Cancer chemopreventive properties of orally bioavailable flavonoids--methylated versus unmethylated flavones

Abstract

Poor oral bioavailability has been a major limitation for the successful use of dietary flavonoids as cancer chemopreventive agents. In this study, we examined fully methylated flavones as promising improved agents. In the human oral SCC-9 cancer cells, 5,7-dimethoxyflavone and 5,7,4'-trimethoxyflavone were both 10 times more potent inhibitors of cell proliferation (IC(50) values 5-8 microM) than the corresponding unmethylated analogs chrysin and apigenin. Flow cytometry indicated that both methylated flavones arrested the SCC-9 cells in the G1 phase with a concomitant decrease in the S phase, dramatically different from the unmethylated analogs, which promoted G2/M phase arrest. Both methylated compounds inhibited the proliferation of two other cancer cell lines with very little effect on two immortalized normal cell lines. Examination of additional flavone structures indicated that methylated flavones in general have antiproliferative properties. Finally, we demonstrated that 5,7-dimethoxyflavone, in contrast to its unmethylated analog chrysin, was well absorbed and had high oral bioavailability as well as tissue accumulation in vivo in the rat. Thus, fully methylated flavones appear to have great potential as cancer chemopreventive/chemotherapeutic agents, in particular in oral cancer.

Fig. 1

Structures of flavones used in this study.

Fig. 2

Effect of the methylated flavones 5,7-DMF and 5,7,4’-TMF compared to the unmethylated analogs chrysin and apigenin on SCC-9 cell proliferation. Cell proliferation, expressed as percent of control (DMSO-treatment), was measured as BrdU incorporation into cellular DNA after a 24-h exposure of the cells to the flavones. Mean values ± SEM are shown (n = 10). The numbers shown in the figure are the calculated IC₅₀ values. * significantly lower than control, P < 0.05. # significantly higher than control, P < 0.05.

Fig. 3

SCC-9 cell uptake of chrysin and 5,7-DMF. The cell monolayers were incubated with 25 μM flavones in complete medium for various time. * significantly higher than 0.5, 2 and 6 hr uptake, P < 0.05; (n = 6).

Fig. 4

Effect of 5,7-DMF (●) compared to chrysin (□) on the proliferation of cancer cells (A and B) and noncancer cells (C and D). A, FaDu cells;B, MCF-7 cells;C, HET-1A cells and D, BEAS-2B cells. Cell proliferation, expressed as percent of control (DMSO-treatment) was measured as BrdU incorporation into cellular DNA after a 24-h exposure of the cells to the flavones. Mean values ± SEM are shown (n = 10). The numbers shown in the figure are the calculated IC₅₀ values.

Fig. 5

Effect of 5,7-DMF (A) compared to chrysin (B) and 5,7,4’-TMF (C) compared to apigenin (D) on SCC-9 cell cycle progression. The cells were exposed to varying concentrations of the flavones for 48 h. The percentage of cells in G1, S and G2/M phase was measured by flow cytometry after propidium iodide staining. Mean values of 3 experiments with duplicate samples are shown. * significantly different from control, P < 0.05 or better.

Fig. 6

Plasma and tissue levels of 5,7-DMF and chrysin after oral administration of 5 mg/kg in rats. A. Plasma 5,7-DMF (no chrysin could be detected at any time-point);B. 5,7-DMF in post-absorption tissues, liver (○), lung (■), and kidney (Δ) (chrysin was barely detectable in a few animals and tissues);C. 5,7-DMF (□) and chrysin (●) in the proximal 2 cm of the colon with associated fecal pellet. The data represent the mean ± SEM of 5 animals at each time-point.