ENHANCED CYTOTOXICITY OF NUCLEOSIDE ANALOGS BY OVEREXPRESSION OF MITOCHONDRIAL DEOXYGUANOSINE KINASE IN CANCER CELL-LINES

The cytotoxic anti-cancer purine nucleoside analogs 2-chloro-2'-deoxya denosine (CdA), 9-beta-D-arabinofuranosylguanine (araG), and 2',2'-dif luorodeoxyguanosine (dFdG) are phosphorylated by human mitochondrial d eoxyguanosine kinase (dGK) in vitro. We overexpressed dGK as a fusion protein to the green fluorescent protein in the human pancreatic cance r cell lines PanC-1 and MIA PaCa-2 to determine the importance of dGK- mediated nucleoside analog phosphorylation, The transfected cells show ed mitochondrial fluorescence patterns, and the mitochondrial location s of endogenous and overexpressed dGK were verified by Western blot an alysis of cell extracts with polyclonal anti-dGK antibodies. The incre ase of dGR activity in the overexpressing cells was similar to 4-fold. These cell lines exhibited increased sensitivity to CdA, araG, and dF dG as compared with the untransfected parent cell lines. This is, to o ur knowledge, the first demonstration of a correlation between the act ivity of a mitochondrial deoxyribonucleoside kinase and the cytotoxici ty of nucleoside analogs. Our data imply that the dGK activity is rate -limiting for the efficacy of nucleoside analogs in the cell lines inv estigated.