11 research outputs found
Structural Determinants of the Dictyostatin Chemotype for Tubulin Binding Affinity and Antitumor Activity Against Taxane- and Epothilone-Resistant Cancer Cells
A combined biochemical, structural, and cell biology characterization of dictyostatin is described, which enables an improved understanding of the structural determinants responsible for the high-affinity binding of this anticancer agent to the taxane site in microtubules (MTs). The study reveals that this macrolide is highly optimized for MT binding and that only a few of the structural modifications featured in a library of synthetic analogues resulted in small gains in binding affinity. The high efficiency of the dictyostatin chemotype in overcoming various kinds of clinically relevant resistance mechanisms highlights its potential for therapeutic development for the treatment of drug-resistant tumors. A structural explanation is advanced to account for the synergy observed between dictyostatin and taxanes on the basis of their differential effects on the MT lattice. The X-ray crystal structure of a tubulin−dictyostatin complex and additional molecular modeling have allowed the rationalization of the structure−activity relationships for a set of synthetic dictyostatin analogues, including the highly active hybrid 12 with discodermolide. Altogether, the work reported here is anticipated to facilitate the improved design and synthesis of more efficacious dictyostatin analogues and hybrids with other MT-stabilizing agents.We thank Peter T. Northcote for peloruside A, W.-S. Fang for Flutax-2, K. H. Altmann for epothilone D, Dr. Paraskevi Giannakakou (Weill Cornell Medical Center, New York) for the 1A9, PTX10, PTX22, and A8 cell lines, and Prof. Richard Ludueñ a (University of Texas) for the HeLa βIII-transfected cells. We thank Matadero INCOVA (Segovia) for the calf brains for tubulin purification. This work was supported in part by grants BIO2013-42984-R (J.F.D.) and SAF2012-39760-C02-02 (F.G.) from Ministerio de Economia y Competitividad, grant S2010/ ́ BMD-2457 BIPEDD2 from Comunidad Autonoma de Madrid ́ (F.G. and J.F.D.), and the Swiss National Science Foundation grants 310030B_138659 and 31003A_166608 (M.O.S.). The authors acknowledge networking contribution by the COST Action CM1407 “Challenging organic syntheses inspired by naturefrom natural products chemistry to drug discovery” and the COST action CM1470. I.P. thanks the EPSRC and AstraZeneca for funding, Dr. John Leonard (AstraZeneca) for useful discussions, Dr. Stuart Mickel (Novartis) for the provision of chemicals, and the EPSRC UK National Mass Spectrometry Facility at Swansea University for mass spectra
Structural Determinants of the Dictyostatin Chemotype for Tubulin Binding Affinity and Antitumor Activity Against Taxane- and Epothilone-Resistant Cancer Cells
A combined biochemical, structural, and cell biology characterization of dictyostatin is described, which enables an improved understanding of the structural determinants responsible for the high-affinity binding of this anticancer agent to the taxane site in microtubules (MTs). The study reveals that this macrolide is highly optimized for MT binding and that only a few of the structural modifications featured in a library of synthetic analogues resulted in small gains in binding affinity. The high efficiency of the dictyostatin chemotype in overcoming various kinds of clinically relevant resistance mechanisms highlights its potential for therapeutic development for the treatment of drug-resistant tumors. A structural explanation is advanced to account for the synergy observed between dictyostatin and taxanes on the basis of their differential effects on the MT lattice. The X-ray crystal structure of a tubulin−dictyostatin complex and additional molecular modeling have allowed the rationalization of the structure−activity relationships for a set of synthetic dictyostatin analogues, including the highly active hybrid 12 with discodermolide. Altogether, the work reported here is anticipated to facilitate the improved design and synthesis of more efficacious dictyostatin analogues and hybrids with other MT-stabilizing agents.We thank Peter T. Northcote for peloruside A, W.-S. Fang for Flutax-2, K. H. Altmann for epothilone D, Dr. Paraskevi Giannakakou (Weill Cornell Medical Center, New York) for the 1A9, PTX10, PTX22, and A8 cell lines, and Prof. Richard Ludueñ a (University of Texas) for the HeLa βIII-transfected cells. We thank Matadero INCOVA (Segovia) for the calf brains for tubulin purification. This work was supported in part by grants BIO2013-42984-R (J.F.D.) and SAF2012-39760-C02-02 (F.G.) from Ministerio de Economia y Competitividad, grant S2010/ ́ BMD-2457 BIPEDD2 from Comunidad Autonoma de Madrid ́ (F.G. and J.F.D.), and the Swiss National Science Foundation grants 310030B_138659 and 31003A_166608 (M.O.S.). The authors acknowledge networking contribution by the COST Action CM1407 “Challenging organic syntheses inspired by naturefrom natural products chemistry to drug discovery” and the COST action CM1470. I.P. thanks the EPSRC and AstraZeneca for funding, Dr. John Leonard (AstraZeneca) for useful discussions, Dr. Stuart Mickel (Novartis) for the provision of chemicals, and the EPSRC UK National Mass Spectrometry Facility at Swansea University for mass spectra
Intra annual seed production and availability of two morphotypes of Brosimum rubescens taubert in forests of the Colombian Amazon
Evaluation of appendicitis risk prediction models in adults with suspected appendicitis
Background
Appendicitis is the most common general surgical emergency worldwide, but its diagnosis remains challenging. The aim of this study was to determine whether existing risk prediction models can reliably identify patients presenting to hospital in the UK with acute right iliac fossa (RIF) pain who are at low risk of appendicitis.
Methods
A systematic search was completed to identify all existing appendicitis risk prediction models. Models were validated using UK data from an international prospective cohort study that captured consecutive patients aged 16–45 years presenting to hospital with acute RIF in March to June 2017. The main outcome was best achievable model specificity (proportion of patients who did not have appendicitis correctly classified as low risk) whilst maintaining a failure rate below 5 per cent (proportion of patients identified as low risk who actually had appendicitis).
Results
Some 5345 patients across 154 UK hospitals were identified, of which two‐thirds (3613 of 5345, 67·6 per cent) were women. Women were more than twice as likely to undergo surgery with removal of a histologically normal appendix (272 of 964, 28·2 per cent) than men (120 of 993, 12·1 per cent) (relative risk 2·33, 95 per cent c.i. 1·92 to 2·84; P < 0·001). Of 15 validated risk prediction models, the Adult Appendicitis Score performed best (cut‐off score 8 or less, specificity 63·1 per cent, failure rate 3·7 per cent). The Appendicitis Inflammatory Response Score performed best for men (cut‐off score 2 or less, specificity 24·7 per cent, failure rate 2·4 per cent).
Conclusion
Women in the UK had a disproportionate risk of admission without surgical intervention and had high rates of normal appendicectomy. Risk prediction models to support shared decision‐making by identifying adults in the UK at low risk of appendicitis were identified
2'-Deoxyribosyltransferase from Leishmania mexicana, an efficient biocatalyst for one-pot, one-step synthesis of nucleosides from poorly soluble purine bases
Processes catalyzed by enzymes offer numerous advantages over chemical methods although in many occasions the stability of the biocatalysts becomes a serious concern. Traditionally, synthesis of nucleosides using poorly water-soluble purine bases, such as guanine, xanthine, or hypoxanthine, requires alkaline pH and/or high temperatures in order to solubilize the substrate. In this work, we demonstrate that the 2′-deoxyribosyltransferase from Leishmania mexicana (LmPDT) exhibits an unusually high activity and stability under alkaline conditions (pH 8–10) across a broad range of temperatures (30–70 °C) and ionic strengths (0–500 mM NaCl). Conversely, analysis of the crystal structure of LmPDT together with comparisons with hexameric, bacterial homologues revealed the importance of the relationships between the oligomeric state and the active site architecture within this family of enzymes. Moreover, molecular dynamics and docking approaches provided structural insights into the substrate-binding mode. Biochemical characterization of LmPDT identifies the enzyme as a type I NDT (PDT), exhibiting excellent activity, with specific activity values 100- and 4000-fold higher than the ones reported for other PDTs. Interestingly, LmPDT remained stable during 36 h at different pH values at 40 °C. In order to explore the potential of LmPDT as an industrial biocatalyst, enzymatic production of several natural and non-natural therapeutic nucleosides, such as vidarabine (ara A), didanosine (ddI), ddG, or 2′-fluoro-2′-deoxyguanosine, was carried out using poorly water-soluble purines. Noteworthy, this is the first time that the enzymatic synthesis of 2′-fluoro-2′-deoxyguanosine, ara G, and ara H by a 2′-deoxyribosyltransferase is reported
2'-Deoxyribosyltransferase from Leishmania mexicana, an efficient biocatalyst for one-pot, one-step synthesis of nucleosides from poorly soluble purine bases.Fundación Santander, Universidad Europea3.340 JCR (2017) Q2, 46/160 Biotechnology and Applied MicrobiologyUE