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Title: Investigation of the use of histone deacetylase inhibitors for the treatment of inherited disorders of the glycolytic pathway
Author: Makarona, Kalliopi
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2013
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Histone acetylation by histone acetyltransferases (HATs) and deacetylation by histone deacetylases (HDACs) regulate gene expression by activating or repressing transcription, respectively. HDAC inhibitors (HDACIs) are a diverse class of drugs used to treat haemoglobinopathies, urea cycle disorders and several types of malignancies. Recent evidence from genome-wide as well as gene-specific epigenetic studies suggest a model whereby active genes are more likely than silent genes to be hyperacetylated and increase their transcription levels in response to HDACIs, a process underpinned by the dynamic recruitment and antagonistic activities of HATs and HDACs. Based on this model and from a therapeutic perspective, I hypothesised that the ability of HDACIs to increase expression of active genes might be relevant for diseases caused by genes that encode proteins with enzymatic function. HDACI-mediated increase in gene transcription, even in the presence of missense, disease-causing mutations, might lead to increased enzymatic activity and amelioration of the cellular and clinical phenotype. I tested this hypothesis on a group of genes involved in the glycolytic and pentose phosphate pathway (GPPP) which, when mutated, cause chronic or episodic haemolytic anaemia. Using RT-qPCR (B cell lines) and gene expression profiling (primary, in vitro generated human erythroid precursors and CD4+ T cells) I found that of the 17 GPPP genes, only Glucose-6-Phosphate Dehydrogenase (G6PD) mRNA levels increased in response to HDACIs in a time-dependent manner. Epigenetic analysis in B cells by ChIP-qPCR showed that histone hyper-acetylation and increased recruitment of HATs and HDACs underpin the selective G6PD transcriptional activation in response to HDACIs. Pharmacological and genetic assays showed that increase in G6PD transcription was also dependent on Sp1, a generic transcription factor known to recruit both HDACs and HATs. Finally, I directly tested the hypothesis that HDACIs may increase enzymatic activity in G6PD deficient cells. Using B cell lines and primary erythroid cells from patients with G6PD deficiency, I found that HDACIs induce the same epigenetic changes in the mutant as in the wild type G6PD gene; more importantly, they lead to increased levels of the mutant mRNA and protein, associated with an up to 3-fold increase in enzymatic activity. These findings are potentially of great therapeutic significance for correction of G6PD deficiency in up to 300 million individuals worldwide with the polymorphic variants of G6PD deficiency (e.g., G6PDMed and G6PDA-).
Supervisor: Karadimitris, Anastasios Sponsor: Sir Halley Stewart Trust
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available