Anticancer effect of two diterpenoid compounds isolated from Annona glabra Linn.
OBJECTIVE
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Zhang
Yong-hong
YH
First Clinical College, Nanjing University of Traditional Chinese Medicine, Nanjing 210029, China. zyh19511227@sohu.com.
Peng
Hai-yan
HY
Xia
Guo-hao
GH
Wang
Ming-yan
MY
Han
Ying
Y
D031041:Annona / Q000737:chemistry*; D000972:Antineoplastic Agents, Phytogenic / Q000302:isolation & purification / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D002453:Cell Cycle; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D004224:Diterpenes / Q000302:isolation & purification / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D019254:Genes, bcl-2; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology; D010946:Plants, Medicinal / Q000737:chemistry; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein
D000972:Antineoplastic Agents, Phytogenic; C494826:BAX protein, human; D004224:Diterpenes; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; C488835:cunabic acid; C488834:ent-kauran-19-al-17-oic acid