Phosphoproteome profiling provides insight into the mechanism of action for carvedilol‐mediated cancer prevention

Abstract

Recent studies suggest that the β-blocker drug carvedilol prevents skin carcinogenesis but the mechanism is unknown. Carvedilol is one of a few β-blockers identified as biased agonist based on an ability to promote β-arrestin-mediated processes such as ERK phosphorylation. To understand the role of phosphoproteomic signaling in carvedilol's anticancer activity, the mouse epidermal JB6 P+ cells treated with EGF, carvedilol, or their combination were analyzed using the Phospho Explorer Antibody Array containing 1318 site-specific and phospho-specific antibodies of over 30 signaling pathways. The array data indicated that both EGF and carvedilol increased phosphorylation of ERK's cytosolic target P70S6 K while its nuclear target ELK-1 were activated only by EGF; Furthermore, EGF-induced phosphorylation of ELK-1 and c-Jun was attenuated by carvedilol. Subcellular fractionation analysis indicated that ERK nuclear translocation induced by EGF was blocked by co-treatment with carvedilol. Western blot and luciferase reporter assays confirmed that the biased β-blockers carvedilol and alprenolol blocked EGF-induced phosphorylation and activation of c-Jun/AP-1 and ELK-1. Consistently, both carvedilol and alprenolol strongly prevented EGF-induced neoplastic transformation of JB6 P+ cells. Remarkably, oral carvedilol treatment significantly inhibited the growth of A375 melanoma xenograft in SCID mice. As nuclear translocation of ERK is a key step in carcinogenesis, inhibition of this event is proposed as a novel anticancer mechanism for biased β-blockers such as carvedilol.

The β-Blocker Carvedilol Prevented Ultraviolet-Mediated Damage of Murine Epidermal Cells and 3D Human Reconstructed Skin

“…Carvedilol-mediated protective effects were unique as neither UV-nor H 2 O 2 -mediated reductions in cell viability were altered by metoprolol or 4-OHC. This result is consistent with previous reports that only carvedilol, but not 4-OHC and metoprolol, inhibited UV-or EGF-induced NF-κB and AP-1 promoter activity and JB6 P+ neoplastic transformation [7,9]. Resveratrol, used as a positive control in the ROS assay, is a potent antioxidant with relatively poor direct scavenging activity [24].…”

“…Two potential mechanisms for carvedilol's protective effects were examined in these experiments: β-AR blockade and ROS scavenging activity. Metoprolol was used as a control β-blocker without anti-transformation activity [9] and without proven antioxidant property [19], but is a commonly used β-blocker. 4-OHC is an intermediate compound in carvedilol synthesis that acted as a free radical scavenger (i.e., chemical antioxidant) [20].…”

“…Carvedilol is a receptor subtype non-selective β-blocker with α-AR blocking and antioxidant properties [3][4][5]. Recent data demonstrate that carvedilol prevents malignant transformation and carcinogenesis induced by epidermal growth factor (EGF), chemical carcinogens, and UV [6][7][8][9]. However, the cancer preventive mechanism for carvedilol remains unknown.…”

“…However, the cancer preventive mechanism for carvedilol remains unknown. Notably, not all β-blockers had effects on EGF-induced transformation [6,9]. Since all β-blockers inhibit β-ARs, the stand-out cancer preventive activity of carvedilol suggests that the mechanism might be associated with a unique property of carvedilol and that the β-ARs may not be the direct target for carvedilol's cancer preventive activity [10].…”

“…Carvedilol's protective activity was compared with metoprolol, a β-blocker without proven antioxidant property [19], and an intermediate compound for carvedilol synthesis, 4-hydroxycarbazole (4-OHC), which possesses a chemical antioxidant activity [20]. In previous studies, metoprolol and 4-OHC did not show cancer preventive efficacy when compared with carvedilol in vitro and in vivo [7,9]. Furthermore, to confirm carvedilol's photoprotective activity, a reconstituted 3-dimensional (3D) human skin model was used.…”