Accessing active fragments for drug discovery utilising nitroreductase biocatalysis.

Biocatalysis has played a limited role in the early stages of drug discovery. This is often attributed to the limited substrate scope of enzymes not affording access to vast areas of novel chemical space. Here, we have shown a promiscuous nitroreductase enzyme (NR-55) can be used to produce a panel of functionalised anilines from a diverse panel of aryl nitro starting materials. After screening on analytical scale, we show that sixteen substrates could be scaled to 1 mmol scale, with several poly-functional anilines afforded with ease under the standard conditions. The aniline products were also screened for activity against several cell lines of interest, with modest activity observed for one compound. This study demonstrates the potential for nitroreductase biocatalysis to provide access to functional fragments under benign conditions.

Aberrant cognitive empathy in individuals with elevated social anxiety and regulation with emotional working memory training.

Social anxiety may disrupt the empathic process, and well-regulated empathy is critical for navigating the social world. Two studies aimed to further understand empathy in the context of social anxiety. Study 1 compared individuals with elevated or normative social anxiety on a measure assessing cognitive and affective empathy for positive and negative emotions conveyed by other people ("targets"), completed under social threat. Relative to individuals with normative social anxiety, individuals with elevated social anxiety had greater cognitive empathy and no differences in affective empathy, regardless of emotion type. As greater cognitive empathy can be maladaptive, Study 2 tested whether this could be down-regulated. Individuals with elevated social anxiety underwent emotional working memory training (eWMT) for negative emotional information, or control training (CT). Effects on an empathy measure completed under social threat were assessed. Cognitive empathy for negative emotions decreased following eWMT but not CT, and this was only evident for those with higher pre-training working memory capacity. Cognitive empathy for positive emotions and affective empathy were not affected. Overall, social anxiety is associated with aberrant elevated cognitive empathy for negative and positive emotions, and the deviation in cognitive empathy for negative emotions can be regulated with eWMT for certain individuals.Trial registration: Australian New Zealand Clinical Trials Registry identifier: ACTRN12618001196235..

Indolo[2,3-e]benzazocines and indolo[2,3-f]benzazonines and their copper(II) complexes as microtubule destabilizing agents.

A series of four indolo[2,3-e]benzazocines HL1-HL4 and two indolo[2,3-f]benzazonines HL5 and HL6, as well as their respective copper(II) complexes 1-6, were synthesized and characterized by 1H and 13C NMR spectroscopy, ESI mass spectrometry, single crystal X-ray diffraction (SC-XRD) and combustion analysis (C, H, N). SC-XRD studies of precursors Vd, VIa·0.5MeOH, of ligands HL4 and HL6·DCM, and complexes 2·2DMF, 5·2DMF, 5'·iPrOH·MeOH provided insights into the energetically favored conformations of eight- and nine-membered heterocycles in the four-ring systems. In addition, proton dissociation constants (pKa) of HL1, HL2 and HL5, complexes 1, 2 and 5, overall stability constants (log ß) of 1, 2 and 5 in 30% (v/v) DMSO/H2O at 298 K, as well as thermodynamic solubility of HL1-HL6 and 1-6 in aqueous solution at pH 7.4 were determined by UV-vis spectroscopy. All compounds were tested for antiproliferative activity against Colo320, Colo205 and MCF-7 cell lines and showed IC50 values in the low micromolar to sub-micromolar concentration range, while some of them (HL1, HL5 and HL6, 1, 2 and 6) showed remarkable selectivity towards malignant cell lines. Ethidium bromide displacement studies provided evidence that DNA is not the primary target for these drugs. Rather, inhibition of tubulin assembly is likely the underlying mechanism responsible for their antiproliferative activity. Tubulin disassembly experiments showed that HL1 and 1 are effective microtubule destabilizing agents binding to the colchicine site. This was also confirmed by molecular modelling investigations. To the best of our knowledge, complex 1 is the first reported transition metal complex to effectively bind to the tubulin-colchicine pocket.

Shifting as a key executive function underlying cognitive restructuring for individuals with elevated social anxiety.

OBJECTIVES: Previous studies have examined the relationship between executive functions and performance on cognitive behavioural therapy (CBT) tasks, such as cognitive restructuring. However, previous studies have used samples of older adults and only traditional measures of executive functions involving non-emotional stimuli. This study extends previous research to examine the specific executive function of shifting with regard to non-emotional and emotional stimuli and its relationship with cognitive restructuring, in a sample of young to middle-aged adults with elevated social anxiety. DESIGN: Cross-sectional study. METHODS: Participants (N = 49) completed a standard Wisconsin Card Sorting Test (WCST), an emotional version of the WCST (eWCST), and a cognitive restructuring task prior to an impromptu speech task. Per cent perseverative errors (an indicator of shifting) from the WCST and eWCST, along with planned covariates, were used to predict three indicators of cognitive restructuring task performance: task response quality (production of helpful alternative thoughts), change in belief in negative thought, and peak anxiety during speech. RESULTS: As expected, higher per cent perseverative errors (i.e., poorer shifting) on the WCST predicted poorer ability during the cognitive restructuring task to produce helpful alternative thoughts to a negative thought about the impending speech task. However, WCST per cent perseverative errors did not predict the other indicators of cognitive restructuring task performance. eWCST per cent perseverative errors did not predict any of the indicators of cognitive restructuring task performance. CONCLUSIONS: The standard WCST may be sensitive to capturing the type of mental flexibility which is important for producing helpful alternative thinking during cognitive restructuring. PRACTITIONER POINTS: Poorer shifting ability with regard to non-emotional stimuli in clients with elevated social anxiety may be related to poorer ability to produce helpful alternative thoughts during cognitive restructuring. For clinicians whose clients with elevated social anxiety are having difficulty with generating alternative thoughts during cognitive restructuring, clinicians should consider poor shifting ability as a potential contributing factor. Clinicians may need to provide further support for such clients during cognitive restructuring (e.g., greater emphasis on Socratic questioning to better facilitate alternative thinking).

Discovery and Optimization of DNA Gyrase and Topoisomerase IV Inhibitors with Potent Activity against Fluoroquinolone-Resistant Gram-Positive Bacteria.

Herein, we describe the discovery and optimization of a novel series that inhibits bacterial DNA gyrase and topoisomerase IV via binding to, and stabilization of, DNA cleavage complexes. Optimization of this series led to the identification of compound 25, which has potent activity against Gram-positive bacteria, a favorable in vitro safety profile, and excellent in vivo pharmacokinetic properties. Compound 25 was found to be efficacious against fluoroquinolone-sensitive Staphylococcus aureus infection in a mouse thigh model at lower doses than moxifloxacin. An X-ray crystal structure of the ternary complex formed by topoisomerase IV from Klebsiella pneumoniae, compound 25, and cleaved DNA indicates that this compound does not engage in a water-metal ion bridge interaction and forms no direct contacts with residues in the quinolone resistance determining region (QRDR). This suggests a structural basis for the reduced impact of QRDR mutations on antibacterial activity of 25 compared to fluoroquinolones.

Topoisomerase Inhibitors Addressing Fluoroquinolone Resistance in Gram-Negative Bacteria.

Since their discovery over 5 decades ago, quinolone antibiotics have found enormous success as broad spectrum agents that exert their activity through dual inhibition of bacterial DNA gyrase and topoisomerase IV. Increasing rates of resistance, driven largely by target-based mutations in the GyrA/ParC quinolone resistance determining region, have eroded the utility and threaten the future use of this vital class of antibiotics. Herein we describe the discovery and optimization of a series of 4-(aminomethyl)quinolin-2(1H)-ones, exemplified by 34, that inhibit bacterial DNA gyrase and topoisomerase IV and display potent activity against ciprofloxacin-resistant Gram-negative pathogens. X-ray crystallography reveals that 34 occupies the classical quinolone binding site in the topoisomerase IV-DNA cleavage complex but does not form significant contacts with residues in the quinolone resistance determining region.