Cathepsins B and L and their inhibitors stefin B and cystatin C as markers for malignant progression of benign meningiomas

Meningiomas are, in general, slowly growing benign tumors attached to the dura mater and composed of neoplastic meningothelial (arachnoidal) cells. They have a wide range of histopathological appearances and are classified, according to the aggressiveness of their growth and the risk of recurrence, as WHO grade I (benign) meningiomas, WHO grade II (atypical) meningiomas and WHO grade III anaplastic (malignant) meningiomas. As invasion of normal tissue may occur in all grades, independent biological markers are needed to identify the more aggressive and recurrent meningiomas. The lysosomal cysteine proteinases, cathepsins B and L, have been associated with tumor invasiveness and the aim of this study was therefore to evaluate them, together with their endogenous inhibitors stefin B and cystatin C, as potential markers for the aggressiveness of meningiomas. The expression of cathepsins B and L and their inhibitors stefin B and cystatin C in 21 benign (grade I) and 9 atypical (grade II) meningiomas has been compared by immunohistochemical staining, QRT-PCR and Northern blot analysis. The protein levels of cathepsins B (p=0.050) and L (p=0.019) were found to be significantly higher in atypical than in benign meningiomas. In contrast, their mRNA levels did not differ, indicating that the synthesis of cathepsins was accelerated at the translational level. Protein and mRNA levels of stefin B (p= 0.007), but not cystatin C, were significantly lower in atypical compared with benign meningiomas. The expression of cathepsins and inhibitors was not different between central and peripheral meningioma tissue or between histological subtypes of meningiomas, with the exception of cathepsin L, the level of which was significantly lower in transitional meningiomas. We conclude that higher protein levels of cathepsins B and L and lower mRNA levels of stefin B are potential diagnostic markers for invasive and aggressive behavior of meningiomas. The diagnostic and prognostic value for relapse of meningioma needs to be confirmed in a larger population of patients.     

Analysis of Gene Expression Profiling in Meningioma: Deregulated Signaling Pathways Associated with Meningioma and EGFL6 Overexpression in Benign Meningioma Tissue and Serum

Molecular mechanisms underlying the pathogenesis of meningioma are not fully elucidated. In this study, we established differential gene expression profiles between meningiomas and brain arachnoidal tissue by using Affymetrix GeneChip Human U133 Plus 2.0 Array. KEGG pathway analysis demonstrated that PI3K/Akt and TGFβ signaling pathways were up-regulated in fibroblastic meningioma, and focal adhesion and ECM-receptor interaction pathways were activated in anaplastic meningioma. EGFL6 was one of the most up-regulated genes in fibroblastic meningioma by microarray analysis. Quantitative real-time PCR demonstrated that benign meningiomas had significantly higher levels of EGFL6 mRNA than brain arachnoidal tissue and atypical and anaplastic meningiomas (P<0.001). EGFL6 gene was also highly expressed in ovarian cancer, but expressed lowly in other investigated tumors. ELISA analysis showed that patients with benign meningiomas and ovarian cancers had the highest serum levels of EGFL6 (mean concentration: 672 pg/ml for benign meningiomas, and 616 pg/ml for ovarian cancers). Healthy people and patients with other tumors, however, had low levels of serum EGFL6. In conclusion, we proposed that activation of PI3K/Akt and integrin-mediated signaling pathways was involved in the pathogenesis of benign and anaplastic meningiomas, respectively. We also presented evidence that EGFL6 was overexpressed in benign meningioma tissues and serum.     

Benign and atypical meningioma metabolic signatures by high-resolution magic-angle spinning molecular profiling

Meningiomas are neoplasms that arise from the leptomeningeal covering of the brain and spinal cord, accounting for 15%-20% of CNS tumors. The WHO classifies meningiomas into three histological grades: benign, atypical, and anaplasic in accordance with the clinical prognosis. Atypical and anaplasic meningiomas tend to recur. Sometimes, meningiomas with histological diagnosis of benign meningioma show clinical characteristics of atypical meningioma. In this context, high-resolution magic-angle spinning (HR-MAS) spectroscopy of intact tissue from brain tumor biopsies has shown great potential as a support diagnostic tool. In this work, we show differences between benign and atypical meningiomas in HR-MAS molecular profiles of meningioma biopsies. Metabolic differences between meningioma grades include changes in the levels of glutathione. Glutathione role in cancer is still unclear, as it may act both as protective and pathogenic factor. Glutamine and glutamate, which are related to glutathione metabolism and have been associated with tumor recurrence, are also increased in atypical meningiomas. Other metabolites associated with tumor malignancy that show statistically significant differences between benign and atypical meningiomas include phosphocholine and phosphoethanolamine. Overall, this work suggests that the additional information obtained by NMR metabolomics applied to biopsies of human meningiomas may be useful for assessing tumor grade and determining optimum treatment strategies.     

Transglutaminase 2 Expression Is Increased as a Function of Malignancy Grade and Negatively Regulates Cell Growth in Meningioma

Most meningiomas are benign, but some clinical-aggressive tumors exhibit brain invasion and cannot be resected without significant complications. To identify molecular markers for these clinically-aggressive meningiomas, we performed microarray analyses on 24 primary cultures from 21 meningiomas and 3 arachnoid membranes. Using this approach, increased transglutaminase 2 (TGM2) expression was observed, which was subsequently validated in an independent set of 82 meningiomas by immunohistochemistry. Importantly, the TGM2 expression level was associated with increasing WHO malignancy grade as well as meningioma recurrence. Inhibition of TGM2 function by siRNA or cystamine induced meningioma cell death, which was associated with reduced AKT phosphorylation and caspase-3 activation. Collectively, these findings suggest that TGM2 expression increases as a function of malignancy grade and tumor recurrence and that inhibition of TGM2 reduces meningioma cell growth.     

Molecular Biological Determinations of Meningioma Progression and Recurrence

Meningiomas are tumors that arise from the coverings of the brain or spinal cord. 5% of the cases turn into malignant forms with aggressive clinical behavior and increased risk of tumor recurrence. One hundred and five patients with meningiomas were operated by open surgery. To investigate predictors of meningioma recurrence in total 124 samples of 105 patients were investigated by iFISH. Dual-probe hybridization was performed to access chromosomal alterations of chromosomes 1p-, 9p- and 22q. Additionally, methylation of TIMP3 and p16 was analyzed with MS-PCR. Of the 105 investigated tumors 59.1% (62/105) were WHO grade I, 33.3% (35/105) were WHO grade II and 7.7% (8/105) were anaplastic meningiomas (grade III), respectively. The histopathological data correlates with the recurrence rate of the investigated meningiomas. Hypermethylation of TIMP3 was detected in 13.3% of all meningiomas: 10.9% in WHO grade I meningiomas, 25.0% in grade II and 14.3% in grade III meningiomas, respectively. No correlation of TIMP3 hypermethylation with tumor recurrence or WHO grade (p = 0.2) was observed. Interestingly, deletion of 1p36 emerged as a significant predictor of shorter overall survival (log rank test, p<0.001), whereas TIMP3 promoter methylation had no significant effect on overall survival (log rank test, p = 0.799). The results of the current study support the finding that the deletion of chromosome 1p is an independent marker of meningioma recurrence and progression (p = 0.0097). Therefore the measurement of genetic aberrations in meningiomas allows in a combined histological approach a more precise assessment of the prognosis of meningiomas than histopathology alone.     

Functional genomics approaches for the study of sporadic adrenal tumor pathogenesis: clinical implications

Although sporadic adrenal tumors are frequently encountered in the general population their pathogenesis is not well elucidated. The advent of functional genomics/bioinformatics tools enabling large scale comprehensive genome expression profiling should contribute to significant progress in this field. Some studies have already been published describing gene expression profiles of benign and malignant adrenocortical tumors and phaeochromocytomas. Several genes coding for growth factors and their receptors, enzymes involved in steroid hormone biosynthesis, genes related to the regulation of cell cycle, cell proliferation, adhesion and intracellular metabolism have been found to be up- or downregulated in various tumors. Some alterations in gene expression appear so specific for certain tumor types that their application in diagnosis, determination of prognosis and the choice of therapy can be envisaged. In this short review, the authors will present a synopsis of these recent findings that seem to open new perspectives in adrenal tumor pathogenesis, with emphasis on changes in steroidogenic enzyme expression profiles and highlighting possible clinical implications.     

Incidence,mortality and outcome of meningiomas: A population-based study from Germany

BackgroundMeningiomas are mostly benign tumors that originate from the coverings of the brain and spinal cord. Compared to malignant glial tumors, meningiomas are relatively understudied with regard to their risk factors and epidemiology. In particular, population-based data on cancer burden and patient outcomes are scant.MethodsPopulation-based data from Saarland, a federal state in South-Western Germany, were used; the data included 992 patients diagnosed with a first meningioma between 2000 and 2015. Incidence and mortality rates—as well as estimates of observed and relative survival and cumulative incidence of tumor recurrence up to 10 years after diagnosis—were derived by sex, age, WHO grade, and whether or not the patient had undergone surgery.ResultsThis population-based study not only included patients treated in the regional university hospital but also those treated elsewhere or patients without any surgical treatment. The mean age of the patients at diagnosis was 63 years, and 70%, 28% and 3% had WHO grade I, II and III meningiomas, respectively. Ten-year observed and relative survival of all patients combined was 72% and 91% respectively. Tumor-related mortality varied by sex and increased with age at diagnosis and the WHO grade of the tumor. The overall 10-year cumulative incidence of meningioma recurrence was 9%.ConclusionThis analysis represents the first modern population-based analysis of meningioma incidence and mortality and outcomes of patients with such neoplasms in Germany. Derived from an unselected sample of patients, this study may fill a hitherto existing gap in the literature on meningiomas.     

The extent of whole‐genome copy number alterations predicts aggressive features in primary melanomas

Recent evidence indicates that melanoma comprises distinct types of tumors and suggests that specific morphological features may help predict its clinical behavior. Using a SNP‐array approach, we quantified chromosomal copy number alterations (CNA) across the whole genome in 41 primary melanomas and found a high degree of heterogeneity in their genomic asset. Association analysis correlating the number and relative length of CNA with clinical, morphological, and dermoscopic attributes of melanoma revealed that features of aggressiveness were strongly linked to the overall amount of genomic damage. Furthermore, we observed that melanoma progression and survival were mainly affected by a low number of large chromosome losses and a high number of small gains. We identified the alterations most frequently associated with aggressive melanoma, and by integrating our data with publicly available gene expression profiles, we identified five genes which expression was found to be necessary for melanoma cells proliferation. In conclusion, this work provides new evidence that the phenotypic heterogeneity of melanoma reflects a parallel genetic diversity and lays the basis to define novel strategies for a more precise prognostic stratification of patients.     

Alpha-synuclein contributes to malignant progression of human meningioma via the Akt/mTOR pathway

Background

The aim of this study is to explore the expression of alpha-synuclein (α-synuclein) in benign, atypical, and anaplastic meningiomas and determine its role in the malignant progression of meningiomas.

Methods

Expression of α-synuclein was measured in 44 meningioma samples by real-time PCR analysis. The effects of overexpression or knockdown of α-synuclein on meningioma cell growth, invasiveness, and tumorigenicity were determined.

Results

Atypical and anaplastic meningiomas displayed significantly greater levels of α-synuclein mRNA, relative to benign tumors. Depletion of α-synuclein decreased cell proliferation and colony formation and promoted apoptosis in IOMM-Lee meningioma cells, whereas overexpression of α-synuclein facilitated cell proliferation and colony formation in CH-157MN meningioma cells. Silencing of α-synuclein attenuated IOMM-Lee cell migration and invasion. In contrast, ectopic expression of α-synuclein increased the invasiveness of CH-157MN cells. In vivo studies further demonstrated that downregulation of α-synuclein significantly retarded meningioma growth in nude mice. At the molecular level, the phosphorylation levels of Akt, mTOR, p70S6K and 4EBP were significantly decreased in α-synuclein-depleted IOMM-Lee cells.

Conclusions

In conclusion, α-synuclein upregulation contributes to aggressive phenotypes of meningiomas via the Akt/mTOR pathway and thus represents a potential therapeutic target for malignant meningiomas.

     

TP53 mutations in human meningiomas

Overexpression of p53 has been reported to play a role in the development of neoplasms of the central nervous system. Meningiomas are generally benign intracranial tumors originating from the meninges. Overexpression of the p53 protein in meningiomas and an association with histological type and recurrence has been reported. Mutation of the TP53 gene leads to a more stable p53 protein in quantities high enough for detection by immunohistochemistry. In the search for these mutations the core domain of the TP53 gene of meningiomas has been analyzed. Only a very low incidence of mutations was reported. The apparent discordance between overexpression of p53 protein and TP53 gene mutations may be explained by mutations located outside the core domain. This issue was addressed in the present study. All 11 exons of 17 meningiomas were analyzed for DNA alterations by PCR single-strand conformation polymorphism (PCR-SSCP) analysis with subsequent sequencing. PCR-SSCP analysis showed a various number of band shifts and nucleotide alterations, caused either by alterations in the flanking introns or common polymorphisms (codon 36 and 72). The allele frequencies of the polymorphisms found in this small population of tumors resemble the frequencies reported in the literature. In addition, three nucleotide changes located in introns 2, 3 and 7 were found in 11, 3 and 4, respectively, of 17 specimens. Based on this study and on reports by others we conclude that it is not very likely that TP53 mutations are involved in the etiology of meningiomas.     

RhoA as a mediator of clinically relevant androgen action in prostate cancer cells

Recently, we have identified serum response factor (SRF) as a mediator of clinically relevant androgen receptor (AR) action in prostate cancer (PCa). Genes that rely on SRF for androgen responsiveness represent a small fraction of androgen-regulated genes, but distinguish benign from malignant prostate, correlate with aggressive disease, and are associated with biochemical recurrence. Thus, understanding the mechanism(s) by which SRF conveys androgen regulation to its target genes may provide novel opportunities to target clinically relevant androgen signaling. Here, we show that the small GTPase ras homolog family member A (RhoA) mediates androgen-responsiveness of more than half of SRF target genes. Interference with expression of RhoA, activity of the RhoA effector Rho-associated coiled-coil containing protein kinase 1 (ROCK), and actin polymerization necessary for nuclear translocation of the SRF cofactor megakaryocytic acute leukemia (MAL) prevented full androgen regulation of SRF target genes. Androgen treatment induced RhoA activation, increased the nuclear content of MAL, and led to MAL recruitment to the promoter of the SRF target gene FHL2. In clinical specimens RhoA expression was higher in PCa cells than benign prostate cells, and elevated RhoA expression levels were associated with aggressive disease features and decreased disease-free survival after radical prostatectomy. Overexpression of RhoA markedly increased the androgen-responsiveness of select SRF target genes, in a manner that depends on its GTPase activity. The use of isogenic cell lines and a xenograft model that mimics the transition from androgen-stimulated to castration-recurrent PCa indicated that RhoA levels are not altered during disease progression, suggesting that RhoA expression levels in the primary tumor determine disease aggressiveness. Androgen-responsiveness of SRF target genes in castration-recurrent PCa cells continued to rely on AR, RhoA, SRF, and MAL and the presence of intact SRF binding sites. Silencing of RhoA, use of Rho-associated coiled-coil containing protein kinase 1 inhibitors, or an inhibitor of SRF-MAL interaction attenuated (androgen-regulated) cell viability and blunted PCa cell migration. Taken together, these studies demonstrate that the RhoA signaling axis mediates clinically relevant AR action in PCa.     

Somatic mutations in the neurofibromatosis type 2 gene in sporadic meningiomas

Meningiomas are benign tumors of the central nervous system. Although usually sporadic, they can occur in patients affected by the autosomal dominant syndrome, neurofibromatosis type 2 (NF2). The NF2 gene has recently been isolated from chromosome 22. The presence of germline mutations in NF2 patients and the loss of heterozygosity (LOH) on 22q in NF2 tumors support the hypothesis that the NF2 gene acts as a tumor suppressor. Cytogenetic and LOH studies have suggested that the gene responsible for the development of meningiomas is located in the region of 22q in which the NF2 gene maps. The meningioma gene could therefore be the NF2 gene itself. Recently, somatic mutations of the NF2 gene have been identified in sporadic meningiomas, thus supporting the hypothesis that the NF2 gene is also important in meningioma pathogenesis. In this study, we analyzed sixty-one sporadic meningiomas for LOH of 22q and for mutations in the NF2 gene. LOH was detected in 36 of the 60 informative tumors. Single-strand conformational polymorphism analysis was used to identify nine mutations in five of the eight exons of the NF2 gene studied. The nine tumors with an altered NF2 gene also showed LOH for 22q markers. These results further support the hypothesis that mutations in the NF2 gene are a critical pathogenetic event in at least some meningiomas.     

Meningioma: a cytogenetic model of a complex benign human tumor, including data on 394 karyotyped cases

Meningioma is the most frequent tumor of neuroectodermal origin in humans. It is usually benign. Only a minority of cases shows progression to an anaplastic tumor (WHO grade II and III). Meningioma is generally a sporadic tumor. Multiple and familial cases are rare and mostly associated with (hereditary) neurofibromatosis 2 (NF2). Meningiomas show an unexpectedly high recurrence rate. Also, completely removed low-grade tumors can recur. Recurrence and multiplicity are correlated with the formation of a peritumoral edema. On the cytogenetic level, meningioma is the best-studied tumor in humans. Grade I tumors show either uniform monosomy 22 or a diploid karyotype. The majority of high-grade, but only a minority of low-grade, meningiomas show loss of merlin, a cytoskeleton-cytoplasm-linker protein. Merlin is the product of the NF2 gene located on chromosome 22. A second tumor suppressor gene on chromosome 22 has not yet been detected. In contrast to other solid tumors, progression of meningiomas is correlated with increasing hypodiploidy, showing characteristic clonal evolutions that mostly include chromosomes 14, 18, and 19 and, more rarely, 6 and 10. Structural aberrations are infrequent, except for the loss of the short arm of chromosome 1, which appears to be the decisive step for anaplastic growth. Comparative histochemical and molecular cytogenetic studies point to the alkaline phosphatase gene (ALPL, liver-bone-kidney type) located on 1p36.1-->p34 as a candidate tumor suppressor gene. A model is proposed that tries to explain - with a minimum number of essential steps - the origin, progression, infiltration, and recurrence of meningiomas.     

Genome wide DNA copy number analysis of serous type ovarian carcinomas identifies genetic markers predictive of clinical outcome

Ovarian cancer is the fifth leading cause of cancer death in women. Ovarian cancers display a high degree of complex genetic alterations involving many oncogenes and tumor suppressor genes. Analysis of the association between genetic alterations and clinical endpoints such as survival will lead to improved patient management via genetic stratification of patients into clinically relevant subgroups. In this study, we aim to define subgroups of high-grade serous ovarian carcinomas that differ with respect to prognosis and overall survival. Genome-wide DNA copy number alterations (CNAs) were measured in 72 clinically annotated, high-grade serous tumors using high-resolution oligonucleotide arrays. Two clinically annotated, independent cohorts were used for validation. Unsupervised hierarchical clustering of copy number data derived from the 72 patient cohort resulted in two clusters with significant difference in progression free survival (PFS) and a marginal difference in overall survival (OS). GISTIC analysis of the two clusters identified altered regions unique to each cluster. Supervised clustering of two independent large cohorts of high-grade serous tumors using the classification scheme derived from the two initial clusters validated our results and identified 8 genomic regions that are distinctly different among the subgroups. These 8 regions map to 8p21.3, 8p23.2, 12p12.1, 17p11.2, 17p12, 19q12, 20q11.21 and 20q13.12; and harbor potential oncogenes and tumor suppressor genes that are likely to be involved in the pathogenesis of ovarian carcinoma. We have identified a set of genetic alterations that could be used for stratification of high-grade serous tumors into clinically relevant treatment subgroups.     

Probable destructive meningioma in an archaeological adult male skull from Alaska

Several intracranial pathological conditions can affect the bones of the skull. The most common cause of these conditions is tumor, but infection and other diseases are also known to affect the bones of the skull. Distinguishing between the various causes of intracranial skeletal pathology in archaeological human remains is usually a challenging exercise, and a specific diagnosis will often be impossible. Meningiomas are tumors that arise in arachnoid tissues embedded in the outer layer of the dura. Because of this association, they occur almost exclusively in the skull and vertebral column. Usually meningiomas are slow-growing tumors that do not metastasize to other organs and tissues of the body. However, rare cases can be malignant and, even when meningiomas are benign, their presence and growth can adversely affect the nervous and vascular supply to other tissues in the skull and vertebral column. Their effect on adjacent bone tissue varies from stimulating bone-forming lesions to causing highly destructive lesions. A few examples of meningioma have been described in the paleopathological literature. Most of these cases are bone-stimulating meningiomas. The case presented here is a probable example of a highly destructive meningioma of the skull base, with unilateral extension into the left side of the cranium. This case is compared with a modern clinical case of destructive intracranial meningioma that was documented both radiographically and pathologically. Destructive meningiomas can be confused with other pathological conditions, including benign and malignant tumors. Criteria for differentiating the diagnostic options are reviewed.     

脑膜瘤WHO 分级及其基因学研究进展

脑膜瘤是颅内第二大常见肿瘤。WHO根据组织学及肿瘤生物学将脑膜瘤分为3级。目前,随着脑膜瘤研究的深入及肿瘤分子生物学的发展,脑膜瘤在基因学方面的研究越来越受到重视。其中包括原癌基因的激活和抑癌基因的失活。现阶段,比较公认的与肿瘤包括脑膜瘤形成有关的原癌基因主要有c-myc基因、bcl-2基因、Survivin基因及mdm2基因等,抑癌基因主要有p53基因、p73基因、NF-2基因及p16基因等。这些基因通过不同的机制发生不同的改变最终参与了不同级别脑膜瘤的发生和发展。现就其研究进展做一综述。     

Proportion of S-phase tumor cells measured by flow cytometry is an independent prognostic factor in meningioma tumors.

Meningiomas are tumors in arachnoid cells which represent up to one fifth of all intracranial tumors and up to a quarter of spinal neoplasias. Although meningiomas have classically been considered to be benign tumors, it has also been well-established that they show a heterogeneous clinical outcome. To the best of our knowledge no study has yet been performed in which the independent prognostic value of both DNA ploidy and cell cycle has been simultaneously assessed in a large series of meningioma tumors. The aim of the present study was to prospectively explore the prognostic value of DNA ploidy status and the proliferative rate of tumor cells in a series of 105 consecutive meningioma patients studied at diagnosis. Both the presence of DNA aneuploidy and the proportion of S-phase tumor cells were analyzed in all cases in fresh tumors obtained during diagnostic surgery. From the technical point of view, we followed the recommendations of the Consensus Conference on Flow Cytometry DNA Analysis held in October 1992. Our results show that meningioma tumors display a relatively low incidence of DNA aneuploidy (14%), and they usually show a low proliferative rate (mean percentage of S-phase cells of 1.3 +/- 0.3%). The presence of DNA aneuploidy was associated with a higher incidence of aggressive histopathologic subtypes (P = 0.045), a greater age (P = 0.009), location at the cerebral convexity (P = 0.004), and a greater proportion of S-phase cells (P = 0.005). In contrast, no significant association between the DNA ploidy status of meningioma patients and their disease-free survival was found (P = 0. 1). Regarding the proliferative activity of neoplastic cells, we found that a high proportion of S-phase cells (>1.8%) was associated with a significantly lower mean age (P = 0.007), aggressive histopathologic subtypes (P = 0.03), a higher incidence of DNA aneuploidy (P = 0.004), and a significantly shorter disease-free survival (P < 0.004). Multivariate analysis of prognostic factors showed that the proportion of S-phase tumor cells was the most powerful independent prognostic factor in meningioma patients (P = 0.02). In summary, we conclude that the proportion of S-phase tumor cells represents the individual parameter with the highest value for predicting disease-free survival in meningioma patients.