Trait-stratified genome-wide association study identifies novel and diverse genetic associations with serologic and cytokine phenotypes in systemic lupus erythematosus

Abstract

Introduction: Systemic lupus erythematosus (SLE) is a highly heterogeneous disorder, characterized by differences in autoantibody profile, serum cytokines, and clinical manifestations. SLE-associated autoantibodies and high serum interferon alpha (IFN-α) are important heritable phenotypes in SLE which are correlated with each other, and play a role in disease pathogenesis. These two heritable risk factors are shared between ancestral backgrounds. The aim of the study was to detect genetic factors associated with autoantibody profiles and serum IFN-α in SLE.

Methods: We undertook a case-case genome-wide association study of SLE patients stratified by ancestry and extremes of phenotype in serology and serum IFN-α. Single nucleotide polymorphisms (SNPs) in seven loci were selected for follow-up in a large independent cohort of 538 SLE patients and 522 controls using a multi-step screening approach based on novel metrics and expert database review. The seven loci were: leucine-rich repeat containing 20 (LRRC20); protein phosphatase 1 H (PPM1H); lysophosphatidic acid receptor 1 (LPAR1); ankyrin repeat and sterile alpha motif domain 1A (ANKS1A); protein tyrosine phosphatase, receptor type M (PTPRM); ephrin A5 (EFNA5); and V-set and immunoglobulin domain containing 2 (VSIG2).

Results: SNPs in the LRRC20, PPM1H, LPAR1, ANKS1A, and VSIG2 loci each demonstrated strong association with a particular serologic profile (all odds ratios > 2.2 and P < 3.5 × 10-4). Each of these serologic profiles was associated with increased serum IFN-α. SNPs in both PTPRM and LRRC20 were associated with increased serum IFN-α independent of serologic profile (P = 2.2 × 10-6 and P = 2.6 × 10-3 respectively). None of the SNPs were strongly associated with SLE in case-control analysis, suggesting that the major impact of these variants will be upon subphenotypes in SLE.

Conclusions: This study demonstrates the power of using serologic and cytokine subphenotypes to elucidate genetic factors involved in complex autoimmune disease. The distinct associations observed emphasize the heterogeneity of molecular pathogenesis in SLE, and the need for stratification by subphenotypes in genetic studies. We hypothesize that these genetic variants play a role in disease manifestations and severity in SLE.

Figure 1

Serum IFN-α levels in SLE patients stratified by SNP genotype at PTPRM (a) and LRRC20 (b). Y-axis shows the serum IFN-α activity score as outlined in the Methods section. Bars show the median, error bars show the interquartile range. Because IFN-α levels were far from normally distributed, we used a non-parametric version of the two sample t-test (non-parametric Mann-Whitney U) to test if IFN-α levels varied significantly between the two homozygous genotype groups.

Figure 2

Serum IFN-α levels in SLE patients stratified by genotype at PPM1 H (a), LPAR1 (b), ANKS1A (c), VSIG2 (d), and EFNA5 (e). Minor allele homozygotes are combined with heterozygotes for analysis. Patients are stratified first by genotype, and secondarily by the serologic factor associated with the particular SNP in Table 1, as indicated in the legend below the X-axis. Y-axis shows the serum IFN-α activity score as outlined in the Methods section. Bars show the median, error bars show the interquartile range. P-values indicate two column comparisons between the bars on the graph indicated by the line, and are calculated using the Mann-Whitney U test. Significant differences are observed either between subjects with the same genotype but with differing serological profiles (a through d), or between subjects with different genotypes but sharing the same serologic profile (e).

Figure 3

Diagrams of the patterns of association observed between SNP genotype, serologic profile, and serum IFN-α. "Gene" represents the genetic variation in the locus studied, and "Ab" represents the particular serologic profile associated with that locus. Connecting lines between nodes indicate associations, and arrowheads show the hypothesized direction of the relationship. The loci demonstrating each pattern are indicated following the letter labeling the panel. The dashed line in C. indicates that a suggestive association was seen between the EFNA5 SNP genotype and the same serologic profile which is linked to IFN-α, but this serologic association did not withstand Bonferroni correction for multiple comparisons.