Tumor Biol. (2015) 36:4591–4596 DOI 10.1007/s13277-015-3104-0
RESEARCH ARTICLE
XRCC1 polymorphisms associated with survival among Chinese bladder cancer patients receiving epirubicin and mitomycin C Xiaheng Deng & Xiaolei Zhang & Yidong Cheng & Xiao Yang & Ruizhe Zhao & Xuzhong Liu & Xiao Li & Chao Qin & Qiang Lu & Changjun Yin
Received: 3 December 2014 / Accepted: 12 January 2015 / Published online: 25 January 2015 # International Society of Oncology and BioMarkers (ISOBM) 2015
Abstract The association between DNA repair gene polymorphisms and bladder cancer risk has been widely studied. However, only few studies have examined the correlation between bladder cancer and instillation agent sensitivity. The aim of this study was to examine the association between polymorphisms of DNA repair genes, namely X-ray repair cross-complementing group I (XRCC1) rs2854509 and rs3213255, and bladder cancer recurrence risk. We recruited 244 patients (130 treated with epirubicin and 114 treated with Xiaheng Deng, Xiaolei Zhang and Yidong Cheng contributed equally to this work. X. Deng : X. Zhang : Y. Cheng : X. Yang : R. Zhao : X. Liu : X. Li : C. Qin : Q. Lu (*) : C. Yin Department of Urology, The First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, 210029 Nanjing, People’s Republic of China e-mail:
[email protected] X. Deng e-mail:
[email protected] X. Zhang e-mail:
[email protected] Y. Cheng e-mail:
[email protected] X. Yang e-mail:
[email protected] R. Zhao e-mail:
[email protected]
mitomycin C). Genomic DNA was used to examine the XRCC1 rs2854509 and rs3213255 genotypes by Taqman PCR analysis. Combination analysis of XRCC1 rs2854509 and rs3213255 and examination of XRCC1 diplotypes were performed to reveal possible correlations. The rs2854509 CC and rs3213255 TT genotypes conferred shorter survival times than the rs2854509 AC/AA and rs3213255 CC/CT genotypes in patients treated with epirubicin, but not in those treated with mitomycin C (MMC) in adjusted models [hazard ratio (HR)= 0.23, 95 % confidence interval (CI)=0.10–0.53 for rs2854509 AC+AA compared with CC; HR=0.17, 95 % CI=0.06–0.46 for rs3213255 CC+CT compared with TT]. Combination analysis showed significantly increased recurrence-free survival (RFS) among patients simultaneously carrying the rs2854509 AC/AA and rs3213255 CC/CT genotypes with an HR of 0.15 (95 % CI=0.05–0.45) compared to those carrying other genotypes. Diplotype analysis demonstrated that the A-C/C-T diplotype is associated with a lower risk of recurrence compared with the common wild C-T/C-T diplotype (HR=0.17, 95 % CI=0.06–0.51). Our results suggest that the rs2854509 CC and rs3213255 TT genotypes confer higher sensitivity to epirubicin instillation. Moreover, the A-C/C-T diplotype presents significantly lower recurrence risk than other diplotypes. Keywords Bladder cancer . XRCC1 . Epirubicin . Mitomycin C . Chemosensitive
X. Liu e-mail:
[email protected] X. Li e-mail:
[email protected]
Introduction
C. Qin e-mail:
[email protected]
Bladder cancer is projected to be the most common genitourinary malignancy worldwide in 2014 [1]. In recent years, the incidence of bladder cancer has continued to increase in China as well. Approximately 75–80 % of newly diagnosed cases of
C. Yin e-mail:
[email protected]
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bladder cancer present superficial tumours; 50–70 % of these superficial tumours relapse within 5 years, and 10–20 % progress to a more aggressive disease [2]. Survival of patients with bladder cancer varies extensively and depends on tumour stage and grade at diagnosis, age, general health of the patient and applied treatments [3]. The 5-year survival rate of patients with bladder cancer is 72.8 % for men and 69.3 % for women [4]. Over 80 % of all bladder cancers are non-invasive (Ta/T1) at diagnosis and present good prognosis in response to transurethral resection followed by intravesical chemoprophylaxis (e.g., epirubicin and mitomycin C (MMC)) or immunoprophylaxis (Bacillus Calmette–Guerin, BCG) [5]. Epirubicin, which belongs to the anthracycline antibiotic family and is a derivative of doxorubicin, has a molecular weight higher than 580 kDa. In Europe, epirubicin is extensively used because of its comparable or higher efficacy and lower toxicity compared with equivalent doses of doxorubicin. Ali-ElDein et al. [6] reported that patients treated with epirubicin exhibited significantly low likelihood of recurrence; in addition, the incidence rates of local toxicity were 15.6 and 36.7 % in patients treated with epirubicin and doxorubicin, respectively. MMC is an alkylating agent that functions by inhibiting DNA synthesis. It has a molecular weight of 334 kDa and minimal systemic absorption. Thus, MMC and epirubicin present a minimal risk of systemic toxicity and can immediately be instilled after transurethral resection (TUR). Sylvester et al. [7] concluded that a dose of cytotoxic chemotherapy immediately after TUR results in 39 % decrease in odds of recurrence and 47–36 % actual decrease in recurrence rate. The responses of patients to chemo treatments involve complex biological mechanisms. Variation in individual responses to drugs may be reduced by tailoring therapies according to individual genetic profiles, which is broadly referred to as pharmacogenetics. DNA repair is crucial in maintaining genetic stability and protecting against cancer. Polymorphisms in DNA repair genes may alter repair capacity and individual susceptibility to adjuvant and intravesical treatments [8]. X-ray repair cross-complementing group I (XRCC1) protein performs a key role in the base excision repair (BER) pathway, which is involved in repair of DNA single-strand breaks after exposure to reactive oxygen species, ionising radiation or alkylating agents [9]. Clinical study has demonstrated the association between XRCC1 polymorphisms and treatment response to platinum chemotherapy in advanced non-small cell lung cancer (NSCLC) patients [10]. Tengstrom et al. found that XRCC1 rs25487 polymorphisms with the AA genotype predicted breast cancer-specific survival among 75 patients treated with adjuvant platinum-based chemotherapy [11]. XRCC1 rs2854509 and rs3213255 confer longer survival times than other genotypes, as indicated by haplotype analysis in patients treated with systemic therapeutic agents [12]. In this work, we aimed to confirm and extend these findings by
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examining the influence of these polymorphisms on clinical outcomes among bladder cancer patients. Specifically, we investigated the association between XRCC1 rs2854509 and rs3213255 haplotypes among patients treated with intravesical chemotherapy and subsequently conducted diplotype analysis.
Methods and patients From March 2007 to September 2012, 244 patients diagnosed with bladder cancer were recruited from the First Affiliated Hospital of Nanjing Medical University. This prospective study was approved by the Institutional Review Board of Nanjing Medical University. Written informed consent was provided by all patients who participated in the study. Patients were excluded from participating if they had had a history of cancer or metastasised cancer of unknown origin. Of the 244 patients included in this work, 130 were treated with epirubicin and 114 patients were given MMC to prevent cancer recurrence. All subjects were personally interviewed prior to recruitment to collect demographic data and clinical characteristics, including age, gender, tobacco and alcohol consumption, and self-reported family history of cancer. Histological grade of tumour was assessed according to a recently published WHO consensus. Pathology slides from radical nephrectomy or core biopsy were independently reviewed by two pathologists who concomitantly confirmed the diagnosis of bladder cancer. Patients were classified into two subgroups according to histopathological grade (WHO 2004, grading of urothelial papilloma): low risk and high risk. Individuals who smoked daily for over a year were defined as smokers, whereas those who did not were considered non-smokers. Individuals who drank alcohol at least three times per week for more than 6 months were defined as drinkers, whereas those who did not were considered non-drinkers. About 24 h after transurethral resection (TUR), epirubicin and MMC were instilled into the bladder. About 50 mg of epirubicin or 30 g of MMC was administered per week for 8 weeks after TUR. This dosage was maintained per month for 12 months or longer. The date of recurrence was obtained from inpatient and outpatient records or from patients’ families via follow-up; the absence of disease beyond this date was considered non-recurrence. Genotype determination Genomic DNA of each individual was extracted from 150 μL of EDTA-anticoagulated peripheral blood samples using a DNA extraction kit (Tiangen Biotech, Beijing, China) following the manufacturer’s instructions. Single-nucleotide polymorphisms were genotyped using the TaqMan genotyping
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assay (Applied Biosystems, Foster City, CA, USA) for singlenucleotide polymorphism (SNP). SDS 2.4 software was used for allelic discrimination. The primers, probes and reaction conditions for each SNP are available upon request (Table 1). Four negative controls were included in each plate, and 5 % of the samples were randomly selected for repeated genotyping to verify the results. All results were 100 % consistent. Each assay was conducted in a 384-well ABI 7900HT real-time PCR system (Applied Biosystems, Foster City, CA, USA), and each reaction was run in triplicate. Primers, probes and reaction conditions for each SNP analysis are available upon request. Amplification was performed under the following conditions: 50 °C for 2 min, 95 °C for 10 min and 45 cycles of 95 °C for 15 s and 60 °C for 1 min. Statistical analysis Recurrence-free survival (RFS) was defined as the time from the first instillation of epirubicin or MMC to the first recurrence of bladder cancer. RFS was estimated using the Kaplan– Meier method, and log-rank test was used to compare different survival rates. Hazard ratios (HRs) and their 95 % confidence intervals (CIs) were derived from univariate and multivariate Cox proportional hazard models. All analyses were carried out using SPSS 13.0. A two-sided P value that was
