Doxycycline Inhibition of a Pseudotyped Virus Transduction Does Not Translate to Inhibition of SARS-CoV-2 Infectivity

The rapid spread of the pandemic caused by the SARS-CoV-2 virus has created an unusual situation, with rapid searches for compounds to interfere with the biological processes exploited by the virus. Doxycycline, with its pleiotropic effects, including anti-viral activity, has been proposed as a therapeutic candidate for COVID-19 and about twenty clinical trials have started since the beginning of the pandemic. To gain information on the activity of doxycycline against SARS-CoV-2 infection and clarify some of the conflicting clinical data published, we designed in vitro binding tests and infection studies with a pseudotyped virus expressing the spike protein, as well as a clinically isolated SARS-CoV-2 strain. Doxycycline inhibited the transduction of the pseudotyped virus in Vero E6 and HEK-293 T cells stably expressing human receptor angiotensin-converting enzyme 2 but did not affect the entry and replication of SARS-CoV-2. Although this conclusion is apparently disappointing, it is paradigmatic of an experimental approach aimed at developing an integrated multidisciplinary platform which can shed light on the mechanisms of action of potential anti-COVID-19 compounds. To avoid wasting precious time and resources, we believe very stringent experimental criteria are needed in the preclinical phase, including infectivity studies with clinically isolated SARS-CoV-2, before moving on to (futile) clinical trials.

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Doxycycline inhibition of a pseudotyped virus transduction does not translate to inhibition of SARS-CoV-2 infectivity

10.1101/2021.07.30.454436 ◽

2021 ◽

Author(s):

Luisa Diomede ◽

Sara Baroni ◽

Ada De Luigi ◽

Arianna Piotti ◽

Jacopo Lucchetti ◽

...

Keyword(s):

Clinical Trials ◽

Experimental Approach ◽

Pseudotyped Virus ◽

Biological Processes ◽

Hek 293 ◽

Angiotensin Converting Enzyme 2 ◽

Preclinical Phase ◽

Vitro Binding ◽

Human Receptor

The pandemic caused by the SARS-CoV-2 has created the need of compounds able to interfere with the biological processes exploited by the virus. Doxycycline, with its pleiotropic effects, including anti-viral activity, has been proposed as a therapeutic candidate for COVID-19 and about twenty clinical trials have started since the beginning of the pandemic. To gain information on the activity of doxycycline against SARS-CoV-2 infection and clarify some of the conflicting clinical data published, we designed in vitro binding tests and infection studies with a pseudotyped virus expressing the spike protein, as well as a clinically isolated SARS-CoV-2 strain. Doxycycline inhibited the transduction of the pseudotyped virus in Vero E6 and HEK-293 T cells stably expressing human receptor angiotensin-converting enzyme 2 but did not affect the entry and replication of SARS-CoV-2. Although this conclusion is apparently disappointing, it is paradigmatic of an experimental approach aimed at developing an integrated multidisciplinary platform. To avoid wasting precious time and resources we believe very stringent experimental criteria are needed in the preclinical phase, including infectious studies with SARS-CoV-2 in the platform before moving on to [failed] clinical trials.

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SARS-CoV-2 Infection Depends on Cellular Heparan Sulfate and ACE2

10.1101/2020.07.14.201616 ◽

2020 ◽

Cited By ~ 7

Author(s):

Thomas Mandel Clausen ◽

Daniel R. Sandoval ◽

Charlotte B. Spliid ◽

Jessica Pihl ◽

Chelsea D. Painter ◽

...

Keyword(s):

Protein Binding ◽

Heparan Sulfate ◽

Docking Studies ◽

Spike Protein ◽

Pseudotyped Virus ◽

Angiotensin Converting Enzyme 2 ◽

Viral Attachment ◽

Vitro Binding ◽

Domain Docking

AbstractWe show that SARS-CoV-2 spike protein interacts with cell surface heparan sulfate and angiotensin converting enzyme 2 (ACE2) through its Receptor Binding Domain. Docking studies suggest a putative heparin/heparan sulfate-binding site adjacent to the domain that binds to ACE2. In vitro, binding of ACE2 and heparin to spike protein ectodomains occurs independently and a ternary complex can be generated using heparin as a template. Contrary to studies with purified components, spike protein binding to heparan sulfate and ACE2 on cells occurs codependently. Unfractionated heparin, non-anticoagulant heparin, treatment with heparin lyases, and purified lung heparan sulfate potently block spike protein binding and infection by spike protein-pseudotyped virus and SARS-CoV-2 virus. These findings support a model for SARS-CoV-2 infection in which viral attachment and infection involves formation of a complex between heparan sulfate and ACE2. Manipulation of heparan sulfate or inhibition of viral adhesion by exogenous heparin may represent new therapeutic opportunities.

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Uncontrolled mass use of hydroxychloroquine

International Journal of Research in Pharmaceutical Sciences ◽

10.26452/ijrps.v11ispl1.2734 ◽

2020 ◽

Vol 11 (SPL1) ◽

pp. 396-398

Author(s):

Tugolbai Tagaev ◽

Sagynali Mamatov ◽

Farida Imanalieva ◽

Vityala Yethindra ◽

Altynai Zhumabekova ◽

...

Keyword(s):

Clinical Trials ◽

Therapeutic Drug Monitoring ◽

Severe Acute Respiratory Syndrome ◽

Adverse Effects ◽

Medical Advice ◽

Therapeutic Drug ◽

Converting Enzyme ◽

Weak Base ◽

Angiotensin Converting Enzyme 2

Hydroxychloroquine (HCQ) has previously been shown to inhibit coronavirus replication in vitro. But antiviral properties mechanisms are not well known, HCQ is a weak base that accumulates in lysosomes, modifies their pH, and interferes with some enzymes. In the lack of confirmed efficacy, the initial potential risk is not to expose patients to adverse effects. However, results from preliminary clinical studies have drawn inconclusive results regarding the efficacy of HCQ in coronavirus disease 2019 (COVID-19), due to several important weaknesses in research methodologies. Hypokalemia often occurs in patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), possibly due to the particular tropism of SARS-CoV-2 with regard to Angiotensin-converting enzyme 2 (ACE2). The wide use of HCQ, even against medical advice, will show an impact on ongoing clinical trials. It is important that we can recruite COVID-19 patients in these research studies to generate appropriate data regarding drugs that show promising efficacy against COVID-19. Currently, only doctors should be allowed to prescribe HCQ, and treatment should be confined to hospital settings, with proper cardiac and therapeutic drug monitoring.

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A Pervasive Review on New Advancements of Nano Vaccines on Covid-19 Pandemic

International Journal of Pharmaceutical Sciences Review and Research ◽

10.47583/ijpsrr.2021.v70i02.015 ◽

2021 ◽

Vol 70 (2) ◽

Author(s):

Ranajit Nath ◽

Ambika Mandal ◽

Ratul Bhowmik ◽

Ratna Roy ◽

Riya Biswas ◽

...

Keyword(s):

Clinical Trials ◽

Immune Responses ◽

Viral Vector ◽

Efficient Tool ◽

Angiotensin Converting Enzyme 2 ◽

Adaptive Immune ◽

Successful Vaccine ◽

Rapid Spread ◽

Health Organization

The infection that causes COVID-19 may be a pathogen referred to as severe acute respiratory syndrome coronavirus-2 (SARS-CoV2) and is believed to possess originated from China's Wuhan Province. The rapid spread of coronavirus disease 2019 (COVID-19) has become a worldwide concern, with the planet Health Organization (WHO) declaring it an epidemic on March, 2020. To enter the cells, SARS-CoV-2 S requires angiotensin-converting enzyme 2 (ACE2). Many existing vaccines have drawbacks like insufficient system stimulation, in vivo instability, high toxicity, the need for a chilly chain, and multiple administration. A nanotechnology is an efficient tool for addressing these issues. A successful vaccine against SARS-CoV-2 infection is predicted to stimulate innate and adaptive immune responses and protects against severe sorts of coronavirus disease 2019 (COVID-19). Different strategies are introduced because the go after an efficient and safe vaccination has begun. Currently, the foremost common vaccine types studied in clinical trials include viral vector-based vaccinations, genetic vaccines, attenuated vaccines, and protein-based vaccines. during this review, we cover the foremost promising anti-COVID-19 vaccine clinical trials also as various vaccination strategies to shed more light on the continued clinical trials. it's also discussed how nanotechnology is often wont to better understand the pathology of the present pandemic, also as how our understanding of SARS-CoV-2 cellular uptake and toxicity can influence future nanotoxicological considerations and nanomedicine design of safe yet effective nanomaterials.

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Dalbavancin binds ACE2 to block its interaction with SARS-CoV-2 spike protein and is effective in inhibiting SARS-CoV-2 infection in animal models

Cell Research ◽

10.1038/s41422-020-00450-0 ◽

2020 ◽

Vol 31 (1) ◽

pp. 17-24 ◽

Cited By ~ 1

Author(s):

Gan Wang ◽

Meng-Li Yang ◽

Zi-Lei Duan ◽

Feng-Liang Liu ◽

Lin Jin ◽

...

Keyword(s):

Clinical Trials ◽

Spike Protein ◽

Drug Candidate ◽

Peptide Drug ◽

Converting Enzyme ◽

Angiotensin Converting Enzyme 2 ◽

High Affinity ◽

Plasma Half Life

AbstractInfection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic worldwide. Currently, however, no effective drug or vaccine is available to treat or prevent the resulting coronavirus disease 2019 (COVID-19). Here, we report our discovery of a promising anti-COVID-19 drug candidate, the lipoglycopeptide antibiotic dalbavancin, based on virtual screening of the FDA-approved peptide drug library combined with in vitro and in vivo functional antiviral assays. Our results showed that dalbavancin directly binds to human angiotensin-converting enzyme 2 (ACE2) with high affinity, thereby blocking its interaction with the SARS-CoV-2 spike protein. Furthermore, dalbavancin effectively prevents SARS-CoV-2 replication in Vero E6 cells with an EC50 of ~12 nM. In both mouse and rhesus macaque models, viral replication and histopathological injuries caused by SARS-CoV-2 infection are significantly inhibited by dalbavancin administration. Given its high safety and long plasma half-life (8–10 days) shown in previous clinical trials, our data indicate that dalbavancin is a promising anti-COVID-19 drug candidate.

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A recombinant protein SARS-CoV-2 candidate vaccine elicits high-titer neutralizing antibodies in macaques.

10.1101/2020.12.20.422693 ◽

2020 ◽

Author(s):

Gary Baisa ◽

David Rancour ◽

Keith Mansfield ◽

Monika Burns ◽

Lori Martin ◽

...

Keyword(s):

Neutralizing Antibodies ◽

High Titer ◽

Pseudotyped Virus ◽

Binding Assays ◽

Neutralizing Activity ◽

Terminal Domain ◽

Vitro Binding ◽

Cynomolgus Macaques ◽

Human Igg1

Vaccines that generate robust and long-lived protective immunity against SARS-CoV-2 infection are urgently required. We assessed the potential of vaccine candidates based on the SARS-CoV-2 spike in cynomolgus macaques (M. fascicularis) by examining their ability to generate spike binding antibodies with neutralizing activity. Antigens were derived from two distinct regions of the spike S1 subunit, either the N-terminal domain (NTD) or an extended C-terminal domain containing the receptor-binding domain (RBD) and were fused to the human IgG1 Fc domain. Three groups of 2 animals each were immunized with either each antigen, alone or in combination. The development of antibody responses was evaluated through 20 weeks post-immunization. A robust IgG response to the spike protein was detected as early as 2 weeks after immunization with either protein and was maintained for over 20 weeks. Sera from animals immunized with antigens derived from the RBD were able to prevent binding of soluble spike proteins to the ACE2 receptor, shown by in vitro binding assays, while sera from animals immunized with the NTD alone lacked this activity. Crucially, sera from animals immunized with the RBD but not the NTD had potent neutralizing activity against SARS-CoV-2 pseudotyped virus, with titers in excess of 10,000, greatly exceeding that typically found in convalescent humans. Neutralizing activity persisted for more than 20 weeks. These data support the utility of spike subunit-based antigens as a vaccine for use in humans.

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Binding of SARS-CoV-2 spike protein to ACE2 is disabled by thiol-based drugs; evidence from in vitro SARS-CoV-2 infection studies

10.1101/2020.12.08.415505 ◽

2020 ◽

Author(s):

Kritika Khanna ◽

Wilfred Raymond ◽

Annabelle R. Charbit ◽

Jing Jin ◽

Irina Gitlin ◽

...

Keyword(s):

Host Cells ◽

Surface Glycoprotein ◽

Spike Protein ◽

Pseudotyped Virus ◽

Cell Infection ◽

Live Virus ◽

Angiotensin Converting Enzyme 2 ◽

Novel Treatments ◽

Binding Interface

AbstractCoronavirus disease 2019 (COVID-19) is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and the SARS-CoV-2 spike protein is an envelope glycoprotein that binds angiotensin converting enzyme 2 as an entry receptor. The capacity of enveloped viruses to infect host cells depends on a precise thiol/disulfide balance in their surface glycoprotein complexes. To determine if cystines in the SARS-CoV-2 spike protein maintain a native binding interface that can be disrupted by drugs that cleave cystines, we tested if thiol-based drugs have efficacy in receptor binding and cell infection assays. We found that thiol-based drugs, cysteamine and WR-1065 (the active metabolite of amifostine) in particular, decrease binding of SARS-CoV-2 spike protein to its receptor, decrease the entry efficiency of SARS-CoV-2 spike pseudotyped virus, and inhibit SARS-CoV-2 live virus infection. Our findings uncover a vulnerability of SARS-CoV-2 to thiol-based drugs and provide rationale to test thiol-based drugs, especially cysteamine and amifostine, as novel treatments for COVID-19.One Sentence SummaryThiol-based drugs decrease binding of SARS-CoV-2 spike protein to its receptor and inhibit SARS-CoV-2 cell entry.

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In Vitro and in Vivo Comparison of Binding of 99m-Tc-Iabeled Anti-CEA MAb F33-104 with 99m-Tc-labeled Anti-CEA MAb BW431/26

Nuklearmedizin ◽

10.1055/s-0038-1632202 ◽

1999 ◽

Vol 38 (04) ◽

pp. 115-119

Author(s):

N. Oriuchi ◽

S. Sugiyama ◽

M. Kuroki ◽

Y. Matsuoka ◽

S. Tanada ◽

...

Keyword(s):

Nude Mice ◽

Binding Capacity ◽

Colon Adenocarcinoma ◽

Human Colon ◽

Cell Binding ◽

Vitro Binding ◽

Labeling Method ◽

Human Colon Adenocarcinoma

Summary Aim: The purpose of this study was to assess the potential for radioimmunodetection (RAID) of murine anti-carcinoembryonic antigen (CEA) monoclonal antibody (MAb) F33-104 labeled with technetium-99m (99m-Tc) by a reduction-mediated labeling method. Methods: The binding capacity of 99m-Tc-labeled anti-CEA MAb F33-104 with CEA by means of in vitro procedures such as immunoradiometric assay and cell binding assay and the biodistribution of 99m-Tc-labeled anti-CEA MAb F33-104 in normal nude mice and nude mice bearing human colon adenocarcinoma LS180 tumor were investigated and compared with 99m-Tc-labeled anti-CEA MAb BW431/26. Results: The in vitro binding rate of 99m-Tc-labeled anti-CEA MAb F33-104 with CEA in solution and attached to the cell membrane was significantly higher than 99m-Tclabeled anti-CEA MAb BW431/261 (31.4 ± 0.95% vs. 11.9 ± 0.55% at 100 ng/mL of soluble CEA, 83.5 ± 2.84% vs. 54.0 ± 2.54% at 107 of LS 180 cells). In vivo, accumulation of 99m-Tc-labeled anti-CEA MAb F33-104 was higher at 18 h postinjection than 99m-Tc-labeled anti-CEA MAb BW431/26 (20.1 ± 3.50% ID/g vs. 14.4 ± 3.30% ID/g). 99m-Tcactivity in the kidneys of nude mice bearing tumor was higher at 18 h postinjection than at 3 h (12.8 ± 2.10% ID/g vs. 8.01 ± 2.40% ID/g of 99m-Tc-labeled anti-CEA MAb F33-104, 10.7 ± 1.70% ID/g vs. 8.10 ± 1.75% ID/g of 99m-Tc-labeled anti-CEA MAb BW431/26). Conclusion: 99m-Tc-labeled anti-CEA MAb F33-104 is a potential novel agent for RAID of recurrent colorectal cancer.

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Discovery of New AKT1 Inhibitors by Combination of In silico Structure Based Virtual Screening Approaches and Biological Evaluations

10.26434/chemrxiv.7591202 ◽

2019 ◽

Author(s):

Filip Fratev ◽

Denisse A. Gutierrez ◽

Renato J. Aguilera ◽

suman sirimulla

Keyword(s):

Virtual Screening ◽

Success Rate ◽

Protein Interactions ◽

In Silico ◽

Biological Data ◽

In Vitro Binding ◽

Vitro Binding ◽

Screening Protocol ◽

Screening Approaches

AKT1 is emerging as a useful target for treating cancer. Herein, we discovered a new set of ligands that inhibit the AKT1, as shown by in vitro binding and cell line studies, using a newly designed virtual screening protocol that combines structure-based pharmacophore and docking screens. Taking together with the biological data, the combination of structure based pharamcophore and docking methods demonstrated reasonable success rate in identifying new inhibitors (60-70%) proving the success of aforementioned approach. A detail analysis of the ligand-protein interactions was performed explaining observed activities.<br>

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Modulation of Matrix Metalloproteinases by Plant-Derived Products

Current Cancer Drug Targets ◽

10.2174/1568009620666201120144838 ◽

2020 ◽

Vol 20 ◽

Author(s):

Nur Najmi Mohamad Anuar ◽

Nurul Iman Natasya Zulkafali ◽

Azizah Ugusman

Keyword(s):

Clinical Trials ◽

Natural Products ◽

Matrix Metalloproteinases ◽

Animal Studies ◽

Current Knowledge ◽

In Vitro Models ◽

In Vivo Studies ◽

Extracellular Matrix Components

: Matrix metalloproteinases (MMPs) are a group of zinc-dependent metallo-endopeptidase that are responsible towards the degradation, repair and remodelling of extracellular matrix components. MMPs play an important role in maintaining a normal physiological function and preventing diseases such as cancer and cardiovascular diseases. Natural products derived from plants have been used as traditional medicine for centuries. Its active compounds, such as catechin, resveratrol and quercetin, are suggested to play an important role as MMPs inhibitors, thereby opening new insights into their applications in many fields, such as pharmaceutical, cosmetic and food industries. This review summarises the current knowledge on plant-derived natural products with MMP-modulating activities. Most of the reviewed plant-derived products exhibit an inhibitory activity on MMPs. Amongst MMPs, MMP-2 and MMP-9 are the most studied. The expression of MMPs is inhibited through respective signalling pathways, such as MAPK, NF-κB and PI3 kinase pathways, which contribute to the reduction in cancer cell behaviours, such as proliferation and migration. Most studies have employed in vitro models, but a limited number of animal studies and clinical trials have been conducted. Even though plant-derived products show promising results in modulating MMPs, more in vivo studies and clinical trials are needed to support their therapeutic applications in the future.

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