Abstract B019: Investigating the role of antigen-presenting fibroblasts in the tumor microenvironment of non-small cell lung cancer
Authors: Hinshaw Rachael , Espinoza Carlos , Zhang Yaqing , Pasca di Magliano Marina , Galban Stefanie 
Published: October 2024

KRAS mutations are common in non-small cell lung cancer (NSCLC) and often co-occur with TP53 mutations. Although the KRAS oncogene was once considered ‘undruggable,’ FDA approval of KRASG12C inhibitors and promising pre-clinical data from KRASG12D inhibitors highlight the potential for targeted therapies. The tumor microenvironment (TME) of NSCLC comprises immune and stromal cells, including antigen-presenting cancer-associated fibroblasts (for simplicity, hereafter ‘apCAFs,’ regardless of cancer association), a population that expresses markers that interact with and affect T cell activation, exhaustion and differentiation. However, the role of apCAFs in tumorigenesis, how they interact with T cells, and their response to KRAS inhibitors remain unknown. A crucial need exists to characterize apCAFs to optimize therapeutic approaches. We used an inducible and reversible KrasG12D lung cancer mouse model (L-iKras) to perform single cell RNA sequencing analyses. Lung tissue from two each (1 male/1 female) of control mice, mice with KrasG12D turned on with doxycycline (ON), and mice with KrasG12D turned on for 20 weeks and off for 1 week via doxycycline withdrawal (OFF) were included. Fibroblasts were clustered and stratified using the Seurat package. One population resembled apCAFs expressing Pdgfra, Clec3b, Col1a1, and Col1a2 and antigen-presenting markers H2-Ab1, H2-Aa, and Cd74. This population increased in ON and OFF groups compared to controls and expressed genes required for three signals of T cell activation: engagement of the T cell receptor with the peptide- MHC complex of an antigen-presenting cell (APC); binding of costimulatory proteins CD28 or CTLA4 (T cell) with CD80/CD86 (APC); and secreted cytokines (APC) initiating T cell differentiation. This apCAF population showed increased markers for immune checkpoints Cd274 (PDL1) and Cd86, which can inhibit or exhaust T cells, upon KRAS inhibition. Further, these apCAFs upregulated markers involved with the induction of Tregs upon KRAS inhibition (Tgfb1). Treg populations were increased, and CD8 T cell populations were decreased in the OFF group compared to the ON group. These findings suggest that apCAFs are induced in the TME during tumorigenesis and interact with tumor-infiltrating T cells. Further, we hypothesize that apCAFs become immunosuppressive upon KrasG12D inhibition, activate Tregs, and induce CD8 T cell exhaustion. Interestingly, upregulation of immune checkpoints in apCAFs suggests that these cells may respond well to checkpoint therapies, which could be harnessed for future therapeutic approaches in NSCLC. The focus of our future studies will be to validate our findings in an orthotopic lung cancer mouse model developed by our lab, which utilizes KrasG12D-inducible and reversible lung cancer cell lines derived from L- iKras. With this model we will further characterize apCAF-T cell interactions, investigate the impact of KRAS inhibitors on apCAFs, and identify the role of apCAFs in tumor responses to combinations of KRAS inhibitors and checkpoint inhibitors.

Citation Format: Rachael Hinshaw, Carlos Espinoza, Yaqing Zhang, Marina Pasca di Magliano, Stefanie Galban. Investigating the role of antigen-presenting fibroblasts in the tumor microenvironment of non-small cell lung cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr B019.