Potential of PSMA-targeting radioligand therapy for malignant primary and secondary brain tumours using super-selective intra-arterial administration: a single centre, open label, non-randomised prospective imaging study.

BACKGROUND: The aim of this study was to provide quantitative evidence for the potential of PSMA-targeting radioligand therapy (RLT) as treatment approach for malignant brain tumours, and to explore whether tumour uptake could be enhanced by super-selective intra-arterial (ssIA)-administration. METHODS: Ten patients (n = 5 high-grade glioma, n = 5 brain metastasis) received 1.5 MBq/kg [68Ga]Ga-PSMA-11 intravenously and, within 7 days, intra-arterially (i.e., selectively in tumour-feeding arteries), followed twice by PET-MRI at 90, 165 and 240 min post-injection. Patient safety was monitored for each procedure. Standardised uptake values (SUVs) were obtained for tumour, healthy-brain, salivary glands and liver. Tumour-to-salivary-gland (T/SG) and tumour-to-liver (T/L) uptake-ratios were calculated. FINDINGS: No adverse events requiring study termination occurred. All patients showed uptake of [68Ga]Ga-PSMA-11 at the tumour site. Uptake was a median 15-fold higher following ssIA-administration (SUVmax median: 142.8, IQR: 102.8-245.9) compared to IV-administration (10.5, IQR:7.5-13.0). According to the bootstrap analysis, mean SUVmax after ssIA (168.8, 95% CI: 110.6-227.0) was well beyond the 95% confidence-interval of IV administration (10.5, 95% CI: 8.4-12.7). Uptake in healthy-brain was negligible, independent of administration route (SUVmean <0.1-0.1). Off-target uptake was comparable, resulting in more favourable T/SG- and T/L-ratios of 8.4 (IQR: 4.4-11.5) and 26.5 (IQR: 14.0-46.4) following ssIA, versus 0.5 (IQR: 0.4-0.7) and 1.8 (IQR: 1.0-2.7) for IV-administration. INTERPRETATION: ssIA-administration is safe and leads to a median fifteen-fold higher radioligand uptake at the tumour site, therewith qualifying more patients for treatment and enhancing the potential of therapy. These results open new avenues for the development of effective RLT-based treatment strategies for patients with brain tumours. FUNDING: Semmy Foundation.

An autologous ex vivo model for exploring patient-specific responses to viro-immunotherapy in glioblastoma.

Oncolytic virus (OV) clinical trials have demonstrated remarkable efficacy in subsets of patients with glioblastoma (GBM). However, the lack of tools to predict this response hinders the advancement of a more personalized application of OV therapy. In this study, we characterize an ex vivo co-culture system designed to examine the immune response to OV infection of patient-derived GBM neurospheres in the presence of autologous peripheral blood mononuclear cells (PBMCs). Co-culture conditions were optimized to retain viability and functionality of both tumor cells and PBMCs, effectively recapitulating the well-recognized immunosuppressive effects of GBM. Following OV infection, we observed elevated secretion of pro-inflammatory cytokines and chemokines, including interferon γ, tumor necrosis factor α, CXCL9, and CXCL10, and marked changes in immune cell activation markers. Importantly, OV treatment induced unique patient-specific immune responses. In summary, our co-culture platform presents an avenue for personalized screening of viro-immunotherapies in GBM, offering promise as a potential tool for future patient stratification in OV therapy.

[18F]FET PET/MRI: An Accurate Technique for Detection of Small Functional Pituitary Tumors.

Small functional pituitary tumors can cause severely disabling symptoms and early death. The gold standard diagnostic approach includes laboratory tests and MRI, with or without inferior petrosal sinus sampling (IPSS). In up to 40% of patients, however, the source of excess hormone production remains unidentified or uncertain. This excludes patients from surgical, Gamma Knife, and CyberKnife therapy and adversely affects overall cure rates. We here assess the diagnostic yield of O-(2-[18F]fluoroethyl)-l-tyrosine ([18F]FET) PET/MRI for detection of small functional pituitary tumors in these patients. Methods: This retrospective analysis included patients with Cushing disease (CD) but prior negative or inconclusive MRI results who underwent [18F]FET PET/MRI between February 1, 2021, and December 1, 2022. PET/MR images and MR images alone were evaluated by experienced nuclear radiologists, neuroradiologists, or radiologists. Postoperative tissue analysis (when performed) was used as a reference standard to assess diagnostic metrics (i.e., sensitivity and positive predictive value). Results were also compared with previously obtained MR images, preceding IPSS, and clinical or biochemical follow-up. Results: Twenty-two patients (68% female; mean age ± SD, 48 ± 15 y; range, 24-68 y) were scanned. All patients showed a clear metabolic focus on [18F]FET PET, whereas reading of the MRI alone yielded a suspected lesion in only 50%. Fifteen patients underwent surgery directed at the [18F]FET-positive focus. Tissue analysis confirmed a pituitary adenoma/pituitary neuroendocrine tumor of the corticotroph cell type (TPIT lineage) in 10 of 15 and a pituicytoma in 1 of 15, rendering a sensitivity of 100% and a positive predictive value of 73%. Lateralization was more accurate with [18F]FET PET/MRI than with IPSS in 33%. Twelve of 16 (75%) patients who received surgical, Gamma Knife, or CyberKnife therapy after [18F]FET PET/MRI reached short-term remission. Conclusion: [18F]FET PET/MRI shows a high diagnostic yield for localizing small functional pituitary tumors. This multimodal imaging technique provides a welcome improvement for diagnosis, planning of surgery, and clinical outcome in patients with Cushing disease, particularly those with repeated negative or inconclusive MRI results with or without IPSS.

PET Imaging and Protein Expression of Prostate-Specific Membrane Antigen in Glioblastoma: A Multicenter Inventory Study.

Upregulation of prostate-specific membrane antigen (PSMA) in neovasculature has been described in glioblastoma multiforme (GBM), whereas vasculature in nonaffected brain shows hardly any expression of PSMA. It is unclear whether PSMA-targeting tracer uptake on PET is based on PSMA-specific binding to neovasculature or aspecific uptake in tumor. Here, we quantified uptake of various PSMA-targeting tracers in GBM and correlated this with PSMA expression in tumor biopsy samples from the same patients. Methods: Fourteen patients diagnosed with de novo (n = 8) or recurrent (n = 6) GBM underwent a preoperative PET scan after injection of 1.5 MBq/kg [68Ga]Ga-PSMA-11 (n = 7), 200 MBq of [18F]DCFpyl (n = 3), or 200 MBq of [18F]PSMA-1007 (n = 4). Uptake in tumor and tumor-to-background ratios, with contralateral nonaffected brain as background, were determined. In a subset of patients, PSMA expression levels from different regions in the tumor tissue samples (n = 40), determined using immunohistochemistry (n = 35) or RNA sequencing (n = 13), were correlated with tracer uptake on PET. Results: Moderate to high (SUVmax, 1.3-20.0) heterogeneous uptake was found in all tumors irrespective of the tracer type used. Uptake in nonaffected brain was low, resulting in high tumor-to-background ratios (6.1-359.0) calculated by dividing SUVmax of tumor by SUVmax of background. Immunohistochemistry showed variable PSMA expression on endothelial cells of tumor microvasculature, as well as on dispersed individual cells (of unknown origin), and granular staining of the neuropil. No correlation was found between in vivo uptake and PSMA expression levels (for immunohistochemistry, r = -0.173, P = 0.320; for RNA, r = -0.033, P = 0.915). Conclusion: Our results indicate the potential use of various PSMA-targeting tracers in GBM. However, we found no correlation between PSMA expression levels on immunohistochemistry and uptake intensity on PET. Whether this may be explained by methodologic reasons, such as the inability to measure functionally active PSMA with immunohistochemistry, tracer pharmacokinetics, or the contribution of a disturbed blood-brain barrier to tracer retention, should still be investigated.

Mimics of Autoimmune Encephalitis: Validation of the 2016 Clinical Autoimmune Encephalitis Criteria.

BACKGROUND AND OBJECTIVES: The clinical criteria for autoimmune encephalitis (AE) were proposed by Graus et al. in 2016. In this study, the AE criteria were validated in the real world, and common AE mimics were described. In addition, criteria for probable anti-LGI1 encephalitis were proposed and validated. METHODS: In this retrospective cohort study, patients referred to our national referral center with suspicion of AE and specific neuroinflammatory disorders with similar clinical presentations were included from July 2016 to December 2019. Exclusion criteria were pure cerebellar or peripheral nerve system disorders. All patients were evaluated according to the AE criteria. RESULTS: In total, 239 patients were included (56% female; median age 42 years, range 1-85). AE was diagnosed in 104 patients (44%) and AE mimics in 109 patients (46%). The most common AE mimics and misdiagnoses were neuroinflammatory CNS disorders (26%), psychiatric disorders (19%), epilepsy with a noninflammatory cause (13%), CNS infections (7%), neurodegenerative diseases (7%), and CNS neoplasms (6%). Common confounding factors were mesiotemporal lesions on brain MRI (17%) and false-positive antibodies in serum (12%). Additional mesiotemporal features (involvement extralimbic structures, enhancement, diffusion restriction) were observed more frequently in AE mimics compared with AE (61% vs 24%; p = 0.005). AE criteria showed the following sensitivity and specificity: possible AE, 83% (95% CI 74-89) and 27% (95% CI 20-36); definite autoimmune limbic encephalitis (LE), 10% (95% CI 5-17) and 98% (95% CI 94-100); and probable anti-NMDAR encephalitis, 50% (95% CI 26-74) and 96% (95% CI 92-98), respectively. Specificity of the criteria for probable seronegative AE was 99% (95% CI 96-100). The newly proposed criteria for probable anti-LGI1 encephalitis showed a sensitivity of 66% (95% CI 47-81) and specificity of 96% (95% CI 93-98). DISCUSSION: AE mimics occur frequently. Common pitfalls in AE misdiagnosis are mesiotemporal lesions (predominantly with atypical features) and false-positive serum antibodies. As expected, the specificity of the criteria for possible AE is low because these criteria represent the minimal requirements for entry in the diagnostic algorithm for AE. Criteria for probable AE (-LGI1, -NMDAR, seronegative) and definite autoimmune LE are applicable for decisions on immunotherapy in early disease stage, as specificity is high.

Local delivery of hrBMP4 as an anticancer therapy in patients with recurrent glioblastoma: a first-in-human phase 1 dose escalation trial.

BACKGROUND: This Phase 1 study evaluates the intra- and peritumoral administration by convection enhanced delivery (CED) of human recombinant Bone Morphogenetic Protein 4 (hrBMP4) - an inhibitory regulator of cancer stem cells (CSCs) - in recurrent glioblastoma. METHODS: In a 3 + 3 dose escalation design, over four to six days, fifteen recurrent glioblastoma patients received, by CED, one of five doses of hrBMP4 ranging from 0·5 to 18 mg. Patients were followed by periodic physical, neurological, blood testing, magnetic resonance imaging (MRI) and quality of life evaluations. The primary objective of this first-in-human study was to determine the safety, dose-limiting toxicities (DLT) and maximum tolerated dose (MTD) of hrBMP4. Secondary objectives were to assess potential efficacy and systemic exposure to hrBMP4 upon intracerebral infusion. RESULTS: Intra- and peritumoral infusion of hrBMP4 was safe and well-tolerated. We observed no serious adverse events related to this drug. Neither MTD nor DLT were reached. Three patients had increased hrBMP4 serum levels at the end of infusion, which normalized within 4 weeks, without sign of toxicity. One patient showed partial response and two patients a complete (local) tumor response, which was maintained until the most recent follow-up, 57 and 30 months post-hrBMP4. Tumor growth was inhibited in areas permeated by hrBMP4. CONCLUSION: Local delivery of hrBMP4 in and around recurring glioblastoma is safe and well-tolerated. Three patients responded to the treatment. A complete response and long-term survival occurred in two of them. This warrants further clinical studies on this novel treatment targeting glioblastoma CSCs. TRIAL REGISTRATION: ClinicaTrials.gov identifier: NCT02869243.

Ex vivo drug sensitivity screening predicts response to temozolomide in glioblastoma patients and identifies candidate biomarkers.

BACKGROUND: Patient-derived glioma stem-like cells (GSCs) have become the gold-standard in neuro-oncological research; however, it remains to be established whether loss of in situ microenvironment affects the clinically-predictive value of this model. We implemented a GSC monolayer system to investigate in situ-in vitro molecular correspondence and the relationship between in vitro and patient response to temozolomide (TMZ). METHODS: DNA/RNA-sequencing was performed on 56 glioblastoma tissues and 19 derived GSC cultures. Sensitivity to TMZ was screened across 66 GSC cultures. Viability readouts were related to clinical parameters of corresponding patients and whole-transcriptome data. RESULTS: Tumour DNA and RNA sequences revealed strong similarity to corresponding GSCs despite loss of neuronal and immune interactions. In vitro TMZ screening yielded three response categories which significantly correlated with patient survival, therewith providing more specific prediction than the binary MGMT marker. Transcriptome analysis identified 121 genes related to TMZ sensitivity of which 21were validated in external datasets. CONCLUSION: GSCs retain patient-unique hallmark gene expressions despite loss of their natural environment. Drug screening using GSCs predicted patient response to TMZ more specifically than MGMT status, while transcriptome analysis identified potential biomarkers for this response. GSC drug screening therefore provides a tool to improve drug development and precision medicine for glioblastoma.

Quantitative proteomics of small numbers of closely-related cells: Selection of the optimal method for a clinical setting.

Mass spectrometry (MS)-based proteomics profiling has undoubtedly increased the knowledge about cellular processes and functions. However, its applicability for paucicellular sample analyses is currently limited. Although new approaches have been developed for single-cell studies, most of them have not (yet) been standardized and/or require highly specific (often home-built) devices, thereby limiting their broad implementation, particularly in non-specialized settings. To select an optimal MS-oriented proteomics approach applicable in translational research and clinical settings, we assessed 10 different sample preparation procedures in paucicellular samples of closely-related cell types. Particularly, five cell lysis protocols using different chemistries and mechanical forces were combined with two sample clean-up techniques (C18 filter- and SP3-based), followed by tandem mass tag (TMT)-based protein quantification. The evaluation was structured in three phases: first, cell lines from hematopoietic (THP-1) and non-hematopoietic (HT-29) origins were used to test the approaches showing the combination of a urea-based lysis buffer with the SP3 bead-based clean-up system as the best performer. Parameters such as reproducibility, accessibility, spatial distribution, ease of use, processing time and cost were considered. In the second phase, the performance of the method was tested on maturation-related cell populations: three different monocyte subsets from peripheral blood and, for the first time, macrophages/microglia (MAC) from glioblastoma samples, together with T cells from both tissues. The analysis of 50,000 cells down to only 2,500 cells revealed different protein expression profiles associated with the distinct cell populations. Accordingly, a closer relationship was observed between non-classical monocytes and MAC, with the latter showing the co-expression of M1 and M2 macrophage markers, although pro-tumoral and anti-inflammatory proteins were more represented. In the third phase, the results were validated by high-end spectral flow cytometry on paired monocyte/MAC samples to further determine the sensitivity of the MS approach selected. Finally, the feasibility of the method was proven in 194 additional samples corresponding to 38 different cell types, including cells from different tissue origins, cellular lineages, maturation stages and stimuli. In summary, we selected a reproducible, easy-to-implement sample preparation method for MS-based proteomic characterization of paucicellular samples, also applicable in the setting of functionally closely-related cell populations.

Complicated Clinical Course in Incipient Gigantism Due to Treatment-resistant Aryl Hydrocarbon Receptor-Interacting Protein-mutated Pediatric Somatotropinoma.

Background: Our objective was to describe the clinical course and treatment challenges in a very young patient with a pituitary adenoma due to a novel aryl hydrocarbon receptor-interacting protein (AIP) gene mutation, highlighting the limitations of somatostatin receptor immunohistochemistry to predict clinical responses to somatostatin analogs in acromegaly. Case Report: We report the case of a 7-year-old boy presenting with headache, visual field defects, and accelerated growth following failure to thrive. The laboratory results showed high insulin-like growth factor I (IGF-I) (standardised deviation scores ( +3.49) and prolactin levels (0.5 nmol/L), and magnetic resonance imaging identified a pituitary macroadenoma. Tumoral/hormonal control could not be achieved despite 3 neurosurgical procedures, each time with apparent total resection or with lanreotide or pasireotide. IGF-I levels decreased with the GH receptor antagonist pegvisomant. The loss of somatostatin receptor 5 was observed between the second and third tumor resection. In vitro, no effect on tumoral GH release by pasireotide (with/without cabergoline) was observed. Genetic analysis revealed a novel germline AIP mutation: p.Tyr202∗ (pathogenic; class 4). Discussion: In vitro response of tumor tissue to somatostatin may better predict tumoral in vivo responses of somatostatin analogs than somatostatin receptor immunohistochemistry. Conclusion: We identified a novel pathologic AIP mutation that was associated with incipient acrogigantism in an extremely young patient who had a complicated course of disease. Growth acceleration can be masked due to failure to thrive. Tumoral growth hormone release in vivo may be predicted with in vitro exposure to somatostatin receptor analogs, as it cannot be assumed that all AIP-mutated somatotropinomas respond well to pasireotide.

Between-hospital variation in time to glioblastoma surgery: a report from the Quality Registry Neuro Surgery in the Netherlands.

OBJECTIVE: Patients with glioblastoma are often scheduled for urgent elective surgery. Currently, the impact of the waiting period until glioblastoma surgery is undetermined. In this national quality registry study, the authors determined the wait times until surgery for patients with glioblastoma, the risk factors associated with wait times, and the risk-standardized variation in time to surgery between Dutch hospitals. The associations between time to surgery and patient outcomes were also explored. METHODS: Data from all 4589 patients who underwent first-time glioblastoma surgery between 2014 and 2019 in the Netherlands were collected by 13 hospitals in the Quality Registry Neuro Surgery. Time to surgery comprised 1) the time from first MR scan to surgery (MTS), and 2) the time from first neurosurgical consultation to surgery (CTS). Long MTS was defined as more than 21 days and long CTS as more than 14 days. Potential risk factors were analyzed in multivariable logistic regression models. The standardized rate of long time to surgery was analyzed using funnel plots. Patient outcomes including Karnofsky Performance Scale (KPS) score change, complications, and survival were analyzed by multivariable logistic regression and proportional hazards models. RESULTS: The median overall MTS and CTS were 18 and 9 days, respectively. Overall, 2576 patients (56%) had an MTS within 3 weeks and 3069 (67%) had a CTS within 2 weeks. Long MTS was significantly associated with older age, higher preoperative KPS score, higher American Society of Anesthesiologists comorbidity class, season, lower hospital case volume, university affiliation, and resection. Long CTS was significantly associated with higher baseline KPS score, university affiliation, resection, more recent year of treatment, and season. In funnel plots, considerable practice variation was observed between hospitals in patients with long times to surgery. Fewer patients with KPS score improvement were observed after a long time until resection. Long CTS was associated with longer survival. Complications and KPS score decline were not associated with time to surgery. CONCLUSIONS: Considerable between-hospital variation among Dutch hospitals was observed in the time to glioblastoma surgery. A long time to resection impeded KPS score improvement, and therefore, patients who may improve should be identified for more urgent resection. Longer survival was observed in patients selected for longer time until surgery after neurosurgical consultation (CTS).

Intraoperative B-Mode Ultrasound Guided Surgery and the Extent of Glioblastoma Resection: A Randomized Controlled Trial.

BACKGROUND: Intraoperative MRI and 5-aminolaevulinic acid guided surgery are useful to maximize the extent of glioblastoma resection. Intraoperative ultrasound is used as a time-and cost-effective alternative, but its value has never been assessed in a trial. The goal of this randomized controlled trial was to assess the value of intraoperative B-mode ultrasound guided surgery on the extent of glioblastoma resection. MATERIALS AND METHODS: In this randomized controlled trial, patients of 18 years or older with a newly diagnosed presumed glioblastoma, deemed totally resectable, presenting at the Erasmus MC (Rotterdam, The Netherlands) were enrolled and randomized (1:1) into intraoperative B-mode ultrasound guided surgery or resection under standard neuronavigation. The primary outcome of this study was complete contrast-enhancing tumor resection, assessed quantitatively by a blinded neuroradiologist on pre- and post-operative MRI scans. This trial was registered with ClinicalTrials.gov (NCT03531333). RESULTS: We enrolled 50 patients between November 1, 2016 and October 30, 2019. Analysis was done in 23 of 25 (92%) patients in the intraoperative B-mode ultrasound group and 24 of 25 (96%) patients in the standard surgery group. Eight (35%) of 23 patients in the intraoperative B-mode ultrasound group and two (8%) of 24 patients in the standard surgery group underwent complete resection (p=0.036). Baseline characteristics, neurological outcome, functional performance, quality of life, complication rates, overall survival and progression-free survival did not differ between treatment groups (p>0.05). CONCLUSIONS: Intraoperative B-mode ultrasound enables complete resection more often than standard surgery without harming patients and can be considered to maximize the extent of glioblastoma resection during surgery.

Between-hospital variation in rates of complications and decline of patient performance after glioblastoma surgery in the dutch Quality Registry Neuro Surgery.

INTRODUCTION: For decisions on glioblastoma surgery, the risk of complications and decline in performance is decisive. In this study, we determine the rate of complications and performance decline after resections and biopsies in a national quality registry, their risk factors and the risk-standardized variation between institutions. METHODS: Data from all 3288 adults with first-time glioblastoma surgery at 13 hospitals were obtained from a prospective population-based Quality Registry Neuro Surgery in the Netherlands between 2013 and 2017. Patients were stratified by biopsies and resections. Complications were categorized as Clavien-Dindo grades II and higher. Performance decline was considered a deterioration of more than 10 Karnofsky points at 6 weeks. Risk factors were evaluated in multivariable logistic regression analysis. Patient-specific expected and observed complications and performance declines were summarized for institutions and analyzed in funnel plots. RESULTS: For 2271 resections, the overall complication rate was 20 % and 16 % declined in performance. For 1017 biopsies, the overall complication rate was 11 % and 30 % declined in performance. Patient-related characteristics were significant risk factors for complications and performance decline, i.e. higher age, lower baseline Karnofsky, higher ASA classification, and the surgical procedure. Hospital characteristics, i.e. case volume, university affiliation and biopsy percentage, were not. In three institutes the observed complication rate was significantly less than expected. In one institute significantly more performance declines were observed than expected, and in one institute significantly less. CONCLUSIONS: Patient characteristics, but not case volume, were risk factors for complications and performance decline after glioblastoma surgery. After risk-standardization, hospitals varied in complications and performance declines.

Malignant Glioma In Vitro Models: On the Utilization of Stem-like Cells.

Recent publications on the molecular characterization of malignant glioma have had profound impact on the appreciation of tumoral heterogeneity within and between patients. Both these phenomena are implicated in the variability in clinical outcome between patients, as well as the inevitable recurrence of these tumors after conventional treatment. The advent of selective cell culture protocols for the propagation of patient-derived glioma stem-like cells (GSCs) provides researchers the ability to selectively study the cells that could be at the root of tumor proliferation and resistance to therapy. As these techniques are widely applied in contemporary studies and becoming the preferred in vitro model, molecular characterization of GSCs is considered pivotal for the identification and advancement of novel therapies for this devastating disease. This review aims to provide an overview of canonical molecular alterations defining subtypes of malignant glioma as derived from genotypic, transcriptomic and epigenetic profiling in relation to their representation in GSC models. The distribution of these hallmark alterations as found in characterization studies of GSCs is compared between publications. Finally, conclusions of these studies with respect to coverage of driving alterations and translational relevance are provided. By doing so, we provide a contemporary overview of scientific results derived from GSC models and hopefully create appreciation of the advantages and caveats of utilizing these models for studying malignant glioma.

ABT-888 enhances cytotoxic effects of temozolomide independent of MGMT status in serum free cultured glioma cells.

BACKGROUND: The current standard of care for Glioblastoma Multiforme (GBM) consists of fractionated focal irradiation with concomitant temozolomide (TMZ) chemotherapy. A promising strategy to increase the efficacy of TMZ is through interference with the DNA damage repair machinery, by poly(ADP-ribose) polymerase protein inhibition(PARPi). The objective of the present study was to investigate the therapeutic benefit of combination therapy in patient-derived glioma stem-like cells (GSC). METHODS: Combination therapy feasibility was tested on established GBM cell lines U373 and T98. We developed an in vitro drug-screening assay based on GSC cultures derived from a panel of primary patient tissue samples (n = 20) to evaluate the effect of PARPi (ABT-888) monotherapy and combination therapy with TMZ. Therapeutic effect was assessed by viability, double stranded breaks, apoptosis and autophagy assays and longitudinal microscopic cell monitoring was performed. O-6-methylguanine-DNA methyltransferase (MGMT) status was determined by methylation assay and protein expression by western blots. RESULTS: PARPi monotherapy was found to decrease viability by more than 25% in 4 of the 20 GSCs (20%) at 10 µM. TMZ monotherapy at 50 µM and 100 µM was effective in 12 and 14 of the 20 GSCs, respectively. TMZ resistance to 100 µM was found in 7 of 8 MGMT protein positive cultures. Potentiation of TMZ therapy through PARPi was found in 90% (n = 20) of GSCs, of which 6 were initially resistant and 7 were sensitive to TMZ monotherapy. Increased induction of double stranded breaks and apoptosis were noted in responsive GSCs. There was a trend noted, albeit statistically insignificant, of increased autophagy both in western blots and accumulation of autophagosomes. CONCLUSION: PARPi mediated potentiation of TMZ is independent of TMZ sensitivity and can override MGMT(-) mediated resistance when administered simultaneously. Response to combination therapy was associated with increased double strand breaks induction, and coincided by increased apoptosis and autophagy. PARPi addition potentiates TMZ treatment in primary GSCs. PARPi could potentially enhance the therapeutic efficacy of the standard of care in GBM.

Locally-delivered T-cell-derived cellular vehicles efficiently track and deliver adenovirus delta24-RGD to infiltrating glioma.

Oncolytic adenoviral vectors are a promising alternative for the treatment of glioblastoma. Recent publications have demonstrated the advantages of shielding viral particles within cellular vehicles (CVs), which can be targeted towards the tumor microenvironment. Here, we studied T-cells, often having a natural capacity to target tumors, for their feasibility as a CV to deliver the oncolytic adenovirus, Delta24-RGD, to glioblastoma. The Jurkat T-cell line was assessed in co-culture with the glioblastoma stem cell (GSC) line, MGG8, for the optimal transfer conditions of Delta24-RGD in vitro. The effect of intraparenchymal and tail vein injections on intratumoral virus distribution and overall survival was addressed in an orthotopic glioma stem cell (GSC)-based xenograft model. Jurkat T-cells were demonstrated to facilitate the amplification and transfer of Delta24-RGD onto GSCs. Delta24-RGD dosing and incubation time were found to influence the migratory ability of T-cells towards GSCs. Injection of Delta24-RGD-loaded T-cells into the brains of GSC-bearing mice led to migration towards the tumor and dispersion of the virus within the tumor core and infiltrative zones. This occurred after injection into the ipsilateral hemisphere, as well as into the non-tumor-bearing hemisphere. We found that T-cell-mediated delivery of Delta24-RGD led to the inhibition of tumor growth compared to non-treated controls, resulting in prolonged survival (p = 0.007). Systemic administration of virus-loaded T-cells resulted in intratumoral viral delivery, albeit at low levels. Based on these findings, we conclude that T-cell-based CVs are a feasible approach to local Delta24-RGD delivery in glioblastoma, although efficient systemic targeting requires further improvement.

Serum-free culture success of glial tumors is related to specific molecular profiles and expression of extracellular matrix-associated gene modules.

BACKGROUND: Recent molecular characterization studies have identified clinically relevant molecular subtypes to coexist within the same histological entities of glioma. Comparative studies between serum-supplemented and serum-free (SF) culture conditions have demonstrated that SF conditions select for glioma stem-like cells, which superiorly conserve genomic alterations. However, neither the representation of molecular subtypes within SF culture assays nor the molecular distinctions between successful and nonsuccessful attempts have been elucidated. METHODS: A cohort of 261 glioma samples from varying histological grades was documented for SF culture success and clinical outcome. Gene expression and single nucleotide polymorphism arrays were interrogated on a panel of tumors for comparative analysis of SF+ (successful cultures) and SF- (unsuccessful cultures). RESULTS: SF culture outcome was correlated with tumor grade, while no relation was found between SF+ and patient overall survival. Copy number-based hierarchical clustering revealed an absolute separation between SF+ and SF- parental tumors. All SF+ cultures are derived from tumors that are isocitrate dehydrogenase 1 (IDH1) wild type, chromosome 7 amplified, and chromosome 10q deleted. SF- cultures derived from IDH1 mutant tumors demonstrated a fade-out of mutated cells during the first passages. SF+ tumors were enriched for The Cancer Genome Atlas Classical subtype and intrinsic glioma subtype-18. Comparative gene ontology analysis between SF+ and SF- tumors demonstrated enrichment for modules associated with extracellular matrix composition, Hox-gene signaling, and inflammation. CONCLUSION: SF cultures are derived from a subset of parental tumors with a shared molecular background including enrichment for extracellular matrix-associated gene modules. These results provide leads to develop enhanced culture protocols for glioma samples not propagatable under current SF conditions.

Laser speckle imaging identification of increases in cortical microcirculatory blood flow induced by motor activity during awake craniotomy.

OBJECT: The goal of awake neurosurgery is to maximize resection of brain lesions with minimal injury to functional brain areas. Laser speckle imaging (LSI) is a noninvasive macroscopic technique with high spatial and temporal resolution used to monitor changes in capillary perfusion. In this study, the authors hypothesized that LSI can be useful as a noncontact method of functional brain mapping during awake craniotomy for tumor removal. Such a modality would be an advance in this type of neurosurgery since current practice involves the application of invasive intraoperative single-point electrocortical (electrode) stimulation and measurements. METHODS: After opening the dura mater, patients were woken up, and LSI was set up to image the exposed brain area. Patients were instructed to follow a rest-activation-rest protocol in which activation consisted of the hand-clenching motor task. Subsequently, exposed brain areas were mapped for functional motor areas by using standard electrocortical stimulation (ECS). Changes in the LSI signal were analyzed offline and compared with the results of ECS. RESULTS: In functional motor areas of the hand mapped with ECS, cortical blood flow measured using LSI significantly increased from 2052 ± 818 AU to 2471 ± 675 AU during hand clenching, whereas capillary blood flow did not change in the control regions (areas mapped using ECS with no functional activity). CONCLUSIONS: The main finding of this study was that changes in laser speckle perfusion as a measure of cortical microvascular blood flow when performing a motor task with the hand relate well to the ECS map. The authors have shown the feasibility of using LSI for direct visualization of cortical microcirculatory blood flow changes during neurosurgery.