A genome-wide association study of total bilirubin and cholelithiasis risk in sickle cell anemia

Abstract

Serum bilirubin levels have been associated with polymorphisms in the UGT1A1 promoter in normal populations and in patients with hemolytic anemias, including sickle cell anemia. When hemolysis occurs circulating heme increases, leading to elevated bilirubin levels and an increased incidence of cholelithiasis. We performed the first genome-wide association study (GWAS) of bilirubin levels and cholelithiasis risk in a discovery cohort of 1,117 sickle cell anemia patients. We found 15 single nucleotide polymorphisms (SNPs) associated with total bilirubin levels at the genome-wide significance level (p value <5 × 10(-8)). SNPs in UGT1A1, UGT1A3, UGT1A6, UGT1A8 and UGT1A10, different isoforms within the UGT1A locus, were identified (most significant rs887829, p = 9.08 × 10(-25)). All of these associations were validated in 4 independent sets of sickle cell anemia patients. We tested the association of the 15 SNPs with cholelithiasis in the discovery cohort and found a significant association (most significant p value 1.15 × 10(-4)). These results confirm that the UGT1A region is the major regulator of bilirubin metabolism in African Americans with sickle cell anemia, similar to what is observed in other ethnicities.

Figure 1. Plot of serum bilirubin among 90 sibling pairs in the CSSCD (A) and 200 pairs of unrelated individuals randomly selected from the CSSCD (B).

In each scatter plot, the x- and y-axes show levels of total bilirubin. The correlation coefficient in the 90 sibling pairs was 0.27, while the average correlation coefficient of bilirubin levels in the pairs of unrelated individuals was −0.002.

Figure 2. Summary of the GWAS data from the CSSCD Cohort.

The Manhattan plot (A) displays the –log10(p value) of the associations tested in the CSSCD cohort using the additive model. Color bands represent chromosomes, and SNPs are ordered by their physical position within each chromosome. The large spike in chromosome 2 corresponds to the UGT1A1, UGT1A3, UGT1A8 and UGT1A10 regions. The QQ-plot (B) displays the observed (y-axis) versus expected (x-axis) –log10 (p-value). From the QQ plot, there is minimal to no inflation in the test statistic.

Figure 3. LD Structure in the CSSCD Cohort.

LD plots for regions in genes UGT1A1, UGT1A3, UGT1A4, UGT1A5, UGT1A6, UGT1A7, UGT1A8, UGT1A9 and UGT1A10 on chromosome 2 in the CSSCD subjects. The LD plot was generated using Haploview 4.2. Each diamond represents the D’ value between two SNPs. The LD color scheme is: white D’<1 and LOD<2, blue D’ = 1 and LOD<2; shades of pinkish-red D’<1 and LOD≥2 and bright red D’ = 1 and LOD≥2.