Cancer associated fibroblast-T cell crosstalk promotes purinergic synthesis in non-small cell lung cancer (NSCLC)

The most abundant stromal cells of the tumor microenvironment (TME), cancer-associated fibroblasts (CAFs), inhibit the cytotoxic T cell response in solid tumors. CD39 and CD73 together deplete extracellular ATP in the TME to produce adenosine, which can promote tumor immune evasion. We aimed to investigate the role of CAF-T-cell crosstalk in the adenosine pathway in NSCLC. CD4⁺ and CD8⁺ T cells were flow sorted from in vitro culture with or without CAFs, and RNA-Seq was performed using the nCounter platform (NanoString). The expression of cell surface markers involved in adenosine production was assessed using flow cytometry in CAFs and peripheral T cells from healthy donors or early NSCLC patients following in vitro co-culture. Purinergic synthesis of AMP and adenosine was measured by HPLC. TCGA data was used to investigate the translational relevance of adenosine signaling in NSCLC, and NanoString Geomx was performed to investigate differences in the transcriptomics of tumor stromas with a high versus low adenosine signature. CD4⁺ and CD8⁺ T cells isolated from in vitro co-culture with CAFs show upregulated expression of CD39 and CD73, which results in functionally relevant production of AMP and adenosine. TCGA data illustrates that adenosine signaling is predictive of poor outcome in lung squamous cell carcinoma. Spatial transcriptomics (GeoMx) of the tumor stroma of early untreated NSCLC patients shows a downregulation of immune-related genes in lung squamous-cell carcinoma. CAF T cell crosstalk promotes the expression of CD39 and CD73 on CAFs and T cells, resulting in an increased synthesis of AMP and adenosine in vitro.