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1.
Sci Rep ; 12(1): 8145, 2022 May 17.
Article in English | MEDLINE | ID: covidwho-1864761

ABSTRACT

Wild boar (Sus scrofa L.) is one of the large mammals most spread worldwide, highly adaptable, and its population rapidly increased in many areas in Europe, including Italy, where Tuscany is considered particularly suitable for wild boar. Wild boars are potential hosts for different etiological agents, such as Brucella spp., Leptospira spp. and Pseudorabies virus and they can contribute to maintain and/or to disseminate some bacterial or viral pathogens to humans and domestic animals, above all-in free-range farms. In order to identify hypothetical genomic regions associated with these infection diseases, 96 samples of wild boars hunted in Tuscany during the 2018-2019 and 2019-2020 hunting seasons were considered. Diagnosis was achieved by serological tests and 42 Pseudorabies, 31 Leptospira and 15 Brucella positive animals were identified. All animals were genotyped with Geneseek Genomic Profiler Porcine HD (70 k) and a genome-wide scan was then performed. Significant markers were highlighted for Pseudorabies (two SNPs), Brucella (seven SNPs), and Leptospira (four SNPs) and they were located within, or nearby, 29 annotated genes on chromosome 6, 9, 12, 13, 14 and 18. Eight genes are implicated in viral (SEC14L1, JMJD6, SRSF2, TMPRSS2, MX1, MX2) or bacterial (COL8A1, SPIRE1) infections, seven genes (MFSD11, METTL23, CTTNBP2, BACE2, IMPA2, MPPE1 and GNAL) are involved in mental disorders and one gene (MGAT5B) is related to the Golgi complex. Results presented here provide interesting starting points for future research, validation studies and fine mapping of candidate genes involved in bacterial and viral infections in wild boar.


Subject(s)
Brucella , Leptospira , Pseudorabies , Swine Diseases , Virus Diseases , Animals , Carrier Proteins , Humans , Italy/epidemiology , Jumonji Domain-Containing Histone Demethylases , Leptospira/genetics , Phosphoprotein Phosphatases , Sus scrofa , Swine , Virus Diseases/epidemiology
2.
Biosens Bioelectron ; 212: 114406, 2022 Sep 15.
Article in English | MEDLINE | ID: covidwho-1850711

ABSTRACT

Coronavirus Disease 2019 (COVID-19) pandemic has shown the need for early diagnosis to manage infectious disease outbreaks. Here, we report a label free electrochemical Fluorine-Doped Tin Oxide (FTO) Immunosensor coupled with gold nanorods (GNRs) as an electron carrier for ultrasensitive detection of the Receptor Binding Domain (RBD) of SARS CoV-2 Spike protein. The RBD gene was cloned, and expressed in-house with confirmed molecular weight of ∼31 kDa via Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF). RBD antibodies (Ab) were generated to be used as a bioreceptor for sensor fabrication, and characterized using SDS-PAGE, Western Blot, and Enzyme-Linked Immunosorbent Assay (ELISA). GNRs were fabricated on the electrode surface, followed by immobilization of RBD Ab. The conjugation steps were confirmed by UV-Vis Spectroscopy, Dynamic Light Scattering (DLS), Atomic Force Microscopy (AFM), Transmission Electron Microscopy (TEM), Cyclic Voltammetry (CV), and Differential Pulse Voltammetry (DPV). The fabricated electrode was further optimized for maximum efficiency and output. The detection limit of the developed electrode was determined as 0.73 fM for RBD antigen (Ag). Furthermore, the patient nasopharyngeal samples were collected in Viral Transport Media (VTM), and tested on the sensor surface that resulted in detection of SARS CoV-2 within 30 s, which was further validated via Reverse Transcription-Polymerase Chain Reaction (RT-PCR). Moreover, the immunosensor showed good repeatability, storage stability, and minimal cross reactivity against Middle East Respiratory Syndrome (MERS) spike protein. Along with ease of fabrication, the electrodes show future miniaturization potential for extensive and rapid screening of populations for COVID-19.


Subject(s)
Biosensing Techniques , COVID-19 , Nanotubes , Biosensing Techniques/methods , COVID-19/diagnosis , Carrier Proteins , Gold , Humans , Immunoassay/methods , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/analysis
3.
Emerg Microbes Infect ; 11(1): 1058-1071, 2022 Dec.
Article in English | MEDLINE | ID: covidwho-1752040

ABSTRACT

Safe, efficacious, and deployable vaccines are urgently needed to control COVID-19 in the large-scale vaccination campaigns. We report here the preclinical studies of an approved protein subunit vaccine against COVID-19, ZF2001, which contains tandem-repeat dimeric receptor-binding domain (RBD) protein with alum-based adjuvant. We assessed vaccine immunogenicity and efficacy in both mice and non-human primates (NHPs). ZF2001 induced high levels of RBD-binding and SARS-CoV-2 neutralizing antibody in both mice and non-human primates, and elicited balanced TH1/TH2 cellular responses in NHPs. Two doses of ZF2001 protected Ad-hACE2-transduced mice against SARS-CoV-2 infection, as detected by reduced viral RNA and relieved lung injuries. In NHPs, vaccination of either 25 µg or 50 µg ZF2001 prevented infection with SARS-CoV-2 in lung, trachea, and bronchi, with milder lung lesions. No evidence of disease enhancement was observed in both animal models. ZF2001 has been approved for emergency use in China, Uzbekistan, Indonesia, and Columbia. The high safety, immunogenicity, and protection efficacy in both mice and NHPs found in this preclinical study was consistent with the results in human clinical trials.


Subject(s)
COVID-19 Vaccines , COVID-19 , Animals , Antibodies, Neutralizing , Antibodies, Viral , COVID-19/prevention & control , Carrier Proteins , Humans , Immunogenicity, Vaccine , Mice , Mice, Inbred BALB C , Primates , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/genetics , Vaccines, Subunit
4.
Life Sci Alliance ; 5(6)2022 06.
Article in English | MEDLINE | ID: covidwho-1732625

ABSTRACT

Inflammasomes are cytosolic innate immune sensors of pathogen infection and cellular damage that induce caspase-1-mediated inflammation upon activation. Although inflammation is protective, uncontrolled excessive inflammation can cause inflammatory diseases and can be detrimental, such as in coronavirus disease (COVID-19). However, the underlying mechanisms that control inflammasome activation are incompletely understood. Here we report that the leucine-rich repeat (LRR) protein ribonuclease inhibitor (RNH1), which shares homology with LRRs of NLRP (nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain containing) proteins, attenuates inflammasome activation. Deletion of RNH1 in macrophages increases interleukin (IL)-1ß production and caspase-1 activation in response to inflammasome stimulation. Mechanistically, RNH1 decreases pro-IL-1ß expression and induces proteasome-mediated caspase-1 degradation. Corroborating this, mouse models of monosodium urate (MSU)-induced peritonitis and lipopolysaccharide (LPS)-induced endotoxemia, which are dependent on caspase-1, respectively, show increased neutrophil infiltration and lethality in Rnh1 -/- mice compared with wild-type mice. Furthermore, RNH1 protein levels were negatively related with disease severity and inflammation in hospitalized COVID-19 patients. We propose that RNH1 is a new inflammasome regulator with relevance to COVID-19 severity.


Subject(s)
COVID-19/pathology , Carrier Proteins/metabolism , Inflammasomes/metabolism , /metabolism , Animals , COVID-19/immunology , Caspase 1/metabolism , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , NF-kappa B/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Patient Acuity , Proteasome Endopeptidase Complex/metabolism
5.
Cells ; 11(4)2022 02 11.
Article in English | MEDLINE | ID: covidwho-1688673

ABSTRACT

Transmembrane proteins of adherens and tight junctions are known targets for viruses and bacterial toxins. The coronavirus receptor ACE2 has been localized at the apical surface of epithelial cells, but it is not clear whether ACE2 is localized at apical Cell-Cell junctions and whether it associates with junctional proteins. Here we explored the expression and localization of ACE2 and its association with transmembrane and tight junction proteins in epithelial tissues and cultured cells by data mining, immunoblotting, immunofluorescence microscopy, and co-immunoprecipitation experiments. ACE2 mRNA is abundant in epithelial tissues, where its expression correlates with the expression of the tight junction proteins cingulin and occludin. In cultured epithelial cells ACE2 mRNA is upregulated upon differentiation and ACE2 protein is widely expressed and co-immunoprecipitates with the transmembrane proteins ADAM17 and CD9. We show by immunofluorescence microscopy that ACE2 colocalizes with ADAM17 and CD9 and the tight junction protein cingulin at apical junctions of intestinal (Caco-2), mammary (Eph4) and kidney (mCCD) epithelial cells. These observations identify ACE2, ADAM17 and CD9 as new epithelial junctional transmembrane proteins and suggest that the cytokine-enhanced endocytic internalization of junction-associated protein complexes comprising ACE2 may promote coronavirus entry.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Intercellular Junctions/metabolism , Intercellular Junctions/virology , ADAM17 Protein/metabolism , Adherens Junctions/metabolism , Angiotensin-Converting Enzyme 2/genetics , Cadherins/metabolism , Carrier Proteins/metabolism , Cell Line , Cell Membrane Permeability , Coronavirus/metabolism , Epithelial Cells/metabolism , Epithelial Cells/virology , Gene Expression/genetics , Tetraspanin 29/metabolism , Tight Junction Proteins/metabolism , Tight Junctions/metabolism , Transcriptome/genetics
6.
J Biol Inorg Chem ; 27(2): 261-269, 2022 03.
Article in English | MEDLINE | ID: covidwho-1680896

ABSTRACT

Metal ions seem to play important roles in the pathogenesis of the novel coronavirus disease of 2019 (Covid-19) and are under investigation as potential prognostic markers and supplements in therapeutic procedures. The present study was aimed at assessing the relationship between the most abundant essential microelements (iron, zinc and copper) and their major binding proteins in the circulation in the early stage of infection. The concentration of zinc ions was measured to be higher in infected than in healthy persons, as well as ratios zinc/albumin and zinc/alpha-2-macroglobulin. Increased zinc levels could be attributed to cellular redistribution of zinc ions or to a use of zinc supplementation (zinc concentration was above the upper reference limit in one-third of infected individuals). Immunoblot analysis of protein molecular forms revealed that infected persons had greater amounts of proteinase-bound alpha-2-macroglobulin tetramer and albumin monomer than healthy individuals. The quantities of these forms were correlated with the concentration of zinc ions (r = 0.42 and 0.55, respectively) in healthy persons, but correlations were lost in infected individuals, most likely due to very high zinc concentrations in some participants which were not proportionally followed by changes in the distribution of protein species. Although we still have to wait for a firm confirmation of the involvement of zinc in beneficial defense mechanisms in patients with Covid-19, it seems that this ion may contribute to the existence of circulating protein forms which are the most optimal.


Subject(s)
COVID-19 , Carrier Proteins/genetics , Trace Elements , Copper , Humans , Iron , SARS-CoV-2 , Zinc
7.
Front Immunol ; 12: 828115, 2021.
Article in English | MEDLINE | ID: covidwho-1680008

ABSTRACT

Transient receptor potential vanilloid 4 (TRPV4) is a non-selective mechanosensitive ion channel expressed by various macrophage populations. Recent reports have characterized the role of TRPV4 in shaping the activity and phenotype of macrophages to influence the innate immune response to pathogen exposure and inflammation. TRPV4 has been studied extensively in the context of inflammation and inflammatory pain. Although TRPV4 activity has been generally described as pro-inflammatory, emerging evidence suggests a more complex role where this channel may also contribute to anti-inflammatory activities. However, detailed understanding of how TRPV4 may influence the initiation, maintenance, and resolution of inflammatory disease remains limited. This review highlights recent insights into the cellular processes through which TRPV4 contributes to pathological conditions and immune processes, with a focus on macrophage biology. The potential use of high-throughput and omics methods as an unbiased approach for studying the functional outcomes of TRPV4 activation is also discussed.


Subject(s)
Gene Expression Regulation , Macrophages/immunology , Macrophages/metabolism , Signal Transduction , TRPV Cation Channels/genetics , TRPV Cation Channels/metabolism , Animals , Carrier Proteins , Disease Management , Disease Susceptibility , Energy Metabolism , Humans , Ligands , Macrophage Activation/genetics , Macrophage Activation/immunology , Mechanotransduction, Cellular , Molecular Targeted Therapy , Protein Binding
9.
Front Immunol ; 12: 767347, 2021.
Article in English | MEDLINE | ID: covidwho-1528823

ABSTRACT

Infection with SARS-CoV-2 triggers the simultaneous activation of innate inflammatory pathways including the complement system and the kallikrein-kinin system (KKS) generating in the process potent vasoactive peptides that contribute to severe acute respiratory syndrome (SARS) and multi-organ failure. The genome of SARS-CoV-2 encodes four major structural proteins - the spike (S) protein, nucleocapsid (N) protein, membrane (M) protein, and the envelope (E) protein. However, the role of these proteins in either binding to or activation of the complement system and/or the KKS is still incompletely understood. In these studies, we used: solid phase ELISA, hemolytic assay and surface plasmon resonance (SPR) techniques to examine if recombinant proteins corresponding to S1, N, M and E: (a) bind to C1q, gC1qR, FXII and high molecular weight kininogen (HK), and (b) activate complement and/or the KKS. Our data show that the viral proteins: (a) bind C1q and activate the classical pathway of complement, (b) bind FXII and HK, and activate the KKS in normal human plasma to generate bradykinin and (c) bind to gC1qR, the receptor for the globular heads of C1q (gC1q) which in turn could serve as a platform for the activation of both the complement system and KKS. Collectively, our data indicate that the SARS-CoV-2 viral particle can independently activate major innate inflammatory pathways for maximal damage and efficiency. Therefore, if efficient therapeutic modalities for the treatment of COVID-19 are to be designed, a strategy that includes blockade of the four major structural proteins may provide the best option.


Subject(s)
Antigens, Viral/immunology , COVID-19/immunology , Complement System Proteins/immunology , Kallikrein-Kinin System , SARS-CoV-2/immunology , Viral Structural Proteins/immunology , Carrier Proteins/genetics , Carrier Proteins/immunology , Hemolysis , Humans , Mitochondrial Proteins/genetics , Mitochondrial Proteins/immunology , Recombinant Proteins/immunology , Viral Structural Proteins/genetics
10.
Bioessays ; 43(12): e2100158, 2021 12.
Article in English | MEDLINE | ID: covidwho-1525412

ABSTRACT

Severe COVID-19 is often accompanied by coagulopathies such as thrombocytopenia and abnormal clotting. Rarely, such complications follow SARS-CoV-2 vaccination. The cause of these coagulopathies is unknown. It is hypothesized that coagulopathies accompanying SARS-CoV-2 infections and vaccinations result from bacterial co-infections that synergize with virus-induced autoimmunity due to antigenic mimicry of blood proteins by both bacterial and viral antigens. Coagulopathies occur mainly in severe COVID-19 characterized by bacterial co-infections with Streptococci, Staphylococci, Klebsiella, Escherichia coli, and Acinetobacter baumannii. These bacteria express unusually large numbers of antigens mimicking human blood antigens, as do both SARS-CoV-2 and adenoviruses. Bacteria mimic cardiolipin, prothrombin, albumin, and platelet factor 4 (PF4). SARS-CoV-2 mimics complement factors, Rh antigens, platelet phosphodiesterases, Factors IX and X, von Willebrand Factor (VWF), and VWF protease ADAMTS13. Adenoviruses mimic prothrombin and platelet factor 4. Bacterial prophylaxis, avoidance of vaccinating bacterially infected individuals, and antigen deletion for vaccines may reduce coagulopathy risk. Also see the video abstract here: https://youtu.be/zWDOsghrPg8.


Subject(s)
COVID-19 , Coinfection , Autoantibodies , Autoimmunity , Bacteria , COVID-19 Vaccines , Cardiolipins , Carrier Proteins , Humans , Platelet Factor 4 , Prothrombin , SARS-CoV-2
11.
Clin Ther ; 43(10): 1706-1727, 2021 10.
Article in English | MEDLINE | ID: covidwho-1525737

ABSTRACT

PURPOSE: Neutralizing antibodies can reduce SARS-CoV-2 cellular entry, viral titers, and pathologic damage. CT-P59 (regdanvimab), a SARS-CoV-2 neutralizing monoclonal antibody, was examined in 2 randomized, double-blind, placebo-controlled, single ascending dose, Phase I studies. METHODS: In study 1.1, healthy adults were sequentially enrolled to receive CT-P59 10, 20, 40, or 80 mg/kg or placebo. In study 1.2, adult patients with mild SARS-CoV-2 infection were enrolled to receive CT-P59 20, 40, or 80 mg/kg or placebo. Primary objectives of both studies were safety and tolerability up to day 14 after infusion. Secondary end points included pharmacokinetic properties. Study 1.2 also measured virology and clinical efficacy. FINDINGS: Thirty-two individuals were randomized to study 1.1 (6 per CT-P59 dose cohort and 8 in the placebo cohort). By day 14 after infusion, adverse events (AEs) were reported in 2 individuals receiving CT-P59 20 mg/kg (headache and elevated C-reactive protein levels) and 1 receiving CT-P59 40 mg/kg (pyrexia) (all Common Terminology Criteria for Adverse Events grade 1). In study 1.2, 18 patients were randomized (5 per dose cohort and 3 in the placebo cohort). Sixteen AEs were reported in 10 patients receiving CT-P59. No AEs in either study led to study discontinuation. Greater reductions in viral titers were reported with CT-P59 than placebo in those with maximum titers >105 copies/mL. Mean time to recovery was 3.39 versus 5.25 days. IMPLICATIONS: CT-P59 exhibited a promising safety profile in healthy individuals and patients with mild SARS-CoV-2 infection, with potential antiviral and clinical efficacy in patients with mild SARS-CoV-2 infection. ClinicalTrials.gov identifier: NCT04525079 (study 1.1) and NCT04593641 (study 1.2).


Subject(s)
COVID-19 , SARS-CoV-2 , Adult , Antibodies, Monoclonal/adverse effects , Antibodies, Monoclonal, Humanized , Antibodies, Neutralizing , Carrier Proteins , Double-Blind Method , Humans , Immunoglobulin G
12.
Cell Rep ; 37(8): 110049, 2021 11 23.
Article in English | MEDLINE | ID: covidwho-1509642

ABSTRACT

Positive-strand RNA viruses replicate in close association with rearranged intracellular membranes. For hepatitis C virus (HCV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), these rearrangements comprise endoplasmic reticulum (ER)-derived double membrane vesicles (DMVs) serving as RNA replication sites. Cellular factors involved in DMV biogenesis are poorly defined. Here, we show that despite structural similarity of viral DMVs with autophagosomes, conventional macroautophagy is dispensable for HCV and SARS-CoV-2 replication. However, both viruses exploit factors involved in autophagosome formation, most notably class III phosphatidylinositol 3-kinase (PI3K). As revealed with a biosensor, PI3K is activated in cells infected with either virus to produce phosphatidylinositol 3-phosphate (PI3P) while kinase complex inhibition or depletion profoundly reduces replication and viral DMV formation. The PI3P-binding protein DFCP1, recruited to omegasomes in early steps of autophagosome formation, participates in replication and DMV formation of both viruses. These results indicate that phylogenetically unrelated HCV and SARS-CoV-2 exploit similar components of the autophagy machinery to create their replication organelles.


Subject(s)
Autophagy/physiology , Hepacivirus/physiology , SARS-CoV-2/physiology , Viral Replication Compartments/metabolism , Autophagosomes/metabolism , Carrier Proteins/metabolism , Class III Phosphatidylinositol 3-Kinases/antagonists & inhibitors , Class III Phosphatidylinositol 3-Kinases/metabolism , Humans , Phosphatidylinositol Phosphates/metabolism , RNA, Viral/biosynthesis , Viral Nonstructural Proteins/metabolism , Virus Replication
13.
Eur J Clin Microbiol Infect Dis ; 40(11): 2295-2303, 2021 Nov.
Article in English | MEDLINE | ID: covidwho-1479485

ABSTRACT

The aim of this study is to present the first nationwide microbiological and epidemiological study of invasive group A Streptococcus (iGAS) disease in Spain. One thousand eight hundred ninety-three iGAS isolates were analyzed over 2007-2019. emm typing was performed by sequencing the gene's variable 5' end, exotoxin genes were identified by PCR, and antimicrobial susceptibility explored via the E test and disk diffusion. Five hundred twenty-three isolates were associated with sepsis, 292 with cellulitis, 232 with scarlet fever, 153 with pneumonia, 141 with streptococcal toxic shock syndrome, and 94 with necrotizing fasciitis. The most prevalent emm types were emm1 (449/1893 isolates), emm89 (210/1893), emm3 (208/1893), emm4 (150/1893), emm12 (112/1893) emm6 (107/1893), emm87 (89/1893), emm28 (88/1893), emm75 (78/1893), emm77 (78/1893), emm11 (58/1893), and emm22 (35/1893). emm1, emm3, emm4, and emm6 were the predominant types affecting children (mostly respiratory infections), while emm11, emm77, and emm89 prevailed in the elderly (mostly skin infections). Each emm type was associated with one or more exotoxin gene (spe, sme, and ssa) profiles. speA was detected in 660 isolates, speB in 1829, speC in 1014, speF in 1826, speG in 1651, speJ in 716, speH in 331, smeZ in 720, and ssa in 512. Isolates with speA were associated with the most severe infections. Penicillin susceptibility was universal. Two hundred twenty-four isolates were resistant to tetracycline, 169 to erythromycin, and 81 to clindamycin. Tetracycline, erythromycin, and clindamycin resistance rates declined over the study period. The above information could serve as the basis for continued surveillance efforts designed to control disease cause by this bacterium.


Subject(s)
Streptococcal Infections/microbiology , Streptococcus pyogenes/isolation & purification , Adolescent , Adult , Aged , Aged, 80 and over , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Carrier Proteins/genetics , Carrier Proteins/metabolism , Child , Child, Preschool , Erythromycin/pharmacology , Exotoxins/genetics , Exotoxins/metabolism , Female , Humans , Infant , Male , Membrane Proteins/genetics , Membrane Proteins/metabolism , Microbial Sensitivity Tests , Middle Aged , Penicillins/pharmacology , Spain/epidemiology , Streptococcal Infections/epidemiology , Streptococcus pyogenes/classification , Streptococcus pyogenes/drug effects , Streptococcus pyogenes/genetics , Young Adult
14.
Front Immunol ; 12: 757691, 2021.
Article in English | MEDLINE | ID: covidwho-1463478

ABSTRACT

The increase in confirmed COVID-19 cases and SARS-CoV-2 variants calls for the development of safe and broad cross-protective vaccines. The RBD of the spike protein was considered to be a safe and effective candidate antigen. However, the low immunogenicity limited its application in vaccine development. Herein, we designed and obtained an RBD heptamer (mHla-RBD) based on a carrier protein-aided assembly strategy. The molecular weight of mHla-RBD is up to 450 kDa, approximately 10 times higher than that of the RBD monomer. When formulated with alum adjuvant, mHla-RBD immunization significantly increased the immunogenicity of RBD, as indicated by increased titers of RBD-specific antibodies, neutralizing antibodies, Th2 cellular immune response, and pseudovirus neutralization activity, when compared to RBD monomer. Furthermore, we confirmed that RBD-specific antibodies predominantly target conformational epitopes, which was approximately 200 times that targeting linear epitopes. Finally, a pseudovirus neutralization assay revealed that neutralizing antibodies induced by mHla-RBD against different SARS-CoV-2 variants were comparable to those against the wild-type virus and showed broad-spectrum neutralizing activity toward different SARS-CoV-2 variants. Our results demonstrated that mHla-RBD is a promising candidate antigen for development of SARS-CoV-2 vaccines and the mHla could serve as a universal carrier protein for antigen design.


Subject(s)
Bacterial Proteins/metabolism , COVID-19 Vaccines/immunology , COVID-19/immunology , Carrier Proteins/metabolism , Hemolysin Proteins/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/metabolism , Th2 Cells/immunology , Animals , Antibodies, Neutralizing/metabolism , Antibodies, Viral/metabolism , Broadly Neutralizing Antibodies/metabolism , Cell Line , Escherichia coli Proteins , Humans , Lymphocyte Activation , Mice , Mice, Inbred BALB C , Protein Domains/genetics , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology
15.
Clin Ther ; 43(10): 1706-1727, 2021 10.
Article in English | MEDLINE | ID: covidwho-1433085

ABSTRACT

PURPOSE: Neutralizing antibodies can reduce SARS-CoV-2 cellular entry, viral titers, and pathologic damage. CT-P59 (regdanvimab), a SARS-CoV-2 neutralizing monoclonal antibody, was examined in 2 randomized, double-blind, placebo-controlled, single ascending dose, Phase I studies. METHODS: In study 1.1, healthy adults were sequentially enrolled to receive CT-P59 10, 20, 40, or 80 mg/kg or placebo. In study 1.2, adult patients with mild SARS-CoV-2 infection were enrolled to receive CT-P59 20, 40, or 80 mg/kg or placebo. Primary objectives of both studies were safety and tolerability up to day 14 after infusion. Secondary end points included pharmacokinetic properties. Study 1.2 also measured virology and clinical efficacy. FINDINGS: Thirty-two individuals were randomized to study 1.1 (6 per CT-P59 dose cohort and 8 in the placebo cohort). By day 14 after infusion, adverse events (AEs) were reported in 2 individuals receiving CT-P59 20 mg/kg (headache and elevated C-reactive protein levels) and 1 receiving CT-P59 40 mg/kg (pyrexia) (all Common Terminology Criteria for Adverse Events grade 1). In study 1.2, 18 patients were randomized (5 per dose cohort and 3 in the placebo cohort). Sixteen AEs were reported in 10 patients receiving CT-P59. No AEs in either study led to study discontinuation. Greater reductions in viral titers were reported with CT-P59 than placebo in those with maximum titers >105 copies/mL. Mean time to recovery was 3.39 versus 5.25 days. IMPLICATIONS: CT-P59 exhibited a promising safety profile in healthy individuals and patients with mild SARS-CoV-2 infection, with potential antiviral and clinical efficacy in patients with mild SARS-CoV-2 infection. ClinicalTrials.gov identifier: NCT04525079 (study 1.1) and NCT04593641 (study 1.2).


Subject(s)
COVID-19 , SARS-CoV-2 , Adult , Antibodies, Monoclonal/adverse effects , Antibodies, Monoclonal, Humanized , Antibodies, Neutralizing , Carrier Proteins , Double-Blind Method , Humans , Immunoglobulin G
16.
Nat Commun ; 12(1): 5061, 2021 08 17.
Article in English | MEDLINE | ID: covidwho-1361634

ABSTRACT

The extent to which immune responses to natural infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and immunization with vaccines protect against variants of concern (VOC) is of increasing importance. Accordingly, here we analyse antibodies and T cells of a recently vaccinated, UK cohort, alongside those recovering from natural infection in early 2020. We show that neutralization of the VOC compared to a reference isolate of the original circulating lineage, B, is reduced: more profoundly against B.1.351 than for B.1.1.7, and in responses to infection or a single dose of vaccine than to a second dose of vaccine. Importantly, high magnitude T cell responses are generated after two vaccine doses, with the majority of the T cell response directed against epitopes that are conserved between the prototype isolate B and the VOC. Vaccination is required to generate high potency immune responses to protect against these and other emergent variants.


Subject(s)
COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/immunology , COVID-19/immunology , COVID-19/prevention & control , SARS-CoV-2/immunology , Angiotensin-Converting Enzyme 2/immunology , Animals , Antibodies, Monoclonal/blood , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/isolation & purification , Antibodies, Neutralizing/metabolism , Antibodies, Viral/blood , Antibodies, Viral/immunology , Carrier Proteins , Epitopes , Humans , Immunity , SARS-CoV-2/drug effects , T-Lymphocytes/immunology
17.
Proteins ; 90(1): 164-175, 2022 01.
Article in English | MEDLINE | ID: covidwho-1340286

ABSTRACT

TMEM106B is an integral membrane protein of late endosomes and lysosomes involved in neuronal function, its overexpression being associated with familial frontotemporal lobar degeneration, and point mutation linked to hypomyelination. It has also been identified in multiple screens for host proteins required for productive SARS-CoV-2 infection. Because standard approaches to understand TMEM106B at the sequence level find no homology to other proteins, it has remained a protein of unknown function. Here, the standard tool PSI-BLAST was used in a nonstandard way to show that the lumenal portion of TMEM106B is a member of the late embryogenesis abundant-2 (LEA-2) domain superfamily. More sensitive tools (HMMER, HHpred, and trRosetta) extended this to predict LEA-2 domains in two yeast proteins. One is Vac7, a regulator of PI(3,5)P2 production in the degradative vacuole, equivalent to the lysosome, which has a LEA-2 domain in its lumenal domain. The other is Tag1, another vacuolar protein, which signals to terminate autophagy and has three LEA-2 domains in its lumenal domain. Further analysis of LEA-2 structures indicated that LEA-2 domains have a long, conserved lipid-binding groove. This implies that TMEM106B, Vac7, and Tag1 may all be lipid transfer proteins in the lumen of late endocytic organelles.


Subject(s)
Carrier Proteins/metabolism , Membrane Proteins/chemistry , Membrane Proteins/metabolism , Nerve Tissue Proteins/chemistry , Nerve Tissue Proteins/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Computational Biology/methods , Cytoplasm/metabolism , Humans , Lysosomes , Membrane Glycoproteins/chemistry , Models, Molecular , Protein Conformation , Protein Domains , Saccharomyces cerevisiae Proteins/chemistry , Vacuoles/metabolism
18.
Front Immunol ; 12: 708149, 2021.
Article in English | MEDLINE | ID: covidwho-1337643

ABSTRACT

Microbial translocation (MT) and intestinal damage (ID) are poorly explored in COVID-19. Aims were to assess whether alteration of gut permeability and cell integrity characterize COVID-19 patients, whether it is more pronounced in severe infections and whether it influences the development of subsequent bloodstream infection (BSI). Furthermore, we looked at the potential predictive role of TM and ID markers on Intensive Care Unit (ICU) admission and in-hospital mortality. Over March-July 2020, 45 COVID-19 patients were enrolled. Markers of MT [LPB (Lipopolysacharide Binding Protein) and EndoCab IgM] and ID [I-FABP (Intestinal Fatty Acid Binding Protein)] were evaluated at COVID-19 diagnosis and after 7 days. As a control group, age- and gender-matched healthy donors (HDs) enrolled during the same study period were included. Median age was 66 (56-71) years. Twenty-one (46.6%) were admitted to ICU and mortality was 22% (10/45). Compared to HD, a high degree of MT and ID was observed. ICU patients had higher levels of MT, but not of ID, than non-ICU ones. Likewise, patients with BSI had lower EndoCab IgM than non-BSI. Interestingly, patients with high degree of MT and low ID were likely to be admitted to ICU (AUC 0.822). Patients with COVID-19 exhibited high level of MT, especially subjects admitted to ICU. COVID-19 is associated with gut permeability.


Subject(s)
COVID-19/metabolism , Intestinal Mucosa/metabolism , SARS-CoV-2/physiology , Acute-Phase Proteins/metabolism , Aged , Biomarkers/metabolism , COVID-19/diagnosis , COVID-19/mortality , COVID-19/pathology , Carrier Proteins/metabolism , Disease Progression , Fatty Acid-Binding Proteins/metabolism , Female , Humans , Intensive Care Units , Intestinal Mucosa/pathology , Male , Membrane Glycoproteins/metabolism , Middle Aged , Predictive Value of Tests , Prognosis , Survival Analysis , Tight Junctions/metabolism
19.
Eur J Med Res ; 26(1): 75, 2021 Jul 13.
Article in English | MEDLINE | ID: covidwho-1309928

ABSTRACT

BACKGROUND: The aim of this study was to evaluate the expression of four up/down-regulated inflammatory miRNAs and their mRNA targets in the serum samples of COVID-19 patients with different grades. Also, we investigated the relative expression of these miRNAs and mRNAs during hospitalization. METHODS: In this cross-sectional study, 5 mL of blood sample were taken from COVID-19 patients with different grades and during hospitalization from several health centers of Yazd, Tehran, and Zahedan province of Iran from December 20, 2020 to March 2, 2021. The relative expression of miRNAs and mRNAs was evaluated by q-PCR. RESULTS: We found that the relative expression of hsa-miR-31-3p, hsa-miR-29a-3p, and hsa-miR-126-3p was significantly decreased and the relative expression of their mRNA targets (ZMYM5, COL5A3, and CAMSAP1) was significantly increased with the increase of disease grade. Conversely, the relative expression of hsa-miR-17-3p was significantly increased and its mRNA target (DICER1) was significantly decreased with the increase of disease grade. This pattern was exactly seen during hospitalization of COVID-19 patients who did not respond to treatment. In COVID-19 patients who responded to treatment, the expression of selected miRNAs and their mRNA targets returned to the normal level. A negative significant correlation was seen between (1) the expression of hsa-miR-31-3p and ZMYM5, (2) hsa-miR-29a-3p and COL5A3, (3) hsa-miR-126-3p and CAMSAP1, and (4) hsa-miR-17-3p and DICER1 in COVID-19 patients with any grade (P < 0.05) and during hospitalization. CONCLUSIONS: In this study, we gained a more accurate understanding of the expression of up/down-regulated inflammatory miRNAs in the blood of COVID-19 patients. The obtained data may help us in the diagnosis and prognosis of COVID-19. TRIAL REGISTRATION: The ethics committee of Zahedan University of Medical Sciences, Zahedan, Iran. (Ethical Code: IR.ZAUMS.REC.1399.316) was registered for this project.


Subject(s)
COVID-19/genetics , Gene Expression Profiling , MicroRNAs/genetics , RNA, Messenger/genetics , COVID-19/blood , COVID-19/virology , Carrier Proteins/genetics , Collagen/genetics , Cross-Sectional Studies , DEAD-box RNA Helicases/genetics , Hospitalization/statistics & numerical data , Humans , Iran , Microtubule-Associated Proteins/genetics , Nuclear Proteins/genetics , Ribonuclease III/genetics , SARS-CoV-2/physiology , Severity of Illness Index
20.
Cell Host Microbe ; 29(7): 1137-1150.e6, 2021 07 14.
Article in English | MEDLINE | ID: covidwho-1252574

ABSTRACT

While the standard regimen of the BNT162b2 mRNA vaccine for SARS-CoV-2 includes two doses administered 3 weeks apart, some public health authorities are spacing these doses, raising concerns about efficacy. However, data indicate that a single dose can be up to 90% effective starting 14 days post-administration. To assess the mechanisms contributing to protection, we analyzed humoral and T cell responses three weeks after a single BNT162b2 dose. We observed weak neutralizing activity elicited in SARS-CoV-2 naive individuals but strong anti-receptor binding domain and spike antibodies with Fc-mediated effector functions and cellular CD4+ T cell responses. In previously infected individuals, a single dose boosted all humoral and T cell responses, with strong correlations between T helper and antibody immunity. Our results highlight the potential role of Fc-mediated effector functions and T cell responses in vaccine efficacy. They also provide support for spacing doses to vaccinate more individuals in conditions of vaccine scarcity.


Subject(s)
Antibodies, Viral/immunology , COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/immunology , COVID-19/immunology , SARS-CoV-2/immunology , T-Lymphocytes/immunology , Adult , Antibodies, Neutralizing/immunology , Antibodies, Viral/chemistry , Betacoronavirus , COVID-19/prevention & control , Carrier Proteins , Female , Humans , Immunity , Immunoglobulin Fc Fragments , Male , Middle Aged , Vaccination , Vaccines, Synthetic/immunology , Young Adult
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