Active Uptake of Oxycodone at Both the Blood-Cerebrospinal Fluid Barrier and The Blood-Brain Barrier without Sex Differences: A Rat Microdialysis Study.

BACKGROUND: Oxycodone active uptake across the blood-brain barrier (BBB) is associated with the putative proton-coupled organic cation (H+/OC) antiporter system. Yet, the activity of this system at the blood-cerebrospinal fluid barrier (BCSFB) is not fully understood. Additionally, sex differences in systemic pharmacokinetics and pharmacodynamics of oxycodone has been reported, but whether the previous observations involve sex differences in the function of the H+/OC antiporter system remain unknown. The objective of this study was, therefore, to investigate the extent of oxycodone transport across the BBB and the BCSFB in female and male Sprague-Dawley rats using microdialysis. METHODS: Microdialysis probes were implanted in the blood and two of the following brain locations: striatum and lateral ventricle or cisterna magna. Oxycodone was administered as an intravenous infusion, and dialysate, blood and brain were collected. Unbound partition coefficients (Kp,uu) were calculated to understand the extent of oxycodone transport across the blood-brain barriers. Non-compartmental analysis was conducted using Phoenix 64 WinNonlin. GraphPad Prism version 9.0.0 was used to perform t-tests, one-way and two-way analysis of variance followed by Tukey's or Sídák's multiple comparison tests. Differences were considered significant at p < 0.05. RESULTS: The extent of transport at the BBB measured in striatum was 4.44 ± 1.02 (Kp,uu,STR), in the lateral ventricle 3.41 ± 0.74 (Kp,uu,LV) and in cisterna magna 2.68 ± 1.01 (Kp,uu,CM). These Kp,uu values indicate that the extent of oxycodone transport is significantly lower at the BCSFB compared with that at the BBB, but still confirm the presence of active uptake at both blood-brain interfaces. No significant sex differences were observed in neither the extent of oxycodone delivery to the brain, nor in the systemic pharmacokinetics of oxycodone. CONCLUSIONS: The findings clearly show that active uptake is present at both the BCSFB and the BBB. Despite some underestimation of the extent of oxycodone delivery to the brain, CSF may be an acceptable surrogate of brain ISF for oxycodone, and potentially also other drugs actively transported into the brain via the H+/OC antiporter system.

Neuropharmacokinetic visualization of regional and subregional unbound antipsychotic drug transport across the blood-brain barrier.

Comprehensive determination of the extent of drug transport across the region-specific blood-brain barrier (BBB) is a major challenge in preclinical studies. Multiple approaches are needed to determine the regional free (unbound) drug concentration at which a drug engages with its therapeutic target. We present an approach that merges in vivo and in vitro neuropharmacokinetic investigations with mass spectrometry imaging to quantify and visualize both the extent of unbound drug BBB transport and the post-BBB cerebral distribution of drugs at regional and subregional levels. Direct imaging of the antipsychotic drugs risperidone, clozapine, and olanzapine using this approach enabled differentiation of regional and subregional BBB transport characteristics at 20-µm resolution in small brain regions, which could not be achieved by other means. Our approach allows investigation of heterogeneity in BBB transport and presents new possibilities for molecular psychiatrists by facilitating interpretation of regional target-site exposure results and decision-making.

Brain Distribution of Drugs: Pharmacokinetic Considerations.

It is crucial to understand the basic principles of drug transport, from the site of delivery to the site of action within the CNS, in order to evaluate the possible utility of a new drug candidate for CNS action, or possible CNS side effects of non-CNS targeting drugs. This includes pharmacokinetic aspects of drug concentration-time profiles in plasma and brain, blood-brain barrier transport and drug distribution within the brain parenchyma as well as elimination processes from the brain. Knowledge of anatomical and physiological aspects connected with drug delivery is crucial in this context. The chapter is intended for professionals working in the field of CNS drug development and summarizes key pharmacokinetic principles and state-of-the-art experimental methodologies to assess brain drug disposition. Key parameters, describing the extent of unbound (free) drug across brain barriers, in particular blood-brain and blood-cerebrospinal fluid barriers, are presented along with their application in drug development. Special emphasis is given to brain intracellular pharmacokinetics and its role in evaluating target engagement. Fundamental neuropharmacokinetic differences between small molecular drugs and biologicals are discussed and critical knowledge gaps are outlined.

In Vivo Studies of Drug BBB Transport: Translational Challenges and the Role of Brain Imaging.

Drug delivery to the brain is challenging to study due to the complexity of the barriers of the central nervous system (CNS). The present chapter describes and compares experimental methods such as microdialysis, two-photon laser scanning fluorescence microscopy and positron emission tomography (PET) that can be used for in vivo studies of drug transport across the blood-brain barrier (BBB). The selection of appropriate method is based on the research question, and the different methods will in most cases provide complementary information. Attention is also given to the fact that the BBB might undergo changes in integrity, protein expression and other morphological alterations as a result of disease. The use of animal models of human disease is therefore also discussed. Special emphasis is given to translational aspects of the different methods and readouts.

Perspectives on Nanodelivery to the Brain: Prerequisites for Successful Brain Treatment.

Nanocarriers (NCs) are promising tools to improve drug delivery across the blood-brain barrier (BBB) for more effective treatment of brain disorders, although there is a scarcity of clinical translation of brain-directed NCs. In order to drive the development of brain-oriented NCs toward clinical success, it is essential to understand the prerequisites for nanodelivery to be successful in brain treatment. In this Perspective, we present how pharmacokinetic/pharmacodynamic (PK/PD), formulation and nanotoxicity factors impact the therapeutic success of brain-specific nanodelivery. Properties including high loading efficiency, slow in vivo drug release, long systemic circulation, an increase in unbound brain-to-plasma concentration/exposure ratio (Kp,uu,brain), high drug potency, and minimal nanotoxicity are prerequisites that should preferably be combined to maximize the therapeutic potential of a brain-targeted NC. The PK of brain-directed NCs needs to be evaluated in a more therapeutically relevant manner, focusing on the released, unbound drug. It is more crucial to increase the Kp,uu,brain than to improve the ability of the NC to cross the BBB in its intact form. Brain-targeted NCs, which are mostly developed for treating brain tumors, including metastases, should aim to enhance drug delivery not just to tumor regions with disrupted BBB, but equally important to regions with intact BBB where the drugs themselves have problems reaching. This article provides critical insights into how a brain-targeted nanoformulation needs to be designed and optimized to achieve therapeutic success in the brain.

Mechanistic Study on the Use of the l-Type Amino Acid Transporter 1 for Brain Intracellular Delivery of Ketoprofen via Prodrug: A Novel Approach Supporting the Development of Prodrugs for Intracellular Targets.

l-Type amino acid transporter 1 (LAT1), selectively expressed at the blood-brain barrier (BBB) and brain parenchymal cells, mediates brain delivery of drugs and prodrugs such as l-dopa and gabapentin. Although knowledge about BBB transport of LAT1-utilizing prodrugs is available, there is a lack of quantitative information about brain intracellular delivery and influence of prodrugs on the transporter's physiological state. We studied the LAT1-mediated intrabrain distribution of a recently developed prodrug of the cyclooxygenase inhibitor ketoprofen as well as its impact on transporter protein expression and function (i.e., amino acid exchange) using brain slice method in mice and rats. The intrabrain distribution of the prodrug was 16 times higher than that of ketoprofen. LAT1 involvement in brain cellular barrier uptake of the prodrug was confirmed, reflected by a higher unbound brain intracellular compared to brain extracellular fluid concentration. The prodrug did not alter LAT1 protein expression and amino acid exchange. Integration of derived parameters with previously performed in vivo pharmacokinetic study using the Combinatory Mapping Approach allowed to estimate the brain extra- and intracellular levels of unbound ketoprofen, prodrug, and released parent drug. The overall efficiency of plasma to brain intracellular delivery of prodrug-released ketoprofen was 11 times higher than after ketoprofen dosing. In summary, this study provides quantitative information supporting the use of the LAT1-mediated prodrug approach for enhanced brain delivery of drugs with intracellular targets.

Rate and extent of mitragynine and 7-hydroxymitragynine blood-brain barrier transport and their intra-brain distribution: the missing link in pharmacodynamic studies.

Mitragyna speciosa is reported to be beneficial for the management of chronic pain and opioid withdrawal in the evolving opioid epidemic. Data on the blood-brain barrier (BBB) transport of mitragynine and 7-hydroxymitragynine, the active compounds of the plant, are still lacking and inconclusive. Here, we present for the first time the rate and the extent of mitragynine and 7-hydroxymitragynine transport across the BBB, with an investigation of their post-BBB intra-brain distribution. We utilized an in vitro BBB model to study the rate of BBB permeation of the compounds and their interaction with efflux transporter P-glycoprotein (P-gp). Mitragynine showed higher apical-to-basolateral (A-B, i.e. blood-to-brain side) permeability than 7-hydroxymitragynine. 7-Hydroxymitragynine showed a tendency to efflux, with efflux ratio (B-A/A-B) of 1.39. Both were found to inhibit the P-gp and are also subject to efflux by the P-gp. Assessment of the extent of BBB transport in vivo in rats from unbound brain to plasma concentration ratios (Kp,uu,brain ) revealed extensive efflux of both compounds, with less than 10 percent of unbound mitragynine and 7-hydroxymitragynine in plasma crossing the BBB. By contrast, the extent of intra-brain distribution was significantly different, with mitragynine having 18-fold higher brain tissue uptake in brain slice assay compared with 7-hydroxymitragynine. Mitragynine showed a moderate capacity to accumulate inside brain parenchymal cells, while 7-hydroxymitragynine showed restricted cellular barrier transport. The presented findings from this systematic investigation of brain pharmacokinetics of mitragynine and 7-hydroxymitragynine are essential for design and interpretation of in vivo experiments aiming to establish exposure-response relationship.

In Vivo Quantitative Understanding of PEGylated Liposome's Influence on Brain Delivery of Diphenhydramine.

Despite the promising features of liposomes as brain drug delivery vehicles, it remains uncertain how they influence the brain uptake in vivo. In order to gain a better fundamental understanding of the interaction between liposomes and the blood-brain barrier (BBB), it is indispensable to test if liposomes affect drugs with different BBB transport properties (active influx or efflux) differently. The aim of this study was to quantitatively evaluate how PEGylated (PEG) liposomes influence brain delivery of diphenhydramine (DPH), a drug with active influx at the BBB, in rats. The brain uptake of DPH after 30 min intravenous infusion of free DPH, PEG liposomal DPH, or free DPH + empty PEG liposomes was compared by determining the unbound DPH concentrations in brain interstitial fluid and plasma with microdialysis. Regular blood samples were taken to measure total DPH concentrations in plasma. Free DPH was actively taken up into the brain time-dependently, with higher uptake at early time points followed by an unbound brain-to-plasma exposure ratio ( Kp,uu) of 3.0. The encapsulation in PEG liposomes significantly decreased brain uptake of DPH, with a reduction of Kp,uu to 1.5 ( p < 0.05). When empty PEG liposomes were coadministered with free drug, DPH brain uptake had a tendency to decrease ( Kp,uu 2.3), and DPH was found to bind to the liposomes. This study showed that PEG liposomes decreased the brain delivery of DPH in a complex manner, contributing to the understanding of the intricate interactions between drug, liposomes, and the BBB.

Combined PET and microdialysis for in vivo estimation of drug blood-brain barrier transport and brain unbound concentrations.

Methods to investigate blood-brain barrier transport and pharmacologically active drug concentrations in the human brain are limited and data translation between species is challenging. Hence, there is a need to further develop the read-out of techniques like positron emission tomography (PET) for studying neuropharmacokinetics. PET has a high translational applicability from rodents to man and measures total drug concentrations in vivo. The aim of the present study was to investigate the possibility of translating total drug concentrations, acquired through PET, to unbound concentrations, resembling those measured in the interstitial fluid by microdialysis sampling. Simultaneous PET scanning and brain microdialysis sampling were performed in rats throughout a 60min infusion of [N-methyl-11C]oxycodone in combination with a therapeutic dose of oxycodone and during a 60min follow up period after the end of infusion. The oxycodone concentrations acquired with PET were converted into unbound concentrations by compensating for brain tissue binding and brain intracellular distribution, using the unbound volume of distribution in brain (Vu,brain), and were compared to microdialysis measurements of unbound concentrations. A good congruence between the methods was observed throughout the infusion. However, an accumulating divergence in the acquired PET and microdialysis data was apparent and became more pronounced during the elimination phase, most likely due to the passage of radioactive metabolites into the brain. In conclusion, the study showed that PET can be used to translate non-invasively measured total drug concentrations into unbound concentrations as long as the contribution of radiolabelled metabolites is minor or can be compensated for.

Quantitative Assessment of Drug Delivery to Tissues and Association with Phospholipidosis: A Case Study with Two Structurally Related Diamines in Development.

Drug induced phospholipidosis (PLD) may be observed in the preclinical phase of drug development and pose strategic questions. As lysosomes have a central role in pathogenesis of PLD, assessment of lysosomal concentrations is important for understanding the pharmacokinetic basis of PLD manifestation and forecast of potential clinical appearance. Herein we present a systematic approach to provide insight into tissue-specific PLD by evaluation of unbound intracellular and lysosomal (reflecting acidic organelles) concentrations of two structurally related diprotic amines, GRT1 and GRT2. Their intratissue distribution was assessed using brain and lung slice assays. GRT1 induced PLD both in vitro and in vivo. GRT1 showed a high intracellular accumulation that was more pronounced in the lung, but did not cause cerebral PLD due to its effective efflux at the blood-brain barrier. Compared to GRT1, GRT2 revealed higher interstitial fluid concentrations in lung and brain, but more than 30-fold lower lysosomal trapping capacity. No signs of PLD were seen with GRT2. The different profile of GRT2 relative to GRT1 is due to a structural change resulting in a reduced basicity of one amino group. Hence, by distinct chemical modifications, undesired lysosomal trapping can be separated from desired drug delivery into different organs. In summary, assessment of intracellular unbound concentrations was instrumental in delineating the intercompound and intertissue differences in PLD induction in vivo and could be applied for identification of potential lysosomotropic compounds in drug development.

Improved method for quantitative analysis of the cyclotide kalata B1 in plasma and brain homogenate.

This study provides a new method for quantifying the cyclotide kalata B1 in both plasma and brain homogenate. Cyclotides are ultra-stable peptides with three disulfide bonds that are interesting from a drug development perspective as they can be used as scaffolds. In this study we describe a new validated LC-MS/MS method with high sensitivity and specificity for kalata B1. The limit of quantification was 2 ng/mL in plasma and 5 ng/gmL in brain homogenate. The method was linear in the range 2-10,000 ng/mL for plasma and 5-2000 ng/g for brain. Liquid Chromatographic separation was performed on a HyPurity C18 column, 50 × 4.6 mm, 3 µm particle size. The method had inter- and intra-day precision and accuracy levels <15% and 12% respectively. Applying the method to in vivo plasma samples and brain homogenate samples from equilibrium dialysis yielded satisfying results and was able to describe the plasma pharmacokinetics and brain tissue binding of kalata B1. The described method is quick, reproducible and well suited to quantifying kalata B1 in biological matrices.

Exploring Factors Causing Low Brain Penetration of the Opioid Peptide DAMGO through Experimental Methods and Modeling.

To advance the development of peptide analogues for improved treatment of pain, we need to learn more about the blood-brain barrier transport of these substances. A low penetration into the brain, with an unbound brain to blood ratio, Kp,uu, of 0.08, is an important reason for the lack of effect of the enkephalin analogue DAMGO (H-Tyr-d-Ala-Gly-MePhe-Gly-ol) according to earlier findings. The aim of this study was to investigate the role of efflux transporters, metabolism in the brain, and/or elimination through interstitial fluid bulk flow for the brain exposure of DAMGO. The in vivo brain distribution of DAMGO was evaluated using microdialysis in the rat. Data were analyzed with population modeling which resulted in a clearance into the brain of 1.1 and an efflux clearance 14 µL/min/g_brain. The efflux clearance was thus much higher than the bulk flow known from the literature. Coadministration with the efflux transporter inhibitors cyclosporin A and elacridar in vivo did not affect Kp,uu. The permeability of DAMGO in the Caco-2 assay was very low, of the same size as mannitol. The efflux ratio was <2 and not influenced by cyclosporin A or elacridar. These results indicate that the well-known efflux transporters Pgp and Bcrp are not responsible for the higher efflux of DAMGO, which opens up for an important role of other transporters at the BBB.

In vivo Functional Evaluation of Increased Brain Delivery of the Opioid Peptide DAMGO by Glutathione-PEGylated Liposomes.

PURPOSE: The purpose of this study was to evaluate formulation factors causing improvement in brain delivery of a small peptide after encapsulation into a targeted nanocarrier in vivo. METHODS: The evaluation was performed in rats using microdialysis, which enabled continuous sampling of the released drug in both the brain (striatum) and blood. Uptake in brain could thereby be studied in terms of therapeutically active, released drug. RESULTS: We found that encapsulation of the peptide DAMGO in fast-releasing polyethylene glycol (PEG)ylated liposomes, either with or without the specific brain targeting ligand glutathione (GSH), doubled the uptake of DAMGO into the rat brain. The increased brain delivery was observed only when the drug was encapsulated into the liposomes, thus excluding any effects of the liposomes themselves on the blood-brain barrier integrity as a possible mechanism. The addition of a GSH coating on the liposomes did not result in an additional increase in DAMGO concentrations in the brain, in contrast to earlier studies on GSH coating. This may be caused by differences in the characteristics of the encapsulated compounds and the composition of the liposome formulations. CONCLUSIONS: We were able to show that encapsulation into PEGylated liposomes of a peptide with limited brain delivery could double the drug uptake into the brain without using a specific brain targeting ligand.

Intracerebral microdialysis in blood-brain barrier drug research with focus on nanodelivery.

Microdialysis has contributed significantly to advance the understanding of BBB transport of drugs and to reveal key aspects of BBB transport, including quantifying active efflux and active uptake. Microdialysis studies on pharmacokinetic-pharmacodynamic relationships have given in-depth understanding of the processes involved. Recently, nanodelivery to the brain has been investigated with microdialysis, contributing to nanodelivery science by giving quantitative information on the possible success of different delivery vehicles and how they are involved in BBB transport.

Molecular properties determining unbound intracellular and extracellular brain exposure of CNS drug candidates.

In the present work we sought to gain a mechanistic understanding of the physicochemical properties that influence the transport of unbound drug across the blood-brain barrier (BBB) as well as the intra- and extracellular drug exposure in the brain. Interpretable molecular descriptors that significantly contribute to the three key neuropharmacokinetic properties related to BBB drug transport (Kp,uu,brain), intracellular accumulation (Kp,uu,cell), and binding and distribution in the brain (Vu,brain) for a set of 40 compounds were identified using partial least-squares (PLS) analysis. The tailoring of drug properties for improved brain exposure includes decreasing the polarity and/or hydrogen bonding capacity. The design of CNS drug candidates with intracellular targets may benefit from an increase in basicity and/or the number of hydrogen bond donors. Applying this knowledge in drug discovery chemistry programs will allow designing compounds with more desirable CNS pharmacokinetic properties.

Mechanistic understanding of brain drug disposition to optimize the selection of potential neurotherapeutics in drug discovery.

PURPOSE: The current project was undertaken with the aim to propose and test an in-depth integrative analysis of neuropharmacokinetic (neuroPK) properties of new chemical entities (NCEs), thereby optimizing the routine of evaluation and selection of novel neurotherapeutics. METHODS: Forty compounds covering a wide range of physicochemical properties and various CNS targets were investigated. The combinatory mapping approach was used for the assessment of the extent of blood-brain and cellular barriers transport via estimation of unbound-compound brain (Kp,uu,brain) and cell (Kp,uu,cell) partitioning coefficients. Intra-brain distribution was evaluated using the brain slice method. Intra- and sub-cellular distribution was estimated via calculation of unbound-drug cytosolic and lysosomal partitioning coefficients. RESULTS: Assessment of Kp,uu,brain revealed extensive variability in the brain penetration properties across compounds, with a prevalence of compounds actively effluxed at the blood-brain barrier. Kp,uu,cell was valuable for identification of compounds with a tendency to accumulate intracellularly. Prediction of cytosolic and lysosomal partitioning provided insight into the subcellular accumulation. Integration of the neuroPK parameters with pharmacodynamic readouts demonstrated the value of the proposed approach in the evaluation of target engagement and NCE selection. CONCLUSIONS: With the rather easily-performed combinatory mapping approach, it was possible to provide quantitative information supporting the decision making in the drug discovery setting.

Analysis of the contributing role of drug transport across biological barriers in the development and treatment of chemotherapy-induced peripheral neuropathy.

BACKGROUND: Chemotherapy-induced peripheral neuropathy (CIPN) represents a major unmet medical need that currently has no preventive and/or curative treatment. This is, among others, driven by a poor understanding of the contributive role of drug transport across biological barriers to target-site exposure. METHODS: Here, we systematically investigated the transport of 11 small-molecule drugs, both, associated and not with CIPN development, at conventional (dorsal root ganglia, sciatic nerve) and non-conventional (brain, spinal cord, skeletal muscle) CIPN sites. We developed a Combinatory Mapping Approach for CIPN, CMA-CIPN, combining in vivo and in vitro elements. RESULTS: Using CMA-CIPN, we determined the unbound tissue-to-plasma concentration ratio (Kp,uu) and the unbound intracellular-to-extracellular concentration ratio (Kp,uu,cell), to quantitatively assess the extent of unbound drug transport across endothelial interfaces and parenchymal cellular barriers of investigated CIPN-sites, respectively, in a rat model. The analysis revealed that unique pharmacokinetic characteristics underly time-dependent accumulation of the CIPN-positive drugs paclitaxel and vincristine at conventional (dorsal root ganglia and sciatic nerve) and non-conventional (skeletal muscle) CIPN sites. Investigated CIPN-positive drugs displayed intracellular accumulation contrary to CIPN-negative drugs nilotinib and methotrexate, which lacked this feature in all investigated tissues. CONCLUSIONS: Hence, high unbound drug intracellular and extracellular exposure at target sites, driven by an interplay of drug transport across the endothelial and parenchymal cellular barriers, is a predisposing factor to CIPN development for CIPN-positive drugs. Critical drug-specific features of unbound drug disposition at various CIPN- sites provide invaluable insights into understanding the pharmacological/toxicological effects at the target-sites which will inform new strategies for monitoring and treatment of CIPN.

Chain length of dietary alkylresorcinols affects their in vivo elimination kinetics in rats.

Two phenolic acids, 3,5-dihydroxybenzoic acid (DHBA) and 3-(3,5-dihydroxyphenyl)- propanoic acid (DHPPA), are the major metabolites of cereal alkylresorcinols (ARs). Like their precursors, AR metabolites have been suggested as biomarkers for intake of whole-grain wheat and rye and as such could aid the understanding of diet-disease associations. This study estimated and compared pharmacokinetic parameters of ARs and their metabolites in rats and investigated differences in metabolite formation after ingestion of different AR homologs. Rats were i.v. infused for 30 min with 2, 12, or 23 µmol/kg DHBA or DHPPA or orally given the same amounts of the AR homologs, C17:0 and C25:0. Repeated plasma samples, obtained from rats for 6 h (i.v.) or 36 h (oral), were simultaneously analyzed for ARs and their metabolites by GC-mass spectrometry. Pharmacokinetic parameters were estimated by population-based compartmental modeling and noncompartmental calculation. A 1-compartment model best described C25:0 pharmacokinetics, whereas C17:0 and AR metabolites best fitted 2-compartment models. Combined models for simultaneous prediction of AR and metabolite concentration were more complex, with less reliable estimates of pharmacokinetic parameters. Although the AUC of C17:0 was lower than that of C25:0 (P < 0.05), the total amount and composition of AR metabolites did not differ between rats given C17:0 or C25:0. The elimination half-life of ARs and their metabolites increased with length of the side chain (P-trend < 0.001) and ranged from 1.2 h (DHBA) to 8.8 h (C25:0). The formation of AR metabolites was slower than their elimination, indicating that the rate of AR metabolism and not excretion of DHBA and DHPPA determines their plasma concentrations in rats.

Enhanced brain delivery of the opioid peptide DAMGO in glutathione pegylated liposomes: a microdialysis study.

Glutathione PEGylated (GSH-PEG) liposomes were evaluated for their ability to enhance and prolong blood-to-brain drug delivery of the opioid peptide DAMGO (H-Tyr-d-Ala-Gly-MePhe-Gly-ol). An intravenous loading dose of DAMGO followed by a 2 h constant rate infusion was administered to rats, and after a washout period of 1 h, GSH-PEG liposomal DAMGO was administered using a similar dosing regimen. DAMGO and GSH-PEG liposomal DAMGO were also administered as a 10 min infusion to compare the disposition of the two formulations. Microdialysis made it possible to determine free DAMGO in brain and plasma, while the GSH-PEG liposomal encapsulated DAMGO was measured with regular plasma sampling. The antinociceptive effect of DAMGO was determined with the tail-flick method. All samples were analyzed using liquid chromatography-tandem mass spectrometry. The short infusion of DAMGO resulted in a fast decline of the peptide concentration in plasma with a half-life of 9.2 ± 2.1 min. Encapsulation in GSH-PEG liposomes prolonged the half-life to 6.9 ± 2.3 h. Free DAMGO entered the brain to a limited extent with a steady state ratio between unbound drug concentrations in brain interstitial fluid and in blood (Kp,uu) of 0.09 ± 0.04. GSH-PEG liposomes significantly increased the brain exposure of DAMGO to a Kp,uu of 0.21 ± 0.17 (p < 0.05). By monitoring the released, active substance in both blood and brain interstitial fluid over time, we were able to demonstrate that GSH-PEG liposomes offer a promising platform for enhancing and prolonging the delivery of drugs to the brain.

Differential Blood-Brain Barrier Transport and Cell Uptake of Cyclic Peptides In Vivo and In Vitro.

The blood-brain barrier (BBB) poses major challenges to drug delivery to the CNS. SFTI-1 and kalata B1 are cyclic cell-penetrating peptides (cCPPs) with high potential to be used as scaffolds for drug delivery. We here studied their transport across the BBB and distribution within the brain to gauge the potential of these two cCPPs as scaffolds for CNS drugs. In a rat model, SFTI-1 exhibited, for a peptide, high extent of BBB transport with a partitioning of unbound SFTI-1 across the BBB, Kp,uu,brain, of 13%, while only 0.5% of kalata B1 equilibrated across the BBB. By contrast, kalata B1, but not SFTI-1, readily entered neural cells. SFTI-1, but not kalata B1, could be a potential CNS delivery scaffold for drugs directed to extracellular targets. These findings indicate that differences between the BBB transport and cellular uptake abilities of CPPs are crucial in the development of peptide scaffolds.