Giang Thanh Le

Differential effects of selective HDAC inhibitors on macrophage inflammatory responses to the Toll‐like receptor 4 agonist LPS

Broad‐spectrum inhibitors of HDACs are therapeutic in many inflammatory disease models but exacerbated disease in a mouse model of atherosclerosis. HDAC inhibitors have anti‐ and proinflammatory effects on macrophages in vitro. We report here that several broad‐spectrum HDAC inhibitors, including TSA and SAHA, suppressed the LPS‐induced mRNA expression of the proinflammatory mediators Edn‐1, Ccl‐7/MCP‐3, and Il‐12p40 but amplified the expression of the proatherogenic factors Cox‐2 and Pai‐1/serpine1 in primary mouse BMM. Similar effects were also apparent in LPS‐stimulated TEPM and HMDM. The pro‐ and anti‐inflammatory effects of TSA were separable over a concentration range, implying that individual HDACs have differential effects on macrophage inflammatory responses. The HDAC1‐selective inhibitor, MS‐275, retained proinflammatory effects (amplification of LPS‐induced expression of Cox‐2 and Pai‐1 in BMM) but suppressed only some inflammatory responses. In contrast, 17a (a reportedly HDAC6‐selective inhibitor) retained anti‐inflammatory but not proinflammatory properties. Despite this, HDAC6−/− macrophages showed normal LPS‐induced expression of HDAC‐dependent inflammatory genes, arguing that the anti‐inflammatory effects of 17a are not a result of inhibition of HDAC6 alone. Thus, 17a provides a tool to identify individual HDACs with proinflammatory properties.

Antifibrotic activity of an inhibitor of histone deacetylases in DOCA-salt hypertensive rats

Background and purpose:  Histone deacetylases (HDACs) silence genes by deacetylating lysine residues in histones and other proteins. HDAC inhibitors represent a new class of compounds with anti‐inflammatory activity. This study investigated whether treatment with a broad spectrum HDAC inhibitor, suberoylanilide hydroxamic acid (SAHA), would prevent cardiac fibrosis, part of the cardiovascular remodelling in deoxycorticosterone acetate (DOCA)‐salt rats.

Potent Antimalarial Activity of Histone Deacetylase Inhibitor Analogues

ABSTRACT The malaria parasite Plasmodium falciparum has at least five putative histone deacetylase (HDAC) enzymes, which have been proposed as new antimalarial drug targets and may play roles in regulating gene transcription, like the better-known and more intensively studied human HDACs (hHDACs). Fourteen new compounds derived from l-cysteine or 2-aminosuberic acid were designed to inhibit P. falciparum HDAC-1 (PfHDAC-1) based on homology modeling with human class I and class II HDAC enzymes. The compounds displayed highly potent antiproliferative activity against drug-resistant (Dd2) or drug sensitive (3D7) strains of P. falciparum in vitro (50% inhibitory concentration of 13 to 334 nM). Unlike known hHDAC inhibitors, some of these new compounds were significantly more toxic to P. falciparum parasites than to mammalian cells. The compounds inhibited P. falciparum growth in erythrocytes at both the early and late stages of the parasites life cycle and caused altered histone acetylation patterns (hyperacetylation), which is a marker of HDAC inhibition in mammalian cells. These results support PfHDAC enzymes as being promising targets for new antimalarial drugs.

Modulating human proteinase activated receptor 2 with a novel antagonist (GB88) and agonist (GB110)

BACKGROUND AND PURPOSE Many cells express proteinase activated receptor 2 (PAR2) on their plasma membrane. PAR2 is activated by proteolytic enzymes, such as trypsin and tryptase that cleave the receptor N‐terminus, inititating signalling to intracellular G proteins. Studies on PAR2 have relied heavily upon activating effects of proteases and peptide agonists that lack stability and bioavailability in vivo.

Molecular diversity through sugar scaffolds.

Monosaccharides provide an excellent platform to tailor molecular diversity by appending desired substituents at selected positions around the sugar scaffold. The presence of five functionalized and stereo-controlled centres on the sugar scaffolds gives the chemist plenty of scope to custom design molecules to a pharmacophore model. This review focuses on the peptidomimetic developments in this area, as well as the concept of tailoring structural and functional diversity in a library using carbohydrate scaffolds and how this can lead to increased hit rates and rapid identification of leads, which has promising prospects for drug development.

Novel agonists and antagonists for human protease activated receptor 2.

Human protease activated receptor 2 (PAR2) is a G protein-coupled receptor that is associated with inflammatory diseases and cancers. PAR2 is activated by serine proteases that cleave its N-terminus and by synthetic peptides corresponding to the new N-terminus. Peptide agonists are widely used to characterize physiological roles for PAR2 but typically have low potency (e.g., SLIGKV-NH(2), SLIGRL-NH(2)), uncertain target selectivity, and poor bioavailability, limiting their usefulness for specifically interrogating PAR2 in vivo. Structure-activity relationships were used to derive new PAR2 agonists and antagonists containing nonpeptidic moieties. Agonist GB110 (19, EC(50) 0.28 μM) selectively induced PAR2-, but not PAR1-, mediated intracellular Ca(2+) release in HT29 human colorectal carcinoma cells. Antagonist GB83 (36, IC(50) 2 μM) is the first compound at micromolar concentrations to reversibly inhibit PAR2 activation by both proteases and other PAR2 agonists (e.g., trypsin, 2f-furoyl-LIGRLO-NH(2), 19). The new compounds are selective for PAR2 over PAR1, serum stable, and suitable for modulating PAR2 in disease models.

Agonists and Antagonists of Protease Activated Receptors (PARs)

Protease activated receptors (PARs) are a category of G-protein coupled receptors (GPCRs) implicated in the progression of a wide range of diseases, including thrombosis, inflammatory disorders, and proliferative diseases. Signal transduction via PARs proceeds via an unusual activation mechanism. Instead of being activated through direct interaction with an extracellular signal like most GPCRs, they are self-activated following cleavage of their extracellular N-terminus by serine proteases to generate a new receptor N-terminus that acts as an intramolecular ligand by folding back onto itself and triggering receptor activation. Short synthetic peptides corresponding to this newly exposed N-terminal tethered ligand can activate three of the four known PARs in the absence of proteases, and such PAR activating peptides (PAR-APs) have served as templates for agonist/antagonist development. In fact much of the evidence for involvement of PARs in diseases has relied upon use of PAR-APs, often of low potency and uncertain selectivity. This review summarizes current structures of PAR agonists and antagonists, the need for more selective and more potent PAR ligands that activate or antagonize this intriguing class of receptors, and outlines the background relevant to PAR activation, assay methods, and physiological properties anticipated for PAR ligands.

Inhibitors of TACE and Caspase-1 as anti-inflammatory drugs

TNF-alpha neutralising agents such as Infliximab (Remicade), Etanercept (Enbrel) and the IL-1 receptor antagonist Anakinra (Kineret), are currently used clinically for the treatment of many inflammatory diseases such as Crohns disease, rheumatoid arthritis, ankylosing spondylitis, juvenile rheumatoid arthritis, psoriatic arthritis and psoriasis. These protein preparations are expensive to manufacture and administer, need to be injected and can cause allergic reactions. An alternative approach to lowering the levels of TNF-alpha and IL-1beta in inflammatory disease, is to inhibit the enzymes that generate these cytokines using cheaper small molecules. This paper is a broad overview of the progress that has been achieved so far, with respect to small molecule inhibitor design and pharmacological studies (in animals and humans), for the metalloprotease Tumour Necrosis Factor-alpha Converting Enzyme (TACE) and the cysteine protease Caspase-1 (Interleukin-1beta Converting Enzyme, ICE). Inhibitors of these two enzymes are currently considered to be good therapeutic targets that have the potential to provide relatively inexpensive and orally bioavailable anti-inflammatory agents in the future.

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.