A new frontier in pharmacology: the endoplasmic reticulum as a regulated export pathway in health and disease

The endoplasmic reticulum (ER), the first secretory compartment of eukaryotic cells, co-ordinates the biogenesis and export of all membrane-bound and soluble cargo molecules to the cell surface. ER function is now recognised to have unprecedented links with signalling pathways regulating cell growth and differentiation and host physiology. Misfolding and aggregation of newly synthesised proteins in the ER or alterations in ER processing of cargo mediated by pathogens is responsible for a broad range of diseases including cystic fibrosis, emphysema and neuropathies such as Alzheimer's disease. The central, integrative role of the ER in determining cell physiology in health and disease represents an untapped area for pharmacological intervention. This review focuses on the potential use of pharmacological agents to modulate cargo selection, folding and degradation in the ER with the goal of alleviating ER export disease. In addition, implementation of novel technologies that utilise normal ER function to store and release biologically active substances of therapeutic relevance are presented as a new frontier in drug delivery.     

BCPT Nordic Prize in Basic and Clinical Pharmacology and Toxicology for 2012 to Michael J. Mulvany

Oestrogen exerts vasculoprotective effects in different experimental settings through inhibition of vascular smooth muscle cell proliferation, stimulation of nitric oxide production and attenuation of inflammation. Although these oestrogen‐evoked beneficial effects have been attributed to oestrogen receptor alpha (ERα), also ER beta (ERβ) and the novel ER G protein‐coupled receptor 30 (GPR30)/G protein‐coupled ER1 probably play significant roles in vascular oestrogen signalling. Oestrogen‐evoked vasculoprotective effects are well documented in various experimental models, but the underlying mechanisms are still incompletely understood. The age hypothesis represents an interesting and promising model to explain the discrepancy between experimental data showing beneficial vascular effects of oestrogen treatment and the clinical findings on hormone replacement therapy obtained in big epidemiology surveys, where no protective effect from supplementation with oestrogen is observed. Identification of novel ERs expressed also in the vascular system offers exciting opportunities for the future to find and characterize the mechanisms behind oestrogen‐evoked beneficial effects in vascular health and disease. Importantly, some vascular effects of pharmacological concentrations of oestrogen are ER‐independent, suggesting that oestrogen besides its specific effects through ERα, ERβ and GPR30 also affects vascular function via ER‐independent mechanisms probably reflecting interaction of the hydrophobic oestrogen molecule with cell membrane properties. In this MiniReview, we focus on the importance of these different vascular ER subtypes in health and disease.     

Targeted-mitochondria antioxidants therapeutic implications in inflammatory bowel disease

The role of the mitochondria, particularly its implications in cellular redox and oxidative balance is important for cell survival, and is involved in a variety of human diseases including inflammatory bowel disease (IBD). In this review we address mitochondria physiology and reactive oxygen species as cognate to IBD. Several studies have reported altered mitochondria function in intestinal epithelium in IBD. However, mitochondrion significance in this disease has not been much investigated. Finally, we suggest potential for antioxidants targeted to mitochondria in IBD therapy as a treatment strategy.     

内质网应激与神经元退行性变

内质网(endoplasmic reticulum,ER)是蛋白质修饰、折叠和钙贮存的场所。ER内未折叠或错折叠蛋白积聚和钙平衡失调均可导致ER应激。早期的ER应激或未折叠蛋白反应,是一种自身代偿过程,对细胞起到保护作用,而长期、严重的ER应激则会诱导细胞凋亡及死亡。研究发现,ER应激在许多神经退行性疾病的病理机制中起重要作用。然而,确切的机制目前仍不清楚。该文就ER应激在神经元退行性变中的作用作一综述。     

Putative Involvement of Endocrine Disruptors in the Alzheimer's Disease Via the Insulin-Regulated Aminopeptidase/GLUT4 Pathway

It has been well established that there is a connection between type II diabetes (DMTII) and Alzheimer''s disease (AD). In fact, the increase in AD incidence may be an emerging complication of DMTII. Both pathologies are related to estradiol (E₂) exposure; on the one hand, estrogen receptors (ER) are emerging as important modulators of glucose homeostasis through ß-pancreatic cell function; on the other hand, brain bioenergetic and cognitive deficits have been related to the down regulation of brain ERs, contributing to women ageing and AD susceptibility, both related to the reduction in estradiol levels and the deficits in brain metabolism. Here we discuss that environmental contaminants with estrogenic capacity such as bisphenol A (BPA) could develop pharmacological effects similar to those of E₂, which could affect ß-pancreatic cell function by increasing the biosynthesis of glucose-induced insulin after extranuclear ER binding. BPA-induced hyperinsulinemia would promote the translocation of glucose transporter 4 (GLUT4), which is located next to insulin-regulated aminopeptidase (IRAP) in intracellular vesicles. In insulin-responsive tissues, IRAP and GLUT 4 are routed together to the cell surface after insulin stimulation. IRAP is also the angiotensin IV (AngIV) receptor, and AngIV associates the brain renin-angiotensin system (bRAS) with AD, since AngIV is related to learning, memory, emotional responses, and processing of sensory information not only through its inhibitory effect on IRAP but also through the stimulation of glucose uptake by increasing the presence of IRAP/GLUT4 at the cell surface. Thus, the IRAP/GLUT4 pathway is an emerging target for pharmacological intervention against AD.     

Differential regulation of the cell-surface targeting and function of beta- and alpha1-adrenergic receptors by Rab1 GTPase in cardiac myocytes

The molecular mechanism underlying the export from the endoplasmic reticulum (ER) to the cell surface and its role in the regulation of signaling of adrenergic receptors (ARs) remain largely unknown. In this report, we determined the role of Rab1, a Ras-like GTPase that coordinates protein transport specifically from the ER to the Golgi, in the cell surface targeting and function of endogenous beta- and alpha1-ARs in neonatal rat ventricular myocytes. Adenovirus-driven expression of Rab1 into myocytes selectively increased the cell-surface number of alpha1-AR, but not beta-AR, whereas the dominant-negative mutant Rab1N124I significantly reduced the cell-surface expression of beta-AR and alpha1-AR. Brefeldin A inhibited beta-AR and alpha1-AR export and antagonized the Rab1 effect on alpha1-AR expression. Manipulation of Rab1 function similarly influenced the transport of alpha1A- and alpha1B-ARs as well as beta1- and beta2-ARs. Fluorescent microscopy analysis demonstrated that expression of Rab1N124I and Rab1 small interfering RNA induced a marked accumulation of GFP-tagged beta2-AR and alpha1B-AR in the ER. Consistent with the effects on receptor cell-surface targeting, Rab1 selectively enhanced ERK1/2 activation and hypertrophic growth in response to the alpha1-AR agonist phenylephrine but not to the beta-AR agonist isoproterenol. Rab1N124I inhibited both agonist-mediated ERK1/2 activation and hypertrophic growth in neonatal myocytes. These results demonstrate that the cell-surface targeting and signaling of beta- and alpha1-ARs require Rab1 and are differentially modulated by augmentation of Rab1 function. Our data provide strong evidence implicating the ER-to-Golgi traffic as a site for selective manipulation of distinct AR function in cardiac myocytes.     

An evaluation of memantine ER + donepezil for the treatment of Alzheimer’s disease

ABSTRACT

Introduction: Alzheimer’s disease (AD) is the most common neurodegenerative disease worldwide and carries an immense societal burden. Unfortunately, no curative or disease-modifying treatment has yet been discovered. The currently approved medications are symptomatic. They include two classes: the cholinesterase inhibitors, such as donepezil, and the NMDA receptor antagonist memantine. Most evidence has shown that combining both classes is superior to monotherapy but may complicate the treatment regimen for patients and families. Namzaric®, a fixed dose combination of donepezil and memantine extended-release (ER) (FDC memantine ER/donepezil), was recently approved by the U.S. Food and Drug Administration (FDA) for patients with moderate to severe AD and warrants further consideration as a clinically useful and advantageous pharmacotherapy in AD.

Areas covered: This review discusses the pharmacological properties, efficacy, and safety/tolerability data of this FDC memantine ER/donepezil as well as its benefits and disadvantages for patients and families. A literature search using PubMed was conducted using Namzaric, donepezil, memantine, AD, and medication adherence as keywords.

Expert opinion: Aside from its cost, FDC memantine ER/donepezil improves adherence to medication and reduces caregiver burden. It allows patients to benefit from combination therapy as the disease progresses, especially in those with dysphagia, poor adherence and limited caregiver support.     

Protein folding in the endoplasmic reticulum and the unfolded protein response

In all eukaryotic cells, the endoplasmic reticulum (ER) is an intracellular organelle where folding and assembly occurs for proteins destined to the extracellular space, plasma membrane, and the exo/endocytic compartments (Kaufman 1999). As a protein-folding compartment, the ER is exquisitely sensitive to alterations in homeostasis, and provides stringent quality control systems to ensure that only correctly folded proteins transit to the Golgi and unfolded or misfolded proteins are retained and ultimately degraded. A number of biochemical and physiological stimuli, such as perturbation in calcium homeostasis or redox status, elevated secretory protein synthesis, expression of misfolded proteins, sugar/glucose deprivation, altered glycosylation, and overloading of cholesterol can disrupt ER homeostasis, impose stress to the ER, and subsequently lead to accumulation of unfolded or misfolded proteins in the ER lumen. The ER has evolved highly specific signaling pathways called the unfolded protein response (UPR) to cope with the accumulation of unfolded or misfolded proteins. Elucidation of the molecular mechanisms by which accumulation of unfolded proteins in the ER transmits a signal to the cytoplasm and nucleus has led to major new insights into the diverse cellular and physiological processes that are regulated by the UPR. This chapter summarizes how cells respond to the accumulation of unfolded proteins in the cell and the relevance of these signaling pathways to human physiology and disease.     

Controlling protein transport by small molecules

Many proteins are transported from the nucleus to the cytoplasm by the exportin CRM1, which recognizes cargo proteins through a leucine rich nuclear export signal (NES). This nuclear export process can be inhibited by several small molecules, both natural products and fully synthetic compounds. The structural basis for the inhibition of nuclear export by leptomycin (LMB) based on disruption of the protein/protein interaction between CRM1 and cargo proteins is discussed. The chemistry and inhibition of nucleocytoplasmic transport of leptomycin, anguinomycin and derivatives, goniothalamin, JBIR-02, valtrate, dihydrovaltrate, ACA, peumusolide A and several synthetic compounds is presented. Consequences for the design of nuclear export inhibitors are discussed, and the potential of these compounds as anticancer agents is evaluated.     

The failure of mitochondria leads to neurodegeneration: Do mitochondria need a jump start?

Mitochondria are the power engine generating biochemical energy in the cell. Mitochondrial dysfunction and bioenergy deficiency is closely linked to the pathogenesis of neurodegenerative disorders. Mitochondria play a variety of roles by integrating extracellular signals and executing important intracellular events in neuronal survival and death. In this context, the regulation of mitochondrial function via therapeutic approaches may exert some salutary and neuroprotective mechanisms. Understanding the relationship of mitochondria-dependent pathogenesis may provide important pharmacological utility in the treatment of neurodegenerative conditions such as Alzheimer's disease, amyotrophic lateral sclerosis, Huntington's disease and Parkinson's disease. Indeed, the modulation of mitochondrial pathways is rapidly emerging as a novel therapeutic target. This review focuses on how mitochondria are involved in neurodegeneration and what therapeutics are available to target mitochondrial pathways.     

Trabecular meshwork as a new target for the treatment of glaucoma

Glaucoma, a group of optic neuropathies, is one of the leading causes of irreversible blindness in the world. It is characterized by degeneration of the optic nerve and progressive visual field loss, often associated with elevated intraocular pressure (IOP). In primary open-angle glaucoma, the most common form of the disease, IOP occurs as a result of abnormally increased resistance to drainage of aqueous humor through the conventional outflow system, which comprises the trabecular meshwork and the Schlemm's canal. The pharmacological treatment of glaucoma has been classically aimed at lowering elevated IOP, either decreasing the production of aqueous humor or improving its outflow. Increasing knowledge of trabecular meshwork physiology shows that this tissue has unique morphologic and functional properties involved in the regulation of aqueous humor outflow. Although trabecular meshwork physiology is yet to be fully revealed, ion channels involved in cell contractility or cell volume regulation, or those capable of responding to high pressure, have been described and may be considered promising pharmacological targets for the treatment of glaucoma. The cytoskeleton architecture of the trabecular meshwork cell is thought to be an important regulator of aqueous humor outflow. Gene technology directed at discovering genes linked to the development of glaucoma or to those upregulated in response to elevated IOP is challenging research but provides an insight into future gene therapy. New tools to study trabecular meshwork physiology have recently been developed, including the use of lentivirus for gene delivery or fusion proteins with a protein transduction domain. These vectors are targeted specifically to the trabecular meshwork and are powerful techniques with broad applications for future gene therapy or as new forms of drug delivery.     

The endoplasmic reticulum as a potential therapeutic target in nonalcoholic fatty liver disease

The endoplasmic reticulum (ER) has emerged as a key to understanding the development and consequences of hepatic fat accumulation in nonalcoholic fatty liver disease (NAFLD). An essential function of this organelle is the proper assembly of proteins that are destined for intracellular organelles and the cell surface. Recent evidence suggests that chemical chaperones that enhance the functional capacity of the ER improve liver function in obesity and NAFLD. These chaperones may therefore provide a novel potential therapeutic strategy in NAFLD.     

磷酸二酯酶3抑制剂的功能和临床应用

磷酸二酯酶3（PDE3）是cAMP的水解酶,参与人体许多生理活动的调节。磷酸二酯酶3抑制剂升高细胞内cAMP水平,具有强心、血管舒张、抗血小板凝聚、抗增殖等作用,在心力衰竭和间歇性跛行等症的治疗中具有临床价值。     

The role of COX-2 in heart pathology

Cyclooxygenase-2 (COX-2) is a key enzyme in the production of prostaglandins, and an important anti-inflammation drug target. Recent focus has been placed on the role of COX-2 in heart function and pathology, due to the finding that specific COX-2 inhibitors significantly increased the risk of heart disease in chronic users. However, the exact role of COX-2 in cardiac physiology and disease remains controversial due to the conflicting data reported. Roughly equal numbers of reports have shown either a detrimental role or a protective role for COX-2 in heart in experimental models. Here we attempt to provide a background on the more general roles of COX-2 in pathophysiology, as well as molecular mechanisms employed to control COX-2 expression. This background provides a basis for better understanding the functional role of COX-2 in human heart pathologies, based on the results of COX-2 pharmacological inhibitor studies in humans as well as COX-2 expression in human heart disease. Furthermore, we will explore the experimental evidence implicating different intracellular molecular signaling cascades that regulate COX-2 expression in cardiomyocytes. All of this data permits a more mechanistic understanding of the published studies using pharmacological inhibitors of COX-2 in experimental models of heart pathology. Lastly, we will examine the use of genetic manipulation of COX-2 in mice as one of the future avenues in an attempt to resolve the role of COX-2 in cardiac physiology and pathology.     

Molecular approaches to the treatment, prophylaxis, and diagnosis of Alzheimer's disease: endoplasmic reticulum stress and immunological stress in pathogenesis of Alzheimer's disease

Alzheimer's disease (AD) is an age-related neurodegenerative disorder, accompanied by neuronal loss and the formation of senile plaques in the brain. Glial cells, such as microglia, have been shown to be activated and induce chronic inflammatory responses in AD brain. The endoplasmic reticulum (ER) functions to facilitate protein folding. However, ER stress occurs when cells are exposed to stress. Mounting evidence suggests that ER stress is involved in the pathology of AD. Meanwhile, recent findings suggested crosstalk between ER stress and immune function. However, the mechanisms linking the progression of AD with ER and immunological stress are still not clear. In the present paper, we review and discuss recent results regarding the mechanism of AD pathogenesis, focusing on ER stress and immunological stress.     

Nuclear export of proteins and drug resistance in cancer

The intracellular location of a protein is crucial to its normal functioning in a cell. Cancer cells utilize the normal processes of nuclear-cytoplasmic transport through the nuclear pore complex of a cell to effectively evade anti-neoplastic mechanisms. CRM1-mediated export is increased in various cancers. Proteins that are exported in cancer include tumor-suppressive proteins such as retinoblastoma, APC, p53, BRAC1, FOXO proteins, INI1/hSNF5, galectin-3, Bok, nucleophosmin, RASSF2, Merlin, p21(CIP), p27(KIP1), N-WASP/FAK, estradiol receptor and Tob, drug targets topoisomerase I and IIα and BCR-ABL, and the molecular chaperone protein Hsp90. Here, we review in detail the current processes and known structures involved in the export of a protein through the nuclear pore complex. We also discuss the export receptor molecule CRM1 and its binding to the leucine-rich nuclear export signal of the cargo protein and the formation of a nuclear export trimer with RanGTP. The therapeutic potential of various CRM1 inhibitors will be addressed, including leptomycin B, ratjadone, KOS-2464, and specific small molecule inhibitors of CRM1, N-azolylacrylate analogs, FOXO export inhibitors, valtrate, acetoxychavicol acetate, CBS9106, and SINE inhibitors. We will also discuss examples of how drug resistance may be reversed by targeting the exported proteins topoisomerase IIα, BCR-ABL, and galectin-3. As effective and less toxic CRM1 export inhibitors become available, they may be used as both single agents and in combination with current chemotherapeutic drugs. We believe that the future development of low-toxicity, small-molecule CRM1 inhibitors may provide a new approach to treating cancer.     

Selective estrogen receptor-beta (SERM-beta) compounds modulate raphe nuclei tryptophan hydroxylase-1 (TPH-1) mRNA expression and cause antidepressant-like effects in the forced swim test

Estrogen acts through two molecularly distinct receptors termed estrogen receptor alpha (ERα) and estrogen receptor beta (ERβ) which bind estradiol with similar affinities and mediate the effects of estrogen throughout the body. ERα plays a major role in reproductive physiology and behavior, and mediates classic estrogen signaling in such tissues as the uterus, mammary gland, and skeleton. ERβ, however, modulates estrogen signaling in the ovary, the immune system, prostate, gastrointestinal tract, and hypothalamus, and there is some evidence that ERβ can regulate ERα activity. Moreover, ERβ knockout studies and receptor distribution analyses in the CNS suggest that this receptor may play a role in the modulation of mood and cognition. In recent years several ERβ-specific compounds (selective estrogen receptor beta modulators; SERM-beta) have become available, and research suggests potential utility of these compounds in menopausal symptom relief, breast cancer prevention, diseases that have an inflammatory component, osteoporosis, cardiovascular disease, and inflammatory bowel disease, as well as modulation of mood, and anxiety. Here we demonstrate an antidepressant-like effect obtained using two SERM-beta compounds, SERM-beta1 and SERM-beta2. These compounds exhibit full agonist activity at ERβ in a cell based estrogen response element (ERE) transactivation assay. SERM-beta1 and 2 are non-proliferative with respect to breast as determined using the MCF-7 breast cancer cell-based assay and non-proliferative in the uterus as determined by assessing the effects of SERM-beta compounds on immature rat uterine weight and murine uterine weight. In vivo SERM-beta1 and 2 are brain penetrant and display dose dependent efficacy in the murine dorsal raphe assays for induction of tryptophan hydroxylase mRNA and progesterone receptor protein. These compounds show activity in the murine forced swim test and promote hippocampal neurogenesis acutely in rats. Taken together these data suggest that ERβ may play an important role in modulating mood and the ERβ specific compounds described herein will be useful tools for probing the utility of an ERβ agonist for treating neuroendocrine-related mood disturbance and menopausal symptoms.     

Clathrin/AP-2-dependent endocytosis: a novel playground for the pharmacological toolbox?

Endocytosis is a vital process for mammalian cells by which they communicate with their environment, internalize nutrients, hormones, or growth factors, or take up extracellular fluids and particles. The best studied among the various pathways to ingest material from the extracellular side is clathrin/AP-2-mediated endocytosis. The past several years have allowed us to gain unprecedented molecular insights into the role of the heterotetrameric AP-2 adaptor complex as a central protein-protein and protein-lipid interaction hub at the plasmalemma. During the initial stages of clathrin-coated pit formation, AP-2 interacts with phosphoinositides and cargo membrane proteins as well as with a variety of accessory proteins and clathrin to coordinate clathrin coat polymerization with membrane deformation and cargo recruitment. In addition, a growing list of alternative adaptors provides opportunity for clathrin-dependent cargo selective pathways of internalization and endosomal sorting. Many of these interactions are now understood in structural detail and are thus amenable to pharmacological interference. In this review we will summarize our present state of knowledge about AP-2 and its partners in endocytosis and delineate potential strategies for pharmacological manipulations.