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Schwannomatosis is the third major form of neurofibromatosis and is characterized by the development of multiple schwannomas in the absence of bilateral vestibular schwannomas. The 2011 Schwannomatosis Update was organized by the Children’s Tumor Foundation (www.ctf.org) and held in Los Angeles, CA, from June 5–8, 2011. This article summarizes the highlights presented at the Conference and represents the “state-of-the-field” in 2011. Genetic studies indicate that constitutional mutations in the SMARCB1 tumor suppressor gene occur in 40–50% of familial cases and in 8–10% of sporadic cases of schwannomatosis. Tumorigenesis is thought to occur through a four-hit, three-step model, beginning with a germline mutation in SMARCB1 (hit 1), followed by loss of a portion of chromosome 22 that contains the second SMARCB1 allele and one NF2 allele (hits 2 and 3), followed by mutation of the remaining wild-type NF2 allele (hit 4). Insights from research on HIV and pediatric rhabdoid tumors have shed light on potential molecular pathways that are dysregulated in schwannomatosis-related schwannomas. Mouse models of schwannomatosis have been developed and promise to further expand our understanding of tumorigenesis and the tumor microenvironment. Clinical reports have described the occurrence of intracranial meningiomas in schwannomatosis patients and in families with germline SMARCB1 mutations. The authors propose updated diagnostic criteria to incorporate new clinical and genetic findings since 2005. In the next 5 years, the authors expect that advances in basic research in the pathogenesis of schwannomatosis will lead toward clinical investigations of potential drug therapies.

Objective: Clinically validated biomarkers for neurofibromatosis 1 (NF1), neurofibromatosis 2 (NF2), and schwannomatosis (SWN) have not been identified to date. The biomarker working group9s goals are to (1) define biomarker needs in NF1, NF2, and SWN; (2) summarize existing data on biomarkers in NF1, NF2, and SWN; (3) outline recommendations for sample collection and biomarker development; and (4) standardize sample collection and methodology protocols where possible to promote comparison between studies by publishing standard operating procedures (SOPs). Methods: The biomarker group reviewed published data on biomarkers in NF1, NF2, and SWN and on biobanking efforts outside these diseases via literature search, defined the need for biomarkers in NF, and developed recommendations in a series of consensus meetings. Results: We describe existing biomarkers in NF and report consensus recommendations for SOP and a minimal clinical dataset to accompany samples derived from patients with NF1, NF2, and SWN in decentralized biobanks. Conclusions: These recommendations are intended to provide clinicians and researchers with a common set of guidelines to collect and store biospecimens and for establishment of biobanks for NF1, NF2, and SWN.

INTRODUCTION: A significant minority of meningiomas are difficult to treat with surgery or radiotherapy, and there are limited chemotherapeutic alternatives. This study aims to better understand the pathways that are active in these tumours, in order to direct future treatment strategies. METHOD: We have investigated the expression of several growth factors and their signalling pathways in 30 meningiomas, using immunohistochemistry and Western blots. Expression was correlated with presumed NF2 gene status, using fluorescent in situ hybridisation to look for chromosome 22q loss. RESULTS: Membrane expression of VEGFR and PDGFR-beta was seen in 83% of tumours, Axl in 70%, EGFR in 50% and IGFR in 47%. Expression was similar in low and high grade tumours, but membrane EGFR expression was not seen in tumours showing chromosome 22q loss. Expression of the ligands IGF, NRG, VEGF, Gas 6, and downstream signalling proteins, Mek, Erk, Jnk and Akt, and pS6RP, was widespread. Our findings suggest that the majority of meningiomas express and show activation of multiple growth factor receptors and their downstream signalling pathways irrespective of tumour grade. CONCLUSION: In addition to previously reported receptors, Axl may also offer an attractive therapeutic target. Our findings also suggest that anti-EGFR based therapies may be less effective in meningiomas with 22q loss.

Neurofibromatosis type 2 (NF2) is a tumor suppressor syndrome characterized by bilateral vestibular schwannomas (VS) which often result in deafness despite aggressive management. Meningiomas, ependymomas, and other cranial nerve and peripheral schwannomas are also commonly found in NF2 and collectively lead to major neurologic morbidity and mortality. Traditionally, the overall survival rate in patients with NF2 is estimated to be 38% at 20 years from diagnosis. Hence, there is a desperate need for new, effective therapies. Recent progress in understanding the molecular basis of NF2 related tumors has aided in the identification of potential therapeutic targets and emerging clinical therapies. In June 2010, representatives of the international NF2 research and clinical community convened under the leadership of Drs. D. Gareth Evans (University of Manchester) and Marco Giovannini (House Research Institute) to review the state of NF2 treatment and clinical trials. This manuscript summarizes the expert opinions about current treatments for NF2 associated tumors and recommendations for advancing therapies emerging from that meeting. The development of effective therapies for NF2 associated tumors has the potential for significant clinical advancement not only for patients with NF2 but for thousands of neuro-oncology patients afflicted with these tumors.

The 2011 annual meeting of the Children’s Tumor Foundation, the annual gathering of the neurofibromatosis (NF) research and clinical communities, was attended by 330 participants who discussed integration of new signaling pathways into NF research, the appreciation for NF mutations in sporadic cancers, and an expanding pre-clinical and clinical agenda. NF1, NF2, and schwannomatosis collectively affect approximately 100,000 persons in US, and result from mutations in different genes. Benign tumors of NF1 (neurofibroma and optic pathway glioma) and NF2 (schwannoma, ependymoma, and meningioma) and schwannomatosis (schwannoma) can cause significant morbidity, and there are no proven drug treatments for any form of NF. Each disorder is associated with additional manifestations causing morbidity. The research presentations described in this review covered basic science, preclinical testing, and results from clinical trials, and demonstrate the remarkable strides being taken toward understanding of and progress toward treatments for these disorders based on the close interaction among scientists and clinicians.

Deficiency of the tumor suppressor protein merlin leads to the development of benign tumors of the nervous system such as schwannomas, ependymomas and meningiomas. These tumors can occur spontaneously or as part of a tumor predisposition syndrome called neurofibromatosis type 2 (NF2), which involves multiple tumors. Schwannomas are the hallmark tumors of NF2 and are the most frequent and well-characterized of the merlin-deficient tumors. Surgery or radiotherapy are used to treat single tumors and can leave the patient with substantial morbidity. Limitations of other treatment options for merlin-deficient tumors, such as the lack of effectiveness of chemotherapy, have led to an urgent requirement for new pharmaceutical therapies. Merlin-deficient tumors are genetically well-defined, which allows rational testing of new molecular therapies that have been developed and successfully used to treat various cancers in the past few years. This Review centers on four key families of receptor tyrosine kinases-the ErbB family, platelet-derived growth factor receptor β, insulin-like growth factor 1 receptor, and vascular endothelial growth factor receptors-focusing on their role in schwannoma pathobiology and the therapeutic potential of targeting these receptors and their downstream signaling pathways.

Vestibular schwannomas (VS) arising sporadically or in patients with neurofibromatosis type 2 (NF2) consistently lack expression of Merlin, a tumor suppressor. Conventional treatment options include surgery and radiotherapy but there is no validated medical option. Recent evidence suggests that Merlin deficiency may result in abnormal activation of receptor tyrosine kinases (RTKs) and downstream signaling, promoting tumor growth. Although small-molecule RTK inhibitors are widely available for clinical use, no such therapy has been validated in patients with VS. To screen for RTK activation, surgical VS specimens from patients with and without NF2 were analyzed by phospho-RTK profiling arrays. Downstream signaling pathway activation was analyzed by phospho-MAPK arrays. Activated RTKs and downstream kinases were validated immunohistochemically in corresponding formalin-fixed, paraffin-embedded tissues. Phospho-RTK arrays and immunohistochemistry showed consistent overexpression and activation of EGFR family receptors and evidence of ERK1/2 downstream signaling was observed in all samples analyzed (n = 11). Based on the findings, the small-molecule EGFR/ErbB2 kinase inhibitor lapatinib was selected for evaluation of target inhibition and treatment efficacy in our in vitro human schwannoma model. EGFR/ErbB2 targeted therapy with lapatinib inhibited ErbB2 phosphorylation and survivin upregulation, as well as downstream ERK1/2 and AKT activation, resulting in decreased proliferation. We conclude that EGFR family receptor activation is a consistent feature of both sporadic and NF2-related VS. Molecular targeted therapy with lapatinib downregulates survivin and has antiproliferative activity in a preclinical VS model. Based on these findings, a clinical trial with lapatinib for the treatment of VS is currently underway.

The epidemiology of motor neurone disease (MND) in the counties of Devon and Cornwall in the southwest of England has not previously been studied. A previous study of England and Wales has, however, reported a very high death certification rate of MND in Devon. This study was carried out to establish the prevalence and incidence of MND in Devon and Cornwall and make comparisons with published rates in other populations. We attempted to identify all cases of MND diagnosed in Devon and Cornwall between 2002 and 2007. Case identification was centred on the major hospitals in the two counties and multiple sources of ascertainment were used. All identified cases had their case notes reviewed to establish the diagnosis and classify by type of MND. Point prevalence of MND was established for September 1st, 2007. The overall incidence rate standardised to the 2001 United Kingdom population was 2.52 per 100,000 (95% confidence interval 2.20-2.84). The incidence rate was significantly higher in males (P < 0.001). The estimated male to female incidence ratio was 2.10 (95% CI 1.61-2.73). The crude incidence rate in Cornwall was 3.78 per 100,000 (95% CI 3.03-4.53) and this was significantly higher (P = 0.011) than the rate in Devon, which was 2.61 per 100,000 (95% CI 2.19-3.04). The standardised incidence rate for the study period in Devon was 2.26 per 100,000 (95% CI 1.91-2.60) and in Cornwall it was 3.06 per 100,000 (95% CI 2.44-3.68). The overall standardised point prevalence rate was 5.66 per 100,000 (95% CI 4.49-6.83). The incidence rate of MND in our study is similar to reported findings in large prospective studies of the disease. There is a significant difference between the incidence rates in Devon and Cornwall. There is a need to establish a prospective MND Register to accurately document the epidemiological characteristics of the disease in the two counties.

A minority of meningiomas are difficult to treat with surgery or radiotherapy, and chemotherapeutic alternatives are limited. This study aims to better understand pathways that are active in meningiomas, in order to direct future treatment strategies. We investigated the expression and activation of multiple growth factor receptors, their ligands and downstream signalling pathways in 30 meningiomas using immunohistochemistry. Expression was correlated with chromosome 22q loss. Membrane expression of VEGF receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR)β was seen in 83% of tumors, Axl in 70%, EGFR in 50% and insulin-like growth factor receptor in 47%. Expression was similar in low- and high-grade tumors, but membrane EGFR expression was not seen in tumors showing chromosome 22q loss (P 0.05). Expression of ligands (IGF, NRG, VEGF, Gas 6), and signalling proteins (Mek, Erk, Jnk, Akt) and pS6RP, was widespread. Western blot confirmed widespread Axl expression and supported selective expression of EGFR in NF2-intact meningiomas. The majority of meningiomas express and show activation of multiple growth factor receptors and their signalling pathways, irrespective of tumor grade. In addition to previously reported receptors, Axl offers a new therapeutic target. The findings also suggest that anti-EGFR based therapies may be less effective in meningiomas with 22q loss.

Mutation detection in the neurofibromatosis type 2 (NF2) gene is challenging because when combining mutation detection methods such as single-strand conformational polymorphism and heteroduplex analysis, denaturing gradient gel electrophoresis, and direct sequencing of aberrant polymerase chain reaction (PCR) fragments only 30 to 60% of the constitutional mutations are detected. Because large deletions and complete chromosome rearrangements are also described methods such as microarray-comparative genomic hybridization and fluorescence in situ hybridization are also used. The one type of mutation often missed corresponds to deletions encompassing one or few exons. To detect this type we have developed a swift and reliable method. We perform a gene dosage analysis with two fluorescent multiplex PCR assays that amplify 15 of the 17 NF2 exons. The labeled PCR products are quantified and gene dose is calculated with respect to controls. We tested the reliability of this method with DNA from eight NF2 patients with known heterozygous NF2 deletions, eight controls and four unknown NF2 patients. In all of the patients with known heterozygous deletions we found in several exons a reduction of gene dosage to 50 to 69%. In one NF2 patient with previously unknown mutation and a severe phenotype we found the gene dosage of two exons reduced by 50% indicating a deletion of these two exons on one allele. This finding was validated by reverse transcriptase-PCR on fibroblast and schwannoma cell cultures of this patient and cDNA sequencing. Our gene dosage assay will detect deletions of one or more exons as well as gross deletions of the whole coding region of the gene. It can complement the existing screening methods because it is faster and easier.

This MRI study was performed to evaluate in vivo alterations of the spinal cord in defined subgroups of motor neuron diseases. Standard MRI examinations of the cervical and thoracic spinal cord in sporadic amyotrophic lateral sclerosis (ALS; n = 39), sporadic lower motor neuron disease (LMND; n = 19), Kennedy’s disease (KD; n = 19) and a control group (n = 96) were analyzed with respect to spinal cord signal changes and the thickness of the spinal cord. No significant changes in thickness or signal alterations were observed when comparing ALS, LMND and control groups with one another. However, in KD patients significant upper spinal cord atrophy was detected at the cervical level as compared with all other groups. At the thoracic level, KD patients had significant upper cord atrophy as compared with controls and LMND. Marked atrophy of the upper spinal cord seems to be a feature of the KD-associated central-peripheral distal axonopathy.

Medulloblastoma is the commonest brain tumor in childhood and in a minority of patients is associated with an underlying genetic disorder such as Gorlin syndrome or familial adenomatous polyposis. Increased susceptibility to certain tumors, including neuroblastoma and some hematological malignancies, is recognized in disorders caused by mutations in genes encoding components of the RAS signaling pathway which include Noonan syndrome, Noonan syndrome with multiple lentigines (NSML; formerly called LEOPARD syndrome), Costello syndrome, Cardiofaciocutaneous syndrome, Legius syndrome, and Neurofibromatosis type 1 (NF1), collectively termed RASopathies. Although an association between medulloblastoma and NF1 has been reported, this tumor has not previously been reported in other RASopathies. We present a patient with NSML caused by the recurrent PTPN11 mutation c.1403C > T (p.Thr468Met) in whom medulloblastoma was diagnosed at age 10 years. Medulloblastoma could therefore be part of the tumor spectrum associated with this disorder.

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.

The neurofibromatosis type 2 (NF2) tumor suppressor merlin plays an important role in contact inhibition of cell growth. It shows high similarity to the ezrin-radixin-moesin (ERM) proteins with which it shares three distinct structural domains: a highly conserved N-terminal FERM (Four-point-one, ERM) domain (residues 1-335), an α-helical domain (residues 336-505), and a C-terminal tail domain (CTT, residues 506-595) that can associate with the FERM domain to modulate the accessibility to its ligands, which include the scaffolding protein EBP50 (reviewed in (Fehon et al., 2010)). ERMs are activated in part by phosphorylation of a conserved threonine in the CTT that reduces the FERM/CTT association to convert the protein from a closed to a more open active state. Similar to ERMs, merlin is regulated by phosphorylation on a unique CTT residue (Ser518) (reviewed in (Bretscher et al., 2002)). Limited evidence suggests that similar to ERMs, phosphorylation of Ser518 reduces the FERM/CTT association (Rong et al., 2004; Shaw et al., 2001). However, in contrast to the ERMs, studies consistently show that this phosphorylation event leads to deactivation of merlin (Shaw et al., 2001; Surace et al., 2004) and a binary switch model has emerged that non-phosphorylated closed merlin represents the active state and phosphorylation opens the molecule to inactivate it. This model presents a paradox, as for example the interaction of merlin with growth factor receptors through EBP50 is required for growth regulation (Curto et al., 2007; James et al., 2004), yet would not be expected to occur with closed merlin. We set out to rigorously test the current model by characterizing a spectrum of merlin variants with different degrees of association of the CTT with the FERM domain. Since the merlin FERM/CTT interaction is not as strong as in ezrin (Nguyen et al., 2001), we engineered (based on similarities in the CTT of merlin and EBP50; Figure S1A, B) a merlin mutant (designated ‘AR’) having a more stable closed form. As strongly open controls, we used a mutant missing the last two residues (ΔEL) and the splice variant isoform 2 (M2) with altered CTT (Nguyen et al., 2001). We compared these with wild type (WT) merlin, the phosphodeficient S518A mutant, the phosphomimetic S518D (Shaw et al., 2001) and S518E mutants, and a merlin fragment genuinely phosphorylated on Ser518. First we compared the interaction of the variant Cter fragments (residues 342-end) with the Nter (residues 1-341) in vitro. As predicted, the AR Cter bound Nter most effectively (Figure 1A, B). Surprisingly and in contrast to earlier findings, S518A Cter showed reduced Nter binding, whereas Cter containing S518D or S518E mutations exhibited increased binding. Similar results were obtained with shorter fragments containing only the CTT and FERM domains (Figure S1C, D) and when the Cter fragments were used to recover Nter expressed in cells (Figure S1E). Moreover, when these fragments were co-expressed in 293T cells, Cter containing S518D and AR mutations efficiently recovered the Nter, but WT, S518A, M2, and ΔEL constructs recovered little or none (Figure 1C). To directly determine the effect of S518 phosphorylation on the Nter/Cter interaction, WT Cter was phosphorylated in vitro. Phosphorylation (~30% yield) resulted in an ATP-dependent shifted band specifically recognized by a S518-phosphomerlin antibody and not seen with S518A (Figure 1D). Immobilized Nter bound the phosphorylated Cter about 10-fold more effectively, binding about 11% of the input compared to 1% for the unphosphorylated species (Figure 1E), directly demonstrating that phosphorylation of S518 enhances the FERM/CTT interaction. Figure 1 A. GST-Cter fragments bound to glutathione-Sepharose beads were used to recover untagged Nter expressed in bacteria. B. Quantification of Nter binding (mean ± SD). C. 293 cells expressing HA-Nter and either vector (V) or Myc-tagged Cter (Myc-Cter). ... Since the binding site for EBP50 on the FERM domain is masked by bound CTT (Bretscher et al., 2002), the accessibility of this site reflects full-length merlin’s conformation. Full-length AR mutant expressed in 293 cells fails to bind immobilized EBP50 and is therefore highly closed, whereas ΔEL bound as effectively as the free FERM domain. WT (which is subject to phosphorylation in vivo) and S518D bound poorly, whereas S518A bound more efficiently (Figure 1F). Similar results were obtained with bacterially purified proteins (Figure S1F). Taken together, our results document a hierarchy of closure, with the AR mutant the most tightly closed, followed by S518D, which is more closed than unmodified WT, then S518A, and finally M2 and ΔEL which are fully open. We assessed the ability of these mutants to control cell growth, using the nonfunctional L64P mutant (Gutmann et al., 1998) as a control. All constructs, except mutant L64P, localized mostly to the plasma membrane, indicating that both open and closed conformations of merlin associate with the plasma membrane (data not shown). Mutant L64P and the more closed mutants AR and S518D failed to block the proliferation of merlin-deficient human schwannoma cells, whereas WT merlin, and the more open S518A and M2 were proficient at growth control, even though their expression level was significantly lower compared to the closed mutants (Figures 1G-H). Consistent results were obtained with mouse 3T3 cells and primary human schwannoma cells. In all cases a more open conformation correlates with higher growth control activity and enhances association with growth factor receptors through EBP50 (Figure S1G-J). In accord with the rheostat model of conformational regulation suggested for ERM proteins (Li et al., 2007), these results show that WT merlin tends not to be fully open or closed, but varies in its degree of openness depending on its circumstances. In contrast to previous work (Shaw et al., 2001; Surace et al., 2004), our studies support a model in which the conformationally open state of merlin is the one that favors growth control, with S518 phosphorylation being a central regulatory event enhancing the FERM/CTT association, tipping the balance toward a more (but not fully) closed molecule with reduced ability to control cell growth. Unmodified merlin (like S518A) is in a more open state and is therefore more active in growth control. Since the amount of active merlin necessary for growth control may vary between different experimental settings, it is not surprising to see the apparently variable results found in the field. That S518A is not as open as M2 or ΔEL and S518D is not as closed as AR implies that additional factors can further regulate the balance between open and closed states. With this new model in hand, and the AR and ΔEL mutants available as tightly closed and open forms of merlin, the stage is set for studies to identify and characterize in detail the effectors that selectively associate with merlin in its different conformations and how these impinge on the regulation of cell growth and other merlin functions.

Purpose of the review Secondary axonal atrophy is common in most if not all demyelinating neuropathies and is likely responsible for the majority of clinical symptoms. We review clinical, electrophysiological and morphological evidence for secondary axonal atrophy in demyelinating neuropathies and summarize recent hypotheses on possible pathomechanisms. Recent findings Elucidation of genetic defects responsible for hereditary demyelinating neuropathies and insights into axon-Schwann cell interactions have allowed longitudinal studies of genetically defined demyelinating neuropathies and research into the pathomechanism of secondary axonal atrophy. Summary There is ample clinical electrophysiological and electropathological evidence that secondary axonal atrophy is found in hereditary and demyelinating neuropathies. Recognizing secondary axonal atrophy as a main cause for clinical disability in demyelinating neuropathies is important for the clinician and may reveal a therapeutic target common to all different forms of demyelinating neuropathies.

The neurofibromatoses (NF) are autosomal dominant genetic disorders that encompass the rare diseases NF1, NF2, and schwannomatosis. The NFs affect more people worldwide than Duchenne muscular dystrophy and Huntington's disease combined. NF1 and NF2 are caused by mutations of known tumor suppressor genes (NF1 and NF2, respectively). For schwannomatosis, although mutations in SMARCB1 were identified in a subpopulation of schwannomatosis patients, additional causative gene mutations are still to be discovered. Individuals with NF1 may demonstrate manifestations in multiple organ systems, including tumors of the nervous system, learning disabilities, and physical disfigurement. NF2 ultimately can cause deafness, cranial nerve deficits, and additional severe morbidities caused by tumors of the nervous system. Unmanageable pain is a key finding in patients with schwannomatosis. Although today there is no marketed treatment for NF-related tumors, a significant number of clinical trials have become available. In addition, significant preclinical efforts have led to a more rational selection of potential drug candidates for NF trials. An important element in fueling this progress is the sharing of knowledge. For over 20 years the Children's Tumor Foundation has convened an annual NF Conference, bringing together NF professionals to share novel findings, ideas, and build collaborations. The 2012 NF Conference held in New Orleans hosted over 350 NF researchers and clinicians. This article provides a synthesis of the highlights presented at the conference and as such, is a “state-of-the-field” for NF research in 2012.

Focal peri-ictal abnormalities on cerebral imaging are well described [1], most commonly mimicking tumours [2, 5]. More extensive abnormalities resembling cerebral ischaemia are more rarely described. Here we describe a case of extensive peri-ictal cerebral oedema. A 37 year-old man with a known craniopharyngioma and secondary epilepsy presented following a cluster of tonic-clonic seizures occurring over the course of a day. At the time of admission he was experiencing generalised tonic-clonic seizures every 10 min. He was apyrexial and normotensive. Neurological examination showed rightsided hemiparesis, right hemianopia and dysphasia. His white blood cell count and C reactive protein were normal. Treatment with phenytoin and lorazepam resulted in resolution of his seizures. He had been diagnosed with craniopharyngioma at the age of 20 when he presented with a 4-year history of partial seizures manifesting as vacant episodes or right arm focal motor jerks. MRI scan at the time revealed the tumour to be compressing the left temporal and frontal lobes, and engulfing but not compromising the left middle cerebral artery (MCA). It was thought that the tumour was not resectable and his epilepsy was treated with sodium valproate and topiramate. He continued to experience monthly tonic-clonic seizures with partial seizures occurring at least weekly. On admission following his seizure cluster, CT scan of the head showed low attenuation of grey and white matter in the left hemisphere with the craniopharyngioma unchanged from previous scans (Fig. 1a–c). The appearances were suggestive of oedema and there was concern that it might have been of ischaemic etiology caused by tumour invasion into the left MCA. However, CT angiogram of the intracranial vessels demonstrated patent arteries in both hemispheres. An MRI scan on day 4 demonstrated oedema in the posterior frontal, parietal and occipital lobes, mainly affecting the cortical grey matter (Fig. 1d,e). Diffusion weighted imaging (DWI) revealed increased signal intensity in the same distribution; there was no evidence of acute restricted diffusion abnormality (Fig. 1f). The patient’s neurological deficit gradually resolved over the following 2 weeks. An MRI scan performed 5 months later showed complete resolution of oedema (Fig. 2), and his seizure frequency improved such that he only experienced very occasional complex partial seizures. Brain oedema is well described following prolonged epileptic seizures, including partial status [6, 8, 9], nonconvulsive status [3] and convulsive status [1, 7, 8]. The focal changes on imaging can imitate a tumour [5, 9] or ischaemic lesion [9]. Indeed, the focality of the changes has been suggested to be a useful marker for seizure focus [7]. Our case is unusual in having extensive oedematous involvement of one hemisphere. The changes evident on imaging are thought to be due to a combination of vasogenic and cytotoxic oedema due C. Carroll (&) C. O. Hanemann Department of Clinical Neurobiology, Peninsula Medical School, The John Bull Building, Tamar Science Park, Plymouth PL6 8BU, UK e-mail: camille.carroll@pms.ac.uk

Advances in molecular biology have resulted in novel therapy for neurofibromatosis 2-related (NF2) tumours, highlighting the need for robust outcome measures. The disease-focused NF2 impact on quality of life (NFTI-QOL) patient questionnaire was assessed as an outcome measure for treatment in a multi-centre study. NFTI-QOL was related to clinician-rated severity (ClinSev) and genetic severity (GenSev) over repeated visits. Data were evaluated for 288 NF2 patients (n = 464 visits) attending the English national NF2 clinics from 2010 to 2012. The male-to-female ratio was equal and the mean age was 42.2 (SD 17.8) years. The analysis included NFTI-QOL eight-item score, ClinSev graded as mild, moderate, or severe, and GenSev as a rank order of the number of NF2 mutations (graded as mild, moderate, severe). The mean (SD) 8.7 (5.4) score for NFTI-QOL for either a first visit or all visits 9.2 (5.4) was similar to the published norm of 9.4 (5.5), with no significant relationships with age or gender. NFTI-QOL internal reliability was good, with a Cronbach’s alpha score of 0.85 and test re-test reliability r = 0.84. NFTI related to ClinSev (r = 0.41, p < 0.001; r = 0.46 for all visits), but weakly to GenSev (r = 0.16, p < 0.05; r = 0.15 for all visits). ClinSev related to GenSev (r = 0.41, p < 0.001; r = 0.42 for all visits). NFTI-QOL showed a good reliability and ability to detect significant longitudinal changes in the QOL of individuals. The moderate relationships of NFTI-QOL with clinician- and genetic-rated severity suggest that NFTI-QOL taps into NF2 patient experiences that are not encompassed by ClinSev rating or genotype.

The neurofibromatoses (NF)--including neurofibromatosis 1 (NF1), neurofibromatosis 2 (NF2), and schwannomatosis--are related tumor-suppressor syndromes characterized by a predisposition to multiple tumor types and other disease manifestations, which often result in functional disability, reduced quality of life, pain, and, in some cases, malignancy. With increasing knowledge of the biology and pathogenesis of NF, clinical trials with targeted agents directed at NF tumors have become available. Most clinical trials for patients with NF have used designs and endpoints similar to oncology trials. However, differences in the disease manifestations and natural history of NF (compared to cancers) require the development of new designs and endpoints to perform meaningful NF clinical trials. The Response Evaluation in Neurofibromatosis and Schwannomatosis (REiNS) International Collaboration was established in 2011 at the Children's Tumor Foundation meeting to achieve consensus within the NF community about the design of future clinical trials, with a specific emphasis on endpoints. The REiNS Collaboration includes 7 working groups that focus on imaging of tumor response; functional, visual, patient-reported, and neurocognitive outcomes; whole-body MRI; and disease biomarkers. This supplement includes the first series of recommendations by the REiNS Collaboration. The hope is that these recommendations will be used by members of the group and by researchers outside of the REiNS International Collaboration to standardize the measurement of outcomes and thus improve clinical trials for patients with NF. Ultimately, we plan to engage industry partners and national regulatory agencies in this process to facilitate the approval of drugs for patients with NF.

The hereditary neuropathy Charcot–Marie–Tooth (CMT) type 1A is, in the majority of cases, caused by duplication of the gene for the peripheral myelin protein PMP22, which leads to abnormally increased PMP22 expression. Recent in vitro and in vivo data indicate a novel function of PMP22 in Schwann-cell growth and differentiation other than its role in myelination, and suggest that over-production of PMP22 leads to a new Schwann-cell phenotype in CMT1A. Taking these data into account, we developed a new hypothesis on the pathogenesis of CMT1A neuropathy: that the defective myelin stability and turnover observed in the disease is caused by altered PMP22 gene dosage and its resultant effect on abnormal Schwann-cell growth and differentiation.

SummaryIt is currently unclear whether Merlin/NF2 suppresses tumorigenesis by activating upstream components of the Hippo pathway at the plasma membrane or by inhibiting the E3 ubiquitin ligase CRL4DCAF1 in the nucleus. We found that derepressed CRL4DCAF1 promotes YAP- and TEAD-dependent transcription by ubiquitylating and, thereby, inhibiting Lats1 and 2 in the nucleus. Genetic epistasis experiments and analysis of tumor-derived missense mutations indicate that this signaling connection sustains the oncogenicity of Merlin-deficient tumor cells. Analysis of clinical samples confirms that this pathway operates in NF2-mutant tumors. We conclude that derepressed CRL4DCAF1 promotes activation of YAP by inhibiting Lats1 and 2 in the nucleus.

Loss of the Merlin tumour suppressor causes abnormal de-differentiation and proliferation of Schwann cells and formation of schwannoma tumours in patients with neurofibromatosis type 2. Within the mature peripheral nerve the normal development, differentiation and maintenance of myelinating and non-myelinating Schwann cells is regulated by a network of transcription factors that include SOX10, OCT6 (now known as POU3F1), NFATC4 and KROX20 (also known as Egr2). We have examined for the first time how their regulation of Schwann cell development is disrupted in primary human schwannoma cells. We find that induction of both KROX20 and OCT6 is impaired, whereas enforced expression of KROX20 drives both myelin gene expression and cell cycle arrest in Merlin-null cells. Importantly, we show that human schwannoma cells have reduced expression of SOX10 protein and messenger RNA. Analysis of mouse SOX10-null Schwann cells shows they display many of the characteristics of human schwannoma cells, including increased expression of platelet derived growth factor receptor beta (PDGFRB) messenger RNA and protein, enhanced proliferation, increased focal adhesions and schwannoma-like morphology. Correspondingly, reintroduction of SOX10 into human Merlin-null cells restores the ability of these cells to induce KROX20 and myelin protein zero (MPZ), localizes NFATC4 to the nucleus, reduces cell proliferation and suppresses PDGFRB expression. Thus, we propose that loss of the SOX10 protein, which is vital for normal Schwann cell development, is also key to the pathology of Merlin-null schwannoma tumours.

The autosomal dominant disorder neurofibromatosis type 2 (NF2) is a hereditary tumor syndrome caused by inactivation of the NF2 tumor suppressor gene, encoding merlin. Apart from tumors affecting the peripheral and central nervous systems, most NF2 patients develop peripheral neuropathies. This peripheral nerve disease can occur in the absence of nerve-damaging tumors, suggesting an etiology that is independent of gross tumor burden. We discovered that merlin isoform 2 (merlin-iso2) has a specific function in maintaining axonal integrity and propose that reduced axonal NF2 gene dosage leads to NF2-associated polyneuropathy. We identified a merlin-iso2-dependent complex that promotes activation of the GTPase RhoA, enabling downstream Rho-associated kinase to promote neurofilament heavy chain phosphorylation. Merlin-iso2-deficient mice exhibited impaired locomotor capacities, delayed sensory reactions and electrophysiological signs of axonal neuropathy. Sciatic nerves from these mice and sural nerve biopsies from NF2 patients revealed reduced phosphorylation of the neurofilament H subunit, decreased interfilament spacings and irregularly shaped axons.

Integrin-mediated activation of PAK (p21-activated kinase) causes phosphorylation and inactivation of the FERM (4.1, ezrin, radixin, moesin) domain-containing protein Merlin, which is encoded by the NF2 (neurofibromatosis type 2) tumor suppressor gene. Conversely, cadherin engagement inactivates PAK, thus leading to accumulation of unphosphorylated Merlin. Current models imply that Merlin inhibits cell proliferation by inhibiting mitogenic signaling at or near the plasma membrane. We have recently shown that the unphosphorylated, growth-inhibiting form of Merlin accumulates in the nucleus and binds to the E3 ubiquitin ligase CRL4(DCAF1) to suppress its activity. Depletion of DCAF1 blocks the hyperproliferation caused by inactivation of Merlin. Conversely, expression of a Merlin-insensitive DCAF1 mutant counteracts the antimitogenic effect of Merlin. Expression of Merlin or silencing of DCAF1 in Nf2-deficient cells induce an overlapping, tumor-suppressive program of gene expression. Mutations present in some tumors from NF2 patients disrupt Merlin's ability to interact with or inhibit CRL4(DCAF1). Lastly, depletion of DCAF1 inhibits the hyperproliferation of Schwannoma cells isolated from NF2 patients and suppresses the oncogenic potential of Merlin-deficient tumor cell lines. Current studies are aimed at identifying the substrates and mechanism of action of CRL4(DCAF1) and examining its role in NF2-dependent tumorigenesis in mouse models. We propose that Merlin mediates contact inhibition and suppresses tumorigenesis by translocating to the nucleus to inhibit CRL4(DCAF1).

Integrin-mediated activation of PAK (p21-activated kinase) causes phosphorylation and inactivation of the FERM (4.1, ezrin, radixin, moesin) domain–containing protein Merlin, which is encoded by the NF2 ( neurofibromatosis type 2 ) tumor suppressor gene. Conversely, cadherin engagement inactivates PAK, thus leading to accumulation of unphosphorylated Merlin. Current models imply that Merlin inhibits cell proliferation by inhibiting mitogenic signaling at or near the plasma membrane. We have recently shown that the unphosphorylated, growth-inhibiting form of Merlin accumulates in the nucleus and binds to the E3 ubiquitin ligase CRL4 DCAF1 to suppress its activity. Depletion of DCAF1 blocks the hyperproliferation caused by inactivation of Merlin. Conversely, expression of a Merlin-insensitive DCAF1 mutant counteracts the antimitogenic effect of Merlin. Expression of Merlin or silencing of DCAF1 in Nf2- deficient cells induce an overlapping, tumor-suppressive program of gene expression. Mutations present in some tumors from NF2 patients disrupt Merlin’s ability to interact with or inhibit CRL4 DCAF1 . Lastly, depletion of DCAF1 inhibits the hyperproliferation of Schwannoma cells isolated from NF2 patients and suppresses the oncogenic potential of Merlin-deficient tumor cell lines. Current studies are aimed at identifying the substrates and mechanism of action of CRL4 DCAF1 and examining its role in NF2-dependent tumorigenesis in mouse models. We propose that Merlin mediates contact inhibition and suppresses tumorigenesis by translocating to the nucleus to inhibit CRL4 DCAF1 .

Cannabinoids have been shown to have a beneficial effect in both animal models of multiple sclerosis (MS) and human disease, although the mechanisms of action are unclear. We examined expression of the major cannabinoid receptors [(CBRs) cannabinoid receptor 1 (CB1) and cannabinoid receptor 2 (CB2)] and a key enzyme involved in synthesis of the endocannabinoid anandamide [N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD)] in autopsy brain samples from patients with MS. CB1 was expressed in neurons, injured axons, oligodendrocytes, macrophages/microglia, some astrocytes, endothelial cells, smooth muscle cells and pericytes. CB2 and NAPE-PLD were localized to cerebral endothelial cells, pericytes, smooth muscle cells, astrocytes and macrophages/microglia. NAPE-PLD immunoreactivity was also seen in neurons. Endothelial CB2 expression was greatest in chronic inactive plaques, and in areas was seen in segments of endothelium where the endothelial expression of adhesion molecules (VCAM-1 and ICAM-1) was focally undetectable, and was often expressed in areas of blood-brain barrier damage. Vascular density was increased in chronic active plaques and normal-appearing white matter compared with controls. These data support findings from animal models which suggest a role for the endocannabinoid system in the MS, particularly in the regulation of endothelial leukocyte adhesion and the cellular response to injury.

This review focuses on altered signaling pathways in schwannoma and identification of molecular markers. Platelet derived growth factors PDGFR and ErbB family of receptor kinase are overexpressed in the schwannoma. PDGFR-β inhibitors with a specific inhibitor AG1296 completely inhibited both basal and PDGF mediated proliferation of human scwannoma cells. Sorafenib, a PDGFR inhibitor currently applied for advanced renal cancer treatment also effectively reducing tyrosine kinase activity and decreased cell proliferation in human schwannoma cells at low concentration. This suggests that this drug could be a promising tool in the treatment of schwannoma.

Angiogenesis plays rate limiting roles in tumor growth and invasion. Angiogenesis inhibition has been proposed as a general strategy to fight against cancers. This book covers different therapeutic targets for angiogenesis interventions with emphasis on clinical development of antiangiogenic drugs that target antiangiogenic cascade in different forms of cancers like breast cancer, multiple myeloma, renal carcinoma, gastrointestinal, prostate and lung cancer. Surrogate predictive markers of antiangiogenic therapy in novel preclinical models recapitulating tumor microenvironment together with genomic studies are special features of this book. Gene therapy approaches with emphasis on viral vectors have been explored in preclinical model with direct and indirect angiogenesis inhibitors. Recent advances in clinical application with antiangiogenic therapy resulted in encouraging clinical results are also detailed. It includes chapters that bridge the gap between basic science and clinical application of current knowledge of cancer. Different chapters are contributed by distinguished scientists and clinicians around the world making it an important addition to the bookshelves of clinicians, academics, researchers, students and institutions.

Charcot-Marie-Tooth disease 1A (CMT1A) is a hereditary demyelinating peripheral neuropathy, associated with a DNA duplication on chromosome 17p11.2. A related disorder in the mouse, trembler (Tr), maps to mouse chromosome 11 which has syntenic homology to human chromosome 17p. Recently, the peripheral myelin protein-22 (pmp-22) gene was identified as the likely Tr locus. We have constructed a partial yeast artificial chromosome contig spanning the CMT1A gene region and mapped the PMP-22 gene to the duplicated region. These observations further implicate PMP-22 as a candidate gene for CMT1A, and suggest that over-expression of this gene may be one mechanism that produces the CMT1A phenotype.

SummaryCurrent models imply that the FERM domain protein Merlin, encoded by the tumor suppressor NF2, inhibits mitogenic signaling at or near the plasma membrane. Here, we show that the closed, growth-inhibitory form of Merlin accumulates in the nucleus, binds to the E3 ubiquitin ligase CRL4DCAF1, and suppresses its activity. Depletion of DCAF1 blocks the promitogenic effect of inactivation of Merlin. Conversely, enforced expression of a Merlin-insensitive mutant of DCAF1 counteracts the antimitogenic effect of Merlin. Re-expression of Merlin and silencing of DCAF1 implement a similar, tumor-suppressive program of gene expression. Tumor-derived mutations invariably disrupt Merlin's ability to interact with or inhibit CRL4DCAF1. Finally, depletion of DCAF1 inhibits the hyperproliferation of Schwannoma cells from NF2 patients and suppresses the oncogenic potential of Merlin-deficient tumor cell lines. We propose that Merlin suppresses tumorigenesis by translocating to the nucleus to inhibit CRL4DCAF1.