Chemoresistance In Glioblastoma Multiforme Research Articles

Role of Non-coding RNAs in the Response of Glioblastoma to Temozolomide.

Chemotherapy and radiotherapy are widely used in clinical practice across the globe as cancer treatments. Intrinsic or acquired chemoresistance poses a significant problem for medical practitioners and researchers, causing tumor recurrence and metastasis. The most dangerous kind of malignant brain tumor is called glioblastoma multiforme (GBM) that often recurs following surgery. The most often used medication for treating GBM is temozolomide chemotherapy; however, most patients eventually become resistant. Researchers are studying preclinical models that accurately reflect human disease and can be used to speed up drug development to overcome chemoresistance in GBM. Non-coding RNAs (ncRNAs) have been shown to be substantial in regulating tumor development and facilitating treatment resistance in several cancers, such as GBM. In this work, we mentioned the mechanisms of how different ncRNAs (microRNAs, long non-coding RNAs, circular RNAs) can regulate temozolomide chemosensitivity in GBM. We also address the role of these ncRNAs encapsulated inside secreted exosomes.

Functional importance of glucose transporters and chromatin epigenetic factors in Glioblastoma Multiforme (GBM): possible therapeutics.

Glioblastoma Multiforme (GBM) is an aggressive brain cancer affecting glial cells and is chemo- and radio-resistant. Glucose is considered the most vital energy source for cancer cell proliferation. During metabolism, hexose molecules will be transported into the cells via transmembrane proteins known as glucose transporter (GLUT). Among them, GLUT-1 and GLUT-3 play pivotal roles in glucose transport in GBM. Knockdown studies have established the role of GLUT-1, and GLUT-3 mediated glucose transport in GBM cells, providing insight into GLUT-mediated cancer signaling and cancer aggressiveness. This review focussed on the vital role of GLUT-1 and GLUT-3 proteins, which regulate glucose transport. Recent studies have identified the role of GLUT inhibitors in effective cancer prevention. Several of them are in clinical trials. Understanding and functional approaches towards glucose-mediated cell metabolism and chromatin epigenetics will provide valuable insights into the mechanism of cancer aggressiveness, cancer stemness, and chemo-resistance in Glioblastoma Multiforme (GBM). This review summarizes the role of GLUT inhibitors, micro-RNAs, and long non-coding RNAs that aid in inhibiting glucose uptake by the GBM cells and other cancer cells leading to the identification of potential therapeutic, prognostic as well as diagnostic markers. Furthermore, the involvement of epigenetic factors, such as microRNAs, in regulating glycolytic genes was demonstrated.

Glioblastoma Multiforme: A Review

In recent years, many research groups have begun to utilize bioengineered in vitro models of cancer to study mechanisms of disease progression, test drug candidates, and develop platforms to advance personalized drug treatment options. Due to advances in cell and tissue engineering over the last few decades, there are now a myriad of tools that can be used to create such in vitro systems. In this review, we describe the considerations one must take when developing model systems that accurately mimic the in vivo tumor microenvironment (TME) and can be used to answer specific scientific questions. We will summarize the importance of cell sourcing in models with one or multiple cell types and outline the importance of choosing biomaterials that accurately mimic the native extracellular matrix (ECM) of the tumor or tissue that is being modeled. We then provide examples of how these two components can be used in concert in a variety of model form factors and conclude by discussing how biofabrication techniques such as bioprinting and organ-on-a-chip fabrication can be used to create highly reproducible complex in vitro models. Since this topic has a broad range of applications, we use the final section of the review to dive deeper into one type of cancer, glioblastoma, to illustrate how these components come together to further our knowledge of cancer biology and move us closer to developing novel drugs and systems that improve patient outcomes. Glioblastomas are universally fatal cancers and contain self-renewing glioblastoma stem cells (GSCs) that initiate tumors. Traditional anticancer drug development based on in vitro cultures tends to select targets with low therapeutic indices and fails to effectively represent the tumour microenvironment's impacts. Glioblastoma multiforme (GBM) is the most common and aggressive form of brain cancer, with treatment options often constrained due to inherent resistance of malignant cells to conventional therapy. We investigated the impact of triggering programmed cell death (PCD) by using BH3 mimetic drugs in human GBM cell lines. Glioblastoma multiforme (GBM) has been characterized by the high incidence, therapy tolerance and relapse. The molecular events controlling GBM resistant to chemotherapy temozolomide (TMZ) remain to be elusive. Here, we identified WNT signaling was amplified by TMZ and mediated drug response in GBM. We found O6-methylguanine DNA methyltransferase (MGMT) was redundant to WNT-mediated chemoresistance, which was highly associated with p53 mutation status. In GBM with p53 mutation, loss of function of p53 downregulated miR-34a expression, which represses transcription of WNT ligand 6 (WNT6) by directly binding to 3′ UTR of WNT6 mRNA, leading to activation of WNT signaling, and the eventual WNT-mediated chemoresistance to TMZ. Combined treatment of TMZ with WNT inhibitor or miR34a mimic induced drug sensitivity of p53-mutant GBM cells and extended survival in xenograft mice in vivo. Our findings provide insight into understanding the molecular mechanism of GBM chemoresistance to TMZ and facilitating to develop novel treatment strategy to combat p53-mutant GBM by targeting miR-34a/WNT6 axis. Advantages and drawbacks of dexamethasone in glioblastoma multiforme: Dexamethasone has been used for many years to treat brain edema and inflammation caused by GBM.Several investigations have shown that dexamethasone also exerts antitumoral effects against GBM.Unfortunately, steroids are associated with various undesirable side effects. Herein, we review pre-clinical and clinical applications of Dexamethasone in GBM. The most widespread, malignant, and deadliest type of glial tumor is glioblastoma multiforme (GBM). Despite radiation, chemotherapy, and radical surgery, the median survival of afflicted individuals is about 12 months. Unfortunately, existing therapeutic interventions are abysmal. Dexamethasone (Dex), a synthetic glucocorticoid, has been used for many years to treat brain edema and inflammation caused by GBM. Several investigations have recently shown that Dex also exerts antitumoral effects against GBM. On the other hand, more recent disputed findings have questioned the long-held dogma of Dex treatment for GBM. Unfortunately, steroids are associated with various undesirable side effects, including severe immunosuppression and metabolic changes like hyperglycemia, which may impair the survival of GBM patients. Current ideas and concerns about Dex's effects on GBM cerebral edema, cell proliferation, migration, and its clinical outcomes were investigated in this study. Glioblastoma multiforme (GBM) is one of the most common, most formidable, and deadliest malignant types of primary astrocytoma with a poor prognosis. At present, the standard of care includes surgical tumor resection, followed by radiation therapy concomitant with chemotherapy and temozolomide. New developments and significant advances in the treatment of GBM have been achieved ..........

Downregulated ferroptosis-related gene SQLE facilitates temozolomide chemoresistance, and invasion and affects immune regulation in glioblastoma.

Chemoresistance in patients with glioblastoma multiforme (GBM) is a common reason hindering the success of treatment. Recently, ferroptosis has been reported to be associated with chemoresistance in different types of cancer, while the role of ferroptosis-related genes in GBM have not been fully elucidated. This study aimed to demonstrate the roles and mechanism of ferroptosis-related genes in chemoresistance and metastasis of GBM. First, two candidate genes, squalene epoxidase (SQLE) and FANCD2, were identified to be associated with ferroptosis-related chemoresistance in GBM from three temozolomide (TMZ) therapeutic datasets and one ferroptosis-related gene dataset. Then, comprehensive bio-informatics data from different databases testified that SQLE was significantly downregulated both in GBM tissue and cells and displayed a better prognosis in GBM. Clinical data identified lower expression of SQLE was significantly associated with WHO grade and 1p/19q codeletion. Moreover, through in vitro experiments, SQLE was confirmed to suppress ERK-mediated TMZ chemoresistance and metastasis of GBM cells. The KEGG analysis of SQLE-associated co-expressed genes indicated SQLE was potentially involved in the cell cycle. Furthermore, SQLE was found to have the most significant correlations with tumor-infiltrating lymphocytes and immunomodulators. These findings highlighted that SQLE could be a potential target and a biomarker for therapy and prognosis of patients with GBM.

Ubiquitin-Specific Protease 6 n-Terminal-like Protein (USP6NL) and the Epidermal Growth Factor Receptor (EGFR) Signaling Axis Regulates Ubiquitin-Mediated DNA Repair and Temozolomide-Resistance in Glioblastoma.

Glioblastoma multiforme (GBM) is the most malignant glioma, with a 30–60% epidermal growth factor receptor (EGFR) mutation. This mutation is associated with unrestricted cell growth and increases the possibility of cancer invasion. Patients with EGFR-mutated GBM often develop resistance to the available treatment modalities and higher recurrence rates. The drug resistance observed is associated with multiple genetic or epigenetic factors. The ubiquitin-specific protease 6 N-terminal-like protein (USP6NL) is a GTPase-activating protein that functions as a deubiquitinating enzyme and regulates endocytosis and signal transduction. It is highly expressed in many cancer types and may promote the growth and proliferation of cancer cells. We hypothesized that USP6NL affects GBM chemoresistance and tumorigenesis, and that its inhibition may be a novel therapeutic strategy for GBM treatment. The USP6NL level, together with EGFR expression in human GBM tissue samples and cell lines associated with therapy resistance, tumor growth, and cancer invasion, were investigated. Its pivotal roles and potential mechanism in modulating tumor growth, and the key mechanism associated with therapy resistance of GBM cells, were studied, both in vitro and in vivo. Herein, we found that deubiquitinase USP6NL and growth factor receptor EGFR were strongly associated with the oncogenicity and resistance of GBM, both in vitro and in vivo, toward temozolomide, as evidenced by enhanced migration, invasion, and acquisition of a highly invasive and drug-resistant phenotype by the GBM cells. Furthermore, abrogation of USP6NL reversed the properties of GBM cells and resensitized them toward temozolomide by enhancing autophagy and reducing the DNA damage repair response. Our results provide novel insights into the probable mechanism through which USP6NL/EGFR signaling might suppress the anticancer therapeutic response, induce cancer invasiveness, and facilitate reduced sensitivity to temozolomide treatment in GBM in an autolysosome-dependent manner. Therefore, controlling the USP6NL may offer an alternative, but efficient, therapeutic strategy for targeting and eradicating otherwise resistant and recurrent phenotypes of aggressive GBM cells.

PLGA-Nano-Encapsulated Disulfiram Inhibits Hypoxia-Induced NF-κB, Cancer Stem Cells, and Targets Glioblastoma In Vitro and In Vivo.

Glioblastoma stem cell (GSC) is the major cause of glioblastoma multiforme (GBM) chemotherapy failure. Hypoxia is one of the determinants of GSC. NF-κB plays a pivotal link between hypoxia and cancer stem cells (CSCs). Disulfiram, an antialcoholism drug, has very strong NF-κB-inhibiting and anti-CSC activity. In this study, the in vitro anti-GSC activity of disulfiram and in vivo anti-GBM efficacy of poly lactic-co-glycolic acid nanoparticle-encapsulated disulfiram (DS-PLGA) were examined. We attempt to elucidate the molecular network between hypoxia and GSCs and also examined the anti-GSC activity of disulfiram in vitro and in vivo. The influence of GSCs and hypoxia on GBM chemoresistance and invasiveness was studied in hypoxic and spheroid cultures. The molecular regulatory roles of NF-κB, hypoxia-inducible factor-1α (HIF1α), and HIF2α were investigated using stably transfected U373MG cell lines. The hypoxia in neurospheres determines the cancer stem cell characteristics of the sphere-cultured GBM cell lines (U87MG, U251MG, U373MG). NF-κB is located at a higher hierarchical position than HIF1α/HIF2α in hypoxic regulatory network and plays a key role in hypoxia-induced GSC characters. DS inhibits NF-κB activity and targets hypoxia-induced GSCs. It showed selective toxicity to GBM cells, eradicates GSCs, and blocks migration and invasion at very low concentrations. DS-PLGA efficaciously inhibits orthotopic and subcutaneous U87MG xenograft in mouse models with no toxicity to vital organs.

WNT signaling modulates chemoresistance to temozolomide in p53-mutant glioblastoma multiforme

Glioblastoma multiforme (GBM) has been characterized by the high incidence, therapy tolerance and relapse. The molecular events controlling GBM resistant to chemotherapy temozolomide (TMZ) remain to be elusive. Here, we identified WNT signaling was amplified by TMZ and mediated drug response in GBM. We found O6-methylguanine DNA methyltransferase (MGMT) was redundant to WNT-mediated chemoresistance, which was highly associated with p53 mutation status. In GBM with p53 mutation, loss of function of p53 downregulated miR-34a expression, which represses transcription of WNT ligand 6 (WNT6) by directly binding to 3' UTR of WNT6 mRNA, leading to activation of WNT signaling, and the eventual WNT-mediated chemoresistance to TMZ. Combined treatment of TMZ with WNT inhibitor or miR34a mimic induced drug sensitivity of p53-mutant GBM cells and extended survival in xenograft mice in vivo. Our findings provide insight into understanding the molecular mechanism of GBM chemoresistance to TMZ and facilitating to develop novel treatment strategy to combat p53-mutant GBM by targeting miR-34a/WNT6 axis.

Interfering with mitochondrial dynamics sensitizes glioblastoma multiforme to temozolomide chemotherapy.

Glioblastoma multiforme (GBM) is a primary tumour of the central nervous system (CNS) that exhibits the highest degree of malignancy. Radiotherapy and chemotherapy are essential to prolong the survival time of patients. However, clinical work has demonstrated that sensitivity of GBM to chemotherapy decreases with time. The phenomenon of multi‐drug resistance (MDR) reminds us that there may exist some fundamental mechanisms in the process of chemo‐resistance. We tried to explore the mechanism of GBM chemo‐resistance from the perspective of energy metabolism. First, we found that the oxidative phosphorylation (OXPHOS) level of SHG44 and U87 cells increased under TMZ treatment. In further studies, it was found that the expression of PINK1 and mitophagy flux downstream was downregulated in GBM cells, which were secondary to the upregulation of TP53 in tumour cells under TMZ treatment. At the same time, we examined the mitochondrial morphology in tumour cells and found that the size of mitochondria in tumour cells increased under the treatment of TMZ, which originated from the regulation of AMPK on the subcellular localization of Drp1 under the condition of unbalanced energy supply and demand in tumour cells. The accumulation of mitochondrial mass and the optimization of mitochondrial quality accounted for the increased oxidative phosphorylation, and interruption of the mitochondrial fusion process downregulated the efficiency of oxidative phosphorylation and sensitized GBM cells to TMZ, which was also confirmed in the in vivo experiment. What is more, interfering with this process is an innovative strategy to overcome the chemo‐resistance of GBM cells.

Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells.

Glioblastoma multiforme (GBM) is a malignant primary brain tumor. The 5-year relative survival rate of patients with GBM remains <30% on average despite aggressive treatments, and secondary therapy fails in 90% of patients. In chemotherapeutic failure, detoxification proteins are crucial to the activity of chemotherapy drugs. Usually, glutathione S-transferase (GST) superfamily members act as detoxification enzymes by activating xenobiotic metabolites through conjugation with glutathione in healthy cells. However, some overexpressed GSTs not only increase GST activity but also trigger chemotherapy resistance and tumorigenesis-related signaling transductions. Whether GSTM3 is involved in GBM chemoresistance remains unclear. In the current study, we found that T98G, a GBM cell line with pre-existing temozolomide (TMZ) resistance, has high glycolysis and GSTM3 expression. GSTM3 knockdown in T98G decreased glycolysis ability through lactate dehydrogenase A activity reduction. Moreover, it increased TMZ toxicity and decreased invasion ability. Furthermore, we provide next-generation sequencing–based identification of significantly changed messenger RNAs of T98G cells with GSTM3 knockdown for further research. GSTM3 was downregulated in intrinsic TMZ-resistant T98G with a change in the expression levels of some essential glycolysis-related genes. Thus, GSTM3 was associated with glycolysis in chemotherapeutic resistance in T98G cells. Our findings provide new insight into the GSTM3 mechanism in recurring GBM.

Hydrogel matrix presence and composition influence drug responses of encapsulated glioblastoma spheroids

Glioblastoma multiforme (GBM) is the most aggressive brain tumor with median patient survival of 12–15 months. To facilitate treatment development, bioengineered GBM models that adequately recapitulate the in vivo tumor microenvironment are needed. Matrix-encapsulated multicellular spheroids represent such model because they recapitulate solid tumor characteristics, such as dimensionality, cell-cell, and cell-matrix interactions. Yet, there is no consensus as to which matrix properties are key to improving the predictive capacity of spheroid-based drug screening platforms. We used a hydrogel-encapsulated GBM spheroid model, where matrix properties were independently altered to investigate their effect on GBM spheroid characteristics and drug responsiveness. We focused on hydrogel degradability, tuned via enzymatically degradable crosslinkers, and hydrogel adhesiveness, tuned via integrin ligands. We observed increased cellular infiltration of GBM spheroids and increased resistance to temozolomide in degradable, adhesive hydrogels compared to spheroids in non-degradable, non-adhesive hydrogels or to free-floating spheroids. Further, a higher infiltration index was noted for spheroids in adhesive compared to non-adhesive degradable hydrogels. For spheroids in degradable hydrogels, we determined that infiltrating cells were more susceptible to temozolomide compared to cells in the spheroid core. The temozolomide susceptibility of the infiltrating cells was independent of integrin adhesion. We could not attribute differential drug responses to differential cellular proliferation or to limited drug penetration into the hydrogel matrix. Our results suggest that cell-matrix interactions guide GBM spheroid drug responsiveness and that further elucidation of these interactions could enable the engineering of more predictive drug screening platforms. Statement of significanceGlioblastoma multiforme (GBM) multicellular spheroids hold promise for drug screening and development as they better mimic in vivo cellular responses to therapeutics compared to monolayer cultures. Traditional spheroid models lack an external extracellular matrix (ECM) and fail to mimic the mechanical, physical, and biochemical cues seen in the GBM microenvironment. While embedding spheroids in hydrogel matrices has been shown to better recapitulate the tumor microenvironment, there is still limited understanding as to the key matrix properties that govern spheroid responsiveness to drugs. Here we decoupled and independently altered matrix properties such as degradability, via an enzymatically degradable peptide crosslinker, and cell adhesion, via an adhesive ligand, giving further insight into what matrix properties contribute to GBM chemoresistance.

Wnt/β-catenin Antagonists: Exploring New Avenues to Trigger Old Drugs in Alleviating Glioblastoma Multiforme.

Glioblastoma Multiforme (GBM) is one of the most heterogeneous primary brain tumors with high mortality. In spite of the current therapeutic approaches, the survival rate remains poor, with death occurring within 12 to 15 months after the preliminary diagnosis. This warrants the need for an effective treatment modality. The Wnt/β-catenin pathway is presumably the most noteworthy pathway upregulated in almost 80% of GBM cases, contributing to tumor initiation, progression, and survival. Therefore, therapeutic strategies targeting key components of the Wnt/β-catenin cascade using established genotoxic agents like temozolomide and pharmacological inhibitors would be an effective approach to modulate the Wnt/β-catenin pathway. Recently, drug repurposing by means of effective combination therapy has gained importance in various solid tumors, including GBM, by targeting two or more proteins in a single pathway, thereby possessing the ability to overcome the hurdle implicated by chemoresistance in GBM. In this context, by employing computational tools, an attempt has been made to find out the novel combinations against the Wnt/β-catenin signalling pathway. We have explored the binding interactions of three conventional drugs - namely temozolomide, metformin and chloroquine - along with three natural compounds, viz. epigallocatechin gallate, naringenin and phloroglucinol, on the major receptors of Wnt/β-catenin signalling. It was noted that all the experimental compounds showed profound interaction with two major receptors of the Wnt/β-catenin pathway. To the best of our knowledge, this study is the first of its kind to characterize the combined interactions of the aforementioned drugs with the Wnt/β-catenin signalling in silico, and this will putatively open up new avenues for combination therapies in GBM treatment.

Temozolomide Induces the Acquisition of Invasive Phenotype by O6-Methylguanine-DNA Methyltransferase (MGMT)+ Glioblastoma Cells in a Snail-1/Cx43-Dependent Manner

Glioblastoma multiforme (GBM) recurrences after temozolomide (TMZ) treatment result from the expansion of drug-resistant and potentially invasive GBM cells. This process is facilitated by O6-Methylguanine-DNA Methyltransferase (MGMT), which counteracts alkylating TMZ activity. We traced the expansion of invasive cell lineages under persistent chemotherapeutic stress in MGMTlow (U87) and MGMThigh (T98G) GBM populations to look into the mechanisms of TMZ-induced microevolution of GBM invasiveness. TMZ treatment induced short-term, pro-invasive phenotypic shifts of U87 cells, in the absence of Snail-1 activation. They were illustrated by a transient induction of their motility and followed by the hypertrophy and the signs of senescence in scarce U87 sub-populations that survived long-term TMZ stress. In turn, MGMThigh T98G cells reacted to the long-term TMZ treatment with the permanent induction of invasiveness. Ectopic Snail-1 down-regulation attenuated this effect, whereas its up-regulation augmented T98G invasiveness. MGMTlow and MGMThigh cells both reacted to the long-term TMZ stress with the induction of Cx43 expression. However, only in MGMThigh T98G populations, Cx43 was directly involved in the induction of invasiveness, as manifested by the induction of T98G invasiveness after ectopic Cx43 up-regulation and by the opposite effect after Cx43 down-regulation. Collectively, Snail-1/Cx43-dependent signaling participates in the long-term TMZ-induced microevolution of the invasive GBM front. High MGMT activity remains a prerequisite for this process, even though MGMT-related GBM chemoresistance is not necessary for its initiation.

Moschamindole induces glioma cell apoptosis by blocking Mia40-dependent mitochondrial intermembrane space assembly and oxidative respiration.

Glioblastoma multiforme (GBM) is the most frequent, lethal, and aggressive tumor of the central nervous system in adults. In this study, we found for the first time that moschamindole (MCD), a rare phenolic amide with 8/6/6/5/5 rings, is a major bioactive constituent derived from Phragmites communis Trin (Poaceae) that exhibits a potential cytotoxic effect on both TMZ-resistant GBM cell lines and xenograft models. MCD-induced intrinsic apoptosis signals and mitochondrial dysfunction were confirmed by cell cycle arrest, caspase-3/7 activation, and membrane potential depolarization. Furthermore, investigations exploring the mechanism showed that MCD specifically inhibits Mia40-mediated oxidative folding of mitochondrial intermembrane space (IMS) proteins via PCR assay and immunoblot analysis. MCD relies on its positive charge to associate with mitochondrial oxidative respiration, thus blocking energy metabolism and inducing apoptosis. Overexpression and upregulation of Mia40 were proven to reverse MCD-induced apoptosis and were correlated with the chemoresistance of GBM in vitro and in vivo, respectively. Taken together, our study demonstrates that Mia40 is a potential target of the chemoresistance of glioblastoma and suggests that MCD might be a potential agent for the individualized treatment of chemoresistant GBM based on mitochondrial metabolic characteristics and Mia40 expression.

The novel roles of virus infection-associated gene CDKN1A in chemoresistance and immune infiltration of glioblastoma.

Chemoresistance is a common limitation for successful treatment of glioblastoma multiforme (GBM). Recently, virus infections have been demonstrated to be associated with tumorigenesis and chemoresistance in tumors. However, the role of infection-related genes in GBM haven’t been clearly demonstrated. Here, we explored the roles and mechanisms of human T-lymphotropic virus type-1 (HTLV-1) infections in tumorigenesis and chemoresistance in GBM. Four candidate genes, CDKN1A, MSX1, MYC and CHEK2, were identified to be the codifferentially expressed genes between three temozolomide (TMZ) chemotherapy datasets and one HTLV-1 infection gene set. Next, comprehensive bioinformatics data from several databases indicated that only CDKN1A was significantly upregulated in both GBM tissues and cells and showed the greatest prognostic value in GBM patients. Clinical data identified the correlations between CDKN1A expression and clinicopathological parameters of GBM patients. Moreover, CDKN1A was found to be involved in AKT-mediated TMZ resistance of glioma cells. In addition, KEGG analysis of CDKN1A-associated coexpression genes showed that CDKN1A was potentially involved in complement and coagulation cascades pathways in GBM. Finally, TISIDB database was used to investigate the role of CDKN1A in tumor-immune system interactions in GBM. These findings enhanced our understanding of the roles of CDKN1A in tumorigenesis and therapy response in GBM.

Targeting the ABC transporter ABCB5 sensitizes glioblastoma to temozolomide-induced apoptosis through a cell-cycle checkpoint regulation mechanism

Glioblastoma multiforme (GBM) is a malignant brain tumor with a poor prognosis resulting from tumor resistance to anticancer therapy and a high recurrence rate. Compelling evidence suggests that this is driven by subpopulations of cancer stem cells (CSCs) with tumor-initiating potential. ABC subfamily B member 5 (ABCB5) has been identified as a molecular marker for distinct subsets of chemoresistant tumor-initiating cell populations in diverse human malignancies. In the current study, we examined the potential role of ABCB5 in growth and chemoresistance of GBM. We found that ABCB5 is expressed in primary GBM tumors, in which its expression was significantly correlated with the CSC marker protein CD133 and with overall poor survival. Moreover, ABCB5 was also expressed by CD133-positive CSCs in the established human U-87 MG, LN-18, and LN-229 GBM cell lines. Antibody- or shRNA-mediated functional ABCB5 blockade inhibited proliferation and survival of GBM cells and sensitized them to temozolomide (TMZ)-induced apoptosis in vitro Likewise, in in vivo human GBM xenograft experiments with immunodeficient mice, mAb treatment inhibited growth of mutant TP53, WT PTEN LN-229 tumors, and sensitized LN-229 tumors to TMZ therapy. Mechanistically, we demonstrate that ABCB5 blockade inhibits TMZ-induced G2/M arrest and augments TMZ-mediated cell death. Our results identify ABCB5 as a GBM chemoresistance marker and point to the potential utility of targeting ABCB5 to improve current GBM therapies.

Novel approaches to combat chemoresistance against glioblastomas

The poor prognosis of glioblastoma multiforme (GBM) patients is in part due to resistance to current standard-of-care treatments including chemotherapy [predominantly temozolomide (TMZ; Temodar)], radiation therapy and an anti-angiogenic therapy [an antibody against the vascular endothelial growth factor (bevacizumab; Avastin)], resulting in recurrent tumors. Several recurrent GBM tumors are commonly resistant to either TMZ, radiation or bevacizumab, which contributes to the low survival rate for GBM patients. This review will focus on novel targets and therapeutic approaches that are currently being considered to combat GBM chemoresistance. One of these therapeutic options is a small molecule called OKlahoma Nitrone 007 (OKN-007), which was discovered to inhibit the transforming growth factor β1 pathway, reduce TMZ-resistance and enhance TMZ-sensitivity. OKN-007 is currently an investigational new drug in clinical trials for both newly-diagnosed and recurrent GBM patients. Another novel target is ELTD1 (epidermal growth factor, latrophilin and seven transmembrane domain-containing protein 1; alternatively known as ADGRL4, Adhesion G protein-coupled receptor L4), which we used a monoclonal antibody against, where a therapy against it was found to inhibit Notch 1 in a pre-clinical GBM xenograft model. Notch 1 is known to be associated with chemoresistance in GBM. Other potential therapeutic targets to combat GBM chemoresistance include the phosphoinositide 3-kinase pathway, nuclear factor-κB, the hepatocyte/scatter factor (c-MET), the epidermal growth factor receptor, and the tumor microenvironment.

R406 elicits anti-Warburg effect via Syk-dependent and -independent mechanisms to trigger apoptosis in glioma stem cells

Given that glioma stem cells (GSCs) play a critical role in the initiation and chemoresistance in glioblastoma multiforme (GBM), targeting GSCs is an attractive strategy to treat GBM. Utilizing an anti-cancer compound library, we identified R406, the active metabolite of a FDA-approved Syk inhibitor for immune thrombocytopenia (ITP), with remarkable cytotoxicity against GSCs but not normal neural stem cells. R406 significantly inhibited neurosphere formation and triggered apoptosis in GSCs. R406 induced a metabolic shift from glycolysis to oxidative phosphorylation (OXPHOS) and subsequently production of excess ROS in GSCs. R406 also diminished tumor growth and efficiently sensitized gliomas to temozolomide in GSC-initiating xenograft mouse models. Mechanistically, the anti-GSC effect of R406 was due to the disruption of Syk/PI3K signaling in Syk-positive GSCs and PI3K/Akt pathway in Syk-negative GSCs respectively. Overall, these findings not only identify R406 as a promising GSC-targeting agent but also reveal the important role of Syk and PI3K pathways in the regulation of energy metabolism in GSCs.

Cell surface vimentin-targeted monoclonal antibody 86C increases sensitivity to temozolomide in glioma stem cells

Glioblastoma multiforme (GBM) is the most prevalent and aggressive brain tumor. The current standard therapy, which includes radiation and chemotherapy, is frequently ineffective partially because of drug resistance and poor penetration of the blood-brain barrier. Reducing resistance and increasing sensitivity to chemotherapy may improve outcomes. Glioma stem cells (GSCs) are a source of relapse and chemoresistance in GBM; sensitization of GSCs to temozoliomide (TMZ), the primary chemotherapeutic agent used to treat GBM, is therefore integral for therapeutic efficacy. We previously discovered a unique tumor-specific target, cell surface vimentin (CSV), on patient-derived GSCs. In this study, we found that the anti-CSV monoclonal antibody 86C efficiently increased GSC sensitivity to TMZ. The combination TMZ+86C induced significantly greater antitumor effects than TMZ alone in eight of 12 GSC lines. TMZ+86C–sensitive GSCs had higher CSV expression overall and faster CSV resurfacing among CSV− GSCs compared with TMZ+86C–resistant GSCs. Finally, TMZ+86C increased apoptosis of tumor cells and prolonged survival compared with either drug alone in GBM mouse models. The combination of TMZ+86C represents a promising strategy to reverse GSC chemoresistance.

Delivery of siRNA in vitro and in vivo using PEI-capped porous silicon nanoparticles to silence MRP1 and inhibit proliferation in glioblastoma

BackgroundMultidrug resistance-associated protein 1 (MRP1) overexpression plays a major role in chemoresistance in glioblastoma multiforme (GBM) contributing to its notorious deadly nature. Although MRP1-siRNA transfection to GBM in vitro has been shown to sensitise the cells to drug, MRP1 silencing in vivo and the phenotypic influence on the tumour and normal tissues upon MRP1 down-regulation have not been established. Here, porous silicon nanoparticles (pSiNPs) that enable high-capacity loading and delivery of siRNA are applied in vitro and in vivo.ResultWe established pSiNPs with polyethyleneimine (PEI) capping that enables high-capacity loading of siRNA (92 µg of siRNA/mg PEI-pSiNPs), and optimised release profile (70% released between 24 and 48 h). These pSiNPs are biocompatible, and demonstrate cellular uptake and effective knockdown of MRP1 expression in GBM by 30%. Also, siRNA delivery was found to significantly reduce GBM proliferation as an associated effect. This effect is likely mediated by the attenuation of MRP1 transmembrane transport, followed by cell cycle arrest. MRP1 silencing in GBM tumour using MRP1-siRNA loaded pSiNPs was demonstrated in mice (82% reduction at the protein level 48 h post-injection), and it also produced antiproliferative effect in GBM by reducing the population of proliferative cells. These results indicate that in vitro observations are translatable in vivo. No histopathological signs of acute damage were observed in other MRP1-expressing organs despite collateral downregulations.ConclusionsThis study proposes the potential of efficient MRP1-siRNA delivery by using PEI-capped pSiNPs in achieving a dual therapeutic role of directly attenuating the growth of GBM while sensitising residual tumour cells to the effects of chemotherapy post-resection.

Endoplasmic reticulum chaperone prolyl 4-hydroxylase, beta polypeptide (P4HB) promotes malignant phenotypes in glioma via MAPK signaling.

Endoplasmic reticulum (ER) chaperone Prolyl 4-hydroxylase, beta polypeptide (P4HB) has previously been identified as a novel target for chemoresistance in glioblastoma multiforme (GBM). Yet its functional roles in glioma carcinogenesis remain elusive. In clinical analysis using human glioma specimens and Gene Expression Omnibus (GEO) profiles, we found that aberrant expression of P4HB was correlated with high-grade malignancy and an angiogenic phenotype in glioma. Furthermore, P4HB upregulation conferred malignant characteristics including proliferation, invasion, migration and angiogenesis in vitro, and increased tumor growth in vivo via the mitogen-activated protein kinase (MAPK) signaling pathway. Pathway analysis suggested genetic and pharmacologic inhibition of P4HB suppressed MAPK expression and its downstream targets were involved in angiogenesis and invasion. This is the first study that demonstrates the oncogenic roles of P4HB and its underlying mechanism in glioma. Since tumor invasion and Vascularisation are typical hallmarks in malignant glioma, our findings uncover a promising anti-glioma mechanism through P4HB-mediated retardation of MAPK signal transduction.