SRA

Colorectal tumors are often densely infiltrated by immune cells that have a role in surveillance and modulation of tumor progression but are burdened by exhaustion mechanisms fueled by the tumor microenvironment, which must be counteracted to achieve control. Here, we deployed a multidimensional approach to unravel the T cell functional landscape in tumor and peritumoral tissues from primary colorectal cancers and liver metastases. We found that T-cells are mainly localized at the front edge and that tumor-infiltrating T cells co-express multiple inhibitory receptors and highlighted CD39 as the major driver of exhaustion in both primary and metastatic colorectal tumors. By CRISPR/Cas9 genome editing tools, we simultaneously redirected T-cell specificity employing a novel T-cell receptor targeting the HER-2 antigen, and disrupted CD39, thus generating triple-knockout engineered lymphocytes. We showed that the absence of CD39 confers HER2-specific T cells a functional advantage in eliminating HER2+ patient-derived organoids, starring the relevance of the CD39 axis for further exploitation in adoptive T-cell therapy strategies to treat primary and metastatic colorectal cancer. Overall design: 37 tissues derived from non-neoplastic, peritumoral and tumoral areas of 13 patients diagnosed with primary Colorectal Cancer were dissociated to obtain a single cell suspension using GentleMacs dissociator (Miltenyi).