Integration of multiple lineage measurements from the same single cell reconstructs parallel tumor evolution

The dynamics of tumorigenesis shares many similarities with Darwinian evolution. Elevated mutation rates result in a genetically diverse population of cells competing for limited resources in the tumor. Natural selection ultimately leads to clonal expansion of the fittest genotypes resulting in aggressive tumor outgrowth and dissemination. Here we mimic clonal expansion by evolving a colon cancer organoid model for over 100 cellular generations while simultaneously monitoring clone size, copy number variants (CNVs), and single nucleotide variants (SNVs) in thousands of individual cells. This offers the unique opportunity to combine multiple lines of evidence to establish reliable clonal evolution trees that reveal the order of events in which chromosomal deletions and amplifications occur and allow the identification of chromosomal aberrations that recur multiple times within the same population. For example, we observe recurrent sequential loss of chromosome 4 after loss of chromosome 18 in multiple unique tumor clones. Reanalysis of clinical data indeed suggests that this sequential loss is often observed in colon tumors and is associated with poor survival. Our results suggest that organoid evolution models complemented with integrated single-cell sequencing technology provide a powerful platform to study and ultimately control tumor evolution in detail.