Two are better than one: A case study of the biferrocenyl-thymidine "click" conjugate biFcT as novel ferroptosis-inducing agent in lung cancer cells.

Ferroptosis is one of the programmed cell death pathways. The detailed molecular mechanisms underlying ferroptosis remain a subject of intense study, but it is well established that they involve iron-dependent lipid peroxidation and cell membrane damage. Ferroptosis inducers are attractive candidates for anticancer drugs as they can bypass drug resistance developed by cancer cells against proapoptotic drugs. In this work, we demonstrate that a "click" biferrocenyl-thymidine conjugate (biFcT) induces ferroptosis in lung cancer cells. Its mechanism of action involves reactive oxygen species (ROS) generation, lipid peroxidation, and protein carbonylation. Although biFcT also induces ROS in mitochondria, it does not lead to effector caspase-3 activation and apoptosis. The ferroptosis inhibitor ferrostatin-1 effectively abrogates the anticancer activity of biFcT. Furthermore, we found that biFcT exerts its deleterious activity independently of the cellular content of the selenoprotein glutathione peroxidase-4 (GPX4)-an enzyme that protects cells from ferroptosis. Another therapeutic advantage of biFcT is its selectivity for lung cancer cells over nontumorigenic human bronchial epithelium Beas-2B cells. To the best of our knowledge, this is only the eighth report on ferrocene-containing ferroptosis activators to date and the first to demonstrate their therapeutic potential in lung cancer cells.