DƯỢC LÍ Goodman & Gilman's The Pharmacological Basis of Therapeutics 12th, 2010

Regulation of Transporter Expression. Transporter
expression can be regulated transcriptionally in response
to drug treatment and pathophysiological conditions,
resulting in induction or down-regulation of transporter
mRNAs. Recent studies have described important roles
of type II nuclear receptors, which form heterodimers
with the 9-cis-retinoic acid receptor (RXR), in regulating
drug-metabolizing enzymes and transporters (see
Table 6–4 and Figure 6–12) (Kullak-Ublick et al., 2004;
Wang and LeCluyse, 2003). Such receptors include
pregnane X receptor (PXR/NR1I2), constitutive
androstane receptor (CAR/NR1I3), farnesoid X receptor
(FXR/ NR1H4), PPARα (peroxisome proliferator-activated
receptor α), and retinoic acid receptor (RAR).
Except for CAR, these are ligand-activated nuclear
receptors that, as heterodimers with RXR, bind specific
elements in the enhancer regions of target genes. CAR
has constitutive transcriptional activity that is antagonized
by inverse agonists such as androstenol and
androstanol and induced by barbiturates. PXR, also
referred to as steroid X receptor (SXR) in humans, is
activated by synthetic and endogenous steroids, bile
acids, and drugs such as clotrimazole, phenobarbital,
rifampicin, sulfinpyrazone, ritonavir, carbamazepine,
phenytoin, sulfadimidine, paclitaxel, and hyperforin (a
constituent of St. John’s wort). Table 5–1 summarizes
the effects of drug activation of type II nuclear receptors
on expression of transporters. The potency of activators
of PXR varies among species such that rodents are not
necessarily a model for effects in humans. There is an
overlap of substrates between CYP3A4 and P-glycoprotein,
and PXR mediates coinduction of CYP3A4 and
P-glycoprotein, supporting their synergy in efficient
detoxification.
DNA methylation is one of the mechanisms underlying the
epigenetic control of gene expression. Reportedly, the tissue-selective
expression of transporters is achieved by DNA methylation (silencing
in the transporter-negative tissues) as well as by transactivation
in the transporter-positive tissues. Transporters subjected to epigenetic
control include OAT3, URAT1, OCT2, Oatp1b2, Ntcp, and
PEPT2 in the SLC families; and MDR1, BCRP, BSEP, and
ABCG5/ABCG8 (Aoki et al., 2008; Imai et al., 2009; Kikuchi et al.,
2006; Turner et al., 2006; Uchiumi et al., 1993).
MOLECULAR STRUCTURES
OF TRANSPORTERS
Predictions of secondary structure of membrane transport
proteins based on hydropathy analysis indicate that
membrane transporters in the SLC and ABC superfamilies
are multi-membrane-spanning proteins. A typical
predicted secondary structure of the ABC transporter
MRP2 (ABCC2) is shown in Figure 5–6. However,
understanding the secondary structure of a membrane
transporter provides little information on how the
transporter functions to translocate its substrates. For
this, information on the tertiary structure of the transporter
is needed, along with complementary molecular
information about the residues in the transporter that
are involved in the recognition, association, and dissociation
of its substrates. X-ray diffraction data on representative
membrane transporters illustrate some basic
structural properties of membrane transporters.
ABC Transporter Crystal Structures. To date, four full ABCs have
been crystallized; three are importers and one is an exporter reminiscent
of human ABC transporters (Figure 5–7). The importers are the
vitamin B 12
transporter BtuCD from E. coli (Locher et al., 2002),
the metal-chelate-type transporter HI1470/1 from H. influenzae
(Pinkett et al., 2007), and the molybdate/tungstate transporter
ModBC from A. fulgidus (Hollenstein et al., 2007). The exporter is
Sav1866, a multidrug resistance transporter from S. aureus (Dawson
et al., 2006, 2007). The nucleotide-binding domains (NBDs), which
are present in the cytoplasm, are considered the motor domains of
ABC transporters and contain conserved motifs (e.g., Walker-A
motif, ABC signature motif) that participate in binding and hydrolysis
of ATP. Crystal structures of all four full ABC transporters show
two NBDs, which are in contact with each other, and a conserved
fold. The transmembrane domains of Sav1866 serve as a good model
for the basic architecture of human ABC transporters. Note how the
transmembrane domains of Sav1866 extend into the cytoplasm and
how in the observed crystal structure, the two major bundles are visible
at the extracelluar surface. The mechanism, shared by these ABC
transporters, appears to involve binding of ATP to the NBDs, which
subsequently triggers an outward-facing conformation of the transporters.
Dissociation of the hydrolysis products of ATP appears to
result in an inward-facing conformation. In the case of drug extrusion,
when ATP binds, the transporters open to the outside, releasing
their substrates to the extracellular media. Upon dissociation of the
hydrolysis products, the transporters return to the inward-facing conformation,
permitting the binding of ATP and substrate.
Lactose Permease Symporter (LacY). Lactose permease is a
bacterial transporter that belongs to the major facilitator superfamily
(MFS). This transporter is a proton-coupled symporter. A highresolution
X-ray crystal structure has been obtained for the protonated
form of a mutant of LacY (C154G) (Abramson et al., 2003)
(Figure 5–8). LacY consists of two units of six membrane-spanning
α-helices. The crystal structure locates substrate at the interface of
the two units and in the middle of the membrane. This location is
consistent with an alternating-access transport mechanism in which
the substrate recognition site is accessible to the cytosolic and then
the extracellular surface but not to both simultaneously (Figure 5–9).
Eight helices form the surface of the hydrophilic cavity, and each
contains proline and glycine residues that result in kinks in the cavity.
From LacY, we now know that, as in the case of MsbA, six membrane-spanning
α-helices are critical structural units for transport
by LacY.
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CHAPTER 5
MEMBRANE TRANSPORTERS AND DRUG RESPONSE