Aspartic protease inhibitors. An integrated approach for the design andsynthesis of diaminodiol-based peptidomimetics.

Aspartic proteases play key roles in a variety of pathologies, including acquired immunodeficiency syndrome. Peptidomimetic inhibitors can act as drugs to combat these pathologies. We have developed an integrated methodology for preparing human immunodeficiency virus (HIV)-1 aspartic protease diaminodiol inhibitors, based on a computational method that predicts the potential inhibitory activity of the designed structures in terms of calculated enzyme-inhibitor complexation energies. This is combined with a versatile synthetic strategy that couples a high degree of stereochemical control in the central diaminodiol module with complete flexibility in the choice of side chains in the core and in flanking residues. A series of 23 tetrameric, pentameric and hexameric inhibitors, with a wide range of calculated relative complexation energies (-47.2 to +117 kJ.mol-1) and predicted hydrophobicities (logPo/w = 1.8-8.4) was thus assembled from readily available amino acids and carboxylic acids. The IC50 values for these compounds ranged from 3.2 nM to 90 microM, allowing study of correlations between structure and activity, and individuation of factors other than calculated complexation energies that determine the inhibition potency. Multivariable regression analysis revealed the importance of side-chain bulkiness and rigidity at the P2, P2' positions, suggesting possible improvements for the prediction process used to select candidate structures.     

Industrial-scale, genomics-based drug design and discovery.

The demands on drug discovery organizations have increased dramatically in recent years, partly because of the need to identify novel targets that are both relevant to disease and chemically tractable. This is leading to an industrial approach to traditional biology and chemistry, inspired in part by the revolution in genomics. The purpose of this article is to highlight the flow of investigation from gene sequence of potential therapeutic targets, through mRNA and protein expression, to protein structure and drug design. To deal with this scale of activity, many commercial and public organizations have been established and some of the key players will be listed in this article.     

Analogs design,synthesis and biological evaluation of peptidomimetics with potential anti-HCV activity

Two series of peptidomimetics were designed, prepared and evaluated for their anti-HCV activity. One series possesses a C-terminal carboxylate functionality. In the other series, the electrophilic vinyl sulfonate moiety was introduced as a novel class of HCV NS3/4A protease inhibitors. In vitro based studies were then performed to evaluate the efficacies of the inhibitors using Human hepatoma cells, with the vinyl sulfonate ester (10) in particular, found to have highly potent anti-HCV activity with an EC₅₀ = 0.296 μM. Finally, molecular modeling studies were performed through docking of the synthesized compounds in the HCV NS3/4A protease active site to assess their binding modes with the enzyme and gain further insight into their structure–activity relationships.     

Synthesis of bioactive and fluorescent pyridine-triazole-coumarin peptidomimetics through sequential click-multicomponent reactions

An easy synthetic protocol for the peptide randomization of 3,5-dicyano pyridine derivatives by linking the pyridine core with a coumarin chromophore spaced by a linker triazole via copper (I) catalyzed [3 + 2] azide-alkyne cycloaddition (CuAAC) is described. The new peptidomimetics thus obtained are extended rule of 5 (eRo5) molecules suitable for the development of therapeutic agents for undruggable targets. The structural and photophysical properties of the molecules are also promising for the development of potential bio imaging agents based on these molecules.     

The reactions of phenylglyoxal and related reagents with amino acids.

1. The reaction of phenylglyoxal (PGO), glyoxal (GO), and methylglyoxal (MGO) with amino acids were investigated at mild pH values at 25 degrees. These aldehydes reacted most rapidly with arginine and the rate of reaction increased with increasing pH values. Histidine, cystine, glycine, tryptophan, asparagine, glutamine, and lysine reacted with these aldehydes at significant but various rates, depending on the pH and the kind of the reagent used. The reactions with these amino acids seemed to involve both the alpha-amino groups and the side chain groups, and no significant reaction appeared to occur with the side chain alone except with those of arginine, lysine, and cysteine. These reagents were similarly reactive with the guanidinium group of arginine, but PGO appeared to be much less reactive with the epsilone-amino group of lysine than MGO and GO. The other ordinary amino acids were very much less reactive or did not react at all with these reagents, with the exception of cysteine. 2. Di-PGO-L-arginine was prepared from Nalpha-benzyloxycarbonyl-L-arginine, and di-PGO-methylguanidine from methylguanidine, and the stoichiometry of the reaction of two PGO molecules with one guanidino group was confirmed. A glyoxal derivative of L-arginine (GO-arginine) was prepared by reaction of glyoxal with arginine. GO-arginine was fairly unstable, especially at higher pH values. A similar derivative (MGO-arginine) was also found to be formed by reaction of MGO with L-arginine, and was similarly unstable. These derivatives, however, did not regenerate arginine upon acid hydrolysis.     

New fluorophore-forming reactions for histochemical visualization of N-acetylated and tertiary indolamines using glyoxylic acid,aluminum-formaldehyde and trifluoroacetic acid anhydride as reagents

Summary N-acetylated and tertiary indolamines, some of which are possible neurotransmitter candidates in the CNS, cannot be visualized with the standard Falck-Hillarp histofluorescence method and very little is known about their cellular localization. The present investigation demonstrates that glyoxylic acid (GA), formaldehyde (FA) in combination with aluminum ions (the ALFA method) and trifluoroacetic acid anhydride (TFAA) are capable of forming fluorescent compounds from N-acetylated (e.g. melatonin and N-acetyl-5-hydroxytryptamine) and tertiary (e.g. bufotenin) indolamines in histochemical protein models. With GA and FA-aluminum more vigorous reaction conditions were required for demonstration of these compounds compared to those needed for optimal visualization of primary catecholand indolamines (prolonged reaction time and higher concentration of GA and FA and aluminum ions). The fluorophore formation from N-acetylated and tertiary indolamines, which represents a new reaction principle in amine fluorescence histochemistry, is proposed to proceed as follows. In the first step, the indole reacts in 2-position with the reagent. The intermediate formed is dehydrated in the second step, yielding a strongly fluorescent 2-methylene derivative, which either per se or as the corresponding autoxidized dimer constitutes the main fluorophore. TFAA and related anhydrides represent new and potent reagents for histochemical visualization of N-acetylated indolamines such as melatonin. In contrast to the GA and ALFA reactions the optimal formation of fluorphores with TFAA required only mild reaction conditions (2–10 min at 0–20° C). The main fluorophore formed from melatonin has been identified and the reaction with TFAA is proposed to proceed as follows. An unstable intermediate, the isoimidinium carboxylate, is formed in the first step and this compound is then cyclized to form the fluorophore, 6-methoxy-1-methyl-3,4-dihydro--carboline. The GA and ALFA methods are already widely used for visualization of catecholamine systems. The fluorescence microscopical and microspectrofluorometric analysis did not, however, veveal any specific structures containing N-acetylated or tertiary indolamines in the rat CNS. The TFAA reaction was highly specific for N-acetylated indolamines when applied to protein models. However, in tissue a disturbing background fluorescence appeared, which under all reaction conditions tested, developed concomitantly with the specific fluorescence from melatonin. The problem with this background reaction has to be solved before the TFAA reaction can be applied for demonstration of N-acetylated indolamines in tissue.     

Recognizing and reporting adverse drug reactions.

Although physicians in practice are most likely to see patients with adverse drug reactions, they may fail to recognize an adverse effect or to attribute it to a drug effect and, when recognized, they may fail to report serious reactions to the US Food and Drug Administration (FDA). To recognize and attribute an adverse event to a drug effect, physicians should review the patient''s clinical course, looking at patient risk factors, the known adverse reactions to the suspected drug, and the likelihood of a causal relationship between the drug and the adverse event-based on the temporal relationship, response to stopping or restarting the drug, and whether other factors could explain the reaction. Once an adverse drug reaction has been identified, the patient should be informed and appropriate documentation made in the patient''s medical record. Serious known reactions and all reactions to newly released drugs or those not previously known to occur (even if the certainty is low) should be reported to the FDA.     

Modelling G-protein-coupled receptors for drug design.

The G-protein coupled receptors form a large and diverse multi-gene superfamily with many important physiological functions. As such, they have become important targets in pharmaceutical research. Molecular modelling and site-directed mutagenesis have played an important role in our increasing understanding of the structural basis of drug action at these receptors. Aspects of this understanding, how these techniques can be used within a drug-design programme, and remaining challenges for the future are reviewed.