Essential metals such as Fe play a key role in cancer cell proliferation and metastasis. The novel dual conjugate chelator deferasirox N-ethyleneamine triapine (DefNEtTrp) exploits this Fe-dependency of cancer to fight the disease by sequestering intracellular labile Fe and inducing both apoptosis and ferroptosis. Previous work has confirmed DefNEtTrp to show broad spectrum anticancer activity and display selectivity for cancer cells. This study further characterizes the therapeutic efficacy and mechanisms of action of DefNEtTrp by assessing intracellular metalation of the compound, serum albumin docking for solubility and drug delivery, and the prominence and utility of off-target interactions with Cu. To investigate intracellular interactions with Fe and Cu, proteomic profiling and fluorescence microscopy were employed. Western blotting quantified changes in Fe- and Cu-handling proteins in A549 lung cancer cells treated with DefNEtTrp, correlating such variation with altered intracellular metal levels. Fluorescence microscopy mapped drug uptake and metalation within cells. DefNEtTrp was also docked on bovine serum albumin (BSA) and screened against cancer and healthy cells to assess its compatibility with a carrier and cytotoxicity in the bound form. Finally, ESI-MS, UV-vis, FT-IR spectroscopy, and elemental analysis characterized its ability coordination Cu(II) ions at both the deferasirox and triapine moieties in a tridentate fashion. Results indicate that DefNEtTrp interacts intracellularly with both Fe and Cu, docks on the drug-binding site Sudlow Site II of BSA while retaining cytotoxic activity, and is capable of coordinating Cu(II) to form a complex with independent anticancer potential. These findings highlight DefNEtTrp as a multifunctional therapeutic with bi-metal intracellular chelation activity and carrier-mediated delivery capabilities.
