Nucleoside metabolism
enzymes are determinants of chemotherapeutic
drug activity. The
nucleoside salvage
enzyme deoxycytidine kinase (dCK) activates
gemcitabine (2', 2'-difluoro-2'-deoxycytidine) and is negatively regulated by
deoxycytidine triphosphate (
dCTP). Reduction of
dCTP in
tumor cells could, therefore, enhance
gemcitabine activity. Mitochondrial
thymidine kinase 2 (TK2) phosphorylates
deoxycytidine to generate
dCTP. We hypothesized that: (1) TK2 modulates human
tumor cell sensitivity to
gemcitabine, and (2) antisense knockdown of TK2 would decrease
dCTP and increase dCK activity and
gemcitabine activation.
siRNA downregulation of TK2 sensitized MCF7 and HeLa cells (high and moderate TK2) but not A549 cells (low TK2) to
gemcitabine. Combined treatment with TK2
siRNA and
gemcitabine increased dCK. We also hypothesized that TK2
siRNA-induced
drug sensitization results in mitochondrial damage that enhances
gemcitabine effectiveness. TK2
siRNA and
gemcitabine decreased mitochondrial redox status,
DNA content, and activity. This is the first demonstration of a direct role for TK2 in
gemcitabine resistance, or any independent role in
cancer drug resistance, and further distinguishes TK2 function from that of other
dTMP-producing
enzymes [cytosolic TK1 and
thymidylate synthase (TS)].
siRNA knockdown of TK1 and/or TS did not sensitize
cancer cells to
gemcitabine indicating that, among the 3
enzymes, only TK2 is a candidate therapeutic target for combination with
gemcitabine.