MOLECULAR MEDICINE: Found in Translation

Abstract

Studies of the major hemoglobin disorders, β-thalassemia and sickle cell disease (SCD), have laid a foundation for molecular medicine. While enormous progress has been made in understanding gene structure and regulation, translating molecular insights to therapy for the many individuals affected with these disorders has been challenging. Advances in three activities have recently converged to bring novel genetic and potentially curative treatments to clinical trials. First, improved lentiviral vectors for gene transfer into hematopoietic stem cells have revived somatic gene therapy for blood disorders. Second, elucidation of regulatory factors and mechanisms that control the normal developmental switch from fetal to adult hemoglobin has provided a route to reactivation of the fetal form for therapy. Third, revolutionary methods of gene engineering permit molecular insights to be leveraged for patients. Here I review how the promise of molecular medicine to bring transformative treatments to the clinical arena is finally being realized.

Figure 1.. The fetal-to-adult hemoglobin switch

The critical transition from HbF(α₂γ₂) to HbA (α₂β₂) entails a transcriptional switch from engagement of the locus control region (LCR) with the γ-globin genes to the β-globin gene. Two repressor proteins BCL11A and LCR (ZBTB7A) initiate the switch by blocking γ-globin transcription and preventing access by the LCR. Repression of γ-globin gene expression allows interaction of β-globin gene with the LCR and transcription activation.

Figure 2.. Mechanism of γ-globin gene repression

The sequence of the γ-globin gene promoter is depicted. The DNA-binding motifs of the two repressors LRF/ZBTB7A and BCL11A are highlighted in blue and red, respectively, with base substitutions observed in patients with HPFH noted below the sequence. Both BCL11A and LRF physically interact with the multicomponent NuRD complex. Binding of BCL11A and NF-Y, an activator that binds a CCAAT box (underlined in yellow) that overlaps a proximal BCL11A motif, oppose each other’s action. BCL11A displaces NF-Y by steric hindrance in the promoter. The boxed region above the sequence indicates a 13bp deletion observed in HPFH and also generated frequently upon CRISPR/Cas9 editing of the promoter . Figure modified from Orkin and Bauer .

Figure 3.. Genetics of BCL11A and HbF output

The consequences of genetic alterations at the BCL11A locus for HbF expression are summarized. The panels depicting common low and high HbF genotypes at the BCL11A locus illustrate the effect of common genetic variation, as measured in GWAS, on BCL11A expression and HbF expression. Natural variants at BCL11A lie within the erythroid-specific enhancer of the BCL11A gene and determine the level of transcription and output of BCL11A protein. The “high HbF” allele is associated with reduced BCL11A protein and a modest increase in HbF. Deletion of the erythroid-specific enhancer nearly abolishes BCL11A expression, leading to marked increase in HbF, as shown in the third panel. Therapeutic gene editing of the erythroid enhancer, specifically at the “Achilles heel” region, greatly reduces BCL11A expression but not to the extent of the enhancer deletion. Nonetheless, the level of HbF expression is significantly increased and is sufficient to ameliorate the severity of either β-thalassemia or SCD. Figure modified from Hardison and Blobel .