Japanese Journal of Clinical Oncology Advance Access published online on December 3, 2008
Japanese Journal of Clinical Oncology, doi:10.1093/jjco/hyn130
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© The Author (2008). Published by Oxford University Press. All rights reserved
Prognostic Significance of X-ray Cross-complementing Group 1 T-77C Polymorphism in Resected Non-small Cell Lung Cancer
1 Institute of Medicine, Chung Shan Medical University, Taichung
2 Institute of Medical and Molecular Toxicology, Chung Shan Medical University, Taichung
3 Lung Cancer Research Center, Chung Shan Medical University, Taichung
4 Department of Surgery, China Medical University, Taichung, Taiwan, Republic of China
5 Department of Internal Medicine, Da Chien General Hospital, Miaoli
6 Department of Surgery, Chung Shan Medical University, Taichung
For reprints and all correspondence: Huei Lee, Lung Cancer Research Center, Institute of Medical and Molecular Toxicology, Chung Shan Medical University, No. 110, Sec. 2, Chien-Kuo N. Rd., Taichung, Taiwan, Republic of China
Received July 31, 2008; accepted October 21, 2008
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Abstract |
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 TOP Abstract INTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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Objective: A novel T-77C polymorphism in the promoter region of the DNA repair gene X-ray cross-complementing group 1 (XRCC1) may modulate its transcription to increase the risk of lung cancer. Here, we attempt to clarify: (i) whether the XRCC1 T-77C polymorphism was associated with lung cancer risk in Taiwanese and (ii) whether this polymorphism could act as a prognostic indicator to predict the clinical outcome of non-small-cell lung cancer (NSCLC) patients.
Methods: A total of 294 primary lung cancer patients and 288 potential controls were recruited into our study. Clinical data were collected. The genotypes of XRCC1 T-77C were identified by polymerase chain reaction.
Results: Our case–control study showed that the XRCC1 T-77C polymorphism was not associated with the risk of lung cancer in Taiwanese patients. To verify the impact of the XRCC1 T-77C polymorphism on the clinical outcome of NSCLC, survival analysis showed that patients with TT had a lower survival rate than those with the TC + CC genotype (33.1% versus 48.8%, P = 0.031). The Cox regression analysis further indicated that patients with the TT genotype had a 1.84-fold risk compared with those with the TC + CC genotype (95% CI, 1.16–2.86, P = 0.008).
Conclusion: Our results suggest that XRCC1 T-77C variants (TC + CC) may act as a favorable prognostic indicator of resected NSCLC.
Key Words: XRCC1 genetic polymorphism • NSCLC • prognosis
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INTRODUCTION |
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TOPAbstractINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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Lung cancer is the leading cause of cancer death worldwide (1). Despite every effort to improve the treatment result in these patients, the overall 5-year survival rate for non-small-cell lung cancer (NSCLC) remains at less than 20% (2,3). Although a number of clinical prognostic factors have an influence on the overall survival rate of NSCLC, none of these has been shown to have a precise relationship with the response and sensitivity to various treatment modalities. Therefore, other prognostic factors for NSCLC, including molecular markers that are based on an individual’s sensitivity to a treatment regimen, are needed.
Recently, more and more evidence shows that polymorphic variations in DNA repair genes can explain inter-individual differences in survival with a cisplatin-based treatment regimen in NSCLC patients independent of performance status, the primary clinical prognostic factor (4,5). Understanding the correlation between DNA repair genotypes and survival in NSCLC can help to elucidate how certain polymorphic variations can adversely or favorably influence the clinical outcome. Theoretically, these polymorphisms can favorably influence the response to chemotherapy through the inefficient repair of DNA damage and the enhancement of chemotherapy sensitivity, leading us to hypothesize that an inefficient DNA repair mechanism in these patients could make them more chemosensitive (6,7).
X-ray cross-complementing group 1 (XRCC1) is one of the major DNA repair proteins involved in the base excision repair pathway. A functional polymorphism in the XRCC1 gene may lead to decreased DNA repair capacity and thus confer an inherited predisposition to cancer risk. There are two large and outstanding case–control studies to confirm that novel T-77C polymorphisms may contribute to the lung caner development owing to the decreased transcriptional activity of the C-allele-containing promoter with higher affinity to Sp1 binding (8,9). The first study comprising 710 patients with incident lung cancer and 710 cancer-free controls found that compared with the -77TT wild-type homozygote, the variant genotypes were associated with a significant increase in the risk of lung cancer (8). In contrast, no significant association was observed between the other two exon variants (Arg194Trp and Arg399Gln) and lung cancer risk (8). The second study examined 1024 patients and 1118 controls to show that the -77T>C polymorphism was significantly associated with an increased lung cancer risk (9).
On the basis of these findings, we conducted a case–control study in Taiwanese patients to investigate the correlation of XRCC1 T-77C polymorphisms with lung cancer risk and its prognostic value in resected NSCLC patients.
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PATIENTS AND METHODS |
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TOPAbstractINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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Study Subjects
Since 1993, 294 patients who were newly diagnosed and pathologically confirmed with lung cancer were recruited from Taichung Veterans General Hospital, located in Central Taiwan. All of these patients were asked to submit written informed consent approved by the Institutional Review Board. The disease stage and histological types of tumors were determined by pathology findings. Of the 294 cases with histologically confirmed primary lung cancer, 155 (52.7%) were adenocarcinoma (ADC) and 139 (42.2%) were squamous cell carcinoma (SCC). During our study period, our lung cancer patients with Stages II–IV were treated under the same protocol after surgery. When tumor recurrence or metatstasis was found in these patients, radiotherapy and/or chemotherapy were used (alone or in combination) to control the disease progression. Unfortunately, the relevant information of 23 of 294 patients was lost, and 86 of 271 patients were shown to have the information of tumor recurrence and/or metastasis. Among these, 29 patients received radiotherapy, and 43 patients received cisplatin, gemcitabine, cisplatin plus gemcitabine or cisplatin plus taxol chemotherapy as first-line chemotherapy. Median follow-up time was 35.9 months (range, 0.1–135.4 months). There were 190 deaths. Meanwhile, 288 potential controls were randomly selected from consecutive patients with no history of cancer. They were admitted to the same teaching medical center for physical check-up. Demographic data, including age, gender and smoking status, were collected on each of these individuals from patient interviews and a review of the hospital charts, with informed consent. In this study, non-smokers were defined as having smoked less than two packs during their lifetime.
Genomic DNA of Adjacent Normal Lung Tissues from Lung Cancer Patients and Blood Samples from Non-Cancer Controls
Adjacent normal lung tissues from lung cancer patients and venous bloods from non-cancer control subjects were enrolled for the collection of genomic DNA. The blood cells were isolated by the Ficoll-Paque method. Frozen tissues were homogenized in 10 mM Tris, 0.1 M NaCl, 25 mM EDTA (pH 8.0) and 0.5% SDS on ice. The aqueous supernatant was incubated with RNase A and RNase T1 (250 µg/ml, Sigma Chemical Co.) at 37°C for 60 min, followed by digestion with proteinase K (10 µg/ml, Merck) at 55°C for 12 h. The digestion was extracted twice with phenol:chloroform:isoamyl alcohol (25:24:1, v/v/v). Then, sodium acetate (0.3 M final concentration) was added to the aqueous supernatant. DNA was precipitated with ethanol at –20°C and dissolved in water.
T-77C Polymorphisms of XRCC1
Genotyping of the T-77C polymorphism of the XRCC1 gene was performed by polymerase chain reaction (PCR) amplification using the primer set of 5'-GAG GAA ACG CTC GTT GCT AAG-3' and 5'-TCC TCA TTA ATT CCC TCA CGT C-3' (9). The amplified products were digested with BsrBI and analyzed by electrophoresis on a 3% agarose gel. Detailed information of the PCR assays can be found in Hao et al. (2004) (10).
Statistical Analysis
Statistical analysis was performed using the SPSS statistical software program (Version 11.0 SPSS Inc., Chicago, IL, USA). Comparisons of age, gender and smoking status between case and control groups were performed by Student's t-test or 
2-test. An unconditional logistic model was employed to obtain the odds ratio (OR) and 95% confidence interval (95% CI) for each variable between lung cancer case and control groups. Overall survival duration of lung cancer patients was calculated from the first day of treatment until death or last known follow-up. Survival probabilities were estimated using the Kaplan–Meier method and assessed using a log-rank test for each variable. Subsequently, a multiple Cox regression model was performed to obtain the adjusted hazard ratio (HR) and 95% CI for potential prognostic factors in lung cancer patients. All P values were calculated from two-tailed statistical tests.
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RESULTS |
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TOPAbstractINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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Basic Characteristics of Study Subjects
The basic information on study subjects is summarized in Table 1. A significant difference was found between the lung cases and controls in gender distribution (t-test, P < 0.001). Mean age was 66 years (range: 26–84 years) for lung cancer patients and 57 years (range: 31–85 years) for control subjects at recruitment. Meanwhile, proportions of male and smoker were not different in the lung cancer cases compared with the controls. In addition, a relatively higher proportion of lung cancers were diagnosed at Stage I (36.1%) and Stage III (42.5%).
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XRCC1 T-77C Genotypes and Risk of Lung Cancer
Previous Chinese studies have shown an association between XRCC1 T-77C genotypes and lung cancer risk (8,9). Here, we performed a case–control study to assess whether XRCC1 T-77C genotypes could be related with the risk of lung cancer in Taiwan. Among the control participants, the XRCC1 T-77C genotype distribution was in Hardy–Weinberg equilibrium (
2 = 0.09, P = 0.98). Our data showed that the genotype frequencies of XRCC1 T-77C in the cases and controls were not different (TT, 85.4% versus 86.8%; TC, 13.6% versus 12.8%; CC, 1.0% versus 0.3%, respectively). Although subjects with the CC genotype had a 2.99-fold risk of lung cancer compared with those with the TT genotype, it was not statistically significant (95% CI, 0.31–28.92; Table 2). This was due to the fact that the frequency of the CC genotype in both the cases and controls was too low to assess the lung cancer risk. When the CC genotype was combined with the TC genotype, these subjects (TC + CC) did not appear to have a higher lung cancer risk than those with the TT genotype (OR, 1.13; 95% CI, 0.70–1.80; Table 2). We further examined the association between XRCC1 T-77C polymorphism and lung cancer risk by viewing the stratification by tumor type, gender, age and smoking status. We found a trend for an increased risk existed only in SCC (OR, 1.37; 95% CI, 0.79–2.40; P = 0.263; Table 3) and male patients (OR, 1.53; 95% CI, 0.84–2.76; P = 0.159).
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The Influence of XRCC1 T-77C Polymorphism on the Clinical Outcome of Lung Cancer
The influence of clinical parameters on the outcome of lung cancer patients was assessed by a log-rank test. Among the parameters, the patients’ survival rate was significantly associated with tumor stage, T and N values and XRCC1 T-77C genotypes (Table 4). Lung cancer patients with Stage I (53.8%) had a significantly higher survival rate than those with different Stage II (38.8%), Stage III (20.8%) and Stage IV (14.3%), respectively. Similarly, patients with T1 + T2 (39.8%) and N0 (51.1%) had a significantly higher survival rate than those with T3 + T4 (20.0%; P < 0.0001) and N1 + N2 + N3 (20.9%; P < 0.0001), respectively. More interestingly, patients with the TC + CC genotypes (48.8%) had a markedly higher survival rate than those with the TT genotypes (33.1%; P = 0.031, log-rank test; Fig. 1).
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Cox’s regression analysis further indicated that patients with the TT genotype had a 1.84 HR compared with those with the TC + CC genotype, suggesting that the TT genotype may act as an independent poor prognostic factor of resected NSCLC (95% CI, 0.35–0.86; P = 0.008; Table 5). As expected, tumor stage had an independent prognostic value in these patients (HR, 2.64; 95% CI, 1.89–3.69; P < 0.0001). In addition, the poor prognosis of ADC and the male gender, compared with its counterparts, were notable as independent prognostic factors in this study population (HR, 1.62; 95% CI, 1.15–2.28; P = 0.005 for tumor type; HR, 1.54; 95% CI, 1.03–2.31; P = 0.035 for gender factor).
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DISCUSSION |
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TOPAbstractINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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Based on this case–control study, the XRCC1 T-77C polymorphism was not associated with the risk of lung cancer in Central Taiwan (Table 2). This finding was not consistent with previous reports, which have shown that the XRCC1 T-77C polymorphism was associated with the risk of lung cancer in Northern (9) and Southern China (8) (TC + CC versus TT: OR, 1.46; 95% CI, 1.18–1.82 for Northern China; OR, 1.55; 95% CI, 1.21–1.98 for Southern China). Meanwhile, the impact of XRCC1 TC + CC genotypes on lung cancer risk was more pronounced for subjects with SCC, males, smokers and those younger than 57 years. Interestingly, a similar trend was also found in the present study, although the results did not reach statistical significance (Table 3). No association of XRCC1 T-77C polymorphism with lung cancer risk in this study could be due to the fact that the prevalence of TC + CC genotypes in the lung cancer group of Taiwanese was higher than that of the lung cancer group of Chinese (14.6% for Taiwanese versus 23.5% for Chinese). Meanwhile, the frequency of TC + CC genotypes in Taiwanese non-cancer controls was relatively similar to that of Chinese non-cancer controls (13.2% for Taiwanese controls versus 17.4% for Chinese controls). Nevertheless, the different impacts of XRCC1 -77T > C polymorphism on lung cancer risk of Chinese in Mainland China and Chinese living in Taiwan necessitate increasing the numbers of study populations in further investigations.
The association of two other genetic polymorphisms of XRCC1, Arg194Trp and Arg399Gln, with lung cancer risk, was not observed by our study (data not shown). Consistent results were also reported by studies of Japanese and Northeastern Chinese (11,12). Additionally, the prognostic value of two genetic polymorphisms was also not found in our studied population (data not shown). However, previous reports indicated that XRCC1 Arg399Gln polymorphisms may be an important prognostic factor in lung cancer patients receiving platinum chemotherapy (13,14). In addition, the XRCC1 polymorphism Arg194Trp had an effect on the survival rate of lung cancer patients treated with radiotherapy (7). In Cox regression analysis, male and ADC had worse prognosis compared with female and SCC (Table 5). We did not see the prognostic value in the category of ADC or SCC and female or male (data not shown). Having stratified smoking status, TT genotype can act as an independent poor survival indicator in non-smoking lung cancer (HR, 2.04; 95% CI, 1.09–3.85; P = 0.027), but not in smoking lung cancer. Thus, the XRCC1 T-77C polymorphism may be used to potentially predict the clinical outcome of lung cancer patients who undergo thoracic surgical therapeutics, particularly in non-smokers. However, the prognostic significance of the XRCC1 polymorphism was not observed in Stage I patients without radio- and/or chemotherapy (P = 0.294). To our knowledge, this is the first report to indicate the prognostic significance of the XRCC1 T-77C polymorphism in resected NSCLC patients.
In theory, DNA repair gene polymorphisms could affect the clinical outcome of lung cancer in two opposing ways. Polymorphisms, contributing to suboptimal DNA repair in the tumor, could lead to more biologically aggressive tumors; conversely, these same polymorphisms could favorably influence the response to platinum agents through the inefficient removal of platinum–DNA adducts. Our preliminary real-time RT–PCR data support the finding of previous study showing that XRCC1 mRNA expression levels in tumors with the TT genotype were significantly higher than those of the TC + CC genotypes (9). We thus suggest that a poorer clinical outcome in patients with the XRCC1 TT genotype could be due to tumors with lower drug sensitivity than those with the XRCC1 TC + CC genotype. In conclusion, a simple detection of the XRCC1 T-77C polymorphism from peripheral bloods may be useful to predict the clinical outcome of resected NSCLC in choosing a feasible clinical treatment for these patients.
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Conflict of interest statement |
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TOPAbstractINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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None declared.
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Funding |
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TOPAbstractINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONConflict of interest statementFundingReferences |
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This work was supported by grants from the National Science Council (NSC96-2628-B-040-002-MY3, NSC97-2314-B-040-027-MY3), The Executive Yuan, Republic of China.
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References |
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