Unconjugated
Pancreatic cancer remains the fourth most common cause of cancer-related death in the United States. Potent therapeutic strategies are urgently needed for pancreatic cancer. Cucurmosin is a novel type 1 ribosome-inactivating protein (RIP) isolated from the sarcocarp of Cucurbita moschata (pumpkin). Due to its cytotoxicity, cucurmosin can inhibit tumor cell proliferation through induction of apoptosis on tumor cells, but the specific mechanism is still unclear. We explored the function of cucurmosin in BxPC-3 pancreatic cancer cells using multiple cellular and molecular approaches such as 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry, reverse transcription polymerase chain reaction (RT-PCR), Western blotting and transmission electron microscopy for observing typical changes and formation of apoptotic bodies. We found that cucurmosin inhibited the proliferation of BxPC-3 cells in a time- and dose-dependent manner, and increased the cell population in the G0-G1 phase. With increasing concentration of cucurmosin, the expression of EGFR, p-PI3K, Akt, p-Akt, mTOR, p-mTOR, P70S6K-α, p-P70S6K-α, 4E-BP1 and p-4E-BP1 at the protein level was decreased, whereas the expression of p-Bad and caspase-9 was elevated. However, the mRNA expression of EGFR did not change. These findings suggest that cucurmosin can down-regulate the expression of EGFR by targeting. Cucurmosin induces the apoptosis of BxPC-3 pancreatic cancer cells via the PI3K/Akt/mTOR signaling pathway.
BACKGROUND:
Pancreatic cancer treatment is limited and effective drugs are needed. We investigated cucurmosin (CUS)-induced apoptosis in cystic fibrosis pancreatic adenocarcinoma cells (CFPAC-1) and a possible mechanism of action to evaluate the clinical application potential of this new Type I ribosome-inactivating protein.
METHODS:
We analyzed the growth inhibition and apoptosis of CFPAC-1 cells via methylthiazol tetrazolium assay and fluorescence-activated cell sorting. Western blot was used to analyze the protein levels of caspase 3, bcl-2, caspase 9, platelet-derived growth factor receptor (PDGFR)-β, PI3K, Akt, p-Akt, the mammalian target of rapamycin (mTOR), p-mTOR, P70S6K-α, p-P70S6K-α, 4E-BP1, p-4E-BP1 and p-Bad after CUS intervention. The mRNA expression of PDGFR-β was analyzed using reverse transcription polymerase chain reaction.
RESULTS:
CUS inhibited the proliferation of pancreatic cancer cells. The induction of apoptosis depended on the CUS dose and incubation time. The drug inhibited all of the examined proteins in the PI3K/Akt/mTOR signalling pathway and induced the active fragments of caspase 3 and caspase 9. CUS downregulated PDGFR-β expression but no significant change was observed at the mRNA level.
CONCLUSION:
CUS strongly inhibits the growth of CFPAC-1 by inducing cell apoptosis. CUS downregulated the expression of PDGFR-β at the protein level and induced the apoptosis of CFPAC-1 through the PI3K/Akt/mTOR signalling pathway.
