Combined prophylactic and therapeutic use maximizes hydroxychloroquine anti-SARS-CoV-2 effects<i>in vitro</i>

Clementi, Nicola; Criscuolo, Elena; Diotti, Roberta Antonia; Ferrarese, Roberto; Castelli, Matteo; Burioni, Roberto; Clementi, Massimo; Mancini, Nicasio

doi:10.1101/2020.03.29.014407

Cited by 10 publications

(9 citation statements)

References 11 publications

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“…For cellular invasion assays, Vero cells were plated at 2.5 × 10 5 cells per well in 24-well plates and incubated with EMEM (Sigma-Aldrich, Italy) supplemented with 10% (v/v) foetal calf serum (complete medium). Twenty four hours later, cells were incubated with porcine mucosal heparin (6.25–200 μg mL −1 ; Celsus Laboratories, Cincinnati, United States) in 300 µL of complete medium, 1 hour prior to infection and then incubated with virus solution containing 50 plaque forming units (PFUs) of either Italy/UniSR1/2020 isolate (GISAID accession ID: EPI_ISL_413489 18 or, SARS-CoV HSR-1 (EID 2004). After incubation for 1 hour at 37°C, supernatants were discarded and 500 µL of 1% (w/v) methylcellulose (Sigma-Aldrich, Italy) overlay (in complete medium) were added to each well.…”

Section: Methodsmentioning

confidence: 99%

“…Heparin Inhibits Cellular Invasion by SARS-CoV-2 Mycroft-West et al 1701 units (PFUs) of either Italy/UniSR1/2020 isolate (GISAID accession ID: EPI_ISL_413489 18 or, SARS-CoV HSR-1 (EID 2004). After incubation for 1 hour at 37°C, supernatants were discarded and 500 µL of 1% (w/v) methylcellulose (Sigma-Aldrich, Italy) overlay (in complete medium) were added to each well.…”

Section: Methods Vero Cell Culture and Assaysmentioning

confidence: 99%

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Heparin Inhibits Cellular Invasion by SARS-CoV-2: Structural Dependence of the Interaction of the Spike S1 Receptor-Binding Domain with Heparin

et al. 2020

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The dependence of development and homeostasis in animals on the interaction of hundreds of extracellular regulatory proteins with the peri- and extracellular glycosaminoglycan heparan sulfate (HS) is exploited by many microbial pathogens as a means of adherence and invasion. Heparin, a widely used anticoagulant drug, is structurally similar to HS and is a common experimental proxy. Exogenous heparin prevents infection by a range of viruses, including S-associated coronavirus isolate HSR1. Here, we show that heparin inhibits severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) invasion of Vero cells by up to 80% at doses achievable through prophylaxis and, particularly relevant, within the range deliverable by nebulisation. Surface plasmon resonance and circular dichroism spectroscopy demonstrate that heparin and enoxaparin, a low-molecular-weight heparin which is a clinical anticoagulant, bind and induce a conformational change in the spike (S1) protein receptor-binding domain (S1 RBD) of SARS-CoV-2. A library of heparin derivatives and size-defined fragments were used to probe the structural basis of this interaction. Binding to the RBD is more strongly dependent on the presence of 2-O or 6-O sulfate groups than on N-sulfation and a hexasaccharide is the minimum size required for secondary structural changes to be induced in the RBD. It is likely that inhibition of viral infection arises from an overlap between the binding sites of heparin/HS on S1 RBD and that of the angiotensin-converting enzyme 2. The results suggest a route for the rapid development of a first-line therapeutic by repurposing heparin and its derivatives as antiviral agents against SARS-CoV-2 and other members of the Coronaviridae.

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Section: Methodsmentioning

confidence: 99%

Section: Methods Vero Cell Culture and Assaysmentioning

confidence: 99%

Heparin Inhibits Cellular Invasion by SARS-CoV-2: Structural Dependence of the Interaction of the Spike S1 Receptor-Binding Domain with Heparin

et al. 2020

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“…While waiting for specific therapeutic treatments and/or vaccines against SARS-CoV-2, efforts initially focused on repurposing drugs that have previously shown broad-spectrum antiviral activity against other coronaviruses. Lopinavir/ritonavir, type I interferon (IFN), hydroxychloroquine, and remdesivir were among the first investigational treatments to be tested on the basis of their in vitro activity against SARS-CoV-2 (3)(4)(5)(6)(7)(8)(9). Activity of combined anti-HIV agent lopinavir/ritonavir against novel coronaviruses occurs via inhibition of 3chymotrypsin-like protease-dependent proteolysis preventing cleavage into viral proteins (3,4).…”

Section: Introductionmentioning

confidence: 99%

Antiviral drugs in hospitalized patients with COVID-19 - the DisCoVeRy trial

Ader¹,

Peiffer-Smadja²,

Poissy³

et al. 2021

Preprint

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BackgroundLopinavir/ritonavir, lopinavir/ritonavir-interferon (IFN)-β-1a and hydroxychloroquine efficacy for COVID-19 have been evaluated, but detailed evaluation is lacking.ObjectiveTo determine the efficacy of lopinavir/ritonavir, lopinavir/ritonavir-IFN-β-1a, hydroxychloroquine or remdesivir for improving the clinical, virological outcomes in COVID-19 inpatients.DesignOpen-label, randomized, adaptive, controlled trial.SettingMulti-center trial with patients from France.Participants583 COVID-19 inpatients requiring oxygen and/or ventilatory supportInterventionStandard of care (SoC, control), SoC plus lopinavir/ritonavir (400 mg lopinavir and 100 mg ritonavir every 12h for 14 days), SoC plus lopinavir/ritonavir plus IFN-ß-1a (44 μg of subcutaneous IFN-ß-1a on days 1, 3, and 6), SoC plus hydroxychloroquine (400 mg twice on day 1 then 400 mg once daily for 9 days) or SoC plus remdesivir (200 mg intravenously on day 1 then 100 mg once-daily for hospitalization duration or 10 days).MeasurementsThe primary outcome was the clinical status at day 15, measured by the WHO 7-point ordinal scale. Secondary outcomes included SARS-CoV-2 quantification in respiratory specimens and safety analyses.ResultsAdjusted Odds Ratio (aOR) for the WHO 7-point ordinal scale were not in favor of investigational treatments: lopinavir/ritonavir versus control, aOR 0.83, 95%CI, 0.55 to 1.26, P=0.39; lopinavir/ritonavir-IFN-β-1a versus control, aOR 0.69, 95%CI, 0.45 to 1.04, P=0.08; hydroxychloroquine versus control, aOR 0.93, 95%CI, 0.62 to 1.41, P=0.75. No significant effect on SARS-CoV-2 RNA clearance in respiratory tract was evidenced. Lopinavir/ritonavir-containing treatments were significantly associated with more SAE.LimitationsNot a placebo-controlled, no anti-inflammatory agents tested.ConclusionNo improvement of the clinical status at day 15 nor SARS-CoV-2 RNA clearance in respiratory tract specimens by studied drugs. This comforts the recent Solidarity findings.RegistrationNCT04315948.FundingPHRC 2020, Dim OneHealth, REACTing

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“…Currently, there are multiple clinical trials underway to investigate the potential use of hydroxychloroquine and chloroquine alone or in combination against SARS-CoV-2 (Table 5). Some of the in vitro and in vivo results obtained with chloroquine and hydroxychloroquine supported their anti-viral role against SARS-CoV-2, (Andreani et al, 2020;Clementi et al, 2020;Fantini et al, 2020;Gao et al, 2020;Gautret et al, 2020;Liu et al, 2020;Weston et al, 2020;Yao et al, 2020), however, results of Gautret et al (2020) faced severe criticism because of small sample size, overruling type I errors, inconsistency between study protocols and lack of blinding and a control arm even though the treatment resulted in viral load reduction. It is also very important to reproduce the in vitro results obtained with chloroquine in the in vivo studies and clinical trials to establish it as a safe and effective anti-SARS-CoV-2 drug.…”

Section: Results Of Clinical Trials Done So Far With Chloroquine Against Covid-19mentioning

confidence: 98%

Repurposing Chloroquine Against Multiple Diseases With Special Attention to SARS-CoV-2 and Associated Toxicity

Kamat

Kumari

2021

Front. Pharmacol.

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Chloroquine and its derivatives have been used since ages to treat malaria and have also been approved by the FDA to treat autoimmune diseases. The drug employs pH-dependent inhibition of functioning and signalling of the endosome, lysosome and trans-Golgi network, immunomodulatory actions, inhibition of autophagy and interference with receptor binding to treat cancer and many viral diseases. The ongoing pandemic of COVID-19 has brought the whole world on the knees, seeking an urgent hunt for an anti-SARS-CoV-2 drug. Chloroquine has shown to inhibit receptor binding of the viral particles, interferes with their replication and inhibits “cytokine storm”. Though multiple modes of actions have been employed by chloroquine against multiple diseases, viral diseases can provide an added advantage to establish the anti–SARS-CoV-2 mechanism, the in vitro and in vivo trials against SARS-CoV-2 have yielded mixed results. The toxicological effects and dosage optimization of chloroquine have been studied for many diseases, though it needs a proper evaluation again as chloroquine is also associated with several toxicities. Moreover, the drug is inexpensive and is readily available in many countries. Though much of the hope has been created by chloroquine and its derivatives against multiple diseases, repurposing it against SARS-CoV-2 requires large scale, collaborative, randomized and unbiased clinical trials to avoid false promises. This review summarizes the use and the mechanism of chloroquine against multiple diseases, its side-effects, mechanisms and the different clinical trials ongoing against “COVID-19”.

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Combined prophylactic and therapeutic use maximizes hydroxychloroquine anti-SARS-CoV-2 effectsin vitro

Cited by 10 publications

References 11 publications

Heparin Inhibits Cellular Invasion by SARS-CoV-2: Structural Dependence of the Interaction of the Spike S1 Receptor-Binding Domain with Heparin

Heparin Inhibits Cellular Invasion by SARS-CoV-2: Structural Dependence of the Interaction of the Spike S1 Receptor-Binding Domain with Heparin

Antiviral drugs in hospitalized patients with COVID-19 - the DisCoVeRy trial

Repurposing Chloroquine Against Multiple Diseases With Special Attention to SARS-CoV-2 and Associated Toxicity

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