Molecular study of frequency of mosaicism in neurofibromatosis 2 patients with bilateral vestibular schwannomas.
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Neurofibromatosis 2 (NF2) is a severe autosomal dominant disorder that predisposes to multiple tumours of the nervous system. About half of all patients are founders with clinically unaffected parents. The purpose of the present study was to examine the extent to which mosaicism is present in NF2 founders. A total of 233 NF2 founders with bilateral vestibular schwannomas (BVS) were screened by exon scanning. NF2 mutations were detected in the blood samples of 122 patients (52%). In 10 of the 122 cases, the ratio of mutant to normal alleles was obviously less than 1, suggesting mosaicism. Tumour specimens were available from 35 of the 111 subjects in whom no mutation could be detected in blood specimens. Mutational analysis by exon scanning detected typical NF2 mutations in 21 of the 35 tumours. In nine subjects, the alterations found in tumours could be confirmed to be the constitutional mutation based on finding of identical mutations in pathologically and/or anatomically distinct second tumours. In six other subjects with only a single tumour available, allelic loss of the NF2 gene was found in addition to the mutation in each tumour, suggesting that either the mutation or the deletion of the NF2 gene is probably the constitutional genetic alteration. Our results suggest that failure to find constitutional mutations in blood specimen from these 15 patients was not because of the limitation of the applied screening technique, but the lack of the mutations in their leucocytes, best explained by mosaicism. Extrapolating the rate (15/35 = 43%) of mosaicism in these 35 cases to the 111 NF2 founders with no constitutional NF2 mutations found in their blood, we inferred 48 mosaic subjects (111 x 0.429). Adding the 10 mosaic cases detected directly in blood specimens, we estimate the rate of mosaicism to be 24.8% (58/233) in our cohort of 233 NF2 founders with bilateral vestibular schwannomas. (+info)
Quinidine impairs proliferation of neurofibromatosis type 2-deficient human malignant mesothelioma cells.
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BACKGROUND: Human malignant mesotheliomas (HMMs) are aggressive tumors that arise from the mesothelium. They respond poorly to conventional tumor treatment and outcome is often fatal. Inactivating mutations of the neurofibromatosis type 2 (NF2) tumor suppressor gene merlin have been described in nearly 60% of primary malignant mesothelioma and in approximately 20% of the mesothelioma cell lines. Studies regarding human NF2 schwannoma cells revealed a higher proliferation and a larger noninactivating K(+) outward current compared with controls. The enhanced proliferation of merlin-deficient NF2 schwannoma cells could be reduced in the presence of quinidine, a K(+) channel blocker, whereas the proliferation of normal Schwann cells is not affected. The current study was undertaken to evaluate the effect of quinidine on the proliferation of HMM cell lines in relation to their NF2 status. METHODS: Proliferation analyses using bromodeoxyuridine incorporation was performed by immunocytochemical staining and fluorescence assisted cell sorting. The patch-clamp technique was applied for electrophysiologic characterization of the HMM cell lines. The cytochrome P450 2D6 locus, known to be mutated at high frequencies in NF2 patients and to be specifically inhibited by quinidine, was screened for mutations by cycle sequencing. RESULTS: Quinidine selectively reduces the proliferation of merlin-deficient HMM cell lines by causing a G(0)/G(1) arrest, whereas the proliferation rates of merlin-expressing HMM cell lines remain unchanged. The effect of quinidine on the proliferation of HMM cell lines appears to be correlated with the NF2 gene status but not with the K(+) outward current. No relation to cytochrome P450 2D6 mutations was detected. CONCLUSIONS: Quinidine or quinidine analogs are of potential therapeutic interest for the subset of merlin-deficient mesothelioma tumors. (+info)
Upregulation of the Rac1/JNK signaling pathway in primary human schwannoma cells.
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Schwann cells lacking the tumor-suppressor-protein merlin tend in man to build benign tumors (schwannoma). We observed that characteristic features of these cells which are relevant to tumorigenicity resemble those described in cells with high Rac activity. Moreover this small GTPase also phosphorylates merlin via PAK activation. We hypothesized that merlin deficiency might cause an activation of Rac and its dependent signaling pathways, in particular the pro-tumorigenic JNK pathway. We show an enhanced activation of Rac1 in primary human schwannoma cells, find both Rac and its effector PAK at the membrane where they colocalize, and describe increased levels of phosphorylated JNK in the nucleus of these cells. Further we describe regulation at post-transcriptional level with upregulated protein, but not mRNA levels for Rac1, and JNK1/2. We conclude that merlin regulates Rac activation, and suggest that this is important for human schwannoma cell dedifferentiation. (+info)
Clinical experience with gamma knife stereotactic radiosurgery in the management of vestibular schwannomas secondary to type 2 neurofibromatosis.
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OBJECTIVE: To evaluate the results of stereotactic radiosurgery treating vestibular schwannomas secondary to type 2 neurofibromatosis. METHODS: A retrospective review of 122 type 2 neurofibromatosis vestibular schwannomas consecutively treated in 96 patients. Tumour control was assessed by recourse to surgical intervention, by serial radiological imaging, and by the calculation of relative growth ratios in patients (n=29) habouring untreated contralateral tumours to act as internal controls. Hearing function was assessed with Gardner-Robertson grades and with averaged pure tone audiogram thresholds. Other complications are detailed. RESULTS: Applying current techniques, eight years after radiosurgery it was estimated that 20% of patients will have undergone surgery for their tumour, 50% will have radiologically controlled tumours, and in 30% there will be some variable concern about tumour control, but up to that time they will have been managed conservatively. Relative growth ratios one and two years after treatment indicate that radiosurgery confers a significant (p=0.01) advantage over the natural history of the disease. Analysis of these ratios beyond two years was precluded by the need to intervene and radiosurgically treat the contralateral control tumours in more than 50% of the cases. This growth control was achieved with 40% of patients retaining their Gardner-Robertson hearing grades three years after treatment, (40% having some deterioration in grade, 20% becoming deaf). Pure tone audiogram results suggest some progressive long term hearing loss, although interpretation of this is difficult. Facial and trigeminal neuropathy occurred in 5% and 2%. CONCLUSIONS: Radiosurgery is a valuable minimally invasive alternative treatment for these tumours. For most patients, it controls growth or defers the need for surgery, or both. There is a price in terms of hearing function, although this may compare favourably with the deafness associated with the natural history of the disease, and with surgery. In deciding on therapy, patients should be aware of this treatment option. (+info)
Bilateral optic nerve sheath meningiomas in a patient with neurofibromatosis type 2.
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A 34-year-old woman who presented with hearing loss and tinnitus was found to have reduced vision bilaterally. Computed tomography scan revealed bilateral acoustic neuromas and bilateral optic nerve sheath meningiomas. The presence of bilateral acoustic neuromas fulfils the criteria for the diagnosis of central neurofibromatosis (neurofibromatosis type 2). Although this is the first report of bilateral optic nerve sheath meningioma in neurofibromatosis type 2, meningiomas are commoner in this dominantly inherited disorder, than in its absence and both forms of central nervous system tumour may be caused by loss of tumour suppressor genes on chromosome 22. (+info)
Merlin, the product of the Nf2 tumor suppressor gene, is an inhibitor of the p21-activated kinase, Pak1.
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The Nf2 tumor suppressor gene codes for merlin, a protein whose function has been elusive. We describe a novel interaction between merlin and p21-activated kinase 1 (Pak1), which is dynamic and facilitated upon increased cellular confluence. Merlin inhibits the activation of Pak1, as the loss of merlin expression results in the inappropriate activation of Pak1 under conditions associated with low basal activity. Conversely, the overexpression of merlin in cells that display a high basal activity of Pak1 resulted in the inhibition of Pak1 activation. This inhibitory function of merlin is mediated through its binding to the Pak1 PBD and by inhibiting Pak1 recruitment to focal adhesions. This link provides a possible mechanism for the effect of loss of merlin expression in tumorigenesis. (+info)
Identification of recurrent regions of chromosome loss and gain in vestibular schwannomas using comparative genomic hybridisation.
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BACKGROUND: Schwannomas are benign tumours of the nervous system that are usually sporadic but also occur in the inherited disorder neurofibromatosis type 2 (NF2). The NF2 gene is a tumour suppressor on chromosome 22. Loss of expression of the NF2 protein product, merlin, is universal in both sporadic and NF2 related schwannomas. The GTPase signalling molecules RhoA and Rac1 regulate merlin function, but to date only mutation in the NF2 gene has been identified as a causal event in schwannoma formation. METHODS: Comparative genomic hybridisation (CGH) was used to screen 76 vestibular schwannomas from 76 patients (66 sporadic and 10 NF2 related) to identify other chromosome regions that may harbour genes involved in the tumorigenesis. RESULTS: The most common change was loss on chromosome 22, which was more frequent in sporadic than in NF2 related tumours. Importantly, eight tumours (10%) showed gain of copy number on chromosome 9q34. Each of the two NF2 patients who had received stereotactic radiotherapy had non-chromosome 22 changes, whereas only one of eight non-irradiated NF2 patients had any chromosome changes. Three tumours had gain on 17q, which has also been reported in malignant peripheral nerve sheath tumours that are associated with neurofibromatosis type 1. Other sites that were identified in three or fewer tumours were regions on chromosomes 10, 11, 13, 16, 19, 20, X, and Y. CONCLUSIONS: These findings should be verified using techniques that can detect smaller genetic changes, such as microarray-CGH. (+info)
CpG island methylation in sporadic and neurofibromatis type 2-associated schwannomas.
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PURPOSE: The purpose of this research was to examine the DNA methylation profile of schwannomas. EXPERIMENTAL DESIGN: We examined the DNA methylation status of 12 tumor-related genes (NF2, RB1, p14(ARF), p16(INK4a), p73, TIMP-3, MGMT, DAPK, THBS1, caspase-8, TP53, and GSTP1) in 44 sporadic and/or NF2-associated schwannomas using methylation-specific PCR. RESULTS: The most frequently methylated genes were THBS1 (36%), p73 (27%), MGMT (20%), NF2 (18%), and TIMP-3 (18%). The RB1/p16INK4a gene pair displayed aberrant methylayed alleles in 15% of cases, whereas methylation was relatively rare in the other genes (<5%). Methylation was tumor specific because it was absent in two nonneoplastic nerve sheath samples and two nonneoplastic brain samples studied as controls. CONCLUSIONS: Our findings indicate that aberrant methylation seems to be a mechanism for NF2 gene inactivation, considered an early step in schwannoma tumorigenesis, and as well, aberrant hypermethylation of other tumor-related genes might represent secondary events that also contribute to the development of these tumors. (+info)