Comparison of the tumor and lymph node immune microenvironment in early non-small cell lung cancer through multimodal single cell sequencing.

Abstract Background/Purpose: Previous studies have analyzed the tumor and local immune microenvironments in lung cancers and suggest immune modulation is associated with worse clinical outcome. However, the tumor-immune microenvironment in early-stage lung tumors and lymph nodes (LNs) have not been fully described. We aim to compare cell states in the immune microenvironments between lung tumors and LNs through multi-modal profiling of the transcriptome and surface proteins. Methods: Needle biopsy samples were taken from 10 treatment-naive early-stage lung cancer patients undergoing lung cancer resections. Tissues were obtained from normal lung, lung tumor, and multiple mediastinal LNs, and processed with scRNA-seq along with the Total-Seq C panel to quantify the levels of 130 cell surface proteins. In total, 98,337 cells (4,661 normal lung; 52,119 tumor; 42,117 LN) were identified with a median of 1,401 genes and 23 protein features detected per cell. Weighted-Nearest Neighbor analysis from the Seurat R package was applied to integrate the CITE-seq and RNA-seq level data for clustering cells into subpopulations. Results: Six broad cell populations were identified including T/NK, myeloid (CD14+), B (CD19+), mast (TPSAB1+), pDC (IRF8+), and epithelial (EPCAM+) cells. Protein expression measured with CITE-seq revealed additional T cell populations not captured by scRNA-seq. Preliminary results have identified cell populations enriched in LNs, which include naive CD4+ (CD4, FHIT, LEF1, CCR7) and CD8+ T cells (CD8A, TCF7, LEF1, CD27, CCR7). Additionally, we have identified numerous other CD8+ T-cell subpopulations enriched in tumors, including resident memory CD8+ T cells (KLRC1, ITGAE, ITGA1, VIM, JUN). Furthermore, immune populations enriched in LNs were similarly shared across different patients, while those enriched in tumors displayed patient-level specificity. Conclusion: Single-cell profiling reveals diversity in immune cell populations between LNs, tumor, and adjacent normal tissue in early-stage LUAD. The results suggest the composition of immune cell type is consistent across LNs but more heterogeneous in the tumor and adjacent normal tissue. In the future, we aim to determine if these markers for various immune subpopulations are associated with survival, recurrence, aggressiveness, and predict responses for neoadjuvant treatments, which could improve prognosis and patient quality of life. Citation Format: Zhan Xi, Yusuke Koga, Shannon McDermott, Jennifer E. Beane, Sarah A. Mazzilli, Kei Suzuki, Joshua D. Campbell. Comparison of the tumor and lymph node immune microenvironment in early non-small cell lung cancer through multimodal single cell sequencing. [abstract]. In: Proceedings of the AACR Special Conference: Precision Prevention, Early Detection, and Interception of Cancer; 2022 Nov 17-19; Austin, TX. Philadelphia (PA): AACR; Can Prev Res 2023;16(1 Suppl): Abstract nr P075.