Tetrahydropyrazolo[1,5-a]Pyrimidine-3-Carboxamide and N-Benzyl-6′,7′-Dihydrospiro[Piperidine-4,4′-Thieno[3,2-c]Pyran] analogues with bactericidal efficacy against Mycobacterium tuberculosis targeting MmpL3

Mycobacterium tuberculosis is a major human pathogen and the causative agent for the pulmonary disease, tuberculosis (TB). Current treatment programs to combat TB are under threat due to the emergence of multi-drug and extensively-drug resistant TB. As part of our efforts towards the discovery of new anti-tubercular leads, a number of potent tetrahydropyrazolo[1,5-a]pyrimidine-3-ca​rboxamide(THPP) and N-benzyl-6′,7′-dihydrospiro[piperidine-4,​4′-thieno[3,2-c]pyran](Spiro) analogues were recently identified against Mycobacterium tuberculosis and Mycobacterium bovis BCG through a high-throughput whole-cell screening campaign. Herein, we describe the attractive in vitro and in vivo anti-tubercular profiles of both lead series. The generation of M. tuberculosis spontaneous mutants and subsequent whole genome sequencing of several resistant mutants identified single mutations in the essential mmpL3 gene. This ‘genetic phenotype’ was further confirmed by a ‘chemical phenotype’, whereby M. bovis BCG treated with both the THPP and Spiro series resulted in the accumulation of trehalose monomycolate. In vivo efficacy evaluation of two optimized THPP and Spiro leads showed how the compounds were able to reduce >2 logs bacterial cfu counts in the lungs of infected mice

CYP450 phenotyping and metabolite identification of quinine by accurate mass UPLC-MS analysis: a possible metabolic link to blackwater fever

BACKGROUND: The naturally occurring alkaloid drug, quinine is commonly used for the treatment of severe malaria. Despite centuries of use, its metabolism is still not fully understood, and may play a role in the haemolytic disorders associated with the drug. METHODS: Incubations of quinine with CYPs 1A2, 2C9, 2C19, 2D6, and 3A4 were conducted, and the metabolites were characterized by accurate mass UPLC-MS(E) analysis. Reactive oxygen species generation was also measured in human erythrocytes incubated in the presence of quinine with and without microsomes. RESULTS: The metabolites 3-hydroxyquinine, 2’-oxoquininone, and O-desmethylquinine were observed after incubation with CYPs 3A4 (3-hydroxyquinine and 2’-oxoquininone) and 2D6 (O-desmethylquinine). In addition, multiple hydroxylations were observed both on the quinoline core and the quinuclidine ring system. Of the five primary abundance CYPs tested, 3A4, 2D6, 2C9, and 2C19 all demonstrated activity toward quinine, while 1A2 did not. Further, quinine produced robust dose-dependent oxidative stress in human erythrocytes in the presence of microsomes. CONCLUSIONS: Taken in context, these data suggest a CYP-mediated link between quinine metabolism and the poorly understood haemolytic condition known as blackwater fever, often associated with quinine ingestion

Perspectives on Astrophysics Based on Atomic, Molecular, and Optical (AMO) Techniques

About two generations ago, a large part of AMO science was dominated by experimental high energy collision studies and perturbative theoretical methods. Since then, AMO science has undergone a transition and is now dominated by quantum, ultracold, and ultrafast studies. But in the process, the field has passed over the complexity that lies between these two extremes. Most of the Universe resides in this intermediate region. We put forward that the next frontier for AMO science is to explore the AMO complexity that describes most of the Cosmos.Comment: White paper submission to the Decadal Assessment and Outlook Report on Atomic, Molecular, and Optical (AMO) Science (AMO 2020

CLEC-2 and Syk in the megakaryocytic/platelet lineage are essential for development

The C-type lectin receptor CLEC-2 signals through a pathway that is critically dependent on the tyrosine kinase Syk. We show that homozygous loss of either protein results in defects in brain vascular and lymphatic development, lung inflation and perinatal lethality. Furthermore, we find that conditional deletion of Syk in the haematopoietic lineage, or conditional deletion of CLEC-2 or Syk in the megakaryocyte/platelet lineage, also causes defects in brain vascular and lymphatic development, although the mice are viable. In contrast, conditional deletion of Syk in other haematopoietic lineages had no effect on viability or brain vasculature and lymphatic development. We show that platelets, but not platelet releasate, modulate the migration and intercellular adhesion of lymphatic endothelial cells through a pathway that is dependent on CLEC-2 and Syk. These studies demonstrate that megakaryocyte/platelet expression of CLEC-2 and Syk is required for normal brain vasculature and lymphatic development and that platelet CLEC-2 and Syk directly modulate lymphatic endothelial cell behaviour in vitro

Stellar Astrophysics and Exoplanet Science with the Maunakea Spectroscopic Explorer (MSE)

The Maunakea Spectroscopic Explorer (MSE) is a planned 11.25-m aperture facility with a 1.5 square degree field of view that will be fully dedicated to multi-object spectroscopy. A rebirth of the 3.6m Canada-France-Hawaii Telescope on Maunakea, MSE will use 4332 fibers operating at three different resolving powers (R ~ 2500, 6000, 40000) across a wavelength range of 0.36-1.8mum, with dynamical fiber positioning that allows fibers to match the exposure times of individual objects. MSE will enable spectroscopic surveys with unprecedented scale and sensitivity by collecting millions of spectra per year down to limiting magnitudes of g ~ 20-24 mag, with a nominal velocity precision of ~100 m/s in high-resolution mode. This white paper describes science cases for stellar astrophysics and exoplanet science using MSE, including the discovery and atmospheric characterization of exoplanets and substellar objects, stellar physics with star clusters, asteroseismology of solar-like oscillators and opacity-driven pulsators, studies of stellar rotation, activity, and multiplicity, as well as the chemical characterization of AGB and extremely metal-poor stars.Comment: 31 pages, 11 figures; To appear as a chapter for the Detailed Science Case of the Maunakea Spectroscopic Explore

A Bioinformatics Classifier and Database for Heme-Copper Oxygen Reductases

Background: Heme-copper oxygen reductases (HCOs) are the last enzymatic complexes of most aerobic respiratory chains, reducing dioxygen to water and translocating up to four protons across the inner mitochondrial membrane (eukaryotes) or cytoplasmatic membrane (prokaryotes). The number of completely sequenced genomes is expanding exponentially, and concomitantly, the number and taxonomic distribution of HCO sequences. These enzymes were initially classified into three different types being this classification recently challenged. Methodology:We reanalyzed the classification scheme and developed a new bioinformatics classifier for the HCO and Nitric oxide reductases (NOR), which we benchmark against a manually derived gold standard sequence set. It is able to classify any given sequence of subunit I from HCO and NOR with a global recall and precision both of 99.8%. We use this tool to classify this protein family in 552 completely sequenced genomes. Conclusions: We concluded that the new and broader data set supports three functional and evolutionary groups of HCOs. Homology between NORs and HCOs is shown and NORs closest relationship with C Type HCOs demonstrated. We established and made available a classification web tool and an integrated Heme-Copper Oxygen reductase and NOR protein database (www.evocell.org/hco)

Molecular modeling of a tandem two pore domain potassium channel reveals a putative binding Site for general anesthetics

[Image: see text] Anesthetics are thought to mediate a portion of their activity via binding to and modulation of potassium channels. In particular, tandem pore potassium channels (K2P) are transmembrane ion channels whose current is modulated by the presence of general anesthetics and whose genetic absence has been shown to confer a level of anesthetic resistance. While the exact molecular structure of all K2P forms remains unknown, significant progress has been made toward understanding their structure and interactions with anesthetics via the methods of molecular modeling, coupled with the recently released higher resolution structures of homologous potassium channels to act as templates. Such models reveal the convergence of amino acid regions that are known to modulate anesthetic activity onto a common three- dimensional cavity that forms a putative anesthetic binding site. The model successfully predicts additional important residues that are also involved in the putative binding site as validated by the results of suggested experimental mutations. Such a model can now be used to further predict other amino acid residues that may be intimately involved in the target-based structure–activity relationships that are necessary for anesthetic binding

Suppression of LPS-induced inflammatory responses in macrophages infected with Leishmania

<p>Abstract</p> <p>Background</p> <p>Chronic inflammation activated by macrophage innate pathogen recognition receptors such as TLR4 can lead to a range of inflammatory diseases, including atherosclerosis, Crohn's disease, arthritis and cancer. Unlike many microbes, the kinetoplastid protozoan pathogen <it>Leishmania </it>has been shown to avoid and even actively suppress host inflammatory cytokine responses, such as LPS-induced IL-12 production. The nature and scope of <it>Leishmania</it>-mediated inflammatory cytokine suppression, however, is not well characterized. Advancing our knowledge of such microbe-mediated cytokine suppression may provide new avenues for therapeutic intervention in inflammatory disease.</p> <p>Methods</p> <p>We explored the kinetics of a range of cytokine and chemokine responses in primary murine macrophages stimulated with LPS in the presence versus absence of two clinically distinct species of <it>Leishmania </it>using sensitive multiplex cytokine analyses. To confirm that these effects were parasite-specific, we compared the effects of <it>Leishmania </it>uptake on LPS-induced cytokine expression with uptake of inert latex beads.</p> <p>Results</p> <p>Whilst <it>Leishmania </it>uptake alone did not induce significant levels of any cytokine analysed in this study, <it>Leishmania </it>uptake in the presence of LPS caused parasite-specific suppression of certain LPS-induced pro-inflammatory cytokines, including IL-12, IL-17 and IL-6. Interestingly, <it>L. amazonensis </it>was generally more suppressive than <it>L. major</it>. We also found that other LPS-induced proinflammatory cytokines, such as IL-1α, TNF-α and the chemokines MIP-1α and MCP-1 and also the anti-inflammatory cytokine IL-10, were augmented during <it>Leishmania </it>uptake, in a parasite-specific manner.</p> <p>Conclusions</p> <p>During uptake by macrophages, <it>Leishmania </it>evades the activation of a broad range of cytokines and chemokines. Further, in the presence of a strong inflammatory stimulus, <it>Leishmania </it>suppresses certain proinflammatory cytokine responses in a parasite-specific manner, however it augments the production of other proinflammatory cytokines. Our findings highlight the complexity of inflammatory cytokine signalling regulation in the context of the macrophage and <it>Leishmania </it>interaction and confirm the utility of the <it>Leishmania</it>/macrophage infection model as an experimental system for further studies of inflammatory regulation. Such studies may advance the development of therapies against inflammatory disease.</p

The Long-Baseline Neutrino Experiment: Exploring Fundamental Symmetries of the Universe

The preponderance of matter over antimatter in the early Universe, the dynamics of the supernova bursts that produced the heavy elements necessary for life and whether protons eventually decay --- these mysteries at the forefront of particle physics and astrophysics are key to understanding the early evolution of our Universe, its current state and its eventual fate. The Long-Baseline Neutrino Experiment (LBNE) represents an extensively developed plan for a world-class experiment dedicated to addressing these questions. LBNE is conceived around three central components: (1) a new, high-intensity neutrino source generated from a megawatt-class proton accelerator at Fermi National Accelerator Laboratory, (2) a near neutrino detector just downstream of the source, and (3) a massive liquid argon time-projection chamber deployed as a far detector deep underground at the Sanford Underground Research Facility. This facility, located at the site of the former Homestake Mine in Lead, South Dakota, is approximately 1,300 km from the neutrino source at Fermilab -- a distance (baseline) that delivers optimal sensitivity to neutrino charge-parity symmetry violation and mass ordering effects. This ambitious yet cost-effective design incorporates scalability and flexibility and can accommodate a variety of upgrades and contributions. With its exceptional combination of experimental configuration, technical capabilities, and potential for transformative discoveries, LBNE promises to be a vital facility for the field of particle physics worldwide, providing physicists from around the globe with opportunities to collaborate in a twenty to thirty year program of exciting science. In this document we provide a comprehensive overview of LBNE's scientific objectives, its place in the landscape of neutrino physics worldwide, the technologies it will incorporate and the capabilities it will possess.Comment: Major update of previous version. This is the reference document for LBNE science program and current status. Chapters 1, 3, and 9 provide a comprehensive overview of LBNE's scientific objectives, its place in the landscape of neutrino physics worldwide, the technologies it will incorporate and the capabilities it will possess. 288 pages, 116 figure

Salve Regina Arboretum Ten Year Plan to Reach Level III Accreditation

The Salve Regina University Arboretum, located in Newport, Rhode Island is currently registered as a Level II arboretum and is intertwined with the city of Newport Arboretum. The university now has intentions to reach Level III status, as part of a ten-year plan. This plan was developed by the students of the Spring 2018 BIO 255: Conservation Biology course, instructed by Dr. Jameson Chace, Associate Professor of biology at Salve Regina University. As part of a curriculum geared towards civic engagement, the class focused on creating and optimizing strategies that can be applied to the ten-year plan. These strategies were applied to the plan categorically: a team to inventory the current tree collection; a team to develop formal educational programming; a team for informal educational programming; a team to establish goals for conservation initiative related to the arboretum; a team dedicated to research related to arboreta; and a team to develop a list of species of special interest to add to the arboretum in the coming years. In the following document, each team’s strategies for the ten-year plan are outlined. Each of the components of this plan incorporate means to fulfill the conditions to meet Level III arboretum status so that the arboretum can apply for official registration. The aforementioned teams were tasked with designing a foundation on which to work up from. This includes formal educational programming to be applied to classroom settings and informal educational programming which can be applied to community outreach-based settings. The teams that worked to strengthen the arboretum’s mission of conservation focused on researching trees that can fit into the current landscape while providing some sort of benefit to the surrounding flora/fauna. Further, many of the species of interest, such as the chestnut, hold historical value to the greater Rhode Island region. In all, the Salve Regina Arboretum must achieve a total of 500 unique species of trees and woody plants as part of its efforts to apply for Level III status. In addition to the programming and research performed so far by the student teams, the arboretum must also hire a curator to manage the programming and to oversee the arboretum as a whole. Additionally, the arboretum must continue to actively collaborate with other arboreta and should encourage scientific research. It is important to recognize that the Salve Regina University Arboretum has already been utilized in the field of microbiology and has gained some attention at the university as a resource for further research and investigation. This ten year plan, along with resources within in it, is designed to provide a list of potential guidelines and ideas that can be applied for the arboretum’s benefit and growth. The Salve Regina University arboretum is a continually growing and developing part of the greater Newport, Rhode Island community, and will continue to strengthen its mission and that of the university which oversees its success.https://digitalcommons.salve.edu/bio255_arboretum/1000/thumbnail.jp