Current
Medicinal Chemistry -Central Nervous System Agents, Volume 2, No. 3, 2002
Comparative Anatomy, Physiology and In Vitro
Models of the Blood-Brain and Blood-Retina Barrier Pp. 175-186
Burkhard Schlosshauer and Heiko Steuer
Blood-Brain Barrier Penetration and Drug
Development from an Industrial Point of View Pp. 187-201
Katharina Mertsch and Jochen Maas
In Vitro Models of the Blood-Brain Barrier:
When to Use Which? Pp.
203-209
A(Bert).G. de Boer and P.J. Gaillard
Computational Approaches for the Prediction
of Blood-Brain Barrier Permeation Pp. 211-227
Wolfgang Sippl
Lipophilicity and Other Parameters Affecting
Brain Penetration Pp.
229-240
F. Atkinson, S. Cole, C. Green and H. van de Waterbeemd
Transport of Drugs Across the Blood-Brain
Barrier by Nanoparticles Pp.
241-249
Jörg Kreuter
[Back to top] Comparative Anatomy, Physiology and In Vitro
Models of the Blood-Brain and Blood-Retina Barrier
Burkhard
Schlosshauer and Heiko Steuer
The blood-brain barrier (BBB) represents a functional
interface between the blood stream and the neuronal microenvironment. Distinct
cellular and molecular features of brain microvessel endothelial cells result
in barrier and carrier functions that guarantee exclusion of adverse components
such as neurotoxic metabolites on the one hand and selective passage of
essential nutrients on the other hand. Circumventricular organs are brain
structures that lack BBB characteristics, allowing for hormone-mediated
interactions between e.g. the pituitary and distant organs. The retina, as an integral
part of the central nervous system, is enclosed by the pigment epithelium,
which functions as a barrier interface between the systemic blood vessels of
the neighboring choroid and the retina. Various BBB-specific markers, tight
junction components, and carrier systems including amino acid and saccharide
transporters have been cloned. P-glycoprotein has been of special interest
because this efflux pump counteracts entry of numerous therapeutically relevant
drugs into the nervous system. Various in vitro systems of the retinal pigment
epithelium (RPE) have been established and employed to analyze pharmacological
aspects and pathological cell interactions. The most advanced systems are
organotypic cultures and acute preparations of the RPE, i.e. fully intact
tissue sheets that can be used as in vivo-like BBB models for transport studies
and drug profiling.
[Back to top] Blood-Brain Barrier Penetration and Drug
Development from an Industrial Point of View
Katharina Mertsch and Jochen Maas
To be effective as therapeutic agents, centrally acting drugs must cross the blood-brain barrier (BBB). Conversely, to be devoid of unwanted CNS effects, peripherally acting drugs must show limited brain accessibility. This demonstrates clearly the need for different methods to assess the blood-brain penetration at different levels of project development in industry. Since the experimental determination of blood-brain partitioning is difficult, time consuming and expensive, also other methods like in-silico approaches mainly based on physicochemical properties like solubility, lipophilicity, molecular size, hydrogen-bonding capacity and charge are used. Approaches for drug delivery and drug modification are also reviewed in the present article.
In vitro cellular models based on cell cultures growing in
two-chamber systems for transport studies or isolated microvessels play an
important role for compound screening. To achieve and use the full potential of
these models a characterization of anatomical, physiological and biochemical
properties is needed. The more strict the criteria for BBB models the better
prediction of penetration and cellular mechanisms. Unfortunately, the
throughput decreases often in parallel. Therefore, though high-throughput
assays as the MDCK/CACO assay or artificial membrane assays are used but they
still suffer from low predictability for specifically transported substances
(transporters, Pglycoprotein, brain specific receptors) due to differences
between peripheral epithelial cells and brain endothelial cells.The animal
experiment with radiolabelled and non-radiolabelled compound not only have the
highest level and the highest predictability but the highest cost and lowest
throughput as well. There is no “golden rule“ for approaching brain penetration
in industry but models available are used often in parallel (in silico, in
vitro, in vivo).
[Back to top] In Vitro Models of the Blood-Brain Barrier:
When to Use Which?
In this paper the various BBB systems have been described
including the various isolation procedures of brain capillary endothelial cells
(BCEC), the culture of monolayers of BCEC and endothelial cell lines, and the
coculture of BCEC with various types of astrocytes. The transference of results
between BBB culture systems has been discussed together with the application of
the various BCEC co-culture systems in research. It is concluded that there is
a need for interlaboratory validation of BBB data, particularly BBB transport
data.
[Back to top] Computational Approaches for the Prediction
of Blood-Brain Barrier Permeation
Wolfgang
Sippl
Recently, one of the key trends in the pharmaceutical industry has been the integration of what has traditionally been considered ‘development’ activities into the early phases of the drug discovery process. The aim of this integration is the prompt identification and elimination of candidate molecules that are unlikely to survive later phases in drug development. Combinatorial chemistry and high throughput screening techniques have enormously increased the possibility of finding new lead structures. Applying these techniques millions of compounds can be generated but most of them show poor biopharmaceutical properties. Identifying and removing compounds with poor properties at an early stage is strongly demanded to save both time and costs. Because biopharmaceutical parameters, such as the blood-brain permeation, cannot be determined for a large number of compounds, alternative evaluation methods are desirable.
In the last thirty years a variety of theoretical
transport and permeation models have been developed to describe mathematically
how a drug is passively transported and how a compound is able to pass a
membrane. Progress in understanding the role of physicochemical properties in
membrane permeability relevant to important processes such as blood-brain
barrier permeation,brings rational drug design more within reach. Several new
methods able to estimate rapidly the biopharmaceutical properties on the basis
of molecular structures have been developed recently. This article will review
the most important and recent techniques in this field and will discuss their applicability
in the drug discovery process.
[Back to top] Lipophilicity and Other Parameters Affecting
Brain Penetration
F. Atkinson, S. Cole, C. Green and H. van de
Waterbeemd
Over the past 50 years a number of efforts have been made
to relate physicochemical parameters to the ability of molecules to cross the
blood-brain barrier. Predominantly drugs enter the brain cells by transcellular
passive diffusion through cells while paracellular transport is not considered
significant due to the tight junctions between cells. Early work focused on
correlations of brain uptake with a measured value of lipophilicity.
Computational models were developed to model this parameter and the important
structural characteristics of molecules, e.g. size and hydrogen-bonding
capacity. New molecular descriptors, such as polar surface area and solvent
free energies, have been generated and used. Practical methodology for
predicting passive diffusion has also diversified with the use of cell monolayers
and artificial membranes. These methods need to be validated against
appropriate in vivo data and there is a need to consider the brain penetration
data itself. Brain uptake is often expressed as partitioning of drugs into
whole brain from blood or plasma, but the usual receptor targets for drugs are
in the aqueous environment, extra-cellular fluid (ECF) surrounding the cell. In
the brain, ECF concentrations are generally regarded to be reasonably well
represented by cerebro-spinal fluid (CSF) concentrations. However there is
limited literature data on CSF concentrations. In the blood-brain barrier the
transporter P-glycoprotein (P-gp) is known to limit brain-uptake of certain
compounds. In addition some compounds, including sugar and amino acids, may be
actively transported into the brain. Models for brain penetration in the future
are likely to include a number of in silico computed parameters or a number of
physical measurements to allow contributions of passive and active transport to
be considered.
[Back to top] Transport of Drugs Across the Blood-Brain Barrier by
Nanoparticles
Jörg
Kreuter
Poly(butyl cyanoacrylate) nanoparticles coated with polysorbate 80 (Tween® 80) enable the transport of a number of drugs across the blood-brain barrier (BBB) into the brain following intravenous injection. These drugs include the hexapeptide endorphin dalargin, the dipeptide kytorphin, loperamide, tubocurarine, doxorubicin, and the NMDAreceptor antagonists MRZ 2/576 and MRZ 2/596.
After binding to the polysorbate-coated particles, dalargin as well as loperamide exhibited a dose-dependent antinociceptive effect after i.v. injection as determined by the tail-flick as well as by the hot plate test. This effect was accompanied by a Straub reaction and was totally inhibited by pretreatment with naloxone, indicating that it is a central effect and not peripheral analgesia. After brain perfusion of rats with tubocurarine bound to the polysorbate-coated nanoparticles epileptic spikes were observable in the EEC of the rats but not with the controls. Other very interesting results were obtained with the NMDA-receptor antagonists MRZ 2/576 and MRZ 2/596. The very short anticonvulsive response of MRZ 2/576 was increased from below 30 to 300 min, and the transport across the BBB of the non-penetrating MRZ 2/596 was enabled after i.v. injection. Intravenous injection of polysorbate 80-coated nanoparticles loaded with doxorubicin (5 mg/kg) achieved very high brain levels of 6 µg/g brain tissue while all the controls, including uncoated nanoparticles and doxorubicin solutions mixed with polysorbate, did not reach the analytical detection limit of 0.1 µg/g. Moreover, experiments with the extremely aggressive glioblastoma 101/8 transplanted intracranially showed a long term survival for 6 months of 40 % of the rats after intravenous injection of the polysorbate 80-coated nanoparticle preparation (3 x 1.5 mg/kg). The surviving animals were sacrificed after this time and showed total remission by histological investigation. Untreated controls died within 10 - 20 days, the animals in the six other control groups between 10 – 50 days.
The mechanism of the drug transport across the blood-brain
barrier with the nanoparticles requires further elucidation. The most likely
mechanism at present appears to be endocytotic uptake by the brain capillary
endothelial cells followed either by release of the drugs in these cells and
diffusion into the brain or by transcytosis. Endocytotic uptake of the
polysorbate-coated nanoparticles but not of uncoated particles has been shown
with bovine, murine, rat, and human brain capillary endothelial cells. After
injection of the nanoparticles, apolipoprotein E (apo E) or apo B adsorption of
the particles seems to occur as already shown in vitro, followed by interaction
with the LDL receptor and endocytotic uptake. This scenario is rather likely
since both apolipoproteins can interact with the LDL-receptor. They may then be
taken up like the naturally occurring lipoprotein particles.