摘要
目的探讨安罗替尼联合抗程序性死亡受体-1(抗PD-1)单抗通过抑制血管内皮生长因子/原癌基因(VEGF/c-Kit)调控Wnt/β-catenin通路来抑制肺腺癌的可能机制。方法建立C57小鼠荷瘤模型,并分为阴性对照组、安罗替尼组、抗PD-1单抗组和安罗替尼联合抗PD-1单抗组,每组8只。连续给药25 d后,比较各组小鼠肿瘤体积的变化。采用基因富集分析(GSEA)探讨癌症基因图谱(TCGA)数据库中肺腺癌(LUAD)队列中c-Kit和VEGF富集的相关通路;采用CIBERSORTx分析c-Kit和VEGFR对22种免疫细胞浸润的影响;分析c-Kit和VEGFR与免疫相关分子的相关性。采用细胞转染敲低c-Kit表达并分组,对照组为正常LLC细胞,si-c-Kit组为siRNA敲低c-Kit表达,安罗替尼处理组为安罗替尼处理LLC细胞。采用Western Blot实验检测各组细胞c-Kit、Wnt1、β-catenin、干扰素-γ(IFN-γ)、肿瘤坏死因子-α(TNF-α)、PD-L1、c-Myc、c-Jun表达。结果与阴性对照组比较,安罗替尼组、PD-1单抗组、安罗替尼联合PD-1单抗组均可抑制肿瘤的生长,差异有统计学意义(P<0.05);与安罗替尼组、PD-1单抗组比较,安罗替尼联合PD-1单抗组可抑制肿瘤的生长,差异有统计学意义(P<0.05)。GSEA表明,IFN-γ通路富集在c-Kit低表达样本中,Wnt/β-catenin通路富集在c-Kit高表达样本中,血管生成及IL-6/JAK/STAT3通路富集在VEGFR高表达样本中,差异均有统计学意义(P<0.05)。CIBERSORTx分析发现,c-Kit高表达样本中有更少的抗肿瘤免疫细胞以及更多的免疫抑制细胞,差异有统计学意义(P<0.05)。VEGFR低表达样本中有更多的抗肿瘤免疫细胞以及更少免疫抑制细胞,差异有统计学意义(P<0.05)。c-Kit与颗粒酶A(GZMA)、颗粒酶B(GZMB)、IFN-γ等促免疫分子呈负相关(r<0,P<0.05),与CD274(PD-L1)、转化生长因子β(TGF-β)、T细胞免疫球蛋白结构域和粘蛋白结构域-3(TIM3)等免疫抑制分子呈正相关(r>0,P<0.05)。VEGFR与GZMA、TNF-α、GZMB、IFN-γ等免�
Objective To explore the possible mechanism of arotinib combined with anti-programmed death receptor-1(anti-PD-1)in inhibiting lung adenocarcinoma by regulating Wnt/β-catenin pathway through inhibiting vascular endothelial growth factor/c-Kit(VEGF/c-Kit).Methods A lung adenocarcinoma model was established in C57 mice,and the mice were divided into negative control group,arotinib group,anti-PD-1 monoclonal antibody group and arotinib combined with anti-PD-1 monoclonal antibody group,with 8 mice in each group.After continuous administration for 25 days,the change in tumor volume in each group was compared.Gene Set Enrichment Analysis(GSEA)was used to explore the pathways related to the enrichment of c-Kit and VEGF in the lung adenocarcinoma(LUAD)cohort of the Cancer Genome Atlas(TCGA)database;the CIBERSORTx was used to analyze the influence of c-Kit and VEGFR on the infiltration of 22 types of immune cells;the correlations of c-Kit and VEGFR with immune-related molecules were analyzed.Cell transfection was used to knock down c-Kit expression and the cells were divided into groups,the control group was normal LLC cells,the si-c-Kit group was siRNA knocked down c-Kit,and the arotinib treatment group was LLC cells processed with arotinib.Western Blot assay was used to detect expressions of c-Kit,Wnt1,β-catenin,gamma interferon(IFN-γ),tumor necrosis factor-alpha(TNF-α),PD-L1,c-Myc and c-Jun.Results Compared with the negative control group,tumor growth was significantly inhibited in the arotinib group,PD-1 monoclonal antibody group,and the combination of arotinib and PD-1 monoclonal antibody group(P<0.05).Compared with the arotinib group and PD-1 monoclonal antibody group,the tumor growth was significantly inhibited in the combination of arotinib and PD-1 monoclonal antibody group(P<0.05).GSEA analysis showed that the IFN-γpathway was enriched in c-Kit low expression samples,Wnt/β-catenin pathway was enriched in c-Kit high expression samples,while the angiogenesis and IL-6/JAK/STAT3 pathway were enriched in VEGFR
作者
周泽军
谢海燕
张宇
石文
夏宁
ZHOU Zejun;XIE Haiyan;ZHANG Yu;SHI Wen;XIA Ning(the First Department of Respiratory,Chest Department Area of Nanjing Brain Hospital,Former Nanjing Chest Hospital,Nanjing,Jiangsu,210029)
出处
《实用临床医药杂志》
2023年第17期51-56,共6页
Journal of Clinical Medicine in Practice
基金
2017年度南京市医学科技发展资金一般性课题项目(YKK17185)。
