Effect of Protease Activated Receptor‐2 antagonist in the transition from endometrium to endometriosis and ovarian cancer

Abstract

Endogenous proteases are associated with the ovulation process and inflammation, among other human physiological processes. Still their role in the transition from normal tissue to cancer development remains uncertain. For example, the Protease activated receptor 2 (PAR‐2) is a G Protein Couple receptor expressed in mostly in epithelial cells, including the female reproductive system. Previous work from our lab showed that the PAR‐2 antagonist‐FSLLRY‐NH2 inhibited the cell‐mediated inflammation in the airways. The aim of this study is to evaluate the effect of the PAR‐2 antagonist when a normal cell progresses to a cancerous one, using trypsin as a gold standard enzyme for Par‐2 activation. In this case, the transition from normal endometrium, to endometriosis, ovarian. We hypothesized that PAR‐2 antagonist will decrease the activation of its receptor reducing the transition to endometriosis and ovarian cancer. To test this, we performed western blot to determine PAR‐2 expression in endometriosis (12Z) cells, human endometrial stromal cells (HESC) cells, ovarian cancer (A2780) cells, and breast cancer (MCF‐7) cells. In addition, we quantified the activation of PAR‐2 receptor using a Fluorogenic intracellular calcium assay in human normal endometrial (uterine) epithelial cells (HEUEC), ovarian cancer (Caov‐3), and MCF‐7 cells as internal positive control. We also performed the BrdU proliferation assay on these cells. Our results showed a higher expression (p<0.05) of PAR‐2 in A2780 cells (ovarian aggressive cancer) when compared with the other cells lines. The intracellular calcium assay resulted in a significant reduction (p<0.05) once exposed to the PAR‐2 antagonist in the HEUEC cells and Caov‐3 when compared to cells treated with trypsin (p<0.05). In contrast, there was not significant difference in the MCF‐7 cell line upon exposure to treatments. We observed a significant decrease in MCF‐7 proliferation when treated with the PAR‐2 antagonist in comparison to trypsin treatment. Our results suggested that PAR‐2 antagonist can be used as treatment or adjunct therapy for specific female reproductive cancers. Next steps of this study will include the active full‐length PAR‐2 recombinant and test other concentrations of the antagonist to determine the LD50 in the selected cells. Proliferating markers will be measured for all the previous cell lines as well for endometrial cancer.