Abstract 3568: Ferroptosis induced by xCT abrogation is dependent on GPX4 protein expression regulated by L-cystine/cysteine metabolism

Abstract

Abstract Nutrients in the tumor microenvironment play an essential role in tumor development and proudly impact cancer metabolism. The xCT antiporter, which mediates cellular uptake of L-cystine coupled with glutamate export, has been considered as a promising therapeutic target due in part to its high expression in cancer cells to promote L-cystine uptake as a precursor for glutathione (GSH) synthesis to maintain redox homeostasis. Although inhibition of xCT has been shown to have anticancer effect in multiple models, emerging evidence also reveals that blocking xCT function could confer survival advantages to cancer cells. These opposite observations suggest that xCT inhibition could be a double-edged sword, and its impact on the overall outcome (cancer-inhibitory or tumor-promoting) is likely context-dependent via yet unknown mechanisms. Here, we show that the expression of glutathione peroxidase 4 (GPX4) is a key molecular determinant that decides the cell fate when xCT is suppressed. In cells with low or no expression of GPX4, inhibition of xCT causes severe ROS stress leading to massive cell death via ferroptosis. In contrast, high expression of GPX4 confers cancer cells the ability to escape ROS-induced ferroptosis. Mechanistic study suggests that the intracellular L-cysteine plays a major role in regulating GPX4 protein level, likely via direct binding to GPX4 and stabilize the protein. Pharmacological and genetic disruption of xCT in cancer cells induces dramatic ferroptosis in culture conditions lacking L-cysteine, whereas the presence of L-cysteine in culture or in vivo renders cancer cells resistance to xCT abrogation. Importantly, genetic knockout of xCT by CRISPR/cas9 reduces cancer cell proliferation in culture but promotes tumor growth in vivo, suggesting that the metabolic changes induced by xCT disruption favor cell growth in vivo. Biochemical and molecular analyses show that the in vivo microenvironment containing L-cysteine enables the xCT-disrupted cells to maintain a high GPX4 level, and thus prevents ferroptosis. Furthermore, metabolic flux using U-13C5 glutamine reveals that xCT abrogation leads to intracellular accumulation of more glutamate, which promotes cancer cell survival and proliferation in the low glucose tissue microenvironment in vivo. Our study has identified GPX4 as a key molecule that determines cell fate under metabolic stress induced by xCT suppression, and suggests that modulation of L-cystine/cysteine metabolism might be an effective strategy to overcome ferroptosis-resistant cancer cells. Citation Format: Yanyu Zhang, Peng Huang. Ferroptosis induced by xCT abrogation is dependent on GPX4 protein expression regulated by L-cystine/cysteine metabolism [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3568.