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1.
Int J Mol Sci ; 22(16)2021 Aug 13.
Article in English | MEDLINE | ID: covidwho-1662675

ABSTRACT

Iron oxide nanoparticles and single domain antibodies from camelids (VHHs) have been increasingly recognized for their potential uses for medical diagnosis and treatment. However, there have been relatively few detailed characterizations of their pharmacokinetics (PK). The aim of this study was to develop imaging methods and pharmacokinetic models to aid the future development of a novel family of brain MRI molecular contrast agents. An efficient near-infrared (NIR) imaging method was established to monitor VHH and VHH conjugated nanoparticle kinetics in mice using a hybrid approach: kinetics in blood were assessed by direct sampling, and kinetics in kidney, liver, and brain were assessed by serial in vivo NIR imaging. These studies were performed under "basal" circumstances in which the VHH constructs and VHH-conjugated nanoparticles do not substantially interact with targets nor cross the blood brain barrier. Using this approach, we constructed a five-compartment PK model that fits the data well for single VHHs, engineered VHH trimers, and iron oxide nanoparticles conjugated to VHH trimers. The establishment of the feasibility of these methods lays a foundation for future PK studies of candidate brain MRI molecular contrast agents.


Subject(s)
Camelids, New World/immunology , Kidney/chemistry , Liver/chemistry , Magnetic Iron Oxide Nanoparticles/chemistry , Single-Domain Antibodies/administration & dosage , Administration, Intravenous , Animals , Brain Chemistry , Female , Fluorometry , Humans , Mice , Models, Theoretical , Particle Size , Single-Domain Antibodies/blood , Single-Domain Antibodies/chemistry
2.
Proc Natl Acad Sci U S A ; 118(44)2021 11 02.
Article in English | MEDLINE | ID: covidwho-1470027

ABSTRACT

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in over 100 million infections and millions of deaths. Effective vaccines remain the best hope of curtailing SARS-CoV-2 transmission, morbidity, and mortality. The vaccines in current use require cold storage and sophisticated manufacturing capacity, which complicates their distribution, especially in less developed countries. We report the development of a candidate SARS-CoV-2 vaccine that is purely protein based and directly targets antigen-presenting cells. It consists of the SARS-CoV-2 Spike receptor-binding domain (SpikeRBD) fused to an alpaca-derived nanobody that recognizes class II major histocompatibility complex antigens (VHHMHCII). This vaccine elicits robust humoral and cellular immunity against SARS-CoV-2 and its variants. Both young and aged mice immunized with two doses of VHHMHCII-SpikeRBD elicit high-titer binding and neutralizing antibodies. Immunization also induces strong cellular immunity, including a robust CD8 T cell response. VHHMHCII-SpikeRBD is stable for at least 7 d at room temperature and can be lyophilized without loss of efficacy.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19 Vaccines/pharmacology , COVID-19/immunology , COVID-19/prevention & control , Pandemics , SARS-CoV-2/immunology , Amino Acid Sequence , Animals , Antibodies, Neutralizing/biosynthesis , Antibodies, Viral/biosynthesis , Antigen-Presenting Cells/immunology , CD8-Positive T-Lymphocytes/immunology , COVID-19/epidemiology , COVID-19 Vaccines/administration & dosage , Camelids, New World/immunology , Female , Histocompatibility Antigens Class II/immunology , Humans , Immunity, Cellular , Immunity, Humoral , Immunization, Secondary , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Transgenic , Pandemics/prevention & control , Recombinant Fusion Proteins/administration & dosage , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/immunology , SARS-CoV-2/genetics , Single-Domain Antibodies/administration & dosage , Single-Domain Antibodies/immunology , Spike Glycoprotein, Coronavirus/administration & dosage , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology
3.
PLoS Pathog ; 17(10): e1009542, 2021 10.
Article in English | MEDLINE | ID: covidwho-1468184

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes the disease COVID-19 can lead to serious symptoms, such as severe pneumonia, in the elderly and those with underlying medical conditions. While vaccines are now available, they do not work for everyone and therapeutic drugs are still needed, particularly for treating life-threatening conditions. Here, we showed nasal delivery of a new, unmodified camelid single-domain antibody (VHH), termed K-874A, effectively inhibited SARS-CoV-2 titers in infected lungs of Syrian hamsters without causing weight loss and cytokine induction. In vitro studies demonstrated that K-874A neutralized SARS-CoV-2 in both VeroE6/TMPRSS2 and human lung-derived alveolar organoid cells. Unlike other drug candidates, K-874A blocks viral membrane fusion rather than viral attachment. Cryo-electron microscopy revealed K-874A bound between the receptor binding domain and N-terminal domain of the virus S protein. Further, infected cells treated with K-874A produced fewer virus progeny that were less infective. We propose that direct administration of K-874A to the lung could be a new treatment for preventing the reinfection of amplified virus in COVID-19 patients.


Subject(s)
Antibodies, Viral/administration & dosage , Antiviral Agents/administration & dosage , COVID-19 , Single-Domain Antibodies/administration & dosage , Virus Attachment/drug effects , Administration, Intranasal , Animals , Chlorocebus aethiops , Cricetinae , Disease Models, Animal , Humans , Mesocricetus , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/immunology , Vero Cells
4.
Nat Commun ; 12(1): 5469, 2021 09 22.
Article in English | MEDLINE | ID: covidwho-1434103

ABSTRACT

SARS-CoV-2 remains a global threat to human health particularly as escape mutants emerge. There is an unmet need for effective treatments against COVID-19 for which neutralizing single domain antibodies (nanobodies) have significant potential. Their small size and stability mean that nanobodies are compatible with respiratory administration. We report four nanobodies (C5, H3, C1, F2) engineered as homotrimers with pmolar affinity for the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Crystal structures show C5 and H3 overlap the ACE2 epitope, whilst C1 and F2 bind to a different epitope. Cryo Electron Microscopy shows C5 binding results in an all down arrangement of the Spike protein. C1, H3 and C5 all neutralize the Victoria strain, and the highly transmissible Alpha (B.1.1.7 first identified in Kent, UK) strain and C1 also neutralizes the Beta (B.1.35, first identified in South Africa). Administration of C5-trimer via the respiratory route showed potent therapeutic efficacy in the Syrian hamster model of COVID-19 and separately, effective prophylaxis. The molecule was similarly potent by intraperitoneal injection.


Subject(s)
Antibodies, Neutralizing/pharmacology , COVID-19/drug therapy , Single-Domain Antibodies/pharmacology , Spike Glycoprotein, Coronavirus/metabolism , Administration, Intranasal , Animals , Antibodies, Neutralizing/administration & dosage , Antibodies, Neutralizing/genetics , Antibodies, Neutralizing/immunology , Cryoelectron Microscopy , Crystallography, X-Ray , Disease Models, Animal , Dose-Response Relationship, Immunologic , Epitopes/chemistry , Epitopes/metabolism , Female , Male , Mesocricetus , Neutralization Tests , SARS-CoV-2/drug effects , Single-Domain Antibodies/administration & dosage , Single-Domain Antibodies/immunology , Single-Domain Antibodies/metabolism , Spike Glycoprotein, Coronavirus/chemistry
5.
Sci Adv ; 7(22)2021 05.
Article in English | MEDLINE | ID: covidwho-1247309

ABSTRACT

Globally, there is an urgency to develop effective, low-cost therapeutic interventions for coronavirus disease 2019 (COVID-19). We previously generated the stable and ultrapotent homotrimeric Pittsburgh inhalable Nanobody 21 (PiN-21). Using Syrian hamsters that model moderate to severe COVID-19 disease, we demonstrate the high efficacy of PiN-21 to prevent and treat SARS-CoV-2 infection. Intranasal delivery of PiN-21 at 0.6 mg/kg protects infected animals from weight loss and substantially reduces viral burdens in both lower and upper airways compared to control. Aerosol delivery of PiN-21 facilitates deposition throughout the respiratory tract and dose minimization to 0.2 mg/kg. Inhalation treatment quickly reverses animals' weight loss after infection, decreases lung viral titers by 6 logs leading to drastically mitigated lung pathology, and prevents viral pneumonia. Combined with the marked stability and low production cost, this innovative therapy may provide a convenient and cost-effective option to mitigate the ongoing pandemic.


Subject(s)
COVID-19/drug therapy , COVID-19/prevention & control , SARS-CoV-2/drug effects , Single-Domain Antibodies/administration & dosage , Administration, Inhalation , Aerosols/administration & dosage , Animals , Disease Models, Animal , Female , Male , Mesocricetus , Pandemics/prevention & control , Pneumonia, Viral/drug therapy , Pneumonia, Viral/prevention & control , Viral Load/drug effects
6.
Front Immunol ; 11: 581076, 2020.
Article in English | MEDLINE | ID: covidwho-1116666

ABSTRACT

COVID-19 has become difficult to contain in our interconnected world. In this article, we discuss some approaches that could reduce the consequences of COVID-19. We elaborate upon the utility of camelid single-domain antibodies (sdAbs), also referred to as nanobodies, which are naturally poised to neutralize viruses without enhancing its infectivity. Smaller sized sdAbs can be easily selected using microbes or the subcellular organelle display methods and can neutralize SARS-CoV2 infectivity. We also discuss issues related to their production using scalable platforms. The favorable outcome of the infection is evident in patients when the inflammatory response is adequately curtailed. Therefore, we discuss approaches to mitigate hyperinflammatory reactions initiated by SARS-CoV2 but orchestrated by immune mediators.


Subject(s)
Antibodies, Viral/administration & dosage , Antiviral Agents/administration & dosage , COVID-19/drug therapy , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Single-Domain Antibodies/administration & dosage , COVID-19/immunology , COVID-19/virology , Humans , SARS-CoV-2/genetics , SARS-CoV-2/immunology
8.
J Thromb Thrombolysis ; 52(2): 468-470, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1002138

ABSTRACT

Acquired thrombotic thrombocytopenic purpura (TTP) is an autoimmune disease that can be triggered by different events, including viral infections. It presents as thrombotic microangiopathy and can lead to severe complications that often require management in the intensive care unit (ICU). We report a patient who presented with acquired TTP following COVID-19 infection. A 44-year-old woman presented to the emergency department with severe anemia, acute kidney injury and respiratory failure due to COVID-19. Clinical and laboratory findings were suggestive for thrombotic microangiopathy. On day 8 laboratory tests confirmed the diagnosis of acquired TTP. The patient needed 14 plasma exchanges, treatment with steroids, rituximab and caplacizumab and 18 days of mechanical ventilation. She completely recovered and was discharged home on day 51. Acquired TTP can be triggered by different events leading to immune stimulation. COVID-19 has been associated with different inflammatory and auto-immune diseases. Considering the temporal sequence and the lack of other possible causes, we suggest that COVID-19 infection could have been the triggering factor in the development of TTP. Since other similar cases have already been described, possible association between COVID and TTP deserves further investigation.


Subject(s)
COVID-19 , Plasma Exchange/methods , Purpura, Thrombotic Thrombocytopenic , Respiration, Artificial/methods , Respiratory Insufficiency , Rituximab/administration & dosage , Single-Domain Antibodies/administration & dosage , Acute Kidney Injury/diagnosis , Acute Kidney Injury/etiology , Adult , COVID-19/complications , COVID-19/immunology , COVID-19/physiopathology , COVID-19/therapy , Female , Fibrinolytic Agents/administration & dosage , Humans , Immunologic Factors/administration & dosage , Male , Purpura, Thrombotic Thrombocytopenic/blood , Purpura, Thrombotic Thrombocytopenic/etiology , Purpura, Thrombotic Thrombocytopenic/physiopathology , Purpura, Thrombotic Thrombocytopenic/therapy , Respiratory Insufficiency/diagnosis , Respiratory Insufficiency/etiology , Respiratory Insufficiency/therapy , SARS-CoV-2/isolation & purification , SARS-CoV-2/pathogenicity , Thrombotic Microangiopathies/diagnosis , Thrombotic Microangiopathies/etiology , Treatment Outcome
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