The Study Between RUNX3 Polymorphisms And Gastric Cancer Susceptibility And Survival

Gastric cancer is one of the most prevalent malignant tumors worldwide. It is the fourth most common cancer and second leading cause of cancer death worldwide. Almost 42% of the cases occur in China. Gastric cancer is a disease with a complex, multifactor etiology, including interactions between genetic makeup and environmental factors. Despite some environmental agents such as Helicobacter pylori (Hp) infection, salt-preserved foods consumption, and tobacco smoking have been found to be the major risk factors for gastric cancer, only a fraction of individuals exposed to these factors develop gastric cancer during their lifetime, suggesting that genetic variation is an important risk determinant in gastric carcinogenesis.Single nucleotide polymorphisms (SNPs) are the most common forms of genetic variations of human genomic DNA and core message of human desease susceptibility. In the study of molecular epidemiology and genetics, SNPs have been widely used for etiology explore, therapy and prognosis evaluation. As an important transcription factor, RUNX3 participates in both TGF-βand Wnt signal parthways. RUNX3 is involved in neurogenesis of the dorsal root ganglia, T-cell differentiation and gastric epithelial cell oncogenesis. It functions as a tumor suppressor gene in various types of cancers.In the present study, we investigated the association between RUNX3 gene genetic variants and gastric cancer patients susceptibility and survival. The study will help us to elucidate the molecular mechanisms involving in gastric carcinogenesis as well as raise awareness of clinical prognosis. It is also beneficial for the individualized prevention, intervention and treatment.Part I The study between RUNX3 polymorphisms and gastric cancer susceptibilityAs a key member of the runt related transcription factors (RUNXs), RUNX3 is closely related with gastric carcinogenesis and cancer development. In this study, we used a hospital-based case-control study to validate the association between RUNX3 tSNPs and risk of gastric cancer.The study subjects consisted of 312 histologically confirmed gastric cancer patients and 329 age (±5 years) and sex matched cancer-free controls. The selected ten tSNPs were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The real-time RT-PCR was used to detect the mRNA expression of gastric tissues adjacent to tumor. The crude and adjusted odds ratios (ORs) and their 95% confidence interval (CIs) were obtained to assess the association between the RUNX3 polymorphisms and gastric cancer risk using unconditional univariate and multivariate logistic regression models. The Mann–Whitney U test was used to compare the RUNX3 mRNA expression levels within groups.The results indicated that the variant TC and CC genotypes of SNP3 rs11249206 were associated with a significantly increased risk of gastric cancer compared with the wild-type TT genotype (adjusted OR = 2.61, 95% CI = 1.79-3.80 for TC and 1.75, 1.03-2.99 for CC). For the SNP7 rs760805, compared with the TT, the variant AA genotypes was associated with a statistically significantly increased risk of gastric cancer (1.82, 1.14-2.92 for AA). Similarly, a significantly increased risk of gastric cancer was observed for the GA genotype (2.49, 1.61-3.84) and GG genotype (1.69, 1.05-2.72) in the SNP8 rs2236852, compared with the AA genotype. In the combined analysis of SNP3, SNP7 and SNP8, we found that individuals with 4-6 variant (risk) alleles had a significantly higher risk of gastric cancer (adjusted OR = 2.00, 95% CI = 1.41-2.85) than those with 1-3 risk alleles. Further stratification analysis showed that this increased risk was more pronounced among subgroups of age≥65 years (3.58, 2.00-6.38), never smokers (2.48, 1.58-3.89), and never drinkers (2.61, 1.70-3.99). The mRNA analyses showed that individuals with SNP8 GA and GG genotypes had higer expression than those with AA genotype (1.63±0.56 vs. 1.18±0.50, P = 0.011).In conclusion, RUNX3 gene polymorphisms may contribute to the risk of gastric cancer susceptibility in a Chinese population. Larger studies with more functional research are needed to confirm our findings.Part II The study between RUNX3 polymorphisms and gastric cancer prognosisMany published studies show that the RUNX3 expression is closely related with cancer prognosis. In this study, we aimed to validate the associaotion of RUNX3 genetic variants and gastric cancer survival. A total of 934 gastric cancer cases diagnosed with pathologic examination and undergone a surgical operation were enrolled as the study subjects. The tSNPs were selected similar with Part I. Immunohistochemical study was performed on the gastric tissues. Survival curves were plotted by Kaplan-Meier method and the differences of survival time were tested by the Log-rank test. Univariate or multivariate Cox regression models were fitted to estimate the hazard ratios (HRs) and 95% CIs for the associations and the effect of RUNX3 polymorphisms and the survival of gastric cancer. The Mann–Whitney U test was used to compare the RUNX3 protein expression levels within groups.In the current study, we observed that the tumor size (≥5.0 cm), the diffuse type, the depth of tumor invasion (≥T2), lymph node metastasis, distant metastasis and clinical stage (≥III) are poor prognosis factors (Log-rank P = 0.004 for distant metastasis, P < 0.001 for all the rests). Univariate Cox regression analysis revealed that patients carrying the RUNX3 SNP10 rs2282718 GG/AG genotypes had a significantly 27% increased death risk (adjusted HR = 1.27, 95% CI = 1.00-1.61) compared with those with the AA genotype, and this risk effect was more pronounced in female, no-self digestive disease, non-cardia carcinoma, diffuse type and clinical staging in stage IV. Finally, distant metastasis (Log-rank P < 0.001; HR = 1.61, 95% CI = 1.01-2.39), lymph node metastasis (Log-rank P = 0.009; HR = 1.67, 95% CI = 1.28-2.19), diffuse type (Log-rank P = 0.011; HR = 1.41, 95% CI = 1.08-1.84) and RUNX3 SNP10 AA (Log-rank P = 0.024; HR = 1.39, 95% CI = 1.04-1.85) were selected into the Cox stepwise regression model in the non-cardia carcinoma patients.In the intestinal type patients, individuals with the SNP1 rs6672420 AA genotype had a 1.04 fold increased death risk (adjusted HR = 2.04, 95% CI = 1.26-3.30) compared with those with the TT/AT genotypes, and individuals with the SNP2 rs11249208 GG genotype had a 91% increased death risk (adjusted HR = 1.91, 95% CI = 1.17-3.10) compared with those with the AA/AG genotypes. In the non-cardia patients, individuals with the SNP3 rs11249206 CT/CC genotypes had a 16% decreased death risk (adjusted HR = 0.84, 95% CI = 0.66-1.06) compared with those with the TT genotype; individuals with the SNP7 rs760805 AT/AA genotypes had a 24% decreased death risk (adjusted HR = 0.76, 95% CI = 0.59-0.96) compared with those with the TT genotype; for the SNP8 rs2236852, individuals with the CT/CC genotypes had a 19% decreased death risk (adjusted HR = 0.81, 95% CI = 0.63-1.05) compared with those with the TT genotype. In the combined analyses of SNP3,SNP7 and SNP8, we found that individuals with 3 protective genotypes had a significantly lower death risk (adjusted HR = 0.66, 95% CI = 0.51-0.86) than those with 0-2 protective genotypes, and this protecticve effect was more pronounced in younger patients (≤60 years), male, no-self digestive diseases, smaller tumor diameter (≤5.0 cm), no-distant metastasis and no-lymph node metastasis, depth of tumor invasion (T2), clinical staging in stage II and no-chemotherapy patients. In the stepwise Cox regression analyses of the non-cardia patients, we found the individuals with 3 protective genotypes were favorable prognosis factors (Log-rank P < 0.001, HR = 0.64, 95% CI = 0.50-0.83). The distant metastasis (Log-rank P = 0.026; HR = 1.57, 95% CI = 1.05-2.36), lymph node metastasis (Log-rank P < 0.001; HR = 1.73, 95% CI = 1.33-2.26), diffuse type (Log-rank P = 0.013; HR = 1.40, 95% CI = 1.08-1.83) were poor prognosis factors.Our results indicated that the RUNX3 tSNPs were associated with gastric cancer survival. Further studies with larger sample size are warranted and these tSNPs may be used as biomarkers for gastric cancer survival.