Association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with susceptibility to breast cancer – a meta-analysis

Summary

Background: Previous studies have evaluated the association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with a risk of breast cancer in different populations, but the results remain inconsistent and inconclusive. Thus, we performed this meta-analysis to explore the associations. Methods: A comprehensive literature search in PubMed, EMBASE, Web of Science, Scopus, SciELO, SID, and CNKI for all eligible studies published up to October 1, 2020. The pooled odds ratios (ORs) with 95% confidence intervals (CIs) were used to evaluate the intensity of associations. Results: A total of 12 case-control studies including seven studies with 2,370 cases and 2,314 controls on IL-8 -251T>A, and five studies with 900 cases and 882 con­trols on IL-18 -607C>A polymorphism were selected. Pooled data showed that IL-8 -251T>A (AT vs. TT: OR= 1.187; 95% CI 1.038–1.356; P = 0.012) and IL-18 -607C>A polymorphisms (A vs. T: OR = 1.205; 95% CI 1.055–1.377; P = 0.006; AA vs. TT: OR = 1.379; 95% CI 1.056–1.802; P = 018; and AA vs. AT+TT: OR = 1.329; 95% CI 1.053–1.678; P = 0.017) were associated with increased risk of breast cancer in overall. Moreover, when the studies were stratified by ethnicity, the IL-8 -251T>A was significantly associated with breast cancer risk in Africans. Publication bias tests provide no evidence of presence of publication bias in a meta-analysis. Conclusion: This meta-analysis results revealed that the IL-8 -251T>A and IL-18 -607C>A polymorphisms are associated with susceptibility to breast cancer. However, further multicenter studies with larger sample sizes in different ethnicities are required to make a better assessment of these associations.

Keywords:

meta-analysis – breast cancer – association – interleukin-8 – interleukin-18

Introduction

Breast cancer is the most common form of cancer and the leading cause of death in women of all ethnic groups [1,2], especially in case of low- and middle-income developing countries due to screening barriers [3]. In 2018, an estimated 266,000 new cases of breast cancer will be dia­gnosed with 41,000 cases of deaths, were estimated to occur in women in the United States [4]. In the literature, obesity, hormone replacement therapy, radiation, age, and family history has been described as a pivotal risk factor for breast cancer development [5–8]. Approximately 4% of cases dia­gnosed with breast cancer in the United States are younger than 40 years and a report by National Cancer Institute showed that the breast cancer incidence rate in women aged 21–54 years was 33.6% of all breast cancer  cases [9,10].

Despite its high prevalence and rapid progress in molecular bio­logy of cancer achieved in the last few decades, the exact mechanism of breast cancer is still poorly understood [11,12]. It is evident that genetic and environmental factors play a role in the development of breast cancer [13,14]. Moreover, there is also a relationship between social environment and breast cancer development and woman cope with a breast cancer dia­gnosis. Epidemiological studies have demonstrated that single nucleotide polymorphisms (SNPs) in different interleukins might be involved in development of breast cancer [15,16].

Recently, the roles of interleukin 8 (IL-8) and IL-18 have been extensively studied in the development of breast cancer. Human IL-8 enhances the immune system against cancer cells and also modifies the tumor microenvironment facilities [17]. Increasing evidence demonstrates that human IL-8 is considerably expressed in ER-, PR- and HER-2/neu+ breast cancer cells, but highly correlated with invasiveness and metastatic of both ER- and ER+ cells by twofold, indicating the invasion-promoting role of IL-8 [17–20]. IL-8 is associated with growth receptors expressed on the surface of breast cancer cells. However, the mechanisms by which IL-8 contributes to breast cancer progression have remained poorly understood [21]. Moreover, IL‐18 may also have role in development of breast cancer. Previous studies revealed that serum levels of IL-18 were higher in advanced than in early stages and higher in metastatic than in non-metastatic breast cancer cases [22]. In the recent decade, several studies have investigated the influence of IL-8 -251T>A and IL-18 -607C>A polymorphisms in breast cancer, but their results remained controversial. Therefore, we performed meta-analyses to evaluate and summarize the contribution of the IL-8 -251T>A and IL-18 -607C>A polymorphisms to breast cancer susceptibility.

Materials and methods

Identification and eligibility of relevant studies

This study was performed according to the Preferred reporting items for systematic reviews and meta-analyses (PRISMA) statement on reporting meta- -analysis. We conducted a comprehensive literature on PubMed, EMBASE, Web of Science, Science Citation Index (SCI), Springer Link, CNKI, Wanfang, OVID, SID, EBSCO and Science Direct databases to identify potential studies evaluated the association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with a risk of breast cancer. The search keywords used were as follows: (‘’Breast Cancer’’ OR ‘‘Tumor’’ OR ‘’Cancer’’ OR ‘’Neoplasm’’) AND (‘’Interlukine-8 ‘’ OR ‘’IL-8’’ OR ‘’-251A>T’’ OR ‘’rs4073’’) AND (‘’Interlukine-18 ‘’ OR ‘’IL-18’’ OR ‘’-607C>A’’ OR ‘’rs1946518’’) AND (‘’Gene’’ OR ‘’Polymorphism’’ OR ‘’SNPs’’ OR ‘’Mutation’’ OR ‘’Variation’’ OR ‘’Allele’’). Then, the references of all retrieved publications and reviews to identify other potential relevant studies were checked manually. The final literature search was updated on October 1, 2020, with no restrictions on publication year or methodological filter. Data for the largest sample set or most recent published articles were included when data from the same set of cases were used more than once within a publication.

Selection criteria

The eligible studies included in the current meta-analysis, must meet all the following criteria:

1) genetic association studies on association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with breast cancer;

2) in human beings;

3) studies with case-control or cohort design;

4) allele and genotype distribution for both IL-8 -251 T>A and IL-18 -607 C>A polymorphisms provided to estimate the odds ratios (ORs) and 95% confidence intervals (CIs);

5) articles published in English, Chinese, Portuguese and Farsi (with an English abstract).

The studies were excluded if one of the following criteria was fulfilled:

1) studies not about IL-8 -251T>A and IL-18 -607C>A polymorphisms and breast cancer;

2) studies on other polymorphisms at IL-8 and IL-18 genes;

3) animal models, cell lines and in vitro studies;

4) no control population;

5) linkage and family-based studies;

6) data unavailable or insufficient for calculating allele and genotypes frequencies;

7) abstracts, posters, case reports, case series, editorials, letters, editorial articles, conference presentations, com­ments, reviews, meta-analyses;

8) overlapping data or duplicate of previous publication.

Data extraction

The necessary data were carefully extracted by two observers from eligible studies according to the mentioned criteria. Any disagreement between two authors was discussed with the third author until a consensus was achieved. If the data were not presented, the corresponding authors were contacted to request extra information. The following characteristics were collected from each study: last name of first author, year of publication, country of origin, ethnicity of study participants, source of controls (hospital based or population based), genotyping methods, genotypic/allelic distributions for both IL-8 -251T>A and IL-18 -607C>A polymorphisms in cases and controls, the number of cases and control genotypes, minor allele frequency (MAFs) and Hardy-Weinberg equilibrium (HWE) in controls.

Statistical Analysis

The strength of associations of IL-8 -251T>A and IL-18 -607C>A polymorphisms with susceptibility to breast cancer was estimated by ORs with 95% confidence intervals (95% CIs). The significance of the pooled effect size was examined by the Z-test. The associations were evaluated under all five genetic models, i.e. allele (B vs. A), homozygote (BB vs. AA), heterozygote (BA vs. AA), dominant (BB+BA vs. AA), and recessive (BB vs. BA+AA). To take into account the possibility of between-study heterogeneity, the Cochran’s Q test was used, in which P < 0.10 indicated a significant heterogeneity. Moreover, I² statistic (range of 0–100%) was also employed to qualify the heterogeneity, where a lower value represents non-significant heterogeneity and a higher value represents a high degree of between study heterogeneity. A fixed-effect method (the Mantel-Haenszel method) was applied to calculate the pooled ORs and the corresponding 95% CIs for all genetic models which did not show significant heterogeneity; otherwise a random-effect method (the DerSimonian and Laird method) was adopted. Subgroup analyses were conducted by stratification of ethnicity to identifying potential source of heterogeneity. HWE was tested in control groups of each study by c² test to assess the latent bias resulting from the deviation of the genotype distribution; P > 0.05 were considered to have reliable and representative controls. The sensitivity analysis was carried out by omitting one study at a time to determine the underlying effects of each single study on the pooled data. Furthermore, we conducted the sensitivity analyses again to delete those studies deviating from HWE and calculate the pooled ORs for the remainder of the studies. Publication bias was performed by the construction of Begg’s funnel plot and Egger’s regression analysis. All of the statistical calculations were performed using Comprehensive Meta-Analysis (CMA) software, version 2.0 (Biostat, USA). For all analyses, statistical significance was assumed at P < 0.05, unless otherwise stated.

Results

Selected studies characteristics

The selection process of eligible studies is presented in Scheme 1. Initially, 108 studies were obtained through publication search in electronic databases, and one study was identified from other sources. Irrelevant articles were excluded by evaluating the titles and abstracts. Therefore, 76 publications were deleted for obvious irrelevance. Finally, 12 case-control studies including seven studies [23–29] with 2,370 cases and 2,314 controls on IL-8 -251T>A polymorphism, and 5 studies [30–34] with 900 cases and 882 controls on IL-18 -607C>A polymorphisms were selected to the meta-analysis. Tab. 1 describes principal characteristics of selected studies. All the studies were published between December 2004 and March 2018. The studies have been carried out in UK, Tunisia, Iran, China, and Brazil. Among these studies, eight studies were conducted among Asians, two studies among Caucasians and two studies among Africans. Seven different genotyping methods were used: ARMS-PCR, AS-PCR, TaqMan, ASO-PCR, PCR-RFLP, MALDI-TOF and direct sequencing. The genotype, allele and MAF in each study for both IL-8 -251T>A and IL-18 -607C>A polymorphisms are shown in Tab. 1. More­- over, the distribution of genotypes in the controls was in agreement with HWE for all selected studies, except for one study on IL-8 -251T>A polymorphism   (Tab. 1).

Quantitative synthesis

IL-8 -251T>A polymorphism

The association between IL-8 -251T>A polymorphism and breast cancer risk is shown in Tab. 2. Pooled data showed that there was a significant association between IL-8 -251T>A polymorphism and an increased risk of breast cancer under heterozygote model (AT vs. TT: OR = 1.187; 95% CI 1.038–1.356, P = 0.012, Fig. 2A). When stratified analysis by ethnicity, a significant association was found between IL-8 -251T>A polymorphism and breast cancer risk in Africans under all five genetic models, i.e. allele (A vs. T: OR = 1.398; 95% CI 1.174–1.663; P ≤ 0.001), homozygote (AA vs. TT: OR = 2.083; 95% CI 1.529–2.839; P ≤ 0.001), heterozygote (AT vs. TT: OR = 1.411; 95% CI 1.066–1.867; P = 0.016), dominant (AA+AT vs. TT: OR = 1.676, 95% CI 1.295–2.168; P ≤ 0.001), and recessive (AA vs. AT+TT: OR = 1.364; 95% CI 1.055–1.764; P = 0.018), but not in Asians and Caucasians (Tab. 2).

IL-18 -607C>A polymorphism

Results of pooled analysis for IL-18 -607C>A polymorphism and breast cancer risk are summarized in detail in Tab. 2. Pooled data showed that the IL-18 -607C>A polymorphism was significantly associated with an increased risk of breast cancer under three genetic models, i.e. allele (A vs. T: OR = 1.205; 95% CI 1.055–1.377; P = 0.006, Fig. 2B), homozygote (AA vs. TT: OR = 1.379; 95% CI 1.056–1.802, P = 018), and recessive (AA vs. AT+TT: OR = 1.329; 95% CI 1.053–1.678; P = 0.017). However, stratified analysis by ethnicity failed to show a significant association between the IL-18 -607C>A polymorphism and breast cancer in Asians (Tab. 2).

Heterogeneity test

In this meta-analysis there was statistical significance between-study heterogeneity for IL-8 -251T>A polymorphism under four genetic models, i.e. allele (A vs. T: I² = 83.32; PH ≤ 0.001), homozygote (AA vs. TT: I² = 87.78; P_H ≤ 0.001), dominant (AA+AT vs. TT: I² = 66.65; P_H = ?) and recessive (AA vs. AT+TT: I² = 85.38; P_H ≤ 0.001), models and IL-18 -607 C>A polymorphism under dominant model (AA+AC vs. CC: I² = 68.65; P_H = 0.013). Therefore, we conducted subgroup analyses by ethnicity to explain the potential source of heterogeneity. Results showed that the heterogeneity disappeared in the subgroup analysis among Asians, Caucasians and Africans, indicating that ethnicity might be the major source of heterogeneity for IL-8 -251T>A polymorphism (Tab. 2).

Sensitivity analysis

While omitting each individual study any time, sensitivity analysis was applied to detect the influence of each study on the pooled OR by repeating the meta-analysis. Sensitivity analyses results showed that the significance of the pooled data for both for IL-8 -251 T>A and IL-18 -607 C>A polymorphisms was not affected by any single study in the overall population. In addition, when we excluded the studies out of HWE, the statistical significance of the results did not change.

Publication bias

Begg’s funnel plot and Egger’s test were used to evaluate the publication bias of the literature for IL-8 -251T>A and IL-18 -607C>A polymorphisms. No publication bias was detected with either the Begg’s funnel plot or the Egger’s tests under all five genetic models in overall and subgroup analysis by ethnicity (Tab. 2). Funnel plots for IL-8 -251T>A polymorphism in allele model and IL-18 -607C>A polymorphism in recessive model were showed in Graphs 3A, B.

Discussion

Human IL-8, also known as neutrophil chemotactic factor, has significant potential as a prognostic and predictive bio­marker in various inflammatory conditions and malignancies [35]. It plays a key role in the recruitment of neutrophils and other immune cells to the site of infection [36]. IL-8 is mapped on chromosome 4q13-q21, contains four exons, a proximal promoter region and has a length of 5191 bp [37,38]. Moreover, human IL-18, a member of the IL-1 family, was initially identified as a protein that induces interferon g (IFNg) production [39,40]. It is a pro-inflammatory chemokine, plays a key role in the initiation, modulation, and maintenance of the gastrointestinal inflammatory response. Human IL-18 gene is mapped on 11q22.2–22.3, contains six exons and several genetic polymorphisms, especially in the promoter region [41]. A functional polymorphism at position -251 of the IL-8 promoter region has been identified in 2000, which has effect on IL-8 gene expression or secretion [42].

In this study, we evaluated the association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with breast cancer risk based on all available studies. By pooling all eligible studies, we found the IL-8 -251T>A and IL-18 -607C>A polymorphisms are significantly associated with an increased risk of breast cancer in the global population. Previous epidemiological studies have reported that these polymorphisms were associated with an increased risk of breast cancer in different populations such as Chinese, Iranian and Danish. However, subgroup analyses by ethnicity showed lack of association between IL-8 -251T>A and IL-18 -607C>A polymorphisms and a risk of breast cancer in Asians and Caucasians. This difference is a common finding in a meta-analysis and could be explained by two reasons. First, because of the complex nature of breast cancer, it is unlikely that a single nucleotide polymorphism in IL-8 and IL-18 genes would be associated with an increased risk of breast cancer, without an interaction from other polymorphic susceptibility genes. Second, other factors, such as age, hormone therapy, family history, life style and different environmental exposure can also influence the development of breast cancer. In 2014, Wang et al evaluated the association between IL-8 -251T>A polymorphism and breast cancer risk in a meta-analysis of five case-control studies [15]. Their results showed that IL-8 -251T>A polymorphism was significantly associated with an increased risk of breast cancer under four genetic models, i.e. allele (A vs. T, OR = 1.21; 95% CI 1.01–1.45), heterozygote (AT vs. TT, OR = 1.28; 95% CI 1.07–1.53), dominant (AA+TT vs. TT: OR = 1.34, 95% CI 1.03–1.74) and recessive (AA vs. AT+TT: OR = 1.25; 95% CI 1.05–1.49). In 2015, Li et al have examined the association of IL-18 -607C>A polymorphism with risk of breast cancer in a meta-analysis of three case-control studies [43]. They have found that IL-18 -607C>A poly­morphism was significantly associated with an increased risk of breast cancer under three genetic models, i.e. allele (A vs. C: OR =1.33; 95% CI 1.00–1.75; P_H = 0.155), homozygote (AA vs. CC: OR = 1.80; 95% CI 1.02–3.21, P_H = 0.162) and dominant (AA+CA vs. CC: OR = 1.33; 95% CI 1.00–1.78; P_H = 0.546). Considering that the samples of breast cancer cases and controls in the previous meta-analyses were quite small, their results might not have enough statistical power. Moreover, their estimates were not adjusted OR values such as ethnicity, which might be caused to inaccurate results. Therefore, our meta-analysis gave strong evidence of an association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with a risk of breast cancer in global population and by ethnicity.

Between-study heterogeneity significantly affects a meta-analysis result. Several factors such as diversity in study populations, study design, sample size, source of controls, genotyping method, and WHE might contribute to potential sources of heterogeneity [44–46]. There was a significant heterogeneity in this meta-analysis mostly for IL-8 -251T>A polymorphism. Thus, due to the significant heterogeneity, we applied the random-effects model to calculate the pooled ORs, which could provide stable results. Subgroup analyses by ethnicity showed that the heterogeneity significantly decreased in Asians, Caucasians and Africans, indicating that ethnicity might be the major source of heterogeneity for IL-8 -251T>A  polymorphism.

We did not observe any publication bias in both IL-8 -251T>A and IL-18 -607C>A polymorphisms, demonstrating that the results of this meta-analysis are stable. However, it is important to note the limitations of our meta-analysis. First, although an increased risk of breast cancer was observed to be associated with IL-8 -251T>A and IL-18 -607C>A polymorphisms; the sample size for both polymorphisms was not large enough to provide enough statistical power. Second, in the current meta-analysis studies from Caucasians, Asians, Africans and mixed populations were involved, although the number of studies was relatively small and results might not have enough statistical power to obtain the association of IL-8 -251T>A and IL-18 -607C>A polymorphisms with breast cancer. Third, only published studies in English were included; it is possible that some relevant published or unpublished studies with null results were missed, which might have biased the results and causing a language bias. Fourth, several important confounding factors, such as age, smoking, drinking, family history of breast cancer, environmental exposures and lifestyle, were not considered for stratification analysis because relevant data was insufficient in the primary reports. Finally, the lack of original data in the eligible studies limited the evaluation of the effects of gene-gene and gene-environment interactions on IL-8 -251T>A and IL-18 -607C>A polymorphisms and breast cancer risk.

Conclusion

Our results showed that IL-8 -251T>A and IL-18 -607C>A polymorphisms are significantly associated with increased susceptibility to breast cancer. Our findings also indicate that the IL-8 -251T>A polymorphism also plays an important role in the development of breast cancer in Africans. Future studies with large sample sizes and more ethnic groups are needed to confirm our findings. More­over, IL-8 -251T>A, IL-18 -607C>A polymorphisms, other interleukin polymorphisms and gene-gene interactions should also be considered in future studies.

Ethical approval and consent  to participate

The ethical approval was not required for this study, as it is a systematic review and meta-analysis.

Availability of data and material

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Funding

No funding was obtained for this study.

Authors Contribution

Conceived and designed the study and experiments: MF, FA and SAD

Performed the experiments: HN and MKZ

Analyzed the data: SHS, SK and HN

Contributed reagents/materials/analysis tools: MZS, SAD and FA

Wrote the paper: MF, SAD and HN

All authors have read and approved the manuscript.

Acknowledgements

I would like to express my sincere gratitude to Professor Seyed Mehdi Kalantar for his motivation, knowledge and support during the course of this research.

The authors declare they have no potential conflicts of interest concerning drugs, products,or services used in the study.
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Submitted/Obdrženo: 16. 8. 2020
Accepted/Přijato: 24. 1. 2021

Prof. Seyed Alireza Dastgheib
Department of Medical Genetics,
School of Medicine,
Shiraz University of Medical
Sciences, Shiraz, Iran
e-mail: dastgheibsa@gmail.com