Therapeutically relevant targets of Twist1 dimers in glioma

神经胶质瘤中 Twist1 二聚体的治疗相关靶点

• 批准号: 9011554
• 负责人: ROBERT C ROSTOMILY
• 金额: $ 38.06万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-02-15 至 2016-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9011554
• 关键词: Address; Affinity; Affinity Chromatography; Behavior; Binding; Bioinformatics; Carcinoma; Categories; Cell Line; Cells; ChIP-seq; Co-Immunoprecipitations; Complex; DNA Binding; DNA Binding Domain; Databases; Development; Dimerization; Epithelial; Failure; Family; Gene Chips; Gene Expression; Gene Expression Profile; Gene Expression Profiling; Genes; Genetic Transcription; Glioblastoma; Glioma; Growth; Homo; Human; In Vitro; Malignant - descriptor; Malignant Glioma; Malignant Neoplasms; Malignant neoplasm of brain; Maps; Mass Spectrum Analysis; Mediating; Mesenchymal; Methods; Molecular Analysis; Molecular Profiling; Multiple Partners; Neoplasm Metastasis; Pathway interactions; Pharmaceutical Preparations; Phenocopy; Phenotype; Phosphorylation; Play; Population; Property; Proteins; Proteomics; Regulation; Resistance; Role; Stem cells; TWIST1 gene; Testing; Therapeutic; Therapeutic Equivalency; To specify; Translating; Tumorigenicity; Work; angiogenesis; base; chromatin immunoprecipitation; clinically relevant; dimer; epithelial to mesenchymal transition; in vitro testing; in vivo; inhibitor/antagonist; innovation; insight; knock-down; malignant phenotype; malignant state; mutant; nerve stem cell; periostin; public health relevance; self-renewal; stem; stemness; targeted treatment; therapeutic target; therapy resistant; tool; transcription factor; tumor; tumor progression; tumorigenic

 DESCRIPTION (provided by applicant): Glioblastoma (GBM) is among the most lethal human cancers, but despite decades of extensive effort, median survivals remain one year, or less. Because invasion and stemness are inter-connected properties that drive much of glioma malignancy, genes that co-regulate these phenotypes are robust therapeutic targets. One such candidate is TWIST1 (TW), a bHLH transcription factor (TF) central to epithelial mesenchymal transition (EMT), invasion and stemness and metastasis in carcinomas and normal development. In gliomas TW is upregulated in higher grade tumors and promotes GBM cell invasion and stemness in vitro and viability and growth of glioma stem cells (GSCs) in vivo. These findings strongly support its potential as a therapeutic target for GBM but identifying drug inhibitors of TFs has proven difficult by traditional means. The purpose of this proposal then is t develop indirect methods for targeting TW in GSCs through downstream genes essential to its malignant activity. TW must form a homo or heterodimers with other bHLH proteins to regulate gene transcription and the specific dimer motif defines unique phenotypes and gene expression sub-networks. The activation of unique gene expression patterns, or sub-networks, specific TW dimer partners which also map to specific malignant phenotypes will be leveraged to identify targets critical for TW activation of GSC invasion and survival. Further, phosphorylation of TW regulates dimer partner affinities and invasive behavior of GBM cells. Based on these observations this proposal will test the hypothesis that TW dimer specific pathways (sub-networks) can identify therapeutic targets to inhibit TW-regulated malignancy. AIM 1 employs proteomics to profile TW bHLH family binding partners associated with increased or decreased invasion and stemness. To facilitate this screen differential invasion and dimer affinity will be introduced through expression of pro- invasive wild-type and an anti-invasive TW phospho-mutant. In AIM2 candidate dimers validated to regulate TW binding and invasive or stem-like phenotypes will be tested to establish their impact on tumorigenicity and malignant phenotypes (invasion, proliferation, survival and angiogenesis) in vivo. In AIM3 gene expression arrays and ChIP-seq will define TW dimer specific sub-networks and identify direct and indirectly regulated TW targets. Candidate targets rank-ordered by comprehensive bioinformatic analysis of GO functional categories, pathways and expression in public GBM databases will then be tested in vitro and in vivo to validate equivalency with TW inhibition in selected GSC lines. This innovative approach is expected to identify TW dimer specific gene expression sub-networks and targets that functionally phenocopy the effects of TW inhibition on invasion, GSC tumorigenicity and glioma malignancy. This strategy is expected to have therapeutic relevance that translates to other TFs currently considered "undruggable". Of significance, these studies will provide new mechanistic insight with broad implications for understanding the role of TW in development, EMT and the many other cancers in which TW promotes malignancy.

 描述（申请人提供）：胶质母细胞瘤（GBM）是最致命的人类癌症之一，但尽管经过数十年的广泛努力，中位生存期仍为一年或更短。由于侵袭和干性是驱动胶质瘤恶性肿瘤的相互关联的特性，共同调节这些表型的基因是强大的治疗靶点。一种这样的候选物是TWIST 1（TW），一种bHLH转录因子（TF），其在上皮间质转化（EMT）、癌中的侵袭和干性以及转移和正常发育中起中心作用。在神经胶质瘤中，TW在更高级别的肿瘤中上调，并在体外促进GBM细胞侵袭和干性，在体内促进神经胶质瘤干细胞（GSC）的活力和生长。这些发现强烈支持其作为GBM的治疗靶点的潜力，但通过传统手段鉴定TF的药物抑制剂已被证明是困难的。因此，本提案的目的是开发通过其恶性活性所必需的下游基因靶向GSC中TW的间接方法。TW必须与其他bHLH蛋白形成同源或异源二聚体来调节基因转录，并且特定的二聚体基序定义了独特的表型和基因表达子网络。将利用独特基因表达模式或子网络、也映射到特定恶性表型的特定TW二聚体伴侣的激活来鉴定对GSC侵袭和存活的TW激活至关重要的靶标。此外，TW的磷酸化调节GBM细胞的二聚体伴侣亲和力和侵袭行为。基于这些观察结果，该提议将检验TW二聚体特异性途径（子网络）可以鉴定治疗靶点以抑制TW调节的恶性肿瘤的假设。目的1采用蛋白质组学分析TW bHLH家族结合伴侣与增加或减少的侵袭和干性。为了促进该筛选，将通过表达促侵袭野生型和抗侵袭TW磷酸突变体来引入差异侵袭和二聚体亲和力。在AIM 2中，将测试经验证可调节TW结合和侵袭性或干细胞样表型的候选二聚体，以确定其对体内致瘤性和恶性表型（侵袭、增殖、存活和血管生成）的影响。在AIM 3基因表达阵列和ChIP-seq中，将定义TW二聚体特异性子网络，并识别直接和间接调节的TW靶标。然后将在体外和体内测试通过对公共GBM数据库中的GO功能类别、途径和表达进行综合生物信息学分析排序的候选靶标，以验证与所选GSC系中TW抑制的等效性。这一创新 这种方法有望鉴定TW二聚体特异性基因表达子网络和靶点，其功能表型复制TW抑制对侵袭、GSC致瘤性和胶质瘤恶性度的影响。预期该策略具有治疗相关性，其转化为目前被认为“不可用药”的其他TF。重要的是，这些研究将为理解TW在发育、EMT和TW促进恶性肿瘤的许多其他癌症中的作用提供新的机制见解。