Breast cancer is the most commonly diagnosed cancer worldwide, yet its proteomic profile shows substantial overlap with other cancer types, making differential diagnosis challenging. This study aimed to identify a panel of plasma protein biomarkers capable of distinguishing breast cancer from eleven other cancer types using a publicly available pan-cancer proteomic dataset (Álvez et al., 2023), comprising 1,477 patients and approximately 1,463 proteins measured by the Proximity Extension Assay (PEA).
Three supervised machine learning (ML) models were applied: Elastic Net Regression (ENR), Random Forest (RF), and Support Vector Machine (SVM), to identify the most discriminative proteins for breast cancer classification. Variable importance scores from all three models were combined to generate a consensus-ranked panel of 70 proteins, with five proteins (FLT3, CD244, ENPP2, JCHAIN, and AHSP) consistently identified across all models. Each model was trained and evaluated using four protein subsets (top 10, top 30, top 70, and all proteins) and assessed using accuracy, area under the receiver operating characteristic curve (AUC), sensitivity, and specificity. Model robustness was further evaluated using repeated 10-fold cross-validation (20 repetitions).
Classification performance improved with increasing protein panel size across all models. When all proteins were included, the ENR model achieved the best performance, with an accuracy of 0.72 and an AUC of 0.78 on the independent test dataset. A reduced 30-protein panel retained competitive discriminative ability (AUC = 0.71–0.73), supporting its potential for future targeted assay development. In repeated cross-validation, ENR demonstrated the highest and most stable performance (mean accuracy = 74.2%, mean Kappa = 0.48), outperforming RF. Functional enrichment analysis revealed that the top candidate proteins were enriched in cell adhesion, immune regulation, and IL-17 signaling pathways, consistent with known mechanisms of tumor invasion and immune evasion in breast cancer. However, the identified proteins should be considered candidate biomarkers, and their diagnostic utility requires validation in independent external cohorts and prospective clinical studies.
These findings demonstrate that plasma proteomics combined with ML can distinguish breast cancer from other cancer types in a pan-cancer setting and identify promising candidate proteins for further experimental validation and assessment in future clinical studies.