ABC transporters andcytochromes P450 in the humancentral nervous system: influenceon brain pharmacokineticsand contribution toneurodegenerative disordersFabien Dutheil, Aude Jacob, Sandrine Dauchy, Philippe Beaune,Jean-Michel Scherrmann, Xavier Decleves & Marie-Anne Loriot††Universite Paris Descartes, 45 rue des Saints-Peres, 75270 Paris Cedex 06, France
Importance of the field: The identification of xenobiotic metabolizing
enzymes (i.e., CYP) and transporters (i.e., ABC transporters) (XMET) in the
human brain, including the BBB, raises the question whether these transporters
and enzymes have specific functions in brain physiology, neuropharmacology
and toxicology.
Areas covered in this review: Relevant literature was identified using PubMed
search articles published up to March 2010. Search terms included ‘ABC
transporters and P450 or CYP’, ‘drug metabolism, effect and toxicity’ and
‘neurodegenerative disease (Alzheimer and Parkinson diseases)’ restricted to
the field of ‘brain or human brain’.
What the reader will gain: This review aims to provide a better understanding
of XMET functions in the human brain and show their pharmacological
importance for improving drug delivery and efficacy and also for managing
their side effects. Finally, the impact of brain XMET activity during neurode-
generative processes is discussed, giving an opportunity to identify new
markers of human brain diseases.
Take home message: During the last 2 decades, much evidence concerning
the specific distribution patterns of XMET, their induction by xenobiotics
and endobiotics and their genetic variations have made cerebral ABC trans-
porters and CYP enzymes key elements in the way individual patients respond
to centrally acting drugs.
Keywords: ATP-binding cassette transporters, CYP enzymes, human brain,
neurodegenerative diseases, xenobiotics
Expert Opin. Drug Metab. Toxicol. (2010) 6(10):1161-1174
1. Introduction
Xenobiotics are chemical compounds that are foreign to the body. They are foundthroughout our environment, in the air, water and our food. Exposure to them isunavoidable, whether it is deliberate (e.g., drugs) or involuntary (e.g., pollutants).Xenobiotics must be converted to their polar hydrophilic metabolites before theycan be detoxified and eliminated from the body. The cytochrome P450 (CYP)enzymes and ATP-binding cassette (ABC) transporters form the main system formetabolizing and transporting xenobiotics (xenobiotic metabolizing enzymes andtransporters; XMETs). The presence of these XMETs within the brain is importantfor the bioactivation of the xenobiotics and hence damage to cells in this target organ
for both centrally-acting drugs and environmental toxins thatcross the blood brain barrier (BBB) [1]. The lipophilic characterof several xenobiotics enables them to diffuse across BBB andenter the brain parenchyma, although the specific ABC trans-porters in the plasma membrane of brain microvessel endothe-lial cells may limit their brain distribution. The limited capacityof neurons to regenerate makes the brain extremely vulnerableto damage by toxic compounds. The expression and regulationof XMET is tissue-specific and cell type-specific, and the brainhas its own unique complement of them [2-4]. These XMETscan metabolize and transport a vast array of compounds, includ-ing centrally-acting drugs, neurotoxins, neurotransmitters andneurosteroids [5-7]. The identification of ABC transporters andCYPs in the brain, including the BBB, raises the questionwhether these transporters and enzymes have specific functionsin brain physiology, neuropharmacology and toxicology.Individuals respond differently to centrally-acting thera-
peutic drugs, and their response is not always predicted bythe concentration of drug in their blood plasma [8]. Drugsthat act on the central nervous system (CNS) may be metab-olized in situ within the brain, and alterations in the degree ofin situ metabolism may contribute to a great variability in theresponses of individuals to a drug. In the same way, ABCtransporters may limit the penetration of a CNS drug intothe brain and, therefore, its efficacy. The concentrations ofbrain ABC transporters and CYPs are determined by an indi-vidual’s genotype and also by their exposure to environmentalinducers and/or repressors.The brain cellular architecture is specific and complex com-
prising of the brain parenchyma with cell bodies of astrocytes,neurons, oligodendrocytes and microglia, and the BBB withbrain endothelial cells, foot astrocytes and pericytes. Thisreview focuses on the nature and distribution of the XMETin the human brain parenchyma and at the BBB. We examinethe recent developments in our understanding of the roleof these proteins in brain physiology, in the response to
xenobiotics (drugs, toxic compounds) and in the pathogenesisof neurodegenerative disorders.
2. CYP enzymes in the human brain
The first studies that established the existence of brain CYPswere done on rats in 1977. CYP monooxygenase activity wasfound in rat brain microsomes [9]. Two years later, Guengerichand Mason and Marietta et al. confirmed the catalytic capaci-ties and inducible properties of these enzymes in thebrain [10,11]. Although microsomal CYPs were present, theiractivity was only 2 -- 5% of the CYP activity in the liver, repre-senting ~ 30 pmol/ml of microsomal protein. Marietta et al.also demonstrated that cerebral CYPs were 3 -- 170 times lessactive than hepatic CYPs when acting on exogenous com-pounds such as meperidine, hexobarbital and benzo[a]pyr-ene [11]. However, Fishman et al. published new information,in 1980, on the role of mitochondrial CYP in cerebral homeo-stasis [12]. Estradiol-2-hydroxylase, also known as CYP1A1, wasvery active just before the preovulatory release of luteinizinghormone. This metabolic enhancement was measured in themicrosomal, mitochondrial and synaptosomal fractions [12].Walther et al. subsequently showed that there was a CYP inthe mitochondrial inner membrane whose activity was compa-rable to that of microsomal CYP [13]. Thus, the amount ofmicrosomal CYP in the rat brain is 6 -- 100 pmol/mg of totalproteins, while the amount of mitochondrial CYP is around75 -- 220 pmol/mg of total proteins [14-16]. The human cerebralcortex has a similar concentration of mitochondrial CYP,100 -- 300 pmol/mg of total proteins [16].
Unlike the liver, the cytoarchitectural organization and cellfunctions in the brain are extremely variable and the concen-trations of CYPs in certain specific neurons can be as high asor higher than those in hepatocytes [17]. These brain enzymesare unlikely to contribute to overall drug metabolism, butthey may contribute to variations in the drug response ofindividuals through local in situ metabolism [18]. Brain CYPsare enzymatically active in vitro, and we and others haverecently identified the cofactors and coenzymes needed forthem to be active in vivo in their immediate environment [2,4].The presence of the enzymes, CYP reductase and ferredoxinreductase, which are required for a functional enzymaticsystem [16,19] supports the existence of active enzymes, as previ-ously documented in human and rodent brains by measuringmicrosomal monooxygenase activities and enzyme inhibitionstudies [4]. Moreover, a recent study has assessed the importanceof brain CYP activity in the analgesic action of µ-opioids [20].Conroy et al. produced a mutant mouse with reduced brainneuron-specific P450 activity. These mice showed much lowermorphine anti-nociception than controls. Pharmacologicalinhibition of brain P450 arachidonate epoxygenases also blocksmorphine anti-nociception in mice and rats [20]. Furthermore,ABC transporters, particularly ABCB1 (MDR1, P-gp), werefound in the human BBB 20 years ago [21]. Almost 1000original articles have been published on it since their discovery,
Article highlights.
. Functional xenobiotic metabolizing enzymes andtransporters (XMET) including ATP-binding cassettetransporters and P450 enzymes are present in humanbrain both in the parenchyma and the blood brainbarrier (BBB) according to a specific pattern.
. Their activities towards endogenous and exogenoussubstrates have pharmacological andtoxicological consequences.
. A better understanding of their role will allowdeveloping more efficient centrally active drugs andmodulating metabolic pathways involved inneurodegenerative diseases.
. Further studies are required to elucidate the role ofXMET and the molecular mechanisms modulating theirexpression and activity and to identify the metaboliteend products biologically active in the human brain.
This box summarizes key points contained in the article.
ABC transporters and cytochromes P450 in the human central nervous system
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but there have been few studies on the drug-metabolizing con-tent of the BBB. We recently determined the gene expressionprofiles of the CYPs and ABC transporters at the humanBBB and in the brain parenchyma [3].
2.1 CYP in the human brain parenchymaA total of 20 isoforms of human CYP (CYP1A1, 1A2, 1B1,2B6, 2C8, 2D6, 2E1, 3A4, 3A5, 8A1, 11A1, 11B1, 11B2,17A1, 19A1, 21A2, 26A1, 26B1, 27B1 and 46A1) havebeen identified in the brain. But only a limited number ofthem have been extensively studied: CYP1A1, 1A2, 2B6,2D6, 2E1 and 46A1. Most of them are widely distributedin the cortex, cerebellum, basal ganglia, hippocampus, sub-stantia nigra, medulla oblongata, pons and so on (Figure 1),but the reported data vary greatly. The discrepancies betweenthe studies are probably due to the different techniques usedto identify the CYPs. Their mRNAs or cDNA have beenidentified by real-time PCR, qualitative RT-PCR, northernblotting, Southern blotting, RNA dot blot, FISH and in situhybridization. The CYP proteins have been detected by west-ern blotting or immunostaining. The conflicting resultsbetween studies are partly due to problems of primer or anti-body specificity and sensitivity because of the great sequencehomology between CYPs [22]. We recently examined the24 genes encoding all the known human CYPs of families 1,2 and 3 and CYP46A1 using highly specific primers thathad been tested and validated in several tissues to avoid anyrisk of nonspecific amplification [23]. We found a wide varietyof xenobiotic-metabolizing enzymes in the human brain anddemonstrated their apparent selective distribution in specificparts of the brain. The major transcripts corresponding toCYP46A1, 2J2, 2U1, 1B1, 2E1 and 2D6 were present in allstructures of the human brain, but the concentrations ofmRNAs encoding each of them were unequally distributedin the different brain regions [2]. CYP transcripts had beendetected previously in whole human brain extracts [24], buttheir distribution and the presence of the corresponding pro-teins was unknown or poorly documented [24]. Our studies onCYP proteins confirmed the presence of extensively studiedCYPs such as CYP1B1 and 2D6, and we used specific anti-bodies to determine the distribution patterns of CYP2J2,CYP2U1 and CYP46A1 in the cells at different parts ofthe human brain. The main CYPs (CYP1B1, 2D6, 2J2,2U1 and 46A1) were found in the mitochondrial and micro-somal fractions of CYP proteins in the human frontal lobe,hippocampus, substantia nigra and cerebellum, but the distri-butions of these five isoforms in specific cell types all differed.CYP46A1 protein was present only in neuronal cells such asthe pyramidal cells of the frontal cortex, the CA1 -- 4 pyrami-dal cells and the granule cell of the dentate gyrus of the hippo-campus, and in cerebellar Purkinje cells. CYP1B1 protein wasmainly detected in microglial cells and to a lesser extent in theastrocytes of these four locations. Immunohistochemical stud-ies showed CYP2D6, 2J2 and 2U1 in both neuronal cells andastrocytes, but the amounts varied with the brain region.
CYP2J2 and 2U1 proteins were predominantly found in theastrocytes surrounding the cerebral blood vessels of the frontallobe and cerebellum, and these two isoforms were presentin both neurons and astrocytes of the hippocampus. Wealso identified other CYP isoforms, such as CYP2E1 andCYP2B6, in human cerebral cells. CYP2E1 protein wasdetected in specific cells at several brain locations [25]. Thisenzyme was mainly found in the pyramidal neurons and glialcells of the frontal cortex, in the pyramidal cells CA2 andCA3 and in the granular cells of the hippocampus dentategyrus [25]. CYP2B6 is also widely distributed throughout thebrain but its basal concentration is generally low [26]. Thisenzyme is present in both the neurons and glial cells of theneocortex layer I, including the astrocytes surrounding cere-bral blood vessels, where it colocalizes with glial fibrillaryacidic protein [25]. All these data are summarized in Figure 2A.
2.2 CYP in the human brain microvesselsThe BBB is composed of three main cellular elements, themicrovessel endothelial cells sealed by tight junctions, theend-feet of the astrocytes sheathing the microvessels andthe pericytes that share the basal membrane with endothelialcells (Figure 2B). They form a dynamic neurovascular unitthat is the first line of defense for the brain against unwantedcompounds. The entry of many compounds into the brain,including numerous commercial drugs, is also restricted byABC transporters; their presence in the BBB is detailed later.The BBB is not only a physical barrier but also a metabolicbarrier because of the drug-metabolizing enzymes, especiallythe CYPs, in the endothelial cells [27]. Several functionalCYPs have been found in isolated rodent brain interfaces [28].However, little is known about the CYPs at the human BBB.We recently used highly specific primers to detect the mRNAof 23 CYP isoforms from families 1, 2 and 3 in isolatedhuman brain microvessels. CYP1B1 and 2U1 mRNA werethe main CYPs in the BBB [3]. There was 14 times moreCYP1B1 mRNA in the microvessels than in the cerebral cor-tex, and the corresponding protein was also detected. In con-trast, the CYP2U1 gene was similarly expressed in isolatedmicrovessels and in the brain cortex. The CYP2D6,CYP2E1, CYP2J2 and CYP2R1 genes were alsoweakly expressed.
3. ABC transporters in the human brain
The human ABC transporter superfamily is a ubiquitous groupof 48 membrane proteins organized into seven subfamilies.These ABC transporters use the energy from ATP hydrolysisto transport endogenous and exogenous compounds acrosscell membranes. They are widely distributed in normal tissuesand play a key role in protecting the entire body against toxinsand xenobiotics by preventing their penetration (into the brain,testis or placenta) or excreting them (via the liver, kidney, gas-trointestinal tract) [21,29]. ABC transporters are also stronglyimplicated in the development of the multi-drug resistance
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(MDR) phenotype in cancer cells [30]. This review focuses onthe expression and function of ABC transporters in the humanBBB, even if some ABC transporters are also expressed inastrocytes [31,32] and pericytes [32].The first ABC transporter detected at the human BBB was
ABCB1 (MDR1, P-gp), which was initially discovered in1976 [33,49]. ABCB1 is principally present at the luminalside of the brain capillaries [32,34], but has also been found inboth the luminal and abluminal membranes of endothelialcells [32]. The ABCC (MDR associated-protein) family mem-bers are also present at the human BBB, but their expressionand functional activity are still a matter for discussion. WhileABCC4 and ABCC5 have been firmly located at the luminalside of the BBB [35], the cellular and subcellular distributionsof ABCC1 mRNA and protein are still unclear. Most studieshave not detected ABCC1 in the human brain or at thehuman BBB [36,37]. But, two studies reported findingABCC1 at the luminal side of human brain capillary endothe-lial cells [34,35]. Other ABCC transporters are not present atthe human BBB (ABCC2) or the concentrations of theirmRNAs are very low (ABCC2, ABCC3) [34,35]. Recently, weshowed that ABCC5 transcripts were the most abundantABCC mRNAs in isolated human brain microvessels [3].ABCG2 (breast cancer resistance protein), a more recently
discovered ABC transporter, is found in capillary and venousendothelium cells of many normal human tissues [38], andespecially on the luminal side of the human BBB [36]. Whilethe presence of some ABC transporters in the human BBBremains controversial, ABCB1, ABCC4 -- 5 and ABCG2 areprobably the main ABC transporters at the human BBB,where they act as the gatekeepers of the brain (Figure 2B).Their expression vary during the development of the humanCNS [39] and in disorders such as epilepsy. The genes encod-ing ABCB1, ABCC2 and ABCC5 are all overexpressed in thebrain microvessels endothelial cells of epileptic humans,whereas ABCC1 is not [40]. Thus, they are involved in bothphysiological and pathological situations.Some recent studies have focused on the relative abundance
of ABC transporters in the human brain and the BBB. A com-parative analysis of the expressions of the ABCG2, ABCB1 andABCC1 genes in homogenates of human non-malignant braintissue and human glioblastoma multiform tumors by real-timequantitative PCR showed that the expressions of ABCG2and ABCC1 were similar, higher than the expression ofABCB1 [41]. A recent study on human cerebral microvesselsisolated from patients suffering from epilepsy or glioma estab-lished the expression profiles of the genes encoding the mainABC transporters at the human BBB. Analyses of mRNA indi-cated that ABCG2 was seven times more expressed thanABCB1 in human brain microvessels, whereas ABCC4 andABCC5 were much less expressed [3]. This was also true for por-cine brain endothelium, where ABCB1 levels determined bynorthern blot analysis were lower than those of ABCG2 [42].As the relative expression of ABC transporters varies
between species [43], it is difficult to find a good animal model
with which to predict the permeability of drugs across thehuman BBB. We need to know the relative expression ofthe main ABC transporters in the human BBB in order tobetter understand the pharmacoresistance and pharmacoki-netics of CNS drugs. These data will help improve drug deliv-ery to the CNS. Additional studies are required to assess therelative amounts of ABC transporter proteins at the humanBBB to confirm that the gene expression profile agrees withthe protein profile so as to better elucidate the implicationof ABC transporters at the BBB.
4. Regulation of ABC transporters and CYPsin the human brain
The amounts of CYPs in the brain can be regulated differentlyfrom those of hepatic CYPs. The concentration of CYP2B6 inthe brain regions of smokers and alcoholics is abnormally high,particularly in the cerebellar Purkinje cells, granular cell layerand hippocampal pyramidal neurons [25]. Like CYP2B6, theconcentration of CYP2D6 protein in the brains of alcoholicsis greater than in those of non-alcoholics. It is particularlyhigh in the putamen, globus pallidus and substantia nigra,despite the fact that it is not elevated in the liver [26].CYP2E1 expression is induced by ethanol and nicotine [25].Smokers have significantly more CYP2E1 protein in severalbrain regions than do non-smokers [26]. The CYP2E1 synthesisis increased when human cultured neuroblastoma cells aretreated with nicotine [25]. Pollutants such as 2,3,7,8-tetrachloro-dibenzo-p-dioxin (or dioxin), an aromatic halogen hydrocar-bon, and b-naphtoflavone are also involved in the inductionof CYP1A1 and 1A2 synthesis in the CNS [44,45].
Transcription factors (TFs) such as nuclear receptors (NRs)and PAS/ARNT family members such as the aryl hydrocar-bon receptor (AhR) are involved in the regulation of ABCtransporter and CYP expression in peripheral tissues [46]. Weshowed that LXRb, RXRb, PPAR-d, RXRa, PPAR-a andPPAR-g are, in the order of appearance, the main TFs inthe human brain that have a location-specific action [2] inline with an earlier Japanese study [47] suggesting the in situregulation of CYP synthesis.
The cerebral distributions of several major regulation path-ways involving the AhR, NR1H2/LXRb or NR1C/PPARsubfamilies correlate well with those of the correspondingCYP target genes. High concentrations of AhR transcriptshave been found in the dura mater, which also has largeamounts of CYP1A1, CYP2S1 and CYP1B1, which can beinduced in an AhR-dependent manner [2]. The epoxygenaseCYP2J2 synthesizes EETs and other metabolites that activateboth NR1C1/PPAR-a and NR1C3/PPAR-g . Finally, recentstudies have clearly demonstrated that PPAR and LXR areNRs that are activated by fatty acid and cholesterol derivates,respectively, and control the expression of a range of genesinvolved in lipid metabolism and inflammation [48].
The pregnane X receptor (PXR) induces the expression ofABCB1, CYP3A4 and CYP2C9, the constitutive androstane
ABC transporters and cytochromes P450 in the human central nervous system
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receptor (CAR) the synthesis of CYP2B6, while the AhRmainly regulates the expression of the CYP1A and CYP1Bgenes [46]. Although PXR, CAR and AhR have been found inthe rodent BBB, the findings remain controversial [50,51]. Thereis little evidence for the presence of TFs that can stimulate thesynthesis of ABC transporters and CYP in the human BBB.Significant amounts of AhR have been found in isolatedhuman brain microvessels [3] and in human cerebral microves-sel endothelial cells (hCMEC/D3). These hCMEC/D3 cellsare a promising in vitro model of the human BBB [52]. In con-trast, PXR and CAR were barely detected in both human brainmicrovessels and hCMEC/D3 cells [3,52,53].
The majority of studies examining XMET regulationfocused at the transcriptional levels. However, some studiespointed out the implication of epigenetic modifications inXMET expression. For example, Shafaati et al. have shown ade-repression of CYP46A1 expression following a treatmentwith an histone deacetylase inhibitor [54]. RegardingABCB1, in vitro studies showed that epigenetic modulationssuch as modifications of methylation or acetylation statusmay lead to modulation of ABCB1 expression [55]. To date,epigenetic regulation of CYPs and ABC transporters withinthe human brain remains unknown.
5. Drug metabolism and pharmacokineticaspects
The BBB containing ABCB1, ABCG2, ABCC4 andABCC5 at the luminal membrane of the cerebral endothelialcells may be considered as the first line of defense againstthe entry of their substrates, limiting their distribution inthe CNS and thus their effects on the CNS (Table 1). Oncea drug has been able to reach the intracellular space, drug-metabolizing enzymes may be considered either as a seconddetoxifying barrier or as a CYP-based toxicological orpharmacological activation. In this review, we focus on theeffect of drug-metabolizing enzymes in the human brain asthey may increase or decrease efficacy and/or toxicity ofCNS-targeted drugs.
CYPs generally metabolize drugs to form hydrophilicmetabolites that are readily excreted from the body. However,the production of such metabolites in the brain could result intheir prolonged presence and slower clearance from the brain.Thus, cerebral metabolism could modulate the pharmacolog-ical response and explain, in part, the variability in responseto centrally psychoactive drugs due to differences in brainCYP-mediated metabolism between individuals [18]. Indeed,there is often a poor correlation between the plasma concentra-tions of neuroleptics and antidepressants and their therapeuticeffects. Hence, in situ metabolism may modulate the responseto these drugs. Brain microsomes slowly metabolize drugs suchas debrisoquine, sparteine and dextrometorphan, probablydue to CYP2D6 activity [56,57]. Because CYP2D6 metabolizesa wide range of centrally-acting drugs such as analgesics,anti-dementia drugs, b-blockers, tricyclic antidepressants,
anti-psychotics, monoamine oxidase-inhibitors and vasodilata-tors [58], it could be important to better understand thecontribution of this isoform to drug metabolism in the brain.
Minor drug metabolic pathways could also produce signif-icant pharmacological or toxicological responses, particularlyif they occur at the site of action. For example, a minor met-abolic pathway for codeine results in the formation of mor-phine that is the O-demethylated metabolite produced byCYP2D6. Chen et al. showed that at least the initial analgesiceffects of codeine are due to morphine produced in the brain,not in the liver [59]. Others have shown that this synthesisoccurs in the cerebral regions containing the µ-opioid recep-tors [60]. Several publications have found no link between theside effects of newer antidepressants, or the clinical responseto them and the CYP2D6 genotype [61,62], even thoughthe plasma concentrations of these drugs are influenced bythe same genotypes [61].
Because CYPs are present in the CNS and the brain is thetarget of centrally-acting drugs, their concentrations may beparticularly important for determining the response of anindividual to centrally-acting substances [22].
6. ABC transporters and CYP inneurodegenerative disorders
The identification of high concentrations of XMET in thebrain strongly suggests that the environment influences cere-bral functions. Several publications have provided new signif-icant insights into the distribution of the ABC transportersand CYP involved in the transport and metabolism of bothexogenous and endogenous substances. Previous reportshave provided evidence for the biochemical basis of neurode-generative disorders. This review examines the role of ABCtransporters and CYP in Parkinson’s and Alzheimer’s diseases.
6.1 Parkinson’s disease, CYP2D6 and ABCB1The etiology of Parkinson’s disease (PD) is usually multi-factorial, involving environmental factors and their interac-tion with susceptible genes [63]. People who have beenexposed to 1-methyl-4phenyl-1,2,3,6-tetrahydropyridine(MPTP) develop Parkinsonism due to the inhibition of mito-chondrial function in dopaminergic neurons [64]. MPTPbecomes active when it is metabolized to 1-methyl-4-phenyl-pyridinium (MPP+), which has a chemical structure similar tothat of the herbicide paraquat.
While environmental chemicals can increase the risk ofPD, host factors that influence their uptake, metabolismand distribution in the body may modulate the risk to agiven individual. Many substances can induce PD. MPTP,an analog of meperidine (or pethidine) which induces PDin humans and primates, is metabolized to MPP+ by themonoamine oxidase B in astrocytes [65]. MPTP is themain toxic compound used to generate a model of PD.This neurotoxic chemical selectively destroys substantianigra cells by inhibiting neurone mitochondrial respiration.
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CYP2D6 catalyzes the N-demethylatio of MPTP [66], whichdecreases its neurotoxic effect. Thus, CYP2D6 helps protectagainst environmental stress. In contrast, organophosphatepesticides such as parathion and chlorpyrifos are metabolizedby CYP2D6 to metabolites that can be toxic for theCNS [67].Epidemiological studies have shown that exposure to pesti-
cides may be associated with PD [68]. Organochlorines tend toaccumulate in lipid-rich organs such as the brain [69], and postmortem studies found higher concentrations of dieldrin andlindane in PD brains than in control brains [70]. Several orga-nochlorines including DDT and cyclodienes (e.g., heptachlor,endosulfan, dieldrin) have selective effects on striatal dopami-nergic neurons [71]. Dieldrin is more toxic for dopaminergiccells than for non-dopaminergic cells [72] and depletes braindopamine by increasing the production of the dopamineand vesicular monoamine transporters in the presynaptic ter-minals of dopaminergic neurons in the striatum of mice [73].Dieldrin also causes apoptotic cell death, mitochondrial dys-function and protein aggregation [74], and induces oxidativedamage in the mouse nigrostriatal dopamine system [75].Elbaz et al. studied French farmers exposed to pesticidesand showed that those with a poor metabolizer CYP2D6
phenotype were at an increased risk of PD (odds ratio: 4.7;95% CI 1.3 -- 17.4) [6].
In summary, it seems likely that the metabolism of pesticidesby brain CYPs is linked to the increased risk of PD, particularlyduring professional exposure, and possibly to the susceptibilityof an individual to PD.
The ABCB1 gene contains > 40 single nucleotide polymor-phisms, some of them linked to changes in the expressionand/or function of this ABC transporter [76]. Many studieshave investigated the relationship between ABCB1 polymor-phisms and the risk of developing PD, but only two studieshave detected a direct association to date [77-80]. Therefore,ABCB1 variants are now thought to rather act indirectly as asusceptibility factor or modulator of the risk of PD in con-junction with exposure to known risk factors of PD such aspesticides [68]. Three studies have underlined the increasedrisk of developing PD for carriers of variant C3435T orG2677[A,T] ABCB1 alleles and have been exposed topesticides [77,80,81].
Bartels et al. conducted two in vivo studies using [11C]-verapamil PET to assess the functional ABCB1 status at theBBB of patients suffering from PD. They found thatABCB1 function was not altered in the early stages of PD,
Table 1. List of substrates and modulators of CYPs and ABC transporters in human brain.
ABC transporters and cytochromes P450 in the human central nervous system
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but its function was decreased in specific brain regions ofthose with advanced PD, implying that the BBB had becomeless protective [82].
Thus, ABCB1 may be a factor in the susceptibility of PDpatients and also explains their ongoing neurodegeneration.
6.2 Alzheimer’s disease and ABC transportersAlzheimer’s disease (AD) is characterized by the accumulationand deposition of amyloid b (Ab) peptides in the brain paren-chyma and cerebral vessels. This leads to senile plaques andcerebral amyloid angiopathy (CAA) that is toxic to both neu-rons and cerebral endothelial cells [83,84]. An imbalancebetween the production of Ab peptides and their clearancefrom the brain or the aberrant trafficking of Ab peptidesacross the BBB are the two main hypotheses advanced toexplain the accumulation of Ab in the brain and the develop-ment of CAA in AD patients [85]. Ab peptides are transportedacross the BBB by receptor-mediated transcytosis: low-density lipoprotein receptor-related protein-1 is involved inthe efflux of Ab peptides from the brain, whereas the receptorfor advanced glycation end products is involved in shuttlingAb from the circulation into the brain [85,86]. Two ABC trans-porters, ABCB1 and ABCG2, have also recently been linkedto the transport of Ab peptides at the BBB. Thus,ABCB1 transports Ab peptides in vitro [87,88]. In vivo studieson Abcb1-null mice and wild-type mice showed thatABCB1 prevented the deposition of Ab in the brains of thewild-type mice as compared to the Abcb1-null mice [89].Vogelgesang et al. found no significant association betweenABCB1 polymorphisms in humans and the risk of AD, butthey highlighted an inverse correlation between Ab depositionrates and the cerebrovascular ABCB1 expression [90]. Theyalso established that the cerebral blood vessels of non-demented elderly subjects had a low ABCB1 content butabundant CAA [91]. However, it is not clear whether the lossof ABCB1 is the cause or consequence of Ab deposition.Xiong et al. showed the amount of ABCG2 in the brains ofAD/CAA patients was higher than in age-matched controls.They also established that the brains of Abcg2-null mice accu-mulated significantly more Ab peptides than did those ofwild-type mice [92]. The implication of both ABCB1 andABCG2 in preventing Ab from entering the brain was alsoconfirmed in vitro using a human brain endothelial cell line,hCMEC/D3 [93].
The above data thus strongly suggest that the ABC trans-porters, ABCB1 and ABCG2, act to protect the brain by pre-venting Ab deposition, and hence against the developmentof AD.
7. Conclusion
There is still great interest in ABC transporters and CYPenzymes because of their significant implication in the trans-port and metabolism of substances by the brain. Their activityand their subcellular and cellular distributions in the brain
parenchyma and at the BBB underline their importance formaintaining the physiology and homeostasis of the brain.Their specific distribution patterns, their induction by xeno-biotics and endobiotics and their genetic variations makeABC transporters and CYP enzymes key elements in theway individual patients respond to centrally acting drugs. Italso suggests that ABC transporters and CYPs are implicatedin the development of neurodegenerative disorders. Thus, abetter understanding of XMET functions in the human brainis of major pharmacological importance because it may pro-vide a useful tool for improving drug delivery and efficacyand also for managing their side effects. It is also an opportu-nity to identify new markers of human brain diseases. Last, abetter understanding of cerebral ABC transporters and CYPenzymes could contribute greatly to the development andoptimization of new therapeutic strategies.
8. Expert opinion
The metabolism and transport of xenobiotics in the humanbrain are a very attractive field of research because of theirimpact on the variations in individual response to drugs actingon the CNS, and susceptibility to psychiatric and neurode-generative disorders. The activities of the XMET towardsendogenous and exogenous compounds in the brain showthe importance of these proteins in brain physiology, neuro-pharmacology, development and diseases. A xenobiotic mustcross the BBB before it can reach its targets within the brainparenchyma. It is equally important to characterize the BBBand the brain parenchyma in terms of their XMET distribu-tion patterns. It is now clear that XMETs are present at theBBB and in the brain parenchyma and several recent studieshave clearly shown that the subcellular and cellular patternsof XMET distribution in these two brain structures differ dra-matically. It is also essential to identify their substrates and themetabolites that are produced in situ in order to better under-stand the pharmacological and toxicological effects of xenobi-otics. The levels of CYPs and ABC transporters in the brainare probably too low to influence the overall pharmacokinet-ics of the drugs. Nevertheless, the presence of a functionalXMET in the brain could partly explain why the differentresponses of individuals to centrally acting drugs do notalways correlate with the plasma drug concentrations. Altera-tions in brain XMET levels through genetic variation orthrough induction by xenobiotics (including drugs, nicotineor ethanol) would result in variations in local metabolism,which may contribute to the inter-individual variations seenin efficacy, drug--drug or drug--endobiotic interactions, andside effects of drugs that enter and act on the CNS. Further-more, it is probably crucial to assess the XMET activity inthe human BBB for a better understanding of the pharma-coresistance and the pharmacokinetics of drugs in order toimprove drug delivery. We, therefore, need to identify theenvironmental and genetic factors that modulate the activityof cerebral XMET in order to predict a xenobiotic response.
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We believe that CNS drug development should use specificin vitro and in vivo models to determine how new chemicalentities transported by the BBB and metabolized by thebrain. However, the complexity of cerebral structure andphysiology plus the multiple functions of the BBB makeit difficult to develop such reliable models for predictingthe metabolism and transport of xenobiotics that targetthe brain.The transcriptional and post-transcriptional regulation of
XMET activity in the brain may involve molecular mecha-nisms that differ from those occurring in the liver and otherextra-hepatic tissues. Thus, the classical inducers of XMETspecifically activate TFs (PXR, CAR) found at low levels inthe brain. The regulatory pathways probably depend notonly on the presence or the absence of TFs, but also on thecellular environment provided by biochemical mediators andtransduction signaling specific to brain cells. The recent map-ping studies on the human brain identified the major isoformsof CYPs and ABC transporters that can be potential targetsfor identifying new drugs or significant modulators of theireffects. Thus, the pharmacological inhibition or induction ofbrain CYP or ABC transporter activities can block or mediatedrug effects. A quantitative picture of XMET in the brainregions, cells and subcellular organelles will help establishtheir relative contributions to the response to xenobiotics.The cellular and regional distribution of XMET may beresponsible for neuroprotective or neurotoxic properties,depending on the biological activities of the metabolic endproducts generated by these enzymes. The location of CYPenzymes in the mitochondria could reflect a specific brainfunction for these proteins. XMET may be involved in theneuroprotection or toxic effect of neurotoxins that causeneurodegenerative disease, especially in some brain regions
where XMET concentrations are high. The involvement ofmany XMETs in the metabolism of endogenous compoundsand xenobiotics provides the opportunity to develop pharma-cological tools for modulating the biochemical pathwaysimplicated in psychiatric or neurodegenerative disorders. Forinstance, the lipid mediators produced by the activity ofCYPs are important endogenous regulators of cell prolifera-tion, differentiation, oxidative stress, neuroinflammation andapoptosis. In addition, CYP epoxygenases, which catalyzethe formation of vasoactive compounds during the arachi-donic acid cascade, may actively contribute to the regulationof the cerebral blood flow. Interestingly, while endobioticmolecules can influence the oxidation of xenobiotics byCYP enzymes, the endobiotic--xenobiotic interactions maybe of great importance for establishing the pharmacologicaland toxicological effects of xenobiotics on the brain. Finally,we need to understand how the XMET functions in orderto improve drug efficacy and explain their side effects. Suchinformation may also identify new markers of human braindiseases. The tools and concepts needed to document the rolesof CYP enzymes and ABC transporters in brain physiologyand the pathology and treatment of neurological disordersare now available.
Acknowledgements
F Dutheil and A Jacob contributed equally to the work.X Decleves English text was edited by O Parkes. X Declevesand M-A Loriot contributed equally to this review.
Declaration of interest
This work was supported by a grant from Servier Technology.
ABC transporters and cytochromes P450 in the human central nervous system
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