Malignant gliomas are aggressive brain tumors with limited therapeutic options, and improvements in treatment require a deeper molecular understanding of this disease. As in other cancers, recent ...studies have identified highly tumorigenic subpopulations within malignant gliomas, known generally as cancer stem cells. Here, we demonstrate that glioma stem cells (GSCs) produce nitric oxide via elevated nitric oxide synthase-2 (NOS2) expression. GSCs depend on NOS2 activity for growth and tumorigenicity, distinguishing them from non-GSCs and normal neural progenitors. Gene expression profiling identified many
NOS2-regulated genes, including the cell-cycle inhibitor cell division autoantigen-1 (
CDA1). Further, high
NOS2 expression correlates with decreased survival in human glioma patients, and NOS2 inhibition slows glioma growth in a murine intracranial model. These data provide insight into how GSCs are mechanistically distinct from their less tumorigenic counterparts and suggest that NOS2 inhibition may be an efficacious approach to treating this devastating disease.
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► Glioma stem cell proliferation is supported by nitric oxide synthase-2 (NOS2) ► Nonstem glioma cells and normal neural stem cells are not dependent on NOS2 ► NOS2 affects glioma stem cell gene expression, including cell-cycle inhibitor CDA1 ► Pharmacological inhibition of NOS2 attenuates glioma growth in vivo
Controlling reactive oxygen species (ROS) at sustainable levels can drive multiple facets of tumor biology, including within the cancer stem cell (CSC) population. Tight regulation of ROS is one key ...component in CSCs that drives disease recurrence, cell signaling, and therapeutic resistance. While ROS are well-appreciated to need oxygen and are a product of oxidative phosphorylation, there are also important roles for ROS under hypoxia. As hypoxia promotes and sustains major stemness pathways, further consideration of ROS impacts on CSCs in the tumor microenvironment is important. Furthermore, glycolytic shifts that occur in cancer and may be promoted by hypoxia are associated with multiple mechanisms to mitigate oxidative stress. This altered metabolism provides survival advantages that sustain malignant features, such as proliferation and self-renewal, while producing the necessary antioxidants that reduce damage from oxidative stress. Finally, disease recurrence is believed to be attributed to therapy resistant CSCs which can be quiescent and have changes in redox status. Effective DNA damage response pathways and/or a slow-cycling state can protect CSCs from the genomic catastrophe induced by irradiation and genotoxic agents. This review will explore the delicate, yet complex, relationship between ROS and its pleiotropic role in modulating the CSC.
Glioblastomas are highly lethal cancers that contain cellular hierarchies with self-renewing cancer stem cells that can propagate tumors in secondary transplant assays. The potential significance of ...cancer stem cells in cancer biology has been demonstrated by studies showing contributions to therapeutic resistance, angiogenesis, and tumor dispersal. We recently reported that physiologic oxygen levels differentially induce hypoxia inducible factor-2α (HIF2α) levels in cancer stem cells. HIF1α functioned in proliferation and survival of all cancer cells but also was activated in normal neural progenitors suggesting a potentially restricted therapeutic index while HIF2α was essential in only in cancer stem cells and was not expressed by normal neural progenitors demonstrating HIF2α is a cancer stem cell specific target. We now extend these studies to examine the role of hypoxia in regulating tumor cell plasticity. We find that hypoxia promotes the self-renewal capability of the stem and non-stem population as well as promoting a more stem-like phenotype in the non-stem population with increased neurosphere formation as well as upregulation of important stem cell factors, such as OCT4, NANOG, and c-MYC. The importance of HIF2α was further supported as forced expression of non-degradable HIF2α induced a cancer stem cell marker and augmented the tumorigenic potential of the non-stem population. This novel finding may indicate a specific role of HIF2α in promoting glioma tumorigenesis. The unexpected plasticity of the non-stem glioma population and the stem-like phenotype emphasizes the importance of developing therapeutic strategies targeting the microenvironmental influence on the tumor in addition to cancer stem cells.
Like all cancers, brain tumors require a continuous source of energy and molecular resources for new cell production. In normal brain, glucose is an essential neuronal fuel, but the blood-brain ...barrier limits its delivery. We now report that nutrient restriction contributes to tumor progression by enriching for brain tumor initiating cells (BTICs) owing to preferential BTIC survival and to adaptation of non-BTICs through acquisition of BTIC features. BTICs outcompete for glucose uptake by co-opting the high affinity neuronal glucose transporter, type 3 (Glut3, SLC2A3). BTICs preferentially express Glut3, and targeting Glut3 inhibits BTIC growth and tumorigenic potential. Glut3, but not Glut1, correlates with poor survival in brain tumors and other cancers; thus, tumor initiating cells may extract nutrients with high affinity. As altered metabolism represents a cancer hallmark, metabolic reprogramming may maintain the tumor hierarchy and portend poor prognosis.
Glioblastomas are lethal cancers characterized by florid angiogenesis promoted in part by glioma stem cells (GSCs). Because hypoxia regulates angiogenesis, we examined hypoxic responses in GSCs. We ...now demonstrate that hypoxia-inducible factor HIF2α and multiple HIF-regulated genes are preferentially expressed in GSCs in comparison to non-stem tumor cells and normal neural progenitors. In tumor specimens, HIF2α colocalizes with cancer stem cell markers. Targeting HIFs in GSCs inhibits self-renewal, proliferation, and survival in vitro, and attenuates tumor initiation potential of GSCs in vivo. Analysis of a molecular database reveals that
HIF2A expression correlates with poor glioma patient survival. Our results demonstrate that GSCs differentially respond to hypoxia with distinct HIF induction patterns, and HIF2α might represent a promising target for antiglioblastoma therapies.
Glioblastomas display cellular hierarchies containing tumor-propagating glioblastoma stem cells (GSCs). STAT3 is a critical signaling node in GSC maintenance but molecular mechanisms underlying STAT3 ...activation in GSCs are poorly defined. Here we demonstrate that the bone marrow X-linked (BMX) nonreceptor tyrosine kinase activates STAT3 signaling to maintain self-renewal and tumorigenic potential of GSCs. BMX is differentially expressed in GSCs relative to nonstem cancer cells and neural progenitors. BMX knockdown potently inhibited STAT3 activation, expression of GSC transcription factors, and growth of GSC-derived intracranial tumors. Constitutively active STAT3 rescued the effects of BMX downregulation, supporting that BMX signals through STAT3 in GSCs. These data demonstrate that BMX represents a GSC therapeutic target and reinforces the importance of STAT3 signaling in stem-like cancer phenotypes.
► BMX is preferentially expressed in glioblastoma stem cells ► BMX activates STAT3 signaling specifically in glioblastoma stem cells ► BMX controls the expression of key glioblastoma stem cell transcription factors ► Targeting BMX disrupts tumorigenic potential of glioblastoma stem cells
Many solid cancers display cellular hierarchies with self-renewing, tumorigenic stemlike cells, or cancer-initiating cells (CICs) at the apex. Whereas CICs often exhibit relative resistance to ...conventional cancer therapies, they also receive critical maintenance cues from supportive stromal elements that also respond to cytotoxic therapies. To interrogate the interplay between chemotherapy and CICs, we investigated cellular heterogeneity in human colorectal cancers. Colorectal CICs were resistant to conventional chemotherapy in cell-autonomous assays, but CIC chemoresistance was also increased by cancer-associated fibroblasts (CAFs). Comparative analysis of matched colorectal cancer specimens from patients before and after cytotoxic treatment revealed a significant increase in CAFs. Chemotherapy-treated human CAFs promoted CIC self-renewal and in vivo tumor growth associated with increased secretion of specific cytokines and chemokines, including interleukin-17A (IL-17A). Exogenous IL-17A increased CIC self-renewal and invasion, and targeting IL-17A signaling impaired CIC growth. Notably, IL-17A was overexpressed by colorectal CAFs in response to chemotherapy with expression validated directly in patient-derived specimens without culture. These data suggest that chemotherapy induces remodeling of the tumor microenvironment to support the tumor cellular hierarchy through secreted factors. Incorporating simultaneous disruption of CIC mechanisms and interplay with the tumor microenvironment could optimize therapeutic targeting of cancer.
The Human Protein Atlas (HPA) aims to map human proteins via multiple technologies including imaging, proteomics and transcriptomics. Access of the HPA data is mainly via web-based interface allowing ...views of individual proteins, which may not be optimal for data analysis of a gene set, or automatic retrieval of original images.
HPAanalyze is an R package for retrieving and performing exploratory analysis of data from HPA. HPAanalyze provides functionality for importing data tables and xml files from HPA, exporting and visualizing data, as well as downloading all staining images of interest. The package is free, open source, and available via Bioconductor and GitHub. We provide examples of the use of HPAanalyze to investigate proteins altered in the deadly brain tumor glioblastoma. For example, we confirm Epidermal Growth Factor Receptor elevation and Phosphatase and Tensin Homolog loss and suggest the importance of the GTP Cyclohydrolase I/Tetrahydrobiopterin pathway. Additionally, we provide an interactive website for non-programmers to explore and visualize data without the use of R.
HPAanalyze integrates into the R workflow with the tidyverse framework, and it can be used in combination with Bioconductor packages for easy analysis of HPA data.
Radiotherapy represents the most effective nonsurgical treatments for gliomas. However, gliomas are highly radioresistant and recurrence is nearly universal. Results from our laboratory and other ...groups suggest that cancer stem cells contribute to radioresistance in gliomas and breast cancers. The Notch pathway is critically implicated in stem cell fate determination and cancer. In this study, we show that inhibition of Notch pathway with γ‐secretase inhibitors (GSIs) renders the glioma stem cells more sensitive to radiation at clinically relevant doses. GSIs enhance radiation‐induced cell death and impair clonogenic survival of glioma stem cells but not non‐stem glioma cells. Expression of the constitutively active intracellular domains of Notch1 or Notch2 protect glioma stem cells against radiation. Notch inhibition with GSIs does not alter the DNA damage response of glioma stem cells after radiation but rather reduces Akt activity and Mcl‐1 levels. Finally, knockdown of Notch1 or Notch2 sensitizes glioma stem cells to radiation and impairs xenograft tumor formation. Taken together, our results suggest a critical role of Notch signaling to regulate radioresistance of glioma stem cells. Inhibition of Notch signaling holds promise to improve the efficiency of current radiotherapy in glioma treatment. STEM CELLS 2010;28:17–28
Malignant gliomas rank among the most lethal cancers. Gliomas display a striking cellular heterogeneity with a hierarchy of differentiation states. Recent studies support the existence of cancer stem ...cells in gliomas that are functionally defined by their capacity for extensive self-renewal and formation of secondary tumors that phenocopy the original tumors. As the c-Myc oncoprotein has recognized roles in normal stem cell biology, we hypothesized that c-Myc may contribute to cancer stem cell biology as these cells share characteristics with normal stem cells.
Based on previous methods that we and others have employed, tumor cell populations were enriched or depleted for cancer stem cells using the stem cell marker CD133 (Prominin-1). We characterized c-Myc expression in matched tumor cell populations using real time PCR, immunoblotting, immunofluorescence and flow cytometry. Here we report that c-Myc is highly expressed in glioma cancer stem cells relative to non-stem glioma cells. To interrogate the significance of c-Myc expression in glioma cancer stem cells, we targeted its expression using lentivirally transduced short hairpin RNA (shRNA). Knockdown of c-Myc in glioma cancer stem cells reduced proliferation with concomitant cell cycle arrest in the G(0)/G(1) phase and increased apoptosis. Non-stem glioma cells displayed limited dependence on c-Myc expression for survival and proliferation. Further, glioma cancer stem cells with decreased c-Myc levels failed to form neurospheres in vitro or tumors when xenotransplanted into the brains of immunocompromised mice.
These findings support a central role of c-Myc in regulating proliferation and survival of glioma cancer stem cells. Targeting core stem cell pathways may offer improved therapeutic approaches for advanced cancers.