Identification of Potential Biomarkers for Breast Cancer Based on Salivary Metabolomics

Xinyu Jiang^1*Yumei Jia¹Bo Zhang¹Kai Yang¹lijiang yang¹Yang Li^2*

• ¹Hunan University of Chinese Medicine, Changsha, China
• ²Hunan Cancer Hospital, Changsha, China

Abstract: Introduction: Breast cancer (BC) remains the most prevalent malignant tumor in women worldwide and a leading cause of cancer-related mortality. Early screening is essential to improve prognosis, yet current diagnostic methods are often invasive or lack sensitivity. Saliva is an accessible and non-invasive biofluid containing various metabolites that reflect systemic physiological and pathological changes. Thus, salivary metabolomics may provide novel insights into breast cancer-associated metabolic alterations and support the development of early diagnostic strategies. Objectives: To explore the salivary metabolomic profile of breast cancer patients and identify potential non-invasive biomarkers for early breast cancer screening. Methods: Saliva samples were collected from a screening set consisting of 30 BC patients and 20 normal controls (NC) volunteers. untargeted metabolomics approach was performed using liquid chromatography–tandem mass spectrometry (LC-MS/MS). Principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA), along with KEGG pathway enrichment and receiver operating characteristic (ROC) curve analyses, were employed to characterize metabolic differences and identify potential biomarkers. Additionally, saliva samples from a validation set (52 BC patients and 52 NC volunteers) were collected. Enzyme-linked immunosorbent assay (ELISA) was used to quantify potential biomarkers, and their diagnostic performance was evaluated through ROC curve analysis. Results: A total of 101 differential metabolites were identified, including 81 upregulated and 20 downregulated compounds. Screening identified 2-aminonicotinic acid and theobromine as potential diagnostic biomarkers. Analysis of the validation set demonstrated that 2-aminonicotinic acid (AUC: 0.81, cut-off: 5.88 ng/mL) and theobromine (AUC: 0.75, cut-off: 5.27 ng/mL) exhibit promising diagnostic potential. Conclusion: The salivary metabolome of breast cancer patients displays distinct changes compared to healthy individuals. Salivary 2-aminonicotinic acid and theobromine emerge as promising non-invasive biomarkers for breast cancer detection. Nevertheless, larger-scale validation studies are warranted to substantiate their specificity and clinical utility.