Uncovering Biosynthetic Relationships Between Antifungal Nonadrides and Octadrides

Maleidrides are a class of bioactive secondary metabolites unique to filamentous fungi, which contain one or more maleic anhydrides fused to a 7-, 8- or 9- membered carbocycle (named heptadrides, octadrides and nonadrides respectively). Herein structural and biosynthetic studies on the antifungal octadride, zopfiellin, and nonadrides scytalidin, deoxyscytalidin and castaneiolide are described. A combination of genome sequencing, bioinformatic analyses, gene disruptions, biotransformations, isotopic feeding studies, NMR and X-ray crystallography revealed that they share a common biosynthetic pathway, diverging only after the nonadride deoxyscytalidin. 5-Hydroxylation of deoxyscytalidin occurs prior to ring contraction in the zopfiellin pathway of Diffractella curvata. In Scytalidium album, 6-hydroxylation-confirmed as being catalysed by the α-ketoglutarate dependent oxidoreductase ScyL2-converts deoxyscytalidin to scytalidin, in the final step in the scytalidin pathway. Feeding scytalidin to a zopfiellin PKS knockout strain led to the production of the nonadride castaneiolide and two novel ring-open maleidrides. © The Royal Society of Chemistry

Novel nonadride, heptadride and maleic acid metabolites from the byssochlamic acid producer Byssochlamys fulva IMI 40021-an insight into the biosynthesis of maleidrides

The filamentous fungus Byssochlamys fulva strain IMI 40021 produces (+)-byssochlamic acid 1, its novel dihydroanalogue 2 and four related secondary metabolites. Agnestadrides A, 17 and B, 18 constitute a novel class of seven-membered ring, maleic anhydride-containing (hence termed heptadride) natural products. The putative maleic anhydride precursor 5 for both nonadride and heptadride biosynthesis was isolated as a fermentation product for the first time and its structure confirmed by synthesis. Acid 5 undergoes facile decarboxylation to anhydride 6. The generic term maleidrides is proposed to encompass biosynthetically-related compounds containing maleic anhydride moieties fused to an alicyclic ring, varying in size and substituents

Inside out: Fusing 3D imaging modalities for the internal and external investigation of multi-material museum objects

3D imaging methods are increasingly employed in cultural heritage research to analyse and document objects in museum collections. In this work, we provide an interactive visualisation plugin for the open-source software Blender, to combine and inspect two complementary 3D imaging modalities: CT images, which capture the interior; and surface scans, which capture the exterior. 3D CT scan data can be visualised, both as volumetric representation and as orthogonal slices, and a 3D surface scan can be registered onto the CT data. It allows users to simultaneously and interactively inspect these modalities and to virtually cut through an object. It also provides tools for generating output images and videos for research and public outreach purposes. The plugin workflow was applied to four case studies from the collections of the Rijksmuseum, Amsterdam, and the British Museum, London. The plugin is published open-source together with detailed guidelines and a practice dataset

Adding 6 months of androgen deprivation therapy to postoperative radiotherapy for prostate cancer: a comparison of short-course versus no androgen deprivation therapy in the RADICALS-HD randomised controlled trial

Background Previous evidence indicates that adjuvant, short-course androgen deprivation therapy (ADT) improves metastasis-free survival when given with primary radiotherapy for intermediate-risk and high-risk localised prostate cancer. However, the value of ADT with postoperative radiotherapy after radical prostatectomy is unclear. Methods RADICALS-HD was an international randomised controlled trial to test the efficacy of ADT used in combination with postoperative radiotherapy for prostate cancer. Key eligibility criteria were indication for radiotherapy after radical prostatectomy for prostate cancer, prostate-specific antigen less than 5 ng/mL, absence of metastatic disease, and written consent. Participants were randomly assigned (1:1) to radiotherapy alone (no ADT) or radiotherapy with 6 months of ADT (short-course ADT), using monthly subcutaneous gonadotropin-releasing hormone analogue injections, daily oral bicalutamide monotherapy 150 mg, or monthly subcutaneous degarelix. Randomisation was done centrally through minimisation with a random element, stratified by Gleason score, positive margins, radiotherapy timing, planned radiotherapy schedule, and planned type of ADT, in a computerised system. The allocated treatment was not masked. The primary outcome measure was metastasis-free survival, defined as distant metastasis arising from prostate cancer or death from any cause. Standard survival analysis methods were used, accounting for randomisation stratification factors. The trial had 80% power with two-sided α of 5% to detect an absolute increase in 10-year metastasis-free survival from 80% to 86% (hazard ratio [HR] 0·67). Analyses followed the intention-to-treat principle. The trial is registered with the ISRCTN registry, ISRCTN40814031, and ClinicalTrials.gov, NCT00541047. Findings Between Nov 22, 2007, and June 29, 2015, 1480 patients (median age 66 years [IQR 61–69]) were randomly assigned to receive no ADT (n=737) or short-course ADT (n=743) in addition to postoperative radiotherapy at 121 centres in Canada, Denmark, Ireland, and the UK. With a median follow-up of 9·0 years (IQR 7·1–10·1), metastasis-free survival events were reported for 268 participants (142 in the no ADT group and 126 in the short-course ADT group; HR 0·886 [95% CI 0·688–1·140], p=0·35). 10-year metastasis-free survival was 79·2% (95% CI 75·4–82·5) in the no ADT group and 80·4% (76·6–83·6) in the short-course ADT group. Toxicity of grade 3 or higher was reported for 121 (17%) of 737 participants in the no ADT group and 100 (14%) of 743 in the short-course ADT group (p=0·15), with no treatment-related deaths. Interpretation Metastatic disease is uncommon following postoperative bed radiotherapy after radical prostatectomy. Adding 6 months of ADT to this radiotherapy did not improve metastasis-free survival compared with no ADT. These findings do not support the use of short-course ADT with postoperative radiotherapy in this patient population

Duration of androgen deprivation therapy with postoperative radiotherapy for prostate cancer: a comparison of long-course versus short-course androgen deprivation therapy in the RADICALS-HD randomised trial

Background Previous evidence supports androgen deprivation therapy (ADT) with primary radiotherapy as initial treatment for intermediate-risk and high-risk localised prostate cancer. However, the use and optimal duration of ADT with postoperative radiotherapy after radical prostatectomy remains uncertain. Methods RADICALS-HD was a randomised controlled trial of ADT duration within the RADICALS protocol. Here, we report on the comparison of short-course versus long-course ADT. Key eligibility criteria were indication for radiotherapy after previous radical prostatectomy for prostate cancer, prostate-specific antigen less than 5 ng/mL, absence of metastatic disease, and written consent. Participants were randomly assigned (1:1) to add 6 months of ADT (short-course ADT) or 24 months of ADT (long-course ADT) to radiotherapy, using subcutaneous gonadotrophin-releasing hormone analogue (monthly in the short-course ADT group and 3-monthly in the long-course ADT group), daily oral bicalutamide monotherapy 150 mg, or monthly subcutaneous degarelix. Randomisation was done centrally through minimisation with a random element, stratified by Gleason score, positive margins, radiotherapy timing, planned radiotherapy schedule, and planned type of ADT, in a computerised system. The allocated treatment was not masked. The primary outcome measure was metastasis-free survival, defined as metastasis arising from prostate cancer or death from any cause. The comparison had more than 80% power with two-sided α of 5% to detect an absolute increase in 10-year metastasis-free survival from 75% to 81% (hazard ratio [HR] 0·72). Standard time-to-event analyses were used. Analyses followed intention-to-treat principle. The trial is registered with the ISRCTN registry, ISRCTN40814031, and ClinicalTrials.gov , NCT00541047 . Findings Between Jan 30, 2008, and July 7, 2015, 1523 patients (median age 65 years, IQR 60–69) were randomly assigned to receive short-course ADT (n=761) or long-course ADT (n=762) in addition to postoperative radiotherapy at 138 centres in Canada, Denmark, Ireland, and the UK. With a median follow-up of 8·9 years (7·0–10·0), 313 metastasis-free survival events were reported overall (174 in the short-course ADT group and 139 in the long-course ADT group; HR 0·773 [95% CI 0·612–0·975]; p=0·029). 10-year metastasis-free survival was 71·9% (95% CI 67·6–75·7) in the short-course ADT group and 78·1% (74·2–81·5) in the long-course ADT group. Toxicity of grade 3 or higher was reported for 105 (14%) of 753 participants in the short-course ADT group and 142 (19%) of 757 participants in the long-course ADT group (p=0·025), with no treatment-related deaths. Interpretation Compared with adding 6 months of ADT, adding 24 months of ADT improved metastasis-free survival in people receiving postoperative radiotherapy. For individuals who can accept the additional duration of adverse effects, long-course ADT should be offered with postoperative radiotherapy. Funding Cancer Research UK, UK Research and Innovation (formerly Medical Research Council), and Canadian Cancer Society

Quantitative energy-dispersive x-ray diffraction for identification of counterfeit medicines: a preliminary study

The prevalence of counterfeit and substandard medicines has been growing rapidly over the past decade, and fast, nondestructive techniques for their detection are urgently needed to counter this trend. In this study, energy-dispersive X-ray diffraction (EDXRD) combined with chemometrics was assessed for its effectiveness in quantitative analysis of compressed powder mixtures. Although EDXRD produces lower-resolution diffraction patterns than angular-dispersive X-ray diffraction (ADXRD), it is of interest for this application as it carries the advantage of allowing the analysis of tablets within their packaging, due to the higher energy X-rays used. A series of caffeine, paracetamol and microcrystalline cellulose mixtures were prepared with compositions between 0 - 100 weight% in 20 weight% steps (22 samples in total, including a centroid mixture), and were pressed into tablets. EDXRD spectra were collected in triplicate, and a principal component analysis (PCA) separated these into their correct positions in the ternary mixture design. A partial least-squares (PLS) regression model calibrated using this training set was validated using both segmented cross-validation, and with a test set of six samples (mixtures in 8:1:1 and 5⅓:2⅓:2⅓ ratios) – the latter giving a root-mean square error of prediction (RMSEP) of 1.30, 2.25 and 2.03 weight% for caffeine, paracetamol and cellulose respectively. These initial results are promising, with RMSEP values on a par with those reported in the ADXRD literature

Lead identification of benzimidazolone and azabenzimidazolone arylsulfonamides as CC-chemokine receptor 4 (CCR4) antagonists

A knowledge-based library of 2,3-dichlorophenylsulfonyl derivatives of commercially available aryl amines was synthesised and screened as human CCR4 antagonists, in order to identify a suitable hit for the start of a lead-optimisation programme. Hits were required to be more potent than an existing indazole series, have better physicochemical properties (c log P 116 μg/mL), and be stable to acid and light. The benzimidazol-2-one core was identified as a hit suitable for further investigation. Substitution at N1 with small alkyl groups was tolerated; however, these analogues were inactive in the whole blood assay (pA2 <5). Azabenzimidazolone analogues were all found to be active, with compound 38 exhibiting whole blood activity of 6.1, low molecular weight (389) and chrom log D7.4 (2.4), high LE (0.43), and solubility (152 μg/mL). In addition, 38 had human serum albumin binding of around 93% and met all the criteria for progression to lead optimisation

Uncovering biosynthetic relationships between antifungal nonadrides and octadrides

Maleidrides are a class of bioactive secondary metabolites unique to filamentous fungi, which contain one or more maleic anhydrides fused to a 7-, 8- or 9- membered carbocycle (named heptadrides, octadrides and nonadrides respectively). Herein structural and biosynthetic studies on the antifungal octadride, zopfiellin, and nonadrides scytalidin, deoxyscytalidin and castaneiolide are described. A combination of genome sequencing, bioinformatic analyses, gene disruptions, biotransformations, isotopic feeding studies, NMR and X-ray crystallography revealed that they share a common biosynthetic pathway, diverging only after the nonadride deoxyscytalidin. 5-Hydroxylation of deoxyscytalidin occurs prior to ring contraction in the zopfiellin pathway of Diffractella curvata. In Scytalidium album, 6-hydroxylation-confirmed as being catalysed by the α-ketoglutarate dependent oxidoreductase ScyL2-converts deoxyscytalidin to scytalidin, in the final step in the scytalidin pathway. Feeding scytalidin to a zopfiellin PKS knockout strain led to the production of the nonadride castaneiolide and two novel ring-open maleidrides. © The Royal Society of Chemistry

Enabling 3D CT-scanning of cultural heritage objects using only in-house 2D X-ray equipment in museums

Visualizing the internal structure of museum objects is a crucial step in acquiring knowledge about the origin, state, and composition of cultural heritage artifacts. Among the most powerful techniques for exposing the interior of museum objects is computed tomography (CT), a technique that computationally forms a 3D image using hundreds of radiographs acquired in a full circular range. However, the lack of affordable and versatile CT equipment in museums, combined with the challenge of transporting precious collection objects, currently keeps this technique out of reach for most cultural heritage applications. We propose an approach for creating accurate CT reconstructions using only standard 2D radiography equipment already available in most larger museums. Specifically, we demonstrate that a combination of basic X-ray imaging equipment, a tailored marker-based image acquisition protocol, and sophisticated data-processing algorithms, can achieve 3D imaging of collection objects without the need for a costly CT imaging system. We implemented this approach in the British Museum (London), the J. Paul Getty Museum (Los Angeles), and the Rijksmuseum (Amsterdam). Our work paves the way for broad facilitation and adoption of CT technology across museums worldwide