Demographic Characteristics of Subjects
The study included 105 cases and 108 controls. The characteristics of all subjects are provided in Table 1. The gender and age distribution differed between the cases and controls (men, 60% and 47%, respectively; women, 40% and 53%, respectively). The mean age (± SD) of the cases and controls was 60.4 years (± 8.8) and 60.0 years (± 9.2), respectively. The demographics of the subjects’ lifestyle habits were evaluated. Drinking status, smoking and betel chewing were more prevalent among the cases than the controls. The distribution frequency of a family history of cancer between cases and controls statistically differed (P < 0.05). The results showed no difference between cases and controls in terms of body mass index (BMI) and marital status (P = 0.681 and P = 0.945, respectively).
Association Between Environmental Factors and EC Risk
The correlations of environmental exposure with EC risk are presented in Table S2. The primary outcomes of the multivariable analysis are depicted in this table, and after backward conditional selection analysis, our results showed that alcohol drinking, smoking and family history of cancer significantly increase the risk of EC (P < 0.05). BMI, marital status and betel chewing were not significantly associated with EC.
Association Between the Methylation Status of P16 and TP53 and Risk of EC
Table 2 provides the results of gene methylation status relative to EC, which revealed that P16 methylation had an increased EC risk and was statistically significant (ORadj = 5.24, 95% CI: 2.57–10.66). Furthermore, our study also found that TP53 methylation was significantly associated with EC at (P < 0.001).
Relationships Between P16 and TP53 Methylation and Environmental Factors
The relationship between P16 and TP53 methylation and environmental factors were explored in all 105 EC cases and 108 controls. As presented in Tables S3 and S4, drinking status and smoking were associated with increased risk of P16 methylation (ORadj = 2.49, 95% CI: 1.71–3.13, P =0.029; ORadj = 2.36, 95% CI: 1.36–4.75, P = 0.003, respectively). Moreover, the risk of TP53 methylation was slightly higher in subjects who were drinking alcohol and smoking (an increased OR), and the risk associated with these factors was statistically significant (ORadj = 2.06, 95% CI: 1.71–3.70, P =0.013; ORadj = 1.61, 95% CI: 1.15–3.21, P = 0.041, respectively).
The Effect of Interactions Between P16 and TP53 Methylation and Their Interactions with Environmental Factors on the Risk of EC
The results found that the combined effects between P16 methylation and alcohol drinking and smoking on EC risk existed (ORc = 2.43, 95% CI: 1.41–4.38, P =0.002; ORc = 2.75, 95% CI: 1.73–5.10, P =0.001 respectively), whereas no interaction between P16 methylation and environmental factors on the EC risk was showed, as seen in Table S5. As for the TP53 gene, its methylation and some environmental factors have combined effects on the risk of EC (P < 0.05) (Table S6). Additionally, the results in Table S7 illustrate that P16 methylation did interact with TP53 methylation on EC risk.
Characteristics of EC Patients
Of the 105 subjects who were recruited as EC cases in this study, all EC patients were included in this 10-year follow-up study. The association between demographic clinicopathological and EC prognosis was analysed, as presented in Table S8. Although the correlation between each demographic characteristic and prognosis of EC patients was not statistically significant, age, gender, BMI, family history of cancer, marital status and betel chewing were still used as the adjustment factors in analysing the relationship between clinical characteristics and EC prognosis, and these factors were common confounder factors in this study. Multivariate analysis based on Cox proportional hazard regression revealed that TNM stage, histology grading (poor differentiation) and lymph node metastasis were statistically significantly associated with EC prognosis (P < 0.05) (Table 3). The results from backward condition selected suggested that EC patients with TNM stage III had marginally poorer prognoses (HR = 1.25, 95% CI: 1.44- 3.42, P = 0.046) and EC patients with stage IV had seriously poorer prognoses (HR = 2.68, 95% CI: 1.04- 6.93, P = 0.041). In addition, metastasis was also associated with poorer prognosis of EC patients (HR = 1.52, 95% CI: 1.21–2.48, P = 0.013) (Table S9).
Association Between P16 and TP53 Methylation Status and EC Prognosis
In this study, the potential impact of P16 and TP53 methylation on EC prognosis was investigated. Our data are shown in Table 4. When compared with non-methylation, both P16 and TP53 methylation were strongly associated with EC prognosis (HR = 2.82, 95% CI: 1.13- 5.06; HR = 2.95, 95% CI: 1.34- 5.47, respectively). In Figure 1, our results show the association between P16 and TP53 methylation status and EC patients’ prognosis by the Kaplan-Meier method. Briefly, this study found that DNA methylation of P16 and TP53 was associated with patients’ overall survival. In addition, EC patients who had P16 and TP53 methylation showed significantly shorter overall survival than those who had non-methylation status (P = 0.027; P = 0.007, respectively).