利用微滴数字PCR进行BRAF V600E突变与PD-L1表达的相关性分析  

Correlation between BRAF mutation and PD-L1 expression using droplet digital PCR

作　　者：段芳芳 樊金星 易丽君 徐红艳[2] 杨文萍[2] 杜香平 李红[1] DUAN Fang-fang;FAN Jin-xing;YI Li-jun;XU Hong-yan;YANG Wen-ping;DU Xiang-ping;LI Hong(Central Laboratory,Nanchang University Affiliated Children’s Hospital/Jiangxi Provincial Children’s Hospital,Nanchang 330006,China;Department of Pathology,Nanchang University Affiliated Children’s Hospital/Jiangxi Provincial Children’s Hospital,Nanchang 330006,China;Department of Orthopedics,Nanchang University Affiliated Children’s Hospital/Jiangxi Provincial Children’s Hospital,Nanchang 330006,China)

机构地区：[1]南昌大学附属儿童医院/江西省儿童医院中心实验室,南昌330006 [2]南昌大学附属儿童医院/江西省儿童医院病理科,南昌330006 [3]南昌大学附属儿童医院/江西省儿童医院骨科,南昌330006

出　　处：《临床与实验病理学杂志》2022年第7期826-830,共5页Chinese Journal of Clinical and Experimental Pathology

基　　金：江西省重点研发计划(20202BBGL73035);江西省卫健委科技计划(202110100);江西省中医药科技计划(2020A0016)。

摘　　要：目的探讨BRAF V600E突变与PD-L1表达的相关性及其表型特征。方法采用免疫组化EnVision两步法对34例朗格汉斯细胞组织细胞增生症(Langerhans cell histiocytosis,LCH)组织中PD-L1的表达进行检测;qRT-PCR和微滴数字PCR(droplet digital PCR,ddPCR)技术对BRAF V600E表达水平进行定量并比较两种方法的一致性和灵敏度;分析BRAF V600E携带者的临床病理学特征。结果qRT-PCR法检测LCH患者的BRAF V600E突变率为76.47%(26/34),ddPCR法为82.35%(28/34),两种方法检测的突变率一致性高(Kappa=0.82,P<0.001),且ddPCR灵敏度更高;BRAF V600E高表达者的PD-L1表达水平亦增加(r=0.44,P<0.05);BRAF V600E携带者累及多个重要脏器受损。结论采用qRT-PCR、ddPCR法检测LCH组织中BRAF V600E突变率一致性高,且ddPCR法具有更高的灵敏度;BRAF V600E与PD-L1表达呈正相关。Purpose To investigate the correlation between BRAF V600E mutation and PD-L1 expression and to summarize BRAF variant’s clinical characteristics.Methods Paraffin-embedded tissue samples were retrospectively collected from 34 Langerhans cell histiocytosis(LCH)patients.Immunohistochemistry of EnVision two-step method was applied to detect PD-L1 expression and quantitative analysis was performed.BRAF V600E gene expression levels were quantified by qRT-PCR and droplet digital PCR(ddPCR)techniques,and the consistency and sensitivity of the two quantitative methods were compared.The clinicopathological features of BRAF V600E carriers were analyzed retrospectively at the same time.Results The detection rate of BRAF V600E by qRT-PCR was 76.47%(26/34),while ddPCR was 82.35%(28/34),the two methods showed a very high consistency(Kappa=0.82,P<0.001)and the sensitivity of ddPCR was significant.Patients with enormous BRAF V600E expression also showed increased PD-L1 expression(r=0.44,P<0.05).BRAF V600E carriers tended to have multiple high-risk organs and tissues damaged.Conclusion BRAF V600E gene detection in children’s LCH adopts qRT-PCR and ddPCR methods with good consistency and ddPCR method has better sensitivity.BRAF V600E and PD-L1 expression display a positive correlation.

关 键 词：朗格汉斯细胞组织细胞增生症 微滴数字PCR BRAF V600E PD-L1 分子诊断 

分 类 号：R733[医药卫生—肿瘤]