5674

JS-K (O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate) is a nitric oxide (NO)-releasing, glutathione-S-transferase (GST)-activated prodrug. Its potent antineoplastic activity has been documented in vitro and in vivo; however its mechanism(s) of action/molecular target(s) are not completely understood. Growth inhibitory effects of JS-K have been most studied in leukemia and multiple myeloma cells. We investigated the effects of JS-K on growth of a panel of 20 human lung adenocarcinoma cell lines. JS-K inhibited growth of all cell lines with IC50 concentrations ranging from 0.2 to 10 µM. JS-K was most effective against cell lines with high levels of endogenous peroxides, with IC50 values for these cell lines in the range 0.2 - 0.5 µM. Three cell lines were chosen for further studies - H1944, characterized by low level of endogenous peroxides and resistance to JS-K, and H1703 and H1734, with significantly higher levels of peroxides and ten-fold greater sensitivity to JS-K. A 2- to 10-fold increase in intracellular nitric oxide level as a result of JS-K treatment was confirmed in all 3 cell lines using 4-amino-5-methylamino-2',7'-difluorofluorescein (DAF) fluorescence. However, a 2- to 3-fold increase in intracellular ROS/RNS levels, measured by 2',7'-dichlorodihydrofluorescein (DCF) fluorescence after a 30 min treatment with 1 µM JS-K, was observed only for cell lines with high basal level of ROS. The elevated intracellular ROS/RNS stress resulted in DNA damage, with a significant 2-fold increase in DNA strand break damage (comet assay) after a 90 min incubation with 1 µM JS-K and massive DNA damage with 10 µM JS-K. Initiation of apoptosis also occurred, as indicated by activation of caspases 3 and 9 after 24 hours incubation with 1 µM JS-K. The results indicate that JS-K has potential as a therapeutic agent against human lung cancers, especially those that express high intracellular levels of peroxides. Cellular and chemical pathways involved are under study.
 Funded in part by N01-CO-12400

99th AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA