Exploring the Therapeutic Potential of Stem Cell Biology in Gliomas

TSCs中去调节分化途径的分子理解： 干细胞自我更新和分化之间的平衡由各种细胞控制， 对正常组织稳态至关重要的内在和外在因素。尽管 TSC和正常干细胞之间广泛的表型和功能相似性， TSCs的分化潜能并不完全正常。区分的阐明 在正常干细胞和TSC中起作用的通路对于充分理解 肿瘤发生，并可能导致新的治疗靶点。我们发现了一组 来源于人原发性胶质母细胞瘤的TSC中的失调的分化途径。我们 证明骨形态发生蛋白（BMP）介导的和睫状神经营养的 CNTF介导的Janus激酶/信号转导和转录激活因子 (Jak/STAT）途径在成人脑源性TSC中引起不同的生物学后果 与正常的神经干细胞（NSC）相比。与非常早期的胚胎NSC一样，0308 TSCs 在CNTF刺激后，BMP反应性增殖并引起边缘STAT3活化。 然而，不像正常的神经干细胞在以后的发展阶段，获得反应， CNTF触发的STAT3激活与BMP受体1B表达增加相关 （BMPR1B），0308细胞不表达Zeste增强子同源物2继发的BMPR1B （EZH2）依赖性BMPR1B启动子高甲基化。BMPR1B在0308 TIC中的强制表达 通过转基因表达或启动子的去甲基化， 分化能力，并诱导其致瘤性的丧失，不仅通过 BMP介导的途径，但也通过CNTF介导的Jak/STAT激活。对54名小学生的调查 人类胶质母细胞瘤显示，大约20%的人BMPR1B表达受到抑制， 与启动子高甲基化有关。综合起来，这些数据表明， 胶质母细胞瘤TSCs亚群中BMP发育分化途径的失调 不仅通过使TIC对正常细胞的敏感性降低， 分化线索，但也通过转换，否则细胞抑制信号，以促增殖 信号.通过使用GBM TSC进行广泛的体外和体内表征， 分化诱导剂如视黄酸证明这些TSC 有效分化并阻止增殖。我们已经证明视黄酸 治疗通过降低CD133阳性细胞的比例达到细胞生长抑制作用， 脑TSCs的推定标志物，来自肿瘤并通过诱导分化成星形胶质细胞 脉有趣的是，用放疗和化疗预处理的GBM TIC亚组， 在体内的药物，没有显示出显着的视黄酸介导的分化。阐发 分子机制的研究将为预测药物敏感性提供重要线索。 辨证论治。TSC在以下方面的表征： 分化诱导剂进一步揭示了传统胶质瘤细胞的局限性， 在血清中生长的细胞系。例如，视黄酸处理和CNTF暴露有效地诱导 在大多数GBM TIC中，但不是传统细胞系的分化。这促使我们 问题是，许多潜在的肿瘤抑制基因和/或细胞抑制基因先前是否 在细胞系中研究，没有被识别。鉴于潜在的技术支持小组的数量不断增加， 从生物信息学方法和技术中鉴定胶质母细胞瘤TSCs中的癌基因 在实验室积累了干细胞培养的专业知识，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。

Molecular understanding 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 TSCs and normal stem cells, the differentiation potentials of TSCs are not entirely normal. Elucidation of the differentiation pathways operative in both normal stem cells and TSCs will be critical for fully understanding tumorigenesis and will likely lead to novel therapeutic targets. We have identified a set of deregulated differentiation pathways in TSCs derived from human primary glioblastoma. We demonstrated that both bone morphogenetic protein (BMP)-mediated and ciliary neurotrophic factor (CNTF)-mediated Janus kinase/signal transducer and activator of transcription (Jak/STAT) pathways elicit distinct biological consequences in adult brain derived TSCs compared to normal neural stem cells (NSCs). Like very early embryonic NSCs, 0308 TSCs proliferate in response to BMP and elicit marginal STAT3 activation after CNTF challenge. However, unlike normal NSCs in later developmental stages that acquire responsiveness to CNTF-triggered STAT3 activation in association with increased expression of BMP receptor 1B (BMPR1B), 0308 cells do not express BMPR1B secondary to Enhancer of Zeste homolog 2 (EZH2)-dependent BMPR1B promoter hypermethylation. Forced expression of BMPR1B in 0308 TICs either by transgene expression or demethylation of the promoter fully restores their differentiation capabilities and induces loss of their tumorigenicity not only via a BMP-mediated pathway but also by CNTF-mediated Jak/STAT activation. A survey of 54 primary human glioblastomas reveals that approximately 20% have suppressed expression of BMPR1B associated with promoter hypermethylation. Taken together, these data implicate that deregulation of the BMP developmental differentiation pathway in a subset of glioblastoma TSCs contributes to their tumorigenic phenotype by not only desensitizing TIC to normal differentiation cues, but also by converting otherwise cytostatic signals to pro-proliferative signals. Extensive in vitro and in vivo characterization of GBM TSCs by using differentiation-inducing agents such as retinoic acid demonstrated that these TSCs differentiate efficiently and stop proliferation. We have demonstrated that retinoic acid treatment achieve cyctostatic effect by decreasing the proportion of CD133 positive cells, a putative marker for brain TSCs, from tumors and by inducing differentiation into astroglial lineage. Interestingly, a subset of GBM TICs pretreated with radiation and chemotherapeutic agents in vivo, do not reveal significant retinoic acid-mediated differentiation. Elucidation of underlying molecular mechanism will provide important clue for predicting sensitivity of differentiation therapeutic approach. Characterization of TSCs in aspect of differentiation-inducing agents further revealed the limitation of traditional glioma cell lines grown in serum. For example, retinoic acid treatment and CNTF exposure potently induce differentiation in most GBM 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 TSGs and oncogenes in glioblastoma TSCs identified from bioinformatics approach 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.