Physiologically Based Pharmacokinetic Modelling of Drug Penetration Across the Blood–Brain Barrier—Towards a Mechanistic IVIVE-Based Approach

Predicting the penetration of drugs across the human blood–brain barrier (BBB) is a significant challenge during their development. A variety of in vitro systems representing the BBB have been described, but the optimal use of these data in terms of extrapolation to human unbound brain concentration profiles remains to be fully exploited. Physiologically based pharmacokinetic (PBPK) modelling of drug disposition in the central nervous system (CNS) currently consists of fitting preclinical in vivo data to compartmental models in order to estimate the permeability and efflux of drugs across the BBB. The increasingly popular approach of using in vitro–in vivo extrapolation (IVIVE) to generate PBPK model input parameters could provide a more mechanistic basis for the interspecies translation of preclinical models of the CNS. However, a major hurdle exists in verifying these predictions with observed data, since human brain concentrations can’t be directly measured. Therefore a combination of IVIVE-based and empirical modelling approaches based on preclinical data are currently required. In this review, we summarise the existing PBPK models of the CNS in the literature, and we evaluate the current opportunities and limitations of potential IVIVE strategies for PBPK modelling of BBB penetration.     

Drug targeting to brain: a systematic approach to study the factors,parameters and approaches for prediction of permeability of drugs across BBB

Introduction: Drug targeting to brain by circumventing the physiological barriers is a prerequisite for drugs acting on central nervous system (CNS) and therapeutic potential of many drugs can be improved by effectively targeting the drug(s) to brain.

Areas covered: Present review describes blood–brain barrier (BBB), drug transport mechanisms and factors affecting drug transportation across BBB along with in vitro BBB models; and the approaches for evaluation of permeability of drug across BBB.

Expert opinion: The development of a still awaited perfect in vitro model to mimic BBB is a challenging task. System biologist, network biologist and computational technologist should come together to integrate the role of transporters, physiological and pathophysiological complexity of BBB to replicate vascular properties of the brain microcapillaries as a suitable model to facilitate the high-throughput screening of CNS acting biomolecules.     

Strategies to optimize brain penetration in drug discovery

The kinetics of brain penetration has two components, extent and rate. Achieving a high extent of brain penetration is an important focus for central nervous system (CNS) drug discovery. Optimal brain penetration can be achieved by reducing efflux transport at the blood-brain barrier (BBB), and it is critical to ensure that a high total brain/plasma ratio (the most commonly used parameter for measuring brain penetration) is due to efflux transport activity at the BBB and not related to high non-specific brain tissue binding or low plasma binding. Rapid brain penetration is essential for those drugs that require fast onset of action in the CNS. This can be achieved by increasing passive permeability and reducing brain tissue binding.     

Development of a physiologically based pharmacokinetic model for the rat central nervous system and determination of an in vitro-in vivo scaling methodology for the blood-brain barrier permeability of two transporter substrates, morphine and oxycodone

A whole-body physiologically based pharmacokinetic (PBPK) model was developed for the prediction of unbound drug concentration-time profiles in the rat brain, in which drug transfer across the blood–brain barrier (BBB) was treated mechanistically by separating the parameters governing the rate (permeability) of BBB transfer from brain binding. An in vitro–in vivo scaling strategy based on Caco-2 cell permeability was proposed to extrapolate the active transporter-driven component of this permeability, in which a relative activity factor, RAF, was estimated by fitting the model to rat in vivo profiles. This scaling factor could be interpreted as the ratio of transporter activity between the in vitro system and the in vivo BBB, for a given drug in a given in vitro system. Morphine and oxycodone were selected to evaluate this strategy, as substrates of BBB-located efflux and influx transporters, respectively. After estimation of their respective RAFs using the rat model, the PBPK model was used to simulate human brain concentration profiles assuming the same RAF, and the implications of this were discussed. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 101:4277–4292, 2012     

In vitro blood–brain barrier model adapted to repeated-dose toxicological screening

Toxicity to the central nervous system (CNS) is a key feature in the toxicological profile of compounds and there is a growing interest to use in vitro cell assays.The blood–brain barrier (BBB) is a highly restrictive barrier that preserves homeostasis within the brain microenvironment. By modelling the BBB it is possible to investigate whether a compound is likely to compromise its functionality, which would cause unwanted effects on brain cells. These investigations are usually performed using a single exposure to drugs, whereas CNS side effects usually result from repeated exposures.The main objective of this study was to adapt our established BBB model to the evaluation of repeated-dose toxicity at the BBB.Studies were undertaken within the European Predict-IV consortium to study the effect on BBB permeability of 12 selected drugs after 14 days of repeated treatment to a single pre-selected concentration.Compared to single exposure, a 100-fold lower colchicine concentration in 14 days repeated-dose treatment was toxic. This demonstrates the importance to evaluate the BBB toxicity in repeated-dose testing. Finally, the potentiating effects of cyclosporin A on the BBB toxicity of colchicine illustrate the possibility to use in vitro BBB models to make risk assessment of drug–drug interactions.     

Molecular mechanisms of drug influx and efflux transport at the blood-brain barrier

The blood-brain barrier (BBB) segregates the circulating blood from interstitial fluid in the brain and restricts drug permeability into the brain. Recent studies have revealed that the BBB exhibits not only blood-to-brain influx transport for the supply of nutrients, but also brain-to-blood efflux transport to excrete drugs and endogenous compounds. The influx transport system allows drugs to enter the brain. (L)-DOPA is transported into the brain by the large neutral amino acid transport system, system L. A cationic mu-opioid peptide analogue enters the brain by adsorptive-mediated endocytosis. In contrast, efflux transport limits the distribution of drugs in the brain. The ATP binding cassette transporter B1 (ABCB1) mediates the efflux transport of lipophilic drugs at the BBB by using ATP energy. Furthermore, organic anion transporter 3 (OAT3) is expressed at the BBB and mediates the efflux transport of homovanillic acid, a dopamine metabolite. This efflux transport is also likely to be involved in the transport of anionic drugs such as 6-mercaptopurine and acyclovir. Clarifying the BBB transport could give us important information allowing the development of better CNS drugs and improving our understanding of the relationship between CNS diseases and BBB functions.     

Contribution of Carrier-Mediated Transport Systems to the Blood–Brain Barrier as a Supporting and Protecting Interface for the Brain; Importance for CNS Drug Discovery and Development

The blood–brain barrier (BBB) forms an interface between the circulating blood and the brain and possesses various carrier-mediated transport systems for small molecules to support and protect CNS function. For example, the blood-to-brain influx transport systems supply nutrients, such as glucose and amino acids. Consequently, xenobiotic drugs recognized by influx transporters are expected to have high permeability across the BBB. On the other hand, efflux transporters, including ATP-binding cassette transporters such as P-glycoprotein located at the luminal membrane of endothelial cells, function as clearance systems for metabolites and neurotoxic compounds produced in the brain. Drugs recognized by these transporters are expected to show low BBB permeability and low distribution to the brain. Despite recent progress, the transport mechanisms at the BBB have not been fully clarified yet, especially in humans. However, an understanding of the human BBB transport system is critical, because species differences mean that it can be difficult to extrapolate data obtained in experimental animals during drug development to humans. Recent progress in methodologies is allowing us to address this issue. Positron emission tomography can be used to evaluate the activity of human BBB transport systems in vivo. Proteomic studies may also provide important insights into human BBB function. Construction of a human BBB transporter atlas would be a most important advance from the viewpoint of CNS drug discovery and drug delivery to the brain.     

In vitro primary human and animal cell-based blood-brain barrier models as a screening tool in drug discovery

Brain penetration is characterized by its extent and rate and is influenced by drug physicochemical properties, plasma exposure, plasma and brain protein binding and BBB permeability. This raises questions related to physiology, interspecies differences and in vitro/in vivo extrapolation. We herein discuss the use of in vitro human and animal BBB model as a tool to improve CNS compound selection. These cell-based BBB models are characterized by low paracellular permeation, well-developed tight junctions and functional efflux transporters. A study of twenty drugs shows similar compound ranking between rat and human models although with a 2-fold higher permeability in rat. cLogP < 5, PSA < 120 ?, MW < 450 were confirmed as essential for CNS drugs. An in vitro/in vivo correlation in rat (R2 = 0.67; P = 2 × 10??) was highlighted when in vitro permeability and efflux were considered together with plasma exposure and free fraction. The cell-based BBB model is suitable to optimize CNS-drug selection, to study interspecies differences and then to support human brain exposure prediction.     

The impact of efflux transporters in the brain on the development of drugs for CNS disorders

The development of drugs to treat disorders of the CNS requires consideration of achievable brain concentrations. Factors that influence the brain concentrations of drugs include the rate of transport into the brain across the blood-brain barrier (BBB), metabolic stability of the drug, and active transport out of the brain by efflux mechanisms. To date, three classes of transporter have been implicated in the efflux of drugs from the brain: multidrug resistance transporters, monocarboxylic acid transporters, and organic ion transporters. Each of the three classes comprises multiple transporters, each of which has multiple substrates, and the combined substrate profile of these transporters includes a large number of commonly used drugs. This system of transporters may therefore provide a mechanism through which the penetration of CNS-targeted drugs into the brain is effectively minimised. The action of these efflux transporters at the BBB may be reflected in the clinic as the minimal effectiveness of drugs targeted at CNS disorders, including HIV dementia, epilepsy, CNS-based pain, meningitis and brain cancers. Therefore, modulation of these efflux transporters by design of inhibitors and/or design of compounds that have minimal affinity for these transporters may well enhance the treatment of intractable CNS disorders.     

Blood-brain barrier permeation and efflux exclusion of anticholinergics used in the treatment of overactive bladder

Overactive bladder (OAB) is a common condition, particularly in the elderly. Anticholinergic agents are the mainstay of pharmacological treatment of OAB; however, many anticholinergics can cross the blood-brain barrier (BBB) and may cause central nervous system (CNS) effects, including cognitive deficits, which can be especially detrimental in older patients. Many anticholinergics have the potential to cause adverse CNS effects due to muscarinic (M(1)) receptor binding in the brain. Of note, permeability of the BBB increases with age and can also be affected by trauma, stress, and some diseases and medications. Passive crossing of a molecule across the BBB into the brain is dependent upon its physicochemical properties. Molecular characteristics that hinder passive BBB penetration include a large molecular size, positive or negative ionic charge at physiological pH, and a hydrophilic structure. Active transport across the BBB is dependent upon protein-mediated transporter systems, such as that of permeability-glycoprotein (P-gp), which occurs only for P-gp substrates, such as trospium chloride, darifenacin and fesoterodine. Reliance on active transport can be problematic since genetic polymorphisms of P-gp exist, and many commonly used drugs and even some foods are P-gp inhibitors or are substrates themselves and, due to competition, can reduce the amount of the drug that is actively transported out of the CNS. Therefore, for drugs that are preferred not to cross into the CNS, such as potent anticholinergics intended for the bladder, it is optimal to have minimal passive crossing of the BBB, although it may also be beneficial for the drug to be a substrate for an active efflux transport system. Anticholinergics demonstrate different propensities to cross the BBB. Darifenacin, fesoterodine and trospium chloride are substrates for P-gp and, therefore, are actively transported away from the brain. In addition, trospium chloride has not been detected in cerebrospinal fluid assays and does not appear to have significant CNS penetration. This article reviews the properties of anticholinergics that affect BBB penetration and active transport out of the CNS, discusses issues of increased BBB permeability in patients with OAB, and examines the clinical implications of BBB penetration on adverse events associated with anticholinergics.     

Minimizing Drug Exposure in the CNS while Maintaining Good Oral Absorption

In some drug discovery approaches, it is advantageous to restrict the access of compounds to the CNS to minimize the risk of side effects. By choosing appropriate physicochemical properties and building in the ability to act as substrates for active efflux transporters, it is possible to achieve CNS restriction and still retain sufficient absorption through the intestinal epithelium to retain good oral bioavailability. Potential risks in employing this approach are considered.For drugs that are required to act at targets outside of the central nervous system (CNS), it may be advantageous to minimize drug exposure in the CNS. Many instances exist where side effects have been attributed to on- or off-target actions of a drug in the CNS that lead to issues of safety and tolerability. Furthermore, in the research phase, the ability to test a novel pharmacological mechanism could be limited by such side effects.The first generation of histamine H1 antagonists used for the treatment of allergic reactions serves as an example, whereby diphenhydramine, while effective as an antiallergic agent, also caused somnolence and other CNS side effects as a result of engagement with H1 receptors in the brain. The second generation agents, for example, cetirizine, had reduced side effects with reduced somnolence at therapeutic doses, while the third generation, including fexofenadine, were free of sedation at doses higher than those used for treatment of allergic reactions. This progression resulted from increasing CNS restriction of these agents, thereby increasing their peripheral H1 selectivity.¹ Other examples include antimuscarinic agents used for the treatment of overactive bladder, which act by binding to muscarinic receptors in the bladder detrusor muscle. Effects such as cognitive impairment, particularly in elderly patients, have been reported for agents such as oxybutynin, which penetrate the CNS readily and are thus able to interact with centrally located muscarinic receptors. Other agents such as darifenacin and 5-hydroxymethyltolterodine (active metabolite of fesoterodine) are not associated with CNS side effects and are largely excluded from the CNS.²Therefore, a general approach that may be advantageous when considering peripherally located drug targets is to restrict the access of compounds to the CNS while maintaining appropriate exposure in peripheral tissues. This may apply particularly when the peripheral therapeutic target is known to be present in the CNS but whose engagement there is not required for desired pharmacological activity. However, it also represents a general means of minimizing risk of unexpected off-target effects in the CNS, thereby increasing therapeutic index.The properties of the brain capillary vascular endothelium that supply blood to the CNS provide a barrier to the free exchange of blood-borne solutes. Efficient tight junctions between adjacent brain vascular endothelial cells (BVECs) restrict passage of solutes between adjacent cells (paracellular movement) so that to traverse the endothelium, compounds have to cross the BVEC plasma membrane (transcellular movement). Hence, the physicochemical properties of a brain penetrant compound need to be compatible with the ability to diffuse passively across the plasma membrane and/or participate in active uptake. In addition, ATP-dependent transporter proteins such as P-glycoprotein (P-gp) and Breast Cancer Resistance Protein (BCRP), expressed in the BVEC apical membrane, are capable of ejecting substrate compounds from the cell. These features of the BVECs constituting the blood–brain barrier (BBB) offer opportunities to design compounds with properties that exploit the requirement for transcellular movement and presence of transporter proteins, to achieve the goal of restricted CNS penetration.However, the properties of orally administered compounds should also be compatible with those required for absorption across the intestinal epithelium that acts as a permeability barrier in the gastrointestinal tract. Molecular weight (MW) < 500, polar surface area (PSA) < 140, and <10 rotatable bonds have been associated with good oral absorption, while MW < 450 and PSA < 70 have been indicated as requirements for good CNS penetration.^3,4 Hence, to favor restriction from the CNS while allowing good absorption in the gastrointestinal tract may point to an area of compatibility of MW of 450–500 and PSA of 70–140. Like the BVECs, the intestinal epithelium contains several efflux transporter proteins, including P-gp and BCRP, expressed on the apical membrane of intestinal epithelial cells (enterocytes) (Figure ​(Figure11).Open in a separate windowFigure 1Schematic diagram of the distribution of transporter proteins in the intestinal epithelium and brain vascular endothelium.P-gp and BCRP are expressed at comparable levels in human brain capillaries, and in mouse gene knockout studies, it has been shown that they may both contribute to exclusion of substrates from the brain.⁵ This suggests that design of compounds that act as substrates for both P-gp and BCRP may maximize their CNS restriction. Indeed, P-gp and BCRP display considerable overlap in their substrates (e.g., imatinib is a substrate of both), although some compounds are exclusively substrates of one or the other (e.g., cetirizine is P-gp only). Increasing MW and PSA increases the likelihood of compounds to act as substrates of P-gp. Additional features include possession of hydrogen bond acceptors and modest ionization potential (acid pK_a > 4; basic pK_a < 8). These features broadly align with those identified for balancing CNS restriction and intestinal absorption.Targeting efflux transporters as part of a drug discovery strategy may suggest a conundrum if efflux transporter expression in enterocytes renders CNS restriction and good oral absorption incompatible. However, this could be a misconception as there are several instances of drugs that are substrates of P-gp and BCRP, CNS restricted, and possess good oral bioavailability. Considering drug doses commonly prescribed for clinical use (10–500 mg) and the resulting range of drug concentrations likely to exist in the gastrointestinal lumen following an oral dose (assuming dissolution in ∼250 mL), P-gp is often likely to be saturated by drug substrates in the gut, given that the K_m for P-gp is usually in the range 1–100 μM.⁶ In contrast, systemic unbound drug concentrations are likely to be in the submicromolar range and hence unlikely to be at concentrations sufficient to saturate transporters in the BVECs. For example, the antitumor agent imatinib is a P-gp and BCRP substrate, with limited brain exposure and high oral bioavailability.⁷ The unbound plasma C_max of imatinib following a dose of 400 mg is approximately 250 nM and is unlikely to saturate P-gp or BCRP at the BBB. The antiviral protease inhibitors ritonavir and indinavir serve as other examples of CNS restricted P-gp substrates having high (60–78%) oral bioavailability.⁵Steady state brain concentrations of a compound result from the net effect of passive and active movements across the BBB, so strategies designed to exclude compounds from the brain could focus on active and passive processes. Maintaining very low passive permeability such that equilibrium between blood and brain tissue is not allowed to occur may have the drawback of impairing intestinal absorption. While, in this case, a nonoral dose route could be explored, oral administration is usually the preferred dose route. In our opinion, the strategy most likely to deliver CNS restriction with good oral absorption is to maintain an efflux rate at the BBB that greatly exceeds influx rate, whereby efflux is mediated by P-gp and BCRP against a background of low-moderate passive permeability. We have utilized this approach successfully at Pfizer to design CNS restricted orally bioavailable ligands.⁵ A series of CNS restricted histamine H3 antagonists was designed to minimize clinical adverse events such as insomnia that would otherwise be observed. Optimizing PSA, reducing passive permeability, and introduction of activity as P-gp and BCRP substrates led to demonstration of CNS restriction in in vivo tissue partition experiments in rat. Good oral bioavailability (>50%) was maintained in rat while brain receptor occupancy data confirmed that CNS restriction was maintained over 7 days of dosing, and electroencephalography data demonstrated the desired TI for efficacy over insomnia.While the H3 antagonist approach dealt with an extracellular target, the design of CNS-restricted drug candidates for intracellular drug targets must incorporate sufficient cellular permeability to reach the site of action, yet maintain low BBB penetration. Therefore, the use of cell-based primary screens together with timely in vivo efficacy and CNS restriction experiments is vital to ensure that candidate compounds combine efficacy and CNS restriction. By application of this approach, we have developed CNS restricted ligands (rat unbound brain:plasma ratio 0.015) for an intracellular target having high cellular potencies (IC₅₀ ≤ 20 nM) combined with good oral absorption, as demonstrated by linear pharmacokinetics over a wide dose range (0.25–1000 mg/kg) in preclinical rodent safety studies.There are identifiable risks associated with building in P-gp and BCRP active efflux to a drug approach, some of which can be addressed by evaluation of clinical data. A drug–drug interaction (DDI), potentially leading to unwanted CNS penetration, could arise if a P-gp substrate is concomitantly administered with a P-gp inhibitor. However, considering the free drug exposures expected at the BBB, only a very potent P-gp inhibitor could be expected to elicit a significant effect. DDI associated with absorption could be expected, given P-gp expression along the intestinal epithelium. Nevertheless, clinical data obtained with the P-gp substrate digoxin suggest that in the majority of cases when a P-gp inhibitor and substrate are coadministered, the digoxin AUC change was less than 2-fold.⁸ It is also possible that P-gp substrates will display nonlinear dose versus exposure relationships, depending on their Km for P-gp. However, as metabolism by CYP3A4, and hence first-pass extraction, often accompany P-gp affinity,⁶ it may be difficult to assess the contribution of each enzyme to any nonlinearity observed. Presently, our ability to accurately predict absorption of P-gp and BCRP substrates is limited until more quantitative information on intestinal transporter expression become available. A number of polymorphisms of P-gp and BCRP are present in the human population that could lead to interpatient variability. For instance, the MDR1 gene single nucleotide polymorphism C3435T is linked to decreased duodenal P-gp expression and modest increases in digoxin exposure. Similarly, changes in BBB permeability and P-gp expression may occur with aging and in certain disease states that may alter the degree of CNS restriction. Finally, a significant concern in compound selection for clinical studies may be whether CNS restriction measured preclinically accurately predicts that which occurs in human. Many preclinical evaluations are conducted in rodents whose transporter expression profile at the BBB differs from human. Furthermore, a number of recent studies indicate that the degree of CNS restriction can exhibit species differences whereby higher primate species, including human, may display significantly higher CNS exposure than in rodents.⁵In conclusion, designing in CNS restriction can be used to improve drug safety. Targeting the efflux transporters P-gp and BCRP alongside modest passive permeability can confer significant CNS restriction while retaining good oral bioavailability, cell penetration, and pharmacological activity. However, there are identifiable risks with this strategy that may be clarified as further clinical data emerge.     

Murine in vitro model of the blood–brain barrier for evaluating drug transport

In vitro blood–brain barrier (BBB) models help predict brain uptake of potential central nervous system drug candidates. Current in vitro models are composed of brain microvascular endothelial cells (BMEC) that are isolated from rat, bovine, or porcine. However, most in vivo studies on drug transport through the BBB are performed in small laboratory animals, specially mouse and thus murine in vitro BBB models serve as better surrogates to correlate with these studies. Here we describe the functional characterization of a reproducible in vitro model composed of murine BMEC co-cultured with rat primary astrocytes in the presence of biochemical inducing agents. The co-cultures presented high TEER and low sodium fluorescein permeability. Expression of specific BBB tight junction proteins (occludin, claudin-5, ZO-1) and the functionality of transporters (Pgp, GLUT1) were detected by immunocytochemistry and Western blotting. These results indicated a 2.5-fold increase in the expression levels of these proteins in the presence of astrocytes. In addition, a high correlation coefficient (0.98) was obtained between the permeability of a series of hydrophobic and hydrophilic drugs and their corresponding in vivo values. These results together establish the utility of this murine model for future drug transport, pathological, and pharmacological characterizations of the BBB.     

New experimental models of the blood-brain barrier for CNS drug discovery

Introduction: The blood-brain barrier (BBB) is a dynamic biological interface which actively controls the passage of substances between the blood and the central nervous system (CNS). From a biological and functional standpoint, the BBB plays a crucial role in maintaining brain homeostasis inasmuch that deterioration of BBB functions are prodromal to many CNS disorders. Conversely, the BBB hinders the delivery of drugs targeting the brain to treat a variety of neurological diseases.

Area covered: This article reviews recent technological improvements and innovation in the field of BBB modeling including static and dynamic cell-based platforms, microfluidic systems and the use of stem cells and 3D printing technologies. Additionally, the authors laid out a roadmap for the integration of microfluidics and stem cell biology as a holistic approach for the development of novel in vitro BBB platforms.

Expert opinion: Development of effective CNS drugs has been hindered by the lack of reliable strategies to mimic the BBB and cerebrovascular impairments in vitro. Technological advancements in BBB modeling have fostered the development of highly integrative and quasi- physiological in vitro platforms to support the process of drug discovery. These advanced in vitro tools are likely to further current understanding of the cerebrovascular modulatory mechanisms.     

Blood-brain barrier drug discovery for central nervous system infections

Central nervous system (CNS) infections are formidable diseases with high rates of morbidity and mortality. Since the majority of antimicrobial agents discovered so far do not cross the blood-brain barrier (BBB), the treatment of CNS infections is a major challenge issue. The development of drugs to treat those diseases requires consideration of achievable brain concentrations by targeting the following question. How can the chemistry and biology of the BBB, and infectomics be exploited for the development of drugs against CNS infections? To date drug targeting approaches, such as chemistry-based, biology-based, and infectomics-based, have been implicated in the development of drugs for treatment of CNS infections. The chemistry-based strategies rely on lipid-mediated BBB drug transport as substances that readily permeate the BBB. These usually include small molecular weight of lipophilic or hydrophobic molecules. The biology-based strategies depend on endogenous BBB transport systems, including carrier-mediated transport (CMT), active efflux transport (AET), and receptor-mediated transport (RMT). These transporters play important roles in the influxes and/or effluxes of drugs including antimicrobial agents in brain capillary endothelial cells that form the BBB. Both microbial and host signatures of infectomes, which can be dissected by infectomics, provide invaluable fountains in the search for novel antimicrobial therapies. Key markers associated with the mechanisms of neuronal injury may be identified, and thus, provide important targets for the prevention and treatment of CNS infections. This review focuses on the major BBB drug targeting strategies in the development of therapeutics for CNS infections. A combination of these strategies will ultimately lead to improved treatments.     

Strategies for increasing drug delivery to the brain: focus on brain lymphoma

The blood-brain barrier (BBB) is a gate that controls the influx and efflux of a wide variety of substances and consequently restricts the delivery of drugs into the central nervous system (CNS). Brain tumours may disrupt the function of this barrier locally and nonhomogeneously. Therefore, the delivery of drugs to brain tumours has long been a controversial subject. The current concept is that inadequate drug delivery is a major factor that explains the unsatisfactory response of chemosensitive brain tumours. Various strategies have been devised to circumvent the BBB in order to increase drug delivery to the CNS. The various approaches can be categorised as those that attempt to increase delivery of intravascularly administered drugs, and those that attempt to increase delivery by local drug administration. Strategies that increase delivery of intravascularly injected drugs can manipulate either the drugs or the capillary permeability of the various barriers (BBB or blood-tumour barrier), or may attempt to increase plasma concentration or the fraction of the drug reaching the tumour (high-dose chemotherapy, intra-arterial injection). Neurotoxicity is a major concern with increased penetration of drugs into the CNS or when local delivery is practised. Systemic toxicity remains the limiting factor for most methods that use intravascular delivery. This review evaluates the strategies used to increase drug delivery in view of current knowledge of drug pharmacokinetics and its relevance to clinical studies of chemosensitive brain tumours. The main focus is on primary CNS lymphoma, as it is a chemosensitive brain tumour and its management routinely utilises specialised strategies to enhance drug delivery to the affected CNS compartments.     

Physiological function of blood-brain barrier transporters as the CNS supporting and protecting system

The blood-brain barrier (BBB) segregates the circulating blood from interstitial fluid in the brain and restricts drug permeability into the brain. Our latest studies have revealed that the BBB transporters play important physiological roles in maintaining the brain environment. For an energy-storing system, the creatine transporter localized at the brain capillary endothelial cells (BCECs) mediates the supply of creatine from the blood to the brain. The BBB is involved in the brain-to-blood efflux transport of gamma-aminobutyric acid, and GAT2/BGT-1 mediates this transport process. BCECs also express serotonin and norepinephrine transporters. Organic anion transporter 3 (OAT3) and ASCT2 are localized at the abluminal membrane of the BCECs. OAT3 is involved in the brain-to-blood efflux of a dopamine metabolite, a uremic toxin, and thiopurine nucleobase analogues. ASCT2 plays a role in L-isomer-selective aspartic acid efflux transport at the BBB. Dehydroepiandrosterone sulfate and small neutral amino acids undergo brain-to-blood efflux transport mediated by organic anion transporting polypeptide 2 and ATA2, respectively. The BBB transporters are regulated by various factors: ATA2 by osmolarity, taurine transporter by tumor necrosis factor-alpha, and L-cystine/L-glutamic acid exchange transporter by oxidative stress. Clarifying the physiological roles of BBB transport systems should give important information allowing the development of better central nervous system (CNS) drugs and improving our understanding of the relationship between CNS disorders and BBB function.     

Strategies to assess blood–brain barrier penetration

Background: The principles and screening strategies for brain penetration in drug discovery are important in identifying drug candidates with desirable CNS properties. Objective: Define key variables and assays that are essential for determining brain penetration. Methods: This review covers issues, methods, and strategies for assessing brain penetration of small molecules in drug discovery. Results/conclusion: Brain penetration is assessed using both initial rate and extent at steady-state. Unbound drug is the active species that exerts pharmacological effects. Low brain penetration can be due to low blood–brain barrier (BBB) permeability, P-glycoprotein (Pgp) efflux, or high plasma protein binding. Successful methods include: parallel artificial membrane permeability assay (PAMPA)-BBB permeability, MDR1-MDCKII for Pgp efflux, B-P dialysis for fraction unbound, and in vivo B/P ratio to extrapolate unbound brain drug concentration.     

Hypoxic Stress and Inflammatory Pain Disrupt Blood-Brain Barrier Tight Junctions: Implications for Drug Delivery to the Central Nervous System

A functional blood-brain barrier (BBB) is necessary to maintain central nervous system (CNS) homeostasis. Many diseases affecting the CNS, however, alter the functional integrity of the BBB. It has been shown that various diseases and physiological stressors can impact the BBB’s ability to selectively restrict passage of substances from the blood to the brain. Modifications of the BBB’s permeability properties can potentially contribute to the pathophysiology of CNS diseases and result in altered brain delivery of therapeutic agents. Hypoxia and/or inflammation are central components of a number of diseases affecting the CNS. A number of studies indicate hypoxia or inflammatory pain increase BBB paracellular permeability, induce changes in the expression and/or localization of tight junction proteins, and affect CNS drug uptake. In this review, we look at what is currently known with regard to BBB disruption following a hypoxic or inflammatory insult in vivo. Potential mechanisms involved in altering tight junction components at the BBB are also discussed. A more detailed understanding of the mediators involved in changing BBB functional integrity in response to hypoxia or inflammatory pain could potentially lead to new treatments for CNS diseases with hypoxic or inflammatory components. Additionally, greater insight into the mechanisms involved in TJ rearrangement at the BBB may lead to novel strategies to pharmacologically increase delivery of drugs to the CNS.     

The use of microdialysis in CNS drug delivery studies. Pharmacokinetic perspectives and results with analgesics and antiepileptics

Microdialysis can give simultaneous information on unbound drug concentration-time profiles in brain extracellular fluid (ECF) and blood, separating the information on blood-brain barrier (BBB) processes from confounding factors such as binding to brain tissue or proteins in blood. This makes microdialysis suitable for studies on CNS drug delivery. It is possible to quantify influx and efflux processes at the BBB in vivo, and to relate brain ECF concentrations to central drug action. The half-life in brain ECF vs. the half-life in blood gives information on rate-limiting steps in drug delivery and elimination from the CNS. Examples are given on microdialysis studies of analgesic and antiepileptic drugs.     

New aspects of the blood-brain barrier transporters; its physiological roles in the central nervous system

The blood-brain barrier (BBB) segregates the circulating blood from interstitial fluid in the brain, and restricts drug permeability into the brain. Our latest studies have revealed that the BBB transporters play important physiological roles in maintaining the brain milieu. The BBB supplies creatine to the brain for an energy-storing system, and creatine transporter localized at the brain capillary endothelial cells (BCECs) is involved in BBB creatine transport. The BBB is involved in the brain-to-blood efflux transport of the suppressive neurotransmitter, gamma-aminobutyric acid, and GAT2/BGT-1 mediates this transport process. BCECs also express serotonin and norepinephrine transporters. Organic anion transporter 3 (OAT3) and ASCT2 are localized at the abluminal membrane of the BCECs. OAT3 is involved in the brain-to-blood efflux of a dopamine metabolite, a uremic toxin and thiopurine nucleobase analogs. ASCT2 plays a role in L-isomer-selective aspartic acid efflux transport at the BBB. Dehydroepiandrosterone sulfate and small neutral amino acids undergo brain-to-blood efflux transport mediated by organic anion transporting polypeptide 2 and ATA2, respectively. The BBB transporters are regulated by various factors, ATA2 by osmolarity, taurine transporter by TNF-alpha, and L-cystine/L-glutamic acid exchange transporter by oxidative stress. Clarifying the physiological roles of BBB transport systems should give us important information allowing the development of better CNS drugs and improving our understanding of the relationship between CNS disorders and BBB function.