Christoffer Bundgaard, Christian Jes Nyberg Jensen, Mats Garmer Discovery DMPK, H...
Transcript of Christoffer Bundgaard, Christian Jes Nyberg Jensen, Mats Garmer Discovery DMPK, H...
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Species comparison of in vivo P-glycoprotein mediated brain efflux using mdr1a deficient rats and mice
Christoffer Bundgaard, Christian Jes Nyberg Jensen, Mats Garmer Discovery DMPK, H. Lundbeck A/S, DK-2500 Valby, Denmark
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Copyright 2011 by the American Society for Pharmacology and Experimental Therapeutics.
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Running title: Brain efflux comparison between P-gp KO rats and mice
Corresponding author:
Christoffer Bundgaard
H. Lundbeck A/S
Discovery DMPK
9 Ottiliavej
DK-2500 Valby, Denmark
Phone: +45 3630 1311
Email:[email protected]
Number of text pages: 31
Number of tables: 3
Number of figures: 2
Number of references: 35
Number of words
In Abstract: 250
In Introduction: 736
In Discussion: 1497
Nonstandard Abbreviations: ABC, ATP-binding cassette; BBB, Blood-brain barrier;
CNS, central nervous system; P-gp, P-glycoprotein (mdr, multidrug resistance); KO,
Knockout; Kp, Total brain to plasma concentration ratio; Kp,uu, Unbound brain to
unbound plasma concentration ratio; fu,plasma, Unbound fraction in plasma; fu,brain,
Unbound fraction in brain; WT, Wild-type
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Abstract
The experiments described herein compared the extent of in vivo P-gp mediated
brain efflux between rats and mice for a set of known CNS compounds. With use of
newly introduced genetically modified mdr1a deficient rats and their gene-competent
counterparts, the brain to plasma distribution was assessed and compared to the
distribution pattern in mdr1a deficient and wild-type mice. Four compounds
(aripiprazole, citalopram, risperidone and venlafaxine) were administered using a
continuous subcutaneous osmotic minipump infusion paradigm. Steady-state brain
and plasma concentrations of the compounds, including selected metabolites (9-OH
risperidone, O-desmethyl-venlafaxine and N-desmethyl-venlafaxine) were measured
in mdr1a deficient rats and mice and their wild-type counterparts along with their free
fractions to determine total and unbound brain to plasma distribution between
genotypes within and between species. The results revealed qualitative as well as
quantitative similarities between P-gp functionality in vivo at the blood-brain barrier
level in rats and mice. All tested compounds were shown to have significantly higher
brain to plasma distribution in both mdr1a deficient rats and mice compared to their
wild-type counterparts. Moreover, the relative enhancement in extent of brain
penetration between mdr1a deficient and wild-type rats could be directly correlated to
the enhancement ratios obtained in mice. From the unbound brain to unbound
plasma distributions (Kp,uu), the impact of P-gp to the overall brain penetration
capabilities showed minor differences between rats and mice for the compounds
tested. In conclusion, a comparable functional role of P-gp between rats and mice
with respect to brain efflux mediated by this transporter is suggested.
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Introduction
The physiology of the blood-brain barrier (BBB) with its endothelial cells lining the
brain capillaries connected by tight junctions limits the extent of brain penetration for
many xenobiotics. In addition to the physical barrier, numerous transport proteins are
present at the BBB acting as efflux pumps to actively exclude a wide variety of
chemically diverse compounds from the central nervous system (CNS). One of the
most studied members of this group of transporters is P-glycoprotein (P-gp, ABCB1),
a mammalian plasma membrane phospho-glycoprotein encoded by the multidrug
resistance 1 (MDR1) gene (mdr1 in rodents). P-gp belongs to the ATP-binding
cassette (ABC) transporter superfamily and is located in the brain in the luminal
membrane of the endothelial cells but is also expressed in other organs (Thiebaut et
al, 1987). For compounds designed to exert their effect in the CNS, drug-P-gp
interactions could influence the potential efficacy by reducing the brain concentration
due to this efflux mechanism. Thus, P-gp has been shown in mice to have a profound
influence on brain concentrations of many types of drug substrates including
antidepressants and antipsychotics (Ejsing et al., 2007; Linnet and Ejsing, 2008).
Consequently, screening for P-gp substrate recognition and assessment of the
potential impact hereof are therefore important parts of the profile evaluation of new
chemical entities, particularly those targeted to the CNS (Wager et al., 2011).
During lead optimization programs and drug candidate characterization, in
vivo pharmacology and pharmacokinetic studies are often performed in rats and
mice. In such studies it is important to understand the potential differences in the
functional activity of P-gp between these species with respect to drug distribution to
the brain in order to make proper comparisons and extrapolations (Kim et al., 2008;
Syvanen et al., 2009). In both rats and mice, P-gp is the most highly expressed ABC
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protein in brain endothelial capillary cells (Warren et al., 2009). Rats and mice shares
two genes encoding for P-gp, mdr1a and mdr1b, which are similarly expressed
between the two species and in both of these species, mdr1a is the predominant
form in the brain (Croop et al., 1989; Kwan et al., 2003). In addition there is a high
degree of homology in the amino acid content of P-gp between rats and mice of
around 93% (Hubbard et al., 2007). Studies investigating differences in P-gp
functionality between rats and mice have primarily been conducted in vitro applying
efflux ratios derived from bidirectional transport studies in cell systems transfected
with mdr1 from each species. These studies suggest some correlation between rat
and mouse P-gp mediated efflux but also that differences may exist depending on
the substrate (Booth-Genthe et al., 2006; Katoh et al., 2006; Takeuchi et al., 2006).
However, comparison of efflux ratios between species from in vitro transport studies
is circumvented with indecisive biological challenges such as total protein expression
and density of the functional transporter in each cell line.
With respect to in vivo studies investigating quantitative species differences in
P-gp function between rats and mice, very limited amount of data have been
published. Cutler et al. (2006) used infusion of the P-gp inhibitor GF-120918
(elacridar) and reported that similar total plasma concentrations of the inhibitor were
required in rats and mice to achieve the same relative increase in the brain to plasma
distribution ratio of an undisclosed P-gp substrate. However, species differences in
plasma protein binding of the inhibitor or substrate are confounding factors that may
influence the total plasma concentrations required to inhibit P-gp. Thus, such
chemically induced knockout studies are not readily comparable between species.
With the advent of mdr1a/b deficient mice either from genetically knockout
modifications or mutant origin (Schinkel et al., 1996), these animals has become
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widely applied in drug discovery to assess the role of P-gp in vivo on drug disposition
including BBB transport studies. These transporter deficient animal models have
suffered from only being available in mice species making quantitative interspecies
comparisons difficult. Recently however, mdr1a knockout rats became commercially
available through the application of Zinc Finger Nuclease technology enabling
targeted gene knockout (Geurts et al., 2009; Chu et al., 2011). The purpose of this
study was therefore to compare the P-gp functionality between mdr1a deficient rats
and mice and there gene-competent counterparts (wild-types) with respect to in vivo
brain distribution. A range of known CNS compounds and their selected metabolites
previously recognized to be P-gp substrates were evaluated in these animal models
using a similar experimental design across species.
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Methods
Animals
Homozygous mdr1a knockout and wild-type rats (bodyweight 350-500 g) were
obtained from SAGETM Labs, Sigma Life Science, Sigma-Aldrich Co. (SD-
Abcb1atm1sage). Mdr1a deficient mice and CF-1TM (non-Swiss) outbred background
mice (bodyweight 25-45 g) were purchased from Charles River Laboratories
(Crl:CF1-Abcb1amds mutants). All animals arrived at the animal facility at least five
days before being used in experiments. During acclimatization, animals were housed
in pairs under controlled conditions of temperature (21 ± 2 °C), relative humidity (55 ±
5%) and a 12-h light–dark cycle (lights on at 6:00 am). Food and tap water were
freely available in the home cage. Approval of all animal procedures was granted by
The Danish National Committee for Ethics in Animal Experimentation.
Drugs and dosing
The administered test compounds included citalopram, venlafaxine, risperidone and
aripiprazole which all are extensively used CNS compounds. In addition, these
compounds and their selected metabolites represent a relative wide span in their
functional extent of substrate affinity towards P-gp at the BBB level in mice (Linnet
and Ejsing, 2008). Alzet osmotic minipumps (Durect Corporation, Cupertino, CA)
were used to deliver the test compounds in steady-state dosing regiments. Alzet
model 2ML1 (delivery rate 10 µl/h) and model 2001 (delivery rate 1 µl/h) were applied
for rats and mice respectively. Citalopram (hydrobromide, H. Lundbeck A/S),
venlafaxine (hydrochloride, Sigma) and risperidone (free base, Sigma) were
dissolved in 0.9% NaCl. Aripiprazole (free base, Sequoia Research) was dissolved in
20% hydroxylpropyl-β-cyclodextrin, pH4. Test compounds were used in the following
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concentrations (as free base) in the pumps with corresponding average doses
calculated based on body weight and pump delivery rate: Citalopram: rats 15 mg/ml
(~9 mg/kg/day), mice 6 mg/ml (~5 mg/kg/day). Venlafaxine: rats 15 mg/ml (~9
mg/kg/day), mice 20 mg/ml (~12 mg/kg/day). Risperidone: rats 10 mg/ml (~5
mg/kg/day), mice 4 mg/ml (~4 mg/kg/day). Aripiprazole: rats 12 mg/ml (~6
mg/kg/day), mice 3 mg/ml (~3 mg/kg/day). Prior to implantation, the pumps were
equilibrated by soaking them overnight in sterile saline solution at 37 ºC. Before
surgery animals received Rimadyl (carprofen) subcutaneously as analgesic
treatment. During brief isoflurane anesthesia, minipumps were implanted
subcutaneously on the back of the animals under aseptic conditions. Postoperative
treatment was provided by Temgesic (buprenorphine) on the day of surgery and
Rimadyl the following two days. 3-5 wild-type and mdr1a deficient rats and mice were
used in each treatment group. The continuous delivery was maintained for 3 days in
both mice and rats to attain steady-state after which the animals were decapitated
and the brains dissected out. Trunk blood was collected in EDTA-coated tubes and
plasma were harvested following centrifugation (5000 g for 10 min at 4 °C). Brain
homogenate was prepared by homogenizing the whole brain with 70% acetonitrile
(1:4 v/v) followed by centrifugation and collection of the supernatant. Plasma and
brain supernatant samples were frozen at -80°C until analysis.
Bioanalysis
Rat and mice plasma and brain homogenate samples were prepared by precipitation
and addition of generic internal standard. Plasma and brain concentrations of dosed
test compounds and selected metabolites (O-desmethyl venlafaxine, N-desmethyl
venlafaxine and 9-OH-risperidone) were determined using UltraPerformance LC
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chromatography (Acquity UPLC, Waters, Milford, MA, USA) followed by MS/MS
detection using a Sciex-API 4000 MS (AB Sciex, Foster City, CA, USA) (Erichsen et
al., 2010). The lower limit of quantification was typically 0.5 ng/ml in plasma and 1
ng/g in brain (peak S/N > 5).
Free fraction determinations
Plasma protein binding and non-specific binding in brain homogenates were
determined in naïve male rats (Sprague-Dawley) and mice (Crl:NMRI(Han)) by use of
equilibrium dialysis. The procedure was modified from Kalvass and Maurer (2002).
Dialysis membranes (96 well formats, cut-off 12-14 kDa, HTDialysis) was soaked in
PBS/ethanol 80:20 and rinsed in deionized water before use. Brain homogenate or
plasma was added to the donor side and spiked with test compound to a final
concentration of 1 µM. The receiver side contained phosphate buffer (pH 7.4).
Equilibrium dialysis was performed by incubating at 37°C for 5 h. Subsequently, the
compound concentration was measured in the buffer phase using liquid
chromatography-mass spectrometry (LC-MS). The free fraction in plasma (fu,plasma)
and intact brain tissue (fu,brain) was calculated according to Kalvass and Maurer
(2002) with their dilution factor in brain taken into account. For all compounds,
recovery following incubation were in the range of 91-109 %.
Data analysis and statistics
Brain to plasma distribution ratios (Kp) were calculated from the measured total drug
levels in the respective matrices (Cbrain,total and Cplasma,total). To compare brain to
plasma distribution in mdr1a deficient and wild-type animals between species, Kp
values were normalized and expressed relative to wild-types (100%) for each
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species. In addition, the unbound brain to unbound plasma partition coefficients,
designated Kp,uu, were calculated by associating free fractions with total brain to
plasma distribution at steady-state for each species as Cbrain,total × fu,brain / Cplasma,total ×
fu,plasma (Hammarlund-Udenaes et al., 1997; Gupta et al., 2006). Student’s t-test was
used to compare brain to plasma distribution ratios data between mdr1a deficient and
wild-type animals. Statistical and regression analyses were performed using
GraphPad Prism® (Vers. 4.02, GraphPad Software, USA).
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Results
Following 3 days of continuous treatment using minipumps, the four test compounds
and three selected formed metabolites were measured in plasma and brains of wild-
type and mdr1a deficient rats and mice. The observed total plasma and brain
concentrations and corresponding Kp values are summarized in Table 1 for rats and
Table 2 for mice. For all compounds except 9-OH risperidone there were no
statistically significant differences between the systemic plasma exposure in mdr1a
deficient animals and their wild-type counterparts. For 9-OH risperidone, plasma
concentrations were significantly higher in mdr1a deficient mice as compared to wild-
type mice (p<0.05). This trend was also observed for 9-OH risperidone in mdr1a
deficient rats versus wild-type rats although not to a statistically significant degree.
From the derived Kp values, all the tested compounds tended to display higher brain
distribution in mice as compared to rats when comparing the species within in each
genotype. Thus, risperidone, 9-OH risperidone and aripiprazole all had Kp values
below 0.22 in wild-type rats. Among wild-type mice, 9-OH risperidone and
aripiprazole had the lowest Kp values of 0.28 and 0.49. Of the compounds evaluated,
citalopram exhibited the highest Kp value in both species.
All the tested compounds in the data set were identified as being substrates
for P-gp at the BBB level in both rats and mice. Thus, from the Kp values, all the
tested compounds were shown to have significantly higher brain distribution in both
mdr1a deficient rats and mice compared to their wild-type counterparts. To
quantitatively compare the impact of P-gp on brain distribution capabilities of the
compounds between rats and mice, Kp values were normalized to account for
differences in absolute total plasma and brain exposure. Figure 1 shows the
normalized Kp values for the test compounds in mdr1a deficient animals relative to
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their wild-type controls (set as 100%) for each species. All the compounds exhibited
significant enhancement in their normalized Kp value in mdr1a deficient animals. In
both rats and mice, the most pronounced effect of P-gp mediated brain efflux was
observed for risperidone (835% ±197 enhancement in mdr1a deficient rats; 737%
±221 in mice) and its metabolite 9-OH risperidone (1100% ±274 enhancement in
mdr1a deficient rats; 1200% ±155 in mice). The weakest effect of P-gp on the relative
brain distribution was seen with venlafaxine (192% ±18 enhancement in mdr1a
deficient rats; 176% ±20 in mice) and citalopram (330% ±27 enhancement in mdr1a
deficient rats; 201% ±8 in mice). As for the most pronounced P-gp substrates, the
weaker relative effects of P-gp on brain distribution of venlafaxine and citalopram
were also shown to be present in both species. When ranking the impact of P-gp on
brain distribution across all tested compounds, considerable overlap between rats in
mice was observed. In rats this was (low to high) venlafaxine, O-desmethyl
venlafaxine, citalopram, aripiprazole, N-desmethyl venlafaxine, risperidone, 9-OH
risperidone. The corresponding ranking for mice was venlafaxine, citalopram,
aripiprazole, O-desmethyl venlafaxine, N-desmethyl venlafaxine, risperidone, 9-OH
risperidone. The overall relationship between the enhancements in the normalized Kp
values in mdr1a deficient rats and mice is visualized in Figure 2. Regression analysis
revealed a linear correlation between the two species with a slope close to unity
(y=1.08x - 102; r2=0.95, p=0.0002).
To assess the impact of P-gp deficiency on the brain to plasma distribution of
the compounds independent of binding to plasma proteins and brain tissue, the
plasma and brain free fractions determined in vitro were combined with measured
total brain and plasma concentrations at steady-state to calculate Kp,uu for each
compound in wild-type and mdr1a deficient animals (Table 3). In wild-type rats all
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compounds had Kp,uu values well below unity consistent with active efflux dominating
the BBB distribution of these compounds. The Kp,uu values in mdr1a deficient rats
were increased to around unity for the majority of the compounds. In mice, all the
compounds except for citalopram exhibited Kp,uu values below unity. Citalopram was
shown to have a Kp,uu value of 2.4 in wild-type mice which was increased to 4.8 in
mdr1a deficient mice whereas the other compounds were increased to values closer
to unity as compared to respective values in wild-type animals.
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Discussion
The experiments described herein compared the prevalence and magnitude of in
vivo P-gp mediated brain efflux between rats and mice for a set of known CNS
compounds. In order to compare the extent of brain distribution between rats and
mice and the impact of P-gp, all experiments were performed at steady-state. This
design allowed for calculations of Kp and Kp,uu between brain and plasma with
species dependent pharmacokinetics taken into account.
From the Kp values obtained in mdr1a deficient rats and mice and their wild-
type counterparts, the experiments revealed qualitative as well as quantitative
species similarities between P-gp functionality at the BBB level. Thus, all compounds
evaluated were shown to be substrates in both species. Moreover, as shown in
Figure 2, the extent of increase in the normalized Kp values between mdr1a deficient
animals and there wild-type counterparts could be directly correlated between rats
and mice. The indication of a comparable quantitative role of in vivo P-gp
functionality at the BBB between rats and mice observed in this study corroborate
with expression levels and protein homology of mdr1 between these species (Kim et
al., 2008). Also in vitro studies in mdr1a transfected cells indicate a comparable
functional role of P-gp between these species although some substrate dependency
has been reported (Booth-Genthe et al., 2006; Katoh et al., 2006; Takeuchi et al.,
2006).
The tested compounds have previously been reported to be substrates for P-
gp in vivo at the BBB in mice as shown by an increased brain to plasma distribution
in P-gp knockout versus to wild-type mice. Citalopram has been shown to have 2-3
fold higher Kp value in P-gp knockout mice (Uhr and Grauer, 2003; Doran et al.,
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2005; Uhr et al., 2008) in line with the present mice data (201% ±8 increase). In
mdr1a deficient rats, citalopram showed an enhancement ratio of 330% ±27 which
could indicate that this compound has a slightly higher propensity for P-gp mediated
brain efflux in rats as compared to mice. Assessment of bidirectional transport
properties across the BBB including impact of P-gp can be based on the ratio
between unbound concentrations in brain and plasma at equilibrium conditions,
expressed as Kp,uu. If Kp,uu is close to unity, the BBB transport is predominantly
mediated by passive diffusion, or the impact of influx and efflux transport is equal. If
Kp,uu is below unity, efflux transport may explain this whereas a Kp,uu above unity is an
indication that active uptake processes are involved (Hammarlund-Udenaes et al.,
1997; Hammarlund-Udenaes et al., 2008). Interestingly, citalopram displayed a Kp,uu
value in wild-type mice of 2.4 which was increased to 4.8 in mdr1a deficient mice,
indicating that, despite clear P-gp mediated efflux, some involvement of uptake
transporters is also suggested to contribute to the overall BBB distribution of
citalopram. This pattern was also observed for citalopram in rats although to a lesser
extent. A putative involvement of active uptake is supported by in vitro transport
studies across bovine brain endothelial cells, where citalopram has been reported to
be transported by means of a carrier-mediated mechanism (Rochat et al., 1999).
Thus, a possible species dependent affinity to uptake transporters could in theory
influence the functional transport activity at the BBB differently between mdr1a
deficient rats and mice although this remains to be explored.
Aripiprazole has previously been shown to be a substrate for P-gp in mice with
a 2.7-fold increase in Kp in knockout mice following sub-chronic minipump treatment
(Kirschbaum et al., 2010). In the present study a similar increase in Kp was observed
in mdr1a deficient mice (269% ±39 increase). Like for citalopram, a slightly higher
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increase in Kp for aripiprazole was attained in mdr1a deficient rats compared to wild-
type rats (338% ±38 increase) which cannot rule out some species difference of P-gp
functionality for this compound. The Kp,uu values of aripiprazole in mdr1a deficient
rats and mice were increased to reach similar values and did not reached unity.
However, some uncertainty in these derived values is to be expected as the free
fractions of aripiprazole were extremely low in plasma as well as in brain.
Risperidone and its metabolite 9-OH risperidone are well studied P-gp
substrates in mice with profound P-gp effects on extent of brain penetration shown by
at least 10 times higher Kp values in knockout mice in a range of different studies
(Wang et al., 2004; Doran et al., 2005; Ejsing et al., 2005). These reports aligned well
with the present results where these compounds were the most pronounced
substrates in the test set with increases in Kp values in similar ranges in both mice
and rats. In both mdr1a deficient rats and mice, Kp,uu values for risperidone and 9-OH
risperidone were increased to values very close to unity, suggesting P-gp to be the
main gatekeeper for brain penetration of these substances in both species.
Brain entry of venlafaxine and its demethylated metabolites O-desmethyl-
venlafaxine and N-desmethyl-venlafaxine have previously been shown to be limited
by P-gp using knockout mice with brain penetration enhancement ratios between 2
and 4 in mice lacking P-gp following sub-chronic minipump treatment (Uhr et al.,
2008; Karlsson et al., 2011). In the current experiments similar enhancement ratios
were observed in mice. Venlafaxine and O-desmethyl-venlafaxine displayed equal
relative enhancement ratios between mice and rats lacking P-gp. N-desmethyl-
venlafaxine had a slightly higher enhancement ratio in rats (634% ±117) as
compared to mice (461% ±13). However, relatively high variability in the Kp data of
formed N-desmethyl-venlafaxine was observed in rats. In regards to this, dosing of
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this metabolite could be considered as means to a more accurate determination of
potential species differences in the P-gp mediated effects of this compound. Kp,uu
estimates of venlafaxine and its metabolites in mdr1a deficient and wild-type animals
indicated that passive diffusion was the dominating factor for BBB distribution without
the presence of P-gp across the parent-metabolite system. Kp,uu values of N-
desmethyl-venlafaxine in rats and O-desmethyl-venlafaxine in mice increased to
values above 2 in the mdr1a deficient species but the aforementioned variability in N-
desmethyl-venlafaxine exposure and the very high free fraction of O-desmethyl-
venlafaxine in mice plasma suggests that these values should be interpreted with
caution.
Development of the genetically mdr1 deficient rat model, provides a simple
tool for evaluating the functional consequence of P-gp efflux at the BBB by the
measurement of drug concentrations in brain and plasma at steady-state. However,
some drug and/or species dependent limitations in this model and methodology likely
exist that may affect interpretation of the obtained Kp and Kp,uu estimates. Firstly, it
has been shown that knockout of P-gp in mice can result in alterations in the mRNA
expression levels of other transporter proteins (Cisternino et al., 2004). Thus, if the
compound under investigation has substrate affinity for other efflux or uptake
transporters besides P-gp, this could potentially affect the brain to plasma distribution
obtained in mdr1 deficient models. A very recent characterization of the mdr1a
deficient rat model suggest that only minor compensatory changes in expression of
other transporter related genes occur at the BBB level in male rats lacking P-gp (Chu
et al., 2011). Secondly, continuous drug administration may increase P-gp
functionality at the BBB as a result of mdr1 induction. This has been suggested for
venlafaxine in vitro in human cell systems (Ehret et al., 2007; Bachmeier et al., 2011)
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and in vivo in rats following 3 weeks of continuous treatment (de Klerk et al., 2010)
and. Thus, if species differences exist in terms of liability and extent of mdr1
induction following sub-chronic treatment, this could theoretically hamper a
quantitative comparison of P-gp mediated efflux between rats and mice. Thirdly,
intrabrain drug distribution governed by other processes than nonspecific binding
may potentially impact the estimates of Kp,uu in the present study. Hence, determining
brain free fractions using the homogenate methodology only measures nonspecific
binding and does therefore not account for other intrabrain distributional aspects
such as sequestration into lysosomes which might affect cellular retention for basic
compounds. Also, The homogenization process may alter the binding properties by
unmasking binding sites that are not accessible to a drug in intact brain tissue.
However, recent studies suggest that a reasonable correlation can be established
between nonspecific binding from homogenates and binding obtained using brain
slices with a preserved cellular structure (Friden et al., 2011).
By use of the same experimental methodology in rats in mice, the validity of
the mdr1a deficient rat model was confirmed in the present study where all selected
P-gp substrates identified in mice was also recognized in rats. This model thus allows
for a direct means to assess the potential influence of P-gp on pharmacokinetic and
pharmacodynamic parameters in rats without the need for chemical P-gp inhibitors.
Besides qualitative similarities in the P-gp functionality between rats and mice, the
data obtained here also indicates substantial quantitative similarities in the extent of
P-gp mediated efflux at the BBB between these species. This information might
prove useful when extrapolating existing knowledge on P-gp mediated efflux
obtained in mice to the rat species during pharmacokinetic and in vivo
pharmacological evaluations.
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Acknowledgements
Mona Elster is acknowledged for technical assistance.
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Authorship Contributions
Participated in research design: Bundgaard, Jensen
Conducted experiments: Jensen, Garmer
Performed data analysis: Garmer, Bundgaard
Wrote or contributed to the writing of the manuscript: Bundgaard
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References
Bachmeier CJ, Beaulieu-Abdelahad D, Ganey NJ, Mullan MJ, and Levin GM (2011)
Induction of drug efflux protein expression by venlafaxine but not desvenlafaxine.
Biopharm Drug Dispos 32: 233-244.
Booth-Genthe CL, Louie SW, Carlini EJ, Li B, Leake BF, Eisenhandler R, Hochman
JH, Mei Q, Kim RB, Rushmore TH, and Yamazaki M (2006) Development and
characterization of LLC-PK1 cells containing Sprague-Dawley rat Abcb1a (Mdr1a):
comparison of rat P-glycoprotein transport to human and mouse. J Pharmacol
Toxicol Methods 54: 78-89.
Chu X, Zhang Z, Yabut J, Horwitz S, Levorse J, Li XQ, Zhu L, Lederman H, Ortiga R,
Strauss J, Li X, Owens KA, Dragovic J, Vogt TF, Evers R, and Shin MK (2011)
Characterization of mdr1a/P-glycoprotein knockout rats generated by zinc finger
nucleases. Mol Pharmacol doi: 10.1124/mol.111.074179.
Cisternino S, Mercier C, Bourasset F, Roux F, and Scherrmann JM (2004)
Expression, up-regulation, and transport activity of the multidrug-resistance protein
Abcg2 at the mouse blood-brain barrier. Cancer Res 64: 3296-3301.
Croop JM, Raymond M, Haber D, Devault A, Arceci RJ, Gros P, and Housman DE
(1989) The three mouse multidrug resistance (mdr) genes are expressed in a tissue-
specific manner in normal mouse tissues. Mol Cell Biol 9: 1346-1350.
Cutler L, Howes C, Deeks NJ, Buck TL, and Jeffrey P (2006) Development of a P-
glycoprotein knockout model in rodents to define species differences in its functional
effect at the blood-brain barrier. J Pharm Sci 95: 1944-1953.
This article has not been copyedited and formatted. The final version may differ from this version.DMD Fast Forward. Published on November 23, 2011 as DOI: 10.1124/dmd.111.043083
at ASPE
T Journals on O
ctober 31, 2020dm
d.aspetjournals.orgD
ownloaded from
DMD #43083
22
de Klerk OL, Bosker FJ, Willemsen AT, Van WA, Visser AK, de JT, Dagyte G, den
Boer JA, Dierckx RA, and Meerlo P (2010) Chronic stress and antidepressant
treatment have opposite effects on P-glycoprotein at the blood-brain barrier: an
experimental PET study in rats. J Psychopharmacol 24: 1237-1242.
Doran A, Obach RS, Smith BJ, Hosea NA, Becker S, Callegari E, Chen C, Chen X,
Choo E, Cianfrogna J, Cox LM, Gibbs JP, Gibbs MA, Hatch H, Hop CE, Kasman IN,
Laperle J, Liu J, Liu X, Logman M, Maclin D, Nedza FM, Nelson F, Olson E,
Rahematpura S, Raunig D, Rogers S, Schmidt K, Spracklin DK, Szewc M, Troutman
M, Tseng E, Tu M, Van Deusen JW, Venkatakrishnan K, Walens G, Wang EQ, Wong
D, Yasgar AS, and Zhang C (2005) The impact of P-glycoprotein on the disposition of
drugs targeted for indications of the central nervous system: evaluation using the
MDR1A/1B knockout mouse model. Drug Metab Dispos 33: 165-174.
Ehret MJ, Levin GM, Narasimhan M, and Rathinavelu A (2007) Venlafaxine induces
P-glycoprotein in human Caco-2 cells. Hum Psychopharmacol 22: 49-53.
Ejsing TB, Morling N, and Linnet K (2007) A review on the relation between the brain-
serum concentration ratio of drugs and the influence of P-glycoprotein. Drug Metabol
Drug Interact 22: 113-129.
Ejsing TB, Pedersen AD, and Linnet K (2005) P-glycoprotein interaction with
risperidone and 9-OH-risperidone studied in vitro, in knock-out mice and in drug-drug
interaction experiments. Hum Psychopharmacol 20: 493-500.
Erichsen MN, Huynh TH, Abrahamsen B, Bastlund JF, Bundgaard C, Monrad O,
Bekker-Jensen A, Nielsen CW, Frydenvang K, Jensen AA, and Bunch L (2010)
Structure-activity relationship study of first selective inhibitor of excitatory amino acid
This article has not been copyedited and formatted. The final version may differ from this version.DMD Fast Forward. Published on November 23, 2011 as DOI: 10.1124/dmd.111.043083
at ASPE
T Journals on O
ctober 31, 2020dm
d.aspetjournals.orgD
ownloaded from
DMD #43083
23
transporter subtype 1: 2-Amino-4-(4-methoxyphenyl)-7-(naphthalen-1-yl)-5-oxo-
5,6,7,8-tetrahydro-4 H-chromene-3-carbonitrile (UCPH-101). J Med Chem 53: 7180-
7191.
Friden M, Bergstrom F, Wan H, Rehngren M, Ahlin G, Hammarlund-Udenaes M, and
Bredberg U (2011) Measurement of unbound drug exposure in brain: modeling of pH
partitioning explains diverging results between the brain slice and brain homogenate
methods. Drug Metab Dispos 39: 353-362.
Geurts AM, Cost GJ, Freyvert Y, Zeitler B, Miller JC, Choi VM, Jenkins SS, Wood A,
Cui X, Meng X, Vincent A, Lam S, Michalkiewicz M, Schilling R, Foeckler J, Kalloway
S, Weiler H, Menoret S, Anegon I, Davis GD, Zhang L, Rebar EJ, Gregory PD, Urnov
FD, Jacob HJ, and Buelow R (2009) Knockout rats via embryo microinjection of zinc-
finger nucleases. Science 325: 433.
Gupta A, Chatelain P, Massingham R, Jonsson EN, and Hammarlund-Udenaes M
(2006) Brain distribution of cetirizine enantiomers: comparison of three different
tissue-to-plasma partition coefficients: K(p), K(p,u), and K(p,uu). Drug Metab Dispos
34: 318-323.
Hammarlund-Udenaes M, Friden M, Syvanen S, and Gupta A (2008) On the rate and
extent of drug delivery to the brain. Pharm Res 25: 1737-1750.
Hammarlund-Udenaes M, Paalzow LK, and de Lange EC (1997) Drug equilibration
across the blood-brain barrier-pharmacokinetic considerations based on the
microdialysis method. Pharm Res 14: 128-134.
Hubbard TJ, Aken BL, Beal K, Ballester B, Caccamo M, Chen Y, Clarke L, Coates G,
Cunningham F, Cutts T, Down T, Dyer SC, Fitzgerald S, Fernandez-Banet J, Graf S,
This article has not been copyedited and formatted. The final version may differ from this version.DMD Fast Forward. Published on November 23, 2011 as DOI: 10.1124/dmd.111.043083
at ASPE
T Journals on O
ctober 31, 2020dm
d.aspetjournals.orgD
ownloaded from
DMD #43083
24
Haider S, Hammond M, Herrero J, Holland R, Howe K, Howe K, Johnson N, Kahari
A, Keefe D, Kokocinski F, Kulesha E, Lawson D, Longden I, Melsopp C, Megy K,
Meidl P, Ouverdin B, Parker A, Prlic A, Rice S, Rios D, Schuster M, Sealy I, Severin
J, Slater G, Smedley D, Spudich G, Trevanion S, Vilella A, Vogel J, White S, Wood
M, Cox T, Curwen V, Durbin R, Fernandez-Suarez XM, Flicek P, Kasprzyk A, Proctor
G, Searle S, Smith J, Ureta-Vidal A, and Birney E (2007) Ensembl 2007. Nucleic
Acids Res 35: D610-D617.
Kalvass JC, and Maurer TS (2002) Influence of nonspecific brain and plasma binding
on CNS exposure: implications for rational drug discovery. Biopharm Drug Dispos 23:
327-338.
Karlsson L, Hiemke C, Carlsson B, Josefsson M, Ahlner J, Bengtsson F, Schmitt U,
and Kugelberg FC (2011) Effects on enantiomeric drug disposition and open-field
behavior after chronic treatment with venlafaxine in the P-glycoprotein knockout mice
model. Psychopharmacology (Berl ) 215: 367-377.
Katoh M, Suzuyama N, Takeuchi T, Yoshitomi S, Asahi S, and Yokoi T (2006) Kinetic
analyses for species differences in P-glycoprotein-mediated drug transport. J Pharm
Sci 95: 2673-2683.
Kim IW, Booth-Genthe C, and Ambudkar SV (2008) Relationship between drugs and
functional activity of various mammalian P-glycoproteins (ABCB1). Mini Rev Med
Chem 8: 193-200.
Kirschbaum KM, Uhr M, Holthoewer D, Namendorf C, Pietrzik C, Hiemke C, and
Schmitt U (2010) Pharmacokinetics of acute and sub-chronic aripiprazole in P-
glycoprotein deficient mice. Neuropharmacology 59: 474-479.
This article has not been copyedited and formatted. The final version may differ from this version.DMD Fast Forward. Published on November 23, 2011 as DOI: 10.1124/dmd.111.043083
at ASPE
T Journals on O
ctober 31, 2020dm
d.aspetjournals.orgD
ownloaded from
DMD #43083
25
Kwan P, Sills GJ, Butler E, Gant TW, and Brodie MJ (2003) Differential expression of
multidrug resistance genes in naive rat brain. Neurosci Lett 339: 33-36.
Linnet K, and Ejsing TB (2008) A review on the impact of P-glycoprotein on the
penetration of drugs into the brain. Focus on psychotropic drugs. Eur
Neuropsychopharmacol 18: 157-169.
Rochat B, Baumann P, and Audus KL (1999) Transport mechanisms for the
antidepressant citalopram in brain microvessel endothelium. Brain Res 831: 229-236.
Schinkel AH, Wagenaar E, Mol CA, and van DL (1996) P-glycoprotein in the blood-
brain barrier of mice influences the brain penetration and pharmacological activity of
many drugs. J Clin Invest 97: 2517-2524.
Syvanen S, Lindhe O, Palner M, Kornum BR, Rahman O, Langstrom B, Knudsen
GM, and Hammarlund-Udenaes M (2009) Species differences in blood-brain barrier
transport of three positron emission tomography radioligands with emphasis on P-
glycoprotein transport. Drug Metab Dispos 37: 635-643.
Takeuchi T, Yoshitomi S, Higuchi T, Ikemoto K, Niwa S, Ebihara T, Katoh M, Yokoi
T, and Asahi S (2006) Establishment and characterization of the transformants
stably-expressing MDR1 derived from various animal species in LLC-PK1. Pharm
Res 23: 1460-1472.
Thiebaut F, Tsuruo T, Hamada H, Gottesman MM, Pastan I, and Willingham MC
(1987) Cellular localization of the multidrug-resistance gene product P-glycoprotein in
normal human tissues. Proc Natl Acad Sci USA 84: 7735-7738.
This article has not been copyedited and formatted. The final version may differ from this version.DMD Fast Forward. Published on November 23, 2011 as DOI: 10.1124/dmd.111.043083
at ASPE
T Journals on O
ctober 31, 2020dm
d.aspetjournals.orgD
ownloaded from
DMD #43083
26
Uhr M, and Grauer MT (2003) abcb1ab P-glycoprotein is involved in the uptake of
citalopram and trimipramine into the brain of mice. J Psychiatr Res 37: 179-185.
Uhr M, Tontsch A, Namendorf C, Ripke S, Lucae S, Ising M, Dose T, Ebinger M,
Rosenhagen M, Kohli M, Kloiber S, Salyakina D, Bettecken T, Specht M, Putz B,
Binder EB, Muller-Myhsok B, and Holsboer F (2008) Polymorphisms in the drug
transporter gene ABCB1 predict antidepressant treatment response in depression.
Neuron 57: 203-209.
Wager TT, Villalobos A, Verhoest PR, Hou X, and Shaffer C (2011) Strategies to
optimize the brain availability of central nervous system drug candidates. Expert Opin
Drug Discov 6: 371-381.
Wang JS, Ruan Y, Taylor RM, Donovan JL, Markowitz JS, and DeVane CL (2004)
The brain entry of risperidone and 9-hydroxyrisperidone is greatly limited by P-
glycoprotein. Int J Neuropsychopharmacol 7: 415-419.
Warren MS, Zerangue N, Woodford K, Roberts LM, Tate EH, Feng B, Li C,
Feuerstein TJ, Gibbs J, Smith B, de Morais SM, Dower WJ, and Koller KJ (2009)
Comparative gene expression profiles of ABC transporters in brain microvessel
endothelial cells and brain in five species including human. Pharmacol Res 59: 404-
413.
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Footnotes
Reprint requests:
Christoffer Bundgaard, PhD
H. Lundbeck A/S
Discovery DMPK
DK-2500 Valby, Denmark
Phone: +45 3630 1311
Email:[email protected]
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Legends for figures Fig. 1. Relative brain to plasma distribution for the 4 tested compounds and their
selected metabolites in mdr1a deficient rats and mice and their wild-type
counterparts following 3-day continuous minipump treatment. Data are displayed as
relative Kp values expressed as average percentage of Kp values in wild-types
(100%) ± relative SEM). Statistical difference were assessed between mdr1a
deficient and wild-type animals within each species (*p<0.05; **p<0.01; ***p<0.001).
Fig. 2. Relationship between the enhancements in the normalized Kp values in mdr1a
deficient rats and mice (designated P-gp defrat/WTrat and P-gp defmice/WTmice
respectively). Dotted line represents the line of unity and the solid line the resulting fit
of the linear regression. Kp values are shown as average ± relative SEM obtained in
each species.
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Table 1. Total plasma and brain concentrations and derived Kp values of
administered test compounds and formed metabolites in wild-type and mdr1a
deficient rats following 3-day continuous minipump treatment. Values are shown as
mean ± SEM (n=3-5).
Compound Wild-type rats Mdr1a deficient rats
Plasma (ng/ml)
Brain (ng/g) Kp
Plasma (ng/ml)
Brain (ng/g) Kp
Citalopram 27 ± 2.4 265 ± 16 10 ± 0.78 35 ± 5.8 1198 ± 289 33 ± 2.7
Venlafaxine 50 ± 3.7 158 ± 4.6 3.3 ± 0.29 63 ± 11 389 ± 33 6.3 ± 0.60
O-Des-venlafaxine 4.5 ± 1.4 4.4 ± 0.89 1.0 ± 0.10 11 ± 3.3 33 ± 7.8 3.3 ± 0.38
N-Des-venlafaxine 6.8 ± 1.5 9.3 ± 1.1 1.4 ± 0.14 10 ± 4.2 79 ± 21 8.9 ± 1.7
Risperidone 194 ± 34 41 ± 0.88 0.22 ± 0.04 168 ± 25 295 ± 36 1.9 ± 0.44
9-OH Risperidone 153 ± 24 22 ± 1.6 0.15 ± 0.03 192 ± 41 289 ± 14 1.7 ± 0.41
Aripiprazole 18 ± 5.3 3.0 ± 0.60 0.19 ± 0.04 22 ± 1.0 14 ± 2.1 0.64 ± 0.07
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Table 2. Total plasma and brain concentrations and derived Kp values of
administered test compounds and formed metabolites in wild-type and mdr1a
deficient mice following 3-day continuous minipump treatment. Values are shown as
mean ± SEM (n=3-5).
Compound Wild-type mice Mdr1a deficient mice
Plasma (ng/ml)
Brain (ng/g)
Kp Plasma (ng/ml)
Brain (ng/g)
Kp
Citalopram 8.8 ± 0.40 191± 19 22 ± 2.2 8.2 ± 0.7 361 ± 33 44 ± 1.6
Venlafaxine 85 ± 9.8 403 ± 63 4.7 ± 0.40 105 ± 16 828 ± 70 8.3 ± 1.0
O-Des-venlafaxine 5.7 ± 0.35 10 ± 0.87 1.8 ± 1.13 7.5 ± 1.6 40 ± 6.9 5.7 ± 0.58
N-Des-venlafaxine 17 ± 2.8 25 ± 5.1 1.5 ± 0.12 11 ± 1.7 77 ± 11 7.0 ± 0.20
Risperidone 43 ± 10 21 ± 2.3 0.52 ± 0.08 40 ± 10 149 ± 16 3.8 ± 1.1
9-OH Risperidone 45 ± 8.4 13 ± 1.7 0.28 ± 0.01 110 ± 26 366 ± 91 3.4 ± 0.44
Aripiprazole 51 ± 8.7 25 ± 4.8 0.49 ± 0.04 32 ± 6.2 41 ± 7.2 1.3 ± 0.17
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Table 3. Free fraction values obtained in plasma (fu,plasma) and brain homogenate
(fu,brain) and calculated unbound brain to plasma partition coefficients (Kp,uu) in wild-
type (WT) and mdr1a deficient (P-gp def.) rats and mice. fu-values are shown as
average of two independent determinations.
Compound
Free fraction, rats (%) Kp,uu, rats
Free fraction, mice (%) Kp,uu, mice
fu,plasma fu,brain WT P-gp def.
fu,plasma fu,brain WT
P-gp def.
Citalopram 50 3.1 0.61 2.1 32 3.5 2.4 4.8
Venlafaxine 64 9.2 0.45 0.89 >99* 18 0.86 1.4
O-Des-venlafaxine 89 35 0.38 1.2 >99* 45 0.80 2.4
N-Des-venlafaxine 32 8.0 0.34 2.0 80 16 0.29 1.4
Risperidone 11 9.0 0.17 1.4 27 5.8 0.11 0.81
9-OH Risperidone 20 15 0.11 1.1 54 18 0.10 1.1
Aripiprazole 0.10 0.10 0.19 0.64 0.20 0.10 0.25 0.64
*fu,plasma values of 99% was used to estimate Kp,uu for venlafaxine and O-Des-venlafaxine.
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