Cell cycle arrest has a potent suppressive effect on ferroptosis, a form of regulated cell death caused by excessive lipid peroxidation. Mechanistically, cell cycle arrest induces diacylglycerol acyltransferase (DGAT)-dependent lipid droplet formation, which sequesters polyunsaturated fatty acids (PUFAs) that accumulate in arrested cells as triacylglycerols (TAGs). This results in reduced ferroptosis. DGAT inhibition shifts PUFAs from TAGs to phospholipids (PLs), making arrested cells more susceptible to ferroptosis. The study found that slow-cycling, therapy-resistant cancer cells, such as 5-fluorouracil-resistant cells, exhibit increased lipid droplet levels and ferroptosis resistance. Combined treatment with ferroptosis inducers and DGAT inhibitors effectively suppresses the growth of 5-fluorouracil-resistant tumors by inducing ferroptosis. These findings suggest that cell cycle arrest drives ferroptosis resistance and provide a therapeutic strategy to target slow-cycling therapy-resistant cancers.Cell cycle arrest has a potent suppressive effect on ferroptosis, a form of regulated cell death caused by excessive lipid peroxidation. Mechanistically, cell cycle arrest induces diacylglycerol acyltransferase (DGAT)-dependent lipid droplet formation, which sequesters polyunsaturated fatty acids (PUFAs) that accumulate in arrested cells as triacylglycerols (TAGs). This results in reduced ferroptosis. DGAT inhibition shifts PUFAs from TAGs to phospholipids (PLs), making arrested cells more susceptible to ferroptosis. The study found that slow-cycling, therapy-resistant cancer cells, such as 5-fluorouracil-resistant cells, exhibit increased lipid droplet levels and ferroptosis resistance. Combined treatment with ferroptosis inducers and DGAT inhibitors effectively suppresses the growth of 5-fluorouracil-resistant tumors by inducing ferroptosis. These findings suggest that cell cycle arrest drives ferroptosis resistance and provide a therapeutic strategy to target slow-cycling therapy-resistant cancers.