Current Drug Metabolism, Volume 2, Number 4, 2001
Membrane Transport
Heterodimeric Amino Acid Transporters:
Molecular Biology and Pathological and Pharmacological Relevance Pp. 339-354
Yoshikatsu Kanai and Hitoshi Endou
Contribution of Specific Transport Systems to
Anthracycline Transport in Tumor and Normal Cells Pp. 355-366
Kazuki Nagasawa, Katsuhito Nagai, Noriaki
Ohnishi, Teruyoshi Yokoyama, and Sadaki Fujimoto
The MRP Family and Anticancer Drug Metabolism Pp. 367-377
T. Suzuki, K. Nishio and S. Tanabe
Effect of Methylxanthine Derivatives on
Doxorubicin Transport and Antitumor Activity Pp. 379-395
A. Kakuyama (nee Iwazaki) and Y. Sadzuka
Interactions of Liposomes with Cells In Vitro
and In Vivo: Opsonins and Receptors Pp. 397-409
Tatsuhiro Ishida, Hideyoshi Harashima and
Hiroshi Kiwada
Application of an In Vivo Brain Microdialysis
Technique to Studies of Drug Transport Across the Blood-Brain Barrier Pp. 411-423
Y. Deguchi and K. Morimoto
[Back to top] Heterodimeric Amino Acid Transporters:
Molecular Biology and Pathological and Pharmacological Relevance
Yoshikatsu Kanai and Hitoshi Endou
In the last decade, a lot of amino acid transporters were
identified by molecular cloning and assigned to the classically characterized
amino acid transport systems. Among them, ones, which belong to the
heterodimeric amino acid transporter family, are unique because of their broad
substrate selectivity and their pathological implications as well as their
structural features. The heterodimeric amino acid transporter family is a
subfamily of SLC7 solute transporter family, which includes 14-transmembrane
cationic amino acid transporters as well as 12-transmembrane heterodimeric
amino acid transporters. The members of heterodimeric amino acid transporter
family are linked via a disulfide bond to single membrane spanning type II
membrane glycoproteins such as 4F2hc (4F2 heavy chain) and rBAT (related to b0, +-amino acid
transporter). Six members are associated with 4F2hc and one is linked to rBAT.
The neutral amino acid transporter of this family seems to rely on the
hydrophobic interactions for their substrate recognition, which can explain
their broad substrate selectivity. Because of this characteristic, they can
permeate amino-acid-related drugs and contribute to the pharmacokinetics of
these drugs. A neutral amino acid transporter LAT1 (L-type amino acid
transporter 1) has actually been shown to be present at the
blood-brain-barrier. Because the members of the heterodimeric amino acid
transporter family exhibit variety of substrate selectivity, it is proposed
that these family members have been diverged from the prototype neutral amino
acid transporter such as LAT1 by acquiring the mechanisms for the recognition
of electric charges on the substrate amino acid side chains. The dysfunction or
hyperfunction of the members of the heterodimeric amino acid transporter family
are involved in some diseases and pathologic conditions. The genetic defects of
the renal and intestinal transporters BAT1/b0,+AT (b0,+-type amino acid transporter
1/b0,+-type
amino acid transporter) and y+LAT1 (y+L-type amino acid transporter 1) result in the amino
aciduria with sever clinical symptoms such as cystinuria and lysinuric protein
intolerance, respectively. LAT1 is proposed to be involved in the progression
of malignant tumor. xCT (x-C-type transporter) functions to protect cells against
oxidative stress, while its over-function may be damaging neurons leading to
the exacerbation of brain damage after brain ischemia. Therefore, these
transporters would be candidates for therapeutic targets based on new
strategies. Through the interaction with the associating proteins, the
transporters of this family would be endowed with more possibility to be regulated
via intracellular and extracellular signalling pathways, which is critical to
tune the transporter functions to meet the metabolic requirements of cells.
[Back to top] Contribution of Specific Transport Systems to Anthracycline
Transport in Tumor and Normal Cells
Kazuki Nagasawa, Katsuhito Nagai, Noriaki
Ohnishi, Teruyoshi Yokoyama, and Sadaki Fujimoto
Anthracycline antibiotics are very effective neoplastic
agents widely used clinically. However, because of their many adverse effects
(e.g. cardiotoxicity, leukopenia and alopecia), their clinical use has been
limited. In order to minimize their adverse effects in clinical cancer
chemotherapy, anthracyclines must be selectively transported into tumor cells.
If there are differences in transport characteristics between tumor and normal
cells, it should be possible to establish a strategy for selectively delivering
anthracyclines to tumor cells on the basis of the differences. In human
cultured leukemia HL60 cells, as tumor cells, and human fresh mononuclear
cells, as normal cells, doxorubicin, pirarubicin, daunorubicin and idarubicin
were incorporated via a common carrier-mediated system, but the carriers were
different in the two cell types. In HL60 cells, it was indicated that a
nucleoside transport system contributed, at least in part, to the transport of
doxorubicin and pirarubicin, but not daunorubicin and idarubicin, and its
contribution to pirarubicin transport was found in other tumor cells, i.e.
mouse ovarian sarcoma M5076 and Ehrlich ascites carcinoma cells. On the other
hand, in mononuclear cells, there was no involvement of a nucleoside transport
system for the four anthracyclines examined. Therefore, we thought that with
the modification of an anthracycline molecule as a substrate for the nucleoside
transport system, the anthracycline could be delivered selectively to tumor
cells.
[Back to top] The MRP Family and Anticancer Drug Metabolism
T. Suzuki, K. Nishio and S. Tanabe
Acquirement of drug resistance by tumor cells is a major
chemotherapeutic problem. It is well known that typical multidrug resistance is
caused by P-glycoprotein and multidrug resistance related protein (MRP1) which
belong to the ATP binding cassette (ABC) transporter family. Ishikawa proposed
that the ATP-dependent glutathione-S-conjugate export pump (GS-X pump) and
phase III detoxification system are essential to drug metabolism, and this
constituted a new concept in drug metabolism and the detoxification of xenobiotics.
The GS-X pump has been revealed to belong to the ABC transporter family and
suggested to the contribution to anticancer drug resistance. The GS-X pump
actively effluxes the glutathione S-platinum (GS-Pt) complex. We cloned novel
ABC transporter cDNA from the PC-14/CDDP cell line, and the cloned cDNA was
designated as a short-type MRP homologue, SMRP. Further investigation suggested
that SMRP is a splicing variant of MRP5. The MRP5 mRNA levels in tumors from
lung cancer patients treated with platinum regimen were significantly higher
than in tumors from patients treated with non-platinum regimens, and the MRP5
expression levels were correlate with the GCS expression levels that is the
rate-limiting step enzyme in glutathione biosynthesis. These results suggested
that MRP5 take part in the function of GS-X pump. Recently many transporter
molecules belong to the ABC transporter family such as MRP family have been
identified, and appear to express in various human tissues. It can be presumed
that their molecules are affected by the disposition and metabolism of drugs,
but their substrates are still unclear. If the substrate specificity is
revealed in the future, it is expected that the anticancer agents transporter,
moreover anti cancer drug resistance mechanisms, can be clarified. This review
is cited in the cisplatin resistance and the GS-X pump, and finally describes
an overview of the MRPs substrates recently clarified, mainly about anticancer
drugs.
[Back to top] Effect of Methylxanthine Derivatives on
Doxorubicin Transport and Antitumor Activity
A. Kakuyama (nee Iwazaki) and Y. Sadzuka
Biochemical modulation, which is more effective with the use
of antitumor agents, has recently played very important role in cancer
chemotherapy. In this review, it was reported that some of the methylxanthine
derivatives, e.g. caffeine, were useful for modulator and attempted to defined
the relation between the effect of methylxanthine derivatives on the
doxorubicin transport and antitumor activity. Caffeine and theobromine
inhibited the doxorubicin efflux from tumor cells, increased the doxorubicin
concentration in a tumor, and enhanced the antitumor effect of doxorubicin.
However, the caffeine metabolites, which had no effect on the doxorubicin
efflux, did not increase antitumor activity. Moreover, caffeine and theobromine
did not enhance the side toxicity of doxorubicin on the lipid peroxide level,
DNA biosynthesis and the doxorubicin concentrations in normal tissues.
Moreover, we investigated the effect of the combination of
doxorubicin with caffeine or theobromine on the change in cyclic adenosine
3',5'-monophosphate (cyclic AMP) in tissues in vivo, and the effect of cyclic
AMP on doxorubicin efflux in vitro, and measured the distribution of caffeine
and theobromine in normal and tumor tissues. In Ehrlich ascites carcinoma
bearing mice, the level of cyclic AMP in a tumor was decreased by doxorubicin.
With the combination of caffeine or theobromine and doxorubicin, the cyclic AMP
level recovered to the control level. This tendency was not seen in normal
tissues (heart and liver). Moreover, the doxorubicin efflux from the Ehrlich
cells was inhibited on the addition of cyclic AMP in vitro. And the caffeine
concentration in the tumors was the same as that in the heart, and was
increased in combination with doxorubicin compared with that in the
caffeine-only group during the 4 hr after caffeine treatment. Furthermore, the
doxorubicin efflux was promoted by the supply of energy (addition of glucose),
influx was decreased relatively, doxorubicin efflux needs the existence of
glucose and the inhibition of energy related drug export pump by caffeine
induced inhibition of doxorubicin efflux. The treatment of doxorubicin nor
caffeine, and any treatment schedule did not change the amount and appearance
of GLUT 1 as glucose transporter on Ehrlich ascites carcinoma cell.
For the mentioned above, we thought as concerns the increase
of antitumor activity of doxorubicin by caffeine which is xanthine derivatives
as follows. Caffeine distributes, high level in tumor, keeps the cyclic AMP
level, and effects glucose transport or doxorubicin transport depend on energy
and inhibits doxorubicin efflux. And then DNA synthesis was increased with the
maintenance the concentration of doxorubicin in tumor. These action did not
show in normal tissues, caffeine did not influence the side toxicity of
doxorubicin. These results suggested that caffeine which is one of xanthine
derivatives will be useful for biochemical modulator.
[Back to top] Interactions of Liposomes with Cells In Vitro
and In Vivo: Opsonins and Receptors
Tatsuhiro Ishida, Hideyoshi Harashima and
Hiroshi Kiwada
A number of studies have appeared recently on the underlying
mechanisms of liposome-cell interactions under in vitro conditions, in which
isolated cell populations or cell lines were used. However, our knowledge of
how liposomes interact with cells and the parameters that influence this in
vivo is limited. We will summarize and discuss the relevant studies on this
matter in this article. In addition, researchers in this field have long been
aware of the interaction of liposomes with blood (or serum/plasma) proteins in
vivo and their potential role in the process of the clearance of liposomes from
the circulation. Some of the ‘opsonizing’ proteins, such as complement
components, immunoglobulins, which enhance the interactions of liposomes with
‘phagocytic cells’ have been identified. However, the issue of which types of
opsonins determine the fate of liposomes in vivo and how liposomal
physicochemical properties such as size, charge and fluidity play an important
role in the process of liposome clearance is not clear. Our own observations of
one of opsonins, complement component are reviewed herein.
As opposed to the fate of conventional liposomes, we briefly
touch on the interaction of surface-modified liposomes, which are designed to
avoid interactions with blood proteins and/or cells (sterically stabilized
liposomes, long-circulating liposomes) and to actively target specific cells or
tissues (targeted liposomes: immunoliposomes). Blood proteins such as opsonins
are not usually thought to play an important role in the clearance of such
liposomes.
[Back to top] Application of an In Vivo Brain Microdialysis Technique to Studies of
Drug Transport Across the Blood-Brain Barrier
Y. Deguchi and K. Morimoto
There is a wide range of methods available for studying the transport of drugs across the blood-brain barrier (BBB) which is equipped with several systems to transport drugs as well as endogenous nutrients and waste products. The in vivo brain microdialysis technique, which allows direct sampling of the brain interstitial fluid (ISF), is a powerful means of characterizing influx and efflux transport across the BBB. In this paper, we review our results from the successful application of this technique to BBB drug transport studies. The drugs investigated include novel and CNS-active peptides, some agents that are actively removed from the brain ISF across the BBB, and a brain-directed prodrug.