Discovery of a small-molecule HIV-1 integrase inhibitor-binding site

Herein, we report the identification of a unique HIV-1 integrase (IN) inhibitor-binding site using photoaffinity labeling and mass spectrometric analysis. We chemically incorporated a photo-activatable benzophenone moiety into a series of coumarin-containing IN inhibitors. A representative of this series was covalently photo-crosslinked with the IN core domain and subjected to HPLC purification. Fractions were subsequently analyzed by using MALDI-MS and electrospray ionization (ESI)-MS to identify photo-crosslinked products. In this fashion, a single binding site for an inhibitor located within the tryptic peptide 128AACWWAGIK136 was identified. Site-directed mutagenesis followed by in vitro inhibition assays resulted in the identification of two specific amino acid residues, C130 and W132, in which substitutions resulted in a marked resistance to the IN inhibitors. Docking studies suggested a specific disruption in functional oligomeric IN complex formation. The combined approach of photo-affinity labeling/MS analysis with site-directed mutagenesis/molecular modeling is a powerful approach for elucidating inhibitor-binding sites of proteins at the atomic level. This approach is especially important for the study of proteins that are not amenable to traditional x-ray crystallography and NMR techniques. This type of structural information can help illuminate processes of inhibitor resistance and thereby facilitate the design of more potent second-generation inhibitors

Drug-Class Specific Impact of Antivirals on the Reproductive Capacity of HIV

Predictive markers linking drug efficacy to clinical outcome are a key component in the drug discovery and development process. In HIV infection, two different measures, viral load decay and phenotypic assays, are used to assess drug efficacy in vivo and in vitro. For the newly introduced class of integrase inhibitors, a huge discrepancy between these two measures of efficacy was observed. Hence, a thorough understanding of the relation between these two measures of drug efficacy is imperative for guiding future drug discovery and development activities in HIV. In this article, we developed a novel viral dynamics model, which allows for a mechanistic integration of the mode of action of all approved drugs and drugs in late clinical trials. Subsequently, we established a link between in vivo and in vitro measures of drug efficacy, and extract important determinants of drug efficacy in vivo. The analysis is based on a new quantity—the reproductive capacity—that represents in mathematical terms the in vivo analog of the read-out of a phenotypic assay. Our results suggest a drug-class specific impact of antivirals on the total amount of viral replication. Moreover, we showed that the (drug-)target half life, dominated by immune-system related clearance processes, is a key characteristic that affects both the emergence of resistance as well as the in vitro–in vivo correlation of efficacy measures in HIV treatment. We found that protease- and maturation inhibitors, due to their target half-life, decrease the total amount of viral replication and the emergence of resistance most efficiently

Three-Dimensional Quantitative Structural Activity Relationship (3D-QSAR) Studies of Some 1,5-Diarylpyrazoles: Analogue Based Design of Selective Cyclooxygenase-2 Inhibitors

Selective cyclooxygenase inhibitors have attracted much attention in recent times in the design of new non-steroidal anti-inflammatory drugs (NSAID). 3D-QSAR studies have been performed on a series of 1,5-diarylpyrazoles that act as selective cyclooxygenase-2 (COX-2) inhibitors, using three different methods: comparative molecular field analysis (CoMFA) with partial least squares (PLS) fit; molecular field analysis (MFA) and; receptor surface analysis (RSA) with genetic function algorithms (GFA). The analyses were carried out on 30 analogues of which 25 were used in the training set and the rest considered for the test set. These studies produced reasonably good predictive models with high cross-validated and conventional r2 values in all the three cases

Three-Dimensional Quantitative Structural Activity Relationship (3D-QSAR) Studies of Some 1,5-Diarylpyrazoles: Analogue Based Design of Selective Cyclooxygenase-2 Inhibitors

Selective cyclooxygenase inhibitors have attracted much attention in recent times in the design of new non-steroidal anti-inflammatory drugs (NSAID). 3D-QSAR studies have been performed on a series of 1,5-diarylpyrazoles that act as selective cyclooxygenase-2 (COX-2) inhibitors, using three different methods: comparative molecular field analysis (CoMFA) with partial least squares (PLS) fit; molecular field analysis (MFA) and; receptor surface analysis (RSA) with genetic function algorithms (GFA). The analyses were carried out on 30 analogues of which 25 were used in the training set and the rest considered for the test set. These studies produced reasonably good predictive models with high cross-validated and conventional r2 values in all the three cases

An approach to the stereo-controlled synthesis of polycyclic derivatives of L-4-thiazolidinecarboxilic acid active against HIV-1 integrase

Herein, we describe a new strategy for the preparation of thiazolothiazepine-based inhibitors of human immunodeficiency virus type-1 integrase (IN). The present method allows facile preparation of the title compounds in a single enantiomeric form starting from l-4-thiazolidinecarboxylic acid. This method could be easily extended to the synthesis of several analogs derived from optically active cyclic aminoacids. We also present a putative model showing the interaction between l- and d-isomers of compound 1 in the IN active site. A sensibly lower IC(50) value was found for (-)-1 over racemic-1 in an anti-IN assay