Structural and Biochemical Basis for Development of Diketo Acid Inhibitors Targeting the Cap-Snatching Endonuclease of the Ebinur Lake Virus (Order: Bunyavirales).

The Bunyavirales contain many important human pathogens that lack an antiviral therapy. The cap-snatching endonuclease (EN) of segmented negative-strand RNA viruses is an attractive target for broad-spectrum antivirals due to its essential role in initiating viral transcription. L-742,001, a previously reported diketo acid inhibitor against influenza virus EN, demonstrated potent EN inhibition and antiviral activity on various bunyaviruses. However, the precise inhibitory mechanism of the compound is still poorly understood. We recently characterized a highly active EN from Ebinur Lake virus (EBIV), a newly identified member of the Orthobunyavirus genus, and obtained its high-resolution structures, paving the way for structure-guided inhibitor development. Here, nine L-742,001 derivatives were designed and synthesized de novo, and their structure-activity relationship with EBIV EN was studied. In vitro biochemical data showed that the compounds inhibited the EBIV EN activity with different levels and could be divided into three categories. Five representative compounds were selected for further cell-based antiviral assay, and the results largely agreed with those of the EN assays. Furthermore, the precise binding modes of L-742,001 and its derivatives in EN were revealed by determining the high-resolution crystal structures of EN-inhibitor complexes, which suggested that the p-chlorobenzene is essential for the inhibitory activity and the flexible phenyl has the greatest exploration potential. This study provides an important basis for the structure-based design and optimization of inhibitors targeting EN of segmented negative-strand RNA viruses. IMPORTANCE The Bunyavirales contain many important human pathogens such as Crimean-Congo hemorrhagic fever virus and Lassa virus that pose serious threats to public health; however, currently there are no specific antiviral drugs against these viruses. The diketo acid inhibitor L-742,001 is a potential drug as it inactivates the cap-snatching endonuclease (EN) encoded by bunyaviruses. Here, we designed and synthesized nine L-742,001 derivatives and assessed the structure-activity relationship using EN of the newly identified Ebinur Lake virus (EBIV) as a research model. Our results revealed that the p-chlorobenzene of this broad-spectrum EN inhibitor is crucial for the inhibitory activity and the flexible phenyl "arm" has the best potential for further optimization. As cap-snatching ENs are present not only in bunyaviruses but also in influenza viruses, our data provide important guidelines for the development of novel and more potent diketo acid-based antiviral drugs against those viruses.

Boronic Acids as Prospective Inhibitors of Metallo-ß-Lactamases: Efficient Chemical Reaction in the Enzymatic Active Site Revealed by Molecular Modeling.

Boronic acids are prospective compounds in inhibition of metallo-ß-lactamases as they form covalent adducts with the catalytic hydroxide anion in the enzymatic active site upon binding. We compare this chemical reaction in the active site of the New Delhi metallo-ß-lactamase (NDM-1) with the hydrolysis of the antibacterial drug imipenem. The nucleophilic attack occurs with the energy barrier of 14 kcal/mol for imipenem and simultaneously upon binding a boronic acid inhibitor. A boron atom of an inhibitor exhibits stronger electrophilic properties than the carbonyl carbon atom of imipenem in a solution that is quantified by atomic Fukui indices. Upon forming the prereaction complex between NDM-1 and inhibitor, the lone electron pair of the nucleophile interacts with the vacant p-orbital of boron that facilitates the chemical reaction. We analyze a set of boronic acid compounds with the benzo[b]thiophene core complexed with the NDM-1 and propose quantitative structure-sroperty relationship (QSPR) equations that can predict IC50 values from the calculated descriptors of electron density. These relations are applied to classify other boronic acids with the same core found in the database of chemical compounds, PubChem, and proposed ourselves. We demonstrate that the IC50 values for all considered benzo[b]thiophene-containing boronic acid inhibitors are 30-70 µM.

Lipoteichoic Acid Biosynthesis Inhibitors as Potent Inhibitors of S. aureus and E. faecalis Growth and Biofilm Formation.

Methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecalis (VRE) have been deemed as serious threats by the CDC. Many chronic MRSA and VRE infections are due to biofilm formation. Biofilm are considered to be between 10-10,000 times more resistant to antibiotics, and therefore new chemical entities that inhibit and/or eradicate biofilm formation are needed. Teichoic acids, such as lipoteichoic acids (LTAs) and wall teichoic acids (WTAs), play pivotal roles in Gram-positive bacteria's ability to grow, replicate, and form biofilms, making the inhibition of these teichoic acids a promising approach to fight infections by biofilm forming bacteria. Here, we describe the potent biofilm inhibition activity against MRSA and VRE biofilms by two LTA biosynthesis inhibitors HSGN-94 and HSGN-189 with MBICs as low as 0.0625 µg/mL against MRSA biofilms and 0.5 µg/mL against VRE biofilms. Additionally, both HSGN-94 and HSGN-189 were shown to potently synergize with the WTA inhibitor Tunicamycin in inhibiting MRSA and VRE biofilm formation.

Vonoprazan: A Novel and Potent Alternative in the Treatment of Acid-Related Diseases.

Although proton pump inhibitors (PPIs) have been used widely, acid-related diseases are still associated with a huge burden on the health care system. Recently, the efficacy and safety of a new acid suppressant named vonoprazan in the treatment of acid-related diseases have been evaluated by a series of studies. As a novel potassium-competitive acid blocker, vonoprazan may provide reversible acid suppression by preventing K+ from binding to gastric H+/K+-ATPase. It has been clinically used for the short-term treatment of gastroesophageal reflux disease (GERD), peptic ulcer disease and Helicobacter pylori (H. pylori) infection in Japan. The healing rate of GERD and gastric ulcers by vonoprazan is more than 95 and 90%, respectively; also, it is effective in curing PPI-resistant GERD. It increases H. pylori eradication rate to more than 88% as part of both first-line and second-line therapy. It is also effective in the eradication of clarithromycin-resistant H. pylori strains. All of these short-term studies show vonoprazan is safe and well-tolerated. As a safe and effective acid inhibitor, vonoprazan might be a novel alternative in the treatment of acid-related diseases.

The Structure of the Zika Virus Protease, NS2B/NS3pro.

In this chapter, we first briefly review the history of Zika virus (ZIKV) over the past 70 years since its discovery. We then focus on the ZIKV NS2B/NS3 protease, a major potential target for anti-ZIKV therapeutics. We describe the structure of the complex between Zika virus NS2B-NS3 protease and a peptide boronic-acid inhibitor that we determined in early 2016. We then review other structural studies on the Zika virus protease, which have been published in the past few months. Three different types of construct for the protease have been investigated by X-ray crystallography and NMR spectroscopy: the traditional "linked" construct comprising the NS2B cofactor, a Gly4SerGly4 linker, and the NS3pro chain; a construct where the linker has been replaced by Lys-Thr-Gly-Lys-Arg, which leads to autocleavage; and the bimolecular "unlinked" protease consisting of the NS2B cofactor segment and NS3pro. In complex with an inhibitor, the protease adopts a closed, "active" conformation with the NS2B chain wrapped around the NS3pro and contributing to the S2 pocket. In the ligand-free state, the Gly4SerGly4-linked enzyme adopts an open or relaxed conformation, with the C-terminal half of the NS2B cofactor highly flexible and disordered. Very surprisingly, however, the "unlinked", bimolecular protease has been reported to adopt the closed conformation in the crystal, even though, apparently, no peptide was bound to the substrate-binding site. The Gly4SerGly4-linked enzyme has been used successfully in drug discovery efforts.

Biomolecular interactions between the antibacterial ceftolozane and the human inflammatory disease target ADAM17: a drug repurposing study.

Inhibition of a disintegrin and metalloproteinase-17 (ADAM17), a metzincin, is proposed as a novel therapeutic strategy to suppress overproduction of the proinflammatory cytokine TNF-α in rheumatoid arthritis and inflammatory bowel disease. Existing ADAM17 inhibitors generate toxic metabolites in-vivo or haven't progressed in clinical trials. Previous studies suggest that ligands which bind to ADAM17 active site by interacting with the Zn ion and L-shaped hydrophobic S1'- and S3'-pockets and forming favorable hydrogen bonds could act as potential ADAM17 inhibitors. Here, we investigated whether the FDA-approved anti-bacterial drug ceftolozane, a cephalosporin containing aromatic groups and carboxyl groups as probable zinc binding groups (ZBGs), forms non-covalent interactions resulting in its binding in the active site of ADAM17. In this study, the density functional theory (DFT), molecular docking and molecular dynamics calculations with the catalytic chain of ADAM17 show that carboxyl group of ceftolozane acts as moderate ZBG, and its extended geometry forms hydrogen bonds and hydrophobic interactions resulting in a binding affinity comparable to the co-crystallized known ADAM17 inhibitor. The favorable binding interactions identified here suggest the potential of ceftolozane to modulate ADAM17 activity in inflammatory diseases. ADAM17 cleaves and releases epidermal growth factor (EGF) ligands from the cell surface. The shed EGF ligands then bind to the EGF receptors to drive embryonic development. Therefore, our findings also suggest that use of ceftolozane during pregnancy may inhibit ADAM17-mediated shedding of EGF and thus increase the risk of birth defects in humans.Communicated by Ramaswamy H. Sarma.

Risk factors and outcomes of Clostridium difficile infection in hospitalized patients.

BACKGROUND: The aim of this study was to identify risk factors for Clostridium difficile infection (CDI) and its attributable mortality and to propose methods to prevent CDI and improve patients' outcomes. METHODS: CDI was defined as diarrheal patients with stool samples that were positive for C. difficile toxin. Clinical presentations of all patients with CDI and two times as many age- and sex-matched culture-negative controls at the Chang Gung Memorial Hospital in 2014 were identified and compared by multivariate, nonparametric, and Kaplan-Meier survival analysis. RESULTS: There were no significant differences in ages, sex, or Charlson comorbidity indexes between the CDI group (n = 42) and the control group (n = 86). The multivariate analysis indicated that underlying peptic ulcer disease and previous use of gastric acid inhibitors or third-generation cephalosporins for at least 3 days were significantly more common in patients with CDI than in the controls. Charlson scores were associated with mortality due to CDI. Recommended treatment using oral vancomycin to treat patients with Charlson score ≥ 5 and oral metronidazole or vancomycin to treat those with moderate underlying disease (Charlson score ≥ 2 and ≤ 5) significantly increased survival in these patients (p = 0.001). CONCLUSIONS: Oral vancomycin given to patients with high Charlson scores and oral metronidazole or vancomycin to patients with moderate Charlson scores decreased mortality due to CDI. Restricting the use of third-generation cephalosporins and gastric acid inhibitors is recommended to prevent CDI in hospitalized patients.

Delamanid: A new armor in combating drug-resistant tuberculosis.

Intense search has been made in the discovery of newer anti-TB drugs to tackle the issues such as drug resistance, HIV co-infection and risk of drug-drug interactions in the management of TB. Delamanid, a newer mycobacterial cell wall synthesis inhibitor, received a conditional approval from European medicines agency (EMA) for the treatment of MDR-TB. Preclinical and clinical studies have shown that delamanid has high potency, least risk for drug-drug interactions and better tolerability.

A hIAPP-derived all-D-amino-acid inhibits hIAPP fibrillation efficiently at membrane surface by targeting α-helical oligomeric intermediates.

A variety of peptides and peptide derivatives have been constructed using the "ß-sheet core segment" of amyloid proteins as inhibitors of amyloidogenic fibrillation. A novel all-D-amino-acid from hIAPP ß-sheet core segment (hIAPP 22-27) is demonstrated to inhibit hIAPP fibril formation efficiently both at the phospholipid membrane and in bulk solution. The inhibitor terminates hIAPP aggregation to the α-helical oligomeric intermediates at the membrane surface, whereas it stops the aggregation at the stage of ß-sheet oligomeric intermediates in bulk solution. This is the first evidence that the inhibition mechanism of the inhibitor at membrane surface is significantly different from that in bulk solution.

Pralatrexate alone or in combination with bexarotene: long-term tolerability in relapsed/refractory mycosis fungoides.

BACKGROUND: This study aimed to assess the long-term tolerability of pralatrexate alone or in combination with oral bexarotene for relapsed or refractory mycosis fungoides (MF). PATIENTS AND METHODS: Patients with MF in this report were participants in 1 of 2 multicenter trials. During the dose-ranging phase I/II study, participants were treated with pralatrexate alone for 3 of 4 weeks. During a second phase I/II dose-ranging combination trial, participants were treated with pralatrexate at 15 mg/m(2)/wk for 3 of 4 weeks combined with 150 to 300 mg/m(2) of daily oral bexarotene. RESULTS: Twenty-six patients were enrolled at our center, including 12 receiving pralatrexate and 14 receiving pralatrexate plus bexarotene. Four of 12 patients (33%) treated with pralatrexate alone responded. Of 14 patients treated with bexarotene plus pralatrexate, 7 (50%) responded. Ten participants, with a median age of 71 years (range, 41-82 years), received more than 9 cycles of pralatrexate, including 3 receiving pralatrexate and 7 receiving combination therapy. Median time to response was 15.75 weeks (range, 4-24 weeks), and the median duration of response was 26.75 weeks (range, 8.5-49.5 weeks). The most common adverse event (AE) was mucositis in 8 (80%) patients. Other AEs of any grade included arthralgias (n = 1), headache (n = 1), neutropenia (n = 5), and skin necrosis (n = 2). Two patients initially had lower leg tumors that responded to therapy, leaving residual chronic leg ulcers. CONCLUSION: Pralatrexate alone or in combination with low-dose oral bexarotene is well tolerated and capable of providing long-term responses in patients of advanced age with advanced-stage MF.

A clinical analysis of intensive gastrointestinal lavage method for treatment of 100 severe cases with stress ulcer bleeding in neurosurgery department / 中国中西医结合急救杂志

Objective To discuss the intensive gastric cavitary lavage method in treatment of stress ulcer bleeding(SUB).Methods The therapeutic effects of 100 patients with severe SUB in intensive care unit (ICU)from July 2005 to February 2008 treated with intensive gastric cavity lavage method were observed and analyzed.Results In these 100 cases,the bleeding was stopped successfully in 97 cases,the therapeutic effective rate being 97%;there were 17 cases died of complicated pulmonary infection(17%)and 3 cases of bleeding(3%),the total fatality rate being 20%.All the other cases were cured.Conclusion Intensive gastrointestinal lavage therapy is an effective method as acid inhibitor for treatment of SUB,but it can decrease the inhabitant flora in the stomach to pass through gastric-pulmonary route,thus the incidence of pulmonary infection was reduced.

The Diagnostic Strategy in Gastroesophageal Reflux Disease and the Role of Acid Inhibition Therapeutic Test / 中国医师杂志

14,and/or pH4% of the total time at 24-hour esophageal pH metry were considered to be pathologic acid reflux(GERD).All patients underwent a diagnostic test by receiving lansoprazloe 30mg or placebo twice daily for 7 days through a cross-over design.A symptom reduction of over 75% was considered positive.Results 18(17%) cases of esophagitis were found in 104 patients by endoscopy.83(80%) patients experienced a typical symptom hearburn.87(84%) patients showed pathologic acid reflux detected from pH metry and they were considered to be GERD.85(82%) of 104 patients had positive result in therapeutic test.There was a significant correlation between both the diagnosis obtained from a trial of lansoprazole and the diagnosis obtained from pH metry(P

EFFECT OF ACID INHIBITOR ON THE FUNCTION AND ULTRASTRUCTURE OF GASTRIC PARIETAL CELLS IN RATS UNDER STRESS / 解放军医学杂志

The aim of this study was to demonstrate the effect of acid inhibitor on the function and ultrastructure of gastric parietal cells in rats under immersion and restraint stress. Thirty two male SD rats were randomly divided into control, stress, omeprazole and cimetidine treated groups. The gastric mucosal ulcer index (UI) and gastric juice pH were measured. The ultrastructural changes of parietal cells were observed by transmission electronic microscopy (TEM). The UI was decreased and the pH value was increased obviously in omeprazole and cimetidine treated groups compared with stress group ( P