Genetic modifiers of Hb E/β0 thalassemia identified by a two-stage genome-wide association study

First, we estimated and compared the allele frequencies of approximately 110,000 gene-based single nucleotide polymorphisms (SNPs) in pooled DNAs from different severity groups. The 756 SNPs that showed reproducible allelic differences at P < 0.02 by pooling were selected for individual genotyping.After adjustment for age, gender and geographic region, logistic regression models showed 50 SNPs significantly associated with disease severity (P < 0.05) after Bonferroni adjustment for multiple testing. Forty-one SNPs in a large LD block within the β-globin gene cluster had major alleles associated with severe disease. The most significant was bthal_bg200 (odds ratio (OR) = 5.56, P = 2.6 × 10-13). Seven SNPs in two distinct LD blocks within a region centromeric to the β-globin gene cluster that contains many olfactory receptor genes were also associated with disease severity; rs3886223 had the strongest association (OR = 3.03, P = 3.7 × 10-11). Several previously unreported SNPs were also significantly associated with disease severity.These results suggest that there may be an additional regulatory region centromeric to the β-globin gene cluster that affects disease severity by modulating fetal hemoglobin expression.β-Thalassemia is a genetic disease in which an abnormal β-globin gene results in decreased (β+ thalassemia) or completely absent (β0 thalassemia) production of the normal β-globin chain [1]. The hemoglobin E (HbE) allele, point mutation (G→A) in codon 26 (Glu→Lys) of the β-globin gene, can induce alternative splicing and thus result in decreased β-globin E chains [2,3]. Homozygosity for the HbE allele results in hypochromic microcytosis and minimal anemia, but the compound heterozygous condition, HbE/β0 thalassemia causes a surprisingly variable anemia, ranging from nearly asymptomatic states to severe anemia and transfusion-dependency [4-6]. Hb E/β0 thalassemia is most prevalent in and around Thailand, and with large numbers affected in other countries in S