Ab Production Research Articles

FcγRIIB modulates splenic germinal center response against immune subversion during acute influenza A virus infection

BackgroundB cells are essential for providing humoral protection against acute influenza A virus (IAV) infection. FcγRIIB, a regulator of antibody (Ab) production, influences immune responses during pathogen infections, but its specific impact on humoral protection and B cell-mediated responses against IAV remains unclear. MethodsTo investigate FcγRIIB's role in host defense and B cell function during acute IAV infection, we generated mice with systemic FcγRIIB deficiency, functional impairment, and B cell-specific FcγRIIB deletion. We infected these mice with PR8 (H1N1) or Hkx31 (H3N2) IAVs and evaluated body weight preservation, survival rates, Ab production, viral neutralization, Ab affinity maturation, and germinal center B cell development. ResultsMice lacking FcγRIIB or with impaired function showed improved protection, preserved body weight, and increased survival rates during IAV infection. Notably, mice with haploinsufficient FcγRIIB function displayed protective effects. Selective deficiency of FcγRIIB in B cells led to enhanced Ab production, resulting in elevated IAV-specific Abs in the serum with superior viral neutralizing potency. However, the impact on the affinity maturation index of virus-specific Abs was modest. Accordingly, FcγRIIB-deficient B cells maintained normal germinal center B cell development during IAV infection, whereas wild-type mice exhibited delayed differentiation. ConclusionOur research underscores the pivotal role of FcγRIIB in host defense and B cell-mediated immunity during acute IAV infection. Additionally, our discoveries hold implications for antiviral treatments, particularly during the initial stages of IAV infection, aimed at enhancing the host's humoral immune response.

Developing Topics.

Alzheimer's disease (AD) is associated with the accumulation of neuropathological beta-amyloid (Ab) plaques, which is thought to be caused by an imbalance between Ab overproduction and dysfunctional Ab clearance. Both animal and human studies have shown that increased cerebrospinal fluid (CSF) levels of Ab peptides, especially Ab-40 and Ab-38 due to their high solubility, may be indicative of overall Ab dysregulation in preclinical AD, years before pathological Ab plaques begin to aggregate. The apolipoprotein E (APOE) e4 allele, which is the strongest genetic risk factor for AD, has been linked to impaired Ab clearance and disruption of the amyloid precursor protein (APP) pathway that is responsible for Ab production. However, the association between early Ab dysregulation and other genetic risk variants remains understudied. Thus, we aimed to explore whether genetic risk for AD is related to early markers of Ab overproduction by examining associations between polygenic risk scores (PRS) and CSF Ab-40 and Ab-38 in an AD-vulnerable population. We selected 88 non-demented Veterans (Age: M = 68.22, SD = 3.75 years) from the Alzheimer's Disease Neuroimaging Initiative-Department of Defense (ADNI-DOD) database with available genetic markers and CSF measures for Ab-40 and Ab-38. Genetic propensity for AD was assessed using genome-wide PRS, both with and without the APOE region, employing a p-value threshold < 0.01. CSF Ab concentrations were measured using a 2D-UPLC-tandem mass spectrometry method outlined by ADNI-DOD. Partial spearman's correlations related PRS to CSF Ab-38 and Ab-40 levels adjusting for age. CSF Ab-40 levels were positively associated with PRS for AD (rhopartial = 0.22, p = 0.037). When variants in the APOE region were excluded from the score, the relationship between CSF Ab-40 and PRS remained significant (rhopartial = 0.23, p = 0.035). The relationship between PRS for AD and CSF Ab-38 levels was not significant, regardless of whether APOE regions were included in the score (all p's > 0.1). Genetic risk for AD may lead to early Ab overproduction, as evidenced by elevated CSF Ab levels, particularly Ab-40. Other genetic variants, in addition to APOE, may play an integral role in disrupting the APP pathway and dysregulating Ab production.

A Novel Adenovirus (Adv)-Vaccinated Murine Model Simulates the Prothrombotic Vaccine-Induced Immune Thrombotic Thrombocytopenia (VITT) Clinical Disorder

Background: VITT has emerged as a rare (10 -5 to 10 -6 vaccinated individuals), but life-threatening side-effect of post-AdV-based COVID-19 vaccination, characterized by rapidly progressive catastrophic thrombosis, thrombocytopenia and high levels of platelet-activating anti-platelet factor 4 (PF4) antibodies (Abs). While the disorder resembles heparin-induced thrombocytopenia (HIT) in that both anti-PF4 Abs activate platelets via FcγRIIA in the presence of PF4, the Abs differ in their requirements for heparin, i.e. HIT Abs require heparin, but VITT Abs do not. Moreover, alanine substitutions in PF4 revealed distinct antigenic sites in these two prothrombotic disorders. We observed that the defined VITT antigenic site, unlike the HIT antigenic site, is conserved on a second platelet-specific chemokine structurally similar to PF4, neutrophil activating peptide 2 (NAP2) and its precursor platelet-basic protein (PBP) and proposed that PBP as well as PF4 may be able to induce VITT. Objective: We have now generated a murine model of VITT (mVITT) to better understand the pathogenesis of this immune disorder and to examine whether PBP may be involved in this process. Methods: Transgenic FcγRIIA mice (n&amp;gt;5/group) that lack mPF4 (PF4 KO), but express human (h) PF4 were injected intramuscularly with 5x10 9 viral particles/mouse of ChadOx1-nCoV-19 (“nCoV-19 AdV”) leading to expression of the COVID-19 spike protein or control ChadOx-GFP, (“GFP AdV”) expressing the green fluorescent protein. To augment mVITT Ab production, mice also received CpG oligodeoxynucleotides (ODN 2395 or “ODN”, 10µg/mouse i.p for 5 days) as an adjuvant. In these latter experiments, ODN only- and carrier alone- (physiologic buffered saline, “PBS”) injected mice served as controls. The clinical course, blood counts and ELISA for Abs to spike protein, hPF4 and mPBP/NAP2 were followed once a week and organs were harvested for histology at the endpoint. Results: Only mice that received the nCoV-19 developed anti-spike protein Abs. In the studies without ODN, 1 out of the 20 mice injected with either nCoV-19 or GFP AdV developed progressive thrombocytopenia, massive thrombi in the lungs, heart and brain, and died at day 43, one day after receiving a second dose of GFP. This mouse developed a high titer of anti-mNAP2 Abs, but no significant increase in anti-hPF4 ( Table 1). In the studies with ODN adjuvant, 2 out of the 5 mice injected with nCoV-19 AdV and one of the 5 mice injected with GFP AdV developed thrombocytopenia and died at days 14, 39 and 40 after the 1 st dose and prior to being able to receive a second dose. Gross histology of the brains showed areas of infarction and/or hemorrhage. All three affected mice had raised anti-hPF4 Abs ( Table 1). Further characterization of these mice revealed newly formed anti-hPF4 Abs that were heparin independent. Moreover, two monoclonal (mo) Abs, a humanized version of KKO (G4 KKO) that only binds to the HIT antigenic site and 1E12 shown to bind to the VITT antigenic site, were tested for their ability to block the binding of mVITT Ab to hPF4. Only 1E12 blocked that binding (Figure 1). None of the ODN or PBS control mice developed thrombocytopenia or died ( Table 1). Conclusions: We have developed a murine model to simulate the development of VITT that supports AdV:chemokine complex formation as being key to the development of Abs and the prothrombotic state in VITT. This model indicates that anti-PBP/NAP2 Abs can develop after AdV immunization. We also show that the inclusion of an oligonucleotide adjuvant switched antigenic targeting to hPF4. The anti-hPF4 Ab did not develop when adjuvant alone was given, and the resultant Abs clearly bound as VITT-like Abs and not HIT Abs. The basis for the apparent ability of ODN to switch which chemokine is immunogenic will need future study. This model system may be useful in designing AdV that do not induce VITT as backbone for vaccines in the future as well.

The trajectory of human B-cell function, immune deficiency, and allergy revealed by inborn errors of immunity.

The essential role of B cells is to produce protective immunoglobulins (Ig) that recognize, neutralize, and clear invading pathogens. This results from the integration of signals provided by pathogens or vaccines and the stimulatory microenvironment within sites of immune activation, such as secondary lymphoid tissues, that drive mature B cells to differentiate into memory B cells and antibody (Ab)-secreting plasma cells. In this context, B cells undergo several molecular events including Ig class switching and somatic hypermutation that results in the production of high-affinity Ag-specific Abs of different classes, enabling effective pathogen neutralization and long-lived humoral immunity. However, perturbations to these key signaling pathways underpin immune dyscrasias including immune deficiency and autoimmunity or allergy. Inborn errors of immunity that disrupt critical immune pathways have identified non-redundant requirements for eliciting and maintaining humoral immune memory but concomitantly prevent immune dysregulation. Here, we will discuss our studies on human B cells, and how our investigation of cytokine signaling in B cells have identified fundamental requirements for memory B-cell formation, Ab production as well as regulating Ig class switching in the context of protective versus allergic immune responses.

Humoral and Cellular Immunity Are Significantly Affected in Renal Transplant Recipients, following Vaccination with BNT162b2.

Renal transplant recipients (RTRs) tend to mount weaker immune responses to vaccinations, including vaccines against the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Humoral immunity was assessed using anti-receptor binding domain (RBD) and neutralizing antibodies (NAb) serum levels measured by ELISA, and cellular immunity was assessed using T-, B-, NK, natural killer-like T (NKT)-cell subpopulations, and monocytes measured by flow cytometry, and also specific T-cell immunity, at predefined time points after BNT162b2 vaccination, in 57 adult RTRs. Administration of three booster doses was necessary to achieve anti-RBD and NAb protective levels in almost all patients (92.98%). Ab production, at several time points, was positively correlated with the corresponding renal function and inversely correlated with hemodialysis vintage (HDV) and treatment with mycophenolic acid (MPA). A gradual rise in several cell subpopulations, including total lymphocytes (p = 0.026), memory B cells (p = 0.028), activated CD4 (p = 0.005), and CD8 cells (p = 0.001), was observed even after the third vaccination dose, while a significant reduction in CD3+PD1+ (p = 0.002), NKT (p = 0.011), and activated NKT cells (p = 0.034) was noted during the same time interval. Moreover, SARS-CoV-2-specific T-cells were present in 41% of the patients who were unable to develop Nabs, and their positivity rates four months after the second dose were in inverse correlation with monocytes (p = 0.045) and NKT cells (p = 0.01). SARS-CoV-2-specific T-cell responses preceded the humoral ones, while two booster doses were needed for this group of immunocompromised patients to mount a protective immune response.

Cutting Edge: TLR2 Signaling in B Cells Promotes Autoreactivity to DNA via IL-6 Secretion.

Autoantibodies to chromatin and dsDNA are a hallmark of systemic lupus erythematosus (SLE). In a mouse model of monogenic human SLE caused by DNASE1L3 deficiency, the anti-DNA response is dependent on endosomal nucleic acid-sensing TLRs TLR7 and TLR9. In this study, we report that this response also required TLR2, a surface receptor for microbial products that is primarily expressed on myeloid cells. Cell transfers into lymphopenic DNASE1L3-deficient mice showed that TLR2 was required for anti-DNA Ab production by lymphocytes. TLR2 was detectably expressed on B cells and facilitated the production of IL-6 by B cells activated in the presence of microbial products. Accordingly, treatment with broad-spectrum antibiotics or Ab-mediated blockade of IL-6 delayed the anti-DNA response in DNASE1L3-deficient mice. These studies reveal an unexpected B cell-intrinsic role of TLR2 in systemic autoreactivity to DNA, and they suggest that microbial products may synergize with self-DNA in the activation of autoreactive B cells in SLE.

B-1 B Cell-Derived Natural Antibodies against N-Acetyl-d-Glucosamine Suppress Autoimmune Diabetes Pathogenesis.

Environmental factors and host microbiota strongly influence type 1 diabetes (T1D) progression. We report that neonatal immunization with group A Streptococcus suppresses T1D development in NOD mice by promoting clonal expansion of N-acetyl-d-glucosamine (GlcNAc)-specific B-1 B cells that recognize pancreatic β cell-derived Ags bearing GlcNAc-containing posttranslational modifications. Early exposure to Lancefield group A cell-wall carbohydrate Ags increased production of GlcNAc-reactive serum Abs and enhanced localization of innate-like GlcNAc-specific B cells to pancreatic tissue during T1D pathogenesis. We show that B-1 B cell-derived GlcNAc-specific IgM engages apoptosis-associated β cell Ags, thereby suppressing diabetogenic T cell activation. Likewise, adoptively transferring GlcNAc-reactive B-1 B cells significantly delayed T1D development in naive recipients. Collectively, these data underscore potentially protective involvement of innate-like B cells and natural Abs in T1D progression. These findings suggest that previously reported associations of reduced T1D risk after GAS infection are B cell dependent and demonstrate the potential for targeting the natural Ab repertoire in considering therapeutic strategies for T1D.

VH2+ Antigen-Experienced B Cells in the Cerebrospinal Fluid Are Expanded and Enriched in Pediatric Anti-NMDA Receptor Encephalitis.

Pediatric and adult autoimmune encephalitis (AE) are often associated with Abs to the NR1 subunit of the N-methyl-d-aspartate (NMDA) receptor (NMDAR). Very little is known regarding the cerebrospinal fluid humoral immune profile and Ab genetics associated with pediatric anti-NMDAR-AE. Using a combination of cellular, molecular, and immunogenetics tools, we collected cerebrospinal fluid from pediatric subjects and generated 1) flow cytometry data to calculate the frequency of B cell subtypes in the cerebrospinal fluid of pediatric subjects with anti-NMDAR-AE and controls, 2) a panel of recombinant human Abs from a pediatric case of anti-NMDAR-AE that was refractory to treatment, and 3) a detailed analysis of the Ab genes that bound the NR1 subunit of the NMDAR. Ag-experienced B cells including memory cells, plasmablasts, and Ab-secreting cells were expanded in the pediatric anti-NMDAR-AE cohort, but not in the controls. These Ag-experienced B cells in the cerebrospinal fluid of a pediatric case of NMDAR-AE that was refractory to treatment had expanded use of variable H chain family 2 (VH2) genes with high somatic hypermutation that all bound to the NR1 subunit of the NMDAR. A CDR3 motif was identified in this refractory case that likely drove early stage activation and expansion of naive B cells to Ab-secreting cells, facilitating autoimmunity associated with pediatric anti-NMDAR-AE through the production of Abs that bind NR1. These features of humoral immune responses in the cerebrospinal fluid of pediatric anti-NMDAR-AE patients may be relevant for clinical diagnosis and treatment.

Pre-existing CD4 T cell help boosts antibody responses but has limited impact on germinal center, antigen-specific B cell frequencies after influenza infection.

The influenza virus is a persistent burden on global health, with seasonal vaccines providing incomplete protection. CD4+ T cells help shape B cell and antibody responses; however, the selectivity of help and the effect on various antigen-specific B cell populations have not been fully elucidated. Here, we studied the specificity, selectivity, and influence of nucleoprotein (NP) CD4+ T cells on the magnitude and quality of hemagglutinin (HA) and NP-specific B cells and antibody responses. We identified immunodominant peptides and showed that peptide immunization was sufficient to induce CD4+ cells with Th1 and Tfh phenotypes. Surprisingly, while preexisting CD4+ T cells enhanced the influx of total germinal center (GC) B cells in the mediastinal lymph node after infection, this was not reflected by an increase in the frequency of antigen-specific cells within the GC. Furthermore, we demonstrated that NP-specific help was able to accelerate the kinetics and magnitude of the Ab response for NP but not for HA. Overall, our results showed that pre-existing CD4+ T cells provide strong cognate help during immunization or infection to enhance Ab production but not antigen-specific GC or memory B cells.

Proteolipid Protein-Induced Mouse Model of Multiple Sclerosis Requires B Cell-Mediated Antigen Presentation.

The pathogenic role B cells play in multiple sclerosis is underscored by the success of B cell depletion therapies. Yet, it remains unclear how B cells contribute to disease, although it is increasingly accepted that mechanisms beyond Ab production are involved. Better understanding of pathogenic interactions between B cells and autoreactive CD4 T cells will be critical for novel therapeutics. To focus the investigation on B cell:CD4 T cell interactions invivo and invitro, we previously developed a B cell-dependent, Ab-independent experimental autoimmune encephalomyelitis (EAE) mouse model driven by a peptide encompassing the extracellular domains of myelin proteolipid protein (PLPECD). In this study, we demonstrate that B cell depletion significantly inhibited PLPECD-induced EAE disease, blunted PLPECD-elicited delayed-type hypersensitivity reactions invivo, and reduced CD4 T cell activation, proliferation, and proinflammatory cytokine production. Further, PLPECD-reactive CD4 T cells sourced from B cell-depleted donor mice failed to transfer EAE to naive recipients. Importantly, we identified B cell-mediated Ag presentation as the critical mechanism explaining B cell dependence in PLPECD-induced EAE, where bone marrow chimeric mice harboring a B cell-restricted MHC class II deficiency failed to develop EAE. B cells were ultimately observed to restimulate significantly higher Ag-specific proliferation from PLP178-191-reactive CD4 T cells compared with dendritic cells when provided PLPECD peptide in head-to-head cultures. We therefore conclude that PLPECD-induced EAE features a required pathogenic B cell-mediated Ag presentation function, providing for investigable B cell:CD4 T cell interactions in the context of autoimmune demyelinating disease.

Productive Germinal Center Responses Depend on the Nature of Stimuli Received by Anti-Insulin B Cells in Type 1 Diabetes-Prone Mice.

Islet autoantibodies, including those directed at insulin, predict type 1 diabetes (T1D) in mice and humans and signal immune tolerance breach by B lymphocytes. High-affinity insulin autoantibodies and T follicular helper cell involvement implicate germinal centers (GCs) in T1D. The VH125SD BCR transgenic model, in which 1-2% of peripheral B lymphocytes recognize insulin, enables direct study of insulin-binding B cells. Our prior studies showed that anti-insulin B cell receptor transgene site-directed to H chain locus mice fail to generate insulin Ab following T-dependent immunization, but it was unclear whether anti-insulin B cells were blocked for GC initiation, survival, or differentiation into Ab-secreting cells. Here, we show that insulin-binding B cells in T1D-prone anti-insulin B cell receptor transgene site-directed to H chain locus mice can spontaneously adopt a GC phenotype and undergo class switching to the IgG1 isotype, with little if any switching to IgG2b. T-dependent immunizations with insulin SRBC or insulin CFA drove anti-insulin B lymphocytes to adopt a GC phenotype, despite blunted insulin Ab production. Dual immunization against self (insulin) and foreign (4-hydroxy-3-nitrophenylacetyl hapten conjugated to keyhole limpet hemocyanin) Ags showed an anti-insulin (but not anti-4-hydroxy-3-nitrophenylacetyl) Ab block that tracked with increased expression of the apoptosis marker, activated caspase 3, in self-reactive GC B cells. Finally, T-independent immunization with insulin conjugated to Brucella abortus ring test Ag released immune tolerance to allow robust expansion of anti-insulin GC B cells and IgG-switched insulin Ab production. Overall, these data pinpoint GC survival and Ab-secreting cell differentiation as immune tolerance blocks that limit T-dependent, but not T-independent, stimulation of anti-insulin B cell responses.

Natural Microbial Exposure from the Earliest Natural Time Point Enhances Immune Development by Expanding Immune Cell Progenitors and Mature Immune Cells.

Microbial experience fundamentally shapes immunity, particularly during the perinatal period when the immune system is underdeveloped, and novel microbial encounters are common. Most animal models are raised in specific pathogen-free (SPF) conditions with relatively uniform microbial communities. How SPF housing conditions alter early-life immune development relative to natural microbial exposure (NME) has not been thoroughly investigated. In this article, we compare immune development in SPF-raised mice with mice born from immunologically experienced mothers in microbially diverse environments. NME induced broad immune cell expansion, including naive cells, suggesting mechanisms besides activation-induced proliferation contribute to the increase in immune cell numbers. We found NME conditions also expanded immune cell progenitor cell populations in the bone marrow, suggesting microbial experience enhances immune development at the earliest stages of immune cell differentiation. Multiple immune functions characteristically impaired in infants were also enhanced by NME, including T cell memory and Th1 polarization, B cell class switching and Ab production, proinflammatory cytokine expression, and bacterial clearance after Listeria monocytogenes challenge. Collectively, our studies reveal numerous impairments in immune development in SPF conditions relative to natural immune development.

Investigation of the frequencies of various B cell populations in non-responder healthcare workers in comparison with responders to hepatitis B virus vaccination.

Hepatitis B is a major global health problem. More than 90% of hepatitis B-vaccinated immunocompetent adults become fully immune. The main purpose of vaccination is immunization. Whether non-responders have a lower percentage of total or antigen-specific memory B cells in comparison with responders is still controversial. We aimed to assess and compare the frequency of various B cell subpopulations in non-responders and responders. Fourteen responders and 14 non-responders of hospital healthcare workers were enrolled in this study. We used flow cytometry to evaluate various CD19+ B cell subpopulations using fluorescent-labeled antibodies against CD19, CD10, CD21, CD27 and IgM and ELISA to evaluate total anti-HBs antibodies. We found no significant differences in the frequency of various B cell subpopulations between the non-responder and responder groups. Furthermore, the frequency of the isotype-switched memory B cell population was significantly higher in the atypical memory B cell subset compared with the classical memory B cell subset in the responder and total groups (p=0.010 and 0.003, respectively). Responders and non-responders to HBsAg vaccine had comparable memory B cell populations. Whether anti-HBs Ab production has a correlation with the level of class switching in B lymphocytes in healthy vaccinated individuals needs further investigation.

Essential role of an ERV-derived Env38 protein in adaptive humoral immunity against an exogenous SVCV infection in a zebrafish model.

Endogenous retroviruses (ERVs) are the relics of ancient retroviruses occupying a substantial fraction of vertebrate genomes. However, knowledge about the functional association of ERVs with cellular activities remains limited. Recently, we have identified approximately 3,315 ERVs from zebrafish at genome-wide level, among which 421 ERVs were actively expressed in response to the infection of Spring viraemia of carp virus (SVCV). These findings demonstrated the previously unrecognized activity of ERVs in zebrafish immunity, thereby making zebrafish an attractive model organism for deciphering the interplay among ERVs, exogenous invading viruses, and host immunity. In the present study, we investigated the functional role of an envelope protein (Env38) derived from an ERV-E5.1.38-DanRer element in zebrafish adaptive immunity against SVCV in view of its strong responsiveness to SVCV infection. This Env38 is a glycosylated membrane protein mainly distributed on MHC-II+ antigen-presenting cells (APCs). By performing blockade and knockdown/knockout assays, we found that the deficiency of Env38 markedly impaired the activation of SVCV-induced CD4+ T cells and thereby led to the inhibition of IgM+/IgZ+ B cell proliferation, IgM/IgZ Ab production, and zebrafish defense against SVCV challenge. Mechanistically, Env38 activates CD4+ T cells by promoting the formation of pMHC-TCR-CD4 complex via cross-linking MHC-II and CD4 molecules between APCs and CD4+ T cells, wherein the surface subunit (SU) of Env38 associates with the second immunoglobin domain of CD4 (CD4-D2) and the first α1 domain of MHC-IIα (MHC-IIα1). Notably, the expression and functionality of Env38 was strongly induced by zebrafish IFNφ1, indicating that env38 acts as an IFN-stimulating gene (ISG) regulated by IFN signaling. To the best of our knowledge, this study is the first to identify the involvement of an Env protein in host immune defense against an exogenous invading virus by promoting the initial activation of adaptive humoral immunity. It improved the current understanding of the cooperation between ERVs and host adaptive immunity.

Bacterial expression systems based on Tymovirus-like particles for the presentation of vaccine antigens.

Virus-like particles (VLPs) are virus-derived artificial nanostructures that resemble a native virus-stimulating immune system through highly repetitive surface structures. Improved safety profiles, flexibility in vaccine construction, and the ease of VLP production and purification have highlighted VLPs as attractive candidates for universal vaccine platform generation, although exploration of different types of expression systems for their development is needed. Here, we demonstrate the construction of several simple Escherichia coli expression systems for the generation of eggplant mosaic virus (EMV) VLP-derived vaccines. We used different principles of antigen incorporation, including direct fusion of EMV coat protein (CP) with major cat allergen Feld1, coexpression of antigen containing and unmodified (mosaic) EMV CPs, and two coexpression variants of EMV VLPs and antigen using synthetic zipper pair 18/17 (SYNZIP 18/17), and coiled-coil forming peptides E and K (Ecoil/Kcoil). Recombinant Fel d 1 chemically coupled to EMV VLPs was included as control experiments. All EMV-Feld1 variants were expressed in E. coli, formed Tymovirus-like VLPs, and were used for immunological evaluation in healthy mice. The immunogenicity of these newly developed vaccine candidates demonstrated high titers of Feld1-specific Ab production; however, a comparably high immune response against carrier EMV was also observed. Antibody avidity tests revealed very specific Ab production (more than 50% specificity) for four out of the five vaccine candidates. Native Feld1 recognition and subclass-specific antibody tests suggested that the EMV-SZ18/17-Feld1 complex and chemically coupled EMV-Feld1 vaccines may possess characteristics for further development.

IL-4 Predicts the Efficacy of a Candidate Antioxycodone Vaccine and Alters Vaccine-Specific Antibody-Secreting Cell Proliferation in Mice.

Opioid use disorders (OUDs) are a public health concern in the United States and worldwide. Current medications for OUDs may trigger side effects and are often heavily regulated. A novel treatment strategy to be used alone or in combination with existing medications is active immunization with antiopioid vaccines, which stimulate production of opioid-specific Abs that bind to the target drug and prevent its distribution to the brain. Although antiopioid vaccines have shown promising preclinical efficacy, prior clinical evaluations of vaccines targeting stimulants indicate that efficacy is limited to a subset of subjects who achieve optimal Ab responses. We have previously reported that depletion of IL-4 with a mAb increased opioid-specific IgG2a and total IgG, and it increased the number of germinal centers and germinal center T follicular helper cells in response to antiopioid vaccines via type I IL-4 signaling. The current study further investigates the mechanisms associated with IL-4-mediated increases in efficacy and whether IL-4 depletion affects specific processes involved in germinal center formation, including affinity maturation, class switching, and plasma cell differentiation in mice. Additionally, results demonstrate that preimmunization production of IL-4 after ex vivo whole blood stimulation predicted invivo vaccine-induced Ab titers in outbred mice. Such mechanistic studies are critical for rational design of next-generation vaccine formulations, and they support the use of IL-4 as a predictive biomarker in ongoing OUD vaccine clinical studies.

Associations of HLA Polymorphisms with Anti-SARS-CoV-2 Spike and Neutralizing Antibody Titers in Japanese Rheumatoid Arthritis Patients Vaccinated with BNT162b2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes Coronavirus Disease 2019. Anti-SARS-CoV-2 spike (S) and neutralizing antibodies (Abs) are measured to evaluate the efficacy of vaccines. Human leukocyte antigen (HLA) may be associated with vaccine efficacy. Here, we investigated the association of HLA polymorphisms with the production of anti-SARS-CoV-2 S or neutralizing Abs in vaccinated rheumatoid arthritis (RA) patients in Japan. Genotyping of DRB1 and DQB1 was conducted in 87 Japanese RA patients vaccinated with BNT162b2. Associations of allele or haplotype carrier frequencies with anti-SARS-CoV-2 S or neutralizing Abs were examined. DRB1*12:01 was significantly positively associated with the production of S Ab (p = 0.0225, odds ratio [OR] 6.08, 95% confidence interval [CI] 1.32-28.03). The DQB1*03:01 allele carrier frequency tended to be higher in high responders of S Ab. Allele carrier frequencies of DRB1*15:01 (p = 0.0102, OR 9.26, 95% CI 1.65-52.01) and DQB1*06:02 (p = 0.0373, OR 7.00, 95% CI 1.18-41.36) were higher in responders of neutralizing Ab. Haplotype and two-locus analyses of DRB1 and DQB1 suggested that DRB1 alleles were the primary drivers of these associations. Logistic regression analysis showed associations of these alleles independent of clinical characteristics. Independent associations were found between HLA alleles and anti-SARS-CoV-2 Ab production by vaccinated RA patients.

WT1 Trio Peptide-Based Cancer Vaccine for Rare Cancers Expressing Shared Target WT1.

No standard treatment has been established for most rare cancers. Here, we report a clinical trial of a biweekly WT1 tri-peptide-based vaccine for recurrent or advanced rare cancers. Due to the insufficient number of patients available for a traditional clinical trial, the trial was designed for rare cancers expressing shared target molecule WT1. The recruitment criteria included WT1-expressing tumors as well as HLA-A*24:02 or 02:01. The primary endpoints were immunoglobulin G (IgG) antibody (Ab) production against the WT1-235 cytotoxic T lymphocyte (CTL) epitope and delayed-type hypersensitivity (DTH) skin reactions to targeted WT1 CTL epitopes. The secondary endpoints were safety and clinical efficacy. Forty-five patients received WT1 Trio, and 25 (55.6%) completed the 3-month protocol treatment. WT1-235 IgG Ab was positive in 88.0% of patients treated with WT1 Trio at 3 months, significantly higher than 62.5% of the weekly WT1-235 CTL peptide vaccine. The DTH positivity rate in WT1 Trio was 62.9%, which was not significantly different from 60.7% in the WT1-235 CTL peptide vaccine. The WT1 Trio safety was confirmed without severe treatment-related adverse events, except grade 3 myasthenia gravis-like symptoms observed in a patient with thymic cancer. Fifteen (33.3%) patients achieved stable disease after 3 months of treatment. In conclusion, the biweekly WT1 Trio vaccine containing the WT1-332 helper T lymphocyte peptide induced more robust immune responses targeting WT1 than the weekly WT1-235 CTL peptide vaccine. Therefore, WT1-targeted immunotherapy may be a potential therapeutic strategy for rare cancers.

Impact of Treatment with Anti-CD20 Monoclonal Antibody on the Production of Neutralizing Antibody Against Anti-SARS-CoV-2 Vaccination in Mature B-Cell Neoplasms.

Anti-CD20 monoclonal antibodies (MoAbs), rituximab (RIT), and obinutuzumab (OBZ) are the central components of immunochemotherapy for B-cell lymphoma (BCL). However, these agents potentially cause B-cell depletion, resulting in the impairment of antibody (Ab) production. During the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic, the optimal prediction of Ab response against anti-SARS-CoV-2 vaccination is critically important in patients with BCL treated by B-cell depletion therapeutics to prevent coronavirus disease 2019 (COVID-19). We investigated the effect of using RIT and/or OBZ on the Ab response in 131 patients with various types of BCL who received the second SARS-CoV-2 mRNA vaccine either after, during, or before immunochemotherapy containing B-cell-depleting moiety between June and November 2021 at seven institutes belonging to the Kyoto Clinical Hematology Study Group. The SARS-Cov-2 neutralizing Ab (nAb) was measured from 14 to 207 days after the second vaccination dose using the iFlash3000 automatic analyzer and the iFlash-2019-nCoV Nab kit. Among 86 patients who received the vaccine within 12 months after B-cell depletion therapy, 8 (9.3%) were seropositive. In 30 patients who received the vaccine after 12 months from B-cell depletion therapy, 22 (73%) were seropositive. In 15 patients who were subjected to B-cell depletion therapy after vaccination, 2 (13%) were seropositive. The multivariate analysis indicated that an interval of 12 months between B-cell depletion therapy and the subsequent vaccination was significantly associated with effective Ab production. Receiver operating characteristic curve analysis identified the optimal threshold period after anti-CD20 MoAb treatment, which determines the seropositivity against SARS-CoV-2, to be 342 days. The use of anti-CD20 MoAb within 12 months before vaccination is a critical risk for poor Ab response against anti-SARS-CoV-2 vaccination in patients with BCL.

Immune responses to inactivated COVID-19 vaccine were decreased in Chinese patients with chronic respiratory diseases.

Purpose: The effectiveness of inactivated vaccines against acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2), the causative agent of coronavirus disease 2019 (COVID-19), has become a global concern. Hence, the aim of this study was to evaluate vaccine safety and to assess immune responses in individuals with chronic respiratory disease (CRD) following a two-dose vaccination. Methods: The study cohort included 191 participants (112 adult CRD patients and 79 healthy controls [HCs]) at least 21 (range, 21-159) days after a second vaccination. Frequencies of memory B cells (MBCs) subsets and titers of SARS-CoV-2 neutralizing antibodies (NAbs) and anti-receptor binding domain (RBD) IgG antibodies (Abs) were analyzed. Results: As compared to the HCs, CRD patients had lower seropositivity rates and titers of both anti-RBD IgG Abs and NAbs, in addition to lower frequencies of RBD-specific MBCs (all, p < 0.05). At 3 months, CRD patients had lower seropositivity rates and titers of anti-RBD IgG Abs than the HCs (p < 0.05). For CoronaVac, the seropositivity rates of both Abs were lower in patients with old pulmonary tuberculosis than HCs. For BBIBP-CorV, the seropositivity rates of CoV-2 NAbs were lower in patients with chronic obstructive pulmonary disease than HCs (all, p < 0.05). Meanwhile, there was no significant difference in overall adverse events between the CRD patients and HCs. Univariate and multivariate analyses identified the time interval following a second vaccination as a risk factor for the production of anti-RBD IgG Abs and CoV-2 NAbs, while the CoronaVac had a positive effect on the titers of both Abs. Female was identified as a protective factor for CoV-2 NAb levels. Conclusion: Inactivated COVID-19 vaccines were safe and well tolerated by CRD patients but resulted in lower Ab responses and the frequencies of RBD-specific MBCs. Therefore, CRD patients should be prioritized for booster vaccinations.