Study On CYTL1 And Systemic Immune Index In Predicting Prognosis Of Locally Advanced Cervical Squamous Carcinoma With Concurrent Chemotherapy

In the 1990s,in order to explore the treatment of locally advanced cervical cancer(LACC),the gynecological oncology community in the United States made useful explorations and published research results based on 5 large-sample randomized controlled trials in 1999.Compared with radiotherapy alone,concurrent chemoradiotherapy based on cisplatin could significantly improve the survival rate of patients and reduce the mortality rate by 30%～50%.These were the five famous trials on the treatment of locally advanced cervical cancer.And the treatment of concurrent radiotherapy and combined chemotherapy had been confirmed as the treatment mode of locally advanced cervical cancer.However,the treatment effect was still unsatisfactory.In recent years,people had explored targeted and immunotherapy combined with concurrent radiotherapy and chemotherapy to treat LACC,looking forward to better clinical treatment effects.However,looking up the literature we found that there was a lack of economic prediction models for predicting the effects of LACC concurrent radiotherapy and chemotherapy.Advances in high-throughput experimental technology and bioinformatics could sensitively detect DNA methylation and chromatin changes,and not only for tumor and other tissue biopsy methods,but also for non-invasive detection methods,for example,body fluid testing.Of course,people were looking for more ways.However,the prediction model based on genetic testing to predict the effect of LACSC concurrent radiotherapy and chemotherapy had not been reported yet.And most studies focused on early cervical cancer or patients with distant metastases.Another way,most studies did not distinguish pathological types.So,we use regression method to formulate a prediction model for predicting the effects of concurrent radiotherapy and chemotherapy for locally advanced cervical cancer,in order to provide important information about patients at risk of treatment failure,and guide patients to choose reasonable treatment plans during initial treatment,and avoid unnecessary treatment harm.And finally,the treatment effect will be improved.The tumor microenvironment is considered to be an important determinant of tumor behavior.In the tumor microenvironment,the inflammatory response plays a key role in tumor occurrence and development.Inflammation and immune cell infiltration which involved in defense against injury and infection can also induce tumor progression and metastasis.These findings indicate that there is an inflammatory tumor microenvironment in the patient’s body,which affects the patient’s response to anti-tumor therapy.Cytokine-like 1(CYTL1),as an emerging inflammatory chemotactic factor,was first discovered in bone marrow and cord blood mononuclear cells about 10 years ago,and these cells carry CD34 surface markers.CYTL1 lacks the main structural features of classic chemokines.However,a study by Tomczak and Pisabarro showed that CYTL1 may adopt a chemokine fold similar to IL-8,similar to CCL2,and also has the necessary features in the three-dimensional structure of CYTL1to identify the chemokine receptor CCR2.Wang et al.,verified that CYTL1 had chemotactic activity,and CYTL1 seemed to have anti-inflammatory function.More and more evidence supports the fact that CYTL1 plays a key role in inflammatory diseases.It plays an important role in maintaining cartilage homeostasis,promoting immune homeostasis,and participating in the growth and metastasis of tumor cells.At present,there is no report on the relationship between CYTL1 and cervical squamous cell carcinoma.Recently,systemic inflammation indicators based on the counts of peripheral blood lymphocytes,neutrophils,and platelets are considered to be better indicators of local immune response and systemic inflammation,because they are important in many tumors(such as hepatocellular carcinoma).High prognostic value and postoperative studies have been conducted for patients with esophageal cancer,colorectal cancer,small cell lung cancer,and cervical cancer.However,PLR,NLR,LMR,and SII have not obtained systematic evaluation in the prognostic value of locally advanced cervical squamous cell carcinoma undergoing concurrent radiotherapy and chemotherapy.The purpose of this study is to establish a LACSC prognostic model through bioinformation analysis.The expression of CYTL1 and immune-infiltrating cells are analyzed by immunohistochemical assay to study their effect on the prognosis of LACC.We also compare the effects of systemic inflammation and local cervical inflammation-related markers on predicting the prognosis of LACSC patients with concurrent radiotherapy and chemotherapy,to explore a cheaper and more convenient and feasible clinical prediction model.We hope that this model can further guide the selection of initial treatment options for LACSC and avoid over-or under-treatment.PartⅠBioinformatics Study on the Relationship between CYTL1 and Locally Advanced Squamous Carcinoma and PrognosisMethods1.The public genetic testing data published in the GEO database was used as the main source of the search.According to the proposed inclusion and exclusion criteria,studies that meet the system evaluation criteria were screened to look for different genes.2.Cervical cancer cases with available clinical information and complete genomic m RNA expression data were obtained from the Cancer Genome Atlas(TCGA).Our study conducted univariate and multivariate COX regression based on DEGs to find differentially expressed genes related to survival and build risk models.3.Based on the risk model,patients meeting the retrieval requirements in TCGA were divided into high and low risk groups.By using the GISTIC 2.0 data in TIMER,differential expression of immune checkpoints and infiltration of immune cells in the two groups were evaluated.4.The gene of CYTL1 was selected.And we analyzed the correlation between tumor infiltrating immune cells and CYTL1 in TCGA.Results1.The GEO2R platform was used to screen the DEGs between the treatment response group(Control group)and the non-response group(N Control group)in the GSE56363 data set.A total of 1021 DEGs were found,of which 663 up-regulated and358 down-regulated.Fun Rich software was uesd to perform GO and KEGG analysis on DEGs in the LACSC control group versus the non-control group.The main signal pathways were:immune response,cytokine interaction,tumor signal pathway,chemokine signal pathway,platelet activation.2.Multivariate regression analysis showed that there were 15 DEGs related to survival,namely FSCN1,NDC1,TNFRSF18,SH3BP5,GPC1,MT3,FBLN5,IQSEC2,CYTL1,NDN,CKB,HIC1,KDELR2,RALGDS,SLCO3A1(Table 5,Figure 3).The risk model was established based on 15 DEGs as follows:PI=FSCN1*0.006063427+NDC1*0.104381469+TNFRSF18*0.019971714+SH3BP5*0.162984858+GPC1*0.009462412+MT3*0.03888099+FBLN5*0.055523035+IQSEC2*0.13623059+CYTL1*0.087325043+NDN*0.083397188+CKB*(-0.006571571)+HIC1*(-0.320144023)+KDELR2*0.006354518+RALGDS*(-0.032461904)+SLCO3A1*(-0.058777737).The time-dependent AUC curve suggested that the model was effective in predicting the outcome.The univariate and multivariate regression comparisons were made between the risk model and classical clinical indicators,and we found that the risk model was an independent prognostic factor Single-factor.3.Common immune checkpoints in high and low risk groups were analyzed according to the COX risk model,and the violin chart was used to display.The common immune checkpoint components were all adjusted up.There was no statistically significant in the expressions of CD274,CTLA4,HAVCR2,PDCD1LG2,and TIGIT between the two groups.The P values(95%CI)were 0.496(-0.15,0.08),0.109(-0.23,0.04),0.515(-0.18,0.09),0.871(-0.13,0.12),0.071(-0.24,0.02),respectively.And the expression of LAG3 and PDCD1 were statistically different,and the P values(95%CI)were 0.020(-0.28,-0.01),0.020(-0.27,-0.02).Percentage stacked bar graphs was used to show the percentage of 22 immune cell infiltrations of each patient in the high-and low-risk groups in the training set,validation set,and the entire population.It could be seen that there were differences in the CD8+T cells,CD4+T cells,mast cells infiltration of monocytes,and neutrophils between the high and low risk groups.The expression of CD8~+T cells increased in the low-risk group,P=0.002.The expression of silent CD4~+T cells increased in the high-risk group,P=0.015.The expression of monocytes increased in the high-risk group,P=0.016.The expression of silent mast cells increased in the low-risk group,P=0.019.The expression of activated mast cells increased in the high-risk group,P=0.001.The expression of neutrophils increased in the high-risk group,P=0.026.4.The correlation analysis between CYTL1 and tumor immune infiltrating cells was carried out by R software.The obvious correlation with 22 immune cells are B cells(Cor=0.182,P=0.009),activated dendritic cells(Cor=0.147,P=0.035),resting mast cells(Cor=0.189,P=0.006),activated natural killer cells(Cor=-0.146,P=0.035),CD4+activated memory T cells(Cor=-0.139,P=0.046),CD4+resting memory T cells(Cor=0.38,P=1.651e-08),CD8+T cells(Cor=-0.259,P=1.672e-04),and helper T cells(Cor=-0.287,P=2.823e-05).Conclusions1.As far as we know,this is the first study using bioinformatics to construct a prognostic model of cervical cancer.2.Tumor immune infiltrating cells and immune checkpoints affect the prognosis of cervical cancer.3.As an emerging inflammatory chemotactic factor,CYTL1 may become an indicator which affects the tumor immune infiltration level and prognosis of locally advanced cervical cancer.PartⅡAnalysis of CYTL1 expression in Locally Advanced Cervical Squamous CarcinomaMethods1.From october 2013 to december 2015,patients with locally advanced cervical squamous cancer who were hospitalized with radical concurrent chemoradiotherapy in the First Affiliated Hospital of Zhengzhou University were collected.And cervical squamous cell carcinoma was confirmed by pathological biopsy.All cases were approved by the Ethics Committee of the First Affiliated Hospital of Zhengzhou University.FIGO staging in 2009 was adopted.2.Immunohistochemical assay was used to detect the expression of CYTL1,the CD4~+T cells and CD8~+T cells infiltration in different cervical tissues were also detected.Then we analyzed the correlation between the expression of CYTL1 with the CD4~+T cells and CD8~+T cells infiltration.3.The correlation between CYTL1 and clinicopathological age was analyzed.The Kaplan-Meier univariate survival was used to analyze the impact of CYTL1 on overall survival(OS).Results1.Compared with the normal cervical tissue and Control group,the CYTL1nuclear staining depth was the strongest under the microscope in the N Control group(P=0.010).2.The expression of CYTL1 was negatively correlated with the infiltration of CD8~+T cells in the N Control group.3.The expression of CYTL1 in LACC had no significant correlation with T stage,N stage,tumor size or age.4.Survival analysis showed that the prognosis of high expression of CYTL1 was significantly worse than that of low expression of CYTL1.5.Survival analysis showed that the prognosis of CD8~+T cell high-level infiltration was better than low-level infiltration(P=0.036).The prognosis of high expression CYTL1 was significantly worse than that of low expression CYTL1(P=0.023).Conclusions1.CYTL1 is highly expressed in N Control group of LACC when patients received concurrent radiotherapy and chemotherapy.2.The expression of CYTL1 affecte the level of tumor immune infiltration,and have no obvious correlation with tumor pathological characteristics.3.The expression of CYTL1 affecte the treatment effect of LACC when patients received concurrent radiotherapy and chemotherapy.Part Ⅲ Study on Systemic Inflammation Index and Therapeutic Effect of Locally Advanced Cervical Squamous Carcinoma Before Concurrent Radiotherapy and ChemotherapyMethods1.Kaplan-Meier univariate survival analysis was used to analyze the value of clinicopathological features,systemic immune inflammatory response markers(NLR,PLR,LMR,SII),lymphocytes,neutrophils,and monocytes in predicting the outcome of concurrent radiotherapy and chemotherapy in patients with LACC.Meaningful variables of univariate analysis were included in COX proportional hazard model for multivariate analysis,so as to further clarify the independent prognostic factors affecting the prognosis of concurrent radiotherapy and chemotherapy in patients with LACSC.2.Based on the results of multivariate analysis,meaningful inflammatory variables were combined to construct a new prognostic scoring model.Results1.Univariate analysis showed age(P=0.024),degree of chemotherapy completion(P=0.095),lymph node metastasis(P=0.182),complete remission(CR,including imaging CR and pathological CR,P=0.000),disease control(Control,the disease state of imaging and pathological examination at the 6-month follow-up after treatment,P=0.000),PLR(P=0.006),NLR(P=0.089),LMR(P=0.021),SII(P=0.087),Hb(P=0.004)are all prognostic factors affecting OS.2.Multivariate regression analysis found that CYTL1(P=0.001),PLR(P=0.004),and TNM staging(P=0.002),were independent risk factors affecting OS in the case population of this study.3.Before concurrent radiotherapy and chemotherapy,CYTL1 strong expression(+and 2+)or PLR>123.36 was recorded as 1 point respectively,while CYTL1 weak expression(–or weak+)or PLR≤123.36 was recorded as 0 points respectively.Therefore,the study population is divided into 3 groups(a score of 0 was divided into low-risk group,1 was divided into middle-risk group and 2 was divided into high-risk group).The average survival time of the three groups was 57.8,44.8,34.5 months,respectively(P=0.013).Conclusions1.This study preliminarily proved that systemic inflammation index PLR and cervical cancer local factor CYTL1 were independent risk factors for OS in LACC patients who initially received radical concurrent radiotherapy and chemotherapy.2.The new inflammation-related prognostic scoring model based on PLR and CYTL1 was a simple,feasible,low-cost prognostic model that could be evaluated before radical concurrent radiotherapy and chemotherapy in locally advanced cervical squamous cell carcinoma.