We present CSD-CrossMiner, a novel tool for pharmacophore-based searches in crystal structure databases. Intuitive pharmacophore queries describing, among others, protein-ligand interaction patterns, ligand scaffolds, or protein environments can be built and modified interactively. Matching crystal structures are overlaid onto the query and visualized as soon as they are available, enabling the researcher to quickly modify a hypothesis on the fly. We exemplify the utility of the approach by showing applications relevant to real-world drug discovery projects, including the identification of novel fragments for a specific protein environment or scaffold hopping. The ability to concurrently search protein-ligand binding sites extracted from the Protein Data Bank (PDB) and small organic molecules from the Cambridge Structural Database (CSD) using the same pharmacophore query further emphasizes the flexibility of CSD-CrossMiner. We believe that CSD-CrossMiner closes an important gap in mining structural data and will allow users to extract more value from the growing number of available crystal structures.

Small modifications of the molecular structure of a ligand sometimes cause strong gains in binding affinity to a protein target, rendering a weakly active chemical series suddenly attractive for further optimization. Our goal in this study is to better rationalize and predict the occurrence of such interaction hot-spots in receptor binding sites. To this end, we introduce two new concepts into the computational description of molecular recognition. First, we take a broader view of noncovalent interactions and describe protein-ligand binding with a comprehensive set of favorable and unfavorable contact types, including for example halogen bonding and orthogonal multipolar interactions. Second, we go beyond the commonly used pairwise additive treatment of atomic interactions and use a small world network approach to describe how interactions are modulated by their environment. This approach allows us to capture local cooperativity effects and considerably improves the performance of a newly derived empirical scoring function, ScorpionScore. More importantly, however, we demonstrate how an intuitive visualization of key intermolecular interactions, interaction networks, and binding hot-spots supports the identification and rationalization of tight ligand binding.

A clustering method based on finding the largest set of disconnected fragments that two chemical compounds have in common is shown to be able to group structures in a way that is ideally suited to medicinal chemistry programs. We describe how markedly improved results can be obtained by using a similarity metric that accounts not just for the size of the shared fragments but also on their relative arrangement in the two parent compounds. The use of a physiochemical atom typing scheme is also shown to provide significant contributions. Results from calculations using a test set consisting of actives from nine different important biological target proteins demonstrate the strengths of our clustering method and the advantages over other approaches that are widely used throughout the pharmaceutical industry.

Two methods for the synthesis of 4-acetylamino benzoic acids substituted at the 3-position with imidazoles are described. Many of the compounds are inhibitors of influenza virus sialidases with levels of activity similar to the recently described 4-acetylamino-3-guanidino-benzoic acid (BANA 113).

4-Amino- and 4-guanidino-4H-pyran-6-carboxamides 4 and 5 related to zanamivir (GG167) are a new class of inhibitors of influenza virus sialidases. Structure-activity studies reveal that, in general, secondary amides are weak inhibitors of both influenza A and B viral sialidases. However, tertiary amides, which contain one or more small alkyl groups, show much greater inhibitory activity, particularly against the influenza A virus enzyme. The sialidase inhibitory activities of these compounds correlate well with their in vitro antiviral efficacy, and several of the most potent analogues displayed useful antiviral activity in vivo when evaluated in a mouse model of influenza A virus infection. Carboxamides which were highly active sialidase inhibitors in vitro also showed good antiviral activity in the mouse efficacy model of influenza A infection when administered intranasally but displayed modest activity when delivered by the intraperitoneal route.

The first paper in this series (see previous article) described structure-activity studies of carboxamide analogues of zanamivir binding to influenza virus sialidase types A and B and showed that inhibitory activity of these compounds was much greater against influenza A enzyme. To understand the large differences in affinities, a number of protein-ligand complexes have been investigated using crystallography and molecular dynamics. The crystallographic studies show that the binding of ligands containing tertiary amide groups is accompanied by the formation of an intramolecular planar salt bridge between two amino acid residues in the active site of the enzyme. It is proposed that the unexpected strong binding of these inhibitors is a result of the burial of hydrophobic surface area and salt-bridge formation in an environment of low dielectric. In sialidase from type A virus, binding of the carboxamide moeity and salt-bridge formation have only a minor effect on the positions of the surrounding residues, whereas in type B enzyme, significant distortion of the protein is observed. The results suggest that the decreased affinity in enzyme from influenza B is directly correlated with the small changes that occur in the amino acid residue interactions accompanying ligand binding. Molecular dynamics calculations have shown that the tendency for salt-bridge formation is greater in influenza A sialidase than influenza B sialidase and that this tendency is a useful descriptor for the prediction of inhibitor potency.

A series of substituted 4-amino-4H-pyran-2-carboxylic acid 6-triazoles and oxadiazoles are described. The compounds are inhibitors of influenza virus sialidases which, in general, show a similar pattern of activity and selectivity to recently reported 4H-pyran-2-carboxylic acid 6-carboxamides.

A molecular dynamics/energy-minimisation protocol has been used to analyse the structural and energetic effects of functional group substitution on the binding of a series of C4-modified 2-deoxy-2,3-didehydro-N-acetylneuraminic acid inhibitors to influenza virus sialidase. Based on the crystal structure of sialidase, a conformational searching protocol, incorporating multiple randomisation steps in a molecular dynamics simulation was used to generate a range of minimum-energy structures. The calculations were useful for predicting the number, location, and orientation of structural water molecules within protein-ligand complexes. Relative binding energies were calculated for the series of complexes using several empirical molecular modelling approaches. Energies were computed using molecular-mechanics-derived interactions as the sum of pairwise atomic nonbonded energies, and in a more rigorous manner including solvation effects as the change in total electrostatic energy of complexation, using a continuum-electrostatics (CE) approach. The CE approach exhibited the superior correlation with observed affinities. Both methods showed definite trends in observed and calculated binding affinities; in both cases inhibitors with a positively charged C4 substituent formed the tightest binding to the enzyme, as observed experimentally.

In this article we describe how the World Wide Web (WWW or Web) has been employed to provide access to computational chemistry software and protein structure data via program macros. We show how the combination of Web technology and macros can automate both the running of chemistry software and the execution of complex operations on protein structures. The current version of the system supports the molecular visualization packages GRASP, RASMOL, MOLVIEWER-OGL and INSIGHT95, and the ligand design tool GRID and includes more than 175 in-house protein-ligand complexes. The approach enables inexperienced users to confidently make full use of sophisticated modeling techniques by offering only sensible options, hiding parameter settings, and controlling program invocation and macro execution. Our interface provides both the expert and non-expert alike with powerful tools for protein structure visualization, molecular modeling, and rational drug design.

The structure-activity relationships of a series of 4-amino and guanidino-4H-pyran-2-carboxylic acid 6-carboxamides are described. These compounds represent a new class of inhibitor of influenza sialidases and are particularly active against influenza A sialidase. The binding of the N-phenethyl-N-propylamide 41 to influenza A and B sialidases has been investigated using X-ray crystallography and molecular dynamics simulations. Our results suggest that formation of a hitherto unobserved intramolecular salt bridge within the enzymes may account for the observed activity and selectivity of the series.

The synthesis of a series of 2-(5-fluoro-1H-indol-3-yl)ethyl spiropiperidines is described together with their tachykinin NK2 receptor affinities measured in a rat colon binding assay. Equivalent NK2 receptor binding affinity was observed for the spirooxazolidinone 3-benzyl-8-[2-(5-fluoro-1H-indol-3-yl)ethyl]-1-oxa-3,8-diazaspiro[4,5]decan-2-one (3a), the imidazolidinone 3-benzyl-8-[2-(5-fluoro-1H-indol-3-yl)ethyl]-1,3,8-triazaspiro[4,5]decan-2-one (3s), and the pyrrolidinone 2-benzyl-8-[2-(5-fluoro-1H-indol-3-yl)ethyl]-2,8-diazaspiro[4,5]decan-3-one (3t). Substitution in the phenyl ring of compound 3a produced no significant enhancement in NK2 binding affinity. Replacement of the phenyl ring in 3a with other aromatic rings resulted in a significant loss in binding affinity. Compound 3a was shown to be a potent NK2 receptor antagonist in guinea pig trachea where it also demonstrated 1000-fold selectivity for NK2 receptors over NK1. In the anesthetized guinea pig, compound 3a administered by the intravenous or oral route displayed potent and long-lasting antagonist activity against NK2 receptor agonist induced bronchoconstriction.

A mathematical model has been developed for the systematic calculation of the ligand arrangements in metal carbonyl clusters. The model is founded on the assumption that the ligand shell geometries are dependent upon: (a) the ligand-ligand repulsions, (b) the geometries of the metal cores and (c) the electronic interactions between the metal cores and the ligands. The agreement between minimum energy configurations and the structures determined by crystallography is good for nearly all small and medium sized metal carbonyl clusters [M(x)(CO)y](x = 2 to 7, y = 8 to 21). The model is described, together with the results of the calculations performed for the octahedral metal clusters systems [M6(CO)16], [M6(CO)17] and [M6(CO)18]. The presentation of the results is facilitated by use of a scheme that separates the minimum energy structures into groups according to the number and type of bridging ligands they contain.