Ageing induces tissue-specific transcriptomic changes in Caenorhabditis elegans.

Ageing is a complex process with common and distinct features across tissues. Unveiling the underlying processes driving ageing in individual tissues is indispensable to decipher the mechanisms of organismal longevity. Caenorhabditis elegans is a well-established model organism that has spearheaded ageing research with the discovery of numerous genetic pathways controlling its lifespan. However, it remains challenging to dissect the ageing of worm tissues due to the limited description of tissue pathology and access to tissue-specific molecular changes during ageing. In this study, we isolated cells from five major tissues in young and old worms and profiled the age-induced transcriptomic changes within these tissues. We observed a striking diversity of ageing across tissues and identified different sets of longevity regulators therein. In addition, we found novel tissue-specific factors, including irx-1 and myrf-2, which control the integrity of the intestinal barrier and sarcomere structure during ageing respectively. This study demonstrates the complexity of ageing across worm tissues and highlights the power of tissue-specific transcriptomic profiling during ageing, which can serve as a resource to the field.

Quantitative Determination of Water in Organic Liquids by Ambient Mass Spectrometry.

This study uses a rapid tandem mass-spectrometry method to determine water content in complex organic solutions. Emphasis is placed on trace-water analysis by a fast and accurate alternative to the Karl-Fischer method. In this new method, water is captured by a charge-labeled molecular probe. Water binds strongly with high specificity to the strongly electrophilic aldehyde site in a charge-labelled molecule (N-methylpyridinium); competitive binding by other analytes is effectively discriminated against in the mass-measurement step. Quantitative determinations are made over a wide concentration range, 0.001 % (10 ppm) to 99 %, with better than 10 % relative standard deviation, along with short (1 min) analysis times using small sample volumes (several µL). Applications include water measurement in simple organic solvents, for example, deuterated solvents, as well as in complex mixtures, for example, organic reaction mixtures. Additionally, this method allows for water monitoring in levitated droplets. Mechanistic investigations into the impact of water on important chemical processes in organic synthesis and environmental science are reported.

Conversion of oxadiazolo[3,4-b]pyrazines to imidazo[4,5-b]pyrazines via a tandem reduction-cyclization sequence generates new mitochondrial uncouplers.

We report new mitochondrial uncouplers derived from the conversion of [1,2,5]oxadiazolo[3,4-b]pyrazines to 1H-imidazo[4,5-b]pyrazines. The in situ Fe-mediated reduction of the oxadiazole fragment followed by cyclization gave access to imidazopyrazines in moderate to good yields. A selection of orthoesters also allowed functionalization on the 2-position of the imidazole ring. This method afforded a variety of imidazopyrazine derivatives with varying substitution on the 2, 5 and 6 positions. Our studies suggest that both a 2-trifluoromethyl group and N-methylation are crucial for mitochondrial uncoupling capacity.

Uniform High-k Amorphous Native Oxide Synthesized by Oxygen Plasma for Top-Gated Transistors.

The integration of high-k gate dielectrics with two-dimensional (2D) semiconducting channel materials is essential for high-performance and low-power electronics. However, the conformal deposition of a uniform high-k dielectric with sub-1 nm equivalent oxide thickness (EOT) and high interface quality on high-mobility 2D semiconductors is still challenging. Here, we report a facile approach to synthesize a uniform high-k (εr ∼ 22) amorphous native oxide Bi2SeOx on the high-mobility 2D semiconducting Bi2O2Se using O2 plasma at room temperature. The conformal native oxide can directly serve as gate dielectrics with EOT of ∼0.9 nm, while the original properties of underlying 2D Bi2O2Se is preserved. Furthermore, high-resolution area-selective oxidation of Bi2O2Se is achieved to fabricate discrete electronic components. This facile integration of a high-mobility 2D semiconductor and its high-k native oxide holds high promise for next-generation nanoelectronics.

Anibamine and Its Analogues: Potent Antiplasmodial Agents from Aniba citrifolia.

In our continuing search for novel natural products with antiplasmodial activity, an extract of Aniba citrifolia was found to have good activity, with an IC50 value less than 1.25 µg/mL. After bioassay-directed fractionation, the known indolizinium alkaloid anibamine (1) and the new indolizinium alkaloid anibamine B (2) were isolated as the major bioactive constituents, with antiplasmodial IC50 values of 0.170 and 0.244 µM against the drug-resistant Dd2 strain of Plasmodium falciparum. The new coumarin anibomarin A (3), the new norneolignan anibignan A (5), and six known neolignans (7-12) were also obtained. The structures of all the isolated compounds were determined based on analyses of 1D and 2D NMR spectroscopic and mass spectrometric data, and the absolute configuration of anibignan A (5) was assigned from its ECD spectrum. Evaluation of a library of 28 anibamine analogues (13-40) indicated that quaternary charged analogues had IC50 values as low as 58 nM, while uncharged analogues were inactive or significantly less active. Assessment of the potential effects of anibamine and its analogues on the intraerythrocytic stages and morphological development of P. falciparum revealed substantial activity against ring stages for compounds with two C-10 side chains, while those with only one C-10 side chain exhibited substantial activity against trophozoite stages, suggesting different mechanisms of action.

Molecular recognition of a branched peptide with HIV-1 Rev Response Element (RRE) RNA.

Interaction of HIV-1 rev response element (RRE) RNA with its cognate protein, Rev, is critical for HIV-1 replication. Understanding the mode of interaction between RRE RNA and ligands at the binding site can facilitate RNA molecular recognition as well as provide a strategy for developing anti-HIV therapeutics. Our approach utilizes branched peptides as a scaffold for multivalent binding to RRE IIB (high affinity rev binding site) with incorporation of unnatural amino acids to increase affinity via non-canonical interactions with the RNA. Previous high throughput screening of a 46,656-member library revealed several hits that bound RRE IIB RNA in the sub-micromolar range. In particular, the lead compound, 4B3, displayed a Kd value of 410 nM and demonstrated selectivity towards RRE. A ribonuclease protection assay revealed that 4B3 binds to the stem-loop structure of RRE IIB RNA, which was confirmed by SHAPE analysis with 234 nt long NL4-3 RRE RNA. Our studies further indicated interaction of 4B3 with both primary and secondary Rev binding sites.

Structural Evidence for Rifampicin Monooxygenase Inactivating Rifampicin by Cleaving Its Ansa-Bridge.

Rifampicin monooxygenase (RIFMO) decreases the potency of rifampicin (RIF) by converting it to oxidative products. Further decomposition of RIF has been observed in bacteria producing RIFMO and contributes to RIFMO-mediated drug resistance. Here we report the first crystal structure of RIFMO in complex with the hydroxylated RIF product. The 2.10 Å resolution structure reveals a breach of the ansa aliphatic chain of RIF between naphthoquinone C2 and amide N1. Our data suggest that RIFMO catalyzes the hydroxylation of RIF at the C2 atom followed by cleavage of the ansa linkage, which leads to inactivation of the antibiotic by preventing key contacts with the RNA polymerase target.

Evidence for the Formation of a Radical-Mediated Flavin-N5 Covalent Intermediate.

The redox-neutral reaction catalyzed by 2-haloacrylate hydratase (2-HAH) leads to the conversion of 2-chloroacrylate to pyruvate. Previous mechanistic studies demonstrated the formation of a flavin-iminium ion as an important intermediate in the 2-HAH catalytic cycle. Time-resolved flavin absorbance studies were performed in this study, and the data showed that the enzyme is capable of stabilizing both anionic and neutral flavin semiquinone species. The presence of a radical scavenger decreases the activity in a concentration-dependent manner. These data are consistent with the flavin iminium intermediate occurring by radical recombination.

Flavin-N5 Covalent Intermediate in a Nonredox Dehalogenation Reaction Catalyzed by an Atypical Flavoenzyme.

The flavin-dependent enzyme 2-haloacrylate hydratase (2-HAH) catalyzes the conversion of 2-chloroacrylate, a major component in the manufacture of acrylic polymers, to pyruvate. The enzyme was expressed in Escherichia coli, purified, and characterized. 2-HAH was shown to be monomeric in solution and contained a non-covalent, yet tightly bound, flavin adenine dinucleotide (FAD). Although the catalyzed reaction was redox-neutral, 2-HAH was active only in the reduced state. A covalent flavin-substrate intermediate, consistent with the flavin-acrylate iminium ion, was trapped with cyanoborohydride and characterized by mass spectrometry. Small-angle X-ray scattering was consistent with 2-HAH belonging to the succinate dehydrogenase/fumarate reductase family of flavoproteins. These studies establish 2-HAH as a novel noncanonical flavoenzyme.

Novel Bioactive Natural Products Isolated from Madagascar Plants and Marine Organisms (2009-2017).

Madagascar's rain forests and tropical dry forests are home to numerous endemic plant species and the island is considered a biodiversity hotspot. About 80% of the Madagascan (Malagasy) population relies on traditional medicines that have been proven to contain a variety of biologically active compounds. In the search for bioactive compounds from Madagascan biodiversity, we accessed and collected most of the literature dealing with the isolation, structure elucidation, and biological activities of organic small molecules originating from Madagascan plants and marine organisms. Since we published the first review of this work in 2009 (Curr. Med. Chem., 17, 2010, Hou and Harinantenaina), the present paper covers the isolation, structures, and bioactivity of 182 new secondary metabolites isolated from Malagasy higher plants and marine organisms in the last seven years (2009-2017).

Antiplasmodial Sesquiterpenoid Lactones from Trichospira verticillata: Structure Elucidation by Spectroscopic Methods and Comparison of Experimental and Calculated ECD Data.

A dichloromethane extract of Trichospira verticillata from the Natural Products Discovery Institute was discovered to have good antiplasmodial activity (IC50 ∼5 µg/mL). After purification by liquid-liquid partition and C18 reversed-phase HPLC, four new germacranolide-type sesquiterpenoid lactones named trichospirolides A-D (1-4) were isolated. The structures of the new compounds were elucidated by analysis of their 1D and 2D NMR and MS data. The relative and absolute configurations were assigned based on a comparison of calculated and experimental ECD and UV spectra, specific rotations, internuclear distances, and coupling constants for all possible diastereomers for each compound. Among these four compounds, the conjugated dienone 1 displayed the most potent antiplasmodial activity, with an IC50 value of 1.5 µM.

Substrate-Assisted, Transition-Metal-Free Diboration of Alkynamides with Mixed Diboron: Regio- and Stereoselective Access to trans-1,2-Vinyldiboronates.

A substrate-assisted diboration of alkynamides using the unsymmetrical pinacolato-1,8-diaminonaphthalenato diboron (pinBBdan) is described. The transition-metal-free reaction proceeds in a regio- and stereoselective fashion to exclusively afford trans-vinyldiboronates in good to excellent yields. Notably, Bdan and Bpin are installed on the α- and ß-carbon atoms, respectively.

In Crystallo Capture of a Covalent Intermediate in the UDP-Galactopyranose Mutase Reaction.

UDP-galactopyranose mutase (UGM) plays an essential role in galactofuranose biosynthesis in pathogens by catalyzing the conversion of UDP-galactopyranose to UDP-galactofuranose. Here we report the first crystal structure of a covalent intermediate in the UGM reaction. The 2.3 Å resolution structure reveals UDP bound in the active site and galactopyranose linked to the FAD through a covalent bond between the anomeric C of galactopyranose and N5 of the FAD. The structure confirms the role of the flavin as nucleophile and supports the hypothesis that the proton destined for O5 of galactofuranose is shuttled from N5 of the FAD via O4 of the FAD.

Design, synthesis and structure activity relationship of potent pan-PIM kinase inhibitors derived from the pyridyl carboxamide scaffold.

The Pim proteins (1, 2 and 3) are serine/threonine kinases that have been found to be upregulated in many hematological malignancies and solid tumors. As a result of overlapping functions among the three isoforms, inhibition of all three Pim kinases has become an attractive strategy for cancer therapy. Herein we describe our efforts in identifying potent pan-PIM inhibitors that are derived from our previously reported pyridyl carboxamide scaffold as part of a medicinal chemistry strategy to address metabolic stability.

Pim2 is required for maintaining multiple myeloma cell growth through modulating TSC2 phosphorylation.

Multiple myeloma (MM) is the second most common hematologic malignancy. Despite recent treatment advances, it remains incurable. Here, we report that Pim2 kinase expression is highly elevated in MM cells and demonstrate that it is required for MM cell proliferation. Functional interference of Pim2 activity either by short hairpin RNAs or by a potent and selective small-molecule inhibitor leads to significant inhibition of MM cell proliferation. Pim inhibition results in a significant decrease of mammalian target of rapamycin C1 (mTOR-C1) activity, which is critical for cell proliferation. We identify TSC2, a negative regulator of mTOR-C1, as a novel Pim2 substrate and show that Pim2 directly phosphorylates TSC2 on Ser-1798 and relieves the suppression of TSC2 on mTOR-C1. These findings support Pim2 as a promising therapeutic target for MM and define a novel Pim2-TSC2-mTOR-C1 pathway that drives MM proliferation.

Isolation of antiplasmodial anthraquinones from Kniphofia ensifolia, and synthesis and structure-activity relationships of related compounds.

Bioassay-guided separation of the South African plant Kniphofia ensifolia for antiplasmodial activity led to the isolation of two new anthraquinones, named kniphofiones A and B (3 and 4), together with three known bioactive anthraquinone monomers (1, 2 and 5), and four known bisanthraquinones (6-9). The structures of the two new compounds were elucidated based on analyses of their 1D and 2D NMR spectra and mass spectrometric data. The dimeric compounds 6 and 7 displayed the strongest antiplasmodial activity among all the isolated compounds, with IC50 values of 0.4 ± 0.1 and 0.2 ± 0.1 µM, respectively. The two new compounds displayed modest activities, with IC50 values of 26 ± 4 and 9 ± 1 µM, respectively. Due to the synthetic accessibility of the new compounds and the increased activity shown by the dimeric compounds, a structure-activity relationship study was conducted. As a result, one analogue of kniphofione B (4), the caffeic acid derivative of aloe-emodin, was found to have the highest activity among all the aloe-emodin derivatives, with an IC50 value of 1.3 ± 0.2 µM.

BLMP-1 is a critical temporal regulator of dietary-restriction-induced response in Caenorhabditis elegans.

The extrinsic diet and the intrinsic developmental programs are intertwined. Although extensive research has been conducted on how nutrition regulates development, whether and how developmental programs control the timing of nutritional responses remain barely known. Here, we report that a developmental timing regulator, BLMP-1/BLIMP1, governs the temporal response to dietary restriction (DR). At the end of larval development, BLMP-1 is induced and interacts with DR-activated PHA-4/FOXA, a key transcription factor responding to the reduced nutrition. By integrating temporal and nutritional signaling, the DR response regulates many development-related genes, including gska-3/GSK3ß, through BLMP-1-PHA-4 at the onset of adulthood. Upon DR, a precocious activation of BLMP-1 in early larval stages impairs neuronal development through gska-3, whereas the increase of gska-3 by BLMP-1-PHA-4 at the last larval stage suppresses WNT signaling in adulthood for DR-induced longevity. Our findings reveal a temporal checkpoint of the DR response that protects larval development and promotes adult health.

Liver-Selective Imidazolopyrazine Mitochondrial Uncoupler SHD865 Reverses Adiposity and Glucose Intolerance in Mice.

Excess body fat is a risk factor for metabolic diseases and is a leading preventable cause of morbidity and mortality worldwide. There is a strong need to find new treatments that decrease the burden of obesity and lower the risk of obesity-related comorbidities, including cardiovascular disease and type 2 diabetes. Pharmacologic mitochondrial uncouplers represent a potential treatment for obesity through their ability to increase nutrient oxidation. Herein, we report the in vitro and in vivo characterization of compound SHD865, the first compound to be studied in vivo in a newly discovered class of imidazolopyrazine mitochondrial uncouplers. SHD865 is a derivative of the furazanopyrazine uncoupler BAM15. SHD865 is a milder mitochondrial uncoupler than BAM15 that results in a lower maximal respiration rate. In a mouse model of diet-induced adiposity, 6-week treatment with SHD865 completely restored normal body composition and glucose tolerance to levels like those of chow-fed controls, without altering food intake. SHD865 treatment also corrected liver steatosis and plasma hyperlipidemia to normal levels comparable with chow-fed controls. SHD865 has maximal oral bioavailability in rats and slow clearance in human microsomes and hepatocytes. Collectively, these data identify the potential of imidazolopyrazine mitochondrial uncouplers as drug candidates for the treatment of obesity-related disorders.

Antiproliferative Homoisoflavonoids and Bufatrienolides from Urginea depressa.

Investigation of the South African plant Urginea depressa Baker (Asparagaceae Juss.) for antiproliferative activity against the A2780 ovarian cancer cell line led to the isolation of the six new homoisoflavonoids urgineanins A-F (1-6), the two known bufatrienolides 7 and 9, and the new bufatrienolides urginins B and C (8 and 10). Structures were elucidated based on analysis of their 1D and 2D NMR spectra, electronic circular dichroism, and mass spectrometric data. Five of the six new homoisoflavonoids had good antiproliferative activity against the A2780 ovarian cancer, A2058 melanoma, and H522-T1 human non-small-cell lung cancer cells, and urgineanin A (1) had submicromolar activity against all three cell lines. The four bufatrienolides 7-10 had strong antiproliferative activity against the same cell line, with IC50 values of 24.1, 11.2, 111, and 40.6 nM, respectively.

Two antiproliferative triterpene saponins from Nematostylis anthophylla from the highlands of Central Madagascar.

Investigation of the endemic Madagascan plant Nematostylis anthophylla (Rubiaceae) for antiproliferative activity against the A2780 ovarian cancer cell line led to the isolation of the known triterpene saponin randianin (1), and the two new bioactive triterpene saponins 2"-O-acetylrandianin (2) and 6"-O-acetylrandianin (3). The structures of the two new compounds were elucidated based on analysis of their 1D- and 2D-NMR spectra, and mass spectrometric data. The three isolated triterpene saponins displayed moderate but selective antiproliferative activities, with IC(50) values of 1.2, 1.7, and 2.2 µM, respectively, against the A2780 ovarian cancer, but only weak inhibitions of the proliferation of A2058 melanoma and the H522 lung cancer cell lines.