Gliotoxin triggers cell death through multifaceted targeting of cancer-inducing genes in breast cancer therapy.

Fungal secondary metabolites have a long history of contributing to pharmaceuticals, notably in the development of antibiotics and immunosuppressants. Harnessing their potent bioactivities, these compounds are now being explored for cancer therapy, by targeting and disrupting the genes that induce cancer progression. The current study explores the anticancer potential of gliotoxin, a fungal secondary metabolite, which encompasses a multi-faceted approach integrating computational predictions, molecular dynamics simulations, and comprehensive experimental validations. In-silico studies have identified potential gliotoxin targets, including MAPK1, NFKB1, HIF1A, TDP1, TRIM24, and CTSD which are involved in critical pathways in cancer such as the NF-κB signaling pathway, MAPK/ERK signaling pathway, hypoxia signaling pathway, Wnt/β-catenin pathway, and other essential cellular processes. The gene expression analysis results indicated all the identified targets are overexpressed in various breast cancer subtypes. Subsequent molecular docking and dynamics simulations have revealed stable binding of gliotoxin with TDP1 and HIF1A. Cell viability assays exhibited a dose-dependent decreasing pattern with its remarkable IC50 values of 0.32, 0.14, and 0.53 μM for MDA-MB-231, MDA-MB-468, and MCF-7 cells, respectively. Likewise, in 3D tumor spheroids, gliotoxin exhibited a notable decrease in viability indicating its effectiveness against solid tumors. Furthermore, gene expression studies using Real-time PCR revealed a reduction of expression of cancer-inducing genes, MAPK1, HIF1A, TDP1, and TRIM24 upon gliotoxin treatment. These findings collectively underscore the promising anticancer potential of gliotoxin through multi-targeting cancer-promoting genes, positioning it as a promising therapeutic option for breast cancer.