Abstract 3021: Development of small molecule direct AMPK activators for the treatment of cancer
Authors: Hitoshi Yasumichi , Jenkins Yonchu , Li Yingwu , Sampang Elmer , Xu Xiang , Dong Guodong , Huang Jianing , Lin Nan , Goff Dane , Shaw Simon , Boralsky Luke , Singh Rajinder , Issakani Sarkiz D. , Payan Donald G. 
Published: July 2016

Abstract
5’-AMP-activated protein kinase (AMPK) is a key sensor of cellular energy status and is critical for maintaining energy homeostasis under conditions of nutrient stress. During cellular transformation, metabolic reprogramming enables the aberrant growth and proliferation of tumor cells. Both positive and negative roles for AMPK in tumor cell proliferation and survival have been reported. However, only a limited number of studies addressed this question with potent direct AMPK activators. AMPK exists as heterotrimers composed of the catalytic subunit α and reguratory subunits β and γ. We expressed the full length of all three human AMPK subunits in insect cells, purified the heterotrimer complexes, and used them for biochemical screening and characterization of AMPK activators. The purified complexes displayed basal activity, which was further enhanced by AMP. The compounds we identified potently activated the complexes in vitro at AC(2X)s (the concentration that gives a twofold activation) of 0.001-0.3 μM. Importantly, the compounds up-regulated substrate phosphorylation (pS79 Acetyl-CoA Carboxylase) and/or auto-phosphorylation (pT172 AMPKα) in multiple cancer cell lines including HepG2 hepatoma cells, A549 liver kinase B1 (LKB1) null lung cancer cells, and MOLM14 myeloid leukemia cells, indicating activation was irrespective of functional status of LKB1, which is a key AMPK-activation kinase. Activation of AMPK by the compounds was also confirmed using native AMPK isolated from normal tissues and tumor cells. We further investigated anti-proliferative effects of the compounds and found that up-regulation of AMPK kinase activity was correlated with anti-proliferative effects in A549 and MOLM14, but not in HepG2, suggesting that positive effects of direct AMPK activators could be cell-type dependent. Interestingly, we identified compounds that display comparable AMPK activation in HepG2 and A549 yet possessed divergent activities on proliferation across a panel of tumor lines. Analysis of cellular signaling across several of these tumor lines with this set of the compounds revealed dose-dependent effects on mTORC1 substrates, feedback signaling to PI3K and mTORC2, and inhibition of kinases downstream of RAF. Direct activation of AMPK could be a good therapeutic strategy for the treatment of subsets of cancers.
Citation Format: Yasumichi Hitoshi, Yonchu Jenkins, Yingwu Li, Elmer Sampang, Xiang Xu, Guodong Dong, Jianing Huang, Nan Lin, Dane Goff, Simon Shaw, Luke Boralsky, Rajinder Singh, Sarkiz D. Issakani, Donald G. Payan. Development of small molecule direct AMPK activators for the treatment of cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3021.