cp-31398 and Endometrial-Neoplasms

Mutation in the tumor suppressor gene p53 is the most frequent molecular defect in endometrial carcinoma (EC). Recently, CP-31398, a p53-stabilizing compound, has been indicated to possess the ability to alter the expression of non-p53 target genes in addition to p53 downstream genes in tumor cells. Herein, we explore the alternative mechanisms underlying the restoration of EC tumor suppressor function in mutant p53 by CP-31398. A p53-mutated EC cell was constructed in AN3CA cells with restored or partial loss of Slug using lentiviral vectors, followed by treatment with 25 μM CP-31398. A p53-independent mechanism of CP-31398 was confirmed by the interaction between mouse double minute 2 homolog (MDM2) and Slug AN3CA cells treated with IWR-1 (inhibitor of Wnt response 1). Furthermore, the AN3CA cells were treated with short hairpin RNA against Slug, Wnt-specific activators (LiCl) or inhibitors (XAV-939) followed by CP-31398 treatment. Moreover, AN3CA cell proliferation and apoptosis were examined. A tumorigenicity assay was conducted in nude mice. CP-31398 could promote the apoptosis of p53-mutated EC cells, while Slug reversed this effect. Slug ubiquitination was found to occur via binding of Slug to MDM2 in AN3CA cells. We found that CP-31398 increased the GSK-3ß, p-Slug, Puma, Wtp53, and Bax expressions whereas Wnt, Mtp-53, Slug, Bcl-2, and Ki-67 expressions were decreased. However, these findings were reversed following the activation of the Wnt pathway and overexpression of Slug. Finally, the in vivo experimental evidence confirmed that CP-31398 with depleted Slug suppressed tumor growth by downregulating the Slug. Collectively, CP-31398-regulated Slug downregulation represses the p53-mutated EC via the p53/Wnt/Puma pathway.

Topics: Animals; Apoptosis; Cell Line, Tumor; Cell Proliferation; Endometrial Neoplasms; Female; Humans; Mice, Nude; Proto-Oncogene Proteins c-mdm2; Pyrimidines; Tumor Suppressor Protein p53

Endometrial cancer (EC) is one of the most common malignancies of the female reproductive system, and metastasis is a major cause of mortality. In this study, we aimed to explore the role of CP‑31398 in the migration, invasion and apoptosis of EC cells by its regulation of the expression of the murine double minute 2 (MDM2) gene. For this purpose, EC tissues and adjacent normal tissues were collected, and the positive expression rate of MDM2 in these tissues was assessed. Subsequently, the cellular 50% inhibitory concentration (IC50) of CP‑31398 was measured. The EC RL95‑2 and KLE cell lines had a higher MDM2 expression and were thus selected for use in subsequent experiments. The EC cells were then treated with CP‑31398 (2 µg/ml), and were transfected with siRNA against MDM2 or an MDM2 overexpression plasmid in order to examine the effects of CP‑31398 and MDM2 on EC cell activities. The expression of p53, p21, Bad, Bax, B‑cell lymphoma‑2 (Bcl‑2), cytochrome c (Cyt‑c), caspase‑3, Cox‑2, matrix metalloproteinase (MMP)‑2 and MMP‑9 was measured to further confirm the effects of CP‑31398 on cell migration, invasion and apoptosis. Our results indicated that MDM2 was highly expressed in EC tissues. Notably, EC cell viability decreased with the increasing concentrations of CP‑31398. The EC cells treated with CP‑31398 or siRNA against MDM2 exhibited an increased apoptosis and a suppressed migration and invasion, corresponding to an increased expression of p53, p21, Bad, Bax, Cyt‑c and caspase‑3, as well as to a decreased expression of Bcl‑2, Cox‑2, MMP‑2 and MMP‑9. Moreover, treatment with CP‑31398 and siRNA against MDM2 further enhanced these effects. Taken together, the findings of this study indicate that the CP‑31398‑mediated downregulation of MDM2 may suppress EC progression via its inhibitory role in EC cell migration, invasion and resistance to apoptosis. Therefore, treatment with CP‑31398 may prove to be possible therapeutic strategy for EC.

Topics: Adult; Aged; Cell Line, Tumor; Cell Movement; Cell Survival; Down-Regulation; Endometrial Neoplasms; Female; Gene Expression Regulation, Neoplastic; Humans; Inhibitory Concentration 50; Middle Aged; Neoplasm Invasiveness; Proto-Oncogene Proteins c-mdm2; Pyrimidines; RNA, Messenger; RNA, Small Interfering