Cantharidin Inhibit The Growth And Invasion Ability Of Pancreatic Cancer Cell

Objective To find out the effect of Cantharidin on the growth ability of pancreatic cancer cell line PANC-1 and CFPAC-1. Methods Cell proliferation was determined by MTT assay. Clone formation ability was determined by clony forming assay. Cell cycle and apoptosis were tested by flow cytometry. Gene expressions were tested using RT-PCR and Western blot. Results The inhibition on PANC-1 and CFPAC-1 growth was increased with cantharidin concentration and time course. After PANC-1 was administered with cantharidin, the clone formation ability significantly decreased from 61.00% to 28.00%(P<0.01). After CFPAC-1 was administered with cantharidin, the clone formation ability significantly decreased from 38.00% to 10.00%(P<0.01).After pancreatic cancer cells was administered with 10μmol/L cantharidin for 48h, the ratio of PANC-1 G2/M phase significantly increased from (16.47?1.66%)to(42.71?3.64%)(P<0.01),the ratio of CFPAC-1 G2/M phase significantly increased from (15.30?0.72%)to(25.26?3.14%)(P<0.01).After PANC-1 was administered with 10μmol/L cantharidin for 24h, the apoptosis rate of experimental group(24.89?4.80%) was more higher than the untreated group(7.35?2.44%)(P<0.01). After CFPAC-1 was administered with 10μmol/L cantharidin for 24h, the apoptosis rate of experimental group(24.70 ?1.92%) was more higher than the untreated group(6.39?3.95%)(P<0.01).These results showed that cantharidin caused G2/M cell-cycle arrest and apoptosis of PANC-1.. Cantharidin caused G2/M cell-cycle arrest that was accompanied by the down-regulation of CDK1 and up-regulation of p21 expression. It induced apoptosis and elevated the expressions of pro-apoptotic factors, TNF-α, TRAILR1, TRAILR2, Bad, Bak, and Bid, and decreased the expression of anti-apoptotic Bcl-2.Conclusion Cantharidin can significantly inhibit the growth of pancreatic cancer cells, making it an attractive candidate compound for pancreatic cancer therapy. Objective: We have found that Cantharidin as PP2A inhibitor ,inhibited pancreatic cancer cell growth, although the mechanism involved is still unclear. PP2A (protein phosphatase 2A) is a multimeric serine/threonine phosphatase which can dephosphorylate multiple kinases. IKK (IκB kinase), the key regulator of NF-κB pathway, is a direct substrate of PP2A. As activation of NF-κB pathway can trigger apoptosis in some circumstance, we tried to investigate whether Cantharidin could induce apoptosis in pancreatic cancer cells through activation of NF-κB pathway.Methods: Proliferation was determined by MTT. Apoptosis was tested by flow cytometry. Gene expressions were tested using RT-PCR and Western blot. Activation of NF-κB pathway was measured using Western blot and luciferase reporter gene assay. Results: Treatment with Cantharidin induced activation of NF-κB pathway through phosphorylation of IKKα, phosphorylation and degradation of IκBα, and nuclear translocation of p65. Using dominant negative -IKKα, dominant negative -IκBα, Bay 11-7082 (an inhibitor of IκBαphosphorylation) or p65 specific shRNA, we could not only block the activation of NF-κB pathway, but also attenuate the cytotoxicity of PP2A inhibitors, suggesting the anti-cancer effect of Cantharidin is executed in a NF-κB pathway dependent manner. We further find out that the transcriptional activation of NF-κB pathway increased the expression of downstream pro-apoptotic genes, TNF-α, TRAILR1, and TRAILR2. Conclusion: We provided convincing evidence that Cantharidin induced cytotoxic effects against pancreatic cancer cells through persistent activation of the NF-κB pathway.