Clinicopathologic Significance Of Structural Alterations Of P16 Protein In Esophageal Squamous Cell Carcinoma

Objective: The purpose of this study was to intensively investigate the clinicopathologic significance of the alternations of p16 protein three-dimensional structure in human esophageal squamous cell carcinoma (HSSC) and their relation with the p16 mRNA expression by Comparative RT-PCR so as to provide a reliable indicator and create a new way for the research on clinical prevention and treatment of HSSC.Methods: Polymerase chain reaction single strand comformational polymorphism (PCR-SSCP), DNA sequencing, computerized three-dimensional protein-modeling, reverse transcription polymerase chain reaction (RT-PCR), and computerized image analysis were utilized.Results: 1. Thirty-three cases with p16 gene abnormality including sixteen point mutations and seventeen deletions, which were defined as Group I , were detected in sixty-nine cases of HSSC. No p16 gene abnormality was found in the rest of thirty-six cases, which were classified as Group II. There were significant differences in lymph node metastasis and distance metastasis between these groups (P<0.05). However, no significant differences in sex, age, size of tumor, invasion's depth of tumor (P>0.05) were found. 2. In the twenty-six cases out of thirty-three HSSCs which showed p16 gene mutation (Group I ), the alternations of the amino acid residues were located in the p16-protein domains. We classified this subgroup as Group I b. The rest of seven cases with their p16 mutations beyond the domains and far from thep16-CDK4/6 binding site, were defined as Group I a. The significant differences in lymph node metastasis, distance metastasis and stage of clinical (P<0.05) were found between the two subgroups.3. The semi-quantitative results of p16 mRNA analyzed by the paired t-test between twenty-six HSSC and normal tissues revealed that the level of p16 mRNA expression in cancer was significantly decreased in cancer tissues (P<0.01). The analysis of p16 expression level in 11 HSSC samples, which is abnormal in p16 gene, showed the tendency of decreased p16 expression in group I b compared to group I a.Conclusion: The mutations of p16 gene will alter the formation of p16 protein. The four ankyrin repeats are the critical regions of p16-protein. The abnormalities in the ankyrin repeats will seriously alter the 3D structure of p16 protein and affect the activity of p16 protein, resulting in lymph node metastasis, distance metastasis, and clinically advanced stage. They render an authentic evidence for the selection of the clinic cases with high risk of metastasis.