Figure 4.

Phendione does not chelate intracellular cations. (A) Quantification of fluorescent intensity in the cells labeled with zinc indicator by flow cytometry. A375 (top) and A375 BRAFi-resistant (bottom) cells were treated with vehicle, phendione or cell-permeable zinc chelator TPEN for 3 h. At the end of treatment, 20 µM zinquin ethyl ester was added into medium to stain Zn²⁺ in live cells. Unstained cells were used as negative control for flow cytometry. (***) P < 0.001 by unpaired two-tailed Student's t-test. (B) Growth inhibition curves of A375 (top) and A375 BRAFi-resistant (bottom) cells treated with phendione or zinc chelator TPEN prepared in regular medium or medium containing 15 µM ZnCl₂. (C) Immunoblot of whole-cell lysate collected after the treatment of vehicle or phendione prepared in regular medium or medium containing 15 µM ZnCl₂ as indicated. (D,E) Growth inhibition curves of A375 (top) and A375 BRAFi-resistant (bottom) cells after 96 h of treatment with phendione or ion chelators DFOM or DFP prepared in regular medium or medium containing 50 µM FeCl₃ (D) or 10 µM CuSO₄ (E).