bims-malgli 2021-09-12 papers

bims-malgli

Biomed News

on Biology of malignant gliomas

Issue of 2021–09–12
twenty-two papers selected by
Oltea Sampetrean, Keio University

Downregulated CLIP3 induces radioresistance by enhancing stemness and glycolytic flux in glioblastoma.
Nonredundant, isoform-specific roles of HDAC1 in glioma stem cells.
In Vitro and In Vivo Enhancement of Temozolomide Effect in Human Glioblastoma by Non-Invasive Application of Cold Atmospheric Plasma.
The Novel Antitumor Compound HCA Promotes Glioma Cell Death by Inducing Endoplasmic Reticulum Stress and Autophagy.
Combination checkpoint therapy with anti-PD-1 and anti-BTLA results in a synergistic therapeutic effect against murine glioblastoma.
Inhibition of microglial EZH2 leads to anti-tumoral effects in pediatric diffuse midline gliomas.
Cilium induction triggers differentiation of glioma stem cells.
Slow Off-Rate Modified Aptamer (SOMAmer) Proteomic Analysis of Patient-Derived Malignant Glioma Identifies Distinct Cellular Proteomes.
Tumor-Associated Microglia and Macrophages in the Glioblastoma Microenvironment and Their Implications for Therapy.
Gelatin methacrylate hydrogels culture model for glioblastoma cells enriches for mesenchymal-like state and models interactions with immune cells.
A map of the altered glioma metabolism.
Nuclear IMPDH Filaments in Human Gliomas.
Impairment of Autophagic Flux Participates in the Antitumor Effects of TAT-Cx43266-283 in Glioblastoma Stem Cells.
Histone deacetylase inhibitors enhance estrogen receptor beta expression and augment agonist-mediated tumor suppression in glioblastoma.
Viral targeting of glioblastoma stem cells with patient-specific genetic and post-translational p53 deregulations.
Regulatory network of PD1 signaling is associated with prognosis in glioblastoma multiforme.
Anti-tumor effects of vitamin D in glioblastoma: mechanism and therapeutic implications.
Grading of adult diffuse gliomas according to the 2021 WHO Classification of Tumors of the Central Nervous System.
Short-term fasting in glioma patients: analysis of diet diaries and metabolic parameters of the ERGO2 trial.
Glioblastoma signature in the DNA of blood-derived cells.
Localized blood-brain barrier opening in infiltrating gliomas with MRI-guided acoustic emissions-controlled focused ultrasound.
Immune cell deconvolution of bulk DNA methylation data reveals an association with methylation class, key somatic alterations, and cell state in glial/glioneuronal tumors.

J Exp Clin Cancer Res. 2021 Sep 06. 40(1): 282

Downregulated CLIP3 induces radioresistance by enhancing stemness and glycolytic flux in glioblastoma.

Hyunkoo Kang, Sungmin Lee, Kyeongmin Kim, Jaewan Jeon, Seok-Gu Kang, HyeSook Youn, Hae Yu Kim, BuHyun Youn.

   BACKGROUND: Glioblastoma Multiforme (GBM) is a malignant primary brain tumor in which the standard treatment, ionizing radiation (IR), achieves a median survival of about 15 months. GBM harbors glioblastoma stem-like cells (GSCs), which play a crucial role in therapeutic resistance and recurrence.
METHODS: Patient-derived GSCs, GBM cell lines, intracranial GBM xenografts, and GBM sections were used to measure mRNA and protein expression and determine the related molecular mechanisms by qRT-PCR, immunoblot, immunoprecipitation, immunofluorescence, OCR, ECAR, live-cell imaging, and immunohistochemistry. Orthotopic GBM xenograft models were applied to investigate tumor inhibitory effects of glimepiride combined with radiotherapy.
RESULTS: We report that GSCs that survive standard treatment radiation upregulate Speedy/RINGO cell cycle regulator family member A (Spy1) and downregulate CAP-Gly domain containing linker protein 3 (CLIP3, also known as CLIPR-59). We discovered that Spy1 activation and CLIP3 inhibition coordinately shift GBM cell glucose metabolism to favor glycolysis via two cellular processes: transcriptional regulation of CLIP3 and facilitating Glucose transporter 3 (GLUT3) trafficking to cellular membranes in GBM cells. Importantly, in combination with IR, glimepiride, an FDA-approved medication used to treat type 2 diabetes mellitus, disrupts GSCs maintenance and suppresses glycolytic activity by restoring CLIP3 function. In addition, combining radiotherapy and glimepiride significantly reduced GBM growth and improved survival in a GBM orthotopic xenograft mouse model.
CONCLUSIONS: Our data suggest that radioresistant GBM cells exhibit enhanced stemness and glycolytic activity mediated by the Spy1-CLIP3 axis. Thus, glimepiride could be an attractive strategy for overcoming radioresistance and recurrence by rescuing CLIP3 expression.

Keywords:  CLIP3; Glimepiride; Glioblastoma; Glioblastoma stem-like cells; Radioresistance

DOI:  https://doi.org/10.1186/s13046-021-02077-4
JCI Insight. 2021 Sep 08. pii: 149232. [Epub ahead of print]6(17):

Nonredundant, isoform-specific roles of HDAC1 in glioma stem cells.

Costanza Lo Cascio, James B McNamara, Ernesto L Melendez, Erika M Lewis, Matthew E Dufault, Nader Sanai, Christopher L Plaisier, Shwetal Mehta.

  Glioblastoma (GBM) is characterized by an aberrant yet druggable epigenetic landscape. One major family of epigenetic regulators, the histone deacetylases (HDACs), are considered promising therapeutic targets for GBM due to their repressive influences on transcription. Although HDACs share redundant functions and common substrates, the unique isoform-specific roles of different HDACs in GBM remain unclear. In neural stem cells, HDAC2 is the indispensable deacetylase to ensure normal brain development and survival in the absence of HDAC1. Surprisingly, we find that HDAC1 is the essential class I deacetylase in glioma stem cells, and its loss is not compensated for by HDAC2. Using cell-based and biochemical assays, transcriptomic analyses, and patient-derived xenograft models, we find that knockdown of HDAC1 alone has profound effects on the glioma stem cell phenotype in a p53-dependent manner. We demonstrate marked suppression in tumor growth upon targeting of HDAC1 and identify compensatory pathways that provide insights into combination therapies for GBM. Our study highlights the importance of HDAC1 in GBM and the need to develop isoform-specific drugs.

Keywords:  Brain cancer; Cancer; Epigenetics; Oncology; Stem cells

DOI:  https://doi.org/10.1172/jci.insight.149232
Cancers (Basel). 2021 Sep 06. pii: 4485. [Epub ahead of print]13(17):

In Vitro and In Vivo Enhancement of Temozolomide Effect in Human Glioblastoma by Non-Invasive Application of Cold Atmospheric Plasma.

Vikas Soni, Manish Adhikari, Hayk Simonyan, Li Lin, Jonathan H Sherman, Colin N Young, Michael Keidar.

  Glioblastoma (GBM) is one of the most aggressive forms of adult brain cancers and is highly resistant to treatment, with a median survival of 12-18 months after diagnosis. The poor survival is due to its infiltrative pattern of invasion into the normal brain parenchyma, the diffuse nature of its growth, and its ability to quickly grow, spread, and relapse. Temozolomide is a well-known FDA-approved alkylating chemotherapy agent used for the treatment of high-grade malignant gliomas, and it has been shown to improve overall survival. However, in most cases, the tumor relapses. In recent years, CAP has been used as an emerging technology for cancer therapy. The purpose of this study was to implement a combination therapy of CAP and TMZ to enhance the effect of TMZ and apparently sensitize GBMs. In vitro evaluations in TMZ-sensitive and resistant GBM cell lines established a CAP chemotherapy enhancement and potential sensitization effect across various ranges of CAP jet application. This was further supported with in vivo findings demonstrating that a single CAP jet applied non-invasively through the skull potentially sensitizes GBM to subsequent treatment with TMZ. Gene functional enrichment analysis further demonstrated that co-treatment with CAP and TMZ resulted in a downregulation of cell cycle pathway genes. These observations indicate that CAP can be potentially useful in sensitizing GBM to chemotherapy and for the treatment of glioblastoma as a non-invasive translational therapy.

Keywords:  RNA-sequencing; bone; cancer therapy; cold atmospheric plasma; electromagnetic waves; gene functional enrichment analysis; glioblastoma; plasma medicine; reactive oxygen and nitrogen species; sensitization; temozolomide; xenografts

DOI:  https://doi.org/10.3390/cancers13174485
Cancers (Basel). 2021 Aug 26. pii: 4290. [Epub ahead of print]13(17):

The Novel Antitumor Compound HCA Promotes Glioma Cell Death by Inducing Endoplasmic Reticulum Stress and Autophagy.

Roberto Beteta-Göbel, Javier Fernández-Díaz, Laura Arbona-González, Raquel Rodríguez-Lorca, Manuel Torres, Xavier Busquets, Paula Fernández-García, Pablo V Escribá, Victoria Lladó.

  Glioblastoma (GBM) is the most common and aggressive type of primary brain tumor in adults, and the median survival of patients with GBM is 14.5 months. Melitherapy is an innovative therapeutic approach to treat different diseases, including cancer, and it is based on the regulation of cell membrane composition and structure, which modulates relevant signal pathways. Here, we have tested the effects of 2-hydroxycervonic acid (HCA) on GBM cells and xenograft tumors. HCA was taken up by cells and it compromised the survival of several human GBM cell lines in vitro, as well as the in vivo growth of xenograft tumors (mice) derived from these cells. HCA appeared to enhance ER stress/UPR signaling, which consequently induced autophagic cell death of the GBM tumor cells. This negative effect of HCA on GBM cells may be mediated by the JNK/c-Jun/CHOP/BiP axis, and it also seems to be provoked by the cellular metabolite of HCA, C21:5n-3 (heneicosapentaenoic acid). These results demonstrate the efficacy of the melitherapeutic treatment used and the potential of using C21:5n-3 as an efficacy biomarker for this treatment. Given the safety profile in animal models, the data presented here provide evidence that HCA warrants further clinical study as a potential therapy for GBM, currently an important unmet medical need.

Keywords:  ER stress; HCA; autophagy; glioma; membrane lipid therapy

DOI:  https://doi.org/10.3390/cancers13174290
Oncoimmunology. 2021 ;10(1): 1956142

Combination checkpoint therapy with anti-PD-1 and anti-BTLA results in a synergistic therapeutic effect against murine glioblastoma.

John Choi, Ravi Medikonda, Laura Saleh, Timothy Kim, Ayush Pant, Siddhartha Srivastava, Young-Hoon Kim, Christina Jackson, Luqing Tong, Denis Routkevitch, Christopher Jackson, Dimitrios Mathios, Tianna Zhao, Hyerim Cho, Henry Brem, Michael Lim.

  Clinical trials involving anti-programmed cell death protein-1 (anti-PD-1) failed to demonstrate improved overall survival in glioblastoma (GBM) patients. This may be due to the expression of alternative checkpoints such as B- and T- lymphocyte attenuator (BTLA) on several immune cell types including regulatory T cells. Murine GBM models indicate that there is significant upregulation of BTLA in the tumor microenvironment (TME) with associated T cell exhaustion. We investigate the use of antibodies against BTLA and PD-1 on reversing immunosuppression and increasing long-term survival in a murine GBM model. C57BL/6 J mice were implanted with the murine glioma cell line GL261 and randomized into 4 arms: (i) control, (ii) anti-PD-1, (iii) anti-BTLA, and (iv) anti-PD-1 + anti-BTLA. Kaplan-Meier curves were generated for all arms. Flow cytometric analysis of blood and brains were done on days 11 and 16 post-tumor implantation. Tumor-bearing mice treated with a combination of anti-PD-1 and anti-BTLA therapy experienced improved overall long-term survival (60%) compared to anti-PD-1 (20%) or anti-BTLA (0%) alone (P = .003). Compared to monotherapy with anti-PD-1, mice treated with combination therapy also demonstrated increased expression of CD4+ IFN-γ (P < .0001) and CD8+ IFN-γ (P = .0365), as well as decreased levels of CD4+ FoxP3+ regulatory T cells on day 16 in the brain (P = .0136). This is the first preclinical investigation into the effects of combination checkpoint blockade with anti-PD-1 and anti-BTLA treatment in GBM. We also show a direct effect on activated immune cell populations such as CD4+ and CD8 + T cells and immunosuppressive regulatory T cells through this combination therapy.

Keywords:  B and T lymphocyte attenuator; anti-BTLA; anti-PD-1; glioblastoma; glioblastoma immunotherapy; immune checkpoint inhibitor therapy

DOI:  https://doi.org/10.1080/2162402X.2021.1956142
Neurooncol Adv. 2021 Jan-Dec;3(1):3(1): vdab096

Inhibition of microglial EZH2 leads to anti-tumoral effects in pediatric diffuse midline gliomas.

Lily Keane, Mathilde Cheray, Dalel Saidi, Caoimhe Kirby, Lara Friess, Patricia Gonzalez-Rodriguez, Maren Elisabeth Gerdes, Kathleen Grabert, Barry W McColl, Bertrand Joseph.

   Background: Diffuse intrinsic pontine gliomas (DIPG), within diffuse midline gliomas are aggressive pediatric brain tumors characterized by histone H3-K27M mutation. Small-molecule inhibitors for the EZH2-H3K27 histone methyltransferase have shown promise in preclinical animal models of DIPG, despite having little effect on DIPG cells in vitro. Therefore, we hypothesized that the effect of EZH2 inhibition could be mediated through targeting of this histone modifying enzyme in tumor-associated microglia.
Methods: Primary DIPG tissues, and cocultures between microglia and patient-derived DIPG or -pediatric high-grade glioma (pHGG) cell lines, were used to establish the H3-K27M status of each cell type. Antisense RNA strategies were used to target EZH2 gene expression in both microglia and glioma cells. Microglia anti-tumoral properties were assessed by gene expression profile, tumor cell invasion capacity, microglial phagocytic activity, and associated tumor cell death.
Results: In primary DIPG tissues, microglia do not carry the H3-K27M mutation, otherwise characteristic of the cancer cells. Activation of a microglial tumor-supportive phenotype by pHGG, independently of their H3-K27M status, is associated with a transient H3K27me3 downregulation. Repression of EZH2 in DIPG cells has no impact on tumor cell survival or their ability to activate microglia. However, repression of EZH2 in microglia induces an anti-tumor phenotype resulting in decreased cancer cell invasion capability, increased microglial phagocytosis, and tumor-related cell death.
Conclusions: These results indicate that microglia, beyond the tumor cells, contribute to the observed response of DIPG to EZH2 inhibition. Results highlight the potential importance of microglia as a new therapeutic avenue in DIPG.

Keywords:  DIPG; EZH2; H3K27me3; anti-tumoral; microglia

DOI:  https://doi.org/10.1093/noajnl/vdab096
Cell Rep. 2021 Sep 07. pii: S2211-1247(21)01099-8. [Epub ahead of print]36(10): 109656

Cilium induction triggers differentiation of glioma stem cells.

Gladiola Goranci-Buzhala, Aruljothi Mariappan, Lucia Ricci-Vitiani, Natasa Josipovic, Simone Pacioni, Marco Gottardo, Johannes Ptok, Heiner Schaal, Giuliano Callaini, Krishnaraj Rajalingam, Brian Dynlacht, Kamyar Hadian, Argyris Papantonis, Roberto Pallini, Jay Gopalakrishnan.

  Glioblastoma multiforme (GBM) possesses glioma stem cells (GSCs) that promote self-renewal, tumor propagation, and relapse. Understanding the mechanisms of GSCs self-renewal can offer targeted therapeutic interventions. However, insufficient knowledge of GSCs' fundamental biology is a significant bottleneck hindering these efforts. Here, we show that patient-derived GSCs recruit elevated levels of proteins that ensure the temporal cilium disassembly, leading to suppressed ciliogenesis. Depleting the cilia disassembly complex components is sufficient to induce ciliogenesis in a subset of GSCs via relocating platelet-derived growth factor receptor-alpha (PDGFR-α) to a newly induced cilium. Importantly, restoring ciliogenesis enabled GSCs to switch from self-renewal to differentiation. Finally, using an organoid-based glioma invasion assay and brain xenografts in mice, we establish that ciliogenesis-induced differentiation can prevent the infiltration of GSCs into the brain. Our findings illustrate a role for cilium as a molecular switch in determining GSCs' fate and suggest cilium induction as an attractive strategy to intervene in GSCs proliferation.

Keywords:  brain organoids; cell cycle; cilium checkpoint; confocal 3D imaging; glioblastoma; invasion assay; primary cilium; tissue clearing

DOI:  https://doi.org/10.1016/j.celrep.2021.109656
Int J Mol Sci. 2021 Sep 03. pii: 9566. [Epub ahead of print]22(17):

Slow Off-Rate Modified Aptamer (SOMAmer) Proteomic Analysis of Patient-Derived Malignant Glioma Identifies Distinct Cellular Proteomes.

Thatchawan Thanasupawat, Aleksandra Glogowska, Christopher Pascoe, Sai Nivedita Krishnan, Maliha Munir, Farhana Begum, Jason Beiko, Jerry Krcek, Marc R Del Bigio, Marshall Pitz, Yaoqing Shen, Victor Spicer, Kevin M Coombs, John Wilkins, Sabine Hombach-Klonisch, Thomas Klonisch.

  Malignant gliomas derive from brain glial cells and represent >75% of primary brain tumors. This includes anaplastic astrocytoma (grade III; AS), the most common and fatal glioblastoma multiforme (grade IV; GBM), and oligodendroglioma (ODG). We have generated patient-derived AS, GBM, and ODG cell models to study disease mechanisms and test patient-centered therapeutic strategies. We have used an aptamer-based high-throughput SOMAscan® 1.3K assay to determine the proteomic profiles of 1307 different analytes. SOMAscan® proteomes of AS and GBM self-organized into closely adjacent proteomes which were clearly distinct from ODG proteomes. GBM self-organized into four proteomic clusters of which SOMAscan® cluster 4 proteome predicted a highly inter-connected proteomic network. Several up- and down-regulated proteins relevant to glioma were successfully validated in GBM cell isolates across different SOMAscan® clusters and in corresponding GBM tissues. Slow off-rate modified aptamer proteomics is an attractive analytical tool for rapid proteomic stratification of different malignant gliomas and identified cluster-specific SOMAscan® signatures and functionalities in patient GBM cells.

Keywords:  SOMAmers; glioblastoma; glioma; patient cell isolates; proteomic clusters

DOI:  https://doi.org/10.3390/ijms22179566
Cancers (Basel). 2021 Aug 24. pii: 4255. [Epub ahead of print]13(17):

Tumor-Associated Microglia and Macrophages in the Glioblastoma Microenvironment and Their Implications for Therapy.

Rikke Sick Andersen, Atul Anand, Dylan Scott Lykke Harwood, Bjarne Winther Kristensen.

  Glioblastoma is the most frequent and malignant primary brain tumor. Standard of care includes surgery followed by radiation and temozolomide chemotherapy. Despite treatment, patients have a poor prognosis with a median survival of less than 15 months. The poor prognosis is associated with an increased abundance of tumor-associated microglia and macrophages (TAMs), which are known to play a role in creating a pro-tumorigenic environment and aiding tumor progression. Most treatment strategies are directed against glioblastoma cells; however, accumulating evidence suggests targeting of TAMs as a promising therapeutic strategy. While TAMs are typically dichotomously classified as M1 and M2 phenotypes, recent studies utilizing single cell technologies have identified expression pattern differences, which is beginning to give a deeper understanding of the heterogeneous subpopulations of TAMs in glioblastomas. In this review, we evaluate the role of TAMs in the glioblastoma microenvironment and discuss how their interactions with cancer cells have an extensive impact on glioblastoma progression and treatment resistance. Finally, we summarize the effects and challenges of therapeutic strategies, which specifically aim to target TAMs.

Keywords:  TAM; crosstalk; glioblastoma; microenvironment; therapeutic strategies; tumor-associated microglia and macrophages

DOI:  https://doi.org/10.3390/cancers13174255
Sci Rep. 2021 Sep 06. 11(1): 17727

Gelatin methacrylate hydrogels culture model for glioblastoma cells enriches for mesenchymal-like state and models interactions with immune cells.

Nameeta Shah, Pavan M Hallur, Raksha A Ganesh, Pranali Sonpatki, Divya Naik, Komal Prasad Chandrachari, Ralph B Puchalski, Aditya Chaubey.

Glioblastoma is the most lethal primary malignant brain tumor in adults. Simplified two-dimensional (2D) cell culture and neurospheres in vitro models fail to recapitulate the complexity of the tumor microenvironment, limiting its ability to predict therapeutic response. Three-dimensional (3D) scaffold-based models have emerged as a promising alternative for addressing these concerns. One such 3D system is gelatin methacrylate (GelMA) hydrogels, and we aimed to understand the suitability of using this system to mimic treatment-resistant glioblastoma cells that reside in specific niches. We characterized the phenotype of patient-derived glioma cells cultured in GelMA hydrogels (3D-GMH) for their tumorigenic properties using invasion and chemoresponse assays. In addition, we used integrated single-cell and spatial transcriptome analysis to compare cells cultured in 3D-GMH to neoplastic cells in vivo. Finally, we assessed tumor-immune cell interactions with a macrophage infiltration assay and a cytokine array. We show that the 3D-GMH system enriches treatment-resistant mesenchymal cells that are not represented in neurosphere cultures. Cells cultured in 3D-GMH resemble a mesenchymal-like cellular phenotype found in perivascular and hypoxic regions and recruit macrophages by secreting cytokines, a hallmark of the mesenchymal phenotype. Our 3D-GMH model effectively mimics the phenotype of glioma cells that are found in the perivascular and hypoxic niches of the glioblastoma core in situ, in contrast to the neurosphere cultures that enrich cells of the infiltrative edge of the tumor. This contrast highlights the need for due diligence in selecting an appropriate model when designing a study's objectives.

DOI: https://doi.org/10.1038/s41598-021-97059-z
Trends Mol Med. 2021 Sep 03. pii: S1471-4914(21)00196-9. [Epub ahead of print]

A map of the altered glioma metabolism.

Ruhi Deshmukh, Maria Francesca Allega, Saverio Tardito.

  The frequent occurrence of neomorphic isocitrate dehydrogenase 1 (IDH1) mutations in low-grade glioma led to an IDH-centric classification of these tumors. However, exploiting metabolic alterations of glioma for diagnostic imaging and treatment has marginally improved patients' prognosis. Here we discuss the nutritional microenvironment of glioma, shaped by the distinctive dependence of the brain on glucose and ketone bodies for energy, and on amino acids for neurotransmission. We highlight the progress in metabolic applications for glioma diagnosis and therapy, and present a map that streamlines the rewired glioma metabolism. The map illustrates the altered reactions in central carbon and nitrogen metabolism that drive glioma biology, and represent metabolic vulnerabilities with translational potential.

Keywords:  IDH1 mutation; cancer metabolism; glioblastoma; glioma

DOI:  https://doi.org/10.1016/j.molmed.2021.07.011
J Neuropathol Exp Neurol. 2021 Sep 08. pii: nlab090. [Epub ahead of print]

Nuclear IMPDH Filaments in Human Gliomas.

Narges Ahangari, David G Munoz, Josee Coulombe, Douglas A Gray, Elizabeth C Engle, Long Cheng, John Woulfe.

  The analysis of nuclear morphology plays an important role in glioma diagnosis and grading. We previously described intranuclear rods (rods) labeled with the SDL.3D10 monoclonal antibody against class III beta-tubulin (TUBB3) in human ependymomas. In a cohort of adult diffuse gliomas, we identified nuclear rods in 71.1% of IDH mutant lower-grade gliomas and 13.7% of IDH wild-type glioblastomas (GBMs). The presence of nuclear rods was associated with significantly longer postoperative survival in younger (≤65) GBM patients. Consistent with this, nuclear rods were mutually exclusive with Ki67 staining and their prevalence in cell nuclei inversely correlated with the Ki67 proliferation index. In addition, rod-containing nuclei showed a relative depletion of lamin B1, suggesting a possible association with senescence. To gain insight into their functional significance, we addressed their antigenic properties. Using a TUBB3-null mouse model, we demonstrate that the SDL.3D10 antibody does not bind TUBB3 in rods but recognizes an unknown antigen. In the present study, we show that rods show immunoreactivity for the nucleotide synthesizing enzymes inosine monophosphate dehydrogenase (IMPDH) and cytidine triphosphate synthetase. By analogy with the IMPDH filaments that have been described previously, we postulate that rods regulate the activity of nucleotide-synthesizing enzymes in the nucleus by sequestration, with important implications for glioma behavior.

Keywords:  Cytoophidia; Glioblastoma; Glioma; IMPDH filaments; Nuclear rods

DOI:  https://doi.org/10.1093/jnen/nlab090
Cancers (Basel). 2021 Aug 24. pii: 4262. [Epub ahead of print]13(17):

Impairment of Autophagic Flux Participates in the Antitumor Effects of TAT-Cx43266-283 in Glioblastoma Stem Cells.

Sara G Pelaz, Claudia Ollauri-Ibáñez, Concepción Lillo, Arantxa Tabernero.

  Autophagy is a physiological process by which various damaged or non-essential cytosolic components are recycled, contributing to cell survival under stress conditions. In cancer, autophagy can have antitumor or protumor effects depending on the developmental stage. Here, we use Western blotting, immunochemistry, and transmission electron microscopy to demonstrate that the antitumor peptide TAT-Cx43266-283, a c-Src inhibitor, blocks autophagic flux in glioblastoma stem cells (GSCs) under basal and nutrient-deprived conditions. Upon nutrient deprivation, GSCs acquired a dormant-like phenotype that was disrupted by inhibition of autophagy with TAT-Cx43266-283 or chloroquine (a classic autophagy inhibitor), leading to GSC death. Remarkably, dasatinib, a clinically available c-Src inhibitor, could not replicate TAT-Cx43266-283 effect on dormant GSCs, revealing for the first time the possible involvement of pathways other than c-Src in TAT-Cx43266-283 effect. TAT-Cx43266-283 exerts an antitumor effect both in nutrient-complete and nutrient-deprived environments, which constitutes an advantage over chloroquine and dasatinib, whose effects depend on nutrient environment. Finally, our analysis of the levels of autophagy-related proteins in healthy and glioma donors suggests that autophagy is upregulated in glioblastoma, further supporting the interest in inhibiting this process in the most aggressive brain tumor and the potential use of TAT-Cx43266-283 as a therapy for this type of cancer.

Keywords:  autophagy; c-Src; cell-penetrating peptide; connexin43; glioblastoma; glioblastoma stem cells

DOI:  https://doi.org/10.3390/cancers13174262
Neurooncol Adv. 2021 Jan-Dec;3(1):3(1): vdab099

Histone deacetylase inhibitors enhance estrogen receptor beta expression and augment agonist-mediated tumor suppression in glioblastoma.

Uday P Pratap, Gangadhara R Sareddy, Zexuan Liu, Prabhakar Pitta Venkata, Junhao Liu, Weiwei Tang, Kristin A Altwegg, Behnam Ebrahimi, Xiaonan Li, Rajeshwar R Tekmal, Suryavathi Viswanadhapalli, Stanton McHardy, Andrew J Brenner, Ratna K Vadlamudi.

   Background: Glioblastomas (GBMs) are the most lethal primary brain tumors. Estrogen receptor β (ESR2/ERβ) function as a tumor suppressor in GBM, however, ERβ expression is commonly suppressed during glioma progression. In this study, we examined whether drugs that reverse epigenetic modifications will enhance ERβ expression and augment ERβ agonist-mediated tumor suppression.
Methods: We tested the utility of epigenetic drugs which act as an inhibitor of histone deacetylases (HDACs), histone methylases, and BET enzymes. Mechanistic studies utilized RT-qPCR, chromatin immunoprecipitation (ChIP), and western blotting. Cell viability, apoptosis, colony formation, and invasion were measured using in vitro assays. An orthotopic GBM model was used to test the efficacy of in vivo.
Results: Of all inhibitors tested, HDACi (panobinostat and romidepsin) showed the potential to increase the expression of ERβ in GBM cells. Treatment with HDACi uniquely upregulated ERβ isoform 1 expression that functions as a tumor suppressor but not ERβ isoform 5 that drives oncogenic functions. Further, combination therapy of HDACi with the ERβ agonist, LY500307, potently reduced cell viability, invasion, colony formation, and enhanced apoptosis. Mechanistic studies showed that HDACi induced ERβ is functional, as it enhanced ERβ reporter activities and ERβ target genes expression. ChIP analysis confirmed alterations in the histone acetylation at the ERβ and its target gene promoters. In orthotopic GBM model, combination therapy of panobinostat and LY500307 enhanced survival of tumor-bearing mice.
Conclusions: Our results suggest that the combination therapy of HDACi and LY500307 provides therapeutic utility in overcoming the suppression of ERβ expression that commonly occurs in GBM progression.

Keywords:  epigenetic drugs; estrogen receptor β; glioblastoma; panobinostat; romidepsin

DOI:  https://doi.org/10.1093/noajnl/vdab099
Cell Rep. 2021 Sep 07. pii: S2211-1247(21)01117-7. [Epub ahead of print]36(10): 109673

Viral targeting of glioblastoma stem cells with patient-specific genetic and post-translational p53 deregulations.

Jon Gil-Ranedo, Carlos Gallego-García, José M Almendral.

  Cancer therapy urges targeting of malignant subsets within self-renewing heterogeneous stem cell populations. We dissect the genetic and functional heterogeneity of human glioblastoma stem cells (GSCs) within patients by their innate responses to non-pathogenic mouse parvoviruses that are tightly restrained by cellular physiology. GSC neurospheres accumulate assembled capsids but restrict viral NS1 cytotoxic protein expression by an innate PKR/eIF2α-P response counteractable by electric pulses. NS1 triggers a comprehensive DNA damage response involving cell-cycle arrest, neurosphere disorganization, and bystander disruption of GSC-derived brain tumor architecture in rodent models. GSCs and cancer cell lines permissive to parvovirus genome replication require p53-Ser15 phosphorylation (Pp53S15). NS1 expression is enhanced by exogeneous Pp53S15 induction but repressed by wtp53. Consistently, patient-specific GSC subpopulations harboring p53 gain-of-function mutants and/or Pp53S15 are selective viral targets. This study provides a molecular foundation for personalized biosafe viral therapies against devastating glioblastoma and other cancers with deregulated p53 signaling.

Keywords:  DDR; PKR; brain tumor MRI; cancer virotherapy; genetic heterogeneity; glioblastoma stem cells; neurospheres; p53 GOF mutants; p53-Ser15 phosphorylation; parvovirus

DOI:  https://doi.org/10.1016/j.celrep.2021.109673
Cancer Res. 2021 Sep 07. pii: canres.0730.2021. [Epub ahead of print]

Regulatory network of PD1 signaling is associated with prognosis in glioblastoma multiforme.

Camila M Lopes-Ramos, Tatiana Belova, Tess H Brunner, Marouen Ben Guebila, Daniel Osorio, John Quackenbush, Marieke L Kuijjer.

Glioblastoma is an aggressive cancer of the brain and spine. While analysis of glioblastoma 'omics data has somewhat improved our understanding of the disease, it has not led to direct improvement in patient survival. Cancer survival is often characterized by differences in gene expression, but the mechanisms that drive these differences are generally unknown. We therefore set out to model the regulatory mechanisms associated with glioblastoma survival. We inferred individual patient gene regulatory networks using data from two different expression platforms from The Cancer Genome Atlas. We performed comparative network analysis between patients with long- and short-term survival. Seven pathways were identified as associated with survival, all of them involved in immune signaling; differential regulation of PD1 signaling was validated to correspond with outcome in an independent dataset from the German Glioma Network. In this pathway, transcriptional repression of genes for which treatment options are available was lost in short-term survivors; this was independent of mutational burden and only weakly associated with T-cell infiltration. Collectively, these results provide a new way to stratify glioblastoma patients that uses network features as biomarkers to predict survival. They also identify new potential therapeutic interventions, underscoring the value of analyzing gene regulatory networks in individual cancer patients.

DOI: https://doi.org/10.1158/0008-5472.CAN-21-0730
Lab Invest. 2021 Sep 09.

Anti-tumor effects of vitamin D in glioblastoma: mechanism and therapeutic implications.

Carmen Sze-Ching Lo, Karrie Mei-Yee Kiang, Gilberto Ka-Kit Leung.

Glioma is the most prevalent primary brain tumor in adults among which glioblastoma is the most malignant and lethal subtype. Its common resistance to conventional chemotherapeutics calls for the development of alternative or concomitant treatment. Taking advantage of its endocrine function as a neurosteroid, vitamin D has become a target of interest to be used in conjunction with different chemotherapies. In this article, we review the mechanisms through which vitamin D and its analogs induce anti-tumor activity in glioblastoma, and the practical issues relevant to their potential application based on in vitro and in vivo studies. Vitamin D has largely been reported to promote cell cycle arrest and induce cell death to suppress tumor growth in glioblastoma. Glioblastoma cells treated with vitamin D have also shown reduced migratory and invasive phenotypes, and reduced stemness. It is worth noting that vitamin D analogs are able to produce similar inhibitory actions without causing adverse effects such as hypercalcemia in vivo. Upregulation of vitamin D receptors by vitamin D and its analogs may also play a role in enhancing its anti-tumor activity. Based on current findings and taking into consideration its potential cancer-protective effects, the clinical application of vitamin D in glioblastoma treatment and prevention will be discussed. With some study findings subject to controversy, further investigation is warranted to elucidate the mechanism of action of vitamin D and to evaluate relevant issues regarding its treatment efficacy and potential clinical application.

DOI: https://doi.org/10.1038/s41374-021-00673-8
Lab Invest. 2021 Sep 09.

Grading of adult diffuse gliomas according to the 2021 WHO Classification of Tumors of the Central Nervous System.

Takashi Komori.

The grading of gliomas based on histological features has been a subject of debate for several decades. A consensus has not yet been reached because of technical limitations and inter-observer variations. While the traditional grading system has failed to stratify the risk of IDH-mutant astrocytoma, canonical histological and proliferative markers may be applicable to the risk stratification of IDH-wild-type astrocytoma. Numerous studies have examined molecular markers in order to obtain more clinically relevant information that will improve the risk stratification of gliomas. The CDKN2A/B homozygous deletion for IDH-mutant astrocytoma and the following three criteria for IDH-wild-type astrocytoma: the concurrent gain of whole chromosome 7 and loss of whole chromosome 10, TERT promoter mutations, and EGFR amplification, were identified as independent molecular markers of the worst clinical outcomes. Therefore, the 2021 World Health Organization (WHO) Classification of Tumors of the Central Nervous System adopted these molecular markers into the revised grading criteria of IDH-mutant and -wild-type astrocytoma, respectively, as a grading system within tumor types. Of note, several recent studies have shown that some low-grade IDH-wild-type astrocytoma lacking both the molecular glioblastoma signature and genetic alterations typical of pediatric-type gliomas may demonstrate a relatively indolent clinical course, suggesting the existence of lower-grade adult IDH-wild-type astrocytoma. In terms of oligodendroglioma, IDH-mutant, and 1p/19q codeleted, consistent makers that predict poor outcomes have not yet been identified, and, thus, the current criteria have remained unchanged. Molecular testing to fulfill the revised WHO criteria is, however, not always available worldwide, and in that case, an integrated diagnosis combining all available complementary information is highly recommended. This review discusses controversial issues surrounding legacy grading systems and newly identified potential genetic markers of adult diffuse gliomas and provides perspectives on future grading systems.

DOI: https://doi.org/10.1038/s41374-021-00667-6
Eur J Nutr. 2021 Sep 06.

Short-term fasting in glioma patients: analysis of diet diaries and metabolic parameters of the ERGO2 trial.

Martin Voss, Katharina J Wenger, Nina von Mettenheim, Jörg Bojunga, Manuela Vetter, Bianca Diehl, Kea Franz, Ruediger Gerlach, Michael W Ronellenfitsch, Patrick N Harter, Elke Hattingen, Joachim P Steinbach, Claus Rödel, Johannes Rieger.

   PURPOSE: The prospective, randomized ERGO2 trial investigated the effect of calorie-restricted ketogenic diet and intermittent fasting (KD-IF) on re-irradiation for recurrent brain tumors. The study did not meet its primary endpoint of improved progression-free survival in comparison to standard diet (SD). We here report the results of the quality of life/neurocognition and a detailed analysis of the diet diaries.
METHODS: 50 patients were randomized 1:1 to re-irradiation combined with either SD or KD-IF. The KD-IF schedule included 3 days of ketogenic diet (KD: 21-23 kcal/kg/d, carbohydrate intake limited to 50 g/d), followed by 3 days of fasting and again 3 days of KD. Follow-up included examination of cognition, quality of life and serum samples.
RESULTS: The 20 patients who completed KD-IF met the prespecified goals for calorie and carbohydrate restriction. Substantial decreases in leptin and insulin and an increase in uric acid were observed. The SD group, of note, had a lower calorie intake than expected (21 kcal/kg/d instead of 30 kcal/kg/d). Neither quality of life nor cognition were affected by the diet. Low glucose emerged as a significant prognostic parameter in a best responder analysis.
CONCLUSION: The strict caloric goals of the ERGO2 trial were tolerated well by patients with recurrent brain cancer. The short diet schedule led to significant metabolic changes with low glucose emerging as a candidate marker of better prognosis. The unexpected lower calorie intake of the control group complicates the interpretation of the results. Clinicaltrials.gov number: NCT01754350; Registration: 21.12.2012.

Keywords:  Fasting; Glioblastoma; Glucose; Ketogenic diet; Leptin; Radiation

DOI:  https://doi.org/10.1007/s00394-021-02666-1
PLoS One. 2021 ;16(9): e0256831

Glioblastoma signature in the DNA of blood-derived cells.

Siddharth Jain, Bijan Mazaheri, Netanel Raviv, Jehoshua Bruck.

Current approach for the detection of cancer is based on identifying genetic mutations typical to tumor cells. This approach is effective only when cancer has already emerged, however, it might be in a stage too advanced for effective treatment. Cancer is caused by the continuous accumulation of mutations; is it possible to measure the time-dependent information of mutation accumulation and predict the emergence of cancer? We hypothesize that the mutation history derived from the tandem repeat regions in blood-derived DNA carries information about the accumulation of the cancer driver mutations in other tissues. To validate our hypothesis, we computed the mutation histories from the tandem repeat regions in blood-derived exomic DNA of 3874 TCGA patients with different cancer types and found a statistically significant signal with specificity ranging from 66% to 93% differentiating Glioblastoma patients from other cancer patients. Our approach and findings offer a new direction for future cancer prediction and early cancer detection based on information derived from blood-derived DNA.

DOI: https://doi.org/10.1371/journal.pone.0256831
Proc Natl Acad Sci U S A. 2021 Sep 14. pii: e2103280118. [Epub ahead of print]118(37):

Localized blood-brain barrier opening in infiltrating gliomas with MRI-guided acoustic emissions-controlled focused ultrasound.

Pavlos Anastasiadis, Dheeraj Gandhi, Yutong Guo, Abdul-Kareem Ahmed, Soren M Bentzen, Costas Arvanitis, Graeme F Woodworth.

  Pharmacological treatment of gliomas and other brain-infiltrating tumors remains challenging due to limited delivery of most therapeutics across the blood-brain barrier (BBB). Transcranial MRI-guided focused ultrasound (FUS), an emerging technology for noninvasive brain treatments, enables transient opening of the BBB through acoustic activation of circulating microbubbles. Here, we evaluate the safety and utility of transcranial microbubble-enhanced FUS (MB-FUS) for spatially targeted BBB opening in patients with infiltrating gliomas. In this Phase 0 clinical trial (NCT03322813), we conducted comparative and quantitative analyses of FUS exposures (sonications) and their effects on gliomas using MRI, histopathology, microbubble acoustic emissions (harmonic dose [HD]), and fluorescence-guided surgery metrics. Contrast-enhanced MRI and histopathology indicated safe and reproducible BBB opening in all patients. These observations occurred using a power cycling closed feedback loop controller, with the power varying by nearly an order of magnitude on average. This range underscores the need for monitoring and titrating the exposure on a patient-by-patient basis. We found a positive correlation between microbubble acoustic emissions (HD) and MR-evident BBB opening (P = 0.07) and associated interstitial changes (P < 0.01), demonstrating the unique capability to titrate the MB-FUS effects in gliomas. Importantly, we identified a 2.2-fold increase of fluorescein accumulation in MB-FUS-treated compared to untreated nonenhancing tumor tissues (P < 0.01) while accounting for vascular density. Collectively, this study demonstrates the capabilities of MB-FUS for safe, localized, controlled BBB opening and highlights the potential of this technology to improve the surgical and pharmacologic treatment of brain tumors.

Keywords:  acoustic emissions; blood–brain barrier; focused ultrasound; glioma; microbubbles

DOI:  https://doi.org/10.1073/pnas.2103280118
Acta Neuropathol Commun. 2021 09 08. 9(1): 148

Immune cell deconvolution of bulk DNA methylation data reveals an association with methylation class, key somatic alterations, and cell state in glial/glioneuronal tumors.

Omkar Singh, Drew Pratt, Kenneth Aldape.

  It is recognized that the tumor microenvironment (TME) plays a critical role in the biology of cancer. To better understand the role of immune cell components in CNS tumors, we applied a deconvolution approach to bulk DNA methylation array data in a large set of newly profiled samples (n = 741) as well as samples from external data sources (n = 3311) of methylation-defined glial and glioneuronal tumors. Using the cell-type proportion data as input, we used dimensionality reduction to visualize sample-wise patterns that emerge from the cell type proportion estimations. In IDH-wildtype glioblastomas (n = 2,072), we identified distinct tumor clusters based on immune cell proportion and demonstrated an association with oncogenic alterations such as EGFR amplification and CDKN2A/B homozygous deletion. We also investigated the immune cluster-specific distribution of four malignant cellular states (AC-like, OPC-like, MES-like and NPC-like) in the IDH-wildtype cohort. We identified two major immune-based subgroups of IDH-mutant gliomas, which largely aligned with 1p/19q co-deletion status. Non-codeleted gliomas showed distinct proportions of a key genomic aberration (CDKN2A/B loss) among immune cell-based groups. We also observed significant positive correlations between monocyte proportion and expression of PD-L1 and PD-L2 (R = 0.54 and 0.68, respectively). Overall, the findings highlight specific roles of the TME in biology and classification of CNS tumors, where specific immune cell admixtures correlate with tumor types and genomic alterations.

Keywords:  Deconvolution; Genomic aberrations; Immunotherapy; Tumor microenvironment

DOI:  https://doi.org/10.1186/s40478-021-01249-9