A Genome-Wide Association Study Suggests New Susceptibility Loci for Primary Antiphospholipid Syndrome

Abstract

Objective: Primary antiphospholipid syndrome (PAPS) is a rare autoimmune disease characterized by the presence of antiphospholipid antibodies and the occurrence of thrombotic events and pregnancy complications. Our study aimed to identify novel genetic susceptibility loci associated with PAPS.

Methods: We performed a genome-wide association study comprising 5,485 individuals (482 affected individuals) of European ancestry. Significant and suggestive independent variants from a meta-analysis of approximately 7 million variants were evaluated for functional and biological process enrichment. The genetic risk variability for PAPS in different populations was also assessed. Hierarchical clustering, Mahalanobis distance, and Dirichlet Process Mixtures with uncertainty clustering methods were used to assess genetic similarities between PAPS and other immune-mediated diseases.

Results: We revealed genetic associations with PAPS in a regulatory locus within the HLA class II region near HLA-DRA and in STAT1-STAT4 with a genome-wide level of significance; 34 additional suggestive genetic susceptibility loci for PAPS were also identified. The disease risk allele near HLA-DRA is associated with overexpression of HLA-DRB6, HLA-DRB9, HLA-DQA2, and HLA-DQB2 in immune cells, vascular tissue, and nervous tissue. This association is independent of the association between PAPS and HLA-DRB1*1302. Functional analyses highlighted immune-related pathways in PAPS-associated loci. The comparison with other immune-mediated diseases revealed a close genetic relatedness to neuromyelitis optica, systemic sclerosis, and Sjögren syndrome, suggesting co-localized causal variations close to STAT1-STAT4, TNPO3, and BLK.

Conclusion: This study represents a comprehensive large-scale genetic analysis for PAPS and provides new insights into the genetic basis and pathophysiology of this rare disease.

Figure 1.

Results of the meta-analysis among the five PAPS cohorts included in this study. (A) Manhattan plot depicting the results across the whole genome. Y and X axes refer to the −log₁₀ p-values and chromosome positions, respectively. The red horizontal line represents the genome-wide association threshold (p-value <5×10⁻⁸) and the blue line represents the suggestive threshold (p-value<1×10⁻⁵). (B) Regional plot for the genome-wide associated signals in chromosome 2 and (C) chromosome 6, respectively. The y-axis of the regional plots corresponds to the −log₁₀ p-values of evaluated variants while the x-axis indicates variants’ positions. Purple diamonds correspond to the most associated variants in each region. The rest of the variants are colored according to their degree of linkage disequilibrium (LD) with each top signal based on pairwise r² values from the European population from the 1000 Genomes Project. The grey dashed line corresponds to the genome-wide association threshold (p-value<5×10⁻⁸) and the blue line to the estimated recombination rates. Genes within the selected regions are shown at the bottom of the plots.

Figure 2.

Violin plots representing the differences in gene expression levels in whole blood (y-axis) of the HLA genes HLA-DRB6, HLA-DRB9, HLA-DPB2, HLA-DQA2 and HLA-DQB2 depending on the genotypes of the PAPS-associated variant within the HLA region, rs9269041 (x-axis). When the PAPS-risk allele is present (rs9269041-A), higher gene expression levels are observed.

Figure 3.

Functional annotation results for the variants (A) rs35930163 and (B) rs9411271 located close to the genes DNM3 and GPSM1, respectively. Left panels indicate epigenetic mark binding at each variant. Central panels represent the physical chromatin interactions established by the regions containing each variant in the lymphoblastoid cell line GM12878; the red lines represent all the interactions of each region while the yellow ones refer to the interaction with the indicated genes. The right panels show violin plots representing the differences in expression levels (y-axis) of the interacting genes depending on the genotypes of each PAPS-associated variant (x-axis).

Figure 4.

Results of etiologic similarity analyses of PAPS with other immune-mediated diseases (IMDs). (A) Hierarchical clustering of PAPS with other immune-mediated diseases GWAS data and self-reported disease traits in UK Biobank, FinnGen Project, and The International Multiple Sclerosis Genetics Consortium. PAPS appears in boldface. Heatmaps indicate delta values for each disease on each component PC1-PC13, with grey indicating 0 (no difference from control), and darker shades of blue or magenta showing departure from controls in one direction or the other. A filled circle indicates that the delta value was significantly non-zero with a significance of FDR <1%, while an unfilled circle indicates a significance of FDR <5%. (B) Top 20 closest Mahalanobis distances to PAPS of all tested traits. The Y axis displays the distance between the center of the distribution (PAPS) and different points (the IMDs). (C) Clustering analysis of PAPS with IMDs using the Dirichlet Process Mixtures with uncertainty method (DPMUnc). Features in which PAPS was significant at FDR 5% level are depicted, comparing PC1 versus PC13 (top) and PC1 versus PC2 (bottom).