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Simple Summary: Recent advancements in cancer research have proven immunotherapies to be a promising strategy for the treatment of hematological malignancies. The bispecific antibody (BsAb) format was developed to overcome the issues of monoclonal antibody-based therapies. T-cell engagers (TCEs) are BsAbs, which directly activate T-cells and their anti-tumor features, ultimately resulting in the lysis of the targeted tumor cells. In 2014, the FDA approved blinatumomab for treatment of acute lymphoblastic leukemia. As of November 2023, seven clinically approved TCE therapies are on the market. In this paper, we summarized the technical basis of the TCE technology, its application in hematology, and its current issues and prospects. Recent advancements in cancer immunotherapy have made directing the cellular immune response onto cancer cells a promising strategy for the treatment of hematological malignancies. The introduction of monoclonal antibody-based (mAbs) targeted therapy has significantly improved the prognosis for hematological patients. Facing the issues of mAb-based therapies, a novel bispecific antibody (BsAb) format was developed. T-cell engagers (TCEs) are BsAbs, which simultaneously target tumor-associated antigens on tumor cells and CD3 molecules present on T-cells. This mechanism allows for the direct activation of T-cells and their anti-tumor features, ultimately resulting in the lysis of tumor cells. In 2014, the FDA approved blinatumomab, a TCE directed to CD3 and CD19 for treatment of acute lymphoblastic leukemia. Since then, numerous TCEs have been developed, allowing for treating different hematological malignancies such as acute myeloid leukemia, multiple myeloma, and non-Hodgkin lymphoma and Hodgkin lymphoma. As of November 2023, seven clinically approved TCE therapies are on the market. TCE-based therapies still have their limitations; however, improving the properties of TCEs, as well as combining TCE-based therapies with other forms of treatment, give hope to find the cures for currently terminal diseases. In this paper, we summarized the technical basis of the TCE technology, its application in hematology, and its current issues and prospects. [ABSTRACT FROM AUTHOR]

Simple Summary: This article addresses the topic of primary immunodeficiencies, with particular emphasis on antibody deficiencies with near-normal immunoglobulin levels or hyperimmunoglobulinemia. This paper goes beyond genetics and emphasizes the importance of the immune system and particularly immune checkpoints and Epstein–Barr virus (EBV) reactivation in the context of these disorders. The article delves into the immune dysregulations occurring in the course of this type of disease and the potential role of EBV reactivation, which affects the clinical picture of patients and, in the future, may contribute to the development of cancer, especially those related to hematological malignancies. Disturbances observed in the immunopathogenesis of the presented diseases go beyond the accepted scheme, with the development of PID largely associated only with genetic disorders, and the article emphasizes that the regulation of immunity and virus reactivation also contributes to the progression of PID. This study delves into the intricate landscape of primary immunodeficiencies, with a particular focus on antibody deficiencies characterized by near-normal immunoglobulin levels or hyperimmunoglobulinemia. Contrary to the conventional focus on genetic dysregulation, these studies investigate the key roles of immune checkpoints, such as PD-1/PD-L1, CTLA-4/CD86, and CD200R/CD200, on selected subpopulations of T and B lymphocytes and their serum concentrations of soluble forms in patients recruited for the studies in healthy volunteers. In addition, the studies also show the role of Epstein–Barr virus (EBV) reactivation and interactions with tested pathways of immune checkpoints involved in the immunopathogenesis of this disease. By examining the context of antibody deficiencies, this study sheds light on the nuanced interplay of factors beyond genetics, particularly the immune dysregulations that occur in the course of this type of disease and the potential role of EBV reactivation, which affects the clinical presentation of patients and may contribute to the development of cancer in the future, especially related to hematological malignancies. [ABSTRACT FROM AUTHOR]

Mutations and the glycosylation of epitopes can convert immunogenic epitopes into non-immunogenic ones via natural selection or evolutionary pressure, thereby decreasing their sensitivity to neutralizing antibodies. Based on Thomas Francis's theory, memory B and T cells induced during primary infections or vaccination will freeze the new mutated epitopes specific to naïve B and T cells from the repertoire. On this basis, some researchers argue that the current vaccines derived from the previous strains of the SARS-CoV-2 virus do not increase immunity and may also prevent the immune response against new epitopes. However, evidence shows that even if the binding affinity is reduced, the previous antibodies or T cell receptors (TCRs) can still bind to this new epitope of the Beta, Gamma, and Delta variant if their concentration is high enough (from a booster injection) and neutralize the virus. This paper presents some convincing immunological reasons that may challenge this theory and argue for the continuation of universal vaccination to prevent further mutations of the SARS-CoV-2 virus. Simultaneously, the information presented can be used to develop vaccines that target novel epitopes or create new recombinant drugs that do not lose their effectiveness when the virus mutates. [ABSTRACT FROM AUTHOR]

The cells of acute myeloid leukemia are defined by clonal growth and heterogenous immunophenotypes. Chimeric antigen receptors (CARs) commonly recognize molecular targets by single-chain antibody fragments (scFvs) specific to a tumor-associated antigen. However, ScFvs may form aggregates, thus stimulating tonic CAR T-cell activation and reducing CAR T-cell functioning in vivo. Harnessing natural ligands as recognition parts of CARs, specific targeting of membrane receptors can be achieved. Previously, we presented ligand-based Flt3-CAR T-cells targeting the Flt3 receptor. The extracellular part of Flt3-CAR consisted of full-size Flt3Lg. Meanwhile, upon recognition, Flt3-CAR may potentially activate Flt3, triggering proliferative signaling in blast cells. Moreover, the long-lasting presence of Flt3Lg may lead to Flt3 downregulation. In this paper, we present mutated Flt3Lg-based Flt3m-CAR ('m'—for 'mutant') T-cells targeting Flt3. The extracellular part of Flt3m-CAR consists of full-length Flt3Lg-L27P. We have determined that ED50 for recombinant Flt3Lg-L27P produced in CHO cells is at least 10-fold higher than for the wild-type Flt3Lg. We show that the mutation in the recognizing domain of Flt3m-CAR did not affect the specificity of Flt3m-CAR T-cells when compared to Flt3-CAR T-cells. Flt3m-CAR T-cells combine the specificity of ligand–receptor recognition with reduced Flt3Lg-L27P bioactivity, leading to potentially safer immunotherapy. [ABSTRACT FROM AUTHOR]

Humans continue to be at risk from the Zika virus. Although there have been significant research advancements regarding Zika, the absence of a vaccine or approved treatment poses further challenges for healthcare providers. In this study, we developed a microparticulate Zika vaccine using an inactivated whole Zika virus as the antigen that can be administered pain-free via intranasal (IN) immunization. These microparticles (MP) were formulated using a double emulsion method developed by our lab. We explored a prime dose and two-booster-dose vaccination strategy using MPL-A® and Alhydrogel® as adjuvants to further stimulate the immune response. MPL-A® induces a Th1-mediated immune response and Alhydrogel® (alum) induces a Th2-mediated immune response. There was a high recovery yield of MPs, less than 5 µm in size, and particle charge of −19.42 ± 0.66 mV. IN immunization of Zika MP vaccine and the adjuvanted Zika MP vaccine showed a robust humoral response as indicated by several antibodies (IgA, IgM, and IgG) and several IgG subtypes (IgG1, IgG2a, and IgG3). Vaccine MP elicited a balance Th1- and Th2-mediated immune response. Immune organs, such as the spleen and lymph nodes, exhibited a significant increase in CD4+ helper and CD8+ cytotoxic T-cell cellular response in both vaccine groups. Zika MP vaccine and adjuvanted Zika MP vaccine displayed a robust memory response (CD27 and CD45R) in the spleen and lymph nodes. Adjuvanted vaccine-induced higher Zika-specific intracellular cytokines than the unadjuvanted vaccine. Our results suggest that more than one dose or multiple doses may be necessary to achieve necessary immunological responses. Compared to unvaccinated mice, the Zika vaccine MP and adjuvanted MP vaccine when administered via intranasal route demonstrated robust humoral, cellular, and memory responses. In this pre-clinical study, we established a pain-free microparticulate Zika vaccine that produced a significant immune response when administered intranasally. [ABSTRACT FROM AUTHOR]

V-set immunoglobulin domain-containing 4 (VSIG4) is a B7 family protein with known roles as a C3 fragment complement receptor involved in pathogen clearance and a negative regulator of T cell activation by an undetermined mechanism. VSIG4 expression is specific for tumor-associated and select tissue-resident macrophages. Increased expression of VSIG4 has been associated with worse survival in multiple cancer indications. Based upon computational analysis of transcript data across thousands of tumor and normal tissue samples, we hypothesized that VSIG4 has an important role in promoting M2-like immune suppressive macrophages and that targeting VSIG4 could relieve VSIG4-mediated macrophage suppression by repolarizing tumor-associated macrophages (TAMs) to an inflammatory phenotype. We have also observed a cancer-specific pattern of VSIG4 isoform distribution, implying a change in the functional regulation in cancer. Through a series of in vitro, in vivo, and ex vivo assays we demonstrate that anti-VSIG4 antibodies repolarize M2 macrophages and induce an immune response culminating in T cell activation. Anti-VSIG4 antibodies induce pro-inflammatory cytokines in M-CSF plus IL-10-driven human monocyte-derived M2c macrophages. Across patient-derived tumor samples from multiple tumor types, anti-VSIG4 treatment resulted in the upregulation of cytokines associated with TAM repolarization and T cell activation and chemokines involved in immune cell recruitment. VSIG4 blockade is also efficacious in a syngeneic mouse model as monotherapy as it enhances efficacy in combination with anti-PD-1, and the effect is dependent on the systemic availability of CD8+ T cells. Thus, VSIG4 represents a promising new target capable of triggering an anti-cancer response via multiple key immune mechanisms. [ABSTRACT FROM AUTHOR]

Before the emergence of SARS-CoV-1, MERS-CoV, and most recently, SARS-CoV-2, four other coronaviruses (the alpha coronaviruses NL63 and 229E and the beta coronaviruses OC43 and HKU1) had already been circulating in the human population. These circulating coronaviruses all cause mild respiratory illness during the winter seasons, and most people are already infected in early life. Could antibodies and/or T cells, especially against the beta coronaviruses, have offered some form of protection against (severe) COVID-19 caused by infection with SARS-CoV-2? Related is the question of whether survivors of SARS-CoV-1 or MERS-CoV would be relatively protected against SARS-CoV-2. More importantly, would humoral and cellular immunological memory generated during the SARS-CoV-2 pandemic, either by infection or vaccination, offer protection against future coronaviruses? Or rather than protection, could antibody-dependent enhancement have taken place, a mechanism by which circulating corona antibodies enhance the severity of COVID-19? Another related phenomenon, the original antigenic sin, would also predict that the effectiveness of the immune response to future coronaviruses would be impaired because of the reactivation of memory against irrelevant epitopes. The currently available evidence indicates that latter scenarios are highly unlikely and that especially cytotoxic memory T cells directed against conserved epitopes of human coronaviruses could at least offer partial protection against future coronaviruses. [ABSTRACT FROM AUTHOR]

The clinical development of an effective Chlamydia vaccine requires in-depth understanding of how well protective pre-clinical immune signatures translate to humans. Here, we report a comparative immunological characterization of CTH522/CAF®01 in female mice and humans. We find a range of immune signatures that translate from mouse to human, including a Th1/Th17 cytokine profile and antibody functionality. We identify vaccine-induced T cell epitopes, conserved among Chlamydia serovars, and previously found in infected individuals. Using the mouse model, we show that the common immune signature protected against ascending infection in mice, and vaccine induced antibodies could delay bacterial ascension to the oviduct, as well as development of pathology, in a T cell depleted mouse model. Finally, we demonstrate long-lasting immunity and protection of mice one year after vaccination. Based on the results obtained in the present study, we propose to further investigate CTH522/CAF®01 in a phase IIb study. Authors present a comparative immunological characterisation of Chlamydia vaccine, CTH522/CAF®01, in mice and humans. Findings suggest the mouse to be a good predictor of human immunity to the Chlamydia vaccine CTH522/CAF®01, and long-lasting protection in the mouse further supports the development of this promising vaccine candidate. [ABSTRACT FROM AUTHOR]

Human cytomegalovirus (HCMV) remains the most common congenital infection and infectious complication in immunocompromised patients. The most successful HCMV vaccine to date, an HCMV glycoprotein B (gB) subunit vaccine adjuvanted with MF59, achieved 50% efficacy against primary HCMV infection. A previous study demonstrated that gB/MF59 vaccinees were less frequently infected with HCMV gB genotype strains most similar to the vaccine strain than strains encoding genetically distinct gB genotypes, suggesting strain-specific immunity accounted for the limited efficacy. To determine whether vaccination with multiple HCMV gB genotypes could increase the breadth of anti-HCMV gB humoral and cellular responses, we immunized 18 female rabbits with monovalent (gB-1), bivalent (gB-1+gB-3), or pentavalent (gB-1+gB-2+gB-3+gB-4+gB-5) gB lipid nanoparticle-encapsulated nucleoside-modified RNA (mRNA–LNP) vaccines. The multivalent vaccine groups did not demonstrate a higher magnitude or breadth of the IgG response to the gB ectodomain or cell-associated gB compared to that of the monovalent vaccine. Also, the multivalent vaccines did not show an increase in the breadth of neutralization activity and antibody-dependent cellular phagocytosis against HCMV strains encoding distinct gB genotypes. Interestingly, peripheral blood mononuclear cell-derived gB-2-specific T-cell responses elicited by multivalent vaccines were of a higher magnitude compared to that of monovalent vaccinated animals against a vaccine-mismatched gB genotype at peak immunogenicity. Yet, no statistical differences were observed in T cell response against gB-3 and gB-5 variable regions among the three vaccine groups. Our data suggests that the inclusion of multivalent gB antigens is not an effective strategy to increase the breadth of anti-HCMV gB antibody and T cell responses. Understanding how to increase the HCMV vaccine protection breadth will be essential to improve the vaccine efficacy. [ABSTRACT FROM AUTHOR]

The regulatory T (Treg) cells constitute a functionally defined subpopulation of T cells that modulate the immune system and maintain immune tolerance through suppression of the development of autoimmune responses to self-antigens and allergic reactions to external antigens. Reduction in the number or function of Treg cells has been suggested as a key immune abnormality underlying the development of autoimmune and allergic diseases. In vitro studies have demonstrated that purified polyvalent immunoglobulin G (IgG) from multiple healthy blood donors can exert immunomodulatory effects on Treg cells. Incubation of polyvalent human IgG with purified CD4+CD25high T cells increased the intracellular expression of interleukin (IL)-10. Intravenous administration of polyvalent human IgG induced significant expansions of CD4+ Foxp3+ Treg cells and clinical improvements in patients with autoimmune diseases. In human clinical trials, intramuscular administration of autologous total IgG significantly increased the percentage of IL-10-producing CD4+ Treg cells in the peripheral blood of healthy subjects and provided significant clinical improvements in patients with atopic dermatitis. These results suggest a clinical usefulness of polyvalent IgG-induced activation of Treg cells in human subjects. This review proposes a new hypothesis for immune tolerance mechanism by integrating the pre-existing “idiotypic network theory” and “Treg cell theory” into an “anti-idiotypic Treg cell theory.” Based on this hypothesis, an “active anti-idiotypic therapy” for allergic and autoimmune diseases using autologous polyvalent IgG (as immunizing antigens) is suggested as follows: (1) Intramuscular or subcutaneous administration of autologous polyvalent IgG produces numerous immunogenic peptides derived from idiotypes of autologous IgG through processing of dendritic cells, and these peptides activate anti-idiotypic Treg cells in the same subject. (2) Activated anti-idiotypic Treg cells secrete IL-10 and suppress Th2 cell response to allergens and autoimmune T cell response to selfantigens. (3) These events can induce a long-term clinical improvements in patients with allergic and autoimmune diseases. Further studies are needed to evaluate the detailed molecular mechanism underlying polyvalent IgG-induced Treg cell activation and the clinical usefulness of this immunomodulatory therapy for autoimmune and allergic diseases. [ABSTRACT FROM AUTHOR]

The conformational landscapes of peptide/human leucocyte antigen (pHLA) protein complexes encompassing tumor neoantigens provide a rationale for target selection towards autologous T cell, vaccine, and antibody-based therapeutic modalities. Here, using complementary biophysical and computational methods, we characterize recurrent RAS55-64 Q61 neoepitopes presented by the common HLA-A*01:01 allotype. We integrate sparse NMR restraints with Rosetta docking to determine the solution structure of NRASQ61K/HLA-A*01:01, which enables modeling of other common RAS55-64 neoepitopes. Hydrogen/deuterium exchange mass spectrometry experiments alongside molecular dynamics simulations reveal differences in solvent accessibility and conformational plasticity across a panel of common Q61 neoepitopes that are relevant for recognition by immunoreceptors. Finally, we predict binding and provide structural models of NRASQ61K antigens spanning the entire HLA allelic landscape, together with in vitro validation for HLA-A*01:191, HLA-B*15:01, and HLA-C*08:02. Our work provides a basis to delineate the solution surface features and immunogenicity of clinically relevant neoepitope/HLA targets for cancer therapy. The authors use an integrative structural biology approach to elucidate molecular features of the RAS Q61 family of public tumor antigens. This information can be used to develop targeted therapeutics to combat a range of cancers. [ABSTRACT FROM AUTHOR]

Background: Classical swine fever and porcine reproductive and respiratory syndrome have seriously affected the development of the swine breeding industry in China. Vaccine immunization remains the main way to prevent these infections. The aim of this study was to establish an optimized protocol for vaccine immunization against classical swine fever virus (CSFV) and porcine reproductive and respiratory syndrome virus (PRRSV). Methods: Blood samples were collected from the anterior vena cava of pigs after immunization, and blood indices, secreted levels of specific antibodies and neutralizing antibodies associated with humoral immunity, the proliferation capacity of T lymphocytes as a measure of cellular immunity, and secreted levels of IFN-γ and TNF-α were determined. Results: The results showed that simultaneous immunization against CSFV and PRRSV infections induced strong and specific humoral and T-cellular immune responses, high levels of cytokine IFN-γ secretion and delayed secretion of cytokine TNF-α. Moreover, significantly higher lymphocyte percentages and red blood cell and leukocyte counts were found in the group simultaneously immunized against CSFV and PRRSV. However, no statistically significant differences were observed in hemoglobin values, neutrophil counts, and median cell percentages among the S + PRRS, PRRS-S, and S-PRRS groups. Conclusion: This study demonstrated that simultaneous immunization against CSFV and PRRSV had the advantages of inducing a rapid, enhanced, and long-lasting immune response. These findings provide a theoretical basis for the establishment of a reasonable and optimized vaccine immunization protocol against CSFV and PRRSV in combination with a variety of other vaccine inoculations. [ABSTRACT FROM AUTHOR]

Respiratory syncytial virus (RSV) causes a high disease burden in older adults. An effective vaccine for this RSV-primed population may need to boost/elicit robust RSV-neutralizing antibody responses and recall/induce RSV-specific T cell responses. To inform the selection of the vaccine formulation for older adults, RSVPreF3 (RSV fusion glycoprotein engineered to maintain the prefusion conformation) with/without AS01 adjuvant was evaluated in mice and bovine RSV infection-primed cattle. In mice, RSVPreF3/AS01 elicited robust RSV-A/B-specific neutralization titers and RSV F-specific polyfunctional CD4+ T cell responses exceeding those induced by non-adjuvanted RSVPreF3. In primed bovines, RSVPreF3/AS01 tended to induce higher pre-/post-vaccination fold-increases in RSV-A/B-specific neutralization titers relative to non-adjuvanted and Alum-adjuvanted RSVPreF3 formulations, and elicited higher RSV F-specific CD4+ T cell frequencies relative to the non-adjuvanted vaccine. Though AS01 adjuvanticity varied by animal species and priming status, RSVPreF3/AS01 elicited/boosted RSV-A/B-specific neutralization titers and RSV F-specific CD4+ T cell responses in both animal models, which supported its further clinical evaluation as prophylactic candidate vaccine for older adults. [ABSTRACT FROM AUTHOR]

HIV affects more than 38 million people worldwide. Although HIV can be effectively treated by lifelong combination antiretroviral therapy, only a handful of patients have been cured. Therapeutic vaccines that induce robust de novo immune responses targeting HIV proteins and latent reservoirs will likely be integral for functional HIV cure. Our study shows that immunization of naïve rhesus macaques with arenavirus-derived vaccine vectors encoding simian immunodeficiency virus (SIVSME543 Gag, Env, and Pol) immunogens is safe, immunogenic, and efficacious. Immunization induced robust SIV-specific CD8+ and CD4+ T-cell responses with expanded cellular breadth, polyfunctionality, and Env-binding antibodies with antibody-dependent cellular cytotoxicity. Vaccinated animals had significant reductions in median SIV viral load (1.45-log10 copies/mL) after SIVMAC251 challenge compared with placebo. Peak viral control correlated with the breadth of Gag-specific T cells and tier 1 neutralizing antibodies. These results support clinical investigation of arenavirus-based vectors as a central component of therapeutic vaccination for HIV cure. [ABSTRACT FROM AUTHOR]

Accurate structural information plays a crucial role in comprehending biological processes and designing drugs. Indeed, the remarkable precision of the AlphaFold2 has facilitated significant advancements in predicting molecular structures, encompassing antibodies and antigens. This breakthrough has paved the way for rational drug design, ushering in new possibilities in the field of pharmaceutical development. Within this study, performing analysis and humanization guided by the structures predicted by AlphaFold2. Notably, the resulting humanized antibody, h3D5-hIgG1, demonstrated exceptional binding affinity to the PD-L1 protein. The KD value of parental antibody 3D5-hIgG1 was increased by nearly 7 times after humanization. Both h3D5-hIgG1 and 3D5-hIgG1 bound to cells expressing human PD-L1 with EC50 values of 5.13 and 9.92nM, respectively. Humanization resulted in a twofold increase in the binding capacity of the antibody, with h3D5-hIgG1 exhibiting superior performance compared to the parental antibody 3D5-hIgG1. Furthermore, h3D5-hIgG1 promoted cytokine secretion of T cells, and significantly suppressed MC38-hPD-L1 tumor growth. This study highlights the potential for artificial intelligence-assisted drug development, which is poised to become a prominent trend in the future. [ABSTRACT FROM AUTHOR]

Combinational immunotherapy of dendritic cell (DC) vaccines and anti‐programmed cell death protein 1 antibodies (aPD1) has been regarded as a promising strategy for cancer treatment because it not only induces tumor‐specific T cell immune responses, but also prevents failure of T cell functions by the immune suppressive milieu of tumors. Microneedles have emerged as an innovative platform for efficient transdermal immunotherapies. However, co‐delivery of DC vaccines and aPD1 via microneedles has not been studied since conventional microneedle platforms are unsuitable for fragile therapeutics like living cells and antibodies. This study employs our newly invented cryomicroneedles (cryoMNs) to co‐deliver DC vaccines and aPD1 for the combinational immunotherapy. CryoMNs are fabricated by stepwise cryogenic micromoulding of cryogenic medium with pre‐suspended DCs and aPD1, which are further integrated with a homemade handle for convenient application. The viability of DCs in cryoMNs remains above 85%. CryoMNs are mechanically strong enough to insert into porcine and mouse skin, successfully releasing DCs and aPD1 inside skin tissue after melting. Co‐delivery of ovalbumin (OVA)‐pulsed DCs (OVA‐DCs) and aPD1 via cryoMNs induced higher antigen‐specific cellular immune responses compared with the mono‐delivery of OVA‐DCs or aPD1. Finally, administration with cryoMNs co‐encapsulated with OVA‐DCs and aPD1 increases the infiltration of effector T cells in the tumor, resulting in stronger anti‐tumor therapeutic efficacy in both prophylactic and therapeutic melanoma models compared with administration with cryoMNs loaded with OVA‐DCs or aPD1. This study demonstrates the great potential of cryoMNs as a co‐delivery system of therapeutic cells and biomacromolecules for combinational therapies. [ABSTRACT FROM AUTHOR]

Among the unknowns in decoding the pathogenesis of SARS-CoV-2 persistent symptoms in Long Covid is whether there is a contributory role of abnormal immunity during acute infection. It has been proposed that Long Covid is a consequence of either an excessive or inadequate initial immune response. Here, we analyze SARS-CoV-2 humoral and cellular immunity in 86 healthcare workers with laboratory confirmed mild or asymptomatic SARS-CoV-2 infection during the first wave. Symptom questionnaires allow stratification into those with persistent symptoms and those without for comparison. During the period up to 18-weeks post-infection, we observe no difference in antibody responses to spike RBD or nucleoprotein, virus neutralization, or T cell responses. Also, there is no difference in the profile of antibody waning. Analysis at 1-year, after two vaccine doses, comparing those with persistent symptoms to those without, again shows similar SARS-CoV-2 immunity. Thus, quantitative differences in these measured parameters of SARS-CoV-2 adaptive immunity following mild or asymptomatic acute infection are unlikely to have contributed to Long Covid causality. ClinicalTrials.gov (NCT04318314). Authors utilise a cohort of healthcare workers, infected during the first wave of the SARS-CoV-2 pandemic, to assess symptom persistence and humoral and cellular immunity. [ABSTRACT FROM AUTHOR]

The intestinal epithelial receptor Guanylyl Cyclase C (GUCY2C) is a tumor-associated cell surface antigen expressed across gastrointestinal malignancies that can serve as an efficacious target for colorectal cancer immunotherapy. Here, we describe a yeast surface-display approach combined with an orthogonal peptide-based mapping strategy to identify the GUCY2C binding epitope of a novel anti-GUCY2CxCD3 bispecific antibody (BsAb) that recently advanced into the clinic for the treatment of cancer. The target epitope was localized to the N-terminal helix H2 of human GUCY2C, which enabled the determination of the crystal structure of the minimal GUCY2C epitope in complex with the anti-GUCY2C antibody domain. To understand if this minimal epitope covers the entire antibody binding region and to investigate the impact of epitope position on the antibody's activity, we further determined the structure of this interaction in the context of the full-length extracellular domain (ECD) of GUCY2C. We found that this epitope is positioned on the protruding membrane-distal helical region of GUCY2C and that its specific location on the surface of GUCY2C dictates the close spatial proximity of the two antigen arms in a diabody arrangement essential to the tumor killing activity of GUCY2CxCD3 BsAb. [ABSTRACT FROM AUTHOR]

One of the most preferable characteristics for a COVID-19 vaccine candidate is the ability to reduce transmission and infection of SARS-CoV-2, in addition to disease prevention. Unlike intramuscular vaccines, intranasal COVID-19 vaccines may offer this by generating mucosal immunity. In this open-label, randomised, multicentre, phase 3 clinical trial (CTRI/2022/02/40065; ClinicalTrials.gov: NCT05522335), healthy adults were randomised to receive two doses, 28 days apart, of either intranasal adenoviral vectored SARS-CoV-2 vaccine (BBV154) or licensed intramuscular vaccine, Covaxin®. Between April 16 and June 4, 2022, we enrolled 3160 subjects of whom, 2971 received 2 doses of BBV154 and 161 received Covaxin. On Day 42, 14 days after the second dose, BBV154 induced significant serum neutralization antibody titers against the ancestral (Wuhan) virus, which met the pre-defined superiority criterion for BBV154 over Covaxin®. Further, both vaccines showed cross protection against Omicron BA.5 variant. Salivary IgA titers were found to be higher in BBV154. In addition, extensive evaluation of T cell immunity revealed comparable responses in both cohorts due to prior infection. However, BBV154 showed significantly more ancestral specific IgA-secreting plasmablasts, post vaccination, whereas Covaxin recipients showed significant Omicron specific IgA-secreting plasmablasts only at day 42. Both vaccines were well tolerated. Overall reported solicited reactions were 6.9% and 25.5% and unsolicited reactions were 1.2% and 3.1% in BBV154 and Covaxin® participants respectively. [ABSTRACT FROM AUTHOR]

Background: Infections with Plasmodium ovale are widely distributed but rarely investigated, and the resulting burden of disease has been underestimated. Plasmodium ovale curtisi Duffy binding protein domain region II (PocDBP-RII) is an essential ligand for reticulocyte recognition and host cell invasion by P. ovale curtisi. However, the genomic variation, antigenicity and immunogenicity of PocDBP-RII remain major knowledge gaps. Methods: A total of 93 P. ovale curtisi samples were collected from migrant workers who returned to China from 17 countries in Africa between 2012 and 2016. The genetic polymorphism, natural selection and copy number variation (CNV) were investigated by sequencing and real-time PCR. The antigenicity and immunogenicity of the recombinant PocDBP-RII (rPocDBP-RII) protein were further examined, and the humoral and cellular responses of immunized mice were assessed using protein microarrays and flow cytometry. Results: Efficiently expressed and purified rPocDBP-RII (39 kDa) was successfully used as an antigen for immunization in mice. The haplotype diversity (Hd) of PocDBP-RII gene was 0.105, and the nucleotide diversity index (π) was 0.00011. No increased copy number was found among the collected isolates of P. ovale curtisi. Furthermore, rPocDBP-RII induced persistent antigen-specific antibody production with a serum IgG antibody titer of 1: 16,000. IFN-γ-producing T cells, rather than IL-10-producing cells, were activated in response to the stimulation of rPocDBP-RII. Compared to PBS-immunized mice (negative control), there was a higher percentage of CD4+CD44highCD62L− T cells (effector memory T cells) and CD8+CD44highCD62L+ T cells (central memory T cells) in rPocDBP-RII‑immunized mice. Conclusions: PocDBP-RII sequences were highly conserved in clinical isolates of P. ovale curtisi. rPocDBP-RII protein could mediate protective blood-stage immunity through IFN-γ-producing CD4+ and CD8+ T cells and memory T cells, in addition to inducing specific antibodies. Our results suggested that rPocDBP-RII protein has potential as a vaccine candidate to provide assessment and guidance for malaria control and elimination activities. [ABSTRACT FROM AUTHOR]

Anti-CTLA-4 antibodies have successfully elicited durable tumor regression in the clinic; however, long-term benefit is limited to a subset of patients for select cancer indications. The incomplete understanding of their mechanism of action has hindered efforts at improvement, with conflicting hypotheses proposing either antagonism of the CTLA-4: B7 axis or Fc effector-mediated regulatory T cell (Treg) depletion governing efficacy. Here, we report the engineering of a nonantagonistic CTLA-4 binding domain (b1s1e2) that depletes intratumoral Tregs as an Fc fusion. Comparison of b1s1e2-Fc to 9d9, an antagonistic anti-CTLA-4 antibody, allowed for interrogation of the separate contributions of CTLA-4 antagonism and Treg depletion to efficacy. Despite equivalent levels of intratumoral Treg depletion, 9d9 achieved more long-term cures than b1s1e2-Fc in MC38 tumors, demonstrating that CTLA-4 antagonism provided additional survival benefit. Consistent with prior reports that CTLA-4 antagonism enhances priming, treatment with 9d9, but not b1s1e2-Fc, increased the percentage of activated T cells in the tumor-draining lymph node (tdLN). Treg depletion with either construct was restricted to the tumor due to insufficient surface CTLA-4 expression on Tregs in other compartments. Through intratumoral administration of diphtheria toxin in Foxp3-DTR mice, we show that depletion of both intratumoral and nodal Tregs provided even greater survival benefit than 9d9, consistent with Treg-driven restraint of priming in the tdLN. Our data demonstrate that anti-CTLA-4 therapies require both CTLA-4 antagonism and intratumoral Treg depletion for maximum efficacy--but that potential future therapies also capable of depleting nodal Tregs could show efficacy in the absence of CTLA-4 antagonism. [ABSTRACT FROM AUTHOR]

Cytomegalovirus (CMV) is known to alter circulating effector memory or re-expressing CD45RA+ (TemRA) T-cell numbers, but whether Epstein–Barr virus (EBV) does the same or this is amplified during a CMV and EBV co-infection is unclear. Immune cell numbers in blood of children and young, middle-aged, and senior adults (n = 336) were determined with flow cytometry, and additional multivariate linear regression, intra-group correlation, and cluster analyses were performed. Compared to non-infected controls, CMV-seropositive individuals from all age groups had more immune cell variance, and CMV+ EBV− senior adults had more late-differentiated CD4+ and CD8+ TemRA and CD4+ effector memory T-cells. EBV-seropositive children and young adults had a more equal immune cell composition than non-infected controls, and CMV− EBV+ senior adults had more intermediate/late-differentiated CD4+ TemRA and effector memory T-cells than non-infected controls. CMV and EBV co-infected young and middle-aged adults with an elevated BMI and anti-CMV antibody levels had a similar immune cell composition as senior adults, and CMV+ EBV+ middle-aged adults had more late-differentiated CD8+ TemRA, effector memory, and HLA-DR+ CD38− T-cells than CMV+ EBV− controls. This study identified changes in T-cell numbers in CMV- or EBV-seropositive individuals and that some CMV and EBV co-infected young and middle-aged adults had an aging-related T-cell phenotype. [ABSTRACT FROM AUTHOR]

Objective: The kinetics of immune responses to various SARS-CoV-2 vaccines in cancer patients were investigated. Methods: In total, 57 cancer patients who received BNT162b2-RNA or BBIBP-CorV vaccines were enrolled. Cellular and humoral immunity were assessed at three-time points, before the first vaccine dose and 14–21 days after the first and second doses. Chemiluminescent microparticle immunoassay was used to evaluate SARS-CoV-2 anti-spike IgG response, and QuantiFERON® SARS-CoV-2 kit assessed T-cell response. Results: Data showed that cancer patients' CD4+ and CD8+ T cell-median IFN-γ secretion of SARS-CoV-2 antigens increased after the first and second vaccine doses (p = 0.027 and p = 0.042). BNT162b2 vaccinees had significantly higher IFN-γ levels to CD4+ and CD8+ T cell epitopes than BBIBP-CorV vaccinees (p = 0.028). There was a positive correlation between IgG antibody titer and T cell response regardless of vaccine type (p < 0.05). Conclusions: This study is one of the first to investigate cellular and humoral immune responses to SARS-CoV-2 immunization in cancer patients on active therapy after each vaccine dose. COVID-19 immunizations helped cancer patients develop an effective immune response. Understanding the cellular and humoral immune response to COVID-19 in cancer patients undergoing active treatment is necessary to improve vaccines and avoid future SARS pandemics. [ABSTRACT FROM AUTHOR]

Agents that can simultaneously activate latent HIV, increase immune activation and enhance the killing of latently-infected cells represent promising approaches for HIV cure. Here, we develop and evaluate a trispecific antibody (Ab), N6/αCD3-αCD28, that targets three independent proteins: (1) the HIV envelope via the broadly reactive CD4-binding site Ab, N6; (2) the T cell antigen CD3; and (3) the co-stimulatory molecule CD28. We find that the trispecific significantly increases antigen-specific T-cell activation and cytokine release in both CD4+ and CD8+ T cells. Co-culturing CD4+ with autologous CD8+ T cells from ART-suppressed HIV+ donors with N6/αCD3-αCD28, results in activation of latently-infected cells and their elimination by activated CD8+ T cells. This trispecific antibody mediates CD4+ and CD8+ T-cell activation in non-human primates and is well tolerated in vivo. This HIV-directed antibody therefore merits further development as a potential intervention for the eradication of latent HIV infection. One of the main hurdles to curing HIV infection are viral reservoirs. Here, the authors develop a trispecific antibody and demonstrate its ability to simultaneously activate and target latently HIV−1 infected cells for elimination by T cells as an alternative strategy for HIV cure. [ABSTRACT FROM AUTHOR]

Bi-specific T-cell engager antibodies (BiTEs) are synthetic fusion molecules that combine multiple antibody-binding domains to induce active contact between T-cells and antigen expressing cells in the body. Blinatumomab, a CD19-CD3 BiTE is now a widely used therapy for relapsed B-cell malignancies, and similar BiTE therapeutics have shown promise for treating various other forms of cancer. The current process for new BiTE development is time consuming and costly, requiring characterization of the individual antigen binding domains, followed by bi-specific design, protein production, purification, and eventually functional screening. Here, we sought to establish a more cost-efficient approach for generating novel BiTE sequences and assessing bioactivity through a function first approach without purification. We generate a plasmid with a bi-modular structure to allow high-throughput exchange of either binding arm, enabling rapid screening of novel tumour-targeting single chain variable (scFv) domains in combination with the well-characterized OKT3 scFv CD3-targeting domain. We also demonstrate two systems for high throughput functional screening of BiTE proteins based on Jurkat T cells (referred to as BiTE-J). Using BiTE-J we evaluate four EGFRvIII-scFv sequenced in BiTE format, identifying two constructs with superior activity for redirecting T-cells against the EGFRvIII-tumour specific antigen. We also confirm activity in primary T cells, where novel EGFRvIII-BiTEs induced T cell activation and antigen selective tumor killing. We finally demonstrate similar exchange the CD3-interacting element of our bi-modular plasmid. By testing several novel CD3-targeting scFv elements for activity in EGFRvIII-targeted BiTEs, we were able to identify highly active BiTE molecules with desirable functional activity for downstream development. In summary, BiTE-J presents a low cost, high-throughput method for the rapid assessment of novel BiTE molecules without the need for purification and quantification. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 has resulted in high levels of morbidity and mortality world-wide, and severe complications can occur in older populations. Humoral immunity induced by authorized vaccines wanes within 6 months, and frequent boosts may only offer transient protection. GRT-R910 is an investigational self-amplifying mRNA (samRNA)-based SARS-CoV-2 vaccine delivering full-length Spike and selected conserved non-Spike T cell epitopes. This study reports interim analyses for a phase I open-label dose-escalation trial evaluating GRT-R910 in previously vaccinated healthy older adults (NCT05148962). Primary endpoints of safety and tolerability were assessed. Most solicited local and systemic adverse events (AEs) following GRT-R910 dosing were mild to moderate and transient, and no treatment-related serious AEs were observed. The secondary endpoint of immunogenicity was assessed via IgG binding assays, neutralization assays, interferon-gamma ELISpot, and intracellular cytokine staining. Neutralizing antibody titers against ancestral Spike and variants of concern were boosted or induced by GRT-R910 and, contrasting to authorized vaccines, persisted through at least 6 months after the booster dose. GRT-R910 increased and/or broadened functional Spike-specific T cell responses and primed functional T cell responses to conserved non-Spike epitopes. This study is limited due to small sample size, and additional data from ongoing studies will be required to corroborate these interim findings. Palmer et al. present interim findings from their clinical trial; they describe the reactogenicity and immunogenicity results of a self-amplifying mRNA SARS-CoV-2 booster vaccine within an older population (≥60 years of age). [ABSTRACT FROM AUTHOR]

With recent advances in immune checkpoint inhibitors (ICIs), immunotherapy has become the standard treatment for various malignant tumors. Their indications and dosages have been determined empirically, taking individually conducted clinical trials into consideration, but without a standard method to evaluate them. Here we establish an advanced imaging system to visualize human PD-1 microclusters, in which a minimal T cell receptor (TCR) signaling unit co-localizes with the inhibitory co-receptor PD-1 in vitro. In these microclusters PD-1 dephosphorylates both the TCR/CD3 complex and its downstream signaling molecules via the recruitment of a phosphatase, SHP2, upon stimulation with the ligand hPD-L1. In this system, blocking antibodies for hPD-1-hPD-L1 binding inhibits hPD-1 microcluster formation, and each therapeutic antibody (pembrolizumab, nivolumab, durvalumab and atezolizumab) is characterized by a proprietary optimal concentration and combinatorial efficiency enhancement. We propose that our imaging system could digitally evaluate PD-1-mediated T cell suppression to evaluate their clinical usefulness and to develop the most suitable combinations among ICIs or between ICIs and conventional cancer treatments. Immune checkpoint inhibitors are now routinely used in cancer therapy, however, the dosage and integration into conventional cancer therapy is determined via empirical experience rather than mechanistic rationale. Here authors establish an advanced single-molecule imaging method, by with which they are directly monitoring and evaluating the effect of immune checkpoint inhibitors on T cell signaling. [ABSTRACT FROM AUTHOR]

Simple Summary: Visceral leishmaniasis is a tropical neglected disease caused by infection with Leishmania parasites. There are no human vaccines and the current drug treatments for infected patients are hampered by toxicity and the development of resistance. In our study, we have used an immunotherapeutic approach, which combines a vaccine and drug, to treat mice infected with the parasite. This involved injecting a candidate Leishmania vaccine ChimT with the adjuvant monophosphoryl lipid A (MPLA, which enhances the immune response) and a treatment drug amphotericin B (AmpB). We found that the combined immunotherapy of ChimT/MPLA/AmpB significantly reduced the number of parasites within infected internal organs of the animals and increased the production of protective antibodies. Organ toxicity was also lower with the ChimT/MPLA/AmpB immunotherapy, suggesting that the inclusion of the vaccine and adjuvant reduced the inherent toxicity of the drug. In addition, the ChimT vaccine stimulated in vitro mouse white cells to kill three different Leishmania parasites that had invaded the cells. We conclude that the combination of ChimT/MPLA/AmpB could be considered for further studies as an immunotherapy for Leishmania infection. Visceral leishmaniasis (VL) in the Americas is a chronic systemic disease caused by infection with Leishmania infantum parasites. The toxicity of antileishmanial drugs, long treatment course and limited efficacy are significant concerns that hamper adequate treatment against the disease. Studies have shown the promise of an immunotherapeutics approach, combining antileishmanial drugs to reduce the parasitism and vaccine immunogens to activate the host immune system. In the current study, we developed an immunotherapy using a recombinant T cell epitope-based chimeric protein, ChimT, previously shown to be protective against Leishmania infantum, with the adjuvant monophosphoryl lipid A (MPLA) and amphotericin B (AmpB) as the antileishmanial drug. BALB/c mice were infected with L. infantum stationary promastigotes and later they received saline or were treated with AmpB, MPLA, ChimT/Amp, ChimT/MPLA or ChimT/MPLA/AmpB. The combination of ChimT/MPLA/AmpB significantly reduced the parasite load in mouse organs (p < 0.05) and induced a Th1-type immune response, which was characterized by higher ratios of anti-ChimT and anti-parasite IgG2a:IgG1 antibodies, increased IFN-γ mRNA and IFN-γ and IL-12 cytokines and accompanied by lower levels of IL-4 and IL-10 cytokines, when compared to other treatments and controls (all p < 0.05). Organ toxicity was also lower with the ChimT/MPLA/AmpB immunotherapy, suggesting that the inclusion of the vaccine and adjuvant ameliorated the toxicity of AmpB to some degree. In addition, the ChimT vaccine alone stimulated in vitro murine macrophages to significantly kill three different internalized species of Leishmania parasites and to produce Th1-type cytokines into the culture supernatants. To conclude, our data suggest that the combination of ChimT/MPLA/AmpB could be considered for further studies as an immunotherapy for L. infantum infection. [ABSTRACT FROM AUTHOR]

Herd immunity achieved through mass vaccination is an effective approach to prevent contagious diseases. Nonetheless, emerging SARS-CoV-2 variants with frequent mutations largely evaded humoral immunity induced by Spike-based COVID-19 vaccines. Herein, we develop a lipid nanoparticle (LNP)-formulated mRNA-based T-cell-inducing antigen, which targeted three SARS-CoV-2 proteome regions that enriched human HLA-I epitopes (HLA-EPs). Immunization of HLA-EPs induces potent cellular responses to prevent SARS-CoV-2 infection in humanized HLA-A*02:01/DR1 and HLA-A*11:01/DR1 transgenic mice. Of note, the sequences of HLA-EPs are highly conserved among SARS-CoV-2 variants of concern. In humanized HLA-transgenic mice and female rhesus macaques, dual immunization with the LNP-formulated mRNAs encoding HLA-EPs and the receptor-binding domain of the SARS-CoV-2 B.1.351 variant (RBDbeta) is more efficacious in preventing infection of SARS-CoV-2 Beta and Omicron BA.1 variants than single immunization of LNP-RBDbeta. This study demonstrates the necessity to strengthen the vaccine effectiveness by comprehensively stimulating both humoral and cellular responses, thereby offering insight for optimizing the design of COVID-19 vaccines. The authors show that an mRNA-based T-cell-inducing antigen combined with the receptor-binding domain of the SARS-CoV-2 spike protein strengthens the COVID19 vaccine against SARS-CoV-2 variants, suggesting improved vaccine designs that comprehensively stimulate both humoral and cellular responses. [ABSTRACT FROM AUTHOR]

Mucosal immunity plays a significant role in the first-line defense against viruses transmitted and infected through the respiratory system, such as SARS-CoV-2. However, the lack of effective and safe adjuvants currently limits the development of COVID-19 mucosal vaccines. In the current study, we prepare an intranasal vaccine containing cationic crosslinked carbon dots (CCD) and a SARS-CoV-2 antigen, RBD-HR with spontaneous antigen particlization. Intranasal immunization with CCD/RBD-HR induces high levels of antibodies with broad-spectrum neutralization against authentic viruses/pseudoviruses of Omicron-included variants and protects immunized female BALB/c mice from Omicron infection. Despite strong systemic cellular immune response stimulation, the intranasal CCD/RBD-HR vaccine also induces potent mucosal immunity as determined by the generation of tissue-resident T cells in the lungs and airway. Moreover, CCD/RBD-HR not only activates professional antigen-presenting cells (APCs), dendritic cells, but also effectively targets nasal epithelial cells, promotes antigen binding via sialic acid, and surprisingly provokes the antigen-presenting of nasal epithelial cells. We demonstrate that CCD is a promising intranasal vaccine adjuvant for provoking strong mucosal immunity and might be a candidate adjuvant for intranasal vaccine development for many types of infectious diseases, including COVID-19. Lei et al. show that intranasal immunisation with their vaccine candidate is able to broadly protect mice from SARS-CoV-2 infection, including Omicron variants. [ABSTRACT FROM AUTHOR]

Kidney transplanted recipients (KTR) are at high risk of severe SARS-CoV-2 infection due to immunosuppressive therapy. Although several studies reported antibody production in KTR after vaccination, data related to immunity to the Omicron (B.1.1.529) variant are sparse. Herein, we analyzed anti-SARS-CoV-2 immune response in seven KTR and eight healthy controls after the second and third dose of the mRNA vaccine (BNT162b2). A significant increase in neutralizing antibody (nAb) titers were detected against pseudoviruses expressing the Wuhan-Hu-1 spike (S) protein after the third dose in both groups, although nAbs in KTR were lower than controls. nAbs against pseudoviruses expressing the Omicron S protein were low in both groups, with no increase after the 3rd dose in KTR. Reactivity of CD4+ T cells after boosting was observed when cells were challenged with Wuhan-Hu-1 S peptides, while Omicron S peptides were less effective in both groups. IFN-γ production was detected in KTR in response to ancestral S peptides, confirming antigen-specific T cell activation. Our study demonstrates that the 3rd mRNA dose induces T cell response against Wuhan-Hu-1 spike peptides in KTR, and an increment in the humoral immunity. Instead, humoral and cellular immunity to Omicron variant immunogenic peptides were low in both KTR and healthy vaccinated subjects. [ABSTRACT FROM AUTHOR]

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of the Coronavirus Disease 2019 (COVID-19) pandemic, which is still a health issue worldwide mostly due to a high rate of contagiousness conferred by the high-affinity binding between cell viral receptors, Angiotensin-Converting Enzyme 2 (ACE2) and SARS-CoV-2 Spike protein. Therapies have been developed that rely on the use of antibodies or the induction of their production (vaccination), but despite vaccination being still largely protective, the efficacy of antibody-based therapies wanes with the advent of new viral variants. Chimeric Antigen Receptor (CAR) therapy has shown promise for tumors and has also been proposed for COVID-19 treatment, but as recognition of CARs still relies on antibody-derived sequences, they will still be hampered by the high evasion capacity of the virus. In this manuscript, we show the results from CAR-like constructs with a recognition domain based on the ACE2 viral receptor, whose ability to bind the virus will not wane, as Spike/ACE2 interaction is pivotal for viral entry. Moreover, we have developed a CAR construct based on an affinity-optimized ACE2 and showed that both wild-type and affinity-optimized ACE2 CARs drive activation of a T cell line in response to SARS-CoV-2 Spike protein expressed on a pulmonary cell line. Our work sets the stage for the development of CAR-like constructs against infectious agents that would not be affected by viral escape mutations and could be developed as soon as the receptor is identified. [ABSTRACT FROM AUTHOR]

The induction of antiviral innate immunity by systemic immunization with live virus can be employed to positively impact the response to therapeutic vaccination. We previously demonstrated that systemic immunization with a non-replicating MVA encoding CD40 ligand (CD40L) enhances innate immune cell activation and function, and triggers potent antitumor CD8+ T cell responses in different murine tumor models. Antitumor efficacy was increased when combined with tumor targeting antibodies. Here we report the development of TAEK-VAC-HerBy (TVH), a first-in-class human tumor antibody enhanced killing (TAEK) vaccine based on the non-replicating MVA-BN viral vector. It encodes the membrane bound form of human CD40L, HER2 and the transcription factor Brachyury. TVH is designed for therapeutic use in HER2- or Brachyury-expressing cancer patients in combination with tumor targeting antibodies. To preclude possible oncogenic activities in infected cells and to prevent binding of vaccine-encoded HER2 by monoclonal antibodies trastuzumab and pertuzumab, genetic modifications of HER2 were introduced in the vaccine. Brachyury was genetically modified to prevent nuclear localization of the protein thereby inhibiting its transcriptional activity. CD40L encoded in TVH enhanced human leukocyte activation and cytokine secretion in vitro. Lastly, TVH intravenous administration to non-human primates was proven immunogenic and safe in a repeat-dose toxicity study. Nonclinical data presented here highlight TVH as a first-in-class immunotherapeutic vaccine platform currently under clinical investigation. [ABSTRACT FROM AUTHOR]

Simple Summary: Flow cytometry is a technique that allows identifying different cell populations based on their morphological characteristics and the presence of proteins in the membrane or inside the cell. It is widely used in biomedical research and clinical diagnosis; however, in the veterinary area, its use has been limited due to the reduced availability of fluorochrome-conjugated antibodies that recognize specific proteins. In order to implement multiparametric analyses in the study of bovine tuberculosis, this work designed two panels composed of different conjugates and tested them to identify helper, cytotoxic, activated, and memory T cells in heifers positively and negatively tested for tuberculin (diagnostic test for bovine tuberculosis). Both panels allowed the identification of the cells of interest; in addition, a higher number of activated and memory cells were observed in tuberculin-positive heifers. These panels can also be used in immunopathogenesis studies, as well as in the evaluation of vaccines in cattle. Flow cytometry (FC) is widely used in microbiology, immunology, hematology, and oncology. In the veterinary field, FC enabled the study of the immune response in cattle infected with different pathogens, as well as vaccine testing. However, few fluorochrome-conjugated antibodies recognize bovine antigens, limiting the possible benefits of FC and the implementation of multiparametric analysis for more complex studies. Two cytometry panels with five colors each were designed and implemented for the study and identification of populations and subpopulations of T cells derived from the peripheral blood mononuclear cells of dairy heifers. Both panels detected differences in T cell subpopulations between heifers positively and negatively tested for tuberculin; they detected overexpression of CD25+ and CD45RO+ in tuberculin-positive heifers after stimulation with a culture filtrate protein extract (CFPE) from Mycobacterium bovis (M. bovis). We identified subpopulations of T cells from peripheral blood mononuclear cells using two multicolor panels. These panels could be used to analyze total bovine blood in immunopathogenic studies and vaccine development. The same strategy could be implemented in other species of veterinary interest. [ABSTRACT FROM AUTHOR]

Exhaustion of chimeric antigen receptor (CAR) T cells is one of the limitations for CAR T efficacy in solid tumors and for tumor recurrence after initial CAR T treatment. Tumor treatment with a combination of programmed cell death receptor-1 (PD-1)/programmed cell death ligand-1 (PD-L1) blockage and CD28-based CAR T cells has been intensively studied. However, it remains largely unclear whether autocrine single-chain variable fragments (scFv) PD-L1 antibody can improve 4-1BB-based CAR T cell anti-tumor activity and revert CAR T cell exhaustion. Here, we studied T cells engineered with autocrine PD-L1 scFv and 4-1BB-containing CAR. The antitumor activity and exhaustion of CAR T cells were investigated in vitro and in a xenograft cancer model using NCG mice. CAR T cells with autocrine PD-L1 scFv antibody demonstrate enhanced anti-tumor activity in solid tumors and hematologic malignancies by blocking the PD-1/PD-L1 signaling. Importantly, we found that CAR T exhaustion was largely diminished by autocrine PD-L1 scFv antibody in vivo. As such, 4-1BB CAR T with autocrine PD-L1 scFv antibody combined the power of CAR T cells and the immune checkpoint inhibitor, thereby increasing the anti-tumor immune function and CAR T persistence, providing a cell therapy solution for a better clinical outcome. [ABSTRACT FROM AUTHOR]

Combining optimized spike (S) protein-encoding mRNA vaccines to target multiple SARS-CoV-2 variants could improve control of the COVID-19 pandemic. We compare monovalent and bivalent mRNA vaccines encoding B.1.351 (Beta) and/or B.1.617.2 (Delta) SARS-CoV-2 S-protein in a transgenic mouse and a Wistar rat model. The blended low-dose bivalent mRNA vaccine contains half the mRNA of each respective monovalent vaccine, but induces comparable neutralizing antibody titres, enrichment of lung-resident memory CD8+ T cells, antigen-specific CD4+ and CD8+ responses, and protects transgenic female mice from SARS-CoV-2 lethality. The bivalent mRNA vaccine significantly reduces viral replication in both Beta- and Delta-challenged mice. Sera from bivalent mRNA vaccine immunized female Wistar rats also contain neutralizing antibodies against the B.1.1.529 (Omicron BA.1 and BA.5) variants. These data suggest that low-dose and fit-for-purpose multivalent mRNA vaccines encoding distinct S-proteins are feasible approaches for extending the coverage of vaccines for emerging and co-circulating SARS-CoV-2 variants. Here the authors show efficacy of a low-dose, unmodified, bivalent mRNA vaccine against SARS-CoV-2 variants in two female rodent models and find that combination of mRNA encoding Beta and Delta Spike sequences induces broadly neutralizing antibodies and robust T-cell responses. [ABSTRACT FROM AUTHOR]

Objectives To analyse humoral and cellular immune response to mRNA COVID-19 vaccines in patients with GCA. Methods Consecutive patients with a diagnosis of GCA receiving two doses of BNT162b2 vaccine were assessed at baseline and 3 weeks from the second vaccine dose. Healthy subjects (n  = 51) were included as controls (HC). Humoral response was assessed with Spike-specific IgG antibody response (S-IgG) and neutralizing antibodies (NtAb). Specific T cell response was assessed by enzyme linked immunosorbent spot (ELISpot). Results Of 56 included patients with GCA, 44 were eligible after exclusion of previous evidence of COVID-19 and incomplete follow-up. A significant proportion of patients with GCA (91%) demonstrated antibody (S-IgG) response, but this was significantly lower than HCs (100%); P  < 0.0001. Neutralizing activity was not detected in 16% of patients with GCA. Antibody titres (S-IgG and NtAb) were significantly lower compared with HCs. Humoral response (S-IgG and NtAb) was significantly hampered by treatment with MTX. Cellular response was lacking in 30% of patients with GCA (vs 0% in HCs; P  < 0.0001). Cellular response was significantly influenced by the levels of baseline peripheral T-lymphocytes and by glucocorticoid treatment. Treatment with tocilizumab did not affect any level of the immune response elicited by vaccination. Conclusions Although patients with GCA apparently achieve a robust antibody seroconversion, there is a significant impairment of the neutralizing activity. MTX significantly reduced all levels of the humoral response. Up to one-third of patients do not develop a cellular immune protection in response to COVID-19 vaccination. [ABSTRACT FROM AUTHOR]

Diabetes mellitus is a group of metabolic disorders with different etiologies, pathogeneses and clinical pictures, characterized by chronic hyperglycemia due to abnormal insulin secretion or action. Type 1 diabetes mellitus is the most common type of diabetes mellitus in children and adolescents, accounting for about 90% of diabetes in the population under the age of 18. The etiopathogenesis of type 1 diabetes is multifactorial. The disease occurs as a result of the interaction of three factors: genetic predisposition, environmental factors and the immune response. Research in recent years has focused on the involvement of Epstein–Barr virus (EBV) in the pathogenesis of type I diabetes. The goals of treating type 1 diabetes include maintaining blood-glucose, fructosamine and glycated hemoglobin (HbA1c) levels; therefore, the main purpose of this study was to evaluate the effect of EBV infection on the activation of selected immune cells, fructosamine levels and HbA1c levels in children with type I diabetes. Based on our study, we found a lower percentage of CD8+ T lymphocytes with expression of the CD69 molecule in patients with anti-VCA antibodies in the IgG class, and a lower percentage of CD8+ T lymphocytes with expression of the CD25+ molecule in patients with anti-EBNA-1 antibodies in the IgG class, which may indicate limited control of the immune system during EBV infection in patients. There was a lower percentage of CD3+CD4+ T lymphocytes secreting IL-4 in the study group, indicating that a deficiency in IL-4 production may be related to the development of type 1 diabetes. There was an increase in the percentage of CD4+CD3+IL-10 lymphocytes in the study group with anti-VCA antibodies present in the IgG class and anti-EBNA-1 antibodies in the IgG class compared to the patients without antibodies. In addition, there was a significant increase in fructosamine levels and higher glycated hemoglobin levels in the study group with antibodies to EBV antigens. In addition, an increase in the percentage of T lymphocytes with a CD4+CD3+IL-17+ phenotype in the patients with anti-VCA IgG antibodies was confirmed, and higher HbA1c levels may suggest that EBV infection is accompanied by an increase in IL-17 secretion. [ABSTRACT FROM AUTHOR]

Simple Summary: Therapeutic options for rare cancers are frequently limited and less effective than for common cancers. Therefore, novel therapeutic strategies to treat rare cancers are urgently required. Our clinical study on rare cancers showed that the biweekly WT1 Trio peptide vaccine comprising two WT1-cytotoxic T lymphocyte (CTL)-peptides and one WT1-helper T lymphocyte-peptide induced more robust immune responses targeting WT1 than the weekly WT1-CTL peptide (WT1-235) vaccine. In addition, the safety of WT1 Trio was confirmed without severe treatment-related adverse events, except grade 3 myasthenia gravis-like symptoms observed in a patient with thymic cancer (TC). Fifteen (33.3%) of the 45 patients with recurrent or advanced rare cancers, including malignant glioma, soft-tissue sarcoma, TC, and malignant pleural mesothelioma, achieved stable disease after 3 months of protocol treatment. Therefore, since WT1 is widely overexpressed in rare cancers, WT1-targeted immunotherapy may be a therapeutic strategy for rare cancers. 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. [ABSTRACT FROM AUTHOR]

Vaccines are among the most effective medical countermeasures against infectious diseases. The emergence of the Coronavirus Disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spurred scientific strategies to fight against the disease. Since 2020, in response to the pandemic, many vaccines based on different platforms have been under development, among which mRNA, adenoviral vectors, and subunit vaccines have been clinically approved for use in humans. These first-generation COVID-19 vaccines largely target the viral spike (S) protein and are aimed at eliciting potent neutralizing antibodies. With the emergence of SARS-CoV-2 variants, particularly the highly transmissible Omicron strains, S-based vaccine strategies have faced a continuing challenge of strong immune escape by variants. The coronavirus nucleocapsid (N) protein is a viral protein that induces strong T-cell immunity and is more conserved than S protein across different SARS-CoV-2 variants. Inclusion of N protein in the development of COVID-19 vaccines has been reported. Here, we briefly review and discuss COVID-19, current S-protein-based vaccine strategies, the immunobiology of N protein in SARS-CoV-2 host immunity, and nextgeneration vaccine strategies involving N protein to combat current and emerging SARS-CoV-2 variants. [ABSTRACT FROM AUTHOR]

In a representative cohort of immunocompromised young people, we observed marked differences in humoral and cellular responses to a standard two-dose mRNA vaccination regime between disease categories. In conclusion, although immunocompromised young people display attenuated humoral and cellular responses to vaccination in general, there are marked differences in vaccine responses between disease subgroups, not completely explained by immunosuppression used. First, given the heterogeneity in vaccine responses, some patient groups who qualify for an additional primary vaccination dose, for example JIA patients, may not benefit significantly, resulting in suboptimal use of vaccines and vaccination capacity. [Extracted from the article]

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Background. Liangxue Tongyu prescription (LTP) is a commonly used formula for acute intracerebral hemorrhage (AICH) in clinical practice that has significant ameliorative effects on neurological deficits and gastrointestinal dysfunction, yet the mechanism remains elusive. The aim of this study was to investigate the pathway by which LTP alleviates brain damage in AICH rats. Methods. The AICH rat models were established by autologous caudal arterial blood injection. The neurological function scores were evaluated before and after treatment. The water content and the volume of Evans blue staining in the brain were measured to reflect the degree of brain damage. RT-PCR was used to detect the inflammatory factors of the brain. Western blotting was used to detect the expression of the tight junction proteins zonula occludens 1 (ZO-1), occludin (OCLN), and claudin (CLDN) in the brain and colon, followed by mucin 2 (MUC2), secretory immunoglobulin A (SIgA), and G protein-coupled receptor 43 (GPR43) in the colon. Flow cytometry was used to detect the ratios of helper T cells 17 (Th17) and regulatory T cells (Treg) in peripheral blood, and the vagus nerve (VN) discharge signals were collected. Results. LTP reduced the brain damage of the AICH rats. Compared with the model group, LTP significantly improved the permeability of the colonic mucosa, promoted the secretion of MUC2, SigA, and GPR43 in the colon, and regulated the immune balance of peripheral T cells. The AICH rats had significantly faster VN discharge rates and lower amplitudes than normal rats, and these abnormalities were corrected in the LTP and probiotics groups. Conclusion. LTP can effectively reduce the degree of brain damage in AICH rats, and the mechanism may be that it can play a neuroprotective role by regulating the function of the intestinal mucosal barrier. [ABSTRACT FROM AUTHOR]

It is known that schistosome-derived antigens induce innate and adaptive immune responses that are essential for the formation of hepatic immunopathology. Here, we screened and synthesized four peptides derived from Schistosoma japonicum (S. japonicum) heat shock protein 90α (Sjp90α-1, -2, -3, and -4), which is widely expressed in adults and eggs of the genus S. japonicum and induces remarkable immune reactions. To define the antigenicity of these peptides, we stimulated splenocytes with peptides, and the results showed that only the Sjp90α-1 peptide could predominately induce the activation of dendritic cells (DCs) and macrophages as well as alter the proportion of follicular helper T (Tfh) cells. Next, CD4+ T cells were purified and cocultured with mouse bone-marrow-derived DCs (BMDCs) with or without Sjp90α-1 peptide stimulation in vitro, and the results showed that Sjp90α-1-stimulated BMDCs can significantly induce CD4+ T-cell differentiation into Tfh cells, while the direct stimulation of CD4+ T cells with Sjp90α-1 did not induce Tfh cells, indicating that the Sjp90α-1 peptide promotes Tfh cell differentiation depending on the presence of DCs. Furthermore, we selected and prepared an Sjp90α-1-peptide-based antibody and illustrated that it has excellent reactivity with the immunizing peptide and detects a single band of 29 kDa corresponding to the Sjp90α protein. The immunolocalization results showed that the protein recognized by this Sjp90α-1-peptide-based antibody is present in the mature eggs and the tegument of adults, implying that the parasite-derived peptide has a potential interaction with the host immune system. Finally, we evaluated antipeptide IgG antibodies and revealed a significantly higher level of anti-Sjp90α-1 peptide IgG antibodies in mice 3 weeks after S. japonicum infection. In conclusion, we illustrate that these synthetic peptides warrant further investigation by evaluating their antigen-specific immune response and their ability to efficiently induce Tfh cells. Moreover, they may constitute a potentially helpful method for the laboratory diagnosis of schistosomiasis japonica. [ABSTRACT FROM AUTHOR]

Hopefully, studies on host immune response will accelerate vaccine development and will result in new commercially available vaccines in the future. Vaccines are one of the greatest achievements of modern medicine, offering an effective way to fight and control infectious diseases. In contrast to many anti-viral and anti-bacterial vaccines available for human and animal use, the paucity of anti-parasitic vaccines shows discrepancies in the development of effective immunisation strategies depending on the pathogen. Whereas the first vaccines were based on attenuated or killed pathogens, purified pathogen antigens (subunit vaccines) were subsequently employed. [Extracted from the article]

A paucity of effector T cells within tumors renders pancreatic ductal adenocarcinoma (PDAC) resistant to immune checkpoint therapies. While several under-development approaches target immune-suppressive cells in the tumor microenvironment, there is less focus on improving T cell function. Here we show that inhibiting vasoactive intestinal peptide receptor (VIP-R) signaling enhances anti-tumor immunity in murine PDAC models. In silico data mining and immunohistochemistry analysis of primary tumors indicate overexpression of the neuropeptide vasoactive intestinal peptide (VIP) in human PDAC tumors. Elevated VIP levels are also present in PDAC patient plasma and supernatants of cultured PDAC cells. Furthermore, T cells up-regulate VIP receptors after activation, identifying the VIP signaling pathway as a potential target to enhance T cell function. In mouse PDAC models, VIP-R antagonist peptides synergize with anti-PD-1 antibody treatment in improving T cell recruitment into the tumors, activation of tumor-antigen-specific T cells, and inhibition of T cell exhaustion. In contrast to the limited single-agent activity of anti-PD1 antibodies or VIP-R antagonist peptides, combining both therapies eliminate tumors in up to 40% of animals. Furthermore, tumor-free mice resist tumor re-challenge, indicating anti-cancer immunological memory generation. VIP-R signaling thus represents a tumor-protective immune-modulatory pathway that is targetable in PDAC. Poor antitumor response of pancreatic cancer to immunotherapies is a major barrier to effective disease management. Herein we show that pancreatic cancers overexpress vasoactive intestinal peptide, and pharmacological inhibition of its signaling significantly enhances responsiveness of pancreatic ductal adenocarcinoma to immune checkpoint therapy, thus improving overall survival in mouse models. [ABSTRACT FROM AUTHOR]

Antibodies against the spike protein of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) can drive adaptive evolution in immunocompromised patients with chronic infection. Here we longitudinally analyze SARS-CoV-2 sequences in a B cell-depleted, lymphoma patient with chronic, ultimately fatal infection, and identify three mutations in the spike protein that dampen convalescent plasma-mediated neutralization of SARS-CoV-2. Additionally, four mutations emerge in non-spike regions encoding three CD8 T cell epitopes, including one nucleoprotein epitope affected by two mutations. Recognition of each mutant peptide by CD8 T cells from convalescent donors is reduced compared to its ancestral peptide, with additive effects resulting from double mutations. Querying public SARS-CoV-2 sequences shows that these mutations have independently emerged as homoplasies in circulating lineages. Our data thus suggest that potential impacts of CD8 T cells on SARS-CoV-2 mutations, at least in those with humoral immunodeficiency, warrant further investigation to inform on vaccine design. SARS-CoV-2 mutations associated with the escape from antibody-mediated neutralization have been widely reported. Here, in a patient with defective antibody responses, the authors find a potential association between SARS-CoV-2 mutations and CD8 T alterations to implicate possible contributions of CD8 T cells in evasion of SARS-CoV-2 from host immunity. [ABSTRACT FROM AUTHOR]

Although the Omicron variant of the SARS-CoV-2 virus shows resistance to neutralizing antibody, it retains susceptibility to the cellular immune response. Here we characterize vaccine-induced T cells specific for various SARS-CoV-2 variants and identified HLA-A*24:02-restricted CD8+ T cells that strongly suppress Omicron BA.1 replication in vitro. Mutagenesis analyses revealed that a G446S mutation, located just outside the N-terminus of the cognate epitope, augmented TCR recognition of this variant. In contrast, no enhanced suppression of replication is observed against cells infected with the prototype, Omicron BA.2, and Delta variants that express G446. The enhancing effect of the G446S mutation is lost when target cells are treated with inhibitors of tripeptidyl peptidase II, a protein that mediates antigen processing. These ex vivo analysis and in vitro results demonstrate that the G446S mutation in the Omicron BA.1 variant affects antigen processing/presentation and potentiates antiviral activity by vaccine-induced T cells, leading to enhanced T cell recognition towards emerging variants. Mutations in the spike of SARS-CoV-2 can result in the escape of the neutralising antibody response but may retain susceptibility to the cellular immune response. Here the authors show the G446S mutation in the spike protein of Omicron BA.1 is associated with altered antigen presentation and potentiates activation of specific T cell immunity. [ABSTRACT FROM AUTHOR]

To better understand the development of SARS-CoV-2-specific immunity over time, a detailed evaluation of humoral and cellular responses is required. Here, we characterize anti-Spike (S) IgA and IgG in a representative population-based cohort of 431 SARS-CoV-2-infected individuals up to 217 days after diagnosis, demonstrating that 85% develop and maintain anti-S responses. In a subsample of 64 participants, we further assess anti-Nucleocapsid (N) IgG, neutralizing antibody activity, and T cell responses to Membrane (M), N, and S proteins. In contrast to S-specific antibody responses, anti-N IgG levels decline substantially over time and neutralizing activity toward Delta and Omicron variants is low to non-existent within just weeks of Wildtype SARS-CoV-2 infection. Virus-specific T cells are detectable in most participants, albeit more variable than antibody responses. Cluster analyses of the co-evolution of antibody and T cell responses within individuals identify five distinct trajectories characterized by specific immune patterns and clinical factors. These findings demonstrate the relevant heterogeneity in humoral and cellular immunity to SARS-CoV-2 while also identifying consistent patterns where antibody and T cell responses may work in a compensatory manner to provide protection. The persistence of the immune response to SARS-CoV-2 after recovery from infection is an indicator for subsequent protection against infection. Here the authors follow recovered patients and measure antibody and T cell responses and find that these two parts of the immune response may have different longevity. [ABSTRACT FROM AUTHOR]