Durable CD8 T Cell Memory against SARS-CoV-2 by Prime/Boost and Multi-Dose Vaccination: Considerations on Inter-Dose Time Intervals.

Facing the COVID-19 pandemic, anti-SARS-CoV-2 vaccines were developed at unprecedented pace, productively exploiting contemporary fundamental research and prior art. Large-scale use of anti-SARS-CoV-2 vaccines has greatly limited severe morbidity and mortality. Protection has been correlated with high serum titres of neutralizing antibodies capable of blocking the interaction between the viral surface protein spike and the host SARS-CoV-2 receptor, ACE-2. Yet, vaccine-induced protection subsides over time, and breakthrough infections are commonly observed, mostly reflecting the decay of neutralizing antibodies and the emergence of variant viruses with mutant spike proteins. Memory CD8 T cells are a potent weapon against viruses, as they are against tumour cells. Anti-SARS-CoV-2 memory CD8 T cells are induced by either natural infection or vaccination and can be potentially exploited against spike-mutated viruses. We offer here an overview of current research about the induction of anti-SARS-CoV-2 memory CD8 T cells by vaccination, in the context of prior knowledge on vaccines and on fundamental mechanisms of immunological memory. We focus particularly on how vaccination by two doses (prime/boost) or more (boosters) promotes differentiation of memory CD8 T cells, and on how the time-length of inter-dose intervals may influence the magnitude and persistence of CD8 T cell memory.

Adjuvant effect of TLR7 agonist adsorbed on aluminum hydroxide (AS37): A phase I randomized, dose escalation study of an AS37-adjuvanted meningococcal C conjugated vaccine.

An adjuvant system (AS37) has been developed containing a synthetic toll-like receptor agonist (TLR7a). We conducted a phase I randomized, observer-blind, dose-escalation study to assess the safety and immunogenicity of an investigational AS37-adjuvanted meningococcus C (MenC) conjugate vaccine in healthy adults (NCT02639351). A control group received a licensed MenC conjugate alum-adjuvanted vaccine. Eighty participants were randomized to receive one dose of control or investigational vaccine containing AS37 (TLR7a dose 12.5, 25, 50, 100 µg). All vaccines were well tolerated, apart from in the TLR7a 100 µg dose group, which had three reports (18.8%) of severe systemic adverse events. Four weeks after vaccination, human complement serum bactericidal assay seroresponse rates against MenC were 56-81% in all groups, and ELISA seroresponses were ≥81% for all AS37-adjuvanted vaccine groups (100% in 50 and 100 µg dose groups) and 88% in the control group. Antibody responses were maintained at six months after vaccination.

Vaccine adjuvant MF59 promotes retention of unprocessed antigen in lymph node macrophage compartments and follicular dendritic cells.

Ag retention within lymph nodes (LNs) upon vaccination is critical for the development of adaptive immune responses, because it facilitates the encounter of the Ag with cognate lymphocytes. During a secondary exposure of the immune system to an Ag, immune complexes (ICs) that contain the unprocessed Ag are captured by subcapsular sinus macrophages and are transferred onto follicular dendritic cells, where they persist for weeks, facilitating Ag presentation to cognate memory B cells. The impact of adjuvants on Ag retention within the draining LNs is unknown. In this article, we provide the first evidence, to our knowledge, that the oil-in-water emulsion adjuvant MF59 localizes in subcapsular sinus and medullary macrophage compartments of mouse draining LNs, where it persists for at least 2 wk. In addition, we demonstrate that MF59 promotes accumulation of the unprocessed Ag within these LN compartments and facilitates the consequent deposition of the IC-trapped Ag onto activated follicular dendritic cells. These findings correlate with the ability of MF59 to boost germinal center generation and Ag-specific Ab titers. Our data suggest that the adjuvant effect of MF59 is, at least in part, due to an enhancement of IC-bound Ag retention within the LN and offer insights to improve the efficacy of new vaccine adjuvants.

The Human Pathogen Streptococcus pyogenes Releases Lipoproteins as Lipoprotein-rich Membrane Vesicles.

Bacterial lipoproteins are attractive vaccine candidates because they represent a major class of cell surface-exposed proteins in many bacteria and are considered as potential pathogen-associated molecular patterns sensed by Toll-like receptors with built-in adjuvanticity. Although Gram-negative lipoproteins have been extensively characterized, little is known about Gram-positive lipoproteins. We isolated from Streptococcus pyogenes a large amount of lipoproteins organized in vesicles. These vesicles were obtained by weakening the bacterial cell wall with a sublethal concentration of penicillin. Lipid and proteomic analysis of the vesicles revealed that they were enriched in phosphatidylglycerol and almost exclusively composed of lipoproteins. In association with lipoproteins, a few hypothetical proteins, penicillin-binding proteins, and several members of the ExPortal, a membrane microdomain responsible for the maturation of secreted proteins, were identified. The typical lipidic moiety was apparently not necessary for lipoprotein insertion in the vesicle bilayer because they were also recovered from the isogenic diacylglyceryl transferase deletion mutant. The vesicles were not able to activate specific Toll-like receptor 2, indicating that lipoproteins organized in these vesicular structures do not act as pathogen-associated molecular patterns. In light of these findings, we propose to name these new structures Lipoprotein-rich Membrane Vesicles.

Modulation of endotoxicity of Shigella generalized modules for membrane antigens (GMMA) by genetic lipid A modifications: relative activation of TLR4 and TLR2 pathways in different mutants.

Outer membrane particles from Gram-negative bacteria are attractive vaccine candidates as they present surface antigens in their natural context. We previously developed a high yield production process for genetically derived particles, called generalized modules for membrane antigens (GMMA), from Shigella. As GMMA are derived from the outer membrane, they contain immunostimulatory components, especially lipopolysaccharide (LPS). We examined ways of reducing their reactogenicity by modifying lipid A, the endotoxic part of LPS, through deletion of late acyltransferase genes, msbB or htrB, in GMMA-producing Shigella sonnei and Shigella flexneri strains. GMMA with resulting penta-acylated lipid A from the msbB mutants showed a 600-fold reduced ability, and GMMA from the S. sonnei ΔhtrB mutant showed a 60,000-fold reduced ability compared with GMMA with wild-type lipid A to stimulate human Toll-like receptor 4 (TLR4) in a reporter cell line. In human peripheral blood mononuclear cells, GMMA with penta-acylated lipid A showed a marked reduction in induction of inflammatory cytokines (S. sonnei ΔhtrB, 800-fold; ΔmsbB mutants, 300-fold). We found that the residual activity of these GMMA is largely due to non-lipid A-related TLR2 activation. In contrast, in the S. flexneri ΔhtrB mutant, a compensatory lipid A palmitoleoylation resulted in GMMA with hexa-acylated lipid A with ∼10-fold higher activity to stimulate peripheral blood mononuclear cells than GMMA with penta-acylated lipid A, mostly due to retained TLR4 activity. Thus, for use as vaccines, GMMA will likely require lipid A penta-acylation. The results identify the relative contributions of TLR4 and TLR2 activation by GMMA, which need to be taken into consideration for GMMA vaccine development.

CD8 T-cell priming upon mRNA vaccination is restricted to bone-marrow-derived antigen-presenting cells and may involve antigen transfer from myocytes.

Self-amplifying mRNAs (SAM(®) ) are a novel class of nucleic acid vaccines, delivered by a non-viral delivery system. They are effective at eliciting potent and protective immune responses and are being developed as a platform technology with potential to be used for a broad range of targets. However, their mechanism of action has not been fully elucidated. To date, no evidence of in vivo transduction of professional antigen-presenting cells (APCs) by SAM vector has been reported, while the antigen expression has been shown to occur mostly in the muscle fibres. Here we show that bone-marrow-derived APCs rather than muscle cells are responsible for induction of MHC class-I restricted CD8 T cells in vivo, but direct transfection of APCs by SAM vectors is not required. Based on all our in vivo and in vitro data we propose that upon SAM vaccination the antigen is expressed within muscle cells and then transferred to APCs, suggesting cross-priming as the prevalent mechanism for priming the CD8 T-cell response by SAM vaccines.

Development and Application of a Novel SPE-Method for Bioassay-Guided Fractionation of Marine Extracts.

The biological diversity of marine habitats is a unique source of chemical compounds with potential use as pharmaceuticals, cosmetics and dietary supplements. However, biological screening and chemical analysis of marine extracts pose specific technical constraints and require adequate sample preparation. Here we report an improved method on Solid Phase Extraction (SPE) to fractionate organic extracts containing high concentration of salt that hampers the recovery of secondary metabolites. The procedure uses a water suspension to load the extracts on a poly(styrene-divynylbenzene)-based support and a stepwise organic solvent elution to effectively desalt and fractionate the organic components. The novel protocol has been tested on MeOH-soluble material from three model organisms (Reniera sarai, Dendrilla membranosa and Amphidinium carterae) and was validated on a small panel of 47 marine samples, including sponges and protists, within discovery programs for identification of immuno-stimulatory and anti-infective natural products.

Ultrastructural Visualization of Vaccine Adjuvant Uptake In Vitro and In Vivo.

Adjuvants are substances that enhance adaptive immune responses when formulated in a vaccine. Alum and MF59 are two vaccine adjuvants licensed for human vaccination. Their mode of action has not been completely elucidated. Here we show the first ultrastructural visualization of Alum and MF59 interaction with immune cells in vitro and in vivo. We observed that Alum is engulfed by cells as inclusions of laminae that are detectable within draining lymph nodes. MF59 is instead engulfed by cells in vitro as low-electron-dense lipid-like inclusions that display a vesicle pattern, as confirmed by confocal microscopy using fluorescently labeled MF59. However, lipid-like inclusions with different high- and low-electron-dense content are detected within cells of draining lymph nodes when injecting MF59. As high-electron-dense lipid-like inclusions are also detected upon injection of Alum, our results suggest that the low-electron-dense inclusions are formed by engulfed MF59, whereas the high-electron-dense inclusions are proper lipid inclusions. Thus, we demonstrated that vaccine adjuvants are engulfed as inclusions by lymph node cells and hypothesize that adjuvant treatment may modify lipid metabolism.

Adjuvanticity of the oil-in-water emulsion MF59 is independent of Nlrp3 inflammasome but requires the adaptor protein MyD88.

Oil-in-water emulsions have been successfully used to increase the efficacy, immunogenicity, and cross-protection of human vaccines; however, their mechanism of action is still largely unknown. Nlrp3 inflammasome has been previously associated to the activity of alum, another adjuvant broadly used in human vaccines, and MyD88 adaptor protein is required for the adjuvanticity of most Toll-like receptor agonists. We compared the contribution of Nlrp3 and MyD88 to the adjuvanticity of alum, the oil-in-water emulsion MF59, and complete Freund's adjuvant in mice using a three-component vaccine against serogroup B Neisseria meningitidis (rMenB). Although the basal antibody responses to the nonadjuvanted rMenB vaccine were largely dependent on Nlrp3, the high-level antibody responses induced by alum, MF59, or complete Freund's adjuvant did not require Nlrp3. Surprisingly, we found that MF59 requires MyD88 to enhance bactericidal antibody responses to the rMenB vaccine. Because MF59 did not activate any of the Toll-like receptors in vitro, we propose that MF59 requires MyD88 for a Toll-like receptor-independent signaling pathway.

The scientific journey of a novel adjuvant (AS37) from bench to bedside.

A decade ago, we described a new approach to discover next generation adjuvants, identifying small-molecule immune potentiators (SMIPs) as Toll-like receptor (TLR)7 agonists. We also optimally formulated these drugs through adsorption to aluminum salts (alum), allowing them to be evaluated with a range of established and early-stage vaccines. Early proof-of-concept studies showed that a TLR7 agonist (TLR7a)-based SMIP, when adsorbed to alum, could perform as an effective adjuvant for a variety of different antigens, in both small and large animals. Studies in rodents demonstrated that the adjuvant enhanced immunogenicity of a recombinant protein-based vaccine against Staphylococcus aureus, and also showed potential to improve existing vaccines against pertussis or meningococcal infection. Extensive evaluations showed that the adjuvant was effective in non-human primates (NHPs), exploiting a mechanism of action that was consistent across the different animal models. The adjuvant formulation (named AS37) has now been advanced into clinical evaluation. A systems biology-based evaluation of the phase I clinical data with a meningococcal C conjugate vaccine showed that the AS37-adjuvanted formulation had an acceptable safety profile, was potent, and activated the expected immune pathways in humans, which was consistent with observations from the NHP studies. In the intervening decade, several alternative TLR7 agonists have also emerged and advanced into clinical development, such as the alum adsorbed TLR7/8 SMIP present in a widely distributed COVID-19 vaccine. This review summarizes the research and early development of the new adjuvant AS37, with an emphasis on the steps taken to allow its progression into clinical evaluations.

A point mutation in the amino terminus of TLR7 abolishes signaling without affecting ligand binding.

TLR7 is the mammalian receptor for ssRNA and some nucleotide-like small molecules. We have generated a mouse by N-nitrose-N'-ethyl urea mutagenesis in which threonine 68 of TLR7 was substituted with isoleucine. Cells bearing this mutant TLR7 lost the sensitivity to the small-molecule TLR7 agonist resiquimod, hence the name TLR7(rsq1). In this work, we report the characterization of this mutant protein. Similar to the wild-type counterpart, TLR7(rsq1) localizes to the endoplasmic reticulum and is expressed at normal levels in both primary cells and reconstituted 293T cells. In addition to small-molecule TLR7 agonists, TLR7(rsq1) fails to be activated by ssRNA. Whole-transcriptome analysis demonstrates that TLR7 is the exclusive and indispensable receptor for both classes of ligands, consistent with the fact that both ligands induce highly similar transcriptional signatures in TLR7(wt/wt) splenocytes. Thus, TLR7(rsq1) is a bona fide phenocopy of the TLR7 null mouse. Because TLR7(rsq1) binds to ssRNA, our studies imply that the N-terminal portion of TLR7 triggers a yet to be identified event on TLR7. TLR7(rsq1) mice might represent a valuable tool to help elucidate novel aspects of TLR7 biology.

Systems analysis of human responses to an aluminium hydroxide-adsorbed TLR7 agonist (AS37) adjuvanted vaccine reveals a dose-dependent and specific activation of the interferon-mediated antiviral response.

The candidate Adjuvant System AS37 contains a synthetic toll-like receptor agonist (TLR7a) adsorbed to alum. In a phase I study (NCT02639351), healthy adults were randomised to receive one dose of licensed alum-adjuvanted meningococcal serogroup C (MenC-CRM197) conjugate vaccine (control) or MenC-CRM197 conjugate vaccine adjuvanted with AS37 (TLR7a dose 12.5, 25, 50 or 100 µg). A subset of 66 participants consented to characterisation of peripheral whole blood transcriptomic responses, systemic cytokine/chemokine responses and multiple myeloid and lymphoid cell responses as exploratory study endpoints. Blood samples were collected pre-vaccination, 6 and 24 h post-vaccination, and 3, 7, 28 and 180 days post-vaccination. The gene expression profile in whole blood showed an early, AS37-specific transcriptome response that peaked at 24 h, increased with TLR7a dose up to 50 µg and generally resolved within one week. Five clusters of differentially expressed genes were identified, including those involved in the interferon-mediated antiviral response. Evaluation of 30 cytokines/chemokines by multiplex assay showed an increased level of interferon-induced chemokine CXCL10 (IP-10) at 24 h and 3 days post-vaccination in the AS37-adjuvanted vaccine groups. Increases in activated plasmacytoid dendritic cells (pDC) and intermediate monocytes were detected 3 days post-vaccination in the AS37-adjuvanted vaccine groups. T follicular helper (Tfh) cells increased 7 days post-vaccination and were maintained at 28 days post-vaccination, particularly in the AS37-adjuvanted vaccine groups. Moreover, most of the subjects that received vaccine containing 25, 50 and 100 µg TLR7a showed an increased MenC-specific memory B cell responses versus baseline. These data show that the adsorption of TLR7a to alum promotes an immune signature consistent with TLR7 engagement, with up-regulation of interferon-inducible genes, cytokines and frequency of activated pDC, intermediate monocytes, MenC-specific memory B cells and Tfh cells. TLR7a 25-50 µg can be considered the optimal dose for AS37, particularly for the adjuvanted MenC-CRM197 conjugate vaccine.

Differential IL-17 production and mannan recognition contribute to fungal pathogenicity and commensalism.

In this study, we present evidence of differential Th17 responses in human monocyte-derived dendritic cells exposed to the pathogenic Candida albicans or the nonpathogenic Saccharomyces cerevisiae. We use different forms of the microorganisms, cells, hyphae, and spores, as a toolbox to dissect the role of surface mannan in the fungal immune response. In contrast to the S. cerevisiae yeast cell-induced Th1 response, dendritic cells stimulated with spores or C. albicans hyphae induce cellular responses shifted toward Th17 differentiation. The differential recognition of specific mannan structures is the master regulator of the discrimination between harmful and harmless fungi. The switch between spores and yeast is crucial for the commensalism of S. cerevisiae and depends on the use of a different receptor repertoire. Understanding the role of cell wall recognition during infection might lead to understanding the boundaries between safety and pathogenicity.

A Novel Toll-Like Receptor 2 Agonist Protects Mice in a Prophylactic Treatment Model Against Challenge With Bacillus anthracis.

Current therapies for anthrax include the use of antibiotics (i.e., doxycycline, and ciprofloxacin), an anthrax vaccine (BioThrax) and Bacillus anthracis-specific, monoclonal antibody (mAb) (i.e., Raxibacumab and obiltoxaximab). In this study, we investigated the activity of immunomodulators, which potentiate inflammatory responses through innate immune receptors. The rationale for the use of innate immune receptor agonists as adjunctive immunomodulators for infectious diseases is based on the concept that augmentation of host defense should promote the antimicrobial mechanism of the host. Our aim was to explore the anti-B. anthracis effector function of Toll-like receptor (TLR) agonists using a mouse model. Amongst the six TLR ligands tested, Pam3CSK4 (TLR1/2 ligand) was the best at protecting mice from lethal challenge of B. anthracis. We then evaluated the activity of a novel TLR2 ligand, DA-98-WW07. DA-98-WW07 demonstrated enhanced protection in B. anthracis infected mice. The surviving mice that received DA-98-WW07 when re-challenged with B. anthracis 20 days post the first infection showed increased survival rate. Moreover, ciprofloxacin, when treated in adjunct with a suboptimal concentration of DA-98-WW07 demonstrated augmented activity in protecting mice from B. anthracis infection. Taken together, we report the prophylactic treatment potential of DA-98-WW07 for anthrax and the utility of immunomodulators in combination with an antibiotic to treat infections caused by the B. anthracis bacterium.

The respiratory syncytial virus (RSV) prefusion F-protein functional antibody repertoire in adult healthy donors.

Respiratory syncytial virus (RSV) is the leading cause of death from lower respiratory tract infection in infants and children, and is responsible for considerable morbidity and mortality in older adults. Vaccines for pregnant women and elderly which are in phase III clinical studies target people with pre-existing natural immunity against RSV. To investigate the background immunity which will be impacted by vaccination, we single cell-sorted human memory B cells and dissected functional and genetic features of neutralizing antibodies (nAbs) induced by natural infection. Most nAbs recognized both the prefusion and postfusion conformations of the RSV F-protein (cross-binders) while a smaller fraction bound exclusively to the prefusion conformation. Cross-binder nAbs used a wide array of gene rearrangements, while preF-binder nAbs derived mostly from the expansion of B-cell clonotypes from the IGHV1 germline. This latter class of nAbs recognizes an epitope located between Site Ø, Site II, and Site V on the F-protein, identifying an important site of pathogen vulnerability.

Antibody avidity, persistence, and response to antigen recall: comparison of vaccine adjuvants.

Differences in innate immune 'imprinting' between vaccine adjuvants may mediate dissimilar effects on the quantity/quality of persisting adaptive responses. We compared antibody avidity maturation, antibody/memory B cell/CD4+ T cell response durability, and recall responses to non-adjuvanted fractional-dose antigen administered 1-year post-immunization (Day [D]360), between hepatitis B vaccines containing Adjuvant System (AS)01B, AS01E, AS03, AS04, or Alum (NCT00805389). Both the antibody and B cell levels ranked similarly (AS01B/E/AS03 > AS04 > Alum) at peak response, at D360, and following their increases post-antigen recall (D390). Proportions of high-avidity antibodies increased post-dose 2 across all groups and persisted at D360, but avidity maturation appeared to be more strongly promoted by AS vs. Alum. Post-antigen recall, frequencies of subjects with high-avidity antibodies increased only markedly in the AS groups. Among the AS, total antibody responses were lowest for AS04. However, proportions of high-avidity antibodies were similar between groups, suggesting that MPL in AS04 contributes to avidity maturation. Specific combinations of immunoenhancers in the AS, regardless of their individual nature, increase antibody persistence and avidity maturation.

Inhibition of natural killer cells through engagement of CD81 by the major hepatitis C virus envelope protein.

The immune response against hepatitis C virus (HCV) is rarely effective at clearing the virus, resulting in approximately 170 million chronic HCV infections worldwide. Here we report that ligation of an HCV receptor (CD81) inhibits natural killer (NK) cells. Cross-linking of CD81 by the major envelope protein of HCV (HCV-E2) or anti-CD81 antibodies blocks NK cell activation, cytokine production, cytotoxic granule release, and proliferation. This inhibitory effect was observed using both activated and resting NK cells. Conversely, on NK-like T cell clones, including those expressing NK cell inhibitory receptors, CD81 ligation delivered a costimulatory signal. Engagement of CD81 on NK cells blocks tyrosine phosphorylation through a mechanism which is distinct from the negative signaling pathways associated with NK cell inhibitory receptors for major histocompatibility complex class I. These results implicate HCV-E2-mediated inhibition of NK cells as an efficient HCV evasion strategy targeting the early antiviral activities of NK cells and allowing the virus to establish itself as a chronic infection.

Dataset of antibody variable region sequence features inferred from a respiratory syncytial virus fusion protein-specific B cell receptor repertoire induced by natural infection of a healthy adult.

Respiratory syncytial virus (RSV) is the primary cause for acute lower respiratory syndrome in children younger than 5 years. Research on B cell repertoires and antibodies binding the RSV fusion protein (RSV F) is of major interest in the development of potential vaccine candidates and therapies. B cell receptors (BCRs) which have higher affinities for a specific antigen are preferentially selected for B cell clonal expansion in germinal center reactions. Consequently, antigen-specific BCR repertoires share common features, as for instance preferential variable gene usage, variable region mutation levels or lengths of the heavy chain complementarity-determining region 3. Since RSV repeatedly infects every person throughout life, memory B cells (MBC) expressing RSV F-binding BCRs circulate in the blood of healthy adults. This dataset of BCR variable region sequence features was derived from single cell-sorted RSV F-directed MBCs of a healthy adult blood donor [1]. The dataset was produced with publicly available data analysis software programs and scripts, which facilitates integration or comparison with antibody sequence repertoire data of different individuals derived with the same or comparable data analysis approaches and tools.

The respiratory syncytial virus fusion protein-specific B cell receptor repertoire reshaped by post-fusion subunit vaccination.

Respiratory syncytial virus (RSV) is the major cause of acute lower respiratory illness in children of less than 5 years of age which usually results in hospitalization or even in death. Vaccine development is hampered in consequence of a failed vaccine trial with fatalities in the 1960s. Even though research has been more focused on the RSV fusion protein in its pre-fusion conformation, maternal vaccination with post-fusion protein (post F) was considered as a promising vaccine strategy for passive immunization of babies, because post F preserves very potent neutralizing epitopes. We extensively analyzed post F-binding B cell receptor (BCR) repertoires of three vaccinees who received a post F-subunit vaccine in the context of a first-in-human, Phase 1, randomized, observer-blind, placebo-controlled clinical trial (ClinicalTrials.gov Identifier: NCT02298179). In order to compare the vaccine-induced BCR repertoires with BCR repertoires induced by natural infection, we also analyzed pre F- and post F-binding BCRs isolated from a healthy blood donor with relatively high F-binding memory B cell (MBC) frequencies. Analysis of the vaccine-induced repertoires revealed that preferentially VH4-encoded BCRs were expanded in response to vaccination. Estimation of antigen-driven selection further demonstrated that expanded BCRs accumulated positively selected replacement mutations which substantiated the hypothesis that post F-vaccination induces diversification of VH4-encoded BCRs in germinal centers. Comparison of the vaccine-induced BCR repertoires with clonally related pre and post F-binding BCRs of the healthy blood donor suggested that the vaccine expanded pre/post F cross-reactive MBCs. Interestingly, several vaccine-induced BCRs shared stereotypic VDJ gene junctions with known neutralizing Abs. Once expressed for functional characterization, the selected monoclonal Abs demonstrated the predicted neutralization activities in plaque reduction neutralization assays indicating that the post F-vaccine induced expansion of neutralizing BCRs.

Serafino Zappacosta: An Enlightened Mentor and Educator.

With this article, the authors aim to honor the memory of Serafino Zappacosta, who had been their mentor during the early years of their career in science. The authors discuss how the combination of Serafino Zappacosta's extraordinary commitment to teaching and passion for science created a fostering educational environment that led to the creation of the "Ruggero Ceppellini Advanced School of Immunology." The review also illustrates how the research on the MHC and the inspirational scientific context in the Zappacosta's laboratory influenced the authors' early scientific interests, and subsequent professional work as immunologists.