The Association Between CXCL12-G801A Polymorphism And Non-small Cell Lung Cancer Risk In The Chinese Han Population

Background and objectiveLung cancer, the main cause of global cancer deaths, accounts for about 20% on the mortality of all malignant tumors, of which more than 80% is non-small cell lung cancer (NSCLC). According to the released data of world health organization (WHO):the incidence (1.2 million/year) and mortality (1.1 million/year) of lung cancer are ranking first in all the cancer, and there is an upward trend in its morbidity and mortality. According to the data published by cancer prevention and control office in China, since the 1970s, mortality of lung cancer in China rose by nearly five times. A lot of research shows that:smoking is closely related to the pathogenesis of lung cancer and smokers are nearly 10 times higher risk of lung cancer than non-smokers. In addition, China currently has more than a third of world’s smoking population and due to changes in lung cancer incidence rates lagging about 20 to 30 years compared with smoking, in the near future, proportion of lung cancer morbidity and mortality worldwide will be more prominent. Lung cancer has become a threat to both global and China, therefore, it is one of the important diseases of residents’ lives and health and it is a major public health problem to be resolved. Along with the advance level of lung cancer molecular biology research, the research on non-small cell lung cancer genetic direction is also more and more.Research results of the Human Genome Project (HGP) show that 99.9% of different individual genome is the same and there is only minimal (0.1%) of genetic differences in the sequence, mainly single nucleotide polymorphisms (SNPs). SNP is refers to the genetic level caused by a single nucleotide mutation of DNA sequence polymorphism and the frequency is not less than 1% in the group including single-base transformation, transversion and insertions or deletions. In the human genome, three or four alleles are very rare form of SNP, so simple SNPS also refers to the double allele and the combination of two different alleles is called the genotype. A huge number of SNPs exist in the human genome, some of which may influence the expression of gene and the structure or function of encoded protein, and then influence the individual risk of disease, drug reactivity and the prognosis of different individuals. High-density, representative, genetic stability and detection of characteristics such as easy to implement of SNPs in the gene sequence embodies the superiority of a new generation of genetic markers, became the third generation of genetic markers after the first generation of restriction fragment length polymorphism markers and the second generation of microsatellite marker, and had been accepted rapidly as a very active content in the field of genetic research. According to the position of gene, SNP area can be divided into the genes that code for SNPS (Coding SNPS, cSNP) and gene regulation area SNPS (Regulatory SNPS, rSNP). According to whether to change the coding of amino acids, cSNP can be divided into Synonymous cSNP and Non-Synonymous cSNP. Synonymous cSNP refers to the changes in the base sequence does not affect the protein amino acid sequence, while non-Synonymous cSNP may change the encoding of the kinds of amino acid residues to change the structure and function of the protein. In addition, through the influence and the transcription factor or combination of microRNAs, the regulation of rSNP can cause changes in the level of gene expression or mRNA stability and the change of the translation process, which affect the normal cell function. As a result, these potential functional SNPs is generally preferred for study on the relationship in the development of disease and prognosis for malignant tumors, including lung cancer, which will provide a scientific basis for prevention and clinical treatment.Research has found that the cell cycle disorder is the main mechanism of tumor and the core of the change in cell malignant is the normal cell cycle process destroyed making tumor cells enter the cell cycle process, changing relevant regulatory proteins and signal transduction pathways and losing their differentiation capacity to undergo malignant proliferation. In addition to the cell cycle regulation factor, chemokines is another type of family closely related to the development of tumor. Chemokine is a kind of molecular, weight of 8-12 kD, at least two conservative half creatine (Cys) secretory type single protein that can activate and affect white blood cells, to participate in the immune adjustment and immune pathological reaction. According to its N-terminal sequence Cys, chemokines can be divided into four categories:CXC (a), CC (β), C (γ) and CX3C (δ). Chemokines, mainly through G protein coupling with its specific receptor, involved in the generation of hematopoietic cells, white blood cell migration and homing, development of embryo, and many other normal life activities, but also involved in inflammatory diseases, viral infection, graft rejection, malignant tumor, and many other pathological process.In the development of tumor, different chemokines play a different role:part of chemokines can promote tumor proliferation, promote angiogenesis, inducing tumor cell move, and adhere to endothelial cells, accelerate the spread of the tumor and metastasis; part of chemokines can inhibit the growth of tumor cells and the form of blood vessels, stimulate the body specific and nonspecific immune response, thus inhibiting tumor growth and metastasis. Research shows that most of the tumors can secrete a variety of abnormally high levels of chemokines and their expression profiles of difference can determine the kinds of tumor infiltrating leukocytes and infiltrating degree, and tumor metastasis and similar the formation process of inflammatory cells infiltration:scroll, involving cell adhesion and migration of endothelial. In addition, there are multiple reports found, CXC (a) and CC (β) and the expression of chemokine disorders can affect the occurrence progress and prognosis of a variety of tumors, including lung cancer. Lung cancer cell line, for example, has down regulated the expression of CXCL12 and CXCL12 apparent silence is an independent predictor of prognosis of NSCLC, high expression and CCL22 without disease progression of lung cancer was significantly associated with survival and recurrence risk.Chemotactic factor CXCL12 that matrix derived factor (stromal cell-derived factor, SDF-1), in development of tumor, mature lymphocytes and homing, regulating immune inflammatory reaction, promoting angiogenesis plays an important role. In coding gene transcription 3’the translation section of CXCL12 is 801th of guanine (G) to the polymorphism of adenine (A) mutations (remember CXCL12-3’ G801A, rs1801157), the gene loci polymorphisms and asthma, the correlation of HIV infection and transplant rejection has been reported. In recent years, many studies have shown that CXCL12-3 ’G801A gene polymorphisms involved in tumor formation, development, invasion and metastasis of multiple processes. The invasion and metastasis is lead to malignant tumor treatment of the main causes of failure.From what has been discussed above, a series of evidence shows that chemokine family related gene expression or function of the change in tumor genesis, including NSCLC, play an important role in the development and prognosis in NSCLC. So far, however, there is less about the gene polymorphism and the prognosis of NSCLC analysis, which has published literature and is only focus on a handful of genes a few sites or only study of European and American people. The purpose of this study is to detect CXCL12-G801A genotype distribution through the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of non-small cell lung cancer, and further investigate CXCL12-G801A gene polymorphism of han population in our country and the influence of non-small cell lung cancer susceptibility.Methods1.408 cases of hospitalized patients pathological diagnosed with non-small cell lung cancer from September 2009 to October 2009. In addition, we recruit 303 healthy individuals to conduct the study (table 1). Non-small cell lung cancer patients did not receive preoperative radiotherapy or chemotherapy. Recruited 303 healthy volunteers were han Chinese residents from surrounding provinces, cities, and between the two group of patients there is no blood relationship. Collection of non-small cell lung cancer group and healthy group patient’s age, gender, and smoking history. Study plan for ethics committee approved the participation unit and the subjects were informed and agreed to take part in.2. Respectively take on an empty stomach peripheral blood 5 ml, proteinase K digestion-saturated sodium chloride salting out method is used to extract the genomic DNA of both groups, and by PCR-RFLP analysis method and PIRA-polymerase chain reaction (PCR) to detect CXCL12-G801A three basic genotype (G/G, A/A and A/G) frequency distribution.3. Statistical analysis of experimental data SPSS 18.0 statistical software was used for analysis. Case group and control group the frequencies of alleles and genotypes with gene counting method. Case group and control group in demographic characteristics between genotype frequency and chi-square test was used to assess the differences. Measurement data to x±s. Inspection level of significance (P< 0.05).Results1. Judgment of CXCL12-G801A genotypesAfter Mspl enzyme reaction, Mutant allele G/G homozygous was cut to two bands as 202 and 100 bp, A/A homozygote for 302 bp and A/G heterozygote showed 3 stripes as 302,202 and 100 bp2. General data of the case group and healthy controlsThere was no significant difference of gender composition in patients between non-small cell lung cancer group and healthy controls (x= 3.384,.P=0.066)3. Age of the two groupsThe average age in experimental group and control group were 61.31±10.0/ 62.50±9.86 years respectively. There was no significant difference in patients’ age between non-small cell lung cancer group and healthy controls. (P=0.365)4. Analysis on CXCL12-G801A SNPs and risk of non-small cell lung cancerG/G genotype frequency of CXCL12-G801A in the experimental group and control group were 59.56% and 66.37% respectively. A/A genotype frequency of CXCL12-G801A in the experimental group and control group were 10.29% and 7.92% respectively. A/G hybrid frequency of CXCL12-G801A in the experimental group and control group were 30.15% and 25.74% respectively. There was no statistically significant difference of CXCL12-G801A genotype frequency between the two groups (χ2=3.527, P=0.171).5. The relationship between CXCL12-G801A alleles distribution and non-small cell lung cancer susceptibilityGenotypes and alleles of CXCL12-G801A in healthy controls (χ2= 0.880, P=0.644) and NSCLC group (x2= 0.835, P=0.659) were in Hardy Weinberg equilibrium, the distribution of polymorphic alleles G and A frequency of CXCL12-G801A in NSCLC group and control group are 74.63%,79.21%,25.37% and 20.79% respectively, and there was no statistically significant difference of CXCL12-G801A polymorphic alleles G and A frequency between the two groups (χ2=2.495, P =0.114).6. Through the study of the experimental group and the control group, we have not observed any clinical parameters and popular genotypes that significantly associated with CXCL12-G801A polymorphism).7. According to the pathological classification and lung cancer staging, there was no statistical significance among the distribution of the three basic genotype in CXCL12-G801A(G/G, A/A andA/G)(x2= 1.349, P=0.509; χ2= 12.141,P=0.059). CXCL12-G801A gene polymorphism was not related with pathological classification and lung cancer staging in non-small cell lung cancer.8、CXCL12-G801A gene polymorphism was not related with EGFR mutation in adenocarcinoma of lung (χ2=1.196, P=0.550).9.1-year survival rate was not statistically different in three basic genotype in CXCL12-G801A(G/G, A/A andA/G) (χ2= 3.096, P=0.213).Conclusions:1. The distribution of three basic genotype (G/G and A/A, A/G) of CXCL12-G801A has no relevance with the incidence of non-small cell lung cancer. CXCL12-G801A gene polymorphism is not related with the risk of non-small cell lung cancer. CXCL12-G801A gene polymorphism is not a risk factor for non-small cell lung cancer in Chinese Han population.2. The distribution of A and G polymorphism allele of CXCL12-G801A has no relevance with the incidence of non-small cell lung cancer. The distribution of A and G polymorphism allele of CXCL12-G801A is not a risk factor for non-small cell lung cancer in Chinese Han population.3. The distribution of three basic genotype (G/G and A/A, A/G) of CXCL12-G801A has no correlation with pathological classification and lung cancer staging of non-small cell lung cancer in Chinese Han population.4. CXCL12-G801A gene polymorphism was not related with EGFR mutation in adenocarcinoma of lung in Chinese Han population.5. CXCL12-G801A gene polymorphism was not related with 1-year survival rate.