Structure, function and pathophysiology of protease activated receptors.

Discovered in the 1990s, protease activated receptors(1) (PARs) are membrane-spanning cell surface proteins that belong to the G protein coupled receptor (GPCR) family. A defining feature of these receptors is their irreversible activation by proteases; mainly serine. Proteolytic agonists remove the PAR extracellular amino terminal pro-domain to expose a new amino terminus, or tethered ligand, that binds intramolecularly to induce intracellular signal transduction via a number of molecular pathways that regulate a variety of cellular responses.

Amyloid formation from an α-helix peptide bundle is seeded by 3(10)-helix aggregates.

Transformation of proteins and peptides to fibrillar aggregates rich in β sheets underlies many diseases, but mechanistic details of these structural transitions are poorly understood. To simulate aggregation, four equivalents of a water-soluble, α-helical (65 %) amphipathic peptide (AEQLLQEAEQLLQEL) were assembled in parallel on an oxazole-containing macrocyclic scaffold. The resulting 4α-helix bundle is monomeric and even more α helical (85 %), but it is also unstable at pH 4 and undergoes concentration-dependent conversion to β-sheet aggregates and amyloid fibrils.

Ex vivo activity of histone deacetylase inhibitors against multidrug-resistant clinical isolates of Plasmodium falciparum and P. vivax.

Histone acetylation plays an important role in regulating gene transcription and silencing in Plasmodium falciparum. Histone deacetylase (HDAC) inhibitors, particularly those of the hydroxamate class, have been shown to have potent in vitro activity against drug-resistant and -sensitive laboratory strains of P. falciparum, raising their potential as a new class of antimalarial compounds.

Profiling gene expression induced by protease-activated receptor 2 (PAR2) activation in human kidney cells.

Protease-Activated Receptor-2 (PAR2) has been implicated through genetic knockout mice with cytokine regulation and arthritis development. Many studies have associated PAR2 with inflammatory conditions (arthritis, airways inflammation, IBD) and key events in tumor progression (angiogenesis, metastasis), but they have relied heavily on the use of single agonists to identify physiological roles for PAR2. However such probes are now known not to be highly selective for PAR2, and thus precisely what PAR2 does and what mechanisms of downstream regulation are truly affected remain obscure.

Novel helix-constrained nociceptin derivatives are potent agonists and antagonists of ERK phosphorylation and thermal analgesia in mice.

The nociceptin opioid peptide receptor (NOP, NOR, ORL-1) is a GPCR that recognizes nociceptin, a 17-residue peptide hormone. Nociceptin regulates pain transmission, learning, memory, anxiety, locomotion, cardiovascular and respiratory stress, food intake, and immunity. Nociceptin was constrained using an optimized helix-inducing cyclization strategy to produce the most potent NOP agonist (EC50 = 40 pM) and antagonist (IC50 = 7.

Antimalarial histone deacetylase inhibitors containing cinnamate or NSAID components.

Malaria is the most lethal parasite-mediated tropical infectious disease, killing 1-2 million people each year. An emerging drug target is the enzyme Plasmodium falciparum histone deacetylase 1 (PfHDAC1). We report 26 compounds designed to bind the zinc and exterior surface around the entrance to the active site of PfHDAC1, 16 displaying potent in vitro antimalarial activity (IC(50)<100 nM) against P.

Inhibitors selective for HDAC6 in enzymes and cells.

Histone deacetylase inhibitors with anticancer or anti-inflammatory activity bind to Class I or Class I and II HDAC enzymes. Here we compare selectivity of inhibitors of a Class II HDAC enzyme (HDAC6) and find one that retains high selectivity in macrophages.

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.

Downsizing human, bacterial, and viral proteins to short water-stable alpha helices that maintain biological potency.

Recombinant proteins are important therapeutics due to potent, highly specific, and nontoxic actions in vivo. However, they are expensive medicines to manufacture, chemically unstable, and difficult to administer with low patient uptake and compliance. Small molecule drugs are cheaper and more bioavailable, but less target-specific in vivo and often have associated side effects.

Selective hexapeptide agonists and antagonists for human complement C3a receptor.

Human anaphylatoxin C3a, formed through cleavage of complement protein C3, is a potent effector of innate immunity via activation of its G protein coupled receptor, human C3aR. Previously reported short peptide ligands for this receptor either have low potency or lack receptor selectivity. Here we report the first small peptide agonists that are both potent and selective for human C3aR, derived from structure-activity relationships of peptides based on the C-terminus of C3a.

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.

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.

Noncovalent tripeptidyl benzyl- and cyclohexyl-amine inhibitors of the cysteine protease caspase-1.

Potent and noncovalent inhibitors of caspase-1 were produced by incorporating a secondary amine (reduced amide) isostere in place of the conventional electrophile (e.g., aldehyde) that normally confers high potency to cysteine protease inhibitors.

Inflammatory lipid mediators in adipocyte function and obesity.

Survival of multicellular organisms depends on their ability to fight infection, metabolize nutrients, and store energy for times of need. Unsurprisingly, therefore, immunoregulatory and metabolic mechanisms interact in human conditions such as obesity. Both infiltrating immunoinflammatory cells and adipocytes play critical roles in the modulation of metabolic homeostasis, so it is important to understand factors that regulate both adipocyte and immune cell function.

Left- and right-handed alpha-helical turns in homo- and hetero-chiral helical scaffolds.

Proteins typically consist of right-handed alpha helices, whereas left-handed alpha helices are rare in nature. Peptides of 20 amino acids or less corresponding to protein helices do not form thermodynamically stable alpha helices in water away from protein environments. The smallest known water-stable right- (alpha(R)) and left- (alpha(L)) handed alpha helices are reported, each stabilized in cyclic pentapeptide units containing all L- or all D-amino acids.

In silico screening of small molecule libraries using the dengue virus envelope E protein has identified compounds with antiviral activity against multiple flaviviruses.

The flaviviruses comprise a large group of related viruses, many of which pose a significant global human health threat, most notably the dengue viruses (DENV), West Nile virus (WNV) and yellow fever virus (YFV). Flaviviruses enter host cells via fusion of the viral and cellular membranes, a process mediated by the major viral envelope protein E as it undergoes a low pH induced conformational change in the endosomal compartment of the host cell. This essential entry stage in the flavivirus life cycle provides an attractive target for the development of antiviral agents.

Structure-activity relationships for substrate-based inhibitors of human complement factor B.

Human complement is a cascading network of plasma proteins important in immune defense, cooperatively effecting recognition, opsonization, destruction, and removal of pathogens and infected/damaged cells. Overstimulated or unregulated complement activation can result in immunoinflammatory diseases. Key serine proteases in this cascade are difficult to study due to their multiprotein composition, short lifetimes, formation on membranes, or serum circulation as inactive zymogens.

Complement component C2, inhibiting a latent serine protease in the classical pathway of complement activation.

The innate immune response to infection or injury involves an antigen-antibody triggered classical pathway (CP) of complement activation, in which soluble and cell surface plasma proteins cooperatively effect elimination of foreign organisms and damaged host cells. However, protracted or dysfunctional complement activation can lead to inflammatory diseases. Complement component 2 bound to C4b is cleaved by classical (C1s) or lectin (MASP2) proteases to produce C4bC2a, a very short-lived C3 convertase (t(1/2) 2 min) that in turn cleaves C3 to C3a and C3b, leading ultimately to formation of Membrane Attack Complex (MAC) and lysis of bacteria and damaged cells.

A cyclic beta-strand tripeptide with an alpha-helix like CD spectrum.

A protein alpha-helix is defined by 3.6 amino acids per turn. Cyclization of the tripeptide Alanine-Leucine-Glutamate through a side chain to the N-terminus lactam bond produces cyclo-(1,3)-[ALE]-NH(2) which displays a circular dichroism spectrum typical of an alpha-helix backbone.

The use of live-animal micro-computed tomography to determine the effect of a novel phospholipase A2 inhibitor on alveolar bone loss in an in vivo mouse model of periodontitis.

Background And Objective: Live-animal micro-computed tomography is a new and promising technique that can be used to quantify changes in bone volume for periodontal disease models. The major aim of this study was to develop the methodology of live-animal micro-computed tomography and to determine the effect of a novel secretory phospholipase A2 inhibitor on alveolar bone loss.

Material And Methods: Periodontitis was induced in mice by oral infection with Porphyromonas gingivalis over a period of 13 wk, and live-animal micro-computed tomography scans were taken at different time-points to determine bone volume changes with disease progression.

Histone deacetylase inhibitors in inflammatory disease.

Lysine acetylation is becoming increasingly appreciated as a key post-translational modification in the endogenous regulation of protein function. The so-called histone acetyl transferases (HATs) and histone deacetylases (HDACs), best known for their roles in controlling chromatin remodeling via histone acetylation/deacetylation, are now known to modify a large number of non-histone proteins to control diverse cell processes. In relation to inflammation, acetylation modulates the activity or function of cytokine receptors, nuclear hormone receptors, intracellular signaling molecules and transcription factors.

Targeting histone deacetylase inhibitors for anti-malarial therapy.

It is now clear that histone acetylation plays key roles in regulating gene transcription in both eukaryotes and prokaryotes, the acetylated form inducing gene expression while deacetylation silences genes. Recent studies have identified roles for histone acetyltransferases (HATs) and/or histone deacetylases (HDACs) in a number of parasites including Entamoeba histolytica, Toxoplasma gondii, Schistosoma mansoni, Cryptosporidium sp., Leishmania donovani, Neospora caninum, and Plasmodium falciparum.