Assessing Cell Activities rather than Identities to Interpret Intra-Tumor Phenotypic Diversity and Its Dynamics

Despite advances in single-cell and molecular techniques, it is still unclear how to best quantify phenotypic heterogeneity in cancer cells that evolved beyond normal, known classifications. We present an approach to phenotypically characterize cells based on their activities rather than static classifications. We validated the detectability of specific activities (epithelial-mesenchymal transition, glycolysis) in single cells, using targeted RT-qPCR analyses and in vitro inductions. We analyzed 50 established activity signatures as a basis for phenotypic description in public data and computed cell-cell distances in 28,513 cells from 85 patients and 8 public datasets. Despite not relying on any classification, our measure correlated with standard diversity indices in populations of known structure. We identified bottlenecks as phenotypic diversity reduced upon colorectal cancer initiation. This suggests that focusing on what cancer cells do rather than what they are can quantify phenotypic diversity in universal fashion, to better understand and predict intra-tumor heterogeneity dynamics.