Induction Of Specific Antibodies Research Articles

A Global Perspective on Oral Vaccination of Wildlife against Rabies.

The long-term mitigation of human-domestic animal-wildlife conflicts is complex and difficult. Over the last 50 yr, the primary biomedical concepts and actualized collaborative global field applications of oral rabies vaccination to wildlife serve as one dramatic example that revolutionized the field of infectious disease management of free-ranging animals. Oral vaccination of wildlife occurred in diverse locales within Africa, Eurasia, the Middle East, and North America. Although rabies is not a candidate for eradication, over a billion doses of vaccine-laden baits distributed strategically by hand, at baiting stations, or via aircraft, resulted in widespread disease prevention, control, or local disease elimination among mesocarnivores. Pure, potent, safe, and efficacious vaccines consisted of either modified-live, highly attenuated, or recombinant viruses contained within attractive, edible baits. Since the late 1970s, major free-ranging target species have included coyotes (Canis latrans), foxes (Urocyon cinereoargenteus; Vulpes vulpes), jackals (Canis aureus; Lupulella mesomelas), raccoons (Procyon lotor), raccoon dogs (Nyctereutes procyonoides), and skunks (Mephitis mephitis). Operational progress has occurred in all but the latter species. Programmatic evaluations of oral rabies vaccination success have included: demonstration of biomarkers incorporated within vaccine-laden baits in target species as representative of bait contact; serological measurement of the induction of specific rabies virus neutralizing antibodies, indicative of an immune response to vaccine; and most importantly, the decreasing detection of rabies virus antigens in the brains of collected animals via enhanced laboratory-based surveillance, as evidence of management impact. Although often conceived mistakenly as a panacea, such cost-effective technology applied to free-ranging wildlife represents a real-world, One Health application benefiting agriculture, conservation biology, and public health. Based upon lessons learned with oral rabies vaccination of mesocarnivores, opportunities for future extension to other taxa and additional diseases will have far-reaching, transdisciplinary benefits.

Recent advances in peptide-based nanovaccines for re-emerging and emerging infectious diseases

Peptide-based nanovaccines have emerged as promising strategies for combating re-emerging and emerging infectious diseases. They exhibit excellent immunogenicity and therapeutic potential. They have shown the ability to elicit robust immune responses, including activation of antigen-presenting cells, induction of specific antibodies and T-cell responses, and generation of memory immune cells. This comprehensive review article aims to provide a thorough overview of recent advances in the field, including self-assembling peptide-based nanovaccines, immunological mechanisms, structural design approaches, and utilization of various nanomaterials. Overall, peptide-based nanovaccines hold great promise in combating infectious diseases. Precise design and assembly of targeted and tailored immune responses will enable effective prevention, treatment, and long-term protection. Further research is needed to optimize their efficacy, safety, and clinical translation. The knowledge gained from these studies will pave the way for a future with more effective immunotherapeutic interventions against infectious diseases.

The impact of BNT162b2 mRNA vaccine on adaptive and innate immune responses

The mRNA-based BNT162b2 protects against severe disease and mortality caused by SARS-CoV-2 via induction of specific antibody and T-cell responses. Much less is known about its broad effects on immune responses against other pathogens. Here, we investigated the adaptive immune responses induced by BNT162b2 vaccination against various SARS-CoV-2 variants and its effects on the responsiveness of immune cells upon stimulation with heterologous stimuli. BNT162b2 vaccination induced effective humoral and cellular immunity against SARS-CoV-2 that started to wane after six months. We also observed long-term transcriptional changes in immune cells after vaccination. Additionally, vaccination with BNT162b2 modulated innate immune responses as measured by inflammatory cytokine production after stimulation - higher IL-1/IL-6 release and decreased IFN-α production. Altogether, these data expand our knowledge regarding the overall immunological effects of this new class of vaccines and underline the need for additional studies to elucidate their effects on both innate and adaptive immune responses.

T-cell specific antibody induction versus corticosteroid induction immunosuppression for liver transplant recipients: a meta-analysis

Corticosteroids remain the mainstay of immunosuppression for liver transplant recipients despite several serious complications including infection, hepatitis C virus (HCV) recurrence, diabetes mellitus (DM), and hypertension. We attempted to compare the safety and efficacy of T-cell specific antibody induction with complete corticosteroid avoidance. We searched MEDLINE, EMBASE, and Cochrane central library. Randomized controlled trials comparing T-cell specific antibody induction with corticosteroid induction immunosuppression were included. Our primary outcome was the incidence of biopsy-proven acute rejection. Eleven trials involving 1683 patients were included. The incidence of acute rejection was not significantly different between the antibody and steroid induction groups (risk ratio [RR] 0.85, 95% confidence interval [CI] 0.72, 1.01, P = 0.06, I2 = 0%). However, T-cell specific antibody induction significantly reduced the risk of cytomegalovirus infection (RR 0.48, 95% CI 0.33, 0.70, P = 0.0002, I2 = 3%), HCV recurrence (RR 0.89, 95% CI 0.80, 0.99, P = 0.03, I2 = 0%), DM (RR 0.41, 95% CI 0.32, 0.54, P < 0.0001, I2 = 0%) and hypertension (RR 0.71, 95% CI 0.55, 0.90, P = 0.005, I2 = 35%). Trial sequential analysis for acute rejection showed that the cumulative z-curve did not cross the Trial sequential boundary and the required information size was not reached. T-cell specific antibody induction compared to corticosteroid induction seems to significantly reduce opportunistic infections including cytomegalovirus infection and HCV recurrence and metabolic complications including DM and hypertension. However, given the insufficient study power, low quality of evidence, and heterogeneous immunosuppressive regimens, our results should be cautiously appreciated.

Expression of a recombinant ASFV P30 protein and production of monoclonal antibodies.

African Swine Fever (ASF) is an infectious disease that affects domestic pig and wild boar populations. The ASF Virus (ASFV) has a genome characterized by a very complex DNA (170-193 kb) that encodes for more than 200 different proteins. Among these, the highly immunogenic phosphoprotein p30 plays a fundamental role in the induction of specific antibodies. To date, the lack of a vaccine against the disease requires continuous studies to improve knowledge about the virus and the development of new tests in addition to virological ones. The aim of this work was to produce specific monoclonal antibodies (mAbs) against the p30 protein of ASFV, which could find useful applications in routine diagnostics and the implementation of new diagnostic tools. ASFV p30 encoding gene was amplified and used for the generation of the recombinant baculovirus by transfection of the Sf21 insect cells. The recombinant protein was analyzed by immunofluorescence assay, purified, and used for mice Balb-c immunization. The hybridomas obtained were cultured and screened, using an indirect Enzyme-linked Immunosorbent Assay (iELISA), in order to select clones that secrete the mAbs of interest. The expression of recombinant p30 protein was assessed using direct immunofluorescence. The purified p30 protein fractions were analyzed by Coomassie gels staining confirming the presence of bands with a molecular weight of 30 kDa and used for the immunization of Balb-c mice. Six clones of pure hybridomas secreting the specific mAbs against recombinant p30 were obtained and tested in iELISA. The mAbs were also characterized by Western blot and immunofluorescence assay. The best results were obtained with the anti-p30 mAb 2B8E10 clone which showed high reactivity with both recombinant and viral p30 protein, respectively. In this work, a recombinant p30 protein produced in an insect cell system was purified and used to immunize Balb-c mice. Six anti-p30 mAbs-secreting hybridomas clone cells were obtained. These mAbs displayed high reactivity against the recombinant protein, but only 2B8E10 mAb showed excellent functionality against the p30 protein produced by ASFV. These results open the possibility to develop different diagnostic assays.

A new S. suis serotype 3 infection model in pigs: lack of effect of buprenorphine treatment to reduce distress

BackgroundStreptoccocus suis (S. suis) is a major porcine pathogen causing meningitis, septicemia, arthritis and endocarditis. These diseases severely impair welfare of pigs. Experimental studies in pigs are important to better understand the pathogenesis and to identify protective antigens, as so far there is no vaccine available protecting against various serotypes (cps). Due to the severity of disease, application of appropriate refinement strategies in experimental S. suis infections is essential to reduce distress imposed on the piglets without jeopardizing the scientific output. The objectives of this study were to evaluate buprenorphine treatment as a refinement measure and serum cortisol levels as a distress read out parameter in a new S. suis cps3 infection model in pigs.ResultsIntravenous application of 2 × 108 CFU of S. suis cps3 (sly+, mrp+) to 6-week-old piglets led to severe morbidity in approximately 50% of the animals. Main pathological findings included suppurative meningoencephalitis and arthritis as well as fibrinosuppurative endocarditis. Buprenorphine treatment (0.05 mg/kg every 8 h) did not prevent signs of severe pain, high clinical scores, moderate to severe pathologies or high levels of serum cortisol in single severely affected piglets. Significant differences in the course of leukocytosis, induction of specific antibodies and bactericidal immunity were not recorded between groups with or w/o buprenorphine treatment. Of note, clinically unobtrusive piglets showed serum cortisol levels at 2 and 5 days post infectionem (dpi) comparable to the levels prior to infection with cps3. Cortisol levels in serum were significantly increased in piglets euthanized due to severe disease in comparison to clinically unobtrusive pigs.ConclusionsDifferent clinical courses and pathologies are induced after intravenous challenge of piglets with 2 × 108 CFU of this S. suis cps3 strain. The chosen protocol of buprenorphine application does not prevent severe distress in this infection model. Important parameters of the humoral immune response, such as the level of IgM binding to S. suis cps3, do not appear to be affected by buprenorphine treatment. Serum cortisol is a meaningful parameter to measure distress in piglets experimentally infected with S. suis and to evaluate refinement strategies. In this intravenous model, which includes close clinical monitoring and different humane endpoints, clinics and cortisol levels suggest convalescence in surviving piglets within 5 days following experimental infection.

Induction of protein specific antibody by carbonated hydroxy apatite as a candidate for mucosal vaccine adjuvant.

Buccal mucosae are considered as a site for vaccine delivery since they are relatively abundant with antigen-presenting dendritic cells, mainly Langerhans cells. In this study, we formulated carbonated hydroxy apatite (CHA) with ovalbumin (OVA) (denoted as CHA-OVA), incorporated it into bilayer buccal membrane to form hydrogel films containing CHA-OVA complex for vaccination via buccal mucosae. Ethylcellulose blend with polyethylene glycol 400 were used as impermeable backing layer. Physical properties of all tested buccal membranes were found suitable for mucosal application. In vitro and ex vivo release study showed there was no burst release of OVA found from all tested formula. From the in vivo examination, rabbit buccal mucosae vaccinated by mucoadhesive membranes containing CHA-OVA complex demonstrated mucosal specific antibody induction, represented the potential of CHA as a candidate of needle-free vaccine adjuvant. Future research is awaiting to investigate proper CHA crystallinity in complex with protein against targeted diseases.

Short-Term Effect of SARS-CoV-2 Spike Protein Receptor-Binding Domain-Specific Antibody Induction on Neutrophil-Mediated Immune Response in Mice.

Vaccination protects against COVID-19 via the spike protein receptor-binding domain (RBD)-specific antibody formation, but it also affects the innate immunity. The effects of specific antibody induction on neutrophils that can cause severe respiratory inflammation are important, though not completely investigated. In the present study, using a mouse model mimicking SARS-CoV-2 virus particle inhalation, we investigated neutrophil phenotype and activity alterations in the presence of RBD-specific antibodies. Mice were immunized with RBD and a week after a strong antibody response establishment received 100 nm particles in the RBD solution. Control mice received injections of a phosphate buffer instead of RBD. We show that the application of 100 nm particles in the RBD solution elevates neutrophil recruitment to the blood and the airways of RBD-immunized mice rather than in control mice. Analysis of bone marrow cells of mice with induced RBD-specific antibodies revealed the increased population of CXCR2+CD101+ neutrophils. These neutrophils did not demonstrate an enhanced ability of neutrophil extracellular traps (NETs) formation compared to the neutrophils from control mice. Thus, the induction of RBD-specific antibodies stimulates the activation of mature neutrophils that react to RBD-coated particles without triggering excessive inflammation.

Opinion on the Safety of BioProtein® by the Scientific Panel on Animal Feed of the Norwegian Scientific Committee for Food Safety

Opinion on the Safety of BioProtein® by the Scientific Panel on Animal Feed of the Norwegian Scientific Committee for Food Safety

Superoxide Dismutase Prevents SARS-CoV-2-Induced Plasma Cell Apoptosis and Stabilizes Specific Antibody Induction.

The infection of coronavirus disease (COVID-19) seriously threatens human life. It is urgent to generate effective and safe specific antibodies (Abs) against the pathogenic elements of COVID-19. Mice were immunized with SARS-CoV-2 spike protein antigens: S ectodomain-1 (CoV, in short) mixed in Alum adjuvant for 2 times and boosted with CoV weekly for 6 times. A portion of mice were treated with Maotai liquor (MTL, in short) or/and heat stress (HS) together with CoV boosting. We observed that the anti-CoV Ab was successfully induced in mice that received the CoV/Alum immunization for 2 times. However, upon boosting with CoV, the CoV Ab production diminished progressively; spleen CoV Ab-producing plasma cell counts reduced, in which substantial CoV-specific Ab-producing plasma cells (sPC) were apoptotic. Apparent oxidative stress signs were observed in sPCs; the results were reproduced by exposing sPCs to CoV in the culture. The presence of MTL or/and HS prevented the CoV-induced oxidative stress in sPCs and promoted and stabilized the CoV Ab production in mice in re-exposure to CoV. In summary, CoV/Alum immunization can successfully induce CoV Ab production in mice that declines upon reexposure to CoV. Concurrent administration of MTL/HS stabilizes and promotes the CoV Ab production in mice.

Induction of Functional Specific Antibodies, IgG-Secreting Plasmablasts and Memory B Cells Following BCG Vaccination.

Tuberculosis (TB) is a major global health problem and the only currently-licensed vaccine, BCG, is inadequate. Many TB vaccine candidates are designed to be given as a boost to BCG; an understanding of the BCG-induced immune response is therefore critical, and the opportunity to relate this to circumstances where BCG does confer protection may direct the design of more efficacious vaccines. While the T cell response to BCG vaccination has been well-characterized, there is a paucity of literature on the humoral response. We demonstrate BCG vaccine-mediated induction of specific antibodies in different human populations and macaque species which represent important preclinical models for TB vaccine development. We observe a strong correlation between antibody titers in serum versus plasma with modestly higher titers in serum. We also report for the first time the rapid and transient induction of antibody-secreting plasmablasts following BCG vaccination, together with a robust and durable memory B cell response in humans. Finally, we demonstrate a functional role for BCG vaccine-induced specific antibodies in opsonizing mycobacteria and enhancing macrophage phagocytosis in vitro, which may contribute to the BCG vaccine-mediated control of mycobacterial growth observed. Taken together, our findings indicate that the humoral immune response in the context of BCG vaccination merits further attention to determine whether TB vaccine candidates could benefit from the induction of humoral as well as cellular immunity.

Enhanced cross protection by hetero prime-boost vaccination with recombinant influenza viruses containing chimeric hemagglutinin-M2e epitopes

Annual repeat influenza vaccination raises concerns about protective efficacy against mismatched viruses. We investigated the impact of heterologous prime-boost vaccination on inducing cross protection by designing recombinant influenza viruses with chimeric hemagglutinin (HA) carrying M2 extracellular domains (M2e-HA). Heterologous prime-boost vaccination of C57BL/6 mice with M2e-HA chimeric virus more effectively induced M2e and HA stalk specific IgG antibodies correlating with cross protection than homologous prime-boost vaccination. Induction of M2e and HA stalk specific IgG antibodies was compromised in 1-year old mice, indicating significant aging effects on priming subdominant M2e and HA stalk IgG antibody responses. This study demonstrates that a heterologous prime-boost strategy with recombinant influenza virus expressing extra M2e epitopes provides more effective cross protection than homologous vaccination.

Prompt Reduction in CRP, IL-6, IFN-γ, IP-10, and MCP-1 and a Relatively Low Basal Ratio of Ferritin/CRP Is Possibly Associated With the Efficacy of Tocilizumab Monotherapy in Severely to Critically Ill Patients With COVID-19.

Background and Aim: Tocilizumab, a humanized anti-IL-6 receptor antibody, has been used to treat severely to critically ill patients with COVID-19. A living systematic review with meta-analysis of recent RCTs indicates that the combination therapy of corticosteroids and tocilizumab produce better outcomes, while previous observational studies suggest that tocilizumab monotherapy is beneficial for substantial numbers of patients. However, what patients could respond to tocilizumab monotherapy remained unknown.Methods: In this retrospective study we evaluated the effects of tocilizumab monotherapy on the clinical characteristics, serum biomediator levels, viral elimination, and specific IgG antibody induction in 13 severely to critically ill patients and compared with those of dexamethasone monotherapy and dexamethasone plus tocilizumab.Results: A single tocilizumab administration led to a rapid improvement in clinical characteristics, inflammatory findings, and oxygen supply in 7 of 11 patients with severe COVID-19, and could recover from mechanical ventilation management (MVM) in 2 patients with critically ill COVID-19. Four patients exhibited rapidly worsening even after tocilizumab administration and required MVM and additional methylprednisolone treatment. Tocilizumab did not delay viral elimination or inhibit IgG production specific for the virus, whereas dexamethasone inhibited IgG induction. A multiplex cytokine array system revealed a significant increase in the serum expression of 54 out of 80 biomediators in patients with COVID-19 compared with that in healthy controls. Compared with those who promptly recovered in response to tocilizumab, patients requiring MVM showed a significantly higher ratio of basal level of ferritin/CRP and a persistent increase in the levels of CRP and specific cytokines and chemokines including IL-6, IFN-γ, IP-10, and MCP-1. The basal high ratio of ferritin/CRP was also associated with clinical deterioration even in patients treated with dexamethasone and tocilizumab.Conclusion: Tocilizumab as monotherapy has substantial beneficial effects in some patients with severe COVID-19, who showed a relatively low level of the ratio of ferritin/CRP and prompt reduction in CRP, IL-6, IFN-γ, IP-10, and MCP-1. The high ratio of ferritin/CRP is associated with rapid worsening of pneumonia. Further evaluation is warranted to clarify whether tocilizumab monotherapy or its combination with corticosteroid is preferred for severely to critically ill patients with COVID-19.

Immune Response to Therapeutic Staphylococcal Bacteriophages in Mammals: Kinetics of Induction, Immunogenic Structural Proteins, Natural and Induced Antibodies.

Bacteriophages are able to affect the human immune system. Phage-specific antibodies are considered as major factors shaping phage pharmacokinetics and bioavailability. So far, general knowledge of phage antigenicity nevertheless remains extremely limited. Here we present comparative studies of immunogenicity in two therapeutic bacteriophages, A3R and 676Z, active against Staphylococcus aureus, routinely applied in patients at the Phage Therapy Unit, Poland. Comparison of the overall ability of whole phages to induce specific antibodies in a murine model revealed typical kinetics of IgM and IgG induction by these two phages. In further studies we identified the location of four phage proteins in the virions, with the focus on the external capsid head (Mcp) or tail sheath (TmpH) or an unidentified precise location (ORF059 and ORF096), and we confirmed their role as structural proteins of these viruses. Next, we compared the immune response elicited by these proteins after phage administration in mice. Similar to that in T4 phage, Mcp was the major element of the capsid that induced specific antibodies. Studies of protein-specific sera revealed that antibodies specific to ORF096 were able to neutralize antibacterial activity of the phages. In humans (population level), none of the studied proteins plays a particular role in the induction of specific antibodies; thus none potentially affects in a particular way the effectiveness of A3R and 676Z. Also in patients subjected to phage therapy, we did not observe increased specific immune responses to the investigated proteins.

Structure and Immunogenicity of the Bordetella pertussis LOS-Derived Oligosaccharides in the Endosomal-Like Pre-Processing Mice Model.

Glycoproteins are processed endosomally prior to presentation to T cells and subsequent induction of specific antibodies. The sugar part of glycoconjugate may be degraded while the type of the process depends on the features of the particular structure. The generated carbohydrate epitopes may differ from native structures and influence immunogenicity of the antigens. We have devised a model of endosomal-like pre-processing of Bordetella pertussis 186 oligosaccharides (OSs) to verify how it affects the immunogenicity of their conjugates. The glycoconjugates of structurally defined forms of the dodecasaccharide OS were synthesized and their immunogenicity was assessed using immunochemical methods. The structural features of the oligosaccharides and their sensitivity to deamination were analyzed by NMR spectroscopy. The distal trisaccharide-comprising pentasaccharide conjugated to a protein was the most effective in inducing immune response against the B. pertussis 186 LOS and the immune response to the complete OS conjugates was significantly lower. This could be explained by the loss of the distal trisaccharide during the in-cell deamination process suggesting that the native structure is not optimal for a vaccine antigen. Consequently, our research has shown that designing of new glycoconjugate vaccines requires the antigen structures to be verified in context of possible endosomal reactions beforehand.

Pullulan-Coated Iron Oxide Nanoparticles for Blood-Stage Malaria Vaccine Delivery.

Vaccines against blood-stage malaria often aim to induce antibodies to neutralize parasite entry into red blood cells, interferon gamma (IFNγ) produced by T helper 1 (Th1) CD4+ T cells or interleukin 4 (IL-4) produced by T helper 2 (Th2) cells to provide B cell help. One vaccine delivery method for suitable putative malaria protein antigens is the use of nanoparticles as vaccine carriers. It has been previously shown that antigen conjugated to inorganic nanoparticles in the viral-particle size range (~40–60 nm) can induce protective antibodies and T cells against malaria antigens in a rodent malaria challenge model. Herein, it is shown that biodegradable pullulan-coated iron oxide nanoparticles (pIONPs) can be synthesized in this same size range. The pIONPs are non-toxic and do not induce conventional pro-inflammatory cytokines in vitro and in vivo. We show that murine blood-stage antigen MSP4/5 from Plasmodium yoelii could be chemically conjugated to pIONPs and the use of these conjugates as immunogens led to the induction of both specific antibodies and IFNγ CD4+ T cells reactive to MSP4/5 in mice, comparable to responses to MSP4/5 mixed with classical adjuvants (e.g., CpG or Alum) that preferentially induce Th1 or Th2 cells individually. These results suggest that biodegradable pIONPs warrant further exploration as carriers for developing blood-stage malaria vaccines.

Development of an Antigen Delivery Platform Using Lactobacillus acidophilus Decorated With Heterologous Proteins: A Sheep in Wolf's Clothing Story.

S-layers are bacterial structures present on the surface of several Gram-positive and Gram-negative bacteria that play a role in bacterial protection. In Lactobacillus acidophilus (L. acidophilus ATCC 4356), the S-layer is mainly composed of the protein SlpA. A tandem of two copies of the protein domain SLP-A (pfam: 03217) was identified at the C-terminal of SlpA, being this double SLP-A protein domain (in short dSLP-A) necessary and sufficient for the association of the protein to the L. acidophilus cell wall. A variety of proteins fused to the dSLP-A domain were able to spontaneously associate with high affinity to the cell wall of L. acidophilus and Bacillus subtilis var. natto, in a process that we termed decoration. Binding of dSLP-A-containing-proteins to L. acidophilus was stable at conditions that mimic the gastrointestinal transit in terms of pH, proteases, and bile salts. To evaluate if protein decoration of L. acidophilus can be adapted to generate an oral vaccine platform, a chimeric antigen derived from the bacterial pathogen Shiga-toxin-producing Escherichia coli (STEC) was constructed by fusing the sequences encoding the polypeptides EspA36–192, Intimin653–953, Tir240–378, and H7 flagellin352–374 (EITH7) to the dSLP-A domain (EITH7-dSLP-A). Recombinantly expressed EITH7-dSLP-A protein was affinity purified and combined with L. acidophilus cultures to allow the association of the chimeric antigen to the bacterial surface. EITH7-decorated L. acidophilus was orally administered to BALB/c mice and the induction of anti-EITH7 specific antibodies in sera and feces determined by ELISA. Mice presenting significantly higher anti-EITH7 antibodies titers were able to control more efficiently an experimental STEC infection than mice that received the non-decorated L. acidophilus carrier, indicating that antigen-decorated L. acidophilus can be adapted as a mucosal immunization delivery platform to elicit a protective immune response for vaccine purposes.

Anti-tumor efficacy and immune responses induced by a novel therapeutic HPV-specific gorilla adenovirus off-the-shelf immunotherapeutic (PRGN-2009)

Abstract There are about 44,000 cases of Human Papilloma Virus (HPV) associated cancer each year in the US, most commonly caused by HPV 16 and 18. Prophylactic vaccines have been successful at preventing healthy patients from acquiring HPV infections via the induction of HPV specific antibodies. To treat established HPV-associated malignancies, however, the continued development of new therapies is necessary. One promising new modality is HPV immunotherapeutics, which target the HPV 16 and 18 oncoproteins E6 and E7 with the goal to activate antigen specific cytotoxic T cells. Here, we evaluated PRGN-2009, a novel gorilla adenovirus GAd HPV off-the-shelf immunotherapeutic containing multiple CTL agonist epitopes of E6 and E7 to enhance the immune responses. Three weekly administrations with PRGN-2009 in the TC-1 mouse model led to decreases in tumor volume and weight, and significant increases in HPV16 E6-specific splenocytes evaluated by IFNg ELISPOT. PRGN-2009 also increased the tumor-infiltrating CD8 and CD4 T cells, as well as multi-functional (IFNg+, GzmB+) CD8 T cells in the tumor microenvironment (TME). Additionally, PRGN-2009 was tested in SiHa, a human HPV16+ cervical tumor, in the NSG β2m−/− PBMC humanized mouse model. Weekly PRGN-2009 administrations led to a reduction in tumor and a trending increase in CD8 and CD4 T cells in the tumor; IHC confirmed the increase of CD8 T cells into the TME. These studies provide the first evaluation of the GAd HPV off-the-shelf immunotherapeutic PRGN-2009 and its therapeutic impact against HPV-associated cancers.

Immunology and the elder eye

AbstractSeveral eye diseases occur more frequently in the elderly, such as age‐related macular degeneration (AMD), primary open angle glaucoma and intraocular melanoma. Inflammation has become an important player, and while in the past AMD was considered purely a vessel disease, it has become clear that macrophages and complement play an important role in the development of the leaky vessels. The function of macrophages changes when we age. As Makitie and Kivela had shown that macrophages have a prognostic value in uveal melanoma, we started to investigate the role of aging in the development of ocular tumors in a mouse model. Ly observed that depletion of macrophages had no effect in young mice, but prevented intraocular tumor growth in old mice. Maat, Bronkhorst and Brouwer showed that specific chromosomal abnormalities in a uveal melanoma are associated with prognosis, influx of macrophages and a high vessel density.While working on this, I learned about a patient with glaucoma who had the impression that her visual field had decreased when she had the flu. Around the same time, Dr Dong Feng Chen of the Schepens Eye Research Institute in Boston developed a murine glaucoma model. A Dutch PhD student, Dr Khanh Vu, set out to determine whether inflammation plays a role in glaucoma, and indeed, glaucomatous damage following high eye pressure increased through the induction of immune responses. High eye pressure led to influx of macrophages and T cells into the retina, and the induction of specific antibodies and T cell responses. The T cells are directed against heat shock proteins, which become expressed after trauma. Blocking T cell immune response limited the loss of retinal ganglion cells and axons in the mouse after glaucoma induction. The anti‐heat shock immune response did not develop when the mouse lacked a microbiome. Patients with glaucoma also showed anti‐heat shock protein antibodies and T cell responses.These data show that inflammation plays a role in several age‐related eye disease, including age‐related macular degeneration, uveal melanoma and glaucoma.Makitie T. et al. Tumor‐infiltrating macrophages (CD68+ cells) and prognosis in malignant uveal melanoma. Invest Ophthalmol Vis Sci 2001; 42: 1414‐1421. Bronkhorst I.H. et al. Detection of M2 macrophages in uveal melanoma and relation with survival. Invest Ophtalmol Vis Sci 2011; 52: 643‐650. Doi: 10.1167/iovs.10‐5979. Ly L.V. et al. In aged mice, outgrowth of intraocular melanoma depends on proangiogenic.M2 type macrophages. J Immunol. 2010; 185: 3481‐3488. Chen H. et al. Commensal microflora‐induced T cell responses mediate progressive neurodegeneration in glaucoma. Nat Commun 2018; Aug 10;9(1): 3209. Doi: 10.1038/s41467‐018‐05681‐9

Induction of Phage-Specific Antibodies by Two Therapeutic Staphylococcal Bacteriophages Administered per os.

In therapeutic phage applications oral administration is a common and well-accepted delivery route. Phages applied per os may elicit a specific humoral response, which may in turn affect phage activity. We present specific anti-phage antibody induction in mice receiving therapeutic staphylococcal bacteriophage A3R or 676Z in drinking water. The schedule comprised: (1) primary exposure to phages for 100 days, followed by (2) diet without phage for 120 days, and (3) secondary exposure to the same phage for 44 days. Both phages induced specific antibodies in blood (IgM, IgG, IgA), even though poor to ineffective translocation of the phages to blood was observed. IgM reached a maximum on day 22, IgG increased from day 22 until the end of the experiment. Specific IgA in the blood and in the gut were induced simultaneously within about 2 months; the IgA level gradually decreased when phage was removed from the diet. Importantly, phage-specific IgA was the limiting factor for phage activity in the gastrointestinal tract. Multicopy proteins (major capsid protein and tail morphogenetic protein H) contributed significantly to phage immunogenicity (IgG), while the baseplate protein gpORF096 did not induce a significant response. Microbiome composition assessment by next-generation sequencing (NGS) revealed that no important changes correlated with phage treatment.