Cellular and humoral responses after second and third SARS-CoV-2 vaccinations in patients with autoimmune diseases treated with rituximab: specific T cell immunity remains longer and plays a protective role against SARS-CoV-2 reinfections.

Background: Humoral and cellular immune responses are known to be crucial for patients to recover from COVID-19 and to protect them against SARS-CoV-2 reinfection once infected or vaccinated. Objectives: This study aimed to investigate humoral and T cell responses to SARS-CoV-2 vaccination in patients with autoimmune diseases after the second and third vaccine doses while on rituximab and their potential protective role against reinfection. Methods: Ten COVID-19-naïve patients were included. Three time points were used for monitoring cellular and humoral responses: pre-vaccine to exclude virus exposure (time point 1) and post-second and post-third vaccine (time points 2 and 3). Specific IgG antibodies were monitored by Luminex and T cells against SARS-CoV-2 spike-protein by ELISpot and CoVITEST. All episodes of symptomatic COVID-19 were recorded. Results: Nine patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis and one with an undifferentiated autoimmune disease were included. Nine patients received mRNA vaccines. The last rituximab infusion was administered for a mean (SD) of 15 (10) weeks before the first vaccine and six patients were CD19-B cell-depleted. After a mean (SD) of 19 (10) and 16 (2) days from the second and third vaccine dose, IgG anti-SARS-CoV-2 antibodies were detected in six (60%) and eight (80%) patients, respectively. All patients developed specific T cell responses by ELISpot and CoVITEST in time points 2 and 3. Previous B cell depletion correlated with anti-SARS-CoV-2 IgG levels. Nine (90%) patients developed mild COVID-19 after a median of 7 months of the third dose. Conclusion: Rituximab in patients with autoimmune diseases reduces humoral responses but does not avoid the development of T cell responses to SARS-CoV-2 vaccination, which remain present after a booster dose. A steady cellular immunity appears to be protective against subsequent reinfections.

Acute and long-term immune responses to SARS-CoV-2 infection in unvaccinated children and young adults with inborn errors of immunity.

Purpose: To describe SARS-CoV-2 infection outcome in unvaccinated children and young adults with inborn errors of immunity (IEI) and to compare their specific acute and long-term immune responses with a sex-, age-, and severity-matched healthy population (HC). Methods: Unvaccinated IEI patients up to 22 years old infected with SARS-CoV-2 were recruited along with a cohort of HC. SARS-CoV-2 serology and ELISpot were performed in the acute phase of infection (up to 6 weeks) and at 3, 6, 9, and 12 months. Results: Twenty-five IEI patients (median age 14.3 years, min.-max. range 4.5-22.8; 15/25 males; syndromic combined immunodeficiencies: 48.0%, antibody deficiencies: 16.0%) and 17 HC (median age 15.3 years, min.-max. range 5.4-20.0; 6/17 males, 35.3%) were included. Pneumonia occurred in 4/25 IEI patients. In the acute phase SARS-CoV-2 specific immunoglobulins were positive in all HC but in only half of IEI in whom it could be measured (n=17/25): IgG+ 58.8% (10/17) (p=0.009); IgM+ 41.2% (7/17)(p<0.001); IgA+ 52.9% (9/17)(p=0.003). Quantitative response (index) was also lower compared with HC: IgG IEI (3.1 ± 4.4) vs. HC (3.5 ± 1.5)(p=0.06); IgM IEI (1.9 ± 2.4) vs. HC (3.9 ± 2.4)(p=0.007); IgA IEI (3.3 ± 4.7) vs. HC (4.6 ± 2.5)(p=0.04). ELISpots positivity was qualitatively lower in IEI vs. HC (S-ELISpot IEI: 3/11, 27.3% vs. HC: 10/11, 90.9%; p=0.008; N-ELISpot IEI: 3/9, 33.3% vs. HC: 11/11, 100%; p=0.002) and also quantitatively lower (S-ELISpot IEI: mean index 3.2 ± 5.0 vs. HC 21.2 ± 17.0; p=0.001; N-ELISpot IEI: mean index 9.3 ± 16.6 vs. HC: 39.1 ± 23.7; p=0.004). As for long term response, SARS-CoV-2-IgM+ at 6 months was qualitatively lower in IEI(3/8, 37.5% vs. 9/10 HC: 90.0%; p=0.043), and quantitatively lower in all serologies IgG, M, and A (IEI n=9, 1.1 ± 0.9 vs. HC n=10, 2.1 ± 0.9, p=0.03; IEI n=9, 1.3 ± 1.5 vs. HC n=10, 2.9 ± 2.8, p=0.02; and IEI n=9, 0.6 ± 0.5 vs. HC n=10, 1.7 ± 0.8, p=0.002 -respectively) but there were no differences at remaining time points. Conclusions: Our IEI pediatric cohort had a higher COVID-19 pneumonia rate than the general age-range population, with lower humoral and cellular responses in the acute phase (even lower compared to the reported IEI serological response after SARS-CoV-2 vaccination), and weaker humoral responses at 6 months after infection compared with HC.

Acute and long-term immune responses to SARS-CoV-2 infection in unvaccinated children and young adults with inborn errors of immunity

Purpose To describe SARS-CoV-2 infection outcome in unvaccinated children and young adults with inborn errors of immunity (IEI) and to compare their specific acute and long-term immune responses with a sex-, age-, and severity-matched healthy population (HC). Methods Unvaccinated IEI patients up to 22 years old infected with SARS-CoV-2 were recruited along with a cohort of HC. SARS-CoV-2 serology and ELISpot were performed in the acute phase of infection (up to 6 weeks) and at 3, 6, 9, and 12 months. Results Twenty-five IEI patients (median age 14.3 years, min.-max. range 4.5-22.8;15/25 males;syndromic combined immunodeficiencies: 48.0%, antibody deficiencies: 16.0%) and 17 HC (median age 15.3 years, min.-max. range 5.4-20.0;6/17 males, 35.3%) were included. Pneumonia occurred in 4/25 IEI patients. In the acute phase SARS-CoV-2 specific immunoglobulins were positive in all HC but in only half of IEI in whom it could be measured (n=17/25): IgG+ 58.8% (10/17) (p=0.009);IgM+ 41.2% (7/17)(p<0.001);IgA+ 52.9% (9/17)(p=0.003). Quantitative response (index) was also lower compared with HC: IgG IEI (3.1 ± 4.4) vs. HC (3.5 ± 1.5)(p=0.06);IgM IEI (1.9 ± 2.4) vs. HC (3.9 ± 2.4)(p=0.007);IgA IEI (3.3 ± 4.7) vs. HC (4.6 ± 2.5)(p=0.04). ELISpots positivity was qualitatively lower in IEI vs. HC (S-ELISpot IEI: 3/11, 27.3% vs. HC: 10/11, 90.9%;p=0.008;N-ELISpot IEI: 3/9, 33.3% vs. HC: 11/11, 100%;p=0.002) and also quantitatively lower (S-ELISpot IEI: mean index 3.2 ± 5.0 vs. HC 21.2 ± 17.0;p=0.001;N-ELISpot IEI: mean index 9.3 ± 16.6 vs. HC: 39.1 ± 23.7;p=0.004). As for long term response, SARS-CoV-2-IgM+ at 6 months was qualitatively lower in IEI(3/8, 37.5% vs. 9/10 HC: 90.0%;p=0.043), and quantitatively lower in all serologies IgG, M, and A (IEI n=9, 1.1 ± 0.9 vs. HC n=10, 2.1 ± 0.9, p=0.03;IEI n=9, 1.3 ± 1.5 vs. HC n=10, 2.9 ± 2.8, p=0.02;and IEI n=9, 0.6 ± 0.5 vs. HC n=10, 1.7 ± 0.8, p=0.002 –respectively) but there were no differences at remaining time points. Conclusions Our IEI pediatric cohort had a higher COVID-19 pneumonia rate than the general age-range population, with lower humoral and cellular responses in the acute phase (even lower compared to the reported IEI serological response after SARS-CoV-2 vaccination), and weaker humoral responses at 6 months after infection compared with HC. Graphical

CoVITEST: A Fast and Reliable Method to Monitor Anti-SARS-CoV-2 Specific T Cells From Whole Blood

Cellular and humoral immune responses are essential for COVID-19 recovery and protection against SARS-CoV-2 reinfection. To date, the evaluation of SARS-CoV-2 immune protection has mainly focused on antibody detection, generally disregarding the cellular response, or placing it in a secondary position. This phenomenon may be explained by the complex nature of the assays needed to analyze cellular immunity compared with the technically simple and automated detection of antibodies. Nevertheless, a large body of evidence supports the relevance of the T cell’s role in protection against SARS-CoV-2, especially in vulnerable individuals with a weakened immune system (such as the population over 65 and patients with immunodeficiencies). Here we propose to use CoVITEST (Covid19 anti-Viral Immunity based on T cells for Evaluation in a Simple Test), a fast, affordable and accessible in-house assay that, together with a diagnostic matrix, allows us to determine those patients who might be protected with SARS-CoV-2-reactive T cells. The method was established using healthy SARS-CoV-2-naïve donors pre- and post-vaccination (n=30), and further validated with convalescent COVID-19 donors (n=51) in a side-by-side comparison with the gold standard IFN-γ ELISpot. We demonstrated that our CoVITEST presented reliable and comparable results to those obtained with the ELISpot technique in a considerably shorter time (less than 8 hours). In conclusion, we present a simple but reliable assay to determine cellular immunity against SARS-CoV-2 that can be used routinely during this pandemic to monitor the immune status in vulnerable patients and thereby adjust their therapeutic approaches. This method might indeed help to optimize and improve decision-making protocols for re-vaccination against SARS-CoV-2, at least for some population subsets.