Comprehensive characterization of early-stage Non-Small Cell Lung cancers with multi-modal data integration

Non-small cell lung cancer (NSCLC) is one of the leading causes of cancer-related death despite the availability of therapies targeting the tumor microenvironment (TME) and/or the tumor. Immunotherapy and novel targeted therapy have improved the management of NSCLC, but the heterogeneity of the disease still hampers treatment efficacy as well as further therapeutic advances. Here, we used spatial single-cell data complemented with bulk RNAseq and whole exome sequencing to investigate the TME of 192 resected, mostly early-stage NSCLC tumors and to characterize associations of genetic, clinical and lifestyle factors with cellular and histological properties of the TME. We found, that the TME of squamous cell carcinoma (LUSC) harbored stronger signs of inflammation and immune exhaustion than adenocarcinoma (LUAD), but interestingly that elevated PD-L1 expression was associated with inflammation and markers of lymphocyte activation/exhaustion only in LUAD. Smoking correlated with T cell infiltration and TP53 mutation in LUAD, and TP53 mutation was associated with proliferation of tumor cells. In both histologies, naïve and BCL2 ⁺ T cells decreased with higher clinical stages. EGFR -driven tumors showed fewer proliferating, activated and exhausted T cells, but more CD4 T cells and HLA-DR+ tumor cells, than tumors with wild type EGFR. Multi-modal integration of our four data types showed that most variation in our cohort was along the axes of histology, smoking, TP53 mutation, and inflammation. Our integration identified histology-specific prognostic signatures, with a LUSC-enriched profile of proliferation, inflammation, and smoking associated with poor prognosis in LUAD. In summary, we provide a rich, multimodal, single-cell characterization of a large NSCLC cohort as a resource and suggest that future investigations of biomarkers for ICI in NSCLC will benefit from stratification by histology.