Identification of Ferroptosis as a Novel Mechanism for Antitumor Activity of 2-phenylphthalazin-2-iums in Breast Cancer.

Quaternary benzo[c]phenanthridine alkaloids (QBAs), an important class of natural isoquinoline alkaloids, possess potential multiple anti-cancer activities. By structural simulation, 29 2-phenylphthalazin-2-iums with anti-cancer effects were designed and synthesized in our previous study. Breast cancer is one of the most frequent malignant tumors, which is closely related to the high mortality rate in women. The role and underlying mechanism of 2-phenylphthalazin-2-iums in repressing the growth and metastasis of human breast cancer were explored. The role of 2-phenylphthalazin-2-iums in regulating the proliferation and invasion of MCF-7 cells was evaluated in vitro and in a xenograft model. After treatment with 3',5'-diCl isomer (29), the reactive oxygen species (ROS) levels, Fe2+ concentration and glutathione (GSH) contents were assessed to evaluate the effect of the 2-arylphthalazine salts on facilitating ferroptosis in MCF-7 cells. Compound 29 exhibited a dose- and time-dependent cytotoxicity on MCF-7 cells. Compound 29 treatment effectively repressed the growth and metastasis of breast cancer in a xenograft model. We verified for the first time that compound 29 triggered ferroptosis of MCF-7 cells, as evidenced by increased Fe2+ concentration, ROS level, and decreased GSH content. Mechanistically, compound 29 induced oxidative damage and ferroptosis via regulating p53/SALC7A11 axis. Iron chelator deferoxamine restored the proliferation and invasion of MCF-7 cells treated with compound 29 through inhibiting ferroptosis. Significantly, compound 29 inhibited the growth of human breast cancer cells MCF-7-derived tumor xenograft (CDX) models. Thus, 2-phenylphthalazin-2-iums represent promising leads for the development of novel anti-breast cancer agents in the future.