Saccharomyces cerevisiae oral immunization in mice using multi-antigen of the African swine fever virus elicits a robust immune response.

African swine fever virus (ASFV) is one of the most complex viruses. ASFV is a serious threat to the global swine industry because no commercial vaccines against this virus are currently available except in Vietnam. Moreover, ASFV is highly stable in the environment and can survive in water, feed, and aerosols for a long time. ASFV is transmitted through the digestive and respiratory tract. Mucosal immunity is the first line of defense against ASFV. Saccharomyces cerevisiae (SC), which has been certified by the U.S. Food and Drug Administration and has a generally recognized as safe status in the food industry, was used for oral immunization in this study. ASFV antigens were effectively expressed in recombinant SC strains with high DNA copy numbers and stable growth though surface display technology and chromosome engineering (δ-integration). The recombinant SC strains containing eight ASFV antigens-KP177R, E183L, E199L, CP204L, E248R, EP402R, B602L, and B646L- induced strong humoral and mucosal immune responses in mice. There was no antigenic competition, and these antigens induced Th1 and Th2 cellular immune responses. Therefore, the oral immunization strategy using recombinant SC strains containing multiple ASFV antigens demonstrate potential for future testing in swine, including challenge studies to evaluate its efficacy as a vaccine against ASFV.

A Novel Whole Yeast-Based Subunit Oral Vaccine Against Eimeria tenella in Chickens.

Cheap, easy-to-produce oral vaccines are needed for control of coccidiosis in chickens to reduce the impact of this disease on welfare and economic performance. Saccharomyces cerevisiae yeast expressing three Eimeria tenella antigens were developed and delivered as heat-killed, freeze-dried whole yeast oral vaccines to chickens in four separate studies. After vaccination, E. tenella replication was reduced following low dose challenge (250 oocysts) in Hy-Line Brown layer chickens (p<0.01). Similarly, caecal lesion score was reduced in Hy-Line Brown layer chickens vaccinated using a mixture of S. cerevisiae expressing EtAMA1, EtIMP1 and EtMIC3 following pathogenic-level challenge (4,000 E. tenella oocysts; p<0.01). Mean body weight gain post-challenge with 15,000 E. tenella oocysts was significantly increased in vaccinated Cobb500 broiler chickens compared to mock-vaccinated controls (p<0.01). Thus, inactivated recombinant yeast vaccines offer cost-effective and scalable opportunities for control of coccidiosis, with relevance to broiler production and chickens reared in low-and middle-income countries (LMICs).

Evaluation and selection of potent fluorescent immunosensors by combining fluorescent peptide and nanobodies displayed on yeast surface.

Quenchbody (Q-body) is a quench-based fluorescent immunosensor labeled with fluorescent dye(s) near the antigen-binding site of an antibody. Q-bodies can detect a range of target molecules rapidly and directly. However, because Q-bodies show different antigen responses depending on the antibody used, time-consuming optimization of the Q-body structure is often necessary, and a high-throughput screening method for discriminating and selecting good Q-bodies is required. Here, we aimed to develop a molecular display method of nanobody-based "mini Q-bodies" by combining yeast surface display and coiled-coil forming E4/K4 peptide-based fluorescence labeling. As a result, the yeast-displayed mini Q-body recognizing the anti-cancer agent methotrexate (MTX) showed significant quenching and MTX-dependent dequenching on cells. To demonstrate the applicability of the developed method to select highly responsive mini Q-bodies, a small nanobody library consisting of 30 variants that recognize human serum albumin was used as a model. The best variant, showing a 2.4-fold signal increase, was obtained through selection by flow cytometry. Furthermore, the same nanobody prepared from Escherichia coli also worked as a mini Q-body after dye labeling. The described approach will be applied to quickly obtain well-behaved Q-bodies and other fluorescent biosensors for various targets through directed evolutionary approaches.

Polyethyleneimine-complexed charge-reversed yeast cell walls for the enhanced oral delivery of pseudovirus-based antigens.

Oral vaccination has wide applicability in poor areas, particularly during the epidemic periods of infectious diseases. However, successful oral antigen delivery and immune activation remain highly challenging due to the instability of vaccines in gastric acid and the low capture of antigens in the intestine. Here, we present a facile approach for the preparation of a robust oral delivery system via encapsulating antigen-carrying pseudoviruses inside positively charged polyethyleneimine-modified yeast capsules (P-YC). By virtue of the physical barrier role and surface ß-glucan of YC, encapsulated pseudoviruses can be protected from gastric insult and delivered into Peyer's patches via uptake mediated by microfold cells located in the intestinal epithelium. Given the ability to carry diverse antigens, the enhanced oral delivery of pseudoviruses achieved by P-YC provides a versatile platform for the development of various oral vaccines.

SPINK7 Recognizes Fungi and Initiates Hemocyte-Mediated Immune Defense Against Fungal Infections.

Serine protease inhibitors of Kazal-type (SPINKs) were widely identified in vertebrates and invertebrates, and played regulatory roles in digestion, coagulation, and fibrinolysis. In this study, we reported the important role of SPINK7 in regulating immune defense of silkworm, Bombyx mori. SPINK7 contains three Kazal domains and has 6 conserved cysteine residues in each domain. Quantitative real-time PCR analyses revealed that SPINK7 was exclusively expressed in hemocytes and was upregulated after infection with two fungi, Saccharomyces cerevisiae and Candida albicans. Enzyme activity inhibition test showed that SPINK7 significantly inhibited the activity of proteinase K from C. albicans. Additionally, SPINK7 inhibited the growth of three fungal spores, including S. cerevisiae, C. albicans, and Beauveria bassiana. The pathogen-associated molecular patterns (PAMP) binding assays suggested that SPINK7 could bind to ß-D-glucan and agglutinate B. bassiana and C. albicans. In vitro assays were performed using SPINK7-coated agarose beads, and indicated that SPINK7 promoted encapsulation and melanization of agarose beads by B. mori hemocytes. Furthermore, co-localization studies using immunofluorescence revealed that SPINK7 induced hemocytes to aggregate and entrap the fungi spores of B. bassiana and C. albicans. Our study revealed that SPINK7 could recognize fungal PAMP and induce the aggregation, melanization, and encapsulation of hemocytes, and provided valuable clues for understanding the innate immunity and cellular immunity in insects.

Yeasts in nanotechnology-enabled oral vaccine and gene delivery.

Oral vaccine and gene delivery systems must be engineered to withstand several different physiological environments, such as those present in the oral cavity, stomach, and jejunum, each of which exhibits varying pH levels and enzyme distributions. Additionally, these systems must be designed to ensure appropriate gastrointestinal absorption and tissue/cellular targeting properties. Yeasts-based delivery vehicles are excellent candidates for oral vaccine and oral gene therapies as many species possess cellular characteristics resulting in enhanced resistance to the harsh gastrointestinal (GI) environment and facilitated passage across the mucosal barrier. Yeast capsules can stimulate and modulate host immune responses, which is beneficial for vaccine efficacy. In addition, recombinant modification of yeasts to express cell penetrating proteins and injection mechanisms along with efficient cell adhering capabilities can potentially improve transfection rates of genetic material. In this literature review, we present evidence supporting the beneficial role yeast-based delivery systems can play in increasing the efficacy of oral administration of vaccines and gene therapies.

Enhanced immune response by vacuoles isolated from Saccharomyces cerevisiae in RAW 264.7 macrophages.

Vacuoles are membrane vesicles in eukaryotic cells, the digestive system of cells that break down substances absorbed outside the cell and digest the useless components of the cell itself. Researches on anticancer and intractable diseases using vacuoles are being actively conducted. The practical application of the present study to animals requires the determination of the biocompatibility of vacuole. In the present study, we evaluated the effects of vacuoles isolated from Saccharomyces cerevisiae in RAW 264.7 cells. This showed a significant increase in the production of nitric oxide (NO) produced by macrophage activity. Using Reactive Oxygen Species (ROS) assay, we identified that ROS is increased in a manner dependent on vacuole concentration. Western blot analysis showed that vacuole concentration-dependently increased protein levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2). Therefore, iNOS expression was stimulated to induce NO production. In addition, pro-inflammatory cytokines levels promoted, such as interleukin (IL) 6 (IL-6) and tumor necrosis factor (TNF) α (TNF-α). In summary, vacuoles activate the immune response of macrophages by promoting the production of immune-mediated transporters NO, ROS, and pro-inflammatory cytokines.

Fecal Bacteria Implicated in Biofilm Production Are Enriched and Associate to Gastrointestinal Symptoms in Patients With APECED - A Pilot Study.

Backgrounds and Aims: APECED is a rare autoimmune disease caused by mutations in the Autoimmune Regulator gene. A significant proportion of patients also have gastrointestinal symptoms, including malabsorption, chronic diarrhea, and obstipation. The pathological background of the gastrointestinal symptoms remains incompletely understood and involves multiple factors, with autoimmunity being the most common underlying cause. Patients with APECED have increased immune responses against gut commensals. Our objective was to evaluate whether the intestinal microbiota composition, predicted functions or fungal abundance differ between Finnish patients with APECED and healthy controls, and whether these associate to the patients' clinical phenotype and gastrointestinal symptoms. Methods: DNA was isolated from fecal samples from 15 patients with APECED (median age 46.4 years) together with 15 samples from body mass index matched healthy controls. DNA samples were subjected to analysis of the gut microbiota using 16S rRNA gene amplicon sequencing, imputed metagenomics using the PICRUSt2 algorithm, and quantitative PCR for fungi. Extensive correlations of the microbiota with patient characteristics were determined. Results: Analysis of gut microbiota indicated that both alpha- and beta-diversity were altered in patients with APECED compared to healthy controls. The fraction of Faecalibacterium was reduced in patients with APECED while that of Atopobium spp. and several gram-negative genera previously implicated in biofilm formation, e.g. Veillonella, Prevotella, Megasphaera and Heamophilus, were increased in parallel to lipopolysaccharide (LPS) synthesis in imputed metagenomics. The differences in gut microbiota were linked to patient characteristics, especially the presence of anti-Saccharomyces cerevisiae antibodies (ASCA) and severity of gastrointestinal symptoms. Conclusions: Gut microbiota of patients with APECED is altered and enriched with predominantly gram-negative bacterial taxa that may promote biofilm formation and lead to increased exposure to LPS in the patients. The most pronounced alterations in the microbiota were associated with more severe gastrointestinal symptoms.

Expression of antibody fragments in Saccharomyces cerevisiae strains evolved for enhanced protein secretion.

Monoclonal antibodies, antibody fragments and fusion proteins derived thereof have revolutionized the practice of medicine. Major challenges faced by the biopharmaceutical industry are however high production costs, long processing times and low productivities associated with their production in mammalian cell lines. The yeast Saccharomyces cerevisiae, a well-characterized eukaryotic cell factory possessing the capacity of post-translational modifications, has been industrially exploited as a secretion host for production of a range of products, including pharmaceuticals. However, due to the incompatible surface glycosylation, few antibody molecules have been functionally expressed in S. cerevisiae. Here, three non-glycosylated antibody fragments from human and the Camelidae family were chosen for expression in a S. cerevisiae strain (HA) previously evolved for high α-amylase secretion. These included the Fab fragment Ranibizumab (Ran), the scFv peptide Pexelizumab (Pex), and a nanobody consisting of a single V-type domain (Nan). Both secretion and biological activities of the antibody fragments were confirmed. In addition, the secretion level of each protein was compared in the wild type (LA) and two evolved strains (HA and MA) with different secretory capacities. We found that the secretion of Ran and Nan was positively correlated with the strains' secretory capacity, while Pex was most efficiently secreted in the parental strain. To investigate the mechanisms for different secretion abilities in these selected yeast strains for the different antibody fragments, RNA-seq analysis was performed. The results showed that several bioprocesses were significantly enriched for differentially expressed genes when comparing the enriched terms between HA.Nan vs. LA.Nan and HA.Pex vs. LA.Pex, including amino acid metabolism, protein synthesis, cell cycle and others, which indicates that there are unique physiological needs for each antibody fragment secretion.

The superposition anti-viral activity of porcine tri-subtype interferon expressed by Saccharomyces cerevisiae.

Interferon (IFN)-mediated antiviral responses are central to host defense against viral infection. Porcine viral infection has emerged as a serious hazard for the pig industry. The construction of an engineered Saccharomyces cerevisiae strain that efficiently produces porcine IFN has demonstrated several advantages. It can be easily fed to pigs, which helps in reducing antibiotic residues in pork and improve meat quality. In this study, the stable expression of several porcine IFN molecules (pIFN-α1, pIFN-ß, pIFN-λ1, pIFN-λ1-ß, pIFN-λ1-ß-α1) were determined using an engineered S. cerevisiae system. With the YeastFab assembly method, the complete transcriptional units containing promoter (GPD), secretory peptide (α-mating factor), target gene (IFN) and terminator (ADH1) were successfully constructed using the characteristics of type II restriction endonuclease, and then integrated into the chromosomes Ⅳ and XVI of ST1814 yeast host strain, respectively. The expression kinetics of recombinant pIFNs were further analyzed. Synergism in the expression level of IFN receptor, antiviral protein, and viral loading was observed in viral-cell infection model treated with different porcine IFN subtypes. The porcine reproductive and respiratory syndrome viral load and antibody titer in serum decreased significantly after oral administration of IFN expression yeast fermentation broth. These findings indicate the potential efficacy of multi-valent pIFNs expressing S. cerevisiae as a potent feed material to prevent viral infections of pigs.

Yeast display of MHC-II enables rapid identification of peptide ligands from protein antigens (RIPPA).

CD4+ T cells orchestrate adaptive immune responses via binding of antigens to their receptors through specific peptide/MHC-II complexes. To study these responses, it is essential to identify protein-derived MHC-II peptide ligands that constitute epitopes for T cell recognition. However, generating cells expressing single MHC-II alleles and isolating these proteins for use in peptide elution or binding studies is time consuming. Here, we express human MHC alleles (HLA-DR4 and HLA-DQ6) as native, noncovalent αß dimers on yeast cells for direct flow cytometry-based screening of peptide ligands from selected antigens. We demonstrate rapid, accurate identification of DQ6 ligands from pre-pro-hypocretin, a narcolepsy-related immunogenic target. We also identify 20 DR4-binding SARS-CoV-2 spike peptides homologous to SARS-CoV-1 epitopes, and one spike peptide overlapping with the reported SARS-CoV-2 epitope recognized by CD4+ T cells from unexposed individuals carrying DR4 subtypes. Our method is optimized for immediate application upon the emergence of novel pathogens.

A humoral factor, hemolymph proteinase 8, elicits a cellular defense response of nodule formation in Bombyx mori larvae in association with recognition by C-type lectins.

Previously, we found that nodule formation, a cellular defense response in insects, is regulated by humoral factors called C-type lectins in the hemolymph. To elucidate the factors that elicit nodule formation following the recognition of microorganisms by C-type lectins, a reproducible quantitative in vitro assay system was constructed. Then, using this system, the inhibitory activities of antisera raised against hemolymph proteases (HPs), serine protease homologues (SPHs), and pathogen-associated molecular pattern (PAMP)-recognition proteins were assessed. Among the antisera raised against HP and SPH, only that against HP8, a terminal proteinase that activates Spätzle, consistently inhibited in-vitro nodule-like aggregate formation in all three tested microorganisms, Micrococcus luteus, Escherichia coli, and Saccharomyces cerevisiae. Antisera raised against C-type lectins, BmLBP, and BmMBP also inhibited nodule-like aggregate formation, while those against ß-glucan recognition proteins and peptidoglycan recognition protein-S1 did not. Microorganisms pretreated with hemolymph, which contains HP8 and C-type lectins, also induced nodule-like aggregate formation, indicating that nodulation factors are present on microbial cells. Furthermore, antisera raised against HP8, BmLBP, and BmMBP showed inhibitory activities in the in vivo nodule formation system using Bombyx mori larvae. Thus, two humoral factors in the hemolymph of B. mori larvae, BmHP8 and C-type lectins, were found to play significant roles in eliciting the cellular defense response of nodule formation.

High immune efficacy against different avian influenza H5N1 viruses due to oral administration of a Saccharomyces cerevisiae-based vaccine in chickens.

A safe and effective vaccine is the best way to control large-scale highly pathogenic avian influenza virus (HPAI) A (H5N1) outbreaks. Saccharomyces cerevisiae (S. cerevisiae) is an ideal mucosal delivery vector for vaccine development, and we have previously shown that conventional administration of a S. cerevisiae-based vaccine (EBY100/pYD1-HA) via injection led to protection against the homologous H5N1 virus in a mouse model. Because the diameter of S. cerevisiae is approximately 10 µm, which results in a severe inflammation by injection route, therefore, oral administration is a more suitable approach for EBY100/pYD1-HA conferring protection in poultry. We extended our work by evaluating the immunogenicity and protective efficacy of oral vaccination with EBY100/pYD1-HA in the chicken model. Oral immunization with EBY100/pYD1-HA could induce robust serum IgG, mucosal IgA and cellular immune responses. Importantly, EBY100/pYD1-HA provided protection against challenges with a homologous and a heterologous H5N1 viruses. These findings suggest that EBY100/pYD1-HA, a promising H5N1 oral vaccine candidate, can avoid potential reassortment of other avian influenza viruses in oral administration of live virus vaccines and overcome the limitations of conventional injection routes. Importantly, this platform will be able to provide opportunities for broader applications in poultry during HPAI A (H5N1) outbreaks.

Molecular characterization and biological effect of a C-type lectin receptor in Qihe crucian carp, Carassius auratus.

C-type lectin receptors, as the important members of pattern-recognition receptors, play the crucial roles in the innate immune system, which discriminate self and non-self by recognizing and binding the carbohydrates on the surface of microorganism. In this study, we identified a C-type lectin receptor gene in Qihe crucian carp Carassius auratus (named as CaCLR). The full-length cDNA of CaCLR was composed of 1130 bp, with a 226 bp 5'-untranslated region (UTR), a 792 bp ORF encoding a 263aa protein, and a 112 bp 3'-UTR with a polyadenylation signal sequence AATAAA and a poly (A) tail. The predicted amino acid sequence of CaCLR is a single transmembrane receptor with a typical carbohydrate recognition domain (CRD) at its C-terminus. With regard to the mRNA transcript of CaCLR, it was ubiquitously detected in the tested tissues, among which it was the most abundant in head kidney. The temporal expressions of CaCLR were obviously up-regulated in liver, spleen, kidney, and head kidney after Aeromonas hydrophila and poly I: C challenge, respectively, and the patterns of expression changes were in a time-depended manner. The recombinant CaCLR (rCaCLR) purified from Escherichia coli BL21 (DE3), exhibited strong binding ability with lipopolysaccharide (LPS), peptidoglycan (PGN), ß-Glucan, and Mannan, as well as five microorganisms including fungus (Saccharomyces cerevisiae), Gram-negative bacteria (A. hydrophila, E. coli and Vibrio anguillarum), and Gram-positive bacteria (Micrococcus lysodeikticus). In the presence of rCaCLR, the eliminating capacity against A. hydrophila could be enhanced in C. auratus. Taken together, CaCLR is involved in the antibacterial defense in C. auratus.

Serological markers facilitate the diagnosis of Crohn's disease.

Background and aim: The diagnosis of Crohn's disease (CD) is challenging. Ongoing search for biomarkers to facilitate the diagnosis is a worthwhile endeavor. The aim of this study was to explore the role of serological markers in the diagnosis of CD at an inflammatory bowel disease (IBD) referral center.Methods: This was a retrospective study including 196 suspected CD patients. The expression of ASCA-IgG, ASCA-IgA, AYMA-IgG, AYCA-IgA, FI2Y-IgG, and pANCA in the patient's serum was determined by enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescence (IF).Results: ASCA was a relatively specific marker for CD (p = 0.0005), but not AYMA-IgG, AYCA-IgA, F12Y-IgG (p = 0.5936, 0.7974, 0.1085, respectively). However, a high sensitivity of 96.77% (95% CI 90.19%-99.83%) was noted for ASCA+/FI2Y+ to identify CD patients among the suspected cases, albeit with low PPV. The more combinations of serological markers, the higher sensitivity, and NPV. No correlation was found between the age of onset or disease location and the expression of ASCA, AYMA, AYCA, FI2Y, or pANCA. There was no significant difference between the expression of ASCA and the disease behavior at diagnosis (p = 0.3307). However, a decreased proportion of AYMA+ CD patients was found in those who received surgery compared with their non-surgical counterparts (p = 0.0488).Conclusions: ASCA was found to be the most accurate serological marker for the differential diagnosis of CD. Combinations of ASCA, AYMA, AYCA, and FI2Y improved diagnostic accuracy of CD.

Saccharomyces cerevisiae as a skin physiology, pathology, and treatment model.

Saccharomyces cerevisiae serves as a useful model in experimental biology. Within dermatology research, several studies have examined this organism's role in skin physiology, pathology, and treatment. Saccharomyces cerevisiae has been used to explore the mechanisms of melanogenesis as its extract inhibits key enzymes involved in melanogenesis and melanosome transfer. Additionally, the lack of probiotic intestinal Saccharomyces cerevisiae has been associated with psoriasis, potentially related to the anti-inflammatory effects of the yeast. Furthermore, antibodies against Saccharomyces cerevisiae have been observed in skin conditions, including atopic dermatitis. Saccharomyces cerevisiae may even cause skin infections, such as septic emboli in a patient with acute myelogenous leukemia. Lastly, Saccharomyces cerevisiae has potential use in vaccine development against melanoma and is utilized to study various treatment modalities such as zinc pyrithione, an ingredient often used in anti-dandruff shampoo.

Selenization of S. cerevisiae increases its protective potential in experimental autoimmune encephalomyelitis by triggering an intestinal immunomodulatory loop.

Multiple sclerosis is an autoimmune disease that affects the myelinated central nervous system (CNS) neurons and triggers physical and cognitive disabilities. Conventional therapy is based on disease-modifying drugs that control disease severity but can also be deleterious. Complementary medicines have been adopted and evidence indicates that yeast supplements can improve symptoms mainly by modulating the immune response. In this investigation, we evaluated the therapeutic potential of Saccharomyces cerevisiae and its selenized derivative (Selemax) in experimental autoimmune encephalomyelitis (EAE). Female C57BL/6 mice submitted to EAE induction were orally supplemented with these yeasts by gavage from day 0 to day 14 after EAE induction. Both supplements determined significant reduction in clinical signs concomitantly with diminished Th1 immune response in CNS, increased proportion of Foxp3+ lymphocytes in inguinal and mesenteric lymph nodes and increased microbiota diversity. However, Selemax was more effective clinically and immunologically; it reduced disease prevalence more sharply, increased the proportion of CD103+ dendritic cells expressing high levels of PD-L1 in mesenteric lymph nodes and reduced the intestinal inflammatory process more strongly than S. cerevisiae. These results suggest a clear gut-brain axis modulation by selenized S. cerevisiae and suggest their inclusion in clinical trials.

Characterization of a novel yeast phase-specific antigen expressed during in vitro thermal phase transition of Talaromyces marneffei.

Talaromyces marneffei is a dimorphic fungus that has emerged as an opportunistic pathogen particularly in individuals with HIV/AIDS. Since its dimorphism has been associated with its virulence, the transition from mold to yeast-like cells might be important for fungal pathogenesis, including its survival inside of phagocytic host cells. We investigated the expression of yeast antigen of T. marneffei using a yeast-specific monoclonal antibody (MAb) 4D1 during phase transition. We found that MAb 4D1 recognizes and binds to antigenic epitopes on the surface of yeast cells. Antibody to antigenic determinant binding was associated with time of exposure, mold to yeast conversion, and mammalian temperature. We also demonstrated that MAb 4D1 binds to and recognizes conidia to yeast cells' transition inside of a human monocyte-like THP-1 cells line. Our studies are important because we demonstrated that MAb 4D1 can be used as a tool to study T. marneffei virulence, furthering the understanding of the therapeutic potential of passive immunity in this fungal pathogenesis.

Harnessing antifungal immunity in pursuit of a Staphylococcus aureus vaccine strategy.

Staphylococcus aureus (S. aureus) is one of the most common bacterial infections worldwide, and antibiotic resistant strains such as Methicillin-Resistant S. aureus (MRSA) are a major threat and burden to public health. MRSA not only infects immunocompromised patients but also healthy individuals and has rapidly spread from the healthcare setting to the outside community. However, all vaccines tested in clinical trials to date have failed. Immunocompromised individuals such as patients with HIV or decreased levels of CD4+ T cells are highly susceptible to S. aureus infections, and they are also at increased risk of developing fungal infections. We therefore wondered whether stimulation of antifungal immunity might promote the type of immune responses needed for effective host defense against S. aureus. Here we show that vaccination of mice with a fungal ß-glucan particle (GP) loaded with S. aureus antigens provides protective immunity to S. aureus. We generated glucan particles loaded with the four S. aureus proteins ClfA, IsdA, MntC, and SdrE, creating the 4X-SA-GP vaccine. Vaccination of mice with three doses of 4X-SA-GP promoted protection in a systemic model of S. aureus infection with a significant reduction in the bacterial burden in the spleen and kidneys. 4X-SA-GP vaccination induced antigen-specific Th1 and Th17 CD4+ T cell and antibody responses and provided long-term protection. This work suggests that the GP vaccine system has potential as a novel approach to developing vaccines for S. aureus.

A Case Report of Sequential Use of a Yeast-CEA Therapeutic Cancer Vaccine and Anti-PD-L1 Inhibitor in Metastatic Medullary Thyroid Cancer.

Medullary thyroid cancer (MTC) accounts for ~4% of all thyroid malignancies. MTC derives from the neural crest and secretes calcitonin (CTN) and carcinoembryonic antigen (CEA). Unlike differentiated thyroid cancer, MTC does not uptake iodine and I-131 RAI (radioactive iodine) treatment is ineffective. Patients with metastatic disease are candidates for FDA-approved agents with either vandetanib or cabozantinib; however, adverse effects limit their use. There are ongoing trials exploring the role of less toxic immunotherapies in patients with MTC. We present a 61-year-old male with the diagnosis of MTC and persistent local recurrence despite multiple surgeries. He was started on sunitinib, but ultimately its use was limited by toxicity. He then presented to the National Cancer Institute (NCI) and was enrolled on a clinical trial with heat-killed yeast-CEA vaccine (NCT01856920) and his calcitonin doubling time improved in 3 months. He then came off vaccine for elective surgery. After surgery, his calcitonin was rising and he enrolled on a phase I trial of avelumab, a programmed death-ligand 1 (PD-L1) inhibitor (NCT01772004). Thereafter, his calcitonin decreased > 40% on 5 consecutive evaluations. His tumor was subsequently found to express PD-L1. CEA-specific T cells were increased following vaccination, and a number of potential immune-enhancing changes were noted in the peripheral immunome over the course of sequential immunotherapy treatment. Although calcitonin declines do not always directly correlate with clinical responses, this response is noteworthy and highlights the potential for immunotherapy or sequential immunotherapy in metastatic or unresectable MTC.