Synthesis of classical and a nonclassical 2-amino-4-oxo-6-methyl-5-substituted pyrrolo[2,3-d]pyrimidine antifolate inhibitors of thymidylate synthase

Compounds 2-5 were designed as potential antifolate nonpolyglutamatable inh ibitors of thymidylate synthase (TS). These analogues are structurally rela ted to 2-amino-4-oxo-5-substituted quinazolines and 2-amino-4-oxo-5-substit uted pyrrolo[2,3-d]pyrimidines which have shown excellent inhibition of TS and, for the quinazoline, significant promise as clinically useful antitumo r agents. Compounds 2-4 were synthesized by appropriate amine exchange reac tions on pivaloyl-protected 5-dimethylaminomethyl-substituted 6-methyl pyrr olo[2,3-d]pyrimidine 7 which in turn was obtained from the Mannich reaction of pivaloylated-6-methyl pyrrolo[2,3-d]pyrimidine 6. In instances where th e amine exchange reaction was sluggish, the Mannich base was quaternized wi th methyl iodide which afforded much faster exchange reaction with improved yields. For compound 5, 4-mercaptopyridine was used as the nucleophile and reacted with 7. The analogues 2-4 inhibited Lactobacillus casei (1c) TS an d recombinant human (h) TS with IC50 in the 10(-4) to. 10(-5) M range. Comp ound 5 inhibited 1cTS and hTS 20% at 26 and 25 mu M, respectively. In addit ion, compound 5 inhibited the growth of Pneumocystis carinii and Toxoplasma gondii cells in culture by 76% at 32 x 10(-6) M and 50% at 831 x 10(-6) M, respectively.