Abstract
The aim of this study is to investigate the A(2B)AR-dependence of okhotoside A(1)-1 cytotoxic and antiproliferative action on triple-negative MDA-MB-231 breast cancer cells using monolayer and 3D culture approaches. Earlier triterpene glycoside okhotoside A(1)-1 (Okh) was isolated from the sea cucumbers Cucumaria djakonovi and C. conicospermium and its selective cytotoxicity against MDA-MB-231 vs. non-tumorigenic MCF-10A cells was reported. Now it has been found that the A(2B) adenosine receptor (A(2B)AR) is one of the molecular targets for Okh and its antiproliferative effect is A(2B)AR-dependent. Molecular docking studies suggested a unique behavior for Okh demonstrating two highly probable binding modes with comparable affinity, when the aglycone is immersed in the binding pocket, or alternatively, the carbohydrate moiety occupies the site. The glycoside modulated cAMP and intracellular Ca(2+) levels in an A(2B)AR-dependent manner, which accompanied by the suppression of p38 MAPK and ERK1/2 phosphorylation, and blocked cell cycle progression. Okh induced mitochondrial dysfunction, characterized by increased ROS production and loss of the mitochondrial membrane potential (ΔΨm), which led to the upregulation of APAF-1 and cytochrome C, activation of caspases-9 and -3, and initiation of apoptosis. The antitumor potential of Okh was confirmed in a 3D culture of MDA-MB-231 cells and was more significant than those of another A(2B)AR-targeted triterpene glycoside cucumarioside A(0)-1 and cisplatin.