基于治疗前及早期宿主基因表达谱研究慢性丙型肝炎患者对IFN/RBV应答差异的机制

The mechanisms of distinct response to Interferon and Ribavirin treament in Chronic Hepatitis C patients are not yet clear. Our exploratory study based on early gene expression profiles of PBMCs showed that carcinoembryonic antigen-related cell adhesion molecule 8 (CEACAM8) was significantly associated with viralogic response. For patients without achieving sustained virologic response (NonSVR), the level of CEACAM8 mRNA is 13.8-fold higher than sustained virologic response patients at pretreatment, and its temporal change pattern also exhibits significant difference between two groups. In addition,the differentially expressed genes between NonSVR and SVR patients at pretreatment include a cluster of genes involved in response to other organism, such as DEFA1B and DEFA3. One thing deserved to be mentioned is that they were the same to CEACAM8, with a significantly higher expression level in NonSVR patients compared with SVR patients. This may suggest that the high expression of defense system related genes before treatment are unfavorable to HCV clearance. This study aims to increase sample size to validate the findings. On the other hand, increasing the sample size and enhancing the corresponding statistical power may reveal more significant genes and pathways. This study not only can provide prediction factors for nonresponser at early phase, making them avoid the various side effects and waste of cost by treatment, but also can provide theoretical explaination for the distinct response.

慢性丙型肝炎患者对干扰素联合利巴韦林治疗应答个体差异的机制尚未完全明确。前期工作中，我们基于治疗早期外周血单个核细胞全基因组mRNA表达谱，发现癌胚抗原相关细胞粘附因子8(CEACAM8)的表达水平与病毒学应答显著相关：未获得持续病毒学应答(NonSVR)患者治疗前CEACAM8表达是获得持续病毒学应答（SVR）患者的13.8倍，表达动力学也存在显著差异。治疗前差异表达基因还包含一簇对其他物种应答基因(DEFA1B, DEFA3等)，且同CEACAM8，均表现出在NonSVR患者的表达水平显著高于SVR患者。提示治疗前防御系统相关基因的高表达反而可能是造成疗效不佳的潜在机制。本项目拟扩大样本量，对结果证实。同时样本量的增加，统计效能的提高，可能发现更有意义的基因及通路。这不仅能为进一步阐明个体应答差异机制提供理论依据,也能为无应答提供早期预测因子,避免治疗带来的各种副作用及成本上的浪费。

慢性丙型肝炎患者对干扰素治疗应答个体差异的机制尚未完全明确。新近，IFNL4 ss469415590(TT/ΔG)被证明相比于rs12979860对干扰素疗效影响更显著。我们对中国汉族人群，447位慢丙肝患者（含328位接受干扰素联合利巴韦林治疗），129位自发清除者及169位健康对照进行了ss469415590分型。自发清除者携带TT/TT基因型比例显著高于慢丙肝患者(95.3% vs. 88.8%, P=0.027)。1b型患者ss469415590分型与SVR独立相关(OR=3.247，95%CI=1.038–10.159, P=0.043)，与治疗过程中病毒学应答（RVR，cEVR，EVR及ETVR）独立相关，最小OR为3.73。特别对于高病毒载量患者（≥4×105IU/ml），携带ΔG等位基因的患者获得SVR的可能性极低(7.1%，阴性预测值92.9%)。但在2a型患者，未发现此位点与疗效间的相关性；而肝功是最显著因素。AST>70IU/L可获得100%SVR。与AST≤35IU/L患者比，提高了37.0%。ss154949590与rs12979860位点完全连锁不平衡。总之，ss154949590 TT/TT基因型利于自发清除及1b型患者干扰素治疗后病毒学应答，但与2a型无关。高病毒载量1b型不推荐用干扰素治疗，DAA药物可能是更好的选择。2型患者若AST>70，干扰素可成为一个选择。在转录水平研究与应答相关基因，检测了干扰素治疗极早期PBMC全基因组表达水平。发现基线CEACAM8的表达水平与应答显著相关。为了验证结果，我们又筛选了30位应答者和20位无应答者，对PBMC进行PCR验证，但目前尚未完成该检测，无定论性结论。通过基因表达谱建立基因之间的共表达网络，发现被干扰素诱导基因在不同时间点基本形成两个簇，一簇为以ISG15等为代表的ISGs，一簇为炎性细胞因子趋化因子。这两促基因之间通过几个关键的分子连接（TLR7，CCL20，MIR155HG等），重要的是两簇之间呈现负相关关系。这可能提示ISG15等干扰素刺激基因不仅发挥清除病毒的作用，还有抑制炎性细胞因子趋化因子表达的作用。DAA疗效显著。但耐药变异影响疗效。对NCBI核酸数据库病毒株和吉林省地区38例慢丙肝患者进行病毒序列测序分析，发现耐药相关位点NS5A 区Y93H流行率显著高于全球。