Synthesis of quaternised 2-aminopyrimido[4,5-d]pyrimidin-4(3H)-ones and their biological activity with dihydrofolate reductase

Markus G. Gebauer, Carolyn McKinlay, Jill E. Gready*

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    78 Citations (Scopus)

    Abstract

    In a program to design and develop mechanism-based compounds active as substrates and inhibitors of dihydrofolate reductase (DHFR), we report the synthesis and physical properties of the 6-methyl- (7), 8-methyl- (8a), and 8-ethyl- (8b) derivatives of the parent 2-aminopyrimido[4,5-d]pyrimidin-4-(3H)-one (6). These compounds are the first members of a class of heterocycles related to 8-alkylpterins (N8-alkyl-2-aminopteridin-4(8H)-ones) (2a-2c), which have been shown to be novel substrates for DHFR. Three methods were developed for the synthesis of target compounds 7, 8a and 8b; however, the optimum yields (1-8%) could not be improved because the products decomposed by ring opening (e.g. to 2,4-diamino-5-methyliminomethylpyrimidin-6(1H)-one (9)) under the reaction conditions. The marked π-electron deficiency of compounds 7, 8a and 8b is the likely cause for the susceptibility of the quaternised pyrimidine ring in the related cations 10, 15a and 15b, respectively, to add nucleophiles, thus promoting the opening of the pyrimidopyrimidine ring system. 1H-NMR spectroscopic studies of compounds 7, 8a and 8b revealed a fast and reversible covalent hydration of the associated cations across the C7-N8 bond for the N6-methyl derivative 7 and across the N6-C7 bond for the N8-methyl derivative 8a. UV spectroscopic studies of methyl derivatives 7 and 8a as well as the parent heterocycle 6 showed that protonation of the latter occurred at N1, while methylation with iodomethane proceeded at N6 and N8. The basicities of the N-methyl derivatives 7 and 8a (pKa ca. 5.5) are similar to those of 8-alkylpterins 2; this is an essential element of the design to promote binding to DHFR in their protonated form. Enzyme kinetics of 7, 8a and 8b with chicken DHFR confirmed our predictions that they are substrates, with apparent Km values of 3.8, 0.08, and 0.65 mM, and apparent Vmax values of 0.47, 2.27, and 0.30 nmol L-1 min-1 (for enzyme concentration 0.122 μM), respectively. The parent compound 6 was not a substrate.

    Original languageEnglish
    Pages (from-to)719-728
    Number of pages10
    JournalEuropean Journal of Medicinal Chemistry
    Volume38
    Issue number7-8
    DOIs
    Publication statusPublished - 2003

    Fingerprint

    Dive into the research topics of 'Synthesis of quaternised 2-aminopyrimido[4,5-d]pyrimidin-4(3H)-ones and their biological activity with dihydrofolate reductase'. Together they form a unique fingerprint.

    Cite this