Calophyllum brasiliense Extracts Induced Apoptosis in Human Breast Adenocarcinoma Cells

Abstract

Aims: Apoptosis, or programmed cell death, is linked to several mechanisms of cell growth control. The present work aimed at evaluating the induction of apoptosis in MCF-7 human breast adenocarcinoma cell by Calophyllum brasiliense. Study design: The tests were performed in triplicates in the apoptosis assays and sextuplicates in the cytotoxic assays, to each group, and the data expressed the mean +/- standard deviations. The cytotoxicity IC50s were obtained based on nonlinear regression curve fit. Two-way ANOVA and Tukey’s tests were applied in the apoptosis analyses. Place and duration of study: The work was done at the Center for Research in Biodiversity (Cell Culture Laboratory, and Phytochemistry Laboratory), and Research Center (Molecular Biology Laboratory), Paulista University, between Jan 2019 and Dec 2019. Methodology: Two aqueous extracts, obtained from the stem (STE) and from the leaves (LFE) of Calophyllum brasiliense by a 24-h maceration, were submitted to a cytotoxic assay against MCF-7 breast cancer cell lines at the concentrations of 0.01 µg/ml, 0.1 µg/ml, 1.0 µg/ml, 10 µg/ml and 100 µg/ml. They were also subjected to the evaluation of apoptosis and necrosis cell death induction at concentrations of 50, 100 and 200 μg/ml, after 6 h, 12 h and 24 h. Curcumin was used as a reference drug for both cytotoxic (50 mM, 5.0 mM, 0.5 mM, 0.05 mM and 0.005 mM ) and apoptosis/necrosis (12.5 μM, 25 μM and 50 μM / 6 h, 12 h and 24 h) assays. Apoptosis and necrosis were accessed by the use of annexin V and 7-AAD, in a two-channel flow cytometer. Results: In terms of the cytotoxic activity, STE (IC50 7.86 µg/ml) was more toxic than LFE (IC50 74.35 µg/ml), and curcumin IC50 was 0.00159 µg/ml. STE induced 21.19 % and LFE, 20.63 %, in comparison to 13.4% of apoptosis induction by curcumin. The results of apoptosis induction in the cancer cells were achieved at 24 h, extract concentrations at 100 µg/ml. Conclusion: Both the extracts, STE and LFE, were cytotoxic against MCF-7 breast cancer cell line, and induced more apoptosis in MCF-7 cells than curcumin, suggesting that they are high potential sources of natural product-inducing apoptosis agents to be used against adenocarcinoma breast cells.