title: Nonclassical 2,4-diamino-5-aryl-6-ethylpyrimidine antifolates: activity as inhibitors of dihydrofolate reductase from Pneumocystis carinii and Toxoplasma gondii and as antitumor agents.
abstract: Twelve novel 2,4-diamino-5-(4'-benzylamino)- and 2,4-diamino-5[4'-(N-methylbenzylamino)-3'-nitrophenyl]-6-ethylp yrimidines bearing 4-substituents on the benzylamino or N-methylbenzylamino aryl ring were synthesized and evaluated as nonclassical inhibitors of Pneumocystis carinii and Toxoplasma gondii dihydrofolate reductase (DHFR). Compounds were prepared by reaction of 2,4-diamino-5-(4'-chloro-3'-nitrophenyl)- (8) or 2,4-diamino-5-(4'-fluoro-3'-nitrophenyl)-6-ethylpyrimidine (15) with the appropriate 4-substituted (CO2H, CO2Me, SO2NH2, dioxolan-2-yl, CHO, dimethyloxazolin-2-yl) benzylamine or N-methylbenzylamine derivative. Compounds 25-29 were synthesized from 2,4-diamino-5-{4'-[N-(4"-carboxybenzyl)amino]-3'-nitrophenyl}-6- ethylpyrimidine (10) and the corresponding amine (NH3, MeNH2, Me2NH, piperidine, diethyl L-glutamate) via isobutyl mixed anhydride coupling; hydrolysis of the diethyl L-glutamate 29 afforded the L-glutamate analogue 30. The compounds exhibited potent inhibitory activity against T. gondii (IC50 values 0.0018-0.14 microM) and rat liver (IC50 values 0.0029-0.27 microM) DHFR, with a 4-substituent invariably enhancing binding to both enzymes relative to the unsubstituted benzoprim (5) or methylbenzoprim (6). Modest selectivity for T. gondii enzyme was observed with several analogues, whereas all of the compounds were relatively weak inhibitors of P. carinii DHFR and exhibited no selectivity. Selected analogues were evaluated for in vivo antitumor activity against the methotrexate-resistant M5076 murine reticulosarcoma, with 2,4-diamino-5-{4'-[N-[4"-(N"-methylcarbamoyl)benzyl]-N- methylamino]-3'-nitrophenyl}-6-ethylpyrimidine (14) (Ki for rat liver DHFR = 0.00035 +/- 0.00029 nM) combining significant antitumor activity with minimal toxicity.
authors.affiliation: Department of Chemistry, University, Newcastle upon Tyne, U.K.
authors.forename: C
authors.initials: C
authors.lastname: Robson
authors.forename: M A
authors.initials: MA
authors.lastname: Meek
authors.forename: J D
authors.initials: JD
authors.lastname: Grunwaldt
authors.forename: P A
authors.initials: PA
authors.lastname: Lambert
authors.forename: S F
authors.initials: SF
authors.lastname: Queener
authors.forename: D
authors.initials: D
authors.lastname: Schmidt
authors.forename: R J
authors.initials: RJ
authors.lastname: Griffin
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D005493:Folic Acid Antagonists / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D007202:Indicators and Reagents; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D016403:Lymphoma, Large B-Cell, Diffuse / Q000188:drug therapy*; D051379:Mice; D011010:Pneumocystis / Q000201:enzymology*; D011743:Pyrimidines / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D051381:Rats; D013329:Structure-Activity Relationship; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism*; D014122:Toxoplasma / Q000201:enzymology*
chemical_list: D000970:Antineoplastic Agents; D005493:Folic Acid Antagonists; D007202:Indicators and Reagents; D011743:Pyrimidines; D013762:Tetrahydrofolate Dehydrogenase