Role of CNS efflux drug transporters in antiepileptic drug delivery: overcoming CNS efflux drug transport

Experimental support for the transporter hypothesis of drug resistance in epilepsies has triggered efforts developing and validating approaches to overcome enhanced blood-brain barrier efflux transport. Testing in rodent models has rendered proof-of-concept for an add-on therapy with antiepileptic drugs. However, further development of the approach would require tolerability considerations as efflux transporters serve an important protective function throughout the body limiting distribution of harmful xenobiotics. Relevant progress has been made in the elucidation of mechanisms driving up-regulation of the multidrug transporter P-glycoprotein in response to seizure activity. Based on this knowledge, novel strategies have been evaluated targeting the signaling cascade that regulates P-glycoprotein in the epileptic brain. Further concepts might include by-passing blood-brain barrier transporters by intracerebral administration or by encapsulation of antiepileptic drugs in nano-sized carrier systems. It is important to note that the future perspectives of respective approaches are still questionable based on the limited evidence for a clinical relevance of transporter expression. Thus, techniques are urgently needed for non-invasive assessment of blood-brain barrier transporter function. Respective techniques would allow testing for a clinical correlation between pharmacosensitivity and transporter function, validating therapeutic strategies targeting efflux transporters and selecting patients with transporter over-expression for respective clinical trials. Provided that further clinical data render support for the transporter hypothesis, the main question remains whether patients exist in which transporter over-expression is the predominant mechanism of drug resistance and in which overcoming drug efflux is equivalent with overcoming drug resistance. Imaging techniques might provide a tool to address these questions in clinical epileptology. However, the complex pharmacological interactions between antiepileptic drugs, radiotracers, and transporter modulators used in these approaches as well as interindividual differences in the brain pathology might hamper clear-cut conclusions and limit the diagnostic significance.     

Drug efflux transporters in the CNS

The central nervous system (CNS) contains important cellular barriers that maintain homeostasis by protecting the brain from circulating toxins and through the elimination of toxic metabolites generated in the brain. The barriers that limit the concentration of toxins and xenobiotics in the interstitial fluids of the CNS are the capillary endothelial cells of the blood-brain barrier (BBB) and the epithelial cells of the blood-cerebrospinal fluid barrier (BCSFB). Both of these barriers have cellular tight junctions and express transport systems which serve to actively transport nutrients into the brain, and actively efflux toxic metabolites and xenobiotics out of the brain. This review will focus on the expression and function of selected drug efflux transporters in these two barriers, specifically the multidrug resistance transporter, p-glycoprotein, and various organic anion transporters, such as multidrug resistance-associated proteins, organic anion transporter polypeptides, and organic anion transporters. These transport systems are increasingly recognized as important determinants of drug distribution to, and elimination from, different compartments of the CNS. Consequences of drug efflux transporters in barriers of the CNS include limiting the distribution of substrates that are beneficial to treat CNS diseases, and increasing the possibility of drug-drug interactions that may lead to untoward toxicities. Therefore, the study of these transporters is important in examining the various determinants of drug delivery to the CNS.     

Microdialysis in drug discovery

Microdialysis has been developed during the last 25 years by several authors primarily to study brain function and changes in levels of endogenous compounds such as neurotransmitters or metabolites. The development of microdialysis for the purpose of measuring drugs was initiated during the late eighties. This technique provides a means of continuous plasma sampling without repeated blood sampling and the applicability to the study of drug metabolism and pharmacokinetics in experimental animals and human. Also, the microdialysis technique allows the study of plasma protein binding and the saturation of protein binding. The implantation of the microdialysis probe in other tissues and organs, like central nervous system, adipose tissue and heart, allows the study of drug distribution. On the other hand, the measurement of endogenous substances using the microdialysis technique permits the study of the effect of drugs on neurotransmission and metabolism. Moreover, as this technique allows the simultaneous determination of different physiological parameters such as blood pressure, locomotor and convulsive activity, it is a suitable tool for pharmacokinetic-pharmacodynamic studies of drugs and pharmacokinetic-pharmacodynamic (PK-PD) modeling. Lastly, the reverse microdialysis is a powerful technique for the study of local actions of drugs in different tissues such as specific brain nuclei, myocardium, liver or skeletal muscle. So, this article reviewed the vast applications of the microdialysis technique for the study of pharmacokinetic and pharmacodynamic properties of drugs.     

ABC transporters in the CNS - an inventory

In the present review we provide a summary of ATP-binding cassette (ABC) transporters in the central nervous system (CNS). Our review is focused on transporters of the ABC A, B, C, D, and G families that have been detected in the cells of the neurovascular unit/blood-brain barrier including brain capillary endothelial cells, pericytes, astrocytes, and neurons, as well as in other brain cells, such as microglia, oligodendrocytes, and choroid plexus epithelial cells. In this review, we provide an overview, organized by ABC family, of transporter expression, localization, and function. We summarize recent findings on ABC transporter regulation in the CNS and address the role of ABC transporters in CNS diseases including brain cancer, seizures/epilepsy, and Alzheimer's disease. Finally, we discuss new therapeutic strategies focused on ABC transporters in CNS disease.     

Microdialysis in clinical drug delivery studies

The introduction of in vivo microdialysis (MD) to clinical pharmacological studies has opened the opportunity to obtain previously inaccessible information about the drug distribution process to the clinically relevant target site. The aim of this review is to provide a comprehensive overview of the current literature about MD in drug delivery studies from a clinical perspective. In particular the application of MD in clinical--antimicrobial, oncological and transdermal--and neurological research will be described and the scope of MD in pharmacokinetic-pharmacodynamic (PK-PD) studies will be discussed. It is concluded that MD has a great potential for both academic and industrial research, and may become the method of choice for drug distribution studies in humans.     

Microdialysis and drug delivery to the eye

The eye presents unique challenges in both the development of tools for elucidating drug disposition as well as for the development of modes of drug delivery for treatment of ocular diseases. In this paper, we present a discussion of the anatomical and physiological characteristics and limitations present in the eye for microdialysis sampling of endogenous substrates and xenobiotics. To date, over twenty papers describing microdialysis approaches for assessment of ocular drug delivery and endogenous substrate characterization have been published. Although the majority of papers describe sampling of vitreous humor, recent efforts have been directed towards ocular anterior segment sampling using microdialysis. With this approach, an appreciable reduction in animal use has been realized. In addition, simultaneous examination of administered drug and endogenous substrates modulated by the drug is possible with this approach, facilitating construction of ocular pharmacokinetic/pharmacodynamic relationships through use of relevant surrogate markers.     

Microdialysis in mice for drug delivery research

Intracerebral microdialysis was first performed in the mouse at the end of the 1980s. Most microdialysis studies on mice were confined to neuropharmacology and changes in neurotransmitter concentrations up to 1995, although pharmacological studies were done on other tissues like the skin, kidney and implanted tumors. The use of microdialysis in mice for pharmacokinetic and drug delivery studies owes much to the recent availability of genetically engineered mice, such as mice in which the genes encoding multiple drug resistance have been knocked out. The quantitative microdialysis of blood and various tissue fluids of the mouse is now feasible and the recent development of specific microdialysis devices for use in mice should facilitate its use in these small animals. This review covers the technical aspects of microdialysis in the mouse and includes references to many of the published studies on pharmacokinetics and drug delivery.     

Placental drug transporters

Any treatment of a pregnant woman with medication (drugs) de facto results in the treatment of her unborn child, even when her unborn child is not the target of drug therapy. This is because, in most instances, the placenta is not a complete barrier to the passage of drugs from the maternal to the fetal compartment. This barrier is in part due to the presence of various efflux transporters in the placenta. The placenta is also richly endowed with influx transporters. In this article, we will review the physiological characteristics of the placenta and how it functions as a barrier to passage of drugs into the fetal compartment. In addition, we will review placental transporters that are important in modulating the exposure of the fetus to drugs and, therefore, the efficacy and toxicity of such drugs towards the fetus.     

ABC transporters: unvalidated therapeutic targets in cancer and the CNS

The discovery of the multidrug transporter P-glycoprotein (Pgp) over 35 years ago in drug resistant cells prompted several decades of work attempting to overcome drug resistance by inhibition of drug efflux. Despite convincing laboratory data showing that drug transport can be inhibited in vitro, efforts to translate this discovery to the clinic have not succeeded. Since overexpression of Pgp and related transporters including ABCG2 and members of the ABCC family have been linked with poor outcome, it remains a reasonable hypothesis that this poor outcome is linked to reduction of drug exposure by efflux, and thus to drug resistance. In this review, we will discuss the question of whether ABC transporters mediate drug resistance in cancer through a reduction in drug accumulation in tumors, and whether the "Pgp inhibition hypothesis" might be wrong. The hypothesis, which holds that increased chemotherapy effectiveness can be achieved by inhibiting Pgp-mediated drug efflux has only been validated in model systems. Possible explanations for the failure to validate this clinically include the existence of other modulators of drug accumulation and uptake in tumors. Despite these difficulties, a potential role has emerged for drug transporters as therapeutic targets in the central nervous system (CNS). Both lines of investigation point to the need for imaging agents to facilitate the study of drug accumulation in human cancer. This is a critical need for targeted therapies where an important dose-response relationship is likely to exist, and where drug resistance renders many of the novel targeted agents ineffective in a subset of patients.     

Pharmacogenetics of drug transporters in the enterohepatic circulation

This article summarizes the impact of the pharmacogenetics of drug transporters expressed in the enterohepatic circulation on the pharmacokinetics and pharmacodynamics of drugs. The role of pharmacogenetics in the function of drug transporter proteins in vitro is now well established and evidence is rapidly accumulating from in vivo pharmacokinetic studies, which suggests that genetic variants of drug transporter proteins can translate into clinically relevant phenotypes. However, a large amount of conflicting information on the clinical relevance of drug transporter proteins has so far precluded the emergence of a clear picture regarding the role of drug transporter pharmacogenetics in medical practice. This is very well exemplified by the case of P-glycoprotein (MDR1, ABCB1). The challenge is now to develop pharmacogenetic models with sufficient predictive power to allow for translation into drug therapy. This will require a combination of pharmacogenetics of drug transporters, drug metabolism and pharmacodynamics of the respective drugs.     

Role of transporters in drug interactions

Over the past few decades, a tremendous amount of work has been done on the molecular characterization of transport proteins in animals and humans, leading to a better understanding of the physiological roles of a number of transport proteins. Furthermore, there is increasing preclinical and clinical evidence to support the importance of transport proteins in the pharmacokinetics and toxicokinetics of a wide variety of structurally diverse drugs. As a consequence, the degree of expression and functionality of transport proteins may directly affect the therapeutic effectiveness, safety and target specificity of drugs. Recently, there has also been increased awareness about potential drug-drug, drug-herb and drug-food interactions involving transporters. Traditionally, a change in metabolic clearance of a drug, particularly via cytochrome P450-mediated metabolism, has been considered the cause of many clinically important drug interactions. However, increasing evidence suggests that some drug interactions result from changes in the activity and/or expression of drug transporters. Accordingly, assessment of the clinical relevance of transporter-mediated drug interactions has become a regulatory issue during the drug approval process and also the evaluation of drug interaction potential has become an integral part of risk assessment during drug development processes. Therefore, this review will highlight the role of some selected drug transporters in drug interactions, as well as their clinical implication.     

Microdialysis for pharmacokinetic analysis of drug transport to the brain

The intracerebral microdialysis technique represents an important tool for monitoring free drug concentrations in brain extracellular fluid (brain(EcF)) as a function of time. With knowledge of associated free plasma concentrations, it provides information on blood-brain barrier (BBB) drug transport. However, as the implantation of the microdialysis probe evokes tissue reactions, it should be established if the BBB characteristics are maintained under particular microdialysis experimental conditions. Several studies have been performed to evaluate the use of intracerebral microdialysis as a technique to measure drug transport across the BBB and to measure regional pharmacokinetics of drugs in the brain. Under carefully controlled conditions, the intracerebral microdialysis data did reflect passive BBB transport under normal conditions, as well as changes induced by hyperosmolar opening or by the presence of a tumor in the brain. Studies on active BBB transport by the mdr1a-encoded P-glycoprotein (Pgp) were performed, comparing mdr1a(-/-) with wild-type mice. Microdialysis surgery and experimental procedures did not affect Pgp functionality, but the latter did influence in vivo concentration recovery, which was in line with theoretical predictions. It is concluded that intracerebral microdialysis provides meaningful data on drug transport to the brain, only if appropriate methods are applied to determine in vivo concentration recovery.     

Pharmacogenomics of drug transporters: the next drug delivery challenge

Scientifically, the third millennium begins with a major triumph--the publishing of the human genomic map, which is destined to have a momentous impact on the quality of life in our time. Disease prevention, individualized medicine, and genotyped-based medicine will soon become a reality. Pharmacogenetics, the forerunner of pharmacogenomics, began in the 1950s with a series of observations relating drug response to various genetic factors. It took almost two more decades for scientists to discover that cytochrome p450 2D6 was responsible for the metabolism of many drugs. This landmark discovery helped focus attention on how gene expression could impact the response to drugs. The stage was set for a revolution in therapeutics some 30 years later as the Human Genome Project crossed the finishing line triumphantly. A parallel development in drug delivery that may also benefit from the fruits of the Human Genome Project is the growing acceptance/awareness of drug transporters as a gateway to epithelial drug transport. This presentation addresses an area in need of attention: the possible impact of genetic polymorphism of drug transporters in pharmacokinetics and the challenge it poses in drug delivery.     

转运体在药物经肝脏清除过程中的作用

肝脏在药物的体内清除过程中具有重要作用,它不仅是药物代谢的主要场所,还控制着药物及其代谢物的胆汁排泄过程。转运体是控制细胞内外物质传输的一类功能性膜蛋白,其在肝脏有广泛表达,并能对药物进入肝细胞以及排泄至胆汁的过程进行调控,因而,对于肝脏清除过程具有重要作用。本文从肝脏中重要转运体的分布、功能以及底物选择性出发,对其在药物的肝脏清除中的作用、由其引起的药物药物相互作用以及重要转运体的基因多态性研究进行了综述。     

The role of transporters in drug interactions

Transport proteins play an important role in the adsorption, distribution and elimination of a wide variety of drugs. Therefore, it is not surprising that transporter-based drug interactions can occur in the clinic. These interactions can lead to changes in toxicity and/or efficacy of the affected drug. Here, we review such interactions and ask if these interactions could have been predicted from in vitro data. Conducting such in vitro–in vivo correlation is important for predicting future transporter-based drug interactions.     

小鼠微透析技术及其在药物传递系统中的应用

微透析技术（mierodialysis，MD）是由早期神经化学实验室中的灌流取样技术发展而来的一项新技术，后逐渐应用于药学领域，并得到快速发展。早期MD主要以大鼠为实验动物，进行药物传递系统（drug delivery system，DDS）的研究。但是大鼠的实验模型较匮乏，小鼠基因工程（mice genetic engineering，MGE）的出现和迅速发展则弥补了这一不足。     

Cerebral expression of drug transporters in epilepsy

Over-expression of drug efflux transporters at the level of the blood-brain barrier (BBB) has been proposed as a mechanism responsible for multidrug resistance. Drug transporters in epileptogenic tissue are not only expressed in endothelial cells at the BBB, but also in other brain parenchymal cells, such as astrocytes, microglia and neurons, suggesting a complex cell type-specific regulation under pathological conditions associated with epilepsy. This review focuses on the cerebral expression patterns of several classes of well-known membrane drug transporters such as P-glycoprotein (Pgp), and multidrug resistance-associated proteins (MRPs) in the epileptogenic brain. Both experimental and clinical evidence of epilepsy-associated cerebral drug transporter regulation and the possible mechanisms underlying drug transporter regulation are discussed. Knowledge of the cerebral expression patterns of drug transporters in normal and epileptogenic brain will provide relevant information to guide strategies attempting to overcome drug resistance by targeting specific transporters.     

Microdialysis: the Key to Physiologically Based Model Prediction of Human CNS Target Site Concentrations

Despite the enormous research efforts that have been put into the development of central nervous system (CNS) drugs, the success rate in this area is still disappointing. To increase the successful rate in the clinical trials, first the problem of predicting human CNS drug distribution should be solved. As it is the unbound drug that equilibrates over membranes and is able to interact with targets, especially knowledge on unbound extracellular drug concentration-time profiles in different CNS compartments is important. The only technique able to provide such information in vivo is microdialysis. Also, obtaining CNS drug distribution data from human subjects is highly limited, and therefore, we have to rely on preclinical approaches combined with physiologically based pharmacokinetic (PBPK) modeling, taking unbound drug CNS concentrations into account. The next step is then to link local CNS pharmacokinetics to target interaction kinetics and CNS drug effects. In this review, system properties and small-molecule drug properties that together govern CNS drug distribution are summarized. Furthermore, the currently available approaches on prediction of CNS pharmacokinetics are discussed, including in vitro, in vivo, ex vivo, and in silico approaches, with special focus on the powerful combination of in vivo microdialysis and PBPK modeling. Also, sources of variability on drug kinetics in the CNS are discussed. Finally, remaining gaps and challenges are highlighted and future directions are suggested.