Abstract 3299: Expression of exchange protein activated by cAMP (Epac) in melanoma and its role in cell migration (2010)

Abstract Background: We previously reported that Epac, a guanine nucleotide exchange factor directly activated by cAMP, increases cell migration in melanoma. Epac1, a major isoform of Epac, increases melanoma cell migration via production of heparan sulfate (HS), a major extracellular glycosaminoglycan. However, the frequency and level of expression of Epac1 in human melanoma tissues is largely unknown. Furthermore, it is still unclear if Epac1's role in heparan sulfate production and cell migration is universal among different melanomas. Methods: Epac1 mRNA expression was examined in melanoma cell lines at different melanoma progression stages, and in human fresh frozen tissue of four different stages, i.e., primary melanoma, regional dermal metastatic melanoma, lymph node metastatic melanoma and distant metastatic melanoma. Also, melanoma and melanocyte cell lines were subjected to cell migration assay with the Boyden chambers. Epac1 expression was ablated by shRNA to examine involvement of Epac1 in cell migration and HS production. Results: There was a tendency toward higher Epac1 mRNA expression with progression of melanoma stage, i.e., Epac1 mRNA was higher in metastatic melanoma (MM) and vertical growth phase (VGP) melanoma cell lines than in a melanocyte cell line. In human melanoma samples, Epac1 mRNA expression was higher in regional dermal (p=0.04, n=10) and lymph node (p=0.03, n=10) metastatic melanoma than in primary melanoma, but not in distant metastatic melanoma; however, 4 out of 10 distant metastatic melanoma showed more than 2-fold increase of Epac1 mRNA expression compared to primary melanoma. These data suggest that Epac1 expression correlates with melanoma stage. When cells were incubated with 8-pMeOPT-2′-O-Me-cAMP, an Epac-specific agonist, cell migration was increased in melanoma cell lines but not in melanocytes. Heparatinase, a heparan sulfate-degrading enzyme, inhibited 8-pMeOPT-2′-O-Me-cAMP-induced cell migration in 2 metastatic melanoma cell lines, SK-Mel-2 (64.6±7.5 vs. 35.2±5.5 cells/field, p&amp;lt;0.05, n=4) and SK-Mel-187 (70.2±6.6 vs 43.2±9 cells/field, p&amp;lt;0.05, n=4) in parallel with decreased HS production. When Epac1 expression was ablated by shRNA, cell migration was decreased (SK-Mel-2, 92.6±4 vs. 49.3±1 cells/field, p&amp;lt;0.05, n=4; SK-Mel-187, 77.9±9 vs. 31.6±5 cells/field, p&amp;lt;0.05, n=4) in parallel with decreased HS production (SK-Mel-2,13 vs. 6.8±0.6 μg/ml, p&amp;lt;0.05, n=4; SK-Mel-187, 46.6±6.2 vs. 26.1±8.2 μg/ml, p&amp;lt;0.05, n=4). These data suggest that Epac1 expression plays a major role in melanoma cell migration through heparan sulfate production, and Epac's role is universal in metastatic melanoma cells. Conclusion: Epac1 expression increases with the progression of melanoma. Epac1 mediates melanoma cell migration via HS production in different melanoma cell lines. These data suggest that Epac1 could be a target for treating patients with melanoma. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3299.