The cellular genomic diversity, regulatory states and networking of the metastatic colorectal cancer microenvironment

ABSTRACTMetastatic colorectal cancer (mCRC) involves tumor cells seeding distant organs. Metastatic CRC genome biology has intrinsic properties related to heterogeneous clonal tumor cells and extrinsic properties of the cellular niche of the tumor microenvironment (TME). To characterize mCRC’s cell types and states, we used single-cell RNA sequencing (scRNA-seq) and DNA sequencing (scDNA-seq) on metastases in the liver and omentum, a tissue lining the abdomen. We performed scRNA-seq on 49,637 cells derived from mCRC, paired normal tissue and peripheral blood. We performed whole genome scDNA-seq on 3,683 mCRC cells from the same samples. These metastases had intrinsic heterogeneity, with clone-specific copy number variation and lineage differentiation. For the extrinsic niche, TME macrophages had a cellular state resembling cirrhotic macrophages and foam cells with indicators of increased activity for extracellular matrix (ECM) organization. TME fibroblasts had an expression program influencing ECM composition that was not observed in matched normal tissue. Only a few CD8 T cells were present among the liver mCRCs, indicating immune exclusion. Receptor-ligand analysis revealed that TME macrophages and fibroblasts formed an interactome, thus providing intercellular coordination which increased T cell exhaustion and exclusion. For one mCRC, we used in a patient-derived ex vivo tissue model that captures the TME components of the original metastatic tumor, applied specific perturbations to the TME immune cells and evaluated cellular state changes with scRNA-seq. Overall, our study identified tumor cell clonal variation, characterized specific TME properties of the CRC metastatic niche and demonstrated an experimental model of perturbing the cellular TME milieu.