Quercetin-induced inhibition and synergistic activity with cisplatin - a chemotherapeutic strategy for nasopharyngeal carcinoma cells

Abstract

Background: Nasopharyngeal carcinoma (NPC) is a unique tumour of epithelial origin with a distinct geographical distribution, genetic predisposition and environmental as well as dietary influence as aetiological factors. Standard NPC treatment regimes, such as radiotherapy and concurrent chemotherapy with cytotoxic drugs, can produce undesirable complications often associated with significant toxicity. Here, we report the effects of a widely distributed flavonoid, quercetin, on cell proliferation, apoptosis and cell cycle arrest. The effects of combining quercetin and cisplatin on human NPC cells were explored.

Methods: Cell proliferation was monitored by the dynamic, impedance-based cell analyzer (xCELLigence system) and the MTS assay. Ki67 proliferation antigen and fatty acid synthase (FASN) level was examined by Western blotting. Flow cytometry was also carried out to study the effects of quercetin on cell cycle and apoptosis status.

Results: At 100 μM, quercetin inhibited cell proliferation and decreased expression of FASN and Ki67 antigen. Cell cycle analysis revealed a substantial increase in the proportion of cells in the G2/M phase. We also demonstrated the enhanced cytotoxic effects of quercetin treatment in concomitant with the chemotherapeutic drug, cisplatin, in cultured NPC cells. The combination index (CI) value of quercetin-cisplatin combination was < 1, indicating synergism.

Conclusions: Our study showed that quercetin exhibited synergistic effects with cisplatin against NPC cells. Dose-reduction index (DRI) values > 1 implied the possibility of reducing the cisplatin dosage required to treat NPC, with the addition of quercetin. In turn, this could reduce the risk of cisplatin-associated toxicity. The potential of combining quercetin with cisplatin as a chemotherapeutic strategy for treatment of NPC should be explored further.

Figure 1

Growth curves generated from acquired cell index depicting the effect of quercetin on (A) HK1 and (B) C666-1 cell proliferation. 100 μM quercetin inhibited HK1 and C666-1 cell proliferation. The graph shown is a representative of triplicate experiments.

Figure 2

Immunoblot for FASN and Ki67 24 hours after quercetin treatment. HK1 and C666-1 cells treated with quercetin expressed lower amount of FASN and Ki67 compared to untreated cells. Detection of β-actin served as a loading control. The experiment was repeated twice and consistent results were obtained. A representative result is shown.

Figure 3

Flow cytometry analysis using annexin V-FITC/PI double staining depicting apoptosis. The lower right and upper right quadrants represent cells undergoing apoptosis. Images shown are representatives of three independent experiments. The average percentage ± sd of total apoptotic cells is shown.

Figure 4

Cell cycle analysis by BrdU labelling in untreated and quercetin-treated cells. Increasing percentages of cells in G2/M phase and sub-G₁ peak were observed. A representative of three independent experiments is shown. The average percentage ± sd of total cells in the G2/M phase is shown.

Figure 5

Dose–response curves of quercetin-treated HK1 (A) and C666-1 (B) cells. The average percentage viable ± sd relative to vehicle control cells is shown.

Figure 6

Bar graphs showing viability after cisplatin or simultaneous treatment with cisplatin and quercetin in HK1 (A) and C666-1 (B) cells, determined by the MTS assay. The control group on the extreme left is assigned 100% viability. The combination of quercetin and cisplatin is more active than cisplatin alone. Results are presented as average percentage viable ± sd.

Figure 7

F_a-CI plots for HK1 (A) and C666-1 (B), respectively, showed a synergistic cytotoxic effect for simultaneous treatment of a fixed combination ratio of quercetin and cisplatin. In the F_a-CI plot, the dashed line (CI = 1) indicates an additive reaction between the two substances. Values above and below this dashed line imply antagonism and synergism, respectively.