Epen-22. Single-cell RNA sequencing identifies upregulation of IKZF1 in PFA2 myeloid subpopulation driving an anti-tumor phenotype

We have previously shown immune gene phenotype variations between posterior fossa ependymoma subgroups. PFA1 tumors chronically secrete IL-6, which pushes the infiltrating myeloid cells to an immune suppressive function. In contrast, PFA2 tumors have a more immune activated phenotype and have a better prognosis. The objective of this study was to use single-cell(sc) RNAseq to descriptively characterize the infiltrating myeloid cells. We analyzed approximately 8500 cells from 21 PFA patient samples and used advanced machine learning techniques to identify distinct myeloid and lymphoid subpopulations. The myeloid compartment was difficult to interrupt as the data shows a continuum of gene expression profiles exist within PFA1 and PFA2. Through lineage tracing, we were able to tease out that PFA2 myeloid cells expressed more genes associated with an anti-viral response (MHC II, TNF-a, interferon-gamma signaling); while PFA1 myeloid cells had genes associated with an immune suppressive phenotype (angiogenesis, wound healing, IL-10). Specifically, we found expression of IKZF1 was upregulated in PFA2 myeloid cells. IKZF1 regulates differentiation of myeloid cells toward M1 or M2 phenotype through upregulation of either IRF5 or IRF4 respectively. IRF5 expression correlated with IKZF1, being predominately expressed in the PFA2 myeloid cell subset. IKZF1 is also involved in T-cell activation. While we have not completed our characterization of the T-cell subpopulation, we did find significantly more T-cell infiltration in PFA2 than PFA1. Moving forward these studies will provide us with valuable information regarding the molecular switches involved in the tumor-immune microenvironment and to better develop immunotherapy for PFA ependymoma

EPEN-11. TUMOR DIFFERENTIATION IMPACTS THE BIOLOGY OF RECURRENCE IN CHILDHOOD POSTERIOR FOSSA EPENDYMOMA

Ependymoma (EPN) of childhood is curable in only 50% of cases, with recurrences in the remainder that are refractory to treatment. In recent years significant advances have been made in understanding the molecular and cellular biology of EPN. Recent studies show that PFA subgroup EPN are comprised of multiple neoplastic subpopulations that show undifferentiated, differentiated and mesenchymal characteristics. These studies focused on tumor at presentation, with recurrent EPN being less well understood. In the present longitudinal study we examine changes in neoplastic cell heterogeneity in serial presentations of PFA EPN using deconvolution (Cibersort) of bulk RNAseq data. Analysis of a cohort of 48 PFA EPN presenting at Children’s Colorado showed survival and PFA1/PFA2 subtype assignment was associated with the proportion of individual neoplastic subpopulations as determined by deconvolution. Tumors that subsequently regrew had a significantly higher estimated proportion of undifferentiated EPN cells (UEC) at presentation, than those that were non-recurrent after 5 years follow-up. This outcome association potentially age related, as UEC proportions are significantly higher in PFA arising in children < 1 year old who have a particularly poor prognosis. Changes in PFA neoplastic subpopulations at recurrence was performed in two cohorts of patients from Children’s Colorado (n=23) and Nottingham, UK (n=15). As a whole, no subpopulation proportion was significantly changed at recurrence. However, separation of PFA into subtypes PFA1 and PFA2 revealed an increase in the proportion of the cilia-differentiated EPN cell subpopulation is more frequent event in PFA1 (15/24), and rare in PFA2 (2/11). Changes in other neoplastic subpopulations at recurrence were smaller and only seen in PFA1, both UEC and mesenchymal subpopulations being lower at recurrence. In summary, only PFA1 showed dynamic changes in neoplastic subpopulation proportions at recurrence, with potential impacts on transcriptomic based-subgroup assignment, whereas PFA2 proportions remained largely stable

EPEN-07. SINGLE-CELL RNA SEQUENCING IDENTIFIES A UNIQUE MYELOID SUBPOPULATION ASSOCIATED WITH MESENCHYMAL TUMOR SUBPOPULATION IN POOR OUTCOME PEDIATRIC EPENDYMOMA

We have previously shown immune gene phenotype variations between posterior fossa ependymoma subgroups. PFA1 tumors chronically secrete IL-6, which induces secretion of myeloid cell IL-8 and pushes the infiltrating myeloid cells to an immune suppressive function. In contrast, PFA2 tumors have a more immune activated phenotype associated with a better prognosis. The objective of this study was to use single-cell(sc) RNAseq to descriptively characterize the infiltrating myeloid cells. We analyzed approximately 8500 cells from 21 PFA patient samples. Using advanced machine learning, we identified eight myeloid cell subpopulations with unique gene expression profiles. Interestingly, only one subpopulation was significantly enriched in PFA1 tumors. This subpopulation, denoted as the hypoxia myeloid subpopulation, was defined by genes associated with angiogenesis, response to hypoxia, wound healing, cell migration, neutrophil activation and response to oxygen levels. These myeloid cells also share similar gene expression profile to a mesenchymal tumor subpopulation (MEC) enriched in PFA1 and associated with poor outcome in EPN patients. This tumor subpopulation was the only population expressing IL-6. Using immunohistochemistry, we found the hypoxia myeloid located in regions of tumor necrosis and perivascular niches. The MEC cells were also more abundant in these regions. In an independent single-cell cytokine release assay, we identified eight subpopulations of functional myeloid cells. One subpopulation significantly secreted IL-8, which represented the hypoxia subpopulation based on IL-8 gene expression in the scRNAseq dataset. This data suggests the tumor necrosis resulting in the development of MEC tumor subpopulation is driving the immune suppressive myeloid phenotype in PFA1 tumors through polarization of myeloid cells to the hypoxia subpopulation. Further studies are needed to determine how these myeloid cells interact with the lymphocyte subpopulation

EPEN-07. SINGLE-CELL RNA SEQUENCING IDENTIFIES A UNIQUE MYELOID SUBPOPULATION ASSOCIATED WITH MESENCHYMAL TUMOR SUBPOPULATION IN POOR OUTCOME PEDIATRIC EPENDYMOMA

We have previously shown immune gene phenotype variations between posterior fossa ependymoma subgroups. PFA1 tumors chronically secrete IL-6, which induces secretion of myeloid cell IL-8 and pushes the infiltrating myeloid cells to an immune suppressive function. In contrast, PFA2 tumors have a more immune activated phenotype associated with a better prognosis. The objective of this study was to use single-cell(sc) RNAseq to descriptively characterize the infiltrating myeloid cells. We analyzed approximately 8500 cells from 21 PFA patient samples. Using advanced machine learning, we identified eight myeloid cell subpopulations with unique gene expression profiles. Interestingly, only one subpopulation was significantly enriched in PFA1 tumors. This subpopulation, denoted as the hypoxia myeloid subpopulation, was defined by genes associated with angiogenesis, response to hypoxia, wound healing, cell migration, neutrophil activation and response to oxygen levels. These myeloid cells also share similar gene expression profile to a mesenchymal tumor subpopulation (MEC) enriched in PFA1 and associated with poor outcome in EPN patients. This tumor subpopulation was the only population expressing IL-6. Using immunohistochemistry, we found the hypoxia myeloid located in regions of tumor necrosis and perivascular niches. The MEC cells were also more abundant in these regions. In an independent single-cell cytokine release assay, we identified eight subpopulations of functional myeloid cells. One subpopulation significantly secreted IL-8, which represented the hypoxia subpopulation based on IL-8 gene expression in the scRNAseq dataset. This data suggests the tumor necrosis resulting in the development of MEC tumor subpopulation is driving the immune suppressive myeloid phenotype in PFA1 tumors through polarization of myeloid cells to the hypoxia subpopulation. Further studies are needed to determine how these myeloid cells interact with the lymphocyte subpopulation

EPEN-07. SINGLE-CELL RNA SEQUENCING IDENTIFIES A UNIQUE MYELOID SUBPOPULATION ASSOCIATED WITH MESENCHYMAL TUMOR SUBPOPULATION IN POOR OUTCOME PEDIATRIC EPENDYMOMA

We have previously shown immune gene phenotype variations between posterior fossa ependymoma subgroups. PFA1 tumors chronically secrete IL-6, which induces secretion of myeloid cell IL-8 and pushes the infiltrating myeloid cells to an immune suppressive function. In contrast, PFA2 tumors have a more immune activated phenotype associated with a better prognosis. The objective of this study was to use single-cell(sc) RNAseq to descriptively characterize the infiltrating myeloid cells. We analyzed approximately 8500 cells from 21 PFA patient samples. Using advanced machine learning, we identified eight myeloid cell subpopulations with unique gene expression profiles. Interestingly, only one subpopulation was significantly enriched in PFA1 tumors. This subpopulation, denoted as the hypoxia myeloid subpopulation, was defined by genes associated with angiogenesis, response to hypoxia, wound healing, cell migration, neutrophil activation and response to oxygen levels. These myeloid cells also share similar gene expression profile to a mesenchymal tumor subpopulation (MEC) enriched in PFA1 and associated with poor outcome in EPN patients. This tumor subpopulation was the only population expressing IL-6. Using immunohistochemistry, we found the hypoxia myeloid located in regions of tumor necrosis and perivascular niches. The MEC cells were also more abundant in these regions. In an independent single-cell cytokine release assay, we identified eight subpopulations of functional myeloid cells. One subpopulation significantly secreted IL-8, which represented the hypoxia subpopulation based on IL-8 gene expression in the scRNAseq dataset. This data suggests the tumor necrosis resulting in the development of MEC tumor subpopulation is driving the immune suppressive myeloid phenotype in PFA1 tumors through polarization of myeloid cells to the hypoxia subpopulation. Further studies are needed to determine how these myeloid cells interact with the lymphocyte subpopulation

EPEN-07. SINGLE-CELL RNA SEQUENCING IDENTIFIES A UNIQUE MYELOID SUBPOPULATION ASSOCIATED WITH MESENCHYMAL TUMOR SUBPOPULATION IN POOR OUTCOME PEDIATRIC EPENDYMOMA

We have previously shown immune gene phenotype variations between posterior fossa ependymoma subgroups. PFA1 tumors chronically secrete IL-6, which induces secretion of myeloid cell IL-8 and pushes the infiltrating myeloid cells to an immune suppressive function. In contrast, PFA2 tumors have a more immune activated phenotype associated with a better prognosis. The objective of this study was to use single-cell(sc) RNAseq to descriptively characterize the infiltrating myeloid cells. We analyzed approximately 8500 cells from 21 PFA patient samples. Using advanced machine learning, we identified eight myeloid cell subpopulations with unique gene expression profiles. Interestingly, only one subpopulation was significantly enriched in PFA1 tumors. This subpopulation, denoted as the hypoxia myeloid subpopulation, was defined by genes associated with angiogenesis, response to hypoxia, wound healing, cell migration, neutrophil activation and response to oxygen levels. These myeloid cells also share similar gene expression profile to a mesenchymal tumor subpopulation (MEC) enriched in PFA1 and associated with poor outcome in EPN patients. This tumor subpopulation was the only population expressing IL-6. Using immunohistochemistry, we found the hypoxia myeloid located in regions of tumor necrosis and perivascular niches. The MEC cells were also more abundant in these regions. In an independent single-cell cytokine release assay, we identified eight subpopulations of functional myeloid cells. One subpopulation significantly secreted IL-8, which represented the hypoxia subpopulation based on IL-8 gene expression in the scRNAseq dataset. This data suggests the tumor necrosis resulting in the development of MEC tumor subpopulation is driving the immune suppressive myeloid phenotype in PFA1 tumors through polarization of myeloid cells to the hypoxia subpopulation. Further studies are needed to determine how these myeloid cells interact with the lymphocyte subpopulation

Medulloblastoma Exome Sequencing Uncovers Subtype-Specific Somatic Mutations

Medulloblastomas are the most common malignant brain tumors in children1. Identifying and understanding the genetic events that drive these tumors is critical for the development of more effective diagnostic, prognostic and therapeutic strategies. Recently, our group and others described distinct molecular subtypes of medulloblastoma based on transcriptional and copy number profiles2–5. Here, we utilized whole exome hybrid capture and deep sequencing to identify somatic mutations across the coding regions of 92 primary medulloblastoma/normal pairs. Overall, medulloblastomas exhibit low mutation rates consistent with other pediatric tumors, with a median of 0.35 non-silent mutations per megabase. We identified twelve genes mutated at statistically significant frequencies, including previously known mutated genes in medulloblastoma such as CTNNB1, PTCH1, MLL2, SMARCA4 and TP53. Recurrent somatic mutations were identified in an RNA helicase gene, DDX3X, often concurrent with CTNNB1 mutations, and in the nuclear co-repressor (N-CoR) complex genes GPS2, BCOR, and LDB1, novel findings in medulloblastoma. We show that mutant DDX3X potentiates transactivation of a TCF promoter and enhances cell viability in combination with mutant but not wild type beta-catenin. Together, our study reveals the alteration of Wnt, Hedgehog, histone methyltransferase and now N-CoR pathways across medulloblastomas and within specific subtypes of this disease, and nominates the RNA helicase DDX3X as a component of pathogenic beta-catenin signaling in medulloblastoma

The Forest Observation System, building a global reference dataset for remote sensing of forest biomass

International audienceForest biomass is an essential indicator for monitoring the Earth's ecosystems and climate. It is a critical input to greenhouse gas accounting, estimation of carbon losses and forest degradation, assessment of renewable energy potential, and for developing climate change mitigation policies such as REDD+, among others. Wall-to-wall mapping of aboveground biomass (aGB) is now possible with satellite remote sensing (RS). However, RS methods require extant, up-to-date, reliable, representative and comparable in situ data for calibration and validation. Here, we present the Forest Observation System (FOS) initiative, an international cooperation to establish and maintain a global in situ forest biomass database. aGB and canopy height estimates with their associated uncertainties are derived at a 0.25 ha scale from field measurements made in permanent research plots across the world's forests. all plot estimates are geolocated and have a size that allows for direct comparison with many RS measurements. The FOS offers the potential to improve the accuracy of RS-based biomass products while developing new synergies between the RS and ground-based ecosystem research communities

Global, regional, and national burden of neurological disorders during 1990-2015 : a systematic analysis for the Global Burden of Disease Study 2015

Background Comparable data on the global and country-specific burden of neurological disorders and their trends are crucial for health-care planning and resource allocation. The Global Burden of Diseases, Injuries, and Risk Factors (GBD) Study provides such information but does not routinely aggregate results that are of interest to clinicians specialising in neurological conditions. In this systematic analysis, we quantified the global disease burden due to neurological disorders in 2015 and its relationship with country development level. Methods We estimated global and country-specific prevalence, mortality, disability-adjusted life-years (DALYs), years of life lost (YLLs), and years lived with disability (YLDs) for various neurological disorders that in the GBD classification have been previously spread across multiple disease groupings. The more inclusive grouping of neurological disorders included stroke, meningitis, encephalitis, tetanus, Alzheimer's disease and other dementias, Parkinson's disease, epilepsy, multiple sclerosis, motor neuron disease, migraine, tension-type headache, medication overuse headache, brain and nervous system cancers, and a residual category of other neurological disorders. We also analysed results based on the Socio-demographic Index (SDI), a compound measure of income per capita, education, and fertility, to identify patterns associated with development and how countries fare against expected outcomes relative to their level of development. Findings Neurological disorders ranked as the leading cause group of DALYs in 2015 (250.7 [95% uncertainty interval (UI) 229.1 to 274.7] million, comprising 10.2% of global DALYs) and the second-leading cause group of deaths (9.4 [9.1 to 9.7] million], comprising 16.8% of global deaths). The most prevalent neurological disorders were tensiontype headache (1505 9 [UI 1337.3 to 1681.6 million cases]), migraine (958.8 [872.1 to 1055.6] million), medication overuse headache (58.5 [50.8 to 67.4 million]), and Alzheimer's disease and other dementias (46.0 [40.2 to 52.7 million]). Between 1990 and 2015, the number of deaths from neurological disorders increased by 36.7%, and the number of DALYs by 7.4%. These increases occurred despite decreases in age-standardised rates of death and DALYs of 26.1% and 29.7%, respectively; stroke and communicable neurological disorders were responsible for most of these decreases. Communicable neurological disorders were the largest cause of DALYs in countries with low SDI. Stroke rates were highest at middle levels of SDI and lowest at the highest SDI. Most of the changes in DALY rates of neurological disorders with development were driven by changes in YLLs. Interpretation Neurological disorders are an important cause of disability and death worldwide. Globally, the burden of neurological disorders has increased substantially over the past 25 years because of expanding population numbers and ageing, despite substantial decreases in mortality rates from stroke and communicable neurological disorders. The number of patients who will need care by clinicians with expertise in neurological conditions will continue to grow in coming decades. Policy makers and health-care providers should be aware of these trends to provide adequate services.Peer reviewe