Challenging the control through a single-cell perspective on normal adjacent tissue in colorectal cancer

While normal adjacent tissue (NAT) is used as a standard control in cancer studies, NAT in close proximity to the tumor can display tumor-like molecular features, which are associated with a higher risk of local recurrence in colorectal cancer (CRC). To investigate the cellular composition and molecular features of NAT in detail, we analyzed single-cell RNA sequencing and spatial transcriptomics datasets of CRC tissue, corresponding NAT, and healthy colorectal biopsies. Our findings revealed significant differences in both cellular composition and gene expression profiles between healthy, NAT, and tumor tissues. We identified differentially expressed genes between healthy and normal adjacent tissue for each cell type, as well as tumor-associated copy number variations within NAT. Furthermore, we identified gene panels that effectively distinguish between healthy tissue and tissues exhibiting tumor-associated gene expression profiles, including NAT and tumor tissue. Finally, we developed a pharmacological strategy to reverse tumor-associated gene expression patterns in vitro.