1. Introduction
Colorectal cancer (CRC) is highly prevalent globally, ranking as the second most common cancer among women and the third most common among men [
1,
2,
3]. The incidence and mortality rates of CRC vary significantly among countries, with differences of up to tenfold [
2,
3,
4]. Several factors contribute to this variation, including meat consumption, cigarette smoking, and exposure to carcinogens, which account for approximately 85% of CRC cases [
5,
6].
In Taiwan, CRC is a significant health concern, with the highest incidence rate among all types of cancer and ranking third in terms of mortality, following lung cancer and hepatoma. The presence of a familial cancer history in approximately 15–20% of CRC cases [
7,
8], indicates the potential contribution of genetic factors to the development of CRC. In recent years, a plethora of genetic biomarkers associated with CRC have been identified [
9,
10,
11,
12,
13,
14], and there is still considerable interest in identifying additional genetic susceptibility factors and investigating the interactions between genetic factors and other risk factors. Gaining a better understanding of the genetic contributions to CRC can assist scientists in developing more targeted and precise approaches to cancer prevention and therapy.
MicroRNAs (miRNAs) are single-stranded, non-coding RNAs that function as negative regulators of the expression level of genes [
15]. They play pivotal roles in diverse biological processes, encompassing embryonic development, cell proliferation, apoptosis, tissue remodeling, and notably, the process of carcinogenesis [
16,
17]. Genetic variations have been identified in both miRNA genes and their target genes, and these variations have been associated with a wide range of human diseases. Meanwhile, there is mounting evidence supporting the idea that dysregulated miRNAs play critical roles in tumorigenesis [
18,
19]. In CRC, several miRNAs have been identified to modulate cell proliferation, migration, apoptosis, and response to radiation in both CRC cells and patient samples. These miRNAs include miRNA 133A [
20], miRNA 483-3p [
21], miRNA 652 [
22], and miRNA-627-5p [
23]. Considering their dysregulation and significant correlation with numerous types of cancer, miRNAs emerge as promising targets for novel therapeutic strategies. Moreover, their secretion into extracellular fluids positions them as promising biomarkers for accessing tumor initiation, progression, metastasis, and tumor survival. However, the genetic contribution of miRNAs to the development of cancer has not yet been thoroughly investigated.
Single nucleotide polymorphisms (SNPs) are subtle genetic variations that commonly occur within miRNA genes. These genetic variations can potentially affect the expression and function of specific miRNAs, thereby contributing to tumorigenesis [
24]. In the recent decade, accumulated studies have examined the relationship between numerous SNPs in miRNAs and the risk of CRC [
25]. Among these SNPs,
mir146a rs2910164 is the most extensively studied. The C allele of this SNP reduces the nuclear processing efficiency of pri-mir146a, resulting in a less stable structure and decreased mir146a expression [
26]. In 2020, Santos and colleagues reported that mir146a expression within the tumor tissues was significantly higher among the CRC patients carrying the
mir146a rs2910164 GG genotype compared to those carrying the GC or CC genotypes [
27]. Regarding the association between this SNP and CRC risk, while the majority of the literature indicates no association [
27,
28,
29,
30,
31,
32,
33,
34], there are specific studies that have found associations within certain populations [
35,
36,
37]. The CC genotype has been associated with increased CRC risks in populations from Greece [
38], Korea [
39], and China [
35]. However, contrasting associations have been reported in Lithuania [
40] and China [
41,
42]. Another frequently studied miRNA SNP is mir196a-2 rs11614913. Zhan and colleagues reported that this SNP influences RNA maturation and its interaction with other molecules [
43]. Several studies have investigated its association with CRC risk, and the results have been inconsistent [
28,
29,
30,
31,
33,
37,
38,
40,
42,
43,
44,
45,
46,
47,
48].
In this study, we aim to investigate the associations of the
mir146a and
mir196a-2 SNPs with the risk of CRC among a Taiwanese population. The physical locations of these two SNPs are illustrated in
Figure 1. Additionally, we aim to assess the potential interactions between genotypes and factors such as age, gender, smoking, and alcohol consumption in relation to CRC risk. Furthermore, we explored the correlation between genotypes and miRNA expression. Importantly, we conducted a thorough review of the existing literature and presented a concise discussion of its findings.
3. Discussion
In this study, we provided evidence that the
mir196a-2 rs11614913 CC genotype and C allele may contribute to a higher risk of CRC (
Table 2 and
Table 4). The
mir146a rs2910164 SNP was not associated with CRC risk in Taiwan (
Table 3). Our findings are consistent with a previous study by Zhan, which reported an association between the
mir196a-2 rs11614913 C allele and increased CRC risk in China, [
43]. However, two studies have reported conflicting results, suggesting that individuals carrying the
mir196a-2 rs11614913 TT genotypic pattern are at higher risk of developing CRC than those carrying the CC genotypic pattern in Iran and China [
42,
45]. The inconsistency between the findings of Zhan and Lv, despite studying similar populations, cannot be attributed to ethnic heterogeneity but rather may be influenced by sampling bias or other factors. It is worth noting that the CC genotype frequency in Lv’s study is unusually low. For Europeans, it appears that
mir196a-2 rs11614913 is not associated with CRC susceptibility [
28,
29,
38,
40]. For Japanese, the genotype of
mir196a-2 rs11614913 is not associated with CRC susceptibility either [
31]. Thus far, the studies that have reported a positive association between
mir196a-2 rs11614913 and CRC risk have focused on Asian populations ([
43] and the current study). The conflicting results observed in Iranian [
45,
48] and Chinese [
42,
43,
47] populations may be resolved through further investigations involving larger sample sizes. It should be emphasized that the frequency of the T allele of mir196a-2 rs11614913 varies significantly across different ethnicities, ranging from 18.8% in Africans, 39.4% in Europeans, 41.1% in Mexicans, 30.7% in South Asians, and 54.8% in East Asians (
Supplemental Table S2). The T allele frequency in our controls (56.4%) is similar to that in East Asians (54.8%). The T allele represents the major allele in East Asians but is the minor allele in all other ethnicities. Conducting additional studies on different populations could provide insights and help reconcile the discrepancies. All the literature investigating the associations between
mir196a-2 rs11614913 genotypes and CRC risk was summarized in
Table 6, including the current study.
Quantification of mir196a-2 expression levels in serum can offer functional evidence that supports the role of mir196a-2 in the etiology of CRC. However, previous studies often lack the necessary data in this regard. Circulating miRNAs are believed to be encapsulated in exosomes, which protect them from degradation by RNase enzymes. Therefore, the expression of mir196a-2 could potentially serve as a measurable biomarker for CRC. In our study, we observed significantly higher expression levels of mir196a-2 carrying the homozygous variant genotype (CC) of
mir196a-2 rs11614913 compared to those carrying the wild-type TT genotype. This finding provides biological plausibility for the association of CC genotypes with increased CRC risk. The observed difference in RNA expression levels aligns with the genotypic data, supporting that the CC genotype has a significant effect on CRC risk. These findings are consistent with a previous report by Zhan, which reported higher expression levels of mir196a-2 in tumor tissue of patients carrying the CT and CC genotypes compared to those with the TT genotype [
43].
The genotypes of
mir196a-2 rs11614913 may have a significant impact on various unresolved aspects of CRC. For example, several studies have examined the potential prognostic value of rs11614913 in Asian CRC patients. In 2018, Pao and colleagues showed that CRC patients carrying the rs11614913 CC genotype had a shorter overall survival time among 188 Taiwanese CRC cases [
49]. Similarly, in 2011, Jang and colleagues reported that the heterozygous TC genotype, not the homozygous one, may serve as a risk factor for unfavorable overall survival in 446 Korean CRC patients [
50]. Further investigations are necessary to validate these findings, as they were conducted exclusively in Asian populations. However, these findings hold potential clinical significance and emphasize the importance of exploring the prognostic value of mir196a-2 rs11614913 in other populations as well.
This study has several limitations that should be acknowledged. Firstly, the absence of CRC tissues prevented us from comparing mir196a-2 expression levels between CRC tissues and adjacent normal tissues. However, the measurements of serum mir196a-2 expression in controls provided an important genotype–phenotype (gene expression) correlation, supporting the observed association between genotypes and CRC risk. Secondly, the CRC patients included in our study were heterogeneous in terms of clinical features and treatments, and some patients were lost to follow-up, which hindered the analyses on the association between the mir196a-2 SNP and CRC prognosis. Thirdly, this investigation was conducted within a single medical center, China Medical University Hospital (CMUH). Although we have extinguished the possibility of population heterogeneity, further multi-center studies with larger sample sizes in the future are important to validate the current findings. Finally, given the small sample size, we only studied two SNPs. Future studies should include a more comprehensive, genome-wide analysis of SNPs in a significantly larger number of subjects.
In conclusion, the findings of the current study demonstrate that the C allele and CC genotype of mir196a-2 rs11614913 are closely correlated to the increased CRC risk among the Taiwanese. Moreover, the CC genotype is correlated with significantly higher levels of serum mir196a-2 expression. Therefore, in conjunction with mir196a-2 rs11614913 genotyping, the elevated serum levels of mir196a-2 may serve as a novel circulating marker for the early detection of CRC.