间质源性OSTEONECTIN蛋白在胰腺癌细胞内的异常积聚促进胰腺癌侵袭转移的机制研究

Radical pancreatectomy is an effective treatment for pancreatic cancer. However, only a small proportion of patients with resected disease could achieve a long-term survival after surgery. We previously identified a subgroup of patients who did not benefit from radical pancreatectomy and had even worse survival compared with those with locally advanced disease who failed to receive pancreatectomy. These patients present a serological characteristic of CEA+/CA125+/CA19-9≥1,000U/mL preoperatively, and have an extremely high metastatic potential. Further confirming the existence of the subgroup of patients using the multicenter databases is the focus of this study. On the other hand, we would focus on the molecular characteristics of the specific subgroup with aiming to explore the potential targets for it. High-throughput screening technologies, including exome sequencing and whole-genome sequencing, together with bioinformatics analysis were performed on clinical samples. The data were validated in the independent samples or the TCGA database. The results showed that ECM3-OSTEONECTIN signaling was activated in the subgroup of patients who did not benefit from pancreatectomy. OSTEONECTIN, as key factor in the signaling, is not only expressed in tumor stroma, but also is abnormally accumulated in tumor cells themselves. Using the loss-or-gain function analysis based on in vitro and in vivo models, we found that the abnormal expression of OSTEONECTIN in pancreatic cancer cells derived more from extrinsic stromal protein. In contrast, endogenous expression of OSTEONECTIN in pancreatic cancer cells is suppressed by hypermethylation in its promoter region. High levels of OSTEONECTIN enhance the invasiveness of pancreatic cancer cells by activing ERK/MAPK signaling. Meanwhile it upregulates cytokines including MCP-2, SDF-1, IL-8, M-CSF-1, subsequently recruiting HLA-LR+/CD206+ macrophages and contributing the metastatic potential of pancreatic cancer. In this regard, OSTEONECTIN may be a new target aiming to improve the operation efficacy and overall survival of patients with pancreatic cancer. The effectiveness of intervention for it would be explored in this study. Altogether, we reveal for the first time a novel mechanism facilitating oncogenic process by OSTEONECTIN in pancreatic cancer cells and provide the basis for a new strategy to realize “personalized operation” of pancreatic cancer.

本课题组前期从单中心逾千例的可切除胰腺癌临床样本中甄别出一组以高转移潜能，预后极差为特征的“手术不获益”亚群。本项目通过高通量筛选技术和生物信息学分析，证实间质ECM3-OSTEONECTIN Signaling活化是该亚群胰腺癌的分子特征；并初步阐明除间质外，OSTEONECTIN蛋白以外源性摄取的方式在胰腺癌细胞内异常积聚，激活ERK/MAPK/Cytokines信号发挥促癌功能。后续研究一方面拟收集多中心数据验证“手术不获益”亚群的临床特征和分子标记；另一方面将在多个体内外模型上全面验证OSTEONECTIN功能，并利用多种基因操控技术或蛋白-蛋白相互作用分析寻找OSTEONECTIN调控促癌的关键分子和效应细胞，探讨多层次干预OSTEONECTIN的有效性。最终为改善“手术不获益”亚群预后提供生物学靶点，也为胰腺癌“个体化综合治疗”的开展提供参考。

根治手术是胰腺癌病人获得长期生存的唯一希望。但迄今为止，手术疗效依然不佳。前期首次甄别出一个具有独特临床特征的胰腺癌“手术不获益”亚群后，本项目针对该亚群分子特征进行探索。基于一系列高通量检测及生物信息学分析，并通过基因调控技术及体内外多个模型验证，我们证实间质ECM3-OSTEONECTIN Signaling及其核心分子OSTEONECTIN的活化是驱动胰腺癌“手术不获益”亚群恶性进展的关键分子信号。进一步研究发现间质分子OSTEONECTIN除表达于胰腺癌间质外，更通过胞外摄取的方式在癌细胞内异常积聚；从而激活下游“ERK/MARK-细胞趋化因子”信号，上调间质内巨噬细胞上Siglec15表达诱导其M2-型极化，促进胰腺癌转移。因此特异性针对癌细胞内OSTEONECTIN表达或活化以及“癌细胞-Siglec15+巨噬细胞”交互作用有望成为“手术不获益”亚群乃至整体胰腺癌病人的有效干预靶点。从而这一研究为“个体化诊治”在胰腺癌中的全面开展提供了临床可借鉴模式。