Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.785803
Title: The aetiology of pathogenic mutations in acute myeloid leukaemia
Author: Md Ahid, Mohd Fadly
ISNI:       0000 0004 7971 2974
Awarding Body: Newcastle University
Current Institution: University of Newcastle upon Tyne
Date of Award: 2019
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Abstract:
Relapse in acute myeloid leukaemia (AML) is driven by additional cooperating somatic mutations that were acquired (or selected for) after chemotherapy, where the relapsing clone evolves from a cell carrying leukaemia-initiating genetic lesions, including gene fusions such as RUNX1/ETO. Some fusion genes are thought to confer a mutator phenotype that predisposes cells to the acquisition of cooperating mutations. As such, it is important to understand how chemotherapy and fusion gene expression contribute to mutagenesis in relapsing AML. The effect of chemotherapeutic agents on the mutation frequency was determined at the thymidine kinase (TK) and hypoxanthine-guanine phosphoribosyltransferase (HPRT) loci in a fully controlled cell model system using TK6 human lymphoblastoid cell lines. Both daunorubicin and cytarabine were mutagenic to DNA at the TK and HPRT loci. Comparison of the daunorubicin-treated and vehicle-treated mutational spectra at the HPRT coding region revealed a significant increase in large deletions in daunorubicin-treated cells relative to vehicletreated cells, demonstrating daunorubicin as a powerful mutagen, inducing almost exclusively gene deletions presumably via strand break induction. The effect of RUNX1/ETO fusion gene expression on mutation frequency was also determined both spontaneously and after chemotherapy treatment using a cell line transduced with the full-length RUNX1/ETO fusion gene. RUNX1/ETO significantly increases spontaneous mutation frequency at both TK and HPRT loci; however, there is no strong evidence that RUNX1/ETO fusion gene expression sensitises cells to chemotherapy-induced mutation. Interrogation of the spontaneous HPRT base substitution spectrum revealed that RUNX1/ETO significantly increases T:A > G:C transversions in vitro, and particularly at the central base position of 5'ApTpA3' / 5'TpApT3' sequences, although this observation was not evident in primary t(8;21) AML. A flow cytometric method was established to evaluate the effect of RUNX1/ETO fusion gene expression on mutation frequency at specific base residues in an AML relevant gene. RUNX1/ETO significantly increase the type A exon 12 NPM1 mutation frequency over time, suggesting this fusion protein predisposes cells to the acquisition of somatic mutation at loci relevant to leukaemogenesis. Relapsed AML is associated with the acquisition of additional somatic mutations which are thought to drive phenotypic adaptability driving clonal selection during treatment and evolution of leukaemic clones. Analysis of high-throughput exome ii sequencing of a small cohort of matched presentation and relapse AML samples revealed clonal evolution patterns in AML, characterised by the continuous acquisition of additional somatic mutations during disease progression and provided important insight on the clonal origins of relapsed AML. Data generated from these studies demonstrate the in vitro mutagenicity of chemotherapeutic agents used in AML remission induction treatment. Furthermore, RUNX1/ETO fusion gene expression increases spontaneous mutation rate, including mutation at the NPM1 locus. However, there is insufficient evidence from the induced mutation spectrum to discern the underlying mechanism of RUNX1/ETO-driven mutagenesis. Taken together, these data inform on the aetiology of somatic mutations in AML.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.785803  DOI: Not available
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