Persistence of Immune Response Elicited by Three Doses of mRNA Vaccine against SARS-CoV-2 in a Cohort of Patients with Solid Tumors: A One-Year Follow-Up.

The role and durability of the immunogenicity of the BNT162b2 mRNA vaccine against severe acute respiratory virus 2 (SARS-CoV-2), in cancer patients one year after receiving the third dose have to be elucidated. We have prospectively evaluated the long-term immunogenicity of the third dose of the SARS-CoV-2 BNT162b2 mRNA vaccine in 55 patients undergoing active treatment. Neutralizing antibody (NT Ab) titers against Omicron variants and total anti-trimeric S IgG levels were measured one year after the third dose. Heparinized whole-blood samples were used for the assessment of the SARS-CoV-2 interferon-γ release assay (IGRA). Thirty-seven patients (67.3%) showed positive total anti-trimeric S IgG one year after the third dose. Looking at the T-cell response against the spike protein, the frequency of responder patients did not decrease significantly between six and twelve months after the third dose. Finally, less than 20% of cancer patients showed an undetectable NT Ab titer against BA.1 and BA.5 variants of concern (VOCs). Underlying therapies seem to not affect the magnitude or frequency of the immune response. Our work underlines the persistence of humoral and cellular immune responses against BNT162b2 in a cohort of cancer patients one year after receiving the third dose, regardless of the type of underlying therapy.

Characterization of CCP: Can We Use Past Convalescent Plasma from COVID-19 Patients for Treatment of New Emerging Variants?

Background and Objectives: New SARS-CoV-2 variants may impact the effectiveness of previously stored convalescent plasma (CCP). We defined levels of anti-delta and anti-omicron SARS-CoV-2 neutralizing antibodies (Nt-Abs) and investigated possible differences of past CCP Nt-Abs responses related to donor location in North and South Italy. Methods: Serum from 153 donors recovered from SARS-COV-2 infection (98 from northern and 55 from southern Italy) were analyzed for Nt-Abs characterization using our in house microneutralization assay. Results were compared to anti-Spike IgG measured by chemiluminescent assay (CLIA) to define a possible agreement with a more affordable test. Results: delta Nt-Abs titer in comparison to the reference strain (PV10734 D614G) showed a reduction of 82% in northern and 77% in southern Italy groups. Omicron Nt-Abs titer showed a reduction of 97%. CCP corresponding to Nt-Abs titer > 1:80 showed a median of 1365 BAU/mL for delta strain and 653 BAU/mL for reference strain. We found no statistical differences between Nt-Abs responses in North and South CCP donors. Conclusions: Not all past CCP could be used to treat patients with SARS-CoV-2 delta and omicron infections due to the lack of specific Nt-Abs. For the moment, the neutralization test remains the gold standard to select potential CCP donors. Interestingly, our study did not find NT-Abs differences between plasma collected from donors living in different areas of Italy.

Immune Response to BNT162b2 in Solid Organ Transplant Recipients: Negative Impact of Mycophenolate and High Responsiveness of SARS-CoV-2 Recovered Subjects against Delta Variant.

The immunogenicity of severe acute respiratory syndrome 2 virus (SARS-CoV-2) vaccines in immunocompromised patients remains to be further explored. Here, we evaluated the immunogenicity elicited by complete vaccination with BNT162b2 vaccine in solid organ transplant recipients (SOTRs). A cohort of 110 SOTRs from Northern Italy were vaccinated with two doses of BNT162b2 mRNA vaccine and prospectively monitored at baseline and after 42 days. Both SARS-CoV-2 naïve and recovered subjects were included. Humoral response elicited by vaccination, including SARS-CoV-2 neutralizing antibodies (SARS-CoV-2 NT Abs), was evaluated; additionally, ex-vivo ELISpot assay was performed for the quantification of Spike-specific T-cell response. Results were compared with those obtained in a cohort of healthy subjects. In a subset of patients, humoral and T-cell responses against delta variant were also evaluated. Less than 20% of transplanted subjects developed a positive humoral and cell-mediated response after complete vaccination schedule. Overall, median levels of immune response elicited by vaccination were significantly lower with respect to controls in SARS-CoV-2 naïve transplant, but not in SARS-CoV-2 recovered transplanted patients. Additionally, a significant impairment of both humoral and cell-mediated response was observed in mycophenolate-treated patients. Positive delta-SARS-CoV-2 NT Abs levels were detected in almost all the SARS-CoV-2 recovered subjects but not in previously uninfected patients. Our study supports previous observations of a low level of seroconversion after vaccination in transplanted patients.

Humoral and cell-mediated response against SARS-CoV-2 variants elicited by mRNA vaccine BNT162b2 in healthcare workers: a longitudinal observational study.

OBJECTIVES: To assess the humoral and cell-mediated response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) elicited by the mRNA BNT162b2 vaccine in SARS-CoV-2-experienced and -naive subjects against a reference strain and SARS-CoV-2 variants. METHODS: The humoral response (including neutralizing antibodies) and T-cell-mediated response elicited by BNT162b2 vaccine in 145 healthcare workers (both naive and positive for previous SARS-CoV-2 infection) were evaluated. In a subset of subjects, the effect of SARS-CoV-2 variants on antibody level and cell-mediated response was also investigated. RESULTS: Overall, 125/127 naive subjects (98.4%) developed both neutralizing antibodies and specific T cells after the second dose of vaccine. Moreover, the antibody and T-cell responses were effective against viral variants since SARS-CoV-2 NT Abs were still detectable in 55/68 (80.9%) and 25/29 (86.2%) naive subjects when sera were challenged against ß and δ variants, respectively. T-cell response was less affected, with no significant difference in the frequency of responders (p 0.369). Of note, two doses of vaccine were able to elicit sustained neutralizing antibody activity against all the SARS-CoV-2 variants tested in SARS-CoV-2-experienced subjects. CONCLUSIONS: BNT162b2 vaccine elicited a sustained humoral and cell-mediated response in immunocompetent subjects after two-dose administration of the vaccine, and the response seemed to be less affected by SARS-CoV-2 variants, the only exceptions being the ß and δ variants. Increased immunogenicity, also against SARS-CoV-2 variant strains, was observed in SARS-CoV-2-experienced subjects. These results suggest that triple exposure to SARS-CoV-2 antigens might be proposed as valuable strategy for vaccination campaigns.

SARS-CoV-2 vaccine breakthrough infections with the alpha variant are asymptomatic or mildly symptomatic among health care workers.

Vaccine breakthrough SARS-CoV-2 infection has been monitored in 3720 healthcare workers receiving 2 doses of BNT162b2. SARS-CoV-2 infection is detected in 33 subjects, with a 100-day cumulative incidence of 0.93%. Vaccine protection against acquisition of SARS-CoV-2 infection is 83% (95%CI: 58-93%) in the overall population and 93% (95%CI: 69-99%) in SARS-CoV-2-experienced subjects, when compared with a non-vaccinated control group from the same Institution, in which SARS-CoV-2 infection occurs in 20/346 subjects (100-day cumulative incidence: 5.78%). The infection is symptomatic in 16 (48%) vaccinated subjects vs 17 (85%) controls (p = 0.01). All analyzed patients, in whom the amount of viral RNA was sufficient for genome sequencing, results infected by the alpha variant. Antibody and T-cell responses are not reduced in subjects with breakthrough infection. Evidence of virus transmission, determined by contact tracing, is observed in two (6.1%) cases. This real-world data support the protective effect of BNT162b2 vaccine. A triple antigenic exposure, such as two-dose vaccine schedule in experienced subjects, may confer a higher protection.

Incidence of SARS-CoV-2 infection in health care workers from Northern Italy based on antibody status: immune protection from secondary infection- A retrospective observational case-controlled study.

OBJECTIVE: The protection from SARS-CoV-2 infection induced by SARS-CoV-2 anti-S1 and anti-S2 IgG antibody positivity resulting from natural infection was evaluated. METHODS: The frequency of SARS-CoV-2 infection (as determined by virus RNA detection) was evaluated in a group of 1,460 seropositive and a control group of 8,150 seronegative healthcare workers in three Centres of Northern Italy in the period June-November 2020. Neutralizing serum titers were analyzed in seropositive subjects with or without secondary SARS-CoV-2 infection. RESULTS: During the 6-month survey, 1.78% seropositive subjects developed secondary SARS-CoV-2 infection while 6.63% seronegative controls developed primary infection (odds ratio: 0.26; 95% confidence interval: 0.17-0.38). Secondary infection was associated with low or absent serum neutralizing titer (p<0.01) and was mildly symptomatic in 45.8% cases vs 71.4% symptomatic primary infections (odds ratio: 0.34; 95% confidence interval: 0.16-0.78). CONCLUSIONS: Immunity from natural infection appears protective from secondary infection; therefore, vaccination of seronegative subjects might be prioritized.

Seroprevalence of SARS-CoV-2 in blood donors from the Lodi Red Zone and adjacent Lodi metropolitan and suburban area.

OBJECTIVES: To define the seroprevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in blood donors (referred to the first lockdown area (Lodi Red-Zone) of the Lombardy region and in a contiguous area that was not included in the first lockdown); to define the agreement between a commercial serological assay and a reference microneutralization assay; and to evaluate the persistence of SARS-CoV-2 neutralizing antibodies in a cohort of blood donors. METHODS: Blood donors referred to the first lockdown area in Lombardy Region and the neighbouring area were analysed for SARS-CoV-2 IgG-specific antibodies during the period 18 March to 24 June 2020. Serum samples were analysed using both a chemiluminescent immunoassay (LIAISON® SARS-CoV-2 S1/S2 IgG, DiaSorin) for the quantitative characterization of SARS-CoV-2 anti-S1 and anti-S2 IgG antibodies and a neutralizing antibodies (NT-Abs) assay. RESULTS: In the period from 18 March to 24 June, 1922 blood donors were tested for the presence of SARS-CoV-2 IgG showing a prevalence of 378/1922 (19.7%). A subgroup of 1139 blood donors were tested in parallel with a SARS-CoV-2 IgG assay and a microneutralization assay showing a prevalence of 22.2% and 21.6%, respectively. SARS-CoV-2 IgG quantification was correlated with NT-Abs titres. In 78.2% of participants the NT-Abs titre was maintained, but in 15.8% it decreased by one four-fold dilution and in 6.0% it increased by one four-fold dilution. CONCLUSIONS: The duration of immunity of SARS-CoV-2 is crucial for the course of the pandemic and for this reason the monitoring of NT Abs is important. Despite a stable NT-Abs titre being observed in the majority of blood donors, our findings need to be validated in a long-term period of follow up.

Persistence of SARS-CoV-2-specific B and T cell responses in convalescent COVID-19 patients 6-8 months after the infection.

BACKGROUND: Monitoring the adaptive immune responses during the natural course of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection provides useful information for the development of vaccination strategies against this virus and its emerging variants. We thus profiled the serum anti-SARS-CoV-2 antibody (Ab) levels and specific memory B and T cell responses in convalescent coronavirus disease 2019 (COVID-19) patients. METHODS: A total of 119 samples from 88 convalescent donors who experienced mild to critical disease were tested for the presence of elevated anti-spike and anti-receptor binding domain Ab levels over a period of 8 months. In addition, the levels of SARS-CoV-2 neutralizing Abs and specific memory B and T cell responses were tested in a subset of samples. FINDINGS: Anti-SARS-CoV-2 Abs were present in 85% of the samples collected within 4 weeks after the onset of symptoms in COVID-19 patients. Levels of specific immunoglobulin M (IgM)/IgA Abs declined after 1 month, while levels of specific IgG Abs and plasma neutralizing activities remained relatively stable up to 6 months after diagnosis. Anti-SARS-CoV-2 IgG Abs were still present, although at a significantly lower level, in 80% of the samples collected at 6-8 months after symptom onset. SARS-CoV-2-specific memory B and T cell responses developed with time and were persistent in all of the patients followed up for 6-8 months. CONCLUSIONS: Our data suggest that protective adaptive immunity following natural infection of SARS-CoV-2 may persist for at least 6-8 months, regardless of disease severity. Development of medium- or long-term protective immunity through vaccination may thus be possible. FUNDING: This project was supported by the European Union's Horizon 2020 research and innovation programme (ATAC, no. 101003650), the Italian Ministry of Health (Ricerca Finalizzata grant no. GR-2013-02358399), the Center for Innovative Medicine, and the Swedish Research Council. J.A. was supported by the SciLifeLab/KAW national COVID-19 research program project grant 2020.

SARS-CoV-2 Virologic and Immunologic Correlates in Patients with Olfactory and Taste Disorders

The main object of the study was to investigate the SARS-CoV-2 molecular and serological pattern in patients with mild symptoms including anosmia and ageusia. A cohort of 69 patients with olfactory and taste disorders (OTDs) were enrolled and prospectively monitored. Serological and molecular assays for the characterization of SARS-CoV-2 IgG and SARS-CoV-2 RNA, respectively, were performed at the time of enrolment and after 7 and 14 days. Patients were stratified according to the symptoms"onset. A total of 52 patients (75.4%) were diagnosed as COVID-19 positive being SARS-CoV-2 RNA and/or SARS-CoV-2 IgG positive. The remaining 17 (24.6%) were negative for COVID-19 and excluded from the analysis. We reported that only 34 out of 52 patients (65.4%) were positive for SARS-CoV-2 RNA. Moreover, the median time from onset of symptoms and enrolment was significantly higher in those patients with negative SARS-CoV-2 RNA in nasal swabs, suggesting that symptoms might last longer than SARS-CoV-2 replication. The great majority of patients (80%) developed SARS-CoV-2 IgG at three weeks after symptoms"onset while the detectability of SARS-CoV-2 RNA dramatically decreased over time, suggesting the crucial role of combination of molecular and serological assays for the diagnosis of COVID-19 in those patients reporting mild symptoms.

Clinical and Analytical Performance of an Automated Serological Test That Identifies S1/S2-Neutralizing IgG in COVID-19 Patients Semiquantitatively.

In the coronavirus (CoV) disease 2019 (COVID-19) pandemic, highly selective serological testing is essential to define exposure to severe acute respiratory syndrome CoV 2 (SARS-CoV-2). Many tests have been developed, yet with variable speeds to first results, and are of unknown quality, particularly when considering the prediction of neutralizing capacity. The LIAISON SARS-CoV-2 S1/S2 IgG assay was designed to measure antibodies against the SARS-CoV-2 native S1/S2 proteins in a standardized automated chemiluminescence assay. The clinical and analytical performances of the test were validated in an observational study using residual samples (>1,500) with a positive or negative COVID-19 diagnosis. The LIAISON SARS-CoV-2 S1/S2 IgG assay proved to be highly selective and specific and offered semiquantitative measures of serum or plasma levels of anti-S1/S2 IgG with neutralizing activity. The assay's diagnostic sensitivities were 91.3% and 95.7% at >5 or ≥15 days from diagnosis, respectively, and 100% when assessed against a neutralizing assay. The assay's specificity ranged between 97% and 98.5%. The average imprecision of the assay was a <5% coefficient of variation. Assay performance at 2 different cutoffs was evaluated to optimize predictive values. The automated LIAISON SARS-CoV-2 S1/S2 IgG assay brings efficient, sensitive, specific, and precise serological testing to the laboratory, with the capacity to test large amounts of samples per day; first results are available within 35 min, with a throughput of 170 tests/hour. The semiquantitative results provided by the test also associate with the presence of neutralizing antibodies and may provide a useful tool for the large-scale screening of convalescent-phase plasma for safe therapeutic use.

Mechanism of baricitinib supports artificial intelligence-predicted testing in COVID-19 patients.

Baricitinib is an oral Janus kinase (JAK)1/JAK2 inhibitor approved for the treatment of rheumatoid arthritis (RA) that was independently predicted, using artificial intelligence (AI) algorithms, to be useful for COVID-19 infection via proposed anti-cytokine effects and as an inhibitor of host cell viral propagation. We evaluated the in vitro pharmacology of baricitinib across relevant leukocyte subpopulations coupled to its in vivo pharmacokinetics and showed it inhibited signaling of cytokines implicated in COVID-19 infection. We validated the AI-predicted biochemical inhibitory effects of baricitinib on human numb-associated kinase (hNAK) members measuring nanomolar affinities for AAK1, BIKE, and GAK. Inhibition of NAKs led to reduced viral infectivity with baricitinib using human primary liver spheroids. These effects occurred at exposure levels seen clinically. In a case series of patients with bilateral COVID-19 pneumonia, baricitinib treatment was associated with clinical and radiologic recovery, a rapid decline in SARS-CoV-2 viral load, inflammatory markers, and IL-6 levels. Collectively, these data support further evaluation of the anti-cytokine and anti-viral activity of baricitinib and support its assessment in randomized trials in hospitalized COVID-19 patients.