Conserved transcriptional connectivity of regulatory T cells in the tumor microenvironment informs novel combination cancer therapy strategies [mouse]

While regulatory T (Treg) cells are traditionally viewed as professional suppressors of antigen presenting and effector T cells in both autoimmunity and cancer, recent findings of distinct Treg functions in tissue maintenance suggest that their regulatory purview extends to a wider range of cells and is broader than previously assumed. To elucidate tumoral Treg “connectivity” to diverse tumor-supporting accessory cell types, we explored immediate early changes in their single cell transcriptomes upon punctual Treg depletion in experimental lung cancer and injury-induced inflammation. Prior to any notable T cell activation and inflammation, fibroblasts, endothelial and myeloid cells exhibited pronounced changes in their gene expression in both cancer and injury settings. Factor analysis revealed shared Treg-dependent gene programs, foremost, prominent upregulation of VEGF signaling-related genes upon Treg deprivation in either setting, as well as in Treg-poor vs Treg-rich human lung adenocarcinomas. Accordingly, punctual Treg depletion combined with short-term VEGF blockade showed markedly improved control of PD-1 blockade-resistant lung adenocarcinoma progression in mice compared to the corresponding monotherapies, highlighting a promising factor-based querying approach to elucidating novel rational combination treatments of solid organ cancers. Three mouse sets of data are deposited. 1. KP Lung Adenocarcinoma and stromal cells were collected from KrasLSL-G12D/WTTrp53fl/fl Foxp3GFP-DTR (KP-DTR) mice harboring ~3 months old Adenicarcinomas, following 48 hr treatment with Diphteria toxin (DT) to deplete Tregs, or PBS as control. Cells were isolated by Fluorescence-activated cell sorting (FACS), gating on CD45, CD31, GP38, MHCII CD11b, GR1 CD4 and CD8 and analysed by Bulk RNA-seq (08820, 08233) 2. KP Lung Adenocarcinoma and stromal cells were collected from KrasLSL-G12D/WTTrp53fl/fl Foxp3GFP-DTR (KP-DTR) mice harboring ~3 months old Adenicarcinomas, following 48 hr treatment with Diphteria toxin (DT) to deplete Tregs, or PBS as control. Cells were isolated by FACS gating on CD45- and CD45+ signal and analyzed using scRNAseq (07734). 3. KP-DTR mice were treatead with bleumycine for 21 days, followed by 48 hr treatment with Diphteria toxin (DT) to deplete Tregs, or PBS as control. Cells were isolated by FACS gating on CD45- CD45+ and EpCAM+ signal and analyzed using scRNAseq (10506)