Abstract 1855: NUC-1031 overcomes resistance associated with gemcitabine in cancer patients

Abstract

Abstract NUC-1031 (NuCana plc) is a phosphoramidate transformation of gemcitabine and the first anti-cancer compound based on ProTide technology (Slusarczyk et al., 2014). It overcomes the main resistance mechanisms associated with gemcitabine and is currently in Phase I, II and III trials for biliary, ovarian, and pancreatic cancers respectively. High expression of dCK is required for activation of gemcitabine, which competes directly with deoxycytidine (dC) as a substrate for dCK. We aimed to compare the mode of action of NUC-1031 with gemcitabine. A panel of pancreatic and ovarian cancer cell lines were treated for 2h to 24h with NUC-1031 or gemcitabine at the EC50 dose and were observed for 96h. Cell morphology and death were observed using time-lapse microscopy. Cell cycle analysis was performed using BrdU labeling and flow cytometry. Exogenous dC was added to the cells either when they were plated or at the time of the treatment, to observe the effect on the sensitivity of cells to NUC-1031 and gemcitabine. Variation of expression of dCK was determined using RT-qPCR and western blotting. Cytospin, followed by immunocytochemistry, was performed to determine the intracellular localization of dCK. Expression of dCK was assessed in tissue samples from patients in the clinical studies using a modified Allred scoring system. After 48h, most cells treated with NUC-1031 show a 24h period when they neither divide nor die, after which they then undergo apoptosis. By contrast, cells treated with gemcitabine start to die sooner, showing features of necrosis and apoptosis. A higher proportion of cells treated with NUC-1031 exhibited a G2/M arrest compared to those treated with gemcitabine or control samples. Protein expression of dCK was increased in MiaPaCa2 cells soon after treatment with gemcitabine compared with NUC-1031. The pool of endogenous nucleotides was altered by addition of dC, inducing significant resistance to gemcitabine, both when dC was added at the time of the treatment and when cells were plated (EC50 respectively 2.5 and 9 times higher than in control cells). NUC-1031 was associated with a moderate decrease in sensitivity when dC was added at the time of the treatment and when cells were plated (EC50 3 times higher than in control cells). Immunohistochemistry of 50 primary tumours from patients enrolled in a Phase 1 study for NUC-1031 showed low-level dCK expression in almost all tumours studied. Our findings indicate that NUC-1031 has a more targeted mode of action to induce cell death than gemcitabine. Also, although a small amount of dCK may be required for maximal sensitivity to NUC-1031, dCK levels are not a limiting step for NUC-1031 efficacy. Citation Format: Jennifer Bré, Awa Sarr, Peter Mullen, In Hwa Um, Sarah P. Blagden, Paul A. Reynolds, David J. Harrison. NUC-1031 overcomes resistance associated with gemcitabine in cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1855.