巨噬细胞通过外泌体/XRN1通路降解胰腺导管上皮细胞BRCA1/2 mRNA引发基因组不稳定的机制

Chronic inflammation is a risk factor of pancreatic cancer. However, the mechanisms involved are still unclear. Previous studies have shown that inflammation can inhibit the expression of BRCA1. Deficiencies of BRCA1 and BRCA2 increase the incidence of pancreatic cancer. Since BRCA1 and BRCA2 participate in the homologous recombination (HR) repair of DNA double-strand break (DSB), and play an important role in maintaining genomic stability, it is possible that inflammation might promote the occurrence of pancreatic cancer through repressing expressions of BRCA1 and BRCA2. Based on our serial studies, which were supported by previous NSFC grants, we found out that marcrophages, the dominant population of inflammatory cells, could repress expressions of BRCA1 and BRCA2 in pancreatic ductal epithelial cells (PDECs). We hypothesize that marcrophages could up-regulate several miRNAs in PDECs through TGF-β pathway. These miRNAs include miR-1224-3p and miR-526b-5p et. al., which could target BRCA1 and BRCA2 mRNAs. Binding of miRNAs could stimulate 5' decapping of BRCA1 and BRCA2 mRNAs. Moreover, macrophages might deliver XRN1, the key 5' exoribonuclease, into PDECs through exosome manner. XRN1 can recognize and degrade 5' decapped mRNAs. These two mechanisms may synergistically result in the selective degradation of BRCA1 and BRCA2 mRNAs. Down-regulations of BRCA1 and BRCA2 inhibit homologous recombination (HR)-mediating DNA double-strand break (DSB) repair, leading to genomic instability and mutation accumulation, which will increase the risk of tumorigenesis. In the present study, we will verify the mechanisms involved in the regulation of BRCA1 and BRCA2 expressions through macrophage/TGF-β/miRNA, and macrophage/exosome/XRN1 pathways. Moreover, by using several well-established animal models, we will investigate the participation of BRCA1 and BRCA2 deficiencies in the genomic instability. Our study will not only establish a new paradigm that significantly affects our views on how chronic inflammation leads to the occurrence of pancreatic cancer, but will also lay groundwork for discovering new markers for the early diagnosis of deadly pancreatic cancer.

慢性炎症促胰腺癌发生。申请人发现炎症细胞主要群体—巨噬细胞，能够抑制胰腺导管上皮细胞（PDEC）中DNA损伤修复关键基因BRCA1、BRCA2表达。基于前期研究提出科研假说：巨噬细胞一方面通过TGF-β通路，上调PDEC中miR-1224-3p、miR-526b-5p等靶向BRCA1/2 mRNA的miRNA；另一方面通过外泌体，向PDEC输送5'核酸外切酶XRN1。XRN1能够降解结合了miRNA的mRNA。两方面协作，降解BRCA1/2 mRNA，下调其表达，进而抑制DNA同源重组修复，导致基因组不稳定，从而增加恶变几率。基于前期建立的多种体内体外模型，我们将研究巨噬细胞/外泌体/XRN1，和巨噬细胞/TGF-β/miRNA信号通路，对PDEC中BRCA1/2表达的调控；以及BRCA1/2缺陷对DNA损伤修复和基因组稳定性的影响。从而为揭示胰腺“炎癌转化”的分子机制奠定一定的理论基础。

胰腺导管腺癌（pancreatic ductal adenocarcinoma，PDAC）是一种恶性程度很高且预后极差的肿瘤。深入研究胰腺癌的发生机制对胰腺癌的预防、早期诊断和治疗具有指导性意义。我们的研究取得以下成果：阐释慢性胰腺炎炎症微环境可以通过抑制BRCA1、BRCA2的表达，抑制同源重组修复（HR），细胞倾向于采用非同源重组末端链接（NHEJ）进行DNA双链损伤（DSB）修复，造成基因组不稳定，从而增加癌变的风险。巨噬细胞通过外泌体途径下调BRCA1、BRCA2的表达。巨噬细胞通过外泌体途径向胰腺导管上皮细胞输送XRN1，促进胰腺导管上皮细胞中的BRCA1、BRCA2 mRNA的降解，导致BRCA1、BRCA2表达下调。巨噬细胞外泌体中存在hsa-miR-146a-5p，可以结合BRCA1、BRCA2的3’-UTR区域，与XRN1相互协作，减少胰腺导管上皮细胞中BRCA1、BRCA2的表达。通过在巨噬细胞特异性敲除XRN1工具鼠（Lyz2 cre；XRN1 flox/flox）中诱导慢性胰腺炎，检测胰腺组织中BRCA1、BRCA2的表达，证明慢性胰腺炎中的巨噬细胞通过XRN1通路影响胰腺组织中BRCA1、BRCA2的表达。同时，我们构建了一种靶向胰腺癌递送BRCA1/2 siRNA以诱导胰腺癌DNA同源重组损伤修复（HR）缺陷的工程化外泌体，该工程化外泌体向胰腺癌靶向递送BRCA1/2 siRNA造成HR通路损伤从而影响DNA同源重组损伤修复能力缺陷。为治疗胰腺癌提供新的治疗靶点和治疗策略。