Qu K, Liu SS, Wang ZX, Huang ZC, Liu SN, Chang HL, Xu XS, Lin T, Dong YF, Liu C. Polymorphisms of glutathione S-transferase genes and survival of resected hepatocellular carcinoma patients. World J Gastroenterol 2015; 21(14): 4310-4322 [PMID: 25892883 DOI: 10.3748/wjg.v21.i14.4310]
Corresponding Author of This Article
Chang Liu, MD, PhD, Department of Hepatobiliary Surgery, the First Affiliated Hospital of Medical College, Xi’an Jiaotong University, No. 277 Yanta West Road, Xi’an 710061, Shaanxi Province, China. liuchangdoctor@163.com
Research Domain of This Article
Gastroenterology & Hepatology
Article-Type of This Article
Prospective Study
Open-Access Policy of This Article
This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Kai Qu, Su-Shun Liu, Zhi-Xin Wang, Zi-Chao Huang, Si-Nan Liu, Hu-Lin Chang, Xin-Sen Xu, Ting Lin, Chang Liu, Department of Hepatobiliary Surgery, The First Affiliated Hospital of Medical College, Xi’an Jiaotong University, Xi’an 710061, Shaanxi Province, China
Ya-Feng Dong, Department of Obstetrics and Gynecology, University of Kansas School of Medicine, Kansas City, KS 66160, United States
ORCID number: $[AuthorORCIDs]
Author contributions: Qu K and Liu C designed the research and wrote this paper; Liu SS, Wang ZX and Huang ZC performed the statistical analysis; Liu SN, Chang HL and Xu XS were involved in collecting patient data; Qu K and Lin T took part in literature searches; Dong YF edited the manuscript.
Supported by Grants from National Natural Science Foundation of China, No. 81201549 and No. 81272644; and the Project of Innovative Research Team for Key Science and Technology in Xi’an Jiaotong University, No. 2003KCJ-23.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Chang Liu, MD, PhD, Department of Hepatobiliary Surgery, the First Affiliated Hospital of Medical College, Xi’an Jiaotong University, No. 277 Yanta West Road, Xi’an 710061, Shaanxi Province, China. liuchangdoctor@163.com
Telephone: +86-29-82654746 Fax: +86-29-82654746
Received: July 24, 2014 Peer-review started: July 24, 2014 First decision: August 10, 2014 Revised: August 19, 2014 Accepted: October 21, 2014 Article in press: October 21, 2014 Published online: April 14, 2015 Processing time: 265 Days and 7.2 Hours
Abstract
AIM: To investigate the effects of single nucleotide polymorphisms (SNPs) in glutathione S-transferase (GST) genes on survival of hepatocellular carcinoma (HCC) patients.
METHODS: Twelve tagging SNPs in GST genes (including GSTA1, GSTA4, GSTM2, GSTM3, GSTO1, GSTO2 and GSTP1) were genotyped using Sequenom MassARRAY iPLEX genotyping method in a cohort of 214 Chinese patients with resected HCC. The Cox proportional hazards model and log-rank test were performed to determine the SNPs related to outcome. Additionally, stratified analysis was performed at each level of the demographic and clinical variables. An SNP-gene expression association model was further established to investigate the correlation between SNP and gene expression.
RESULTS: Two SNPs (GSTO2: rs7085725 and GSTP1: rs4147581) were significantly associated with overall survival in HCC patients (P = 0.035 and 0.042, respectively). In stratified analysis, they were more significantly associated with overall survival in patients with younger age, male gender and cirrhosis. We further investigated cumulative effects of these two SNPs on overall survival in HCC patients. Compared with the patients carrying no unfavorable genotypes, those carrying 2 unfavorable genotypes had a 1.70-fold increased risk of death (P < 0.001). The cumulative effects were more significant in those patients with younger age, male gender and cirrhosis (HR = 2.00, 1.94 and 1.97, respectively; all P < 0.001). Additionally, we found that heavy smoking resulted in a significantly worse overall survival in those patients carrying variant alleles of rs7085725 (HR = 2.07, 95%CI: 1.13-3.76, P = 0.018). The distributions of GSTO2: rs7085725 and GSTP1: rs4147581 genotypes were associated with altered gene expression and contributed to influences on overall survival.
CONCLUSION: Our study provides the first evidence that GSTO2 and GSTP1 gene polymorphisms may serve as independent prognostic markers for HCC patients.
Core tip: To determine the prognostic value of single nucleotide polymorphisms (SNPs) in GST genes in hepatocellular carcinoma (HCC) after liver resection, we analyzed the association between 12 tagging SNPs in GST genes and outcome of 214 HCC patients who underwent liver resection. Our data showed two SNPs (GSTO2: rs7085725 and GSTP1: rs4147581) to be significantly associated with overall survival in HCC patients. These two SNPs used in combination were more powerful prognostic markers for HCC patients. Our study provides the first evidence that GSTO2 and GSTP1 gene polymorphisms may serve as independent prognostic markers for HCC patients.
Citation: Qu K, Liu SS, Wang ZX, Huang ZC, Liu SN, Chang HL, Xu XS, Lin T, Dong YF, Liu C. Polymorphisms of glutathione S-transferase genes and survival of resected hepatocellular carcinoma patients. World J Gastroenterol 2015; 21(14): 4310-4322
Glutathione S-transferases (GSTs) are a broadly expressed family of phase II isoenzymes that protect normal cells against damage induced by hepatitis virus or aflatoxin-related hepatocarcinogen[1]. Recently, emerging evidence indicates that the detoxifying properties of the GSTs are genetically determined[2]. In human, several classes of GST genes exist: α (GSTA), μ (GSTM), π (GSTP), θ (GSTT), and ω (GSTO) with one or more genes in each class[3,4]. Altered expression of these GST genes has been suggested to increase the risk of cancer[4-6]. Polymorphisms in these genes have been shown to produce significant alterations in the metabolism of many substrates, including carcinogens and chemotherapeutic agents[7]. As a result, these polymorphisms also have been suggested to be associated with the risk of many malignances[2,8-16]. Some of them have also been suggested to affect the survival status of cancer patients[8-10,14].
Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and the third most common cause of cancer mortality, resulting in about 500000 deaths per year. Half of HCC cases and deaths were estimated to occur in China[17]. Most of these cases in China arise due to the chronic infection with hepatitis virus, the common carcinogen to HCC[18]. Not only hepatitis virus infection, but also other environmental carcinogens, such as aflatoxin or chemical carcinogens, would contribute to hepatic carcinogenesis[19]. The metabolizing enzymes for detoxification or activation of these carcinogens to lower toxicity, potentially affect the individual cancer risk and progression of HCC. Evidence has shown that some SNPs of GSTs were associated with HCC risk[12]. However, none of these polymorphisms are investigated in relation to HCC survival.
In this study, we assessed the effects of 12 tagging SNPs in seven GST genes GSTA1, GSTA4, GSTM2, GSTM3, GSTO1, GSTO2 and GSTP1 on 5-year survival in a cohort of 214 Chinese HCC patients receiving surgical treatment. To the best of our knowledge, this is the first study to investigate the predictive value of SNPs in these GST genes for HCC prognosis in Chinese population.
MATERIALS AND METHODS
Study population
A total of 214 Han Chinese HCC patients were recruited at the Department of Hepatobiliary Surgery, the First Affiliated Hospital of Xi’an Jiaotong University in Xi’an, China. These patients were recruited between January 2009 and October 2013. HCC diagnosis was based on the National Comprehensive Cancer Network (NCCN) clinical practice guidelines in oncology. All subjects included in this study had histopathologically confirmed HCC. All cases received surgical treatment and had no previous history of other cancers or cancer-related treatments. There were no recruitment restrictions on age at diagnosis, gender and tumor stage. Participants received other treatments (such as chemotherapy, interventional therapy or biological target therapy) after surgical operation were excluded for further analysis. Ethics committee of the First Affiliated Hospital of Xi’an Jiaotong University approved this study. Written informed consent was obtained from each participant before enrollment.
Another 86 HCC patients derived from Queen Mary Hospital, the University of Hong Kong, were recruited as a validated cohort (Hong Kong Cohort). All genetic and SNP data were obtained from two microarray series (GSE28127 and GSE22058) in the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database (http://www.ncbi.nlm.nih.gov/geo/).
Demographic and clinical data
Detailed demographic and clinical information was collected through in-person interview, medical chart review, or consultation with treating physicians. The demographic data included the date of diagnosis, smoking behavior, liver cirrhosis, Child-Pugh score, serum alpha-fetoprotein (AFP), and tumor-node-metastasis (TNM) stage. To calculate pack-years of smoking, the average of number of cigarettes smoked per day was divided by 20 to give packs per day and multiplied by the total number of years of smoking. At 3-mo intervals, follow-up information on death of patients was updated by a trained clinical specialist through on-site interview, direct calling or medical chart review. The latest follow-up data in this analysis were obtained in January 2014. Demographic and clinical characteristics of HCC patients are shown in Table 1.
Table 1 Demographic and clinical characteristics of hepatocellular carcinoma patients.
Variable
Our cohort
Hong Kong cohort
n
%
n
%
Total
214
100
401
100
Age (yr)
≤ 55
108
50.5
NA
NA
> 55
106
49.5
NA
NA
Gender
Male
176
82.2
322
80.3
Female
38
17.8
79
19.7
Smoking behavior
Yes
70
32.7
223
55.6
No
144
67.3
178
44.4
Cirrhosis
Present
126
58.9
229
57.1
Absent
88
41.1
172
42.9
Child-Pugh score
A
172
80.4
390
97.3
B
42
19.6
11
2.7
AFP level (ng/mL)
< 200
100
46.7
NA
NA
≥ 200
114
53.3
NA
NA
Tumor size
< 5 cm
102
47.7
NA
NA
≥ 5 cm
112
52.3
NA
NA
TNM stage
I + II
119
55.6
177
44.1
III
95
44.4
224
55.9
Outcome
Dead
111
51.9
272
67.8
Alive
103
48.1
129
32.2
Tagging SNP selection and genotyping
Tagging SNP selection was based on data from the NIEHS (National Institute of Environmental Health Sciences, http://snpinfo.niehs.nih.gov/snpinfo/snptag.htm). Together with SNPs within a gene, we also considered polymorphisms in the 5’- and 3’-flanking regions up to 5 kb. We chose common SNPs with a minor allele frequency (MAF) ≥ 0.05 in a population of Chinese Han in Beijing (CHB) and an SNP design score cutoff ≥ 0.6 (indicating a higher probability for a successful assay). To select tagging SNPs, we used a pairwise tagging approach with r2≥ 0.8. Finally, a total of 12 potentially tagging SNPs in 7 genes were selected, including GSTA1: rs6917150; GSTA4: rs385636; GSTM2: rs638820; GSTM3: rs1109138, rs7483; GSTO1: rs2282326, rs17116779; GSTO2: rs156699, rs7085725, rs157077; GSTP1: rs4147581, rs2370141. Information of selected tagging SNPs is shown in Table 2 and Figure 1.
Figure 1 Relevant pictures of polymorphisms in seven GST genes.
A: GSTA1 (Gene ID: 2946); B: GSTM2 (Gene ID: 2946); C: GSTA4 (Gene ID: 2941); D: GSTM3 (Gene ID: 2947); E: GSTO1 (Gene ID: 9446); F: GSTO2 (Gene ID: 119391); G: GSTP1 (Gene ID: 2950). Genotype data were obtained from HapMap database (http://hapmap.ncbi.nlm.nih.gov) and SNPs were sorted by LD similarity using NIEHS tools (http://snpinfo.niehs.nih.gov). Twelve selected SNPs were labeled by red block.
Table 2 Summarized information of selected tagging SNPs in GST genes.
Gene
Tagging SNPs
bp position
Ref SNP alleles
MAF_CHB
GSTA1
rs6917150
Chr 6: 52760414
T/C
0.338
GSTA4
rs385636
Chr 6: 52948896
A/G
0.174
GSTM2
rs638820
Chr 1: 110011429
T/C
0.344
GSTM3
rs1109138
Chr 1: 110079472
G/A
0.167
rs7483
Chr 1: 110081224
A/G
0.244
GSTO1
rs2282326
Chr 10: 106010388
A/C
0.244
rs17116779
Chr 10: 106021185
G/A
0.078
GSTO2
rs156699
Chr 10: 106043631
T/C
0.222
rs7085725
Chr 10: 106050199
T/C
0.122
rs157077
Chr 10: 106027884
A/G
0.378
GSTP1
rs4147581
Chr 11: 67108161
G/C
0.273
rs2370141
Chr 11: 67105105
A/G
0.181
Genomic DNA extraction and genotyping
Genomic DNA was extracted from peripheral blood samples using E.Z.N.A. Blood DNA Midi Kit (Omega Bio-Tek, Norcross, GA, United States) according to the manufacture’s instructions. Genotyping was performed using Sequenom MassARRAY iPLEX genotyping system (Sequenom Inc, San Diego, CA, United States). Laboratory personnel conducting genotyping were blinded to patients’ information.
Statistical analysis
All statistical analyses were performed using the SPSS 21.0 statistical package for Microsoft Windows (SPSS, Chicago, IL, United States). The survival time was defined as the period from the date of first treatment to the date of death or last follow-up. Hazard ratios (HRs) and 95%CIs were estimated from a Cox proportional hazards model. Kaplan-Meier curve and log-rank test were also used to assess the differences of overall survival. All statistical tests were two-sided and P < 0.05 was considered statistically significant.
RESULTS
Characteristics of the study population
For the 214 patients included, the median age at the time of HCC diagnosis was 55 years (range: 20-84 years). The majority of patients (82.2%, 176/214) were men and 32.7% (70/214) of all patients had smoking behavior. About 58.9% (126/214) of patients had cirrhosis and 80.4% (172/214) had Child-Pugh score A. There were 52.3% (112) of patients with tumor size ≥ of 5 cm and 53.3% (114) with significantly increased serum AFP (200 ng/mL). The percentages of patients with TNM stage I/II or stage III disease were 55.6% (119/214) and 44.4% (95/214), respectively. The median follow-up time was 24 mo (ranging from 3 to 60 mo). By the last follow-up, 51.9% (111/214) of patients were dead, and the median survival time for the entire cohort of patients was 33.6 mo.
Association between individual SNPs and overall survival in HCC patients
We assessed the effect of 14 tagging SNPs in seven GST genes on death of HCC patients using the Cox regression model. When the dominant genetic model was tested, our data showed that SNPs rs7085725 in GSTO2 and rs4147581 in GSTP1 gene were significantly associated with the overall survival of HCC patients (Table 3). Patients carrying at least one variant allele of rs7085725 (WV + VV genotypes) had a significantly increased risk of death (HR = 1.60; 95%CI: 1.03-2.47), meanwhile, those carrying WV + VV genotypes of rs4147581 had a decreased risk of death (HR = 0.68; 95%CI: 0.46-0.99), when compared with those carrying homozygous wild-type alleles (WW genotype) (P = 0.035 and 0.042, respectively). Kaplan-Meier analysis showed a significantly shorter median survival time in patients with WV + VV genotypes of SNP rs7085725 than those with WW genotype (19.8 mo vs 36.6 mo; log rank P = 0.03) (Figure 2A). In contrast, patients with WV + VV genotypes of SNP rs4147581 had a longer median survival time than those with WW genotype (33.9 mo vs 26.8 mo; log rank P = 0.04) (Figure 2B).
Significant association of rs7085725 and rs4147581 with overall survival in HCC patients
We further analyzed the effect of SNPs rs7085725 and rs4147581 on the overall survival in HCC patients stratified by demographic and clinicopathological characteristics. As shown in Table 4, the significantly increased death risk conferred by SNP rs7085725 was observed in younger patients [HR (95%CI) = 1.86 (1.02-3.38); P = 0.044], male patients [HR (95%CI) = 1.83 (1.14-2.94); P = 0.013], smoking patients [HR (95%CI) = 2.59 (1.27-5.27); P = 0.09], and those with cirrhosis [HR (95%CI) = 2.00 (1.11-3.59); P = 0.020], Child-Pugh score A [HR (95%CI:) = 2.28 (1.32-3.93); P = 0.003] and AFP ≥ 200 ng/mL [HR (95%CI) = 2.40 (1.34-4.29); P = 0.003]. Meanwhile, the significantly decreased death risk conferred by SNP rs4147581 was observed in younger patients [HR (95%CI) = 0.52 (0.30-0.92), P = 0.024], male patients [HR (95%CI) = 0.64 (0.42-0.99), P = 0.042], non-smoking patients [HR (95%CI) = 0.58 (0.36-0.93); P = 0.024], and those with cirrhosis [HR (95%CI) = 0.50 (0.29-0.85); P = 0.011], Child-Pugh score B [HR (95%CI) = 0.45 (0.22-0.90); P = 0.024], AFP < 200 ng/mL [HR (95%CI) = 0.51 (0.29-0.89); P = 0.018] and tumor size ≥ 5 cm [HR (95%CI) = 0.60 (0.37-1.00); P = 0.048]. Interestingly, both of the two SNPs were significantly associated with overall survival in patients with younger age, male gender and cirrhosis. Log-rank test further indicated significant differences in overall survival between WV + VV and WW genotypes of SNPs rs7085725 or rs4147581 in those patients with above-mentioned factors (Figure 3).
Figure 3 Kaplan-Meier curves for hepatocellular carcinoma patients carrying genetic variants of GSTO2: rs7085725 and GSTP1: rs4147581 in stratified subgroups.
A and B: In patients with age < 55 years; C and D: In patients with male gender; E and F: In patients with cirrhosis. WW: Homozygous wild-type genotype, WV: Heterozygous genotype; VV: Homozygous variant genotype.
Table 4 Stratified analysis on the association between GSTO2: rs7085725 and GSTP1: rs4147581 genotype and overall survival of hepatocellular carcinoma patients.
Cumulative effect of unfavorable genotypes on overall survival
To further assess the cumulative effects of genetic variants of SNPs rs7085725 and rs4147581 on overall survival in HCC, we did a joint analysis by including the two SNPs (Table 5). The unfavorable genotypes were defined as WV or VV for rs7085725 and WW for rs4147581. When using group 1 (with 0 unfavorable genotype) as reference, HCC patients in group 3 (with two unfavorable genotypes) had a 1.70-fold increased risk of death (95%CI: 1.30-2.22; P < 0.001), when compared with those in the reference group. A significant dose-response trend was observed (P for trend = 0.004). Furthermore, the death risk of carriers of two unfavorable genotypes was significantly increased among HCC patients with younger age, male gender or cirrhosis, when compared with the reference group (HR = 2.00, 1.94 and 1.97, respectively; P < 0.001). Kaplan-Meier analysis showed that there was significantly decreased overall survival in patients carrying two unfavorable genotypes, when compared with others, either in all patients or in stratified subgroups (Figure 4).
Figure 4 Kaplan-Meier curves for hepatocellular carcinoma patients carrying unfavorable genotypes.
A: In all patients; B: In patients with age < 55 years; C: In patients with male gender; D: In patients with cirrhosis. Patients were classified into two groups, according to number of unfavorable genotypes carried. Group 1 + 2 carried 0 or 1 unfavorable genotype, while Group 3 carried 2 unfavorable genotypes. Unfavorable genotypes were GSTO2: rs7085725 (WV or VV) and GSTP1: rs4147581 (WW).
Table 5 Cumulative effects of unfavorable genotypes on overall survival of hepatocellular carcinoma patients.
Modulating effects of smoking on overall survival in HCC patients by rs7085725
Previous report from our center showed that cigarette smoking was associated with an increased risk of death in HCC[20]. As a metabolizing enzyme, GSTP1 has been extensively investigated in the activation and detoxification of pro-carcinogens in tobacco smoke[11,21]. In line with this evidence, we further assessed the modulating effects on the association between smoking and overall patient survival, stratified by SNPs rs7085725 and rs4147581 (Table 6). We observed that the survival time of those patients carrying variant alleles of rs7085725 were adversely affected by smoking exposure [HR (95%CI) = 1.52 (1.02-2.26); P = 0.042]. This significant adverse effect on patient survival conferred by smoking was also observed in smoking quantity-stratified analysis [HR (95%CI) = 2.07 (1.13-3.76); P = 0.018]. Kaplan-Meier analysis further suggested that smoking exposure adversely affected overall survival of variant allele carriers of rs7085725, either in smoking behavior- or in smoking quantity-stratified analysis (Figure 5).
Genotype distributions of rs7085725 and rs4147581 in HCC patients with different gene expression
To further investigate the association between SNP and gene expression, we developed an SNP-gene expression model by integrating genotyping and gene expression data from Hong Kong Cohort. We selected two SNPs rs11191994 and rs614080 to predict genotypes of rs7085725 and rs4147581, respectively. Both of the two SNPs were completely linked with rs7085725 or rs4147581 [linkage disequilibrium (LD): r2 = 1.000, D’ = 1.000]. As shown in Figure 6, there was a significant difference in the genotype distribution of rs7085725 between patients with high and median GSTO2 expression levels (P = 0.042). Additionally, the genotype distributions of rs4147581 in low and median GSTP1 expression groups were also significantly different (P = 0.011). We found that patients carrying WV + VV genotypes of rs7085725 were more frequent in high GSTO2 expression subgroup, while rs4147581 WW allele carriers were dominant in low GSTP1 expression subgroup (Figure 6).
Figure 6 Comparison of genotype distribution in hepatocellular carcinoma patients with different GSTO2 and GSTP1 expression.
A: rs7085725 genotype distribution in different GSTO2 expression groups. rs7085725 genotype was predicted according to rs11191994 (LD r2 = 1.000, D’ = 1.000); B: rs4147581 genotype distribution in different GSTP1 expression groups. rs4147581 genotype was predicted according to rs614080 (LD r2 = 1.000, D’ = 1.000). WV: Heterozygous genotype; VV: Homozygous variant genotype; NS: Not significant.
DISCUSSION
In this study, we evaluated the prognostic value of 12 tagging polymorphisms from seven GST genes in Chinese HCC patients. The most important finding was that SNPs GSTO2: rs7085725 and GSTP1: rs4147581 were significantly associated with the overall survival of HCC patients (P = 0.035 and P = 0.042, respectively). Our data indicated that SNPs GSTO2: rs7085725 and GSTP1: rs4147581 used alone or in combination were potential prognostic markers for HCC patients, especially in those patients with younger age, male gender and cirrhosis. If further validated, the two SNPs may potentially be developed as simple non-invasive predictive biomarkers for HCC patients.
GSTP1, an extensively studied member of GSTs, acts as a part of the protection system against a wide range of potentially harmful cytotoxic compounds. It was observed that altered expression of GSTP1 existed in liver cancer cell lines[22], and in more than 77.8% of HBV-associated HCC tissues[23]. Previous studied have extensively demonstrated that GSTP1 polymorphisms would contribute to the increased risk and poor outcome of many cancers, including esophageal cancer[24], colorectal cancer[8], glioma[10], breast cancer[2,25], prostate cancer[26] and ovarian cancer[9,27]. However, little is known to date about their effect on HCC risk and prognosis. Chen et al[13] reported that GSTP1 gene polymorphisms might be considered as factors increasing the susceptibility to HCC in Taiwanese aged ≤ 57 years. In the present study, we similarly identified a tagging SNP of GSTP1 (rs4147581) as a predictive biomarker in HCC patients aged ≤ 55 years. Besides younger age, we additionally identified that GSTP1: rs4147581 polymorphism was associated with the overall survival in those patients with male gender, non-smoking behavior, liver cirrhosis, Child-Pugh score B, AFP < 200 ng/mL, or tumor size ≥ 5 cm, all of which suggested this SNP as a more powerful predictor than previous SNP markers.
The function of GSTP1: rs4147581 to date remains to be unclear. In the present study, we first reported that genotype distribution of rs4147581 was associated with GSTP1 expression. It has been reported that GSTP1: rs4147581 was associated with DNA methylation[28], which has been clearly demonstrated to play a critical role in down-regulating GSTP1 expression in HCC[6,22,23]. Considering that loss of GSTP1 expression has been suggested to increase the risk of DNA damage[6], we put forward the hypothesis that rs4147581 may control GSTP1 expression by influencing DNA methylation, and consequently induce high risk of carcinogenesis and cancer progression.
GSTO2 gene encodes omega class GST member GSTO2, which exhibits dehydroascorbate (DHA) reductase activity in addition to novel thioltransferase, and monomethylarsonate reductase activities[29,30]. GSTO2 has 70-100 times higher DHA reductase (DHAR) activity than GSTO1, another GST omega class GST member, and is considered to be the most active DHAR in mammalian cells[30]. Previous studies have extensively demonstrated that GSTO2 polymorphisms might contribute to the increased risk of colorectal cancer[31], gastric cancer[32], ovarian cancer[33] and leukemia[34]. This finding suggested the hypothesis that GSTO2 gene polymorphisms might contribute to the increased risk and outcome of HCC. Although Marahatta et al[35] did not find a difference in genotypic distribution for GSTO2 polymorphism between HCC patients and controls, limited number of patients (only 28 cases of HCC and 98 controls) recruited in their study might contribute to this negative result. In our study, we identified that GSTO2: rs7085725 polymorphism was associated with overall survival of HCC patients. When combined with GSTP1: rs4147581, the predictive power of GSTO2: rs7085725 increased, especially in those younger, male and cirrhotic patients.
Recently, emerging evidence has shown that GSTO2 gene polymorphism is associated with lung function modified by smoking exposure[36], and contributes to the development of lung cancer[37]. These reports suggest the critical role of GSTO2: rs7085725 in smoking related HCC risk. Our data indicated the significant association between GSTO2: rs7085725 and smoking behavior, which always predicts a relatively poor survival[21,38]. We further investigated the interaction between SNP GSTO2: rs7085725 and smoking exposure in modulating HCC patient survival. Interestingly, these variant allele carriers of GSTO2: rs7085725 were more susceptible to smoking exposure, when compared with wild type allele carriers. All the above results clearly demonstrated that the population carrying variant alleles of GSTO2: rs7085725 might obtain more benefit of smoking cessation, which may guide those patients to make better decision.
It has been reported that the polymorphisms in the GSTO2 gene affect its expression levels in Alzheimer and Parkinson disease[39]; however, the influence of rs7085725 on GSTO2 expression in HCC remains unclear. Our data showed that rs7085725 minor allele carriers were significantly increased in high GSTO2 expression subgroup, when compared with median and low expression subgroups. Although the biological function of GSTO2 has not been clearly demonstrated, it has been observed that the expression of GSTO1, which was an important paralog gene of GSTO2 and exhibited very similar biological function to GSTO2, was over-expressed in chemo- and radio-resistant cancer cells[40,41]. Besides, elevation of GSTO1 protein in colorectal cancer cells was also involved in cell invasion and metastasis[42]. Above evidence suggested that high GSTO2 expression might contribute to tumor progression. This hypothesis might partly explain why rs7085725 WV + VV allele carriers always had worse overall survival. To date there is no report of regulating mechanism of rs7085725 on GSTO2 expression. The functional prediction indicates that rs7085725 is located in the 3’-UTR of the GSTO2 gene. Previous evidence showed that SNP sequences in 3’-UTRs of many other genes are clearly involved in the regulation of mRNA expression either by providing mutated binding sites for proteins and microRNAs to alter mRNA stability, or by forming hairpin loop structures to stabilize and thus slow down mRNA degradation. This suggests a possibility of elevated expression regulation on the GSTO2 gene by rs7085725. However, all given explanations are speculative and merit further research.
There are several strengths in this study. The patients analyzed in this study were enrolled from Xi’an and adjacent area, and these patients were all surgically treated to remove the primary tumors by the same medical center. The highly homogenous patient characteristics and treatments, as well as low rate of patient loss to follow-up, greatly reduced the confounding effects of the heterogeneous therapeutics modalities in many other similar biomarker studies of HCC prognosis. The limitations of our study include the generalizability issue, because our study was restricted to Han Chinese. Further evaluation is necessary to determine whether these findings can be generalized to other ethnic groups. Moreover, the moderate sample size limited the validity of some stratified analyses. Following validation using larger independent populations is warranted.
In conclusion, our study presents the first epidemiological evidence supporting a role of GSTO2: rs7085725 and GSTP1: rs4147581 in the prognosis prediction in a Chinese population of HCC patients.
ACKNOWLEDGMENTS
We sincerely thank Dr. Ya-Min Liu and his colleague at Department of Cardiology and Peripheral Vascular Medicine, The First Affiliated Hospital of Medical College, Xi’an Jiaotong University for their support in sample collection. We also thank researchers at Queen Mary Hospital, the University of Hong Kong, who uploaded datasets to GEO database.
COMMENTS
Background
Previous studies have revealed that several GST genes are highly polymorphic, with particular genotypes shown to be associated with cancer risk and progression. Evidence has shown that some single nucleotide polymorphisms (SNPs) of GST were associated with HCC risk. However, none of these polymorphisms are investigated in relation to survival of patients with hepatocellular carcinoma (HCC).
Research frontiers
HCC is one of the most common malignancies worldwide, especially in China, and it largely remains an incurable disease. Traditional clinicopathological features such as tumor stage, histological grade and concentration of serum AFP seem to be insufficient to predict clinical outcomes in HCC patients after surgical treatment. Therefore, it is extremely urgent to explore novel biomarkers to discriminate patient with different clinical outcomes and direct future treatment for HCC patients.
Innovations and breakthroughs
In recent decades, widespread efforts have been made to investigate tumor tissue and serological markers that would predict survival of resected HCC patients. However, to date there have been very few studies to explore the prognostic value of genetic markers. This study presents the first epidemiological evidence supporting a role of genetic markers in the prognosis prediction in a Chinese population of HCC patients.
Applications
By establishing the novel prognostic predictive genetic markers, this study may represent a future strategy for cancer prediction in the follow-up of patients with HCC.
Terminology
The term “genetic polymorphism” refers to genetic variants within the population that allow evolution by natural selection. It is defined by the occurrence in the same population of multiple discrete allelic states of which at least two have a high frequency (conventionally of 1% or more). Genetic polymorphism as a predictive marker for clinical outcomes has been extensively investigated.
Peer-review
The manuscript investigates the effects of SNPs in GST genes on survival of HCC patients, and suggests that the GSTO2 and GSTP1 gene polymorphisms may serve as independent prognostic markers for HCC patients. This study may be very useful for a large number of hospitals worldwide.
Footnotes
P- Reviewer: Guo YM, Luo HS S- Editor: Yu J L- Editor: Wang TQ E- Editor: Ma S
Zhang YJ, Chen Y, Ahsan H, Lunn RM, Chen SY, Lee PH, Chen CJ, Santella RM. Silencing of glutathione S-transferase P1 by promoter hypermethylation and its relationship to environmental chemical carcinogens in hepatocellular carcinoma.Cancer Lett. 2005;221:135-143.
[PubMed] [DOI][Cited in This Article: ][Cited by in Crossref: 78][Cited by in F6Publishing: 80][Article Influence: 4.2][Reference Citation Analysis (0)]
Ding S, Gong BD, Yu J, Gu J, Zhang HY, Shang ZB, Fei Q, Wang P, Zhu JD. Methylation profile of the promoter CpG islands of 14 “drug-resistance” genes in hepatocellular carcinoma.World J Gastroenterol. 2004;10:3433-3440.
[PubMed] [DOI][Cited in This Article: ]
Zhong S, Tang MW, Yeo W, Liu C, Lo YM, Johnson PJ. Silencing of GSTP1 gene by CpG island DNA hypermethylation in HBV-associated hepatocellular carcinomas.Clin Cancer Res. 2002;8:1087-1092.
[PubMed] [DOI][Cited in This Article: ]
Pongstaporn W, Rochanawutanon M, Wilailak S, Linasamita V, Weerakiat S, Petmitr S. Genetic alterations in chromosome 10q24.3 and glutathione S-transferase omega 2 gene polymorphism in ovarian cancer.J Exp Clin Cancer Res. 2006;25:107-114.
[PubMed] [DOI][Cited in This Article: ]
Allen M, Zou F, Chai HS, Younkin CS, Miles R, Nair AA, Crook JE, Pankratz VS, Carrasquillo MM, Rowley CN. Glutathione S-transferase omega genes in Alzheimer and Parkinson disease risk, age-at-diagnosis and brain gene expression: an association study with mechanistic implications.Mol Neurodegener. 2012;7:13.
[PubMed] [DOI][Cited in This Article: ][Cited by in Crossref: 58][Cited by in F6Publishing: 68][Article Influence: 5.7][Reference Citation Analysis (0)]
