Genome-Wide DNA Methylation Profiling of Erythroid Progenitors and Its Role in Foetal Haemoglobin Reactivation in Sickle Cell Disease: An Original Epigenetic Therapeutic Target Discovery Study

This dissertation conducts genome-wide DNA methylation profiling of erythroid progenitors to identify novel epigenetic loci regulating foetal haemoglobin (HbF) reactivation in sickle cell disease (SCD), providing an original epigenetic target discovery framework applicable to therapeutic development. HbF elevation is the most clinically validated modulator of SCD severity, but the complete epigenetic regulatory landscape controlling HbF silencing in adult erythropoiesis remains incompletely characterised, particularly in African-ancestry erythroid progenitors. Erythroid burst-forming units were isolated from peripheral blood of 60 HbSS patients stratified by HbF tertile (low below 5%, medium 5 to 15%, high above 15%) and 20 HbAA controls at the University of Nigeria Teaching Hospital and National Hospital Abuja. Reduced representation bisulfite sequencing (RRBS) was performed at an average coverage of 30x. CpG methylation differences were identified by MethylKit and Bismark. Differential methylation regions (DMRs) were integrated with ATAC-seq chromatin accessibility data and H3K27ac ChIP-seq from publicly available erythroid datasets. The dissertation identifies 37 novel DMRs in high-HbF versus low-HbF SCD patients, including three hypermethylated regions in BCL11A enhancer elements not previously linked to HbF regulation. An original Erythroid HbF Methylation Atlas for Nigerian SCD patients is constructed. CRISPR-dCas9-TET1 targeted demethylation at the top novel DMR induced a mean 5.8 percent HbF increase in ex vivo erythroid differentiation experiments, providing proof-of-concept for targeted epigenetic therapeutic strategy. Keywords: DNA methylation, foetal haemoglobin, erythroid progenitors, epigenetics, sickle cell disease.