The impacts of natural product miltirone and the CYP2D6 pharmacogenetic phenotype on fluoxetine metabolism.

Introduction: To study the effects of drug-induced CYP2D6 activity inhibition and genetic polymorphisms on fluoxetine metabolism, rat liver microsomes (RLMs) and SD rats were used to investigate the potential drug‒drug interactions (DDIs), and CYP2D6 http://muchong.com/t-10728934-1 recombinant baculosomes were prepared and subjected to catalytic reactivity studies. Methods and Results: All analytes were detected by ultraperformance liquid chromatography-tandem mass spectrometry (UPLC‒MS/MS). After screening for 27 targeted natural products, miltirone was identified as having obvious inhibitory effect on fluoxetine metabolism in RLMs. In vivo, the concentration of fluoxetine in rat blood increased markedly after miltirone administration. The molecular docking results showed that miltirone bound more strongly to CYP2D6 than fluoxetine, and PHE120 may be the key residue leading to the inhibition of CYP2D6-mediated fluoxetine N-demethylation by miltirone. In terms of the genetic polymorphism of CYP2D6 on fluoxetine metabolism, the intrinsic clearance values of most variants were significantly altered. Among these variants, CYP2D6*92 and CYP2D6*96/Q424X were found to be catalytically inactive for fluoxetine metabolism, five variants (CYP2D6*89/L142S, *97/F457L, *R497, *V342M and *R344Q) exhibited markedly increased clearance values (>125.07%) and seven variants (CYP2D6*2, *10, *87/A5V, *93/T249P, *E215K, *R25Q and *R440C) exhibited significantly decreased clearance values (from 6.62% to 66.79%) compared to those of the wild-type. Conclusion: Our results suggest that more attention should be given to subjects in the clinic who take fluoxetine and also carry one of these infrequent CYP2D6 alleles or are coadministered drugs containing miltirone.

Cerastecins inhibit membrane lipooligosaccharide transport in drug-resistant Acinetobacter baumannii.

Carbapenem-resistant Acinetobacter baumannii infections have limited treatment options. Synthesis, transport and placement of lipopolysaccharide or lipooligosaccharide (LOS) in the outer membrane of Gram-negative bacteria are important for bacterial virulence and survival. Here we describe the cerastecins, inhibitors of the A. baumannii transporter MsbA, an LOS flippase. These molecules are potent and bactericidal against A. baumannii, including clinical carbapenem-resistant Acinetobacter baumannii isolates. Using cryo-electron microscopy and biochemical analysis, we show that the cerastecins adopt a serpentine configuration in the central vault of the MsbA dimer, stalling the enzyme and uncoupling ATP hydrolysis from substrate flipping. A derivative with optimized potency and pharmacokinetic properties showed efficacy in murine models of bloodstream or pulmonary A. baumannii infection. While resistance development is inevitable, targeting a clinically unexploited mechanism avoids existing antibiotic resistance mechanisms. Although clinical validation of LOS transport remains undetermined, the cerastecins may open a path to narrow-spectrum treatment modalities for important nosocomial infections.

Design, Synthesis of Hydrogen Peroxide Response AIE Fluorescence Probes Based on Imidazo [1,2-a] Pyridine.

Hydrogen peroxide (H2O2), a significant member of reactive oxygen species, plays a crucial role in oxidative stress and cell signaling. Abnormal levels of H2O2 in the body can induce damage or even impair body function, leading to the development of certain diseases. Therefore, real-time monitoring of H2O2 in living cells is very important. In this work, the aggregation-induced emission fluorescence probe 2-(2-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl) benzyl) oxy) phenyl) imidazo [1,2-a] pyridine (B2) was designed and synthesized, which enables the long-term tracing of H2O2 in living cells. The addition of H2O2 to probe B2 results in a dramatic fluorescence enhancement around 500 nm. Notably, B2 can visualize both exogenous and endogenous H2O2 in living cells. The synthesis method for B2 is simple, has a high yield, and utilizes readily available materials. It exhibits advantages such as low toxicity, photostability, and good biocompatibility. Consequently, the developed fluorescent probe in this study has great potential as a reliable tool for determining H2O2 in living cells.

Ganodermanontriol regulates tumor-associated M2 macrophage polarization in gastric cancer.

AIM: We focused on investigating the role and mechanism of ganodermanontriol (GAN) in regulating the M2 polarization of tumor-associated macrophages in the gastric cancer microenvironment. METHODS: M2 polarization of RAW264.7 macrophages was induced by IL-4 or co-culture with MFC, and the expression levels of M1 macrophage markers (TNF-α, IFN-γ, IL-1ß) and M2 macrophage markers (IL-10, TGF-ß, Arg-1) were detected by enzyme-linked immunosorbed assay (ELISA). The protein expression was assayed by Western-Blotting. For in vitro experiments, a tumor-bearing mouse model was established, with which the CD206 level was detected by histochemistry, and the binding mode between GAN and STAT6 was simulated through molecular dynamics. RESULTS: Both IL-4 and MFC could induce the M2 polarization of macrophages. GAN could inhibit such polarization, which produced unobvious effects on M1 markers, but could suppress the levels of M2 markers. GAN could inhibit the phosphorylated expression of STAT6, and M2 macrophages treated by it had a weakened ability to promote malignant behavior of MFC. According to the results of in vitro experiments, GAN could inhibit tumor growth, suppress the tissue infiltration of CD206 cells, and inhibit the phosphorylated expression of STAT6. CONCLUSION: Our results show that GAN can inhibit the M2 macrophage polarization in gastric cancer microenvironment, whose mechanism of action is associated with the regulation of STAT6 phosphorylation.

Twin-Field Quantum Key Distribution without Phase Locking.

Twin-field quantum key distribution (TF-QKD) has emerged as a promising solution for practical quantum communication over long-haul fiber. However, previous demonstrations on TF-QKD require the phase locking technique to coherently control the twin light fields, inevitably complicating the system with extra fiber channels and peripheral hardware. Here, we propose and demonstrate an approach to recover the single-photon interference pattern and realize TF-QKD without phase locking. Our approach separates the communication time into reference frames and quantum frames, where the reference frames serve as a flexible scheme for establishing the global phase reference. To do so, we develop a tailored algorithm based on fast Fourier transform to efficiently reconcile the phase reference via data postprocessing. We demonstrate no-phase-locking TF-QKD from short to long distances over standard optical fibers. At 50-km standard fiber, we produce a high secret key rate (SKR) of 1.27 Mbit/s, while at 504-km standard fiber, we obtain the repeaterlike key rate scaling with a SKR of 34 times higher than the repeaterless secret key capacity. Our work provides a scalable and practical solution to TF-QKD, thus representing an important step towards its wide applications.

3D culture of the spinal cord with roots as an ex vivo model for comparative studies of motor and sensory nerve regeneration.

Motor and sensory nerves exhibit tissue-specific structural and functional features. However, in vitro models designed to reflect tissue-specific differences between motor and sensory nerve regeneration have rarely been reported. Here, by embedding the spinal cord with roots (SCWR) in a 3D hydrogel environment, we compared the nerve regeneration processes between the ventral and dorsal roots. The 3D hydrogel environment induced an outward migration of neurons in the gray matter of the spinal cord, which allowed the long-term survival of motor neurons. Tuj1 immunofluorescence labeling confirmed the regeneration of neurites from both the ventral and dorsal roots. Next, we detected asymmetric ventral and dorsal root regeneration in response to nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF), and we observed motor and sensory Schwann cell phenotypes in the regenerated ventral and dorsal roots, respectively. Moreover, based on the SCWR model, we identified a targeted effect of collagen VI on sensory nerve fasciculation and characterized the protein expression profiles correlating to motor/sensory-specific nerve regeneration. These results suggest that the SCWR model can serve as a valuable ex vivo model for comparative study of motor and sensory nerve regeneration and for pharmacodynamic evaluations.

A review on carbon quantum dots: Synthesis, photoluminescence mechanisms and applications.

Carbon quantum dots (CQDs), having outstanding biocompatibility, attractive catalytic performance, excellent optical properties, and valuable environment friendliness, are emerging as a new paradigm to design luminescent devices and show great potential in application fields such as biomedical sensors, optical and photonic devices. Furthermore, CQDs are known as one of the most promising carbon-based nanomaterials in the 21st century. Therefore, they have attracted a lot of attention since they were first discovered in 2004. In this review, we explain the accepted photoluminescence mechanism of CQDs, including fluorescence and phosphorescence. There are two main types of synthesis strategies: top-down approach and bottom-up approach. At the same time, the main application fields, including ion detection, anti-counterfeiting, biological imaging, food safety, sensors, lubrication additives, are reviewed. Finally, the existing bottlenecks, pending problems and prospects for the future of CQDs are discussed.

Total Synthesis of Darobactin A.

The collaborative total synthesis of darobactin A, a recently isolated antibiotic that selectively targets Gram-negative bacteria, has been accomplished in a convergent fashion with a longest linear sequence of 16 steps from d-Garner's aldehyde and l-serine. Scalable routes toward three non-canonical amino acids were developed to enable the synthesis. The closure of the bismacrocycle was realized through sequential, halogen-selective Larock indole syntheses, where the proper order of cyclizations proved crucial for the formation of the desired atropisomer of the natural product.

Promoted Hydrogenolysis of Furan Aldehydes to 2,5-Dimethylfuran by Defect Engineering on Pd/NiCo2 O4.

Catalytic hydrogenolysis of biobased furan aldehydes (i. e., 5-methylfurfural, 5-hydroxymethylfurfural) to 2,5-dimethylfuran has gained extensive interest for biomass-derived fuels and chemicals. Herein, a class of NiCo2 O4 -supported palladium with considerable oxygen defects was synthesized by hydrogen plasma etching and phosphating methods. The oxygen defects not only promoted the hydrogenation of the C=O group but also enhanced the accessibility of coordinatively unsaturated metal cations with Lewis acidity for the hydrogenolysis of the C-OH group. Meanwhile, the additional Brønsted acidity in Pd/NiCo2 O4-x obtained by phosphating could further strengthen the hydrogenolysis ability by the etherification route of C-OH. Finally, Pd/NiCo2 O4-x exhibited the most effective performance with 2,5-dimethylfuran yields of 92.9 and 90.5 % from 5-methylfurfural and 5-hydroxymethylfurfural, respectively. These catalytic mechanisms were confirmed by in-situ infrared spectroscopy and control experiments. Furthermore, the catalyst showed outstanding recycling stability. This work shows powerful synergistic catalysis in the hydrogenolysis reaction by multifunctional active sites.

Advancing Structure Characterization of PS-80 by Charge-Reduced Mass Spectrometry and Software-Assisted Composition Analysis.

The commercially available Polysorbate 80 (PS-80) is a highly heterogeneous product. It is a complex and structurally diverse mixture consisting of polymeric species containing polyoxyethylenes (POEs), fatty acid esters, with/or without a carbohydrate core. The core is primarily sorbitan, with some isosorbide and sorbitol. Depending on the sources of fatty acids and the degrees of esterification, multiple combinations of fatty acid esters are commonly observed. A number of POE intermediates, such as polyoxyethylene glycols, POE-sorbitans, POE-isosorbides, and an array of fatty acid esters from these intermediates remain in the raw material as well. The complex composition of PS-80 is difficult to control and poses a significant characterization challenge for its use in the pharmaceutical industry. Here, we present a novel solution for PS-80 characterization using ultra high-performance liquid chromatography coupled with charge-reduction high resolution mass spectrometry. Post column co-infusion of triethylamine focused the signal into mainly singly charged molecular ions and reduced the extent of in-source fragmentation, resulting in a simpler ion map and enhanced measurement of PS-80 species. The data processing workflow is designed to programmatically identify PS-80 component classes and reduce the burden of manually analyzing complex MS data. The 2-dimensional graphical representation of the data helps visualize these features. Together, these innovative methodologies enabled us to analyze components in PS-80 with unprecedented detail and shall be a useful tool to study formulation and stability of pharmaceutical preparations. The power of this approach was demonstrated by comparing the composition of PS-80 obtained from different vendors.

Shp2 regulates PM2.5-induced airway epithelial barrier dysfunction by modulating ERK1/2 signaling pathway.

The impact of fine particulate matter (PM2.5) on public health has received increasing attention. Through various biochemical mechanisms, PM2.5 alters the normal structure and function of the airway epithelium, causing epithelial barrier dysfunction. Src homology domain 2-containing protein tyrosine phosphatase 2 (Shp2) has been implicated in various respiratory diseases; however, its role in PM2.5-induced epithelial barrier dysfunction remains unclear. Herein, we assessed the regulatory effects of Shp2 on PM2.5-mediated epithelial barrier function and tight junction (TJ) protein expression in both mice and human pulmonary epithelial (16HBE) cells. We observed that Shp2 levels were upregulated and claudin-4 levels were downregulated after PM2.5 stimulation in vivo and in vitro. Mice were exposed to PM2.5 to induce acute lung injury, and disrupted epithelial barrier function, with decreased transepithelial electrical resistance (TER) and increased paracellular flux that was observed in 16HBE cells. In contrast, the selective inhibition or knockdown of Shp2 retained airway epithelial barrier function and reversed claudin-4 downregulation that triggered by PM2.5, and these effects may occur through the ERK1/2 MAPK signaling pathway. These data highlight an important role of Shp2 in PM2.5-induced airway epithelial barrier dysfunction and suggest a possible new course of therapy for PM2.5-induced respiratory diseases.

A Simple and Low-Cost Method for Fabrication of Polydimethylsiloxane Microfludic Chips.

The conventional fabrication methods for enrichment microfluidic devices require cleanroom, which are costly and time-consuming. Developing a facile and low-cost method to fabricate microfluidic chips could stimulate the progress of the applications of those chips. Here, we present an easy method for fabrication of a complete PDMS (Polydimethylsiloxane) microfluidic chip used for ion and protein enrichment. The method consists of three main fabrication steps: PDMS microchannels ablation by co2 laser, nation membrane deposition, and oxygen plasma assist bonding under pressure. To fabricate a desired microchannel, the laser ablation parameters, containing laser power and ablation speed, were analyzed. The parameters for oxygen plasma assist bonding were also investigated to improve the bonding quality of the chips (low dimension loss and high bonding strength). The following Rhodamine B enrichment tests demonstrate that the presented method allows fabrication of microfluidic chips with precise dimensions and leakage free.

Development of a highly efficient decontamination approach for ceftolozane in the pharmaceutical manufacturing environment.

The ß-lactam core is a key structure responsible for inducing both IgE-mediated acute-onset hypersensitivity and T-cell-mediated delayed-onset hypersensitivity with penicillins in humans. There is essentially no clinically significant immunologic cross-reactivity noted between the ß-lactam cores of penicillins and cephalosporins based on challenge studies in humans. The side-chains appear to be more important in inducing IgE-mediated acute-onset hypersensitivity and T-cell delayed-onset hypersensitivity with cephalosporins in humans. Despite these clinical findings, the U. S. Food and Drug Administration (FDA) still requires the level of ß-lactam-related antibiotic residues to be controlled at very low levels in manufacturing facilities. Ceftolozane is Merck & Co., Inc., Kenilworth, NJ, USA's (MSD's) 5th generation broad spectrum cephalosporin antibiotic against gram-negative bacteria. In searching for the optimal decontamination method of ceftolozane, most methods were found to be very slow in opening the ß-lactam ring in ceftolozane. Moreover, most of the previously reported decontamination methods applied analytical methods that only monitored the disappearance of the parent molecule as the endpoint of degradation. In this way, many of the ß-lactam-containing degradation products could be overlooked. In order to develop an efficient decontamination solution for ceftolozane, a sensitive ultra high performance liquid chromatography-high resolution-electrospray ionization-tandem mass spectrometry (UHPLC-HRMS/MS) method was first developed to ensure the detection of the ß-lactam ring in all degradation products. Through online UHPLC-UV-HRMS monitoring, 2.5 N KOH in 50% aqueous MeOH or 50% aqueous EtOH was identified as the best condition to fully degrade the ß-lactam ring in ceftolozane. This decontamination could be done within 15 min, even at 100 mg/mL concentration, and thus enable a quick turnaround time for equipment cleaning in the ß-lactam manufacturing facility. This method was also successfully applied to 12 other commercially available ß-lactam antibiotics.

Unexpected Propargylic Retro-Brook Rearrangements in Alkynes.

Retro-Brook rearrangements refer to the intramolecular migration of a silyl group from oxygen to carbon. In this study, we report a novel propargylic retro-Brook rearrangement observed in terminal alkynes bearing a silyl ether moiety. Retro-Brook rearrangements involving [1,2]-, [1,4]-, and [1,5]-migrations are described, affording propargylsilanes in reasonable yield. The reaction mechanism was investigated experimentally by deuterium quenching and rationalized by density functional theory calculations. The terminal alkyne and the subsequent propargyl/allenyl dianion were shown to be crucial for the reaction favoring the retro-Brook rearrangement product over the Brook rearrangement. The second deprotonation at the propargylic position was determined to be the rate-limiting step. In addition, a gas-phase Brook-type rearrangement of the propargylsilanes was observed under GC-MS conditions. This observation was also further confirmed by DFT calculations.

MK-8353: Discovery of an Orally Bioavailable Dual Mechanism ERK Inhibitor for Oncology.

The emergence and evolution of new immunological cancer therapies has sparked a rapidly growing interest in discovering novel pathways to treat cancer. Toward this aim, a novel series of pyrrolidine derivatives (compound 5) were identified as potent inhibitors of ERK1/2 with excellent kinase selectivity and dual mechanism of action but suffered from poor pharmacokinetics (PK). The challenge of PK was overcome by the discovery of a novel 3(S)-thiomethyl pyrrolidine analog 7. Lead optimization through focused structure-activity relationship led to the discovery of a clinical candidate MK-8353 suitable for twice daily oral dosing as a potential new cancer therapeutic.

Chemoselective Peptide Modification via Photocatalytic Tryptophan ß-Position Conjugation.

Targeting tryptophan is a promising strategy to achieve high levels of selectivity for peptide or protein modification. A chemoselective peptide modification method via photocatalytic tryptophan ß-position conjugation has been discovered. This transformation has good substrate scope for both peptide and Michael acceptor, and has good chemoselectivity versus other amino acid residues. The endogenous peptides, glucagon and GLP-1 amide, were both successfully conjugated at the tryptophan ß-position. Insulin was studied as a nontryptophan control molecule, resulting in exclusive B-chain C-terminal-selective decarboxylative conjugation. This transformation provides a novel approach toward peptide modification to support the discovery of new therapeutic peptides, protein labeling and bioconjugation.

Discovery of 3(S)-thiomethyl pyrrolidine ERK inhibitors for oncology.

Compound 5 (SCH772984) was identified as a potent inhibitor of ERK1/2 with excellent selectivity against a panel of kinases (0/231 kinases tested @ 100 nM) and good cell proliferation activity, but suffered from poor PK (rat AUC PK @10 mpk = 0 µM h; F% = 0) which precluded further development. In an effort to identify novel ERK inhibitors with improved PK properties with respect to 5, a systematic exploration of sterics and composition at the 3-position of the pyrrolidine led to the discovery of a novel 3(S)-thiomethyl pyrrolidine analog 28 with vastly improved PK (rat AUC PK @10 mpk = 26 µM h; F% = 70).

Discovery of a highly potent orally bioavailable imidazo-[1, 2-a]pyrazine Aurora inhibitor.

Imidazo-[1, 2-a]pyrazine 1 is a potent inhibitor of Aurora A and B kinase in vitro and is effective in in vivo tumor models, but has poor oral bioavailbility and is unsuitable for oral dosing. We describe herein our effort to improve oral exposure in this class, resulting ultimately in the identification of a potent Aurora inhibitor 16, which exhibited good drug exposure levels across species upon oral dosing, and showed excellent in vivo efficacy in a mouse xenograft tumor model when dosed orally.

The synthesis of 2,3,6-trisubstituted 1-oxo-1,2-dihydroisoquinolines as potent CRTh2 antagonists.

New synthetic methods were developed for the preparation of 2,3,6-trisubstituted 1-oxo-1,2-dihydroisoquinolines as CRTh2 antagonists. The isoquinolinone core could be constructed before the introduction of substitution groups or synthesized through a catalytic intramolecular cyclization reaction with desired substitution groups properly installed. These synthetic strategies have helped to accelerate the SAR development of this series, and potent lead compounds were identified in both the CRTh2 receptor binding assay and the CD11b biomarker assay.

Unusual (+/-)-electrospray ionization induced fragmentation: Structural elucidation of an in-process synthetic intermediate of doravirine (MK-1439) using liquid chromatography/high-resolution tandem mass spectrometry and two-dimensional nuclear magnetic resonance.

RATIONALE: During the development of a novel synthetic route to doravirine (1), a human immunodeficiency type 1 virus (HIV-1) nonnucleoside reverse transcriptase inhibitor (NNRTI), an unanticipated reaction intermediate, methyl (Z)-2-(3-chloro-5-cyanophenoxy)-5-(3-(3-chloro-5-cyanophenoxy)-2-oxo-4-(trifluoromethyl)pyridin-1(2H)-yl)-5-ethoxy-3-(trifluoromethyl)pent-2-enoate (2), was isolated. Moreover, an unusual electrospray ionization (ESI)-induced fragmentation was observed for 2. Hence, efforts were made towards the understanding of the structure of 2, which was crucial for the understanding of the reaction mechanism. METHODS: The isolated impurity was fully characterized by liquid chromatography coupled with high-resolution tandem mass spectrometry (LC/HRMS/MS), hydrogen/deuterium (H/D) exchange, and an ensemble of two-dimensional nuclear magnetic resonance (2D-NMR) techniques. Density functional theory (DFT) calculations were also conducted. RESULTS: An unusual ESI-induced fragmentation was observed for intermediate 2, giving an ion for half of the molecule in the positive ion mode, with the other half of the molecule affording an ion in the negative ion mode. CONCLUSIONS: To the best of our knowledge, this unique ESI-induced fragmentation has not been previously reported in the literature. The underlying mechanism was explored and is supported by DFT calculations, which could greatly help the structural characterization of unknown impurities with similar structural features using ESI-MS in the future. Copyright © 2017 John Wiley & Sons, Ltd.