Multi-Algorithm Machine Learning Benchmarking for Pan-Cancer Classification from Tumour-Educated Platelet RNA Sequencing

Tumour-educated platelets (TEPs) carry cancer-type-specific RNA signatures accessible through whole-blood RNA sequencing, but systematic multi-algorithm benchmarking with quantified statistical uncertainty had not been applied to the GSE68086 dataset, the fields primary reference cohort. We applied an end-to-end transcriptomic and machine learning framework to 280 whole-blood platelet RNA-seq samples from six cancer types (non-small cell lung cancer, colorectal cancer, glioblastoma multiforme, hepatobiliary cancer, breast cancer, and pancreatic cancer) and healthy donors. After a standardised preprocessing and normalisation pipeline, seven supervised classifiers - Logistic Regression, SVM (RBF), XGBoost, LightGBM, Random Forest, K-Nearest Neighbours, and a Multilayer Perceptron were benchmarked using stratified 5-fold cross-validation and a held-out test set. Statistical uncertainty was quantified via 2,000-resample percentile bootstrap confidence intervals. Multinomial Logistic Regression achieved the highest test macro F1-score (0.522) and macro-averaged ROC-AUC (0.869), both substantially above the seven-class chance level (1/7 {approx} 0.14). SHAP analysis of the Random Forest classifier identified IFITM3 as the globally dominant TEP biomarker; cancer-type-specific discriminators included ATP5PD (hepatobiliary cancer), C6orf62 (NSCLC and pancreatic cancer), VPS13C (healthy donors), and TMSB4Y (breast cancer). Gene Ontology and KEGG pathway enrichment corroborated the biological specificity of identified transcriptomic signatures. These results support the diagnostic potential of TEP transcriptomics as a multi-class liquid biopsy platform and provide a methodologically transparent, reproducible reference framework for future blood-based cancer classification studies.