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Chikungunya virus (CHIKV) is an Arbovirus that is transmitted to humans primarily by the mosquito species Aedes aegypti. Infection with this pathogen is often associated with fever, rash and arthralgia. Neither a vaccine nor an antiviral drug is available for the prevention or treatment of this disease. Albeit considered a tropical pathogen, adaptation of the virus to the mosquito species Aedes albopictus, which is also very common in temperate zones, has resulted in recent outbreaks in Europe and the US. In the present study, we report on the discovery of a novel series of compounds that inhibit CHIKV replication in the low μM range. In particular, we initially performed a virtual screening simulation of ∼5 million compounds on the CHIKV nsP2, the viral protease, after which we investigated and explored the Structure-Activity Relationships of the hit identified in silico. Overall, a series of 26 compounds, including the original hit, was evaluated in a virus-cell-based CPE reduction assay. The study of such selective inhibitors will contribute to a better understanding of the CHIKV replication cycle and may represents a first step towards the development of a clinical candidate drug for the treatment of this disease., (Copyright © 2013 Elsevier B.V. All rights reserved.)

We herein report a novel double pro-drug approach applied to the anti-HCV agent 2'-beta-C-methyl guanosine. A phosphoramidate ProTide motif and a 6-O-methoxy base pro-drug moiety are combined to generate lipophilic prodrugs of the monophosphate of the guanine nucleoside. Modification of the ester and amino acid moieties lead to a compound INX-08189 that exhibits 10nM potency in the HCV genotype 1b subgenomic replicon, thus being 500 times more potent than the parent nucleoside. The potency of the lead compound INX-08189 was shown to be consistent with intracellular 2'-C-methyl guanosine triphosphate levels in primary human hepatocytes. The separated diastereomers of INX-08189 were shown to have similar activity in the replicon assay and were also shown to be similar substrates for enzyme processing. INX-08189 has completed investigational new drug enabling studies and has been progressed into human clinical trials for the treatment of chronic HCV infection., (2010 Elsevier Ltd. All rights reserved.)

Computer-aided drug design has seen constantly increasing application over the past two decades in every area of drug discovery. It can offer significant advantages over conventional approaches, being far less expensive and faster than conventional methods, or offering the possibility to predict molecular behaviours that cannot be elucidated in any other way. Recent developments in software and hardware make it possible to simulate increasingly complex molecular environments, widening the applicability of in silico studies from the interactions of small molecules with key protein residues, to the simulation of the dynamic evolution of complex biological systems with atomic resolution. Antiviral research offers several open challenges, from a biological, biochemical and pharmaceutical point of view. Computational approaches are already providing some answers and will undoubtedly give more in the near future. Here, we present a brief overview of the cutting-edge computational methods that play a major role in present and future antiviral research.

The biological importance of microtubules in mitosis and cell division makes them an interesting target for the development of anticancer agents. Small molecules such as benzo[b]furans are attractive as inhibitors of tubulin polymerization. Thus, a new class of inhibitors of tubulin polymerization based on the 2-(3',4',5'-trimethoxybenzoyl)-benzo[b]furan molecular skeleton, with electron-donating (Me, OMe or OH) or electron-withdrawing (F, Cl and Br) substituents on the benzene ring, was synthesized and evaluated for antiproliferative activity, inhibition of tubulin polymerization and cell cycle effects. Adding a methyl group at the C-3 position resulted in increased activity. The most promising compound in this series was 2-(3',4',5'-trimethoxybenzoyl)-3-methyl-6-ethoxy-benzo[b]furan, which inhibits cancer cell growth at nanomolar concentrations and interacts strongly with tubulin by binding to the colchicine site.

Structure-based drug design is a creative process that displays several features that make it closer to human reasoning than to machine automation. However, very often the user intervention is limited to the preparation of the input and analysis of the output of a computer simulation. In some cases, allowing human intervention directly in the process could improve the quality of the results by applying the researcher intuition directly into the simulation. Haptic technology has been previously explored as a useful method to interact with a chemical system. However, the need of expensive hardware and the lack of accessible software have limited the use of this technology to date. Here we are reporting the implementation of a haptic-based molecular mechanics environment aimed for interactive drug design and ligand optimization, using an easily accessible software/hardware combination.

Chalcones are characterized by possessing an enone moiety between two aromatic rings. A series of chalcone-like agents, in which the double bond of the enone system is embedded within a thiophene ring, were synthesized and evaluated for antiproliferative activity and inhibition of tubulin assembly and colchicine binding to tubulin. The replacement of the double bond with a thiophene maintains antiproliferative activity and therefore must not significantly alter the relative conformation of the two aryl rings. The synthesized compounds were found to inhibit the growth of several cancer cell lines at nanomolar to low micromolar concentrations. In general, all compounds having significant antiproliferative activity inhibited tubulin polymerization with an IC(50)<2microM. Several of these compounds caused K562 cells to arrest in the G2/M phase of the cell cycle.

The 2-(3,4,5-trimethoxybenzoyl)-2-aminoindole nucleus was used as the fundamental structure for the synthesis of compounds modified with respect to positions C-4 to C-7 with different moieties (chloro, methyl, or methoxy). Additional structural variations concerned the indole nitrogen, which was alkylated with small alkyl groups such as methyl or ethyl. We have identified 1-methyl-2-(3,4,5-trimethoxybenzoyl)-3-amino-7-methoxyindole as a new highly potent antiproliferative agent that targets tubulin at the colchicine binding site and leads to apoptotic cell death.

Carbocyclic nucleoside analogues are catabolically stable since they are resistant to phosphorolytic cleavage by pyrimidine nucleoside phosphorylase enzymes. The carbocyclic analogue (C-BCNA) of the highly potent and selective anti-VZV bicyclic nucleoside analogue (BCNA) 6-pentylphenylfuro[2,3-d]pyrimidine-2'-deoxyribose was synthesized using carbocyclic 2'-deoxyuridine as starting material. C-BCNA was found to be chemically more stable than the furano lead, but it was shown to be significantly less antivirally active than its parent nucleoside analogue. It was noted to have a 10-fold lower inhibitory activity against the VZV-encoded thymidine kinase. This reduction of activity may be attributed to the different conformation of the sugar and base, as predicted by computational studies and supported by NMR studies. However, other factors besides affinity for VZV-TK must account for the greatly reduced antiviral potency.

We measured the levels of glutamine, aspartic acid, glutamic acid and GABA in cortex and hippocampus of mice acutely treated with i.p. human recombinant interleukin-2 (IL-2) or human recombinant interleukin-1 beta (IL-1 beta). Administration of IL-2 (5.0 micrograms kg-1) induced a slight but statistically significant increase in glutamine concentrations in both brain areas, while similar administration of IL-1 beta (20 micrograms kg-1) significantly reduced the hippocampal levels of glutamine, glutamic acid and GABA. Our data suggest that brain amino acid pathways are involved in the central modifications induced by IL-1 beta.

The influence of alprazolam, chlordiazepoxide and midazolam on the antinociceptive effect of subcutaneous morphine was investigated in rats, using the tail-flick test. After intraperitoneal administration, all drugs significantly enhanced the morphine-induced analgesia. Both the benzodiazepine receptor antagonist RO 15-3505 and the benzodiazepine receptor inverse agonist FG 7142 antagonized the potentiating effect of alprazolam, chlordiazepoxide and midazolam. Our results suggest that the interaction between benzodiazepines and opioids in modulating nociceptive responses involves primarily benzodiazepine receptors and that different pathways are involved in the anxiolytic and pro-analgesic actions of benzodiazepines.

Concentration of neurotransmitter amino acids (Tau, Gly, Asp, Glu, Gln, Ala, GABA) were measured in rat striatum following varying exposure (320 to 1408 h) to high intensity (50 Hz) electric fields. Tissue extracts in methanol, after drying, were derivatized with dansyl chloride and the amino acids quantitated by high-pressure liquid chromatography with ultraviolet detection. Short exposures (320 h) to 100 kV/m field induced a decrease in almost all tested amino acids. Longer exposure times (640 h) to 25 and 100 kV/m were only associated with a decrease in Tau. A further increase of the exposure time (1240 and 1408 h) both at 25 and 180 kV/m were mainly associated with a reduction of the amino acid levels. It is concluded that electric fields in the range 20-180 kV/m generate bimodal variations in neurotransmitter amino acids with troughs at very short and longer exposure times, independent from the field strength.

The effects of a series of adenosine derivatives on morphine antinoceptive effect were investigated in rats by the 'tail-flick' method. 2-Chloroadenosine (CADO) and L-N6-phenylisopropyladenosine (L-PIA), given intraperitoneally, caused decreased morphine antinociception. Intracerebroventricular injections of CADO, L-PIA and 5'-N-ethylcarboxamide adenosine (NECA), but not of 2'-deoxyadenosine, antagonized morphine antinociception. The effects of both central and peripheral injections of CADO and L-PIA on morphine antinociception were partially reversed by caffeine. Intracerebroventricular injection of dibutyryl-cyclic 3', 5' adenosine monophosphate (db cyclic AMP) had no effect on morphine antinociception. These data indicate that adenosine plays a role in morphine-induced antinociception. The results are discussed in terms of postulated effects of adenosine derivatives on adenylate cyclase.

1 Intracerebroventricular administration of diazepam or midazolam decreased the antinociceptive effect of morphine in rats as measured by the "tail flick" method. 2 Midazolam, injected into the periaqueductal grey matter (PAG) antagonized the analgesic effect of morphine. The action of midazolam was partially reversed by bicuculline. 3 These findings support the view that the effect of benzodiazephines on morphine antinociception may be mediated through gamma-aminobutyric acid receptors.

We describe a high-performance liquid chromatographic method, using electrochemical detection, for the determination of pirprofen in cerebrospinal fluid (CSF), plasma, and synovial fluid (SF). A C-18 column with a mobile phase containing acetonitrile acetate:phosphate buffer (pH 3.0) was employed. Samples were added with phosphoric acid, then extracted into dichloromethane, evaporated, and injected into the chromatograph. A detection potential of +0.85 V was applied on the basis of current-potential curves. Good linearity was found for each fluid in the expected range of therapeutic concentrations. The detection limit was 0.1 ng/mL for CSF, and 1 ng/mL for plasma and SF, with a recovery greater than 96% and intraday coefficient of variation less than 5% in all cases. The main advantages of this method include high specificity and sensitivity which allow the analysis of CSF and the use of small volumes of plasma and SF. The application of the method for the analysis of plasma and SF samples and the kinetic profile in CSF are shown.

The pharmacokinetic disposition of buflomedil was compared in humans after oral administration of solution, tablets and film tablets. Six healthy male volunteers received a single oral dose of the three different dosage formulations. Blood and urine samples were taken before dosing and at selected times over 24 h and 72 h, resp., after dosing. The concentration of the drug in samples was measured by gas-chromatography with nitrogen detector. The absorption of buflomedil was faster after solution administration, while other plasma parameters did not show any major differences. Also the amount of drug excreted in urines was higher with solution dosing.

1 Muscimol, a gamma-aminobutyric acid (GABA) receptor agonist, when injected intraventricularly antagonizes the antinociceptive effect of morphine given either subcutaneously or intraventricularly. The antagonistic effect of muscimol on morphine analgesia appears to be linearly related. 2 This finding provides support for the view that a GABA-ergic system is involved in morphine analgesia.