Phytochemical profile and proteomic landscape associated with antineoplastic activity of Zingiber cassumunar Roxb. Essential oil in canine melanoma.

Canine malignant melanoma is an aggressive neoplasm with limited remedial options. Therefore, the development of novel antineoplastic agents is urgently needed. This study investigated the medicinal potential and molecular mechanisms of Zingiber cassumunar Roxb. essential oil (CEO) against canine melanoma. Phytochemical profiling via GC-MS identified 22 compounds, with monoterpenoids (90.99%) predominating, specifically sabinene (53.25%) and terpinen-4-ol (17.99%). CEO exhibited potent, concentration-dependent cytotoxicity across four canine melanoma cell lines, with a mean IC₅₀ of approximately 0.006% v/v. Quantitative proteomic mapping of CEO-treated M5 cells identified a total of 222 differentially expressed proteins associated with systemic cellular collapse. Key mechanisms included significant genotoxic stress (upregulation of H2AX and histone variants), endoplasmic reticulum stress (suppression of CALR and HSPB1), and the activation of a coordinated necroptosis/mitophagy axis (modulation of Drp1, VDAC, and p62). Protein-protein interaction network analysis further identified SLC2A1 as a central metabolic hub with high-confidence connectivity to conventional chemotherapies, suggesting that CEO-induced metabolic disruption converges onto core pathways targeted by 5-fluorouracil and imatinib. These findings suggest that CEO induces complex cell death signatures through coordinated nuclear and organelle stress. Ultimately, it should be considered as a promising candidate for future translational research and functional validation.