The HIF1?/JMY pathway promotes glioblastoma stem-like cell invasiveness after irradiation.
ABSTRACT: Human glioblastoma (GBM) is the most common primary malignant brain tumor. A minor subpopulation of cancer cells, known as glioma stem-like cells (GSCs), are thought to play a major role in tumor relapse due to their stem cell-like properties, their high resistance to conventional treatments and their high invasion capacity. We show that ionizing radiation specifically enhances the motility and invasiveness of human GSCs through the stabilization and nuclear accumulation of the hypoxia-inducible factor 1? (HIF1?), which in turn transcriptionally activates the Junction-mediating and regulatory protein (JMY). Finally, JMY accumulates in the cytoplasm where it stimulates GSC migration via its actin nucleation-promoting activity. Targeting JMY could thus open the way to the development of new therapeutic strategies to improve the efficacy of radiotherapy and prevent glioma recurrence.
Project description:Chemotherapy plays a significant role in glioma treatment; however, it has limited effectiveness in extending the life expectancies of glioma patients. Traditional studies have attributed this lack of efficacy to glioma stem cells (GSCs) and their high resistance to chemotherapy, and hypoxia worsens this issue. In contrast, hyperoxia effectively alleviates hypoxia in glioma and sensitizes glioma cells to chemotherapy. In a summary of traditional studies, the majority of researchers overlooked the influence of hypoxia on differentiated cells because they only focused on the maintenance of GSCs stemness, which thus resulted in chemoresistance. Because of this background, we hypothesized that GSCs may be induced through dedifferentiation under hypoxic conditions, and hypoxia maintains GSCs stemness, which thus leads to resistance to chemotherapy. In contrast, hyperoxia inhibits the dedifferentiation process and promotes GSCs differentiation, which increases the sensitization of glioma cells to chemotherapy. Hypoxia-inducible factor-1? (HIF1?) contributes substantially to the stemness maintenance of GSCs and resistance of glioma to chemotherapy; thus, we investigated whether HIF1? regulates the resistance or sensitization of glioma cells to chemotherapy in different oxygen levels. It highlights a novel viewpoint on glioma chemosensitivity from the transformation between dedifferentiation and differentiation in different oxygen levels.
Project description:Tumor radioresistance leading to local therapy failure remains a major obstacle for successful treatment of high-grade glioma. We hypothesized that distinct radiobiological features of particle therapy with carbon ions may circumvent glioma radioresistance. We demonstrate that carbon irradiation (CIR) efficiently eradicates radioresistant patient-derived glioma stem cells (GSCs), leading to growth inhibition and prolonged survival. The impact of CIR at the tumor-stroma interface was further investigated in 2 syngeneic mouse and 2 orthotopic GSC xenograft models. Intriguingly, tumor regressions and long-term local controls were observed at doses greater than or equal to 15-Gy CIR. Fractionated CIR further prolonged survival. The enhanced relative biological effectiveness of CIR in vivo was attributed to its potent antiangiogenic effects and eradication of radioresistant hypoxic tumor cells. Blockade of the HIF1-?/stromal cell-derived factor 1/CXCR4 axis by CIR reduced the recruitment of microglia and myeloid-derived suppressor cells (CD11b+Gr1+). Consequently, CIR abrogated M2-like immune polarization and enhanced the influx of CD8+ cells, generating an immunopermissive niche. We report that radiotherapy with carbon ions could surmount several central glioma resistance mechanisms by eradicating hypoxic and stem cell-like tumor cells, as well as modulating the glioma niche toward an antiangiogenic and less immunosuppressive state. Conclusively, potentially novel rationales for CIR in conquering glioma radioresistance are provided.
Project description:"Glioma Stem Cells" (GSCs) are known to play a role in glioblastoma (GBM) recurrence. Homologous recombination (HR) defects and cell cycle checkpoint abnormalities can contribute concurrently to the radioresistance of GSCs. DNA repair protein RAD51 homolog 1 (RAD51) is a crucial protein for HR and its inhibition has been shown to sensitize GSCs to irradiation. The aim of this study was to examine the consequences of ionizing radiation (IR) for cell cycle progression in GSCs. In addition, we intended to assess the potential effect of RAD51 inhibition on cell cycle progression. Five radiosensitive GSC lines and five GSC lines that were previously characterized as radioresistant were exposed to 4Gy IR, and cell cycle analysis was done by fluorescence-activated cell sorting (FACS) at 24, 48, 72, and 96 h with or without RAD51 inhibitor. Following 4Gy IR, all GSC lines presented a significant increase in G2 phase at 24 h, which was maintained over 72 h. In the presence of RAD51 inhibitor, radioresistant GSCs showed delayed G2 arrest post-irradiation for up to 48 h. This study demonstrates that all GSCs can promote G2 arrest in response to radiation-induced DNA damage. However, following RAD51 inhibition, the cell cycle checkpoint response differed. This study contributes to the characterization of the radioresistance mechanisms of GSCs, thereby supporting the rationale of targeting RAD51-dependent repair pathways in view of radiosensitizing GSCs.
Project description:Glioblastoma multiforme (GBM) is one of the most aggressive human malignancies with a poor patient prognosis. Ionizing radiation either alone or adjuvant after surgery is part of standard treatment for GBM but remains primarily noncurative. The mechanisms underlying tumor radioresistance are manifold and, in part, accredited to a special subpopulation of tumorigenic cells. The so-called glioma stem cells (GSC) are bestowed with the exclusive ability to self-renew and repopulate the tumor and have been reported to be less sensitive to radiation-induced damage through preferential activation of DNA damage checkpoint responses and increased capacity for DNA damage repair. During each fraction of radiation, non-stem cancer cells (CC) die and GSCs become enriched and potentially increase in number, which may lead to accelerated repopulation. We propose a cellular Potts model that simulates the kinetics of GSCs and CCs in glioblastoma growth and radiation response. We parameterize and validate this model with experimental data of the U87-MG human glioblastoma cell line. Simulations are conducted to estimate GSC symmetric and asymmetric division rates and explore potential mechanisms for increased GSC fractions after irradiation. Simulations reveal that in addition to their higher radioresistance, a shift from asymmetric to symmetric division or a fast cycle of GSCs following fractionated radiation treatment is required to yield results that match experimental observations. We hypothesize a constitutive activation of stem cell division kinetics signaling pathways during fractionated treatment, which contributes to the frequently observed accelerated repopulation after therapeutic irradiation.
Project description:The standard treatment for Glioblastoma multiforme (GBM) is surgical resection and subsequent radiotherapy and chemotherapy. Surgical resection of GBM is typically restricted because of its invasive growth, which results in residual tumor cells including glioma stem cells (GSCs) and differentiated cells. Recurrence has been previously thought to occur as a result of these GSCs, and hypoxic microenvironment maintains the GSCs stemness also plays an important role. Summarizing traditional studies and we find many researchers ignored the influence of hypoxia on differentiated cells. We hypothesized that the residual differentiated cells may be dedifferentiated to GSC-like cells under hypoxia and play a crucial role in the rapid, high-frequency recurrence of GBM. Therefore, isolated CD133-CD15-NESTIN- cells were prepared as single-cell culture and treated with hypoxia. More than 95% of the surviving single differentiated CD133-CD15-NESTIN- cell dedifferentiated into tumorigenic CD133+CD15+NESTIN+ GSCs, and this process was regulated by hypoxia inducible factor-1?. Moreover, the serum also played an important role in this dedifferentiation. These findings challenge the traditional glioma cell heterogeneity model, cell division model and glioma malignancy development model. Our study also highlights the mechanism of GBM recurrence and the importance of anti-hypoxia therapy. In addition to GSCs, residual differentiated tumor cells also substantially contribute to treatment resistance and the rapid, high recurrence of GBM.
Project description:Glioblastoma is the most common malignant brain tumour in adults. The failure of current therapies can be ascribed to glioma stem cells (GSCs), which can rapidly repopulate the tumour following the initial treatment. The study of histone deacetylase inhibitors, such as valproic acid (VPA), is becoming an attractive field in cancer research. However, the exact mechanisms underlying its anti-cancer effect remain to be elucidated due to its pleiotropic effects on several cell-signalling pathways. Ingenuity Pathway Analysis (IPA) bioinformatics analysis was performed on genome-wide data regarding GSCs methylome to identify the signalling pathways mainly affected by methylation changes induced by VPA. Real time PCR and luciferase reporter assay were used to better investigate VPA effects on Wnt/?-catenin signalling pathway. VPA effect on GSC proliferation was evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) and Trypan blue assays. Finally, VPA impact on GSC motility was demonstrated by Boyden chamber assay and further confirmed evaluating the expression levels or localisation, through western blot or immunofluorescence, of Twist1, Snail1, E-Cadherin and N-Cadherin. The bioinformatics analyses performed on GSCs methylome highlighted that Wnt/?-catenin signalling was affected by the methylation changes induced by VPA, which could influence its activation status. In particular, we pointed out a general activation of this pathway after VPA exposure, which was accompanied by an inhibitory potential on GSCs proliferation. Finally, we also proved VPA's ability to inhibit GSCs invasion through Snail1 and Twist1 downregulation and E-Cadherin relocalisation. VPA treatment may represent a new, interesting therapeutic approach to affect GSC proliferation and motility, but further investigations are certainly needed.
Project description:Plastic phenotype convention between glioma stem cells (GSCs) and non-stem tumor cells (NSTCs) significantly fuels glioblastoma heterogeneity that causes therapeutic failure. Recent progressions indicate that glucose metabolic reprogramming could drive cell fates. However, the metabolic pattern of GSCs and NSTCs and its association with tumor cell phenotypes remain largely unknown. Here we found that GSCs were more glycolytic than NSTCs, and voltage-dependent anion channel 2 (VDAC2), a mitochondrial membrane protein, was critical for metabolic switching between GSCs and NSTCs to affect their phenotypes. VDAC2 was highly expressed in NSTCs relative to GSCs and coupled a glycolytic rate-limiting enzyme platelet-type of phosphofructokinase (PFKP) on mitochondrion to inhibit PFKP-mediated glycolysis required for GSC maintenance. Disruption of VDAC2 induced dedifferentiation of NSTCs to acquire GSC features, including the enhanced self-renewal, preferential expression of GSC markers, and increased tumorigenicity. Inversely, enforced expression ofVDAC2 impaired the self-renewal and highly tumorigenic properties of GSCs. PFK inhibitor clotrimazole compromised the effect of VDAC2 disruption on glycolytic reprogramming and GSC phenotypic transition. Clinically, VDAC2 expression inversely correlated with glioma grades (Immunohistochemical staining scores of VDAC2 were 4.7?±?2.8, 3.2?±?1.9, and 1.9?±?1.9 for grade II, grade III, and IV, respectively, p?<?0.05 for all) and the patients with high expression of VDAC2 had longer overall survival than those with low expression of VDAC2 (p?=?0.0008). In conclusion, we demonstrate that VDAC2 is a new glycolytic regulator controlling the phenotype transition between glioma stem cells and non-stem cells and may serves as a new prognostic indicator and a potential therapeutic target for glioma patients.
Project description:Glioblastoma is one of the most common neoplasms in the central nervous system characterized by limited immune response and unlimited expansion capability. Cancer stem cells (GSCs), a small fraction of the tumor cells, possess a pivotal regulation capability in the tumor microenvironment with a superior proliferation ability. We aimed to reveal the interaction between glioma stem cells (GSCs) and immune cells during tumorigenesis. Single-cell sequencing data from seven surgical specimens of glioblastoma patients and patient-derived GSCs cocultured with peripheral leukocytes were used for the analysis. Cell grouping and trajectory analysis were performed using Seurat and Monocle 3 packages in R software. The gene set of Cancer Genome Anatomy Project was used to define different cell types. Cells with the ability of proliferation and differentiation in glioblastoma tissue were defined as GSCs, which had a similar expression pattern to that in the GSCs in vitro. Astrocytes in glioblastoma were mainly derived from differentiated GSCs, while oligodendrocytes were most likely to be derived from different precursor cells. No remarkable evolutionary trajectory was observed among the subgroups of T cells in glioblastoma. The immune checkpoint interaction between GSCs and immune cells was changed from stimulatory to inhibitory during tumorigenesis. The patient-derived GSCs system is an ideal model for GSC research. The above research revealed that the interaction pattern between GSC glioma stem cells and immune cells during tumorigenesis provides a theoretical basis for GSC glioma stem cell-targeted immunotherapy.
Project description:Recombinant erythropoietin (EPO) is a growth factor used in the treatment of chemotherapy-induced anemia, but recent studies suggest that EPO may accelerate cancer growth. Although several cancers express EPO receptors (EPORs), the mechanism by which EPOR promotes tumor growth remains poorly understood. Glioblastomas display a cellular hierarchy of self-renewal and tumor propagation restricted to glioma stem cells (GSCs). We find that GSCs express higher levels of EPOR than matched non-stem glioma cells. Prospective enrichment for EPOR on GSCs increased neurosphere formation, suggesting that EPOR can select for a subset of GSCs with increased self-renewal capacity. Targeting EPOR expression with lentiviral mediated short hairpin RNA (shRNA) reduced GSC growth, survival, and neurosphere formation capacity, defining a crucial role for EPOR in GSC maintenance. We further find that STAT3 is an important mediator of EPOR signals in GSCs. EPOR knockdown attenuated the basal activation of STAT3 present in GSCs, and a small molecule inhibitor of STAT3 reduced GSC growth and survival. EPOR signaling was critical for survival in vivo, as targeting EPOR expression decreased GSC tumorigenic potential. Elevated EPOR expression also associated with poor patient outcome. Thus, EPOR on GSCs promotes tumor growth and may explain the poor survival of cancer patients treated with EPO.