Targeting the fibroblast-immune cell crosstalk to relieve immune suppression in the pancreatic cancer microenvironment

摘要 胰腺癌的特点是广泛的纤维炎症反应，或肿瘤间质。在免疫期间 间质内细胞丰富，它们在很大程度上是免疫抑制的，因此胰腺癌 对免疫治疗基本无反应。胰腺癌基因工程小鼠模型的建立 概括了胰腺癌的逐步发展过程，是研究前驱病变的理想工具，例如 胰腺上皮内瘤变(Panin)。对Panin阶段免疫渗入的分析显示， 免疫抑制在很早就建立起来，并先于恶性进展。其作用机制 在胰腺癌中免疫抑制的建立的基础仍不清楚 了解它们对于设计新的胰腺癌化疗方法具有重要意义。 癌症。我们之前使用单细胞RNA测序来表征人类的基因表达谱 胰腺癌免疫浸润性。然后，我们绘制了微环境中潜在的细胞-细胞相互作用图 基于配体和受体的相互表达。在预测的相互作用中，我们确定了WNT 肿瘤细胞表达的配体驱动胰腺癌T细胞亚群的信号激活 和肿瘤相关成纤维细胞(CAF)。我们和其他人之前曾将不适当的激活 胚胎信号通路，包括WNT信号，作为胰腺癌的特征。WNT信令 是胰腺癌中被激活的核心通路之一。我们之前的研究表明，消融上皮性WNT 信号转导抑制胰腺癌的发生，但WNT在胰腺癌中的潜在作用 微环境，特别是免疫细胞中的情况尚不清楚。CAF表达WNT家族的几种配体； 为了抑制它们的表达，我们灭活了PORCN(豪猪)，这是一种需要 胰腺成纤维细胞中WNT配体的酰化和分泌。然后我们移植了胰腺癌细胞 并观察到生长速度减慢。要确定T细胞激活WNT信号是否对此很重要 结果，我们产生了编码蛋白质TCF1的转录因子Tcf7被灭活的小鼠 在CD4+T细胞中。在这些动物中，我们观察到移植的肿瘤生长减少，CAF发生变化 表型，并增强抗肿瘤免疫的激活。在这项提案中，我们计划在我们初步的基础上 资料剖析WNT信号在胰腺癌中驱动免疫抑制的机制。从长远来看 目标是设计靶向方法，可能逆转这种疾病的免疫抑制。

ABSTRACT Pancreatic cancer is characterized by an extensive fibroinflammatory reaction, or tumor stroma. While immune cells are abundant within the stroma, they are largely immune suppressive, and therefore pancreatic cancer is largely unresponsive to immunotherapy. Genetically engineered mouse models of pancreatic cancer recapitulate the stepwise progression of pancreatic cancer, and are ideal to study precursor lesions, such as pancreatic intraepithelial neoplasia (PanIN). Analysis of the immune infiltrates at the PanIN stage revealed that immune suppression is established very early on and precedes malignant progression. The mechanisms underlying the establishment of the immune suppression in pancreatic cancer remain unknown and understanding them is of fundamental importance to design new chemotherapy approaches for pancreatic cancer. We previously used single cell RNA sequencing to characterize gene expression profiles in the human pancreatic cancer immune infiltrate. We then mapped potential cell-cell interactions within the microenvironment based on reciprocal expression of ligands and receptors. Among predicted interactions, we identified WNT signaling activation in the T cell compartment of pancreatic cancer, driven by ligands expressed by tumor cells and cancer associated fibroblasts (CAFs). We and others have previously associated inappropriate activation of embryonic signaling pathways, including WNT signaling, as a characteristic of pancreatic cancer. WNT signaling is one of the core pathways activated in pancreatic cancer. We previously showed that ablation of epithelial WNT signaling inhibits the onset of pancreatic carcinogenesis, but the potential role of WNT in the pancreatic cancer microenvironment and specifically in immune cells is unknown. CAFs express several ligands of the WNT family; to ablate their expression, we inactivated PORCN (PORCUPINE), a transmembrane enzyme required for acylation and secretion of WNT ligands, in pancreatic fibroblasts. We then transplanted pancreatic cancer cells and observed reduced growth. To determine whether T cell activation of WNT signaling was important for this effect, we generated mice where the transcription factors TCF7, encoding for the protein TCF1, was inactivated in CD4+ T cells. In these animals, we observed reduced growth of transplanted tumors, alterations in the CAF phenotype, and increased activation of anti-tumor immunity. In this proposal, we plan to build on our preliminary data to dissect the mechanism of WNT signaling driven immune suppression in pancreatic cancer. The long term goal is to design targeting approaches that might reverse immune suppression in this disease.