Abstract

Abstract 2641

Abstract:

Only few genes appear to strongly regulate HbF levels in adults with sickle cell disease (SCD). We aim to: (1) Extend these observations to children with SCD, who likely have better preserved marrow capacity; (2) Assess whether these same genes and other previously identified candidates (Ma et al., 2007) associate with HbF response to HU.

Methods:

We performed a retrospective analysis from 6 sites (see author affiliations) of children age 5–21 with HbSS or HbS-B thalassemia, untreated with HU or treated for > 6 months at comparable indications and dosing, using %HbF at steady state (baseline) and on HU at or near maximal tolerated dose (MTD), defined as >20mg/kg/day. Subject adherence to HU was assessed by report to their hematology clinician. Siblings were excluded to ensure genetic independence. Candidate 36 SNPs from 2 groups of genes were genotyped: 1) those from reported GWAS: 15 SNPs in BCL11A, 3 in HBS1L-MYB intron, 5' site in B globin, plus sar1; and 2) 15 candidate SNPs exhibiting the largest effect size on HbF with HU treatment (Ma, 2007). SNP genotyping (minor allele frequency (MAF) ranging from 0.10 to 0.50) was performed on the Sequenom MassArray iPLEX platform. (SNP sequences are available.) Duplicate samples assured genotype concordance. Genotype frequency distribution at each SNP was tested for deviations from Hardy-Weinberg equilibrium. MAFs were comparable across our 6 sites, to allele frequencies in HapMap for CEU populations, and CSSCD (Lettre et al., 2008), confirming validity of pooling SNP genotype data from the sites. Using HbF as a continuous trait, genetic associations were assessed from a total of 80 children, 29 of whom are on HU, between each of the 36 SNPs and: a) baseline %HbF; b) %HbF on HU treatment; c) delta %HbF (HU treatment - baseline). For each model, linear regression analysis was used to test quantitative trait and disease trait SNP associations assuming an additive effect for each copy of the minor allele on the phenotype. Resultant p-values were assessed for significance using Bonferroni adjustment for multiple testing.

Results:

Of the 80 children, comparing the 51 not on HU to those 29 on HU, no significance differences were seen in the distribution and average of baseline %HbF (9.2 vs. 8.9, p=0.820). SNP analyses are summarized in Table 1. 8 SNPs were nominally significantly associated with baseline %HbF. Direction of SNP association differed among these SNPs; some MAF may be reversed in this population compared to those previously reported. For %HbF on HU, the B globin SNP was significantly associated. The delta %HbF on HU is significantly associated with the B globin SNP and nominally so with BCLA11 and SAR1A gene. Our preliminary data begin to extend findings of specific genetic variants regulating HbF to children with SCD. Early data suggest that HbF in response to HU may share some of the mechanisms governing baseline HbF in SCD, not surprising given the commonality of enhanced erythropoiesis. Subject recruitment and analyses are on-going.

Table 1.

SNP associations: baseline %HbF, %HbF on HU, delta %HbF on HU.

%HbF baseline%HbF on HUDelta %HbF
ChrSNPGeneReferenceNβPNβPNβP
rs243027 BCL11A Sedgewick, 2008 80 −0.03 0.015 43 −0.01 0.628 29 −0.03 0.187 
rs243081 BCL11A Menzel, 2007 80 −0.02 0.018 43 −0.02 0.294 29 −0.02 0.384 
rs6729815 BCL11A Nuinoon, 2010 80 0.00 0.870 43 0.03 0.258 29 −0.05 0.046 
rs9399137 HBS1L-MYB Lettre, 2008 80 0.04 0.031 43 0.04 0.453 29 0.05 0.439 
rs487278 PDE7B Ma, 2007 80 0.02 0.038 43 0.00 0.822 29 0.01 0.653 
rs3889124 SOX17 Ma, 2007 80 −0.02 0.020 43 −0.01 0.813 29 0.01 0.791 
rs765587 TOX Ma, 2007 80 0.03 0.026 43 0.01 0.660 29 0.01 0.475 
10 rs4282891 SAR1A Ma, 2007 78 0.05 0.012 41 −0.08 0.144 28 0.16 0.045 
11 rs7482144 β-globin Lettre, 2008 80 0.04 0.043 43 0.13 0.015 29 −0.20 0.009 
%HbF baseline%HbF on HUDelta %HbF
ChrSNPGeneReferenceNβPNβPNβP
rs243027 BCL11A Sedgewick, 2008 80 −0.03 0.015 43 −0.01 0.628 29 −0.03 0.187 
rs243081 BCL11A Menzel, 2007 80 −0.02 0.018 43 −0.02 0.294 29 −0.02 0.384 
rs6729815 BCL11A Nuinoon, 2010 80 0.00 0.870 43 0.03 0.258 29 −0.05 0.046 
rs9399137 HBS1L-MYB Lettre, 2008 80 0.04 0.031 43 0.04 0.453 29 0.05 0.439 
rs487278 PDE7B Ma, 2007 80 0.02 0.038 43 0.00 0.822 29 0.01 0.653 
rs3889124 SOX17 Ma, 2007 80 −0.02 0.020 43 −0.01 0.813 29 0.01 0.791 
rs765587 TOX Ma, 2007 80 0.03 0.026 43 0.01 0.660 29 0.01 0.475 
10 rs4282891 SAR1A Ma, 2007 78 0.05 0.012 41 −0.08 0.144 28 0.16 0.045 
11 rs7482144 β-globin Lettre, 2008 80 0.04 0.043 43 0.13 0.015 29 −0.20 0.009 
Disclosure:

Off Label Use: Hydroxyurea has not been FDA approved for use in children with sickle cell disease, a topic of the submitted abstract.

Author notes

*

Asterisk with author names denotes non-ASH members.