Neurosurgical application of olaparib from a thermo-responsive paste potentiates DNA damage to prolong survival in malignant glioma.

BACKGROUND: There is increased pan-cancer specific interest in repurposing the poly adenosine diphosphate-ribose polymerase-1 (PARP-1) inhibitor, olaparib, for newly diagnosed or recurrent isocitrate dehydrogenase wild type glioblastoma. We explore whether intra-cavity delivery of olaparib confers a survival benefit in a pre-clinical high-grade glioma model. METHODS: Primary tumor RNA sequencing data was used to determine PARP-1 as a target in the glioblastoma infiltrative margin. We assessed radiosensitization conferred by olaparib alone and concomitant to genotoxic insults in vitro using clonal growth assays, cell cycle analysis and immunocytochemistry, and in vivo upon post-surgical delivery from a temperature-sensitive polymeric paste. RESULTS: RNA-sequencing confirmed PARP-1 as a viable therapy target in glioblastoma infiltrative disease. Acute exposure of glioma cells to olaparib impaired proliferation and induced late-stage apoptosis associated with DNA damage in vitro, potentiated by radiation. Using high-grade glioma orthotopic allografts, a long-term overall survival benefit was observed upon interstitial olaparib delivery concomitant with radiotherapy, compared to systemic olaparib and standard glioblastoma treatment. Combined delivery of olaparib with either temozolomide or etoposide increased long-term survival, suggestive of olaparib functioning as DNA damage sensitizer. CONCLUSIONS: Collectively, our data support a rationale for localized olaparib delivery concomitant with the current clinical regimen for malignant glioma treatment.

Bipolar electrochemical growth of conductive microwires for cancer spheroid integration: a step forward in conductive biological circuitry.

The field of bioelectronics is developing exponentially. There is now a drive to interface electronics with biology for the development of new technologies to improve our understanding of electrical forces in biology. This builds on our recently published work in which we show wireless electrochemistry could be used to grow bioelectronic functional circuitry in 2D cell layers. To date our ability to merge electronics with in situ with biology is 3D limited. In this study, we aimed to further develop the wireless electrochemical approach for the self-assembly of microwires in situ with custom-designed and fabricated 3D cancer spheroids. Unlike traditional electrochemical methods that rely on direct electrical connections to induce currents, our technique utilises bipolar electrodes that operate independently of physical wired connections. These electrodes enable redox reactions through the application of an external electric field. Specifically, feeder electrodes connected to a power supply generate an electric field, while the bipolar electrodes, not physically connected to the feeder electrodes, facilitate the reduction of silver ions from the solution. This process occurs upon applying a voltage across the feeder electrodes, resulting in the formation of self-assembled microwires between the cancer spheroids.Thereby, creating interlinked bioelectronic circuitry with cancer spheroids. We demonstrate that a direct current was needed to stimulate the growth of conductive microwires in the presence of cell spheroids. Microwire growth was successful when using 50 V (0.5 kV/cm) of DC applied to a single spheroid of approximately 800 µm in diameter but could not be achieved with alternating currents. This represents the first proof of the concept of using wireless electrochemistry to grow conductive structures with 3D mammalian cell spheroids.

Radiogenomics as an Integrated Approach to Glioblastoma Precision Medicine.

PURPOSE OF REVIEW: Isocitrate dehydrogenase wild-type glioblastoma is the most aggressive primary brain tumour in adults. Its infiltrative nature and heterogeneity confer a dismal prognosis, despite multimodal treatment. Precision medicine is increasingly advocated to improve survival rates in glioblastoma management; however, conventional neuroimaging techniques are insufficient in providing the detail required for accurate diagnosis of this complex condition. RECENT FINDINGS: Advanced magnetic resonance imaging allows more comprehensive understanding of the tumour microenvironment. Combining diffusion and perfusion magnetic resonance imaging to create a multiparametric scan enhances diagnostic power and can overcome the unreliability of tumour characterisation by standard imaging. Recent progress in deep learning algorithms establishes their remarkable ability in image-recognition tasks. Integrating these with multiparametric scans could transform the diagnosis and monitoring of patients by ensuring that the entire tumour is captured. As a corollary, radiomics has emerged as a powerful approach to offer insights into diagnosis, prognosis, treatment, and tumour response through extraction of information from radiological scans, and transformation of these tumour characteristics into quantitative data. Radiogenomics, which links imaging features with genomic profiles, has exhibited its ability in characterising glioblastoma, and determining therapeutic response, with the potential to revolutionise management of glioblastoma. The integration of deep learning algorithms into radiogenomic models has established an automated, highly reproducible means to predict glioblastoma molecular signatures, further aiding prognosis and targeted therapy. However, challenges including lack of large cohorts, absence of standardised guidelines and the 'black-box' nature of deep learning algorithms, must first be overcome before this workflow can be applied in clinical practice.

Free drug and ROS-responsive nanoparticle delivery of synergistic doxorubicin and olaparib combinations to triple negative breast cancer models.

Combinations of the topoisomerase II inhibitor doxorubicin and the poly (ADP-ribose) polymerase inhibitor olaparib offer potential drug-drug synergy for the treatment of triple negative breast cancers (TNBC). In this study we performed in vitro screening of combinations of these drugs, administered directly or encapsulated within polymer nanoparticles, in both 2D and in 3D spheroid models of breast cancer. A variety of assays were used to evaluate drug potency, and calculations of combination index (CI) values indicated that synergistic effects of drug combinations occurred in a molar-ratio dependent manner. It is suggested that the mechanisms of synergy were related to enhancement of DNA damage as shown by the level of double-strand DNA breaks, and mechanisms of antagonism associated with mitochondrial mediated cell survival, as indicated by reactive oxygen species (ROS) generation. Enhanced drug delivery and potency was observed with nanoparticle formulations, with a greater extent of doxorubicin localised to cell nuclei as evidenced by microscopy, and higher cytotoxicity at the same time points compared to free drugs. Together, the work presented identifies specific combinations of doxorubicin and olaparib which were most effective in a panel of TNBC cell lines, explores the mechanisms by which these combined agents might act, and shows that formulation of these drug combinations into polymeric nanoparticles at specific ratios conserves synergistic action and enhanced potency in vitro compared to the free drugs.

Wireless electrical-molecular quantum signalling for cancer cell apoptosis.

Quantum biological tunnelling for electron transfer is involved in controlling essential functions for life such as cellular respiration and homoeostasis. Understanding and controlling the quantum effects in biology has the potential to modulate biological functions. Here we merge wireless nano-electrochemical tools with cancer cells for control over electron transfer to trigger cancer cell death. Gold bipolar nanoelectrodes functionalized with redox-active cytochrome c and a redox mediator zinc porphyrin are developed as electric-field-stimulating bio-actuators, termed bio-nanoantennae. We show that a remote electrical input regulates electron transport between these redox molecules, which results in quantum biological tunnelling for electron transfer to trigger apoptosis in patient-derived cancer cells in a selective manner. Transcriptomics data show that the electric-field-induced bio-nanoantenna targets the cancer cells in a unique manner, representing electrically induced control of molecular signalling. The work shows the potential of quantum-based medical diagnostics and treatments.

Chain-extension in hyperbranched polymers alters tissue distribution and cytotoxicity profiles in orthotopic models of triple negative breast cancers.

The therapeutic efficacy of nanomedicines is highly dependent on their access to target sites in the body, and this in turn is markedly affected by their size, shape and transport properties in tissue. Although there have been many studies in this area, the ability to design nanomaterials with optimal physicochemical properties for in vivo efficacy remains a significant challenge. In particular, it is often difficult to quantify the detailed effects of cancer drug delivery systems in vivo as tumour volume reduction, a commonly reported marker of efficacy, does not always correlate with cytotoxicity in tumour tissue. Here, we studied the behaviour in vivo of two specific poly(2-hydroxypropyl methacrylamide) (pHPMA) pro-drugs, with hyperbranched and chain-extended branched architectures, redox-responsive backbone components, and pH-sensitive linkers to the anti-cancer drug doxorubicin. Evaluation of the biodistribution of these polymers following systemic injection indicated differences in the circulation time and organ distribution of the two polymers, despite their very similar hydrodynamic radii (∼10 and 15 nm) and architectures. In addition, both polymers showed improved tumour accumulation in orthotopic triple-negative breast cancers in mice, and decreased accumulation in healthy tissue, as compared to free doxorubicin, even though neither polymer-doxorubicin pro-drug decreased overall tumour volume as much as the free drug under the dosing regimens selected. However, the results of histopathological examinations by haematoxylin and eosin, and TUNEL staining indicated a higher population of apoptotic cells in the tumours for both polymer pro-drug treatments, and in turn a lower population of apoptotic cells in the heart, liver and spleen, as compared to free doxorubicin treatment. These data suggest that the penetration of these polymer pro-drugs was enhanced in tumour tissue relative to free doxorubicin, and that the combination of size, architecture, bioresponsive backbone and drug linker degradation yielded greater efficacy for the polymers as measured by biomarkers than that of tumour volume. We suggest therefore that the effects of nanomedicines may be different at various length scales relative to small molecule free drugs, and that penetration into tumour tissue for some nanomedicines may not be as problematic as prior reports have suggested. Furthermore, the data indicate that dual-responsive crosslinked polymer-prodrugs in this study may be effective nanomedicines for breast cancer chemotherapy, and that endpoints beyond tumour volume reduction can be valuable in selecting candidates for pre-clinical trials.

Spatially resolved transcriptomic profiles reveal unique defining molecular features of infiltrative 5ALA-metabolizing cells associated with glioblastoma recurrence.

BACKGROUND: Spatiotemporal heterogeneity originating from genomic and transcriptional variation was found to contribute to subtype switching in isocitrate dehydrogenase-1 wild-type glioblastoma (GBM) prior to and upon recurrence. Fluorescence-guided neurosurgical resection utilizing 5-aminolevulinic acid (5ALA) enables intraoperative visualization of infiltrative tumors outside the magnetic resonance imaging contrast-enhanced regions. The cell population and functional status of tumor responsible for enhancing 5ALA-metabolism to fluorescence-active PpIX remain elusive. The close spatial proximity of 5ALA-metabolizing (5ALA +) cells to residual disease remaining post-surgery renders 5ALA + biology an early a priori proxy of GBM recurrence, which is poorly understood. METHODS: We performed spatially resolved bulk RNA profiling (SPRP) analysis of unsorted Core, Rim, Invasive margin tissue, and FACS-isolated 5ALA + /5ALA - cells from the invasive margin across IDH-wt GBM patients (N = 10) coupled with histological, radiographic, and two-photon excitation fluorescence microscopic analyses. Deconvolution of SPRP followed by functional analyses was performed using CIBEROSRTx and UCell enrichment algorithms, respectively. We further investigated the spatial architecture of 5ALA + enriched regions by analyzing spatial transcriptomics from an independent IDH-wt GBM cohort (N = 16). Lastly, we performed survival analysis using Cox Proportinal-Hazards model on large GBM cohorts. RESULTS: SPRP analysis integrated with single-cell and spatial transcriptomics uncovered that the GBM molecular subtype heterogeneity is likely to manifest regionally in a cell-type-specific manner. Infiltrative 5ALA + cell population(s) harboring transcriptionally concordant GBM and myeloid cells with mesenchymal subtype, -active wound response, and glycolytic metabolic signature, was shown to reside within the invasive margin spatially distinct from the tumor core. The spatial co-localization of the infiltrating MES GBM and myeloid cells within the 5ALA + region indicates PpIX fluorescence can effectively be utilized to resect the immune reactive zone beyond the tumor core. Finally, 5ALA + gene signatures were associated with poor survival and recurrence in GBM, signifying that the transition from primary to recurrent GBM is not discrete but rather a continuum whereby primary infiltrative 5ALA + remnant tumor cells more closely resemble the eventual recurrent GBM. CONCLUSIONS: Elucidating the unique molecular and cellular features of the 5ALA + population within tumor invasive margin opens up unique possibilities to develop more effective treatments to delay or block GBM recurrence, and warrants commencement of such treatments as early as possible post-surgical resection of the primary neoplasm.

Untargeted Metabolomic Characterization of Glioblastoma Intra-Tumor Heterogeneity Using OrbiSIMS.

Glioblastoma (GBM) is an incurable brain cancer with a median survival of less than two years from diagnosis. The standard treatment of GBM is multimodality therapy comprising surgical resection, radiation, and chemotherapy. However, prognosis remains poor, and there is an urgent need for effective anticancer drugs. Since different regions of a single GBM contain multiple cancer subpopulations ("intra-tumor heterogeneity"), this likely accounts for therapy failure as certain cancer cells can escape from immune surveillance and therapeutic threats. Here, we present metabolomic data generated using the Orbitrap secondary ion mass spectrometry (OrbiSIMS) technique to investigate brain tumor metabolism within its highly heterogeneous tumor microenvironment. Our results demonstrate that an OrbiSIMS-based untargeted metabolomics method was able to discriminate morphologically distinct regions (viable, necrotic, and non-cancerous) within single tumors from formalin-fixed paraffin-embedded tissue archives. Specifically, cancer cells from necrotic regions were separated from viable GBM cells based on a set of metabolites including cytosine, phosphate, purine, xanthine, and 8-hydroxy-7-methylguanine. Moreover, we mapped ubiquitous metabolites across necrotic and viable regions into metabolic pathways, which allowed for the discovery of tryptophan metabolism that was likely essential for GBM cellular survival. In summary, this study first demonstrated the capability of OrbiSIMS for in situ investigation of GBM intra-tumor heterogeneity, and the acquired information can potentially help improve our understanding of cancer metabolism and develop new therapies that can effectively target multiple subpopulations within a tumor.

Low-Density Lipoprotein Pathway Is a Ubiquitous Metabolic Vulnerability in High Grade Glioma Amenable for Nanotherapeutic Delivery.

Metabolic reprogramming, through increased uptake of cholesterol in the form of low-density lipoproteins (LDL), is one way by which cancer cells, including high grade gliomas (HGG), maintain their rapid growth. In this study, we determined LDL receptor (LDLR) expression in HGGs using immunohistochemistry on tissue microarrays from intra- and inter tumour regions of 36 adult and 133 paediatric patients to confirm LDLR as a therapeutic target. Additionally, we analysed expression levels in three representative cell line models to confirm their future utility to test LDLR-targeted nanoparticle uptake, retention, and cytotoxicity. Our data show widespread LDLR expression in adult and paediatric cohorts, but with significant intra-tumour variation observed between the core and either rim or invasive regions of adult HGG. Expression was independent of paediatric tumour grade or identified clinicopathological factors. LDLR-expressing tumour cells localized preferentially within perivascular niches, also with significant adult intra-tumour variation. We demonstrated variable levels of LDLR expression in all cell lines, confirming their suitability as models to test LDLR-targeted nanotherapy delivery. Overall, our study reveals the LDLR pathway as a ubiquitous metabolic vulnerability in high grade gliomas across all ages, amenable to future consideration of LDL-mediated nanoparticle/drug delivery to potentially circumvent tumour heterogeneity.

Childhood Brain Tumors: A Review of Strategies to Translate CNS Drug Delivery to Clinical Trials.

Brain and spinal tumors affect 1 in 1000 people by 25 years of age, and have diverse histological, biological, anatomical and dissemination characteristics. A mortality of 30-40% means the majority are cured, although two-thirds have life-long disability, linked to accumulated brain injury that is acquired prior to diagnosis, and after surgery or chemo-radiotherapy. Only four drugs have been licensed globally for brain tumors in 40 years and only one for children. Most new cancer drugs in clinical trials do not cross the blood-brain barrier (BBB). Techniques to enhance brain tumor drug delivery are explored in this review, and cover those that augment penetration of the BBB, and those that bypass the BBB. Developing appropriate delivery techniques could improve patient outcomes by ensuring efficacious drug exposure to tumors (including those that are drug-resistant), reducing systemic toxicities and targeting leptomeningeal metastases. Together, this drug delivery strategy seeks to enhance the efficacy of new drugs and enable re-evaluation of existing drugs that might have previously failed because of inadequate delivery. A literature review of repurposed drugs is reported, and a range of preclinical brain tumor models available for translational development are explored.

Stimuli-Responsive Multifunctional Nanomedicine for Enhanced Glioblastoma Chemotherapy Augments Multistage Blood-to-Brain Trafficking and Tumor Targeting.

Minimal therapeutic advances have been achieved over the past two decades for glioblastoma (GBM), which remains an unmet clinical need. Here, hypothesis-driven stimuli-responsive nanoparticles (NPs) for docetaxel (DTX) delivery to GBM are reported, with multifunctional features that circumvent insufficient blood-brain barrier (BBB) trafficking and lack of GBM targeting-two major hurdles for anti-GBM therapies. NPs are dual-surface tailored with a i) brain-targeted acid-responsive Angiopep-2 moiety that triggers NP structural rearrangement within BBB endosomal vesicles, and ii) L-Histidine moiety that provides NP preferential accumulation into GBM cells post-BBB crossing. In tumor invasive margin patient cells, the stimuli-responsive multifunctional NPs target GBM cells, enhance cell uptake by 12-fold, and induce three times higher cytotoxicity in 2D and 3D cell models. Moreover, the in vitro BBB permeability is increased by threefold. A biodistribution in vivo trial confirms a threefold enhancement of NP accumulation into the brain. Last, the in vivo antitumor efficacy is validated in GBM orthotopic models following intratumoral and intravenous administration. Median survival and number of long-term survivors are increased by 50%. Altogether, a preclinical proof of concept supports these stimuli-responsive multifunctional NPs as an effective anti-GBM multistage chemotherapeutic strategy, with ability to respond to multiple fronts of the GBM microenvironment.

Evaluation of cannabidiol nanoparticles and nanoemulsion biodistribution in the central nervous system after intrathecal administration for the treatment of pain.

We investigated how the biodistribution of cannabidiol (CBD) within the central nervous system (CNS) is influenced by two different formulations, an oil-in-water (O/W) nanoemulsion and polymer-coated nanoparticles (PCNPs). We observed that both CBD formulations administered were preferentially retained in the spinal cord, with high concentrations reaching the brain within 10 min of administration. The CBD nanoemulsion reached Cmax in the brain at 210 ng/g within 120 min (Tmax), whereas the CBD PCNPs had a Cmax of 94 ng/g at 30 min (Tmax), indicating that rapid brain delivery can be achieved through the use of PCNPs. Moreover, the AUC0-4h of CBD in the brain was increased 3.7-fold through the delivery of the nanoemulsion as opposed to the PCNPs, indicating higher retention of CBD at this site. Both formulations exhibited immediate anti-nociceptive effects in comparison to the respective blank formulations.

Development of nanoparticle loaded microneedles for drug delivery to a brain tumour resection site.

Systemic drug delivery to the central nervous system (CNS) has been historically impeded by the presence of the blood brain barrier rendering many therapies inefficacious to any cancer cells residing within the brain. Therefore, local drug delivery systems are being developed to overcome this shortfall. Here we have manufactured polymeric microneedle (MN) patches, which can be anchored within a resection cavity site following surgical removal of a tumour such as isocitrate dehydrogenase wild type glioblastoma (GBM). These MN patches have been loaded with polymer coated nanoparticles (NPs) containing cannabidiol (CBD) or olaparib (OLA) and applied to an in vitro brain simulant and ex vivo rat brain tissue to assess drug release and distance of penetration. MN patches loaded with methylene blue dye were placed into a cavity of 0.6 % agarose to simulate brain tissue. The results showed that clear channels were generated by the MNs and the dye spread laterally throughout the agarose. When loaded with CBD-NPs, the agarose showed a CBD concentration of 12.5 µg/g at 0.5 cm from the MN insertion site. Furthermore, high performance liquid chromatography of ex vivo brain tissue following CBD-NP/MN patch insertion showed successful delivery of 59.6 µg/g into the brain tissue. Similarly, OLA-NP loaded MN patches showed delivery of 5.2 µg/g OLA into agarose gel at 0.5 cm distance from the insertion site. Orbitrap secondary ion mass spectrometry (OrbiSIMS) analysis confirmed the presence of OLA and the MN patch at up to 6 mm away from the insertion site following its application to a rat brain hemisphere. This data has provided insight into the capabilities and versatility of MN patches for use in local brain drug delivery, giving promise for future research.

Electric field responsive nanotransducers for glioblastoma.

BACKGROUND: Electric field therapies such as Tumor Treating Fields (TTFields) have emerged as a bioelectronic treatment for isocitrate dehydrogenase wild-type and IDH mutant grade 4 astrocytoma Glioblastoma (GBM). TTFields rely on alternating current (AC) electric fields (EF) leading to the disruption of dipole alignment and induced dielectrophoresis (DEP) during cytokinesis. Although TTFields have a favourable side effect profile, particularly compared to cytotoxic chemotherapy, survival benefits remain limited (~ 4.9 months) after an extensive treatment regime (20 hours/day for 18 months). The cost of the technology also limits its clinical adoption worldwide. Therefore, the discovery of new technology that can enhance both the therapeutic efficiency and efficacy of these TTFields will be of great benefit to cancer treatment and decrease healthcare costs worldwide. METHODS: In this work, we report the role of electrically conductive gold (GNPs), dielectric silica oxide (SiO2), and semiconductor zinc oxide (ZnO) nanoparticles (NPs) as transducers for enhancing EF mediated anticancer effects on patient derived GBM cells. Physicochemical properties of these NPs were analyzed using spectroscopic, electron microscopy, and light-scattering techniques. RESULTS: In vitro TTFields studies indicated an enhanced reduction in the metabolic activity of patient-derived Glioma INvasive marginal (GIN 28) and Glioma contrast enhanced core (GCE 28) GBM As per our journal style, article titles should not include capitalised letters unless these are proper nouns/acronyms. We have therefore used the article title "Electric field responsive nanotransducers for glioblastoma" as opposed to "Electric Field Responsive Nanotransducers for Glioblastoma" as given in the submission system. Please check if this is correct.cells in groups treated with NPs vs. control groups, irrespective of NPs dielectric properties. Our results indicate the inorganic NPs used in this work enhance the intracellular EF effects that could be due to the virtue of bipolar dielectrophoretic and electrophoretic effects. CONCLUSIONS: This work presents preliminary evidence which could help to improve future EF applications for bioelectronic medicine. Furthermore, the merits of spherical morphology, excellent colloidal stability, and low toxicity, make these NPs ideal for future studies for elucidating the detailed mechanism and efficacy upon their delivery in GBM preclinical models.

Lipoprotein Deprivation Reveals a Cholesterol-Dependent Therapeutic Vulnerability in Diffuse Glioma Metabolism.

Poor outcomes associated with diffuse high-grade gliomas occur in both adults and children, despite substantial progress made in the molecular characterisation of the disease. Targeting the metabolic requirements of cancer cells represents an alternative therapeutic strategy to overcome the redundancy associated with cell signalling. Cholesterol is an integral component of cell membranes and is required by cancer cells to maintain growth and may also drive transformation. Here, we show that removal of exogenous cholesterol in the form of lipoproteins from culture medium was detrimental to the growth of two paediatric diffuse glioma cell lines, KNS42 and SF188, in association with S-phase elongation and a transcriptomic program, indicating dysregulated cholesterol homeostasis. Interrogation of metabolic perturbations under lipoprotein-deficient conditions revealed a reduced abundance of taurine-related metabolites and cholesterol ester species. Pharmacological reduction in intracellular cholesterol via decreased uptake and increased export was simulated using the liver X receptor agonist LXR-623, which reduced cellular viability in both adult and paediatric models of diffuse glioma, although the mechanism appeared to be cholesterol-independent in the latter. These results provide proof-of-principle for further assessment of liver X receptor agonists in paediatric diffuse glioma to complement the currently approved therapeutic regimens and expand the options available to clinicians to treat this highly debilitating disease.

Metabolic modeling-based drug repurposing in Glioblastoma.

The manifestation of intra- and inter-tumor heterogeneity hinders the development of ubiquitous cancer treatments, thus requiring a tailored therapy for each cancer type. Specifically, the reprogramming of cellular metabolism has been identified as a source of potential drug targets. Drug discovery is a long and resource-demanding process aiming at identifying and testing compounds early in the drug development pipeline. While drug repurposing efforts (i.e., inspecting readily available approved drugs) can be supported by a mechanistic rationale, strategies to further reduce and prioritize the list of potential candidates are still needed to facilitate feasible studies. Although a variety of 'omics' data are widely gathered, a standard integration method with modeling approaches is lacking. For instance, flux balance analysis is a metabolic modeling technique that mainly relies on the stoichiometry of the metabolic network. However, exploring the network's topology typically neglects biologically relevant information. Here we introduce Transcriptomics-Informed Stoichiometric Modelling And Network analysis (TISMAN) in a recombinant innovation manner, allowing identification and validation of genes as targets for drug repurposing using glioblastoma as an exemplar.

Transcription Profile and Pathway Analysis of the Endocannabinoid Receptor Inverse Agonist AM630 in the Core and Infiltrative Boundary of Human Glioblastoma Cells.

BACKGROUND: We have previously reported that the endocannabinoid receptor inverse agonist AM630 is a potent inhibitor of isocitrade dehydrogenase-1 wild-type glioblastoma (GBM) core tumour cell proliferation. To uncover the mechanism behind the anti-tumour effects we have performed a transcriptional analysis of AM630 activity both in the tumour core cells (U87) and the invasive margin cells (GIN-8), the latter representing a better proxy of post-surgical residual disease. RESULTS: The core and invasive margin cells exhibited markedly different gene expression profiles and only the core cells had high expression of a potential AM630 target, the CB1 receptor. Both cell types had moderate expression of the HTR2B serotonin receptor, a reported AM630 target. We found that the AM630 driven transcriptional response was substantially higher in the central cells than in the invasive margin cells, with the former driving the up regulation of immune response and the down regulation of cell cycle and metastatic pathways and correlating with transcriptional responses driven by established anti-neoplastics as well as serotonin receptor antagonists. CONCLUSION: Our results highlight the different gene sets involved in the core and invasive margin cell lines derived from GBM and an associated marked difference in responsiveness to AM630. Our findings identify AM630 as an anti-neoplastic drug in the context of the core cells, showing a high correlation with the activity of known antiproliferative drugs. However, we reveal a key set of similarities between the two cell lines that may inform therapeutic intervention.

Detection of Label-Free Drugs within Brain Tissue Using Orbitrap Secondary Ion Mass Spectrometry as a Complement to Neuro-Oncological Drug Delivery.

Historically, pre-clinical neuro-oncological drug delivery studies have exhaustively relied upon overall animal survival as an exclusive measure of efficacy. However, with no adopted methodology to both image and quantitate brain parenchyma penetration of label-free drugs, an absence of efficacy typically hampers clinical translational potential, rather than encourage re-formulation of drug compounds using nanocarriers to achieve greater tissue penetration. OrbiSIMS, a next-generation analytical instrument for label-free imaging, combines the high resolving power of an OrbiTrapTM mass spectrometer with the relatively high spatial resolution of secondary ion mass spectrometry. Here, we develop an ex vivo pipeline using OrbiSIMS to accurately detect brain penetration of drug compounds. Secondary ion spectra were acquired for a panel of drugs (etoposide, olaparib, gemcitabine, vorinostat and dasatinib) under preclinical consideration for the treatment of isocitrate dehydrogenase-1 wild-type glioblastoma. Each drug demonstrated diagnostic secondary ions (all present molecular ions [M-H]− which could be discriminated from brain analytes when spiked at >20 µg/mg tissue. Olaparib/dasatinib and olaparib/etoposide dual combinations are shown as exemplars for the capability of OrbiSIMS to discriminate distinct drug ions simultaneously. Furthermore, we demonstrate the imaging capability of OrbiSIMS to simultaneously illustrate label-free drug location and brain chemistry. Our work encourages the neuro-oncology community to consider mass spectrometry imaging modalities to complement in vivo efficacy studies, as an analytical tool to assess brain distribution of systemically administered drugs, or localised brain penetration of drugs released from micro- or nano-scale biomaterials.

Genome-Wide Expression and Anti-Proliferative Effects of Electric Field Therapy on Pediatric and Adult Brain Tumors.

The lack of treatment options for high-grade brain tumors has led to searches for alternative therapeutic modalities. Electrical field therapy is one such area. The Optune™ system is an FDA-approved novel device that delivers continuous alternating electric fields (tumor treating fields-TTFields) to the patient for the treatment of primary and recurrent Glioblastoma multiforme (GBM). Various mechanisms have been proposed to explain the effects of TTFields and other electrical therapies. Here, we present the first study of genome-wide expression of electrotherapy (delivered via TTFields or Deep Brain Stimulation (DBS)) on brain tumor cell lines. The effects of electric fields were assessed through gene expression arrays and combinational effects with chemotherapies. We observed that both DBS and TTFields significantly affected brain tumor cell line viability, with DBS promoting G0-phase accumulation and TTFields promoting G2-phase accumulation. Both treatments may be used to augment the efficacy of chemotherapy in vitro. Genome-wide expression assessment demonstrated significant overlap between the different electrical treatments, suggesting novel interactions with mitochondrial functioning and promoting endoplasmic reticulum stress. We demonstrate the in vitro efficacy of electric fields against adult and pediatric high-grade brain tumors and elucidate potential mechanisms of action for future study.

Investigating histidinylated highly branched poly(lysine) for siRNA delivery.

The temporary silencing of disease-associated genes utilising short interfering RNA (siRNA) is a potent and selective route for addressing a wide range of life limiting disorders. However, the few clinically approved siRNA therapies rely on lipid based formulations, which although potent, provide limited chemical space to tune the stability, efficacy and tissue selectivity. In this study, we investigated the role of molar mass and histidinylation for poly(lysine) based non-viral vectors, synthesised through a fully aqueous thermal condensation polymerisation. Formulation and in vitro studies revealed that higher molar mass derivatives yielded smaller polyplexes attributed to a greater affinity for siRNA at lower N/P ratios yielding greater transfection efficiency, albeit with some cytotoxicity. Histidinylation had a negligible effect on formulation size, yet imparted a moderate improvement in biocompatibility, but did not provide any meaningful improvement over silencing efficiency compared to non-histidinylated derivatives. This was attributed to a greater degree of cellular internalisation for non-histidinylated analogues, which was enhanced with the higher molar mass material.