Investigating the effects of PARP and ATR inhibition for the treatment of high-risk neuroblastoma

Abstract

Neuroblastoma (NB) is the commonest extra-cranial malignant solid tumour of childhood and one of the most difficult to cure. Long term survival of high-risk NB (HR-NB) currently remains less than 50% at 5 years despite intensive high-dose multimodal treatment. DNA damage response (DDR) defects are frequently observed in HR-NB including allelic deletion and loss of function mutations in key DDR genes, leading to cell cycle checkpoint dysfunction, and oncogene induced replication stress (RS). Cancer cells with defective cell cycle checkpoint signalling and/or increased oncogene-driven RS are acutely dependent on the DNA damage sensor kinase ATR. The aim of this thesis is to identify determinants of sensitivity to ATR and PARP inhibition, alone and in combination, in preclinical models of NB. The baseline level of RS and activity of DDR kinases ATM and ATR was characterised in a panel NB cell lines as well the mutation and/or deletion status of genes previously reported to be synthetically lethal with PARP or ATR inhibition. MYCN-induced RS resulted in vulnerability to ATR and PARP inhibition, by VE821 and olaparib, respectively. ATM deficiency was also associated with sensitivity to ATR inhibition. VE-821 sensitised NB cell lines to olaparib cytotoxicity, which was synergistic in all cell lines. There was a significant positive correlation between fold sensitisation by VE-821 and MYCN protein expression. To obtain greater mechanistic understanding of the synergy between ATR and PARP inhibition, the effect of dual inhibition on markers of RS, cell cycle checkpoints and DNA repair were measured. VE-821 abrogated olaparib-induced S and G2 checkpoint arrest, increased olaparib-induced markers of RS and reduced homologous recombination repair foci. Taken together the work presented in this thesis provides strong rationale for the introduction of ATR and PARP inhibitors for the treatment of HR-NB exhibiting high levels of RS.