The apoptotic potential of the most active compounds was investigated and they were able to induce apoptosis being compound 12 the best inducer. The caspase-3, -8 and -9 activities were measured. The results obtained showed that compounds 12, 21 and 23 induce apoptosis via the activation of caspase-8, whereas compound 20 induces apoptosis via caspase-9.
Inmunoblot analysis of the expression of p53, Bax, Bcl-2, Bcl-x1 and IAPs in RAW 264.7 cells was also carried out. When cells were exposed to 5 μM of the different compounds, expression levels of p53 and Bax increased whereas levels of antiapoptotic proteins such as Bcl-2, Bcl-x1 and IAPs decreased.
In conclusion, kaurene derivatives (12, 20, 21 and 23) induce apoptosis via both the mitochondrial and membrane death receptor pathways, involving the Bcl-2 family proteins. Taken together these results provide a role of kaurene derivatives as apoptotic inducers in tumor cells and suggest their potential application as antitumor agents.