Epidemiological, clinical and experimental evidence suggests a link between type 2 diabetes and Alzheimer's disease (AD). Insulin modulates metabolism of ␤-amyloid precursor protein (APP) in neurons, decreasing the intracellular accumulation of ␤-amyloid (A␤) peptides, which are pivotal in AD pathogenesis. The present study investigates whether the widely prescribed insulin-sensitizing drug, metformin (Glucophage R ), affects APP metabolism and A␤ generation in various cell models. We demonstrate that metformin, at doses that lead to activation of the AMP-activated protein kinase (AMPK), significantly increases the generation of both intracellular and extracellular A␤ species. Furthermore, the effect of metformin on A␤ generation is mediated by transcriptional up-regulation of ␤-secretase (BACE1), which results in an elevated protein level and increased enzymatic activity. Unlike insulin, metformin exerts no effect on A␤ degradation. In addition, we found that glucose deprivation and various tyrphostins, known inhibitors of insulin-like growth factors/insulin receptor tyrosine kinases, do not modulate the effect of metformin on A␤. Finally, inhibition of AMP-activated protein kinase (AMPK) by the pharmacological inhibitor Compound C largely suppresses metformin's effect on A␤ generation and BACE1 transcription, suggesting an AMPK-dependent mechanism. Although insulin and metformin display opposing effects on A␤ generation, in combined use, metformin enhances insulin's effect in reducing A␤ levels. Our findings suggest a potentially harmful consequence of this widely prescribed antidiabetic drug when used as a monotherapy in elderly diabetic patients. A lzheimer's disease (AD) is a devastating neurodegenerative disorder, with aging, genetic, and environmental factors contributing to its development and progression. AD is not only characterized by pathological deposition of A␤ peptides and neurofibrillary tangles but is also associated with microgliamediated inflammation and dysregulated lipid homeostasis and glucose metabolism. Amyloid peptides are derived from sequential proteolytic cleavages of full-length amyloid precursor protein (APP) by ␤-secretase (BACE1) and ␥-secretase. Full-length APP can undergo alternative processing by ␣-secretase, releasing a soluble fragment (sAPP␣) extracellularly, which precludes A␤ formation. Compelling evidence indicates that A␤, especially the oligomers, are toxic to neurons; excessive generation and accumulation of A␤ peptides in neurons is believed to initiate the pathological cascade in AD (1-3).Epidemiological studies strongly suggest that metabolic defects correlate with the functional alterations associated with aging of the brain and with AD pathogenesis (4-11). The vast majority of AD cases are late onset and sporadic in origin with aging being the most profound risk factor. Insulin signaling is known to be involved in the process of brain aging (12)(13)(14)(15)(16)(17)(18)(19)(20). Insulin dysfunction/resistance in diabetes mellitus (DM) is not only a common syndrome ...

Organic anion-transporting polypeptide 1A2 (OATP1A2) is a drug uptake transporter known for broad substrate specificity, including many drugs in clinical use. Therefore, genetic variation in SLCO1A2 may have important implications to the disposition and tissue penetration of substrate drugs. In the present study, we demonstrate OATP1A2 protein expression in human brain capillary and renal distal nephron using immunohistochemistry. We also determined the extent of single nucleotide polymorphisms in SLCO1A2 upon analyses of ethnically defined genomic DNA samples (n ‫؍‬ 95 each for African-, Chinese-, European-, and Hispanic-Americans). We identified six nonsynonymous polymorphisms within the coding region of SLCO1A2 (T38C (I13T), A516C (E172D), G559A (A187T), A382T (N128Y), A404T (N135I), and C2003G (T668S)), the allelic frequencies of which appeared to be ethnicitydependent. In vitro functional assessment revealed that the A516C and A404T variants had markedly reduced capacity for mediating the cellular uptake of OATP1A2 substrates, estrone 3-sulfate and two ␦-opioid receptor agonists, deltorphin II, and [D-penicillamine 2,5 ]-enkephalin. On the other hand, the G559A and C2003G variants appeared to have substrate-dependent changes in transport activity. Cell surface biotinylation and immunofluorescence confocal microscopy suggested that altered plasma membrane expression of the transporter may contribute to reduced transport activity associated with the A516C, A404T, and C2003G variants. The A404T (N135I) variant also showed a shift in the apparent molecular size, indicative of alterations in glycosylation status. Taken together, these data suggest that SLCO1A2 polymorphisms may be an important yet unrecognized contributor to interindividual variability in drug disposition and central nervous system entry of substrate drugs.During the past decade, there has been an increasing recognition of the critical interplay between drug transporters and drug-metabolizing enzymes as determinants of drug disposition and response. Indeed, the extent of targeted tissue entry for many drugs may be facilitated by drug transporters, which are often expressed in a tissue-specific manner with broad substrate specificities. Among the uptake transporters, members of the organic anion-transporting polypeptides (human, OATPs 1 ; rodents, Oatps) have been shown to be expressed in organs such as the central nervous system, liver, and intestine and mediate the cellular uptake of a large number of structurally divergent compounds (1). Within this family, OATP1A2 (SLCO1A2, also known as human OATP-A or OATP1) was the first human OATP to be cloned and characterized (2). OATP1A2 mRNA has been detected in various tissues including the brain, liver, and kidney (2, 3). Substrates of OATP1A2 include endogenous compounds such as bile acids, steroid hormones, and their conjugates, thyroid hormones, as well as drugs including fexofenadine, ouabain, peptides (e.g. deltorphin II, [D-penicillamine 2,5 ]enkephalin, DPDPE), and the toxin, microcystin (2, ...

Cardiac hypertrophy is initiated as an adaptive response to sustained overload but progresses pathologically as heart failure ensues1. Here we report that genetic loss of APJ confers resistance to chronic pressure overload by dramatically reducing myocardial hypertrophy and heart failure. In contrast, mice lacking apelin (the endogenous APJ ligand) remain sensitive, suggesting an apelin independent function of APJ. Freshly isolated APJ-null cardiomyocytes exhibit an attenuated response to stretch, indicating that APJ is a mechano-sensor. Activation of APJ by stretch increases cardiomyocyte cell size and induces molecular markers of hypertrophy. Whereas apelin stimulates APJ to activate Gαi and elicits a protective response, stretch signals in an APJ-dependent G-protein-independent fashion to induce hypertrophy. Stretch-mediated hypertrophy is prevented by knockdown of β-arrestins or by pharmacological doses of apelin acting through Gαi. Taken together, our data indicate that APJ is a bifunctional receptor for both mechanical stretch and for the endogenous peptide apelin. By sensing the balance between these stimuli, APJ occupies a pivotal point linking sustained overload to cardiomyocyte hypertrophy.

The proteasome is a vital cellular machine that maintains protein homeostasis, which is of particular importance in multiple myeloma and possibly other cancers. Targeting proteasome 20S peptidase activity with bortezomib and carfilzomib has been widely used to treat myeloma. However, not all patients respond, and those that do eventually suffer relapse. Therefore, there is an urgent and unmet need to develop novel drugs that target proteostasis through different mechanisms. We identified quinoline-8-thiol (8TQ) as a first-in-class inhibitor of the proteasome 19S subunit Rpn11. A derivative of 8TQ, capzimin, shows >5-fold selectivity for Rpn11 over the related JAMM proteases and >2 logs less activity towards metalloenzymes. Capzimin stabilized proteasome substrates, induced an unfolded protein response, and blocked proliferation of cancer cells, including those resistant to bortezomib. Proteomic analysis revealed that capzimin stabilized a subset of polyubiquitinated substrates. Identification of capzimin offers an alternative path to develop proteasome inhibitors for cancer therapy.

SUMMARY Hepatocyte Nuclear Factor (HNF)4α is a central regulator of gene expression in cell types that play a critical role in metabolic homeostasis, including hepatocytes, enterocytes, and pancreatic β-cells. Although fatty acids were found to occupy the HNF4α ligand-binding pocket and proposed to act as ligands, there is controversy about both the nature of HNF4α ligands as well as the physiological role of the binding. Here, we report the discovery of potent synthetic HNF4α antagonists through a high-throughput screen for effectors of the human insulin promoter. These molecules bound to HNF4α with high affinity and modulated the expression of known HNF4α target genes. Notably, they were found to be selectively cytotoxic to cancer cell lines in vitro and in vivo, although in vivo potency was limited by suboptimal pharmacokinetic properties. The discovery of bioactive modulators for HNF4α raises the possibility that diseases involving HNF4α, such as diabetes and cancer, might be amenable to pharmacologic intervention by modulation of HNF4α activity.