Segregation of non‐p.R132H mutations in IDH1 in distinct molecular subtypes of glioma

Mutations in the gene encoding the isocitrate dehydrogenase 1 gene (IDH1) occur at a high frequency (up to 80%) in many different subtypes of glioma. In this study, we have screened for IDH1 mutations in a cohort of 496 gliomas. IDH1 mutations were most frequently observed in low grade gliomas with c.395G>A (p.R132H) representing >90% of all IDH1 mutations. Interestingly, non‐p.R132H mutations segregate in distinct histological and molecular subtypes of glioma. Histologically, they occur sporadically in classic oligodendrogliomas and at significantly higher frequency in other grade II and III gliomas. Genetically, non‐p.R132H mutations occur in tumors with TP53 mutation, are virtually absent in tumors with loss of heterozygosity on 1p and 19q and accumulate in distinct (gene‐expression profiling based) intrinsic molecular subtypes. The IDH1 mutation type does not affect patient survival. Our results were validated on an independent sample cohort, indicating that the IDH1 mutation spectrum may aid glioma subtype classification. Functional differences between p.R132H and non‐p.R132H mutated IDH1 may explain the segregation in distinct glioma subtypes. © 2010 Wiley‐Liss, Inc.     

IDH‐mutant brainstem gliomas in adolescent and young adult patients: Report of three cases and review of the literature

Isocitrate dehydrogenase (IDH) mutations are rare in pediatric and adolescent gliomas. We recently identified three adolescent/young adult (AYA) patients with IDH‐mutant low grade gliomas of the brainstem with several key clinicopathologic and molecular features in common. We discuss these three cases and review the current literature.     

Molecular subtypes of glioma identified by genome‐wide methylation profiling

Recent studies have indicated a prognostic role for genome‐wide methylation in gliomas: Tumors that show an overall increase in DNA methylation at CpG sites (CIMP+; CpG island methylator phenotype) have a more favorable prognosis than CIMP? gliomas. Here, we have determined whether methylation profiling can identify more and clinically relevant molecular subtypes of glioma by performing genome‐wide methylation profiling on 138 glial brain tumors of all histological diagnosis. Hopach (Hierarchical ordered partitioning and collapsing hybrid) clustering using the 1,000 most variable CpGs identified three distinct glioma subtypes (C+_1p19q, C+_wt, and C?) and one adult brain subtype. All “C+_1p19q” and “C+_wt” tumors were CIMP+ whereas most (50/54) “C?” tumors were CIMP?. The C? subtype gliomas contained many glioblastomas and all pilocytic astrocytomas. 1p19q LOH was frequent in the C+_1p19q subtype. Other genetic changes (IDH1 mutation and EGFR amplification) and gene‐expression based molecular subtypes also segregated in distinct methylation subtypes, demonstrating that these subtypes are also genetically distinct. Each subtype was associated with its own prognosis: median survival for C?, C+_1p19q, and C+_wt tumors was 1.18, 5.00, and 2.62 years, respectively. The prognostic value of these methylation subtypes was validated on an external dataset from the TCGA. Analysis of recurrences of 14 primary tumors samples indicates that shifts between some C+_wt and C+_1p/19q tumors can occur between the primary and recurrent tumor, but CIMP status remained stable. Our data demonstrate that methylation profiling identifies at least three prognostically relevant subtypes of glioma that can aid diagnosis and potentially guide treatment for patients. © 2013 Wiley Periodicals, Inc.     

Epithelioid Glioblastomas and Anaplastic Epithelioid Pleomorphic Xanthoastrocytomas—Same Entity or First Cousins?

Epithelioid glioblastoma (eGBM) and pleomorphic xanthoastrocytoma (PXA) with anaplastically transformed foci (ePXA) show overlapping features. Eleven eGBMs and 5 ePXAs were reviewed and studied immunohistochemically. Fluorescence in situ hybridization for EGFR amplification, PTEN deletion and ODZ3 deletion was also performed, with Ilumina 450 methylome analysis obtained in five cases. The average age for eGBM was 30.9 (range 2–79) years, including five pediatric cases and a M : F ratio of 4.5. The ePXA patients had a M : F ratio of 4 and averaged 21.2 (range 10–38) years in age, including two pediatric cases. Six eGBMs and two ePXAs recurred (median recurrence interval of 12 and 3.3 months, respectively). All tumors were composed of solid sheets of loosely cohesive, “melanoma‐like” cells with only limited infiltration. ePXAs showed lower grade foci with classic features of PXA. Both tumor types showed focal expression of epithelial and glial markers, retained INI1 and BRG1 expression, occasional CD34 positivity, and lack of mutant IDH1 (R132H) immunoreactivity. BRAF V600E mutation was present in four eGBMs and four ePXAs. ODZ3 deletion was detected in seven eGBMs and two ePXAs. EGFR amplification was absent. Methylome analysis showed that one ePXA and one eGBM clustered with PXAs, one eGBM clustered with low‐grade gliomas, and two eGBMs clustered with pediatric‐type glioblastomas. Common histologic, immunohistochemical, molecular and clinical features found in eGBM and ePXA suggest that they are closely related or the same entity. If the latter is true, the nomenclature and WHO grading remains to be resolved.     

Bithalamic gliomas may be molecularly distinct from their unilateral high‐grade counterparts

Bithalamic gliomas are rare cancers diagnosed based on poorly defined radiologic criteria. Infiltrative astrocytomas account for most cases. While some previous studies reported dismal outcomes for patients with bithalamic gliomas irrespective of therapy and histologic grade, others described better prognoses even without anticancer therapy. Little is known about their molecular characteristics. We reviewed clinical, radiologic, and histologic features of patients with bithalamic gliomas treated at our institution over 15 years. Targeted sequencing of mutational hotspots in H3F3A, HIST1H3B, IDH1/2, and BRAF, and genome‐wide analysis of DNA methylation and copy number abnormalities was performed in available tumors. Eleven patients with bithalamic gliomas were identified. Their median age at diagnosis was 4.8 years (range: 1–15.7). Additional involvement of the brainstem, basal ganglia, and cerebral lobes occurred in 11, 9, and 3 cases, respectively. All patients presented with hydrocephalus. Two‐thirds of the patients had a histologic diagnosis of anaplastic astrocytoma. Despite aggressive therapy, our youngest patient, the only one diagnosed before 1 year of age, is the sole long‐term survivor. DNA methylation could be performed in seven tumors, all of which clustered with the RTK I ‘PDGFRA’ subgroup by unsupervised hierarchical analysis of methylation array against a previously published cohort of 59 pediatric high‐grade gliomas. Sequencing of hotspots mutations could be done in 10 tumors, none of which harbored H3F3A p.K27 and/or the respective DNA methylation signature, and any other hotspot mutations. Amplification of MDM4 (n = 2), PDGFRA (n = 2), and ID2 combined with MYCN (n = 1) were observed in 7 tumors available for analysis. In comparison with the previously published experience with unilateral high‐grade thalamic astrocytomas where H3F3A p.K27 was present in two‐thirds of cases, the absence of this molecular subgroup in bithalamic gliomas was striking. This finding suggests that unilateral and bithalamic high‐grade gliomas may represent two distinct molecular entities.     

The hypermethylation of the O6-methylguanine-DNA methyltransferase gene promoter in gliomas--correlation with array comparative genome hybridization results and IDH1 mutation

The use of molecular markers in the diagnostics of gliomas aids histopathological diagnosis and allows their further classification into clinically significant subgroups. The aim of this study was to characterize the methylation pattern of the O(6) -methylguanine-DNA methyltransferase (MGMT) promoter, gene copy number aberrations, and isocitrate dehydrogenase I (IDH1) mutation in gliomas. We studied 51 gliomas (15 oligodendrogliomas, 18 oligoastrocytomas, 3 astrocytomas, and 15 glioblastomas) by pyrosequencing, array comparative genome hybridization (CGH), and immunohistochemistry. MGMT hypermethylation was observed in 100% of oligoastrocytomas, 93% of oligodendrogliomas, and 47% of glioblastomas. The most frequently altered chromosomal regions were deletions of 1p31.1/21.1-22.2 and 19q13.3qter in oligodendroglial tumors, and losses of 9p21.3, 10q25.3qter, and 10q26.13-26.2 in glioblastomas. Deletions on 9p and 10q, and gain of 7p were associated with the unmethylated MGMT phenotype, whereas deletion of 19q and oligodendroglial morphology was associated with MGMT hypermethylation. IDH1 mutation showed positive correlation with MGMT hypermethylation and loss of 1p/19q. Our results suggest that MGMT promoter methylation, analyzed by pyrosequencing, is a frequent event in oligodendroglial tumors, and it correlates with IDH1 mutation and 19q loss in gliomas. Pyrosequencing proved a good method for assessing the degree of MGMT methylation in formalin-fixed paraffin-embedded glioma samples. However, further studies are needed to confirm a clinically relevant cut-off point for MGMT methylation in gliomas.     

2‐Hydoxyglutarate: D/Riving Pathology in gLiomaS

Common pathways and mechanisms can be found in both cancers and inborn errors of metabolism. 2‐Hydroxyglutarate (2‐HG) acidurias and isocitrate dehydrogenase (IDH) 1/2 mutant tumors are examples of this phenomenon. 2‐HG can exist in two chiral forms, D(R)‐2‐HG and L(S)‐2‐HG, which are elevated in D‐ and L‐acidurias, respectively. D‐2‐HG was subsequently discovered to be synthesized in IDH 1/2 mutant tumors including ～70% of intermediate‐grade gliomas and secondary glioblastomas (GBM). Recent studies have revealed that L‐2‐HG is generated in hypoxia in IDH wild‐type tumors. Both 2‐HG enantiomers have similar structures as α‐ketoglutarate (α‐KG) and can competitively inhibit α‐KG‐dependent enzymes. This inhibition modulates numerous cellular processes, including histone and DNA methylation, and can ultimately impact oncogenesis. D‐2‐HG can be detected in vivo in glioma patients and animal models using advanced imaging modalities. Finally, pharmacologic inhibitors of mutant IDH 1/2 attenuate the production of D‐2‐HG and show great promise as therapeutic agents.     

Congenital tumors of the central nervous system: an institutional review of 64 cases with emphasis on tumors with unique histologic and molecular characteristics

Congenital brain tumors are rare accounting for 0.5%–1.9% of all pediatric brain tumors. While different criteria have been used to classify a tumor as congenital, those diagnosed prior to 6 months of age are considered to be “probably” congenital in origin. We performed an institutional review of all central nervous system (CNS) tumors (surgical and autopsy specimens from 1990 to 2019) in patients less than 6 months old. Sixty‐four unique cases were identified, and these accounted for 2.0% of all CNS tumor specimens at our institution. The most common tumor types were high‐grade gliomas, low‐grade gliomas and medulloblastomas. Atypical teratoid rhabdoid tumors, choroid plexus tumors and germ cell tumors also accounted for a significant portion of the cohort. Seven tumors were diagnosed prenatally. The most common clinical presentation at diagnosis was increased head circumference. At the conclusion of the study, over half of the patients were alive including all patients with WHO grade I and II tumors. Ninety‐two percent of cases were classifiable using the 2016 WHO system, and when available, molecular findings supported the histologic diagnoses. However, several gliomas had unusual histologic features and did not correspond to a well‐defined entity. Molecular testing was essential for accurate classification of a subset of these tumors, and several high‐grade gliomas exhibited fusions considered unique to infantile gliomas, including those involving the MET, ALK and NTRK genes. To our knowledge, this cohort represents the largest single‐institution study of congenital CNS tumors and highlights many ways in which congenital CNS tumors are distinct from CNS tumors of older pediatric patients and adults.     

Diffuse Midline Gliomas with Histone H3‐K27M Mutation: A Series of 47 Cases Assessing the Spectrum of Morphologic Variation and Associated Genetic Alterations

Somatic mutations of the H3F3A and HIST1H3B genes encoding the histone H3 variants, H3.3 and H3.1, were recently identified in high‐grade gliomas arising in the thalamus, pons and spinal cord of children and young adults. However, the complete range of patients and locations in which these tumors arise, as well as the morphologic spectrum and associated genetic alterations remain undefined. Here, we describe a series of 47 diffuse midline gliomas with histone H3‐K27M mutation. The 25 male and 22 female patients ranged in age from 2 to 65 years (median = 14). Tumors were centered not only in the pons, thalamus, and spinal cord, but also in the third ventricle, hypothalamus, pineal region and cerebellum. Patients with pontine tumors were younger (median = 7 years) than those with thalamic (median = 24 years) or spinal (median = 25 years) tumors. A wide morphologic spectrum was encountered including gliomas with giant cells, epithelioid and rhabdoid cells, primitive neuroectodermal tumor (PNET)‐like foci, neuropil‐like islands, pilomyxoid features, ependymal‐like areas, sarcomatous transformation, ganglionic differentiation and pleomorphic xanthoastrocytoma (PXA)‐like areas. In this series, histone H3‐K27M mutation was mutually exclusive with IDH1 mutation and EGFR amplification, rarely co‐occurred with BRAF‐V600E mutation, and was commonly associated with p53 overexpression, ATRX loss (except in pontine gliomas), and monosomy 10.     

Chromatin Remodeling Defects in Pediatric and Young Adult Glioblastoma: A Tale of a Variant Histone 3 Tail

Primary brain tumors occur in 8 out of 100 000 people and are the leading cause of cancer‐related death in children. Among brain tumors, high‐grade astrocytomas (HGAs) including glioblastoma multiforme (GBM) are aggressive and are lethal human cancers. Despite decades of concerted therapeutic efforts, HGAs remain essentially incurable in adults and children. Recent discoveries have revolutionized our understanding of these tumors in children and young adults. Recurrent somatic driver mutations in the tail of histone 3 variant 3 (H3.3), leading to amino acid substitutions at key residues, namely lysine (K) 27 (K27M) and glycine 34 (G34R/G34V), were identified as a new molecular mechanism in pediatric GBM. These mutations represent the pediatric counterpart of the recurrent mutations in isocitrate dehydrogenases (IDH) identified in young adult gliomas and provide a much‐needed new pathway that can be targeted for therapeutic development. This review will provide an overview of the potential role of these mutations in altering chromatin structure and affecting specific molecular pathways ultimately leading to gliomagenesis. The distinct changes in chromatin structure and the specific downstream events induced by each mutation need characterizing independently if progress is to be made in tackling this devastating cancer.     

Multimodal molecular analysis of astroblastoma enables reclassification of most cases into more specific molecular entities

Astroblastoma is a rare and controversial glioma with variable clinical behavior. The diagnosis currently rests on histologic findings of a circumscribed glioma with astroblastomatous pseudorosettes and vascular hyalinization. Immunohistochemical studies have suggested different oncogenic drivers, such as BRAF p.V600E, but very few cases have been studied using genome‐wide methodologies. Recent genomic profiling identified a subset of CNS embryonal tumors with astroblastoma‐like morphology that harbored MN1 gene fusions, termed “CNS high‐grade neuroepithelial tumors with MN1 alteration” (CNS‐HGNET‐MN1). To further characterize the genetic alterations that drive astroblastomas, we performed targeted next‐generation sequencing (NGS) of 500 cancer‐associated genes in a series of eight cases. We correlated these findings with break‐apart fluorescence in situ hybridization (FISH) analysis of the MN1 locus and genome‐wide DNA methylation profiling. Four cases showed MN1 alteration by FISH, including two pediatric cases that lacked other pathogenic alterations, and two adult cases that harbored other cancer‐associated gene mutations or copy number alterations (eg, CDKN2A/B homozygous deletion, TP53, ATM and TERT promoter mutations). Three of these cases grouped with the CNS‐HGNET‐MN1 entity by methylation profiling. Two of four MN1 intact cases by FISH showed genetic features of either anaplastic pleomorphic xanthoastrocytoma (BRAF p.V600E mutation, CDKN2A/B homozygous deletion and TERT promoter mutation) or IDH‐wildtype glioblastoma (trisomy 7, monosomy 10, CDK4 amplification and TP53, NRAS and TERT promoter mutations) and these cases had an aggressive clinical course. Two clinically indolent cases remained unclassifiable despite multimodal molecular analysis. We conclude that astroblastoma histology is not specific for any entity including CNS‐HGNET‐MN1, and that additional genetic characterization should be considered for astroblastomas, as a number of these tumors likely contain a methylation profile or genetic alterations that suggest classification as other tumor entities. Our heterogeneous molecular findings help to explain the clinical unpredictability of astroblastoma.     

IDH1 mutations in gliomas: first series from a tertiary care centre in India with comprehensive review of literature

Object

Mutations of the gene encoding isocitrate dehydrogenase (IDH) have been shown in a significant proportion of diffuse gliomas. These mutations are specific to gliomas and their utility for diagnosis and prognostication of these tumors is being proclaimed. The present study was conducted with the aim of assessing frequency of IDH1 mutations in gliomas, their correlation with other molecular alterations along with a comprehensive review of available literature.

Methods

A total of 100 gliomas of various grades and subtypes from Indian patients were screened for assessing frequency of IDH1 mutations. The findings were correlated with TP53 mutations, 1p/19q deletion, EGFR amplification and PTEN deletion status. The detailed comprehensive review of literature was performed comparing all studies available till date.

Results

IDH1 mutations in codon 132 were observed in 46% cases. The frequency was 68.8% in grade II, 85.7% in grade III and 12.8% in GBMs. R132H mutation was most frequent (84.8%). Overall frequency of these mutations was relatively higher in oligodendroglial tumours as compared to astrocytic phenotype (66.7% versus 38.4%; p = 0.06). Primary GBMs showed IDH1 mutation in only 4.4% cases. In contrast, 66.7% of secondary GBMs harboured this alteration. Patients with IDH1 mutations were significantly younger as compared to those without mutation (p = 0.001). There was a significant correlation between IDH1 mutation and TP53 mutation (p = 0.004). Although IDH1 mutation showed a positive correlation with 1p/19q deletion, the association was not statistically significant (p = 0.653). There was no correlation with EGFR amplification or PTEN deletion.

Conclusion

IDH1 mutations are present in large proportion of Indian patients with diffuse astrocytic and oligodendroglial neoplasms similar to the reported literature form west. The frequency is lower in primary GBMs and as compared to secondary GBMs. Association with younger age and positive correlation with TP53 mutation and 1p/19q loss is observed. More importantly it is emerging as an independent prognostic marker. Hence the greatest challenge now is establishing a reliable user friendly test for incorporating this novel genetic alteration to routine clinical practice.     

Alterations in the RB1 pathway in low-grade diffuse gliomas lacking common genetic alterations

We recently reported that the vast majority (>90%) of low‐grade diffuse gliomas (diffuse astrocytoma, oligoastrocytoma and oligodendroglioma) carry at least one of the following genetic alterations: IDH1/2 mutation, TP53 mutation or 1p/19q loss. Only 7% of cases were triple‐negative (ie, lacking any of these alterations). In the present study, array comparative genomic hybridization (CGH) in 15 triple‐negative WHO grade II gliomas (eight diffuse astrocytomas and seven oligodendrogliomas) showed loss at 9p21 (p14^ARF, p15^INK4b, p16^INK4a loci) and 13q14–13q32 (containing the RB1 locus) in three and two cases, respectively. Further analyses in 31 triple‐negative cases as well as a total of 160 non‐triple‐negative cases revealed that alterations in the RB1 pathway (homozygous deletion and promoter methylation of the p15^INK4b, p16^INK4a and RB1 genes) were significantly more frequent in triple‐negative (26%) than in non‐triple‐negative cases (11%; P = 0.0371). Multivariate analysis after adjustment for age, histology and treatment showed that RB1 pathway alterations were significantly associated with unfavorable outcome for patients with low‐grade diffuse glioma [hazard ratio, 3.024 (1.279–6.631); P = 0.0057]. These results suggest that a fraction of low‐grade diffuse gliomas lacking common genetic alterations may develop through a distinct genetic pathway, which may include loss of cell‐cycle control regulated by the RB1 pathway.     

IDH1R132H在中枢神经系统胶质瘤中的表达及其鉴别诊断意义

目的 探讨异柠檬酸盐脱氢酶1基因(isocitrate dehydrogenase 1 gene,IDH1)突变的表达产物IDH1R132H在人中枢神经系统胶质瘤中的表达及其在鉴别诊断中的意义.方法 应用免疫组织化学EnVision法检测不同级别胶质瘤75例(包括WHOⅡ级33例,Ⅲ级20例和Ⅳ级22例)与各种原因造成的胶质增生性脑组织中IDH1R132H的表达情况,并与p53的表达情况进行比较分析.结果 IDH1R132H在WHOⅡ级、Ⅲ级和Ⅳ级胶质瘤的表达阳性率分别为57.6%(19/33)、40.0%(8/20)和27.3%(6/22),差异有统计学意义(P=0.024).IDH1R132H的具体表达部位在胶质瘤细胞的胞质、突起以及部分细胞核.除肿瘤主体部分密集增生的肿瘤细胞呈阳性表达以外,包括皮质和白质在内的肿瘤周边区域单个或散在的肿瘤细胞也呈强阳性表达.IDH1R132H阳性病例有额叶受累为主的倾向.胶质增生组织、毛细胞型星形细胞瘤均为阴性表达.另外,p53在WHOⅡ级、Ⅲ级和Ⅳ级胶质瘤的表达阳性率分别为42.4%(14/33)、65.0%(13/20)和77.3%(17/22).IDH1R132H的表达与p53的表达之间没有明确的相关性(P=0.766).结论 IDH1R132H在各级别胶质瘤中均有表达,且随着肿瘤级别的增高呈表达下降趋势.IDH1R132H可以作为鉴别低级别胶质瘤和胶质增生性病变的一个有用的分子标志物.

Abstract：

Objective To investigate the immunohistochemical expression of isocitrate dehydrogenase 1 gene ( IDH1 ) R132H in glioma and its diagnostic utility. Methods Immunohistochemical study of IDH1R132H expression was performed on formalin-fixed paraffin-embedded tissue samples of 75 gliomas, including 33 cases of grade Ⅱ , 20 cases of grade Ⅲ and 22 cases of grade Ⅳ tumors. Six cases of pilocytic astrocytoma and 12 cases of gliosis were used as controls. Results Nineteen in 33 cases of grade Ⅱ (57.6%), 8 in 20 cases of grade Ⅲ (40. 0% ), 6 in 22 cases of grade Ⅳ (27. 3% ) showed positive cytoplasmic staining of IDH1R132H. Scattered invasive glioma cells at the tumor periphery also expressed IDH1R132H. Gliomas involving the frontal lobe showed more strong IDH1R132H staining. In contrast, none of the pilocytic astrocytomas and gliosis showed IDH1R132H staining. Moreover, the rate of p53 immunopositivities were 42. 4% ( 14/33 ) in grade Ⅱ , 65.0% (13/20) in grade Ⅲ and 77.3% (17/22) in grade Ⅳ gliomas. There were no statistic correlations between expression of IDH1R132H and p53.Conclusion IDH1R132H tends to express preferentially in low-grade gliomas, and it thus may serve as a valuable marker in distinguishing low grade gliomas from gliosis.     

Isocitrate dehydrogenase 1 analysis differentiates gangliogliomas from infiltrative gliomas

Recent work has identified novel point mutations in isocitrate dehydrogenase 1 (IDH1) in the majority of the World Health Organization grades II and III infiltrative gliomas and secondary grade IV glioblastomas. Gangliogliomas consist of neoplastic ganglion and glial cells and, in contrast to infiltrative gliomas, are generally indolent. Yet distinguishing between a ganglioglioma and an infiltrative glioma with admixed gray matter can be difficult, perhaps accounting for some "gangliogliomas" that ultimately show aggressive behavior. In this multi-institutional study, 98 cases originally diagnosed as ganglioglioma were analyzed for IDH1 mutations, 86 of which had follow-up data available. Eight cases (8.2%) were positive for R132H IDH1 mutations; six had silent IDH2 mutations and two had nonsense IDH2 mutations. The presence of mutant IDH1 in gangliogliomas correlated with a greater risk of recurrence (P=0.0007) and malignant transformation and/or death (P<0.0001) compared with tumors that were IDH1 wild type. Furthermore, the age of patients with IDH1-mutant gangliogliomas was higher than those without mutations (25.5 vs. 46.1 years, P=0.0033). IDH1/2 testing of tumors suspected of being gangliogliomas may therefore be advisable, particularly in the adult population.     

Molecular classification of low-grade diffuse gliomas

The current World Health Organization classification recognizes three histological types of grade II low-grade diffuse glioma (diffuse astrocytoma, oligoastrocytoma, and oligodendroglioma). However, the diagnostic criteria, in particular for oligoastrocytoma, are highly subjective. The aim of our study was to establish genetic profiles for diffuse gliomas and to estimate their predictive impact. In this study, we screened 360 World Health Organization grade II gliomas for mutations in the IDH1, IDH2, and TP53 genes and for 1p/19q loss and correlated these with clinical outcome. Most tumors (86%) were characterized genetically by TP53 mutation plus IDH1/2 mutation (32%), 1p/19q loss plus IDH1/2 mutation (37%), or IDH1/2 mutation only (17%). TP53 mutations only or 1p/19q loss only was rare (2 and 3%, respectively). The median survival of patients with TP53 mutation ± IDH1/2 mutation was significantly shorter than that of patients with 1p/19q loss ± IDH1/2 mutation (51.8 months vs. 58.7 months, respectively; P = 0.0037). Multivariate analysis with adjustment for age and treatment confirmed these results (P = 0.0087) and also revealed that TP53 mutation is a significant prognostic marker for shorter survival (P = 0.0005) and 1p/19q loss for longer survival (P = 0.0002), while IDH1/2 mutations are not prognostic (P = 0.8737). The molecular classification on the basis of IDH1/2 mutation, TP53 mutation, and 1p/19q loss has power similar to histological classification and avoids the ambiguity inherent to the diagnosis of oligoastrocytoma.     

Characterization of R132H Mutation-specific IDH1 Antibody Binding in Brain Tumors

Heterozygous point mutations of isocitrate dehydrogenase (IDH)1 codon 132 are frequent in grade II and III gliomas. Recently, we reported an antibody specific for the IDH1R132H mutation. Here we investigate the capability of this antibody to differentiate wild type and mutated IDH1 protein in central nervous system (CNS) tumors by Western blot and immunohistochemistry. Results of protein analysis are correlated to sequencing data. In Western blot, anti-IDH1R132H mouse monoclonal antibody mIDH1R132H detected a specific band only in mutated tumors. Immunohistochemistry of 345 primary brain tumors demonstrated a strong cytoplasmic and weaker nuclear staining in 122 cases. Correlation with direct sequencing of 186 cases resulted in consensus of 177 cases. Genetic retesting of cases with conflicting findings resulted in a match of 186/186 cases, with all discrepancies resolving in favor of immunohistochemistry. Intriguing is the ability of mIDH1R132H to detect single infiltrating tumor cells. The very high frequency and the distribution of this mutation among specific brain tumor entities allow the highly sensitive and specific discrimination of various tumors by immunohistochemistry, such as anaplastic astrocytoma from primary glioblastoma or diffuse astrocytoma World Health Organization (WHO) grade II from pilocytic astrocytoma or ependymoma. Noteworthy is the discrimination of the infiltrating edge of tumors with IDH1 mutation from reactive gliosis.     

PDGFRA Amplification is Common in Pediatric and Adult High‐Grade Astrocytomas and Identifies a Poor Prognostic Group in IDH1 Mutant Glioblastoma

High‐grade astrocytomas (HGAs), corresponding to World Health Organization grades III (anaplastic astrocytoma) and IV (glioblastoma; GBM), are biologically aggressive, and their molecular classification is increasingly relevant to clinical management. PDGFRA amplification is common in HGAs, although its prognostic significance remains unclear. Using fluorescence in situ hybridization (FISH), the most sensitive technique for detecting PDGFRA copy number gains, we determined PDGFRA amplification status in 123 pediatric and 263 adult HGAs. A range of PDGFRA FISH patterns were identified and cases were scored as non‐amplified (normal and polysomy) or amplified (low‐level and high‐level). PDGFRA amplification was frequent in pediatric (29.3%) and adult (20.9%) tumors. Amplification was not prognostic in pediatric HGAs. In adult tumors diagnosed initially as GBM, the presence of combined PDGFRA amplification and isocitrate dehydrogenase 1 (IDH1)^R132H mutation was a significant independent prognostic factor (P = 0.01). In HGAs, PDGFRA amplification is common and can manifest as high‐level and focal or low‐level amplifications. Our data indicate that the latter is more prevalent than previously reported with copy number averaging techniques. To our knowledge, this is the largest survey of PDGFRA status in adult and pediatric HGAs and suggests PDGFRA amplification increases with grade and is associated with a less favorable prognosis in IDH1 mutant de novo GBMs.     

Prognostic and radiographic correlates of a prospectively collected molecularly profiled cohort of IDH1/2‐wildtype astrocytomas

Background: In the molecular era, the relevance of tumor grade for prognostication of IDH1/2‐wildtype (WT) gliomas has been debated. It has been suggested that histologic grade II and III astrocytomas with molecular features of glioblastoma, IDH1/2‐WT have a similar prognosis to glioblastoma and should be considered for the same clinical trials. Methods: We integrated prospective clinical sequencing from 564 patients with IDH1/2‐WT gliomas (26 grade II, 71 grade III and 467 grade IV) with clinical and radiographic data to assess associations between molecular features, grade and outcome. Results: Compared to histologic grade IV IDH1/2‐WT astrocytomas, histologic grade II astrocytomas harbor fewer chromosome 7/10 alterations (P = 0.04), EGFR amplifications (P = 0.022) and alterations in cell‐cycle effectors (P = 1.9e‐11), but a similar frequency of TERT promoter mutations. In contrast, there is no difference in the frequency of these canonical molecular features in histologic grade III vs. IV IDH1/2‐WT disease. Progression‐free (PFS) and overall survival (OS) for histologic grade II tumors were significantly longer than grade III tumors (P = 0.02 and P = 0.008, respectively), whereas there was no difference in PFS and OS for histologic grade III compared to grade IV tumors. Median PFS for histologic grade II, III and IV tumors was 19, 11 and 9 months, respectively. Median OS for the same tumors was 44, 23 and 23 months, respectively. In histologic grade II and III IDH1/2 WT tumors, gliomatosis is associated with the absence of cell‐cycle alterations (P = 0.008) and enriched in grade II features (P = 0.1) and alterations in the PI3K‐AKT pathway (P = 0.09). Conclusions: Grade II histology has genotypic and phenotypic associations with prognostic implications in IDH1/2‐WT astrocytomas.