Interleukin-62/lymphocyte as a proposed predictive index for COVID-19 patients treated with monoclonal antibodies.

In a convenience sample of 93 patients treated with monoclonal antibodies (moAbs) against SARS-CoV-2, the interleukin-62/lymphocyte count ratio (IL-62/LC) was able to predict clinical worsening both in early stages of COVID-19 and in oxygen-requiring patients. Moreover, we analysed 18 most at-risk patients with asymptomatic or mild disease treated with both moAbs and antiviral treatment and found that only 2 had clinical progression, while patients with a similar risk were reported to have an unfavourable outcome in most cases from recent data. In only one of our 18 patients, clinical progression was attributable to COVID-19, and in the other cases, clinical progression was observed despite IL-62/LC being above the risk cut-off. In conclusion, IL-62/LC may be a valuable method to identify patients requiring more aggressive treatments both in earlier and later stages of the disease; however, most at-risk patients can be protected from clinical worsening by combining moAbs and antivirals, even if levels of the IL-62/LC biomarker are lower than the risk cut-off.

Severe and mild-moderate SARS-CoV-2 vaccinated patients show different frequencies of IFNγ-releasing cells: An exploratory study.

BACKGROUND: Despite an apparent effective vaccination, some patients are admitted to the hospital after SARS-CoV-2 infection. The role of adaptive immunity in COVID-19 is growing; nonetheless, differences in the spike-specific immune responses between patients requiring or not hospitalization for SARS-CoV-2 infection remains to be evaluated. In this study, we aim to evaluate the spike-specific immune response in patients with mild-moderate or severeSARS-CoV-2 infection, after breakthrough infection following two doses of BNT162b2 mRNA vaccine. METHODS: We included three cohorts of 15 cases which received the two BNT162b2 vaccine doses in previous 4 to 7 months: 1) patients with severe COVID-19; 2) patients with mild-moderate COVID-19 and 3) vaccinated individuals with a negative SARS-CoV-2 molecular pharyngeal swab (healthy subjects). Anti-S1 and anti-S2 specific SARS-CoV-2 IgM and IgG titers were measured through a chemiluminescence immunoassay technology. In addition, the frequencies of IFNγ-releasing cells were measured by ELISpot. RESULTS: The spike-specific IFNγ-releasing cells were significantly lower in severe patients (8 [0; 26] s.f.c.×106), as compared to mild-moderate patients (135 [64; 159] s.f.c.×106; p<0.001) and healthy subjects (103 [50; 188] s.f.c.×106; p<0.001). The anti-Spike protein IgG levels were similar among the three cohorts of cases (p = 0.098). All cases had an IgM titer below the analytic sensitivity of the test. The Receiver Operating Curve analysis indicated the rate of spike-specific IFNγ-releasing cells can discriminate correctly severe COVID-19 and mild-moderate patients (AUC: 0.9289; 95%CI: 0.8376-1.000; p< 0.0001), with a diagnostic specificity of 100% for s.f.c. > 81.2 x 106. CONCLUSIONS: 2-doses vaccinated patients requiring hospitalization for severe COVID-19 show a cellular-mediated immune response lower than mild-moderate or healthy subjects, despite similar antibody titers.

The Spread of SARS-CoV-2 Omicron Variant in CALABRIA: A Spatio-Temporal Report of Viral Genome Evolution.

We investigated the evolution of SARS-CoV-2 spread in Calabria, Southern Italy, in 2022. A total of 272 RNA isolates from nasopharyngeal swabs of individuals infected with SARS-CoV-2 were sequenced by whole genome sequencing (N = 172) and/or Sanger sequencing (N = 100). Analysis of diffusion of Omicron variants in Calabria revealed the prevalence of 10 different sub-lineages (recombinant BA.1/BA.2, BA.1, BA.1.1, BA.2, BA.2.9, BA.2.10, BA.2.12.1, BA.4, BA.5, BE.1). We observed that Omicron spread in Calabria presented a similar trend as in Italy, with some notable exceptions: BA.1 disappeared in April in Calabria but not in the rest of Italy; recombinant BA.1/BA.2 showed higher frequency in Calabria (13%) than in the rest of Italy (0.02%); BA.2.9, BA.4 and BA.5 emerged in Calabria later than in other Italian regions. In addition, Calabria Omicron presented 16 non-canonical mutations in the S protein and 151 non-canonical mutations in non-structural proteins. Most non-canonical mutations in the S protein occurred mainly in BA.5 whereas non-canonical mutations in non-structural or accessory proteins (ORF1ab, ORF3a, ORF8 and N) were identified in BA.2 and BA.5 sub-lineages. In conclusion, the data reported here underscore the importance of monitoring the entire SARS-CoV-2 genome.

Lessons learned and implications of early therapies for coronavirus disease in a territorial service centre in the Calabria region: a retrospective study.

BACKGROUND: Monoclonal antibodies (mAbs) and antivirals have been approved for early therapy of coronavirus disease (COVID-19), however, in the real-life setting, there are difficulties to prescribe these therapies within few days from symptom onset as recommended, and effectiveness of combined use of these drugs have been hypothesised in most-at-risk patients (such as those immunocompromised) but data supporting this strategy are limited. METHODS: We describe the real-life experience of SARS-CoV-2 antivirals and/or monoclonal antibodies (mAbs) and focus on the hospitalisation rate due to the progression of COVID-19. Clinical results obtained through our risk-stratification algorithm and benefits achieved through a strategic proximity territorial centre are provided. We also report a case series with an in-depth evaluation of SARS-CoV-2 genome in relationship with treatment strategy and clinical evolution of patients. RESULTS: Two hundred eighty-eight patients were analysed; 94/288 (32.6%) patients were treated with mAb monotherapy, 171/288 (59.4%) patients were treated with antivirals, and 23/288 (8%) patients received both mAbs and one antiviral drug. Haematological malignancies were more frequent in patients treated with combination therapy than in the other groups (p = 0.0003). There was a substantial increase in the number of treated patients since the opening of the centre dedicated to early therapies for COVID-19. The provided disease-management and treatment appeared to be effective since 98.6% patients recovered without hospital admission. Moreover, combination therapy with mAbs and antivirals seemed successful because all patients admitted to the hospital for COVID-19 did not receive such therapies, while none of the most-at-risk patients treated with combination therapy were hospitalized or reported adverse events. CONCLUSIONS: A low rate of COVID-19 progression requiring hospital admission was observed in patients included in this study. The dedicated COVID-19 proximity territorial service appeared to strengthen the regional sanitary system, avoiding the overwhelming of other services. Importantly, our results also support early combination therapy: it is possible that this strategy reduces the emergence of escape mutants of SARS-CoV-2, thereby increasing efficacy of early treatment, especially in immunocompromised individuals.

Spike-specific T-cell responses in patients with COVID-19 successfully treated with neutralizing monoclonal antibodies against SARS-CoV-2.

OBJECTIVES: Neutralizing monoclonal antibodies (moAbs) improves clinical outcomes in patients with COVID-19 when administered during the initial days of infection. The action of moAbs may impair the generation or maintenance of effective immune memory, similar to that demonstrated in other viral diseases. We aimed to evaluate short-term memory T-cell responses in patients effectively treated with bamlanivimab/etesevimab, casirivimab/imdevimab, or sotrovimab (SOT). METHODS: Spike (S)-specific T-cell responses were analyzed in 23 patients with COVID-19 (vaccinated or unvaccinated) before and after a median of 50 (range: 28-93) days from moAb treatment, compared with 11 vaccinated healthy controls. T-cell responses were measured by interferon-γ-enzyme-linked immunospot and flow cytometric activation-induced marker assay. RESULTS: No statistically significant difference in S-specific T-cell responses was observed between patients treated with moAb and vaccinated healthy controls. Bamlanivimab/etesevimab and casirivimab/imdevimab groups showed significant increases in cellular responses in paired baseline/postrecovery series, as well as vaccinated patients receiving SOT. In contrast, unvaccinated patients prescribed SOT presented no statistically significant increases in T-cell-responses, suggesting diverse impacts of different moAbs on the evolution of S-specific T-cell responses in vaccinated and unvaccinated patients. CONCLUSION: The moAbs did not hinder short-term memory S-specific T-cell responses in the overall group of patients; however, differences among moAbs must be further investigated both in vaccinated and unvaccinated individuals.

Dynamics of Viral Infection and Evolution of SARS-CoV-2 Variants in the Calabria Area of Southern Italy

In this study, we report on the results of SARS-CoV-2 surveillance performed in an area of Southern Italy for 12 months (from March 2021 to February 2022). To this study, we have sequenced RNA from 609 isolates. We have identified circulating VOCs by Sanger sequencing of the S gene and defined their genotypes by whole-genome NGS sequencing of 157 representative isolates. Our results indicated that B.1 and Alpha were the only circulating lineages in Calabria in March 2021;while Alpha remained the most common variant between April 2021 and May 2021 (90 and 73%, respectively), we observed a concomitant decrease in B.1 cases and appearance of Gamma cases (6 and 21%, respectively);C.36.3 and Delta appeared in June 2021 (6 and 3%, respectively);Delta became dominant in July 2021 while Alpha continued to reduce (46 and 48%, respectively). In August 2021, Delta became the only circulating variant until the end of December 2021. As of January 2022, Omicron emerged and took over Delta (72 and 28%, respectively). No patient carrying Beta, Iota, Mu, or Eta variants was identified in this survey. Among the genomes identified in this study, some were distributed all over Europe (B1_S477N, Alpha_L5F, Delta_T95, Delta_G181V, and Delta_A222V), some were distributed in the majority of Italian regions (B1_S477N, B1_Q675H, Delta_T95I and Delta_A222V), and some were present mainly in Calabria (B1_S477N_T29I, B1_S477N_T29I_E484Q, Alpha_A67S, Alpha_A701S, and Alpha_T724I). Prediction analysis of the effects of mutations on the immune response (i.e., binding to class I MHC and/or recognition of T cells) indicated that T29I in B.1 variant;A701S in Alpha variant;and T19R in Delta variant were predicted to impair binding to class I MHC whereas the mutations A67S identified in Alpha;E484K identified in Gamma;and E156G and ΔF157/R158 identified in Delta were predicted to impair recognition by T cells. In conclusion, we report on the results of SARS-CoV-2 surveillance in Regione Calabria in the period between March 2021 and February 2022, identified variants that were enriched mainly in Calabria, and predicted the effects of identified mutations on host immune response.

Whole-genome analysis of SARS-CoV-2 in a 2020 infection cluster in a nursing home of Southern Italy.

BACKGROUND: Nursing homes have represented important hotspots of viral spread during the initial wave of COVID-19 pandemics. The proximity of patients inside nursing homes allows investigate the dynamics of viral transmission, which may help understand SARS-Cov2 biology and spread. METHODS: SARS-CoV-2 viral genomes obtained from 46 patients infected in an outbreak inside a nursing home in Calabria region (South Italy) were analyzed by Next Generation Sequencing. We also investigated the evolution of viral genomes in 8 patients for which multiple swabs were available. Phylogenetic analysis and haplotype reconstruction were carried out with IQ-TREE software and RegressHaplo tool, respectively. RESULTS: All viral strains isolated from patients infected in the nursing home were classified as B.1 lineage, clade G. Overall, 14 major single nucleotide variations (SNVs) (frequency > 80%) and 12 minor SNVs (frequency comprised between 20% and 80%) were identified with reference to the Wuhan-H-1 sequence (NC_045512.2). All patients presented the same 6 major SNVs: D614G in the S gene; P4715L, ntC3037T (F924F) and S5398P in Orf1ab gene; ntC26681T (F53F) in the M gene; and ntC241T in the non-coding UTR region. However, haplotype reconstruction identified a founder haplotype (Hap A) in 36 patients carrying only the 6 common SNVs indicated above, and 10 other haplotypes (Hap BK) derived from Hap A in the remaining 10 patients. Notably, no significant association between a specific viral haplotype and clinical parameters was found. CONCLUSION: The predominant viral strain responsible for the infection in a nursing home in Calabria was the B.1 lineage (clade G). Viral genomes were classified into 11 haplotypes (Hap A in 36 patients, Hap BK in the remaining patients).

Clinical characteristics and predictors of mortality associated with COVID-19 in elderly patients from a long-term care facility.

Since December 2019, coronavirus disease 2019 (COVID-19) pandemic has spread from China all over the world and many COVID-19 outbreaks have been reported in long-term care facilities (LCTF). However, data on clinical characteristics and prognostic factors in such settings are scarce. We conducted a retrospective, observational cohort study to assess clinical characteristics and baseline predictors of mortality of COVID-19 patients hospitalized after an outbreak of SARS-CoV-2 infection in a LTCF. A total of 50 patients were included. Mean age was 80 years (SD, 12 years), and 24/50 (57.1%) patients were males. The overall in-hospital mortality rate was 32%. At Cox regression analysis, significant predictors of in-hospital mortality were: hypernatremia (HR 9.12), lymphocyte count < 1000 cells/µL (HR 7.45), cardiovascular diseases other than hypertension (HR 6.41), and higher levels of serum interleukin-6 (IL-6, pg/mL) (HR 1.005). Our study shows a high in-hospital mortality rate in a cohort of elderly patients with COVID-19 and hypernatremia, lymphopenia, CVD other than hypertension, and higher IL-6 serum levels were identified as independent predictors of in-hospital mortality. Given the small population size as major limitation of our study, further investigations are necessary to better understand and confirm our findings in elderly patients.

COVID-19: High-JAKing of the Inflammatory "Flight" by Ruxolitinib to Avoid the Cytokine Storm.

Since SARS-CoV-2 outbreak in December 2019, world health-system has been severely impacted with increased hospitalization, Intensive-Care-Unit (ICU) access and high mortality rates, mostly due to severe acute respiratory failure and multi-organ failure. Excessive and uncontrolled release of proinflammatory cytokines (cytokine release/storm syndrome, CRS) have been linked to the development of these events. The recent advancements of immunotherapy for the treatment of hematologic and solid tumors shed light on many of the molecular mechanisms underlying this phenomenon, thus rendering desirable a multidisciplinary approach to improve COVID-19 patients' outcome. Indeed, currently available therapeutic-strategies to overcome CRS, should be urgently evaluated for their capability of reducing COVID-19 mortality. Notably, COVID-19 shares different pathogenic aspects with acute graft-versus-host-disease (aGVHD), hemophagocytic-lymphohistiocytosis (HLH), myelofibrosis, and CAR-T-associated CRS. Specifically, similarly to aGVHD, an induced tissue damage (caused by the virus) leads to increased cytokine release (TNFα and IL-6) which in turn leads to exaggerated dendritic cells, macrophages (like in HLH) and lymphocytes (as in CAR-T) activation, immune-cells migration, and tissue-damage (including late-stage fibrosis, similar to myelofibrosis). Janus Kinase (JAK) signaling represents a molecular hub linking all these events, rendering JAK-inhibitors suitable to limit deleterious effects of an overwhelming inflammatory-response. Accordingly, ruxolitinib is the only selective JAK1 and JAK2-inhibitor approved for the treatment of myelofibrosis and aGVHD. Here, we discuss, from a molecular and hematological point of view, the rationale for targeting JAK signaling in the management of COVID-19 patients and report the clinical results of a patient admitted to ICU among the firsts to be treated with ruxolitinib in Italy.

Use of subcutaneous tocilizumab in patients with COVID-19 pneumonia

Over the last months, pandemic SARS-CoV-2 caused a significant challenge for clinicians. Unfortunately, no approved and validated treatments are available. Intravenous tocilizumab, an antirheumatic drug, seems to be promising in counteracting cytokine storm caused by SARS-CoV-2 infection with associated clinical improvements. We report herein a case series of patients with COVID-19 pneumonia who were treated with tocilizumab administrated, for the first time, subcutaneously with good clinical and radiological outcomes. This article is protected by copyright. All rights reserved.