Antigen-specific Systemic Immune Responses Research Articles

Abstract CT052: A phase IB study of mRNA-based active cancer vaccine, BI1361849, combined with durvalumab and tremelimumab immunotherapy in patients with non-small cell lung cancer (NSCLC)

Abstract Introduction. BI1361849 is an RNA-based cancer vaccine, encoding for 6 tumor-associated antigens, including MUC1, survivin, NY-ESO-1, 5T4, MAGE-C2 and MAGE-C1, to balance systemic antigen-specific immune response. This open-label multicenter 2-arm Phase IB study evaluated the safety and preliminary efficacy of the addition of BI1361849 to durvalumab with or without tremelimumab for NSCLC. Methods. Patients with metastatic NSCLC were enrolled in one of the two arms, Arm A: BI1361849 + durvalumab and Arm B: BI1361849 + durvalumab+ tremelimumab, to determine Recommended Combination Dose (RCD) for durvalumab according to a standard 3 + 3 design with a starting dose of 1500 mg and possible de-escalation to 750 mg, and a fixed total dose of BI1361849 at 960 μg and tremelimumab at 75 mg. Primary endpoints were safety and tolerability. Secondary endpoints were progression free survival (PFS), objective response rate (ORR), disease control rate (DCR), duration of response (DOR), and overall survival (OS), based on irRECIST and RECIST. Statistical analysis included descriptive statistics, Fisher exact test, and Kaplan-Meier method. Results. A total of 57 patients were treated with 23 patients in Arm A and 34 patients in Arm B for safety analysis. Among them, 36 patients received at least 75% of the scheduled doses of durvalumab and tremelimumab and at least 4 vaccine doses in the first 2 cycles to meet efficacy analysis requirement with 17 patients in Arm A and 19 patients in Arm B. The RCD was durvalumab at 1500 mg, BI 1361849 at 960 μg, and tremelimumab at 75 mg. Both arms had comparable treatment related adverse events (56-57%) with generally low grade and manageable within current guidelines. Both arms had a comparable rate of treatment discontinuation (22-24%). For efficacy analysis, patients in Arm A had 29% ORR and 71% DCR, median months of 10, 7.3 5.7, and not reached for DOR, irPFS, PFS, and OS, respectively. In contrast, Arm B yielded 11% ORR and 53% DCR, median months of 6, 2.5, 2.5 and 10 for DOR, irPFS, PFS, and OS, respectively. Conclusions. Overall, the doublet and triplet regimens were well tolerated. In comparison to recent published studies treated with durvalumab and/or tremelimumab, incorporation of BI1361849 to durvalumab yielded comparable or better treatment response, PFS, and OS than durvalumab alone or with chemotherapy. However, tremelimumab did not add clinical benefit. Citation Format: Dung-Tsa Chen, Joshua Sabari, Jonathan Thompson, Jiaxin Niu, Hirva Mamdani, Ram Thapa, Zachary Thompson, Thanmai Posina, Aileen Ryan, Ralph Venhaus, Toni Ricciardi, Kristen Ramirez, Tawee Tanvetyanon, Jhanelle Gray. A phase IB study of mRNA-based active cancer vaccine, BI1361849, combined with durvalumab and tremelimumab immunotherapy in patients with non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT052.

Enhanced Severe Acute Respiratory Syndrome Coronavirus 2 Antigen-Specific Systemic Immune Responses in Multisystem Inflammatory Syndrome in Children and Reversal After Recovery.

BackgroundMultisystem inflammatory syndrome in children (MIS-C) presents with inflammation and pathology of multiple organs in the pediatric population in the weeks following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.MethodsWe characterized the SARS-CoV-2 antigen–specific cytokine and chemokine responses in children with MIS-C, coronavirus disease 2019 (COVID-19), and other infectious diseases.ResultsMIS-C is characterized by elevated levels of type 1 (interferon-γ, interleukin [IL] 2), type 2 (IL-4, IL-13), type 17 (IL-17), and other proinflammatory cytokines (IL-1α, IL-6, IL-12p70, IL-18, and granulocyte-macrophage colony-stimulating factor) in comparison to COVID-19 and other infectious diseases following stimulation with SARS-CoV-2–specific antigens. Similarly, upon SARS-CoV-2 antigen stimulation, CCL2, CCL3, and CXCL10 chemokines were significantly elevated in children with MIS-C in comparison to the other 2 groups. Principal component analysis based on these cytokines and chemokines could clearly distinguish MIS-C from both COVID-19 and other infections. In addition, these responses were significantly diminished and normalized 6–9 months after recovery.ConclusionsOur data suggest that MIS-C is characterized by an enhanced production of cytokines and chemokines that may be associated with disease pathogenesis.

Characterization of an inactivated whole-virus bivalent vaccine that induces balanced protective immunity against coxsackievirus A6 and A10 in mice

Coxsackievirus A6 (CVA6) and CVA10 are two of the major pathogens associated with hand, foot and mouth disease (HFMD) in children. The majority of CVA6 and CVA10 infections result in mild, self-limiting episodes (fever and herpangina) in pediatric populations; however, in some cases, can proceed to severe neurological disease and death. Efforts to mitigate viral transmission to decrease the morbidity and mortality associated with infection would be greatly strengthened by the availability of an efficacious CVA6 and CVA10 bivalent vaccine. Here we report the immunogenicity and protective efficacy of a bivalent combination vaccine comprised of formaldehyde-inactivated, whole-virus CVA6 and CVA10. We demonstrate that subcutaneous delivery of the bivalent vaccine can induce antigen-specific systemic immune responses, particularly the induction of polyfunctional T cells, which elicit active immunization to achieve a protection rate of >80% in the infected neonatal mice. Furthermore, passive transfer of the antisera from vaccinated mice potently protected recipient mice against CVA6 and CVA10 challenge. Importantly, the bivalent vaccine could induce high levels of IgG and neutralizing antibodies in adult female mice and the maternal antibody transmitted to the recipient mice played an important role in controlling homotypic and heterotypic CVA6 and CVA10 infections and viral replication in vivo. Collectively, these findings indicate that there is no immunological interference between the two antigens with respect to their ability to induce virus-specific immune responses, and thus provides proof-of-concept for further development of multivalent vaccines for broad protection against HFMD.

Abstract 4558: Immunogenicity and antitumor efficacy of live attenuated Salmonella typhimurium-based oral T-cell vaccines VXM01m, VXM04m and VXM06m

Abstract Significant progresses have been recently accomplished in immuno-oncology and in the development of cancer immunotherapies. However, novel solutions are necessary to overcome the peripheral tolerance and the immunosuppressive tumor microenvironment that prevent the eradication of cancer. VAXIMM is developing first-in-kind Salmonella typhi Ty21a-based oral T-cell vaccination platform for the initiation of anti-tumor cellular immune responses via a unique mode-of-action. This study summarizes the immunogenicity and preclinical anti-cancer efficacy for Salmonella typhimurium murine vaccines VXM01m, VXM04m and VXM06m which encode murine vascular endothelial growth factor receptor 2 (VEGFR2), mesothelin (MSLN) and Wilm's tumor 1 (WT1) protein antigens, respectively. Immunokinetic studies were performed in healthy animals treated with 1010 CFU/dose of either the control Salmonella typhimurium empty vector VXM0m_empty, or Salmonella typhimurium VXM01m, VXM06m or VXM04m, and antigen-specific T cells were detected with MHC class I pentamers by flow cytometry in the spleen. The anti-tumor efficacy of VXM01m and VXM04m was evaluated in the Panc02 syngeneic model of pancreatic adenocarcinoma overexpressing MSLN, and the anti-cancer activity of VXM06m was evaluated in the FBL-3 disseminated model of erythroleukemia expressing WT1, in a prime-boost setting. Treatment with each of the vaccines VXM01m, VXM04m and VXM06m induce a peak antigen-specific systemic immune response 7 to 10 days after the last vaccination. Treatment of Panc02 tumor-bearing mice with VXM01m and VXM04m single agents resulted in a significant reduction in the tumor growth rate, compared to the control group, with a median T/C of 37.6% and 19.4% respectively, 35 days after tumor challenge. Treatment of mice bearing FBL-3 leukemia with VXM06m generated a rapid and sustained anti-tumor effect with 100% (10 out of 10) of surviving animals 175 days after leukemia challenge. In contrast, treatment with VXM0_empty did not show any anti-cancer effect, with a median survival of 45 days and 0% (0 out of 10) of cancer regression. VXM01m, VXM04m and VMX06m were tolerated at the effective doses and have demonstrated consistent anti-cancer activities with substantial T cell responses in different animal tumor models. This study provides further evidence that VAXIMM’s versatile oral T-cell vaccination platform can be used to stimulate anti-tumor immunity against various tumor-associated antigens. Further studies of VAXIMM's cancer vaccine candidates, as monotherapy as well as in combination, are warranted. Citation Format: Sébastien Wieckowski, Lilli Podola, Marco Springer, Iris Kobl, Zina Koob, Caroline Mignard, Amine Adda Berkane, Ming Wei, Albrecht Meichle, Klaus Breiner, Matthias Schroff, Philipp Beckhove, Heinz Lubenau. Immunogenicity and antitumor efficacy of live attenuated Salmonella typhimurium-based oral T-cell vaccines VXM01m, VXM04m and VXM06m [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4558. doi:10.1158/1538-7445.AM2017-4558

Anti-PD-1 increases the clonality and activity of tumor infiltrating antigen specific T cells induced by a potent immune therapy consisting of vaccine and metronomic cyclophosphamide

BackgroundFuture cancer immunotherapies will combine multiple treatments to generate functional immune responses to cancer antigens through synergistic, multi-modal mechanisms. In this study we explored the combination of three distinct immunotherapies:...

Intranasal Vaccination against HIV-1 with Adenoviral Vector-Based Nanocomplex Using Synthetic TLR-4 Agonist Peptide as Adjuvant.

Recombinant type 5 adenovirus (rAd5) vaccines hold the promise to prevent HIV-1 infections. Intranasal vaccination not only stimulates systemic immunity but also elicits mucosal immunity that provides first defense for mucosally transmitted diseases like HIV-1. Adjuvants such as TLR agonists are usually codelivered with antigens to enhance the immunogenicity of vaccines. Here, we present a rAd5 vaccine delivery system using DEG-PEI as the carrier. Adenovirus encoding HIV gag was used as antigen, and was complexed with DEG-PEI polymer via electrostatic interaction. A novel synthetic TLR-4 agonist, RS09, was either chemically linked with DEG-PEI (DP-RS09) or physically mixed with it(DP/RS09) to enhance the immunogenticity of rAd5 vaccine. After intranasal immunization, the systemic antigen-specific immune responses and cytotoxicity T lymphocytes responses induced by DP-RS09-rAd5 and DP/RS09-rAd5 were analyzed. The mucosal secretory IgA level was detected in both nasal and vaginal washes to determine the mucosal immunity. Furthermore, cytokine productions on RAW264.7 cells were tested after preincubation with TLR-4 pathway inhibitors. The results indicated that DEG-PEI could facilitate the intranasal delivery of rAd5 vaccine. Both chemically linked (DP-RS09) and physically mixed RS09 (DP/RS09) could further enhance the mucosal immunity of rAd5 vaccine via TLR-4 pathway. This RS09 adjuvanted DEG-PEI polymer represents a potential intranasal vaccine delivery system and may have a wider application for other viral vectors.

Development of a poly (lactic-co-glycolic acid) particle vaccine to protect against house dust mite induced allergy.

Poly(lactic-co-glycolic acid) (PLGA) particles carrying antigen and adjuvant is a promising vaccine system which has been shown to stimulate systemic antigen-specific immune responses. In this study, we investigated the relationship of (i) the sizes of PLGA particle and (ii) the presence of cytosine-phosphate-guanine motifs (CpG), with the extent and type of immune response stimulated against Dermatophagoides pteronyssinus-2 (Der p2) antigen. Different sizes of PLGA particles encapsulating CpG were prepared using a double emulsion solvent evaporation method. Mice were vaccinated with Der p2 and different sizes of empty or CpG-loaded PLGA particles. Vaccinated mice were exposed to daily intranasal instillation of Der p2 for 10 days followed by euthanization to estimate leukocyte accumulation in bronchoalveolar lavage (BAL) fluids, antibody profiles, and airway hyperresponsiveness. PLGA particles showed a size-dependent decrease in the proportion of eosinophils found in BAL fluids. Mice vaccinated with the Der p2 coated on 9-μm-sized empty PLGA particles showed increased levels of IgE and IgG1 antibodies as well as increased airway hyperresponsiveness. All sizes of PLGA particles encapsulating CpG prevented airway hyperresponsiveness after Der p2 exposures. Inflammatory responses to Der p2 exposure were significantly reduced when smaller PLGA particles were used for vaccination. In addition, encapsulating CpG in PLGA particles increased IgG2a secretion. This study shows that the size of PLGA particles used for vaccination plays a major role in the prevention of house dust mite-induced allergy and that incorporation of CpG into the PLGA particles preferentially develops a Th1-type immune response.

Intranasal immunization protects against Acinetobacter baumannii-associated pneumonia in mice

Multidrug-resistant Acinetobacter baumannii has become an important causative agent of healthcare associated infections. Hospital- and community-acquired pneumonia is the most common clinical manifestation of A. baumannii infection worldwide and is often associated with high mortality. Most experimental vaccine studies to date have evaluated vaccines against systemic A. baumannii infections following systemic immunization. We recently demonstrated that a mouse model of respiratory A. baumannii infection using the strain LAC-4 results in disease progression that is similar to that observed in humans. Here we used this model in conjunction with an inactivated whole cell vaccine to evaluate the feasibility of developing protective mucosal vaccines against respiratory A. baumannii infection and to investigate the potential mechanism of protection of such vaccines. Our results showed that intranasal immunization with formalin-killed whole cells of the LAC-4 strain elicited mucosal and systemic antigen-specific immune responses, and protected mice against lethal intranasal or intraperitoneal challenges. Compared to naïve mice, immunized mice had significantly fewer bacteria in their lungs, and the pathogen was barely detectable in blood and spleens at 24h post challenge, indicating the ability of immunized mice to control extrapulmonary dissemination of the pathogen. Mechanistic studies using gene-deficient mice, neutropenic mice, or passive immunization showed that B cells and neutrophils, but not FcRγ, played crucial roles in the protection against respiratory A. baumannii challenge of intranasally immunized mice whereas passive transfer of hyperimmune sera only prolonged the survival time of challenged mice by 48h. These results provide immunological insights for the rational design of novel mucosal vaccines to protect against respiratory A. baumannii infection and demonstrate the feasibility to develop such vaccines.

Intravenous genetic mesothelin vaccine based on human adenovirus 40 inhibits growth and metastasis of pancreatic cancer.

High pancreatic cancer mortality and poor prognosis are caused by the difficulty for early diagnosis and extremely low rates of resection because of metastasis. Mesothelin overexpression in pancreatic cancer is a remarkable biomarker for tumor progression, especially for invasion and metastasis. Here, we generated a novel replication-defective recombinant adenovirus 40 (rAd40), whose gene delivery properties are totally different from a conventional rAd5. In this study, we have identified intravenous administration with rAd40 expressing mouse mesothelin (Msln) as an effective prophylactic cancer vaccine against metastatic lesions of pancreatic cancer in mice. Intravenous administration of rAd40 (rAd40 i.v.) achieved transgene delivery in wider range of organs compared to rAd5 i.v., while rAd5 was distributed mainly to the liver, spleen, and lungs. Additionally, rAd40 i.v. showed less transduction of the liver or inflammatory responses, resulted in reduced liver toxicity compared to rAd5 i.v. Also, more robust systemic antigen-specific immune responses were stimulated by rAd40 i.v. Pretreatment with a single ovalbumin-expressing rAd40 i.v. prevented tumor growth in mouse subcutaneous models of ovalbumin-expressing pancreatic cancer. When used with Msln-expressing rAd40 i.v., Msln protein expression and metastases were suppressed in a syngeneic orthotopic mouse model of pancreatic cancer, corresponding to the detection of Msln- and tumor-specific cytotoxic T lymphocyte (CTL). Our novel methods generated antitumor effects against antigen-expressing tumors through antigen- and tumor-specific CTL-mediated immunity. Thus, our results indicate that a rAd40-based intravenous vaccine provides a new strategy for the effective control of metastatic pancreatic cancer and novel therapy against other cancers and infectious diseases.

Biodegradable Particles as Vaccine Delivery Systems: Size Matters

Poly(lactide-co-glycolide) (PLGA) particles have strong potential as antigen delivery systems. The size of PLGA particles used to vaccinate mice can affect the magnitude of the antigen-specific immune response stimulated. In this study, we fabricated and characterized 17 μm, 7 μm, 1 μm, and 300 nm PLGA particles coloaded with a model antigen ovalbumin (OVA) and CpG oligodeoxynucleotides (CpG ODN). PLGA particles demonstrated a size-dependent burst release followed by a more sustained release of encapsulated molecules. PLGA particles that were 300 nm in size showed the highest internalization by, and maximum activation of, dendritic cells. The systemic antigen-specific immune response to vaccination was measured after administration of two intraperitoneal injections, 7 days apart, of 100 μg OVA and 50 μg CpG ODN in C57BL/6 mice. In vivo studies showed that 300 nm sized PLGA particles generated the highest antigen-specific cytotoxic T cell responses by days 14 and 21. These mice also showed the highest IgG2a:IgG1 ratio of OVA-specific antibodies on day 28. This study suggests that the smaller the PLGA particle used to deliver antigen and adjuvants the stronger the antigen-specific cytotoxic T cell response generated.

Lactobacillus rhamnosus GG and Bifidobacterium animalis MB5 Induce Intestinal but Not Systemic Antigen-Specific Hyporesponsiveness in Ovalbumin-Immunized Rats

Probiotics may modulate the host immune response by mechanisms not yet fully understood. We evaluated the modulation of intestinal and systemic antigen-specific immune response by Lactobacillus rhamnosus GG (LGG) or Bifidobacterium animalis MB5 in tolerized and immunized rats. Three groups of rats received orally LGG, B. animalis, or PBS (control) for 28 d. Each group was divided into two subgroups of tolerized or immunized rats receiving orally ovalbumin (OVA; 7 mg) or PBS on d 7, 9, and 11. All rats were immunized with OVA (300 μg) on d 14 and 21. In tolerized rats, the OVA-induced proliferative response of mesenteric lymph nodes (MLN) and spleen cells did not differ from control, indicating that the two probiotics maintained the tolerance. LGG and B. animalis in immunized rats reduced the OVA-induced proliferative response in MLN (P < 0.01) but not in spleen, whereas the proliferative response to anti-CD3 and concanavalin A of MLN and spleen cells as well as the delayed-type hypersensitivity reaction were not affected by probiotic treatment, indicating OVA-specific hyporesponsiveness restricted to intestinal immunity. This hyporesponsiveness was associated with CD4+CD25+Foxp3+ T cell expansion (P < 0.01) and increased IL-10 and TGFβ after LGG (P < 0.05), and increased apoptosis after B. animalis (P < 0.001) in MLN. In conclusion, we report a novel activity of LGG and B. animalis in inducing OVA-specific hyporesponsiveness in MLN of OVA-immunized rats that can be useful for a therapeutic strategy to prevent undesirable reactions to immunogenic antigens in the gut.

Thermal-sensitive hydrogel as adjuvant-free vaccine delivery system for H5N1 intranasal immunization

For H5N1 influenza immunization, we developed a thermal-sensitive hydrogel as intranasal vaccine delivery system, which was formulated with N-[(2-hydroxy-3-trimethylammonium) propyl] chitosan chloride (HTCC) and α, β-glycerophosphate (α, β-GP). The flowing solution of HTCC/GP under room temperature could gelate rapidly at body temperature, which significantly prolonged the H5N1 split antigen residence time in nasal cavity. This system also enhanced the transepithelial transport via the paracellular routes due to the disorganization of ZO-1 protein in nasal epithelial tissue. In comparison to naked H5N1 split antigen and MF59 adjuvanted antigen, as designed hydrogel/H5N1 vaccine induced greater antigen-specific systemic immune responses and mucosal IgA immunity without adjuvants. Furthermore, a boosted cellular and humoral response was also obtained by examination of IFN-γ and IL-4 cytokines, respectively. In addition, hydrogel based formulation promoted the antigen-specific CD8 + T cell immune memory as determined by the proportion of central and effector memory CD8 + T cells in nasal associated lymphoid tissue (NALT). These results demonstrate that the HTCC hydrogel has potential as an adjuvant-free platform for H5N1 split antigen intranasal vaccination.

Photodynamic therapy of tumors can lead to development of systemic antigen-specific immune response

Background: The mechanism by which the immune system can effectively recognize and destroy tumors is dependent on recognition of tumor antigens. The molecular identity of a number of these antigens has recently been identified and several immunotherapies have explored them as targets. Photodynamic therapy (PDT) is an anti-cancer modality that uses a non-toxic photosensitizer and visible light to produce cytotoxic reactive oxygen species that destroy tumors. PDT has been shown to lead to local destruction of tumors as well as to induction of anti-tumor immune response. Methodology/Principal Findings: We used a pair of equally lethal BALB/c colon adenocarcinomas, CT26 wild-type (CT26WT) and CT26.CL25 that expressed a tumor antigen, b-galactosidase (b-gal), and we treated them with vascular PDT. All mice bearing antigen-positive, but not antigen-negative tumors were cured and resistant to rechallenge. T lymphocytes isolated from cured mice were able to specifically lyse antigen positive cells and recognize the epitope derived from betagalactosidase antigen. PDT was capable of destroying distant, untreated, established, antigen-expressing tumors in 70% of the mice. The remaining 30% escaped destruction due to loss of expression of tumor antigen. The PDT anti-tumor effects were completely abrogated in the absence of the adaptive immune response. Conclusion: Understanding the role of antigen-expression in PDT immune response may allow application of PDT in metastatic as well as localized disease. To the best of our knowledge, this is the first time that PDT has been shown to lead to systemic, antigen- specific anti-tumor immunity.

Topical and mucosal liposomes for vaccine delivery

Mucosal (and in minor extent transcutanous) stimulation can induce local or distant mucosa secretory IgA. Liposomes and other vesicles as mucosal and transcutaneous adjuvants are attractive alternatives to parenteral vaccination. Liposomes can be massively produced under good manufacturing practices and stored for long periods, at high antigen/vesicle mass ratios. However, their uptake by antigen-presenting cells (APC) at the inductive sites remains as a major challenge. As neurotoxicity is a major concern in intranasal delivery, complexes between archaeosomes and calcium as well as cationic liposomes complexed with plasmids encoding for antigenic proteins could safely elicit secretory and systemic antigen-specific immune responses. Oral bilosomes generate intense immune responses that remain to be tested against challenge, but the admixing with toxins or derivatives is mandatory to reduce the amount of antigen. Most of the current experimental designs, however, underestimate the mucus blanket 100- to 1000-fold thicker than a 100-nm diameter liposome, which has first to be penetrated to access the underlying M cells. Overall, designing mucoadhesive chemoenzymatic resistant liposomes, or selectively targeted to M cells, has produced less relevant results than tailoring the liposomes to make them mucus penetrating. Opposing, the nearly 10 µm thickness stratum corneum interposed between liposomes and underlying APC can be surpassed by ultradeformable liposomes (UDL), with lipid matrices that penetrate up to the limit with the viable epidermis. UDL made of phospholipids and detergents, proved to be better transfection agents than conventional liposomes and niosomes, without the toxicity of ethosomes, in the absence of classical immunomodulators.

Photodynamic Therapy of Tumors Can Lead to Development of Systemic Antigen-Specific Immune Response

BackgroundThe mechanism by which the immune system can effectively recognize and destroy tumors is dependent on recognition of tumor antigens. The molecular identity of a number of these antigens has recently been identified and several immunotherapies have explored them as targets. Photodynamic therapy (PDT) is an anti-cancer modality that uses a non-toxic photosensitizer and visible light to produce cytotoxic reactive oxygen species that destroy tumors. PDT has been shown to lead to local destruction of tumors as well as to induction of anti-tumor immune response.Methodology/Principal FindingsWe used a pair of equally lethal BALB/c colon adenocarcinomas, CT26 wild-type (CT26WT) and CT26.CL25 that expressed a tumor antigen, β-galactosidase (β-gal), and we treated them with vascular PDT. All mice bearing antigen-positive, but not antigen-negative tumors were cured and resistant to rechallenge. T lymphocytes isolated from cured mice were able to specifically lyse antigen positive cells and recognize the epitope derived from beta-galactosidase antigen. PDT was capable of destroying distant, untreated, established, antigen-expressing tumors in 70% of the mice. The remaining 30% escaped destruction due to loss of expression of tumor antigen. The PDT anti-tumor effects were completely abrogated in the absence of the adaptive immune response.ConclusionUnderstanding the role of antigen-expression in PDT immune response may allow application of PDT in metastatic as well as localized disease. To the best of our knowledge, this is the first time that PDT has been shown to lead to systemic, antigen- specific anti-tumor immunity.

Modulation of systemic antigen-specific immune responses by oral antigen in humans

Oral antigen uptake can induce systemic immune responses ranging from tolerance to immunity. However, the underlying mechanisms are poorly understood, especially in humans. Here, keyhole limpet hemocyanin (KLH), a neoantigen which has been used in earlier studies of oral tolerance, was fed in a repeated low-dose and a single high-dose protocol to healthy volunteers. KLH-specific CD4(+) T-cell proliferation and cytokine production, as well as KLH-specific serum Ab and the effects of oral KLH on a subsequent parenterally induced systemic immune response, were analyzed. Repeated low-dose oral KLH alone induced antigen-specific CD4(+) T cells positive predominantly for the gut-homing receptor integrin β7 and the cytokines IL-2 and TNF-α; some CD4(+) T cells also produced IL-4. Oral feeding of KLH accelerated a subsequent parenterally induced systemic CD4(+) T-cell response. The cytokine pattern of KLH-specific CD4(+) T cells shifted toward more IL-4- and IL-10- and less IFN-γ-, IL-2- and TNF-α-producing cells. The parenterally induced systemic KLH-specific B-cell response was accelerated and amplified by oral KLH. The impact of single high-dose oral KLH on antigen-specific immune responses was less pronounced compared with repeated low-dose oral KLH. These findings suggest that oral antigen can effectively modulate subsequently induced systemic antigen-specific immune responses. Immunomodulation by oral antigen may offer new therapeutic strategies for Th type1-mediated inflammatory diseases and for the development of vaccination strategies.

Archaeal lipid mucosal vaccine adjuvant and delivery system

Archaeal polar lipids are being evaluated as adjuvants/vaccine delivery systems for mucosal vaccines that can provide protection against pathogens that enter the human host via the mucosal surfaces. Archaeosomes, liposomes made from polar lipids extracted from Archaea, with encapsulated antigens elicit strong antigen-specific systemic immune responses upon systemic or intranasal immunization, but fail to generate mucosal immune responses. However, intranasal immunization of mice with the archaeal lipid mucosal vaccine adjuvant and delivery (AMVAD) system, obtained by the interaction of archaeosomes/antigens with multivalent cations, induces robust, antigen-specific IgA responses in nasal and vaginal mucosa, feces, bile, and serum. In addition, strong antigen-specific systemic antibody (serum IgG, IgG1 and IgG2a) and cell-mediated responses, including CD8+ cytotoxic T lymphocyte, are generated. The responses are sustained over time and are subject to good memory-boost responses. The AMVAD formulations are stable during storage, have a good safety profile and show protective efficacy in a murine model of infection/challenge.

Antigen-sensitized CD4+CD62Llow memory/effector T helper 2 cells can induce airway hyperresponsiveness in an antigen free setting

BackgroundAirway hyperresponsiveness (AHR) is one of the most prominent features of asthma, however, precise mechanisms for its induction have not been fully elucidated. We previously reported that systemic antigen sensitization alone directly induces AHR before development of eosinophilic airway inflammation in a mouse model of allergic airway inflammation, which suggests a critical role of antigen-specific systemic immune response itself in the induction of AHR. In the present study, we examined this possibility by cell transfer experiment, and then analyzed which cell source was essential for this process.MethodsBALB/c mice were immunized with ovalbumin (OVA) twice. Spleen cells were obtained from the mice and were transferred in naive mice. Four days later, AHR was assessed. We carried out bronchoalveolar lavage (BAL) to analyze inflammation and cytokine production in the lung. Fluorescence and immunohistochemical studies were performed to identify T cells recruiting and proliferating in the lung or in the gut of the recipient. To determine the essential phenotype, spleen cells were column purified by antibody-coated microbeads with negative or positive selection, and transferred. Then, AHR was assessed.ResultsTransfer of spleen cells obtained from OVA-sensitized mice induced a moderate, but significant, AHR without airway antigen challenge in naive mice without airway eosinophilia. Immunization with T helper (Th) 1 elicited antigen (OVA with complete Freund's adjuvant) did not induce the AHR. Transferred cells distributed among organs, and the cells proliferated in an antigen free setting for at least three days in the lung. This transfer-induced AHR persisted for one week. Interleukin-4 and 5 in the BAL fluid increased in the transferred mice. Immunoglobulin E was not involved in this transfer-induced AHR. Transfer of in vitro polarized CD4+ Th2 cells, but not Th1 cells, induced AHR. We finally clarified that CD4+CD62Llow memory/effector T cells recruited in the lung and proliferated, thus induced AHR.ConclusionThese results suggest that antigen-sensitized memory/effector Th2 cells themselves play an important role for induction of basal AHR in an antigen free, eosinophil-independent setting. Therefore, regulation of CD4+ T cell-mediated immune response itself could be a critical therapeutic target for allergic asthma.

In Vivo IL-10 Gene Delivery Suppresses Airway Eosinophilia and Hyperreactivity by Down-Regulating APC Functions and Migration without Impairing the Antigen-Specific Systemic Immune Response in a Mouse Model of Allergic Airway Inflammation

IL-10 is an immunosuppressive cytokine. Although previous studies have reported that exogenous delivery of IL-10 reduced airway inflammation in experimental allergic airway inflammation, the mechanism of action has not been fully clarified. In this report, we elucidated a mechanism of action of IL-10 in vivo. BALB/c mice were immunized and aerosol challenged with OVA-Ag. We delivered the IL-10 gene to the mice before systemic sensitization or during aerosol Ag challenge by administering an IL-10-producing plasmid vector. Not only presensitization delivery of IL-10, as reported, but also delivery during inflammation strongly suppressed the development of airway eosinophilia and hyperreactivity. Presensitization delivery suppressed the Ag-specific Th2-type immune response in both the lung and spleen. In contrast, delivery in the effector phase suppressed the Th2 response only in the lung, whereas that in the spleen was not affected. IL-10 gene delivery did not induce the development of a regulatory phenotype of T cells or dendritic cells; rather, it suppressed the overall functions of CD11c(+) APCs of the lung such as Ag-presenting capacity, cytokine production, and transportation of OVA-Ag to lymph nodes, thus attenuating Th2-mediated allergic airway inflammation. Further, IL-10 revealed a distinct immunosuppressive effect in the presence of Ag and APCs. These results suggest that suppression of APC function in the lung, the site of immune response, played a critical role in the IL-10-mediated suppression of Ag-induced airway inflammation and hyperreactivity. Therefore, if delivered selectively, IL-10 could site specifically suppress the Ag-specific immune response without affecting systemic immune responses.

Dendritic Cells Targeting Flt3 Ligand As Molecular Mucosal Adjuvant

Nasal antigen plus mucosal adjuvant delivery has emerged as perhaps the most effective route for induction of both mucosal and peripheral immune response. It has been shown that dendritic cells (DCs) played essential roles for the induction of both immunity and tolerance. Thus, nasal application of CpG ODN targeting plasmacytoid DCs effectively induced antigen-specific immunity. Our recent studies showed that nasal or oral immunization with protein antigen, ovalbumin plus plasmid encoding Flt3 ligand (pFL) cDNA as adjuvant effectively elicited OVA-specific immune responses in both mucosal and systemic lymphoid tissues. The numbers of (DCs) in mice given pFL as mucosal adjuvant were significantly increased. Further, these expanded DCs were activated and expressed MHC II and co-stimulatory molecules (CD40, CD80 and CD86). These results show that mucosal delivery of pFL is DCs targeting adjuvant system that effectively induce both mucosal and systemic antigen-specific immune responses.