Mechanisms of Hedgehog signaling in glioblastoma

项目总结 胶质母细胞瘤是一种致命的肿瘤，约占所有恶性原发脑癌的一半。 胶质母细胞瘤干细胞存在于受肿瘤影响的可互换的细胞状态中。 微环境，关于细胞机制的信息很少。 胶质母细胞瘤细胞和肿瘤间质之间的通讯。引导基因的Hedgehog途径 对发育和成体干细胞动态平衡至关重要的表达程序对生存至关重要 胶质母细胞瘤干细胞。矛盾的是，一项临床试验表明，刺猬的药物抑制 途径加速胶质母细胞瘤的生长。这些临床数据提供了一个机会来重新评估 Hedgehog通路在肿瘤中的作用，并提示Hedgehog信号在胶质母细胞瘤干细胞中的作用 细胞仍有待阐明。这项建议侧重于了解如何抑制Hedgehog途径 在肿瘤、细胞和分子水平上驱动胶质母细胞瘤。为了做到这一点，我们的提案结合了一部小说 胶质母细胞瘤的星形细胞器官模型，便于肿瘤内细胞相互作用的实时成像 微环境。当与CRISPR干扰、药理学和小鼠同基因和 患者来源的胶质母细胞瘤异种移植模型，我们创新的器官系统将促进以前 理解胶质母细胞瘤生物学的实验是不可能的。根据我们初步的有机化合物，在体内，和 在本申请中提供的单细胞RNA测序数据中，我们假设癌细胞-外源性 肿瘤微环境中的Hedgehog信号通过以下途径抑制胶质母细胞瘤干细胞的更新和侵袭 诱导分化因子，重塑细胞外基质。我们将通过定义以下内容来检验这一假设 Hedgehog信号如何通过胶质母细胞瘤微环境进行转导；确定癌细胞 外源性Hedgehog信号通过诱导骨形态发生蛋白信号抑制胶质母细胞瘤干细胞； 并确定Hedgehog信号是否通过诱导调控基因来抑制胶质母细胞瘤细胞的侵袭 通过细胞外基质进行迁移。这种方法是基于这样一个前提，即通过研究 Hedgehog信号在胶质母细胞瘤中的机制，我们将发现对胶质母细胞瘤细胞如何 与肿瘤微环境沟通。事实上，我们对相互作用的了解令人惊讶地少之又少 胶质母细胞瘤细胞和肿瘤微环境之间的关系几乎不会影响癌症的进展 关于星形胶质细胞如何影响胶质母细胞瘤的侵袭，星形胶质细胞是成人大脑中的主要细胞类型 或肿瘤的发生。虽然这项建议的目的是广泛地提高我们对 Hedgehog信号在胶质母细胞瘤中的机制，本研究的长期目标是了解这些 过程足够好，可以考虑有针对性的治疗策略，以提高胶质母细胞瘤的存活率。因此， 这项工作将解释为什么Hedgehog通路抑制意外地加速胶质母细胞瘤的生长，阐明 靶向机制，并为胶质母细胞瘤患者的新临床试验提供临床前基础。

Project summary Glioblastomas are deadly tumors and account for approximately half of all malignant primary brain cancers. Glioblastoma stem cells exist in interchangeable cellular states that are influenced by the tumor microenvironment, and there is a paucity of information concerning the cellular mechanisms responsible for communicating between glioblastoma cells and the tumor stroma. The Hedgehog pathway, which directs gene expression programs that are essential for development and adult stem cell homeostasis, is critical for survival of glioblastoma stem cells. Paradoxically, a clinical trial suggested that pharmacologic inhibition of the Hedgehog pathway accelerates glioblastoma growth. These clinical data provide an opportunity to re-evaluate how the Hedgehog pathway functions in cancer, and suggest that the role of Hedgehog signaling in glioblastoma stem cells remains to be elucidated. This proposal focuses on understanding how inhibition of the Hedgehog pathway drives glioblastoma at the tumor, cellular, and molecular scale. To do so, our proposal incorporates a novel astrocyte organoid model of glioblastoma that facilitates live imaging of cellular interactions within the tumor microenvironment. When integrated with CRISPR interference, pharmacology, and mouse syngeneic and patient derived xenograft models of glioblastoma, our innovative organoid system will facilitate previously impossible experiments to understand glioblastoma biology. Based on our preliminary organoid, in vivo, and single cell RNA sequencing data presented in this application, we hypothesize that cancer cell-extrinsic Hedgehog signaling in the tumor microenvironment restrains glioblastoma stem cell-renewal and invasion by inducing differentiation factors and remodeling the extracellular matrix. We will test this hypothesis by defining how Hedgehog signals are transduced through the glioblastoma microenvironment; determining if cancer-cell extrinsic Hedgehog signaling restrains glioblastoma stem cells by inducing bone morphogenic protein signaling; and determining if Hedgehog signaling restrains glioblastoma cell invasion by inducing genes that regulate migration through the extracellular matrix. This approached is based on the premise that by studying the mechanisms of Hedgehog signaling in glioblastoma, we will discover new insights into how glioblastoma cells communicate with the tumor microenvironment. Indeed, we know surprisingly little about how interactions between glioblastoma cells and the tumor microenvironment influence cancer progression, and almost nothing about how astrocytes, which comprise the majority of cell types in the adult brain, influence glioblastoma invasion or tumorigenesis. Though the objective of this proposal is to broadly improve our understanding of the mechanisms of Hedgehog signaling in glioblastoma, a long-term goal of this research is to understand these processes well enough to consider targeted therapeutic strategies to improve survival from glioblastoma. Thus, this work will explain why Hedgehog pathway inhibition unexpectedly accelerates glioblastoma growth, elucidate targetable mechanisms, and provide the preclinical basis for new clinical trials in glioblastoma patients.