative phenotype (negative for ER and PR expression and lack ErbB2 overexpression) that correlates with aggressive cancer and limited treatment options. We have developed an in vivo screen for defining the role of specific kinases in tumorigenesis and metastasis of breast cancer cells. Triple negative and triple positive breast cancer lines were used for shRNA-mediated knock down of specific kinases. The genetically altered lines (expressing luciferase in addition to specific shRNA gene knockdown) are injected into the mammary fat pad of female SCID mice. Tumor growth and vascularization is monitored longitudinally over an 8 week period using a micro-probe ultrasound system. Metastasis is monitored by bioluminescence imaging. The assay provides an in vivo screen for analysis of proteins that control the growth, vascularization and metastasis of breast tumors. MAP3Ks are the first tier of kinases regulating the MAP kinase signaling pathways that lead to the activation of the MAPKs ERKl/2, p38, JNK and ERKS. MAPSKs control expression of genes important for regulating the cell cycle, cytokine and protease expression and apoptosis. In a screen of 9 MAP3Ks, MEKK2 was identified as a key regulator of metastasis using MDA-MB-231 (triple negative basal) and BT474 (triple positive luminal) breast adenocarcinoma cells in the in vivo tumorigenesis assay. MEKK2 is a MAP3K that regulates the activation of the JNK and ERK5 pathways via activation of MKK7 and MEKS. We have shown that MEKK2 expression is required for EGFR (ErbBI) and ErbB2/Neu activation of ERKS in MDA-MB-231 and BT474 cells, respectively. Our hypothesis is that the MAP3K MEKK2 functions as a critical signaling node within the cell signaling network stimulating tumor growth and metastasis in response to ErbB and possibly other tyrosine kinases. The goal of this proposal is to genetically define the role of MEKK2 in triple negative breast cancer tumor growth and metastasis and to develop a MEKK2 small molecule inhibitor. Specific aim 1 involves defining the role of MEKK2-MEK5-ERK5 signaling in tumorigenesis and metastasis of triple negative breast cancer cells using the in vivo xenograft assay. In specific aim 2, we propose to elucidate the mechanism by which MEKK2 gets activated by ErbB1/2 using MEKK2 mutants in biochemical and cell-based activation assays. In specific aim 3, a small molecule biochemical screen will be developed to identify compounds that specifically inhibit MEKK2 kinase activity. MEKK2 inhibitors will be tested for MAPK pathway specificity in cell-based assays and profiled for specificity against the kinome. In future studies, these MEKK2 inhibitors will be tested for anti-tumor efficacy in genetically engineered mouse models (GEMMs) of breast cancer.

抽象的： 大约15-20％的乳腺癌具有与侵略性癌症和有限的治疗选择相关的三重阴性表型（ER和PR表达负面，缺乏ERBB2过表达）。我们开发了一个体内筛选，以定义特定激酶在乳腺癌细胞的肿瘤发生和转移中的作用。三重阴性和三阳性乳腺癌系用于SHRNA介导的特定激酶的敲低。遗传改变的线（表达荧光素酶以外，除了特定的shRNA基因敲低外）被注入雌性SCID小鼠的乳腺脂肪垫中。肿瘤生长和血管化是 使用微型探针超声系统在8周内纵向监视。通过生物发光成像监测转移。该测定提供了一个体内筛选，用于分析控制乳腺肿瘤的生长，血管化和转移的蛋白质。 MAP3KS是调节地图的激酶的第一层 激酶信号通路会导致MAPKS ERKL/2，P38，JNK和ERK的激活。 MAPSK控制基因的表达对于调节细胞周期，细胞因子和蛋白酶表达和凋亡至关重要。在9个MAP3KS的屏幕中，MEKK2使用MDA-MB-231确定为转移的关键调节剂 （三阴性基底）和BT474（三阳性腔）乳腺腺癌细胞中的体内肿瘤发生分析。 MEKK2是一个MAP3K，可通过激活MKK7和MEK来调节JNK和ERK5途径的激活。我们已经表明，EGFR（ERBBI）需要MEKK2表达 MDA-MB-231和BT474细胞中ERBB2/NEU激活。我们的假设是MAP3K MEKK2在细胞信号网络中起作用，刺激肿瘤生长和转移，响应于ERBB以及可能其他酪氨酸激酶。该提案的目的是从基因上定义MEKK2在三重阴性乳腺癌肿瘤生长和转移中的作用，并开发MEKK2小分子抑制剂。具体目标1涉及使用体内异种移植测定法定义MEKK2-MEK5-ERK5信号传导在肿瘤发生和三阴性乳腺癌细胞转移中的作用。具体 AIM 2，我们建议阐明使用MEKK2突变体在生化和基于细胞的激活测定中使用MEKK2突变体激活MEKK2的机制。在特定的目标3中，将开发一个小分子生化筛选，以鉴定特异性抑制MEKK2激酶活性的化合物。 MEKK2抑制剂将是 测试了基于细胞测定的MAPK途径特异性，并介绍了针对Kinome的特异性。在未来的研究中，这些MEKK2抑制剂将在乳腺癌的基因工程小鼠模型（GEMM）中测试抗肿瘤功效。