Metastin and its variant forms suppress migration of pancreatic cancer cells

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Abstract

Metastin, a post-translationally modified variant of KiSS1, was recently identified as an endogenous peptide agonist for a novel G-protein coupled receptor, hOT7T175 (AXOR12, GPR54). In this study, we analyzed the role of KiSS1 and hOT7T175 in both pancreatic cancer tissues and pancreatic cancer cell lines. Furthermore, we synthesized novel short variant forms of metastin and tested the inhibitory effect of those variants on in vitro cell functions that are relevant to metastasis. Pancreatic cancer tissues showed significantly lower expression of KiSS1 mRNA than normal tissues (p=0.018), while cancer tissues showed significantly higher expression of hOT7T175 mRNA than normal pancreatic tissues (p=0.027). In human pancreatic cancer cell lines, KiSS1 mRNA was highly expressed in 2 out of 6 pancreatic cancer cell lines, while hOT7T175 mRNA was expressed in all cell lines at various degrees. PANC-1 cells showed the highest expression of hOT7T175. Exogenous metastin did not suppress cell proliferation but significantly reduced the in vitro migration of PANC-1 cells (p<0.01). Metastin induced activation of ERK1 in PANC-1 and AsPC-1 cells. Finally, we synthesized 3 novel short variant forms of metastin, FM053a2TFA, FM059a2TFA, and FM052a4TFA. These metastin variants significantly suppressed the migration of PANC-1 cells and activated ERK1. These data suggest that the metastin receptor, hOT7T175, is one of the promising targets for suppression of metastasis, and that small metastin variants could be an anti-metastatic agent to pancreatic cancer.

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Materials and methods

Cell culture. Pancreatic cancer cell lines, AsPC-1, BxPC-3, Capan-2, CFPAC-1, PANC-1, and SUIT-2 were purchased from the American Type Culture Collection. Cells were cultured as monolayers in the appropriate medium supplemented with 10% fetal bovine serum, 100 U/ml penicillin, and 100 μg/ml streptomycin at 37 °C in a humid atmosphere of 5% CO2/95% air. As for AsPC-1 and PANC-1, upon reaching 80% confluence, the medium was removed, the cells were washed in phosphate-buffered saline (PBS) and

Expression of KiSS1 and hOT7T175 in pancreatic cancer tissues

First, we measured the mRNA expression levels of KiSS1 and hOT7T175 in 30 pancreatic cancer tissues and in 5 adjacent normal pancreatic tissues. All the normal pancreatic tissues (5/5) and 14/30 of pancreatic ductal carcinoma tissues expressed KiSS1 mRNA (Fig. 1A). The expression level of KiSS1 mRNA in pancreatic cancer tissues was significantly lower than normal pancreatic tissues (p=0.018). In contrast, all the pancreatic cancer tissues (30/30) and normal pancreatic tissues (5/5) expressed

Discussion

In this study, we have demonstrated for the first time that hOT7T175 is expressed in pancreatic cancer tissues, but KiSS1 is less expressed when compared to normal pancreatic tissues. These results are in agreement with the analysis by other investigators in ovarian cancer, breast cancer, and colon cancer [2], [5]. Several reports indicated that KiSS1 and its receptor hOT7T175 are also highly expressed in placenta [4], [5]. The placenta is an invasive tissue, and there are similarities in the

Acknowledgements

This study was supported by a Grant-in-Aid for Scientific Research (#15390395) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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