Gamma gene expression in haemoglobin disorders

Transcription

1 Gamma gene expression in haemoglobin disorders Innovative therapies for Red Cell and Iron related disorders EHA / ESH : April 16-18, Cascais, Portugal Swee Lay Thein King s College London School of Medicine / King s College Hospital London, UK

2 Outline Conditions associated with increased HbF (α2γ2) hereditary persistence of fetal haemoglobin (HPFH) Pancellular and heterocellular forms HbF persistence in adults as a quantitative trait Dissection of genetic architecture underlying variability of HbF persistence Plausible mechanisms for increased HbF expression Implications for β haemoglobin disorders

3 Hereditary Persistence of Fetal Haemoglobin ( HPFH ) genetically determined persistence of fetal haemoglobin production into adult life in the absence of any haematological disorder Pancellular or heterocellular, based on HbF distribution among erythrocytes

4 Deletions of β globin gene complex kb ε Gγ Aγ ψβ δ β % HbF in heterozygotes β thalassaemia 2-8 δβ fusion Lepore δ thalassaemia Corfu γβ fusion Kenya G γ A o γ (δβ) thalassaemia 8-18 G γ ( Aγδβ) o thalassaemia G γ A o γ (δβ) HPFH (ε Gγ A o γδβ) thalassaemia (Bill Wood, WIMM, Oxord )

5 Non-deletion HPFH Tunisian C % Black -202 C -> G % Greek -196 C -> T 8.6 % Black/Sardinian -175 T -> C % Japanese -114 C -> T % Australian -114 C -> G 9 % Gγ Black -202 C -> T % British -198 T -> C 3-10 % Greek -201 C -> T 10.2 % Brazilian -195 C -> G 5-7 % Black -175 T -> C Italian/Chinese -196 C -> T 12-21% % Greek/Black -117 G -> A % Black -114 to -102 deleted % Aγ Georgia -114 C -> T 3-5 %

6 Mendelian forms of HPFH are rare and do not explain common HbF variation in healthy adults or in patients with sickle cell disease or β thalassaemia Underlying background of variable HbF persistence in adults Persistence of HbF production inherited as a quantitative trait (QT) Twin studies show that the trait is largely genetically controlled (heritability =0.89)

7 HBB cluster on chromosome 11p and Xmn1 Gγ site. α 2G γ 2 ; α 2A γ 2 (HbF) α 2 δ 2 (HbA 2 ) α 2 β 2 (HbA/HbS) HBE HBG2 HBG1 HBBP1 HBD HBB (pseudogene) β globin haplotype Hinc II Xmn I Hind III Hind III Hinc II Hinc II Ava II Hpa I Bam HI HbS -Senegal HbS -Benin HbS Bantu (CAR) HbS -Arab/Indian Swiss Type HPFH NA

8 Chr.6q23.3 The BCL11A gene on chromosome 2p15 chromosome Mb 60.5 Mb 60.6 Mb 3 BCL11A 5 3 adjacent sequences Intron 2 British European Sardinian European GWAS GWAS African-American and African-Brazilian Sickle Patients African British Sickle Patients Chinese and Thai β Thalassaemia Heterozygotes Replicate Replicate Replicate

9 Chr.6q23.3 The HBS1L-MYB locus on chromosome 6q23 chromosome 6 ALDH8A1 HBS1L MYB AHI1 PDE7B Mb Mb HMIP 3 (23 kb) HBS1L HMIP 2 (24 kb) HMIP 1 (7.5 kb) MYB ex 3 ex 2 ex 1 ex 1a ex 1 ex 2 ex 3 European Twins Jamaican Jamaican African haplotypes African - American sickle patients Chinese β thal Carriers

10 BCL11A Stage-specific repressor of γ globin gene? High F BCL11A genotype associated with reduced BCL11A expression Analysis of BCL11A expression BCL11A acts as a silencer of γ-globin gene expression, based on modulation of BCL11A levels Bcl11a / mice fail to silence the expression of human γ-globin genes. Sankaran et al., Science 322, (2008) Sankaran et al., Nature 2009;460:

11 The HBS1L MYB intergenic region contains distal regulatory elements ALDH8A1 HBS1L MYB AHI1 PDE7B Mb Mb DNase I HS sites K562 HS1 HS2 HS3 Cons. GATA-1 motifs Eryth GATA-1 HMIP-1 HMIP-2 HMIP-3 RNAPII AcH3 Transcripts HBS1L Intergenic activity MYB Wahlberg et al Blood 2009;114:1254

12 Impact of HbF QTLs on β haemoglobin disorders

13 Impact of HbF QTLs on β haemoglobin disorders XmnI-Gγ, HBS1L-MYB and BCL11A effect present in Asian Indians, Chinese, Thais and Sardinians with/without β thalassaemia, HbE/β thal, African- Americans, Brazilians and African-British with sickle cell anaemia In African-American sickle patients, the 3 major QTLs account for up to 20% HbF variation with corresponding reduction in acute pain rate γ globin (?%) 6q (3-7%) Other factors (80%) BCL11A (7-12%) Sardinia : BCL11A, HBS1L-MYB and α thalassaemia account for 75% of variable phenotypic severity in beta thalassaemia Thailand : BCL11A, HBS1L-MYB and XmnI-Gγ associated with both disease severity and HbF levels in HbE/β thal (Thein et al PNAS 2007; Menzel et al Nat. Genet 2007; Uda et al PNAS 2008; Lettre et al PNAS 2008; So C-C et al J. Med. Genet. 2008; Sedgewick et al Blood Cells Molecules 2008; Galanello et al Blood 2009;Nuinoon et al Hum Genet 2010)

14 Plausible mechanisms of HbF modulation: 1. BCL11A - Direct effect on γ globin gene expression inhibition of γ globin gene expression 2. HBS1L MYB intergenic region: Alteration of kinetics of erythroid maturation and differentiation, mimicking stress erythropoiesis with release of early erythroid progenitors that synthesise more HbF Challenge: 1. Delineate causal variant(s), physiological pathways involved in increased γ globin gene expression 2. Identify other QTLs to improve prediction of ability to produce HbF; and molecular network of different HbF-modifying loci

15 Conclusions and implications for therapy 1. Molecular basis for persistent HbF expression in adults extremely heterogeneous 2. Mendelian forms deletion HPFH or γ promoter mutations clearly identified genotypically and phenotypically. No unifying simple explanation for increased HbF in those caused by deletions 3. Common variation (heterocellular HPFH) contributed by several linked and unlinked loci, three major QTLs - XmnI-Gγ, HMIP on 6q23, and BCL11A account for 50% of F cell variance. Increased HbF appears to involve multiple pathways. 4. Current pharmacological induction involves multiple pathways, including acceleration of erythroid maturation and differentiation and offer proof-ofprinciple of mechanism underlying disease 5. Downregulating BCL11A expression for therapeutic reactivation of HbF is an attractive option 6. Elucidation of HbF genetics allows improved prediction of ability to produce HbF : prediction of disease severity, genetic counselling and PND.