ABSTRACT: Both genetic and environmental factors are thought to be causal in gliomagenesis. Several genes have been implicated in glioma development, but the putative role of a major immunity-related gene complex member, immunoglobulin heavy chain ? (IGHG) has not been evaluated. Prior observations that IGHG-encoded ? marker (GM) allotypes exhibit differential sensitivity to an immunoevasion strategy of cytomegalovirus, a pathogen implicated as a promoter of gliomagenesis, has lead us to hypothesize that these determinants are risk factors for glioma. To test this hypothesis, we genotyped the IGHG locus comprising the GM alleles, specifically GM alleles 3 and 17, of 120 glioma patients and 133 controls via TaqMan® genotyping assay. To assess the associations between GM genotypes and the risk of glioma, we applied an unconditional multivariate logistic regression analysis adjusted for potential confounding variables. In comparison to subjects who were homozygous for the GM 17 allele, the GM 3 homozygotes were over twice as likely, and the GM 3/17 heterozygotes were over three times as likely, to develop glioma. Similar results were achieved when analyzed by combining the data corresponding to alleles GM 3 and GM 3/17 in a dominant model. The GM 3/17 genotype and the combination of GM 3 and GM 3/17 were found to be further associated with over 3 times increased risk for high-grade astrocytoma (grades III-IV). Allele frequency analyses also showed an increased risk for gliomas and high-grade astrocytoma in association with GM 3. Our findings support the premise that the GM 3 allele may present risk for the development of glioma, possibly by modulating immunity to cytomegalovirus.
Project description:Background:The genomic characterization of sporadically arising gliomas has delineated molecularly and clinically distinct subclasses of disease. However, less is known about the molecular nature of gliomas that are familial in origin. We performed molecular subtyping of 163 tumor specimens from individuals with a family history of glioma and integrated germline and somatic genomic data to characterize the pathogenesis of 20 tumors in additional detail. Methods:Immunohistochemical analyses were performed on formalin-fixed, paraffin-embedded tumor sections to determine molecular subtypes of glioma. For 20 cases, tumor DNA was exome sequenced on an Illumina HiSeq 2000 platform and copy number profiling was performed on the Illumina HumanOmniExpress BeadChip. Genotypes at glioma risk polymorphisms were determined from germline DNA profiled on the Illumina Infinium OncoArray and deleterious germline mutations were identified from germline sequencing data. Results:All 3 molecular subtypes of sporadic glioma were represented in the overall case series, including molecular glioblastoma (n = 102), oligodendroglioma (n = 21), and astrocytoma (n = 20). Detailed profiling of 20 of these cases showed characteristic subtype-specific alterations at frequencies comparable to sporadic glioma cases. All 20 cases had alterations in genes regulating telomere length. Frequencies of common glioma risk alleles were similar to those among sporadic cases, and correlations between risk alleles and same-gene somatic mutations were not observed. Conclusions:This study illustrates that the molecular characteristics of familial tumors profiled largely recapitulate what is known about sporadic glioma and that both germline and somatic molecular features target common core pathways involved in gliomagenesis. Key Points:1. Familial and sporadic gliomas display highly comparable molecular landscapes. 2. Germline and somatic molecular events target common core pathways involved in gliomagenesis. 3. Carriage of germline glioma risk variants is not associated with somatic events in the same gene.
Project description:Immunoglobulin G (IgG) proteins are known for the huge diversity of the variable domains of their heavy and light chains, aimed at protecting each individual against foreign antigens. The IgG also harbor specific polymorphism concentrated in the CH2 and CH3-CHS constant regions located on the Fc fragment of their heavy chains. But this individual particularity relies only on a few amino acids among which some could make accurate sequence determination a challenge for mass spectrometry-based techniques.The purpose of the study was to bring a molecular validation of proteomic results by the sequencing of encoding DNA fragments. It was performed using ten individual samples (DNA and sera) selected on the basis of their Gm (gamma marker) allotype polymorphism in order to cover the main immunoglobulin heavy gamma (IGHG) gene diversity. Gm allotypes, reflecting part of this diversity, were determined by a serological method. On its side, the IGH locus comprises four functional IGHG genes totalizing 34 alleles and encoding the four IgG subclasses. The genomic study focused on the nucleotide polymorphism of the CH2 and CH3-CHS exons and of the intron. Despite strong sequence identity, four pairs of specific gene amplification primers could be designed. Additional primers were identified to perform the subsequent sequencing. The nucleotide sequences obtained were first assigned to a specific IGHG gene, and then IGHG alleles were deduced using a home-made decision tree reading of the nucleotide sequences. IGHG amino acid (AA) alleles were determined by mass spectrometry. Identical results were found at 95% between alleles identified by proteomics and those deduced from genomics. These results validate the proteomic approach which could be used for diagnostic purposes, namely for a mother-and-child differential IGHG detection in a context of suspicion of congenital infection.
Project description:BACKGROUND:Gliomas are the most common aggressive brain tumors and have many complex pathological types. Previous reports have discovered that genetic mutations are associated with the risk of glioma. However, it is unclear whether uniform genetic mutations exist difference between glioma and its two pathological types in the Han Chinese population. MATERIALS AND METHODS:We evaluated 20 SNPs of 703 glioma cases (338 astrocytoma cases, 122 glioblastoma cases) and 635 controls in a Han Chinese population using ?(2) test and genetic model analysis. RESULTS:In three case-control studies, we found rs9288516 in XRCC5 gene showed a decreased risk of glioma (OR, 0.85; 95% CI, 0.73-0.99; P = 0.042) and glioblastoma (OR, 0.70; 95% CI, 0.52-0.92; P = 0.001) in the allele model. We identified rs414805 in RPA3 gene showed an increased risk of glioblastoma in allele model (OR, 1.38; 95% CI, 1.00-1.89; P = 0.047) and dominant model (OR, 1.57; 95% CI, 1.05-2.35; P = 0.027), analysis respectively. Meanwhile, rs2297440 in RTEL1 gene showed an increased risk of glioma (OR, 1.30; 95% CI, 1.10-1.54; P = 0.002) and astrocytoma (OR, 1.26; 95% CI, 1.02-1.54; P = 0.029) in the allele model. In addition, we also observed a haplotype of "GCT" in the RTEL1 gene with an increased risk of astrocytoma (P = 0.005). CONCLUSIONS:Polymorphisms in the XRCC5, RPA3 and RTEL1 genes, combinating with previous reaserches, are associated with glioma developing. However, those genes mutations may play different roles in the glioma, astrocytoma and glioblastoma, respectively.
Project description:Glioma, the most common central nervous system cancer in adults, has poor prognosis. Here we identify a new SNP associated with glioma risk, rs1920116 (near TERC), that reached genome-wide significance (Pcombined = 8.3 × 10(-9)) in a meta-analysis of genome-wide association studies (GWAS) of high-grade glioma and replication data (1,644 cases and 7,736 controls). This region has previously been associated with mean leukocyte telomere length (LTL). We therefore examined the relationship between LTL and both this new risk locus and other previously established risk loci for glioma using data from a recent GWAS of LTL (n = 37,684 individuals). Alleles associated with glioma risk near TERC and TERT were strongly associated with longer LTL (P = 5.5 × 10(-20) and 4.4 × 10(-19), respectively). In contrast, risk-associated alleles near RTEL1 were inconsistently associated with LTL, suggesting the presence of distinct causal alleles. No other risk loci for glioma were associated with LTL. The identification of risk alleles for glioma near TERC and TERT that also associate with telomere length implicates telomerase in gliomagenesis.
Project description:Immunoglobulin G (IgG) proteins are known for the huge diversity of the variable domains of their heavy and light chains, aimed at protecting each individual against foreign antigens. The IgG also harbor specific polymorphism concentrated in the CH2 and CH3-CHS constant regions located on the Fc fragment of their heavy chains. But this individual particularity relies only on a few amino acids among which some could make accurate sequence determination a challenge for mass spectrometry-based techniques. The purpose of the study was to bring a molecular validation of proteomic results by the sequencing of encoding DNA fragments. It was performed using ten individual samples (DNA and sera) selected on the basis of their Gm (gamma marker) allotype polymorphism in order to cover the main immunoglobulin heavy gamma (IGHG) gene diversity. Gm allotypes, reflecting part of this diversity, were determined by a serological method. On its side, the IGH locus comprises four functional IGHG genes totalizing 34 alleles and encoding the four IgG subclasses. The genomic study focused on the nucleotide polymorphism of the CH2 and CH3-CHS exons and of the intron. Despite strong sequence identity, four pairs of specific gene amplification primers could be designed. Additional primers were identified to perform the subsequent sequencing. The nucleotide sequences obtained were first assigned to a specific IGHG gene, and then IGHG alleles were deduced using a home-made decision tree reading of the nucleotide sequences. IGHG amino acid (AA) alleles were determined by mass spectrometry. Identical results were found at 95% between alleles identified by proteomics and those deduced from genomics. These results validate the proteomic approach which could be used for diagnostic purposes, namely for a mother-and-child differential IGHG detection in a context of suspicion of congenital infection.
Project description:Identifying modifiers of glioma risk in patients with type I neurofibromatosis (NF1) could help support personalized tumor surveillance, advance understanding of gliomagenesis, and potentially identify novel therapeutic targets. Here, we report genetic polymorphisms in the human adenylate cyclase gene adenylate cyclase 8 (ADCY8) that correlate with glioma risk in NF1 in a sex-specific manner, elevating risk in females while reducing risk in males. This finding extends earlier evidence of a role for cAMP in gliomagenesis based on results in a genetically engineered mouse model (Nf1 GEM). Thus, sexually dimorphic cAMP signaling might render males and females differentially sensitive to variation in cAMP levels. Using male and female Nf1 GEM, we found significant sex differences exist in cAMP regulation and in the growth-promoting effects of cAMP suppression. Overall, our results establish a sex-specific role for cAMP regulation in human gliomagenesis, specifically identifying ADCY8 as a modifier of glioma risk in NF1.
Project description:Point mutations at Arg132 of the cytoplasmic NADP(+)-dependent isocitrate dehydrogenase 1 (IDH1) occur frequently in gliomas and result in a gain of function to produce the "oncometabolite" D-2-hydroxyglutarate (D-2HG). The mutated IDH1 allele is usually associated with a wild-type IDH1 allele (heterozygous) in cancer. Here, we identify 2 gliomas that underwent loss of the wild-type IDH1 allele but retained the mutant IDH1 allele following tumor progression from World Health Organization (WHO) grade III anaplastic astrocytomas to WHO grade IV glioblastomas. Intratumoral D-2HG was 14-fold lower in the glioblastomas lacking wild-type IDH1 than in glioblastomas with heterozygous IDH1 mutations. To characterize the contribution of wild-type IDH1 to cancer cell D-2HG production, we established an IDH1-mutated astrocytoma (IMA) cell line from a WHO grade III anaplastic astrocytoma. Disruption of the wild-type IDH1 allele in IMA cells by gene targeting resulted in an 87-fold decrease in cellular D-2HG levels, showing that both wild-type and mutant IDH1 alleles are required for D-2HG production in glioma cells. Expression of wild-type IDH1 was also critical for mutant IDH1-associated D-2HG production in the colorectal cancer cell line HCT116. These insights may aid in the development of therapeutic strategies to target IDH1-mutated cancers.
Project description:KIAA1549-BRAF is the most frequently identified genetic mutation in sporadic pilocytic astrocytoma (PA), creating a fusion BRAF (f-BRAF) protein with increased BRAF activity. Fusion-BRAF-expressing neural stem cells (NSCs) exhibit increased cell growth and can generate glioma-like lesions following injection into the cerebella of naïve mice. Increased Iba1+ monocyte (microglia) infiltration is associated with murine f-BRAF-expressing NSC-induced glioma-like lesion formation, suggesting that f-BRAF-expressing NSCs attract microglia to establish a microenvironment supportive of tumorigenesis. Herein, we identify Ccl2 as the chemokine produced by f-BRAF-expressing NSCs, which is critical for creating a permissive stroma for gliomagenesis. In addition, f-BRAF regulation of Ccl2 production operates in an ERK- and NFκB-dependent manner in cerebellar NSCs. Finally, Ccr2-mediated microglia recruitment is required for glioma-like lesion formation in vivo, as tumor do not form in Ccr2-deficient mice following f-BRAF-expressing NSC injection. Collectively, these results demonstrate that f-BRAF expression creates a supportive tumor microenvironment through NFκB-mediated Ccl2 production and microglia recruitment.
Project description:Cytomegalovirus (CMV) has been ubiquitously detected within high-grade gliomas, but its role in gliomagenesis has not been fully elicited.Glioblastoma multiforme (GBM) tumors were analyzed by flow cytometry to determine CMV antigen expression within various glioma-associated immune populations. The glioma cancer stem cell (gCSC) CMV interleukin (IL)-10 production was determined by ELISA. Human monocytes were stimulated with recombinant CMV IL-10 and levels of expression of p-STAT3, VEGF (vascular endothelial growth factor), TGF-?, viral IE1, and pp65 were determined by flow cytometry. The influence of CMV IL-10-treated monocytes on gCSC biology was ascertained by functional assays.CMV showed a tropism for macrophages (M?)/microglia and CD133+ gCSCs within GBMs. The gCSCs produce CMV IL-10, which induces human monocytes (the precursor to the central nervous system M?s/microglia) to assume an M2 immunosuppressive phenotype (as manifested by downmodulation of the major histocompatibility complex and costimulatory molecules) while upregulating immunoinhibitory B7-H1. CMV IL-10 also induces expression of viral IE1, a modulator of viral replication and transcription in the monocytes. Finally, the CMV IL-10-treated monocytes produced angiogenic VEGF, immunosuppressive TGF-?, and enhanced migration of gCSCs.CMV triggers a feedforward mechanism of gliomagenesis by inducing tumor-supportive monocytes.
Project description:Glioblastoma multiformes (GBMs) are high-grade astrocytomas and the most common brain malignancies. Primary GBMs are often associated with disturbed RAS signaling, and expression of oncogenic HRAS results in a malignant phenotype in glioma cell lines. Secondary GBMs arise from lower-grade astrocytomas, have slower progression than primary tumors, and contain IDH1 mutations in over 70% of cases. Despite significant amount of accumulating genomic and transcriptomic data, the fundamental mechanistic differences of gliomagenesis in these two types of high-grade astrocytoma remain poorly understood. Only a few studies have attempted to investigate the proteome, phosphorylation signaling, and epigenetic regulation in astrocytoma. In the present study, we applied quantitative phosphoproteomics to identify the main signaling differences between oncogenic HRAS and mutant IDH1-driven glioma cells as models of primary and secondary GBM, respectively. Our analysis confirms the driving roles of the MAPK and PI3K/mTOR signaling pathways in HRAS driven cells and additionally uncovers dysregulation of other signaling pathways. Although a subset of the signaling changes mediated by HRAS could be reversed by a MEK inhibitor, dual inhibition of MEK and PI3K resulted in more complete reversal of the phosphorylation patterns produced by HRAS expression. In contrast, cells expressing mutant IDH1 did not show significant activation of MAPK or PI3K/mTOR pathways. Instead, global downregulation of protein expression was observed. Targeted proteomic analysis of histone modifications identified significant histone methylation, acetylation, and butyrylation changes in the mutant IDH1 expressing cells, consistent with a global transcriptional repressive state. Our findings offer novel mechanistic insight linking mutant IDH1 associated inhibition of histone demethylases with specific histone modification changes to produce global transcriptional repression in secondary glioblastoma. Our proteomic datasets are available for download and provide a comprehensive catalogue of alterations in protein abundance, phosphorylation, and histone modifications in oncogenic HRAS and IDH1 driven astrocytoma cells beyond the transcriptomic level.