Phage N15-Based Vectors for Gene Cloning and Expression in Bacteria and Mammalian Cells.

Bacteriophage N15 is the first virus known to deliver linear prophage into During its lysogenic cycle, N15 protelomerase (TelN) resolves its telomerase occupancy site () into hairpin telomeres. This protects the N15 prophage from bacterial exonuclease degradation, enabling it to stably replicate as a linear plasmid in . Interestingly, purely proteinaceous TelN can retain phage DNA linearization and hairpin formation without involving host- or phage-derived intermediates or cofactors in the heterologous environment.

Large BACs transfect more efficiently in circular topology.

The effect of DNA topology on transfection efficiency of mammalian cells has been widely tested on plasmids smaller than 10 kb, but little is known for larger DNA vectors carrying intact genomic DNA containing introns, exons, and regulatory regions. Here, we demonstrate that circular BACs transfect more efficiently than covalently closed linear BACs. We found up to 3.

Utility of Animal Models to Understand Human Alzheimer's Disease, Using the Mastermind Research Approach to Avoid Unnecessary Further Sacrifices of Animals.

To diagnose and treat early-stage (preclinical) Alzheimer's disease (AD) patients, we need body-fluid-based biomarkers that reflect the processes that occur in this stage, but current knowledge on associated processes is lacking. As human studies on (possible) onset and early-stage AD would be extremely expensive and time-consuming, we investigate the potential value of animal AD models to help to fill this knowledge gap. We provide a comprehensive overview of processes associated with AD pathogenesis and biomarkers, current knowledge on AD-related biomarkers derived from on human and animal brains and body fluids, comparisons of biomarkers obtained in human AD and frequently used animal AD models, and emerging body-fluid-based biomarkers.

A Self-Replicating Linear DNA.

TelN and are a unique DNA linearization unit isolated from bacteriophage N15. While being transferable, the TelN cleaving-rejoining activities remained stable to function on in both bacterial and mammalian environments. However, TelN contribution in linear plasmid replication in mammalian cells remains unknown.

Physiologically Based Modeling Approach to Predict Dopamine D2 Receptor Occupancy of Antipsychotics in Brain: Translation From Rat to Human.

Receptor occupancy (RO) is a translational biomarker for assessing drug efficacy and safety. We aimed to apply a physiologically based pharmacokinetic (PBPK) modeling approach to predict the brain dopamine D2 RO time profiles of antipsychotics. Clozapine and risperidone were modeled together with their active metabolites, norclozapine and paliperidone, First, in PK-Sim a rat PBPK model was developed and optimized using literature plasma PK data.

Intranasal delivery of a novel acetylcholinesterase inhibitor HLS-3 for treatment of Alzheimer's disease.

Aim: The present study aims to investigate the pharmacokinetics and pharmacodynamics of HLS-3, a tacrine dimer with high anti-acetylcholinesterase activity for the treatment of Alzheimer's disease.

Main Methods: In vitro Calu-3 and Caco-2 cell monolayer transport and liver microsomal incubation studies of HLS-3 were carried out to evaluate its nasal epithelium and intestinal membrane permeability, transporters involved in absorption and hepatic metabolism. In vivo pharmacokinetics of HLS-3 followed by central and peripheral cholinergic mediated responses and ex vivo AChE activities in rats via oral and intranasal administrations were further investigated and compared.

Prediction of human CNS pharmacokinetics using a physiologically-based pharmacokinetic modeling approach.

Knowledge of drug concentration-time profiles at the central nervous system (CNS) target-site is critically important for rational development of CNS targeted drugs. Our aim was to translate a recently published comprehensive CNS physiologically-based pharmacokinetic (PBPK) model from rat to human, and to predict drug concentration-time profiles in multiple CNS compartments on available human data of four drugs (acetaminophen, oxycodone, morphine and phenytoin). Values of the system-specific parameters in the rat CNS PBPK model were replaced by corresponding human values.

Development of a population pharmacokinetic model to predict brain distribution and dopamine D2 receptor occupancy of raclopride in non-anesthetized rat.

Background: Raclopride is a selective antagonist of the dopamine D2 receptor. It is one of the most frequently used in vivo D2 tracers (at low doses) for assessing drug-induced receptor occupancy (RO) in animals and humans. It is also commonly used as a pharmacological blocker (at high doses) to occupy the available D2 receptors and antagonize the action of dopamine or drugs on D2 in preclinical studies.

Novel CNS drug discovery and development approach: model-based integration to predict neuro-pharmacokinetics and pharmacodynamics.

CNS drug development has been hampered by inadequate consideration of CNS pharmacokinetic (PK), pharmacodynamics (PD) and disease complexity (reductionist approach). Improvement is required via integrative model-based approaches. Areas covered: The authors summarize factors that have played a role in the high attrition rate of CNS compounds.

Predicting Drug Concentration-Time Profiles in Multiple CNS Compartments Using a Comprehensive Physiologically-Based Pharmacokinetic Model.

Drug development targeting the central nervous system (CNS) is challenging due to poor predictability of drug concentrations in various CNS compartments. We developed a generic physiologically based pharmacokinetic (PBPK) model for prediction of drug concentrations in physiologically relevant CNS compartments. System-specific and drug-specific model parameters were derived from literature and in silico predictions.

Brain Uptake of Bioactive Flavones in Scutellariae Radix and Its Relationship to Anxiolytic Effect in Mice.

Scutellariae Radix (SR) and its bioactive flavones elicit a variety of effects in the brain. However, the brain uptake of individual SR flavones and its relationship to the elicited effects after SR administration remain unknown. Moreover, previous studies seldom measured pharmacokinetic and pharmacodynamic outcomes simultaneously.

Kinetics for Drug Discovery: an industry-driven effort to target drug residence time.

A considerable number of approved drugs show non-equilibrium binding characteristics, emphasizing the potential role of drug residence times for in vivo efficacy. Therefore, a detailed understanding of the kinetics of association and dissociation of a target-ligand complex might provide crucial insight into the molecular mechanism-of-action of a compound. This deeper understanding will help to improve decision making in drug discovery, thus leading to a better selection of interesting compounds to be profiled further.

A Generic Multi-Compartmental CNS Distribution Model Structure for 9 Drugs Allows Prediction of Human Brain Target Site Concentrations.

Purpose: Predicting target site drug concentration in the brain is of key importance for the successful development of drugs acting on the central nervous system. We propose a generic mathematical model to describe the pharmacokinetics in brain compartments, and apply this model to predict human brain disposition.

Methods: A mathematical model consisting of several physiological brain compartments in the rat was developed using rich concentration-time profiles from nine structurally diverse drugs in plasma, brain extracellular fluid, and two cerebrospinal fluid compartments.

Revealing the Neuroendocrine Response After Remoxipride Treatment Using Multi-Biomarker Discovery and Quantifying It by PK/PD Modeling.

To reveal unknown and potentially important mechanisms of drug action, multi-biomarker discovery approaches are increasingly used. Time-course relationships between drug action and multi-biomarker profiles, however, are typically missing, while such relationships will provide increased insight in the underlying body processes. The aim of this study was to investigate the effect of the dopamine D2 antagonist remoxipride on the neuroendocrine system.

Mechanistic models enable the rational use of in vitro drug-target binding kinetics for better drug effects in patients.

Introduction: Drug-target binding kinetics are major determinants of the time course of drug action for several drugs, as clearly described for the irreversible binders omeprazole and aspirin. This supports the increasing interest to incorporate newly developed high-throughput assays for drug-target binding kinetics in drug discovery. A meaningful application of in vitro drug-target binding kinetics in drug discovery requires insight into the relation between in vivo drug effect and in vitro measured drug-target binding kinetics.

Identification and disposition of novel mono-hydroxyl mefenamic acid and their potentially toxic 1-O-acyl-glucuronides in vivo.

Mefenamic acid (MEF) is a widely prescribed non-steroidal anti-inflammatory drug that has been found associated with rare but severe cases of hepatotoxicity, nephrotoxicity and gastrointestinal toxicity. The formation of protein-reactive acylating metabolites such as 1-O-acyl-MEF glucuronide (MEFG) and 3'-hydroxymethyl-MEF 1-O-acyl-glucuronide is one proposed cause. In addition to the well-reported 3'-hydroxymethyl-MEF, two mono-hydroxyl-MEF (OH-MEFs) were recently identified in vitro.

Herb-drug interactions between Scutellariae Radix and mefenamic acid: Simultaneous investigation of pharmacokinetics, anti-inflammatory effect and gastric damage in rats.

Ethnopharmacological Relevance: Scutellariae Radix (SR), the dried root of Scutellariae baicalensis Georgi, has a lot in common with non-steroidal anti-inflammatory drugs (NSAIDs). Their similarities in therapeutic action (anti-inflammation) and metabolic pathways (phase II metabolisms) may lead to co-administration by patients with the potential of pharmacokinetic and/or pharmacodynamic interactions. The current study aims to investigate the potential interactions between SR and an NSAID, mefenamic acid (MEF), on the overall pharmacokinetic dispositions, anti-inflammatory effects and adverse effects in rats.

Development of a SPE-LC/MS/MS method for simultaneous quantification of baicalein, wogonin, oroxylin A and their glucuronides baicalin, wogonoside and oroxyloside in rats and its application to brain uptake and plasma pharmacokinetic studies.

This study aims to identify and quantify the six major bioactive flavones of the traditional Chinese medicine Scutellariae Radix (RS), including baicalein, baicalin, wogonin, wogonoside, oroxylin A and oroxyloside in rat after oral administration of a standardized RS extract. A novel, sensitive and selective method for simultaneous determination of these six analytes in rat brain and plasma using solid phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC/MS/MS) was developed and fully validated. The lower limits of quantification (LLOQs) for the six RS flavones in brain tissue were 0.

Development, characterization and application of in situ gel systems for intranasal delivery of tacrine.

The present study aimed to develop an in situ gel formulation for intranasal delivery of tacrine (THA), an anti-Alzheimer's drug. Thermosensitive polymer Pluronic F-127 was used to prepare THA in situ gels. Sol-gel transition temperature (Tsol-gel), rheological properties, in vitro release, and in vivo nasal mucociliary transport time were optimized.

Pharmacokinetic comparison between the long-term anesthetized, short-term anesthetized and conscious rat models in nasal drug delivery.

Purpose: To investigate the pharmacokinetic differences between the common nasal delivery models.

Methods: In three different rat models [long-term anesthetized (with nasal surgery), short-term anesthetized (without nasal surgery) and conscious models], tacrine and loxapine were administered via nasal, intravenous and oral routes, and the plasma pharmacokinetics were compared among different models.

Results: Systemic exposures of both drugs and their metabolites were consistently higher in long-term anesthetized model after all routes of administration in comparison to that of conscious model.

Alterations in the CNS effects of anti-epileptic drugs by Chinese herbal medicines.

Introduction: Concomitant use of anti-epileptic drugs (AEDs) and Chinese herbal medicines (CHMs) is increasing globally. However, information summarizing how CHMs might alter the CNS effects of AEDs is lacking.

Areas Covered: A systematic review of the English-language articles in evidence-based databases was performed.

Brain disposition and catalepsy after intranasal delivery of loxapine: role of metabolism in PK/PD of intranasal CNS drugs.

Purpose: To elucidate the role of metabolism in the pharmacokinetics and pharmacodynamics of intranasal loxapine in conscious animals.

Methods: At pre-determined time points after intranasal or oral loxapine administration, levels of loxapine, loxapine metabolites, and neurotransmitters in rat brain were quantified after catalepsy assessments (block test and paw test). Cataleptogenicity of loxapine was also compared with its metabolites.

Regioselective biotransformation of CNS drugs and its clinical impact on adverse drug reactions.

Introduction: Adverse drug reactions (ADRs) continue to be one of the major causes of failure in drug development while limiting the clinical utilities of many drugs. Contribution of the metabolites formed in vivo to ADRs could be more significant than we might have expected.

Areas Covered: This review focuses on the relationship between regioselectivity in biotransformation and the ADRs of drugs acting on the central nervous system (CNS).