Meningeal hemangiopericytomas: A clinicopathological study with emphasis on MGMT (O6‐methylguanine‐DNA methyltransferase) promoter methylation status

Meningeal hemangiopericytomas (HPCs) are aggressive dural‐based tumors, for which no prognostic or predictive marker has been identified. Gross total resection is treatment of choice, but not easily achieved; hence, alkylating agents like temozolomide (TMZ) are now being tried. O⁶‐methylguanine‐DNA methyltransferase (MGMT) promoter methylation has proven prognostic and predictive value in glioblastomas. This study evaluates MGMT promoter methylation in meningeal HPCs to determine its role in HPC oncogenesis and its association with patient outcome. Meningeal HPCs diagnosed between 2002 and 2011 were retrieved and clinicopathological features reviewed. MGMT promoter methylation status was assessed by methylation‐specific polymerase chain reaction (MSP) and immunohistochemistry (IHC) for MGMT protein. HPCs accounted for 1.1% of all CNS tumors. Forty cases were analyzed; the majority were adults (mean age = 41.4 years). Seventy percent were primary and 30% were recurrent tumors; 60% were grade II and 40% were grade III. MGMT promoter methylation was identified in 45% of cases, including Grade II (54.2%) and Grade III (31.3%) (P = 0.203). Promoter methylation was significantly (P = 0.035) more frequent in primary (57.1%) than in recurrent (16.7%) tumors. No correlation was noted between MGMT promoter methylation by MSP and MGMT protein expression by IHC, or with progression‐free survival. Thus, a significant proportion of HPCs demonstrate MGMT promoter methylation, suggesting possible susceptibility to TMZ. As promoter methylation is more frequent in primary tumors, TMZ may serve as a therapeutic option in residual primary tumors. Epigenetic inactivation of MGMT in HPCs necessitates the assessment of prognostic and predictive value of MGMT promoter methylation in HPCs in larger clinical trials.     

A comparative immunohistochemistry of O6‐methylguanine–DNA methyltransferase and p53 in diffusely infiltrating astrocytomas

The DNA repair protein O⁶‐methylguanine–DNA methyltransferase (MGMT) removes mutagenic adducts from the O⁶ position of guanine, thereby protecting the genome against guanine : cytosine to adenine : thymine transition and, meanwhile, conferring tumor resistance to many anticancer alkylating agents commonly used in the treatment of malignant gliomas. Studies on the involvement of p53 protein in expression of the MGMT gene have provided conflicting results regarding the relation between p53 protein and MGMT gene expression. To examine the potential immunostaining pattern of MGMT expression and to evaluate the possible relationship between p53 and MGMT regulation, we assessed MGMT and p53 accumulation on 35 cases of diffusely infiltrating astrocytomas. With a few cases showing cytoplasmic staining, MGMT accumulation was mainly nuclear. The percentage of labeled tumor cells was lower in high‐grade astrocytomas than in low‐grade astrocytomas (P < 0.05). Additionally, p53‐immunopositive tumor cells were usually immunonegative to MGMT. Thus, it is suggested that MGMT expression is reduced during malignant transformation of diffusely infiltrating astrocytomas, and that mutant p53 protein might be associated with down regulation of the MGMT expression.     

Pitfalls in the assessment of MGMT expression and in its correlation with survival in diffuse astrocytomas: proposal of a feasible immunohistochemical approach

Immunohistochemical studies showed that O₆-methylguanine-DNA methyltransferase (MGMT) protein expression is negatively associated with survival in glioblastomas treated with alkylating agents in accordance with previous results of methylation-specific PCR. Implementation of this data in routine clinical diagnostics is limited due to often inappropriate study designs, e.g. pooling of tumor entities, WHO grades or primary and secondary glioblastomas, disregard concerning the infiltration zone or various epidemiological factors. The aim of our study was to evaluate MGMT expression and its prognostic value taking into consideration the aforementioned deficiencies. For this, 162 astrocytic tumors WHO II–IV (36 diffuse astrocytomas WHO II, 51 anaplastic astrocytomas, 75 primary glioblastomas) as well as 25 glioblastoma infiltration zones and 19 glioblastoma relapses were analyzed for immunohistochemical MGMT protein expression using tissue microarray technique. Expression of MGMT significantly decreased from WHO grade II (25.6%) to glioblastoma (16.8%, p = 0.01) with lowest levels in grade III tumors (10.2%, II/III p < 0.0001). Significant negative associations of MGMT and survival were detected for WHO grade II and IV (p = 0.003 and 0.013). The optimal cut-off value of MGMT positive nuclei in primary glioblastomas discriminating patients with significantly different survival rates was at 15% (Log–Rank p = 0.0002). Individual relapse tumors showed changes of MGMT expression to a varying degree. The infiltration zone demonstrated a significant increase of MGMT (p < 0.0001). We conclude that immunohistochemical MGMT assessment has potential as a powerful diagnostic tool but analysis should only be performed in a grade dependent manner, before radio-/chemotherapy and with special attention to the infiltration zone of diffuse astrocytomas.     

Immunohistochemical evaluation of O6‐methylguanine DNA methyltransferase (MGMT) expression in 117 cases of glioblastoma

Temozolomide (TMZ) is an oral alkylating agent which is widely used in the treatment of glioblastoma (GBM) and is composed of astrocytic and/or oligodendroglial tumors, and the evaluation of O⁶‐methylguanine DNA methyltransferase (MGMT) expression is important to predict the response to TMZ therapy. In this study, we conducted immunohistochemical analysis of 117 cases of Japanese GBM including 19 cases of GBM with oligodendroglioma component (GBMO), using a scoring system for quantitative evaluation of staining intensity and proportion of MGMT, and performed survival analysis of these patients. Immunohistochemically, 55 cases (47%) were positive for MGMT with various intensities and proportions (total score (TS) ≥ 2), while 62 cases (53%) were negative (TS = 0). The distribution of MGMT expression pattern was not affected by any clinicopathological parameters such as the histological subtype (GBM vs. GBMO), age and gender. The survival analysis of these patients revealed that the minimal expression of MGMT (TS ≥ 2) was a significant unfavorable prognostic factor (P < 0.001) as well as resectability (P = 0.004). Moreover, multivariate analysis showed that minimal MGMT expression in GBM was the most potent independent predictor for progression free survival (P < 0.001) and also overall patient survival (P < 0.001). This is the first report employing the scoring system for both staining intensity and proportion to evaluate immunohistochemical MGMT expression in GBM. In addition, our results emphases the clinicopathological values of the immunohistochemical approach for MGMT expression in glioma patients as a routine laboratory examination.     

Immunohistochemical profiles of IDH1, MGMT and P53: Practical significance for prognostication of patients with diffuse gliomas

Genetic and epigenetic status, including mutations of isocitrate dehydrogenase (IDH) and TP53 and methylation of O⁶‐methylguanine‐DNA methyltransferase (MGMT), are associated with the development of various types of glioma and are useful for prognostication. Here, using routinely available histology sections from 312 patients with diffuse gliomas, we performed immunohistochemistry using antibodies specific for IDH1 mutation, MGMT methylation status, and aberrant p53 expression to evaluate the possible prognostic significance of these features. With regard to overall survival (OS), univariate analysis indicated that an IDH1‐positive profile in patients with glioblastoma (GBM), anaplastic astrocytoma (AA), anaplastic oligoastrocytoma and oligodendroglioma, or a MGMT‐negative profile in patients with GBM and AA were significantly associated with a favorable outcome. Multivariate analysis revealed that both profiles were independent factors influencing prognosis. The OS of patients with IDH1‐positive/MGMT‐negative profiles was significantly longer than that of patients with negative/negative and negative/positive profiles. A p53 profile was not an independent prognostic factor. However, for GBM/AA patients with IDH1‐negative/MGMT‐negative profiles, p53 overexpression was significantly associated with an unfavorable outcome. Thus, the immunohistochemical profiles of IDH1 and MGMT are of considerable significance in gliomas, and a combination of IDH1, MGMT and p53 profiles may be useful for prognostication of GBM/AA.     

Carbonic anhydrase IX is a prognostic biomarker in glioblastoma multiforme

The identification of prognostic factors in patients with glioblastoma multiforme (GBM) represents an area of increasing interest. Carbonic anhydrase IX (CA‐IX), a hypoxia marker, correlates with tumor progression in a variety of human cancers. However, the role of CA‐IX in GBM remains largely unknown. In the present study, we evaluated the prognostic role of CA‐IX in GBM patients. In total, 66 consecutive patients with GBM who received concomitant chemoradiotherapy and adjuvant chemotherapy with temozolomide were retrospectively reviewed, and all patients received temozolomide chemotherapy for at least 3 months. Kaplan–Meier curves and log‐rank tests were used for analysis of progression‐free survival (PFS) and overall survival (OS), and a multivariate Cox proportional hazard model was employed to identify factors with an independent effect on survival. The median OS was longer in patients with low levels of CA‐IX expression (18 months) compared to patients overexpressing CA‐IX (9 months) (P = 0.004). There was not a statistically significant difference in median PFS (3.5 vs. 8 months, P = 0.054) between patients with high or low levels of CA‐IX expression. In multivariate analysis, the variables that were identified as significant prognostic factors for OS were preoperative Karnofsky performance scale score (KPS) (hazard ratio (HR), 3.703; P = 0.001), CA‐IX overexpression (HR, 1.967; P = 0.019), and incomplete adjuvant temozolomide treatment (HR, 2.241; P = 0.003) and gross‐total resection (HR, 1.956; P = 0.034). Our findings indicated that CA‐IX may be a potential prognostic biomarker in the treatment of GBM.     

Methylation status of the MGMT gene promoter fails to predict the clinical outcome of glioblastoma patients treated with ACNU plus cisplatin

We analyzed the methylation status of the O6‐methylguanine‐DNA methyltransferase (MGMT) promoter using a methylation‐specific polymerase chain reaction (MSP) in glioblastoma patients treated with 1‐(4‐amino‐2‐methyl‐5‐pyrimidinyl)methyl‐3‐(2‐chloroethyl)‐3‐nitrosourea (ACNU) plus cisplatin followed by radiation therapy. Forty‐eight patients with interpretable MSP results were included in this study. The MGMT promoter was methylated in 26 patients (54.2%, methylated group) and unmethylated in 22 patients (45.8%, unmethylated group). Comparison of clinical outcomes between the two groups revealed that the methylation status of the MGMT gene promoter was not a prognostic factor for overall survival (P = 0.516) or a predictive factor for radiological response to ACNU plus cisplatin treatment (P = 0.529). The most noteworthy explanation for the result is that the synergistic antitumor effects of ACNU and cisplatin resulting from inactivation of MGMT by cisplatin in MGMT active tumors offset the drug resistance.     

幕上大脑半球高级别星形细胞瘤预后的影响因素分析

目的探讨与幕上大脑半球高级别星形细胞瘤生存预后相关的临床因素。方法回顾接受手术与术后辅助性放疗及化疗的97例高级别星形细胞瘤的临床资料,其中间变性星形细胞瘤(AA)60例,胶质母细胞瘤(GB)37例。随访其生存状况,用Kaplan-Meier单因素生存分析与COX多元回归分析作统计学处理。结果间变性星形细胞瘤和胶质母细胞瘤的无进展生存时间(PFS)分别为18和10个月,两者总生存时间(OS)分别为21和12个月;年龄“≤40岁”与“>40岁”者PFS各为18和10个月,OS各为21和13个月;治疗前“KPS≥80”与“<80”PFS各为15和10个月,OS为18和12个月;术前有抽搐症状与否的PFS各为21和11个月,OS为24和14个月;术前MR检查肿瘤强化与否的PFS各为11和18个月,OS为14和21个月。单因素分析显示,上述各因素与病人PFS和OS相关(P<0.05)。性别、肿瘤直径、肿瘤部位、肿瘤切除程度与病人PFS、OS关系无统计学意义(P>0.05)。COX回归分析显示年龄大小、肿瘤级别的高低、治疗前KPS评分、术前有无抽搐症状是影响预后的显著因素。结论年龄较小、高KPS评分、低病理级别及术前有抽搐症状被提示是高级别星形细胞瘤患者获得较长生存期的保护因素,而性别、肿瘤部位、肿瘤大小和手术切除程度对预后影响无统计学意义。肿瘤强化与预后的关系有待进一步研究证实。     

影响胶质母细胞瘤切除术后的预后因素分析

目的 探讨影响胶质母细胞瘤切除术后的预后因素.方法 对山东省三家医院1999-2004年经手术治疗且随访资料完整的205例胶质母细胞瘤进行回顾性研究,Kaplan-Meier单因素分析筛选预后相关因素,并通过Cox回归模型对筛选出的相关因素进行多因素分析.结果 患者中位生存期为12.0个月,术后6、12、18、24个月的积累生存率分别为82%、52%、27%、17%.多因素分析显示年龄、肿瘤部位、术前KPS评分、手术切除程度、术后放疗、化疗是影响预后的主要因素.结论 经多因素分析证实肿瘤的切除程度、术后放疗和化疗均能显著影响胶质母细胞瘤的预后,其中放疗是最具统计学意义的治疗方式.     

Novel biomarker,phosphorylated T‐LAK cell‐originated protein kinase (p‐TOPK) can predict outcome in primary central nervous system lymphoma

This study aimed to assess whether T‐lymphokine‐activated killer cell‐originated protein kinase (TOPK) can be a potent novel biomarker to predict the outcome in patients with primary central nervous system lymphoma (PCNSL). This study enrolled 20 patients who were histologically diagnosed as having diffuse large B‐cell type PCNSL between 2005 and 2015. Using surgical specimens, the expression of TOPK and phosphorylated TOPK (p‐TOPK) was analyzed on immunohistochemistry. Clinical features such as age, sex, Karnofsky performance status (KPS), ocular involvement, deep brain structure involvement, the number of lesions, chemotherapy and radiation therapy were also collected. Impacts of TOPK/p‐TOPK expression on their progression‐free survival (PFS) and overall survival (OS) were examined with multivariate analysis. Median PFS/OS were 24.2 and 39.0 months, respectively. On immunostaining, the mean percentage of TOPK‐positive cells was 35.5 ± 20.8%, and the mean number of p‐TOPK‐positive cells was 13.7 ± 15.7 cells/mm². The higher expression of p‐TOPK was significantly related to multiple lesions (P = 0.003). Multivariate analysis demonstrated that only the higher expression of p‐TOPK was an independent predictor to shorten both PFS (P = 0.029; hazard ratio (HR), 5.5; 95% confidential interval (CI), 1.2–25.3) and OS (P = 0.014; HR, 7.7; 95% CI, 1.5–41.3). These findings strongly suggest that p‐TOPK may be a potent biomarker to determine the outcome of patients with PCNSL and to develop novel drugs to treat PCNSL.     

Genetic pathways to glioblastomas

Glioblastomas, the most frequent and malignant human brain tumors, may develop de novo (primary glioblastoma) or by progression from low‐grade or anaplastic astrocytoma (secondary glioblastoma). These glioblastoma subtypes constitute distinct disease entities that affect patients of different ages and develop through different genetic pathways. Our recent population‐based study in the Canton of Zürich, Switzerland, shows that primary glioblastomas develop in older patients (mean age, 62 years) and typically show LOH on chromosome 10q (69%) and other genetic alterations (EGFR amplification, TP53 mutations, p16^INK4a deletion, and PTEN mutations) at frequencies of 24–34%. Secondary glioblastomas develop in younger patients (mean, 45 years) and frequently show TP53 mutations (65%) and LOH 10q (63%). Common to both primary and secondary glioblastoma is LOH on 10q, distal to the PTEN locus; a putative suppressor gene at 10q25‐qter may be responsible for the glioblastoma phenotype. Of the TP53 point mutations in secondary glioblastomas, 57% were located in hotspot codons 248 and 273, while in primary glioblastomas, mutations were more widely distributed. Furthermore, G:C→A:T mutations at CpG sites were more frequent in secondary than in primary glioblastomas (56% vs 30%). These data suggest that the TP53 mutations in these glioblastoma subtypes arise through different mechanisms. There is evidence that G:C→A:T transition mutations at CpG sites in the TP53 gene are significantly more frequent in low‐grade astrocytomas with promoter methylation of the O⁶‐methylguanine‐DNA methyltransferase (MGMT) gene than in those without methylation. This suggests that, in addition to deamination of 5‐methylcytosine (the best known mechanism of formation of G:C→A:T transitions at CpG sites), involvement of alkylating agents that produce O⁶‐methylguanine or related adducts recognized by MGMT cannot be excluded in the pathway leading to secondary glioblastomas.     

Expression of O6‐methylguanine DNA methyltransferase (MGMT) and immunohistochemical analysis of 12 pineal parenchymal tumors

Pineal parenchymal tumors (PPTs) are rare neoplasms which occupy less than 1% of primary CNS tumors. Because of their rare incidence, previous reports on PPTs are limited in number and the useful molecular markers for deciding histological grading and even selecting chemotherapy are undetermined. In this study, we conducted immunohistochemical analysis of 12 PPT specimens, especially for expression of O⁶‐methylguanine DNA methyltransferase (MGMT) to assess whether temozolomide (TMZ) could serve as a possible alternative therapy for PPTs. We analyzed 12 PPTs, consisting of three pineocytomas, six PPTs of intermediate differentiation (PPTIDs), and three pineoblastomas. Immunohistochemical analysis was performed using antibodies against MGMT, synaptophysin, neurofilament protein (NF), p53, and neuronal nuclear antigen (NeuN). Immunohistochemically, 11 out of 12 cases were positive for MGMT. The mean MIB‐1 labeling index was less than 1% in pineocytoma, 3.5% in PPTID, and 10.5% in pineoblastoma. All 12 cases were positive for synaptophysin and 11 cases, except one PPTID case, showed positive for NF. Nuclear staining of NeuN was negative in all cases although cytoplasmic staining of NeuN was observed in five cases. No case was positive for p53. Eleven out of 12 cases of PPTs demonstrated MGMT expression, suggesting chemoresistancy to TMZ treatment. This is the first report showing MGMT expression in PPTs. In addition, MIB‐1 labeling index correlated with WHO grade, although the immunoreactivity of synaptophysin, NF, NeuN and p53 did not correlate with the histological grade.     

Gliosarcomas lack BRAFV600E mutation,but a subset exhibit β‐catenin nuclear localization

Gliosarcoma (GS) is a rare subtype of glioblastoma (GBM) characterized by both glial and mesenchymal components. Unlike GBM, there are no specific prognostic markers, and optimized treatments for patients with GS do not exist. Recent reports describe BRAF^V600E mutation in malignant peripheral nerve sheath tumors, and aberrant Wnt signaling and CTNNB1 (β‐catenin gene) mutations have been described in GBM. We sought to determine whether GS tumors harbor BRAF^V600E mutations or aberrant Wnt signaling, as indicated by nuclear localization of β‐catenin, by immunohistochemical detection. Forty‐eight (48) cases of primary and secondary adult GS (including recurrent ones) were evaluated by immunohistochemical techniques for the presence of nuclear β‐catenin and the BRAF^V600E mutation. A small subset (6/46, 13%) showed nuclear localization of β‐catenin. None of the cases harbored BRAF^V600E mutations (0/48). These results are the first to describe the presence of Wnt signaling pathway abnormalities, manifested by nuclear β‐catenin, in a subset, as well as the lack of BRAF^V600E mutation in GS. We propose a potential role for Wnt pathway alterations in the pathogenesis of a subset of GS.     

胶质瘤MGMT启动子甲基化定量检测的MS-HRM方法建立及评估

目的建立甲基化特异高分辨率溶解曲线(Methylation sensitive high resolution melting curve,MS-HRM)定量检测胶质瘤MGMT基因启动子甲基化的方法,用于指导胶质瘤患者术后化疗及预后判断。方法从标本库随机选取胶质瘤组织37例,提取DNA进行甲基化修饰,应用MS-HRM方法和甲基化特异性PCR(methylation specific PCR,MSP)进行MGMT基因启动子区甲基化检测。结果 MSP方法检测显示部分甲基化标本29例(78.4%),较甲基化(13.5%)和未甲基化(8.1%)差异显著。MS-HRM发现MGMT基因启动子甲基化水平在10%~25%和25%~50%的标本分别有14例(37.8%)和17例(49.5%)。结论成功建立MS-HRM检测胶质瘤MGMT启动子甲基化的方法,该方法特异性高、灵敏度强和可重复性,有望应用于临床胶质瘤MGMT启动子甲基化高通量定量检测,以指导个体化治疗。     

胼胝体胶质瘤的预后影响因素(附60例分析)

目的 分析与胼胝体胶质瘤患者预后相关的临床因素.方法 回顾性分析第二军医大学长征医院神经外科自1995年1月至2007年12月收治、接受手术和行术后放、化疗的60例胼胝体胶质瘤患者的临床资料,随访其生存状况,分析性别、年龄、肿瘤部位、术前机能状况标准(KPS)评分、术前有无癫痫、肿瘤病理级别、肿瘤大小、影像学强化、手术切除程度对胼胝体胶质瘤患者生存预后的影响.结果 Kaplan-Meier法单因素分析显示年龄小、KPS评分高和胶质瘤病理级别低患者无进展生存时间(PFS)及总生存时间(OS)均较长,胶质瘤部位不同患者OS不同,相比差异均有统计学意义(P<0.05);Cox多元回归分析显示年龄、肿瘤病理级别为胼胝体胶质瘤患者PFS的影响因素,而年龄、病理级别、手术切除程度是OS的影响因素.结论 患者年龄较小、肿瘤病理级别低及切除程度高是胼胝体胶质瘤患者预后的保护因素,采取积极正确的治疗策略可使患者的预后得到改善.     

MGMT promoter gene methylation in pediatric glioblastoma: analysis using MS-MLPA

Purpose  

Promoter methylation of the O⁶-methylguanine-DNA-methyltransferase (MGMT) gene is widely recognized as an important predictive factor in the treatment of glioblastoma (GBM) patients with temozolomide. However, data regarding the methylation status of the MGMT promoter in pediatric GBM are yet to be elucidated.     

Radiation‐induced glioblastoma in a medulloblastoma patient: A case report with molecular features

We report a case of glioblastoma (GBM) occurring 8 years after radiation therapy for a medulloblastoma. A 15‐year‐old boy underwent surgery and radiotherapy for a medulloblastoma and 8 years later he developed a second tumor at the same site. The second lesion showed different histological and molecular features, was diagnosed as a glioblastoma and fulfilled the criteria of radiation‐induced neoplasm. Mutational analysis of the p53 gene showed a C to G transition at codon 176 in tumor DNA. LOH was detected at 17p and 19q. The tumor also showed O6‐methylguanine‐DNA methyl‐transferase (MGMT) promoter methylation and no amplification of EGF receptor. In conclusion, the radiation‐induced MGMT hyper‐methylation and p53 mutations may have a role in the development of a subgroup of radio‐induced glioma (RIG), suggesting that these molecular alterations directly cooperate in the genesis of the post‐irradiation GBM. Moreover RIGs seem to be a heterogeneous group of tumors that may resemble either primary or secondary GBM.     

Amplification of the PDGFRA,KIT and KDR genes in glioblastoma: a population‐based study

The aim of this study was to establish the frequency of amplification of tyrosine kinase receptor genes PDGFRA, KIT and KDR (VEGFR2) at 4q12 in glioblastomas at a population level, and to assess whether such alterations have any clinical impact. Screening of 390 glioblastomas from a population‐based study by differential PCR revealed amplification of the PDGFRA, KIT and KDR genes in 33 (8.5%), 17 (4.4%) and 13 (3.3%) glioblastomas, respectively. None of these alterations was prognostic for overall survival. Patients with glioblastoma showing KIT amplification were significantly younger than those with glioblastoma showing no amplification (51.7 ± 21.7 years vs. 59.3 ± 13.1 years; P = 0.0231). Twelve glioblastomas showed concurrent amplification of the PDGFRA, KIT and KDR genes, whereas 18 glioblastomas showed PDGFRA amplification only. A significant inverse association was observed between KIT amplification and EGFR amplification (P = 0.0260), whereas a borderline positive association was found between KIT amplification and TP53 mutation (P = 0.0579). No significant difference was observed in the frequency of amplification of these genes in primary and secondary glioblastomas or in glioblastomas with and without IDH1 mutations, suggesting that amplification of PDGFRA, KIT and KDR may be implicated in the pathogenesis of a small fraction of both subtypes of glioblastoma.