Brain-specific Delivery Research Articles

A critical evaluation of drug delivery from ligand modified nanoparticles: Confounding small molecule distribution and efficacy in the central nervous system

In this work, we sought to test how surface modification of poly(lactic-co-glycolic acid) (PLGA) nanoparticles with peptide ligand alters the brain specific delivery of encapsulated molecules. For biodistribution studies, nanoparticles modified with rabies virus glycoprotein (RVG29) were loaded with small molecule drug surrogates and administered to healthy mice by lateral tail vein injection. Mice were perfused 2h after injection and major anatomical regions of the CNS were dissected (striatum, midbrain, cerebellum, hippocampus, cortex, olfactory bulb, brainstem, and cervical, thoracic, lumbar and sacral spinal cord). For functional studies, surface modified nanoparticles were loaded with the chemotherapeutic camptothecin (CPT) and administered to mice bearing intracranial GL261-Luc2 gliomas. Outcome measures included tumor growth, as measured by bioluminescent imaging, and median survival time. We observed that small molecule delivery from PLGA nanoparticles varied by as much as 150% for different tissue regions within the CNS. These differences were directly correlated to regional differences in cerebral blood volume. Although the presence of RVG29 enhanced apparent brain delivery for multiple small molecule payloads, we observed minimal evidence for targeting to muscle or spinal cord, which are the known sites for rabies virus entry into the CNS, and enhancements in brain delivery were not prolonged due to an apparent aqueous instability of the RVG29 ligand. Furthermore, we have identified concerning differences in apparent delivery kinetics as measured by different payloads: nanoparticle encapsulated DiR was observed to accumulate in the brain, whereas encapsulated Nile red was rapidly cleared. Although systemically administered CPT loaded nanoparticles slowed the growth of orthotopic brain tumors to prolong survival, the presence of RVG29 did not enhance therapeutic efficacy compared to control nanoparticles. These data are consistent with a model of delivery of hydrophobic small molecules to the brain that does not rely on internalization of polymer nanoparticles in target tissue. We discuss an important risk for discordance between biodistribution, as typically measured by drug surrogate, and therapeutic outcome, as determined by clinically relevant measurement of drug function in a disease model. These results pose critical considerations for the methods used to design and evaluate targeted drug delivery systems in vivo.

Targeted Brain Delivery of Bioactive Molecules Using Nanocarriers

Delivery of the bioactives to the brain is the utmost challenging task to cope with brain diseases. Brain is protected with blood brain barrier, blood-CSF barrier and efflux systems, which controls the entry of body as well as foreign compounds to access the brain cells. Only nutrients, which are essential for normal metabolism, can enter into the brain. In the shadow of this fact new strategies are being investigated to facilitate the entry of administered therapeutic compound into the brain. Active targeting is a evolving approach, which uses ligand and suitable carrier for the site-specific delivery and is being recently achieved by the use of nanocarriers. These nanocarriers are nanosized systems, which act as a cargo for the encapsulated drugs. At the same time, endo- or exogenous ligand can be attached to these nanocarriers to recognize specific receptors on brain capillary endothelium leading to delivery of drug in the vicinity of brain cells. Their potential is under immense investigation to increase the therapeutics outcome in the treatment of brain related problems. This review deals with the recent advances in nanocarriers based novel strategies for effective brain specific delivery.

Cerebral hypoperfusion-assisted intra-arterial deposition of liposomes in normal and glioma-bearing rats.

Optimizing liposomal vehicles for targeted delivery to the brain has important implications for the treatment of brain tumors. The promise of efficient, brain-specific delivery of chemotherapeutic compounds via liposomal vehicles has yet to be achieved in clinical practice. Intra-arterial injection of specially designed liposomes may facilitate efficient delivery to the brain and to gliomas. To test the hypothesis that cationic liposomes may be effectively delivered to both normal and glioma-bearing brain tissue utilizing a strategy of intra-arterial injection during transient cerebral hypoperfusion. Cationic, anionic, and neutral liposomes were separately injected via the internal carotid artery of healthy rats during transient cerebral hypoperfusion. Rats bearing C6 gliomas were similarly injected with cationic liposomes. Liposomes were loaded with DilC18(5) dye whose concentrations can be measured by light absorbance and fluorescence methods. After intra-arterial injection, a robust uptake of cationic in comparison with anionic and neutral liposomes into brain parenchyma was observed by diffuse reflectance spectroscopy. Postmortem multispectral fluorescence imaging revealed that liposomal cationic charge was associated with more efficient delivery to the brain. Cationic liposomes were also readily observed within glioma tissue after intra-arterial injection. However, over time, cationic liposomes were retained longer and at higher concentrations in the surrounding, peritumoral brain than in the tumor core. This study demonstrates the feasibility of cationic liposome delivery to brain and glioma tissue after intra-arterial injection. Highly cationic liposomes directly delivered to the brain via an intracarotid route may represent an effective method for delivering antiglioma agents.

Pathogen-inspired drug delivery to the central nervous system

For as long as the human blood-brain barrier (BBB) has been evolving to exclude bloodborne agents from the central nervous system (CNS), pathogens have adopted a multitude of strategies to bypass it. Some pathogens, notably viruses and certain bacteria, enter the CNS in whole form, achieving direct physical passage through endothelial or neuronal cells to infect the brain. Other pathogens, including bacteria and multicellular eukaryotic organisms, secrete toxins that preferentially interact with specific cell types to exert a broad range of biological effects on peripheral and central neurons. In this review, we will discuss the directed mechanisms that viruses, bacteria, and the toxins secreted by higher order organisms use to enter the CNS. Our goal is to identify ligand-mediated strategies that could be used to improve the brain-specific delivery of engineered nanocarriers, including polymers, lipids, biologically sourced materials, and imaging agents.

Brain-specific delivery of dopamine mediated by n,n-dimethyl amino group for the treatment of Parkinson's disease.

Parkinson's disease (PD) has become one of the most deadly diseases due to a lack of effective treatment. Herein, N-3,4-bis(pivaloyloxy)dopamine-3-(dimethylamino)propanamide (PDDP), a brain-specific derivative of dopamine, was designed and synthesized, which consists of a brain targeted ligand, N,N-dimethyl amino group, and two dipivaloyloxy groups for lipophilic modification. PDDP was investigated both in vitro and in vivo by comparing with L-DOPA and another derivative (BPD) without N,N-dimethyl amino group. PDDP showed a more pronounced accumulation in mouse brain microvascular endothelial cells (bEnd.3) than BPD via an active transport process. The increased cellular uptake of PDDP was proven to be mediated by putative pyrilamine cationic transporters. Following intravenous administration, the concentration of PDDP in the brain was 269.28-fold and 6.41-fold higher than that of L-DOPA and BPD at 5 min, respectively. Additionally, PDDP effectively attenuated the striatum lesion caused by 6-hydroxydopamine (6-OHDA) in rats. More importantly, PDDP presented antioxidant and antiapoptotic effects on 6-OHDA-induced toxicity in human neuroblastoma cells (SH-SY5Y). Thus, N,N-dimethyl amino group-based PDDP represents an effective and safe treatment for PD.

Polymersomes conjugated with des-octanoyl ghrelin for the delivery of therapeutic and imaging agents into brain tissues

The effective protection of the blood–brain barrier (BBB) from tight junctions and efflux transport systems ultimately results in the limited entry of 95% of drug/gene candidates, which are potentially beneficial for central nervous system (CNS) diseases. In order to enhance the brain-specific delivery, in this study we developed a targeting carrier system, which consists of poly(carboxyl ethylene glycol-g-glutamate)-co-poly(distearin-g-glutamate) (CPEGGM-PDSGM) polymersomes with the conjugation of des-octanoyl ghrelin. Des-octanoyl ghrelin across the BBB was reported to be unidirectional (blood-to-brain direction). However, there is no report about the conjugation of des-octanoyl ghrelin to a drug carrier system to confer the BBB targeting property through des-octanoyl ghrelin binding sites mediated endocytosis. To qualitatively and quantitatively investigate this carrier's properties, coumarin 6, Cy5.5 and met-enkephalin were individually encapsulated in these polymersomes. The experimental results showed that the cellular uptake was significantly higher for des-octanoyl ghrelin-conjugated polymersomes (GPs) than unconjugated polymersomes when co-incubated with the BBB cells. In addition, an enhanced accumulation in brain together with a reduced accumulation in liver and spleen was observed in animal study, indicating better brain selectivity for the GPs. In a hot-plate test, a significant inhibition of nociceptive response could be achieved for an intravenous injection of GPs encapsulated with met-enkephalin. The overall results demonstrated that GPs own a great potential for targeting delivery of drug across the BBB to treat CNS diseases.

Tween 80 containing lipid nanoemulsions for delivery of indinavir to brain

Indinavir is a protease inhibitor used in the treatment of HIV infection. However, it has limited efficacy in eradicating the virus in the brain due to efflux by P-glycoprotein (P-gp) expressed at the blood–brain barrier (BBB). The objective of this work was to develop an o/w lipid nanoemulsion (LNE) of indinavir using Tween 80 as co-emulsifier to improve its brain specific delivery. LNEs were prepared with different compositions and were characterized for globule size, polydispersity index, zeta potential and in vitro drug release. Five formulations were then evaluated for drug content, entrapment efficiency and stability after which brain uptake studies were carried out using fluorescent labeled LNEs and pharmacokinetic (PK) and tissue distribution studies were conducted after intravenous administration in mice. Brain uptake of indinavir was shown to be improved for a 1% Tween 80 containing formulation (F5) compared to a formulation containing 0.3% cholesterol (F2). In PK studies, the brain level of indinavir subsequent to administration of F5 was significantly (P<0.05) higher than produced by administration of a drug solution (2.44-fold) or a control nanoemulsion (F1) (1.48-fold) or formulation F2 (1.6-fold). The increased brain specific accumulation of indinavir from F5 is probably due to enhanced low density lipoprotein-mediated endocytosis and P-gp inhibition by Tween 80 at the BBB. These results suggest Tween 80 containing LNEs could provide a simple but effective means of delivering indinavir to brain.

Brain delivery of transferrin coupled indinavir submicron lipid emulsions—Pharmacokinetics and tissue distribution

Indinavir, an antiretroviral protease inhibitor used in treatment of HIV infection has limited penetration into brain due to efflux of P-glycoprotein. The aim of this work was to develop transferrin coupled submicron lipid emulsions (SLEs) containing indinavir for delivery to brain. Stearylamine containing SLEs were prepared, characterized, tested for stability and optimized formulation (SLE-4) was developed. Transferrin was coupled to get SLE-6 by water soluble EDC method and purified by gel filtration. The coupled transferrin was quantified by modified Bradford dye assay method. The fluorescent dye (DiD oil) incorporated SLEs were used to check the brain specific delivery of SLEs. The in vivo pharmacokinetic and tissue distribution were conducted in mice. During pharmacokinetic studies, there was no significant difference in the serum levels of indinavir from SLE-1, SLE-4 and SLE-6 formulations at all time points. In tissue distribution studies the therapeutic availability (TA) of indinavir in brain from SLE-6 was 4.69, 3.1 and 1.7 times higher than drug solution, SLE-1 and SLE-4 respectively whereas, the TA of indinavir from SLE-4 was 2.76 and 1.82 times the drug solution and SLE-1. The brain to serum ratios with SLE-6 were above one indicates the brain specific delivery. The brain delivery of indinavir was improved with transferrin ligand attachment to SLEs by receptor mediated transcytosis.

Development of Indinavir Submicron Lipid Emulsions Loaded with Lipoamino Acids—In Vivo Pharmacokinetics and Brain-Specific Delivery

The aim of our present work was to develop indinavir O/W submicron lipid emulsions (SLEs) loaded with lipoamino acids for specific delivery to brain. Tetradecyl aspartic acid (A) and decyl glutamic acid (G) loaded stable SLEs of indinavir having a mean size range of 210-220 nm and average zeta potential of -23.54±1.2 mV were developed using homogenization and ultrasonication. The cumulative % drug release from different SLEs varied in between 26% and 85%. The formulations, SLE, SLE-A3, and SLE-G3 were stable to the centrifugal stress, dilution stress, and storage at RT. The total drug content and entrapment efficiency were determined by HPLC method. During pharmacokinetic studies in male Wistar rats there was no significant difference in the serum levels of indinavir for SLE, SLE-A3 and SLE-G3 formulations at all time points. In tissue distribution studies, the therapeutic availability (TA) of indinavir in brain and kidneys for SLE-A3 were 4.27- and 2.66-fold whereas for SLE-G3 were 2.94 and 2.12 times, respectively, higher than that of indinavir solution. But when compared with that of SLE, in brain tissue the levels of indinavir from SLE-G3 and SLE-A3 varied in between 2.5- and 3.38-fold. While in case of the kidney, it was between 1.23- and 1.54-fold only. However, the TA is not significantly different in tissues like the heart, liver, and spleen. Thus, brain-specific delivery of indinavir was improved by including tetradecyl aspartic acid and decyl glutamic acid in submicron lipid emulsions.

Brain specific delivery of pegylated indinavir submicron lipid emulsions

The aim of this study was to develop stable parenteral pegylated indinavir submicron lipid emulsions (SLEs) for improving brain specific delivery. The O/W SLEs were prepared by homogenization and ultra sonication process. The sizes of oil globules varied from 241.5 to 296.4 nm and zeta potential from −26.6 to −42.4 mV. During in vitro drug release studies the cumulative amount of drug released within 12 h from SLE-5, DSP2-3 and DPP5-3 was 71.8 ± 0.76, 66.09 ± 1.45 and 68.33 ± 1.29, respectively. The total drug content and entrapment efficiencies were determined. The optimized formulations were stable for the effect of centrifugal stress, thermal stress, dilution stress and storage. In vivo pharmacokinetic and tissue distribution studies were performed in Swiss albino mice, the therapeutic availability (TA) of DSP2-3 was 3.59 times and 2.36 times in comparison to drug solution and SLE-5 respectively, where as DPP5-3 showed TA 2.8 and 1.84 times the drug solution and SLE-5, respectively. The brain to serum ratio of indinavir from DSP2-3 and DPP5-3 varied between 0.4 and 0.7 at all time points indicated the preferential accumulation of drug in brain. In conclusion, pegylated SLEs improved brain specific delivery of indinavir and will be useful in treating chronic HIV infection.

Transferrin-appended PEGylated nanoparticles for temozolomide delivery to brain: in vitro characterisation

Polymer-based nanotechnologies are proposed to be an alternative for drug administration, delivery and targeting to those of conventional formulations. The blood brain barrier is frequently a rate-limiting factor in determining permeation of a drug into brain. In this study, the surface-engineered long-circulating PLGA nanoparticles (NPs) were assessed for brain-specific delivery. Long circulating NPs of PLGA- and PEG-synthesised copolymer were prepared by emulsification solvent evaporation method. Further, the surface of PEGylated NPs was modified by anchoring transferrin (Tf) ligand for receptor-mediated targeting to brain. NPs were characterised for shape and size, zeta potential, entrapment efficiency and in vitro drug release. In vitro cytotoxicity studies were performed on human cancer cell lines. Confocal Laser Scanning Microscopy studies show the enhanced uptake of Tf-appended PEGylated NPs and their localisation in the brain tissues. Hence, the specific role of Tf ligand on PEGylated NPs for brain delivery was confirmed.

Brain‐Specific Delivery of Naproxen Using Different Carrier Systems

Naproxen is one of the most potent NSAIDs and plays an important role in the treatment of neurodegenerative diseases. Poor brain delivery of naproxen at therapeutic doses, in addition to its serious gastrointestinal side effects, has prompted research into the development of a specific carrier system that is capable of delivering naproxen to the brain at smaller doses. The purpose of this study was to evaluate two brain-specific carrier systems of naproxen. The first was the dihydropyridine/pyridinium redox system that utilized a lipophilic chemical delivery system coupled to the carboxylic acid group of naproxen through an ethanolamine linker. Secondly, an ascorbic acid system, which has reducing properties and acts as a biological carrier through sodium-dependent vitamin-C transporter, was used for brain-specific delivery of naproxen. The prepared prodrugs were stable in aqueous buffers (pH 1.2 and 7.4) and rapidly hydrolyzed in biological fluids. Bioavailability studies revealed that both prodrugs 10 and 17 were rapidly cleared from blood with half lives of about 1 h, which will likely decrease systemic adverse effects. The rapid clearance from the blood was accompanied by an increase in the prodrug concentration in the brain, which occurred as a result of the prodrug being more locked in compared to the parent drug naproxen.

Identification of Peptide Ligands for Targeting to the Blood-Brain Barrier

PurposeTransport of drugs to the brain is limited by the blood-brain barrier. New, specific brain endothelium ligands can facilitate brain-specific delivery of drugs.MethodsWe used phage display in an in situ brain perfusion model to screen for new brain endothelium peptide ligands.ResultsTwo phage clones, displaying 15 amino acid-peptides (GLA and GYR) that were selected for brain binding in the mouse model, showed significant binding to human brain endothelium (hCMEC/D3), compared to a random control phage. This binding was not seen for other human endothelial cells (HUVEC). Binding to hCMEC/D3 cells was dose dependent. When phage GLA and GYR were individually perfused through the murine brain, their ability to bind to the brain was 6-fold (GLA) and 5-fold (GYR) higher than the control phage. When compared to lung perfusion, phage showed an 8.5-fold (GYR) and 48-fold (GLA) preference for brain over lung compared to the control.ConclusionsThese results indicate that two new peptide ligands have been identified that may be used for specific targeting of drugs to the blood-brain barrier.

Enhanced Brain Targeting of Tegafur Using Novel Lactyl Cholesterol Liposome as a Carrier

A series of lactyl cholesterols as targeting liposome ligands for brain-specific delivery were synthesized. The lactic acid protected by benzyl group was coupled with cholesteryl oligoethylene carboxylate derivatives 3, then deprotected in tetrahydrofuran by Pd/C with H2 to obtain the title compounds La-Lf. As a model drug, tegafur was entrapped by liposmes and lipsomes coated with these ligands for in vivo study, which showed that the ligand could enhance the ability of liposome delivering tegafur across the blood-brain barrier. Keywords: Liposome, Liposome ligands, Monocarboxylic acid transport, Tegafur, Blood-brain barrier, Brain targeting

Nanotechnology approach for drug addiction therapy: Gene silencing using delivery of gold nanorod-siRNA nanoplex in dopaminergic neurons

Drug abuse is a worldwide health concern in which addiction involves activation of the dopaminergic signaling pathway in the brain. Here, we introduce a nanotechnology approach that utilizes gold nanorod-DARPP-32 siRNA complexes (nanoplexes) that target this dopaminergic signaling pathway in the brain. The shift in the localized longitudinal plasmon resonance peak of gold nanorods (GNRs) was used to show their interaction with siRNA. Plasmonic enhanced dark field imaging was used to visualize the uptake of these nanoplexes in dopaminergic neurons in vitro. Gene silencing of the nanoplexes in these cells was evidenced by the reduction in the expression of key proteins (DARPP-32, ERK, and PP-1) belonging to this pathway, with no observed cytotoxicity. Moreover, these nanoplexes were shown to transmigrate across an in vitro model of the blood-brain barrier (BBB). Therefore, these nanoplexes appear to be suited for brain-specific delivery of appropriate siRNA for therapy of drug addiction and other brain diseases.

Molecular Pathologies of and Enzyme Replacement Therapies for Lysosomal Diseases

Lysosomal diseases comprise a group of inherited disorders resulting from defects of lysosomal enzymes and their cofactors, and in many of them the nervous system is affected. Recently, enzyme replacement therapy with recombinant lysosomal enzymes has been clinically available for several lysosomal diseases. Such enzyme replacement therapies can improve non-neurological disorders but is not effective for neurological ones. In this review, we discuss the molecular pathologies of lysosomal diseases from the protein structural aspect, current enzyme replacement therapies, and attempts to develop enzyme replacement therapies effective for lysosomal diseases associated with neurological disorders, i.e., production of enzymes, brain-specific delivery and incorporation of lysosomal enzymes into cells.

Targeted brain delivery of AZT via transferrin anchored pegylated albumin nanoparticles

Hydrophilic drugs/peptides have poor cross Blood–brain permeability. Various drug delivery systems with diverse surfacial characteristics have been reported for effective translocation of drugs across Blood–brain barrier. In present investigation, the potential of engineered albumin nanoparticles was evaluated for brain specific delivery after intravenous administration. Long circulatory PEGylated albumin nanoparticles encapsulating water-soluble antiviral drug azidothymidine (AZT) were prepared by ultra-emulsification method using chemical cross-linking by glutaraldehyde. Surface of the PEGylated nanoparticles was modified by anchoring transferrin as a ligand for brain targeting. Nanoparticles were characterized for their size, polydispersity, surfacial charge, drug loading and in vitro drug release. Fluorescence studies revealed the enhanced uptake of transferrin-anchored nanoparticles in the brain tissues when compared with unmodified nanoparticles. In vivo evaluation was carried out on albino rats to evaluate tissue distribution of engineered nanoparticles after intravenous administration. A significant (*P < 0.01) enhancement of brain localization of AZT was observed for transferrin anchored pegylated albumin nanopariticles (Tf-PEG-NPs). Hence, the specific role of transferrin ligand on nanoparticles for brain targeting was confirmed.

1, 2-dihydroisoquinoline-N-acetic acid derivatives as new carriers for brain-specific delivery II: delivery of phenethylamine as model drug.

N-alkyloxycarbonylmethyl-1, 2-dihydroisoquinolin-4-carboxylic acid derivatives 7a-c synthesized as new carriers for brain specific delivery. The design of the carrier systems are based on sequential hydrolysis at the acetic acid ester group linked to dihydroisoquinoline nitrogen followed by ring oxidation and formation of quaternary isoquinolinium derivatives which are then hydrolyzed to release the drug. Once the carrier system is administered, a sequential enzymatic process will take place resulting in significant increase in its rate of oxidation, the key factor in brain specific delivery. The chemical stability of the synthesized carrier system was investigated in aqueous buffer solutions and ferricyanide reagent and proofed to be quite stable against hydration and oxidation during formulation and storage. Furthermore, enzymatic stability was also investigated in 80 % human plasma and 20 % rabbit brain homogenate. Both oxidation and hydrolysis were found to take place; however, hydrolysis was the major route. In vivo distribution of the ethyl ester derivative 7b studied in rats and showed that the concentration of the quaternary product is increasing in the brain and cleared from blood with time.

New Carrier for Specific Delivery of Drugs to the Brain

1,4-Dihydropyridines were thoroughly investigated as carriers for specific drug delivery to the brain and were found very efficient. The main problem which arises in their application in pharmaceutical preparations, is the short shelf-life time due to hydration and/or oxidation. To overcome these problems, a new carrier system is suggested. Many of 1,4-dihydropyridine-3,5-dicarboxylate derivatives are used as calcium channel blockers which are known to have long shelf-life time, and at the same time, they are safe, with no reported neurotoxicity. Since efficiency of brain specific delivery depends on the rate of oxidation of the dihydropyridine carrier, (the faster the rate, the higher the efficiency), these 3,5-dicarbonyl compounds have almost of no brain-specific delivery properties. N-alkoxycarbonylmethyl derivatives of 1,4-dihydropyridine-3,5-dicarboxylate, a new carrier system is suggested and expected to be stable enough for formulation and storage. The drug moiety will be connected to the 3,5-dicarbonyl groups in the form of esters or amides. The suggested drug-carrier once delivered and distributed in the body, will be subjected to several sequential enzymatic hydrolytic and oxidative processes. The D-carrier is designed so that, the rate of hydrolysis of the ester group at the nitrogen atom should be faster than that of the hydrolysis and release of the drug moiety. The stability and efficiency of brain specific delivery of model drugs were investigated. The in vitro and in vivo studies proved the efficiency of brain specific delivery of the carrier and at the same time its shelf life stability. The results suggest that the designed carrier is promising to be used in application of pharmaceutical formulation.

In Vitro and In Vivo Evaluations of Dihydroquinoline- and Dihydroisoquinoline-based Targetor Moieties for Brain-specific Chemical Delivery Systems

Brain-targeted delivery of various drugs can be successfully achieved by chemical delivery systems (CDS) that contain a 1,4-dihydropyridine-based redox targetor moiety and undergo a sequential metabolism. However, the susceptibility of this moiety toward hydration in acidic media may limit the shelf-life of such compounds in aqueous formulation. Here, a systematic investigation of the chemical stability toward oxidation and hydration of ester and amide derivatives of 3-substituted 1,4-dihydropyridine, 1,4-dihydroquinoline, and 4-substituted 1,2-dihydroisoquinoline is reported, together with the in vitro stability and in vivo (rat) distribution of isoquinoline-based testosterone and hydrocortisone chemical delivery systems, which were selected as having the most suitable acid-resistant targetor moieties.