4,4'-dimethoxychalcone induces thermosensitive impairment of SIRT3-associated mitochondrial signaling and apoptotic cell death with therapeutic potential in cancer.

SIRT3, a key mitochondrial deacetylase that regulates metabolic adaptation, oxidative stress, and apoptosis in cancer cells, has emerged as a promising therapeutic target in cancer treatment. 4,4'-dimethoxychalcone (DMC) is a natural chalcone derivative extracted from Angelica keiskei. In this study, we demonstrated that DMC induces thermosensitive impairment in SIRT3-associated mitochondrial signaling. A non-lethal temperature (42 °C) facilitates the interaction between DMC and the catalytic pocket of SIRT3, leading to preferential suppression of SIRT3 activity and consequently sensitizing HeLa cells to robust apoptotic signaling. This inhibition was accompanied by increased SOD2 acetylation and reduced SOD2 abundance, driving accumulation of oxidative stress, which triggers loss of mitochondrial membrane potential (ΔΨm), and pronounced caspase activation. Concomitantly, γ-H2AX nuclear foci formation and dual-phase cell-cycle arrest at the G1/S and G2/M checkpoints were associated with activation of p53/p21 and ATM/Chk2 signaling. In addition, cell migratory capacity was significantly impaired, as evidenced by reduced expression of MMP2 and MMP9. Notably, this inhibition impairs autophagic and mitophagic flux as well as lysosomal function, thereby preventing cytoprotective responses and facilitating cell death. Importantly, enforced SIRT3 overexpression reverses these effects, restores mitochondrial function, reactivates autophagic activity, and attenuates apoptotic cell death. Collectively, our findings identify DMC as a thermosensitive inhibitor of SIRT3 and provide mechanistic insights into its potential to disrupt mitochondrial homeostasis for targeted cancer therapy.