Synergistic Combination of Hypericin-Mediated Photodynamic Therapy and HIF-1α siRNA Inhibits Triple-Negative Breast Cancer via Inducing PANoptosis in vitro.

BACKGROUND: Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer associated with high global morbidity and mortality. Hypericin is a natural photosensitizer that can preferentially accumulate in tumor tissues. Photodynamic therapy (PDT) is an emerging anticancer strategy that eliminates tumor cells through reactive oxygen species (ROS) generation by light-activated photosensitizers in the presence of molecular oxygen. However, PDT-induced oxygen consumption may further aggravate tumor hypoxia. In this study, we evaluated the synergistic antitumor effects of hypericin-mediated PDT combined with siHIF-1α (HYP-siHIF) in TNBC cells. METHODS: The MTT assay was performed to evaluate the effects of HYP-PDT alone or in combination with siHIF on TNBC cell viability. Intracellular ROS levels were assessed using the fluorescent probe DCFH-DA. RNA sequencing (RNA-seq) analysis was performed to explore the potential molecular changes induced by HYP-siHIF treatment. Transmission electron microscopy (TEM) and Hoechst staining were used to evaluate cellular morphological alterations, while flow cytometry was performed to assess apoptosis. Western blotting was used to detect the expression levels of proteins associated with cell death pathways. RESULTS: HYP-siHIF significantly inhibited TNBC cell viability, increased intracellular ROS accumulation, and induced morphological and molecular features associated with apoptosis, pyroptosis, and necroptosis. Mechanistically, this study provides evidence that HYP-siHIF activates PANoptosis-related cell death pathways following PDT treatment. For apoptosis, HYP-siHIF increased Bax expression, reduced Bcl-2 expression, and enhanced the cleavage of caspase-9/3/7. For pyroptosis, HYP-siHIF increased NLRP3 expression and promoted the cleavage of GSDMD and GSDME. For necroptosis, HYP-siHIF significantly increased the phosphorylation levels of RIP3 and MLKL. CONCLUSION: HYP-PDT and siHIF exhibit synergistic antitumor effects, which are associated with the activation of PANoptosis-related cell death pathways. HYP-siHIF may represent a potential therapeutic approach for TNBC that warrants further mechanistic and translational investigation.