Investigating the molecular mechanisms of fraction size sensitivity in irradiated cells
Thesis or Dissertation
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Fractionated radiotherapy is used clinically when it spares healthy tissue relative to the cancer; the healthy tissue is described as more fraction size sensitive. However, a molecular understanding of the mechanisms that determine this sensitivity are limited. The recently demonstrated response of breast and prostate cancers to hypofractionation highlights the need to understand the mechanisms of fraction size sensitivity in order to improve dose regimens. We must also develop biomarkers and drugs that leverage this sensitivity to individualise and improve radiotherapy outcomes. DNA double-strand breaks (DSB) are the most deleterious form of damage caused by irradiation due to their potential for misrepair. The cell cycle stage influences the availability of DSB repair pathways. There is a tight inverse association between fraction size sensitivity and proliferation rate. We hypothesise that the enhanced fidelity of Homologous Recombination (HR) repair in S/G2 phase decreases fraction size sensitivity through a reduction of misrepair. Using a non-cancerous repair-proficient fibroblast model, we show that fraction size sensitivity is comparable across all cell cycle phases by clonogenic survival. Chromosome aberration analyses are consistent with survival and suggest that mis-repair events in G1 and G2 phase cells are spared by fractionation. We conclude that the availability of HR in G2 does not impact fraction size sensitivity. Using y-H2AX foci as a surrogate for DSBs we show that induction and kinetics of repair do not alter between fractions, however after complete repair persistent foci are shown to increase with dose and are spared with fractionation, suggesting a role for unrepaired DSBs. The chromatin environment can impact repair pathway choice, we demonstrate through global changes to chromatin state with histone deacetylase inhibition (HDAC) and CRISPR-Cas9 guided BRG1 mutations that these changes do not impact fraction size sensitivity. Finally, a single cell sequencing approach begins to establish a mutational signature for irradiation.
Radiotherapy - Methods
Clinical Academic Radiotherapy (Yarnold)
License start date
Institute of Cancer Research (University Of London)