AMP-activated protein kinase (AMPK) is a key sensor and regulator of intracellular and whole-body energy metabolism. We have identified a thienopyridone family of AMPK activators. A-769662 directly stimulated partially purified rat liver AMPK (EC50 = 0.8 microM) and inhibited fatty acid synthesis in primary rat hepatocytes (IC50 = 3.2 microM). Short-term treatment of normal Sprague Dawley rats with A-769662 decreased liver malonyl CoA levels and the respiratory exchange ratio, VCO2/VO2, indicating an increased rate of whole-body fatty acid oxidation. Treatment of ob/ob mice with 30 mg/kg b.i.d. A-769662 decreased hepatic expression of PEPCK, G6Pase, and FAS, lowered plasma glucose by 40%, reduced body weight gain and significantly decreased both plasma and liver triglyceride levels. These results demonstrate that small molecule-mediated activation of AMPK in vivo is feasible and represents a promising approach for the treatment of type 2 diabetes and the metabolic syndrome.

The nontransmembrane protein tyrosine phosphatase SHP-2 plays a critical role in growth factor and cytokine signaling pathways. Previous studies revealed that a fraction of SHP-2 moves to focal contacts upon integrin engagement and that SHP-2 binds to SHP substrate 1 (SHPS- Complex processes such as cell growth, differentiation, and migration require the integration of multiple types of extracellular signals, including those delivered by growth factors, cytokines, and hormones (soluble signals), and solid-state signals, transmitted by cell-cell and cell-extracellular matrix (ECM) interactions. Most of these signaling pathways involve changes in cellular tyrosyl phosphorylation. Tyrosyl phosphorylation is regulated by protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). Although many PTKs are implicated in signaling pathways for both soluble and solid-state signals, the roles of specific PTPs are less well defined.1Multiple reverse-genetic studies suggest that the nontransmembrane PTP SHP-2 is a required positive (i.e., signalenhancing) component of growth factor and cytokine signal transduction pathways (for reviews, see references 35, 56, and 63). Consistent with these studies, fibroblasts from mice containing a deletion of SHP-2 exon 3 (hereafter referred to as SHP-2 mutant mice), which express low levels of a defective SHP-2 protein that lacks its N-terminal SH2 domain (48), exhibit impaired mitogen-activated protein kinase (MAPK) activation in response to fibroblast growth factor (FGF), epidermal growth factor, and insulin-like growth factor I (48, 55). SHP-2 functions similarly in lower organisms. Dominant-negative SHP-2 blocks FGF-induced mesoderm induction in Xenopus ectodermal explants and completion of gastrulation in early embryos, leading to severe tail truncations (38, 57). Likewise, corkscrew (csw), the Drosophila SHP-2 homolog (39, 40), is required for multiple receptor tyrosine kinase (RTK) pathways involved in early development (e.g., Torso, Sevenless, Breathless, and Drosophila EGF receptor) (1,39,40), and the recently described Caenorhabditis elegans homolog, Ptp-2, is a component of the Let-23 pathway (13).The precise mechanism by which SHP-2 orthologs function, as well as their specific target(s), has remained unclear. SHP-2 binds directly to and may dephosphorylate some growth factor and cytokine receptors (for reviews, see references 35 and 63). In other pathways, however, SHP-2 binds to distinct signaling intermediates. One class of SHP-2 binding proteins, exemplified by the Drosophila daughter of sevenless (dos) gene product, consists of an N-terminal pleckstrin homology domain and multiple proline-rich stretches and potential tyrosyl phosphorylation sites (42). Dos is essential for Sevenless signaling (15,42) and may be a direct substrate for Csw (15). Mammalian cells express several groups of molecules with overall topology similar to that of Dos (for a review, see reference 63). including insulin receptor substrate family members (for a review, see reference 64), ...

Phosphatidylinositol 3-kinase (PI3K) activation is necessary for many insulin-induced metabolic and mitogenic responses. However, it is unclear whether PI3K activation is sufficient for any of these effects. To address this question we increased PI3K activity in differentiated 3T3-L1 adipocytes by adenovirus-mediated expression of both the inter-SH2 region of the regulatory p85 subunit of PI3K (iSH2) and the catalytic p110␣ subunit (p110). Coexpression resulted in PI3K activity that exceeded insulin-stimulated activity by two-to fivefold in cytosol, total membranes, and the low density microsome (LDM) fraction, the site of greatest insulin stimulation. While insulin increased glucose transport 15-fold, coexpression of iSH2-p110 increased transport 5.2-؎ 0.7-fold with a parallel increase in GLUT4 translocation to the plasma membrane. Constitutive activation of PI3K had no effect on maximally insulin-stimulated glucose transport. Neither basal nor insulinstimulated activity of glycogen synthase or mitogen-activated protein kinase was altered by iSH2-p110 coexpression. DNA synthesis was increased twofold by insulin in control 3T3-L1 adipocytes transduced with ␤-galactosidase-encoding recombinant adenovirus, while iSH2-p110 coexpression increased DNA synthesis fivefold. These data indicate that (i) increased PI3K activity is sufficient to activate some but not all metabolic responses to insulin, (ii) activation of PI3K to levels exceeding the effect of insulin in adipocyte LDM results in only a partial stimulation of glucose transport, and (iii) increased PI3K activity in the absence of growth factor or oncoprotein stimulation is a potent stimulus of DNA synthesis.Insulin is pivotal in the regulation of glucose homeostasis. One of the primary actions of insulin is the stimulation of glucose transport into insulin-sensitive tissues by eliciting translocation of the major insulin-responsive glucose transporter, GLUT4, from an intracellular pool to the plasma membrane. Resistance to this stimulatory effect of insulin is a major pathologic feature of both type I and type II diabetes (5, 43). In most insulin-resistant states the expression of GLUT4 in skeletal muscle, the major site of insulin-stimulated glucose disposal, is not reduced (16), suggesting a defect in the signalling cascade or the cellular elements linking the insulin receptor to GLUT4 translocation, fusion with the plasma membrane, and/or activation (15). Although the number of molecules known to be involved in insulin signalling is increasing rapidly (2), the exact signalling pathways responsible for glucose transporter translocation remain to be defined. Ultimately these pathways could be attractive targets for novel therapeutic strategies to reduce or prevent insulin resistance.Insulin stimulation of phosphatidylinositol 3-kinase (PI3K) was first observed by Ruderman et al. (31). Subsequently, several studies using either the PI3K inhibitors wortmannin and LY 294002 (3, 27) or dominant negative approaches (22) demonstrated that stimulation of PI3K activ...