Children with medulloblastoma and ependymoma are treated with a multidisciplinary approach that incorporates surgery, radiotherapy, and chemotherapy; however, overall survival rates for patients with high-risk disease remain unsatisfactory. Data indicate that plant-derived cannabinoids are effective against adult glioblastoma; however, preclinical evidence supporting their use in pediatric brain cancers is lacking. Here we investigated the potential role for Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD) in medulloblastoma and ependymoma. Dose-dependent cytotoxicity of medulloblastoma and ependymoma cells was induced by THC and CBD in vitro, and a synergistic reduction in viability was observed when both drugs were combined. Mechanistically, cannabinoids induced cell cycle arrest, in part by the production of reactive oxygen species, autophagy, and apoptosis; however, this did not translate to increased survival in orthotopic transplant models despite being well tolerated. We also tested the combination of cannabinoids with the medulloblastoma drug cyclophosphamide, and despite some in vitro synergism, no survival advantage was observed in vivo. Consequently, clinical benefit from the use of cannabinoids in the treatment of high-grade medulloblastoma and ependymoma is expected to be limited. This study emphasizes the importance of preclinical models in validating therapeutic agent efficacy prior to clinical trials, ensuring that enrolled patients are afforded the most promising therapies available.

We recently reported that the offspring of mice treated during pregnancy with the microbial-derived immunomodulator OM-85 manifest striking resistance postnatally to allergic airways inflammation, and localised the potential treatment target to the fetal cDC progenitor compartment which expands to increase the pool of precursors available at birth, enabling accelerated postnatal seeding of the lung mucosal cDC network required for establishment of immunological homeostasis in the airways. Here, we profile maternal OM-85 treatment-associated transcriptomic signatures in fetal bone marrow, and identify a series of immunometabolic pathways which provide essential metabolites for accelerated myelopoiesis, that are hallmarks of classical “immune training”. In addition, the cDC progenitor compartment displayed treatment-associated activation of the XBP1-ERN1 signalling axis which has previously been shown to be essential for tissue survival of cDC, particularly within the lung microenvironment. Our forerunner studies indicate uniquely rapid turnover of airway mucosal cDCs at baseline, with further large-scale upregulation of population dynamics during aeroallergen and/or pathogen challenge. XBP1-ERN1 signalling plays a key role in mitigation of ER stress-associated toxicity which frequently accompanies DC hyper-activation during intense immunoinflammatory responses, and we suggest that enhanced capacity for XBP1-ERN1-dependent cDC survival within the airway mucosal tissue microenvironment may be a crucial element of the OM-85-mediated transplacental “innate immune training” process which results in enhanced resistance to airway inflammatory disease during the high-risk early postnatal period.

Rationale: Atopic asthma is a persistent disease characterized by intermittent wheeze and progressive loss of lung function. The disease is thought to be driven primarily by chronic aeroallergen-induced Th2-associated airways inflammation. However, the vast majority of atopics do not develop asthma-related wheeze, despite ongoing exposure to aeroallergens to which they are strongly sensitized, indicating that additional pathogenic mechanism(s) operate in conjunction with Th2 immunity to drive asthma pathogenesis. Objectives: Employ systems level analyses to identify inflammation-associated gene networks operative at baseline in sputum-derived RNA from house dust mite-sensitized (HDM ^s ) subjects with/without wheezing history; identify networks characteristic of the ongoing asthmatic state. All subjects resided in the constitutively-HDMhigh Perth environment. Methods: Genome wide expression profiling by RNASeq followed by gene coexpression network analysis. Measurements/Results: HDM ^s -nonwheezers displayed baseline gene expression in sputum including IL-5, IL-13 and CCL17. HDM ^s -wheezers showed equivalent expression of these classical Th2-effector genes but their overall baseline sputum signatures were more complex, comprising hundreds of Th2-associated and epithelial-associated genes, networked into two separate coexpression modules. The first module was connected by the hubs EGFR, ERBB2, CDH1 and IL-13. The second module was associated with CDHR3, and contained genes that control mucociliary clearance. Conclusions: Our findings provide new insight into the inflammatory mechanisms operative at baseline in the airway mucosal microenvironment in atopic asthmatics undergoing natural perennial aeroallergen exposure. The molecular mechanism(s) that determine susceptibility to asthma amongst these subjects involve interactions between Th2-and epithelial function-associated genes within a complex co-expression network, which is not operative in equivalently sensitized/exposed atopic non-asthmatics. Funding: This study was funded by the Asthma Foundation WA, the Department of Health WA, and the NHMRC. AB is funded by a BrightSpark Foundation McCusker Fellowship. GLH is a NHMRC Fellow. AG is supported by the McCusker Charitable Foundation Bioinformatics Centre. ACJ is a recipient of an Australian Postgraduate Award and a Top-Up Award from the University of Western Australia.

Infection-associated inflammatory stress during pregnancy is the most common cause of fetal growth restriction and/or miscarriage. Treatment strategies for protection of at-risk mothers are limited to a narrow range of vaccines, which do not cover the bulk of the common pathogens most frequently encountered. Employing mouse models, we demonstrate that oral treatment during pregnancy with a microbial-derived immunomodulator (OM85 ^TM ), currently used clinically for attenuation of infection-associated airway inflammatory symptoms in infants-adults, markedly reduces risk for fetal loss/growth restriction resulting from maternal challenge with bacterial LPS or influenza. Focusing on LPS exposure, we demonstrate that the key molecular indices of maternal inflammatory stress, notably high levels of RANTES, MIP-1a, CCL2, IL-8 and G-CSF in gestational tissues/serum, are abrogated by OM85 pretreatment. Systems-level analyses conducted in parallel employing RNASeq revealed that OM85 pretreatment selectively tunes LPS-induced activation in maternal gestational tissues for attenuated expression of TNF-, IL1-, and IFNg-driven that drive production of these pro-inflammatory cytokines, without constraining Type1-IFN-associated networks central to first-line anti-microbial defense. This study suggests that broad-spectrum protection-of-pregnancy against infection-associated inflammatory stress, without compromising capacity for efficient pathogen eradication, represents an achievable therapeutic goal. Disclosure: This study was funded principally by Nation Health and Medical Research Council (NHMRC) of Australia with supplementary support provided by OM Pharma (Geneva, Switzerland). CP is an employee of OM Pharma (Vifor Pharma). The other authors declare that they have no conflict of interest.

Background: Multiple regulatory mechanisms have been identified employing conventional hypothesis-driven approaches as contributing to allergen-specific immunotherapy outcomes, but understanding of how these integrate to maintain immunological homeostasis is incomplete. Objective: To explore the potential for unbiased systems-level gene co-expression network analysis to advance understanding of immunotherapy mechanisms. Methods: We profiled genome-wide allergen-specific Th-memory responses prospectively across 24mths of subcutaneous immunotherapy (SCIT) in 25 rhinitics, documenting changes in immunoinflammatory pathways and associated co-expression networks and their relationships to symptom scores to 36mths. Results: Prior to immunotherapy, mite-specific Th-memory response networks involved multiple discrete co-expression modules including those related to Th2-, Type1-IFN-, Inflammation-, and FOXP3/IL2-associated signalling. A signature comprising 109 genes correlated with symptom scores, and these mapped to cytokine signalling/T-cell activation-associated pathways, with upstream drivers including hallmark Th1/Th2-and inflammation-associated genes. Reanalysis after 3.5mths SCIT updosing detected minimal changes to pathway/upstream regulator profiles despite 32.5% reduction in symptoms, however network analysis revealed underlying merging of FOXP3/IL2-with Inflammation-and Th2-associated modules. By 12mths on SCIT, symptoms had reduced by 41% without further significant changes to pathway/upstream regulator or network profiles. Continuing SCIT to 24mths stabilised symptoms at 47% of baseline, accompanied by upregulation of the Type1-IFN-associated network module and its merging into the Th2/FOXP3/IL2/Inflammation module. Conclusions: SCIT stimulates progressive integration of Th-memory-associated Th2-,FOXP3/IL2-, Inflammation-, and Type1-IFN-signalling subnetworks, forming a single highly integrated co-expression network module, maximising potential for stable homeostatic control of allergen-specific Th2 responses via cross-regulation. Th2-anatogonistic Type1-IFN signalling may play a key role in stabilising clinical effects of SCIT. Clinical Implication: Stabilisation of the clinical effectiveness of SCIT involves recruitment of Th2-antagonistic Type 1 IFN-dependent signalling into the overall gene co-expression network underlying the allergen-specific Th-memory response, and this does not occur until the 2 ^nd year of treatment. Capsule summary: SCIT-induced rewiring of the gene network governing allergen-specific Th2-memory, as opposed to selective upregulation of genes associated with regulatory functions, underlies the clinical effectiveness of immunotherapy.