Structure–activity relationship of ipglycermide binding to phosphoglycerate mutases

Catalysis of human phosphoglycerate mutase is dependent on a 2,3-bisphosphoglycerate cofactor (dPGM), whereas the nonhomologous isozyme in many parasitic species is cofactor independent (iPGM). This mechanistic and phylogenetic diversity offers an opportunity for selective pharmacologic targeting of glycolysis in disease-causing organisms. We previously discovered ipglycermide, a potent inhibitor of iPGM, from a large combinatorial cyclic peptide library. To fully delineate the ipglycermide pharmacophore, herein we construct a detailed structure–activity relationship using 280 substituted ipglycermide analogs. Binding affinities of these analogs to immobilized Caenorhabditis elegans iPGM, measured as fold enrichment relative to the index residue by deep sequencing of an mRNA display library, illuminated the significance of each amino acid to the pharmacophore. Using cocrystal structures and binding kinetics, we show that the high affinity of ipglycermide for iPGM orthologs, from Brugia malayi, Onchocerca volvulus, Dirofilaria immitis, and Escherichia coli, is achieved by a codependence between (1) the off-rate mediated by the macrocycle Cys14 thiolate coordination to an active-site Zn2+ in the iPGM phosphatase domain and (2) shape complementarity surrounding the macrocyclic core at the phosphotransferase–phosphatase domain interface. Our results show that the high-affinity binding of ipglycermide to iPGMs freezes these structurally dynamic enzymes into an inactive, stable complex

A target-agnostic screen identifies approved drugs to stabilize the endoplasmic reticulum-resident proteome

Endoplasmic reticulum (ER) dysregulation is associated with pathologies including neurodegenerative, muscular, and diabetic conditions. Depletion of ER calcium can lead to the loss of resident proteins in a process termed exodosis. To identify compounds that attenuate the redistribution of ER proteins under pathological conditions, we performed a quantitative high-throughput screen using the Gaussia luciferase (GLuc)-secreted ER calcium modulated protein (SERCaMP) assay, which monitors secretion of ER-resident proteins triggered by calcium depletion. We identify several clinically used drugs, including bromocriptine, and further characterize them using assays to measure effects on ER calcium, ER stress, and ER exodosis. Bromocriptine elicits protective effects in cell-based models of exodosis as well as in vivo models of stroke and diabetes. Bromocriptine analogs with reduced dopamine receptor activity retain similar efficacy in stabilizing the ER proteome, indicating a non-canonical mechanism of action. This study describes a strategic approach to identify small-molecule drugs capable of improving ER proteostasis in human disease conditions.Peer reviewe

Kinetic and Structural Investigations of Novel Inhibitors of Human Epithelial 15-Lipoxygenase-2

Human epithelial 15-lipoxygenase-2 (h15-LOX-2, ALOX15B) is expressed in many tissues and has been implicated in atherosclerosis, cystic fibrosis and ferroptosis. However, there are few reported potent/selective inhibitors that are active ex vivo. In the current work, we report newly discovered molecules that are more potent and structurally distinct from our previous inhibitors, MLS000545091 and MLS000536924 (Jameson et al, PLoS One, 2014, 9, e104094), in that they contain a central imidazole ring, which is substituted at the 1-position with a phenyl moiety and with a benzylthio moiety at the 2-position. The initial three molecules were mixed-type, non-reductive inhibitors, with IC(50) values of 0.34 ± 0.05 μM for MLS000327069, 0.53 ± 0.04 μM for MLS000327186 and 0.87 ± 0.06 μM for MLS000327206 and greater than 50-fold selectivity versus h5-LOX, h12-LOX, h15-LOX-1, COX-1 and COX-2. A small set of focused analogs was synthesized to demonstrate the validity of the hits. In addition, a binding model was developed for the three imidazole inhibitors based on computational docking and a co-structure of h15-LOX-2 with MLS000536924. Hydrogen/deuterium exchange (HDX) results indicate a similar binding mode between MLS000536924 and MLS000327069, however, the latter restricts protein motion of helix-α2 more, consistent with its greater potency. Given these results, we designed, docked, and synthesized novel inhibitors of the imidazole scaffold and confirmed our binding mode hypothesis. Importantly, four of the five inhibitors mentioned above are active in an h15-LOX-2/HEK293 cell assay and thus they could be important tool compounds in gaining a better understanding of h15-LOX-2’s role in human biology. As such, a suite of similar pharmacophores that target h15-LOX-2 both in vitro and ex vivo are presented in the hope of developing them as therapeutic agents

Quantitative High-Throughput Screening Identifies 8-Hydroxyquinolines as Cell-Active Histone Demethylase Inhibitors

Small molecule modulators of epigenetic processes are currently sought as basic probes for biochemical mechanisms, and as starting points for development of therapeutic agents. N(epsilon)-Methylation of lysine residues on histone tails is one of a number of post-translational modifications that together enable transcriptional regulation. Histone lysine demethylases antagonize the action of histone methyltransferases in a site- and methylation state-specific manner. N(epsilon)-Methyllysine demethylases that use 2-oxoglutarate as co-factor are associated with diverse human diseases, including cancer, inflammation and X-linked mental retardation; they are proposed as targets for the therapeutic modulation of transcription. There are few reports on the identification of templates that are amenable to development as potent inhibitors in vivo and large diverse collections have yet to be exploited for the discovery of demethylase inhibitors

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Environmentally benign reaction: Synthesis of sydnone chaIcones under solvent-free conditions

2556-2557An easy, simple and eco-friendly synthesis of 4-(1'-oxo-3'-aryl-2'-propenyl)-3-arylsydnones is reported by grinding 4-acetyl-3-arylsydnones with various aryl aldehydes in a mortar with pestle

One pot reaction: Synthesis, characterization and biological activity of 3-alkyl/aryl-9- substituted 1,2,4-triazolo[3,4-<i>b</i>] [1,3,4]quinolinothiadiazepines

211-214Reaction of 6-substituted-2-chloro-3-formylquinoline 1 and 3-substituted-4-amino-5-mercapto-1,2,4-triazole 2 gave the novel thiadiazepine derivatives 5 rather than expected Schiff bases 4. Alternatively, compounds 5 were also prepared by the reaction of 2 with 6-substitutcri quinolones 3. The structures of the newly synthesized compounds have been proposed on the basis of elemental analysis, IR,1H NMR and mass spectral data. Some of the new synthetic compounds were also screened for their antibacterial and antifungal activity. Most of them showed significant activity

<span style="font-size:14.0pt;font-family:"Times New Roman";color:#1A1A1A;mso-bidi-font-weight: bold" lang="EN-IN">Reaction of anilinoacet hydrazides with <span style="font-size:14.0pt;font-family:"Times New Roman";mso-fareast-font-family: HiddenHorzOCR;color:#1A1A1A;mso-bidi-font-weight:bold" lang="EN-IN">α<span style="font-size:14.0pt;font-family:HiddenHorzOCR;mso-hansi-font-family: "Times New Roman";mso-bidi-font-family:HiddenHorzOCR;color:#1A1A1A;mso-bidi-font-weight: bold" lang="EN-IN">,<span style="font-size:14.0pt;font-family:"Times New Roman"; mso-fareast-font-family:HiddenHorzOCR;color:#1A1A1A;mso-bidi-font-weight:bold" lang="EN-IN">β<span style="font-size:14.0pt;font-family:HiddenHorzOCR;mso-hansi-font-family: "Times New Roman";mso-bidi-font-family:HiddenHorzOCR;color:#1A1A1A;mso-bidi-font-weight: bold" lang="EN-IN">-acetylenic <span style="font-size:14.0pt;font-family: "Times New Roman";color:#1A1A1A;mso-bidi-font-weight:bold" lang="EN-IN">ketones and structural characterization of new compounds </span></span></span></span></span></span>

597-602Reactions of 1-aryl-3 -(5-nitro-2-thienyl/furyl)-2-propyne-1-one 3 with anilinoacet hydrazides 4<span style="font-size:14.0pt;font-family: " times="" new="" roman";color:#1a1a1a;mso-bidi-font-weight:bold"="" lang="EN-IN"> give the 1-anilinoacetyl-3-(5-nitro-2-thienyl/furyl)-5-aryl-5-hydroxypyrazolines 5-6 rather than the expected pyrazoles 7-<b style="mso-bidi-font-weight: normal">8. Various attempts have been made for the conversion of 5-6 to 7-8. The studies also reveal that the carbonyl group in 4 is responsible for the formation of hydroxypyrazolines. The structures of newly synthesized compounds have been proposed on the basis of elemental analysis and spectral data, and in some cases through the synthesis of authentic samples by alternative routes. </span