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Inhibition of thymidylate synthase by the diastereoisomers of leucovorin

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Summary

The clinical formulation of leucovorin (5-CHO-FH4) is a mixture of diastereoisomers with markedly different pharmacologic properties. Comparatively little information is available concerning the cellular pharmacology of reduced folate stereoisomers, due largely to the difficulty in preparing sufficient quantities of these compounds for in vitro use. Recent improvements in HPLC technology have now facilitated this process, enabling studies of folate stereochemistry on a larger scale. Using purified (6R) and (6S) leucovorin, we examined the effects of these compounds on the enzymatic activity ofLactobacillus casei thymidylate synthase (TS) in a cell-free system. The natural (6S), unnatural (6R), and racemic (6R,S) leucovorin preparations inhibited TS activity by 50% at concentrations of 0.11, 2.1, and 0.52 mM, respectively. Dixon plots demonstrated the inhibition to be competitive, with Ki values of 85µM, 1.59 mM, and 385µM for (6S), (6R), and (6R,S) leucovorin, respectively. In view of the high doses of leucovorin given clinically and the slow clearance of the unnatural isomer, our observations suggest that leucovorin may have important direct inhibitory effects on folate-requiring enzymes.

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Abbreviations

FH4 :

l-tetrahydrofolic acid

FH2 :

7,8-dihydrofolic acid

CH2FH4 :

5,10-methylenetetrahydrofolic acid

5-CHO-FH4 :

leucovorin, 5-formyl-tetrahydrofolic acid, LV

10-CHO-FH4 :

10-formyltetrahydrofolic acid

5-CH3FH4 :

5-methyltetrahydrofolic acid

5-FdUMP:

5-fluorodeoxyuridine monophosphate

dTMP:

deoxythymidine monophosphate

TS:

thymidylate synthase

ΔA:

change in absorbance

dUMP:

deoxyuridine monophosphate

5-FU:

5-fluorouracil

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  1. Section of Hematology/Oncology, University of Chicago Pritzker School of Medicine, 60637, Chicago, IL, USA

    Pauline P. Lee & Richard L. Schilsky

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  1. Pauline P. Lee
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  2. Richard L. Schilsky
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Lee, P.P., Schilsky, R.L. Inhibition of thymidylate synthase by the diastereoisomers of leucovorin. Cancer Chemother Pharmacol 26, 273–277 (1990). https://doi.org/10.1007/BF02897229

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