Abstract B14: Characterization of p53-HDM2 inhibitor in nucleotide excision repair deficient cell line

Abstract

Abstract The ubiquitin (Ub)/proteasome system has a special role in the control on the cellular concentration of p53 tumor suppressor, a master regulator of cell survival and death. Over the last years, many groups have elucidated the mechanisms of Ub ligase–mediated p53 ubiquitylation under various conditions. One pivotal regulatory mechanism is HDM2-mediated p53 degradation, but it can also export p53 from nucleus impairing its transcriptional activity or by binding p53 transactivation domain, that inhibits p53 transcriptional activity. Rad23 can directly bind Ub and the proteasome and has been shown to act as an adaptor that shuttles the ubiquitylated proteins to the proteasome. Rad23 is required for the degradation of p53 and one possible helper of Rad23 is its binding protein XPC, a key factor for nucleotide excision repair (NER). Mutations in XP proteins can cause xeroderma pigmentosum (XP), a genetic disease that often leads to high rates of skin cancer. Pathogenic mutations in p53 are found in more than 50% of all human malignancies, which makes p53 the most frequently mutated gene in transformed cells in humans. In cancers in which the p53 gene is not mutated, the function of the p53 pathway is often suppressed through overexpression or deregulation of HDM2. We aimed to study the impact of HDM2-P53 inhibition in XPC-deficient fibroblasts by treating these cells with the drug CGM9097, which is a potent inhibitor of HDM2-p53 protein-protein interaction used in preclinical trials. In this poster we present preliminary data. We firstly evaluated CGM9097 cytotoxicity in XPC deficient and corrected cells, combined with 5-fluorouracil (5FU) treatment. XPC deficient cells were more sensitive to 5FU+ CGM9097 than normal fibroblast (MRC5) and correct fibroblast (XPC+ XPC gene) (P<0,05). Protein analysis by Western blotting showed dose-response expression of p53 in both lineages when treated with 5FU, with the higher expression level within 6 h. When cells were treated with 5FU+ CGM9097, XPC-deficient cells presented high levels of p53 even in the untreated fraction compared to corrected cell line, suggesting an impairment of p53 degradation in the absence of XPC. These preliminary data suggest that XPC has an important role in the cytotoxicity of 5FU in cells with impairment of p53. P53 mutant cancers are sensitive to some chemotherapeutic drugs, but the reported sensitivity is generally modest (2-3 fold), and this can be explained by the fact that p53 affects the GGR NER sub pathway through GADD45. In addition, p53-mediated DNA repair may be counterbalanced by the opposing response of apoptosis, also regulated in part by p53. Thus, there appears to be much potential to exploit DNA repair defects in p53 mutant cancers. Citation Format: Douglas Felipe de Lima Silva, Ana Rafaela de Souza Timoteo, Lucymara Fassarela Agnez Lima, Tirzah Braz Petta Lajus. Characterization of p53-HDM2 inhibitor in nucleotide excision repair deficient cell line [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B14.