| Objective: Lung cancer, the most common malignant tumor threatens the health and life of human beings with its incidence and mortality are increasing rapidly. Lung cancer is the first killer of cancer-related deaths both in men and women in the world. Previous studies have demonstrated that cigarette smoking, various occupational exposures and carcinogens in heavily polluted air are the risk factors to lung cancer. Although it is well recognized that most of lung cancer cases are attributable to these factors, only 15% of all smokers develop lung cancer and less than 10% of all diagnosed lung cancer are among non-smokers. Recently, along with the development of the molecular biology, the effect of hereditary susceptibility has been focused on the research of lung cancer.Angiogenesis is an important step in the development of tumor and is necessary for primary tumor growth, invasiveness, and metastasis. Vascular endothelial growth factor (VEGF) is one of the powerful factors to facilitate angiogenesis as we know so far. Currently, studies show that some tumors may induce a higher serumal, urinary or intratumoral VEGF level, for instance, breast cancer, gastric cancer, lung cancer, prostatic carcinoma, and esophageal carcinoma. VEGF gene has high incidence of polymorphism. At least 30 single nucleotide polymorphisms (SNP) have been identified in the VEGF gene. This study was designed to investigate the relationship between two single nucleotide polymorphisms (-2578C/A and +405G/C) in the promoter region of VEGF gene and the susceptibilitiy of lung cancer in North China.Methods: This hospital-based case-control study comprised 172 patients with lung cancer (62 squamous cell carcinomas, 41 adenocarcinomas, 17 adenosquamous carcinoma,26 small cell lung carcinomas, and 26 other pathological types including large cell carcinoma, clear-cell carcinoma, signet-ring cell carcinoma, carcinoma sarcomatodes, bronchioloalveolar carcinoma, and so on included) and 184 frequency-matched healthy controls with no evidence of disease. Five milliliter of venous blood from each subject was drawn in Vacutainer tubes containing EDTA and stored at 4℃, meanwhile the medical history and personal data of each subject was obtained. The genomic DNA was extracted within one week after bleeding by using proteinase K digestion followed by a salting out procedure. Genotypes of the VEGF-2578C/A and +405G/C SNPs were analyzed by polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP) method and primer-introduced restriction analysis PCR (PIRA-PCR).Statistical analysis was performed using SPSS11.5 software package. A probability level of 5% (P<0.05) was considered significant. Hardy-Weinberg analysis was performed by comparing the observed and expected genotype frequencies in study groups using Chi-square test. The age difference of cases and frequency-matched controls was validated by the t-test. Comparison of the VEGF gene genotype and allelotype distribution in cancer patients and healthy controls was performed by means of two-sided contingency tables using Chi-square test. The VEGF -2578C/A and +405G/C haplotype frequencies and linkage disequilibrium coefficient were estimated by using the EH linkage software and 2LD program, respectively. The odds ratio (OR) and 95% confidence Interval (CI) were calculated by using an unconditional logistic regression model and adjusted by age, gender and smoking status accordingly.Results: 1 The frequency of smokers in lung cancer patients (58.1%) was significantly higher than that in healthy controls (42.4%) (χ2 =8.200, P value= 0.004). Therefore smoking may increased the risk of developing lung cancer (age and gender adjusted OR=2.448, 95%CI=1.382-4.337, respectively). 2 The distribution of the VEGF-2578C/A SNP genotypes among healthy controls did not significantly deviate from that expected by Hardy-Weinberg equilibrium (P value>0.05). In lung cancer patients, frequency of the C and A allele were 70.9% and 29.1% respectively; in healthy controls, the frequencies were 78.5% and 21.5% respectively. The frequency of the A allele in lung cancer patients was significantly higher than that in healthy controls (χ2= 5.064, P value=0.020). The frequencies of three genotypes (C/C, C/A and A/A) in healthy controls were 60.9%, 35.3% and 3.8% respectively, and 49.4%, 43.0% and 7.6% respectively in lung cancer patients. (Because of the low frequency of A/A genotype, the C/A and A/A genotype were combined). The frequencies of the two genotypes (C/C and C/A+A/A) in patients and healthy controls were 49.4%, 50.6% and 60.9%, 39.1%, respectively. There was significant difference in genotype distributions of the VEGF -2578C/A polymorphism between the two groups (χ2=4.265, P value=0.033). Compared with individuals with the C/C genotype, individuals with the C/A+A/A genotype had significantly higher risk to develop lung cancer (age, gender and smoking status adjusted OR=2.019, 95%CI=1.290-3.158. When stratified by smoking status, compared with individuals with the C/C genotype, the C/A+A/A genotype increased the risk in developing lung cancer in smokers (age and gender adjusted OR=2.043, 95%CI=1.090-3.827) With the subjects being stratified by tumor histology, compared with individuals with the C/C genotype, individuals with the C/A+A/A genotype carried an increased risk of lung cancer in squamous cell carcinomas (SCC) (age, gender smoking status adjusted OR=2.073, 95%CI=1.025-4.191). 3 The distribution of the VEGF +405G/C SNP genotypes among healthy controls did not significantly deviate from that expected by Hardy-Weinberg equilibrium (P value>0.05). When the overall lung cancer cases were compared with the controls, no significant difference was found in the distributions of genotype and allele of VEGF +405G/C (P value>0.05). When the overall lung cancer cases were categorized by smoking status, however, the G/C+C/C genotype was associated with a significantly increased risk of smokers compared with the G/G genotype (age and gender adjusted OR=2.402, 95%CI=1.177-4.904). When stratified by tumor histology, compared with individuals with the G/G genotype, individuals with the G/C+C/C genotype carried an increased risk of lung cancer in squamous cell carcinomas (SCC) (age, gender smoking status adjusted OR=2.210, 95%CI=1.081-4.517). 4 The combined effect of VEGF -2578C/A and +405G/C SNP on lung cancer was analyzed. It was shown that the two SNPs were not in linkage disequilibrium (D'=0.27149).Conclusions: 1 Smoking may increase the risk of developing lung cancer. 2 VEGF-2578C/A SNP was associated with the risk of lung cancer. When stratified by smoking status, compared with individuals with the C/C genotype, the C/A+A/A genotype increased the risk of lung cancer in smokers. When stratified by tumor histology, compared with the C/C genotype, individuals with the C/A+A/A genotype carried an increased risk of lung cancer in squamous cell carcinomas (SCC). 3 The combined G/C+C/C genotype of VEGF+405G/C SNP was associated with a significantly increased risk of smokers, compared with the G/G genotype. When stratified by tumor histology, compared with the G/G genotype, individuals with the G/C+C/C genotype carried an increased risk of lung cancer in squamous cell carcinomas (SCC). |
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