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

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

Simposio; COST ACTION CM1407; 2017

Cancer still remains a major cause of disease anddeath worldwide and thus new therapeutic options are a high priority for mostof researchers. The genus Melia is well-known as a rich and valuable source of bioactivetetranortriterpenoids known as limonoids. Inprevious research conducted by our group, the fractionationof a kernel extract obtained from Meliaazedarach L. led to the isolation of meliartenin (1), anantifeedant limonoid, which exists as a mixture with its tautomeric isomer12-hydroxyamoorastatin [1].In this study, the antitumor activity of 1 was evaluated over a panel of humantumor-derived cell lines, resulting in a cytotoxic effect that was highlyselective of the human colorectal carcinoma cell line HCT116 over the rest ofthe assayed cell lines. Mechanistic analysis revealed that treatment with 1 induced a time-dependent increase inthe number of cells transiting the S-phase of the cell cycle, with a similaractivity from 0.4 uM to that obtained with the positive control hydroxyurea(HU). Subsequent detection with an anti-BrdU antibody confirmed a sustainedanti-proliferative effect on HCT116 cells. Inaddition to slowing down the cellular progression through S-phase, flowcytometry analysis after SYTOX Red stain showed that 1 induced cell death in a time-dependent manner, reachingpercentages of death higher than 53% from 0.2 μM at 72h. Furthermore, Annexin V/SYTOX Red double staining demonstrated that thiscompound was able to induce apoptosis in HCT116 cells with values higher than70% at 72 h. It was also found that 1 was not only capable of killingHCT116 but also strongly impaired its clonogenic potential at a concentrationas low as 0.04 uM. In the light of our findings weevaluated whether the biological effects mediated by 1 were related to the induction and activity of p53 and itsdownstream target p21. The increased expression profile of these proteins wassimilar to the treatment with HU. It was found that p53-dependent apoptosis was responsible for thecytotoxicity observed in HCT116. The fact that p53, but not p21, waslinked to the cytotoxic effect mediated by this compound strongly suggests that only thepro-apoptotic functions of p53 are critical for this effect. Taken together, theresults described in this work position compound 1 as a strong antitumoral agent, with great potential for targetingp53+ tumors (Lorenc:2017kw)