In our study, TNF -α-857 C/T genetic variant containing the T allele was associated with a significantly increased risk for gastric cancer. SNP-SNP interaction in the TNF gene including TNF -α-857 C/T or TNF -α-1031 T/C genetic variant was associated with a risk for gastric cancer among smokers but not among non-smokers. Moreover this effect of the TNF -α-857 C/T genetic variant was ascertained in both haplotype and MDR analysis.
Previous studies reported that TNF gene polymorphisms modified the risk of gastric cancer [5, 8–11], while others did not find a significant association [6, 7, 12–15]. This inconsistency may be due to the small number of cases, differences in ethnic populations and SNP selection. Two recent meta-analyses assessed the association between TNF -α polymorphisms and risk of gastric cancer [33, 34]. According to these papers, TNF -α-308 polymorphism had a significantly increased risk for gastric cancer but TNF -α-857 polymorphism only showed a marginally significant risk due to the small number of studies. In contrast to the meta-analysis, our results indicated that TNF -α-857 polymorphism was associated with a significantly increased risk for gastric cancer but other SNPs including TNF -α-308 did not show a significant association. Our findings about the single SNP effect of TNF -α-857 polymorphism can help clarify results of the previous meta-analyses. Inconsistency in TNF -α-308 polymorphism may be due to an ethnic difference since TNF -α-308 AA polymorphism is rare in the East Asian population. Also, we were not able to clearly evaluate TNF -α-308 AA polymorphism and gastric cancer because none of our gastric cancer cases had the TNF -α-308 AA polymorphism.
In haplotype-pairs and MDR analysis, we observed a genetic combined effect among TNF SNPs. Individual haplotype-pairs including TNF -α-1031, -863 and -857 were consistently associated with a significantly increased risk of gastric cancer. Compared to the single SNP effect of the TNF -α-857 polymorphism, a greater odds ratio for haplotype-pairs including TNF -α-857 indicated that a synergistic interaction among TNF SNPs was more strongly associated with gastric cancer development. These results were nearly replicated in the MDR analysis. Our consistent findings from the different statistical methodologies are quite meaningful.
TNF cytokine may interact with cigarette smoking to promote gastric cancer development. Biologically, cigarette smoking can activate systemic inflammations  and augment the level of TNF -α through changes in inflammatory markers or cytokine level in animal models [36–38]. Although these findings have not been fully reproduced in humans, several papers suggest an indirect connection between cigarette smoking and TNF gene in the etiology of gastric cancer [39–42]. Some studies reported that circulating concentrations of TNF -α were increased and higher TNF -α level was associated with such diseases in smokers compared to non-smokers [39–42]. In one of these studies, higher TNF -α levels were presented among smokers, especially subjects with the 857 T allele and rare haplotype of the TNF -α promoter . Similar to our findings, the study concluded that TNF -α-857 polymorphism was especially susceptible to the hazards of smoking and the TNF cytokine was strongly affected by cigarette smoking.
The combined effect between smoking and TNF gene was definitely expressed in our haplotype-pairs and MDR analysis. Haplotype-pairs formed with TNF -α-1031, -863 and -857 were associated with a significantly greater increased risk for gastric cancer only among smokers but not among non-smokers. Both single SNP and haplotype-pair effect of TNF -α-857 T allele were related with an increased risk among smokers. Haplotype-pairs including TNF -α-1031 and -863 genetic variation that did not show significance for single SNP effect were also associated with a significantly increased risk of gastric cancer among smokers only. In contrast to TNF -α-857, TNF -α-1031 and -863 may interact mutually based on haplotype specificity and not allele specificity, and this effect may be strongly affected by smoking. If only allele specificity by high-producing or mutant alleles plays a role as risk factors for gastric cancer, haplotype-pairs composed of a greater number of high-producing or mutant alleles may have an even greater risk. Based on this hypothesis, CAT haplotype should be the most powerful risk factor for gastric cancer development and CAC haplotype composed of two high-producing alleles should show a greater risk than CCC or TCT haplotype that includes only one high-producing allele. However, we were not able to fully test this hypothesis because only four haplotypes, TCC, TCT, CCC and CAC, were observed in our population. Considering our results, independent allele specificity of TNF -α-857 T and mutual haplotype specificity of TNF -α-1031 and -863 may be more important risk factors rather than the total number of high-producing alleles, especially for smokers. Moreover, these results were reproduced in all haploblocks regardless of different SNP combinations and were nearly replicated in the MDR analysis. This supports our conclusion that the interaction between the TNF gene and smoking may play a crucial role in the etiology of gastric cancer.
Although, to our knowledge, this is the first study to report an interaction between the TNF gene and cigarette smoking on gastric cancer development, our study had several limitations. First, because of a small number of gastric cancer cases and small sample size, we did not have sufficient statistical power and were not able to stratify on different factors. Second, although we had information on cardiac or non-cardiac cancer of gastric cancer cases, because of our small number of cases, we were not able to observe a difference for gastric cancer risk according to cancer type. Moreover, we did not collect information on cancer histology. Finally, a part of SNPs which is related to signal pathway (NF-κB) from H. pylori to cytokine gene expression were genotyped. Other cytokines, such as interleukin s and GM-CSF, which are involved in NF-κB pathway, were not considered for analysis so we examined only a small portion of signal pathway for gastric carcinogenesis.
In spite of these limitations, our study had several strengths. This is a population-based, nested case-control study that is free of biases that are common in retrospective studies. Additionally, we matched cases and controls according to basic confounders, such as age and sex, and significant confounding factors selected in the full model. Furthermore, we used various approaches to detect the potential association between genetic and environmental factors on gastric cancer, and derived consistent results through different approaches. Finally, the minor allele frequencies (MAFs) of all cytokine genes analyzed in our study showed very similar results in the Korean, Chinese, and Japanese Hapmap projects [43, 44] and thus, our results are applicable to most East-Asian populations. On the basis of this study results, we will be able to make more conclusive evidence in future studies.