Validation of Aztreonam-Avibactam Susceptibility Testing Using Digitally Dispensed Custom Panels

ABSTRACT Aztreonam-avibactam is a combination antimicrobial agent with activity against carbapenemase-producing Enterobacteriaceae (CPE) with metallo-β-lactamases (MβLs). Although aztreonam-avibactam is not yet approved by the U.S. Food and Drug Administration (FDA), clinicians can administer this combination by using two FDA-approved drugs: aztreonam and ceftazidime-avibactam. This combination of drugs is recommended by multiple experts for treatment of serious infections caused by MβL-producing CPE. At present, in vitro antimicrobial susceptibility testing (AST) of aztreonam-avibactam is not commercially available; thus, most clinicians receive no laboratory-based guidance that can support consideration of aztreonam-avibactam for serious CPE infections. Here, we report our internal validation for aztreonam-avibactam AST by reference broth microdilution (BMD) according to Clinical and Laboratory Standards Institute (CLSI) guidelines. The validation was performed using custom frozen reference BMD panels prepared in-house at the Centers for Disease Control and Prevention (CDC). In addition, we took this opportunity to evaluate a new panel-making method using a digital dispenser, the Hewlett Packard (HP) D300e. Our studies demonstrate that the performance characteristics of digitally dispensed panels were equivalent to those of conventionally prepared frozen reference BMD panels for a number of drugs, including aztreonam-avibactam. We found the HP D300e digital dispenser to be easy to use and to provide the capacity to prepare complex drug panels. Our findings will help other clinical and public health laboratories implement susceptibility testing for aztreonam-avibactam.

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Assessing the in vitro impact of ceftazidime on aztreonam/avibactam susceptibility testing for highly resistant MBL-producing Enterobacterales

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkaa531 ◽

2020 ◽

Author(s):

Amelia Bhatnagar ◽

Eric M Ransom ◽

María-José Machado ◽

Sandra Boyd ◽

Natashia Reese ◽

...

Keyword(s):

Susceptibility Testing ◽

Antagonistic Effect ◽

Vitro Activity ◽

In Vitro Activity ◽

Hewlett Packard ◽

Antibiotic Resistant ◽

Doubling Dilution ◽

Approved Drugs ◽

Fda Approved Drugs

Abstract Background Aztreonam/avibactam is a combination agent that shows promise in treating infections caused by highly antibiotic-resistant MBL-producing Enterobacterales. This combination can be achieved by combining two FDA-approved drugs: ceftazidime/avibactam and aztreonam. It is unknown whether ceftazidime in the combination ceftazidime/aztreonam/avibactam has a synergistic or antagonistic effect on the in vitro activity of aztreonam/avibactam by significantly increasing or decreasing the MIC. Objectives To determine whether increasing ceftazidime concentrations affect the MICs of aztreonam/avibactam alone. Methods A custom 8 × 8 chequerboard broth microdilution (BMD) panel was made using a digital dispenser (Hewlett-Packard, Corvallis, OR, USA). The panel included orthogonal 2-fold dilution series of aztreonam and ceftazidime ranging from 0.5 to 64 mg/L. Avibactam concentration was kept constant at 4 mg/L throughout the chequerboard. Thirty-seven Enterobacterales isolates from the CDC & FDA Antibiotic Resistance Isolate Bank or CDC’s internal collection with intermediate or resistant interpretations to aztreonam and ceftazidime/avibactam were included for testing. All isolates harboured at least one of the following MBL genes: blaIMP, blaNDM or blaVIM. Results Regardless of the concentration of ceftazidime, aztreonam/avibactam with ceftazidime MICs for all 37 isolates were within one 2-fold doubling dilution of the aztreonam/avibactam MIC. Conclusions Ceftazidime, in the combination ceftazidime/avibactam/aztreonam, did not affect the in vitro activity of aztreonam/avibactam in this sample of isolates. These findings can help assure clinical and public health laboratories that testing of aztreonam/avibactam by BMD can act as a reliable surrogate test when the combination of ceftazidime/avibactam and aztreonam is being considered for treatment of highly antibiotic-resistant MBL-producing Enterobacterales.

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Identification of 3-chymotrypsin like protease (3CLPro) inhibitors as potential anti-SARS-CoV-2 agents

Communications Biology ◽

10.1038/s42003-020-01577-x ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Vicky Mody ◽

Joanna Ho ◽

Savannah Wills ◽

Ahmed Mawri ◽

Latasha Lawson ◽

...

Keyword(s):

Inhibitory Effect ◽

Dynamics Simulation ◽

Simulation Studies ◽

Inhibitory Effects ◽

Major Threat ◽

Main Protease ◽

Approved Drugs ◽

Fda Approved Drugs ◽

Computational Molecular Modeling

AbstractEmerging outbreak of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection is a major threat to public health. The morbidity is increasing due to lack of SARS-CoV-2 specific drugs. Herein, we have identified potential drugs that target the 3-chymotrypsin like protease (3CLpro), the main protease that is pivotal for the replication of SARS-CoV-2. Computational molecular modeling was used to screen 3987 FDA approved drugs, and 47 drugs were selected to study their inhibitory effects on SARS-CoV-2 specific 3CLpro enzyme in vitro. Our results indicate that boceprevir, ombitasvir, paritaprevir, tipranavir, ivermectin, and micafungin exhibited inhibitory effect towards 3CLpro enzymatic activity. The 100 ns molecular dynamics simulation studies showed that ivermectin may require homodimeric form of 3CLpro enzyme for its inhibitory activity. In summary, these molecules could be useful to develop highly specific therapeutically viable drugs to inhibit the SARS-CoV-2 replication either alone or in combination with drugs specific for other SARS-CoV-2 viral targets.

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Targeting SARS-CoV-2 Polymerase with New Nucleoside Analogues

Molecules ◽

10.3390/molecules26113461 ◽

2021 ◽

Vol 26 (11) ◽

pp. 3461

Author(s):

Vasiliki Daikopoulou ◽

Panagiotis Apostolou ◽

Sofia Mourati ◽

Ioanna Vlachou ◽

Maria Gougousi ◽

...

Keyword(s):

Inhibitory Activity ◽

Drug Repositioning ◽

Nucleoside Analogues ◽

In Vitro Assays ◽

Rna Dependent Rna Polymerase ◽

Sugar Moiety ◽

The Family ◽

Approved Drugs ◽

Fda Approved Drugs

Despite the fact that COVID-19 vaccines are already available on the market, there have not been any effective FDA-approved drugs to treat this disease. There are several already known drugs that through drug repositioning have shown an inhibitory activity against SARS-CoV-2 RNA-dependent RNA polymerase. These drugs are included in the family of nucleoside analogues. In our efforts, we synthesized a group of new nucleoside analogues, which are modified at the sugar moiety that is replaced by a quinazoline entity. Different nucleobase derivatives are used in order to increase the inhibition. Five new nucleoside analogues were evaluated with in vitro assays for targeting polymerase of SARS-CoV-2.

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Identification of FDA-approved drugs as novel allosteric inhibitors of human executioner caspases

10.1101/356956 ◽

2018 ◽

Author(s):

R. N. V. Krishna Deepak ◽

Ahmad Abdullah ◽

Priti Talwar ◽

Hao Fan ◽

Palaniyandi Ravanan

Keyword(s):

Active Site ◽

Caspase 3 ◽

Specific Activity ◽

Special Importance ◽

Allosteric Inhibitors ◽

Dimerization Interface ◽

Executioner Caspases ◽

Approved Drugs ◽

Fda Approved Drugs

AbstractThe regulation of apoptosis is a tightly-coordinated process and caspases are its chief regulators. Of special importance are the executioner caspases, caspase-3/7, the activation of which irreversibly sets the cell on the path of death. Dysregulation of apoptosis, particularly an increased rate of cell death lies at the root of numerous human diseases. Although several peptide-based inhibitors targeting the homologous active site region of caspases have been developed, owing to their non-specific activity and poor pharmacological properties their use has largely been restricted. Thus, we sought to identify FDA-approved drugs that could be repurposed as novel allosteric inhibitors of caspase-3/7. In this study, we virtually screened a catalog of FDA-approved drugs targeting an allosteric pocket located at the dimerization interface of caspase-3/7. From among the top-scoring hits we short-listed five compounds for experimental validation. Our enzymatic assays using recombinant caspase-3 suggested that four out of the five drugs effectively inhibited caspase-3 enzymatic activity in vitro with IC50 values ranging ~10-55 μM. Structural analysis of the docking poses show the four compounds forming specific non-covalent interactions at the allosteric pocket suggesting that these molecules could disrupt the adjacently-located active site. In summary, we report the identification of four novel non-peptide allosteric inhibitors of caspase-3/7 from among FDA-approved drugs.

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Virtual screening as therapeutic strategy of COVID-19 targeting angiotensin-converting enzyme 2

Frontiers in Molecular Immunology ◽

10.25082/fmi.2021.01.002 ◽

2021 ◽

Vol 2 (1) ◽

pp. 16-27

Author(s):

Zahra Sharifinia ◽

◽

Samira Asadi ◽

Mahyar Irani ◽

Abdollah Allahverdi ◽

...

Keyword(s):

Virtual Screening ◽

Angiotensin Converting Enzyme ◽

Host Cells ◽

Spike Protein ◽

Potential Drug ◽

Converting Enzyme ◽

Angiotensin Converting Enzyme 2 ◽

Approved Drugs ◽

Fda Approved Drugs

Objective: The receptor-binding domain (RBD) of the S1 domain of the SARS-CoV- 2 Spike protein performs a key role in the interaction with Angiotensin-converting enzyme 2 (ACE2), leading to both subsequent S2 domain-mediated membrane fusion and incorporation of viral RNA in host cells. Methods: In this study, we investigated the inhibitor’s targeted compounds through existing human ACE2 drugs to use as a future viral invasion. 54 FDA approved drugs were selected to assess their binding affinity to the ACE2 receptor. The structurebased methods via computational ones have been used for virtual screening of the best drugs from the drug database. Key Findings: The ligands “Cinacalcet” and “Levomefolic acid” highaffinity scores can be a potential drug preventing Spike protein of SARS-CoV-2 and human ACE2 interaction. Levomefolic acid from vitamin B family was proved to be a potential drug as a spike protein inhibitor in previous clinical and computational studies. Besides that, in this study, the capability of Levomefolic acid to avoid ACE2 and Spike protein of SARS-CoV-2 interaction is indicated. Therefore, it is worth to consider this drug for more in vitro investigations as ACE2 and Spike protein inhibition candidate. Conclusion: The two Cinacalcet and Levomefolic acid are the two ligands that have highest energy binding for human ACE2 blocking among 54 FDA approved drugs.

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DDRE-28. IDENTIFICATION OF SYNERGISTIC DRUG COMBINATIONS FOR THE THERAPY OF IDHmut GLIOMA

Neuro-Oncology ◽

10.1093/neuonc/noab196.312 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi80-vi80

Author(s):

Rolf Warta ◽

Florian Stammler ◽

Andreas Unterberg ◽

Christel Herold-Mende

Keyword(s):

Single Agent ◽

Therapy Response ◽

Drug Combinations ◽

Drug Concentrations ◽

Brain Barrier Permeability ◽

Approved Drugs ◽

Fda Approved Drugs ◽

Biological Differences ◽

Synergistic Drug Combinations

Abstract OBJECTIVE Isocitrate Dehydrogenase (IDH) mutation in glioma results in a multitude of biological differences with consequences for survival and therapy response. Therefore, IDH mutated (IDHmut) and wildtype (IDHwt) tumors are regarded as separate entities with the need for adjusted therapy like the combination of procarbazine, CCNU and vincristine (PCV). However, as vincristine has often severe side effects like neuropathy new effective therapy options are required. Therefore, we searched for combinations of FDA-approved drugs which effectively inhibit the growth of IDHmut cells in vitro. METHODS We tested different drug combinations of a drug library consisting of 146 FDA-approved drugs on two established IDHmut GSC lines. Based on a previous single agent drug screen, six drugs were selected (Idarubicin, Ixazumib, Ponatinib, Neratinib, Romidepsin) to be combined with all 146 drugs of the library. Cell viability was assessed by the CellTiterGlo 3D assay (Promega) in 96 well plates, while Caspase-Glo 3/7 3D assay was used to measure induction of apoptosis. RESULTS Out of 1460 drug combinations tested altogether 21 synergistic drug combinations could be identified and validated. The combination with the highest blood-brain-barrier permeability score was further investigated. Finally, drug-concentrations elucidating the highest synergistic effect on proliferation was further studied in a 8-point dose-response matrix followed by validation in additional four IDHmut GSC lines. CONCLUSION This work can lay the foundation for future improvements of the therapy of patients suffering from LGGs.

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A platform utilizing Drosophila ovulation for nonhormonal contraceptive screening

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2026403118 ◽

2021 ◽

Vol 118 (28) ◽

pp. e2026403118

Author(s):

Kewa Jiang ◽

Jiyang Zhang ◽

Yuping Huang ◽

Yingzheng Wang ◽

Shuo Xiao ◽

...

Keyword(s):

Ex Vivo ◽

Unmet Need ◽

Dependent Manner ◽

Female Reproduction ◽

Contraceptive Agents ◽

Follicle Rupture ◽

Approved Drugs ◽

Fda Approved Drugs

A significant unmet need for new contraceptive options for both women and men remains due to side-effect profiles, medical concerns, and the inconvenience of many currently available contraceptive products. Unfortunately, the development of novel nonsteroidal female contraceptive medicine has been stalled in the last couple of decades due to the lack of effective screening platforms. Drosophila utilizes conserved signaling pathways for follicle rupture, a final step in ovulation that is essential for female reproduction. Therefore, we explored the potential to use Drosophila as a model to screen compounds that could inhibit follicle rupture and be nonsteroidal contraceptive candidates. Using our ex vivo follicle rupture assay, we screened 1,172 Food and Drug Administration (FDA)–approved drugs and identified six drugs that could inhibit Drosophila follicle rupture in a dose-dependent manner. In addition, we characterized the molecular actions of these drugs in the inhibition of adrenergic signaling and follicle rupture. Furthermore, we validated that three of the four drugs consistently inhibited mouse follicle rupture in vitro and that two of them did not affect progesterone production. Finally, we showed that chlorpromazine, one of the candidate drugs, can significantly inhibit mouse follicle rupture in vivo. Our work suggests that Drosophila ovulation is a valuable platform for identifying lead compounds for nonsteroidal contraceptive development and highlights the potential of these FDA-approved drugs as novel nonsteroidal contraceptive agents.

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Sulfisoxazole does not inhibit the secretion of small extracellular vesicles

10.1101/2020.03.15.988055 ◽

2020 ◽

Cited By ~ 1

Author(s):

Pamali Fonseka ◽

Sai V Chitti ◽

Rahul Sanwlani ◽

Suresh Mathivanan

Keyword(s):

Cancer Cells ◽

Extracellular Vesicles ◽

Breast Cancer Cells ◽

Drug Repurposing ◽

Tumor Burden ◽

Immunocompetent Mouse ◽

Approved Drugs ◽

Fda Approved Drugs

AbstractRecently, the study by Im et al. focused on blocking the release of extracellular vesicles (EVs) by cancer cells, as a strategy to block metastasis, by deploying a drug repurposing screen. Upon screening the library of FDA approved drugs in breast cancer cells in vitro, the authors reported the ability of the antibiotic Sulfisoxazole (SFX) in inhibiting EV biogenesis and secretion. SFX was also effective in reducing breast primary tumor burden and blocking metastasis in immunocompromised and immunocompetent mouse models. As we seek a compound to block EV biogenesis and secretion in our current in vivo studies, we intended to use SFX and hence performed in vitro characterization as the first step. However, treatment of two cancer cells with SFX did not reduce the amount of EVs as reported by the authors.

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Repurposing inhibitors for SARS-CoV2 main protease

10.21203/rs.3.rs-41342/v1 ◽

2020 ◽

Author(s):

Kumar Sharp

Keyword(s):

Viral Replication ◽

Active Sites ◽

Drug Repurposing ◽

Chemotherapeutic Drugs ◽

Short Term ◽

Main Protease ◽

Potential Binding ◽

Approved Drugs ◽

Fda Approved Drugs

Abstract SARS-CoV2 main protease is important for viral replication and one of the most potential targets for drug development in this current pandemic. Drug repurposing is a promising field to provide potential short-term acceptable therapy for management of coronavirus till a specific anti-viral for coronavirus is developed. In-silico drug repurposing screening is the current fastest way to repurpose drugs by targeting active sites in fraction of seconds. In this study, SARS-CoV2 main protease is being targeted by 1050 FDA-approved drugs to inhibit its activity thereby interfering with viral replication. Chemotherapeutic drugs and anti-retroviral drugs have shown potential binding as inhibitor. In-vitro and clinical trials required to establish final fact.

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Protoporphyrin IX and verteporfin prevent SARS-CoV-2 infection in vitro and in a mouse model expressing human ACE2

10.1101/2020.04.30.071290 ◽

2020 ◽

Cited By ~ 3

Author(s):

Chenjian Gu ◽

Yang Wu ◽

Huimin Guo ◽

Yuanfei Zhu ◽

Wei Xu ◽

...

Keyword(s):

Cytopathic Effect ◽

Antiviral Agents ◽

Protoporphyrin Ix ◽

Ring Structure ◽

Receptor Binding Domain ◽

Viral Receptor ◽

Margin Of Safety ◽

Approved Drugs ◽

Fda Approved Drugs

AbstractThe SARS-CoV-2 infection is spreading rapidly worldwide. Efficacious antiviral therapeutics against SARS-CoV-2 is urgently needed. Here, we discovered that protoporphyrin IX (PpIX) and verteporfin, two FDA-approved drugs, completely inhibited the cytopathic effect produced by SARS-CoV-2 infection at 1.25 μM and 0.31 μM respectively, and their EC50 values of reduction of viral RNA were at nanomolar concentrations. The selectivity indices of PpIX and verteporfin were 952.74 and 368.93, respectively, suggesting broad margin of safety. Importantly, PpIX and verteporfin prevented SARS-CoV-2 infection in mice adenovirally transduced with human ACE2. The compounds, sharing a porphyrin ring structure, were shown to bind viral receptor ACE2 and interfere with the interaction between ACE2 and the receptor-binding domain of viral S protein. Our study suggests that PpIX and verteporfin are potent antiviral agents against SARS-CoV-2 infection and sheds new light on developing novel chemoprophylaxis and chemotherapy against SARS-CoV-2.

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