Targeting the Endothelin-1 pathway to reduce invasion and chemoresistance in gallbladder cancer cells.

BACKGROUND: Gallbladder cancer (GBC) is a prevalent and deadly biliary tract carcinoma, often diagnosed at advanced stages with limited treatment options. The 5-year survival rate varies widely from 4 to 60%, mainly due to differences in disease stage detection. With only a small fraction of patients having resectable tumors and a high incidence of metastasis, advanced GBC stages are characterized by significant chemoresistance. Identification of new therapeutic targets is crucial, and recent studies have shown that the Endothelin-1 (ET-1) signaling pathway, involving ETAR and/or ETBR receptors (ETRs), plays a crucial role in promoting tumor aggressiveness in various cancer models. Blocking one or both receptors has been reported to reduce invasiveness and chemoresistance in cancers like ovarian, prostate, and colon. Furthermore, transcriptomic studies have associated ET-1 levels with late stages of GBC; however, it remains unclear whether its signaling or its inhibition has implications for its aggressiveness. Although the role of ET-1 signaling in gallbladder physiology is minimally understood, its significance in other tumor models leads us to hypothesize its involvement in GBC malignancy. RESULTS: In this study, we investigated the expression of ET-1 pathway proteins in three GBC cell lines and a primary GBC culture. Our findings demonstrated that both ETAR and ETBR receptors are expressed in GBC cells and tumor samples. Moreover, we successfully down-regulated ET-1 signaling using a non-selective ETR antagonist, Macitentan, which resulted in reduced migratory and invasive capacities of GBC cells. Additionally, Macitentan treatment chemosensitized the cells to Gemcitabine, a commonly used therapy for GBC. CONCLUSION: For the first time, we reveal the role of the ET-1 pathway in GBC cells, providing insight into the potential therapeutic targeting of its receptors to mitigate invasion and chemoresistance in this cancer with limited treatment options. These findings pave the way for further exploration of Macitentan or other ETR antagonists as potential therapeutic strategies for GBC management. In summary, our study represents a groundbreaking contribution to the field by providing the first evidence of the ET 1 pathway's pivotal role in modulating the behavior and aggressiveness of GBC cells, shedding new light on potential therapeutic targets.

Co-Expression of Immunohistochemical Markers MRP2, CXCR4, and PD-L1 in Gallbladder Tumors Is Associated with Prolonged Patient Survival.

Gallbladder cancer (GBC) is a rare pathology in Western countries. However, it constitutes a relevant health problem in Asia and Latin America, with a high mortality in middle-aged Chilean women. The limited therapeutic options for GBC require the identification of targetable proteins with prognostic value for improving clinical management support. We evaluated the expression of targetable proteins, including three epithelial tumor markers, four proteins associated with multidrug and apoptosis resistance, and eleven immunological markers in 241 primary gallbladder adenocarcinomas. We investigated correlations between tumor marker expression, the primary tumor staging, and GBC patients' survival using automated immunohistochemistry, a semi-automatic method for image analysis, univariate and multivariate statistical analyses, and machine learning algorithms. Our data show a significant association between the expression of MRP2 (p = 0.0028), CXCR4 (p = 0.0423), and PD-L1 (p = 0.0264), and a better prognosis for patients with late-stage primary tumors. The expression of the MRP2/CXCR4/PD-L1 cluster of markers discriminates among short-, medium-, and long-term patient survival, with an ROC of significant prognostic value (AUC = 0.85, p = 0.0012). Moreover, a high MRP2/CXCR4/PD-L1 co-expression is associated with increased survival time (30 vs. 6 months, p = 0.0025) in GBC patients, regardless of tumor stage. Hence, our results suggest that the MRP2/CXCR4/PD-L1 cluster could potentially be a prognostic marker for GBC.

Glioblastoma Microenvironment and Invasiveness: New Insights and Therapeutic Targets.

Glioblastoma (GBM) is the most common and malignant primary brain cancer in adults. Without treatment the mean patient survival is approximately 6 months, which can be extended to 15 months with the use of multimodal therapies. The low effectiveness of GBM therapies is mainly due to the tumor infiltration into the healthy brain tissue, which depends on GBM cells' interaction with the tumor microenvironment (TME). The interaction of GBM cells with the TME involves cellular components such as stem-like cells, glia, endothelial cells, and non-cellular components such as the extracellular matrix, enhanced hypoxia, and soluble factors such as adenosine, which promote GBM's invasiveness. However, here we highlight the role of 3D patient-derived glioblastoma organoids cultures as a new platform for study of the modeling of TME and invasiveness. In this review, the mechanisms involved in GBM-microenvironment interaction are described and discussed, proposing potential prognosis biomarkers and new therapeutic targets.

Cancer Stem Cell and Aggressiveness Traits Are Promoted by Stable Endothelin-Converting Enzyme-1c in Glioblastoma Cells.

Glioblastoma (GBM) is the most common and aggressive type of brain tumor due to its elevated recurrence following treatments. This is mainly mediated by a subpopulation of cells with stemness traits termed glioblastoma stem-like cells (GSCs), which are extremely resistant to anti-neoplastic drugs. Thus, an advancement in the understanding of the molecular processes underlying GSC occurrence should contribute significantly towards progress in reducing aggressiveness. High levels of endothelin-converting enzyme-1 (ECE1), key for endothelin-1 (ET-1) peptide activation, have been linked to the malignant progression of GBM. There are four known isoforms of ECE1 that activate ET-1, which only differ in their cytoplasmic N-terminal sequences. Isoform ECE1c is phosphorylated at Ser-18 and Ser-20 by protein kinase CK2, which increases its stability and hence promotes aggressiveness traits in colon cancer cells. In order to study whether ECE1c exerts a malignant effect in GBM, we designed an ECE1c mutant by switching a putative ubiquitination lysine proximal to the phospho-serines Lys-6-to-Arg (i.e., K6R). This ECE1cK6R mutant was stably expressed in U87MG, T98G, and U251 GBM cells, and their behavior was compared to either mock or wild-type ECE1c-expressing clone cells. ECE1cK6R behaved as a highly stable protein in all cell lines, and its expression promoted self-renewal and the enrichment of a stem-like population characterized by enhanced neurospheroid formation, as well as increased expression of stem-like surface markers. These ECE1cK6R-derived GSC-like cells also displayed enhanced resistance to the GBM-related chemotherapy drugs temozolomide and gemcitabine and increased expression of the ABCG2 efflux pump. In addition, ECE1cK6R cells displayed enhanced metastasis-associated traits, such as the modulation of adhesion and the enhancement of cell migration and invasion. In conclusion, the acquisition of a GSC-like phenotype, together with heightened chemoresistance and invasiveness traits, allows us to suggest phospho-ECE1c as a novel marker for poor prognosis as well as a potential therapeutic target for GBM.

The low affinity A2B adenosine receptor enhances migratory and invasive capacity in vitro and angiogenesis in vivo of glioblastoma stem-like cells.

Glioblastoma (GBM) is the most common and deadly malignant brain tumor, with a median survival of 15 to 17 months for a patient. GBM contains a cellular subpopulation known as GBM stem-like cells (GSCs) that persist in hypoxic niches and are capable of infiltrating into healthy brain tissue. For this reason, GSCs are considered one of the main culprits for GBM recurrence. A hypoxic microenvironment increases extracellular adenosine levels, activating the low affinity A2B adenosine receptor (A2BAR). Adenosine, through A2BAR, is capable of modulating invasiveness. However, its role in the invasion/migration of hypoxic-GSCs is still unknown. This study aims to understand the importance of A2BAR in modulating the migratory/invasive capacity of GSCs under hypoxia. Data analysis from The Cancer Genome Atlas (TCGA) program correlates A2BAR expression with high-grade glioma and hypoxic necrotic areas. U87MG and primary culture-derived GSCs under hypoxic conditions (0.5% O2) increased A2BAR mRNA and protein levels. As expected, the migratory and invasive capacity of GSCs increased under hypoxia, which was counteracted by blocking A2BAR, through the downregulation of MMP9 activity and epithelial-mesenchymal transition marker expression. Finally, in a xenograft mouse model, we demonstrate that treatment with MRS1754 did not affect the tumor volume but could decrease blood vessel formation and VEGF expression. Our results suggest that extracellular adenosine, through the activation of A2BAR, enhances the migratory and invasive capacity of GSCs in vitro under hypoxic conditions. Targeting A2BAR can be an effective therapy for GBM recurrence.

A2B Adenosine Receptor Enhances Chemoresistance of Glioblastoma Stem-Like Cells under Hypoxia: New Insights into MRP3 Transporter Function.

Glioblastoma is the most common and aggressive primary brain tumor, characterized by its high chemoresistance and the presence of a cell subpopulation that persists under hypoxic niches, called glioblastoma stem-like cells (GSCs). The chemoresistance of GSCs is mediated in part by adenosine signaling and ABC transporters, which extrude drugs outside the cell, such as the multidrug resistance-associated proteins (MRPs) subfamily. Adenosine promotes MRP1-dependent chemoresistance under normoxia. However, adenosine/MRPs-dependent chemoresistance under hypoxia has not been studied until now. Transcript and protein levels were determined by RT-qPCR and Western blot, respectively. MRP extrusion capacity was determined by intracellular 5 (6)-Carboxyfluorescein diacetate (CFDA) accumulation. Cell viability was measured by MTS assays. Cell cycle and apoptosis were determined by flow cytometry. Here, we show for the first time that MRP3 expression is induced under hypoxia through the A2B adenosine receptor. Hypoxia enhances MRP-dependent extrusion capacity and the chemoresistance of GSCs. Meanwhile, MRP3 knockdown decreases GSC viability under hypoxia. Downregulation of the A2B receptor decreases MRP3 expression and chemosensibilizes GSCs treated with teniposide under hypoxia. These data suggest that hypoxia-dependent activation of A2B adenosine receptor promotes survival of GSCs through MRP3 induction.

Adapt to Persist: Glioblastoma Microenvironment and Epigenetic Regulation on Cell Plasticity.

Glioblastoma (GBM) is the most frequent and aggressive brain tumor, characterized by great resistance to treatments, as well as inter- and intra-tumoral heterogeneity. GBM exhibits infiltration, vascularization and hypoxia-associated necrosis, characteristics that shape a unique microenvironment in which diverse cell types are integrated. A subpopulation of cells denominated GBM stem-like cells (GSCs) exhibits multipotency and self-renewal capacity. GSCs are considered the conductors of tumor progression due to their high tumorigenic capacity, enhanced proliferation, invasion and therapeutic resistance compared to non-GSCs cells. GSCs have been classified into two molecular subtypes: proneural and mesenchymal, the latter showing a more aggressive phenotype. Tumor microenvironment and therapy can induce a proneural-to-mesenchymal transition, as a mechanism of adaptation and resistance to treatments. In addition, GSCs can transition between quiescent and proliferative substates, allowing them to persist in different niches and adapt to different stages of tumor progression. Three niches have been described for GSCs: hypoxic/necrotic, invasive and perivascular, enhancing metabolic changes and cellular interactions shaping GSCs phenotype through metabolic changes and cellular interactions that favor their stemness. The phenotypic flexibility of GSCs to adapt to each niche is modulated by dynamic epigenetic modifications. Methylases, demethylases and histone deacetylase are deregulated in GSCs, allowing them to unlock transcriptional programs that are necessary for cell survival and plasticity. In this review, we described the effects of GSCs plasticity on GBM progression, discussing the role of GSCs niches on modulating their phenotype. Finally, we described epigenetic alterations in GSCs that are important for stemness, cell fate and therapeutic resistance.

Phosphorylation of Endothelin-Converting Enzyme-1c at Serines 18 and 20 by CK2 Promotes Aggressiveness Traits in Colorectal Cancer Cells.

Endothelin-converting enzyme-1 (ECE1) activates the endothelin-1 peptide, which upregulates pathways that are related to diverse hallmarks of cancer. ECE1 is expressed as four isoforms differing in their N-terminal domains. Protein kinase CK2 phosphorylates the N-terminus of isoform ECE1c, enhancing its stability and promoting invasiveness of colorectal cancer cells. However, the specific residues in ECE1c that are phosphorylated by CK2 and how this phosphorylation promotes invasiveness was unknown. Here we demonstrate that Ser-18 and Ser-20 are the bona fide residues phosphorylated by CK2 in ECE1c. Thus, biphospho-mimetic ECE1cDD and biphospho-resistant ECE1cAA mutants were constructed and stably expressed in different colorectal cancer cells through lentiviral transduction. Biphospho-mimetic ECE1cDD displayed the highest stability in cells, even in the presence of the specific CK2 inhibitor silmitasertib. Concordantly, ECE1cDD-expressing cells showed enhanced hallmarks of cancer, such as proliferation, migration, invasiveness, and self-renewal capacities. Conversely, cells expressing the less-stable biphospho-resistant ECE1cAA showed a reduction in these features, but also displayed an important sensitization to 5-fluorouracil, an antineoplastic agent traditionally used as therapy in colorectal cancer patients. Altogether, these findings suggest that phosphorylation of ECE1c at Ser-18 and Ser-20 by CK2 promotes aggressiveness in colorectal cancer cells. Therefore, phospho-ECE1c may constitute a novel biomarker of poor prognosis and CK2 inhibition may be envisioned as a potential therapy for colorectal cancer patients.

Endothelin-converting enzyme-1c promotes stem cell traits and aggressiveness in colorectal cancer cells.

Endothelin-1 is a mitogenic peptide that activates several proliferation, survival, and invasiveness pathways. The effects of endothelin-1 rely on its activation by endothelin-converting enzyme-1 (ECE1), which is expressed as four isoforms with different cytoplasmic N termini. Recently, isoform ECE1c has been suggested to have a role in cancer aggressiveness. The N terminus of ECE1c is phosphorylated by protein kinase CK2 (also known as casein kinase 2), and this enhances its stability and promotes invasiveness in colorectal cancer cells. However, it is not known how phosphorylation improves stability and why this is correlated with increased aggressiveness. We hypothesized that CK2 phosphorylation protects ECE1c from N-terminal ubiquitination and, consequently, from proteasomal degradation. Here, we show that lysine 6 is the bona fide residue involved in ubiquitination of ECE1c and its mutation to arginine (ECE1cK6R ) significantly impairs proteasomal degradation, thereby augmenting ECE1c stability, even in the presence of the CK2 inhibitor silmitasertib. Furthermore, colorectal cancer cells overexpressing ECE1cK6R displayed enhanced cancer stem cell (CSC) traits, including increased stemness gene expression, chemoresistance, self-renewal, and colony formation and spheroid formation in vitro, as well as enhanced tumor growth and metastasis in vivo. These findings suggest that CK2-dependent phosphorylation enhances ECE1c stability, promoting an increase in CSC-like traits. Therefore, phospho-ECE1c may be a biomarker of poor prognosis and a potential therapeutic target for colorectal cancer.

Adenosine Depletion as A New Strategy to Decrease Glioblastoma Stem-Like Cells Aggressiveness.

Glioblastoma is the brain tumor with the worst prognosis. This is mainly due to a cell subpopulation with an extremely aggressive potential, called glioblastoma stem-like cells (GSCs). These cells produce high levels of extracellular adenosine, which are increased even more under hypoxic conditions. Under hypoxia, adenosine signaling is related to HIF-2α expression, enhancing cell aggressiveness. Adenosine can be degraded using recombinant adenosine deaminase (ADA) to revert its pathological effects. The aim of this study was to degrade adenosine using ADA in order to decrease malignancy of GSCs. Adenosine depletion was performed using recombinant ADA. Migration and invasion were measured by transwell and matrigel-coated transwell assay, respectively. HIF-2α-dependent cell migration/invasion decreased in GSCs treated with ADA under hypoxia. MRPs-mediated chemoresistance and colony formation decreased in treatment with ADA. In conclusion, adenosine depletion using adenosine deaminase decreases GSCs aggressiveness.

Endothelin-converting enzyme-1 in cancer aggressiveness.

The endothelin-1 (ET-1) axis contributes to the pathophysiology of several cancers by promoting tumor development and progression. This peptide is activated from its precursor, big ET-1, by endothelin-converting enzyme-1 (ECE-1). Active ET-1 binds to its cognate G-coupled receptor, ETAR, which transduces the signal to the inside of the cell. ET-1 has a short half-life of about 90 s, so its biological effects are completely dependent on its enzymatic activation by ECE-1. Expression of ECE-1 is elevated in several tumors and cancer cell lines. There are four ECE-1 isoforms -ECE-1a, -1b, -1c, and -1d- which vary in terms of their subcellular localization and, in some cases, their effects on cancer-related properties such as proliferation and invasiveness. In this article, we review findings on the role of ECE-1, particularly isoform ECE-1c, in oncogenesis and malignant progression. We also review evidence regarding ECE-1 expression in several types of tumors and cancer cell lines. Recent findings from our laboratory and others allow us to speculate on the mechanism by which ECE-1c promotes cancer aggressiveness. Finally, we evaluate potential post-translational modifications of ECE-1c, highlighting phosphorylation by several kinases, as well as evidence pointing to a putative, non-canonical, ET-1-independent mechanism for promoting invasiveness. Taken together, current evidence suggests that ECE-1c contributes to cancer aggressiveness and plays a putative role as a key regulator of cancer progression. Therefore, we propose that this protein is a promising target for prognostic and therapeutic purposes.

Glioma progression in diabesity.

Diabetes mellitus, obesity, and cancer are diseases that in recent years have caused a large number of deaths worldwide, so have been in the front line of biomedical research. On the other hand, obesity is a risk factor for several types of cancer and type 2 diabetes mellitus. The metabolic disorder and global inflammatory environment seen in obese patients is also critical for the treatment of both diabetes mellitus and gliomas. Several molecules are increased in patients with obesity and are considered risk factors in the failure of multimodal therapies for diabetes mellitus and gliomas. These molecules include adenosine, insulin, adenosine deaminases, adenosine kinase, lipids, as well as adenosine receptors, adenosine membrane transporters, and the immune response. The role of adenosine will be explained in depth since it is a nucleoside aberrantly increased in patients with these diseases, is one of the main causes of diabetes mellitus progression and the failure of glioma therapies. In addition, the role of type 2 diabetes mellitus/obesity, i.e., diabesity, and its implication in glioma treatment is discussed.

CK2 inhibition with silmitasertib promotes methuosis-like cell death associated to catastrophic massive vacuolization of colorectal cancer cells.

Protein kinase CK2 is a highly conserved and constitutively active Ser/Thr-kinase that phosphorylates a large number of substrates, resulting in increased cell proliferation and survival. A known target of CK2 is Akt, a player in the PI3K/Akt/mTORC1 signaling pathway, which is aberrantly activated in 32% of colorectal cancer (CRC) patients. On the other hand, mTORC1 plays an important role in the regulation of protein synthesis, cell growth, and autophagy. Some studies suggest that CK2 regulates mTORC1 in several cancers. The most recently developed CK2 inhibitor, silmitasertib (formerly CX-4945), has been tested in phase I/II trials for cholangiocarcinoma and multiple myeloma. This drug has been shown to induce autophagy and enhance apoptosis in pancreatic cancer cells and to promote apoptosis in non-small cell lung cancer cells. Nevertheless, it has not been tested in studies for CRC patients. We show in this work that inhibition of CK2 with silmitasertib decreases in vitro tumorigenesis of CRC cells in response to G2/M arrest, which correlates with mTORC1 inhibition and formation of large cytoplasmic vacuoles. Notably, molecular markers indicate that these vacuoles derive from massive macropinocytosis. Altogether, these findings suggest that an aberrantly elevated expression/activity of CK2 may play a key role in CRC, promoting cell viability and proliferation in untreated cells, however, its inhibition with silmitasertib promotes methuosis-like cell death associated to massive catastrophic vacuolization, accounting for decreased tumorigenicity at later times. These characteristics of silmitasertib support a potential therapeutic use in CRC patients and probably other CK2-dependent cancers.

Extracellular adenosine promotes cell migration/invasion of Glioblastoma Stem-like Cells through A3 Adenosine Receptor activation under hypoxia.

Glioblastoma (GBM) is the brain tumor with the worst prognosis composed of a cell subpopulation called Glioblastoma Stem-like Cells (GSCs) responsible for tumor recurrence mediated by cell invasion. GSCs persist in a hypoxic microenvironment which promotes extracellular adenosine production and activation of the A3 Adenosine Receptor (A3AR), therefore, the aim of this study was to determine the role of extracellular adenosine and A3AR on GSCs invasion under hypoxia. GSCs were obtained from a U87MG cell line and primary cultures of GBM patients, and then incubated under normoxia or hypoxia. Gene expression was evaluated by RNAseq, RT-qPCR, and western blot. Cell migration was measured by spreading and transwell boyden chamber assays; cell invasion was evaluated by Matrigel-coated transwell, ex vivo brain slice, and in vivo xenograft assays. The contribution of A3AR on cell migration/invasion was evaluated using the A3AR antagonist, MRS1220. Extracellular adenosine production was higher under hypoxia than normoxia, mainly by the catalytic action of the prostatic acid phosphatase (PAP), promoting cell migration/invasion in a HIF-2-dependent process. A3AR blockade decreased cell migration/invasion and the expression of Epithelial-Mesenchymal Transition markers. In conclusion, high levels of extracellular adenosine production enhance cell migration/invasion of GSCs, through HIF-2/PAP-dependent activation of A3AR under hypoxia.

FK506 Attenuates the MRP1-Mediated Chemoresistant Phenotype in Glioblastoma Stem-Like Cells.

Poor response to current treatments for glioblastoma has been attributed to the presence of glioblastoma stem-like cells (GSCs). GSCs are able to expel antitumor drugs to the extracellular medium using the multidrug resistance-associated protein 1 (MRP1) transporter. Tacrolimus (FK506) has been identified as an MRP1 regulator in differentiated glioblastoma (GBM) cells (non-GSCs); however, the effect of FK506 on GSCs is currently unknown. The objective of the following research is to evaluate the effect of FK506 on the MRP1-related chemo-resistant phenotype of GSCs. For this, U87MG and C6 glioma cell lines were used to generate non-GSCs and GSCs. mRNA and MRP1-positive cells were evaluated by RT-qPCR and flow cytometry, respectively. A Carboxyfluorescein Diacetate (CFDA)-retention assay was performed to evaluate the MRP1 activity. Apoptosis and MTT assays were employed to evaluate the cytotoxic effects of FK506 plus Vincristine (MRP1 substrate). GSC-derived subcutaneous tumors were generated to evaluate the in vivo effect of FK506/Vincristine treatment. No differences in transcript levels and positive cells for MRP1 were observed in FK506-treated cells. Lesser cell viability, increased apoptosis, and CFDA-retention in the FK506/Vincristine-treated cells were observed. In vivo, the FK506/Vincristine treatment decreased the tumor size as well as ki67, Glial Fibrillary Acidic Protein (GFAP), and nestin expression. We conclude that FK506 confers a chemo-sensitive phenotype to MRP1-drug substrate in GSCs.

Current natural therapies in the treatment against glioblastoma.

Glioblastoma (GBM) is the most common and aggressive brain tumor, which causes the highest number of deaths worldwide. It is a highly vascularized tumor, infiltrative, and its tumorigenic capacity is exacerbated. All these hallmarks are therapeutic targets in GBM treatment, including surgical removal followed by radiotherapy and chemotherapy. Current therapies have not been sufficient for the effective patient's management, so the classic therapies have had to expand and incorporate new alternative treatments, including natural compounds. This review summarizes natural products and their physiological effects in in vitro and in vivo models of GBM, specifically by modulating signaling pathways involved in angiogenesis, cell migration/invasion, cell viability, apoptosis, and chemoresistance. The most important aspects of natural products and their derivatives were described in relation to its antitumoral effects. As a final result, it can be obtained that within the compounds with more evidence that supports or suggests its clinical use are the cannabinoids, terpenes, and curcumin, because many have been shown to have a significant effect in decreasing the progress of GBM through known mechanisms, such as chemo-sensitization or decrease migration and cell invasion. Natural compounds emerge as promising therapies to attack the progress of GBM.

The Adenosine A3 Receptor Regulates Differentiation of Glioblastoma Stem-Like Cells to Endothelial Cells under Hypoxia.

Glioblastoma (GBM) is a neoplasm characterized by an extensive blood vessel network. Hypoxic niches of GBM can induce tumorigenic properties of a small cell subpopulation called Glioblastoma stem-like cells (GSCs) and can also increase extracellular adenosine generation which activates the A3 adenosine receptor (A3AR). Moreover, GSCs potentiates the persistent neovascularization in GBM. The aim of this study was to determine if A3AR blockade can reduce the vasculogenesis mediated by the differentiation of GSCs to Endothelial Cells (ECs) under hypoxia. We evaluated the expression of endothelial cell markers (CD31, CD34, CD144, and vWF) by fluorescence-activated cell sorting (FACS), and vascular endothelial growth factor (VEGF) secretion by ELISA using MRS1220 (A3AR antagonist) under hypoxia. We validate our results using U87MG-GSCs A3AR knockout (GSCsA3-KO). The effect of MRS1220 on blood vessel formation was evaluated in vivo using a subcutaneous GSCs-tumor model. GSCs increased extracellular adenosine production and A3AR expression under hypoxia. Hypoxia also increased the percentage of GSCs positive for endothelial cell markers and VEGF secretion, which was in turn prevented when using MRS1220 and in GSCsA3-KO. Finally, in vivo treatment with MRS1220 reduced tumor size and blood vessel formation. Blockade of A3AR decreases the differentiation of GSCs to ECs under hypoxia and in vivo blood vessel formation.

Role of extracellular vesicles in glioma progression.

The role of extracellular vesicles in cancer biology has emerged as a focus of the study of great importance and has been shown to directly influence tumour development in several cancers including brain tumours, such as gliomas. Gliomas are the most aggressive brain tumours, and in the last time, a considerable effort has been made to understand their biology. Studies focus in the signalling pathways involved in the processes of angiogenesis, viability, drug resistance and immune response evasion, as well as gliomas ability to infiltrate healthy tissue, a phenomenon regulated by the migratory and invasive capacity of the cells within a tumour. In this review, we summarize the different types and classifications of extracellular vesicles, their intravesicular content, and their role in the regulation of tumour progression processes in glioma.

Current Therapeutic Alternatives and New Perspectives in Glioblastoma Multiforme.

BACKGROUND: In the last two decades, there have been significant technological advances in the early detection of brain tumors. However, no notable improvements have been observed in the treatment of Glioblastoma Multiforme (GBM), the most common brain neoplasm coupled with the worst prognosis. GBM is characterized by an extensive resistance to a broad spectrum of anti-tumor drugs. This property is the result of a phenomenon known as Multiple Drug Resistance (MDR), which significantly limits noninvasive alternative therapies. This limitation is primarily due to the activity of ABC transporters and proteins related with DNA repair such as the MGMT enzyme. Due to the high mortality rate in GBM patients and current treatment deficits, new therapeutic strategies for this type of neoplasm are of vital importance. METHODS: In this review, proposed treatments for GBM, including the use of alkylating agents with MGMT inhibitors, MDR modulators, and immunotherapies are discussed. We focused our bibliographic research on papers containing in vitro, in vivo, and clinical phase analysis published over the last 20 years. RESULTS: Several studies have demonstrated good results using alkylating agents plus MGMT inhibitors, although without great improvements in survival. The use of modulators of ABC transporters enhances the effects of chemotherapy, proving it an effective complementary therapy. Immunotherapies have undergone significant developments as a directed and personalized approach for GBM treatment. CONCLUSION: The use of alternative complementary therapies discussed in this review could increase the survival of GBM patients; however, additional clinical phase analysis and the generation of new treatment protocols are required.