Abstract
PURPOSE: While combination BRAF and MEK inhibitor treatment in BRAF (V600E) -mutant cancers results in a response, treatment resistance and toxicity are common. Ferroptosis is an iron-dependent form of non-apoptotic cell death. BRAF inhibition has been associated with increased sensitivity to ferroptosis that is dependent on Glutathione Peroxidase 4 (GPX4). EXPERIMENTAL DESIGN: In vitro , ex vivo , and in vivo models of anaplastic thyroid cancer (ATC) were used to evaluate the anticancer activity of combination BRAF inhibition and ferroptosis induction. RESULTS: Targeting key regulators of ferroptosis-GPX4, using RSL3 and ML162, and system X (c) (-) , using erastin-induced significant cell death in all ATC cell lines. Combination of dabrafenib and RSL3 synergistically increased cell death in BRAF (V600E) -mutant ATC cells, and significantly inhibited cellular migration and colony formation. Mechanistically, lipid peroxidation, reactive oxygen species levels, and intracellular Fe (2+) increased significantly with combination treatment compared with each agent alone. Analysis of cell membrane iron importers and exporters showed significantly lower expression of ferroportin-1 (an iron exporter), suggesting the synergistic anticancer activity was due to increased iron accumulation and oxidative stress, leading to enhanced ferroptotic cell death. BRAF (V600E) -mutant ATC cell spheroids showed synergistic cell death with dabrafenib and RSL3 treatment. In vivo , combination dabrafenib and ferroptosis induction (by targeting GPX4 using C18, and system X (c) (-) with IKE) significantly inhibited tumor growth in an orthotopic ATC mouse model. Additionally, dabrafenib-resistant BRAF (V600E) -mutant ATC cells were more sensitive to ferroptosis induction than parental cells. CONCLUSIONS: Dual targeting of BRAF (V600E) and ferroptosis results in synergistic anticancer activity and overcomes resistance to BRAF inhibition.