Cell-cell interactome of the hematopoietic niche and its changes in acute myeloid leukemia.

The bone marrow (BM) is a complex microenvironment, coordinating the production of billions of blood cells every day. Despite its essential role and its relevance to hematopoietic diseases, this environment remains poorly characterized. Here we present a high-resolution characterization of the niche in health and acute myeloid leukemia (AML) by establishing a single-cell gene expression database of 339,381 BM cells. We found significant changes in cell type proportions and gene expression in AML, indicating that the entire niche is disrupted. We then predicted interactions between hematopoietic stem and progenitor cells (HSPCs) and other BM cell types, revealing a remarkable expansion of predicted interactions in AML that promote HSPC-cell adhesion, immunosuppression, and cytokine signaling. In particular, predicted interactions involving transforming growth factor ß1 (TGFB1) become widespread, and we show that this can drive AML cell quiescence in vitro. Our results highlight potential mechanisms of enhanced AML-HSPC competitiveness and a skewed microenvironment, fostering AML growth.

A genotype-to-phenotype approach suggests under-reporting of single nucleotide variants in nephrocystin-1 (NPHP1) related disease (UK 100,000 Genomes Project).

Autosomal recessive whole gene deletions of nephrocystin-1 (NPHP1) result in abnormal structure and function of the primary cilia. These deletions can result in a tubulointerstitial kidney disease known as nephronophthisis and retinal (Senior-Løken syndrome) and neurological (Joubert syndrome) diseases. Nephronophthisis is a common cause of end-stage kidney disease (ESKD) in children and up to 1% of adult onset ESKD. Single nucleotide variants (SNVs) and small insertions and deletions (Indels) have been less well characterised. We used a gene pathogenicity scoring system (GenePy) and a genotype-to-phenotype approach on individuals recruited to the UK Genomics England (GEL) 100,000 Genomes Project (100kGP) (n = 78,050). This approach identified all participants with NPHP1-related diseases reported by NHS Genomics Medical Centres and an additional eight participants. Extreme NPHP1 gene scores, often underpinned by clear recessive inheritance, were observed in patients from diverse recruitment categories, including cancer, suggesting the possibility of a more widespread disease than previously appreciated. In total, ten participants had homozygous CNV deletions with eight homozygous or compound heterozygous with SNVs. Our data also reveals strong in-silico evidence that approximately 44% of NPHP1 related disease may be due to SNVs with AlphaFold structural modelling evidence for a significant impact on protein structure. This study suggests historical under-reporting of SNVS in NPHP1 related diseases compared with CNVs.

Supervised Machine Learning Classifies Inflammatory Bowel Disease Patients by Subtype Using Whole Exome Sequencing Data.

BACKGROUND: Inflammatory bowel disease [IBD] is a chronic inflammatory disorder with two main subtypes: Crohn's disease [CD] and ulcerative colitis [UC]. Prompt subtype diagnosis enables the correct treatment to be administered. Using genomic data, we aimed to assess machine learning [ML] to classify patients according to IBD subtype. METHODS: Whole exome sequencing [WES] from paediatric/adult IBD patients was processed using an in-house bioinformatics pipeline. These data were condensed into the per-gene, per-individual genomic burden score, GenePy. Data were split into training and testing datasets [80/20]. Feature selection with a linear support vector classifier, and hyperparameter tuning with Bayesian Optimisation, were performed [training data]. The supervised ML method random forest was utilised to classify patients as CD or UC, using three panels: 1] all available genes; 2] autoimmune genes; 3] 'IBD' genes. ML results were assessed using area under the receiver operating characteristics curve [AUROC], sensitivity, and specificity on the testing dataset. RESULTS: A total of 906 patients were included in analysis [600 CD, 306 UC]. Training data included 488 patients, balanced according to the minority class of UC. The autoimmune gene panel generated the best performing ML model [AUROC = 0.68], outperforming an IBD gene panel [AUROC = 0.61]. NOD2 was the top gene for discriminating CD and UC, regardless of the gene panel used. Lack of variation in genes with high GenePy scores in CD patients was the best classifier of a diagnosis of UC. DISCUSSION: We demonstrate promising classification of patients by subtype using random forest and WES data. Focusing on specific subgroups of patients, with larger datasets, may result in better classification.

The Pediatric Crohn Disease Morbidity Index (PCD-MI): Development of a Tool to Assess Long-Term Disease Burden Using a Data-Driven Approach.

BACKGROUND/OBJECTIVE: Heterogeneity and chronicity of Crohn disease (CD) make prediction of outcomes difficult. To date, no longitudinal measure can quantify burden over a patient's disease course, preventing assessment and integration into predictive modeling. Here, we aimed to demonstrate the feasibility of constructing a data driven, longitudinal disease burden score. METHODS: Literature was reviewed for tools used in assessment of CD activity. Themes were identified to construct a pediatric CD morbidity index (PCD-MI). Scores were assigned to variables. Data were extracted automatically from the electronic patient records at Southampton Children's Hospital, diagnosed from 2012 to 2019 (inclusive). PCD-MI scores were calculated, adjusted for duration of follow up and assessed for variation (ANOVA) and distribution (Kolmogorov-Smirnov). RESULTS: Nineteen clinical/biological features across five themes were included in the PCD-MI including blood/fecal/radiological/endoscopic results, medication usage, surgery, growth parameters, and extraintestinal manifestations. Maximal score was 100 after accounting for follow-up duration. PCD-MI was assessed in 66 patients, mean age 12.5 years. Following quality filtering, 9528 blood/fecal test results and 1309 growth measures were included. Mean PCD-MI score was 14.95 (range 2.2-32.5); data were normally distributed ( P = 0.2) with 25% of patients having a PCD-MI < 10. There was no difference in the mean PCD-MI when split by year of diagnosis, F -statistic 1.625, P = 0.147. CONCLUSIONS: PCD-MI is a calculatable measure for a cohort of patients diagnosed over an 8-year period, integrating a wide-range of data with potential to determine high or low disease burden. Future iterations of the PCD-MI require refinement of included features, optimized scores, and validation on external cohorts.

Prediction of Crohn's Disease Stricturing Phenotype Using a NOD2-derived Genomic Biomarker.

BACKGROUND: Crohn's disease (CD) is highly heterogenous and may be complicated by stricturing behavior. Personalized prediction of stricturing will inform management. We aimed to create a stricturing risk stratification model using genomic/clinical data. METHODS: Exome sequencing was performed on CD patients, and phenotype data retrieved. Biallelic variants in NOD2 were identified. NOD2 was converted into a per-patient deleteriousness metric ("GenePy"). Using training data, patients were stratified into risk groups for fibrotic stricturing using NOD2. Findings were validated in a testing data set. Models were modified to include disease location at diagnosis. Cox proportional hazards assessed performance. RESULTS: Six hundred forty-five patients were included (373 children and 272 adults); 48 patients fulfilled criteria for monogenic NOD2-related disease (7.4%), 24 of whom had strictures. NOD2 GenePy scores stratified patients in training data into 2 risk groups. Within testing data, 30 of 161 patients (18.6%) were classified as high-risk based on the NOD2 biomarker, with stricturing in 17 of 30 (56.7%). In the low-risk group, 28 of 131 (21.4%) had stricturing behavior. Cox proportional hazards using the NOD2 risk groups demonstrated a hazard ratio (HR) of 2.092 (P = 2.4 × 10-5), between risk groups. Limiting analysis to patients diagnosed aged < 18-years improved performance (HR-3.164, P = 1 × 10-6). Models were modified to include disease location, such as terminal ileal (TI) disease or not. Inclusion of NOD2 risk groups added significant additional utility to prediction models. High-risk group pediatric patients presenting with TI disease had a HR of 4.89 (P = 2.3 × 10-5) compared with the low-risk group patients without TI disease. CONCLUSIONS: A NOD2 genomic biomarker predicts stricturing risk, with prognostic power improved in pediatric-onset CD. Implementation into a clinical setting can help personalize management.

NOD2 is a well-established risk gene for development of Crohn's disease and stricturing behavior. Here we demonstrate NOD2 can be utilized as a genomic biomarker, stratifying patients into 2 stricturing risk groups. Further refinement using disease location at diagnosis improved risk stratification.

A novel variant in GATM causes idiopathic renal Fanconi syndrome and predicts progression to end-stage kidney disease.

Renal Fanconi syndrome (RFS) is a generalised disorder of the proximal convoluted tubule. Many genes have been associated with RFS including those that cause systemic disorders such as cystinosis, as well as isolated RFS. We discuss the case of a 10-year-old female who presented with leg pain and raised creatinine on a screening blood test. Her mother has RFS and required a kidney transplant in her thirties. Further investigations confirmed RFS in the daughter. Exome sequencing was performed on the affected mother, child, and unaffected father. We identified a novel variant in GATM; c.965G>C p.(Arg322Pro) segregating dominantly in the mother and daughter. We validated our finding with molecular dynamics simulations and demonstrated a dynamic signature that differentiates our variant and two previously identified pathogenic variants in GATM from wildtype. Genetic testing has uncovered a novel pathogenic variant that predicts progression to end stage kidney failure and has important implications for family planning and cascade testing. We recommend that GATM is screened for in children presenting with RFS, in addition to adults, particularly with kidney failure, who may have had previous negative gene testing.

NOD2 in Crohn's Disease-Unfinished Business.

Studies of Crohn's disease have consistently implicated NOD2 as the most important gene in disease pathogenesis since first being identified in 2001. Thereafter, genome-wide association, next-generation sequencing and functional analyses have all confirmed a key role for NOD2, but despite this, NOD2 also has significant unresolved complexity. More recent studies have reinvigorated an early hypothesis that NOD2 may be a single-gene cause of disease, and the distinct ileal stricturing phenotype seen with NOD2-related disease presents an opportunity for personalized diagnosis, disease prediction and targeted therapy. The genomics of NOD2 has much that remains unknown, including the role of rare variation, phasing of variants across the haplotype block and the role of variation in the NOD2-regulatory regions. Here, we discuss the evidence and the unmet needs of NOD2 research, based on recently published evidence, and suggest methods that may meet these requirements.

Evidence of a genetically driven metabolomic signature in actively inflamed Crohn's disease.

Crohn's disease (CD) is characterised by chronic inflammation. We aimed to identify a relationship between plasma inflammatory metabolomic signature and genomic data in CD using blood plasma metabolic profiles. Proton NMR spectroscopy were achieved for 228 paediatric CD patients. Regression (OPLS) modelling and machine learning (ML) approaches were independently applied to establish the metabolic inflammatory signature, which was correlated against gene-level pathogenicity scores generated for all patients and functional enrichment was analysed. OPLS modelling of metabolomic spectra from unfasted patients revealed distinctive shifts in plasma metabolites corresponding to regions of the spectrum assigned to N-acetyl glycoprotein, glycerol and phenylalanine that were highly correlated (R2 = 0.62) with C-reactive protein levels. The same metabolomic signature was independently identified using ML to predict patient inflammation status. Correlation of the individual peaks comprising this metabolomic signature of inflammation with pathogenic burden across 15,854 unselected genes identified significant enrichment for genes functioning within 'intrinsic component of membrane' (p = 0.003) and 'inflammatory bowel disease (IBD)' (p = 0.003). The seven genes contributing IBD enrichment are critical regulators of pro-inflammatory signaling. Overall, a metabolomic signature of inflammation can be detected from blood plasma in CD. This signal is correlated with pathogenic mutation in pro-inflammatory immune response genes.

A Systematic Review of Artificial Intelligence and Machine Learning Applications to Inflammatory Bowel Disease, with Practical Guidelines for Interpretation.

BACKGROUND: Inflammatory bowel disease (IBD) is a gastrointestinal chronic disease with an unpredictable disease course. Computational methods such as machine learning (ML) have the potential to stratify IBD patients for the provision of individualized care. The use of ML methods for IBD was surveyed, with an additional focus on how the field has changed over time. METHODS: On May 6, 2021, a systematic review was conducted through a search of MEDLINE and Embase databases, with the search structure ("machine learning" OR "artificial intelligence") AND ("Crohn* Disease" OR "Ulcerative Colitis" OR "Inflammatory Bowel Disease"). Exclusion criteria included studies not written in English, no human patient data, publication before 2001, studies that were not peer reviewed, nonautoimmune disease comorbidity research, and record types that were not primary research. RESULTS: Seventy-eight (of 409) records met the inclusion criteria. Random forest methods were most prevalent, and there was an increase in neural networks, mainly applied to imaging data sets. The main applications of ML to clinical tasks were diagnosis (18 of 78), disease course (22 of 78), and disease severity (16 of 78). The median sample size was 263. Clinical and microbiome-related data sets were most popular. Five percent of studies used an external data set after training and testing for additional model validation. DISCUSSION: Availability of longitudinal and deep phenotyping data could lead to better modeling. Machine learning pipelines that consider imbalanced data and that feature selection only on training data will generate more generalizable models. Machine learning models are increasingly being applied to more complex clinical tasks for specific phenotypes, indicating progress towards personalized medicine for IBD.

Exome sequencing identifies a disease variant of the mitochondrial ATP-Mg/Pi carrier SLC25A25 in two families with kidney stones.

BACKGROUND: Calcium kidney stones are common and recurrences are often not preventable by available empiric remedies. Their etiology is multifactorial and polygenic, and an increasing number of genes are implicated. Their identification will enable improved management. METHODS: DNA from three stone-formers in a Southampton family (UK) and two from an Italian family were analyzed independently by whole exome sequencing and selected variants were genotyped across all available members of both pedigrees. A disease variant of SLC25A25 (OMIM 608745), encoding the mitochondrial ATP-Mg/Pi carrier 3 (APC3) was identified, and analyzed structurally and functionally with respect to its calcium-regulated transport activity. RESULTS: All five patients had a heterozygous dominant SLC25A25 variant (rs140777921; GRCh37.p13: chr 9 130868670 G>C; p.Gln349His; Reference Sequence NM_001006641.3). Non-stone formers also carried the variant indicating incomplete penetrance. Modeling suggests that the variant lacks a conserved polar interaction, which may cause structural instability. Calcium-regulated ATP transport was reduced to ~20% of the wild type, showing a large reduction in function. CONCLUSION: The transporter is important in regulating mitochondrial ATP production. This rare variant may increase urine lithogenicity through impaired provision of ATP for solute transport processes in the kidney, and/or for purinergic signaling. Variants found in other genes may compound this abnormality.

Expression profile of the matricellular protein periostin in paediatric inflammatory bowel disease.

The precise role of periostin, an extra-cellular matrix protein, in inflammatory bowel disease (IBD) is unclear. Here, we investigated periostin in paediatric IBD including its relationship with disease activity, clinical outcomes, genomic variation and expression in the colonic tissue. Plasma periostin was analysed using ELISA in 144 paediatric patients and 38 controls. Plasma levels were assessed against validated disease activity indices in IBD and clinical outcomes. An immuno-fluorescence for periostin and detailed isoform-expression analysis in the colonic tissue was performed in 23 individuals. We integrated a whole-gene based burden metric 'GenePy' to assess the impact of variation in POSTN and 23 other genes functionally connected to periostin. We found that plasma periostin levels were significantly increased during remission compared to active Crohn's disease. The immuno-fluorescence analysis demonstrated enhanced peri-cryptal ring patterns in patients compared to controls, present throughout inflamed, as well as macroscopically non-inflamed colonic tissue. Interestingly, the pattern of isoforms remained unchanged during bowel inflammation compared to healthy controls. In addition to its role during the inflammatory processes in IBD, periostin may have an additional prominent role in mucosal repair. Additional studies will be necessary to understand its role in the pathogenesis, repair and fibrosis in IBD.

Growth failure is rare in a contemporary cohort of paediatric inflammatory bowel disease patients.

AIM: We assessed growth in a paediatric inflammatory bowel disease (PIBD) cohort. METHODS: Paediatric inflammatory bowel disease patients were eligible if they were diagnosed at Southampton Children's Hospital from 2011 to 2018. Weight and height standard deviation scores (SDS) were retrieved. Mean SDS values, SDS change and anti-TNF status were analysed at diagnosis and during follow-up. RESULTS: Four hundred and ninety patients were included, 313 with Crohn's disease (CD). CD patients presented with mean height SDS -0.13, -0.1 at 1-year, -0.11 at 2-years and -0.03 at 5 years, reflecting preserved linear growth. There was no significant height-SDS change from diagnosis to 5-year follow-up, +0.12, 95%-CI: 0.48 to -0.24. Mean weight-SDS at diagnosis was -0.39, driven by CD patients (-0.65). Mean weight-SDS approached 0 after 1 year and remained at the 50th centile throughout follow-up. Growth in ulcerative colitis was maintained. In multivariable regression males had worse height growth from diagnosis to transition (P = .036). Anti-TNF treatment (P = .013) and surgical resection (P = .005) were also associated with poorer linear growth. Patients treated with anti-TNF therapy had lower height-SDS compared to those never treated with anti-TNF at 1 year (-0.2 vs -0.01, P = .22), 2-years (-0.27 vs -0.01, P = .07) and 5 years (-0.21 vs 0.25, P = .051). CONCLUSION: Height was generally maintained in Crohn's disease, and impaired linear growth was rare in this cohort.

Ileal Transcriptomic Analysis in Paediatric Crohn's Disease Reveals IL17- and NOD-signalling Expression Signatures in Treatment-naïve Patients and Identifies Epithelial Cells Driving Differentially Expressed Genes.

BACKGROUND AND AIMS: Crohn's disease [CD] arises through host-environment interaction. Abnormal gene expression results from disturbed pathway activation or response to bacteria. We aimed to determine activated pathways and driving cell types in paediatric CD. METHODS: We employed contemporary targeted autoimmune RNA sequencing, in parallel to single-cell sequencing, to ileal tissue derived from paediatric CD and controls. Weighted gene co-expression network analysis [WGCNA] was performed and differentially expressed genes [DEGs] were determined. We integrated clinical data to determine co-expression modules associated with outcomes. RESULTS: In all, 27 treatment-naive CD [TN-CD], 26 established CD patients and 17 controls were included. WGCNA revealed a 31-gene signature characterising TN-CD patients, but not established CD, nor controls. The CSF3R gene is a hub within this module and is key in neutrophil expansion and differentiation. Antimicrobial genes, including S100A12 and the calprotectin subunit S100A9, were significantly upregulated in TN CD compared with controls [p = 2.61 x 10-15 and p = 9.13 x 10-14, respectively] and established CD [both p = 0.0055]. Gene-enrichment analysis confirmed upregulation of the IL17-, NOD- and Oncostatin-M-signalling pathways in TN-CD patients, identified in both WGCNA and DEG analyses. An upregulated gene signature was enriched for transcripts promoting Th17-cell differentiation and correlated with prolonged time to relapse [correlation-coefficient-0.36, p = 0.07]. Single-cell sequencing of TN-CD patients identified specialised epithelial cells driving differential expression of S100A9. Cell groups, determined by single-cell gene expression, demonstrated enrichment of IL17-signalling in monocytes and epithelial cells. CONCLUSIONS: Ileal tissue from treatment-naïve paediatric patients is significantly upregulated for genes driving IL17-, NOD- and Oncostatin-M-signalling. This signal is driven by a distinct subset of epithelial cells expressing antimicrobial gene transcripts.

Genetic Sequencing of Pediatric Patients Identifies Mutations in Monogenic Inflammatory Bowel Disease Genes that Translate to Distinct Clinical Phenotypes.

OBJECTIVES: Monogenic inflammatory bowel disease (IBD) comprises rare Mendelian causes of gut inflammation, often presenting in infants with severe and atypical disease. This study aimed to identify clinically relevant variants within 68 monogenic IBD genes in an unselected pediatric IBD cohort. METHODS: Whole exome sequencing was performed on patients with pediatric-onset disease. Variants fulfilling the American College of Medical Genetics criteria as "pathogenic" or "likely pathogenic" were assessed against phenotype at diagnosis and follow-up. Individual patient variants were assessed and processed to generate a per-gene, per-individual, deleteriousness score. RESULTS: Four hundred one patients were included, and the median age of disease-onset was 11.92 years. In total, 11.5% of patients harbored a monogenic variant. TRIM22-related disease was implicated in 5 patients. A pathogenic mutation in the Wiskott-Aldrich syndrome (WAS) gene was confirmed in 2 male children with severe pancolonic inflammation and primary sclerosing cholangitis. In total, 7.3% of patients with Crohn's disease had apparent autosomal recessive, monogenic NOD2-related disease. Compared with non-NOD2 Crohn's disease, these patients had a marked stricturing phenotype (odds ratio 11.52, significant after correction for disease location) and had undergone significantly more intestinal resections (odds ratio 10.75). Variants in ADA, FERMT1, and LRBA did not meet the criteria for monogenic disease in any patients; however, case-control analysis of mutation burden significantly implicated these genes in disease etiology. DISCUSSION: Routine whole exome sequencing in pediatric patients with IBD results in a precise molecular diagnosis for a subset of patients with IBD, providing the opportunity to personalize therapy. NOD2 status informs risk of stricturing disease requiring surgery, allowing clinicians to direct prognosis and intervention.

Analysis and Hierarchical Clustering of Blood Results Before Diagnosis in Pediatric Inflammatory Bowel Disease.

BACKGROUND: Pediatric inflammatory bowel disease (PIBD) is associated with a diagnostic delay. Blood tests are a routine part of the work-up in children with chronic abdominal symptoms (pain, diarrhea). Normal blood tests cannot exclude PIBD. We analyzed blood results at diagnosis over a 5-year period. METHODS: Patients diagnosed from 2013 to 2017 were identified from the Southampton-PIBD database. Results were obtained up to 100 days before diagnostic endoscopy. Erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), albumin, hemoglobin, platelets, packed cell volume (PCV), white cell count (WCC), and alanine transferase (ALT) were analyzed. Hierarchical clustering was applied to normalized results. RESULTS: Two hundred fifty-six patients were included (Crohn's disease [CD], 151; ulcerative colitis [UC], 95; IBD-unclassified, 10; median age, 13.48 years; 36.7% female). Hierarchical clustering of patients revealed novel groupings enriched for CD and UC, characterized by specific patterns of results. In PIBD, 9% presented with all normal blood tests, 21.9% with normal CRP and ESR. Abnormal results were seen in all tests (ESR, 56.4% of patients; CRP, 53.4%; albumin, 28%; hemoglobin, 61.9%; platelets, 55.6%; PCV, 64.6%; WCC, 22.7%; and ALT, 7.2%). Normal inflammatory markers were more common in UC compared with CD (UC, 34%; CD, 15.8%; P = 0.0035). UC (14.4% normal) presented with all normal results more frequently than CD (CD, 5.3%; P = 0.02). CRP, ESR, and platelets were significantly higher in CD compared with UC. Albumin and hemoglobin were significantly lower. CONCLUSIONS: Most cases of PIBD present with >1 abnormal blood result, although 1/11 patients presents with normal blood tests and 1/5 present with normal inflammatory markers. Hierarchical clustering offers the potential to produce novel groupings to inform disease categorization and best management.

Review article: the genetics of the human leucocyte antigen region in inflammatory bowel disease.

BACKGROUND: The human leucocyte antigen (HLA) complex, located at chromosome 6p21.3 is a highly polymorphic region containing the classical class I and II HLA genes. The region is highly associated with inflammatory bowel disease (IBD), largely through genome-wide association studies (GWAS). AIMS: To review the role of HLA in immune function, summarise data on risk/protective HLA genotypes for IBD, discuss the role of HLA in IBD pathogenesis, treatment and examine limitations that might be addressed by future research. METHODS: An organised search strategy was used to collate articles describing HLA genes in IBD, including Crohn's disease and ulcerative colitis. RESULTS: All classical HLA genes with variation (including HLA-A, B, C, DRB1, DQA1, DQB1, DPA1 and DPB1) harbour IBD-associated genotypes. The most implicated gene is HLA-DRB1, with HLA-DRB1*03:01 the most associated risk allele in both Crohn's disease and ulcerative colitis. Elucidating precise disease associations is challenging due to high linkage disequilibrium between HLA genotypes. The mechanisms by which risk alleles cause disease are multifactorial, with the best evidence indicating structural and electrostatic alteration impacting antigen binding and downstream signalling. Adverse medication events have been associated with HLA genotypes including with thiopurines (pancreatitis) and anti-TNF agents (antibody formation). CONCLUSIONS: The HLA complex is associated with multiple risk/protective alleles for IBD. Future research utilising long-read technology, ascertainment of zygosity and integration in disease modelling will improve the functional understanding and clinical translation of genetic findings.

Systematic Review of Somatic Mutations in Splenic Marginal Zone Lymphoma.

The aims of this systematic review are to refine the catalogue of somatic variants in splenic marginal zone lymphoma (SMZL) and to provide a well-annotated, manually curated database of high-confidence somatic mutations to facilitate variant interpretation for further biological studies and future clinical implementation. Two independent reviewers systematically searched PubMed and Ovid in January 2019 and included studies that sequenced SMZL cases with confirmed diagnosis. The database included fourteen studies, comprising 2817 variants in over 1000 genes from 475 cases. We confirmed the high prevalence of NOTCH2, KLF2 and TP53 mutations and analysis of targeted genes further implicated TNFAIP3, KMT2D, and TRAF3 as recurrent targets of somatic mutation based on their high incidence across studies. The major limitations we encountered were the low number of patients with whole-genome, unbiased analysis and the relative sensitivities of differing sequencing approaches. Overall, we showed that there is little concordance between whole exome sequencing studies of SMZL. We strongly support the continuing unbiased analysis of the SMZL genome for mutations in all protein-coding genes and provide a valuable database resource to facilitate this endeavour that will ultimately improve our understanding of SMZL pathobiology.

Increased prevalence of anti-TNF therapy in paediatric inflammatory bowel disease is associated with a decline in surgical resections during childhood.

BACKGROUND: Anti-tumour necrosis factor-α (anti-TNF) therapy use has risen in paediatric-onset inflammatory bowel disease (PIBD). Whether this has translated into preventing/delaying childhood surgery is uncertain. The Wessex PIBD cohort was analysed for trends in anti-TNF-therapy and surgery. AIM: To assess patients diagnosed with PIBD within Wessex from 1997 to 2017. The prevalence of anti-TNF-therapy and yearly surgery rates (resection and perianal) during childhood (<18 years) were analysed (Pearson's correlation, multivariate regression, Fisher's exact). RESULTS: Eight-hundred-and-twenty-five children were included (498 Crohn's disease, 272 ulcerative colitis, 55 IBD-unclassified), mean age at diagnosis 13.6 years (1.6-17.6), 39.6% female. The prevalence of anti-TNF-treated patients increased from 5.1% to 27.1% (2007-2017), P = 0.0001. Surgical resection-rate fell (7.1%-1.5%, P = 0.001), driven by a decrease in Crohn's disease resections (8.9%-2.3%, P = 0.001). Perianal surgery and ulcerative colitis resection-rates were unchanged. Time from diagnosis to resection increased (1.6-2.8 years, P = 0.028) but mean age at resection was unchanged. Patients undergoing resections during childhood were diagnosed at a younger age in the most recent 5 years (2007-2011 = 13.1 years, 2013-2017 = 11.9 years, P = 0.014). Resection-rate in anti-TNF-therapy treated (16.1%) or untreated (12.2%) was no different (P = 0.25). Patients started on anti-TNF-therapy <3 years post-diagnosis (11.6%) vs later (28.6%) had a reduction in resections, P = 0.047. Anti-TNF-therapy prevalence was the only significant predictor of resection-rate using multivariate regression (P = 0.011). CONCLUSIONS: The prevalence of anti-TNF-therapy increased significantly, alongside a decrease in surgical resection-rate. Patients diagnosed at younger ages still underwent surgery during childhood. Anti-TNF-therapy may reduce the need for surgical intervention in childhood, thereby influencing the natural history of PIBD.

Analysis of Mutation and Loss of Heterozygosity by Whole-Exome Sequencing Yields Insights into Pseudomyxoma Peritonei.

Pseudomyxoma peritonei (PMP) is a clinical syndrome characterized by gross mucinous ascites originating from a disseminated intraperitoneal neoplasm. Although typically confined to the abdomen, mortality is high if untreated. Biomarkers, including genetic mutation profiles, may aid treatment selection and decision making. We applied whole-exome sequencing to five patients diagnosed with low-grade appendiceal mucinous neoplasms, using paired tumor and germline samples identify biomarkers. Multiple bioinformatic approaches were applied to these data to assess both somatic mutation profiles and loss of heterozygosity events. Mutation profiles of the tumors were consistent with deamination of methylcytosine being the prevailing mechanism. Pathogenic mutations were identified in both KRAS and GNAS in all samples, and further mutations in genes implicated in PMP, namely FGFR2, APC, SMAD2, and FAT4. No TP53 somatic mutations were identified, matching expectations for low-grade tumors. Four of five samples exhibited clonal loss of heterozygosity; these regions were further examined and found to contain genes harboring pathogenic somatic mutations in some samples. RNF43 was hereby implicated in the pathogenesis of PMP of appendiceal origin, having previously been found to increase sensitivity to Wnt signaling and to have involvement in similar mucinous tumors. In conclusion, we have investigated the mutation profile of PMP of appendiceal origin and provided the first report of RNF43 involvement in its progression.