Suppression of Glioblastoma Stem Cells by Kruppel-Like Factor 9

描述(申请人提供)：胶质母细胞瘤干细胞(或肿瘤干细胞，CSCs)保持多系分化的能力，并有效地增殖肿瘤异种移植，准确地概括了临床胶质母细胞瘤的复杂组织病理学。识别和靶向调节GBM-CSC表型的分子机制对于最终耗尽其CSCs中的肿瘤具有很大的希望，目前认为CSCs在治疗耐药和肿瘤复发中起着重要作用。Kruppel-like factor9(KLF9)是一种知之甚少的转录因子，以前与癌症或干细胞没有明显的联系。我们最近发现KLF9能有效地诱导胶质母细胞瘤肿瘤干细胞(GBM-CSC)分化，抑制GBM-CSC的自我更新，并抑制GBM-CSC来源的肿瘤移植瘤的生长。KLF9的这些肿瘤抑制作用被发现部分是直接抑制Notch1转录的结果。KLF9可能调节一个广泛的转录网络，因为它识别转录启动子中常见的GC-GT BTE位点。这一建议是基于这样的普遍假设，即KLF9调节转录网络，抑制GBM-CSCs的致癌表型和治疗耐药。这项建议的总体目标是确定KLF9的S治疗和肿瘤抑制作用及其转录靶点，并开发一种直接细胞穿透形式的KLF9用于潜在的临床翻译。目的#1将利用富含GBM-CSCs的人GBM球体形成细胞及其原位移植瘤来确定KLF9如何调节GBM-CSC对放射和替莫唑胺化疗的反应，这两种细胞毒方法目前是GBM治疗的主要方式。目的#2利用芯片、基因表达阵列和广泛的生物信息学分析，鉴定在GBM-CSCs中受KLF9调控的转录网络。目的#3将使用体内RCAS/TV-a系统来确定KLF9是否调节多潜能神经干细胞/祖细胞的体内转化和随后的胶质瘤形成。目的#4利用羧基末端多聚精氨酸(KLF9-11R)修饰的KLF9制备直接穿透细胞的KLF9蛋白，用于靶向GBM-CSC。我们发现KLF9在人GBM-CSCs中具有分化、抑制肿瘤和辐射增敏作用，这一发现具有广泛的生物学和临床可翻译意义。这些实验的积极结果将显著影响识别和了解肿瘤干细胞的分子调控因子，并最终控制它们的生长、命运和脑癌治疗的化疗/放射敏感性的目标。 公共卫生相关性：肿瘤起始的肿瘤干细胞(CSCs)在胶质母细胞瘤(GBM)和其他实体恶性肿瘤的治疗抵抗和复发中起着重要作用。我们最近发现Kruppel样因子9转录因子在GBM-CSCs中具有强大的分化和肿瘤抑制功能。该方案的目的是确定KLF9在GBM-CSCs中的肿瘤抑制/治疗增敏作用和转录靶点，并开发一种直接细胞穿透形式的KLF9用于潜在的治疗性翻译。

DESCRIPTION (provided by applicant): Glioblastoma stem-like cells (or cancer stem cells, CSCs) maintain a capacity for multi-lineage differentiation and efficiently propagate tumor xenografts that accurately recapitulate the complex histopathology of clinical glioblastoma. Identifying and targeting the molecular mechanisms that regulate the GBM-CSC phenotype holds great promise for ultimately depleting tumors of their CSCs that are currently believed to have a major role in therapeutic resistance and tumor recurrence. Kruppel-like factor 9 (KLF9) is a poorly understood transcription factor with no significant previous link to cancer or stem cells. We recently found that KLF9 potently induces glioblastoma cancer stem cell (GBM-CSC) differentiation, inhibits GBM-CSC self-renewal, and suppresses the growth of GBM-CSC derived tumor xenografts. These tumor suppressing effects of KLF9 were found to result in part from the direct repression of Notch1 transcription. KLF9 is likely to modulate an extensive transcriptional network since it recognizes GC-GT BTE sites that are common in transcriptional promoters. This proposal is based on the general hypothesis that KLF9 regulates a transcriptional network that suppresses the oncogenic phenotype and therapeutic resistance of GBM-CSCs. The general goals of this proposal are to determine KLF9's therapeutic and tumor suppressing effects, its transcriptional targets, and to develop a direct cell-penetrating form of KLF9 for potential clinical translation. Aim #1 will use human GBM sphere-forming cells enriched in GBM-CSCs and their orthotopic tumor xenografts to determine how KLF9 modulates GBM-CSC responses to radiation and temozolomide chemotherapy, cytotoxic modalities that are currently the mainstay of GBM therapy. Aim #2 will identify the transcriptional networks regulated by KLF9 in GBM-CSCs using ChIP-Chip, gene expression array, and extensive bioinformatics analyses. Aim #3 will use the in vivo RCAS/tv-a system to determine if KLF9 modulates in vivo transformation of multipotent neural stem/progenitor cells and subsequent gliomagenesis. Aim #4 will use KLF9 modified with carboxy-terminal poly-arginines (KLF9-11R) to develop direct cell- penetrating KLF9 protein for targeting GBM-CSCs. Our discovery that KLF9 induction is differentiating, tumor suppressing, and radiation sensitizing in human GBM-CSCs is novel with broad biological and clinical translatable implications. Positive results from these experiments will significantly impact the goals to identify and understand molecular regulators of neoplastic stem cells and to ultimately control their growth, fate, and chemo/radiation sensitivity for brain cancer therapy. PUBLIC HEALTH RELEVANCE: Tumor-initiating cancer stem-like cells (CSCs) play a prominent role in the therapeutic resistance and recurrence of glioblastoma (GBM) and other solid malignancies. We recently found that the Kruppel-like factor 9 transcription factor has potent differentiating and tumor suppressing functions in GBM-CSCs. The goals of this proposal are to determine tumor suppressing/treatment sensitizing effects and transcriptional targets of KLF9 in GBM-CSCs and to develop a direct cell penetrating form of KLF9 for potential therapeutic translation.