A549 lung adenocarcinoma cells were treated with vehicle, camptothecins CPT, TPT, 9-CPT, or 10-CPT, the prodrug IRN, its active metabolite SN-38, non-camptothecin GENZ, non-canonical inhibitor SW, or LPO inducer CHP, each at 0.1 μM for 72 h. (a) Active Top1 poisons induce ROS. Live cell imaging of CellROX Deep Red (red) and brightfield (grey). Scale bar = 10 μM. (b) Induction of ROS and LPO by Top1 poisons are linked. Plot displays mean fluorescence intensity for CellROX analysis of ROS (MFI CellROX, mean ± SD) and the fraction of cells probed with BODIPY-C11 LPO sensor that display high fluorescence (% LPO+ cells, mean ± SD) for each Top1 poison (R2 = 0.85). (c) Top1 poisons induce cellular accumulation of HNE. Competitive ELISA of cell lysate with anti-HNE-adduct antibody (HNE adducts (μM), mean ± SD, n = 2). *, P ≤ 0.05 (Kruskal-Wallis). (d) Top1 poisons deplete glutathione (GSH). Assay of GSH in deproteinated cell lysate (GSH (μM), mean ± SD, n = 2). *, P ≤ 0.05 (Kruskal-Wallis). (e) Top1 poisons induce ALDH enzyme activity. AldeRed flow cytometry ALDH activity assay (610 nm). Percent ALDHHI cells shown. See also , , and .