The BCA2 & AMPK paradigm: Unraveling the cancer connection.

摘要

Adenosine monophosphate-activated kinase (AMPK), a master regulator of cellular energy homeostasis, has emerged as a promising molecular target in the prevention of breast cancer, and phase II and III clinical trials using the FDA-approved, AMPK activating, anti-diabetic drug metformin are promising in this regard, but the question of why metformin is protective for some women but not others still remains. B&barbelow;reast C&barbelow;ancer A&barbelow;ssociated Gene 2&barbelow; (BCA2/Rabring7/RNF115), a novel RING-finger ubiquitin E3 ligase, is overexpressed in >50% of breast tumors. Herein, I hypothesized that BCA2 is an endogenous inhibitor of AMPK activation in breast cancer cells and that BCA2 inhibition would therefore increase the efficacy of metformin. My hypothesis is strongly supported by the finding that BCA2 overexpression inhibited both basal and inducible Thr172 phosphorylation/activation of AMPKalpha1, while BCA2-specific siRNA enhanced pAMPKalpha1. Furthermore, the AMPK-suppressive function of BCA2 requires its E3 ligase-specific RING domain, suggesting that BCA2 targets a critical protein controlling (de)phosphorylation of pAMPKalpha1 for degradation. A large scale proteomics analysis has revealed that PP2A and ITPR3 may be promising leads in this regard and should be further investigated in relation to AMPK activation by specifically CaMKK. Activation of AMPK by metformin not only triggered growth a inhibitory signal, but also increased BCA2 protein (but not mRNA) levels, which correlated positively with activation of AKT and could be curbed by an AMPK inhibitor, suggesting a potential feedback mechanism from AMPK to pAkt to BCA2. Finally, BCA2 siRNA, or inhibition of its upstream stabilizing kinase AKT, increased the growth-inhibitory effect of metformin in multiple breast cancer cell lines, supporting the conclusion that BCA2 weakens metformin's efficacy in breast cancer cells. My data therefore suggests that metformin in combination with a BCA2 inhibitor may be a more effective breast cancer treatment strategy than metformin alone. On this account, a first generation of "specific" BCA2 inhibitors, as well as DSF inhibitors, is in existence and all compounds have been evaluated for their cancer cell anti-proliferative effects and should be further tested in combination with metformin. The studies performed in this dissertation provide new grounds for the development of BCA2 as a novel anti-cancer drug target, and in addition, provide awareness for the potential limitations metformin use in the clinic may have.