Behavior Of Glioma Stem Cells Research Articles

SENP1-Mediated deSUMOylation Regulates the Tumor Remodeling of Glioma Stem Cells Under Hypoxic Stress.

Objectives: This study aimed to investigate the effect of specific small ubiquitin-like modifier (SUMO) proteases 1 (SENP1)-mediated deSUMOylation on the malignant behavior of glioma stem cells (GSCs) under hypoxia conditions and evaluate the clinical value of prevention in glioma patients. Introductions: Under hypoxic conditions, upregulated hypoxia-inducible factor 1α (HIF1α) expression in GSCs activates Wnt/β-catenin signaling pathways, which provide rich nutritional support for glioblastoma (GBM). SENP1-mediated deSUMOylation stabilizes the expression of HIF1α and β-catenin, leading to the occurrence of GSCs-initiated tumorigenesis. Targeting SENP1-mediated deSUMOylation may suppress the malignancy of GSCs and disrupt GBM progression. Methods: The expression of SENP1 in different World Health Organization grades was observed by immunohistochemistry and western blot. Lentivirus-packaged SENP1shRNA downregulated the expression of SENP1 in GSCs, and the downregulated results were verified by western blotting and polymerase chain reaction. The effects of LV-SENP1shRNA on the migration and proliferation of GSCs were detected by scratch and cloning experiments. The effect of LV-SENP1shRNA on the tumor formation ability of GSCs was observed in nude mice. Immunoprecipitation clarified the mechanism of SENP1 regulating the malignant behavior of GSCs under hypoxia. The correlation between the expression level of SENP1 and the survival of glioma patients was determined by statistical analysis. Results: SENP1 expression in GSCs derived from clinical samples was upregulated in GBM. SUMOylation was observed in GSCs in vitro, and deSUMOylation, accompanied by an increase in SENP1 expression, was induced by hypoxia. SENP1 expression was downregulated in GSCs with lentivirus-mediated stable transfection, which attenuated the proliferation and differentiation of GSCs, thus diminishing tumorigenesis. Mechanistically, HIF1α induced activation of Wnt/β-catenin, which depended on SENP1-mediated deSUMOylation, promoting GSC-driven GBM growth under the hypoxia microenvironment. Conclusion: Our findings indicate that SENP1-mediated deSUMOylation as a feature of GSCs is essential for GBM maintenance, suggesting that targeting SENP1 against GSCs may effectively improve GBM therapeutic efficacy.

Deciphering the Role of Autophagy in Treatment of Resistance Mechanisms in Glioblastoma.

Autophagy is a process essential for cellular energy consumption, survival, and defense mechanisms. The role of autophagy in several types of human cancers has been explicitly explained; however, the underlying molecular mechanism of autophagy in glioblastoma remains ambiguous. Autophagy is thought to be a “double-edged sword”, and its effect on tumorigenesis varies with cell type. On the other hand, autophagy may play a significant role in the resistance mechanisms against various therapies. Therefore, it is of the utmost importance to gain insight into the molecular mechanisms deriving the autophagy-mediated therapeutic resistance and designing improved treatment strategies for glioblastoma. In this review, we discuss autophagy mechanisms, specifically its pro-survival and growth-suppressing mechanisms in glioblastomas. In addition, we try to shed some light on the autophagy-mediated activation of the cellular mechanisms supporting radioresistance and chemoresistance in glioblastoma. This review also highlights autophagy’s involvement in glioma stem cell behavior, underlining its role as a potential molecular target for therapeutic interventions.

A three-dimensional (3D) organotypic microfluidic model for glioma stem cells – Vascular interactions

Glioblastoma (GBM) is one of the deadliest forms of cancer. Despite many treatment options, prognosis of GBM remains dismal with a 5-year survival rate of 4.7%. Even then, tumors often recur after treatment. Tumor recurrence is hypothesized to be driven by glioma stem cell (GSC) populations which are highly tumorigenic, invasive, and resistant to several forms of therapy. GSCs are often concentrated around the tumor vasculature, referred to as the vascular niche, which are known to provide microenvironmental cues to maintain GSC stemness, promote invasion, and resistance to therapies. In this work, we developed a 3D organotypic microfluidic platform, integrated with hydrogel-based biomaterials, to mimic the GSC vascular niche and study the influence of endothelial cells (ECs) on patient-derived GSC behavior and identify signaling cues that mediate their invasion and phenotype. The established microvascular network enhanced GSC migration within a 3D hydrogel, promoted invasive morphology as well as maintained GSC proliferation rates and phenotype (Nestin, SOX2, CD44). Notably, we compared migration behavior to in vivo mice model and found similar invasive morphology suggesting that our microfluidic system could represent a physiologically relevant in vivo microenvironment. Moreover, we confirmed that CXCL12-CXCR4 signaling is involved in promoting GSC invasion in a 3D vascular microenvironment by utilizing a CXCR4 antagonist (AMD3100), while also demonstrating the effectiveness of the microfluidic as a drug screening assay. Our model presents a potential ex vivo platform for studying the interplay of GSCs with its surrounding microenvironment as well as development of future therapeutic strategies tailored toward disrupting key molecular pathways involved in GSC regulatory mechanisms.

Implication of connexin30 on the stemness of glioma: connexin30 reverses the malignant phenotype of glioma by modulating IGF-1R, CD133 and cMyc.

Gap-junctional intercellular communication (GJIC) plays a major role in the malignant growth of glioma. Although the mechanistic aspects of GJIC have been extensively studied, the role of connexins in the regulation of the malignant behavior of glioma stem cells (GSCs) remains unclear. In our previous studies, we have shown that connexin30 can interfere with the insulin-like growth factor 1 receptor (IGF-1R), which is known for self-renewal and pluripotency. Following our earlier in vitro observation, in this work, we aimed to study the consequence of this influence of Cx30 on IGF-1R by evaluating the marker of GSCs, CD133 and oncoprotein, cMyc. We strengthened our basis by examining human glioma samples of different grades as well as rat C6 xenografts (Cx30-transfected and -non-transfected C6 cells) along with the sphere formation assays in vitro. Investigation of stemness-related CD133 and cMyc in human samples and rat xenografts exhibited a reciprocal relationship between Cx30 and IGF-1R in the low and high grades (HG) of glioma. Cx30 was completely abolished in HG; levels of IGF-1R, CD133 and cMyc expression were positively correlated with HG. Cx30 transfection could attenuate the malignant burden of glioma in rat xenografts. Cx30 transfection also altered the tumor sphere formation of C6 glioma cells in vitro, an important property of GSCs, and there was a significant reduction of CD133 and cMyc expression by Cx30 both in vitro and in vivo. These factors indicate that dysfunction of Cx30 plays a crucial role in the prevention of the stemness of glioma, and the exploitation of this feature will help in the management of glioma.

Linc00152 promotes malignant progression of glioma stem cells by regulating miR-103a-3p/FEZF1/CDC25A pathway

BackgroundGlioma is one of the most frequent intracranial malignant tumors. LncRNAs have been identified as new modulators in the origination and progression of glioma.MethodsQuantitative real-time PCR were conducted to evaluate the expression of linc00152 and miRNA-103a-3p in glioma tissues and cells. Western blot were used to determine the expression of FEZF1 and CDC25A in glioma tissues and cells. Stable knockdown of linc00152 or over-expression of miR-103a-3p in glioma stem cells (GSCs) were established to explore the function of linc00152 and miR-103a-3p in GSCs. Further, luciferase reports were used to investigate the correlation between linc00152 and miR-103a-3p. Cell Counting Kit-8, transwell assays, and flow cytometry were used to investigate the function of linc00152 and miR-103a-3p in GSC malignant biological behaviors. ChIP assays were employed to ascertain the correlations between FEZF1 and CDC25A.ResultsLinc00152 was up-regulated in glioma tissues as well as in GSCs. Knockdown of linc00152 inhibited cell proliferation, migration and invasion, while promoted GSC apoptosis. Linc00152 regulated the malignant behavior of GSCs by binding to miR-103a-3p, which functions as a tumor suppressor. In addition, knockdown of linc00152 down-regulated forebrain embryonic zinc finger protein 1 (FEZF1), a direct target of miR-103a-3p which played an oncogenic role in GSCs. FEZF1 elevated promoter activities and up-regulated expression of the oncogenic gene cell division cycle 25A (CDC25A). CDC25A over-expression activated the PI3K/AKT pathways, which regulated the malignant behavior of GSCs.ConclusionsLinc00152/miR-103a-3p/FEZF1/CDC25A axis plays a novel role in regulating the malignant behavior of GSCs, which may be a new potential therapeutic strategy for glioma therapy.

MiR-200b inhibits CD133+ glioma cells by targeting the AKT pathway

MicroRNA-200b (miR-200b) is a tumor suppressor in multiple tumor types, including gastric cancer, breast cancer, ovarian cancer and glioma. The biological significance of a known normal and cancer stem cell marker, CD133, remains elusive. The aim of the present study was to identify the function and mechinism of miR-200b in suppressing CD133+ glioma cells. CD133+ glioma cells were sorted by flow cytometry. The expression of miR-200b, Ki67, GAP43, GFAP and CD133 were tested by reverse transcription-quantitative polymerase chain reaction. The binding of miR-200b to prominin 1 (PROM1) was certificated by luciferase reporter assay. Cell proliferation was analyzed by bromodeoxyuridine staining. The protein level of CD133, p-AKT, AKT and Notch1 was detected by western blot analysis. Analysis of glioma samples revealed that CD133 expression is negatively associated with miR-200b. PROM1, which is the gene that codes CD133, was certified to be a target of miR-200b. miR-200b expression inhibited the stemness properties and division of the CD133+ glioma cells. Our results identified a miR-200b/CD133/PI3K/Akt signaling axis, exploring the fundamental role of miR-200b and CD133 in glioma stem cell behavior.

Epigenetic targeting of glioma stem cells: Short-term and long-term treatments with valproic acid modulate DNA methylation and differentiation behavior, but not temozolomide sensitivity.

Glioblastoma (GBM) is the most aggressive tumor of the central nervous system. GBM is a fatal tumor, incurable by conventional therapies. One of the factors underlying tumor recurrence and poor long-term survival is the presence of a cancer stem-like cell population, termed glioma stem cells (GSCs), which is particularly resistant to chemotherapy and radiotherapy and supports tumor self-renewal. The aim of the present study was to evaluate the impact and difference in effects of short-term and long‑term treatments with valproic acid (VPA), a histone deacetylase inhibitor, on seven GSC lines. We investigated for the first time the changes in the genome-wide DNA methylation profile and the differentiation behavior of GSCs induced by short-term and long-term VPA treatments. Moreover, we verified VPA sensitivity after long-term VPA pretreatment and, notably, the results provide evidence of a subpopulation more resistant to further VPA treatments. Finally, since short-term VPA treatment induced a reversal of the MGMT methylation status, we aimed to sensitize GSCs to temozolomide, the drug commonly used for this tumor, using this regimen. The overall data highlighted the heterogeneous behavior of GSC lines that is representative of tumor heterogeneity in GBM. The VPA effects were variable among these cell lines in terms of pro‑differentiating ability and DNA methylation switch. Here, we attempted to identify a suitable therapy for the eradication of the stem cell subpopulation, which is mandatory to achieve an effective treatment for this tumor. Differentiation-inducing and epigenetic therapies are the most promising approaches to affect the multiple properties of GSCs and, finally, defeat GBM.

MiR-608 inhibits the migration and invasion of glioma stem cells by targeting macrophage migration inhibitory factor.

Glioma stem cells (GSCs) contribute to the malignant biological behavior of these tumors and have also been shown to be resistant to radiation and chemotherapy. Recently, a variety of microRNAs (miRNAs) has been found to present altered expression and to play an important oncogenic role or tumor-suppressive function in cancer stem cells (CSCs). microRNA-608 (miR-608) is one of the newly discovered microRNAs, and its biological functions remain unclear. Human macrophage migration inhibitory factor (MIF) is a well known oncogene associated with tumor recurrence and the poor prognosis of gliomas. In the present study, we found that miR-608 negatively regulated the gene expression of MIF at the post‑transcriptional level and plays a tumor-suppressive role by targeting MIF in GSCs. We found that miR-608 expression was significantly downregulated and the expression levels of the MIF gene and protein showed an increase in the GSCs. miR-608 overexpression significantly attenuated the proliferation, migration and invasion, and induced the apoptosis of GSCs. The dual-luciferase reporter system revealed that the 3'UTR of MIF is a direct target of miR-608, and miR-608 negatively regulates the gene expression of MIF at the post-transcriptional level by targeting its 3'UTR. Furthermore, we demonstrated that miR-608 overexpression inhibited the malignant behavior of GSCs by downregulating MIF. Western blot results showed that the inhibition of MIF resulted in the inactivation of the PI3K/AKT and JNK pathways. These results demonstrate that miR-608 acts as a potential tumor suppressor and provide insight into new therapeutic targets for malignant glioma.

MiR-218-5p inhibits the stem cell properties and invasive ability of the A2B5⁺CD133⁻ subgroup of human glioma stem cells.

MicroRNAs (miRs) act as oncogenes or tumor-suppressor genes, and regulate the proliferation, apoptosis, invasion, differentiation, angiogenesis and behavior of glioma stem cells, which are important in glioma development and recurrence. The present study was performed to investigate the impact of miR-218-5p on stem cell properties and invasive ability of the A2B5+CD133- human glioma stem cell subgroup. qRT-PCR was used to detect miR-218-5p expression in non-cancerous brain and human glioma tissues, human glioma cell lines and human glioma stem cell lines. Lentivirus vectors encoding miR-218-5p and anti-miR-218-5p were constructed and stably transfected into A2B5+CD133- SHG-139s cells. Neurosphere formation Cell Counting Kit-8 (CCK-8) and Transwell assays, immunofluorescence and qRT-PCR analyses were used to explore the role of miR-218-5p in SHG-139s cells. qRT-PCR analysis showed that miR-218-5p expression was lower in human glioma tissues and cells than in non-cancerous brain tissues and normal human astrocyte cells, and lower in A2B5+CD133- (SHG-139s) cells than in CD133+ (SU2 and U87s) cells. The CCK-8 assay demonstrated that the growth curve was significantly inhibited in the miR-218-5p-SHG-139s cells compared to the miR-control, blank and anti-miR‑218-5p groups. The neurosphere formation assay indicated that upregulation of miR‑218-5p expression inhibited SHG-139s neurosphere formation. Immunofluorescence staining and qRT-PCR showed that miR-218-5p reduced stem cell marker (A2B5, nestin, PLAGL2, ALDH1 and Sox2) expression compared with the controls; however, immunofluorescence staining analysis showed that upregulation of miR-218-5p expression led to no difference in CD133 expression. miR‑218-5p reduced SHG-139s cell invasiveness in the Transwell assay and reduced MMP9 expression as detected in qRT-PCR and immunofluorescence analyses. All differences were statistically significant. miR-218-5p expression was lower in human glioma tissues, cells and the A2B5+CD133- human glioma stem cell subgroup. miR-218-5p may be a tumor-suppressor gene in glioma that functions by upregulating miR-218-5p expression, which inhibits the stem cell properties and invasive properties of SHG-139s cells.

SC-02 * CONVERSION OF DIFFERENTIATED CANCER CELLS INTO CANCER STEM-LIKE CELLS IN A GLIOBLASTOMA MODEL AFTER PRIMARY CHEMOTHERAPY

Glioblastoma multiforme patients have a poor prognosis due to therapeutic resistance and tumor relapse. It has been suggested that gliomas are driven by a rare subset of tumor cells known as glioma stem cells (GSCs). This hypothesis states that only a few GSCs are able to divide, differentiate and initiate a new tumor. It has also been shown that this subpopulation is more resistant to conventional therapies than its differentiated counterpart. In order to understand glioma recurrence post therapy, we investigated the behavior of GSCs after primary chemotherapy. We first show that exposure of patient-derived as well as established glioma cell lines to therapeutic doses of temozolomide (TMZ), the most commonly used anti-glioma chemotherapy, consistently increases the GSC pool over time both in vitro and in vivo. Secondly, lineage-tracing analysis of the expanded GSC pool suggests that such amplification is a result of a phenotypic shift in the non-GSC population to GSC-like state in the presence of TMZ. The newly converted GSC population expresses markers associated with pluripotency and stemness, such as CD133, SOX2, Oct4 and Nestin. Furthermore, we show that intracranial implantation of the newly converted GSCs in nude mice results in a more efficient grafting and invasive phenotype. Taken together, these findings provide the first evidence that glioma cells exposed to chemotherapeutic agents are able to interconvert between non-GSCs and GSCs, thereby replenishing the original tumor population, leading to a more infiltrative phenotype and enhanced chemoresistance. This may represent a potential mechanism for therapeutic relapse.

Conversion of differentiated cancer cells into cancer stem-like cells in a glioblastoma model after primary chemotherapy

Glioblastoma multiforme patients have a poor prognosis due to therapeutic resistance and tumor relapse. It has been suggested that gliomas are driven by a rare subset of tumor cells known as glioma stem cells (GSCs). This hypothesis states that only a few GSCs are able to divide, differentiate, and initiate a new tumor. It has also been shown that this subpopulation is more resistant to conventional therapies than its differentiated counterpart. In order to understand glioma recurrence post therapy, we investigated the behavior of GSCs after primary chemotherapy. We first show that exposure of patient-derived as well as established glioma cell lines to therapeutic doses of temozolomide (TMZ), the most commonly used antiglioma chemotherapy, consistently increases the GSC pool over time both in vitro and in vivo. Secondly, lineage-tracing analysis of the expanded GSC pool suggests that such amplification is a result of a phenotypic shift in the non-GSC population to a GSC-like state in the presence of TMZ. The newly converted GSC population expresses markers associated with pluripotency and stemness, such as CD133, SOX2, Oct4, and Nestin. Furthermore, we show that intracranial implantation of the newly converted GSCs in nude mice results in a more efficient grafting and invasive phenotype. Taken together, these findings provide the first evidence that glioma cells exposed to chemotherapeutic agents are able to interconvert between non-GSCs and GSCs, thereby replenishing the original tumor population, leading to a more infiltrative phenotype and enhanced chemoresistance. This may represent a potential mechanism for therapeutic relapse.

Activation of PI3K/Akt pathway by CD133-p85 interaction promotes tumorigenic capacity of glioma stem cells

The biological significance of a known normal and cancer stem cell marker CD133 remains elusive. We now demonstrate that the phosphorylation of tyrosine-828 residue in CD133 C-terminal cytoplasmic domain mediates direct interaction between CD133 and phosphoinositide 3-kinase (PI3K) 85 kDa regulatory subunit (p85), resulting in preferential activation of PI3K/protein kinase B (Akt) pathway in glioma stem cell (GSC) relative to matched nonstem cell. CD133 knockdown potently inhibits the activity of PI3K/Akt pathway with an accompanying reduction in the self-renewal and tumorigenicity of GSC. The inhibitory effects of CD133 knockdown could be completely rescued by expression of WT CD133, but not its p85-binding deficient Y828F mutant. Analysis of glioma samples reveals that CD133 Y828 phosphorylation level is correlated with histopathological grade and overlaps with Akt activation. Our results identify the CD133/PI3K/Akt signaling axis, exploring the fundamental role of CD133 in glioma stem cell behavior.

MicroRNA Functions and Potential Clinical Utility in Glioblastoma

A flurry of recent research has established that microRNAs have fundamental roles in the biology of the aggressive brain tumor, glioblastoma multiforme. MicroRNAs function in multiple hallmark biological characteristics of glioblastoma, including cell proliferation, invasion, glioma stem cell behavior, and angiogenesis as well as sensitivity to conventional therapies. Many of these microRNAs intersect with key pathways in glioblastoma, indicating their importance in the disease. MicroRNAs also have prognostic potential, may be useful biomarkers and can be used to define subclasses of glioblastoma. MicroRNAs have also shown therapeutic activity in preclinical cancer models.

MicroRNA Functions and Potential Clinical Utility in Glioblastoma

A flurry of recent research has established that microRNAs have fundamental roles in the biology of the aggressive brain tumor, glioblastoma multiforme. MicroRNAs function in multiple hallmark biological characteristics of glioblastoma, including cell proliferation, invasion, glioma stem cell behavior, and angiogenesis as well as sensitivity to conventional therapies. Many of these microRNAs intersect with key pathways in glioblastoma, indicating their importance in the disease. MicroRNAs also have prognostic potential, may be useful biomarkers and can be used to define subclasses of glioblastoma. MicroRNAs have also shown therapeutic activity in preclinical cancer models. Keywords: MicroRNA, glioblastoma, therapeutic, biomarker, subclassification, biological regulatory networks, non-coding RNAs

Distribution of CD133 reveals glioma stem cells self-renew through symmetric and asymmetric cell divisions

Malignant gliomas contain a population of self-renewing tumorigenic stem-like cells; however, it remains unclear how these glioma stem cells (GSCs) self-renew or generate cellular diversity at the single-cell level. Asymmetric cell division is a proposed mechanism to maintain cancer stem cells, yet the modes of cell division that GSCs utilize remain undetermined. Here, we used single-cell analyses to evaluate the cell division behavior of GSCs. Lineage-tracing analysis revealed that the majority of GSCs were generated through expansive symmetric cell division and not through asymmetric cell division. The majority of differentiated progeny was generated through symmetric pro-commitment divisions under expansion conditions and in the absence of growth factors, occurred mainly through asymmetric cell divisions. Mitotic pair analysis detected asymmetric CD133 segregation and not any other GSC marker in a fraction of mitoses, some of which were associated with Numb asymmetry. Under growth factor withdrawal conditions, the proportion of asymmetric CD133 divisions increased, congruent with the increase in asymmetric cell divisions observed in the lineage-tracing studies. Using single-cell-based observation, we provide definitive evidence that GSCs are capable of different modes of cell division and that the generation of cellular diversity occurs mainly through symmetric cell division, not through asymmetric cell division.

167 MicroRNA-34 family in triple-negative/basal-like breast cancer

MicroRNAs are endogenously expressed regulatory noncoding RNAs. Altered expression levels of several microRNAs have been observed in glioblastomas. Functions and direct mRNA targets for these microRNAs have been relatively well studied over the last years. According to these data, it is now evident, that impairment of microRNA regulatory network is one of the key mechanisms in glioblastoma pathogenesis. MicroRNA deregulation is involved in processes such as cell proliferation, apoptosis, cell cycle regulation, invasion, glioma stem cell behavior, and angiogenesis. In this review, we summarize the current knowledge of miRNA functions in glioblastoma with an emphasis on its significance in glioblastoma oncogenic signaling and its potential to serve as a disease biomarker and a novel therapeutic target in oncology.

MicroRNA involvement in glioblastoma pathogenesis

MicroRNAs are endogenously expressed regulatory noncoding RNAs. Altered expression levels of several microRNAs have been observed in glioblastomas. Functions and direct mRNA targets for these microRNAs have been relatively well studied over the last years. According to these data, it is now evident, that impairment of microRNA regulatory network is one of the key mechanisms in glioblastoma pathogenesis. MicroRNA deregulation is involved in processes such as cell proliferation, apoptosis, cell cycle regulation, invasion, glioma stem cell behavior, and angiogenesis. In this review, we summarize the current knowledge of miRNA functions in glioblastoma with an emphasis on its significance in glioblastoma oncogenic signaling and its potential to serve as a disease biomarker and a novel therapeutic target in oncology.

Emerging functions of microRNAs in glioblastoma

Distinct patterns of microRNA expression have been observed in glioblastomas. The functional significance of some of these microRNAs is beginning to emerge. This data indicates that microRNAs play roles in multiple hallmark biological characteristics of glioblastoma, including cell proliferation, invasion, glioma stem cell behavior, and angiogenesis. Research in this area is quickly gathering pace and is illuminating important aspects of the disease that may ultimately lead to novel therapeutic interventions, as well as diagnostic and prognostic tools for brain tumors.