Glioblastoma-initiating Cells Research Articles

Intracranial Assessment of Androgen Receptor Antagonists in Mice Bearing Human Glioblastoma Implants.

The median survival time of patients with an aggressive brain tumor, glioblastoma, is still poor due to ineffective treatment. The discovery of androgen receptor (AR) expression in 56% of cases offers a potential breakthrough. AR antagonists, including bicalutamide and enzalutamide, induce dose-dependent cell death in glioblastoma and glioblastoma-initiating cell lines (GIC). Oral enzalutamide at 20 mg/kg reduces subcutaneous human glioblastoma xenografts by 72% (p = 0.0027). We aimed to further investigate the efficacy of AR antagonists in intracranial models of human glioblastoma. In U87MG intracranial models, nude mice administered Xtandi (enzalutamide) at 20 mg/kg and 50 mg/kg demonstrated a significant improvement in survival compared to the control group (p = 0.24 and p < 0.001, respectively), confirming a dose-response relationship. Additionally, we developed a newly reformulated version of bicalutamide, named "soluble bicalutamide (Bic-sol)", with a remarkable 1000-fold increase in solubility. This reformulation significantly enhanced bicalutamide levels within brain tissue, reaching 176% of the control formulation's area under the curve. In the U87MG intracranial model, both 2 mg/kg and 4 mg/kg of Bic-sol exhibited significant efficacy compared to the vehicle-treated group (p = 0.0177 and p = 0.00364, respectively). Furthermore, combination therapy with 8 mg/kg Bic-sol and Temozolomide (TMZ) demonstrated superior efficacy compared to either Bic-sol or TMZ as monotherapies (p = 0.00706 and p = 0.0184, respectively). In the ZH-161 GIC mouse model, the group treated with 8 mg/kg Bic-sol as monotherapy had a significantly longer lifespan than the groups treated with TMZ or the vehicle (p < 0.001). Our study demonstrated the efficacy of androgen receptor antagonists in extending the lifespan of mice with intracranial human glioblastoma, suggesting a promising approach to enhance patient outcomes in the fight against this challenging disease.

Uptake of H-ferritin by Glioblastoma stem cells and its impact on their invasion capacity.

Iron acquisition is key to maintaining cell survival and function. Cancer cells in general are considered to have an insatiable iron need. Iron delivery via the transferrin/transferrin receptor pathway has been the canonical iron uptake mechanism. Recently, however, our laboratory and others have explored the ability of ferritin, particularly the H-subunit, to deliver iron to a variety of cell types. Here, we investigate whether Glioblastoma (GBM) initiating cells (GICs), a small population of stem-like cells, are known for their iron addiction and invasive nature acquire exogenous ferritin, as a source of iron. We further assess the functional impact of ferritin uptake on the invasion capacity of the GICs. To establish that H-ferritin can bind to human GBM, tissue-binding assays were performed on samples collected at the time of surgery. To interrogate the functional consequences of H-ferritin uptake, we utilized two patient-derived GIC lines. We further describe H-ferritin's impact on GIC invasion capacity using a 3D invasion assay. H-ferritin bound to human GBM tissue at the amount of binding was influenced by sex. GIC lines showed uptake of H-ferritin protein via transferrin receptor. FTH1 uptake correlated with a significant decrease in the invasion capacity of the cells. H-ferritin uptake was associated with a significant decrease in the invasion-related protein Rap1A. These findings indicate that extracellular H-ferritin participates in iron acquisition to GBMs and patient-derived GICs. The functional significance of the increased iron delivery by H-ferritin is a decreased invasion capacity of GICs potentially via reduction of Rap1A protein levels.

ADAMTS1 Supports Endothelial Plasticity of Glioblastoma Cells with Relevance for Glioma Progression.

Gliomas in general and the more advanced glioblastomas (GBM) in particular are the most usual tumors of the central nervous system with poor prognosis. GBM patients develop resistance to distinct therapies, in part due to the existence of tumor cell subpopulations with stem-like properties that participate in trans-differentiation events. Within the complex tumor microenvironment, the involvement of extracellular proteases remains poorly understood. The extracellular protease ADAMTS1 has already been reported to contribute to the plasticity of cancer cells. Accordingly, this basic knowledge and the current availability of massive sequencing data from human gliomas, reinforced the development of this work. We first performed an in silico study of ADAMTS1 and endothelial markers in human gliomas, providing the basis to further assess these molecules in several primary glioblastoma-initiating cells and established GBM cells with the ability to acquire an endothelial-like phenotype. Using a co-culture approach of endothelial and GBM cells, we noticed a relevant function of ADAMTS1 in GBM cells leading the organization of endothelial-like networks and, even more significantly, we found a blockade of the formation of tumor-spheres and a deficient response to hypoxia in the absence of ADAMTS1. Our data support a chief role of this protease modulating the phenotypic plasticity of GBM.

Suppressing Dazl modulates tumorigenicity and stemness in human glioblastoma cells

BackgroundGlioblastoma is devastating cancer with a high frequency of occurrence and poor survival rate and it is urgent to discover novel glioblastoma-specific antigens for the therapy. Cancer-germline genes are known to be related to the formation and progression of several cancer types by promoting tumor transformation. Dazl is one such germline gene and is up-regulated in a few germ cell cancers. In this study, we analyzed the expression of Dazl in human glioblastoma tissues and cells, and investigated its significance in proliferation, migration, invasion and chemoresistance of the glioblastoma cell lines.MethodsWe evaluated the expression of Dazl in different pathologic grades of glioblastoma tissues by immunohistochemistry. We assessed the expression of Dazl in glioblastoma cells and normal human astrocytes (NHA) cells by western blotting and RT-qPCR. Then we generated Dazl knockout glioblastoma cell lines using the CRISPR/Cas9 gene-editing technology to explore the cellular function of Dazl. We detected the proliferation and germline traits via CCK-8 assays and alkaline phosphatase staining, respectively. Boyden chamber assays were performed to measure glioblastoma cell migration and invasion. Crystal violet staining was used to determine the number of viable cells after the treatment of Doxorubicin and Temozolomide. Finally, we used subcutaneous xenograft studies to measure the growth of tumors in vivo.ResultsWe found that Dazl was upregulated in glioblastoma tissues and glioblastoma cell lines. Dazl knockdown glioblastoma cells showed decreased cellular proliferation, migration, invasion, and resistance in vitro, and inhibited the initiation of glioblastoma in vivo. The glioblastoma cell lines A172, U251, and LN229 were found to express stem cell markers CD133, Oct4, Nanog, and Sox2. The expression of these markers was downregulated in Dazl-deficient cells.ConclusionsOur results indicated that Dazl contributes to the tumorigenicity of glioblastoma via reducing cell stemness. Therefore, cancer-germline genes might represent a new paradigm of glioblastoma-initiating cells in the treatment of malignant tumors.

Growth-Inhibitory Activity of Bone Morphogenetic Protein 4 in Human Glioblastoma Cell Lines Is Heterogeneous and Dependent on Reduced SOX2 Expression.

Glioblastoma multiforme continues to have a dismal prognosis. Even though detailed information on the genetic aberrations in cell signaling and cell-cycle checkpoint control is available, no effective targeted treatment has been developed. Despite the advanced molecular defects, glioblastoma cells may have remnants of normal growth-inhibitory pathways, such as the bone morphogenetic protein (BMP) signaling pathway. We have evaluated the growth-inhibitory effect of BMP4 across a broad spectrum of patient samples, using a panel of 40 human glioblastoma initiating cell (GIC) cultures. A wide range of responsiveness was observed. BMP4 sensitivity was positively correlated with a proneural mRNA expression profile, high SOX2 activity, and BMP4-dependent upregulation of genes associated with inhibition of the MAPK pathway, as demonstrated by gene set enrichment analysis. BMP4 response in sensitive cells was mediated by the canonical BMP receptor pathway involving SMAD1/5/9 phosphorylation and SMAD4 expression. SOX2 was consistently downregulated in BMP4-treated cells. Forced expression of SOX2 attenuated the BMP4 sensitivity including a reduced upregulation of MAPK-inhibitory genes, implying a functional relationship between SOX2 downregulation and sensitivity. The results show an extensive heterogeneity in BMP4 responsiveness among GICs and identify a BMP4-sensitive subgroup, in which SOX2 is a mediator of the response. IMPLICATIONS: Development of agonists targeting the BMP signaling pathway in glioblastoma is an attractive avenue toward a better treatment. Our study may help find biomarkers that predict the outcome of such treatment and enable stratification of patients.

Polycomb-mediated repression of EphrinA5 promotes growth and invasion of glioblastoma

Glioblastoma (GBM) is the most common and most aggressive intrinsic brain tumour in adults. Integrated transcriptomic and epigenomic analyses of glioblastoma initiating cells (GIC) in a mouse model uncovered a novel epigenetic regulation of EfnA5. In this model, Bmi1 enhances H3K27me3 at the EfnA5 locus and reinforces repression of selected target genes in a cellular context-dependent fashion. EfnA5 mediates Bmi1-dependent proliferation and invasion in vitro and tumour formation in an allograft model. Importantly, we show that this novel Polycomb feed-forward loop is also active in human GIC and we provide pre-clinical evidence of druggability of the EFNA5 signalling pathway in GBM xenografts overexpressing Bmi1.

TMIC-22. IDENTIFICATION OF CANCER-ASSOCIATED FIBROBLASTS IN GLIOBLASTOMA and Defining Their Protumoral Effects

Abstract While cancer-associated fibroblasts (CAFs) and their pro-tumoral effects have been demonstrated in systemic cancers, CAFs had been presumed absent in glioblastoma given the lack of normal fibroblasts in the healthy brain. Here, we show that 5–26% (mean=12%) of cells in human glioblastomas express CAF markers α-SMA or PDGFR-β, morphologically resemble fibroblasts, and transcriptomically resemble by RNA-seq CAFs from other cancers. Glioblastoma CAFs were chemotactically attracted to glioblastoma-initiating stem cells (P=0.02). While glioblastoma CAFs did not affect differentiated glioblastoma cell proliferation (P=0.4), CAFs increased glioblastoma stem cell proliferation (P=0.002) and expression of glioblastoma stem cell-associated genes (P&lt; 0.001). To identify mediators of CAF/glioblastoma stem cell interactions, we created a resource of inferred crosstalk by mapping the expression of receptors to that of their cognate ligands/agonists, using our RNA-seq results from glioblastoma CAFs and stem cells, revealing PDGF-β/PDGFR and osteopontin/CD44 to mediate stem cell recruitment of CAFs and CAF enrichment of stem cells, as confirmed by blocking antibodies (P=0.02–0.03). CAFs also render the glioblastoma microenvironment more pro-tumoral by promoting M2 polarization of tumor-associated macrophages (P=0.01), an effect we found to arise from unique CAF production of the EDA splice variant of fibronectin binding toll-like receptor 4 (TLR4), a known EDA receptor expressed by macrophages (P=0.02). In patient glioblastomas, CAFs were enriched 3-fold in the subventricular zone (SVZ) (P=0.04) which houses the neural stem cells that generate glioblastoma stem cells. SVZs from epilepsy cases or autopsies of glioblastoma-containing brains without ventricular involvement lacked CAFs. Depleting CAFs in xenografts derived from neurosphere-containing glioblastoma stem cells slowed their growth in vivo (P&lt; 0.001). These findings are among the first to identify and profile glioblastoma CAFs. CAF recruitment by glioblastoma stem cells and creation of a pro-tumoral microenvironment in the perivascular niche housing glioblastoma stem cells, particularly in the SVZ, makes them an intriguing therapeutic target.

Glioblastoma initiating cells are sensitive to histone demethylase inhibition due to epigenetic deregulation.

Tumor-initiating cells are a subpopulation of cells that have self-renewal capacity to regenerate a tumor. Here, we identify stem cell-like chromatin features in human glioblastoma initiating cells (GICs) and link them to a loss of the repressive histone H3 lysine 9 trimethylation (H3K9me3) mark. Increasing H3K9me3 levels by histone demethylase inhibition led to cell death in GICs but not in their differentiated counterparts. The induction of apoptosis was accompanied by a loss of the activating H3 lysine 9 acetylation (H3K9ac) modification and accumulation of DNA damage and downregulation of DNA damage response genes. Upon knockdown of histone demethylases, KDM4C and KDM7A both differentiation and DNA damage were induced. Thus, the H3K9me3-H3K9ac equilibrium is crucial for GIC viability and represents a chromatin feature that can be exploited to specifically target this tumor subpopulation.

A novel 3D nanofibre scaffold conserves the plasticity of glioblastoma stem cell invasion by regulating galectin-3 and integrin-\u03b21 expression

Glioblastoma Multiforme (GBM) invasiveness renders complete surgical resection impossible and highly invasive Glioblastoma Initiating Cells (GICs) are responsible for tumour recurrence. Their dissemination occurs along pre-existing fibrillary brain structures comprising the aligned myelinated fibres of the corpus callosum (CC) and the laminin (LN)-rich basal lamina of blood vessels. The extracellular matrix (ECM) of these environments regulates GIC migration, but the underlying mechanisms remain largely unknown. In order to recapitulate the composition and the topographic properties of the cerebral ECM in the migration of GICs, we have set up a new aligned polyacrylonitrile (PAN)-derived nanofiber (NF) scaffold. This system is suitable for drug screening as well as discrimination of the migration potential of different glioblastoma stem cells. Functionalisation with LN increases the spatial anisotropy of migration and modulates its mode from collective to single cell migration. Mechanistically, equally similar to what has been observed for mesenchymal migration of GBM in vivo, is the upregulation of galectin-3 and integrin-β1 in Gli4 cells migrating on our NF scaffold. Downregulation of Calpain-2 in GICs migrating in vivo along the CC and in vitro on LN-coated NF underlines a difference in the turnover of focal adhesion (FA) molecules between single-cell and collective types of migration.

The oncogenic roles of 27-hydroxycholesterol in glioblastoma.

Glioblastoma is the most frequent primary malignant brain tumor in adults. Oxysterols are oxidation products of cholesterol generated by enzymatic reactions. 27-hydroxycholesterol (27-HC), an oxysterol, is an abundant metabolite of cholesterol. 27-HC significantly accelerates mammary cancer growth, proliferation and progression in experimental models. However, to the best of our knowledge, the effect of 27-HC on glioblastoma has not been studied. Therefore, the present study aimed to determine the exact role of 27-HC in glioblastoma. The present study demonstrated that 27-HC promoted proliferation, epithelial to mesenchymal transition, colony formation, migration and invasion of U251 and U118 MG glioblastoma cells. Treatment with 27-HC was also associated with an increase in the formation of glioblastoma-initiating cells in U251 and U118 MG cell lines. Additionally, it was observed that high levels of 27-HC in glioblastoma tissues were associated with poor outcome in patients. In conclusion, 27-HC, a primary metabolite of cholesterol, may serve an important role in the progression of glioblastoma.

The efficacy and toxicity of ATM inhibition in glioblastoma initiating cells-driven tumor models.

The Ataxia Telangiectasia Mutated (ATM)-mediated DNA damage response (DDR) is a major mechanism of resistance of glioblastoma (GB) - initiating cells (GICs) to radiotherapy. The closely related Ataxia Telangiectasia and Rad3-related protein (ATR) is also involved in tumor resistance to radio- and chemotherapy. It has been shown that pharmacological inhibition of ATM protein may overcome the DDR-mediated resistance in GICs and significantly radiosensitize GIC-driven GB. Albeit not essential for life as shown by the decade-long lifespan of AT patients, the ATM protein may be involved in a number of important functions other than the response to DNA damage. We discuss our current knowledge about the toxicity of pharmacologic inhibition of ATM and ATR proteins.

Inhibiting Integrin β8 to Differentiate and Radiosensitize Glioblastoma-Initiating Cells.

Glioblastomas (GB) are malignant brain tumors with poor prognosis despite treatment with surgery and radio/chemotherapy. These tumors are defined by an important cellular heterogeneity and notably contain a subpopulation of GB-initiating cells (GIC), which contribute to tumor aggressiveness, resistance, and recurrence. Some integrins are specifically expressed by GICs and could be actionable targets to improve GB treatment. Here, integrin β8 (ITGB8) was identified as a potential selective target in this highly tumorigenic GIC subpopulation. Using several patient-derived primocultures, it was demonstrated that ITGB8 is overexpressed in GICs compared with their differentiated progeny. Furthermore, ITGB8 is also overexpressed in GB, and its overexpression is correlated with poor prognosis and with the expression of several other classic stem cell markers. Moreover, inhibiting ITGB8 diminished several main GIC characteristics and features, including self-renewal ability, stemness, migration potential, and tumor formation capacity. Blockade of ITGB8 significantly impaired GIC cell viability via apoptosis induction. Finally, the combination of radiotherapy and ITGB8 targeting radiosensitized GICs through postmitotic cell death. IMPLICATIONS: This study identifies ITGB8 as a new selective marker for GICs and as a promising therapeutic target in combination with chemo/radiotherapy for the treatment of highly aggressive brain tumors.

Theranostic nanoparticles enhance the response of glioblastomas to radiation.

Despite considerable progress with our understanding of glioblastoma multiforme (GBM) and the precise delivery of radiotherapy, the prognosis for GBM patients is still unfavorable with tumor recurrence due to radioresistance being a major concern. We recently developed a cross-linked iron oxide nanoparticle conjugated to azademethylcolchicine (CLIO-ICT) to target and eradicate a subpopulation of quiescent cells, glioblastoma initiating cells (GICs), which could be a reason for radioresistance and tumor relapse. The purpose of our study was to investigate if CLIO-ICT has an additive therapeutic effect to enhance the response of GBMs to ionizing radiation.Methods: NSG™ mice bearing human GBMs and C57BL/6J mice bearing murine GBMs received CLIO-ICT, radiation, or combination treatment. The mice underwent pre- and post-treatment magnetic resonance imaging (MRI) scans, bioluminescence imaging (BLI), and histological analysis. Tumor nanoparticle enhancement, tumor flux, microvessel density, GIC, and apoptosis markers were compared between different groups using a one-way ANOVA and two-tailed Mann-Whitney test. Additional NSG™ mice underwent survival analyses with Kaplan-Meier curves and a log rank (Mantel-Cox) test.Results: At 2 weeks post-treatment, BLI and MRI scans revealed significant reduction in tumor size for CLIO-ICT plus radiation treated tumors compared to monotherapy or vehicle-treated tumors. Combining CLIO-ICT with radiation therapy significantly decreased microvessel density, decreased GICs, increased caspase-3 expression, and prolonged the survival of GBM-bearing mice. CLIO-ICT delivery to GBM could be monitored with MRI. and was not significantly different before and after radiation. There was no significant caspase-3 expression in normal brain at therapeutic doses of CLIO-ICT administered.Conclusion: Our data shows additive anti-tumor effects of CLIO-ICT nanoparticles in combination with radiotherapy. The combination therapy proposed here could potentially be a clinically translatable strategy for treating GBMs.

MiR-186 reverses cisplatin resistance and inhibits the�formation of the glioblastoma-initiating cell�phenotype by degrading Yin Yang 1 in glioblastoma

Glioblastoma multiforme (GBM) is among the most devastating types of cancer, with a median survival of <1year. Despite the development of new surgical and radiation techniques, and the use of multiple anti‑neoplastic drugs, effective treatment strategies for malignant gliomas have not yet been developed. The limited efficacy of current treatments reflects the resistance of glioblastoma cells to cytotoxic agents. In this study, using western blot analysis, we found that YinYang1(YY1) expression was increased in cisplatin‑resistant glioblastoma U87MG cells (U87MG‑CR). We observed that the silencing of YY1 sensitized the U87MG‑CR cells to cisplatin and that the overexpression of YY1 promoted the resistance of LN‑229 glioblastoma cells to cisplatin, as shown by MTT assay. Using sphere formation assay, we also found that the silencing of YY1 inhibited the formation of the glioblastoma‑initiating cell (GIC) phenotype in the U87MG‑CR cells. In addition, the results of RT‑qPCR revealed that miR‑186 expression was decreased in U87MG‑CR cells. Using RT‑PCR and western blot analysis, we observed that overexpression of miR‑186 inhibited YY1 expression in U87MG‑CR cells. The overexpression of miR‑186 also reversed cisplatin resistance and the formation of the GIC phenotype in glioblastoma cells. On the whole, the findings of this study demonstrate that miR‑186 reverses cisplatin resistance and inhibits the formation of the GIC phenotype by degrading YY1 in glioblastoma.

Inhibition of sonic hedgehog and PI3K/Akt/mTOR pathways cooperate in suppressing survival, self-renewal and tumorigenic potential of glioblastoma-initiating cells.

Since PI3K/Akt/mTOR and sonic hedgehog (SHH) signaling pathways are highly activated in glioblastoma-initiating cells (GICs), we examined the effects of inhibiting these pathways on GIC characteristics and tumor growth in mice. NVP-LDE-225 (inhibitor of Smoothened) inhibited the expression of Gli1, Gli2, Smoothened, Patched1, and Patched2, and induced the expression of SuFu, whereas NVP-BEZ-235 (dual inhibitor of PI3K and mTOR) inhibited the expression of p-PI3K, p-Akt, p-mTOR, and p-p70S6K. NVP-LDE-225 co-operated with NVP-BEZ-235 in inhibiting the self-renewal capacity of GICs, expression of pluripotency maintaining factors (Nanog, c-Myc, Oct4, and Sox2), Musashi1, cyclin D1, and Bcl-2, and transcription and expression of Gli, and in inducing the expression of cleaved caspase-3, cleaved PARP and Bim. Additionally, NVP-LDE-225 co-operated with NVP-BEZ-235 in inhibiting epithelial-mesenchymal transition. Finally, the combination of NVP-LDE-225 and NVP-BEZ-235 was superior in inhibiting tumor growth, regulating the expression of pluripotency promoting factors, stem cell markers, cell cycle, and cell proliferation, and modulating EMT compared to single agent alone. In conclusion, the combined inhibition of PI3K/Akt/mTOR and SHH pathways was superior to single pathway inhibition in suppressing glioblastoma growth by targeting GICs.

Aurora A plays a dual role in migration and survival of human glioblastoma cells according to the CXCL12 concentration

Primary glioblastoma is the most frequent human brain tumor in adults and is generally fatal due to tumor recurrence. We previously demonstrated that glioblastoma-initiating cells invade the subventricular zones and promote their radio-resistance in response to the local release of the CXCL12 chemokine. In this work, we show that the mitotic Aurora A kinase (AurA) is activated through the CXCL12–CXCR4 pathway in an ERK1/2-dependent manner. Moreover, the CXCL12–ERK1/2 signaling induces the expression of Ajuba, the main cofactor of AurA, which allows the auto-phosphorylation of AurA.We show that AurA contributes to glioblastoma cell survival, radio-resistance, self-renewal, and proliferation regardless of the exogenous stimulation with CXCL12. On the other hand, AurA triggers the CXCL12-mediated migration of glioblastoma cells in vitro as well as the invasion of the subventricular zone in xenograft experiments. Moreover, AurA regulates cytoskeletal proteins (i.e., Actin and Vimentin) and favors the pro-migratory activity of the Rho-GTPase CDC42 in response to CXCL12. Altogether, these results show that AurA, a well-known kinase of the mitotic machinery, may play alternative roles in human glioblastoma according to the CXCL12 concentration.

MicroRNA-132 induces temozolomide resistance and promotes the formation of cancer stem cell phenotypes by targeting tumor suppressor candidate3 in glioblastoma.

The prognosis of patients suffering from glioblastoma [also referred to as glioblastoma multiforme (GBM)] is dismal despite multimodal therapy. Chemotherapy with temozolomide may suppress tumor growth for a certain period of time (a few months); however, invariable tumor recurrence suggests that glioblastoma initiating cells (GICs) render these tumors persistant. Thus, the understanding of the molecular mechanisms of action of GICs as regards their role in the progression of GBM is important as such knowledge will be helpful in the discovery of novel drug targets, as well as in the design of novel therapeutic strategies for more effective treatment of the disease. In this study, we found that tumor suppressor candidate 3 (TUSC3) was downregulated in temozolomide-resistant U87MG cells (U87MG-res cells) and its restoration sensitized U87MG-res cells to temozolomide. TUSC3 was able to inhibit the formation of GIC phenotypes in the U87MG-res cells. The overexpression of microRNA (miR)-132 inhibited TUSC3 protein expression in the U87MG cells. However, its overexpression did not degrade TUSC3 mRNA expression in the cells. miR-132 was upregulated in the U87MG-res cells and its overexpression induced temozolomide resistance and the formation of cancer stem cell phenotypes in the U87MG cells. Thus, our data indicate that miR-132 induces temozolo-mide resistance and promotes the formation of cancer stem cell phenotypes by targeting TUSC3 in glioblastoma.

Interest of integrins targeting in glioblastoma according to tumor heterogeneity and cancer stem cell paradigm: an update.

Glioblastomas are malignant brain tumors with dismal prognosis despite standard treatment with surgery and radio/chemotherapy. These tumors are defined by an important cellular heterogeneity and notably contain a particular subpopulation of Glioblastoma-initiating cells, which recapitulate the heterogeneity of the original Glioblastoma. In order to classify these heterogeneous tumors, genomic profiling has also been undertaken to classify these heterogeneous tumors into several subtypes. Current research focuses on developing therapies, which could take into account this cellular and genomic heterogeneity. Among these targets, integrins are the subject of numerous studies since these extracellular matrix transmembrane receptors notably controls tumor invasion and progression. Moreover, some of these integrins are considered as membrane markers for the Glioblastoma-initiating cells subpopulation. We reviewed here integrin expression according to glioblastoma molecular subtypes and cell heterogeneity. We discussed their roles in glioblastoma invasion, angiogenesis, therapeutic resistance, stemness and microenvironment modulations, and provide an overview of clinical trials investigating integrins in glioblastomas. This review highlights that specific integrins could be identified as selective glioblastoma cells markers and that their targeting represents new diagnostic and/or therapeutic strategies.

Radioresistance of mesenchymal glioblastoma initiating cells correlates with patient outcome and is associated with activation of inflammatory program.

Glioblastoma (GBM) still remains an incurable disease being radiotherapy (RT) the mainstay treatment. Glioblastoma intra-tumoral heterogeneity and Glioblastoma-Initiating Cells (GICs) challenge the design of effective therapies. We investigated GICs and non-GICs response to RT in a paired in-vitro model and addressed molecular programs activated in GICs after RT. Established GICs heterogeneously expressed several GICs markers and displayed a mesenchymal signature. Upon fractionated RT, GICs reported higher radioresistance compared to non-GICs and showed lower α- and β-values, according to the Linear Quadratic Model interpretation of the survival curves. Moreover, a significant correlation was observed between GICs radiosensitivity and patient disease-free survival. Transcriptome analysis of GICs after acquisition of a radioresistant phenotype reported significant activation of Proneural-to-Mesenchymal transition (PMT) and pro-inflammatory pathways, being STAT3 and IL6 the major players. Our findings support a leading role of mesenchymal GICs in defining patient response to RT and provide the grounds for targeted therapies based on the blockade of inflammatory pathways to overcome GBM radioresistance.

β-escin selectively targets the glioblastoma-initiating cell population and reduces cell viability.

Glioblastoma multiforme (GBM) is a highly aggressive tumour of the central nervous system and is associated with an extremely poor prognosis. Within GBM exists a subpopulation of cells, glioblastoma-initiating cells (GIC), which possess the characteristics of progenitor cells, have the ability to initiate tumour growth and resist to current treatment strategies. We aimed at identifying novel specific inhibitors of GIC expansion through use of a large-scale chemical screen of approved small molecules. Here, we report the identification of the natural compound β-escin as a selective inhibitor of GIC viability. Indeed, β-escin was significantly cytotoxic in nine patient-derived GIC, whilst exhibiting no substantial effect on the other human cancer or control cell lines tested. In addition, β-escin was more effective at reducing GIC growth than current clinically used cytotoxic agents. We further show that β-escin triggers caspase-dependent cell death combined with a loss of stemness properties. However, blocking apoptosis could not rescue the β-escin-induced reduction in sphere formation or stemness marker activity, indicating that β-escin directly modifies the stem identity of GIC, independent of the induction of cell death. Thus, this study has repositioned β-escin as a promising potential candidate to selectively target the aggressive population of initiating cells within GBM.