Full text of DOI:10.1038/sj.tpj.6500044

The Pharmacogenomics Journal (2001) 1, 178–186
 2001 Nature Publishing Group All rights reserved 1470-269X/01 $15.00
www.nature.com/tpj
REVIEW
Recent advances in P450 research
JL Raucy1,2
ABSTRACT
P450 enzymes comprise a superfamily of heme-containing proteins that cata-
SW Allen1,2
lyze oxidative metabolism of structurally diverse chemicals. Over the past few
years, there has been significant progress in P450 research on many fronts
and the information gained is currently being applied to both drug develop-
ment and clinical practice. Recently, a major accomplishment occurred when
the structure of a mammalian P450 was determined by crystallography.
Results from these studies will have a major impact on understanding struc-
ture-activity relationships of P450 enzymes and promote prediction of drug
interactions. In addition, new technologies have facilitated the identification
of several new P450 alleles. This information will profoundly affect our under-
standing of the causes attributed to interindividual variations in drug
responses and link these differences to efficacy or toxicity of many thera-
peutic agents. Finally, the recent accomplishments towards constructing
P450 null animals have afforded determination of the role of these enzymes
in toxicity. Moreover, advances have been made towards the construction
of humanized transgenic animals and plants. Overall, the outcome of recent
developments in the P450 arena will be safer and more efficient drug ther-
apies. The Pharmacogenomics Journal (2001) 1, 178–186.
1La Jolla Institute for Molecular Medicine, San
Diego, CA 92121, USA; 2Puracyp Inc, San
Diego, CA 92121, USA
Correspondence:
JL Raucy,La Jolla Institute for Molecular
Medicine,4570 Executive Dr,Suite 208,
San Diego,CA 92121,USA
Tel: +1 858 587 8788 ext 116
Fax: +1 858 587 6742
E-mail: jraucyȰljimm.org
Keywords: P450; CYP; P450 structure; humanized mice; transgenic mice; polymor-
phisms; drug metabolism
The first report of an unknown carbon monoxide binding pigment in rat liver
microsomes was in 1958. Four years later, the hemoprotein nature of the
microsomal pigment was elucidated and named cytochrome P450 because of its
unique 450-nm optical absorption peak which occurred when the cytochrome
was in its reduced carbon monoxide bound form. It was a year later, 1963, when
the function of P450 as a monooxygenase in the metabolism of 17-hydroxypro-
gesterone was discovered.1 Since that time, an enormous amount of literature
concerning P450 has accumulated.
From the 1960s to the present, research in the P450 field has involved identifi-
cation of their catalytic properties by testing thousands of compounds. In the
1970s, regulatory mechanisms and multiplicity of P450 enzymes began to be
elucidated, and in the 1980s and 1990s, cloning and identification of additional
genes occurred. To date, about 100 P450 genes have been identified in mam-
malian species. Each of these enzymes appears to be variously expressed in differ-
ent organs or cell types and can change during the development of a species. In
addition to the mammalian enzymes, numerous and interesting P450 genes are
also being described in microbes, fungi, plants and insects.2
Cytochrome P450 (CYP) is now regarded as a superfamily of heme-containing
enzymes that have diversified from a single ancestral protein to many different
forms that are widely distributed in nature. The mammalian P450 gene family
dates back about a billion years when it evolved from CYP51 (lanosterol 14␣-
Received: 1 May 2001
Revised: 31 July 2001
Accepted: 2 August 2001

Recent advances in P450 research
JL Raucy and SW Allen
179
demethylase) of Ascomycete fungi.3 However, it is generally tural core6 (Figure 1). Moreover, a region that is associated
believed that the xenobiotic metabolizing enzymes in famil-
ies 1–4 evolved later, about 600 million years ago. This was
a time when animals became terrestrial and P450s in these
families evolved due to the interaction between plants and
animals.4
with substrate binding and conserved throughout the P450
family is the center portion of the I helix.7 The microbial
P450s contain four ␤ sheets and approximately 13 ␣ helices
with an insertion termed the ‘J’ helix. The conserved struc-
tural core consists of a four-helix bundle, comprising helices
D, E, I, and L and helices J and K. Helix I contains a highly
conserved threonine with an acidic residue just N-terminal
to this which is positioned over the pyrrole ring B in the
active site. Helix K contains the absolutely conserved Glu-
X-X-Arg motif that may be involved in stabilizing the core
structure. Helix L forms part of the heme-binding region.
There are two sets of structurally conserved ␤ sheets: ␤ sheet
1 containing five strands and ␤ sheet 2 with two strands.
These sheets help form the hydrophobic substrate access
channel. As is true for all P450s, there is a structurally con-
served consensus sequence on the proximal face of the
heme containing the absolutely conserved cysteine residue.
This residue forms the 5th ligand of the heme iron and is
the reason for the characteristic 450-nm Soret absorbance
found in the carbon monoxide bound protein. All the P450
structures contain a coil termed the ‘meander’ which is
highly conserved in structure and is also located on the
proximal face of the protein.6
Because there had not been a crystal structure of a mam-
malian P450, in 1992 Gotoh,8 using the structure of CYP101
identified regions that he believed were involved in sub-
strate recognition designating them with an SRS (Substrate
Recognition Site) number. These SRS regions were extrapo-
lated to the mammalian microsomal P450s (Figure 1). While
most of the SRS regions were actually believed to be in the
active site/heme pocket and involved in substrate orien-
tation, helix A, the F–G loop and ␤ strands 1–1, and 1–2
appear to be involved in substrate recognition. The variable
regions believed to be associated with substrate binding are
helices A, B, B′, F, and G and their adjacent loops. Loops B-
B′ and B′-C line the active site (SRS-1) and helices F and G
and their loops form part of the access channel and the ceil-
ing of the active site (SRS-2 and SRS-3). The ␤ turn at the
end of the ␤ sheet 4 protrudes into the active site (SRS-6) as
does the region at the N-terminus of ␤ strand 1–4 (SRS-5). As
previously mentioned for the microbial enzymes, the other
region associated with substrate binding and conserved
throughout the P450 family is the center portion of helix I
(SRS-4).6,7
P450 enzymes are targets for recent research because they
are involved in a vast array of biological processes and cata-
lyze chemicals by a variety of oxidative mechanisms. The
result of these oxidations may be activation or inactivation
of a substrate. Almost any moderately to highly hydro-
phobic organic chemical is likely to be a substrate for a
P450.2 These chemicals include not only exogenous com-
pounds such as drugs, pollutants, pesticides and natural pro-
ducts, but also many endogenous chemicals including steroids,
eicosanoids, and other important regulatory molecules.
Because CYP genes play key roles in so many areas, the
need to understand the diversity, functions and regulation
of these enzymes continues. Studies on CYP which have
shown a remarkable expansion in the past 40 years, have
mainly focused on mammalian P450s participating in the
metabolism of xenobiotic compounds, particularly drugs,
and the biosynthesis and metabolism of steroid hormones
and other lipids.1 Although many new aspects of P450
research have been forthcoming including the elucidation
of receptors involved in xenobiotic regulation of certain
CYPs, this review will limit its focus to three areas of investi-
gation including new advances in P450 structure, on
describing polymorphisms within the P450 superfamily,
and finally, the use of transgenic animals for characterizing
P450 functions.
P450 STRUCTURE
P450s are a complex gene superfamily comprised of over 70
families. They function to metabolize xenobiotics and to
synthesize biologically active compounds such as steroids,
prostaglandins and arachidonate metabolites. Although all
P450s have a similar structural core, each has a preferred set
of substrates and a unique mechanism of substrate recog-
nition. The method in which P450s recognize their sub-
strates has largely eluded investigators in the field for years,
thereby making it difficult to fully determine their function.
In order to better understand the functionality of individual
CYPS, it has become important to determine their structure.
Understanding P450 structure has been extremely difficult
especially for the mammalian enzymes because of their
hydrophobicity. Thus, only the structures of the soluble
enzymes have been determined. These studies began in 1985
with the publication of the first crystalline structure of
P450cam. Since then, several soluble bacterial P450s have
been crystallized and their structures determined including
a recently crystallized enzyme, CYP51 from Mycobacterium
tuberculosis.5 Of the bacterial enzymes, P450cam (CYP101)
and P450BM3 (CYP102) are probably most well under-
stood.6 Between these two bacterial P450s, there is about
20% sequence identity and 47% similarity.
Recently a major milestone was achieved with the first
crystallization and structural analysis of a microsomal eucar-
yotic P450.9 The difficulty in preparing crystalline structures
of a microsomal P450 was overcome by engineering a more
soluble enzyme. CYP2C5, a progesterone hydroxylase, was
modified to increase solubility and promote crystallization.
Removal of the N-terminal trans domain and the addition of
a 4-residue histidine tag to the C-terminus resulted in P450
constructs that were more soluble and fully active. Other
mutations between amino acid residues 201 to 210 were also
All P450s have a similar three-dimensional structure with made to diminish membrane interaction and aggregation of
a similar three-dimensinal structure with a conserved struc- CYP2C5.10 This engineered P450 was used to produce dif-
www.nature.com/tpj

Recent advances in P450 research
JL Raucy and SW Allen
180
Figure 1 A cartoon illustrating the secondary structure of P450 enzymes. The turquoise banners represent helices,blue arrows represent
the ␤-sheets,and the brown lines represent the random coils connecting the helices and strands. The approximate SRS regions are
represented by a gray square. The sizes of the elements are not in proportion to their actual length. The figure is a modified representation
of the two domains and was adapted from Peterson and Graham.6
fraction quality crystals that have yielded the first mam- the electrostatic dipole of the P450 to maximize the attrac-
malian P450 structure.9
tion between the enzyme and reductase.9. The entrance to
Once the structure of CYP2C5 had been determined, dif- the putative substrate access channel between the F–G loop
ferences between the microbial P450, CYP102, and CYP2C5 and the N-terminal ␤ sheet system is located in this mem-
were identified.9 The topological elements that define the brane attachment surface. This suggests that lipophilic sub-
active site of CYP2C5 are the same as those seen in microbial strates and products could enter and exit directly from the
P450s, but the spatial arrangement differs. Only SRS-4 bilayer. An alternative substrate access channel is suggested
located in the center of helix I where it crosses the heme is by the relatively loose packing of the B–C loop with helices
highly conserved. The relative positions of the remaining G and I. This alternate route may allow substrates to enter
SRSs are more varied among P450 structures and their along helix I and would allow metabolites with increased
locations in CYP2C5 differ considerably from those of water solubility to exit into the cytoplasm. All of the resi-
CYP102. There are significant structural differences between dues that contact or nearly contact the docked progesterone
the substrate binding site of this P450 and those of soluble in CYP2C5 are in the six predicted SRSs.9 Results from these
microbial P450s as the spatial organization is poorly conserved. studies suggest mechanisms for the interaction between
The most divergent topological regions between CYP2C5 mammalian P450s and their substrates.
and CYP102 are the N-terminal ␤ sheet domain and the F–
More recently, a crystal structure was obtained for a
G loop. The N-terminal domain of CYP2C5 is rotated rela- ligand-binding domain of the human nuclear pregnane X
tive to the heme-binding site when compared to the CYP102 receptor (PXR) or steroid and xenobiotic receptor (SXR).11
structure.9 However, the spatial organization of the heme- In addition, the same ligand-binding domain complexed to
binding site of CYP2C5 is similar to the soluble microbial the cholesterol-lowering agent, SR12813 was crystallized.
P450s. Also similar to microbial P450s is the axial ligand at This receptor plays an integral role in the regulation of the
the fifth coordination site of the heme iron which is pro- drug metabolizing P450, CYP3A4, by modulating its
vided by Cys432. Coordination of the heme iron by a cyst- expression in response to certain xenobiotics. Thus
eine plays a central role in the capacity of P450s to catalyze determining its structure can provide information regarding
the scission of the dioxygen bound to the 6th coordination which xenobiotics may affect expression of CYP3A4. From
site of the heme iron.
the crystalline structure, it was determined that a small
Examination of the crystalline structure revealed that the number of polar residues are spaced throughout the smooth
orientation of CYP2C5 relative to the membrane, positions hydrophobic ligand-binding pocket of hPXR. The unique
The Pharmacogenomics Journal

Recent advances in P450 research
JL Raucy and SW Allen
181
composition of the ligand pocket allows hPXR to bind a hybridization coupled with real-time PCR or micro array
diverse set of chemicals and also permits a single ligand to analysis.17 Currently, allele identification by DNA-chip
dock in multiple orientations.11 These results provide analysis is expensive and few reports describe the use of this
important insights into how hPXR interacts with xenobiotics. technology for screening single-nucleotide polymorphisms
(SNPs). However, real-time PCR is being used with more fre-
P450 POLYMORPHISMS
quency. In this assay, the hybridization probes are com-
Due to the recent sequencing of the human genome, a great bined with the traditional forward and reverse primers. Gen-
deal of activity has occurred in the identification of new erally, hybridization is performed with two different
genes and allelic variants. Examination of the genome oligonucleotides that hybridize to two adjacent internal
revealed that only about 3% is coding sequence and the fre- sequences during amplification. One probe is labeled with a
quency of polymorphisms in these regions is about 1 in reporter fluorochrome and the other with a quencher fluor-
1000 bases.12 Genetic polymorphisms are defined as allelic ochrome. After hybridization, the probes are in close prox-
variants that occur with a frequency of 1% or greater within imity and the emitted fluorescence is quantitated. Differen-
the human population.13 Many polymorphisms will be tiation of different alleles is performed by determining the
silent, but about 68% of the possible mutations will result melting curves after PCR. PCR products are then subjected
in an amino acid substitution.3 Geneticists consider any to PCR direct sequencing. Using real time PCR and gener-
allele that persists in a population at a frequency of Ͻ1 in ation of the melting profiles of an allele-specific fluorescent
106 as having a reason for existing which can include selec- probe, a polymorphism was identified at codon 432 in the
tive pressures such as diet or environmental constituents.14 CYP1B1 gene, named CYP1B1*2.18 Geneotype frequency
In order for organisms to handle selective pressures such was determined to be 0.4 for the CYP1B1*2 allele.
as chemicals in the environment, the xenobiotic or drug
In addition, allele-specific hybridization coupled with PCR
metabolizing enzymes evolved and with that evolution can be used to screen many samples at a given time. For
came a great deal of heterogeneity in xenobiotic metaboliz- example, CYP2D6 was amplified from DNA isolated from 22
ing capabilities. Genetic heterogeneity has been increasingly samples of human liver. The allelic frequencies of the more
recognized as a significant source of variation in drug common CYP2D6*4 and *5 variants were found to be 22%
response.15 This variation is largely due to inherited differ- and 9%, respectively.19 In another study, the CYP2E1 alleles,
ences that can influence the pharmacokinetics and hence c1 and c2 were analyzed in 102 blood samples of Japanese
pharmacology of drug substrates. The effects can be pro- volunteers.20 The frequencies of the normal and mutant
found toxicity or reduced efficacy of drugs metabolized by alleles were found to be 0.83 and 0.17, respectively. With
such polymorphic enzymes.12 A new area of investigation, these technologies, the number of known defective P450
pharmacogenomics, has emerged that is applicable to the alleles is increasing at a rapid pace. The catalytic activity of
enzymes involved in the metabolism of xenobiotics and the the mutant P450s can be divided into three classifications,
molecular events associated with polymorphisms. Pharma- those where the enzyme is absent producing no activity,
cogenomics has become a popular field that will eventually those with an enzyme exhibiting diminished activity, and a
explain why drugs work better in some individuals than third group containing P450s that possess enhanced meta-
they do in others.16
bolic capabilities (Table 1).
The molecular genetic basis for inherited traits in meta-
Completely inactive alleles have been found for many
bolic enzymes began to be elucidated in the late 1980s with P540s including CYP2D6, CYP2C19, and CYP2A6 (Table 1).
the initial cloning and characterization of a polymorphic The homozygous presence of such alleles leads to a total
human gene encoding the drug metabolizing enzyme, debri- absence of enzyme and an impaired ability to metabolize
soquine hydroxylase (CYP2D6).12 Since that time, many drugs specific for these enzymes, hence the poor metabolizer
polymorphisms have been defined for the drug metaboliz- (PM) phenotype. More than 30 different defective CYP2D6
ing enzymes. Of the enzymes exhibiting polymorphisms, alleles and about 70 CYP2D6 variants have been identified.4
the CYPs or P450 enzymes have been best characterized and Of these, the six most common defective alleles will predict
play a significant role in individual differences in response the PM phenotype.21
to therapeutics. Many P450-dependent drug-metabolism
Genetic polymorphisms exhibiting inactive alleles are also
reactions are performed by polymorphic enzymes.4 Indeed, associated with CYP2C19, the S-mephenytoin hydroxylase.
of the different human P450 genes identified thus far, about The CYP2C19 polymorphism affects as many as 20% of Asi-
60% of these exhibit polymorphic sites that affect the pro- ans but only 3% of Caucasians. Indeed, a marked difference
tein sequence. Some are known and have been associated occurs in the frequency of the PM phenotype between Cau-
with the ability to metabolize drugs. Other polymorphic casians and Orientals.22 However, there are few differences
sites will be outside the coding region and may influence between Caucasians and Blacks in the frequency of the
gene expression.3
A
continuously updated website CYP2C19 PM phenotype. Therefore, certain racial differ-
(http://www.imm.ki.se/CYPalleles/) provides a major mech- ences in expression of CYP2C19 can affect drug elimination
anism for viewing newly identified P450 alleles and rel- and response. At least nine different defective CYP2C19
evant references.
alleles have been identified.23–27 Of these CYP2C19*2 and
Rapid progress in identifying polymorphisms has been CYPC19*3 contain mutations that create an aberrant splice
made possible by new techniques including allele-specific site and a premature stop codon and are the most common
www.nature.com/tpj

Recent advances in P450 research
JL Raucy and SW Allen
182
Table 1 CYP alleles and enzyme activity
mutant alleles (Table 1). However, additional mutations tive splicing and protein truncation resulting in the absence
must be present in Caucasians given that the known alleles of CYP3A5 from tissues of certain individuals.30
do not account for all PMs identified.
In addition to defective CYP genes, there are also alleles
Five different defective CYP2A6 alleles have also been that cause diminished or altered enzyme activity.4 The most
described thus far including CYP2A6*2 which encodes an common CYP2D6 allele in the Chinese is CYP2D6*10, Pro34-
inactive enzyme resulting from a Leu160His substitution, and Ser substitution, resulting in an enzyme that exhibits an
CYP2A6*3 probably generated through gene conversion impaired folding capacity and the expression of a functional
between CYP2A6 and CYP2A7, a pseudogene, causing an enzyme is diminished. The most frequent CYP2D6 variant
inactive product28 (Table 1). The allele frequency is ෂ1–3% in Black African (Zimbabwean) and Afro-American popu-
for CYP2A6*2 in Caucasians whereas a deletion of the lations appears to be CYP2D6*17. The enzyme exhibits a
CYP2A6 gene appears to be very common among Asian five-fold higher Km for codeine. An additional CYP pos-
populations.4 In addition, a novel single nucleotide poly- sessing mutant alleles with altered activity is CYP2C9.31 Two
morphism (SNP) that alters stability and activity of CYP2A6, variant CYP2C9 enzymes are known and both exhibit
CYP2A6*8, in Japanese has recently been described.29 A SNP amino acid substitutions. CYP2C9*2 encodes an enzyme
at T1412C resulted in Ile471Thr substitution. The frequency of with an Arg144Cys mutation that displays impaired func-
this polymorphism was high, 15.7%, and caused a reduction tional interactions with P450 reductase. The other variant,
in stability of the heterologously expressed enzyme. More- CYP2C9*3 yields an enzyme with an Ile359Leu that has a
over, the mutant exhibited coumarin hydroxylase activity lower affinity for many substrates (Table 1). Additional
but lacked the ability to C-oxidize nicotine. SNPs in other CYP2A6 genes have been found with dimished activity
P450s that affect expression have recently been found. including CYP2A6*6,32 CYP2A6*7,29 and CYP2A6*9.33
CYP3A5*3 and CYP3A5*6 exhibit SNPs that cause alterna- CYP2C8 was also found to exhibit polymorphisms that
The Pharmacogenomics Journal

Recent advances in P450 research
JL Raucy and SW Allen
183
resulted in altered kinetic parameters. CYP2C8*2 and *3
Recently a Cyp1a1 knockout mouse line was constructed42
have higher Km and Vmax values for the 6 ␣-hydroxylation and when hepatic enzyme activities associated with Cyp1a1
of paclitaxel when compared to wild-type CYP2C8, were assessed in microsomes from control and TCDD-
CYP2C8*1.34
treated Cyp1a1 (−/−) animals, a significant loss of aryl
Ultra-rapid metabolism is caused by the occurrence of hydrocarbon hydroxylase activity (AHH) and ethoxyresoru-
duplicated, multiduplicated or amplified genes.4 At present, fin O-deethylase activity (EROD) was noted, when compared
alleles with 2, 3, 4, 5 and 13 gene copies in tandem have with microsomes from treated and untreated wild-type
been reported for CYP2D6 and the number of individuals mice. Because Cyp1a1 plays a key role in the metabolic
carrying multiple gene copies of this enzyme is highest in potentiation of many environmental carcinogens, mutagens
Ethiopia and Saudi Arabia where up to a third of the popu- and toxins, this mouse line is invaluable for carcinogenesis
lation (29%) displays this genotype (Table 1). Another and toxicity studies. Similarly, Cyp1a2 plays a role in meta-
example of an enzyme with an allele that exhibits greater bolic activation of many xenobiotics making this P450
metabolic capabilities when compared to its wild-type important in carcinogenesis studies. Thus, mouse lines have
counterpart is CYP3A5. This P450 is polymorphically also been constructed that lack Cyp1a2.43,44
expressed at high levels in a minority of Caucasians. This
CYP1B1 shares many properties with CYP1A1 such as the
polymorphism is due to SNPs in the promoter region of the metabolic activation of polycyclic aromatic hydrocarbons
CYP3A5 gene and individuals with the CYP3A5*1 allele (PAHs). In addition to activating polycyclic aromatic hydro-
constitutively express large amounts of the enzyme30 carbons, CYP1B1 functions as a physiological regulator in
(Table 1). The occurrence of this P450 is also affected by that a deficiency in the enzyme has been linked to congeni-
race. CYP3A5 was more frequently expressed in African tal glaucoma in humans. The exact role of CYP1B1 in this
Americans (60%) than in Caucasians (33%). This P450 rep- disorder is not yet known. Because catalytic activity and
resents 50% of the hepatic CYP3A content in certain indi- expression patterns between mouse and humans are fairly
viduals expressing the enzyme, suggesting that CYP3A5 is well conserved, a Cyp1b1 null mouse has recently been con-
important in interindividual and interracial differences in structed.45 These mice are almost completely protected from
reactions catalyzed by the CYP3A enzymes. A SNP was also acute bone marrow cytotoxicity initiated by 7,12-dimethyl-
found in another P450, CYP2B6, which resulted in benz[a]anthracene (DMBA). In addition, the preleukemic
enhanced catalytic activity due to a substitutions from effects of DMBA were ablated and these animals did not
Gln172His. The frequency of this allele was 19.9% in the exhibit appreciable amounts of DMBA-DNA adducts in bone
Japanese population.35
marrow when compared to wild-type mice. Cyp1b1-null
mice that were intragastrically exposed to DMBA revealed
an attenuated incidence of lymphomas when compared to
wild-type mice.46 Such observations suggest that tissue-
USE OF GENETICALLY ALTERED ANIMALS TO STUDY
P450 FUNCTION
The role of human CYPs in metabolism of xenobiotics specific expression of CYP1B1 may contribute to cancer sus-
including therapeutic agents has been studied in vitro ceptibility in humans.
through the use of inhibitory antibodies or specific inhibi-
CYP2E1 metabolizes a large number of low-molecular
tors and in animals by CYP-specific inducers. Recent pro- weight chemicals such as industrial solvents and chemicals
gress in molecular biology has facilitated the engineering of of toxicological and carcinogenic significance. This P450 is
animal models that do not express CYPs or receptors constitutively expressed and induced in liver and other
involved in the regulation of P450s.36–40 These null mutants tissues by treatment with ethanol and other small molecular
have enabled direct studies of the in vivo role of P450 weight compounds. A Cyp2e1-null mouse line has been
enzymes in the toxicity, metabolism and carcinogenesis of established47 and used to investigate the role of CYP2E1 in
xenobiotics.
toxicity associated with benzene and acetaminophen. Myel-
The first report of a CYP null mouse was in 1995 describ- otoxicity due to benzene exposure is a consequence of
ing the Cyp1a2 knockout animals41 and since that time metabolism by CYP2E1, thus the null mice were consider-
many other mouse lines have been developed. These null ably more resistant to benzene-induced cytotoxicity than
mouse lines are generally lacking one of the xenobiotic-met- wild-type mice.48 Similar results were obtained when aceta-
abolizing CYPs that may be associated with adverse drug minophen was the substrate. This analgesic is also metabol-
reactions, but have not been linked to serious developmen- ized to a reactive intermediate by CYP2E1.49 Mice lacking
tal or physiological defects or pathological phenotypes. The Cyp2e1 were more resistant to acetaminophen-mediated
construction of these mouse lines has led to the conclusion hepatotoxicity when compared to wild-type mice, con-
that xenobiotic-metabolizing CYPs may only be required for firming that CYP2E1 is responsible for mediating toxicity
metabolism of foreign chemicals36 and that they are not from this drug. These results are in good agreement with
essential for mammalian development or physiological in vitro studies, which demonstrate that microsomes from
homeostasis.37 Moreover, those CYPs with conserved animals treated with ethanol exhibit greater formation of
activity and regulation across species, CYP1A1, 1A2, 1B1, the reactive acetaminophen metabolite.
and 2E1, have been good candidates for null mouse con-
The broad and overlapping substrate specificities of P450s
struction.37 These null mouse lines provide relevant infor- can confound identifying specific functions of individual
mation on the functions of their human counterparts.
CYPs. Thus, targeting one P450 for homologous recombi-
www.nature.com/tpj

Recent advances in P450 research
JL Raucy and SW Allen
184
nation may result in a mouse in which another P450 substi- mouse model cannot be used to study the effects of inducers
tutes for the role of the deleted enzyme. Such a response on the human CYP4A subfamily as it has recently been dem-
raises the possibility that deletion of several P450s in one onstrated that human hepatic CYP4A is not inducible by
mouse line could give insight into the contribution of these peroxisome proliferators. The reason for this refractoriness
enzymes to the metabolism of a single substrate. To date, may be because there are insufficient levels of PPAR␣ in
only one double-null mouse line, in which both Cyp1a2 human liver.57
and Cyp2e1 were disrupted, has been constructed.50 These
The orphan nuclear receptor, pregnane X receptor
mice were highly resistant to acetaminophen toxicity when (PXR)/steroid and xenobiotic receptor (SXR)42 is important
compared to wild-type, or mice lacking either Cyp1a2 or in xenobiotic-mediated induction of the CYP3A genes58 and
Cyp2e1, indicating that both CYP2E1 and CYP1A2 contrib- CYP2C8.59 To more fully understand the role of this receptor
ute to the metabolism and ultimate toxicity associated with in the regulation of CYP3A and species differences associa-
acetaminophen.40 Thus, double-null mice can provide an in ted with inducibility of this gene family, PXR null and
vivo model for studying the combined function of CYPs in humanized PXR mouse lines were constructed.60 The loss of
chemically-induced toxicity.
PXR did not alter basal expression of mouse Cyp3a. How-
Several receptors involved in the regulation of P450s have ever, the Cyp3a gene was no longer inducible by prototypi-
been targeted for disruption. The aryl hydrocarbon receptor cal inducers such as dexamethasone and PCN, indicating a
(AhR) is a transcriptional regulatory protein, which binds role for this receptor in mediating xenobiotic induction of
to DNA response elements known as DRE (dioxin response Cyp3a. This mouse line was also used to examine whether
element). Because the [Ah] gene battery comprises at least PXR played a role in induction of Cyp3a by phenobarbital,
six and probably many more genes, the AhR controls consti- a known inducer of the CYP3A gene family. In mice lacking
tutive expression of several xenobiotic-metabolizing PXR, Cyp3a was induced by phenobarbital.61 Interestingly,
enzymes. AhR-null mice have been constructed by several the phenobarbital-mediated induction of Cyp3a was
laboratories43,51,52 and these animals exhibited immune sys- accompanied by induction of a second P450, Cyp2b10, and
tem impairment and hepatic fibrosis confirming that AhR this enzyme was consistently higher in the PXR null mice
has a developmental and physiological role. Activation of than wild-type. Cyp2b10 is also up-regulated by xenobiotics
the AhR by polycyclic aromatic hydrocarbons (PAHs) and such as phenobarbital and chemically related compounds
associated compounds is associated with disruption of mul- by the constitutive androstane receptor (CAR). Results from
tiple hormone systems resulting in developmental and these experiments suggest that CAR also regulates Cyp3a
reproductive impairment.53 This may explain why mice expression. To more fully understand species differences,
lacking the AhR exhibit developmental alterations. When transgenic animals lacking mouse PXR, but containing
used to assess toxicity, these AhR-null mice were resistant to human PXR were examined for Cyp3a inducibility by
the acute toxicity from high levels of TCDD that induced inducers known to enhance expression of human CYP3A.
pathological responses in the liver of wild-type mice.53,54 In Human PXR caused constitutive up-regulation of the mouse
addition, treatment of null and wild-type mice with Cyp3a gene and was responsive to human-specific inducers
benzo[a]pyrene resulted in tumor formation in either skin such as rifampicin.60 In wild-type mice, rifampicin does not
or liver of control animals, but mice lacking the AhR pos- induce Cyp3a. These results demonstrate that the species
sessed no apparent tumors.51 These mice present an origin of the receptor rather than the structure of Cyp3A
important tool for determining the toxic effects associated genes, dictates the pattern of Cyp3a inducibility.
with dioxin and PAH exposure.
Future directions that are already beginning to be realized
In response to peroxisome proliferators such as fibric acid with the recent development of a humanized PXR mouse
derivatives, the nuclear receptor peroxisome proliferator- line, include additional lines that express human P450s in
activated receptor ␣(PPAR␣) regulates transcription of genes a background devoid of the corresponding mouse P450s. A
encoding peroxisomal, mitochondrial, and some P450 humanized mouse line would provide an animal model for
enzymes involved in fatty acid oxidation. PPAR␣ null mice the in vivo expression of human CYPs and would enable
have been constructed.55,56 Upon treatment with the known determination of the role of specific human CYPs in chemi-
peroxisome proliferator clofibrate, no proliferator plei- cal metabolism, toxicity, and carcinogenesis. A transgenic
otropic response was observed indicating that PPAR␣ is mouse line expressing the human CYP1A2 in the pancreas
involved in this response.55 Thus, mice lacking PPAR␣ are was established.62 The level of CYP1A2 expressed in pancre-
refractory to induction of genes encoding fatty acid meta- atic microsomes from transgenic mice was comparable to
bolizing enzymes, such as Cyp4a, peroxisome proliferation that of the endogenously expressed CYP1A2 in liver. In
and carcinogenesis.55 Moreover, short-term starvation of terms of activity, the transgenic pancreas exhibited one-
these PPAR␣-null mice caused hepatic steatosis, myocardial third to one-half of estrone oxidation by mouse liver micro-
lipid accumulation, and hypoglycemia, a phenotype similar somes from wild-type animals. Humanized mice containing
to that of humans with genetic defects in mitochondrial CYP4B1 in the liver were also constructed.63 Hepatic micro-
fatty acid oxidation enzymes.56 Thus, PPAR␣ has a critical somes from these mice catalyzed the ␻-hydroxylation of
role in the transcriptional regulatory response to fasting and laurate and also activated 2-aminofluorene. These activities
the PPAR␣-null mice may be useful as a model for abnor- were not observed in control mice. Additionally, a trans-
malities of human fatty acid utilization. Unfortunately, this genic mouse line expressing CYP3A7, the human fetal form of
The Pharmacogenomics Journal

Recent advances in P450 research
JL Raucy and SW Allen
185
CYP3A, has been developed.64 Collectively, these mouse lines drug chemical elimination. While in vitro systems now play a
provide useful tools to study human CYPs and their relation major role in determining xenobiotic metabolism, in vivo
to chemical toxicity, carcinogenesis and teratogenesis.
models, such as the transgenic mouse, are increasingly being
In addition to transgenic animals, humanized transgenic evaluated with the hope of better predicting human responses
plants have been established.65,66 Plants containing human to therapeutic agents.
CYPs can be used for phytoremediation of soil and
REFERENCES
groundwater pollution. For example, plants containing
1
2
3
4
Omura T. Forty years of cytochrome P450. Biochem Biophys Res Com-
mun 1999; 266: 690–698.
CYP2E1 exhibit a profound increase in metabolism of a
common contaminant, tricholethylene (TCE), up to 640-
fold above null plants.66 These transgenic plants could be
used for efficient remediation of many sites contaminated
with halogenated hydrocarbons. Plants containing other
CYPs such as, CYP1A1, CYP2B6 or CYP2C19 are also useful
for breeding of herbicide-tolerant crops as well as for phyto-
remediation of environmental contaminants.65
Stegeman JJ, Livingstone DR. Forms and functions of cytochrome
P450. Comp Biochem Physiol [C] 1998; 121: 1–3.
Nelson DR. Cytochrome P450 and the individuality of species. Arch
Biochem Biophys 1999; 369: 1–10.
Ingelman-Sundberg M, Oscarson M, McLellan RA. Polymorphic human
cytochrome P450 enzymes: an opportunity for individualized drug
treatment. Trends Pharmacol Sci 1999; 20: 342–349.
5
6
Podust L, Poulos T, Waterman M. Crystal structure of cytochrome P450
14␣-sterol demethylase (CYP51) from Mycobacterium tuberculosis in
complex with azole inhibitors. Proc Natl Acad Sci USA 2001; 98:
3068–3073.
Peterson JA, Graham SE. A close family resemblance: the importance
of structure in understanding cytochromes P450. Structure 1998; 6:
1079–1085.
CONCLUDING REMARKS
The present review describes some of the most recent and
important advances in the P450 field. Many other areas of
P450 research not discussed here, have progressed and pro-
vided exciting and important new information. Technologi-
cal advances have facilitated our understanding of mech-
anisms of regulation, toxicity and carcinogenesis. New P450
enzymes are currently being identified that exhibit dual
roles; they metabolize xenobiotics and have important
physiological functions. In the drug metabolism area a great
deal of progress has been aimed at identifying new polymor-
phic sites which could be of clinical and toxicological
importance because these polymorphisms are correlated
with the incidence of drug-induced toxicity and environ-
mentally related cancers. In addition, genetic tests are being
developed that will gradually replace probe drugs as the pri-
mary tool for screening populations for CYP polymor-
phisms. Microchip array technology is underway to assay
the P450 profile of an individual in a single hybridization.
Knowledge of this profile would characterize a person’s risk of
adverse drug reactions and possible predisposition to disease.
7
8
Graham SE, Peterson JA. How similar are P450s and what can their
differences teach us? Arch Biochem Biophys 1999; 369: 24–29.
Gotoh O. Substrate recognition sites in cytochrome P450 family 2
(CYP2) proteins inferred from comparative analyses of amino acid and
coding nucleotide sequences. J Biol Chem 1992; 267: 83–90.
Williams PA, Cosme J, Sridhar V, Johnson EF, McRee DE. Mammalian
microsomal cytochrome P450 monooxygenase: structural adaptations for
membrane binding and functional diversity. Mol Cell 2000; 5: 121–131.
9
10 Cosme J, Johnson EF. Engineering microsomal cytochrome P4502C5
to be a soluble, monomeric enzyme—mutations that alter aggre-
gation, phospholipid dependence of catalysis, and membrane binding.
J Biol Chem 2000; 275: 2545–2553.
11 Watkins RE, Wisely GB, Moore LB, Collins JL, Lambert MH, Williams SP
et al. The human nuclear xenobiotic receptor PXR: structural determi-
nants of directed promiscuity. Science 2001; 292: 2329–2333.
12 Evans WE, Relling MV. Pharmacogenomics: translating functional gen-
omics into rational therapeutics. Genome 1999; 286: 487–491.
13 Nebert DW. Suggestions for the nomenclature of human alleles: rel-
evance to ecogenetics, pharmacogenetics and molecular epidemi-
ology. Pharmacogenetics 2000; 10: 279–290.
14 Dover G. Molecular drive in multigene families: how biological novel-
ties arise, spread and are assimilated. Trends Genet 1986; 2: 159–165.
15 Hodgson J, Marshall A. Pharmacogenomics: will the regulators
approve? Nat Biotechnol 1998; 16: 13–15.
The first crystallization of
a mammalian P450 that
occurred recently will significantly advance structure-
activity investigations. It may now be possible to determine
how a substrate will interact with the active site of a given
P450 enzyme. The technology recently developed for crys-
tallizing a mammalian P450 can also provide insights that will
facilitate the crystallization of additional enzymes. Perhaps
one day we may understand how a single enzyme such as
CYP3A4, can metabolize so many diverse compounds. Also
described here is the current use of mice with disrupted P450
genes. These mice are providing appropriate in vivo models for
investigating the role of specific P450s in chemically-mediated
toxicity and carcinogenicity. They will be particularly useful
in the future towards understanding the metabolic basis of
increased or decreased human risks to chemical carcinogenesis
due to polymorphisms in CYP enzymes. Another area that
16 Marshall A. Getting the right drug into the right patient. Nat Biotechnol
1998; 16: 9–12.
17 Howell WM, Jobs M, Gyllensten U, Brookes AJ. Dynamic allele-specific
hybridization: a new method for scoring single nucleotide polymor-
phisms. Nat Biotechnol 1999; 17: 88.
18 Bruning T, Able J, Koch B, Lorenzen K, Harth V, Donat S et al. Real-
time PCR-analysis of the cytocrhome P450 1B1 codon 432-polymor-
phims. Arch Toxicol 1999; 73: 427–436.
19 Scarlett LA, Madani S, Shen DD, Ho RJY. Development and characteriz-
ation of a rapid and comprehensive genotyping assay to detect the
most common variants in cytochrome P450 2D6. Pharm Res 2000; 17:
242–246.
20 Sohda T. Allele-specific polymerase chain reaction for genotyping
human cytochrome P450 2E1. J Clin Lab Anal 1999; 13: 205–208.
21 Shimada T, Tsumura F, Yamazaki H, Guengerich FP, Inoue K. Charac-
terization of ( )-bufuraolol hydroxylation activities in liver microsomes
of Japanese and Caucasian subjects genotyped for CYP2D6. Pharmaco-
genetics 2001; 11: 143–156.
may potentially evolve from CYP gene targeting is the bioengi- 22 Wedlund PJ. The CYP2C19 enzyme polymorphism. Pharmacology
2000; 61: 174–183.
23 de Morais SM, Wilkinson GR, Blaisdell J, Meyer UA, Nakamura K,
neering of additional mice that expresses human CYPs, phase
II enzymes, or drug transporters. These animal models will
provide an in vivo system more similar to the human situation
that will be beneficial to assessing the role of each protein in
Goldstein JA. Identification of a new genetic defect responsible for the
polymorphism of (S)-mephenytoin metabolism in Japanese. Mol Phar-
macol 1994; 46: 594–598.
www.nature.com/tpj

Recent advances in P450 research
JL Raucy and SW Allen
186
24 Ibeanu GC, Blaisdell J, Ferguson RJ, Ghanayem BI, Brosen K, Benhamou
S et al. A novel transversion in the intron 5 donor splice junction of
CYP2C19 and a sequence polymorphism in exon 3 contribute to the
poor metabolizer phenotype for the anticonvulsant drug S-mepheny-
toin. J Pharmacol Exp Ther 1999; 290: 635–640.
25 Ferguson RJ, de Morais SMF, Benhamou S, Bouchardy C, Blaisdell J,
Ibeanu G et al. A new genetic defect in human CYP2C19. Mutation of
the initiation codon is responsible for poor metabolism of S-mepheny-
toin. J Pharmacol Exp Ther 1998; 284: 356–361.
26 Ibeanu GC, Blaisdell J, Ghanayem BI, Beyeler C, Benhamou S, Bouch-
ardy C et al. An additional defective allele, CYP2C19*5, contributes to
the S-mephenytoin poor metabolizer phenotype in Caucasians. Pharm-
acogenetics 1998; 8: 129–135.
27 Ibeanu GC, Goldstein JA, Meyer U, Benhamou S, Bouchardy C, Dayer
P et al. Identification of new human CYP2C19 alleles (CYP2C19*6 and
CYP2C19*2B) in a Caucasian poor metabolizer of mephenytoin. J Phar-
macol Exp Ther 1998; 286: 1490–1495.
45 Heidel SM, MacWilliams PS, Baird WM, Dashwood M, Buters JTM,
Gonzalez FJ et al. Cytrochrome P4501B1 mediates induction of bone
marrow cytoxocity and preleukemia cells in mice treated with 7,12-
dmethylbenz[a]anthracene. Cancer Res 2000; 60: 3454–3460.
46 Buters JTM, Sakai S, Richter T, Pineau T, Alexander DL, Savas U et al.
Cytochrome P450 CYP1B1 determines susceptibility to 7,12-dimethyl-
benz[a]anthracene-induced lymphomas. Proc Natl Acad Sci USA 1999;
96: 1977–1982.
47 Lee SST, Buters JTM, Pineau T, Fernandez-Salguero P, Gonzalez FJ. Role
of CYP2E1 in the hepatotoxicity of acetaminophen. J Biol Chem 1996;
271: 12063–12067.
48 Valentine CR, Valentine JL, Seng J, Leakey J, Casciano D. The use of
transgenic cell lines for evaluating toxic metabolites of carbamazepine.
Cell Biol Toxicol 1996; 12: 155–165.
49 Raucy JL, Lasker JM, Lieber CS, Black M. Acetaminophen activation by
human liver cytochromes P450IIE1 and P450IA2. Arch Biochem Biophys
1989; 271: 270–283.
28 Nunoya K, Yokoi T, Takahashi Y, Kimura K, Kinoshita M, Kamataki T.
Homologous unequal cross-over within the human CYP2A gene cluster as
a mechanism for the deletion of the entire CYP2A6 gene associated with
the poor metabolizer phenotype. J Biochem (Tokyo) 1999; 126: 402–407.
29 Ariyoshi N, Sawamura Y, Kamataki T. A novel single nucleotide poly-
morphism altering stability and activity of CYP2A6. Biochem Biophys
Res Commun 2001; 281: 810–814.
30 Kuehl P, Zhang J, Lin Y, Lamba J, Mahfoud A, Schuetz J et al. Sequence
diversity in CYP3A promoters and characterization of the genetic basis
of polymorphic CYP3A5 expression. Nature Genet 2001; 27: 383–391.
31 Sullivan-Klose TH, Ghanayem BI, Bell DA, Zhang ZY, Kaminsky LS,
Shenfield GM et al. The role of the CYP2C9-Leu359 allelic variant in the
tolbutamide polymorphism. Pharmacogenetics 1996; 6: 341–349.
32 Kitagawa K, Kunugita N, Kitagawa M, Kawamoto T. CYP2A6*6, a novel
polymorphism in cytochrome P4502A6, has a single amino acid substi-
tution (R128Q) that inactivates enzymatic activity. J Biol Chem 2001;
276: 17830–17835.
33 Pitarque M, Von Richter O, Oke B, Berkkan H, Oscarson M, Ingelman-
Sunberg M. Identification of a single nucleotide polymorphism in the
TATA box of the CYP2A6 gene: impairment of its promoter activity.
Biochem Biophys Res Commun 2001; 284: 455–460.
34 Dai D, Blaisdell J, Ghahayem B, Anders E-M. Genetic polymorphism of
human CYP2C8 and their effects on metabolism of anticancer drug
paclitaxel. FASEB J 2000; 14: A1338.
35 Ariyosh N, Miyazaki M, Toide K, Swamura Y, Kamataki T. A single
nucleotide polymorphism of CYP2B6 found in Japanese enhances cata-
lytic activity by autoactivation. Biochem Biophys Res Commun 2001;
281: 1256–1260.
36 Gonzalez FJ. The study of xenobiotic-metabolizing enzymes and their
role in toxicity in vivo using targeted gene disruption. Toxicol Lett
1998; 103: 161–166.
37 Buters JTM, DoehmerJ, Gonzalez FJ. Cytochrome P450-null mice. Drug
Metab Rev 1999; 31: 437–447.
38 Westphal H. Transgenic mammals and biotechnology. FASEB J 1989;
3: 117–120.
50 Zaher H, Buters JTM, Ward JM, Bruno MK, Lucas AM, Stern ST et al.
Protection against acetaminophen toxicity in CYP1A2 and CYP2E1
double-null mice. Toxicol Appl Pharmacol 1998; 152: 193–199.
51 Shimizu Y, Nakatsuru Y, Ichinose M, Takahashi Y, Kume H, Mimura J
et al. Benzo[a]pyrene carcinogenicity is lost in mice lacking the aryl
hydrocarbon receptor. Proc Natl Acad Sci USA 2000; 97: 779–782.
52 Fernandez-Salguero P, Pineau T, Hilbert DM, McPhail T, Lee S, Kimura
S et al. Immune system impairment and hepatic fibrosis in mice lacking
the dioxin-binding Ah receptor. Science 1995; 268: 722–726.
53 Gonzalez FJ, Fernandez-Salguero P. Anthony YH Lu commemorative
issue. The aryl hydrocarbon receptor. Studies using the AhR-null mice.
Drug Metab Dispos 1998; 26: 1194–1198.
54 Fernandez-Salguero P, Hilbert D, Rudikoff S, Ward J, Gonzalez FJ. Aryl-
hydrocarbon receptor-deficient mice are resistant to 2,3,7,8-tetrachlo-
rodibenzo-p-dioxin toxicity. Toxicol Appl Pharmacol 1996; 140: 173–179.
55 Lee SS, Gonzalez FJ. Targeted disruption of the peroxisome prolifer-
ator-activated receptor alpha gene, PPAR alpha. Ann NY Acad Sci 1996;
804: 524–529.
56 Leone TC, Weinheimer CJ, Kelly DP. A critical role for the peroxisome
proliferator-activated receptor
␣ (PPAR␣) in the cellular fasting
response: the PPAR␣-null mouse as a model of fatty acid oxidation
disorders. Proc Natl Acad Sci USA 1999; 96: 7473–7478.
57 Palmer CNA, Hsu MH, Griffin KJ, Raucy JL, Johnson EF. Peroxisome
proliferator activated receptor-␣ expression in human liver. Mol Pharm-
acol 1998; 53: 14–22.
58 Kliewer SA, Moore JT, Wade L, Staudinger JL, Watson MA, Jones SA et
al. An orphan nuclear receptor activated by pregnanes defines a novel
steroid signaling pathway. Cell 1998; 92: 73–82.
59 Synold TW, Dussault I, Forman BM. The orphan nuclear receptor SXR
coordinately regulates drug metabolism and efflux. Nat. Med 2001; 7:
584–590.
60 Xie W, Barwick JL, Downes M, Blumberg B, Simon CM, Nelson MC et
al. Humanized xenobiotic response in mice expressing nuclear recep-
tor SXR. Nature 2000; 406: 435–439.
61 Xie W, Barwick J, Simon C, Pierce AM, Safe S, Blumberg B et al.
Reciprocal activation of xenobiotic response genes by nuclear recep-
tors SXR/PXR and CAR. Genes Dev 2000; 14: 3014–3023.
62 Ueno T, Tamura S, Frels WI, Shou M, Gonzalez FJ, Kimura S. A trans-
genic mouse expressing human CYP1A2 in the pancreas. Biochem
Pharmacol 2000; 60: 857–863.
63 Imaoka S, Hayashi K, Hiroi T, Yabusaki Y, Kamataki T, Funae Y. A trans-
genic mouse expressing human CYP4B1 in the liver. Biochem Biophys
Res Commun 2001; 284: 757–762.
39 McKinnon RA, Nebert DW. Cytochrome P450 knockout mice: new
toxicological models. Clin Exp Pharmacol Physiol 1998; 25: 783–787.
40 Ghanayem BI, Wang H, Sumner S. Using cytochrome P450 gene
knock-out mice to study chemical metabolism, toxicity, and carcino-
genicity. Toxicol Pathol 2000; 28: 839–850.
41 Pineau T, Fernandez-Salguero P, Lee SST, McPhail T, Ward JM, Gonza-
lez FJ. Neonatal lethality associated with respiratory distress in mice
lacking cytochrome P450 1A2. Proc Natl Acad Sci USA 1995: 92:
5134–5138.
64 Li Y, Yokoi T, Kitamura R, Sasaki M, Gunji M, Katsuki M et al. Establish-
ment of transgenic mice carrying human fetus-specific CYP3A7. Arch
Biochem Biophys 1996; 329: 235–240.
65 Inui H, Kodama T, Ohkawa Y, Ohkawa H. Herbicide metabolism and
cross-tolerance in transgenic potato plants co-expressing human
CYP1A1, CYP2B6, and CYP2C19. Pestic Biochem Physiol 2000; 66:
116–129.
66 Doty SL, Shang TQ, Wilson AM, Tangen J, Westergreen AD, Newman
LA et al. Enhanced metabolism of halogenated hydrocarbons in trans-
genic plants containing mammalian cytochrome P450 2E1. Proc Natl
Acad Sci USA 2000; 97: 6287–6291.
42 Dalton TP, Dieter MZ, Matlib RS, Childs NL, Shertzer HG, Genter MB
et al. Targeted knockout of cyp1a1 gene does not alter hepatic consti-
tutive expression of other genes in the mouse [Ah] battery. Biochem
Biophys Res Commun 2000; 267: 184–189.
43 Schmidt JV, Su GH-T, Reddy JK, Simon M, Bradfield CA. Characteriz-
ation of a murine Ahr null allele: involvement of the Ah receptor in
hepatic growth and development. Proc Natl Acad Sci USA 1996; 93:
6731–6736.
44 Liang HCL, Li H, McKinnon RA, Duffy JJ, Potter SS, Puga et al.
Cyp1a2(−/−) null mutant mice develop normally but show deficient
drug metabolism. Proc Natl Acad Sci USA 1996; 93 1671–1676.
The Pharmacogenomics Journal