Discovery of 5-substituted pyrrolo[2,3-d]pyrimidine antifolates as dual acting inhibitors of glycinamide ribonucleotide formyltransferase and 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase in de novo purine nucleotide biosynthesis: implications of inhibiting 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase to AMPK activation and anti-tumor activity

• Published in: Journal of medicinal chemistry Vol. 56; no. 24; pp. 10016 - 10032
• Main Authors: Mitchell-Ryan, Shermaine, Wang, Yiqiang, Raghavan, Sudhir, Ravindra, Manasa Punaha, Hales, Eric, Orr, Steven, Cherian, Christina, Hou, Zhanjun, Matherly, Larry H., Gangjee, Aleem
• Format: Journal Article
• Language: English
• Published: 11.12.2013
• ISSN: 0022-2623; 1520-4804
• DOI: 10.1021/jm401328u

We synthesized 5-substituted pyrrolo[2,3- d ]pyrimidine antifolates (compounds 5 – 10 ) with 1 to 6 bridge carbons and a benozyl ring in the side chain as antitumor agents. Compound 8 with a 4-carbon bridge was the most active analog and potently inhibited proliferation of folate receptor (FR) α-expressing Chinese hamster ovary and KB human tumor cells. Growth inhibition was reversed completely or in part by excess folic acid, indicating that FRα is involved in cellular uptake, and resulted in S-phase accumulation and apoptosis. Anti-proliferative effects of compound 8 toward KB cells were protected by excess adenosine but not thymidine, establishing de novo purine nucleotide biosynthesis as the targeted pathway. However, 5-aminoimidazole-4-carboxamide (AICA) protection was incomplete, suggesting inhibition of both AICA ribonucleotide formyltransferase (AICARFTase) and glycinamide ribonucleotide formyltransferase (GARFTase). Inhibition of GARFTase and AICARFTase by compound 8 was confirmed by cellular metabolic assays and resulted in ATP pool depletion. To our knowledge, this is the first example of an antifolate that acts as a dual inhibitor of GARFTase and AICARFTase as its principal mechanism of action.