Abstract 2604: Functional analysis of p53 codon 72 polymorphism with cysteine substitution in cancer cells

Abstract

Abstract The p53 codon p72 polymorphism with proline (p53P72)/arginine (p53R72) substitution is associated with cancer incidence. Cysteine(C) substitution at p53 codon 72 polymorphism (p53C72) is rarely studied in cancer patients. A patient who developed AML and who carries p53C72 was described (El-Deiry, 2022 WIN Symposium). The patient’s family has high rates of cancer susceptibility in the women some of whom carry FANCC alterations and other alleles. To examine the functional characteristics of different p53 alterations at codon 72, we generated plasmids expressing p53C72, p53 R72 or p53P72. The variants of p53 were overexpressed in p53-null cancer cells by transient transfection of the plasmids. We tested functions of the p53 codon 72 polymorphisms (p53C72, p53R72 and p53P72) in two cancer cell lines (Soas-2 and HCT116 p53 null) with different expression levels of the p53 variants at different time points. To examine the effect of codon 73 variants on p53 transactivation, we performed a PG13-Luciferase reporter assay and examined the endogenous p53 targets at the protein level. The luciferase-reporter assay showed that all the three variants of the p53 codon p72 variants increased PG-13 luciferase expression at early time points. The Western blot assay showed that the p53C72, the p53R73 and the p53P72 increased endogenous p21, DR5, puma and MDM2 at the protein levels at different time points. The p53C72 increased the p53 targets and PG13-driven luciferase expression much higher than the p53R72 and the p53P72. These results suggest that the p53 codon 72 variants (C72, R72 and P72) retain wild-type activity for p53-mediated transcription, and p53C72 has the highest potency among them. We examined the effect of the p53 variants on cell growth and death. PARP cleavage (a marker of cell apoptosis) was induced in cells overexpressing all the three p53 codon 72 variants. We observed a lesser potency of p53C72 to induce PARP cleavage compared to p53R72 and p53P72. Colony formation assay showed a suppressive effect of p53C72, p53R72 and p53P72 on cancer cell growth. Our studies demonstrate some differences of in wild-type functions of p53 p53C72, p53R72 and p53P72 alleles. Citation Format: Shengliang Zhang, Wafik S. El-Deiry. Functional analysis of p53 codon 72 polymorphism with cysteine substitution in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2604.