Barbera D. C. van Schaik

High-throughput sequencing of B-cell and T-cell immune receptor repertoires provides unprecedented insight into adaptive immune responses. The data produced are structured by clonal relationships and somatic mutation signatures, and yield extremely rich information in sequence-derived features, including physicochemical properties and compositional patterns. However, integrated analysis across datasets, conditions, and time points remains challenging. Current analytical tools typically focus only on certain features within individual repertoires, without enabling integrated, multivariable comparisons across datasets, conditions, and time points to address their diversity and variability. Here we present AbSolution, a user-friendly and flexible interactive application for comprehensive exploration of immune repertoires and their sequence-based properties. AbSolution enables multiscale analysis of thousands of sequence-derived features across receptor regions, while accounting for V(D)J usage, clonal composition and experimental groupings. We demonstrate its utility by identifying distinct sequence-based profiles associated with dominant (highly abundant) and non-dominant B-cell clones in peripheral blood BCR repertoires from patients with idiopathic inflammatory myopathies, and with antigen-responsive T-cell populations over time in a longitudinal in vitro antigen-stimulation dataset. Through interactive, interlinked visualizations, statistical feature selection and multi-sample comparisons, AbSolution facilitates integrated feature profiling that supports the interpretation of immune selection processes and enables systematic analysis of complex repertoire datasets.

Reproducibility of computational research is often challenging despite established guidelines and best practices. Translating these guidelines into practical applications remains difficult. Here, we present ENCORE, an approach to enhance transparency and reproducibility by guiding researchers in how to structure and document a computational project. ENCORE builds on previous efforts in computational reproducibility and integrates all project components into a standardized file system structure. It utilizes pre-defined files as documentation templates, leverages GitHub for software versioning, and includes an HTML-based navigator. ENCORE is designed to be agnostic to the type of computational project, data, programming language, and ICT infrastructure, and does not rely on specific software tools. We also share our group's experience using ENCORE, highlighting that the most significant challenge to the routine adoption of approaches like ours is the lack of incentives to motivate researchers to dedicate sufficient time and effort to ensure reproducibility.

Although hundreds of genome-wide association studies-implicated loci have been reported for adult obesity-related traits, less is known about the genetics specific for early-onset obesity and with only a few studies conducted in non-European populations to date. Searching for additional genetic variants associated with childhood obesity, we performed a trans-ancestral meta-analysis of 30 studies consisting of up to 13 005 cases (≥95th percentile of body mass index (BMI) achieved 2–18 years old) and 15 599 controls (consistently METTL15). The variant was nominally associated with only the European subgroup analysis but had a consistent direction of effect in other ethnicities. We then utilized trans-ancestral Bayesian analysis to narrow down the location of the probable causal variant at each genome-wide significant signal. Of all the fine-mapped loci, we were able to narrow down the causative variant at four known loci to fewer than 10 single nucleotide polymorphisms (SNPs) (FAIM2, GNPDA2, MC4R and SEC16B loci). In conclusion, an ethnically diverse setting has enabled us to both identify an additional pediatric obesity locus and further fine-map existing loci.

Birth weight variation is influenced by fetal and maternal genetic and non-genetic factors, and has been reproducibly associated with future cardio-metabolic health outcomes. In expanded genome-wide association analyses of own birth weight (n = 321,223) and offspring birth weight (n = 230,069 mothers), we identified 190 independent association signals (129 of which are novel). We used structural equation modeling to decompose the contributions of direct fetal and indirect maternal genetic effects, then applied Mendelian randomization to illuminate causal pathways. For example, both indirect maternal and direct fetal genetic effects drive the observational relationship between lower birth weight and higher later blood pressure: maternal blood pressure-raising alleles reduce offspring birth weight, but only direct fetal effects of these alleles, once inherited, increase later offspring blood pressure. Using maternal birth weight-lowering genotypes to proxy for an adverse intrauterine environment provided no evidence that it causally raises offspring blood pressure, indicating that the inverse birth weight–blood pressure association is attributable to genetic effects, and not to intrauterine programming.

Genetic and immunological evidence clearly points to a role for T cells in the pathogenesis of rheumatoid arthritis (RA). Selective targeting of such disease-associated T cell clones might be highly effective while having few side effects. However, such selective targeting may only be feasible if the same T cell clones dominate the immune response at different sites of inflammation. We leveraged high-throughput technology to quantitatively assess whether different T cell clones dominate the inflammatory infiltrate at various sites of inflammation in this prototypic autoimmune disease. In 13 RA patients, we performed quantitative next-generation sequencing–based human TCRβ repertoire analysis in simultaneously obtained samples from inflamed synovial tissue (ST) from distinct locations within one joint, from multiple joints, and from synovial fluid (SF) and peripheral blood (PB). Identical TCRβ clones dominate inflammatory responses in ST samples taken from different locations within a single joint and when sampled in different joints. Although overall ST–SF overlap was comparable to higher ST–ST values, the overlap in dominant TCRβ clones in ST–SF comparisons was much lower than ST–ST and comparable to the low ST–PB overlap. In individual RA patients, a limited number of TCRβ clones dominate the immune response in the inflamed ST regardless of the location within a joint and which joint undergoes biopsy; in contrast, there is limited overlap of ST with SF or PB TCR repertoires. This limited breadth of the T cell response in ST of the individual RA patient indicates that development of immunotherapies that selectively modulate dominant T cell responses might be feasible.

Background A phase characterised by the presence of specific autoantibodies and arthralgia’s in the absence of clinically evident synovial inflammation often precedes the onset of rheumatoid arthritis (RA). However, only a subset of these RA-risk individuals will develop active disease in the short term¹. Recent findings show that dominant B-cell receptor (BCR) clones in peripheral blood can accurately predict imminent onset of arthritis in these RA-risk individuals.² Objectives To validate the predictive role of BCR clones in peripheral blood in RA-risk individuals in a larger cohort. Methods The BCR repertoire in peripheral blood was analysed using next-generation BCR sequencing in a prospective cohort study of 129 RA-risk individuals from Reade. Like earlier, BCR clones expanded beyond 0.5% of the total repertoire were labelled highly expanded clones (HECs), shortly referred to as dominant BCR clones, and individuals were labelled BCR-positive if peripheral blood at study baseline showed ≥5 dominant BCR clones. Results We observed that the number of dominant BCR clones was increased in RA-risk individuals who developed arthritis within 3 years, compared to RA-risk individuals who did not 10.6±5.3 vs 2.2±2.8 (mean ±SD; pRA-risk individuals (data not shown). Conclusions In this external validation cohort we could replicate the fact that dominant BCR clones in peripheral blood predict imminent onset of clinical symptoms of RA in seropositive arthralgia patients with high accuracy. Furthermore, a highly significant association correlating a higher number of dominant BCR clones with higher risk was shown. We hope these results will support evaluation of early interventions that prevent onset of arthritis. References [1] Bos WH, et al. Ann. Rheum. Dis2010;69. [2] Tak PP, et al. Ann. Rheum. Dis2017;76. Acknowledgements Intellectual property PCT/EP2017/067047. N de Vries, PP Tak, ME Doorenspleet, PL Klarenbeek. ‘METHOD FOR DETERMINING THE RISK OF DEVELOPING ARTHRITIS’. Disclosure of Interest A. Musters: None declared, M. van Beers-Tas: None declared, M. Doorenspleet: None declared, P. Klarenbeek: None declared, B. van Schaik: None declared, A. van Kampen: None declared, F. Baas: None declared, D. van Schaardenburg: None declared, N. de Vries Grant/research support from: IMI (ABIRISK, BeTheCure), CTMM (TRACER), LSH (MODIRA), Pfizer, Roche, Janssen, GSK, Consultant for: UCB, MSD

Small insertions and deletions (indels) and large structural variations (SVs) are major contributors to human genetic diversity and disease. However, mutation rates and characteristics of de novo indels and SVs in the general population have remained largely unexplored. We report 332 validated de novo structural changes identified in whole genomes of 250 families, including complex indels, retrotransposon insertions, and interchromosomal events. These data indicate a mutation rate of 2.94 indels (1–20 bp) and 0.16 SVs (>20 bp) per generation. De novo structural changes affect on average 4.1 kbp of genomic sequence and 29 coding bases per generation, which is 91 and 52 times more nucleotides than de novo substitutions, respectively. This contrasts with the equal genomic footprint of inherited SVs and substitutions. An excess of structural changes originated on paternal haplotypes. Additionally, we observed a nonuniform distribution of de novo SVs across offspring. These results reveal the importance of different mutational mechanisms to changes in human genome structure across generations.

Variants associated with blood lipid levels may be population-specific. To identify low-frequency variants associated with this phenotype, population-specific reference panels may be used. Here we impute nine large Dutch biobanks (~35,000 samples) with the population-specific reference panel created by the Genome of the Netherlands Project and perform association testing with blood lipid levels. We report the discovery of five novel associations at four loci (P value −4), including a rare missense variant in ABCA6 (rs77542162, p.Cys1359Arg, frequency 0.034), which is predicted to be deleterious. The frequency of this ABCA6 variant is 3.65-fold increased in the Dutch and its effect (β_LDL-C=0.135, β_TC=0.140) is estimated to be very similar to those observed for single variants in well-known lipid genes, such as LDLR.

Human cytomegalovirus (hCMV) infection is characterized by a vast expansion of resting effector-type virus-specific T cells in the circulation. In mice, IL-7Rα-expressing cells contain the precursors for long-lived antigen-experienced CD8⁺ T cells, but it is unclear if similar mechanisms operate to maintain these pools in humans. Here we studied whether IL-7Rα-expressing cells obtained from peripheral blood (PB) or lymph nodes (LNs) sustain the circulating effector-type hCMV-specific pool.