The role of adenosine A3 receptors in cytotoxicity of the breast cancer cell lines

The nucleoside adenosine is present within cells and body fluids of all living organisms and its production, both intra- and extracellularly, is tightly coupled to energy consumption resulting in increased level of extracellular adenosine. The physiological effects of adenosine are mediated through four pharmacologically and biochemically distinct adenosine receptors (AR), i.e. A1, A2A, A2B and A3. Al and A3 AR generally couple to Gi proteins, whereas A2A and A2B receptors activate Gs proteins. Although the presence of these receptors has been reported in both normal and cancer cells, no data is available regarding breast cancer. Therefore, this study was aimed to investigate the existence and possible role of A3 AR in ER+ MCF- 7 and ER- MDA-MB468 breast cancer cell lines. The cell lines were cultured in RPMI-1640 medium and incubated with different concentrations of IB-MECA (0.1-100 µM), A3 selective agonist, and MRS-1220 (0.1-10 µM), a highly selective antagonist for different time periods (24 - 72 hr). MTT viability test was used to evaluate the proliferative and cytotoxic response. Then, mRNA was isolated and reverse transeriptase polymerase chain reaction (RT -PCR) was used. The results showed that IB-MECA at concentrations higher than 10 µM results in a significant cell death (p<0.05), which reached the maximum level 48 hr after the experiment in both cell lines. In addition, pretreatment with MRS-1220 at 0.1 and l µM prevented the cytotoxic effect of IB-MECA at 30 and 60 µM, indicating that A3 receptor is present on both cell lines. Application of MRS-1220 had no effect when used alone. Further confirmation was provided by the-application of RT -PCR method. Since both cell lines were responsive to the A3 agonist treatment and as of the presence of the receptor mRNA in RT -PCR analysis, it is concluded that this membrane receptor exists in the breast cancer cell lines, irrespective of ER status. These data are new and informative, emphasizing the role of A3 receptor in the cell cytotoxicity, which may introduce a new perspective on the mode of action and possible application of adenosine in breast cancer.