LEPRE1联合PD-L1免疫治疗生物标志物对不同亚型食管鳞癌患者免疫疗效及预后评估的价值

Value of LEPRE1 combined with programmed cell death 1 ligand 1(PD-L1)immunotherapy biomarkers in evaluating the immune efficacy and prognosis of patients with different subtypes of esophageal squamous cell carcinoma

目的评估LEPRE1联合PD-L1免疫治疗生物标志物对不同亚型食管鳞癌免疫疗效及预后价值。方法以2011年-2017年新疆医科大学附属肿瘤医院胸外科收治的125例食管鳞癌(Esophageal squamous cell carcinoma,ESCC)患者为研究对象,采用免疫组化方法测定LEPRE1、PD-L1和CD8+T的蛋白表达水平,肿瘤综合阳性评分(CPS)确定三者共表达状态并将患者进行特征分组;在ESCC患者队列的mRNA表达转录组数据集中,通过相关性分析来研究LEPRE1和PD-L1及相关免疫检查点抑制剂之间的关系;利用R中“limma”包对转录组数据进行临床病理特征和基因突变分析;通过基因本体(Gene Ontology,GO)富集和通路GSEA(Gene Set Enrichment Analysis)富集和肿瘤微环境中免疫检查点抑制剂、免疫调节因子、免疫细胞的分析,探究LEPRE1和PD-L1不同表达的分组可能参与调控的信号转导通路;通过免疫治疗反应在不同免疫治疗队列中验证联合分组的临床免疫治疗应答效果以及临床预后。结果在125例ESCC患者的转录组数据集中根据LEPRE1和PD-L1的表达将患者分为4个亚组,LEPRE1高表达亚组、PD-L1高表达亚组、LEPRE1和PD-L1双高表达亚组和双低表达亚组;CPS分组免疫组化结果初步显示PD-L1高表达亚组与CD8免疫细胞浸润呈正相关;相关性分析结果显示免疫检查点抑制剂与PD-L1成显著正相关(P<0.01);临床病理特征和基因突变相关性分析结果显示LEPRE1高表达亚组更易出现LRP2突变(P<0.1),而PD-L1高表达亚组中更易出现RELN和LRP1B类型的基因改变(P<0.1);GO富集和GSEA富集分析结果表明LEPRE1高表达亚组参与调控ECM相关通路,而PD-L1高表达亚组则在免疫反应调节等通路富集;肿瘤微环境免疫相关分子特征结果显示,与LEPRE1高表达亚组相比,PD-L1高表达亚组的免疫检查点抑制剂、免疫调节因子和免疫细胞指标升高,差异具有统计学意义(P<0.05);临床免疫反应应答结果显示,PD-L1高表达亚组中的临床患者免疫反应应答比例显著高于其余分组且患者预后更佳(P=0.04),新生抗原和肿瘤突变负荷所反应的免疫治疗效果也显著优于其余分组(P<0.05);在GSE91061、RCC Braun、PRJEB23709免疫治疗数据集中,PD-L1高表达亚组预后良好,差异具有统计学意义(P<0.05)。结论LEPRE1与PD-L1联合免疫生物标志物模式有助于将患者分为不同的ESCC亚型,可以实施更精准的治疗方案提高免疫治疗疗效,为ESCC诊断和治疗提供了新思路。

Objective To evaluate the immune efficacy and prognostic value of LEPRE1 combined with pro-grammed cell death 1 ligand 1(PD-L1)immunotherapy biomarkers in different subtypes of esophageal squamous cell carcinoma.Methods A total of 125 patients with esophageal squamous cell carcinoma(ES-CC)treated in the hospital from 2011 to 2017 were selected as subjects.The protein expression levels of LEPRE1,PD-L1 and CD8+T were determined by immunohistochemical method,and the co-expression status of LEPRE1,PD-L1 and CD8+T was determined by tumor comprehensive positive score(CPS).In the mRNA expression transcriptional group data set of the cohort of ESCC patients,the relationship be-tween LEPRE1,PD-L1 and related immune checkpoint inhibitors was studied by correlation analysis,and the clinicopathological characteristics and gene mutation of the transcriptional group data were analyzed by"limma"package in R.Through the enrichment of gene ontology(GO)and pathway of Gene Set Enrich-ment Analysis(GSEA)and the analysis of immune checkpoint inhibitors,immunomodulatory factors and immune cells in tumor microenvironment,this study deeply explored the signal transduction pathways that may be regulated by different expression groups of LEPRE1 and PD-L1,and verify the clinical immuno-therapy response effect and clinical prognosis of combined groups in different immunotherapy cohorts.Results According to the expression of LEPRE1 and PD-L1,the patients with ESCC were divided into 4 subgroups:LEPRE1 high expression subgroup,PD-L1 high expression subgroup,LEPRE1 and PD-L1 double high expression subgroup and double low expression subgroup.The immunohistochemical results of CPS group showed that there was a positive correlation between PD-L1 high expression subgroup and CD8 immune cell infiltration.The results of correlation analysis showed that there was a significant posi-tive correlation between immune checkpoint inhibitors and PD-L1(P<0.01).The results of correlation analysis between clinicopathological features and gene mutation showed that LRP2 mutation was more likely to occur in high LEPRE1 subgroup(P<0.1).However,genetic changes of RELN and LRP1B types were more likely to occur in the high PD-L1 subgroup(P<0.1).The results of GO enrichment and GSEA enrichment analysis showed that the high LEPRE1 subgroup was involved in the regulation of ECM related pathway,while the PD-L1 high expression subgroup was enriched in immune response regulation and other pathways.The results of immune-related molecular characteristics in tumor microenvironment showed that the immune checkpoint inhibitors,immunomodulatory factors and immune cell indexes in PD-L1 high expression subgroup were significantly higher than those in PD-L1 high expression subgroup(P<0.05).The results of clinical immune response showed that the proportion of clinical patients in the sub-group with high expression of PD-L1 was significantly higher than that in other groups,and the prognosis was better(P=0.04).The immunotherapeutic effect of new antigen and tumor mutation load was also significantly better than that of the other groups(P<0.05).In the GSE91061,RCC Braun and PRJEB23709 immunotherapy data sets,the prognosis of the subgroup with high expression of PD-L1 was good,and the difference was statistically significant(P<0.05).Conclusion The immunophenotyping of LEPRE1/PD-L1 co-expression as a biomarker is helpful to classify patients into different ESCC subtypes,improve the efficacy of immunotherapy with more accurate treatment,and provide new ideas for the diag-nosis and treatment of ESCC.