Identifying mechanisms involved in the resistance of homologous recombination defective Dictyostelium cells to ART inhibitors

Thesis

Poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi) are toxic to cells with defects in homologous recombination (HR), which is exploited in the clinic as therapy of BRCA1/2-mutated tumours. Although the concept of synthetic lethality between PARPi and HR deficiency is well-known, the exact mechanisms resulting in cell death are still unclear. Here, we establish Dictyostelium as model to investigate the molecular basis of synthetic lethality of HR dysfunction with PARPi. We show that disrupting the HR-critical factor exonuclease I (exo1) sensitises Dictyostelium cells to PARPi. We further identify that HR inhibition is toxic for cells disrupted in the single-strand break responding PARP Adprt2, whereas an additional loss of the non-homologous end-joining (NHEJ)-promoting PARP Adprt1a renders cells resistant to the treatment. Accordingly, a defect in NHEJ suppresses PARPi toxicity to HR-defective exo1^- strains which is not dependent on the alternative NHEJ (alt-NHEJ) factors Lig3 and PolQ but instead due to re-activation of HR. Furthermore, inhibiting Mre11 re-sensitises dnapkcs^-exo1^- cells to PARPi, indicating that redundancy between nucleases induce PARPi resistance. To identify further mechanims that suppress PARPi toxicity to HR-defective cells, we perform a screen to select for PARPi-resistant exo1^- cells. Using whole genome sequencing we detect mutations that might be accountable for the tolerance of HR-compromised cells to PARPi. Taken together, our data indicate that NHEJ drives the toxicity of PARPi in the absence of HR and that its disruption results in PARPi resistance due to nuclease redundancy, which restores HR.

Authors: Kolb, A

• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford