中枢神经系统肿瘤CDKN2基因的丢失

目的探讨中枢神经系统肿瘤CDKN2基因的丢失情况。方法采用聚合酶链反应(PCR)对81例脑、脊髓肿瘤手术标本中CDKN2基因的丢失情况进行了检测。结果胶质瘤CDKN2基因丢失率为60％,且高级别胶质瘤基因丢失率显著高于低级别肿瘤;脑膜瘤、神经鞘瘤、垂体腺瘤及转移瘤亦存在不同程度CDKN2基因的丢失。结论CDKN2基因的丢失与中枢神经系统肿瘤的发生、发展有一定的关系。     

CDKN2A deletion in pediatric versus adult glioblastomas and predictive value of p16 immunohistochemistry

Cell cycle regulator genes are major target of mutation in many human malignancies including glioblastomas (GBMs). CDKN2A is one such tumor suppressor gene which encodes p16INK4a protein and serves as an inhibitor of cell cycle progression. Very few studies are available regarding the association of CDKN2A deletion with p16 protein expression in GBMs. There is limited data on the frequency of CDKN2A deletion in different age groups. The aim of the present study was to analyze the frequency of CDKN2A gene deletions in GBM and correlate CDKN2A deletional status with (i) age of the patient (ii) p16 protein expression and (iii) other genetic alterations, namely EGFR amplification and TP53 mutation. A combined retrospective and prospective study was conducted. Sixty seven cases were included. The patients were grouped into pediatric (≤18 years), young adults (19‐40 years) and older adults (>40 years). CDKN2A and EGFR status were assessed by Fluorescence in situ Hybridization.TP53 mutation was analyzed by PCR based method. p16 expression was assessed using immunohistochemistry. CDKN2A deletion was noted in 40.3% cases of GBM with majority being homozygous deletion (74%). It was commoner in primary GBMs (65.8%) and cases with EGFR amplification (50%). A variable frequency of CDKN2A was observed in older adults (42.3%), young adults (44%), and pediatric patients (31.25%). The difference however was not statistically significant. There was statistically significant association between CDKN2A deletion and p16 immunonegativity with a high negative predictive value of immunohistochemistry.     

Comparative genomic hybridization analysis of genomic alterations in benign,atypical and anaplastic meningiomas

BACKGROUND: Meningiomas are common tumors of the central nervous system. Although most are benign tumors, approximately 10% show a histologic progression to a higher malignancy grade similar to atypical (GII) and anaplastic (GIII) meningiomas. Monosomy 22q12 is the most frequent genetic alteration detected in these tumors, but failure of detection of 22q mutations in about 40% of tumors which are indistinguishable from meningiomas with 22q deletions with respect to clinical and histopathologic features, makes it apparent that an alternative mechanism is responsible for the initiation of meningioma. Moreover, little is known about genetic alterations during malignant progression of meningioma. PURPOSE: In order to determine the genetic pathways underlying the development of meningioma, 15 benign (WHO grade I), 7 atypical (WHO grade II) and 3 anaplastic (WHO grade III), sporadic meningiomas were screened by Comparative Genomic Hybridization (CGH). RESULTS: Statistical analysis revealed a significant correlation between the number of chromosomal imbalances and the tumor grade; the numbers of total alterations detected per tumor were 2.20 (2.24 for GI, 10.00 (1.17 for GII and 14.66 (1.15 for GIII. The most frequent abnormality seen in benign tumors was loss on 22q (47%). The second alteration was 1p deletion (33%) and this abnormality was also the common aberration in three tumors without CGH detected 22q deletion. In GII, aberrations most commonly identified were losses on 1p (6/7 cases), 22q (5/7 cases), 10q (4/7 cases), 14q and 18q (3/7 cases) as well as gains on 15q and 17q (3/7 cases). In GIII, genomic loss on 1p was the most commonly observed abnormality (3/3). Losses on 9p, 10q, 14q, 15q, 18q and 22q as well as gains on 12q, 15q and 18p were the other genomic alterations detected by CGH. Combined 1p/14q deletions were encountered in 2/15 benign, 3/7 atypical and 2/3 anaplastic meningiomas. By CGH, DNA sequences on 17q21-qter were seen to be amplified in 1/7 GII and 2/3 GIII, whereas highly amplified DNA sequences on 12q13-qter, 20q and 22q11-q12 were seen in one GII, two GII/one GIII, and one GIII, respectively. CONCLUSION: It was concluded that chromosomal deletion from 1p could play a major role in the initiation and progression of meningiomas and that 1p/14q deletions could be a primary focus of further detailed assessment of tumour genesis.     

Molecular heterogeneity of oligodendrogliomas suggests alternative pathways in tumor progression

OBJECTIVE: To identify different genetic molecular profiles in oligodendrogliomas and to evaluate their prognostic significance. METHODS: The main genetic alterations reported in glial tumors were investigated in 26 oligodendrogliomas (10 World Health Organization grade II and 16 World Health Organization grade III). Correlation between identified molecular changes and pathologic grade or clinical course was subsequently analyzed using univariate and multivariate statistical analyses. RESULTS: Loss of heterozygosity (LOH) on chromosome 1p, 19q, and 10; P16/CDKN2A homozygous deletion; EGFR (epidermal growth factor receptor) amplification; and TP53 and PTEN mutations were observed in 14 (54%), 15 (58%), 9 (35%), 7 (27%), 5 (19%), 1 (4%), and 0 cases. LOH 1p and 19q were tightly associated (p < 0.0001). A mutual exclusion was found between LOH 1p/19q and EGFR amplification (p = 0.01), P16/CDKN2A deletions (p = 0.001), or LOH on 10q (p = 0.03), suggesting the existence of distinct genetic subsets in oligodendrogliomas. On univariate analysis, age <50 years (p = 0.002) and LOH 1p (p = 0.01) were associated with a longer progression-free survival (PFS) whereas LOH 10q (p = 0.03) and EGFR amplification (p = 0.007) were associated with a worse PFS. In multivariate analyses, age <50 years (p = 0.001) and LOH 1p (p = 0.006) remained independent predictive factors for PFS. CONCLUSION: These results provide evidence for two alternative molecular pathways of progression in oligodendrogliomas. The first one is associated with loss of 1p and 19q and the second one with P16/CDKN2A deletion, 10q loss, and EGFR amplification. The findings confirm the value of loss of 1p as predictor of longer progression-free survival; in addition, the study demonstrates the unfavorable impact of 10q loss and EGFR amplification on the prognosis.     

颅内血管外皮细胞瘤鉴别诊断分析

目的 颅内血管外皮细胞瘤(HPC),较为罕见,局部复发和远处转移率较高.临床上常需与血管瘤型脑膜瘤、孤立性纤维瘤(SFT)相鉴别,本组采用免疫组化方法来鉴别这三种肿瘤.方法 采用免疫组化方法对1993至2010年收治的11例HPC患者进行回顾性分析,并基于染色结果重新评估临床诊断.结果 根据免疫组化结果,在11例中有1例因CD34和Bcl-2染色阳性被重新诊断为SFT,有1例因EMA强阳性被诊断为血管瘤型脑膜瘤.随访期内,有3例HPC患者复发,1例HPC患者死亡.结论 考虑到SFT和血管瘤型脑膜瘤较HPC预后良好,初步诊断时应严格进行免疫组化检查,对这三种肿瘤的鉴别诊断起着重要作用.

Abstract：

Objective Intracranial hemangiopericytomas (HPCs) were rare neoplasms with relatively high rates of recurrence and extracranial metastasis. The differential diagnoses from angiomatous meningiomas and solitary fibrous tumors(SFTs) are very important. This study is aimed to reveal differences in the specific immunohistochemical features of HPCs, angiomatous meningiomas and SFTs by newly developed immunohistochemical methods. Method Eleven sequential cases of HPC diagnosed in Renji hospital between 1993 and 2010 were retrospectively analyzed by immunohistochemical staining for CD34,Bcl -2 and epithelial membrane antigen ( EMA ). The cases were then re - evaluated and newly diagnosed based on the results of the immunohistochemical stainings. Results One of the eleven cases was newly diagnosed as SFTs and nine were reconfirmed as HPCs, based on the immunohistochemical studies for CD34,Bcl - 2 and EMA staining. One case was newly diagnosed as angiomatous meningioma on the basis of a strong EMA staining positivity. Three HPCs recurred and one HPC patient died of the tumor. Conclusions Given that SFTs and angiomatous meningioma have a considerably better prognosis than HPCs, it is important to carry out meticulous immunohistochemical examinations for primary diagnosis.     

Clinicopathological and molecular characteristics of pediatric meningiomas

Molecular and clinical characteristics of pediatric meningiomas are poorly defined. Therefore, we analyzed clinical, morphological and molecular profiles of pediatric meningiomas. Forty pediatric meningiomas from January 2002 to June 2015 were studied. 1p36, 14q32 and 22q‐deletion were assessed by fluorescent in situ hybridization and mutations of most relevant exons of AKT, SMO, KLF4, TRAF and pTERT using sequencing. Expression of GAB1, stathmin, progesterone receptor (PR), p53 along with MIB‐1 LI was examined using immunohistochemistry. There were 36 sporadic and four NF2 associated meningiomas. Among sporadic meningiomas, the majority (72.2%) of cases harbored 22q‐deletion. Difference in frequency of combined 1p/14q deletion in Grade‐I versus Grade‐II/III tumors was not significant (13.7% vs 28.5%, P = 0.57). PR immunoreactivity was seen in 65.5% of Grade‐I and 14.2% of Grade‐II/III tumors (P = 0.03). The majority (97.2%) of meningiomas were immunonegative for p53. Stathmin and GAB co‐expression was observed in 58.3% of cases. Notably, AKT, SMO, KLF4, TRAF7 (exon 17) and pTERT mutations were seen in none of the cases analyzed. 1p/14q codeletion was frequent in skull base as compared to non‐skull base meningiomas (23% vs 11.1%, P = 0.37). All NF2 meningiomas harbored 22q‐deletion and showed GAB and stathmin co‐expression while none showed 1p/14q loss. Pediatric meningiomas share certain phenotypic and cytogenetic characteristics with adult counterparts, but GAB and stathmin co‐expression in the majority of cases and non‐significant difference in frequency of 1p/14q co‐deletion between low‐ and high‐grade meningiomas indicate an inherently aggressive nature. Characteristic AKT/SMO, KLF4/TRAF7 and pTERT genetic alterations seen in adults are distinctly absent in pediatric meningiomas.     

Radiation-induced meningioma: a distinct molecular genetic pattern?

Radiation-induced meningiomas arise after low-dose irradiation treatment of certain medical conditions and are recognized as clinically separate from sporadic meningioma. These tumors are often aggressive or malignant, they are likely to be multiple, and they have a high recurrence rate following treatment compared with sporadic meningiomas. To understand the molecular mechanism by which radiation-induced meningioma (RIM) arise, we compared genetic changes in 7 RIM and 8 sporadic meningioma (SM) samples. The presence of mutations in the 17 exons of the neurofibromatosis type 2 (NF2) gene, which has been shown to be inactivated in sporadic meningiomas, was analyzed in RIM and SM using single-strand conformation polymorphism (SSCP) and DNA sequencing. In contrast to SM, which showed NF2 mutations in 50% of specimens, no mutations were found in RIM. In addition, Western blot analysis of schwannomin/merlin protein, the NF2 gene product, demonstrated protein levels comparable to normal brain in 4/4 RIM tumor samples analyzed. Loss of heterozygosity (LOH) of genomic regions, which were reported for SM, was also analyzed in all cases of RIM using 22 polymorphic DNA markers. Allele losses were found on chromosomes 1p (4/7), 9p (2/7), 19q (2/7), 22q (2/7), and 18q (1/7). From these observations we conclude that unlike sporadic meningiomas, NF2 gene inactivation and chromosome 22q deletions are far less frequent in RIM, and their role in meningioma development following low dose irradiation is less significant. Other chromosomal lesions, especially loss of 1p, possibly induced by irradiation, may be more important in the development of these tumors.     

Tumour necrosis and microvascular proliferation are associated with 9p deletion and CDKN2A alterations in 1p/19q-deleted oligodendrogliomas

A subset of oligodendrogliomas and oligoastrocytomas has been associated with 1p/19q deletion. Subsequently, this genetic alteration was linked to chemosensitivity and classic histology of oligodendrogliomas. Tumoural progression includes deletions of 9p, 10q and alterations of CDKN2A. However, these (epi)genetic changes have not been associated with specific histological features. In a series of 45 gliomas including oligodendrogliomas, oligoastrocytomas and astrocytomas, deletions of chromosomal regions implied in these tumours (1p, 9p, 10, 17p13, 19q and 22) were looked for by microsatellite analysis. Tumours that were deleted for 1p and 19q were selected. Subsequently, presence of deletions in the other studied regions, (epi)genetic changes in p14ARF, CDKN2A and CDKN2B, as well as histological features, were associated to these tumours. 1p/19q deletion was observed in 22 tumours. Twenty-one of them presented regions of classic histology of oligodendroglioma. A deletion of 9p was found in eight of them, always in association with tumour necrosis and/or microvascular proliferation. In addition, (epi)genetic alterations of CDKN2A were observed in 71% of these tumours. Presence of regions of classic histology of oligodendroglioma in a tumour sample is predictive of 1p/19q deletions. Necrosis and/or microvascular proliferation are signs of an additional 9p deletion. Finally, as CDKN2A (epi)genetic alterations were found in 71% of the 1p/19q/9p-deleted oligodendrogliomas, CDKN2A may have a role in oligodendroglioma-associated microvascular proliferation.     

Molecular genetic alterations in glioblastomas with oligodendroglial component

Glioblastoma multiforme is the most malignant astrocytic glioma and usually resistant to chemotherapy. A small fraction of glioblastomas may contain areas with histological features of oligodendroglial differentiation. To determine the molecular genetic alterations in such "glioblastomas with oligodendroglial component", we investigated 13 of these tumors for genetic alterations and/or expression of the TP53, CDKN2A, PTEN, and EGFR genes. In addition, we performed microsatellite analyses for loss of heterozygosity (LOH) on chromosome arms 1p, 19q and 10q. None of tumors showed evidence for LOH on 10q. LOH on 1p was detected in 3 tumors, 1 of which additionally showed LOH on 19q. The 3 tumors with LOH on 1p showed neither TP53 mutations nor nuclear p53 accumulation. In contrast, 9 of 10 tumors without demonstrated losses on 1p showed nuclear p53 accumulation. TP53 mutations were identified in 3 of these cases. Further aberrations detected were epidermal growth factor receptor (EGFR) overexpression (3 of 13 tumors), homozygous CDKN2A deletion (2 of 11 tumors), and PTEN mutation (1 of 13 tumors). Taken together, our results indicate that "glioblastomas with oligodendroglial component" carry heterogeneous genetic alterations. LOH on 10q, PTEN mutation, and homozygous CDKN2A deletion appear to be less common in these tumors as compared to ordinary glioblastomas. Furthermore, a subset of these tumors demonstrates LOH on 1p, i.e., an alteration that has recently been linked to chemosensitivity and good prognosis in anaplastic oligodendrogliomas.     

EZH2 expression in gliomas: Correlation with CDKN2A gene deletion/ p16 loss and MIB‐1 proliferation index

Enhancer of zeste homolog 2 (EZH2) mediated down‐regulation of CDKN2A/p16 has been observed in cell lines as well as in a few carcinomas. However, there is no study correlating EZH2 expression with CDKN2A/p16 status in gliomas. Hence, the present study was conducted to evaluate EZH2 expression in astrocytic and oligodendroglial tumors and correlate with CDKN2A/p16 status as well as MIB‐1 labeling index (LI). Gliomas of all grades (n = 118) were studied using immunohistochemistry to assess EZH2, p16 and MIB‐1 LI and fluorescence in situ hybrization to evaluate CDKN2A gene status. EZH2 expression and CDKN2A homozygous deletion (HD) were both significantly more frequent in high‐grade gliomas (HGG). Further, strong EZH2 expression (LI ≥ 25%) was significantly more common in HGGs without CDKN2A HD (48.7%; 19/39) as compared to cases with deletion (15.8%; 3/19). Loss of p16 expression was noted in 100% and 51.3% of CDKN2A deleted and non‐deleted tumors, respectively. Notably, 80% (16/20) of the CDKN2A non‐deleted HGGs with p16 loss had strong EZH2 expression, in contrast to only 15.8% (3/19) in the deleted group. Loss of p16 expression significantly correlated with MIB‐1 LI, irrespective of EZH2 status. Thus, this study shows that EZH2 expression correlates with tumor grade in both astrocytic and oligodendroglial tumors and hence can be used as a diagnostic marker to differentiate between low and HGGs. Further, this is the first report demonstrating an inverse correlation of strong EZH2 expression with CDKN2A HD in HGGs. Loss of p16 protein expression is mostly attributable to CDKN2A HD and correlates significantly with MIB‐1 LI. Notably, our study for the first time suggests a possible epigenetic mechanism of p16 loss in CDKN2A non‐deleted HGGs mediated by strong EZH2 expression. A hypothetical model for control of proliferative activity in low versus HGGs is therefore proposed.     

Accumulation of wild-type p53 in meningiomas

The p53 tumour suppressor gene contributes to the regulation of DNA repair and the initiation of apoptosis. Mutations of this gene and nuclear accumulation of its protein product are features of a large variety of turnours. A histological spectrum of meningiomas, including anaplastic examples and a meningosarcoma. was analysed for accumulation of the p53 protein and mutations in exons 4–9 of the p53 gene. No mutations were found, but 9134 (26%) tumours showed accumulation of the p53 protein. The p53-positive meningiomas came from across the histological spectrum and were not restricted to the anaplastic group. The accumulation of wild-type p53 in a proportion of meningiomas may reflect this gene's role in DNA repair, or its enhanced stability when bound to cellular proteins such as the mdm-2 gene product.     

Systemic metastasis in glioblastoma may represent the emergence of neoplastic subclones

Glioblastomas only rarely metastasize to sites outside the central nervous system, for reasons that are poorly understood. We report the clinicopathological and molecular genetic findings in 6 patients with metastatic glioblastoma. Four patients were under the age of 32 and all but 1 patient died within 2 yr of diagnosis. The number of metastases ranged from 1 to 3. At the time of death, 3 patients had apparent tumor control at their primary site. We evaluated DNA from both primary and metastatic glioblastomas for genetic alterations commonly found in glioblastomas: TP53 mutations, CDKN2A/p16 deletions, EGFR amplification, and allelic loss of chromosomes 1p, 10q and 19q. Four of 6 cases had TP53 mutations and only single cases had EGFR amplification, CDKN2A/p16 deletions, or allelic loss of 1p, 10q and 19q; 2 cases had no detectable genetic alterations. In 2 cases, the primary and metastatic tumors had identical genotypes. Remarkably, however, 2 cases had different TP53 alterations in the primary and metastatic lesions, or among the metastatic tumors, which suggests that some metastatic deposits may represent emergence of subclones that were not necessarily dominant in the primary tumor. The present observations and a review of the recent literature demonstrate that metastatic glioblastomas tend to occur in younger adults who do not follow long clinical courses, and may be characterized by TP53 mutations and differential clonal selection.     

Chromosome 1p and 14q FISH analysis in clinicopathologic subsets of meningioma: diagnostic and prognostic implications

The second most frequently reported genetic abnormalities in meningiomas after 22q loss are deletions of 1p and 14q. To assess the potential diagnostic and prognostic utility of these chromosomal alterations, we studied 180 well-characterized meningiomas using dual-color fluorescence in situ hybridization (FISH) with DNA probes localized to 1p32, 1p36, 14q13, and 14q32. Our cohort consisted of 77 benign (grade I), 74 atypical (grade II), and 29 anaplastic (grade III) meningiomas. Benign and atypical meningiomas were further stratified into subsets of recurring (despite gross total resection) vs non-recurring (at least 10 yr of follow-up) and mitotically active vs brain invasive subsets, respectively. Losses of 1p and 14q losses were identified in 23% and 31% of benign, 56% and 57% of atypical, and 75% and 67% of anaplastic meningiomas, respectively (p < 0.001 for 1p; p = 0.004 for 14q). Combined 1p/14q deletions were encountered in 7% benign. 39% atypical, and 63% anaplastic meningiomas (p < 0.001). Benign non-recurring meningiomas were less likely to harbor 14q deletions than recurring examples (17% vs 50%, p = 0.013). There was a trend for anaplastic meningiomas with 14q deletions and atypical meningiomas with combined 1p/14q deletions to have poorer overall survivals, though neither reached statistical significance. We conclude that 1p and 14q deletions are highly associated with increasing histologic grade and play an important role in meningioma tumor progression. Furthermore, 14q FISH analysis may aid in assessing recurrence risk in histologically benign meningiomas.     

Homozygous deletion of the survival motor neuron 2 gene is a prognostic factor in sporadic ALS

BACKGROUND: Spinal muscular atrophy (SMA) results from mutations of the survival motor neuron (SMN) gene on chromosome 5. The SMN gene exists in two highly homologous copies, telomeric (SMN1) and centromeric (SMN2). SMA is caused by mutations in SMN1 but not SMN2. The clinical phenotype of SMA appears to be related to the expression of SMN2. Patients suffering from the milder forms of SMA carry more copies of the SMN2 gene compared with patients with more severe SMA. It is suggested that the SMN2 gene is translated into an at least partially functional protein that protects against loss of motor neurons. OBJECTIVE: To investigate whether genetic mechanisms implicated in motor neuron death in SMA have a role in ALS. METHODS: The presence of deletions of exons 7 and 8 of SMN1 and SMN2 was determined in 110 patients with sporadic ALS and compared with 100 unaffected controls. RESULTS: The presence of a homozygous SMN2 deletion was overrepresented in patients with ALS compared with controls (16% versus 4%; OR, 4.4; 95% CI, 1.4 to 13.5). Patients with a homozygous SMN2 deletion had a shorter median time of survival (p < 0.009). Furthermore, multivariate regression analysis showed that the presence of an SMN2 deletion was independently associated with survival time (p < 0.02). No homozygous deletions in SMN1 were found. Carrier status of SMA appeared to be equally present in patients and controls (1 in 20). CONCLUSION: These results indicate that, similar to SMA, the SMN2 gene can act as a prognostic factor and may therefore be a phenotypic modifier in sporadic ALS. Increasing the expression of the SMN2 gene may provide a strategy for treatment of motor neuron disease.     

A reevaluation of the primary diagnosis of hemangiopericytoma and the clinical importance of differential diagnosis from solitary fibrous tumor of the central nervous system

Objectives

Hemangiopericytomas (HPCs) are rare neoplasms with relatively high rates of recurrence and extracranial metastasis. Though the differential diagnoses from angiomatous meningiomas and from solitary fibrous tumors (SFTs) are both important, the latter diagnosis is somewhat more important in light of the benign prognosis of SFTs and the difficulties in distinguishing SFTs from HPCs. Newly developed immunohistochemical methods reveal differences in the specific immunohistochemical features of HPCs and SFTs. To elucidate whether SFTs have been misdiagnosed as HPCs in the past, our group used recent immunohistochemical methods to re-evaluate tissues that had been originally diagnosed as HPCs. We also compared the clinical features of these cases.

Patients and methods

Thirteen sequential cases of HPC diagnosed in Kanazawa University Hospital and Kumamoto University Hospital between 1970 and 2006 were retrospectively analyzed by immunohistochemical staining for CD34, Bcl-2, epithelial membrane antigen (EMA), vimentin, and S100 protein, and by measurement of the MIB-1 labeling index (LI). The cases were then re-evaluated and newly diagnosed based on the results of the immunohistochemical stainings. The clinical course of each case was also evaluated.

Results

Four of the 13 cases were newly diagnosed as SFTs and eight were reconfirmed as HPCs, based on the immunohistochemical studies for CD34, Bcl-2, and reticulin staining. One case was newly diagnosed as meningioma on the basis of a strong EMA positivity. The MIB-1 LI was less than 1% in 12 of the cases. In two cases, one case of HPC and the other of meningioma, the MIB-1 LI was relatively high, 8% and 4% respectively. All eight of the HPCs recurred, and 5 of the HPC patients died of the disease. Only one case of the SFTs recurred.

Conclusion

Our study suggests that a relatively high percentage of the tumors diagnosed as HPCs in the past may have in fact been intracranial SFTs. Immunohistochemical examinations of CD34, Bcl-2, and reticulin stains are keys for the differential diagnosis. Given that SFTs have a considerably better prognosis than HPCs, it is important to carry out meticulous immunohistochemical examinations for the primary diagnosis.     

p53 and PTEN gene mutations in gemistocytic astrocytomas

The gemistocytic astrocytoma is a histological variant of diffuse astrocytomas and is characterised by the presence of large, GFAP-expressing neoplastic astrocytes (gemistocytes) and a tendency towards rapid progression to glioblastoma. In this study, we analyzed 28 gemistocytic astrocytomas (mean fraction of gemistocytes, 35.0 ± 9.9%) for mutations in the p53 and PTEN (MMAC1) tumour suppressor genes. Single strand conformation polymorphism (SSCP), followed by direct DNA sequencing of p53 exons 5–8, revealed a mutation in 23 of 28 (82%) cases. Regional analysis of four tumours revealed identical p53 mutations in gemistocytic and fibrillary tumour areas. In contrast, none of 15 gemistocytic astrocytomas (WHO Grade II) and only two of 11 (18%) anaplastic gemistocytic astrocytomas (WHO Grade III) contained a PTEN mutation. Of these, one was a 1 bp deletion in codon 345 and the other a 1 bp insertion in intron 4. Differential PCR did not reveal homozygous PTEN deletion in any of the tumours analysed. These results indicate that p53 mutations are a genetic hallmark of gemistocytic astrocytomas, whilst PTEN mutations are absent in low-grade and rare in anaplastic gemistocytic astrocytomas. Received: 16 February 1998 / Accepted: 2 March 1998     

Spastin mutation screening in Chinese patients with pure hereditary spastic paraplegia

BackgroundHereditary spastic paraplegia (HSP) is a clinically and genetically heterogeneous group of neurodegenerative diseases. Mutations in the spastin (SPAST) gene are the most common cause of pure HSP. However, few data are available regarding the clinical and genetic spectrum of HSP among Chinese patients.MethodsClinical data were collected at diagnosis and follow-up of 42 Chinese patients with pure HSP. All seventeen exons of the SPAST gene were directly sequenced. Additionally, we used a multiplex ligation dependent probe amplification (MLPA) assay targeting the SPAST gene to evaluate large exon deletion or insertion mutations in patients without SPAST point mutations.ResultsThe age of disease onset of our patients was 19.6 ± 14.4 years. Six novel variations were found, including three missense mutations (p. L363P, p. D441V, and p. S595R), one insertion (c.1511dupT (p. Y505Ifs*7)), and two larger deletions (exons 5–17 and exons 10–17). Four previously reported mutations, including p. S399L, c.1215_c.1219delTATAA (p. N405Kfs*36), exon 1 deletion, and exon 16 deletion, were detected. The SPAST mutation rate was 40% (4/10) in Chinese familial patients and 33.33% (7/21) in Chinese sporadic pure HSP patients. The frequency of large deletions was high in both AD-HSP (20%, 2/10) and sporadic HSP (14.28%, 3/21).ConclusionSPAST mutations are common in Chinese patients with pure HSP. Large exon deletions are an important cause of AD-HSP and sporadic pure HSP in Chinese patients. Large fragment tests should be performed to explore large SPAST mutations in familial and sporadic HSP patients without SPAST point mutations.