Penicillide Triggers Immunogenic Pyroptosis in Tumor Cells via Oxidative Stress/NF-κB/GSDMD Signaling Axis

Area of Science:

• Oncology
• Immunology
• Cell Biology

Background:

• Immunogenic cell death (ICD) converts dying tumor cells into an endogenous vaccine.
• Clinically applicable pyroptosis-inducing agents are scarce.
• Penicillide (PEN), a fungal metabolite, has not been previously examined for anticancer activity.

Purpose of the Study:

• To investigate the anticancer activity of Penicillide (PEN).
• To determine the mechanism of cell death induced by PEN.
• To evaluate the potential of PEN as an immunogenic cell death (ICD)-inducing agent for cancer immunotherapy.

Main Methods:

• Cytotoxicity assays across six human cancer cell lines.
• Electron microscopy to observe cell membrane damage.
• Analysis of the NLRP3/caspase-1/GSDMD axis activation.
• Assessment of oxidative stress markers (GSH, MDA, ROS).
• Evaluation of NF-κB signaling pathway activation.
• Detection of ICD hallmarks (calreticulin exposure, HMGB1 translocation, ATP release).
• In vivo tumor models in Balb/c mice to assess vaccine efficacy.

Main Results:

• PEN induces potent, concentration-dependent cytotoxicity by triggering pyroptosis, not apoptosis, necroptosis, ferroptosis, or autophagy.
• PEN causes cell membrane damage and activates the NLRP3/caspase-1/GSDMD axis.
• PEN induces mitochondrial dysfunction, glutathione depletion, elevated ROS, and activates NF-κB signaling.
• PEN-treated cells exhibit ICD hallmarks and act as an effective prophylactic vaccine in mice, delaying tumor outgrowth and enhancing anti-tumor immune cell infiltration.

Conclusions:

• PEN is a novel pyroptosis-inducing agent with significant anticancer activity.
• PEN couples oxidative stress and NF-κB signaling to elicit pyroptosis and immunogenic cell death.
• PEN demonstrates translational potential for ICD-based cancer immunotherapy by generating anti-tumor immunity.