title: The impact of hypoxic treatment on the expression of phosphoglycerate kinase and the cytotoxicity of troxacitabine and gemcitabine.
abstract: beta-L-Dioxolane-cytidine (L-OddC, Troxacitabine, BCH-4556), 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 triphosphate form (L-OddCTP) in nuclear DNA. Phosphoglycerate kinase (PGK), a downstream protein of hypoxia-inducible-factor-1alpha (HIF-1alpha), is responsible for the phosphorylation of the diphosphate to the triphosphate of L-OddC. In this study, we studied the impact of hypoxia on the metabolism and the cytotoxicity of L-OddC and beta-d-2',2'-difluorodeoxycytidine (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 after exposure to drugs for one generation. Dimethyloxallyl glycine enhanced the cytotoxicity of L-OddC but not dFdC in Panc-1 cells under normoxic conditions. Overexpression 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-1alpha 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.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA.
authors.forename: Wing
authors.initials: W
authors.lastname: Lam
authors.forename: Chung-Hang
authors.initials: CH
authors.lastname: Leung
authors.forename: Scott
authors.initials: S
authors.lastname: Bussom
authors.forename: Yung-Chi
authors.initials: YC
authors.lastname: Cheng
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000633:toxicity*; D000970:Antineoplastic Agents / Q000633:toxicity*; D002277:Carcinoma / Q000473:pathology; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000473:pathology; D003596:Cytosine / Q000031:analogs & derivatives* / Q000633:toxicity; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000378:metabolism; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000633:toxicity; D004148:Dioxolanes / Q000633:toxicity*; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology; D010190:Pancreatic Neoplasms / Q000473:pathology; D010735:Phosphoglycerate Kinase / Q000378:metabolism*; D012208:Rhabdomyosarcoma / Q000473:pathology; D000093542:Gemcitabine
chemical_list: D000964:Antimetabolites, Antineoplastic; D000970:Antineoplastic Agents; D004148:Dioxolanes; D003841:Deoxycytidine; C074908:troxacitabine; D003596:Cytosine; D010735:Phosphoglycerate Kinase; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D000093542:Gemcitabine