P53 tumor suppresor is required for efficient execution of the death program following treatment with a cytotoxic limonoid obtained from Melia azedarach

GONZALEZ MARÍA LAURA; PALACIOS, SARA M.; CARPINELLA MARÍA CECILIA; VILLAFAÑEZ, FLORENCIA; LAIOLO JERÓNIMO; SORIA, GASTÓN; JORAY MARIANA BELÉN; CRESPO, MARÍA INÉS; BOCCO, JOSÉ LUIS

Lisboa

Simposio; COST ACTION CM1407; 2017

Cancer still remains a major cause of disease and death worldwide and thus new therapeutic options are a high priority for most of researchers. The genus Melia is well-known as a rich and valuable source of bioactive tetranortriterpenoids known as limonoids. In previous research conducted by our group, the fractionation of a kernel extract obtained from Melia azedarach L. led to the isolation of meliartenin (1), an antifeedant limonoid, which exists as a mixture with its tautomeric isomer 12-hydroxyamoorastatin [1].In this study, the antitumor activity of 1 was evaluated over a panel of human tumor-derived cell lines, resulting in a cytotoxic effect that was highly selective of the human colorectal carcinoma cell line HCT116 over the rest of the assayed cell lines. Mechanistic analysis revealed that treatment with 1 induced a time-dependent increase in the number of cells transiting the S-phase of the cell cycle, with a similar activity from 0.4 uM to that obtained with the positive control hydroxyurea (HU). Subsequent detection with an anti-BrdU antibody confirmed a sustained anti-proliferative effect on HCT116 cells. In addition to slowing down the cellular progression through S-phase, flow cytometry analysis after SYTOX Red stain showed that 1 induced cell death in a time-dependent manner, reaching percentages of death higher than 53% from 0.2 μM at 72 h. Furthermore, Annexin V/SYTOX Red double staining demonstrated that this compound was able to induce apoptosis in HCT116 cells with values higher than 70% at 72 h. It was also found that 1 was not only capable of killing HCT116 but also strongly impaired its clonogenic potential at a concentration as low as 0.04 uM. In the light of our findings we evaluated whether the biological effects mediated by 1 were related to the induction and activity of p53 and its downstream target p21. The increased expression profile of these proteins was similar to the treatment with HU. It was found that p53-dependent apoptosis was responsible for the cytotoxicity observed in HCT116. The fact that p53, but not p21, was linked to the cytotoxic effect mediated by this compound strongly suggests that only the pro-apoptotic functions of p53 are critical for this effect. Taken together, the results described in this work position compound 1 as a strong antitumoral agent, with great potential for targeting p53+ tumors{Lorenc:2017kw}.