Design of Novel, Potent, Noncovalent Inhibitors of Thrombin with Nonbasic P-1 Substructures:  Rapid Structure−Activity Studies by Solid-Phase Synthesis

Wc, Lumma; Km, Witherup; Tj, Tucker; Sf, Brady; Jt, Sisko; Am, Naylor-Olsen; Sd, Lewis; Bj, Lucas; Jp, Vacca

doi:10.1021/jm9706933

Cited by 83 publications

(59 citation statements)

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“…1A), which is usually called an induced fit effect (10,11,18). Thrombin, a serine protease, is a very important drug target in the coagulation cascade for many thromboembolic diseases such as deep vein thrombosis, myocardial infarction, and pulmonary embolism (20,21,22). With the discovery of a neutral P1 substitute of the native substrates, a new generation of more potent inhibitors were designed with high levels of bioavailability and good pharmacokinetic properties, among which 2-…”

Section: Resultsmentioning

confidence: 99%

“…are representative (20,22). In the binding complexes of CDA and CDB, the structures of the protein are essentially the same.…”

Section: Resultsmentioning

confidence: 99%

“…The computational cost of this method is comparable with normal FEP, and it can be very easily generalized to more complicated systems of pharmaceutical interest. We apply this method on two systems; (a) the L99A mutant of the T4 lysozyme (T4L/L99A), (18,19) a popular model system with an engineered nonpolar binding pocket where the structural reorganization happens in the protein, and (b) thrombin (Factor IIa), (20,21,22) an important drug target in the coagulation cascade where the structural reorganization happens in the ligand. (See Fig.…”

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confidence: 99%

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On achieving high accuracy and reliability in the calculation of relative protein–ligand binding affinities

Wang

Berne²,

Friesner³

2012

Proc. Natl. Acad. Sci. U.S.A.

230

351

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We apply a free energy perturbation simulation method, free energy perturbation/replica exchange with solute tempering, to two modifications of protein-ligand complexes that lead to significant conformational changes, the first in the protein and the second in the ligand. The approach is shown to facilitate sampling in these challenging cases where high free energy barriers separate the initial and final conformations and leads to superior convergence of the free energy as demonstrated both by consistency of the results (independence from the starting conformation) and agreement with experimental binding affinity data. The second case, consisting of two neutral thrombin ligands that are taken from a recent medicinal chemistry program for this interesting pharmaceutical target, is of particular significance in that it demonstrates that good results can be obtained for large, complex ligands, as opposed to relatively simple model systems. To achieve quantitative agreement with experiment in the thrombin case, a next generation force field, Optimized Potentials for Liquid Simulations 2.0, is required, which provides superior charges and torsional parameters as compared to earlier alternatives.enhanced sampling | protein-ligand binding affinity | structural reorganization | lead optimization | drug design B iological processes often depend on protein-ligand binding so that accurate prediction of protein-ligand binding affinities is of central importance in structural based drug design (1-4). Among the existing methods used to calculate these binding affinities in explicit solvent, free energy perturbation (FEP) simulations provide one of the most rigorous simulation techniques. Usually FEP is applied in the lead optimization stage of structure based drug design and is used to rank-order a series of congeneric ligands to choose the most potent ones for further investigation (1-4).Despite the potentially large impact that FEP could have on structure based drug design projects, practical applications in an industrial context have been limited over the past decade. High accuracy and reliability in the methodology are required to make productive decisions about compound modification during late stage lead optimization, but neither has yet been demonstrated by existing implementations. Two types of challenges stand in the way of developing FEP into a true engineering platform for drug candidate optimization. Firstly, converging explicit solvent simulations to the desired precision is far from trivial, even with the immense computing power that is currently available using low cost multiprocessor clusters or cloud computing platforms of various types. Secondly, errors in the potential energy models must be reduced to the point where they lead to errors in a converged calculation that are smaller than the desired errors in relative binding affinities compared to experiment. In our estimation these errors are on the order of 0.5-1.0 kcal∕mole mean unsigned error for a typical late stage lead optimization effort in a drug dis...

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Section: Resultsmentioning

confidence: 99%

“…are representative (20,22). In the binding complexes of CDA and CDB, the structures of the protein are essentially the same.…”

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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On achieving high accuracy and reliability in the calculation of relative protein–ligand binding affinities

Wang

Berne²,

Friesner³

2012

Proc. Natl. Acad. Sci. U.S.A.

230

351

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“…However Lumma et al decided to use the P1 benzyl derivative as a lead for the development of neutral P1 analogs of 22 in order to improve the oral bioavailability of this series of compounds. 71 Again taking advantage of the simplified synthesis of the noncovalent inhibitors, resinbased rapid analog synthesis was used to prepare a variety of compounds with uncharged P1 groups. This led to the development of a 2,5-dichlorophenyl derivative 29 which is a potent and highly selective compound (the selectivity of 29 is consistent with the proposal that the active site of thrombin is larger and more hydrophobic than that of trypsin).…”

Section: • Sandersonmentioning

confidence: 99%

Small, noncovalent serine protease inhibitors

Sanderson

1999

Med. Res. Rev.

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“…65 The complete elimination of a basic P1 residue was tolerated, compensated for introduction of chloro substituents to form favourable interactions with the S1 pocket (Tyr228). 65 1.8 replaced the highly lipophilic diphenylalanine P3-residue with Dcyclohexylglycine and extra polar substituent for hydrogen bonding was introduced through an ether link to the 2-position of P1.…”

Section: 4mentioning

confidence: 99%

Towards serine protease inhibitors

Chow¹

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Serine proteases are among the most important proteolytic enzymes and important mediators of a wide range of physiological processes. Thus potent and selective inhibitors of these proteases are viable therapeutic candidates, since dysregulation of serine proteases has been linked to many diseases. The aim of this thesis is to design and synthesize inhibitors of trypsin-like serine proteases, including some proteases for which inhibitor development has proved elusive to date, and evaluate their effects in enzyme assays relevant to inflammatory and infectious diseases.Chapter 1 reviews the biological importance of serine proteases, their mode(s) of action, the approaches employed to develop inhibitors and the current status of the field of inhibiting the serine proteases: Tryptase, Dengue Virus NS2B/NS3pro, West Nile Virus NS2B/NS3pro and Complement Factor B.Chapter 2 focuses on human mast cell ß-tryptase, a trypsin-like serine protease implicated in airway diseases, inflammatory conditions, cancer and tumour growth. A binding pocket located at the S2' position, not previously exploited in inhibitor development, was examined and successfully targeted using two approaches. The first approach involved a rationally designed P1 scaffold, where moderately active hybrid tryptase inhibitors were produced and the S2' pocket was identified as a viable target pocket. Electrophilic isosteres (aldehyde and α-keto amide) were used in a second approach producing potent and selective inhibitors of the human tryptase.Chapter 3 describes a peptidomimetic approach to afford inhibitors of two proteases, DENV NS2B/NS3pro and WNV NS2B/NS3pro, that are essential for replication of Dengue and West Nile viruses respectively. The basic residues of the parent tripeptide inhibitors were replaced by non-natural amino acids and the binding requirements of subsites were mapped. Rationally designed amino acid mimetics complementary to the target subsite were identified, and active peptidomimetic inhibitors of DENV and WNV NS2B/NS3pro were produced with moderate success.Chapter 4 examines two isolated serine proteases, DENV NS2B/NS3pro protease and complement Factor B protease, that are catalytically inactive at neutral pH and display optimal proteolytic activity under basic conditions. The oxyanion holes of these two proteases were found to be defective in the ligand-free state, however full restoration of function was observed in a ligandbound state. An electrophilic isostere, α-keto amide, was incorporated in peptidic inhibitors to target the oxyanion hole residues and to allow occupation of the prime-binding sites of these two proteases, but only moderately potent inhibitors were identified for these proteases using this approach, despite it being successful for tryptase.iv Chapter 5 employs a screening campaign to identify potential new inhibitors of DENV NS2B/NS3pro. A library of ~2000 structurally diverse natural products was assayed against DENV NS2B-NS3pro. Phloeodictine X was successfully identified as a potent inhibitor of thi...

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Design of Novel, Potent, Noncovalent Inhibitors of Thrombin with Nonbasic P-1 Substructures: Rapid Structure−Activity Studies by Solid-Phase Synthesis

Cited by 83 publications

References 5 publications

On achieving high accuracy and reliability in the calculation of relative protein–ligand binding affinities

On achieving high accuracy and reliability in the calculation of relative protein–ligand binding affinities

Small, noncovalent serine protease inhibitors

Towards serine protease inhibitors

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