Expression profile of HERVs and inflammatory mediators detected in nasal mucosa as a predictive biomarker of COVID-19 severity.

Introduction: Our research group and others demonstrated the implication of the human endogenous retroviruses (HERVs) in SARS-CoV-2 infection and their association with disease progression, suggesting HERVs as contributing factors in COVID-19 immunopathology. To identify early predictive biomarkers of the COVID-19 severity, we analyzed the expression of HERVs and inflammatory mediators in SARS-CoV-2-positive and -negative nasopharyngeal/oropharyngeal swabs with respect to biochemical parameters and clinical outcome. Methods: Residuals of swab samples (20 SARS-CoV-2-negative and 43 SARS-CoV-2-positive) were collected during the first wave of the pandemic and expression levels of HERVs and inflammatory mediators were analyzed by qRT-Real time PCR. Results: The results obtained show that infection with SARS-CoV-2 resulted in a general increase in the expression of HERVs and mediators of the immune response. In particular, SARS-CoV-2 infection is associated with increased expression of HERV-K and HERV-W, IL-1ß, IL-6, IL-17, TNF-α, MCP-1, INF-γ, TLR-3, and TLR-7, while lower levels of IL-10, IFN-α, IFN-ß, and TLR-4 were found in individuals who underwent hospitalization. Moreover, higher expression of HERV-W, IL-1ß, IL-6, IFN-α, and IFN-ß reflected the respiratory outcome of patients during hospitalization. Interestingly, a machine learning model was able to classify hospitalized vs not hospitalized patients with good accuracy based on the expression levels of HERV-K, HERV-W, IL-6, TNF-a, TLR-3, TLR-7, and the N gene of SARS-CoV-2. These latest biomarkers also correlated with parameters of coagulation and inflammation. Discussion: Overall, the present results suggest HERVs as contributing elements in COVID-19 and early genomic biomarkers to predict COVID-19 severity and disease outcome.

Rapid System to Detect Variants of SARS-CoV-2 in Nasopharyngeal Swabs.

Currently, the reference method for identifying the presence of variants of SARS-CoV-2 is whole genome sequencing. Although it is less expensive than in the past, it is still time-consuming, and interpreting the results is difficult, requiring staff with specific skills who are not always available in diagnostic laboratories. The test presented in this study aimed to detect, using traditional real-time PCR, the presence of the main variants described for the spike protein of the SARS-CoV-2 genome. The primers and probes were designed to detect the main deletions that characterize the different variants. The amplification targets were deletions in the S gene: 25-27, 69-70, 241-243, and 157-158. In the ORF1a gene, the deletion 3675-3677 was chosen. Some of these mutations can be considered specific variants, while others can be identified by the simultaneous presence of one or more deletions. We avoided using point mutations in order to improve the speed of the test. Our test can help clinical and medical microbiologists quickly recognize the presence of variants in biological samples (particularly nasopharyngeal swabs). The test can also be used to identify variants of the virus that could potentially be more diffusive as well as not responsive to the vaccine.

Thymosin alpha 1 restores the immune homeostasis in lymphocytes during Post-Acute sequelae of SARS-CoV-2 infection.

The complex alterations of the immune system and the immune-mediated multiorgan injury plays a key role in host response to SARS-CoV-2 infection and in the pathogenesis of COVID-19, being also associated with adverse outcomes. Thymosin alpha 1 (Tα1) is one of the molecules used in the treatment of COVID-19, as it is known to restore the homeostasis of the immune system during infections and cancer. The use of Tα1 in COVID-19 patients had been widely used in China and in COVID-19 patients, it has been shown to decrease hospitalization rate, especially in those with greater disease severity, and reduce mortality by restoring lymphocytopenia and more specifically, depleted T cells. Persistent dysregulation with depletion of naive B and T cell subpopulations and expansion of memory T cells suggest a chronic stimulation of the immune response in individuals with post-acute sequelae of SARS-CoV-2 infection (PASC). Our data obtained from an ex vivo study, showed that in PASC individuals with a chronically altered immune response, Tα1 improve the restoration of an appropriate response, most evident in those with more severe illness and who need respiratory support during acute phase, and in those with specific systemic and psychiatric symptoms of PASC, confirming Tα1 treatment being more effective in compromised patients. The results obtained, along with promising reports on recent trials on Tα1 administration in patients with COVID-19, offer new insights into intervention also for those patients with long-lasting inflammation with post-infectious symptoms, some of which have a delayed onset.

B- and T-Cell Responses After SARS-CoV-2 Vaccination in Patients With Multiple Sclerosis Receiving Disease Modifying Therapies: Immunological Patterns and Clinical Implications.

Background: Vaccination campaign to contrast the spread of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has raised the issue of vaccine immunogenicity in special populations such as people with multiple sclerosis (PwMS) on highly effective disease modifying treatments (DMTs). While humoral responses to SARS-CoV-2 mRNA vaccines have been well characterized in the general population and in PwMS, very little is known about cell-mediated responses in conferring protection from SARS-CoV-2 infection and severe coronavirus disease-2019 (COVID-19). Methods: PwMS on ocrelizumab, fingolimod or natalizumab, vaccinated with two doses of mRNABNT162b2 (Comirnaty®) vaccine were enrolled. Anti-Spike (S) and anti-Nucleoprotein (N) antibody titers, IFN-gamma production upon S and N peptide libraries stimulation, peripheral blood lymphocyte absolute counts were assessed after at least 1 month and within 4 months from vaccine second dose administration. A group of age and sex matched healthy donors (HD) were included as reference group. Statistical analysis was performed using GraphPad Prism 8.2.1. Results: Thirty PwMS and 9 HDs were enrolled. All the patients were negative for anti-N antibody detection, nor reported previous symptoms of COVID-19. Peripheral blood lymphocyte counts were assessed in PwMS showing: (i) reduction of circulating B-lymphocytes in PwMS on ocrelizumab; (ii) reduction of peripheral blood B- and T-lymphocyte absolute counts in PwMS on fingolimod and (iii) normal B- and T-lymphocyte absolute counts with an increase in circulating CD16+CD56+ NK-cells in PwMS on natalizumab. Three patterns of immunological responses were identified in PwMS. In patients on ocrelizumab, anti-S antibody were lacking or reduced, while T-cell responses were normal. In patients on fingolimod both anti-S titers and T-cell mediated responses were impaired. In patients on natalizumab both anti-S titers and T-cell responses were present and comparable to those observed in HD. Conclusions: The evaluation of T-cell responses, anti-S titers and peripheral blood lymphocyte absolute count in PwMS on DMTs can help to better characterize the immunological response after SARS-CoV-2 vaccination. The evaluation of T-cell responses in longitudinal cohorts of PwMS will help to clarify their protective role in preventing SARS-CoV-2 infection and severe COVID-19. The correlation between DMT treatment and immunological responses to SARS-CoV-2 vaccines could help to better evaluate vaccination strategies in PwMS.

Multidistrict Host-Pathogen Interaction during COVID-19 and the Development Post-Infection Chronic Inflammation.

Due to the presence of the ACE2 receptor in different tissues (nasopharynx, lung, nervous tissue, intestine, liver), the COVID-19 disease involves several organs in our bodies. SARS-CoV-2 is able to infect different cell types, spreading to different districts. In the host, an uncontrolled and altered immunological response is triggered, leading to cytokine storm, lymphopenia, and cellular exhaustion. Hence, respiratory distress syndrome (ARDS) and systemic multi-organ dysfunction syndrome (MODS) are established. This scenario is also reflected in the composition of the microbiota, the balance of which is regulated by the interaction with the immune system. A change in microbial diversity has been demonstrated in COVID-19 patients compared with healthy donors, with an increase in potentially pathogenic microbial genera. In addition to other symptoms, particularly neurological, the occurrence of dysbiosis persists after the SARS-CoV-2 infection, characterizing the post-acute COVID syndrome. This review will describe and contextualize the role of the immune system in unbalance and dysbiosis during SARS-CoV-2 infection, from the acute phase to the post-COVID-19 phase. Considering the tight relationship between the immune system and the gut-brain axis, the analysis of new, multidistrict parameters should be aimed at understanding and addressing chronic multisystem dysfunction related to COVID-19.

Expression analysis of miRNA hsa-let7b-5p in naso-oropharyngeal swabs of COVID-19 patients supports its role in regulating ACE2 and DPP4 receptors.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is the novel coronavirus responsible for worldwide coronavirus disease (COVID-19). We previously observed that Angiotensin-converting enzyme 2 (ACE2) and Dipeptidyl peptidase-4 (DPP4) are significantly overexpressed in naso-oropharyngeal swabs (NPS) of COVID-19 patients, suggesting their putative functional role in the disease progression. ACE2 and DPP4 overexpression in COVID-19 patients may be associated to epigenetic mechanism, such as miRNA differential expression. We investigated if hsa-let7b-5p, reported to target both ACE2 and DPP4 transcripts, could be involved in the regulation of these genes. We verified that the inhibition and overexpression of hsa-let7b-5p matched to a modulation of both ACE2 and DPP4 levels. Then, we observed a statistically significant downregulation (FC = -1.5; p < 0.05) of hsa-let7b-5p in the same COVID-19 and control samples of our previous study. This is the first study that shows hsa-let7b-5p low expression in naso-oropharyngeal swabs of COVID-19 patients and demonstrates a functional role of this miR in regulating ACE2 and DPP4 levels. These data suggest the involvement of hsa-let7b-5p in the regulation of genes necessary for SARS-CoV-2 infections and its putative role as a therapeutic target for COVID-19.

Increased Mild Vaccine-Related Side Effects and Higher Specific Antibody Titers in Health Care Workers with Previous SARS-CoV-2 Infection after the mRNA BNT162b2 Vaccine.

BACKGROUND: to evaluate whether prior SARS-CoV-2 infection affects side effects and specific antibody production after vaccination with BNT162b2. METHODS: We included 1106 health care workers vaccinated with BNT162b2. We assessed whether prior SARS-CoV-2 infection affects the number and type of side effects and performed a nested case-control analysis comparing plasma levels of specific IgG titers between SARS-CoV-2-naïve and previously infected subjects after the first and the second vaccine doses. RESULTS: After the first dose, SARS-CoV-2-naïve subjects experienced side effects more often than SARS-CoV-2 naïve subjects. Individuals with prior SARS-CoV-2 infection more often reported pain at the injection site, weakness, and fever than SARS-CoV-2-naïve subjects. After the second dose, the frequency of side effects was similar in the two groups. All subjects with prior SARS-CoV-2 infection developed either a high (>100 AU/mL) or intermediate (10-100 AU/mL) antibody titer. Among SARS-CoV-2-naïve subjects, the majority developed an intermediate titer. After the second dose, a high (>2000 AU/mL) antibody titer was more common among subjects with prior SARS-CoV-2 infection. CONCLUSIONS: vaccine-related side effects and a higher anti-SARS-CoV-2-RBD IgG titer were more common in subjects with previous infection than in SARS-CoV-2-naïve after the first, but not after the second dose of the BNT162b2 vaccine.

Infection Rate of Respiratory Viruses in the Pandemic SARS-CoV-2 Period Considering Symptomatic Patients: Two Years of Ongoing Observations.

BACKGROUND: In the last two years, the SARS-CoV-2 pandemic has determined radical changes in human behaviors and lifestyles, with a drastic reduction in socialization due to physical distancing and self-isolation. These changes have also been reflected in the epidemiological patterns of common respiratory viruses. For this reason, early discrimination of respiratory viruses is important as new variants emerge. METHODS: Nasopharyngeal swabs of 2554 patients, with clinically suspected Acute Respiratory Infections (ARIs) from October 2019 to November 2021, were collected to detect 1 or more of the 23 common respiratory pathogens, especially viruses, via BioFilmArray RP2.1plus, including SARS-CoV-2. Demographical characteristics and epidemiological analyses were performed as well as a laboratory features profile of positive patients. RESULTS: An observational study on 2300 patients (254 patients were excluded because of missing data) including 1560 men and 760 women, median age of 64.5 years, was carried out. Considering the respiratory virus research request, most of the patients were admitted to the Emergency Medicine Department (41.2%, of patients), whereas 29.5% were admitted to the Infectious Diseases Department. The most frequently detected pathogens included SARS-CoV-2 (31.06%, 707/2300, from March 2020 to November 2021), InfA-B (1.86%, 43/2300), HCoV (2.17% 50/2300), and HSRV (1.65%, 38/2300). Interestingly, coinfection rates decreased dramatically in the SARS-CoV-2 pandemic period. The significative decrease in positive rate of SARS-CoV-2 was associated with the massive vaccination. CONCLUSION: This study represents a dynamic picture of the epidemiological curve of common respiratory viruses during the two years of pandemic, with a disregarded trend for additional viruses. Our results showed that SARS-CoV-2 had a preferential tropism for the respiratory tract without co-existing with other viruses. The possible causes were attributable either to the use of masks, social isolation, or to specific respiratory receptors mostly available for this virus, external and internal lifestyle factors, vaccination campaigns, and emergence of new SARS-CoV-2 variants.

Interaction of Pelargonium sidoides Compounds with Lactoferrin and SARS-CoV-2: Insights from Molecular Simulations.

(1) Background: Pelargonium sidoides extracts and lactoferrin are two important natural, anti-inflammatory, and antiviral agents, which can interfere with the early stages of SARS-CoV-2 infection. Molecular docking and molecular dynamics simulation approaches have been applied to check for the occurrence of interactions of the Pelargonium sidoides compounds with lactoferrin and with SARS-CoV-2 components. (2) Methods: Computational methods have been applied to confirm the hypothesis of a direct interaction between PEL compounds and the lactoferrin protein and between Pelargonium sidoides compounds and SARS-CoV-2 Spike, 3CLPro, RdRp proteins, and membrane. Selected high-score complexes were structurally investigated through classical molecular dynamics simulation, while the interaction energies were evaluated using the molecular mechanics energies combined with generalized Born and surface area continuum solvation method. (3) Results: Computational analyses suggested that Pelargonium sidoides extracts can interact with lactoferrin without altering its structural and dynamical properties. Furthermore, Pelargonium sidoides compounds should have the ability to interfere with the Spike glycoprotein, the 3CLPro, and the lipid membrane, probably affecting the functional properties of the proteins inserted in the double layer. (4) Conclusion: Our findings suggest that Pelargonium sidoides may interfere with the mechanism of infection of SARS-CoV-2, especially in the early stages.

B- and T-Cell Responses After SARS-CoV-2 Vaccination in Patients With Multiple Sclerosis Receiving Disease Modifying Therapies: Immunological Patterns and Clinical Implications

Background Vaccination campaign to contrast the spread of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has raised the issue of vaccine immunogenicity in special populations such as people with multiple sclerosis (PwMS) on highly effective disease modifying treatments (DMTs). While humoral responses to SARS-CoV-2 mRNA vaccines have been well characterized in the general population and in PwMS, very little is known about cell-mediated responses in conferring protection from SARS-CoV-2 infection and severe coronavirus disease-2019 (COVID-19). Methods PwMS on ocrelizumab, fingolimod or natalizumab, vaccinated with two doses of mRNABNT162b2 (Comirnaty®) vaccine were enrolled. Anti-Spike (S) and anti-Nucleoprotein (N) antibody titers, IFN-gamma production upon S and N peptide libraries stimulation, peripheral blood lymphocyte absolute counts were assessed after at least 1 month and within 4 months from vaccine second dose administration. A group of age and sex matched healthy donors (HD) were included as reference group. Statistical analysis was performed using GraphPad Prism 8.2.1. Results Thirty PwMS and 9 HDs were enrolled. All the patients were negative for anti-N antibody detection, nor reported previous symptoms of COVID-19. Peripheral blood lymphocyte counts were assessed in PwMS showing: (i) reduction of circulating B-lymphocytes in PwMS on ocrelizumab;(ii) reduction of peripheral blood B- and T-lymphocyte absolute counts in PwMS on fingolimod and (iii) normal B- and T-lymphocyte absolute counts with an increase in circulating CD16+CD56+ NK-cells in PwMS on natalizumab. Three patterns of immunological responses were identified in PwMS. In patients on ocrelizumab, anti-S antibody were lacking or reduced, while T-cell responses were normal. In patients on fingolimod both anti-S titers and T-cell mediated responses were impaired. In patients on natalizumab both anti-S titers and T-cell responses were present and comparable to those observed in HD. Conclusions The evaluation of T-cell responses, anti-S titers and peripheral blood lymphocyte absolute count in PwMS on DMTs can help to better characterize the immunological response after SARS-CoV-2 vaccination. The evaluation of T-cell responses in longitudinal cohorts of PwMS will help to clarify their protective role in preventing SARS-CoV-2 infection and severe COVID-19. The correlation between DMT treatment and immunological responses to SARS-CoV-2 vaccines could help to better evaluate vaccination strategies in PwMS.

First Case of a COVID-19 Patient Infected by Delta AY.4 with a Rare Deletion Leading to a N Gene Target Failure by a Specific Real Time PCR Assay: Novel Omicron VOC Might Be Doing Similar Scenario?

Herein, we report a case of an Italian male infected by Delta sublineage AY.4 harboring an atypical deletion, leading to a N gene target failure (NGTF) by a commercial molecular assay for SARS-CoV-2 diagnosis (AllplexTM SARS-CoV-2 Assay, Seegene). A 59-year-old unvaccinated patient was hospitalized for pulmonary embolism, with first negative results obtained by both molecular and antigen tests. After several days of viral negativity, he presented positive results for E and RdRP/S genes, but negative in N gene. Negativity in N gene was repeatedly confirmed in the following days. Suspecting an infection by the Omicron variant, SARS-CoV-2 genome sequencing was rapidly performed from nasopharyngeal swab by MiSeq and revealed the presence of the Delta sublineage AY.4 variant with an atypical deletion of six nucleotides, leading to G214-G215 deletion in the Nucleocapsid, thus responsible for NGTF. The analysis of GISAID sequences (N = 2,618,373 12 January 2022) showed that G214-G215 deletion is rarely occurring in most circulating Delta lineages and sublineages in the globe and Europe, with an overall prevalence never exceeding 0.2%. Hence, this study highlights the importance to perform SARS-CoV-2 sequencing and to characterize novel mutations/deletions that could jeopardize the proper interpretation of molecular diagnostic tests. Based on these assumptions, the role of deletions in the recently identified Omicron variant deserves further investigation.

Reduced Titers of Circulating Anti-SARS-CoV-2 Antibodies and Risk of COVID-19 Infection in Healthcare Workers during the Nine Months after Immunization with the BNT162b2 mRNA Vaccine.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has had a tremendous impact on health services; hundreds of thousands of healthcare workers (HCWs) have died from coronavirus disease 2019 (COVID-19). The introduction of the BNT162b2 mRNA vaccine in Italy provided recipients with significant protection against COVID-19 within one to two weeks after the administration of the second of the two recommended doses. While the vaccine induces a robust T cell response, the protective role of factors and pathways other than those related to memory B cell responses to specific SARS-CoV-2 antigens remains unclear. This retrospective study aimed to evaluate the determinants of serological protection in a group of vaccinated HCWs (n = 793) by evaluating circulating levels of antiviral spike receptor-binding domain (S-RBD) antibodies during the nine-month period following vaccination. We found that 99.5% of the HCWs who received the two doses of the BNT162b2 vaccine developed protective antibodies that were maintained at detectable levels for as long as 250 days after the second dose of the vaccine. Multivariate analysis was performed on anti-S-RBD titers in a subgroup of participants (n = 173) that were evaluated twice during this period. The results of this analysis reveal that the antibody titer observed at the second time point was significantly related to the magnitude of the primary response, the time that had elapsed between the first and the second evaluation, and a previous history of SARS-CoV-2 infection. Of importance is the finding that despite waning antibody titers following vaccination, none of the study participants contracted severe COVID-19 during the observational period.

High CD169 Monocyte/Lymphocyte Ratio Reflects Immunophenotype Disruption and Oxygen Need in COVID-19 Patients.

BACKGROUND: Sialoadhesin (CD169) has been found to be overexpressed in the blood of COVID-19 patients and identified as a biomarker in early disease. We analyzed CD169 in the blood cells of COVID-19 patients to assess its role as a predictive marker of disease progression and clinical outcomes. METHODS: The ratio of the median fluorescence intensity of CD169 between monocytes and lymphocytes (CD169 RMFI) was analyzed by flow cytometry in blood samples of COVID-19 patients (COV) and healthy donors (HDs) and correlated with immunophenotyping, inflammatory markers, cytokine mRNA expression, pulmonary involvement, and disease progression. RESULTS: CD169 RMFI was high in COV but not in HDs, and it correlated with CD8 T-cell senescence and exhaustion markers, as well as with B-cell maturation and differentiation in COV. CD169 RMFI correlated with blood cytokine mRNA levels, inflammatory markers, and pneumonia severity in patients who were untreated at sampling, and was associated with the respiratory outcome throughout hospitalization. Finally, we also report the first evidence of the specific ability of the spike protein of SARS-CoV-2 to trigger CD169 RMFI in a dose-dependent manner in parallel with IL-6 and IL-10 gene transcription in HD PBMCs stimulated in vitro. CONCLUSION: CD169 is induced by the spike protein and should be considered as an early biomarker for evaluating immune dysfunction and respiratory outcomes in COVID-19 patients.

Post-Mortem RT-PCR Assay for SARS-CoV-2 RNA in COVID-19 Patients' Corneal Epithelium, Conjunctival and Nasopharyngeal Swabs.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) disease has been described to possibly be associated with ocular surface disturbances. However, whether the virus could invade ocular tissues still remains elusive. In the present study, we tried to investigate the post-mortem presence of SARS-CoV-2 RNA in corneal epithelium gathered by patients with an ante-mortem confirmed diagnosis of Coronavirus disease-19 (COVID-19). Cadavers with an ante-mortem confirmed diagnosis of moderate to severe COVID-19 were examined. Clinical and demographic features were retrieved from hospital patients' notes. For each cadaver, corneal scrapings, conjunctival swabs (CS) and nasopharyngeal swabs (NPS) were collected to perform real-time reverse transcriptase polymerase chain reaction ((RT)-PCR) for SARS-CoV-2. Fourteen consecutive cadavers with an ante-mortem confirmed diagnosis of moderate to severe COVID-19 were examined. The last NPS performed ante-mortem confirmed SARS-CoV-2 infection in 12/14 (85.7%) patients. The mean death-to-swab time (DtS) was 3.15 ± 0.5 (2.10-5.1) h. The post-mortem NPS and CS found positive for SARS-CoV-2 RNA were 9/14 (64.3%) and 3/28 (10.7%), respectively. None of the corneal epithelium scrapes tested positive to RT-PCR for SARS-CoV-2 RNA. These data promote the SARS-CoV-2 as not able to contaminate the post-mortem corneal epithelium, while it can persist in different other structures of the ocular surface (i.e., the conjunctiva). It is reasonable to assume that such a contamination can occur ante-mortem too.

T-cell responses to SARS-CoV-2 in multiple sclerosis patients treated with ocrelizumab healed from COVID-19 with absent or low anti-spike antibody titers.

BACKGROUND: Disease modifying therapies for multiple sclerosis (MS) can impair the specific immune response to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Specifically, it is recognized that ocrelizumab reduces or abrogates anti-SARS-CoV-2 antibody production after natural infection or vaccination, while very little is known about T-cell responses. METHODS: We developed an interferon (IFN)-γ release assay (IGRA) to detect T-cell responses specific to SARS-CoV-2 after overnight stimulation of whole blood with peptide libraries covering the immunodominant sequence domains of the Spike glycoprotein (S) and the Nucleocapsid phosphoprotein (N). RESULTS: Five patients with MS receiving ocrelizumab treatment for at least 1 year and recovered from SARS-CoV-2 infection were enrolled in the study. Despite the absence or the very low concentration of anti-S antibodies, a T-cell response was detectable in all the five MS patients. These results are in accordance with the marked reduction of peripheral B-lymphocyte absolute counts induced by ocrelizumab, that, conversely, did not affect peripheral blood T-lymphocyte subset absolute and relative counts and CD4/CD8 ratio. CONCLUSIONS: The detection of specific T-cell responses to SARS-CoV-2 in patients receiving B-cell depleting therapies represents a useful tool to improve the diagnostic approach in SARS-CoV-2 infection and to accurately assess the immunological response after natural infection or vaccination.

Evidence of the pathogenic HERV-W envelope expression in T lymphocytes in association with the respiratory outcome of COVID-19 patients.

BACKGROUND: Despite an impressive effort in clinical research, no standard therapeutic approach for coronavirus disease 2019 (COVID-19) patients has been established, highlighting the need to identify early biomarkers for predicting disease progression and new therapeutic interventions for patient management. The present study aimed to evaluate the involvement of the human endogenous retrovirus -W envelope (HERV-W ENV) in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection considering recent findings that HERVs are activated in response to infectious agents and lead to various immunopathological effects. We analysed HERV-W ENV expression in blood cells of COVID-19 patients in correlation with clinical characteristics and have discussed its potential role in the outcome of the disease. METHODS: We analysed HERV-W ENV expression in blood samples of COVID-19 patients and healthy donors by flow cytometry and quantitative reverse transcriptase PCR analysis, and evaluated its correlation with clinical signs, inflammatory markers, cytokine expression, and disease progression. FINDINGS: HERV-W ENV was highly expressed in the leukocytes of COVID-19 patients but not in those of healthy donors. Its expression correlated with the markers of T-cell differentiation and exhaustion and blood cytokine levels. The percentage of HERV-W ENV-positive lymphocytes correlated with inflammatory markers and pneumonia severity in COVID-19 patients. Notably, HERV-W ENV expression reflects the respiratory outcome of patients during hospitalization. INTERPRETATION: Given the known immuno- and neuro-pathogenicity of HERV-W ENV protein, it could promote certain pathogenic features of COVID-19 and therefore serve as a biomarker to predict clinical progression of disease and open to further studies for therapeutic intervention. FUNDING: Information available at the end of the manuscript.

Inhibition of HECT E3 ligases as potential therapy for COVID-19.

SARS-CoV-2 is responsible for the ongoing world-wide pandemic which has already taken more than two million lives. Effective treatments are urgently needed. The enzymatic activity of the HECT-E3 ligase family members has been implicated in the cell egression phase of deadly RNA viruses such as Ebola through direct interaction of its VP40 Protein. Here we report that HECT-E3 ligase family members such as NEDD4 and WWP1 interact with and ubiquitylate the SARS-CoV-2 Spike protein. Furthermore, we find that HECT family members are overexpressed in primary samples derived from COVID-19 infected patients and COVID-19 mouse models. Importantly, rare germline activating variants in the NEDD4 and WWP1 genes are associated with severe COVID-19 cases. Critically, I3C, a natural NEDD4 and WWP1 inhibitor from Brassicaceae, displays potent antiviral effects and inhibits viral egression. In conclusion, we identify the HECT family members of E3 ligases as likely novel biomarkers for COVID-19, as well as new potential targets of therapeutic strategy easily testable in clinical trials in view of the established well-tolerated nature of the Brassicaceae natural compounds.

Thymosin Alpha 1 Mitigates Cytokine Storm in Blood Cells From Coronavirus Disease 2019 Patients.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is characterized by immune-mediated lung injury and complex alterations of the immune system, such as lymphopenia and cytokine storm, that have been associated with adverse outcomes underlining a fundamental role of host response in severe acute respiratory syndrome coronavirus 2 infection and the pathogenesis of the disease. Thymosin alpha 1 (Tα1) is one of the molecules used in the management of COVID-19, because it is known to restore the homeostasis of the immune system during infections and cancer. METHODS: In this study, we captured the interconnected biological processes regulated by Tα1 in CD8+ T cells under inflammatory conditions. RESULTS: Genes associated with cytokine signaling and production were upregulated in blood cells from patients with COVID-19, and the ex vivo treatment with Tα1-mitigated cytokine expression, and inhibited lymphocyte activation in a CD8+ T-cell subset specifically. CONCLUSION: These data suggest the potential role of Tα1 in modulating the immune response homeostasis and the cytokine storm in vivo.

Clinical validation of a second generation anti-SARS-CoV-2 IgG and IgM automated chemiluminescent immunoassay.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has proven to be extremely contagious and has spread rapidly all over the world. A key aspect in limiting the virus diffusion is to ensure early and accurate diagnosis. Serological assays could be an alternative in increasing testing capabilities, particularly when used as part of an algorithmic approach combined with molecular analysis. The aim of this study was to evaluate the diagnostic accuracy of a second generation chemiluminescent automated immunoassay able to detect anti-SARS-CoV-2 immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies. Data are carried out on healthy subjects and other infectious diseases pre-pandemic sera, as controls, and on two different coronavirus disease 2019 hospitalized patient groups (early and late infection time). Data obtained have been analyzed in terms of precision, linearity, sensitivity and specificity. Specificities are: 100% for anti-SARS-CoV-2 IgG and 98% for anti-SARS-CoV-2 IgM, in all patient groups. Sensitivities are: 97%, 100%, and 98% for anti-SARS-CoV-2 IgG and 87%, 83%, and 86% for anti-SARS-CoV-2 IgM in the early infection, in the late infection and in the total patient group, respectively. The Mindray anti-SARS-CoV-2 IgG and IgM assays demonstrated higher sensitivity and specificity, indicating that IgG and IgM simultaneous detection is useful even in the early phases of infection.

Breast cancer patients with hormone neoadjuvant bridging therapy due to asymptomatic Corona virus infection. Case report, clinical and histopathologic findings.

INTRODUCTION: Breast cancer management during COVID-19 pandemic has changed and in case of COVID-19 patients with simultaneous neoplasia, it has been strongly recommended to treat Sars-CoV-2 infection firstly. PRESENTATION OF CASE: We reported a case of a 53-years-old women with early breast cancer and simultaneous asymptomatic SARS-CoV-2 infection. According to COVID-19 breast cancer recommendations she underwent hormone neoadjuvant treatment as a bridging therapy for surgery. Six months from the diagnosis, after virus eradication, patient underwent breast surgery. No SARS-CoV-2 RNA was found both in the surgical specimen and sentinel lymph node but micrometastasis were reported. During the last follow-up, the patient was in good clinical condition and started the adjuvant chemotherapy. DISCUSSION: COVID-19 outbreak determined the publication of temporary recommendation leading to an extensive use of neoadjuvant chemotherapy in breast cancer patients. Although endocrine therapy is a mainstay in the adjuvant treatment, its role in the neoadjuvant schedule is unclear. CONCLUSION: Upfront awake surgery should be preferred especially in asymptomatic COVID-19 patient with early breast cancer when monitoring of tumor response is not feasible.