Inhibition of Bacterial Dihydrofolate Reductase by 6‐Alkyl‐2,4‐diaminopyrimidines

Abstract

A series of (±)-6-alkyl-2,4-diaminopyrimidine-based inhibitors of bacterial dihydrofolate reductase (DHFR) have been prepared and evaluated for biological potency against Bacillus anthracis and Staphylococcus aureus. Biological studies reveal attenuated activity relative to earlier structures lacking substitution at C6 of the diaminopyrimidine moiety, though minimum inhibitory concentration (MIC) values are in the 0.125–8 μg/mL range for both organisms. This effect was rationalized from previous three-dimensional X-ray structure studies that indicate the presence of a side pocket containing two water molecules adjacent to the main binding pocket. Because of the hydrophobic nature of the substitutions at C6 the main interactions are with protein residues Leu20 and Leu28. These interactions lead to a minor conformational change in the protein, which opens the pocket containing these waters such that it is continuous with the main binding pocket. These water molecules are reported to play a critical role in the catalytic reaction. This highlights a new area for inhibitor expansion within the limited architectural variation at the catalytic site of bacterial DHFR.

Modified 2,4-diaminopyrimidine-based dihydrofolate reductase inhibitors as potential drug scaffolds against Bacillus anthracis

“…This was followed by a palladium(II) acetate-mediated Heck reaction of the resulting ketone 2 with (±)-1-(1-alkyl-2(1 H )-phthalazinyl)-2-propen-1-ones 3a and 3b in DMF at 140 °C to give the target molecules 4a (79%) and 4b (75%), respectively. 7, 8, 18, 19 …”

“…7, 8 Briefly, the minimum inhibitory concentration (MIC) was the lowest concentration of compound needed to block growth of a standardized culture of B. anthracis Sterne as measured by turbidity at 600 nm and by visual inspection. 27 Enzymatic activity was reconstituted in vitro with saturating concentrations of co-factor and dihydrofolate reductase.…”

“…7, 10 The second alterations were carried out at R 4 on the DAP ring, but increased activity was not observed. 8 In a further quest for improved potency over RAB1 and BN-53, the current study investigated analogs resulting from oxidation of the methylene bridge linking the DAP fragment to the central ring, as well as changes at R 2 and R 3 , and assessed the biological activities of the resulting compounds.…”

Evaluation of New Dihydrophthalazine-Appended 2,4-Diaminopyrimidines against Bacillus anthracis: Improved Syntheses Using a New Pincer Complex

“…It was observed earlier that any alteration of R 1 , at the C-1 stereocenter of the dihydrophthalazine unit, tended to modulate interaction of the protein surface with the surrounding solvent, and thus, the inhibitory activity [ 13 , 15 ]. Alternatively, changes at the R 2 , R 3 and R 4 positions on the ring disrupted the compound orientation in the binding pocket, which resulted in attenuated potency [ 14 , 16 ]. Numerous compounds with changes at the R 1 position were synthesized and evaluated, but derivatives with sensitive functional groups at this position proved challenging to prepare.…”

4,5-Dimethylbenzene-1,2-dimethanol

“…The rigid planarity of the ring system allows close positioning of reactive centers such that additional fused rings can be readily appended. We recently needed multi-gram quantities of the title compound as a building block to prepare 6,7-dimethylphthalazine for use in the synthesis of dihydrophthalazine-based antibiotics [1][2][3][4][5][6]. Though the compound is commercially available, it is quite expensive, and the supplier was unable to meet our requirements.…”