Fluorination at the 4 position alters the substrate behavior of l-glutamine and l-glutamate: Implications for positron emission tomography of neoplasias

Metabolism of the 2S,4S- and 2S,4R diastereoisomers of 4-FGln and 4-FGln. The solid arrows indicate the proposed metabolic pathways for the 2S,4S- and 2S,4R diastereoisomers of 4-FGln and 4F-Glu, while the broken arrows indicate the less favored pathways. [Display omitted] •Activity with glutamine transaminase K: Gln≈(2S,4S)-4-FGln>>(2S,4R)-4-FGln.•Activity with glutamine transaminase l: Gln>>(2S,4R)-4-FGln≈(2S,4S)-4-FGln.•Activity with glutaminase (short form): Glu>(2S,4S)-4-FGln>>(2S,4R)-4-FGln.•Activity with glutamine synthetase: Glu>(2S,4S)-4-FGlu≈(2S,4R)-4-FGlu.•(2S,4S)-4-F-(3-Fluoropropyl)glutamine is likely largely metabolically inert in vivo. Two 4-fluoro-l-glutamine diastereoisomers [(2S,4R)-4-FGln, (2S,4S)-4-FGln] were previously developed for positron emission tomography. Label uptake into two tumor cell types was greater with [18F](2S,4R)-4-FGln than with [18F](2S,4S)-4-FGln. In the present work we investigated the enzymology of two diastereoisomers of 4-FGln, two diastereoisomers of 4-fluoroglutamate (4-FGlu) (potential metabolites of the 4-FGln diastereoisomers) and another fluoro-derivative of l-glutamine [(2S,4S)-4-(3-fluoropropyl)glutamine (FP-Gln)]. The two 4-FGlu diastereoisomers were found to be moderate-to-good substrates relative to l-glutamate of glutamate dehydrogenase, aspartate aminotransferase and alanine aminotransferase. Additionally, alanine aminotransferase was shown to catalyze an unusual γ-elimination reaction with both 4-FGlu diastereoisomers. Both 4-FGlu diastereoisomers were shown to be poor substrates, but strong inhibitors of glutamine synthetase. Both 4-FGln diastereoisomers were shown to be poor substrates compared to l-glutamine of glutamine transaminase L and α-aminoadipate aminotransferase. However, (2S,4R)-4-FGln was found to be a poor substrate of glutamine transaminase K, whereas (2S,4S)-4-FGln was shown to be an excellent substrate. By contrast, FP-Gln was found to be a poor substrate of all enzymes examined. Evidently, substitution of H in position 4 by F in l-glutamine/l-glutamate has moderate-to-profound effects on enzyme-catalyzed reactions. The present results: 1) show that 4-FGln and 4-FGlu diastereoisomers may be useful for studying active site topology of glutamate- and glutamine-utilizing enzymes; 2) provide a framework for understanding possible metabolic transformations in tumors of 18F-labeled (2S,4R)-4-FGln, (2S,4S)-4-FGln, (2S,4R)-4-FGlu or (2S,4S)-4-FGlu; and 3) show that [18F]FP-Gln is likely t