Received for publication December 15, 2006.
Revised June 9, 2007.
Accepted for publication June 12, 2007.

The Impact of Hypoxic treatment on the Expression of Phosphoglycerate Kinase (PGK) and the Cytotoxicity of L-OddC (Troxacitabine) and dFdC (Gemcitabine)

Abstract

Beta-L-Dioxolane-cytidine (L-OddC, BCH-4556, Troxacitabine), a novel L-configuration deoxycytidine analog, is under clinical trials for treating cancer. The cytotoxicity of L-OddC is dependent on the amount of the triphoshate form (L-OddCTP) in nuclear DNA. Phosphoglycerate kinase (PGK), a downstream protein of hypoxia-inducible-factor-1 (HIF-1), is responsible for the phosphorylation of the dihosphate to the triphosphate of L-OddC. Here, we studied the impact of hypoxia on the metabolism and the cytotoxicity of L-OddC and dFdC in several human tumor cell lines including HepG2, Hep3B, A673, Panc-1 and RKO. Hypoxic treatment induced the protein expression of PGK 3-fold but had no effect on the protein expression of APE-1, dCK, CMPK and nM23 H1. Hypoxic treatment increased L-OddCTP formation and incorporation of L-OddC into DNA but it decreased the uptake and incorporation of dFdC which correlated with the reduction of hENT1, hENT2 and hCNT2 expression. Using a clonogenic assay, hypoxic treatment of cells made them 2 to 3-fold more susceptible to L-OddC but not to dFdC following exposure to drugs for one generation. Dimethyloxallyl glycine enhanced the cytotoxicity of L-OddC but not dFdC in Panc-1 cells under normoxic conditions. Over-expression or down-regulation of PGK using transient transfection of pcDNA5-PGK, or inducible shRNA in RKO cells affected the cytotoxicity of L-OddC but not that of dFdC. The knockdown of HIF-1 in inducible shRNA in RKO cells reduced the cytotoxicity of L-OddC but not dFdC under hypoxic conditions. In conclusion, hypoxia is an important factor that may potentiate the activity of L-OddC.

Key words: Nucleoside/Nucleotide, Nucleoside/Nucleotide derivatives