Study of association between codon72 of p53 (rs1042522) gene single nucleotide polymorphisms(SNP) with the risk of colorectal cancer in khuzestan province
Study of association between codon72 of p53 (rs1042522) gene single nucleotide polymorphisms(SNP) with the risk of colorectal cancer in khuzestan province
Introduction: Cancer is caused by uncontrolled proliferation of cells and it is the second leading cause of death in the world and the third leading cause of death in Iran. Colorectal cancer is the growth of cancer cells in the colon or rectum and it is the second most common cancer among men after lung and prostate cancer and the third most common cancer among women after lung and breast cancer.
The p53 protein is a transcription factor that is inactive in half of cancers and activated by DNA damage and induces transcription of genes involved in cell cycle control and apoptosis, and Codon 72 polymorphism of this gene can cause changes in its structure and activity.
Methods: This polymorphism was studied among the population with colorectal cancer in Khuzestan province by RFLP method and 112 patients with colorectal cancer and 100 control samples of this polymorphism were studied.
Sampling was performed in containers containing EDTA and osmotic shock was used to extract the cell extract; and DNA-bound proteins were then digested by ionic detergents. The cellular proteins were then precipitated with phenol and chloroform or saturated NaCl solution, and finally, DNA precipitation was performed with cold ethanol.
To evaluate the quantity and quality of DNA, electrophoresis methods on agarose gel and nanodrop spectrophotometry were used. In this experiment, the polymorphism of the P53 gene was studied and part of the sequence of this gene was PCR, which contained the desired nucleotide, and after sectioning, the band difference was detected by the restriction enzyme.
After performing RFLP and observing the pattern of different bands, sequences related to different patterns were sequenced and the results were compared with the natural gene sequences available at the NCBI site and the relevant genotype was determined. The test results were analyzed using SPSS statistical software, version 21.
Results: The P53 gene PCR product is a sequence of 199 nucleotides, and the restricted enzyme BstUI cleaves the sequence when the target nucleotide is G; Therefore, the presence of a 199 nucleotide band on the gel electrophoresis indicates the proline amino acid codon, and the presence of two nuclei 113 and 86 nucleotides on the gel electrophoresis indicates arginine in the codon 72 of the P53 gene; Also, seeing three nucleotide bands 199, 86 and 113 indicates that the sample is heterozygous in this case.
In the control group, 41% of the respondents have arginine / arginine genotype, 46% have arginine / proline genotype and 13% have proline / proline genotype; While in the patient group, 32.1% of the respondents have arginine / arginine genotype, 43.8% have arginine / proline genotype and 24.1% have proline / proline genotype. The amount of chi-square (chi-square) obtained by comparing the frequencies of the two groups in the three types of p53 gene genotype is equal to 4.655 and there is no statistically significant difference between the genotype in the control group and the patient in terms of p53 gene genotype. The proline / proline genotype increases the chance of developing colorectal cancer by 2.365 times and is also statistically significant, given that the p-value level is less than 0.05.
Conclusion: Mutation in P53 is one of the most abundant single nucleotide genetic changes in human cancers. During polymorphism, codon 72 of exon 4 of the P53 gene, nucleotide C is converted to G, resulting in the conversion of proline CCC to arginine CGC, altering the original structure of P53. Although both structures function normally in binding activities to specific DNA sequences, they do show some functional differences. The arginine P53 allele shows greater apoptotic potency than the proline allele, and extensive research has been conducted on this polymorphism and cancer. Therefore, in this study, the effect of converting codon 72 of p53 gene to amino acid arginine in colorectal cancer was not observed in Khuzestan province.