Carvedilol inhibits EGF-mediated JB6 P+ colony formation through a mechanism independent of adrenoceptors

Abstract

Carvedilol is reported to prevent cancers in humans and animal models. However, a molecular mechanism has yet to be established, and the extent to which other P-blockers are chemopreventive remains relatively unknown. A comparative pharmacological approach was utilized with the expectation that a mechanism of action could be devised. JB6 CI 41-5a (JB6 P+) murine epidermal cells were used to elucidate the chemopreventative properties of beta-blockers, as JB6 P+ cells recapitulate in vivo tumor promotion and chemoprevention. The initial hypothesis was that beta-blockers that are GRK/beta-arrestin biased agonists, like carvedilol, are chemopreventive. Sixteen beta-blockers of different classes, isoproterenol, and HEAT HCI were individually co-administered with epidermal growth factor (EGF) to JB6 P+ cells to examine the chemopreventative properties of each ligand. Cytotoxicity was examined to ensure that the anti-transformation effects of each ligand were not due to cellular growth inhibition. Many of the examined p-blockers suppressed EGF-induced JB6 P+ cell transformation in a non-cytotoxic and concentration-dependent manner. However, the IC50 values are high for the most potent inhibitors (243, 326, and 431 nM for carvedilol, labetalol, and alprenolol, respectively) and there is no correlation between pharmacological properties and inhibition of transformation. Therefore, the role of alpha 1- and beta 2-adrenergic receptors (AR) was examined by standard competition assays and shRNA targeting beta 2-ARs, the only beta-AR expressed in JB6 P+ cells. The results reveal that pharmacological inhibition of alpha 1- and beta 2-ARs and genetic knockdown of beta 2-ARs did not abrogate carvedilol-mediated inhibition of EGF-induced JB6 P+ cell transformation. Furthermore, topical administration of carvedilol protected mice from UV-induced skin damage, while genetic ablation of beta 2-ARs increased carvedilol-mediated effects. Therefore, the prevailing hypothesis that the chemopreventive property of carvedilol is mediated through P-ARs is not supported by this data.