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Dysregulated immune proteins in plasma in the UK Biobank predict Multiple Myeloma 12 years before clinical diagnosis

KLF5 Is a Key Regulator of IMiD-Induced Neutropenia

Although immunomodulatory drugs (IMiDs, lenalidomide [LEN] & pomalidomide [POM]) have had a huge impact on the therapeutic landscape in multiple myeloma (MM), IMiD-induced neutropenia remains a clinical challenge. Understanding the molecular basis of IMiD-associated myelosuppression could improve the rational design of novel agents which mitigate this important side effect whilst retaining therapeutic efficacy against MM cells. We have previously established an experimental system to carry out single cell multiomic analyses of neutrophil differentiation ex vivo, allowing us to study the impact of exposure to IMiDs, which caused a maturation impairment and decrease in the abundance of differentiating granulocytes1. These findings aligned with previous clinical observations of a myeloid maturation arrest which is associated with IMiD-induced neutropenia in MM patients, with Cereblon (CRBN)-driven IKZF1 degradation previously implicated as the underlying cause. However, restoration of IKZF1 expression only partially rescues the IMiD-associated neutrophil maturation arrest, suggesting that other mechanisms might also contribute. The purpose of the current study was to interrogate our single cell multiomic dataset to characterise the impact of IMiDs on the dynamic genome regulatory landscape during neutrophil development and thereby identify putative novel neo-substrates that might contribute to myelosuppression. Analysis of single cell RNA seq (n=3 donors and 111,109 cells) and multiome datasets (n=3 donors and 47,452 cells) of ex vivo neutrophil differentiation and its perturbation by IMiDs, allowed us to construct a gene regulatory map to identify putative driver genes implicated in the pathobiology of IMiD-mediated neutrophil maturation arrest. Combined gene expression and chromatin accessibility analysis based on the detection of 330,334 peak-to-gene links correlated chromatin architecture to neutrophil terminal differentiation potential. Myeloid progenitors contained relaxed chromatin, whilst IMiD-associated abnormal myeloid precursors states were characterized by premature chromatin compaction (as reflected by a 30% decline in filtered peaks) and priming towards a pro-apoptotic state, despite bearing gene expression profiles largely reminiscent of intact neutrophils and precursors. This finding was corroborated by the reduction of accessible enhancer loci (4.5-14.8%), suggesting a repression of transcriptional activity in the treated populations. To enrich for candidate transcription factors, we selected motifs that were not altered in expression at the transcript level in the granulocyte-monocyte progenitor and metamyelocyte clusters but showed marked IMiD-induced changes in their associated regulons at the transcript level. Furthermore, ATAC-seq footprinting allowed us to infer altered transcription factor binding dynamics in the same cells. As expected, IKZF1 met these criteria alongside SPI1,CEBPA, ZBTB7A, ZBTB7B, KLF5, PPARA, PPARG additional candidate genes. To further prioritise candidate neo-substrates, we intersected our data with an extensive proteomic screening to enrich for CRBN-interacting small molecules. Amongst the genes overlapping in both our multiomic analysis and proteomic screen was KLF5, a known transcription factor involved in the regulation of neutrophil differentiation in mice. Moreover, we also identified downregulation of KLF5 binding partners C/EBPβ and C/EBPδ and the downstream target PPARG alongside a gradual switch from the overarching (during normal granulopoiesis) glycolytic metabolic machinery towards increased fatty acid oxidation. This surrogate pathway has been previously associated with skewed neutrophil effector functions and described in genome-wide association studies (GWAS) focusing on abnormal neutrophil counts. Intracellular FACS analysis confirmed rapid KLF5 degradation upon exposure to IMiDs, confirming that this is a likely IMiD-associated CRBN neo-substrate. Proximity assays, rescue studies and conformation capture assays to dissect the relevant key regulatory elements are underway. Our study has identified KLF5 as a putative novel regulator of IMiD-induced neutropenia which might inform the rational design of new therapeutic agents to mitigate this problematic side effect of widely used drugs in MM. 1.https://doi.org/10.1182/blood-2023-182041

Constructing a Computational Workflow for the Identification of Novel Cellular and Molecular Drivers of Human Granulopoiesis

Neutrophils are crucial immune cells with complex and heterogeneous transcriptional programs. To understand the dynamic changes governing human granulopoiesis, we previously developed an ex vivo myeloid cell differentiation assay using human mobilised peripheral blood CD34+ cells from healthy donors and conducted single cell multiomic analysis. We constructed a multimodal atlas of human granulopoiesis that captured the full spectrum of human myeloid cells spanning from early myeloid progenitors through to neutrophil populations, thus enabling the study the genome regulatory events underlying neutrophil maturation with unprecedented resolution1. Our aim was to design a computational analytical workflow enabling the exploration of the cellular and molecular drivers of normal human granulopoiesis alongside the precise characterisation of the dynamic changes in the genome regulatory landscape dictating myeloid cell commitment and terminal neutrophil differentiation. Our dataset included transcriptomic (scRNA; n=3 donors and 29,300 cells) and simultaneous profiled gene expression and chromatin accessibility (scGEX & scATAC; n=3 donors and 17,219 cells) analyses from mature neutrophils and myeloid precursors. We successfully co-embedded our data with a published reference dataset encompassing neutrophils and myeloid progenitors from heathy donors, confirming the overlap of the transcriptomic signatures. The transition from transcriptome-based to epigenome-based cluster labels improved annotation resolution, enabling precise identification of previously unexplored subtypes within neutrophil and precursor populations. Further investigation of both modalities revealed distinct chromatin conformations, but almost identical transcriptomic signatures, for clusters of band and mature neutrophils approaching the end of their lifespan. The exhausted pro-apoptotic neutrophils displayed dynamically increased chromatin compaction as evidenced by a 42.1% decline in accessible peaks detected, indicating an earlier stage towards programmed cell death commitment compared to robust neutrophils. Next, we used our resource atlas to interrogate the underlying genome regulatory networks and characterise cell-type-specific cis regulatory elements (CREs) involved in human granulopoiesis. We coupled transcription factor (TF) and target gene identification using Scanpy with SCENIC analysis, enabling a comparative study focusing on open chromatin regions. We then linked the pre-identified cell-type-specific activities for 178 differentially present TFs to chromatin accessibility changes using ArchR and performed motif enrichment and peak-to-gene linkage identification using the integrated scGEX dataset. As a result, we were able to underpin the connection between neutrophil terminal differentiation potential and the dynamic changes in chromatin architecture. We analysed the role of our candidate TFs by imputing regulatory single nucleotide polymorphisms (SNPs) from published genome-wide association studies (GWAS) focusing on neutrophil biology and absolute counts. This showed an increased transcriptional activity for our candidate TFs, with an overall enrichment frequency of detected regulatory SNPs, within the metamyelocyte compartment, thus further highlighting the role of this differentiation stage as a regulatory switch towards terminal neutrophil maturation. We used reference chromatin immunoprecipitation sequencing (ChIP-seq) data from mature neutrophils and progenitors focusing on histone modification marks and CTCF sites to further annotate de novo genome-wide cell-type-specific CREs using REgulamentary2. These findings were visualised using Multi-Dimensional Viewer, a comprehensive analytical interpretation tool, generating a publicly available resource atlas benefiting from the seamless interaction with this multiomic dataset. We present a computational analytical workflow that enables large-scale multiomic data mining to study the molecular underpinnings of normal neutrophil development, by precisely characterising the dynamical changes of the regulatory landscape during human granulopoiesis and generating testable hypothesis through regulatory SNPs assessment. 1. https://doi.org/10.1182/blood-2023-182041 2. https://doi.org/10.1101/2024.05.24.595662

Benchmarking transformer-based models for medical record deidentification: A single centre, multi-specialty evaluation

IRAK3 is upregulated in rheumatoid arthritis synovium and delays the onset of experimental arthritis

<jats:p>Tumour necrosis factor (TNF) is a potent inducer of endotoxin tolerance-associated molecules, such as interleukin-1 receptor-associated kinase 3 (IRAK3), and also a therapeutic target in inflammatory autoimmune diseases, as it upregulates the production of inflammatory mediators. The role of IRAK3 was assessed in rheumatoid arthritis (RA), a disease which is amenable to TNF blockade. As a variant of IRAK3 lacks the death domain required for its canonical role, isoform expression was determined in different inflammatory milieu by immunoblotting. RA synovial explant expression of IRAK3 was measured by qPCR. The expression of the larger, “classical” IRAK3 isoform predominated in macrophages treated with various stimuli. The expression of IRAK3 was higher in RA synovium compared to osteoarthritis synovium. Using collagen-induced arthritis, a murine model of RA, the immunomodulatory role of IRAK3 was investigated with wild-type (WT) and IRAK3-deficient mice expressing the MHC-II A<jats:sup>q</jats:sup> allele. Disease progression was significantly accelerated in IRAK3<jats:sup>−/−</jats:sup> mice. In addition, the circulating levels of IL-1β were greater, and there were fewer Tregs both before and after the onset of disease. Inflammatory gene expression was higher in the arthritic paws of IRAK3<jats:sup>−/−</jats:sup> mice. This study demonstrates that IRAK3 deficiency accelerates the progression of arthritis and increases molecular markers of disease severity.”</jats:p>

Primary Care Resource Utilisation and Costs of Fragility Fractures in Postmenopausal Women in the UK Using CPRD Data Mapped to OMOP Common Data Model

Large Scale Propensity Score Matching With Time-Stratification in Older Women With or Without Fractures in the UK

INCIDENCE OF IMMINENT SUBSEQUENT FRACTURE IN POSTMENOPAUSAL WOMEN AS CAPTURED IN PRIMARY CARE COMPARED TO HOSPITAL RECORDS IN UK AND SPAIN

HOSPITAL RESOURCE UTILIZATION AND COSTS OF IMMINENT SUBSEQUENT FRACTURES IN POSTMENOPAUSAL WOMEN: A DISTRIBUTED NETWORK ANALYSIS USING DATA FROM THE UK AND SPAIN MAPPED TO OMOP COMMON DATA MODEL

Feasibility of a Federated Network Analysis Using Real-World Data Mapped to OMOP Common Data Model to Estimate Healthcare Resource Utilisation and Costs of Imminent Subsequent Fracture

Treatment of Systemic Lupus Erythematosus: Analysis of Treatment Patterns in Adult and Paediatric Patients Across Four European Countries

DARWIN EU®-Trends of Prescription Opioid Use in Europe

Ethnicity data resource in population-wide health records: completeness, coverage and granularity of diversity

COLLATERAL EFFECTS OF THE COVID-19 PANDEMIC ON ENDOCRINE TREATMENTS FOR BREAST AND PROSTATE CANCER IN THE UK: IMPLICATIONS FOR BONE HEALTH

The Risks of Immune-oand Inflammatory Related Post-Acute Sequelae of COVID-19: A Network Study in 6 European Countries, the US, and Korea

Natural History of Dermatomyositis and Polymyositis in Adults and Paediatric Populations: A Network Cohort Study Part of DARWIN EU®

Prevalence of Dermatomyositis and Polymyositis in Adults and Paediatric Populations: A Network Cohort Study Part of DARWIN EU®

Incidence, Prevalence, and Survival of Breast Cancer in the United Kingdom From 2000-2021: A Population-Based Cohort Study

PREDICTING LONG TERM CANCER SURVIVAL FOR HEALTH TECHNOLOGY ASSESSMENT: A MULTINATIONAL COHORT STUDY

Discontinuation of Preventative Treatments in Older People with Complex Health Needs: Self-Controlled Case Series Analyses of Preventable Cardiovascular Events