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1.
Cells ; 11(1)2021 12 24.
Article in English | MEDLINE | ID: covidwho-1580994

ABSTRACT

The ongoing pandemic of coronavirus disease-2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), needs better treatment options both at antiviral and anti-inflammatory levels. It has been demonstrated that the aminothiol cysteamine, an already human applied drug, and its disulfide product of oxidation, cystamine, have anti-infective properties targeting viruses, bacteria, and parasites. To determine whether these compounds exert antiviral effects against SARS-CoV-2, we used different in vitro viral infected cell-based assays. Moreover, since cysteamine has also immune-modulatory activity, we investigated its ability to modulate SARS-CoV-2-specific immune response in vitro in blood samples from COVID-19 patients. We found that cysteamine and cystamine decreased SARS-CoV-2-induced cytopathic effects (CPE) in Vero E6 cells. Interestingly, the antiviral action was independent of the treatment time respect to SARS-CoV-2 infection. Moreover, cysteamine and cystamine significantly decreased viral production in Vero E6 and Calu-3 cells. Finally, cysteamine and cystamine have an anti-inflammatory effect, as they significantly decrease the SARS-CoV-2 specific IFN-γ production in vitro in blood samples from COVID-19 patients. Overall, our findings suggest that cysteamine and cystamine exert direct antiviral actions against SARS-CoV-2 and have in vitro immunomodulatory effects, thus providing a rational to test these compounds as a novel therapy for COVID-19.

2.
Front Mol Biosci ; 8: 752616, 2021.
Article in English | MEDLINE | ID: covidwho-1572298

ABSTRACT

Although lung fibrosis has a major impact in COVID-19 disease, its pathogenesis is incompletely understood. In particular, no direct evidence of pleura implication in COVID-19-related fibrotic damage has been reported so far. In this study, the expression of epithelial cytokeratins and Wilms tumor 1 (WT1), specific markers of mesothelial cells (MCs), was analyzed in COVID-19 and unrelated pleura autoptic samples. SARS-CoV-2 replication was analyzed by RT-PCR and confocal microscopy in MeT5A, a pleura MC line. SARS-CoV-2 receptors were analyzed by RT-PCR and western blot. Inflammatory cytokines from the supernatants of SARS-CoV-2-infected MeT5A cells were analysed by Luminex and ELLA assays. Immunohistochemistry of COVID-19 pleura patients highlighted disruption of pleura monolayer and fibrosis of the sub-mesothelial stroma, with the presence of MCs with fibroblastoid morphology in the sub-mesothelial stroma, but no evidence of direct infection in vivo. Interestingly, we found evidence of ACE2 expression in MCs from pleura of COVID-19 patients. In vitro analysis shown that MeT5A cells expressed ACE2, TMPRSS2, ADAM17 and NRP1, plasma membrane receptors implicated in SARS-CoV-2 cell entry and infectivity. Moreover, MeT5A cells sustained SARS-CoV-2 replication and productive infection. Infected MeT5A cells produced interferons, inflammatory cytokines and metalloproteases. Overall, our data highlight the potential role of pleura MCs as promoters of the fibrotic reaction and regulators of the immune response upon SARS-CoV-2 infection.

3.
Preprint in English | EuropePMC | ID: ppcovidwho-294112

ABSTRACT

Background: Data on SARS-CoV-2 vaccine immunogenicity in PLWH are currently limited. Aim of the study was to investigate immunogenicity according to current CD4 T-cell count and predictive role on immune response to vaccination in PLWH.<br><br>Methods: PLWH attending a single-center SARS-CoV-2 vaccination program in Italy, were included in a prospective evaluation for immunogenicity after receiving BNT162b2 or mRNA-1273. PLWH were stratified according to current CD4 T-cell count (severe immunodeficiency, SID: <200/mm 3 ;minor immunodeficiency, MID: 200-500/mm 3 ;no immunodeficiency, NID: >500/mm 3 ). RBD-binding IgG, SARS-CoV-2 neutralizing antibodies (nAbs) and cell-mediated (IFN-γ/IL-2) immune response were measured. As control group, not-matched HIV-negative healthcare workers (HCWs) were used.<br><br>Findings: Participants were 166 PLWH (SID=32;MID=56;NID=78) on ART. After 1 month from the booster dose, detectable RBD-binding IgG in 86.7% of SID, in 100% of MID, in 98.7% of NID (SID vs NID, p=0.021) and nAbs (titre ≥1:10) in 70.0%, 88.2% and 93.1%, respectively (SID vs NID, p=0.002), were elicited. Compared to NID, magnitude of anti-RBD, nAbs and IFN-γ production was significantly lower in SID and comparable in MID. After adjusting for confounders, current CD4 T-cell <200/mm 3 significantly predicted a poor magnitude of anti-RDB, nAbs and IFN-γ production. As compared with HCWs, SID elicited a consistently reduced immunogenicity for all parameters, MID only a reduced RBD-binding antibody response, NID a comparable response to HIV-negative controls for all parameters.<br><br>Interpretation: Neutralizing and cell-mediated immune response against SARS-CoV-2 were elicited in most of PLWH receiving ART, albeit significantly poorer in those with current CD4 T-cell <200/mm 3 versus those with CD4 T-cell >500/mm 3 and HIV-negative controls. A marginal decreased immunogenicity than HCWs was also observed in PLWH with CD4 T-cell 200-500/mm 3 , whereas immune response elicited in PLWH with a CD4 T-cell >500/mm 3 was comparable to HIV-negative population.<br><br>Funding: Italian Ministry of Health;European Commission, European Virus Archive – GLOBAL.<br><br>Declaration of Interest: None to declare. <br><br>Ethical Approval: The study was approved by the Scientific Committee of the Italian Drug Agency (AIFA) and by the Ethical Committee of the Lazzaro Spallanzani Institute, as National Review Board for COVID-19 pandemic in Italy (approval number 323/2021).

4.
Neurology ; 2021 Nov 22.
Article in English | MEDLINE | ID: covidwho-1528702

ABSTRACT

OBJECTIVE: To evaluate the immune-specific response after the full SARS-CoV-2 vaccination of multiple sclerosis (MS) patients treated with different Disease Modifying drugs by the detection of both serological- and T-cell responses. METHODS: Health care workers (HCWs) and MS patients, having completed the two-dose schedule of an mRNA-based vaccine against SARS-CoV-2 in the last 2-4 weeks, were enrolled from two parallel prospective studies conducted in Rome, Italy, at the National Institute for Infectious diseases Spallanzani-IRCSS and San Camillo Forlanini Hospital. Serological response was evaluated by quantifying the Region-Binding-Domain (RBD) and neutralizing-antibodies. Cell-mediated response was analyzed by a whole-blood test quantifying interferon (IFN)-γ response to spike peptides. Cells responding to spike stimulation were identified by FACS analysis. RESULTS: We prospectively enrolled 186 vaccinated individuals: 78 HCWs and 108 MS patients. Twenty-eight MS patients were treated with IFN-ß, 35 with fingolimod, 20 with cladribine, and 25 with ocrelizumab. A lower anti-RBD-antibody response rate was found in patients treated with ocrelizumab (40%, p<0.0001) and fingolimod (85.7%, p=0.0023) compared to HCWs and patients treated with cladribine or IFN-ß. Anti-RBD-antibody median titer was lower in patients treated with ocrelizumab (p<0.0001), fingolimod (p<0.0001) and cladribine (p=0.010) compared to HCWs and IFN-ß-treated patients. Importantly, serum neutralizing activity was present in all the HCWs tested and only in a minority of the fingolimod-treated patients (16.6%). T-cell-specific response was detected in the majority of MS patients (62%), albeit with significantly lower IFN-γ levels compared to HCWs. The lowest frequency of T-cell response was found in fingolimod-treated patients (14.3%). T-cell-specific response correlated with lymphocyte count and anti-RBD antibody titer (rho=0.554, p<0.0001 and rho=0.255, p=0.0078 respectively). Finally, IFN-γ T-cell response was mediated by both CD4+ and CD8+ T cells. CONCLUSION: mRNA vaccines induce both humoral and cell-mediated specific immune responses against spike peptides in all HCWs and in the majority of MS patients. These results carry relevant implications for managing vaccinations suggesting to promote vaccination in all treated MS patients. CLASSIFICATION OF EVIDENCE: This study provides Class III data that COVID mRNA vaccination induces both humoral and cell-mediated specific immune responses against viral spike proteins in a majority of MS patients.

5.
NPJ Vaccines ; 6(1): 131, 2021 Nov 04.
Article in English | MEDLINE | ID: covidwho-1503480

ABSTRACT

Here we report on the humoral and cellular immune response in eight volunteers who autonomously chose to adhere to the Italian national COVID-19 vaccination campaign more than 3 months after receiving a single-administration GRAd-COV2 vaccine candidate in the context of the phase-1 clinical trial. We observed a clear boost of both binding/neutralizing antibodies as well as T-cell responses upon receipt of the heterologous BNT162b2 or ChAdOx1-nCOV19 vaccines. These results, despite the limitation of the small sample size, support the concept that a single dose of an adenoviral vaccine may represent an ideal tool to effectively prime a balanced immune response, which can be boosted to high levels by a single dose of a different vaccine platform.

6.
Liver Int ; 2021 Oct 31.
Article in English | MEDLINE | ID: covidwho-1488231

ABSTRACT

Limited data are available on risks and benefits of anti-SARS-CoV2 vaccination in solid organ transplant recipients, and weaker responses have been described. At the Italian National Institute for Infectious Diseases, 61 liver transplant recipients underwent testing to describe the dynamics of humoral and cell-mediated immune response after two doses of anti-SARS-CoV2 mRNA vaccines and compared with 51 healthy controls. Humoral response was measured by quantifying both anti-spike and neutralizing antibodies; cell-mediated response was measured by PBMC proliferation assay with IFN-γ and IL-2 production. Liver transplant recipients showed lower response rates compared with controls in both humoral and cellular arms; shorter time since transplantation and multi-drug immunosuppressive regimen containing mycophenolate mofetil were predictive of reduced response to vaccination. Specific antibody and cytokine production, though reduced, were highly correlated in transplant recipients.

8.
JAMA Ophthalmol ; 139(9): 1041, 2021 09 01.
Article in English | MEDLINE | ID: covidwho-1453506

Subject(s)
COVID-19 , SARS-CoV-2 , Eye , Humans
9.
Mol Ther ; 2021 Sep 20.
Article in English | MEDLINE | ID: covidwho-1450246

ABSTRACT

The COVID-19 pandemic caused by SARS-CoV-2 has made the development of safe and effective vaccines a critical priority. To date, four vaccines have been approved by European and American authorities for preventing COVID-19, but the development of additional vaccine platforms with improved supply and logistics profiles remains a pressing need. Here we report the preclinical evaluation of a novel COVID-19 vaccine candidate based on the electroporation of engineered, synthetic cDNA encoding a viral antigen in the skeletal muscle. We constructed a set of prototype DNA vaccines expressing various forms of the SARS-CoV-2 spike (S) protein and assessed their immunogenicity in animal models. Among them, COVID-eVax-a DNA plasmid encoding a secreted monomeric form of SARS-CoV-2 S protein receptor-binding domain (RBD)-induced the most potent anti-SARS-CoV-2 neutralizing antibody responses (including against the current most common variants of concern) and a robust T cell response. Upon challenge with SARS-CoV-2, immunized K18-hACE2 transgenic mice showed reduced weight loss, improved pulmonary function, and lower viral replication in the lungs and brain. COVID-eVax conferred significant protection to ferrets upon SARS-CoV-2 challenge. In summary, this study identifies COVID-eVax as an ideal COVID-19 vaccine candidate suitable for clinical development. Accordingly, a combined phase I-II trial has recently started.

10.
Front Immunol ; 12: 740249, 2021.
Article in English | MEDLINE | ID: covidwho-1448730

ABSTRACT

Objective: To assess in rheumatoid arthritis (RA) patients, treated with different immunosuppressive therapies, the induction of SARS-CoV-2-specific immune response after vaccination in terms of anti-region-binding-domain (RBD)-antibody- and T-cell-specific responses against spike, and the vaccine safety in terms of clinical impact on disease activity. Methods: Health care workers (HCWs) and RA patients, having completed the BNT162b2-mRNA vaccination in the last 2 weeks, were enrolled. Serological response was evaluated by quantifying anti-RBD antibodies, while the cell-mediated response was evaluated by a whole-blood test quantifying the interferon (IFN)-γ-response to spike peptides. FACS analysis was performed to identify the cells responding to spike stimulation. RA disease activity was evaluated by clinical examination through the DAS28crp, and local and/or systemic clinical adverse events were registered. In RA patients, the ongoing therapeutic regimen was modified during the vaccination period according to the American College of Rheumatology indications. Results: We prospectively enrolled 167 HCWs and 35 RA patients. Anti-RBD-antibodies were detected in almost all patients (34/35, 97%), although the titer was significantly reduced in patients under CTLA-4-inhibitors (median: 465 BAU/mL, IQR: 103-1189, p<0.001) or IL-6-inhibitors (median: 492 BAU/mL, IQR: 161-1007, p<0.001) compared to HCWs (median: 2351 BAU/mL, IQR: 1389-3748). T-cell-specific response scored positive in most of RA patients [24/35, (69%)] with significantly lower IFN-γ levels in patients under biological therapy such as IL-6-inhibitors (median: 33.2 pg/mL, IQR: 6.1-73.9, p<0.001), CTLA-4-inhibitors (median: 10.9 pg/mL, IQR: 3.7-36.7, p<0.001), and TNF-α-inhibitors (median: 89.6 pg/mL, IQR: 17.8-224, p=0.002) compared to HCWs (median: 343 pg/mL, IQR: 188-756). A significant correlation between the anti-RBD-antibody titer and spike-IFN-γ-specific T-cell response was found in RA patients (rho=0.432, p=0.009). IFN-γ T-cell response was mediated by CD4+ and CD8+ T cells. Finally, no significant increase in disease activity was found in RA patients following vaccination. Conclusion: This study showed for the first time that antibody-specific and whole-blood spike-specific T-cell responses induced by the COVID-19 mRNA-vaccine were present in the majority of RA patients, who underwent a strategy of temporary suspension of immunosuppressive treatment during vaccine administration. However, the magnitude of specific responses was dependent on the immunosuppressive therapy administered. In RA patients, BNT162b2 vaccine was safe and disease activity remained stable.


Subject(s)
Antibodies, Viral/immunology , Arthritis, Rheumatoid/therapy , COVID-19 Vaccines/immunology , Immunotherapy/adverse effects , T-Lymphocytes/immunology , Aged , Arthritis, Rheumatoid/immunology , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/immunology , COVID-19/prevention & control , Female , Humans , Interferon-gamma/immunology , Lymphocyte Count , Male , Middle Aged , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/cytology , Vaccines, Synthetic/immunology
11.
Cells ; 10(10)2021 09 26.
Article in English | MEDLINE | ID: covidwho-1438527

ABSTRACT

Specific memory B cells and antibodies are a reliable read-out of vaccine efficacy. We analysed these biomarkers after one and two doses of BNT162b2 vaccine. The second dose significantly increases the level of highly specific memory B cells and antibodies. Two months after the second dose, specific antibody levels decline, but highly specific memory B cells continue to increase, thus predicting a sustained protection from COVID-19. We show that although mucosal IgA is not induced by the vaccination, memory B cells migrate in response to inflammation and secrete IgA at mucosal sites. We show that the first vaccine dose may lead to an insufficient number of highly specific memory B cells and low concentration of serum antibodies, thus leaving vaccinees without the immune robustness needed to ensure viral elimination and herd immunity. We also clarify that the reduction of serum antibodies does not diminish the force and duration of the immune protection induced by vaccination. The vaccine does not induce sterilizing immunity. Infection after vaccination may be caused by the lack of local preventive immunity because of the absence of mucosal IgA.


Subject(s)
Antibodies, Viral/immunology , B-Lymphocytes/cytology , COVID-19 Vaccines/therapeutic use , COVID-19/immunology , COVID-19/prevention & control , Immunoglobulin A/immunology , Immunologic Memory , Adult , Antibodies, Neutralizing/blood , Antigens, Viral/immunology , B-Lymphocytes/immunology , Cryopreservation , Female , Health Personnel , Healthy Volunteers , Hospitals, Pediatric , Humans , Immunoglobulin G , Immunoglobulin M/immunology , Lactation , Male , Middle Aged , Mucous Membrane/immunology , Patient Safety , SARS-CoV-2 , Vaccination
12.
Front Immunol ; 12: 690322, 2021.
Article in English | MEDLINE | ID: covidwho-1403471

ABSTRACT

A convalescent, non-severe, patient with COVID-19 was enrolled as a hyper-immune plasma voluntary donor by the Immuno-Hematology and Transfusion Unit of the Regina Elena National Cancer Institute in Rome, under the TSUNAMI national study criteria. During a nearly 6-month period (May-October 2020), the patient was closely monitored and underwent four hyperimmune plasma collections. Serum SARS-CoV-2 (anti-S + anti-N) IgG and IgM, anti-S1 IgA, and neutralizing titers (NTs) were measured. Anti-SARS-CoV-2 antibody levels steadily decreased. No correlation was found between anti-S/anti-N IgG and IgM levels and viral NT, measured by either a microneutralization test or the surrogate RBD/ACE2-binding inhibition test. Conversely, NTs directly correlated with anti-S1 IgA levels. Hyperimmune donor plasma, administered to five SARS-CoV-2 patients with persistent, severe COVID-19 symptoms, induced short-term clinical and pathological improvement. Reported data suggest that high NTs can persist longer than expected, thus widening hyperimmune plasma source, availability, and potential use. In vitro RBD/ACE2-binding inhibition test is confirmed as a convenient surrogate index for neutralizing activity and patients' follow-up, suitable for clinical settings where biosafety level 3 facilities are not available. IgA levels may correlate with serum neutralizing activity and represent a further independent index for patient evaluation.


Subject(s)
Antibodies, Neutralizing/administration & dosage , Antibodies, Viral/administration & dosage , COVID-19/therapy , SARS-CoV-2/immunology , Aged , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Blood Donors , COVID-19/immunology , COVID-19/virology , Humans , Immunization, Passive , Immunoglobulin A/administration & dosage , Immunoglobulin A/blood , Immunoglobulin A/immunology , Male , Middle Aged , Spike Glycoprotein, Coronavirus/immunology , Time Factors , Treatment Outcome
13.
Clin Chem Lab Med ; 59(12): 2010-2018, 2021 Nov 25.
Article in English | MEDLINE | ID: covidwho-1398962

ABSTRACT

OBJECTIVES: Simple and standardized methods to establish correlates to vaccine-elicited SARS-CoV-2 protection are needed. METHODS: An observational study on antibody response to a mRNA vaccine (Comirnaty) was performed on health care workers (V, n=120). Recovered COVID-19 patients (N, n=94) were used for comparison. Antibody response was evaluated by a quantitative anti-receptor binding domain IgG (anti-RBD) commercial assay and by virus microneutralization test (MNT), in order to establish a threshold of anti-RBD binding antibody units (BAU) able to predict a robust (≥1:80) MNT titer. RESULTS: Significant correlation between BAU and MNT titers was found in both V and N, being stronger in V (rs=0.91 and 0.57 respectively, p<0.001); a higher incremental trend starting from MNT titer 1:80 was observed in the V group. The 99% probability of high MNT titer (≥1:80) was reached at 1,814 and 3,564 BAU/mL, and the area under the receiver operating characteristic (ROC) curve was 0.99 (CI: 0.99-1.00) and 0.78 (CI: 0.67-0.86) in V and N, respectively. CONCLUSIONS: A threshold of 2,000 BAU/mL is highly predictive of strong MNT response in vaccinated individuals and may represent a good surrogate marker of protective response. It remains to be established whether the present results can be extended to BAU titers obtained with other assays.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , Immunity, Humoral , Vaccines, Synthetic/immunology , Adult , Aged , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Area Under Curve , COVID-19/immunology , COVID-19/virology , COVID-19 Vaccines/administration & dosage , Female , Health Personnel , Humans , Logistic Models , Male , Middle Aged , Neutralization Tests , ROC Curve , SARS-CoV-2/isolation & purification , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Vaccines, Synthetic/administration & dosage , Young Adult
14.
JAMA Ophthalmol ; 139(9): 1041, 2021 09 01.
Article in English | MEDLINE | ID: covidwho-1330280

Subject(s)
COVID-19 , SARS-CoV-2 , Eye , Humans
15.
Viruses ; 12(10)2020 10 20.
Article in English | MEDLINE | ID: covidwho-1305819

ABSTRACT

BACKGROUND: RT-PCR on nasopharyngeal (NPS)/oropharyngeal swabs is the gold standard for diagnosis of SARS-CoV-2 infection and viral load monitoring. Oral fluid (OF) is an alternate clinical sample, easy and safer to collect and could be useful for COVID-19 diagnosis, monitoring viral load and shedding. METHODS: Optimal assay conditions and analytical sensitivity were established for the commercial Simplexa™ COVID-19 Direct assay adapted to OF matrix. The assay was used to test 337 OF and NPS specimens collected in parallel from 164 hospitalized patients; 50 bronchoalveolar lavage (BAL) specimens from a subgroup of severe COVID-19 cases were also analysed. RESULTS: Using Simplexa™ COVID-19 Direct on OF matrix, 100% analytical detection down to 1 TCID50/mL (corresponding to 4 × 103 copies (cp)/mL) was observed. No crossreaction with other viruses transmitted through the respiratory toute was observed. Parallel testing of 337 OF and NPS samples showed highly concordant results (κ = 0.831; 95 % CI = 0.771-0.891), and high correlation of Ct values (r = 0.921; p < 0.0001). High concordance and elevated correlation was observed also between OF and BAL. Prolonged viral RNA shedding was observed up to 100 days from symptoms onset (DSO), with 32% and 29% positivity observed in OF and NPS samples, respectively, collected between 60 and 100 DSO. CONCLUSIONS: Simplexa™ COVID-19 Direct assays on OF have high sensitivity and specificity to detect SARS-CoV-2 RNA and provide an alternative to NPS for diagnosis and monitoring SARS-CoV-2 shedding.


Subject(s)
Betacoronavirus/physiology , Clinical Laboratory Techniques/methods , Coronavirus Infections/virology , Pneumonia, Viral/virology , Virus Shedding/physiology , Adult , Aged , Betacoronavirus/genetics , Body Fluids/virology , COVID-19 , COVID-19 Testing , COVID-19 Vaccines , Coronavirus Infections/diagnosis , Diagnostic Tests, Routine , Female , Humans , Male , Middle Aged , Molecular Diagnostic Techniques/methods , Pandemics , Pharynx/virology , Pneumonia, Viral/diagnosis , RNA, Viral/analysis , SARS-CoV-2 , Sensitivity and Specificity , Specimen Handling , Viral Load
16.
Biomed Res Int ; 2021: 8856018, 2021.
Article in English | MEDLINE | ID: covidwho-1303204

ABSTRACT

Coronaviruses (CoVs) are enveloped nonsegmented positive-sense RNA viruses belonging to the family Coronaviridae that contain the largest genome among RNA viruses. Their genome encodes 4 major structural proteins, and among them, the Spike (S) protein plays a crucial role in determining the viral tropism. It mediates viral attachment to the host cell, fusion to the membranes, and cell entry using cellular proteases as activators. Several in vitro models have been developed to study the CoVs entry, pathogenesis, and possible therapeutic approaches. This article is aimed at summarizing the current knowledge about the use of relevant methodologies and cell lines permissive for CoV life cycle studies. The synthesis of this information can be useful for setting up specific experimental procedures. We also discuss different strategies for inhibiting the binding of the S protein to the cell receptors and the fusion process which may offer opportunities for therapeutic intervention.


Subject(s)
Antiviral Agents , Coronaviridae , Models, Biological , Viral Tropism , Virus Internalization , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , COVID-19 , Cells, Cultured , Coronaviridae/drug effects , Coronaviridae/metabolism , Coronaviridae/pathogenicity , Coronaviridae/physiology , Coronaviridae Infections , Humans , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
17.
Int J Infect Dis ; 107: 247-250, 2021 Jun.
Article in English | MEDLINE | ID: covidwho-1300800

ABSTRACT

Prolonged B-cell depletion due to anti-CD20 monoclonal antibody (mAbs) therapy impairs the adaptive immune response, causing severe manifestations during COronaVIrus Disease-2019 (COVID-19). The cases of two patients under anti-CD20 therapy who experienced prolonged and severe COVID-19 successfully treated with mAbs against Severe Acute Respiratory Syndrome-CoV-2 spike proteins are reported.


Subject(s)
Antibodies, Monoclonal/therapeutic use , B-Lymphocytes/immunology , COVID-19/complications , Lymphocyte Depletion/adverse effects , SARS-CoV-2 , Antigens, CD20/immunology , COVID-19/drug therapy , Female , Humans , Male , Middle Aged , Severity of Illness Index
18.
Antibodies (Basel) ; 10(3)2021 Jul 05.
Article in English | MEDLINE | ID: covidwho-1295739

ABSTRACT

We report the isolation of two human IgG1k monoclonal antibodies (mAbs) directed against the SARS-CoV-2 spike protein. These mAbs were isolated from two donors who had recovered from COVID-19 infection during the first pandemic peak in the Lombardy region of Italy, the first European and initially most affected region in March 2020. We used the method of EBV immortalization of purified memory B cells and supernatant screening with a spike S1/2 assay for mAb isolation. This method allowed rapid isolation of clones, with one donor showing about 7% of clones positive against spike protein, whereas the other donor did not produce positive clones out of 91 tested. RNA was extracted from positive clones 39-47 days post-EBV infection, allowing VH and VL sequencing. The same clones were sequenced again after a further 100 days in culture, showing that no mutation had taken place during in vitro expansion. The B cell clones could be expanded in culture for more than 4 months after EBV immortalization and secreted the antibodies stably during that time, allowing to purify mg quantities of each mAb for functional assays without generating recombinant proteins. Unfortunately, neither mAb had significant neutralizing activity in a virus infection assay with several different SARS-CoV-2 isolates. The antibody sequences are made freely available.

20.
Microorganisms ; 9(6)2021 Jun 16.
Article in English | MEDLINE | ID: covidwho-1278501

ABSTRACT

Vaccination is the main public health measure to reduce SARS-CoV-2 transmission and hospitalization, and a massive worldwide scientific effort resulted in the rapid development of effective vaccines. This work aimed to define the dynamics of humoral and cell-mediated immune response in a cohort of health care workers (HCWs) who received a two-dose BNT162b2-mRNA vaccination. The serological response was evaluated by quantifying the anti-RBD and neutralizing antibodies. The cell-mediated response was performed by a whole blood test quantifying Th1 cytokines (IFN-γ, TNF-α, IL-2), produced in response to spike peptides. The BNT162b2-mRNA vaccine induced both humoral and cell-mediated immune responses against spike peptides in virtually all HCWs without previous SARS-CoV-2 infection, with a moderate inverse relation with age in the anti-RBD response. Spike-specific T cells produced several Th1 cytokines (IFN-γ, TNF-α, and IL-2), which correlated with the specific-serological response. Overall, our study describes the ability of the BNT162b2 mRNA vaccine to elicit a coordinated neutralizing humoral and spike-specific T cell response in HCWs. Assessing the dynamics of these parameters by an easy immune monitoring protocol can allow for the evaluation of the persistence of the vaccine response in order to define the optimal vaccination strategy.

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