Meta-analysis:Abnormal β-catenin Expression In Gastric Cancer In Asia Population-a Prognostic Factor

Background and objectiveGastric cancer (GC) is the fourth most common cancer globally, and was the second most common cause of death due to malignancies worldwide. Especially in Asia,the incidence of GC is high. Incidence of gastric cancer in our country ranked after the lung cancer in the second place, the gastric cancer in China accounts for about 40 percent of global new cases. Multiple environmental factors including chronic Helicobacter pylori (H. pylori) infection, hereditary factors and dietary factors have been implicated in the initiation of gastric carcinogenesis. Although the understanding of neoplastic progression has improved in the past decade, the prognosis of patients with advanced GC remains relatively poor. The invasiveness and easy metastasis of cancer is the cause of poor prognosis of patients with gastric cancer. Consequently, it is important to uncover the biological mechanisms underlying progression of the disease and develop strategies to intervene in this process.Epithelial-mesenchymal transition (EMT), a developmental process in which epithelial cells show reduced intercellular adhesion and acquire migratory fibroblastoid properties, is considered to be critical for invasive and metastatic progression in cancer. EMT is associated with the down-regulation of epithelial markers, aberrant upregulation of mesenchymal markers and abnormal translocation of β-catenin in several human cancers including GC. As a central molecule in the int/Wingless family (Wnt) signaling pathway, P-catenin expression is localized in the membrane, cytoplasm and nucleus.β-catenin has a dual role depending on its intracellular localization. Membranous expression of β-catenin is linked to E-cadherin and the actin cytoskeleton, and is responsible for cell-to-cell adhesion and exerting a restrictive effect on tumor growth. Cytoplasmic and nuclear β-catenin are mainly involved in regulation of the Wnt signaling pathway. When a Wnt ligand engages with its receptors, the scaffold protein Axin translocates to the transmembranous receptor complex and inhibits the destruction complex. Thus, cytoplasmic P-catenin escapes degradation, accumulates in the cytoplasm and finally translocates to the nucleus. After localizing to the nucleus, β-catenin activates a target gene expression program with loss of E-cadherin, linking EMT to Wnt signaling. In addition, Wnt signaling is also connected to EMT by activation of snail2 and ZEB1 via other Wnt target genes. Taken together, these findings suggest that different forms of β-catenin contribute to a feed-forward loop in invading cancer cells.Deregulated β-catenin is involved in the development of multiple tumors. In the presence of a Wnt signal, activation of P-catenin is found in about 30% of GC patients. Moreover, potential biological mechanisms have been proposed, such as the CagA+ strain of H. pylori which is thought to activate P-catenin to promote intestinal transdifferentiation in gastric epithelial cells, and accumulating evidence indicates that inflammation induced by COX-2/PGE2 and the Wnt pathway plays a critical role in GC development. These findings provide evidence that β-catenin is important in the development and progression of GC and may be significantly associated with prognosis in patients with GC. When evaluating the expression of variant localization, controversial results have been observed. The morbidity and mortality of GS is high in Asia, especially in China, to reveal the relationship between β-catenin and GS in Asian population is important. Thus, this meta-analysis was performed to evaluate the prognostic significance of β-catenin immunohistochemical expression in patients with GC.MATERIALS AND METHODSLiterature searchWe searched the Medline(PubMed),Embase,Web of Science,Cochrane Library,Vip,Wanfang and Cnki databases using the following terms and all possible combinations:"β-catenin", "beta-catenin", "Wnt", "Gastric Neoplasms", "Gastric Cancer", "Gastric Carcinoma", "Gastric Tumor", "Stomach Cancer", "Stomach Carcinoma", "Stomach Neoplasms", "Stomach Tumor", "GC" and "Prognosis." The search ended on January,2015, and no lower date limit was used. The citation lists associated with the studies were used to identify additional eligible studies. The reviews and bibliographies were also manually inspected to find related articles.Inclusion and exclusion criteriaThe studies were included in our meta-analysis if they met the following inclusion criteria:(1) Evaluation of the relationships between β-catenin expression and overall survival (OS) or clinicopathological features of GC in Asians;(2) The specimens derived from GC patients;(3) β-catenin expression examined by immunohistochemistry;(4) Publications in English and Chinese;(5) Sufficient information provided to estimate the HR and its 95%CI, and information to estimate the OR in the analysis of clinicopathological features. The following articles were excluded:(1) non-English and non-Chinese language articles;(2) letters, case reports, reviews, and conference abstracts without original data;(3) articles from which the relevant data could not be extracted; and overlapping articles or those with duplicate data;(4) the definition of abnormal β-catenin expression was unclear.Data extraction and assessmentThe database recorded the most relevant data comprising author’s name, year of publication, antibody source, definition of β-catenin expression, study location, number of patients and tumor characteristics. Study quality was assessed according to the Newcastle-Ottawa quality assessment scale. The score assessed eight items of methodology, and grouped them into three major classifications:selection, comparability and outcome. A maximum score of 1 was graded for each item, except that related to comparability, which allowed for 2. For quality, scores ranged from 0 (lowest) to 9 (highest), and studies with more than 5 points were rated as qualified.Statistical analysisThe impact of β-catenin expression on OS was measured by HR for quantitative aggregation. The most accurate approach was to obtain the HR estimate and 95%CI directly from the paper, or calculate them using the parameters offered in the manuscript. Otherwise, Kaplan-Meier curves were read by Engauge Digitizer version 4.1. Heterogeneity across studies was evaluated using a x2-based Q statistical test. The I2 statistic was also calculated for study heterogeneity. I2> 50 and P≤ 0.10 indicated a lack of heterogeneity among the studies. For studies with P> 0.10, the pooled OR and HR estimates of each study were calculated by the fixed-effects model. For studies with P≤ 0.10, the random-effects model was used. For the pooled analysis of the correlation between β-catenin expression and clinicopathological features [Lauren classification, tumor-node-metastasis (TNM) stage, the depth of invasion, lymph node metastasis, distant metastasis, grade of differentiation and vascular invasion], ORs and their 95%CIs were combined to estimate the effect.Sensitivity analysis was also conducted by sequential omission of individual studies to evaluate stability of the results. In addition, publication bias was assessed using funnel plots. HRs and ORs and their variance were calculated and pooled using the RevMan systematic review and meta-analysis software package (Review Manager Version 5.2).RESULTSAfter screening the abstract and full-text of the included articles,40 articles were finally included in the quantitative analysis of the prognostic value of β-catenin expression in GC and the relationship between β-catenin expression and clinicopathological features. The quality of the including 40 articles were assessed as high.In general, aberrant β-catenin expression, which consisted of positive cytoplasmic and nuclear expression and negative membranous expression in GC, demonstrated a significant increase in mortality risk as compared to regular β-catenin expression (combined HR= 1.83,95%CI:1.21～2.78). However, a significant degree of heterogeneity was observed (I2= 83%, P< 0.00001). We performed a subgroup analysis using the definition of β-catenin expression. β-catenin abnormal expression in membranous, nuclear and cytoplasm was associated with poor prognosis (HR=2.98, 95%CI:1.52～5.84). No significant relationship between OS of GC and the other two patterns of nuclear accumulation(HR=1.39,95%CI:0.82～2.36) and loss of membranous expression(HR=1.31,95%CI:0.23～7.41). Significant heterogeneity was observed(I2= 77%～93%).We evaluated the correlation between β-catenin immunohistochemical expression and the clinicopathological characteristics of GC. β-catenin expression was significantly associated with the size of GC(>5cm vs<5cm:OR=1.31, 95%CI:1.00～1.71). Significant heterogeneity was not observed(I2= 0%). The subgroup analysis showed that loss of membranous expression of β-catenin, the nuclear and cytoplasm accumulation of β-catenin and loss of membranous expression and cytoplasm accumulation of β-catenin had no significant association with GC size. Significant heterogeneity was not observed in the subgroup.P-catenin expression was significantly associated with Lauren classification(intestinal-type vs diffuse-type:OR= 0.40,95%CI:0.23～0.71). Significant heterogeneity was observed(I2= 65%).The subgroup analysis showed that the nuclear accumulation of β-catenin had no significant association with Lauren classification(OR=1.36,95%CI:0.59-3.14). β-catenin abnormal expression in membranous, nuclear and cytoplasm(OR=0.36,95%CI:0.19～0.69), the and cytoplasm accumulation of β-catenin(OR=0.16,95%CI:0.06～0.47) and loss of membranous expression of β-catenin(OR=0.20,95%CI:0.10～0.44) was significantly associated with Lauren classification. Significant heterogeneity was not observed in the subgroup(I2=0%～48%).β-catenin expression was significantly associated with grade of differentiation(G3/G4 vs G1/G2:OR=3.29,95% CI:2.67～4.06). Significant heterogeneity was not observed(I2=31%). The subgroup analysis showed that the nuclear accumulation of β-catenin(OR=2.16,95%CI:0.69～6.72) and loss of membranous expression of β-catenin(OR=1.79,95%CI:0.79～4.02) had no significant association with grade of differentiation. P-catenin abnormal expression in membranous, nuclear and cytoplasm(OR=3.80,95%CI:2.68～5.40), the nuclear and cytoplasm accumulation of β-catenin(OR=2.67,95%CI:1.74～4.08) and loss of membranous expression and cytoplasm accumulation of β-catenin(OR=3.93, 95%CI:2.66～5.81) was significantly associated with grade of differentiation. Significant heterogeneity was not observed(I2= 0%～48%).β-catenin expression was significantly associated with lymph node metastasis (positive vs negative:OR=2.98,95%CI:2.24～3.96). Significant heterogeneity was observed(I2= 53%). The subgroup analysis showed that the nuclear accumulation of β-catenin(OR=1.75,95%CI:0.35～8.87), loss of membranous expression of β-catenin(OR=2.28,95%CI:0.60～8.68) and the nuclear and cytoplasm accumulation of β-catenin (OR=1.89,95%CI:0.74～4.87) had no significant association with lymph node metastasis. Loss of membranous expression, nuclear and cytoplasm accumulation of β-catenin(OR=2.91,95%CI:2.15～3.94) and the and cytoplasm accumulation of p-catenin(OR=2.52,95%CI:1.12～5.64) and loss of membranous expression and cytoplasm accumulation of β-catenin(OR=5.58,95%CI:2.76-11.29) was significantly associated with lymph node metastasis. Significant heterogeneity was not observed(I2= 0%-48%).β-catenin expression was significantly associated with distant metastasis (positive vs negative:OR=4.07,95%CI:2.48～6.68). Significant heterogeneity was not observed(I2= 0%). The subgroup analysis showed that P-catenin abnormal expression in membranous, nuclear and cytoplasm(OR=7.11,95%CI:2.53～20.01), loss of membranous expression and cytoplasm accumulation of β-catenin(OR=4.41, 95%CI:1.99～9.78) and the and cytoplasm accumulation of β-catenin(OR=2.38, 95%CI:1.04～5.44) Significant heterogeneity was not observed(I2= 0%～0%).β-catenin expression was significantly associated with the depth of invasion(G3-G4 vs G1-G2:OR=2.43,95%CI:1.78～3.31). Significant heterogeneity was observed(I2= 58%). The subgroup analysis showed that the nuclear accumulation of β-catenin(OR=1.61,95%CI:0.31～8.40) and loss of membranous expression of β-catenin had no significant association with the depth of invasion. β-catenin abnormal expression in membranous, nuclear and cytoplasm(OR=2.43, 95%CI:1.50～3.94), the and cytoplasm accumulation of β-catenin(OR=2.91, 95%CI:1.37～6.18), the nuclear and cytoplasm accumulation of β-catenin(OR=2.26, 95%CI:1.22～4.20) and loss of membranous expression and cytoplasm accumulation of β-catenin(OR=2.56,95%CI:1.23～5.33) was significantly associated with the depth of invasion. Significant heterogeneity was observed(I2= 51%～72%).β-catenin expression was significantly associated with TNM stage(T3-T4 vs T1-T2:OR=2.78,95%CI:1.93～4.00). Significant heterogeneity was observed(I2 =61%).The subgroup analysis showed that the nuclear accumulation of P-catenin(OR=1.49,95%CI:0.45～4.96), loss of membranous expression of β-catenin(OR=1.98,95%CI:0.74～5.35) and the nuclear and cytoplasm accumulation of β-catenin(OR=1.59,95%CI:0.60-4.24) had no significant association with TNM stage. β-catenin abnormal expression in membranous, nuclear and cytoplasm(OR=3.25,95%CI:2.03-5.19) and loss of membranous expression and cytoplasm accumulation of β-catenin(OR=5.92,95%CI:3.30～10.63) was significantly associated with TNM stage. Significant heterogeneity was not observed(I2= 0%-47%).In the meta-analysis of clinicopathological parameters, we found that data from Sun et al created a significant bias in the pooled ORs of TNM stage, the depth of invasion and lymph node metastasis, as well as increased unexpected inter-study heterogeneity. Moreover, Shun’s study of early GC was considered unsuitable for the analysis of lymph node metastasis. The funnel plots of publication bias did not exhibit significant asymmetry in the analysis of OS and histopathological features.DiscussionIn recent years, β-catenin and its associate E-cadherin were proved to be not only static components of adherens junctions, but important mediators of downstream Wnt signaling cascades. As described above, the different localization of β-catenin contributes to cell-cell adhesion or transcriptional activation of responsive target genes. GSK3β, APC, and axin are negative regulators of the Wnt pathway and are functionally assembled in the destruction complex, where β-catenin can be efficiently regulated. This leads to subsequent over expression of free cytoplasmic P-catenin and exerts a nuclear function without control of tumorigenesis and progression.This meta-analysis aimed to examine the association between P-catenin expression, OS and clinicopathological characteristics of GC. We assessed the outcomes of patients with GC in 10 studies, and found that P-catenin expression significantly predicted poor OS in GC patients.Subgroup analysis revealed that the combined abnormal pattern of β-catenin influenced the survival of GC patients. Loss of membranous expression and nuclear accumulation of β-catenin was associated with poor prognosis. Our result revealed that β-catenin immunohistochemical expression was associated with poor OS.When evaluating the OR in histopathology, significant correlations were observed between abnormal β-catenin expression and clinicopathological features including Lauren classification, lymph node metastasis, distant metastasis, grade of differentiation, GC size, TNM stage and the depth of invasion. These results suggested that β-catenin may predict metastasis in patients with advanced GC. Subgroup analysis showed that the OR of combined abnormal pattern of P-catenin was similar to the summational OR of all abnormal pattern. The combined abnormal pattern of β-catenin expression may predict metastasis in patients with advanced GC.A previous meta-analysis indicated that decreased E-cadherin expression was related to poor OS of gastric carcinoma, as well as being significantly correlated with differentiation, invasion and metastasis. The studies included in our meta-analysis also showed that a decrease in E-cadherin or abnormal β-catenin or both weakened cell-cell adhesion and resulted in cell spread, allowing cancer cell infiltration and metastasis. Taking these findings into account, we suggest that abnormalβ-catenin expression represents an independent risk factor together with E-cadherin expression for the occurrence and development of GC. However, the role of different β-catenin location in gastric tumor biology remains unclear and should be investigated in future studies.Our analysis indicated that abnormal expression of P-catenin associated with tumor invasion and metastasis, may be able to become a targeted therapy based on the intervention Wnt/β-catenin signaling pathway in gastric cancer. But the Wnt/β-catenin signaling pathway was associated with lots of normal function in human, so the direction of future research should continue to explore the mechanism of GC and to find out specific Wnt/β-catenin signaling pathway marker molecules of GS.We found highly significant heterogeneity between the included studies. In the stratified analysis of OS, heterogeneity was not significant in certain subgroup analyses. These findings suggest that the definition of β-catenin account for this heterogeneity. Sensitivity analysis showed that the OR from Sun et al created unexpected heterogeneity. Shun’s study on early GC also contributed to heterogeneity in the estimate of OR for lymph node metastasis. A potential source of bias may be associated with the approach of extrapolating HRs. If HRs were not directly reported in the studies, we calculated them from the data provided in the papers and from the survival curves, assuming that censored observations were identically distributed. This approach did not completely eliminate inaccuracy during extraction of the survival rates. The estimated HR might thus be less reliable than when obtained directly from published statistics.ConclusionThis meta-analysis revealed that β-catenin immunohistochemical expression was associated with poor OS and histopathological features such as Lauren classification, lymph node metastasis, distant metastasis, grade of differentiation, GC size, TNM stage and the depth of invasion in patients with GC in Asians. Abnormal β-catenin expression may be a predictive factor of poor prognosis in GC patients, and might predict invasion and metastasis.