HLA基因多态性介导的高危型EB病毒慢性感染及其交互作用与鼻咽癌发病关系的前瞻性流行病学研究

Genetic and environmental factors are causally related to nasopharyngeal carcinoma(NPC).We have identified that Human leukocyte antigen (HLA) polymorphisms, EBV chronic activation and high-risk EBV subtype (RPMS1 G>A variation) were the risk factors of NPC and also found that HLA genotype had related with different EBV subtype infections. Thus, we put forward the hypothesis that high-risk HLA genotype may interact and mediate EBV chronic infection and EBV high-risk subtype infection, and HLA high-risk genotypes collaborate with EBV to induce the occurrence of NPC. However, the hypothesis still has not been confirmed due to lack of epidemiological evidences. Based on the established NPC screening cohort (60000 participants) and the EBV different reactivation cohort (1800 participants), we plan to carry out a multi-phase study to explore the synergy function for NPC between HLA genetic and EBV infection. Genotyping of HLA SNPS and EBV high-risk SNP in saliva are tested using the Agena Bioscience Mass Array platform, EBV DNA and genome expression in the local nasopharynx are detected with real-time PCR and gene expression profile chip, respectively. Causal mediation analysis methods are used to perform joint interaction and mediation analysis of the effects of HLA genetic variants and EBV high-risk SNP or EBV chronic infection on NPC. Specifically, we decompose the total effect of each HLA genetic variant in the possible presence of the EBV genetic variant and EBV chronic infection on NPC into four components, with which the host variant may interact. With the longitudinal data analysis model, the affection for EBV high-risk subtype infection of nasopharyngeal mucosal and genome expression will be analyzed. This study will provide the prospective evidence for revealing the genetic-variant-by-EBV interaction and EBV mediation of genetic effects on NPC risk and help to illustrate the EBV high-risk subtype natural infection history and carcinogenic mechanism.

遗传和环境因素在鼻咽癌（NPC）发病中起重要作用。我们已证实人类白细胞抗原（HLA）基因多态性、EBV慢性激活和EBV高危亚型（RPMS1基因G>A突变）感染是其危险因素。还发现二者间存在相关关系。因此提出HLA介导EBV慢性感染和选择感染高危型EBV两条通路而致癌,并产生交互作用。但该假说尚缺乏流行病学证据。在前期建立6万NPC筛查及0.18万 EBV不同再激活人群队列基础上，拟开展多阶段研究。质谱技术检测白细胞HLA易感基因位点和唾液EBV亚型，定量PCR和基因表达谱芯片检测鼻咽脱落细胞EBVDNA和基因组表达。结合病因中介模型分析HLA-EBV协同作用的超额发病风险并分解其中的交互和中介作用大小。运用纵向数据模型分析EBV高危型在鼻咽粘膜癌变前感染规律及其对基因组影响。本研究为阐明基因-病毒交互作用与NPC发病关系提供前瞻性证据，有助揭示EBV高危亚型感染自然史及其致癌机制。

本研究首先开展了基于鼻咽癌高发区的病例对照设计，利用多重扩增子捕获技术检测已报道的鼻咽癌易感基因，多因素分析发现位于HLA-DQA1基因3’非翻译区的rs1064994和HCG9基因区域的rs5009448碱基变异与鼻咽癌发病风险相关。并在一个EBV不同再激活水平的人群队列中发现此两个易感位点与EBV再激活水平降低相关，得出此两个易感基因通过介导EB病毒再激活水平降低鼻咽癌发病风险的结论。进而通过中介效应分析发现，位点rs1064994对鼻咽癌的总超额相对风险度为-0.61，其中基因-病毒交互效应为-0.20，占比33.34%；中介效应为-0.26，占比42.48%。rs5009448对鼻咽癌的总超额相对风险度为-0.29，基因病毒交互效应为-0.17，占比57.86%，二者的中介效应为-0.07，占比25.26%。在不同再激活水平的队列人群中，利用飞行质谱方法检测唾液标本中的EB病毒亚型，EB病毒的高危亚型（BALF2基因162215 C>A，162476 T>C和163364C>T）与EB病毒的再激活水平无显著关联。对两阶段病例对照研究的关联分析和检测EB病毒高危亚型的人群中基因关联分析进行meta分析发现，位点rs3095341，rs4122190，rs9366775和rs4999716是潜在的鼻咽癌与高危EB病毒亚型共同差异位点。最终，对不同EB病毒再激活水平的队列人群进行随访，发现不同EB病毒再激活水平的人群中EB病毒DNA含量存在着显著的差异，证实EB病毒的再激活导致鼻咽原发部位EB病毒负荷增加及感染上皮细胞机会增加的病因学结论。本研究为阐明基因-病毒交互作用与鼻咽癌发病关系提供前瞻性证据，分析了基因-病毒交互作用和中介效应的大小，为鼻咽癌的防治提供了重要的流行病学证据和病因学证据。整个项目在SCI杂志发表论文11篇，申请专利2项，培养研究生4名。