Meta-analysis of the relationship between MnSOD polymorphism and cancer in the Turkish and Cypriot population MnSOD polimorfizmi ile kanser arasındaki ilişkinin Türk ve Kıbrıs toplumundaki meta

Objectives: The association between manganese superoxide dismutase (MnSOD) p.Val16Ala polymorphism and cancer has been shown in various studies. The aim of this study is to investigate the relationship between MnSOD polymorphism (V/V, V/A, A/A) and cancer in the Turkish and Cypriot population through meta-analysis. Material and methods: The present study included meta-analysis of 14 publications covering 2413 cancer patients and 2907 healthy control groups from 2005 to 2016. Pooled odds ratio (OR) and 95% confidence interval (CI) were calculated using the random effect model of DerSimonian and Laird for each study. Publication bias was checked with funnel plot by Begg’s and Egger’s test statistics. Results: Meta-analysis of MnSOD polymorphism was performed in the additive model (AV vs. VV; OR = 1.133, 95% CI: 1.002–1.282), allele contrast (A vs. V; OR = 1.016, 95% CI: 0.930–1.278), homozygote model (AA vs. VV; OR = 0.983, 95% CI: 0.839–1.153), dominant model (AA + AV vs. VV; OR = 1.090, 95% CI: 0.971–1.223) and recessive model (AA vs. AV + VV; OR = 0.924, 95% CI: 0.803–1.064). The A/V genotype polymorphism was found be significant for cancer. Conclusion: The frequency of the A/V heterozygote genotype of the MnSOD polymorphisms is found to be higher in the Cypriot and Turkish populations than any other genotype.


Introduction
Manganese superoxide dismutase (MnSOD) is an antioxidant enzyme located in the mitochondrial matrix [1]. MnSOD that has tumor suppressor activity, is encoded by a gene located on chromosome 6q25.3 [2]. MnSOD plays an important role in the protection of cells from oxidative damage induced by reactive oxygen species (ROS) and catalyzes the destruction of superoxide radicals in hydrogen peroxide and oxygen [1]. MnSOD is a nuclear protein transported after translocation through the N-terminal signal sequence in mitochondria [3,4]. The signal sequence is important for protein transport activity by mitochondria [5]. The polymorphism on the second exon of the MnSOD gene (SOD2), and the change from valine to alanine (Val16Ala) position at the amino acid position 16, results in the signal sequence at position -9 (Ala-9Val) of the enzyme MnSOD (rs4880) [6]. This polymorphism may alter the mitochondrial transport and structural conformation of MnSOD [6,7]. Consequently, alanine-containing protein exhibitions normally transport 30-40% more active enzyme forms than the valine form of the enzyme [8]. The low expression of MnSOD is suggested to be responsible for different types of cancer. Additionally, this protein overexpression inhibits the proliferation of the cancerous cell, indicating that it is a tumor suppressor gene [9]. MnSOD may act as a tumor suppressor by altering pathways, including cellular apoptosis and proliferation [10].
The association between MnSOD Val16Ala polymorphism and cancer has been shown in various studies. The Val allele and Val/Val genotype are found to be related with an increased risk of bladder and lung cancers [11,12]. Ala allele is related with the increased risk of prostate, ovarian and breast cancers [13][14][15][16][17]. However, the results of some molecular epidemiological studies have presented   contradictory results regarding the relationship between MnSOD Ala-9Val gene polymorphism and cancer risk [18,19]. Ala allele was only observed in individuals with breast cancer risk in only three populations, among many others [9,13,15,20]. The Val allele is associated with lung and bladder cancer; while Woodson et al. and Kang et al. showed that the Ala allele was related with prostate cancer [12,17,21]. The aim of this study is to investigate the relationship between MnSOD polymorphism and cancer in the Turkish and Cypriot populations through meta-analysis.

Literature search
For the meta-analysis, PubMed, Web-of-Science, Turkish Medline, Turkish Journal of Medical Science, Journal of Turkish Oncology, the Turkish Council of Higher Education Theses Database and Google scholar were searched and data was obtained from the electronic databases using the keywords "MnSOD polymorphism, cancer, Turkish population and Cypriot population". Moreover, keywords were searched in both Turkish and English languages.

Selection criteria
The case-control groups of the studies on MnSOD polymorphism and cancer related to the Cypriot and Turkish population were included in the meta-analysis. In the investigation of the MnSOD polymorphism (V/V, V/A and A/A genotype) and cancer association, the genotype frequencies were detailed in the control and case groups and constituted the criteria for inclusion in the meta-analysis study. In the present study, publications in the control group that did not match the Hardy-Weinberg equality   were not excluded from the meta-analysis. There is no consensus on the use of control groups that do not comply with the Hardy-Weinberg equation (HWE) in statistical analysis [22]. It has also been observed that this situation was not accepted as the exclusion criteria in similar metaanalyses of different populations [23,24] (Figure 1).

Exclusion criterion
The control group was not included in the studies that were not composed of healthy individuals, the letter of the editor, meta-analysis publications and the studies without a control group or only healthy individuals were not included in the meta-analysis.

Statistical analysis
Meta-analysis was performed using STATA software version 14.2 (StataCorp, College, TX, USA). The pooled odds ratio (OR) and 95% confidence interval (CI) values for each study were calculated using the random effect model of DerSimonian and Laird and forest plots were   generated. As the studies were conducted on the Turkish and Cypriot population, a random effect model is used for heterogeneity. Publication bias was checked using the Begg's and Egger's test regression statistics. A funnels-plot graph was used to assess neutrality. Deviations from HWE in the control groups were calculated using the Genhwcci Stata module and power of the Q-test was calculated by using the PowerQ Stata module. A p < 0.05 was considered significant.

Results
A total of 12 publications and two unpublished theses covering 2413 cancer patients and 2907 healthy control groups were analyzed. The patients included breast (68.59%), prostate (11.02%), bladder (6.51%), head and neck and laryngeal (5.76%), lung (4.14%), ovarian (2.28%), and renal cell (1.70%) cancers. In analyses of the MnSOD gene; the frequencies of V/V, V/A and A/A genotypes were found to be 36.15%, 44.86% and 18.99% in the control group and 35.02%, 47.37% and 17.61% in the cancer group, respectively (Table 1). Ala and Val alleles were found as 58.58%, 41.42% in the control group and 58.70% and 41.30% in the cancer group, respectively.
Meta-analysis results, Heterogeneity test, Publication Bias results and Power analysis are given in Table 2. There was a significant correlation between the additive genetic model (AV vs. VV: OR = 1.133, 95% CI: 1.002-1.282), and cancer ( Figure 2). The risk of cancer in heterozygous individuals is increased 1.13-fold. The highest risk group in this model is lung cancer. In heterozygous individuals in the Turkish population,     (Figures 4-7). Higher heterogeneity (I 2 > 50%) was determined in all models. In this meta-analysis study, evaluated in terms of Publication Bias, no bias was found (Figure 8) except for the recessive genetic model (p = 0.014).

Discussion
Meta-analysis is conducted to evaluate the results of working with a large number of different studies that belong to a specific domain. The meta-analysis findings are the result of statistical evaluation of large-scale aggregated studies [31,32]. The aim is to investigate the relationship between MnSOD polymorphism (A/A, A/V, V/V) and cancer by using meta-analysis. For this purpose, 14 publications were examined.
Recently, different polymorphisms of the MnSOD gene have been noted in the etiology of various cancers. The MnSOD Val16Ala polymorphism has been shown to be associated with the development of various types of cancer such as PCA, breast and lung [14,17,[37][38][39][40][41][42][43]. In a meta-analysis study on prostate cancer, it was observed that the MnSOD Val16Ala polymorphism increased the   [10,[44][45][46]. Prostate cancer cells were increased in hydrogen peroxide levels. H 2 O 2 is an intracellular oxidase, which induces DNA degradation in prostate cancer cells [47]. The different ala allele has been associated with an increased risk of breast cancer among premenopausal women with low antioxidant consumption [48].
Val16Ala polymorphism is associated with breast cancer. Bergman et al. [49] found that Val/Val and Val/ Ala [OR = 2.7; (95% CI), 2.2-5.5 and OR = 3.0; 95% CI: 1.4-6.5] have increased breast cancer risk in individuals with genotypes. The Ala allele has been found to be associated with prostate cancer risk in the Turkish population [14]. In this meta-analysis study conducted on the Cypriot and Turkish populations, breast cancer risk (OR = 0.97, 95% CI: 0.83, 1.13) was found to increase in the A/V heteozygotic model. In the homozygous, recessive, and dominant models of the MnSOD polymorphisms, it   is associated with a decreased risk of lung cancer. In the study by Sun et al. [50], the Ala allele has been shown to be associated with prostate and esophageal cancer risk, but not lung cancer. Low MnSOD expressions in the cells lead to increased ROS in the mitochondria. Increased accumulated ROS causes genomic instability, DNA, protein and lipid breakdown [51,52]. In a study, supporting this finding, MnSOD Ala/Ala genotype was associated with a 1.7-fold increased risk of prostate cancer, a 1.9-fold increase risk of esophageal cancer, and a significant risk of breast cancer [15,17,53] In conclusion, our meta-analysis with the Cyprus and Turkish populations showed that the A/V heterozygote genotype of MnSOD polymorphism is significant, and that genetic factors play a role in cancer development. Increase of ROS due to MnSOD polymorphism in the mitochondria may cause mtDNA degradation, which leads to cancer risk. More related studies to extend the effective risk factors in cancer patients from these populations are needed.