Abstract 3624: A novel, selective DNA-PK inhibitor sensitizes poor prognosis chronic lymphocytic leukemia (CLL) cells to mitoxantrone

Abstract

Abstract Approximately 20% of Chronic Lymphocytic Leukaemia (CLL) cases carry 11q deletions and 10% carry 17p deletions (associated with loss of p53 function), both of which are poor prognostic markers. Ataxia telangiectasia mutated kinase (ATM) is located on chromosome 11q23 and 30% of del11q patients lack ATM function. Previous work has shown that the DNA-PK inhibitor NU7441 chemosensitises CLL cells to fludarabine ex vivo (Willmore E et al 2008), and monotherapy with KU60648, a dual inhibitor of PI3kδ and DNA-PK has been shown to be effective in ATM deficient cells (Riabinska A et al 2013). Furthermore, a dual DNA-PK/mTOR inhibitor, CC-115, is now in clinical trials (NCT01353625). Here, we aimed to validate the selective inhibition of DNA-PK over related kinases as a therapeutic approach in CLL, using a novel, more selective compound (NDD0004, cell-free IC50 10nM). Both primary CLL cells and an isogenic pair of DNA-PK-deficient and proficient HCT116 cells were used as models. Primary CLL cells were cultured ex vivo and treated with the topoisomerase II poison mitoxantrone in the presence and absence of NU7441 and NDD0004. Cell viability was determined after 48h by XTT assay. Apoptosis and DNA damage (yH2AX foci) were measured by caspase 3/7GloAssay and immunofluorescence, respectively. ATM function was determined by the cells ability to autophosphorylate ATM on serine 1981 after ionising radiation. Chemosensitisation to mitoxantrone in primary CLL cells was substantial and it was higher with NDD0004 than with NU7441 (mean 202- vs. 69-fold, ranges 1-1000 vs. 1-543-fold). Intriguingly, cells lacking ATM or p53 function were also sensitised by DNA-PK inhibition (mean 15- and 21-fold, respectively). HCT116 cells lacking DNA-PK were not chemosensitised by NDD0004 but DNA-PK proficient cells were sensitised 5-fold. Mitoxantrone-induced DSBs (as measured by yH2AX foci) were repaired by 24h, but persisted in the presence of DNA-PK inhibitors. In keeping with this observation, apoptosis as measured by caspase 3/7 activity was potentiated by DNA-PK inhibitors. NDD0004 and KU60648 were also evaluated in a panel of ATM deficient (HT144, H1395) and ATM proficient (A549, A375) cell lines. ATM status had no impact on (a) the level of chemosensitisation to mitoxantrone by both inhibitors and (b) single agent activity for NDD0004 and KU60648. In conclusion, our data show that selective inhibition of DNA-PK chemosensitises primary CLL cells even when p53 is mutated or ATM function is lost. Current work is focused on identifying biomarkers which predict good response to DNA-PK inhibitors. Citation Format: Gesa Junge. A novel, selective DNA-PK inhibitor sensitizes poor prognosis chronic lymphocytic leukemia (CLL) cells to mitoxantrone. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3624. doi:10.1158/1538-7445.AM2015-3624