FGFR4 Gly388Arg polymorphism contributes to prostate cancer development and progression: A meta-analysis of 2618 cases and 2305 controls

Background Fibroblast growth factor receptor 4 (FGFR4) displays multiple biological activities, including mitogenic and angiogenic activity, and plays important roles in the etiology and progression of prostate cancer. Gly388Arg polymorphism in FGFR4 gene has been reported to be involved in prostate cancer incidence and aggressiveness in several studies. To derive a more precise estimation of the relationship, a meta-analysis was performed. Methods Odds ratios (ORs) with 95% confidence intervals (CIs) were estimated to assess the association. Results The Arg388 allele increased prostate cancer risk compared with Gly388 allele (OR = 1.17, 95% CI = 1.07-1.29). When stratified by race, there was a significantly increased prostate cancer risk in Asian and Caucasian populations. Moreover, prostate cancer patients with Arg/Arg genotype had a 1.34-fold increased risk of advanced prostate cancer (95% CI: 1.03-1.74) compared with those with Gly/Gly+Gly/Arg genotype. Conclusion This meta-analysis showed the evidence that FGFR4 Gly388Arg polymorphism was associated with an increased risk of prostate cancer development and progression, suggesting that FGFR4 Gly388Arg polymorphism could be a marker for prostate cancer development and progression.


Background
Prostate cancer is the most frequently diagnosed solid tumor and the second leading cause of cancer-related death among American men, with an estimated 192,280 new cases and 27,360 deaths in the United States in 2009 [1]. The etiology of human prostate cancer is complex and largely remains unknown.
Fibroblast growth factor receptor 4 (FGFR4) belongs to the family of fibroblast growth factor receptors (FGFR1-4), which display multiple biological activities, including mitogenic and angiogenic activity, with a consequent crucial role in cell differentiation, development, hormonal and proliferative signaling in response to more than 20 known ligands [2,3]. In light of its involvement in the regulation of essential biologic mechanisms, FGF signaling is also likely to play a role in tumor growth and progression; indeed, dysreglation of this pathway has been demonstrated in several tumor types [3]. Recently, FGFR4 was found to be more abundantly expressed in malignant than benign prostate cells and in vitro suppression of FGFR4 expression effectively blocked prostate cancer proliferation and invasion [4]. Moreover, strong expression of FGFR4 in prostate cancer cells, as assessed by immunohistochemistry, is significantly associated with increased clinical stage and tumor grade and decreased patient survival [5].
A germ line polymorphism in FGFR4 gene, resulting in different expression of FGFR4 containing either glycine (Gly 388 ) or arginine (Arg 388 ) at codon 388 in the transmembrane domain of the receptor was identified several years ago. In addition, the FGFR4 Arg 388 allele may predispose cancer patients to disease progression, based on the reported significant association between FGFR4 genotype and tumor aggressiveness or patients' survival in several cancers [6,7].
To date, several studies had been reported to focus on the association between this polymorphism and incidence and aggressiveness of prostate cancer [4,[8][9][10][11][12]. However, a single study may be too underpowered to detect a possible small effect of the polymorphism on prostate cancer, especially when the sample size is relatively small. Hence, we carried out a meta-analysis of all eligible case-control studies to derive a more precise estimation of the association of FGFR4 Gly 388 Arg polymorphism with prostate cancer.

Publication search
PubMed and EMBASE were searched (the last search update on the 1 st Nov. 2010) using the search terms: 'FGFR4 or fibroblast growth factor receptor 4', 'polymorphism', 'Gly 388 Arg or rs351855' and 'prostate cancer or prostate neoplasm'. All published English language papers with available full text matching the eligible criteria were retrieved. In addition, we checked all the references of relevant reviews and eligible articles that our search retrieved. Two investigators (BX and SQC) searched the literature and extracted data independently.

Inclusion, exclusion criteria and data abstraction
For inclusion in the meta-analysis, the identified articles had to provide information on: (1) evaluation of FGFR4 Gly 388 Arg polymorphism and prostate cancer risk, (2) using a case-control design and (3) containing information about available genotype frequency that can help infer the results in the papers. Major reasons for the exclusion of studies were: (1) no control population; (2) no usable data reported; (3) duplicates. For each of the eligible case-control studies, the following data were collected: the first author's last name, year of publication, country of origin, ethnicity, numbers of genotyped cases and controls, genotyping methods.

Statistical analysis
The strength of the association between the FGFR4 Gly 388 Arg polymorphism and prostate cancer risk was measured by ORs with 95% confidence intervals (CIs). We explored the association between allele Arg 388 and prostate cancer development and progression, as well as homozygote comparison (Arg/Arg vs. Gly/Gly), dominant genetic model [(Gly/Arg+Arg/Arg) vs. Gly/Gly] and recessive model [Arg/Arg vs. (Gly/Gly+ Gly/Arg)]. Heterogeneity assumption was checked by a chi-squarebased Q-test [13]. A P-value of more than 0.05 for the Q-test indicated a lack of heterogeneity among the studies, so the summary OR estimate of each study was calculated by the fixed-effects model (the Mantel-Haenszel method). Otherwise, the random effects model (DerSimonian and Laird method) was used [14,15]. The significance of the pooled OR was determined by the Z-test, and P < 0.05 was considered as statistically significant. To evaluate the ethnic-specific effect, subgroup analysis was conducted on the basis of different ethnicities.
Evidence of publication bias was determined using Begg's [16] and Egger's [17] formal statistical test and by visual inspection of the funnel plot. All statistical analyses were performed with Stata software (version 10.0; StataCorp LP, College Station, TX), using two-sided P values.

Study characteristics
Using the searching terms, seven papers were reviewed in the two online databases. The study of Spinola et al. [18] was focused on the association between FGFR4 Gly 388 Arg polymorphism and lung cancer risk, and the studies of Wang et al. [12] and Sahadevan et al. [4] were not epidemiological association studies, so they were all excluded in present study. In the four papers left, Fitz-Gerald et al. [8] and Wang et al. [11] provided data on both African-American and Caucasian. Overall, four articles (six studies) with 2618 prostate cancer cases and 2305 controls were retrieved based on the search criteria for prostate cancer susceptibility related to the FGFR4 Gly 388 Arg polymorphism. Study characteristics are summarized in Table 1. All the studies used frequencymatched controls to the cases by the age, sex or ethnicity, and the distribution of genotypes in the controls was consistent with Hardy-Weinberg equilibrium in all studies. Moreover, among the four articles, three [8,10,11] mentioned the association between FGFR4 Gly 388 Arg polymorphism and progression of prostate cancer. The stratifications for pathological parameters of the cases in the three articles were also shown in Table 1. The cases of the three articles were all stratified by Gleason score and tumor stage. However, the classification standard of Gleason score was not uniform; thus, we only focused on the association between FGFR4 Gly 388 Arg polymorphism and tumor stage (advanced vs. localized). Advanced stage corresponded to T3 stage in the study of Wang et al. [11], regional/distant stage in the study of FitzGerald et al. [8], and stage C+D in the study of Ma et al. [

Quantitative synthesis
We observed a wide variation of Arg 388 allele frequencies across different ethnicities. The frequency of Arg 388 allele was 28.90% among Caucasian controls and 41.57% among Asian controls, which were significantly higher than that in African-American controls (11.49%, P < 0.01).
Overall, the combined result based on all studies showed the evidence of an association between the increased risk of prostate cancer and the variant genotypes in different genetic models. As shown in Table 2 and Figure 1, the Arg 388 allele increased overall prostate cancer risk compared with Gly 388 allele (OR = 1.17, 95% CI = 1.07-1.29). Significant main effects were also observed in dominate genetic model (OR = 1.21, 95% CI = 1.08-1.36).
When stratifying for race, results were similar. Specially, significantly increased risk was found among Caucasian populations (Arg 388 and Gly 388 comparison: OR = In addition, when concerning tumor stage and FGFR4 Gly 388 Arg polymorphism, patients with prostate cancer with Arg/Arg genotype had a 1.34-fold increased risk of advanced or metastatic prostate cancer (95% CI: 1.03-1.74) compared with the Gly/Gly+Gly/Arg genotype (seen Figure 2).

Sensitivity analysis
Sensitivity analysis was performed by sequential omission of individual studies. The pooled 95% CI for Arg 388 vs. Gly 388 was consistently over 1.0, indicating that the results of this meta-analysis are stable.

Publication bias
Begg's funnel plot and Egger's test were performed to assess the publication bias. The shape of the funnel plots seemed   symmetrical in the comparison of the Arg 388 vs. Gly 388 ( Figure 3). Furthermore, Egger's test was used to provide statistical evidence for funnel plot symmetry (t = 1.30, P = 0.26), suggesting that no publication bias was exist.

Discussion
The present meta-analysis, including 2,618 cases and 2,305 controls from six published studies, explored the association between FGFR4 Gly 388 Arg polymorphism and development and progression of prostate cancer. To the best of our knowledge, this is the first meta-analysis to explore FGFR4 Gly 388 Arg polymorphism in development and progression of prostate cancer. The results indicated that FGFR4 Arg 388 allele is a potential risk factor for developing and progressing prostate cancer. These findings may be biologically plausible. The FGFR4 Gly 388 Arg polymorphism results in an amino acid change in the transmembrane domain of the receptor, which may alter the activity of the receptor. FGFR4 is the activator of the MAPK signaling cascade, yet it is a principal receptor for key mitogenic FGFs in prostate cancer cells [19][20][21]. There was evidence that FGFR4 contributed to progression in liver, lung, colon tumors [22] and prostate cancer [12]. The effects of FGFR4 Arg 388 allele may also predispose cancer patients to disease progression, based on the reported significant association between FGFR4 genotype and tumor aggressiveness (lymph node involvement, advanced stage) or patients' survival [6,7], and the results about its biological role on cancer cell motility and invasiveness [11]. In our meta-analysis, we found that subjects carrying Arg 388 were associated with higher risk of developing and progressing prostate cancer than those with the wild-type allele, which confirmed the hypothesis described above. Some limitations of this meta-analysis should be acknowledged. First of all, the control populations were not uniform. Healthy populations as well as non-cancer patients like BPH patients were included. Some individuals in the control group are likely to develop cancer in subsequent years though they had no clinical symptoms at the time of investigation. Misclassification bias results in deviation of genotype distribution in the controls. Second, prostate cancer, as a complex disease, was considered as the result of combined effects of multifactors, including inherited and environmental factors [23], however, no such data was observed in previous studies. Thus, our result was only based on unadjusted estimates. Lacking of the information for the data analysis may cause serious confounding bias. Third, the effect Figure 1 Forest plot of prostate cancer risk associated with FGFR4 Gly 388 Arg polymorphism (Arg allele vs. Gly allele). The squares and horizontal lines correspond to the study-specific OR and 95% CI. The area of the squares reflects the weight (inverse of the variance). The diamond represents the summary OR and 95% CI. Figure 2 Forest plot of prostate cancer progression associated with FGFR4 Gly 388 Arg polymorphism (Arg/Arg vs. Gly/Gly+Gly/ Arg). The squares and horizontal lines correspond to the studyspecific OR and 95% CI. The area of the squares reflects the weight (inverse of the variance). The diamond represents the summary OR and 95% CI. of the polymorphism was relatively trivial with small ORs. We need further studies with larger number participants to confirm the effect.
Our meta-analysis also had some advantages. First, disease progression status as tumor stage was taken into account in present study. Second, data in present study were pooled from different studies, which significantly increased statistical power of the analysis. Third, the quality of studies included in our meta-analysis was satisfactory and cruelly met our inclusion criterion. Fourth, the distribution of genotypes in the controls was consistent with Hardy-Weinberg equilibrium in all studies. We further performed sensitivity analysis to detect the stability of the meta-analysis, and the results did not alter the pattern of association and revealed that the risk effect of Arg 388 was stable. In addition, publication bias was not detected in present study, indicating that our findings seemed not to be due to biased publications.

Conclusions
Our meta-analysis showed the evidence that FGFR4 Arg 388 allele was associated with an increased risk of prostate cancer development and progression, suggesting that FGFR4 Gly 388 Arg polymorphism could be a marker for prostate cancer development and progression. Based on the limitations of present study list above, further prospective researches using standardized unbiased methods, and larger numbers of worldwide participants are expected to examine the association to confirm our results, and other possible confounding risk factors like age, life style, and familial history should also be controlled when it was assessed. Moreover, gene-gene and gene-environment interactions should also be considered.