Bradburn, AK; (2017) Effect of DNA Damage and Repair Mechanisms in Human Haematopoiesis and their role in Chemoresistance of Acute Myeloid Leukaemia. Doctoral thesis , UCL (University College London).

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Abstract

Most standard AML chemotherapy regimes utilize Ara-C, a cytotoxic compound inflicting DNA damage in proliferating cells and inducing cell death. Comparing normal and malignant haematopoietic cells in their response to Ara-C will determine whether differing responses to DNA damage and activation of DNA repair mechanisms contribute to chemoresistance. Moreover, DNA repair in primitive human haematopoiesis and LIC’s is still not well established. Using AML cell lines, cord blood and CD34+ AML patient samples, a characterization of the DNA damage response post Ara-C treatment was performed. AML cell lines were shown to have a range of sensitivities with significant increases in DNA damage after Ara-C exposure. However, cell lines resistant to Ara-C, were quicker to recover their DNA damage, with a faster revival of cell numbers and reduction in apoptosis. Similar findings were also observed when using ionizing radiation as a source of DNA damage. Analysis of the DNA repair mechanisms non-homologous end joining (NHEJ) and homologous recombination (HR) by 53BP1 and RAD51 localization respectively, showed NHEJ was preferentially used in response to Ara-C, with low rates of HR. Normal haematopoietic stem and progenitor populations were compared with leukaemia initiating and tumour populations of CD34+ AML patient samples in vivo. Normal stem populations showed significant increases in apoptosis with enrichment in cells in G0 after exposure to Ara-C. Progenitor cells were more refractory to treatment and displayed a G1/S-phase accumulation. When observing AML patient samples it appeared they either behaved in a progenitor, or a stem like manner, showing heterogeneous responses overall. Just like the AML cell lines NHEJ was primarily observed, with some basal level of activation. Overall there are differences in normal stem progenitor and cells in response to Ara-C, with AML patient samples displaying a more heterogeneous response to DNA damage.

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