Serum Proteomic Analysis Using Gel‐Based Liquid Chromatography Tandem Mass Spectrometry Reveals Differences Between Canine Oral Malignancies and Non‐Malignant Conditions

BACKGROUND: Canine oral cancers are difficult to manage due to complex biology and a lack of non-invasive biomarkers. Proteomic approaches, particularly gel-based liquid chromatography-tandem mass spectrometry (GeLC-MS/MS), have been used on tissue and saliva, but serum remains obscure despite its clinical accessibility and ability to reflect systemic disease. OBJECTIVES: This study evaluated GeLC-MS/MS for serum proteomic profiling in canine oral malignancies, compared to benign and healthy conditions. METHODS: We analysed 62 serum samples from dogs with oral melanoma (OM, n = 28), oral squamous cell carcinoma (OSCC, n = 10), benign tumours (BN, n = 12) and controls (healthy/periodontitis, n = 12) using GeLC-MS/MS-based proteomics. RESULTS: Significant protein expression differences emerged across groups. In OM and OSCC, phosphodiesterase 4D (PDE4D) was upregulated, while ornithine decarboxylase antizyme 3 (OAZ3), centriolar coiled-coil protein 110 (CCP110), non-specific serine/threonine protein kinase 8 (NEK8), receptor-type tyrosine-protein phosphatase F (PTPRF) and interleukin 23 receptor (IL23R) were downregulated. These proteins are linked to critical pathways, including insulin signalling, insulin resistance, adherens junctions and cell cycle regulation, highlighting their roles in cancer progression and showing potential interactions with common chemotherapy drugs like doxorubicin, cisplatin and cyclophosphamide. CONCLUSIONS: This study demonstrates that GeLC-MS/MS-based serum proteomics can successfully identify candidate biomarkers for canine oral malignancies. The discovery of these protein signatures represents promising diagnostic and prognostic targets, with the potential to guide chemotherapeutic selection and improve clinical outcomes in dogs with oral cancer.