KCa3.1 inhibition switches the phenotype of glioma-infiltrating microglia/macrophages

Among the strategies adopted by glioma to successfully invade the brain parenchyma is turning the infiltrating microglia/macrophages (M/MΦ) into allies, by shifting them toward an anti-inflammatory, pro-tumor phenotype. Both glioma and infiltrating M/MΦ cells express the Ca(2+)-activated K(+) channel (KCa3.1), and the inhibition of KCa3.1 activity on glioma cells reduces tumor infiltration in the healthy brain parenchyma. We wondered whether KCa3.1 inhibition could prevent the acquisition of a pro-tumor phenotype by M/MΦ cells, thus contributing to reduce glioma development. With this aim, we studied microglia cultured in glioma-conditioned medium or treated with IL-4, as well as M/MΦ cells acutely isolated from glioma-bearing mice and from human glioma biopsies. Under these different conditions, M/MΦ were always polarized toward an anti-inflammatory state, and preventing KCa3.1 activation by 1-[(2-Chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34), we observed a switch toward a pro-inflammatory, antitumor phenotype. We identified FAK and PI3K/AKT as the molecular mechanisms involved in this phenotype switch, activated in sequence after KCa3.1. Anti-inflammatory M/MΦ have higher expression levels of KCa3.1 mRNA (kcnn4) that are reduced by KCa3.1 inhibition. In line with these findings, TRAM-34 treatment, in vivo, significantly reduced the size of tumors in glioma-bearing mice. Our data indicate that KCa3.1 channels are involved in the inhibitory effects exerted by the glioma microenvironment on infiltrating M/MΦ, suggesting a possible role as therapeutic targets in glioma

Immune factors preceding diagnosis of glioma: a Prostate Lung Colorectal Ovarian Cancer Screening Trial nested case-control study.

BackgroundEpidemiological studies of adult glioma have identified genetic and environmental risk factors, but much remains unclear. The aim of the current study was to evaluate anthropometric, disease-related, and prediagnostic immune-related factors for relationship with glioma risk.MethodsWe conducted a nested case-control study among the intervention arm of the Prostate, Lung, Colorectal, and Ovarian Cancer (PLCO) Screening Trial. One hundred and twenty-four glioma cases were identified and each matched to four controls. Baseline characteristics were collected at enrollment and were evaluated for association with glioma status. Serum specimens were collected at yearly intervals and were analyzed for immune-related factors including TGF-β1, TNF-α, total IgE, and allergen-specific IgE. Immune factors were evaluated at baseline in a multivariate conditional logistic regression model, along with one additional model that incorporated the latest available measurement.ResultsA family history of glioma among first-degree relatives was associated with increased glioma risk (OR = 4.41, P = .002). In multivariate modeling of immune factors at baseline, increased respiratory allergen-specific IgE was inversely associated with glioma risk (OR for allergen-specific IgE > 0.35 PAU/L: 0.59, P = .03). A logistic regression model that incorporated the latest available measurements found a similar association for allergen-specific IgE (P = .005) and showed that elevated TGF-β1 was associated with increased glioma risk (P-value for trend <.0001).ConclusionThe results from this prospective prediagnostic study suggest that several immune-related factors are associated with glioma risk. The association observed for TGF-β1 when sampling closer to the time of diagnosis may reflect the nascent brain tumor's feedback on immune function

Differential sensitivity of Glioma stem cells to Aurora kinase A inhibitors: implications for stem cell mitosis and centrosome dynamics

Glioma stem-cell-like cells are considered to be responsible for treatment resistance and tumour recurrence following chemo-radiation in glioblastoma patients, but specific targets by which to kill the cancer stem cell population remain elusive. A characteristic feature of stem cells is their ability to undergo both symmetric and asymmetric cell divisions. In this study we have analysed specific features of glioma stem cell mitosis. We found that glioma stem cells appear to be highly prone to undergo aberrant cell division and polyploidization. Moreover, we discovered a pronounced change in the dynamic of mitotic centrosome maturation in these cells. Accordingly, glioma stem cell survival appeared to be strongly dependent on Aurora A activity. Unlike differentiated cells, glioma stem cells responded to moderate Aurora A inhibition with spindle defects, polyploidization and a dramatic increase in cellular senescence, and were selectively sensitive to Aurora A and Plk1 inhibitor treatment. Our study proposes inhibition of centrosomal kinases as a novel strategy to selectively target glioma stem cells

Selective Targeting to Glioma with Nucleic Acid Aptamers

Malignant glioma is characterised by a rapid growth rate and high capacity for invasive infiltration to surrounding brain tissue; hence, diagnosis and treatment is difficult and patient survival is poor. Aptamers contribute a promising and unique technology for the in vitro imaging of live cells and tissues, with a potentially bright future in clinical diagnostics and therapeutics for malignant glioma. The binding selectivity, uptake capacity and binding target of two DNA aptamers, SA43 and SA44, were investigated in glioma cells and patient tissues. The binding assay showed that SA43 and SA44 bound with strong affinity (Kd, 21.56 ± 4.60 nM and Kd, 21.11 ± 3.30 nM respectively) to the target U87MG cells. Quantitative analysis by flow cytometry showed that the aptamers were able to actively internalise in U87MG and 1321N1 glioma cells compared to the non-cancerous and non-glioma cell types. Confocal microscopy confirmed staining in the cytoplasm, and co-localisation studies with endoplasmic reticulum, Golgi apparatus and lysosomal markers suggested internalisation and compartmentalisation within the endomembrane system. Both aptamers selectively bound to Ku 70 and Ku 80 DNA repair proteins as determined by aptoprecipitation (AP) followed by mass spectrometry analysis and confirmation by Western blot. In addition, aptohistochemical (AHC) staining on paraffin embedded, formalin fixed patient tissues revealed that the binding selectivity was significantly higher for SA43 aptamer in glioma tissues (grade I, II, III and IV) compared to the non-cancerous tissues, whereas SA44 did not show selectivity towards glioma tissues. The results indicate that SA43 aptamer can differentiate between glioma and non-cancerous cells and tissues and therefore, shows promise for histological diagnosis of glioma

KCa3.1 channel inhibition sensitizes malignant gliomas to temozolomide treatment

Malignant gliomas are among the most frequent and aggressive cerebral tumors, characterized by high proliferative and invasive indexes. Standard therapy for patients, after surgery and radiotherapy, consists of temozolomide (TMZ), a methylating agent that blocks tumor cell proliferation. Currently, there are no therapies aimed at reducing tumor cell invasion. Ion channels are candidate molecular targets involved in glioma cell migration and infiltration into the brain parenchyma. In this paper we demonstrate that: i) blockade of the calcium-activated potassium channel KCa3.1 with TRAM-34 has co-adjuvant effects with TMZ, reducing GL261 glioma cell migration, invasion and colony forming activity, increasing apoptosis, and forcing cells to pass the G2/M cell cycle phase, likely through cdc2 de-phosphorylation; ii) KCa3.1 silencing potentiates the inhibitory effect of TMZ on glioma cell viability; iii) the combination of TMZ/TRAM-34 attenuates the toxic effects of glioma conditioned medium on neuronal cultures, through a microglia dependent mechanism since the effect is abolished by clodronate-induced microglia killing; iv) TMZ/TRAM-34 co-treatment increases the number of apoptotic tumor cells, and the mean survival time in a syngeneic mouse glioma model (C57BL6 mice implanted with GL261 cells); v) TMZ/TRAM-34 co-treatment reduces cell viability of GBM cells and cancer stem cells (CSC) freshly isolated from patients.Taken together, these data suggest a new therapeutic approach for malignant glioma, targeting both glioma cell proliferating and migration, and demonstrate that TMZ/TRAM-34 co-treatment affects both glioma cells and infiltrating microglia, resulting in an overall reduction of tumor cell progression

Glioma Diagnosis Aid through CNNs and Fuzzy-C Means for MRI

Glioma is a type of brain tumor that causes mortality in many cases. Early diagnosis is an important factor. Typically, it is detected through MRI and then either a treatment is applied, or it is removed through surgery. Deep-learning techniques are becoming popular in medical applications and image-based diagnosis. Convolutional Neural Networks are the preferred architecture for object detection and classification in images. In this paper, we present a study to evaluate the efficiency of using CNNs for diagnosis aids in glioma detection and the improvement of the method when using a clustering method (Fuzzy C-means) for preprocessing the input MRI dataset. Results offered an accuracy improvement from 0.77 to 0.81 when using Fuzzy C-Means.Ministerio de Economía y Competitividad TEC2016-77785-

Whole tumor RNA-sequencing and deconvolution reveal a clinically-prognostic PTEN/PI3K-regulated glioma transcriptional signature

The concept that solid tumors are maintained by a productive interplay between neoplastic and non-neoplastic elements has gained traction with the demonstration that stromal fibroblasts and immune system cells dictate cancer development and progression. While less studied, brain tumor (glioma) biology is likewise influenced by non-neoplastic immune system cells (macrophages and microglia) which interact with neoplastic glioma cells to create a unique physiological state (glioma ecosystem) distinct from that found in the normal tissue. To explore this neoplastic ground state, we leveraged several preclinical mouse models of neurofibromatosis type 1 (NF1) optic glioma, a low-grade astrocytoma whose formation and maintenance requires productive interactions between non-neoplastic and neoplastic cells, and employed whole tumor RNA-sequencing and mathematical deconvolution strategies to characterize this low-grade glioma ecosystem as an aggregate of cellular and acellular elements. Using this approach, we demonstrate that optic gliomas generated by altering the germlin

Enriched environment reduces glioma growth through immune and non-immune mechanisms in mice

Mice exposed to standard (SE) or enriched environment (EE) were transplanted with murine or human glioma cells and differences in tumour development were evaluated. We report that EE exposure affects: (i) tumour size, increasing mice survival; (ii) glioma establishment, proliferation and invasion; (iii) microglia/macrophage (M/Mφ) activation; (iv) natural killer (NK) cell infiltration and activation; and (v) cerebral levels of IL-15 and BDNF. Direct infusion of IL-15 or BDNF in the brain of mice transplanted with glioma significantly reduces tumour growth. We demonstrate that brain infusion of IL-15 increases the frequency of NK cell infiltrating the tumour and that NK cell depletion reduces the efficacy of EE and IL-15 on tumour size and of EE on mice survival. BDNF infusion reduces M/Mφ infiltration and CD68 immunoreactivity in tumour mass and reduces glioma migration inhibiting the small G protein RhoA through the truncated TrkB.T1 receptor. These results suggest alternative approaches for glioma treatment

Electrophysiology of glioma: a Rho GTPase-activating protein reduces tumor growth and spares neuron structure and function

Background. Glioblastomas are the most aggressive type of brain tumor. A successful treatment should aim at halting tumor growth and protecting neuronal cells to prevent functional deficits and cognitive deterioration. Here, we exploited a Rho GTPase-activating bacterial protein toxin, cytotoxic necrotizing factor 1 (CNF1), to interfere with glioma cell growth in vitro and vivo. We also investigated whether this toxin spares neuron structure and function in peritumoral areas. Methods. We performed a microarray transcriptomic and in-depth proteomic analysis to characterize the molecular changes triggered by CNF1 in glioma cells. We also examined tumor cell senescence and growth in vehicle-and CNF1-treated glioma-bearing mice. Electrophysiological and morphological techniques were used to investigate neuronal alterations in peritumoral cortical areas. Results. Administration of CNF1 triggered molecular and morphological hallmarks of senescence in mouse and human glioma cells in vitro. CNF1 treatment in vivo induced glioma cell senescence and potently reduced tumor volumes. In peritumoral areas of glioma-bearing mice, neurons showed a shrunken dendritic arbor and severe functional alterations such as increased spontaneous activity and reduced visual responsiveness. CNF1 treatment enhanced dendritic length and improved several physiological properties of pyramidal neurons, demonstrating functional preservation of the cortical network. Conclusions. Our findings demonstrate that CNF1 reduces glioma volume while at the same time maintaining the physiological and structural properties of peritumoral neurons. These data indicate a promising strategy for the development of more effective antiglioma therapies