Combined Detection Of Dickkopf-1 Subtype Classification Autoantibodies As Biomarkers For The Diagnosis And Prognosis Of Non-small Cell Lung Cancer

Background:Early diagnosis and screening of lung cancer is of great significance in preventing long-term progression and improving prognosis of lung cancer.LDCT(Low-dose Computed Tomography)is the primary method for clinical screening for high-risk lung cancer.But there are some problems cannot be ignored,such as false positive rate,excessive diagnosis,radiation damage and so on.The search for noninvasive methods of high sensitivity and specificity is a new trend in lung cancer screening and early diagnosis.Serum biomarkers such as tumor-associated antigens and autoantibodies have become a hotspot in the early diagnosis of lung cancer.In the early stage of the tumor,the immune system is able to recognize the abnormal expression of tumor antigens,activate the immune response,and produce a large number of autoantibodies.Autoantibodies have a long half-life in the blood,which can be detected in the early stage of the tumor,and may be used as an early diagnostic marker.Autoantibodies have the effect of immune surveillance in tumorigenesis and development.It is found that autoantibodies may be related to the prognosis of various types of tumors.However,the current research conclusion is contradictory and the mechanism is not clear.Therefore,the role of autoantibodies in early diagnosis and prognosis of tumors is worth further study.Objective: The present study aimed to utilize peptide array to identify epitopes of DKK1 autoantibodies and detect the antibodies that are linked to the epitopes resulting from DKK1.To explore the role of different subtypes of DKK1 autoantibodies in the diagnosis and prognosis of non-small cell lung cancer.Methods: Epitope mapping by peptide array-based serum screening of NSCLC patients and healthy controls was performed.Competitive ELISA was used to verify that DKK1 autoantibodies in serum could specifically bind to synthetic peptides and then detect the level of DKK1 autoantibodies in 209 lung cancer patients and 99 healthy controls by indirect peptide ELISA.To analyze the relationship between DKK1 autoantibody subtypes and clinicopathological characteristics,and the diagnostic value and prognostic value in non-small cell lung cancer.Results: 1.Four highly reactive epitopes were identified,including peptides 37-54(Pep A),67-84(Pep B),145-156(Pep C)and 247-261(Pep D).The DKK1 autoantibodies are further classified into subtypes according to response to different epitopes.2.The levels of autoantibodies were considerably higher in sera of lung cancer compared with the healthy controls(P<0.001).In M0 patients,DKK1 antibodies to Pep B,Pep C,Pep D were significantly higher than M1 patients(P=0.007,P=0.007,P=0.001),but no significant differences in DKK1 antibodies to Pep A in M0 and M1 patients(P>0.05)3.As indicated in the relationship between DKK1 autoantibodies subtypes and clinicopathological characteristics,DKK1 autoantibodies to Pep,Pep C,Pep D were correlated with NSCLC clinical staging(P=0.001;P=0.001;P=0.002)and distant metastasis(P=0.007;P=0.007;P=0.001).4.DKK1 autoantibodies as NSCLC diagnostic markers,AUC of DKK1 autoantibodies to the four peptides were 0.744(95%CI=0.684-0.799),0.809(95%CI=0.756-0.854),0.740(95%CI=0.684-0.791),0.767(95%CI=0.712-0.814)respectively.While there were no significant AUC changes between four autoantibodies subtypes.AUC of combined detection the four types of autoantibodies were 0.821(95%CI=0.764-0.868),the sensitivity and specificity were 58.1% and 85.3%.In the I and II phase NSCLC,AUC of DKK1 autoantibodies to the four peptides were 0.744(95%CI=0.684-0.799),0.809(95%CI=0.756-0.854),0.740(95%CI=0.684-0.791),0.767(95%CI=0.712-0.814)respectively.AUC of combined detection the four types of autoantibodies were 0.706(95%CI=0.611-0.790)、0.808(95%CI=0.732-0.871)、0.723(95%CI=0.640-0.796)、0.780(95%CI=0.704-0.843).AUC of ROC curves of DKK1 autoantibodies to the four peptides were 0.818(95%CI=0.719-0.894),the sensitivity and specificity were 76.9% and 75.9%.5.High levels of Pep B autoantibody was remarkably associated with better OS and PFS(P=0.004;P=0.006).However,there is no statistical significance in other autoantibodies subtypes.Subsequent Cox regression analysis disclosed that antibody to Pep B was an independent prognostic factor for NSCLC(OS: p=0.008,HR=0.435,95%CI:0.236-0.802;PFS: p=0.032,HR=0.533,95%CI:0.322-0.950).Conclusions: The expression of four types of DKK1 autoantibodies in NSCLC were up-regulated,which may be diagnostic markers of NSCLC.Pep B autoantibody was associated with distant metastases and clinical staging and positively correlated with overall survival and progression-free survival of NSCLC.What’s more,Pep B autoantibody was a good independent prognostic factor for NSCLC,suggesting that Pep B autoantibody may play an important role in the development of NSCLC.