EB病毒相关胃癌体内外模型的建立及其在靶向药物联合免疫细胞治疗策略中的应用

Gastric cancer is one of the most common malignant tumors, possessing more obvious heterogeneity when comparing with other cancers. As a result, it is particularly necessary to conduct individualized treatment for gastric cancer. The TCGA molecular typing divide gastric cancer into four categories: EBVaGC, as one of the categories, have PI3KCA hypermutations,CDKN2A methylation, JAK2, PD-L1/2 amplification, and specific antigen expression characteristics from EBV. It’s the perfect entry points to the immunotherapy but now still no research model and treatment strategy. We have successfully built the CTC detection platform and the PDX model for gastric cancer, and we will: (1) analyzing Immune index that related to prognosis by connecting with immunology characteristic in adjacent tissues or peripheral blood of cancer;(2)constructing the Epstein-Barr virus-associated gastric cancer model in vitro and in vivo and human immune reconstitute animal model in tumor bearing mice basing on patient-derived tumor xenograft (PDX) and circulating tumor cell (CTC); (3) screening PI3K, CDK4/6, JAK2 targeted inhibitor through above-mentioned Epstein-Barr virus-associated gastric cancer cells and PDX model, and promoting the intervention plan to split the virus;(4)By establishing anthropogenic tumor-burdened model in mice, we'll evaluate the treatment with NK cells、EB virus's speciality in recognition vaccine-related to PD - L1 antibody and curative effect of double drug combination. The study will provide accurate treatment for EB virus associated gastric cancer, effectively solve problems about high heterogeneity and poor effects and explore new pathways for tumor individualized treatment.

胃癌具有比其它肿瘤更明显的异质性，探索胃癌个体化治疗尤为重要。TCGA分子分型将胃癌分为4类，其中EBV相关胃癌有PI3KCA高频突变，CDKN2A甲基化，JAK2、PD-L1、PD-L2扩增，具有EBV来源特异性抗原表达特点，是免疫治疗极佳切入点，目前尚无反映该类细胞特征的研究模型与治疗策略。课题组前期建立了CTC检测平台及胃癌PDX研究模型，为此将1）结合肿瘤周边及外周血免疫细胞特征，分析预后相关免疫指标；2）基于原位肿瘤PDX模型及循环肿瘤细胞建立原代EBV相关胃癌模型；人源性免疫重建荷瘤小鼠模型；3）以上述EBV相关胃癌细胞和PDX模型筛选CDK4/6，JAK2等靶向抑制剂；促进病毒产生裂解的干预方案；4）以小鼠模型评价NK细胞治疗，EBV特异识别DC疫苗联合PD-1抗体的叠加疗效。本研究将为EBV相关胃癌提供精准治疗，有效解决胃癌高异质、疗效差难题，为肿瘤个性化治疗探索新途径。

2014年，癌症基因组图谱计划（TCGA）将胃癌分子分型为4类：EBV相关型，微卫星不稳定型，基因学稳定型和染色体不稳定型。其中EB病毒（EBV）阳性的胃癌约占10%。然而，EBV相关胃癌研究尚有诸多问题尚未解决。本项目针对现阶段（1）缺乏EBV+胃癌预后与机体免疫反应相互关系的研究和（2）缺乏直接反映EBV+胃癌疗效的体内检测与评估手段等两大问题，进行了系统性研究并取得以下进展：（1）我们对肿瘤生物学国家重点实验室组织样本库中132例胃癌组织通过原位杂交方法（临床诊断金标准）筛选出5例EBV+的胃癌患者。我们对EBV+胃癌患者和EBV-胃癌患者组织提取RNA并进行RNA-Seq以比对其组织的免疫学特性。我们发现EBV+胃癌患者组织中Th1、Th17、Treg、CD8+ cytotoxic T比例明显高于EBV-胃癌患者组织。通过对EBV+胃癌患者和EBV-胃癌患者组织的表达谱芯片分析，我们发现EBV+胃癌患者组织中CXCL9、CXCL10和CXCL11表达明显高于EBV-胃癌患者组织，以上差异分子均富集在chemokine信号通路，提示EBV感染可能主要通过与CXCR3相互作用，分泌相关细胞因子以招募免疫细胞，进而杀伤肿瘤细胞。（2）为进一步在体内验证以上免疫特性，我们正利用EBV+的胃癌患者组织分别构建PDX免疫缺陷荷瘤小鼠模型和人源性免疫重建荷瘤小鼠模型，目前已构建成瘤6例EBV+的胃癌PDX免疫缺陷荷瘤小鼠模型。本项目将为EBV+的胃癌患者评估靶向药物及免疫治疗的疗效并探索其分子机制提供重要的实验依据和研究模型。