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1.
Genome Biol ; 23(1): 55, 2022 02 16.
Article in English | MEDLINE | ID: covidwho-1785167

ABSTRACT

BACKGROUND: Multiplexing of samples in single-cell RNA-seq studies allows a significant reduction of the experimental costs, straightforward identification of doublets, increased cell throughput, and reduction of sample-specific batch effects. Recently published multiplexing techniques using oligo-conjugated antibodies or -lipids allow barcoding sample-specific cells, a process called "hashing." RESULTS: Here, we compare the hashing performance of TotalSeq-A and -C antibodies, custom synthesized lipids and MULTI-seq lipid hashes in four cell lines, both for single-cell RNA-seq and single-nucleus RNA-seq. We also compare TotalSeq-B antibodies with CellPlex reagents (10x Genomics) on human PBMCs and TotalSeq-B with different lipids on primary mouse tissues. Hashing efficiency was evaluated using the intrinsic genetic variation of the cell lines and mouse strains. Antibody hashing was further evaluated on clinical samples using PBMCs from healthy and SARS-CoV-2 infected patients, where we demonstrate a more affordable approach for large single-cell sequencing clinical studies, while simultaneously reducing batch effects. CONCLUSIONS: Benchmarking of different hashing strategies and computational pipelines indicates that correct demultiplexing can be achieved with both lipid- and antibody-hashed human cells and nuclei, with MULTISeqDemux as the preferred demultiplexing function and antibody-based hashing as the most efficient protocol on cells. On nuclei datasets, lipid hashing delivers the best results. Lipid hashing also outperforms antibodies on cells isolated from mouse brain. However, antibodies demonstrate better results on tissues like spleen or lung.


Subject(s)
COVID-19/blood , Sequence Analysis, RNA/methods , Single-Cell Analysis/methods , Animals , Antibodies/chemistry , Case-Control Studies , Cell Line, Tumor , Cell Nucleus/chemistry , Humans , Lipids/chemistry , Mice, Inbred BALB C , Mice, Inbred C57BL , Neutrophils/chemistry , Neutrophils/immunology , Neutrophils/virology
2.
Trends Biotechnol ; 40(4): 463-481, 2022 04.
Article in English | MEDLINE | ID: covidwho-1735006

ABSTRACT

Humoral immunity is divided into the cellular B cell and protein-level antibody responses. High-throughput sequencing has advanced our understanding of both these fundamental aspects of B cell immunology as well as aspects pertaining to vaccine and therapeutics biotechnology. Although the protein-level serum and mucosal antibody repertoire make major contributions to humoral protection, the sequence composition and dynamics of antibody repertoires remain underexplored. This limits insight into important immunological and biotechnological parameters such as the number of antigen-specific antibodies, which are for example, relevant for pathogen neutralization, microbiota regulation, severity of autoimmunity, and therapeutic efficacy. High-resolution mass spectrometry (MS) has allowed initial insights into the antibody repertoire. We outline current challenges in MS-based sequence analysis of antibody repertoires and propose strategies for their resolution.


Subject(s)
Antibodies , High-Throughput Nucleotide Sequencing , Antibodies/chemistry , Antigens , B-Lymphocytes , High-Throughput Nucleotide Sequencing/methods , Mass Spectrometry
3.
Int J Biol Macromol ; 200: 438-448, 2022 Mar 01.
Article in English | MEDLINE | ID: covidwho-1633972

ABSTRACT

As SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) continues to inflict chaos globally, a new variant officially known as B.1.1.529 was reported in South Africa and was found to harbor 30 mutations in the spike protein. It is too early to speculate on transmission and hospitalizations. Hence, more analyses are required, particularly to connect the genomic patterns to the phenotypic attributes to reveal the binding differences and antibody response for this variant, which can then be used for therapeutic interventions. Given the urgency of the required analysis and data on the B.1.1.529 variant, we have performed a detailed investigation to provide an understanding of the impact of these novel mutations on the structure, function, and binding of RBD to hACE2 and mAb to the NTD of the spike protein. The differences in the binding pattern between the wild type and B.1.1.529 variant complexes revealed that the key substitutions Asn417, Ser446, Arg493, and Arg498 in the B.1.1.529 RBD caused additional interactions with hACE2 and the loss of key residues in the B.1.1.529 NTD resulted in decreased interactions with three CDR regions (1-3) in the mAb. Further investigation revealed that B.1.1.529 displayed a stable dynamic that follows a global stability trend. In addition, the dissociation constant (KD), hydrogen bonding analysis, and binding free energy calculations further validated the findings. Hydrogen bonding analysis demonstrated that significant hydrogen bonding reprogramming took place, which revealed key differences in the binding. The total binding free energy using MM/GBSA and MM/PBSA further validated the docking results and demonstrated significant variations in the binding. This study is the first to provide a basis for the higher infectivity of the new SARS-CoV-2 variants and provides a strong impetus for the development of novel drugs against them.


Subject(s)
Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Antibodies/chemistry , Antibodies/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/metabolism , Humans , Hydrogen Bonding , Immune Evasion , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Binding/immunology , Protein Domains/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
4.
Clin Transl Med ; 11(12): e668, 2021 12.
Article in English | MEDLINE | ID: covidwho-1568016

ABSTRACT

The level of postvaccine protection depends on two factors: antibodies and T-cell responses. While the first one is relatively easily measured, the measuring of the second one is a difficult problem. The recent studies indicate that the first one may be a good proxy for the protection, at least for SARS-CoV-2. The massive data currently gathered by both researcher and citizen scientists may be pivotal in confirming this observation, and the collective body of evidence is growing daily. This leads to an acceptance of IgG antibody levels as an accessible biomarker of individual's protection. With enormous and immediate need for assessing patient condition at the point of care, quantitative antibody analysis remains the most effective and efficient way to assess the protection against the disease. Let us not discount importance of reference points in the turmoil of current pandemics.


Subject(s)
Antibodies, Viral/chemistry , Antibodies/chemistry , Biomarkers/metabolism , COVID-19/blood , COVID-19/immunology , Antibody Specificity , Humans , Immune System , Immunity , Immunoglobulin G/metabolism , Intensive Care Units , Pandemics , Point-of-Care Systems , SARS-CoV-2 , Serologic Tests/methods , Serologic Tests/standards , Vaccines
5.
Protein Sci ; 31(1): 141-146, 2022 01.
Article in English | MEDLINE | ID: covidwho-1520274

ABSTRACT

The antibody repertoires of individuals and groups have been used to explore disease states, understand vaccine responses, and drive therapeutic development. The arrival of B-cell receptor repertoire sequencing has enabled researchers to get a snapshot of these antibody repertoires, and as more data are generated, increasingly in-depth studies are possible. However, most publicly available data only exist as raw FASTQ files, making the data hard to access, process, and compare. The Observed Antibody Space (OAS) database was created in 2018 to offer clean, annotated, and translated repertoire data. In this paper, we describe an update to OAS that has been driven by the increasing volume of data and the appearance of paired (VH/VL) sequence data. OAS is now accessible via a new web server, with standardized search parameters and a new sequence-based search option. The new database provides both nucleotides and amino acids for every sequence, with additional sequence annotations to make the data Minimal Information about Adaptive Immune Receptor Repertoire compliant, and comments on potential problems with the sequence. OAS now contains 25 new studies, including severe acute respiratory syndrome coronavirus 2 data and paired sequencing data. The new database is accessible at http://opig.stats.ox.ac.uk/webapps/oas/, and all data are freely available for download.


Subject(s)
Antibodies/chemistry , Databases, Protein , Amino Acid Sequence , Animals , Antibodies/immunology , COVID-19/immunology , Humans , Immunoglobulin Heavy Chains/chemistry , Immunoglobulin Heavy Chains/immunology , Immunoglobulin Light Chains/chemistry , Immunoglobulin Light Chains/immunology , Immunoglobulin Variable Region/chemistry , Immunoglobulin Variable Region/immunology , SARS-CoV-2/immunology
6.
Adv Sci (Weinh) ; 8(23): e2103266, 2021 12.
Article in English | MEDLINE | ID: covidwho-1479368

ABSTRACT

Activation of endothelial cells following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is thought to be the primary driver for the increasingly recognized thrombotic complications in coronavirus disease 2019 patients, potentially due to the SARS-CoV-2 Spike protein binding to the human angiotensin-converting enzyme 2 (hACE2). Vaccination therapies use the same Spike sequence or protein to boost host immune response as a protective mechanism against SARS-CoV-2 infection. As a result, cases of thrombotic events are reported following vaccination. Although vaccines are generally considered safe, due to genetic heterogeneity, age, or the presence of comorbidities in the population worldwide, the prediction of severe adverse outcome in patients remains a challenge. To elucidate Spike proteins underlying patient-specific-vascular thrombosis, the human microcirculation environment is recapitulated using a novel microfluidic platform coated with human endothelial cells and exposed to patient specific whole blood. Here, the blood coagulation effect is tested after exposure to Spike protein in nanoparticles and Spike variant D614G in viral vectors and the results are corroborated using live SARS-CoV-2. Of note, two potential strategies are also examined to reduce blood clot formation, by using nanoliposome-hACE2 and anti-Interleukin (IL) 6 antibodies.


Subject(s)
Blood Coagulation/physiology , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/metabolism , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Antibodies/chemistry , Antibodies/immunology , Antibodies/metabolism , COVID-19/diagnosis , COVID-19/virology , Endothelial Cells/chemistry , Endothelial Cells/cytology , Endothelial Cells/metabolism , Fibrin/chemistry , Fibrin/metabolism , Genetic Vectors/genetics , Genetic Vectors/metabolism , Humans , Interleukin-6/immunology , Liposomes/chemistry , Microfluidics/methods , Mutation , Nanoparticles/chemistry , Platelet Aggregation , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/analysis , Spike Glycoprotein, Coronavirus/genetics
7.
Science ; 372(6537)2021 04 02.
Article in English | MEDLINE | ID: covidwho-1166346

ABSTRACT

Multivalent display of receptor-engaging antibodies or ligands can enhance their activity. Instead of achieving multivalency by attachment to preexisting scaffolds, here we unite form and function by the computational design of nanocages in which one structural component is an antibody or Fc-ligand fusion and the second is a designed antibody-binding homo-oligomer that drives nanocage assembly. Structures of eight nanocages determined by electron microscopy spanning dihedral, tetrahedral, octahedral, and icosahedral architectures with 2, 6, 12, and 30 antibodies per nanocage, respectively, closely match the corresponding computational models. Antibody nanocages targeting cell surface receptors enhance signaling compared with free antibodies or Fc-fusions in death receptor 5 (DR5)-mediated apoptosis, angiopoietin-1 receptor (Tie2)-mediated angiogenesis, CD40 activation, and T cell proliferation. Nanocage assembly also increases severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudovirus neutralization by α-SARS-CoV-2 monoclonal antibodies and Fc-angiotensin-converting enzyme 2 (ACE2) fusion proteins.


Subject(s)
Antibodies/chemistry , Antibodies/immunology , Nanostructures , Protein Engineering , Signal Transduction , Angiopoietins/chemistry , Angiopoietins/immunology , Angiopoietins/metabolism , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/immunology , Antibodies, Viral/chemistry , Antibodies, Viral/immunology , B-Lymphocytes/immunology , CD40 Antigens/chemistry , CD40 Antigens/immunology , CD40 Antigens/metabolism , Cell Line, Tumor , Cell Proliferation , Computer Simulation , Genes, Synthetic , Humans , Immunoglobulin Fc Fragments/chemistry , Lymphocyte Activation , Models, Molecular , Protein Binding , Receptor, TIE-2/metabolism , Receptors, TNF-Related Apoptosis-Inducing Ligand/immunology , Receptors, TNF-Related Apoptosis-Inducing Ligand/metabolism , SARS-CoV-2/immunology , T-Lymphocytes/immunology , T-Lymphocytes/physiology
8.
Nature ; 589(7843): 630-632, 2021 01.
Article in English | MEDLINE | ID: covidwho-1049956
9.
Chembiochem ; 22(8): 1371-1378, 2021 04 16.
Article in English | MEDLINE | ID: covidwho-985970

ABSTRACT

Infectious diseases are a continues threat to human health and the economy worldwide. The latest example is the global pandemic of COVID-19 caused by SARS-CoV-2. Antibody therapy and vaccines are promising approaches to treat the disease; however, they have bottlenecks: they might have low efficacy or narrow breadth due to the continuous emergence of new strains of the virus or antibodies could cause antibody-dependent enhancement (ADE) of infection. To address these bottlenecks, I propose the use of 24-meric ferritin for the synthesis of mosaic nanocages to deliver a cocktail of antibodies or nanobodies alone or in combination with another therapeutic, like a nucleotide analogue, to mimic the viral entry process and deceive the virus, or to develop mosaic vaccines. I argue that available data showing the effectiveness of ferritin-antibody conjugates in targeting specific cells and ferritin-haemagglutinin nanocages in developing influenza vaccines strongly support my proposals.


Subject(s)
Antiviral Agents/chemistry , Ferritins/chemistry , Nanostructures/chemistry , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Animals , Antibodies/chemistry , Antibodies/therapeutic use , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , COVID-19/drug therapy , COVID-19/prevention & control , COVID-19/virology , COVID-19 Vaccines/chemistry , COVID-19 Vaccines/immunology , Drug Carriers/chemistry , Ferritins/metabolism , Humans , Mice , SARS-CoV-2/isolation & purification , Serine Endopeptidases/chemistry , Serine Endopeptidases/metabolism , Virus Internalization/drug effects
10.
Med Hypotheses ; 145: 110343, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-842726

ABSTRACT

ABO blood groups is a cheap and affordable test that can be immediately retrieved from COVID-19 patients at the diagnosis. There is increasing evidence that non-O blood groups have both higher susceptibility and higher severity of COVID-19 infections. The reason behind such relationship seems elusive. Regarding susceptibility, Non-O individuals have Anti-A antibodies which can prevent viral entry across ACE-2 receptors, moreover, Non-O individuals are at higher risk of autoimmunity, hypercoagulable state, and dysbiosis resulting in an augmented tendency for vascular inflammatory sequelae of COVID-19. We can conclude, on the diagnostic level, that ABO blood groups can be potentially used for risk stratification of affected COVID-19 patients, to anticipate the deterioration of patients at higher risk for complications. On a therapeutic level, plasma from normal O blood group individuals might potentially replace the use of convalescent serum for the treatment of COVID-19.


Subject(s)
ABO Blood-Group System , COVID-19 Serological Testing/methods , COVID-19/blood , COVID-19/diagnosis , Risk Assessment/methods , Antibodies/chemistry , Autoimmunity , COVID-19/immunology , COVID-19/therapy , Disease Progression , Female , Furin/metabolism , Gastrointestinal Microbiome , Humans , Immunization, Passive , Male , Pandemics , Thrombosis
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