A novel tri-allelic insertion/deletion polymorphism in the promoter of p21(Waf1/Cip1) and the association with gastric cancer

Abstract

Aims: p21(Waf1/Cip1) is a cyclin-dependent kinase inhibitor that is pivotal in arresting cellular growth in terminal cell differentiation and apoptosis. Thus, the existence of natural variants of p21(Waf1/Cip1) could be linked to specific cancer. The purpose of this report was to identify a novel tri-allelic insertion/deletion (INDEL) polymorphism (rs4135235) involving a poly-T sequence in the promoter region of p21(Waf1/Cip1) gene and to explore its role in gastric cancer (GC).

Method: A total of unrelated 676 subjects (376 GC patients; 300 cancer-free controls) were enrolled in the study, and genomic DNA was obtained from each subject for genotyping. PCR-directed sequencing technique was used to detect the genotypes of the polymorphism. TA cloning was used to confirm the existence of three alleles. Odds ratios (ORs) and 95% confidence intervals (CIs) were estimated by unconditional logistic regression analysis.

Results: Six genotypes (9T/9T, 10T/10T, 11T/11T, 9T/10T, 10T/11T, and 9T/11T) and three alleles (9Ts, 10Ts, and 11Ts) were identified among all study subjects. GC cases were different from the control group with over-representation of 9T/11T heterozygotes (19.7% vs. 12.3%) and under-representation of 10T/10T homozygotes (18.4% vs. 20.7%). Compared with those carrying 10T/10T, individuals with 9T/11T increased the susceptibility for GC (OR=1.797, 95%CI=1.065-3.031).

Conclusion: Our findings confirmed the existence of a tri-allelic polymorphism in the promoter of p21(Waf1/Cip1). It has also shown the heterozygous genotype 9T/11T to be a potential risk factor for GC in the Chinese population.

FIG. 1.

Sequencing chromatograms of p21-1933 INDEL polymorphism by PCR direct sequencing. All sequencing chromatograms could be classified into two groups: Group 1 containing three homozygotes with clear and single peak and Group 2 containing three heterozygotes with double peaks. (A) The homozygous sequence of 9T/9T. (B) The homozygous sequence of 10T/10T. (C) The homozygous sequence of 11T/11T. (D) The heterozygous sequence of 9T/10T. (E) The heterozygous sequence of 10T/11T. (F) The heterozygous sequence of 9T/11T.

FIG. 2.

Analysis for variation by DHPLC. Three homozygous genotypes were one-peaked. Three heterozygous genotypes were two-peaked. DHPLC, denaturing high-performance liquid chromatography.