Nuclear Medicine and Biology 30 (2003) 31–37

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Characterization of 3-[125I]iodo-␣-methyl-L-tyrosine transport via
human L-type amino acid transporter 1
Naoto Shikanoa,*, Yoshikatsu Kanaib, Keiichi Kawaic, Nobuyoshi Ishikawaa, Hitoshi EndoubaDepartment of Radiological Sciences, Ibaraki Prefectural University of Health Sciences, 4669-2 Ami Ami-machi, Inashiki-gun,Ibaraki 300-0394 Japanb
Department of Pharmacology and Toxicology, Kyorin University School of Medicine, 6-20-2 Shinkawa, Mitaka, Tokyo 181-8611, Japanc
School of Health Sciences, Faculty of Medicine, Kanazawa University, 5-11-80 Kodatsuno, Kanazawa-city, Ishikawa 920-0942, Japan
Received 30 April 2002; received in revised form 6 June 2002; accepted 29 June 2002

Abstract
We examined transport of 3-[125I]iodo-␣-methyl-L-tyrosine ([125I]IMT) in Xenopus laevis oocytes co-expressing human L-type aminoacid transporter 1 (a component of system L) and human 4F2hc. Human LAT1 mediated transport of [125I]IMT. [125I]IMT uptake wasdecreased by the presence of L-isomers of Cys, Leu, Ileu, Phe, Met, Tyr, His, Trp and Val and D-isomers of Leu, Phe and Met. Human
LAT1-mediated [125I]IMT uptake was highly stereoselective for the L-isomers of Tyr, His, Trp, Val and Ileu. To examine the effects of
3-iodination and ␣-methylation on IMT transport, kinetic parameters of IMT were compared with those of mother Tyr and 3-[125I]iodoL-tyrosine (3-I-Tyr). Uptake of Tyr, 3-I-Tyr and [125I]IMT followed Michaelis-Menten kinetics, with Km values of 29.0 ⫾ 5.1, 12.6 ⫾ 6.1
and 22.6 ⫾ 4.1 ␮M, respectively. Neither the ␣-methyl group nor the size of the 3-iodinated Tyr residue was an obstacle to transport via
hLAT1. Furthermore, affinity of IMT for hLAT1 is higher than that of the natural parent tyrosine. The level of efflux mediated by hLAT1
was highly stimulated by extracellularly applied L-Leu, suggesting exchange of [125I]IMT and L-Leu via hLAT1. © 2002 Elsevier Science
Inc. All rights reserved.
Keywords: Membrane transport; 3-[123I]Iodo-␣-methyl-L-tyrosine; Artificial amino acid; L-type amino acid transporter 1; 4F2hc; Xenopus laevis oocyte

1. Introduction
3-[123I]Iodo-␣-methyl-L-tyrosine ([123I]IMT, Fig. 1) was
developed as a functional imaging agent for Tyr transportmechanisms in the brain [1] and pancreas [2]. It is a nonmetabolizable artificial amino acid that has also been usedclinically for SPECT imaging of tumors [3].
In cultured type 86HG-39 human glioma cells, membrane transport of [123I]IMT is dominated by
2-aminobicyclo[2.2.1]heptane-2-carboxylic acid (BCH)sensitive transport; i.e., amino acid transport system L [4].
The kinetics of [123I]IMT transport have also been studied
in rat C6 glioma cells [5], human GOS3 glioma cells [6] andhuman Ewing’s sarcoma cells [7]. The studies of [123I]IMT
uptake in glioma cells revealed that the majority of transporttakes place via the Na⫹-independent system L (⬎ 70%) andthat relatively minor uptake takes place via the Na⫹-depen* Corresponding author. Tel.: ⫹81-298-40-2217; fax: ⫹81-298-402317.
E-mail address: sikano@ipu.ac.jp (N. Shikano).

dent system B0,⫹ (⬍ 20%). Lahoutte et al. reported thatsystem T also mediates [123I]IMT transport into U266 human myeloma cells [8].
The transport of large neutral amino acids with branchedor aromatic side chains is mediated by amino acid transportsystem L [9]. System L is a Na⫹-independent neutral aminoacid transport system, and is thought to be a major route bywhich cells obtain branched or aromatic amino acids [9].
System L uptake has been defined as uptake of an aminoacid in Na⫹-free buffer which is subject to inhibition by anexcess of BCH [10].
In 1998, by expression cloning from C6 rat glioma cells,
we isolated cDNA encoding a Na⫹-independent transporterthat acts as a component of amino acid transport system L[11], and named the protein L-type amino acid transporter 1(LAT1). This protein, which has 12 trans-membrane domains, preferentially transports large neutral amino acids
such as Leu, Ile, Val, Phe, Tyr, Trp, Met and His [11]. In theperformance of its function, it forms a heterodimer with asingle membrane-spanning protein called 4F2hc, a type IImembrane glycoprotein [11].

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N. Shikano et al. / Nuclear Medicine and Biology 30 (2003) 31–37

Fig. 1. Chemical structures.

It has been suggested that amino acid transporters are
upregulated in transformed cells to support the high-levelprotein synthesis required for continuous cell growth andproliferation [9]. Some human tumor cell lines (e.g., Hela,
HL-60, KG-1 and U937) express both human LAT1(hLAT1) and the heavy chain of human 4F2 (h4F2hc) at
high levels [12].
In the present study, we examined aspects of transport of[125I]IMT via hLAT1 (uptake, inhibition and kinetics) using
Xenopus laevis oocytes co-expressing hLAT1 and h4F2hc.
To examine the effects of 3-iodination and ␣-methylation
on IMT transport, kinetic parameters of IMT were compared with those of mother Tyr and 3-I-Tyr.

2. Materials and methods
All animal experiments were approved by the ethicalcommittee of our university.
2.1. Preparation of amino acids
Reagent grade chemicals (Sigma-Aldrich, Germany)
were used in this experiment. [125I]NaI (8.1 ⫻ 1019 Bq/mol)
was obtained from Amersham Pharmacia (U.K.). No-carrier-added [125I]IMT and 3-I-Tyr were prepared using theconventional chloramine-T method, as described elsewhere[1,2]. Synthesis of inactive IMT was performed according
to the method of Krummeich et al. [13]. L-[14C(U)]-Tyr was
obtained from American Radiolabeled Chemicals (USA).
2.2. Transport characterization
Human LAT1 (hLAT1) cDNA (GenBank/EMBL/DDBJ
accession no. AB18009) was isolated from an oligo-dTprimed cDNA library prepared from human teratocarci-

noma PA-1 cell poly(A)⫹ RNA (Clontech, USA), using a
fragment corresponding to nucleotides 1135 to 1529 of rat
LAT1 cDNA (GenBank/EMBL/DDBJ accession no.
AB015432) as a probe, as described elsewhere [10,14]. The
amino acid sequence of this hLAT1 cDNA was 100% identical to that of the hLAT1 cDNA sequence published by
Prassad et al. [15].
For functional expression in Xenopus laevis oocytes,
cRNAs for hLAT1 and human 4F2hc (h4F2hc) (GenBank/
EMBL/DDBJ accession no. AB018010) were obtained byin vitro transcription using T3 RNA polymerase for hLAT1
in pBluescript II SK- (Stratagene, USA) linearized with
XhoI and T7 RNA polymerase for human 4F2hc (h4F2hc)
in pZL1 (Life Technologies Inc., USA) linearized with
BamHI, as described elsewhere [16]. Examination of expression in Xenopus laevis oocytes was performed according to the methods of Kanai et al. [17], with minor modifications. Xenopus laevis oocytes were injected with 17.6 ng
of hLAT1 cRNA and 7.4 ng of h4F2hc cRNA at a final
molar ratio of 1 : 1. For expression of only hLAT1 or only
h4F2hc in Xenopus laevis oocytes, 17.6 ng of hLAT1 cRNAor 7.4 ng of h4F2hc cRNA was injected. Two days after
injection, the uptake of radio-labeled amino acids in Na⫹free uptake solution (100 mM choline-Cl, 2 mM KCl, 1 mM
CaCl2, 1 mM MgCl2, 10 mM HEPES, 5 mM Tris, pH 7.4)
containing 18.5 to 74 kBq/ml 125I-labeled amino acids wasmeasured.
For the Na⫹-containing uptake solution, the choline-Cl
present in the Na⫹-free uptake solution was replaced with
NaCl. For the Cl- -free uptake solution, the Cl- present in theNa⫹-containing uptake solution was replaced with gluconate anion. Preliminary experiments to determine thetime-course of no– carrier-added (n.c.a.) [125I]IMT uptake
into oocytes expressing hLAT1 indicated that the uptake
was linearly dependent on incubation time up to 45 min(data not shown). Therefore, for all subsequent experiments,

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33

except for Km and Vmax determination. Results from therepresentative experiments are shown in the figures.

3. Results
3.1. Preparation of amino acids
Labeling of ␣-methyl-L-tyrosine and L-tyrosine produced [125I]IMT and 3-I-Tyr, respectively, at labeling efficiency of more than 80% for both compounds. After purification, no-carrier-added [125I]IMT and 3-I-Tyr withradiochemical purities greater than 95% were obtained. For
both compounds, purification by reverse-phase HPLC
yielded inactive IMT at a rate of 68%.
3.2. Transport characterization

Fig. 2. [125I]IMT uptake into Xenopus laevis oocytes co-expressing hLAT1
and h4F2hc. Co-expression of hLAT1 and h4F2hc resulted in high uptakeof [125I]IMT. *, P⬍0.001.

amino acid uptake rate was measured for 30 min and expressed as pmol/oocyte/min.
Km and Vmax of the substrates Tyr, 3-I-Tyr and IMT
were determined by double reciprocal plot analysis of
hLAT1-mediated amino acid uptake measured at 1, 3, 10,30, 100 and 300 ␮M. hLAT1-mediated amino acid uptake
was calculated as the difference between mean uptake ofoocytes injected with both hLAT1 and h4F2hc cRNAs andmean uptake of control oocytes injected with water. For the
inhibition study, the oocytes were incubated in the presenceof 3 mM of 1 of 21 different natural L- and D-amino acids:
Gly, Ala, Ser, Thr, Cys, Leu, Ileu, Phe, Met, Tyr, His, Trp,
Val, Asn, Gln, Asp, Glu, Lys, Arg, Pro and Cyst.
For the efflux measurement, 100 nl (74 Bq) of [125I]IMT
was injected into oocytes using a fine-tipped glass micropipette, as described elsewhere [18,19]. Oocytes were individually incubated for 5 min in ice-cold Na⫹-free uptake
solution, and then transferred to Na⫹-free uptake solution
with or without 20 ␮fM nonradiolabeled L-leucine maintained at room temperature (18 –22°C). The radioactivity ofthe medium and the residual radioactivity of the oocytes
were measured. The efflux values were expressed as percent
radioactivity (radioactivity of medium/[radioactivity of medium ⫹ radioactivity of oocytes] ⫻ 100%) [18,19].
For the uptake and efflux measurements, 7 to 8 oocytes
were used for each data point. Each data point in the figures
represents the mean⫾standard deviation of uptake (n ⫽7– 8). To confirm the reproducibility of results, 3 separate
experiments using different batches of oocytes and in-vitro–transcribed cRNA were performed for each measurement,

As shown in Fig. 2, hLAT1 requires h4F2hc for functional ([125I]IMT uptake) expression in Xenopus laevis oocytes. The uptake of [125I]IMT by Xenopus laevis oocytes
expressing only hLAT1 was 16-fold greater (p⬍0.001) thanthat of the control oocytes (injected with water instead ofcRNA). Uptake of [125I]IMT by Xenopus laevis oocytes
co-expressing h4F2hc and hLAT1 was 13-fold greater thanthat of oocytes expressing only hLAT1.
The uptake of [125I]IMT by co-expressing oocytes in thepresence and absence of Na⫹ is compared in Fig. 3.

Fig. 3. Na⫹ independence of [125I]IMT transport in oocytes expressing
hLAT1 and h4F2hc. There was no significant difference in uptake of[125I]IMT (n.c.a.) between the standard uptake solution and the Na⫹-free
uptake solution, in which Na⫹ was replaced with the same concentration ofcholine. *, P⬍0.001.

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Fig. 4. The effects of L- and D-amino acids on hLAT1-mediated transport of [125I]IMT. (a) Inhibition of hLAT1-mediated [125I]IMT (n.c.a.) uptake by
L-amino acids in Na⫹-free uptake solution. Human LAT1-mediated [125I]IMT (n.c.a.) uptake was measured in the presence of 3 mM nonradiolabeled
L-amino acids. Values are expressed as percentage of the control value ([125I]IMT uptake in the absence of inhibitors). (b) Inhibition of hLAT1-mediated[125I]IMT (n.c.a.) uptake by D-amino acids. Same experimental procedures as described in (a).

[125I]IMT uptake was not dependent on Na⫹ or Cl- (data notshown).
As shown in Fig. 4 (A), the uptake of [125I]IMT was
markedly inhibited by the presence of Cys, Leu, Ileu, Phe,
Met, Tyr, His, Trp and Val. Ala, Ser, Thr, Asn and Gln had
relatively weaker inhibitory effects on hLAT1-mediated[125I]IMT uptake. As shown in Fig. 4 (B), D-isomers ofLeu, Phe and Met strongly inhibited hLAT1-mediated[125I]IMT uptake. In contrast, hLAT1-mediated [125I]IMT
uptake was highly stereoselective for the L-isomers of Tyr,
His, Trp, Val and Ileu.
The uptake was saturable and fit the Michaelis-Menten
curve (Fig. 5). The kinetic parameters obtained are shown in
Table 1. Uptake of Tyr, 3-I-Tyr and IMT followed Michaelis-Menten kinetics with Km values of 29.0 ⫾ 5.1, 12.6 ⫾6.1 and 22.6 ⫾ 4.1 ␮M (mean ⫾ standard deviation of 4
separate experiments), respectively.
The level of [125I]IMT efflux mediated by hLAT1 was
fairly low. However, as shown in Fig. 6, L-Leu applied
extracellularly greatly stimulated the efflux of [125I]IMT(preloaded substrate). This suggests amino acid exchange
via hLAT1.

4. Discussion
In the first step of their metabolism, neutral amino acids
are taken into cells by membrane transport and pooled as
free amino acids. They are then used as monomers ofstructural proteins, materials for biosynthesis of neurotransmitters, or substrates for energy metabolism [9]. Because of
its high metabolic stability, [125I]IMT is not metabolized
and remains a free amino acid in vivo [1]. This artificial
amino acid [125I]IMT has generated much interest since it
was demonstrated that its uptake specifically reflects increased amino acid membrane transport in gliomas [3,20].
Membrane transport of [123I]IMT has been studied using
cultured human 86HG-39 glioma cells [4], rat C6 glioma
cells [5], human GOS3 glioma cells [6], and human Ewing’s
sarcoma cells [7]. The majority of transport of [123I]IMT inglioma cells involves the Na⫹-independent system L (⬎

N. Shikano et al. / Nuclear Medicine and Biology 30 (2003) 31–37

35

Fig. 5. Concentration dependence of hLAT1-mediated [125I]IMT uptake. The hLAT1-mediated [125I]IMT uptake in oocytes co-expressing hLAT1 and h4F2hc
was measured for 1, 3, 10, 30, 100 and 300 ␮M nonradiolabeled IMT in Na⫹-free uptake solution. The uptake was plotted against IMT concentration. (b)Double reciprocal plot of the IMT uptake, from which kinetic parameters were determined.

70%), and the Na⫹-dependent system B0,⫹ is responsible
for a relatively minor amount of transport (⬍ 20%). The
Michaelis constant Km of [125I]IMT was found to be quite
high (20 – 45 ␮M) in the above-mentioned cell line studies[4,5,6,7]. In U266 human myeloma cells, system L contributes 39.0% of [123I]IMT influx, and system T contributes
43.8% [8]. System L transports amino acids with bulky
and/or branched chains, and has a broad substrate specificity[9,11,12]. System T is more specific, and favors aromatic
amino acids [9]. System T has only been found on themembrane of red blood cells and hepatocytes, whereas system L is present in most cell types [9]. The transport of mostneutral amino acids involves several distinct systems withoverlapping substrate specificities [9]. The main systems
transporting neutral amino acids can be divided into Na⫹dependent transport systems (systems A, ASC, B0,⫹ and B0)
and Na⫹-independent transport systems (systems L and T)[9].
Cell line studies have revealed that systems L, A, T and
B0,⫹ (and/or B0) play a role in [123I]IMT transport [7,8].
However, it has also been shown that the Na⫹-independent

carrier systems b0,⫹ and y⫹ do not play a role in the[123I]IMT uptake process [7,21]. It is unclear whether system y⫹L is involved in [123I]IMT uptake [7].
In the present study, the kinetic parameters determined inTable 1 reveal correlation between structure and transport

Table 1
Kinetic parameters of amino acid substrates
Amino acid

Km (␮M)

Vmax

Tyr
3-I-Tyr
IMT

29.0 ⫾ 5.1
12.6 ⫾ 6.1
22.6 ⫾ 4.1(1.00)
1.60
0.81

Km values ⫽ mean ⫾ standard deviation of 4 separate experiments.

Fig. 6. [125I]IMT efflux via hLAT1. Efflux of radioactivity from oocytes
expressing hLAT1 and h4F2hc in Na⫹-free uptake solution containing 0␮M (hatched columns) or 100 ␮M (black columns) of nonlabeled L-Leu
was measured. Values are expressed as percentage of total radioactivity
loaded into the oocytes. *, P⬍0.001.

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N. Shikano et al. / Nuclear Medicine and Biology 30 (2003) 31–37

function. The molecular weight of Tyr is about 181, and theatomic weight of the stable isotope of iodine is 127. The
atomic weight of iodine accounts for approximately 41%
and 38% of the molecular weight of 3-I-Tyr and IMT,
respectively. Although hLAT1 preferentially transports
neutral amino acids with bulky hydrophobic residues [11,12], we assumed that the weight added by introducing
iodine into the benzene ring of Tyr would decrease theaffinity of [125I]IMT membrane transport via hLAT1. However, our results were clearly contrary to this prediction. Not
only did hLAT1 exhibit high degrees of affinity(Km⫽10 –30 ␮M) for all 3 substrates, Tyr, 3-I-Tyr and IMT,
but it exhibited higher affinity for [125I]IMT than for motherTyr. The present Km value for [125I]IMT agrees with thevalues obtained in previous cell line studies. Thus, hLAT1
mediates transport of [125I]IMT as well as Tyr and 3-I-Tyr,indicating that this transporter has adequate space for the␣-methyl group of [125I]IMT to pass through the transmembrane domains of the protein.
Thus, introduction of hydrophobic iodine into the benzene ring of Tyr results in higher transport affinity. Neitherthe ␣-methyl group nor the size of the 3-iodinated Tyr
residue was an obstacle to transport via hLAT1. In contrast,
hLAT1 was unable to transport a 3,5-diodinated Tyr residue(data not shown).
It has been reported that system L-mediated [123I]IMTtransport and h4F2hc antigen expression significantly correlate with each other, and that both correlate with theproliferation rate of human glioma cells in vitro [22]. As
shown in Fig. 2, hLAT1 requires h4F2hc for functional([125I]IMT uptake) expression in Xenopus laevis oocytes.
The results in Fig. 3 show that the transport of [125I]IMT
was Na⫹-independent. Radioactivity of oocytes injectedwith cRNA significantly increased after uptake incubation,compared to control oocytes. These findings are strong
evidence for the occurrence of hLAT1-mediated transportof [125I]IMT in Xenopus laevis oocytes injected withhLAT1 cRNA.
Because it has been shown that D-amino acids are endogenously present in mammalian tissues [23], both D- andL-amino acids were used as inhibitors. Fig. 4 shows that
[125I]IMT competed for transport with L-isomers of Cys,
Leu, Ileu, Phe, Met, Tyr, His, Trip and Val and D-isomers
of Leu, Phe and Met. High stereoselectivity for Tyr, His,
Trip, Val and Ileu was observed. L-isomers of small neutralamino acids (Gly, Ala, Ser and Thr) slightly inhibited[125I]IMT uptake. Neither basic amino acids (Lys and Arg)
nor acidic amino acids (Glu and Asp) inhibited [125I]IMT
uptake.
As shown in Fig. 6, L-Leu applied extracellularly induced the efflux of [125I]IMT (preloaded substrate). It has
been suggested that hLAT1 acts as an intracellular Gln
exchanger in the influx of substrates [12]. It has been reported that Asn and Gln are transported with relatively low
affinity [12]. However, it has also been reported that thepools of free Asn and Gln in cells are considerably larger

than those of other amino acids, such as Leu, Ileu, Phe, Met,
Tyr, His, Trip and Val [24]. Other amino acid exchangetransporters have been reported [25]; e.g., the cysteine/
glutamate exchange transporter xCT, the cDNA of which
has been cloned [26]. The free Glu pools in cells are also
large [24].
Human LAT1 is expressed in a wide variety of organs.
Its highest expression levels are in the brain, placenta, testis,bone marrow and fetal liver, followed by peripheral leukocytes, lymph nodes and the thymus [12]. In a previous
study, rat LAT1 was found to act as a system L transporterat the blood-brain barrier [27].
Given present and previous findings of experiments using [123I]IMT, this molecule can be expected to facilitate
future research on tumors and cerebral function, and be
useful in SPECT research into amino acid transport.
Of the genes identified thus far by the human genome
project, about 3% are believed to function as membrane or
intracellular transporters. Furthermore, about 30% of currently available clinical drugs target transporters or channels. The goals of membrane transport research include
isolation of all human genes involved in drug transport,
clarification of sites of expression of these genes and distribution of their gene products, identification of the geneproduct substrates, and characterization of inhibitory mechanisms. Further studies of transport of IMT via L-typeamino acid transporters are currently being conducted by
the present authors.

5. Conclusions
We have clarified characteristics of membrane transportof [125I]IMT via the amino acid transporter hLAT1 (a component of system L) using molecular biological methods
and hLAT1 cDNA. [125I]IMT was confirmed to be thesubstrate transported by hLAT1. Uptake and inhibition ofthis compound were examined by co-expressing hLAT1 andh4F2hc in Xenopus laevis oocytes. [125I]IMT uptake wasdecreased by the presence of L-isomers of Cys, Leu, Ileu,
Phe, Met, Tyr, His, Trp and Val and D-isomers of Leu, Phe
and Met. Human LAT1-mediated [125I]IMT uptake was
highly stereoselective for the L-isomers of Tyr, His, Trp,
Val and Ileu. The uptake was Na⫹-independent, and was
activated by h4F2hc. The affinity of hLAT1-mediated transport for [125I]IMT (Km⫽22.6 ␮M) was greater than formother Tyr (Km⫽29.0 ␮M).

Acknowledgments
We would like to thank T. Murakami, M. Kikuchi, M.
Hira, K. Hamazaki, M. Nishikido, M. Hakamada and R.
Sasaki (Ibaraki Prefectural University) for their technicalassistance. We are grateful to J. Inatomi, K.D. Kim (Kyorin
University School of Medicine) for providing cDNA. This

N. Shikano et al. / Nuclear Medicine and Biology 30 (2003) 31–37

work was supported by Grants-in-Aid for Scientific Research (# 10770451, # 14770498 and # 13557075) from the
Ministry of Education, Science, Sports and Culture of Japanand the Japan Society for Promotion of Science. Financial
support was also provided by the IPU Project Research(9808-3, 0118-1 and 0220-1) and Ibaraki Prefectural University Grants-in-Aid for the Encouragement of Young Scientists 2001 and 2002.

References[1] K. Kawai, Y. Fujibayashi, H. Saji, Y. Yonekura, J. Konishi, A.
Kubodera, A. Yokoyama, A strategy for study of cerebral amino acidtransport using iodine-123-labeled amino acid radiopharmaceutical:
3-iodo-alpha-methyl-L-tyrosine, J. Nucl. Med. 32 (1991) 819 – 824.
[2] K. Kawai, Y. Fujibayashi, H. Saji, Y. Yonekura, J. Konishi, A.
Kubodera, A. Yokoyama, An artificial amino acid radiopharmaceutical for single photon emission computed tomographic study ofpancreatic amino acid transports 123I-3-iod-alpha-methyl-L-tyrosine,Ann. Nucl. Med. 6 (1992) 169 –175.
[3] K-J. Langen, N. Roosen, H.H. Coenen, J.T. Kuikka, T. Kuwert, H.
Herzog, G. Stocklin, L.E. Feinendegen, Brain and brain tumor uptakeof L-3-[123I]iodo-alpha-methyl tyrosine: competition with natural Lamino acids, J. Nucl. Med. 31 (1990) 281–286.
[4] K-J. Langen, H. Muhlensiepen, M. Holschbach, H. Hautzel, P.
Jansen, H.H. Coenen, Transport mechanisms of 3-[123I]iodo-␣-methlyl-L–tyrosine in a human glioma cell line: comparison with [3Hmethyl]-L-methionine, J. Nucl. Med. 41 (2000) 1250 –1255.
[5] B. Reimann, F. Stoegbauer, K. Kopka, H. Halffer, M. Lasic, A.
Schirmacher, T. Kuwert, M. Weckesser, B. Ringelstein, O. Schober,
Kinetics of 3-[123I]iodo-L-␣-methlyl tyrosine transport in rat C6 glioma cells, Eur. J. Nucl. Med. 26 (1999) 1274 –1278.
[6] B. Riemann, K. Kopka, F. Stogbauer, H. Halfter, S. Ketteler, T.Q. Vu
Phan, C. Franzius, M. Weckesser, B. Ringelstein, O. Schober, Kinetic
parameters of 3-[123I]iodo-L-␣-methlyl tyrosine [123I]IMT) transport
in human GOS3 glioma cells, Nucl. Med. Biol. 28 (2001) 293–297.
[7] C. Franzius, K. Kopka, F. Valen, V. Eckervogt, B. Riemann, J. Sciuk,O. Schober, Characterization of 3-[123I]iodo-L-␣-methlyl tyrosine
transport ([123I]IMT) transport into human Ewing’s sarcoma cells in
vitro, Nucl. Med. Biol. 28 (2001) 123–128.
[8] T. Lahotte, V. Caveliers, L. Dierickx, M. Vekeman, H. Everaert, J.
Mertens, A. Bossuyt, In vitro characterization of the influx of
3-[125I]iodo-L-␣-methlyl tyrosine and 2-[125I]iodo-L-tyrosine into
U266 human myeloma cells: Evidence for System T transport, Nucl.
Med. Biol. 28 (2001) 129 –134.
[9] H.N. Christensen, Role of amino acid transport and counter-transport
in nutrition and metabolism, Physiol. Rev. 70 (1990) 43–77.
[10] H. Segawa, Y. Fukasawa, K. Miyamoto, E. Takeda, H. Endou, Y.
Kanai, Identification and functional characterization of a Na⫹-independent neutral amino acid transporter with broad substrate selectivity, J. Biol. Chem. 274 (1999) 19745–19751.
[11] Y. Kanai, H. Segawa, K. Miyamoto, H. Uchino, E. Takeda, H. Endou,
Expression cloning and characterization of a transporter for large
neutral amino acids activated by heavy chain of 4F2 antigen (CD98),
J. Biol. Chem. 273 (1998) 23629 –23632.
[12] O. Yanagida, Y. Kanai, A. Chairoungdua, D.K. Kim, H. Segawa, T.
Nii, S.H. Cha, H. Matsuo, J. Fukushima, Y. Fukasawa, Y. Tani, Y.

37

Taketani, H. Uchino, J.Y. Kim, J. Inatomi, I. Okayasu, K. Miyamoto,
E. Takeda, T. Goya, H. Endou, Human L-type amino acid transporter
1 (LAT1): characterization of function and expression in tumor celllines, Biochm. Biophys. Acta. 1514 (2001) 291–302.
[13] C. Krummeich, M. Holschbach, G. Stocklin, Direct n.c.a. electrophilic radioiodination of tyrosine analogues: their in vivo stability andbrain-uptake in mice Appl. Radiat. Isot. 45 (1994) 929 –935.
[14] A. Chairoungdua, H. Segawa, J.Y. Kim, K. Miyamoto, J. Haga, Y.
Fukui, K. Mizoguchi, H. Ito, E. Takeda, H. Endou, Y. Kanai, Identification of an amino acid transporter associated with the cystinuriarelated type II membrane glycoprotein, J. Biol. Chem. 274 (1999)
28845–28848.
[15] P.D. Prasad, H. Wang, H. Huang, R. Kekuda, D.P. Rajan, F.H.
Leibach, V. Ganapathy, Human LAT1, a subunit of system L aminoacid transporter: molecular cloning and transport function, Biochem.
Biophys. Res. Commun. 255 (1999) 283–288.
[16] Y. Kanai, M.A. Hediger, Primary structure and functional characterization of a high-affinity glutamate transporter, Nature 360 (1992)
467– 471.
[17] Y. Kanai, S. Nussberger, M.F. Romero, W.F. Boron, S.C. Hebert,
M.A. Hediger, Electrogenic properties of the epithelial and neuronalhigh affinity glutamate transporter, J. Biol. Chem. 270 (1995) 16561–
16568.
[18] Y. Kanai, Y. Fukasawa, H.S. Cha, H. Segawa, A. Chairoungdua, D.K.
Kim, H. Matsuo, J.Y. Kim, K. Miyamoto, E. Takeda, H. Endou,Transport properties of a system y⫹L neutral and basic amino acidtransporter: Insights into the mechanisms of substrate recognition,
J. Biol. Chem. 275 (2000) 20787–20793.
[19] Y. Fukasawa, H. Segawa, J.Y. Kim, A. Chairoungdua, D.K. Kim, H.
Matsuo, S.H. Cha, H. Endou, Y. Kanai, Identification and characterization of a Na⫹-independent neutral amino acid transporter thatassociates with 4F2 heavy chain and exhibits substrate selectivity forsmall neutral D- and L-amino acids, J. Biol. Chem. 275 (2000)
9690 –9698.
[20] P.L. Jager, W. Vaalburg, J. Pruim, E. G.E. de Vries, K-J. Langen,
D.A. Piers, Radiolabeled amino acids: Basic aspects and clinicalapplications in oncology, J. Nucl. Med. 42 (2001) 432– 445.
[21] R. Deves, C.A.R. Boyd, Transporters for cationic amino acids inanimal cells: discovery, structure and function, Phys. Rev. 78 (1998)
487–545.
[22] K.-J. Langen, R. Bonnie, H. Muhlensiepen, P. Jansen, S. Broer, M.
Holschbach, H.H. Coenen, 3-[123I]iodo-␣-methyl-l-tyrosine transportand 4F2 antigen expression in human glioma cells, Nucl. Med. Biol.
28 (2001) 5–11.
[23] A. Hashimoto, T. Oka, Free D-aspartate and D-serine in the mammalian brain and periphery, Proc. Neuro. Boil. 52 (1997) 325–353.
[24] H.H. Tallan, S. Moore, W.H. Stein, Studies on the free amino acids
and related compounds in the tissues of the cat, J. Biol. Chem. 211(1954) 927–939.
[25] M. Palacin, R. Estevenz, E. Bertran, A. Zorano, Molecular biology of
mammalian plasma membrane amino acid transports, Physiol. Rev.
78 (1998) 969 –1054.
[26] H. Sato, M. Tamba, T. Ishii, S. Bannai, Cloning and expression of aplasma membrane cysteine/glutamate exchange transporter composedof two distinct proteins, J. Biol. Chem. 274 (1999) 11455–11458.
[27] H. Matsuo, S. Tsukada, T. Nakata, A. Chairoungdua, D.K. Kim, S.H.
Cha, J. Inatomi, H. Yorifuji, J. Fukuda, H. Endou, Y. Kanai, Expression of a system L neutral amino acid transporter at the blood-brain
barrier, Neuro. Report 11 (2000) 3507–3511.