Structure-Based Design of Nonpeptidic Thrombin Inhibitors: Exploring the D-Pocket and the Oxyanion Hole

Betschmann, Patrick; Sahli, Stefan; Diederich, François; Obst, Ulrike; Gramlich, Völker

doi:10.1002/1522-2675(200205)85:5<1210::aid-hlca1210>3.0.co;2-t

Cited by 27 publications

(23 citation statements)

References 22 publications

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“…1,7 The extremely high, 1609-fold selectivity of the 4-chlorophenyl derivative (±)-12 is by far the largest measured for all tricyclic inhibitors prepared so far. 1,2, 16 The inhibitory constants in the series of imide inhibitors (±)-1-(±)-8 vary greatly. If the substituent on the benzyl ring in the D pocket -the only variable in the series -is considered, K i /M increases in the sequence F < OCH 2 O < Cl < H < OMe < OH < N pyr Br (Table 1).…”

Section: Biological Resultsmentioning

confidence: 99%

“…14 In both steps, the nucleophile Scheme 1 Synthesis of the tricyclic imide inhibitors. Reagents and conditions: (i) CH 3 CN, 80 • C, [14][15][16] h; (±)-16 (44%)/(±)-18 (53%), (±)-17 (35%)/(±)-19 (18%), (±)-24 (41%)/(±)-25 (38%); (ii) CuCN, DMF, 165 • C, 24 h,or [Pd 2 (dba) 3 ] dppf, Zn(CN) 2 , DMF, 120 • C, 24 h; (±)-20 (78%), (±)-21 (32%); (iii) MeOH, HCl(g), CH 2 Cl 2 , 4 • C, 28-35 h; (iv) NH 3 , MeOH, 65 • C, 3 h; (±)-5 (49%), (±)-6 (21%), (±)-7 (28%), (±)-8 (40% from (±)-20); (v) BBr 3 , CH 2 Cl 2 , −50 • C → 25 • C, 8 h. DMF = dimethylformamide, dba = dibenzylideneacetone, dppf = diphenylphosphinoferrocene. Exo and endo refer to the orientation of the 4-bromophenyl substituent at C(4) with respect to the bicyclic perhydropyrrolo [3,4-c]pyrrole scaffold, and cis and trans to the position of this 4-bromophenyl ring with respect to the configuration of C(8a) at the fusion of the two pentagons in the perhydropyrrolizidine bicycle (for atom numbering, see Fig.…”

Section: Introductionmentioning

confidence: 99%

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Multipolar interactions in the D pocket of thrombin: large differences between tricyclic imide and lactam inhibitors

et al. 2006

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Two series of tricyclic inhibitors of the serine protease thrombin, imides (+/-)-1-(+/-)-8 and lactams (+/-)-9-(+/-)-13, were analysed to evaluate contributions of orthogonal multipolar interactions with the backbone C=O moiety of Asn98 to the free enthalpy of protein-ligand complexation. The lactam derivatives are much more potent and more selective inhibitors (K(i) values between 0.065 and 0.005 microM, selectivity for thrombin over trypsin between 361- and 1609-fold) than the imide compounds (Ki values between 0.057 and 23.7 microM, selectivity for thrombin over trypsin between 3- and 67-fold). The increase in potency and selectivity is explained by the favorable occupancy of the P-pocket of thrombin by the additional isopropyl substituent in the lactam derivatives. The nature of the substituent on the benzyl ring filling the D pocket strongly influences binding potency in the imide series, with Ki values increasing in the sequence: F < OCH2O < Cl < H < OMe < OH < N(pyr)<< Br. This sequence can be explained by both steric fit and the occurrence of orthogonal multipolar interactions with the backbone C[double bond, length as m-dash]O moiety of Asn98. In contrast, the substituent on the benzyl ring hardly affects the ligand potency in the lactam series. This discrepancy was clarified by the comparison of X-ray structures solved for co-crystals of thrombin with imide and lactam ligands. Whereas the benzyl substituents in the imide inhibitors are sufficiently close (< or =3.5 Angstroms) to the C=O group of Asn98 to allow for attractive orthogonal multipolar interactions, the distances in the lactam series are too large (> or =4 Angstroms) for attractive dipolar contacts to be effective.

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Section: Biological Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Multipolar interactions in the D pocket of thrombin: large differences between tricyclic imide and lactam inhibitors

et al. 2006

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“…7, 9–12 Additionally, subsites S1’, S2’ and S3’ and the oxyanion hole have also been considered in the design of stronger and more selective inhibitors. 10, 14 …”

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Synthesis and biochemical evaluation of triazole/tetrazole-containing sulfonamides against thrombin and related serine proteases

Siles¹,

Kawasaki²,

Ross³

et al. 2011

Bioorganic & Medicinal Chemistry Letters

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A small library of 25 triazole/tetrazole-based sulfonamides have been synthesized and further evaluated for their inhibitory activity against thrombin, trypsin, tryptase and chymase. In general, the triazole-based sulfonamides inhibited thrombin more efficiently than the tetrazole counterparts. Particularly, compound 26 showed strong thrombin inhibition (Ki =880 nM) and significant selectivity against other human related serine proteases like trypsin (Ki =729 µM). Thrombin binding affinity of the same compound was determined by ITC and demonstrated that the binding of this new triazole-based scaffold is enthalpically driven, making it a good candidate for further development.

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“…Early crystal structures revealed a well-defined active site [3] [4], and, accordingly, structure-based design has been intensively applied to the generation of new inhibitors of this enzyme [5 ± 7] (for some recent reports, see [8]). Previous investigations with this class of tricyclic inhibitors addressed in greater detail the molecular-recognition features of the spacious hydrophobic D-pocket (D: distal) [9a,b] [10] [12], the tight hydrophobic P-pocket (P: proximal) [9b] [10], and the selectivity pocket S1 (Fig. 1; inhibitory constant K i 7 nm, selectivity over the digestive serine protease trypsin K i (trypsin)/ K i (thrombin) 740) and confirmed the binding modes of several derivatives at the thrombin active site by crystal-structure analysis [9] [10].…”

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confidence: 99%

“…In particular, we were intrigued by the possibility to bind to the H 2 O molecule found in the oxyanion hole in the crystal structure of ()-1, H-bonded to the NÀH residues of Gly193 and Ser195. A series of tricyclic imide inhibitors was prepared starting with the 1,3-dipolar cycloaddition between the azomethine ylide, formed in situ from l-(4R)-hydroxyproline (3) and 4-bromobenzaldehyde (4), and N-piperonylmaleimide (5) in DMF [9] [12] [15] (Scheme 1). 1) with a H-bond donor center would be particularly suitable for bonding to the H 2 O molecule.…”

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confidence: 99%

Enantiomerically Pure Thrombin Inhibitors for Exploring the Molecular‐Recognition Features of the Oxyanion Hole

Schärer

Morgenthaler

Seiler

et al. 2004

Helvetica Chimica Acta

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A new route via intermediate pseudoenantiomers was developed to synthesize racemic and enantiomerically pure new non‐peptidic inhibitors of thrombin, a key serine protease in the blood‐coagulation cascade. These ligands feature a conformationally rigid tricyclic core and are decorated with substituents to fill the major binding pockets (distal (D), proximal (P), selectivity (S1), and oxyanion hole) at the thrombin active site (Fig. 1). The key step in the preparation of the new inhibitors is the 1,3‐dipolar cycloaddition between an optically active azomethine ylide, prepared in situ from L‐(4R)‐hydroxyproline and 4‐bromobenzaldehyde, and N‐piperonylmaleimide (Scheme 1). According to this protocol, tricyclic imide (compounds (±)‐15‐(±)‐18 and (+)‐21) and lactam (compound (+)‐2) inhibitors with OH or ether substituents at C(7) in the proline‐derived pyrrolidine ring were synthesized to specifically explore the binding features of the oxyanion hole (Schemes 2–4). Biological assays (Table) showed that the polar oxyanion hole in thrombin is not suitable for the accommodation of bulky substituents of low polarity, thereby confirming previous findings. In contrast, tricyclic lactam (+)‐2 (Ki=9 nM, Ki(trypsin)/Ki(thrombin)=1055) and tricyclic imide (+)‐21 (Ki=36 nM, Ki(trypsin)/Ki(thrombin)=50) with OH‐substituents at the (R)‐configured C(7)‐atom are among the most‐potent and most‐selective thrombin inhibitors in their respective classes, prepared today. While initial modeling predicted H‐bonding between the OH group at C(7) in (+)‐2 and (+)‐21 with the H2O molecule bound in the oxyanion hole (Fig. 2), the X‐ray crystal structure of the complex of (+)‐21 (Fig. 7, b) revealed a different interaction for this group. The propionate side chain of Glu192 undergoes a conformational change, thereby re‐orienting towards the OH group at C(7) under formation of a very short ionic H‐bond (OH⋅⋅⋅−OOC; d(O⋅⋅⋅O)=2.4 Å). The energetic contribution of this H‐bond, however, is negligible, due to its location on the surface of the protein and the unfavorable conformation of the H‐bonded propionate side chain.

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Structure-Based Design of Nonpeptidic Thrombin Inhibitors: Exploring the D-Pocket and the Oxyanion Hole

Cited by 27 publications

References 22 publications

Multipolar interactions in the D pocket of thrombin: large differences between tricyclic imide and lactam inhibitors

Multipolar interactions in the D pocket of thrombin: large differences between tricyclic imide and lactam inhibitors

Synthesis and biochemical evaluation of triazole/tetrazole-containing sulfonamides against thrombin and related serine proteases

Enantiomerically Pure Thrombin Inhibitors for Exploring the Molecular‐Recognition Features of the Oxyanion Hole

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