Association of polymorphisms in MALAT1 with the risk of endometrial cancer in Southern Chinese women

Abstract Background Endometrial cancer is the most common gynecologic malignancy worldwide. Polymorphisms in MALAT1 have been demonstrated to play critical roles in cancer. However, the roles of MALAT1 polymorphisms in the etiology of endometrial cancer have not been well documented. Methods We genotyped three MALAT1 polymorphisms in 249 endometrial cancer cases and 446 cancer‐free female controls using quantitative polymerase chain reaction with TaqMan probes. To estimate the association between MALAT1 polymorphisms (rs591291 C>T, rs664589 C>G, and rs4102217 G>C) and the risk of endometrial cancer, an unconditional logistic regression model was conducted to calculate the odds ratio (OR) and the 95% confidence interval (CI), adjusting for surgery history, menopause, number of deliveries, BMI, and FIGO stage. Results We found that the MALAT1 rs664589 C>G polymorphism was significantly associated with endometrial cancer risk (heterogeneous: adjusted OR = 0.57, 95% CI = 0.34‐0.93, P = .026; homogenous: adjusted OR = 3.74, 95% CI = 1.12‐12.45, P = .032; and recessive: adjusted OR = 4.06, 95% CI = 1.22‐13.48, P = .022). Stratified analysis further demonstrated that the MALAT1 rs664589 C>G polymorphism significantly increased the risk of endometrial cancer susceptibility in patients with no history of surgery, more deliveries, BMI between 25 and 29.9, and FIGO stages II‐III. Compared with the wild‐type GCG haplotype carriers, individuals with CGG haplotypes had a higher risk of developing endometrial cancer. Conclusion The MALAT1 rs664589 C>G polymorphism was associated with a significant increase in endometrial cancer risk.


| Patients and controls
In the present case-control study, 249 endometrial cancer patients were enrolled at The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University (WMU) from February 2007 to February 2017. The diagnosis of endometrial cancer was confirmed in all cases by histological examination of tissues from biopsy or resected specimens. Cancer-free controls (n = 446) were also recruited at the same hospital during routine physical examinations, and we excluded participants who had been diagnosed with malignant neoplasms or had a family history of cancers. This study was approved by The Second Affiliated Hospital and Yuying Children's Hospital of WMU, and written informed consent was obtained from all patients.
The TIAN quick FFPE DNA Kit (Qiagen Inc) was applied to extract genomic DNA from all patients from paraffin-embedded tissues, while genomic DNA from the controls was extracted from peripheral blood specimens using the TIANamp Blood DNA Kit (TianGen Biotech Co. Ltd.) as described previously. 16,17 The AUV absorption spectrophotometer was used to detect DNA purity and concentration (NanoDrop Technologies Inc).
Genotyping analysis was performed by real-time PCR with TaqMan PCR master mix and ABI Prism 7900HT genetic detection system. The details of the analysis procedures have been previously described. 18 In addition, approximately 5% of the samples were randomly selected as positive controls and negative controls for assessing the accuracy of the genotyping results.

| Statistical analysis
The goodness-of-fit chi-square test was adopted to evaluate departure from Hardy-Weinberg equilibrium (HWE) for the selected polymorphisms in control subjects. The heterogeneity of the genotypes and ages between patients and controls was evaluated by using a two-sided chi-square test. The association between MALAT1 polymorphisms and endometrial cancer risk was assessed by an unconditional logistic regression model, calculated as crude and adjusted odds ratio (OR) and 95% confidence interval (CI). Additionally, stratified analyses were performed by surgery history, menopause, number of deliveries, BMI, and FIGO stage. All statistical tests were carried out by SAS software (version 9.4; SAS Institute), with a twosided P-value < .05 considered statistically significant.

| Characteristics of the study participants
In the present study, we enrolled 249 endometrial cancer patients with an average age of 54.60 ± 9.09 months and 446 cancer-free controls with an average age of 53.03 ± 10.84 months. The frequency distributions of selected demographic characteristics of the cases and controls are shown in Table 1. There was no significant difference between cases and controls in age (P = .053), surgery history (P = .949), menopause (P = .075), number of deliveries (P = .110), and BMI (P = .063).

| Association between selected MALAT1 polymorphisms and endometrial cancer risk
We investigated the association with endometrial cancer of each of the three selected MALAT1 polymorphisms. First, the HWE test was used to identify genotyping errors in cancer-free controls. As shown in Table 2, all the genotype distributions were in accordance with HWE (P = .255 for rs591291, P = .853 for rs664589, P = .720 for rs4102217). Next, single-locus analysis indicated that MALAT1 rs664589 C>G was significantly associated with endometrial cancer risk (heterogeneous: adjusted OR = 0.57, 95% CI = 0.34-0.93, P = .026; homogenous: adjusted OR = 3.74, 95% CI = 1.12-12.45, P = .032; and recessive: adjusted OR = 4.06, 95% CI = 1.22-13.48, P = .022). However, other genetic associations with endometrial cancer risk were not discovered in the present study. When the three SNPs were combined, we found that carriers of 2-3 genotypes did not have a significantly increased risk of endometrial cancer at an OR of 1.25 (95% CI = 0.85-1.84, P = .263).

| Stratified analysis
We further explored the association of MALAT1 rs664589 C>G polymorphisms with endometrial cancer susceptibility by stratified analysis. The rs664589 CG/CC genotypes significantly increased endometrial cancer susceptibility in the women with no surgery history, more deliveries, BMI between 25.0 and 29.9, and FIGO stages II-III (Table 3).

| MALAT1 haplotypes and endometrial cancer risk
As shown in Table 4, MALAT1 haplotypes were available for analysis.

| D ISCUSS I ON
In recent years, increasing evidence has indicated that lncRNAs play

| CON CLUS ION
The present study provides evidence that MALAT1 rs664589 C>G significantly increased the risk of endometrial cancer. These findings need to be verified or linked with functional studies.  Abbreviations: CI, confidence interval; OR, odds ratio. a Adjusted for surgery history, menopause, delivery, and BMI.

ACK N OWLED G M ENTS
The results were in bold, if the 95% CI excluded 1 or P < .05.