bims-malgli Biomed News
on Biology of malignant gliomas
Issue of 2022‒01‒16
fifteen papers selected by
Oltea Sampetrean
Keio University

  1. Int J Mol Sci. 2021 Dec 28. pii: 297. [Epub ahead of print]23(1):
      Stemness reprogramming remains a largely unaddressed principal cause of lethality in glioblastoma (GBM). It is therefore of utmost importance to identify and target mechanisms that are essential for GBM stemness and self-renewal. Previously, we implicated BIRC3 as an essential mediator of therapeutic resistance and survival adaptation in GBM. In this study, we present novel evidence that BIRC3 has an essential noncanonical role in GBM self-renewal and stemness reprogramming. We demonstrate that BIRC3 drives stemness reprogramming of human GBM cell lines, mouse GBM cell lines and patient-derived GBM stem cells (GSCs) through regulation of BMP4 signaling axis. Specifically, BIRC3 induces stemness reprogramming in GBM through downstream inactivation of BMP4 signaling. RNA-Seq interrogation of the stemness reprogramming hypoxic (pseudopalisading necrosis and perinecrosis) niche in GBM patient tissues further validated the high BIRC3/low BMP4 expression correlation. BIRC3 knockout upregulated BMP4 expression and prevented stemness reprogramming of GBM models. Furthermore, siRNA silencing of BMP4 restored stemness reprogramming of BIRC3 knockout in GBM models. In vivo silencing of BIRC3 suppressed tumor initiation and progression in GBM orthotopic intracranial xenografts. The stemness reprograming of both GSCs and non-GSCs populations highlights the impact of BIRC3 on intra-tumoral cellular heterogeneity GBM. Our study has identified a novel function of BIRC3 that can be targeted to reverse stemness programming of GBM.
    Keywords:  BIRC3; BMP4; GBM; brain tumor; cancer stem cell; stemness
  2. Am J Cancer Res. 2021 ;11(12): 5933-5950
      Glioblastoma multiforme (GBM) is the most common and aggressive form of brain tumors and the hardest type of cancer to treat. Therapies targeting developmental pathways, such as Notch, eliminate neoplastic glioma cells, but their efficacy can be limited by various mechanisms. Combination regimens may represent a good opportunity for effective therapies with durable effects. We used low doses of the γ-secretase inhibitor RO4929097 (GSI), to block the Notch pathway activity, in combination with Resveratrol (RSV) and we evidenced the mechanisms of autophagy/apoptosis transition in GBM cells. Resveratrol and GSI combination results in the synergistic induction of cell death together with the block of the autophagic flux evidenced by a sustained increase of LC3-II and p62 protein content, due to the dramatic reduction of CDK4, an important regulator of lysosomal function. The ectopic overexpression of the constitutive active CDK4 mutant, greatly counteracted the RSV+GSI induced block of the autophagy. Triggering autophagy in RSV+GSI-treated cells, which have impaired lysosomal function, caused the collapse of the system and a following apoptosis. For instance, by combining the CDK4 mutant as well as the early stage autophagy inhibitor, 3-methyladenina, abolished the RSV+GSI induced caspases activation. The initiator caspases (caspases-8 and -9), effector caspase (caspase-3) and its downstream substrate PARP were induced after RSV+GSI exposure as well as the percentage of the TUNEL positive cells. Moreover, the pro-apoptotic signaling MAPK p38 was activated while the pro-survival MAPK p42/p44 signaling was inhibited. In short, we establish the role of CDK4 in the regulation of autophagy/apoptosis transition induced by RSV and GSI in GBM cells. This new synergistic therapeutic combination, increasing the accumulation of autophagosomes, may have therapeutic value for GBM patients.
    Keywords:  Atg12; brain; cancer; cdk4; cell death; gamma secretase inhibitor
  3. Nanomedicine (Lond). 2022 Jan 10.
      Rapid vascular growth, infiltrative cells and high tumor heterogenicity are some glioblastoma multiforme (GBM) characteristics, making it the most lethal form of brain cancer. Low efficacy of the conventional treatment modalities leads to rampant disease progression and a median survival of 15 months. Magnetic nanoparticles (MNPs), due to their unique physical features/inherent abilities, have emerged as a suitable theranostic platform for targeted GBM treatment. Thus, new strategies are being designed to enhance the efficiency of existing therapeutic techniques such as chemotherapy, radiotherapy, and so on, using MNPs. Herein, the limitations of the current therapeutic strategies, the role of MNPs in mitigating those inadequacies, recent advances in the MNP-based theranostics of GBM and possible future directions are discussed.
    Keywords:  chemotherapy; gene therapy; glioblastoma; hyperthermia; imaging; magnetic nanoparticles; radiotherapy
  4. Clin Cancer Res. 2022 Jan 14. pii: clincanres.0947.2021. [Epub ahead of print]
      PURPOSE: Patients with glioblastoma (GBM) are treated with radiation therapy (RT) and temozolomide (TMZ). These treatments may cause prolonged systemic lymphopenia, which itself is associated with poor outcomes. NT-I7 is a long-acting IL-7 that expands CD4 and CD8 T cell numbers in humans and mice. We tested whether NT-I7 prevents systemic lymphopenia and improves survival in mouse models of GBM.EXPERIMENTAL DESIGN: C57BL/6 mice bearing intracranial tumors (GL261 or CT2A) were treated with RT (1.8 Gy/day x 5 days), TMZ (33 mg/kg/day x 5 days), and/or NT-I7 (10 mg/kg on the final day of RT). We followed the mice for survival while serially analyzing levels of circulating T lymphocytes. We assessed regulatory T cells (Treg) and cytotoxic T lymphocytes in the tumor microenvironment, cervical lymph nodes, spleen, and thymus; and hematopoietic stem and progenitor cells (HSPCs) in the bone marrow.
    RESULTS: GBM tumor-bearing mice treated with RT+NT-I7 increased T lymphocytes in the lymph nodes, thymus, and spleen, enhanced IFNγ production, and decreased Treg cells in the tumor which was associated with a significant increase in survival. NT-I7 also enhanced central memory and effector memory CD8 T cells in lymphoid organs and tumor. Depleting CD8 T cells abrogated the effects of NT-I7. Furthermore, NT-I7 treatment decreased progenitor cells in the bone marrow.
    CONCLUSION: In orthotopic glioma-bearing mice, NT-I7 mitigates radiation-related lymphopenia, increases cytotoxic CD8 T lymphocytes systemically and in the tumor, and improves survival. A phase I/II trial to evaluate NT-I7 in patients with high-grade gliomas is ongoing (NCT03687957).
  5. Sci Rep. 2022 Jan 10. 12(1): 424
      Glioma is the most common form of malignant primary brain tumours in adults. Their highly invasive nature makes the disease incurable to date, emphasizing the importance of better understanding the mechanisms driving glioma invasion. Glial fibrillary acidic protein (GFAP) is an intermediate filament protein that is characteristic for astrocyte- and neural stem cell-derived gliomas. Glioma malignancy is associated with changes in GFAP alternative splicing, as the canonical isoform GFAPα is downregulated in higher-grade tumours, leading to increased dominance of the GFAPδ isoform in the network. In this study, we used intravital imaging and an ex vivo brain slice invasion model. We show that the GFAPδ and GFAPα isoforms differentially regulate the tumour dynamics of glioma cells. Depletion of either isoform increases the migratory capacity of glioma cells. Remarkably, GFAPδ-depleted cells migrate randomly through the brain tissue, whereas GFAPα-depleted cells show a directionally persistent invasion into the brain parenchyma. This study shows that distinct compositions of the GFAPnetwork lead to specific migratory dynamics and behaviours of gliomas.
  6. Commun Biol. 2022 Jan 11. 5(1): 22
      Glioma stem cells (GSCs) contribute to the pathogenesis of glioblastoma, the most malignant form of glioma. The implication and underlying mechanisms of SMAD specific E3 ubiquitin protein ligase 2 (SMURF2) on the GSC phenotypes remain unknown. We previously demonstrated that SMURF2 phosphorylation at Thr249 (SMURF2Thr249) activates its E3 ubiquitin ligase activity. Here, we demonstrate that SMURF2Thr249 phosphorylation plays an essential role in maintaining GSC stemness and tumorigenicity. SMURF2 silencing augmented the self-renewal potential and tumorigenicity of patient-derived GSCs. The SMURF2Thr249 phosphorylation level was low in human glioblastoma pathology specimens. Introduction of the SMURF2T249A mutant resulted in increased stemness and tumorigenicity of GSCs, recapitulating the SMURF2 silencing. Moreover, the inactivation of SMURF2Thr249 phosphorylation increases TGF-β receptor (TGFBR) protein stability. Indeed, TGFBR1 knockdown markedly counteracted the GSC phenotypes by SMURF2T249A mutant. These findings highlight the importance of SMURF2Thr249 phosphorylation in maintaining GSC phenotypes, thereby demonstrating a potential target for GSC-directed therapy.
  7. J Immunother Cancer. 2022 Jan;pii: e003289. [Epub ahead of print]10(1):
      PURPOSE: Glioblastoma (GBM) patients suffer from a dismal prognosis, with standard of care therapy inevitably leading to therapy-resistant recurrent tumors. The presence of cancer stem cells (CSCs) drives the extensive heterogeneity seen in GBM, prompting the need for novel therapies specifically targeting this subset of tumor-driving cells. Here, we identify CD70 as a potential therapeutic target for recurrent GBM CSCs.EXPERIMENTAL DESIGN: In the current study, we identified the relevance and functional influence of CD70 on primary and recurrent GBM cells, and further define its function using established stem cell assays. We use CD70 knockdown studies, subsequent RNAseq pathway analysis, and in vivo xenotransplantation to validate CD70's role in GBM. Next, we developed and tested an anti-CD70 chimeric antigen receptor (CAR)-T therapy, which we validated in vitro and in vivo using our established preclinical model of human GBM. Lastly, we explored the importance of CD70 in the tumor immune microenvironment (TIME) by assessing the presence of its receptor, CD27, in immune infiltrates derived from freshly resected GBM tumor samples.
    RESULTS: CD70 expression is elevated in recurrent GBM and CD70 knockdown reduces tumorigenicity in vitro and in vivo. CD70 CAR-T therapy significantly improves prognosis in vivo. We also found CD27 to be present on the cell surface of multiple relevant GBM TIME cell populations, notably putative M1 macrophages and CD4 T cells.
    CONCLUSION: CD70 plays a key role in recurrent GBM cell aggressiveness and maintenance. Immunotherapeutic targeting of CD70 significantly improves survival in animal models and the CD70/CD27 axis may be a viable polytherapeutic avenue to co-target both GBM and its TIME.
    Keywords:  antigens; brain neoplasms; cell engineering; chimeric antigen; immunotherapy; neoplasm; receptors
  8. Clin Cancer Res. 2022 Jan 12. pii: clincanres.2347.2021. [Epub ahead of print]
      PURPOSE: Oncolytic herpes simplex virus-1 (oHSV) infection of brain tumors activates NOTCH, however the consequences of NOTCH on oHSV induced immunotherapy is largely unknown. Here we evaluated the impact of NOTCH blockade on virus induced immunotherapy.EXPERIMENTAL DESIGN: RNA sequencing (RNA-seq), TCGA data analysis, flow cytometry, Luminex- and ELISA-based assays, brain tumor animal models, and serum analysis of recurrent glioblastoma (GBM) patients treated with oHSV was used to evaluate the effect of NOTCH signaling on virus induced immunotherapy.
    RESULTS: TCGA data analysis of patients with grade IV glioma and oHSV treatment of experimental brain tumors in mice showed that NOTCH signaling significantly correlated with a higher myeloid cell infiltration. Immunofluorescence staining and RNA-seq uncovered a significant induction of Jag1 (NOTCH ligand) expression in infiltrating myeloid cells upon oHSV infection. Jag1 expressing macrophages further spread NOTCH activation in the tumor microenvironment (TME). NOTCH activated macrophages increased the secretion of CCL2 which further amplified myeloid derived suppressor cells (MDSCs). CCL2 and IL-10 induction was also observed in serum of recurrent GBM patients treated with oHSV (rQnestin34.5) (NCT03152318). Pharmacologic blockade of NOTCH signaling rescued the oHSV induced immunosuppressive TME and activated a CD8 dependent anti-tumor memory response, resulting in a therapeutic benefit.
    CONCLUSIONS: NOTCH induced immunosuppressive myeloid cell recruitment limited anti-tumor immunity. Translationally, these findings support the use of NOTCH inhibition in conjunction with oHSV therapy.
  9. Nat Commun. 2022 Jan 10. 13(1): 116
      Glioblastoma is an aggressive form of brain cancer with well-established patterns of intra-tumoral heterogeneity implicated in treatment resistance and progression. While regional and single cell transcriptomic variations of glioblastoma have been recently resolved, downstream phenotype-level proteomic programs have yet to be assigned across glioblastoma's hallmark histomorphologic niches. Here, we leverage mass spectrometry to spatially align abundance levels of 4,794 proteins to distinct histologic patterns across 20 patients and propose diverse molecular programs operational within these regional tumor compartments. Using machine learning, we overlay concordant transcriptional information, and define two distinct proteogenomic programs, MYC- and KRAS-axis hereon, that cooperate with hypoxia to produce a tri-dimensional model of intra-tumoral heterogeneity. Moreover, we highlight differential drug sensitivities and relative chemoresistance in glioblastoma cell lines with enhanced KRAS programs. Importantly, these pharmacological differences are less pronounced in transcriptional glioblastoma subgroups suggesting that this model may provide insights for targeting heterogeneity and overcoming therapy resistance.
  10. Dev Cell. 2022 Jan 10. pii: S1534-5807(21)00996-5. [Epub ahead of print]57(1): 32-46.e8
      We test the hypothesis that glioblastoma harbors quiescent cancer stem cells that evade anti-proliferative therapies. Functional characterization of spontaneous glioblastomas from genetically engineered mice reveals essential quiescent stem-like cells that can be directly isolated from tumors. A derived quiescent cancer-stem-cell-specific gene expression signature is enriched in pre-formed patient GBM xenograft single-cell clusters that lack proliferative gene expression. A refined human 118-gene signature is preserved in quiescent single-cell populations from primary and recurrent human glioblastomas. The F3 cell-surface receptor mRNA, expressed in the conserved signature, identifies quiescent tumor cells by antibody immunohistochemistry. F3-antibody-sorted glioblastoma cells exhibit stem cell gene expression, enhance self-renewal in culture, drive tumor initiation and serial transplantation, and reconstitute tumor heterogeneity. Upon chemotherapy, the spared cancer stem cell pool becomes activated and accelerates transition to proliferation. These results help explain conventional treatment failure and lay a conceptual framework for alternative therapies.
    Keywords:  F3 receptor; cancer stem cells; chemoresistance; glioblastoma; heterogeneity; quiescence; recurrence; single-cell RNA sequencing; temozolomide
  11. Neuro Oncol. 2022 Jan 06. pii: noac004. [Epub ahead of print]
      BACKGROUND: There is an extensive literature highlighting the utility of blood-based liquid biopsies in several extracranial tumors for diagnosis and monitoring.METHODS: The RANO (Response Assessment in Neuro-Oncology) group developed a multidisciplinary international Task Force to review the English literature on liquid biopsy in gliomas focusing on the most frequently used techniques, i.e. circulating tumor DNA, circulating tumor cells and extracellular vesicles in blood and CSF.
    RESULTS: ctDNA has a higher sensitivity and capacity to represent the spatial and temporal heterogeneity in comparison to circulating tumor cells. Exosomes have the advantages to cross an intact blood-brain barrier and carry also RNA, miRNA and proteins. Several clinical applications of liquid biopsies are suggested: to establish a diagnosis when tissue is not available, monitor the residual disease after surgery, distinguish progression from pseudoprogression, and predict outcome.
    CONCLUSIONS: There is need for standardization of biofluid collection, choice of analyte, and detection strategies along with rigorous testing in future clinical trials to validate findings and enable entry into clinical practice.
    Keywords:  CSF; circulating tumor cells; ctDNA; extracellular vesicles; gliomas
  12. Nat Protoc. 2022 Jan 12.
      When modeling disease in the laboratory, it is important to use clinically relevant models. Patient-derived human brain cells grown in vitro to study and test potential treatments provide such a model. Here, we present simple, highly reproducible coordinated procedures that can be used to routinely culture most cell types found in the human brain from single neurosurgically excised brain specimens. The cell types that can be cultured include dissociated cultures of neurons, astrocytes, microglia, pericytes and brain endothelial and neural precursor cells, as well as explant cultures of the leptomeninges, cortical slice cultures and brain tumor cells. The initial setup of cultures takes ~2 h, and the cells are ready for further experiments within days to weeks. The resulting cells can be studied as purified or mixed population cultures, slice cultures and explant-derived cultures. This protocol therefore enables the investigation of human brain cells to facilitate translation of neuroscience research to the clinic.