Exploring the Therapeutic Potential of Stem Cell Biology in Gliomas

我们已经进行了实验，以提高我们的理解的分子机制的失调分化途径在TSCs：干细胞自我更新和分化之间的微妙的平衡是由各种细胞的内在和外在因素，是至关重要的正常组织的稳态控制。尽管GSC和正常干细胞之间存在广泛的表型和功能相似性，但GSC的分化潜力并不完全正常。阐明在正常干细胞和GSC中起作用的分化途径对于充分理解肿瘤发生至关重要，并可能导致新的治疗靶点。我们还鉴定了一组在源自人原代GBM的GSC中去调节的分化途径。阐明其分子机制将为预测分化治疗方案的敏感性提供重要线索。肿瘤干细胞在分化诱导剂方面的特性进一步揭示了传统血清培养胶质瘤细胞系的局限性。例如，视黄酸处理和CNTF暴露有效地诱导大多数GBM肿瘤起始细胞（TIC）的分化，但不诱导传统细胞系的分化。这促使我们质疑是否许多潜在的肿瘤抑制基因和/或细胞生长抑制基因先前在细胞系中研究，没有被识别。鉴于从生物信息学方法和实验室积累的干细胞培养技术专业知识中鉴定出的胶质母细胞瘤TSC中潜在的GSG和癌基因数量不断增加，我们建立了筛选系统来研究这些基因在干细胞培养中的功能。此外，我们在关键项目之一，即了解我们的胶质瘤TSCs和正常神经干细胞（NSC）之间的基因组和分子信号传导相似性和差异方面取得了重大进展。我们已经在增殖和分化条件下对7种不同的GBM衍生的TSC和正常胚胎NSC系进行了非常大规模的研究，并获得了高通量mRNA和microRNA谱。自2014年11月，当吉尔伯特博士启动新的胶质瘤干细胞转化研究项目时，我们又创建了6个胶质瘤源性TSC，引入了3个IDH突变的TSC，随后进行了计算分析，以表征这些GSC中的遗传学和信号通路，使这些细胞系能够用于探索胶质瘤干细胞生物学的治疗潜力，如代谢变化和免疫反应。此外，我们通过合作和材料转让协议获得了重要的细胞系，为实验室提供了出色的资源。目前，NOB实验室共有62种人类肿瘤细胞系。作为该项目的一部分，这些细胞系中的每一种都已被广泛地分子表征，包括全外显子组测序、甲基化组分析和RNA测序。这些数据被上传到一个数据库系统中，以便为NOB的所有研究小组进行临床前研究分析和选择最佳模型系统。

We have performed experiments to improve our understanding of the molecular mechanisms of deregulated differentiation pathways in TSCs: The delicate balance between stem cell self-renewal and differentiation is controlled by various cell intrinsic and extrinsic factors that are critical for normal tissue homeostasis. Despite extensive phenotypic and functional similarities between GSCs and normal stem cells, the differentiation potentials of GSCs are not entirely normal. Elucidation of the differentiation pathways that are operative in both normal stem cells and GSCs will be critical for fully understanding tumorigenesis and will likely lead to novel therapeutic targets. We have also identified a set of deregulated differentiation pathways in GSCs derived from human primary GBM. Elucidation of underlying molecular mechanism will provide important clues for predicting sensitivity of differentiation therapeutic approach. Characterization of TSCs in aspect of differentiation-inducing agents further revealed the limitations of traditional glioma cell lines grown in serum. For example, retinoic acid treatment and CNTF exposure potently induce differentiation in most GBM tumor initiate cells (TICs) but not of traditional cell lines. This prompted us to question whether many of potential tumor suppressors and/or cytostatic genes previously studied in cell lines, were not recognized. Given the ever-increasing number of potential GSGs and oncogenes in glioblastoma TSCs identified from bioinformatics approaches and technical expertise of stem cell culture accumulated in the laboratories, we have set up screening systems to study the function of these genes in stem cell cultures. In addition, we have made significant progress on one of keystone projects that is to understand the genomic and molecular signaling similarities and differences between our glioma TSCs and normal neural stem cells (NSC). We have performed a very large scale study of 7 different GBM-derived TSCs and normal embryonic NSC lines under both proliferative and differentiating conditions and derived high-throughput mRNA and microRNAs profiling. Since November of 2014, when Dr Gilbert initiated the new glioma stem cell translational study project, we have created 6 more glioma-derived TSCs, introduced 3 IDH- mutated TSCs, and subsequently performed the computational analyses for characterization of the genetics and signaling pathways in these GSCs, enabling these cell lines to be used to explore the therapeutic potential of glioma stem cell biology, such as metabolic changes and immunological reactions. Additionally, we have through collaborations and Material Transfer Agreements obtained important cell lines that provide an outstanding resource for the laboratory. In total, there are now 62 human tumor lines in the NOB Laboratory. As part of this project, each of these cell lines has been extensively molecularly characterized including whole exome sequencing, analysis of the methylome and RNA sequencing. These data are being uploaded into a database system to enable analysis and selection of optimal model systems for preclinical studies for all of the research groups in the NOB.