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Objective This study aims to assess the feasibility and reliability of our endoscopic trans-nasal technique for the repair of cribriform and sellar high-flow cerebrospinal fluid (CSF) leaks. Methods A comparison between patients suffering from high-flow rhinorrhea and treated through a free grafting endoscopic technique or the “parachute” technique, our nasal packing proposal, was performed. Results Thirty-three patients were included. The mean age was 52 years (range: 36–68 years). The etiology of the CSF leaks was iatrogenic in 16 cases (48.5%), traumatic in 5 cases (15.2%), spontaneous in 11 cases (33.3%), and related to anterior skull base tumors in 1 case (3%). The bone defect affected the sphenoidal sinus in 20 cases (60.6%), the cribriform plate of the ethmoid in 10 cases (30.3%), and both the sphenoid and ethmoid in 3 cases (9.1%). The mean size of bone defects was 8.5 ± 3.9 mm. The median follow-up was 28 (64) months. A CSF leak recurrence occurred in no cases treated with the parachute technique and in 3 cases that underwent conventional endoscopic treatments. The CSF leak recurrences were associated with 2 iatrogenic and 1 post-traumatic fistula. All the CSF leak recurrences underwent the parachute technique, not showing second recurrences. Conclusions Our results suggest that the parachute technique is simple, safe, and effective. We recommend it as an alternative treatment to vascular flaps for the treatment of high-flow and recurrent fistulas.

Gliomas are brain tumors that are treated with surgical resection. Prognosis is influenced by the extent of resection and postoperative neurological status. As consequence, given the extreme interindividual and interhemispheric variability of subcortical white matter (WM) surgical planning requires to be patient's tailored. According to the “connectionist model,” there is a huge variability among both cortical areas and subcortical WM in all human beings, and it is known that brain is able to reorganize itself and to adapt to WM lesions. Brain magnetic resonance imaging diffusion tensor imaging (DTI) tractography allows visualization of WM bundles. Nowadays DTI tractography is widely available in the clinical setting for presurgical planning. Arcuate fasciculus (AF) is a long WM bundle that connects the Broca's and Wernicke's regions with a complex anatomical architecture and important role in language functions. Thus, its preservation is important for the postoperative outcome, and DTI tractography is usually performed for planning surgery within the language-dominant hemisphere. High variability among individuals and an asymmetrical pattern has been reported for this WM bundle. However, the functional relevance of AF in the contralateral non-dominant hemisphere in case of tumoral or surgical lesion of the language-dominant AF is unclear. This review focuses on AF anatomy with special attention to its asymmetry in both normal and pathological conditions and how it may be explored with preoperative tools for planning surgery on gliomas in language areas. Based on the findings available in literature, we finally speculate about the potential role of preoperative evaluation of the WM contralateral to the surgical site.

Glioblastoma is an extremely heterogeneous disease. Treatment failure and tumor recurrence primarily reflect the presence in the tumor core (TC) of the glioma stem cells (GSCs), and secondly the contribution, still to be defined, of the peritumoral brain zone (PBZ). Using the array-CGH platform, we deepened the genomic knowledge about the different components of GBM and we identified new specific biomarkers useful for new therapies. We firstly investigated the genomic profile of 20 TCs of GBM, then, for 14 cases and 7 cases, respectively, we compared these genomic profiles with those of the related GSC cultures and PBZ biopsies. The analysis on 20 TCs confirmed the intertumoral heterogeneity and a high percentage of copy number alterations (CNAs) in GBM canonical pathways. Comparing the genomic profiles of 14 TC-GSC pairs, we evidenced a robust similarity among the two samples of each patient. The shared imbalanced genes are related to the development and progression of cancer and in metabolic pathways, as shown by bioinformatic analysis using DAVID. Finally, the comparison between 7 TC-PBZ pairs leads to identifying PBZ-unique alterations, which it has been identified, require further investigation.

Introduction: Glymphatic fluid circulation may be considered the lymphatic system of the brain and the main role of such system seems to be played by aquaporins (AQPs), a family of proteins which regulates water exchange, in particular AQP4 and 1. Alterations of glymphatic fluid circulation through AQPs variations are now emerging as central elements in the pathophysiology of different brain conditions, like hydrocephalus. This systematic review provides an insight about the role of AQPs in hydrocephalus establishment and compensation, investigating their possible role as diagnostic tools or therapeutic targets. Methods: PubMed database was screened searching for the relevant existing literature in English language published until February 29th 2020, according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) Statement. Results: A total of 40 articles met the inclusion criteria for our systematic analysis. AQP4 resulted the most studied water channel, followed by AQP1. The changes in cerebrospinal fluid (CSF), brain parenchyma and choroid plexus (CP) in different hydrocephalus type were analyzed. Moreover, important pharmacological interactions regarding AQP and molecules or conditions were discussed. A very interesting result is the general consensus on increase of AQP4 in hydrocephalic patients, unless in patients suffering from idiopathic normal pressure hydrocephalus, where AQP4 shows a tendency in reduction. Conclusion: AQP seem to play a central role in the pathophysiology of hydrocephalus and in its compensation mechanisms. Further studies are required to definitively establish their precise roles and their quantitative changes to allow their utilization as diagnostic tools or therapeutic targets.

The study of glioblastoma genomic profiles permits the use of CNAs load to identify patients with poor prognosis and the understanding of the genomic signatures essential for the disease maintenance and the identification of new potential biomarkers.

Recent evidences suggest the involvement of the Vacuolar H+ ATPase (V-ATPase) in the development and/or progression of Glioblastoma Multiforme (GBM). This proton pump could be a valid therapeutic target but more in-depth studies are necessary. The aim of this study is to better define the in vitro effects on Glioma Stem Cell (GSC) primary cultures viability of single and combined treatment with Bafilomycin-A1 (Baf-A1), a V-ATPase inhibitor, and Temozolomide (TMZ), the chemotherapeutic agent currently used to treat GBM patients. We found out that GSC were resistant to TMZ and more sensitive to treatments with Baf-A1 and that the two drugs exerted a synergistic effect when administered together.

Background The transsphenoidal approach is considered the gold standard for resection of pituitary adenomas and other sellar region lesions. This approach is guided by a few fundamental anatomic landmarks that conduct the surgeon toward the sellar floor. Some anatomic structures may vary a lot (e.g., intrasphenoidal septa, intercarotid distance) and may be difficult to identify. Pneumatization and conformation of the sphenoidal sinus (SS) plays a key role in accessing the floor of the sella and other skull base structures. A poorly pneumatized SS may be a relative contraindication to the transsphenoidal approach. We analyzed outcome and complications in transsphenoidal surgery for sellar lesions with a difficult SS. Methods We analyzed 243 consecutive patients who underwent a transsphenoidal approach for sellar lesions. Patients with poor pneumatization of the SS were included. Neurosurgical and endocrinologic outcomes were reported. Results Successful treatment using a transsphenoidal approach with neuronavigation and Doppler ultrasound was achieved in 15 patients with a low degree of pneumatization of the SS. A pituitary adenoma was present in 13 of 15 patients. Endocrinologic and neurosurgical outcomes were similar to patients with normal pneumatization of the SS, showing a cure of disease in 6 of 9 patients with functioning adenomas and an improvement of symptoms in cases of nonfunctioning adenomas. Conclusions Patients with a poorly pneumatized SS can be treated safely with a transsphenoidal approach using image guidance techniques to avoid major neurovascular complications.

Background and purpose: Glioblastoma (GBM) is the most aggressive and frequent subtype of all malignant gliomas. At the time of recurrence, therapeutic options are lacking. Ortataxel, a second-generation taxane was reported to be effective in pre-clinical and phase I clinical studies. The aim of this study was to evaluate a potential therapeutic activity of ortataxel in patients with GBM recurring after surgery and first line treatment. Methods: In this phase II study, according to a two stage design, adult patients with histologically confirmed GBM in recurrence after surgery or biopsy, standard radiotherapy and chemotherapy with temozolomide were considered eligible. Patients included were treated with ortataxel 75mg/m2 i.v. every 3weeks until disease progression. The primary objective of the study was to evaluate the activity of ortataxel in terms of progression free survival (PFS) at 6months after the enrollment. PFS, overall survival at 9months after the enrollment, objective response rate, compliance and safety were evaluated as secondary endpoints. Results: Between Nov 26, 2013 and Dec 12, 2015, 40 patients were recruited across six centres. The number of patients alive and free from progression at 6months after the enrollment, observed in the first stage was four (11.4%), out of 35 patients included in the analysis, below the minimum number of events (7 out of 33) required to continue the study with the second stage The most important toxicities were neutropenia and hepatotoxicity that occurred in 13.2% of patients and leukopenia that occurred in 15.8% of patients. Conclusion: Overall ortataxel treatment fail to demonstrate a significant activity in recurrent GBM patients. However in a limited number of patients the drug produced a benefit that lasted for a long time. Trial registration: This study is registered with ClinicalTrials.gov, number NCT01989884.

Cerebrospinal fluid (CSF) disorders are challenging conditions in neurosurgical practice. The majority of CSF is contained in the basal cisterns of the brain, which are subarachnoid compartments that communicate with each other, and contribute to the circulation of CSF. Yasargil et al. (1976) was the first to provide the systematic classification and naming of the basal cisterns. The lamina terminalis (LT) starts from the gyrus rectus and descends to the lateral aspect of the optic chiasm. It is a thick arachnoidal membrane delineating the anterior wall of the third ventricle that borders the LT cistern. With the introduction of the operating microscope and the progressive development of modern neurosurgery, the arachnoid and basal cisterns have been used as surgical corridors in order to reach deep areas of the brain and to release CSF for brain relaxation. In this way, the LT is used as a surgical corridor for the treatment of several conditions such as obstructive hydrocephalus and diencephalic tumors. In this chapter, we will describe the anatomy of the LT, possible conditions treated by opening the LT, the different surgical approaches to opening the LT, along with their advantages and disadvantages.

Traumatic brain injuries (TBIs) are common neurosurgical emergencies that, in many cases, require fast decision-making and surgical management. In particular, the surgical strategy in these cases may unpredictably be modified because traumatic brain lesions may change over time from the first radiological picture to the time of surgery. For this reason, real-time imaging may offer the opportunity to overcome such problems and help neurosurgeons to get a quick management of intraoperative problems related to unexpected evolution of traumatic brain lesions.

Context Treatment of primary spinal syringomyelia is still controversial. Among others, shunting syrinx fluid to the subarachnoid, peritoneal or pleural space has been utilized with varying success. Shunt obstruction, migration, and infection represent the most common complications of these procedures. Findings The authors present the case of an 81-year-old woman who developed an unusual neurological deterioration resembling a subacute posttraumatic ascending myelopathy (SPAM) after the insertion of a syringosubarachnoid shunt for the treatment of slow-growing D10 syringomyelia. Conclusion/clinical relevance To date, no cases of SPAM secondary to the insertion of a syringosubarachnoid shunt for the treatment of syringomyelia have been reported. The potential pathogenesis related to this phenomenon is discussed.

Several hypotheses have been proposed regarding the mechanisms underlying meningioma-related hyperostosis. In this study, we investigated the role of osteoprotegerin (OPG), insulin-like growth factor 1 (IGF-1), endothelin 1 (ET-1), and bone morphogenetic protein (BMP) 2 and 4.A total of 149 patients (39 males and 110 females; mean age, 62 years) who underwent surgery were included. Depending on the relationship with the bone, meningiomas were classified as hyperostotic, osteolytic, infiltrative, or unrelated. Expression of OPG, and IGF-1, ET-1, BMP-2, and BMP-4 was evaluated by tissue microarray analysis of surgical samples.Our series comprised 132 cases of grade I, 14 cases of grade II, and 3 cases of grade III meningiomas, according to the World Health Organization classification. Based on preoperative computed tomography scan, the cases were classified as follows: hyperostotic, n = 11; osteolytic, n = 11; infiltrative, n = 15; unrelated to the bone, n = 108. Four cases were excluded from the statistical analysis. Using receiver operating characteristic curve analysis, we identified a 2% cutoff for the mean value of IGF-1 that discriminated between osteolytic and osteoblastic lesions; cases with a mean IGF-1 expression of2% were classified as osteolytic (P = 0.0046), whereas those with a mean OPG expression of10% were classified as osteolytic (P = 0.048). No other significant relationships were found.Expression of OPG and expression of IGF-1 were found to be associated with the development of hyperostosis. Preliminary findings suggest that hyperostosis can be caused by an overexpression of osteogenic molecules that influence osteoblast/osteoclast activity. Based on our results, further studies on hyperostotic bony tissue in meningiomas are needed to better understand how meningiomas influence bone overproduction.

Patients affected by a high-grade glioma (HGG) have a poor prognosis with a median survival of 12–16 months. Such poor prognosis affects the perception of the remaining life by patients and the neuropsychological status can strongly affect every-day functioning of these patients. Monitoring changes of neuropsychological functioning (NPF) overtime may provide better clinical information and optimize the neuro-oncological management. The aims of our work were (1) to investigate the feasibility of a complex neuropsychological battery in HGG patients before and during follow-up after surgery; (2) to study the neuropsychological profile of patients affected by HGGs and their relation with the disease status (relapse/death) across time after surgery. One hundred two patients who received surgery for HGG between 2011 and 2017 were studied. All clinical data were prospectively recorded. NPF was assessed during the neuro-oncological follow-up through the Milano-Bicocca Battery (MIBIB). Statistical analysis was performed on the neuropsychological results of the tests administered. First, MIBIB proved to be suitable for patients with HGG tumors before and after surgery, and during long-term follow-up; it also showed a cluster structure representative of the principal cognitive domains. Second, we found a steep decline in the neuropsychological profile before death and/or tumor relapse for the 52% of the neuropsychological tests administered. Complex neuropsychological batteries can be administered to HGG patients before and during follow-up after surgery. There is a correlation between neuropsychological deterioration and tumor relapse and/or death, which may reflect a progressive damage to cognitive functions due to tumor infiltration and progression.

Purpose: To present a single-center experience on pituitary tumor apoplexy and a review of literature focusing on predisposing and precipitating factors. Methods: Clinical presentation of our series of cases. Contemporary published literature is also reviewed. Results: The definition of this syndrome has not been consistent although now the majority of authors agree to definite it as an acute condition caused by hemorrhage or infarction of a pre-existing pituitary adenoma. Different predisposing and precipitating factors have been described in literature; among these antithrombotic and anticoagulant drugs, seem to play relevant roles. The clinical cases observed in our clinic confirm these data and suggest a probable association between elderly patients taking anticoagulant therapy and pituitary apoplexy adenoma. Conclusion: Pituitary tumor apoplexy remains a challenging disease in relation to difficulties in correct diagnosis and thus in appropriate treatment. Antithrombotic/anticoagulant therapy may have an important role as precipitating factor. When a pituitary disorder is known, great care should be taken in the prescription of anticoagulant therapy.

Aquaporin4 (AQP4) is a water channel protein located at astrocyte foot processes that plays a role in glymphatic system, a highly organized fluid transport pathway which seems to be involved in Alzheimer's disease (AD) and normal pressure hydrocephalus (NPH) pathophysiology. Cerebrospinal fluid (CSF) AQP4 levels were determined in 11 patients with AD, 10 patients with NPH, and 9 controls. We found significantly reduced AQP4 in AD patients, a trend in reduction in NPH patients, and a correlation between AQP4 and amyloid-β CSF levels. This study indicates the potential role of AQP4 and glymphatic system in neurodegenerative diseases pathophysiology.

Background Constructional apraxia (CA) is a neuropsychological impairment of either basic perceptual and motor abilities or executive functions, in the absence of any kind of motor or perceptual deficit. Considering patients with focal brain tumors, CA is common in left or right parietal and parieto-occipital lesions. In neuropsychology, the Rey-Osterrieth Complex Figure Test (ROCFT; or parallel forms) is commonly used for the assessment of CA. This study stems from a clinical observation of a difficulty with CA tests for the majority of older neurosurgical patients without occipitoparietal lesions. Methods Patients were tested at 3 points: before surgery, 3 months after surgery, and 12 months after surgery. Thirty patients (15 meningiomas and 15 glioblastomas) were studied retrospectively. Results Older patients with focal brain lesions, regardless of the nature of the tumor, performed poorly at CA tests. More than 50% of patients obtained pathologic results at all 3 times considered. Conclusions Our findings suggest that as CA complex tests involve multiple domains, poor results in copy task may reflect a global cognitive deficit of older patients with tumors, without a specific constructional praxis deficit. CA complex tests (such as the ROCFT) do not give significant informations about visuo-constructional abilities.

Surgical treatment of elderly patients with meningioma is has proved to be safe, especially when patients are selected using dedicated surgical scores. These scores take into account tumor size, edema, location and patient's co-morbidities. Neuropsychological functioning (NPF) of this kind of patients has been poorly studied in literature and it is not taken into account by these scores. Aim of our study was to describe the long-term outcome in terms of NPF of elderly patients undergoing surgery. Patients older than 70 years of age affected by intracranial meningioma and selected with the Clinical-Radiological Grading Score were included in our study. Neuropsychological testing was performed using a dedicated battery of tests before surgery, 3 and 12 months after surgery. Clinical, neurological and radiological outcomes were studied as well. Forty-one patients with a median age of 74 years were included in this study. Preoperatively only 1/41 patients showed a normal NPF with all tests scoring normally. Four out of 39 patients showed a complete neuropsychological recovery after 3 months; while 10/37 patients had a complete recovery after 12 months. NPF showed a trend of progressive improvement after surgery. Our study is the first experience reported in literature describing a long term follow-up in elderly patients after surgery for intracranial meningioma. In our series, surgery determined an improvement of NPF over time; especially with a low complication rate related to the selection of patients obtained through the CRGS. Further studies need to be performed in order to understand how brain edema, tumor size, volume and tumor location affect NPF in both short and long term.

Gene expression analysis provides an effective methodology to identify clinically relevant genes implicated in intervertebral disc (IVD) pathology. The analysis of gene profile in mesenchymal stem cells (MSCs) from human herniated IVD (H-IVD) and degenerated IVD (D-IVD) has not yet been investigated. We present in this study a characterization of MSCs isolated from clinically categorized H-IVD and D-IVD disc samples. H-IVD-MSCs and D-IVD-MSCs showed multipotent mesenchymal differentiation ability, expressing positivity for adipogenic, osteogenic, and chondrogenic markers with an immunophenotypical profile representative of MSCs. FACS analyses revealed a higher expression of CD44 in D-IVD-MSCs compared to H-IVD-MSCs. Gene expression profile revealed that most genes under investigation displayed large variations and were not significantly different in the two types of analyzed IVD-MSCs. Conversely, the gene expression of osteopontin (OPN), a protein involved in bone matrix mineralization and extracellular matrix destruction, was found markedly increased (more than 400-fold) in D-IVD-MSCs compared to H-IVD-MSCs. Moreover, the OPN protein expression was detectable only in D-IVD-MSCs, and its levels were directly related with D-IVD severity. These findings suggest that an abnormal expression of OPN in D-IVD-MSCs occurs and plays a pivotal role in the pathophysiological process of human disc degeneration. We speculate that the regulation of the OPN pathway might be a therapeutic target to counteract disc degeneration.

High grade gliomas (HGG) are tumors with a rapidly progressive course and the standard of care consists of surgery and chemo-radiotherapy. Elderly patients with HGG usually have a worse prognosis due to their comorbidities and difficulties in accessing or completing adjuvant treatments. The purpose of our study was to assess the influence of pre-operative factors (MMSE, age, sex, KPS, tumor volume) on the post-operative access to chemo-radiotherapy in the elderly population. In addition, the influence of the access to adjuvant therapies on overall survival (OS) was assessed. We retrospectively reviewed our consecutive case series of 117 elderly patients (≥65 years) with HGG treated in our Institution. All the clinical records regarding age, sex, tumor location, MMSE, KPS, access to adjuvant treatments and OS were analyzed. 72 males and 45 females with a median age of 71 years were analyzed. Adjuvant therapies were considered; concomitant chemo-radiotherapy with standard radiotherapy or hypofractionated radiation regimen. 84 patients had access to adjuvant therapies. Access to therapies was associated with a median age of 71(range 66–80) years, a median MMSE of 26(range 5–30), and a median tumor volume of 24 cm3(range 1–140). The median OS was 13 months for patients who had access to adjuvant therapies and 5 months for patients who did not. In the elderly patients with HGG, the MMSE, age and tumor volume were predictive of post-surgery access to adjuvant treatments. OS was significantly longer in elderly patients with HGG who had access to post-surgery chemo-radiotherapy.

The optic radiation (OR) is a white matter bundle with a very complex anatomy. Its anterior component bends sharply around the tip of the temporal horn, forming the Meyer's loop (ML), the sparing of which during surgery is crucial to preserve visual function. Defining its exact anatomy and accurately identifying its position remain challenging, even with diffusion tensor imaging (DTI) tractography and the most refined tracking procedure. We have developed an alternative tracking technique to detect the ML position.We performed DTI studies in 26 patients undergoing resection of a temporo-parieto-occipital lesion. We then reconstructed the ORs of each patient using 2 techniques (the first developed by our team, the other taken from the literature), using the same tracking software and parameters. We evaluated the accuracy of each technique measuring 3 distances that define the ML position. We created 5 data groups and compared the 2 techniques. Finally, we compared our results with the results from 8 anatomic dissection studies and other tractographic studies.Our findings show that our technique allows a more accurate definition of the ML position. We found a statistically significant (P0.05) difference for all the distances between the 2 techniques; our results resemble those obtained in dissection studies. Our technique is also easy to perform and repeatable.Our tracking technique may be of marked interest for the evaluation and anatomic definition of the ML position, particularly for neurosurgeons approaching the anterior temporal region.

Background: The V-ATPase proton pump controls acidification of intra and extra-cellular milieu in both physiological and pathological conditions. We previously showed that some V-ATPase subunits are enriched in glioma stem cells and in patients with poor survival. In this study, we investigated how expression of a GBM-like V-ATPase pump influences the non-neoplastic brain microenvironment. Methods: Large oncosome (LO) vesicles were isolated from primary glioblastoma (GBM) neurospheres, or from patient sera, and co-cultured with primary neoplastic or non-neoplastic brain cells. LO transcript and protein contents were analyzed by qPCR, immunoblotting and immunogold staining. Activation of pathways in recipient cells was determined at gene and protein expression levels. V-ATPase activity was impaired by Bafilomycin A1 or gene silencing. Findings: GBM neurospheres influence their non-neoplastic microenvironment by delivering the V-ATPase subunit V1G1 and the homeobox genes HOXA7, HOXA10, and POU3F2 to recipient cells via LO. LOs reprogram recipient cells to proliferate, grow as spheres and to migrate. Moreover, LOs are particularly abundant in the circulation of GBM patients with short survival time. Finally, impairment of V-ATPase reduces LOs activity. Interpretation: We identified a novel mechanism adopted by glioma stem cells to promote disease progression via LO-mediated reprogramming of their microenvironment. Our data provide preliminary evidence for future development of LO-based liquid biopsies and suggest a novel potential strategy to contrast glioma progression. Funding Statement: This work was supported by Fondazione Cariplo (2014-1148 to VV) and by the Italian Minister of Health-Ricerca Corrente program 2017 (to SF). Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: Primary culture were obtained from fresh glioma specimens using surgical surplus of neoplastic or non-neoplastic tissue. All patients received a gross total resection of the tumor at Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico (Milan, Italy) and no patient received neoadjuvant therapy. The study was approved by an Institutional Review Board (IRB#275/2013) and signed informed consent was obtained from all enrolled subjects.

Cancer cells use specific V-ATPase subunits to activate oncogenic pathways. We investigated deregulation of V-ATPase and associated signaling in aggressive gliomas. In vivo, we found that V-ATPase subunit expression correlates with glioma growth. Using patients' series, we observed that glioblastoma (GBM) and relapsed gliomas display a specific V-ATPase signature. Such signature resolves the heterogeneous class of IDH-wild type lower-grade gliomas, identifying the patients with worse prognosis. Molecularly, GBM-like V-ATPase expression correlated with switch in epigenetic enzymes and upregulation of homeobox genes. GBM neurospheres influenced the non-neoplastic microenvironment by delivering the V-ATPase subunit V1G1 and homeobox genes HOXA7, HOXA10, and POU3F2 to recipient cells via large oncosomes (LO). LO reprogrammed recipient cells to grow as spheres and to migrate. Moreover, LO were particularly abundant in the circulation of GBM and of patients who died during follow-up. Finally, impairment of V-ATPase reduced LO activity. These data identify a global cell fate change that underlies glioma aggressiveness and suggest new entry points for glioma stratification, follow-up, and therapy. Funding Statement: This work was supported by Fondazione Cariplo (2014-1148 to VV), Fondazione IRCCS Ca’ Granda, and Fondazione INGM Grant in Molecular Medicine 2014 (to VV) and by the Ricerca Corrente program 2017 (to SF). AMS and MF were supported by a Fellowship from the Doctorate School in Molecular and Translational Medicine of Milan University. Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: Animal experiments were carried out in compliance with the institutional guidelines for the care and use of experimental animals (European Directive 2010/63/UE and the Italian law 26/2014), authorized by the Italian Ministry of Health and approved by the Animal Use and Care Committee of the University of Milan. The patient series of the study: The study was approved by an Institutional Review Board (IRB#275/2013) and signed informed consent was obtained from all patients.

Brain metastases from NSCLC are associated with a poor prognosis, and local radiotherapy is the most effective therapeutic strategy. The oncofetal protein IMP3 has been studied extensively, and evidence suggests that its expression is related to shorter overall survival and a more aggressive phenotype in solid malignancies. Here, the prognostic role of IMP3 was investigated in a cohort of patients with NSCLC brain metastases in correlation with survival and tumor histotype. A series of 42 NSCLC brain metastases samples was analyzed by tissue microarray and immunohistochemical staining for IMP3. IMP3 expression was associated with shorter overall survival in the whole series and in subgroups of metastases from non-neuroendocrine pulmonary malignancies and adenocarcinoma metastases. These results indicated that IMP3 is a strong prognostic factor in non-neuroendocrine brain metastases and in particular in patients with adenocarcinoma metastases.

Background The thalamus is a deep-seated and crucial structure for the sensorimotor system. It has been long considered a surgically inaccessible area because of the morbidity associated with surgical resections. Astrocytomas of the thalamus are usually treated with bioptic procedures followed by adjuvant treatments. Intraoperative neurophysiologic monitoring (IONM) allows safe and satisfactory resections of lobar gliomas, but few data are available for thalamic lesions. The aim of this study was to review the outcome of a small series of patients with thalamic astrocytomas that were treated with surgical resection with the aid of IONM. Methods Surgical resection with IONM was performed in 5 patients with thalamic astrocytomas (1 grade I, 1 grade II, 2 grade III, 1 grade IV). Two astrocytomas were in the dominant hemisphere. Preoperative and postoperative neuropsychological assessments were performed in 3 patients. IONM was tailored to the individual patient and consisted of transcranial motor evoked potential monitoring, cortical motor evoked potential monitoring, somatosensory evoked potential monitoring, direct electrical stimulation, electroencephalography, and electrocorticography. Results None of the patients experienced permanent motor deficits; 2 patients had a transient hemiparesis requiring rehabilitation; 1 patient had a transient aphasia, and 1 patient had permanent aphasia. None of the patients had intraoperative seizures, but 1 patient experienced postoperative transient status epilepticus. The extent of resection on postoperative volumetric magnetic resonance imaging was >70% in all cases. Conclusions Surgical resection of thalamic astrocytomas appeared to be effective and relatively safe when guided by IONM. Larger series of patients are required to confirm these preliminary data.

Background Osteopetroses are a heterogeneous group of heritable disorders characterized by increased bone density as the result of defective osteoclast-mediated bone resorption. The autosomal-dominant osteopetrosis type I (ADO-I) is defined by the presence of osteosclerosis involving mainly the skull bones, variably associated with compression of the foramina of cranial nerves and vascular structures, hypertelorism, exophthalmos, and less commonly with hydrocephalus, pseudotumor, and Chiari malformation type I. Case Description We describe an adult patient with ADO-I presenting with an atypical association of clinical manifestations that required a tailored management. On admission, the patient complained about chronic headache, recurrent sinusitis, and postnasal drip. Findings of the examination didn't show clear signs of increased intracranial pressure, whereas imaging studies revealed thickening of the skull bones and an unexpected fistula associated with anterior ethmoidal meningoencephalocele. Some days after endoscopic transnasal closure of the fistula, a severe hypertensive hydrocephalus developed, which required a prompt ventriculoperitoneal shunt placement, complicated by a diffuse subarachnoid hemorrhage. The 6-month follow-up showed complete recovery. Conclusions After reviewing the literature, we can confirm that ours was the second case of an adult ADO-I patient associated with anterior ethmoidal meningoencephalocele, the first one needing a combined treatment of the encephalocele and hydrocephalus. Because ADO-I is a rare disease with a wide spectrum of clinical manifestations, our case can represent a prototype for the future management of similar cases.

Objective Peripheral nerve sheath tumors (PNSTs) are tumors arising from the neural sheath cells. Surgery plays a central role in the management of this disease, with the purpose of obtaining radical tumor's resection and at the same time providing the best outcome. We retrospectively analyzed 53 PNSTs in 42 patients in an attempt to identify some factors that may improve surgical outcome. Material and Methods Clinical, histologic, and imaging data of 42 patients with PNSTs treated at our Institute between 2001 and 2012 were collected and analyzed. We evaluated the outcome 1 month and 6 month after surgery using three clinical parameters (pain, motor deficits, and sensory deficits) in relation to different histotypes, the presence of neurofibromatosis type 1, tumor location, and duration of symptoms before treatment. Results The best functional results were observed in patients having neurofibromas; the worst outcomes were observed in patients with malignant PNSTs. The other factors were not associated with outcome. Conclusion The timing of surgery is the most important predictive factor of surgical outcome, being the only factor that allows to improve the outcome. With the current study, we want to stress the importance of treating PNSTs as soon as possible to provide the best outcome possible.

Endoscopic third ventriculostomy (ETV) is considered the gold standard treatment for obstructive hydrocephalus due to partial or complete obstruction of cerebrospinal fl uid (CSF) ventricular pathways caused by mass lesions. However long-term effi cacy of this procedure remains controversial as treatment of chronic adult hydrocephalus due to stenosis of Sylvian acqueduct [late-onset idiopathic aqueductal stenosis (LIAS)]. The authors describe clinical presentation, diagnostic investigations in patients affected by LIAS, and defi ne their clinical and radiological outcome after ETV. From January 2003 to December 2008, 13 consecutive LIAS patients treated by ETV were retrospectively reviewed. Pre- and post-operative clinical and radiological fi ndings, including conventional and phase-contrast (PC) cine magnetic resonance imaging (MRI) were investigated. ETV was successfully performed in all patients. Patient’s neurological condition improved. No one required a second ETV procedure or shunt implantation. Clinical and radiological results reveal a satisfactory outcome of LIAS patients treated by ETV. At follow-up a clinical improvement could be demonstrated in all cases. Selection criteria of LIAS patients seem to be crucial to obtain satisfactory and long-lasting results. Even in elderly patients with chronic hydrocephalus, ETV can be considered the treatment of choice.

Background The lumbar infusion test (LIT) and tap test (TT) have previously been described for the diagnosis and selection of appropriate surgical candidates in idiopathic normal pressure hydrocephalus (iNPH). Methods We retrospectively reviewed 81 consecutive patients with a clinical diagnosis of iNPH selected for supplementary testing. Clinical evaluation was scored with the Japanese Grading Scale for Normal Pressure Hydrocephalus, the Global Deterioration Score, and the modified Rankin Scale (mRS). The test protocol included a cerebrospinal fluid pressure monitoring (PMi), an LIT, and a TT. Patients were selected for surgery if outflow resistance was ≥14 mm Hg/mL/minute or if a clinical improvement was recorded after TT. Results Sixty-eight patients were selected for ventriculoperitoneal shunting; 72.8% had a positive PMi or LIT, 74.1% had a positive TT, and 63.0% were positive to both tests. Complications were all transient. Clinical evaluation at 12 months after shunting showed a global improvement in 60 patients (88.2%). Overall, 75.0% of patients had no significant disability (mRS score, 1 and 2), 20.6% had an mRS score of 3 or 4, and 4.4% had severe disability after surgery. The positive predictive value of PMi/LIT, TT, or both combined was similar (89.8, 90.0, and 88.2%); however, 21.7% of patients who improved after surgery were selected with either a positive LIT or TT alone. Conclusions LIT and TT are complementary and they can easily be combined in sequence with a low complication rate and high probability of selecting patients with iNPH who may benefit from ventriculoperitoneal shunt surgery.

Familial hemiplegic migraine (FHM) type 1 is a rare monogenic dominant autosomal disease due to CACNA1A gene mutations. Besides the classical phenotype, mutations on CACNA1A gene are associated with a broader spectrum of clinical features including cerebellar ataxia, making FHM1 a complex channelopathy. We report the case of a patient carrying the p.Arg583Gln mutation affected by hemiplegic migraine and late onset ataxia and we performed a literature review about the clinical features of p.Arg583Gln. Although p.Arg583Gln mutations are associated with a heterogeneous phenotype, carriers present cerebellar signs which consisted generally in ataxia and dysmetria, with intention tremor appearing mostly in advanced age, often progressive and permanent. The heterogeneous spectrum of CACNA1A gene mutations probably causes sporadic hemiplegic migraine (SHM) to be misdiagnosed. Given the therapeutic opportunities, SHM/FHM1 should be considered in differential diagnosis of patients with cerebellar ataxia and migraine with aura.

Brain metastasis are the most common neoplastic lesions of the nervous system. Many cancer patients are diagnosed on the basis of a first clinical presentation of cancer on the basis of a single or multiple brain lesions. Brain metastases are manifestations of primary disease progression and often determine a poor prognosis. Not all patients with a brain metastases undergo surgery: many are submitted to alternative or palliative treatments. Management of patients with brain metastases is still controversial, and many studies have been developed to determine which is the best therapy. Furthermore, management of patients operated for a brain metastasis is often difficult. Chemotherapy, stereotactic radiosurgery, panencephalic radiation therapy, and surgery, in combination or alone, are the means most commonly used. We report our experience in the management of a ten-year series of surgical brain metastasis and discuss our results in the preoperative and postoperative management of this complex condition.

Background: Recent evidences highlighted that GBM secreted microvesicles (EVs), particularly exosomes (Exo) and large oncosomes (LO), play a major role in the cross-talk between tumor cell and non-neoplastic parenchyma. How GBMs manage to thrive in a highly unfavorable, acidic microenvironment is still unclear, but recent work from our group has identified the vacuolar pump H+-ATPase (V-ATPase) as an important effector of GBM growth and glioma stem cells (GSC) maintenance. Additionally, in ExoCarta database V-ATPase subunits have been described in Exo from different cancer cell types. Taken together, these data identify V-ATPase as an important driver of gliomagenesis, and a novel, actionable therapeutic target for disease intervention. However, the role of V-ATPase in reprogramming the GBM microenvironment has not been previously investigated. Methods: Exo and LO were isolated by an Invitrogen kit and serial centrifugation, respectively, from media of patients’ derived GBM neurospheres, enriched in GSC (NS, n=12) or differentiated cultures (n=8). For EVs internalization studies, Exo or LO were stained using FM 1-43 FX dye and the process was followed live for 30’ and at selected time points (30’-4h-24h), using a confocal microscopy or flow cytometry (FACS). Electron microscopy, FACS (of Exo stained with CellTrace and SytoRNA in combination with CD63 coated beads), Nanosight and immunoblotting (for CD63, CD9 and Clathrin) analyses were used to confirm EVs subtypes. Cultures from patients’ derived brain tumor margins or primary GBM (differentiated and not) were used as EVs-recipient cells. miRNA profiling was performed using Taqman Low density arrays and analyzed by R packages. Gene Ontology analysis was performed by DAVID. The study was approved by the Institutional Ethical Committee. Results: NS are able to produce different EVs, which are internalized by recipient cells after 4 and up to 24 hours of co-culture. Both Exo than LO from NS are able to significantly increase cell growth in recipient cells (brain tumor margins and primary GMB differentiated monolayers), and this effect is stronger with EVs produced by NS with higher V-ATPase expression (V-ATPaseHIGH NS). Primary GBM cells after co-culture with EVs are able to produce a higher number of NS and V-ATPase activity block by BafilomycinA1 in NS-producing EVs completely revert this effect. Finally, the co-culture of V-ATPaseLOW NS with EVs from V-ATPaseHIGH NS increases their motility in collagen matrixes. At molecular level, profiling of Exo-derived miRNAs distinguishes differentiated cultures from NS and, among NS, V-ATPaseHIGH cultures. In silico analysis and annotation of miRNA target genes from V-ATPaseHIGH–derived Exo showed an enrichment of cancer, cell cycle and PI3K/Akt pathways. Conclusions: Altogether, these data point toward the central role of different EV types in GBM communication and suggest a role of the V-ATPase proton pump in regulating EV’s contents. Citation Format: Irene Bertolini, Andrea Terrasi, Andrea Di Cristofori, Silvano Bosari, Valentina Vaira. V-ATPase control of EV signaling in glioma stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2889. doi:10.1158/1538-7445.AM2017-2889

Morphologic criteria illustrated in WHO guidelines are the most significant prognostic factor in human gliomas, but novel biomarkers are needed to identify patients with a poorer outcome. The present study examined the expression of the oncofetal protein IMP3 in a series of 135 patients affected by high-grade (grade III and IV) gliomas, correlating the results with proliferative activity, molecular parameters, and clinical and follow-up data. Overall, IMP3 expression was higher in glioblastomas (68%) than in grade III tumors (20%,P<0.0001), and IMP3-positive high-grade gliomas showed a shorter overall and disease-free survival than negative ones (P=0.0002andP=0.006, resp.). IMP3 expression was significantly associated with the absence of mutations of IDH1 gene (P=0.0001) and with the unmethylated phenotype of MGMT in high-grade gliomas (P=0.004). High Ki67 levels were correlated with better prognosis in glioblastomas but IMP3 expression was not correlated with the proliferation index. These findings confirm the role of IMP3 as a marker of poor outcome, also in consideration of its association with IDH1 wild-type phenotype and MGMT unmethylated status. The data suggest that IMP3 staining could identify a subgroup of patients with poor prognosis and at risk of recurrence in high-grade gliomas.

INTRODUCTION: Intra-operative neurophysiological monitoring and brain mapping techniques represent nowadays the gold standard for surgical resection of brain tumors. Brain mapping usually requires the exposure of a large area of brain cortex in order to identify the different eloquent sites (e.g. primary motor strip, speech arrest sites). Thus, relatively large classic fronto-temporal craniotomies are generally performed. Nevertheless, in recent years the use of small craniotomies targeted on the tumor has gained popularity thanks to Neuronavigation systems and to the low morbidity rates associated with shorter surgical procedures and reduced anesthestic drug administration. Aim of this study was to review our experience which combines extensive neurophysiological monitoring and small craniotomies. Length of surgical time, intra-operative and post-operative complications, length of stay, extent of resection rate and the neurological outcomes were assessed. PATIENTS AND METHODS: Eighty-five consecutive patients (54 Males, 31 Females; Median Age 55 years, range 12-77) underwent surgery with brain mapping and intra-operative neurophysiological monitoring for resection of a brain tumor in eloquent location under asleep-asleep (62 patients) or asleep-awake (23 patients) anesthesia. Pre- and post-operative assessment included in-depth neuropsychological evaluation and MRI. Tumor histologies were the following: 74 gliomas (60 HGG, 14 LGG), 6 metastases and 5 other tumor histologies (2 cavernomas, 2, meningiomas, 1 AVM). The bone flap was tailored to the size of the tumor; no cortex other than that which was strictly necessary for tumor resection was exposed. The intra-operative neurophysiological monitoring included: EEG, SSEP, transcranial MEP and/or cortical MEP, ECoG; direct electrical stimulation (DES) was performed using bipolar and monopolar probes. RESULTS: Median surgical time was 230 minutes (range 90-390). Intra-operative complications included 9 seizures (3 requiring pharmacological treatment, 6 regressing after cold irrigation) and 5 electrical seizure. Post-operative neurosurgical complications were the following: 8 seizures and 4 re-operation for hematoma. Seventy-three patients (86%) could be discharged home while 12 (14%) were transferred to rehabilitation. In only 3 patients (3.5%) a permanent new neurological deficit could be noted at 3-month follow-up. Median post-operative stay was 7 days (range 3-45). CONCLUSIONS: In our experience a small craniotomy supported by extensive neurophysiological monitoring and brain mapping permitted to achieve satisfactory results in terms of complications, hospitalization, and neurological outcomes.

BACKGROUND: Glioblastoma (GBM) is a primary malignant brain tumor and its evolution is generally rapidly fatal. The current standard of care for this disease is surgical resection followed by concomitant radiationtherapy plus chemiotherapy as described by Stupp and Colleagues. In this study we described our experience in a series of 100 consecutive patients affected by GBM treated with modified Stupp regimen at “Fondazione IRCCS Ca' Granda Ospedale Policlinico Maggiore” in Milan, Italy. METHODS: One hundred patients with newly diagnosed, histologically confirmed GBM were submitted to concomitant chemo-radiation therapy. Subjects who were not able to complete the concomitant phase of chemo-radiation therapy were excluded from this study.The diagnosis was obtained after surgical resection or stereotactic biopsy and then all the patients were treated with radiationtherapy plus continuous daily temozolomide (TMZ) according to a modified Stupp's schedule. In the adjuvant phase we administered a modified schedule consisting of 150mg/mq of TMZ, from day 1 to 5 and 75 mg/mq of TMZ from day 6 to 10 for 12 cycles instead of the 6 standard cycles proposed by Stupp and Colleagues. At recurrence, various strategies were chosen according the patients clinical and radiological status. After the concomitant phase, patients underwent a regular follow-up with a clinical examination and a brain MRI with gadolinium every 3 months. MGMT status was assessed according the method described by Hegi et al. in 2005 and was retrospectively re-evaluated with PCR real time (Pyromark Q96ID System). RESULTS: One hundred patients affected by GBM were included (60 males; 40 females; median age 61). The median KPS at presentation was 70 (range= 40- 100) which remained stable after 6 months. Median Ki67 was 10%. In 63 patients MGMT was methylated (38 males; 25 females. Median TTP-1 was of 12 months) and in 32 patients were MGMT un-methylated (male 21; females11; median TTP-1 of 9 months); in 5 cases we have no data. Surgical resection was done in 92 patients whereas only in 8 cases weproceeded with stereotactic biopsy alone. Gross total resection (>95%) was achieved in 57 patients; while 35 patients receibed a sub-total resection (85-95%).Median Overall survival (OS) was 16 months and TTP-1 and time to tumor progression after recurrence (TTP-2) was 11 months in MGMT methylated patients and 4 months in MGMT un-methylated patients. At a median follow up of 12 months, 24 months and 36 months our OS was 81%, 33% and 20% respectively compared to the historical group described by Stupp in 2005 which were 61% at 12 months and 27.5% at 24 months and 16% at 36 months. Data about the comparison of MGMT status assessment will be discussed at the time of the meeting. CONCLUSION: According the data collected, our modified adjuvant schedule can be useful for increasing the therapeutic effect of TMZ and can contribute to prolong the TTP.

Based on a retrospective study, in a previous report we developed a grading system, called ‘Clinical Radiological Grading System’ (CRGS), in order to standardise surgical indications in patients older than 70 years affected by intracranial meningiomas. Various prognostic factors were taken into consideration: clinical conditions, comorbidities, neurological status, size and size of the lesion and peritumoral edema. We then performed a prospective cross-sectional study including 90 consecutively recruited patients (from 1990 to 2000), 70 years of age or older, affected by meningiomas in whom the decision whether to operate was based on the CRGS score. Our findings showed that patients with a score lower than 10 had a poor prognosis regardless of surgical treatment, those with a score between 10 and 12 had a prognosis positively influenced by surgery, and those with a score higher than 12 had a good prognosis regardless of surgical treatment. A further validation of our scale has been made in a second prospective study, which is discussed in the present chapter. We also analyzed similar studies performed by other authors and compare our grading system to alternative scores which have been created afterwards.

Early and delayed cognitive dysfunctions are an understudied issue after aneurysmal subarachnoid hemorrhage (aSAH), irrespective of neurological outcome. The aim of this study was to describe early and delayed changes of cognitive functions, activities of everyday life, and quality of life in aSAH patients.

BACKGROUND: Patients with high grade gliomas (HGG) have a poor prognosis and experience cognitive deficits which depend on location, size, mass effect and edema of the tumor. Neuropsychological functioning (NPF) can be assessed by a battery of tests which objectivise the cognitive status of the patients. These tests are time-consuming and fatiguing for the patients, nevertheless could give relevant information about the course of the disease. Aim of our work was to describe overtime the NPF of patients affected by HGG and to correlate the NPF scores with tumor recurrence and survival. MATERIAL AND METHODS: We examined the NPF in 102 patients operated for HGG. Patients were followed longitudinally with the Milano-Bicocca Battery for neuropsychological assessment of low grade gliomas. Patients were evaluated before surgery and few days after surgery. Then, neuropsychological and neuro-oncological follow-up were performed every three months. We first explored the correlational structure of our neuropsychological variables through hierarchical clustering in the R package Hmisc. We then investigated the impact on patients’ outcome of the variables selected through hierarchical clustering via non-linear regression, with fixed/random effects. Time to tumor recurrence/death was selected as the main dependent variable. RESULTS: Sixty-seven males and 35 females (median age 59, range 18–81) were included. Starting from a battery of 27 neuropsychological tests, 10 tests could be selected from the statistical analysis (cluster structure, condition number K of 51.16) since they were strictly correlated with the other tests; thus, these 10 tests were used for the further statistical analysis. Pitting patients’ neuropsychological profiles against tumor relapse/death, 5 cognitive tests showed a significant correlation: Token test (p=0.003), Object Naming test (p=0.02), Digit Memory Span (p=0.001), Corsi Block test (p=0.004), Semantic Fluency test (p=0.003). A steep decline for all these tests emerged a few weeks before tumor relapse/death. Analyzing the NPF overtime, all the tests worsened immediately after surgery, recovered steadily within 2–3 months and then remained stable until progression. CONCLUSION: The tests which showed to be more informative about the patients NPF were Token test, Object Naming test, Digit Memory Span, Corsi Block test, Semantic Fluency test. These tests showed a significant decline few weeks before tumor relapse/death.

// Andrea Di Cristofori 1,2,* , Stefano Ferrero 3,4,* , Irene Bertolini 1,3 , Gabriella Gaudioso 1,3 , Maria Veronica Russo 1,3 , Valeria Berno 5 , Marco Vanini 6 , Marco Locatelli 2 , Mario Zavanone 1,2 , Paolo Rampini 2 , Thomas Vaccari 7 , Manuela Caroli 2 and Valentina Vaira 3,5 1 Department of Pathophysiology and Organ Transplantation, University of Milan, Milan, Italy 2 Division of Neurosurgery, Fondazione IRCCS Ca’ Granda, Ospedale Maggiore Policlinico, Milan, Italy 3 Division of Pathology, Fondazione IRCCS Ca’ Granda, Ospedale Maggiore Policlinico, Milan, Italy 4 Department of Biomedical, Surgical and Dental Sciences, University of Milan, Milan, Italy 5 Istituto Nazionale Genetica Molecolare “Romeo ed Enrica Invernizzi”, Milan, Italy 6 Surgical Pathology Unit, St. Anna Hospital, Como, Italy 7 IFOM - The FIRC Institute of Molecular Oncology, Milan, Italy * These authors have equally contributed to this work Correspondence to: Manuela Caroli, email: // Valentina Vaira, email: // Keywords : glioblastoma, vacuolar H+-ATPase, bafilomycin A1, cancer stem cells, organotypic tissue cultures Received : January 16, 2015 Accepted : May 02, 2015 Published : May 22, 2015 Abstract The vacuolar H + ATPase (V-ATPase) is a proton pump responsible for acidification of cellular microenvironments, an activity exploited by tumors to survive, proliferate and resist to therapy. Despite few observations, the role of V-ATPase in human tumorigenesis remains unclear. We investigated the expression of ATP6V0C, ATP6V0A2, encoding two subunits belonging to the V-ATPase V0 sector and ATP6V1C, ATP6V1G1, ATPT6V1G2, ATP6V1G3, which are part of the V1 sector, in series of adult gliomas and in cancer stem cell-enriched neurospheres isolated from glioblastoma (GBM) patients. ATP6V1G1 expression resulted significantly upregulated in tissues of patients with GBM and correlated with shorter patients’ overall survival independent of clinical variables. ATP6V1G1 knockdown in GBM neurospheres hampered sphere-forming ability, induced cell death, and decreased matrix invasion, a phenotype not observed in GBM monolayer cultures. Treating GBM organotypic cultures or neurospheres with the selective V-ATPase inhibitor bafilomycin A1 reproduced the effects of ATP6V1G1 siRNA and strongly suppressed expression of the stem cell markers Nestin, CD133 and transcription factors SALL2 and POU3F2 in neurospheres. These data point to ATP6V1G1 as a novel marker of poor prognosis in GBM patients and identify V-ATPase inhibition as an innovative therapeutic strategy for GBM.