We performed a multistage genome-wide association study of melanoma. In a discovery cohort of 1804 melanoma cases and 1026 controls, we identified loci at chromosomes 15q13.1 (HERC2/OCA2 region) and 16q24.3 (MC1R) regions that reached genome-wide significance within this study and also found strong evidence for genetic effects on susceptibility to melanoma from markers on chromosome 9p21.3 in the p16/ARF region and on chromosome 1q21.3 (ARNT/LASS2/ANXA9 region). The most significant single-nucleotide polymorphisms (SNPs) in the 15q13.1 locus (rs1129038 and rs12913832) lie within a genomic region that has profound effects on eye and skin color; notably, 50% of variability in eye color is associated with variation in the SNP rs12913832. Because eye and skin colors vary across European populations, we further evaluated the associations of the significant SNPs after carefully adjusting for European substructure. We also evaluated the top 10 most significant SNPs by using data from three other genome-wide scans. Additional in silico data provided replication of the findings from the most significant region on chromosome 1q21.3 rs7412746 (P = 6 × 10−10). Together, these data identified several candidate genes for additional studies to identify causal variants predisposing to increased risk for developing melanoma.

Chromosome 5p15.33 has been identified by genome-wide association studies as one of the regions that associate with lung cancer risk. A few single-nucleotide polymorphisms (SNPs) in the telomerase reverse transcriptase (TERT) and cleft lip and palate transmembrane 1-like (CLPTM1L) genes located in this region have shown consistent associations. We performed dense genotyping of SNPs in this region to refine the previously reported association signals for lung cancer risk. Two hundred and fifteen SNPs were genotyped on an Illumina iSelect panel, in a hospital-based case–control study of 1681 lung cancer cases and 1235 unaffected controls. Association was tested using unconditional logistic regression, while adjusting for age, sex and pack-years smoked. Furthermore, since many of the SNPs were in linkage disequilibrium (LD), haplotype blocks were constructed, from which tagging SNPs at an r2 threshold of ≥0.95 were included in a stepwise forward selection logistic regression model. Of the 215 SNPs, 69 were significant at P < 0.05 in univariate analysis; of these, 35 SNPs meeting the r2 threshold were included in the multiple logistic regression model. Two SNPs, rs370348 (odds ratio = 0.76, P = 1.6 × 10−6) and rs4975538 (odds ratio = 1.18, P = 0.005), significantly associated with risk in the overall sample. Among ever smokers, rs4975615 (odds ratio = 0.75, P = 1.2 × 10−4) and rs4975538 (odds ratio = 1.26, P = 0.002) were significant, whereas among never-smokers, rs451360 (odds ratio = 0.62, P = 7.6 × 10−5) was significant. We refined the consistent association signal in this region, allowing for the considerable LD between SNPs and identified four novel SNPs that were independently and significantly associated with lung cancer risk. Results of these analyses strongly suggest effects on risk from several loci in the TERT/CLPTM1L region.

DNA repair genes are important for maintaining genomic stability and limiting carcinogenesis. We analyzed all single nucleotide polymorphisms (SNPs) of 125 DNA repair genes covered by the Illumina HumanHap300 (v1.1) BeadChips in a previously conducted genome-wide association study (GWAS) of 1,154 lung cancer cases and 1,137 controls and replicated the top-hits of XRCC4 SNPs in an independent set of 597 cases and 611 controls in Texas populations. We found that six of 20 XRCC4 SNPs were associated with a decreased risk of lung cancer with a P value of 0.01 or lower in the discovery dataset, of which the most significant SNP was rs10040363 (P for allelic test = 4.89 ×10−4). Moreover, the data in this region allowed us to impute a potentially functional SNP rs2075685 (imputed P for allelic test = 1.3 ×10−3). A luciferase reporter assay demonstrated that the rs2075685G>T change in the XRCC4 promoter increased expression of the gene. In the replication study of rs10040363, rs1478486, rs9293329, and rs2075685, however, only rs10040363 achieved a borderline association with a decreased risk of lung cancer in a dominant model (adjusted OR = 0.80, 95% CI = 0.62–1.03, P = 0.079). In the final combined analysis of both the Texas GWAS discovery and replication datasets, the strength of the association was increased for rs10040363 (adjusted OR = 0.77, 95% CI = 0.66–0.89, Pdominant = 5×10−4 and P for trend = 5×10−4) and rs1478486 (adjusted OR = 0.82, 95% CI = 0.71 −0.94, Pdominant = 6×10−3 and P for trend = 3.5×10−3). Finally, we conducted a meta-analysis of these XRCC4 SNPs with available data from published GWA studies of lung cancer with a total of 12,312 cases and 47,921 controls, in which none of these XRCC4 SNPs was associated with lung cancer risk. It appeared that rs2075685, although associated with increased expression of a reporter gene and lung cancer risk in the Texas populations, did not have an effect on lung cancer risk in other populations. This study underscores the importance of replication using published data in larger populations.

Published genome-wide association studies (GWASs) have identified few variants in the known biological pathways involved in lung cancer etiology. To mine the possibly hidden causal single nucleotide polymorphisms (SNPs), we explored all SNPs in the extrinsic apoptosis pathway from our published GWAS dataset for 1154 lung cancer cases and 1137 cancer-free controls. In an initial association analysis of 611 tagSNPs in 41 apoptosis-related genes, we identified only 10 tagSNPs associated with lung cancer risk with a P value <10−2, including four tagSNPs in DAPK1 and three tagSNPs in TNFSF8. Unlike DAPK1 SNPs, TNFSF8 rs2181033 tagged other four predicted functional but untyped SNPs (rs776576, rs776577, rs31813148 and rs2075533) in the promoter region. Therefore, we further tested binding affinity of these four SNPs by performing the electrophoretic mobility shift assay. We found that only rs2075533T allele modified levels of nuclear proteins bound to DNA, leading to significantly decreased expression of luciferase reporter constructs by 5- to –10-fold in H1299, HeLa and HCT116 cell lines compared with the C allele. We also performed a replication study of the untyped rs2075533 in an independent Texas population but did not confirm the protective effect. We further performed a mini meta-analysis for SNPs of TNFSF8 obtained from other four published lung cancer GWASs with 12  214 cases and 47  721 controls, and we found that only rs3181366 (r2 = 0.69 with the untyped rs2075533) was associated to lung cancer risk (P = 0.008). Our findings suggest a possible role of novel TNFSF8 variants in susceptibility to lung cancer.

Alopecia areata (AA) is among the most highly prevalent human autoimmune diseases, leading to disfiguring hair loss due to the collapse of immune privilege of the hair follicle and subsequent autoimmune attack1,2. The genetic basis of AA is largely unknown. We undertook a genome-wide association study (GWAS) in a sample of 1,054 cases and 3,278 controls and identified 139 single nucleotide polymorphisms that are significantly associated with AA (P ≤ 5 × 10−7). Here we show an association with genomic regions containing several genes controlling the activation and proliferation of regulatory T cells (Treg cells), cytotoxic T lymphocyte-associated antigen 4 (CTLA4), interleukin (IL)-2/IL-21, IL-2 receptor A (IL-2RA; CD25) and Eos (also known as Ikaros family zinc finger 4; IKZF4), as well as the human leukocyte antigen (HLA) region. We also find association evidence for regions containing genes expressed in the hair follicle itself (PRDX5 and STX17). A region of strong association resides within the ULBP (cytomegalovirus UL16-binding protein) gene cluster on chromosome 6q25.1, encoding activating ligands of the natural killer cell receptor NKG2D that have not previously been implicated in an autoimmune disease. By probing the role of ULBP3 in disease pathogenesis, we also show that its expression in lesional scalp from patients with AA is markedly upregulated in the hair follicle dermal sheath during active disease. This study provides evidence for the involvement of both innate and acquired immunity in the pathogenesis of AA. We have defined the genetic underpinnings of AA, placing it within the context of shared pathways among autoimmune diseases, and implicating a novel disease mechanism, the upregulation of ULBP ligands, in triggering autoimmunity.

Genomic deletions have long been known to play a causative role in microdeletion syndromes. Recent whole-genome genetic studies have shown that deletions can increase the risk for several psychiatric disorders, suggesting that genomic deletions play an important role in the genetic basis of complex traits. However, the association between genomic deletions and common, complex diseases has not yet been systematically investigated in gene mapping studies. Likelihood-based statistical methods for identifying disease-associated deletions have recently been developed for familial studies of parent-offspring trios. The purpose of this study is to develop statistical approaches for detecting genomic deletions associated with complex disease in case–control studies. Our methods are designed to be used with dense single nucleotide polymorphism (SNP) genotypes to detect deletions in large-scale or whole-genome genetic studies. As more and more SNP genotype data for genome-wide association studies become available, development of sophisticated statistical approaches will be needed that use these data. Our proposed statistical methods are designed to be used in SNP-by-SNP analyses and in cluster analyses based on combined evidence from multiple SNPs. We found that these methods are useful for detecting disease-associated deletions and are robust in the presence of linkage disequilibrium using simulated SNP data sets. Furthermore, we applied the proposed statistical methods to SNP genotype data of chromosome 6p for 868 rheumatoid arthritis patients and 1,197 controls from the North American Rheumatoid Arthritis Consortium. We detected disease-associated deletions within the region of human leukocyte antigen in which genomic deletions were previously discovered in rheumatoid arthritis patients.

We performed linkage analysis on families with rheumatoid arthritis, stratifying by ethnic origin. We compared results using either Kong and Cox nonparametric LOD scores or MOD score analysis using the software GeneHunter MODSCORE. We first applied SNPLINK to remove markers showing excess linkage disequilibrium from the SNPs in the Illumina IV SNP Linkage panel. In this analysis there were 659 self-reported Caucasian families and 29 self-reported Hispanic families in the NARAC collection. Chromosome 19 yielded MOD scores > 3.00 in the Hispanic group, while chromosomes 2, 6, 7, 11, and XY had MOD scores > 3.00 in the Caucasian group. We performed simulation studies to evaluate the empirical distribution of the MOD score for autosomal loci separately in Hispanics and Caucasians. Results showed genome-wide significant evidence for linkage in Caucasians for chromosomes 2q and 6p, but no significant evidence for any linkages in the Hispanics, including little evidence for linkage to chromosome 6p in this group. An examination of the difference of phenotypes in two ethnic groups suggested significantly earlier mean age of onset, higher percentage of anti-cyclic citrullinated peptide positive people, and lower percentage of affected people carrying shared epitopes in Hispanics than those in Caucasians. A larger sample size of the Hispanic group is needed to identify linkage regions.

BACKGROUND

Rheumatoid arthritis has a complex mode of inheritance. Although HLA-DRB1 and PTPN22 are well-established susceptibility loci, other genes that confer a modest level of risk have been identified recently. We carried out a genomewide association analysis to identify additional genetic loci associated with an increased risk of rheumatoid arthritis.

METHODS

We genotyped 317,503 single-nucleotide polymorphisms (SNPs) in a combined case-control study of 1522 case subjects with rheumatoid arthritis and 1850 matched control subjects. The patients were seropositive for autoantibodies against cyclic citrullinated peptide (CCP). We obtained samples from two data sets, the North American Rheumatoid Arthritis Consortium (NARAC) and the Swedish Epidemiological Investigation of Rheumatoid Arthritis (EIRA). Results from NARAC and EIRA for 297,086 SNPs that passed quality-control filters were combined with the use of Cochran-Mantel-Haenszel stratified analysis. SNPs showing a significant association with disease (P<1×10-8) were genotyped in an independent set of case subjects with anti-CCP-positive rheumatoid arthritis (485 from NARAC and 512 from EIRA) and in control subjects (1282 from NARAC and 495 from EIRA).

RESULTS

We observed associations between disease and variants in the major-histocompatibility-complex locus, in PTPN22, and in a SNP (rs3761847) on chromosome 9 for all samples tested, the latter with an odds ratio of 1.32 (95% confidence interval, 1.23 to 1.42; P = 4×10-14). The SNP is in linkage disequilibrium with two genes relevant to chronic inflammation: TRAF1 (encoding tumor necrosis factor receptor-associated factor 1) and C5 (encoding complement component 5).

CONCLUSIONS

A common genetic variant at the TRAF1-C5 locus on chromosome 9 is associated with an increased risk of anti-CCP-positive rheumatoid arthritis.

BACKGROUND

Rheumatoid arthritis is a chronic inflammatory disease with a substantial genetic component. Susceptibility to disease has been linked with a region on chromosome 2q.

METHODS

We tested single-nucleotide polymorphisms (SNPs) in and around 13 candidate genes within the previously linked chromosome 2q region for association with rheumatoid arthritis. We then performed fine mapping of the STAT1-STAT4 region in a total of 1620 case patients with established rheumatoid arthritis and 2635 controls, all from North America. Implicated SNPs were further tested in an independent case-control series of 1529 patients with early rheumatoid arthritis and 881 controls, all from Sweden, and in a total of 1039 case patients and 1248 controls from three series of patients with systemic lupus erythematosus.

RESULTS

A SNP haplotype in the third intron of STAT4 was associated with susceptibility to both rheumatoid arthritis and systemic lupus erythematosus. The minor alleles of the haplotype-defining SNPs were present in 27% of chromosomes of patients with established rheumatoid arthritis, as compared with 22% of those of controls (for the SNP rs7574865, P = 2.81×10-7; odds ratio for having the risk allele in chromosomes of patients vs. those of controls, 1.32). The association was replicated in Swedish patients with recent-onset rheumatoid arthritis (P = 0.02) and matched controls. The haplotype marked by rs7574865 was strongly associated with lupus, being present on 31% of chromosomes of case patients and 22% of those of controls (P = 1.87×10-9; odds ratio for having the risk allele in chromosomes of patients vs. those of controls, 1.55). Homozygosity of the risk allele, as compared with absence of the allele, was associated with a more than doubled risk for lupus and a 60% increased risk for rheumatoid arthritis.

CONCLUSIONS

A haplotype of STAT4 is associated with increased risk for both rheumatoid arthritis and systemic lupus erythematosus, suggesting a shared pathway for these illnesses.