Correlation between membrane proteins and sizes of extracellular vesicles and particles: A potential signature for cancer diagnosis

Extracellular vesicles and particles (EVPs) are recognized as ideal liquid biopsy tools for cancer detection, and membrane proteins are commonly used EVP biomarkers. However, bulk analysis of EVP membrane protein biomarkers typically fails to meet the clinical requirement for diagnostic accuracy. We investigated the correlation between the membrane protein expression level, the binding kinetics to aptamers and the sizes of EVPs with interferometric plasmonic microscopy (iPM), and demonstrated the implementation of the correlative signature to determine cancer types. Using EVPs collected from both cell model and clinical plasma samples with liver, lung, breast, or prostate cancer, we found that the selective set of membrane protein expression levels of five protein markers and their binding kinetics were highly heterogeneous across various sizes of EVPs, resulting in the low overall accuracy (<50%) in cancer classification with bulk analysis of all populations. By grouping the EVPs into three subpopulations according to their sizes, the overall accuracy could be increased to about 70%. We further grouped the EVPs into subpopulations with a 10 nm interval in sizes and analysed the correlation between the membrane proteins and sizes with a machine learning algorithm. The results show that the overall accuracy to discriminate cancer types could be improved to 85%. Therefore, this work highlights the significance of size‐dependent subtyping of EVPs and suggests that the correlation between the selective set of membrane proteins and sizes of EVP can serve as a signature for clinical cancer diagnosis.