Effect of N-linked glycosylation removal on PD-L1 expression in esophageal squamous cell carcinoma and metastatic lymph nodes
Objective To investigate the effect of N-linked glycation modification on the expression of programmed death ligand 1(PD-L1)in esophageal squamous cell carcinoma tissue and its relationship with clinical and pathological features.Methods PD-L1 22C3 antibody was used to perform immunohistochemical staining on esophageal squamous cell carcino-ma tissue chips,MCF-7,and NCI-H226 cell sections after standard treatment or peptide N-glycosidase F(PNGase F)treatment,respectively.Tumor cell proportion score(TPS),combined positive score(CPS),or histological score(H-score)were used to evaluate the expression level of PD-L1 in tissues and cell sections.TPS 1％and CPS 10 were defined as PD-L1 positive expression.The consistency of PD-L1 expression between the primary lesion and metastatic lymph node tissue was analyzed;The relationship between PD-L1 expression in the primary lesion tissue and demographic and clinical pathological characteristics was analyzed.Results Among the 50 patient tissue samples,24 normal esophage-al mucosal tissues,48 primary lesion tissues,and 32 metastatic lymph node tissues were eligible for PD-L1 expression e-valuation.After standard treatment,PD-L1 expression was negative in normal esophageal mucosal tissue;The positive rates of PD-L1 TPS,CPS,and median H-score in the primary lesion tissue were 50.00％,29.17％,and 1.41 points,re-spectively;The positive rates of PD-L1 TPS,CPS,and median H-score in metastatic lymph node tissue were 50.00％,68.75％,and 2.02 points,respectively.Five cases of normal esophageal mucosal tissue showed focal PD-L1 expression after PNGase F treatment;The positive rates of PD-L1 TPS,CPS,and median H-score in the primary lesion tissue were 56.25％,52.08％,and 3.55 points,respectively;The positive rates of PD-L1 TPS,CPS,and median H-score in meta-static lymph node tissue were 59.38％,81.25％,and 10.33 points,respectively.After PNGase F treatment,the PD-L1 TPS,CPS,and H-score of the primary lesion and metastatic lymph node tissue were higher than those of the standard treatment,all P＜0.001.The PD-L1 TPS and H-score of NCI-H226 cells treated with PNGase F were higher than those of the standard treatment,with t values of-18.087 and-33.588,and P values of 0.003 and 0.001,respectively.The consistency of PD-L1 TPS and CPS positive expression in the primary lesion and its metastatic lymph node tissues of 32 patients after standard treatment and PNGase F treatment was poor.The proportion of PD-L1 TPS and CPS positive pa-tients in the primary lesion and/or metastatic lymph node tissue after PNGase F treatment was 78.13％and 87.50％,re-spectively.The TPS positive rate of PNGase F treatment combined with detection of primary lesion tissue was higher than that of standard treatment,x2=5.497,P=0.019;The CPS positive rate was higher than the standard treatment,with=23.127 and P＜0.001.There were no statistically significant differences in PD-L1 TPS and CPS positivity rates a-mong different genders,ages,maximum tumor diameter,pathological grading,N-stage,and primary lymph node metas-tasis tissues,with P＞0.05.Conclusions PNGase F treatment can enhance the staining intensity of PD-L1 22C3 antibod-y.PD-L1 expression exhibits heterogeneity between the primary and metastatic lymph node tissues of esophageal cancer.PNGase F treatment of esophageal cancer tissue while evaluating both primary and metastatic lymph node tissues can im-prove the PD-L1 positive detection rate.
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