Synthesis and antitumor activity of a novel series of 6-substituted pyrrolo[2,3-d]pyrimidines as potential nonclassical antifolates targeting both thymidylate and purine nucleotide biosynthesis

摘要

A novel series of 2-amino-4-oxo-6-substituted pyrrolo[2,3-d]pyrimidines were designed and synthesized as potential nonclassical antifolates targeting both thymidylate and purine nucleotide biosynthesis. Condensation of 2,4-diamino-6-hydroxypyrimidine with ethyl-4-chloroacetoacetate and subsequent hydrolysis afforded the key intermediate, 2-amino-4-oxo-pyrrolo[2,3-d]pyrimidin-6-yl-acetic acid. Coupling with various amino acid methyl esters followed by saponification and condensation with 3(aminomethyl)pyridine provided target compounds 1-9. The new compounds exhibited micromolar to submicromolar antiproliferative potencies against a panel of tumor cell lines including KB, A549 and HepG2. Growth inhibition of compound 2 toward KB cells resulted in cytotoxicity and G1/G2-phase accumulation, and was partially protected by excess thymidine and adenosine, but was completely reversed in the combination of thymidine and adenosine, indicating both thymidylate and de novo purine nucleotide synthesis as the targeted pathway. However, 5-aminoimidazole-4-carboxamide (AICA) protection was incomplete, suggesting inhibition of both glycinamide ribonucleotide formyltransferase (GARFTase) and AICA ribonucleotide formyltransferase (AICARFTase). The results of the docking studies show that 2 could bind and inhibit both thymidylate synthase (TS) and the two folate-dependent purine biosynthetic enzymes (GARFTase and AICARFFase), which is consistent with the results of in vitro metabolic assays. Our studies establish that compound 2 is an excellent lead analog as a multitargeted antifolate for further structure optimization. (C) 2015 Elsevier Masson SAS. All rights reserved.