Plexin-B2 function in glioma invasion and glioma stem cell maintenance

Plexin-B2 在神经胶质瘤侵袭和神经胶质瘤干细胞维持中的功能

• 批准号: 9106872
• 负责人: Roland Horst Friedel
• 金额: $ 38.32万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9106872
• 关键词: Actins; Address; Adopted; Biological Assay; Brain; Brain Neoplasms; CRISPR/Cas technology; Cadherins; Cell Adhesion Molecules; Cell Line; Cell Maintenance; Cell Proliferation; Cells; Characteristics; Clustered Regularly Interspaced Short Palindromic Repeats; Corpus Callosum; Cues; Cytoskeleton; DNA Sequence Alteration; Data; Databases; Excision; Exhibits; Family; Gene Targeting; Genes; Genetic Epistasis; Glioblastoma; Glioma; Growth; Guanosine Triphosphate Phosphohydrolases; Heterogeneity; Human; In Vitro; Infiltrative Growth; Investigation; Knock-out; Knowledge; Link; Maintenance; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of brain; Mediating; Migration Assay; Modeling; Molecular; Monomeric GTP-Binding Proteins; Mutation; Neural Cell Adhesion Molecule L1; Operative Surgical Procedures; Pathway interactions; Patients; Phenotype; Pilot Projects; Play; Primary Brain Neoplasms; Property; Recurrence; Resistance; Role; SEMA3F gene; STAT3 gene; Semaphorins; Signal Pathway; Signal Transduction; Slice; Source; Stem Cell Factor; Stem cells; Structure; Technology; Testing; Transplantation; Treatment Protocols; Tumorigenicity; Twin Multiple Birth; Up-Regulation; Vascularization; arm; axon guidance; base; cancer stem cell; cell growth; cell motility; cell type; cohort; combinatorial; glioma cell line; in vivo; insight; knock-down; member; migration; mutant; neoplastic cell; new therapeutic target; novel; novel marker; novel therapeutics; outcome forecast; overexpression; plexin; public health relevance; radiation resistance; receptor; receptor binding; rho; rho GTP-Binding Proteins; self-renewal; small hairpin RNA; stem; stem cell niche; stemness; targeted treatment; therapy resistant; transcription factor; transcriptome sequencing; tumor; tumor progression; tumorigenic

 DESCRIPTION (provided by applicant): Glioblastoma multiforme (GBM) remains the most deadly form of brain cancer. There is a dire need to better define the molecular pathways that regulate the malignant features of GBM, in particular, its invasiveness and the persistence of glioma stem cells (GSC), also known as tumor propagating cells, which are thought to be the main source of tumor recurrence due to their tumorigenic potential and therapy resistance. Plexin-B2 is an axon guidance receptor originally identified as a gene highly upregulated in glioma. Our recent patient database analyses found that upregulation of Plexin-B2 correlates with glioma grade and poor survival. This prompted us to investigate the mechanistic basis of this important link. Our studies in human ATCC glioma cell lines demonstrated that Plexin-B2 activation alters actin cytoskeleton and promotes cell migration through activation of Rho GTPases. In intracranial transplants, Plexin-B2 shRNA knockdown hindered invasion and disrupted glioma vascularization. Our new preliminary data from patient-derived GSC lines suggest that besides a promigratory role, Plexin-B2 may also regulate glioma stem cell phenotypes, including stem cell marker expression, self-renewal capacity, proliferation and differentiation potential. Building on these exciting results, we propose to test the central hypothesis that Plexin-B2 plays a multifaceted role in promoting glioma invasion and stem cell characteristics. To facilitate our studies, we have adopted CRISPR gene editing technology to generate bi-allelic mutations of Plexin-B2 in patient-derived GSC lines. In Aim 1, we focus on glioma invasion and ask if Plexin-B2 signaling influences cytoskeletal dynamics and cellular motility of GBM cells by performing culture assays, brain slice cultures, and intracranial transplants. By comparing cell migration of GSCs and their differentiated progeny, this set of studies also addresses whether migratory potential is contingent on differentiation hierarchy. In Aim 2, we focus on glioma stem cell characteristics and ask if Plexin-B2 contributes to stem cell marker expression, self-renewal capacity, proliferation and differentiation potential, and treatment resistance. We will investigate in survival cohorts with GSC intracranial transplants if Plexin-B2 deletion confers survival benefit and if it reduces radiation resistance. In Aim 3, we focus on functional targets and downstream effectors of Plexin-B2 signaling in mediating migration and stem cell properties. We ask which intracellular domain of Plexin-B2 is the main signaling arm, if cellular migration is mediated through modulation of Rho GTPases, whether STAT3 functions as a core signaling hub, and if stem cell factors SOX2 or OLIG2 function downstream of STAT3 for stemness maintenance. Lastly, we will study the adhesion molecules L1CAM and cadherins/protocadherins, the most significantly regulated group of genes that were identified in our recent RNA-Seq analysis of Plexin-B2 knockout GSC. Understanding how Plexin-B2 regulates the malignant features of GBM will open up possibilities for new targets to inhibit glioma growth and invasion.

 描述（由申请人提供）：多形性胶质母细胞瘤（GBM）仍然是最致命的脑癌形式。迫切需要更好地定义调节GBM的恶性特征的分子途径，特别是其侵袭性和胶质瘤干细胞（GSC）（也称为肿瘤增殖细胞）的持续性，由于其致瘤潜力和治疗抗性，其被认为是肿瘤复发的主要来源。丛蛋白-B2是一种轴突导向受体，最初被确定为在胶质瘤中高度上调的基因。我们最近的患者数据库分析发现，丛蛋白-B2的上调与胶质瘤分级和不良生存率相关。这促使我们研究这一重要环节的机制基础。我们在人ATCC神经胶质瘤细胞系中的研究表明，丛蛋白-B2活化改变肌动蛋白细胞骨架，并通过Rho GTP酶的活化促进细胞迁移。在颅内移植中，丛状蛋白-B2 shRNA敲低阻碍了侵袭并破坏了胶质瘤血管形成。我们来自患者来源的GSC系的新的初步数据表明，除了促迁移作用，丛蛋白-B2还可以调节胶质瘤干细胞表型，包括干细胞标志物表达，自我更新能力，增殖和分化潜力。基于这些令人兴奋的结果，我们建议测试中心假设，即丛蛋白B2在促进胶质瘤侵袭和干细胞特性方面发挥多方面的作用。为了促进我们的研究，我们采用CRISPR基因编辑技术在患者来源的GSC系中产生丛蛋白-B2的双等位基因突变。在目标1中，我们专注于胶质瘤侵袭，并询问丛蛋白-B2信号是否通过进行培养测定、脑切片培养和颅内移植影响GBM细胞的细胞骨架动力学和细胞运动性。通过比较GSC及其分化后代的细胞迁移，这组研究还解决了迁移潜力是否取决于分化等级。在目标2中，我们关注胶质瘤干细胞的特征，并询问丛蛋白-B2是否有助于干细胞标志物表达、自我更新能力、增殖和分化潜力以及治疗抗性。我们将在GSC颅内移植的生存队列中研究丛蛋白-B2缺失是否带来生存益处以及是否降低辐射抗性。在目标3中，我们专注于丛蛋白-B2信号传导在介导迁移和干细胞特性中的功能靶标和下游效应物。我们询问丛蛋白-B2的哪个胞内结构域是主要的信号传导臂，细胞迁移是否通过Rho GTP酶的调节介导，STAT 3是否作为核心信号传导枢纽发挥作用，以及干细胞因子SOX 2或OLIG 2是否在STAT 3下游发挥作用以维持干性。最后，我们将研究粘附分子L1 CAM和钙粘蛋白/原钙粘蛋白，这是我们最近对丛蛋白-B2敲除GSC的RNA-Seq分析中鉴定的最显著调节的基因组。了解丛蛋白B2如何调节GBM的恶性特征将为抑制胶质瘤生长和侵袭的新靶点开辟可能性。