Induction of cell cycle-dependent cytotoxicity and apoptosis by new heterodinucleoside phosphate dimers of 5-fluorodeoxyuridine in PC-3 human prostate cancer cells

Fluorodeoxyuridine (5-FdUrd) is an antineoplastic agent with clinical activ ity against different types of solid rumours. To enhance the effectiveness of this drug, we have synthesised new heterodinucleoside phosphate dimers o f 5-FdUrd. These dimers were compared to 5-FdUrd for their cytotoxic effect and the cell cycle dependence of cytotoxicity, as well as for their capaci ty to induce apoptosis and inhibit thymidylate synthetase (TS) in androgen- independent human PC-3 prostate tumour cells. Incubation of the cells with the dimers N-4-palmitoyl-2'-deoxycytidylyl-(3'-->5')-5-fluoro-2'-deoxyuridi ne (dCpam-5-FdUrd) and 2'-deoxy-5-flourouridylyl-(3'-->5')-2'-deoxy-5-fluor o-N-4-octadecylcytidine(5-FdUrd-5-FdC18) resulted in a marked cytotoxicity with IC50 Values of 4 muM, similar to 5-FdUrd. In contrast to 5-FdUrd, 100% toxicity was achieved with concentrations of 100-200 muM 5-FdUrd-5-FdC18. Flow cytometric analysis revealed an increase in the cell population in S-p hase after treatment with 5-FdUrd, 5-FdUrd-5-FdC18, and dCpam-5-FdUrd from 36 to 63%, 50%, and 77%, respectively. dCpam-5-FdUrd was more potent than 5 -FdUrd in arresting the cell cycle. Significant S-phase arrest was indicate d by a decreased proportion of cells in G1- and G2/M-phases. Cell cycle arr est and inhibition of cell proliferation were followed by apoptosis, as sho wn by a 6- to 8-fold increased binding of Apo2.7 antibody, a 9- to 11-fold increase in caspase-3 activity, DNA fragmentation, and by cell morphology s howing the appearance of apoptotic bodies. Importantly, 5-FdUrd-5-FdC18 inc reased the number of apoptotic cells to 160% compared to 5-FdUrd under the same conditions. As with 5-FdUrd, the two dimers also inhibited TS in a tim e- and concentration dependent manner, although requiring 100-fold higher c oncentrations. In conclusion, dCpam-5-FdUrd and 5-FdUrd-5-FdC18 exert stron ger cytotoxicity and induce more S-phase arrest and apoptosis than does 5-F dUrd in PC-3 cells, suggesting their potential role in the treatment of hum an prostate cancer. BIOCHEM PHARMACOL 60;12:1887-1896, 2000. (C) 2000 Elsev ier Science Inc.