How does fenbendazole induce pyroptosis in breast cancer cells?

Based on the provided sources, fenbendazole (FBZ) induces pyroptosis—a highly inflammatory form of programmed cell death—in breast cancer cells through the p53/HK2/caspase-3/GSDME signaling pathway.

Here is the step-by-step breakdown of this mechanism:

1. Inhibition of Glycolysis via the p53/HK2 Axis

Fenbendazole targets the metabolic reprogramming of breast cancer cells by acting on the p53-glycolysis axis. FBZ significantly upregulates the p53 tumor suppressor protein, which subsequently downregulates the expression of several critical glycolytic enzymes. The most potent inhibition occurs with Hexokinase 2 (HK2), a critical enzyme for tumor metabolism. By inhibiting HK2, fenbendazole severely suppresses aerobic glycolysis, resulting in reduced glucose consumption, decreased lactate production, and lowered ATP (energy) levels in the cancer cells.

2. Activation of Caspase-3 and GSDME

The disruption of glycolysis and the inhibition of HK2 serve as a trigger that activates the caspase-3/GSDME cascade. Fenbendazole treatment significantly activates caspase-3. Once activated, cleaved caspase-3 acts on Gasdermin E (GSDME), cleaving it into its active N-terminal fragment (GSDME-NT).

3. Membrane Pore Formation and Morphological Changes

The newly formed active GSDME fragments drive the execution phase of pyroptosis by forming pores in the cell membrane. This causes the breast cancer cells to undergo hallmark pyroptotic morphological changes, including severe cell swelling, membrane rupture, and the formation of large blisters.

4. Release of Pro-Inflammatory Cytokines

As the cell membrane is ruptured by GSDME pores, the dying cancer cells rapidly release intracellular contents into the surrounding microenvironment. This includes a significant release of lactate dehydrogenase (LDH) and mature pro-inflammatory cytokines, specifically IL-18 and IL-1β.

A Note on Cell Death Plasticity

Researchers found that GSDME is the key trigger for this specific type of cell death. When GSDME is deliberately knocked down or silenced in laboratory settings, the release of inflammatory markers (LDH, IL-18, IL-1β) drops significantly. Interestingly, turning off this pyroptotic pathway does not stop fenbendazole from killing the breast cancer cells; instead, the drug demonstrates “plasticity” by compensating and triggering standard apoptosis (non-inflammatory cell death) to eliminate the tumor cells.