Search

BackgroundDisabling pansclerotic morphea (DPM) is a rare systemic inflammatory disorder, characterized by poor wound healing, fibrosis, cytopenias, hypogammaglobulinemia, and squamous-cell carcinoma. The cause is unknown, and mortality is high.MethodsWe evaluated four patients from three unrelated families with an autosomal dominant pattern of inheritance of DPM. Genomic sequencing independently identified three heterozygous variants in a specific region of the gene that encodes signal transducer and activator of transcription 4 (STAT4). Primary skin fibroblast and cell-line assays were used to define the functional nature of the genetic defect. We also assayed gene expression using single-cell RNA sequencing of peripheral-blood mononuclear cells to identify inflammatory pathways that may be affected in DPM and that may respond to therapy.ResultsGenome sequencing revealed three novel heterozygous missense gain-of-function variants in STAT4. In vitro, primary skin fibroblasts showed enhanced interleukin-6 secretion, with impaired wound healing, contraction of the collagen matrix, and matrix secretion. Inhibition of Janus kinase (JAK)-STAT signaling with ruxolitinib led to improvement in the hyperinflammatory fibroblast phenotype in vitro and resolution of inflammatory markers and clinical symptoms in treated patients, without adverse effects. Single-cell RNA sequencing revealed expression patterns consistent with an immunodysregulatory phenotype that were appropriately modified through JAK inhibition.ConclusionsGain-of-function variants in STAT4 caused DPM in the families that we studied. The JAK inhibitor ruxolitinib attenuated the dermatologic and inflammatory phenotype in vitro and in the affected family members. (Funded by the American Academy of Allergy, Asthma, and Immunology Foundation and others.).

Reversible modification of proteins with linkage-specific ubiquitin chains is critical for intracellular signaling. Information on physiological roles and underlying mechanisms of particular ubiquitin linkages during human development are limited. Here, relying on genomic constraint scores, we identify 10 patients with multiple congenital anomalies caused by hemizygous variants in OTUD5, encoding a K48/K63 linkage-specific deubiquitylase. By studying these mutations, we find that OTUD5 controls neuroectodermal differentiation through cleaving K48-linked ubiquitin chains to counteract degradation of select chromatin regulators (e.g., ARID1A/B, histone deacetylase 2, and HCF1), mutations of which underlie diseases that exhibit phenotypic overlap with OTUD5 patients. Loss of OTUD5 during differentiation leads to less accessible chromatin at neuroectodermal enhancers and aberrant gene expression. Our study describes a previously unidentified disorder we name LINKED (LINKage-specific deubiquitylation deficiency-induced Embryonic Defects) syndrome and reveals linkage-specific ubiquitin cleavage from chromatin remodelers as an essential signaling mode that coordinates chromatin remodeling during embryogenesis.

Systemic sclerosis (SSc) is a clinically heterogeneous autoimmune disease characterized by mutually exclusive autoantibodies directed against distinct nuclear antigens. We examined HLA associations in SSc and its autoantibody subsets in a large, newly recruited African American (AA) cohort and among European Americans (EA). In the AA population, the African ancestry-predominant HLA-DRB1*08:04 and HLA-DRB1*11:02 alleles were associated with overall SSc risk, and the HLA-DRB1*08:04 allele was strongly associated with the severe antifibrillarin (AFA) antibody subset of SSc (odds ratio = 7.4). These African ancestry-predominant alleles may help explain the increased frequency and severity of SSc among the AA population. In the EA population, the HLA-DPB1*13:01 and HLA-DRB1*07:01 alleles were more strongly associated with antitopoisomerase (ATA) and anticentromere antibody-positive subsets of SSc, respectively, than with overall SSc risk, emphasizing the importance of HLA in defining autoantibody subtypes. The association of the HLA-DPB1*13:01 allele with the ATA+ subset of SSc in both AA and EA patients demonstrated a transancestry effect. A direct correlation between SSc prevalence and HLA-DPB1*13:01 allele frequency in multiple populations was observed (r = 0.98, P = 3 × 10-6). Conditional analysis in the autoantibody subsets of SSc revealed several associated amino acid residues, mostly in the peptide-binding groove of the class II HLA molecules. Using HLA α/β allelic heterodimers, we bioinformatically predicted immunodominant peptides of topoisomerase 1, fibrillarin, and centromere protein A and discovered that they are homologous to viral protein sequences from the Mimiviridae and Phycodnaviridae families. Taken together, these data suggest a possible link between HLA alleles, autoantibodies, and environmental triggers in the pathogenesis of SSc.

We conducted a genome-wide association study of rheumatoid arthritis in 2,418 cases and 4,504 controls from North America and identified an association at the REL locus, encoding c-Rel, on chromosome 2p13 (rs13031237, P = 6.01 x 10(-10)). Replication in independent case-control datasets comprising 2,604 cases and 2,882 controls confirmed this association, yielding an allelic OR = 1.25 (P = 3.08 x 10(-14)) for marker rs13031237 and an allelic OR = 1.21 (P = 2.60 x 10(-11)) for marker rs13017599 in the combined dataset. The combined dataset also provides definitive support for associations at both CTLA4 (rs231735; OR = 0.85; P = 6.25 x 10(-9)) and BLK (rs2736340; OR = 1.19; P = 5.69 x 10(-9)). c-Rel is an NF-kappaB family member with distinct functional properties in hematopoietic cells, and its association with rheumatoid arthritis suggests disease pathways that involve other recently identified rheumatoid arthritis susceptibility genes including CD40, TRAF1, TNFAIP3 and PRKCQ.

Rheumatoid arthritis (RA) is a chronic, systemic autoimmune disease affecting both joints and extra-articular tissues. Although some genetic risk factors for RA are well-established, most notably HLA-DRB1 and PTPN22, these markers do not fully account for the observed heritability. To identify additional susceptibility loci, we carried out a multi-tiered, case-control association study, genotyping 25,966 putative functional SNPs in 475 white North American RA patients and 475 matched controls. Significant markers were genotyped in two additional, independent, white case-control sample sets (661 cases/1322 controls from North America and 596 cases/705 controls from The Netherlands) identifying a SNP, rs1953126, on chromosome 9q33.2 that was significantly associated with RA (OR(common) = 1.28, trend P(comb) = 1.45E-06). Through a comprehensive fine-scale-mapping SNP-selection procedure, 137 additional SNPs in a 668 kb region from MEGF9 to STOM on 9q33.2 were chosen for follow-up genotyping in a staged-approach. Significant single marker results (P(comb) 5.41E-09). The observed association patterns for these SNPs had heightened statistical significance and a higher degree of consistency across sample sets. In addition, the allele frequencies for these SNPs displayed reduced variability between control groups when compared to other SNPs. Lastly, in combination with the other two known genetic risk factors, HLA-DRB1 and PTPN22, the variants reported here generate more than a 45-fold RA-risk differential.

Objective: To determine whether systemic juvenile idiopathic arthritis (JIA) susceptibility loci that were identified by candidate gene studies demonstrate association with systemic JIA in the largest study population assembled to date. Methods: Single‐nucleotide polymorphisms (SNPs) from 11 previously reported systemic JIA risk loci were examined for association in 9 populations, including 770 patients with systemic JIA and 6,947 controls. The effect of systemic JIA–associated SNPs on gene expression was evaluated in silico in paired whole genome and RNA sequencing data from the lymphoblastoid cell lines (LCLs) of 373 European subjects from the 1000 Genomes Project. Responses of systemic JIA–associated SNPs to anakinra treatment were evaluated in 38 US patients for whom treatment response data were available. Results: We found no association between the previously reported 26 SNPs and systemic JIA. Expanded analysis of the regions containing the 26 SNPs revealed only 1 significant association: the promoter region of IL1RN (P < 1 × 10–4). Systemic JIA–associated SNPs correlated with IL1RN expression in LCLs, with an inverse correlation between systemic JIA risk and IL1RN expression. The presence of homozygous IL1RN high expression alleles correlated strongly with a lack of response to anakinra therapy (odds ratio 28.7 [95% confidence interval 3.2–255.8]). Conclusion: In our study, IL1RN was the only candidate locus associated with systemic JIA. The implicated SNPs are among the strongest known determinants of IL1RN and interleukin‐1 receptor antagonist levels, linking low expression with increased systemic JIA risk. Homozygous high expression alleles predicted nonresponsiveness to anakinra therapy, making them ideal candidate biomarkers to guide systemic JIA treatment. This study is an important first step toward the personalized treatment of systemic JIA. [ABSTRACT FROM AUTHOR]

Objective To describe the genotypes, phenotypes, immunophenotypes, and treatments of PAPA syndrome (pyogenic sterile arthritis, pyoderma gangrenosum, and acne), a rare autoinflammatory disease, in 5 patients. Methods Clinical information was gathered from medical records and through interviews with 5 patients from 4 kindreds. PSTPIP1 ( CD2BP1) exon 10 and exon 11 sequencing was performed in each patient. Neutrophil granule content and cytokine levels were determined in plasma and stimulated peripheral blood mononuclear cells (PBMCs) from patients and controls. Results We identified 2 previously described PAPA syndrome-associated PSTPIP1 mutations, A230T and E250Q, and a novel change, E250K. Disease penetrance was incomplete, with variable expressivity. The cutaneous manifestations included pathergy, cystic acne, and pyoderma gangrenosum. Interleukin-1β (IL-1β) and circulating neutrophil granule enzyme levels were markedly elevated in patients compared to those in controls. PBMC stimulation studies demonstrated impaired production of IL-10 and enhanced production of granulocyte-macrophage colony-stimulating factor. Good resolution of pyoderma gangrenosum was achieved in 3 patients with tumor necrosis factor α (TNFα) blockade treatment. Conclusion This analysis of 5 patients demonstrates that mutations in PSTPIP1 are incompletely penetrant and variably expressed in the PAPA syndrome. Neutrophil granule proteins are markedly elevated ex vivo and in the plasma, and elevated levels might be compatible with a diagnosis of PAPA syndrome. TNFα blockade appears to be effective in treating the cutaneous manifestations of PAPA syndrome. [ABSTRACT FROM AUTHOR]

This article discusses findings of a study which investigated the structural effect of disease-causing mutations on cryopyrin to gain better understanding of the impact of disease-associated mutations on protein function. Also known as cryopyrin-associated periodic syndromes, the cryopyrinopathies are rare autoinflammatory diseases that have been characterized at the molecular level with the identification of mutations in the cold-induced autoinflammatory syndrome 1 (CIAS1).

Familial Mediterranean fever (FMF; MIM 249100) is an autosomal recessive disease characterized by recurrent attacks of fever with synovial, pleural or peritoneal inflammation. The disease is caused by mutations in the gene encoding the pyrin protein. Human population studies have revealed extremely high allele frequencies for several different pyrin mutations, leading to the conclusion that the mutant alleles confer a selective advantage. Here we examine the ret finger protein (rfp) domain (which contains most of the disease-causing mutations) of pyrin during primate evolution. Amino acids that cause human disease are often present as wild type in other species. This is true at positions 653 (a novel mutation), 680, 681, 726, 744 and 761. For several of these human mutations, the mutant represents the reappearance of an ancestral amino acid state. Examination of lineage-specific dN/dS ratios revealed a pattern consistent with the signature of episodic positive selection. Our data, together with previous human population studies, indicate that selective pressures may have caused functional evolution of pyrin in humans and other primates. [ABSTRACT FROM AUTHOR]

Mutations of the tumor necrosis factor receptor 1 (TNFRSF1A) gene underly susceptibility to a subset of autosomal dominant recurrent fevers (ADRFs). We report on a two-generation six-member Dutch family in which a novel R92P mutation and reduced plasma TNFRSFIA levels were found in all the children, including two who are unaffected. However, only the daughter proband and father exhibited a typical TNF-receptor associated periodic syndrome (TRAPS) phenotype. PCR-RFLP analysis revealed that the mutation was not present in 120 control chromosomes from unaffected Dutch individuals. As this R92P mutation is present in two unaffected carriers it appears to be less penetrant than previously reported TNFRSF1A mutations involving cysteine residues in the extracellular domains. [ABSTRACT FROM AUTHOR]

The pyrin domain was first noted in the familial Mediterranean fever protein from which it takes its name. It belongs to a structural superfamily that includes death domains, death effector domains and caspase activation and recruitment domains. Several genes underlying autoinflammatory diseases have at least one of these four death domain-fold motifs. Mutations in CIAS1, encoding cryopyrin, a leukocyte-specific member of this growing superfamily, have now been shown to cause both familial cold autoinflammatory syndrome and Muckle-Wells syndrome. These new findings add to the growing body of evidence that the dysregulation of leukocyte apoptosis may be a common molecular pathway leading to inflammatory disease. [ABSTRACT FROM AUTHOR]

Abstract: Affected relatives are essential for pedigree linkage analysis, however, they cause a violation of the independent sample assumption in case–control association studies. To avoid the correlation between samples, a common practice is to take only one affected sample per pedigree in association analysis. Although several methods exist in handling correlated samples, they are still not widely used in part because these are not easily implemented, or because they are not widely known. We advocate the effective sample size method as a simple and accessible approach for case–control association analysis with correlated samples. This method modifies the chi-square test statistic, p-value, and 95% confidence interval of the odds-ratio by replacing the apparent number of allele or genotype counts with the effective ones in the standard formula, without the need for specialized computer programs. We present a simple formula for calculating effective sample size for many types of relative pairs and relative sets. For allele frequency estimation, the effective sample size method captures the variance inflation exactly. For genotype frequency, simulations showed that effective sample size provides a satisfactory approximation. A gene which is previously identified as a type 1 diabetes susceptibility locus, the interferon-induced helicase gene (IFIH1), is shown to be significantly associated with rheumatoid arthritis when the effective sample size method is applied. This significant association is not established if only one affected sib per pedigree were used in the association analysis. Relationship between the effective sample size method and other methods – the generalized estimation equation, variance of eigenvalues for correlation matrices, and genomic controls – are discussed. [Copyright &y& Elsevier]

Summary: NLRP3 nucleates the inflammasome, a protein complex responsible for cleavage of prointerleukin-1β (IL-1β) to its active form. Mutations in the NLRP3 gene cause the autoinflammatory disease spectrum cryopyrin-associated periodic syndromes (CAPS). The central role of IL-1β in CAPS is supported by the response to IL-1-targeted therapy. We developed two Nlrp3 mutant knockin mouse strains to model CAPS to examine the role of other inflammatory mediators and adaptive immune responses in an innate immune-driven disease. These mice had systemic inflammation and poor growth, similar to some human CAPS patients, and demonstrated early mortality, primarily mediated by myeloid cells. Mating these mutant mice to various gene mutant backgrounds showed that the mouse disease phenotype required an intact inflammasome, was only partially dependent on IL-1β, and was independent of T cells. These data suggest that CAPS are true inflammasome-mediated diseases and provide insight for more common inflammatory disorders. [Copyright &y& Elsevier]