Abstract CT401: NUC-1031: A novel ProTide that overcomes the key cancer resistance mechanisms associated with poor survival

Abstract

Abstract Introduction: Chemotherapeutic resistance is a common issue that severely limits the clinical utility of all nucleoside analogues. The key cellular mechanisms causing resistance are limited expression of activating kinases and nucleoside transporters, and overexpression of catabolising enzymes. Phosphoramidate chemistry has been applied to nucleoside analogues to generate a new class of anti-cancer agents, ProTides, which overcome all these resistance mechanisms. Here, we report potent in vitro and in vivo biological activity of NUC-1031, a gemcitabine-derived ProTide. Methods: The cytotoxicity of the ProTide NUC-1031 and gemcitabine was monitored through in vitro viability assays. Both compounds were tested against a broad range of cancer cell lines including the pancreatic lines MiaPaCa-2 and BxPC-3 (partially or fully resistant to gemcitabine). Mimicking cancer resistance conditions, the activating enzyme, deoxycytidine kinase (dCK) was inhibited with deoxycytidine, and the nucleoside transporter, hENT1, was blocked with dipyridamole. The resulting intracellular concentrations of the active moiety, dFdCTP, were then measured. The ability of gemcitabine and NUC-1031 to resist degradation by cytidine deaminase (CDA) was assessed using absorption spectroscopy. Activity of NUC-1031 and gemcitabine were further evaluated in MiaPaCa-2 and BxPC-3 nude mouse xenograft models. Results: NUC-1031 was more cytotoxic than gemcitabine in vitro, with a lower EC50. NUC-1031 activity was not significantly affected by dCK inhibition (EC50= 0.2nM to 0.7nM in the presence of deoxycytidine) whereas gemcitabine cytotoxicity was reduced by 75 fold (EC50= 1.4nM to 104.9nM). Inhibition of nucleoside transport with dipyridamole significantly decreased gemcitabine cytotoxicity (EC50= 611μM rose to >2000μM). In contrast, NUC-1031 cytotoxic activity remained unchanged and significantly lower than gemcitabine (EC50= 190μM to 162μM). Measured intracellular concentrations of dFdCTP were consistent with these results. Additionally, gemcitabine was catabolized within one minute of exposure to CDA, while NUC-1031 was impervious to deamination. Finally, in pancreatic cancer xenografts, at 11, 14, 21 & 25 days, NUC-1031 showed greater reduction in tumor volumes compared to gemcitabine and control. Conclusions: The novel ProTide, NUC-1031, overcomes the key cancer resistance mechanisms associated with gemcitabine. Notably, NUC-1031 reaches up to 20 fold higher intracellular levels of dFdCTP, even in resistant cells. A Phase I clinical study of NUC-1031 is nearing completion and has provided significant disease control in both patients refractory to, or deemed unsuitable for, gemcitabine therapy. Citation Format: Essam A. Ghazaly, Magdalena Slusarczyk, Malcolm Mason, John Gribben, Christopher McGuigan, Sarah Blagden. NUC-1031: A novel ProTide that overcomes the key cancer resistance mechanisms associated with poor survival. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr CT401. doi:10.1158/1538-7445.AM2014-CT401