Discovery of novel cathepsin S inhibitors by pharmacophore-based virtual high-throughput screening

The cysteine protease cathepsin S (CatS) is involved in the pathogenesis of autoimmune disorders, atherosclerosis, and obesity. Therefore, it represents a promising pharmacological target for drug development. We generated ligand-based and structure-based pharmacophore models for noncovalent and covalent CatS inhibitors to perform virtual high-throughput screening of chemical databases in order to discover novel scaffolds for CatS inhibitors. An in vitro evaluation of the resulting 15 structures revealed seven CatS inhibitors with kinetic constants in the low micromolar range. These compounds can be subjected to further chemical modifications to obtain drugs for the treatment of autoimmune disorders and atherosclerosis.

Power combining techniques into unbalanced loads for Class-E and inverse Class-E amplifiers

A power combining strategy for Class-E and inverse Class-E amplifiers operating at high frequencies such that they can operate into unbalanced loads is proposed. This power combining method is particularly important for the inverse Class-E amplifier configuration whose single-stage topology is naturally limited for small-to-medium power applications. Design examples for the power combining synthesis of classical Class-E and then inverse Class-E amplifiers with specification 3 V-1.5 W-2.5 GHz are given. For this specification, it is shown that a three-branch combiner has a natural 50 V output impedance. The resulting circuits are simulated within Agilent Advanced Design Systems environment with good agreement to theoretical prediction. Further the performance of the proposed circuits when operated in a Linear amplification using Nonlinear Components transmitter configuration whereby two-branch amplifiers are driven with constant amplitude conjugate input phase signals is investigated.

Series-L/parallel-tuned comparison with shunt-C/series-tuned class-E power amplifier

An analysis of the operation of a series-L/parallel-tuned class-E amplifier and its equivalence to the classic shunt-C/series-tuned class-E amplifier are presented. The first reported closed form design equations for the series-L/parallel-tuned topology operating under ideal switching conditions are given. Furthermore, a design procedure is introduced that allows the effect that nonzero switch resistance has on amplifier performance efficiency to be accounted for. The technique developed allows optimal circuit components to be found for a given device series resistance. For a relatively high value of switching device ON series resistance of 4O, drain efficiency of around 66% for the series-L/parallel-tuned topology, and 73% for the shunt-C/series-tuned topology appear to be the theoretical limits. At lower switching device series resistance levels, the efficiency performance of each type are similar, but the series-L/parallel-tuned topology offers some advantages in terms of its potential for MMIC realisation. Theoretical analysis is confirmed by numerical simulation for a 500mW (27dBm), 10% bandwidth, 5 V series-L/parallel-tuned, then, shunt-C/series-tuned class E power amplifier, operating at 2.5 GHz, and excellent agreement between theory and simulation results is achieved. The theoretical work presented in the paper should facilitate the design of high-efficiency switched amplifiers at frequencies commensurate with the needs of modern mobile wireless applications in the microwave frequency range, where intrinsically low-output-capacitance MMIC switching devices such as pHEMTs are to be used.

Novel genomic methods for drug discovery and mechanism-based toxicological assessment

Genomics encompasses a range of powerful technologies that can be applied at all levels of gene expression, from transcription to mRNA translation. Collectively, these technologies have great potential for improving drug discovery, both target and molecule recognition, and development. In this article we review the current and potential future status of established and novel genomic methods within drug discovery.

The masses and radii of HD 186753B and TYC7096-222-1B: the discovery of two M-dwarfs that eclipse A-type stars

We present observations of two new single-lined eclipsing binaries, both consisting of an Am star and an M-dwarf, discovered by the Wide Angle Search for Planets transit photometry survey. Using WASP photometry and spectroscopic measurements we find that HD 186753B has an orbital period of P=1.9194 days, a mass of M=0.24±0.02~M? and radius of R=0.31+0.06-0.06~R?; and that TCY7096-222-1B has an orbital period of P=8.9582 days, a mass of between 0.29 and 0.54 M? depending on eccentricity and radius of R=0.263+0.02-0.07~R?. We find that the Am stars have relatively low rotational velocities that closely match the orbital velocities of the M-dwarfs, suggesting that they have been “spun-down” by the M-dwarfs.

Discovery of multiple neuropeptide families in the phylum Platyhelminthes

Available evidence shows that short amidated neuropeptides are widespread and have important functions within the nervous systems of all flatworms (phylum Platyhelminthes) examined, and could therefore represent a starting point for new lead drug compounds with which to combat parasitic helminth infections. However, only a handful of these peptides have been characterised, the rigorous exploration of the flatworm peptide signalling repertoire having been hindered by the dearth of flatworm genomic data. Through searches of both expressed sequence tags and genomic resources using the basic local alignment search tool (BLAST), we describe 96 neuropeptides on 60 precursors from 10 flatworm species. Most of these (51 predicted peptides on 14 precursors) are novel and are apparently restricted to flatworms; the remainder comprise nine recognised peptide families including FMRFamide-like (FLPs), neuropeptide F (NPF)-like, myomodulin-like, buccalin-like and neuropeptide FF (NPFF)-like peptides; notably, the latter have only previously been reported in vertebrates. Selected peptides were localised immunocytochemically to the Schistosoma mansoni nervous system. We also describe several novel flatworm NPFs with structural features characteristic of the vertebrate neuropeptide Y (NPY) superfamily, previously unreported characteristics which support the common ancestry of flatworm NPFs with the NPY-superfamily. Our dataset provides a springboard for investigation of the functional biology and therapeutic potential of neuropeptides in flatworms, simultaneously launching flatworm neurobiology into the post-genomic era. (C) 2009 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

Peptide DV-28 amide: Prototype of a novel class of bradykinin receptor antagonist isolated from the defensive skin secretion of bombinid toads

Kinestatin, isolated from the skin of the Chinese toad, Bombina maxima, was the first bradykinin B2 receptor antagonist identified in amphibians. Molecular cloning established that it is co-encoded with the bradykinin-related peptide, maximakinin, within one of several skin kininogens. To examine other species within the genus Bombina for the presence of structural homologues of kinestatin, we subjected skin secretion of the toad, Bombina orientalis, to HPLC fractionation with subsequent bioassay of fractions for antagonism of bradykinin activity using an isolated rat tail artery smooth muscle preparation. A single fraction was located that inhibited bradykinin-induced relaxation of rat arterial smooth muscle and MALDI-TOF analysis of this fraction revealed that it contained a single peptide of molecular mass 3198.5 Da. Further primary structural analysis of this peptide showed that it was a 28-mer with an N-terminal Asp (D) residue and a C-terminal Val (V) residue that was amidated. The peptide was named DV-28 amide in accordance with these primary structural attributes. Synthetic DV-28 amide replicated the observed bradykinin antagonistic effect within the smooth muscle bioassay in a dose-dependent manner. In addition, it was observed to inhibit the proliferation of human microvessel endothelial cells (HMECs) as assessed by MTT assay. Bioinformatic analysis revealed that DV-28 amide was, like kinestatin, co-encoded with a bradykinin receptor agonist on one of two skin kininogens identified in B. orientalis. DV-28 amide thus represents a novel class of bradykinin antagonist from skin secretions of bombinid toads that appear to be a rich source of such novel peptides.

GK-22 amide: a novel myotropic peptide from the skin secretion of the bamboo leaf odorous frog, Odorrana versabilis

The skin secretions produced by many amphibians are formidable chemical/biological weapons deployed as a defence against predators. Bioactive peptides are often the predominant class of biochemical within these secretions and the inventory of such remains incomplete with each individual taxon producing unique cocktails contained within which are some signature peptides, such as bradykinins and tachykinins. These secretions have been the source of many peptides subsequently found to have structural homologues in vertebrate neuroendocrine systems (bombesin/GRP; sauvagine/CRF; caerulein/CCK) and vice versa (bradykinin, CGRP, NMU). They are thus unequivocally intriguing resources for novel bioactive peptide discovery. Here we describe a novel 22-mer amidated peptide, named GK-22 amide (N-terminal Gly (G) and C-terminal Lys (K) amide) with an internal disulphide bridge between Cys (C) 11 and 20 from the skin secretion of Odorrana versabilis. Molecular cloning indicated that it is encoded as a single copy on a biosynthetic precursor of 59 amino acid residues consisting of a signal peptide, an acidic amino acid residue-rich spacer domain and a mature peptide encoding domain flanked N-terminally by a classical -Lys-Arg- (KR) propeptide convertase processing site and C-terminally by a Gly (G) residue amide donor. A synthetic replicate of this peptide produced potent and dose-dependent contraction of the smooth muscle of rat urinary bladder. GK-22 amide thus represents the prototype of a novel class of myotropic peptide from amphibian skin and its discovery illustrates the continuing potential of this resource to this end.

Molecular cloning and deduced organization of the pHpG/CNP precursor from the venom of the African forest viper, Atheris squamigera

The discovery that the hypotensive sequela of envenomation by the South American viper, Bothrops jararaca, was mediated by peptides, represented a milestone in drug discovery research that led to the introduction of ACE inhibitors. These bradykinin-potentiating peptides (BPPs) have been found in the venoms of many species of viper and molecular cloning of biosynthetic precursors has revealed that each encodes several different BPPs in tandem with a single copy of a C-type natriuretic peptide (CNP) located at the C-terminus. Venoms of the African forest vipers (Atheris) have been poorly studied possibly because they do not represent a major danger to humans. However, initial studies have indicated that they contain some of the “classical” protein toxins of viper venoms and a novel class of peptide, the polyglycine/polyhistidine (pGpH) peptides. These peptides occur in several molecular forms with different numbers of repetitive glycine and histidine repeats. We have cloned the biosynthetic precursor of A. squamigera pGpH peptides from a venom-derived cDNA library and have confirmed that a single copy of CNP is located at the C-terminus and additionally that, like BPPs in other vipers, pGpH peptides are encoded in tandem within this single precursor. Solid phase peptide synthesis of pGpH peptides has proven to be extremely difficult but is progressing and acquisition of synthetic replicates of each peptide is a necessary prerequisite for systematic pharmacological characterisation as establishment of a biological function for these peptides remains elusive. pGpH peptides may prove to play a role as fundamental as that of the BPPs.

Helokinestatins: A novel family of bioactive peptides with drug-lead potential fromthe venomof the Gila Monster (Heloderma suspectum)

Venom of the Gila Monster (Heloderma suspectum) has proven to be an unlikely source of lead compounds (exendins) for the development of new injectable peptide therapeutics for the treatment of Type 2 diabetes. However, no systematic searches for new classes of bioactive peptides in lizard venom have appeared until recently. Here we describe the discovery of a new class of peptides – the helokinestatins – from H. suspectum venom, their structural characterisation and that of their biosynthetic precursors from cloned cDNA. In addition, we have subjected members of the family to preliminary pharmacological characterisation. Helokinestatins 1–6 are a family of proline-rich peptides containing 10–15 amino acid residues terminating in a common -Pro-Arg.OH motif. They are encoded in tandem within two virtually identical biosynthetic precursors of 177 and 178 amino acid residues, differing by only a single Pro residue. Each precursor also encodes a single copy of a C-type natriuretic peptide located at the C-terminus. Synthetic replicates of all helokinestatins were shown to be devoid of any direct action on the smooth muscle of rat tail artery but were found to be potent inhibitors of bradykinin-induced relaxation in this preparation in a manner that is suggestive of a non-competitive mechanism. Helokinestatin-3 (VPPPPLQMPLIPR) and helokinestatin-5 (VPPPLQMPLIPR) were found to be most potent in this respect causing almost complete inhibition of bradykinin-induced relaxation. Helokinestatins and BPPs may have a shared evolutionary history but the former do not inhibit ACE. The bradykinin inhibitory potential of helokinestatins may be exploited in the local control of chronic inflammation.

Digital Baseband Predistortion Based Linearized Broadband Inverse Class-E Power Amplifier

A newly introduced inverse class-E power amplifier (PA) was designed, simulated, fabricated, and characterized. The PA operated at 2.26 GHz and delivered 20.4-dBm output power with peak drain efficiency (DE) of 65% and power gain of 12 dB. Broadband performance was achieved across a 300-Mitz bandwidth with DE of better than 50% and 1-dB output-power flatness. The concept of enhanced injection predistortion with a capability to selectively suppress unwanted sub-frequency components and hence suitable for memory effects minimization is described coupled with a new technique that facilitates an accurate measurement of the phase of the third-order intermodulation (IM3) products. A robust iterative computational algorithm proposed in this paper dispenses with the need for manual tuning of amplitude and phase of the IM3 injected signals as commonly employed in the previous publications. The constructed inverse class-E PA was subjected to a nonconstant envelope 16 quadrature amplitude modulation signal and was linearized using combined lookup table (LUT) and enhanced injection technique from which superior properties from each technique can be simultaneously adopted. The proposed method resulted in 0.7% measured error vector magnitude (in rms) and 34-dB adjacent channel leakage power ratio improvement, which was 10 dB better than that achieved using the LUT predistortion alone.