( 18 F) ( 40: 23 30, 2003) 3 1 ( ) 3) 18 F-FDG 1,2) 1,2) 5,6) 6,7) 14 C-FDG 18 F-FDG F-FDM-6-

Transcription

1 23 18 F-FDG D- ( 18 F) * ** * * * 18 F-FDG 14 C 14 C-FDG B 75% FDG 18 F-FDG 18 F-FDG-6-18 F-FDG F-FDM-6- PET FDG 18 F-FDG FDG ( 40: 23 30, 2003) I [ 18 F] D- ( 18 F-FDG ) ( ) 1,2) * ** ( ) 18 F-FDG 3) 18 F-FDG 4) 70 1,2) 5,6) 6,7)

2 (2003) 8,9) FDG FDG-6-10,11) in vitro 18 F-FDG 18 F-FDG-6-12) 18 F-FDG FDG II. 1. SD (1,500 g, 4 C 10 ) (PBS Dulbecco s Formula) 50 ml 2 5 ml ( MEK- 5158) /ml 2. ( ) 0.5 ml 0 1 mol/l 25 µl ( 0 50 mmol/ l) ( BF-200) 37 C C- (Du Pont 1 mmol/l) 14 C- FDG (American Radiolabeled Chemicals FDG 1 mmol/l) 37 C ml (0.1 mmol/l 1% (2,000 g 5 ) 0.5 mmol/l 0.75 ml ( 0.15 ml BECKMAN Ready Cap TM (Beckman ) (Beckman LS6000SC) 3. ( ) 0.5 ml 0 20 mmol/l B (Aldrich Chemical ) 25 µl ( B 0 1 mmol/l) 37 C C- ( 1 mmol/l) 14 C-FDG (FDG 1 mmol/l) 37 C Table 1 37 C 30 1 mol/l 0.5 ml (2,000 g 6 )

3 18 F-FDG 25 Table 1 Reaction mixture for measurement of hexokinase activity 18 F-FDG injectable : 50 µl Hexokinase : 4 U 40 mmol/l ATP* : 200 µl 170 mmol/l MgCl 2 : 100 µl 50 mmol/l Tris HCl (ph 8.0) : 100 µl PBS: Dulbecco s formula : Adequate Total volume : 1 ml Hexokinase (Boehringer Mannheim GmbH, Germany) *ATP: Adenosine 5 -triphosphate (Boehringer Mannheim GmbH, Germany) Table 4 Rf value of metabolites Rf value Metabolites Before AP* After AP* treatment treatment 18 F-FDG F-FDM F-FDG-6-phosphate F-FDM-6-phosphate *AP: Alkaline phosphatase Table 2 Reaction mixture for measurement of PGI* activity Reaction mixture of Table 1 : 100 µl PGI : 14 U or 70 U 50 mmol/l Tris HCl (ph 8.0) : 50 µl PBS: Dulbecco s formula : Adequate Total volume : 1 ml * PGI: Phosphoglucose isomerase (Boehringer Mannheim GmbH, Germany) Table 3 Reaction mixture for dephosphorylation Reaction mixture of Table 2 : 100 µl Alkaline phosphatase : 20 U 50 mmol/l Tris HCl (ph 8.0) : 50 µl PBS: Dulbecco s formula : Adequate Total volume : 1 ml Alkaline phosphatase (Boehringer Mannheim GmbH, Germany) ( 5 1 1) 5. Table 2 37 C Table 3 37 C 30 Fig. 1 Uptake of 14 C-glucose ( ) or 14 C-FDG ( ) into erythrocytes. The plots are shown as percent of radioactivity in erythrocytes to untreated with glucose. (mean, n 4) 1 mol/l 0.5 ml (2,000 g 6 ) ( 5 1 1) 6. Rijn 13) 60 (2 20 cm 10% 2 )

4 (2003) Fig. 2 Uptake of 14 C-glucose ( ) or 14 C-FDG ( ) into erythrocytes. The plots are shown as percent of radioactivity in erythrocytes to untreated with cytochalasin B. (mean, n 4) Fig. 4 Thin-layer chromatogram of reaction mixture (Table 3) after dephosphorylation with alkaline phosphatase. Isomerization was reacted with 70 U PGI* for 90 min. *PGI: Phosphoglucose isomerase BAS2000) Rf Table 4 III. 1. ( ) 14 C- 14 C-FDG Fig. 3 Thin-layer chromatogram of reaction mixture (Table 1) after phosphorylation. (95 5) 5 cm 6.5 cm 8 cm, 9.5 cm, 11 cm, 12.5 cm 15.5 cm 7 ( 14 C- 50 mmol/l 6% (Fig. 1) 14 C-FDG 50 mmol/l 11% (Fig. 1) 2. ( ) 14 C- 14 C-FDG B 14 C- 0.1 mmol/l B (Fig. 2) 14 C-FDG B 26% (Fig. 2)

5 18 F-FDG F-FDG 100% 18 F-FDG- (Fig. 3) F-FDG FDG D- ( 18 F-FDM ) (Fig. 4) 14 U 30 4% 70 U 90 27% 18 F-FDM (Table 5) Table 5 Isomerization of 18 F-FDG-6-phosphate to 18 F-FDM-6-phosphate by PGI* Amount of PGI Reaction times 18 F-FDG-6-phosphate 18 F-FDM-6-phosphate 14 U 30 min 96.3% 3.7% 14 U 90 min 92.2% 7.9% 70 U 30 min 85.1% 14.9% 70 U 90 min 73.2% 26.8% Note: The amount of 18 F-FDG-6-phosphate and 18 F-FDM-6-phosphate were estimated after dephosphorylation with alkaline phosphatase by Thin-layer chromatography. *PGI: Phosphoglucose isomerase Fig. 5 Scheme for the accumulation of FDG in the tumor cells.

6 (2003) IV. 14 C-FDG 14 C-FDG 14 C-FDG B 75% 14 C-FDG FDG B 25% 18 F-FDG 100% 18 F-FDG 18 F-FDG 18 F-FDG 18 F-FDG 2 14) 18 F-FDG 18 F-FDG F-FDG 15) (Fig. 5) 19 F-NMR 19 F-FDG FDG FDM ) 16) 14 C-FDG 14 C-FDM 17) 18 F in vitro 18 F-FDG 18 F- FDM 18 F-FDG 18 F-FDG 18 F-FDM 18 F- FDG 18 F- 18 F-FDM 18 F-FDM 18 F-FDM 18 F-FDG 18) V. 14 C 14 C-FDG FDG 18 F-FDG 18 F-FDG 18 F-FDG 18 F-FDM 18 F-FDG PET FDG 18 F-FDG FDG

7 18 F-FDG 29 1) Warburg O: On the origin of cancer cells. Science 1956; 123: ) Warburg O: The Metabolism of Tumors. Constable, London, 1930: ) Som P, Atkins HL, Bandoypadyay D, Fowler JS, MacGregor RR, Matsui K, et al: A fluorinated glucose analog, 2-fluoro-2-deoxy-D-glucose (F-18): Nontoxic tracer for rapid tumor detection. J Nucl Med 1980; 21: ) : 18 F-FDG D- ( 18 F) 1999; 36: ) Yamamoto T, Seino Y, Fukumoto H, Koh G, Yano H, Inagaki N, et al: Over-expression of facilitative glucose transporter genes in human cancer. Biochem Biophys Res Commun 1990; 170: ) Younes M, Lechago LV, Somoano JR, Mosharaf M, Lechago J: Wide Expression of the human erythrocyte glucose transporter Glut 1 in human cancers. Cancer Res 1996; 56: ) Brown RS, Leung JY, Fisher SJ, Frey KA, Ethier SP, Wahl RL: Intratumoral distribution of tritiated-fdg in breast carcinoma: Correlation between Glut-1 expression and FDG uptake. J Nucl Med 1996; 37: ) Weber G: Enzymology of cancer cells. (Second of two parts) New Eng J Med 1977; 296: ) Monakov NK, Neistadt EL, Shavlovskii MM, Shvartsman AL, Neifakh SA: Physicochemical properties and isoenzyme composition of hexokinase from normal and malignant human tissues. J Natl Cancer Inst 1978; 61: ) Pouremad R, Wyrwicz AM: Cerebral metabolism of fluorodeoxyglucose measured with 19 F NMR spectroscopy. NMR in Biomed 1991; 4: ) Shinohara S, Kanazawa Y, Kojima M: Evaluation of energy metabolism in brain using epimerization of 2- deoxy-2-fluoro-d-glucose by 19 F NMR: The effect of anesthesia. Magn Reson Med 1991; 21: ) Gallagher BM, Fowler JS, Gutterson NI, MacGregor RR, Wan CN, Wolf AP: Metabolic trapping as a principle of radiopharmaceutical design: Some factors responsible for the biodistribution of [ 18 F] 2-deoxy-2- fluoro-d-glucose. J Nucl Med 1978; 19: ) Rijn CJSV, Herscheid JDM, Visser GWM, Hoekstra A: On the streoselectivity of the reaction of [ 18 F] acetylhypofluororite with glucals. Int J Appl Radiat Isot 1985; 36: ) Bessell EM, Thomas P: The deoxyfluoro-d-glucose 6- phosphate and their effect on yeast glucose phosphate isomerase. Biochem J 1973; 131: ) Nelson CA, Wang JQ, Leav I, Crane PD: The interaction among glucose transport, hexokinase, and glucose-6-phosphatase with respect to 3 H-2-deoxyglucose retention in murine tumor models. Nucl Med Biol 1996; 23: ) Kanazawa Y, Yamane H, Shinohara S, Kuribayashi S, Momozono Y, Yamato Y, et al: 2-deoxy-2-fluoro-Dglucose as a functional probe for NMR: the unique metabolism beyond its 6-phosphate. J Neurochem 1996; 66: ) Schmidt MFG, Biely P, Kratky Z, Schwarz RT: Metabolism of 2-deoxy-2-fluoro-D-[ 3 H]glucose and 2- deoxy-2-fluoro-d-[ 3 H]mannose in yeast and chickembryo cells. Eur J Biochem 1978; 87: ) : PET 1985; 34:

8 30 Summary Uptake of FDG (2-fluoro-2-deoxy-D-glucose) as a Tumor Imaging Agent into Erythrocytes and Accumulation of FDG in Tumor Cells Yoshihito MINOSAKO*, Masahiro NEMOTO**, Sento INO*, Yoshifumi SHIRAKAMI* and Miki KURAMI* *Research Centre, Research & Development Division, Nihon Medi-physics Co., Ltd. **Department of Basic Sciences, Japanese Red Cross Hokkaido College of Nursing Fluorine-18-2-fluoro-2-deoxy-D-glucose ( 18 F-FDG) injectable was developed as a tumor imaging agent reflecting glucose metabolism. In membrane transportation studies, the uptake of 14 C-FDG into erythrocytes decreased with an increase in glucose concentration, and Cytochalasin B, inhibitor of glucose transporter (GLUT), blocked the uptake about 75%. The results means FDG is transported into tumor cells mainly by GLUT as glucose analogues. 18 F- FDG is recognized to be phosphorylated to 18 F-FDG- 6-phosphate with hexokinase. We found that FDG- 6-phosphate was further isomerized to 18 F-FDM-6- phosphate by phosphoglucose isomerase (PGI) in vitro. About 27% 18 F-FDM-6-phosphate was generated at the reaction with 70 U PGI for 90 min. These results show that the 18 F-FDG injectable manufactured by the commercial supply system has equivalent properties; membrane transportation characteristic and enzyme affinity, to FDG synthesized at each PET institution. Key words: 18 F-FDG, PET, GLUT, Hexokinase, Metabolism.