Thrombin benzamidine inhibitors

The LIE method has also been used with continuum electrostatic methods. Nicolotti et al. (27) have published a nice report on this use of the LIE method for screening benzamidine inhibitors of thrombin. Another recent study used the atom-bond electronegativity equalization method on force field with the generalized Born (GB) continuum electrostatics approach to predict binding free energies of HIV-1 protease inhibitors (28). 

Table 2 A, Values Obtained for ftw-Benzamidine Inhibitors of Matriptase and Thrombin...

D, H W Hoeffken, D Crosse, J Stuerzebecher, P D Martin, B F P Edwards and W Bode 1992. Refined 2.3 Angstroms X-Ray Crystal Structure of Bovine Thrombin Complexes Formed witli he 3 Benzamidine and Arginine-Based Thrombin Inhibitors NAPAP, 4-TAPAP and MQPA A Starting Point for Improving Antithrombotics. Journal of Molecular Biology 226 1085-1099. 

Figure 3. Chemical structures of the thrombin inhibitors for which calculations were carried out in Ref. 46 (BZA is benzamidine).

Sugano et al. studied the membrane permeation of 51 benzamidine-based thrombin inhibitors in a rat everted sac permeability model [197]. They reported significant membrane permeabilities in this in vitro model, which they attributed to passive paracellular transport, a different absorption mechanism to transcellular permeability. 

Sugano, K., Yoshida, S., Takaku, M., Haramura, M., Saitoh, R., Nabughi, Y., and Ushio, H. Quantitative structure-intestinal permeability relationship of benzamidine analogue thrombin inhibitor. Bioorg. Med. Chem. Lett. 2000, 10, 1939-1942. 

Brandstetter H, Turk D, Hoeffken W, Grosse D, Sturzebecher J, Martin PD, Edwards BFP, Bode W. X-ray crystal structure of thrombin complexes with the benzamidine- and arginine-base inhibitors NAPAP, 4-TAPAP and MQPA a starting point for elaborating improved antithrombotics. J Mol Biol 1992 226 1085-1099. 

Nicolotti O, Giangreco I, Miscioscia TF et al (2010) Screening of benzamidine-based thrombin inhibitors via a linear interaction energy in continuum electrostatics model. J Comput Aided Mol Des 24(2) 117-129... 

The set of 51 benzamidine-based thrombin inhibitors was taken from the study of Sugano et al. (2000). Experimental rat everted sac permeabilities were expressed as log(ES A) values.2 The experimental permeability in this assay is expressed as ratio of outer (mucosal side) concentration of the drug and inner (serosal side) concentration after 1 h incubation of the everted sac of rat small intestine. All molecules were treated in their neutral form and converted into their 3D structures using CORINA (Sadowski et al. 1992). From GRID molecular interaction fields for water, dry, and carbonyl oxygen probes, a set of 72 VolSurf descriptors (Cruciani et al. 2000) was computed and analysed as described above. 

Fig. 8. Correlation of VolSurf descriptors with intestinal paracellular passive absorption for 51 benzamidine-based thrombin inhibitors. Left Predicted versus experimental permeability log(ESA) from the 4 component PLS model. Right PLS loadings showing the importance of VolSurf descriptors to the prediction of paracellular permeability.

Recently some non-basic Si anchoring groups have been incorporated in the molecules of some thrombin inhibitors [54,115,116]. The presence of guanidine-Zbenzamidino moieties in such compounds was critical. Three series of derivatives using benzamidine and sulfaguanidine as lead molecules have been synthesized and tested (62-64) against human thrombin. Forty... 

Figure 7.6 Fragment generation and matching using the Shapelets approach, (a) A molecular fragment is described by its Shapelets (paraboloids) decomposition of the solvent-accessible surface (mesh), (b) Result of fragments matching to a reference structure. A benzamidine building-block was matched to a thrombin inhibitor by Shapelets-mdiic g.

Based on these findings, two new classes of potent thrombin inhibitors have been discovered by a three-step process [6-8]. First, a collection of small, basic molecules has been tested in thrombin and trypsin assays iV-amidinopiperidine has been selected as being more active than benzamidine and slightly selective for thrombin. Secondly, substituents attached to the 3-position of amidinopiperidine containing two hydrophobic moieties mediate low nano-... 

Human thrombin A lipophilic chlorophenyl fragment was identified by fragment-based screening by X-ray as a suitable substitute for the Si benzamidine group of conventional thrombin inhibitors. 190... 

Fig. 7.3 The simplest thrombin inhibitors, benzamidine 2 (left) and APPA, 3 p-amidino phenyl pyruvate.

Burkhard et al. [159] developed a software tool for rigid-body molecular docking called SAN DOCK. The method is based on matching surfaces with mapped properties and is not described in detail here. The authors applied SAN DOCK to searching for new thrombin inhibitors. One of the hits was p-amino-benzamidine, a small molecule with ten non-hydrogen atoms, was experimentally validated. The structure predicted for the complex was in agreement with the crystallographic structure determined afterwards. 

Figure 8.11 Top Two bioisosteres of a phenolic group (Mewshaw, R.E., et al. New generation dopaminergic agents. Part 8 Heterocyclic bioisosteres that exploit the 7-OH-2-(Aminomethyl)chroman D2 template. Bioorg. Med. Chem. Lett. 2002, 12, 271-274.). Bottom Aminoisoquinoline bioisosteres of a benzamidine (Rewinkel, J.B.M., et al. 1-Aminoisoquinoline as benzamidine isosteres in the design and synthesis of orally active thrombin inhibitors. Bioorg. Med. Chem. Lett. 1999, 9, 685-690.)...

The thrombin inhibitor NAPAP (Figure 8.11) features an amidine group that s crucial for its interaction with an active site aspartate of the enzyme. Unfortunately, benzamidines like these tend to have poor cell permeability. The 1-aminoisoquinoline bioisostere, 11, although itself much less active than NAPAP, led to the synthesis of other aminoisoquinolines like 12, which was almost 10-fold more potent and > 10-fold more cell-permeable than NAPAP. ... 

Rewinkel, J.B.M., et al. 1-Aminoisoquinoline as benzamidine isosteres in the design and synthesis of orally active thrombin inhibitors. Bioorg. Med. Chem. Lett. 1999, 9, 685-690. 

S.3.2 Immobilized Synthetic Thrombin Inhibitors Based on Benzamidine to Reduce Coagulation Activation... 

In the following, we will focus on low thrombogenic material surfaces prepared by covalent attachment of synthetic thrombin inhibitors based on benzamidine (benzenecarboximidamide) [136-138] on the top of macroscopic flat surfaces precoated with reactive maleic anhydride copolymer thin... 

The structures of two benzamidine-type thrombin inhibitors (hirudin analogs), a small benzamidine derivative (1) and a NAPAP (Mx-(2-naphthyl-sulphonyl-glycyl)-DL-p-amidinophenylalanyl-piperidine) analog (2), are displayed in Figure 6.35. The presence of the primary amino group allows for an effective attachment of the active compounds onto the reactive polymeric carrier (Fig. 6.36). 

Consequently, benzamidine-modified material surfaces show a promising potential for the development of thrombin-inhibiting biomaterials to be applied in blood-contacting devices. However, due to their permanent character, immobilized inhibitors are difficult to dose. 

---