• Glioblastoma multiforme
• Immunotherapy
• Signaling pathways
• Translational element

• Glioblastoma/therapy
• Glioblastoma/immunology
• Humans
• Brain Neoplasms/therapy
• Brain Neoplasms/immunology
• Immunotherapy/methods
• Tumor Microenvironment/immunology
• Immune Checkpoint Inhibitors/therapeutic use
• Oncolytic Virotherapy/methods
• Immunomodulation

• cell markers
• glioma stem cells
• nanomedicine
• niches
• signaling pathways
• targeted delivery

• Humans
• Brain Neoplasms/pathology
• Nanomedicine
• Glioma/pathology
• Glioblastoma/metabolism
• Neoplastic Stem Cells/metabolism
• Cell Line, Tumor
• Tumor Microenvironment

• GLI signaling
• Shh signaling
• glioma invasion
• scaffold proteins
• tumor stem cells

• Cell Line, Tumor
• Gene Expression Regulation, Neoplastic
• Glioblastoma/pathology
• Glioma/pathology
• Hedgehog Proteins/genetics
• Humans
• Membrane Proteins/genetics
• Membrane Proteins/metabolism
• Neoplastic Stem Cells/metabolism
• Transcription Factors/genetics
• Tumor Suppressor Proteins/genetics
• Tumor Suppressor Proteins/metabolism
• Zinc Finger Protein GLI1/genetics
• Zinc Finger Protein GLI1/metabolism

• P30 AG066546 NIA NIH HHS
• R21 NS106537 NINDS NIH HHS
• National Institutes of Health

Glioblastoma is the product of accumulated genetic and epigenetic alteration where tumor cells support each other through cellular communication mechanisms and deregulated signalling processes. The autocrine and paracrine pathways between the intracellular and extracellular milieu is mediated by connexin 43, the main gap junction-forming protein driving glioblastoma progression. In this scenario, sonic hedgehog pathway, a key deregulated pathway involved in cell network signalling may affect connexin 43 expression, promoting glioblastoma pathobiology. In this study, we sought to explore how the modulation of the sonic hedgehog affects connexin 43 inducing glioblastoma hallmarks. To do this we evaluated biological effects of sonic hedgehog pathway modulation by purmorphamine and cyclopamine, a smoothened agonist and antagonist, respectively. We revealed that cell migration and proliferation are associated with connexin 43 expression upon sonic hedgehog modulation. Our study suggests that sonic hedgehog and connexin 43 axis may represent a potential therapeutic strategy for glioblastoma.

Glioblastoma (GBM) represents the most common primary brain tumor within the adult population. Current therapeutic options are still limited by high rate of recurrences and signalling axes that promote GBM aggressiveness. The contribution of gap junctions (GJs) to tumor growth and progression has been proven by experimental evidence. Concomitantly, tumor microenvironment has received increasing interest as a critical process in dysregulation and homeostatic escape, finding a close link between molecular mechanisms involved in connexin 43 (CX43)-based intercellular communication and tumorigenesis. Moreover, evidence has come to suggest a crucial role of sonic hedgehog (SHH) signalling pathway in GBM proliferation, cell fate and differentiation. Herein, we used two human GBM cell lines, modulating SHH signalling and CX43-based intercellular communication in in vitro models using proliferation and migration assays. Our evidence suggests that modulation of the SHH effector smoothened (SMO), by using a known agonist (i.e., purmorphamine) and a known antagonist (i.e., cyclopamine), affects the CX43 expression levels and therefore the related functions. Moreover, SMO activation also increased cell proliferation and migration. Importantly, inhibition of CX43 channels was able to prevent SMO-induced effects. SHH pathway and CX43 interplay acts inducing tumorigenic program and supporting cell migration, likely representing druggable targets to develop new therapeutic strategies for GBM.

• GBM
• GLI1
• connexin
• gap junction
• smoothened

• Akt
• GLI
• S6K1
• SHH
• cell signaling
• glioma

• Glioblastoma
• Glioma stem cell
• SH3GL3
• STAT3
• Therapeutic target
• Tumor suppressor

• Chromatin remodeling
• DNA methylation
• Epigenetics
• Glioblastoma
• Histone deacetylase inhibitors

This paper examines the effect of TET1 expression on survival in glioma patients using open-access data from the Genomic Data Commons. A neural network-based survival model was built on expression data from a selection of genes most affected by TET1 knockdown with a median cross-validated survival concordance of 82.5%. A synthetic experiment was then conducted that linked two separately trained neural networks: a multitask model estimating cancer hallmark gene expression from TET1 expression, and a survival neural network. This experiment quantified the mediation of the TET1 survival effect through eight cancer hallmarks: apoptosis, cell cycle, cell death, cell motility, DNA repair, immune response, two phosphorylation pathways, and a randomized gene sets. Immune response, DNA repair, and apoptosis displayed greater mediation than the randomized gene set. Cell motility was inversely associated with only 12.5% mediated concordance. We propose the neural network linkage mediation experiment as an approach to collecting evidence of hazard mediation relationships with prognostic capacity useful for designing interventions.

Diffusely infiltrating gliomas (DIGs) are difficult to completely resect and are associated with a high rate of tumor relapse and progression from low- to high-grade glioma. In particular, optimized short-term culture-enriching patient-derived glioma stem cells (GSCs) are essential for customizing the therapeutic strategy based on clinically feasible in vitro drug screening for a wide range of DIGs, owing to the high inter-tumoral heterogeneity. Herein, we constructed a novel high-throughput culture condition screening platform called ‘GFSCAN’, which evaluated the cellular growth rates of GSCs for each DIG sample in 132 serum-free combinations, using 13 previously reported growth factors closely associated with glioma aggressiveness. In total, 72 patient-derived GSCs with available genomic profiles were tested in GFSCAN to explore the association between cellular growth rates in specific growth factor combinations and genomic/molecular backgrounds, including isocitrate dehydrogenase 1 (IDH1) mutation, chromosome arm 1p and 19q co-deletion, ATRX chromatin remodeler alteration, and transcriptional subtype. GSCs were clustered according to the dependency on epidermal growth factor and basic fibroblast growth factor (E&F), and isocitrate dehydrogenase 1 (IDH1) wild-type GSCs showed higher E&F dependencies than IDH1 mutant GSCs. More importantly, we elucidated optimal combinations for IDH1 mutant glioblastoma and lower grade glioma GSCs with low dependencies on E&F, which could be an aid in clinical decision-making for these DIGs. Thus, we demonstrated the utility of GFSCAN in personalizing in vitro cultivation to nominate personalized therapeutic options, in a clinically relevant time frame, for individual DIG patients, where standard clinical options have been exhausted.

• diffuse infiltrating glioma
• genomic profiling
• glioma stem cells
• growth factor
• personalized medicine
• short-term cultivation screening platform

• 1H-NMR spectroscopy
• Glioma
• Hh pathway
• Isoflavones
• Metabolomics

The invasive capacity of GBM is one of the key tumoral features associated with treatment resistance, recurrence, and poor overall survival. The molecular machinery underlying GBM invasiveness comprises an intricate network of signaling pathways and interactions with the extracellular matrix and host cells. Among them, PI3k/Akt, Wnt, Hedgehog, and NFkB play a crucial role in the cellular processes related to invasion. A better understanding of these pathways could potentially help in developing new therapeutic approaches with better outcomes. Nevertheless, despite significant advances made over the last decade on these molecular and cellular mechanisms, they have not been translated into the clinical practice. Moreover, targeting the infiltrative tumor and its significance regarding outcome is still a major clinical challenge. For instance, the pre- and intraoperative methods used to identify the infiltrative tumor are limited when trying to accurately define the tumor boundaries and the burden of tumor cells in the infiltrated parenchyma. Besides, the impact of treating the infiltrative tumor remains unclear. Here we aim to highlight the molecular and clinical hallmarks of invasion in GBM.

Glioblastoma is one of the deadliest malignancies and is virtually incurable. Accumulating evidence indicates that a small population of cells with a stem-like phenotype is the major culprit of tumor recurrence. Enhanced DNA repair capacity and expression of stemness marker genes are the main characteristics of these cells. Elimination of this population might delay or prevent tumor recurrence following radiochemotherapy. The aim of this study was to analyze whether interference with the Hedgehog signaling (Hh) pathway or combined Hh/Notch blockade using small-molecule inhibitors can efficiently target these cancer stem cells and sensitize them to therapy. Using tumor sphere lines and primary patient-derived glioma cultures we demonstrate that the Hh pathway inhibitor GANT61 (GANT) and the arsenic trioxide (ATO)-mediated Hh/Notch inhibition are capable to synergistically induce cell death in combination with the natural anticancer agent (−)-Gossypol (Gos). Only ATO in combination with Gos also strongly decreased stemness marker expression and prevented sphere formation and recovery. These synergistic effects were associated with distinct proteomic changes indicating diminished DNA repair and markedly reduced stemness. Finally, using an organotypic brain slice transplantation model, we show that combined ATO/Gos treatment elicits strong growth inhibition or even complete elimination of tumors. Collectively, our data show for the first time that ATO and Gos, two drugs that can be used in the clinic, represent a promising targeted therapy approach for the synergistic elimination of glioma stem-like cells.

• DNA damage
• Hedgehog
• Notch
• cancer stem cells
• glioblastoma

• Brain Neoplasms/genetics
• Brain Neoplasms/metabolism
• Cell Line, Tumor
• Cell Survival/drug effects
• Forkhead Box Protein M1/genetics
• Gene Expression Regulation, Neoplastic/drug effects
• Gene Knockdown Techniques
• Glioma/genetics
• Glioma/metabolism
• HeLa Cells
• Humans
• Neoplastic Stem Cells/drug effects
• Neoplastic Stem Cells/metabolism
• Nerve Tissue Proteins/antagonists & inhibitors
• Nerve Tissue Proteins/genetics
• Nuclear Proteins/antagonists & inhibitors
• Nuclear Proteins/genetics
• Pyridines/pharmacology
• Pyrimidines/pharmacology
• Repressor Proteins/antagonists & inhibitors
• Repressor Proteins/genetics
• Zinc Finger Protein GLI1/antagonists & inhibitors
• Zinc Finger Protein GLI1/genetics
• Zinc Finger Protein Gli2/antagonists & inhibitors
• Zinc Finger Protein Gli2/genetics
• Zinc Finger Protein Gli3/antagonists & inhibitors
• Zinc Finger Protein Gli3/genetics

• Russian Foundation for Assistance to Small Innovative Enterprises (FASIE)
• Russian Foundation for Basic Research

Many cancers have similar aberrations in various signaling cascades with crucial roles in cellular proliferation, differentiation, and morphogenesis. Dysregulation of signal cascades that play integral roles during early cellular development is well known to be a central feature of many malignancies. One such signaling cascade is the Wnt/β-catenin pathway, which has a profound effect on stem cell proliferation, migration, and differentiation. This pathway is dysregulated in numerous cell types, underscoring its global oncogenetic potential. This review highlights regulators and downstream effectors of this receptor cascade and addresses the increasingly apparent crosstalk of Wnt with other tumorigenic signaling pathways. As understanding of the genetic and epigenetic changes unique to these malignancies increases, identifying the regulatory mechanisms unique to the Wnt/β-catenin pathway and similarly aberrant receptor pathways will be imperative.

• Cell signaling
• Glioma stem cell
• Tumorigenicity
• Wnt
• β-catenin

The canonical hedgehog (HH) pathway is a multicomponent signaling cascade (HH, protein patched homolog 1 (PTCH1), smoothened (SMO)) that plays a pivotal role during embryonic development through activation of downstream effector molecules, namely glioma-associated oncogene homolog 1 (GLI1), GLI2 and GLI3. Activation of GLIs must be tightly regulated as they modulate target genes which control tissue patterning, stem cell maintenance, and differentiation during development. However, dysregulation or mutations in HH signaling leads to genomic instability (GI) and various cancers, for example, germline mutation in PTCH1 lead to Gorlin syndrome, a condition where patients develop numerous basal cell carcinomas and rarely rhabdomyosarcoma (RMS). Activating mutations in SMO have also been recognized in sporadic cases of medulloblastoma and SMO is overexpressed in many other cancers. Recently, studies in several human cancers have shown that GLI1 expression is independent from HH ligand and canonical intracellular signaling through PTCH and SMO. In fact, this aberrantly regulated GLI1 has been linked to several non-canonical oncogenic growth signals such as Kirsten rat sarcoma viral oncogene homolog (KRAS), avian myelocytomatosis virus oncogene cellular homolog (C-MYC), transforming growth factor β (TGFβ), wingless-type MMTV integration site family (WNT) and β-catenin. Recent studies from our lab and other independent studies demonstrate that aberrantly expressed GLI1 influences the integrity of several DNA damage response and repair signals, and if altered, these networks can contribute to GI and impact tumor response to chemo- and radiation therapies. Furthermore, the ineffectiveness of SMO inhibitors in clinical studies argues for the development of GLI1-specific inhibitors in order to develop effective therapeutic modalities to treat these tumors. In this review, we focus on summarizing current understanding of the molecular, biochemical and cellular basis for aberrant GLI1 expression and discuss GLI1-mediated HH signaling on DNA damage responses, carcinogenesis and chemoresistance.

• DNA damage
• GLI1
• carcinogenesis
• chemoresistance
• hedgehog signaling

• Hedgehog signaling pathway
• Shh protein
• glioblastoma multiforme
• transcription factor GLI

• Animals
• Cell Movement
• Cell Proliferation
• Central Nervous System Neoplasms/metabolism
• Central Nervous System Neoplasms/pathology
• Drug Resistance, Neoplasm
• Glioma/metabolism
• Glioma/pathology
• Hedgehog Proteins/genetics
• Hedgehog Proteins/metabolism
• Humans
• Signal Transduction
• Transcription Factors/genetics
• Transcription Factors/metabolism
• Zinc Finger Protein GLI1

• Wnt and SHH signaling pathways
• bottleneck nodes
• casein kinase 1, alpha 1 (CSNK1A1)
• gene expression
• glioblastoma
• glioma-associated oncogene 2 (Gli2) proteins
• integrative analysis
• protein–protein interaction networks
• simultaneous targeting
• β-catenin (CTNNB1)

• cancer stem cells
• glioblastoma
• neural stem cells

Glioblastoma Multiforme (GBM) is a grade IV astrocytoma, with a median survival of 14.6 mo. Within GBM, stem-like cells, namely glioblastoma stem cells (GSCs), have the ability to self-renew, differentiate into distinct lineages within the tumor and initiate tumor xenografts in immunocompromised animal models. More importantly, GSCs utilize cell-autonomous and tumor microenvironment-mediated mechanisms to overcome current therapeutic approaches. They are, therefore, very important therapeutic targets. Although the functional criteria defining GSCs are well defined, their molecular characteristics, the mechanisms whereby they establish the cellular hierarchy within tumors, and their contribution to tumor heterogeneity are not well understood. This review is aimed at summarizing current findings about GSCs and their therapeutic importance from a molecular and cellular point of view. A better characterization of GSCs is crucial for designing effective GSC-targeted therapies.

• Glioblastoma
• Glioblastoma stem cells
• Self-renewal
• Differentiation
• Molecular markers
• Therapy resistance

• P30 CA016087 NCI NIH HHS
• R21 NS087241 NINDS NIH HHS
• T32 GM007308 NIGMS NIH HHS
• UL1 TR000038 NCATS NIH HHS

Glioblastomas are the most lethal primary brain tumor that frequently relapse or progress as focal masses after radiation, suggesting that a fraction of tumor cells are responsible for the tumor regrowth. The identification of a brain tumor cell subpopulation with potent tumorigenic activity supports the cancer stem cell hypothesis in solid tumors. The goal of this study is to determine a methodology for the establishment of primary human glioblastoma cell lines. Our aim is achieved by taking the following approaches: (i) the establishment of primary glioblastoma cell culture; (ii) isolation of neurospheres derived from glioblastoma primary cultures; (iii) selection of CD133 cells from neurospheres, (iv) formation of subspheres in the CD133-positive population, (v) study of the expression level of GFAP, CD133, Nestin, Nanog, CD34, Sox2, CD44, and CD90 markers on tumor subspheres. Hence, we described a successful method for isolation of CD133-positive cell population and establishment of glioblastoma neurospheres from this primary culture, which are more robust than the ones derived straight from the tumor. Pointed out that the neurospheres derived from glioblastoma primary culture showed 29% more cells expressing CD133 then the ones straight tumor-derived, denoting a higher concentration of CD133-positive cells in the neurospheres derived from glioblastoma primary culture. These CD133-positive fractions were able to further generate subspheres. The subspheres derived from glioblastoma primary culture presented a well-defined morphology while the ones derived from the fresh tumor were sparce and less robust. And the negative fraction of CD133 cells was unable to generate subspheres. The tumor subspheres expressed GFAP, CD133, Nestin, Nanog, CD44, and CD90. Also, the present study describes an optimization of neurospheres/subspheres isolation from glioblastoma primary culture by selection of CD133-positive adherent stem cell.

• CD133
• adherent stem cell
• glioblastoma
• methods
• neurosphere/subspheres
• primary culture

• Hsa-miR-338-3p
• colorectal carcinoma
• invasion
• migration
• smoothened

• Epigenetics
• Glioma stem cells
• Hypoxia
• Microenvironment
• Therapeutic resistance

• Covariance estimation
• GLASSO
• Hierarchical penalty
• LASSO
• Multiple response
• Regression
• Sparsity

• R01 CA149569 NCI NIH HHS

• Animals
• Brain Neoplasms/genetics
• Brain Neoplasms/metabolism
• Brain Neoplasms/pathology
• Cell Line, Tumor
• Cell Proliferation
• Drug Resistance, Neoplasm/genetics
• Extracellular Matrix Proteins/genetics
• Extracellular Matrix Proteins/metabolism
• Extracellular Matrix Proteins/physiology
• Gene Expression Regulation, Neoplastic/drug effects
• Glioma/genetics
• Glioma/metabolism
• Glioma/pathology
• HEK293 Cells
• Humans
• Mice
• Mice, Nude
• Paracrine Communication/genetics
• Paracrine Communication/physiology
• RNA, Small Interfering/pharmacology
• Rats
• Receptor, Notch1/genetics
• Receptor, Notch1/metabolism
• Receptor, Notch1/physiology
• Xenograft Model Antitumor Assays

• R01 CA152065 NCI NIH HHS
• 1R01CA152065-01 NCI NIH HHS

Emerging laboratory and clinical investigations demonstrate that Hedgehog signaling (Hh) represents a novel therapeutic target in various human cancers. This conserved signaling pathway precisely regulates self-renewal and terminal differentiation in embryonic development, but is typically silenced in adult tissues, with reactivation usually only during tissue repair. Aberrant Hh pathway signaling has been implicated in the pathogenesis, self-renewal, and chemotherapy resistance of a growing number of solid and hematologic malignancies. Major components of the Hh pathway include the Hh ligands (Sonic, Desert, and Indian), the transmembrane receptor Patched, the signal transducer Smoothened (Smo), and transcription factors Gli1–3 which regulate the transcription of Hh target genes. Mutations in Hh pathway genes, increased Hh signaling in tumor stroma, and Hh overexpression in self-renewing cells (cancer stem cells) have been described, and these different modes of Hh signaling have implications for the design of Hh pathway inhibitors and their integration into conventional treatment regimens. Discovery of a naturally-occurring Smo inhibitor, cyclopamine, and the identification of Hh pathway mutations and over expression in cancer cells prompted the development of several cyclopamine derivatives. Encouraging laboratory and in vivo data has resulted in Phase I and II clinical trials of Smo inhibitors. In this review, we will discuss the current understanding of Hh pathway signaling in malignancy and Smo antagonists in development. Recent data with these agents shows that they are well-tolerated and may be effective for subsets of patients. Challenges remain for appropriate patient selection and the optimal combination and sequence of these targeted therapies into current treatment paradigms.

• Smoothened inhibitors
• cancer stem cells
• hedgehog pathway