Anticancer effect of two diterpenoid compounds isolated from Annona glabra Linn

To study the inhibitory effect of two diterpenoid compounds isolated from Annona glabra Linn (Cunabic acid and ent-kauran-19-al-17-oic acid) on the proliferation of Human Liver Cancer (HLC) cell line SMMC-7721 and its mechanism.

Inhibition of cell proliferation was measured by MTT assay. The morphological changes of SMMC-7721 cells were observed under inverted phase-contrast microscope, fluorescent microscope, transmission electron microscope (TEM), and scanning electron microscope (SEM). Flow cytometer (FCM) was used to calculate the cell apoptotic rate, and immunohistochemical staining was used to observe the regulation of gene expression.

The proliferation of SMMC-7721 cells was obviously inhibited after being treated with Cunabic acid at the concentration great than 5 micromol/L and ent-kauran-19-al-17-oic acid great than 10 micromol/L. The biggest inhibitory effect was 81.05% when treated with Cunabic acid at the concentration of 25 micromol/L. The effect had a linear relationship with concentration. The result indicated that drug-treated cells exhibit typical morphological changes of apoptosis, including condensed chromatin and a reduction in volume. Sub-G0/G1 peak was found by FCM analysis and the cell cycle was arrested at G0/G1 stage. The apoptotic rates of the cells treated by Cunabic acid and ent-kauran-19-al-17-oic acid were 43.31% and 24.95%, respectively. It was visualized by immunohistochemical staining that the drugs down-regulated the gene expression of bcl-2 gene and up-regulated that of bax gene.

The two diterpenoid compounds isolated from Annona glabra Linn, Cunabic acid and ent-kauran-19-al-17-oic acid can obviously inhibit the proliferation of HLC cell line SMMC-7721. The mechanism is correlated with the induction of cell apoptosis by down-regulating the gene expression of bcl-2 gene and up-regulating that of bax gene.