cyclin-d1 and Ependymoma

Nuclear factor-kappa B (NF-κB) signaling is an intracellular signaling pathway involved in inflammatory responses and the pathogenesis of various cancers, including ependymoma, which is a rare and chemotherapy-resistant glioma. Several isoforms of fusion proteins that consist of a nuclear protein, zinc finger translocation associated (ZFTA), and RELA (ZFTA-RELA), an NF-κB-signaling effector transcription factor, cause excessive activation of the NF-κB signaling pathway and result in supratentorial ependymomas (ST-EPN-RELA). As inhibitors of NF-κB activity induced by ZFTA-RELA are expected to be therapeutic agents for ST-EPN-RELA, we established an NF-κB responsive luciferase reporter cell line that expresses the most common isoform of ZFTA-RELA in a doxycycline-dependent manner. Using this reporter cell line, we screened fungus extracts for compounds that inhibit the NF-κB activity induced by ZFTA-RELA expression and identified aszonalenin, an alkaloid from Aspergillus novofumigatus. We also purified analogs of aszonalenin, namely acetylaszonalenin and epi-aszonalenin B and C. In a luciferase assay using cells constitutively expressing luciferase (counter assay), acetylaszonalenin and epi-aszonalenin C showed non-specific inhibition of the luciferase activity. Aszonalenin and epi-aszonalenin B inhibited the NF-κB responsive luciferase activity by expressing ZFTA-RELA more strongly than the luciferase activity in the counter assay. The upregulation of endogenous NF-κB responsive genes, such as CCND1, ICAM1, and L1CAM, by ZFTA-RELA expression was inhibited by epi-aszonalenin B, but not by aszonalenin. This study suggests that epi-aszonalenin B may be a lead compound for the therapeutic development of ST-EPN-RELA.

Topics: Aspergillus; Blotting, Western; Cyclin D1; Doxycycline; Ependymoma; Gene Expression Regulation, Neoplastic; HEK293 Cells; HeLa Cells; Humans; Indole Alkaloids; Intercellular Adhesion Molecule-1; Molecular Structure; Neural Cell Adhesion Molecule L1; NF-kappa B; Nuclear Proteins; Oncogene Proteins, Fusion; Reverse Transcriptase Polymerase Chain Reaction; Transcription Factor RelA

Supratentorial extraventricular ependymomas (STEEs) are relatively rare ependymomas, and their pathologic and genetic characteristics are still poorly understood. The aim of this study was to determine the histologic, immunohistochemical, and RELA fusion features, as well as to clarify in more detail the clinical courses of STEEs. Data from a total of 43 patients with STEEs was analyzed retrospectively. The status of RELA fusion was evaluated using fluorescence in situ hybridization. The expression levels of L1CAM, p65, cyclin D1, and p53 were assessed using immunohistochemistry. Progression-free survival and overall survival were calculated via Kaplan-Meier estimation using the log-rank test. Among all 43 STEEs, 65.1% (28/43) are positive for RELA fusion. Interestingly, almost half of the patients with RELA fusion-positive ependymomas are adults (13/28), and 89.3% (25/28) cases are anaplastic ependymomas, which suggests that RELA fusion testing is necessary in adults with STEEs. We investigated the immunohistochemical status of p65, L1CAM and CCND1 protein expression for their ability to predict RELA fusion status. RELA fusion-positive STEEs are frequently associated with expression of p65 (85.2%), L1CAM (85.2%), and CCND1 (81.5%). The accuracy of predicting RELA fusion status was much higher when the expression of p65 and L1CAM was combined, that is, when both were immunopositive. The status of RELA fusion, p53 overexpression, and extent of tumor resection are significantly associated with prognosis.

Topics: Adolescent; Adult; Biomarkers, Tumor; Biopsy; Child; Child, Preschool; Cyclin D1; Disease Progression; Ependymoma; Female; Gene Fusion; Genetic Predisposition to Disease; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Infant; Male; Middle Aged; Neural Cell Adhesion Molecule L1; Predictive Value of Tests; Progression-Free Survival; Retrospective Studies; Supratentorial Neoplasms; Time Factors; Transcription Factor RelA; Tumor Suppressor Protein p53; Young Adult

Ependymomas (EPNs) are the third most common brain tumor in children. These tumors are resistant to available chemotherapeutic treatments, therefore new effective targeted therapeutics must be identified. Increasing evidence shows epigenetic alterations including histone posttranslational modifications (PTMs), are associated with malignancy, chemotherapeutic resistance and prognosis for pediatric EPNs. In this study we examined histone PTMs in EPNs and identified potential targets to improve chemotherapeutic efficacy.. Global histone H3 lysine 4 trimethylation (H3K4me3) levels were detected in pediatric EPN tumor samples with immunohistochemistry and immunoblots. Candidate genes conferring therapeutic resistance were profiled in pediatric EPN tumor samples with micro-array. Promoter H3K4me3 was examined for two candidate genes, CCND1 and ERBB2, with chromatin-immunoprecipitation coupled with real-time PCR (ChIP-PCR). These methods and MTS assay were used to verify a relationship between H3K4me3 levels and CCND1 and ERBB2, and to investigate cell viability in response to chemotherapeutic drugs in primary cultured pediatric EPN cells.. H3K4me3 levels positively correlate with WHO grade malignancy in pediatric EPNs and are associated with progression free survival in patients with posterior fossa group A EPNs (PF-EPN-A). Reduction of H3K4me3 by silencing its methyltransferase SETD1A, in primary cultured EPN cells increased cell response to chemotherapy.. Our results support the development of a novel treatment that targets H3K4me3 to increase chemotherapeutic efficacy in pediatric PF-EPN-A tumors.

Topics: Carboplatin; Cell Survival; Child, Preschool; Cyclin D1; Drug Resistance, Neoplasm; Ependymoma; Female; Gene Expression Regulation, Neoplastic; Histone-Lysine N-Methyltransferase; Histones; Humans; Male; Pediatrics; Primary Cell Culture; Promoter Regions, Genetic; Receptor, ErbB-2; Vincristine

Ependymal tumors are pathologically defined intrinsic neoplasms originating in the intracranial compartments or the spinal cord that affect both children and adults. The recently integrated classification of ependymomas based on both histological and molecular characteristics is capable of subgrouping patients with various prognoses. However, the application of histological and molecular markers in Chinese patients with ependymomas has rarely been reported. We aimed to demonstrate the significance of histological characteristics, the v-relavian reticuloendotheliosis viral oncogene homolog A (RELA) fusions and other molecular features in ependymal tumors.. We reviewed the histological characteristics of ependymal tumors using conventional pathological slides and investigate the RELA fusions and Cylclin D1 (CCND1) amplification by Fluorescence in situ hybridization (FISH) and trimethylation of histone 3 lysine 27 (H3K27me3) expression by immunohistochemistry (IHC) methods. SPSS software was used to analyze the data.. We demonstrated that hypercellularity, atypia, microvascular proliferation, necrosis, mitosis, and an elevated Ki-67 index, were tightly associated with an advanced tumor grade. Tumor location, necrosis, mitosis and the Ki-67 index were related to the survival of the ependymomas, but Ki67 was the only independent prognostic factor. Additionally, RELA fusions, mostly presented in pediatric grade III intracranial ependymomas, indicated decreased survival times of patients, and closely related to the patients' age, tumor grade, cellularity, cellular atypia, necrosis and Ki67 index in the intracranial ependymal tumors, whereas reduction of H3K27me3 predicted the worse prognosis in ependymal tumors.. Histological and molecular features facilitate tumor grading and prognostic predictions for ependymal tumors in Chinese patients.

Topics: Adolescent; Adult; Biomarkers, Tumor; Brain Neoplasms; Child; China; Cyclin D1; Ependymoma; Female; Follow-Up Studies; Histones; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Kaplan-Meier Estimate; Ki-67 Antigen; Male; Necrosis; Neoplasm Grading; Prognosis; Spinal Cord Neoplasms; Transcription Factor RelA; Young Adult

Ependymomas are tumors of the CNS. Although cyclin D1 overexpression has been related to several cancers, its prognostic value in ependymomas has not yet been fully established. We evaluated cyclin D1 expression by an immunohistochemistry analysis of 149 samples of ependymomas, including some relapses, corresponding to 121 patients. Eighty-one patients were adults, 60 were intracranial cases and 92 tumors were grade II. Gross total resection (GTR) was achieved in 62% of cases, and relapse was confirmed in 41.4% of cases. Cyclin D1 protein expression was analyzed by immunohistochemistry and scored with a labeling index (LI) calculated as the percentage of positively stained cells by intensity. We also analyzed expression of CCND1 and NOTCH1 in 33 samples of ependymoma by quantitative real-time PCR. A correlation between cyclin D1 LI score and anaplastic cases (P < 0.001), supratentorial location (P < 0.001) and age (P = 0.001) were observed. A stratified analysis demonstrated that cyclin D1 protein expression was strong in tumors with a supratentorial location, independent of the histological grade or age. Relapse was more frequent in cases with a higher cyclin D1 LI score (P = 0.046), and correlation with progression-free survival was observed in cases with GTR (P = 0.002). Only spinal canal tumor location and GTR were suggestive markers of PFS in multivarite analyses. Higher expression levels were observed in anaplastic cases for CCND1 (P = 0.002), in supratentorial cases for CCND1 (P = 0.008) and NOTCH1 (P = 0.011). There were correlations between the cyclin D1 mRNA and protein expression levels (P < 0.0001) and between CCND1 and NOTCH1 expression levels (P = 0.003). Higher cyclin D1 LI was predominant in supratentorial location and predict relapse in GTR cases. Cyclin D1 could be used as an immunohistochemical marker to guide follow-up and treatment in these cases.

Topics: Adolescent; Adult; Biomarkers, Tumor; Cyclin D1; Disease-Free Survival; Ependymoma; Female; Humans; Immunohistochemistry; Male; Neoplasm Recurrence, Local; Receptor, Notch1; Supratentorial Neoplasms; Young Adult

Currently, there are few effective adjuvant therapies for pediatric ependymoma outside confocal radiation, and prognosis remains poor. The phosphoinositide 3-kinase (PI3K) pathway is one of the most commonly activated pathways in cancer. PI3Ks transduce signals from growth factors and cytokines, resulting in the phosphorylation and activation of AKT, which in turn induces changes in cell growth, proliferation, and apoptosis.. PI3K pathway status was analyzed in ependymoma using gene expression data and immunohistochemical analysis of phosphorylated AKT (P-AKT). The effect of the PI3K pathway on cell proliferation was investigated by immunohistochemical analysis of cyclin D1 and Ki67, plus in vitro functional analysis. To identify a potential mechanism of PI3K pathway activation, PTEN protein expression and the mutation status of PI3K catalytic subunit α-isoform gene (PIK3CA) was investigated.. Genes in the pathway displayed significantly higher expression in supratentorial than in posterior fossa and spinal ependymomas. P-AKT protein expression, indicating pathway activation, was seen in 72% of tumors (n = 169) and P-AKT expression was found to be an independent marker of a poorer progression-free survival. A significant association between PI3K pathway activation and cell proliferation was identified, suggesting that pathway activation was influencing this process. PTEN protein loss was not associated with P-AKT staining and no mutations were identified in PIK3CA.. Our results suggest that the PI3K pathway could act as a biomarker, not only identifying patients with a worse prognosis but also those that could be treated with therapies targeted against the pathway, a strategy potentially effective in a high percentage of ependymoma patients.

Topics: Biomarkers, Tumor; Brain Neoplasms; Cell Line, Tumor; Cell Proliferation; Child; Child, Preschool; Class I Phosphatidylinositol 3-Kinases; Cyclin D1; Disease-Free Survival; Ependymoma; Female; Humans; Kaplan-Meier Estimate; Male; Molecular Targeted Therapy; Phosphatidylinositol 3-Kinases; Prognosis; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Signal Transduction; Transcriptome

The molecular genetic events involved in the development of ependymal neoplasms are not well understood. This study retrospectively examines 41 ependymomas; in all tumors the cyclin D1 labeling index (LI) (percent of positive immunostaining tumor cells) was correlated with MIB-1 LI and outcome. The study included 41 patients (25 males and 16 females) ranging in age from 1.5 to 70 years (mean, 30.7 years). Twenty-five patients underwent a subtotal resection or biopsy, and 16 underwent a gross total resection. Thirty-two patents had an ordinary or tanycytic ependymoma, and 9 had anaplastic/malignant tumors. The cyclin D1 LI ranged from 0 to 8.2 (mean, 0.7); 21 tumors had an LI of 0, and 8 had an LI of >1.0. The MIB-1 LI ranged from 0.1 to 34 (mean, 4.6); 16 tumors had an LI >2.0. All 8 patients with a cyclin D1 LI of >1.0 had an MIB-1 LI of >2.0. The tumors in the 6 of 8 patients with a cyclin D1 LI of >1.0 were classified as anaplastic/malignant tumors. The findings at the most recent follow-up were as follows: alive with no evidence of tumor (n = 11; mean, 64.1 months); died with evidence of tumor (n = 11; mean, 45.9 months); alive with tumor (n = 10; mean, 39.0 months). Two patients were alive with evidence of residual disease at follow-up intervals of 7 and 16 months but were lost to further follow-up. The remaining 6 patients either were lost to follow-up or else died in the immediate postoperative period. Cyclin D1 and MIB-1 LI did not reliably correlate with clinical outcome or recurrence. All tumors with an elevated cyclin D1 LI also had an elevated MIB-1 LI; however, the converse was not true. An elevated cyclin D1 LI (>1.0 in this study) appeared to be associated with anaplastic/malignant histology; however, cyclin D1 LI along with MIB-1 LI and histology did not always reliably correlate with clinical outcome.

Topics: Adolescent; Adult; Aged; Antigens, Nuclear; Biomarkers, Tumor; Brain Neoplasms; Child; Child, Preschool; Cyclin D1; Ependymoma; Female; Humans; Immunohistochemistry; Infant; Ki-67 Antigen; Male; Middle Aged; Nuclear Proteins; Spinal Cord Neoplasms

While several genetic alterations associated with the evolution of the astrocytomas have been identified, the molecular basis of non-astrocytic brain tumors has remained largely unknown. In this study, p15, p16, CDK4 and cyclin D1 genes were analyzed in 69 nonastrocytic human brain tumors, including 17 oligodendrogliomas, 16 medulloblastomas/primitive neuroectodermal tumors (PNETs), 14 ependymomas and 22 meningiomas. Southern blot analysis of DNA from frozen samples showed no homozygous deletions in p15 or p16 genes in any of these tumors. No mobility shift was found by PCR-single-strand conformation polymorphism (PCR-SSCP) analysis in exons 1 and 2 of the p15 gene and exons 1 and 2 of the p16 genes, except for one oligodendroglioma. Direct sequencing of DNA from this tumor showed a G --> A transition at nucleotide 436 (codon 140) in exon 2 of the p16 gene, which is a common polymorphism. Southern blot analyses revealed no amplification of CDK4 and cyclin D1 genes in any of the neoplasms analyzed. In contrast to astrocytic brain tumors, which show frequent loss of the p16 gene and amplification of the CDK4 gene, alteration of these genes appears to be rare in other neoplasms of the human nervous system.

Topics: Astrocytoma; Base Sequence; Blotting, Southern; Brain Neoplasms; Cerebellar Neoplasms; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinases; Cyclins; DNA Primers; Ependymoma; Humans; Medulloblastoma; Meningeal Neoplasms; Meningioma; Molecular Sequence Data; Neuroectodermal Tumors; Oligodendroglioma; Oligonucleotide Probes; Oncogene Proteins; Point Mutation; Polymerase Chain Reaction; Polymorphism, Single-Stranded Conformational; Proto-Oncogene Proteins; Reference Values