X-3, a mangiferin derivative, stimulates AMP-activated protein kinase and reduces hyperglycemia and obesity in db/db mice Jun Han a , Jia Yi b , Fengying Liang b , Bo Jiang b , Ying Xiao b , Shouhong Gao b , Na Yang b , Honggang Hu c , Wei-Fen Xie a , Wansheng Chen b, * a Department of Gastroenterology, Changzheng Hospital, Second Military Medical University, Shanghai, China b Department of Pharmacy, Changzheng Hospital, Second Military Medical University, Shanghai, China c Department of Organic Chemistry, School of Pharmacy, Second Military Medical University, Shanghai, China ARTICLE INFO Article history: Received 12 May 2014 Received in revised form 12 January 2015 Accepted 6 February 2015 Available online 11 February 2015 Keywords: AMP-activated protein kinase Animal experiment Anti-diabetic Compounds A B ST R AC T Diabetes mellitus is a major health concern, affecting nearly 10% of the population. Here we describe a potential novel therapeutic agent for this disease, X-3, a derivative of mangiferin. Therapeutic adminis- tration of X-3 significantly and dose-dependently reduced plasma glucose and triglycerides in db/db mice following 8 week-treatments. Treatment with X-3 dose-dependently increased the number of insulin- positive β-cell mass. Importantly, X-3 did not cause any death or signs of toxicity in acute toxicity studies. Study of mechanism of action revealed that X-3 increased glucose uptake in parallel with increased phos- phorylation of AMP-activated protein kinase (AMPK) in 3T3-L1 cells. It activates AMPK in both LKB1- dependent and -independent manner. Furthermore, administration of X-3 resulted in activation of AMPK and its downstream target, acetyl-CoA carboxylase (ACC) in the hypothalamus, liver, muscle and adipose tissues of C57BL/6 mice. An 80 mg/kg X-3 was more potent than metformin at 500 mg/kg in the hypo- thalamus, and interscapular fat tissues, potent than MF at the same dose in the liver. Thus, we conclude that X-3 is a promising new class of AMPK activating drug, and can potentially be used in the treatment of type 2 diabetes. © 2015 Published by Elsevier Ireland Ltd. 1. Introduction Mangiferin (MF) is a xanthonoid found in mangoes and Anemarrhena asphodeloides rhizomes (Miura et al., 2001). Mangiferin has been used in India for the treatments of arteriosclerosis, cor- onary heart disease and diabetes. It has been shown that MF exhibits antidiabetic (Ichiki et al., 1998; Muruganandan et al., 2005; Yoshikawa et al., 2001), hypolipidemic and antiatherogenic prop- erties (Guo et al., 2011; Muruganandan et al., 2002, 2005) by reducing plasma total cholesterol, triglycerides, low density lipoprotein- cholesterol (LDL-C) and increasing high density lipoprotein (HDL- C) (Muruganandan et al., 2005). Niu et al. has recently reported that MF decreased plasma FFA in hyperlipidemic rats and activated AMPK in liver, whereas there is no sufficient evidence to show that MF could alone activate AMPK at the cellular level (Niu et al., 2012). More- over the major shortcomings of MF are its poor solubility and oral bioavailability (Cai et al., 2010). AMPK is a major cellular energy sensor and a master regulator of metabolic homeostasis (Viollet et al., 2009; Zhang et al., 2009). AMPK is a heterotrimeric protein kinase consisting of a catalytic (α) and two regulatory subunits (β and γ) (Hardie et al., 2006; Viollet et al., 2006). AMPK are activated by two distinct signals: a Ca 2+ - dependent pathway mediated by calcium/calmodulin-dependent protein kinase kinase β (CaMKKβ) and an AMP-dependent pathway mediated by LKB1 (Sanders et al., 2007). Under conditions of energy depletion, AMPK inhibits ATP-consuming pathways (e.g., fatty acid synthesis, cholesterol synthesis, protein synthesis and gluconeo- genesis) and stimulates ATP-generating processes (e.g., fatty acid oxidation and glycolysis), thus restoring overall cellular energy ho- meostasis (Carling, 2004; Zhang et al., 2009). In addition, AMPK activation acutely increases glucose uptake (via Glut1 and Glut4) and glycolysis (Hue et al., 2002; Jing and Ismail-Beigi, 2006; Jones and Dohm, 1997). In lipid metabolism, AMPK activation results in the phosphorylation and inactivation of ACC (Carling et al., 1987), a direct AMPK substrate, leading to decreased conversion of acetyl- CoA to malonyl CoA. Malonyl CoA allosterically inhibits carnitine palmitoyl-CoA transferase (CPT1), the rate-limiting step in trans- port of long chain acyl-CoAs into mitochondria for oxidation (Lochhead et al., 2000; McGarry and Brown, 1997). Therefore, a re- duction in malonyl CoA levels increases fatty acid oxidation. AMPK, independently of insulin, is able to phosphorylate Akt sub- strate AS160, while inhibition of AS160 is believed to allow increased GLUT4 membrane localization and glucose uptake (Sano et al., 2007). * Corresponding author. Changzheng Hospital, 415 Fengyang Rd., Huangpu District, Shanghai 200003, China. Tel.: +862181871349; fax: +862181886181. E-mail address: wschen126@126.com (W. Chen). http://dx.doi.org/10.1016/j.mce.2015.02.008 0303-7207/© 2015 Published by Elsevier Ireland Ltd. Molecular and Cellular Endocrinology 405 (2015) 63–73 Contents lists available at ScienceDirect Molecular and Cellular Endocrinology journal homepage: www.elsevier.com/locate/mce