Whole-cell Vaccine Research Articles

Lack of evidence supporting a role of IFN-β and TGF-β in differential polarization of Bordetella pertussis specific-T cell responses

Bordetella Pertussis (BP) vaccine-induced immunity is waning worldwide despite excellent vaccine coverage. Replacement of the whole-cell inactivated vaccine (wP) by an acellular subunit vaccine (aP) is thought to play a major role and to be associated with the recurrence of whooping cough. Previously, we detected that the polarization towards a Th2 and Th1/Th17 response in aP and wP vaccinees, respectively, persists upon aP boosting in adolescents and adults. Additionally, IL-9 and TGF-β were found to be up-regulated in aP-primed donors and network analysis further identified IFN-β as a potential upstream regulator of IL-17 and IL-9. Based on these findings, we hypothesized that IFN-β produced following aP vaccination may lead to increased IL-9 and decreased IL-17 production. Also, due to the well characterized role of TGF-β in both Th17 and Th9 differentiation, we put forth that TGF-β addition to BP-stimulated CD4 + T cells might modulate IL-17 and IL-9 production. To test this hypothesis, we stimulated in vitro cultures of PBMC or isolated naive CD4 + T cells from aP vs wP donors with a pool of BP epitopes and assessed the effect of IFN-β or TGF-β in proliferative responses as well as in the cytokine secretion of IL-4, IL-9, IL-17, and IFN-γ. IFN-β reduced BP-specific proliferation in PBMC as well as cytokine production but increased IL-9, IL-4, and IFN-γ cytokines in naïve CD4 + T cells. These effects were independent of the childhood vaccination received by the donors. Similarly, TGF-β reduced BP-specific proliferation in PBMC but induced proliferation in naïve CD4 + T cells. However, stimulation was associated with a generalized inhibition of cytokine production regardless of the original aP or wP vaccination received by the donors. Our study suggests that key T cell functions such as cytokine secretion are under the control of antigen stimulation and environmental cues but molecular pathways different than the ones investigated here might underlie the long-lasting differential cytokine production associated with aP- vs wP-priming in childhood vaccination.

Chemoimmunotherapy for the treatment of prostate cancer: Insights from mathematical modelling

In this article, we aim to apply mathematical modelling by means of eight first-order ordinary differential equations to investigate the response of prostate cancer and the immune system to an allogenic whole-cell cancer vaccine and metronomic chemotherapy. Research to formulate our mathematical model was based on the clinical study developed by Kronik et al. In order to provide an accurate mathematical description of the global dynamics of our system, we apply the Localization of Compact Invariant Sets method to calculate upper bounds for all cell populations and define the so-called localizing domain. Further, by Lyapunov’s direct method and LaSalle’s invariance principle we study the asymptotic stability of the system and determine conditions for a global attractor existence. The latter allow us to establish sufficient conditions to ensure prostate cancer elimination by applying the metronomic chemotherapy treatment. In silico experiments were performed by setting six different initial tumor sizes and considering two cases: one with the sole administration of metronomic chemotherapy, and another with the combined application of chemoimmunotherapy. Numerical simulations are consistent with our analytical results as they successfully illustrate that the prostate cancer cells population is eliminated in both considered cases.

Parent reported outcomes to measure satisfaction, acceptability, and daily life impact after vaccination with whole-cell and acellular pertussis vaccine in Chile

AimTo compare parental satisfaction and impact on daily life among parents of children receiving whole-cell pentavalent + oral polio vaccine (Arm1) with an acellular hexavalent vaccine (Hexaxim; Arm2). MethodsSelf-administered electronic questionnaire at vaccination and one week later in six community health clinics of metropolitan Santiago, Chile, exploring parent-reported outcomes on satisfaction, acceptability, and impact on daily life after immunization. Univariate and multivariate analyses were conducted to determine differences in the responses in both groups (α = 0.05). ResultsThe study enrolled 800 participants and 65% (222 in Arm1, 296 in Arm2) were included for according-to-protocol analysis. Demographic characteristics were comparable, except for a higher proportion of mothers answering the questionnaire at the 6-month visit. Regardless of the study arm, parental knowledge and perception of the immunization practices were good, and there were no differences in vaccination experiences in the prior 5 years. However, satisfaction with vaccination and intention to vaccinate were statistically significantly higher in Arm2 after the 6-month visit. Also, more parents in Arm2 reported no disruption in several aspects of the everyday activities of the parent, the child, and other children in the household. Parents in Arm2 were more likely to be satisfied with the vaccine received (OR 2.82; 95% CI, 1.22–7.07); return for other vaccine dose (OR 2.62; 95% CI, 1.45–4.84); follow a healthcare professional recommendation (OR 2.24; 95% CI, 1.57–3.21); and, to be confident that the vaccine will not disrupt the family’s daily routine (OR 1.89; 95% CI, 1.32–2.71). ConclusionsOverall, satisfaction, intention for future vaccination, and lower impact on the family daily routine were significantly better in the group receiving the hexavalent vaccine. We also found that health care providers’ recommendations to vaccinate and participants’ access to health services were important factors favoring immunization.

The second coming: The comeback of the live vaccines.

Live vaccines are gradually replaced by protein-based vaccines given the latter's better safety. But live vaccines seem to be more efficient via stimulation of the innate immune system.

Transversal sero-epidemiological study of Bordetella pertussis in Tehran, Iran.

Pertussis remains endemic despite high vaccine coverage in infants and toddlers. Pertussis vaccines confer protection but immunity wanes overtime and boosters are needed in a lifetime. Iran, eligible for the Expanded Program on Immunization that includes the primary immunization, implemented two additional booster doses using a whole-cell vaccine (wPV) at 18 months-old and about 6 years-old. Duration of protection induced by the wPVs currently in use and their impact as pre-school booster are not well documented. This study aimed at assessing vaccination compliance and at estimating the duration of protection conferred by vaccination with wPV in children aged < 15 years in Tehran, Iran. Detailed information on vaccination history and capillary blood samples were obtained from 1047 children aged 3-15 years who completed the 3 doses-primary pertussis immunization, in Tehran. Anti-pertussis toxin IgG levels were quantified by ELISA. Compliance was very high with 93.3% of children who received the three primary and 1st booster doses in a timely manner. Timeliness of the 2nd booster was lower (63.3%). Rate of seropositive samples continuously and significantly increased from 1-2 to 5-6 years after 1st booster attaining 30.4% of children exhibiting serological sign of recent contact with B. pertussis. Second booster dating back 1 or 2 years was associated with high antibody titers, which significantly decreased within 3 years from injection. Among children who received 2nd booster injection more than 2 years before serum analysis, seroprevalence of pertussis infection was 8.4% and seropositivity rate was higher from the 10 years-old group. Seropositivity in children aged 6-7 years with no 2nd booster supports the need for a vaccination at that age. Adolescent booster may also be considered.

Nanoparticle Delivery of MnO2 and Antiangiogenic Therapy to Overcome Hypoxia-Driven Tumor Escape and Suppress Hepatocellular Carcinoma.

Antiangiogenic therapy is widely administered in many cancers, and the antiangiogenic drug sorafenib offers moderate benefits in advanced hepatocellular carcinoma (HCC). However, antiangiogenic therapy can also lead to hypoxia-driven angiogenesis and immunosuppression in the tumor microenvironment (TME) and metastasis. Here, we report the synthesis and evaluation of NanoMnSor, a tumor-targeted, nanoparticle drug carrier that efficiently codelivers oxygen-generating MnO2 and sorafenib into HCC. We found that MnO2 not only alleviates hypoxia by catalyzing the decomposition of H2O2 to oxygen but also enhances pH/redox-responsive T1-weighted magnetic resonance imaging and drug-release properties upon decomposition into Mn2+ ions in the TME. Moreover, macrophages exposed to MnO2 displayed increased mRNA associated with the immunostimulatory M1 phenotype. We further show that NanoMnSor treatment leads to sorafenib-induced decrease in tumor vascularization and significantly suppresses primary tumor growth and distal metastasis, resulting in improved overall survival in a mouse orthotopic HCC model. Furthermore, NanoMnSor reprograms the immunosuppressive TME by reducing the hypoxia-induced tumor infiltration of tumor-associated macrophages, promoting macrophage polarization toward the immunostimulatory M1 phenotype, and increasing the number of CD8+ cytotoxic T cells in tumors, thereby augmenting the efficacy of anti-PD-1 antibody and whole-cell cancer vaccine immunotherapies. Our study demonstrates the potential of oxygen-generating nanoparticles to deliver antiangiogenic agents, efficiently modulate the hypoxic TME, and overcome hypoxia-driven drug resistance, thereby providing therapeutic benefit in cancer.

Abstract 1072: Lymphangiogenesis-inducing vaccines for melanoma treatment elicit potent tumor-specific T cell immunity and long-term tumor control

Abstract We recently demonstrated that the growth of tumor-associated lymphatic vessels (lymphangiogenesis) promotes immune infiltration in melanoma (Lund et al. 2016) and increases T cell activation following immunotherapy, resulting in strongly enhanced therapeutic efficacy (Fankhauser et a. 2017). On the other hand, tumor lymphangiogenesis is known to support cancer cell dissemination through the lymphatic route and metastasis formation. Therefore, inducing lymphangiogesis within the tumor site to boost anti-tumor immunity appears as a potentially unsafe therapeutic approach since it might also result in increased tumor metastatic potential. In this study, we sought to exploit the immune-promoting functions of lymphatics remotely from the tumor by developing a lymphangiogenesis-inducing cancer vaccine that mimics the microenvironment of a lymphangiogenic tumor. Whole-cell lymphangiogenic vaccines were generated using lethally irradiated mouse melanoma cells genetically engineered to overexpress the lymphatic-specific growth factor VEGFC, in combination with topical immune adjuvants (VEGFC vax). VEGFC vax was compared to control vaccines with identical formulation but lacking VEGFC overexpression and to a vaccine composed of GM-CSF-overexpressing tumor cells (GVAX), a vaccination approach that has been widely used in clinical studies. Upon intradermal injection in mice, VEGFC vax induced extensive lymphangiogenesis at the vaccine site and increased antigen transport to vaccine-draining lymph nodes. Analogously to what observed in lymphangiogenic tumors, we found that naïve T cells significantly infiltrated lymphangiogenic vaccine sites and could undergo in situ priming and activation. Using VEGFC vaccines containing either B16-F10 mouse melanoma cells or melanoma cells obtained from tumors growing in BrafV600EPten−/- transgenic mice, we demonstrated that VEGFC vax elicits a potent tumor-specific T cell response. Importantly, VEGFC vaccination resulted in the mounting of a broad T cell immunity directed against multiple B16-F10-associated antigens. In both prophylactic and therapeutic settings, VEGFC vax provided effective tumor control and long-term protection against B16-F10 melanomas. In the present study we introduced for the first time the concept of lymphangenesis induction as a tool to increase cancer vaccine potency and we provided a proof of efficacy of lymphangiogenic vaccines in preclinical melanoma models. Citation Format: Maria Stella Sasso, Nikolaos Mitrousis, Sylvie Hauert, Priscilla S. Briquez, Yue Wang, Jun Ishihara, Jeffrey A. Hubbell, Melody A. Swartz. Lymphangiogenesis-inducing vaccines for melanoma treatment elicit potent tumor-specific T cell immunity and long-term tumor control [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1072.

Whole-cell pertussis vaccine in early infancy for the prevention of allergy

Whole-cell pertussis vaccine in early infancy for the prevention of allergy

Study of the Immunobiological Properties of Surface Protein-Containing Antigens of Streptococcus pneumoniae Serotype 6B

Relevance . Prevention of pneumococcal infection in the Russian Federation is carried out by imported polysaccharide and conjugated vaccines without taking into account the circulation of clinically significant isolates, which leads to the growth of previously rare genetic lines and serotypes that are not part of the vaccines, and does not protect against the carriage. Serotype-independent vaccines are being developed based on protective proteins and variants of whole-cell vaccines capable of providing a cross-protective effect against pneumococcus. Objective . Investigation of the protective properties of surface protein-containing antigens isolated from the immunogenic strain S. pneumoniae 6B No. 296 and their influence on key effectors of innate immunity. Materials and methods . S. pneumoniae 6B No. 296 was cultured in a semi-synthetic medium under stationary conditions at 5% CO 2 for 5-7 hours. From inactivated bacterial cells, the experimental initial protein-containing preparation was obtained - an aqueous extract (A/E), from which a 30-100 kDa fraction (FR) isolated, and from the sterile culture supernatant, the protein-containing preparation - supernatant (S/N); in the preparations, the protein content was determined. Immunobiological properties were studied after double intraperitoneal immunization of BALB/c mice. The protective activity was determined after infection with virulent strains of S. pneumoniae serotypes 6B No. 1121 and No. 3. Phagocytic activity was studied by the number of granulocytes that absorbed heat-killed FITC-labeled S. aureus cells. The expression of Toll-like receptors (TLRs) and the subpopulation structure of mouse spleen lymphocytes were investigated after staining with FITC/PE-labeled monoclonal antibodies using flow cytometry. Statistical analysis of materials was carried out using parametric and non-parametric methods using the software package «Statistica for Windows», ver. 7.0 (Stat Soft, Inc). Results . The protective effect of FR upon infection with virulent strains of homologous (6B No. 1121) and heterologous serotypes (3 No. 3) was established; S/N protected mice when infected with strain 3 No. 3. All the studied drugs led to an increase in the number of phagocytic cells (the greatest effect was observed upon immunization with FR and S/N) and stimulated the expression of TLR2 and TLR4 positive cells (there was an increase in the number of TLR2-expressing cells during immunization with FR compared with A/E). When studying the immunophenotype of mouse lymphocytes, it was noted that the preparations induced the expression of effectors of innate and adaptive immunity. Conclusions . As a result of the studies, it was shown that the protective surface protein-containing fraction with MM 30-100 kDa is from tested experimental preparations the most active stimulator of innate and adaptive immunity, and requires further study to determine the possibility of using the serotype-independent pneumococcal vaccine in the development.

Comparison of whole-cell versus acellular pertussis vaccine effectiveness in school clusters of pertussis, France, 2013

Objectives and methodWe conducted a prospective study in 2013 to compare the whole-cell versus acellular pertussis vaccines effectiveness and duration of protection, following the occurrence of pertussis clusters. ResultsDuring seven school outbreaks, we identified 102 clinical pertussis cases, including 10 cases biologically confirmed by Bordetella pertussis specific PCR, among a cohort of 305 children in 2nd to 6th grade. The risk of pertussis when vaccinated with an acellular vaccine alone was 1.6 (RR=1.6; 95% CI=1.1–2.5) times higher than when vaccinated with a whole-cell vaccine or using a combined schedule. ConclusionsThe limited duration of protection conferred by the acellular vaccine reinforces the 2013 introduction of the pertussis booster at six years old.

BNIP3L Is a New Autophagy Related Prognostic Biomarker for Melanoma Patients Treated With AGI-101H.

Skin melanoma belongs to the most invasive malignancies with no cure for a progressing disease. Personalized therapy would allow for the selection of patients that will benefit from treatment. For this purpose, proper predictive biomarkers must be defined. Allogeneic whole-cell gene-modified therapeutic melanoma vaccine (AGI-101H) was applied in advanced melanoma patients. Humoral responses were analyzed using SEREX, and in silico gene expression analysis in TCGA melanoma patients was performed. A specific antibody response was raised against an antigen identified as BNIP3L, which correlated with a good prognosis. Moreover, AGI-101H directs an immune response against autophagy, as BNIP3L is a marker of this process. Medium and high expression of BNIP3L was also linked with longer overall survival. BNIP3L is a candidate prognostic marker of clinical outcome of melanoma patients treated with AGI-101H, and may be considered as a prediction marker for patient survival.

Outer Membrane Vesicles of Helicobacter pylori 7.13 as Adjuvants Promote Protective Efficacy Against Helicobacter pylori Infection.

Helicobacter pylori(H. pylori), a gram-negative bacterium in the human stomach with global prevalence, is relevant to chronic gastrointestinal diseases. Due to its increasing drug resistance and the low protective efficacy of some anti-H. pylori vaccines, it is necessary to find a suitable adjuvant to improve antigen efficiency. In our previous study, we determined that outer membrane vesicles (OMVs), a multicomponent secretion generated by gram-negative bacteria, of H. pylori were safe and could induce long-term and robust immune responses against H. pylori in mice. In this study, we employed two common vaccines, outer membrane proteins (OMPs) and whole cell vaccine (WCV) to assess the adjuvanticity of OMVs in mice. A standard adjuvant, cholera toxin (CT), was used as a control. Purified H. pylori OMVs used as adjuvants generated lasting anti-H. pylori resistance for 12 weeks. Additionally, both systematic and gastric mucosal immunity, as well as humoral immunity, of mice immunized with vaccine and OMVs combinations were significantly enhanced. Moreover, OMVs efficiently promoted Th1 immune response, but the response was skewed toward Th2 and Th17 immunity when compared with that induced by the CT adjuvant. Most importantly, OMVs as adjuvants enhanced the eradication of H. pylori. Thus, OMVs have potential applications as adjuvants in the development of a new generation of vaccines to treat H. pylori infection.

Pertussis vaccination: problems, possible solutions

Relevance. The growth of pertussis is observed in all countries of the world. The main reasons for this are: decreased level of vaccination, loss of antibodies after vaccination, new methods of examination, changes in the antigenic structure of the pathogen. Solution: vaccination of pregnant women, increasing the level of vaccination, age-related revaccinations. The prospect is to create new vaccines. Purpose. Determine the frequency of untimely vaccination against whooping cough in children of the first years of life, the duration of immunity preservation after the completed set of vaccinations, and the rationale for the catch-up immunization scheme. Results. The timeliness of vaccination was evaluated in 675 children according to vaccination certificates and form112/y. It turned out that 9,21% of children received the first vaccination after a first year old, the second – already 24,10%, the third more than 34%. 49,75% of children included in the analysis had revaccination by the 2 year old. Antibody titers were evaluated after vaccination and 1 revaccination in 114 children with informed parental consent. Through the year after whole cell vaccine did not have protective titers of 17 of the 58 children (29,3%), the grafted acellular – 15 of 56 (26.8 percent). After 3 years was examined 39 children (25 – grafted acellular vaccine and the whole-cell 14). 44,0% (11 children) and 50,0% (7 children) did not have protective titers, respectively. The data coincided with the results of serological monitoring of Rospotrebnadzor in St. Petersburg. The need for catch-up immunization and the introduction of age-related revaccinations against pertussis was confirmed. Catch-up immunization schemes have been developed. Conclusions . Almost half of children are vaccinated with a violation of the calendar, the primary complex of vaccinations, both whole-cell and acellular vaccines, does not provide long-term protection. It is necessary to introduce catchup immunization and age-related revaccinations.

Toward an Alum Free Mono-Component Monovalent Pertussis Vaccine: A Cytokine Response Assay.

Current evidence indicates the resurgence of whooping cough despite high coverage of whole-cell (wP) and acellular (aP) pertussis vaccines. To investigate the cytokine response to a genetically inactivated protein containing the S1 subunit of pertussis toxin (PTS1) with and without the Listeriolysin O (LLO-PTS1), in comparison with current wP and aP vaccines in the mice model. Thirty-six female NMRI mice aged 8 to 12 weeks (25 ± 5 g) were divided into six groups, including control (n=6) and five treated groups (n=6/each). Treated groups received intraperitoneal injection of recombinant PTS1, recombinant fusion LLO-PTS1, aP, wP, and sham (phosphate-buffered saline), whereas the control group did not receive anything. After 60 days, the serum levels of IFN-γ, IL-4, and IL-17 cytokines were evaluated by ELISA method. Our findings showed LLO-PTS1 significantly increased IL-17 and IL-4 cytokines compared with wP and aP vaccines. IFN-γ failed to increase substantially in the LLO-PTS1 group compared to others, but it was non-inferior to standard vaccines. Our alum free mono-component monovalent recombinant fusion protein (LLO-PTS1) could bear the capacity to stimulate the release of IFN-γ similar to wP and aP vaccines in the mouse model. Besides, it showed better results in stimulating the release of IL-17 and IL-4 response. This study can be regarded as a platform for further probes in booster pertussis vaccine development.

Combined RNAseq and ChIPseq Analyses of the BvgA Virulence Regulator of Bordetella pertussis.

Bordetella pertussis regulates the production of its virulence factors by the two-component system BvgAS. In the virulence phase, BvgS phosphorylates BvgA, which then activates the transcription of virulence-activated genes (vags). In the avirulence phase, such as during growth in the presence of MgSO4, BvgA is not phosphorylated and the vags are not expressed. Instead, a set of virulence-repressed genes (vrgs) is expressed. Here, we performed transcriptome sequencing (RNAseq) analyses on B. pertussis cultivated with or without MgSO4 and on a BvgA-deficient Tohama I derivative. We observed that 146 genes were less expressed under modulating conditions or in the BvgA-deficient strain than under the nonmodulating condition, while 130 genes were more expressed. Some of the genes code for proteins with regulatory functions, suggesting a BvgA/S regulation cascade. To determine which genes are directly regulated by BvgA, we performed chromatin immunoprecipitation sequencing (ChIPseq) analyses. We identified 148 BvgA-binding sites, 91 within putative promoter regions, 52 within open reading frames, and 5 in noncoding regions. Among the former, 32 are in BvgA-regulated putative promoter regions. Some vags, such as dnt and fhaL, contain no BvgA-binding site, suggesting indirect BvgA regulation. Unexpectedly, BvgA also bound to some vrg putative promoter regions. Together, these observations indicate an unrecognized complexity of BvgA/S biology.IMPORTANCE Bordetella pertussis, the etiological agent of whooping cough, remains a major global health problem. Despite the global usage of whole-cell vaccines since the 1950s and of acellular vaccines in the 1990s, it still is one of the most prevalent vaccine-preventable diseases in industrialized countries. Virulence of B. pertussis is controlled by BvgA/S, a two-component system responsible for upregulation of virulence-activated genes (vags) and downregulation of virulence-repressed genes (vrgs). By transcriptome sequencing (RNAseq) analyses, we identified more than 270 vags or vrgs, and chromatin immunoprecipitation sequencing (ChIPseq) analyses revealed 148 BvgA-binding sites, 91 within putative promoter regions, 52 within open reading frames, and 5 in noncoding regions. Some vags, such as dnt and fhaL, do not contain a BvgA-binding site, suggesting indirect regulation. In contrast, several vrgs and some genes not identified by RNAseq analyses under laboratory conditions contain strong BvgA-binding sites, indicating previously unappreciated complexities of BvgA/S biology.

Comparative genomics of whole-cell pertussis vaccine strains from India

BackgroundDespite high vaccination coverage using acellular (ACV) and whole-cell pertussis (WCV) vaccines, the resurgence of pertussis is observed globally. Genetic divergence in circulating strains of Bordetella pertussis has been reported as one of the contributing factors for the resurgence of the disease. Our current knowledge of B. pertussis genetic evolution in circulating strains is mostly based on studies conducted in countries using ACVs targeting only a few antigens used in the production of ACVs. To better understand the adaptation to vaccine-induced selection pressure, it will be essential to study B. pertussis populations in developing countries which are using WCVs. India is a significant user and global supplier of WCVs. We report here comparative genome analyses of vaccine and clinical isolates reported from India. Whole-genome sequences obtained from vaccine strains: WCV (J445, J446, J447 and J448), ACV (BP165) were compared with Tohama-I reference strain and recently reported clinical isolates from India (BPD1, BPD2). Core genome-based phylogenetic analysis was also performed using 166 isolates reported from countries using ACV.ResultsWhole-genome analysis of vaccine and clinical isolates reported from India revealed high genetic similarity and conserved genome among strains. Phylogenetic analysis showed that clinical and vaccine strains share genetic closeness with reference strain Tohama-I. The allelic profile of vaccine strains (J445:ptxP1/ptxA2/prn1/fim2–1/fim3–1; J446: ptxP2/ptxA4/prn7/fim2–2/fim3–1; J447 and J448: ptxP1/ptxA1/ prn1/fim2–1/fim3–1), which matched entirely with clinical isolates (BPD1:ptxP1/ptxA1/prn1/fim2–1 and BPD2: ptxP1/ptxA1/prn1/fim2–1) reported from India. Multi-locus sequence typing (MLST) demonstrated the presence of dominant sequence types ST2 and primitive ST1 in vaccine strains which will allow better coverage against circulating strains of B. pertussis.ConclusionsThe study provides a detailed characterization of vaccine and clinical strains reported from India, which will further facilitate epidemiological studies on genetic shifts in countries which are using WCVs in their immunization programs.

Neoantigen-based EpiGVAX vaccine initiates antitumor immunity in colorectal cancer.

Metastatic colorectal cancer (CRC) is poorly immunogenic, with limited neoantigens that can be targeted by cancer vaccine. Previous approaches to upregulate neoantigen have had limited success. In this study, we investigated the role of a DNA methyltransferase inhibitor (DNMTi), 5-aza-2'-deoxycytidine (DAC), in inducing cancer testis antigen (CTA) expression and evaluated the antitumor efficacy of a combinatorial approach with an epigenetically regulated cancer vaccine EpiGVAX and DAC. A murine model of metastatic CRC treated with combination therapy with an irradiated whole-cell CRC vaccine (GVAX) and DAC was used to assess the antitumor efficacy. DAC significantly induced expression of CTAs in CRC, including a new CTA Tra-P1A with a known neoepitope, P1A. Epigenetically modified EpiGVAX with DAC improved survival outcomes of GVAX. Using the epigenetically regulated antigen Tra-P1A as an example, our study suggests that the improved efficacy of EpiGVAX with DAC may due in part to the enhanced antigen-specific antitumor immune responses. This study shows that epigenetic therapy with DNMTi can not only induce new CTA expression but may also sensitize tumor cells for immunotherapy. Neoantigen-based EpiGVAX combined with DAC can improve the antitumor efficacy of GVAX by inducing antigen-specific antitumor T cell responses to epigenetically regulated proteins.

Daily pool testing of nasopharyngeal self-obtained swaps of healthcare professionals with definite COVID-19 exposition.

The coronavirus disease 2019 (COVID-19) pandemic challenges the national healthcare systems worldwide. Specifically, the potential risks for healthcare professionals (HCP) and quarantine measures can affect entire hospital units within hours. COVID-19 is transmitted via aerosols (cough or respiratory droplets) as well as contact with bodily fluids, or from contaminated surfaces [1] and therefore, specific situations that are known to produce high concentrations of aerosols like orotracheal intubation may lead to infection of HCPs [2]. The high-risk situations can usually be controlled and HCPs are able to prepare themselves. On the other hand, the staff of emergency departments are often confronted with unusual presentations of any disease including COVID-19 and therefore may be exposed unwillingly to infectious patients in early phases of care. There are no studies with COVID-19 cases available, but data from severe acute respiratory syndrome (SARS) and influenza strongly suggest that except for high-risk procedures as mentioned above, a surgical face mask is protective if it fits well, which is more important than the type of the mask itself [3,4] (https://apps.who.int/iris/handle/10665/331215; assessed 14 March 2020). We report about 10 HCPs who were exposed to an index case with COVID-19 and a highly positive PCR-test result. The patient was a 21-year-old man, who did not belong to a risk category as defined at that time by exposure to a confirmed case of COVID-19 or travel to a risk zone. He was presented by the Emergency Medical Services early in the morning with slightly disturbed vigilance and mental confusion. The primary hypothesis was meningitis or encephalitis. At triage, an elevated temperature of 38.5°C was detected and the patient reported respiratory symptoms such as rhinorrhea and dry cough without fever for 14 days. The symptoms had aggravated during the last couple of days prior to presentation. He also reported having been vaccinated against rabies, diphtheria and tetanus toxoids combined with whole-cell pertussis vaccine and Japanese encephalitis the day before. At triage, a facemask was placed on the patient’s face. A cranial computed tomography-scan and liquor test was normal. After a negative influenza A/B and respiratory syncytial virus test, the patient was sent home with the recommendation of bed rest. Later the SARS-CoV-2 screening PCR of the influenza swaps was reported positive. During the 7 h of treatment, 10 HCPs, 3 doctors and 7 nurses were exposed <2 m distance and at least once without facemasks. The patient was wearing a surgical facemask at all times. Gloves were worn at patient contact at all times, but no safety goggles or safety gown. No aerosol-generating procedures had been performed. All HCPs were sent in home isolation. From day 5, we started to analyze daily nasal-pharyngeal swaps, obtained by the HCPs themselves, using a real-time RT-PCR for SARS-CoV-2 performed on MagNa Pure 96 and Light Cycler 480 instruments [5]. The test was done from pooled specimen and remained negative until day 14. Two of the HCPs developed a sore throat, but none had fever. On day 14, additional swaps were taken from outside and analyzed separately. The tests of all 10 HCPs were negative. These data suggest that outside of high-risk situations, a surgical facemask, gloves and standard hand hygiene is sufficient to protect HCPs that usually follow basic hygiene recommendations, and that short exposition in this context might have a low risk of infection. Acknowledgements Conflicts of interest There are no conflicts of interest.

Two for one: Identification of P. aeruginosa and B. pertussis Cross-species Antigens

Abstract Development of effective vaccines requires logical design and formulation of two main components: antigens and adjuvants. Some compounds, called immunogens, can assume both of these roles. Notably, Bordetella pertussis (Bp) and some of its antigenic products have been found to adjuvant immune responses to other vaccines or vaccine components. We chose to explore this ability by using B. pertussis and pertussis toxin as adjuvants for a whole cell vaccine (WCV) against Pseudomonas aeruginosa (Pa). Pa is a highly antibiotic resistant Gram-negative opportunistic pathogen for which there are no approved vaccines or immunotherapies. We formulated vaccines containing Pa with Bp or pertussis toxin to determine if these components could modulate the immune response to PaWCV, as measured by quantification of bacterial burden and immune cell response after an acute pneumonia challenge. We determined that these additions to the PaWCV were associated with increased clearance of bacteria after challenge and increased Pa-binding immunoglobulins in the blood and airway. Additionally, we determined that the BpWCV alone was sufficient to induce protection from acute Pa infection. This correlated with production of antibodies that bind Pa, indicating that there is at least one cross-reactive antigen that could be providing protection. Through utilization of immunoblotting of protein fractions of Pa and mass spectroscopy, we identified this protective antigen. It will be used to formulate a subunit vaccine and tested for efficacy in protection from Pseudomonas infection. This type of cross-protection is novel within the field and could lead to the identification of a protective antigen against P. aeruginosa in B. pertussis.

Uptake and activation profile of murine dendritic cells in response to stimulation with functionalized iron oxide nanoparticles

Abstract Subunit vaccines typically have better safety profiles than live attenuated or killed whole-cell vaccines, but are often less immunogenic and less efficacious when deployed alone. Thus, immunological adjuvants are frequently formulated with subunit vaccines to improve efficacy. However, few vaccine adjuvants are FDA-approved; and for many adjuvants their mechanisms of action are poorly understood. We previously explored the use of iron oxide (IO) nanoparticles (NPs) in subunit vaccine delivery and show that the IO NPs also possess adjuvant-like qualities, eliminating the need for additional adjuvants in the vaccine formulation. This study further dissects the mode of action of IO NPs as immunological adjuvants by examining the relationship of particle size and uptake on the activation profile of murine bone marrow derived dendritic cells (BMDCs) in response to in vitro stimulation with functionalized IO NPs. Using IO NPs of sizes ranging 5 – 30 nm, we showed by flow cytometry immunophenotyping that only IOs of 20nm and 30nm could induce an activated BMDC subset. The level of NP uptake by BMDCs, as measured by Prussian Blue staining, also varied with particle size, with IO of 5nm being the most efficiently internalized. Intriguingly, the levels of BMDC activation did not correlate with the level of IO NP uptake. We hypothesize only IO NPs taken up via selective endocytic pathways will activate BMDCs. The production of cytokine/chemokines by these BMDCs as further characterizations of the adjuvant-like profiles of IO NPs in relation to particle size and cellular uptake is currently being examined. These studies begin to dissect the mode of action of IO NPs as a self-adjuvanted vaccine delivery system on the innate-adaptive immune interface.