The Potential Role of Firboblast Activation Protein as a Natural Killer Cell Immune Checkpoint in Pancreatic Cancer

项目总结 免疫治疗在很大程度上对胰腺癌无效，部分原因是周围密集的间质。 纤维化，造成免疫抑制的微环境。纤维化的主要细胞成分， 胰腺星状细胞(PSCs)是以成纤维细胞激活蛋白(FAP)表达升高为特征的细胞。FAP 是一种II型跨膜丝氨酸蛋白酶，在正常胰腺中表达最低，但在 胰腺癌FAP在90%的病变中过度表达，并与较差的临床预后相关。这里 我们使用了一种新的体外共培养系统，该系统利用了初级供体来源的PSCs和人类自然杀伤细胞 (NK)细胞株NK92，以评估PSCs和NK细胞之间的关系。我们测试了NK细胞的能力 杀死PSCs，并监测FAP表达和激活标志物。我们还评估了FAP的效果 体外抑制NK细胞活性和体内胰腺肿瘤清除。我们发现NK细胞被激活 通过和杀伤PSCs，可能通过NK细胞表面受体NKG2D识别PSCs上的MICA/B。在直接 与PSCs接触，PSCs下调FAP，而NK细胞上调FAP。这是第一次发现NK细胞。 已被证明可产生FAP，FAP的诱导是通过细胞间的接触来介导的。此外， NK细胞表达FAP与失活表型有关。FAP抑制增强小鼠NK细胞杀伤活性 PSCs在体外和体内增强胰腺肿瘤清除。抑制FAP的抗肿瘤活性为 通过添加抗PD-1治疗而增强。基于这些发现，我假设FAP具有NK功能 细胞免疫检查站。FAP在NK细胞活化后表达，以减轻细胞毒作用，并可 抑制增强抗肿瘤免疫功能。为了验证这一假设，我的目标是确定FAP的机制 NK细胞的诱导(目标1)，评估这些发现的临床相关性(目标1A和1D)，并操纵 增强体外和体内抗肿瘤免疫反应的FAP活性(目标2)。成功完成 这些目标将确定调节FAP表达的因素，并进一步了解FAP是如何调节的 免疫反应。这些发现将填补围绕FAP表达调控的知识空白 为增强抗肿瘤免疫活性提供了新的途径。

PROJECT SUMMARY Immunotherapy has been largely ineffective in pancreatic cancer, partially due to the surrounding dense stromal fibrosis which creates an immunosuppressive microenvironment. The main cellular component of this fibrosis, pancreatic stellate cells (PSCs), are marked by elevated expression of fibroblast activation protein (FAP). FAP is a type II transmembrane serine protease that is minimally expressed in normal pancreas, however, in pancreatic cancer FAP is overexpressed in 90% of lesions and is associated with worse clinical outcomes. Here we use a novel in vitro co-culturing system that utilizes primary donor-derived PSCs and a human natural killer (NK) cell line, NK92, to assess the relationship between PSCs and NK cells. We tested the ability of NK cells to kill PSCs and monitored for FAP expression and markers of activation. We also assessed the effect of FAP inhibition on NK cell activity in vitro and pancreatic tumor clearance in vivo. We found that NK cells are activated by and kill PSCs, potentially via NK cell surface receptor NKG2D recognition of MICA/B on PSCs. Upon direct contact with PSCs, PSCs downregulate FAP, however, NK cells upregulate FAP. This is the first-time NK cells have been shown to produce FAP and that induction of FAP is mediated by cell-to-cell contact. Furthermore, FAP expression by NK cells is associated with an inactivate phenotype. FAP inhibition enhanced NK killing of PSCs in vitro and enhanced pancreatic tumor clearance in vivo. The anti-tumor activity of FAP inhibition was enhanced by addition of anti-PD-1 therapy. Based on these findings, I hypothesize that FAP functions as an NK cell immune checkpoint. FAP is expressed in NK cells after activation to attenuate cytotoxicity and can be inhibited to enhance anti-tumor immunity. To test this hypothesis, I aim to determine mechanisms of FAP induction in NK cells (Aim 1), assess the clinical relevance of these findings (Aim 1A and 1D) and manipulate FAP activity to enhance in vitro and in vivo anti-tumor immune responses (Aim 2). Successful completion of these aims will identify factors that regulate FAP expression and further our understanding of how FAP regulates the immune response. These findings will fill the gap in knowledge surrounding regulators of FAP expression and provide new approaches to enhance anti-tumor immune activity.