3‑Hydroxy-Beta-Lactam Inhibitors of Dihydrofolate Synthetase.

Area of Science:

• Medicinal Chemistry
• Enzyme Inhibition
• Drug Discovery

Background:

• The 3-hydroxy-β-lactam (3-HβL) moiety is derived from tabtoxinine-β-lactam (TβL), a known glutamine synthetase inhibitor.
• Dihydrofolate synthetase (DHFS) is a critical enzyme in folate metabolism, making it a target for antifolate drugs.

Purpose of the Study:

• To repurpose the 3-HβL group as a pharmacophore for developing novel dihydrofolate synthetase (DHFS) inhibitors.
• To investigate the potential of 3-HβL as a general pharmacophore for ATP-dependent carboxylate-amine ligase superfamily inhibitors.

Main Methods:

• In vitro steady-state kinetics and enzyme-coupled assays were employed to assess DHFS inhibition.
• Molecular modeling was utilized to understand the interaction of the 3-HβL group with DHFS.
• Optimized synthesis of key intermediates, including 3-(p-aminophenyl)-3-HβL, was performed using Henry and Grignard reactions.

Main Results:

• Replacement of the carboxyl group of p-aminobenzoic acid (PABA) with a 3-HβL moiety yielded potent DHFS inhibitors.
• The 3-HβL group was validated as essential for DHFS inhibition.
• Successful synthesis and conjugation of the PABA analogue to a pterin mimic created a functional antifolate scaffold.

Conclusions:

• The 3-hydroxy-β-lactam (3-HβL) group serves as a potent pharmacophore for dihydrofolate synthetase (DHFS) inhibition.
• This study provides the first evidence for the broad applicability of 3-HβL in targeting ATP-dependent carboxylate-amine ligases.
• The findings suggest potential for developing new antifolates with significant therapeutic applications.