Identification and Characterization of Cancer-Associated Fibroblast Subpopulations in Lung Adenocarcinoma

SIMPLE SUMMARY: Considering that cancer-associated fibroblasts (CAFs) facilitate cancer cell motility, invasiveness, and drug resistance, heterogeneity within the CAF population may be of major significance during cancer progression and metastasis. Through pseudotime trajectory analysis of single-cell RNA sequencing data, we revealed several subpopulations of lung CAFs with distinct gene expression patterns, namely, immunosuppressive, neoantigen-presenting, myofibroblastic, and proliferative CAFs. The knockdown of KPNA2, one of the neoantigen-presenting CAF-specific markers, attenuated CAF invasiveness, suggesting that CAF subtype markers may represent therapeutic targets within the tumor microenvironment of lung cancer patients. ABSTRACT: Cancer-associated fibroblasts (CAFs) reside within the tumor microenvironment, facilitating cancer progression and metastasis via direct and indirect interactions with cancer cells and other stromal cell types. CAFs are composed of heterogeneous subpopulations of activated fibroblasts, including myofibroblastic, inflammatory, and immunosuppressive CAFs. In this study, we sought to identify subpopulations of CAFs isolated from human lung adenocarcinomas and describe their transcriptomic and functional characteristics through single-cell RNA sequencing (scRNA-seq) and subsequent bioinformatics analyses. Cell trajectory analysis of combined total and THY1 + CAFs revealed two branching points with five distinct branches. Based on Gene Ontology analysis, we denoted Branch 1 as “immunosuppressive”, Branch 2 as “neoantigen presenting”, Branch 4 as “myofibroblastic”, and Branch 5 as “proliferative” CAFs. We selected representative branch-specific markers and measured their expression levels in total and THY1 + CAFs. We also investigated the effects of these markers on CAF activity under coculture with lung cancer cells. This study describes novel subpopulations of CAFs in lung adenocarcinoma, highlighting their potential value as therapeutic targets.