Long-term efficacy of the peptide-based COVID-19 T cell activator CoVac-1 in healthy adults (preprint)

Background: T cell immunity is key for the control of viral infections including SARS-CoV-2, in particular with regard to immune memory and protection against arising genetic variants. Method: We recently evaluated a peptide-based SARS-CoV-2 T cell activator termed CoVac-1 in a first-in-human clinical trial and observed a favorable safety profile and induction of poly-specific T cell responses until month 3. Here, we report on long-term safety and efficacy data of CoVac-1 in healthy adults until month 12. Findings: CoVac-1 is well tolerated without long-term immune-related side effects and induces long-lasting anti-viral T cell responses in 100% of study participants. Potent expandability of CD4+ and CD8+ T cells targeting multiple different CoVac-1 T cell epitopes was observed 6 and 12 months after one single dose of CoVac-1. T cell responses were associated with the severity and the number of local adverse events at injection site. Beyond induction of T cell immunity, 89% of study participants developed CoVac-1-specific IgG antibody titers which associated with the intensity of the T cell response, indicating that CoVac-1-specific CD4+ T cells support the induction of B cell responses. Vaccination with approved COVID-19 vaccines boosted CoVac-1-specific T cell responses. Overall, a low SARS-CoV-2 infection rate was observed in the study population (8.3% of participants until month 12). Interpretation: Together, a single application of CoVac-1 elicits long-lived and broad SARS-CoV-2-specific T cell immunity, which further supports the current evaluation of our T cell activator in patients with congenital or acquired B cell defects (NCT04954469). Funding: This trial is funded by the Ministry of Science, Research and the Arts Baden-Wuerttemberg., Germany

Maintained imbalance of triglycerides, apolipoproteins, energy metabolites and cytokines in long-term COVID-19 syndrome (LTCS) patients (preprint)

Deep metabolomic, proteomic and immunologic phenotyping of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) patients have matched a wide diversity of clinical symptoms with potential biomarkers for coronavirus disease 2019 (COVID-19). Within here, several studies described the role of metabolites, lipoproteins and inflammation markers during infection and in recovered patients. In fact, after SARS-CoV-2 viral infection almost 20-30% of patients experience persistent symptoms even after 12 weeks of recovery which has been defined as long-term COVID-19 syndrome (LTCS). Emerging evidence revealed that a dysregulated immune system and persisting inflammation could be one of the key drivers of LTCS. However, how these small biomolecules such as metabolites, lipoprotein, cytokines and chemokines altogether govern pathophysiology is largely underexplored. Thus, a clear understanding of how these parameters in an integrated fashion could predict the disease course may help to stratify LTCS patients from acute COVID-19 or recovered specimens and would help to elucidate a potential mechanistic role of these biomolecules during the disease course. Here, we report an integrated analysis of blood serum and plasma by in vitro diagnostics research NMR spectroscopy and flow cytometry-based cytokine quantification in a total of 125 individuals (healthy controls (HC; n=73), recovered (n=12), acute (n=7) and LTCS (n=33)). We identified that in LTCS patients lactate and pyruvate were significantly different from either healthy controls or acute COVID-19 patients. Further correlational analysis of cytokines and metabolites indicated that creatine, glutamine, and high-density lipoprotein (HDL) phospholipids were distributed differentially amongst patients or individuals. Of note, triglycerides and several lipoproteins (apolipoproteins Apo-A1 and A2) in LTCS patients demonstrate COVID-19-like alterations compared to HC. Interestingly, LTCS and acute COVID-19 samples were distinguished mostly by their creatinine, phenylalanine, succinate, 3-hydroxybutyrate (3-HB) and glucose concentrations, illustrating an imbalanced energy metabolism. Most of the cytokines and chemokines were present at low levels in LTCS patients compared with HC except IL-18 chemokine, which tended to be higher in LTCS patients and correlated positively with several amino acids (creatine, histidine, leucine, and valine), metabolites (lactate and 3-HB) and lipoproteins. The identification of these persisting plasma metabolites, lipoprotein and inflammation alterations will help to better stratify LTCS patients from other diseases and could help to predict the ongoing severity of LTCS patients.

Antibody binding and ACE2 binding inhibition is significantly reduced for the Omicron variant compared to all other variants of concern (preprint)

The rapid emergence of the Omicron variant and its large number of mutations has led to its classification as a variant of concern (VOC) by the WHO. Initial studies on the neutralizing response towards this variant within convalescent and vaccinated individuals have identified substantial reductions. However many of these sample sets used in these studies were either small, uniform in nature, or were compared only to wild-type (WT) or, at most, a few other VOC. Here, we assessed IgG binding, (Angiotensin-Converting Enzyme 2) ACE2 binding inhibition, and antibody binding dynamics for the omicron variant compared to all other VOC and variants of interest (VOI), in a large cohort of infected, vaccinated, and infected and then vaccinated individuals. While omicron was capable of binding to ACE2 efficiently, antibodies elicited by infection or immunization showed reduced IgG binding and ACE2 binding inhibition compared to WT and all VOC. Among vaccinated samples, antibody binding responses towards omicron were only improved following administration of a third dose. Overall, our results identify that omicron can still bind ACE2 while pre-existing antibodies can bind omicron. The extent of the mutations appear to inhibit the development of a neutralizing response, and as a result, omicron remains capable of evading immune control.

Reduced serum neutralization capacity against SARS-CoV-2 variants in a multiplex ACE2 RBD competition assay (preprint)

As global vaccination campaigns against SARS-CoV-2 proceed, there is emerging interest in the longevity of immune protection, especially with regard to increasingly infectious virus variants. Neutralizing antibodies (Nabs) targeting the receptor binding domain (RBD) of SARS-CoV-2 are promising correlates of protective immunity and have been successfully used for prevention and therapy. To assess neutralizing capacity, we developed a bead-based multiplex ACE2 RBD competition assay as a large scalable, time-, cost-, and material-saving alternative to infectious live-virus neutralization tests. By mimicking the interaction between ACE2 and RBD, this assay detects the presence of Nabs against SARS-CoV2 in serum. Using this multiplex approach allows the simultaneous analysis of Nabs against all SARS-CoV-2 variants of concern and variants of interest in a single well. Following validation, we analyzed 325 serum samples from 186 COVID-19 patients of varying severity. Neutralization capacity was reduced for all variants examined compared to wild-type, especially for those displaying the E484K mutation. The neutralizing immune response itself, while highly individualistic, positively correlates with IgG levels. Neutralization capacity also correlated with disease severity up to WHO grade 7, after which it reduced.

Severe COVID-19 infection is associated with increased antibody-mediated platelet apoptosis (preprint)

The pathophysiology of COVID-19 associated thrombosis seems to be multifactorial, involving interplay between cellular and plasmatic elements of the hemostasis. We hypothesized that COVID-19 is accompanied by platelet apoptosis with subsequent alteration of the coagulation system. We investigated depolarization of mitochondrial inner transmembrane potential ({Delta}{Psi}m), cytosolic calcium (Ca2+) concentration, and phosphatidylserine (PS) externalization by flow cytometry. Platelets from intensive care unit (ICU) COVID-19 patients (n=21) showed higher {Delta}{Psi}m depolarization, cytosolic Ca2+ concentration and PS externalization, compared to healthy controls (n=18) and COVID-19 non-ICU patients (n=4). Moreover significant higher cytosolic Ca2+ concentration and PS was observed compared to septic ICU control group (ICU control). In ICU control group (n=5; non-COVID-19 ICU) cytosolic Ca2+ concentration and PS externalization was comparable to healthy control, with an increase in {Delta}{Psi}m depolarization. Sera from ICU COVID-19 patients induced significant increase in apoptosis markers ({Delta}{Psi}m depolarization, cytosolic Ca2+ concentration and PS externalization) compared to healthy volunteer and septic ICU control. Interestingly, immunoglobulin G (IgG) fractions from COVID-19 patients induced an Fc gamma receptor IIA dependent platelet apoptosis ({Delta}{Psi}m depolarization, cytosolic Ca2+ concentration and PS externalization). Enhanced PS externalization in platelets from ICU COVID-19 patients was associated with increased sequential organ failure assessment (SOFA) score (r=0.5635) and D-Dimer (r=0.4473). Most importantly, patients with thrombosis had significantly higher PS externalization compared to those without. The strong correlations between apoptosis markers and increased D-Dimer levels as well as the incidence of thrombosis may indicate that antibody-mediated platelet apoptosis potentially contributes to sustained increased thromboembolic risk in ICU COVID-19 patients.