MAP4K4 inhibition enhances the anti-ferroptosis effect of the FTO/YAP/ATF4 axis in doxorubicin-induced cardiotoxicity

BACKGROUND: Doxorubicin (DOX) is a highly effective chemotherapeutic agent, but its clinical application was constrained by irreversible cardiotoxicity. Increasing evidence indicates that ferroptosis is a central mechanism of DOX-induced cardiotoxicity (DIC). Both the m⁶A demethylase FTO and yes-associated protein (YAP) are found to suppress ferroptosis in DIC. Importantly, both YAP and its upstream negative regulator, MAP4K4, are potential targets of FTO. However, whether FTO regulates DOX-induced cardiac ferroptosis via the MAP4K4/YAP signaling axis remains unexplored. METHODS: FTO and YAP expressions were analyzed in DOX-treated cardiac tissues and cardiomyocytes. The effect of FTO on ferroptosis, as well as on the expression of YAP, ATF4, and MAP4K4, was evaluated in DOX-treated mouse heart and cardiomyocytes using cardiac-specific FTO AAV delivery or lentiviral transfection. Pharmacological inhibition of YAP, ATF4, or MAP4K4 was further performed to investigate the underlying mechanisms. In addition, the m⁶A modifications of FTO on MAP4K4 and YAP transcripts were examined. RESULTS: FTO and YAP were significantly reduced in DOX-treated cardiac tissues and cardiomyocytes. FTO overexpression markedly attenuated DIC by inhibiting ferroptosis by enhancing YAP and ATF4 expressions. Inhibition of YAP or ATF4 significantly abolished the protective and anti-ferroptotic effects of FTO. FTO also increased MAP4K4 expression, whereas MAP4K4 inhibition further enhanced the anti-ferroptotic effects mediated by the FTO/YAP/ATF4 axis. Mechanistically, FTO promoted m⁶A demethylation of MAP4K4 and YAP transcripts and enhanced their mRNA stability. CONCLUSIONS: FTO protects against DIC by inhibiting ferroptosis via up-regulation of the YAP/ATF4 axis, and inhibition of MAP4K4 further enhances the FTO/YAP/ATF4-mediated cardioprotective and anti-ferroptotic effects.