The apoptosis-inducing effect of the
triterpene saponins, namely,
ursolic acid and its natural derivative,
methyl-ursolate beta-D-
glucoside on A431 human
epidermoid carcinoma cells was studied. The cells treated with 5-50 microg/ml of
ursolic acid resulted in a dose- and time-dependent decrease in cell number, due to an increase of apoptotic cells as evidenced by MTT assay together with morphological changes. The highest dose (50 microg/ml) of
ursolic acid resulted in approximately 90% inhibition in
tumor cell growth after 96 hours of treatment and 60% of apoptosis after 48 hours. To the contrary, when the same treatment was carried out with
methyl-ursolate beta-D-
glucoside, after 96 hours of treatment the percentage of cell growth inhibition was found to be only 30% at the dose of 50 microg/ml and the value of apoptosis did not exceed 10%. Similarly to these results,
ursolic acid effectively induced proteolytic activation of
caspase-3 protease in a dose-dependent manner while its derivative showed only weak activity in this
enzyme assay. The addition of
DEVD-CHO prior to
ursolic acid and
methyl-ursolate beta-D-
glucoside treatment effectively prevented the loss of
triterpenes-induced viability. In summary, the
triterpene saponins investigated contain an apoptotic-inducing activity in A431 cells and in the case of
ursolic acid it is associated with proteolytic activation of
caspase-3 and/or other similar
caspases. Our results also indicated that methylation of COOH-28 together with the glycosylation of C3 of
ursolic acid have a strong impact on its antitumor activity.