Polyclonal Response Research Articles

Evaluating novel COVID-19 vaccines in the current chapter of the pandemic

Evaluating novel COVID-19 vaccines in the current chapter of the pandemic

1676P Active immunization with a multi-peptide B cell vaccine, targeting trastuzumab and pertuzumab binding sites, prevents the formation of HER-2/neu expressing lung metastases

The passive application of Trastuzumab and Pertuzumab has demonstrated remarkable success in the clinical outcome of patients with Her-2/neu positive metastatic breast cancer. However, active immunization with mimotope/B cell epitope-based vaccines can induce the production of the corresponding mAbs by the patient’s immune system. We have developed a B cell peptide-based Her-2/neu vaccine (HerVaxx) comprising Trastuzumab’s binding site. In clinical settings, the vaccine has been shown to reduce primary tumor growth by inducing polyclonal anti-tumor immune responses and immunological memory. The vaccine was also shown to result in loss of Her-2/neu expression, a phenomenon similarly observed in the clinic after treatment with Trastuzumab. Applying a Her-2/neu lung metastasis mouse model, here we tested a multi-peptide B cell vaccine containing HerVaxx and a mimotope of Pertuzumab for the prevention of lung metastases formation and investigated the loss of Her-2/neu expression. Mice were vaccinated with the multi-peptide vaccine, tail-vein injected with mammary carcinoma cells expressing human Her-2/neu, and histological evaluations of the mice lungs were carried out. A significantly reduced lung metastasis formation by active immunization with either the mimotope of Pertuzumab, HerVaxx, or the combination of both, was observed and associated with decreased lung weights and increased CD4+ and CD8+ T cells infiltration. Markedly, along with the overall reduction of lung weights and Her-2 positive metastases, the formation of Her-2/neu-negative tumors but with increased PD-L1 expression was observed. Our data might result in a multi-peptide B cell Her-2/neu vaccine serving as a secondary intervention in adjuvant settings to prevent tumor recurrence and spread. Furthermore, the loss of Her-2/neu expression suggests a combination therapy involving an anti-PD-L1 immune checkpoint inhibitor or an active immunization with a mimotope from PD-L1. Such combination therapy to alternately target Her-2/neu and PD-L1 might be adapted to the stage and progression phase of the disease and result in the remission of the metastases.

Characterization of prefusion-F-specific antibodies elicited by natural infection with human metapneumovirus.

Human metapneumovirus (hMPV) is a major cause of acute respiratory infections in infants and older adults, for which no vaccines or therapeutics are available. The viral fusion (F)glycoprotein is required for entry and is the primary target of neutralizing antibodies; however, little is known about the humoral immune response generated from natural infection. Here, using prefusion-stabilized F proteins to interrogate memory B cells from two older adults, we obtain over 700 paired non-IgM antibody sequences representing 563 clonotypes, indicative of a highly polyclonal response. Characterization of 136 monoclonal antibodies reveals broad recognition of the protein surface, with potently neutralizing antibodies targeting each antigenic site. Cryo-EM studies further reveal two non-canonical sites and the molecular basis for recognition of the apex of hMPV F by two prefusion-specific neutralizing antibodies. Collectively, these results provide insight into the humoral response to hMPV infection in older adults and will help guide vaccine development.

Structural evolution of severe acute respiratory syndrome coronavirus 2: Implications for adhesivity to angiotensin-converting enzyme 2 receptors and vaccines

The Omicron BA.5 variant currently accounts for the majority of cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (https://covid.cdc.gov/covid-data-tracker/#variant-proportions). This subvariant evolved from the Omicron lineage and became more contagious. Furthermore, data suggest that Omicron sublineages may evade polyclonal neutralizing antibody responses elicited by primary vaccine series [1,2]. Of note, development of antibodies to SARS-CoV-2 largely revolves around the viral spike (S) glycoprotein [3], which mediates host cell entry by binding to a cell surface receptor, angiotensin-converting enzyme 2 (ACE2, Fig. 1) [4].

Structure-selected RBM immunogens prime polyclonal memory responses that neutralize SARS-CoV-2 variants of concern.

Successful control of the COVID-19 pandemic depends on vaccines that prevent transmission. The full-length Spike protein is highly immunogenic but the majority of antibodies do not target the virus: ACE2 interface. In an effort to affect the quality of the antibody response focusing it to the receptor-binding motif (RBM) we generated a series of conformationally-constrained immunogens by inserting solvent-exposed RBM amino acid residues into hypervariable loops of an immunoglobulin molecule. Priming C57BL/6 mice with plasmid (p)DNA encoding these constructs yielded a rapid memory response to booster immunization with recombinant Spike protein. Immune sera antibodies bound strongly to the purified receptor-binding domain (RBD) and Spike proteins. pDNA primed for a consistent response with antibodies efficient at neutralizing authentic WA1 virus and three variants of concern (VOC), B.1.351, B.1.617.2, and BA.1. We demonstrate that immunogens built on structure selection can be used to influence the quality of the antibody response by focusing it to a conserved site of vulnerability shared between wildtype virus and VOCs, resulting in neutralizing antibodies across variants.

Both COVID-19 infection and vaccination induce high-affinity cross-clade responses to SARS-CoV-2 variants

SummaryThe B.1.1.529 (omicron) variant has rapidly supplanted most other SARS-CoV-2 variants. Using microfluidics-based antibody affinity profiling (MAAP), we have characterized affinity and IgG concentration in the plasma of 39 individuals with multiple trajectories of SARS-CoV-2 infection and/or vaccination. Antibody affinity was similar against the wild-type, delta, and omicron variants (KA ranges: 122 ± 155, 159 ± 148, 211 ± 307 μM-1, respectively), indicating a surprisingly broad and mature cross-clade immune response. Postinfectious and vaccinated subjects showed different IgG profiles, with IgG3 (p-value = 0.002) against spike being more prominent in the former group. Lastly, we found that the ELISA titers correlated linearly with measured concentrations (R = 0.72) but not with affinity (R = 0.29). These findings suggest that the wild-type and delta spike induce a polyclonal immune response capable of binding the omicron spike with similar affinity. Changes in titers were primarily driven by antibody concentration, suggesting that B-cell expansion, rather than affinity maturation, dominated the response after infection or vaccination.

Genetic and structural basis of the human anti-α-galactosyl antibody response

Humans lack the capacity to produce the Galα1-3Galβ1-4GlcNAc (α-gal) glycan, and produce anti-α-gal antibodies upon exposure to the carbohydrate on a diverse set of immunogens, including commensal gut bacteria, malaria parasites, cetuximab, and tick proteins. Here we use X-ray crystallographic analysis of antibodies from α-gal knockout mice and humans in complex with the glycan to reveal a common binding motif, centered on a germline-encoded tryptophan residue at Kabat position 33 (W33) of the complementarity-determining region of the variable heavy chain (CDRH1). Immunoglobulin sequencing of anti-α-gal B cells in healthy humans and tick-induced mammalian meat anaphylaxis patients revealed preferential use of heavy chain germline IGHV3-7, encoding W33, among an otherwise highly polyclonal antibody response. Antigen binding was critically dependent on the presence of the germline-encoded W33 residue for all of the analyzed antibodies; moreover, introduction of the W33 motif into naive IGHV3-23 antibody phage libraries enabled the rapid selection of α-gal binders. Our results outline structural and genetic factors that shape the human anti-α-galactosyl antibody response, and provide a framework for future therapeutics development.

Antigenic imprinting in SARS-CoV-2.

Antigenic imprinting in SARS-CoV-2.

The SARS-CoV-2 Delta variant induces an antibody response largely focused on class 1 and 2 antibody epitopes.

Exposure histories to SARS-CoV-2 variants and vaccinations will shape the specificity of antibody responses. To understand the specificity of Delta-elicited antibody immunity, we characterize the polyclonal antibody response elicited by primary or mRNA vaccine-breakthrough Delta infections. Both types of infection elicit a neutralizing antibody response focused heavily on the receptor-binding domain (RBD). We use deep mutational scanning to show that mutations to the RBD's class 1 and class 2 epitopes, including sites 417, 478, and 484-486 often reduce binding of these Delta-elicited antibodies. The anti-Delta antibody response is more similar to that elicited by early 2020 viruses than the Beta variant, with mutations to the class 1 and 2, but not class 3 epitopes, having the largest effects on polyclonal antibody binding. In addition, mutations to the class 1 epitope (e.g., K417N) tend to have larger effects on antibody binding and neutralization in the Delta spike than in the D614G spike, both for vaccine- and Delta-infection-elicited antibodies. These results help elucidate how the antigenic impacts of SARS-CoV-2 mutations depend on exposure history.

T Cell-Intrinsic Interleukin 17 Receptor A Signaling Supports the Establishment of Chronic Murine Gammaherpesvirus 68 Infection.

Gammaherpesviruses, such as human Epstein-Barr virus (EBV) and murine gammaherpesvirus 68 (MHV68), are species-specific, ubiquitous pathogens that are associated with multiple cancers, including B cell lymphomas. These viruses have a natural tropism for B cells and usurp B cell differentiation to drive a unique and robust polyclonal germinal center response to establish a long-term latent reservoir in memory B cells. The robust polyclonal germinal center response driven by gammaherpesvirus infection increases the risk for B cell transformation. Unsurprisingly, many gammaherpesvirus cancers are derived from germinal center or post-germinal center B cells. The viral and host factors that influence the gammaherpesvirus-driven germinal center response are not clearly defined. We previously showed that host interleukin 17 receptor A (IL-17RA) signaling promotes the establishment of chronic MHV68 infection and the MHV68-driven germinal center response. In this study, we found that T cell-intrinsic IL-17RA signaling recapitulates some proviral aspects of global IL-17RA signaling during MHV68 infection. Specifically, we found that T cell-intrinsic IL-17RA signaling supports the MHV68-driven germinal center response, the establishment of latency in the spleen, and viral reactivation in the spleen and peritoneal cavity. Our study unveils an unexpected finding where the T cell-specific IL-17RA signaling supports the establishment of a latent reservoir of a B cell-tropic gammaherpesvirus. IMPORTANCE Gammaherpesviruses, such as human EBV, establish lifelong infection in >95% of adults and are associated with B cell lymphomas. Gammaherpesviruses usurp the germinal center response to establish latent infection, and the germinal center B cells are thought to be the target of viral transformation. We previously found that global expression of IL-17RA promotes the establishment of chronic MHV68 infection and the MHV68-driven germinal center response. In this study, we showed that T cell-intrinsic IL-17RA signaling is necessary to promote the MHV68-driven germinal center response by supporting CD4+ T follicular helper cell expansion. We also found that T cell-intrinsic IL-17RA signaling contributes to but is not solely responsible for the systemic proviral role of IL-17RA signaling, highlighting the multifaceted function of IL-17RA signaling during MHV68 infection.

Complexity of Viral Epitope Surfaces as Evasive Targets for Vaccines and Therapeutic Antibodies.

The dynamic interplay between virus and host plays out across many interacting surfaces as virus and host evolve continually in response to one another. In particular, epitope-paratope interactions (EPIs) between viral antigen and host antibodies drive much of this evolutionary race. In this review, we describe a series of recent studies examining aspects of epitope complexity that go beyond two interacting protein surfaces as EPIs are typically understood. To structure our discussion, we present a framework for understanding epitope complexity as a spectrum along a series of axes, focusing primarily on 1) epitope biochemical complexity (e.g., epitopes involving N-glycans) and 2) antigen conformational/dynamic complexity (e.g., epitopes with differential properties depending on antigen state or fold-axis). We highlight additional epitope complexity factors including epitope tertiary/quaternary structure, which contribute to epistatic relationships between epitope residues within- or adjacent-to a given epitope, as well as epitope overlap resulting from polyclonal antibody responses, which is relevant when assessing antigenic pressure against a given epitope. Finally, we discuss how these different forms of epitope complexity can limit EPI analyses and therapeutic antibody development, as well as recent efforts to overcome these limitations.

IMPRINTER: An open label, multicenter, dose escalation/expansion, phase 1 study of imu-201 (PD1-Vaxx), a B-cell immunotherapy as monotherapy or in combination with atezolizumab, in adults with non-small cell lung cancer (IMU.201.101).

e21134 Background: Therapies with monoclonal antibodies targeting PD-1 and its ligands are associated with remarkable outcomes and have revolutionized cancer treatment (Honey 2017). However, patients treated with PD-1/PD-L1 blockade may develop “a primary or secondary resistance” to therapy (Sharma, Hu-Lieskovan et al. 2017). Contrary to monoclonal antibodies, chimeric B-cell cancer vaccines have the advantage of producing polyclonal B-cell antibodies that can potentially induce memory B- and T-cell responses, while reducing immune evasion and suppression. The hypothesis is that a polyclonal induced B-cell antibody response will be more effective or as effective with improved safety over current monoclonal antibody therapy. IMU-201 (PD1-Vaxx) is being developed using an active immunization approach to treat cancers that overexpress PD-L1 by inducing the production of anti-PD-1 antibodies with a peptide epitope designed to stimulate polyclonal antibodies against PD-1 (Kaumaya et al. 2020). Methods: The IMPRINTER study is an ongoing open-label dose escalation study of IMU-201 as monotherapy (Phase 1) or in combination with atezolizumab (Phase 1b) for patients with PD-L1 expressing non-small cell lung cancer (NSCLC). All patients enrolled in Phase 1 of the study must have previously received an immune checkpoint inhibitor and experienced disease progression. The primary objective is to evaluate the safety and tolerability of IMU-201 and identify the optimal biological dose (OBD). The secondary objective is to evaluate the efficacy of IMU-201 as monotherapy and in combination with atezolizumab. Humoral and cellular immunogenicity data will be evaluated, including IMU-201 and PD-1 specific antibodies (IgG, IgM), vaccine-specific cytokine levels, and regulatory and effector T and B cells. IMU-201 is administered by intramuscular (IM) injection on Day 1, Day 15, and Day 29. Dose-limiting toxicity (DLT) assessment is completed after 29 days on treatment. Tumor progression is evaluated according to RECIST 1.1 at Day 43 then every 42 days until progression or withdrawal. Results: In Phase 1, four patients were enrolled into each of the three cohorts at 10 μg/dose, 50 μg/dose and 100 μg/dose IMU-201 with no DLTs observed. In the 10 μg/dose cohort, one patient achieved CR and one patient SD; in the 50 μg/dose cohort, four patients achieved SD; and in the 100 μg/dose cohort, one patient achieved PR and two patients achieved SD. Within the 100 μg/dose cohort, one patient experienced an immune related pneumonitis after two IMU-201 administrations and discontinued from study treatment. Conclusions: IMU-201 had no observed DLT and demonstrated preliminary signs of efficacy. The study will therefore move into Phase 1b with IMU-201 being assessed in combination with atezolizumab. Clinical trial information: NCT04432207.

Analysis of a SARS-CoV-2 convalescent cohort identified a common strategy for escape of vaccine-induced anti-RBD antibodies by Beta and Omicron variants.

SummaryBackgroundEvolutionary pressure has led to the emergence of SARS-CoV-2 variants, with the most recent Omicron variant containing an unparalleled 30 mutations in the spike protein. Many of these mutations are expected to increase immune evasion, thus making breakthrough cases and re-infection more common.MethodsFrom June 2020 to December 2021 serial blood samples (initial post recovery, 6 months, 12 months) were collected from a COVID-19 convalescent cohort in Boston, MA. Plasma was isolated for use in Mesoscale Discovery based antibody binding assays. Unvaccinated donors or those vaccinated prior to the primary blood draw were excluded from this analysis, as were those who did not have at least two blood draws. Wilcoxon signed rank tests were used to compare pre- and post-vaccination titers and antibody response against different variants, while McNemar tests were used to compare the proportions of achieving ≥ 4 fold increases against different variants.FindingsForty-eight COVID convalescent donors with post-infection vaccination (hybrid immunity) were studied to evaluate the levels of cross-reactive antibodies pre- and post- vaccination against various SARS-CoV-2 Spike and RBD proteins. Vaccination with BNT162b2, mRNA-1273 or Ad26.COV2.S led to a 6.3 to 7.8 fold increase in anti-Spike antibody titers and a 7·0 to 7·4 fold increase in anti-WT, Alpha and Delta RBD antibody. However, a lower response was observed for Beta and Omicron RBDs with only 7/48 (15%) and 15/48 (31%) donors having a ≥4 fold increase in post-vaccination titers against Beta and Omicron RBDs. Structural analysis of the Beta and Omicron RBDs reveal a shared immune escape strategy involving residues K417-E484-N501 that is exploited by these variants of concern.InterpretationThrough mutations of the K417-E484-N501 triad, SARS-CoV-2 has evolved to evade neutralization by the class I/II anti-RBD antibody fraction of hybrid immunity plasma as the polyclonal antibody response post-vaccination shows limitations in the ability to solve the structural requirements to bind the mutant RBDs.FundingMassachusetts Consortium on Pathogen Readiness (280870.5116709.0016) and the National Institute of Allergy and Infectious Diseases (1R01AI161152-01A1).

T cell-intrinsic IL-17RA signaling in the spleen supports the establishment of chronic gammaherpesvirus infection.

Abstract Human gammaherpesviruses, such as Epstein-Barr virus (EBV), are human specific gammaherpesviruses that establish lifelong infection in >95% of adults and are associated with multiple cancers, including B cell lymphomas. These viruses have a natural tropism for B cells and usurp B cell differentiation to drive a robust polyclonal germinal center response, needed to establish a latent reservoir in memory B cells. The viral and host factors that influence the gammaherpesvirus-driven germinal center response are not clearly defined. Murine gammaherpesvirus 68 (MHV68) is a natural rodent pathogen that is genetically and biologically related to human gammaherpesviruses and provides a tractable animal model to study viral and host factors that impact chronic infection. Utilizing MHV68, we discovered that IL-17RA signaling is proviral and supports the establishment of chronic gammaherpesvirus infection. Specifically, loss of IL-17RA signaling during MHV68 infection significantly attenuated viral latency, reactivation, and the germinal center response. Importantly, the mechanism by which IL-17A signaling promotes MHV68-driven germinal center response and latency remained unclear. To gain mechanistic insight, we generated a mouse model of T cell specific IL-17RA deficiency. Excitingly, loss of IL-17RA specifically in T cells during MHV68 infection attenuated viral latency, reactivation, and the gammaherpesvirus-driven germinal center response in the spleen, similar to what was observed in the global IL-17RA−/− model. As T cells are not infected by MHV68, our finding unveils a novel crosstalk between T cell intrinsic IL-17A signaling and gammaherpesvirus infection of B cells, including virus-driven B cell differentiation. Supported by American Cancer Society Postdoctoral Award (134165-PF-19-176-01-MPC, C.N.J.)

Severe acute respiratory syndrome coronavirus 2 vaccination induces short-term polyclonal antibody response against seasonal endemic coronaviruses

Abstract Antibody (Ab) responses induced by endemic human coronaviruses (HCoVs) may affect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) specific Ab production due to cross-reactivity among these HCoVs. To explore this hypothesis, we collected serum samples of healthy healthcare workers (HCW) at four time-points: pre-vaccination, post-vaccination (1st and 2nd dose) and 6 months post-vaccination. We used N protein peptide-based enzyme-linked immunosorbent assays (ELISA) specific for alphaCoVs (NL63 and 229E) and betaCoVs (OC43 and HKU1) as well as N and S peptide-based ELISA for SARS-CoV-2. Low levels of SARS-CoV-2 N protein-specific Abs were present in some individuals prior to vaccination. The level remained constant following one or two doses of vaccine and declined after 6 months. SARS-CoV-2 S protein-specific Abs level were low in most individuals prior to vaccination, increased sharply following 1st and 2nd vaccinations and declined by 6 months. We observed low levels of pre-existing IgG Abs against endemic HCoVs prior to COVID-19 vaccination. However, like SARS-CoV-2-specific IgG Ab titers, they increased (1.6–1.9-fold) following SARS-CoV-2 vaccination and waned 6 months later. IgG Abs surge targeting HCoV N protein suggest polyclonal activation of memory B cell responses induced by the S gene based COVID-19 vaccines. The levels of IgG Abs against endemic HCoVs were 1.1–1.5 times higher in SARS-CoV-2 seropositive vs. seronegative individuals, suggesting serological cross-reactivity between SARS-CoV-2 and other HCoVs. In summary, short-term immunity (six months) against common cold causing HCoVs can be reactivated following COVID-19 vaccination however it requires booster dose for long term protection.

A dormant T-cell population with autoimmune potential exhibits low self-reactivity and infiltrates islets in type 1 diabetes.

The contribution of low-affinity T cells to autoimmunity in the context of polyclonal T-cell responses is understudied due to the limitations in their capture by tetrameric reagents and low level of activation in response to antigenic stimulation. As a result, low-affinity T cells are often disregarded as nonantigen-specific cells irrelevant to the immune response. Our study aimed to assess how the level of self-antigen reactivity shapes T-cell lineage and effector responses in the context of spontaneous tissue-specific autoimmunity observed in NOD mice. Using multicolor flow cytometry in combination with Nur77GFP reporter of TCR signaling, we identified a dormant population of T cells that infiltrated the pancreatic islets of prediabetic NOD mice, which exhibited reduced levels of self-tissue reactivity based on expression of CD5 and Nur77GFP . We showed that these CD5low T cells had a unique TCR repertoire and exhibited low activation and minimal effector function; however, induced rapid diabetes upon transfer. The CD4+ CD5low T-cell population displayed transcriptional signature of central memory T cells, consistent with the ability to acquire effector function post-transfer. Transcriptional profile of CD5low T cells was similar to T cells expressing a low-affinity TCR, indicating TCR affinity to be an important factor in shaping CD5low T-cell phenotype and function at the tissue site. Overall, our study suggests that autoimmune tissue can maintain a reservoir of undifferentiated central memory-like autoreactive T cells with pathogenic effector potential that might be an important source for effector T cells during long-term chronic autoimmunity.

Efficacy and Selectivity of Monovalent and Bivalent Vaccination Strategies to Protect against Exposure to Carfentanil, Fentanyl, and Their Mixtures in Rats.

Drug-related fatal overdoses have significantly increased in the past decade due to the widespread availability of illicit fentanyl and other potent synthetic opioids such as carfentanil. Deliberate or accidental consumption or exposure to carfentanil, fentanyl, and their mixture induces respiratory depression and bradycardia that can be difficult to reverse with the opioid receptor antagonist naloxone. Vaccines offer a promising strategy to reduce the incidence of fatalities associated with fentanyl-related substances, as well as treatment for opioid use disorder (OUD). This study reports monovalent and bivalent vaccination strategies that elicit polyclonal antibody responses effective in protecting against the pharmacological actions of carfentanil, fentanyl, or carfentanil/fentanyl mixtures. Rats were prophylactically immunized with individual conjugate vaccines containing either carfentanil- or fentanyl-based haptens, or their combination in bivalent vaccine formulations, and then challenged with carfentanil, fentanyl, or their mixture. First, these studies identified a lead vaccine protective against carfentanil-induced antinociception, respiratory depression, and bradycardia. Then, efficacy against both carfentanil and fentanyl was achieved through bivalent vaccination strategies that combined lead anti-carfentanil and anti-fentanyl vaccines via either heterologous prime/boost or co-administration immunization regimens. These preclinical data support the development of vaccines as a viable strategy to prevent toxicity from exposure to excessive doses of carfentanil, fentanyl, or their mixtures.

Enhancing and stabilization of cord blood regulatory T-cell suppressive function by human mesenchymal stem cell (MSC)-derived exosomes.

FOXP3+ regulatory T cells (Tregs) are central to maintaining peripheral tolerance and immune homeostasis. They have the potential to be developed as a cellular therapy to treat various clinical ailments such as autoimmune disorders, inflammatory diseases and to improve transplantation outcomes. However, a major question remains whether Tregs can persist and exert their function effectively in a disease state, where a broad spectrum of inflammatory mediators could inactivate Tregs. In this study, we investigated the potential of mesenchymal stem cell (MSC)-derived exosomes to promote and sustain Tregs function. MSC-conditioned media (MSC-CM) cultured Tregs were more suppressive in both polyclonal and allogeneic responses and were resistant to inflammatory stimulation in vitro compared with the controls. A similar enhancement of Treg function was also observed by culturing Tregs with MSC-derived exosomes alone. The enhanced suppressive activity and stability of Treg cultured in MSC-CM was reduced when exosomes were depleted from MSC-CM. We identified that MSC-derived exosomes could upregulate the expression of LC3(II/I), phosphorylate Jak3 and Stat5 to promote Treg survival, and regulate FOXP3 expression in Tregs. Overall, our study demonstrates that MSC-derived exosomes are capable of enhancing Hucb-Tregs function and stability by activating autophagy and Stat5 signalling pathways. Our findings provide a strong rationale for utilizing MSC-derived exosomes as an effective strategy to enhance Treg function, and improve the overall Tregs-based cell therapy landscape.

6 month serologic response to the Pfizer-BioNTech COVID-19 vaccine among healthcare workers.

AimHealthcare workers (HCWs) were among the first group of people vaccinated with the Pfizer-BioNTech Covid-19 vaccine (BNT162b2). Characterization of the kinetics of antibody response to vaccination is important to devise future vaccination strategies. To better characterize the antibody response to BNT162b2, we analyzed the kinetics of IgG and IgM antibody response to 5 different SARS-CoV-2 epitopes over a period of 6 months.Methods and resultsAn observational single-centered study was conducted to evaluate the temporal dynamics of anti-SARS-CoV-2 antibodies following immunization with two doses of BNT162b2. Anti-SARS-CoV-2 antibodies were assessed using the Maverick SARS-CoV-2 multi-antigen panel (Genalyte Inc.). Healthcare workers aged ≥18 receiving BNT162b2 vaccination who self-reported no prior symptoms of COVID-19 nor prior COVID-19 PCR test positivity, were included in this study. HCWs developed an IgG antibody response to SARS-CoV-2 Spike S1, Spike S1 receptor binding domain (RBD), Spike S1S2 and Spike S2 after vaccination. IgG response was observed at two weeks following immunization in most participant samples and continued to increase at week 4, but subsequently decreased significantly starting at 3 months and up to 6 months. In contrast, IgM response to respective epitopes was minimal.ConclusionMultiplex results demonstrate that, contrary to natural infection, immunization with BNT162b2 produces minimal anti-Spike IgM response. Polyclonal IgG response to Spike declined at 3 months and continued to do so up to 6 months.

Fc galactosylation of anti-platelet human IgG1 alloantibodies enhances complement activation on platelets.

Approximately 20% of patients receiving multiple platelet transfusions develop platelet alloantibodies, which can be directed against human leukocyte antigens (HLA) and, to a lesser extent, against human platelet antigens (HPA). These antibodies can lead to the rapid clearance of donor platelets, presumably through IgG-Fc receptor (FcγR)-mediated phagocytosis or via complement activation, resulting in platelet refractoriness. Strikingly, not all patients with anti-HLA or -HPA antibodies develop platelet refractoriness upon unmatched platelet transfusions. Previously, we found that IgG Fc glycosylation of anti-HLA antibodies was highly variable between patients with platelet refractoriness, especially with respect to galactosylation and sialylation of the Fc-bound sugar moiety. Here, we produced recombinant glycoengineered anti-HLA and anti-HPA-1a monoclonal antibodies with varying Fc galactosylation and sialylation levels and studied their ability to activate the classical complement pathway. We observed that anti-HLA monoclonal antibodies with different specificities, binding simultaneously to the same HLA-molecules, or anti-HLA in combination with anti-HPA-1a monoclonal antibodies interacted synergistically with C1q, the first component of the classical pathway. Elevated Fc galactosylation and, to a lesser extent, sialylation significantly increased the complement-activating properties of anti-HLA and anti-HPA-1a monoclonal antibodies. We propose that both the breadth of the polyclonal immune response, with recognition of different HLA epitopes and in some cases HPA antigens, and the type of Fc glycosylation can provide an optimal stoichiometry for C1q binding and subsequent complement activation. These factors can shift the effect of a platelet alloimmune response to a clinically relevant response, leading to complement-mediated clearance of donor platelets, as observed in platelet refractoriness.