Effects of nucleoside analogues, lipophilic prodrugs and elaidic acids on core signaling pathways in cancer cells

Objectives: Nucleoside analogs such as gemcitabine (GEM; dFdC) and cytarabine (Ara-C) require nucleoside transporters to enter cells, and deficiency in equilibrative nucleoside transporter 1 (ENT1) can lead to resistance to these drugs. To facilitate transport-independent uptake, prodrugs with a fatty acid chain attached to the 5’-position of the ribose group of gemcitabine or cytarabine were developed (CP-4126 and CP-4055, respectively). As antimetabolites can activate cellular survival pathways, we investigated whether the prodrugs or their side-chains had similar or decreased effects. Methods: Two cell lines A549 (non-small cell lung cancer) and WiDr (colon cancer cells) were exposed for 2-24hr to IC50 concentrations of GEM, Ara-C, CP-4126, CP4055 and elaidic acid (EA) concentrations corresponding to the CP-4126 and CP-4055 IC50. Cells were harvested and analyzed for proteins in cell survival pathways (p-AKT/AKT, p-ERK/ERK, p-P38/P38, GSK-3β/pGSK-3β) by using Western Blotting. Results: All drugs and their derivatives showed time- and cell-line-dependent effects. In A549 cells, GEM, CP-4126 and EA-4126 decreased the p-AKT/AKT ratio at 2 and 24 hr. For the p-ERK/ERK ratio, GEM, EA-4126, Ara-C, CP-4045 and EA-4055 exposure led to an increase after 6 hr in A549 cells. Interestingly, Ara-C, CP-4055 and EA-4055 decreased p-ERK/ERK ratio in WiDr cells after 4 hr. In A549 cells, the p-GSK-3β/GSK-3β ratio decreased after exposure to Ara-C and CP-4055 but in WiDr cells increased after 24 hr. In A549 cells treatment with Ara-C, CP-4055 and EA-4126 decreased the p-P38/P38 after 6 hr. Conclusions: The findings suggest that both parent drugs, prodrugs, and the EA chain influence cell survival and signaling pathways.