ABSTRACT
Homozygous deletion of CDKN2A/B was recently incorporated into the World Health Organization classification for grade 3 meningiomas. While this marker is overall rare in meningiomas, its relationship to other CDKN2A alterations on a transcriptomic, epigenomic, and copy number level has not yet been determined. We therefore utilized multidimensional molecular data of 1577 meningioma samples from 6 independent cohorts enriched for clinically aggressive meningiomas to comprehensively interrogate the spectrum of CDKN2A alterations through DNA methylation, copy number variation, transcriptomics, and proteomics using an integrated molecular approach. Homozygous CDKN2A/B deletions were identified in only 7.1% of cases but were associated with significantly poorer outcomes compared to tumors without these deletions. Heterozygous CDKN2A/B deletions were identified in 2.6% of cases and had similarly poor outcomes as those with homozygous deletions. Among tumors with intact CDKN2A/B (without a homozygous or heterozygous deletion), we found a distinct difference in outcome based on mRNA expression of CDKN2A, with meningiomas that had elevated mRNA expression (CDKN2Ahigh) having a significantly shorter time to recurrence. The expression of CDKN2A was independently prognostic after accounting for copy number loss and consistently increased with WHO grade and more aggressive molecular and methylation groups irrespective of cohort. Despite the discordant and mutually exclusive status of the CDKN2A gene in these groups, both CDKN2Ahigh meningiomas and meningiomas with CDKN2A deletions were enriched for similar cell cycle pathways but at different checkpoints. High mRNA expression of CDKN2A was also associated with gene hypermethylation, Rb-deficiency, and lack of response to CDK inhibition. p16 immunohistochemistry could not reliably differentiate between meningiomas with and without CDKN2A deletions but appeared to correlate better with mRNA expression. These findings support the role of CDKN2A mRNA expression as a biomarker of clinically aggressive meningiomas with potential therapeutic implications.
Subject(s)
Meningeal Neoplasms , Meningioma , Humans , Genes, p16 , Meningioma/genetics , Cyclin-Dependent Kinase Inhibitor p16/genetics , Transcriptome , DNA Copy Number Variations , Homozygote , Sequence Deletion , Meningeal Neoplasms/geneticsABSTRACT
INTRODUCTION: Meningioma is the most common primary brain tumor. Most meningiomas are benign; however, a subset of these tumors can be aggressive, presenting with early or multiple tumor recurrences that are refractory to neurosurgical resection and radiotherapy. There is no standard systemic therapy for these patients, and post-surgical management of these patients is usually complicated due to lack of accurate prediction for tumor progression. METHODS: In this review, we summarise the crucial immunosuppressive role of checkpoint regulators, including PD-1 and PD-L1 interacting in the tumor microenvironment, which has led to efforts aimed at targeting this axis. RESULTS: Since their discovery, checkpoint inhibitors have significantly improved the outcome in many types of cancers. Currently, targeted therapy for PD-1 and PD-L1 proteins are being tested in several ongoing clinical trials for brain tumors such as glioblastoma. More recently, there have been some reports implicating increased PD-L1 expression in high-grade (WHO grades II and III) meningiomas. Several clinical trials are underway to assess the efficacy of checkpoint inhibitors in the therapeutic management of patients with aggressive meningiomas. Here, we review the immune suppressive microenvironment in meningiomas, and then focus on clinical and pathological characterization and tumor heterogeneity with respect to PD-L1 expression as well as challenges associated with the assessment of PD-L1 expression in meningioma. CONCLUSION: We conclude with a brief review of ongoing clinical trials using checkpoint inhibitors for the treatment of high-grade and refractory meningiomas.
Subject(s)
B7-H1 Antigen/genetics , Brain Neoplasms/genetics , Genes, cdc/genetics , Meningioma/genetics , Animals , B7-H1 Antigen/biosynthesis , Brain Neoplasms/pathology , Brain Neoplasms/therapy , Humans , Immunohistochemistry , Meningioma/pathology , Meningioma/therapyABSTRACT
Liquid biopsy, as a non-invasive technique for cancer diagnosis, has emerged as a major step forward in conquering tumors. Current practice in diagnosis of central nervous system (CNS) tumors involves invasive acquisition of tumor biopsy upon detection of tumor on neuroimaging. Liquid biopsy enables non-invasive, rapid, precise and, in particular, real-time cancer detection, prognosis and treatment monitoring, especially for CNS tumors. This approach can also uncover the heterogeneity of these tumors and will likely replace tissue biopsy in the future. Key components of liquid biopsy mainly include circulating tumor cells (CTC), circulating tumor nucleic acids (ctDNA, miRNA) and exosomes and samples can be obtained from the cerebrospinal fluid, plasma and serum of patients with CNS malignancies. This review covers current progress in application of liquid biopsies for diagnosis and monitoring of CNS malignancies.
Subject(s)
Central Nervous System Neoplasms/diagnosis , Liquid Biopsy/methods , Liquid Biopsy/trends , Biomarkers, Tumor/blood , Biomarkers, Tumor/cerebrospinal fluid , Central Nervous System Neoplasms/metabolism , Circulating Tumor DNA/blood , Exosomes/pathology , Humans , MicroRNAs , Neoplastic Cells, Circulating/pathology , PrognosisABSTRACT
von Hippel-Lindau (VHL) disease is a rare familial cancer predisposition syndrome caused by a loss or mutation in a single gene,VHL, but it exhibits a wide phenotypic variability that can be categorized into distinct subtypes. The phenotypic variability has been largely argued to be attributable to the extent of deregulation of the α subunit of hypoxia-inducible factor α, a well established target of VHL E3 ubiquitin ligase, ECV (Elongins/Cul2/VHL). Here, we show that erythropoietin receptor (EPOR) is hydroxylated on proline 419 and 426 via prolyl hydroxylase 3. EPOR hydroxylation is required for binding to the ß domain of VHL and polyubiquitylation via ECV, leading to increased EPOR turnover. In addition, several type-specific VHL disease-causing mutants, including those that have retained proper binding and regulation of hypoxia-inducible factor α, showed a severe defect in binding prolyl hydroxylated EPOR peptides. These results identify EPOR as the secondbona fidehydroxylation-dependent substrate of VHL that potentially influences oxygen homeostasis and contributes to the complex genotype-phenotype correlation in VHL disease.
Subject(s)
Oxygen/metabolism , Proteolysis , Receptors, Erythropoietin/metabolism , Signal Transduction , Ubiquitination , Von Hippel-Lindau Tumor Suppressor Protein/metabolism , Basic Helix-Loop-Helix Transcription Factors/genetics , Basic Helix-Loop-Helix Transcription Factors/metabolism , Cullin Proteins/genetics , Cullin Proteins/metabolism , HEK293 Cells , Humans , Receptors, Erythropoietin/genetics , Von Hippel-Lindau Tumor Suppressor Protein/genetics , von Hippel-Lindau Disease/genetics , von Hippel-Lindau Disease/metabolism , von Hippel-Lindau Disease/pathologyABSTRACT
Mutations in Ras GTPase and various other components of the Ras signaling pathways are among the most common genetic alterations in human cancers and also have been identified in several familial developmental syndromes. Over the past few decades it has become clear that the activity or the oncogenic potential of Ras is dependent on the nonreceptor tyrosine kinase Src to promote the Ras/Raf/MAPK pathway essential for proliferation, differentiation, and survival of eukaryotic cells. However, no direct relationship between Ras and Src has been established. We show here that Src binds to and phosphorylates GTP-, but not GDP-, loaded Ras on a conserved Y32 residue within the switch I region in vitro and that in vivo, Ras-Y32 phosphorylation markedly reduces the binding to effector Raf and concomitantly increases binding to GTPase-activating proteins and the rate of GTP hydrolysis. These results suggest that, in the context of predetermined crystallographic structures, Ras-Y32 serves as an Src-dependent keystone regulatory residue that modulates Ras GTPase activity and ensures unidirectionality to the Ras GTPase cycle.
Subject(s)
GTP Phosphohydrolases/metabolism , Membrane Proteins/metabolism , Phosphotyrosine/metabolism , src-Family Kinases/metabolism , Amino Acid Sequence , Animals , Cell Line , GTP Phosphohydrolases/chemistry , GTPase-Activating Proteins/metabolism , Guanosine Triphosphate/metabolism , Humans , Hydrolysis , Membrane Proteins/chemistry , Mice , Models, Molecular , Molecular Sequence Data , Mutant Proteins/metabolism , Phosphorylation , Protein Binding , raf Kinases/metabolismABSTRACT
BACKGROUND: Diffuse gliomas represent over 80% of malignant brain tumors ranging from low-grade to aggressive high-grade lesions. Within isocitrate dehydrogenase (IDH)-mutant gliomas, there is a high variability in survival and a need to more accurately predict outcome. METHODS: To identify and characterize a predictive signature of outcome in gliomas, we utilized an integrative molecular analysis (using methylation, mRNA, copy number variation (CNV), and mutation data), analyzing a total of 729 IDH-mutant samples including a test set of 99 from University Health Network (UHN) and 2 validation cohorts including the German Cancer Research Center (DKFZ) and The Cancer Genome Atlas (TCGA). RESULTS: Cox regression analysis of methylation data from the UHN cohort identified CpG-based signatures that split the glioma cohort into 2 prognostic groups strongly predicting survival that were validated using 2 independent cohorts from TCGA and DKFZ (all P-valuesâ <â .0001). The methylation signatures that predicted poor outcomes also exhibited high CNV instability and hypermethylation of HOX gene probes. Integrated multi-platform analyses using mRNA and methylation (iRM) showed that parallel HOX gene overexpression and simultaneous hypermethylation were significantly associated with increased mutational load, high aneuploidy, and worse survival (P-valueâ <â .0001). A 7-HOX gene signature was developed and validated using the most significantly associated HOX genes with patient outcome in both 1p/19q codeleted and non-codeleted IDHmut gliomas. CONCLUSIONS: HOX gene methylation and expression provide important prognostic information in IDH-mutant gliomas that are not captured by current molecular diagnostics. A 7-HOX gene signature of outcome shows significant survival differences in both 1p/19q codeleted and non-codeleted IDH-mutant gliomas.
Subject(s)
Brain Neoplasms , Glioma , Humans , Genes, Homeobox , Isocitrate Dehydrogenase/genetics , DNA Copy Number Variations , Glioma/pathology , Brain Neoplasms/pathology , Mutation , RNA, MessengerABSTRACT
Malignant peripheral nerve sheath tumor (MPNST) is a highly aggressive sarcoma, and a lethal neurofibromatosis type 1-related malignancy, with little progress made on treatment strategies. Here, we apply a multiplatform integrated molecular analysis on 108 tumors spanning the spectrum of peripheral nerve sheath tumors to identify candidate drivers of MPNST that can serve as therapeutic targets. Unsupervised analyses of methylome and transcriptome profiles identify two distinct subgroups of MPNSTs with unique targetable oncogenic programs. We establish two subgroups of MPNSTs: SHH pathway activation in MPNST-G1 and WNT/ß-catenin/CCND1 pathway activation in MPNST-G2. Single nuclei RNA sequencing characterizes the complex cellular architecture and demonstrate that malignant cells from MPNST-G1 and MPNST-G2 have neural crest-like and Schwann cell precursor-like cell characteristics, respectively. Further, in pre-clinical models of MPNST we confirm that inhibiting SHH pathway in MPNST-G1 prevent growth and malignant progression, providing the rational for investigating these treatments in clinical trials.
Subject(s)
Nerve Sheath Neoplasms , Neurofibromatosis 1 , Neurofibrosarcoma , Humans , Neurofibrosarcoma/genetics , Neurofibrosarcoma/metabolism , Nerve Sheath Neoplasms/genetics , Nerve Sheath Neoplasms/metabolism , Nerve Sheath Neoplasms/pathology , Neurofibromatosis 1/genetics , Schwann Cells/metabolism , Wnt Signaling Pathway/geneticsABSTRACT
The mechanism that leads to the inverse relationship between heightened cellular proliferation and the cessation of elastic fibers production, observed during formation of the arterial occlusions and dermal scars, is not fully understood. Because the retinoblastoma protein (Rb), responsible for cell cycle initiation, has also been implicated in insulin-like growth factor-I-mediated signaling stimulating elastin gene activation, we explored whether differential phosphorylation of Rb by various cyclin·cyclin-dependent kinase complexes would be responsible for promoting either elastogenic or pro-proliferative signals. We first tested cultures of dermal fibroblasts derived from Costello syndrome patients, in which heightened proliferation driven by mutated oncogenic H-Ras coincides with inhibition of elastogenesis. We found that Costello syndrome fibroblasts display elevated level of Rb phosphorylation on serine 780 (Ser(P)-780-Rb) and that pharmacological inhibition of Ras with radicicol, Mek/Erk with PD98059, or cyclin-dependent kinase 4 with PD0332991 not only leads to down-regulation of Ser(P)-780-Rb levels but also enhances Rb phosphorylation on threonine-821 (Thr(P)-821-Rb), which coincides with the recovery of elastin production. Then we demonstrated that treatment of normal skin fibroblasts with the pro-proliferative PDGF BB also up-regulates Ser(P)-780-Rb levels, but treatment with the pro-elastogenic insulin-like growth factor-I activates cyclinE-cdk2 complex to phosphorylate Rb on Thr-821. Importantly, we have established that elevation of Thr(P)-821-Rb promotes Rb binding to the Sp1 transcription factor and that successive binding of the Rb-Sp1 complex to the retinoblastoma control element within the elastin gene promoter stimulates tropoelastin transcription. In summary, we provide novel insight into the role of Rb in mediating the inverse relationship between elastogenesis and cellular proliferation.
Subject(s)
Cell Proliferation , Dermis/metabolism , Elastin/biosynthesis , Fibroblasts/metabolism , Retinoblastoma Protein/metabolism , Adolescent , Adult , Becaplermin , Cells, Cultured , Costello Syndrome , Dermis/pathology , Elastin/genetics , Extracellular Signal-Regulated MAP Kinases/antagonists & inhibitors , Extracellular Signal-Regulated MAP Kinases/genetics , Extracellular Signal-Regulated MAP Kinases/metabolism , Female , Fibroblasts/pathology , Flavonoids/pharmacology , Humans , Insulin-Like Growth Factor I/genetics , Insulin-Like Growth Factor I/metabolism , Phosphorylation/genetics , Platelet-Derived Growth Factor/genetics , Platelet-Derived Growth Factor/metabolism , Proto-Oncogene Proteins c-sis , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Retinoblastoma Protein/genetics , Sp1 Transcription Factor/genetics , Sp1 Transcription Factor/metabolism , Up-Regulation/drug effects , Up-Regulation/geneticsABSTRACT
The results of our in vitro experiments indicate that exposing cultured human aortic smooth muscle cells and dermal fibroblasts to 39 to 41 °C induces a significant up-regulation in the net deposition of elastic fibers, but not of collagen I or fibronectin, and also decreases the deposition of chondroitin sulfate-containing moieties. We further demonstrate that mild hyperthermia also rectifies the insufficient elastogenesis notable in cultures of fibroblasts derived from the stretch-marked skin of adult patients and in cultures of dermal fibroblasts from children with Costello syndrome, which is characterized by the accumulation of chondroitin 6-sulfate glycosaminoglycans that induce shedding and inactivation of the 67-kDa elastin-binding protein. We have previously established that this protein serves as a reusable chaperone for tropoelastin and that its recycling is essential for the normal deposition of elastic fibers. We now report that hyperthermia not only inhibits deposition of chondroitin 6-sulfate moieties and the consequent preservation of elastin-binding protein molecules but also induces their faster recycling. This, in turn, triggers a more efficient preservation of tropoelastin, enhancement of its secretion and extracellular assembly into elastic fibers. The presented results encourage using mild hyperthermia to restore elastic fiber production in damaged adult skin and to enhance elastogenesis in children with genetic elastinopathies.
Subject(s)
Costello Syndrome/metabolism , Elastic Tissue/metabolism , Fibroblasts/metabolism , Heat-Shock Response , Receptors, Cell Surface/metabolism , Tropoelastin/metabolism , Adult , Cells, Cultured , Child , Child, Preschool , Costello Syndrome/pathology , Elastic Tissue/pathology , Fibroblasts/pathology , Glycosaminoglycans , Humans , Infant , MaleABSTRACT
One of the most prominent features of glioblastoma (GBM) is hyper-vascularization. Bone marrow-derived macrophages are actively recruited to the tumor and referred to as glioma-associated macrophages (GAMs) which are thought to provide a critical role in tumor neo-vascularization. However, the mechanisms by which GAMs regulate endothelial cells (ECs) in the process of tumor vascularization and response to anti-angiogenic therapy (AATx) is not well-understood. Here we show that GBM cells secrete IL-8 and CCL2 which stimulate GAMs to produce TNFα. Subsequently, TNFα induces a distinct gene expression signature of activated ECs including VCAM-1, ICAM-1, CXCL5, and CXCL10. Inhibition of TNFα blocks GAM-induced EC activation both in vitro and in vivo and improve survival in mouse glioma models. Importantly we show that high TNFα expression predicts worse response to Bevacizumab in GBM patients. We further demonstrated in mouse model that treatment with B20.4.1.1, the mouse analog of Bevacizumab, increased macrophage recruitment to the tumor area and correlated with upregulated TNFα expression in GAMs and increased EC activation, which may be responsible for the failure of AATx in GBMs. These results suggest TNFα is a novel therapeutic that may reverse resistance to AATx. Future clinical studies should be aimed at inhibiting TNFα as a concurrent therapy in GBMs.
Subject(s)
Brain Neoplasms/pathology , Drug Resistance, Neoplasm/physiology , Glioma/pathology , Macrophages/metabolism , Neovascularization, Pathologic/metabolism , Tumor Necrosis Factor-alpha/metabolism , Angiogenesis Inhibitors/pharmacology , Animals , Brain Neoplasms/metabolism , Endothelial Cells/metabolism , Glioma/metabolism , Humans , Mice , Neovascularization, Pathologic/pathology , Xenograft Model Antitumor AssaysABSTRACT
We recently established that the subunit of cell surface-residing elastin receptor, neuraminidase-1 (Neu1), can desialylate adjacent insulin-like growth factor 1 receptors (IGF-1R) of arterial smooth muscle cells, thereby quenching their proliferative response to insulin-like growth factor II. In this study, we explored whether Neu1 would also desialylate the insulin receptors (IR), as well as the IGF-1R on rat skeletal L6 myoblasts, and whether desialylation of IR and IGF-1R would affect a net proliferative effect of insulin. First, we found that physiological (0.5-1 nM) and high therapeutic (10 nM) insulin concentrations induced a modest increase in proliferation rate of cultured L6 myoblasts. While IR kinase inhibitor could abolish the mitogenic effect of these insulin concentrations, the observed more pronounced proliferative response to supraphysiological concentration (100 nM) of insulin could be eliminated only by specific inhibition of IGF-1R. Then, we found that treatment of L6 cells with mouse-derived Neu1 or with Clostridium perfringens neuraminidase caused desialylation of IR, which coincided with a significant increase of their proliferative response to lower (0.5-10 nM) concentrations of insulin. In contrast, experimental desialylation of IGF-1R coincided with elimination of the heightened proliferative response of L6 myoblasts to 100 nM insulin. Importantly, we also found that inhibition of endogenous Neu1 abolished the increase in proliferation of L6 cells induced by 1 and 10 nM of insulin, but amplified the proliferative effect of 100 nM insulin. We therefore conclude that desialylation of both IR and IGF-1R by Neu1 controls the net proliferative response of skeletal myoblasts to insulin.
Subject(s)
Insulin/pharmacology , Myoblasts/cytology , Myoblasts/drug effects , Receptor, IGF Type 1/metabolism , Receptor, Insulin/metabolism , Repressor Proteins/metabolism , Sialic Acids/metabolism , Animals , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Mice , Myoblasts/metabolism , Rats , Receptor, IGF Type 1/chemistry , Receptor, Insulin/chemistry , Ubiquitin-Protein LigasesABSTRACT
Capicua (CIC) is a transcriptional repressor that counteracts activation of genes in response to receptor tyrosine kinase (RTK)/Ras/ERK signaling. Following activation of RTK, ERK enters the nucleus and serine-phosphorylates CIC, releasing it from its targets to permit gene expression. We recently showed that ERK triggers ubiquitin-mediated degradation of CIC in glioblastoma (GBM). In this study, we examined whether another important downstream effector of RTK/EGFR, the non-RTK c-Src, affects CIC repressor function in GBM. We found that c-Src binds and tyrosine-phosphorylates CIC on residue 1455 to promote nuclear export of CIC. On the other hand, CIC-mutant allele (CIC-Y1455F), that escapes c-Src-mediated tyrosine phosphorylation, remains localized to the nucleus and retains strong repressor function against CIC targets, the oncogenic transcription factors ETV1 and ETV5. Furthermore, we show that the orally available Src family kinase inhibitor, dasatinib, which prevents EGF-mediated tyrosine phosphorylation of CIC and attenuates elevated ETV1 and ETV5 levels, reduces viability of GBM cells and glioma stem cells (GSC), but not of their control cells with undetectable c-Src activity. In fact, GBM cells and GSC expressing the tyrosine-defective CIC mutant (Y1455F) lose sensitivity to dasatinib, further endorsing the effect of dasatinib on Src-mediated tyrosine phosphorylation of CIC. These findings elucidate important mechanisms of CIC regulation and provide the rationale to target c-Src alongside ERK pathway inhibitors as a way to fully restore CIC tumor suppressor function in neoplasms such as GBM. IMPLICATIONS: c-Src tyrosine-phosphorylates CIC exports to cytoplasm and inactivates its repressor function in GBM.
Subject(s)
Biomarkers, Tumor/metabolism , CSK Tyrosine-Protein Kinase/metabolism , Gene Expression Regulation, Neoplastic , Glioblastoma/pathology , Repressor Proteins/antagonists & inhibitors , Animals , Apoptosis , Biomarkers, Tumor/genetics , CSK Tyrosine-Protein Kinase/genetics , Cell Proliferation , Dasatinib/pharmacology , Glioblastoma/drug therapy , Glioblastoma/genetics , Glioblastoma/metabolism , Humans , Male , Mice , Mice, Inbred NOD , Mice, SCID , Phosphorylation , Protein Kinase Inhibitors/pharmacology , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
We recently established that the elastin-binding protein, which is identical to the spliced variant of beta-galactosidase, forms a cell surface-targeted complex with two proteins considered "classic lysosomal enzymes": protective protein/cathepsin A and neuraminidase-1 (Neu1). We also found that cell surface-residing Neu1 can desialylate neighboring microfibrillar glycoproteins and facilitate the deposition of insoluble elastin, which contributes to the maintenance of cellular quiescence. Here we provide evidence that cell surface-residing Neu1 contributes to a novel mechanism that limits cellular proliferation by desialylating cell membrane-residing sialoglycoproteins that directly propagate mitogenic signals. We demonstrated that treatment of cultured human aortic smooth muscle cells (SMCs) with either a sialidase inhibitor or an antibody that blocks Neu1 activity induced significant up-regulation in SMC proliferation in response to fetal bovine serum. Conversely, treatment with Clostridium perfringens neuraminidase (which is highly homologous to Neu1) decreased SMC proliferation, even in cultures that did not deposit elastin. Further, we found that pretreatment of aortic SMCs with exogenous neuraminidase abolished their mitogenic responses to recombinant platelet-derived growth factor (PDGF)-BB and insulin-like growth factor (IGF)-2 and that sialidosis fibroblasts (which are exclusively deficient in Neu1) were more responsive to PDGF-BB and IGF-2 compared with normal fibroblasts. Furthermore, we provide direct evidence that neuraminidase caused the desialylation of both PDGF and IGF-1 receptors and diminished the intracellular signals induced by the mitogenic ligands PDGF-BB and IGF-2.
Subject(s)
Cell Membrane/metabolism , Insulin-Like Growth Factor II/metabolism , N-Acetylneuraminic Acid/metabolism , Neuraminidase/metabolism , Platelet-Derived Growth Factor/metabolism , Protein Subunits/metabolism , Receptors, Cell Surface/metabolism , Animals , Aorta/cytology , Aorta/enzymology , Becaplermin , Cell Membrane/drug effects , Cell Proliferation/drug effects , Cells, Cultured , Child, Preschool , Elastic Tissue/drug effects , Elastic Tissue/enzymology , Elastin/metabolism , Fibroblasts/drug effects , Fibroblasts/enzymology , Fibroblasts/pathology , Humans , Infant , Insulin-Like Growth Factor II/pharmacology , Mitogens/metabolism , Mitogens/pharmacology , Mucolipidoses/enzymology , Mucolipidoses/pathology , Myocytes, Smooth Muscle/cytology , Myocytes, Smooth Muscle/drug effects , Myocytes, Smooth Muscle/enzymology , Neuraminidase/antagonists & inhibitors , Neuraminidase/pharmacology , Platelet-Derived Growth Factor/pharmacology , Proto-Oncogene Proteins c-sis , Receptor, IGF Type 1/metabolism , Receptors, Platelet-Derived Growth Factor/metabolism , Recombinant Proteins/pharmacology , SwineABSTRACT
Capicua (CIC) is a transcriptional repressor that counteracts activation of genes downstream of receptor tyrosine kinase (RTK)/Ras/ERK signaling. It is well-established that tumorigenesis, especially in glioblastoma (GBM), is attributed to hyperactive RTK/Ras/ERK signaling. While CIC is mutated in other tumors, here we show that CIC has a tumor suppressive function in GBM through an alternative mechanism. We find that CIC protein levels are negligible in GBM due to continuous proteasome-mediated degradation, which is mediated by the E3 ligase PJA1 and show that this occurs through binding of CIC to its DNA target and phosphorylation on residue S173. PJA1 knockdown increased CIC stability and extended survival using in-vivo models of GBM. Deletion of the ERK binding site resulted in stabilization of CIC and increased therapeutic efficacy of ERK inhibition in GBM models. Our results provide a rationale to target CIC degradation in Ras/ERK-driven tumors, including GBM, to increase efficacy of ERK inhibitors.
Subject(s)
Glioblastoma/metabolism , Glioblastoma/pathology , Repressor Proteins/metabolism , Animals , Carcinogenesis/metabolism , Carcinogenesis/pathology , Cell Line, Tumor , Cell Proliferation/genetics , Cell Proliferation/physiology , Cell Transformation, Neoplastic/metabolism , Cell Transformation, Neoplastic/pathology , Drosophila , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Gene Expression Regulation, Developmental/genetics , Gene Expression Regulation, Developmental/physiology , HEK293 Cells , Humans , Male , Mice, Inbred C57BL , Mice, Transgenic , Mutation , Repressor Proteins/genetics , Signal Transduction/genetics , Signal Transduction/physiologyABSTRACT
PURPOSE: Hexokinase II (HK2) protein expression is elevated in glioblastoma (GBM), and we have shown that HK2 could serve as an effective therapeutic target for GBM. Here, we interrogated compounds that target HK2 effectively and restrict tumor growth in cell lines, patient-derived glioma stem cells (GSCs), and mouse models of GBM.Experimental Design: We performed a screen using a set of 15 drugs that were predicted to inhibit the HK2-associated gene signature. We next determined the EC50 of the compounds by treating glioma cell lines and GSCs. Selected compounds showing significant impact in vitro were used to treat mice and examine their effect on survival and tumor characteristics. The effect of compounds on the metabolic activity in glioma cells was also assessed in vitro. RESULTS: This screen identified the azole class of antifungals as inhibitors of tumor metabolism. Among the compounds tested, ketoconazole and posaconazole displayed the greatest inhibitory effect on GBM both in vitro and in vivo. Treatment of mice bearing GBM with ketoconazole and posaconazole increased their survival, reduced tumor cell proliferation, and decreased tumor metabolism. In addition, treatment with azoles resulted in increased proportion of apoptotic cells. CONCLUSIONS: Overall, we provide evidence that azoles exert their effect by targeting genes and pathways regulated by HK2. These findings shed light on the action of azoles in GBM. Combined with existing literature and preclinical results, these data support the value of repurposing azoles in GBM clinical trials.
Subject(s)
Antineoplastic Agents/pharmacology , Hexokinase/antagonists & inhibitors , Ketoconazole/pharmacology , Triazoles/pharmacology , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Disease Models, Animal , Dose-Response Relationship, Drug , Energy Metabolism/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Glioblastoma/genetics , Glioblastoma/metabolism , Hexokinase/genetics , Hexokinase/metabolism , Humans , Male , Mice , Xenograft Model Antitumor AssaysABSTRACT
The link between GBM molecular subtype and response to treatment remains undefined. In this issue of Cancer Cell, Cosset and colleagues define a subpopulation of patients within the proneural/classical subtype sensitive to integrin blockade because of a Glut3 addiction. These findings reveal context-dependent druggable vulnerability in a subpopulation of GBM.
Subject(s)
Brain Neoplasms , Glioblastoma , Adult , HumansABSTRACT
Pseudoxanthoma elasticum (PXE) is a heritable disorder mainly characterized by calcified elastic fibers in cutaneous, ocular, and vascular tissues. PXE is caused by mutations in ABCC6, a gene encoding an ABC transporter predominantly expressed in liver and kidneys. The functional relationship between ABCC6 and elastic fiber calcification is unknown. We speculated that ABCC6 deficiency in PXE patients induces a persistent imbalance in circulating metabolite(s), which may impair the synthetic abilities of normal elastoblasts or specifically alter elastic fiber assembly. Therefore, we compared the deposition of elastic fiber proteins in cultures of fibroblasts derived from PXE and unaffected individuals. PXE fibroblasts cultured with normal human serum expressed and deposited increased amounts of proteins, but structurally normal elastic fibers. Interestingly, normal and PXE fibroblasts as well as normal smooth muscle cells deposited abnormal aggregates of elastic fibers when maintained in the presence of serum from PXE patients. The expression of tropoelastin and other elastic fiber-associated genes was not significantly modulated by the presence of PXE serum. These results indicated that certain metabolites present in PXE sera interfered with the normal assembly of elastic fibers in vitro and suggested that PXE is a primary metabolic disorder with secondary connective tissue manifestations.
Subject(s)
Blood Proteins/pharmacology , Elastin/metabolism , Multidrug Resistance-Associated Proteins/genetics , Pseudoxanthoma Elasticum/blood , Pseudoxanthoma Elasticum/physiopathology , Blood Proteins/analysis , Cardiovascular System/pathology , Cardiovascular System/physiopathology , Cells, Cultured , Child , Connective Tissue/pathology , Connective Tissue/physiopathology , Elastic Tissue/chemistry , Eye/pathology , Eye/physiopathology , Female , Fibroblasts/chemistry , Fibroblasts/pathology , Gene Expression Regulation , Humans , Male , Metabolic Diseases/blood , Metabolic Diseases/etiology , Metabolic Diseases/genetics , Metabolic Diseases/physiopathology , Multidrug Resistance-Associated Proteins/deficiency , Multidrug Resistance-Associated Proteins/physiology , Muscle, Smooth/cytology , Muscle, Smooth/physiopathology , Mutation , Pseudoxanthoma Elasticum/genetics , Pseudoxanthoma Elasticum/metabolism , Skin/pathology , Skin/physiopathology , Tropoelastin/genetics , Tropoelastin/physiologyABSTRACT
Ras is phosphorylated on a conserved tyrosine at position 32 within the switch I region via Src kinase. This phosphorylation inhibits the binding of effector Raf while promoting the engagement of GTPase-activating protein (GAP) and GTP hydrolysis. Here we identify SHP2 as the ubiquitously expressed tyrosine phosphatase that preferentially binds to and dephosphorylates Ras to increase its association with Raf and activate downstream proliferative Ras/ERK/MAPK signalling. In comparison to normal astrocytes, SHP2 activity is elevated in astrocytes isolated from glioblastoma multiforme (GBM)-prone H-Ras(12V) knock-in mice as well as in glioma cell lines and patient-derived GBM specimens exhibiting hyperactive Ras. Pharmacologic inhibition of SHP2 activity attenuates cell proliferation, soft-agar colony formation and orthotopic GBM growth in NOD/SCID mice and decelerates the progression of low-grade astrocytoma to GBM in a spontaneous transgenic glioma mouse model. These results identify SHP2 as a direct activator of Ras and a potential therapeutic target for cancers driven by a previously 'undruggable' oncogenic or hyperactive Ras.
Subject(s)
Glioblastoma/enzymology , Oncogene Protein p21(ras)/metabolism , Protein Tyrosine Phosphatase, Non-Receptor Type 11/metabolism , Animals , Carcinogenesis , Cell Line , Female , Glioblastoma/genetics , Glioblastoma/metabolism , Glioblastoma/pathology , Humans , Male , Mice , Mice, Inbred ICR , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Oncogene Protein p21(ras)/genetics , Phosphorylation , Protein Tyrosine Phosphatase, Non-Receptor Type 11/geneticsABSTRACT
Granulocyte-macrophage colony-stimulating factor (GM-CSF) and the related cytokines interleukin (IL)-3 and IL-5 regulate the production and functional activation of hematopoietic cells. GM-CSF acts on monocytes/macrophages and granulocytes, and several chronic inflammatory diseases and a number of haematological malignancies such as Juvenile myelomonocytic leukaemia (JMML) are associated with deregulated GM-CSF receptor (GMR) signaling. The downregulation of GMR downstream signaling is mediated in part by the clearance of activated GMR via the proteasome, which is dependent on the ubiquitylation of ßc signaling subunit of GMR via an unknown E3 ubiquitin ligase. Here, we show that suppressor of cytokine signaling 1 (SOCS-1), best known for its ability to promote ubiquitin-mediated degradation of the non-receptor tyrosine kinase Janus kinase 2 (JAK2), also targets GMRßc for ubiquitin-mediated degradation and attenuates GM-CSF-induced downstream signaling.