5-Fluorouracil and its active metabolite FdUMP cause DNA damage in human SW620 colon adenocarcinoma cell line.

5-Fluorouracil (5-FU) is an antineoplasic drug widely used to treat cancer. Its cytotoxic effect has been principally ascribed to the misincorporation of fluoronucleotides into DNA and RNA during their synthesis, and the inhibition of thymidylate synthase (TS) by FdUMP (one of the 5-FU active metabolites), which leads to nucleotide pool imbalance. In the present study, we compared the ability of 5-FU and FdUMP to induce apoptosis and to influence the cell cycle progression in human colon SW620 adenocarcinoma cells in regards to their genotoxic and clastogenic activities. Our study demonstrates that 5-FU induces SSB, DSB and apoptosis earlier than FdUMP. Interestingly, while both drugs are able to induce apoptosis, their effect on the cell cycle progression differed. Indeed, 5-FU induces an arrest in G1/S while FdUMP causes an arrest in G2/M. Independently of the temporal difference in strand breaks and apoptosis induction, as well as the differential cell cycle modulation, both drugs presented similar clastogenic effects. The different pattern of cell cycle arrest suggests that the two drugs induce different types of primary DNA lesions that could lead to the activation of different checkpoints and recruit different DNA repair pathways.

Targeted drug delivery to chemoresistant cells: folic acid derivatization of FdUMP[10] enhances cytotoxicity toward 5-FU-resistant human colorectal tumor cells.

Current chemotherapy protocols that include fluoropyrimidines, such as 5-fluorouracil (5-FU), are limited by the development of chemoresistance during the course of treatment. Our laboratory has developed a novel class of fluoropyrimidines, FdUMP[N], that are oligodeoxynucleotides (ODNs) composed of some number, N, of 5-fluoro-2'-deoxyuridine-5'-O-monphosphate (FdUMP) nucleotides. Novel synthetic procedures are described that permit conjugation of folic acid to the 5'-OH of FdUMP[10] via a phosphodiester linkage using automated synthesis. The synthetic methods developed are generally applicable for ODN conjugation with folic acid. The folic acid conjugate FA-FdUMP[10] showed improved cytotoxicity toward human colorectal tumor cells (H630), and 5-FU-resistant colorectal tumor cells (H630-10). Enhanced cytotoxicity was observed for FA-FdUMP[10] relative to nonconjugated FdUMP[10] for cells grown under folate-restricted conditions, consistent with cellular uptake being, in part, receptor-mediated. Folate receptor alpha (FRalpha) mRNA was shown by RT-PCR to be overexpressed 26.3-fold in 5-FU-resistant H630-10 cells relative to H630 cells. Thus, FA-FdUMP[N] may prove useful for the treatment of 5-FU-resistant malignancies.

The novel heterodinucleoside dimer 5-FdU-NOAC is a potent cytotoxic drug and a p53-independent inducer of apoptosis in the androgen-independent human prostate cancer cell lines PC-3 and DU-145.

BACKGROUND: We analyzed the cytotoxic properties of the new heterodinucleoside phosphate dimer 5-FdU-NOAC, which is composed of the cytotoxic drugs 5-FdU and N(4)-octadecyl-1-beta-D-arabinofuranosylcytosine (NOAC) against human prostate tumor cells. METHODS: 5-FdU-NOAC effects on cell proliferation, cell cycle distribution, thymidylate synthase activity, and apoptosis were investigated in vitro in the two human prostate carcinoma cell lines DU-145 and PC-3 and compared to cells treated with the corresponding single drugs 5-FdU and NOAC. RESULTS: Treatment of the cells with 5-FdU-NOAC resulted in IC(50) values of 3.9-5 microM and in a complete inhibition of cell proliferation at 200 microM after 96 hr compared to 5-FdU, where 10% of the cells remained resistant. Flow cytometric analysis revealed cell cycle perturbations in S-phase only in the DU-145 cells. 5-FdU-NOAC caused 50% inhibition of thymidylate synthase after 90 min at 0.6 microM in both cell lines. Apoptotic cell fractions in DU-145 (66%) and in PC-3 (34%) cells were found after treatment with 5-FdU-NOAC for 96 hr. DNA fragmentation further confirmed the induction of apoptosis. CONCLUSIONS: 5-FdU-NOAC inhibits thymidylate synthase and cell cycle progression causing proliferation arrest and apoptosis in DU-145 and PC-3 cells, suggesting a potential role of 5-FdU-NOAC for the treatment of prostate cancer.

Increased cytotoxicity and decreased in vivo toxicity of FdUMP[10] relative to 5-FU.

The efficacy of treatment with 5-Fluorouracil (5-FU) is limited, in part, by its inefficient conversion to 5-Fluoro-2'-deoxyuridine-5'-O-monophosphate (FdUMP). We present data indicating that FdUMP[10], designed as a pro-drug for intracellular release of FdUMP, is cytotoxic as a consequence of uptake of the multimeric form. FdUMP[10] is stable in cell culture medium, with more than one-half of the material persisting as multimers of at least six nucleotides after a 48 h incubation at 37 degrees C. FdUMP[10] is more than 400 times more cytotoxic than 5-FU towards human colorectal tumor cells (H630). FdUMP[10] also has decreased toxicity in vivo, with doses as high as 200 mg/kg/day (qdx3) administered to Balb/c mice without morbidity, compared to a maximum tolerated dose of 45 mg/kg/day for 5-FU using the same protocol. FdUMP[10] shows reduced sensitivity to OPRTase- and TK-mediated drug resistance, relative to 5-FU and FdU, respectively, and is much more cytotoxic than 5-FU towards cells that overexpress thymidylate synthase. Thus, FdUMP[10] is less susceptible to resistance mechanisms that limit the clinical utility of 5-FU. The increased cytotoxicity, decreased toxicity in vivo, and reduced sensitivity to drug resistance of FdUMP[10], relative to 5-FU, indicates multimeric FdUMP is potentially valuable as an anti-neoplastic agent, either as a single agent, or in combination with 5-FU.

Inhibitory effects of fluorinated pyrimidines, 5'-DFUR, UFT and T-506, in a model of hepatic metastasis of mouse colon 26 adenocarcinoma-assessment of inhibitory activity and adverse reactions at the maximum tolerated dose.

The effects of fluorinated pyrimidines, 5'-DFUR, UFT and T-506, on a mouse model of hepatic metastasis were assessed in regard to inhibitory activity and adverse reactions at the maximum tolerated dose. The model was prepared by injecting the mouse colonic cancer cell line, colon 26, into the portal vein of CDF1 mice. At the treatment regimens employed for 5'-DFUR (1.0 mmol/kg/day, p.o., daily from days 1 to 7), UFT (0.1 mmol/kg/day, p.o., daily from days 1 to 7), and T-506 (0.074 mmol/kg/day, i.v., days 1, 4, 7, and 10), complete inhibition of hepatic metastasis was obtained in six out of seven mice (85.7%) with 5'-DFUR, and in five out of six mice (83.3%) with T-506. Significant inhibition of hepatic metastasis was not achieved with UFT (3/7, 42.9%). 5'-DFUR and T-506 showed the highest rate of inhibition of hepatic metastasis, suggesting that these drugs would be effective for the prophylactic treatment of metastatic disease. 5'-DFUR and UFT exhibited mild adverse reactions such as loss of body weight.