scholarly journals Repurposing Drugs for Mayaro Virus: Identification of EIDD-1931, Favipiravir and Suramin as Mayaro Virus Inhibitors

2021 ◽  
Vol 9 (4) ◽  
pp. 734
Author(s):  
Lana Langendries ◽  
Rana Abdelnabi ◽  
Johan Neyts ◽  
Leen Delang
Keyword(s):  
South America ◽  
High Throughput Screening ◽  
Virus Identification ◽  
Antiviral Compounds ◽  
Infection Models ◽  
Animal Infection ◽  
Orally Bioavailable ◽  
Mayaro Virus ◽  
Cytopathogenic Effect

Despite the emerging threat of the Mayaro virus (MAYV) in Central and South-America, there are no licensed antivirals or vaccines available for this neglected mosquito-borne virus. Here, we optimized a robust antiviral assay based on the inhibition of the cytopathogenic effect that could be used for high-throughput screening to identify MAYV inhibitors. We first evaluated different cell lines and virus inputs to determine the best conditions for a reliable and reproducible antiviral assay. Next, we used this assay to evaluate a panel of antiviral compounds with known activity against other arboviruses. Only three drugs were identified as inhibitors of MAYV: β-D-N4-hydroxycytidine (EIDD-1931), favipiravir and suramin. The in vitro anti-MAYV activity of these antiviral compounds was further confirmed in a virus yield assay. These antivirals can therefore serve as reference compounds for future anti-MAYV compound testing. In addition, it is of interest to further explore the activity of EIDD-1931 and its orally bioavailable pro-drug molnupiravir in animal infection models to determine whether it offers promise for the treatment of MAYV infection.

scholarly journals Use of anInVitroPharmacodynamic Model To Derive a Moxifloxacin Regimen That Optimizes Kill ofYersinia pestisand Prevents Emergence of Resistance

2010 ◽  
Vol 55 (2) ◽  
pp. 822-830 ◽  
Author(s):  
A. Louie ◽  
H. S. Heine ◽  
B. VanScoy ◽  
A. Eichas ◽  
K. Files ◽  
...  
Keyword(s):  
Drug Exposure ◽  
Dose Range ◽  
Human Infection ◽  
Pharmacodynamic Model ◽  
Dose Fractionation ◽  
Time Curve ◽  
Infection Models ◽  
Animal Infection ◽  
Emergence Of Resistance

ABSTRACTYersinia pestis, the causative agent of bubonic, septicemic, and pneumonic plague, is classified as a CDC category A bioterrorism pathogen. Streptomycin and doxycycline are the “gold standards” for the treatment of plague. However, streptomycin is not available in many countries, andY. pestisisolates resistant to streptomycin and doxycycline occur naturally and have been generated in laboratories. Moxifloxacin is a fluoroquinolone antibiotic that demonstrates potent activity againstY. pestisinin vitroand animal infection models. However, the dose and frequency of administration of moxifloxacin that would be predicted to optimize treatment efficacy in humans while preventing the emergence of resistance are unknown. Therefore, dose range and dose fractionation studies for moxifloxacin were conducted forY. pestisin anin vitropharmacodynamic model in which the half-lives of moxifloxacin in human serum were simulated so as to identify the lowest drug exposure and the schedule of administration that are linked with killing ofY. pestisand with the suppression of resistance. In the dose range studies, simulated moxifloxacin regimens of ≥175 mg/day killed drug-susceptible bacteria without resistance amplification. Dose fractionation studies demonstrated that the AUC (area under the concentration-time curve)/MIC ratio predicted kill of drug-susceptibleY. pestis, while theCmax(maximum concentration of the drug in serum)/MIC ratio was linked to resistance prevention. Monte Carlo simulations predicted that moxifloxacin at 400 mg/day would successfully treat human infection due toY. pestisin 99.8% of subjects and would prevent resistance amplification. We conclude that in anin vitropharmacodynamic model, the clinically prescribed moxifloxacin regimen of 400 mg/day is predicted to be highly effective for the treatment ofY. pestisinfections in humans. Studies of moxifloxacin in animal models of plague are warranted.

scholarly journals Phosphate flow in the chemotactic response system of Helicobacter pylori

Microbiology ◽  
2005 ◽  
Vol 151 (10) ◽  
pp. 3299-3311 ◽  
Author(s):  
María-Antonieta Jiménez-Pearson ◽  
Isabel Delany ◽  
Vincenzo Scarlato ◽  
Dagmar Beier
Keyword(s):  
Helicobacter Pylori ◽  
Response Regulator ◽  
Chemotactic Response ◽  
Receiver Domain ◽  
Regulator Protein ◽  
Infection Models ◽  
Animal Infection ◽  
H Pylori ◽  
Response Regulator Protein

It is well established that motility is an essential virulence trait of the human gastric pathogen Helicobacter pylori. Accordingly, chemotaxis contributes to the ability of H. pylori to colonize animal infection models. Chemotactic signal transduction in H. pylori differs from the enterobacterial paradigm in several respects. In addition to a separate CheY response regulator protein (CheY1), H. pylori contains a CheY-like receiver domain (CheY2) which is C-terminally fused to the histidine kinase CheA. Furthermore, the genome of H. pylori encodes three CheV proteins consisting of an N-terminal CheW-like domain and a C-terminal receiver domain, while there are no orthologues of the chemotaxis genes cheB, cheR and cheZ. To obtain insight into the mechanisms controlling the chemotactic response of H. pylori, we investigated the phosphotransfer reactions between the purified two-component signalling modules in vitro. We demonstrate that both CheY1 and CheY2 are phosphorylated by CheA∼P and that the three CheV proteins mediate the dephosphorylation of CheA∼P, but with a clearly reduced efficiency as compared to CheY1 and CheY2. Furthermore, our data indicate retrophosphorylation of CheAY2 by CheY1∼P, suggesting a role of CheY2 as a phosphate sink to modulate the half-life of CheY1∼P.

scholarly journals A simple, fast and reliable scan-based technique as a novel approach to quantify intracellular bacteria

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Meysam Sarshar ◽  
Daniela Scribano ◽  
Giulia Tranquilli ◽  
Marisa Di Pietro ◽  
Simone Filardo ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
High Speed ◽  
Low Cost ◽  
Shigella Flexneri ◽  
Intracellular Bacteria ◽  
Fluorescence Signal ◽  
Chronic Infections ◽  
Novel Approach ◽  
Infection Models

Abstract Background Quantification of intracellular bacteria is fundamental in many areas of cellular and clinical microbiology to study acute and chronic infections. Therefore, rapid, accurate and low-cost methods represent valuable tools in determining bacterial ability to persist and proliferate within eukaryotic cells. Results Herein, we present the first application of the immunofluorescence In-Cell Western (ICW) assay aimed at quantifying intracellular bacteria in in vitro infection models. The performance of this new approach was evaluated in cell culture infection models using three microorganisms with different lifestyles. Two facultative intracellular bacteria, the fast-growing Shigella flexneri and a persistent strain of Escherichia coli, as well as the obligate intracellular bacterium Chlamydia trachomatis were chosen as bacterial models. The ICW assay was performed in parallel with conventional quantification methods, i.e. colony forming units (CFUs) and inclusion forming units (IFUs). The fluorescence signal intensity values from the ICW assay were highly correlated to CFU/IFUs counting and showed coefficients of determination (R2), ranging from 0,92 to 0,99. Conclusions The ICW assay offers several advantages including sensitivity, reproducibility, high speed, operator-independent data acquisition and overtime stability of fluorescence signals. All these features, together with the simplicity in performance, make this assay particularly suitable for high-throughput screening and diagnostic approaches.

scholarly journals Evaluation of Possible Correlations between Antifungal Susceptibilities of Filamentous Fungi In Vitro and Antifungal Treatment Outcomes in Animal Infection Models

1998 ◽  
Vol 42 (2) ◽  
pp. 282-288 ◽  
Author(s):  
F. C. Odds ◽  
F. Van Gerven ◽  
A. Espinel-Ingroff ◽  
M. S. Bartlett ◽  
M. A. Ghannoum ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Filamentous Fungi ◽  
Guinea Pigs ◽  
Survival Times ◽  
Pseudallescheria Boydii ◽  
Infection Models ◽  
Animal Infection ◽  
Effective Doses

ABSTRACT Nine isolates of filamentous fungi previously tested in 11 different laboratories for their susceptibilities to amphotericin B and itraconazole in vitro were injected intravenously into mice and guinea pigs, and responses to treatment with both agents were studied. The experiments were done in a single laboratory. Mean survival times, the percentages of animals surviving 12 days after infection, and culture results for samples of deep organs obtained postmortem were used as markers of antifungal efficacy. Because of variations in organism pathogenicity, interpretable test systems in vivo could not be established for Fusarium spp. in mice or guinea pigs or forPseudallescheria boydii in mice, even with the use of immunosuppressive pretreatments. Among the infections that could be evaluated, some degree of response to the corresponding treatment in vivo was seen in animals infected with each of two Rhizopus arrhizus isolates susceptible to amphotericin B at <0.5 μg/ml and Aspergillus spp. isolates susceptible to itraconazole at <1.0 μg/ml. Conversely, no responses were apparent with infecting strains for which MICs were ≥2 μg/ml (amphotericin B) or ≥1 μg/ml (itraconazole). However, the limitations of the intravenous challenge systems studied mean that no firm conclusion relating MICs in vitro to the lowest effective doses in vivo could be drawn.

scholarly journals Identifying SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of Nsp13 Helicase

2021 ◽  
Author(s):  
Rupert Beale ◽  
Agustina P Bertolin ◽  
Annabel Borg ◽  
Berta Canal ◽  
John FX Diffley ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Drug Targets ◽  
Global Public Health ◽  
Chemical Library ◽  
Enzymatic Assays ◽  
Antiviral Compounds ◽  
Public Health Challenge ◽  
Related Compounds ◽  
Potential Drug Targets

The coronavirus disease 2019 (COVID-19) pandemic, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a global public health challenge. While the efficacy of vaccines against emerging and future virus variants remains unclear, there is a need for therapeutics. Repurposing existing drugs represents a promising and potentially rapid opportunity to find novel antivirals against SARS-CoV-2. The virus encodes at least nine enzymatic activities that are potential drug targets. Here we have expressed, purified and developed enzymatic assays for SARS-CoV-2 nsp13 helicase, a viral replication protein that is essential for the coronavirus life cycle. We screened a custom chemical library of over 5000 previously characterised pharmaceuticals for nsp13 inhibitors using a FRET-based high-throughput screening (HTS) approach. From this, we have identified FPA-124 and several suramin-related compounds as novel inhibitors of nsp13 helicase activity in vitro. We describe the efficacy of these drugs using assays we developed to monitor SARS-CoV-2 growth in Vero E6 cells.

scholarly journals Pharmacokinetics and pharmacodynamics utilizing unbound target tissue exposure as part of a disposition-based rationale for lead optimization of benzoxaboroles in the treatment of Stage 2 Human African Trypanosomiasis

Parasitology ◽  
2013 ◽  
Vol 141 (1) ◽  
pp. 104-118 ◽  
Author(s):  
STEPHEN WRING ◽  
ERIC GAUKEL ◽  
BAKELA NARE ◽  
ROBERT JACOBS ◽  
BETH BEAUDET ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Human African Trypanosomiasis ◽  
Oral Treatment ◽  
Target Tissue ◽  
African Trypanosomiasis ◽  
Infection Models ◽  
Animal Infection ◽  
Dosing Regimens ◽  
Time Kill

SUMMARYThis review presents a progression strategy for the discovery of new anti-parasitic drugs that uses in vitro susceptibility, time-kill and reversibility measures to define the therapeutically relevant exposure required in target tissues of animal infection models. The strategy is exemplified by the discovery of SCYX-7158 as a potential oral treatment for stage 2 (CNS) Human African Trypanosomiasis (HAT). A critique of current treatments for stage 2 HAT is included to provide context for the challenges of achieving target tissue disposition and the need for establishing pharmacokinetic–pharmacodynamic (PK–PD) measures early in the discovery paradigm. The strategy comprises 3 stages. Initially, compounds demonstrating promising in vitro activity and selectivity for the target organism over mammalian cells are advanced to in vitro metabolic stability, barrier permeability and tissue binding assays to establish that they will likely achieve and maintain therapeutic concentrations during in-life efficacy studies. Secondly, in vitro time-kill and reversibility kinetics are employed to correlate exposure (based on unbound concentrations) with in vitro activity, and to identify pharmacodynamic measures that would best predict efficacy. Lastly, this information is used to design dosing regimens for pivotal pharmacokinetic–pharmacodyamic studies in animal infection models.

Discovery of small molecule guanylyl cyclase A receptor positive allosteric modulators

2021 ◽  
Vol 118 (52) ◽  
pp. e2109386118
Author(s):  
S. Jeson Sangaralingham ◽  
Kanupriya Whig ◽  
Satyamaheshwar Peddibhotla ◽  
R. Jason Kirby ◽  
Hampton E. Sessions ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Small Molecule ◽  
Guanylyl Cyclase ◽  
High Throughput Screening ◽  
Metabolic Diseases ◽  
Ex Vivo ◽  
Guanosine Monophosphate ◽  
Cardiomyocyte Hypertrophy ◽  
Orally Bioavailable

The particulate guanylyl cyclase A receptor (GC-A), via activation by its endogenous ligands atrial natriuretic peptide (ANP) and b-type natriuretic peptide (BNP), possesses beneficial biological properties such as blood pressure regulation, natriuresis, suppression of adverse remodeling, inhibition of the renin-angiotensin-aldosterone system, and favorable metabolic actions through the generation of its second messenger cyclic guanosine monophosphate (cGMP). Thus, the GC-A represents an important molecular therapeutic target for cardiovascular disease and its associated risk factors. However, a small molecule that is orally bioavailable and directly targets the GC-A to potentiate cGMP has yet to be discovered. Here, we performed a cell-based high-throughput screening campaign of the NIH Molecular Libraries Small Molecule Repository, and we successfully identified small molecule GC-A positive allosteric modulator (PAM) scaffolds. Further medicinal chemistry structure–activity relationship efforts of the lead scaffold resulted in the development of a GC-A PAM, MCUF-651, which enhanced ANP-mediated cGMP generation in human cardiac, renal, and fat cells and inhibited cardiomyocyte hypertrophy in vitro. Further, binding analysis confirmed MCUF-651 binds to GC-A and selectively enhances the binding of ANP to GC-A. Moreover, MCUF-651 is orally bioavailable in mice and enhances the ability of endogenous ANP and BNP, found in the plasma of normal subjects and patients with hypertension or heart failure, to generate GC-A–mediated cGMP ex vivo. In this work, we report the discovery and development of an oral, small molecule GC-A PAM that holds great potential as a therapeutic for cardiovascular, renal, and metabolic diseases.

First Fluorescent Acetylspermidine Deacetylation Assay for HDAC10 Identifies Inhibitors of Neuroblastoma Cell Colony Growth That Increase Lysosome Accumulation

2020 ◽  
Author(s):  
Daniel Herp ◽  
Johannes Ridinger ◽  
Dina Robaa ◽  
Stephen A. Shinsky ◽  
Karin Schmidtkunz ◽  
...  
Keyword(s):  
Histone Deacetylases ◽  
High Throughput Screening ◽  
Neuroblastoma Cell ◽  
Hdac Inhibitors ◽  
Anticancer Therapy ◽  
Colony Growth ◽  
Autophagic Flux ◽  
Enzymatic Conversion ◽  
Lysine Deacetylase

Histone deacetylases (HDACs) are important epigenetic regulators involved in many diseases, esp. cancer. First HDAC inhibitors have been approved for anticancer therapy and many are in clinical trials. Among the 11 zinc-dependent HDACs, HDAC10 has received relatively little attention by drug discovery campaigns, despite its involvement e.g. in the pathogenesis of neuroblastoma. This is due in part to a lack of robust enzymatic conversion assays. In contrast to the protein lysine deacetylase and deacylase activity of the other HDAC subtypes, it has recently been shown that HDAC10 has strong preferences for deacetylation of oligoamine substrates like spermine or spermidine. Hence, it also termed a polyamine deacetylase (PDAC). Here, we present the first fluorescent enzymatic conversion assay for HDAC10 using an aminocoumarin labelled acetyl spermidine derivative to measure its PDAC activity, which is suitable for high-throughput screening. Using this assay, we identified potent inhibitors of HDAC10 mediated spermidine deacetylation in-vitro. Among those are potent inhibitors of neuroblastoma colony growth in culture that show accumulation of lysosomes, implicating disturbance of autophagic flux.

3D Culture Modelling: An Emerging Approach for Translational Cancer Research in Sarcomas

2020 ◽  
Vol 27 (29) ◽  
pp. 4778-4788 ◽  
Author(s):  
Victoria Heredia-Soto ◽  
Andrés Redondo ◽  
José Juan Pozo Kreilinger ◽  
Virginia Martínez-Marín ◽  
Alberto Berjón ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Cancer Biology ◽  
Soft Tissues ◽  
Three Dimensional ◽  
3D Culture ◽  
Resistance Mechanisms ◽  
In Vitro Models ◽  
Tumour Spheroids ◽  
Current Therapies

Sarcomas are tumours of mesenchymal origin, which can arise in bone or soft tissues. They are rare but frequently quite aggressive and with a poor outcome. New approaches are needed to characterise these tumours and their resistance mechanisms to current therapies, responsible for tumour recurrence and treatment failure. This review is focused on the potential of three-dimensional (3D) in vitro models, including multicellular tumour spheroids (MCTS) and organoids, and the latest data about their utility for the study on important properties for tumour development. The use of spheroids as a particularly valuable alternative for compound high throughput screening (HTS) in different areas of cancer biology is also discussed, which enables the identification of new therapeutic opportunities in commonly resistant tumours.

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