Induction and subsequent decline of S1-specific T cell reactivity after COVID-19 vaccination.

We analyzed magnitude and duration of SARS-CoV-2-specific T cell responses in healthy, infection-naïve subjects receiving COVID-19 vaccines. Overlapping peptides spanning the N-terminal spike 1 (S1) domain of the spike protein triggered secretion of the T cell-derived cytokine interleukin-2 ex vivo in 94/94 whole blood samples from vaccinated subjects at levels exceeding those recorded in all 45 pre-vaccination samples. S1-specific T cell reactivity was stronger in vaccinated subjects compared with subjects recovering from natural COVID-19 and decayed with an estimated half-life of 134 days in the first six months after the 2nd vaccination. We conclude that COVID-19 vaccination induces robust T cell immunity that subsequently declines. EudraCT 2021-000349-42. https://www.clinicaltrialsregister.eu/ctr-search/search?query=2021-000349-42.

An adjuvant-containing cDC1-targeted recombinant fusion vaccine conveys strong protection against murine melanoma growth and metastasis.

Type 1 conventional dendritic cells (cDC1) efficiently cross-present antigens that prime cytotoxic CD8+ T cells. cDC1 therefore constitute conceivable targets in cancer vaccine development. We generated recombinant fusion cancer vaccines that aimed to concomitantly deliver tumor antigen and adjuvant to CD103+ migratory cDC1, following intranasal administration. The fusion vaccine constructs comprised a cDC1-targeting anti-CD103 single chain antibody (aCD103) and a cholera toxin A1 (CTA1) subunit adjuvant, fused with MHC class I and II- or class II-restricted tumor cell antigens to generate a CTA1-I/II-aCD103 vaccine and a CTA1-II-aCD103 vaccine. The immunostimulatory and anti-tumor efficacy of these vaccines was evaluated in murine B16F1-ovalbumin (OVA) melanoma models in C57BL/6 J mice. The CTA1-I/II-aCD103 vaccine was most efficacious and triggered robust tumor antigen-specific CD8+ T cell responses along with a Th17-polarized CD4+ T cell response. This vaccine construct reduced the local growth of implanted B16F1-OVA melanomas and efficiently prevented hematogenous lung metastasis after prophylactic and therapeutic vaccination. Anti-tumor effects of the CTA1-I/II-aCD103 vaccine were antigen-specific and long-lasting. These results imply that adjuvant-containing recombinant fusion vaccines that target and activate cDC1 trigger effective anti-tumor immunity to control tumor growth and metastasis.

Transient and durable T cell reactivity after COVID-19.

This study analyzed whole blood samples (n = 56) retrieved from 30 patients at 1 to 21 (median 9) mo after verified COVID-19 to determine the polarity and duration of antigen-specific T cell reactivity against severe acute respiratory syndrome coronavirus 2-derived antigens. Multimeric peptides spanning the entire nucleocapsid protein triggered strikingly synchronous formation of interleukin (IL)-4, IL-12, IL-13, and IL-17 ex vivo until ∼70 d after confirmed infection, whereafter this reactivity was no longer inducible. In contrast, levels of nucleocapsid-induced IL-2 and interferon-γ remained stable and highly correlated at 3 to 21 mo after infection. Similar cytokine dynamics were observed in unvaccinated, convalescent patients using whole-blood samples stimulated with peptides spanning the N-terminal portion of the spike 1 protein. These results unravel two phases of T cell reactivity following natural COVID-19: an early, synchronous response indicating transient presence of multipolar, antigen-specific T helper (TH) cells followed by an equally synchronous and durable TH1-like reactivity reflecting long-lasting T cell memory.

Impaired SARS-CoV-2-specific T-cell reactivity in patients with cirrhosis following mRNA COVID-19 vaccination.

Background & Aims: Cirrhosis entails elevated risk of COVID-19-associated mortality. This study determined T cell-mediated and antibody reactivity against the spike 1 (S1) protein of SARS-CoV-2 among 48 patients with cirrhosis and 39 healthy controls after mRNA COVID-19 vaccination. Methods: SARS-CoV-2-specific T-cell reactivity was measured by induced level of T cell-derived interferon-γ (IFN-γ) in blood cells stimulated ex vivo with multimeric peptides spanning the N-terminal portion of S1. S1-induced IFN-γ was quantified before and after the 1st and 2nd vaccination (BNT162b2, Pfizer-BioNTech or mRNA-1273, Moderna) alongside serum IgG against the receptor-binding domain (RBD) within S1 (anti-RBD-S1 IgG). Results: T-cell reactivity against S1 was reduced in patients with cirrhosis after the 1st (p <0.001 vs. controls) and 2nd (p <0.001) vaccination. Sixty-eight percent of patients lacked detectable S1-specific T-cell reactivity after the 1st vaccination vs. 19% in controls (odds ratio 0.11, 95% CI 0.03-0.48, p = 0.003) and 36% remained devoid of reactivity after the 2nd vaccination vs. 6% in controls (odds ratio 0.12, 95% CI 0.03-0.59, p = 0.009). T-cell reactivity in cirrhosis remained significantly impaired after correction for potential confounders in multivariable analysis. Advanced cirrhosis (Child-Pugh class B) was associated with absent or lower T-cell responses (p <0.05 vs. Child-Pugh class A). The deficiency of T-cell reactivity was paralleled by lower levels of anti-RBD-S1 IgG after the 1st (p <0.001 vs. controls) and 2nd (p <0.05) vaccination. Conclusions: Patients with cirrhosis show deficient T-cell reactivity against SARS-CoV-2 antigens along with diminished levels of anti-RBD-S1 IgG after dual COVID-19 vaccination, highlighting the need for vigilance and additional preventative measures. Clinical trial registration: EudraCT 2021-000349-42. Lay summary: T cells are a pivotal component in the defence against viruses. We show that patients with cirrhosis have impaired SARS-CoV-2-specific T-cell responses and lower antibody levels after mRNA vaccination against COVID-19 compared with healthy controls. Patients with more advanced liver disease exhibited particularly inferior vaccine responses. These results call for additional preventative measures in these patients.

ADP-ribosylating adjuvant reveals plasticity in cDC1 cells that drive mucosal Th17 cell development and protection against influenza virus infection.

Migratory dendritic cells expressing CD103 are the targets for mucosal vaccines. These belong to either of two lineage-restricted subsets, cDC1 or cDC2 cells, which have been linked to priming of functionally distinct CD4 T cells. However, recent studies have identified plasticity in cDC2 cells with overlapping functions with cDC1 cells, while the converse has not been reported. We genetically engineered a vaccine adjuvant platform that targeted the cholera toxin A1 (CTA1) ADP-ribosylating enzyme to CD103+ cDC1 and cDC2 cells using a single-chain antibody (scFv) to CD103. Unexpectedly, intranasal immunization with the CTA1-svFcCD103 adjuvant modified cDC1 cells to effectively prime Th17 cells, a function previously limited to cDC2 cells. In fact, cDC2 cells were dispensible, while cDC1 cells, lacking in Batf3-/- mice, were critical. Following intranasal immunizations isolated cDC1 cells from mLN exclusively promoted Rorgt+ T cells and IL-17, IL-21, and IL-22 production. Strong CD8 T cell responses through antigen cross presentation by cDC1 cells were also observed. Single-cell RNAseq analysis revealed upregulation of Th17-promoting gene signatures in sorted cDC1 cells. Gene expression in isolated cDC2 cells was largely unaffected. Our finding represents a major shift of paradigm as we have documented functional plasticity in cDC1 cells.

Deficiency of SARS-CoV-2 T-cell responses after vaccination in long-term allo-HSCT survivors translates into abated humoral immunity.

Recipients of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for hematological diseases are at risk of severe disease and death from COVID-19. To determine the safety and immunogenicity of BNT162b2 and mRNA-1273 COVID-19 vaccines, samples from 50 infection-naive allo-HSCT recipients (median, 92 months from transplantation, range, 7-340 months) and 39 healthy controls were analyzed for serum immunoglobulin G (IgG) against the receptor binding domain (RBD) within spike 1 (S1) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2; anti-RBD-S1 IgG) and for SARS-CoV-2-specific T-cell immunity, reflected by induction of T-cell-derived interferon-γ in whole blood stimulated ex vivo with 15-mer SI-spanning peptides with 11 amino acid overlap S1-spanning peptides. The rate of seroconversion was not significantly lower in allo-transplanted patients than in controls with 24% (12/50) and 6% (3/50) of patients remaining seronegative after the first and second vaccination, respectively. However, 58% of transplanted patients lacked T-cell responses against S1 peptides after 1 vaccination compared with 19% of controls (odds ratio [OR] 0.17; P = .009, Fisher's exact test) with a similar trend after the second vaccination where 28% of patients were devoid of detectable specific T-cell immunity, compared with 6% of controls (OR 0.18; P = .02, Fisher's exact test). Importantly, lack of T-cell reactivity to S1 peptides after vaccination heralded substandard levels (<100 BAU/mL) of anti-RBD-S1 IgG 5 to 6 months after the second vaccine dose (OR 8.2; P = .007, Fisher's exact test). We conclude that although allo-HSCT recipients achieve serum anti-RBD-S1 IgG against SARS-CoV-2 after 2 vaccinations, a deficiency of SARS-CoV-2-specific T-cell immunity may subsequently translate into insufficient humoral responses.

Rapid Cytokine Release Assays for Analysis of Severe Acute Respiratory Syndrome Coronavirus 2-Specific T Cells in Whole Blood.

BACKGROUND: Waning of immunoglobulin G (IgG) antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) complicates the diagnosis of past infection. The durability of T-cell memory against SARS-CoV-2 remains unclear, and most current T-cell protocols are unsuited for large-scale automation. METHODS: Whole-blood samples from 31 patients with verified past coronavirus disease 2019 (COVID-19) and 46 controls, of whom 40 received COVID-19 vaccine, were stimulated with peptides spanning the nucleocapsid (NC) or spike 1 (S1) regions of SARS-CoV-2 and analyzed for interferon γ in supernatant plasma. Diagnostic accuracy of these assays was evaluated against serum anti-NC and anti-receptor-binding domain S1-IgG. RESULTS: Induction of interferon γ in whole blood by NC or S1 peptides diagnosed past COVID-19 with high accuracy (area under the receiver operating characteristic curve, 0.93 and 0.95, respectively). In accordance with previous studies, NC-IgG levels rapidly waned with only 5 of 17 patients (29%) remaining seropositive >180 days after infection. By contrast, NC peptide-induced T-cell memory responses remained in 13 of 17 study participants (76%) >180 days after infection (P = .01 for comparison with NC-IgG; McNemar test). After 2 vaccine doses, all 18 donors exhibited S1-specific T-cell memory. CONCLUSIONS: Cytokine release assays for the monitoring of T-cell memory in whole blood may be useful for evaluating complications following unverified past COVID-19 and for long-term assessment of vaccine-induced T-cell immunity. CLINICAL TRIALS REGISTRATION: EudraCT 2021-000349-42.

Receptor-specific TRAIL as a means to achieve targeted elimination of activated hepatic stellate cells.

Activated hepatic stellate cells (HSCs) are known to play a central role in liver fibrosis and their elimination is a crucial step toward the resolution and reversion of liver fibrosis. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a molecule that may contribute to the apoptotic removal of activated HSC through binding to its dedicated receptors. In the present study, we investigated the potential application of recombinant receptor-specific TRAIL proteins in the efficient elimination of activated HSCs. Our finding revealed differential contribution of TRAIL receptors among HSCs populations with activated hepatic stellate cells expresses more TRAIL receptors DR5. In vitro treatment of activated HSCs with DR5-specific or wild-type TRAIL variants induced a significant reduction in viability and extracellular matrix production, whereas no significant decrease in viability was associated with the treatment of cells by DR4-specific TRAIL. Our analysis indicate the successful application of the DR5 receptor-specific TRAIL variant in the targeted elimination of activated HSCs via interference with collagen production and simultaneous induction of apoptosis via activation of the caspase pathway. DR5 receptor-specific TRAIL may thus represent a new therapeutic compound for the treatment of liver fibrosis.

Targeted elimination of activated hepatic stellate cells by an anti-epidermal growth factor-receptor single chain fragment variable antibody-tumor necrosis factor-related apoptosis-inducing ligand (scFv425-sTRAIL).

BACKGROUND: Progressive liver fibrosis is the result of chronic liver injury and is characterized by the excessive accumulation of extracellular matrix that may result in liver failure. Activated hepatic stellate cells are known to play a central role in this process and their elimination is a crucial step towards the resolution and reversion of liver fibrosis. In the present study, we investigated the potential application of an anti-epidermal growth factor receptor single chain fragment variable antibody-tumor necrosis factor-related apoptosis-inducing ligand (scFv425-sTRAIL) fusion protein in the targeted elimination of activated hepatic stellate cells. METHODS: Activated hepatic stellate cells (LX2 cells) were treated by adenovirus-derived scFv425-sTRAIL to evaluate its effect on the viability and extracellular matrix production of this type of cells. RESULTS: In vitro treatment of activated hepatic stellate cells with scFv425-sTRAIL induced a significant reduction in viability (up to 100% reduction) and extracellular matrix production (60% reduction), yet no significant effect was observed on hepatic parenchymal cells. Blockage of the epidermal growth factor receptor (EGFR) by a monoclonal antibody significantly reduced the effectiveness of scFv425-sTRAIL in activated hepatic stellate cells, whereas a reduced effectivity was also observed after inhibition of the caspase pathway. CONCLUSIONS: Evidence is presented for the successful application of the scFv425-sTRAIL fusion protein in the targeted elimination of activated hepatic stellate cells via EGFR and simultaneous activation of the caspase pathway. scFv425-sTRAIL may thus represent a new therapeutic compound against liver fibrosis.

Prevalence of human Metapneumovirus (hMPV) in children with wheezing in Shiraz-Iran.

Human metapneumovirus (hMPV) infection plays an important role in the pediatric respiratory infections. The aim of this study was to determine the relationship between asthma and wheezing with hMPV in hospitalized children. Nasal pharyngeal swabs obtained from 120 children aged 1-60 months, hospitalized during a one year period, were tested for the hMPV by RT-PCR. HMPV was detected in 20 (16.6%) of patients suffering from wheezing. Some patients in addition to wheezing had asthma 10.8%. This infection occurred predominantly from October 2008 to September 2009.