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.
Shikano
Naoto
N
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
Ogura
Masato
M
Sagara
Jun-ichi
J
Nakajima
Syuichi
S
Kobayashi
Masato
M
Baba
Takeshi
T
Yamaguchi
Naoto
N
Iwamura
Yukio
Y
Kubota
Nobuo
N
Kawai
Keiichi
K
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