Gambogic Acid Induces Cell Apoptosis and Inhibits MAPK Pathway in PTEN/p53Prostate Cancer Cells In Vitro and Ex Vivo.
- Author:
Hong PAN
1
;
Li-Yuan LU
2
;
Xue-Qian WANG
2
;
Bin-Xue LI
2
;
Kathleen KELLY
3
;
Hong-Sheng LIN
4
Author Information
- Publication Type:Journal Article
- Keywords: PTEN−/−/p53−/−; apoptosis; gambogic acid; mitogen-activated protein kinase; prostate cancer
- From: Chinese journal of integrative medicine 2018;24(2):109-116
- CountryChina
- Language:English
-
Abstract:
OBJECTIVETo investigate the effect of gambogic acid (GA) on the growth and cell death of castrate resistant prostate cancer (PC) with phosphate and tension homology (PTEN) and p53 genes deleted in vitro and ex vivo, and elucidate the underlying possible molecular mechanisms.
METHODSPTEN/p53PC cells and Los Angeles prostate cancer-4 (LAPC-4) cells were treated with GA for 24 h and 48 h, then cell viability was determined by cell proliferation assay. PTEN/p53PC cells organoids number was calculated under GA treatment for 1 week. In addition, cell titer glo assay was performed to analyze 3 dimensional cell viability of patients derived xenografts (PDX) 170.2 organoids. Flow cytometry was used to detect apoptotic cells treated with GA. And confocal image was performed to detect the apoptotic mitochondrial morphological changes. Apoptotic cell death related protein levels were measured through Western blot (WB) in GA treated cells and organoids. The expression levels of mitogen-activated protein kinases (MAPKs) pathway related ribonucleic acid (RNAs) and proteins were analyzed by reverse transcription polymerase chain reaction (RT-PCR) and WB, respectively.
RESULTSThe treatment of GA significantly reduced cell viability of PTEN/p53PC cells and LAPC-4 in a time- and concentration-dependent manner. In organoids, GA showed strong inhibition towards organoids' numbers and diameters and continuously led to a complete organoids inhibition with GA 150 nmol/L. Ex vivo results validated that GA 1 μmol/L inhibited 44.6% PDX170.2 organoids growth. As for mechanism, flow cytometry detected continuously increased apoptotic portion under GA treatment from 1.98% to 11.78% (6 h) and 29.94% (8 h, P<0.05). In addition, mitochondrial fragmentation emerged in GA treated cells indicated the mitochondrial apoptotic pathway might be involved. Furthermore, WB detected caspases-3, -9 activation and light chain (LC)-3 conversion with GA treatment. WB revealed decreased activity of MAPK pathway and down-regulation of downstream c-fos oncogene RNA level was detected by RT-PCR before undergoing apoptosis (P<0.05).
CONCLUSIONGA was a potent anti-tumor compound as for PTEN/p53PC, which contributed to cell apoptosis via inhibition of the MAPK pathway and c-fos.