Ferroptosis is a form of regulated cell death resulting from iron accumulation and lipid peroxidation. While impaired ferroptosis is tightly linked to human diseases and conditions, the mechanism and regulation of ferroptosis remain largely unknown. Here, we demonstrate that STAT3 is a positive regulator of ferroptosis in human pancreatic ductal adenocarcinoma (PDAC) cell lines. Activation of the MAPK/ERK pathway, but not inhibition of system Xc(-), was required for STAT3 activation during erastin-induced ferroptosis. Importantly, pharmacological inhibition and genetic silencing of STAT3 through small molecules (e.g., cryptotanshinone and S31-201) or siRNA blocked erastin-induced ferroptosis in PDAC cells. Mechanically, STAT3-mediated cathepsin B expression was required for ferroptosis. Consequently, inhibition of lysosome-dependent cell death by pharmacological blockade of cathepsin activity (using CA-074Me) or vacuolar type H+-ATPase (using bafilomycin A1) limited erastin-induced ferroptosis. These studies indicate that ferroptosis is a lysosomal cell death process. (C) 2018 Elsevier Inc. All rights reserved.