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Triple‑negative breast cancer (TNBC) is an aggressive malignancy with limited treatment options and a poor prognosis. The present study investigated the anti‑TNBC effects and underlying mechanisms of cepharanthine (CEP), an isoquinoline alkaloid derived from Stephania cephalantha. The findings showed that CEP inhibited colony formation and induced apoptosis in TNBC cells. Mechanistic investigations revealed that CEP upregulated the pro‑apoptotic protein phorbol‑12‑myristate‑13‑acetate‑induced protein 1 (NOXA), downregulated the anti‑apoptotic protein Bcl‑2 and reduced the mitochondrial membrane potential (ΔΨm). Using quantitative proteomics and limited proteolysis‑coupled mass spectrometry, the present study demonstrated that CEP bound directly to the lysosomal enzymes cathepsin B and cathepsin D, thereby impairing their maturation and suppressing lysosomal degradation. This inhibition triggered the nuclear accumulation of transcription factor EB (TFEB), a factor that regulates the expression of Bcl‑2 family members. These findings indicated that CEP induced apoptosis by inhibiting lysosomal function and activating TFEB, leading to the upregulation of NOXA and downregulation of Bcl‑2. In conclusion, the present study demonstrated the pro‑apoptotic effect of CEP in TNBC cells and identified lysosomal enzymes as the direct target for its mechanism of action. These findings provided a foundation for further investigation of the pharmacological mechanisms of CEP.