Exploring the Therapeutic Potential of Stem Cell Biology in Gliomas

探索干细胞生物学在神经胶质瘤中的治疗潜力

• 批准号: 8157495
• 负责人: Howard Fine
• 金额: $ 71.24万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8157495
• 关键词: Glioma; Therapeutic; stem cell biology

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.

对TSC中失调分化途径的分子理解：干细胞自我更新和分化之间的微妙平衡由对正常组织稳态至关重要的各种细胞内在和外在因素控制。尽管TSC和正常干细胞之间存在广泛的表型和功能相似性，但TSC的分化潜力并不完全正常。阐明在正常干细胞和TSCs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。我们已经确定了一套失调的分化途径，在TSC来自人类原发性胶质母细胞瘤。我们证明，骨形态发生蛋白（BMP）介导的和睫状神经营养因子（CNTF）介导的Janus激酶/信号转导和转录激活因子（Jak/STAT）途径引起不同的生物学后果相比，正常的神经干细胞（NSC）在成人脑来源的TSCs。与非常早期的胚胎NSC一样，0308 TSCs响应BMP增殖，并在CNTF激发后引起边缘STAT 3活化。然而，与发育后期的正常神经干细胞不同，0308细胞不表达BMPR 1B，后者继发于Zeste增强子同源物2（EZH 2）依赖性BMPR 1B启动子超甲基化。通过转基因表达或启动子去甲基化在0308 TIC中强制表达BMPR 1B完全恢复其分化能力，并不仅通过BMP介导的途径而且通过CNTF介导的Jak/STAT活化诱导其致瘤性的丧失。对54例原发性人类胶质母细胞瘤的调查显示，大约20%的肿瘤抑制了与启动子高甲基化相关的BMPR 1B表达。总之，这些数据表明，在胶质母细胞瘤TSC的子集中BMP发育分化途径的失调不仅通过使TIC对正常分化线索脱敏，而且还通过将其他细胞抑制信号转化为促增殖信号而促成其致瘤表型。通过使用分化诱导剂如视黄酸对GBM TSC进行广泛的体外和体内表征，证明这些TSC有效分化并停止增殖。我们已经证明，视黄酸处理通过降低来自肿瘤的CD 133阳性细胞（脑TSC的推定标记物）的比例和通过诱导分化成星形胶质细胞谱系来实现细胞生长抑制作用。有趣的是，在体内用放射和化疗剂预处理的GBM TIC的子集没有显示出显著的视黄酸介导的分化。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胶质瘤细胞系在血清中生长的局限性。例如，视黄酸处理和CNTF暴露有效地诱导大多数GBM 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TSG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。