John Joseph Deadman

Parallel Screening of Low Molecular Weight Fragment Libraries Do Differences in Methodology Affect Hit Identification

Fragment screening is becoming widely accepted as a technique to identify hit compounds for the development of novel lead compounds. In neighboring laboratories, we have recently, and independently, performed a fragment screening campaign on the HIV-1 integrase core domain (IN) using similar commercially purchased fragment libraries. The two campaigns used different screening methods for the preliminary identification of fragment hits; one used saturation transfer difference nuclear magnetic resonance spectroscopy (STD-NMR), and the other used surface plasmon resonance (SPR) spectroscopy. Both initial screens were followed by X-ray crystallography. Using the STD-NMR/X-ray approach, 15 IN/fragment complexes were identified, whereas the SPR/X-ray approach found 6 complexes. In this article, we compare the approaches that were taken by each group and the results obtained, and we look at what factors could potentially influence the final results. We find that despite using different approaches with little overlap of initial hits, both approaches identified binding sites on IN that provided a basis for fragment-based lead discovery and further lead development. Comparison of hits identified in the two studies highlights a key role for both the conditions under which fragment binding is measured and the criteria selected to classify hits.

Small Molecule Inhibitors of the Ledgf Site of Human Immunodeficiency Virus Integrase Identified by Fragment Screening and Structure Based Design.

A fragment-based screen against human immunodeficiency virus type 1 (HIV) integrase led to a number of compounds that bound to the lens epithelium derived growth factor (LEDGF) binding site of the integrase catalytic core domain. We determined the crystallographic structures of complexes of the HIV integrase catalytic core domain for 10 of these compounds and quantitated the binding by surface plasmon resonance. We demonstrate that the compounds inhibit the interaction of LEDGF with HIV integrase in a proximity AlphaScreen assay, an assay for the LEDGF enhancement of HIV integrase strand transfer and in a cell based assay. The compounds identified represent a potential framework for the development of a new series of HIV integrase inhibitors that do not bind to the catalytic site of the enzyme.

Structural basis for a new mechanism of inhibition of HIV-1 integrase identified by fragment screening and structure-based design

Background: HIV-1 integrase is a clinically validated therapeutic target for the treatment of HIV-1 infection, with one approved therapeutic currently on the market. This enzyme represents an attractive target for the development of new inhibitors to HIV-1 that are effective against the current resistance mutations. Methods: A fragment-based screening method employing surface plasmon resonance and NMR was initially used to detect interactions between integrase and fragments. The binding sites of the fragments were elucidated by crystallography and the structural information used to design and synthesize improved ligands. Results: The location of binding of fragments to the catalytic core of integrase was found to be in a previously undescribed binding site, adjacent to the mobile loop. Enzyme assays confirmed that formation of enzyme–fragment complexes inhibits the catalytic activity of integrase and the structural data was utilized to further develop these fragments into more potent novel enzyme inhibitors. Conclusions: We have defined a new site in integrase as a valid region for the structure-based design of allosteric integrase inhibitors. Using a structure-based design process we have improved the activity of the initial fragments 45-fold.

Crystal structure of the HIV-1 integrase core domain in complex with sucrose reveals details of an allosteric inhibitory binding site

MINT‐7713125: IN (uniprotkb:P04585) and IN (uniprotkb:P04585) bind (MI:0407) by X‐ray crystallography (MI:0114)

Design of a series of bicyclic HIV-1 integrase inhibitors. Part 1: selection of the scaffold.

HIV integrase inhibitors based on a novel bicyclic pyrimidinone core is presented. Nine variations of the core scaffold are evaluated leading to optimization of the 6:6 core giving compound 48 with an EC(50) of 3 nM against wild type HIV infected T-cells.

The synthesis of 1-aminobenzylphosphonic acids from benzylidenediphenylmethylamines, for use as structural units in antithrombotic tripeptides

Abstract Acid hydrolyses of O , O -dimethyl or O , O -diethyl 1-(diphenylmethylamino) benzylphosphonate intermediates 2 , formed from the addition at elevated temperature of dimethyl or diethyl phosphite to benzylidenediphenylmethylamines 1 , generates 1-aminobenzylphosphonic acids 3 in good yield.

Crystal Structures of Novel Allosteric Peptide Inhibitors of HIV Integrase Identify New Interactions at the LEDGF Binding Site

An optimised method of solution cyclisation gave us access to a series of peptides including SLKIDNLD (2). We investigated the crystallographic complexes of the HIV integrase (HIV‐IN) catalytic core domain with 13 of the peptides and identified multiple interactions at the binding site, including hydrogen bonds with residues Thr125 and Gln95, that have not previously been described as being accessible within the binding site. We show that the peptides inhibit the interaction of lens epithelium‐derived growth factor (LEDGF) with HIV‐IN in a proximity AlphaScreen assay and in an assay for the LEDGF enhancement of HIV‐IN strand transfer. The interactions identified represent a potential framework for the development of new HIV‐IN inhibitors.

Binaphthyl‐Based Dicationic Peptoids with Therapeutic Potential

[Extract] While the cationic glycopeptide vancomycin has long been regarded as the gold standard for the treatment of recalcitrant Gram-positive bacterial infection, this position has been compromised by the emergence of resistant strains. The first report of such resistance emerged in 1988, and has subsequently widened amongst the enterococci and staphylococci, including methicillin-resistant Staphylococcus aureus (MRSA); 1, 2 cross-resistance to linezolid is also a concern. Some recent chemical strategies for overcoming this resistance have centered on other high molecular weight cyclic peptides, elegantly crafted vancomycin11 or vancomycin aglycone analogues, potent dual-action vancomycin/β-lactam hybrid antibiotics,or large vancomycin dimers. An alternative strategy is to design smaller, simpler cationic peptoids with some related design features to vancomycin which could still interact with the altered peptide-glycan cell-wall moiety in both vancomycin-resistant and -sensitive strains and thus broaden the antibacterial spectrum. Svendsen et al. designed minimal cationic peptidomimetics, and a pharmacophore has been developed for dipeptides which includes the presence of two cationic charges and two hydrophobic units of steric bulk. Subsequently, cationic tripeptide analogues were developed that demonstrated good activity against both Gram-positive (including MRSA) and Gram-negative bacteria, but were not evaluated with respect to vancomycin-resistant strains.

Design of a series of bicyclic HIV-1 integrase inhibitors. Part 2: Azoles: Effective metal chelators

Synthesis of a diverse set of azoles and their utilizations as an amide isostere in the design of HIV integrase inhibitors is described. The Letter identified thiazole, oxazole, and imidazole as the most promising heterocycles. Initial SAR studies indicated that these novel series of integrase inhibitors are amenable to lead optimization. Several compounds with low nanomolar inhibitory potency are reported.

Fragment-based design of ligands targeting a novel site on the integrase enzyme of human immunodeficiency virus 1

Fragment-based screening has been used to identify a novel ligand binding site on HIV-1 integrase. Crystal structures of fragments bound at this site (shown) have been used to design elaborated second-generation compounds that bind with higher affinity and good ligand efficiency.