Abstract
Mesothelin, a secreted protein, is overexpressed in some cancers, including pancreatic cancer. Rescent studies have shown that overexpression of mesothelin significantly increased tumor cell proliferation, and downregulation of mesothelin inhibited cell proliferation in pancreatic cancer cells, but its exact function and mechanism remains unclear. The aim of the present study was to evaluate the effects of mesothelin on proliferation and apoptosis in pancreatic cancer cells with different p53 status and to explore its signal pathway. Mesothelin levels were detected by western blot and RT-PCR assay in human pancreatic cancer AsPC-1, HPAC and Capan-2, Capan-1 and MIA PaCa-2 cell lines. Mesothelin was slienced by shRNA in AsPC-1, Capan-2 and Capan-1 cells with rich mesothelin level, and mesothelin was overexpressed in the HPAC and Capan-2 cells with less mesothelin level. We observed that in the AsPC-1 and Capan-1cells with mt-p53, and Capan-2 cells with wt-p53, shRNA mediated sliencing of the mesothelin significantly increased PUMA and Bax expression and caspase-3 activity, and decreased bcl-2 expression, followed by the reduced proliferation and colony forming capability and increased cell apoptosis. When PUMA was slienced by siRNA in the stable mesothelin shRNA transfected cells, proliferative capability was significantly increased, and apoptosis was decreased. However, in the Capan-2 cells with wt-p53, suppression of the mesothelin significantly increased wt-p53 levels. When p53 was blocked by siRNA in the stable mesothelin shRNA transfected Capan-2 cells, PUMA was inhibited, followed by increased proliferative capability and decreased cell apoptosis. In the HPAC and Capan-2 cells with wt-p53 and in the MIA PaCa-2 cells with mt-p53, overexpression of the mesothelin significantly decreased bax levels and increased bcl-2 levels, followed by increased proliferative and colony forming capability. Furthermore, mesothelin-shRNA-transfected cells exhibited a reduced rate of tumor growth under in vivo conditions. However, mesothelin-transfected cells exhibited a increased rate of tumor growth under in vivo conditions. Our data demonstrated that mesothelin promotes proliferation and inhibited apoptosis through p53-dependent pathway in pancreatic cancer cells with wt-p53, and p53-independent pathway in pancreatic cancer cells with mt-p53. Targeting mesothelin by shRNA is the important method for pancreatic cancer therapy.
Highlights
Pancreatic cancer remains stubbornly resistant to many key cytotoxic chemotherapeutic agents and novel targeted therapies
More endogenous mesothelin introduced caused lower expression of the pro-apoptotic protein Bax. These results indicate that endogenous mesothelin enhanced the expression of the anti-apoptotic proteins Bcl-2 and Mcl-1, and reduced the expression of the pro-apoptotic protein Bax [10].In the present study,we study whether mesothelin regulates proliferation and apoptosis in pancreatic cancer cells through p53-bcl-2/bax pathway
Expression of mesothelin in human pancreatic cancer cell lines We examined mesothelin expression in AsPC-1(p53-null), HPAC(wt-p53) and Capan-2(wt-p53), Capan-1 and MIA PaCa-2(mutant p53)human pancreatic cancer cell lines by western blot and RT-PCR
Summary
Pancreatic cancer remains stubbornly resistant to many key cytotoxic chemotherapeutic agents and novel targeted therapies. Attempts at improving survival in the past 15 years, in advanced disease, have failed This is true even with the introduction of molecularly targeted agents, chosen on the basis of their action on pathways that were supposedly important in pancreatic cancer development and progression [1]. Mutant mice that lacked both copies of the mesothelin gene had no detectable phenotype, and both male and female mice produced healthy offspring, suggesting that mesothelin is not involved in normal growth and development [3]. It has recently found mesothelin is highly expressed in many common epithelial cancers. This differential expression of mesothelin makes it an attractive target for cancer therapy
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