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Background/purpose Glioblastoma (GBM) is the most common primary malignant brain tumor. Sex has been shown to be an important prognostic factor for GBM. The purpose of this study was to develop and independently validate sex-specific nomograms for estimation of individualized GBM survival probabilities using data from 2 independent NRG Oncology clinical trials. Methods This analysis included information on 752 (NRG/RTOG 0525) and 599 (NRG/RTOG 0825) patients with newly diagnosed GBM. The Cox proportional hazard models by sex were developed using NRG/RTOG 0525 and significant variables were identified using a backward selection procedure. The final selected models by sex were then independently validated using NRG/RTOG 0825. Results Final nomograms were built by sex. Age at diagnosis, KPS, MGMT promoter methylation and location of tumor were common significant predictors of survival for both sexes. For both sexes, tumors in the frontal lobes had significantly better survival than tumors of multiple sites. Extent of resection, and use of corticosteroids were significant predictors of survival for males. Conclusions A sex specific nomogram that assesses individualized survival probabilities (6-, 12- and 24-months) for patients with GBM could be more useful than estimation of overall survival as there are factors that differ between males and females. A user friendly online application can be found here—https://npatilshinyappcalculator.shinyapps.io/SexDifferencesInGBM/.

Background Gliomas are the most common type of malignant brain and other CNS tumors, accounting for 80.8% of malignant primary brain and CNS tumors. They cause significant morbidity and mortality. This study investigates the intersection between age and sex to better understand variation of incidence and survival for glioma in the United States. Methods Incidence data from 2000 to 2017 were obtained from CBTRUS, which obtains data from the NPCR and SEER, and survival data from the CDC’s NPCR. Age-adjusted incidence rate ratios (IRR) per 100 000 were generated to compare male-to-female incidence by age group. Cox proportional hazard models were performed by age group, generating hazard ratios to assess male-to-female survival differences. Results Overall, glioma incidence was higher in males. Male-to-female incidence was lowest in ages 0-9 years (IRR: 1.04, 95% CI: 1.01-1.07, P = .003), increasing with age, peaking at 50-59 years (IRR: 1.56, 95% CI: 1.53-1.59, P < .001). Females had worse survival for ages 0-9 (HR: 0.93, 95% CI: 0.87-0.99), though male survival was worse for all other age groups, with the difference highest in those 20-29 years (HR: 1.36, 95% CI: 1.28-1.44). Incidence and survival differences by age and sex also varied by histological subtype of glioma. Conclusions To better understand the variation in glioma incidence and survival, investigating the intersection of age and sex is key. The current work shows that the combined impact of these variables is dependent on glioma subtype. These results contribute to the growing understanding of sex and age differences that impact cancer incidence and survival.

Background:Sex plays an important role in the incidence, prognosis, and mortality of cancers, but often is not considered in disease treatment.Methods:We quantified sex differences in cancer incidence using the United States Cancer Statistics (USCS) public use database and sex differences in cancer survival using Surveillance, Epidemiology, and End Results (SEER) public use data from 2001 to 2016. Age-adjusted male-to-female incidence rate ratios (IRR) with 95% confidence intervals (CI) were generated by primary cancer site, race, and age groups. In addition, age-adjusted hazard ratios with 95% CI by sex within site were generated.Results:In general, cancer incidence and overall survival were lower in males than females, with Kaposi sarcoma (IRR: 9.751; 95% CI, 9.287–10.242; P < 0.001) having highest male-to-female incidence, and thyroid cancers (HR, 1.774; 95% CI, 1.707–1.845) having largest male-to-female survival difference. Asian or Pacific Islanders had particularly high male-to-female incidence in larynx cancers (IRR: 8.199; 95% CI, 7.203–9.363; P < 0.001), relative to other races. Among primary brain tumors, germ cell tumors had the largest male-to-female incidence (IRR: 3.03; 95% CI, 2.798–3.284, P < 0.001).Conclusions:Overall, incidence and survival of cancer vary significantly by sex, with males generally having lower incidence and survival compared with females. Male-to-female incidence differences were also noted across race and age groups. These results provide strong evidence that the fundamental biology of sex differences affects cancers of all types.Impact:This study represents the most recent and comprehensive reporting of sex differences in cancer incidence and survival in the United States. Identifying disadvantaged groups is critical as it can provide useful information to improve cancer survival, as well as to better understand the etiology and pathogenesis of specific cancers.

Significant sex differences exist across cellular, tissue organization, and body system scales to serve the distinct sex-specific functions required for reproduction. They are present in all animals that reproduce sexually and have widespread impacts on normal development, aging, and disease. Observed from the moment of fertilization, sex differences are patterned by sexual differentiation, a lifelong process that involves mechanisms related to sex chromosome complement and the epigenetic and acute activational effects of sex hormones. In this mini-review, we examine evidence for sex differences in cellular responses to DNA damage, their underlying mechanisms, and how they might relate to sex differences in cancer incidence and response to DNA-damaging treatments.

Background Chronotherapy is an innovative approach to improving survival through timed delivery of anti-cancer treatments according to patient daily rhythms. Temozolomide (TMZ) is a standard-of-care chemotherapeutic agent for glioblastoma (GBM). Whether timing of TMZ administration affects GBM patient outcome has not previously been studied. We sought to evaluate maintenance TMZ chronotherapy on GBM patient survival. Methods This retrospective study reviewed patients with newly diagnosed GBM from January 1, 2010 to December 31, 2018 at Washington University School of Medicine who had surgery, chemoradiation, and were prescribed TMZ to be taken in the morning or evening. The Kaplan–Meier method and Cox regression model were used for overall survival (OS) analyses. The propensity score method accounted for potential observational study biases. The restricted mean survival time (RMST) method was performed where the proportional hazard assumption was violated. Results We analyzed 166 eligible GBM patients with a median follow-up of 5.07 years. Patients taking morning TMZ exhibited longer OS compared to evening (median OS, 95% confidence interval [CI] = 1.43, 1.12–1.92 vs 1.13, 0.84–1.58 years) with a significant year 1 RMST difference (−0.09, 95% CI: −0.16 to −0.018). Among MGMT-methylated patients, median OS was 6 months longer for AM patients with significant RMST differences at years 1 (−0.13, 95% CI = −0.24 to −0.019) to 2.5 (−0.43, 95% CI = −0.84 to −0.028). Superiority of morning TMZ at years 1, 2, and 5 (all P < .05) among all patients was supported by RMST difference regression after adjusting for confounders. Conclusions Our study presents preliminary evidence for the benefit of TMZ chronotherapy to GBM patient survival. This impact is more pronounced in MGMT-methylated patients.

Medulloblastoma, a common pediatric malignant central nervous system tumour, represent a small proportion of brain tumours in adults. Previously it has been shown that in adults, Sonic Hedgehog (SHH)-activated tumours predominate, with Wingless-type (WNT) and Group 4 being less common, but molecular risk stratification remains a challenge. We performed an integrated analysis consisting of genome-wide methylation profiling, copy number profiling, somatic nucleotide variants and correlation of clinical variables across a cohort of 191 adult medulloblastoma cases identified through the Medulloblastoma Advanced Genomics International Consortium. We identified 30 WNT, 112 SHH, 6 Group 3, and 41 Group 4 tumours. Patients with SHH tumours were significantly older at diagnosis compared to other subgroups (p < 0.0001). Five-year progression-free survival (PFS) for WNT, SHH, Group 3, and Group 4 tumours was 64.4 (48.0-86.5), 61.9% (51.6-74.2), 80.0% (95% CI 51.6-100.0), and 44.9% (95% CI 28.6-70.7), respectively (p = 0.06). None of the clinical variables (age, sex, metastatic status, extent of resection, chemotherapy, radiotherapy) were associated with subgroup-specific PFS. Survival among patients with SHH tumours was significantly worse for cases with chromosome 3p loss (HR 2.9, 95% CI 1.1-7.6; p = 0.02), chromosome 10q loss (HR 4.6, 95% CI 2.3-9.4; p < 0.0001), chromosome 17p loss (HR 2.3, 95% CI 1.1-4.8; p = 0.02), and PTCH1 mutations (HR 2.6, 95% CI 1.1-6.2; p = 0.04). The prognostic significance of 3p loss and 10q loss persisted in multivariable regression models. For Group 4 tumours, chromosome 8 loss was strongly associated with improved survival, which was validated in a non-overlapping cohort (combined cohort HR 0.2, 95% CI 0.1-0.7; p = 0.007). Unlike in pediatric medulloblastoma, whole chromosome 11 loss in Group 4 and chromosome 14q loss in SHH was not associated with improved survival, where MYCN, GLI2 and MYC amplification were rare. In sum, we report unique subgroup-specific cytogenetic features of adult medulloblastoma, which are distinct from those in younger patients, and correlate with survival disparities. Our findings suggest that clinical trials that incorporate new strategies tailored to high-risk adult medulloblastoma patients are urgently needed.

Diffuse intrinsic pontine glioma (DIPG) arising in the brainstem is the deadliest pediatric brain cancer with nearly 100% fatality and a median survival of

The tumor microenvironment in glioblastoma provides cancer cells with favorable conditions to proliferate and invade surrounding tissues. Macrophages comprise a large portion of the glioblastoma tumor microenvironment (TME) both in terms of volume and function. These cells have been reported to influence tumor progression by modulating immune responses, remodeling extracellular matrix, and providing nutrients to cancer cells among numerous other functions. Radiation therapy forms one of the pillars of glioblastoma management along with surgical resection and chemotherapy. Here we investigated the effects of radiation on macrophage iron metabolism. Using mouse bone-marrow-derived macrophages (BMDMs) we performed in-vitro 59Fe radiotracer assays to study how radiation exposure modified iron trafficking in these cells. We found that low dose radiation at 0.25, 0.5, or 2 Gy from a 60Co source stimulated iron release from the BMDMs with maximal release occurring at 0.5 Gy. Moreover, we observed that iron release was dependent on the amount of serum present in culture media with cells cultured in 20% fetal bovine serum (FBS) showing reduced iron release profiles compared to those cultured in 10% or 1% FBS. Since glioblastoma patients exhibit sexually dimorphic survival outcomes, we investigated whether these radiation-induced responses occurred in a sexually dimorphic pattern. At radiation doses of 0.25 Gy we observed that male macrophages tended to release more iron than female macrophages despite no differences in iron uptake between the sexes – raising the question as to whether differential iron trafficking in response to treatment contributes to the poorer survival outcomes observed in males. Our data suggest that delineating how supporting cells such as macrophages respond to glioblastoma treatment regimens may provide insights into addressing mechanisms of treatment resistance and further our understanding of the sexual dimorphism observed in patient outcomes.

OBJECTIVE Glioblastoma (GBM) is the most common malignant primary brain tumor in adults, with males more commonly affected than females(1.6:1). Despite advancements in treatments, prognosis is dismal with a median overall survival of 15 months. Our aim was to investigate sex as a variable in GBM patient survival after receiving incremental levels of standard-of-care treatment regimens – different extents of surgical resection and different numbers of cycles of adjuvant temozolomide chemotherapy. METHODS Drawing from our extensive multi-institutional brain tumor repository, we investigated GBM subjects with overall survival (OS), extent of resection (EOR), number of temozolomide (TMZ) cycles, and sex data (n=620, males: n=387, females: n=233). Cox proportional hazard ratios were computed to investigate the multivariable predictive value of the patient variables with OS. Patients were then divided into groups based on their sex, EOR (either biopsy, subtotal resection (STR) or gross total resection (GTR)), and TMZ cycles (I: < 6 cycles, II: 7-11 cycles and III: >12 cycles). RESULTS We observed that STR was beneficial for females (HR=0.52; CI=0.33-0.83; p-value=0.013), while for males the benefit was not detected (HR=0.73; CI=0.46-1.15; p-value=0.173) for STR but was detectable for GTR (HR=0.58, CI=0.37-0.90; p-value=0.014). Females receiving 7-11 cycles of TMZ showed a survival benefit (HR=0.52; CI=0.12-0.53; p-value=0.048) while males in the same group did not (HR=0.74; CI=0.46-1.19; p-value=0.21), in comparison to those in group I of TMZ cycles. No sex differences were identified in patients receiving < =6 cycles or >=12 cycles. CONCLUSION Together, our results contribute to the growing literature that sex differences exist in GBM patients, even in response to standard-of-care therapies. This should be accounted for when designing clinical trials for GBM so that we may advance our pursuit to deliver personalized medicine.

Background: High-grade pediatric brain tumors exhibit the highest cancer mortality rates in children. While conventional MRI has been widely adopted for examining pediatric high-grade brain tumors clinically, accurate neuroimaging detection and differentiation of tumor histopathology for improved diagnosis, surgical planning, and treatment evaluation, remains an unmet need in their clinical management. Methods: We employed a novel Diffusion Histology Imaging (DHI) approach employing diffusion basis spectrum imaging (DBSI) derived metrics as the input classifiers for deep neural network analysis. DHI aims to detect, differentiate, and quantify heterogeneous areas in pediatric high-grade brain tumors, which include normal white matter (WM), densely cellular tumor, less densely cellular tumor, infiltrating edge, necrosis, and hemorrhage. Distinct diffusion metric combination would thus indicate the unique distributions of each distinct tumor histology features. Results: DHI, by incorporating DBSI metrics and the deep neural network algorithm, classified pediatric tumor histology with an overall accuracy of 83.3%. Receiver operating analysis (ROC) analysis suggested DHI’s great capability in distinguishing individual tumor histology with AUC values (95%CI) of 0.983 (0.985 - 0.989), 0.961 (0.957 - 0.964), 0.993 (0.992 - 0.994), 0.953 (0.947 - 0.958), 0.974 (0.970 - 0.978) and 0.980 (0.977 - 0.983) for normal WM, densely cellular tumor, less densely cellular tumor, infiltrating edge, necrosis and hemorrhage, respectively. Conclusions: Our results suggest that DBSI-DNN, or DHI, accurately characterized and classified multiple tumor histologic features in pediatric high-grade brain tumors. If these results could be further validated in patients, the novel DHI might emerge as a favorable alternative to the current neuroimaging techniques to better guide biopsy and resection as well as monitor therapeutic response in patients with high-grade brain tumors.

High-grade pediatric brain tumors constitute the highest mortality of cancer-death in children. While conventional MRI has been widely adopted for examining pediatric high-grade brain tumor clinically, accurate neuroimaging detection and differentiation of tumor histopathology for improved diagnosis, surgical planning, and treatment evaluation, remains an unmet need in the clinical management of pediatric brain tumor. We employed a novel Diffusion Histology Imaging (DHI) approach that incorporates diffusion basis spectrum imaging (DBSI) and deep neural network. DHI aims to detect, differentiate, and quantify heterogenous areas in pediatric high-grade brain tumors, which include normal white matter (WM), densely cellular tumor (DC tumor), less densely cellular tumor (LDC tumor), infiltrating edge, necrosis, and hemorrhage. Distinct diffusion metric combination would thus indicate the unique distributions of each distinct tumor histology features. DHI, by incorporating DBSI metrics and the deep neural network algorithm, classified pediatric tumor histology with an overall accuracy of 83.3%. Receiver operating analysis (ROC) analysis suggested DHI’s great capability in distinguishing individual tumor histology with AUC values (95%CI) of 0.983 (0.985-0.989), 0.961 (0.957-0.964), 0.993 (0.992-0.994), 0.953 (0.947-0.958), 0.974 (0.970-0.978) and 0.980 (0.977-0.983) for normal WM, DC tumor, LDC tumor, infiltrating edge, necrosis and hemorrhage, respectively. Our results suggest that DBSI-DNN, or DHI, accurately characterized and classified multiple tumor histologic features in pediatric high-grade brain tumors. If further validated in patients, the novel DHI might emerge as a favorable alternative to the current neuroimaging techniques to better guide biopsy and resection as well as monitor therapeutic response in patients with high-grade brain tumors.

BackgroundSex is recognized as a significant determinant of outcome among glioblastoma patients, but the relative prognostic importance of glioblastoma features has not been thoroughly explored for sex differences.MethodsCombining multi-modal MR images, biomathematical models, and patient clinical information, this investigation assesses which pretreatment variables have a sex-specific impact on the survival of glioblastoma patients. Pretreatment MR images of 494 glioblastoma patients (299 males and 195 females) were segmented to quantify tumor volumes. Cox proportional hazard (CPH) models and Student’s t-tests were used to assess which variables were associated with survival outcomes.ResultsAmong males, tumor (T1Gd) radius was a predictor of overall survival (HR=1.027, p=0.044). Among females, higher tumor cell net invasion rate was a significant detriment to overall survival (HR=1.011, pConclusionDespite similar distributions of the MR imaging parameters between males and females, there was a sex-specific difference in how these parameters related to outcomes, which emphasizes the importance of considering sex as a biological factor when determining patient prognosis and treatment approach.

Significance: The blood–brain barrier (BBB) is a major obstacle to detecting and treating brain tumors. Overcoming this challenge will facilitate the early and accurate detection of brain lesions and guide surgical resection of tumors. Aim: We generated an orthotopic brain tumor model that simulates the pathophysiology of gliomas at early stages; determine the BBB integrity and breakdown over the time course of tumor progression using generic and cancer-targeted near-infrared (NIR) fluorescent molecular probes. Approach: We developed an intracranial tumor xenograft model that rapidly reestablished BBB integrity and monitored tumor progression by bioluminescence imaging. Sham control mice were injected with phosphate-buffered saline only. Fluorescence molecular tomography (FMT) was used to quantify the uptake of tumor-targeted and passive NIR fluorescent imaging agents in orthotopic glioma (U87-GL-GFP PDE7B H217Q cells) tumor model. Cancer-induced and transient (with focused ultrasound, FUS) disruption of BBB integrity was monitored with NIR fluorescent dyes. Results: Stereotactic injection of 50,000 cells into mouse brain allowed rapid reestablishment of BBB integrity within a week, as determined by the inability of both tumor-targeted and generic NIR imaging agents to extravasate into the brain. Tumor-induced BBB disruption was observed 7 weeks after tumor implantation. FUS achieved a similar effect at any time point after reestablishing BBB integrity. While tumor uptake and retention of the passive NIR dye, indocyanine green, was negligible, both actively tumor-targeting agents exhibited selective accumulation in the tumor region. The tumor-targeting molecular probe that clears rapidly from nontumor brain tissue exhibits higher contrast than the analogous vascular-targeting agent and helps delineate tumors from sham control. Conclusions: We highlight the utility of FMT imaging for longitudinal assessment of brain tumors and the interplay between the stages of BBB disruption and molecular probe retention in tumors, with potential application to other neurological diseases.

SUMMARY Over the past decade, wingless-activated (WNT) medulloblastoma has been identified as a candidate for therapy de-escalation based on excellent survival; however, a paucity of relapses has precluded additional analyses of markers of relapse. To address this gap in knowledge, an international cohort of 93 molecularly confirmed WNT MB was assembled, where 5-year progression-free survival is 0.84 (95%, 0.763–0.925) with 15 relapsed individuals identified. Maintenance chemotherapy is identified as a strong predictor of relapse, with individuals receiving high doses of cyclophosphamide or ifosphamide having only one very late molecularly confirmed relapse (p = 0.032). The anatomical location of recurrence is metastatic in 12 of 15 relapses, with 8 of 12 metastatic relapses in the lateral ventricles. Maintenance chemotherapy, specifically cumulative cyclophosphamide doses, is a significant predictor of relapse across WNT MB. Future efforts to de-escalate therapy need to carefully consider not only the radiation dose but also the chemotherapy regimen and the propensity for metastatic relapses., Graphical Abstract, In Brief Nobre et al. analyze a clinically annotated cohort of 93 WNT-activated medulloblastoma using an integrated genomic approach. A maintenance chemotherapy regimen is the strongest predictor of relapse, and cumulative cyclophosphamide doses confer a reduced risk of relapse. Unlike other medulloblastoma subgroups, WNT-activated medulloblastoma recurs most frequently in the lateral ventricles.

Background The median survival for patients with glioblastoma (GBM), the most common primary malignant brain tumor in adults, has remained approximately 1 year for more than 2 decades. Recent advances in the field have identified GBM as a sexually dimorphic disease. It is less prevalent in females and they have better survival compared to males. The molecular mechanism of this difference has not yet been established. Iron is essential for many biological processes supporting tumor growth and its regulation is impacted by sex. Therefore, we interrogated the expression of a key component of cellular iron regulation, the HFE (homeostatic iron regulatory) gene, on sexually dimorphic survival in GBM. Methods We analyzed TCGA microarray gene expression and clinical data of all primary GBM patients (IDH-wild type) to compare tumor mRNA expression of HFE with overall survival, stratified by sex. Results In low HFE expressing tumors (below median expression, n = 220), survival is modulated by both sex and MGMT status, with the combination of female sex and MGMT methylation resulting in over a 10-month survival advantage (P < .0001) over the other groups. Alternatively, expression of HFE above the median (high HFE, n = 240) is associated with significantly worse overall survival in GBM, regardless of MGMT methylation status or patient sex. Gene expression analysis uncovered a correlation between high HFE expression and expression of genes associated with immune function. Conclusions The level of HFE expression in GBM has a sexually dimorphic impact on survival. Whereas HFE expression below the median imparts a survival benefit to females, high HFE expression is associated with significantly worse overall survival regardless of established prognostic factors such as sex or MGMT methylation.

Glioblastoma (GBM) is the most common malignant brain tumor in the United States. Incidence of GBM increases with age, and younger age-at-diagnosis is significantly associated with improved prognosis. While the relationship between candidate GBM risk SNPs and age-at-diagnosis has been explored, genome-wide association studies (GWAS) have not previously been stratified by age. Potential age-specific genetic effects were assessed in autosomal SNPs for GBM patients using data from four previous GWAS. Using age distribution tertiles (18–53, 54–64, 65+) datasets were analyzed using age-stratified logistic regression to generate p values, odds ratios (OR), and 95% confidence intervals (95%CI), and then combined using meta-analysis. There were 4,512 total GBM cases, and 10,582 controls used for analysis. Significant associations were detected at two previously identified SNPs in 7p11.2 (rs723527 [p(54–63)=1.50×10(−9), OR(54–63)=1.28, 95%CI(54–63)=1.18–1.39; p(64+)=2.14×10(−11), OR(64+)=1.32, 95%CI(64+)=1.21–1.43] and rs11979158 [p(54–63)=6.13×10(−8), OR(54–63)=1.35, 95%CI(54–63)=1.21–1.50; p(64+)=2.18×10(−10), OR(64+)=1.42, 95%CI(64+)=1.27–1.58]) but only in persons >54. There was also a significant association at the previously identified lower grade glioma (LGG) risk locus at 8q24.21 (rs55705857) in persons ages 18–53 (p(18–53)=9.30×10(−11), OR(18–53)=1.76, 95%CI(18–53)=1.49–2.10). Within The Cancer Genome Atlas (TCGA) there was higher prevalence of ‘LGG’-like tumor characteristics in GBM samples in those 18–53, with IDH1/2 mutation frequency of 15%, as compared to 2.1% [54–63] and 0.8% [64+] (p=0.0005). Age-specific differences in cancer susceptibility can provide important clues to etiology. The association of a SNP known to confer risk for IDH1/2 mutant glioma and higher prevalence of IDH1/2 mutation within younger individuals 18–53 suggests that more younger individuals may present initially with ‘secondary glioblastoma.’

Focused ultrasound combined with microbubble-mediated intranasal delivery (FUSIN) is a new brain drug delivery technique. FUSIN utilizes the nasal route for direct nose-to-brain drug administration, thereby bypassing the blood-brain barrier (BBB) and minimizing systemic exposure. It also uses FUS-induced microbubble cavitation to enhance transport of intranasally (IN) administered agents to the FUS-targeted brain location. Previous studies have provided proof-of-concept data showing the feasibility of FUSIN to deliver dextran and the brain-derived neurotrophic factor to the caudate putamen of mouse brains. The objective of this study was to evaluate the biodistribution of IN administered gold nanoclusters (AuNCs) and assess the feasibility and short-term safety of FUSIN for the delivery of AuNCs to the brainstem. Three experiments were performed. First, the whole-body biodistribution of IN administered (64)Cu-alloyed AuNCs ((64)Cu-AuNCs) was assessed using in vivo positron emission tomography/computed tomography (PET/CT) and verified with ex vivo gamma counting. Control mice were intravenously (IV) injected with the (64)Cu-AuNCs. Second, (64)Cu-AuNCs and Texas red-labeled AuNCs (TR-AuNCs) were used separately to evaluate FUSIN delivery outcome in the brain. (64)Cu-AuNCs or TR-AuNCs were administered to mice through the nasal route, followed by FUS sonication at the brainstem in the presence of systemically injected microbubbles. The spatial distribution of (64)Cu-AuNCs and TR-AuNCs were examined by autoradiography and fluorescence microscopy of ex vivo brain slices, respectively. Third, histological analysis was performed to evaluate any potential histological damage to the nose and brain after FUSIN treatment. The experimental results revealed that IN administration induced significantly lower (64)Cu-AuNCs accumulation in the blood, lungs, liver, spleen, kidney, and heart compared with IV injection. FUSIN enhanced the delivery of (64)Cu-AuNCs and TR-AuNCs at the FUS-targeted brain region compared with IN delivery alone. No histological-level tissue damage was detected in the nose, trigeminal nerve, and brain. These results suggest that FUSIN is a promising technique for noninvasive, spatially targeted, and safe delivery of nanoparticles to the brain with minimal systemic exposure.

Background Glioblastoma (GBM) is the most common and most malignant glioma. Nonglioblastoma (non-GBM) gliomas (WHO Grades II and III) are invasive and also often fatal. The goal of this study is to determine whether sex differences exist in glioma survival. Methods Data were obtained from the National Cancer Database (NCDB) for years 2010 to 2014. GBM (WHO Grade IV; N = 2073) and non-GBM (WHO Grades II and III; N = 2963) were defined using the histology grouping of the Central Brain Tumor Registry of the United States. Non-GBM was divided into oligodendrogliomas/mixed gliomas and astrocytomas. Sex differences in survival were analyzed using Kaplan–Meier and multivariable Cox proportional hazards models adjusted for known prognostic variables. Results There was a female survival advantage in patients with GBM both in the unadjusted (P = .048) and adjusted (P = .003) models. Unadjusted, median survival was 20.1 months (95% CI: 18.7-21.3 months) for women and 17.8 months (95% CI: 16.9-18.7 months) for men. Adjusted, median survival was 20.4 months (95% CI: 18.9-21.6 months) for women and 17.5 months (95% CI: 16.7-18.3 months) for men. When stratifying by age group (18-55 vs 56+ years at diagnosis), this female survival advantage appeared only in the older group, adjusting for covariates (P = .017). Women (44.1%) had a higher proportion of methylated MGMT (O6-methylguanine-DNA methyltransferase) than men (38.4%). No sex differences were found for non-GBM. Conclusions Using the NCDB data, there was a statistically significant female survival advantage in GBM, but not in non-GBM.

Background Leptomeningeal disease and hydrocephalus are present in up to 30% of patients with diffuse intrinsic pontine glioma (DIPG), however there are no animal models of cerebrospinal fluid (CSF) dissemination. As the tumor-CSF-ependymal microenvironment may play an important role in tumor pathogenesis, we identified characteristics of the Nestin-tumor virus A (Nestin-Tva) genetically engineered mouse model (GEMM) that make it ideal to study the interaction of tumor cells with the CSF and its associated pathways with implications for the development of treatment approaches to address CSF dissemination in DIPG. Methods A Nestin-TVa model of DIPG utilizing the three most common DIPG genetic alterations (H3.3K27M, PDGF-B, p53) was used for this study. All animals underwent MR imaging and a subset underwent histopathologic analysis with H&E and beta-IV tubulin. Results Tumor dissemination within the CSF pathways (ventricles, leptomeninges) was present in 76% (25/33) of animals, with invasion of the choroid plexus, disruption of the ciliated ependyma and regional subependymal fluid accumulation. Ventricular enlargement consistent with hydrocephalus was present in 94% (31/33). Ventricle volume correlated with region specific transependymal CSF flow (periventricular T2 signal), localized anterior to the lateral ventricles. Subependymal tumor cells were also present subjacent to the 4th ventricle in a post-mortem human specimen. Conclusions This is the first study to report CSF pathway tumor dissemination an animal model of DIPG and is representative of CSF dissemination seen clinically. Understanding the CSF-tumor-ependymal microenvironment has significant implications for treatment of DIPG through targeting mechanisms of tumor spread within the CSF pathways.

Sex differences in cancer incidence and survival, including central nervous system tumors, are well documented. Multiple mechanisms contribute to sex differences in health and disease. Recently, the presence of fetal-in-maternal microchimeric cells has been shown to have prognostic significance in breast and colorectal cancers. The frequency and potential role of these cells has not been investigated in brain tumors. We therefore selected two common primary adult brain tumors for this purpose: meningioma, which is sex hormone responsive and has a higher incidence in women, and glioblastoma, which is sex hormone independent and occurs more commonly in men. Quantitative PCR was used to detect the presence of male DNA in tumor samples from women with a positive history of male pregnancy and a diagnosis of either glioblastoma or meningioma. Fluorescence in situ hybridization for the X and Y chromosomes was used to verify the existence of intact male cells within tumor tissue. Fetal microchimerism was found in approximately 80% of glioblastoma cases and 50% of meningioma cases. No correlations were identified between the presence of microchimerism and commonly used clinical or molecular diagnostic features of disease. The impact of fetal microchimeric cells should be evaluated prospectively.

BACKGROUND Significant sex differences exist in cancer, and males have higher incidence and lower survival compared to females for most cancers. No large-scale studies have systematically examined sex differences in incidence and survival across all primary brain histologies. We performed a comprehensive investigation of the differences in incidence and survival in patients diagnosed with primary malignant brain and other CNS tumors by specific histologies. METHODS Age-adjusted incidence rate ratios (IRR) and 95% confidence intervals (945% CI) were generated from the United States Cancer Statistics (USCS) Public Use Database. Data from the Surveillance, Epidemiology, and End-Results (SEER) program were used to calculate overall survival. Data was restricted to patients with histologically or radiologically confirmed, primary malignant tumors diagnosed between 2001 and 2015. Histological groupings were categorized based on the Central Brain Tumor Registry of the United States (CBTRUS). Cox proportional hazards models were used to calculate hazard ratios (HR) adjusted for age for males as compared to females. RESULTS Males exhibited higher incidence than females in all brain histologies except meningioma (IRR=0.83; 95% CI 0.73–0.93) and other neuroepithelial tumors (i.e. polar spongioblastomas and astroblastomas) (IRR=0.48; 95% CI 0.26–0.88). Males experienced better overall survival in germ cell tumors, cysts and heterotopias (HR=0.68; 95% CI 0.49–0.94) compared to females, but were observed to have lower survival in all other histologies. Survival was lowest for males among patients with nerve sheath tumors (HR=2.32; 95% CI 1.31–4.12) and other neoplasms related to the meninges (e.g. chondrosarcomas and chordomas) (HR=2.23; 95% CI 1.24–3.99). Survival in meningioma patients was significantly higher in females (HR=1.49; 95% CI 1.25–1.77). Patients with glioblastomas, had a slightly worse survival outcome in males (HR=1.02, 95% CI 1.00–1.05). CONCLUSION Understanding the role of sex differences is critical for addressing sex based inequalities and needs to be taken into account in clinical paradigms.

Sex differences in the rates of glioblastoma increase with age. There are likely to be multiple biological mechanisms underlying these sex differences. Here, we focused on the changes that occur with aging in the secretomes of male and female astrocytes to determine whether sex differences in chemokine and cytokine secretion, and inflammatory cell recruitment, could be one of those mechanisms. We focused on glioblastoma (GBM) and the role that senescent astrocytes might play in the age-dependent widening of the gap between male and female GBM cases. We found that the senescence associated secretory phenotype of male and female astrocytes significantly differed, notably in the enrichment of Fractalkine (1.33:1 (F:M)), the primary chemoattractant for microglia, in female compared to male SASP. This was in contrast to the greater abundance of tumorigenic growth factors like bFGF (1.25:1 (M:F)), in the male SASP. Implantation of either male or female senescent astrocytes into the brains of mice resulted in the recruitment of microglia to the injection site. Regardless of the recipient mouse sex, female astrocyte implantation resulted in significantly larger microglial infiltrates (2-fold) and greater astrocyte activation, as compared to the injection of equal numbers of male senescent cells. We propose a model for how sex differences in astrocyte SASP could result in lower rates of GBM with age in females: greater microglial attraction to senescent cells results in their enhanced clearance and consequently a reduction in senescence-associated GBM promotion.

PURPOSE Brain tumor related epilepsy (BTE) is a major co-morbidity in patients with glioma. It is difficult to determine whether the use of anti-epileptic drugs is necessary. We attempted to build a machine-learning model to predict the probability of seizure presentation (SP) with glioma. METHODS We trained a random forest classifier using the following variables: volumetric data of pre-treatment MR images (T1Gd and T2-FLAIR sequences), patient demographics (age; sex), and measurements of tumor proliferation (log(ρ)), invasiveness (log(D)) and their relative ratio (log(ρ/D)). Our cohort consisted of 221 patients total. Using an 80-20 ratio, we used 176 patients (76 SP, 100 nSP) for training and the remaining 45 patients (19 SP, 26 nSP) were used for testing. We also trained on male-only and female-only cohorts to evaluate any sex differences in prediction. For training, 108 males (53 SP, 55 nSP) were used and 28 for testing (14 SP, 14 nSP). We used 72 females (21 SP, 49 nSP) for training and 15 (7 SP, 8 nSP) for testing. We corrected for class imbalance in the female cohort before training. Using 10-fold cross-validation and a separate testing set, we measured performance by ROC curve (AUC), accuracy, sensitivity, and specificity of predictions (average of folds in cross validation). RESULTS The female model achieved the highest AUC (0.853) followed by the mixed model (0.726) and the male model (0.651). In the validation set, the accuracy/sensitivity/specificity of the three cohorts were as follows: mixed (0.726/0.696/0.750), female (0.853/0.830/0.875), and male (0.651/0.577/0.722). The performance of the testing set, in terms of accuracy/sensitivity/specificity were: mixed (0.733/0.74/0.73), female (0.8/0.57/1), and male (0.714/0.64/0.79). CONCLUSION We found a negative correlation between seizure probability and size and invasiveness of tumors. Our model shows promising performance on testing set data. Further cohort studies and training is warranted.

Incidence rates for glioblastoma increase exponentially with age in a sex-specific manner, with a steeper increase observed in men. This suggests that age may be a more significant factor in promoting male tumors than female tumors. Drivers in the age-related cancer increase include both the accumulation of DNA damage and microenvironment changes, specifically an increase in the number of senescent cells. Cellular senescence comprises two distinct components – a cell cycle arrest program that prevents proliferation of mutated cells and acts as a potent anti-cancer mechanism, and a paracrine signaling component that can have both pro- and anti-tumorigenic properties. The contribution of senescence to brain tumor incidence and outcome is currently unknown; additionally, no research has been done on whether there are sex differences in mechanisms of senescence induction or in the secretory phenotype of senescent cells. Using both wildtype astrocytes and a glioblastoma model, we show that female cells have a lower threshold for senescence induction in response to oxidative stress, telomere shortening, and DNA damage, and that sex differences in senescence induction in response to DNA damage are in part mediated by differences in the regulation of p21. In addition, we show that male and female senescent astrocytes differ in their secretory phenotypes. Male senescent astrocytes secrete more tumor promoting factors, and conditioned media from male but not female senescent astrocytes promotes tumor cell proliferation. These findings provide a potential explanation for sex differences in glioblastoma incidence rates. Achieving a better understanding of mechanisms of senescence and how they differ in male and female cells could advance development of senescence directed therapies and potentially improve cancer treatment for a range of tumor types.

Sex differences in brain tumor related epilepsy (BTE) as an initial symptom of glioma are rarely investigated. Sex differences are being explored with increased interest more recently. Epidemiological studies have generally indicated overall incidence of epilepsy to be slightly higher in males than females, but a higher incidence of idiopathic generalized epilepsy has been seen in females in some studies. Clinicians regularly face the challenge of anti-epileptic drug prescription determination in patients with newly identified brain mass and would greatly benefit from knowing clinical characteristics in BTE. We analyzed clinical characteristics in patients with pathologically-determined glioma with known seizure status at clinical presentation using non-parametric and Fisher’s Exact tests with the entire cohort and male and female separately. From our clinical data repository, we identified 223 patients (females, n=86 (39%); males, n=137 (61%)) divided by those that presented with seizure (SP, n=96, 43%; F, n= 28; M, n= 68) and those that presented without seizure (nSP, n=127, 57%, F n=58, M n=69). In sex-specific analysis males had a significantly (x2=6.28, p=0.01) higher proportion of SP (49.6%) than females (32.6%). There was a dependence between seizure presentation status and tumor grade in males (x2=5.94, p=0.015). Tumor kinetics in the female cohort showed significantly (p< 0.0001) more diffuse tumors in SP compared to nSP. Right-sided tumors were predominantly nSP, while left-sided tumors were split more evenly between SP and nSP. The predominance of nSP among right-sided tumors seems to be driven by females (74%), male right-sided tumors were split more evenly between SP and nSP. Seizure status is dependent on tumor location for the combined cohort, however not for males (x2=5.23, p=0.16) or females (x2=4.34, p=0.23). Clearly there are sex differences and similarities in clinical characteristics at glioma presentation. Further characterization could lead to improved adherence of national standards for AED prescription.

Glioblastoma (GBM) is the most common and aggressive brain tumor in adults. GBM occurs more commonly in males, but female patients survive significantly longer. Understanding the molecular mechanisms that underlie those sex differences could support novel treatment strategies. In this regard, we found that male and female GBM patient samples differ in their metabolite abundance and that male patients exhibit a significantly higher abundance of TCA cycle metabolites. We confirmed those findings in a murine model of GBM, which has previously yielded important insights into sexual dimorphism in GBM. Strikingly, sex differences in TCA cycle flux were entirely driven by glutamine flux, not glucose flux, suggesting a sex-specific role for glutamine in GBM. Metabolic manipulation through glutamine deprivation resulted in a greater growth inhibition in male GBM cells. Glutamine itself can be utilized for anabolic reactions or it can be converted to glutamate by glutaminase. Only male GBM cells were sensitive to pharmacological glutaminase inhibition with BPTES or CB-839, suggesting that male GBM cells are glutamate dependent while female GBM cells are not. Concordantly, we found significantly higher glutaminase levels in male GBM cells. Furthermore, we found that numerous metabolites (including NADH, ATP, and glutathione) involved in cellular processes downstream of glutamate were more abundant in male GBM cells. In contrast, female GBM cells were resistant to low glutamine conditions and glutaminase inhibitors unless glutamine-synthase activity was disrupted, suggesting that glutamine synthesis might play a more prominent role in female GBM. Together, these data indicate that male and female GBM differ in their metabolic adaptions. Male GBM utilize glutamate to fuel the TCA cycle and mitochondrial activity while female GBM synthesize and utilize glutamine itself. This sexual dimorphism in metabolic reprogramming reveals novel sex specific metabolic targets for GBM and underlines the importance of considering sex in metabolic targeting approaches.

Seizures are common presenting symptoms of primary brain tumors. Mechanisms of epileptogenesis are still unknown and are believed to be multifactorial. Previous studies have indicated correlation of seizure with tumor location. Recent investigations of our group have shown image-based parameters have sex-specific implications for patient outcome. In this retrospective study, we examined the association of tumor location with the probability and risk of seizure in male and female glioma patients.

BackgroundBrain tumor related epilepsy (BTE) is a major co-morbidity related to the management of patients with brain cancer. Despite published practice guidelines recommending against anti-epileptic drug (AED) utilization in patients with gliomas, there is heterogeneity in prescription practices of AEDs in these patients. In an attempt to impact BTE management, we statistically analyzed clinically relevant attributes (sex, age, tumor size, tumor growth kinetics, and tumor location) pertaining to seizure at presentation and used them to build a computational machine learning model to predict the probability of a seizure (at presentation).MethodsFrom our clinical data repository, we identified 223 patients (females, n=86; males, n=137) with pathologically-determined glioma and known seizure status at clinical presentation. Non-parametric and Fisher’s Exact tests were used to identify statistical differences in clinical characteristics. We utilized a random forest machine learning method for generating our predictive models by entire cohort and separated by male and female.FindingsPatients were divided into those that presented with seizure (SP, n=96, 43%; F, n= 28; M, n= 68) and those that presented without seizure (nSP, n=127, 57%, F n=58, M n=69). Females presented with seizures significantly less often than males (x2=6·28, p=0·01). SP patients had significantly smaller T1Gd radius compared to nSP (SP 11·30mm, nSP 18.66mm, pInterpretationDespite heterogeneity across our patient cohort, there is strong evidence of a role for patient sex, tumor size, tumor invasion, and patient age in predicting the incidence of seizures at diagnosis. Future studies, with prospectively detailed data collection, may provide clearer insights into the incidence of seizures through a patient’s treatment course.

Sex differences in the incidence and outcome of human disease are broadly recognized but, in most cases, not sufficiently understood to enable sex-specific approaches to treatment. Glioblastoma (GBM), the most common malignant brain tumor, provides a case in point. Despite well-established differences in incidence and emerging indications of differences in outcome, there are few insights that distinguish male and female GBM at the molecular level or allow specific targeting of these biological differences. Here, using a quantitative imaging-based measure of response, we found that standard therapy is more effective in female compared with male patients with GBM. We then applied a computational algorithm to linked GBM transcriptome and outcome data and identified sex-specific molecular subtypes of GBM in which cell cycle and integrin signaling are the critical determinants of survival for male and female patients, respectively. The clinical relevance of cell cycle and integrin signaling pathway signatures was further established through correlations between gene expression and in vitro chemotherapy sensitivity in a panel of male and female patient-derived GBM cell lines. Together, these results suggest that greater precision in GBM molecular subtyping can be achieved through sex-specific analyses and that improved outcomes for all patients might be accomplished by tailoring treatment to sex differences in molecular mechanisms.

Wingless-activated (WNT) medulloblastoma have been identified over the past decade as being a candidate for therapy de-escalation based on their excellent survival, however a paucity of relapses in any single cohort have precluded any additional analysis of markers of relapse. To address this gap in knowledge, an international cohort of 91 WNT-MB was assembled where we identified 14 relapsed cases where five-year progression free survival was 0.84 (95% 0.763-0.925). Maintenance chemotherapy was identified as a strong predictor of relapse, where no relapses were observed in cases receiving high doses of cyclophosphamide or ifosphamide (p=0.0026). The location of relapse was metastatic in 10/14 of relapses, with 7/10 metastatic relapses presenting along the ependymal lining in the lateral ventricles. Maintenance chemotherapy, specifically cumulative cyclophosphamide dose is a significant predictor of relapse across WNT-MB. Future efforts to de-escalate therapy need to carefully consider not only the radiation dose but also the chemotherapy regimen, and the propensity to metastatic relapses.

Glioblastoma (GBM) is an aggressive brain tumor in children and adults. It occurs more commonly in males, but female patients survive significantly longer. Understanding the molecular mechanisms that underlie those sex differences could support novel treatment strategies. In this regard, we found that male and female GBM patient samples differ in their metabolite abundance and that males exhibit a significantly higher abundance of amino acid metabolites. We confirmed those findings in a murine model of GBM, which has previously yielded important insights into sexual dimorphism in GBM. Furthermore, we found that male GBM cell cultures are significantly more sensitive to amino acid deprivation, which was almost entirely driven by amino acids involved in the synthesis of the antioxidant glutathione. Glutaminase 1 (GLS1) mediates the conversion from glutamine to glutamate, a crucial component of glutathione. We found that male GBM cells exhibited higher levels of GLS1, suggesting they are more dependent on glutamate. Indeed, we found that male GBM cells are more sensitive to pharmacological GLS1 inhibition with the clinical inhibitor CB-839. This correlated with significantly increased reactive oxygen species (ROS) in males compared to females. We further confirmed sex differences in redox state through pharmacological depletion of glutathione that resulted in a significant increase in ROS and cell death in male GBM. Together, these data indicate that male GBM cells are more dependent on glutamine to regulate ROS levels. This reveals novel sex-specific metabolic targets for GBM and underlines the importance of considering sex in metabolic targeting approaches.

Glioblastoma (GBM) is a lethal brain tumor that has a higher incidence and mortality in males compared to females. GBM tumors consume glucose and glutamine as the major fuels for growth and strategies have been developed that disrupt these nutrient utilization pathways. Both pre-clinical and clinical reports have identified the ketogenic diet (KD) as an approach which consists of high fat and low carbohydrates. It reduces circulating glucose and increases ketones resulting in restricted tumor growth. Because of published studies showing that carbohydrate utilization is higher in healthy males, we hypothesized that the KD would have greater inhibition of male GBM growth. We compared sex differences in response to the KD using an established GBM mouse model (Nf1-/-DNp53), which has previously yielded important insights into sex differences in GBM. Flank tumors were implanted and tumor size measurements were performed. Unexpectedly, the KD had no effect on male GBM tumor growth, but female tumors grew significantly slower in mice that were on the KD compared to control diet. To understand how male GBM tumors adapt their metabolism to the KD, we measured serial blood glucose and ketone levels. We discovered expected physiologic responses to the KD with decreased serum glucose and increased serum ketones, suggesting that males adapt to the KD through changes in cell-intrinsic metabolism. Using FDG-PET imaging to investigate glucose utilization, we found that FDG uptake was increased in both tumor and normal brain of male but not female mice that were on the KD compared to control diet. Taken together, our results show that the KD has a greater impact on female GBM tumor growth and that sex differences in metabolic adaptations such as increased glucose transport may underlie the male-specific resistance to the KD. This study has the potential to develop sex-specific metabolic treatment approaches for GBM.

Sex differences in incidence and outcome are consistently observed in cancer, including in the most common malignant brain tumor – glioblastoma (GBM). Women are less likely to develop GBM and have a survival advantage compared to men. Previous research from our lab identified the proteins Rb, p16, and p21 as key mediators of female protection against cellular transformation. Strikingly, these proteins are all primary components of the senescence response, a potent cell-intrinsic anti-cancer mechanism that results in permanent cell cycle arrest, preventing proliferation of damaged cells. We hypothesized that sex differences in GBM incidence and outcome are mediated by differences in senescence induction after DNA damage in male and female cells. Using both wildtype astrocytes and a GBM model, we found that females have a lower threshold for senescence induction in response to oxidative stress, telomere shortening, and treatment with chemotherapy or radiation. Following irradiation, female GBM model cells had higher levels of p21 expression, and p21 levels correlated with the percentage of senescent cells. Knocking out p21 eliminated sex differences in the senescence response following irradiation. Achieving a better understanding of mechanisms of senescence and how they differ in male and female cells could advance development of senescence-directed therapies and potentially improve treatment for brain tumors.

e14525 Background: High grade gliomas (HGG) (the most common being glioblastoma) are the most common primary CNS malignancy in adults. Mainstay of therapy is surgical resection followed by concurrent radiation and temozolomide (TMZ) followed by adjuvant TMZ. Unfortunately, prognosis remains poor and optimization of current therapy is critical. Chronotherapy is defined as improvement in treatment outcomes by maximizing treatment efficacy and minimizing toxicity by administering medications in accordance with biological rhythms of the patient. In a mouse model, there was greater anti-tumor efficacy during morning administration of TMZ. This trial was designed to determine the feasibility and potential clinical impact of chrono-therapeutically administering TMZ in patients with HGG. Methods: Adult patients ( > 18 years) with HGG (WHO Grade III/IV) were eligible. Patients were screened and consented prior to initiation of monthly TMZ therapy. Eligible patients were randomized to TMZ in the morning (AM) before 10AM or in the evening (PM) after 8PM. Pill diaries were recorded for drug administration time and compliance. Fact-Br Quality of Life (QoL) surveys were administered to patients at the time of enrollment in the trial and at the end of treatment to measure differences in QoL in both groups. Circadian rhythm was recorded by Actiwatch. Adverse events (AE), overall survival (OS) and progression free survival (PFS) were measured for each group. Results: At the time of submission, a total of 28 patients were evaluated. 15 patients were in AM group and 13 in PM group. It is feasible for participants to take TMZ per study assignment. There was no significant difference in the QoL based on the Fact-Br dataset in the four main categories of physical well-being, social/family well-being, functional well-being and emotional well-being. The Friedman’s two-way nonparametric ANOVA tests were used to analyze the differences across time points. Cytopenias are a known adverse effect of TMZ. There was a trend towards worsening lymphocyte counts in the AM group compared to PM group, although not statistically significant. There was no statistical significance in PFS or OS in patients with newly diagnosed glioblastoma. Conclusions: Chronotherapy with TMZ is feasible. A trend of worsening lymphocyte counts is noted in AM treatment group compared to PM group but was not statistically significant. No difference in OS or PFS was noted, although sample size was too small to effectively assess this. A larger study will need to be conducted to effectively assess the effect of chronotherapy on survival. Clinical trial information: NCT02781792.

Primary brain tumor incidence rates rise more rapidly in males than females with increasing age. Both pre-neoplastic cells and stromal elements contribute to age-related increases in cancer, and cellular senescence plays a role in the transformation of pre-neoplastic cells. While senescence prevents the proliferation of damaged cells, senescent cells produce senescence-associated secretory phenotypes (SASP) which contribute to the tumor microenvironment. Certain SASP factors favor tumor suppression by inducing senescence in neighboring cells or recruiting immune cells for tumor clearance, while other factors increase tumor proliferation, resistance and relapse. Studies have demonstrated the role of senescence in the growth of solid tumors in various tissues including the liver, pancreas, and prostate. However, no studies have investigated how senescence in resident brain cells influences age-dependent increases in brain tumors. Moreover, there are no data in any tissues about sex differences in cellular senescence and its effect on age-dependent cancer rates. In the brain, astrocytes are the most abundant resident cell type with critical functions in brain structure, physiology, and homeostasis. Thus, we investigated sex differences in astrocyte senescence and whether these differences contribute to the sex disparity in brain tumor rates, specifically glioblastoma which has a male to female ratio of 1.6. Using β-galactosidase staining, we found that murine female astrocytes undergo senescence more readily after oxidative stress than their male counterparts. Furthermore, in vitro studies showed that murine GBM cells cultured in female astrocyte SASP-conditioned media show a different pattern of growth as compared to those cultured in male astrocyte SASP-conditioned media. These results indicate that there are sex differences in astrocyte senescence that influence tumor proliferation. Further investigation into the molecular mechanisms involved in male and female senescent astrocytes will allow us to better understand age-dependent brain tumor growth, which may reveal novel therapeutic targets for treatment.

Background Diagnosis of diffuse intrinsic pontine glioma (DIPG) has relied on imaging studies, since the appearance is pathognomonic, and surgical risk was felt to be high and unlikely to affect therapy. The DIPG Biology and Treatment Study (DIPG-BATS) reported here incorporated a surgical biopsy at presentation and stratified subjects to receive FDA-approved agents chosen on the basis of specific biologic targets. Methods Subjects were eligible for the trial if the clinical features and imaging appearance of a newly diagnosed tumor were consistent with a DIPG. Surgical biopsies were performed after enrollment and prior to definitive treatment. All subjects were treated with conventional external beam radiotherapy with bevacizumab, and then stratified to receive bevacizumab with erlotinib or temozolomide, both agents, or neither agent, based on O6-methylguanine-DNA methyltransferase status and epidermal growth factor receptor expression. Whole-genome sequencing and RNA sequencing were performed but not used for treatment assignment. Results Fifty-three patients were enrolled at 23 institutions, and 50 underwent biopsy. The median age was 6.4 years, with 24 male and 29 female subjects. Surgical biopsies were performed with a specified technique and no deaths were attributed to the procedure. Two subjects experienced grade 3 toxicities during the procedure (apnea, n = 1; hypertension, n = 1). One subject experienced a neurologic deficit (left hemiparesis) that did not fully recover. Of the 50 tumors biopsied, 46 provided sufficient tissue to perform the study assays (92%, two-stage exact binomial 90% CI: 83%–97%). Conclusions Surgical biopsy of DIPGs is technically feasible, associated with acceptable risks, and can provide biologic data that can inform treatment decisions.

BackgroundGenome-wide association studies (GWAS) have identified 25 risk variants for glioma, which explain ~30% of heritable risk. Most glioma histologies occur with significantly higher incidence in males. A sex-stratified analysis ide7ntified sex-specific glioma risk variants, and further analyses using gene- and pathway-based approaches may further elucidate risk variation by sex.MethodsResults from the Glioma International Case-Control Study were used as a testing set, and results from three GWAS were combined via meta-analysis and used as a validation set. Using summary statistics for autosomal SNPs found to be nominally significant (p−6in ⅔ algorithms.Results25 genes within five regions and 19 genes within six regions reached the set significance threshold in at least 2/3 algorithms in males and females, respectively.EGFRandRTEL1-TNFRSF6Bwere significantly associated with all glioma and glioblastoma in males only, and a female-specific association inTERT, all of which remained nominally significant after conditioning on known risk loci. There were nominal associations with the Telomeres, Telomerase, Cellular Aging, and Immortality pathway in both males and females.ConclusionsThese results suggest that there may be biologically relevant significant differences by sex in genetic risk for glioma. Additional gene- and pathway-based analyses may further elucidate the biological processes through which this risk is conferred.

Sex differences in the incidence and outcome of human disease are broadly recognized but in most cases not adequately understood to enable sex-specific approaches to treatment. Glioblastoma (GBM), the most common malignant brain tumor, provides a case in point. Despite well-established differences in incidence, and emerging indications of differences in outcome, there are few insights that distinguish male and female GBM at the molecular level, or allow specific targeting of these biological differences. Here, using a quantitative imaging-based measure of response, we found that temozolomide chemotherapy is more effective in female compared to male GBM patients. We then applied a novel computational algorithm to linked GBM transcriptome and outcome data, and identified novel sex-specific molecular subtypes of GBM in which cell cycle and integrin signaling were identified as the critical determinants of survival for male and female patients, respectively. The clinical utility of cell cycle and integrin signaling pathway signatures was further established through correlations between gene expression and in vitro chemotherapy sensitivity in a panel of male and female patient-derived GBM cell lines. Together these results suggest that greater precision in GBM molecular subtyping can be achieved through sex-specific analyses, and that improved outcome for all patients might be accomplished via tailoring treatment to sex differences in molecular mechanisms.One Sentence SummaryMale and female glioblastoma are biologically distinct and maximal chances for cure may require sex-specific approaches to treatment.

Tumor growth is not solely a consequence of autonomous tumor cell properties. Rather, tumor cells act upon and are acted upon by their microenvironment. It is tumor tissue biology that ultimately determines tumor growth. Thus, we developed a compound library screen for agents that could block essential tumor-promoting effects of the glioblastoma (GBM) perivascular stem cell niche (PVN). We modeled the PVN with three-dimensional primary cultures of human brain microvascular endothelial cells in Matrigel. We previously demonstrated stimulated growth of GBM cells in this PVN model and used this to assay PVN function. We screened the Microsource Spectrum Collection library for drugs that specifically blocked PVN function, without any direct effect on GBM cells themselves. Three candidate PVN-disrupting agents, Iridin, Tigogenin and Triacetylresveratrol (TAR), were identified and evaluated in secondary in vitro screens against a panel of primary GBM isolates as well as in two different in vivo intracranial models. Iridin and TAR significantly inhibited intracranial tumor growth and prolonged survival in these mouse models. Together these data identify Iridin and TAR as drugs with novel GBM tissue disrupting effects and validate the importance of preclinical screens designed to address tumor tissue function rather than the mechanisms of autonomous tumor cell growth.

Sex differences in human health and disease can range from undetectable to profound. Differences in brain tumor rates and outcome are evident in males and females throughout the world and regardless of age. These observations indicate that fundamental aspects of sex determination can impact the biology of brain tumors. It is likely that optimal personalized approaches to the treatment of male and female brain tumor patients will require recognizing and understanding the ways in which the biology of their tumors can differ. It is our view that sex-specific approaches to brain tumor screening and care will be enhanced by rigorously documenting differences in brain tumor rates and outcomes in males and females, and understanding the developmental and evolutionary origins of sex differences. Here we offer such an integrative perspective on brain tumors. It is our intent to encourage the consideration of sex differences in clinical and basic scientific investigations.

The molecular bases for sex differences in cancer remain undefined and how to incorporate them into risk stratification remains undetermined. Given sex differences in metabolism and the inverse correlation between fluorodeoxyglucose (FDG) uptake and survival, we hypothesized that glycolytic phenotyping would improve glioma subtyping. Using retrospectively acquired lower-grade glioma (LGG) transcriptome data from The Cancer Genome Atlas (TCGA), we discovered male-specific decreased survival resulting from glycolytic gene overexpression. Patients within this high-glycolytic group showed significant differences in the presence of key genomic alterations (i.e., 1p/19q codeletion, CIC, EGFR, NF1, PTEN, FUBP1, and IDH mutations) compared with the low-glycolytic group. Although glycolytic stratification defined poor prognostic males independent of grade, histology, TP53, and ATRX mutation status, we unexpectedly found that females with high-glycolytic gene expression and wild-type IDH survived longer than all other wild-type patients. Validation with an independent metabolomics dataset from grade 2 gliomas determined that glycolytic metabolites selectively stratified males and also uncovered a potential sexual dimorphism in pyruvate metabolism. These findings identify a potential synergy between patient sex, tumor metabolism, and genomic alterations in determining outcome for glioma patients.

INTRODUCTION: Diffuse intrinsic pontine glioma (DIPG) remains a devastating and incurable disease. The DIPG-BATs clinical trial incorporates diagnostic biopsy with molecularly determined treatment stratification. Here we present the initial genomic analysis of the DIPG-BATs cohort. METHODS: Children enrolled on the DIPG-BATs clinical trial underwent upfront diagnostic biopsies prior to commencement of therapy. RNA and DNA were extracted from single core biopsies and subjected to whole-genome sequencing (WGS) and RNA-sequencing. Tumor samples were also collected at autopsy if there was parental consent. RESULTS: Fifty-three patients were enrolled on study, of whom 50 underwent biopsy. There were no biopsy-related deaths. A mean of 5ug of RNA and 10ug of DNA were extracted from single frozen cores. WGS on 41 DIPG samples (including eight autopsy samples) revealed a mean mutation rate of 0.753 mutations per Mb. We confirmed TP53, PIK3CA, H3F3A, ACVR1, PPM1D, and HIST1H3B to be recurrently mutated in DIPG. Additional mutations were found in epigenetic modifiers including ASXL1. Copy-number analysis revealed PDGFRA to meet statistical significance as a recurrent amplification peak in DIPG (q

Background Noninvasively predicting early response to therapy in recurrent pediatric brain tumors provides a challenge. 3,4-dihydroxy-6-[18F]fluoro-L-phenylalanine (18F-FDOPA) PET/MRI has not been previously studied as a tool to evaluate early response to antiangiogenic therapy in children. The purpose of this study was to evaluate the safety and feasibility of using 18F-FDOPA PET/MRI to assess response to bevacizumab in children with relapsed brain tumors. Materials and Methods Six patients with recurrent gliomas (5 low-grade, 1 high-grade) planned to undergo treatment with bevacizumab were enrolled. 18F-FDOPA PET/MRI scans were obtained prior to and 4 weeks following the start of treatment, and these were compared with the clinical response determined at the 3-month MRI. The primary PET measure was metabolic tumor volume (MTV) at 10 to 15 min after 18F-FDOPA injection. For each tumor, the MTV was determined by manually defining initial tumor volumes of interest (VOI) and then applying a 1.5-fold threshold relative to the mean standardized uptake value (SUV) of a VOI in the frontal lobe contralateral to the tumor. Results 18F-FDOPA PET/MRI was well tolerated by all patients. All tumors were well visualized with 18F-FDOPA on the initial study, with peak tumor uptake occurring approximately 10 min after injection. Maximum and mean SUVs as well as tumor-to-brain ratios were not predictors of response at 3 months. Changes in MTVs after therapy ranged from 23% to 98% (n = 5). There is a trend towards the percent MTV change seen on the 4-week scan correlating with progression-free survival. Conclusion 18F-FDOPA PET/MRI was well tolerated in pediatric patients and merits further investigation as an early predictor of response to therapy.

// Amy Barone 1,* , Rajarshi Sengupta 1,* , Nicole M. Warrington 1 , Erin Smith 2,3 , Patrick Y. Wen 4,5 , Rolf A. Brekken 6 , Barbara Romagnoli 7 , Garry Douglas 7 , Eric Chevalier 7 , Michael P. Bauer 7 , Klaus Dembowsky 7 , David Piwnica-Worms 2,3,8,9 and Joshua B. Rubin 1,10 1 Department of Pediatrics, Washington University School of Medicine, 660 South Euclid Ave, St. Louis, MO 2 BRIGHT Institute , Washington University School of Medicine, 660 South Euclid Ave, St. Louis, MO 3 Molecular Imaging Center, Mallinckrodt Institute of Radiology , Washington University School of Medicine, 660 South Euclid Ave, St. Louis, MO 4 Center for Neuro-Oncology, Dana Farber/Brigham and Women’s Cancer Center, Brookline Ave, Boston, MA 5 Division of Neuro-Oncology, Department of Neurology, Brigham and Women’s Hospital, Brookline Ave, Boston, MA 6 Hamon Center for Therapeutic Oncology Research, UT Southwestern Medical Center, Harry Hines Blvd. Dallas, TX 7 PolyPhor Ltd, Hegenheimermattweg 125 CH-4123 Allschwil, Switzerland 8 Department of Cell Biology & Physiology , Washington University School of Medicine, 660 South Euclid Ave, St. Louis, MO 9 Department of Cancer Systems Imaging, University of Texas MD Anderson Cancer Center, Holcombe Dr., Houston, TX 10 Department of Anatomy and Neurobiology , Washington University School of Medicine, 660 South Euclid Ave, St. Louis, MO * These authors contributed equally to this work Correspondence: Joshua B. Rubin, email: // Keywords : CXCR4, VEGF, perivascular, glioblastoma, stem cells Received : August 22, 2014 Accepted : September 08, 2014 Published : September 09, 2014 Abstract Glioblastoma recurrence involves the persistence of a subpopulation of cells with enhanced tumor-initiating capacity (TIC) that reside within the perivascular space, or niche (PVN). Anti-angiogenic therapies may prevent the formation of new PVN but have not prevented recurrence in clinical trials, suggesting they cannot abrogate TIC activity. We hypothesized that combining anti-angiogenic therapy with blockade of PVN function would have superior anti-tumor activity. We tested this hypothesis in an established intracranial xenograft model of GBM using a monoclonal antibody specific for murine and human VEGF (mcr84) and a Protein Epitope Mimetic (PEM) CXCR4 antagonist, POL5551. When doses of POL5551 were increased to overcome an mcr84-induced improvement in vascular barrier function, combinatorial therapy significantly inhibited intracranial tumor growth and improved survival. Anti-tumor activity was associated with significant changes in tumor cell proliferation and apoptosis, and a reduction in the numbers of perivascular cells expressing the TIC marker nestin. A direct effect on TICs was demonstrated for POL5551, but not mcr84, in three primary patient-derived GBM isolates. These findings indicate that targeting the structure and function of the PVN has superior anti-tumor effect and provide a strong rationale for clinical evaluation of POL5551 and Avastin in patients with GBM.

Medulloblastoma, the most common malignant childhood brain tumor, exhibits distinct molecular subtypes and cellular origins. Genetic alterations driving medulloblastoma initiation and progression remain poorly understood. Herein, we identify GNAS, encoding the G protein Gαs, as a potent tumor suppressor gene that, when expressed at low levels, defines a subset of aggressive Sonic hedgehog (SHH)-driven human medulloblastomas. Ablation of the single Gnas gene in anatomically distinct progenitors in mice is sufficient to induce Shh-associated medulloblastomas, which recapitulate their human counterparts. Gαs is highly enriched at the primary cilium of granule neuron precursors and suppresses Shh signaling by regulating both the cAMP-dependent pathway and ciliary trafficking of Hedgehog pathway components. Elevation in levels of a Gαs effector, cAMP, effectively inhibits tumor cell proliferation and progression in Gnas-ablated mice. Thus, our gain- and loss-of-function studies identify a previously unrecognized tumor suppressor function for Gαs that can be found consistently across Shh-group medulloblastomas of disparate cellular and anatomical origins, highlighting G protein modulation as a potential therapeutic avenue.

PURPOSE: Despite a growing body of research on the etiology of seizures in the brain tumor population, it is not yet clear why approximately one third of this population presents with seizure. In this retrospective study we seek to assess the impact of tumor location on the risk of seizure among male and female glioma patients, respectively. METHODS: From our multi-institutional database, we selected adult patients with contrast-enhancing gliomas (any grade) and known seizure-presentation status at initial diagnosis. Tumors were segmented on pretreatment T2-FLAIR, T2, and post-gadolinium T1 (T1Gd) MR sequences. We warped cortical and subcortical probabilistic atlases to patient images and estimated tumor burden (%) on structures within each lobe and the deep brain. We calculated risk of seizure for various levels of tumor burden and used the result to create sex-specific risk-maps RESULTS: Our cohort included 128 patients (47 females, 81 males) among whom tumors presented with seizure in 34% of females (n=18) and 58% of males (n=47). Patients with tumors in deep brain structures were identified as at risk for seizure in both males and females. In female patients T2 hyperintensity on 44% of right frontal lobe or 28% of left parietal lobe seem to have an estimated 50%+ risk of seizure. Data indicate that in male patients with left-sided tumors, T2 hyperintensity on 30% or more of frontal, temporal, or parietal lobes results in 50%+ risk of seizure. Male patients with right side tumors, showed a similar level of risk for T2 presence in lateral ventricles and parietal lobe. CONCLUSION: This study reveals that the risk of seizure is specific to the location of glioma, the percentage of tumor burden, and patient sex. These differences could be considered in patient management decisions to more selectively prescribe anticonvulsants and optimize patient care.

BACKGROUND: Gliomas can cause significant changes in normal brain function leading to behavior deficits. Functional connectivity (fc) disruptions may contribute to these deficits. However, investigating the effects of glioma growth on fc in humans is complicated by heterogeneity in lesion size, type, and location across subjects. In this study, we evaluated the effects of different patterns of growth on fc, hemodynamic parameters, and behavior in a controlled mouse glioma model. METHODS: 5x10(4) glioma cells growing in either a circumscribed (U87 PDE7B H217Q, n=13) or infiltrative (U87 PDE7B WT, n=14) pattern were stereotactically injected into the forepaw somatosensory cortex of adult nude mice. We monitored tumor burden with weekly bioluminescence imaging (BLI). Additionally, we evaluated fc with weekly functional connectivity optical intrinsic signal imaging (fcOIS). Fc was calculated between homotopic brain regions. Furthermore, cerebral blood flow (CBF) and hemodynamic lags were assessed. Lastly, we assessed performance on a somatomotor behavior battery during the 7(th) week post-injection. RESULTS: Mice with infiltrative tumors had more tumor burden then mice with circumscribed tumors. Furthermore, infiltrative tumors disrupted fc and produced hemodynamic lags earlier than circumscribed tumors. However, infiltrative tumors didn’t change CBF, while circumscribed tumors increased CBF near the injection site. Lastly, mice with infiltrative tumors exhibited somatomotor deficits, while mice with circumscribed tumors performed similar to sham mice (n=20). CONCLUSIONS: We were able to detect differences in fc disruptions in mice with gliomas growing in different patterns with fcOIS. Infiltrative tumors produced greater tumor burden and fc disruptions than circumscribed tumors. Additionally, hemodynamic lag was a better indicator of tumor burden than CBF. Lastly, infiltrative tumors induced behavior deficits during this study, while circumscribed tumors did not. A better understanding of how fc disruptions contribute to behavior deficits in glioma patients may lead to better patient risk stratification and improved behavior outcomes.

Sex impacts the clinical outcome of glioblastoma (GBM) patients. Iron is a key metabolic driver in glioma biology impacting both the neoplastic cells directly and tumor progression through the immune system. We previously reported the expression of HFE (iron homeostatic) gene has a sexually dimorphic impact on survival in GBM patients. To further interrogate the relationship of iron and sexual dimorphism in GBMs, we expanded our analysis to include the ferrome (profile of genes and proteins involved in iron regulation). These genes were identified using IronChip and then interrogated for sexually dimorphic expression and survival in primary GBM patients using The Cancer Genome Atlas GBM database. Biological pathway analysis indicated that the Toll receptor signaling and immune system signaling pathways are most strongly correlated to sex difference and GBM patient survival. Toll like receptor 4 (TLR4) expression in the bottom quartile is associated with a modest survival benefit for female GBM patients compared to male, but in the top quartile of expression of this gene the survival for males is dramatically greater than that for females (18 months versus 3.6 months). Males expressing top quartile of Dual specificity phosphatases (DUSP1) also known as MKP1 survive 24.2 months whereas females survive 5.8 months. There is no sex effect of DUSP1 in the bottom quartile of expression. These data suggest that top quartile expression of TLR4/DUSP1 are both a benefit for males (extending their survival) and a detriment to females (shortening their survival). Both TLR4 and DUSP1 are richly expressed in macrophages. The data suggest that macrophage infiltration into the tumor could be particularly negatively impactful in females; a finding consistent with our previous reports for the HFE gene. In summary, iron related gene expression is linked to sexual dimorphism in survival and may provide insights on sex specific tumor biology.