## 3 Results
The least efficient compound was the 2'-hydroxy-3'-methoxy derivative (**2**, Figure 5B). In the case of the 4'-hydroxy-3'-methoxy-5'-nitro (**6**) and 2'-chloro-6'-fluoro derivatives (**12**), the relationship to other compounds was more complicated, particularly in the case of the latter, where the reduction was markedly dependent on pH. The results of the copper reduction experiments are summarized in Figure 6.  
Figure 2: Iron chelation by representative compounds. (**A**): 4′-trifluoromethylomently(**9**), (**B**): 4′-dimethylamino(**8**), (**C**): 4′-ethoxy (**7**), (**D**): 3′′-dihydroxy (**3**) and (**E**): 2′-hydroxy-3′-methoxy (**2**). The graphs show different xanthone:iron ions ratios at different pH conditions. The percentage of chelation on the \(y\) axis was calculated using the positive control sample (solvent with iron ions—100% remaining metal ions and hence 0% chelation). The final concentration of the iron ions was 50 uM in all cases. Figure 4: Iron reduction—representative compounds. **(A)**: \(3^{\prime}A^{\prime}\)-dihydroxy **(3)**, **(B)**: \(4^{\prime}\)-trifluoromethyl **(9)**, **(C)**: \(4^{\prime}\)-dimethylmino **(8)** and **(D)**: \(4^{\prime}\)-acetamido **(10)** derivatives. Results at higher pH (negligible reduction) are not shown. The final concentration of iron ions was 50 \(\upmu\)M in all cases. Figure 3: Summary of the differences in iron chelation among the tested compounds. The direction of the arrows shows more active compounds. The numbers represent pH at which the differences were observed; at pH 4.5, the oxidative state is mentioned in parentheses. As a result of their ability to reduce both metals (see likely scenarios in the Supplementary Data, Figure S8), the xanthones can potentially promote iron- and/or copper-based Fenton reaction and, consequently, increase the cytotoxic effects of these metals. To study the potentially cytotoxic effects toward cancer cells, the most active chelator, the 4'-trifluoromethyl derivate (**9**), was selected and tested on the adenoma breast cancer cell line (MCF7/S0.5) and fulvestrant-resistant-derived cell line (MCF7/182R-6). We selected this couple of cell lines in order to investigate if the activity of this compound is affected by this specific type of resistance, which is a common consequence observed in cancer. Metals and the xanthone derivative were tested over a range of concentrations, alone and also in combination, and the results were compared to the vehicle control (DMSO 0.1%) assigned to 100% viability. The results showed that although Fe\({}^{2+}\) is, as expected, not toxic (IC\({}_{50}\) = 2611 uM), the 4'-trifluoromethyl xanthone derivate possessed toxicity similar to that of Cu\({}^{2+}\); the IC\({}_{508}\) were 218 and 300 uM, respectively (Table 1). Next, we selected the concentration of 500 uM of copper, which was strongly cytotoxic itself, and tested different concentrations of the chelator in order to see the cytotoxicity relationship in relation to the chelator:metal ratio. As shown (Figure 7), the copper  
Figure 5: Copper reduction—examples. (**A**): 4′-trifluoromethyl (**9**) and (**B**): 2′-hydroxy-3′-methoxy (**2**) derivative. The arrows show significance vs. the solvent at corresponding pH. The final concentration of cupric ions was 50 uM in all cases. Figure 6: Summary of the differences in copper reduction among the tested compounds. The direction of the arrows shows more active compounds. The numbers represent pH at which the differences were observed. cytotoxicity was apparently decreased by increasing concentrations of the 4'-trifluoromethyl derivate. The same experiment was also performed with iron. The addition of iron markedly protected cells against chelator toxicity (Figure 7). The same experiments were also performed with MCF/7/182R-6 cells. Here, the IC\({}_{50}\) were, as expected, slightly higher than those for MCF/7/S0.5;. The pattern of results was similar, but the protection against copper was very mild (Supplementary data Figure S9).