Project description:Single-cell RNA-sequencing analyses were performed on unfractionated live cell mixtures from pancreatic tumors of KPC mice and wild-type mice to detect different myCAF subpopulations
Project description:Single-cell ATAC-sequencing analyses were performed on unfractionated live cell mixtures from pancreatic tumors of KPC mice and wild-type mice to better understand the epigenetic characteristics of different cell populations
Project description:A major barrier in pancreatic immunotherapy is the \"cold\" TME, characterized by T cell exclusion and a deficiency in cDC1s necessary for antigen presentation. Using the KPC mouse model, we found that while FAP-CD40 monotherapy only delayed tumor growth, combining it with PD1-IL2v induced robust regression. To elucidate the underlying mechanisms, we performed single-cell transcriptome analysis to compare the effects on T cell and myeloid populations across mono- and combination therapies. Our study revealed that the combination uniquely overcomes immune exclusion by generating dense intratumoral T cell-cDC1 clusters (TDCs). These TDCs drive anti-tumor immunity by sustaining local T cell proliferation within the parenchyma, independent of lymph node priming.
Project description:We harvested and sequenced the spontaneous pancreatic tumor generated by a 6-month-old KPC mouse (KrasLSL-G12D; Trp53LSL-R172H; Ptf1a-Cre).
Project description:Bulk RNA sequencing of sorted peri-pancreatic LN cDC1s from different stages of neoplastic development in the KPC mouse model of pancreatic adenocarcinoma.
Project description:During carcinoma progression, mesenchymal stromal cells (MSCs) become recruited to tumors and contribute to the pool of cancer-associated fibroblasts (CAFs). Sub-populations of CAFs, have diverse and incompletely understood effects on disease progression and resistance to therapy. Adipose stromal cells (ASCs), the MSCs from fat tissue increasingly recruited by carcinomas in the context of obesity, have been shown to support cancer progression. Here, the roles of perivascular CAFs expressing platelet-derived growth factor receptor beta (Pdgfrb) and of CAFs expressing non-glycanated decorin (ngDCN), a marker of ASCs, were analyzed. We used mice orthotopically grafted with pancreatic ductal adenocarcinoma (KPC) cells. Ablation of cells expressing thymidine kinase under the control of Pdgfrb promoter with ganciclovir resulted in suppression of primary pancreatic tumor growth. However, ablation Pdgfrb+ cells induced spontaneous metastases to the liver. For depletion of ngDCN+ cells, we used a hunter-killer peptide D-CAN, which has been previously reported to induce apoptosis of ASCs and CAFs in mouse models. Here, D-CAN also had a negative effect on pancreatic tumor growth, however, there was no significant effect on metastatic dissemination. Single cell RNA sequencing of tumors demonstrated that targeting of Pdgfrb+ and ngDCN+ stromal cells had distinct effects on sub-populations of CAFs. Endothelial cell and cancer cell survival was decreased by depletion of either Pdgfrb+ or ngDCN+ stroma. However, pathway analysis revealed that depletion of Pdgfrb+ and ngDCN+ stromal cells has different effects on the markers of cancer cell aggressiveness. D-CAN treatment, and to a less extent Pdgfrb+ cell ablation, suppressed macrophage M2 polarization and increased infiltration of cytotoxic T-lymphocytes. This observation, confirmed in the MyCap graft model of prostate cancer, suggested that ablation of ngDCN+ stroma should synergize with immune checkpoint inhibitors. We tested the effect of D-CAN on the efficacy of anti-PD-L1 antibody in the orthotopic KPC model. Compared to D-CAN and anti-PD-L1 antibody alone, combination treatment had a synergistic effect on tumor growth. Importantly, liver metastases were suppressed by the D-CAN / anti-PD-L1 antibody combination, but not by individual agents. We conclude that improved approaches to target mesenchymal stroma in tumors may be effective in combination with immunotherapy.
