Genome-wide study of longitudinal brain imaging measures of multiple sclerosis progression across six clinical trials.

While the genetics of MS risk susceptibility are well-described, and recent progress has been made on the genetics of disease severity, the genetics of disease progression remain elusive. We therefore investigated the genetic determinants of MS progression on longitudinal brain MRI: change in brain volume (BV) and change in T2 lesion volume (T2LV), reflecting progressive tissue loss and increasing disease burden, respectively. We performed genome-wide association studies of change in BV (N = 3401) and change in T2LV (N = 3513) across six randomized clinical trials from Biogen and Roche/Genentech: ADVANCE, ASCEND, DECIDE, OPERA I & II, and ORATORIO. Analyses were adjusted for randomized treatment arm, age, sex, and ancestry. Results were pooled in a meta-analysis, and were evaluated for enrichment of MS risk variants. Variant colocalization and cell-specific expression analyses were performed using published cohorts. The strongest peaks were in PTPRD (rs77321193-C/A, p = 3.9 × 10-7) for BV change, and NEDD4L (rs11398377-GC/G, p = 9.3 × 10-8) for T2LV change. Evidence of colocalization was observed for NEDD4L, and both genes showed increased expression in neuronal and/or glial populations. No association between MS risk variants and MRI outcomes was observed. In this unique, precompetitive industry partnership, we report putative regions of interest in the neurodevelopmental gene PTPRD, and the ubiquitin ligase gene NEDD4L. These findings are distinct from known MS risk genetics, indicating an added role for genetic progression analyses and informing drug discovery.

Genetic regulation of fetal hemoglobin across global populations.

Human genetic variation has enabled the identification of several key regulators of fetal-to-adult hemoglobin switching, including BCL11A, resulting in therapeutic advances. However, despite the progress made, limited further insights have been obtained to provide a fuller accounting of how genetic variation contributes to the global mechanisms of fetal hemoglobin (HbF) gene regulation. Here, we have conducted a multi-ancestry genome-wide association study of 28,279 individuals from several cohorts spanning 5 continents to define the architecture of human genetic variation impacting HbF. We have identified a total of 178 conditionally independent genome-wide significant or suggestive variants across 14 genomic windows. Importantly, these new data enable us to better define the mechanisms by which HbF switching occurs in vivo. We conduct targeted perturbations to define BACH2 as a new genetically-nominated regulator of hemoglobin switching. We define putative causal variants and underlying mechanisms at the well-studied BCL11A and HBS1L-MYB loci, illuminating the complex variant-driven regulation present at these loci. We additionally show how rare large-effect deletions in the HBB locus can interact with polygenic variation to influence HbF levels. Our study paves the way for the next generation of therapies to more effectively induce HbF in sickle cell disease and ß-thalassemia.

Systematic Mendelian randomization using the human plasma proteome to discover potential therapeutic targets for stroke.

Stroke is the second leading cause of death with substantial unmet therapeutic needs. To identify potential stroke therapeutic targets, we estimate the causal effects of 308 plasma proteins on stroke outcomes in a two-sample Mendelian randomization framework and assess mediation effects by stroke risk factors. We find associations between genetically predicted plasma levels of six proteins and stroke (P ≤ 1.62 × 10-4). The genetic associations with stroke colocalize (Posterior Probability >0.7) with the genetic associations of four proteins (TFPI, TMPRSS5, CD6, CD40). Mendelian randomization supports atrial fibrillation, body mass index, smoking, blood pressure, white matter hyperintensities and type 2 diabetes as stroke risk factors (P ≤ 0.0071). Body mass index, white matter hyperintensity and atrial fibrillation appear to mediate the TFPI, IL6RA, TMPRSS5 associations with stroke. Furthermore, thirty-six proteins are associated with one or more of these risk factors using Mendelian randomization. Our results highlight causal pathways and potential therapeutic targets for stroke.

Immune disease risk variants regulate gene expression dynamics during CD4+ T cell activation.

During activation, T cells undergo extensive gene expression changes that shape the properties of cells to exert their effector function. Understanding the regulation of this process could help explain how genetic variants predispose to immune diseases. Here, we mapped genetic effects on gene expression (expression quantitative trait loci (eQTLs)) using single-cell transcriptomics. We profiled 655,349 CD4+ T cells, capturing transcriptional states of unstimulated cells and three time points of cell activation in 119 healthy individuals. This identified 38 cell clusters, including transient clusters that were only present at individual time points of activation. We found 6,407 genes whose expression was correlated with genetic variation, of which 2,265 (35%) were dynamically regulated during activation. Furthermore, 127 genes were regulated by variants associated with immune-mediated diseases, with significant enrichment for dynamic effects. Our results emphasize the importance of studying context-specific gene expression regulation and provide insights into the mechanisms underlying genetic susceptibility to immune-mediated diseases.

Genome-wide association study identifies kallikrein 5 in type 2 inflammation-low asthma.

BACKGROUND: Clinical studies of type 2 (T2) cytokine-related neutralizing antibodies in asthma have identified a substantial subset of patients with low levels of T2 inflammation who do not benefit from T2 cytokine neutralizing antibody treatment. Non-T2 mechanisms are poorly understood in asthma but represent a redefined unmet medical need. OBJECTIVE: We sought to gain a better understanding of genetic contributions to T2-low asthma. METHODS: We utilized an unbiased genome-wide association study of patients with moderate to severe asthma stratified by T2 serum biomarker periostin. We also performed additional expression and biological analysis for the top genetic hits. RESULTS: We identified a novel protective single nucleotide polymorphism at chr19q13.41, which is selectively associated with T2-low asthma and establishes Kallikrein-related peptidase 5 (KLK5) as the causal gene mediating this association. Heterozygous carriers of the single nucleotide polymorphisms have reduced KLK5 expression. KLK5 is secreted by human bronchial epithelial cells and elevated in asthma bronchial alveolar lavage. T2 cytokines IL-4 and IL-13 downregulate KLK5 in human bronchial epithelial cells. KLK5, dependent on its catalytic function, induces epithelial chemokine/cytokine expression. Finally, overexpression of KLK5 in airway or lack of an endogenous KLK5 inhibitor, SPINK5, leads to spontaneous airway neutrophilic inflammation. CONCLUSION: Our data identify KLK5 to be the causal gene at a novel locus at chr19q13.41 associated with T2-low asthma.

A higher burden of multiple sclerosis genetic risk confers an earlier onset.

BACKGROUND: Age at onset of multiple sclerosis (MS) is an objective, influential predictor of the evolution of MS independent of disease duration. OBJECTIVES: Determine the influence of MS genetic predisposition on age of onset. METHODS: We conducted a comprehensive investigation of MS risk variants and age at onset in 3495 non-Latinx white individuals, including for combinations of HLA-DRB1*15:01 alleles and quintiles of an unweighted genetic risk score (GRS) for 198 of 200 autosomal MS risk variants that reside outside the major histocompatibility complex. RESULTS: The mean age at onset was 32 years, 29% were male, and 46% were HLA-DRB1*15:01 carriers. For those with the greatest genetic risk burden (the highest GRS quintile with two HLA-DRB1*15:01 alleles) were on average 5 years younger at onset (p = 0.002) than those with the lowest genetic risk burden (the lowest GRS quintile with no HLA-DRB1*15:01 alleles). There was a strong inverse relationship between the MS genetic risk burden and age at onset of MS (p < 5 × 10-8). CONCLUSION: We demonstrate a significant gradient between elevated MS genetic risk burden and an earlier onset of MS, suggesting that a higher MS genetic risk burden accelerates onset of the disease.

Systematic single-variant and gene-based association testing of thousands of phenotypes in 394,841 UK Biobank exomes.

Genome-wide association studies have successfully discovered thousands of common variants associated with human diseases and traits, but the landscape of rare variations in human disease has not been explored at scale. Exome-sequencing studies of population biobanks provide an opportunity to systematically evaluate the impact of rare coding variations across a wide range of phenotypes to discover genes and allelic series relevant to human health and disease. Here, we present results from systematic association analyses of 4,529 phenotypes using single-variant and gene tests of 394,841 individuals in the UK Biobank with exome-sequence data. We find that the discovery of genetic associations is tightly linked to frequency and is correlated with metrics of deleteriousness and natural selection. We highlight biological findings elucidated by these data and release the dataset as a public resource alongside the Genebass browser for rapidly exploring rare-variant association results.

Advancing human genetics research and drug discovery through exome sequencing of the UK Biobank.

The UK Biobank Exome Sequencing Consortium (UKB-ESC) is a private-public partnership between the UK Biobank (UKB) and eight biopharmaceutical companies that will complete the sequencing of exomes for all ~500,000 UKB participants. Here, we describe the early results from ~200,000 UKB participants and the features of this project that enabled its success. The biopharmaceutical industry has increasingly used human genetics to improve success in drug discovery. Recognizing the need for large-scale human genetics data, as well as the unique value of the data access and contribution terms of the UKB, the UKB-ESC was formed. As a result, exome data from 200,643 UKB enrollees are now available. These data include ~10 million exonic variants-a rich resource of rare coding variation that is particularly valuable for drug discovery. The UKB-ESC precompetitive collaboration has further strengthened academic and industry ties and has provided teams with an opportunity to interact with and learn from the wider research community.

STXBP6 and B3GNT6 Genes are Associated With Selective IgA Deficiency.

Immunoglobulin A Deficiency (IgAD) is a polygenic primary immune deficiency, with a strong genetic association to the human leukocyte antigen (HLA) region. Previous genome-wide association studies (GWAS) have identified five non-HLA risk loci (IFIH1, PVT1, ATG13-AMBRA1, AHI1 and CLEC16A). In this study, we investigated the genetic interactions between different HLA susceptibility haplotypes and non-MHC genes in IgAD. To do this, we stratified IgAD subjects and healthy controls based on HLA haplotypes (N = 10,993), and then performed GWAS to identify novel genetic regions contributing to IgAD susceptibility. After replicating previously published HLA risk haplotypes, we compared individuals carrying at least one HLA risk allele (HLA-B*08:01-DRB1*03:01-DQB1*02:01 or HLA-DRB1*07:01-DQB1*02:02 or HLA-DRB1*01-DQB1*05:01) with individuals lacking an HLA risk allele. Subsequently, we stratified subjects based on the susceptibility alleles/haplotypes and performed gene-based association analysis using 572,856 SNPs and 24,125 genes. A significant genome-wide association in STXBP6 (rs4097492; p = 7.63 × 10-9) was observed in the cohort carrying at least one MHC risk allele. We also identified a significant gene-based association for B3GNT6 (P Gene = 2.1 × 10-6) in patients not carrying known HLA susceptibility alleles. Our findings indicate that the etiology of IgAD differs depending on the genetic background of HLA susceptibility haplotypes.

Single-cell transcriptomics identifies an effectorness gradient shaping the response of CD4+ T cells to cytokines.

Naïve CD4+ T cells coordinate the immune response by acquiring an effector phenotype in response to cytokines. However, the cytokine responses in memory T cells remain largely understudied. Here we use quantitative proteomics, bulk RNA-seq, and single-cell RNA-seq of over 40,000 human naïve and memory CD4+ T cells to show that responses to cytokines differ substantially between these cell types. Memory T cells are unable to differentiate into the Th2 phenotype, and acquire a Th17-like phenotype in response to iTreg polarization. Single-cell analyses show that T cells constitute a transcriptional continuum that progresses from naïve to central and effector memory T cells, forming an effectorness gradient accompanied by an increase in the expression of chemokines and cytokines. Finally, we show that T cell activation and cytokine responses are influenced by the effectorness gradient. Our results illustrate the heterogeneity of T cell responses, furthering our understanding of inflammation.

Neurology-related protein biomarkers are associated with cognitive ability and brain volume in older age.

Identifying biological correlates of late life cognitive function is important if we are to ascertain biomarkers for, and develop treatments to help reduce, age-related cognitive decline. Here, we investigated the associations between plasma levels of 90 neurology-related proteins (Olink® Proteomics) and general fluid cognitive ability in the Lothian Birth Cohort 1936 (LBC1936, N = 798), Lothian Birth Cohort 1921 (LBC1921, N = 165), and the INTERVAL BioResource (N = 4451). In the LBC1936, 22 of the proteins were significantly associated with general fluid cognitive ability (ß between -0.11 and -0.17). MRI-assessed total brain volume partially mediated the association between 10 of these proteins and general fluid cognitive ability. In an age-matched subsample of INTERVAL, effect sizes for the 22 proteins, although smaller, were all in the same direction as in LBC1936. Plasma levels of a number of neurology-related proteins are associated with general fluid cognitive ability in later life, mediated by brain volume in some cases.

Genetic Study of Severe Prolonged Lymphopenia in Multiple Sclerosis Patients Treated With Dimethyl Fumarate.

In delayed-release dimethyl fumarate (DMF)-treated patients, absolute lymphocyte count (ALC) often declines in the first year and stabilizes thereafter; early declines have been associated with development of severe prolonged lymphopenia (SPL). Prolonged moderate or severe lymphopenia is a known risk factor for progressive multifocal leukoencephalopathy (PML); DMF-associated PML is very rare. It is unknown whether genetic predictors of SPL secondary to DMF treatment exist. We aimed to identify genetic predictors of reduced white blood cell (WBC) counts in DMF-treated multiple sclerosis (MS) patients. Genotyping (N = 1,258) and blood transcriptional profiling (N = 1,133) were performed on MS patients from DEFINE/CONFIRM. ALCs were categorized as: SPL, < 500 cells/µL for ≥6 months; moderate prolonged lymphopenia (MPL), < 800 cells/µL for ≥6 months, excluding SPL; mildly reduced lymphocytes, < 910 cells/µL at any point, excluding SPL and MPL; no lymphopenia, ≥910 cells/µL. Genome-wide association, HLA, and cross-sectional gene expression studies were performed. No common variants, HLA alleles, or expression profiles clinically useful for predicting SPL or MPL were identified. There was no overlap between genetic peaks and genetic loci known to be associated with WBC. Gene expression profiles were not associated with lymphopenia status. A classification model including gene expression features was not more predictive of lymphopenia status than standard covariates. There were no genetic predictors of SPL (or MPL) secondary to DMF treatment. Our results support ALC monitoring during DMF treatment as the most effective way to identify patients at risk of SPL.

Chromatin activity at GWAS loci identifies T cell states driving complex immune diseases.

Immune-disease-associated variants are enriched in active chromatin regions of T cells and macrophages. However, whether these variants function in specific cell states is unknown. Here we stimulated T cells and macrophages in the presence of 13 cytokines and profiled active and open chromatin regions. T cell activation induced major chromatin remodeling, while the presence of cytokines fine-tuned the magnitude of changes. We developed a statistical method that accounts for subtle changes in the chromatin landscape to identify SNP enrichment across cell states. Our results point towards the role of immune-disease-associated variants in early rather than late activation of memory CD4+ T cells, with modest differences across cytokines. Furthermore, variants associated with inflammatory bowel disease are enriched in type 1 T helper (TH1) cells, whereas variants associated with Alzheimer's disease are enriched in different macrophage cell states. Our results represent an in-depth analysis of immune-disease-associated variants across a comprehensive panel of activation states of T cells and macrophages.

Increased DNA methylation of SLFN12 in CD4+ and CD8+ T cells from multiple sclerosis patients.

DNA methylation is an epigenetic mark that is influenced by environmental factors and is associated with changes to gene expression and phenotypes. It may link environmental exposures to disease etiology or indicate important gene pathways involved in disease pathogenesis. We identified genomic regions that are differentially methylated in T cells of patients with relapsing remitting multiple sclerosis (MS) compared to healthy controls. DNA methylation was assessed at 450,000 genomic sites in CD4+ and CD8+ T cells purified from peripheral blood of 94 women with MS and 94 healthy women, and differentially methylated regions were identified using bumphunter. Differential DNA methylation was observed near four loci: MOG/ZFP57, HLA-DRB1, NINJ2/LOC100049716, and SLFN12. Increased methylation of the first exon of the SLFN12 gene was observed in both T cell subtypes and remained present after restricting analyses to samples from patients who had never been on treatment or had been off treatment for more than 2.5 years. Genes near the regions of differential methylation in T cells were assessed for differential expression in whole blood samples from a separate population of 1,329 women with MS and 97 healthy women. Gene expression of HLA-DRB1, NINJ2, and SLFN12 was observed to be decreased in whole blood in MS patients compared to controls. We conclude that T cells from MS patients display regions of differential DNA methylation compared to controls, and corresponding gene expression differences are observed in whole blood. Two of the genes that showed both methylation and expression differences, NINJ2 and SLFN12, have not previously been implicated in MS. SLFN12 is a particularly compelling target of further research, as this gene is known to be down-regulated during T cell activation and up-regulated by type I interferons (IFNs), which are used to treat MS.

Common variants at PVT1, ATG13-AMBRA1, AHI1 and CLEC16A are associated with selective IgA deficiency.

Selective immunoglobulin A deficiency (IgAD) is the most common primary immunodeficiency in Europeans. Our genome-wide association study (GWAS) meta-analysis of 1,635 patients with IgAD and 4,852 controls identified four new significant (P < 5 × 10-8) loci and association with a rare IFIH1 variant (p.Ile923Val). Peak new variants (PVT1, P = 4.3 × 10-11; ATG13-AMBRA1, P = 6.7 × 10-10; AHI1, P = 8.4 × 10-10; CLEC16A, P = 1.4 × 10-9) overlapped with autoimmune markers (3/4) and correlated with 21 putative regulatory variants, including expression quantitative trait loci (eQTLs) for AHI1 and DEXI and DNase hypersensitivity sites in FOXP3+ regulatory T cells. Pathway analysis of the meta-analysis results showed striking association with the KEGG pathway for IgA production (pathway P < 0.0001), with 22 of the 30 annotated pathway genes containing at least one variant with P ≤ 0.05 in the IgAD meta-analysis. These data suggest that a complex network of genetic effects, including genes known to influence the biology of IgA production, contributes to IgAD.

A sequence-based approach demonstrates that balancing selection in classical human leukocyte antigen (HLA) loci is asymmetric.

Balancing selection has maintained human leukocyte antigen (HLA) allele diversity, but it is unclear whether this selection is symmetric (all heterozygotes are comparable and all homozygotes are comparable in terms of fitness) or asymmetric (distinct heterozygote genotypes display greater fitness than others). We tested the hypothesis that HLA is under asymmetric balancing selection in populations by estimating allelic branch lengths from genetic sequence data encoding peptide-binding domains. Significant deviations indicated changes in the ratio of terminal to internal branch lengths. Such deviations could arise even if no individual alleles present a strikingly altered branch length (e.g. if there is an overall distortion, with all or many terminal branches being longer than expected). DQ and DP loci were also analyzed as haplotypes. Using allele frequencies for 419 distinct populations in 10 geographical regions, we examined population differentiation in alleles within and between regions, and the relationship between allelic branch length and frequency. The strongest evidence for asymmetrical balancing selection was observed for HLA-DRB1, HLA-B and HLA-DPA1, with significant deviation (P ≤ 1.1 × 10(-4)) in about half of the populations. There were significant results at all loci except HLA-DQB1/DQA1. We observed moderate genetic variation within and between geographic regions, similar to the rest of the genome. Branch length was not correlated with allele frequency. In conclusion, sequence data suggest that balancing selection in HLA is asymmetric (some heterozygotes enjoy greater fitness than others). Because HLA polymorphism is crucial for pathogen resistance, this may manifest as a frequency-dependent selection with fluctuation in the fitness of specific heterozygotes over time.

The genetics of type I interferon in systemic lupus erythematosus.

The discovery that type I interferon (IFN)-inducible genes were strongly upregulated in peripheral blood in SLE over a decade ago sparked interest in understanding the relationship between type I IFN and SLE. Genome-wide association studies provide strong genetic evidence that type I IFNs are important for SLE risk. Of 47 genetic variants associated with SLE, over half (27/47, 57%) can be linked to type I IFN production or signaling. The recent identification of single gene mutations for disorders that share features with SLE--Aicardi-Goutières syndrome, chilblain lupus, and spondyloenchondrodysplasia--provide additional support for the hypothesis that type I IFNs are central drivers of SLE pathogenesis. These insights provide significant focus for efforts to tackle SLE therapeutically.

CIITA variation in the presence of HLA-DRB1*1501 increases risk for multiple sclerosis.

The MHC class II transactivator gene (CIITA) is an important transcription factor regulating gene required for HLA class II MHC-restricted antigen presentation. Association with HLA class II variation, particularly HLA-DRB1*1501, has been well-established for multiple sclerosis (MS). In addition, the -168A/G CIITA promoter variant (rs3087456) has been reported to be associated with MS. Thus, a multi-stage investigation of variation within CIITA, DRB1*1501 and MS was undertaken in 6108 individuals. In stage 1, 24 SNPs within CIITA were genotyped in 1320 cases and 1363 controls (n = 2683). Rs4774 (missense +1614G/C; G500A) was associated with MS (P = 4.9 x 10(-3)), particularly in DRB1*1501 +individuals (P = 1 x 10(-4)). No association was observed for the -168A/G promoter variant. In stage 2, rs4774 was genotyped in 973 extended families; rs4774*C was also associated with increased risk for MS in DRB1*1501+ families (P = 2.3 x 10(-2)). In a third analysis, rs4774 was tested in cases and controls (stage 1) combined with one case per family (stage 2) for increased power. Rs4774*C was associated with MS (P = 1 x 10(-3)), particularly in DRB1*1501+ cases and controls (P = 1 x 10(-4)). Results obtained from logistic regression analysis showed evidence for interaction between rs4774*C and DRB1*1501 associated with risk for MS (ratio of ORs = 1.72, 95% CI 1.28-2.32, P = 3 x 10(-4)). Furthermore, rs4774*C was associated with DRB1*1501+ MS when conditioned on the presence (OR = 1.67, 95% CI = 1.19-2.37, P = 1.9 x 10(-3)) and absence (OR = 1.49, 95% CI = 1.15-1.95, P = 2.3 x 10(-3)) of CLEC16A rs6498169*G, a putative MS risk allele adjacent to CIITA. Our results provide strong evidence supporting a role for CIITA variation in MS risk, which appears to depend on the presence of DRB1*1501.

Analysis of maternal-offspring HLA compatibility, parent-of-origin effects, and noninherited maternal antigen effects for HLA-DRB1 in systemic lupus erythematosus.

OBJECTIVE: Genetic susceptibility to systemic lupus erythematosus (SLE) is well established, with the HLA class II DRB1 and DQB1 loci demonstrating the strongest association. However, HLA may also influence SLE through novel biologic mechanisms in addition to genetic transmission of risk alleles. Evidence for increased maternal-offspring HLA class II compatibility in SLE and differences in maternal versus paternal transmission rates (parent-of-origin effects) and nontransmission rates (noninherited maternal antigen [NIMA] effects) in other autoimmune diseases have been reported. Thus, we investigated maternal-offspring HLA compatibility, parent-of-origin effects, and NIMA effects at DRB1 in SLE. METHODS: The cohort comprised 707 SLE families and 188 independent healthy maternal-offspring pairs (total of 2,497 individuals). Family-based association tests were conducted to compare transmitted versus nontransmitted alleles (transmission disequilibrium test) and both maternally versus paternally transmitted (parent-of-origin) and nontransmitted alleles (using the chi-square test of heterogeneity). Analyses were stratified according to the sex of the offspring. Maternally affected offspring DRB1 compatibility in SLE families was compared with paternally affected offspring compatibility and with independent control maternal-offspring pairs (using Fisher's test) and was restricted to male and nulligravid female offspring with SLE. RESULTS: As expected, DRB1 was associated with SLE (P < 1 x 10(-4)). However, mothers of children with SLE had similar transmission and nontransmission frequencies for DRB1 alleles when compared with fathers, including those for the known SLE risk alleles HLA-DRB1*0301, *1501, and *0801. No association between maternal-offspring compatibility and SLE was observed. CONCLUSION: Maternal-offspring HLA compatibility, parent-of-origin effects, and NIMA effects at DRB1 are unlikely to play a role in SLE.