Structural basis for potent inhibition of d-amino acid oxidase by thiophene carboxylic acids

A series of thiophene-2-carboxylic acids and thiophene-3-carboxylic acids were identified as a new class of DAO inhibitors. Structure-activity relationship (SAR) studies revealed that small substituents are well-tolerated on the thiophene ring of both the 2-carboxylic acid and 3-carboxylic acid scaffolds. Crystal structures of human DAO in complex with potent thiophene carboxylic acids revealed that Tyr224 was tightly stacked with the thiophene ring of the inhibitors, resulting in the disappearance of the secondary pocket observed with other DAO inhibitors. Molecular dynamics simulations of the complex revealed that Tyr224 preferred the stacked conformation irrespective of whether Tyr224 was stacked or not in the initial state of the simulations. MM/GBSA indicated a substantial hydrophobic interaction between Tyr244 and the thiophene-based inhibitor. In addition, the active site was tightly closed with an extensive network of hydrogen bonds including those from Tyr224 in the stacked conformation. The introduction of a large branched side chain to the thiophene ring markedly decreased potency. These results are in marked contrast to other DAO inhibitors that can gain potency with a branched side chain extending to the secondary pocket due to Tyr224 repositioning. These insights should be of particular importance in future efforts to optimize DAO inhibitors with novel scaffolds

Inhibition of xc⁻ transporter-mediated cystine uptake by sulfasalazine analogs

A series of sulfasalazine analogs were synthesized and tested for their ability to block cystine-glutamate antiporter system xc⁻ using L-[(14)C]cystine as a substrate. Replacement of sulfasalazine\u27s diazo group with an alkyne group led to an equally potent inhibitor, 2-hydroxy-5-((4-(N-pyridin-2-ylsulfamoyl)phenyl)ethynyl)benzoic acid 6. Our SAR studies also revealed that the carboxylate group of sulfasalazine is essential for its inhibitory activity while the phenolic hydroxyl group is dispensable. Truncated analogs lacking an N-pyridin-2-ylsulfamoyl moiety were less potent than sulfasalazine, but may serve as more tractable templates because of their low molecular weight by applying a variety of fragment growing approaches. Given that sulfasalazine is rapidly metabolized through cleavage of the diazo bond, these analogs may possess a more desirable pharmacological profile as system xc- blockers, in particular, for in vivo studies

Structural and computational basis for potent inhibition of glutamate carboxypeptidase II by carbamate-based inhibitors

A series of carbamate-based inhibitors of glutamate carboxypeptidase II (GCPII) were designed and synthesizedusing ZJ-43, N-[[[(1S)-1-carboxy-3-methylbutyl]amino]carbonyl]-L-glutamic acid, as a molecular template inorder to better understand the impact of replacing one of the two nitrogen atoms in the urea-based GCPIIinhibitor with an oxygen atom. Compound 7 containing a C-terminal 2-oxypentanedioic acid was more potentthan compound 5 containing a C-terminal glutamic acid (2-aminopentanedioic acid) despite GCPII’s preferencefor peptides containing an N-terminal glutamate as substrates. Subsequent crystallographic analysis revealedthat ZJ-43 and its two carbamate analogs 5 and 7 with the same (S,S)-stereochemical configuration adopt anearly identical binding mode while (R,S)-carbamate analog 8 containing a D-leucine forms a less extensivehydrogen bonding network. QM and QM/MM calculations have identified no specific interactions in the GCPIIactive site that would distinguish ZJ-43 from compounds 5 and 7 and attributed the higher potency of ZJ-43 andcompound 7 to the free energy changes associated with the transfer of the ligand from bulk solvent to the proteinactive site as a result of the lower ligand strain energy and solvation/desolvation energy. Our findings underscorea broader range of factors that need to be taken into account in predicting ligand-protein binding affinity.These insights should be of particular importance in future efforts to design and develop GCPII inhibitors foroptimal inhibitory potency

6‑Hydroxy-1,2,4-triazine-3,5(2<i>H</i>,4<i>H</i>)‑dione Derivatives as Novel d‑Amino Acid Oxidase Inhibitors

A series of 2-substituted 6-hydroxy-1,2,4-triazine-3,5­(2<i>H</i>,4<i>H</i>)-dione derivatives were synthesized as inhibitors of d-amino acid oxidase (DAAO). Many compounds in this series were found to be potent DAAO inhibitors, with IC₅₀ values in the double-digit nanomolar range. The 6-hydroxy-1,2,4-triazine-3,5­(2<i>H</i>,4<i>H</i>)-dione pharmacophore appears metabolically resistant to <i>O</i>-glucuronidation unlike other structurally related DAAO inhibitors. Among them, 6-hydroxy-2-(naphthalen-1-ylmethyl)-1,2,4-triazine-3,5­(2<i>H</i>,4<i>H</i>)-dione <b>11h</b> was found to be selective over a number of targets and orally available in mice. Furthermore, oral coadministration of d-serine with <b>11h</b> enhanced the plasma levels of d-serine in mice compared to the oral administration of d-serine alone, demonstrating its ability to serve as a pharmacoenhancer of d-serine

Synthesis and SAR of 1‑Hydroxy‑1<i>H</i>‑benzo[<i>d</i>]imidazol-2(3<i>H</i>)‑ones as Inhibitors of d‑Amino Acid Oxidase

A series of 1-hydroxy-1<i>H</i>-benzo­[<i>d</i>]­imidazol-2­(3<i>H</i>)-ones were synthesized and evaluated for their ability to inhibit human and porcine forms of d-amino acid oxidase (DAAO). The inhibitory potency is largely dependent on the size and position of substituents on the benzene ring with IC₅₀ values of the compounds ranging from 70 nM to greater than 100 μM. Structure–activity relationships of this new class of DAAO inhibitors will be presented in detail along with comparisons to previously published SAR data from other classes of DAAO inhibitors. Two of these compounds were given to mice orally together with d-serine to assess their effects on plasma d-serine pharmacokinetics