Biology subjects

Mitchell, J.

Publications and source records attributed to Mitchell, J..

At least 55 records · Page 3Linked to original sources

bioRxiv2022

Clinical application of tumour in normal contamination assessment from whole genome sequencing

The unexpected contamination of normal samples with tumour cells reduces variant detection sensitivity, compromising downstream analyses in canonical tumour-normal analyses. Leveraging whole-genome sequencing data available at Genomics England, we develop a tool for normal sample contamination assessment, which we validate in silico and against minimal residual disease testing. From a systematic review of 771 patients with haematological malignancies and sarcomas, we find contamination across a range of cancer clinical indications and DNA sources, with highest prevalence in saliva samples from acute myeloid leukaemia patients, and sorted CD3+ T-cells from myeloproliferative neoplasms. Further exploration reveals 108 hotspot mutations in genes associated with haematological cancers at risk of being subtracted by standard variant calling pipelines. Our work highlights the importance of contamination assessment for accurate somatic variants detection in research and clinical settings, especially with large-scale sequencing projects being utilised to deliver accurate data from which to make clinical decisions for patient care.

bioinformatics↗

bioRxiv2022

Cancer-driving mutations are enriched in genic regions intolerant to germline variation

Large reference datasets of protein-coding variation in human populations have allowed us to determine which genes and genic sub-regions are intolerant to germline genetic variation. There is also a growing number of genes implicated in severe Mendelian diseases that overlap with genes implicated in cancer. Here, we hypothesized that mitotically mutable genic sub-regions that are intolerant to germline variation are enriched for cancer-driving mutations. We introduce a new metric, OncMTR, which uses 125,748 exomes in the gnomAD database to identify genic sub-regions intolerant to germline variation but enriched for hematologic somatic variants. We demonstrate that OncMTR can significantly predict driver mutations implicated in hematologic malignancies. Divergent OncMTR regions were enriched for cancer-relevant protein domains, and overlaying OncMTR scores on protein structures identified functionally important protein residues. Finally, we performed a rare variant, gene-based collapsing analysis on an independent set of 394,694 exomes from the UK Biobank and find that OncMTR dramatically improves genetic signals for hematologic malignancies. Our web app enables easy visualization of OncMTR scores for each protein-coding gene (https://astrazeneca-cgr-publications.github.io/OncMTR-Viewer/).

genomics↗

bioRxiv2021

Disrupting ATXN1 Nuclear Localization in a Knock-in SCA1 Mouse Model Improves a Spectrum of SCA1-Like Phenotypes and their Brain Region Associated Transcriptomic Profiles

Spinocerebellar ataxia type 1 (SCA1) is a dominant trinucleotide repeat neurodegenerative disease characterized by motor dysfunction, cognitive impairment, and premature death. Degeneration of cerebellar Purkinje cells is a frequent and prominent pathological feature of SCA1. We previously showed that transport of ATXN1 to Purkinje cell nuclei is required for pathology, where mutant ATXN1 alters transcription. To examine the role of ATXN1 nuclear localization broadly in SCA1-like disease pathogenesis, CRISPR-Cas9 was used to develop a mouse with the amino acid alteration (K772T) in the nuclear localization sequence of the expanded ATXN1 protein. Characterization of these mice indicates proper nuclear localization of mutant ATXN1 contributes to many disease-like phenotypes including motor dysfunction, cognitive deficits, and premature lethality. RNA sequencing analysis of genes whose expression was corrected to WT levels in Atxn1175QK772T/2Q mice indicates that transcriptomic aspects of SCA1 pathogenesis differ between the cerebellum, brainstem, cerebral cortex, hippocampus, and striatum.

neuroscience↗

bioRxiv2021

Novel insights on obligate symbiont lifestyle and adaptation to chemosynthetic environment as revealed by the giant tubeworm genome

The mutualism between the giant tubeworm Riftia pachyptila and its endosymbiont Candidatus Endoriftia persephone has been extensively researched over the past 40 years. However, the lack of the host whole genome information has impeded the full comprehension of the genotype/phenotype interface in Riftia. Here we described the high-quality draft genome of Riftia, its complete mitogenome, and tissue-specific transcriptomic data. The Riftia genome presents signs of reductive evolution, with gene family contractions exceeding expansions. Expanded gene families are related to sulphur metabolism, detoxification, anti-oxidative stress, oxygen transport, immune system, and lysosomal digestion, reflecting evolutionary adaptations to the vent environment and endosymbiosis. Despite the derived body plan, the developmental gene repertoire in the gutless tubeworm is extremely conserved with the presence of a near intact and complete Hox cluster. Gene expression analyses establishes that the trophosome is a multi-functional organ marked by intracellular digestion of endosymbionts, storage of excretory products and haematopoietic functions. Overall, the plume and gonad tissues both in contact to the environment harbour highly expressed genes involved with cell cycle, programmed cell death, and immunity indicating a high cell turnover and defence mechanisms against pathogens. We posit that the innate immune system plays a more prominent role into the establishment of the symbiosis during the infection in the larval stage, rather than maintaining the symbiostasis in the trophosome. This genome bridges four decades of physiological research in Riftia, whilst simultaneously provides new insights into the development, whole organism functions and evolution in the giant tubeworm.

evolutionary biology↗

bioRxiv2021

Human cytomegalovirus blocks canonical TGFβ signaling during lytic infection to limit induction of type I interferons

Human cytomegalovirus (HCMV) microRNAs (miRNAs) significantly rewire host signaling pathways to support the viral lifecycle and regulate host cell responses. Here we show that SMAD3 expression is regulated by HCMV miR-UL22A and contributes to the IRF7-mediated induction of type I IFNs and IFN-stimulated genes (ISGs) in human fibroblasts. Addition of exogenous TGF{beta} interferes with the replication of a miR-UL22A mutant virus in a SMAD3-dependent manner in wild type fibroblasts, but not in cells lacking IRF7, indicating that downregulation of SMAD3 expression to limit IFN induction is important for efficient lytic replication. These findings uncover a novel interplay between SMAD3 and innate immunity during HCMV infection and highlight the role of viral miRNAs in modulating these responses. Author SummaryCells trigger the interferon (IFN) response to induce the expression of cellular genes that limit virus replication. In turn, viruses have evolved numerous countermeasures to avoid the effects of IFN signaling. Using a microRNA (miRNA) mutant virus we have uncovered a novel means of regulating the IFN response during human cytomegalovirus (HCMV) infection. Lytic HCMV infection induces the production of TGF{beta}, which binds to the TGF{beta} receptor and activates the receptor-associated SMAD SMAD3. SMAD3, together with IRF7, induces the expression of IFN{beta} and downstream IFN-stimulated genes in human fibroblasts. To counteract this, HCMV miR-UL22A, along with other HCMV gene products, directly targets SMAD3 for downregulation. Infection of fibroblasts with a miR-UL22A mutant virus results in enhanced type I IFN production in a SMAD3-dependent manner and the virus is impaired for growth in the presence of TGF{beta}, but only when both SMAD3 and IRF7 are present, highlighting the unique interaction between TGF{beta} and innate immune signaling.

microbiology↗

bioRxiv2021

Diet High in Soybean Oil Increases Susceptibility to Colitis in Mice

The current American diet is high in soybean oil (SO), which consists of unsaturated fatty acids, most notably linoleic acid (LA, C18:2 omega-6). While LA is an essential fatty acid that must be obtained from the diet, high LA consumption has been linked to the development of inflammatory bowel disease (IBD) in humans. Here, we show that a high fat diet (HFD) based on soybean oil increases susceptibility to colitis in wild-type and IL10 knockout mice. It causes immune dysfunction, decreases colon and crypt length and increases intestinal epithelial barrier permeability; these effects were not observed in low LA HFDs. The SO diet also disrupts the balance of isoforms encoded by the IBD susceptibility gene Hepatocyte Nuclear Factor 4 (HNF4). Both the SO diet and an LA gavage cause gut dysbiosis: the SO diet increases the abundance of an adherent, invasive Escherichia coli (AIEC), which can use LA as a carbon source, and the LA gavage decreases the beneficial bacteria Lactobacillus murinus. Metabolomic analysis of both host-associated and cultured bacteria shows that SO increases levels of LA and oxylipins while decreasing eicosapentaenoic acid (EPA, C20:5 omega-3) and endocannabinoids. Our results suggest that excess LA, obtained from a diet high in soybean oil, increases susceptibility to colitis by alterations in intestinal HNF4, gut microbiota and bioactive metabolites.

physiology↗

bioRxiv2021

The recycling endosome protein Rab25 coordinates collective cell movements in the zebrafish surface epithelium

In emerging epithelial tissues, cells undergo dramatic rearrangements to promote tissue shape changes. Dividing cells remain interconnected via transient cytokinetic bridges. Bridges are cleaved during abscission and currently, the consequences of disrupting abscission in developing epithelia are not well understood. We show that the Rab GTPase, Rab25, localizes near cytokinetic midbodies and likely coordinates abscission through endomembrane trafficking in the epithelium of the zebrafish gastrula during epiboly. In maternal-zygotic Rab25a and Rab25b mutant embryos, morphogenic activity tears open persistent apical cytokinetic bridges that failed to undergo timely abscission. Cytokinesis defects result in anisotropic cell morphologies that are associated with a reduction of contractile actomyosin networks. This slows cell rearrangements and alters the viscoelastic responses of the tissue, all of which likely contribute to delayed epiboly. We present a model in which Rab25 trafficking coordinates cytokinetic bridge abscission and cortical actin density, impacting local cell shape changes and tissue-scale forces.

cell biology↗

bioRxiv2021

A heritable androgenic mechanism of female intrasexual competition in cooperatively breeding meerkats

Female intrasexual competition can be intense in cooperatively breeding species, with the dominant breeder or matriarch limiting reproduction in subordinates via aggression, eviction or infanticide. In males, these tendencies bidirectionally link to testosterone, but in females, there has been no systematic investigation of androgen-mediated behaviour within and across generations. In 22 wild meerkat (Suricata suricatta) clans, we show that matriarchs 1) express peak androgen concentrations during late gestation, 2) when displaying peak feeding competition, dominance, and evictions, and 3) relative to subordinates, produce offspring that are more aggressive in early development. Late-gestation, antiandrogen treatment of matriarchs 4) reduced their dominance behaviour, was associated with infrequent evictions, decreased social centrality within the clan, 5) increased aggression in cohabiting subordinate dams, and 6) reduced their offsprings aggression. These effects implicate androgen-mediated aggression in the operation of female sexual selection, and intergenerational transmission of masculinised phenotypes in the evolution of meerkat cooperative breeding.

animal behavior and cognition↗

bioRxiv2020

Human cytomegalovirus UL7, miR-US5-1 and miR-UL112-3p inactivation of FOXO3a protects CD34+ hematopoietic progenitor cells from apoptosis

Human cytomegalovirus (HCMV) infection of myeloid-lineage cells, such as CD34+ hematopoietic progenitor cells (HPCs) or monocytes results in the upregulation of anti-apoptotic cellular proteins that protect the newly infected cells from programmed cell death. The mechanisms used by HCMV to regulate pro-apoptotic cellular proteins upon infection of CD34+ HPCs has not been fully explored. Here we show that HCMV utilizes pUL7, a secreted protein that signals through the FLT3 receptor, and miR-US5-1 and miR-UL112-3p to reduce the abundance and activity of the pro-apoptotic transcription factor FOXO3a at early times after infection of CD34+ HPCs. Regulation of FOXO3a by pUL7, miR-US5-1 and miR-UL112 results in reduced expression of the pro-apoptotic BCL2L11 transcript and protection of CD34+ HPCs from virus-induced apoptosis. These data highlight the importance of both viral proteins and miRNAs in protecting CD34+ HPCs from apoptosis at early times post-infection, allowing for the establishment of latency and maintenance of viral genome-containing cells. ImportanceHuman cytomegalovirus (HCMV) causes serious disease in immunocompromised individuals and is a significant problem during transplantation. The virus can establish a latent infection in CD34+ hematopoietic progenitor cells (HPCs) and periodically reactivate to cause disease in the absence of an intact immune system. What viral gene products are required for successful establishment of latency are still not fully understood. Here we show that both a viral protein and viral miRNAs are required to prevent apoptosis after infection of CD34+ HPCs. HCMV pUL7 and miRNAs miR-US5-1 and miR-UL112-3p act to limit the expression and activation of the transcription factor FOXO3a which in turn reduces expression of pro-apoptotic gene BCL2L11 and prevents virus-induced apoptosis in CD34+ HPCs.

microbiology↗

bioRxiv2020

Pain-related opioidergic and dopaminergic neurotransmission: Dual Meta-Analyses of PET Radioligand Studies

AO_SCPLOWBSTRACTC_SCPLOWMolecular mechanisms of the interaction between pain and reward associated with pain relief processes in the human brain are still incompletely understood. This is partially due to the invasive nature of the available techniques to visualize and measure metabolic activity. Positron Emission Tomography (PET) radioligand studies using radioactive substances are still the only available modality to date that allows for the investigation of the molecular mechanisms in the human brain. For pain and reward studies, the most commonly studied PET radiotracers are [11C]-carfentanil (CFN) and [11C]- or [18F]-diprenorphine (DPN), which bind to opioid receptors, and [11C]-raclopride (RAC) and [18F]-fallypride (FAL) tracers, which bind to dopamine receptors. The current meta-analysis looks at 15 pain-related studies using opioid radioligands and 8 studies using dopamine radioligands in an effort to consolidate the available data into the most likely activated regions. Our primary goal was to identify regions of shared opioid/dopamine neurotransmission during pain-related experiences. SDM analysis of previously published voxel coordinate data showed that opioidergic activations were strongest in the bilateral caudate, thalamus, right putamen, cingulate gyrus, midbrain, inferior frontal gyrus, and left superior temporal gyrus. The dopaminergic studies showed that the bilateral caudate, thalamus, right putamen, cingulate gyrus, and left putamen had the highest activations. We were able to see a clear overlap between opioid and dopamine activations in a majority of the regions during pain-related processing, though there were some unique areas of dopaminergic activation such as the left putamen. Regions unique to opioidergic activation include the midbrain, inferior frontal gyrus, and left superior temporal gyrus. By investigating the regions of dopaminergic and opioidergic activation, we can potentially provide more targeted treatment to these sets of receptors in patients with pain conditions. These findings could eventually assist in the development of more targeted medication in order to help treat pain conditions and simultaneously prevent physical dependency.

neuroscience↗

bioRxiv2020

Dendritic localization of mRNA in Drosophila Mushroom Body Output Neurons

Memory-relevant neuronal plasticity is believed to require local translation of new proteins at synapses. Understanding this process requires the visualization of the relevant mRNAs within these neuronal compartments. Here we used single-molecule fluorescence in situ hybridization (smFISH) to localize mRNAs at subcellular resolution in the adult Drosophila brain. mRNAs for subunits of nicotinic acetylcholine receptors and kinases could be detected within the dendrites of co-labelled Mushroom Body Output Neurons (MBONs) and their relative abundance showed cell-specificity. Moreover, aversive olfactory learning produced a transient increase in the level of CaMKII mRNA within the dendritic compartments of the {gamma}5{beta}'2a MBONs. Localization of specific mRNAs in MBONs before and after learning represents a critical step towards deciphering the role of dendritic translation in the neuronal plasticity underlying behavioural change in Drosophila.

neuroscience↗

bioRxiv2020

Human papillomavirus 16 positive cervical cancer in Guatemala: The D2 and D3 sublineages differ in integration rate and age of diagnosis

Human papillomavirus (HPV) 16 displays substantial sequence variation; four HPV16 lineages (A, B, C, D) have been described, as well as multiple sub-lineages. To identify molecular events associated with HPV16 carcinogenesis we evaluated viral variation, the integration of HPV16, and somatic mutation in 96 cervical cancer samples from Guatemala. A total of 64% (60/94) of the samples had integrated HPV16 sequences, and integration was associated with an earlier age of diagnosis (P=0.0007) and pre-menopausal disease. HPV16 integration sites were broadly distributed in the genome but in one tumor, HPV16 integrated into the promoter of the interferon regulatory factor 4 (IRF4) gene, which plays an important role in the regulation of the interferon response to viral infection. The HPV16 D2 and D3 sub-lineages were found in 23% and 30% of the tumors, respectively and were significantly associated with adenocarcinoma. D2-positive tumors had a higher rate of integration (P=0.011), earlier age of diagnosis (P=0.012), and a lower rate of somatic mutation (P=0.03). Whereas D3-positive tumors are less likely to integrate, have later age-of-diagnosis, and a higher rate of somatic mutation. In conclusion, Guatemalan cervical tumors have a high frequency of the very high-risk HPV16 D2 and D3 sub-lineages and cervical cancer patients with these variants of HPV16 differ in histology, age of- diagnosis, integration, and somatic mutation frequency. In summary, related lineages of HPV16 have different features of oncogenicity.

cancer biology↗

bioRxiv2019

Dopamine and risky decision-making in pathological and problem gamblers.

Gambling disorder is a behavioral addiction that is associated with impairments in value-based decision-making such as increased temporal discounting and reduced risk-aversion. Dopamine regulates learning and decision-making by modulating information processing throughout fronto-striatal circuits. Although the role of alterations in dopamine neurotransmission in the etiology of gambling disorder is controversial, preliminary evidence suggests that specifically increasing frontal dopamine levels might improve cognitive functioning in pathological and problem gamblers. We therefore examined whether increasing frontal dopamine levels via the catechol-O-methyltransferase (COMT) inhibitor tolcapone would reduce risky choice in a group of pathological and problem gamblers (n=14) in a repeated-measures counter-balanced placebo-controlled double-blind study. Choice data were fit using hierarchical Bayesian parameter estimation and a modeling scheme that combined a risky choice model with the drift diffusion model to account for both choices and response time distributions. Model comparison revealed that the data were best accounted for by a variant of the drift diffusion model with a non-linear modulation of trial-wise drift rates by value differences, confirming recent findings. Contrary to our hypothesis, risk-taking was slightly increased under tolcapone vs. placebo (Cohens d = -.281). Examination of drug effects on diffusion model parameters revealed an increase in the value-dependency of the drift rate (Cohens d = .932) with a simultaneous reduction in the maximum drift rate (Cohens d = -1.84). These results add to previous work on the potential role of COMT inhibitors in behavioral addictions, and show no consistent beneficial effect of tolcapone on risky choice in gambling disorder. Modeling results add to mounting evidence for the applicability of diffusion models in value-based decision-making. Future work should focus on individual genetic, clinical and cognitive factors that might account for the heterogeneity in the effects of COMT inhibition.

neuroscience↗

bioRxiv2019

Metabolic dysregulation of the lysophospholipid/autotaxin axis in the chromosome 9p21 gene SNP rs10757274

AimsCommon chromosome 9p21 SNPs increase coronary heart disease (CHD) risk, independent of \"traditional lipid risk factors\". However, lipids comprise large numbers of structurally-related molecules not measured in traditional risk measurements, and many have inflammatory bioactivities. Here we applied lipidomic and genomic approaches to three model systems, to characterize lipid metabolic changes in common Chr9p21 SNPs which confer [~]30% elevated CHD risk associated with altered expression of ANRIL, a long ncRNA.\n\nMethods and ResultsUntargeted and targeted lipidomics was applied to plasma samples from Northwick Park Heart Study II (NPHSII) homozygotes for AA or GG in rs10757274. Elevated risk GG correlated with reduced lysophosphospholipids (lysoPLs), lysophosphatidic acids (lysoPA) and autotaxin (ATX). Five other risk SNPs did not show this phenotype. Correlation and network analysis showed that lysoPL-lysoPA interconversion was uncoupled from ATX in GG, indicating metabolic dysregulation. To identify candidate genes, transcriptomic data from shRNA downregulation of ANRIL in HEK293 cells was mined. Significantly-altered expression of several lysoPL/lysoPA metabolising enzymes was found (MBOAT2, PLA2G4C, LPCAT2, ACSL6, PNPLA2, PLBD1, PLPP1, PLPP2 and PLPPR2). Next, vascular smooth muscle cells differentiated from iPSCs of individuals homozygous for Chr9p21 risk SNPs were examined. Here, the presence of risk alleles was associated with altered expression of several lysoPL/lysoPA enzymes. Importantly, for several, deletion of the risk locus fully or partially reversed their expression to non-risk haplotype levels: ACSL3, DGKA, PLA2G2A, LPCAT2, LPL, PLA2G3, PNPLA3, PLA2G12A LIPC, LCAT, PLA2G6, ACSL1, MBOAT2.\n\nConclusionA Chr9p21 risk SNP associates with complex alterations in immune-bioactive phospholipids and their enzymatic metabolism. Lipid metabolites and genomic pathways associated with CHD pathogenesis in Chr9p21 and ANRIL-associated disease are demonstrated.\n\nOne sentence summaryInflammatory phospholipid metabolism defines a cardiovascular disease SNP

physiology↗

bioRxiv2019

The Autoimmune Susceptibility Gene, PTPN2, Restricts Expansion of a Novel Mouse Adherent-Invasive E. coli

Inflammatory bowel diseases (IBD) involve genetic and environmental factors that play major roles in disease pathogenesis. Loss-of-function single-nucleotide polymorphisms (SNPs) in the protein tyrosine phosphatase non-receptor type 2 (PTPN2) gene increase the risk of IBD and are associated with altered microbiome population dynamics in IBD. Moreover, expansion of intestinal pathobionts, such as adherent-invasive E. coli (AIEC), is strongly implicated in the pathogenesis of IBD as AIEC increases pro-inflammatory cytokine production and alters tight junction protein regulation suggesting a potential mechanism of pathogen-induced barrier dysfunction and inflammation. The aim of this study was to identify if PTPN2 deficiency disturbs the composition of the intestinal microbiome to promote expansion of specific bacteria with pathogenic properties. In mice constitutively lacking Ptpn2 we identified increased abundance of a novel adherent-invasive E. coli (AIEC) that showed similar adherence and invasion of intestinal epithelial cells, but greater survival in macrophages to the IBD associated AIEC, LF82. Furthermore, we confirmed this novel mouse AIEC (mAIEC) caused disease when administered to germ-free and mice lacking segmented-filamentous bacteria (SFB). Moreover, mAIEC infection increased severity of and prevented recovery from dextran-sodium sulfate (DSS)-induced colitis. mAIEC genome sequence analysis showed >90% similarity to LF82. Interestingly, mAIEC contained distinct attachment genes not found in LF82 thereby also demonstrating the novelty of this AIEC. We show here for the first time that an IBD susceptibility gene, PTPN2, plays a key role in modulating the gut microbiome to protect against a novel pathobiont. This study generates new insights into gene-environment-microbiome interactions in IBD.

microbiology↗

bioRxiv2019

Diverse cell junctions with unique molecular composition in tissues of a sponge (Porifera)

The integrity and organization of animal tissues depends upon specialized protein complexes that mediate adhesion between cells with each other (cadherin-based adherens junctions), and with the extracellular matrix (integrin-based focal adhesions). Reconstructing how and when these cell junctions evolved is central to understanding early tissue evolution in animals. We examined focal adhesion protein homologs in tissues of the freshwater sponge, Ephydatia muelleri (phylum Porifera). We found that sponge homologs of focal adhesion proteins co-precipitate as a complex and localize to cell junctions in sponge tissues. These data support that the adhesion roles of these proteins evolved early, prior to the divergence of sponges and other animals. However, in contrast to the spatially partitioned distribution of cell junctions in epithelia of other animals, focal adhesion proteins were found to be co-distributed with the adherens junction protein Em{beta}-catenin in sponge tissues; both at certain cell-cell and cell-extracellular matrix (ECM) adhesions. Sponge adhesion structures were found to be unique in other ways, too. The basopinacoderm (substrate-attachment epithelium) lacks typical polarity in that cell-ECM adhesions form on both basal and apical surfaces, and compositionally unique cell junctions form at the interface between cells with spicules (siliceous skeletal elements) and between cells and environmental bacteria. These results clarify the diversity, distribution and molecular composition of cell junctions in tissues of E. muelleri, but raise new questions about their function and homology with cell junctions in other animals.

evolutionary biology↗

bioRxiv2019

11β-Hydroxysteroid Dehydrogenase Type 1 inhibition in Idiopathic Intracranial Hypertension: a double-blind randomized controlled trial.

Treatment options for idiopathic intracranial hypertension are limited. The enzyme 11{beta}-hydroxysteroid dehydrogenase type 1 has been implicated in regulating cerebrospinal fluid secretion, and its activity is associated with alterations in intracranial pressure in idiopathic intracranial hypertension. We assessed therapeutic efficacy, safety and tolerability, and investigate indicators of in vivo efficacy of the 11{beta}-hydroxysteroid dehydrogenase type 1 inhibitor AZD4017 compared to placebo in idiopathic intracranial hypertension. A multicenter, UK, 16-week phase II randomized, double-blind, placebo-controlled trial of 12-weeks treatment with AZD4017 or placebo was conducted. Women aged 18 to 55 years with active idiopathic intracranial hypertension (>25cmH2O lumbar puncture opening pressure and active papilledema) were included. Participants received 400mg twice daily of oral AZD4017 compared to matching placebo over 12-weeks. The outcome measures were initial efficacy, safety and tolerability. The primary clinical outcome was lumbar puncture opening pressure at 12 weeks analysed by intention-to-treat. Secondary clinical outcomes were symptoms, visual function, papilledema, headache and anthropological measures. In vivo efficacy was evaluated in the central nervous system and systemically. 31 subjects (mean age 31.2 (SD=6.9) years and BMI 39.2 (SD=12.6) kg/m2) were randomized to AZD4017 (n=17) or placebo (n=14). At 12 weeks, lumbar puncture pressure was lower in the AZD4017 group (29.7 cmH2O) compared with placebo (31.3 cmH2O), but the difference between groups was not statistically significant (mean difference: -2.8, 95% confidence interval: -7.1-1.5; p=0.2). An exploratory analysis assessing mean change in lumbar puncture pressure within each group found a significant decrease in the AZD4017 group (mean change: -4.3 cmH2O (SD=5.7); p=0.009) but not in the placebo group (mean change: -0.3 cmH2O (SD=5.9); p=0.8). AZD4017 was safe, with no withdrawals related to adverse effects. Nine transient drug-related adverse events were reported. One serious adverse event occurred in the placebo group (deterioration requiring shunt surgery). In vivo biomarkers of 11{beta}-hydroxysteroid dehydrogenase type 1 activity (urinary glucocorticoid metabolites, hepatic prednisolone generation and CSF cortisone to cortisol ratios) demonstrated significant enzyme inhibition. This is the first phase 2 randomized controlled trial in idiopathic intracranial hypertension evaluating a novel therapeutic target. AZD4017 was safe, well-tolerated and inhibited 11{beta}-hydroxysteroid dehydrogenase type 1 activity in vivo. Possible clinical benefits were noted in this small cohort. A longer, larger study would now be of interest.

clinical trials↗

bioRxiv2019

HCMV miR-US22 down-regulation of EGR-1 regulates CD34+ hematopoietic progenitor cell proliferation and viral reactivation

Reactivation of latent Human Cytomegalovirus (HCMV) in CD34+ hematopoietic progenitor cells (HPCs) is closely linked to hematopoiesis. Viral latency requires maintenance of the progenitor cell quiescence, while reactivation initiates following mobilization of HPCs to the periphery and differentiation into CD14+ macrophages. Early growth response gene 1 (EGR-1) is a transcription factor activated by Epidermal growth factor receptor (EGFR) signaling that is essential for the maintenance of CD34+ HPC self-renewal in the bone marrow niche. Down-regulation of EGR-1 results in mobilization and differentiation of CD34+ HPC from the bone marrow to the periphery. In the current study we demonstrate that the transcription factor EGR-1 is directly targeted for down-regulation by HCMV miR-US22 that results in decreased proliferation of CD34+ HPCs and a decrease in total hematopoietic colony formation. We also show that an HCMV miR-US22 mutant fails to reactivate in CD34+ HPCs, indicating that expression of EGR-1 inhibits viral reactivation during latency. Since EGR-1 promotes CD34+ HPC self-renewal in the bone marrow niche, HCMV miR-US22 down-regulation of EGR-1 is a necessary step to block HPC self-renewal and proliferation to induce a cellular differentiation pathway necessary to promote reactivation of virus.\n\nAuthor summaryHuman cytomegalovirus (HCMV) is a widespread herpesvirus that persists in the host and remains a significant cause of morbidity and mortality in solid organ and stem cell transplant patients. HCMV latency is complex, and the molecular mechanisms for establishment, maintenance, and reactivation from latency are poorly understood.\n\nQuiescent stem cells in the bone marrow represent a critical reservoir of latent HCMV, and the mobilization and differentiation of these cells is closely linked to viral reactivation from latency. HCMV encodes small regulatory RNAs, called miRNAs that play important roles in the regulation of viral and cellular gene expression. In this study, we show that HCMV miR-US22 targets Early growth response gene 1 (EGR-1) a host transcription factor that is necessary for stem cell quiescence and self-renewal in the bone marrow. Expression of this miR-US22 down-regulates expression of EGR-1 that reduces CD34+ HPCs proliferation and total hematopoietic colony formation. An HCMV miR-US22 mutant is unable to reactivate from latency suggesting that the ability of the miRNA to disrupt CD34+ HPC renewal in the bone marrow niche to initiate a differentiation pathway is critical for viral reactivation.

microbiology↗