and 4.8 ± 0.5%, respectively) in comparison with negative control (94.3 ± 1.5%, viable cells; 1.7 ± 0.9%, early apoptosis and 1.5 ± 0.2%, late apoptosis) (p < 0.05) ( Fig. 5B). Similarly, Dox also caused a significant cell viability decreasing (16.1 ± 0.1%) and early apoptosis rising (83.2 ± 0.1%). Another early marker of the apoptotic process is the depletion of mitochondria membrane potential. In this work, none of the compounds evaluated in 24 h of treatment significantly alter the mitochondrial membrane potential (p > 0.05), suggesting that only the extrinsic pathway was activated within 24 h. However, in 48 h exposure, compound 4 (2 μM) caused depolarization of mitochondrial membrane potential (37.3 ± 4.6%, Fig. 5C) when compared to negative control (4.7 ± 0.6%, p < 0.05). Dox, positive control, cause Selleckchem Antidiabetic Compound Library www.selleckchem.com/products/ldk378.html intense membrane depolarization after 24 h (44.0 ± 2.3%) and 48 h (46.9 ± 5.4%) of incubation. The DNA damage induced by the α-santonin derivatives was evaluated in human mononuclear cells. DNA damages were not detected with the concentrations tested (p > 0.05, data not shown). Sesquiterpene lactones (SLs) are plant-derived compounds often used in traditional medicine against several human diseases such as inflammation and cancer (Ghantous et al., 2010). Previous researches showed no cytotoxic activity of the α-santonin molecule, even at high concentrations (100 μM) (Kim et al., 2002 and Konaklieva ASK1 and Plotkin,
2005). Then, we designed three cytotoxic sesquiterpene lactones based on α-santonin (Arantes et al., 2009; 2010) with activity on different cancer cell lines and low toxicity on PBMC. In this work, we propose the mechanism responsible for this cytotoxicity using the HL-60 cell line as experimental model and the compounds tested (1 and 2 μM) after 24 h of treatment. Initially, we showed that the antiproliferative potential of the α-santonin derivatives is not related to direct
membrane damages, since the trypan and propidium iodide exclusion techniques did reveal membrane permeability of remaining cells. In fact, it is possible that apoptosis or other process might have already compromised cell proliferation, but membrane integrity is still maintained (Kepp et al., 2011). We previously reported that these derivatives did not produce cell membrane disruption of mouse erythrocytes (Arantes et al., 2010). Some studies have been pointed that SLs inhibit tumor growth by selective alkylation of growth-regulatory biological macromolecules, such as DNA and key enzymes, which control cell division, thereby inhibiting a variety of cellular functions, which leads cells into apoptotic death (Fernandes et al., 2008 and Rozenblat et al., 2008). Herein, all molecules reduced BrdU incorporation by HL-60 treated cells, suggesting inhibition of DNA synthesis. Other SLs caused inhibition of DNA synthesis by BrdU test such as enhydin, uvedalin and sonchifolin (Siriwan et al., 2011).