| Background and objectiveThe long-term survival of the esophageal squamous cell carcinoma(ESCC)patients with the local middle and late stages is related to the pathological response after neoadjuvant chemoradiotherapy(NCRT).However,the lack of effective biomarkers to predict the pathological response of ESCC patients after NCRT is a long-standing problem,so a comprehensive and systematic analysis focusing on efficacy of chemoradiotherapy and interactions of these genes in ESCC will provide valuable insights to understand the molecular features.Methods1.we collected genes potentially associated with efficacy of chemoradiotherapy by screening publications on genetic association studies deposited in PubMed(https://www.ncbi.nlm.nih.gov/pubmed).The major biological themes linked with these genes were then revealed by function and biochemical pathway enrichment analysis.2.In the context of protein-protein interaction,a specific protein subnetwork involving genes related to the efficacy of radiotherapy and chemotherapy was constructed using GenRev,and some new genes outside the candidate gene set would be obtained.3.In Kaplan-Meier Plotter online resources(http://kmplot.com/analysis/),we can analysis the correlations between new gene expression and prognosis of ESCC patients.4.MMS22L gene mRNA and protein expression in ESCC cancer tissue,adjacent normal tissue,Kyse150,Eca109,TE-1 and HEEC cell lines were detected by qRT-PCR and Western blot.5.Immunohistochemical staining(IHC)was used to detect MMS22L protein expression in ESCC tissue from endoscopic biopsies.Image-Pro Plus 6.0 was used to quantify the protein expression level of MMS22L.6.The predictive performance of biomarkers was estimated using receiver operating characteristic(ROC)curve analysis.7.After the expression of MMS22L was knocked down,the metastasis and invasive ability of TE-1 cells was detected by Transwell assay.8.After the expression of MMS22L was knocked down,the proliferation ability of TE-1 cells under different drug gradients was measured by CCK8 assay,and the IC50 value of 5-FU was calculated.Results1.We compiled 101 human genes reported to be associated with chemoradiotherapy efficacy by reading the full text of 79 articles from 217 publications collected.Enrichment analysis indicated that collected genes were associated with biochemical pathway related to Platinum Resistance,Ribosomes,Pancreatic Cancer,HIF-1 Signaling Pathway,DNA Repair Complexes,etc.2.A specific protein network of 105 genes related to the efficacy of radiotherapy and/or chemotherapy in ESCC was inferred.Eight new genes potentially associated with chemoradiotherapy efficacy in ESCC were identified,such as CUL3,MUC13,MMS22L,MME,UBC,VAPA,CYP1B1 and UGDH.3.Database analysis shows that ESCC patients with higher mRNA levels of MMS22L(HR=0.33,CI:0.14-0.75,logrank P=0.0052)and MUC 13(HR=0.37,CI:0.14-0.96,logrank P=0.033)had higher overall survival,and ESCC patients with lower mRNA levels of VAPA(HR=2.32,CI:1.04-5.17,logrank P=0.035),CYP1B1(HR=2.67,CI:1.02-6.97,logrank P=0.039)and UBC(R=2.26,CI:1.03-4.96,logrank P=0.038)had higher overall survival,while mRNA expression of UGDH,MME and CUL3 was not associated with ESCC patient survival.4.MMS22L mRNA expression was significantly decreased in 23 ESCC tissues with only undergoing surgery compared with adjacent non-cancerous tissue samples(P<0.01),and Western blot were consistent with the RT-qPCR.5.Except lymph node metastasis(P<0.01),other clinicopathological characteristics genes were not associations with MMS22L protein expression.6.MMS22L protein in responding group(CR plus PR,29 patients)was significantly higher than that in the non-responding groups(SD plus PD)(P<0.01).ROC curve analyses indicated that MMS22L yielded an AUC of 0.847(95%CI:0.7232 to 0.9703;P<0.01)with 100%sensitivity and 65.52%specificity for predicting the efficacy of NCRT in patients with ESCC.7.The mRNA and protein expression of MMS22L in Kyse150,Eca109 and TE-1 was lower than that HEEC,and the difference was statistically significant(P<0.01).8.After knocked down the expression of MMS22L in TE-1 cells,transwell assay results indicated that the invasion and metastasis ability of TE-1 cells increased(P<0.01).9.After knocked down the expression of MMS22L in TE-1 cells,CCK8 assay results showed that the IC50 of 5-FU in TE-1 cells was increased,indicating that TE-1 with low MMS22L were more resistant to 5-FU.ConclusionsSpecific molecular network associated with efficacy of chemoradiotherapy was inferred and some potential related genes were identified in ESCC.MMS22L was significantly downregulated in cancer tissues compared with normal controls,and low MMS22L expression was correlated with Lymph node metastasis in ESCC.Importantly,MMS22L might serve as a biomarker for resistance to NCRT in ESCC. |
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