http://www.mdpi.com/journal/pharmaceuticals Latest open access articles published in Pharmaceuticals at http://www.mdpi.com/journal/pharmaceuticals http://www.mdpi.com/1424-8247/5/5/529 Cannabidiol (CBD), a major phytocannabinoid constituent of cannabis, is attracting growing attention in medicine for its anxiolytic, antipsychotic, antiemetic and anti-inflammatory properties. However, up to this point, a comprehensive literature review of the effects of CBD in humans is lacking. The aim of the present systematic review is to examine the randomized and crossover studies that administered CBD to healthy controls and to clinical patients. A systematic search was performed in the electronic databases PubMed and EMBASE using the key word “cannabidiol”. Both monotherapy and combination studies (e.g., CBD + ∆9-THC) were included. A total of 34 studies were identified: 16 of these were experimental studies, conducted in healthy subjects, and 18 were conducted in clinical populations, including multiple sclerosis (six studies), schizophrenia and bipolar mania (four studies), social anxiety disorder (two studies), neuropathic and cancer pain (two studies), cancer anorexia (one study), Huntington’s disease (one study), insomnia (one study), and epilepsy (one study). Experimental studies indicate that a high-dose of inhaled/intravenous CBD is required to inhibit the effects of a lower dose of ∆9-THC. Moreover, some experimental and clinical studies suggest that oral/oromucosal CBD may prolong and/or intensify ∆9-THC-induced effects, whereas others suggest that it may inhibit ∆9-THC-induced effects. Finally, preliminary clinical trials suggest that high-dose oral CBD (150–600 mg/d) may exert a therapeutic effect for social anxiety disorder, insomnia and epilepsy, but also that it may cause mental sedation. Potential pharmacokinetic and pharmacodynamic explanations for these results are discussed. http://www.mdpi.com/1424-8247/5/5/529 Mon, 21 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-21 5 5 Review 529 552 1424-8247 Cannabidiol in Humans—The Quest for Therapeutic Targets 2012-05-21 doi: 10.3390/ph5050529 Simon Zhornitsky Stéphane Potvin http://www.mdpi.com/1424-8247/5/5/514 Sphingosine 1-phosphate (S1P), a lysophospholipid mediator, is generated from sphingosine by sphingosine kinases and binds five known cell surface receptors. S1P receptor 1 (S1P1) plays an essential role in lymphocyte egress from secondary lymphoid organs (SLO), as evinced by the inability of lymphocytes to exit from the SLO in mice lacking lymphocytic S1P1. Fingolimod hydrochloride (FTY720) is a first-in-class, orally active, S1P receptor modulator with a structure closely related to sphingosine. FTY720 was first synthesized by chemical modification of a natural product, myriocin. FTY720 is effectively converted to an active metabolite, FTY720 phosphate (FTY720-P) by sphingosine kinases. FTY720-P shows high affinity to 4 of the S1P receptors (S1P1, S1P3, S1P4, and S1P5). In particular, FTY720-P strongly induces internalization and degradation of S1P1, inhibits S1P responsiveness of lymphocytes in the SLO, and acts as a functional antagonist at lymphocytic S1P1. Consequently, FTY720 inhibits S1P1-dependent lymphocyte egress from the SLO to decrease circulation of lymphocytes including autoreactive Th17 cells and is highly effective in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). Because FTY720 shows a superior efficacy in relapsing remitting MS patients compared to intramuscular interferon-β-1a (Avonex®), S1P1 is presumed to be a useful target for the therapy of MS. http://www.mdpi.com/1424-8247/5/5/514 Fri, 18 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-18 5 5 Review 514 528 1424-8247 Sphingosine 1-Phosphate Receptor 1 as a Useful Target for Treatment of Multiple Sclerosis 2012-05-18 doi: 10.3390/ph5050514 Kenji Chiba Kunitomo Adachi http://www.mdpi.com/1424-8247/5/5/493 The use of nucleic acid derived aptamers has rapidly expanded since the introduction of SELEX in 1990. Nucleic acid aptamers have demonstrated their ability to target a broad range of molecules in ways that rival antibodies, but advances have been very uneven for different biochemical classes of targets, and clinical applications have been slow to emerge. What sets different aptamers apart from each other and from rivaling molecular recognition platforms, specifically proteins? What advantages do aptamers as a reagent class offer, and how do the chemical properties and selection procedures of aptamers influence their function? Do the building blocks of nucleic acid aptamers dictate inherent limitations in the nature of molecular targets, and do existing aptamers give us insight in how these challenges might be overcome? This review is written as an introduction for potential endusers of aptamer technology who are evaluating the advantages of aptamers as a versatile, affordable, yet highly expandable platform to target a broad range of biological processes or interactions. http://www.mdpi.com/1424-8247/5/5/493 Fri, 18 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-18 5 5 Review 493 513 1424-8247 Selecting Molecular Recognition. What Can Existing Aptamers Tell Us about Their Inherent Recognition Capabilities and Modes of Interaction? 2012-05-18 doi: 10.3390/ph5050493 Qian Zhang Ralf Landgraf http://www.mdpi.com/1424-8247/5/5/481 The viability of the probiotic strain Lactobacillus plantarum subsp. plantarum, after its passage through simulated gastric and pancreatic juices, was evaluated as function of its pre-growth in a medium containing the known prebiotics pectin or inulin, and was compared to glucose used as control. The presence of pectin or inulin did not markedly affect the growth (10.07 log10 colony forming units/mL and 10.28 log10 colony forming units/mL for pectin and inulin respectively versus 10.42 log10 colony forming units/mL obtained for glucose). Pectin and inulin, in contrast to glucose, induced cell stress resistance against gastrointestinal juices (D log101.5 and 2.4 colony forming units/mL respectively, versus D log10 4.0 for glucose). The data were corroborated by the analysis of the protein pattern following stress treatments which, in the case of microbial cells grown with glucose, revealed a more marked protein degradation after the double passage through simulated gastric and intestinal juices. Inulin stimulated the production of the relevant healthy bio-molecule butyrate, which amount was 30% higher respect of growth in the presence of glucose. Inulin and pectin improved cell DPPH scavenging activity, and an impressive hydrophobicity (35.28% and 34.81%, respectively) was observed with respect to the microbial growth in presence of glucose (3.39%). http://www.mdpi.com/1424-8247/5/5/481 Tue, 15 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-15 5 5 Article 481 492 1424-8247 Biochemical Traits, Survival and Biological Properties of the Probiotic Lactobacillus plantarum Grown in the Presence of Prebiotic Inulin and Pectin as Energy Source 2012-05-15 doi: 10.3390/ph5050481 Filomena Nazzaro Florinda Fratianni Pierangelo Orlando Raffaele Coppola http://www.mdpi.com/1424-8247/5/5/469 Non-valvular atrial fibrillation is a recognized risk factor for stroke and systemic embolism. It has been clearly established that warfarin reduces the risk of stroke and systemic embolism in persons with atrial fibrillation and additional risk factors for stroke. The use of warfarin, however, requires frequent monitoring, and there is great variability in patient response to warfarin. Warfarin interacts with several medications and foods. In addition, warfarin use portends a significant risk of bleeding. For these reasons, warfarin is frequently not prescribed to persons for whom the drug would provide a clear benefit. Over the past decade, attempts have been made to develop drugs that are at least as safe and effective as warfarin for the treatment of atrial fibrillation that do not require monitoring nor have as many interactions. Initial studies of compounds in this regard ultimately failed due to safety concerns, but over the past two years two novel agents have been approved by the United States Food and Drug Association for anticoagulation in non-valvular atrial fibrillation, another drug is under review, and additional compounds are being studied. This article will review the use of warfarin and these new agents in the treatment of non-valvular atrial fibrillation. http://www.mdpi.com/1424-8247/5/5/469 Fri, 04 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-04 5 5 Review 469 480 1424-8247 Newer Anticoagulants for Non-Valvular Atrial Fibrillation 2012-05-04 doi: 10.3390/ph5050469 Joseph M. Harburger Wilbert S. Aronow http://www.mdpi.com/1424-8247/5/5/460 A group of benzimidazole analogs of sildenafil, 3-benzimidazolyl-4-methoxy-phenylsulfonylpiperazines 2–4 and 3-benzimidazolyl-4-methoxy-N,N-dimethyl- benzenesulfonamide (5), were efficiently synthesized. Compounds 2–5 were characterized by NMR and MS and contrary to the reported mass spectra of sildenafil, the spectra of the piperazine-containing compounds 2–4 showed a novel fragmentation pattern leading to an m/z = 316. A mechanism for the formation of this fragment was proposed. http://www.mdpi.com/1424-8247/5/5/460 Thu, 03 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-03 5 5 Article 460 468 1424-8247 Synthesis and Spectroscopic Analysis of Novel 1H-Benzo[d]imidazoles Phenyl Sulfonylpiperazines 2012-05-03 doi: 10.3390/ph5050460 Amjad M. Qandil http://www.mdpi.com/1424-8247/5/5/447 Sub-retinal injection of the common AAV2 pseudotypes frequently results in strong transduction of the retinal pigment epithelium (RPE) as well as the retina itself. This has been of benefit to date in human clinical trials using AAV, where the disease target is in the RPE. However, many mutations predisposing to retinal disease are located in the photoreceptor cells, present in the neural retina and not the RPE; in this case the sub-retinal injection route may cause an effective “loss” of therapeutic AAV to the RPE. The avβ5 integrin receptor is highly expressed on the apical surface of the RPE, and is essential to the daily phagocytosis of the outer segment tips of photoreceptor cells. The transduction efficiency of AAV was tested in the retinas of β5−/− mice lacking this receptor and showing defects in photoreceptor outer segment phagocytosis. Following sub-retinal injection of AAV2/5-eGFP, fluorescence was found to be stronger and more widespread in the neural retina of β5−/− mice compared to wild-types with greatly reduced fluorescence in the RPE. Increased levels of the phagocytic signalling protein MFG-E8, the ligand for the avβ5 integrin receptor, is found to have a moderate inhibitory effect on AAV transduction of the retina. However the opposite effect is found when only the integrin-binding domain of MFG-E8, the RGD (Arginine-Glycine-Aspartic acid) domain, was increased. In this case RGD enhanced AAV-mediated retinal transduction relative to RPE transduction. These results are presented for their relevance for the design of AAV-based retinal gene therapy strategies strategies targeting retinal/photoreceptor cells. http://www.mdpi.com/1424-8247/5/5/447 Wed, 02 May 2012 00:00:00 CEST Pharmaceuticals 2012-05-02 5 5 Article 447 459 1424-8247 The Signalling Role of the avβ5-Integrin Can Impact the Efficacy of AAV in Retinal Gene Therapy 2012-05-02 doi: 10.3390/ph5050447 Therese Cronin Daniel C. Chung Ying Yang Emeline F. Nandrot Jean Bennett http://www.mdpi.com/1424-8247/5/5/417 G-protein coupled receptors (GPCRs) are cell surface proteins responsible for translating >80% of extracellular reception to intracellular signals. The extracellular information in the form of neurotransmitters, peptides, ions, odorants etc is converted to intracellular signals via a wide variety of effector molecules activating distinct downstream signaling pathways. All GPCRs share common structural features including an extracellular N-terminal, seven-transmembrane domains (TMs) linked by extracellular/intracellular loops and the C-terminal tail. Recent studies have shown that most GPCRs function as dimers (homo- and/or heterodimers) or even higher order of oligomers. Protein-protein interaction among GPCRs and other receptor proteins play a critical role in the modulation of receptor pharmacology and functions. Although ~50% of the current drugs available in the market target GPCRs, still many GPCRs remain unexplored as potential therapeutic targets, opening immense possibility to discover the role of GPCRs in pathophysiological conditions. This review explores the existing information and future possibilities of GPCRs as tools in clinical pharmacology and is specifically focused for the role of somatostatin receptors (SSTRs) in pathophysiology of diseases and as the potential candidate for drug discovery. http://www.mdpi.com/1424-8247/5/5/417 Fri, 27 Apr 2012 00:00:00 CEST Pharmaceuticals 2012-04-27 5 5 Review 417 446 1424-8247 Pathophysiology of GPCR Homo- and Heterodimerization: Special Emphasis on Somatostatin Receptors 2012-04-27 doi: 10.3390/ph5050417 Rishi K. Somvanshi Ujendra Kumar http://www.mdpi.com/1424-8247/5/5/405 A simple method for attaching immunoglobulin G (IgG) on the cell surface was successfully developed for enhancing phagocytosis of apoptotic tumor cells (ATCs) by dendritic cells (DCs) ex vivo. By conjugating with a poly(ethylene glycol) (PEG)-lipid, named the biocompatible anchor for the membrane (BAM), arbitrary IgG could be incorporated into the cell membrane. In particular, when IgG-BAM conjugates were prepared at the optimal molar ratio of IgG to BAM (1 to 20), almost all cells were efficiently modified with IgG by treatment with IgG-BAM. This simple method was successfully applied to four types of mammalian cells. Furthermore, treatment of ATCs with the IgG-BAM conjugate increased the phagocytosis ratio of ATCs by DCs two-fold when compared to no treatment. This phagocytosis-enhancing effect was nearly identical to treatment with a tumor-specific IgG. Thus, without employing the tumor-specific IgG, which is difficult to obtain for any tumor cells and is expensive, the present method could opsonize ATC with the use of arbitrary IgG. The results strongly indicate that IgG-BAM treatment represents a promising method for opsonizing ATC with human serum IgG, and that this approach will lead to objective clinical responses in DC vaccines. http://www.mdpi.com/1424-8247/5/5/405 Thu, 26 Apr 2012 00:00:00 CEST Pharmaceuticals 2012-04-26 5 5 Article 405 416 1424-8247 Poly(ethylene glycol)-Lipid-Conjugated Antibodies Enhance Dendritic Cell Phagocytosis of Apoptotic Cancer Cells 2012-04-26 doi: 10.3390/ph5050405 Urara Tomita Satoshi Yamaguchi Yoichiro Sugimoto Satoshi Takamori Teruyuki Nagamune http://www.mdpi.com/1424-8247/5/4/398 Accelerated sinus rhythm is an important side effect of inhaled salbutamol which is especially harmful in patients with chronic obstructive pulmonary disease (COPD) and coronary heart disease (CHD). Cross-over, randomized, open label study design. 20 patients (18 males and two females) with COPD stage II–IV and comorbide CHD NYHA class I–III were included. Spirometry with 400 mg salbutamol inhalation was performed at two consecutive days of the study. Patients in group I were prescribed 5 mg ivabradine per os 3 h before salbutamol inhalation solely on the first day of the study and patients of group II received 5 mg ivabradine only on the second day of the study. Salbutamol caused a significant increase of HR by 5.5 bpm (95% CI 0.8; 10.2, p < 0.03). After ivabradine ingestion salbutamol did not change HR significantly by −2.4 bpm (−7.0; 2.3, p = 0.33). The attenuation of HR elevation by ivabradine was significant, p < 0.01. Salbutamol alone increased FEV1 by 6.0% (2.7; 9.3, p < 0.01). This effect was not impaired by ivabradine (FEV1 increase by 7.7% (2.8; 12.6, p < 0.01 versus baseline, p = 0.5 versus no ivabradine). Ivabradine 5 mg per os prevents heart rate acceleration after inhalation of 400 mg salbutamol. Ivabradine has no impact on lung function in patients with moderate-to-very-severe COPD and CHD comorbidity. http://www.mdpi.com/1424-8247/5/4/398 Mon, 16 Apr 2012 00:00:00 CEST Pharmaceuticals 2012-04-16 5 4 Article 398 404 1424-8247 Ivabradine Prevents Heart Rate Acceleration in Patients with Chronic Obstructive Pulmonary Disease and Coronary Heart Disease after Salbutamol Inhalation 2012-04-16 doi: 10.3390/ph5040398 Rustem Zulkarneev Naufal Zagidullin Guzel Abdrahmanova Uta C. Hoppe Shamil Zagidullin http://www.mdpi.com/1424-8247/5/4/384 Atrial Fibrillation (AF) is the most common sustained arrhythmia and 1/6 strokes is attributed to AF. The cornerstone of treatment remains maintaining sinus rhythm or appropriate ventricular rate control in addition to prevention of stroke. Oral anticoagulation therapy (OAC) with vitamin K antagonists (VKAs) has been the gold standard for almost 50 years and a significant reduction in the risk of stroke in patients with AF has been demonstrated. Nonetheless, only 50% of patients with guideline recommendations for OAC treatment actually receive VKAs and half of these will discontinue therapy within 3 to 5 years with only another half achieving therapeutic ranges more than 50% of the time. The aforementioned limitations in addition with frequent blood monitoring have prompted the development of a series of new OAC therapies. The present review focuses on the current pharmacological management for stroke prevention in patients with AF based on current and emerging evidence. http://www.mdpi.com/1424-8247/5/4/384 Thu, 05 Apr 2012 00:00:00 CEST Pharmaceuticals 2012-04-05 5 4 Review 384 397 1424-8247 Stroke Prevention in Atrial Fibrillation: Latest Clinical Trials and Guidelines 2012-04-05 doi: 10.3390/ph5040384 Luciana Armaganijan Dimpi Patel Cristiano Dietrich Carlos A. Morillo http://www.mdpi.com/1424-8247/5/4/369 The number of children with mild neurodevelopmental disorders, such as autism, has been recently increasing in advanced countries. This increase is probably caused by environmental factors rather than genetic factors, because it is unlikely that genetic mutation rates suddenly increased within a short period. Epigenetics is a mechanism that regulates gene expression, depending not on the underlying DNA sequence but on the chemical modifications of DNA and histone proteins. Because mental stress can alter the epigenetic status in neuronal cells, environmental factors may alter brain function through epigenetic changes. However, one advantage of epigenetic changes is their reversibility. Therefore, diseases due to abnormal epigenetic regulation are theoretically treatable. In fact, several drugs for treating mental diseases are known to have restoring effects on aberrant epigenetic statuses, and a novel therapeutic strategy targeting gene has been developed. In this review, we discuss epigenetic mechanisms of congenital and acquired neurodevelopmental disorders, drugs with epigenetic effects, novel therapeutic strategies for epigenetic diseases, and future perspectives in epigenetic medicine. http://www.mdpi.com/1424-8247/5/4/369 Thu, 05 Apr 2012 00:00:00 CEST Pharmaceuticals 2012-04-05 5 4 Review 369 383 1424-8247 Epigenetic Mechanisms and Therapeutic Perspectives for Neurodevelopmental Disorders 2012-04-05 doi: 10.3390/ph5040369 Takeo Kubota Hirasawa Takae Kunio Miyake http://www.mdpi.com/1424-8247/5/4/353 Biosimilars (or follow-on biologics) are a new class of medicine which enters the market subsequent to a previously approved version. They have demonstrated similarity to innovator biologic products in terms of quality, safety, and efficacy. The EMA has taken the lead in the regulatory approval framework for biosimilar products, and WHO has published guidelines on the evaluation of biosimilars in order to facilitate the global harmonization. Based on EMA and WHO guidelines, many other countries such as Canada, Japan and Korea have also issued their own guidance for evaluating follow-on biologics. The US FDA was authorized to approve follow-on biologics by the BPCI Act passed by the US Congress on March 23, 2010, and has just issued a draft guidance in early 2012. The basic concepts and main principles of approving biosimilars are similar among various nations, notwithstanding some differences in regard to the scope, the choice of reference product, and the data requirement. This article reviews the regulatory approval pathway of biosimilar products in different regions. http://www.mdpi.com/1424-8247/5/4/353 Fri, 30 Mar 2012 00:00:00 CEST Pharmaceuticals 2012-03-30 5 4 Review 353 368 1424-8247 On the Regulatory Approval Pathway of Biosimilar Products 2012-03-30 doi: 10.3390/ph5040353 Jun Wang Shein-Chung Chow http://www.mdpi.com/1424-8247/5/4/339 Objective: Shark type II collagen (SCII) is extracted as a glycoprotein from the cartilage of blue shark (Prionace glauca). We aim to confirm the effects of oral tolerance of SCII on inflammatory and immune responses to the ankle joint of rheumatoid-arthritis rats induced by Complete Freund’s Adjuvant (CFA). Materials and Methods: The onset of rheumatoid arthritis (RA) was observed 14 ± x days after injection of CFA. Rats in the control group were treated with acetic acid by oral administration (0.05 mmol kg−1d−1, days 14–28), while rats in experimental groups were treated by oral administration with SCII (1 or 3 mg kg−1d−1, days 14–28), Tripterygium wilfordii polyglycosidium (TWP) (10 mg kg−1d−1, days 14–28), and bovine type II collagen from US (US-CII) (1 mg kg−1d−1, days 14–28), respectively. The severity of arthritis was evaluated by the articular swelling. The immunological indexes observed included delayed type hypersensitivity (DTH) reaction, the level of interleukins 10 (IL-10) in rat blood serum and morphological characterization. Mixed lymphocyte culture (MLC) was performed to investigate the relationship between T cell apoptosis and specific immune tolerance induced by SCII. Results: Treatment with SCII for 2 weeks significantly attenuated the acute inflammation. The rats orally administrated with SCII at the level of 3 mg kg−1d−1 (SCII 3) and US-CII had decreased DTH reaction compared with rats in control group. Rats treated with SCII 3 had the highest level of IL-10 with 102 pg/mL. SCII with concentration of 10 μg/L could help to significantly enhance level of Fas/Apo-1 in T cell in vitro. The result of histological staining indicated that the recovery of the articular membranes of ankle joint in SCII 3 group was greatly enhanced. Conclusions: Our results suggest that appropriate dose of SCII can not only ameliorate symptoms but also modify the disease process of Complete-Freunds-Adjuvant-induced arthritis. Oral administration of SCII might be a potential candidate as a novel drug for further investigation. http://www.mdpi.com/1424-8247/5/4/339 Wed, 28 Mar 2012 00:00:00 CEST Pharmaceuticals 2012-03-28 5 4 Article 339 352 1424-8247 Oral Administration of Shark Type II Collagen Suppresses Complete Freund’s Adjuvant-Induced Rheumatoid Arthritis in Rats 2012-03-28 doi: 10.3390/ph5040339 Lijuan Chen Bin Bao Nanping Wang Jing Xie Wenhui Wu http://www.mdpi.com/1424-8247/5/3/325 Objective: To compare the effects of dorzolamide and timolol add-on therapy in open-angle glaucoma (OAG) patients previously treated with prostaglandin analogue (Pg), by evaluating fluctuations in the intraocular (IOP), blood (BP), ocular perfusion pressures (OPP) and retrobulbar blood flow (RBF) parameters. Methods: 35 OAG patients (35 eyes), 31 women (88.6%) age 63.3 (8.9) years were evaluated in a 3 month randomized, cross-over, single-masked study. During the experiments BP, heart rate, IOP and OPP were assessed 4 times per day (8–12–16–20 h). RBF was measured twice per day (8–20 h) using Color Doppler imaging in the ophthalmic (OA), central retinal (CRA), nasal (nSPCA) and temporal (tSPCA) posterior ciliary arteries. In each vessel, peak systolic velocity (PSV) and end-diastolic velocity (EDV) were assessed and vascular resistance (RI) calculated. Results: Both add-on therapies lowered IOP in a statistically significant manner from 15.7 ± 2.4 mmHg at latanoprost baseline to 14.9 ± 2.2 mmHg using dorzolamide (p < 0.001) and 14.2 ± 1.9 mmHg using timolol (p < 0.001). The IOP lowering effect was statistically significant at 20 h, favoring timolol as compared to dorzolamide (1.4 ± 2.4 vs. 0.2 ± 2.1 mmHg), (p < 0.05). Dorzolamide add-on therapy showed smaller IOP (2.0 ± 1.4), SPP (13.3 ± 7.9), systolic BP (13.5 ± 8.7) and diastolic BP (8.4 ± 5.4) fluctuations as compared to both latanoprost baseline or timolol add-on therapies. Higher difference between morning and evening BP was correlated to decreased evening CRA EDV in the timolol group (c = −0.41; p = 0.01). With increased MAP in the morning or evening hours, we found increased evening OA RI in timolol add-on group (c = 0.400, p = 0.02; c = 0.513, p = 0.002 accordingly). Higher MAP fluctuations were related to impaired RBF parameters during evening hours-decreased CRA EDV (c = −0.408; p = 0.01), increased CRA RI (c = 0.576; p < 0.001) and tSPCA RI (c = 0.356; p = 0.04) in the dorzolamide group and increased nSPCA RI (c = 0.351; p = 0.04) in the timolol add-on group. OPP fluctuations correlated with increased nSPCA RI (c = 0.453; p = 0.006) in the timolol group. OPP fluctuations were not related to IOP fluctuations in both add-on therapies (p < 0.05). Conclusions: Both dorzolamide and timolol add-on therapies lowered IOP in a statistically significant fashion dorzolamide add-on therapy showed lower fluctuations in IOP, SPP and BP. Higher variability of daytime OPP led to impaired RBF parameters in the evening. http://www.mdpi.com/1424-8247/5/3/325 Wed, 21 Mar 2012 00:00:00 CET Pharmaceuticals 2012-03-21 5 3 Article 325 338 1424-8247 Comparison of Intraocular Pressure, Blood Pressure, Ocular Perfusion Pressure and Blood Flow Fluctuations During Dorzolamide Versus Timolol Add-On Therapy in Prostaglandin Analogue Treated Glaucoma Subjects 2012-03-21 doi: 10.3390/ph5030325 Ingrida Januleviciene Lina Siaudvytyte Vaida Diliene Ruta Barsauskaite Daiva Paulaviciute-Baikstiene Brent Siesky Alon Harris http://www.mdpi.com/1424-8247/5/3/317 The main purpose of this paper is to provide an insight into the biological activities of pyrazole derivatives which contain the carbohydrazide moiety. http://www.mdpi.com/1424-8247/5/3/317 Fri, 16 Mar 2012 00:00:00 CET Pharmaceuticals 2012-03-16 5 3 Review 317 324 1424-8247 Pyrazole Carbohydrazide Derivatives of Pharmaceutical Interest 2012-03-16 doi: 10.3390/ph5030317 Luiza Rosaria Sousa Dias Raquel Rocha Silva Salvador http://www.mdpi.com/1424-8247/5/3/297 This minireview provides a brief outline of the peculiar aspects of the preparation of peptidomimetic and pseudopeptidic structures containing heterocycles. In particular novel tricyclic structures are investigated as potential drugs. http://www.mdpi.com/1424-8247/5/3/297 Thu, 08 Mar 2012 00:00:00 CET Pharmaceuticals 2012-03-08 5 3 Review 297 316 1424-8247 Heterocycles in Peptidomimetics and Pseudopeptides: Design and Synthesis 2012-03-08 doi: 10.3390/ph5030297 Iole Cerminara Lucia Chiummiento Maria Funicello Ambra Guarnaccio Paolo Lupattelli http://www.mdpi.com/1424-8247/5/3/279 Dual antiplatelet therapy with acetylsalicylic acid and a thienopyridine, such as clopidogrel, is effective for the secondary prevention of cardiovascular events in patients with acute coronary syndrome, but there is still a substantial residual risk of recurrence. Although anticoagulant therapy with a vitamin K antagonist (e.g. warfarin) in conjunction with antiplatelet therapy has been shown to reduce the risk of cardiovascular events, the rates of bleeding were increased with these combination therapies; hence, triple therapy with warfarin is currently only recommended in patients at low risk of bleeding. In addition, there are other limitations associated with vitamin K antagonist therapy, including the need for routine coagulation monitoring and dose adjustment to maintain the treatment within the therapeutic range. Rivaroxaban is an oral, direct Factor Xa inhibitor; in clinical practice, it is likely that rivaroxaban will be given to patients who also receive antiplatelet therapy, such as clopidogrel. This randomized, non-blinded, three-way crossover study investigated the effect of rivaroxaban on bleeding time when co­administered with clopidogrel. In addition, the influence of clopidogrel on the safety, tolerability, pharmacodynamics and pharmacokinetics of rivaroxaban was investigated. Of 27 healthy male subjects who received a single 300 mg dose of clopidogrel, 14 were identified as clopidogrel responders and were then randomized to the following three treatments: (A) two doses of clopidogrel on two consecutive days (300 mg on day 1; 75 mg on day 2); (B) one dose of rivaroxaban (15 mg); or (C) a combination of treatments A and B (rivaroxaban given on day 2). All treatments were well tolerated. Bleeding time with co­administration of rivaroxaban and clopidogrel was significantly prolonged in four subjects, compared with either drug alone: combination treatment increased the overall least squares-means to 3.77 times baseline (90% confidence interval [CI] 2.82–4.73), compared with 1.13 times baseline (90% CI 0.17–2.09) with rivaroxaban and 1.96 times baseline (90% CI 0.10–2.91) with clopidogrel. Co-administration of clopidogrel had no significant effect on the pharmacokinetics of rivaroxaban and, when compared with rivaroxaban alone, had no further effects on Factor Xa activity or prothrombin time. Inhibition of ADP-stimulated platelet aggregation by clopidogrel was not affected by rivaroxaban. As expected, owing to the mode of action of each study drug, the results of this study demonstrated that co­administration of the Factor Xa inhibitor rivaroxaban and the antiplatelet clopidogrel increased the bleeding time in healthy subjects without affecting other pharmacokinetic or pharmacodynamic parameters of each drug. http://www.mdpi.com/1424-8247/5/3/279 Fri, 24 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-24 5 3 Article 279 296 1424-8247 Effect of Co-Administration of Rivaroxaban and Clopidogrel on Bleeding Time, Pharmacodynamics and Pharmacokinetics: A Phase I Study 2012-02-24 doi: 10.3390/ph5030279 Dagmar Kubitza Michael Becka Wolfgang Mück Stephan Schwers http://www.mdpi.com/1424-8247/5/3/249 Recent discoveries indicate that many G-protein coupled receptors (GPCRs) and channels involved in pain modulation are able to form receptor heteromers. Receptor and channel heteromers often display distinct signaling characteristics, pharmacological properties and physiological function in comparison to monomer/homomer receptor or ion channel counterparts. It may be possible to capitalize on such unique properties to augment therapeutic efficacy while minimizing side effects. For example, drugs specifically targeting heteromers may have greater tissue specificity and analgesic efficacy. This review will focus on current progress in our understanding of roles of heteromeric GPCRs and channels in pain pathways as well as strategies for controlling pain pathways via targeting heteromeric receptors and channels. This approach may be instrumental in the discovery of novel classes of drugs and expand our repertoire of targets for pain pharmacotherapy. http://www.mdpi.com/1424-8247/5/3/249 Thu, 23 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-23 5 3 Review 249 278 1424-8247 Receptor and Channel Heteromers as Pain Targets 2012-02-23 doi: 10.3390/ph5030249 Kelly A. Berg Amol M. Patwardhan Armen N. Akopian http://www.mdpi.com/1424-8247/5/2/236 Probiotics possess potential therapeutic and preventative effects for various diseases and metabolic disorders. One important limitation for the oral delivery of probiotics is the harsh conditions of the upper gastrointestinal tract (GIT) which challenge bacterial viability and activity. One proposed method to surpass this obstacle is the use of microencapsulation to improve the delivery of bacterial cells to the lower GIT. The aim of this study is to use alginate-poly-L-lysine-alginate (APA) microcapsules to encapsulate Lactobacillus fermentum NCIMB 5221 and characterize its enzymatic activity and viability through a simulated GIT. This specific strain, in previous research, was characterized for its inherent ferulic acid esterase (FAE) activity which could prove beneficial in the development of a therapeutic for the treatment and prevention of cancers and metabolic disorders. Our findings demonstrate that the APA microcapsule does not slow the mass transfer of substrate into and that of the FA product out of the microcapsule, while also not impairing bacterial cell viability. The use of simulated gastrointestinal conditions led to a significant 2.5 log difference in viability between the free (1.10 × 104 ± 1.00 × 103 cfu/mL) and the microencapsulated (5.50 × 106 ± 1.00 × 105 cfu/mL) L. fermentum NCIMB 5221 following exposure. The work presented here suggests that APA microencapsulation can be used as an effective oral delivery method for L. fermentum NCIMB 5221, a FAE-active probiotic strain. http://www.mdpi.com/1424-8247/5/2/236 Thu, 16 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-16 5 2 Article 236 248 1424-8247 Probiotic Ferulic Acid Esterase Active Lactobacillus fermentum NCIMB 5221 APA Microcapsules for Oral Delivery: Preparation and in Vitro Characterization 2012-02-16 doi: 10.3390/ph5020236 Catherine Tomaro-Duchesneau Shyamali Saha Meenakshi Malhotra Michael Coussa-Charley Imen Kahouli Mitchell L. Jones Alain Labbé Satya Prakash http://www.mdpi.com/1424-8247/5/2/209 Biologics such as monoclonal antibodies (mAb) and soluble receptors represent new classes of therapeutic agents for treatment of several diseases. High affinity and high specificity biologics can be utilized for variety of clinical purposes. Monoclonal antibodies have been used as diagnostic agents when coupled with radionuclide, immune modulatory agents or in the treatment of cancers. Among other limitations of using large molecules for therapy the actual cost of biologics has become an issue. There is an effort among chemists and biologists to reduce the size of biologics which includes monoclonal antibodies and receptors without a reduction of biological efficacy. Single chain antibody, camel antibodies, Fv fragments are examples of this type of deconstructive process. Small high-affinity peptides have been identified using phage screening. Our laboratory used a structure-based approach to develop small-size peptidomimetics from the three-dimensional structure of proteins with immunoglobulin folds as exemplified by CD4 and antibodies. Peptides derived either from the receptor or their cognate ligand mimics the functions of the parental macromolecule. These constrained peptides not only provide a platform for developing small molecule drugs, but also provide insight into the atomic features of protein-protein interactions. A general overview of the reduction of monoclonal antibodies to small exocyclic peptide and its prospects as a useful diagnostic and as a drug in the treatment of cancer are discussed. http://www.mdpi.com/1424-8247/5/2/209 Thu, 16 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-16 5 2 Review 209 235 1424-8247 Structure Based Antibody-Like Peptidomimetics 2012-02-16 doi: 10.3390/ph5020209 Ramachandran Murali Mark I. Greene http://www.mdpi.com/1424-8247/5/2/189 Scrophularia ningpoensis has been used in China for centuries as a herbal tea to treat various diseases. Based on the numerous animal studies on its pharmaceutical effects and the long time clinical experiences, we studied the molecular and cellular mechanism underlying the bioactivity of aqueous extract of Scrophularia and its isolated compounds. Seven isolated compounds, unlike Scrophularia extract, failed to induce cytotoxicity on HaCaT cells, but their combination improved the effect of extract. Tumor cell line selectivity was not observed, when we studied its cytotoxic effect on melanoma cell lines. The apoptotic and anti-inflammatory effects of Scrophularia extract have been demonstrated on HaCaT cells. The extract induced those effects potentially through affecting the MAPK pathway and inhibition of the NF-κB pathway, Microarray-based bioinformatical analyses on the compound acetoside from Scrophularia revealed a gene expression profile which confirmed our findings with the extract on proliferation inhibition, anti-inflammation and apoptosis. With DNA alkylation as major proposed mechanism of action, we assume acetoside as one of the active compounds in Scrophularia. http://www.mdpi.com/1424-8247/5/2/189 Tue, 14 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-14 5 2 Article 189 208 1424-8247 Effects of Scrophularia ningpoensis Hemsl. on Inhibition of Proliferation, Apoptosis Induction and NF-κB Signaling of Immortalized and Cancer Cell Lines 2012-02-14 doi: 10.3390/ph5020189 Xiao Shen Tolga Eichhorn Henry Johannes Greten Thomas Efferth http://www.mdpi.com/1424-8247/5/2/169 Phosphodiesterase 10A (PDE10A) is a key enzyme of intracellular signal transduction which is involved in the regulation of neurotransmission. The molecular imaging of PDE10A by PET is expected to allow a better understanding of physiological and pathological processes related to PDE10A expression and function in the brain. The aim of this study was to develop a new 18F-labeled PDE10A ligand based on a 6,7-dimethoxy-4-pyrrolidinylquinazoline and to evaluate its properties in biodistribution studies. Nucleophilic substitution of the 7-tosyloxy-analogue led to the 7-[18F]fluoroethoxy-derivative [18F]IV with radiochemical yields of 25% ± 9% (n = 9), high radiochemical purity of ≥99% and specific activities of 110–1,100 GBq/μmol. [18F]IV showed moderate PDE10A affinity (KD,PDE10A = 14 nM) and high metabolic stability in the brain of female CD-1 mice, wherein the radioligand entered rapidly with a peak uptake of 2.3% ID/g in striatum at 5 min p.i. However, ex vivo autoradiographic and in vivo blocking studies revealed no target specific accumulation and demonstrated [18F]IV to be inapplicable for imaging PDE10A with PET. http://www.mdpi.com/1424-8247/5/2/169 Mon, 06 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-06 5 2 Article 169 188 1424-8247 Radiosynthesis and Radiotracer Properties of a 7-(2-[18F]Fluoroethoxy)-6-methoxypyrrolidinylquinazoline for Imaging of Phosphodiesterase 10A with PET 2012-02-06 doi: 10.3390/ph5020169 Uta Funke Winnie Deuther-Conrad Gregor Schwan Aurélie Maisonial Matthias Scheunemann Steffen Fischer Achim Hiller Detlef Briel Peter Brust http://www.mdpi.com/1424-8247/5/2/155 Dabigatran is an oral thrombin inhibitor which has been approved in several countries as an alternative to vitamin-K-antagonists for the prevention of stroke or embolism in atrial fibrillation patients. Dabigatran is introduced into clinical practice, although many issues regarding this drug are still unclear, like laboratory monitoring, use in elderly patients, drug- and food-interactions and use in patients with renal insufficiency. Additionally, there is no antidote for dabigatran. Thus, aim of the present review is to give an overview of concerns and unresolved issues concerning dabigatran. http://www.mdpi.com/1424-8247/5/2/155 Fri, 03 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-03 5 2 Review 155 168 1424-8247 Concerns Regarding the Use of Dabigatran for Stroke Prevention in Atrial Fibrillation 2012-02-03 doi: 10.3390/ph5020155 Claudia Stöllberger Josef Finsterer http://www.mdpi.com/1424-8247/5/2/133 The in vitro shoot culture of a T-DNA insertional mutant of Papaver somniferum L. established by the infection of Agrobacterium rhizogenes MAFF03-01724 accumulated thebaine instead of morphine as a major opium alkaloid. To develop a non-narcotic opium poppy and to gain insight into its genetic background, we have transplanted this mutant to soil, and analyzed its alkaloid content along with the manner of inheritance of T-DNA insertion loci among its selfed progenies. In the transplanted T0 primary mutant, the opium (latex) was found to be rich in thebaine (16.3% of dried opium) by HPLC analysis. The analyses on T-DNA insertion loci by inverse PCR, adaptor-ligation PCR, and quantitative real-time PCR revealed that as many as 18 copies of T-DNAs were integrated into a poppy genome in a highly complicated manner. The number of copies of T-DNAs was decreased to seven in the selected T3 progenies, in which the average thebaine content was 2.4-fold that of the wild type plant. This may indicate that the high thebaine phenotype was increasingly stabilized as the number of T-DNA copies was decreased. In addition, by reverse transcription PCR analysis on selected morphine biosynthetic genes, the expression of codeine 6-O-demethylase was clearly shown to be diminished in the T0 in vitro shoot culture, which can be considered as one of the key factors of altered alkaloid composition. http://www.mdpi.com/1424-8247/5/2/133 Thu, 02 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-02 5 2 Article 133 154 1424-8247 Genetic and Phenotypic Analyses of a Papaver somniferum T-DNA Insertional Mutant with Altered Alkaloid Composition 2012-02-02 doi: 10.3390/ph5020133 Noriaki Kawano Fumiyuki Kiuchi Nobuo Kawahara Kayo Yoshimatsu http://www.mdpi.com/1424-8247/5/2/114 In the last decade, considerable evidence as accumulated to support the development of Transient Receptor Potential Vanilloid 1 (TRPV1) antagonists for the treatment of various chronic pain conditions. Whereas there is a widely accepted rationale for the development of TRPV1 antagonists for the treatment of various inflammatory pain conditions, their development for indications of chronic pain, where conditions of tactical, mechanical and spontaneous pain predominate, is less clear. Preclinical localization and expression studies provide a firm foundation for the use of molecules targeting TRPV1 for conditions of bone pain, osteoarthritis and neuropathic pain. Selective TRPV1 antagonists weakly attenuate tactile and mechanical hypersensivity and are partially effective for behavioral and electrophysiological endpoints that incorporate aspects of spontaneous pain. While initial studies with TRPV1 antagonist in normal human subjects indicate a loss of warm thermal perception, clinical studies assessing allelic variants suggests that TRPV1 may mediate other sensory modalities under certain conditions. The focus of this review is to summarize the current perspectives of TRPV1 for the treatment of conditions beyond those with a primary thermal sensitivity. http://www.mdpi.com/1424-8247/5/2/114 Thu, 02 Feb 2012 00:00:00 CET Pharmaceuticals 2012-02-02 5 2 Review 114 132 1424-8247 TRPV1 Antagonists and Chronic Pain: Beyond Thermal Perception 2012-02-02 doi: 10.3390/ph5020114 Michael R. Brandt Chad E. Beyer Stephen M. Stahl http://www.mdpi.com/1424-8247/5/1/94 A recent expansion of our knowledge about epigenetic changes strongly suggests that epigenetic rather than genetic features better reflect disease development, and consequently, can become more conclusive biomarkers for the detection and diagnosis of different diseases. In this paper we will concentrate on the current advances in DNA methylation studies that demonstrate a direct link between abnormal DNA methylation and a disease. This link can be used to develop diagnostic biomarkers that will precisely identify a particular disease. It also appears that disease-specific DNA methylation patterns undergo unique changes in response to treatment with a particular drug, thus raising the possibility of DNA methylation-based biomarkers for the monitoring of treatment efficacy, for prediction of response to treatment, and for the prognosis of outcome. While biomarkers for oncology are the most obvious applications, other fields of medicine are likely to benefit as well. This potential is demonstrated by DNA methylation-based biomarkers for neurological and psychiatric diseases. A special requirement for a biomarker is the possibility of longitudinal testing. In this regard cell-free circulating DNA from blood is especially interesting because it carries methylation markers specific for a particular disease. Although only a few DNA methylation-based biomarkers have attained clinical relevance, the ongoing efforts to decipher disease-specific methylation patterns are likely to produce additional biomarkers for detection, diagnosis, and monitoring of different diseases in the near future. http://www.mdpi.com/1424-8247/5/1/94 Wed, 18 Jan 2012 00:00:00 CET Pharmaceuticals 2012-01-18 5 1 Review 94 113 1424-8247 DNA Methylation as Clinically Useful Biomarkers—Light at the End of the Tunnel 2012-01-18 doi: 10.3390/ph5010094 Victor V. Levenson Anatoliy A. Melnikov http://www.mdpi.com/1424-8247/5/1/79 The human epidermal growth factor receptor 2 (HER2/neu) is overexpressed in 20–30% of breast cancers and is associated with tumor growth, angiogenesis, and development of distant metastases. Trastuzumab, an anti-HER2 monoclonal antibody, is used for the treatment of HER2 positive breast cancer and clinical efficacy of this agent is dependent on HER2 expression. Targeted PET imaging of HER2 with radiolabeled trastuzumab may be used to determine HER2 expression levels and guide therapy selection. The purpose of the current study was to evaluate a facile 89Zr-trastuzumab preparation method that can be efficiently applied for clinical grade production. Also, relative HER2 expression levels in orthotopic and metastatic breast cancer models were assessed by PET imaging using the 89Zr-trastuzumab produced by this simpler method. http://www.mdpi.com/1424-8247/5/1/79 Thu, 05 Jan 2012 00:00:00 CET Pharmaceuticals 2012-01-05 5 1 Article 79 93 1424-8247 89Zr-Radiolabeled Trastuzumab Imaging in Orthotopic and Metastatic Breast Tumors 2012-01-05 doi: 10.3390/ph5010079 Albert J. Chang Ravindra DeSilva Sandeep Jain Kimberley Lears Buck Rogers Suzanne Lapi http://www.mdpi.com/1424-8247/5/1/61 Cyclodextrin (CyD)-based nanoparticles and polyamidoamine (PAMAM) starburst dendrimers (dendrimers) are used as novel carriers for DNA and RNA. Recently, small interfering RNA (siRNA) complex with β-CyD-containing polycations (CDP) having adamantine-PEG or adamantine-PEG-transferrin underwent a phase I study for treatment of solid tumors. Multifunctional dendrimers can be used for a wide range of biomedical applications, including the interaction and intracellular delivery of DNA and RNA. The present review will address the latest developments in dendrimer conjugates with cyclodextrins for siRNA delivery including the novel sustained release system. http://www.mdpi.com/1424-8247/5/1/61 Fri, 30 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-30 5 1 Review 61 78 1424-8247 Potential Use of Polyamidoamine Dendrimer Conjugates with Cyclodextrins as Novel Carriers for siRNA 2011-12-30 doi: 10.3390/ph5010061 Hidetoshi Arima Keiichi Motoyama Taishi Higashi http://www.mdpi.com/1424-8247/5/1/49 Hepatitis C virus (HCV) replication is dependent on the existence of several highly conserved functional genomic RNA domains. The cis-acting replication element (CRE), located within the 3' end of the NS5B coding region of the HCV genome, has been shown essential for efficient viral replication. Its sequence and structural features determine its involvement in functional interactions with viral RNA-dependent RNA polymerase and distant RNA domains of the viral genome. This work reports the use of an in vitro selection strategy to select aptamer RNA molecules against the complete HCV-CRE. After six selection cycles, five potential target sites were identified within this domain. Inhibition assays using a sample of representative aptamers showed that the selected RNAs significantly inhibit the replication (>80%) of a subgenomic HCV replicon in Huh-7 cell cultures. These results highlight the potential of aptamer RNA molecules as therapeutic antiviral agents. http://www.mdpi.com/1424-8247/5/1/49 Wed, 28 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-28 5 1 Article 49 60 1424-8247 Anti-HCV RNA Aptamers Targeting the Genomic cis-Acting Replication Element 2011-12-28 doi: 10.3390/ph5010049 Soledad Marton Beatriz Berzal-Herranz Eva Garmendia Francisco J. Cueto Alfredo Berzal-Herranz http://www.mdpi.com/1424-8247/5/1/16 One approach to develop successful pain therapies is the modulation of dysfunctional ion channels that contribute to the detection of thermal, mechanical and chemical painful stimuli. These ion channels, known as thermoTRPs, promote the sensitization and activation of primary sensory neurons known as nociceptors. Pharmacological blockade and genetic deletion of thermoTRP have validated these channels as therapeutic targets for pain intervention. Several thermoTRP modulators have progressed towards clinical development, although most failed because of the appearance of unpredicted side effects. Thus, there is yet a need to develop novel channel modulators with improved therapeutic index. Here, we review the current state-of-the art and illustrate new pharmacological paradigms based on TRPV1 that include: (i) the identification of activity-dependent modulators of this thermoTRP channel; (ii) the design of allosteric modulators that interfere with protein-protein interaction involved in the functional coupling of stimulus sensing and gate opening; and (iii) the development of compounds that abrogate the inflammation-mediated increase of receptor expression in the neuronal surface. These new sites of action represent novel strategies to modulate pathologically active TRPV1, while minimizing an effect on the TRPV1 subpopulation involved in physiological and protective roles, thus increasing their potential therapeutic use. http://www.mdpi.com/1424-8247/5/1/16 Tue, 27 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-27 5 1 Review 16 48 1424-8247 New Strategies to Develop Novel Pain Therapies: Addressing Thermoreceptors from Different Points of View 2011-12-27 doi: 10.3390/ph5010016 Asia Fernández-Carvajal Gregorio Fernández-Ballester Isabel Devesa José Manuel González-Ros Antonio Ferrer-Montiel http://www.mdpi.com/1424-8247/5/1/1 Studies from this laboratory have demonstrated the potential of targeting HER2 for therapeutic and imaging applications with medically relevant radionuclides. To expand the repertoire of trastuzumab as a radioimmunoconjugate (RIC) vector, use of 177Lu was investigated. The combination of a 6.7 d half-life, lower energy β−-emissions (500 keV max; 130 keV ave), and an imagable γ-emission make 177Lu an attractive candidate for radioimmunotherapy (RIT) regimens for treatment of larger tumor burdens not possible with α-particle radiation. Radiolabeling trastuzumab-CHX-A″-DTPA with 177Lu was efficient with a specific binding of 60.8 ± 6.8% with HER2 positive SKOV-3 cells. Direct quantitation of tumor targeting and normal tissue uptake was performed with athymic mice bearing subcutaneous and intraperitoneal LS-174T xenografts; a peak tumor %ID/g of 24.70 ± 10.29 (96 h) and 31.70 ± 16.20 (72 h), respectively, was obtained. Normal tissue uptake of the RIC was minimal. Tumor targeting was also demonstrated by γ-scintigraphy. A therapy study administering escalating doses of 177Lu-trastuzumab to mice bearing three day LS-174T i.p. xenografts established the effective therapeutic dose of i.p. administered 177Lu-trastuzumab at 375 μCi with a median survival of 124.5 d while a median survival of 10 d was noted for the control (untreated) group. In conclusion, trastuzumab radiolabeled with 177Lu has potential for treatment of disseminated, HER2 positive, peritoneal disease. http://www.mdpi.com/1424-8247/5/1/1 Thu, 22 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-22 5 1 Article 1 15 1424-8247 Pre-Clinical Assessment of 177Lu-Labeled Trastuzumab Targeting HER2 for Treatment and Management of Cancer Patients with Disseminated Intraperitoneal Disease 2011-12-22 doi: 10.3390/ph5010001 Geoffrey L. Ray Kwamena E. Baidoo Lanea M. M. Keller Paul S. Albert Martin W. Brechbiel Diane E. Milenic http://www.mdpi.com/1424-8247/4/12/1591 RNA interference (RNAi) allows the specific knockdown of tumor relevant genes. To induce RNAi, the delivery of small interfering RNAs (siRNAs) is of crucial importance. This is particularly challenging for their therapeutic applications in vivo. Low molecular weight branched polyethylenimine (PEI) is safe and efficient for nucleic acid delivery including small RNA molecules, based on its ability to electrostatically complex siRNA molecules, thereby protecting them from nuclease degradation. The nanoscale PEI/siRNA complexes are endocytosed by cells prior to intracellular complex release from the lysosome and cytoplasmic release of the siRNAs from the complexes. Chemical modification and ligand decoration of the complexes aim at introducing target tissue specificity and further increased efficacy of PEI-mediated siRNA delivery. CRM197 is a mutated, non-toxic diphtheria toxin (DT) that binds to the membrane-bound precursor of HB-EGF-like growth factor/diphtheria toxin receptor highly expressed in glioblastoma cells. Likewise, the growth factor pleiotrophin (PTN/HB-GAM/HARP) is overexpressed in glioblastoma and is rate limiting for tumor growth, thus representing an attractive target gene for therapeutic knockdown approaches. PEGylation of PEI was performed to reduce the surface charge, and by CRM197 coupling we prepared a modified PEI for siRNA delivery into glioblastoma cells. The novel PEI conjugates were analyzed for their complexation efficiency and optimal mixing ratios, and complexes were physicochemically characterized regarding stability, size and zeta potential. The biological activity of the complexes was confirmed in cell culture by reporter gene knockdown. For the therapeutic treatment of subcutaneous human gliobastoma xenografts in athymic nude mice, we systemically injected the modified PEI/siRNA complexes targeting PTN. Antitumor effects based on PTN knockdown demonstrated the advantage of tumor-targeted CRM197-PEG-PEI/siRNA over untargeted PEG-PEI polyplexes. Thus, we establish targeted CRM197-PEG-PEI-based complexes for siRNA delivery in vivo, and show therapeutic effects of CRM197-PEG-PEI/siRNA-mediated knockdown of PTN. http://www.mdpi.com/1424-8247/4/12/1591 Fri, 16 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-16 4 12 Article 1591 1606 1424-8247 Targeted CRM197-PEG-PEI/siRNA Complexes for Therapeutic RNAi in Glioblastoma 2011-12-16 doi: 10.3390/ph4121591 Sabrina Höbel Chantal C.M. Appeldoorn Pieter J. Gaillard Achim Aigner http://www.mdpi.com/1424-8247/4/12/1578 Friedreich’s ataxia (FRDA) is caused by transcriptional repression of the nuclear FXN gene encoding the essential mitochondrial protein frataxin. Based on the hypothesis that the acetylation state of the histone proteins is responsible for gene silencing in FRDA, previous work in our lab identified a first generation of HDAC inhibitors (pimelic o-aminobenzamides), which increase FXN mRNA in lymphocytes from FRDA patients. Importantly, these compounds also function in a FRDA mouse model to increase FXN mRNA levels in the brain and heart. While the first generation of HDAC inhibitors hold promise as potential therapeutics for FRDA, they have two potential problems: less than optimal brain penetration and metabolic instability in acidic conditions. Extensive optimization focusing on modifying the left benzene ring, linker and the right benzene ring lead to a novel class of HDAC inhibitors that have optimized pharmacological properties (increased brain penetration and acid stability) compared to the previous HDAC inhibitors. This article will describe the chemical synthesis and pharmacological properties of these new HDAC inhibitors. http://www.mdpi.com/1424-8247/4/12/1578 Wed, 14 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-14 4 12 Article 1578 1590 1424-8247 Improved Histone Deacetylase Inhibitors as Therapeutics for the Neurodegenerative Disease Friedreich’s Ataxia: A New Synthetic Route 2011-12-14 doi: 10.3390/ph4121578 Chunping Xu Elisabetta Soragni Vincent Jacques James R. Rusche Joel M. Gottesfeld http://www.mdpi.com/1424-8247/4/12/1551 Angiogenesis, a neovascularization process induced from the existing parent blood vessels, is a prerequisite for many physiological and pathological conditions. Under physiological conditions it is regulated by a balance between endogenous angioinhibitors and angioactivators, and an imbalance between them would lead to pathological conditions such as cancer, age-related macular degeneration (AMD), diabetic retinopathy, cardiovascular diseases, etc. Several proteolytically generated endogenous molecules have been identified which exhibit angioinhibition and/or antitumor activities. These angioinhibitors interact with endothelial and tumor cells by binding to distinct integrins and initiate many of their intracellular signaling mechanisms regulating the cell survival and or apoptotic pathways. The present review will focus on the extracellular matrix derived angioinhibitors, and their mechanisms of actions that point to the clinical significance and therapeutic implications. http://www.mdpi.com/1424-8247/4/12/1551 Fri, 02 Dec 2011 00:00:00 CET Pharmaceuticals 2011-12-02 4 12 Review 1551 1577 1424-8247 Proteolytically Derived Endogenous Angioinhibitors Originating from the Extracellular Matrix 2011-12-02 doi: 10.3390/ph4121551 Chandra Shekhar Boosani Yakkanti A. Sudhakar http://www.mdpi.com/1424-8247/4/12/1535 Eucalyptus cinerea, known as silver dollar tree, has few descriptions in traditional medicine. Chemical composition and antimicrobial properties of the essential oils of leaves, flowers and fruits, collected seasonally, were determined by GC/MS and disk diffusion/MIC, respectively. 1,8-Cineole was the main compound, particularly in fresh leaves—Spring (74.98%), dried leaves—Spring (85.32%), flowers—Winter (78.76%) and fruits—Winter (80.97%). Other compounds were found in the aerial parts in all seasons: α-pinene (2.41% to 10.13%), limonene (1.46% to 4.43%), α-terpineol (1.73% to 11.72%), and α-terpinyl acetate (3.04% to 20.44%). The essential oils showed antimicrobial activities against bacteria and yeasts, with the best results being found for the dried autumn and winter leaves oils (MIC < 0.39 mg/mL) against Streptococcus pyogenes. For the other tested microorganisms the following MIC results were found: Staphylococcus aureus—Dried leaves oil from summer (0.78 mg/mL), Pseudomonas aeruginosa—Flowers oil from autumn and fruits oil from winter (1.56 mg/mL) and Candida albicans—Flowers oil from autumn and fruits oils from winter and spring (0.78 mg/mL). http://www.mdpi.com/1424-8247/4/12/1535 Fri, 25 Nov 2011 00:00:00 CET Pharmaceuticals 2011-11-25 4 12 Article 1535 1550 1424-8247 Essential Oils from Different Plant Parts of Eucalyptus cinerea F. Muell. ex Benth. (Myrtaceae) as a Source of 1,8-Cineole and Their Bioactivities 2011-11-25 doi: 10.3390/ph4121535 Sayonara Mendes Silva Simone Yae Abe Fábio Seigi Murakami Gustavo Frensch Francisco A. Marques Tomoe Nakashima http://www.mdpi.com/1424-8247/4/12/1518 Respiratory viral infections constitute the most frequent reason for medical consultations in the World. They can be associated with a wide range of clinical manifestations ranging from self-limited upper respiratory tract infections to more devastating conditions such as pneumonia. In particular, in serious cases influenza A leads to pneumonia, which is particularly fatal in patients with cardiopulmonary diseases, obesity, young children and the elderly. In the present study, we show a protective effect of the low-molecular weight compound Ingavirin (6-[2-(1H-imidazol-4-yl)ethylamino]-5-oxohexanoic acid) against influenza A (H1N1) virus, human parainfluenza virus and human adenovirus infections in animals. Mortality, weight loss, infectious titer of the virus in tissues and tissue morphology were monitored in the experimental groups of animals. The protective action of Ingavirin was observed as a reduction of infectious titer of the virus in the lung tissue, prolongation of the life of the infected animals, normalization of weight dynamics throughout the course of the disease, lowering of mortality of treated animals compared to a placebo control and normalization of tissue structure. In case of influenza virus infection, the protective activity of Ingavirin was similar to that of the reference compound Tamiflu. Based on the results obtained, Ingavirin should be considered as an important part of anti-viral prophylaxis and therapy. http://www.mdpi.com/1424-8247/4/12/1518 Fri, 25 Nov 2011 00:00:00 CET Pharmaceuticals 2011-11-25 4 12 Article 1518 1534 1424-8247 Activity of Ingavirin (6-[2-(1H-Imidazol-4-yl)ethylamino]-5-oxo-hexanoic Acid) Against Human Respiratory Viruses in in Vivo Experiments 2011-11-25 doi: 10.3390/ph4121518 Vladimir V. Zarubaev Angelica V. Garshinina Nelly A. Kalinina Anna A. Shtro Svetlana V. Belyaevskaya Alexander V. Slita Vladimir E. Nebolsin Oleg I. Kiselev http://www.mdpi.com/1424-8247/4/11/1503 Serine/threonine protein kinase C βII isoform (PKCβII) or the pain receptor transient receptor potential vanilloid 1 (TRPV1) have been separately implicated in mediating heat hyperalgesia during inflammation or diabetic neuropathy. However, detailed information on the role of PKC βII in nociceptive signaling mediated by TRPV1 is lacking. This study presents evidence for activation and translocation of the PKC βII isoform as a signaling event in nociception mediated by activation of TRPV1 by capsaicin. We show that capsaicin induces translocation of cytosolic PKCβII isoform fused with enhanced green fluorescence protein (PKCβII-EGFP) in dorsal root ganglion (DRG) neurons. We also show capsaicin-induced translocation in Chinese Hamster Ovarian (CHO) cells co-transfected with TRPV1 and PKCβII-EGFP, but not in CHO cells expressing PKCβII-EGFP alone. By contrast, the PKC activator phorbol-12-myristate-13-acetate (PMA) induced translocation of PKCβII-EGFP which was sustained and independent of calcium or TRPV1. In addition PMA-induced sensitization of TRPV1 to capsaicin response in DRG neurons was attenuated by PKCβII blocker CGP 53353. Capsaicin response via TRPV1 in the DRG neurons was confirmed by TRPV1 antagonist AMG 9810. These results suggested a novel and potential signaling link between PKCβII and TRPV1. These cell culture models provide a platform for investigating mechanisms of painful neuropathies mediated by nociceptors expressing the pain sensing gene TRPV1, and its regulation by the PKC isoform PKCβII. http://www.mdpi.com/1424-8247/4/11/1503 Fri, 11 Nov 2011 00:00:00 CET Pharmaceuticals 2011-11-11 4 11 Article 1503 1517 1424-8247 Real-Time Translocation and Function of PKCβII Isoform in Response to Nociceptive Signaling via the TRPV1 Pain Receptor 2011-11-11 doi: 10.3390/ph4111503 Sravan Mandadi Patricia J. Armati Basil D. Roufogalis http://www.mdpi.com/1424-8247/4/11/1488 In this study we have focused on the response of SKBR-3 cells to both single 17-DMAG treatment as well as its combination with photodynamic therapy with hypericin. Low concentrations of 17-DMAG without any effect on survival of SKBR-3 cells significantly reduced metabolic activity, viability and cell number when combined with photodynamic therapy with hypericin. Moreover, IC10 concentation of 17-DMAG resulted in significant increase of SKBR-3 cells in G1 phase of the cell cycle, followed by an increase of cells in G2 phase when combined with photodynamic therapy. Furthermore, 17-DMAG already decreased HER2, Akt, P-Erk1/2 and survivin protein levels in SKBR-3 cells a short time after its application. In this regard, 17-DMAG protected also SKBR-3 cells against both P-Erk1/2 as well as survivin upregulations induced by photodynamic therapy with hypericin. Interestingly, IC10 concentration of 17-DMAG led to total depletion of Akt, P-Erk1/2 proteins and to decrease of survivin level at 48 h. On the other hand, 17-DMAG did not change HER2 relative expression in SKBR-3 cells, but caused a significant decrease of HER2 mRNA in MCF-7 cells characterized by low HER2 expression. These results show that targeting HSP90 client proteins increases the efficiency of antineoplastic effect of photodynamic therapy in vitro. http://www.mdpi.com/1424-8247/4/11/1488 Thu, 10 Nov 2011 00:00:00 CET Pharmaceuticals 2011-11-10 4 11 Article 1488 1502 1424-8247 Photodynamic Therapy with Hypericin Improved by Targeting HSP90 Associated Proteins 2011-11-10 doi: 10.3390/ph4111488 Peter Solár Mária Chytilová Zuzana Solárová Ján Mojžiš Peter Ferenc Peter Fedoročko http://www.mdpi.com/1424-8247/4/11/1475 This is a review of key factors for pharmacy and therapeutics committees to consider when developing a therapeutic interchange (TI) program for venous thromboembolism (VTE) prophylaxis. Recent patient safety initiatives aimed at reducing the incidence of hospital-acquired VTE may increase the prescribing of thromboprophylactic agents recommended in VTE management guidelines. As a result, more pharmacy and therapeutics committees may consider TI programs for parenteral anticoagulants. However, the TI of anticoagulants appears challenging at this time. Firstly, the therapeutic equivalence of the commonly prescribed parenteral anticoagulants, enoxaparin, dalteparin and fondaparinux, has not been established. Secondly, because of the wide range of clinical indications for these anticoagulants, a blanket agent-specific TI program could lead to off-label use. Use of an indication-specific TI program could be difficult to manage administratively, and may cause prescribing confusion and errors. Thirdly, careful dosing and contraindications of certain parenteral anticoagulants in special patient populations, such as those with renal impairment, further impact the suitability of these agents for inclusion in TI programs. Finally, although TI may appear to offer lower drug-acquisition costs, it is important to determine its effect on all cost parameters and ultimately ensure that the care of patients requiring VTE prophylaxis is not compromised. http://www.mdpi.com/1424-8247/4/11/1475 Mon, 07 Nov 2011 00:00:00 CET Pharmaceuticals 2011-11-07 4 11 Review 1475 1487 1424-8247 Therapeutic Interchange of Parenteral Anticoagulants: Challenges for Pharmacy and Therapeutics Committees 2011-11-07 doi: 10.3390/ph4111475 Alpesh Amin http://www.mdpi.com/1424-8247/4/11/1450 The design of new drugs with better physiochemical properties, adequate absorption, distribution, metabolism, and excretion, effective pharmacologic potency and lacking toxicity remains is a challenge. Inflammation is the initial trigger of several different diseases, such as Alzheimer’s disease, asthma, atherosclerosis, colitis, rheumatoid arthritis, depression, cancer; and disorders such as obesity and sexual dysfunction. Although inflammation is not the direct cause of these disorders, inflammatory processes often increase related pain and suffering. New anti-inflammatory drugs developed using molecular hybridization techniques to obtain multiple-ligand drugs can act at one or multiple targets, allowing for synergic action and minimizing toxicity. This work is a review of new anti-inflammatory drugs developed using the molecular modification approach. http://www.mdpi.com/1424-8247/4/11/1450 Thu, 27 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-27 4 11 Review 1450 1474 1424-8247 Anti-Inflammatory Drug Design Using a Molecular Hybridization Approach 2011-10-27 doi: 10.3390/ph4111450 Priscila Longhin Bosquesi Thais Regina Ferreira Melo Ednir Oliveira Vizioli Jean Leandro dos Santos Man Chin Chung http://www.mdpi.com/1424-8247/4/11/1434 RNA-based approaches are among the most promising strategies aimed at developing safer and more effective therapeutics. RNA therapeutics include small non-coding miRNAs, small interfering RNA, RNA aptamers and more recently, small activating RNAs. However, major barriers exist to the use of RNAs as therapeutics such as resistance to nucleases present in biological fluids, poor chemical stability, need of specific cell targeted delivery and easy entry into the cell. Such issues have been addressed by several recent reports that show the possibility of introducing chemical modifications in small RNAs to stabilize the molecular conformation and increase by several fold their integrity, while still preserving the functional activity. Further, several aptamers have been developed as excellent candidates for the specific recognition of cell surface targets. In the last few years, by taking advantage of recent advances in the small RNA field, molecular bioconjugates have been designed that permit specific targeting and may act as cargoes for cell internalization of small RNAs acting on gene expression that will be discussed in this review. http://www.mdpi.com/1424-8247/4/11/1434 Thu, 27 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-27 4 11 Review 1434 1449 1424-8247 Coupling Aptamers to Short Interfering RNAs as Therapeutics 2011-10-27 doi: 10.3390/ph4111434 Laura Cerchia Carla Lucia Esposito Simona Camorani Silvia Catuogno Vittorio de Franciscis http://www.mdpi.com/1424-8247/4/11/1423 Alchornea triplinervia (Spreng.) Muell. Arg (Euphorbiaceae) is a medicinal plant commonly used by people living in the Cerrado region of Brazil to treat gastrointestinal ulcers. We previously described the gastroprotective action of methanolic extract (ME) of Alchornea triplinervia and the ethyl acetate fraction (EAF) in increasing of prostaglandin E2 (PGE2) gastric levels in the mucosa. In this work we evaluated the effect of EAF in promoting the healing process in rats with acetic acid-induced gastric ulcers. In addition, toxicity was investigated during treatment with EAF. After 14 days of treatment with EAF, the potent stimulator of gastric cell proliferation contributed to the acceleration of gastric ulcer healing. Upon immunohistochemical analysis, we observed a pronounced expression of COX-2, mainly in the submucosal layer. The 14-day EAF treatment also significantly increased the number of neutrophils in the gastric mucosa regeneration area. The EAF induced angiogenesis on gastric mucosa, observed as an increase of the number of blood vessels supplying the stomach in rats treated with EAF. Oral administration for 14 days of the ethyl acetate fraction from Alchornea triplinervia accelerated the healing of gastric ulcers in rats by promoting epithelial cell proliferation, increasing the number of neutrophils and stimulation of mucus production. This fraction, which contained mainly phenolic compounds, contributed to gastric mucosa healing. http://www.mdpi.com/1424-8247/4/11/1423 Tue, 25 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-25 4 11 Article 1423 1433 1424-8247 Effects of the Ethyl Acetate Fraction of Alchornea triplinervia on Healing Gastric Ulcer in Rats 2011-10-25 doi: 10.3390/ph4111423 Zeila P. Lima Flavia Bonamin Tamara R. Calvo Wagner Vilegas Lourdes C. Santos Ariane L. Rozza Claudia H. Pellizzon Lucia R. M. Rocha Clélia A. Hiruma-Lima http://www.mdpi.com/1424-8247/4/11/1400 The molecular chaperone Hsp90 holds great promise as a cancer drug target, despite some of the initial clinical trials of Hsp90 inhibitor drugs having not lived up to expectation. Effective use of these drugs will benefit greatly from a much more detailed understanding of the factors that contribute to resistance, whether intrinsic or acquired. We review how cell culture studies have revealed a number of different mechanisms whereby cells can be rendered less susceptible to the effects of Hsp90 inhibitor treatment. A major influence is Hsp90 inhibition causing strong induction of the heat shock response, a stress response that increases cellular levels of prosurvival chaperones such as Hsp27 and Hsp70. Another problem seems to be that these inhibitors do not always access the Hsp90 proteins of the mitochondrion, forms of Hsp90 that—in cancer cells—are operating to suppress apoptosis. It should be possible to overcome these drawbacks through the appropriate drug redesign or with the combinatorial use of an Hsp90 inhibitor with a drug that targets either heat shock factor or the chaperone Hsp70. Still though, cells will often differ in the key antiapoptotic versus proapoptotic activities that are dependent on Hsp90, in the key steps in their apoptotic pathways responsive to Hsp90 inhibition or Hsp70 level, as well as the extents to which their survival is dependent on oncogenic tyrosine kinases that are clients of Hsp90. A systems approach will therefore often be required in order to establish the most prominent effects of Hsp90 inhibition in each type of cancer cell. http://www.mdpi.com/1424-8247/4/11/1400 Tue, 25 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-25 4 11 Review 1400 1422 1424-8247 Mechanisms of Resistance to Hsp90 Inhibitor Drugs: A Complex Mosaic Emerges 2011-10-25 doi: 10.3390/ph4111400 Peter W. Piper Stefan H. Millson http://www.mdpi.com/1424-8247/4/10/1381 Non-viral gene therapy requires innovative strategies to achieve higher transfection efficacy. A few years ago, our group proposed bioinspired lipids whoseinteraction with DNA was not based on ionic interactions, but on hydrogen bonds. We thusdeveloped lipids bearing a thiourea head which allowed an interaction with DNAphosphates through hydrogen bonds. After a proof of concept with a lipid bearing threethiourea functions, a molecular and cellular screening was performed by varying all partsof the lipids: the hydrophobic anchor, the spacer, the linker, and the thiourea head. Twolipothiourea-based structures were identified as highly efficient in vitro transfecting agents.The lipothioureas were shown to reduce non specific interactions with cell membranes anddeliver their DNA content intracellularly more efficiently, as compared to cationiclipoplexes. These lipids could deliver siRNA efficiently and allowed specific cell targetingin vitro. In vivo, thiourea lipoplexes presented a longer retention time in the blood and lessaccumulation in the lungs after an intravenous injection in mice. They also inducedluciferase gene expression in muscle and tumor after local administration in mice.Therefore, these novel lipoplexes represent an excellent alternative to cationic lipoplexes astransfecting agents. In this review we will focus on the structure activity studies thatpermitted the identification of the two most efficient thiourea lipids. http://www.mdpi.com/1424-8247/4/10/1381 Mon, 24 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-24 4 10 Review 1381 1399 1424-8247 Lipothioureas as Lipids for Gene Transfection: A Review 2011-10-24 doi: 10.3390/ph4101381 Marie Breton Jeanne Leblond Isabelle Tranchant Daniel Scherman Michel Bessodes Jean Herscovici Nathalie Mignet http://www.mdpi.com/1424-8247/4/10/1355 Stem cell therapy has emerged as a promising new approach in almost every medicine specialty. This vast, heterogeneous family of cells are now both naturally (embryonic and adult stem cells) or artificially obtained (induced pluripotent stem cells or iPSCs) and their fates have become increasingly controllable, thanks to ongoing research in this passionate new field. We are at the beginning of a new era in medicine, with multiple applications for stem cell therapy, not only as a monotherapy, but also as an adjunct to other strategies, such as organ transplantation or standard drug treatment. Regrettably, serious preclinical concerns remain and differentiation, cell fusion, senescence and signalling crosstalk with growth factors and biomaterials are still challenges for this promising multidisciplinary therapeutic modality. Severe burns have several indications for stem cell therapy, including enhancement of wound healing, replacement of damaged skin and perfect skin regeneration – incorporating skin appendages and reduced fibrosis –, as well as systemic effects, such as inflammation, hypermetabolism and immunosuppression. The aim of this review is to describe well established characteristics of stem cells and to delineate new advances in the stem cell field, in the context of burn injury and wound healing. http://www.mdpi.com/1424-8247/4/10/1355 Fri, 21 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-21 4 10 Review 1355 1380 1424-8247 Stem Cell Therapy: A New Treatment for Burns? 2011-10-21 doi: 10.3390/ph4101355 Anna Arno Alexandra H. Smith Patrick H. Blit Mohammed Al Shehab Gerd G. Gauglitz Marc G. Jeschke http://www.mdpi.com/1424-8247/4/10/1328 Ribonucleotide reductase (RR) is a crucial enzyme in de novo DNA synthesis, where it catalyses the rate determining step of dNTP synthesis. RRs consist of a large subunit called RR1 (α), that contains two allosteric sites and one catalytic site, and a small subunit called RR2 (β), which houses a tyrosyl free radical essential for initiating catalysis. The active form of mammalian RR is an anbm hetero oligomer. RR inhibitors are cytotoxic to proliferating cancer cells. In this brief review we will discuss the three classes of RR, the catalytic mechanism of RR, the regulation of the dNTP pool, the substrate selection, the allosteric activation, inactivation by ATP and dATP, and the nucleoside drugs that target RR. We will also discuss possible strategies for developing a new class of drugs that disrupts the RR assembly. http://www.mdpi.com/1424-8247/4/10/1328 Thu, 13 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-13 4 10 Review 1328 1354 1424-8247 Targeting the Large Subunit of Human Ribonucleotide Reductase for Cancer Chemotherapy 2011-10-13 doi: 10.3390/ph4101328 Sanath R. Wijerathna Md. Faiz Ahmad Hai Xu James W. Fairman Andrew Zhang Prem Singh Kaushal Qun Wan Jianying Kiser Chris G. Dealwis http://www.mdpi.com/1424-8247/4/10/1315 The high rate of recidivism in cocaine addiction after prolonged periods of abstinence poses a significant problem for the effective treatment of this condition. Moreover, the neurobiological basis of this relapse phenomenon remains poorly understood. In this review, we will discuss the evidence currently available regarding the neurobiological changes during the extinction of cocaine self-administration. Specifically, we will focus on alterations in the dopaminergic, opioidergic, glutamatergic, cholinergic, serotoninergic and CRF systems described in self-administration experiments and extinction studies after chronic cocaine administration. We will also discuss the differences related to contingent versus non-contingent cocaine administration, which highlights the importance of environmental cues on drug effects and extinction. The findings discussed in this review may aid the development of more effective therapeutic approaches to treat cocaine relapse. http://www.mdpi.com/1424-8247/4/10/1315 Thu, 13 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-13 4 10 Review 1315 1327 1424-8247 Neural Changes Developed during the Extinction of Cocaine Self-Administration Behavior 2011-10-13 doi: 10.3390/ph4101315 Alejandro Higuera-Matas Miguel Miguens Nuria del Olmo Carmen García-Lecumberri Emilio Ambrosio http://www.mdpi.com/1424-8247/4/10/1295 Pelargonium species contribute significantly to the health care of a large population in the Southern African region, as part of a long-standing medical system intimately linked to traditional healing practices. Most notably, extracts of the roots of P. sidoides have commonly been applied for the treatment of dysentery and diarrhoea but only occasionally for respiratory complaints. Clinical trials have shown that a modern aqueous-ethanolic formulation of P. sidoides extracts (EPs® 7630) is an efficacious treatment for disorders of the respiratory tract, for example bronchitis and sinusitis. It should be noted that EPs® 7630 is the most widely investigated extract and therefore is the focus of this review. In order to provide a rationale for its therapeutic activity extracts have been evaluated for antibacterial activity and for their effects on non-specific immune functions. Only moderate direct antibacterial capabilities against a spectrum of bacteria, including Mycobacteria strains, have been noted. In contrast, a large body of in vitro studies has provided convincing evidence for an anti-infective principle associated with activation of the non-specific immune system. Interestingly, significant inhibition of interaction between bacteria and host cells, a key to the pathogenesis of respiratory tract infections, has emerged from recent studies. In addition, antiviral effects have been demonstrated, including inhibition of the replication of respiratory viruses and the enzymes haemagglutinin and neuraminidase. Besides, an increase of cilliary beat frequency of respiratory cells may contribute to the beneficial effects of P. sidoides extracts. This example provides a compelling argument for continuing the exploration of Nature and traditional medical systems as a source of therapeutically useful herbal medicines. http://www.mdpi.com/1424-8247/4/10/1295 Mon, 10 Oct 2011 00:00:00 CEST Pharmaceuticals 2011-10-10 4 10 Review 1295 1314 1424-8247 Antimicrobial, Antiviral and Immunomodulatory Activity Studies of Pelargonium sidoides (EPs® 7630) in the Context of Health Promotion 2011-10-10 doi: 10.3390/ph4101295 Herbert Kolodziej http://www.mdpi.com/1424-8247/4/10/1293 In the published version “Paladini et al. reported that aliskiren 300 mg provided a sustained BP-lowering effect beyond the 24-h dosing interval, with a significantly smaller loss of BP-lowering effect in the 24-48 h period after dose than irbesartan 300 mg or ramipril 10 mg [25]”. Paladini et al. should be Palatini et al., and the cited reference number should be [10], not [25].. In the sentence, “Early data suggest a role for aliskiren in preventing end-organ damage but, considering the ONTARGET results with an ACE-I-ARB combination, outcome studies are needed before the use of aliskiren can be recommended in combination with other RAS inhibitors [18–30]” one more citation number was added [5], so the revised sentence is “Early data suggest a role for aliskiren in preventing end-organ damage but, considering the ONTARGET results with an ACE-I-ARB combination, outcome studies are needed before the use of aliskiren can be recommended in combination with other RAS inhibitors [5,18–30]”. [...] http://www.mdpi.com/1424-8247/4/10/1293 Fri, 30 Sep 2011 00:00:00 CEST Pharmaceuticals 2011-09-30 4 10 Correction 1293 1294 1424-8247 Novo, S. et al. Aliskiren: Just a New Drug for Few Selected Patients or an Innovative Molecule Predestinated to Replace Arbs and Ace-Inhibitors? Pharmaceuticals 2009, 2, 118-124 2011-09-30 doi: 10.3390/ph4101293 Salvatore Novo Giovanni Fazio Elena Raccuglia Antonino Mignano Giuseppina Novo http://www.mdpi.com/1424-8247/4/10/1281 Cross-linked nanoassemblies (CNAs) with a degradable core were prepared for sustained release of 17-N-allylamino-17-demethoxygeldanamycin (17-AAG), a potent inhibitor of heat shock protein 90 (HSP90). The particle size of CNAs ranged between 100 and 250 nm, which changed depending on the cross-linking yields and drug entrapment method. CNAs with a 1% cross-linking yield entrapped 17-AAG in aqueous solutions, yet degraded in 3 hrs. CNAs entrapped 5.2 weight% of 17-AAG as the cross-linking yield increased to 10%, retaining more than 80% of particles for 24 hrs. CNAs with drugs entrapped after the cross-linking reactions were 100 nm and remained stable in both pH 7.4 and 5.0, corresponding to the physiological, tumoral, and intracellular environments. Drug was completely released from CNAs in 48 hrs, which would potentially maximize drug delivery and release efficiency within tumor tissues. Drug release patterns were not negatively affected by changing the cross-linking yields of CNAs. CNAs entrapping 17-AAG suppressed the growth of human non-small cell lung cancer A549 cells as equally effective as free drugs. The results demonstrated that CNAs would be a promising formulation that can be used in aqueous solutions for controlled delivery and release of 17-AAG. http://www.mdpi.com/1424-8247/4/10/1281 Mon, 26 Sep 2011 00:00:00 CEST Pharmaceuticals 2011-09-26 4 10 Article 1281 1292 1424-8247 Degradable Cross-Linked Nanoassemblies as Drug Carriers for Heat Shock Protein 90 Inhibitor 17-N-Allylamino-17-demethoxy-geldanamycin 2011-09-26 doi: 10.3390/ph4101281 Andrei Ponta Shanjida Akter Younsoo Bae http://www.mdpi.com/1424-8247/4/9/1248 Different neuropeptides, all containing a common carboxy-terminal RFamide sequence, have been characterized as ligands of the RFamide peptide receptor family. Currently, five subgroups have been characterized with respect to their N-terminal sequence and hence cover a wide pattern of biological functions, like important neuroendocrine, behavioral, sensory and automatic functions. The RFamide peptide receptor family represents a multiligand/multireceptor system, as many ligands are recognized by several GPCR subtypes within one family. Multireceptor systems are often susceptible to cross-reactions, as their numerous ligands are frequently closely related. In this review we focus on recent results in the field of structure-activity studies as well as mutational exploration of crucial positions within this GPCR system. The review summarizes the reported peptide analogs and recently developed small molecule ligands (agonists and antagonists) to highlight the current understanding of the pharmacophoric elements, required for affinity and activity at the receptor family. Furthermore, we address the biological functions of the ligands and give an overview on their involvement in physiological processes. We provide insights in the knowledge for the design of highly selective ligands for single receptor subtypes to minimize cross-talk and to eliminate effects from interactions within the GPCR system. This will support the drug development of members of the RFamide family. http://www.mdpi.com/1424-8247/4/9/1248 Wed, 21 Sep 2011 00:00:00 CEST Pharmaceuticals 2011-09-21 4 9 Review 1248 1280 1424-8247 RFamide Peptides: Structure, Function, Mechanisms and Pharmaceutical Potential 2011-09-21 doi: 10.3390/ph4091248 Maria Findeisen Daniel Rathmann Annette G. Beck-Sickinger http://www.mdpi.com/1424-8247/4/9/1236 We present a self-organizing map (SOM) approach to predicting macromolecular targets for combinatorial compound libraries. The aim was to study the usefulness of the SOM in combination with a topological pharmacophore representation (CATS) for selecting biologically active compounds from a virtual combinatorial compound collection, taking the multi-component Biginelli dihydropyrimidine reaction as an example. We synthesized a candidate compound from this library, for which the SOM model suggested inhibitory activity against cyclin-dependent kinase 2 (CDK2) and other kinases. The prediction was confirmed in an in vitro panel assay comprising 48 human kinases. We conclude that the computational technique may be used for ligand-based in silico pharmacology studies, off-target prediction, and drug re-purposing, thereby complementing receptor-based approaches. http://www.mdpi.com/1424-8247/4/9/1236 Tue, 20 Sep 2011 00:00:00 CEST Pharmaceuticals 2011-09-20 4 9 Article 1236 1247 1424-8247 Target Profile Prediction and Practical Evaluation of a Biginelli-Type Dihydropyrimidine Compound Library 2011-09-20 doi: 10.3390/ph4091236 Petra Schneider Katharina Stutz Ladina Kasper Sarah Haller Michael Reutlinger Felix Reisen Tim Geppert Gisbert Schneider http://www.mdpi.com/1424-8247/4/9/1216 Aptamers are nucleic acid-based ligands identified through a process of molecular evolution named SELEX (Systematic Evolution of Ligands by Exponential enrichment). During the last 10-15 years, numerous aptamers have been developed specifically against targets present on or associated with the surface of human cells or infectious pathogens such as viruses, bacteria, fungi or parasites. Several of the aptamers have been described as potent probes, rivalling antibodies, for use in flow cytometry or microscopy. Some have also been used as drugs by inhibiting or activating functions of their targets in a manner similar to neutralizing or agonistic antibodies. Additionally, it is straightforward to conjugate aptamers to other agents without losing their affinity and they have successfully been used in vitro and in vivo to deliver drugs, siRNA, nanoparticles or contrast agents to target cells. Hence, aptamers identified against cell surface biomarkers represent a promising class of ligands. This review presents the different strategies of SELEX that have been developed to identify aptamers for cell surface-associated proteins as well as some of the methods that are used to study their binding on living cells. http://www.mdpi.com/1424-8247/4/9/1216 Tue, 20 Sep 2011 00:00:00 CEST Pharmaceuticals 2011-09-20 4 9 Review 1216 1235 1424-8247 Methods To Identify Aptamers against Cell Surface Biomarkers 2011-09-20 doi: 10.3390/ph4091216 Agnes Cibiel Daniel Miotto Dupont Frédéric Ducongé http://www.mdpi.com/1424-8247/4/9/1196 Recently the first community-wide assessments of the prediction of the structures of complexes between proteins and small molecule ligands have been reported in the so-called GPCR Dock 2008 and 2010 assessments. In the current review we discuss the different steps along the protein-ligand modeling workflow by critically analyzing the modeling strategies we used to predict the structures of protein-ligand complexes we submitted to the recent GPCR Dock 2010 challenge. These representative test cases, focusing on the pharmaceutically relevant G Protein-Coupled Receptors, are used to demonstrate the strengths and challenges of the different modeling methods. Our analysis indicates that the proper performance of the sequence alignment, introduction of structural adjustments guided by experimental data, and the usage of experimental data to identify protein-ligand interactions are critical steps in the protein-ligand modeling protocol. http://www.mdpi.com/1424-8247/4/9/1196 Thu, 01 Sep 2011 00:00:00 CEST Pharmaceuticals 2011-09-01 4 9 Review 1196 1215 1424-8247 In Silico Veritas: The Pitfalls and Challenges of Predicting GPCR-Ligand Interactions 2011-09-01 doi: 10.3390/ph4091196 Luc Roumen Marijn P.A. Sanders Bas Vroling Iwan J.P. De Esch Jacob De Vlieg Rob Leurs Jan P.G. Klomp Sander B. Nabuurs Chris De Graaf http://www.mdpi.com/1424-8247/4/8/1183 Mithramycin A (MTM) and histone deacetylase inhibitors (HDACi) are effective therapeutic agents for cancer and neurodegenerative diseases. MTM is a FDA approved aureolic acid-type antibiotic that binds to GC-rich DNA sequences and interferes with Sp1 transcription factor binding to its target sites (GC box). HDACi, on the other hand, modulate the activity of class I and II histone deacetylases. They mediate their protective function, in part, by regulating the acetylation status of histones or transcription factors, including Sp1, and in turn chromatin accessibility to the transcriptional machinery. Because these two classes of structurally and functionally diverse compounds mediate similar therapeutic functions, we investigated whether they act on redundant or synergistic pathways to protect neurons from oxidative death. Non-protective doses of each of the drugs do not synergize to create resistance to oxidative death suggesting that these distinct agents act via a similar pathway. Accordingly, we found that protection by MTM and HDACi is associated with diminished expression of the oncogene, Myc and enhanced expression of a tumor suppressor, p21waf1/cip1. We also find that neuroprotection by MTM or Myc knockdown is associated with downregulation of class I HDAC levels. Our results support a model in which the established antitumor drug MTM or canonical HDACi act via distinct mechanisms to converge on the downregulation of HDAC levels or activity respectively. These findings support the conclusion that an imbalance in histone acetylase and HDAC activity in favor of HDACs is key not only for oncogenic transformation, but also neurodegeneration. http://www.mdpi.com/1424-8247/4/8/1183 Mon, 22 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-22 4 8 Article 1183 1195 1424-8247 Histone Deacetylase Inhibitors and Mithramycin A Impact a Similar Neuroprotective Pathway at a Crossroad between Cancer and Neurodegeneration 2011-08-22 doi: 10.3390/ph4081183 Sama F. Sleiman Jill Berlin Manuela Basso Saravanan S.Karuppagounder Jürgen Rohr Rajiv R. Ratan http://www.mdpi.com/1424-8247/4/8/1171 Frankincense (Boswellia serrata, B. carterii) is used as traditional remedy to treat inflammatory diseases. The molecular effects of the active ingredients, the boswellic acids, on the immune system have previously been studied and verified in several clinical studies. Boswellic acids also inhibit cancer cell growth in vitro and in vivo. The molecular basis of the cytotoxicity of boswellic acids is, however, not fully understood as yet. By mRNA-based microarray, COMPARE, and hierarchical cluster analyses, we identified a panel of genes from diverse functional groups, which were significantly associated with sensitivity or resistance of a- or b-boswellic acids, such as transcription factors, signal transducers, growth regulating genes, genes involved in RNA and protein metabolism and others. This indicates that boswellic acids exert profound cytotoxicity on cancer cells by a multiplicity of molecular mechanisms. http://www.mdpi.com/1424-8247/4/8/1171 Fri, 19 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-19 4 8 Article 1171 1182 1424-8247 Molecular Determinants of the Response of Tumor Cells to Boswellic Acids 2011-08-19 doi: 10.3390/ph4081171 Tolga Eichhorn Henry Johannes Greten Thomas Efferth http://www.mdpi.com/1424-8247/4/8/1158 Preparation of 4-benzyl-2-substituted phthalazin-1-one derivatives 2-8 is reported. Condensation of 4-benzyl-1-chlorophthalazine (9) with a series of different nucleophiles gave 4-benzylphthalazin-1-ylamino derivatives (10-13 and 16) and 4-amino-2-[N'-(4-benzylphthalazin-1-yl)-hydrazino]-6-arylpyrimidine-5-carbonitriles (14a,b). Interaction of 9 with ambident anions was also studied. 5-Benzyl-6,6a,12-triazobenzo[a]-anthracen-7-one (15) is obtained from 9 and anthranilic acid derivatives. Treatment of 16 with (EtO)3CH/Ac2O under reflux afforded the corresponding ethoxymethylene derivative 17, while aqueous ammonium hydroxide treatment afforded carboxamide derivative 18. The structures of the newly synthesized derivatives were confirmed by their elemental analysis, IR, 1H NMR, 13C NMR and mass spectral studies. Antimicrobial activities of some selected compounds were also studied and some of these were found to exhibit promising effects against Gram-positive and Gram-negative bacteria and fungi. http://www.mdpi.com/1424-8247/4/8/1158 Wed, 17 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-17 4 8 Article 1158 1170 1424-8247 Synthesis and Biological Screening of 4-Benzyl-2H-phthalazine Derivatives 2011-08-17 doi: 10.3390/ph4081158 Ashraf H.F. Abd El-Wahab Hany M. Mohamed Ahmed M. El-Agrody Mohammed A. El-Nassag Ahmed H. Bedair http://www.mdpi.com/1424-8247/4/8/1137 Aptamers are functional nucleic acid sequences which can bind specific targets. An artificial combinatorial methodology can identify aptamer sequences for any target molecule, from ions to whole cells. Drug delivery systems seek to increase efficacy and reduce side-effects by concentrating the therapeutic agents at specific disease sites in the body. This is generally achieved by specific targeting of inactivated drug molecules. Aptamers which can bind to various cancer cell types selectively and with high affinity have been exploited in a variety of drug delivery systems for therapeutic purposes. Recent progress in selection of cell-specific aptamers has provided new opportunities in targeted drug delivery. Especially functionalization of nanoparticles with such aptamers has drawn major attention in the biosensor and biomedical areas. Moreover, nucleic acids are recognized as an attractive building materials in nanomachines because of their unique molecular recognition properties and structural features. A active controlled delivery of drugs once targeted to a disease site is a major research challenge. Stimuli-responsive gating is one way of achieving controlled release of nanoparticle cargoes. Recent reports incorporate the structural properties of aptamers in controlled release systems of drug delivering nanoparticles. In this review, the strategies for using functional nucleic acids in creating smart drug delivery devices will be explained. The main focus will be on aptamer-incorporated nanoparticle systems for drug delivery purposes in order to assess the future potential of aptamers in the therapeutic area. Special emphasis will be given to the very recent progress in controlled drug release based on molecular gating achieved with aptamers. http://www.mdpi.com/1424-8247/4/8/1137 Mon, 15 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-15 4 8 Review 1137 1157 1424-8247 Aptamer-Gated Nanoparticles for Smart Drug Delivery 2011-08-15 doi: 10.3390/ph4081137 Veli Cengiz Ozalp Fusun Eyidogan Huseyin Avni Oktem http://www.mdpi.com/1424-8247/4/8/1101 The endocannabinoid system (ECS) has been implicated in many physiological functions, including the regulation of appetite, food intake and energy balance, a crucial involvement in brain reward systems and a role in psychophysiological homeostasis (anxiety and stress responses). We first introduce this important regulatory system and chronicle what is known concerning the signal transduction pathways activated upon the binding of endogenous cannabinoid ligands to the Gi/0-coupled CB1 cannabinoid receptor, as well as its interactions with other hormones and neuromodulators which can modify endocannabinoid signaling in the brain. Anorexia nervosa (AN) and bulimia nervosa (BN) are severe and disabling psychiatric disorders, characterized by profound eating and weight alterations and body image disturbances. Since endocannabinoids modulate eating behavior, it is plausible that endocannabinoid genes may contribute to the biological vulnerability to these diseases. We present and discuss data suggesting an impaired endocannabinoid signaling in these eating disorders, including association of endocannabinoid components gene polymorphisms and altered CB1-receptor expression in AN and BN. Then we discuss recent findings that may provide new avenues for the identification of therapeutic strategies based on the endocannabinod system. In relation with its implications as a reward-related system, the endocannabinoid system is not only a target for cannabis but it also shows interactions with other drugs of abuse. On the other hand, there may be also a possibility to point to the ECS as a potential target for treatment of drug-abuse and addiction. Within this framework we will focus on enzymatic machinery involved in endocannabinoid inactivation (notably fatty acid amide hydrolase or FAAH) as a particularly interesting potential target. Since a deregulated endocannabinoid system may be also related to depression, anxiety and pain symptomatology accompanying drug-withdrawal states, this is an area of relevance to also explore adjuvant treatments for improving these adverse emotional reactions. http://www.mdpi.com/1424-8247/4/8/1101 Wed, 10 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-10 4 8 Review 1101 1136 1424-8247 The Endocannabinoid System as Pharmacological Target Derived from Its CNS Role in Energy Homeostasis and Reward. Applications in Eating Disorders and Addiction 2011-08-10 doi: 10.3390/ph4081101 Maria-Paz Viveros Francisco-Javier Bermúdez-Silva Ana-Belén Lopez-Rodriguez Edward J. Wagner http://www.mdpi.com/1424-8247/4/8/1088 Oxidative stress has been implicated in the pathogenesis of heart failure. Reactive oxygen species (ROS) are produced in the failing myocardium, and ROS cause hypertrophy, apoptosis/cell death and intracellular Ca2+ overload in cardiac myocytes. ROS also cause damage to lipid cell membranes in the process of lipid peroxidation. In this process, several aldehydes, including 4-hydroxy-2-nonenal (HNE), are generated and the amount of HNE is increased in the human failing myocardium. HNE exacerbates the formation of ROS, especially H2O2 and ·OH, in cardiomyocytes and subsequently ROS cause intracellular Ca2+ overload. Treatment with beta-blockers such as metoprolol, carvedilol and bisoprolol reduces the levels of oxidative stress, together with amelioration of heart failure. This reduction could be caused by several possible mechanisms. First, the beta-blocking effect is important, because catecholamines such as isoproterenol and norepinephrine induce oxidative stress in the myocardium. Second, anti-ischemic effects and negative chronotropic effects are also important. Furthermore, direct antioxidative effects of carvedilol contribute to the reduction of oxidative stress. Carvedilol inhibited HNE-induced intracellular Ca2+ overload. Beta-blocker therapy is a useful antioxidative therapy in patients with heart failure. http://www.mdpi.com/1424-8247/4/8/1088 Fri, 05 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-05 4 8 Review 1088 1100 1424-8247 Beta-Blockers and Oxidative Stress in Patients with Heart Failure 2011-08-05 doi: 10.3390/ph4081088 Kazufumi Nakamura Masato Murakami Daiji Miura Kei Yunoki Kenki Enko Masamichi Tanaka Yukihiro Saito Nobuhiro Nishii Toru Miyoshi Masashi Yoshida Hiroki Oe Norihisa Toh Satoshi Nagase Kunihisa Kohno Hiroshi Morita Hiromi Matsubara Kengo F Kusano Tohru Ohe Hiroshi Ito http://www.mdpi.com/1424-8247/4/8/1070 Many pathological conditions are associated with phosphatidylinositol 3-kinase (PI3K) dysfunction, providing an incentive for the study of the effects of PI3K modulation in different aspects of diabetes, cancer, and aging. The PI3K/AKT/mTOR pathway is a key transducer of brain metabolic and mitogenic signals involved in neuronal proliferation, differentiation, and survival. In several models of neurodegenerative diseases associated with aging, the PI3K/AKT pathway has been found to be dysregulated, suggesting that two or more initiating events may trigger disease formation in an age-related manner. The search for chemical compounds able to modulate the activity of the PI3K/AKT/mTOR pathway is emerging as a potential therapeutic strategy for the treatment and/or prevention of some metabolic defects associated with brain aging. In the current review, we summarize some of the critical actions of PI3K in brain function as well as the evidence of its involvement in aging and Alzheimer’s disease. http://www.mdpi.com/1424-8247/4/8/1070 Thu, 04 Aug 2011 00:00:00 CEST Pharmaceuticals 2011-08-04 4 8 Review 1070 1087 1424-8247 The Phosphatidylinositol 3-Kinase/mTor Pathway as a Therapeutic Target for Brain Aging and Neurodegeneration 2011-08-04 doi: 10.3390/ph4081070 David Heras-Sandoval Evangelina Avila-Muñoz Clorinda Arias http://www.mdpi.com/1424-8247/4/7/1052 Vasoinhibins, a family of antiangiogenic peptides derived from prolactin proteolysis, inhibit the vascular effects of several proangiogenic factors, including bradykinin (BK). Here, we report that vasoinhibins block the BK-induced proliferation of bovine umbilical vein endothelial cells. This effect is mediated by the inactivation of endothelial nitric oxide synthase (eNOS), as the NO donor DETA-NONOate reverted vasoinhibin action. It is an experimentally proven fact that the elevation of intracellular Ca2+ levels ([Ca2+]i) upon BK stimulation activates eNOS, and vasoinhibins blocked the BK-mediated activation of phospholipase C and the formation of inositol 1,4,5-triphosphate leading to a reduced release of Ca2+ from intracellular stores. The [Ca2+]i rise evoked by BK also involves the influx of extracellular Ca2+ via canonical transient receptor potential (TRPC) channels. Vasoinhibins likely interfere with TRPC-mediated Ca2+ entry since La3+, which is an enhancer of TRPC4 and TRPC5 channel activity, prevented vasoinhibins from blocking the stimulation by BK of endothelial cell NO production and proliferation, and vasoinhibins reduced the BK-induced increase of TRPC5 mRNA expression. Finally, vasoinhibins prevented the BK-induced phosphorylation of eNOS at Ser1179, a post-translational modification that facilitates Ca2+-calmodulin activation of eNOS. Together, our data show that vasoinhibins, by lowering NO production through the inhibition of both [Ca2+]i mobilization and eNOS phosphorylation, prevent the BK-induced stimulation of endothelial cell proliferation. Thus, vasoinhibins help to regulate BK effects on angiogenesis and vascular homeostasis. http://www.mdpi.com/1424-8247/4/7/1052 Wed, 20 Jul 2011 00:00:00 CEST Pharmaceuticals 2011-07-20 4 7 Article 1052 1069 1424-8247 Vasoinhibins Prevent Bradykinin-Stimulated Endothelial Cell Proliferation by Inactivating eNOS via Reduction of both Intracellular Ca2+ Levels and eNOS Phosphorylation at Ser1179 2011-07-20 doi: 10.3390/ph4071052 Stéphanie Thebault Carmen González Celina García David Arredondo Zamarripa Gabriel Nava Luis Vaca Fernando López-Casillas Gonzalo Martínez De la Escalera Carmen Clapp http://www.mdpi.com/1424-8247/4/7/1032 The behavior of 2-ethoxy-(4H)-3,1-benzoxazin-4-one (1) towards nitrogen nucleo-philes, e.g. ethanolamine, aromatic amines (namely: p-toluidine, p-anisidine, p-hydroxyaniline, o-hydroxyaniline, o-bromoaniline, o-phenylenediamine, p-phenylene- diamine, o-tolidinediamine) p-aminobenzoic acid, glucosamine hydrochloride,  2-amino- nicotinic acid, 1-naphthalenesulfonic acid hydrazide, n-decanoic acid hydrazide, benzoic acid hydrazide, semicarbazide, aminoacids (e.g. D,L-alanine, L-asparagine, L-arginine) and derivatives of 2-aminothiodiazole has been investigated. The behavior of the benzoxazinone towards a selected sulfur nucleophile, L-cysteine, has also been discussed. Formation of an amidine salt as a reaction intermediate has been assumed. The effect of solvent in some reactions has been elucidated. The structures of all the novel quinazoline and quinazolinone derivatives, obtained by heterocyclic ring opening and ring closure were inferred by the IR, MS as well as 1H-NMR spectral analysis. Moreover, the antimicrobial potential of some of the new synthesized derivatives has been evaluated. http://www.mdpi.com/1424-8247/4/7/1032 Thu, 14 Jul 2011 00:00:00 CEST Pharmaceuticals 2011-07-14 4 7 Article 1032 1051 1424-8247 The Uses of 2-Ethoxy-(4H)-3,1-benzoxazin-4-one in the Synthesis of Some Quinazolinone Derivatives of Antimicrobial Activity 2011-07-14 doi: 10.3390/ph4071032 Maher A. El-Hashash Khalid M. Darwish Sameh A. Rizk Fakhry A. El-Bassiouny http://www.mdpi.com/1424-8247/4/7/1019 Extracts of Echinacea species have been used traditionally in North America for the control of symptoms of colds, influenza, and other diseases, and some of them have become very popular as “herbal medicines”. Recent studies have revealed that preparations derived from certain species and plant parts, but not all of them, possess potent antiviral activities, at non-cytotoxic concentrations, particularly against membrane-containing viruses. Thus all strains of human and avian influenza viruses tested (including a Tamiflu-resistant strain), as well as herpes simplex virus, respiratory syncytial virus, and rhinoviruses, were very sensitive to a standardized Echinacea purpurea preparation. In mechanistic studies the influenza virus-specific hemagglutinin and neuraminidase were inhibited. In addition some extracts displayed anti-inflammatory activity in virus-infected cells, and numerous other effects on the expression of cellular genes. Multiple components, either discrete compounds or mixtures, appeared to be responsible for the various antiviral activities. http://www.mdpi.com/1424-8247/4/7/1019 Wed, 13 Jul 2011 00:00:00 CEST Pharmaceuticals 2011-07-13 4 7 Review 1019 1031 1424-8247 Echinacea—A Source of Potent Antivirals for Respiratory Virus Infections 2011-07-13 doi: 10.3390/ph4071019 James Hudson Selvarani Vimalanathan http://www.mdpi.com/1424-8247/4/7/992 The amphetamine analogue 3,4-methylenedioxymethamphetamine (MDMA, ‘ecstasy’) is widely abused as a recreational drug due to its unique psychological effects. Of interest, MDMA causes long-lasting deficits in neurochemical and histological markers of the serotonergic neurons in the brain of different animal species. Such deficits include the decline in the activity of tryptophan hydroxylase in parallel with the loss of 5-HT and its main metabolite 5-hydoxyindoleacetic acid (5-HIAA) along with a lower binding of specific ligands to the 5-HT transporters (SERT). Of concern, reduced 5-HIAA levels in the CSF and SERT density have also been reported in human ecstasy users, what has been interpreted to reflect the loss of serotonergic fibers and terminals. The neurotoxic potential of MDMA has been questioned in recent years based on studies that failed to show the loss of the SERT protein by western blot or the lack of reactive astrogliosis after MDMA exposure. In addition, MDMA produces a long-lasting down-regulation of SERT gene expression; which, on the whole, has been used to invoke neuromodulatory mechanisms as an explanation to MDMA-induced 5-HT deficits. While decreased protein levels do not necessarily reflect neurodegeneration, the opposite is also true, that is, neuroregulatory mechanisms do not preclude the existence of 5-HT terminal degeneration. http://www.mdpi.com/1424-8247/4/7/992 Tue, 05 Jul 2011 00:00:00 CEST Pharmaceuticals 2011-07-05 4 7 Review 992 1018 1424-8247 Methylenedioxymethamphetamine (MDMA, 'Ecstasy'): Neurodegeneration versus Neuromodulation 2011-07-05 doi: 10.3390/ph4070992 Elena Puerta Norberto Aguirre http://www.mdpi.com/1424-8247/4/7/976 Drugs of abuse induce plastic changes in the brain that seem to underlie addictive phenomena. These plastic changes can be structural (morphological) or synaptic (biochemical), and most of them take place in the mesolimbic and mesostriatal circuits. Several addiction-related changes in brain circuits (hypofrontality, sensitization, tolerance) as well as the outcome of treatment have been visualized in addicts to psychostimulants using neuroimaging techniques. Repeated exposure to psychostimulants induces morphological changes such as increase in the number of dendritic spines, changes in the morphology of dendritic spines, and altered cellular coupling through new gap junctions. Repeated exposure to psychostimulants also induces various synaptic adaptations, many of them related to sensitization and neuroplastic processes, that include up- or down-regulation of D1, D2 and D3 dopamine receptors, changes in subunits of G proteins, increased adenylyl cyclase activity, cyclic AMP and protein kinase A in the nucleus accumbens, increased tyrosine hydroxylase enzyme activity, increased calmodulin and activated CaMKII in the ventral tegmental area, and increased deltaFosB, c-Fos and AP-1 binding proteins. Most of these changes are transient, suggesting that more lasting plastic brain adaptations should take place. In this context, protein synthesis inhibitors block the development of sensitization to cocaine, indicating that rearrangement of neural networks must develop for the long-lasting plasticity required for addiction to occur. Self-administration studies indicate the importance of glutamate neurotransmission in neuroplastic changes underlying transition from use to abuse. Finally, plastic changes in the addicted brain are enhanced and aggravated by neuroinflammation and neurotrophic disbalance after repeated psychostimulants. http://www.mdpi.com/1424-8247/4/7/976 Thu, 30 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-30 4 7 Review 976 991 1424-8247 Psychostimulant Drugs and Neuroplasticity 2011-06-30 doi: 10.3390/ph4070976 Emilio Fernandez-Espejo Nieves Rodriguez-Espinosa http://www.mdpi.com/1424-8247/4/7/964 A new, simple, sensitive and accurate spectrophotometric method has been developed for the assay of metoprolol tartrate (MPT), which is based on the complexation of drug with copper(II) [Cu(II)] at pH 6.0, using Britton-Robinson buffer solution, to produce a blue adduct. The latter has a maximum absorbance at 675 nm and obeys Beer’s law within the concentration range 8.5-70 mg/mL. Regression analysis of the calibration data showed a good correlation coefficient (r = 0.998) with a limit of detection of 5.56 mg/mL. The proposed procedure has been successfully applied to the determination of this drug in its tablets. In addition, the spectral data and stability constant for the binuclear copper(II) complex of MPT (Cu2MPT2Cl2) have been reported. http://www.mdpi.com/1424-8247/4/7/964 Tue, 28 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-28 4 7 Article 964 975 1424-8247 Spectrophotometric Determination of Metoprolol Tartrate in Pharmaceutical Dosage Forms on Complex Formation with Cu(II) 2011-06-28 doi: 10.3390/ph4070964 Mustafa Cesme Derya Tarinc Aysegul Golcu http://www.mdpi.com/1424-8247/4/7/950 To determine the hapten number in hapten-carrier protein conjugate matrix-assisted laser desorption/ionization (MALDI) tof mass spectrometry was applied. Highly specific anti-ginsenoside Rb1 and Rg1 monoclonal antibodies (MAbs) were prepared. Ginsenosides were developed on thin layer chromatography (TLC) plates which were covered by a polyvinylidene difluoride (PVDF) membrane resulting in blotting. The membrane was treated with NaIO4 solution to release the aldehyde group on the sugar moiety of the ginsenosides. By treatment of the membrane with a protein solution the ginsenoside-protein conjugation as a Schiff-base occurred, which can function to fix it to the PVDF membrane. A part of the ginsenoside aglycone was reacted with anti-ginsenoside Rb1 MAb, secondary MAb conjugated with enzyme and finally a substrate was added, resulting in a specific and highly sensitive staining that we named Eastern blotting. Furthermore, it makes one-step isolation of ginsenoside Rb1 possible using an immuno-affinity column conjugated with anti-ginsenoside Rb1 MAb. Furthermore, immunoaffinity concentration was carried out allowing high sensitivity analysis of lower concentrations of ginsenoside Rb1 so that several unknown bands could be structurally determined. http://www.mdpi.com/1424-8247/4/7/950 Tue, 28 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-28 4 7 Article 950 963 1424-8247 Monoclonal Antibodies against Small Molecule Natural Products and Their Applications, Eastern Blotting and Knockout Extract 2011-06-28 doi: 10.3390/ph4070950 Yukihiro Shoyama http://www.mdpi.com/1424-8247/4/6/933 The endogenous cannabinoid (eCB) system is emerging as an important component of female reproductive tract physiology. The eCBs anandamide (AEA), 2-arachidonoyl glycerol (2-AG), and N-arachidonoyl glycine (NAGly) were measured in the rat reproductive tract at five time points in the four-day estrous cycle, in acyclic retired breeders (RB), after ovariectomy (OVX), OVX + estrogen (E2), OVX + progesterone (P4), or OVX with E2+P4. eCBs were measured in the uterus, uterine adipose, ovaries, and ovarian adipose using HPLC/MS/MS. Levels of AEA, 2-AG, and NAGly were highest in the estrus phase of the estrous cycle in the uterus, whereas, only NAGly had differences in production in the ovaries across the cycle. All eCBs were lower in RB ovaries; however, the production of eCBs in the uterus of RB and OVX groups was more varied with NAGly showing the lowest levels of production in these groups. Levels of AEA in uterine fat were significantly higher or equivalent to levels in the uterus. However, levels of 2-AG and NAGly were dramatically lower in uterine fat verses the organ. Ovarian fat had significantly lower levels of all three eCBs. These data provide evidence that the hormonal milieu plays a significant and complex role in the production of eCBs in the female rat reproductive tract. http://www.mdpi.com/1424-8247/4/6/933 Thu, 23 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-23 4 6 Article 933 949 1424-8247 Endogenous Cannabinoid Production in the Rat Female Reproductive Tract Is Regulated by Changes in the Hormonal Milieu 2011-06-23 doi: 10.3390/ph4060933 Heather B. Bradshaw Cassandra Allard http://www.mdpi.com/1424-8247/4/6/915 The capacity of the brain to generate new adult neurons is a recent discovery that challenges the old theory of an immutable adult brain. A new and fascinating field of research now focuses on this regenerative process. The two brain systems that constantly produce new adult neurons, known as the adult neurogenic systems, are the dentate gyrus (DG) of the hippocampus and the lateral ventricules/olfactory bulb system. Both systems are involved in memory and learning processes. Different drugs of abuse, such as cocaine and MDMA, have been shown to produce cellular and molecular changes that affect adult neurogenesis. This review summarizes the effects that these drugs have on the adult neurogenic systems. The functional relevance of adult neurogenesis is obscured by the functions of the systems that integrate adult neurons. Therefore, we explore the effects that cocaine and MDMA produce not only on adult neurogenesis, but also on the DG and olfactory bulbs. Finally, we discuss the possible role of new adult neurons in cocaine- and MDMA-induced impairments. We conclude that, although harmful drug effects are produced at multiple physiological and anatomical levels, the specific consequences of reduced hippocampus neurogenesis are unclear and require further exploration. http://www.mdpi.com/1424-8247/4/6/915 Fri, 17 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-17 4 6 Review 915 932 1424-8247 Cocaine and MDMA Induce Cellular and Molecular Changes in Adult Neurogenic Systems: Functional Implications 2011-06-17 doi: 10.3390/ph4060915 Vivian Capilla-Gonzalez Vicente Hernandez-Rabaza http://www.mdpi.com/1424-8247/4/6/880 Sigma1 receptors (σ1Rs) represent a structurally unique class of intracellular proteins that function as chaperones. σ1Rs translocate from the mitochondria-associated membrane to the cell nucleus or cell membrane, and through protein-protein interactions influence several targets, including ion channels, G-protein-coupled receptors, lipids, and other signaling proteins. Several studies have demonstrated that σR antagonists block stimulant-induced behavioral effects, including ambulatory activity, sensitization, and acute toxicities. Curiously, the effects of stimulants have been blocked by σR antagonists tested under place-conditioning but not self-administration procedures, indicating fundamental differences in the mechanisms underlying these two effects. The self administration of σR agonists has been found in subjects previously trained to self administer cocaine. The reinforcing effects of the σR agonists were blocked by σR antagonists. Additionally, σR agonists were found to increase dopamine concentrations in the nucleus accumbens shell, a brain region considered important for the reinforcing effects of abused drugs. Although the effects of the σR agonist, DTG, on dopamine were obtained at doses that approximated those that maintained self administration behavior those of another agonist, PRE-084 required higher doses. The effects of DTG were antagonized by non-selective or a preferential σ2R antagonist but not by a preferential σ1R antagonist. The effects of PRE-084 on dopamine were insensitive to σR antagonists. The data suggest that the self administration of σR agonists is independent of dopamine and the findings are discussed in light of a hypothesis that cocaine has both intracellular actions mediated by σRs, as well as extracellular actions mediated through conventionally studied mechanisms. The co-activation and potential interactions among these mechanisms, in particular those involving the intracellular chaperone σRs, may lead to the pernicious addictive effects of stimulant drugs. http://www.mdpi.com/1424-8247/4/6/880 Fri, 17 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-17 4 6 Review 880 914 1424-8247 A Role for Sigma Receptors in Stimulant Self Administration and Addiction 2011-06-17 doi: 10.3390/ph4060880 Jonathan L. Katz Tsung-Ping Su Takato Hiranita Teruo Hayashi Gianluigi Tanda Theresa Kopajtic Shang-Yi Tsai http://www.mdpi.com/1424-8247/4/6/848 Hearing loss affects hundreds of millions of people worldwide by dampening or cutting off their auditory connection to the world. Current treatments for sensorineural hearing loss (SNHL) with cochlear implants are not perfect, leaving regenerative medicine as the logical avenue to a perfect cure. Multiple routes to regeneration of damaged hair cells have been proposed and are actively pursued. Each route not only requires a keen understanding of the molecular basis of ear development but also faces the practical limitations of stem cell regulation in the delicate inner ear where topology of cell distribution is essential. Improvements in our molecular understanding of the minimal essential genes necessary for hair cell formation and recent advances in stem cell manipulation, such as seen with inducible pluripotent stem cells (iPSCs) and epidermal neural crest stem cells (EPI-NCSCs), have opened new possibilities to advance research in translational stem cell therapies for individuals with hearing loss. Despite this, more detailed network maps of gene expression are needed, including an appreciation for the roles of microRNAs (miRs), key regulators of transcriptional gene networks. To harness the true potential of stem cells for hair cell regeneration, basic science and clinical medicine must work together to expedite the transition from bench to bedside by elucidating the full mechanisms of inner ear hair cell development, including a focus on the role of miRs, and adapting this knowledge safely and efficiently to stem cell technologies. http://www.mdpi.com/1424-8247/4/6/848 Fri, 17 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-17 4 6 Review 848 879 1424-8247 Regeneration of Hair Cells: Making Sense of All the Noise 2011-06-17 doi: 10.3390/ph4060848 Benjamin Kopecky Bernd Fritzsch http://www.mdpi.com/1424-8247/4/6/822 Amphetamine derivatives such as methamphetamine (METH) and 3,4-methylenedioxymethamphetamine (MDMA, “ecstasy”) are widely abused drugs in a recreational context. This has led to concern because of the evidence that they are neurotoxic in animal models and cognitive impairments have been described in heavy abusers. The main targets of these drugs are plasmalemmal and vesicular monoamine transporters, leading to reverse transport and increased monoamine efflux to the synapse. As far as neurotoxicity is concerned, increased reactive oxygen species (ROS) production seems to be one of the main causes. Recent research has demonstrated that blockade of a7 nicotinic acetylcholine receptors (nAChR) inhibits METH- and MDMA-induced ROS production in striatal synaptosomes which is dependent on calcium and on NO-synthase activation. Moreover, a7 nAChR antagonists (methyllycaconitine and memantine) attenuated in vivo the neurotoxicity induced by METH and MDMA, and memantine prevented the cognitive impairment induced by these drugs. Radioligand binding experiments demonstrated that both drugs have affinity to a7 and heteromeric nAChR, with MDMA showing lower Ki values, while fluorescence calcium experiments indicated that MDMA behaves as a partial agonist on a7 and as an antagonist on heteromeric nAChR. Sustained Ca increase led to calpain and caspase-3 activation. In addition, modulatory effects of MDMA on a7 and heteromeric nAChR populations have been found. http://www.mdpi.com/1424-8247/4/6/822 Wed, 15 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-15 4 6 Review 822 847 1424-8247 Neuronal Nicotinic Receptors as New Targets for Amphetamine-Induced Oxidative Damage and Neurotoxicity 2011-06-15 doi: 10.3390/ph4060822 David Pubill Sara Garcia-Ratés Jordi Camarasa Elena Escubedo http://www.mdpi.com/1424-8247/4/6/804 Drug addiction is a chronic illness characterized by high rates of relapse. Relapse to drug use can be triggered by re-exposure to drug-associated cues, stressful events, or the drug itself after a period of abstinence. Pharmacological intervention to reduce the impact of relapse-instigating factors offers a promising target for addiction treatment. Growing evidence has implicated an important role of the orexin/hypocretin system in drug reward and drug-seeking, including animal models of relapse. Here, we review the evidence for the role of orexins in modulating reward and drug-seeking in animal models of addiction and the potential for orexin receptors as specific targets for anti-relapse medication approaches. http://www.mdpi.com/1424-8247/4/6/804 Tue, 14 Jun 2011 00:00:00 CEST Pharmaceuticals 2011-06-14 4 6 Review 804 821 1424-8247 Orexin Receptor Targets for Anti-Relapse Medication Development in Drug Addiction 2011-06-14 doi: 10.3390/ph4060804 Luyi Zhou Wei-Lun Sun Ronald E. See http://www.mdpi.com/1424-8247/4/6/794 Background: Both nifekalant hydrochloride (NIF), a selective IKr blocker, and intravenous amiodarone (AMD), a multi-channel (including IKr blocking) blocker, have been reported to be efficacious for refractory arrhythmias. However, the optimal use of those antiarrhythmic drugs for refractory arrhythmia with severe heart failure has not been established. Intravenous AMD might be effective for arrhythmias refractory to NIF in patients with severe heart failure. Here, we report that intravenous amiodarone was effective in the treatment of nifekalant-resistant in a group of arrhythmia patients with severe heart failure. Methods: Eleven severe heart failure patients who had received intravenous AMD for treatment of NIF-resistant arrhythmias were included in this study, and retrospective analysis was performed. Clinical efficacy (terminative and preventive effects on arrhythmia) of intravenous AMD was evaluated. Results: All cases were emergent cases and had depressed left ventricular ejection fraction (30 ± 13%). Clinical arrhythmias were ventricular fibrillation (VF) in four patients, ventricular tachycardia (VT) in six patients, and atrial fibrillation (AF) in one patient. NIF was administered to all patients by intravenous injection. After administration of NIF, VT/VF/AF was terminated in seven of the 10 patients, but a preventive effect was not obtained in any of the patients (NIF-resistance). Intravenous AMD (maintenance dose: 484 ± 166 mg/day) was effective both in termination (80%) and in prevention (80%) of VT/VF events in those patients. It was also effective in termination (80%) and prevention (60%) of AF events refractory to NIF. During continuous AMD administration, no significant adverse effects or proarrhythmic effects were observed in any of the patients. Five patients died within one month, but there was no arrhythmic deaths. Conclusions: Intravenous AMD was effective in NIF-resistant lethal arrhythmias and was relatively safe in emergent cases with severe heart failure. http://www.mdpi.com/1424-8247/4/6/794 Tue, 31 May 2011 00:00:00 CEST Pharmaceuticals 2011-05-31 4 6 Article 794 803 1424-8247 Use of Intravenous Amiodarone in the Treatment of Nifekalant-Resistant Arrhythmia: A Review of 11 Consecutive Cases with Severe Heart Failure 2011-05-31 doi: 10.3390/ph4060794 Koji Nakagawa Kazufumi Nakamura Kengo Fukushima Kusano Satoshi Nagase Takeshi Tada Masato Murakami Yoshiki Hata Hiroshi Morita Kunihisa Kohno Kazumasa Hina Tohru Ujihira Tohru Ohe Hiroshi Ito http://www.mdpi.com/1424-8247/4/6/782 Vasohibin-1 (VASH1) is a VEGF-inducible gene of endothelial cells (ECs) that acts as a negative feedback regulator of angiogenesis. To further characterize the function of VASH1, we transfected human VASH1 gene into the mouse EC line MS1, established stable VASH1 expressing clones, and determined gene alteration by cDNA microarray analysis. Among the various angiogenesis-related genes, vascular endothelial growth factor type 1 receptor (VEGFR-1) and its alternative spliced form, soluble VEGFR1 (sVEGFR-1), were found to be the most significantly down-regulated genes. Transient overexpression of VASH1 in human umbilical vein endothelial cells confirmed the down-regulation of VEGFR-1 and sVEGFR-1. sVEGFR-1 is a decoy receptor for VEGF and inhibits angiogenesis. Interestingly, when sVEGFR-1 was overexpressed in ECs, it inhibited the expression of VASH1 in turn. These results suggest that VASH1 and sVEGFR-1, two angiogenesis inhibitors, mutually balance their expressions in ECs. http://www.mdpi.com/1424-8247/4/6/782 Tue, 31 May 2011 00:00:00 CEST Pharmaceuticals 2011-05-31 4 6 Article 782 793 1424-8247 Mutual Balance between Vasohibin-1 and Soluble VEGFR-1 in Endothelial Cells 2011-05-31 doi: 10.3390/ph4060782 Hiroki Miyashita Hirotada Suzuki Akihide Ohkuchi Yasufumi Sato http://www.mdpi.com/1424-8247/4/5/770 Although chronic sympathetic activation provides inotropic and chronotropic support to the failing heart, such activation may also have deleterious effects, including the direct cardiotoxic effects of catecholamines, activation of the renin-angiotensin-aldosterone system and an increase in myocardial oxygen demand. These observations indicate that β-blockade might be beneficial in the treatment of heart failure resulting from dilated cardiomyopathy or ischaemic heart disease. Carvedilol is a non-selective β-blocker acting on β1-, β2-, and α1-adrenoceptors. It possesses potent anti-oxidant and anti-apoptotic properties, along with neuroprotective, vasculoprotective, cardioprotective effects, and it has reduced overall mortality in patients with heart failure in controlled clinical trials. Its role in treating cardiomyopathy requires focus. The fact that anthracyclines are cardiotoxic seriously narrows their therapeutic index in cancer therapy. The cardiotoxic risk increases with the cumulative dose and may lead to congestive heart failure and dilated cardiomyopathy in adults and in children. This review focuses on recent research regarding the beneficial effects of carvedilol in the treatment of dilated cardiomyopathy and to revisit the available evidence on the cardioprotection of carvedilol when associated with anthracycline and to explain the mechanisms underlying the benefits of their co-administration. http://www.mdpi.com/1424-8247/4/5/770 Tue, 24 May 2011 00:00:00 CEST Pharmaceuticals 2011-05-24 4 5 Review 770 781 1424-8247 The Role of Carvedilol in the Treatment of Dilated and Anthracyclines-Induced Cardiomyopathy 2011-05-24 doi: 10.3390/ph4050770 Kenichi Watanabe Wawaimuli Arozal Flori R. Sari Somasundaram Arumugam Rajarajan A. Thandavarayan Kenji Suzuki Makoto Kodama http://www.mdpi.com/1424-8247/4/5/758 We have developed a computational method that predicts the positions of active compounds, making it possible to increase activity as a fragment evolution strategy. We refer to the positions of these compounds as the active position. When an active fragment compound is found, the following lead generation process is performed, primarily to increase activity. In the current method, to predict the location of the active position, hydrogen atoms are replaced by small side chains, generating virtual compounds. These virtual compounds are docked to a target protein, and the docking scores (affinities) are examined. The hydrogen atom that gives the virtual compound with good affinity should correspond to the active position and it should be replaced to generate a lead compound. This method was found to work well, with the prediction of the active position being 2 times more efficient than random synthesis. In the current study, 15 examples of lead generation were examined. The probability of finding active positions among all hydrogen atoms was 26%, and the current method accurately predicted 60% of the active positions. http://www.mdpi.com/1424-8247/4/5/758 Fri, 20 May 2011 00:00:00 CEST Pharmaceuticals 2011-05-20 4 5 Article 758 769 1424-8247 Prediction of Positions of Active Compounds Makes It Possible To Increase Activity in Fragment-Based Drug Development 2011-05-20 doi: 10.3390/ph4050758 Yoshifumi Fukunishi http://www.mdpi.com/1424-8247/4/5/741 Eukaryotic cells comprise a set of organelles, surrounded by membranes with a unique composition, which is maintained by a complex synthesis and transport system. Cells also synthesize the proteins destined for secretion. Together, these processes are known as the secretory pathway or exocytosis. In addition, many molecules can be internalized by cells through a process called endocytosis. Chronic and acute alcohol (ethanol) exposure alters the secretion of different essential products, such as hormones, neurotransmitters and others in a variety of cells, including central nervous system cells. This effect could be due to a range of mechanisms, including alcohol-induced alterations in the different steps involved in intracellular transport, such as glycosylation and vesicular transport along cytoskeleton elements. Moreover, alcohol consumption during pregnancy disrupts developmental processes in the central nervous system. No single mechanism has proved sufficient to account for these effects, and multiple factors are likely involved. One such mechanism indicates that ethanol also perturbs protein trafficking. The purpose of this review is to summarize our understanding of how ethanol exposure alters the trafficking of proteins in different cell systems, especially in central nervous system cells (neurons and astrocytes) in adult and developing brains. http://www.mdpi.com/1424-8247/4/5/741 Tue, 17 May 2011 00:00:00 CEST Pharmaceuticals 2011-05-17 4 5 Review 741 757 1424-8247 Protein Traffic Is an Intracellular Target in Alcohol Toxicity 2011-05-17 doi: 10.3390/ph4050741 Guillermo Esteban-Pretel María Pilar Marín Ana M. Romero Xavier Ponsoda Raul Ballestin Juan J. Canales Jaime Renau-Piqueras http://www.mdpi.com/1424-8247/4/5/726 Hormone replacement after menopause has in recent years been the subject of intense scientific debate and public interest and has sparked intense research efforts into the biological effects of estrogens and progestagens. However, there are reasons to believe that the doses used and plasma concentrations produced in a large number of studies casts doubt on important aspects of their validity. The concept of hormesis states that a substance can have diametrically different effects depending on the concentration. Even though estrogens and progestagens have proven prone to this kind of dose-response relation in a multitude of studies, the phenomenon remains clearly underappreciated as exemplified by the fact that it is common practice to only use one hormone dose in animal experiments. If care is not taken to adjust the concentrations of estrogens and progestagens to relevant biological conditions, the significance of the results may be questionable. Our aim is to review examples of female sexual steroids demonstrating bidirectional dose-response relations and to discuss this in the perspective of hormesis. Some examples are highlighted in detail, including the effects on cerebral ischemia, inflammation, cardiovascular diseases and anxiety. Hopefully, better understanding of the hormesis phenomenon may result in improved future designs of studies of female sexual steroids. http://www.mdpi.com/1424-8247/4/5/726 Mon, 16 May 2011 00:00:00 CEST Pharmaceuticals 2011-05-16 4 5 Review 726 740 1424-8247 Hormesis and Female Sex Hormones 2011-05-16 doi: 10.3390/ph4050726 Jakob O. Strom Annette Theodorsson Elvar Theodorsson http://www.mdpi.com/1424-8247/4/5/713 Several, if not all adrenergic β-blockers (β-Bs), accumulate progressively inside secretory vesicles in a time- and concentration-dependent manner, and could be considered to be false neurotransmitters. This transmitter effect is most likely unrelated to their ability to block adrenergic receptors, but it could explain the delay in lowering arterial pressure in hypertensive patients. We have developed a new drug to monitor the accumulation of β-Bs inside living cells, RCTM-3, which fluoresces in the visible spectrum. Here we describe the procedure to synthesize this new compound, as well as its fluorescent properties, pharmacological profile and its accumulation inside the secretory vesicles of PC12 cells. http://www.mdpi.com/1424-8247/4/5/713 Thu, 28 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-28 4 5 Article 713 725 1424-8247 Fluorescent β-Blockers as Tools to Study Presynaptic Mechanisms of Neurosecretion 2011-04-28 doi: 10.3390/ph4050713 Beatriz Beltran Romen Carrillo Tomas Martin Victor S. Martin Jose D. Machado Ricardo Borges http://www.mdpi.com/1424-8247/4/5/681 The number of available and effective antimalarial drugs is quickly dwindling. This is mainly because a number of drug resistance-associated mutations in malaria parasite genes, such as crt, mdr1, dhfr/dhps, and others, have led to widespread resistance to all known classes of antimalarial compounds. Unfortunately, malaria parasites have started to exhibit some level of resistance in Southeast Asia even to the most recently introduced class of drugs, artemisinins. While there is much need, the antimalarial drug development pipeline remains woefully thin, with little chemical diversity, and there is currently no alternative to the precious artemisinins. It is difficult to predict where the next generation of antimalarial drugs will come from; however, there are six major approaches: (i) re-optimizing the use of existing antimalarials by either replacement/rotation or combination approach; (ii) repurposing drugs that are currently used to treat other infections or diseases; (iii) chemically modifying existing antimalarial compounds; (iv) exploring natural sources; (v) large-scale screening of diverse chemical libraries; and (vi) through parasite genome-based (“targeted”) discoveries. When any newly discovered effective antimalarial treatment is used by the populus, we must maintain constant vigilance for both parasite-specific and human-related factors that are likely to hamper its success. This article is neither comprehensive nor conclusive. Our purpose is to provide an overview of antimalarial drug resistance, associated parasite genetic factors (1. Introduction; 2. Emergence of artemisinin resistance in P. falciparum), and the antimalarial drug development pipeline (3. Overview of the global pipeline of antimalarial drugs), and highlight some examples of the aforementioned approaches to future antimalarial treatment. These approaches can be categorized into “short term” (4. Feasible options for now) and “long term” (5. Next generation of antimalarial treatment—Approaches and candidates). However, these two categories are interrelated, and the approaches in both should be implemented in parallel with focus on developing a successful, long-lasting antimalarial chemotherapy. http://www.mdpi.com/1424-8247/4/5/681 Tue, 26 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-26 4 5 Review 681 712 1424-8247 Expanding the Antimalarial Drug Arsenal—Now, But How? 2011-04-26 doi: 10.3390/ph4050681 Brian T. Grimberg Rajeev K. Mehlotra http://www.mdpi.com/1424-8247/4/4/665 Pain is a major symptom in many medical conditions, and often interferes significantly with a person’s quality of life. Although a priority topic in medical research for many years, there are still few analgesic drugs approved for clinical use. One reason is the lack of appropriate animal models that faithfully represent relevant hallmarks associated with human pain. Here we propose zebrafish (Danio rerio) as a novel short-term behavioral model of nociception, and analyse its sensitivity and robustness. Firstly, we injected two different doses of acetic acid as the noxious stimulus. We studied individual locomotor responses of fish to a threshold level of nociception using two recording systems: a video tracking system and an electric biosensor (the MOBS system). We showed that an injection dose of 10% acetic acid resulted in a change in behavior that could be used to study nociception. Secondly, we validated our behavioral model by investigating the effect of the analgesic morphine. In time-course studies, first we looked at the dose-response relationship of morphine and then tested whether the effect of morphine could be modulated by naloxone, an opioid antagonist. Our results suggest that a change in behavioral responses of zebrafish to acetic acid is a reasonable model to test analgesics. The response scales with stimulus intensity, is attenuated by morphine, and the analgesic effect of morphine is blocked with naloxone. The change in behavior of zebrafish associated with the noxious stimulus can be monitored with an electric biosensor that measures changes in water impedance. http://www.mdpi.com/1424-8247/4/4/665 Mon, 18 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-18 4 4 Article 665 680 1424-8247 A Novel Behavioral Fish Model of Nociception for Testing Analgesics 2011-04-18 doi: 10.3390/ph4040665 Ana D. Correia Sérgio R. Cunha Martin Scholze E. Don Stevens http://www.mdpi.com/1424-8247/4/4/652 An understanding of the functional mechanisms of G-protein-coupled receptors (GPCRs) is very important for GPCR-related drug design. We have developed an integrated GPCR database (SEVENS http://sevens.cbrc.jp/) that includes 64,090 reliable GPCR genes comprehensively identified from 56 eukaryote genome sequences, and overviewed the sequences and structure spaces of the GPCRs. In vertebrates, the number of receptors for biological amines, peptides, etc. is conserved in most species, whereas the number of chemosensory receptors for odorant, pheromone, etc. significantly differs among species. The latter receptors tend to be single exon type or a few exon type and show a high ratio in the numbers of GPCRs, whereas some families, such as Class B and Class C receptors, have long lengths due to the presence of many exons. Statistical analyses of amino acid residues reveal that most of the conserved residues in Class A GPCRs are found in the cytoplasmic half regions of transmembrane (TM) helices, while residues characteristic to each subfamily found on the extracellular half regions. The 69 of Protein Data Bank (PDB) entries of complete or fragmentary structures could be mapped on the TM/loop regions of Class A GPCRs covering 14 subfamilies. http://www.mdpi.com/1424-8247/4/4/652 Wed, 13 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-13 4 4 Review 652 664 1424-8247 Functional and Structural Overview of G-Protein-Coupled Receptors Comprehensively Obtained from Genome Sequences 2011-04-13 doi: 10.3390/ph4040652 Makiko Suwa Minoru Sugihara Yukiteru Ono http://www.mdpi.com/1424-8247/4/4/630 We sought to determine the involvement of phosphatidyl inositol 3-kinase (PI3K) and AMP-activated protein kinase (AMPK) in the estrogenic antagonism of the cannabinoid regulation of energy homeostasis. Food intake and body weight were evaluated in ovariectomized female guinea pigs treated s.c. with estradiol benzoate (EB) or its sesame oil vehicle, or the CB1 receptor antagonist AM251 or its cremephor/ethanol/0.9% saline vehicle. AMPK catalytic subunit, PI3K p85α regulatory subunit and proopiomelanocortin (POMC) gene expression was assessed via quantitative RT-PCR in microdissected hypothalamic tissue. Whole-cell patch clamp recordings were performed in hypothalamic slices. Both EB and AM251 decreased food intake and weight gain, and increased AMPKα1, AMPKα2 and PI3K p85α gene expression in the mediobasal hypothalamus. 17β-Estradiol rapidly and markedly attenuated the decreases in glutamatergic miniature excitatory postsynaptic current (mEPSC) frequency caused by the cannabinoid receptor agonist WIN 55,212-2 in POMC neurons. This rapid estrogenic diminution of cannabinoid-induced decreases in mEPSC frequency was blocked by the estrogen receptor (ER) antagonist ICI 182,780 and the PI3K inhibitor PI 828, the latter of which also prevented the AM251-induced increase in mEPSC frequency. In addition, the AMPK activator metformin reversed the EB-induced decreases in food intake and weight gain and restored the ability of WIN 55,212-2 to reduce mEPSC frequency. These data reveal that estrogens physiologically antagonize cannabinoid-induced changes in appetite and POMC neuronal activity by activating PI3K and inhibiting AMPK. As such, they provide insight into the neuroanatomical substrates and signal transduction mechanisms upon which these counter-regulatory factors converge in the control of energy homeostasis. http://www.mdpi.com/1424-8247/4/4/630 Tue, 12 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-12 4 4 Article 630 651 1424-8247 The Role of Phosphatidylinositol-3-Kinase and AMP-Activated Kinase in the Rapid Estrogenic Attenuation of Cannabinoid-Induced Changes in Energy Homeostasis 2011-04-12 doi: 10.3390/ph4040630 Garrett S. Jeffery Kelly C. Peng Edward J. Wagner http://www.mdpi.com/1424-8247/4/4/603 Serotonin (5-HT)3 receptors are the only ligand-gated ion channel of the 5-HT receptors family. They are present both in the peripheral and central nervous system and are localized in several areas involved in mood regulation (e.g., hippocampus or prefrontal cortex). Moreover, they are involved in regulation of neurotransmitter systems implicated in the pathophysiology of major depression (e.g., dopamine or GABA). Clinical and preclinical studies have suggested that 5-HT3 receptors may be a relevant target in the treatment of affective disorders. 5-HT3 receptor agonists seem to counteract the effects of antidepressants in non-clinical models, whereas 5-HT3 receptor antagonists, such as ondansetron, present antidepressant-like activities. In addition, several antidepressants, such as mirtazapine, also target 5-HT3 receptors. In this review, we will report major advances in the research of 5-HT3 receptor’s roles in neuropsychiatric disorders, with special emphasis on mood and anxiety disorders. http://www.mdpi.com/1424-8247/4/4/603 Thu, 07 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-07 4 4 Review 603 629 1424-8247 Role of 5-HT3 Receptors in the Antidepressant Response 2011-04-07 doi: 10.3390/ph4040603 Cécile Bétry Adeline Etiévant Chris Oosterhof Bjarke Ebert Connie Sanchez Nasser Haddjeri http://www.mdpi.com/1424-8247/4/4/590 Some fragrance components of spices used for cooking are known to have an effect on human behavior. The aim of this investigation was to examine the effect of the essential oils of basil (Ocimum formacitratum L.) leaves, lemongrass (Cymbopogon citrates L.) herbs, ki lemo (Litsea cubeba L.) bark, and laja gowah (Alpinia malaccencis Roxb.) rhizomes on locomotor activity in mice and identify the active component(s) that might be responsible for the activity. The effect of the essential oils was studied by a wheel cage method and the active compounds of the essential oils were identified by GC/MS analysis. The essential oils were administered by inhalation at doses of 0.1, 0.3, and 0.5 mL/cage. The results showed that the four essential oils had inhibitory effects on locomotor activity in mice. Inhalation of the essential oils of basil leaves, lemongrass herbs, ki lemo bark, and laja gowah rhizomes showed the highest inhibitory activity at doses of 0.5 (57.64%), 0.1 (55.72%), 0.5 (60.75%), and 0.1 mL/cage (47.09%), respectively. The major volatile compounds 1,8-cineole, α-terpineol, 4-terpineol, citronelol, citronelal, and methyl cinnamate were identified in blood plasma of mice after inhalation of the four oils. These compounds had a significant inhibitory effect on locomotion after inhalation. The volatile compounds of essential oils identified in the blood plasma may correlate with the locomotor-inhibiting properties of the oil when administered by inhalation. http://www.mdpi.com/1424-8247/4/4/590 Wed, 06 Apr 2011 00:00:00 CEST Pharmaceuticals 2011-04-06 4 4 Article 590 602 1424-8247 Analysis of Indonesian Spice Essential Oil Compounds That Inhibit Locomotor Activity in Mice 2011-04-06 doi: 10.3390/ph4040590 Muchtaridi Adjeng Diantini Anas Subarnas http://www.mdpi.com/1424-8247/4/4/567 Hundreds of G protein coupled receptor (GPCR) isotypes integrate and coordinate the function of individual cells mediating signaling between different organs in our bodies. As an aberration of the normal relationships that organize cells’ coexistence, cancer has to deceive cell-cell communication in order to grow and spread. GPCRs play a critical role in this process. Despite the fact that GPCRs represent one of the most common drug targets, current medical practice includes only a few anticancer compounds directly acting on their signaling. Many approaches can be envisaged to target GPCRs involved in oncology. Beyond interfering with GPCRs signaling by using agonists or antagonists to prevent cell proliferation, favor apoptosis, induce maturation, prevent migration, etc., the high specificity of the interaction between the receptors and their ligands can be exploited to deliver toxins, antineoplastic drugs or isotopes to transformed cells. In this review we describe the strategies that are in use, or appear promising, to act directly on GPCRs in the fight against neoplastic transformation and tumor progression. http://www.mdpi.com/1424-8247/4/4/567 Fri, 25 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-25 4 4 Review 567 589 1424-8247 Molecular Approaches To Target GPCRs in Cancer Therapy 2011-03-25 doi: 10.3390/ph4040567 Giulio Innamorati Maria Teresa Valenti Francesco Giovinazzo Luca Dalle Carbonare Marco Parenti Claudio Bassi http://www.mdpi.com/1424-8247/4/3/551 Cardiotoxicity, which results from intense cardiac oxidative stress and inflammation, is the main limiting factor of the anthracyclines. Carvedilol, a beta blocker that is used as a multifunctional neurohormonal antagonist, has been shown to act not only as an anti-oxidant, but also as an anti-inflammatory drug. This study was designed to evaluate whether carvedilol exerts a protective role against inflammation-mediated cardiotoxicity in the daunorubicin (DNR)-induced rats. Carvedilol was administered orally to the rats every day for 6 weeks at a cumulative dose of 9 mg/kg body weight DNR. DNR significantly induced cardiac damage and worsened cardiac function as well as increased cardiac mast cell density, elevating the myocardial protein and mRNA expression levels of tumor necrosis factor-α, vascular cell adhesion molecule-1, inter-cellular adhesion molecule-1, nuclear factor kappa-B, cyclooxygenase-2, monocyte chemotactic protein -1 and interleukin -6 compared to that in the control group. Cotreatment with carvedilol significantly attenuated the myocardial protein and mRNA expression levels of these inflammatory markers, decreased cardiac mast cell density, improved histological cardiac damage and cardiac functions. In conclusion, inflammation plays a significant role in DNR-induced cardiotoxicity, and carvedilol contributes to cardioprotection against inflammation-mediated cardiotoxicity in DNR-induced rats through its anti-inflammatory mechanism. http://www.mdpi.com/1424-8247/4/3/551 Thu, 17 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-17 4 3 Article 551 566 1424-8247 Carvedilol Attenuates Inflammatory-Mediated Cardiotoxicity in Daunorubicin-Induced Rats 2011-03-17 doi: 10.3390/ph4030551 Flori R. Sari Wawaimuli Arozal Kenichi Watanabe Meilei Harima Punniyakoti T. Veeravedu Rajarajan A. Thandavarayan Kenji Suzuki Somasundaram Arumugam Vivian Soetikno Makoto Kodama http://www.mdpi.com/1424-8247/4/3/524 Many signaling molecules involved in G protein-mediated signal transduction, which are present in the lipid rafts and believed to be controlled spatially and temporally, influence the potency and efficacy of neurotransmitter receptors and transporters. This has focus interest on lipid rafts and the notion that these microdomains acts as a kind of signaling platform and thus have an important role in the expression of membrane receptor-mediated signal transduction, cancer, immune responses, neurotransmission, viral infections and various other phenomena due to specific and efficient signaling according to extracellular stimuli. However, the real structure of lipid rafts has not been observed so far due to its small size and a lack of sufficiently sophisticated observation systems. A soft X-ray microscope using a coherent soft X-ray laser in the water window region (2.3–4.4 nm) should prove to be a most powerful tool to observe the dynamic structure of lipid rafts of several tens of nanometers in size in living cells. We have developed for the X-ray microscope a new compact soft X-ray laser using strongly induced plasma high harmonic resonance. We have also developed a time-resolved highly sensitive fluorescence resonance energy transfer (FRET) system and confirmed protein-protein interactions coupled with ligands. The simultaneous use of these new tools for observation of localization of G-protein coupled receptors (GPCRs) in rafts has become an important and optimum tool system to analyze the dynamics of signal transduction through rafts as signaling platform. New technology to visualize rafts is expected to lead to the understanding of those dynamics and innovative development of drug discovery that targets GPCRs localized in lipid rafts. http://www.mdpi.com/1424-8247/4/3/524 Mon, 14 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-14 4 3 Review 524 550 1424-8247 Soft X-ray Laser Microscopy of Lipid Rafts towards GPCR-Based Drug Discovery Using Time-Resolved FRET Spectroscopy 2011-03-14 doi: 10.3390/ph4030524 Motoyoshi Baba Tohru Kozasa Takao Hamakubo Hiroto Kuroda Kazuyuki Masuda Shin Yoneya Tatsuhiko Kodama http://www.mdpi.com/1424-8247/4/3/509 G Protein Coupled Receptors (GPCRs) represent the largest family of membrane proteins in the human genome, are the targets of approximately 25% of all marketed pharmaceuticals, and the focus of intensive research worldwide given that this superfamily of receptors is as varied in function as it is ubiquitously expressed among all cell types. Increasing evidence has shown that the classical two part model of GPCR signaling (one GPCR, one type of heterotrimeric G protein) is grossly oversimplified as many GPCRs can couple to more than one type of G protein, each subunit of the heterotrimeric G protein can activate different downstream effectors, and, surprisingly, other GPCRs can affect receptor behavior in G protein-independent ways. The concept of GPCR heterodimerization, or the physical association of two different types of GPCRs, presents an unexpected mechanism for GPCR regulation and function, and provides a novel target for pharmaceuticals. Here we present a synopsis of the functional consequences of GPCR heterodimerization in both in vitro and in vivo studies, focusing on the concept of GPCRs as allosteric modulators. Typically, an allosteric modulator is a ligand or molecule that alters a receptor’s innate functional properties, but here we propose that in the case of GPCR heterodimers, it is the physical coupling of two receptors that leads to changes in cognate receptor signaling. http://www.mdpi.com/1424-8247/4/3/509 Mon, 14 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-14 4 3 Review 509 523 1424-8247 Functional Consequences of GPCR Heterodimerization: GPCRs as Allosteric Modulators 2011-03-14 doi: 10.3390/ph4030509 Karla K.V. Haack Nael A. McCarty http://www.mdpi.com/1424-8247/4/3/494 Many genetic and infectious diseases can be targeted at the RNA level as RNA is more accessible than DNA. We seek to develop new approaches for detection and tracking RNA in live cells, which is necessary for RNA-based diagnostics and therapy. We recently described a method for RNA visualization in live bacterial cells based on fluorescent protein complementation [1-3]. The RNA is tagged with an RNA aptamer that binds an RNA-binding protein with high affinity. This RNA-binding protein is expressed as two split fragments fused to the fragments of a split fluorescent protein. In the presence of RNA the fragments of the RNA-binding protein bind the aptamer and bring together the fragments of the fluorescent protein, which results in its re-assembly and fluorescence development [1-3]. Here we describe a new version of the RNA labeling method where fluorescent protein complementation is triggered by paired interactions of two different closely-positioned RNA aptamers with two different RNA-binding viral peptides. The new method, which has been developed in bacteria as a model system, uses a smaller ribonucleoprotein complementation complex, as compared with the method using split RNA-binding protein, and it can potentially be applied to a broad variety of RNA targets in both prokaryotic and eukaryotic cells. We also describe experiments exploring background fluorescence in these RNA detection systems and conditions that improve the signal-to-background ratio. http://www.mdpi.com/1424-8247/4/3/494 Thu, 10 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-10 4 3 Article 494 508 1424-8247 RNA Detection in Live Bacterial Cells Using Fluorescent Protein Complementation Triggered by Interaction of Two RNA Aptamers with Two RNA-Binding Peptides 2011-03-10 doi: 10.3390/ph4030494 Hung-Wei Yiu Vadim V. Demidov Paul Toran Charles R. Cantor Natalia E. Broude http://www.mdpi.com/1424-8247/4/3/485 The vasoactive intestinal peptide/pituitary adenylyl cyclase-activating polypepetide (VPAC) receptors are important for many physiologic functions, including glucose homeostasis, neuroprotection, memory, gut function, modulation of the immune system and circadian function. In addition, VPAC receptors have been shown to function in vitro to modulate the infection of HIV by a signal transduction pathway that appears to regulate viral integration. In this article, the affects of VPAC stimulation on HIV infection will be reviewed and approaches for the development of HIV/AIDS therapeutics that target these receptors will be described. Novel HIV/AIDS therapeutics are urgently required to stem the continued spread of this disease, particularly in underdeveloped countries. Drug design to inhibit signaling through VPAC1 and stimulate signaling through VPAC2 could lead to alternative therapies for the treatment and/or prevention of HIV/AIDS. http://www.mdpi.com/1424-8247/4/3/485 Wed, 09 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-09 4 3 Opinion 485 493 1424-8247 Neuropeptide Receptors: Novel Targets for HIV/AIDS Therapeutics 2011-03-09 doi: 10.3390/ph4030485 Donald R. Branch http://www.mdpi.com/1424-8247/4/3/470 Opioid compounds and G-protein coupled opioid receptors (ORs) have been studied widely in terms of central nervous system (CNS) actions relating to pain management and drug abuse. Opioids are also linked to induction of mammalian hibernation, a natural state of tolerance involving prolonged and orchestrated shifts in cellular metabolism, growth and stress resistance. It is not surprising then that OR agonism induces acute or delayed cytoprotective states in myocardium, rendering ORs an attractive target for protection of cardiac tissue from the potentially fatal consequences of ischemic heart disease. Cardiac ORs are implicated in triggering/mediating so-called ‘conditioning’ responses, in which powerful cytoprotection arises following transient receptor ligation prior to or immediately following ischemic insult. These responses involve one or more OR sub-types engaging pro-survival kinase cascades to ultimately modulate cell stress and mitochondrial end-effectors. However, important questions remain regarding the role of endogenous opioids, OR signalling, and the transduction and mediation of these protective responses. We briefly review opioid-mediated cardioprotection, focussing on recent developments in signal transduction, the role of receptor ‘cross-talk’, and the effects of sustained OR ligand activation. http://www.mdpi.com/1424-8247/4/3/470 Fri, 04 Mar 2011 00:00:00 CET Pharmaceuticals 2011-03-04 4 3 Review 470 484 1424-8247 Myocardial Opioid Receptors in Conditioning and Cytoprotection 2011-03-04 doi: 10.3390/ph4030470 Grant Williams-Pritchard John P. Headrick Jason N. Peart http://www.mdpi.com/1424-8247/4/3/457 G protein-coupled receptors (GPCRs) are crucial elements in mammalian signal transduction, and are considered to represent potent drug targets. We have previously developed a GPCR assay system in cultured cells based on complementation of split fragments of click beetle (Pyrearinus termitilluminans) luciferase. The interaction of GPCRs with its target, β-arrestin, resulted in strong emission of bioluminescence upon stimulation with its specific ligand. In this study, we improved precision of the GPCR assay system by using railroad worm (Phrixothrix hirtus) luciferase as an internal control. We generated stable cell lines harboring the railroad worm luciferase and quantitatively evaluate the extent of GPCR-β-arrestin interactions. We showed concentration-dependent bioluminescence responses for four GPCRs: β2-adrenoceptor, endothelin receptor type A, α2-adrenoceptor and human μ-opioid receptor. We also demonstrated that the variation of responses was reduced significantly by normalizing the data with bioluminescence from railroad worm luciferase. This assay system represents a simple and reliable approach for screening drug candidates in a high throughput manner. http://www.mdpi.com/1424-8247/4/3/457 Fri, 25 Feb 2011 00:00:00 CET Pharmaceuticals 2011-02-25 4 3 Article 457 469 1424-8247 Dual-Color Bioluminescence Analysis for Quantitatively Monitoring G-Protein-Coupled Receptor and β-Arrestin Interactions 2011-02-25 doi: 10.3390/ph4030457 A.K.M. Kafi Mitsuru Hattori Naomi Misawa Takeaki Ozawa http://www.mdpi.com/1424-8247/4/3/429 Asthma is a complex, inflammatory disorder characterized by airflow obstruction of variable degrees, bronchial hyper-responsiveness, and airway inflammation. Asthma is caused by environmental factors and a combination of genetic and environmental stimuli. Genetic studies have revealed that multiple loci are involved in the etiology of asthma. Recent cellular, molecular, and animal-model studies have revealed several cellular events that are involved in the progression of asthma, including: increased Th2 cytokines leading to the recruitment of inflammatory cells to the airway, and an increase in the production of reactive oxygen species and mitochondrial dysfunction in the activated inflammatory cells, leading to tissue injury in the bronchial epithelium. Further, aging and animal model studies have revealed that mitochondrial dysfunction and oxidative stress are involved and play a large role in asthma. Recent studies using experimental allergic asthmatic mouse models and peripheral cells and tissues from asthmatic humans have revealed antioxidants as promising treatments for people with asthma. This article summarizes the latest research findings on the involvement of inflammatory changes, and mitochondrial dysfunction/oxidative stress in the development and progression of asthma. This article also addresses the relationship between aging and age-related immunity in triggering asthma, the antioxidant therapeutic strategies in treating people with asthma. http://www.mdpi.com/1424-8247/4/3/429 Fri, 25 Feb 2011 00:00:00 CET Pharmaceuticals 2011-02-25 4 3 Review 429 456 1424-8247 Mitochondrial Dysfunction and Oxidative Stress in Asthma: Implications for Mitochondria-Targeted Antioxidant Therapeutics 2011-02-25 doi: 10.3390/ph4030429 P. Hemachandra Reddy http://www.mdpi.com/1424-8247/4/2/419 An evolution of antibiotic-resistant bacteria has resulted in the need for new antibiotics. β-Lactam based drugs are the most predominantly prescribed antibiotics to combat bacterial infections; however, production of β-lactamases, which catalyze the hydrolysis of the β-lactam bond of this class of antibiotics, by pathogenic bacteria such as Bacillus cereus, are rendering them useless. Some inhibitors of β-lactamases have been found, but there are no inhibitors against a class of β-lactamases known as metallo-β-lactamases, and it has been reported that the number of bacteria that produce metallo-β-lactamases is on the rise. Finding inhibitors of metallo-β-lactamases is thus an urgent necessity. One way to approach the problem is by employing the combinatorial method SELEX. The SELEX method is significant in discovering and producing new classes of inhibitors, as well as providing insight into the development of these inhibitors and paves the way for future aptamer applications that further novel drug discovery. http://www.mdpi.com/1424-8247/4/2/419 Fri, 18 Feb 2011 00:00:00 CET Pharmaceuticals 2011-02-18 4 2 Review 419 428 1424-8247 Metallo-β-Lactamases and Aptamer-Based Inhibition 2011-02-18 doi: 10.3390/ph4020419 Sara R. Schlesinger Mieke J. Lahousse Taylor O. Foster Sung-Kun Kim http://www.mdpi.com/1424-8247/4/2/382 The review summarizes research into the highly relevant topics of cholinesterase and amyloid aggregation inhibitors connected to tacrine congeners, both of which are associated with neurogenerative diseases. Various opinions will be discussed regarding the dual binding site inhibitors which are characterized by increased inhibitor potency against acetylcholin/butyrylcholine esterase and amyloid formation. It is suggested that these compounds can both raise levels of acetylcholine by binding to the active site, and also prevent amyloid aggregation. In connection with this problem, the mono/dual binding of the multifunctional derivatives of tacrine, their mode of action and their neuroprotective activities are reported. The influence of low molecular compounds on protein amyloid aggregation, which might be considered as a potential therapeutic strategy in the treatment of Alzheimer’s disease is also reported. Finally, attention is paid to some physico-chemical factors, such as desolvation energies describing the transfer of the substrate solvated by water, the metal-chelating properties of biometals reacting with amyloid precursor protein, amyloid beta peptide and tau protein. http://www.mdpi.com/1424-8247/4/2/382 Fri, 18 Feb 2011 00:00:00 CET Pharmaceuticals 2011-02-18 4 2 Review 382 418 1424-8247 Neuroactive Multifunctional Tacrine Congeners with Cholinesterase, Anti-Amyloid Aggregation and Neuroprotective Properties 2011-02-18 doi: 10.3390/ph4020382 Maria Kozurkova Slavka Hamulakova Zuzana Gazova Helena Paulikova Pavol Kristian http://www.mdpi.com/1424-8247/4/2/366 Self-injurious behavior (SIB) is a primary reason that individuals with neurodevelopmental disabilities (NDD) are either retained in restrictive environments or are administered psychotropic medication. There are no known causes and no universally accepted treatments for this complex behavior among individuals with NDD. There is developing evidence, however, that individuals exhibiting SIB have a disturbance of the opiate-mediated pain and pleasure system. One hypothesis is that SIB reflects insensitivity to pain and general sensory depression (hypoalgesia), perhaps related to chronic elevation of endogenous opiates. For instance, many self-injurious individuals do not exhibit the usual signs of pain after their “injurious” behavior. Moreover, for some individuals the addictive properties of elevated endogenous opiates (euphoria) may be responsible for maintaining their SIB. In this perspective, SIB may be viewed as an addiction because it supplies the "fix" for tolerant, down-regulated opiate receptors. Reports that levels of endogenous opiates at rest and after SIB episodes predict positive responses to opiate blockers (e.g., naltrexone) provide further support for opiate-mediated SIB and form the basis for a rational treatment strategy. Although the long term effects of opiate blockers on SIB are unknown, reduction in SIB following acute treatment provides support that a specific biological system may be dysregulated in a subgroup of patients. It is concluded that naltrexone produces a clinically significant reduction in the serious and life-threatening behavior of self injury for individuals who have not been responsive to any other type of treatment. Several suggestions and cautions are provided for regimens of naltrexone treatment of SIB. http://www.mdpi.com/1424-8247/4/2/366 Fri, 28 Jan 2011 00:00:00 CET Pharmaceuticals 2011-01-28 4 2 Review 366 381 1424-8247 Opioid Antagonists May Reverse Endogenous Opiate “Dependence” in the Treatment of Self-Injurious Behavior 2011-01-28 doi: 10.3390/ph4020366 Curt A. Sandman Aaron S. Kemp