A new mouse model for studying the pathogenesis and immunobiology of intrahepatic cholangiocarcinoma and improving its immunotherapy

摘要 肝内胆管细胞癌(ICC)是一种侵袭性的原发性肝癌(PLC)，尤其难以 治疗，总的5年生存率为8%。与主要的PLC一样，肝细胞癌(HCC)、ICC 由于胆汁淤积性肝病、非酒精性脂肪性肝病、 二型糖尿病和烟酒滥用。与肝细胞癌不同，肝细胞癌通常在晚期被检测到，当 生存前景不佳，这是目前放射学对其与早期肝细胞癌的不精确鉴别的并发症 技巧。两种肝癌的早期鉴别诊断非常关键，因为它们的处理方式不同，且缺乏 国际刑事法院和肝细胞癌共有的靶向致癌驱动因素。这个问题可以通过识别PLC来解决- 类型不可知的治疗方法，如免疫检查点抑制(ICI)，适用于两种ICC 还有肝癌。然而，尽管基于ICI的治疗被批准用于肝细胞癌，其疗效已经通过以下方式得到改善 与血管内皮生长因子抑制剂联合使用，它们在ICC中的表现很差。这一失败可以部分归因于 对ICC特异性免疫抑制肿瘤微环境(TME)和 缺乏合适的小鼠模型来评估ICI与治疗相结合的能力 解除免疫抑制。我们已经通过开发一种新的ICC模型解决了这个问题，该模型与 以前的小鼠模型不依赖于强制过度表达强大的致癌驱动因素 在没有获得额外突变的选择压力的情况下诱导ICC。我们的MUP-UPA/NRF2法案 内质网应激与转录因子NRF2双电位激活相结合的模型 肝胆祖细胞和成熟肝细胞，表现出健壮、高穿透性、类似人类的逐步 进展为ICC，伴随着表现CD8+T细胞的免疫抑制TME的积聚 细胞排斥。我们计划建立MUP-uPA/NRF2Act小鼠作为研究进化的主要模型 ICC及其免疫抑制药TME，并寻找解除免疫抑制和治疗的方法 增强对现有PD-(L)1抑制剂的反应。为了实现这一目标，我们将全面描述MUP- UPA/NRF2Act小鼠，定义ICC每个阶段的转录、表观遗传和遗传变化 进展，包括胆管反应、胆管上皮内瘤变和已确诊的促结缔组织增生性癌。 我们还将确定肿瘤浸润性淋巴细胞在每个阶段的T细胞受体(TCR)谱系。 恶性进展和挖掘癌细胞转录数据产生的调节因子 促结缔组织增生、免疫抑制和CD8+T细胞排斥。这些研究将得到以下补充 以鉴定靶向分子为目标的免疫表型和免疫耗竭实验 负责建立ICC特异性免疫抑制TME的开关。我们将使用这些信息 确定临床批准的可以克服免疫抑制并极大地提高疗效的治疗方法 以PD-(L)1抑制剂为基础的脑梗塞治疗。

ABSTRACT Intrahepatic cholangiocarcinoma (ICC) is an aggressive primary liver cancer (PLC) that is particularly hard to treat, having an overall 5-year survival rate of 8%. Like the major PLC, hepatocellular carcinoma (HCC), ICC incidence is on the rise due to the ongoing increase in cholestatic liver diseases, non-alcoholic fatty liver diseases, type 2 diabetes and alcohol and tobacco abuse. Unlike HCC, ICC is often detected at an advanced stage when survival prospects are poor, a complication of its imprecise differentiation from early HCC by current radiological techniques. Early differential diagnosis of two PLCs is critical due to their distinct management and the absence of targetable oncogenic drivers shared by ICC and HCC. This problem could be solved by identification of a PLC- type agnostic therapeutic approach, such as immune checkpoint inhibition (ICI), that is applicable to both ICC and HCC. However, while ICI based therapies were approved for HCC and their efficacy has been improved by combination with VEGF inhibitors, they have performed poorly in ICC. This failure can be attributed, in part, to poor mechanistic understanding of the ICC-specific immunosuppressive tumor microenvironment (TME) and lack of suitable mouse models that allow the evaluation of ICI in combination with therapeutics capable of dismantling immunosuppression. We have solved this problem by developing a new ICC model that unlike previous mouse models does not depend on forced overexpression of potent oncogenic drivers that rapidly induce ICC in the absence of selective pressure for acquisition of additional mutations. Our MUP-uPA/NRF2Act model depends on combination of ER stress with activation of transcription factor NRF2 in bipotential hepatobiliary progenitors and mature hepatocytes and shows robust, highly penetrant, human-like stepwise progression towards ICC that is accompanied by the buildup of an immunosuppressive TME exhibiting CD8+ T cell exclusion. We plan to establish the MUP-uPA/NRF2Act mouse as the leading model for studying the evolution of ICC and its immunosuppressive TME and for finding treatments that will dismantle immunosuppression and boost the response to existing PD-(L)1 inhibitors. To accomplish this goal, we will fully characterize the MUP- uPA/NRF2Act mouse, defining the transcriptomic, epigenetic, and genetic alterations at each stage of ICC progression, including ductular reactions, biliary intraepithelial neoplasia and established desmoplastic cancer. We will also define the T cell receptor (TCR) repertoire of tumor infiltrating lymphocytes at each stage of malignant progression and mine the transcriptomic data for cancer cell produced factors that mediate desmoplasia, immunosuppression and CD8+ T cell exclusion. These studies will be complemented by immunophenotyping and immunodepletion experiments whose goal is the identification of targetable molecular switches responsible for establishment of the ICC-specific immunosuppressive TME. We will use this information to identify clinically approved treatments that can overcome immunosuppression and vastly improve the efficacy of ICI therapy based on PD-(L)1 inhibitors.