Abstract
Aims
The aim of this study was to investigate the impact of three single nucleotide polymorphisms (SNPs) within X-Ray Repair Cross Complementary Group 2 (XRCC2) gene and additional gene- abdominal obesity (AO) interaction with endometrial carcinoma (EC) risk.
Methods
Hardy–Weinberg equilibrium was tested for all participants by using SNPstats (online software: http://bioinfo.iconcologia.net/SNPstats). The best SNP–SNP and gene–AO interaction combination among three SNPs within XRCC2 gene and AO was screened using generalized multifactor dimensionality reduction (GMDR).
Results
We employed the logistic regression analysis showed that rs718282-T allele is associated with increased EC risk, adjusted ORs (95%CI) were 1.67 (1.23–2.04). However, we did not find statistical association between rs3218536, and rs3218384 and EC susceptibility. GMDR analysis was used for SNP-SNP- and gene-abdominal obesity analysis. The cross-validation consistency and the testing accuracy for the interaction were calculated. The two-locus model between rs718282 and AO had a testing accuracy of 60.11%, which was significant at the p < .001 level, and this two- locus model was considered as the best model. It provided statistical evidence for rs718282 gene–AO interaction effects. The results indicated that AO influenced the EC risk depending on the rs718282 genotypes. Compared with non- AO subjects with rs718282–CC genotype, AO subjects with rs718282-CT or TT genotype had the highest EC risk, OR (95%CI) was 2.83 (1.67 − 4.02), after covariates adjustment.
Conclusions
Both the rs718282- T allele, and its interaction with AO were associated with increased EC risk.
Introduction
Endometrial carcinoma (EC) is a group of epithelial malignant tumors that occur in the endometrium epithelium and is one of the common malignant tumors in the reproductive system of female [Citation1,Citation2], and accounting for 4.8% of all malignant tumors in women [Citation3]. With mass cancer screening since 2008, the incidence of endometrial cancer in China has surpassed that of cervical cancer, ranking first among gynecological cancers in developed cities [Citation4]. Nearly, 300,000 women worldwide are diagnosed with endometrial cancer each year. Previous study has implied that genetic factors play a role in the development of EC [Citation5]. But the detailed mechanism for this role was not well known. Study also reported that mutations in DNA of double-strand breaks (DSB) repair genes are involved in the pathogenesis of cancers [Citation6]. DSB in DNA may be rectified either by homologous recombination (HR) or non-homologous end joining [Citation7]. X-ray repair cross complementing gene 2 (XRCC2), as a homolog of RAD51, can play an important role in homology recombination (HR) processes [Citation8].
X-Ray Repair Cross Complementary Group 2 (XRCC2) gene is located at 7q36.1, which is one of the candidate genes for tumor function [Citation9,Citation10]. Many previous studies [Citation11–13] have reported that XRCC2 single nucleotide polymorphisms (SNPs) are significantly associated with multiple malignant tumors, including colorectal cancer, ovarian and cervical cancer. Up to now, the development and pathogenesis of EC are not completely clear. And less studies were conducted to the relationship between XRCC2 SNPs and EC risk. The pathogenesis and development mechanism of EC is a complex and multi-stage process. The influencing factors include environmental factors, genetic factors, and their synergistic effects [Citation14,Citation15]. Previous studies have reported that obesity is an important risk factor for endometrial cancer [Citation16,Citation17]. There are too many fat cells in obese people, and the excess fat cells can convert peripheral androgens into estrogen. Therefore, the level of non-antagonistic estrogen acting on the endometrium is high, leading to endometrial hyperplasia or cancer. But to our knowledge, there are few studies on the effect of XRCC2 gene and its synergistic effect with obesity on EC risk. Therefore, this study aims to verify the effect of three SNPs in XRCC2 gene on EC susceptibility, and whether there is interaction between XRCC2 gene and obesity, which has an impact on EC risk.
Materials and methods
Subjects
The study sample was consisted of 906 participants, including 450 patients who were hospitalized in our hospital with EC and 456 normal controls who participated in our hospital for physical examination with health certificate. All patients were clinical diagnosed as endometrial cancer by pathology. Those participants with radiotherapy, chemotherapy history and any other cancer history were excluded from the case group. All the control groups were healthy people without any type of cancer, and matched with the case group according to age (± 3 years old). All participants are from the Han population and any blood and genetic relationship between any two participants. At the time of recruitment, all participants signed the written informed consent form.
Genomic DNA extraction and genotyping
We collected 3 ml of whole blood samples from all participants. According to the manufacturer’s instructions, the genomic DNA of all participants was extracted from the whole blood treated by EDTA using the DNA Blood Mini Kit (Qiagen, Hilden, Germany) and stored in a refrigerator at − 20° C for use. Three selected SNPs were genotyped using restriction fragment length polymorphism based on polymerase chain reaction (RFLP- PCR). The nucleotide sequence of the primer and the detailed description of the four SNPs are shown in . The conditions for genotyping progression were as follows: one initial denaturation step at 95 °C 45 s, 72 °C for 45 s and 72 °C for 60 s, repeated in 35 cycles.
Table 1. Description and primer sequences designed for sequencing 3 SNPs of XRCC2 gene.
Statistical analysis
In this study, SPSS 22.0 was used for statistical analysis. The percentages were calculated for categorical variables and were compared using Chi-squared methods between case and control group, and the means and standard deviations (SDs) were calculated for normally distributed continuous variables and were compared using t-test between case and control group. Logistic regression was employed to investigate the relationship between XRCC2 gene polymorphism and EC susceptibility. The best SNP- SNP and gene- AO interaction combination among three SNPs within XRCC2 gene and AO was screened using generalized multifactor dimensionality reduction (GMDR) [Citation18]. All reported p-values were two-tailed, and those less than 0.05 were considered statistically significant.
Results
General characteristics of study participants with EC and normal controls was shown in . A total of 906 participants were selected, including 450 patients with EC and 456 normal controls. The means of age for all participants is 65.8 ± 11.8 years. There was no significant difference between EC cases and normal controls was observed for the parameters including age, hypertension, T2DM, the rates for smoking and alcohol drinking (all P values were more than .05). In addition, the mean WC values, rates of contraceptive use history and were higher in cases than controls.
Table 2. General characteristics of study participants with GC and normal controls.
In current study, all genotypes of three SNPs are distributed according to Hardy–Weinberg equilibrium. The frequency for the rs718282- T allele is 29.9% in cases and 19.7% in controls, which were significantly different between EC patients and normal controls. Logistic regression testing suggested that rs718282-T allele is associated with increased EC risk, adjusted ORs (95%CI) were 1.67 (1.23–2.04). However, we did not find statistical association between rs3218536, and rs3218384 and EC susceptibility ().
Table 3. Association analysis for three SNPs within XRCC2 gene and EC risk.
shows the results of GMDR analysis for SNP-SNP and gene- AO interaction. The cross-validation consistency and the testing accuracy for the interaction were calculated. The two-locus model between rs718282 and AO had a testing accuracy of 60.11% (p = .001), and a cross-validation consistency of 9/10. Compared with non-AO subjects with rs718282-CC genotype, AO subjects with rs718282-CT or TT genotype had the highest EC risk, OR (95%CI) was 2.83 (1.67–4.02), after covariate adjustment ().
Figure 1. Logistic regression for interaction analysis between rs718282 and AO on EC risk.
Table 4. GMDR analysis for SNP-SNP and gene- AO interaction on EC risk.
Discussion
In this study, we found that rs718282-T allele is associated with increased EC risk. However, we did not find statistical association between rs3218536 or rs3218384 and EC susceptibility. Previously, studies have indicated that XRCC2 gene was associated with several cancers, including colorectal cancer, ovarian and cervical cancer [Citation11–13]. But less study focused on the influence of the XRCC2 gene SNPs on EC risk. In terms of rs718282, some reports suggested that XRCC2-rs718282 is statistically related to the susceptibility of esophageal squamous cell carcinoma and cardiac cancer [Citation19,Citation20]. Michalska et al. [Citation20] firstly performed a case- control study in Polish women, and they suggested that the SNP rs718282 (-41657 C/T) of the XRCC2 gene was significantly associated with increased EC risk in Polish women, which supported the hypothesis that rs718282 polymorphism may be correlated with high incidence of EC. However, this study has some limitations, such as the participants enrolled for this study were all females; therefore, the results of this study are farfetched to represent the male population or the whole population. Secondly, participants enrolled in this study were all Polish women, to date, no study regarding this topic was performed in Chinese population. Lastly, the sample size of this study was relatively small. In our study, we obtained similar results with the fore-mentioned study, and the sample size in our study was larger. Previously, the relationship between rs3218536 (Arg188His) and EC risk have been reported in different populations [Citation21–23], and these studies were all suggested that no significant association between rs3218536 (Arg188His) and EC risk were observed. And in our study, we also found that consistent results.
Generally, the pathogenesis and development of endometrial carcinoma is a complex process, in which many factors are involved [Citation24], many genes, environmental factor and the synergistic effect between genetic and non- genetic factors. Previously, some studies suggested that obesity was an important risk factor for EC incidence [Citation25], especially, abdominal obesity may be the best obesity-related index affecting the risk of EC [Citation26]. In our study, the WC values was significant different between EC patients and normal controls, and the WC values was higher than that in controls. However, in recent years, few studies have reported the effect of the interaction between AO and XRCC2 genes on EC susceptibility. As we know, we firstly investigated the effect of interaction between XRCC2 gene and AO on EC susceptibility. In this study, we performed interaction test using GMDR model, we found that the two-locus model between rs718282 and AO was considered as the best model, which indicating that the interaction between rs718282 and AO have significant influence on EC susceptibility. It provided statistical evidence for rs718282 gene–AO interaction effects on EC risk, which means that AO influenced the EC risk depending on the rs718282 genotypes.
The limitation in this study should also been discussed. Firstly, although it has met the requirement the sample size in our study was relatively small, and the results obtained from our study should be verified in different population with larger sample size. Secondly, the subjects included in this study are all Han people, so the results of this study still need to be summarized in other ethnic groups in China. Thirdly, hormone levels are also important aspect of endometrial carcinoma, but in this study, hormone was not tested for all participants, so we could not investigate the association between SNPs and hormone level. Lastly, genetic predisposition is increasingly recognized as an important factor in EC risk, but we could not exclude the impact of genetic predisposition on results.
In conclusion, we found that the rs718282- T allele, interaction between rs718282 and AO were both associated with increased EC risk.
Authors’ contributions
All authors have read and approved the manuscript.
Wenjuan Tian: definition of intellectual content, literature research, clinical studies
Siyu Cao: experimental studies, data acquisition, data analysis.
Wei Zhang: experimental studies and manuscript preparation
Chenlian Quan: statistical analysis.
Meiqin Zhang: guarantor of integrity of the entire study, study concepts and manuscript editing.
Yan Huang: guarantor of integrity of the entire study, study concepts, study design and manuscript review.
Ethics approval and consent to participate
Each participant understood the process of the study and signed a written informed consent before the start of the study. All study protocols of the current study were approved by ethics committee of Shanghai Medical College of Fudan University. All methods were performed in accordance with the Declaration of Helsinki.
Consent for publication
Not applicable.
Availability of data and material
The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request.
List of abbreviations
| OR | = | odds ratios |
| CI | = | confidence interval |
| XRCC2 | = | X-Ray Repair Cross Complementary Group 2 |
| EC | = | endometrial carcinoma |
| GMDR | = | Generalized multifactor dimensionality reduction |
| SNPs | = | single nucleotide polymorphisms |
| AO | = | abdominal obesity |
| HWE | = | Hardy-Weinberg equilibrium |
| WC | = | waist circumference. |
Acknowledgements
We appreciate the cooperation of the families and individuals who cooperated in this study.
Disclosure statement
No potential conflict of interest was reported by the author(s).
Additional information
Funding
References
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