Stimulation of 125I-3-iodo-alpha-methyl-L-tyrosine uptake in Chinese hamster ovary (CHO-K1) cells by tyrosine esters.
BACKGROUND
Transport of the amino acid analog (123)I-3-iodo-alpha-methyl-L-tyrosine, which is used in clinical SPECT imaging, occurs mainly via L-type amino acid transporter type 1 (LAT1; an amino acid exchanger). As LAT1 is highly expressed in actively proliferating tumors, we made a preliminary investigation of the effects of amino acid esters on enhancement of (125)I-3-iodo-alpha-methyl-L-tyrosine (IMT) uptake via LAT1 in Chinese hamster ovary (CHO-K1) cells.


METHODS
Because the sequence of the CHO-K1 LAT1 gene is not available, we confirmed LAT1 expression through IMT (18.5 kBq) uptake mechanisms using specific inhibitors. L-Gly, L-Ser, L-Leu, L-Phe, L-Met, L-Tyr, D-Tyr, L-Val and L-Lys ethyl/methyl esters were tested in combination with IMT. Time-course studies over a 3-h period were conducted, and the concentration dependence of L-Tyr ethyl and methyl esters (0.001 to 10 mM) in combination with IMT was also examined. For a proof of de-esterification of L- and D-Tyr ethyl and methyl esters in the cells (by enzymatic attack or other cause), the concentration of L- and D-Tyr was analyzed by high-performance liquid chromatography of the esters in phosphate buffer (pH 7.4) and cell homogenates at 37 degrees C or under ice-cold conditions.


RESULTS
Inhibition tests suggested that LAT1 is involved in IMT uptake by CHO-K1 cells. Co-administration of 1 mM of l-Tyr ethyl or methyl ester with IMT produced the greatest enhancement. The de-esterification reaction was stereo selective and temperature dependent in the homogenate. De-esterification kinetics were very fast in the homogenate and very slow in the phosphate buffer.


CONCLUSIONS
The L-Tyr ethyl or methyl esters were the most effective enhancers of IMT uptake into CHO-K1 cells and acted by trans-stimulation of the amino acid exchange function of LAT1. This result suggests that de-esterification in the cells may be caused by enzymatic attack. We will use IMT and L-Tyr ethyl or methyl esters to examine LAT1 function in tumor cells or tissues in vivo.
Department of Radiological Sciences, Center for Medical Sciences and Center for Humanities and Sciences, Ibaraki Prefectural University of Health Sciences, Inashiki-gun, Ibaraki, Japan. sikano@ipu.ac.jp
Naoto
N
Shikano
Masato
M
Ogura
Jun-ichi
J
Sagara
Syuichi
S
Nakajima
Masato
M
Kobayashi
Takeshi
T
Baba
Naoto
N
Yamaguchi
Yukio
Y
Iwamura
Nobuo
N
Kubota
Keiichi
K
Kawai
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D016466:CHO Cells; D002457:Cell Extracts; D006224:Cricetinae; D003412:Cricetulus; D004951:Esterification; D004952:Esters / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology*; D006868:Hydrolysis; D027283:Large Neutral Amino Acid-Transporter 1 / Q000378:metabolism; D008781:Methyltyrosines / Q000378:metabolism*
D002457:Cell Extracts; D004952:Esters; D027283:Large Neutral Amino Acid-Transporter 1; D008781:Methyltyrosines; C030855:3-iodo-alpha-methyltyrosine