AIM: To investigate the association between pre-miR-146a C/G polymorphism and gastric cancer risk.

METHODS: We performed a hospital-based, case-control study using polymerase chain reaction-restriction fragment length polymorphism method in 608 individuals (304 gastric cancer patients and 304 age and sex matched cancer-free controls).

RESULTS: The frequencies of pre-miR-146a C/G genotypes in the case group were significantly different from those in the control groups (P = 0.037). Compared with CC genotype carriers, subjects with the variant genotypes (GC + GG) had a 58% increased risk of gastric cancer (adjusted OR = 1.58, 95% CI: 1.11-2.20, P = 0.009). Moreover, a higher gastric cancer risk was especially evident in younger individuals aged ≤ 58 years, nonsmokers, and males (adjusted OR = 1.76, 95% CI: 1.08-2.87, P = 0.024; adjusted OR = 1.55, 95% CI: 1.06-2.28, P = 0.025; adjusted OR = 1.53, 95% CI: 1.04-2.27, P = 0.033; respectively).

CONCLUSION: Pre-miR-146a C/G polymorphism might be associated with an elevated risk of gastric cancer in Chinese population.

MicroRNAs (miRNAs) are endogenous 19- to 25-nt noncoding RNAs, which could negatively regulate gene expression through suppressing translation or degrading mRNA[1]. Bioinformatic data identified that a single miRNA could bind to a large number of mRNA targets, which could be implicated in disease development[2-8]. Interestingly, recent evidences indicated that miR-146a can play a role of mediator in a wide spectrum of biological processes, such as cell proliferation, differentiation, apoptosis, immune response and tumorigenesis[9-15]. Aberrant expression of miR-146a has been reported as a signature in breast cancer, pancreatic cancer, prostate cancer, and cervical cancer [16-18].

Single nucleotide polymorphism (SNP) is the most common type of genetic variation in the human genome. Polymorphisms in human pre-miRNA genome lesion modify the processing and/or target selection of human miRNAs, which are implicated in cell cycle regulation, and thereby play critical roles in carcinogenesis[19,20]. Calin et al[21] found that the pre-miR-206 precursor mutation was associated with the risk of chronic lymphocytic leukemias (CLL). They also found a germ-line mutation in the miR-16-1-miR-15a primary precursor, which caused low levels of miRNA expression in vivo and in vitro[21].

Pre-miR-146a C/G polymorphism designated rs2910164 is located on chromosome 5, in the stem region opposite to the mature miR-146a sequence[22]. Several epidemiological studies have examined the role of the pre-miR-146a polymorphism in many human cancers, such as hepatocellular carcinoma (HCC), prostate cancer, breast cancer and papillary thyroid carcinoma (PTC)[22-26]. However, the results of these studies were inconsistent. Furthermore, no data are available concerning the association between pre-miR-146a C/G polymorphism and the risk of gastric cancer. Therefore, we have conducted a hospital-based case-control study to investigate the potential link between this polymorphism and gastric cancer in Chinese population.

Recent genome-wide SNP polymorphisms are encouraging and widely performed to help in developing more accurate diagnostic and therapeutic strategies of various kinds of human disorders[29]. In the present study, we for the first time found that the C to G variant in pre-miR-146a conferred an increased risk of gastric cancer in Chinese population. We also found that the elevated gastric cancer risk was especially evident in the individuals aged ≤ 58 years, nonsmokers and males.

The human miR-146a is located on chromosome 5[22]. In general, one miRNA targets thousands of mRNAs and one mRNA is regulated by many miRNAs. Therefore, it is very hard to understand the precise mechanisms of the miRNA in the pathogenesis of human disorders. Nevertheless, various studies have suggested that miR-146a is a NF-κB-dependent gene[30,31]. Upon processing, it could down-regulate levels of IRAK1 and TRAF6 proteins, reducing the activity of the NF-κB signaling pathway, which has been implicated as an important causal link between inflammation and carcinogenesis[9,10,32-34]. To date, aberrant miR-146a expressions have been described in several diseases[16-18,35-38]. Recent reports have indicated that miR-146a levels were increased in the tissues associated with chronic inflammatory diseases such as rheumatoid arthritis and psoriasis, whereas there was no increase of miR-146a levels in tissues obtained from patients with other chronic inflammatory diseases such as the skin from atopic eczema or lung biopsies from mild asthmatics[35-37]. Moreover, elevated miR-146a levels have been reported in PTC, cervical cancer, breast cancer and pancreatic cancer, whereas decreased miR-146a expression is associated with prostate cancer[16-18,38].

A C/G polymorphism (rs2910164) resides at position + 60 relative to the first nucleotide of pre-miR-146a, placing it in the passenger strand[22]. Compared with the major C allele, the minor G allele causes mispairing in the hairpin and a higher dG from -40.3 to -43.1 kcal/mol, suggesting a more stable secondary structure for the G allele[39]. The genetic variant might have an impact on processing the pre-miRNA into the mature microRNA. The mutation in pre-miR-146a gene has been reported to modulate cancer risks, although the results are inconsistent[22-26]. Xu et al[23] showed that male individuals with GG genotype were 2-fold more susceptible to HCC (OR = 2.016, 95% CI: 1.056-3.848, P = 0.034) compared to those with CC genotype. In a case-control study, Xu et al[24] suggested that CC homozygotes exhibited an association with prostate cancer with a 65% decreased risk (95% CI: 0.43-0.99, P = 0.03). Findings by Jazdzewski et al[22] demonstrated that the GC heterozygous state was associated with an increased risk of acquiring PTC (OR = 1.62, P = 0.000007), and both homozygous states were protective with OR = 0.42 for the CC genotype and OR = 0.69 for the GG genotype. Although one study showed that this polymorphism did not appear to be linked with an increased risk of breast cancer[26], another indicated that those breast and ovarian cancer patients who had at least one C allele were diagnosed at an earlier age (median age: 45 vs 56, P = 0.029 for breast cancer; median age: 45 vs 50, P = 0.014 for ovarian cancer)[25]. We noted that, compared with the CC genotype, the G allele carriers had a 55% increased risk of gastric cancer. After adjustment for age, sex, smoking status, residence, hypertension and diabetes, the difference still existed, confirming that the polymorphism was related to the risk of gastric cancer in China. Although the precise mechanisms of miRNA expression regulation are largely unknown and the inconsistent effect of miRNA SNP for cancer diagnosis and prognosis is ambiguous, those results may imply that miRNA expression patterns are associated with the biological and clinical behavior of human cancers.

Our data showed that the polymorphism was associated with the increased risk of gastric cancer among subgroups of the subjects aged ≤ 58 years, whereas not among older subjects. Overwhelming accumulated exposure to environmental carcinogens and weak immune system in older individuals may account for the age difference we observed[28]. An old age is related to an increased risk for gastric cancer, which is more likely due to the age effect rather than the direct genetic effect. Therefore, the polymorphism in the pre-miR-146a may be more evident in early-onset gastric cancer, although this age-specific association needs to be further replicated.

Tobacco smoking is a known cause of gastric cancer[40]. The association between the polymorphism and gastric cancer risk may be masked by the relatively high-level exposure to tobacco carcinogens in smokers so that it is more influential in nonsmokers.

Subsequently, a positive correlation between pre-miR-146a genotype and sex was found. A significantly elevated risk of gastric cancer was observed for the GG + GC genotypes among males but not among females. Xu et al[23] also concluded that male individuals with GG genotype were more susceptible to HCC compared to those with CC genotype. Based on the findings, it would therefore be plausible to expect that the sex-specific effect of pre-miR-146a may be responsible for the different results between males and females, although the exact mechanism remains unclear.

Further stratified analyses by clinicopathologic parameters of gastric cancer showed no significant association between the pre-miR-146a genotypes and clinical pathological parameters of gastric cancer. Gastric cancer is considered to be a complex, multi-step and multi-factorial process. Although our data did not support a causal link between the variant genotypes and gastric cancer progression, we could not exclude the possibility that other factors might impact the interaction between them. Moreover, the size of cases in the subgroups was relatively small; our findings from the stratified analyses should be interpreted with caution before confirmed in further studies.

Several limitations should be addressed in the study. First, lack of information on Helicobacter pylori (H. pylori) infection status did not allow us to analyze the interaction between the polymorphism and H. pylori infection status, because it was unethical to do H. pylori test in every subject. Second, the sample size was relatively small. This may have been underpowered to detect small but real gene-environment associations. Finally, the study was conducted in Chinese population, data should be extrapolated to other ethnic groups cautiously. Nevertheless, our initial data presented valuable insights and interesting information to guide future studies in this area.

In summary, our results suggested that pre-miR-146a C/G polymorphism is associated with the increased risk of gastric cancer in Chinese population, especially in younger individuals, nonsmokers, and males. Further studies are required to elucidate the mechanism underlying the polymorphism and gastric cancer progression.