Results
IDX enhances the cytotoxicity of cisplatin against NPC cell lines
The human parental NPC cell lines C17, C666 and EBV-positive/negative NPC cells (NPC43+ve/ NPC43-ve) used in this study were all positive for ENOX2 expression (Supplementary Figure 1A and B). We first determine the drug concentrations that caused inhibition of 50% cell viability (IC50). IDX inhibited proliferation of NPC43-ve, NPC43+ve, C17 and C666 cells over 120 hours with IC50 values ranging between 2-4 µM (Figure 1A); however, NPC43-ve showed higher resistance to IDX treatment. Based on this initial data, doses ranging from 1-4 µM were selected and used to further assess the antitumor effects on cell migration and apoptosis. Cancer cells treated with IDX showed significantly reduced migration compared with untreated cells, in a dose-dependent manner (Figure 1B). Likewise, IDX dose-dependently increased late-stage apoptotic cells compared with the untreated control. (Figure 1C and D). The cis demonstrated diverse cytotoxicity with IC50 values ranging from 0.2-1.2 µM in the four tested cell lines (Figure 1E). In contrast to IDX, NPC43-ve cells demonstrated pronounced sensitivity, suggesting that cis might be more effective in killing EBV-negative cells. A combination cytotoxicity study evaluated the interaction between IDX (1, 2 and 4 µM) and cis (0.6, 1.2 and 2.6 µM). A combination of IDX and cis enhanced the inhibition of cell proliferation (Figure 1F), with minimal impact observed when cells were co-treated with lowest dose of cis (0.6 µM). Similarly, Figure 1G demonstrated that cell growth was not significantly impacted in cells treated with either cis at 0.6 µM or IDX at 1 µM, while cis at 4 µM exhibited far greater inhibition on cell growth. We therefore selected a combination treatment of cis 1.2 µM with IDX at 2 or 4 µM resulting in ~10-fold and 20-fold reductions in cell growth when compared to IDX and cis treatments alone, respectively (Figure 1H). The combination treatment also resulted in a reduction in NPC43-ve and NPC43+ve cell migration by 6-fold (Supplementary Figure 1C).