ABSTRACT
The deadly complication of brain metastasis (BM) is largely confined to a relatively narrow cross-section of systemic malignancies, suggesting a fundamental role for biological mechanisms shared across commonly brain metastatic tumor types. To identify and characterize such mechanisms, we performed genomic, transcriptional, and proteomic profiling using whole-exome sequencing, mRNA-seq, and reverse-phase protein array analysis in a cohort of the lung, breast, and renal cell carcinomas consisting of BM and patient-matched primary or extracranial metastatic tissues. While no specific genomic alterations were associated with BM, correlations with impaired cellular immunity, upregulated oxidative phosphorylation (OXPHOS), and canonical oncogenic signaling pathways including phosphoinositide 3-kinase (PI3K) signaling, were apparent across multiple tumor histologies. Multiplexed immunofluorescence analysis confirmed significant T cell depletion in BM, indicative of a fundamentally altered immune microenvironment. Moreover, functional studies using in vitro and in vivo modeling demonstrated heightened oxidative metabolism in BM along with sensitivity to OXPHOS inhibition in murine BM models and brain metastatic derivatives relative to isogenic parentals. These findings demonstrate that pathophysiological rewiring of oncogenic signaling, cellular metabolism, and immune microenvironment broadly characterizes BM. Further clarification of this biology will likely reveal promising targets for therapeutic development against BM arising from a broad variety of systemic cancers.
Subject(s)
Brain Neoplasms/pathology , Brain Neoplasms/secondary , DNA Fingerprinting/methods , Genomics/methods , Animals , Base Sequence , Brain Neoplasms/immunology , Cell Survival , Female , Fluorescent Antibody Technique , Gene Expression Regulation, Neoplastic , Humans , Mice , Mice, SCID , Neoplasm Transplantation , Protein Array Analysis , Proteomics , Superoxide Dismutase/metabolism , Survival Analysis , Exome SequencingABSTRACT
Ependymomas (EPN) show site specific genetic alterations and a recent DNA methylation profiling study identified nine molecular subgroups. C11orf95-RELA and YAP1 fusions characterise the RELA and YAP1 molecular subgroups, respectively, of supratentorial (ST)-EPNs. Current guidelines recommend molecular subgrouping over histological grade for accurate prognostication. Clinicopathological features of ST-EPNs in correlation with C11orf95-RELA and YAP1 fusions have been assessed in only few studies. We aimed to study these fusions in EPNs, and identify diagnostic and prognostic markers. qRT-PCR and Sanger Sequencing for the detection of C11orf95-RELA, YAP1-MAMLD1 and YAP1-FAM118B fusion transcripts, gene expression analysis for NFKB1, and immunohistochemistry for p53, MIB-1, nestin, VEGF, and L1CAM were performed. 88 EPNs (10-Grade I and 78-Grade II/III) from all sites were included. RELA fusions were unique to Grade II/III ST-EPNs, detected in 81.4% (22/27) and 18.5% (5/27) of pediatric and adult ST-EPNs respectively. ST-EPNs harbouring RELA fusions showed frequent grade III histology (81.5%), clear cell morphology (70.3%), upregulated NFKB1 expression, MIB-1 labelling indices (LI) ≥ 10% (77.8%), and immunopositivity for nestin (95.7%), VEGF (72%), L1CAM (79%), and p53 (64%). Presence of RELA fusions, L1CAM immunopositivity and MIB-1 LI ≥ 10% associated with poor outcome. L1CAM showed 81% concordance with RELA fusions. YAP1-MAMLD1 fusion was identified in a single RELA fusion negative adult anaplastic ST-EPN. RELA fusions are frequent in ST-EPNs and associate with poor outcome. L1CAM is a surrogate immunohistochemical marker. RELA fusion positive ST-EPNs strongly express nestin indicating increased stemness. Further evaluation of the interactions between NFKB and stem cell pathways is warranted.
Subject(s)
Ependymoma , NF-kappa B/metabolism , Nestin/metabolism , Neural Cell Adhesion Molecule L1/metabolism , Nuclear Proteins/metabolism , Supratentorial Neoplasms , Transcription Factor RelA/metabolism , Adolescent , Adult , Aged , Child , Child, Preschool , Ependymoma/metabolism , Ependymoma/pathology , Ependymoma/physiopathology , Female , Humans , Male , Middle Aged , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Progression-Free Survival , Retrospective Studies , Signal Transduction/physiology , Supratentorial Neoplasms/metabolism , Supratentorial Neoplasms/pathology , Supratentorial Neoplasms/physiopathology , Young AdultABSTRACT
Enhancer of zeste homolog-2(EZH2) is a key epigenetic regulator that functions as oncogene and also known for inducing altered trimethylation of histone at lysine-27 (H3K27me3) mark in various tumors. However, H3K27me3 targets and their precise relationship with gene expression are largely unknown in astrocytic tumors. In this study, we checked EZH2 messenger RNA and protein expression in 90 astrocytic tumors of different grades using quantitative PCR and immunohistochemistry, respectively. Further, genome-wide ChIP-seq analysis for H3K27me3 modification was also performed on 11 glioblastomas (GBMs) and 2 diffuse astrocytoma (DA) samples. Our results showed EZH2 to be highly overexpressed in astrocytic tumors with a significant positive correlation with grade. Interestingly, ChIP-seq mapping revealed distinct differences in genes and pathways targeted by these H3K27me3 modifications between GBM versus DA. Neuroactive ligand receptor pathway was found most enriched in GBM (P = 9.4 × 10-25), whereas DA were found to be enriched in metabolic pathways. Also, GBM showed a higher enrichment of H3K27me3 targets reported in embryonic stem cells and glioma stem cells as compared with DAs. Our results show majority of these H3K27me3 target genes were downregulated, not only due to H3K27me3 modification but also due to concomitant DNA methylation. Further, H3K27me3 modification-associated gene silencing was not restricted to promoter but also present in gene body and transcription start site regions. To the best of our knowledge, this is the first high-resolution genome-wide mapping of H3K27me3 modification in adult astrocytic primary tissue samples of human, highlighting the differences between grades. Interestingly, we identified SLC25A23 as important target of H3K27me3 modification, which was downregulated in GBM and its low expression was associated with poor prognosis in GBMs.
Subject(s)
Antiporters/genetics , Astrocytoma/genetics , DNA Methylation/genetics , Enhancer of Zeste Homolog 2 Protein/genetics , Glioblastoma/genetics , Histone-Lysine N-Methyltransferase/genetics , Mitochondrial Proteins/genetics , Astrocytoma/pathology , Cell Line, Tumor , Enhancer Elements, Genetic/genetics , Enhancer of Zeste Homolog 2 Protein/metabolism , Female , Gene Expression Regulation, Neoplastic , Genome, Human , Glioblastoma/pathology , Histone-Lysine N-Methyltransferase/metabolism , Humans , Male , Neoplasm Grading , Promoter Regions, GeneticABSTRACT
Elevated expression of enhancer of zeste homolog 2 (EZH2), a histone H3K27 methyltransferase, was observed in gliomas harboring telomerase reverse transcriptase (TERT) promoter mutations. Given the known involvement of TERT and EZH2 in glioma progression, the correlation between the two and subsequently its involvement in metabolic programming was investigated. Inhibition of human telomerase reverse transcriptase either pharmacologically or through genetic manipulation not only decreased EZH2 expression, but also (i) abrogated FASN levels, (ii) decreased de novo fatty acid accumulation, and (iii) increased ataxia-telangiectasia-mutated (ATM) phosphorylation levels. Conversely, diminished TERT and FASN levels upon siRNA-mediated EZH2 knockdown indicated a positive correlation between TERT and EZH2. Interestingly, ATM kinase inhibitor rescued TERT inhibition-mediated decrease in FASN and EZH2 levels. Importantly, TERT promoter mutant tumors exhibited greater microsatellite instability, heightened FASN levels and lipid accumulation. Coherent with in vitro findings, pharmacological inhibition of TERT by costunolide decreased lipid accumulation and elevated ATM expression in heterotypic xenograft glioma mouse model. By bringing TERT-EZH2 network at the forefront as driver of dysregulated metabolism, our findings highlight the non-canonical but distinct role of TERT in metabolic reprogramming and DNA damage responses in glioblastoma.
Subject(s)
DNA Damage/physiology , Enhancer of Zeste Homolog 2 Protein/metabolism , Glioblastoma/metabolism , Lipid Metabolism/physiology , Telomerase/metabolism , Animals , Cell Line, Tumor , Gene Expression Regulation, Neoplastic/physiology , Glioblastoma/pathology , Heterografts , Humans , Mice , Mice, NudeABSTRACT
Craniopharyngiomas (CP) are rare benign epithelial tumors, with two histological variants, namely the adamantinomatous variant (ACP) and the rarer papillary variant (PCP). They are locally infiltrative and surgically challenging tumors with severe long term morbidity. CTNNB1 mutations with ß-catenin immunopositivity and BRAFV600E mutations with anti-VE immunopositivity have been recently described in ACPs and PCPs respectively. We aimed to study BRAF and CTNNB1 gene mutations in CPs operated at our institute, and correlate it with clinicopathological parameters including histopathology and immunohistochemistry (IHC) for proteins VE-1 and ß-catenin. A total of 54 CPs diagnosed over 3-year duration were included. IHC for ß-catenin and VE-1 proteins, and Sanger sequencing for CTNNB1 (exon 3) and BRAF (exon 15) genes were performed. CTNNB1 mutations were identified in 63% (27/43) of ACPs while nuclear immunopositivity for ß-catenin was observed in 79% (34/43) of them. Seven ACPs showed ß-catenin immunopositivity in the absence of mutations. BRAFV600E (p.Val600Glu) mutations were observed in 57% of PCPs (4/7), while cytoplasmic immunopositivity for anti-VE1 antibody was observed only in 43% of PCPs (3/7), all of which also harboured BRAFV600E mutations. The mutations and IHC staining patterns of ACPs and PCPs were non-overlapping. Four cases with uncertain histological pattern could be subcategorised into specific variants only following mutation analysis/IHC. The identification of hallmark molecular signatures in the two CP variants holds promise for alternate improved treatment modalities, emphasizing the need for sub-categorization in routine histopathology reporting. IHC for ß-catenin and targeted sequencing for BRAFV600E serve as useful adjuncts.
Subject(s)
Craniopharyngioma/genetics , Craniopharyngioma/pathology , Pituitary Neoplasms/genetics , Pituitary Neoplasms/pathology , Proto-Oncogene Proteins B-raf/genetics , beta Catenin/genetics , Adolescent , Adult , Child , Craniopharyngioma/metabolism , Cytoplasm/metabolism , Cytoplasm/pathology , DNA Mutational Analysis , Diagnosis, Differential , Female , Follow-Up Studies , Humans , Immunohistochemistry , Male , Middle Aged , Mutation , Pituitary Neoplasms/metabolism , Retrospective Studies , beta Catenin/metabolismABSTRACT
Ependymomas are gliomas that recapitulate the ependymal cells microscopically and ultrastructurally. They commonly occur along the ventricular surfaces and central canal of the brain and spinal cord. Intracranial extra-axial ependymoma (IEAE) is a rare entity and is commonly misdiagnosed clinically and radiologically as a meningioma. The histogenesis of such IEAEs is obscure. A novel recurrent oncogenic fusion involving the C11orf95 and RELA genes was recently described in supratentorial ependymomas. A 9-year-old girl presented with a dural based parafalcine mass that, in addition to exhibiting classical immunohistochemical features of an ependymoma, also demonstrated C11orf95-RELA fusion, characteristic of supratentorial ependymomas. We suggest that IEAEs share their histogenesis with their intra-axial counterparts, arising either from dural extension of subcortical, subependymal rests or directly from ectopic dural rests.
Subject(s)
Brain Neoplasms/genetics , Brain Neoplasms/pathology , Ependymoma/genetics , Ependymoma/pathology , Proteins/genetics , Transcription Factor RelA/genetics , Brain/ultrastructure , Brain Neoplasms/diagnostic imaging , Brain Neoplasms/ultrastructure , Child , Ependymoma/diagnostic imaging , Female , Gene Fusion , HumansABSTRACT
BACKGROUND: Functional inactivation of ATRX characterizes large subgroups of malignant gliomas in adults and children. ATRX deficiency in glioma induces widespread chromatin remodeling, driving transcriptional shifts and oncogenic phenotypes. Effective strategies to therapeutically target these broad epigenomic sequelae remain undeveloped. METHODS: We utilized integrated multiomics and the Broad Institute Connectivity Map (CMAP) to identify drug candidates that could potentially revert ATRX-deficient transcriptional changes. We then employed disease-relevant experimental models to evaluate functional phenotypes, coupling these studies with epigenomic profiling to elucidate molecular mechanism(s). RESULTS: CMAP analysis and transcriptional/epigenomic profiling implicated the Class III HDAC Sirtuin2 (SIRT2) as a central mediator of ATRX-deficient cellular phenotypes and a driver of unfavorable prognosis in ATRX-deficient glioma. SIRT2 inhibitors reverted Atrx-deficient transcriptional signatures in murine neuroepithelial progenitor cells (mNPCs), impaired cell migration in Atrx/ATRX-deficient mNPCs and human glioma stem cells (GSCs), and increased expression of senescence markers in glioma models. Moreover, SIRT2 inhibition impaired growth and increased senescence in ATRX-deficient GSCs in vivo. These effects were accompanied by genome-wide shifts in enhancer-associated H3K27ac and H4K16ac marks, with the latter in particular demonstrating compelling transcriptional links to SIRT2-dependent phenotypic reversals. Motif analysis of these data identified the transcription factor KLF16 as a mediator of phenotype reversal in Atrx-deficient cells upon SIRT2 inhibition. CONCLUSIONS: Our findings indicate that SIRT2 inhibition selectively targets ATRX-deficient gliomas for senescence through global chromatin remodeling, while demonstrating more broadly a viable approach to combat complex epigenetic rewiring in cancer.
Subject(s)
Chromatin , Glioma , Adult , Child , Humans , Animals , Mice , Sirtuin 2/genetics , Sirtuin 2/metabolism , Glioma/pathology , X-linked Nuclear Protein/genetics , Kruppel-Like Transcription Factors/geneticsABSTRACT
Patients with cholangiocarcinoma have poor clinical outcomes due to late diagnoses, poor prognoses, and limited treatment strategies. To identify drug combinations for this disease, we have conducted a genome-wide CRISPR screen anchored on the bromodomain and extraterminal domain (BET) PROTAC degrader ARV825, from which we identified anticancer synergy when combined with genetic ablation of members of the mTOR pathway. This combination effect was validated using multiple pharmacological BET and mTOR inhibitors, accompanied by increased levels of apoptosis and cell cycle arrest. In a xenograft model, combined BET degradation and mTOR inhibition induced tumor regression. Mechanistically, the 2 inhibitor classes converged on H3K27ac-marked epigenetic suppression of the serine glycine one carbon (SGOC) metabolism pathway, including the key enzymes PHGDH and PSAT1. Knockdown of PSAT1 was sufficient to replicate synergy with single-agent inhibition of either BET or mTOR. Our results tie together epigenetic regulation, metabolism, and apoptosis induction as key therapeutic targets for further exploration in this underserved disease.
Subject(s)
Cholangiocarcinoma , MTOR Inhibitors , Humans , Epigenesis, Genetic , Clustered Regularly Interspaced Short Palindromic Repeats , Cell Line, Tumor , TOR Serine-Threonine Kinases , Cholangiocarcinoma/drug therapy , Cholangiocarcinoma/geneticsABSTRACT
Stimulating the innate immune system has been explored as a therapeutic option for the treatment of gliomas. Inactivating mutations in ATRX, defining molecular alterations in IDH-mutant astrocytomas, have been implicated in dysfunctional immune signaling. However, little is known about the interplay between ATRX loss and IDH mutation on innate immunity. To explore this, we generated ATRX-deficient glioma models in the presence and absence of the IDH1R132H mutation. ATRX-deficient glioma cells are sensitive to dsRNA-based innate immune agonism and exhibit impaired lethality and increased T-cell infiltration in vivo. However, the presence of IDH1R132H dampens baseline expression of key innate immune genes and cytokines in a manner restored by genetic and pharmacological IDH1R132H inhibition. IDH1R132H co-expression does not interfere with the ATRX deficiency-mediated sensitivity to dsRNA. Thus, ATRX loss primes cells for recognition of dsRNA, while IDH1R132H reversibly masks this priming. This work reveals innate immunity as a therapeutic vulnerability of astrocytomas.
Subject(s)
Astrocytoma , Brain Neoplasms , Glioma , Humans , Brain Neoplasms/genetics , Brain Neoplasms/metabolism , X-linked Nuclear Protein/genetics , Glioma/genetics , Glioma/metabolism , Astrocytoma/genetics , Mutation , Immunity, Innate/genetics , Isocitrate Dehydrogenase/genetics , Isocitrate Dehydrogenase/metabolismABSTRACT
Inactivating mutations in ATRX characterize large subgroups of malignant gliomas in adults and children. ATRX deficiency in glioma induces widespread chromatin remodeling, driving transcriptional shifts and oncogenic phenotypes. Effective strategies to therapeutically target these broad epigenomic sequelae remain undeveloped. We utilized integrated mulit-omics and the Broad Institute Connectivity Map (CMAP) to identify drug candidates that could potentially revert ATRX-deficient transcriptional changes. We then employed disease-relevant experimental models to evaluate functional phenotypes, coupling these studies with epigenomic profiling to elucidate molecular mechanim(s). CMAP analysis and transcriptional/epigenomic profiling implicated the Class III HDAC Sirtuin2 (Sirt2) as a central mediator of ATRX-deficient cellular phenotypes and a driver of unfavorable prognosis in ATRX-deficient glioma. Sirt2 inhibitors reverted Atrx-deficient transcriptional signatures in murine neuroprogenitor cells (mNPCs) and impaired cell migration in Atrx/ATRX-deficient mNPCs and human glioma stem cells (GSCs). While effects on cellular proliferation in these contexts were more modest, markers of senescence significantly increased, suggesting that Sirt2 inhibition promotes terminal differentiation in ATRX-deficient glioma. These phenotypic effects were accompanied by genome-wide shifts in enhancer-associated H3K27ac and H4K16ac marks, with the latter in particular demonstrating compelling transcriptional links to Sirt2-dependent phenotypic reversals. Motif analysis of these data identified the transcription factor KLF16 as a mediator of phenotype reversal in Atrx-deficient cells upon Sirt2 inhibition. Finally, Sirt2 inhibition impaired growth and increased senescence in ATRX-deficient GSCs in vivo . Our findings indicate that Sirt2 inhibition selectively targets ATRX-deficient gliomas through global chromatin remodeling, while demonstrating more broadly a viable approach to combat complex epigenetic rewiring in cancer. One Sentence Summary: Our study demonstrates that SIRT2 inhibition promotes senescence in ATRX-deficient glioma model systems through global epigenomic remodeling, impacting key downstream transcriptional profiles.
ABSTRACT
Stimulating the innate immune system has been explored as a therapeutic option for the treatment of gliomas. Inactivating mutations in ATRX , defining molecular alterations in IDH -mutant astrocytomas, have been implicated in dysfunctional immune signaling. However, little is known about the interplay between ATRX loss and IDH mutation on innate immunity. To explore this, we generated ATRX knockout glioma models in the presence and absence of the IDH1 R 132 H mutation. ATRX-deficient glioma cells were sensitive to dsRNA-based innate immune agonism and exhibited impaired lethality and increased T-cell infiltration in vivo . However, the presence of IDH1 R 132 H dampened baseline expression of key innate immune genes and cytokines in a manner restored by genetic and pharmacological IDH1 R132H inhibition. IDH1 R132H co-expression did not interfere with the ATRX KO-mediated sensitivity to dsRNA. Thus, ATRX loss primes cells for recognition of dsRNA, while IDH1 R132H reversibly masks this priming. This work reveals innate immunity as a therapeutic vulnerability of astrocytoma.
ABSTRACT
Long noncoding RNAs (lncRNA) have emerged as vital molecules governing epithelial-to-mesenchymal transition (EMT) in cancers. Translation regulatory RNA 1 (TRERNA1) is one such lncRNA known to enhance the transcriptional activity of the EMT-transcription factor, Snail. We have previously demonstrated differential upregulation of EMT-transcription factors and cadherin switching across various clinico-pathologic-molecular subclasses of ependymomas (EPN). With an aim to analyze the correlation between the expression of TRERNA1 in EPNs, we performed gene expression analysis for TRERNA1 on 75 Grade II/III EPNs and correlated with tumor site, C11orf95-RELA fusions, age, MIB-1 proliferative indices, and outcome wherever available. Upregulation of gene expression levels of TRERNA1 was seen in intracranial EPNs, with highest expression levels in pediatric posterior fossa EPNs. High TRERNA1 expression was found associated with higher proliferative indices (p = 0.034) and shorter progression free survival (p = 0.002). Our study, for the first time, demonstrates an association between TRERNA1 expressions and pediatric posterior fossa EPNs. Further in-vivo and in-vitro studies are required to confirm these findings and evaluate TRERNA1 as a novel biomarker and potential therapeutic target in childhood PF-EPNs.
Subject(s)
Biomarkers, Tumor/genetics , Ependymoma/genetics , Ependymoma/pathology , RNA, Long Noncoding/genetics , Adolescent , Adult , Child , Child, Preschool , Epithelial-Mesenchymal Transition/genetics , Female , Humans , Infant , Infratentorial Neoplasms/genetics , Infratentorial Neoplasms/pathology , Male , Middle Aged , Retrospective Studies , Spinal Cord Neoplasms/genetics , Spinal Cord Neoplasms/pathology , Supratentorial Neoplasms/genetics , Supratentorial Neoplasms/pathology , Young AdultABSTRACT
Immune check-point blockade (ICB) targeting programmed cell death ligand-1 (PD-L1)/programmed death-1 (PD-1) axis has created paradigm shift in cancer treatment. 'ST-RELA' and 'PF-A' molecular subgroups of ependymomas (EPN) show poor outcomes. We aimed to understand the potential candidature of EPNs for ICB. Supratentorial (ST) Grade II/III EPNs were classified into ST-RELA, ST-YAP, and ST-not otherwise specified (NOS), based on RELA/YAP1 fusion transcripts and/or L1CAM and p65 protein expression. Posterior fossa (PF) EPNs were classified into PF-A and PF-B based on H3K27me3 expression. Immunohistochemistry for PD-L1 and CD8 was performed. RelA protein enrichment at PDL1 promoter site was analysed by chromatin immunoprecipitation-qPCR (ChIP-qPCR). Eighty-three intracranial EPNs were studied. Median tumor infiltrating CD8 + cytotoxic T-lymphocyte (CTL) density was 6/mm2, and was higher in ST-EPNs (median 10/mm2) as compared to PF-EPNs (median 3/mm2). PD-L1 expression was noted in 17/83 (20%) EPNs, including 12/31 ST-RELA and rare ST-NOS (2/12), PF-A (2/25) and PF-B (1/13) EPNs. Twelve EPNs (14%) showed high CTL density and concurrent PD-L1 positivity, of which majority (10/12) were ST-RELA EPNs. Enrichment of RelA protein was seen at PDL1 promoter. Increased CTL densities and upregulation of PD-L1 in ST-RELA ependymomas suggests potential candidature for immunotherapy.
Subject(s)
B7-H1 Antigen/genetics , Ependymoma/immunology , Ependymoma/pathology , T-Lymphocytes, Cytotoxic/pathology , B7-H1 Antigen/metabolism , Ependymoma/genetics , Female , Humans , Immunohistochemistry , Immunotherapy/methods , Male , Neural Cell Adhesion Molecule L1/metabolism , Supratentorial Neoplasms/pathology , Transcription Factor RelA/metabolismABSTRACT
Epithelial-to-mesenchymal transition (EMT) plays an important role in invasion and metastasis of various cancers including gliomas. EMT has also been linked to cancer stem cells and resistance to chemotherapy. An initial in-silico data mining in a published ependymoma (EPN) patient series (GSE21687) revealed up-regulation of EMT transcription factors in tumor samples. Furthermore, quantitative real-time polymerase chain reaction-based gene expression analysis of EMT transcription factors in 96 EPNs showed significant up-regulation of SNAI1, SNAI2, ZEB1, and TWIST1 as compared with normal brain, associated with up-regulation of CDH2/N-cadherin and down-regulation of CDH1/E-cadherin. Although this was observed in varying degrees in all clinicopathological-molecular subgroups of EPNs, it was most evident in supratentorial EPNs harboring fusions of RELA (v-rel avian reticuloendotheliosis viral oncogene homolog A) gene and in posterior fossa EPNs. Immunohistochemistry performed in 60 of the above cases corroborated with gene expression patterns, and immunopositivity for Snail, Slug, Zeb1, and Twist1 was observed in 80%, 80%, 81%, and 63% of all EPNs. Immunopositivity for N-cadherin and E-cadherin was observed in 76.6% and 2% of the cases, respectively. Univariate Cox regression analysis showed that low expression of CDH1/E-cadherin (Pâ¯=â¯.002) and high expression levels of CDH2/N-cadherin (Pâ¯<â¯.001), SNAI1/Snail (Pâ¯=â¯.023), SNAI2/Slug (Pâ¯<â¯.001), and ZEB1 (Pâ¯<â¯.001) were associated with shorter progression-free survival. Here, we report for the first time the existence of EMT-like phenotype in EPNs. These factors could represent new prognostic and therapeutic targets in EPN.
Subject(s)
Biomarkers, Tumor/genetics , Brain Neoplasms/genetics , Ependymoma/genetics , Epithelial-Mesenchymal Transition/genetics , Transcription Factors/genetics , Adolescent , Adult , Aged , Biomarkers, Tumor/analysis , Brain Neoplasms/chemistry , Brain Neoplasms/pathology , Brain Neoplasms/therapy , Case-Control Studies , Child , Child, Preschool , Data Mining , Databases, Genetic , Ependymoma/chemistry , Ependymoma/pathology , Ependymoma/therapy , Female , Gene Expression Profiling/methods , Gene Expression Regulation, Neoplastic , Humans , Immunohistochemistry , In Situ Hybridization, Fluorescence , Infant , Male , Middle Aged , Progression-Free Survival , Real-Time Polymerase Chain Reaction , Risk Factors , Time Factors , Transcription Factors/analysis , Up-Regulation , Young AdultABSTRACT
Recent molecular subgrouping of ependymomas (EPN) by DNA methylation profiling has identified ST-EPN-RELA and PF-EPN-A subgroups to be associated with poor outcome. Snail/Slug are cardinal epithelial-to-mesenchymal transcription factors (EMT-TFs) and are overexpressed in several CNS tumors, including EPNs. A systematic analysis of gene-sets/modules co-expressed with Snail and Slug genes using published expression microarray dataset (GSE27279)identified 634 genes for Snail with enriched TGF-ß, PPAR and PI3K signaling pathways, and 757 genes for Slug with enriched focal adhesion, ECM-receptor interaction and regulation of actin cytoskeleton related pathways. Of 37 genes commonly expressed with both Snail and Slug, IL1R1, a cytokine receptor of interleukin-1 receptor family, was positively correlated with Snail (r=0.43) and Slug (r=0.51), preferentially expressed in ST-EPN-RELA and PF-EPN-A molecular groups, and enriched for pathways related to inflammation, angiogenesis and glycolysis. IL1R1 expression was fairly specific to EPNs among various CNS tumors analyzed. It also showed significant positive correlation with EMT, stemness and MDSC (myeloid derived suppressor cell) markers. Our study reports IL1R1 as a poor prognostic marker associated with EMT-like phenotype and stemness in EPNs. Our findings emphasize the need to further examine and validate IL1R1 as a novel therapeutic target in aggressive subsets of intracranial EPNs.
ABSTRACT
Diffuse leptomeningeal glioneuronal tumor (DL-GNT) is a newly introduced tumor entity of uncertain prognosis characterised by a primary diffuse leptomeningeal growth pattern, oligodendroglial-like morphology and dual glial/neuronal differentiation. Predominantly occurring in children, these tumors present as chronic meningitis and mimic infectious/inflammatory diseases. They are surgically challenging tumors with a high incidence of delayed morbidity and mortality despite low-grade histology. Their molecular genetic profile is not fully elucidated and few reports have identified chromosome 1p and 19q deletions, and BRAF alterations. We present a rare instance of a DL-GNT in a 13-year-old female who presented with slowly progressive and sequential neurological deficits over a 12-month duration. Imaging showed leptomeningeal thickening and spinal lesions. Biopsy from the spinal mass showed histomorphological features characteristic of DL-GNT. Further molecular analysis revealed 1p and 19q co-deletion and H3K27M mutation, while no mutation were identified in IDH, TERT, or BRAF genes. Patient died 4 months after diagnosis. Only one previous case of DL-GNT has been reported to harbour H3K27M mutation. Although H3K27M mutations have been described in rare examples of low-grade glial and glioneuronal tumors, whether DL-GNTs with H3K27M represent a rare growth pattern of the aggressive H3K27M-mutant diffuse midline gliomas needs further clarification.
Subject(s)
Cell Transformation, Neoplastic/genetics , Chromosome Deletion , Chromosomes, Human, Pair 19/genetics , Chromosomes, Human, Pair 1/genetics , Meningeal Neoplasms/genetics , Meningeal Neoplasms/therapy , Mutation , Neuroglia/pathology , Neurons/pathology , Oligodendroglioma/genetics , Oligodendroglioma/therapy , Adolescent , Female , Humans , Meningeal Neoplasms/pathology , Neoplasm Grading , Oligodendroglioma/pathology , Ovarian CystsABSTRACT
Loss of SMARCB1/INI1 expression is considered to be a hallmark for childhood chordomas (CCs). Although mutation/loss of 22q has strongly established the loss of SMARCB1/INI1 in cancers, the cause in CCs remains elusive. Recent studies suggest role of miRNAs in regulation of SMARCB1/INI1 expressions. We examined 5 reported/target predicted miRNAs to SMARCB1/INI1 in SMARCB1/INI1 immunonegative and immunopositive cases, and found upregulation of miR-671-5p and miR-193a-5p in SMARCB1/INI1-immunonegative cases. Notably, these two miRNAs were significantly predicted to target TGF-ß signaling, suggestive of dysregulation of developmental and osteoblast regulation pathway in CCs. Overall, we suggest miR-671-5p- and miR-193a-5p-mediated epigenetic mode of SMARCB1/INI1 loss and downregulated TGF-ß pathway in CCs.
Subject(s)
Chordoma/genetics , Down-Regulation/genetics , Gene Expression Regulation, Neoplastic/genetics , Gene Expression/genetics , Genetic Association Studies , MicroRNAs/genetics , SMARCB1 Protein/genetics , Up-Regulation/genetics , Adult , Child , Epigenesis, Genetic/genetics , Female , Humans , Male , Mutation , Signal Transduction/genetics , Transforming Growth Factor beta/genetics , Transforming Growth Factor beta/metabolismABSTRACT
Enhancer of Zeste homologue2 (EZH2) is an epigenetic regulator that functions as oncogene in astrocytic tumors, however, EZH2 regulation remains little studied. In this study, we measured EZH2 levels in low (Gr-II,DA) and high grade (Gr-IV,GBM) astrocytic tumors and found significant increased EZH2 transcript level with grade(median DA-8.5, GBM-28.9).However, a different trend was reflected in protein levels, with GBMs showing high EZH2 LI(median-26.5) compared to DA (median 0.3). This difference in correlation of EZH2 protein and RNA levels suggested post-transcriptional regulation of EZH2, likely mediated by miRNAs. We selected eleven miRNAs that strongly predicted to target EZH2 and measured their expression. Three miRNAs (miR-26a-5p,miR27a-3p and miR-498) showed significant correlation with EZH2 protein, suggesting them as regulators of EZH2, however miR-26a-5p levels decreased with grade. ChIP analyses revealed H3K27me3 modifications in miR-26a promoter suggesting feedback loop between EZH2 and miR26a. We further measured six downstream miRNA targets of EZH2 and found significant downregulation of four (miR-181a/b and 200b/c) in GBM. Interestingly, EZH2 associated miRNAs were predicted to target 25 genes in glioma-pathway, suggesting their role in tumor formation or progression. Collectively, our work suggests EZH2 and its miRNA interactors may serve as promising biomarkers for progression of astrocytic tumors and may offer novel therapeutic strategies.
Subject(s)
Astrocytoma/genetics , Brain Neoplasms/genetics , Enhancer of Zeste Homolog 2 Protein/genetics , Glioblastoma/genetics , MicroRNAs/genetics , MicroRNAs/physiology , Adult , Aged , Astrocytoma/pathology , Astrocytoma/therapy , Brain Neoplasms/pathology , Brain Neoplasms/therapy , Disease Progression , Enhancer of Zeste Homolog 2 Protein/metabolism , Epistasis, Genetic , Female , Gene Expression/genetics , Glioblastoma/pathology , Glioblastoma/therapy , Humans , Male , Middle Aged , Molecular Targeted Therapy , Neoplasm Staging , Young AdultABSTRACT
BACKGROUND: Chordomas are slow-growing tumors and most commonly involve the sacrum and clivus. Multiple recurrences are frequent. Childhood chordomas are rare and often show exceptionally aggressive behavior, resulting in short survival and a high incidence of metastatic spread. OBJECTIVE: This study examined the histologic features and immunohistochemical profile of pediatric chordomas and compared them with their adult counterparts. METHODS: Nine pediatric and 13 adult cases were included in the study. Childhood chordomas were classified into conventional, atypical, and poorly differentiated types. Immunohistochemistry was performed for cytokeratin, epithelial membrane antigen, vimentin, S100, brachyury, p53, INI1, epidermal growth factor receptor (EGFR), and CD117. Cytogenetic analyses were performed in a subset of tumors for SMARCB1/INI1 locus on 22q chromosome by fluorescent in situ hybridization (FISH) and analysis of the SMARCB1/INI1 gene sequence. RESULTS: All tumors showed expression of cytokeratin, epithelial membrane antigen, S100, vimentin, brachyury, and EGFR. Atypical morphology, p53 expression, higher MIB-1 labelling index (LI), and INI1 loss were more frequently seen in pediatric chordomas as compared with adults. None of the tumors showed CD117 expression. No point mutation in the SMARCB1/INI1 gene was noted in the tumors examined; however, 4 pediatric and 1 adult chordoma showed loss of this locus on FISH analysis. CONCLUSIONS: A subset of pediatric chordomas with atypical histomorphologic features needs to be identified, as they behave in an aggressive manner and require adjuvant therapy. Pediatric chordomas more frequently show p53 expression, INI1 loss, and higher MIB-1 LI as compared with adults, whereas EGFR expression is common to both.