Karre, K. | bioRxiv Science

bioRxiv2025

TCR clonality and TCR clonal expansion in the in situ microenvironment of non-small cell lung cancer

T-cell activation and clonal expansion are essential for the efficacy of immunotherapy in non-small cell lung cancer (NSCLC) patients. Since the distribution of T-cell clones might provide insights into immunogenic mechanisms, we determined the /{beta} TCR clonality using RNA-sequencing from frozen tumor tissue of 182 NSCLC patients and paired the results with extensive in situ image and sequence analyses of the immune microenvironment of NSCLC. TCR clonality (Gini index) patterns ranged from high T-cell clone diversity with high evenness (Gini index low) to clonal dominance with low evenness (Gini index high). TCR clonality in cancer tissue was lower than in matched normal lung (p=0.021). High Gini index correlated strongly with distinct mutations (EGFR, P53), tumor mutation burden (p<0.001), and inflamed tumor phenotypes (PRF1, GZMA, GZMB, INFG) with exhaustion signatures (LAG3, TIGIT, IDO1, PD-1, PD-L1). Correspondingly, PD-1+, CD3+, CD8A+, CD163+, and CD138+ immune cells infiltrated cancer tissue with high TCR clonality. In situ sequencing revealed that dominant T-cell clones were more often of CD8-subtype and tended to approximate the tumor cell compartment (p<0.03). In a checkpoint inhibitor-treated NSCLC patient cohort, high TCR clonality was associated with therapy response (p=0.016) and prolonged survival (p=0.003, median survival 13.8 vs 2.9 months). Our robust analysis pipeline revealed diverse TCR repertoires related to genotypes and immune phenotypes. The in situ positioning of expanded T-cell clones indicated functional impact, which was clinically confirmed in NSCLC patients receiving immunotherapy. One sentence summaryT-cell clone expansion in NSCLC is associated with genetic mutations, immune phenotypes, immunotherapy response, and patient survival.

cancer biology↗

bioRxiv2021

PD-1 expression on NK cells can be related to cytokine stimulation and tissue residency

Although PD-1 was shown to be a hallmark of T cells exhaustion, controversial studies have been reported on the role of PD-1 on NK cells. Here, we found by flow cytometry and single cell RNA sequencing analysis that PD-1 can be expressed on MHC class I-deficient tumor-infiltrating NK cells in vivo. We also demonstrate distinct alterations in the phenotype of PD-1-deficient NK cells which in part could be attributed to a decrease in tumor-infiltrating NK cells in PD-1-deficient mice. NK cells from PD-1-deficient mice exhibited a more mature phenotype which might reduce their capacity to migrate and kill in vivo. Finally, our results demonstrate that PD-L1 molecules in membranes of PD-1-deficient NK cells migrate faster than in NK cells from wildtype mice, suggesting that PD-1 and PD-L1 form cis interactions with each other on NK cells.

immunology↗