Permeable Blood-brain Barrier Research Articles

Feasibility of intranasal human milk as stem cell therapy in preterm infants with intraventricular hemorrhage.

Intraventricular hemorrhage (IVH) is a common cause of preterm brain injury. Fresh parent's own milk (POM) contains pluripotent stem cells (SCs) that produce neuronal cells in-vitro. The permeable neonatal blood brain barrier potentially allows SC delivery. We performed the first prospective trial (clinicaltrials.gov NCT04225286) of feasibility of intranasal POM (IPOM) in preterm infants with IVH and described SC content of POM samples. 37 Infants (mean gestation 27.7 ± 2.6 weeks, birthweight 1030 ± 320 g) with IVH (35.1% grade IV) were recruited from two tertiary Toronto NICUs. IPOM was given ideally twice daily until 28 days of age. Tolerance and adverse reactions were collected and 162 administering providers surveyed. There were no major adverse reactions. Provider surveys suggested acceptability, although potential provider and subject stress requires further study. Milk cell analysis suggests wide variability between parents. This phase 1 study demonstrated IPOM was tolerated and feasible in preterm infants.

Unraveling the link between neuropathy target esterase NTE/SWS, lysosomal storage diseases, inflammation, abnormal fatty acid metabolism, and leaky brain barrier.

Mutations in Drosophila Swiss cheese (SWS) gene or its vertebrate orthologue neuropathy target esterase (NTE) lead to progressive neuronal degeneration in flies and humans. Despite its enzymatic function as a phospholipase is well established, the molecular mechanism responsible for maintaining nervous system integrity remains unclear. In this study, we found that NTE/SWS is present in surface glia that forms the blood-brain barrier (BBB) and that NTE/SWS is important to maintain its structure and permeability. Importantly, BBB glia-specific expression of Drosophila NTE/SWS or human NTE in the sws mutant background fully rescues surface glial organization and partially restores BBB integrity, suggesting a conserved function of NTE/SWS. Interestingly, sws mutant glia showed abnormal organization of plasma membrane domains and tight junction rafts accompanied by the accumulation of lipid droplets, lysosomes, and multilamellar bodies. Since the observed cellular phenotypes closely resemble the characteristics described in a group of metabolic disorders known as lysosomal storage diseases (LSDs), our data established a novel connection between NTE/SWS and these conditions. We found that mutants with defective BBB exhibit elevated levels of fatty acids, which are precursors of eicosanoids and are involved in the inflammatory response. Also, as a consequence of a permeable BBB, several innate immunity factors are upregulated in an age-dependent manner, while BBB glia-specific expression of NTE/SWS normalizes inflammatory response. Treatment with anti-inflammatory agents prevents the abnormal architecture of the BBB, suggesting that inflammation contributes to the maintenance of a healthy brain barrier. Considering the link between a malfunctioning BBB and various neurodegenerative diseases, gaining a deeper understanding of the molecular mechanisms causing inflammation due to a defective BBB could help to promote the use of anti-inflammatory therapies for age-related neurodegeneration.

A Chronic Increase in Blood-Brain Barrier Permeability Facilitates Intraneuronal Deposition of Exogenous Bloodborne Amyloid-Beta1-42 Peptide in the Brain and Leads to Alzheimer's Disease-Relevant Cognitive Changes in a Mouse Model.

Increased blood-brain barrier (BBB) permeability and amyloid-β (Aβ) peptides (especially Aβ1-42) (Aβ42) have been linked to Alzheimer's disease (AD) pathogenesis, but the nature of their involvement in AD-related neuropathological changes leading to cognitive changes remains poorly understood. To test the hypothesis that chronic extravasation of bloodborne Aβ42 peptide and brain-reactive autoantibodies and their entry into the brain parenchyma via a permeable BBB contribute to AD-related pathological changes and cognitive changes in a mouse model. The BBB was rendered chronically permeable through repeated injections of Pertussis toxin (PT), and soluble monomeric, fluorescein isothiocyanate (FITC)-labeled or unlabeled Aβ42 was injected into the tail-vein of 10-month-old male CD1 mice at designated intervals spanning ∼3 months. Acquisition of learned behaviors and long-term retention were assessed via a battery of cognitive and behavioral tests and linked to neuropathological changes. Mice injected with both PT and Aβ42 demonstrated a preferential deficit in the capacity for long-term retention and an increased susceptibility to interference in selective attention compared to mice exposed to PT or saline only. Immunohistochemical analyses revealed increased BBB permeability and entry of bloodborne Aβ42 and immunoglobulin G (IgG) into the brain parenchyma, selective neuronal binding of IgG and neuronal accumulation of Aβ42 in animals injected with both PT and Aβ42 compared to controls. Results highlight the potential synergistic role of BBB compromise and the influx of bloodborne Aβ42 into the brain in both the initiation and progression of neuropathologic and cognitive changes associated with AD.

T Cells in Tick-Borne Flavivirus Encephalitis: A Review of Current Paradigms in Protection and Disease Pathology.

The family Flaviviridae is comprised of a diverse group of arthropod-borne viruses that are the etiological agents of globally relevant diseases in humans. Among these, infection with several of these flaviviruses-including West Nile virus (WNV), Zika virus (ZIKV), Japanese encephalitis virus (JEV), tick-borne encephalitis virus (TBEV), and Powassan virus (POWV)-can result in neuroinvasive disease presenting as meningitis or encephalitis. Factors contributing to the development and resolution of tick-borne flavivirus (TBEV, POWV) infection and neuropathology remain unclear, though many recently undertaken studies have described the virus-host interactions underlying encephalitic disease. With access to neural tissues despite the selectively permeable blood-brain barrier, T cells have emerged as one notable contributor to neuroinflammation. The goal of this review is to summarize the recent advances in tick-borne flavivirus immunology-particularly with respect to T cells-as it pertains to the development of encephalitis. We found that although T cell responses are rarely evaluated in a clinical setting, they are integral in conjunction with antibody responses to restricting the entry of TBFV into the CNS. The extent and means by which they can drive immune pathology, however, merits further study. Understanding the role of the T cell compartment in tick-borne flavivirus encephalitis is instrumental for improving vaccine safety and efficacy, and has implications for treatments and interventions for human disease.

Abstract 2501: A glioblastoma and blood-brain barrier in vitro model to assess the tumor microenvironment and temozolomide resistance

Abstract Glioblastoma multiforme (GBM) is the most common primary brain tumor in adults with a dismal prognosis. Currently, Temozolomide (TMZ) and Bevacizumab are the main approved agents for primary and recurrent GBM, respectively. However, TMZ resistance is frequent and Bevacizumab does not improve overall survival. The heterogeneity of GBM and the presence of a highly selectively permeable blood-brain barrier (BBB) preclude the use of chemotherapeutics approved for other malignancies. Meanwhile, new therapy development is limited in part because most pre-clinical models do not recapitulate the key features of GBM that are relevant to therapy resistance and drug delivery. This is particularly important when validating immunotherapies such as engineered T cell therapy, which has shown some promise in the clinics. We therefore developed an in vitro 3D model that recapitulates the major features of the GBM tumor microenvironment (TME) that are involved in therapy resistance, including the dense tumor core and infiltrating invasive front observed in clinical tumors, surrounded by a perfusable BBB with a physiological permeability. With this model, we investigate the angiogenic nature of GBM, and the role of tumor organization and the TME in TMZ resistance and immunosuppression. We find that TMZ sensitivity decreases as complexity of tumor cell 3D organization increases, and identify pathways potentially associated with TMZ resistance and immunosuppression using proteomics and genomics approaches. Additionally, we find that tumor-associated vasculature has a higher permeability due to VEGFA from GBM tumors, which leads to the downregulation of endothelial junction proteins. Using an IL13Ra2-targetting chimeric antigen receptor (CAR) T cell as a model for engineered T cell therapy, we find that tumor-associated vasculature acts as a physical barrier to CAR-T cell extravasation, significantly impeding CAR-T cell efficacy. We then use the model to screen a CAR library to identify novel CAR combinations that can improve CAR-T cell extravasation and cytotoxicity in GBM. Citation Format: Maxine S. Lam, Joey Aw, Damien Tan, Ragavi Vijayakumar, Khloe S. Gordon, Michael E. Birnbaum, Radoslaw M. Sobota, Giulia Adriani, Andrea Pavesi. A glioblastoma and blood-brain barrier in vitro model to assess the tumor microenvironment and temozolomide resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2501.

Mechanistic insights into immunopathogenesis of murine cerebral malaria: Cues from “young” C57BL/6J and BALB/c mice

Cerebral malaria (CM), a major cause of mortality in children <5 years, presents disparity in pathophysiological features and poor prognosis compared to adults. Adult C57BL/6J mice infected with Plasmodium berghei ANKA (PbA) are widely used to understand CM pathogenesis compared to relatively less prone BALB/c mice; however, age and immune status of the host also influence disease sequelae and cerebral manifestations. Murine models of CM known so far do not project complete disease spectrum of pediatric CM. The present study was designed to dissect and differentiate CM immunopathogenesis in “young” BALB/c and C57BL/6J mice infected with PbA, in search of a competent mouse model mimicking pediatric CM. Multipronged approach including the analysis of blood-brain barrier (BBB) permeability and parasite infiltration, histopathology, nitric oxide levels, and pro/anti-inflammatory (TNF-α, IFN-γ, IL-4, and IL-10) cytokine expression were compared in the cortices of both young BALB/c and C57BL/6J mice. The results illustrate severe course of infection and typical CM like histopathological alterations including monocytic plugging in PbA-infected “young" BALB/c compared to C57BL/6J mice. The decreased expression of tight junction proteins (ZO-1 and Claudin-3) and Evan's blue extravasation was also more evident in BALB/c mice indicating a more permeable BBB. The increased cortical expression of TNF-α, IFN-γ, IL-4, IL-10, iNOS, eNOS, nNOS, and associated activation of brain resident cells in cortices of BALB/c with progressive parasitaemia depicts the cumulative involvement of host immune responses and parasite accumulation in progression of CM. Thus, the incongruity of cytokine balance resulted in worsening of disease manifestation in “young” BALB/c similar to pediatric CM.

Design, synthesis, and structure-activity relationship of caffeine-based triazoles as dual AChE and BACE-1 inhibitors.

Natural products have significantly contributed to drug discovery for neurodegenerative diseases. Caffeine is one of the well-known central nervous system(CNS)-active natural products. Besides its CNS stimulant properties, it is a mild inhibitor of acetylcholinesterase (AChE) and possesses memory-enhancing properties. The present work aimed to improve the AChE inhibition activity of the caffeine. The rationally designed caffeine-based triazoles were synthesized and evaluated in vitro for cholinesterase and β-site amyloid precursor protein cleaving enzyme-1 (BACE-1) inhibitory activities. The attachment of triazole to the caffeine enhances its AChE inhibition activity from half-maximal inhibitory concentration (IC50 ) of 129 µMto0.49 µM (derivative, 6l). The caffeine core interacts with the peripheral anionic site, whereas the benzyl triazole occupies the catalytic anionic site located at the bottom of the active site gorge. The structure-activity relationship revealed that the four-atom ester linker is superior to shorter linkers for connecting the caffeine core to the triazole. The 2,6-difluorobenzyl triazole-linked caffeine derivative, 6d, exhibits dual inhibition of AChE and BACE-1 with IC50 values of 1.43 and 10.9 µM, respectively. The derivative 6d inhibits AChE via a mixed-type mode with an inhibition rate constant(Ki ) value of 2.35 μM, which was corroborated by docking studies. The triazole 6d has an acceptable stability profile in human liver microsomes (t1/2 = 54 min) and was found to possessCNS permeabilitywhen evaluated using the parallel artificial membrane permeability blood-brain barrier assay. The results presented herein warrant investigating caffeine-based triazoles in preclinical models of Alzheimer's disease.

DIPG-21. DIPG cells alter the permeability of the blood-brain barrier in the brainstem leading to treatment failure.

Diffuse Intrinsic Pontine Glioma (DIPG) is an aggressive paediatric high-grade glioma with no effective therapies. The blood-brain barrier (BBB) presents a significant obstacle for delivery of therapeutics into the brain, especially in the brainstem. This study aims to investigate the effect of DIPG cells on the BBB in the brainstem. We hypothesized that the location of DIPG may result in a less permeable BBB than other brain regions. We compared two independent orthotopic models with the same DIPG cells injected in the cortical region or brainstem. We found that treatment with mTOR inhibitor, temsirolimus, significantly extended survival for cortical tumors compared with the same tumor in the brainstem (p=0.0097). Immunohistochemical analysis showed significant reduction of mTOR target, P-p70S6K, in the cortical region compared to brainstem in treated animals, with pharmacokinetic analysis confirming significantly higher temsirolimus levels in the cortical region. These findings suggest that cortical tumors respond better than brainstem tumors, and that a less permeable BBB in brainstem tumors contributes to treatment failure. To understand whether DIPG cells affect the BBB in the brainstem, single-cell RNA sequencing experiments were conducted on vasculature isolated from DIPG and matrigel-injected mice. Gene-ontology overrepresentation analyses identified downregulation in the P38MAPK pathway in endothelial cells from DIPG-injected mice, suggesting the potential for therapeutic manipulation with cytokines. Treatment with SNGR-TNFα, a derivative of an agent successfully used in improving drug penetration in CNS lymphoma patients, in an in vitro BBB/DIPG model significantly reduced transendothelial resistance, and further exploration into the effects on the BBB in vivo is currently being undertaken. Our studies indicate that the intact BBB in the brainstem in DIPG is a major reason for treatment failure, and DIPG cells directly influence the vasculature and response to treatment. This may lead to a novel DIPG treatment strategy and for other brain tumors.

Lithium Induced Parkinsonism: A Case Report

AimsLithium is an effective mood stabiliser for the treatment of the bipolar disorder. Its utility is not restricted to acute mania and prophylactic treatment of the bipolar disorder. Another well-known indication for its use is the treatment of refractory depression. Lithium can cause several adverse effects, and typically the side effects are dose-related. Unlike antipsychotic medications, lithium is rarely associated with drug-induced Parkinsonism.MethodsWe present a case of 78 years old gentleman who was assessed due to complaints suggestive of cognitive impairment. His past psychiatric history revealed that he was admitted to a psychiatric inpatient unit with a diagnosis of treatment-resistant depression in 1991. Lithium therapy was commenced during this admission, and he remained on lithium for 27 years. The patient was clinically stable in terms of the symptoms of depression; however, he reported bilateral postural tremors 20 years after the initiation of lithium therapy. Initially, he was diagnosed with lithium-induced tremor; however, in the following months, his symptoms had worsened, and he developed new motor disturbances, although the serum levels of lithium were within the therapeutic range. On examination, he had classic parkinsonian signs of shuffling gait, muscle rigidity in all four limbs and freezing of gait. DaT-SPECT imaging clarified the diagnosis as drug-induced Parkinsonism. As the daily lithium dosage was stopped, the patient's motor symptoms improved significantly; nevertheless, some of the symptoms persisted.ResultsThe pathophysiological mechanism behind lithium-induced Parkinsonism is unclear. The condition may develop with or without frank lithium toxicity and have diverse presentations. Literature suggests that the risk factors for lithium-induced Parkinsonism appear to be the patients' age, duration of lithium therapy, and serum lithium levels. It has been suggested that older patients have a more permeable blood-brain barrier and decreased renal clearance; hence, serum lithium levels can appear therapeutic, but brain lithium levels may be much higher. Pharmacokinetic drug-drug interactions might also contribute; thus, careful monitoring is essential.Drug-induced Parkinsonism improves with discontinuation of the offending medication; however, 10% of patients will develop a persistent and progressive parkinsonian syndrome.ConclusionThis report aims to emphasise the need to consider lithium-induced Parkinsonism when Parkinson Disease symptoms appear in chronic lithium users and close monitoring of lithium levels in geriatric populations. It is essential to recognise the condition, avoid misdiagnosis and prevent inappropriate use of anti-dopaminergic medications.

Current Strategies to Enhance Delivery of Drugs across the Blood-Brain Barrier.

The blood–brain barrier (BBB) has shown to be a significant obstacle to brain medication delivery. The BBB in a healthy brain is a diffusion barrier that prevents most substances from passing from the blood to the brain; only tiny molecules can pass across the BBB. The BBB is disturbed in specific pathological illnesses such as stroke, diabetes, seizures, multiple sclerosis, Parkinson’s disease, and Alzheimer’s disease. The goal of this study is to offer a general overview of current brain medication delivery techniques and associated topics from the last five years. It is anticipated that this review will stimulate readers to look into new ways to deliver medications to the brain. Following an introduction of the construction and function of the BBB in both healthy and pathological conditions, this review revisits certain contested questions, such as whether nanoparticles may cross the BBB on their own and if medications are selectively delivered to the brain by deliberately targeted nanoparticles. Current non-nanoparticle options are also discussed, including drug delivery via the permeable BBB under pathological circumstances and the use of non-invasive approaches to improve brain medication absorption.

Minocycline decreases blood-brain barrier permeability following aneurysmal subarachnoid hemorrhage: a randomized, double-blind, controlled trial.

Aneurysmal subarachnoid hemorrhage (aSAH)-induced vasospasm is linked to increased inflammatory cell trafficking across a permeable blood-brain barrier (BBB). Elevations in serum levels of matrix metalloprotease 9 (MMP9), a BBB structural protein, have been implicated in the pathogenesis of vasospasm onset. Minocycline is a potent inhibitor of MMP9. The authors sought to detect an effect of minocycline on BBB permeability following aSAH. Patients presenting within 24 hours of symptom onset with imaging confirmed aSAH (Fisher grade 3 or 4) were randomized to high-dose (10 mg/kg) minocycline or placebo. The primary outcome of interest was BBB permeability as quantitated by contrast signal intensity ratios in vascular regions of interest on postbleed day (PBD) 5 magnetic resonance permeability imaging. Secondary outcomes included serum MMP9 levels and radiographic and clinical evidence of vasospasm. A total of 11 patients were randomized to minocycline (n = 6) or control (n = 5) groups. No adverse events or complications attributable to minocycline were reported. High-dose minocycline administration was associated with significantly lower permeability indices on imaging analysis (p < 0.01). There was no significant difference with respect to serum MMP9 levels between groups, although concentrations trended upward in both cohorts. Radiographic vasospasm was noted in 6 patients (minocycline = 3, control = 3), with only 1 patient developing symptoms of clinical vasospasm in the minocycline cohort. There was no difference between cohorts with respect to Lindegaard ratios, transcranial Doppler values, or onset of vasospasm. Minocycline at high doses is well tolerated in the ruptured cerebral aneurysm population. Minocycline curtails breakdown of the BBB following aSAH as evidenced by lower permeability indices, though minocycline did not significantly alter serum MMP9 levels. Larger randomized clinical trials are needed to assess minocycline as a neuroprotectant against aSAH-induced vasospasm. Clinical trial registration no.: NCT04876638 (clinicaltrials.gov).

495 5-ALA PDT and Targeting MEK/ERK Signaling Elicits Synergistic Antitumor Effects in Diffuse Midline Glioma

INTRODUCTION: Diffuse midline gliomas are highly invasive, unresectable tumors in children. To date, there is no effective treatment for DMG. Concurrent radiotherapy (RT) and systemic therapies, the standard of care, has only been successful in providing limited disease control. The major obstacle to therapy is the selectively permeable blood brain barrier (BBB) that limits systemic drug delivery. Given the unmet need for penetrant and minimally invasive DMG treatments, photodynamic therapy (PDT), with the precursor photosensitizing agent 5-aminolevulinic acid (5-ALA), is an oncologic treatment that holds promise. 5-ALA PDT of tumors occurs by targeting tumor cells that accumulate the 5-ALA metabolite, protoporphyrin IX (PPIX), with 635 nm light to create deadly reactive oxygen species (ROS). METHODS: Growth delay assays were conducted for each individual treatment modality or in combination. FDA-approved MEK/ERK inhibitors were administered to mice with aggressive heterotopic allograft DMGs through intraperitoneal injection. Ex vivo immunofluorescence staining was performed on those mice that received 5-ALA PDT treatment. RESULTS: We demonstrated that sub-micromolar levels of 5-ALA PDT and nanomolar levels of MEK inhibitor successfully decrease cell proliferation and induce apoptosis in DMG cell lines. Cell viability assays revealed that drug response differs based on the histone mutation (H3.1 or H3.3) of the line. Mechanisms of decreased cell survival involves the generation of reactive oxygen species that induces programmed cell death. Through the use of a DMG genetically engineered mouse model, we also found 5-ALA PDT to induce apoptosis in vivo. CONCLUSION: The synergistic effects of MEK inhibition and 5-ALA PDT in vitro and apoptotic effects of 5-ALA PDT in vivo, highlights the potential therapeutic efficacy of this treatment modality.

ITVT-01. 5-ALA PDT and Targeting MEK/ERK Signaling Elicits Synergistic Antitumor Effects in Diffuse Midline Glioma

Abstract Diffuse midline gliomas are highly invasive, unresectable tumors in children. To date, there is no effective treatment for DMG. Concurrent radiotherapy (RT) and systemic therapies, the standard of care, has only been successful in providing limited disease control. The major obstacle to therapy is the selectively permeable blood brain barrier (BBB) that limits systemic drug delivery. Given the unmet need for penetrant and minimally invasive DMG treatments, photodynamic therapy (PDT), with the precursor photosensitizing agent 5-aminolevulinic acid (5-ALA), is an oncologic treatment that holds promise. 5-ALA PDT of tumors occurs by targeting tumor cells that accumulate the 5-ALA metabolite, protoporphyrin IX (PPIX), with 635 nm light to create deadly reactive oxygen species (ROS). We explore the synergism of 5-ALA PDT with the MEK inhibitor, trametinib, since the RAS/MEK signaling pathway regulates tumor cell proliferation and survival and has been shown to therapeutically enhance PDT in select carcinoma models. We demonstrated that sub-micromolar levels of 5-ALA PDT and nanomolar levels of trametinib successfully decrease cell proliferation and induce apoptosis in DMG cell lines. Cell viability assays revealed that drug response differs based on the histone mutation (H3.1 or H3.3) of the line. Mechanisms of decreased cell survival involves the generation of reactive oxygen species that induces programmed cell death. Through the use of a DMG genetically engineered mouse model, we also found 5-ALA PDT to induce apoptosis in vivo. The synergistic effects of MEK inhibition and 5-ALA PDT in vitro and apoptotic effects of 5-ALA PDT in vivo, highlights the potential therapeutic efficacy of this treatment modality.

Calcineurin Inhibitor Use and Myoclonus Association. Is There a Clinical Implication?

Background: Calcineurin Inhibitors (CNIs) play a pivotal role in anti rejection therapy for transplant patients. Neurotoxicity is a known side effect that usually manifests as encephalopathy but myoclonus has also been described. Perioperative myoclonus as a manifestation of neurotoxicity, has not been well studied. Methods: We retrospectively reviewed data from 842,762 patients from the Nationwide Inpatient Sample (NIS) database from January 2011 to December 2014. Of those records we compared 56,423 patients requiring CNIs and undergoing Heart Transplant (HT) with 786,339 patients undergoing Coronary Artery Bypass Graft (CABG) surgery as controls. The objective was to study the rates of myoclonus in patients undergoing cardiac surgery, especially those requiring CNIs, and study the outcome of those patients with myoclonus. The NIS database from January 2011 to December 2014 was the source for the analysis. Patients with underlying epilepsy or hypo-ischemic encephalopathy based on ICD-9-CM codes were excluded from the study. Results: A total of 147 patients (0.26%) were found to have myoclonus in the HT group versus 338 patients (0.04%) in the CABG group, p Conclusion: In this study we have compared the rate of myoclonus found in HT patients versus CABG patients. We have identified calcineurin inhibitors as potentially contributing to myoclonus due to its neurotoxic effects. The study also suggests that other disease processes like renal failure may also have an impact on the rate of myoclonus even in the absence of calcineurin inhibitors. Higher rates of myoclonus were seen in patients undergoing HT when compared to patients undergoing CABG, suggesting that CNIs may increase the risk for myoclonus. Myoclonus may be a clinical indicator of patient overall health including a more permeable blood brain barrier. In-Hospital mortality was higher in patients with myoclonus undergoing CABG and a trend towards significance in the HT group suggesting that it may be a marker of poor prognosis. More studies are needed to corroborate our findings.

Bypassing the Blood-Brain Barrier to Treat Brain Cancer: A Systematic Review of the Efficacy of Carmustine Wafer Implant Therapy

Introduction: Gliomas are neoplasms of the central nervous system (CNS). Despite aggressive treatment, median survival of malignant tumors remains poor at 12 - 18 months. Newer treatments allow delivery of therapeutic substances across the selectively permeable blood-brain barrier (BBB). This allows for chemotherapeutics to more easily reach their target location in the CNS. Drug eluting wafers made up of carmustine can be placed in the surgical resection cavity of a tumor and clinical trials to date have demonstrated their utility. Hypothesis: Bypassing the BBB to allow greater accumulation of chemotherapeutics in the CNS will improve clinical outcomes in glioma patients. Methods: Studies from medical literature databases describing trials using carmustine wafers implanted after glioma resection were obtained. To test our hypothesis, the available data using this therapy was compared to current first line treatment data for glioma as described by Stupp and colleagues. The inclusion criterion for efficacy analysis was histopathologically confirmed primary glioma. Exclusion criteria included presence of metastasis or pediatric tumors. Results: 10 studies describing wafer therapy use were initially gathered, encompassing over 500 patients. 6 studies met criteria for treatment efficacy analysis. 4 of 6 (75%) trials exhibited significant survival advantage as compared to control treatment. Furthermore, 3 of the 4 (75%) studies showing significance also demonstrated equal or higher percent increase in overall survival from control as compared to data generated from current first line therapy. Conclusion: Treatments bypassing the BBB are not currently standard-of-care for patients with glioma. We uncovered that most trials using carmustine implants post tumor resection describe increased overall survival, however in specific cohorts. Diverting the BBB in general may also have fewer side effects in contrast to classical routes of therapy. Future work is needed to develop similar therapeutics that improve outcomes in all age, gender, and prognostic risk factor populations.

Predictive medicinal metabolites from Momordica dioica against comorbidity related proteins of SARS-CoV-2 infections

Momordica dioica have proven medicinal potential of antidiabetic, antiviral and immune stimulating properties. Flavonoids and triterpenoids from M. dioica were more extensively investigated for antiviral, antidiabetic and immunomodulatory activities. In this present study, we have predicted the reported bioactive flavonoids and triterpenoids of the plant against the SARS-CoV-2 main protease, RNA-dependent RNA polymerase (RdRp), spike protein, angiotensin converting enzyme (ACE-2) receptor and dipeptidyl peptidase (DPP4) receptor through molecular docking and in silico ADME predictions methods. According to the binding affinities, the two triterpenoids, hederagenin and oleanolic acid exhibited the best docking scores with these proteins than the catechin and quercetin with compared to standard remdesivir, favipiravir and hydroxychloroquine. The in vitro protein-drug studies have also showed significant interaction of catechin and quercetin compounds than standard drugs. The in silico binding studies correlated with the in silico binding studies. Further, M. dioica being used as antidiabetic and its metabolite had significant interaction with DDP4, a comorbidity protein involved in aiding the viral entry. Out of all the natural ligands, quercetin was reported relatively good and safe for humans with high gastrointestinal tract permeability and poor blood brain barrier crossing abilities. Hence, M. dioica phytocompounds reflects promising therapeutic properties against SARS-CoV-2 infections under comorbid conditions such as diabetes, cardiovascular disease and kidney disorders. Communicated by Ramaswamy H. Sarma

Quantitative neurotoxicology: Potential role of artificial intelligence/deep learning approach.

Neurotoxicity studies are important in the preclinical stages of drug development process, because exposure to certain compounds that may enter the brain across a permeable blood brain barrier damages neurons and other supporting cells such as astrocytes. This could, in turn, lead to various neurological disorders such as Parkinson's or Huntington's disease as well as various dementias. Toxicity assessment is often done by pathologists after these exposures by qualitatively or semiquantitatively grading the severity of neurotoxicity in histopathology slides. Quantification of the extent of neurotoxicity supports qualitative histopathological analysis and provides a better understanding of the global extent of brain damage. Stereological techniques such as the utilization of an optical fractionator provide an unbiased quantification of the neuronal damage; however, the process is time-consuming. Advent of whole slide imaging (WSI) introduced digital image analysis which made quantification of neurotoxicity automated, faster and with reduced bias, making statistical comparisons possible. Although automated to a certain level, simple digital image analysis requires manual efforts of experts which is time-consuming and limits analysis of large datasets. Digital image analysis coupled with a deep learning artificial intelligence model provides a good alternative solution to time-consuming stereological and simple digital analysis. Deep learning models could be trained to identify damaged or dead neurons in an automated fashion. This review has focused on and discusses studies demonstrating the role of deep learning in segmentation of brain regions, toxicity detection and quantification of degenerated neurons as well as the estimation of area/volume of degeneration.

Assessment of Macrophage and Blood‐Brain Barrier Function in the Resilient C57BL/6 Mouse and Non‐Resilient Balb/c Mouse

Macrophage activation disrupts the blood‐brain barrier (BBB) in neurological and psychological diseases. However, the contribution of individual differences in macrophage and BBB phenotype in people’s susceptibility to these illnesses is largely unknown. We proposed to use a mouse model of psychological resilience to understand the role of macrophage and BBB function in disease susceptibility. In this model, the adoptive transfer of macrophages between Balb/c and C57BL/6 mice confers susceptibility or resistance, respectively, to development of depressive behaviors, thus highlighting the role of individual differences in macrophage function in brain‐related disorders.Our objective was to determine BBB function (permeability of brain microvascular endothelial cell (BMEC) monolayer), BBB response to endotoxin challenge (E. colilipopolysaccharide (LPS)), and macrophage activation (via LPS) in C57BL/6 mice and Balb/c mice. We hypothesized that 1) Balb/c BMEC monolayer will be innately more permeable than C57BL/6 BMEC monolayer, 2) Balb/c BMECs will be more sensitive to LPS, and 3) LPS‐activated bone‐marrow derived macrophages (BMMs) of Balb/c mice will induce greater disruption of BMECs than C57BL/6 BMMs.BMECs derived from C57BL/6 and Balb/c mice were seeded on 96‐well plates outfitted with electrodes to measure transendothelial electrical resistance (TEER) by Electric Cell‐substrate Impedance Sensing (Applied Biophysics, Inc.). After a stable monolayer was established, cells were treated with LPS (0, 25, 50, 75 or 100 ng/mL), or media conditioned by Balb/c or C57BL/6 BMMs plated at a density of 20,000 cells/cm2. BMMs were treated with 100 ng/mL of LPS (activated) or plain media (non‐activated). After 24 hours, the media was exchanged with fresh plain media, and collected after an additional 24 hours. Experiments were performed on three different days to obtain N=3/group.C57BL/6 BMEC monolayers displayed higher maximal TEER than Balb/c BMEC monolayer (24684 ± 1291 versus 19950 ± 1462 ohms cm2), and were significantly more sensitive to the TEER‐reducing effects of LPS after 12 hours of LPS exposure (FStrain(1, 20) = 7.4, P = 0.01). Media conditioned with LPS‐activated Balb/c BMMs induced significant disruption of C57BL/6 BMECs for up to 24 hours (FStrain(1,20) = 18.9, P = 0.0003), but had no effect on Balb/c BMECs. Media conditioned with non‐activated Balb/c BMMs, or activated or non‐activated C57BL/6 BMMs had no effect on TEER of either types of BMECs.Our study demonstrated that C57BL/6 mice have a less permeable BBB that is more sensitive to endotoxin challenge compared to BBB of Balb/c mice. C57BL/6 mice also possess macrophages that, when activated, are less capable of inducing BBB leakiness compared to Balb/c mice. The innate differences in macrophage and BBB function between the resilient C57BL/6 mouse and the non‐resilient Balb/c mouse likely play a role in their susceptibility to neurological/psychological disorders. We postulate that people’s susceptibility to brain‐related disorders is partly due to the interaction of their genetically‐determined BBB and macrophage phenotypes.Support or Funding InformationThe views expressed in this abstract are those of the authors and do not reflect the official policy or position of the Department of the Army, Department of Defense, or the U.S. Government.

Ultrasmall Core-Shell Silica Nanoparticles for Precision Drug Delivery in a High-Grade Malignant Brain Tumor Model.

Small-molecule inhibitors have revolutionized treatment of certain genomically defined solid cancers. Despite breakthroughs in treating systemic disease, central nervous system (CNS) metastatic progression is common, and advancements in treating CNS malignancies remain sparse. By improving drug penetration across a variably permeable blood-brain barrier and diffusion across intratumoral compartments, more uniform delivery and distribution can be achieved to enhance efficacy. Ultrasmall fluorescent core-shell silica nanoparticles, Cornell prime dots (C' dots), were functionalized with αv integrin-binding (cRGD), or nontargeting (cRAD) peptides, and PET labels (124I, 89Zr) to investigate the utility of dual-modality cRGD-C' dots for enhancing accumulation, distribution, and retention (ADR) in a genetically engineered mouse model of glioblastoma (mGBM). mGBMs were systemically treated with 124I-cRGD- or 124I-cRAD-C' dots and sacrificed at 3 and 96 hours, with concurrent intravital injections of FITC-dextran for mapping blood-brain barrier breakdown and the nuclear stain Hoechst. We further assessed target inhibition and ADR following attachment of dasatinib, creating nanoparticle-drug conjugates (Das-NDCs). Imaging findings were confirmed with ex vivo autoradiography, fluorescence microscopy, and p-S6RP IHC. Improvements in brain tumor delivery and penetration, as well as enhancement in the ADR, were observed following administration of integrin-targeted C' dots, as compared with a nontargeted control. Furthermore, attachment of the small-molecule inhibitor, dasatinib, led to its successful drug delivery throughout mGBM, demonstrated by downstream pathway inhibition. These results demonstrate that highly engineered C' dots are promising drug delivery vehicles capable of navigating the complex physiologic barriers observed in a clinically relevant brain tumor model.

Sugar alcohol provides imaging contrast in cancer detection

Clinical imaging is widely used to detect, characterize and stage cancers in addition to monitoring the therapeutic progress. Magnetic resonance imaging (MRI) aided by contrast agents utilizes the differential relaxivity property of water to distinguish between tumorous and normal tissue. Here, we describe an MRI contrast method for the detection of cancer using a sugar alcohol, maltitol, a common low caloric sugar substitute that exploits the chemical exchange saturation transfer (CEST) property of the labile hydroxyl group protons on maltitol (malCEST). In vitro studies pointed toward concentration and pH-dependent CEST effect peaking at 1 ppm downfield to the water resonance. Studies with control rats showed that intravenously injected maltitol does not cross the intact blood-brain barrier (BBB). In glioma carrying rats, administration of maltitol resulted in the elevation of CEST contrast in the tumor region only owing to permeable BBB. These preliminary results show that this method may lead to the development of maltitol and other sugar alcohol derivatives as MRI contrast agents for a variety of preclinical imaging applications.