How well do liposomes cross the BBB? Some abstracts...

[Guest guest]

I read the posts Sue B. pointed us to on the CFSMEexperimental list.

Very exciting! But I was left unsure exactly how much closer we were

to knowing about BBB penetration by lipoceutical glutathione,

because the example related in the conference call had to do with a

tumor and in some cases brain tumors are associated with a

compromised BBB.

The fourth reference below specifies this:

" Being incapable of penetrating the unimpaired blood-brain barrier

(BBB), these immunoliposomes, nevertheless, may be useful in

delivering drugs to glial brain tumors (which continue to express

GFAP) or to other pathological loci in the brain with a partially

disintegrated BBB. "

On the other hand, at least one patient I know with heavy

neurological symptoms and MRI confirmed brain pathology reported

doing well on lipoceutical glutathione, and while they're too

technical in their language for me to draw any clear conclusion

from, several of the abstracts below seem encouraging.

Rich, any additional thoughts?

ABSTRACTS FROM PUB MED:

1: Int J Pharm. 2005 Jul 25;298(2):274-92. Related Articles, Links

Colloidal carriers and blood-brain barrier (BBB) translocation: A

way to deliver drugs to the brain?

- E, Andrieux K, Gil S, Couvreur P.

Laboratory of Pharmaceutical Technology and Biopharmacy, UMR CNRS

8612, Faculty of Pharmacy, University of Paris-XI, 92296 Chatenay-

Malabry, France.

The major problem in drug delivery to the brain is the presence of

the blood-brain barrier (BBB) which limits drug penetration even if

in certain pathological situations the BBB is partly disrupted.

Therefore, various strategies have been proposed to improve the

delivery of drugs to this tissue. This review presents the status of

the BBB in healthy patients and in pathologies like

neurodegenerative, cerebrovascular and inflammatory diseases. The

second part of this article aims to review the invasive and non-

invasive strategies developed to circumvent the BBB and deliver

drugs into the brain. The use of nanotechnologies (liposomes,

nanoparticles) is especially discussed in the ultimate part of the

review evidencing their potentiality as non-invasive technique in

the brain delivery of drugs with the possibility to target specific

brain tissue thanks to ligand linked to carrier surface.

PMID: 15896933 [PubMed - in process]

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2: J Control Release. 2005 Jun 20;105(1-2):106-19. Related Articles,

Links

Transport of nerve growth factor encapsulated into liposomes across

the blood-brain barrier: in vitro and in vivo studies.

Xie Y, Ye L, Zhang X, Cui W, Lou J, Nagai T, Hou X.

Department of Physical Pharmacy, School of Pharmaceutical Science,

Xueyuan Road #38, Peking University, Beijing 100083, PR China.

A nerve growth factor (NGF) was encapsulated into liposomes in order

to protect it from the enzyme degradation in vivo and promote it

permeability across the blood-brain barrier (BBB). RMP-7, a ligand

to the B2 receptor on brain microvascular endothelial cells (BMVEC),

was combined with 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-n-

[poly(ethylenegly-col)]-hydroxy succinamide (DSPE-PEG-NHS) to obtain

DSPE-PEG-RMP-7. Then DSPE-PEG-RMP-7 was incorporated into the

liposomes' surface to target sterically stabilized liposomes (SSL-T)

to the brain. The highest percent of NGF encapsulated into liposomes

was about 34%, and the average size of liposomes was below 100 nm. A

primary model of BBB was established and evaluated by morphological,

permeability, and transendothelial electrical resistance (TEER). The

BBB model was employed to study the permeability of NGF liposomes in

vitro. The results indicated that the liposomes could enhance

transport of NGF across the BBB. The best transport rate was

received with NGF-SSL-T. The brain distribution of NGF liposomes was

studied in vivo, the amount of NGF in the brain was increased in the

order: NGF-SSL-T>NGF-SSL+RMP-7>NGF-SSL>NGF-L. The maximum

concentration of NGF was recorded in 30 min following the

intravenous injection. In particular, a majority of NGF was

distributed in striatum, hippocampus and cortex, and the

concentration of NGF was relatively lower in olfactory bulb,

cerebellum and brain stem. There was a close relationship between P

(e) (permeability coefficient on in vitro BBB model) and T(e) (brain

targeted coefficient in vivo) for NGF encapsulated into the

liposomes.

PMID: 15893839 [PubMed - in process]

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3: Eur J Pharm Sci. 2005 Jun;25(2-3):299-305. Epub 2005 Apr 18.

Related Articles, Links

Targeting liposomes with protein drugs to the blood-brain barrier in

vitro.

Visser CC, Stevanovic S, Voorwinden LH, van Bloois L, Gaillard PJ,

Danhof M, Crommelin DJ, de Boer AG.

Leiden/Amsterdam Centre for Drug Research (LACDR), Leiden

University, Division of Pharmacology, PO Box 9502, 2300 RA Leiden,

The Netherlands.

In this study, we aim to target pegylated liposomes loaded with

horseradish peroxidase (HRP) and tagged with transferrin (Tf) to the

BBB in vitro. Liposomes were prepared with the post-insertion

technique: micelles of polyethylene glycol (PEG) and PEG-Tf were

inserted into pre-formed liposomes containing HRP. Tf was measured

indirectly by measuring iron via atomic absorption spectroscopy. All

liposomes were around 100 nm in diameter, contained 5-13 microg HRP

per mumol phospholipid and 63-74 Tf molecules per liposome (lipo Tf)

or no Tf (lipo C). Brain capillary endothelial cells (BCEC) were

incubated with liposomes at 4 degrees C (to determine binding) or at

37 degrees C (to determine association, i.e. binding+endocytosis)

and the HRP activity, rather than the HRP amount was determined in

cell lysates. Association of lipo Tf was two- to three-fold higher

than association of lipo C. Surprisingly, the binding of lipo Tf at

4 degrees C was four-fold higher than the association of at 37

degrees C. Most likely this high binding and low endocytosis is

explained by intracellular degradation of endocytosed HRP. In

conclusion, we have shown targeting of liposomes loaded with protein

or peptide drugs to the BCEC and more specifically to the lysosomes.

This is an advantage for the treatment of lysosomal storage disease.

However, drug targeting to other intracellular targets also results

in intracellular degradation of the drug. Our experiments suggest

that liposomes release some of their content within the BBB, making

targeting of liposomes to the TfR on BCEC an attractive approach for

brain drug delivery.

PMID: 15911226 [PubMed - in process]

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4: Drug Deliv. 2005 Jan-Feb;12(1):1-6. Related Articles, Links

PEGylated immunoliposomes directed against brain astrocytes.

Chekhonin VP, Zhirkov YA, Gurina OI, Ryabukhin IA, Lebedev SV,

Kashparov IA, Dmitriyeva TB.

Laboratory of Immunochemistry, Serbsky National Research Centre for

Social and Forensic Psychiatry, Moscow, Russia. chekhonin@...

Polyethylene glycol (PEG)ylated (stealth) immunoliposomes directed

against human gliofibrillary acidic protein (GFAP) were prepared by

coupling the thiolated monoclonal anti-GFAP antibodies with a

maleimide derivative of phosphatidyl ethanolamine of the liposomal

membrane. Experiments with cell cultures demonstrated specific and

competitive binding of these immunoliposomes to embryonic rat brain

astrocytes. Administered intravenously into rats, the

immunoliposomes displayed typical kinetics with elimination half-

lives of 8-15 hr. Being incapable of penetrating the unimpaired

blood-brain barrier (BBB), these immunoliposomes, nevertheless, may

be useful in delivering drugs to glial brain tumors (which continue

to express GFAP) or to other pathological loci in the brain with a

partially disintegrated BBB.

PMID: 15801714 [PubMed - indexed for MEDLINE]

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5: NeuroRx. 2005 Jan;2(1):99-107. Related Articles, Links

Drug transport to brain with targeted liposomes.

Schnyder A, Huwyler J.

Department of Research and Division of Clinical Pharmacology,

University Hospital Basel, CH-4031 Basel, Switzerland.

Antibody-conjugated liposomes or immunoliposomes are particulate

drug carriers that can be used to direct encapsulated drug molecules

to diseased tissues or organs. The present review discusses examples

of successful applications of this technology to achieve drug

transport across the blood-brain barrier. In addition, information

will be provided on practical aspects such as phospholipid

compositions of liposomes, antibody coupling technologies, large-

scale production of liposomes, and obstacles related to drug loading

of the carrier. Prospects of future uses of immunoliposome-based

drug delivery systems such as gene therapy of the brain and clinical

trials are discussed.

PMID: 15717061 [PubMed - in process]

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6: Yao Xue Xue Bao. 2004 Nov;39(11):944-8. Related Articles, Links

[study on nerve growth factor liposomes on crossing blood-brain

barrier in vitro and in vivo]

[Article in Chinese]

Xie Y, Ye LY, Cui W, Xu K, Zhang XB, Lou JN, Hou XP.

Department of Physical-Pharmacy, School of Pharmaceutical Science,

Peking University, Beijing 100083, China.

AIM: To study the permeability of nerve growth factor (NGF)

liposomes (NGF-L, NGF-SSL, NGF-SSL-T) on the blood-brain barrier

(BBB) model and the distribution in vivo, and analyze the

correlation between the results in vitro and in vivo. METHODS: The

BBB model in vitro was established by using mouse brain

microvascullar endothelial cell, and the model was applied to study

the permeability of NGF liposomes. The distribution of NGF of each

group was studied by 125I labeled and SDS-PAGE method. RESULTS: The

highest encapsulation proportion was 34%, and the mean size of NGF

liposomes was below 100 nm. The permeability of NGF liposomes on in

vitro BBB model showed that the liposome could promote NGF to

transport across the BBB, the permeability of NGF-SSL-T was the

highest. The distribution in the brain showed in an order of NGF

concentration NGF-SSL-T > NGF-SSL + RMP-7 > NGF-SSL > NGF-L. There

was a close relationship between P(e) (permeability coefficient on

in vitro BBB model) and BUI (brain uptake constant in vivo).

CONCLUSION: Liposomes can promote NGF to transport across the BBB,

and the transporting ability BBB of NGF-SSL-T which RMP-7

incorporated into the surface of NGF liposomes is the best.

PMID: 15696939 [PubMed - indexed for MEDLINE]

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7: Yao Xue Xue Bao. 2003 Nov;38(11):867-70. Related Articles, Links

[Effect of RMP-7 and its derivatives on the transportation of

liposome into the brain]

[Article in Chinese]

Zhang XB, Jin YG, Xie Y, Xu K, Hou XP.

Department of Physical-pharmacy, School of Pharmacy, Peking

University, Beijing 100083, China.

AIM: To study the action of RMP-7 and its derivative on transporting

liposome across the blood brain barrier (BBB) into the brain.

METHODS: RMP-7 and DSPE-PEG-NHS [[1,2-dioleoyl-sn-glycero-3-

phosphoethanolamine-n-[poly (ethylene-glycol)]-hydroxy succinamide]]

were conjugated together in mild condition and MALDI-TOF-MS (Matrix-

Assisted Laser Desorption-Ionization Time-of-Flight Mass

Spectrometry) was used to determine their molecular ratio. An in

vitro BBB model was established and used to determine in vitro

bioactivity of RMP-7 and its derivative. The fluorescence of brain

slices and the Evens Blue (EB) concentration in the brain, liver,

spleen, lung and kidney of each group were used to evaluate the in

vivo bioactivity of RMP-7 and its derivative on transporting

liposome across the BBB. RESULTS: The average molecular weight (MW)

of the reaction product was 4,900, while those of DSPE-PEG-NHS and

RMP-7 were 3,224 and 1,098. The results demonstrated that RMP-7 was

conjugated to DSPE-PEG-NHS at the molecular ratio of 1:1, so the

product was DSPE-PEG-RMP-7. RMP-7 and DSPE-PEG-RMP-7 was shown to

improve the transporting of peralcohol enzyme across the in vitro

BBB model 2-3 times higher than the peralcohol enzyme only. DSPE-PEG-

RMP-7 could facilitate the transporting of EB into brain more easily

than RMP-7. CONCLUSION: Both RMP-7 and DSPE-PEG-RMP-7 could

facilitate the transporting of liposome across the BBB, especially

DSPE-PEG-RMP-7.

PMID: 14992004 [PubMed - indexed for MEDLINE]

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8: Pharm Res. 2002 Oct;19(10):1430-8. Related Articles, Links

Design and characterization of liposomes containing long-chain N-

acylPEs for brain delivery: penetration of liposomes incorporating

GM1 into the rat brain.

Mora M, Sagrista ML, Trombetta D, Bonina FP, De Pasquale A, Saija A.

Department of Biochemistry and Molecular Biology, Faculty of

Chemistry, University of Barcelona, Spain.

PURPOSE: To develop a suitable liposomal carrier to encapsulate neu

roactive compounds that are stable enough to carry them to the brain

across the blood-brain barrier with the appropriate surface

characteri tics for an effective targeting and for an active

membrane transport. METHODS: Liposomes containing glycosides and a

fusogenic lipid were prepared by extrusion. Photon correlation

spectroscopy, fluorescent spectroscopy, and differential scanning

calorimetry were used to characterize liposomal preparations. Tissue

distribution was determined by using 3H-cholesterylhexadecylether as

a marker. RESULTS: The incorporation of glycoside determinants and N-

palmitoylphosphatidylethanolamine gives liposomes with similar in

tial size, trapped volume, negative surface charge, bilayer

fluidity, and melting temperature, except for monosialoganglioside-

containing liposomes, which showed less negative surface charge and

the highe size, trapped volume and melting temperature. All

glycosilated formulations gave liposomes able to retain up to the

95% of encapsulated carboxyfluorescein after 90 min at physiologic

temperature even in the presence of serum. Monosialoganglioside

liposomes were recovered in the cortex, basal ganglia, and

mesencephalon of both brain hemispheres. The liver uptake was higher

for sulfatide- and glucose-liposomes, whereas the higher blood

levels were observed for glucose- and mannose-liposomes.

CONCLUSIONS: These results show the suitability of such liposomal

formulations to hold encapsulated drugs. Moreover, the brain uptake

of monosialoganglioside liposomes makes them good candidates as drug

delivery systems to the brain.

PMID: 12425459 [PubMed - indexed for MEDLINE]

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9: Cancer. 2001 Oct 1;92(7):1936-42. Related Articles, Links

Long-term stabilization in patients with malignant glioma after

treatment with liposomal doxorubicin.

Fabel K, Dietrich J, Hau P, Wismeth C, Winner B, Przywara S,

Steinbrecher A, Ullrich W, Bogdahn U.

Department of Neurology, University of Regensburg,

Universitaetsstrasse 84, 93053 Regensburg, Germany.

ulrich.bogdahn@...

BACKGROUND: Resistance to chemotherapeutic agents and poor blood-

brain barrier penetration are major limitations in the treatment of

malignant glioma. To improve drug delivery across the blood-brain

barrier, the authors used doxorubicin as liposomal encapsulated

formulation (Caelyx, Scheringh-Plough, Munich, Germany) in therapy

of recurrent malignant glioma. METHODS: Fifteen patients with

recurrent high-grade gliomas were included in the study. Of these,

13 patients could be evaluated, including 6 patients with

glioblastoma, 1 patient with gliosarcoma and 6 patients with

anaplastic astrocytoma. The treatment consisted of liposomal

doxorubicin (20 mg/m(2)), applied intravenously every 2 weeks.

RESULTS: Stabilization of the disease was observed in 54% (7 of 13)

of patients. Partial response and complete response (CR) were not

observed. Median time-to-progression was 11 weeks. Progression free

survival at 12 months was 15%. Median overall survival (OS) after

doxorubicin therapy was 40.0 weeks, whereas the median OS after

diagnosis reached 20.0 months (87.0 weeks). Doxorubicin was well

tolerated, with main side effects being palmoplantar

erythrodysesthesia occurring in 38% and myelotoxicity (World Health

Organization Grade 3-4) in 31% of the patients. CONCLUSIONS:

Doxorubicin has been shown to be a safe treatment with moderate

activity that may lead to long-term stabilization in recurrent high-

grade glioma patients. Of note, median OS after all and after

initiation of recurrence therapy was prolonged in comparison with

the OS in other Phase II studies, as recently described by Wong et

al. (Wong ET, Hess KR, Gleason MJ, Jaeckle KA, Kyritsis AP, Prados

MD, et al. Outcomes and prognostic factors in recurrent glioma

patients enrolled onto phase II clinical trials. J Clin Oncol

1999;17:2572.). Copyright 2001 American Cancer Society.

Publication Types:

Clinical Trial

PMID: 11745268 [PubMed - indexed for MEDLINE]

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10: J Drug Target. 2000;8(6):435-46. Related Articles, Links

Endocytosis and transcytosis of an immunoliposome-based brain drug

delivery system.

Cerletti A, Drewe J, Fricker G, Eberle AN, Huwyler J.

Dept. of Research and Clinical Pharmacology, University Hospital, CH-

4031 Basel, Switzerland.

Immunoliposomes conjugated with the OX26 monoclonal antibody to the

rat transferrin receptor can be used for brain delivery of small

molecules. In the present study the uptake of OX26-immunoliposomes

by target cells as well as their transcytosis across the blood-brain

barrier was investigated. Microscopy of RG2 rat glioma cells

incubated with fluorescence labeled OX26-immunoliposomes revealed

intracellular co-localization of liposomal cargo, the liposomal

membrane bilayer and the OX26 monoclonal antibody. The distinct

particulate staining pattern was indicative for accumulation of OX26-

immunoliposomes within endosomal or lysosomal compartments.

Prolonged incubations demonstrated endosomal release of the

liposomal cargo propidium iodide to the cytoplasm. A maximum of 50%

of propidium iodide was released from the endosomal compartment

after 24 hours of incubation. Transcytosis was studied using an in

vitro model of the blood-brain barrier consisting of immortalized

RBE4 rat brain endothelial cells. OX26-immunoliposomes did permeate

across the RBE4 cell monolayer and showed a permeability coefficient

of P(app) = 1.6 x 10(-5) ml/s. Transport was inhibited at low

temperature, by competition with free OX26 or by exchanging the OX26

monoclonal antibody for an unspecific isotype antibody. Transcytosis

of OX26-immunolipsomes was confirmed in vivo by the brain perfusion

and capillary depletion technique. OX26-immunoliposomes were

detected within the post-vascular compartment of brain parenchyma

(PS product = 2.4 microl/g/min.) and were not associated with the

brain microvasculature.

PMID: 11328669 [PubMed - indexed for MEDLINE]

> Hi all,

>

> There are three incredibly interesting and hopeful posts on

> today. A CFS patient has posted

his

> notes from a conference call that included Loomis (savvy

head

> of the HHV-6 Foundation), Dr. Dharam Ablashi (studies HHV-6),

Rich Van

> Konynenburg (studies glutathione) and a patient who helped bring

them

> together.

>

> Sue ,

> Upstate New York

>

> 80800

> Lipoceutical Glutathione tops Foscarnet, 70 others PART 1 , Aug.

1,

> 4:13 a.m.

>

> 80803

> Lipceutical Glutathione tops Foscarnet, UK Genes PART 2

> Hi, All. Continuing on from PART 1 of this message about the

successful

> in...

> davidhall2020 4:46 am

>

> 80808

> Lipoceutical Glutathione top Foscarnet, The Call PART 3

> Hi, All. Now concluding from PART 1 & 2 of this message regarding

the

> new...

> davidhall2020

> 7:52 am

[Guest guest]

Hi . Rich is on vacation for 2 weeks...but on

may be able to answer your question?

dan

> > Hi all,

> >

> > There are three incredibly interesting and hopeful posts on

> > today. A CFS patient has

posted

> his

> > notes from a conference call that included Loomis (savvy

> head

> > of the HHV-6 Foundation), Dr. Dharam Ablashi (studies HHV-6),

> Rich Van

> > Konynenburg (studies glutathione) and a patient who helped bring

> them

> > together.

> >

> > Sue ,

> > Upstate New York

> >

> > 80800

> > Lipoceutical Glutathione tops Foscarnet, 70 others PART 1 , Aug.

> 1,

> > 4:13 a.m.

> >

> > 80803

> > Lipceutical Glutathione tops Foscarnet, UK Genes PART 2

> > Hi, All. Continuing on from PART 1 of this message about the

> successful

> > in...

> > davidhall2020 4:46 am

> >

> > 80808

> > Lipoceutical Glutathione top Foscarnet, The Call PART 3

> > Hi, All. Now concluding from PART 1 & 2 of this message regarding

> the

> > new...

> > davidhall2020

> > 7:52 am