Single-cell profiling and integrative TCR analysis reveals tumor-mutation associated phenotypes and immune repertoire in lung adenocarcinoma

BackgroundTumor genomics has been linked to prognosis and response to therapies in non-small cell lung cancer (NSCLC) including well-established associations with aggressive disease biology in both KRAS and tp53 mutated tumors. Deciphering the contribution of specific mutations to the composition of the tumor immune microenvironment (TME) has been more complex. Early work suggests that specific mutations may sculpt the TME and affect prognosis and response to immunotherapy, however much remains unknown about how mutations mediate the interaction between tumor cells and immune cell populations. Here, we applied single cell sequencing to investigate the association between TME composition and the presence of KRAS and tp53 mutations in NSCLC focusing on conventional T cell phenotype as well as TCR clonality and predicted specificity.MethodsWe profiled the tumors of 18 patients with lung adenocarcinoma with concomitant tumor genomic profiling using single-cell RNA sequencing with paired protein expression (CITEseq) and single-cell V(D)J immune repertoire analysis. To the immune repertoire we applied GLIPH 2 and clustering to uncover TCR-antigen associations.ResultsWe identified that an exhausted CD8 T cell phenotype that differentially expresses genes including: HAVCR2 (TIM3), PDCD1, LAG3, and ENTPD1 was significantly enriched in tp53 mutated NSCLC (P = 0.013), with a strong trend to suggest that this holds irrespective of KRAS status. This phenotype exhibited a more clonal TCR repertoire compared to other T cell populations, and its clonotypes clustered with known external antigens. Fractional abundance of this phenotype in profiled tumor above the median for the patient cohort was associated with shorter overall survival (P = 0.33, HR = 2.25 [0.44 – 11.15]), and was significantly associated with increasing tumor stage (P = 0.043).ConclusionsWe identified a unique tp53-associated T cell phenotype that we hypothesize infiltrates tp53 mutant NSCLC early in the course of disease and becomes exhausted over time and less able to mediate anti-tumor immunity. This population may underpin the responsiveness of tp53 mutant NSCLC to immune checkpoint inhibitors and may serve as a key biomarker in the future.Legal entity responsible for the studyThe authors.FundingPrincess Margaret Cancer Centre, Canadian Institutes of Health Research (CIHR).DisclosureM. Ghaedi: Financial Interests, Full or part-time Employment: Sanofi Canada. F.A. Shepherd: Financial Interests, Personal, Advisory Board: AstraZeneca; Financial Interests, Personal, Invited Speaker: AstraZeneca; Financial Interests, Institutional, Local PI: AstraZeneca, Novartis. N.B. Leighl: Financial Interests, Personal, Other, CME/independent lectures: MSD, BMS; Financial Interests, Personal, Invited Speaker, independent lectures: Novartis, Takeda; Financial Interests, Personal, Invited Speaker, independent CME lecture: BeiGene; Financial Interests, Personal, Invited Speaker, Independent CME lectures: Janssen; Financial Interests, Institutional, Research Grant: AstraZeneca, Lilly, MSD, Pfizer, Janssen, Inivata/Neogenomics, Novartis; Other, Travel funding to deliver independent CME lectures: AstraZeneca; Other, Travel funding to deliver independent CME lecture: Janssen, MSD, Sanofi, Guardant Health. P.S. Ohashi: Financial Interests, Advisory Board: Providence Therapeutics, Treadwell Therapeutics, Tikva Allocell, Rondo Therapeutics. Inc.; Financial Interests, Research Funding: Providence Therapeutics. A.G. Sacher: Financial Interests, Institutional, Coordinating PI: Genentech-Roche, BMS, AstraZeneca; Financial Interests, Institutional, Local PI: Amgen, Iovance, CRISPR Therapeutics, Merck, Pfizer, GSK, Spectrum, Lilly, BridgeBio, Hotspot Therapeutics. All other authors have declared no conflicts of interest.