Synthesis and in Vitro Biological Activity of New Deaza Analogues of Folic Acid, Aminopterin, and Methotrexate with an L-Ornithine Side Chain

Article abstract of DOI:10.1021/jm00108a032

The 5-deaza and 5,8-dideaza analogues of N^α-pteroyl-L-ornithine (Pter-Orn), the 5-deaza, 8-deaza, and 5,8-dideaza analogues of N^α-(4-amino-4-deoxypteroyl)-L-ornithine (APA-Orn), and the N^δ-carboxymethyl derivative of N^α-(4-amino-4-deoxy-N¹⁰-methylpteroyl)-L-ornithine (m-APA-Orn) were synthesized and tested as inhibitors of dihydrofolate reductase (DHFR) and as inhibitors of tumor cell growth in culture.Reductive amination of 2-acetamido-6-formylpyrido<2,3-d>pyrimidin-4(3H)-one with methyl N^α-(4-aminobenzoyl)-N^δ-(benzyloxycarbonyl)-L-ornithinatefollowed by removal of the blocking groups afforded the 5-deaza analogue of Pter-Orn, whereas N-alkylation of methyl N^α-(4-aminobenzoyl)-N^δ-(benzyloxycarbonyl)-L-ornithinate with 2-amino-6-(bromomethyl)quinazolin-4(3H)-one and deprotection gave the corresponding 5,8-dideaza analogue.Reductive coupling of 2,4-diaminopyrido<2,3-d>pyrimidine-6-carbonitrile and 4-aminobenzoic acid followed by reaction with 96-97percent formic acid yielded 4-amino-4-deoxy-5-deaza-N¹⁰-formylpteroic acid, which on condensation with methyl N^δ-(benzyloxycarbonyl)-L-ornithinate and deprotection gave the 5-deaza analogue of APA-Orn.A similar sequence starting from 2,4-diaminoquinazoline-6-carbonitrile led to the corresponding 5,8-dideaza compound, whereas treatment of 2,4-diaminopyrido<3,2-d>pyrimidine-6-methanol with phosphorus tribromide followed by condensation with methyl N^α-(4-aminobenzoyl)-N^δ-(benzyloxycarbonyl)-L-ornithinate and deprotection afforded the 8-deaza analogue.For the preparation of the N^δ-carboxymethyl derivative of mAPA-Orn, N^α-(benzyloxycarbonyl)-L-ornithine was subjected to N^δ-monoalkylation with glyoxylic acid and sodium cyanoborohydride, followed by N^δ-acylation with ethyl trifluoroacetate, N^α-deprotection by hydrogenolysis, condensation with 4-amino-4-deoxy-N¹⁰-methylpteroic acid, and N^δ-deprotection by gentle treatment with ammonia.The 2,4-diamino derivatives all inhibited the growth of tumor cells in culture, with IC50 values of 0.2-2 μM, and inhibited purified DHFR with IC50 values of 0.02-0.08 μM.Deletion of ring nitrogens and N^δ-carboxymethylation both increased potency in the cell growth assay; however, the ornithine derivatives were less potent than aminopterin or methotrexate.