Intracellular Cytidine Deaminase Regulates Gemcitabine Metabolism in Pancreatic Cancer Cell Lines

• Published in: Drug metabolism and disposition Vol. 48; no. 3; pp. 153 - 158
• Main Authors: Bjånes, Tormod K., Jordheim, Lars Petter, Schjøtt, Jan, Kamceva, Tina, Cros-Perrial, Emeline, Langer, Anika, Ruiz de Garibay, Gorka, Kotopoulis, Spiros, McCormack, Emmet, Riedel, Bettina
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.03.2020; American Society for Pharmacology and Experimental Therapeutics, Inc; American Society for Pharmacology and Experimental Therapeutics (ASPET); The American Society for Pharmacology and Experimental Therapeutics
• Subjects: Adenocarcinoma; Adenocarcinoma - metabolism; Biocompatibility; Biotechnology; Cancer; Carcinoma, Pancreatic Ductal - metabolism; Cell activation; Cell Line, Tumor; Cytidine deaminase; Cytidine Deaminase - metabolism; Deactivation; Deoxycytidine - analogs & derivatives; Deoxycytidine - metabolism; Gemcitabine; Human health and pathology; Humans; Hépatology and Gastroenterology; In vivo methods and tests; Inactivation; Intracellular; Life Sciences; Liquid chromatography; Mass spectrometry; Mass spectroscopy; Metabolites; Pancreatic cancer; Pancreatic Neoplasms - metabolism; Pharmaceutical sciences; Pharmacology; Toxicity; Tumor cell lines; dFdCTP; THU; CDA; DMEM; PDAC; hENT1; dFdCMP; dCK; dFdU; dFdC
• ISSN: 0090-9556; 1521-009X; 1521-009X
• DOI: 10.1124/dmd.119.089334

Cytidine deaminase (CDA) is a determinant of in vivo gemcitabine elimination kinetics and cellular toxicity. The impact of CDA activity in pancreatic ductal adenocarcinoma (PDAC) cell lines has not been elucidated. We hypothesized that CDA regulates gemcitabine flux through its inactivation and activation pathways in PDAC cell lines. Three PDAC cell lines (BxPC-3, MIA PaCa-2, and PANC-1) were incubated with 10 or 100 µM gemcitabine for 60 minutes or 24 hours, with or without tetrahydrouridine, a CDA inhibitor. Extracellular inactive gemcitabine metabolite (dFdU) and intracellular active metabolite (dFdCTP) were quantified with liquid chromatography tandem mass spectrometry. Cellular expression of CDA was assessed with real-time PCR and Western blot. Gemcitabine conversion to dFdU was extensive in BxPC-3 and low in MIA PaCa-2 and PANC-1, in accordance with their respective CDA expression levels. CDA inhibition was associated with low or undetectable dFdU in all three cell lines. After 24 hours gemcitabine incubation, dFdCTP was highest in MIA PaCa-2 and lowest in BxPC-3. CDA inhibition resulted in a profound dFdCTP increase in BxPC-3 but not in MIA PaCa-2 or PANC-1. dFdCTP concentrations were not higher after exposure to 100 versus 10 µM gemcitabine when CDA activities were low (MIA PaCa-2 and PANC-1) or inhibited (BxPC-3). The results suggest a regulatory role of CDA for gemcitabine activation in PDAC cells but within limits related to the capacity in the activation pathway in the cell lines. The importance of cytidine deaminase (CDA) for cellular gemcitabine toxicity, linking a lower activity to higher toxicity, is well described. An underlying assumption is that CDA, by inactivating gemcitabine, limits the amount available for the intracellular activation pathway. Our study is the first to illustrate this regulatory role of CDA in pancreatic ductal adenocarcinoma cell lines by quantifying intracellular and extracellular gemcitabine metabolite concentrations.