Investigating mechanisms of kinase-mediated cell migration in aggressive glioblastoma

研究侵袭性胶质母细胞瘤中激酶介导的细胞迁移机制

• 批准号: 9752228
• 负责人: Kyle P. Mohler
• 金额: $ 6.12万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9752228
• 关键词: Adopted; Adult; Alanine; Amino Acids; Biological; Biological Assay; Brain Neoplasms; Cancer Biology; Cations; Cell Culture Techniques; Cell Proliferation; Cell Survival; Cell Volumes; Cells; Chemicals; Chlorides; Complex; Coupled; Cytoskeleton; Data; Development; Diabetes Mellitus; Disease; Disease Progression; Enzyme-Linked Immunosorbent Assay; Family; Future; Glioblastoma; Glioma; Goals; Homeostasis; Hypertension; Investigation; Ion Channel; Lead; Link; Maintenance; Malignant - descriptor; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Mediation; Microfluidics; Migration Assay; Mutation; Nature; Output; Pathway interactions; Pharmacology; Phenotype; Phosphorylation; Phosphorylation Site; Phosphothreonine; Phosphotransferases; Physiological; Physiological Processes; Physiology; Play; Post-Translational Protein Processing; Preparation; Production; Proline; Proteins; Proteome; Proteomics; Regulation; Reporting; Role; Signal Pathway; Signal Transduction; Site; Speed; Stable Isotope Labeling; Structure; System; Techniques; Translations; Validation; Work; base; cancer cell; cancer invasiveness; cancer type; cell motility; cross reactivity; environmental change; experimental study; human disease; improved; inhibitor/antagonist; insight; kinase inhibitor; knock-down; migration; novel; outcome forecast; prevent; small hairpin RNA; small molecule; small molecule inhibitor; targeted treatment; therapeutic target; tumor progression

Project Summary – Mohler Glioblastoma multiforme (GBM) is the deadliest brain tumor in adults, characterized by rapid progression and poor prognosis due to its highly proliferative and invasive nature. GBM is dynamically heterogeneous with a complex set of inputs ultimately determining a set of outputs (phenotypes) related to cell survival, proliferation, and invasive migration. As a whole, the goals of this project are structured to provide insight into mechanisms of glioblastoma cell migration. The approach is centered on assessing the contributions of underexplored kinase regulatory networks to the phenotypic switch of cancer cells from proliferative to migratory states, underlying aggressive disease progression. As the most abundant post-translational modifications in eukaryotic signaling pathways, protein phosphorylation occupies a central role in regulatory networks and the maintenance of cellular homeostasis, yet their roles in the mediation of cell migration are poorly understood. It has recently been shown that GBM cells can use normal physiological processes for cell migration, such as exploiting ion channels like NKCC1, to promote motility. The Ste20-family kinase SPS1-related proline/alanine-rich kinase (SPAK) relays changes in cell volume to cation-chloride cotransporters (NKCCs and KCCs) to maintain cellular homeostasis and control cell migration. Although previous reports have highlighted the importance of SPAK kinase as a potential therapeutic target for the treatment of GBM, the inability to produce high yields of physiologically phosphorylated kinase, until now, impeded this progress. Leveraging recent advances in orthogonal translation systems and mass spectrometry from the Rinehart lab enables the implementation of a robust pipeline for the identification and validation of lead inhibitor compounds which target physiologically phosphorylated, active SPAK kinase. The impact of candidate SPAK inhibitors on the migration and physiology of GBM cells will be assessed using a microfluidics based 1D cell migration assays. In parallel, the direct interactions and impacts of candidate inhibitor compounds with the GBM proteome will defined using quantitative mass spectrometry techniques. Overall, characterization of small molecule inhibitor compounds identified from this pipeline will provide important mechanistic insight into the role of kinase networks in the regulation of cancer cell migration, expanding potential applications to the broader scope of cancer biology.

项目概要- Mohler 多形性胶质母细胞瘤（GBM）是成人中最致命的脑肿瘤，其特征是快速进展， 由于其高度增殖性和侵袭性，预后差。GBM是动态异构的， 最终确定与细胞存活，增殖， 和入侵性迁移。作为一个整体，本项目的目标是提供深入了解的机制， 胶质母细胞瘤细胞迁移。该方法的核心是评估未充分开发的激酶的贡献 调节网络的癌细胞从增殖到迁移状态的表型转换， 侵袭性疾病进展。作为真核生物信号转导中最丰富的翻译后修饰 蛋白质磷酸化在调节网络和维持细胞内的 然而，它们在介导细胞迁移中的作用知之甚少。最近已经表明 GBM细胞可以使用正常的生理过程进行细胞迁移，例如利用离子通道， NKCC 1，促进运动。Ste 20家族激酶SPS 1相关的脯氨酸/丙氨酸丰富激酶（SPAK）中继 改变细胞体积，以阳离子-氯共转运蛋白（NKCC和KCC）维持细胞稳态 并控制细胞迁移。尽管先前的报道已经强调了SPAK激酶作为一种免疫调节剂的重要性， GBM治疗的潜在治疗靶点，不能产生高产量的生理性 到目前为止，磷酸化激酶阻碍了这一进程。利用正交平移的最新进展 系统和质谱从莱因哈特实验室能够实现一个强大的管道， 鉴定和验证靶向生理磷酸化、活性 SPAK激酶。将研究候选SPAK抑制剂对GBM细胞的迁移和生理学的影响。 使用基于微流体的1D细胞迁移测定法进行评估。与此同时， 将使用定量质谱法确定具有GBM蛋白质组的候选抑制剂化合物 技术.总体而言，从该管道中鉴定的小分子抑制剂化合物的表征将 为激酶网络在调节癌细胞迁移中的作用提供了重要的机制见解， 将潜在应用扩展到更广泛的癌症生物学范围。