Molecular modeling of blood-brain barrier nutrient transporters: in silico basis for evaluation of potential drug delivery to the central nervous system

For drugs that act in the brain, the blood-brain barrier (BBB) is a considerable physical barrier which influences the distribution of drugs to the brain. The BBB is essentially impermeable for hydrophilic and/or charged compounds. Nutrient membrane transporters have an important physiological role in the transport of essential substances across the BBB required for normal brain function. We and others have shown that these transporters may have utility as drug delivery vectors, thereby increasing brain distribution of these compounds via these systems. In this review, we evaluate molecular (in silico) models of BBB transport proteins. Few BBB membrane transporters have been crystallized, but their crystal structures have a possibility for use in homology modeling. Other techniques commonly used are 2D quantitative structure-activity relationships (QSAR), as well as 3D-QSAR techniques including comparative molecular field analysis (CoMFA) and comparative similarity index analysis (CoMSIA). Each of these models provides valuable information for ascertaining their potential basis for BBB transport and brain drug delivery.     

Shedding light on the puzzle of drug-membrane interactions: Experimental techniques and molecular dynamics simulations

Lipid membranes work as barriers, which leads to inevitable drug-membrane interactions in vivo. These interactions affect the pharmacokinetic properties of drugs, such as their diffusion, transport, distribution, and accumulation inside the membrane. Furthermore, these interactions also affect their pharmacodynamic properties with respect to both therapeutic and toxic effects. Experimental membrane models have been used to perform in vitro assessment of the effects of drugs on the biophysical properties of membranes by employing different experimental techniques. In in silico studies, molecular dynamics simulations have been used to provide new insights at an atomistic level, which enables the study of properties that are difficult or even impossible to measure experimentally. Each model and technique has its advantages and disadvantages. Hence, combining different models and techniques is necessary for a more reliable study. In this review, the theoretical backgrounds of these (in vitro and in silico) approaches are presented, followed by a discussion of the pharmacokinetic and pharmacodynamic properties of drugs that are related to their interactions with membranes. All approaches are discussed in parallel to present for a better connection between experimental and simulation studies. Finally, an overview of the molecular dynamics simulation studies used for drug-membrane interactions is provided.     

Human and animal hepatocytes in vitro with extrapolation in vivo

Human and animal hepatocytes are now being used as an in vitro technique to aid drug discovery by predicting the in vivo metabolic pathways of drugs or new chemical entities (NCEs), identifying drug-metabolizing enzymes and predicting their in vivo induction. Because of the difficulty of establishing whether the cytotoxic susceptibility of human hepatocytes to xenobiotics/drugs in vitro could be used to predict in vivo human hepatotoxicity, a comparison of the susceptibility of the hepatocytes of human and animal models to six chemical classes of drugs/xenobiotics in vitro have been related to their in vivo hepatotoxicity and the corresponding activity of their metabolizing enzymes. This study showed that the cytotoxic effectiveness of 16 halobenzenes towards rat hepatocytes in vitro using higher doses and short incubation times correlated well with rat hepatotoxic effectiveness in vivo with lower doses/longer times. The hepatic/hepatocyte xenobiotic metabolizing enzyme activities of various animal species and human have been reviewed for use by veterinarians and research scientists. Where possible, recommendations have been made regarding which animal hepatocyte model is most applicable for modeling the susceptibility to xenobiotic induced hepatotoxicity of those humans with slow versus rapid metabolizing enzyme polymorphisms. These recommendations are based on the best human fit for animal drug/xenobiotic metabolizing enzymes in terms of activity, kinetics and substrate/inhibitor specificity. The use of human hepatocytes from slow versus rapid metabolizing individuals for drug metabolism/cytotoxicity studies; and the research use of freshly isolated rat hepatocytes and "Accelerated Cytotoxicity Mechanism Screening" (ACMS) techniques for identifying drug/xenobiotic reactive metabolites are also described. Using these techniques the molecular hepatocytotoxic mechanisms found in vitro for seven classes of xenobiotics/drugs were found to be similar to the rat hepatotoxic mechanisms reported in vivo.     

心肺复苏后脑缺血再灌注损伤治疗理念及相关动物模型研究进展

心肺复苏后脑缺血再灌注损伤是一个复杂的病理生理变化过程,由多种损伤机制共同参与。自心肺复苏后系统性综合治疗和亚低温治疗在临床上广泛应用后,目前已有多种治疗理念在不同的动物实验和动物模型基础上被提出,包括缺血预处理、药物预处理、缺血后处理、和药物后处理,而后吸入麻醉药对心肺复苏后脑缺血再灌注损伤的保护作用受到了人们的重视,而七氟烷后处理已经成为目前研究的热点之一。为了指导临床上的心肺复苏,人们一直在利用不同动物模型,探究不同保护方法,寻找有效的脑保护药物。而各种治疗理念的提出均是建立在动物实验和动物模型的基础上,窒息性心肺复苏模型模拟围术期气道梗阻,能较贴切的复制临床上由窒息引起的心肺复苏后脑损伤,对将来指导临床复苏具有重大意义。     

A new paradigm in respiratory hygiene: modulating respiratory secretions to contain cough bioaerosol without affecting mucus clearance

Background

Several strategies and devices have been designed to protect health care providers from acquiring transmissible respiratory diseases while providing care. In modulating the physical characteristics of the respiratory secretions to minimize the aerosolization that facilitates transmission of airborne diseases, a fundamental premise is that the prototype drugs have no adverse effect on the first line of respiratory defense, clearance of mucus by ciliary action.

Methods

To assess and demonstrate the primary mechanism of our mucomodulators (XLs), we have built our evidence moving from basic laboratory studies to an ex-vivo model and then to an in-vivo large animal model. We exposed anesthetized dogs without hypersecretion to different dose concentrations of aerosolized XL "B", XL "D" and XL "S". We assessed: cardio-respiratory pattern, tracheal mucus clearance, airway patency, and mucus viscoelastic changes.

Results

Exposure of frog palate mucus to XLs did not affect the clearance of mucus by ciliary action. Dogs maintained normal cardio-respiratory pattern with XL administration. Tracheal mucociliary clearance in anesthetized dogs indicated a sustained 40% mean increase. Tracheal mucus showed increased filance, and there was no mucus retention in the airways.

Conclusion

The ex-vivo frog palate and the in-vivo mammalian models used in this study, appear to be appropriate and complement each other to better assess the effects that our mucomodulators exert on the mucociliary clearance defence mechanism. The physiological function of the mucociliary apparatus was not negatively affected in any of the two epithelial models. Airway mucus crosslinked by mucomodulators is better cleared from an intact airway and normally functioning respiratory system, either due to enhanced interaction with cilia or airflow-dependent mechanisms. Data obtained in this study allow us to assure that we have complied with the fundamental requirement criteria established in the initial phase of developing the concept of mucomodulation: Can we modulate the physical characteristics of the respiratory secretions to reduce aerosolization without affecting normal mucociliary clearance function, or even better improving it?     

Kidney disease models:tools to identify mechanisms and potential therapeutic targets

Acute kidney injury(AKI) and chronic kidney disease(CKD) are worldwide public health problems affecting millions of people and have rapidly increased in prevalence in recent years. Due to the multiple causes of renal failure, many animal models have been developed to advance our understanding of human nephropathy. Among these experimental models, rodents have been extensively used to enable mechanistic understanding of kidney disease induction and progression, as well as to identify potential targets for therapy. In this review, we discuss AKI models induced by surgical operation and drugs or toxins, as well as a variety of CKD models(mainly genetically modified mouse models).Results from recent and ongoing clinical trials and conceptual advances derived from animal models are also explored.     

Lymphatic transport of orally administered drugs

Several therapeutic molecules such as lipophilic drugs and peptides suffer from the problems of low oral bioavailability. Improvement of their bioavailability and simultaneous prevention of the oral degradation of the prone molecules appears to be a challenge. Lymphatic system, which is responsible for the maintenance of fluid balance, immunity and metastatic spread of cancers, is also found to play a major role in the oral absorption of lipids and lipophilic drugs from intestine. The specialized structure of gut associated lymphoid tissue can be utilized as a gateway for the delivery of particulate systems containing drugs. Even though a large gap has existed in the field of lymphatic drug delivery, the introduction of a large number of lipophilic drugs and peptides has brought a renewed interest of research in this area. In this review, the mechanisms of intestinal lymphatic drug transport, approaches taken for the delivery of macromolecules, lipophilic and peptide drugs, biochemical barriers involved in intestinal drug absorption, and animal models used in the studies of intestinal lymphatic drug transport has been discussed.     

Formulating gels for decreased mucociliary transport using rheologic properties: Polyacrylic acids

The purpose of these studies was to identify the rheologic properties of polyacrylic acid gels necessary for optimal reductions in mucociliary clearance. The mucociliary transport of 2 bioadhesive polyacrylic acid polymers, polycarbophil and carbopol, was assessed in vitro by measuring their clerance rates across explants of ciliated bovine tracheal tissue. The viscoelastic properties of polymer gels were measured in the presence of mucus using controlled stress rheometry. Combinations of apparent viscosity (η) and complex modulus (G^*) were found to be the most useful parameters in the identification of polyacrylic acid formulations capable of decreasing mucociliary transport rate (MTR). A narrow range of η and G^* values suitable for reducing mucociliary clearance, while remaining sufficiently fluid for intranasal administration, were identified. The correlations between the rheologic parameters of the polycarbophil gels and their mucociliary transport rates were used to identify other polyacrylic acid gels that also had suitable mucociliary clearance properties, demonstrating that these parameters can be used to direct the optimization of formulations using simple in vitro rheologic testing. Published: April 20, 2007     

Microbial biotransformation: a tool for drug designing

For centuries microbial biotransformation has proved to be an imperative tool in alleviating the production of various chemicals used in food, pharmaceutical, agrochemical and other industries. In the field of pharmaceutical research and development, biotransformation studies have been extensively applied to investigate the metabolism of compounds (leads, lead candidates, etc.) using animal models. The microbial biotransformation phenomenon is then commonly employed in comparing metabolic pathways of drugs and scaling up the metabolites of interest discovered in these animal models for further pharmacological and toxicological evaluation. Microorganisms can conveniently afford drugs difficult obtained via synthesis. The plethora of reported microbial biotransformations along with its added benefits has already invoked further research in bioconversion of novel and structurally complex drugs. This review alternatively discusses the prospect of microbial biotransformation studies as a significant element ameliorating drug discovery and design in terms of cost-effectiveness, environment protection and greater structural diversity as compared to animal models used to study metabolism. To explicate the microbial biotransformation paradigm in drug designing 3 main areas in this aspect have been analyzed: 1—lead expansion: obtaining pharmacologically improved metabolites from bioactive molecules; 2—biosynthesis of precursors/intermediates involved in the production of bioactive molecules; 3—resolution of racemic mixture to obtain enantiomers possessing different pharmacological profiles.     

Mouse models for breast cancer metastasis

Metastasis of cancer cells is the main cause of death in most breast cancer patients. Although markers for early diagnosis and drugs that limit the spread of cancer to other organs have been developed, it is difficult to prevent the relapse of breast cancer. Recent research has highlighted the importance of tumor environment in which communication between tumor cells and the body system occurs. Emerging data have suggested that animal models are a good system to investigate this communication. Therefore, studies with mouse models have been developed as a reasonable method for a systemic approach to understand breast cancer metastasis. In this review, we summarize mouse models of breast cancer and their applications to the study of human breast cancers, and discuss limitation of model system and advanced techniques to overcome it.     

Mucus and the mare: how little we know

Uterine infections are a major cause of infertility, but the role of mucus in equine uterine defense is not well understood. Mucociliary currents play an important role in protecting mucous membranes, including the upper and lower respiratory tracts of mammals, and are required for feeding and oxygenation of many aquatic invertebrates. Although phagocytosis has long been considered the first line of uterine defense in the mare, there are concerns about its efficacy in the uterine lumen. Additional local defenses, such as mucociliary currents, have therefore been proposed. The uterine epithelium exhibits alternating mucus-secreting and ciliated cells supporting a mucopolysaccharide blanket, features shared with mucociliary membranes throughout the animal kingdom. Gross uterine anatomy, such as continuity of uterine and cervical folds, may indicate adaptations to mucociliary clearance. In addition, ciliated cells obtained in uterine lavages often display motility. Disruptions of mucociliary clearance play major roles in pathogenesis of mucosal infections in humans, including pneumonia, chronic sinusitis, and otitis media. Establishing drainage is a major goal of therapy in treatment of chronic sinusitis, hastening return of mucociliary function. Similar disruptions may occur in equine uterine infections, associated with accumulations of uterine fluid, loss of endometrial folds, and cervical trauma. Possible clinical implications of mucociliary clearance in the mare are discussed, however the role of mucociliary clearance in the mare remains speculative.     

痔疮动物模型的研究进展

随着对痔疮新药药效评价的需求,痔疮模型的建立有了初步的发展。目前,大鼠、小鼠、家兔、卷尾猴等动物已被成功的用于痔疮模型的建立。所用方法主要有:巴豆油法、醋酸法、感染法、创伤法、静脉阻断法等,动物模型的成功建立和合理应用将有利于推动痔疮新药的研发,本文将现有痔疮模型创建的原理和方法进行了归纳、总结。     

Animal models of enterovirus 71 infection: applications and limitations

Human enterovirus 71 (EV71) has emerged as a neuroinvasive virus that is responsible for several outbreaks in the Asia-Pacific region over the past 15 years. Appropriate animal models are needed to understand EV71 neuropathogenesis better and to facilitate the development of effective vaccines and drugs. Non-human primate models have been used to characterize and evaluate the neurovirulence of EV71 after the early outbreaks in late 1990s. However, these models were not suitable for assessing the neurovirulence level of the virus and were associated with ethical and economic difficulties in terms of broad application. Several strategies have been applied to develop mouse models of EV71 infection, including strategies that employ virus adaption and immunodeficient hosts. Although these mouse models do not closely mimic human disease, they have been applied to determine the pathogenesis of and treatment and prevention of the disease. EV71 receptor-transgenic mouse models have recently been developed and have significantly advanced our understanding of the biological features of the virus and the host-parasite interactions. Overall, each of these models has advantages and disadvantages, and these models are differentially suited for studies of EV71 pathogenesis and/or the pre-clinical testing of antiviral drugs and vaccines. In this paper, we review the characteristics, applications and limitation of these EV71 animal models, including non-human primate and mouse models.     

痔疮动物模型制作方法研究进展

近年来,在兔、大鼠、小鼠和犬等动物种属成功地建立了巴豆油、醋酸、感染与创伤诱导的痔疮动物模型,每种动物模型各有优缺点,应用合理的动物模型能更好地开展痔的实验和临床研究。随着治痔药物的深入研究,动物模型有很好发展和应用,本文综述了痔疮动物模型制作方法研究进展。     

Extrapolating Antifungal Animal Data to Humans—Is It Reliable?

This article aimed to review animal models of antifungals and identifies human literature to assess if the extrapolation of results is reliable. Animal studies have helped identify area under the concentration curve to minimum inhibitory concentration ratio targets for new drugs and formulations such as isavuconazole and delayed-release posaconazole that have translated to successful outcomes in humans. Models have also been influential in the identification of possible combination therapies for the treatment of aspergillosis, such as voriconazole and echinocandins. However, challenges are endured with animal models when it comes to replicating the pharmacokinetics of humans which has been exemplified with the newest itraconazole formulation. Additionally, animal models have displayed a survival benefit with the use of iron chelators and amphotericin for mucormycosis which was not demonstrated in humans. Animal models have been a staple in the development and optimization of antifungal agents. They afford the ability to investigate uncommon diseases, such as invasive fungal infections, that would otherwise take years and many resources to complete. Although there are many benefits of animal models, there are also shortcomings. This is why the reliability of extrapolating data from animal models to humans is often scrutinized.     

And then there were acyl coenzyme A:cholesterol acyl transferase inhibitors

PURPOSE OF REVIEW: The reputation of acyl coenzyme A:cholesterol acyltransferase (ACAT) inhibitors has changed profoundly from promising new drugs for cardiovascular prevention to drugs without clinical benefits or possibly even with adverse effects. RECENT FINDINGS: ACAT inhibitors decrease the intracellular conversion of free cholesterol into cholesteryl ester in a number of tissues, including intestine, liver and macrophages. In contrast to promising results in experimental animal models, all subsequent clinical studies in humans with ACAT inhibitors failed to show lipid profile changes as well as reductions in surrogate markers for coronary artery disease. In fact, there was even a tendency towards an increase in atheroma burden in the most recent and well executed clinical trials. In addition, the inhibition of this pivotal enzyme in cholesterol esterification may interfere with reverse cholesterol transport. SUMMARY: In our opinion, the consistent negative findings in recent clinical trials have virtually eliminated the chances for this class of drugs to be introduced for cardiovascular prevention. Possible strategies focused on selective ACAT 2 inhibition or the combination of ACAT inhibitors with compounds that stimulate reverse cholesterol transport may prove to have clinical benefit. This will have to await further clinical research in humans, however, as, obviously, rodent models cannot provide reliable data as to the efficacy of this class of drugs in humans.     

Liquid secretion inhibitors reduce mucociliary transport in glandular airways

Because of its possible importance in cystic fibrosis (CF) pulmonary pathogenesis, the effect of anion and liquid secretion inhibitors on airway mucociliary transport was examined. When excised porcine tracheas were treated with ACh to induce gland liquid secretion, the rate of mucociliary transport was increased nearly threefold from 2.5 +/- 0.5 to 6.8 +/- 0.8 mm/min. Pretreatment with both bumetanide and dimethylamiloride (DMA), to respectively inhibit Cl(-) and HCO secretion, significantly reduced mucociliary transport in the presence of ACh by 92%. Pretreatment with the anion channel blocker 5-nitro-2-(3-phenylpropylamino)benzoic acid similarly reduced mucociliary transport in ACh-treated airways by 97%. These agents did not, however, reduce ciliary beat frequency. Luminal application of benzamil to block liquid absorption significantly attenuated the inhibitory effects of bumetanide and DMA on mucociliary transport. We conclude that anion and liquid secretion is essential for normal mucociliary transport in glandular airways. Because the CF transmembrane conductance regulator protein likely mediates Cl(-), HCO, and liquid secretion in normal glands, we speculate that impairment of gland liquid secretion significantly contributes to defective mucociliary transport in CF.     

药物跨膜转运细胞模型影响因素及应用

本文详细介绍了Caco-2细胞系和MDCK细胞系的特点、跨膜转运细胞模型的建立及其影响因素,包括细胞模型的选择、细胞接种密度、细胞单层的紧密性等细胞因素和Transwell多微孔膜的性质等环境因素。概述了国内外关于利用Caco-2和MDCK细胞系作为模型进行药物筛选、药物相互作用和研究药物吸收转运机制等方面的内容及MDCK细胞模型作为肠道模型、肾脏模型及血脑屏障模型的应用。比较了Caco-2细胞和MDCK细胞在肠道模型方面的差别,MDCK细胞主要用于选择性研究药物在小肠吸收及转运机制,特别用于细胞旁被动转运药物的研究,而Caco-2细胞用于双向转运或能量依赖主动转运研究。MDCK细胞模型可在体外培养条件下平稳转染人类MDR1基因,因此可高表达P-gp基因,可作为可用于评估肾脏药物相互作用、快速进行候选药物筛选及研究药物转运机制的理想模型。     

Osteocalcina: nexo de unión entre homeostasis ósea y metabolismo energético

Research in animal models has demonstrated the role of osteocalcin, a bone formation marker, in regulation of energy metabolism. Those studies have led to a new concept of the bone acting as an endocrine organ by secreting osteocalcin, which acts by increasing insulin secretion, lowering plasma glucose, and increasing insulin sensitivity and energy expenditure. Results in humans have been conflicting. On the other hand, antiresorptive drugs used against osteoporosis decrease osteocalcin levels, while anabolic drugs increase osteocalcin levels. However, the effects of these therapies on energy metabolism have not been investigated.     

Percutaneous permeation of enantiomers and racemates of chiral drugs and prediction of their flux ratios using thermal data: a pharmaceutical perspective

Albeit pharmacological, pharmacokinetic, and toxicological differences between enantiomeric pairs or between the pure enantiomers and racemate of chiral drugs are known to exist for decades, we are just beginning to realize that there are apparent differences between these species with respect to their percutaneous permeation as well. Such differences in permeation are likely to be enhanced when chiral drugs are formulated with chiral excipients, necessitating a careful assessment of the effect of formulation excipients on the permeation as well as the overall therapeutic outcomes. The in vitro transport data from the preclinical investigations, using laboratory animal models and/or in vitro cell culture systems, must be carefully validated in vivo as there are differences between these models and the human skin. Mathematical models such as MTMT that utilize the interdependence of certain physicochemical characteristics and percutaneous permeability have a predictive value in assessing the flux behavior of enantiomers and racemates.