Pathogen-Sensing and Regulatory T Cells: Integrated Regulators of Immune Responses

PubMed Central

Grossman, Zvi; Paul, William E.

2014-01-01

We present the concept that pathogen-sensing and Tregs mutually regulate immune responses to conventional and tumor antigens through countervailing effects on dendritic cells. Normally, conventional CD4 T cells recognizing their cognate antigen-presented by a dendritic cell will respond only if the dendritic cell also receives a signal through its pathogen-sensing/ danger / adjuvant recognition systems (the pathogen-sensing triad). However, if Tregs capable of interacting with the same DC are absent, dendritic cells are competent to present antigens, both foreign and self, even without the stimulation provided by the pathogen-sensing triad. Tregs recognizing an antigen presented by the DC that is also presenting antigen to a conventional CD4 T cell will prevent such responses but a signal delivered by a member of the pathogen-sensing traid will overcome the Tregs’inhibitory action and will allow responses to go forward. These considerations take on special meaning for responses to “weak antigens” such as many of the antigens displayed by spontaneous human tumors. PMID:24894087

Presentation of lipid antigens to T cells.

PubMed

Mori, Lucia; De Libero, Gennaro

2008-04-15

T cells specific for lipid antigens participate in regulation of the immune response during infections, tumor immunosurveillance, allergy and autoimmune diseases. T cells recognize lipid antigens as complexes formed with CD1 antigen-presenting molecules, thus resembling recognition of MHC-peptide complexes. The biophysical properties of lipids impose unique mechanisms for their delivery, internalization into antigen-presenting cells, membrane trafficking, processing, and loading of CD1 molecules. Each of these steps is controlled at molecular and celular levels and determines lipid immunogenicity. Lipid antigens may derive from microbes and from the cellular metabolism, thus allowing the immune system to survey a large repertoire of immunogenic molecules. Recognition of lipid antigens facilitates the detection of infectious agents and the initiation of responses involved in immunoregulation and autoimmunity. This review focuses on the presentation mechanisms and specific recognition of self and bacterial lipid antigens and discusses the important open issues.

A Novel Method Linking Antigen Presentation by Human Monocyte-Derived Macrophages to CD8+ T Cell Polyfunctionality

PubMed Central

Short, Kirsty R.; Grant, Emma J.; Vissers, Marloes; Reading, Patrick C.; Diavatopoulos, Dimitri A.; Kedzierska, Katherine

2013-01-01

To understand the interactions between innate and adaptive immunity, and specifically how virally infected macrophages impact T cell function, novel assays examining the ability of macrophages to present antigen to CD8+ T cells are needed. In the present study, we have developed a robust in vitro assay to measure how antigen presentation by human monocyte-derived macrophages (MDMs) affects the functional capacity of autologous CD8+ T cells. The assay is based on the polyfunctional characteristics of antigen-specific CD8+ T cells, and is thus called a Mac-CD8 Polyfunctionality Assay. Following purification of monocytes and their maturation to MDMs, MDMs were pulsed with an antigenic peptide to be presented to CD8+ T cells. Peptide-pulsed MDMs were then incubated with antigen-specific CD8+ T cells in order to assess the efficacy of antigen presentation to T cells. CD8+ T cell polyfunctionality was assessed by staining with mAbs to IFN-γ, TNF-α, and CD107a in a multi-color intracellular cytokine staining assay. To highlight the utility of the Mac-CD8 Polyfunctionality Assay, we assessed the effects of influenza infection on the ability of human macrophages to present antigen to CD8+ T cells. We found that influenza infection of human MDMs can alter the effector efficacy of MDMs to activate more CD8+ T cells with cytotoxic capacity. This has important implications for understanding how the virus-infected macrophages affect adaptive immunity at the site of infection. PMID:24312096

Cationic liposomes promote antigen cross-presentation in dendritic cells by alkalizing the lysosomal pH and limiting the degradation of antigens

PubMed Central

Gao, Jie; Ochyl, Lukasz J; Yang, Ellen; Moon, James J

2017-01-01

Cationic liposomes (CLs) have been widely examined as vaccine delivery nanoparticles since they can form complexes with biomacromolecules, promote delivery of antigens and adjuvant molecules to antigen-presenting cells (APCs), and mediate cellular uptake of vaccine components. CLs are also known to trigger antigen cross-presentation – the process by which APCs internalize extracellular protein antigens, degrade them into minimal CD8+ T-cell epitopes, and present them in the context of major histocompatibility complex-I (MHC-I). However, the precise mechanisms behind CL-mediated induction of cross-presentation and cross-priming of CD8+ T-cells remain to be elucidated. In this study, we have developed two distinct CL systems and examined their impact on the lysosomal pH in dendritic cells (DCs), antigen degradation, and presentation of peptide:MHC-I complexes to antigen-specific CD8+ T-cells. To achieve this, we have used 3β-[N-(N′,N′-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) as the prototypical components of CLs with tertiary amine groups and compared the effect of CLs and anionic liposomes on lysosomal pH, antigen degradation, and cross-presentation by DCs. Our results showed that CLs, but not anionic liposomes, elevated the lysosomal pH in DCs and reduced antigen degradation, thereby promoting cross-presentation and cross-priming of CD8+ T-cell responses. These studies shed new light on CL-mediated cross-presentation and suggest that intracellular fate of vaccine components and subsequent immunological responses can be controlled by rational design of nanomaterials. PMID:28243087

Microfluidic squeezing for intracellular antigen loading in polyclonal B-cells as cellular vaccines

NASA Astrophysics Data System (ADS)

Lee Szeto, Gregory; van Egeren, Debra; Worku, Hermoon; Sharei, Armon; Alejandro, Brian; Park, Clara; Frew, Kirubel; Brefo, Mavis; Mao, Shirley; Heimann, Megan; Langer, Robert; Jensen, Klavs; Irvine, Darrell J.

2015-05-01

B-cells are promising candidate autologous antigen-presenting cells (APCs) to prime antigen-specific T-cells both in vitro and in vivo. However to date, a significant barrier to utilizing B-cells as APCs is their low capacity for non-specific antigen uptake compared to “professional” APCs such as dendritic cells. Here we utilize a microfluidic device that employs many parallel channels to pass single cells through narrow constrictions in high throughput. This microscale “cell squeezing” process creates transient pores in the plasma membrane, enabling intracellular delivery of whole proteins from the surrounding medium into B-cells via mechano-poration. We demonstrate that both resting and activated B-cells process and present antigens delivered via mechano-poration exclusively to antigen-specific CD8+T-cells, and not CD4+T-cells. Squeezed B-cells primed and expanded large numbers of effector CD8+T-cells in vitro that produced effector cytokines critical to cytolytic function, including granzyme B and interferon-γ. Finally, antigen-loaded B-cells were also able to prime antigen-specific CD8+T-cells in vivo when adoptively transferred into mice. Altogether, these data demonstrate crucial proof-of-concept for mechano-poration as an enabling technology for B-cell antigen loading, priming of antigen-specific CD8+T-cells, and decoupling of antigen uptake from B-cell activation.

Prolonged expression of MHC class I - peptide expression in bone marrow derived retrovirus transfected matured dendritic cells by continuous centrifugation in the presence of IL-4.

PubMed

Hettihewa, L M

2011-11-01

Dendritic cells (DCs) are potent antigen presenting cells which proceed from immature to a mature stage during their differentiation. There are several methods of obtaining long lasting mature antigen expressing DCs and different methods show different levels of antigen expressions. We investigated bone marrow derived DCs for the degree of maturation and genetically engineered antigen presentation in the presence of interleukin-4 (IL-4) as a maturity enhancer. DCs and transfected retrovirus were cultured together in the presence of granulocyte-macrophage colony stimulating factor (GMCSF)-IL4, GMCSF +IL4, lipopolysaccharide (LPS). B 7.1, B7.2 and CD11c were measured by the degree of immune fluorescence using enhanced green fluorescent protein (EGFP) shuttled retrovirus transfected antigen. Degree of MHC class I molecule with antigen presentation of antigen was also evaluated by fluorescence activated cell sorting. The antigen presenting capacity of transfected DCs was investigated. Bone marrow DCs were generated in the presence of GMCSF and IL-4 in vitro. Dividing bone marrow cells were infected with EGFP shuttled retrovirus expressing SSP2 by prolonged centrifugation for three consecutive days from day 5, 6 and 7 and continued to culture in the presence of GMSCF and IL-4 until day 8. IL-4 as a cytokine increased the maturation of retrovirus transfected DCs by high expression of B 7-1 and B 7-2. Also, IL-4 induced DC enhanced by the prolonged centrifugation and it was shown by increased antigen presentation of these dendric cells as antigen presenting cell (APC). Cytolytic effects were significantly higher in cytotoxic T cell response (CTLs) mixed with transfected DCs than CTLs mixed with pulsed DCs. There was an enhanced antigen presentation by prolonged expression of antigen loaded MHC class I receptors in DCs in the presence of IL-4 by prolonged centrifugation.

A Single Subset of Dendritic Cells Controls the Cytokine Bias of Natural Killer T Cell Responses to Diverse Glycolipid Antigens

PubMed Central

Arora, Pooja; Baena, Andres; Yu, Karl O.A.; Saini, Neeraj K.; Kharkwal, Shalu S.; Goldberg, Michael F.; Kunnath-Velayudhan, Shajo; Carreño, Leandro J.; Venkataswamy, Manjunatha M.; Kim, John; Lazar-Molnar, Eszter; Lauvau, Gregoire; Chang, Young-tae; Liu, Zheng; Bittman, Robert; Al-Shamkhani, Aymen; Cox, Liam R.; Jervis, Peter J.; Veerapen, Natacha; Besra, Gurdyal S.; Porcelli, Steven A.

2014-01-01

Summary Many hematopoietic cell types express CD1d and are capable of presenting glycolipid antigens to invariant natural killer T cells (iNKT cells). However, the question of which cells are the principal presenters of glycolipid antigens in vivo remains controversial, and it has been suggested that this might vary depending on the structure of a particular glycolipid antigen. Here we have shown that a single type of cell, the CD8α+ DEC-205+ dendritic cell, was mainly responsible for capturing and presenting a variety of different glycolipid antigens, including multiple forms of α-galactosylceramide that stimulate widely divergent cytokine responses. After glycolipid presentation, these dendritic cells rapidly altered their expression of various costimulatory and coinhibitory molecules in a manner that was dependent on the structure of the antigen. These findings show flexibility in the outcome of two-way communication between CD8α+ dendritic cells and iNKT cells, providing a mechanism for biasing toward either proinflammatory or anti-inflammatory responses. PMID:24412610

Nonclassical T Cells and Their Antigens in Tuberculosis

PubMed Central

De Libero, Gennaro; Singhal, Amit; Lepore, Marco; Mori, Lucia

2014-01-01

T cells that recognize nonpeptidic antigens, and thereby are identified as nonclassical, represent important yet poorly characterized effectors of the immune response. They are present in large numbers in circulating blood and tissues and are as abundant as T cells recognizing peptide antigens. Nonclassical T cells exert multiple functions including immunoregulation, tumor control, and protection against infections. They recognize complexes of nonpeptidic antigens such as lipid and glycolipid molecules, vitamin B2 precursors, and phosphorylated metabolites of the mevalonate pathway. Each of these antigens is presented by antigen-presenting molecules other than major histocompatibility complex (MHC), including CD1, MHC class I–related molecule 1 (MR1), and butyrophilin 3A1 (BTN3A1) molecules. Here, we discuss how nonclassical T cells participate in the recognition of mycobacterial antigens and in the mycobacterial-specific immune response. PMID:25059739

Quantitative immunophenotypic analysis of antigen-presenting cells involved in ectromelia virus antigen presentation in BALB/c and C57BL/6 mice.

PubMed

Szulc-Dąbrowska, Lidia; Gieryńska, Małgorzata; Boratyńska-Jasińska, Anna; Martyniszyn, Lech; Winnicka, Anna; Niemiałtowski, Marek G

2013-08-01

During mousepox in resistant (C57BL/6) or susceptible (BALB/c) strains of mice, stimulation of Th1 or Th2 cytokine immune response, respectively, is observed. Because mechanisms of different polarization of T cells remain elusive, in this study, we quantitatively assessed the phenotype of antigen-presenting cells (APCs) involved in ectromelia virus (ECTV) antigen presentation and cluster formation with effector cells in secondary lymphoid organs of BALB/c and C57BL/6 mice. We showed that both strains of mice display similar dynamics and kinetics of viral antigen presentation by CD11c(+) , CD11b(+) , and CD19(+) cells. CD11c(+) and CD11b(+) cells highly participated in viral antigen presentation during all stages of mousepox, whereas CD19(+) cells presented viral peptides later in infection. The main population of dendritic cells (DCs) engaged in ECTV antigen presentation and cell junction formation with effector cells was a population of myeloid CD11b(+) DCs (mDCs). We suggest that, on the one hand, ECTV may differentially affect the functions of APCs depending on the strain of mice. On the other hand, we suggest that some types of APCs, such as mDCs or other DCs subsets, have different abilities to direct the shape of immune response depending on the host resistance to mousepox. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

EVIR: chimeric receptors that enhance dendritic cell cross-dressing with tumor antigens.

PubMed

Squadrito, Mario Leonardo; Cianciaruso, Chiara; Hansen, Sarah K; De Palma, Michele

2018-03-01

We describe a lentivirus-encoded chimeric receptor, termed extracellular vesicle (EV)-internalizing receptor (EVIR), which enables the selective uptake of cancer-cell-derived EVs by dendritic cells (DCs). The EVIR enhances DC presentation of EV-associated tumor antigens to CD8 + T cells primarily through MHCI recycling and cross-dressing. EVIRs should facilitate exploring the mechanisms and implications of horizontal transfer of tumor antigens to antigen-presenting cells.

T cell activation is determined by the number of presented antigens.

PubMed

Deeg, Janosch; Axmann, Markus; Matic, Jovana; Liapis, Anastasia; Depoil, David; Afrose, Jehan; Curado, Silvia; Dustin, Michael L; Spatz, Joachim P

2013-01-01

Antigen recognition is a key event during T cell activation. Here, we introduce nanopatterned antigen arrays that mimic the antigen presenting cell surface during T cell activation. The assessment of activation related events revealed the requirement of a minimal density of 90-140 stimulating major histocompatibility complex class II proteins (pMHC) molecules per μm(2). We demonstrate that these substrates induce T cell responses in a pMHC dose-dependent manner and that the number of presented pMHCs dominates over local pMHC density.

T Cell Activation is Determined by the Number of Presented Antigens

PubMed Central

2013-01-01

Antigen recognition is a key event during T cell activation. Here, we introduce nanopatterned antigen arrays that mimic the antigen presenting cell surface during T cell activation. The assessment of activation related events revealed the requirement of a minimal density of 90–140 stimulating major histocompatibility complex class II proteins (pMHC) molecules per μm2. We demonstrate that these substrates induce T cell responses in a pMHC dose-dependent manner and that the number of presented pMHCs dominates over local pMHC density. PMID:24117051

Dendritic cells cross-present HIV antigens from live as well as apoptotic infected CD4+ T lymphocytes

NASA Astrophysics Data System (ADS)

Marañón, Concepción; Desoutter, Jean-François; Hoeffel, Guillaume; Cohen, William; Hanau, Daniel; Hosmalin, Anne

2004-04-01

A better understanding of the antigen presentation pathways that lead to CD8+ T cell recognition of HIV epitopes in vivo is needed to achieve better immune control of HIV replication. Here, we show that cross-presentation of very small amounts of HIV proteins from apoptotic infected CD4+ T lymphocytes by dendritic cells to CD8+ T cells is much more efficient than other known HIV presentation pathways, i.e., direct presentation of infectious virus or cross-presentation of defective virus. Unexpectedly, dendritic cells also take up actively antigens into endosomes from live infected CD4+ T lymphocytes and cross-present them as efficiently as antigens derived from apoptotic infected cells. Moreover, live infected CD4+ T cells costimulate cross-presenting dendritic cells in the process. Therefore, dendritic cells can present very small amounts of viral proteins from infected T cells either after apoptosis, which is frequent during HIV infection, or not. Thus, if HIV expression is transiently induced while costimulation is enhanced (for instance after IL-2 and IFN immune therapy), this HIV antigen presentation pathway could be exploited to eradicate latently infected reservoirs, which are poorly recognized by patients' immune systems.

Tumor-Derived Microvesicles Modulate Antigen Cross-Processing via Reactive Oxygen Species-Mediated Alkalinization of Phagosomal Compartment in Dendritic Cells.

PubMed

Battisti, Federico; Napoletano, Chiara; Rahimi Koshkaki, Hassan; Belleudi, Francesca; Zizzari, Ilaria Grazia; Ruscito, Ilary; Palchetti, Sara; Bellati, Filippo; Benedetti Panici, Pierluigi; Torrisi, Maria Rosaria; Caracciolo, Giulio; Altieri, Fabio; Nuti, Marianna; Rughetti, Aurelia

2017-01-01

Dendritic cells (DCs) are the only antigen-presenting cells able to prime naïve T cells and cross-prime antigen-specific CD8 + T cells. Their functionality is a requirement for the induction and maintenance of long-lasting cancer immunity. Albeit intensively investigated, the in vivo mechanisms underlying efficient antigen cross-processing and presentation are not fully understood. Several pieces of evidence indicate that antigen transfer to DCs mediated by microvesicles (MVs) enhances antigen immunogenicity. This mechanism is also relevant for cross-presentation of those tumor-associated glycoproteins such as MUC1 that are blocked in HLA class II compartment when internalized by DCs as soluble molecules. Here, we present pieces of evidence that the internalization of tumor-derived MVs modulates antigen-processing machinery of DCs. Employing MVs derived from ovarian cancer ascites fluid and established tumor cell lines, we show that MV uptake modifies DC phagosomal microenvironment, triggering reactive oxygen species (ROS) accumulation and early alkalinization. Indeed, tumor MVs carry radical species and the MV uptake by DCs counteracts the chemically mediated acidification of the phagosomal compartment. Further pieces of evidence suggest that efficacious antigen cross-priming of the MUC1 antigen carried by the tumor MVs results from the early signaling induced by MV internalization and the function of the antigen-processing machinery of DCs. These results strongly support the hypothesis that tumor-derived MVs impact antigen immunogenicity by tuning the antigen-processing machinery of DCs, besides being carrier of tumor antigens. Furthermore, these findings have important implications for the exploitation of MVs as antigenic cell-free immunogen for DC-based therapeutic strategies.

Preserved MHC-II antigen processing and presentation function in chronic HCV infection

PubMed Central

DH, Canaday; CJ, Burant; L, Jones; H, Aung; L, Woc-Colburn; DD, Anthony

2010-01-01

Individuals with chronic HCV infection have impaired response to vaccine, though the etiology remains to be elucidated. Dendritic cells (DC) and monocytes (MN) provide antigen uptake, processing, presentation, and costimulatory functions necessary to achieve optimal immune responses. The integrity of antigen processing and presentation function within these antigen presenting cells (APC) in the setting of HCV infection has been unclear. We used a novel T cell hybridoma system that specifically measures MHC-II antigen processing and presentation function of human APC. Results demonstrate MHC-II antigen processing and presentation function is preserved in both myeloid DC (mDC) and MN in the peripheral blood of chronically HCV-infected individuals, and indicates that an alteration in this function does not likely underlie the defective HCV-infected host response to vaccination. PMID:21055734

A Lipid Based Antigen Delivery System Efficiently Facilitates MHC Class-I Antigen Presentation in Dendritic Cells to Stimulate CD8+ T Cells

NASA Astrophysics Data System (ADS)

Maji, Mithun; Mazumder, Saumyabrata; Bhattacharya, Souparno; Choudhury, Somsubhra Thakur; Sabur, Abdus; Shadab, Md.; Bhattacharya, Pradyot; Ali, Nahid

2016-06-01

The most effective strategy for protection against intracellular infections such as Leishmania is vaccination with live parasites. Use of recombinant proteins avoids the risks associated with live vaccines. However, due to low immunogenicity, they fail to trigger T cell responses particularly of CD8+ cells requisite for persistent immunity. Previously we showed the importance of protein entrapment in cationic liposomes and MPL as adjuvant for elicitation of CD4+ and CD8+ T cell responses for long-term protection. In this study we investigated the role of cationic liposomes on maturation and antigen presentation capacity of dendritic cells (DCs). We observed that cationic liposomes were taken up very efficiently by DCs and transported to different cellular sites. DCs activated with liposomal rgp63 led to efficient presentation of antigen to specific CD4+ and CD8+ T cells. Furthermore, lymphoid CD8+ T cells from liposomal rgp63 immunized mice demonstrated better proliferative ability when co-cultured ex vivo with stimulated DCs. Addition of MPL to vaccine enhanced the antigen presentation by DCs and induced more efficient antigen specific CD8+ T cell responses when compared to free and liposomal antigen. These liposomal formulations presented to CD8+ T cells through TAP-dependent MHC-I pathway offer new possibilities for a safe subunit vaccine.

Inhibitory effects of thymus-independent type 2 antigens on MHC class II-restricted antigen presentation: comparative analysis of carbohydrate structures and the antigen presenting cell.

PubMed

González-Fernández, M; Carrasco-Marín, E; Alvarez-Domínguez, C; Outschoorn, I M; Leyva-Cobián, F

1997-02-25

The role of thymus-independent type 2 (TI-2) antigens (polysaccharides) on the MHC-II-restricted processing of protein antigens was studied in vitro. In general, antigen presentation is inhibited when both peritoneal and splenic macrophages (M phi) as well as Küpffer cells (KC) are preincubated with acidic polysaccharides or branched dextrans. However, the inhibitory effect of neutral polysaccharides was minimal when KC were used as antigen presenting cells (APC). Morphological evaluation of the uptake of fluoresceinated polysaccharides clearly correlates with this selective and differential interference. Polysaccharides do not block MHC-I-restricted antigen presentation. Some chemical characteristics shared by different saccharides seem to be specially related to their potential inhibitory abilities: (i) those where two anomeric carbon atoms of two interlinked sugars and (ii) those containing several sulfate groups per disaccharide repeating unit. No polysaccharide being inhibitory in M phi abrogated antigen processing in other APC: lipopolysaccharide-activated B cells, B lymphoma cells, or dendritic cells (DC). Using radiolabeled polysaccharides it was observed that DC and B cells incorporated less radioactivity as a function of time than M phi. Morphological evaluation of these different APC incubated for extended periods of time with inhibitory concentrations of polysaccharides revealed intense cytoplasmic vacuolization in M phi but not in B cells or DC. The large majority of M phi lysosomes containing polysaccharides fail to fuse with incoming endocytic vesicles and delivery of fluid-phase tracers was reduced, suggesting that indigestible carbohydrates reduced the fusion of these loaded lysosomes with endosomes containing recently internalized tracers. It is suggested that the main causes of this antigen presentation blockade are (i) the chemical characteristics of certain carbohydrates and whether the specific enzymatic machinery for their intracellular degradation exists; and (ii) the different phagocytic abilities of distinct APC populations, fluid-phase pinocytosis and receptor-mediated saccharide uptake, and existence of a differential antigen-processing pathway in M phi and DC or B cells, which could be based on a polysaccharide-inhibited step present in M phi but unaffected or irrelevant in both B cells and DC.

Antigen recognition by H-2-restricted T cells. I. Cell-free antigen processing

PubMed Central

1983-01-01

We examined the ability of a set of cloned chicken ovalbumin (cOVA)- specific, Id-restricted, T cell hybridomas to produce interleukin-2 in response to cOVA presented by the Ia+ B cell lymphoma line, A20-2J. Although viable A20-2J cells presented native, denatured, and fragmented cOVA more or less equally well, A20-2J cells that were glutaraldehyde-fixed could present only enzymatically or chemically fragmented cOVA. These results suggest that antigen fragmentation may be both necessary and sufficient to define accessory cell processing of soluble antigens so that they may be recognized in association with I- region molecules by T cells. PMID:6193218

Extraction of Cell-Wall Polysaccharide Antigen from Streptococci

PubMed Central

Slade, Hutton D.

1965-01-01

Slade, Hutton D. (Northwestern University Medical School, Chicago, Ill., and Max-Planck Institut für Immunbiologie, Freiburg, Germany). Extraction of cell-wall polysaccharide antigen from streptococci. J. Bacteriol. 90:667–672. 1965.—The carbohydrate grouping antigens in the cell walls of streptococci belonging to groups A, E, G, L, and T were extracted with 5% trichloroacetic acid at 90 C. The antigens were removed also from dry whole cells by extraction with trichloroacetic acid followed by treatment with phenol-water. Details of the methods are presented. The antigens obtained by use of either of these procedures were suitable for studies on immunological specificity and chemical structure. Quantitative enzymatic and chemical analyses of two group E antigens and one group T preparation showed the presence of l-rhamnose (22 to 44%), d-glucose (7 to 22%), d-galactose (T antigen only, 26%), glucosamine (2 to 16%), and galactosamine (T antigen only, 3%). In addition, analyses of A and G antigen preparations are presented. The protein and phosphate content of the A and E antigens were about 1% each. Quantitative precipitin curves of these antigens are presented. PMID:16562065

Inhibiting DNA methylation activates cancer testis antigens and expression of the antigen processing and presentation machinery in colon and ovarian cancer cells.

PubMed

Siebenkäs, Cornelia; Chiappinelli, Katherine B; Guzzetta, Angela A; Sharma, Anup; Jeschke, Jana; Vatapalli, Rajita; Baylin, Stephen B; Ahuja, Nita

2017-01-01

Innovative therapies for solid tumors are urgently needed. Recently, therapies that harness the host immune system to fight cancer cells have successfully treated a subset of patients with solid tumors. These responses have been strong and durable but observed in subsets of patients. Work from our group and others has shown that epigenetic therapy, specifically inhibiting the silencing DNA methylation mark, activates immune signaling in tumor cells and can sensitize to immune therapy in murine models. Here we show that colon and ovarian cancer cell lines exhibit lower expression of transcripts involved in antigen processing and presentation to immune cells compared to normal tissues. In addition, treatment with clinically relevant low doses of DNMT inhibitors (that remove DNA methylation) increases expression of both antigen processing and presentation and Cancer Testis Antigens in these cell lines. We confirm that treatment with DNMT inhibitors upregulates expression of the antigen processing and presentation molecules B2M, CALR, CD58, PSMB8, PSMB9 at the RNA and protein level in a wider range of colon and ovarian cancer cell lines and treatment time points than had been described previously. In addition, we show that DNMTi treatment upregulates many Cancer Testis Antigens common to both colon and ovarian cancer. This increase of both antigens and antigen presentation by epigenetic therapy may be one mechanism to sensitize patients to immune therapies.

Spatial separation of the processing and MHC class I loading compartments for cross-presentation of the tumor-associated antigen HER2/neu by human dendritic cells

PubMed Central

Baleeiro, Renato B; Rietscher, René; Diedrich, Andrea; Czaplewska, Justyna A; Lehr, Claus-Michael; Scherließ, Regina; Hanefeld, Andrea; Gottschaldt, Michael; Walden, Peter

2015-01-01

Cross-presentation is the process by which professional antigen presenting cells (APCs) (B cells, dendritic cells (DCs) and macrophages) present endocytosed antigens (Ags) via MHC-I to CD8+ T cells. This process is crucial for induction of adaptive immune responses against tumors and infected cells. The pathways and cellular compartments involved in cross-presentation are unresolved and controversial. Among the cells with cross-presenting capacity, DCs are the most efficient, which was proposed to depend on prevention of endosomal acidification to block degradation of the epitopes. Contrary to this view, we show in this report that some cargoes induce strong endosomal acidification following uptake by human DCs, while others not. Moreover, processing of the tumor-associated antigen HER2/neu delivered in nanoparticles (NP) for cross-presentation of the epitope HER2/neu369–377 on HLA-A2 depended on endosomal acidification and cathepsin activity as well as proteasomes, and newly synthesized HLA class I. However, the HLA-A*0201/HER2/neu369–377 complexes were not found in the endoplasmic reticulum (ER) nor in endolysosomes but in hitherto not described vesicles. The data thus indicate spatial separation of antigen processing and loading of MHC-I for cross-presentation: antigen processing occurs in the uptake compartment and the cytosol whereas MHC-I loading with peptide takes place in a distinct subcellular compartment. The findings further elucidate the cellular pathways involved in the cross-presentation of a full-length, clinically relevant tumor-associated antigen by human DCs, and the impact of the vaccine formulation on antigen processing and CD8+ T cell induction. PMID:26985398

Enhanced activation of human T cell clones specific for virus-like particles expressing the HIV V3 loop in the presence of HIV V3 loop-specific polyclonal antibodies

PubMed Central

Peifang, S.; Pira, G. L.; Fenoglio, D.; Harris, S.; Costa, M. G.; Venturino, V.; Dessì, V.; Layton, G.; Laman, J.; Huisman, J. G.; Manca, F.

1994-01-01

Recombinant virus-like particles (VLP), formed by the yeast Ty p1 protein, carrying the HIV gp120 V3 loop on their surface (V3-VLP) have been tested in vitro for immunogenicity and antigenicity by using VLP p1-specific human CD4+ T cell lines and clones. VLP-specific human T cell lines and clones were generated from normal individuals, indicating that VLP-specific precursor cells present in the peripheral lymphocyte pool can be induced to expand clonally upon antigen challenge in vitro, in the absence of previous immunization. It was also shown that V3-specific polyclonal antibodies enhance V3-VLP-induced activation of VLP-specific T cell clones. Antibody-dependent potentiation has been shown previously in other antigen systems, and it depends on enhanced uptake of complexed antigen by Fc receptor-positive antigen-presenting cells. Since in this case antigen is internalized by presenting cells as a complex, it can be inferred that a similar event of antibody-mediated antigen uptake can take place with V3-specific B cells, resulting in presentation by the B cells of T helper epitopes derived from processing of the VLP p1 moiety. This suggests that T helper cells specific for the carrier VLP p1 protein can be activated to provide help to V3-specific B cells in the presence of the appropriate antigen construct. PMID:7915974

CELL SEPARATION ON ANTIGEN-COATED COLUMNS

PubMed Central

Wigzell, Hans; Andersson, Birger

1969-01-01

Glass and plastic bead columns coated with antigenic protein molecules were used as an immunological filter for cell populations containing immune cells of relevant specificity. A selective elimination of these immune cells from the passing cell suspension was regularly noted and it approached, in some experiments, complete abolition of the specific immune reactivity of the filtered cell population. This specific retention of immune cells by antigenic columns could be selectively blocked by the presence of free antigen molecules in the medium during filtration. The results obtained support the concept of a cell-associated antigen-specific receptor being present on the outer surface of immune cells, displaying the same antigen-binding specificity as the potential product of the cell, the humoral antibody. Using the present bead column system, results were obtained indicating that this receptor was an active product of the immune cells and not any passively adsorbed, cytophilic antibody. Antigenic bead columns may very well constitute a tool for the production in vitro of cell populations being specifically deprived of immune reactivity and allow detailed analysis of the characteristics of the cell-associated antibody of immune cells. PMID:5782770

Allogeneic substitution for nominal antigen-specific T-cell clone reactivity in schistosomiasis.

PubMed Central

Linette, G P; Lammie, P J; Phillips, S M

1986-01-01

The present studies have established the nature of a T-cell clone which demonstrates dual reactivity directed against Schistosoma mansoni antigen presented by syngeneic antigen presenting cells and against allogeneic cells. Clone G4, when stimulated by either antigen (SEA) or allogeneic cells (PL/J), exhibits similar functional and phenotypic characteristics. A subclone of G4, G4A.1, which has been maintained in continuous mixed lymphocyte culture for 12 months (in the absence of SEA), retains comparable reactivity with respect to proliferation and ability to produce lymphokines, transfer delayed-type hypersensitivity, and produce in vitro granulomas in response to SEA. Normal antigenic stimulation is highly contingent upon I-Ab compatibility while antibody blocking experiments map allo-reactivity to I-Eu. The failure of B10.PL spleen cells to stimulate G4, however, suggests that alloreactivity may be directed against the recently described Mls X locus. Both allogeneic and nominal antigen induced T-cell activation are blocked by antibody directed against L3T4A, confirming Class II MHC restriction for both types of stimulation. These studies suggest that stimulation of T cells by either alloantigen or nominal antigen elicits qualitatively similar functional profiles, and further suggest the feasibility of producing large numbers of nominal antigen reactive cloned T cells in the absence of nominal antigen under mixed lymphocyte culture conditions. PMID:2420707

Antigen Cross-Presentation of Immune Complexes

PubMed Central

Platzer, Barbara; Stout, Madeleine; Fiebiger, Edda

2014-01-01

The ability of dendritic cells (DCs) to cross-present tumor antigens has long been a focus of interest to physicians, as well as basic scientists, that aim to establish efficient cell-based cancer immune therapy. A prerequisite for exploiting this pathway for therapeutic purposes is a better understanding of the mechanisms that underlie the induction of tumor-specific cytotoxic T-lymphocyte (CTL) responses when initiated by DCs via cross-presentation. The ability of humans DC to perform cross-presentation is of utmost interest, as this cell type is a main target for cell-based immunotherapy in humans. The outcome of a cross-presentation event is guided by the nature of the antigen, the form of antigen uptake, and the subpopulation of DCs that performs presentation. Generally, CD8α+ DCs are considered to be the most potent cross-presenting DCs. This paradigm, however, only applies to soluble antigens. During adaptive immune responses, immune complexes form when antibodies interact with their specific epitopes on soluble antigens. Immunoglobulin G (IgG) immune complexes target Fc-gamma receptors on DCs to shuttle exogenous antigens efficiently into the cross-presentation pathway. This receptor-mediated cross-presentation pathway is a well-described route for the induction of strong CD8+ T cell responses. IgG-mediated cross-presentation is intriguing because it permits the CD8− DCs, which are commonly considered to be weak cross-presenters, to efficiently cross-present. Engaging multiple DC subtypes for cross-presentation might be a superior strategy to boost CTL responses in vivo. We here summarize our current understanding of how DCs use IgG-complexed antigens for the efficient induction of CTL responses. Because of its importance for human cell therapy, we also review the recent advances in the characterization of cross-presentation properties of human DC subsets. PMID:24744762

MyD88/CD40 Genetic Adjuvant Function in Cutaneous Atypical Antigen-Presenting Cells Contributes to DNA Vaccine Immunogenicity

PubMed Central

Slawin, Kevin M.; Levitt, Jonathan M.; Spencer, David M.

2016-01-01

Therapeutic DNA-based vaccines aim to prime an adaptive host immune response against tumor-associated antigens, eliminating cancer cells primarily through CD8+ cytotoxic T cell-mediated destruction. To be optimally effective, immunological adjuvants are required for the activation of tumor-specific CD8+ T cells responses by DNA vaccination. Here, we describe enhanced anti-tumor efficacy of an in vivo electroporation-delivered DNA vaccine by inclusion of a genetically encoded chimeric MyD88/CD40 (MC) adjuvant, which integrates both innate and adaptive immune signaling pathways. When incorporated into a DNA vaccine, signaling by the MC adjuvant increased antigen-specific CD8+ T cells and promoted elimination of pre-established tumors. Interestingly, MC-enhanced vaccine efficacy did not require direct-expression of either antigen or adjuvant by local antigen-presenting cells, but rather our data supports a key role for MC function in “atypical” antigen-presenting cells of skin. In particular, MC adjuvant-modified keratinocytes increased inflammatory cytokine secretion, upregulated surface MHC class I, and were able to increase in vitro and in vivo priming of antigen-specific CD8+ T cells. Furthermore, in the absence of critical CD8α+/CD103+ cross-priming dendritic cells, MC was still able to promote immune priming in vivo, albeit at a reduced level. Altogether, our data support a mechanism by which MC signaling activates an inflammatory phenotype in atypical antigen-presenting cells within the cutaneous vaccination site, leading to an enhanced CD8+ T cell response against DNA vaccine-encoded antigens, through both CD8α+/CD103+ dendritic cell-dependent and independent pathways. PMID:27741278

IL-10 down-regulates T cell activation by antigen-presenting liver sinusoidal endothelial cells through decreased antigen uptake via the mannose receptor and lowered surface expression of accessory molecules.

PubMed

Knolle, P A; Uhrig, A; Hegenbarth, S; Löser, E; Schmitt, E; Gerken, G; Lohse, A W

1998-12-01

Our study demonstrates that antigen-presenting liver sinusoidal endothelial cells (LSEC) induce production of interferon-gamma (IFN-gamma) from cloned Th1 CD4+ T cells. We show that LSEC used the mannose receptor for antigen uptake, which further strengthened the role of LSEC as antigen-presenting cell (APC) population in the liver. The ability of LSEC to activate cloned CD4+ T cells antigen-specifically was down-regulated by exogenous prostaglandin E2 (PGE2) and by IL-10. We identify two separate mechanisms by which IL-10 down-regulated T cell activation through LSEC. IL-10 decreased the constitutive surface expression of MHC class II as well as of the accessory molecules CD80 and CD86 on LSEC. Furthermore, IL-10 diminished mannose receptor activity in LSEC. Decreased antigen uptake via the mannose receptor and decreased expression of accessory molecules may explain the down-regulation of T cell activation through IL-10. Importantly, the expression of low numbers of antigen on MHC II in the absence of accessory signals on LSEC may lead to induction of anergy in T cells. Because PGE2 and IL-10 are released from LSEC or Kupffer cells (KC) in response to those concentrations of endotoxin found physiologically in portal venous blood, it is possible that the continuous presence of these mediators and their negative effect on the local APC may explain the inability of the liver to induce T cell activation and to clear chronic infections. Our results support the notion that antigen presentation by LSEC in the hepatic microenvironment contributes to the observed inability to mount an effective cell-mediated immune response in the liver.

Collective Genetic Interaction Effects and the Role of Antigen Presenting Cells in Autoimmune Diseases

DTIC Science & Technology

2017-01-12

RESEARCH ARTICLE Collective Genetic Interaction Effects and the Role of Antigen-Presenting Cells in Autoimmune Diseases Hyung Jun Woo*, Chenggang Yu...autoimmunity. Genetic predispositions center around the major histocompatibility complex (MHC) class II loci involved in antigen presentation, the key...helper and regulatory T cells showing strong dis- ease-associated interactions with B cells. Our results provide direct genetic evidence point- ing to

The biology and underlying mechanisms of cross-presentation of exogenous antigens on MHC I molecules

PubMed Central

Cruz, Freidrich M.; Colbert, Jeff D.; Merino, Elena; Kriegsman, Barry A.; Rock, Kenneth L.

2017-01-01

To monitor the health of cells, the immune system tasks antigen presenting cells with gathering antigens from other cells and reporting them to CD8 T cells in the form of peptides bound to MHC I molecules. Most cells would be unable to perform this function because they use their MHC I molecules to exclusively present peptides derived from the cell’s own proteins. However, the immune system evolved mechanisms for dendritic cells and some other phagocytes to sample and present antigens from the extracellular milieu on MHC I through a process called cross-presentation (XPT). How this important task is accomplished, its role in health and disease and its potential for exploitation are the subject of this review. PMID:28125356

Podocytes Are Nonhematopoietic Professional Antigen-Presenting Cells

PubMed Central

Burkard, Miriam; Ölke, Martha; Daniel, Christoph; Amann, Kerstin; Hugo, Christian; Kurts, Christian; Steinkasserer, Alexander; Gessner, André

2013-01-01

Podocytes are essential to the structure and function of the glomerular filtration barrier; however, they also exhibit increased expression of MHC class II molecules under inflammatory conditions, and they remove Ig and immune complexes from the glomerular basement membrane (GBM). This finding suggests that podocytes may act as antigen-presenting cells, taking up and processing antigens to initiate specific T cell responses, similar to professional hematopoietic cells such as dendritic cells or macrophages. Here, MHC–antigen complexes expressed exclusively on podocytes of transgenic mice were sufficient to activate CD8+ T cells in vivo. In addition, deleting MHC class II exclusively on podocytes prevented the induction of experimental anti-GBM nephritis. Podocytes ingested soluble and particulate antigens, activated CD4+ T cells, and crosspresented exogenous antigen on MHC class I molecules to CD8+ T cells. In conclusion, podocytes participate in the antigen-specific activation of adaptive immune responses, providing a potential target for immunotherapies of inflammatory kidney diseases and transplant rejection. PMID:23539760

B-cell acquisition of antigen: Sensing the surface.

PubMed

Knight, Andrew M

2015-06-01

B-cell antigen receptor (BCR) recognition and acquisition of antigen by B cells is the essential first step in the generation of effective antibody responses. As B-cell-mediated antigen presentation is also believed to play a significant role in the activation of CD4(+) Th-cell responses, considerable effort has focused on clarifying the nature of antigen/BCR interactions. Following earlier descriptions of interactions of soluble antigens with the BCR, it is now clear that B cells also recognize, physically extract and present antigens that are tethered to, or integral components of, the surfaces or extracellular matrix of other cells. In this issue of the European Journal of Immunology, Zeng et al. [Eur. J. Immunol. 2015. 45: XXXX-XXXX] examine how the physical property or "stiffness" of the surface displaying antigens to B cells influences the B-cell response. This commentary reports that antigen tethered on "less stiff" surfaces induces increased B-cell activation and antibody responses. I then infer how "sensing the surface" by B cells may represent a new component of the immune system's ability to detect "damage," and how this understanding may influence approaches to clinical therapies where immune activity is either unwanted or desired. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Disruption of HLA class II antigen presentation in Burkitt lymphoma: implication of a 47 000 MW acid labile protein in CD4+ T-cell recognition

PubMed Central

God, Jason M; Zhao, Dan; Cameron, Christine A; Amria, Shereen; Bethard, Jennifer R; Haque, Azizul

2014-01-01

While Burkitt lymphoma (BL) has a well-known defect in HLA class I-mediated antigen presentation, the exact role of BL-associated HLA class II in generating a poor CD4+ T-cell response remains unresolved. Here, we found that BL cells are deficient in their ability to optimally stimulate CD4+ T cells via the HLA class II pathway. This defect in CD4+ T-cell recognition was not associated with low levels of co-stimulatory molecules on BL cells, as addition of external co-stimulation failed to elicit CD4+ T-cell activation by BL. Further, the defect was not caused by faulty antigen/class II interaction, because antigenic peptides bound with measurable affinity to BL-associated class II molecules. Interestingly, functional class II–peptide complexes were formed at acidic pH 5·5, which restored immune recognition. Acidic buffer (pH 5·5) eluate from BL cells contained molecules that impaired class II-mediated antigen presentation and CD4+ T-cell recognition. Biochemical analysis showed that these molecules were greater than 30 000 molecular weight in size, and proteinaceous in nature. In addition, BL was found to have decreased expression of a 47 000 molecular weight enolase-like molecule that enhances class II-mediated antigen presentation in B cells, macrophages and dendritic cells, but not in BL cells. These findings demonstrate that BL likely has multiple defects in HLA class II-mediated antigen presentation and immune recognition, which may be exploited for future immunotherapies. PMID:24628049

Modified Vaccinia Virus Ankara-Infected Dendritic Cells Present CD4+ T-Cell Epitopes by Endogenous Major Histocompatibility Complex Class II Presentation Pathways

PubMed Central

Thiele, Frank; Tao, Sha; Zhang, Yi; Muschaweckh, Andreas; Zollmann, Tina; Protzer, Ulrike; Abele, Rubert

2014-01-01

ABSTRACT CD4+ T lymphocytes play a central role in the immune system and mediate their function after recognition of their respective antigens presented on major histocompatibility complex II (MHCII) molecules on antigen-presenting cells (APCs). Conventionally, phagocytosed antigens are loaded on MHCII for stimulation of CD4+ T cells. Certain epitopes, however, can be processed directly from intracellular antigens and are presented on MHCII (endogenous MHCII presentation). Here we characterized the MHCII antigen presentation pathways that are possibly involved in the immune response upon vaccination with modified vaccinia virus Ankara (MVA), a promising live viral vaccine vector. We established CD4+ T-cell lines specific for MVA-derived epitopes as tools for in vitro analysis of MHCII antigen processing and presentation in MVA-infected APCs. We provide evidence that infected APCs are able to directly transfer endogenous viral proteins into the MHCII pathway to efficiently activate CD4+ T cells. By using knockout mice and chemical inhibitory compounds, we further elucidated the molecular basis, showing that among the various subcellular pathways investigated, proteasomes and autophagy are key players in the endogenous MHCII presentation during MVA infection. Interestingly, although proteasomal processing plays an important role, neither TAP nor LAMP-2 was found to be involved in the peptide transport. Defining the molecular mechanism of MHCII presentation during MVA infection provides a basis for improving MVA-based vaccination strategies by aiming for enhanced CD4+ T-cell activation by directing antigens into the responsible pathways. IMPORTANCE This work contributes significantly to our understanding of the immunogenic properties of pathogens by deciphering antigen processing pathways contributing to efficient activation of antigen-specific CD4+ T cells. We identified autophagosome formation, proteasomal activity, and lysosomal integrity as being crucial for endogenous CD4+ T-cell activation. Since poxvirus vectors such as MVA are already used in clinical trials as recombinant vaccines, the data provide important information for the future design of optimized poxviral vaccines for the study of advanced immunotherapy options. PMID:25520512

Effective antigen presentation to helper T cells by human eosinophils.

PubMed

Farhan, Ruhaifah K; Vickers, Mark A; Ghaemmaghami, Amir M; Hall, Andrew M; Barker, Robert N; Walsh, Garry M

2016-12-01

Although eosinophils are inflammatory cells, there is increasing attention on their immunomodulatory roles. For example, murine eosinophils can present antigen to CD4 + T helper (Th) cells, but it remains unclear whether human eosinophils also have this ability. This study determined whether human eosinophils present a range of antigens, including allergens, to activate Th cells, and characterized their expression of MHC class II and co-stimulatory molecules required for effective presentation. Human peripheral blood eosinophils purified from non-allergic donors were pulsed with the antigens house dust mite extract (HDM), Timothy Grass extract (TG) or Mycobacterium tuberculosis purified protein derivative (PPD), before co-culture with autologous CD4 + Th cells. Proliferative and cytokine responses were measured, with eosinophil expression of HLA-DR/DP/DQ and the co-stimulatory molecules CD40, CD80 and CD86 determined by flow cytometry. Eosinophils pulsed with HDM, TG or PPD drove Th cell proliferation, with the response strength dependent on antigen concentration. The cytokine responses varied with donor and antigen, and were not biased towards any particular Th subset, often including combinations of pro- and anti-inflammatory cytokines. Eosinophils up-regulated surface expression of HLA-DR/DP/DQ, CD80, CD86 and CD40 in culture, increases that were sustained over 5 days when incubated with antigens, including HDM, or the major allergens it contains, Der p I or Der p II. Human eosinophils can, therefore, act as effective antigen-presenting cells to stimulate varied Th cell responses against a panel of antigens including HDM, TG or PPD, an ability that may help to determine the development of allergic disease. © 2016 John Wiley & Sons Ltd.

Exposure to sequestered self-antigens in vivo is not sufficient for the induction of autoimmune diabetes

PubMed Central

Chan, Olivia; Hall, Håkan; Elford, Alisha R.; Yen, Patty; Calzascia, Thomas; Spencer, David M.; Ohashi, Pamela S.

2017-01-01

Although the role of T cells in autoimmunity has been explored for many years, the mechanisms leading to the initial priming of an autoimmune T cell response remain enigmatic. The ‘hit and run’ model suggests that self-antigens released upon cell death can provide the initial signal for a self-sustaining autoimmune response. Using a novel transgenic mouse model where we could induce the release of self-antigens via caspase-dependent apoptosis. We tracked the fate of CD8+ T cells specific for the self-antigen. Our studies demonstrated that antigens released from apoptotic cells were cross-presented by CD11c+ cells in the draining lymph node. This cross-presentation led to proliferation of self-antigen specific T cells, followed by a transient ability to produce IFN-γ, but did not lead to the development of autoimmune diabetes. Using this model we examined the consequences on T cell immunity when apoptosis was combined with dendritic cell maturation signals, an autoimmune susceptible genetic background, and the deletion of Tregs. The results of our study demonstrate that autoimmune diabetes cannot be initiated by the presentation of antigens released from apoptotic cells in vivo even in the presence of factors known to promote autoimmunity. PMID:28257518

Antigen Presentation by Individually Transferred HLA Class I Genes in HLA-A, HLA-B, HLA-C Null Human Cell Line Generated Using the Multiplex CRISPR-Cas9 System.

PubMed

Hong, Cheol-Hwa; Sohn, Hyun-Jung; Lee, Hyun-Joo; Cho, Hyun-Il; Kim, Tai-Gyu

Human leukocyte antigens (HLAs) are essential immune molecules that affect transplantation and adoptive immunotherapy. When hematopoietic stem cells or organs are transplanted with HLA-mismatched recipients, graft-versus-host disease or graft rejection can be induced by allogeneic immune responses. The function of each HLA allele has been studied using HLA-deficient cells generated from mutant cell lines or by RNA interference, zinc finger nuclease, and the CRISPR/Cas9 system. To improve HLA gene editing, we attempted to generate an HLA class I null cell line using the multiplex CRISPR/Cas9 system by targeting exons 2 and 3 of HLA-A, HLA-B, and HLA-C genes simultaneously. Multiplex HLA editing could induce the complete elimination of HLA class I genes by bi-allelic gene disruption on target sites which was defined by flow cytometry and target-specific polymerase chain reaction. Furthermore, artificial antigen-presenting cells were generated by transfer of a single HLA class I allele and co-stimulatory molecules into this novel HLA class I null cell line. Artificial antigen-presenting cells showed HLA-restricted antigen presentation following antigen processing and were successfully used for the efficient generation of tumor antigen-specific cytotoxic T cells in vitro. The efficient editing of HLA genes may provide a basis for universal cellular therapies and transplantation.

Adeno-associated virus capsid antigen presentation is dependent on endosomal escape

PubMed Central

Li, Chengwen; He, Yi; Nicolson, Sarah; Hirsch, Matt; Weinberg, Marc S.; Zhang, Ping; Kafri, Tal; Samulski, R. Jude

2013-01-01

Adeno-associated virus (AAV) vectors are attractive for gene delivery-based therapeutics, but data from recent clinical trials have indicated that AAV capsids induce a cytotoxic T lymphocyte (CTL) response that eliminates transduced cells. In this study, we used traditional pharmacological agents and AAV mutants to elucidate the pathway of capsid cross-presentation in AAV-permissive cells. Endosomal acidification inhibitors blocked AAV2 antigen presentation by over 90%, while proteasome inhibitors completely abrogated antigen presentation. Using mutant viruses that are defective for nuclear entry, we observed a 90% decrease in capsid antigen presentation. Different antigen presentation efficiencies were achieved by selectively mutating virion nuclear localization signals. Low antigen presentation was demonstrated with basic region 1 (BR1) mutants, despite relatively high transduction efficiency, whereas there was no difference in antigen presentation between BR2 and BR3 mutants defective for transduction, as compared with wild-type AAV2. These results suggest that effective AAV2 capsid antigen presentation is dependent on AAV virion escape from the endosome/lysosome for antigen degradation by proteasomes, but is independent of nuclear uncoating. These results should facilitate the design of effective strategies to evade capsid-specific CTL-mediated elimination of AAV-transduced target cells in future clinical trials. PMID:23454772

Carbohydrates and T cells: A sweet twosome

PubMed Central

Avci, Fikri Y.; Li, Xiangming; Tsuji, Moriya; Kasper, Dennis L.

2013-01-01

Carbohydrates as T cell-activating antigens have been generating significant interest. For many years, carbohydrates were thought of as T-independent antigens, however, more recent research had demonstrated that mono- or oligosaccharides glycosidically-linked to peptides can be recognized by T cells. T cell recognition of these glycopeptides depends on the structure of both peptide and glycan portions of the antigen. Subsequently, it was discovered that natural killer T cells recognized glycolipids when presented by the antigen presenting molecule CD1d. A transformative insight into glycan-recognition by T cells occurred when zwitterionic polysaccharides were discovered to bind to and be presented by MHCII to CD4+ T cells. Based on this latter observation, the role that carbohydrate epitopes generated from glycoconjugate vaccines had in activating helper T cells was explored and it was found that these epitopes are presented to specific carbohydrate recognizing T cells through a unique mechanism. Here we review the key interactions between carbohydrate antigens and the adaptive immune system at the molecular, cellular and systems levels exploring the significant biological implications in health and disease. PMID:23757291

Co-ordination of incoming and outgoing traffic in antigen-presenting cells by pattern recognition receptors and T cells.

PubMed

Nair, Priyanka; Amsen, Derk; Blander, J Magarian

2011-12-01

Dendritic cells are innate sentinels of the immune system and potent activators of naÏve T cells. Mechanisms must exist to enable these cells to achieve maximal activation of T cells specific for microbial antigens, while avoiding activation of T cells specific for self-antigens. Here we discuss how a combination of signals from pattern recognition receptors and T cells co-ordinates subcellular trafficking of antigen with both major histocompatibility complex class I and class II molecules and T-cell costimulatory molecules, resulting in the preferential presentation of microbial peptides within a stimulatory context. © 2011 John Wiley & Sons A/S.

Dendritic cells in oral tolerance in the gut.

PubMed

Rescigno, Maria

2011-09-01

Oral tolerance is a process that allows generation of systemic unresponsiveness to food antigens. Hence if the same antigen is introduced systemically even under immunogenic conditions it does not induce immune responsiveness. Dendritic cells (DCs) have been identified as essential players in this process. DCs in the gut are located in a strategic position as they can interact directly with luminal antigens or indirectly after their transcytosis across epithelial cells. DCs can then migrate to associated lymphoid tissues to induce tolerance. Antigen presenting cells in the gut are specialized in function and have divided their labour so that there are cells capable to migrate to the draining mesenteric lymph node for induction of T regulatory cells, while other subsets are resident and are required to enforce tolerance locally in the gut after food antigen exposure. In this review, I shall summarize the characteristics of antigen presenting cells in the gut and their involvement in oral tolerance induction. In addition, I will also emphasize that tolerance to food allergens may be contributed by plasmacytoid DCs in the liver that participate to the elimination or anergy of allergen-specific CD8 T cells. Hence specialized functions are associated to different subsets of antigen presenting cells and different organs. © 2011 Blackwell Publishing Ltd.

CD4+ T cell-mediated rejection of MHC class II-positive tumor cells is dependent on antigen secretion and indirect presentation on host APCs.

PubMed

Haabeth, Ole Audun Werner; Fauskanger, Marte; Manzke, Melanie; Lundin, Katrin U; Corthay, Alexandre; Bogen, Bjarne; Tveita, Anders Aune

2018-05-11

Tumor-specific CD4+ T cells have been shown to mediate efficient anti-tumor immune responses against cancer. Such responses can occur through direct binding to MHC class II (MHC II)-expressing tumor cells or indirectly via activation of professional antigen-presenting cells (APC) that take up and present the tumor antigen. We have previously shown that CD4+ T cells reactive against an epitope within the Ig light chain variable region of a murine B cell lymphoma can reject established tumors. Given the presence of MHC II molecules at the surface of lymphoma cells, we investigated whether MHC II-restricted antigen presentation on tumor cells alone was required for rejection. Variants of the A20 B lymphoma cell line that either secreted or intracellularly retained different versions of the tumor-specific antigen revealed that antigen secretion by the MHC II-expressing tumor cells was essential both for the priming and effector phase of CD4+ T cell-driven anti-tumor immune responses. Consistent with this, genetic ablation of MHC II in tumor cells, both in the case of B lymphoma and B16 melanoma, did not preclude rejection of tumors by tumor antigen-specific CD4+ T cells in vivo. These findings demonstrate that MHC class II expression on tumor cells themselves is not required for CD4+ T cell-mediated rejection, and that indirect display on host APC is sufficient for effective tumor elimination. These results support the importance of tumor-infiltrating APC as mediators of tumor cell killing by CD4+ T cells. Copyright ©2018, American Association for Cancer Research.

Processing of urushiol (poison ivy) hapten by both endogenous and exogenous pathways for presentation to T cells in vitro.

PubMed

Kalish, R S; Wood, J A; LaPorte, A

1994-05-01

The antigen processing requirements for urushiol, the immunogen of poison ivy (Toxicodendron radicans), were tested by presentation of urushiol to cultured human urushiol-responsive T cells. Urushiol was added to antigen-presenting cells (APC) either before or after fixation with paraformaldehyde. Three distinct routes of antigen processing were detected. CD8+ and CD4+ T cells, which were dependent upon processing, proliferated if urushiol was added to APC before fixation, but did not proliferate when urushiol was added to APC after fixation. Processing of urushiol for presentation to CD8+ T cells was inhibited by azide, monensin, and brefeldin A. This suggests that urushiol was processed by the endogenous pathway. In contrast, presentation of urushiol to CD4+ T cells was inhibited by monensin but not by brefeldin A. This was compatible with antigen processing by the endosomal (exogenous) pathway. Finally, certain CD8+ T cells recognized urushiol in the absence of processing. These cells proliferated in response to APC incubated with urushiol after fixation. Classification of contact allergens by antigen processing pathway may predict the relative roles of CD4+ and CD8+ cells in the immunopathogensis of allergic contact dermatitis.

Processing of urushiol (poison ivy) hapten by both endogenous and exogenous pathways for presentation to T cells in vitro.

PubMed Central

Kalish, R S; Wood, J A; LaPorte, A

1994-01-01

The antigen processing requirements for urushiol, the immunogen of poison ivy (Toxicodendron radicans), were tested by presentation of urushiol to cultured human urushiol-responsive T cells. Urushiol was added to antigen-presenting cells (APC) either before or after fixation with paraformaldehyde. Three distinct routes of antigen processing were detected. CD8+ and CD4+ T cells, which were dependent upon processing, proliferated if urushiol was added to APC before fixation, but did not proliferate when urushiol was added to APC after fixation. Processing of urushiol for presentation to CD8+ T cells was inhibited by azide, monensin, and brefeldin A. This suggests that urushiol was processed by the endogenous pathway. In contrast, presentation of urushiol to CD4+ T cells was inhibited by monensin but not by brefeldin A. This was compatible with antigen processing by the endosomal (exogenous) pathway. Finally, certain CD8+ T cells recognized urushiol in the absence of processing. These cells proliferated in response to APC incubated with urushiol after fixation. Classification of contact allergens by antigen processing pathway may predict the relative roles of CD4+ and CD8+ cells in the immunopathogensis of allergic contact dermatitis. Images PMID:7910172

Carbohydrates as T-cell antigens with implications in health and disease.

PubMed

Sun, Lina; Middleton, Dustin R; Wantuch, Paeton L; Ozdilek, Ahmet; Avci, Fikri Y

2016-10-01

Glycosylation is arguably the most ubiquitous post-translational modification on proteins in microbial and mammalian cells. During the past few years, there has been intensive research demonstrating that carbohydrates, either in pure forms or in conjunction with proteins or lipids, evoke and modulate adaptive immune responses. We now know that carbohydrates can be directly recognized by T cells or participate in T-cell stimulation as components of T-cell epitopes. T-cell recognition of carbohydrate antigens takes place via their presentation by major histocompatibility complex pathways on antigen-presenting cells. In this review, we summarize studies on carbohydrates as T-cell antigens modulating adaptive immune responses. Through discussion of glycan-containing antigens, such as glycoproteins, glycolipids, zwitterionic polysaccharides and carbohydrate-based glycoconjugate vaccines, we will illustrate the key molecular and cellular interactions between carbohydrate antigens and T cells and the implications of these interactions in health and disease. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Ubiquitinated Proteins Isolated From Tumor Cells Are Efficient Substrates for Antigen Cross-Presentation.

PubMed

Yu, Guangjie; Moudgil, Tarsem; Cui, Zhihua; Mou, Yongbin; Wang, Lixin; Fox, Bernard A; Hu, Hong-Ming

2017-06-01

We have previously shown that inhibition of the proteasome causes defective ribosomal products to be shunted into autophagosomes and subsequently released from tumor cells as defective ribosomal products in Blebs (DRibbles). These DRibbles serve as an excellent source of antigens for cross-priming of tumor-specific T cells. Here, we examine the role of ubiquitinated proteins (Ub-proteins) in this pathway. Using purified Ub-proteins from tumor cells that express endogenous tumor-associated antigen or exogenous viral antigen, we tested the ability of these proteins to stimulate antigen-specific T-cell responses, by activation of monocyte-derived dendritic cells generated from human peripheral blood mononuclear cells. Compared with total cell lysates, we found that purified Ub-proteins from both a gp100-specific melanoma cell line and from a lung cancer cell line expressing cytomegalovirus pp65 antigen produced a significantly higher level of IFN-γ in gp100- or pp65-specific T cells, respectively. In addition, Ub-proteins from an allogeneic tumor cell line could be used to stimulate tumor-infiltrating lymphocytes isolated and expanded from non-small cell lung cancer patients. These results establish that Ub-proteins provide a relevant source of antigens for cross-priming of antitumor immune responses in a variety of settings, including endogenous melanoma and exogenous viral antigen presentation, as well as antigen-specific tumor-infiltrating lymphocytes. Thus, ubiquitin can be used as an affinity tag to enrich for unknown tumor-specific antigens from tumor cell lysates to stimulate tumor-specific T cells ex vivo or to be used as vaccines to target short-lived proteins.

Immunoglobulins drive terminal maturation of splenic dendritic cells

PubMed Central

Ziętara, Natalia; Łyszkiewicz, Marcin; Puchałka, Jacek; Pei, Gang; Gutierrez, Maximiliano Gabriel; Lienenklaus, Stefan; Hobeika, Elias; Reth, Michael; Martins dos Santos, Vitor A. P.; Krueger, Andreas; Weiss, Siegfried

2013-01-01

Nature and physiological status of antigen-presenting cells, such as dendritic cells DCs, are decisive for the immune reactions elicited. Multiple factors and cell interactions have been described that affect maturation of DCs. Here, we show that DCs arising in the absence of immunoglobulins (Ig) in vivo are impaired in cross-presentation of soluble antigen. This deficiency was due to aberrant cellular targeting of antigen to lysosomes and its rapid degradation. Function of DCs could be restored by transfer of Ig irrespective of antigen specificity and isotype. Modulation of cross-presentation by Ig was inhibited by coapplication of mannan and, thus, likely to be mediated by C-type lectin receptors. This unexpected dependency of splenic DCs on Ig to cross-present antigen provides insights into the interplay between cellular and humoral immunity and the immunomodulatory capacity of Ig. PMID:23345431

Probing the Effector and Suppressive Functions of Human T Cell Subsets Using Antigen-Specific Engineered T Cell Receptors

PubMed Central

Imberg, Keren; Mercer, Frances; Zhong, Shi; Krogsgaard, Michelle; Unutmaz, Derya

2013-01-01

Activation of T cells through the engagement of the T cell receptors (TCRs) with specific peptide-MHC complexes on antigen presenting cells (APCs) is the major determinant for their proliferation, differentiation and display of effector functions. To assess the role of quantity and quality of peptide-MHC presentation in eliciting T cell activation and suppression functions, we genetically engineered human T cells with two TCRs that recognize HLA-A*0201-restricted peptides derived from either HIV or melanoma antigens. The engineered-TCRs are highly functional in both CD8+ and CD4+ T cells as assessed by the upregulation of activation markers, induction of cytokine secretion and cytotoxicity. We further demonstrated that engineered-TCRs can also be expressed on naïve human T cells, which are stimulated through APCs presenting specific peptides to induce T cell proliferation and acquire effector functions. Furthermore, regulatory T cells (Tregs) ectopically expressing the engineered-TCRs are activated in an antigen-specific fashion and suppress T cell proliferation. In this system, the inhibitory activity of peptide-stimulated Tregs require the presence of dendritic cells (DCs) in the culture, either as presenters or as bystander cells, pointing to a critical role for DCs in suppression by Tregs. In conclusion, the engineered-TCR system reported here advances our ability to understand the differentiation pathways of naïve T cells into antigen-specific effector cells and the role of antigen-specific signaling in Treg-mediated immune suppression. PMID:23437112

Vaccinia virus decreases major histocompatibility complex (MHC) class II antigen presentation, T-cell priming, and peptide association with MHC class II

PubMed Central

Rehm, Kristina E; Connor, Ramsey F; Jones, Gwendolyn J B; Yimbu, Kenneth; Mannie, Mark D; Roper, Rachel L

2009-01-01

Vaccinia virus (VACV) is the current live virus vaccine used to protect humans against smallpox and monkeypox, but its use is contraindicated in several populations because of its virulence. It is therefore important to elucidate the immune evasion mechanisms of VACV. We found that VACV infection of antigen-presenting cells (APCs) significantly decreased major histocompatibility complex (MHC) II antigen presentation and decreased synthesis of 13 chemokines and cytokines, suggesting a potent viral mechanism for immune evasion. In these model systems, responding T cells were not directly affected by virus, indicating that VACV directly affects the APC. VACV significantly decreased nitric oxide production by peritoneal exudate cells and the RAW macrophage cell line in response to lipopolysaccharide (LPS) and interferon (IFN)-γ, decreased class II MHC expression on APCs, and induced apoptosis in macrophages and dendritic cells. However, VACV decreased antigen presentation by 1153 B cells without apparent apoptosis induction, indicating that VACV differentially affects B lymphocytes and other APCs. We show that the key mechanism of VACV inhibition of antigen presentation may be its reduction of antigenic peptide loaded into the cleft of MHC class II molecules. These data indicate that VACV evades the host immune response by impairing critical functions of the APC. PMID:20067538

Enhanced Direct Major Histocompatibility Complex Class I Self-Antigen Presentation Induced by Chlamydia Infection

PubMed Central

Cram, Erik D.; Simmons, Ryan S.; Palmer, Amy L.; Hildebrand, William H.; Rockey, Daniel D.

2015-01-01

The direct major histocompatibility complex (MHC) class I antigen presentation pathway ensures intracellular peptides are displayed at the cellular surface for recognition of infected or transformed cells by CD8+ cytotoxic T lymphocytes. Chlamydia spp. are obligate intracellular bacteria and, as such, should be targeted by CD8+ T cells. It is likely that Chlamydia spp. have evolved mechanisms to avoid the CD8+ killer T cell responses by interfering with MHC class I antigen presentation. Using a model system of self-peptide presentation which allows for posttranslational control of the model protein's stability, we tested the ability of various Chlamydia species to alter direct MHC class I antigen presentation. Infection of the JY lymphoblastoid cell line limited the accumulation of a model host protein and increased presentation of the model-protein-derived peptides. Enhanced self-peptide presentation was detected only when presentation was restricted to defective ribosomal products, or DRiPs, and total MHC class I levels remained unaltered. Skewed antigen presentation was dependent on a bacterial synthesized component, as evidenced by reversal of the observed phenotype upon preventing bacterial transcription, translation, and the inhibition of bacterial lipooligosaccharide synthesis. These data suggest that Chlamydia spp. have evolved to alter the host antigen presentation machinery to favor presentation of defective and rapidly degraded forms of self-antigen, possibly as a mechanism to diminish the presentation of peptides derived from bacterial proteins. PMID:26597986

Roles of lymphatic endothelial cells expressing peripheral tissue antigens in CD4 T-cell tolerance induction.

PubMed

Rouhani, Sherin J; Eccles, Jacob D; Riccardi, Priscila; Peske, J David; Tewalt, Eric F; Cohen, Jarish N; Liblau, Roland; Mäkinen, Taija; Engelhard, Victor H

2015-04-10

Lymphatic endothelial cells (LECs) directly express peripheral tissue antigens and induce CD8 T-cell deletional tolerance. LECs express MHC-II molecules, suggesting they might also tolerize CD4 T cells. We demonstrate that when β-galactosidase (β-gal) is expressed in LECs, β-gal-specific CD8 T cells undergo deletion via the PD-1/PD-L1 and LAG-3/MHC-II pathways. In contrast, LECs do not present endogenous β-gal in the context of MHC-II molecules to β-gal-specific CD4 T cells. Lack of presentation is independent of antigen localization, as membrane-bound haemagglutinin and I-Eα are also not presented by MHC-II molecules. LECs express invariant chain and cathepsin L, but not H2-M, suggesting that they cannot load endogenous antigenic peptides onto MHC-II molecules. Importantly, LECs transfer β-gal to dendritic cells, which subsequently present it to induce CD4 T-cell anergy. Therefore, LECs serve as an antigen reservoir for CD4 T-cell tolerance, and MHC-II molecules on LECs are used to induce CD8 T-cell tolerance via LAG-3.

Roles of lymphatic endothelial cells expressing peripheral tissue antigens in CD4 T-cell tolerance induction

PubMed Central

Rouhani, Sherin J.; Eccles, Jacob D.; Riccardi, Priscila; Peske, J. David; Tewalt, Eric F.; Cohen, Jarish N.; Liblau, Roland; Mäkinen, Taija; Engelhard, Victor H.

2015-01-01

Lymphatic endothelial cells (LECs) directly express peripheral tissue antigens and induce CD8 T-cell deletional tolerance. LECs express MHC-II molecules, suggesting they might also tolerize CD4 T cells. We demonstrate that when β-galactosidase (β-gal) is expressed in LECs, β-gal-specific CD8 T cells undergo deletion via the PD-1/PD-L1 and LAG-3/MHC-II pathways. In contrast, LECs do not present endogenous β-gal in the context of MHC-II molecules to β-gal-specific CD4 T cells. Lack of presentation is independent of antigen localization, as membrane-bound haemagglutinin and I-Eα are also not presented by MHC-II molecules. LECs express invariant chain and cathepsin L, but not H2-M, suggesting that they cannot load endogenous antigenic peptides onto MHC-II molecules. Importantly, LECs transfer β-gal to dendritic cells, which subsequently present it to induce CD4 T-cell anergy. Therefore, LECs serve as an antigen reservoir for CD4 T-cell tolerance, and MHC-II molecules on LECs are used to induce CD8 T-cell tolerance via LAG-3. PMID:25857745

Pivotal Advance: Peritoneal cavity B-1 B cells have phagocytic and microbicidal capacities and present phagocytosed antigen to CD4+ T cells

PubMed Central

Parra, David; Rieger, Aja M.; Li, Jun; Zhang, Yong-An; Randall, Louise M.; Hunter, Christopher A.; Barreda, Daniel R.; Sunyer, J. Oriol

2012-01-01

Breaking the long-held paradigm that primary B cells are not phagocytic, several studies have demonstrated recently that B cells from fish, amphibians, and reptilians have a significant phagocytic capacity. Whether such capacity has remained conserved in certain mammalian B cell subsets is presently an enigma. Here, we report a previously unrecognized ability of PerC B-1a and B-1b lymphocytes to phagocytose latex beads and bacteria. In contrast, B-2 lymphocytes had an almost negligible ability to internalize these particles. Upon phagocytosis, B-1a and B-1b cells were able to mature their phagosomes into phagolysosomes and displayed the ability to kill internalized bacteria. Importantly, B-1a and B-1b cells effectively present antigen recovered from phagocytosed particles to CD4+ T cells. However, these cells showed a much lower competence to present soluble antigen or antigen from large, noninternalized particles. B-1 B cells presented particulate and soluble antigen to CD4+ T cells more efficiently than macrophages, whereas DCs were the most potent APCs. The novel phagocytic and microbicidal abilities identified in B-1 B lymphocytes strengthen the innate nature that has long been attributed to these cells. In the context of adaptive immunity, we show that these innate immune processes are relevant, as they enable B-1 B cells to present phagocytosable particulate antigen. These capacities position these cells at the crossroads that link innate with adaptive immune processes. In a broader context, these newly identified capacities of B-1 B cells further support the previously recognized functional, developmental, and evolutionary relationships between these cells and macrophages. PMID:22058420

Pivotal advance: peritoneal cavity B-1 B cells have phagocytic and microbicidal capacities and present phagocytosed antigen to CD4+ T cells.

PubMed

Parra, David; Rieger, Aja M; Li, Jun; Zhang, Yong-An; Randall, Louise M; Hunter, Christopher A; Barreda, Daniel R; Sunyer, J Oriol

2012-04-01

Breaking the long-held paradigm that primary B cells are not phagocytic, several studies have demonstrated recently that B cells from fish, amphibians, and reptilians have a significant phagocytic capacity. Whether such capacity has remained conserved in certain mammalian B cell subsets is presently an enigma. Here, we report a previously unrecognized ability of PerC B-1a and B-1b lymphocytes to phagocytose latex beads and bacteria. In contrast, B-2 lymphocytes had an almost negligible ability to internalize these particles. Upon phagocytosis, B-1a and B-1b cells were able to mature their phagosomes into phagolysosomes and displayed the ability to kill internalized bacteria. Importantly, B-1a and B-1b cells effectively present antigen recovered from phagocytosed particles to CD4(+) T cells. However, these cells showed a much lower competence to present soluble antigen or antigen from large, noninternalized particles. B-1 B cells presented particulate and soluble antigen to CD4(+) T cells more efficiently than macrophages, whereas DCs were the most potent APCs. The novel phagocytic and microbicidal abilities identified in B-1 B lymphocytes strengthen the innate nature that has long been attributed to these cells. In the context of adaptive immunity, we show that these innate immune processes are relevant, as they enable B-1 B cells to present phagocytosable particulate antigen. These capacities position these cells at the crossroads that link innate with adaptive immune processes. In a broader context, these newly identified capacities of B-1 B cells further support the previously recognized functional, developmental, and evolutionary relationships between these cells and macrophages.

Antitumor immune responses mediated by dendritic cells

PubMed Central

Spel, Lotte; Boelens, Jaap-Jan; Nierkens, Stefan; Boes, Marianne

2013-01-01

Dendritic cells (DCs) are essential for the induction of adaptive immune responses against malignant cells by virtue of their capacity to effectively cross-present exogenous antigens to T lymphocytes. Dying cancer cells are indeed a rich source of antigens that may be harnessed for the development of DC-based vaccines. In particular, malignant cells succumbing to apoptosis, rather than necrosis, appear to release antigens in a manner that allows for the elicitation of adaptive immune responses. In this review, we describe the processes that mediate the cross-presentation of antigens released by apoptotic cancer cells to CD8+ T lymphocytes, resulting in the activation of protective tumor-specific immune responses. PMID:24482744

Pros and Cons of Antigen-Presenting Cell Targeted Tumor Vaccines.

PubMed

Goyvaerts, Cleo; Breckpot, Karine

2015-01-01

In therapeutic antitumor vaccination, dendritic cells play the leading role since they decide if, how, when, and where a potent antitumor immune response will take place. Since the disentanglement of the complexity and merit of different antigen-presenting cell subtypes, antitumor immunotherapeutic research started to investigate the potential benefit of targeting these subtypes in situ. This review will discuss which antigen-presenting cell subtypes are at play and how they have been targeted and finally question the true meaning of targeting antitumor-based vaccines.

Alpha-alumina nanoparticles induce efficient autophagy-dependent cross-presentation and potent antitumour response

NASA Astrophysics Data System (ADS)

Li, Haiyan; Li, Yuhuan; Jiao, Jun; Hu, Hong-Ming

2011-10-01

Therapeutic cancer vaccination is an attractive strategy because it induces T cells of the immune system to recognize and kill tumour cells in cancer patients. However, it remains difficult to generate large numbers of T cells that can recognize the antigens on cancer cells using conventional vaccine carrier systems. Here we show that α-Al2O3 nanoparticles can act as an antigen carrier to reduce the amount of antigen required to activate T cells in vitro and in vivo. We found that α-Al2O3 nanoparticles delivered antigens to autophagosomes in dendritic cells, which then presented the antigens to T cells through autophagy. Immunization of mice with α-Al2O3 nanoparticles that are conjugated to either a model tumour antigen or autophagosomes derived from tumour cells resulted in tumour regression. These results suggest that α-Al2O3 nanoparticles may be a promising adjuvant in the development of therapeutic cancer vaccines.

Spatiotemporally and Mechanically Controlled Triggering of Mast Cells using Atomic Force Microscopy

PubMed Central

Hu, Kenneth K.; Bruce, Marc A.; Butte, Manish J.

2014-01-01

Mast cells are thought to be sensitive to mechanical forces, for example, coughing in asthma or pressure in “physical urticarias”. Conversion of mechanical forces to biochemical signals could potentially augment antigenic signaling. Studying the combined effects of mechanical and antigenic cues on mast cells and other hematopoietic cells has been elusive. Here, we present an approach using a modified atomic force microscope cantilever to deliver antigenic signals to mast cells while simultaneously applying mechanical forces. We developed a strategy to concurrently record degranulation events by fluorescence microscopy during antigenic triggering. Finally, we also measured the mechanical forces generated by mast cells while antigen receptors are ligated. We showed that mast cells respond to antigen delivered by the AFM cantilever with prompt degranulation and the generation of strong pushing and pulling forces. We did not discern any relationship between applied mechanical forces and the kinetics of degranulation. These experiments present a new method for dissecting the interactions of mechanical and biochemical cues in signaling responses of immune cells. PMID:24777418

Identifying Regulators of the Immune Response to Dying Cells | Center for Cancer Research

Cancer.gov

Cytotoxic T cells are responsible for carrying out antigen-mediated immune responses against virally-infected and malignant cells. In both cases, cytotoxic T cells are stimulated by interacting with antigen presenting cells, such as dendritic cells (DCs). Infected cells produce virus-specific antigens and pathogen associated molecular patterns, which are recognized by DCs and

Efficient Capsid Antigen Presentation From Adeno-Associated Virus Empty Virions In Vivo.

PubMed

Pei, Xiaolei; Earley, Lauriel Freya; He, Yi; Chen, Xiaojing; Hall, Nikita Elexa; Samulski, Richard Jude; Li, Chengwen

2018-01-01

Adeno-associated virus (AAV) vectors have been successfully applied in clinical trials for hemophilic patients. Although promising, the clinical results suggest that the capsid-specific CD8+T cell response has a negative effect on therapeutic success. In an in vitro analysis using an engineered AAV virus carrying immune-dominant SIINFEKL peptide in the capsid backbone, we have previously demonstrated that capsid antigen presentation from full (genome containing) AAV capsids requires endosome escape and is proteasome dependent and that no capsid antigen presentation is induced from empty virions. In the present study, we examined capsid antigen presentation from administration of empty virions in animal models. In wild-type mice, similar to AAV full particles, capsid antigen presentation from AAV empty virion infection was dose dependent, and the kinetics studies showed that antigen presentation was detected from 2 to 40 days after AAV empty virion administration. In the transporter associated with antigen processing 1 deficient (TAP-/-) mice, capsid antigen presentation was inhibited from both AAV full and empty virions, but higher inhibition was achieved from AAV full particle administration than that from empty virions. This indicates that the pathway of capsid antigen presentation from AAV transduction is dependent on proteasome-mediated degradation of AAV capsids (mainly for full particles) and that the endosomal pathway may also play a role in antigen presentation from empty particles but not full virions. The capsid antigen presentation efficiency from AAV preparations was positively correlated with the amount of empty virions contaminated with full particles. Collectively, the results indicate that contamination of AAV empty virions induces efficient antigen presentation in vivo and the mechanism of capsid antigen presentation from empty virions involves both endosomal and proteasomal pathways. The elucidation of capsid antigen presentation from AAV empty virions may allow us to rationally design effective strategies to prevent elimination of AAV transduced target cells by capsid specific CD8+ T cells.

Efficient Capsid Antigen Presentation From Adeno-Associated Virus Empty Virions In Vivo

PubMed Central

Pei, Xiaolei; Earley, Lauriel Freya; He, Yi; Chen, Xiaojing; Hall, Nikita Elexa; Samulski, Richard Jude; Li, Chengwen

2018-01-01

Adeno-associated virus (AAV) vectors have been successfully applied in clinical trials for hemophilic patients. Although promising, the clinical results suggest that the capsid-specific CD8+T cell response has a negative effect on therapeutic success. In an in vitro analysis using an engineered AAV virus carrying immune-dominant SIINFEKL peptide in the capsid backbone, we have previously demonstrated that capsid antigen presentation from full (genome containing) AAV capsids requires endosome escape and is proteasome dependent and that no capsid antigen presentation is induced from empty virions. In the present study, we examined capsid antigen presentation from administration of empty virions in animal models. In wild-type mice, similar to AAV full particles, capsid antigen presentation from AAV empty virion infection was dose dependent, and the kinetics studies showed that antigen presentation was detected from 2 to 40 days after AAV empty virion administration. In the transporter associated with antigen processing 1 deficient (TAP−/−) mice, capsid antigen presentation was inhibited from both AAV full and empty virions, but higher inhibition was achieved from AAV full particle administration than that from empty virions. This indicates that the pathway of capsid antigen presentation from AAV transduction is dependent on proteasome-mediated degradation of AAV capsids (mainly for full particles) and that the endosomal pathway may also play a role in antigen presentation from empty particles but not full virions. The capsid antigen presentation efficiency from AAV preparations was positively correlated with the amount of empty virions contaminated with full particles. Collectively, the results indicate that contamination of AAV empty virions induces efficient antigen presentation in vivo and the mechanism of capsid antigen presentation from empty virions involves both endosomal and proteasomal pathways. The elucidation of capsid antigen presentation from AAV empty virions may allow us to rationally design effective strategies to prevent elimination of AAV transduced target cells by capsid specific CD8+ T cells. PMID:29725339

Dendritic cell targeted vaccines: Recent progresses and challenges

PubMed Central

Chen, Pengfei; Liu, Xinsheng; Sun, Yuefeng; Zhou, Peng; Wang, Yonglu; Zhang, Yongguang

2016-01-01

ABSTRACT Dendritic cells (DCs) are known to be a set of morphology, structure and function of heterogeneous professional antigen presenting cells (APCs), as well as the strongest functional antigen presenting cells, which can absorb, process and present antigens. As the key regulators of innate and adaptive immune responses, DCs are at the center of the immune system and capable of interacting with both B cells and T cells, thereby manipulating the humoral and cellular immune responses. DCs provide an essential link between the innate and adaptive immunity, and the strong immune activation function of DCs and their properties of natural adjuvants, make them a valuable target for antigen delivery. Targeting antigens to DC-specific endocytic receptors in combination with the relevant antibodies or ligands along with immunostimulatory adjuvants has been recently recognized as a promising strategy for designing an effective vaccine that elicits a strong and durable T cell response against intracellular pathogens and cancer. This opinion article provides a brief summary of the rationales, superiorities and challenges of existing DC-targeting approaches. PMID:26513200

Antibody-Antigen-Adjuvant Conjugates Enable Co-Delivery of Antigen and Adjuvant to Dendritic Cells in Cis but Only Have Partial Targeting Specificity

PubMed Central

Abuknesha, Ram; Uematsu, Satoshi; Akira, Shizuo; Nestle, Frank O.; Diebold, Sandra S.

2012-01-01

Antibody-antigen conjugates, which promote antigen-presentation by dendritic cells (DC) by means of targeted delivery of antigen to particular DC subsets, represent a powerful vaccination approach. To ensure immunity rather than tolerance induction the co-administration of a suitable adjuvant is paramount. However, co-administration of unlinked adjuvant cannot ensure that all cells targeted by the antibody conjugates are appropriately activated. Furthermore, antigen-presenting cells (APC) that do not present the desired antigen are equally strongly activated and could prime undesired responses against self-antigens. We, therefore, were interested in exploring targeted co-delivery of antigen and adjuvant in cis in form of antibody-antigen-adjuvant conjugates for the induction of anti-tumour immunity. In this study, we report on the assembly and characterization of conjugates consisting of DEC205-specific antibody, the model antigen ovalbumin (OVA) and CpG oligodeoxynucleotides (ODN). We show that such conjugates are more potent at inducing cytotoxic T lymphocyte (CTL) responses than control conjugates mixed with soluble CpG. However, our study also reveals that the nucleic acid moiety of such antibody-antigen-adjuvant conjugates alters their binding and uptake and allows delivery of the antigen and the adjuvant to cells partially independently of DEC205. Nevertheless, antibody-antigen-adjuvant conjugates are superior to antibody-free antigen-adjuvant conjugates in priming CTL responses and efficiently induce anti-tumour immunity in the murine B16 pseudo-metastasis model. A better understanding of the role of the antibody moiety is required to inform future conjugate vaccination strategies for efficient induction of anti-tumour responses. PMID:22808118

Immunization Route Dictates Cross-Priming Efficiency and Impacts the Optimal Timing of Adjuvant Delivery

PubMed Central

Bouvier, Isabelle; Jusforgues-Saklani, Hélène; Lim, Annick; Lemaître, Fabrice; Lemercier, Brigitte; Auriau, Charlotte; Nicola, Marie-Anne; Leroy, Sandrine; Law, Helen K.; Bandeira, Antonio; Moon, James J.; Bousso, Philippe; Albert, Matthew L.

2011-01-01

Delivery of cell-associated antigen represents an important strategy for vaccination. While many experimental models have been developed in order to define the critical parameters for efficient cross-priming, few have utilized quantitative methods that permit the study of the endogenous repertoire. Comparing different strategies of immunization, we report that local delivery of cell-associated antigen results in delayed T cell cross-priming due to the increased time required for antigen capture and presentation. In comparison, delivery of disseminated antigen resulted in rapid T cell priming. Surprisingly, local injection of cell-associated antigen, while slower, resulted in the differentiation of a more robust, polyfunctional, effector response. We also evaluated the combination of cell-associated antigen with poly I:C delivery and observed an immunization route-specific effect regarding the optimal timing of innate immune stimulation. These studies highlight the importance of considering the timing and persistence of antigen presentation, and suggest that intradermal injection with delayed adjuvant delivery is the optimal strategy for achieving CD8+ T cell cross-priming. PMID:22566860

TNF-induced target cell killing by CTL activated through cross-presentation.

PubMed

Wohlleber, Dirk; Kashkar, Hamid; Gärtner, Katja; Frings, Marianne K; Odenthal, Margarete; Hegenbarth, Silke; Börner, Carolin; Arnold, Bernd; Hämmerling, Günter; Nieswandt, Bernd; van Rooijen, Nico; Limmer, Andreas; Cederbrant, Karin; Heikenwalder, Mathias; Pasparakis, Manolis; Protzer, Ulrike; Dienes, Hans-Peter; Kurts, Christian; Krönke, Martin; Knolle, Percy A

2012-09-27

Viruses can escape cytotoxic T cell (CTL) immunity by avoiding presentation of viral components via endogenous MHC class I antigen presentation in infected cells. Cross-priming of viral antigens circumvents such immune escape by allowing noninfected dendritic cells to activate virus-specific CTLs, but they remain ineffective against infected cells in which immune escape is functional. Here, we show that cross-presentation of antigen released from adenovirus-infected hepatocytes by liver sinusoidal endothelial cells stimulated cross-primed effector CTLs to release tumor necrosis factor (TNF), which killed virus-infected hepatocytes through caspase activation. TNF receptor signaling specifically eliminated infected hepatocytes that showed impaired anti-apoptotic defense. Thus, CTL immune surveillance against infection relies on two similarly important but distinct effector functions that are both MHC restricted, requiring either direct antigen recognition on target cells and canonical CTL effector function or cross-presentation and a noncanonical effector function mediated by TNF. Copyright © 2012 The Authors. Published by Elsevier Inc. All rights reserved.

The Role of FcRn in Antigen Presentation

PubMed Central

Baker, Kristi; Rath, Timo; Pyzik, Michal; Blumberg, Richard S.

2014-01-01

Immunoglobulins are unique molecules capable of simultaneously recognizing a diverse array of antigens and themselves being recognized by a broad array of receptors. The abundance specifically of the IgG subclass and the variety of signaling receptors to which it binds render this an important immunomodulatory molecule. In addition to the classical Fcγ receptors that bind IgG at the cell surface, the neonatal Fc receptor (FcRn) is a lifelong resident of the endolysosomal system of most hematopoietic cells where it determines the intracellular fate of both IgG and IgG-containing immune complexes (IgG IC). Cross-linking of FcRn by multivalent IgG IC within antigen presenting cells such as dendritic cells initiates specific mechanisms that result in trafficking of the antigen-bearing IgG IC into compartments from which the antigen can successfully be processed into peptide epitopes compatible with loading onto both major histocompatibility complex class I and II molecules. In turn, this enables the synchronous activation of both CD4+ and CD8+ T cell responses against the cognate antigen, thereby bridging the gap between the humoral and cellular branches of the adaptive immune response. Critically, FcRn-driven T cell priming is efficient at very low doses of antigen due to the exquisite sensitivity of the IgG-mediated antigen delivery system through which it operates. FcRn-mediated antigen presentation has important consequences in tissue compartments replete with IgG and serves not only to determine homeostatic immune activation at a variety of sites but also to induce inflammatory responses upon exposure to antigens perceived as foreign. Therapeutically targeting the pathway by which FcRn enables T cell activation in response to IgG IC is thus a highly attractive prospect not only for the treatment of diseases that are driven by immune complexes but also for manipulating local immune responses against defined antigens such as those present during infections and cancer. PMID:25221553

The Effect of Superparamagnetic Iron Oxide Nanoparticle Surface Charge on Antigen Cross-Presentation.

PubMed

Mou, Yongbin; Xing, Yun; Ren, Hongyan; Cui, Zhihua; Zhang, Yu; Yu, Guangjie; Urba, Walter J; Hu, Qingang; Hu, Hongming

2017-12-01

Magnetic nanoparticles (NPs) of superparamagnetic iron oxide (SPIO) have been explored for different kinds of applications in biomedicine, mechanics, and information. Here, we explored the synthetic SPIO NPs as an adjuvant on antigen cross-presentation ability by enhancing the intracellular delivery of antigens into antigen presenting cells (APCs). Particles with different chemical modifications and surface charges were used to study the mechanism of action of antigen delivery. Specifically, two types of magnetic NPs, γFe 2 O 3 /APTS (3-aminopropyltrimethoxysilane) NPs and γFe 2 O 3 /DMSA (meso-2, 3-Dimercaptosuccinic acid) NPs, with the same crystal structure, magnetic properties, and size distribution were prepared. Then, the promotion of T-cell activation via dendritic cells (DCs) was compared among different charged antigen coated NPs. Moreover, the activation of the autophagy, cytosolic delivery of the antigens, and antigen degradation mediated by the proteasome and lysosome were measured. Our results indicated that positive charged γFe 2 O 3 /APTS NPs, but not negative charged γFe 2 O 3 /DMSA NPs, enhanced the cross-presentation ability of DCs. Increased cross-presentation ability induced by γFe 2 O 3 /APTS NPs was associated with increased cytosolic antigen delivery. On the contrary, γFe 2 O 3 /DMSA NPs was associated with rapid autophagy. Overall, our results suggest that antigen delivered in cytoplasm induced by positive charged particles is beneficial for antigen cross-presentation and T-cell activation. NPs modified with different chemistries exhibit diverse biological properties and differ greatly in their adjuvant potentials. Thus, it should be carefully considered many different effects of NPs to design effective and safe adjuvants.

The Effect of Superparamagnetic Iron Oxide Nanoparticle Surface Charge on Antigen Cross-Presentation

NASA Astrophysics Data System (ADS)

Mou, Yongbin; Xing, Yun; Ren, Hongyan; Cui, Zhihua; Zhang, Yu; Yu, Guangjie; Urba, Walter J.; Hu, Qingang; Hu, Hongming

2017-01-01

Magnetic nanoparticles (NPs) of superparamagnetic iron oxide (SPIO) have been explored for different kinds of applications in biomedicine, mechanics, and information. Here, we explored the synthetic SPIO NPs as an adjuvant on antigen cross-presentation ability by enhancing the intracellular delivery of antigens into antigen presenting cells (APCs). Particles with different chemical modifications and surface charges were used to study the mechanism of action of antigen delivery. Specifically, two types of magnetic NPs, γFe2O3/APTS (3-aminopropyltrimethoxysilane) NPs and γFe2O3/DMSA (meso-2, 3-Dimercaptosuccinic acid) NPs, with the same crystal structure, magnetic properties, and size distribution were prepared. Then, the promotion of T-cell activation via dendritic cells (DCs) was compared among different charged antigen coated NPs. Moreover, the activation of the autophagy, cytosolic delivery of the antigens, and antigen degradation mediated by the proteasome and lysosome were measured. Our results indicated that positive charged γFe2O3/APTS NPs, but not negative charged γFe2O3/DMSA NPs, enhanced the cross-presentation ability of DCs. Increased cross-presentation ability induced by γFe2O3/APTS NPs was associated with increased cytosolic antigen delivery. On the contrary, γFe2O3/DMSA NPs was associated with rapid autophagy. Overall, our results suggest that antigen delivered in cytoplasm induced by positive charged particles is beneficial for antigen cross-presentation and T-cell activation. NPs modified with different chemistries exhibit diverse biological properties and differ greatly in their adjuvant potentials. Thus, it should be carefully considered many different effects of NPs to design effective and safe adjuvants.

Langerhans cells utilize CD1a and langerin to efficiently present nonpeptide antigens to T cells

PubMed Central

Hunger, Robert E.; Sieling, Peter A.; Ochoa, Maria Teresa; Sugaya, Makoto; Burdick, Anne E.; Rea, Thomas H.; Brennan, Patrick J.; Belisle, John T.; Blauvelt, Andrew; Porcelli, Steven A.; Modlin, Robert L.

2004-01-01

Langerhans cells (LCs) constitute a subset of DCs that initiate immune responses in skin. Using leprosy as a model, we investigated whether expression of CD1a and langerin, an LC-specific C-type lectin, imparts a specific functional role to LCs. LC-like DCs and freshly isolated epidermal LCs presented nonpeptide antigens of Mycobacterium leprae to T cell clones derived from a leprosy patient in a CD1a-restricted and langerin-dependent manner. LC-like DCs were more efficient at CD1a-restricted antigen presentation than monocyte-derived DCs. LCs in leprosy lesions coexpress CD1a and langerin, placing LCs in position to efficiently present a subset of antigens to T cells as part of the host response to human infectious disease. PMID:14991068

T cell costimulation by chemokine receptors.

PubMed

Molon, Barbara; Gri, Giorgia; Bettella, Monica; Gómez-Moutón, Concepción; Lanzavecchia, Antonio; Martínez-A, Carlos; Mañes, Santos; Viola, Antonella

2005-05-01

Signals mediated by chemokine receptors may compete with T cell receptor stop signals and determine the duration of T cell-antigen-presenting cell interactions. Here we show that during T cell stimulation by antigen-presenting cells, T cell chemokine receptors coupled to G(q) and/or G(11) protein were recruited to the immunological synapse by a G(i)-independent mechanism. When chemokine receptors were sequestered at the immunological synapse, T cells became insensitive to chemotactic gradients, formed more stable conjugates and finally responded with enhanced proliferation and cytokine production. We suggest that chemokine receptor trapping at the immunological synapse enhances T cell activation by improving T cell-antigen-presenting cell attraction and impeding the 'distraction' of successfully engaged T cells by other chemokine sources.

Purification of the Membrane Compartment for Endoplasmic Reticulum-associated Degradation of Exogenous Antigens in Cross-presentation.

PubMed

Imai, Jun; Otani, Mayu; Sakai, Takahiro; Hatta, Shinichi

2017-08-21

Dendritic cells (DCs) are highly capable of processing and presenting internalized exogenous antigens upon major histocompatibility class (MHC) I molecules also known as cross-presentation (CP). CP plays an important role not only in the stimulation of naïve CD8 + T cells and memory CD8 + T cells for infectious and tumor immunity but also in the inactivation of self-acting naïve T cells by T cell anergy or T cell deletion. Although the critical molecular mechanism of CP remains to be elucidated, accumulating evidence indicates that exogenous antigens are processed through endoplasmic reticulum-associated degradation (ERAD) after export from non-classical endocytic compartments. Until recently, characterizations of these endocytic compartments were limited because there were no specific molecular markers other than exogenous antigens. The method described here is a new vesicle isolation protocol, which allows for the purification of these endocytic compartments. Using this purified microsome, we reconstituted the ERAD-like transport, ubiquitination, and processing of the exogenous antigen in vitro, suggesting that the ubiquitin-proteasome system processed the exogenous antigen after export from this cellular compartment. This protocol can be further applied to other cell types to clarify the molecular mechanism of CP.

T-cells from HLA-B*57:01+ human subjects are activated with abacavir through two independent pathways and induce cell death by multiple mechanisms.

PubMed

Bell, Catherine C; Faulkner, Lee; Martinsson, Klara; Farrell, John; Alfirevic, Ana; Tugwood, Jonathan; Pirmohamed, Munir; Naisbitt, Dean J; Park, B Kevin

2013-05-20

Susceptibility to abacavir hypersensitivity has been attributed to possession of the specific human leukocyte antigen allele HLA-B*57:01. HLA-B*57:01-restricted activation of CD8+ T-cells provides a link between the genetic association and the iatrogenic disease. The objectives of this study were to characterize the functionality of drug-responsive CD8+ T-cell clones generated from HLA-B*57:01+ drug-naive subjects and to explore the relationship between abacavir accumulation in antigen presenting cells and the T-cell response. Seventy-four CD8+ clones expressing different Vβ receptors were shown to proliferate and kill target cells via different mechanisms when exposed to abacavir. Certain clones were activated with abacavir in the absence of antigen presenting cells. Analysis of the remaining clones revealed two pathways of drug-dependent T-cell activation. Overnight incubation of antigen presenting cells with abacavir, followed by repeated washing to remove soluble drug, activated approximately 50% of the clones, and the response was blocked by glutaraldehyde fixation. In contrast, a 1 h antigen presenting cell pulse did not activate any of the clones. Accumulation of abacavir in antigen presenting cells was rapid (less than 1 h), and the intracellular concentrations were maintained for 16 h. However, intracellular abacavir was not detectable by mass spectrometry after pulsing. These data suggest that T-cells can be activated by abacavir through a direct interaction with surface and intracellular major histocompatibility complex (MHC) molecules. With the former, abacavir seemingly participates in the MHC T-cell receptor binding interaction. In contrast, the latter pathway likely involves MHC binding peptides displayed as a consequence of abacavir exposure, but not abacavir itself.

The Epstein-Barr Virus Glycoprotein gp150 Forms an Immune-Evasive Glycan Shield at the Surface of Infected Cells

PubMed Central

Gram, Anna M.; Oosenbrug, Timo; Lindenbergh, Marthe F. S.; Büll, Christian; Comvalius, Anouskha; Dickson, Kathryn J. I.; Wiegant, Joop; Vrolijk, Hans; Lebbink, Robert Jan; Wolterbeek, Ron; Adema, Gosse J.; Griffioen, Marieke; Heemskerk, Mirjam H. M.; Tscharke, David C.; Hutt-Fletcher, Lindsey M.; Ressing, Maaike E.

2016-01-01

Cell-mediated immunity plays a key role in host control of viral infection. This is exemplified by life-threatening reactivations of e.g. herpesviruses in individuals with impaired T-cell and/or iNKT cell responses. To allow lifelong persistence and virus production in the face of primed immunity, herpesviruses exploit immune evasion strategies. These include a reduction in viral antigen expression during latency and a number of escape mechanisms that target antigen presentation pathways. Given the plethora of foreign antigens expressed in virus-producing cells, herpesviruses are conceivably most vulnerable to elimination by cell-mediated immunity during the replicative phase of infection. Here, we show that a prototypic herpesvirus, Epstein-Barr virus (EBV), encodes a novel, broadly acting immunoevasin, gp150, that is expressed during the late phase of viral replication. In particular, EBV gp150 inhibits antigen presentation by HLA class I, HLA class II, and the non-classical, lipid-presenting CD1d molecules. The mechanism of gp150-mediated T-cell escape does not depend on degradation of the antigen-presenting molecules nor does it require gp150’s cytoplasmic tail. Through its abundant glycosylation, gp150 creates a shield that impedes surface presentation of antigen. This is an unprecedented immune evasion mechanism for herpesviruses. In view of its likely broader target range, gp150 could additionally have an impact beyond escape of T cell activation. Importantly, B cells infected with a gp150-null mutant EBV displayed rescued levels of surface antigen presentation by HLA class I, HLA class II, and CD1d, supporting an important role for iNKT cells next to classical T cells in fighting EBV infection. At the same time, our results indicate that EBV gp150 prolongs the timespan for producing viral offspring at the most vulnerable stage of the viral life cycle. PMID:27077376

Antigen Cross-Priming of Cell-Associated Proteins is Enhanced by Macroautophagy within the Antigen Donor Cell

PubMed Central

Joubert, Pierre-Emmanuel; Albert, Matthew L.

2012-01-01

Phagocytosis of dying cells constitutes an important mechanism of antigen capture for the cross-priming of CD8+ T cells. This process has been shown to be critical for achieving tumor and viral immunity. While most studies have focused on the mechanisms inherent in the dendritic cell that account for exogenous antigen accessing MHC I, several recent reports have highlighted the important contribution made by the antigen donor cell. Specifically, the cell stress and cell death pathways that precede antigen transfer are now known to impact cross-presentation and cross-priming. Herein, we review the current literature regarding a role for macroautophagy within the antigen donor cell. Further examination of this point of immune regulation is warranted and may contribute to a better understanding of how to optimize immunotherapy for treatment of cancer and chronic infectious disease. PMID:22566942

Functional Macroautophagy Induction by Influenza A Virus without a Contribution to Major Histocompatibility Complex Class II-Restricted Presentation▿†

PubMed Central

Comber, Joseph D.; Robinson, Tara M.; Siciliano, Nicholas A.; Snook, Adam E.; Eisenlohr, Laurence C.

2011-01-01

Major histocompatibility complex (MHC) class II-presented peptides can be derived from both exogenous (extracellular) and endogenous (biosynthesized) sources of antigen. Although several endogenous antigen-processing pathways have been reported, little is known about their relative contributions to global CD4+ T cell responses against complex antigens. Using influenza virus for this purpose, we assessed the role of macroautophagy, a process in which cytosolic proteins are delivered to the lysosome by de novo vesicle formation and membrane fusion. Influenza infection triggered productive macroautophagy, and autophagy-dependent presentation was readily observed with model antigens that naturally traffic to the autophagosome. Furthermore, treatments that enhance or inhibit macroautophagy modulated the level of presentation from these model antigens. However, validated enzyme-linked immunospot (ELISpot) assays of influenza-specific CD4+ T cells from infected mice using a variety of antigen-presenting cells, including primary dendritic cells, revealed no detectable macroautophagy-dependent component. In contrast, the contribution of proteasome-dependent endogenous antigen processing to the global influenza CD4+ response was readily appreciated. The contribution of macroautophagy to the MHC class II-restricted response may vary depending upon the pathogen. PMID:21525345

Approach for Identifying Human Leukocyte Antigen (HLA)-DR Bound Peptides from Scarce Clinical Samples *

PubMed Central

Heyder, Tina; Kohler, Maxie; Tarasova, Nataliya K.; Haag, Sabrina; Rutishauser, Dorothea; Rivera, Natalia V.; Sandin, Charlotta; Mia, Sohel; Malmström, Vivianne; Wheelock, Åsa M.; Wahlström, Jan; Holmdahl, Rikard; Eklund, Anders; Zubarev, Roman A.; Grunewald, Johan; Ytterberg, A. Jimmy

2016-01-01

Immune-mediated diseases strongly associating with human leukocyte antigen (HLA) alleles are likely linked to specific antigens. These antigens are presented to T cells in the form of peptides bound to HLA molecules on antigen presenting cells, e.g. dendritic cells, macrophages or B cells. The identification of HLA-DR-bound peptides presents a valuable tool to investigate the human immunopeptidome. The lung is likely a key player in the activation of potentially auto-aggressive T cells prior to entering target tissues and inducing autoimmune disease. This makes the lung of exceptional interest and presents an ideal paradigm to study the human immunopeptidome and to identify antigenic peptides. Our previous investigation of HLA-DR peptide presentation in the lung required high numbers of cells (800 × 106 bronchoalveolar lavage (BAL) cells). Because BAL from healthy nonsmokers typically contains 10–15 × 106 cells, there is a need for a highly sensitive approach to study immunopeptides in the lungs of individual patients and controls. In this work, we analyzed the HLA-DR immunopeptidome in the lung by an optimized methodology to identify HLA-DR-bound peptides from low cell numbers. We used an Epstein-Barr Virus (EBV) immortalized B cell line and bronchoalveolar lavage (BAL) cells obtained from patients with sarcoidosis, an inflammatory T cell driven disease mainly occurring in the lung. Specifically, membrane complexes were isolated prior to immunoprecipitation, eluted peptides were identified by nanoLC-MS/MS and processed using the in-house developed ClusterMHCII software. With the optimized procedure we were able to identify peptides from 10 × 106 cells, which on average correspond to 10.9 peptides/million cells in EBV-B cells and 9.4 peptides/million cells in BAL cells. This work presents an optimized approach designed to identify HLA-DR-bound peptides from low numbers of cells, enabling the investigation of the BAL immunopeptidome from individual patients and healthy controls in order to identify disease-associated peptides. PMID:27452731

Neutrophils and monocytes transport tumor cell antigens from the peritoneal cavity to secondary lymphoid tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Terasawa, Masao; Nagata, Kisaburo; Kobayashi, Yoshiro

2008-12-12

Antigen-transporting cells take up pathogens, and then migrate from sites of inflammation to secondary lymphoid tissues to induce an immune response. Among antigen-transporting cells, dendritic cells (DCs) are believed to be the most potent and professional antigen-presenting cells that can stimulate naive T cells. However, the cells that transport antigens, tumor cell antigens in particular, have not been clearly identified. In this study we have analyzed what types of cells transport tumor cell antigens to secondary lymphoid tissues. We show that neutrophils, monocytes and macrophages but not DCs engulf X-irradiated P388 leukemic cells after their injection into the peritoneal cavity,more » and that neutrophils and monocytes but not macrophages migrate to the parathymic lymph nodes (pLN), the blood, and then the spleen. The monocytes in the pLN comprise Gr-1{sup -} and Gr-1{sup +} ones, and some of these cells express CD11c. Overall, this study demonstrates that neutrophils and monocytes transport tumor cell antigens from the peritoneal cavity to secondary lymphoid tissues.« less

Equine infectious anemia virus-infected dendritic cells retain antigen presentation capability

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rivera, Julie A.; McGuire, Travis C.

2005-05-10

To determine if equine monocyte-derived dendritic cells (DC) were susceptible to equine infectious anemia virus (EIAV) infection, ex vivo-generated DC were infected with virus in vitro. EIAV antigen was detected by immunofluorescence 3 days post-infection with maximum antigen being detected on day 4, whereas there was no antigen detected in DC incubated with the same amount of heat-inactivated EIAV. No cytolytic activity was observed after EIAV{sub WSU5} infection of DC. These monocyte-derived DC were more effective than macrophages and B cells in stimulating allogenic T lymphocytes. Both infected macrophages and DC stimulated similar levels of memory CTL responses in mixturesmore » of CD8+ and CD4+ cells as detected with {sup 51}Cr-release assays indicating that EIAV infection of DC did not alter antigen presentation. However, EIAV-infected DC were more effective than infected macrophages when used to stimulate memory CTL in isolated CD8+ cells. The maintenance of antigen processing and presenting function by EIAV-infected DC in vitro suggests that this function is maintained during in vivo infection.« less

Antigen-Specific Th17 Cells Are Primed by Distinct and Complementary Dendritic Cell Subsets in Oropharyngeal Candidiasis

PubMed Central

Kirchner, Florian R.; Becattini, Simone; Rülicke, Thomas; Sallusto, Federica; LeibundGut-Landmann, Salomé

2015-01-01

Candida spp. can cause severe and chronic mucocutaneous and systemic infections in immunocompromised individuals. Protection from mucocutaneous candidiasis depends on T helper cells, in particular those secreting IL-17. The events regulating T cell activation and differentiation toward effector fates in response to fungal invasion in different tissues are poorly understood. Here we generated a Candida-specific TCR transgenic mouse reactive to a novel endogenous antigen that is conserved in multiple distant species of Candida, including the clinically highly relevant C. albicans and C. glabrata. Using TCR transgenic T cells in combination with an experimental model of oropharyngeal candidiasis (OPC) we investigated antigen presentation and Th17 priming by different subsets of dendritic cells (DCs) present in the infected oral mucosa. Candida-derived endogenous antigen accesses the draining lymph nodes and is directly presented by migratory DCs. Tissue-resident Flt3L-dependent DCs and CCR2-dependent monocyte-derived DCs collaborate in antigen presentation and T cell priming during OPC. In contrast, Langerhans cells, which are also present in the oral mucosa and have been shown to prime Th17 cells in the skin, are not required for induction of the Candida-specific T cell response upon oral challenge. This highlights the functional compartmentalization of specific DC subsets in different tissues. These data provide important new insights to our understanding of tissue-specific antifungal immunity. PMID:26431538

Robust and Accurate Discrimination of Self/Non-Self Antigen Presentations by Regulatory T Cell Suppression.

PubMed

Furusawa, Chikara; Yamaguchi, Tomoyuki

The immune response by T cells usually discriminates self and non-self antigens, even though the negative selection of self-reactive T cells is imperfect and a certain fraction of T cells can respond to self-antigens. In this study, we construct a simple mathematical model of T cell populations to analyze how such self/non-self discrimination is possible. The results demonstrate that the control of the immune response by regulatory T cells enables a robust and accurate discrimination of self and non-self antigens, even when there is a significant overlap between the affinity distribution of T cells to self and non-self antigens. Here, the number of regulatory T cells in the system acts as a global variable controlling the T cell population dynamics. The present study provides a basis for the development of a quantitative theory for self and non-self discrimination in the immune system and a possible strategy for its experimental verification.

Robust and Accurate Discrimination of Self/Non-Self Antigen Presentations by Regulatory T Cell Suppression

PubMed Central

Furusawa, Chikara; Yamaguchi, Tomoyuki

2016-01-01

The immune response by T cells usually discriminates self and non-self antigens, even though the negative selection of self-reactive T cells is imperfect and a certain fraction of T cells can respond to self-antigens. In this study, we construct a simple mathematical model of T cell populations to analyze how such self/non-self discrimination is possible. The results demonstrate that the control of the immune response by regulatory T cells enables a robust and accurate discrimination of self and non-self antigens, even when there is a significant overlap between the affinity distribution of T cells to self and non-self antigens. Here, the number of regulatory T cells in the system acts as a global variable controlling the T cell population dynamics. The present study provides a basis for the development of a quantitative theory for self and non-self discrimination in the immune system and a possible strategy for its experimental verification. PMID:27668873

Anergic self-reactive B cells present self antigen and respond normally to CD40-dependent T-cell signals but are defective in antigen-receptor-mediated functions.

PubMed Central

Eris, J M; Basten, A; Brink, R; Doherty, K; Kehry, M R; Hodgkin, P D

1994-01-01

B-cell tolerance to soluble protein self antigens such as hen egg lysozyme (HEL) is mediated by clonal anergy. Anergic B cells fail to mount antibody responses even in the presence of carrier-primed T cells, suggesting an inability to activate or respond to T helper cells. To investigate the nature of this defect, B cells from tolerant HEL/anti-HEL double-transgenic mice were incubated with a membrane preparation from activated T-cell clones expressing the CD40 ligand. These membranes, together with interleukin 4 and 5 deliver the downstream antigen-independent CD40-dependent B-cell-activating signals required for productive T-B collaboration. Anergic B cells responded to this stimulus by proliferating and secreting antibody at levels comparable to or better than control B cells. Furthermore, anergic B cells presented HEL acquired in vivo and could present the unrelated antigen, conalbumin, targeted for processing via surface IgD. In contrast, the low immunoglobulin receptor levels on anergic B cells were associated with reduced de novo presentation of HEL and a failure to upregulate costimulatory ligands for CD28. These defects in immunoglobulin-receptor-mediated functions could be overcome in vivo, suggesting a number of mechanisms for induction of autoantibody responses. Images PMID:7514304

A pathway of costimulation that prevents anergy in CD28- T cells: B7- independent costimulation of CD1-restricted T cells

PubMed Central

1995-01-01

A class of molecules that is expressed on antigen presenting cells, exemplified by CD80 (B7), has been found to provide a necessary costimulatory signal for T cell activation and proliferation. CD28 and CTLA4 are the B7 counterreceptors and are expressed on the majority of human CD4+ T cells and many CD8+ T cells. The signal these molecules mediate is distinguished from other costimulatory signals by the finding that T cell recognition of antigen results in a prolonged state of T cell unresponsiveness or anergy, unless these costimulatory molecules are engaged. However, nearly half of the CD8+ and CD4-CD8- T cells lack CD28, and the costimulatory signals required for the activation of such cells are unknown. To understand the pathways of activation used by CD28- T cells, we have examined the costimulatory requirements of antigen-specific CD4-CD8- TCR(+)-alpha/beta circulating T cells that lack the expression of CD28. We have characterized two T cell lines, DN1 and DN6, that recognize a mycobacterial antigen, and are restricted not by major histocompatibility complex class I or II, but by CD1b or CD1c, two members of a family of major histocompatibility complex-related molecules that have been recently implicated in a distinct pathway for antigen presentation. Comparison of antigen-specific cytolytic responses of the DN1 and DN6 T cell lines against antigen-pulsed CD1+ monocytes or CD1+ B lymphoblastoid cell lines (B-LCL) demonstrated that these T cells recognized antigen presented by both types of cells. However, T cell proliferation occurred only when antigen was presented by CD1+ monocytes, indicating that the CD1+ monocytes expressed a costimulatory molecule that the B- LCL transfectants lacked. This hypothesis was confirmed by demonstrating that the T cells became anergic when incubated with the CD1(+)-transfected B-LCL in the presence of antigen, but not in the absence of antigen. The required costimulatory signal occurred by a CD28-independent mechanism since both the CD1+ monocytes and CD1+ B-LCL transfectants expressed B7-1 and B7-2, and DN1 and DN6 lacked surface expression of CD28. We propose that these data define a previously unrecognized pathway of costimulation for T cells distinct from that involving CD28 and its counterreceptors. We suggest that this B7- independent pathway plays a crucial role in the activation and maintenance of tolerance of at least a subset of CD28- T cells. PMID:7500046

Impact of aging on antigen presentation cell function of dendritic cells.

PubMed

Wong, Christine; Goldstein, Daniel R

2013-08-01

Older people exhibit increased mortality to infections and cancer as compared to younger people, indicating that aging impairs immunity. Dendritic cells (DCs) are key for bridging the innate and adaptive arms of the immune system by priming antigen specific T cells. Discerning how aging impacts DC function to initiate adaptive immune responses is of great biomedical importance as this could lead to the development of novel therapeutics to enhance immunity with aging. This review details reports indicating that aging impairs the antigen presenting function of DCs but highlights other studies indicating preserved DC function with aging. How aging impacts antigen presentation by DCs is complex and without a clear unifying biological underpinning. Copyright © 2013 Elsevier Ltd. All rights reserved.

Using Antigen-Specific B Cells to Combine Antibody and T Cell-Based Cancer Immunotherapy.

PubMed

Wennhold, Kerstin; Thelen, Martin; Schlößer, Hans Anton; Haustein, Natalie; Reuter, Sabrina; Garcia-Marquez, Maria; Lechner, Axel; Kobold, Sebastian; Rataj, Felicitas; Utermöhlen, Olaf; Chakupurakal, Geothy; Theurich, Sebastian; Hallek, Michael; Abken, Hinrich; Shimabukuro-Vornhagen, Alexander; von Bergwelt-Baildon, Michael

2017-09-01

Cancer immunotherapy by therapeutic activation of T cells has demonstrated clinical potential. Approaches include checkpoint inhibitors and chimeric antigen receptor T cells. Here, we report the development of an alternative strategy for cellular immunotherapy that combines induction of a tumor-directed T-cell response and antibody secretion without the need for genetic engineering. CD40 ligand stimulation of murine tumor antigen-specific B cells, isolated by antigen-biotin tetramers, resulted in the development of an antigen-presenting phenotype and the induction of a tumor antigen-specific T-cell response. Differentiation of antigen-specific B cells into antibody-secreting plasma cells was achieved by stimulation with IL21, IL4, anti-CD40, and the specific antigen. Combined treatment of tumor-bearing mice with antigen-specific CD40-activated B cells and antigen-specific plasma cells induced a therapeutic antitumor immune response resulting in remission of established tumors. Human CEA or NY-ESO-1-specific B cells were detected in tumor-draining lymph nodes and were able to induce antigen-specific T-cell responses in vitro, indicating that this approach could be translated into clinical applications. Our results describe a technique for the exploitation of B-cell effector functions and provide the rationale for their use in combinatorial cancer immunotherapy. Cancer Immunol Res; 5(9); 730-43. ©2017 AACR . ©2017 American Association for Cancer Research.

Multiplexed immunophenotyping of human antigen-presenting cells in whole blood by polychromatic flow cytometry

PubMed Central

Fung, Erik; Esposito, Laura; Todd, John A.; Wicker, Linda S.

2010-01-01

We describe two modular protocols for immunostaining and multiparameter flow cytometric analysis of major human antigen-presenting cells (dendritic cells, monocytes, B lymphocytes) in minimally manipulated whole blood. Simultaneous detection of up to eight colors is enabled by careful selection and testing of cell-subset-defining monoclonal antibodies (anchor markers) in the appropriate fluorochrome combinations, to demonstrate the quantification of surface expression levels of molecules involved in chemotaxis (e.g. CX3CR1, CCR2), adhesion (e.g. CD11b, CD62L), antigen presentation (e.g. CD83, CD86, CD209) and immune regulation (e.g. CD101) on circulating antigen-presenting cells. Each immunostaining reaction requires as little as 50–100 μl of peripheral whole blood, no density-gradient separation, and the entire procedure from preparation of reagents to flow cytometry can be completed in <5 h. PMID:20134434

Antigen presenting capacity of murine splenic myeloid cells.

PubMed

Hey, Ying-Ying; Quah, Benjamin; O'Neill, Helen C

2017-01-11

The spleen is an important site for hematopoiesis. It supports development of myeloid cells from bone marrow-derived precursors entering from blood. Myeloid subsets in spleen are not well characterised although dendritic cell (DC) subsets are clearly defined in terms of phenotype, development and functional role. Recently a novel dendritic-like cell type in spleen named 'L-DC' was distinguished from other known dendritic and myeloid cells by its distinct phenotype and developmental origin. That study also redefined splenic eosinophils as well as resident and inflammatory monocytes in spleen. L-DC are shown to be distinct from known splenic macrophages and monocyte subsets. Using a new flow cytometric procedure, it has been possible to identify and isolate L-DC in order to assess their functional competence and ability to activate T cells both in vivo and in vitro. L-DC are readily accessible to antigen given intravenously through receptor-mediated endocytosis. They are also capable of CD8 + T cell activation through antigen cross presentation, with subsequent induction of cytotoxic effector T cells. L-DC are MHCII - cells and unable to activate CD4 + T cells, a property which clearly distinguishes them from conventional DC. The myeloid subsets of resident monocytes, inflammatory monocytes, neutrophils and eosinophils, were found to have varying capacities to take up antigen, but were uniformly unable to activate either CD4 + T cells or CD8 + T cells. The results presented here demonstrate that L-DC in spleen are distinct from other myeloid cells in that they can process antigen for CD8 + T cell activation and induction of cytotoxic effector function, while both L-DC and myeloid subsets remain unable to activate CD4 + T cells. The L-DC subset in spleen is therefore distinct as an antigen presenting cell.

The overlooked "nonclassical" functions of major histocompatibility complex (MHC) class II antigens in immune and nonimmune cells.

PubMed

Altomonte, M; Pucillo, C; Maio, M

1999-06-01

Besides their "classical" antigenic peptide-presenting activity, major histocompatibility complex (MHC) class II antigens can activate different cellular functions in immune and nonimmune cells. However, this "nonclassical" role and its functional consequences are still substantially overlooked. In this review, we will focus on these alternative functional properties of MHC class II antigens, to reawaken attention to their present and foreseeable immunobiologic and pathogenetic implications. The main issues that will be addressed concern 1) the role of MHC class II molecules as basic components of exchangeable oligomeric protein complexes with intracellular signaling ability; 2) the nonclassical functions of MHC class II antigens in immune cells; 3) the pathogenetic role of MHC class II antigens in inflammatory/autoimmune and infectious disease; and 4) the functional role of MHC class II antigens in solid malignancies.

Cross–dressers turn on T cells

PubMed Central

YEWDELL, JONATHAN W.; DOLAN, BRIAN P.

2012-01-01

Memory T cells remember viruses from previous infections, providing immunity by facilitating the killing of infected cells. For this, they exploit cross-dressing, the transfer of antigens between antigen-presenting cells. PMID:21455165

Antigen discovery and delivery of subunit vaccines by nonliving bacterial ghost vectors.

PubMed

Walcher, Petra; Mayr, Ulrike B; Azimpour-Tabrizi, Chakameh; Eko, Francis O; Jechlinger, Wolfgang; Mayrhofer, Peter; Alefantis, Tim; Mujer, Cesar V; DelVecchio, Vito G; Lubitz, Werner

2004-12-01

The bacterial ghost (BG) platform system is a novel vaccine delivery system endowed with intrinsic adjuvant properties. BGs are nonliving Gram-negative bacterial cell envelopes which are devoid of their cytoplasmic contents, yet maintain their cellular morphology and antigenic structures, including bioadhesive properties. The main advantages of BGs as carriers of subunit vaccines include their ability to stimulate a high immune response and to target the carrier itself to primary antigen-presenting cells. The intrinsic adjuvant properties of BGs enhance the immune response to target antigens, including T-cell activation and mucosal immunity. Since native and foreign antigens can be carried in the envelope complex of BGs, combination vaccines with multiple antigens of diverse origin can be presented to the immune system simultaneously. Beside the capacity of BGs to function as carriers of protein antigens, they also have a high loading capacity for DNA. Thus, loading BGs with recombinant DNA takes advantage of the excellent bioavailability for DNA-based vaccines and the high expression rates of the DNA-encoded antigens in target cell types such as macrophages and dendritic cells. There are many spaces within BGs including the inner and outer membranes, the periplasmic space and the internal lumen which can carry antigens, DNA or mediators of the immune response. All can be used for subunit antigen to design new vaccine candidates with particle presentation technology. In addition, the fact that BGs can also carry piggyback large-size foreign antigen particles, increases the technologic usefulness of BGs as combination vaccines against viral and bacterial pathogens. Furthermore, the BG antigen carriers can be stored as freeze-dried preparations at room temperature for extended periods without loss of efficacy. The potency, safety and relatively low production cost of BGs offer a significant technical advantage over currently utilized vaccine technologies.

Adaptor protein-3 is required in dendritic cells for optimal Toll-like receptor signaling from phagosomes and antigen presentation to CD4+ T cells

PubMed Central

Mantegazza, Adriana R.; Guttentag, Susan H.; El-Benna, Jamel; Sasai, Miwa; Iwasaki, Akiko; Shen, Hao; Laufer, Terri M.; Marks, Michael S.

2012-01-01

SUMMARY Effective major histocompatibility complex-II (MHC-II) antigen presentation from phagocytosed particles requires phagosome-intrinsic toll-like receptor (TLR) signaling, but the molecular mechanisms underlying TLR delivery to phagosomes and how signaling regulates antigen presentation are incompletely understood. We show a requirement in dendritic cells (DCs) for adaptor protein-3 (AP-3) in efficient TLR recruitment to phagosomes and MHC-II presentation of antigens internalized by phagocytosis but not receptor-mediated endocytosis. DCs from AP-3-deficient pearl mice elicited impaired CD4+ T cell activation and Th1 effector function to particulate antigen in vitro and to recombinant Listeria monocytogenes infection in vivo. Whereas phagolysosome maturation and peptide:MHC-II complex assembly proceeded normally in pearl DCs, peptide:MHC-II export to the cell surface was impeded. This correlated with reduced TLR4 recruitment and proinflammatory signaling from phagosomes by particulate TLR ligands. We propose that AP-3-dependent TLR delivery from endosomes to phagosomes and subsequent signaling mobilize peptide:MHC-II export from intracellular stores. PMID:22560444

Understanding MHC Class I Presentation of Viral Antigens by Human Dendritic Cells as a Basis for Rational Design of Therapeutic Vaccines

PubMed Central

van Montfoort, Nadine; van der Aa, Evelyn; Woltman, Andrea M.

2014-01-01

Effective viral clearance requires the induction of virus-specific CD8+ cytotoxic T lymphocytes (CTL). Since dendritic cells (DC) have a central role in initiating and shaping virus-specific CTL responses, it is important to understand how DC initiate virus-specific CTL responses. Some viruses can directly infect DC, which theoretically allow direct presentation of viral antigens to CTL, but many viruses target other cells than DC and thus the host depends on the cross-presentation of viral antigens by DC to activate virus-specific CTL. Research in mouse models has highly enhanced our understanding of the mechanisms underlying cross-presentation and the dendritic cells (DC) subsets involved, however, these results cannot be readily translated toward the role of human DC in MHC class I-antigen presentation of human viruses. Here, we summarize the insights gained in the past 20 years on MHC class I presentation of viral antigen by human DC and add to the current debate on the capacities of different human DC subsets herein. Furthermore, possible sources of viral antigens and essential DC characteristics for effective induction of virus-specific CTL are evaluated. We conclude that cross-presentation is not only an efficient mechanism exploited by DC to initiate immunity to viruses that do not infect DC but also to viruses that do infect DC, because cross-presentation has many conceptual advantages and bypasses direct immune modulatory effects of the virus on its infected target cells. Since knowledge on the mechanism of viral antigen presentation and the preferred DC subsets is crucial for rational vaccine design, the obtained insights are very instrumental for the development of effective anti-viral immunotherapy. PMID:24795724

MHC structure and function − antigen presentation. Part 2

PubMed Central

Goldberg, Anna Carla; Rizzo, Luiz Vicente

2015-01-01

The second part of this review deals with the molecules and processes involved in the processing and presentation of the antigenic fragments to the T-cell receptor. Though the nature of the antigens presented varies, the most significant class of antigens is proteins, processed within the cell to be then recognized in the form of peptides, a mechanism that confers an extraordinary degree of precision to this mode of immune response. The efficiency and accuracy of this system is also the result of the myriad of mechanisms involved in the processing of proteins and production of peptides, in addition to the capture and recycling of alternative sources aiming to generate further diversity in the presentation to T-cells. PMID:25807243

Multivalent glycopeptide dendrimers for the targeted delivery of antigens to dendritic cells.

PubMed

García-Vallejo, Juan J; Ambrosini, Martino; Overbeek, Annemieke; van Riel, Wilhelmina E; Bloem, Karien; Unger, Wendy W J; Chiodo, Fabrizio; Bolscher, Jan G; Nazmi, Kamran; Kalay, Hakan; van Kooyk, Yvette

2013-04-01

Dendritic cells are the most powerful type of antigen presenting cells. Current immunotherapies targeting dendritic cells have shown a relative degree of success but still require further improvement. One of the most important issues to solve is the efficiency of antigen delivery to dendritic cells in order to achieve an appropriate uptake, processing, and presentation to Ag-specific T cells. C-type lectins have shown to be ideal receptors for the targeting of antigens to dendritic cells and allow the use of their natural ligands - glycans - instead of antibodies. Amongst them, dendritic cell-specific ICAM-3-grabbing non-integrin (DC-SIGN) is an interesting candidate due to its biological properties and the availability of its natural carbohydrate ligands. Using Le(b)-conjugated poly(amido amine) (PAMAM) dendrimers we aimed to characterize the optimal level of multivalency necessary to achieve the desired internalization, lysosomal delivery, Ag-specific T cell proliferation, and cytokine response. Increasing DC-SIGN ligand multivalency directly translated in an enhanced binding, which might also be interesting for blocking purposes. Internalization, routing to lysosomal compartments, antigen presentation and cytokine response could be optimally achieved with glycopeptide dendrimers carrying 16-32 glycan units. This report provides the basis for the design of efficient targeting of peptide antigens for the immunotherapy of cancer, autoimmunity and infectious diseases. Copyright © 2012 Elsevier Ltd. All rights reserved.

Polymer blend particles with defined compositions for targeting antigen to both class I and II antigen presentation pathways

PubMed Central

Tran, Kenny K.; Zhan, Xi; Shen, Hong

2013-01-01

Defense against many persistent and difficult-to-treat diseases requires a combination of humoral, CD4+ and CD8+ T cell responses, which necessitates targeting antigens to both class I and II antigen presentation pathways. In this study, we developed polymer blend particles by mixing two functionally unique polymers, poly(lactide-co-glycolide) (PLGA) and a pH-responsive polymer, poly(dimethylaminoethyl methacrylate-co-propylacrylic acid-co-butyl methacrylate) (DMAEMA-co-PAA-co-BMA). We showed polymer blend particles enabled the delivery of antigens into both class I and II antigen presentation pathways in vitro. Increasing the ratio of the pH-responsive polymer in blend particles increased the degree of class I antigen presentation, while maintaining high levels of class II antigen presentation. In a mouse model, we demonstrated that a significantly higher and sustained level of CD4+ and CD8+ T cell responses, and comparable antibody responses, were elicited with polymer blend particles than PLGA particles and a conventional vaccine, Alum. The polymer blend particles offer a potential vaccine delivery platform to generate a combination of humoral and cell-mediated immune responses that insure robust and long-lasting immunity against many infectious diseases and cancers. PMID:24124123

Peyer’s patches: Organizing B cell responses at the intestinal frontier

PubMed Central

Reboldi, Andrea; Cyster, Jason G

2015-01-01

Summary Secondary lymphoid tissues share the important function of bringing together antigens and rare antigen-specific lymphocytes to foster induction of adaptive immune responses. Peyer’s patches (PPs) are unique compared to other secondary lymphoid tissues in their continual exposure to an enormous diversity of microbiome- and food-derived antigens and in the types of pathogens they encounter. Antigens are delivered to PPs by specialized microfold (M) epithelial cells and they may be captured and presented by resident dendritic cells (DCs). In accord with their state of chronic microbial antigen exposure, PPs exhibit continual germinal center (GC) activity. These GCs contribute to the generation of B cells and plasma cells producing somatically mutated gut antigen-specific IgA antibodies, but have also been suggested to support antigen-nonspecific diversification of the B cell repertoire. Here we review current understanding of how PPs foster B cell encounters with antigen, how they favor isotype switching to the secretory IgA isotype, and how their GC responses may uniquely contribute to mucosal immunity. PMID:27088918

Poly-functional and long-lasting anticancer immune response elicited by a safe attenuated Pseudomonas aeruginosa vector for antigens delivery

PubMed Central

Chauchet, Xavier; Hannani, Dalil; Djebali, Sophia; Laurin, David; Polack, Benoit; Marvel, Jacqueline; Buffat, Laurent; Toussaint, Bertrand; Le Gouëllec, Audrey

2016-01-01

Live-attenuated bacterial vectors for antigens delivery have aroused growing interest in the field of cancer immunotherapy. Their potency to stimulate innate immunity and to promote intracellular antigen delivery into antigen-presenting cells could be exploited to elicit a strong and specific cellular immune response against tumor cells. We previously described genetically-modified and attenuated Pseudomonas aeruginosa vectors able to deliver in vivo protein antigens into antigen-presenting cells, through Type 3 secretion system of the bacteria. Using this approach, we managed to protect immunized mice against aggressive B16 melanoma development in both a prophylactic and therapeutic setting. In this study, we further investigated the antigen-specific CD8+ T cell response, in terms of phenotypic and functional aspects, obtained after immunizations with a killed but metabolically active P. aeruginosa attenuated vector. We demonstrated that P. aeruginosa vaccine induces a highly functional pool of antigen-specific CD8+ T cell able to infiltrate the tumor. Furthermore, multiple immunizations allowed the development of a long-lasting immune response, represented by a pool of predominantly effector memory cells which protected mice against late tumor challenge. Overall, killed but metabolically active P. aeruginosa vector is a safe and promising approach for active and specific antitumor immunotherapy. PMID:28035332

The tyrosine phosphatase PTPN22 discriminates weak self peptides from strong agonist TCR signals.

PubMed

Salmond, Robert J; Brownlie, Rebecca J; Morrison, Vicky L; Zamoyska, Rose

2014-09-01

T cells must be tolerant of self antigens to avoid autoimmunity but responsive to foreign antigens to provide protection against infection. We found that in both naive T cells and effector T cells, the tyrosine phosphatase PTPN22 limited signaling via the T cell antigen receptor (TCR) by weak agonists and self antigens while not impeding responses to strong agonist antigens. T cells lacking PTPN22 showed enhanced formation of conjugates with antigen-presenting cells pulsed with weak peptides, which led to activation of the T cells and their production of inflammatory cytokines. This effect was exacerbated under conditions of lymphopenia, with the formation of potent memory T cells in the absence of PTPN22. Our data address how loss-of-function PTPN22 alleles can lead to the population expansion of effector and/or memory T cells and a predisposition to human autoimmunity.

The CD1 family: serving lipid antigens to T cells since the Mesozoic era.

PubMed

Zajonc, Dirk M

2016-08-01

Class I-like CD1 molecules are in a family of antigen-presenting molecules that bind lipids and lipopeptides, rather than peptides for immune surveillance by T cells. Since CD1 lacks the high degree of polymorphism found in their major histocompatibility complex (MHC) class I molecules, different species express different numbers of CD1 isotypes, likely to be able to present structurally diverse classes of lipid antigens. In this review, we will present a historical overview of the structures of the different human CD1 isotypes and also discuss species-specific adaptations of the lipid-binding groove. We will discuss how single amino acid changes alter the shape and volume of the CD1 binding groove, how these minor changes can give rise to different numbers of binding pockets, and how these pockets affect the lipid repertoire that can be presented by any given CD1 protein. We will compare the structures of various lipid antigens and finally, we will discuss recognition of CD1-presented lipid antigens by antigen receptors on T cells (TCRs).

The CD1 family: serving lipid antigens to T cells since the Mesozoic era

PubMed Central

Zajonc, Dirk M.

2016-01-01

Class I-like CD1 molecules are in a family of antigen-presenting molecules that bind lipids and lipopeptides, rather than peptides for immune surveillance by T cells. Since CD1 lacks the high degree of polymorphism found in their major histocompatibility complex (MHC) class I molecules, different species express different numbers of CD1 isotypes, likely to be able to present structurally diverse classes of lipid antigens. In this review, we will present a historical overview of the structures of the different human CD1 isotypes and also discuss species-specific adaptations of the lipid-binding groove. We will discuss how single amino acid changes alter the shape and volume of the CD1 binding groove, how these minor changes can give rise to different numbers of binding pockets, and how these pockets affect the lipid repertoire that can be presented by any given CD1 protein. We will compare the structures of various lipid antigens and finally, we will discuss recognition of CD1-presented lipid antigens by antigen receptors on T cells (TCRs). PMID:27368414

Identification and characterization of B cell precursors in rat lymphoid tissues. I. Adoptive transfer assays for precursors of TI-1, TI-2, and TD antigen-reactive B cells.

PubMed

Whalen, B J; Goldschneider, I

1993-10-01

Quantitative adoptive transfer assays were developed to detect the precursors of TI-1, TI-2, and TD antigen-reactive B cells in rat lymphoid tissues. Studies on the immune responses in normal and athymic nude rats validate the use of TNP-lipopolysaccharide as a TI-1 antigen, TNP-Ficoll as a TI-2 antigen, and SRBC as a TD antigen in rats. The precursors to these immunologically competent B cells are detected, following transfer into irradiated histocompatible recipients, by their ability to generate expanded populations of antigen-reactive B cells capable of mounting antibody responses (splenic IgM plaque-forming cells) to these antigens. Maximal numbers of antigen-reactive B cells emerge in antigenically naive rats after an interval of 7-12 days following transfer of donor lymphoid cells and decline rapidly thereafter. The delayed responses in adoptive recipients reconstituted with spleen cells are proportional to the numbers of spleen cells transferred and are shown to be primarily donor derived using histocompatible Ig kappa chain alloantigen disparate rat strain combinations. The precursors of TI-1, TI-2, and TD antigen-reactive B cells are present in both donor spleen and bone marrow. However, precursor cells to TI-1 and TD antigens are largely absent from donor lymph node cells, whereas precursors to the TI-2 antigen are as prevalent in donor lymph node as in donor spleen. These results support the hypothesis that newly formed virginal B cells represent transient populations of precursor cells that undergo further proliferation and differentiation in the spleen before acquiring immunological competence. The results also suggest that the precursors of TI-2 antigen-reactive B cells differ developmentally from those of TI-1 and TD antigen-reactive B cells, and that the antigen-reactive progeny of these precursors require additional stimulation in order to join the pool of long-lived peripheral B cells.

Protective antitumor activity through dendritic cell immunization is mediated by NK cell as well as CTL activation.

PubMed

Kim, K D; Kim, J K; Kim, S J; Choe, I S; Chung, T H; Choe, Y K; Lim, J S

1999-08-01

Dendritic cells (DCs) are potent professional antigen-presenting cells (APC) capable of inducing the primary T cell response to antigen. Although tumor cells express target antigens, they are incapable of stimulating a tumor-specific immune response due to a defect in the costimulatory signal that is required for optimal activation of T cells. In this work, we describe a new approach using tumor-DC coculture to improve the antigen presenting capacity of tumor cells, which does not require a source of tumor-associated antigen. Immunization of a weakly immunogenic and progressive tumor cocultured with bone marrow-derived DCs generated an effective tumor vaccine. Immunization with the cocultured DCs was able to induce complete protective immunity against tumor challenges and was effective for the induction of tumor-specific CTL (cytotoxic T lymphocyte) activity. Furthermore, high NK cell activity was observed in mice in which tumors were rejected. In addition, immunization with tumor-pulsed DCs induced delayed tumor growth, but not tumor eradication in tumor-bearing mice. Our results demonstrate that coculture of DCs with tumors generated antitumor immunity due to the NK cell activation as well as tumor-specific T cell. This approach would be useful for designing tumor vaccines using DCs when the information about tumor antigens is limited.

Identification of the soluble HVP-associated antigen of the lymphoblastoid cell line established from lymphomatous baboon (Papio hamadryas).

PubMed

Voevodin, A F; Lapin, B A; Agrba, V Z; Timanovskaya, V V

1978-01-01

A new technique (indirect double immunodiffusion) for detection of EBV-associated soluble antigen and corresponding antibodies has been developed. This technique includes three steps: 1) simple double immunodiffusion with extracts of Raji cells (or other EBV-genome positive cells) and human sera containing antibodies against EBV-associated soluble antigen; 2) extensive washing and treatment with anti-human globulin; 3) extensive washing and treatment with tannic acid. Using this test it was shown that the soluble antigen indistinguishable from EBV-associated soluble antigen was present in KMPG-1 cells producing HVP.

Cross-Priming of Naive Cd8 T Cells against Melanoma Antigens Using Dendritic Cells Loaded with Killed Allogeneic Melanoma Cells

PubMed Central

Berard, Frederic; Blanco, Patrick; Davoust, Jean; Neidhart-Berard, Eve-Marie; Nouri-Shirazi, Mahyar; Taquet, Nicolas; Rimoldi, Donata; Cerottini, Jean Charles; Banchereau, Jacques; Palucka, A. Karolina

2000-01-01

The goal of tumor immunotherapy is to elicit immune responses against autologous tumors. It would be highly desirable that such responses include multiple T cell clones against multiple tumor antigens. This could be obtained using the antigen presenting capacity of dendritic cells (DCs) and cross-priming. That is, one could load the DC with tumor lines of any human histocompatibility leukocyte antigen (HLA) type to elicit T cell responses against the autologous tumor. In this study, we show that human DCs derived from monocytes and loaded with killed melanoma cells prime naive CD45RA+CD27+CD8+ T cells against the four shared melanoma antigens: MAGE-3, gp100, tyrosinase, and MART-1. HLA-A201+ naive T cells primed by DCs loaded with HLA-A201− melanoma cells are able to kill several HLA-A201+ melanoma targets. Cytotoxic T lymphocyte priming towards melanoma antigens is also obtained with cells from metastatic melanoma patients. This demonstration of cross-priming against shared tumor antigens builds the basis for using allogeneic tumor cell lines to deliver tumor antigens to DCs for vaccination protocols. PMID:11104796

Antigen-Specific CD8+ T Cells Fail To Respond to Shigella flexneri ▿

PubMed Central

Jehl, Stephanie P.; Doling, Amy M.; Giddings, Kara S.; Phalipon, Armelle; Sansonetti, Philippe J.; Goldberg, Marcia B.; Starnbach, Michael N.

2011-01-01

CD8+ T lymphocytes often play a primary role in adaptive immunity to cytosolic microbial pathogens. Surprisingly, CD8+ T cells are not required for protective immunity to the enteric pathogen Shigella flexneri, despite the ability of Shigella to actively secrete proteins into the host cytoplasm, a location from which antigenic peptides are processed for presentation to CD8+ T cells. To determine why CD8+ T cells fail to play a role in adaptive immunity to S. flexneri, we investigated whether antigen-specific CD8+ T cells are primed during infection but are unable to confer protection or, alternatively, whether T cells fail to be primed. To test whether Shigella is capable of stimulating an antigen-specific CD8+ T-cell response, we created an S. flexneri strain that constitutively secretes a viral CD8+ T-cell epitope via the Shigella type III secretion system and characterized the CD8+ T-cell response to this strain both in mice and in cultured cells. Surprisingly, no T cells specific for the viral epitope were stimulated in mice infected with this strain, and cells infected with the recombinant strain were not targeted by epitope-specific T cells. Additionally, we found that the usually robust T-cell response to antigens artificially introduced into the cytoplasm of cultured cells was significantly reduced when the antigen-presenting cell was infected with Shigella. Collectively, these results suggest that antigen-specific CD8+ T cells are not primed during S. flexneri infection and, as a result, afford little protection to the host during primary or subsequent infection. PMID:21357720

'Educated' dendritic cells act as messengers from memory to naive T helper cells.

PubMed

Alpan, Oral; Bachelder, Eric; Isil, Eda; Arnheiter, Heinz; Matzinger, Polly

2004-06-01

Ingested antigens lead to the generation of effector T cells that secrete interleukin 4 (IL-4) rather than interferon-gamma (IFN-gamma) and are capable of influencing naive T cells in their immediate environment to do the same. Using chimeric mice generated by aggregation of two genotypically different embryos, we found that the conversion of a naive T cell occurs only if it can interact with the same antigen-presenting cell, although not necessarily the same antigen, as the effector T cell. Using a two-step culture system in vitro, we found that antigen-presenting dendritic cells can act as 'temporal bridges' to relay information from orally immunized memory CD4 T cells to naive CD4 T cells. The orally immunized T cells use IL-4 and IL-10 (but not CD40 ligand) to 'educate' dendritic cells, which in turn induce naive T cells to produce the same cytokines as those produced by the orally immunized memory T cells.

Therapeutic use of Aldara in chronic myeloid leukemia.

PubMed

Marleau, Annette M; Lipton, Jeffrey H; Riordan, Neil H; Ichim, Thomas E

2007-01-25

The potent clinical responses seen in patients with chronic myeloid leukemia (CML) after administration of donor-specific lymphocytes, as well as the correlation between the presence of antigen specific T cells and prolonged remission in these patients, suggests a role for the immunological control of CML. Here we propose Aldara, a clinically used formulation of imiquimod, as an agent for augmenting immune responses to CML antigens. Our proposition is based upon 3 tenets: 1) Endogenous dendritic cells (DC) of CML patients, which are known to be derived from the malignant clone, express and present various leukemic antigens; 2) CML-antigen reactive T cell clones exist in the patient but in many situations are ineffectively stimulated to cause significant hematological responses; and 3) Antigen presentation by mature, activated DC, which endogenously express CML-antigens may endow the pre-existing ineffective T cell responses with ability to control CML progression. The practical use of Aldara as a localized activator of DC in the context of present day leukemic therapeutics, as well as various properties of this unique immune modulator will be discussed.

[Dendritic cell-based therapeutic cancer vaccines].

PubMed

Rizzo, Manglio; Alaniz, Laura; Mazzolini, Guillermo D

In recent years immunotherapy has revolutionized the treatment of patients with advanced cancer. The increased knowledge in the tumor immune-biology has allowed developing rational treatments by manipulation of the immune system with significant clinical impact. This rapid development has significantly changed the prognosis of many tumors without treatment options up to date. Other strategies have explored the use of therapeutic vaccines based on dendritic cells (DC) by inducing antitumor immunity. DC are cells of hematopoietic origin, constitutively expressing molecules capable to present antigens, that are functionally the most potent inducers of the activation and proliferation of antigen specific T lymphocytes. The CD8+ T cells proliferate and acquire cytotoxic capacity after recognizing their specific antigen presented on the surface of DC, although only some types of DC can present antigens internalized from outside the cell to precursors of cytotoxic T lymphocytes (this function is called cross-presentation) requiring translocation mechanisms of complex antigens. The induction of an effective adaptive immune response is considered a good option given its specificity, and prolonged duration of response. The DC, thanks to its particular ability of antigen presentation and lymphocyte stimulation, are able to reverse the poor antitumor immune response experienced by patients with cancer. The DC can be obtained from various sources, using different protocols to generate differentiation and maturation, and are administered by various routes such as subcutaneous, intravenous or intranodal. The wide variety of protocols resulted in heterogeneous clinical responses.

A critical examination of the numerology of antigen-binding cells: evidence for multiple receptor specificities on single cells.

PubMed

Miller, A

1977-01-01

The data available from other laboratories as well as our own on the frequency of cells recognizing major histocompatibility antigens or conventional protein and hapten antigens is critically evaluated. The frequency of specific binding for a large number of antigens is sufficiently high to support the idea that at least part of the antigen-binding cell population must have multiple specificities. Our results suggest that these multiple specific cells result from single cells synthesizing and displaying as many as 50-100 species of receptor, each at a frequency of 10(4) per cell. A model involving gene expansion of constant-region genes is suggested and some auxilliary evidence consistent with such C-gene expansion is presented.

Glycan modification of antigen alters its intracellular routing in dendritic cells, promoting priming of T cells

PubMed Central

Streng-Ouwehand, Ingeborg; Ho, Nataschja I; Litjens, Manja; Kalay, Hakan; Boks, Martine Annemarie; Cornelissen, Lenneke AM; Kaur Singh, Satwinder; Saeland, Eirikur; Garcia-Vallejo, Juan J; Ossendorp, Ferry A; Unger, Wendy WJ; van Kooyk, Yvette

2016-01-01

Antigen uptake by dendritic cells and intracellular routing of antigens to specific compartments is regulated by C-type lectin receptors that recognize glycan structures. We show that the modification of Ovalbumin (OVA) with the glycan-structure LewisX (LeX) re-directs OVA to the C-type lectin receptor MGL1. LeX-modification of OVA favored Th1 skewing of CD4+ T cells and enhanced cross-priming of CD8+ T cells. While cross-presentation of native OVA requires high antigen dose and TLR stimuli, LeX modification reduces the required amount 100-fold and obviates its dependence on TLR signaling. The OVA-LeX-induced enhancement of T cell cross-priming is MGL1-dependent as shown by reduced CD8+ effector T cell frequencies in MGL1-deficient mice. Moreover, MGL1-mediated cross-presentation of OVA-LeX neither required TAP-transporters nor Cathepsin-S and was still observed after prolonged intracellular storage of antigen in Rab11+LAMP1+ compartments. We conclude that controlled neo-glycosylation of antigens can crucially influence intracellular routing of antigens, the nature and strength of immune responses and should be considered for optimizing current vaccination strategies. DOI: http://dx.doi.org/10.7554/eLife.11765.001 PMID:26999763

Comprehensive Analysis of the Activation and Proliferation Kinetics and Effector Functions of Human Lymphocytes, and Antigen Presentation Capacity of Antigen-Presenting Cells in Xenogeneic Graft-Versus-Host Disease.

PubMed

Kawasaki, Yasufumi; Sato, Kazuya; Hayakawa, Hiroko; Takayama, Norihito; Nakano, Hirofumi; Ito, Ryoji; Mashima, Kiyomi; Oh, Iekuni; Minakata, Daisuke; Yamasaki, Ryoko; Morita, Kaoru; Ashizawa, Masahiro; Yamamoto, Chihiro; Hatano, Kaoru; Fujiwara, Shin-Ichiro; Ohmine, Ken; Muroi, Kazuo; Kanda, Yoshinobu

2018-04-17

Xenogeneic graft-versus-host disease (GVHD) models in highly immunodeficient mice are currently being used worldwide to investigate human immune responses against foreign antigens in vivo. However, the individual roles of CD4 + and CD8 + T cells, and donor/host hematopoietic and nonhematopoietic antigen-presenting cells (APCs) in the induction and development of GVHD have not been fully investigated. In the present study, we comprehensively investigated the immune responses of human T cells and the antigen presentation capacity of donor/host hematopoietic and nonhematopoietic APCs in xenogeneic GVHD models using nonobese diabetic/Shi-scid-IL2rg null mice. CD4 + T cells and, to a lesser extent, CD8 + T cells individually mediated potentially lethal GVHD. In addition to inflammatory cytokine production, CD4 + T cells also supported the activation and proliferation of CD8 + T cells. Using bone marrow chimeras, we demonstrated that host hematopoietic, but not nonhematopoietic, APCs play a critical role in the development of CD4 + T cell-mediated GVHD. During early GVHD, we detected 2 distinct populations in memory CD4 + T cells. One population was highly activated and proliferated in major histocompatibility complex antigen (MHC) +/+ mice but not in MHC -/- mice, indicating alloreactive T cells. The other population showed a less activated and slowly proliferative status regardless of host MHC expression, and was associated with higher susceptibility to apoptosis, indicating nonalloreactive T cells in homeostasis-driven proliferation. These observations are clinically relevant to donor T cell response after allogeneic hematopoietic stem cell transplantation. Our findings provide a better understanding of the immunobiology of humanized mice and support the development of novel options for the prevention and treatment for GVHD. Copyright © 2018. Published by Elsevier Inc.

Uptake routes of tumor-antigen MAGE-A3 by dendritic cells determine priming of naïve T-cell subtypes.

PubMed

Moeller, Ines; Spagnoli, Giulio C; Finke, Jürgen; Veelken, Hendrik; Houet, Leonora

2012-11-01

Induction of tumor-antigen-specific T cells in active cancer immunotherapy is generally difficult due to the very low anti-tumoral precursor cytotoxic T cells. By improving tumor-antigen uptake and presentation by dendritic cells (DCs), this problem can be overcome. Focusing on MAGE-A3 protein, frequently expressed in many types of tumors, we analyzed different DC-uptake routes after additional coating the recombinant MAGE-A3 protein with either a specific monoclonal antibody or an immune complex formulation. Opsonization of the protein with antibody resulted in increased DC-uptake compared to the uncoated rhMAGE-A3 protein. This was partly due to Fcγ receptor-dependent internalization. However, unspecific antigen internalization via macropinocytosis also played a role. When analyzing DC-uptake of MAGE-A3 antigen expressed in multiple myeloma cell line U266, pretreatment with proteasome inhibitor bortezomib resulted in increased apoptosis compared to γ-irradiation. Bortezomib-mediated immunogenic apoptosis, characterized by elevated surface expression of hsp90, triggered higher phagocytosis of U266 cells by DCs involving specific DC-derived receptors. We further investigated the impact of antigen delivery on T-cell priming. Induction of CD8(+) T-cell response was favored by stimulating naïve T cells with either antibody-opsonized MAGE-A3 protein or with the bortezomib-pretreated U266 cells, indicating that receptor-mediated uptake favors cross-presentation of antigens. In contrast, CD4(+) T cells were preferentially induced after stimulation with the uncoated protein or protein in the immune complex, both antigen formulations were preferentially internalized by DCs via macropinocytosis. In summary, receptor-mediated DC-uptake mechanisms favored the induction of CD8(+) T cells, relevant for clinical anti-tumor response.

Targeting LAG-3 and PD-1 to Enhance T Cell Activation by Antigen-Presenting Cells

PubMed Central

Lichtenegger, Felix S.; Rothe, Maurine; Schnorfeil, Frauke M.; Deiser, Katrin; Krupka, Christina; Augsberger, Christian; Schlüter, Miriam; Neitz, Julia; Subklewe, Marion

2018-01-01

Immune checkpoint inhibition has been shown to successfully reactivate endogenous T cell responses directed against tumor-associated antigens, resulting in significantly prolonged overall survival in patients with various tumor entities. For malignancies with low endogenous immune responses, this approach has not shown a clear clinical benefit so far. Therapeutic vaccination, particularly dendritic cell (DC) vaccination, is a strategy to induce T cell responses. Interaction of DCs and T cells is dependent on receptor–ligand interactions of various immune checkpoints. In this study, we analyzed the influence of blocking antibodies targeting programmed cell death protein 1 (PD-1), HVEM, CD244, TIM-3, and lymphocyte activation gene 3 (LAG-3) on the proliferation and cytokine secretion of T cells after stimulation with autologous TLR-matured DCs. In this context, we found that LAG-3 blockade resulted in superior T cell activation compared to inhibition of other pathways, including PD-1/PD-L1. This result was consistent across different methods to measure T cell stimulation (proliferation, IFN-γ secretion), various stimulatory antigens (viral and bacterial peptide pool, specific viral antigen, specific tumor antigen), and seen for both CD4+ and CD8+ T cells. Only under conditions with a weak antigenic stimulus, particularly when combining antigen presentation by peripheral blood mononuclear cells with low concentrations of peptides, we observed the highest T cell stimulation with dual blockade of LAG-3 and PD-1 blockade. We conclude that priming of novel immune responses can be strongly enhanced by blockade of LAG-3 or dual blockade of LAG-3 and PD-1, depending on the strength of the antigenic stimulus. PMID:29535740

Targeting LAG-3 and PD-1 to Enhance T Cell Activation by Antigen-Presenting Cells.

PubMed

Lichtenegger, Felix S; Rothe, Maurine; Schnorfeil, Frauke M; Deiser, Katrin; Krupka, Christina; Augsberger, Christian; Schlüter, Miriam; Neitz, Julia; Subklewe, Marion

2018-01-01

Immune checkpoint inhibition has been shown to successfully reactivate endogenous T cell responses directed against tumor-associated antigens, resulting in significantly prolonged overall survival in patients with various tumor entities. For malignancies with low endogenous immune responses, this approach has not shown a clear clinical benefit so far. Therapeutic vaccination, particularly dendritic cell (DC) vaccination, is a strategy to induce T cell responses. Interaction of DCs and T cells is dependent on receptor-ligand interactions of various immune checkpoints. In this study, we analyzed the influence of blocking antibodies targeting programmed cell death protein 1 (PD-1), HVEM, CD244, TIM-3, and lymphocyte activation gene 3 (LAG-3) on the proliferation and cytokine secretion of T cells after stimulation with autologous TLR-matured DCs. In this context, we found that LAG-3 blockade resulted in superior T cell activation compared to inhibition of other pathways, including PD-1/PD-L1. This result was consistent across different methods to measure T cell stimulation (proliferation, IFN-γ secretion), various stimulatory antigens (viral and bacterial peptide pool, specific viral antigen, specific tumor antigen), and seen for both CD4 + and CD8 + T cells. Only under conditions with a weak antigenic stimulus, particularly when combining antigen presentation by peripheral blood mononuclear cells with low concentrations of peptides, we observed the highest T cell stimulation with dual blockade of LAG-3 and PD-1 blockade. We conclude that priming of novel immune responses can be strongly enhanced by blockade of LAG-3 or dual blockade of LAG-3 and PD-1, depending on the strength of the antigenic stimulus.

Novel fusion proteins for the antigen-specific staining and elimination of B cell receptor-positive cell populations demonstrated by a tetanus toxoid fragment C (TTC) model antigen.

PubMed

Klose, Diana; Saunders, Ute; Barth, Stefan; Fischer, Rainer; Jacobi, Annett Marita; Nachreiner, Thomas

2016-02-17

In an earlier study we developed a unique strategy allowing us to specifically eliminate antigen-specific murine B cells via their distinct B cell receptors using a new class of fusion proteins. In the present work we elaborated our idea to demonstrate the feasibility of specifically addressing and eliminating human memory B cells. The present study reveals efficient adaptation of the general approach to selectively target and eradicate human memory B cells. In order to demonstrate the feasibility we engineered a fusion protein following the principle of recombinant immunotoxins by combining a model antigen (tetanus toxoid fragment C, TTC) for B cell receptor targeting and a truncated version of Pseudomonas aeruginosa exotoxin A (ETA') to induce apoptosis after cellular uptake. The TTC-ETA' fusion protein not only selectively bound to a TTC-reactive murine B cell hybridoma cell line in vitro but also to freshly isolated human memory B cells from immunized donors ex vivo. Specific toxicity was confirmed on an antigen-specific population of human CD27(+) memory B cells. This protein engineering strategy can be used as a generalized platform approach for the construction of therapeutic fusion proteins with disease-relevant antigens as B cell receptor-binding domains, offering a promising approach for the specific depletion of autoreactive B-lymphocytes in B cell-driven autoimmune diseases.

Murine Aortic Smooth Muscle Cells Acquire, Though Fail to Present Exogenous Protein Antigens on Major Histocompatibility Complex Class II Molecules

PubMed Central

Maddaluno, Marcella; MacRitchie, Neil; Grassia, Gianluca; Ialenti, Armando; Butcher, John P.; Garside, Paul; Brewer, James M.; Maffia, Pasquale

2014-01-01

In the present study aortic murine smooth muscle cell (SMC) antigen presentation capacity was evaluated using the Eα-GFP/Y-Ae system to visualize antigen uptake through a GFP tag and tracking of Eα peptide/MHCII presentation using the Y-Ae Ab. Stimulation with IFN-γ (100 ng/mL) for 72 h caused a significant (P < 0.01) increase in the percentage of MHC class II positive SMCs, compared with unstimulated cells. Treatment with Eα-GFP (100 μg/mL) for 48 h induced a significant (P < 0.05) increase in the percentage of GFP positive SMCs while it did not affect the percentage of Y-Ae positive cells, being indicative of antigen uptake without its presentation in the context of MHC class II. After IFN-γ-stimulation, ovalbumin- (OVA, 1 mg/mL) or OVA323–339 peptide-(0.5 μg/mL) treated SMCs failed to induce OT-II CD4+ T cell activation/proliferation; this was also accompanied by a lack of expression of key costimulatory molecules (OX40L, CD40, CD70, and CD86) on SMCs. Finally, OVA-treated SMCs failed to induce DO11.10-GFP hybridoma activation, a process independent of costimulation. Our results demonstrate that while murine primary aortic SMCs express MHC class II and can acquire exogenous antigens, they fail to activate T cells through a failure in antigen presentation and a lack of costimulatory molecule expression. PMID:25136640

Antigen presentation by MART-1 adenovirus-transduced interleukin-10-polarized human monocyte-derived dendritic cells

PubMed Central

Mehrotra, Shikhar; Chhabra, Arvind; Chakraborty, Abolokita; Chattopadhyay, Subhasis; Slowik, Mark; Stevens, Robert; Zengou, Ryan; Mathias, Clinton; Butterfield, Lisa H; Dorsky, David I; Economou, James S; Mukherji, Bijay; Chakraborty, Nitya G

2004-01-01

Dendritic cells (DC) play critical roles in generating an immune response and in inducing tolerance. Diverse microenvironmental factors can ‘polarize’ DC toward an immunogenic or non-immunogenic phenotype. Among the various microenvironmental factors, interleukin-10 (IL-10) exhibits a potent immunosuppressive effect on antigen-presenting cells (APC). Here, we show that monocyte-derived DC generated in the presence of IL-10 exhibit a profound down-regulation of many genes that are associated with immune activation and show that the IL-10-grown DC are poor stimulators of CD8+ T cells in a strictly autologous and major histocompatibility complex (MHC) class I-restricted melanoma antigen recognized by T cells (MART-1) epitope presentation system. However, these IL-10-grown DC can efficiently activate the epitope-specific CD8+ T cells when they are made to present the epitope following transduction with an adenoviral vector expressing the MART-1 antigen. In addition, we show that the MART-1 protein colocalizes with the MHC class I protein, equally well, in the iDC and in the DC cultured in presence of IL-10 when both DC types are infected with the viral vector. We also show that the vector transduced DC present the MART-127–35 epitope for a sustained period compared to the peptide pulsed DC. These data suggest that although DCs generated in the presence of IL-10 tend to be non-immunogenic, they are capable of processing and presenting an antigen when the antigen is synthesized within the DC. PMID:15554925

Differences in antigen presentation to MHC class I-and class II- restricted influenza virus-specific cytolytic T lymphocyte clones

PubMed Central

1986-01-01

We have examined requirements for antigen presentation to a panel of MHC class I-and class II-restricted, influenza virus-specific CTL clones by controlling the form of virus presented on the target cell surface. Both H-2K/D- and I region-restricted CTL recognize target cells exposed to infectious virus, but only the I region-restricted clones efficiently lysed histocompatible target cells pulsed with inactivated virus preparations. The isolated influenza hemagglutinin (HA) polypeptide also could sensitize target cells for recognition by class II-restricted, HA-specific CTL, but not by class I-restricted, HA- specific CTL. Inhibition of nascent viral protein synthesis abrogated the ability of target cells to present viral antigen relevant for class I-restricted CTL recognition. Significantly, presentation for class II- restricted recognition was unaffected in target cells exposed to preparations of either inactivated or infectious virus. This differential sensitivity suggested that these H-2I region-restricted CTL recognized viral polypeptides derived from the exogenously introduced virions, rather than viral polypeptides newly synthesized in the infected cell. In support of this contention, treatment of the target cells with the lysosomotropic agent chloroquine abolished recognition of infected target cells by class II-restricted CTL without diminishing class I-restricted recognition of infected target cells. Furthermore, when the influenza HA gene was introduced into target cells without exogenous HA polypeptide, the target cells that expressed the newly synthesized protein product of the HA gene were recognized only by H-2K/D-restricted CTL. These observations suggest that important differences may exist in requirements for antigen presentation between H-2K/D and H-2I region-restricted CTL. These differences may reflect the nature of the antigenic epitopes recognized by these two CTL subsets. PMID:3485173

Brain Ischemia Induces Diversified Neuroantigen-Specific T-Cell Responses That Exacerbate Brain Injury.

PubMed

Jin, Wei-Na; Gonzales, Rayna; Feng, Yan; Wood, Kristofer; Chai, Zhi; Dong, Jing-Fei; La Cava, Antonio; Shi, Fu-Dong; Liu, Qiang

2018-06-01

Autoimmune responses can occur when antigens from the central nervous system are presented to lymphocytes in the periphery or central nervous system in several neurological diseases. However, whether autoimmune responses emerge after brain ischemia and their impact on clinical outcomes remains controversial. We hypothesized that brain ischemia facilitates the genesis of autoimmunity and aggravates ischemic brain injury. Using a mouse strain that harbors a transgenic T-cell receptor to a central nervous system antigen, MOG 35-55 (myelin oligodendrocyte glycoprotein) epitope (2D2), we determined the anatomic location and involvement of antigen-presenting cells in the development of T-cell reactivity after brain ischemia and how T-cell reactivity impacts stroke outcome. Transient middle cerebral artery occlusion and photothrombotic stroke models were used in this study. We also quantified the presence and status of T cells from brain slices of ischemic patients. By coupling transfer of labeled MOG 35-55 -specific (2D2) T cells with tetramer tracking, we show an expansion in reactivity of 2D2 T cells to MOG 91-108 and MOG 103-125 in transient middle cerebral artery occlusion and photothrombotic stroke models. This reactivity and T-cell activation first occur locally in the brain after ischemia. Also, microglia act as antigen-presenting cells that effectively present MOG antigens, and depletion of microglia ablates expansion of 2D2 reactive T cells. Notably, the adoptive transfer of neuroantigen-experienced 2D2 T cells exacerbates Th1/Th17 responses and brain injury. Finally, T-cell activation and MOG-specific T cells are present in the brain of patients with ischemic stroke. Our findings suggest that brain ischemia activates and diversifies T-cell responses locally, which exacerbates ischemic brain injury. © 2018 The Authors.

Human MAIT-cell responses to Escherichia coli: activation, cytokine production, proliferation, and cytotoxicity.

PubMed

Dias, Joana; Sobkowiak, Michał J; Sandberg, Johan K; Leeansyah, Edwin

2016-07-01

Mucosa-associated invariant T cells are a large and relatively recently described innate-like antimicrobial T-cell subset in humans. These cells recognize riboflavin metabolites from a range of microbes presented by evolutionarily conserved major histocompatibility complex, class I-related molecules. Given the innate-like characteristics of mucosa-associated invariant T cells and the novel type of antigens they recognize, new methodology must be developed and existing methods refined to allow comprehensive studies of their role in human immune defense against microbial infection. In this study, we established protocols to examine a range of mucosa-associated invariant T-cell functions as they respond to antigen produced by Escherichia coli These improved and dose- and time-optimized experimental protocols allow detailed studies of MR1-dependent mucosa-associated invariant T-cell responses to Escherichia coli pulsed antigen-presenting cells, as assessed by expression of activation markers and cytokines, by proliferation, and by induction of apoptosis and death in major histocompatibility complex, class I-related-expressing target cells. The novel and optimized protocols establish a framework of methods and open new possibilities to study mucosa-associated invariant T-cell immunobiology, using Escherichia coli as a model antigen. Furthermore, we propose that these robust experimental systems can also be adapted to study mucosa-associated invariant T-cell responses to other microbes and types of antigen-presenting cells. © The Author(s).

Phylogeny of Immune Recognition: Processing and Presentation of Structurally Defined Proteins in Channel Catfish Immune Responses

PubMed Central

Vallejo, Abbe N.; Miller, Norman W.

1991-01-01

This work was undertaken to investigate whether or not antigen processing and presentation are important in channel catfish in vitro secondary immune responses elicited with structurally defined proteins, namely, pigeon heart cytochrome C (pCytC), hen egg lysozyme, and horse myoglobin. The use of in vitro antigen-pulsed and fixed B cells or monocytes as antigen presenting cells (APC) resulted in autologous peripheral blood leukocytes (PBL) responding with vigorous proliferation and antibody production in vitro. In addition, several long-term catfish monocyte lines have been found to function as efficient APC with autologous but not allogeneic responders. Subsequent separation of the responding PBL into sIg- (T-cell-enriched) and B (sIg+) cell subsets showed that both underwent proliferative responses to antigen-pulsed and fixed APC. Moreover, allogeneic cells used as APC were found to induce only strong mixed leukocyte reactions without specific in vitro antibody production. Initial attempts at identifying the immunogenic region(s) of the protein antigens for catfish indicated there are two such regions for pCytC, namely, peptides 66-80 and 81-104. PMID:1668258

Eliciting Epitope-Specific CD8+ T Cell Response by Immunization with Microbial Protein Antigens Formulated with α-Galactosylceramide: Theory, Practice, and Protocols.

PubMed

Gilchuk, Pavlo; Knight, Frances C; Wilson, John T; Joyce, Sebastian

2017-01-01

CD8+ cytotoxic T lymphocytes confer protection against infectious diseases caused by viruses, bacteria, and parasites. Hence, significant efforts have been invested into devising ways to generate CD8+ T cell-targeted vaccines. Generation of microbe-free protein subunit vaccines requires a thorough knowledge of protective target antigens. Such antigens are proteolytically processed peptides presented by MHC class I molecules. To induce a robust antigen-specific CD8+ T cell response through vaccination, it is essential to formulate the antigen with an effective adjuvant. Here, we describe a versatile method for generating high-frequency antigen-specific CD8+ T cells through immunization of mice using the invariant natural killer T cell agonist α-galactosylceramide as the adjuvant.

Tumor necrosis factor-alpha stimulates the production of squamous cell carcinoma antigen in normal squamous cells.

PubMed

Numa, F; Takeda, O; Nakata, M; Nawata, S; Tsunaga, N; Hirabayashi, K; Suminami, Y; Kato, H; Hamanaka, S

1996-01-01

Squamous cell carcinoma (SCC) antigen, a tumor marker of squamous cell carcinoma, is also increased in several nonmalignant skin lesions, e.g. pemphigus. The aim of the present investigation was to determine if tumor necrosis factor-alpha (TNF-alpha), one of the important environmental factors, stimulated the production of SCC antigen in the normal squamous cells. The exposure of normal human epidermal keratinocytes to TNF-alpha (100 IU/ml) for 72 h greatly increased the SCC antigen production. The stimulatory effect of TNF-alpha (1,000 IU/ml) on the production of SCC antigen was also observed in the normal squamous epithelium tissue. These results would be helpful for understanding the increase of SCC antigen in several nonmalignant skin disorders.

Antigen-Presenting Intratumoral B Cells Affect CD4+ TIL Phenotypes in Non-Small Cell Lung Cancer Patients.

PubMed

Bruno, Tullia C; Ebner, Peggy J; Moore, Brandon L; Squalls, Olivia G; Waugh, Katherine A; Eruslanov, Evgeniy B; Singhal, Sunil; Mitchell, John D; Franklin, Wilbur A; Merrick, Daniel T; McCarter, Martin D; Palmer, Brent E; Kern, Jeffrey A; Slansky, Jill E

2017-10-01

Effective immunotherapy options for patients with non-small cell lung cancer (NSCLC) are becoming increasingly available. The immunotherapy focus has been on tumor-infiltrating T cells (TILs); however, tumor-infiltrating B cells (TIL-Bs) have also been reported to correlate with NSCLC patient survival. The function of TIL-Bs in human cancer has been understudied, with little focus on their role as antigen-presenting cells and their influence on CD4 + TILs. Compared with other immune subsets detected in freshly isolated primary tumors from NSCLC patients, we observed increased numbers of intratumoral B cells relative to B cells from tumor-adjacent tissues. Furthermore, we demonstrated that TIL-Bs can efficiently present antigen to CD4 + TILs and alter the CD4 + TIL phenotype using an in vitro antigen-presentation assay. Specifically, we identified three CD4 + TIL responses to TIL-Bs, which we categorized as activated, antigen-associated, and nonresponsive. Within the activated and antigen-associated CD4 + TIL population, activated TIL-Bs (CD19 + CD20 + CD69 + CD27 + CD21 + ) were associated with an effector T-cell response (IFNγ + CD4 + TILs). Alternatively, exhausted TIL-Bs (CD19 + CD20 + CD69 + CD27 - CD21 - ) were associated with a regulatory T-cell phenotype (FoxP3 + CD4 + TILs). Our results demonstrate a new role for TIL-Bs in NSCLC tumors in their interplay with CD4 + TILs in the tumor microenvironment, establishing them as a potential therapeutic target in NSCLC immunotherapy. Cancer Immunol Res; 5(10); 898-907. ©2017 AACR . ©2017 American Association for Cancer Research.

Efficient Culture of Human Naïve and Memory B cells for Use as Antigen-presenting Cells

PubMed Central

Su, Kuei-Ying; Watanabe, Akiko; Yeh, Chen-Hao; Kelsoe, Garnett; Kuraoka, Masayuki

2016-01-01

The ability to culture and expand B cells in vitro has become a useful tool for studying human immunity. A limitation of current methods for human B-cell culture is the capacity to support mature B-cell proliferation. We have developed a culture method to support the efficient activation and proliferation of both naïve and memory human B cells. This culture supports extensive B-cell proliferation, with approximately 103-fold increases following 8 days in culture, and 106-fold increases when cultures are split and cultured for 8 more days. In culture, a significant fraction of naïve B cells undergo isotype switching and differentiate into plasmacytes. Culture-derived (CD) B cells are readily cryopreserved, and when recovered, retain their ability to proliferate and differentiate. Significantly, proliferating CD B cells express high levels of MHCII, CD80, and CD86. CD B cells act as APCs and present both alloantigens and microbial antigens to T cells. We are able to activate and expand antigen-specific memory B cells; these cultured cells are highly effective in presenting antigen to T cells. We have characterized the TCR repertoire of rare antigen-specific CD4+ T cells that proliferated in response to tetanus toxoid (TT) presented by autologous CD B cells. TCR Vβ usage by TT-activated CD4+ T cells differs from both resting and unspecifically activated CD4+ T cells. Moreover, we found that TT-specific TCR Vβ usage by CD4+ T cells was substantially different between donors. This culture method provides a platform for studying the BCR and TCR repertoires within a single individual. PMID:27815447

Autologous cellular vaccine overcomes cancer immunoediting in a mouse model of myeloma.

PubMed

Mazzocco, Marta; Martini, Matteo; Rosato, Antonio; Stefani, Elisabetta; Matucci, Andrea; Dalla Santa, Silvia; De Sanctis, Francesco; Ugel, Stefano; Sandri, Sara; Ferrarini, Giovanna; Cestari, Tiziana; Ferrari, Sergio; Zanovello, Paola; Bronte, Vincenzo; Sartoris, Silvia

2015-09-01

In the Sp6 mouse plasmacytoma model, a whole-cell vaccination with Sp6 cells expressing de novo B7-1 (Sp6/B7) induced anatomically localized and cytotoxic T cell (CTL)-mediated protection against wild-type (WT) Sp6. Both WT Sp6 and Sp6/B7 showed down-regulated expression of MHC H-2 L(d). Increase of H-2 L(d) expression by cDNA transfection (Sp6/B7/L(d)) raised tumour immune protection and shifted most CTL responses towards H-2 L(d)-restricted antigenic epitopes. The tumour-protective responses were not specific for the H-2 L(d)-restricted immunodominant AH1 epitope of the gp70 common mouse tumour antigen, although WT Sp6 and transfectants were able to present it to specific T cells in vitro. Gp70 transcripts, absent in secondary lymphoid organs of naive mice, were detected in immunized mice as well as in splenocytes from naive mice incubated in vitro with supernatants of CTL-lysed Sp6 cell cultures, containing damage-associated molecular patterns (DAMPs). It has been shown that Toll-like receptor triggering induces gp70 expression. Damage-associated molecular patterns are released by CTL-mediated killing of Sp6/B7-Sp6/B7/L(d) cells migrated to draining lymph nodes during immunization and may activate gp70 expression and presentation in most resident antigen-presenting cells. The same could also apply for Mus musculus endogenous ecotropic murine leukaemia virus 1 particles present in Sp6-cytosol, discharged by dying cells and superinfecting antigen-presenting cells. The outcome of such a massive gp70 cross-presentation would probably be tolerogenic for the high-affinity AH1-gp70-specific CTL clones. In this scenario, autologous whole-tumour-cell vaccines rescue tumour-specific immunoprotection by amplification of subdominant tumour antigen responses when those against the immune dominant antigens are lost. © 2015 The Authors. Immunology Published by John Wiley & Sons Ltd.

Autologous cellular vaccine overcomes cancer immunoediting in a mouse model of myeloma

PubMed Central

Mazzocco, Marta; Martini, Matteo; Rosato, Antonio; Stefani, Elisabetta; Matucci, Andrea; Dalla Santa, Silvia; De Sanctis, Francesco; Ugel, Stefano; Sandri, Sara; Ferrarini, Giovanna; Cestari, Tiziana; Ferrari, Sergio; Zanovello, Paola; Bronte, Vincenzo; Sartoris, Silvia

2015-01-01

In the Sp6 mouse plasmacytoma model, a whole-cell vaccination with Sp6 cells expressing de novo B7-1 (Sp6/B7) induced anatomically localized and cytotoxic T cell (CTL) -mediated protection against wild-type (WT) Sp6. Both WT Sp6 and Sp6/B7 showed down-regulated expression of MHC H-2 Ld. Increase of H-2 Ld expression by cDNA transfection (Sp6/B7/Ld) raised tumour immune protection and shifted most CTL responses towards H-2 Ld-restricted antigenic epitopes. The tumour-protective responses were not specific for the H-2 Ld-restricted immunodominant AH1 epitope of the gp70 common mouse tumour antigen, although WT Sp6 and transfectants were able to present it to specific T cells in vitro. Gp70 transcripts, absent in secondary lymphoid organs of naive mice, were detected in immunized mice as well as in splenocytes from naive mice incubated in vitro with supernatants of CTL-lysed Sp6 cell cultures, containing damage-associated molecular patterns (DAMPs). It has been shown that Toll-like receptor triggering induces gp70 expression. Damage-associated molecular patterns are released by CTL-mediated killing of Sp6/B7-Sp6/B7/Ld cells migrated to draining lymph nodes during immunization and may activate gp70 expression and presentation in most resident antigen-presenting cells. The same could also apply for Mus musculus endogenous ecotropic murine leukaemia virus 1 particles present in Sp6-cytosol, discharged by dying cells and superinfecting antigen-presenting cells. The outcome of such a massive gp70 cross-presentation would probably be tolerogenic for the high-affinity AH1-gp70-specific CTL clones. In this scenario, autologous whole-tumour-cell vaccines rescue tumour-specific immunoprotection by amplification of subdominant tumour antigen responses when those against the immune dominant antigens are lost. PMID:25959091

Identification of a dendritic cell receptor that couples sensing of necrosis to immunity.

PubMed

Sancho, David; Joffre, Olivier P; Keller, Anna M; Rogers, Neil C; Martínez, Dolores; Hernanz-Falcón, Patricia; Rosewell, Ian; Reis e Sousa, Caetano

2009-04-16

Injury or impaired clearance of apoptotic cells leads to the pathological accumulation of necrotic corpses, which induce an inflammatory response that initiates tissue repair. In addition, antigens present in necrotic cells can sometimes provoke a specific immune response and it has been argued that necrosis could explain adaptive immunity in seemingly infection-free situations, such as after allograft transplantation or in spontaneous and therapy-induced tumour rejection. In the mouse, the CD8alpha+ subset of dendritic cells phagocytoses dead cell remnants and cross-primes CD8+ T cells against cell-associated antigens. Here we show that CD8alpha+ dendritic cells use CLEC9A (also known as DNGR-1), a recently-characterized C-type lectin, to recognize a preformed signal that is exposed on necrotic cells. Loss or blockade of CLEC9A does not impair the uptake of necrotic cell material by CD8+ dendritic cells, but specifically reduces cross-presentation of dead-cell-associated antigens in vitro and decreases the immunogenicity of necrotic cells in vivo. The function of CLEC9A requires a key tyrosine residue in its intracellular tail that allows the recruitment and activation of the tyrosine kinase SYK, which is also essential for cross-presentation of dead-cell-associated antigens. Thus, CLEC9A functions as a SYK-coupled C-type lectin receptor to mediate sensing of necrosis by the principal dendritic-cell subset involved in regulating cross-priming to cell-associated antigens.

Autophagy-related protein Vps34 controls the homeostasis and function of antigen cross-presenting CD8α+ dendritic cells.

PubMed

Parekh, Vrajesh V; Pabbisetty, Sudheer K; Wu, Lan; Sebzda, Eric; Martinez, Jennifer; Zhang, Jianhua; Van Kaer, Luc

2017-08-01

The class III PI3K Vacuolar protein sorting 34 (Vps34) plays a role in both canonical and noncanonical autophagy, key processes that control the presentation of antigens by dendritic cells (DCs) to naive T lymphocytes. We generated DC-specific Vps34 -deficient mice to assess the contribution of Vps34 to DC functions. We found that DCs from these animals have a partially activated phenotype, spontaneously produce cytokines, and exhibit enhanced activity of the classic MHC class I and class II antigen-presentation pathways. Surprisingly, these animals displayed a defect in the homeostatic maintenance of splenic CD8α + DCs and in the capacity of these cells to cross-present cell corpse-associated antigens to MHC class I-restricted T cells, a property that was associated with defective expression of the T-cell Ig mucin (TIM)-4 receptor. Importantly, mice deficient in the Vps34-associated protein Rubicon, which is critical for a noncanonical form of autophagy called "Light-chain 3 (LC3)-associated phagocytosis" (LAP), lacked such defects. Finally, consistent with their defect in the cross-presentation of apoptotic cells, DC-specific Vps34 -deficient animals developed increased metastases in response to challenge with B16 melanoma cells. Collectively, our studies have revealed a critical role of Vps34 in the regulation of CD8α + DC homeostasis and in the capacity of these cells to process and present antigens associated with apoptotic cells to MHC class I-restricted T cells. Our findings also have important implications for the development of small-molecule inhibitors of Vps34 for therapeutic purposes.

Epitope diversification driven by non-tumor epitope-specific Th1 and Th17 mediates potent antitumor reactivity.

PubMed

Ichikawa, Kosuke; Kagamu, Hiroshi; Koyama, Kenichi; Miyabayashi, Takao; Koshio, Jun; Miura, Satoru; Watanabe, Satoshi; Yoshizawa, Hirohisa; Narita, Ichiei

2012-09-21

MHC class I-restricted peptide-based vaccination therapies have been conducted to treat cancer patients, because CD8⁺ CTL can efficiently induce apoptosis of tumor cells in an MHC class I-restricted epitope-specific manner. Interestingly, clinical responders are known to demonstrate reactivity to epitopes other than those used for vaccination; however, the mechanism underlying how antitumor T cells with diverse specificity are induced is unclear. In this study, we demonstrated that dendritic cells (DCs) that engulfed apoptotic tumor cells in the presence of non-tumor MHC class II-restricted epitope peptides, OVA(323-339), efficiently presented tumor-associated antigens upon effector-dominant CD4⁺ T cell balance against regulatory T cells (Treg) for the OVA(323-339) epitope. Th1 and Th17 induced tumor-associated antigens presentation of DC, while Th2 ameliorated tumor-antigen presentation for CD8⁺ T cells. Blocking experiments with anti-IL-23p19 antibody and anti-IL-23 receptor indicated that an autocrine mechanism of IL-23 likely mediated the diverted tumor-associated antigens presentation of DC. Tumor-associated antigens presentation of DC induced by OVA(323-339) epitope-specific CD4⁺ T cells resulted in facilitated antitumor immunity in both priming and effector phase in vivo. Notably, this immunotherapy did not require pretreatment to reduce Treg induced by tumor. This strategy may have clinical implications for designing effective antitumor immunotherapies. Copyright © 2012 Elsevier Ltd. All rights reserved.

The translocon protein Sec61 mediates antigen transport from endosomes in the cytosol for cross-presentation to CD8(+) T cells.

PubMed

Zehner, Matthias; Marschall, Andrea L; Bos, Erik; Schloetel, Jan-Gero; Kreer, Christoph; Fehrenschild, Dagmar; Limmer, Andreas; Ossendorp, Ferry; Lang, Thorsten; Koster, Abraham J; Dübel, Stefan; Burgdorf, Sven

2015-05-19

The molecular mechanisms regulating antigen translocation into the cytosol for cross-presentation are under controversial debate, mainly because direct data is lacking. Here, we have provided direct evidence that the activity of the endoplasmic reticulum (ER) translocon protein Sec61 is essential for endosome-to-cytosol translocation. We generated a Sec61-specific intrabody, a crucial tool that trapped Sec61 in the ER and prevented its recruitment into endosomes without influencing Sec61 activity and antigen presentation in the ER. Expression of this ER intrabody inhibited antigen translocation and cross-presentation, demonstrating that endosomal Sec61 indeed mediates antigen transport across endosomal membranes. Moreover, we showed that the recruitment of Sec61 toward endosomes, and hence antigen translocation and cross-presentation, is dependent on dendritic cell activation by Toll-like receptor (TLR) ligands. These data shed light on a long-lasting question regarding antigen cross-presentation and point out a role of the ER-associated degradation machinery in compartments distinct from the ER. Copyright © 2015 Elsevier Inc. All rights reserved.

Cooperation between Epstein-Barr Virus Immune Evasion Proteins Spreads Protection from CD8+ T Cell Recognition across All Three Phases of the Lytic Cycle

PubMed Central

Quinn, Laura L.; Zuo, Jianmin; Abbott, Rachel J. M.; Shannon-Lowe, Claire; Tierney, Rosemary J.; Hislop, Andrew D.; Rowe, Martin

2014-01-01

CD8+ T cell responses to Epstein-Barr virus (EBV) lytic cycle expressed antigens display a hierarchy of immunodominance, in which responses to epitopes of immediate-early (IE) and some early (E) antigens are more frequently observed than responses to epitopes of late (L) expressed antigens. It has been proposed that this hierarchy, which correlates with the phase-specific efficiency of antigen presentation, may be due to the influence of viral immune-evasion genes. At least three EBV-encoded genes, BNLF2a, BGLF5 and BILF1, have the potential to inhibit processing and presentation of CD8+ T cell epitopes. Here we examined the relative contribution of these genes to modulation of CD8+ T cell recognition of EBV lytic antigens expressed at different phases of the replication cycle in EBV-transformed B-cells (LCLs) which spontaneously reactivate lytic cycle. Selective shRNA-mediated knockdown of BNLF2a expression led to more efficient recognition of immediate-early (IE)- and early (E)-derived epitopes by CD8+ T cells, while knock down of BILF1 increased recognition of epitopes from E and late (L)-expressed antigens. Contrary to what might have been predicted from previous ectopic expression studies in EBV-negative model cell lines, the shRNA-mediated inhibition of BGLF5 expression in LCLs showed only modest, if any, increase in recognition of epitopes expressed in any phase of lytic cycle. These data indicate that whilst BNLF2a interferes with antigen presentation with diminishing efficiency as lytic cycle progresses (IE>E>>L), interference by BILF1 increases with progression through lytic cycle (IE

Peptide-Conjugated Nanoparticles Reduce Positive Co-stimulatory Expression and T Cell Activity to Induce Tolerance.

PubMed

Kuo, Robert; Saito, Eiji; Miller, Stephen D; Shea, Lonnie D

2017-07-05

Targeted approaches to treat autoimmune diseases would improve upon current therapies that broadly suppress the immune system and lead to detrimental side effects. Antigen-specific tolerance was induced using poly(lactide-co-glycolide) nanoparticles conjugated with disease-relevant antigen to treat a model of multiple sclerosis. Increasing the nanoparticle dose and amount of conjugated antigen both resulted in more durable immune tolerance. To identify active tolerance mechanisms, we investigated downstream cellular and molecular events following nanoparticle internalization by antigen-presenting cells. The initial cell response to nanoparticles indicated suppression of inflammatory signaling pathways. Direct and functional measurement of surface MHC-restricted antigen showed positive correlation with both increasing particle dose from 1 to 100 μg/mL and increasing peptide conjugation by 2-fold. Co-stimulatory analysis of cells expressing MHC-restricted antigen revealed most significant decreases in positive co-stimulatory molecules (CD86, CD80, and CD40) following high doses of nanoparticles with higher peptide conjugation, whereas expression of a negative co-stimulatory molecule (PD-L1) remained high. T cells isolated from mice immunized against myelin proteolipid protein (PLP 139-151 ) were co-cultured with antigen-presenting cells administered PLP 139-151 -conjugated nanoparticles, which resulted in reduced T cell proliferation, increased T cell apoptosis, and a stronger anti-inflammatory response. These findings indicate several potential mechanisms used by peptide-conjugated nanoparticles to induce antigen-specific tolerance. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

Detecting T-cell reactivity to whole cell vaccines

PubMed Central

Brusic, Ana; Hainz, Ursula; Wadleigh, Martha; Neuberg, Donna; Su, Mei; Canning, Christine M.; DeAngelo, Daniel J.; Stone, Richard M.; Lee, Jeng-Shin; Mulligan, Richard C.; Ritz, Jerome; Dranoff, Glenn; Sasada, Tetsuro; Wu, Catherine J.

2012-01-01

BCR-ABL+ K562 cells hold clinical promise as a component of cancer vaccines, either as bystander cells genetically modified to express immunostimulatory molecules, or as a source of leukemia antigens. To develop a method for detecting T-cell reactivity against K562 cell-derived antigens in patients, we exploited the dendritic cell (DC)-mediated cross-presentation of proteins generated from apoptotic cells. We used UVB irradiation to consistently induce apoptosis of K562 cells, which were then fed to autologous DCs. These DCs were used to both stimulate and detect antigen-specific CD8+ T-cell reactivity. As proof-of-concept, we used cross-presented apoptotic influenza matrix protein-expressing K562 cells to elicit reactivity from matrix protein-reactive T cells. Likewise, we used this assay to detect increased anti-CML antigen T-cell reactivity in CML patients that attained long-lasting clinical remissions following immunotherapy (donor lymphocyte infusion), as well as in 2 of 3 CML patients vaccinated with lethally irradiated K562 cells that were modified to secrete high levels of granulocyte macrophage colony-stimulating factor (GM-CSF). This methodology can be readily adapted to examine the effects of other whole tumor cell-based vaccines, a scenario in which the precise tumor antigens that stimulate immune responses are unknown. PMID:23170257

Discovery of a protective Rickettsia prowazekii antigen recognized by CD8+ T cells, RP884, using an in vivo screening platform.

PubMed

Gazi, Michal; Caro-Gomez, Erika; Goez, Yenny; Cespedes, Maria A; Hidalgo, Marylin; Correa, Paula; Valbuena, Gustavo

2013-01-01

Rickettsia prowazekii has been tested for biological warfare due to the high mortality that it produces after aerosol transmission of very low numbers of rickettsiae. Epidemic typhus, the infection caused by these obligately intracellular bacteria, continues to be a threat because it is difficult to diagnose due to initial non-specific symptoms and the lack of commercial diagnostic tests that are sensitive and specific during the initial clinical presentation. A vaccine to prevent epidemic typhus would constitute an effective deterrent to the weaponization of R. prowazekii; however, an effective and safe vaccine is not currently available. Due to the cytoplasmic niche of Rickettsia, CD8(+) T-cells are critical effectors of immunity; however, the identification of antigens recognized by these cells has not been systematically addressed. To help close this gap, we designed an antigen discovery strategy that uses cell-based vaccination with antigen presenting cells expressing microbe's proteins targeted to the MHC class I presentation pathway. We report the use of this method to discover a protective T-cell rickettsial antigen, RP884, among a test subset of rickettsial proteins.

Chemotherapy Enhances Cross-Presentation of Nuclear Tumor Antigens

PubMed Central

Anyaegbu, Chidozie C.; Lake, Richard A.; Heel, Kathy; Robinson, Bruce W.; Fisher, Scott A.

2014-01-01

Cross-presentation of tumor antigen is essential for efficient priming of naïve CD8+ T lymphocytes and induction of effective anti-tumor immunity. We hypothesized that the subcellular location of a tumor antigen could affect the efficiency of cross-presentation, and hence the outcome of anti-tumor responses to that antigen. We compared cross-presentation of a nominal antigen expressed in the nuclear, secretory, or cytoplasmic compartments of B16 melanoma tumors. All tumors expressed similar levels of the antigen. The antigen was cross-presented from all compartments but when the concentration was low, nuclear antigen was less efficiently cross-presented than antigen from other cellular locations. The efficiency of cross-presentation of the nuclear antigen was improved following chemotherapy-induced tumor cell apoptosis and this correlated with an increase in the proportion of effector CTL. These data demonstrate that chemotherapy improves nuclear tumor antigen cross-presentation and could be important for anti-cancer immunotherapies that target nuclear antigens. PMID:25243472

Nuclear location of an endogenously expressed antigen, EBNA1, restricts access to macroautophagy and the range of CD4 epitope display.

PubMed

Leung, Carol S; Haigh, Tracey A; Mackay, Laura K; Rickinson, Alan B; Taylor, Graham S

2010-02-02

Whereas exogenously acquired proteins are the major source of antigens feeding the MHC class II pathway in antigen-presenting cells, some endogenously expressed antigens also access that pathway but the rules governing such access are poorly understood. Here we address this using Epstein-Barr virus (EBV)-coded nuclear antigen EBNA1, a protein naturally expressed in EBV-infected B lymphoblastoid cell lines (LCLs) and a source of multiple CD4(+) T cell epitopes. Using CD4(+) T cell clones against three indicator epitopes, we find that two epitopes are weakly displayed on the LCL surface whereas the third is undetectable, a pattern of limited epitope presentation that is maintained even when nuclear expression of EBNA1 is induced to high supraphysiological levels. Inhibitor and siRNA studies show that, of the two epitopes weakly presented under these conditions, one involves macroautophagy, and the second involves antigen delivery to the MHC II pathway by another endogenous route. In contrast, when EBNA1 is expressed as a cytoplasmic protein, all three CD4 epitopes are processed and presented much more efficiently, and all involve macroautophagy. We conclude that EBNA1's nuclear location limits its accessibility to the macroautophagy pathway and, in consequence, limits the level and range of EBNA1 CD4 epitopes naturally displayed on the infected cell surface.

Leukemia-associated antigens in man.

PubMed

Brown, G; Capellaro, D; Greaves, M

1975-12-01

Rabbit antisera raised against acute lymphoblastic leukemia (ALL) cells were used to distinguish ALL from other leukemias, to identify rare leukemia cells in the bone marrow of patients in remission, and to define human leukemia-associated antigens. Antibody binding was studied with the use of immunofluorescence reagents and the analytic capacity of the Fluorescence Activated Cell Sorter-1 (FACS-1). The results indicated that most non-T-cell ALL have three leukemia-associated antigens on their surface which are absent from normal lymphoid cells: 1) an antigen shared with myelocytes, myeloblastic leukemia cells, and fetal liver (hematopoietic) cells; 2) an antigen shared with a subset of intermediate normoblasts in normal bone marrow and fetal liver; and 3) an antigen found thus far only on non-T-cell ALL and in some acute undifferentiated leukemias, which we therefore regard as a strong candidate for a leukemia-specific antigen. These antigens are absent from a subgroup of ALL patients in which the lymphoblasta express T-cell surface markers. Preliminary studies on the bone marrow samples of patients in remission indicated that rare leukemia cells were present in some samples. The implications of these findings with respect to the heterogeneity and cell origin(s) of ALL, its diagnosis, and its potential monitoring during treatment were discussed.

Pro-Inflammatory Activated Kupffer Cells by Lipids Induce Hepatic NKT Cells Deficiency through Activation-Induced Cell Death

PubMed Central

Tang, Tongfang; Sui, Yongheng; Lian, Min; Li, Zhiping; Hua, Jing

2013-01-01

Background Dietary lipids play an important role in the progression of non-alcoholic fatty liver disease (NAFLD) through alternation of liver innate immune response. Aims The present study was to investigate the effect of lipid on Kupffer cells phenotype and function in vivo and in vitro. And further to investigate the impact of lipid on ability of Kupffer cell lipid antigen presentation to activate NKT cells. Methods Wild type male C57BL/6 mice were fed either normal or high-fat diet. Hepatic steatosis, Kupffer cell abundance, NKT cell number and cytokine gene expression were evaluated. Antigen presentation assay was performed with Kupffer cells treated with certain fatty acids in vitro and co-cultured with NKT cells. Results High-fat diet induced hepatosteatosis, significantly increased Kupffer cells and decreased hepatic NKT cells. Lipid treatment in vivo or in vitro induced increase of pro-inflammatory cytokines gene expression and toll-like receptor 4 (TLR4) expression in Kupffer cells. Kupffer cells expressed high levels of CD1d on cell surface and only presented exogenous lipid antigen to activate NKT cells. Ability of Kupffer cells to present antigen and activate NKT cells was enhanced after lipid treatment. In addition, pro-inflammatory activated Kupffer cells by lipid treatment induced hepatic NKT cells activation-induced apoptosis and necrosis. Conclusion High-fat diet increase Kupffer cells number and induce their pro-inflammatory status. Pro-inflammatory activated Kupfffer cells by lipid promote hepatic NKT cell over-activation and cell death, which lead to further hepatic NKT cell deficiency in the development of NAFLD. PMID:24312613

Pathogenesis and spectrum of autoimmunity.

PubMed

Perl, Andras

2012-01-01

The immune system specifically recognizes and eliminates foreign antigens and, thus, protects integrity of the host. During maturation of the immune system, tolerance mechanisms develop that prevent or inhibit potentially harmful reactivities to self-antigens. Autoreactive B and T cells that are generated during immune responses are eliminated by apoptosis in the thymus, lymph nodes, or peripheral circulation or actively suppressed by regulatory T cells. However, autoreactive cells may survive due to failure of apoptosis or molecular mimicry, i.e., presentation and recognition of cryptic epitopes of self-antigens, or aberrant lymphokine production. Preservation of the host requires the development of immune responses to foreign antigen and tolerance to self-antigens. Autoimmunity results from a breakdown of tolerance to self-antigens through an interplay of genetic and environmental factors.One of the basic functions of the immune system is to specifically recognize and eliminate foreign antigens and, thus, protect integrity of the host. Through rearrangements and somatic mutations of various gene segments encoding T and B cell receptors and antibody molecules, the immune system acquires tremendous diversity. During maturation of the immune system, recognition of self-antigens plays an important role in shaping the repertoires of immune receptors. Tolerance mechanisms develop that prevent or inhibit potentially harmful reactivities to self-antigens. These self-defense mechanisms are mediated on the levels of central and peripheral tolerance, i.e., autoreactive T cells are either eliminated by apoptosis in the thymus, lymph nodes, or peripheral circulation or actively suppressed by regulatory T cells. Likewise, autoreactive B cells are eliminated in the bone marrow or peripheral lymphoid organs. However, immune responses triggered by foreign antigens may be sustained by molecular mimicry, i.e., presentation and recognition of cryptic epitopes of self-antigens. Further downstream, execution of immune responses depends on the functioning of intracellular signaling networks and the cooperation of many cell types communicating via surface receptors, cytokines, chemokines, and antibody molecules. Therefore, autoimmunity represents the end result of the breakdown of one or multiple basic mechanisms of immune tolerance (Table 1).

Antigen-specific helper factors present in the supernatant of concanavalin A-induced spleen cell cultures.

PubMed

Kilburn, D G; Anaka, R

1981-08-01

The supernatants from cultures of concanavalin A-induced spleen cells contained both antigen-specific and nonspecific (Interleukin 2) helper factors (Hf). The antigen-specific factor could be isolated from the supernatant by adsorption onto and elution from antigen-Sepharose immunoadsorbents. Specific Hf was produced in cultures of either immune or nonimmune spleen cells although in the latter case the quantity of Hf was significantly less. The specific Hf did not manifest the thymocyte stimulatory property of 112.

A T-Cell Receptor Breaks the Rules | Center for Cancer Research

Cancer.gov

Most mature T cells function immunologically when a T-cell receptor (TCR) located on the cell surface encounters and engages its ligand, a major histocompatability complex (MHC), which displays a specific part of a target protein called an antigen. This antigen-presenting complex is assembled from one of the dozen or so MHC molecules that every person inherits from their parents; and the antigen fragment, called a peptide epitope, is excised from one of thousands of possible proteins—originally part of an invading pathogen or a cancer cell—that T cells are capable of identifying and attacking. The framework of an MHC molecule holding a centrally displayed or “presented” peptide is what engages the TCR and triggers T-cell action. This role of MHC molecules presenting antigens to the TCR is a central tenet of immunology, with the fit between a TCR and the MHC framework actually “hardwired” into their three-dimensional structures.

Human Macrophages and Dendritic Cells Can Equally Present MART-1 Antigen to CD8+ T Cells after Phagocytosis of Gamma-Irradiated Melanoma Cells

PubMed Central

Barrio, María Marcela; Abes, Riad; Colombo, Marina; Pizzurro, Gabriela; Boix, Charlotte; Roberti, María Paula; Gélizé, Emmanuelle; Rodriguez-Zubieta, Mariana

2012-01-01

Dendritic cells (DC) can achieve cross-presentation of naturally-occurring tumor-associated antigens after phagocytosis and processing of dying tumor cells. They have been used in different clinical settings to vaccinate cancer patients. We have previously used gamma-irradiated MART-1 expressing melanoma cells as a source of antigens to vaccinate melanoma patients by injecting irradiated cells with BCG and GM-CSF or to load immature DC and use them as a vaccine. Other clinical trials have used IFN-gamma activated macrophage killer cells (MAK) to treat cancer patients. However, the clinical use of MAK has been based on their direct tumoricidal activity rather than on their ability to act as antigen-presenting cells to stimulate an adaptive antitumor response. Thus, in the present work, we compared the fate of MART-1 after phagocytosis of gamma-irradiated cells by clinical grade DC or MAK as well as the ability of these cells to cross present MART-1 to CD8+ T cells. Using a high affinity antibody against MART-1, 2A9, which specifically stains melanoma tumors, melanoma cell lines and normal melanocytes, the expression level of MART-1 in melanoma cell lines could be related to their ability to stimulate IFN-gamma production by a MART-1 specific HLA-A*0201-restricted CD8+ T cell clone. Confocal microscopy with Alexa Fluor®647-labelled 2A9 also showed that MART-1 could be detected in tumor cells attached and/or fused to phagocytes and even inside these cells as early as 1 h and up to 24 h or 48 h after initiation of co-cultures between gamma-irradiated melanoma cells and MAK or DC, respectively. Interestingly, MART-1 was cross-presented to MART-1 specific T cells by both MAK and DC co-cultured with melanoma gamma-irradiated cells for different time-points. Thus, naturally occurring MART-1 melanoma antigen can be taken-up from dying melanoma cells into DC or MAK and both cell types can induce specific CD8+ T cell cross-presentation thereafter. PMID:22768350

Cellular Pathway(S) of Antigen Processing and Presentation in Fish APC: Endosomal Involvement and Cell-Free Antigen Presentation

PubMed Central

Vallejo, Abbe N.; Miller, Norman W.; Harvey, Nancy E.; Cuchens, Marvin A.; Warr, Gregory W.

1992-01-01

Studies were conducted to address further the role(s) of antigen processing and presentation in the induction of immune responses in a phylogenetically lower vertebrate, specifically a teleost, the channel catfish. In particular, studies were aimed at determining the subcellular compartments involved in antigen degradation by channel catfish antigen-presenting cells (APC) as well as ascertaining the reexpression of immunogenic peptides on the surfaces of APC. The results showed that exogenous protein antigens were actively endocytosed by APC as detected by flow cytometry. Use of radiolabeled antigen and subcellular fractionation protocols also showed that antigen localized in endosomes/lysosomes. Furthermore, there was an apparent redistribution of antigen between these organelles and the plasma membrane during the course of antigen pulsing. Functional assays for the induction of in vitro antigen-specific proliferation of immune catfish peripheral blood leukocytes (PBL) showed that membrane preparations from antigen-pulsed autologous APC were highly stimulatory. The magnitude of responses elicited with such membrane preparations was very similar to that of PBL cultures stimulated with native antigen-pulsed and fixed intact APC or prefixed intact APC incubated with a peptide fragment of the nominal antigen. Current data further corroborate our previous findings that steps akin to antigen processing and presentation are clearly important in the induction of immune responses in lower vertebrates like fish, in a manner similar to that seen in mammalian systems. Consequently, it would appear that many immune functions among the diverse taxa of vertebrates are remarkably conserved. PMID:1343103

Antigen expression level threshold tunes the fate of CD8 T cells during primary hepatic immune responses.

PubMed

Tay, Szun Szun; Wong, Yik Chun; McDonald, David M; Wood, Nicole A W; Roediger, Ben; Sierro, Frederic; Mcguffog, Claire; Alexander, Ian E; Bishop, G Alex; Gamble, Jennifer R; Weninger, Wolfgang; McCaughan, Geoffrey W; Bertolino, Patrick; Bowen, David G

2014-06-24

CD8 T-cell responses to liver-expressed antigens range from deletional tolerance to full effector differentiation resulting in overt hepatotoxicity. The reasons for these heterogeneous outcomes are not well understood. To identify factors that govern the fate of CD8 T cells activated by hepatocyte-expressed antigen, we exploited recombinant adenoassociated viral vectors that enabled us to vary potential parameters determining these outcomes in vivo. Our findings reveal a threshold of antigen expression within the liver as the dominant factor determining T-cell fate, irrespective of T-cell receptor affinity or antigen cross-presentation. Thus, when a low percentage of hepatocytes expressed cognate antigen, high-affinity T cells developed and maintained effector function, whereas, at a high percentage, they became functionally exhausted and silenced. Exhaustion was not irreversibly determined by initial activation, but was maintained by high intrahepatic antigen load during the early phase of the response; cytolytic function was restored when T cells primed under high antigen load conditions were transferred into an environment of low-level antigen expression. Our study reveals a hierarchy of factors dictating the fate of CD8 T cells during hepatic immune responses, and provides an explanation for the different immune outcomes observed in a variety of immune-mediated liver pathologic conditions.

The Human Leukocyte Antigen–presented Ligandome of B Lymphocytes*

PubMed Central

Hassan, Chopie; Kester, Michel G. D.; de Ru, Arnoud H.; Hombrink, Pleun; Drijfhout, Jan Wouter; Nijveen, Harm; Leunissen, Jack A. M.; Heemskerk, Mirjam H. M.; Falkenburg, J. H. Frederik; van Veelen, Peter A.

2013-01-01

Peptides presented by human leukocyte antigen (HLA) molecules on the cell surface play a crucial role in adaptive immunology, mediating the communication between T cells and antigen presenting cells. Knowledge of these peptides is of pivotal importance in fundamental studies of T cell action and in cellular immunotherapy and transplantation. In this paper we present the in-depth identification and relative quantification of 14,500 peptide ligands constituting the HLA ligandome of B cells. This large number of identified ligands provides general insight into the presented peptide repertoire and antigen presentation. Our uniquely large set of HLA ligands allowed us to characterize in detail the peptides constituting the ligandome in terms of relative abundance, peptide length distribution, physicochemical properties, binding affinity to the HLA molecule, and presence of post-translational modifications. The presented B-lymphocyte ligandome is shown to be a rich source of information by the presence of minor histocompatibility antigens, virus-derived epitopes, and post-translationally modified HLA ligands, and it can be a good starting point for solving a wealth of specific immunological questions. These HLA ligands can form the basis for reversed immunology approaches to identify T cell epitopes based not on in silico predictions but on the bona fide eluted HLA ligandome. PMID:23481700

Protein-scaffold Directed Nanoscale Assembly of T Cell Ligands: Artificial Antigen Presentation with Defined Valency, Density and Ratio.

PubMed

Smith, Mason R; Tolbert, Stephanie V; Wen, Fei

2018-05-07

Tuning antigen presentation to T cells is a critical step in investigating key aspects of T cell activation. However, existing technologies have limited ability to control the spatial and stoichiometric organization of T cell ligands on 3D surfaces. Here, we developed an artificial antigen presentation platform based on protein-scaffold directed assembly that allows fine control over the spatial and stoichiometric organization of T cell ligands on a 3D yeast-cell surface. Using this system, we observed that the T cell activation threshold on a 3D surface is independent of peptide-major histocompatibility complex (pMHC) valency, but instead determined by the overall pMHC surface density. When intercellular adhesion molecule 1 (ICAM-1) was co-assembled with pMHC, it enhanced antigen recognition sensitivity by 6-fold. Further, T cells responded with different magnitudes to varying ratios of pMHC and ICAM-1 and exhibited a maximum response at a ratio of 15% pMHC and 85% ICAM-1, introducing an additional parameter for tuning T cell activation. This protein-scaffold directed assembly technology is readily transferrable to acellular surfaces for translational research as well as large-scale T-cell manufacturing.

Molecular basis of mycobacterial lipid antigen presentation by CD1c and its recognition by αβ T cells

PubMed Central

Roy, Sobhan; Ly, Dalam; Li, Nan-Sheng; Altman, John D.; Piccirilli, Joseph A.; Moody, D. Branch; Adams, Erin J.

2014-01-01

CD1c is a member of the group 1 CD1 family of proteins that are specialized for lipid antigen presentation. Despite high cell surface expression of CD1c on key antigen-presenting cells and the discovery of its mycobacterial lipid antigen presentation capability, the molecular basis of CD1c recognition by T cells is unknown. Here we present a comprehensive functional and molecular analysis of αβ T-cell receptor (TCR) recognition of CD1c presenting mycobacterial phosphomycoketide antigens. Our structure of CD1c with the mycobacterial phosphomycoketide (PM) shows similarities to that of CD1c-mannosyl-β1-phosphomycoketide in that the A' pocket accommodates the mycoketide alkyl chain; however, the phosphate head-group of PM is shifted ∼6 Å in relation to that of mannosyl-β1-PM. We also demonstrate a bona fide interaction between six human TCRs and CD1c-mycoketide complexes, measuring high to moderate affinities. The crystal structure of the DN6 TCR and mutagenic studies reveal a requirement of five complementarity determining region (CDR) loops for CD1c recognition. Furthermore, mutagenesis of CD1c reveals residues in both the α1 and α2 helices involved in TCR recognition, yet not entirely overlapping among the examined TCRs. Unlike patterns for MHC I, no archetypical binding footprint is predicted to be shared by CD1c-reactive TCRs, even when recognizing the same or similar antigens. PMID:25298532

Nonidentity of Some Simian Virus 40-induced Enzymes with Tumor Antigen

PubMed Central

Kit, Saul; Melnick, Joseph L.; Anken, Milton; Dubbs, Del Rose; de Torres, R. A.; Kitahara, Tsunehiro

1967-01-01

The complement-fixing tumor (T) antigen induced by simian virus 40 (SV40) has been prepared from SV40-infected cell cultures, from infected cell cultures treated at the time of infection with 1-β-d-arabinofuranosylcytosine (ara-C), and from SV40-transformed cells. Upon partial purification, the T antigen exhibited the following properties: it was tightly adsorbed by calcium phosphate gel, it was precipitated by acetic acid at pH 5 or by ammonium sulfate at about 20 to 32% saturation, and it had a molecular weight greater than 250,000, as estimated by Sephadex G-200 gel chromatography. In contrast, deoxycytidylate (dCMP) deaminase, thymidylate (dTMP) kinase, and thymidine (dT) kinase were less strongly bound to calcium phosphate and were not precipitated at pH 5; these enzymes also had much lower molecular weights than the T antigen, as did dihydrofolic (FH2) reductase. Furthermore, higher ammonium sulfate concentrations were required to precipitate dCMP deaminase, dTMP kinase, and FH2 reductase activities than to precipitate the T antigen. Another difference was that the T antigen was not stabilized, but dCMP deaminase, dTMP kinase, and dT kinase, were stabilized, respectively, by dCTP, dTMP, and dT or dTTP. Deoxyribonucleic acid (DNA) polymerase activity resembled the T antigen in adsorption to calcium phosphate, in precipitation by ammonium sulfate or at pH 5, and in the rate of inactivation when incubated at 38 C. However, the polymerase activity could be partly separated from the T antigen by Sephadex G-200 gel chromatography. The cell fraction containing partially purified T antigen also contained a soluble complement-fixing antigen (presumably a subunit of the viral capsid) which reacted with hyperimmune monkey sera. The latter antigen was present in very low titers or absent from cell extracts prepared from SV40-infected monkey kidney cell cultures which had been treated with ara-C at the time of infection, or from SV40-transformed mouse kidney (mKS) or hamster tumor (H-50) cells. The T antigen, however, was present in usual amounts in SV40-transformed cells or ara-C treated, infected cells. PMID:4316227

Cytotoxic T lymphocyte recognition of HLA-A/B antigens introduced into EL4 cells by cell-liposome fusion.

PubMed

Engelhard, V H; Powers, G A; Moore, L C; Holterman, M J; Correa-Freire, M C

1984-01-01

HLA-A2 and -B7 antigens were introduced into EL4 (H-2b) cells by cell-liposome fusion and were used as targets or stimulators for cytotoxic T lymphocytes (CTL) generated in C57B1/6 (H-2b) mice. It was found that such EL4-HLA cells were not recognized by CTL that had been raised against either a human cell line bearing these HLA antigens or the purified HLA-A2 and -B7 antigens reconstituted into liposomes. In addition, EL4-HLA cells were not capable of inducing CTL that could recognize a human cell line bearing HLA-A2 and -B7 antigens. Instead, EL4-HLA cells induced CTL that specifically lysed EL4-HLA cells and not human cells expressing HLA-A2 and -B7. CTL recognition required the presence of HLA antigens on the EL4 cell surface and was inhibited by antibodies against either H-2b or HLA-A/B. Monoclonal antibody binding studies showed that the expected polymorphic determinants of the HLA-A2 and -B7 antigens were still present on EL4-HLA cells. However, the specificity of CTL or their precursors that are capable of recognizing HLA-A2 or -B7 was altered after these antigens became associated with the EL4 surface. Possible explanations for these results are discussed.

Peptide-dependent Conformational Fluctuation Determines the Stability of the Human Leukocyte Antigen Class I Complex*

PubMed Central

Yanaka, Saeko; Ueno, Takamasa; Shi, Yi; Qi, Jianxun; Gao, George F.; Tsumoto, Kouhei; Sugase, Kenji

2014-01-01

In immune-mediated control of pathogens, human leukocyte antigen (HLA) class I presents various antigenic peptides to CD8+ T-cells. Long-lived peptide presentation is important for efficient antigen-specific T-cell activation. Presentation time depends on the peptide sequence and the stability of the peptide-HLA complex (pHLA). However, the determinant of peptide-dependent pHLA stability remains elusive. Here, to reveal the pHLA stabilization mechanism, we examined the crystal structures of an HLA class I allomorph in complex with HIV-derived peptides and evaluated site-specific conformational fluctuations using NMR. Although the crystal structures of various pHLAs were almost identical independent of the peptides, fluctuation analyses identified a peptide-dependent minor state that would be more tightly packed toward the peptide. The minor population correlated well with the thermostability and cell surface presentation of pHLA, indicating that this newly identified minor state is important for stabilizing the pHLA and facilitating T-cell recognition. PMID:25028510

From The Cover: Poly- amino ester-containing microparticles enhance the activity of nonviral genetic vaccines

NASA Astrophysics Data System (ADS)

Little, Steven R.; Lynn, David M.; Ge, Qing; Anderson, Daniel G.; Puram, Sidharth V.; Chen, Jianzhu; Eisen, Herman N.; Langer, Robert

2004-06-01

Current nonviral genetic vaccine systems are less effective than viral vaccines, particularly in cancer systems where epitopes can be weakly immunogenic and antigen-presenting cell processing and presentation to T cells is down-regulated. A promising nonviral delivery method for genetic vaccines involves microencapsulation of antigen-encoding DNA, because such particles protect plasmid payloads and target them to phagocytic antigen-presenting cells. However, conventional microparticle formulations composed of poly lactic-co-glycolic acid take too long to release encapsulated payload and fail to induce high levels of target gene expression. Here, we describe a microparticle-based DNA delivery system composed of a degradable, pH-sensitive poly- amino ester and poly lactic-co-glycolic acid. These formulations generate an increase of 3-5 orders of magnitude in transfection efficiency and are potent activators of dendritic cells in vitro. When used as vaccines in vivo, these microparticle formulations, unlike conventional formulations, induce antigen-specific rejection of transplanted syngenic tumor cells.

Rational design of nanoparticles towards targeting antigen-presenting cells and improved T cell priming.

PubMed

Zupančič, Eva; Curato, Caterina; Paisana, Maria; Rodrigues, Catarina; Porat, Ziv; Viana, Ana S; Afonso, Carlos A M; Pinto, João; Gaspar, Rogério; Moreira, João N; Satchi-Fainaro, Ronit; Jung, Steffen; Florindo, Helena F

2017-07-28

Vaccination is a promising strategy to trigger and boost immune responses against cancer or infectious disease. We have designed, synthesized and characterized aliphatic-polyester (poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NP) to investigate how the nature of protein association (adsorbed versus entrapped) and polymer/surfactant concentrations impact on the generation and modulation of antigen-specific immune responses. The ability of the NP formulations to target dendritic cells (DC), be internalized and activate the T cells was characterized and optimized in vitro and in vivo using markers of DC activation and co-stimulatory molecules. Ovalbumin (OVA) was used as a model antigen in combination with the engraftment of CD4 + and CD8 + T cells, carrying a transgenic OVA-responding T cell receptor (TCR), to trace and characterize the activation of antigen-specific CD4 + and CD8 + lymph node T cells upon NP vaccination. Accordingly, the phenotype and frequency of immune cell stimulation induced by the NP loaded with OVA, isolated or in combination with synthetic unmethylated cytosine-phosphate-guanine (CpG) oligodeoxynucleotide (ODN) motifs, were characterized. DC-NP interactions increased with incubation time, presenting internalization values between 50 and 60% and 30-40%, in vitro and in vivo, respectively. Interestingly, animal immunization with antigen-adsorbed NP up-regulated major histocompatibility complex (MHC) class II (MHCII), while NP entrapping the antigen up-regulated MHCI, suggesting a more efficient cross-presentation. On the other hand, rather surprisingly, the surfactant used in the NP formulation had a major impact on the activation of antigen presenting cells (APC). In fact, DC collected from lymph nodes of animals immunized with NP prepared using poly(vinil alcohol) (PVA), as a surfactant, expressed significantly higher levels of CD86, MHCI and MHCII. In addition, those NP prepared with PVA and co-entrapping OVA and the toll-like receptor (TLR) ligand CpG, induced the most profound antigen-specific T cell response, by both CD4 + and CD8 + T cells, in vivo. Overall, our data reveal the impact of NP composition and surface properties on the type and extension of induced immune responses. Deeper understanding on the NP-immune cell crosstalk can guide the rational development of nano-immunotherapeutic systems with improved and specific therapeutic efficacy and avoiding off-target effects. Copyright © 2017 Elsevier B.V. All rights reserved.

Loss of Proliferation and Antigen Presentation Activity following Internalization of Polydispersed Carbon Nanotubes by Primary Lung Epithelial Cells

PubMed Central

Kumari, Mandavi; Sachar, Sumedha; Saxena, Rajiv K.

2012-01-01

Interactions between poly-dispersed acid functionalized single walled carbon nanotubes (AF-SWCNTs) and primary lung epithelial (PLE) cells were studied. Peritoneal macrophages (PMs, known phagocytic cells) were used as positive controls in this study. Recovery of live cells from cultures of PLE cells and PMs was significantly reduced in the presence of AF-SWCNTs, in a time and dose dependent manner. Both PLE cells as well as PMs could take up fluorescence tagged AF-SWCNTs in a time dependent manner and this uptake was significantly blocked by cytochalasin D, an agent that blocks the activity of acto-myosin fibers and therefore the phagocytic activity of cells. Confocal microscopic studies confirmed that AF-SWCNTs were internalized by both PLE cells and PMs. Intra-trachially instilled AF-SWCNTs could also be taken up by lung epithelial cells as well as alveolar macrophages. Freshly isolated PLE cells had significant cell division activity and cell cycling studies indicated that treatment with AF-SWCNTs resulted in a marked reduction in S-phase of the cell cycle. In a previously standardized system to study BCG antigen presentation by PLE cells and PMs to sensitized T helper cells, AF-SWCNTs could significantly lower the antigen presentation ability of both cell types. These results show that mouse primary lung epithelial cells can efficiently internalize AF-SWCNTs and the uptake of nanotubes interfered with biological functions of PLE cells including their ability to present BCG antigens to sensitized T helper cells. PMID:22384094

Synthetic Nanovaccines Against Respiratory Pathogens (SYNARP). Addendum

DTIC Science & Technology

2014-09-01

and c) block ionomer complexes (BIC) for targeted delivery of DNA (or protein) antigen to the antigen presenting cells (APCs) (Platform C). The...immune cells to elicit most efficient immune response. The proposal was focusing on achieving the following specific technical objectives: 1) Develop...muscle in a mouse (Platform B & C). ACCOMPLISHED YEAR 3 Task 1: Determine optimal antigen-containing BPN that activate dendritic cells (DCs

Exosome-based tumor antigens-adjuvant co-delivery utilizing genetically engineered tumor cell-derived exosomes with immunostimulatory CpG DNA.

PubMed

Morishita, Masaki; Takahashi, Yuki; Matsumoto, Akihiro; Nishikawa, Makiya; Takakura, Yoshinobu

2016-12-01

For cancer immunotherapy via tumor antigen vaccination in combination with an adjuvant, major challenges include the identification of a particular tumor antigen and efficient delivery of the antigen as well as adjuvant to antigen-presenting cells. In this study, we proposed an efficient exosome-based tumor antigens-adjuvant co-delivery system using genetically engineered tumor cell-derived exosomes containing endogenous tumor antigens and immunostimulatory CpG DNA. Murine melanoma B16BL6 cells were transfected with a plasmid vector encoding a fusion streptavidin (SAV; a protein that binds to biotin with high affinity)-lactadherin (LA; an exosome-tropic protein) protein, yielding genetically engineered SAV-LA-expressing exosomes (SAV-exo). SAV-exo were combined with biotinylated CpG DNA to prepare CpG DNA-modified exosomes (CpG-SAV-exo). Fluorescent microscopic observation revealed the successful modification of exosomes with CpG DNA by SAV-biotin interaction. CpG-SAV-exo showed efficient and simultaneous delivery of exosomes with CpG DNA to murine dendritic DC2.4 cells in culture. Treatment with CpG-SAV-exo effectively activated DC2.4 cells and enhanced tumor antigen presentation capacity. Immunization with CpG-SAV-exo exhibited stronger in vivo antitumor effects in B16BL6 tumor-bearing mice than simple co-administration of exosomes and CpG DNA. Thus, genetically engineered CpG-SAV-exo is an effective exosome-based tumor antigens-adjuvant co-delivery system that will be useful for cancer immunotherapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Precision cancer immunotherapy: optimizing dendritic cell-based strategies to induce tumor antigen-specific T-cell responses against individual patient tumors.

PubMed

Osada, Takuya; Nagaoka, Koji; Takahara, Masashi; Yang, Xiao Yi; Liu, Cong-Xiao; Guo, Hongtao; Roy Choudhury, Kingshuk; Hobeika, Amy; Hartman, Zachary; Morse, Michael A; Lyerly, H Kim

2015-05-01

Most dendritic cell (DC)-based vaccines have loaded the DC with defined antigens, but loading with autologos tumor-derived antigens would generate DCs that activate personalized tumor-specific T-cell responses. We hypothesized that DC matured with an optimized combination of reagents and loaded with tumor-derived antigens using a clinically feasible electroporation strategy would induce potent antitumor immunity. We first studied the effects on DC maturation and antigen presentation of the addition of picibanil (OK432) to a combination of zoledronic acid, tumor necrosis factor-α, and prostaglandin E2. Using DC matured with the optimized combination, we tested 2 clinically feasible sources of autologous antigen for electroloading, total tumor mRNA or total tumor lysate, to determine which stimulated more potent antigen-specific T cells in vitro and activated more potent antitumor immunity in vivo. The combination of tumor necrosis factor-α/prostaglandin E2/zoledronic acid/OK432 generated DC with high expression of maturation markers and antigen-specific T-cell stimulatory function in vitro. Mature DC electroloaded with tumor-derived mRNA [mRNA electroporated dendritic cell (EPDC)] induced greater expansion of antigen-specific T cells in vitro than DC electroloaded with tumor lysate (lysate EPDC). In a therapeutic model of MC38-carcinoembryonic antigen colon cancer-bearing mice, vaccination with mRNA EPDC induced the most efficient anti-carcinoembryonic antigen cellular immune response, which significantly suppressed tumor growth. In conclusion, mature DC electroloaded with tumor-derived mRNA are a potent cancer vaccine, especially useful when specific tumor antigens for vaccination have not been identified, allowing autologous tumor, and if unavailable, allogeneic cell lines to be used as an unbiased source of antigen. Our data support clinical testing of this strategy.

Skin Dendritic Cell Targeting via Microneedle Arrays Laden with Antigen-Encapsulated Poly-d,l-lactide-co-Glycolide Nanoparticles Induces Efficient Antitumor and Antiviral Immune Responses

PubMed Central

2013-01-01

The efficacious delivery of antigens to antigen-presenting cells (APCs), in particular, to dendritic cells (DCs), and their subsequent activation remains a significant challenge in the development of effective vaccines. This study highlights the potential of dissolving microneedle (MN) arrays laden with nanoencapsulated antigen to increase vaccine immunogenicity by targeting antigen specifically to contiguous DC networks within the skin. Following in situ uptake, skin-resident DCs were able to deliver antigen-encapsulated poly-d,l-lactide-co-glycolide (PGLA) nanoparticles to cutaneous draining lymph nodes where they subsequently induced significant expansion of antigen-specific T cells. Moreover, we show that antigen-encapsulated nanoparticle vaccination via microneedles generated robust antigen-specific cellular immune responses in mice. This approach provided complete protection in vivo against both the development of antigen-expressing B16 melanoma tumors and a murine model of para-influenza, through the activation of antigen-specific cytotoxic CD8+ T cells that resulted in efficient clearance of tumors and virus, respectively. In addition, we show promising findings that nanoencapsulation facilitates antigen retention into skin layers and provides antigen stability in microneedles. Therefore, the use of biodegradable polymeric nanoparticles for selective targeting of antigen to skin DC subsets through dissolvable MNs provides a promising technology for improved vaccination efficacy, compliance, and coverage. PMID:23373658

Co-stimulatory function in primary germinal center responses: CD40 and B7 are required on distinct antigen-presenting cells.

PubMed

Watanabe, Masashi; Fujihara, Chiharu; Radtke, Andrea J; Chiang, Y Jeffrey; Bhatia, Sumeena; Germain, Ronald N; Hodes, Richard J

2017-09-04

T cell-dependent germinal center (GC) responses require coordinated interactions of T cells with two antigen-presenting cell (APC) populations, B cells and dendritic cells (DCs), in the presence of B7- and CD40-dependent co-stimulatory pathways. Contrary to the prevailing paradigm, we found unique cellular requirements for B7 and CD40 expression in primary GC responses to vaccine immunization with protein antigen and adjuvant: B7 was required on DCs but was not required on B cells, whereas CD40 was required on B cells but not on DCs in the generation of antigen-specific follicular helper T cells, antigen-specific GC B cells, and high-affinity class-switched antibody production. There was, in fact, no requirement for coexpression of B7 and CD40 on the same cell in these responses. Our findings support a substantially revised model for co-stimulatory function in the primary GC response, with crucial and distinct contributions of B7- and CD40-dependent pathways expressed by different APC populations and with important implications for understanding how to optimize vaccine responses or limit autoimmunity. This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.

The Role of Dendritic Cell Maturation in the Induction of Insulin-Dependent Diabetes Mellitus.

PubMed

Mbongue, Jacques C; Nieves, Hector A; Torrez, Timothy W; Langridge, William H R

2017-01-01

Dendritic cells (DCs) are the dominant class of antigen-presenting cells in humans and are largely responsible for the initiation and guidance of innate and adaptive immune responses involved in maintenance of immunological homeostasis. Immature dendritic cells (iDCs) phagocytize pathogens and toxic proteins and in endosomal vesicles degrade them into small fragments for presentation on major histocompatibility complex (MHC) II receptor molecules to naïve cognate T cells (Th0). In addition to their role in stimulation of immunity, DCs are involved in the induction and maintenance of immune tolerance toward self-antigens. During activation, the iDCs become mature. Maturation begins when the DCs cease taking up antigens and begin to migrate from their location in peripheral tissues to adjacent lymph nodes or the spleen where during their continued maturation the DCs present stored antigens on surface MHCII receptor molecules to naive Th0 cells. During antigen presentation, the DCs upregulate the biosynthesis of costimulatory receptor molecules CD86, CD80, CD83, and CD40 on their plasma membrane. These activated DC receptor molecules bind cognate CD28 receptors presented on the Th0 cell membrane, which triggers DC secretion of IL-12 or IL-10 cytokines resulting in T cell differentiation into pro- or anti-inflammatory T cell subsets. Although basic concepts involved in the process of iDC activation and guidance of Th0 cell differentiation have been previously documented, they are poorly defined. In this review, we detail what is known about the process of DC maturation and its role in the induction of insulin-dependent diabetes mellitus autoimmunity.

Assembly and Immunological Processing of Polyelectrolyte Multilayers Composed of Antigens and Adjuvants.

PubMed

Chiu, Yu-Chieh; Gammon, Joshua M; Andorko, James I; Tostanoski, Lisa H; Jewell, Christopher M

2016-07-27

While biomaterials provide a platform to control the delivery of vaccines, the recently discovered intrinsic inflammatory characteristics of many polymeric carriers can also complicate rational design because the carrier itself can alter the response to other vaccine components. To address this challenge, we recently developed immune-polyelectrolyte multilayer (iPEMs) capsules electrostatically assembled entirely from peptide antigen and molecular adjuvants. Here, we use iPEMs built from SIINFEKL model antigen and polyIC, a stimulatory toll-like receptor agonist, to investigate the impact of pH on iPEM assembly, the processing and interactions of each iPEM component with primary immune cells, and the role of these interactions during antigen-specific T cell responses in coculture and mice. We discovered that iPEM assembly is pH dependent with respect to both the antigen and adjuvant component. Controlling the pH also allows tuning of the relative loading of SIINFEKL and polyIC in iPEM capsules. During in vitro studies with primary dendritic cells (DCs), iPEM capsules ensure that greater than 95% of cells containing at least one signal (i.e., antigen, adjuvant) also contained the other signal. This codelivery leads to DC maturation and SIINFEKL presentation via the MHC-I antigen presentation pathway, resulting in antigen-specific T cell proliferation and pro-inflammatory cytokine secretion. In mice, iPEM capsules potently expand antigen-specific T cells compared with equivalent admixed formulations. Of note, these enhancements become more pronounced with successive booster injections, suggesting that iPEMs functionally improve memory recall response. Together our results reveal some of the features that can be tuned to modulate the properties of iPEM capsules, and how these modular vaccine structures can be used to enhance interactions with immune cells in vitro and in mice.

Heat-shock proteins as dendritic cell-targeting vaccines – getting warmer

PubMed Central

McNulty, Shaun; Colaco, Camilo A; Blandford, Lucy E; Bailey, Christopher R; Baschieri, Selene; Todryk, Stephen

2013-01-01

Heat-shock proteins (hsp) provide a natural link between innate and adaptive immune responses by combining the ideal properties of antigen carriage (chaperoning), targeting and activation of antigen-presenting cells (APC), including dendritic cells (DC). Targeting is achieved through binding of hsp to distinct cell surface receptors and is followed by antigen internalization, processing and presentation. An improved understanding of the interaction of hsp with DC has driven the development of numerous hsp-containing vaccines, designed to deliver antigens directly to DC. Studies in mice have shown that for cancers, such vaccines generate impressive immune responses and protection from tumour challenge. However, translation to human use, as for many experimental immunotherapies, has been slow partly because of the need to perform trials in patients with advanced cancers, where demonstration of efficacy is challenging. Recently, the properties of hsp have been used for development of prophylactic vaccines against infectious diseases including tuberculosis and meningitis. These hsp-based vaccines, in the form of pathogen-derived hsp–antigen complexes, or recombinant hsp combined with selected antigens in vitro, offer an innovative approach against challenging diseases where broad antigen coverage is critical. PMID:23551234

ESAT-6 Targeting to DEC205+ Antigen Presenting Cells Induces Specific-T Cell Responses against ESAT-6 and Reduces Pulmonary Infection with Virulent Mycobacterium tuberculosis.

PubMed

Silva-Sánchez, Aarón; Meza-Pérez, Selene; Flores-Langarica, Adriana; Donis-Maturano, Luis; Estrada-García, Iris; Calderón-Amador, Juana; Hernández-Pando, Rogelio; Idoyaga, Juliana; Steinman, Ralph M; Flores-Romo, Leopoldo

2015-01-01

Airways infection with Mycobacterium tuberculosis (Mtb) is contained mostly by T cell responses, however, Mtb has developed evasion mechanisms which affect antigen presenting cell (APC) maturation/recruitment delaying the onset of Ag-specific T cell responses. Hypothetically, bypassing the natural infection routes by delivering antigens directly to APCs may overcome the pathogen's naturally evolved evasion mechanisms, thus facilitating the induction of protective immune responses. We generated a murine monoclonal fusion antibody (α-DEC-ESAT) to deliver Early Secretory Antigen Target (ESAT)-6 directly to DEC205+ APCs and to assess its in vivo effects on protection associated responses (IFN-γ production, in vivo CTL killing, and pulmonary mycobacterial load). Treatment with α-DEC-ESAT alone induced ESAT-6-specific IFN-γ producing CD4+ T cells and prime-boost immunization prior to Mtb infection resulted in early influx (d14 post-infection) and increased IFN-γ+ production by specific T cells in the lungs, compared to scarce IFN-γ production in control mice. In vivo CTL killing was quantified in relevant tissues upon transferring target cells loaded with mycobacterial antigens. During infection, α-DEC-ESAT-treated mice showed increased target cell killing in the lungs, where histology revealed cellular infiltrate and considerably reduced bacterial burden. Targeting the mycobacterial antigen ESAT-6 to DEC205+ APCs before infection expands specific T cell clones responsible for early T cell responses (IFN-γ production and CTL activity) and substantially reduces lung bacterial burden. Delivering mycobacterial antigens directly to APCs provides a unique approach to study in vivo the role of APCs and specific T cell responses to assess their potential anti-mycobacterial functions.

ESAT-6 Targeting to DEC205+ Antigen Presenting Cells Induces Specific-T Cell Responses against ESAT-6 and Reduces Pulmonary Infection with Virulent Mycobacterium tuberculosis

PubMed Central

Silva-Sánchez, Aarón; Meza-Pérez, Selene; Flores-Langarica, Adriana; Donis-Maturano, Luis; Estrada-García, Iris; Calderón-Amador, Juana; Hernández-Pando, Rogelio; Idoyaga, Juliana; Flores-Romo, Leopoldo

2015-01-01

Airways infection with Mycobacterium tuberculosis (Mtb) is contained mostly by T cell responses, however, Mtb has developed evasion mechanisms which affect antigen presenting cell (APC) maturation/recruitment delaying the onset of Ag-specific T cell responses. Hypothetically, bypassing the natural infection routes by delivering antigens directly to APCs may overcome the pathogen’s naturally evolved evasion mechanisms, thus facilitating the induction of protective immune responses. We generated a murine monoclonal fusion antibody (α-DEC-ESAT) to deliver Early Secretory Antigen Target (ESAT)-6 directly to DEC205+ APCs and to assess its in vivo effects on protection associated responses (IFN-γ production, in vivo CTL killing, and pulmonary mycobacterial load). Treatment with α-DEC-ESAT alone induced ESAT-6-specific IFN-γ producing CD4+ T cells and prime-boost immunization prior to Mtb infection resulted in early influx (d14 post-infection) and increased IFN-γ+ production by specific T cells in the lungs, compared to scarce IFN-γ production in control mice. In vivo CTL killing was quantified in relevant tissues upon transferring target cells loaded with mycobacterial antigens. During infection, α-DEC-ESAT-treated mice showed increased target cell killing in the lungs, where histology revealed cellular infiltrate and considerably reduced bacterial burden. Targeting the mycobacterial antigen ESAT-6 to DEC205+ APCs before infection expands specific T cell clones responsible for early T cell responses (IFN-γ production and CTL activity) and substantially reduces lung bacterial burden. Delivering mycobacterial antigens directly to APCs provides a unique approach to study in vivo the role of APCs and specific T cell responses to assess their potential anti-mycobacterial functions. PMID:25915045

Identification of Barramundi (Lates calcarifer) DC-SCRIPT, a Specific Molecular Marker for Dendritic Cells in Fish

PubMed Central

Zoccola, Emmanuelle; Delamare-Deboutteville, Jérôme; Barnes, Andrew C.

2015-01-01

Antigen presentation is a critical step bridging innate immune recognition and specific immune memory. In mammals, the process is orchestrated by dendritic cells (DCs) in the lymphatic system, which initiate clonal proliferation of antigen-specific lymphocytes. However, fish lack a classical lymphatic system and there are currently no cellular markers for DCs in fish, thus antigen-presentation in fish is poorly understood. Recently, antigen-presenting cells similar in structure and function to mammalian DCs were identified in various fish, including rainbow trout (Oncorhynchus mykiss) and zebrafish (Danio rerio). The present study aimed to identify a potential molecular marker for DCs in fish and therefore targeted DC-SCRIPT, a well-conserved zinc finger protein that is preferentially expressed in all sub-types of human DCs. Putative dendritic cells were obtained in culture by maturation of spleen and pronephros-derived monocytes. DC-SCRIPT was identified in barramundi by homology using RACE PCR and genome walking. Specific expression of DC-SCRIPT was detected in barramundi cells by Stellaris mRNA FISH, in combination with MHCII expression when exposed to bacterial derived peptidoglycan, suggesting the presence of DCs in L. calcarifer. Moreover, morphological identification was achieved by light microscopy of cytospins prepared from these cultures. The cultured cells were morphologically similar to mammalian and trout DCs. Migration assays determined that these cells have the ability to move towards pathogens and pathogen associated molecular patterns, with a preference for peptidoglycans over lipopolysaccharides. The cells were also strongly phagocytic, engulfing bacteria and rapidly breaking them down. Barramundi DCs induced significant proliferation of responder populations of T-lymphocytes, supporting their role as antigen presenting cells. DC-SCRIPT expression in head kidney was higher 6 and 24 h following intraperitoneal challenge with peptidoglycan and lipopolysaccharide and declined after 3 days relative to PBS-injected controls. Relative expression was also lower in the spleen at 3 days post challenge but increased again at 7 days. As DC-SCRIPT is a constitutively expressed nuclear receptor, independent of immune activation, this may indicate initial migration of immature DCs from head kidney and spleen to the injection site, followed by return to the spleen for maturation and antigen presentation. DC-SCRIPT may be a valuable tool in the investigation of antigen presentation in fish and facilitate optimisation of vaccines and adjuvants for aquaculture. PMID:26173015

A STING-activating nanovaccine for cancer immunotherapy

NASA Astrophysics Data System (ADS)

Luo, Min; Wang, Hua; Wang, Zhaohui; Cai, Haocheng; Lu, Zhigang; Li, Yang; Du, Mingjian; Huang, Gang; Wang, Chensu; Chen, Xiang; Porembka, Matthew R.; Lea, Jayanthi; Frankel, Arthur E.; Fu, Yang-Xin; Chen, Zhijian J.; Gao, Jinming

2017-07-01

The generation of tumour-specific T cells is critically important for cancer immunotherapy. A major challenge in achieving a robust T-cell response is the spatiotemporal orchestration of antigen cross-presentation in antigen-presenting cells with innate stimulation. Here, we report a minimalist nanovaccine, comprising a simple physical mixture of an antigen and a synthetic polymeric nanoparticle, PC7A NP, which generates a strong cytotoxic T-cell response with low systemic cytokine expression. Mechanistically, the PC7A NP achieves efficient cytosolic delivery of tumour antigens to antigen-presenting cells in draining lymph nodes, leading to increased surface presentation while simultaneously activating type I interferon-stimulated genes. This effect is dependent on stimulator of interferon genes (STING), but not the Toll-like receptor or the mitochondrial antiviral-signalling protein (MAVS) pathway. The nanovaccine led to potent tumour growth inhibition in melanoma, colon cancer and human papilloma virus-E6/E7 tumour models. The combination of the PC7A nanovaccine and an anti-PD-1 antibody showed great synergy, with 100% survival over 60 days in a TC-1 tumour model. Rechallenging of these tumour-free animals with TC-1 cells led to complete inhibition of tumour growth, suggesting the generation of long-term antitumour memory. The STING-activating nanovaccine offers a simple, safe and robust strategy in boosting anti-tumour immunity for cancer immunotherapy.

Cell-permeable nanobodies for targeted immunolabelling and antigen manipulation in living cells

NASA Astrophysics Data System (ADS)

Herce, Henry D.; Schumacher, Dominik; Schneider, Anselm F. L.; Ludwig, Anne K.; Mann, Florian A.; Fillies, Marion; Kasper, Marc-André; Reinke, Stefan; Krause, Eberhard; Leonhardt, Heinrich; Cardoso, M. Cristina; Hackenberger, Christian P. R.

2017-08-01

Functional antibody delivery in living cells would enable the labelling and manipulation of intracellular antigens, which constitutes a long-thought goal in cell biology and medicine. Here we present a modular strategy to create functional cell-permeable nanobodies capable of targeted labelling and manipulation of intracellular antigens in living cells. The cell-permeable nanobodies are formed by the site-specific attachment of intracellularly stable (or cleavable) cyclic arginine-rich cell-penetrating peptides to camelid-derived single-chain VHH antibody fragments. We used this strategy for the non-endocytic delivery of two recombinant nanobodies into living cells, which enabled the relocalization of the polymerase clamp PCNA (proliferating cell nuclear antigen) and tumour suppressor p53 to the nucleolus, and thereby allowed the detection of protein-protein interactions that involve these two proteins in living cells. Furthermore, cell-permeable nanobodies permitted the co-transport of therapeutically relevant proteins, such as Mecp2, into the cells. This technology constitutes a major step in the labelling, delivery and targeted manipulation of intracellular antigens. Ultimately, this approach opens the door towards immunostaining in living cells and the expansion of immunotherapies to intracellular antigen targets.

T cell ignorance is bliss: T cells are not tolerized by Langerhans cells presenting human papillomavirus antigens in the absence of costimulation.

PubMed

Woodham, Andrew W; Yan, Lisa; Skeate, Joseph G; van der Veen, Daniel; Brand, Heike H; Wong, Michael K; Da Silva, Diane M; Kast, W Martin

2016-12-01

Human papillomavirus type 16 (HPV16) infections are intra-epithelial, and thus, HPV16 is known to interact with Langerhans cells (LCs), the resident epithelial antigen-presenting cells (APCs). The current paradigm for APC-mediated induction of T cell anergy is through delivery of T cell receptor signals via peptides on MHC molecules (signal 1), but without costimulation (signal 2). We previously demonstrated that LCs exposed to HPV16 in vitro present HPV antigens to T cells without costimulation, but it remained uncertain if such T cells would remain ignorant, become anergic, or in the case of CD4+ T cells, differentiate into Tregs. Here we demonstrate that Tregs were not induced by LCs presenting only signal 1, and through a series of in vitro immunizations show that CD8 + T cells receiving signal 1 + 2 from LCs weeks after consistently receiving signal 1 are capable of robust effector functions. Importantly, this indicates that T cells are not tolerized but instead remain ignorant to HPV, and are activated given the proper signals.

Efficient induction of CD25- iTreg by co-immunization requires strongly antigenic epitopes for T cells.

PubMed

Geng, Shuang; Yu, Yang; Kang, Youmin; Pavlakis, George; Jin, Huali; Li, Jinyao; Hu, Yanxin; Hu, Weibin; Wang, Shuang; Wang, Bin

2011-05-05

We previously showed that co-immunization with a protein antigen and a DNA vaccine coding for the same antigen induces CD40 low IL-10 high tolerogenic DCs, which in turn stimulates the expansion of antigen-specific CD4+CD25-Foxp3+ regulatory T cells (CD25- iTreg). However, it was unclear how to choose the antigen sequence to maximize tolerogenic antigen presentation and, consequently, CD25- iTreg induction. In the present study, we demonstrated the requirement of highly antigenic epitopes for CD25- iTreg induction. Firstly, we showed that the induction of CD25- iTreg by tolerogenic DC can be blocked by anti-MHC-II antibody. Next, both the number and the suppressive activity of CD25- iTreg correlated positively with the overt antigenicity of an epitope to activate T cells. Finally, in a mouse model of dermatitis, highly antigenic epitopes derived from a flea allergen not only induced more CD25- iTreg, but also more effectively prevented allergenic reaction to the allergen than did weakly antigenic epitopes. Our data thus indicate that efficient induction of CD25- iTreg requires highly antigenic peptide epitopes. This finding suggests that highly antigenic epitopes should be used for efficient induction of CD25- iTreg for clinical applications such as flea allergic dermatitis.

A role for mitochondria in antigen processing and presentation

PubMed Central

Bonifaz, Laura C; Cervantes-Silva, Mariana P; Ontiveros-Dotor, Elizabeth; López-Villegas, Edgar O; Sánchez-García, F Javier

2015-01-01

Immune synapse formation is critical for T-lymphocyte activation, and mitochondria have a role in this process, by localizing close to the immune synapse, regulating intracellular calcium concentration, and providing locally required ATP. The interaction between antigen-presenting cells (APCs) and T lymphocytes is a two-way signalling process. However, the role of mitochondria in APCs during this process remains unknown. For APCs to be able to activate T lymphocytes, they must first engage in an antigen-uptake, -processing and -presentation process. Here we show that hen egg white lysozyme (HEL) -loaded B lymphocytes, as a type of APC, undergo a small but significant mitochondrial depolarization by 1–2 hr following antigen exposure, suggesting an increase in their metabolic demands. Inhibition of ATP synthase (oligomycin) or mitochondrial Ca2+ uniporter (MCU) (Ruthenium red) had no effect on antigen uptake. Therefore, antigen processing and antigen presentation were further analysed. Oligomycin treatment reduced the amount of specific MHC–peptide complexes but not total MHC II on the cell membrane of B lymphocytes, which correlated with a decrease in antigen presentation. However, oligomycin also reduced antigen presentation by B lymphocytes, which endogenously express HEL and by B lymphocytes loaded with the HEL48–62 peptide, although to a lesser extent. ATP synthase inhibition and MCU inhibition had a clear inhibitory effect on antigen processing (DQ-OVA). Taken together these results suggest that ATP synthase and MCU are relevant for antigen processing and presentation. Finally, APC mitochondria were found to re-organize towards the APC–T immune synapse. PMID:25251370

Mannan-decorated thiolated Eudragit microspheres for targeting antigen presenting cells via nasal vaccination.

PubMed

Li, Hui-Shan; Singh, Bijay; Park, Tae-Eun; Hong, Zhong-Shan; Kang, Sang-Kee; Cho, Chong-Su; Choi, Yun-Jaie

2015-12-01

Mucosal vaccination of protein as an antigen requires appropriate delivery or adjuvant systems to deliver antigen to mucosal immune cells efficiently and generate valid immune responses. For successful nasal immunization, the obstacles imposed by the normal process of mucociliary clearance which limits residence time of applied antigens and low antigen delivery to antigen presenting cells (APCs) in nasal associated lymphoid tissue (NALT) need to be overcome for the efficient vaccination. Here, we prepared mucoadhesive and mannan-decorated thiolated Eudragit microspheres (Man-TEM) as a nasal vaccine carrier to overcome the limitations. Mucoadhesive thiolated Eudragit (TE) were decorated with mannan for targeting mannose receptors (MR) in antigen presenting cells (APCs) to obtain efficient immune responses. The potential adjuvant ability of Man-TEM for intranasal immunization was confirmed by in vitro and in vivo experiments. In mechanistic study using APCs in vitro, we obtained that Man-TEM enhanced the receptor-mediated endocytosis by stimulating the MR receptors of APCs. The nasal vaccination of OVA-loaded Man-TEM in mice showed higher levels of serum IgG and mucosal sIgA than the soluble OVA group due to the specific recognition of MR of APCs by the mannan in the Man-TEM. These results suggest that mucoadhesive and Man-TEM may be a promising candidate for nasal vaccine delivery system to elicit systemic and mucosal immunity. Copyright © 2015 Elsevier B.V. All rights reserved.

CD8α+ DC trans-presentation of IL-15 to naïve CD8+ T cells produces antigen inexperienced T cells in the periphery with memory phenotype and function

PubMed Central

Sosinowski, Tomasz; White, Jason T.; Cross, Eric; Haluszczak, Catherine; Marrack, Philippa; Gapin, Laurent; Kedl, Ross M.

2013-01-01

Various populations of memory phenotype CD8+ T cells have been described over the last 15–20 years, all of which possess elevated effector functions relative to naïve phenotype cells. Using a technique for isolating antigen specific cells from unprimed hosts, we recently identified a new subset of cells, specific for nominal antigen, but phenotypically and functionally similar to memory cells arising as a result of homeostatic proliferation (HP). We show here that these “Virtual Memory” cells are independent of previously identified “innate memory” cells, arising as a result of their response to IL-15 trans-presentation by lymphoid tissue-resident CD8α+ DCs in the periphery. The absence of IL-15, CD8+ T cell expression of either CD122 or Eomes, or of CD8a+ DCs all lead to the loss of Virtual Memory cells in the host. Our results show that CD8+ T cell homeostatic expansion is an active process within the non-lymphopenic environment, is mediated by IL-15, and produces antigen inexperienced memory cells which retain the capacity to respond to nominal antigen with memory-like function. Preferential engagement of these “Virtual Memory” T cells into a vaccine response could dramatically enhance the rate by which immune protection develops. PMID:23355737

Cross-presentation of IgG-containing immune complexes

PubMed Central

Baker, Kristi; Rath, Timo; Lencer, Wayne I.; Fiebiger, Edda

2012-01-01

IgG is a molecule that functionally combines facets of both innate and adaptive immunity and therefore bridges both arms of the immune system. On the one hand, IgG is created by adaptive immune cells, but can be generated by B cells independently of T cell help. On the other hand, once secreted, IgG can rapidly deliver antigens into intracellular processing pathways, which enable efficient priming of T cell responses towards epitopes from the cognate antigen initially bound by the IgG. While this process has long been known to participate in CD4+ T cell activation, IgG-mediated delivery of exogenous antigens into a major histocompatibility complex (MHC) class I processing pathway has received less attention. The coordinated engagement of IgG with IgG receptors expressed on the cell-surface (FcγR) and within the endolysosomal system (FcRn) is a highly potent means to deliver antigen into processing pathways that promote cross-presentation of MHC class I and presentation of MHC class II-restricted epitopes within the same dendritic cell. This review focuses on the mechanisms by which IgG-containing immune complexes mediate such cross-presentation and the implications that this understanding has for manipulation of immune-mediated diseases that depend upon or are due to the activities of CD8+ T cells. PMID:22847331

Mycobacterium tuberculosis GroEL2 Modulates Dendritic Cell Responses.

PubMed

Georgieva, Maria; Sia, Jonathan Kevin; Bizzell, Erica; Madan-Lala, Ranjna; Rengarajan, Jyothi

2018-02-01

Mycobacterium tuberculosis successfully subverts the host immune response to promote disease progression. In addition to its known intracellular niche in macrophages, M. tuberculosis interferes with the functions of dendritic cells (DCs), which are the primary antigen-presenting cells of the immune system. We previously showed that M. tuberculosis dampens proinflammatory responses and impairs DC functions through the cell envelope-associated serine protease Hip1. Here we present data showing that M. tuberculosis GroEL2, a substrate of Hip1, modulates DC functions. The full-length GroEL2 protein elicited robust proinflammatory responses from DCs and promoted DC maturation and antigen presentation to T cells. In contrast, the cleaved form of GroEL2, which predominates in M. tuberculosis , was poorly immunostimulatory and was unable to promote DC maturation and antigen presentation. Moreover, DCs exposed to full-length, but not cleaved, GroEL2 induced strong antigen-specific gamma interferon (IFN-γ), interleukin-2 (IL-2), and IL-17A cytokine responses from CD4 + T cells. Moreover, the expression of cleaved GroEL2 in the hip1 mutant restored the robust T cell responses to wild-type levels, suggesting that proteolytic cleavage of GroEL2 allows M. tuberculosis to prevent optimal DC-T cell cross talk during M. tuberculosis infection. Copyright © 2018 American Society for Microbiology.

Anti-Yo antibody uptake and interaction with its intracellular target antigen causes Purkinje cell death in rat cerebellar slice cultures: a possible mechanism for paraneoplastic cerebellar degeneration in humans with gynecological or breast cancers.

PubMed

Greenlee, John E; Clawson, Susan A; Hill, Kenneth E; Wood, Blair; Clardy, Stacey L; Tsunoda, Ikuo; Carlson, Noel G

2015-01-01

Anti-Yo antibodies are immunoglobulin G (IgG) autoantibodies reactive with a 62 kDa Purkinje cell cytoplasmic protein. These antibodies are closely associated with paraneoplastic cerebellar degeneration in the setting of gynecological and breast malignancies. We have previously demonstrated that incubation of rat cerebellar slice cultures with patient sera and cerebrospinal fluid containing anti-Yo antibodies resulted in Purkinje cell death. The present study addressed three fundamental questions regarding the role of anti-Yo antibodies in disease pathogenesis: 1) Whether the Purkinje cell cytotoxicity required binding of anti-Yo antibody to its intraneuronal 62 kDa target antigen; 2) whether Purkinje cell death might be initiated by antibody-dependent cellular cytotoxicity rather than intracellular antibody binding; and 3) whether Purkinje cell death might simply be a more general result of intracellular antibody accumulation, rather than of specific antibody-antigen interaction. In our study, incubation of rat cerebellar slice cultures with anti-Yo IgG resulted in intracellular antibody binding, and cell death. Infiltration of the Purkinje cell layer by cells of macrophage/microglia lineage was not observed until extensive cell death was already present. Adsorption of anti-Yo IgG with its 62 kDa target antigen abolished both antibody accumulation and cytotoxicity. Antibodies to other intracellular Purkinje cell proteins were also taken up by Purkinje cells and accumulated intracellularly; these included calbindin, calmodulin, PCP-2, and patient anti-Purkinje cell antibodies not reactive with the 62 kDa Yo antigen. However, intracellular accumulation of these antibodies did not affect Purkinje cell viability. The present study is the first to demonstrate that anti-Yo antibodies cause Purkinje cell death by binding to the intracellular 62 kDa Yo antigen. Anti-Yo antibody cytotoxicity did not involve other antibodies or factors present in patient serum and was not initiated by brain mononuclear cells. Purkinje cell death was not simply due to intraneuronal antibody accumulation.

Anti-Yo Antibody Uptake and Interaction with Its Intracellular Target Antigen Causes Purkinje Cell Death in Rat Cerebellar Slice Cultures: A Possible Mechanism for Paraneoplastic Cerebellar Degeneration in Humans with Gynecological or Breast Cancers

PubMed Central

Greenlee, John E.; Clawson, Susan A.; Hill, Kenneth E.; Wood, Blair; Clardy, Stacey L.; Tsunoda, Ikuo; Carlson, Noel G.

2015-01-01

Anti-Yo antibodies are immunoglobulin G (IgG) autoantibodies reactive with a 62 kDa Purkinje cell cytoplasmic protein. These antibodies are closely associated with paraneoplastic cerebellar degeneration in the setting of gynecological and breast malignancies. We have previously demonstrated that incubation of rat cerebellar slice cultures with patient sera and cerebrospinal fluid containing anti-Yo antibodies resulted in Purkinje cell death. The present study addressed three fundamental questions regarding the role of anti-Yo antibodies in disease pathogenesis: 1) Whether the Purkinje cell cytotoxicity required binding of anti-Yo antibody to its intraneuronal 62 kDa target antigen; 2) whether Purkinje cell death might be initiated by antibody-dependent cellular cytotoxicity rather than intracellular antibody binding; and 3) whether Purkinje cell death might simply be a more general result of intracellular antibody accumulation, rather than of specific antibody-antigen interaction. In our study, incubation of rat cerebellar slice cultures with anti-Yo IgG resulted in intracellular antibody binding, and cell death. Infiltration of the Purkinje cell layer by cells of macrophage/microglia lineage was not observed until extensive cell death was already present. Adsorption of anti-Yo IgG with its 62 kDa target antigen abolished both antibody accumulation and cytotoxicity. Antibodies to other intracellular Purkinje cell proteins were also taken up by Purkinje cells and accumulated intracellularly; these included calbindin, calmodulin, PCP-2, and patient anti-Purkinje cell antibodies not reactive with the 62 kDa Yo antigen. However, intracellular accumulation of these antibodies did not affect Purkinje cell viability. The present study is the first to demonstrate that anti-Yo antibodies cause Purkinje cell death by binding to the intracellular 62 kDa Yo antigen. Anti-Yo antibody cytotoxicity did not involve other antibodies or factors present in patient serum and was not initiated by brain mononuclear cells. Purkinje cell death was not simply due to intraneuronal antibody accumulation. PMID:25885452

Hsp70 enhances presentation of FMDV antigen to bovine CD4+ T cells in vitro

PubMed Central

McLaughlin, Kerry; Seago, Julian; Robinson, Lucy; Kelly, Charles; Charleston, Bryan

2010-01-01

Foot-and-mouth disease virus (FMDV) is the causative agent of a highly contagious acute vesicular disease affecting cloven-hoofed animals, including cattle, sheep and pigs. The current vaccine induces a rapid humoral response, but the duration of the protective antibody response is variable, possibly associated with a variable specific CD4+ T cell response. We investigated the use of heat shock protein 70 (Hsp70) as a molecular chaperone to target viral antigen to the Major Histocompatibility Complex (MHC) class II pathway of antigen presenting cells and generate enhanced MHC II-restricted CD4+ T cell responses in cattle. Monocytes and CD4+ T cells from FMDV vaccinated cattle were stimulated in vitro with complexes of Hsp70 and FMDV peptide, or peptide alone. Hsp70 was found to consistently improve the presentation of a 25-mer FMDV peptide to CD4+ T cells, as measured by T cell proliferation. Complex formation was required for the enhanced effects and Hsp70 alone did not stimulate proliferation. This study provides further evidence that Hsp70:peptide complexes can enhance antigen-specific CD4+ T cell responses in vitro for an important pathogen of livestock. PMID:20167197

T-Cell Surface Antigens and sCD30 as Biomarkers of the Risk of Rejection in Solid Organ Transplantation.

PubMed

Wieland, Eberhard; Shipkova, Maria

2016-04-01

T-cell activation is a characteristic of organ rejection. T cells, located in the draining lymph nodes of the transplant recipient, are faced with non-self-molecules presented by antigen presenting cells and become activated. Activated T cells are characterized by up-regulated surface antigens, such as costimulatory molecules, adhesion molecules, chemokine receptors, and major histocompatibility complex class II molecules. Surface antigen expression can be followed by flow cytometry using monoclonal antibodies in either cell function assays using donor-specific or nonspecific stimulation of isolated cells or whole blood and without stimulation on circulating lymphocytes. Molecules such as CD30 can be proteolytically cleaved off the surface of activated cells in vivo, and the determination of the soluble protein (sCD30) in serum or plasma is performed by immunoassays. As promising biomarkers for rejection and long-term transplant outcome, CD28 (costimulatory receptor for CD80 and CD86), CD154 (CD40 ligand), and sCD30 (tumor necrosis factor receptor superfamily, member 8) have been identified. Whereas cell function assays are time-consuming laboratory-developed tests which are difficult to standardize, commercial assays are frequently available for soluble proteins. Therefore, more data from clinical trials have been published for sCD30 compared with the surface antigens on activated T cells. This short review summarizes the association between selected surface antigens and immunosuppression, and rejection in solid organ transplantation.

The actin cytoskeleton modulates the activation of iNKT cells by segregating CD1d nanoclusters on antigen-presenting cells

PubMed Central

Torreno-Pina, Juan A.; Manzo, Carlo; Salio, Mariolina; Aichinger, Michael C.; Oddone, Anna; Lakadamyali, Melike; Shepherd, Dawn; Besra, Gurdyal S.; Cerundolo, Vincenzo

2016-01-01

Invariant natural killer T (iNKT) cells recognize endogenous and exogenous lipid antigens presented in the context of CD1d molecules. The ability of iNKT cells to recognize endogenous antigens represents a distinct immune recognition strategy, which underscores the constitutive memory phenotype of iNKT cells and their activation during inflammatory conditions. However, the mechanisms regulating such “tonic” activation of iNKT cells remain unclear. Here, we show that the spatiotemporal distribution of CD1d molecules on the surface of antigen-presenting cells (APCs) modulates activation of iNKT cells. By using superresolution microscopy, we show that CD1d molecules form nanoclusters at the cell surface of APCs, and their size and density are constrained by the actin cytoskeleton. Dual-color single-particle tracking revealed that diffusing CD1d nanoclusters are actively arrested by the actin cytoskeleton, preventing their further coalescence. Formation of larger nanoclusters occurs in the absence of interactions between CD1d cytosolic tail and the actin cytoskeleton and correlates with enhanced iNKT cell activation. Importantly and consistently with iNKT cell activation during inflammatory conditions, exposure of APCs to the Toll-like receptor 7/8 agonist R848 increases nanocluster density and iNKT cell activation. Overall, these results define a previously unidentified mechanism that modulates iNKT cell autoreactivity based on the tight control by the APC cytoskeleton of the sizes and densities of endogenous antigen-loaded CD1d nanoclusters. PMID:26798067

Intersection of autophagy with pathways of antigen presentation.

PubMed

Patterson, Natalie L; Mintern, Justine D

2012-12-01

Traditionally, macroautophagy (autophagy) is viewed as a pathway of cell survival. Autophagy ensures the elimination of damaged or unwanted cytosolic components and provides a source of cellular nutrients during periods of stress. Interestingly, autophagy can also directly intersect with, and impact, other major pathways of cellular function. Here, we will review the contribution of autophagy to pathways of antigen presentation. The autophagy machinery acts to modulate both MHCI and MHCII antigen presentation. As such autophagy is an important participant in pathways that elicit host cell immunity and the elimination of infectious pathogens.

Induction of peripheral T cell tolerance and allo/xenoimmunity.

PubMed

Charpentier, B; Alard, P; Hiesse, C; Lantz, O

1994-03-01

It is theoretically impossible or at least difficult to tolerize animals in xenogeneic situation, particularly in non-concordant species. Both humoral defense and several cellular components (T and non T) are offensive weapons which can be directed at myriad of self-antigens in a normal situation, at allo-antigens in abnormal situations, at xeno-antigens in exceptional situations. In either cases, the fine epitopic definition of allo/xeno antigens seen by the TCR is far from being totally known. From recent studies showing the precise requirement of peptides assembly composition in the MHC class I and II groove it should be theoretically possible to tolerize any group of peptides if they are presented, some other, because not presented, may remain ignored, thus apparently tolerized. It is also know, that transplantation tolerance is not a law of "either nothing or all" but a multiple process depending of both the affinity of the TCR and the nature/compositions of the target. In view of the complex array of factors influencing the pathway of T cell activation, three forms of T cell non responsiveness may be suggested in the context of xenogeneic recognition: physical deletion of potentially reactive T cells occurring predominantly in the thymus, non reactivity of T cells resulting from their failure to be influenced by antigens, and anergy possibly due to inappropriate signals from non professional antigen presenting cells. Future investigations must elucidate the requirements for inducing these events in a purpose of xenogeneic organ transplantation.

Probiotic metabolites from Bacillus coagulans GanedenBC30TM support maturation of antigen-presenting cells in vitro

PubMed Central

Benson, Kathleen F; Redman, Kimberlee A; Carter, Steve G; Keller, David; Farmer, Sean; Endres, John R; Jensen, Gitte S

2012-01-01

AIM: To study the effects of probiotic metabolites on maturation stage of antigen-presenting immune cells. METHODS: Ganeden Bacillus coagulans 30 (GBC30) bacterial cultures in log phase were used to isolate the secreted metabolite (MET) fraction. A second fraction was made to generate a crude cell-wall-enriched fraction, by centrifugation and lysis, followed by washing. A preparation of MET was subjected to size exclusion centrifugation, generating three fractions: < 3 kDa, 3-30 kDa, and 30-200 kDa and activities were tested in comparison to crude MET and cell wall in primary cultures of human peripheral blood mononuclear cell (PBMC) as a source of antigen-presenting mononuclear phagocytes. The maturation status of mononuclear phagocytes was evaluated by staining with monoclonal antibodies towards CD14, CD16, CD80 and CD86 and analyzed by flow cytometry. RESULTS: Treatment of PBMC with MET supported maturation of mononuclear phagocytes toward both macrophage and dendritic cell phenotypes. The biological activity unique to the metabolites included a reduction of CD14+ CD16+ pro-inflammatory cells, and this property was associated with the high molecular weight metabolite fraction. Changes were also seen for the dendritic cell maturation markers CD80 and CD86. On CD14dim cells, an increase in both CD80 and CD86 expression was seen, in contrast to a selective increase in CD86 expression on CD14bright cells. The co-expression of CD80 and CD86 indicates effective antigen presentation to T cells and support of T helper cell differentiation. The selective expression of CD86 in the absence of CD80 points to a role in generating T regulatory cells. CONCLUSION: The data show that a primary mechanism of action of GBC30 metabolites involves support of more mature phenotypes of antigen-presenting cells, important for immunological decision-making. PMID:22563167

Dynamics of T cell, antigen-presenting cell, and pathogen interactions during recall responses in the lymph node.

PubMed

Chtanova, Tatyana; Han, Seong-Ji; Schaeffer, Marie; van Dooren, Giel G; Herzmark, Paul; Striepen, Boris; Robey, Ellen A

2009-08-21

Memory T cells circulate through lymph nodes where they are poised to respond rapidly upon re-exposure to a pathogen; however, the dynamics of memory T cell, antigen-presenting cell, and pathogen interactions during recall responses are largely unknown. We used a mouse model of infection with the intracellular protozoan parasite, Toxoplasma gondii, in conjunction with two-photon microscopy, to address this question. After challenge, memory T cells migrated more rapidly than naive T cells, relocalized toward the subcapsular sinus (SCS) near invaded macrophages, and engaged in prolonged interactions with infected cells. Parasite invasion of T cells occurred by direct transfer of the parasite from the target cell into the T cell and corresponded to an antigen-specific increase in the rate of T cell invasion. Our results provide insight into cellular interactions during recall responses and suggest a mechanism of pathogen subversion of the immune response.

Parasite Manipulation of the Invariant Chain and the Peptide Editor H2-DM Affects Major Histocompatibility Complex Class II Antigen Presentation during Toxoplasma gondii Infection

PubMed Central

Nishi, Manami; El-Hage, Sandy; Fox, Barbara A.; Bzik, David J.

2015-01-01

Toxoplasma gondii is an obligate intracellular protozoan parasite. This apicomplexan is the causative agent of toxoplasmosis, a leading cause of central nervous system disease in AIDS. It has long been known that T. gondii interferes with major histocompatibility complex class II (MHC-II) antigen presentation to attenuate CD4+ T cell responses and establish persisting infections. Transcriptional downregulation of MHC-II genes by T. gondii was previously established, but the precise mechanisms inhibiting MHC-II function are currently unknown. Here, we show that, in addition to transcriptional regulation of MHC-II, the parasite modulates the expression of key components of the MHC-II antigen presentation pathway, namely, the MHC-II-associated invariant chain (Ii or CD74) and the peptide editor H2-DM, in professional antigen-presenting cells (pAPCs). Genetic deletion of CD74 restored the ability of infected dendritic cells to present a parasite antigen in the context of MHC-II in vitro. CD74 mRNA and protein levels were, surprisingly, elevated in infected cells, whereas MHC-II and H2-DM expression was inhibited. CD74 accumulated mainly in the endoplasmic reticulum (ER), and this phenotype required live parasites, but not active replication. Finally, we compared the impacts of genetic deletion of CD74 and H2-DM genes on parasite dissemination toward lymphoid organs in mice, as well as activation of CD4+ T cells and interferon gamma (IFN-γ) levels during acute infection. Cyst burdens and survival during the chronic phase of infection were also evaluated in wild-type and knockout mice. These results highlight the fact that the infection is influenced by multiple levels of parasite manipulation of the MHC-II antigen presentation pathway. PMID:26195549

Correlation of the cell surface antigens with stage and grade in cancer of the bladder.

PubMed

Emmott, R C; Javadpour, N; Bergman, S M; Soares, T

1979-01-01

We examined 76 bladder tumors of various stages and grades for the presence of the ABO (H) cell surface antigen, using the specific red cell adherence technique. Of the grade I lesions studied 70 per cent were positive for the cell surface antigen and none of the 26 grade III tumors retained the antigens. When correlated with clinical stage the tumors showed no antigens for those of stages B1 to D, while 12 of 16 stage A lesions were positive for the antigen. When stage A lesions were studied and the findings were correlated with recurrence and metastasis/invasion rates the cell surface antigen was present on the initial tumor in only 1 lesion that recurred at an invasive stage. The findings of this study show that the specific red cell adherence technique may be valuable for predicting malignant potential in low grade, low stage cancer of the bladder. If supported by further investigation this technique may offer the capability of selecting low grade, low stage bladder tumors that are destined to invade or metastasize while they are at curable stages.

Role of Metalloproteases in Vaccinia Virus Epitope Processing for Transporter Associated with Antigen Processing (TAP)-independent Human Leukocyte Antigen (HLA)-B7 Class I Antigen Presentation*

PubMed Central

Lorente, Elena; García, Ruth; Mir, Carmen; Barriga, Alejandro; Lemonnier, François A.; Ramos, Manuel; López, Daniel

2012-01-01

The transporter associated with antigen processing (TAP) translocates the viral proteolytic peptides generated by the proteasome and other proteases in the cytosol to the endoplasmic reticulum lumen. There, they complex with nascent human leukocyte antigen (HLA) class I molecules, which are subsequently recognized by the CD8+ lymphocyte cellular response. However, individuals with nonfunctional TAP complexes or tumor or infected cells with blocked TAP molecules are able to present HLA class I ligands generated by TAP-independent processing pathways. Herein, using a TAP-independent polyclonal vaccinia virus-polyspecific CD8+ T cell line, two conserved vaccinia-derived TAP-independent HLA-B*0702 epitopes were identified. The presentation of these epitopes in normal cells occurs via complex antigen-processing pathways involving the proteasome and/or different subsets of metalloproteinases (amino-, carboxy-, and endoproteases), which were blocked in infected cells with specific chemical inhibitors. These data support the hypothesis that the abundant cellular proteolytic systems contribute to the supply of peptides recognized by the antiviral cellular immune response, thereby facilitating immunosurveillance. These data may explain why TAP-deficient individuals live normal life spans without any increased susceptibility to viral infections. PMID:22298786

Antigen Loss Variants: Catching Hold of Escaping Foes.

PubMed

Vyas, Maulik; Müller, Rolf; Pogge von Strandmann, Elke

2017-01-01

Since mid-1990s, the field of cancer immunotherapy has seen steady growth and selected immunotherapies are now a routine and preferred therapeutic option of certain malignancies. Both active and passive cancer immunotherapies exploit the fact that tumor cells express specific antigens on the cell surface, thereby mounting an immune response specifically against malignant cells. It is well established that cancer cells typically lose surface antigens following natural or therapy-induced selective pressure and these antigen-loss variants are often the population that causes therapy-resistant relapse. CD19 and CD20 antigen loss in acute lymphocytic leukemia and chronic lymphocytic leukemia, respectively, and lineage switching in leukemia associated with mixed lineage leukemia (MLL) gene rearrangements are well-documented evidences in this regard. Although increasing number of novel immunotherapies are being developed, majority of these do not address the control of antigen loss variants. Here, we review the occurrence of antigen loss variants in leukemia and discuss the therapeutic strategies to tackle the same. We also present an approach of dual-targeting immunoligand effectively retargeting NK cells against antigen loss variants in MLL-associated leukemia. Novel immunotherapies simultaneously targeting more than one tumor antigen certainly hold promise to completely eradicate tumor and prevent therapy-resistant relapses.

The role and mechanics of dendritic cells in tumor antigen acquisition and presentation following laser immunotherapy

NASA Astrophysics Data System (ADS)

Laverty, Sean M.; Dawkins, Bryan A.; Chen, Wei R.

2018-02-01

We extend our model of the antitumor immune response initiated by laser-immunotherapy treatment to more closely examine key steps in the immune response 1) tumor antigen acquisition by antigen-presenting dendritic cells (DCs) and 2) cytotoxic T cell (CTL) priming by lymphatic DCs. Specifically we explore the formation of DC-CTL complexes that lead to CTL priming. We find that the bias in the dissociation rate of the complex influences the outcome of treatment. In particular, a bias towards priming favors a rapid activated CTL response and the clearance of tumors.

Exploiting the pliability and lateral mobility of Pickering emulsion for enhanced vaccination

NASA Astrophysics Data System (ADS)

Xia, Yufei; Wu, Jie; Wei, Wei; Du, Yiqun; Wan, Tao; Ma, Xiaowei; An, Wenqi; Guo, Aiying; Miao, Chunyu; Yue, Hua; Li, Shuoguo; Cao, Xuetao; Su, Zhiguo; Ma, Guanghui

2018-02-01

A major challenge in vaccine formulations is the stimulation of both the humoral and cellular immune response for well-defined antigens with high efficacy and safety. Adjuvant research has focused on developing particulate carriers to model the sizes, shapes and compositions of microbes or diseased cells, but not antigen fluidity and pliability. Here, we develop Pickering emulsions--that is, particle-stabilized emulsions that retain the force-dependent deformability and lateral mobility of presented antigens while displaying high biosafety and antigen-loading capabilities. Compared with solid particles and conventional surfactant-stabilized emulsions, the optimized Pickering emulsions enhance the recruitment, antigen uptake and activation of antigen-presenting cells, potently stimulating both humoral and cellular adaptive responses, and thus increasing the survival of mice upon lethal challenge. The pliability and lateral mobility of antigen-loaded Pickering emulsions may provide a facile, effective, safe and broadly applicable strategy to enhance adaptive immunity against infections and diseases.

Exosomal cancer immunotherapy is independent of MHC molecules on exosomes.

PubMed

Hiltbrunner, Stefanie; Larssen, Pia; Eldh, Maria; Martinez-Bravo, Maria-Jose; Wagner, Arnika K; Karlsson, Mikael C I; Gabrielsson, Susanne

2016-06-21

Peptide-loaded exosomes are promising cancer treatment vehicles; however, moderate T cell responses in human clinical trials indicate a need to further understand exosome-induced immunity. We previously demonstrated that antigen-loaded exosomes carry whole protein antigens and require B cells for inducing antigen-specific T cells. Therefore, we investigated the relative importance of exosomal major histocompatibility complex (MHC) class I for the induction of antigen-specific T cell responses and tumour protection. We show that ovalbumin-loaded dendritic cell-derived exosomes from MHCI-/- mice induce antigen-specific T cells at the same magnitude as wild type exosomes. Furthermore, exosomes lacking MHC class I, as well as exosomes with both MHC class I and II mismatch, induced tumour infiltrating T cells and increased overall survival to the same extent as syngeneic exosomes in B16 melanoma. In conclusion, T cell responses are independent of exosomal MHC/peptide complexes if whole antigen is present. This establishes the prospective of using impersonalised exosomes, and will greatly increase the feasibility of designing exosome-based vaccines or therapeutic approaches in humans.

Universal Artificial Antigen Presenting Cells to Selectively Propagate T Cells Expressing Chimeric Antigen Receptor Independent of Specificity

PubMed Central

Rushworth, David; Jena, Bipulendu; Olivares, Simon; Maiti, Sourindra; Briggs, Neima; Somanchi, Srinivas; Dai, Jianliang; Lee, Dean; Cooper, Laurence J. N.

2014-01-01

T cells genetically modified to stably express immunoreceptors are being assessed for therapeutic potential in clinical trials. T cells expressing a chimeric antigen receptor (CAR) are endowed with a new specificity to target tumor-associated antigen (TAA) independent of major histocompatibility complex. Our approach to non-viral gene transfer in T cells uses ex vivo numeric expansion of CAR+ T cells on irradiated artificial antigen presenting cells (aAPC) bearing the targeted TAA. The requirement for aAPC to express a desired TAA limits the human application of CARs with multiple specificities when selective expansion through co-culture with feeder cells is sought. As an alternative to expressing individual TAAs on aAPC, we expressed one ligand that could activate CAR+ T cells for sustained proliferation independent of specificity. We expressed a CAR ligand (designated CARL) that binds the conserved IgG4 extracellular domain of CAR and demonstrated CARL+ aAPC propagate CAR+ T cells of multiple specificities. CARL avoids technical issues and costs associated with deploying clinical-grade aAPC for each TAA targeted by a given CAR. Employing CARL enables one aAPC to numerically expand all CAR+ T cells containing the IgG4 domain, and simplifies expansion, testing, and clinical translation of CAR+ T cells of any specificity. PMID:24714354

Targeting tumor antigens to secreted membrane vesicles in vivo induces efficient antitumor immune responses.

PubMed

Zeelenberg, Ingrid S; Ostrowski, Matias; Krumeich, Sophie; Bobrie, Angélique; Jancic, Carolina; Boissonnas, Alexandre; Delcayre, Alain; Le Pecq, Jean-Bernard; Combadière, Béhazine; Amigorena, Sebastian; Théry, Clotilde

2008-02-15

Expression of non-self antigens by tumors can induce activation of T cells in vivo, although this activation can lead to either immunity or tolerance. CD8+ T-cell activation can be direct (if the tumor expresses MHC class I molecules) or indirect (after the capture and cross-presentation of tumor antigens by dendritic cells). The modes of tumor antigen capture by dendritic cells in vivo remain unclear. Here we examine the immunogenicity of the same model antigen secreted by live tumors either in association with membrane vesicles (exosomes) or as a soluble protein. We have artificially addressed the antigen to secreted vesicles by coupling it to the factor VIII-like C1C2 domain of milk fat globule epidermal growth factor-factor VIII (MFG-E8)/lactadherin. We show that murine fibrosarcoma tumor cells that secrete vesicle-bound antigen grow slower than tumors that secrete soluble antigen in immunocompetent, but not in immunodeficient, host mice. This growth difference is due to the induction of a more potent antigen-specific antitumor immune response in vivo by the vesicle-bound than by the soluble antigen. Finally, in vivo secretion of the vesicle-bound antigen either by tumors or by vaccination with naked DNA protects against soluble antigen-secreting tumors. We conclude that the mode of secretion can determine the immunogenicity of tumor antigens and that manipulation of the mode of antigen secretion may be used to optimize antitumor vaccination protocols.

Enhancement of MHC-I antigen presentation via architectural control of pH-responsive, endosomolytic polymer nanoparticles.

PubMed

Wilson, John T; Postma, Almar; Keller, Salka; Convertine, Anthony J; Moad, Graeme; Rizzardo, Ezio; Meagher, Laurence; Chiefari, John; Stayton, Patrick S

2015-03-01

Protein-based vaccines offer a number of important advantages over organism-based vaccines but generally elicit poor CD8(+) T cell responses. We have previously demonstrated that pH-responsive, endosomolytic polymers can enhance protein antigen delivery to major histocompatibility complex class I (MHC-I) antigen presentation pathways thereby augmenting CD8(+) T cell responses following immunization. Here, we describe a new family of nanocarriers for protein antigen delivery assembled using architecturally distinct pH-responsive polymers. Reversible addition-fragmentation chain transfer (RAFT) polymerization was used to synthesize linear, hyperbranched, and core-crosslinked copolymers of 2-(N,N-diethylamino)ethyl methacrylate (DEAEMA) and butyl methacrylate (BMA) that were subsequently chain extended with a hydrophilic N,N-dimethylacrylamide (DMA) segment copolymerized with thiol-reactive pyridyl disulfide (PDS) groups. In aqueous solution, polymer chains assembled into 25 nm micellar nanoparticles and enabled efficient and reducible conjugation of a thiolated protein antigen, ovalbumin. Polymers demonstrated pH-dependent membrane-destabilizing activity in an erythrocyte lysis assay, with the hyperbranched and cross-linked polymer architectures exhibiting significantly higher hemolysis at pH ≤ 7.0 than the linear diblock. Antigen delivery with the hyperbranched and cross-linked polymer architecture enhanced in vitro MHC-I antigen presentation relative to free antigen, whereas the linear construct did not have a discernible effect. The hyperbranched system elicited a four- to fivefold increase in MHC-I presentation relative to the cross-linked architecture, demonstrating the superior capacity of the hyperbranched architecture in enhancing MHC-I presentation. This work demonstrates that the architecture of pH-responsive, endosomolytic polymers can have dramatic effects on intracellular antigen delivery, and offers a promising strategy for enhancing CD8(+) T cell responses to protein-based vaccines.

Functional analysis of HPV-like particle-activated Langerhans cells in vitro.

PubMed

Yan, Lisa; Woodham, Andrew W; Da Silva, Diane M; Kast, W Martin

2015-01-01

Langerhans cells (LCs) are antigen-presenting cells responsible for initiating an immune response against human papillomaviruses (HPVs) entering the epithelial layer in vivo as they are the first immune cell that HPV comes into contact with. LCs become activated in response to foreign antigens, which causes internal signaling resulting in the increased expression of co-stimulatory molecules and the secretion of inflammatory cytokines. Functionally activated LCs are then capable of migrating to the lymph nodes where they interact with antigen-specific T cells and initiate an adaptive T-cell response in vivo. However, HPV has evolved in a manner that suppresses LC function, and thus the induction of antigen-specific T cells is hindered. While many methods exist to monitor the activity of LCs in vitro, the migration and induction of cytotoxic T cells is ultimately indicative of a functional immune response. Here, methods in analyzing functional migration and induction of antigen-specific T cells after stimulation of LCs with HPV virus-like particles in vitro are described.

Effective collaboration between marginal metallophilic macrophages and CD8+ dendritic cells in the generation of cytotoxic T cells

PubMed Central

Backer, Ronald; Schwandt, Timo; Greuter, Mascha; Oosting, Marije; Jüngerkes, Frank; Tüting, Thomas; Boon, Louis; O’Toole, Tom; Kraal, Georg; Limmer, Andreas; den Haan, Joke M. M.

2009-01-01

The spleen is the lymphoid organ that induces immune responses toward blood-borne pathogens. Specialized macrophages in the splenic marginal zone are strategically positioned to phagocytose pathogens and cell debris, but are not known to play a role in the activation of T-cell responses. Here we demonstrate that splenic marginal metallophilic macrophages (MMM) are essential for cross-presentation of blood-borne antigens by splenic dendritic cells (DCs). Our data demonstrate that antigens targeted to MMM as well as blood-borne adenoviruses are efficiently captured by MMM and exclusively transferred to splenic CD8+ DCs for cross-presentation and for the activation of cytotoxic T lymphocytes. Depletion of macrophages in the marginal zone prevents cytotoxic T-lymphocyte activation by CD8+ DCs after antibody targeting or adenovirus infection. Moreover, we show that tumor antigen targeting to MMM is very effective as antitumor immunotherapy. Our studies point to an important role for splenic MMM in the initial steps of CD8+ T-cell immunity by capturing and concentrating blood-borne antigens and the transfer to cross-presenting DCs which can be used to design vaccination strategies to induce antitumor cytotoxic T-cell immunity. PMID:20018690

Immune-Modulation by Epidermal Growth Factor Receptor Inhibitors: Implication on Anti-Tumor Immunity in Lung Cancer

PubMed Central

Herrmann, Amanda C.; Bernatchez, Chantale; Haymaker, Cara; Molldrem, Jeffrey J.; Hong, Waun Ki; Perez-Soler, Roman

2016-01-01

Skin toxicity is the most common toxicity caused by Epidermal Growth Factor Receptor (EGFR) inhibitors, and has been associated with clinical efficacy. As EGFR inhibitors enhance the expression of antigen presenting molecules in affected skin keratinocytes, they may concurrently facilitate neo-antigen presentation in lung cancer tumor cells contributing to anti-tumor immunity. Here, we investigated the modulatory effect of the EGFR inhibitor, erlotinib on antigen presenting molecules and PD-L1, prominent immune checkpoint protein, of skin keratinocytes and lung cancer cell lines to delineate the link between EGFR signaling pathway inhibition and potential anti-tumor immunity. Erlotinib up-regulated MHC-I and MHC-II proteins on IFNγ treated keratinocytes but abrogated IFNγ-induced expression of PD-L1, suggesting the potential role of infiltrating autoreactive T cells in the damage of keratinocytes in affected skin. Interestingly, the surface expression of MHC-I, MHC-II, and PD-L1 was up-regulated in response to IFNγ more often in lung cancer cell lines sensitive to erlotinib, but only expression of PD-L1 was inhibited by erlotinib. Further, erlotinib significantly increased T cell mediated cytotoxicity on lung cancer cells. Lastly, the analysis of gene expression dataset of 186 lung cancer cell lines from Cancer Cell Line Encyclopedia demonstrated that overexpression of PD-L1 was associated with sensitivity to erlotinib and higher expression of genes related to antigen presenting pathways and IFNγ signaling pathway. Our findings suggest that the EGFR inhibitors can facilitate anti-tumor adaptive immune responses by breaking tolerance especially in EGFR driven lung cancer that are associated with overexpression of PD-L1 and genes related to antigen presentation and inflammation. PMID:27467256

Adoptive T cell cancer therapy

NASA Astrophysics Data System (ADS)

Dzhandzhugazyan, Karine N.; Guldberg, Per; Kirkin, Alexei F.

2018-06-01

Tumour heterogeneity and off-target toxicity are current challenges of cancer immunotherapy. Karine Dzhandzhugazyan, Per Guldberg and Alexei Kirkin discuss how epigenetic induction of tumour antigens in antigen-presenting cells may form the basis for multi-target therapies.

Complement opsonization of HIV-1 results in a different intracellular processing pattern and enhanced MHC class I presentation by dendritic cells

PubMed Central

Tjomsland, Veronica; Ellegård, Rada; Burgener, Adam; Mogk, Kenzie; Che, Karlhans F; Westmacott, Garrett; Hinkula, Jorma; Lifson, Jeffrey D; Larsson, Marie

2013-01-01

Induction of optimal HIV-1-specific T-cell responses, which can contribute to controlling viral infection in vivo, depends on antigen processing and presentation processes occurring in DCs. Opsonization can influence the routing of antigen processing and pathways used for presentation. We studied antigen proteolysis and the role of endocytic receptors in MHC class I (MHCI) and II (MHCII) presentation of antigens derived from HIV-1 in human monocyte-derived immature DCs (IDCs) and mature DCs, comparing free and complement opsonized HIV-1 particles. Opsonization of virions promoted MHCI presentation by DCs, indicating that complement opsonization routes more virions toward the MHCI presentation pathway. Blockade of macrophage mannose receptor (MMR) and β7-integrin enhanced MHCI and MHCII presentation by IDCs and mature DCs, whereas the block of complement receptor 3 decreased MHCI and MHCII presentation. In addition, we found that IDC and MDC proteolytic activities were modulated by HIV-1 exposure; complement-opsonized HIV-1 induced an increased proteasome activity in IDCs. Taken together, these findings indicate that endocytic receptors such as MMR, complement receptor 3, and β7-integrin can promote or disfavor antigen presentation probably by routing HIV-1 into different endosomal compartments with distinct efficiencies for degradation of viral antigens and MHCI and MHCII presentation, and that HIV-1 affects the antigen-processing machinery. PMID:23526630

Chemokine programming dendritic cell antigen response: part II - programming antigen presentation to T lymphocytes by partially maintaining immature dendritic cell phenotype.

PubMed

Park, Jaehyung; Bryers, James D

2013-05-01

In a companion article to this study,(1) the successful programming of a JAWSII dendritic cell (DC) line's antigen uptake and processing was demonstrated based on pre-treatment of DCs with a specific 'cocktail' of select chemokines. Chemokine pre-treatment modulated cytokine production before and after DC maturation [by lipopolysaccharide (LPS)]. After DC maturation, it induced an antigen uptake and processing capacity at levels 36% and 82% higher than in immature DCs, respectively. Such programming proffers a potential new approach to enhance vaccine efficiency. Unfortunately, simply enhancing antigen uptake does not guarantee the desired activation and proliferation of lymphocytes, e.g. CD4(+) T cells. In this study, phenotype changes and antigen presentation capacity of chemokine pre-treated murine bone marrow-derived DCs were examined in long-term co-culture with antigen-specific CD4(+) T cells to quantify how chemokine pre-treatment may impact the adaptive immune response. When a model antigen, ovalbumin (OVA), was added after intentional LPS maturation of chemokine-treated DCs, OVA-biased CD4(+) T-cell proliferation was initiated from ~ 100% more undivided naive T cells as compared to DCs treated only with LPS. Secretion of the cytokines interferon-γ, interleukin-1β, interleukin-2 and interleukin-10 in the CD4(+) T cell : DC co-culture (with or without chemokine pre-treatment) were essentially the same. Chemokine programming of DCs with a 7 : 3 ratio of CCL3 : CCL19 followed by LPS treatment maintained partial immature phenotypes of DCs, as indicated by surface marker (CD80 and CD86) expression over time. Results here and in our companion paper suggest that chemokine programming of DCs may provide a novel immunotherapy strategy to obviate the natural endocytosis limit of DC antigen uptake, thus potentially increasing DC-based vaccine efficiency. © 2012 Blackwell Publishing Ltd.

Characterization of self-T-cell response and antigenic determinants of U1A protein with bone marrow-derived dendritic cells in NZB x NZW F1 mice.

PubMed

Suen, J L; Wu, C H; Chen, Y Y; Wu, W M; Chiang, B L

2001-07-01

Systemic lupus erythematosus (SLE) is characterized by the existence of a heterogeneous group of autoantibodies directed against nuclear intact structures, such as nucleosomes and small nuclear ribonucleoproteins (snRNPs). Autoantibodies against snRNPs are of special interest because they are detectable in the majority of SLE patients. Although the B-cell antigenic determinants have been well characterized, very limited data have been reported in regard to the T-cell epitopes of snRNPs. Furthermore, several studies have demonstrated that determination of the auto-T-cell epitopes recognized by freshly isolated T cells is difficult from unprimed lupus mice when self-antigen-pulsed B cells or macrophages are used as antigen-presenting cells (APCs) in vitro. In the present study, we showed a novel approach for determining the auto-T-cell epitopes, using bone marrow-derived dendritic cells (BMDCs) pulsed with the murine U1A protein - an immunodominant antigen of the U1 snRNPs - which is capable of activating freshly isolated T cells from unprimed (NZB x NZW) F1 (BWF1) mice in vitro. The T-cell epitope area was found to be located at the C-terminus of U1A, overlapping the T-cell epitope of human U1A that has been reported in human SLE. Identification of the autoreactive T-cell epitope(s) in snRNPs will help to elucidate how reciprocal T-B determinant spreading of snRNPs emerges in lupus. The results presented here also indicate that it is feasible to use this approach to further explore strategies to design immunotherapy for patients with lupus.

Tuning B cell responses to antigens by cell polarity and membrane trafficking.

PubMed

Del Valle Batalla, Felipe; Lennon-Dumenil, Ana-María; Yuseff, María-Isabel

2018-06-20

The capacity of B lymphocytes to produce specific antibodies, particularly broadly neutralizing antibodies that provide immunity to viral pathogens has positioned them as valuable therapeutic targets for immunomodulation. To become competent as antibody secreting cells, B cells undergo a series of activation steps, which are triggered by the recognition of antigens frequently displayed on the surface of other presenting cells. Such antigens elicit the formation of an immune synapse (IS), where local cytoskeleton rearrangements coupled to mechanical forces and membrane trafficking orchestrate the extraction and processing of antigens in B cells. In this review, we discuss the molecular mechanisms that regulate polarized membrane trafficking and mechanical properties of the immune synapse, as well as the potential extracellular cues from the environment, which may impact the ability of B cells to sense and acquire antigens at the immune synapse. An integrated view of the diverse cellular mechanisms that shape the immune synapse will provide a better understanding on how B cells are efficiently activated. Copyright © 2018 Elsevier Ltd. All rights reserved.

Flt3L controls the development of radiosensitive dendritic cells in the meninges and choroid plexus of the steady-state mouse brain

PubMed Central

Anandasabapathy, Niroshana; Victora, Gabriel D.; Meredith, Matthew; Feder, Rachel; Dong, Baojun; Kluger, Courtney; Yao, Kaihui; Dustin, Michael L.; Nussenzweig, Michel C.; Steinman, Ralph M.

2011-01-01

Antigen-presenting cells in the disease-free brain have been identified primarily by expression of antigens such as CD11b, CD11c, and MHC II, which can be shared by dendritic cells (DCs), microglia, and monocytes. In this study, starting with the criterion of Flt3 (FMS-like receptor tyrosine kinase 3)-dependent development, we characterize the features of authentic DCs within the meninges and choroid plexus in healthy mouse brains. Analyses of morphology, gene expression, and antigen-presenting function established a close relationship between meningeal and choroid plexus DCs (m/chDCs) and spleen DCs. DCs in both sites shared an intrinsic requirement for Flt3 ligand. Microarrays revealed differences in expression of transcripts encoding surface molecules, transcription factors, pattern recognition receptors, and other genes in m/chDCs compared with monocytes and microglia. Migrating pre-DC progenitors from bone marrow gave rise to m/chDCs that had a 5–7-d half-life. In contrast to microglia, DCs actively present self-antigens and stimulate T cells. Therefore, the meninges and choroid plexus of a steady-state brain contain DCs that derive from local precursors and exhibit a differentiation and antigen-presenting program similar to spleen DCs and distinct from microglia. PMID:21788405

Flt3L controls the development of radiosensitive dendritic cells in the meninges and choroid plexus of the steady-state mouse brain.

PubMed

Anandasabapathy, Niroshana; Victora, Gabriel D; Meredith, Matthew; Feder, Rachel; Dong, Baojun; Kluger, Courtney; Yao, Kaihui; Dustin, Michael L; Nussenzweig, Michel C; Steinman, Ralph M; Liu, Kang

2011-08-01

Antigen-presenting cells in the disease-free brain have been identified primarily by expression of antigens such as CD11b, CD11c, and MHC II, which can be shared by dendritic cells (DCs), microglia, and monocytes. In this study, starting with the criterion of Flt3 (FMS-like receptor tyrosine kinase 3)-dependent development, we characterize the features of authentic DCs within the meninges and choroid plexus in healthy mouse brains. Analyses of morphology, gene expression, and antigen-presenting function established a close relationship between meningeal and choroid plexus DCs (m/chDCs) and spleen DCs. DCs in both sites shared an intrinsic requirement for Flt3 ligand. Microarrays revealed differences in expression of transcripts encoding surface molecules, transcription factors, pattern recognition receptors, and other genes in m/chDCs compared with monocytes and microglia. Migrating pre-DC progenitors from bone marrow gave rise to m/chDCs that had a 5-7-d half-life. In contrast to microglia, DCs actively present self-antigens and stimulate T cells. Therefore, the meninges and choroid plexus of a steady-state brain contain DCs that derive from local precursors and exhibit a differentiation and antigen-presenting program similar to spleen DCs and distinct from microglia.

Lentiviral Protein Transfer Vectors Are an Efficient Vaccine Platform and Induce a Strong Antigen-Specific Cytotoxic T Cell Response

PubMed Central

Uhlig, Katharina M.; Schülke, Stefan; Scheuplein, Vivian A. M.; Malczyk, Anna H.; Reusch, Johannes; Kugelmann, Stefanie; Muth, Anke; Koch, Vivian; Hutzler, Stefan; Bodmer, Bianca S.; Schambach, Axel; Buchholz, Christian J.; Waibler, Zoe; Scheurer, Stephan

2015-01-01

ABSTRACT To induce and trigger innate and adaptive immune responses, antigen-presenting cells (APCs) take up and process antigens. Retroviral particles are capable of transferring not only genetic information but also foreign cargo proteins when they are genetically fused to viral structural proteins. Here, we demonstrate the capacity of lentiviral protein transfer vectors (PTVs) for targeted antigen transfer directly into APCs and thereby induction of cytotoxic T cell responses. Targeting of lentiviral PTVs to APCs can be achieved analogously to gene transfer vectors by pseudotyping the particles with truncated wild-type measles virus (MV) glycoproteins (GPs), which use human SLAM (signaling lymphocyte activation molecule) as a main entry receptor. SLAM is expressed on stimulated lymphocytes and APCs, including dendritic cells. SLAM-targeted PTVs transferred the reporter protein green fluorescent protein (GFP) or Cre recombinase with strict receptor specificity into SLAM-expressing CHO and B cell lines, in contrast to broadly transducing vesicular stomatitis virus G protein (VSV-G) pseudotyped PTVs. Primary myeloid dendritic cells (mDCs) incubated with targeted or nontargeted ovalbumin (Ova)-transferring PTVs stimulated Ova-specific T lymphocytes, especially CD8+ T cells. Administration of Ova-PTVs into SLAM-transgenic and control mice confirmed the observed predominant induction of antigen-specific CD8+ T cells and demonstrated the capacity of protein transfer vectors as suitable vaccines for the induction of antigen-specific immune responses. IMPORTANCE This study demonstrates the specificity and efficacy of antigen transfer by SLAM-targeted and nontargeted lentiviral protein transfer vectors into antigen-presenting cells to trigger antigen-specific immune responses in vitro and in vivo. The observed predominant activation of antigen-specific CD8+ T cells indicates the suitability of SLAM-targeted and also nontargeted PTVs as a vaccine for the induction of cytotoxic immune responses. Since cytotoxic CD8+ T lymphocytes are a mainstay of antitumoral immune responses, PTVs could be engineered for the transfer of specific tumor antigens provoking tailored antitumoral immunity. Therefore, PTVs can be used as safe and efficient alternatives to gene transfer vectors or live attenuated replicating vector platforms, avoiding genotoxicity or general toxicity in highly immunocompromised patients, respectively. Thereby, the potential for easy envelope exchange allows the circumventing of neutralizing antibodies, e.g., during repeated boost immunizations. PMID:26085166

Proteomics show antigen presentation processes in human immune cells after AS03-H5N1 vaccination.

PubMed

Galassie, Allison C; Goll, Johannes B; Samir, Parimal; Jensen, Travis L; Hoek, Kristen L; Howard, Leigh M; Allos, Tara M; Niu, Xinnan; Gordy, Laura E; Creech, C Buddy; Hill, Heather; Joyce, Sebastian; Edwards, Kathryn M; Link, Andrew J

2017-06-01

Adjuvants enhance immunity elicited by vaccines through mechanisms that are poorly understood. Using a systems biology approach, we investigated temporal protein expression changes in five primary human immune cell populations: neutrophils, monocytes, natural killer cells, T cells, and B cells after administration of either an Adjuvant System 03 adjuvanted or unadjuvanted split-virus H5N1 influenza vaccine. Monocytes demonstrated the strongest differential signal between vaccine groups. On day 3 post-vaccination, several antigen presentation-related pathways, including MHC class I-mediated antigen processing and presentation, were enriched in monocytes and neutrophils and expression of HLA class I proteins was increased in the Adjuvant System 03 group. We identified several protein families whose proteomic responses predicted seroprotective antibody responses (>1:40 hemagglutination inhibition titer), including inflammation and oxidative stress proteins at day 1 as well as immunoproteasome subunit (PSME1 and PSME2) and HLA class I proteins at day 3 in monocytes. While comparison between temporal proteomic and transcriptomic results showed little overlap overall, enrichment of the MHC class I antigen processing and presentation pathway in monocytes and neutrophils was confirmed by both approaches. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Leptin deficiency in vivo enhances the ability of splenic dendritic cells to activate T cells

PubMed Central

Ramirez, Oscar

2014-01-01

Leptin is a pleiotropic adipokine that is critical for regulating food intake and energy expenditure and also participates in functions of the immune system, including those of antigen-presenting cells. Here, we assess the effect of leptin deficiency on the function splenic dendritic cells (sDC). sDC from leptin-deficient mice (Lepob) were evaluated ex vivo for phenotype, ability to respond to inflammatory stimuli, to acquire and process antigens and to activate T cells. The data show that Lepob sDC express activation markers similar to controls and respond similarly to LPS activation or anti-CD40 cross-linking. In addition, antigen acquisition and processing by Lepob sDC was similar to controls. However, Lepob sDC elicited higher production of IFN-γ in mixed lymphocyte reactions and increased production of IL-2 by antigen-specific T-cell hybridoma relative to controls. To assess Lepob sDC activation of T cells in vivo, Lepob and control mice were infected systemically with Mycobacterium avium. Lepob mice were significantly better at neutralizing the infection as measured by splenic bacterial load over time. This was mirrored with an increased percentage of activated T cells in M. avium-infected Lepob mice. Thus, although no changes were detected in sDC phenotype, activation, antigen processing or presentation, these DC surprisingly presented an enhanced ability to activate T cells ex vivo and in vivo. These data demonstrate that leptin can modulate DC function and suggest that leptin may dampen T-cell responsiveness in the physiological setting. PMID:24966213

Tumour cell lysate-loaded dendritic cell vaccine induces biochemical and memory immune response in castration-resistant prostate cancer patients

PubMed Central

Reyes, D; Salazar, L; Espinoza, E; Pereda, C; Castellón, E; Valdevenito, R; Huidobro, C; Inés Becker, M; Lladser, A; López, M N; Salazar-Onfray, F

2013-01-01

Background: Recently, we produced a tumour antigen-presenting cells (TAPCells) vaccine using a melanoma cell lysate, called TRIMEL, as an antigen source and an activation factor. Tumour antigen-presenting cells induced immunological responses and increased melanoma patient survival. Herein, we investigated the effect of TAPCells loaded with prostate cancer cell lysates (PCCL) as an antigen source, and TRIMEL as a dendritic cell (DC) activation factor; which were co-injected with the Concholepas concholepas haemocyanin (CCH) as an adjuvant on castration-resistant prostate cancer (CRPC) patients. Methods: The lysate mix capacity, for inducing T-cell activation, was analysed by flow cytometry and Elispot. Delayed-type hypersensitivity (DTH) reaction against PCCL, frequency of CD8+ memory T cells (Tm) in blood and prostate-specific antigen (PSA) levels in serum were measured in treated patients. Results: The lysate mix induced functional mature DCs that were capable of activating PCCL-specific T cells. No relevant adverse reactions were observed. Six out of 14 patients showed a significant decrease in levels of PSA. DTH+ patients showed a prolonged PSA doubling-time after treatment. Expansion of functional central and effector CD8+ Tm were detected. Conclusion: Treatment of CRPC patients with lysate-loaded TAPCells and CCH as an adjuvant is safe: generating biochemical and memory immune responses. However, the limited number of cases requires confirmation in a phase II clinical trial. PMID:23989944

Tumour cell lysate-loaded dendritic cell vaccine induces biochemical and memory immune response in castration-resistant prostate cancer patients.

PubMed

Reyes, D; Salazar, L; Espinoza, E; Pereda, C; Castellón, E; Valdevenito, R; Huidobro, C; Inés Becker, M; Lladser, A; López, M N; Salazar-Onfray, F

2013-09-17

Recently, we produced a tumour antigen-presenting cells (TAPCells) vaccine using a melanoma cell lysate, called TRIMEL, as an antigen source and an activation factor. Tumour antigen-presenting cells induced immunological responses and increased melanoma patient survival. Herein, we investigated the effect of TAPCells loaded with prostate cancer cell lysates (PCCL) as an antigen source, and TRIMEL as a dendritic cell (DC) activation factor; which were co-injected with the Concholepas concholepas haemocyanin (CCH) as an adjuvant on castration-resistant prostate cancer (CRPC) patients. The lysate mix capacity, for inducing T-cell activation, was analysed by flow cytometry and Elispot. Delayed-type hypersensitivity (DTH) reaction against PCCL, frequency of CD8(+) memory T cells (Tm) in blood and prostate-specific antigen (PSA) levels in serum were measured in treated patients. The lysate mix induced functional mature DCs that were capable of activating PCCL-specific T cells. No relevant adverse reactions were observed. Six out of 14 patients showed a significant decrease in levels of PSA. DTH(+) patients showed a prolonged PSA doubling-time after treatment. Expansion of functional central and effector CD8(+) Tm were detected. Treatment of CRPC patients with lysate-loaded TAPCells and CCH as an adjuvant is safe: generating biochemical and memory immune responses. However, the limited number of cases requires confirmation in a phase II clinical trial.

Dry eye syndrome: developments and lifitegrast in perspective

PubMed Central

Lollett, Ivonne V; Galor, Anat

2018-01-01

Dry eye (DE) is a chronic ocular condition with high prevalence and morbidity. It has a complex pathophysiology and is multifactorial in nature. Chronic ocular surface inflammation has emerged as a key component of DE that is capable of perpetuating ocular surface damage and leading to symptoms of ocular pain, discomfort, and visual phenomena. It begins with stress to the ocular surface leading to the production of proinflammatory mediators that induce maturation of resident antigen-presenting cells which then migrate to the lymph nodes to activate CD4 T cells. The specific antigen(s) targeted by these pathogenic CD4+ T cells remains unknown. Two emerging theories include self-antigens by autoreactive CD4 T cells or harmless exogenous antigens in the setting of mucosal immunotolerance loss. These CD4 T cells migrate to the ocular surface causing additional inflammation and damage. Lifitegrast is the second topical anti-inflammatory agent to be approved by the US Food and Drug Administration for the treatment of DE and the first to show improvement in DE symptoms. Lifitegrast works by blocking the interaction between intercellular adhesion molecule-1 and lymphocyte functional associated antigen-1, which has been shown to be critical for the migration of antigen-presenting cells to the lymph nodes as well as CD4+ T cell activation and migration to the ocular surface. In four large multicenter, randomized controlled trials, lifitegrast has proven to be effective in controlling both the signs and symptoms of DE with minimal side effects. Further research should include comparative and combination studies with other anti-inflammatory therapies used for DE. PMID:29391773

Human Parvovirus B19 Induced Apoptotic Bodies Contain Altered Self-Antigens that are Phagocytosed by Antigen Presenting Cells

PubMed Central

Thammasri, Kanoktip; Rauhamäki, Sanna; Wang, Liping; Filippou, Artemis; Kivovich, Violetta; Marjomäki, Varpu; Naides, Stanley J.; Gilbert, Leona

2013-01-01

Human parvovirus B19 (B19V) from the erythrovirus genus is known to be a pathogenic virus in humans. Prevalence of B19V infection has been reported worldwide in all seasons, with a high incidence in the spring. B19V is responsible for erythema infectiosum (fifth disease) commonly seen in children. Its other clinical presentations include arthralgia, arthritis, transient aplastic crisis, chronic anemia, congenital anemia, and hydrops fetalis. In addition, B19V infection has been reported to trigger autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis. However, the mechanisms of B19V participation in autoimmunity are not fully understood. B19V induced chronic disease and persistent infection suggests B19V can serve as a model for viral host interactions and the role of viruses in the pathogenesis of autoimmune diseases. Here we investigate the involvement of B19V in the breakdown of immune tolerance. Previously, we demonstrated that the non-structural protein 1 (NS 1) of B19V induces apoptosis in non-permissive cells lines and that this protein can cleave host DNA as well as form NS1-DNA adducts. Here we provide evidence that through programmed cell death, apoptotic bodies (ApoBods) are generated by B19V NS1 expression in a non-permissive cell line. Characterization of purified ApoBods identified potential self-antigens within them. In particular, signature self-antigens such as Smith, ApoH, DNA, histone H4 and phosphatidylserine associated with autoimmunity were present in these ApoBods. In addition, when purified ApoBods were introduced to differentiated macrophages, recognition, engulfment and uptake occurred. This suggests that B19V can produce a source of self-antigens for immune cell processing. The results support our hypothesis that B19V NS1-DNA adducts, and nucleosomal and lysosomal antigens present in ApoBods created in non-permissive cell lines, are a source of self-antigens. PMID:23776709

A novel T cell receptor single-chain signaling complex mediates antigen-specific T cell activity and tumor control

PubMed Central

Stone, Jennifer D.; Harris, Daniel T.; Soto, Carolina M.; Chervin, Adam S.; Aggen, David H.; Roy, Edward J.; Kranz, David M.

2014-01-01

Adoptive transfer of genetically modified T cells to treat cancer has shown promise in several clinical trials. Two main strategies have been applied to redirect T cells against cancer: 1) introduction of a full-length T cell receptor (TCR) specific for a tumor-associated peptide-MHC, or 2) introduction of a chimeric antigen receptor (CAR), including an antibody fragment specific for a tumor cell surface antigen, linked intracellularly to T cell signaling domains. Each strategy has advantages and disadvantages for clinical applications. Here, we present data on the in vitro and in vivo effectiveness of a single-chain signaling receptor incorporating a TCR variable fragment as the targeting element (referred to as TCR-SCS). This receptor contained a single-chain TCR (Vβ-linker-Vα) from a high-affinity TCR called m33, linked to the intracellular signaling domains of CD28 and CD3ζ. This format avoided mispairing with endogenous TCR chains, and mediated specific T cell activity when expressed in either CD4 or CD8 T cells. TCR-SCS-transduced CD8-negative cells showed an intriguing sensitivity, compared to full-length TCRs, to higher densities of less stable pepMHC targets. T cells that expressed this peptide-specific receptor persisted in vivo, and exhibited polyfunctional responses. Growth of metastatic antigen-positive tumors was significantly inhibited by T cells that expressed this receptor, and tumor cells that escaped were antigen loss variants. TCR-SCS receptors represent an alternative targeting receptor strategy that combines the advantages of single-chain expression, avoidance of TCR chain mispairing, and targeting of intracellular antigens presented in complex with MHC proteins. PMID:25082071

Use of Engineered Exosomes Expressing HLA and Costimulatory Molecules to Generate Antigen-specific CD8+ T Cells for Adoptive Cell Therapy.

PubMed

Kim, Sueon; Sohn, Hyun-Jung; Lee, Hyun-Joo; Sohn, Dae-Hee; Hyun, Seung-Joo; Cho, Hyun-Il; Kim, Tai-Gyu

2017-04-01

Dendritic cell-derived exosomes (DEX) comprise an efficient stimulator of T cells. However, the production of sufficient DEX remains a barrier to their broad applicability in immunotherapeutic approaches. In previous studies, genetically engineered K562 have been used to generate artificial antigen presenting cells (AAPC). Here, we isolated exosomes from K562 cells (referred to as CoEX-A2s) engineered to express human leukocyte antigen (HLA)-A2 and costimulatory molecules such as CD80, CD83, and 41BBL. CoEX-A2s were capable of stimulating antigen-specific CD8 T cells both directly and indirectly via CoEX-A2 cross-dressed cells. Notably, CoEX-A2s also generated similar levels of HCMV pp65-specific and MART1-specific CD8 T cells as DEX in vitro. The results suggest that these novel exosomes may provide a crucial reagent for generating antigen-specific CD8 T cells for adoptive cell therapies against viral infection and tumors.

Survival and signaling changes in antigen presenting cell subsets after radiation

NASA Astrophysics Data System (ADS)

Parker, Jennifer Janell

Radiation therapy is a widely used cancer treatment that has the potential to influence anti-tumor immune responses. Both myeloablative and non-myeloablative radiation are often used as part of preparatory regimens for hematopoetic stem cell transplantation, in combination with other chemotherapy or immuno-modulatory (e.g. Anti-thymocyte globulin (ATG)) therapies for both cytotoxic and immune modulatory purposes. However, the mechanisms responsible for the effect of radiation on antigen presenting cell (APC) responsiveness and radioresistance are poorly understood. The first studies described in this thesis were designed to identify and characterize early radiation-induced signaling changes in antigen presenting cells and to determine the effects of these signaling changes on APC receptor expression and function. The NFkappaB pathway in antigen presenting cells was chosen for study because it is activated by radiation in a wide range of other cell types and plays a vital role in the maintenance and regulation of the immune system. The effects of therapeutically relevant doses radiation (2 and 20 Gy) were compared at various timepoints in the human monocytic cell line (U937) using phospho-flow cytometry staining methods and cytometric analysis. These studies demonstrated that radiation-induced changes in the phosphorylation state of NFkappaB family members that were p53 independent. However, these changes were dependent upon activation of ATM in response to single or double-stranded breaks in DNA, as shown in experiments using an inhibitor of ATM and ATM siRNA knockdown U937 cells. In addition, studies examining the effect of radiation on co-stimulatory receptors with and without inhibition of the NFkappaB pathway via phospho-flow cytometry revealed that radiation-induced phosphorylation of NEMO promoted the activation and functional maturation of U937 cells. Furthermore, functional studies using both phospho-flow cytometry and/or mixed lymphocyte reactions to examine co-stimulatory receptor activation, pro-inflammatory cytokine release, and T cell proliferation with and without radiation and inhibition of the NFkappaB pathway, demonstrated that NEMO is necessary for the activation, maturation, and enhanced responsiveness of human subsets of antigen presenting cells that occur after radiation. These findings provided insight into the mechanism of action of radiation-enhanced promotion of the antigen presenting cell responses. The methods of analysis employed can be used for monitoring immune changes that impact immune modulation in transplantation and tumor vaccines studies. Furthermore, NFkappaB pathway proteins have the potential to serve as biomarkers for optimal antitumor responses. The NBD peptide may also have usefulness as a therapeutic agent for inhibition of graft versus host disease (GVHD) in patients who have undergone transplantation. While the first set of experiments focused on antigen presenting cell responsiveness, the second set of experiments were designed to enhance our understanding of why antigen presenting cells, specifically monocytes and dendritic cells, are more radioresistant than conventional T cells. Flow cytometric analysis of various surface markers and intracellular signaling markers were used to examine the mechanisms behind the radioresistance of antigen presenting cells. The experiments described here showed a hierarchy of radiosensitivity among T cells, with naive CD8 T cells being the most radiosensitive and CD4 memory T cells being the most radioresistant. Antigen presenting cells were found to be significantly more radioresistant than T cell subsets (<10 fold decrease after radiation), and among APC, monocytes were more radiosensitive than either total or conventional dendritic cells. Furthermore APC expressed lower levels of Bax after radiation than T cells, and APC subsets that expressed high levels were also more sensitive to radiation induced cell death. These results demonstrate that T cell and APC subsets are dying by apoptosis after radiation, and that the differential level of Bax expression is an important determinant of the relative radiosensitivity of these immune cell subsets. Again, these findings are clinically relevant to transplant patients and patients with tumors receiving radiation therapy since APC survival may have importance for the generation of anti-tumor immunity and post-transplantation immune sequelae such as GVHD. In addition, elucidation of the mechanism of death of APC and T cell subsets, as described in chapter 3, provides potential markers of cell death that can be correlated to good graft versus tumor (GVT) effects versus bad (tumor recurrence and persistence) GVT effects. Thus, understanding the mechanistic basis for radiation-induced changes in APC and the effect of these changes on survival and function is essential for optimizing the use of radiation in transplantation and tumor vaccine treatment protocols.

Antigen presentation by small intestinal epithelial cells uniquely enhances IFN-γ secretion from CD4{sup +} intestinal intraepithelial lymphocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hatano, Ryo; Yamada, Kiyoshi; Iwamoto, Taku

2013-06-14

Highlights: •Small intestinal epithelial cells (sIECs). •sIECs are able to induce antigen specific proliferation of CD4{sup +} IELs. •sIECs induce markedly enhanced IFN-γ secretion by CD4{sup +} IELs. •Induction of enhanced IFN-γ secretion by sIECs is uniquely observed in CD4{sup +} IELs. -- Abstract: Small intestinal epithelial cells (sIECs) express major histocompatibility complex class II molecules even in a normal condition, and are known to function as antigen presenting cells (APCs) at least in vitro. These findings raised the possibility that sIECs play an important role in inducing immune responses against luminal antigens, especially those of intestinal intraepithelial lymphocytes (IELs)more » and lamina propria lymphocytes (LPLs). We herein showed that antigenic stimulation with sIECs induced markedly greater secretion of interferon-gamma (IFN-γ) by CD4{sup +} IELs, but not interleukin (IL)-4, IL-10 and IL-17 although the proliferative response was prominently lower than that with T cell-depleted splenic APCs. In contrast, no enhanced IFN-γ secretion by CD4{sup +} LPLs and primed splenic CD4{sup +} T cells was observed when stimulated with sIECs. Taken together, these results suggest that sIECs uniquely activate CD4{sup +} IELs and induce remarkable IFN-γ secretion upon antigenic stimulation in vivo.« less

Tumor-associated antigen human chorionic gonadotropin beta contains numerous antigenic determinants recognized by in vitro-induced CD8+ and CD4+ T lymphocytes.

PubMed

Dangles, Virginie; Halberstam, Ilan; Scardino, Antonio; Choppin, Jeannine; Wertheimer, Mireille; Richon, Sophie; Quelvennec, Erwann; Moirand, Romain; Guillet, Jean-Gérard; Kosmatopoulos, Kostas; Bellet, Dominique; Zeliszewski, Dominique

2002-02-01

The beta subunit of human chorionic gonadotropin (hCG beta) is markedly overexpressed by neoplastic cells of differing histological origin including those present in colon, breast, prostate and bladder tumors. We have previously shown that some patients with hCG beta-producing urothelial tumors have circulating T cells that proliferate in response to hCG beta. To make a comprehensive study of hCG beta as a potential target for cancer immunotherapy, we investigated whether hCG beta peptides could induce CD4+ or CD8+ T-cell responses in vitro. By stimulating peripheral blood mononuclear cells (PBMCs) from three donors with mixtures of overlapping 16-mer synthetic peptides analogous to portions of either the hCG beta 20-71 or the hCG beta 102-129 region, we established six CD4+ T-cell lines that proliferated specifically in response to five distinct determinants located within these two hCG beta regions. Three antigenic determinants (hCG beta 52-67, 106-121 and 114-125) were presented by HLA-DR molecules, while the two other antigenic determinants (hCG beta 48-63 and 56-67) were presented by HLA-DQ molecules. Interestingly, one T-cell line specific for peptide hCG beta 106-121 recognized hCG beta peptides comprising, at position 117, either an alanine or an aspartic acid residue, with the latter residue being present within the protein expressed by some tumor cells. In addition, three other hCG beta-derived peptides that exhibited HLA-A*0201 binding ability were able to stimulate CD8+ cytotoxic T cells from two HLA-A*0201 donors. These three immunogenic peptides corresponded to regions hCG beta 40-48, hCG beta 44-52 and hCG beta 75-84. Our results indicate that the tumor-associated antigen hCG beta possesses numerous antigenic determinants liable to stimulate CD4+ and CD8+ T lymphocytes, and might thus be an effective target antigen for the immunotherapy of hCG beta-producing tumors.

The Extended Family of CD1d-Restricted NKT Cells: Sifting through a Mixed Bag of TCRs, Antigens, and Functions

PubMed Central

Macho-Fernandez, Elodie; Brigl, Manfred

2015-01-01

Natural killer T (NKT) cells comprise a family of specialized T cells that recognize lipid antigens presented by CD1d. Based on their T cell receptor (TCR) usage and antigen specificities, CD1d-restricted NKT cells have been divided into two main subsets: type I NKT cells that use a canonical invariant TCR α-chain and recognize α-galactosylceramide (α-GalCer), and type II NKT cells that use a more diverse αβ TCR repertoire and do not recognize α-GalCer. In addition, α-GalCer-reactive NKT cells that use non-canonical αβ TCRs and CD1d-restricted T cells that use γδ or δ/αβ TCRs have recently been identified, revealing further diversity among CD1d-restricted T cells. Importantly, in addition to their distinct antigen specificities, functional differences are beginning to emerge between the different members of the CD1d-restricted T cell family. In this review, while using type I NKT cells as comparison, we will focus on type II NKT cells and the other non-invariant CD1d-restricted T cell subsets, and discuss our current understanding of the antigens they recognize, the formation of stimulatory CD1d/antigen complexes, the modes of TCR-mediated antigen recognition, and the mechanisms and consequences of their activation that underlie their function in antimicrobial responses, anti-tumor immunity, and autoimmunity. PMID:26284062

Antigen Presenting Properties of a Myeloid Dendritic-Like Cell in Murine Spleen.

PubMed

Hey, Ying-Ying; O'Neill, Helen C

This paper distinguishes a rare subset of myeloid dendritic-like cells found in mouse spleen from conventional (c) dendritic cells (DC) in terms of phenotype, function and gene expression. These cells are tentatively named "L-DC" since they resemble dendritic-like cells produced in longterm cultures of spleen. L-DC can be distinguished on the basis of their unique phenotype as CD11bhiCD11cloMHCII-CD43+Ly6C-Ly6G-Siglec-F- cells. They demonstrate similar ability as cDC to uptake and retain complex antigens like mannan via mannose receptors, but much lower ability to endocytose and retain soluble antigen. While L-DC differ from cDC by their inability to activate CD4+ T cells, they are capable of antigen cross-presentation for activation of CD8+ T cells, although less effectively so than the cDC subsets. In terms of gene expression, CD8- cDC and CD8+ cDC are quite distinct from L-DC. CD8+ cDC are distinguishable from the other two subsets by expression of CD24a, Clec9a, Xcr1 and Tlr11, while CD8- cDC are distinguished by expression of Ccnd1 and H-2Eb2. L-DC are distinct from the two cDC subsets through upregulated expression of Clec4a3, Emr4, Itgam, Csf1r and CD300ld. The L-DC gene profile is quite distinct from that of cDC, confirming a myeloid cell type with distinct antigen presenting properties.

Remote control of therapeutic T cells through a small molecule-gated chimeric receptor

PubMed Central

Wu, Chia-Yung; Roybal, Kole T.; Puchner, Elias M.; Onuffer, James; Lim, Wendell A.

2016-01-01

There is growing promise in using engineered cells as therapeutic agents. For example, synthetic Chimeric Antigen Receptors (CARs) can redirect T cells to recognize and eliminate tumor cells expressing specific antigens. Despite promising clinical results, excessive activity and poor control over such engineered T cells can cause severe toxicities. We present the design of “ON-switch” CARs that enable small molecule-control over T cell therapeutic functions, while still retaining antigen specificity. In these split receptors, antigen binding and intracellular signaling components only assemble in the presence of a heterodimerizing small molecule. This titratable pharmacologic regulation could allow physicians to precisely control the timing, location, and dosage of T cell activity, thereby mitigating toxicity. This work illustrates the potential of combining cellular engineering with orthogonal chemical tools to yield safer therapeutic cells that tightly integrate both cell autonomous recognition and user control. PMID:26405231

Antigen presenting cells (APCs) from thermally injured and/or septic rats modulate CD4+ T cell responses of naive rat.

PubMed

Fazal, Nadeem; Raziuddin, Syed; Khan, Mehdi; Al-Ghoul, Walid M

2006-01-01

Regulation of immune response is marked by complex interactions among the cells that recognize and present antigens. Antigen presenting cells (APCs), the antigen presenting cell component of the innate immune response plays an important role in effector CD4+ T cell response. Thermal injury and/or superimposed sepsis in rats' leads to suppressed CD4+ T cell functions. We investigated modulations of CD4+ T cell function by APCs (purified non-T cells) from thermally injured and/or septic rats. Rats were subjected to 30% total body surface area scald burn or exposed to 37 degrees C water (Sham burn) and sepsis was induced by cecal-ligation and puncture (CLP) method. At day 3 post-injury animals were sacrificed and CD4+ T cells and APCs from mesenteric lymph nodes (MLN) were obtained using magnetic microbead isolation procedure. APCs from injured rats were co-cultured with sham rat MLN CD4+ T cells and proliferative responses (thymidine incorporation), phenotypic changes (Flow cytometry), IL-2 production (ELISA) and CTLA-4 mRNA (RT-PCR) were determined in naive rat CD4+ T cells. The data indicate that APCs from thermally injured and/or septic rats when co-cultured with CD4+ T cells suppressed CD4+ T cell effector functions. This lack of CD4+ T cell activation was accompanied with altered co-stimulatory molecules, i.e., CD28 and/or CTLA-4 (CD152). In conclusion, our studies indicated that defective APCs from thermally injured and/or septic rats modulate CD4+ T cell functions via changes in co-stimulatory molecules expressed on naive CD4+ T cells. This altered APC: CD4+ T cell interaction leads to suppressed CD4+ T cell activation of healthy animals.

Immunization with SV40-transformed cells yields mainly MHC-restricted monoclonal antibodies

PubMed Central

1986-01-01

Recognition of antigens on cell surfaces only in the context of the MHC- encoded alloantigens of the presenting cell (self + X) has classically been considered the province of T cells. However, evidence from several sources has indicated that B cells and antibodies can exhibit self + X- restricted recognition as well. This report concerns the mAb response to SV40-transformed H-2b fibroblast cell lines. The specificities of the antibodies obtained have been analyzed for binding to a panel of SV40-transformed H-2-syngeneic, H-2-allogeneic, and H-2b mutant fibroblast cell lines, as well as cell lines not bearing cell surface SV40 transformation-associated antigens. A large proportion of primary C57BL/6 (71%) and BALB/c (68%) splenic B cells responding to in vitro stimulation with SV40-transformed H-2b cells recognize cell surface antigens associated with SV40 transformation only when coexpressed with MHC antigens of the immunizing cell, particularly the Kb molecule, on transformed cells. To extensively define the nature of antigen recognition by these antibodies, we have generated and characterized nine hybridoma antibodies specific for SV40-transformed H-2-syngeneic cell lines. Seven of these hybridoma antibodies recognize SV40- associated transformation antigens in the context of H-2b molecules. Six of these are restricted by the Kb molecule and discriminate among a panel of SV40-transformed Kb mutant cell lines, thus confirming the participation of class I MHC-encoded molecules in the recognition by B cells of cell surface antigens. PMID:3014034

Heat Shock Protein-90 Inhibitors Enhance Antigen Expression on Melanomas and Increase T Cell Recognition of Tumor Cells

PubMed Central

Haggerty, Timothy J.; Dunn, Ian S.; Rose, Lenora B.; Newton, Estelle E.; Pandolfi, Franco; Kurnick, James T.

2014-01-01

In an effort to enhance antigen-specific T cell recognition of cancer cells, we have examined numerous modulators of antigen-expression. In this report we demonstrate that twelve different Hsp90 inhibitors (iHsp90) share the ability to increase the expression of differentiation antigens and MHC Class I antigens. These iHsp90 are active in several molecular and cellular assays on a series of tumor cell lines, including eleven human melanomas, a murine B16 melanoma, and two human glioma-derived cell lines. Intra-cytoplasmic antibody staining showed that all of the tested iHsp90 increased expression of the melanocyte differentiation antigens Melan-A/MART-1, gp100, and TRP-2, as well as MHC Class I. The gliomas showed enhanced gp100 and MHC staining. Quantitative analysis of mRNA levels showed a parallel increase in message transcription, and a reporter assay shows induction of promoter activity for Melan-A/MART-1 gene. In addition, iHsp90 increased recognition of tumor cells by T cells specific for Melan-A/MART-1. In contrast to direct Hsp90 client proteins, the increased levels of full-length differentiation antigens that result from iHsp90 treatment are most likely the result of transcriptional activation of their encoding genes. In combination, these results suggest that iHsp90 improve recognition of tumor cells by T cells specific for a melanoma-associated antigen as a result of increasing the expressed intracellular antigen pool available for processing and presentation by MHC Class I, along with increased levels of MHC Class I itself. As these Hsp90 inhibitors do not interfere with T cell function, they could have potential for use in immunotherapy of cancer. PMID:25503774

Novel mechanisms and approaches in immunotherapy for brain tumors.

PubMed

Finocchiaro, Gaetano; Pellegatta, Serena

2015-01-01

Converging data indicate that the immune system is able to recognize cancer epitopes as non-self and mount an immune reaction that may erase, or temporarily block, tumor growth. The immune pressure supports the amplification of immune resistant tumor clones, creating an immune suppressive environment that leads to the formation of a clinically relevant tumor. These general observations also apply to brain tumors and specifically to gliomas. Cancer immunotherapy strategies are aimed at reverting such immune suppression. Two approaches are already used in the clinics. The first one, peptide immunotherapy, has been oriented to the most aggressive glioma, glioblastoma (GBM) where, in the context of EGFR (epidermal growth factor receptor) amplification, a large deletion arises and creates a novel, cancer-specific antigen, EGFRvIII. The second one is dendritic cell immunotherapy. Dendritic cells are potent antigen presenting cells that can be pulsed with autologous tumor lysate or peptide pp65 from cytomegalovirus (CMV) that is present in GBM but not in normal brain. Antigen presentation by dendritic cells is bolstered by preconditioning their injection site with the tetanus/diphtheria toxoid. The third approach is adoptive cell therapy (ACT) in which tumor-specific T cells can be amplified ex vivo and subsequently re-injected to the patient to lyse cells expressing tumor antigens, increasing survival durably in a fraction of melanoma patients. ACT may also be based on T cell transduction of tumor specific receptors or chimeric antigen receptors (CARs). CARs are powerful tools for immunotherapy but off-target toxicity may be an issue as they do not request MHC presentation for activation. Upcoming clinical trial results will clarify the most effective direction for cancer immunotherapy in gliomas and other cancers with poor prognosis.

Dendritic Cells: A Double-Edged Sword in Immune Responses during Chagas Disease

PubMed Central

Gil-Jaramillo, Natalia; Motta, Flávia N.; Favali, Cecília B. F.; Bastos, Izabela M. D.; Santana, Jaime M.

2016-01-01

Dendritic cells (DCs) are the most important member of the antigen presenting cells group due to their ability to recognize antigen at the infection site and their high specialized antigen internalization capacity. These cells have central role in connecting the innate and adaptive immune responses against Trypanosoma cruzi, the causative agent of Chagas disease. These first line defense cells modulate host immune response depending on type, maturation level, cytokine milieu and DC receptor involved in the interactions with T. cruzi, influencing the development of the disease clinic forms. Here, we present a review of DCs–T. cruzi interactions both in human and murine models, pointing out the parasite ability to manipulate DCs activity for the purpose of evading innate immune response and assuring its own survival and persistence. PMID:27471496

Prevalence of feline leukaemia provirus DNA in feline lymphomas.

PubMed

Weiss, Alexander Th A; Klopfleisch, Robert; Gruber, Achim D

2010-12-01

A significant drop in the prevalence of feline leukaemia virus (FeLV) antigenaemic cats and antigen-associated lymphomas has been observed after the introduction of FeLV vaccination and antigen-testing with removal of persistently antigenaemic cats. However, recent reports have indicated that regressively infected cats may contain FeLV provirus DNA and that lymphoma development may be associated with the presence of provirus alone. In the present study, we investigated the presence of FeLV antigen and provirus DNA in 50 lymphomas by immunohistochemistry and semi-nested polymerase chain reaction, respectively. Interestingly, almost 80% of T-cell lymphomas and 60% of B-cell lymphomas contained provirus DNA while only 21% of T-cell lymphomas and 11% of B-cell lymphomas expressed FeLV antigen. In conclusion, our results support previous hypotheses that vaccination and removal of persistently antigenaemic cats have led to a drop in FeLV antigen-expressing lymphomas. However, FeLV provirus DNA is still present in a high percentage of feline lymphomas. Copyright © 2010 ISFM and AAFP. Published by Elsevier Ltd. All rights reserved.

Dynamic Imaging of CD8(+) T cells and dendritic cells during infection with Toxoplasma gondii.

PubMed

John, Beena; Harris, Tajie H; Tait, Elia D; Wilson, Emma H; Gregg, Beth; Ng, Lai Guan; Mrass, Paulus; Roos, David S; Dzierszinski, Florence; Weninger, Wolfgang; Hunter, Christopher A

2009-07-01

To better understand the initiation of CD8(+) T cell responses during infection, the primary response to the intracellular parasite Toxoplasma gondii was characterized using 2-photon microscopy combined with an experimental system that allowed visualization of dendritic cells (DCs) and parasite specific CD8(+) T cells. Infection with T. gondii induced localization of both these populations to the sub-capsular/interfollicular region of the draining lymph node and DCs were required for the expansion of the T cells. Consistent with current models, in the presence of cognate antigen, the average velocity of CD8(+) T cells decreased. Unexpectedly, infection also resulted in modulation of the behavior of non-parasite specific T cells. This TCR-independent process correlated with the re-modeling of the lymph node micro-architecture and changes in expression of CCL21 and CCL3. Infection also resulted in sustained interactions between the DCs and CD8(+) T cells that were visualized only in the presence of cognate antigen and were limited to an early phase in the response. Infected DCs were rare within the lymph node during this time frame; however, DCs presenting the cognate antigen were detected. Together, these data provide novel insights into the earliest interaction between DCs and CD8(+) T cells and suggest that cross presentation by bystander DCs rather than infected DCs is an important route of antigen presentation during toxoplasmosis.

Dynamic Imaging of CD8+ T Cells and Dendritic Cells during Infection with Toxoplasma gondii

PubMed Central

John, Beena; Harris, Tajie H.; Tait, Elia D.; Wilson, Emma H.; Gregg, Beth; Ng, Lai Guan; Mrass, Paulus; Roos, David S.; Dzierszinski, Florence; Weninger, Wolfgang; Hunter, Christopher A.

2009-01-01

To better understand the initiation of CD8+ T cell responses during infection, the primary response to the intracellular parasite Toxoplasma gondii was characterized using 2-photon microscopy combined with an experimental system that allowed visualization of dendritic cells (DCs) and parasite specific CD8+ T cells. Infection with T. gondii induced localization of both these populations to the sub-capsular/interfollicular region of the draining lymph node and DCs were required for the expansion of the T cells. Consistent with current models, in the presence of cognate antigen, the average velocity of CD8+ T cells decreased. Unexpectedly, infection also resulted in modulation of the behavior of non-parasite specific T cells. This TCR-independent process correlated with the re-modeling of the lymph node micro-architecture and changes in expression of CCL21 and CCL3. Infection also resulted in sustained interactions between the DCs and CD8+ T cells that were visualized only in the presence of cognate antigen and were limited to an early phase in the response. Infected DCs were rare within the lymph node during this time frame; however, DCs presenting the cognate antigen were detected. Together, these data provide novel insights into the earliest interaction between DCs and CD8+ T cells and suggest that cross presentation by bystander DCs rather than infected DCs is an important route of antigen presentation during toxoplasmosis. PMID:19578440

[Role of Langerhans cells in the physiopathology of atopic dermatitis].

PubMed

Bieber, T

1995-12-01

The demonstration of IgE receptors on the surface of epidermal dendritic cells and on other antigen presenting cells is a crucial element in the understanding of the pathophysiological role of these cells in the genesis of atopic disease, and especially the atopic dermatitis (AD). The sensibilisation phase to an aeroallergen at the level of nasal or bronchial mucosa and even at the skin may be mediated by dendritic cells expressing Fc epsilon RI. Distinct forms of AD may then represent the equivalent of the ellicitation phase of the classical allergic contact dermatitis. Fc epsilon RI would lead, via specific IgE, to an efficient antigen capture, to the activation of the dendritic cells and finally to an antigen presentation. Thus, AD may represent the paradigma of an IgE-mediated type IV reaction.

Thalidomide inhibits tumor necrosis factor-alpha production and antigen presentation by Langerhans cells.

PubMed

Deng, Liang; Ding, Wanhong; Granstein, Richard D

2003-11-01

Thalidomide is an effective treatment for several inflammatory and autoimmune disorders including erythema nodosum leprosum, Behcet's syndrome, discoid lupus erythematosus, and Crohn's disease. Thalidomide is believed to exert its anti-inflammatory effects, at least in part, by inhibiting tumor necrosis factor-alpha (TNF-alpha) production by monocytes. We studied the effects of thalidomide on epidermal Langerhans cells (LC). LCs are epidermal antigen-presenting dendritic cells that play important roles in skin immune responses. Using the murine epidermis-derived dendritic cell lines, XS106A from A/J mice and XS52 from BALB/c mice as surrogates for LC, we found that thalidomide inhibited TNF-alpha production in a concentration-dependent manner. Northern blot analysis revealed that thalidomide significantly decreased the peak-induced mRNA level of TNF-alpha in XS106A cells and XS52 cells. We then examined the effect of thalidomide on fresh LC enriched to approximately 98% using positive selection of Ia+ cells with antibodies conjugated to magnetic microspheres. TNF-alpha production was reduced by 67.7% at a thalidomide concentration of 200 microg per mL. Thalidomide also had a profound inhibitory effect on the ability of LC to present antigen to a responsive TH1 clone. Thalidomide inhibits TNF-alpha production and the antigen-presenting ability of epidermal LCs. These mechanisms may contribute to the therapeutic effects observed with this agent.

Induction of the c-myc protooncogene following antigen binding to hapten-specific B cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Snow, E.C.; Fetherston, J.; Zimmer, S.

1986-03-01

Considerable controversy has centered on the role that the surface immunoglobulin (sIg) receptor for antigen plays during the induction of B cell activation. Stimulation by anti-Ig reagents has been shown to activate G/sub 0/ B cells to enter the cell cycle. The binding of thymus-dependent antigens to hapten-specific B cell populations apparently does not result in the movement of the antigen-binding cells (ABC) into the G/sub 1/ stage of the cell cycle. However, the authors have recently demonstrated that antigen binding to such hapten-specific B cells does result in the initiation of the membrane phosphatidylinositol cycle. In the present experiments,more » hapten-specific B cells (80-90% ABC, 99% in G/sub 0/) were incubated with either the correct hapten-carrier conjugate, with the carrier protein, or only media for 2 hours at 37/sup 0/C. At that time, total cellular RNA was isolated and subsequently analyzed by either dot blots or Northern gel techniques. The blots were probed with a (/sup 32/P)-c-myc SstI-Xhol fragment. The results indicate that hapten carrier stimulation of the hapten-specific B cells induces enhanced transcription of the c-myc gene. These observations lend further support to the premise that antigen binding to the sIg receptor results in the transduction to the cell of important signals and implicates the active participation of sIg during the process of antigen-mediated B cell activation.« less

Differential TCR signals for T helper cell programming.

PubMed

Morel, Penelope A

2018-05-02

Upon encounter with their cognate antigen naïve CD4 T cells become activated and are induced to differentiate into several possible T helper (Th) cell subsets. This differentiation depends on a number of factors including antigen presenting cells, cytokines and costimulatory molecules. The strength of the T cell receptor (TCR) signal, related to the affinity of TCR for antigen and antigen dose, has emerged as a dominant factor in determining Th cell fate. Recent studies have revealed that TCR signals of high or low strength do not simply induce quantitatively different signals in the T cells, but rather qualitatively distinct pathways can be induced based on TCR signal strength. This review examines the recent literature in this area and highlights important new developments in our understanding of Th cell differentiation and TCR signal strength. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Analysis of the role of tripeptidyl peptidase II in MHC class I antigen presentation in vivo1

PubMed Central

Kawahara, Masahiro; York, Ian A.; Hearn, Arron; Farfan, Diego; Rock, Kenneth L.

2015-01-01

Previous experiments using enzyme inhibitors and RNAi in cell lysates and cultured cells have suggested that tripeptidyl peptidase II (TPPII) plays a role in creating and destroying MHC class I-presented peptides. However, its precise contribution to these processes has been controversial. To elucidate the importance of TPPII in MHC class I antigen presentation, we analyzed TPPII-deficient gene-trapped mice and cell lines from these animals. In these mice, the expression level of TPPII was reduced by >90% compared to wild-type mice. Thymocytes from TPPII gene-trapped mice displayed more MHC class I on the cell surface, suggesting that TPPII normally limits antigen presentation by destroying peptides overall. TPPII gene-trapped mice responded as well as did wild-type mice to four epitopes from lymphocytic choriomeningitis virus (LCMV). The processing and presentation of peptide precursors with long N-terminal extensions in TPPII gene-trapped embryonic fibroblasts was modestly reduced, but in vivo immunization with recombinant lentiviral or vaccinia virus vectors revealed that such peptide precursors induced an equivalent CD8 T cell response in wild type and TPPII-deficient mice. These data indicate while TPPII contributes to the trimming of peptides with very long N-terminal extensions, TPPII is not essential for generating most MHC class I-presented peptides or for stimulating CTL responses to several antigens in vivo. PMID:19841172

Myelin-reactive antibodies initiate T cell-mediated CNS autoimmune disease by opsonization of endogenous antigen.

PubMed

Kinzel, Silke; Lehmann-Horn, Klaus; Torke, Sebastian; Häusler, Darius; Winkler, Anne; Stadelmann, Christine; Payne, Natalie; Feldmann, Linda; Saiz, Albert; Reindl, Markus; Lalive, Patrice H; Bernard, Claude C; Brück, Wolfgang; Weber, Martin S

2016-07-01

In the pathogenesis of central nervous system (CNS) demyelinating disorders, antigen-specific B cells are implicated to act as potent antigen-presenting cells (APC), eliciting waves of inflammatory CNS infiltration. Here, we provide the first evidence that CNS-reactive antibodies (Ab) are similarly capable of initiating an encephalitogenic immune response by targeting endogenous CNS antigen to otherwise inert myeloid APC. In a transgenic mouse model, constitutive production of Ab against myelin oligodendrocyte glycoprotein (MOG) was sufficient to promote spontaneous experimental autoimmune encephalomyelitis (EAE) in the absence of B cells, when mice endogenously contained MOG-recognizing T cells. Adoptive transfer studies corroborated that anti-MOG Ab triggered activation and expansion of peripheral MOG-specific T cells in an Fc-dependent manner, subsequently causing EAE. To evaluate the underlying mechanism, anti-MOG Ab were added to a co-culture of myeloid APC and MOG-specific T cells. At otherwise undetected concentrations, anti-MOG Ab enabled Fc-mediated APC recognition of intact MOG; internalized, processed and presented MOG activated naïve T cells to differentiate in an encephalitogenic manner. In a series of translational experiments, anti-MOG Ab from two patients with an acute flare of CNS inflammation likewise facilitated detection of human MOG. Jointly, these observations highlight Ab-mediated opsonization of endogenous CNS auto-antigen as a novel disease- and/or relapse-triggering mechanism in CNS demyelinating disorders.

Natural killer T cells in health and disease

PubMed Central

Wu, Lan; Van Kaer, Luc

2013-01-01

Natural killer T (NKT) cells are a subset of T lymphocytes that share surface markers and functional characteristics with both conventional T lymphocytes and natural killer cells. Most NKT cells express a semiinvariant T cell receptor that reacts with glycolipid antigens presented by the major histocompatibility complex class I-related protein CD1d on the surface of antigen-presenting cells. NKT cells become activated during a variety of infections and inflammatory conditions, rapidly producing large amounts of immunomodulatory cytokines. NKT cells can influence the activation state and functional properties of multiple other cell types in the immune system and, thus, modulate immune responses against infectious agents, autoantigens, tumors, tissue grafts and allergens. One attractive aspect of NKT cells is that their immunomodulatory activities can be readily harnessed with cognate glycolipid antigens, such as the marine sponge-derived glycosphingolipid alpha-galactosylceramide. These properties of NKT cells are being exploited for therapeutic intervention to prevent or treat cancer, infections, and autoimmune and inflammatory diseases. PMID:21196373

Transfer of allogeneic CD4+ T cells rescues CD8+ T cells in anti-PD-L1–resistant tumors leading to tumor eradication

PubMed Central

Arina, Ainhoa; Karrison, Theodore; Galka, Eva; Schreiber, Karin; Weichselbaum, Ralph R.; Schreiber, Hans

2017-01-01

Adoptively transferred CD8+ T cells can stabilize the size of solid tumors over long periods of time by exclusively recognizing antigen cross-presented on tumor stroma. However, these tumors eventually escape T cell–mediated growth control. The aim of this study was to eradicate such persistent cancers. In our model, the SIYRYYGL antigen is expressed by cancer cells that lack the MHC-I molecule Kb needed for direct presentation, but the antigen is picked up and cross-presented by tumor stroma. A single injection of antigen-specific 2C CD8+ T cells caused long-term inhibition of tumor growth, but without further intervention, tumors started to progress after approximately 3 months. Escape was associated with reduced numbers of circulating 2C cells. Tumor-infiltrating 2C cells produced significantly less TNFα and expressed more of the “exhaustion” markers PD-1 and Tim-3 than T cells from lymphoid organs. High-dose local ionizing radiation, depletion of myeloid-derived suppressor cells, infusions of additional 2C cells, and antibodies blocking PD-L1 did not prevent tumor escape. In contrast, adoptive transfer of allogeneic CD4+ T cells restored the numbers of circulating Ag-specific CD8+ T cells and their intratumoral function, resulting in tumor eradication. These CD4+ T cells had no antitumor effects in the absence of CD8+ T cells and recognized the alloantigen cross-presented on tumor stroma. CD4+ T cells might also be effective in cancer patients when PD1/PD-L1 blockade does not rescue intratumoral CD8+ T-cell function and tumors persist. PMID:28077434

A review of mechanistic insight and application of pH-sensitive liposomes in drug delivery.

PubMed

Paliwal, Shivani Rai; Paliwal, Rishi; Vyas, Suresh P

2015-05-01

The pH-sensitive liposomes have been extensively used as an alternative to conventional liposomes in effective intracellular delivery of therapeutics/antigen/DNA/diagnostics to various compartments of the target cell. Such liposomes are destabilized under acidic conditions of the endocytotic pathway as they usually contain pH-sensitive lipid components. Therefore, the encapsulated content is delivered into the intracellular bio-environment through destabilization or its fusion with the endosomal membrane. The therapeutic efficacy of pH-sensitive liposomes enables them as biomaterial with commercial utility especially in cancer treatment. In addition, targeting ligands including antibodies can be anchored on the surface of pH-sensitive liposomes to target specific cell surface receptors/antigen present on tumor cells. These vesicles have also been widely explored for antigen delivery and serve as immunological adjuvant to enhance the immune response to antigens. The present review deals with recent research updates on application of pH-sensitive liposomes in chemotherapy/diagnostics/antigen/gene delivery etc.

Spontaneous CD4+ and CD8+ T‐cell responses directed against cancer testis antigens are present in the peripheral blood of testicular cancer patients

PubMed Central

Pearce, Hayden; Hutton, Paul; Chaudhri, Shalini; Porfiri, Emilio; Patel, Prashant; Viney, Richard

2017-01-01

Cancer/testis antigen (CTAg) expression is restricted to spermatogenic cells in an immune‐privileged site within the testis. However, these proteins are expressed aberrantly by malignant cells and T‐cell responses against CTAgs develop in many cancer patients. We investigated the prevalence, magnitude and phenotype of CTAg‐specific T cells in the blood of patients with testicular germ cell tumors (TGCTs). CD8+ and CD4+ T‐cell responses against MAGE‐A family antigens were present in 44% (20/45) of patients’ samples assayed by ex vivo IFN‐γ ELISPOT. The presence of MAGE‐specific CD8+ T cells was further determined following short‐term in vitro expansion through the use of pMHC‐I multimers containing known immunogenic peptides. Longitudinal analysis revealed that the frequency of MAGE‐specific T cells decreased by 89% following orchidectomy suggesting that persistence of tumor antigen is required to sustain CTAg‐specific T‐cell immunity. Notably, this decrease correlated with a decline in the global effector/memory T‐cell pool following treatment. Spontaneous T‐cell immunity against CTAg proteins therefore develops in many patients with testicular cancer and may play an important role in the excellent clinical outcome of patients with this tumor subtype. PMID:28555838

Mycobacterium tuberculosis impairs dendritic cell functions through the serine hydrolase Hip1

PubMed Central

Madan-Lala, Ranjna; Sia, Jonathan Kevin; King, Rebecca; Adekambi, Toidi; Monin, Leticia; Khader, Shabaana A; Pulendran, Bali; Rengarajan, Jyothi

2014-01-01

Mycobacterium tuberculosis (Mtb) is a highly successful human pathogen that primarily resides in host phagocytes, such as macrophages and dendritic cells (DCs), and interferes with their functions. While multiple strategies used by Mtb to modulate macrophage responses have been discovered, interactions between Mtb and DCs are less well understood. DCs are the primary antigen presenting acells (APCs) of the immune system and play a central role in linking innate and adaptive immune responses to microbial pathogens. In this study we show that Mtb impairs DC cytokine secretion, maturation and antigen presentation through the cell envelope-associated serine hydrolase Hip1. Compared to wild type, a hip1 mutant strain of Mtb induced enhanced levels of the key T helper 1 (Th1)-inducing cytokine IL-12, as well as other proinflammatory cytokines (IL-23, IL-6, TNF-α, IL-1β, IL-18) in DCs via MyD88- and TLR2/9-dependent pathways, indicating that Hip1 restricts optimal DC inflammatory responses. Infection with the hip1 mutant also induced higher levels of MHC class II and co-stimulatory molecules, CD40 and CD86, indicating that Mtb impairs DC maturation through Hip1. Further, we show that Mtb promotes sub-optimal antigen presentation, as DCs infected with the hip1 mutant showed increased capacity to present antigen to OT-II- and early secreted antigenic target 6 (ESAT-6)-specific transgenic CD4 T cells and enhanced Th1 and Th17 polarization. Overall, these data show that Mtb impairs DC functions and modulates the nature of antigen-specific T cell responses, with important implications for vaccination strategies. PMID:24659689

Ultraviolet-B radiation induces modulation of antigen presentation of herpes simplex virus by human epidermal cells.

PubMed

van der Molen, R G; Out-Luiting, C; Claas, F H; Norval, M; Koerten, H K; Mommaas, A M

2001-06-01

Although ultraviolet (UV) B radiation is known to be immunosuppressive, there is little information regarding a relevant immunological endpoint to assess human subjects in vivo. Therefore, we have examined the effect of in vivo UV radiation on the ability of human epidermal cells (EC) to present herpes simplex virus (HSV) antigens to memory T cells. Human volunteers, who were seropositive for HSV, were exposed to one minimal erythemal dose (MED) for four consecutive days. EC, prepared from suction blister roofs, were co-cultured with autologous T cells in the presence of HSV. HSV antigen presentation by UV-exposed EC was increased compared with control, nonexposed EC. This up-regulation correlated with an influx of macrophages into the epidermis, which are considered to be associated with UV-induced tolerance. Altering the UV protocol to a sub-erythemal UV dose for four consecutive days or to a single high dose of 2 MED, resulted in suppressed HSV antigen presentation, without the influx of the UV-macrophages. One of the goals of the present study was to eventually use this HSV system to investigate sunscreen immunoprotection. A pilot study with a TiO2-containing sunscreen suggested that the endpoint for UV-induced immunosuppression presented here is promising to be used for human in vivo sunscreen immunoprotection studies.

Evaluation of accessory cell heterogeneity. III. Role of dendritic cells in the in vitro activation of the antibody response to soluble antigens.

PubMed

Erb, P; Ramila, G; Sklenar, I; Kennedy, M; Sunshine, G H

1985-05-01

Dendritic cells and macrophages obtained from spleen and peritoneal exudate were tested as accessory cells for the activation of lymphokine production by T cells, for supporting T-B cooperation and for the induction of antigen-specific T helper cells. Dendritic cells as well as macrophages were able to activate T cells for interleukin-2 secretion and functioned as accessory cells in T-B cooperation, but only macrophages induced T helper cells, which cooperate with B cells by a linked recognition interaction, to soluble antigens. Dendritic cell- and antigen-activated T cells also did not help B cells in the presence of Con A supernatants which contained various T cell- and B cell-stimulatory factors. The failure of dendritic cells to differentiate memory into functional T helper cells, but their efficient accessory cell function in T-B cooperation, where functional T helper cells are already present, can be best explained by a differential accessory cell requirement for T helper cell activation dependent on the differentiation stage of the T helper cell.

The Role of B Cells for in Vivo T Cell Responses to a Friend Virus-Induced Leukemia

NASA Astrophysics Data System (ADS)

Schultz, Kirk R.; Klarnet, Jay P.; Gieni, Randall S.; Hayglass, Kent T.; Greenberg, Philip D.

1990-08-01

B cells can function as antigen-presenting cells and accessory cells for T cell responses. This study evaluated the role of B cells in the induction of protective T cell immunity to a Friend murine leukemia virus (F-MuLV)-induced leukemia (FBL). B cell-deficient mice exhibited significantly reduced tumor-specific CD4^+ helper and CD8^+ cytotoxic T cell responses after priming with FBL or a recombinant vaccinia virus containing F-MuLV antigens. Moreover, these mice had diminished T cell responses to the vaccinia viral antigens. Tumor-primed T cells transferred into B cell-deficient mice effectively eradicated disseminated FBL. Thus, B cells appear necessary for efficient priming but not expression of tumor and viral T cell immunity.

Trans-nodal migration of resident dendritic cells into medullary interfollicular regions initiates immunity to influenza vaccine

PubMed Central

Woodruff, Matthew C.; Heesters, Balthasar A.; Herndon, Caroline N.; Groom, Joanna R.; Thomas, Paul G.; Luster, Andrew D.; Turley, Shannon J.

2014-01-01

Dendritic cells (DCs) are well established as potent antigen-presenting cells critical to adaptive immunity. In vaccination approaches, appropriately stimulating lymph node–resident DCs (LNDCs) is highly relevant to effective immunization. Although LNDCs have been implicated in immune response, their ability to directly drive effective immunity to lymph-borne antigen remains unclear. Using an inactive influenza vaccine model and whole node imaging approaches, we observed surprising responsiveness of LNDC populations to vaccine arrival resulting in a transnodal repositioning into specific antigen collection sites within minutes after immunization. Once there, LNDCs acquired viral antigen and initiated activation of viral specific CD4+ T cells, resulting in germinal center formation and B cell memory in the absence of skin migratory DCs. Together, these results demonstrate an unexpected stimulatory role for LNDCs where they are capable of rapidly locating viral antigen, driving early activation of T cell populations, and independently establishing functional immune response. PMID:25049334

Neutral Polymer Micelle Carriers with pH-Responsive, Endosome-Releasing Activity Modulate Antigen Trafficking to Enhance CD8 T-Cell Responses

PubMed Central

Keller, Salka; Wilson, John T; Patilea, Gabriela I; Kern, Hanna B; Convertine, Anthony J; Stayton, Patrick S

2014-01-01

Synthetic subunit vaccines need to induce CD8+ cytotoxic T-cell (CTL) responses for effective vaccination against intracellular pathogens. Most subunit vaccines primarily generate humoral immune responses, with a weaker than desired CD8+ cytotoxic T-cell response. Here, a neutral, pH-responsive polymer micelle carrier that alters intracellular antigen trafficking was shown to enhance CD8+ T-cell responses with a correlated increase in cytosolic delivery and a decrease in exocytosis. Polymer diblock carriers consisted of a N-(2-hydroxypropyl) methacrylamide corona block with pendant pyridyl disulfide groups for reversible conjugation of thiolated ovalbumin, and a tercopolymer ampholytic core-forming block composed of propylacrylic acid (PAA), dimethylaminoethyl methacrylate (DMAEMA), and butyl methacrylate (BMA). The diblock copolymers self-assembled into 25–30 nm diameter micellar nanoparticles. Conjugation of ovalbumin to the micelles significantly enhanced antigen cross-presentation in vitro relative to free ovalbumin, an unconjugated physical mixture of ovalbumin and polymer, and a non pH-responsive micelle-ovalbumin control. Mechanistic studies in a murine dendritic cell line (DC2.4) demonstrated micelle-mediated enhancements in intracellular antigen retention and cytosolic antigen accumulation. Approximately 90% of initially internalized ovalbumin-conjugated micelles were retained in cells after 1.5 h, compared to only ~40% for controls. Furthermore, cells dosed with conjugates displayed 67-fold higher cytosolic antigen levels relative to soluble ovalbumin 4 h post uptake. Subcutaneous immunization of mice with ovalbumin-polymer conjugates significantly enhanced antigen-specific CD8+ T cell responses (0.4 % IFN-γ+ of CD8+) compared to immunization with soluble protein, ovalbumin and polymer mixture, and the control micelle without endosome-releasing activity. Additionally, pH-responsive carrier facilitated antigen delivery to antigen presenting cells in the draining lymph nodes. As early as 90 min post injection ova-micelle conjugates were associated with 28% and 55% of dendritic cells and macrophages, respectively. After 24 h, conjugates preferentially associated with dendritic cells, affording 30-, 3-, and 3-fold enhancements in uptake relative to free protein, physical mixture, and the non pH-responsive conjugate controls, respectively. These results demonstrate the potential of pH-responsive polymeric micelles for use in vaccine applications that rely on CD8+ T cell activation. PMID:24698946

Neutral polymer micelle carriers with pH-responsive, endosome-releasing activity modulate antigen trafficking to enhance CD8(+) T cell responses.

PubMed

Keller, Salka; Wilson, John T; Patilea, Gabriela I; Kern, Hanna B; Convertine, Anthony J; Stayton, Patrick S

2014-10-10

Synthetic subunit vaccines need to induce CD8(+) cytotoxic T cell (CTL) responses for effective vaccination against intracellular pathogens. Most subunit vaccines primarily generate humoral immune responses, with a weaker than desired CD8(+) cytotoxic T cell response. Here, a neutral, pH-responsive polymer micelle carrier that alters intracellular antigen trafficking was shown to enhance CD8(+) T cell responses with a correlated increase in cytosolic delivery and a decrease in exocytosis. Polymer diblock carriers consisted of a N-(2-hydroxypropyl) methacrylamide corona block with pendent pyridyl disulfide groups for reversible conjugation of thiolated ovalbumin, and a tercopolymer ampholytic core-forming block composed of propylacrylic acid (PAA), dimethylaminoethyl methacrylate (DMAEMA), and butyl methacrylate (BMA). The diblock copolymers self-assembled into 25-30nm diameter micellar nanoparticles. Conjugation of ovalbumin to the micelles significantly enhanced antigen cross-presentation in vitro relative to free ovalbumin, an unconjugated physical mixture of ovalbumin and polymer, and a non-pH-responsive micelle-ovalbumin control. Mechanistic studies in a murine dendritic cell line (DC 2.4) demonstrated micelle-mediated enhancements in intracellular antigen retention and cytosolic antigen accumulation. Approximately 90% of initially internalized ovalbumin-conjugated micelles were retained in cells after 1.5h, compared to only ~40% for controls. Furthermore, cells dosed with conjugates displayed 67-fold higher cytosolic antigen levels relative to soluble ovalbumin 4h post uptake. Subcutaneous immunization of mice with ovalbumin-polymer conjugates significantly enhanced antigen-specific CD8(+) T cell responses (0.4% IFN-γ(+) of CD8(+)) compared to immunization with soluble protein, ovalbumin and polymer mixture, and the control micelle without endosome-releasing activity. Additionally, pH-responsive carrier facilitated antigen delivery to antigen presenting cells in the draining lymph nodes. As early as 90min post injection, ova-micelle conjugates were associated with 28% and 55% of dendritic cells and macrophages, respectively. After 24h, conjugates preferentially associated with dendritic cells, affording 30-, 3-, and 3-fold enhancements in uptake relative to free protein, physical mixture, and the non-pH-responsive conjugate controls, respectively. These results demonstrate the potential of pH-responsive polymeric micelles for use in vaccine applications that rely on CD8(+) T cell activation. Copyright © 2014 Elsevier B.V. All rights reserved.

Pathologic findings in red-tailed hawks (Buteo jamaicensis) and Cooper's hawks (Accipiter cooper) naturally infected with West Nile virus.

PubMed

Wünschmann, Arno; Shivers, Jan; Bender, Jeff; Carroll, Larry; Fuller, Susan; Saggese, Miguel; van Wettere, Arnaud; Redig, Pat

2004-09-01

Carcasses of 13 red-tailed hawks (RTHAs) and 11 Cooper's hawks (COHAs) were tested for West Nile virus (WNV) using WNV-specific reverse transcriptase-polymerase chain reaction (RT-PCR) on fresh brain tissue and WNV-specific immunohistochemistry (IHC) on various organs. Ten COHAs (91%) and 11 RTHAs (85%) were positive for WNV RNA by RT-PCR. All 11 COHAs (100%) and 10 RTHAs (77%) were positive for WNV antigen by IHC. A triad of inflammatory lesions, including chronic lymphoplasmacytic and histiocytic encephalitis, endophthalmitis, and myocarditis, was common in both species. In COHAs, the heart (54%), cerebrum (50%), and eye (45%) were the organs that most commonly contained WNV antigen. The amount of WNV antigen was usually small. In RTHAs, the kidney (38%), cerebrum (38%), cerebellum (38%), and eye (36%) were the organs most commonly containing WNV antigen. Unlike COHAs, larger amounts of WNV antigen were present in the cerebrum of RTHAs. WNV antigen was detected in similar cell populations in both species, including neurons of brain, spinal cord, and retina, pigmented epithelial cells of the retina, epithelial cells of renal medullary tubules, cardiomyocytes, endothelial cells and smooth muscle cells of arteries, dendritic cells of splenic lymph follicles, exocrine pancreatic cells, adrenal cells, and keratinocytes of the skin. The study presents strong evidence that WNV can cause a chronic fatal disease in RTHAs and COHAs. The lesion distribution of WNV infection in both species is variable, but inflammatory lesions are common, and a triad of lesions including encephalitis, myocarditis, and endophthalmitis is indicative of WNV infection in both species.

A T-Cell Receptor Breaks the Rules | Center for Cancer Research

Cancer.gov

Most mature T cells function immunologically when a T-cell receptor (TCR) located on the cell surface encounters and engages its ligand, a major histocompatability complex (MHC), which displays a specific part of a target protein called an antigen. This antigen-presenting complex is assembled from one of the dozen or so MHC molecules that every person inherits from their

Activation of professional antigen presenting cells by acharan sulfate isolated from giant African snail, Achatina fulica.

PubMed

Kim, Hyun-Sun; Lee, Young-Hee; Lee, Young-Ran; Im, Sun-A; Lee, Jae-Kwon; Kim, Yeong Shik; Sim, Joon-Soo; Choi, Hyung Seok; Lee, Chong-Kil

2007-07-01

Acharan sulfate isolated from the giant African snail, Achatina fulica, has been reported to have antitumor activity in vivo. In an effort to determine the mechanisms of its antitumor activity, we examined the effects of acharan sulfate on professional antigen presenting cells (APCs). Acharan sulfate increased the phagocytic activity, the production of cytokines such as TNF-alpha and IL-1beta, and the release of nitric oxide on a macrophage cell line, Raw 264.7 cells. In addition, acharan sulfate induced phenotypic and functional maturation of immature dendritic cells (DCs). Immature DCs cultured with acharan sulfate expressed higher levels of class II MHC molecules and major co-stimulatory molecules such as B7-1, B7-2, and CD40. Functional maturation of immature DCs cultured in the presence of acharan sulfate was confirmed by the increased allostimulatory capacity and IL-12 production. These results suggest that the antitumor activity of acharan sulfate is partly due to the activation of professional antigen presenting cells.

Mechanistic studies of systemic immune responses induced by laser-nanotechnology

NASA Astrophysics Data System (ADS)

Chen, Wei R.; Zhou, Feifan; Henderson, Brock; Vasquez, Bailey; Liu, Hong; Hode, Tomas; Nordquist, Robert E.

2014-02-01

With the help of the specific absorption spectrum of carbon nanotubes, we achieved selective photothermal tumor cell destruction, particularly using a near-infrared laser to reduce potential damage to untargeted tissues. Combined with immunological stimulation, using a novel adjuvant, we also observed the anti-tumor immune responses when treating animal tumors using the laser-nano treatment. In fact, the local application of laser-nano-immunotherapy appeared to result in a systemic curative effect. In our mechanistic study, we found that the laser-nano-immuno treatment can activate antigen-presenting cells, such as dendritic cells (DCs). More importantly, the uptake and presentation of antigens by these antigen presenting cells were significantly enhanced, as shown by the strong binding of tumor cells and DCs as well as the proliferation of T cells caused by the DCs after the DCs had been incubated with laser-nano-immuno treated tumors. These cellular observations provide evidence that a systemic anti-tumor immune response was induced by the combination of laser and nanotechnology.

Routine detection of Epstein-Barr virus specific T-cells in the peripheral blood by flow cytometry

NASA Technical Reports Server (NTRS)

Crucian, B. E.; Stowe, R. P.; Pierson, D. L.; Sams, C. F.

2001-01-01

The ability to detect cytomegalovirus-specific T-cells (CD4(+)) in the peripheral blood by flow cytometry has been recently described by Picker et al. In this method, cells are incubated with viral antigen and responding (cytokine producing) T-cells are then identified by flow cytometry. To date, this technique has not been reliably used to detect Epstein-Barr virus (EBV)-specific T-cells primarily due to the superantigen/mitogenic properties of the virus which non-specifically activate T-cells. By modifying culture conditions under which the antigens are presented, we have overcome this limitation and developed an assay to detect and quantitate EBV-specific T-cells. The detection of cytokine producing T-cells by flow cytometry requires an extremely strong signal (such as culture in the presence of PMA and ionomycin). Our data indicate that in modified culture conditions (early removal of viral antigen) the non-specific activation of T-cells by EBV is reduced, but antigen presentation will continue uninhibited. Using this method, EBV-specific T-cells may be legitimately detected using flow cytometry. No reduction in the numbers of antigen-specific T-cells was observed by the early removal of target antigen when verified using cytomegalovirus antigen (a virus with no non-specific T-cell activation properties). In EBV-seropositive individuals, the phenotype of the EBV-specific cytokine producing T-cells was evaluated using four-color flow cytometry and found to be CD45(+), CD3(+), CD4(+), CD45RA(-), CD69(+), CD25(-). This phenotype indicates the stimulation of circulating previously unactivated memory T-cells. No cytokine production was observed in CD4(+) T-cells from EBV-seronegative individuals, confirming the specificity of this assay. In addition, the use of four color cytometry (CD45, CD3, CD69, IFNgamma/IL-2) allows the total quantitative assessment of EBV-specific T-cells while monitoring the interference of EBV non-specific mitogenic activity. This method may have significant utility for the monitoring of the immune response to latent virus infection/reactivation.

Non-covalent pomegranate (Punica granatum) hydrolyzable tannin-protein complexes modulate antigen uptake, processing and presentation by a T-cell hybridoma line co-cultured with murine peritoneal macrophages.

PubMed

Madrigal-Carballo, Sergio; Haas, Linda; Vestling, Martha; Krueger, Christian G; Reed, Jess D

2016-12-01

In this work we characterize the interaction of pomegranate hydrolyzable tannins (HT) with hen egg-white lysozyme (HEL) and determine the effects of non-covalent tannin-protein complexes on macrophage endocytosis, processing and presentation of antigen. We isolated HT from pomegranate and complex to HEL, the resulting non-covalent tannin-protein complex was characterized by gel electrophoresis and MALDI-TOF MS. Finally, cell culture studies and confocal microscopy imaging were conducted on the non-covalent pomegranate HT-HEL protein complexes to evaluate its effect on macrophage antigen uptake, processing and presentation to T-cell hybridomas. Our results indicate that non-covalent pomegranate HT-HEL protein complexes modulate uptake, processing and antigen presentation by mouse peritoneal macrophages. After 4 h of pre-incubation, only trace amounts of IL-2 were detected in the co-cultures treated with HEL alone, whereas a non-covalent pomegranate HT-HEL complex had already reached maximum IL-2 expression. Pomegranate HT may increase rate of endocytose of HEL and subsequent expression of IL-2 by the T-cell hybridomas.

Langerhans Cells and Their Role in Oral Mucosal Diseases

PubMed Central

Upadhyay, Juhi; Upadhyay, Ram B; Agrawal, Pankaj; Jaitley, Shweta; Shekhar, Rhitu

2013-01-01

Dendritic cells are arguably the most potent antigen-presenting cells and may be the only cells capable of initiating the adaptive immune response. The epithelial residents of dendritic cells are Langerhans cells, which serve as the “sentinels” of the mucosa, altering the immune system not only to pathogen entry but also of tolerance to self antigen and commensal microbes. Oral mucosal Langerhans cells are capable of engaging and internalizing a wide variety of pathogens and have been found responsive to nickel in patients with nickel allergies, oral Candida species, oral lichen planus, lichenoid drug eruptions, graft versus host diseases, periodontal diseases median rhomboid glossitis, human immunodeficiency virus infection, hairy leukoplakia of the tongue, and oral squamous cell carcinoma. Review focuses on the role of antigen-presenting cells in particular Langerhans cells to better understand the mechanisms underlying immune responses. In this review, comprehensive detail about mucosal diseases has been compiled using the PubMed database and through textbooks. PMID:24251267

Mixed isotype class II antigen expression. A novel class II molecule is expressed on a murine B cell lymphoma

PubMed Central

1989-01-01

The structures of Ia molecules expressed by two BALB/c B cell lymphoma lines, A20-1.11 (A20) and 2PK3, were analyzed in an effort to explain the differences in antigen-presenting capacity displayed by these cells. Alloreactive T cell hybridomas specific for I-Ad and antigen- specific, I-Ad-restricted T cells responded well to A20 as the APC. The same alloreactive T cell hybridomas responded weakly or not at all to 2PK3 and the responses of the antigen-specific, I-Ad-restricted T cells were consistently lower to antigen presented by 2PK3 as compared with A20. T cells restricted to I-Ed responded equally well to either A20 or 2PK3 as APC. Additionally 2PK3, but not A20, stimulated a strong syngeneic mixed lymphocyte response. Structural analyses of the Ia antigens revealed that I-A and I-E molecules were expressed by A20, whereas an I-E and a novel I-A-like molecule were expressed by 2PK3. The novel class II molecule was affinity purified from 2PK3 cells using an mAb specific for Ad beta (MK-D6), and this molecule was subsequently shown by an RIA to react with an E alpha-specific mAb (14-4-4S) as well. Chain-specific polyclonal antisera raised against I-A and I-E alpha and beta chains indicated that the 2PK3 "I-A" alpha chain reacted in immunoblot with E alpha-specific and not A alpha-specific antisera, whereas the beta chain reacted with A beta- and not E beta-specific antisera. Peptide map and partial amino acid sequence analyses indicated that the "I-A" molecule expressed by 2PK3 represented a mixed isotype structure resulting from the pairing of Ed alpha with Ad beta. By immunofluorescence staining analysis, 2PK3 did not react with an mAb specific for Ad alpha. 2PK3 was capable of limited antigen presentation through the mixed isotype molecule to I-Ad-restricted OVA-specific T cell hybridomas, although the responses induced were low compared with presentation through I-A on A20. Previous descriptions of the expression of mixed isotype class II molecules in the mouse have resulted primarily from DNA-mediated gene transfer experiments. The results presented indicate that a mixed isotype class II molecule can be expressed naturally. PMID:2647893

In Vivo Cytometry of Antigen-Specific T Cells Using 19F MRI

PubMed Central

Srinivas, Mangala; Turner, Michael S.; Janjic, Jelena M.; Morel, Penelope A.; Laidlaw, David H.; Ahrens, Eric T.

2009-01-01

Noninvasive methods to image the trafficking of phenotypically defined immune cells are paramount as we attempt to understand adaptive immunity. A 19F MRI-based methodology for tracking and quantifying cells of a defined phenotype is presented. These methods were applied to a murine inflammation model using antigen-specific T cells. The T cells that were intracellularly labeled ex vivo with a perfluoropolyether (PFPE) nanoemulsion and cells were transferred to a host receiving a localized inoculation of antigen. Longitudinal 19F MRI over 21 days revealed a dynamic accumulation and clearance of T cells in the lymph node (LN) draining the antigen. The apparent T-cell numbers were calculated in the LN from the time-lapse 19F MRI data. The effect of in vivo T-cell division on the 19F MRI cell quantification accuracy was investigated using fluorescence assays. Overall, in vivo cytometry using PFPE labeling and 19F MRI is broadly applicable to studies of whole-body cell biodistribution. PMID:19585593

Clonal deletion of thymocytes can occur in the cortex with no involvement of the medulla

PubMed Central

McCaughtry, Tom M.; Baldwin, Troy A.; Wilken, Matthew S.; Hogquist, Kristin A.

2008-01-01

The thymic medulla is generally held to be a specialized environment for negative selection. However, many self-reactive thymocytes first encounter ubiquitous self-antigens in the cortex. Cortical epithelial cells are vital for positive selection, but whether such cells can also promote negative selection is controversial. We used the HYcd4 model, where T cell receptor for antigen (TCR) expression is appropriately timed and a ubiquitous self-antigen drives clonal deletion in male mice. We demonstrated unambiguously that this deletion event occurs in the thymic cortex. However, the kinetics in vivo indicated that apoptosis was activated asynchronously relative to TCR activation. We found that radioresistant antigen-presenting cells and, specifically, cortical epithelial cells do not efficiently induce apoptosis, although they do cause TCR activation. Rather, thymocytes undergoing clonal deletion were preferentially associated with rare CD11c+ cortical dendritic cells, and elimination of such cells impaired deletion. PMID:18936237

Immunity to Intracellular Salmonella Depends on Surface-associated Antigens

PubMed Central

Claudi, Beatrice; Mazé, Alain; Schemmer, Anne K.; Kirchhoff, Dennis; Schmidt, Alexander; Burton, Neil; Bumann, Dirk

2012-01-01

Invasive Salmonella infection is an important health problem that is worsening because of rising antimicrobial resistance and changing Salmonella serovar spectrum. Novel vaccines with broad serovar coverage are needed, but suitable protective antigens remain largely unknown. Here, we tested 37 broadly conserved Salmonella antigens in a mouse typhoid fever model, and identified antigen candidates that conferred partial protection against lethal disease. Antigen properties such as high in vivo abundance or immunodominance in convalescent individuals were not required for protectivity, but all promising antigen candidates were associated with the Salmonella surface. Surprisingly, this was not due to superior immunogenicity of surface antigens compared to internal antigens as had been suggested by previous studies and novel findings for CD4 T cell responses to model antigens. Confocal microscopy of infected tissues revealed that many live Salmonella resided alone in infected host macrophages with no damaged Salmonella releasing internal antigens in their vicinity. In the absence of accessible internal antigens, detection of these infected cells might require CD4 T cell recognition of Salmonella surface-associated antigens that could be processed and presented even from intact Salmonella. In conclusion, our findings might pave the way for development of an efficacious Salmonella vaccine with broad serovar coverage, and suggest a similar crucial role of surface antigens for immunity to both extracellular and intracellular pathogens. PMID:23093937

Unusual antigen presentation offers new insight into HIV vaccine design.

PubMed

McMichael, Andrew J; Picker, Louis J

2017-06-01

Recent findings with a rhesus monkey cytomegalovirus based simian immunodeficiency virus vaccine have identified strong CD8+ T cell responses that are restricted by MHC-E. Also mycobacteria specific CD8+ T cells, that are MHC-E restricted, have been identified. MHC-E therefore can present a wide range of epitope peptides to CD8+ T cells, alongside its well defined role in presenting a conserved MHC-class I signal peptide to the NKG2A/C-CD94 receptor on natural killer cells. Here we explore the antigen processing pathways involved in these atypical T cell responses. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Jurkat 76 based triple parameter reporter system to evaluate TCR functions and adoptive T cell strategies.

PubMed

Rosskopf, Sandra; Leitner, Judith; Paster, Wolfgang; Morton, Laura T; Hagedoorn, Renate S; Steinberger, Peter; Heemskerk, Mirjam H M

2018-04-03

Adoptive T cell therapy using TCR transgenic autologous T cells has shown great potential for the treatment of tumor patients. Thorough characterization of genetically reprogrammed T cells is necessary to optimize treatment success. Here, we describe the generation of triple parameter reporter T cells based on the Jurkat 76 T cell line for the evaluation of TCR and chimeric antigen receptor functions as well as adoptive T cell strategies. This Jurkat subline is devoid of endogenous TCR alpha and TCR beta chains, thereby circumventing the problem of TCR miss-pairing and unexpected specificities. The resultant reporter cells allow simultaneous determination of the activity of the transcription factors NF-κB, NFAT and AP-1 that play key roles in T cell activation. Human TCRs directed against tumor and virus antigens were introduced and reporter responses were determined using tumor cell lines endogenously expressing the antigens of interest or via addition of antigenic peptides. Finally, we demonstrate that coexpression of adhesion molecules like CD2 and CD226 as well as CD28 chimeric receptors represents an effective strategy to augment the response of TCR-transgenic reporters to cells presenting cognate antigens.

Intestinal Epithelial Cells Modulate Antigen-Presenting Cell Responses to Bacterial DNA

PubMed Central

Campeau, J. L.; Salim, S. Y.; Albert, E. J.; Hotte, N.

2012-01-01

Intestinal epithelial cells and antigen-presenting cells orchestrate mucosal innate immunity. This study investigated the role of bacterial DNA in modulating epithelial and bone marrow-derived antigen-presenting cells (BM-APCs) and subsequent T-lymphocyte responses. Murine MODE-K epithelial cells and BM-APCs were treated with DNA from either Bifidobacterium breve or Salmonella enterica serovar Dublin directly and under coculture conditions with CD4+ T cells. Apical stimulation of MODE-K cells with S. Dublin DNA enhanced secretion of cytokines from underlying BM-APCs and induced interleukin-17 (IL-17) and gamma interferon (IFN-γ) secretion from CD4+ T cells. Bacterial DNA isolated from either strain induced maturation and increased cytokine secretion from BM-APCs. Conditioned medium from S. Dublin-treated MODE-K cells elicited an increase in cytokine secretion similar to that seen for S. Dublin DNA. Treatment of conditioned medium from MODE-K cells with RNase and protease prevented the S. Dublin-induced increased cytokine secretion. Oral feeding of mice with B. breve DNA resulted in enhanced levels of colonic IL-10 and transforming growth factor β (TGFβ) compared with what was seen for mice treated with S. Dublin DNA. In contrast, feeding mice with S. Dublin DNA increased levels of colonic IL-17 and IL-12p70. T cells from S. Dublin DNA-treated mice secreted high levels of IL-12 and IFN-γ compared to controls and B. breve DNA-treated mice. These results demonstrate that intestinal epithelial cells are able to modulate subsequent antigen-presenting and T-cell responses to bacterial DNA with pathogenic but not commensal bacterial DNA inducing effector CD4+ T lymphocytes. PMID:22615241

Immunological Characterization of Whole Tumour Lysate-Loaded Dendritic Cells for Cancer Immunotherapy

PubMed Central

Ottobrini, Luisa; Biasin, Mara; Borelli, Manuela; Lucignani, Giovanni; Trabattoni, Daria; Clerici, Mario

2016-01-01

Introduction Dendritic cells play a key role as initiators of T-cell responses, and even if tumour antigen-loaded dendritic cells can induce anti-tumour responses, their efficacy has been questioned, suggesting a need to enhance immunization strategies. Matherials & Methods We focused on the characterization of bone marrow-derived dendritic cells pulsed with whole tumour lysate (TAA-DC), as a source of known and unknown antigens, in a mouse model of breast cancer (MMTV-Ras). Dendritic cells were evaluated for antigen uptake and for the expression of MHC class I/II and costimulatory molecules and markers associated with maturation. Results Results showed that antigen-loaded dendritic cells are characterized by a phenotypically semi-mature/mature profile and by the upregulation of genes involved in antigen presentation and T-cell priming. Activated dendritic cells stimulated T-cell proliferation and induced the production of high concentrations of IL-12p70 and IFN-γ but only low levels of IL-10, indicating their ability to elicit a TH1-immune response. Furthermore, administration of Antigen loaded-Dendritic Cells in MMTV-Ras mice evoked a strong anti-tumour response in vivo as demonstrated by a general activation of immunocompetent cells and the release of TH1 cytokines. Conclusion Data herein could be useful in the design of antitumoral DC-based therapies, showing a specific activation of immune system against breast cancer. PMID:26795765

CELLS INVOLVED IN THE IMMUNE RESPONSE

PubMed Central

Abdou, Nabih I.; Richter, Maxwell

1969-01-01

Rabbits were made immunologically tolerant to either human serum albumin or bovine gamma globulin by the neonatal administration of antigen. At 10 wk of age, they were challenged with the tolerogenic antigen and found to be non-responsive. However, these tolerant rabbits could respond with humoral antibody formation directed toward the tolerogenic antigen if they were treated with normal, allogeneic bone marrow or bone marrow obtained from a rabbit made tolerant toward a different antigen. They were incapable of responding if they were given bone marrow obtained from a rabbit previously made tolerant to the tolerogenic antigen. Irradiated rabbits were unable to respond if treated with tolerant bone marrow, but could respond well if given normal bone marrow. Since it has previously been demonstrated that the antibody-forming cell, in an irradiated recipient of allogeneic bone marrow, is of recipient and not donor origin, the data presented strongly indicate that the unresponsive cell in the immunologically tolerant rabbit is the antigen-reactive cell. PMID:4183777

Saponins from soy bean and mung bean inhibit the antigen specific activation of helper T cells by blocking cell cycle progression.

PubMed

Lee, Suk Jun; Bae, Joonbeom; Kim, Sunhee; Jeong, Seonah; Choi, Chang-Yong; Choi, Sang-Pil; Kim, Hyun-Sook; Jung, Woon-Won; Imm, Jee-Young; Kim, Sae Hun; Chun, Taehoon

2013-02-01

Treatment of helper T (Th) cells with saponins from soy bean and mung bean prevented their activation by inhibiting cell proliferation and cytokine secretion. However, the saponins did not affect the expression of major histocompatibility complex class II (A(b)) and co-stimulatory molecule (CD86) on professional antigen-presenting cells. Instead, the saponins directly inhibited Th cell proliferation by blocking the G(1) to S phase cell cycle transition. Moreover, blocking of the cell cycle by the saponins was achieved by decreased expression of cyclin D1 and cyclin E, and constitutive expression of p27(KIP1). Saponins also increased stability of p27(KIP1) in Th cells after antigenic stimulation.

Enhanced B Cell Alloantigen Presentation and Its Epigenetic Dysregulation in Liver Transplant Rejection.

PubMed

Ningappa, M; Ashokkumar, C; Higgs, B W; Sun, Q; Jaffe, R; Mazariegos, G; Li, D; Weeks, D E; Subramaniam, S; Ferrell, R; Hakonarson, H; Sindhi, R

2016-02-01

T cell suppression prevents acute cellular rejection but causes life-threatening infections and malignancies. Previously, liver transplant (LTx) rejection in children was associated with the single-nucleotide polymorphism (SNP) rs9296068 upstream of the HLA-DOA gene. HLA-DOA inhibits B cell presentation of antigen, a potentially novel antirejection drug target. Using archived samples from 122 white pediatric LTx patients (including 77 described previously), we confirmed the association between rs9296068 and LTx rejection (p = 0.001, odds ratio [OR] 2.55). Next-generation sequencing revealed that the putative transcription factor (CCCTC binding factor [CTCF]) binding SNP locus rs2395304, in linkage disequilibrium with rs9296068 (D' 0.578, r(2) = 0.4), is also associated with LTx rejection (p = 0.008, OR 2.34). Furthermore, LTx rejection is associated with enhanced B cell presentation of donor antigen relative to HLA-nonidentical antigen in a novel cell-based assay and with a downregulated HLA-DOA gene in a subset of these children. In lymphoblastoid B (Raji) cells, rs2395304 coimmunoprecipitates with CTCF, and CTCF knockdown with morpholino antisense oligonucleotides enhances alloantigen presentation and downregulates the HLA-DOA gene, reproducing observations made with HLA-DOA knockdown and clinical rejection. Alloantigen presentation is suppressed by inhibitors of methylation and histone deacetylation, reproducing observations made during resolution of rejection. Enhanced donor antigen presentation by B cells and its epigenetic dysregulation via the HLA-DOA gene represent novel opportunities for surveillance and treatment of transplant rejection. © Copyright 2015 The American Society of Transplantation and the American Society of Transplant Surgeons.

Mice completely lacking immunoproteasomes display major alterations in antigen presentation

PubMed Central

Kincaid, Eleanor Z; Che, Jenny W; York, Ian; Escobar, Hernando; Reyes-Vargas, Eduardo; Delgado, Julio C.; Welsh, Raymond M; Karow, Margaret L.; Murphy, Andrew J.; Valenzuela, David M.; Yancopoulos, George D.; Rock, Kenneth L

2011-01-01

The importance of immunoproteasomes to antigen presentation has been unclear because animals totally lacking immunoproteasomes have not been previously developed. Here we show that dendritic cells from mice lacking the three immunoproteasome catalytic subunits display defects in presenting multiple major histocompatability (MHC) class I epitopes. During viral infection in vivo, the presentation of a majority of MHC class I epitopes is markedly reduced in immunoproteasome-deficient animals, while presentation of MHC class II peptides is unaffected. By mass spectrometry the repertoire of MHC class I-presented peptides is ~50% different and these differences are sufficient to stimulate robust transplant rejection of wild type cells in mutant mice. These results indicate that immunoproteasomes play a much more important role in antigen presentation than previously thought. PMID:22197977

An evolving new paradigm: endothelial cells – conditional innate immune cells

PubMed Central

2013-01-01

Endothelial cells (ECs) are a heterogeneous population that fulfills many physiological processes. ECs also actively participate in both innate and adaptive immune responses. ECs are one of the first cell types to detect foreign pathogens and endogenous metabolite-related danger signals in the bloodstream, in which ECs function as danger signal sensors. Treatment with lipopolysaccharide activates ECs, causing the production of pro-inflammatory cytokines and chemokines, which amplify the immune response by recruiting immune cells. Thus, ECs function as immune/inflammation effectors and immune cell mobilizers. ECs also induce cytokine production by immune cells, in which ECs function as immune regulators either by activating or suppressing immune cell function. In addition, under certain conditions, ECs can serve as antigen presenting cells (antigen presenters) by expressing both MHC I and II molecules and presenting endothelial antigens to T cells. These facts along with the new concept of endothelial plasticity suggest that ECs are dynamic cells that respond to extracellular environmental changes and play a meaningful role in immune system function. Based on these novel EC functions, we propose a new paradigm that ECs are conditional innate immune cells. This paradigm provides a novel insight into the functions of ECs in inflammatory/immune pathologies. PMID:23965413

An evolving new paradigm: endothelial cells--conditional innate immune cells.

PubMed

Mai, Jietang; Virtue, Anthony; Shen, Jerry; Wang, Hong; Yang, Xiao-Feng

2013-08-22

Endothelial cells (ECs) are a heterogeneous population that fulfills many physiological processes. ECs also actively participate in both innate and adaptive immune responses. ECs are one of the first cell types to detect foreign pathogens and endogenous metabolite-related danger signals in the bloodstream, in which ECs function as danger signal sensors. Treatment with lipopolysaccharide activates ECs, causing the production of pro-inflammatory cytokines and chemokines, which amplify the immune response by recruiting immune cells. Thus, ECs function as immune/inflammation effectors and immune cell mobilizers. ECs also induce cytokine production by immune cells, in which ECs function as immune regulators either by activating or suppressing immune cell function. In addition, under certain conditions, ECs can serve as antigen presenting cells (antigen presenters) by expressing both MHC I and II molecules and presenting endothelial antigens to T cells. These facts along with the new concept of endothelial plasticity suggest that ECs are dynamic cells that respond to extracellular environmental changes and play a meaningful role in immune system function. Based on these novel EC functions, we propose a new paradigm that ECs are conditional innate immune cells. This paradigm provides a novel insight into the functions of ECs in inflammatory/immune pathologies.

Dysfunctional BLK in common variable immunodeficiency perturbs B-cell proliferation and ability to elicit antigen-specific CD4+ T-cell help.

PubMed

Compeer, Ewoud B; Janssen, Willemijn; van Royen-Kerkhof, Annet; van Gijn, Marielle; van Montfrans, Joris M; Boes, Marianne

2015-05-10

Common Variable Immunodeficiency (CVID) is the most prevalent primary antibody deficiency, and characterized by defective generation of high-affinity antibodies. Patients have therefore increased risk to recurrent infections of the respiratory and intestinal tract. Development of high-affinity antigen-specific antibodies involves two key actions of B-cell receptors (BCR): transmembrane signaling through BCR-complexes to induce B-cell differentiation and proliferation, and BCR-mediated antigen internalization for class-II MHC-mediated presentation to acquire antigen-specific CD4(+) T-cell help.We identified a variant (L3P) in the B-lymphoid tyrosine kinase (BLK) gene of 2 related CVID-patients, which was absent in healthy relatives. BLK belongs to the Src-kinases family and involved in BCR-signaling. Here, we sought to clarify BLK function in healthy human B-cells and its association to CVID.BLK expression was comparable in patient and healthy B-cells. Functional analysis of L3P-BLK showed reduced BCR crosslinking-induced Syk phosphorylation and proliferation, in both primary B-cells and B-LCLs. B-cells expressing L3P-BLK showed accelerated destruction of BCR-internalized antigen and reduced ability to elicit CD40L-expression on antigen-specific CD4(+) T-cells.In conclusion, we found a novel BLK gene variant in CVID-patients that causes suppressed B-cell proliferation and reduced ability of B-cells to elicit antigen-specific CD4(+) T-cell responses. Both these mechanisms may contribute to hypogammaglobulinemia in CVID-patients.

Antigen-presenting cells and materno-fetal tolerance: an emerging role for dendritic cells.

PubMed

Laskarin, Gordana; Kämmerer, Ulrike; Rukavina, Daniel; Thomson, Angus W; Fernandez, Nelson; Blois, Sandra M

2007-09-01

During pregnancy, a delicate balance of innate and adaptive immune responses at the maternal-fetal interface promotes survival of the semi-allogeneic embryo and, at the same time, allows effective immunity to protect the mother from environmental pathogens. As in other tissues, antigen handling and processing in the decidualized endometrium constitutes a primary event in the onset of immune responses and is therefore likely to determine their stimulatory or tolerogenic nature. Maternal antigen-presenting cells [macrophages and dendritic cells (DCs)] are scattered throughout the decidualized endometrium during all stages of pregnancy and appear to be important players in this feto-maternal immune adjustment. This review focuses on the characterization of decidual macrophages and DCs, as well as their involvement in cell-cell interactions within the decidual leukocyte network, which are likely to influence uterine and placental homeostasis as well as the local maternal immune responses to the fetus during pregnancy.

Caspase-3 Is Transiently Activated without Cell Death during Early Antigen Driven Expansion of CD8+ T Cells In Vivo

PubMed Central

McComb, Scott; Mulligan, Rebecca; Sad, Subash

2010-01-01

Background CD8+ T cell responses develop rapidly during infection and are swiftly reduced during contraction, wherein >90% of primed CD8+ T cells are eliminated. The role of apoptotic mechanisms in controlling this rapid proliferation and contraction of CD8+ T cells remains unclear. Surprisingly, evidence has shown non-apoptotic activation of caspase-3 to occur during in vitro T-cell proliferation, but the relevance of these mechanisms to in vivo CD8+ T cell responses has yet to be examined. Methods and Findings We have evaluated the activity of caspase-3, a key downstream inducer of apoptosis, throughout the entirety of a CD8+ T cell response. We utilized two infection models that differ in the intensity, onset and duration of antigen-presentation and inflammation. Expression of cleaved caspase-3 in antigen specific CD8+ T cells was coupled to the timing and strength of antigen presentation in lymphoid organs. We also observed coordinated activation of additional canonical apoptotic markers, including phosphatidylserine exposure. Limiting dilution analysis directly showed that in the presence of IL7, very little cell death occurred in both caspase-3hi and caspase-3low CD8+ T cells. The expression of active caspase-3 peaked before effector phenotype (CD62Llow) CD8+ T cells emerged, and was undetectable in effector-phenotype cells. In addition, OVA-specific CD8+ cells remained active caspase-3low throughout the contraction phase. Conclusions Our results specifically implicate antigen and not inflammation in driving activation of apoptotic mechanisms without cell death in proliferating CD8+ T cells. Furthermore, the contraction of CD8+ T cell response following expansion is likely not mediated by the key downstream apoptosis inducer, caspase-3. PMID:21203525

A Positive Control for Detection of Functional CD4 T Cells in PBMC: The CPI Pool.

PubMed

Schiller, Annemarie; Zhang, Ting; Li, Ruliang; Duechting, Andrea; Sundararaman, Srividya; Przybyla, Anna; Kuerten, Stefanie; Lehmann, Paul V

2017-12-07

Testing of peripheral blood mononuclear cells (PBMC) for immune monitoring purposes requires verification of their functionality. This is of particular concern when the PBMC have been shipped or stored for prolonged periods of time. While the CEF (Cytomegalo-, Epstein-Barr and Flu-virus) peptide pool has become the gold standard for testing CD8 cell functionality, a positive control for CD4 cells is so far lacking. The latter ideally consists of proteins so as to control for the functionality of the antigen processing and presentation compartments, as well. Aiming to generate a positive control for CD4 cells, we first selected 12 protein antigens from infectious/environmental organisms that are ubiquitous: Varicella, Influenza, Parainfluenza, Mumps, Cytomegalovirus, Streptococcus , Mycoplasma , Lactobacillus , Neisseria , Candida , Rubella, and Measles. Of these antigens, three were found to elicited interferon (IFN)-γ-producing CD4 cells in the majority of human test subjects: inactivated cytomegalo-, parainfluenza-, and influenza virions (CPI). While individually none of these three antigens triggered a recall response in all donors, the pool of the three (the 'CPI pool'), did. One hundred percent of 245 human donors tested were found to be CPI positive, including Caucasians, Asians, and African-Americans. Therefore, the CPI pool appears to be suitable to serve as universal positive control for verifying the functionality of CD4 and of antigen presenting cells.

Complex antigen presentation pathway for an HLA-A*0201-restricted epitope from Chikungunya 6K protein

PubMed Central

Lemonnier, François A.; Esteban, Mariano

2017-01-01

Background The adaptive cytotoxic T lymphocyte (CTL)-mediated immune response is critical for clearance of many viral infections. These CTL recognize naturally processed short viral antigenic peptides bound to human leukocyte antigen (HLA) class I molecules on the surface of infected cells. This specific recognition allows the killing of virus-infected cells. The T cell immune T cell response to Chikungunya virus (CHIKV), a mosquito-borne Alphavirus of the Togaviridae family responsible for severe musculoskeletal disorders, has not been fully defined; nonetheless, the importance of HLA class I-restricted immune response in this virus has been hypothesized. Methodology/Principal findings By infection of HLA-A*0201-transgenic mice with a recombinant vaccinia virus that encodes the CHIKV structural polyprotein (rVACV-CHIKV), we identified the first human T cell epitopes from CHIKV. These three novel 6K transmembrane protein-derived epitopes are presented by the common HLA class I molecule, HLA-A*0201. One of these epitopes is processed and presented via a complex pathway that involves proteases from different subcellular locations. Specific chemical inhibitors blocked these events in rVACV-CHIKV-infected cells. Conclusions/Significance Our data have implications not only for the identification of novel Alphavirus and Togaviridae antiviral CTL responses, but also for analyzing presentation of antigen from viruses of different families and orders that use host proteinases to generate their mature envelope proteins. PMID:29084215

CD1a presentation of endogenous antigens by group 2 innate lymphoid cells.

PubMed

Hardman, Clare S; Chen, Yi-Ling; Salimi, Maryam; Jarrett, Rachael; Johnson, David; Järvinen, Valtteri J; Owens, Raymond J; Repapi, Emmanouela; Cousins, David J; Barlow, Jillian L; McKenzie, Andrew N J; Ogg, Graham

2017-12-22

Group 2 innate lymphoid cells (ILC2) are effectors of barrier immunity, with roles in infection, wound healing, and allergy. A proportion of ILC2 express MHCII (major histocompatibility complex II) and are capable of presenting peptide antigens to T cells and amplifying the subsequent adaptive immune response. Recent studies have highlighted the importance of CD1a-reactive T cells in allergy and infection, activated by the presentation of endogenous neolipid antigens and bacterial components. Using a human skin challenge model, we unexpectedly show that human skin-derived ILC2 can express CD1a and are capable of presenting endogenous antigens to T cells. CD1a expression is up-regulated by TSLP (thymic stromal lymphopoietin) at levels observed in the skin of patients with atopic dermatitis, and the response is dependent on PLA2G4A. Furthermore, this pathway is used to sense Staphylococcus aureus by promoting Toll-like receptor-dependent CD1a-reactive T cell responses to endogenous ligands. These findings define a previously unrecognized role for ILC2 in lipid surveillance and identify shared pathways of CD1a- and PLA2G4A-dependent ILC2 inflammation amenable to therapeutic intervention. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Complex antigen presentation pathway for an HLA-A*0201-restricted epitope from Chikungunya 6K protein.

PubMed

Lorente, Elena; Barriga, Alejandro; García-Arriaza, Juan; Lemonnier, François A; Esteban, Mariano; López, Daniel

2017-10-01

The adaptive cytotoxic T lymphocyte (CTL)-mediated immune response is critical for clearance of many viral infections. These CTL recognize naturally processed short viral antigenic peptides bound to human leukocyte antigen (HLA) class I molecules on the surface of infected cells. This specific recognition allows the killing of virus-infected cells. The T cell immune T cell response to Chikungunya virus (CHIKV), a mosquito-borne Alphavirus of the Togaviridae family responsible for severe musculoskeletal disorders, has not been fully defined; nonetheless, the importance of HLA class I-restricted immune response in this virus has been hypothesized. By infection of HLA-A*0201-transgenic mice with a recombinant vaccinia virus that encodes the CHIKV structural polyprotein (rVACV-CHIKV), we identified the first human T cell epitopes from CHIKV. These three novel 6K transmembrane protein-derived epitopes are presented by the common HLA class I molecule, HLA-A*0201. One of these epitopes is processed and presented via a complex pathway that involves proteases from different subcellular locations. Specific chemical inhibitors blocked these events in rVACV-CHIKV-infected cells. Our data have implications not only for the identification of novel Alphavirus and Togaviridae antiviral CTL responses, but also for analyzing presentation of antigen from viruses of different families and orders that use host proteinases to generate their mature envelope proteins.

A lipid-based nano-regulator for cancer immunotherapy (Conference Presentation)

NASA Astrophysics Data System (ADS)

Qian, Yuan; Qiao, Sha; Zhang, Zhihong

2017-02-01

In the application of nanotechnology in cancer immunotherapy, antigen presenting cells (APCs, dendritic cells and macrophages) are preferable target due to their endocytic capacity and suppressed phenotype. Recently, we developed a lipid-based core-shell nanocarrier, which is stabilized by changeable fusion peptides and possesses a sub-30 diameter. With the different peptides, the nanoparticles (NPs) could either target to dendritic cells (DCs) in lymph nodes (LNs) or tumor associated macrophages (TAMs) in tumor environment. After subcutaneous injection, the NPs could targeted deliver the encapsulated antigen peptides (APs) and adjuvants (CpG-ODN) to dendritic cells in LNs, and lead to the antigen presenting and activation of cytotoxic T lymphocytes against tumor. In other case, after systemic administration, the immune regulatory molecules were carried by NPs and targeting delivered to specific immunocytes in tumor microenvironment resulting in the immunosuppressive state broken and tumor growth inhibition.

New Insights into the Mechanism of Inhibition of p53 by Simian Virus 40 Large T Antigen

PubMed Central

Sheppard, Hilary M.; Corneillie, Siska I.; Espiritu, Christine; Gatti, Andrea; Liu, Xuan

1999-01-01

Simian virus 40 (SV40) large tumor antigen (T antigen) has been shown to inhibit p53-dependent transcription by preventing p53 from binding to its cognate cis element. Data presented in this report provide the first direct functional evidence that T antigen, under certain conditions, may also repress p53-dependent transcription by a mechanism in which the transactivation domain of p53 is abrogated while DNA binding is unaffected. Specifically, p53 purified as a complex with T antigen from mouse cells was found to bind DNA as a transcriptionally inactive intact complex, while that purified from human cells was found to bind DNA independently of T antigen and could activate p53-dependent transcription. This difference in activity may be dependent on a different interaction of T antigen with mouse and human p53 and, in addition, on the presence of super T, which is found only in transformed rodent cells. These results suggest that subtle yet important differences exist between the inhibition of p53 by T antigen in mouse and human cells. The implications of this finding with respect to SV40-associated malignancies are discussed. PMID:10082540

Multiphoton microscopy of antigen presenting cells in experimental cancer therapies

NASA Astrophysics Data System (ADS)

Watkins, Simon C.; Papworth, Glenn D.; Spencer, Lori A.; Larregina, Adriana T.; Hackstein, Holger

2002-06-01

The absence of effective conventional therapy for most cancer patients justifies the application of novel, experimental approaches. One alternative to conventional cytotoxic agents is a more defined molecular approach for cancer immune treatment; promotion of the immune system specifically to target and eliminate tumor cells on the basis of expression of tumor-associated antigens (TAA). TAA could be presented to T-cells by professional antigen-presenting cells (APC) that generate a more efficient and effective anti-tumor immune response. In fact, it has been well documented that dendritic cells, the most immunologically potent APC, are capable of recognizing, processing and presenting TAA, in turn initiating a specific antitumor immune response. Results from several laboratories and clinical trials suggested significant but still limited efficacy of TAA-pulsed dendritic cells administered to tumor-bearing hosts. Following such delivery, it is fundamentally necessary to dynamically assess cell abundance within the microenvironment of the tumor in the presence of the appropriate therapeutic agent. Multiphoton microscopy was used to assess the trafficking of pulsed dendritic cells and other APC in skin, lymph nodes and brain of several animal tumor models, following different routes of administration.

A new approach to gene therapy using Sleeping Beauty to genetically modify clinical-grade T cells to target CD19

PubMed Central

Singh, Harjeet; Huls, Helen; Cooper, Laurence JN

2014-01-01

Summary The advent of efficient approaches to the genetic modification of T cells has provided investigators with clinically appealing approaches to improve the potency of tumor-specific clinical grade T cells. For example, gene therapy has been successfully used to enforce expression of chimeric antigen receptors (CAR) that provide T cells with ability to directly recognize tumor-associated antigens without the need for presentation by human leukocyte antigen. Gene transfer of CARs can be undertaken using viral-based and non-viral approaches. We have advanced DNA vectors derived from the Sleeping Beauty (SB) system to avoid the expense and manufacturing difficulty associated with transducing T cells with recombinant viral vectors. After electroporation, the transposon/transposase system improves the efficiency of integration of plasmids used to express CAR and other transgenes in T cells. The SB system combined with artificial antigen-presenting cells (aAPC) can selectively propagate and thus retrieve CAR+ T cells suitable for human application. This review describes the translation of the SB system and aAPC for use in clinical trials and highlights how a nimble and cost-effective approach to developing genetically modified T cells can be used to implement clinical trials infusing next-generation T cells with improved therapeutic potential. PMID:24329797

Identification and characterization of polyclonal αβ T cells with dendritic cell properties

PubMed Central

Kuka, Mirela; Munitic, Ivana; Ashwell, Jonathan D.

2012-01-01

An efficient immune response requires coordination between innate and adaptive immunity, which act through cells different in origin and function. Here we report the identification of thymus-derived αβ TCR+ cells that express CD11c and MHC class II, and require FLT3L for development (TDC). TDC express genes heretofore found uniquely in T cells or DC, as well as a distinctive signature of cytotoxicity-related genes. Unlike other innate T cell subsets, TDC have a polyclonal TCR repertoire andrespond to cognate antigens. However, they differ from conventional T cells in that they do not require help from antigen-presenting cells, respond to TLR-mediated stimulation by producing IL-12 and process and present antigen. The physiologic relevance of TDC, found in mice and humans, is still under investigation, but the fact that they combine key features of T and DC cells suggests that they provide a bridge between the innate and adaptive immune systems. PMID:23187623

Use of human antigen presenting cell gene array profiling to examine the effect of human T-cell leukemia virus type 1 Tax on primary human dendritic cells.

PubMed

Ahuja, Jaya; Kampani, Karan; Datta, Suman; Wigdahl, Brian; Flaig, Katherine E; Jain, Pooja

2006-02-01

Human T-cell leukemia virus type 1 (HTLV-1) is etiologically linked to adult T-cell leukemia and a progressive demyelinating disorder termed HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). One of the most striking features of the immune response in HAM/TSP centers on the expansion of HTLV-1-specific CD8(+) cytotoxic T lymphocyte (CTL) compartment in the peripheral blood and cerebrospinal fluid. More than 90% of the HTLV-1-specific CTLs are directed against the viral Tax (11-19) peptide implying that Tax is available for immune recognition by antigen presenting cells, such as dendritic cells (DCs). DCs obtained from HAM/TSP patients have been shown to be infected with HTLV-1 and exhibit rapid maturation. Therefore, we hypothesized that presentation of Tax peptides by activated DCs to naIve CD8(+) T cells may play an important role in the induction of a Tax-specific CTL response and neurologic dysfunction. In this study, a pathway-specific antigen presenting cell gene array was used to study transcriptional changes induced by exposure of monocyte-derived DCs to extracellular HTLV-1 Tax protein. Approximately 100 genes were differentially expressed including genes encoding toll-like receptors, cell surface receptors, proteins involved in antigen uptake and presentation and adhesion molecules. The differential regulation of chemokines and cytokines characteristic of functional DC activation was also observed by the gene array analyses. Furthermore, the expression pattern of signal transduction genes was also significantly altered. These results have suggested that Tax-mediated DC gene regulation might play a critical role in cellular activation and the mechanisms resulting in HTLV-1-induced disease.

Antigen-capturing nanoparticles improve the abscopal effect and cancer immunotherapy

NASA Astrophysics Data System (ADS)

Min, Yuanzeng; Roche, Kyle C.; Tian, Shaomin; Eblan, Michael J.; McKinnon, Karen P.; Caster, Joseph M.; Chai, Shengjie; Herring, Laura E.; Zhang, Longzhen; Zhang, Tian; Desimone, Joseph M.; Tepper, Joel E.; Vincent, Benjamin G.; Serody, Jonathan S.; Wang, Andrew Z.

2017-09-01

Immunotherapy holds tremendous promise for improving cancer treatment. To administer radiotherapy with immunotherapy has been shown to improve immune responses and can elicit the 'abscopal effect'. Unfortunately, response rates for this strategy remain low. Herein we report an improved cancer immunotherapy approach that utilizes antigen-capturing nanoparticles (AC-NPs). We engineered several AC-NP formulations and demonstrated that the set of protein antigens captured by each AC-NP formulation is dependent on the NP surface properties. We showed that AC-NPs deliver tumour-specific proteins to antigen-presenting cells (APCs) and significantly improve the efficacy of αPD-1 (anti-programmed cell death 1) treatment using the B16F10 melanoma model, generating up to a 20% cure rate compared with 0% without AC-NPs. Mechanistic studies revealed that AC-NPs induced an expansion of CD8+ cytotoxic T cells and increased both CD4+T/Treg and CD8+T/Treg ratios (Treg, regulatory T cells). Our work presents a novel strategy to improve cancer immunotherapy with nanotechnology.

Rheumatoid arthritis vaccine therapies: perspectives and lessons from therapeutic ligand epitope antigen presentation system vaccines for models of rheumatoid arthritis

PubMed Central

Rosenthal, Kenneth S.; Mikecz, Katalin; Steiner, Harold L.; Glant, Tibor T.; Finnegan, Alison; Carambula, Roy E.; Zimmerman, Daniel H.

2016-01-01

The current status of therapeutic vaccines for autoimmune diseases is reviewed with rheumatoid arthritis as the focus. Therapeutic vaccines for autoimmune diseases must regulate or subdue responses to common self-antigens. Ideally, such a vaccine would initiate an antigen-specific modulation of the T-cell immune response that drives the inflammatory disease. Appropriate animal models and types of T helper cells and signature cytokine responses that drive autoimmune disease are also discussed. Interpretation of these animal models must be done cautiously because the means of initiation, autoantigens, and even the signature cytokine and T helper cell (Th1 or Th17) responses that are involved in the disease may differ significantly from those in humans. We describe ligand epitope antigen presentation system vaccine modulation of T-cell autoimmune responses as a strategy for the design of therapeutic vaccines for rheumatoid arthritis, which may also be effective in other autoimmune conditions. PMID:25787143

Use of lectin-functionalized particles for oral immunotherapy

PubMed Central

Diesner, Susanne C; Wang, Xue-Yan; Jensen-Jarolim, Erika; Untersmayr, Eva; Gabor, Franz

2013-01-01

Immunotherapy, in recent times, has found its application in a variety of immunologically mediated diseases. Oral immunotherapy may not only increase patient compliance but may, in particular, also induce both systemic as well as mucosal immune responses, due to mucosal application of active agents. To improve the bioavailability and to trigger strong immunological responses, recent research projects focused on the encapsulation of drugs and antigens into polymer particles. These particles protect the loaded antigen from the harsh conditions in the GI tract. Furthermore, modification of the surface of particles by the use of lectins, such as Aleuria aurantia lectin, wheatgerm agglutinin or Ulex europaeus-I, enhances the binding to epithelial cells, in particular to membranous cells, of the mucosa-associated lymphoid tissue. Membranous cell-specific targeting leads to an improved transepithelial transport of the particle carriers. Thus, enhanced uptake and presentation of the encapsulated antigen by antigen-presenting cells favor strong systemic, but also local, mucosal immune responses. PMID:22834202

Serine Proteases Enhance Immunogenic Antigen Presentation on Lung Cancer Cells

PubMed Central

Peters, Haley L.; Tripathi, Satyendra C.; Kerros, Celine; Katayama, Hiroyuki; Garber, Haven R.; St. John, Lisa S.; Federico, Lorenzo; Meraz, Ismail M.; Roth, Jack A.; Sepesi, Boris; Majidi, Mourad; Ruisaard, Kathryn; Clise-Dwyer, Karen; Roszik, Jason; Gibbons, Don L.; Heymach, John V.; Swisher, Stephen G.; Bernantchez, Chantale; Alatrash, Gheath; Hanash, Samir; Molldrem, Jeffrey J.

2017-01-01

Immunotherapies targeting immune checkpoints have proven efficacious in reducing the burden of lung cancer in patients; however, the antigenic targets of these re-invigorated T cells remain poorly defined. Lung cancer tumors contain tumor-associated macrophages (TAM) and neutrophils, which release the serine proteases neutrophil elastase (NE) and proteinase 3 (P3) into the tumor microenvironment. NE and P3 shape the antitumor adaptive immune response in breast cancer and melanoma. In this report, we demonstrate that lung cancer cells cross-presented the tumor-associated antigen PR1, derived from NE and P3. Additionally, NE and P3 enhanced the expression of human leukocyte antigen (HLA) class I molecules on lung cancer cells and induced unique, endogenous peptides in the immunopeptidome, as detected with mass spectrometry sequencing. Lung cancer patient tissues with high intratumoral TAM were enriched for MHC class I genes and T-cell markers, and patients with high TAM and cytotoxic T lymphocyte (CTL) infiltration had improved overall survival. We confirmed the immunogenicity of unique, endogenous peptides with cytotoxicity assays against lung cancer cell lines, using CTL from healthy donors that had been expanded against select peptides. Finally, CTL specific for serine proteases–induced endogenous peptides were detected in lung cancer patients using peptide/HLA-A2 tetramers and were elevated in tumor-infiltrating lymphocytes. Thus, serine proteases in the tumor microenvironment of lung cancers promote the presentation of HLA class I immunogenic peptides that are expressed by lung cancer cells, thereby increasing the antigen repertoire that can be targeted in lung cancer. PMID:28254787

Frequency and reactivity of antigen-specific T cells were concurrently measured through the combination of artificial antigen-presenting cell, MACS and ELISPOT.

PubMed

Shen, Chuanlai; Xu, Tao; Wu, You; Li, Xiaoe; Xia, Lingzhi; Wang, Wei; Shahzad, Khawar Ali; Zhang, Lei; Wan, Xin; Qiu, Jie

2017-11-27

Conventional peptide-major histocompatibility complex (pMHC) multimer staining, intracellular cytokine staining, and enzyme-linked immunospot (ELISPOT) assay cannot concurrently determine the frequency and reactivity of antigen-specific T cells (AST) in a single assay. In this report, pMHC multimer, magnetic-activated cell sorting (MACS), and ELISPOT techniques have been integrated into a micro well by coupling pMHC multimers onto cell-sized magnetic beads to characterize AST cell populations in a 96-well microplate which pre-coated with cytokine-capture antibodies. This method, termed AAPC-microplate, allows the enumeration and local cytokine production of AST cells in a single assay without using flow cytometry or fluorescence intensity scanning, thus will be widely applicable. Here, ovalbumin 257-264 -specific CD8 + T cells from OT-1 T cell receptor (TCR) transgenic mice were measured. The methodological accuracy, specificity, reproducibility, and sensitivity in enumerating AST cells compared well with conventional pMHC multimer staining. Furthermore, the AAPC-microplate was applied to detect the frequency and reactivity of Hepatitis B virus (HBV) core antigen 18-27 - and surface antigen 183-191 -specific CD8 + T cells for the patients, and was compared with conventional method. This method without the need of high-end instruments may facilitate the routine analysis of patient-specific cellular immune response pattern to a given antigen in translational studies.

A novel system of artificial antigen-presenting cells efficiently stimulates Flu peptide-specific cytotoxic T cells in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Hui; Peng, Ji-Run, E-mail: pengjr@medmail.com.cn; Chen, Peng-Cheng

Highlights: {yields} Adoptive immunotherapy depends on relevant numbers of cytolytic T lymphocytes. {yields} An ideal artificial APCs system was successfully prepared in vivo. {yields} Controlled release of IL-2 leads to much more T-cell expansion. {yields} This system is better than general cellular APCs on T-cell expansion. -- Abstract: Therapeutic numbers of antigen-specific cytotoxic T lymphocytes (CTLs) are key effectors in successful adoptive immunotherapy. However, efficient and reproducible methods to meet the qualification remain poor. To address this issue, we designed the artificial antigen-presenting cell (aAPC) system based on poly(lactic-co-glycolic acid) (PLGA). A modified emulsion method was used for the preparationmore » of PLGA particles encapsulating interleukin-2 (IL-2). Biotinylated molecular ligands for recognition and co-stimulation of T cells were attached to the particle surface through the binding of avidin-biotin. These formed the aAPC system. The function of aAPCs in the proliferation of specific CTLs against human Flu antigen was detected by enzyme-linked immunospot assay (ELISPOT) and MTT staining methods. Finally, we successfully prepared this suitable aAPC system. The results show that IL-2 is released from aAPCs in a sustained manner over 30 days. This dramatically improves the stimulatory capacity of this system as compared to the effect of exogenous addition of cytokine. In addition, our aAPCs promote the proliferation of Flu antigen-specific CTLs more effectively than the autologous cellular APCs. Here, this aAPC platform is proved to be suitable for expansion of human antigen-specific T cells.« less

Role of the Antigen Capture Pathway in the Induction of a Neutralizing Antibody Response to Anthrax Protective Antigen.

PubMed

Verma, Anita; Ngundi, Miriam M; Price, Gregory A; Takeda, Kazuyo; Yu, James; Burns, Drusilla L

2018-02-27

Toxin neutralizing antibodies represent the major mode of protective immunity against a number of toxin-mediated bacterial diseases, including anthrax; however, the cellular mechanisms that lead to optimal neutralizing antibody responses remain ill defined. Here we show that the cellular binding pathway of anthrax protective antigen (PA), the binding component of anthrax toxin, determines the toxin neutralizing antibody response to this antigen. PA, which binds cellular receptors and efficiently enters antigen-presenting cells by receptor-mediated endocytosis, was found to elicit robust anti-PA IgG and toxin neutralizing antibody responses. In contrast, a receptor binding-deficient mutant of PA, which does not bind receptors and only inefficiently enters antigen-presenting cells by macropinocytosis, elicited very poor antibody responses. A chimeric protein consisting of the receptor binding-deficient PA mutant tethered to the binding subunit of cholera toxin, which efficiently enters cells using the cholera toxin receptor rather than the PA receptor, elicited an anti-PA IgG antibody response similar to that elicited by wild-type PA; however, the chimeric protein elicited a poor toxin neutralizing antibody response. Taken together, our results demonstrate that the antigen capture pathway can dictate the magnitudes of the total IgG and toxin neutralizing antibody responses to PA as well as the ratio of the two responses. IMPORTANCE Neutralizing antibodies provide protection against a number of toxin-mediated bacterial diseases by inhibiting toxin action. Therefore, many bacterial vaccines are designed to induce a toxin neutralizing antibody response. We have used protective antigen (PA), the binding component of anthrax toxin, as a model antigen to investigate immune mechanisms important for the induction of robust toxin neutralizing antibody responses. We found that the pathway used by antigen-presenting cells to capture PA dictates the robustness of the neutralizing antibody response to this antigen. These results provide new insights into immune mechanisms that play an important role in the induction of toxin neutralizing antibody responses and may be useful in the design of new vaccines against toxin-mediated bacterial diseases.

A viral, transporter associated with antigen processing (TAP)-independent, high affinity ligand with alternative interactions endogenously presented by the nonclassical human leukocyte antigen E class I molecule.

PubMed

Lorente, Elena; Infantes, Susana; Abia, David; Barnea, Eilon; Beer, Ilan; García, Ruth; Lasala, Fátima; Jiménez, Mercedes; Mir, Carmen; Morreale, Antonio; Admon, Arie; López, Daniel

2012-10-12

The transporter associated with antigen processing (TAP) enables the flow of viral peptides generated in the cytosol by the proteasome and other proteases to the endoplasmic reticulum, where they complex with nascent human leukocyte antigen (HLA) class I. Later, these peptide-HLA class I complexes can be recognized by CD8(+) lymphocytes. Cancerous cells and infected cells in which TAP is blocked, as well as individuals with unusable TAP complexes, are able to present peptides on HLA class I by generating them through TAP-independent processing pathways. Here, we identify a physiologically processed HLA-E ligand derived from the D8L protein in TAP-deficient vaccinia virus-infected cells. This natural high affinity HLA-E class I ligand uses alternative interactions to the anchor motifs previously described to be presented on nonclassical HLA class I molecules. This octameric peptide was also presented on HLA-Cw1 with similar binding affinity on both classical and nonclassical class I molecules. In addition, this viral peptide inhibits HLA-E-mediated cytolysis by natural killer cells. Comparison between the amino acid sequences of the presenting HLA-E and HLA-Cw1 alleles revealed a shared structural motif in both HLA class molecules, which could be related to their observed similar cross-reactivity affinities. This motif consists of several residues located on the floor of the peptide-binding site. These data expand the role of HLA-E as an antigen-presenting molecule.

Polymer nanoparticles for cross-presentation of exogenous antigens and enhanced cytotoxic T-lymphocyte immune response

PubMed Central

Song, Chanyoung; Noh, Young-Woock; Lim, Yong Taik

2016-01-01

Effective induction of an antigen-specific cytotoxic T lymphocyte (CTL) immune response is one of the key goals of cancer immunotherapy. We report the design and fabrication of polyethylenimine (PEI)-coated polymer nanoparticles (NPs) as efficient antigen-delivery carriers that can induce antigen cross-presentation and a strong CTL response. After synthesis of poly(d,l-lactide-co-glycolide) (PLGA) NPs containing ovalbumin (OVA) by the double-emulsion solvent-evaporation method, cationic-charged PLGA NPs were generated by coating them with PEI. In a methyl tetrazolium salt assay, no discernible cytotoxic effect of PEI-coated PLGA (OVA) NPs was observed. The capacity and mechanism of PEI-coated PLGA (OVA) NPs for antigen delivery and cross-presentation on dendritic cells (DCs) were determined by fluorescence microscopy and flow cytometry. PEI-coated PLGA (OVA) NPs were internalized efficiently via phagocytosis or macropinocytosis in DCs and induced efficient cross-presentation of the antigen on MHC class I molecules via both endosome escape and a lysosomal processing mechanism. The DCs treated with PEI-coated PLGA (OVA) NPs induced a release of IL-2 cytokine from OVA-specific CD8-OVA1.3 T cells more efficiently than DCs treated with PLGA (OVA) NPs. Therefore, the PEI-coated PLGA (OVA) NPs can induce antigen cross-presentation and are expected to be used for induction of a strong CTL immune response and for efficient anticancer immunotherapy. PMID:27540289

Band 3 in aging and neurological disease.

PubMed

Kay, M M

1991-01-01

Senescent cell antigen appears on old cells and marks them for death by initiating the binding of IgG autoantibody and subsequent removal by phagocytes in mammals and other vertebrates. We have created a synthetic aging antigen that blocks binding of IgG to senescent cells in vitro. Synthetic senescent cell antigen might be effective in preventing cellular destruction in vivo in certain diseases, and can be used to manipulate cellular life span in situ. Senescent cell antigen is generated by the modification of an important structural and transport membrane molecule, protein band 3. Band 3 is present in cellular, nuclear, Golgi, and mitochondrial membranes as well as in cell membranes. Band 3 proteins in nucleated cells participate in cell surface patching and capping. Band 3 maintains acid-base balance by mediating the exchange of anions (e.g., chloride, bicarbonate), and is the binding site for glycolytic enzymes. It is responsible for CO2 exchange in all tissues and organs. Thus, it is the most heavily used anion transport system in the body. Band 3 is a major transmembrane structural protein which attaches the plasma membrane to the internal cell cytoskeleton by binding to band 2.1 (ankyrin). Oxidation generates senescent cell antigen in situ. Band 3 is present in the central nervous system, and differences have been described in band 3 between young and aging brain tissue. One autosomal recessive neurological disease, choreoacanthocytosis, is associated with band 3 abnormalities. The 150 residues of the carboxyl terminus segment of band 3 appear to be altered. In brains from Alzheimer's disease patients, antibodies to aged band 3 label the amyloid core of classical plaques and the microglial cells located in the middle of the plaque in tissue sections, and an abnormal band 3 in immunoblots. Band 3 protein(s) in mammalian brain performs the same functions as that of erythroid band 3. These functions is anion transport, ankyrin binding, and generation of senescent cell antigen, an aging antigen that terminates the life of cells. Structural similarity of brain and erythroid band 3 is suggested by the reaction of antibodies to synthetic peptides of erythroid band 3 with brain band 3, the inhibition of anion transport by the same inhibitors, and an equal degree of inhibition of brain and erythrocyte anion transport by synthetic peptides of erythroid band 3. One of these segments, pep-COOH, contains antigenic determinants of senescent cell antigen.(ABSTRACT TRUNCATED AT 400 WORDS)

Endothelial microparticles interact with and support the proliferation of T cells

PubMed Central

Wheway, Julie; Latham, Sharissa L; Combes, Valery; Grau, Georges ER

2014-01-01

Endothelial cells (EC) closely interact with circulating lymphocytes. Aggression or activation of the endothelium leads to an increased shedding of EC microparticles (MP). Endothelial MP (EMP) are found in high plasma levels in numerous immunoinflammatory diseases, e.g. atherosclerosis, sepsis, multiple sclerosis and cerebral malaria, supporting their role as effectors and markers of vascular dysfunction. Given our recently described role for human brain microvascular endothelial cells (HBEC) in modulating immune responses we investigated how HBEC-derived MP could interact with and support the proliferation of T cells. Like their mother cells, EMP expressed molecules important for antigen presentation and T cell co-stimulation, i.e., β2-microglobulin, MHC II, CD40 and ICOSL. HBEC were able to take up fluorescently labeled antigens with EMP also containing fluorescent antigens suggestive of antigen carryover from HBEC to EMP. In co-cultures, fluorescently labeled EMP from resting or cytokine-stimulated HBEC formed conjugates with both CD4+ and CD8+ subsets, with higher proportions of T cells binding EMP from cytokine stimulated cells. The increased binding of EMP from cytokine stimulated HBEC to T cells was VCAM-1 and ICAM-1-dependent. Finally, in CFSE T cell proliferation assays using anti-CD3 mAb or T cell mitogens, EMP promoted the proliferation of CD4+ T cells and that of CD8+ T cells in the absence of exogenous stimuli and in the T cell mitogenic stimulation. Our findings provide novel evidence that EMP can enhance T cell activation and potentially ensuing antigen presentation, thereby pointing towards a novel role for MP in neuro-immunological complications of infectious diseases. PMID:25187656

T-Cell Receptor- and CD28-induced Vav1 activity is required for the accumulation of primed T cells into antigenic tissue

PubMed Central

David, Rachel; Ma, Liang; Ivetic, Aleksandar; Takesono, Aya; Ridley, Anne J.; Chai, Jian-Guo; Tybulewicz, Victor; Marelli-Berg, Federica M.

2016-01-01

Localization of primed T cells to antigenic tissue is essential for the development of effective immunity. Together with tissue-selective homing molecules, T-cell receptor (TCR)- and CD28-mediated signals have been shown to promote transendothelial migration of specific T cells into non-lymphoid antigen-rich tissue tissue. However, the cellular and molecular requirements for T-cell accumulation to target tissue following their recruitment are largely undefined. The guanine nucleotide exchange factor (GEF) Vav1 has an integral role in coupling TCR and CD28 to signalling pathways that regulate T cell activation and migration. Here, we have investigated the contribution of TCR- and CD28-induced Vav1 activity to the trafficking and localization of primed HY-specific CD4+ T cells to antigenic sites. Severe migratory defects displayed by Vav1-/- T cells in vitro were fully compensated by a combination of shear flow and chemokines, leading to normal recruitment of Vav1-/- T cells in vivo. In contrast, Vav1-/- T-cell retention into antigen-rich tissue was severely impaired, reflecting their inability to engage in sustained TCR- and CD28-mediated interactions with tissue-resident antigen-presenting cells (APCs). This novel function of APC-induced, TCR- and CD28-mediated Vav1 activity in the regulation of effector T-cell immunity highlights its potential as a therapeutic target in T-cell-mediated tissue damage. PMID:19060239

Methods and compositions for diagnosing and preventing a group B streptococcal infection

DOEpatents

Brady, Linda Jeannine [Gainesville, FL; Seifert, Kyle N [Harrisonburg, VA; Adderson, Elisabeth E [Memphis, TN; Bohnsack, John F [Salt Lake City, UT

2009-09-15

The present invention provides a group B streptococcal (GBS) surface antigen, designated epsilon antigen, that is co-expressed with the delta antigen on a subset of serotype III GBS. Epsilon is expressed on more pathogenic Restriction Digest Pattern (RDP) III-3 GBS, but not on RDP types 1, 2, or 4. Accordingly, the present invention provides compositions and methods for detecting a group B streptococcus serotype III, RDP III-3 strain. Vaccines and methods of identifying agents which inhibit adhesion of a group B streptococcal cell to a host cell are also provided.

Antigen delivery by α2-macroglobulin enhances the cytotoxic T lymphocyte response

PubMed Central

Bowers, Edith V.; Horvath, Jeffrey J.; Bond, Jennifer E.; Cianciolo, George J.; Pizzo, Salvatore V.

2009-01-01

α2M* targets antigens to APCs for rapid internalization, processing, and presentation. When used as an antigen-delivery vehicle, α2M* amplifies MHC class II presentation, as demonstrated by increased antibody titers. Recent evidence, however, suggests that α2M* encapsulation may also enhance antigen-specific CTL immunity. In this study, we demonstrate that α2M*-delivered antigen (OVA) enhances the production of specific in vitro and in vivo CTL responses. Murine splenocytes expressing a transgenic TCR specific for CTL peptide OVA257–264 (SIINFEKL) demonstrated up to 25-fold greater IFN-γ and IL-2 secretion when treated in vitro with α2M*-OVA compared with soluble OVA. The frequency of IFN-γ-producing cells was increased ∼15-fold, as measured by ELISPOT. Expansion of the OVA-specific CD8+ T cell population, as assayed by tetramer binding and [3H]thymidine incorporation, and OVA-specific cell-mediated cytotoxicity, as determined by a flow cytometric assay, were also enhanced significantly by α2M*-OVA. Furthermore, significant CTL responses were observed at antigen doses tenfold lower than those required with OVA alone. Finally, we also observed enhanced humoral and CTL responses by naïve mice following intradermal immunization with α2M*-OVA. These α2M*-OVA-immunized mice demonstrated increased protection against a s.c.-implanted, OVA-expressing tumor, as demonstrated by delayed tumor growth and prolonged animal survival. The observation that α2M*-mediated antigen delivery elicits specific CTL responses suggests the cross-presentation of antigen onto MHC class I. These results support α2M* as an effective antigen-delivery system that may be particularly useful for vaccines based on weakly immunogenic subunits or requiring dose sparing. PMID:19652028

CD8 single-cell gene coexpression reveals three different effector types present at distinct phases of the immune response

PubMed Central

Peixoto, António; Evaristo, César; Munitic, Ivana; Monteiro, Marta; Charbit, Alain; Rocha, Benedita; Veiga-Fernandes, Henrique

2007-01-01

To study in vivo CD8 T cell differentiation, we quantified the coexpression of multiple genes in single cells throughout immune responses. After in vitro activation, CD8 T cells rapidly express effector molecules and cease their expression when the antigen is removed. Gene behavior after in vivo activation, in contrast, was quite heterogeneous. Different mRNAs were induced at very different time points of the response, were transcribed during different time periods, and could decline or persist independently of the antigen load. Consequently, distinct gene coexpression patterns/different cell types were generated at the various phases of the immune responses. During primary stimulation, inflammatory molecules were induced and down-regulated shortly after activation, generating early cells that only mediated inflammation. Cytotoxic T cells were generated at the peak of the primary response, when individual cells simultaneously expressed multiple killer molecules, whereas memory cells lost killer capacity because they no longer coexpressed killer genes. Surprisingly, during secondary responses gene transcription became permanent. Secondary cells recovered after antigen elimination were more efficient killers than cytotoxic T cells present at the peak of the primary response. Thus, primary responses produced two transient effector types. However, after boosting, CD8 T cells differentiate into long-lived killer cells that persist in vivo in the absence of antigen. PMID:17485515

Hypoxia and hypoxia-inducible factor (HIF) downregulate antigen-presenting MHC class I molecules limiting tumor cell recognition by T cells

PubMed Central

Nguyen, Thao; Hatfield, Stephen M.; Ohta, Akio; Sitkovsky, Michail V.

2017-01-01

Human cancers are known to downregulate Major Histocompatibility Complex (MHC) class I expression thereby escaping recognition and rejection by anti-tumor T cells. Here we report that oxygen tension in the tumor microenvironment (TME) serves as an extrinsic cue that regulates antigen presentation by MHC class I molecules. In support of this view, hypoxia is shown to negatively regulate MHC expression in a HIF-dependent manner as evidenced by (i) lower MHC expression in the hypoxic TME in vivo and in hypoxic 3-dimensional (3D) but not 2-dimensional (2D) tumor cell cultures in vitro; (ii) decreased MHC in human renal cell carcinomas with constitutive expression of HIF due to genetic loss of von Hippel-Lindau (VHL) function as compared with isogenically paired cells with restored VHL function, and iii) increased MHC in tumor cells with siRNA-mediated knockdown of HIF. In addition, hypoxia downregulated antigen presenting proteins like TAP 1/2 and LMP7 that are known to have a dominant role in surface display of peptide-MHC complexes. Corroborating oxygen-dependent regulation of MHC antigen presentation, hyperoxia (60% oxygen) transcriptionally upregulated MHC expression and increased levels of TAP2, LMP2 and 7. In conclusion, this study reveals a novel mechanism by which intra-tumoral hypoxia and HIF can potentiate immune escape. It also suggests the use of hyperoxia to improve tumor cell-based cancer vaccines and for mining novel immune epitopes. Furthermore, this study highlights the advantage of 3D cell cultures in reproducing hypoxia-dependent changes observed in the TME. PMID:29155844

Herbal preparation (HemoHIM) enhanced functional maturation of bone marrow-derived dendritic cells mediated toll-like receptor 4.

PubMed

Lee, Sung-Ju; Kim, Jong-Jin; Kang, Kyung-Yun; Hwang, Yun-Ho; Jeong, Gil-Yeon; Jo, Sung-kee; Jung, Uhee; Park, Hae-Ran; Yee, Sung-Tae

2016-02-19

HemoHIM, which is an herbal preparation of three edible herbs (Angelicam gigas Nakai, Cnidium offinale Makino, and Peaonia japonica Miyabe), is known to have various biological and immunological activities, but the modulatory effects of this preparation on dendritic cells (DCs)-mediated immune responses have not been examined previously. DCs are a unique group of white blood cells that initiate primary immune responses by capturing, processing, and presenting antigens to T cells. In the present study, we investigated the effect of HemoHIM on the functional and phenotypic maturation of murine bone marrow-derived dendritic cells (BMDCs) both in vitro and in vivo. The expression of co-stimulatory molecules (CD40, CD80, CD86, MHC I, and MHC II) and the production of cytokines (IL-1β, IL-6, IL-12p70, and TNF-α) were increased by HemoHIM in BMDCs. Furthermore, the antigen-uptake ability of BMDCs was decreased by HemoHIM, and the antigen-presenting ability of HemoHIM-treated mature BMDCs increased TLR4-dependent CD4(+) and CD8(+) T cell responses. Our findings demonstrated that HemoHIM induces TLR4-mediated BMDCs functional and phenotypic maturation through in vivo and in vitro. And our study showed the antigen-presenting ability that HemoHIM-treated mature BMDCs increase CD4(+) and CD8(+) T cell responses by in vitro. These results suggest that HemoHIM has the potential to mediate DC immune responses.

The many sounds of T lymphocyte silence.

PubMed

Melero, Ignacio; Arina, Ainhoa; Chen, Lieping

2005-01-01

It is not unusual for antigens and potentially responsive T cells to co-exist in the same organism while these T cells remain silent and do not mount life-threatening immune responses. A rich array of mechanisms has been proposed to explain these observations. T cell silencing is controlled in multiple levels. Initially, dendritic cells and regulatory T cells appear to play critical roles. In addition, T cell immunity is tightly regulated by a molecular network of cytokines and cell receptor interactions by the opposed surfaces of antigen-presenting cells and T cells. Recognition of a specific antigen is therefore shaped and tuned by co-stimulatory and co-inhibitory receptor-ligand pairs. At last, immunologists are beginning to exploit the rules governing these assorted sounds of T cell silence.

Photochemical Internalization of Peptide Antigens Provides a Novel Strategy to Realize Therapeutic Cancer Vaccination

PubMed Central

Haug, Markus; Brede, Gaute; Håkerud, Monika; Nedberg, Anne Grete; Gederaas, Odrun A.; Flo, Trude H.; Edwards, Victoria T.; Selbo, Pål K.; Høgset, Anders; Halaas, Øyvind

2018-01-01

Effective priming and activation of tumor-specific CD8+ cytotoxic T lymphocytes (CTLs) is crucial for realizing the potential of therapeutic cancer vaccination. This requires cytosolic antigens that feed into the MHC class I presentation pathway, which is not efficiently achieved with most current vaccination technologies. Photochemical internalization (PCI) provides an emerging technology to route endocytosed material to the cytosol of cells, based on light-induced disruption of endosomal membranes using a photosensitizing compound. Here, we investigated the potential of PCI as a novel, minimally invasive, and well-tolerated vaccination technology to induce priming of cancer-specific CTL responses to peptide antigens. We show that PCI effectively promotes delivery of peptide antigens to the cytosol of antigen-presenting cells (APCs) in vitro. This resulted in a 30-fold increase in MHC class I/peptide complex formation and surface presentation, and a subsequent 30- to 100-fold more efficient activation of antigen-specific CTLs compared to using the peptide alone. The effect was found to be highly dependent on the dose of the PCI treatment, where optimal doses promoted maturation of immature dendritic cells, thus also providing an adjuvant effect. The effect of PCI was confirmed in vivo by the successful induction of antigen-specific CTL responses to cancer antigens in C57BL/6 mice following intradermal peptide vaccination using PCI technology. We thus show new and strong evidence that PCI technology holds great potential as a novel strategy for improving the outcome of peptide vaccines aimed at triggering cancer-specific CD8+ CTL responses. PMID:29670624

Food antigen-induced immune responses in Crohn's disease patients and experimental colitis mice.

PubMed

Kawaguchi, Takaaki; Mori, Maiko; Saito, Keiko; Suga, Yasuyo; Hashimoto, Masaki; Sako, Minako; Yoshimura, Naoki; Uo, Michihide; Danjo, Keiko; Ikenoue, Yuka; Oomura, Kaori; Shinozaki, Junko; Mitsui, Akira; Kajiura, Takayuki; Suzuki, Manabu; Takazoe, Masakazu

2015-04-01

In Crohn's disease (CD), the involvement of food antigens in immune responses remains unclear. The objective of this study was to detect immune responses against food antigens in CD patients and examine the mechanism in a mouse model of colitis. We enrolled 98 CD patients, 50 ulcerative colitis patients, and 52 healthy controls (HCs) to compare the levels of serum immunoglobulin (Ig)Gs against 88 foods. The presence of serum IgGs against foods was also examined in interleukin (IL)-10 knockout (KO) mice in which CD4(+) T cell activation by antigenic food protein was assessed. Mice transferred with IL-10 KO cells received diets with or without food antigens, and the development of colitis was evaluated. The prevalence of IgGs against various foods, especially vegetables, grains, and nuts, was significantly higher in CD patients than in HCs. Similarly, the prevalence of IgGs against food proteins was higher in IL-10 KO mice than in BALB/c mice. Beta-conglycinin, identified as an antigenic food proteins in IL-10 KO mice, induced CD4(+) T cell production of interferon-γ and IL-17 through dendritic cell antigen presentation. Elimination of the food antigens ameliorated the development of colitis in mice without altering the composition of their intestinal microbiota. In CD colitis mice, intestinal inflammation via CD4(+) T cell hyperactivation was induced by food antigens associated with high serum IgG levels and was ameliorated by the elimination of food antigens. This disrupted immunological tolerance to food antigen, which might act as an exacerbating factor, remains to be elucidated in CD patients.

Antitoxic Cholera Immunity in Mice: Influence of Antigen Deposition on Antitoxin-Containing Cells and Protective Immunity in Different Parts of the Intestine

PubMed Central

Lange, Stefan; Nygren, Håkan; Svennerholm, Ann-Mari; Holmgren, Jan

1980-01-01

The importance of the mode of antigen presentation (intravenous, oral, or enteral restricted to the lower ileum) in the development of a local immune response and immunological memory for such a response in different parts of the intestine was studied in mice. Cholera toxin was used as antigen and the immune response was assayed by determining both the number of specific antitoxin-containing cells in the lamina propria and protection against experimental cholera. The results showed that all of these routes of antigen presentation could induce significant memory along the entire small intestine. In contrast, the actual production of antitoxin-containing cells or protective immune response elicited by booster immunization was restricted to those parts of the intestine that were directly exposed to antigen; i.e., lower ileum boosting resulted in immunity in the distal ileum but not in the proximal jejunum, whereas oral or intravenous boosting gave a response in both jejunum and ileum. Protection correlated closely with the number of antitoxin-containing cells in the lamina propria (correlation coefficient, 0.88); ≥4,000 antitoxin-containing cells per mm3 conferred solid immunity to cholera toxin-induced diarrhea. The total number of immunoglobulin-containing cells in intestines was not significantly influenced by the specific immunizations. There were four times as many of these cells in the upper jejunum (167,000 cells per mm3) as in the lower ileum, but the proportions of immunoglobulin A-containing cells (80 to 85%), immunoglobulin M-containing cells (14 to 20%), and immunoglobulin G-containing cells (0.4 to 0.9%) were similar in various parts of the intestine. The results indicate a differential dependence on local tissue antigen for the intestinal antibody-secreting cells and their memory cell precursors. PMID:7189747

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens

PubMed Central

Kleiveland, Charlotte R.; Minic, Rajna; Moen, Lars F.; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Eijsink, Vincent G. H.

2016-01-01

ABSTRACT Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum. The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo. The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. IMPORTANCE This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and to anchor this antigen to the bacterial cell wall or to the cell membrane. The recombinant strains elicited proliferative antigen-specific T-cell responses in white blood cells from tuberculosis-positive humans and induced specific immune responses after nasal and oral administrations in mice. PMID:27815271

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens.

PubMed

Kuczkowska, Katarzyna; Kleiveland, Charlotte R; Minic, Rajna; Moen, Lars F; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Mathiesen, Geir; Eijsink, Vincent G H

2017-01-15

Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and to anchor this antigen to the bacterial cell wall or to the cell membrane. The recombinant strains elicited proliferative antigen-specific T-cell responses in white blood cells from tuberculosis-positive humans and induced specific immune responses after nasal and oral administrations in mice. Copyright © 2016 American Society for Microbiology.

Macrophages play an essential role in antigen-specific immune suppression mediated by T CD8⁺ cell-derived exosomes.

PubMed

Nazimek, Katarzyna; Ptak, Wlodzimierz; Nowak, Bernadeta; Ptak, Maria; Askenase, Philip W; Bryniarski, Krzysztof

2015-09-01

Murine contact sensitivity (CS) reaction could be antigen-specifically regulated by T CD8(+) suppressor (Ts) lymphocytes releasing microRNA-150 in antibody light-chain-coated exosomes that were formerly suggested to suppress CS through action on macrophages (Mφ). The present studies investigated the role of Mφ in Ts cell-exosome-mediated antigen-specific suppression as well as modulation of Mφ antigen-presenting function in humoral and cellular immunity by suppressive exosomes. Mice depleted of Mφ by clodronate liposomes could not be tolerized and did not produce suppressive exosomes. Moreover, isolated T effector lymphocytes transferring CS were suppressed by exosomes only in the presence of Mφ, demonstrating the substantial role of Mφ in the generation and action of Ts cell regulatory exosomes. Further, significant decrease of number of splenic B cells producing trinitrophenyl (TNP) -specific antibodies with the alteration of the ratio of serum titres of IgM to IgG was observed in recipients of exosome-treated, antigen-pulsed Mφ and the significant suppression of CS was demonstrated in recipients of exosome-treated, TNP-conjugated Mφ. Additionally, exosome-pulsed, TNP-conjugated Mφ mediated suppression of CS in mice pre-treated with a low-dose of cyclophosphamide, suggesting de novo induction of T regulatory (Treg) lymphocytes. Treg cell involvement in the effector phase of the studied suppression mechanism was proved by unsuccessful tolerization of DEREG mice depleted of Treg lymphocytes. Furthermore, the inhibition of proliferation of CS effector cells cultured with exosome-treated Mφ in a transmembrane manner was observed. Our results demonstrated the essential role of Mφ in antigen-specific immune suppression mediated by Ts cell-derived exosomes and realized by induction of Treg lymphocytes and inhibition of T effector cell proliferation. © 2015 John Wiley & Sons Ltd.

T Cell Receptor Engineering and Analysis Using the Yeast Display Platform

PubMed Central

Smith, Sheena N.; Harris, Daniel T.; Kranz, David M.

2017-01-01

The αβ heterodimeric T cell receptor (TCR) recognizes peptide antigens that are transported to the cell surface as a complex with a protein encoded by the major histocompatibility complex (MHC). T cells thus evolved a strategy to sense these intracellular antigens, and to respond either by eliminating the antigen-presenting cell (e.g. a virus-infected cell) or by secreting factors that recruit the immune system to the site of the antigen. The central role of the TCR in the binding of antigens as peptide-MHC (pepMHC) ligands has now been studied thoroughly. Interestingly, despite their exquisite sensitivity (e.g. T cell activation by as few as 1 to 3 pepMHC complexes on a single target cell), TCRs are known to have relatively low affinities for pepMHC, with KD values in the micromolar range. There has been interest in engineering the affinity of TCRs in order to use this class of molecules in ways similar to now done with antibodies. By doing so, it would be possible to harness the potential of TCRs as therapeutics against a much wider array of antigens that include essentially all intracellular targets. To engineer TCRs, and to analyze their binding features more rapidly, we have used a yeast display system as a platform. Expression and engineering of a single-chain form of the TCR, analogous to scFv fragments from antibodies, allow the TCR to be affinity matured with a variety of possible pepMHC ligands. In addition, the yeast display platform allows one to rapidly generate TCR variants with diverse binding affinities and to analyze specificity and affinity without the need for purification of soluble forms of the TCRs. The present chapter describes the methods for engineering and analyzing single-chain TCRs using yeast display. PMID:26060072

Organic extract of diesel exhaust particles stimulates expression of Ia and costimulatory molecules associated with antigen presentation in rat peripheral blood monocytes but not in alveolar macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Koike, Eiko; Kobayashi, Takahiro

2005-12-15

We hypothesized that diesel exhaust particles (DEP) induce the activation of antigen-presenting cells (APC) in lung. The present study was designed to clarify the following about DEP: (1) whether it affects the expression of Ia and B7 molecules in alveolar macrophages (AM) as a mature cell or in peripheral blood monocytes (PBM) as an immature cell (2) if it affects the antigen-presenting (AP) activity of PBM (3) what component of DEP is responsible for the effects, and (4) whether the effect of DEP is related to oxidative stress. DEP was extracted with methylene chloride. Cells were exposed to whole DEP,more » organic extract, or residual particles for 24 h. Cell-surface molecules were measured by flow cytometry. AP activity was assessed by antigen-specific T cell proliferation. Whole DEP or organic extract significantly increased the expression of Ia and B7 molecules on PBM but not on AM. No significant effect of residual particles was observed. A low concentration of organic extract also increased the AP activity of PBM. When the induction of an antioxidative enzyme was assessed, heme oxygenase-1 protein was found to be significantly increased by exposure to whole DEP, and the organic extract was more effective than the residual particles. Furthermore, the organic extract-induced expression of Ia antigen on PBM was reduced by the addition of an antioxidative agent. These results suggest that DEP may act on immature APC and enhance their AP activity and that the action contributing to oxidative stress may be mediated by organic compounds of DEP.« less

Spontaneous CD4+ and CD8+ T-cell responses directed against cancer testis antigens are present in the peripheral blood of testicular cancer patients.

PubMed

Pearce, Hayden; Hutton, Paul; Chaudhri, Shalini; Porfiri, Emilio; Patel, Prashant; Viney, Richard; Moss, Paul

2017-07-01

Cancer/testis antigen (CTAg) expression is restricted to spermatogenic cells in an immune-privileged site within the testis. However, these proteins are expressed aberrantly by malignant cells and T-cell responses against CTAgs develop in many cancer patients. We investigated the prevalence, magnitude and phenotype of CTAg-specific T cells in the blood of patients with testicular germ cell tumors (TGCTs). CD8 + and CD4 + T-cell responses against MAGE-A family antigens were present in 44% (20/45) of patients' samples assayed by ex vivo IFN-γ ELISPOT. The presence of MAGE-specific CD8 + T cells was further determined following short-term in vitro expansion through the use of pMHC-I multimers containing known immunogenic peptides. Longitudinal analysis revealed that the frequency of MAGE-specific T cells decreased by 89% following orchidectomy suggesting that persistence of tumor antigen is required to sustain CTAg-specific T-cell immunity. Notably, this decrease correlated with a decline in the global effector/memory T-cell pool following treatment. Spontaneous T-cell immunity against CTAg proteins therefore develops in many patients with testicular cancer and may play an important role in the excellent clinical outcome of patients with this tumor subtype. © 2017 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Genetically modified anthrax lethal toxin safely delivers whole HIV protein antigens into the cytosol to induce T cell immunity

NASA Astrophysics Data System (ADS)

Lu, Yichen; Friedman, Rachel; Kushner, Nicholas; Doling, Amy; Thomas, Lawrence; Touzjian, Neal; Starnbach, Michael; Lieberman, Judy

2000-07-01

Bacillus anthrax lethal toxin can be engineered to deliver foreign proteins to the cytosol for antigen presentation to CD8 T cells. Vaccination with modified toxins carrying 8-9 amino acid peptide epitopes induces protective immunity in mice. To evaluate whether large protein antigens can be used with this system, recombinant constructs encoding several HIV antigens up to 500 amino acids were produced. These candidate HIV vaccines are safe in animals and induce CD8 T cells in mice. Constructs encoding gag p24 and nef stimulate gag-specific CD4 proliferation and a secondary cytotoxic T lymphocyte response in HIV-infected donor peripheral blood mononuclear cells in vitro. These results lay the foundation for future clinical vaccine studies.

The effect of the parenteral administration of a rabbit anti-(mouse)-IgD serum on the immune response of mice to sheep erythrocytes.

PubMed Central

Dresser, D W; Parkhouse, R M

1978-01-01

Experiments have been carried out to find out if the administration of an anti-IgD serum to mice interferes in anyway with their immune response to sheep red blood cells. This was done to test a hypothesis that a biological role for IgD might be as a critical cellular receptor for antigen. Our results show that the injection of anti-IgD two days before antigen results in suppression of primary responses and priming (the antigen dependent generation of memory cells) but has no suppressive effect on a secondary response. On this indirect evidence we conclude that it is likely that IgD is present on antigen-sensitive percursor cells but not on memory cells. PMID:367957

Recruitment of bone marrow CD11b+Gr-1+ cells by polymeric nanoparticles for antigen cross-presentation

NASA Astrophysics Data System (ADS)

Yang, Ya-Wun; Luo, Wen-Hui

2017-03-01

The objective of this study was to investigate the function of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) on the activation of antigen-specific CD8+ T cell responses via the CD11b+Gr-1+ myeloid subpopulations in murine bone marrow (BM). PLGA NPs containing ovalbumin (OVA) were fabricated by the double-emulsion method. The CD11b+Gr-1lowLy-6Chigh and CD11b+Gr-1highLy-6Clow subsets from mice bone marrow were sorted and treated with the PLGA/OVA NPs, followed by co-culture with the carboxyfluorescein succinimidyl ester (CFSE)-labelled OT-I CD8+ cells. Co-culture of OT-I CD8+ T cells with PLGA/OVA NPs-primed CD11b+Gr-1+ subsets upregulated the expression of IL-2, TNF-α, INF-γ, granzyme B, and perforin, resulting in proliferation of CD8+ T cells and differentiation into effector cytotoxic T lymphocytes (CTLs). In vivo proliferation of CFSE-labelled OT-I CD8+ cells in response to OVA was also obtained in the animals immunized with PLGA/OVA NPs. The results presented in this study demonstrate the ability of polymeric NPs to recruit two CD11b+Gr-1+ myeloid subsets for effective presentation of exogenous antigen to OT-I CD8+ T cells in the context of major histocompatibility complex (MHC) class I, leading to an induction of antigen-specific cell proliferation and differentiation into effector cells.

Environmental Factors Contribute to β Cell Endoplasmic Reticulum Stress and Neo-Antigen Formation in Type 1 Diabetes

PubMed Central

Marré, Meghan L.; Piganelli, Jon D.

2017-01-01

Type 1 diabetes (T1D) is an autoimmune disease in which immune-mediated targeting and destruction of insulin-producing pancreatic islet β cells leads to chronic hyperglycemia. There are many β cell proteins that are targeted by autoreactive T cells in their native state. However, recent studies have demonstrated that many β cell proteins are recognized as neo-antigens following posttranslational modification (PTM). Although modified neo-antigens are well-established targets of pathology in other autoimmune diseases, the effects of neo-antigens in T1D progression and the mechanisms by which they are generated are not well understood. We have demonstrated that PTM occurs during endoplasmic reticulum (ER) stress, a process to which β cells are uniquely susceptible due to the high rate of insulin production in response to dynamic glucose sensing. In the context of genetic susceptibility to autoimmunity, presentation of these modified neo-antigens may activate autoreactive T cells and cause pathology. However, inherent β cell ER stress and protein PTM do not cause T1D in every genetically susceptible individual, suggesting the contribution of additional factors. Indeed, many environmental factors, such as viral infection, chemicals, or inflammatory cytokines, are associated with T1D onset, but the mechanisms by which these factors lead to disease onset remain unknown. Since these environmental factors also cause ER stress, exposure to these factors may enhance production of neo-antigens, therefore boosting β cell recognition by autoreactive T cells and exacerbating T1D pathogenesis. Therefore, the combined effects of physiological ER stress and the stress that is induced by environmental factors may lead to breaks in peripheral tolerance, contribute to antigen spread, and hasten disease onset. This Hypothesis and Theory article summarizes what is currently known about ER stress and protein PTM in autoimmune diseases including T1D and proposes a role for environmental factors in breaking immune tolerance to β cell antigens through neo-antigen formation. PMID:29033899

Critical role for Sec22b-dependent antigen cross-presentation in antitumor immunity

PubMed Central

Rookhuizen, Derek C.; Joannas, Leonel; Carpier, Jean-Marie; Yatim, Nader; Albert, Matthew L.

2017-01-01

CD8+ T cells mediate antigen-specific immune responses that can induce rejection of solid tumors. In this process, dendritic cells (DCs) are thought to take up tumor antigens, which are processed into peptides and loaded onto MHC-I molecules, a process called “cross-presentation.” Neither the actual contribution of cross-presentation to antitumor immune responses nor the intracellular pathways involved in vivo are clearly established because of the lack of experimental tools to manipulate this process. To develop such tools, we generated mice bearing a conditional DC-specific mutation in the sec22b gene, a critical regulator of endoplasmic reticulum–phagosome traffic required for cross-presentation. DCs from these mice show impaired cross-presentation ex vivo and defective cross-priming of CD8+ T cell responses in vivo. These mice are also defective for antitumor immune responses and are resistant to treatment with anti–PD-1. We conclude that Sec22b-dependent cross-presentation in DCs is required to initiate CD8+ T cell responses to dead cells and to induce effective antitumor immune responses during anti–PD-1 treatment in mice. PMID:28663435

A GMP-compliant protocol to expand and transfect cancer patient T cells with mRNA encoding a tumor-specific chimeric antigen receptor.

PubMed

Krug, Christian; Wiesinger, Manuel; Abken, Hinrich; Schuler-Thurner, Beatrice; Schuler, Gerold; Dörrie, Jan; Schaft, Niels

2014-10-01

Chimeric antigen receptors (CARs), which combine an antibody-derived binding domain (single chain fragment variable) with T-cell-activating signaling domains, have become a promising tool in the adoptive cellular therapy of cancer. Retro- and lenti-viral transductions are currently the standard methods to equip T cells with a CAR; permanent CAR expression, however, harbors several risks like uncontrolled auto-reactivity. Modification of T cells by electroporation with CAR-encoding RNA to achieve transient expression likely circumvents these difficulties. We here present a GMP-compliant protocol to activate and expand T cells for clinical application. The protocol is optimized in particular to produce CAR-modified T cells in clinically sufficient numbers under full GMP-compliance from late-stage cancer patients. This protocol allows the generation of 6.7 × 10(8) CAR-expressing T cells from one patient leukapheresis. The CAR-engineered T cells produced pro-inflammatory cytokines after stimulation with antigen-bearing tumor cells and lysed tumor cells in an antigen-specific manner. This functional capacity was maintained after cryopreservation. Taken together, we provide a clinically applicable protocol to transiently engineer sufficient numbers of antigen-specific patient T cells for use in adoptive cell therapy of cancer.

mRNA Cancer Vaccines-Messages that Prevail.

PubMed

Grunwitz, Christian; Kranz, Lena M

2017-01-01

During the last decade, mRNA became increasingly recognized as a versatile tool for the development of new innovative therapeutics. Especially for vaccine development, mRNA is of outstanding interest and numerous clinical trials have been initiated. Strikingly, all of these studies have proven that large-scale GMP production of mRNA is feasible and concordantly report a favorable safety profile of mRNA vaccines. Induction of T-cell immunity is a multi-faceted process comprising antigen acquisition, antigen processing and presentation, as well as immune stimulation. The effectiveness of mRNA vaccines is critically dependent on making the antigen(s) of interest available to professional antigen-presenting cells, especially DCs. Efficient delivery of mRNA into DCs in vivo remains a major challenge in the mRNA vaccine field. This review summarizes the principles of mRNA vaccines and highlights the importance of in vivo mRNA delivery and recent advances in harnessing their therapeutic potential.

Multiple myeloma presenting as CEA-producing rectal cancer.

PubMed

Talamo, Giampaolo; Barochia, Amitkumar; Zangari, Maurizio; Loughran, Thomas P

2010-03-31

We report the case of a 57-year-old patient with multiple myeloma, characterized by extramedullary involvement of the rectum at presentation. Malignant plasma cells were found to produce carcinoembryonic antigen (CEA), a tumor antigen more commonly associated with rectal adenocarcinomas.

A systems approach to designing next generation vaccines: combining α-galactose modified antigens with nanoparticle platforms

NASA Astrophysics Data System (ADS)

Phanse, Yashdeep; Carrillo-Conde, Brenda R.; Ramer-Tait, Amanda E.; Broderick, Scott; Kong, Chang Sun; Rajan, Krishna; Flick, Ramon; Mandell, Robert B.; Narasimhan, Balaji; Wannemuehler, Michael J.

2014-01-01

Innovative vaccine platforms are needed to develop effective countermeasures against emerging and re-emerging diseases. These platforms should direct antigen internalization by antigen presenting cells and promote immunogenic responses. This work describes an innovative systems approach combining two novel platforms, αGalactose (αGal)-modification of antigens and amphiphilic polyanhydride nanoparticles as vaccine delivery vehicles, to rationally design vaccine formulations. Regimens comprising soluble αGal-modified antigen and nanoparticle-encapsulated unmodified antigen induced a high titer, high avidity antibody response with broader epitope recognition of antigenic peptides than other regimen. Proliferation of antigen-specific CD4+ T cells was also enhanced compared to a traditional adjuvant. Combining the technology platforms and augmenting immune response studies with peptide arrays and informatics analysis provides a new paradigm for rational, systems-based design of next generation vaccine platforms against emerging and re-emerging pathogens.

Nanoengineering approaches to the design of artificial antigen-presenting cells

PubMed Central

Sunshine, Joel C; Green, Jordan J

2014-01-01

Artificial antigen-presenting cells (aAPCs) have shown great initial promise for ex vivo activation of cytotoxic T cells. The development of aAPCs has focused mainly on the choice of proteins to use for surface presentation to T cells when conjugated to various spherical, microscale particles. We review here biomimetic nanoengineering approaches that have been applied to the development of aAPCs that move beyond initial concepts about aAPC development. This article also discusses key technologies that may be enabling for the development of nano- and micro-scale aAPCs with nanoscale features, and suggests several future directions for the field. PMID:23837856

Type 1 Diabetes and NKT Cells: A Report on the 3rd International Workshop on NKT Cells and CD1-Mediated Antigen Presentation, September 2004, Heron Island, QLD, Australia

PubMed Central

Fletcher, Julie M.; Jordan, Margaret A.; Baxter, Alan G.

2004-01-01

NKT cells play a major role in regulating the vigor and character of a broad range of immune responses. Defects in NKT cell numbers and function have been associated with type 1 diabetes, especially in the NOD mouse model. The 3rd International Workshop on NKT Cells and CD1-Mediated Antigen Presentation provided an opportunity for researchers in the field of NKT cell biology to discuss their latest results, many of which have direct relevance to understanding the etiology and pathogenesis of diabetes. PMID:17491677

Natural Killer T Cell Activation Protects Mice Against Experimental Autoimmune Encephalomyelitis

PubMed Central

Singh, Avneesh K.; Wilson, Michael T.; Hong, Seokmann; Olivares-Villagómez, Danyvid; Du, Caigan; Stanic, Aleksandar K.; Joyce, Sebastian; Sriram, Subramaniam; Koezuka, Yasuhiko; Van Kaer, Luc

2001-01-01

Experimental autoimmune encephalomyelitis (EAE) serves as a prototypic model for T cell–mediated autoimmunity. Vα14 natural killer T (NKT) cells are a subset of T lymphocytes that recognize glycolipid antigens presented by the nonpolymorphic major histocompatibility complex (MHC) class I–like protein CD1d. Here, we show that activation of Vα14 NKT cells by the glycosphingolipid α-galactosylceramide (α-GalCer) protects susceptible mice against EAE. β-GalCer, which binds CD1d but is not recognized by NKT cells, failed to protect mice against EAE. Furthermore, α-GalCer was unable to protect CD1d knockout (KO) mice against EAE, indicating the requirement for an intact CD1d antigen presentation pathway. Protection of disease conferred by α-GalCer correlated with its ability to suppress myelin antigen-specific Th1 responses and/or to promote myelin antigen-specific Th2 cell responses. α-GalCer was unable to protect IL-4 KO and IL-10 KO mice against EAE, indicating a critical role for both of these cytokines. Because recognition of α-GalCer by NKT cells is phylogenetically conserved, our findings have identified NKT cells as novel target cells for treatment of inflammatory diseases of the central nervous system. PMID:11748281

Expression of Chicken DEC205 Reflects the Unique Structure and Function of the Avian Immune System

PubMed Central

Staines, Karen; Young, John R.; Butter, Colin

2013-01-01

The generation of appropriate adaptive immune responses relies critically on dendritic cells, about which relatively little is known in chickens, a vital livestock species, in comparison with man and mouse. We cloned and sequenced chicken DEC205 cDNA and used this knowledge to produce quantitative PCR assays and monoclonal antibodies to study expression of DEC205 as well as CD83. The gene structure of DEC205 was identical to those of other species. Transcripts of both genes were found at higher levels in lymphoid tissues and the expression of DEC205 in normal birds had a characteristic distribution in the primary lymphoid organs. In spleen, DEC205 was seen on cells ideally located to trap antigen. In thymus it was found on cells thought to participate in the education of T cells, and in the bursa on cells that may be involved in presentation of antigen to B cells and regulation of B cell migration. The expression of DEC205 on cells other than antigen presenting cells (APC) is also described. Isolated splenocytes strongly expressing DEC205 but not the KUL01 antigen have morphology similar to mammalian dendritic cells and the distinct expression of DEC205 within the avian-specific Bursa of Fabricius alludes to a unique function in this organ of B cell diversification. PMID:23326318

Cancer vaccine--Antigenics.

PubMed

2002-01-01

Antigenics is developing a therapeutic cancer vaccine based on heat-shock proteins (HSPs). The vaccine [HSPPC-96, Oncophage] is in a pivotal phase III clinical trial for renal cancer at 80 clinical sites worldwide. The trial is enrolling at least 500 patients who are randomised to receive surgical removal of the primary tumour followed by out-patient treatment with Oncophage((R)) or surgery only. This study was initiated on the basis of results from a pilot phase I/II study and preliminary results from a phase II study in patients with renal cell cancer. In October 2001, Oncophage was designated as a fast-track product by the Food and Drug Administration in the US for the treatment of renal cell carcinoma. Oncophage is in phase I/II trials in Italy for colorectal cancer (30 patients) and melanoma. The trials in Italy are being conducted at the Istituto dei Tumouri, Milan (in association with Sigma-Tau). Preliminary data from the phase II trial for melanoma was presented at the AACR-NCI-EORTC International Conference in Florida, USA, in October 2001. Oncophage is also in a phase I/II (42 patients) and a phase II trial (84 patients) in the US for renal cell cancer, a phase II trial in the US for non-Hodgkin's lymphoma (35 patients), a phase II trial in the US for sarcoma (20-35 patients), a phase I/II trial in the US for melanoma (36 patients), and phase I/II trials in Germany for gastric (30 patients) and pancreatic cancers. A pilot phase I trial in patients with pancreatic cancer began in the US in 1997 with 5 patients enrolled. In November 2000, Antigenics announced that this trial had been expanded to a phase I/II study which would now include survival as an endpoint and would enroll 5 additional patients. The US trials are being performed at Memorial Sloan-Kettering Cancer Center and the M.D. Anderson Cancer Center. The trials in Germany are being carried out at Johannes Gutenberg-University Hospital, Mainz. Oncophage is an autologous vaccine consisting of purified complexes of tumour-derived HSPs linked to tumour antigen peptides. When these HSPPC are readministered to a patient following surgery or biopsy of the tumour, the antigenic tumour peptides are expressed on the surface of potent antigen-presenting cells of the immune system, such as macrophages and dendritic cells. This stimulates a much more powerful anti-tumour immune response than that generated by expression of the same antigens by the tumour cell. Thus, Antigenics autologous HSP technology is attractive because it is highly specific for individual patients and circumvents the need for identification of specific antigens for individual cancers (i.e. it does not require definition of the antigenic epitopes on cancer cells) and it overcomes the immune tolerance associated with various tumours. Oncophage is manufactured in a 10-hour process from surgically resected autologous tumour. A minimum of 1-3g of tumour tissue is required to produce enough Oncophage for a course of treatment. The major limiting factor for producing Oncophage from a particular cancer is the ability to purify HSP from that cancer. From clinical studies to date, Antigenics has been able to produce HSP from 100, 98, 90, 71 and 30% of colorectal carcinoma, renal cell carcinoma, melanoma, gastric cancer and pancreatic cancer tumours, respectively. The low success rate with pancreatic cancers is because of the high concentration of proteases in that tissue type. HSPs are a family of highly conserved proteins present in the cells of all organisms. They function as molecular chaperones, assisting the correct folding of polypeptides and aiding intracellular protein transport. In addition, HSPs associate with a broad range of peptides derived from intracellular protein degradation, including antigenic peptides produced in tumour cells. Antigenics has exclusively licensed worldwide rights to its HSP immunotherapeutic complexes from Mount Sinai School of Medicine and Fordham University in the USA. On 3 November 1998, Antigenics was issued a US patent (5,830,464) covering immunotherapy in which antigen-presenting cells are isolated and mixed with heat shock protein-antigen complexes purified from patients' tumours. The patent was issued to Fordham University, New York, US, who subsequently licensed it to Antigenics. Antigenics has an agreement with Sigma Tau, under the terms of which the latter company will fund 2 clinical trials in return for an option to market Oncophage in Italy, Portugal, Spain and Switzerland. Antigenics also has an agreement with Medison for marketing of Oncophage in Israel.

CD1-Restricted T Cells at the Crossroad of Innate and Adaptive Immunity.

PubMed

Pereira, Catia S; Macedo, M Fatima

2016-01-01

Lipid-specific T cells comprise a group of T cells that recognize lipids bound to the MHC class I-like CD1 molecules. There are four isoforms of CD1 that are expressed at the surface of antigen presenting cells and therefore capable of presenting lipid antigens: CD1a, CD1b, CD1c, and CD1d. Each one of these isoforms has distinct structural features and cellular localizations, which promotes binding to a broad range of different types of lipids. Lipid antigens originate from either self-tissues or foreign sources, such as bacteria, fungus, or plants and their recognition by CD1-restricted T cells has important implications in infection but also in cancer and autoimmunity. In this review, we describe the characteristics of CD1 molecules and CD1-restricted lipid-specific T cells, highlighting the innate-like and adaptive-like features of different CD1-restricted T cell subtypes.

Antigen recognition in the islets changes with progression of autoimmune islet infiltration

PubMed Central

Lindsay, Robin S.; Corbin, Kaitlin; Mahne, Ashley; Levitt, Bonnie E.; Gebert, Matthew J.; Wigton, Eric J.; Bradley, Brenda J.; Haskins, Kathryn; Jacobelli, Jordan; Tang, Qizhi; Krummel, Matthew F.; Friedman, Rachel S.

2014-01-01

In type 1 diabetes, the pancreatic islets are an important site for therapeutic intervention since immune infiltration of the islets is well established at diagnosis. Therefore, understanding the events that underlie the continued progression of the autoimmune response and islet destruction is critical. Islet infiltration and destruction is an asynchronous process, making it important to analyze the disease process on a single islet basis. To understand how T cell stimulation evolves through the process of islet infiltration we analyzed the dynamics of T cell movement and interactions within individual islets of spontaneously autoimmune non-obese diabetic (NOD) mice. Using both intra-vital and explanted 2-photon islet imaging, we defined a correlation between increased islet infiltration and increased T cell motility. Early T cell arrest was antigen dependent and due, at least in part, to antigen recognition through sustained interactions with CD11c+ antigen presenting cells (APCs). As islet infiltration progressed, T cell motility became antigen-independent, with a loss of T cell arrest and sustained interactions with CD11c+ APCs. These studies suggest that the autoimmune T cell response in the islets may be temporarily dampened during the course of islet infiltration and disease progression. PMID:25505281

Impairment of Immunoproteasome Function by Cigarette Smoke and in Chronic Obstructive Pulmonary Disease.

PubMed

Kammerl, Ilona E; Dann, Angela; Mossina, Alessandra; Brech, Dorothee; Lukas, Christina; Vosyka, Oliver; Nathan, Petra; Conlon, Thomas M; Wagner, Darcy E; Overkleeft, Hermen S; Prasse, Antje; Rosas, Ivan O; Straub, Tobias; Krauss-Etschmann, Susanne; Königshoff, Melanie; Preissler, Gerhard; Winter, Hauke; Lindner, Michael; Hatz, Rudolf; Behr, Jürgen; Heinzelmann, Katharina; Yildirim, Ali Ö; Noessner, Elfriede; Eickelberg, Oliver; Meiners, Silke

2016-06-01

Patients with chronic obstructive pulmonary disease (COPD) and in particular smokers are more susceptible to respiratory infections contributing to acute exacerbations of disease. The immunoproteasome is a specialized type of proteasome destined to improve major histocompatibility complex (MHC) class I-mediated antigen presentation for the resolution of intracellular infections. To characterize immunoproteasome function in COPD and its regulation by cigarette smoke. Immunoproteasome expression and activity were determined in bronchoalveolar lavage (BAL) and lungs of human donors and patients with COPD or idiopathic pulmonary fibrosis (IPF), as well as in cigarette smoke-exposed mice. Smoke-mediated alterations of immunoproteasome activity and MHC I surface expression were analyzed in human blood-derived macrophages. Immunoproteasome-specific MHC I antigen presentation was evaluated in spleen and lung immune cells that had been smoke-exposed in vitro or in vivo. Immunoproteasome and MHC I mRNA expression was reduced in BAL cells of patients with COPD and in isolated alveolar macrophages of patients with COPD or IPF. Exposure of immune cells to cigarette smoke extract in vitro reduced immunoproteasome activity and impaired immunoproteasome-specific MHC I antigen presentation. In vivo, acute cigarette smoke exposure dynamically regulated immunoproteasome function and MHC I antigen presentation in mouse BAL cells. End-stage COPD lungs showed markedly impaired immunoproteasome activities. We here show that the activity of the immunoproteasome is impaired by cigarette smoke resulting in reduced MHC I antigen presentation. Regulation of immunoproteasome function by cigarette smoke may thus alter adaptive immune responses and add to prolonged infections and exacerbations in COPD and IPF.

PLGA nano/micro particles encapsulated with pertussis toxoid (PTd) enhances Th1/Th17 immune response in a murine model.

PubMed

Li, Pan; Asokanathan, Catpagavalli; Liu, Fang; Khaing, Kyi Kyi; Kmiec, Dorota; Wei, Xiaoqing; Song, Bing; Xing, Dorothy; Kong, Deling

2016-11-20

Poly(lactic-co-glycolic acid) (PLGA) based nano/micro particles were investigated as a potential vaccine platform for pertussis antigen. Presentation of pertussis toxoid as nano/micro particles (NP/MP) gave similar antigen-specific IgG responses in mice compared to soluble antigen. Notably, in cell line based assays, it was found that PLGA based nano/micro particles enhanced the phagocytosis of fluorescent antigen-nano/micro particles by J774.2 murine monocyte/macrophage cells compared to soluble antigen. More importantly, when mice were immunised with the antigen-nano/micro particles they significantly increased antigen-specific Th1 cytokines INF-γ and IL-17 secretion in splenocytes after in vitro re-stimulation with heat killed Bordetalla pertussis, indicating the induction of a Th1/Th17 response. Also, presentation of pertussis antigen in a NP/MP formulation is able to provide protection against respiratory infection in a murine model. Thus, the NP/MP formulation may provide an alternative to conventional acellular vaccines to achieve a more balanced Th1/Th2 immune response. Copyright © 2016 Elsevier B.V. All rights reserved.

Heterologous Prime-Boost Immunisation Regimens Against Infectious Diseases

DTIC Science & Technology

2006-08-01

of these cells by boosting. DNA vaccines are good priming agents since they are internalised by antigen presenting cells and can induce antigen...presentation via both MHC class I and class II, thereby inducing both cytotoxic T lymphocytes and type 1-helper T lymphocytes. Successful boosting agents ...assessing prime-boost vaccine combinations for protection against infectious agents . • In a number of prime - boost studies, the inclusion of growth

Antigen-Specific T-Cell Activation Independently of the MHC: Chimeric Antigen Receptor-Redirected T Cells.

PubMed

Chmielewski, Markus; Hombach, Andreas A; Abken, Hinrich

2013-01-01

Adoptive T-cell therapy has recently shown promise in initiating a lasting anti-tumor response with spectacular therapeutic success in some cases. Specific T-cell therapy, however, is limited since a number of cancer cells are not recognized by T cells due to various mechanisms including the limited availability of tumor-specific T cells and deficiencies in antigen processing or major histocompatibility complex (MHC) expression of cancer cells. To make adoptive cell therapy applicable for the broad variety of cancer entities, patient's T cells are engineered ex vivo with pre-defined specificity by a recombinant chimeric antigen receptor (CAR) which consists in the extracellular part of an antibody-derived domain for binding with a "tumor-associated antigen" and in the intracellular part of a T-cell receptor (TCR)-derived signaling moiety for T-cell activation. The specificity of CAR-mediated T-cell recognition is defined by the antibody domain, is independent of MHC presentation and can be extended to any target for which an antibody is available. We discuss the advantages and limitations of MHC-independent T-cell targeting by an engineered CAR in comparison to TCR modified T cells and the impact of the CAR activation threshold on redirected T-cell activation. Finally we review most significant progress recently made in early stage clinical trials to treat cancer.

Malassezia yeasts activate the NLRP3 inflammasome in antigen-presenting cells via Syk-kinase signalling.

PubMed

Kistowska, Magdalena; Fenini, Gabriele; Jankovic, Dragana; Feldmeyer, Laurence; Kerl, Katrin; Bosshard, Philipp; Contassot, Emmanuel; French, Lars E

2014-12-01

Although being a normal part of the skin flora, yeasts of the genus Malassezia are associated with several common dermatologic conditions including pityriasis versicolour, seborrhoeic dermatitis (SD), folliculitis, atopic eczema/dermatitis (AE/AD) and dandruff. While Malassezia spp. are aetiological agents of pityriasis versicolour, a causal role of Malassezia spp. in AE/AD and SD remains to be established. Previous reports have shown that fungi such as Candida albicans and Aspergillus fumigatus are able to efficiently activate the NLRP3 inflammasome leading to robust secretion of the pro-inflammatory cytokine IL-1β. To date, innate immune responses to Malassezia spp. are not well characterized. Here, we show that different Malassezia species could induce NLRP3 inflammasome activation and subsequent IL-1β secretion in human antigen-presenting cells. In contrast, keratinocytes were not able to secrete IL-1β when exposed to Malassezia spp. Moreover, we demonstrate that IL-1β secretion in antigen-presenting cells was dependent on Syk-kinase signalling. Our results identify Malassezia spp. as potential strong inducers of pro-inflammatory responses when taken up by antigen-presenting cells and identify C-type lectin receptors and the NLRP3 inflammasome as crucial actors in this process. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Oral dendritic cells mediate antigen-specific tolerance by stimulating TH1 and regulatory CD4+ T cells.

PubMed

Mascarell, Laurent; Lombardi, Vincent; Louise, Anne; Saint-Lu, Nathalie; Chabre, Henri; Moussu, Hélène; Betbeder, Didier; Balazuc, Anne-Marie; Van Overtvelt, Laurence; Moingeon, Philippe

2008-09-01

A detailed characterization of oral antigen-presenting cells is critical to improve second-generation sublingual allergy vaccines. To characterize oral dendritic cells (DCs) within lingual and buccal tissues from BALB/c mice with respect to their surface phenotype, distribution, and capacity to polarize CD4(+) T-cell responses. In situ analysis of oral DCs was performed by immunohistology. Purified DCs were tested in vitro for their capacity to capture, process, and present the ovalbumin antigen to naive CD4(+) T cells. In vivo priming of ovalbumin-specific T cells adoptively transferred to BALB/c mice was analyzed by cytofluorometry in cervical lymph nodes after sublingual administration of mucoadhesive ovalbumin. Three subsets of oral DCs with a distinct tissue distribution were identified: (1) a minor subset of CD207(+) Langerhans cells located in the mucosa itself, (2) a major subpopulation of CD11b(+)CD11c(-) and CD11b(+)CD11c(+) myeloid DCs at the mucosal/submucosal interface, and (3) B220(+)120G8(+) plasmacytoid DCs found in submucosal tissues. Purified myeloid and plasmacytoid oral DCs capture and process the antigen efficiently and are programmed to elicit IFN-gamma and/or IL-10 production together with a suppressive function in naive CD4(+) T cells. Targeting the ovalbumin antigen to oral DCs in vivo by using mucoadhesive particles establishes tolerance in the absence of cell depletion through the stimulation of IFN-gamma and IL-10-producing CD4(+) regulatory T cells in cervical lymph nodes. The oral immune system is composed of various subsets of tolerogenic DCs organized in a compartmentalized manner and programmed to induce T(H)1/regulatory T-cell responses.

Murine T-Cell Response to Native and Recombinant Protein Antigens of Rickettsia Tsutsugamushi

DTIC Science & Technology

1993-02-01

Wright, and J. Sadoff. 1985. 18-kilodalton protein of Mycobacterium leprae recognized by Immunoenzymatic analysis by monoclonal antibodies of bacte- Vo...determinants and closely resembles T-cell antigenic determinants, Rothbard and Taylor, by the GroEL homolog (65 kDa) of Mycobacterium tuberculo- analysis of...not be completely present in protein that is recognized by 20% of the mycobacterium - peptide 91-110. If this were the core of the antigenic deter

Histological study of cell migration in the dermis of hamsters after immunisation with two different vaccines against visceral leishmaniasis.

PubMed

Moreira, Nádia das Dores; Giunchetti, Rodolfo Cordeiro; Carneiro, Cláudia Martins; Vitoriano-Souza, Juliana; Roatt, Bruno Mendes; Malaquias, Luiz Cosme Cotta; Corrêa-Oliveira, Rodrigo; Reis, Alexandre Barbosa

2009-04-15

Vaccine candidates, including live and/or killed parasites, Leishmania-purified fractions, defined recombinant antigens and antigen-encoding DNA-plasmids have been proposed to use as vaccine anti-Leishmania. More recently, the hamsters have been used to pre-selection of antigens candidate to apply in further experiments using canine model. In this report we evaluated the kinetics of cell migration in dermal inflammatory infiltrate, circulating leukocytes and the presence of nitric oxide (NO)/induced nitric oxide synthase during the early (1-24h) and late (48-168h) periods following inoculation of hamsters with antigenic components of anti-canine visceral leishmaniasis vaccines Leishmune and Leishmania braziliensis antigen (LB) with and without saponin (Sap) adjuvant. Our results show that LB caused an early reduction of lymphocytes in the dermis while Sap and LBSap triggered a late recruitment, suggesting the role of the adjuvant in the traffic of antigen-presenting cells and the induction of lymphocyte migration. In that manner our results suggest that the kinetics of cell migration on hamster model may be of value in the selection of vaccine antigens prior the tests in dogs particularly in respect of the toxicity of the preparations.

Multiple myeloma presenting as CEA-producing rectal cancer

PubMed Central

Talamo, Giampaolo; Barochia, Amitkumar; Zangari, Maurizio; Loughran, Thomas P

2010-01-01

We report the case of a 57-year-old patient with multiple myeloma, characterized by extramedullary involvement of the rectum at presentation. Malignant plasma cells were found to produce carcinoembryonic antigen (CEA), a tumor antigen more commonly associated with rectal adenocarcinomas. PMID:21139949

Serine Proteases Enhance Immunogenic Antigen Presentation on Lung Cancer Cells.

PubMed

Peters, Haley L; Tripathi, Satyendra C; Kerros, Celine; Katayama, Hiroyuki; Garber, Haven R; St John, Lisa S; Federico, Lorenzo; Meraz, Ismail M; Roth, Jack A; Sepesi, Boris; Majidi, Mourad; Ruisaard, Kathryn; Clise-Dwyer, Karen; Roszik, Jason; Gibbons, Don L; Heymach, John V; Swisher, Stephen G; Bernatchez, Chantale; Alatrash, Gheath; Hanash, Samir; Molldrem, Jeffrey J

2017-04-01

Immunotherapies targeting immune checkpoints have proven efficacious in reducing the burden of lung cancer in patients; however, the antigenic targets of these reinvigorated T cells remain poorly defined. Lung cancer tumors contain tumor-associated macrophages (TAM) and neutrophils, which release the serine proteases neutrophil elastase (NE) and proteinase 3 (P3) into the tumor microenvironment. NE and P3 shape the antitumor adaptive immune response in breast cancer and melanoma. In this report, we demonstrate that lung cancer cells cross-presented the tumor-associated antigen PR1, derived from NE and P3. Additionally, NE and P3 enhanced the expression of human leukocyte antigen (HLA) class I molecules on lung cancer cells and induced unique, endogenous peptides in the immunopeptidome, as detected with mass spectrometry sequencing. Lung cancer patient tissues with high intratumoral TAMs were enriched for MHC class I genes and T-cell markers, and patients with high TAM and cytotoxic T lymphocyte (CTL) infiltration had improved overall survival. We confirmed the immunogenicity of unique, endogenous peptides with cytotoxicity assays against lung cancer cell lines, using CTLs from healthy donors that had been expanded against select peptides. Finally, CTLs specific for serine proteases-induced endogenous peptides were detected in lung cancer patients using peptide/HLA-A2 tetramers and were elevated in tumor-infiltrating lymphocytes. Thus, serine proteases in the tumor microenvironment of lung cancers promote the presentation of HLA class I immunogenic peptides that are expressed by lung cancer cells, thereby increasing the antigen repertoire that can be targeted in lung cancer. Cancer Immunol Res; 5(4); 319-29. ©2017 AACR . ©2017 American Association for Cancer Research.

Streptococcus suis Serotype 2 Infection Impairs Interleukin-12 Production and the MHC-II-Restricted Antigen Presentation Capacity of Dendritic Cells

PubMed Central

Letendre, Corinne; Auger, Jean-Philippe; Lemire, Paul; Galbas, Tristan; Gottschalk, Marcelo; Thibodeau, Jacques; Segura, Mariela

2018-01-01

Streptococcus suis is an important swine pathogen and emerging zoonotic agent. Encapsulated strains of S. suis modulate dendritic cell (DC) functions, leading to poorly activated CD4+ T cells. However, the antigen presentation ability of S. suis-stimulated DCs has not been investigated yet. In this work, we aimed to characterize the antigen presentation profiles of S. suis-stimulated DCs, both in vitro and in vivo. Upon direct activation in vitro, S. suis-stimulated murine bone marrow-derived DCs (bmDCs) preserved their antigen capture/processing capacities. However, they showed delayed kinetics of MHC-II expression compared to lipopolysaccharide-stimulated bmDCs. Meanwhile, splenic DCs from infected mice exhibited a compromised MHC-II expression, despite an appropriate expression of maturation markers. To identify potential interfering mechanisms, Class II Major Histocompatibility Complex Transactivator (CIITA) and membrane-associated RING-CH (MARCH)1/8 transcription were studied. S. suis-stimulated DCs maintained low levels of CIITA at early time points, both in vitro and in vivo, which could limit their ability to increase MHC-II synthesis. S. suis-stimulated DCs also displayed sustained/upregulated levels of MARCH1/8, thus possibly leading to MHC-II lysosomal degradation. The bacterial capsular polysaccharide played a partial role in this modulation. Finally, interleukin (IL)-12p70 production was inhibited in splenic DCs from infected mice, a profile compatible with DC indirect activation by pro-inflammatory compounds. Consequently, these cells induced lower levels of IL-2 and TNF-α in an antigen-specific CD4+ T cell presentation assay and blunted T cell CD25 expression. It remains unclear at this stage whether these phenotypical and transcriptional modulations observed in response to S. suis in in vivo infections are part of a bacterial immune evasion strategy or rather a feature common to systemic inflammatory response-inducing agents. However, it appears that the MHC-II-restricted antigen presentation and Th1-polarizing cytokine production capacities of DCs are impaired during S. suis infection. This study highlights the potential consequences of inflammation on the type and magnitude of the immune response elicited by a pathogen. PMID:29899744

A mammalian tRNAHis-containing antigen is recognized by the polymyositis-specific antibody anti-Jo-1.

PubMed Central

Rosa, M D; Hendrick, J P; Lerner, M R; Steitz, J A; Reichlin, M

1983-01-01

The mammalian cell antigen reactive with the autoantibody anti-Jo-1 has been shown to contain tRNAHis. The RNA sequence of this human and mouse cell tRNA was determined in a search for unusual features that might be related to antigenicity. The 5' terminal nucleotide is unique among other sequenced tRNAs in that it is a methylated guanine. The presence of the hypermodified base queuine, which occurs in the wobble position of the anticodon of tRNAHis from several species, was not detected in the tRNAHis immunoprecipitated by anti-Jo-1 from either human HeLa or mouse Friend erytholeukemia cell extracts. The binding of protein(s) appears to confer antigenicity on tRNAHis since either proteinase K treatment or phenol extraction resulted in the loss of immunoprecipitability. However, we have not succeeded in identifying an antigenic protein, and we find that the antigenic complex is not resolved from purified tRNAHis by Sephacryl S-200 column chromatography. Immunofluorescence studies indicate that the antigenic form of tRNAHis is located preferentially in the mammalian cell cytoplasm. The results presented here are discussed in light of an earlier report (1) on the nature of the Jo-1 antigen. Images PMID:6188108

T lymphocyte-derived TNF and IFN-γ repress HFE expression in cancer cells.

PubMed

Reuben, Alexandre; Godin-Ethier, Jessica; Santos, Manuela M; Lapointe, Réjean

2015-06-01

The immune system and tumors are closely intertwined initially upon tumor development. During this period, tumors evolve to promote self-survival through immune escape, including by targeting crucial components involved in the presentation of antigens to the immune system in order to avoid recognition. Accordingly, components involved in MHC I presentation of tumor antigens are often mutated and down-regulated targets in tumors. On the other hand, the immune system has been shown to influence tumors through production of immunosuppressive cytokines, recruitment and polarization of cells favoring or impeding tumor escape or through production of anti-tumor cytokines promoting tumor rejection. We previously discovered that the hemochromatosis protein HFE, a negative regulator of iron absorption, dampens classical MHC I antigen presentation. In this study, we evaluated the impact of activated T lymphocytes purified from peripheral blood mononuclear cells (PBMC) on HFE expression in tumor cell lines. We co-cultured tumor cell lines from melanoma, lung, and kidney cancers with anti-CD3-activated PBMC and established that HFE expression is increased in tumor cell lines compared to healthy tissues, whilst being down-regulated significantly upon exposure to activated PBMC. HFE down-regulation was mediated by both CD4 and CD8 T lymphocytes, through production of soluble mediators, namely TNF and IFN-γ. These results suggest that the immune system may modulate tumor HFE expression in inflammatory conditions in order to regulate MHC I antigen presentation and promote tumor clearance. Copyright © 2015. Published by Elsevier Ltd.

Coevolution of T-cell receptors with MHC and non-MHC ligands

PubMed Central

Castro, Caitlin C.; Luoma, Adrienne M.; Adams, Erin J.

2015-01-01

Summary The structure and amino acid diversity of the T-cell receptor (TCR), similar in nature to that of Fab portions of antibodies, would suggest these proteins have a nearly infinite capacity to recognize antigen. Yet all currently defined native T cells expressing an α and β chain in their TCR can only sense antigen when presented in the context of a major histocompatibility complex (MHC) molecule. This MHC molecule can be one of many that exist in vertebrates, presenting small peptide fragments, lipid molecules, or small molecule metabolites. Here we review the pattern of TCR recognition of MHC molecules throughout a broad sampling of species and T-cell lineages and also touch upon T cells that do not appear to require MHC presentation for their surveillance function. We review the diversity of MHC molecules and information on the corresponding T-cell lineages identified in divergent species. We also discuss TCRs with structural domains unlike that of conventional TCRs of mouse and human. By presenting this broad view of TCR sequence, structure, domain organization, and function, we seek to explore how this receptor has evolved across time and been selected for alternative antigen-recognition capabilities in divergent lineages. PMID:26284470

Immunotherapy and immunoescape in colorectal cancer

PubMed Central

Mazzolini, Guillermo; Murillo, Oihana; Atorrasagasti, Catalina; Dubrot, Juan; Tirapu, Iñigo; Rizzo, Miguel; Arina, Ainhoa; Alfaro, Carlos; Azpilicueta, Arantza; Berasain, Carmen; Perez-Gracia, José L; Gonzalez, Alvaro; Melero, Ignacio

2007-01-01

Immunotherapy encompasses a variety of interventions and techniques with the common goal of eliciting tumor cell destructive immune responses. Colorectal carcinoma often presents as metastatic disease that impedes curative surgery. Novel strategies such as active immunization with dendritic cells (DCs), gene transfer of cytokines into tumor cells or administration of immunostimulatory monoclonal antibodies (such as anti-CD137 or anti-CTLA-4) have been assessed in preclinical studies and are at an early clinical development stage. Importantly, there is accumulating evidence that chemotherapy and immunotherapy can be combined in the treatment of some cases with colorectal cancer, with synergistic potentiation as a result of antigens cross-presented by dendritic cells and/or elimination of competitor or suppressive T lymphocyte populations (regulatory T-cells). However, genetic and epigenetic unstable carcinoma cells frequently evolve mechanisms of immunoevasion that are the result of either loss of antigen presentation, or an active expression of immunosuppressive substances. Some of these actively immunosuppressive mechanisms are inducible by cytokines that signify the arrival of an effector immune response. For example, induction of 2, 3 indoleamine dioxygenase (IDO) by IFNγ in colorectal carcinoma cells. Combinational and balanced strategies fostering antigen presentation, T-cell costimulation and interference with immune regulatory mechanisms will probably take the stage in translational research in the treatment of colorectal carcinoma. PMID:17990348

A CD22-reactive TCR from the T-cell allorepertoire for the treatment of acute lymphoblastic leukemia by TCR gene transfer

PubMed Central

Jahn, Lorenz; Hagedoorn, Renate S.; van der Steen, Dirk M.; Hombrink, Pleun; Kester, Michel G.D.; Schoonakker, Marjolein P.; de Ridder, Daniëlle; van Veelen, Peter A.; Falkenburg, J.H. Frederik; Heemskerk, Mirjam H.M.

2016-01-01

CD22 is currently evaluated as a target-antigen for the treatment of B-cell malignancies using chimeric antigen receptor (CAR)-engineered T-cells or monoclonal antibodies (mAbs). CAR- and mAbs-based immunotherapies have been successfully applied targeting other antigens, however, occurrence of refractory disease to these interventions urges the identification of additional strategies. Here, we identified a TCR recognizing the CD22-derived peptide RPFPPHIQL (CD22RPF) presented in human leukocyte antigen (HLA)-B*07:02. To overcome tolerance to self-antigens such as CD22, we exploited the immunogenicity of allogeneic HLA. CD22RPF-specific T-cell clone 9D4 was isolated from a healthy HLA-B*07:02neg individual, efficiently produced cytokines upon stimulation with primary acute lymphoblastic leukemia and healthy B-cells, but did not react towards healthy hematopoietic and nonhematopoietic cell subsets, including dendritic cells (DCs) and macrophages expressing low levels of CD22. Gene transfer of TCR-9D4 installed potent CD22-specificity onto recipient CD8+ T-cells that recognized and lysed primary B-cell leukemia. TCR-transduced T-cells spared healthy CD22neg hematopoietic cell subsets but weakly lysed CD22low-expressing DCs and macrophages. CD22-specific TCR-engineered T-cells could form an additional immunotherapeutic strategy with a complementary role to CAR- and antibody-based interventions in the treatment of B-cell malignancies. However, CD22 expression on non-B-cells may limit the attractiveness of CD22 as target-antigen in cellular immunotherapy. PMID:27689397

A CD22-reactive TCR from the T-cell allorepertoire for the treatment of acute lymphoblastic leukemia by TCR gene transfer.

PubMed

Jahn, Lorenz; Hagedoorn, Renate S; van der Steen, Dirk M; Hombrink, Pleun; Kester, Michel G D; Schoonakker, Marjolein P; de Ridder, Daniëlle; van Veelen, Peter A; Falkenburg, J H Frederik; Heemskerk, Mirjam H M

2016-11-01

CD22 is currently evaluated as a target-antigen for the treatment of B-cell malignancies using chimeric antigen receptor (CAR)-engineered T-cells or monoclonal antibodies (mAbs). CAR- and mAbs-based immunotherapies have been successfully applied targeting other antigens, however, occurrence of refractory disease to these interventions urges the identification of additional strategies. Here, we identified a TCR recognizing the CD22-derived peptide RPFPPHIQL (CD22RPF) presented in human leukocyte antigen (HLA)-B*07:02. To overcome tolerance to self-antigens such as CD22, we exploited the immunogenicity of allogeneic HLA. CD22RPF-specific T-cell clone 9D4 was isolated from a healthy HLA-B*07:02neg individual, efficiently produced cytokines upon stimulation with primary acute lymphoblastic leukemia and healthy B-cells, but did not react towards healthy hematopoietic and nonhematopoietic cell subsets, including dendritic cells (DCs) and macrophages expressing low levels of CD22. Gene transfer of TCR-9D4 installed potent CD22-specificity onto recipient CD8+ T-cells that recognized and lysed primary B-cell leukemia. TCR-transduced T-cells spared healthy CD22neg hematopoietic cell subsets but weakly lysed CD22low-expressing DCs and macrophages. CD22-specific TCR-engineered T-cells could form an additional immunotherapeutic strategy with a complementary role to CAR- and antibody-based interventions in the treatment of B-cell malignancies. However, CD22 expression on non-B-cells may limit the attractiveness of CD22 as target-antigen in cellular immunotherapy.

In vitro immunization of patient T cells with autologous bone marrow antigen presenting cells pulsed with tumor lysates.

PubMed

Coulon, V; Ravaud, A; Gaston, R; Delaunay, M; Pariente, J L; Verdier, D; Scrivante, V; Gualde, N

2000-12-01

Presentation of cell-associated antigen to T cells is a critical event in the initiation of an anti-tumor immune response but it appears to often be deficient or limiting. Here we report an experimental system for stimulation of human T lymphocytes using autologous antigen presenting cells (APCs) and autologous tumor cells. Two types of APCs were prepared from human bone marrow: MC and DC. MC were produced by using GM-CSF and SCF. DC were obtained with the same cytokines plus IL-4. DC and MC were generated in parallel from the same patients and their phenotypes and capacities to prime T lymphocytes were analyzed and compared. MC were CD14+, CD1a-, CD33+ and HLA-DR+. Two populations of DC were defined: immature DC were uniformly CD1a-; mature DC expressed CD1a, CD80, CD86, HLA-DR, CD54 and CD58 but lacked surface CD14. Stimulation of autologous T lymphocytes was studied by measuring their proliferation and cytotoxic function. In more than 80% of our experiments the proliferation of autologous T lymphocytes cocultured with APC pulsed or not with tumor cell lysates was higher than that of T cells cultured alone. DC were more effective than MC in stimulating proliferation of lymphocytes. The capacity of a patient's autologous bone marrow-derived APC to stimulate T cells when exposed to autologous tumor cell lysates suggest that such antigen-exposed APC may be useful in specific anti-tumor immunotherapy protocols. Copyright 2000 Wiley-Liss, Inc.

T suppressor cells are required for the maintenance of the antigen-induced B-cell unresponsive state in humans

DOE Office of Scientific and Technical Information (OSTI.GOV)

Benveniste, E.; Stevens, R.H.

1983-04-01

Tetanus toxoid immunization of humans generates circulating B cells which secrete IgG anti-tetanus toxoid antibodies (IgG-Tet) when stimulated in vitro with T cells and pokeweed mitogen (PWM). A unique property of these cells is the inhibition of maturation into antibody-secreting plasma cells following a 1-hr in vitro pulse with tetanus toxoid. Studies were undertaken to determine if different T-cell subsets could modulate the in vitro generated B-cell unresponsive state. The addition of OKT4+/OKT8- cells to antigen-treated B cells resulted in a partial reversal of the antigen-induced inhibition of IgG-Tet synthesis. The addition of OKT4-/OKT8+ cells to the treated B cellsmore » caused a suppression of IgG-Tet synthesis comparable to that seen in cultures containing unfractionated T cells. These results indicate that (1) the B-cell unresponsive state generated by antigen treatment is not absolute, (2) the degree of B-cell unresponsiveness results from a balance of suppressor and helper signals, and (3) T-suppressor cells need to be present to induce and maintain the B-cell unresponsive state.« less

Specific Schistosoma mansoni rat T cell clones. I. Generation and functional analysis in vitro and in vivo.

PubMed

Pestel, J; Dissous, C; Dessaint, J P; Louis, J; Engers, H; Capron, A

1985-06-01

In an attempt to determine the role of schistosome-specific T cells in the immune mechanisms developed during schistosomiasis, Schistosoma mansoni-specific T cells and clones were generated in vitro and some of their functions analyzed in vitro and in vivo in the fischer rat model. The data presented here can be summarized as follows: a) Lymph node cells (LNC) from rats primed with the excretory/secretory antigens-incubation products (IPSm) of adult worms proliferate in vitro only in response to the homologous schistosome antigens and not to unrelated antigens (Ag) such as ovalbumin (OVA) or Dipetalonema viteae and Fasciola hepatica parasite extracts. b) After in vitro restimulation of the primed LNC population with IPSm in the presence of antigen-presenting cells (APC) and maintenance in IL 2-containing medium, the frequency of IPSm-specific T cells is increased and the T cells can be restimulated only in the presence of APC possessing the same major histocompatibility complex (MHC) antigens. c) Following appropriate limiting dilution assays (LDA) (1 cell/well), 10 IPSm-specific T cell clones were obtained, and two of four maintained in culture were tested for their helper activity because they expressed only the W3/13+ W3/25+ surface phenotypes. d) The two highly proliferating IPSm-specific T cell clones (G5 and E23) exhibit an IPSm-dependent helper activity, as shown by the increase in IgG production by IPSm-primed B cells. e) IPSm-T cell clone (G5) as well as IPSm-T cell lines when injected in S. mansoni-infested rats can exert an in vivo helper activity, which is characterized by an accelerated production of IgG antibodies specific for the previously identified 30 to 40 kilodaltons (kd) schistosomula surface antigens (Ag). As recent studies have demonstrated that rat monoclonal antibodies recognize some incubation products of adult S. mansoni as well as one of the 30 to 40 kd schistosomula surface antigens, and taking into account the fact that the T cell clones here studied were restimulated either with IPSm or with schistosomulum Ag, it appears that such IPSm-specific T cell clones could be involved in the concomitant immunity mechanisms.

Perspectives for immunotherapy in glioblastoma treatment.

PubMed

Finocchiaro, Gaetano; Pellegatta, Serena

2014-11-01

Avoiding immune destruction is one emerging hallmark of cancer, including glioblastoma. The number of immunotherapy approaches to fight glioblastoma is growing. Here, we review the recent progress in four main areas: dendritic cell immunotherapy, peptide vaccination, chimeric antigen receptors and immune checkpoints. We and others are using dendritic cells to present glioblastoma antigens (whole tumor lysate) to the immune system; our initial data indicate that clinical benefit is associated to increased presence of natural killer cells in the periphery. A pilot study loading dendritic cells with glioblastoma stem-like cells will start soon. Peptide vaccination targeting the epidermal growth factor receptor variant III (EGFRvIII) epitope, present in 25% of glioblastomas, is ongoing. Intriguing results have been obtained by vaccination with three other peptides in pediatric gliomas. Another clinical trial is targeting EGFRvIII by adoptive cell transfer of chimeric antigen receptor. This exciting technology could be suited for a number of other potential epitopes discovered through next-generation sequencing. Finally, antibodies against the immune checkpoints cytotoxic T lymphocyte antigen-4 and programmed cell death-1, which demonstrated efficacy in advanced melanomas, will be used in novel trials for recurrent glioblastoma. In all these studies attention to novel side-effects and to MRI as immunological follow-up to distinguish progression or pseudoprogression will be of critical relevance.

In Situ Modulation of Dendritic Cells by Injectable Thermosensitive Hydrogels for Cancer Vaccines in Mice

PubMed Central

2014-01-01

Attempts to develop cell-based cancer vaccines have shown limited efficacy, partly because transplanted dendritic cells (DCs) do not survive long enough to reach the lymph nodes. The development of biomaterials capable of modulating DCs in situ to enhance antigen uptake and presentation has emerged as a novel method toward developing more efficient cancer vaccines. Here, we propose a two-step hybrid strategy to produce a more robust cell-based cancer vaccine in situ. First, a significant number of DCs are recruited to an injectable thermosensitive mPEG–PLGA hydrogel through sustained release of chemoattractants, in particular, granulocyte-macrophage colony-stimulating factor (GM-CSF). Then, these resident DCs can be loaded with cancer antigens through the use of viral or nonviral vectors. We demonstrate that GM-CSF-releasing mPEG–PLGA hydrogels successfully recruit and house DCs and macrophages, allowing the subsequent introduction of antigens by vectors to activate the resident cells, thus, initiating antigen presentation and triggering immune response. Moreover, this two-step hybrid strategy generates a high level of tumor-specific immunity, as demonstrated in both prophylactic and therapeutic models of murine melanoma. This injectable thermosensitive hydrogel shows great promise as an adjuvant for cancer vaccines, potentially providing a new approach for cell therapies through in situ modulation of cells. PMID:25207465

Involvement of dendritic cells in allograft rejection new implications of dendritic cell-endothelial cell interactions.

PubMed

Schlichting, C L; Schareck, W D; Kofler, S; Weis, M

2007-04-01

For almost half a century immunologists have tried to tear down the MHC barrier, which separates two unrelated individuals during transplantation. Latest experimental data suggest that a breakthrough in vitro is imminent. Dendritic cells (DCs), which activate naïve allo-reactive T-cells (TCs), play a central role in the establishment of allo-antigen-specific immunity. Allograft solid organ rejection is initiated at the foreign endothelial cell (EC) layer, which forms an immunogenic barrier for migrating DCs. Thus, DC/EC interactions might play a crucial role in antigen-specific allograft rejection. Organ rejection is mediated by host allo-reactive TCs, which are activated by donor DCs (direct activation) or host DCs (indirect activation). Direct allo-antigen presentation by regulatory dendritic cells (DCreg) can play an instructive role towards tolerance induction. Several groups established that, DCregs, if transplanted beforehand, enter host thymus, spleen, or bone marrow where they might eventually establish allo-antigen-specific tolerance. A fundamental aspect of DC function is migration throughout the entire organism. After solid organ transplantation, host DCs bind to ECs, invade allograft tissues, and finally transmigrate into lymphoid vessels and secondary lymphoid organs, where they present allo-antigens to naïve host TCs. Recent data suggest that in vitro manipulated DCregs may mediate allo-transplantation tolerance induction. However, the fundamental mechanisms on how such DCregs cause host TCs in the periphery towards tolerance remain unclear. One very promising experimental concept is the simultaneous manipulation of DC direct and indirect TC activation/suppression, towards donor antigen-specific allo-transplantation tolerance. The allo-antigen-specific long-term tolerance induction mediated by DCreg pre-transplantation (with simultaneous short-term immunosuppression) has become reproducible in the laboratory animal setting. Despite the shortcomings of laboratory animal studies, strong promises are deriving from these studies for clinical kidney, heart, and liver transplantation.

Cinnamon extract suppresses experimental colitis through modulation of antigen-presenting cells.

PubMed

Kwon, Ho-Keun; Hwang, Ji-Sun; Lee, Choong-Gu; So, Jae-Seon; Sahoo, Anupama; Im, Chang-Rok; Jeon, Won Kyung; Ko, Byoung Seob; Lee, Sung Haeng; Park, Zee Yong; Im, Sin-Hyeog

2011-02-28

To investigate the anti-inflammatory effects of cinnamon extract and elucidate its mechanisms for targeting the function of antigen presenting cells. Cinnamon extract was used to treat murine macrophage cell line (Raw 264.7), mouse primary antigen-presenting cells (APCs, MHCII(+)) and CD11c(+) dendritic cells to analyze the effects of cinnamon extract on APC function. The mechanisms of action of cinnamon extract on APCs were investigated by analyzing cytokine production, and expression of MHC antigens and co-stimulatory molecules by quantitative real-time PCR and flow cytometry. In addition, the effect of cinnamon extract on antigen presentation capacity and APC-dependent T-cell differentiation were analyzed by [H(3)]-thymidine incorporation and cytokine analysis, respectively. To confirm the anti-inflammatory effects of cinnamon extract in vivo, cinnamon or PBS was orally administered to mice for 20 d followed by induction of experimental colitis with 2,4,6 trinitrobenzenesulfonic acid. The protective effects of cinnamon extract against experimental colitis were measured by checking clinical symptoms, histological analysis and cytokine expression profiles in inflamed tissue. Treatment with cinnamon extract inhibited maturation of MHCII(+) APCs or CD11c(+) dendritic cells (DCs) by suppressing expression of co-stimulatory molecules (B7.1, B7.2, ICOS-L), MHCII and cyclooxygenase (COX)-2. Cinnamon extract induced regulatory DCs (rDCs) that produce low levels of pro-inflammatory cytokines [interleukin (IL)-1β, IL-6, IL-12, interferon (IFN)-γ and tumor necrosis factor (TNF)-α] while expressing high levels of immunoregulatory cytokines (IL-10 and transforming growth factor-β). In addition, rDCs generated by cinnamon extract inhibited APC-dependent T-cell proliferation, and converted CD4(+) T cells into IL-10(high) CD4(+) T cells. Furthermore, oral administration of cinnamon extract inhibited development and progression of intestinal colitis by inhibiting expression of COX-2 and pro-inflammatory cytokines (IL-1β, IFN-γ and TNF-α), while enhancing IL-10 levels. Our study suggests the potential of cinnamon extract as an anti-inflammatory agent by targeting the generation of regulatory APCs and IL-10(+) regulatory T cells.

Isolation and partial characterization of melanoma-associated antigens identified by autologous antibody.

PubMed

Vlock, D R; Scalise, D; Meglin, N; Kirkwood, J M; Ballou, B

1988-06-01

The study of the autologous immune response to cancer avoids the difficulties encountered in the use of xenoantisera and may identify antigens of physiological relevance. However, the low titer and incidence of autologous antibody to melanoma have hampered further evaluation. By utilizing acid dissociation and ultrafiltration of serum, we have been able to augment the detectable autologous immune response to melanoma in the majority of patients studied. In autologous system Y-Mel 84:420, serum S150 demonstrated a rise in titer from 1:32 in native sera to 1:262,044 after dissociation. The antigen detected by S150 was found to be broadly represented on melanoma, glioma, renal cell carcinoma, neuroblastoma, and head and neck carcinoma cell lines. It did not react with bladder or colon carcinoma, fetal fibroblasts, pooled platelets, lymphocytes and red blood cells, or autologous cultured lymphocytes. Using polyacrylamide gel electrophoresis, S150 detects a 66,000-mol wt antigen in spent tissue culture media and serum ultrafiltrate. In cell lysate two bands between 20,000 and 30,000 mol wt are detected by S150. The 66,000-mol wt antigen is sensitive to trypsin digestion and but is resistant to pepsin and heat inactivation. Exposure of spent media to trypsin results in the development of a 24,000-mol wt band that appears to correspond to the antigen detected in the cell lysate. The difference between the antigens detected in the cell lysate as compared with spent media and serum ultrafiltrate may be due to degradation during cell lysis. We conclude that melanoma-associated antigens are present in the serum of patients with melanoma and are shed or secreted by their tumor cells.

Antigen presentation under the influence of 'immune evasion' proteins and its modulation by interferon-gamma: implications for immunotherapy of cytomegalovirus infection with antiviral CD8 T cells.

PubMed

Fink, Annette; Lemmermann, Niels A W; Gillert-Marien, Dorothea; Thomas, Doris; Freitag, Kirsten; Böhm, Verena; Wilhelmi, Vanessa; Reifenberg, Kurt; Reddehase, Matthias J; Holtappels, Rafaela

2012-11-01

Cytomegalovirus (CMV) disease with multiple organ manifestations is the most feared viral complication limiting the success of hematopoietic cell transplantation as a therapy of hematopoietic malignancies. A timely endogenous reconstitution of CD8 T cells controls CMV infection, and adoptive transfer of antiviral CD8 T cells is a therapeutic option to prevent CMV disease by bridging the gap between an early CMV reactivation and delayed endogenous reconstitution of protective immunity. Preclinical research in murine models has provided 'proof of concept' for CD8 T-cell therapy of CMV disease. Protection by CD8 T cells appears to be in conflict with the finding that CMVs encode proteins that inhibit antigen presentation to CD8 T cells by interfering with the constitutive trafficking of peptide-loaded MHC class I molecules (pMHC-I complexes) to the cell surface. Here, we have systematically explored antigen presentation in the presence of the three currently noted immune evasion proteins of murine CMV in all possible combinations and its modulation by pre-treatment of cells with interferon-gamma (IFN-γ). The data reveal improvement in antigen processing by pre-treatment with IFN-γ can almost overrule the inhibitory function of immune evasion molecules in terms of pMHC-I expression levels capable of triggering most of the specific CD8 T cells, though the intensity of stimulation did not retrieve their full functional capacity. Notably, an in vivo conditioning of host tissue cells with IFN-γ in adoptive cell transfer recipients constitutively overexpressing IFN-γ (B6-SAP-IFN-γ mice) enhanced the antiviral efficiency of CD8 T cells in this transgenic cytoimmunotherapy model.

Connective tissue growth factor linked to the E7 tumor antigen generates potent antitumor immune responses mediated by an antiapoptotic mechanism.

PubMed

Cheng, W-F; Chang, M-C; Sun, W-Z; Lee, C-N; Lin, H-W; Su, Y-N; Hsieh, C-Y; Chen, C-A

2008-07-01

A novel method for generating an antigen-specific cancer vaccine and immunotherapy has emerged using a DNA vaccine. However, antigen-presenting cells (APCs) have a limited life span, which hinders their long-term ability to prime antigen-specific T cells. Connective tissue growth factor (CTGF) has a role in cell survival. This study explored the intradermal administration of DNA encoding CTGF with a model tumor antigen, human papilloma virus type 16 E7. Mice vaccinated with CTGF/E7 DNA exhibited a dramatic increase in E7-specific CD4(+) and CD8(+) T-cell precursors. They also showed an impressive antitumor effect against E7-expressing tumors compared with mice vaccinated with the wild-type E7 DNA. The delivery of DNA encoding CTGF and E7 or CTGF alone could prolong the survival of transduced dendritic cells (DCs) in vivo. In addition, CTGF/E7-transduced DCs could enhance a higher number of E7-specific CD8(+) T cells than E7-transduced DCs. By prolonging the survival of APCs, DNA vaccine encoding CTGF linked to a tumor antigen represents an innovative approach to enhance DNA vaccine potency and holds promise for cancer prophylaxis and immunotherapy.

Presence of IgT-C and I-A subregion-encoded determinants on distinct chains of monoclonal antigen-specific augmenting factor derived from a T cell hybridoma

PubMed Central

1983-01-01

Monoclonal antibodies specific for mouse T cell alloantigens, Tindd and Tsud, linked to the Igh-1 locus on chromosome 12, were used to directly define the antigen-binding molecule produced by a cloned hybridoma. The T cell hybridoma, FL10, was established from antigen-binding T cells of A/J mice. FL10 produces an antigen-specific augmenting T cell factor (TaF) that bears a unique I region-controlled determinant (I-A) and has antigen-binding capacity. The Tindd, but not the Tsud, determinant was detected on the surface of FL10. The presence of both Tindd and I-A subregion-controlled determinants on FL10-derived TaF was directly demonstrated by the adsorption of TaF with immunoadsorbents prepared with monoclonal antibodies. The Igh-1-linked T cell alloantigen, Tsud, was not found on TaF. Further experiments indicated that Tindd is present on the antigen-binding polypeptide chain and not on the second chain bearing the I-A determinant. Despite the presence of the Tindd determinant on hybridoma-derived TaF, augmentation induced by TaF was restricted by the H-2 type of the responding mice and not by the Igh-1 allotype. PMID:6189953

[Neoretinal antigen expression: a comparison of anatomical and clinical features of a murine uveoretinitis model].

PubMed

Terrada, C; Pâques, M; Fisson, S; De Kozak, Y; Klatzmann, D; Salomon, B; LeHoang, P; Bodaghi, B

2008-02-01

Uveitis is an inflammation involving the retina. The antigens targeted by the experimental models are located in the pigmentary epithelium-photoreceptor complex. To gain insights into the variations in topographic expression of the antigen in the retina, we studied a new mouse model. and methods: Stable retinal expression of the influenza virus hemagglutinin (HA) was obtained after intravitreal or subretinal injection of recombinant adeno-associated virus carrying HA (AAV-HA). One month later, we transferred HA-specific T cells, followed by a subcutaneous immunization of the cognate antigen emulsified in CFA. The animals were clinically examined with a slit lamp biomicroscope. Infiltration of donor cells was detected by immunostaining on retina flatmounts with anti-Thy-1.1 antibody, and infiltrating cells were studied using FACS analysis. Whatever the location of the HA expression, intraocular inflammation was clinically and histologically detected in all animals, between 10 and 15 days after immunization with HA. Lesions were identified with histopathological analysis. The ocular infiltrate was mostly composed of macrophages and HA-specific T cells in different proportions. The topographic variations of targeted ocular antigens do not seem to modify the development of inflammatory reactions in our model. By targeting different antigen-presenting cells, ocular infiltrating cells are different.

T cell receptor–induced phosphoinositide-3-kinase p110δ activity is required for T cell localization to antigenic tissue in mice

PubMed Central

Jarmin, Sarah J.; David, Rachel; Ma, Liang; Chai, Jan-Guo; Dewchand, Hamlata; Takesono, Aya; Ridley, Anne J.; Okkenhaug, Klaus; Marelli-Berg, Federica M.

2008-01-01

The establishment of T cell–mediated inflammation requires the migration of primed T lymphocytes from the blood stream and their retention in antigenic sites. While naive T lymphocyte recirculation in the lymph and blood is constitutively regulated and occurs in the absence of inflammation, the recruitment of primed T cells to nonlymphoid tissue and their retention at the site are enhanced by various inflammatory signals, including TCR engagement by antigen-displaying endothelium and resident antigen-presenting cells. In this study, we investigated whether signals downstream of TCR ligation mediated by the phosphoinositide-3-kinase (PI3K) subunit p110δ contributed to the regulation of these events. T lymphocytes from mice expressing catalytically inactive p110δ displayed normal constitutive trafficking and migratory responses to nonspecific stimuli. However, these cells lost susceptibility to TCR-induced migration and failed to localize efficiently to antigenic tissue. Importantly, we showed that antigen-induced T cell trafficking and subsequent inflammation was abrogated by selective pharmacological inhibition of PI3K p110δ activity. These observations suggest that pharmacological targeting of p110δ activity is a viable strategy for the therapy of T cell–mediated pathology. PMID:18259608

A structural basis for antigen recognition by the T cell-like lymphocytes of sea lamprey

DOE Office of Scientific and Technical Information (OSTI.GOV)

Deng, Lu; Velikovsky, C. Alejandro; Xu, Gang

Adaptive immunity in jawless vertebrates is mediated by leucine-rich repeat proteins called 'variable lymphocyte receptors' (VLRs). Two types of VLR (A and B) are expressed by mutually exclusive lymphocyte populations in lamprey. VLRB lymphocytes resemble the B cells of jawed vertebrates; VLRA lymphocytes are similar to T cells. We determined the structure of a high-affinity VLRA isolated from lamprey immunized with hen egg white lysozyme (HEL) in unbound and antigen-bound forms. The VLRA-HEL complex demonstrates that certain VLRAs, like {gamma}{delta} T-cell receptors (TCRs) but unlike {alpha}{beta} TCRs, can recognize antigens directly, without a requirement for processing or antigen-presenting molecules. Thus,more » these VLRAs feature the nanomolar affinities of antibodies, the direct recognition of unprocessed antigens of both antibodies and {gamma}{delta} TCRs, and the exclusive expression on the lymphocyte surface that is unique to {alpha}{beta} and {gamma}{delta} TCRs.« less

Identifying Regulators of the Immune Response to Dying Cells | Center for Cancer Research

Cancer.gov

Cytotoxic T cells are responsible for carrying out antigen-mediated immune responses against virally-infected and malignant cells. In both cases, cytotoxic T cells are stimulated by interacting with antigen presenting cells, such as dendritic cells (DCs). Infected cells produce virus-specific antigens and pathogen associated molecular patterns, which are recognized by DCs and lead to robust T cell activation. Dead or dying uninfected cells, on the other hand, release damage associated molecular patterns, but their release does not always appear to be sufficient to induce cytotoxic T cell activity. Tim Greten, M.D., of CCR’s Medical Oncology Branch, and a group of international collaborators set out to understand how immune responses against dying cancer cells are regulated. These processes are likely to be important for improving the efficacy of cancer treatment vaccines, which induce an immune reaction against a patient’s cancer cells.

Antigen Specific Responses and ANA production in B6.Sle1b mice: A role for SAP

PubMed Central

Jennings, Paula; Chan, Alice; Schwartzberg, Pamela; Wakeland, Edward K.; Yuan, Dorothy

2010-01-01

B6.Sle1b mice, which contain the Sle1b gene interval derived from lupus prone NZM2410 mice on a C57BL/6 background, present with gender-biased, highly penetrant anti-nuclear antibody (ANA) production. To obtain some insight into the possible induction mechanism of autoantibodies in these mice we compared antigen specific T dependent (TD) and T independent (TI-II) responses between B6.Sle1b and B6 mice before the development of high ANA titers. Our results show that B6.Sle1b mice mount enhanced responses to a TI-II antigen. Additionally, the memory T cell response generated by a TD antigen was also increased. This enhancement correlates with the greater ability of B cells from B6.Sle1b mice to present antigen to T cells. The SLAM Associated Protein (SAP) is critical for signaling of many of the molecules encoded by the SLAM/CD2 gene cluster, candidates for mediating the Sle1b phenotype; therefore, we also investigated the effect of sap deletion in these strains on the TD and TI-II responses as well as on ANA production. The results of these studies of responses to non-self antigens provide further insight for the mechanism by which responses to self-antigens might be initiated in the context of specific genetic alterations. PMID:18845419

Engineering Dendritic Cells to Enhance Cancer Immunotherapy

PubMed Central

Boudreau, Jeanette E; Bonehill, Aude; Thielemans, Kris; Wan, Yonghong

2011-01-01

Cancer immunotherapy aims to establish immune-mediated control of tumor growth by priming T-cell responses to target tumor-associated antigens. Three signals are required for T-cell activation: (i) presentation of cognate antigen in self MHC molecules; (ii) costimulation by membrane-bound receptor-ligand pairs; and (iii) soluble factors to direct polarization of the ensuing immune response. The ability of dendritic cells (DCs) to provide all three signals required for T-cell activation makes them an ideal cancer vaccine platform. Several strategies have been developed to enhance and control antigen presentation, costimulation, and cytokine production. In this review, we discuss progress toward developing DC-based cancer vaccines by genetic modification using RNA, DNA, and recombinant viruses. Furthermore, the ability of DC-based vaccines to activate natural killer (NK) and B-cells, and the impact of gene modification strategies on these populations is described. Clinical trials using gene-modified DCs have shown modest results, therefore, further considerations for DC manipulation to enhance their clinical efficacy are also discussed. PMID:21468005

Toward a Network Model of MHC Class II-Restricted Antigen Processing

PubMed Central

Miller, Michael A.; Ganesan, Asha Purnima V.; Eisenlohr, Laurence C.

2013-01-01

The standard model of Major Histocompatibility Complex class II (MHCII)-restricted antigen processing depicts a straightforward, linear pathway: internalized antigens are converted into peptides that load in a chaperone dependent manner onto nascent MHCII in the late endosome, the complexes subsequently trafficking to the cell surface for recognition by CD4+ T cells (TCD4+). Several variations on this theme, both moderate and radical, have come to light but these alternatives have remained peripheral, the conventional pathway generally presumed to be the primary driver of TCD4+ responses. Here we continue to press for the conceptual repositioning of these alternatives toward the center while proposing that MHCII processing be thought of less in terms of discrete pathways and more in terms of a network whose major and minor conduits are variable depending upon many factors, including the epitope, the nature of the antigen, the source of the antigen, and the identity of the antigen-presenting cell. PMID:24379819

Activation status of mucosal-associated invariant T cells reflects disease activity and pathology of systemic lupus erythematosus.

PubMed

Chiba, Asako; Tamura, Naoto; Yoshikiyo, Kazunori; Murayama, Goh; Kitagaichi, Mie; Yamaji, Ken; Takasaki, Yoshinari; Miyake, Sachiko

2017-03-14

Mucosal-associated invariant T (MAIT) cells are innate-like lymphocytes constituting a large proportion of peripheral blood T cells expressing αβ T-cell receptor in humans. In this study, we aimed to investigate their involvement in systemic lupus erythematosus (SLE). Peripheral blood MAIT cells from patients with SLE were assessed for their frequency, activation markers, and cell death by flow cytometry. The correlation between plasma cytokine levels and CD69 expression on MAIT cells was analyzed. The major histocompatibility complex class I-related protein MR1-restricted antigen-presenting capacity of antigen-presenting cells was investigated. Cytokine-mediated activation of MAIT cells in the absence of exogenous antigens was also examined. The frequency of MAIT cells was markedly reduced in SLE. The reduced number of MAIT cells was not attributable to the downregulation of surface markers, but it was partially due to the enhanced cell death of MAIT cells, possibly by activation-induced cell death. The CD69 expression levels on MAIT cells in SLE correlated with disease activity. Moreover, monocytes from patients with SLE exhibited increased ability to induce MAIT cell activation. The plasma concentration of interleukin (IL)-6, IL-18, and interferon (IFN)-α positively correlated with the expression levels of CD69 on MAIT cells in SLE. MAIT cells were activated by cytokines, including IFN-α, IL-15, and IL-12 plus IL-18, in the absence of exogenous antigens. These results suggest that MAIT cells reflect the pathological condition of SLE and that their activated status correlates with presence of disease.

A meta-analysis of HLA peptidome composition in different hematological entities: entity-specific dividing lines and “pan-leukemia” antigens

PubMed Central

Walz, Simon; Schuster, Heiko; Berlin, Claudia; Neidert, Marian Christoph; Schemionek, Mirle; Brümmendorf, Tim H.; Vucinic, Vladan; Niederwieser, Dietger; Kanz, Lothar; Salih, Helmut Rainer; Kohlbacher, Oliver; Weisel, Katja; Rammensee, Hans-Georg; Stevanović, Stefan; Walz, Juliane Sarah

2017-01-01

Hematological malignancies (HM) are highly amenable targets for immunotherapeutic intervention and may be effectively treated by antigen-specific T-cell based treatment. Recent studies demonstrate that physiologically occurring anti-cancer T-cell responses in certain HM entities target broadly presented non-mutated epitopes. HLA ligands are thus implied as prime targets for broadly applicable and antigen-specific off-the-shelf compounds. With the aim of assessing the presence of common targets shared among different HM which may enable addressing a larger patient collective we conducted a meta-analysis of 83 mass spectrometry-based HLA peptidome datasets (comprising 40,361 unique peptide identifications) across four major HM (19 AML, 16 CML, 35 CLL, and 13 MM/MCL samples) and investigated similarities and differences within the HLA presented antigenic landscape. We found the cancer HLA peptidome datasets to cluster specifically along entity and lineage lines, suggesting that the immunopeptidome directly reflects the differences in the underlying (tumor-)biology. In line with these findings, we only detected a small set of entity-spanning antigens, which were predominantly characterized by low presentation frequencies within the different patient cohorts. These findings suggest that design of T-cell immunotherapies for the treatment of HM should ideally be conducted in an entity-specific fashion. PMID:28159928

Cancer Immunotherapy Utilized Bubble Liposomes and Ultrasound as Antigen Delivery System

NASA Astrophysics Data System (ADS)

Oda, Yusuke; Otake, Shota; Suzuki, Ryo; Otake, Shota; Nishiie, Norihito; Hirata, Keiichi; Taira, Yuichiro; Utoguchi, Naoki; Maruyama, Kazuo

2010-03-01

In dendritic cells (DCs)-based cancer immunotherapy, it is important to present the epitope peptide derived from tumor associated antigens (TAAs) on MHC class I in order to induce tumor specific cytotoxic T lymphocytes (CTLs). However, MHC class I molecules generally present the epitope peptides derived from endogenous antigens for DCs but not exogenous ones such as TAAs. Recently, we developed the novel liposomal bubbles (Bubble liposomes) encapsulating perfluoropropane nanobubbles. In this study, we attempted to establish the novel antigen delivery system to induce MHC class I presentation using the combination of ultrasound and Bubble liposomes. Using ovalbumin (OVA) as model antigen, the combination of Bubble liposomes and ultrasound exposure for the DC could induce MHC class I presentation. In addition, the viability of DCs was more than 80%. These results suggest that Bubble liposomes might be a novel ultrasound enhanced antigen delivery tool in DC-based cancer immunotherapy.

MECHANISM OF THYMUS-INDEPENDENT IMMUNOCYTE TRIGGERING

PubMed Central

Coutinho, Antonio; Gronowicz, Eva; Bullock, Wesley W.; Möller, Göran

1974-01-01

The present experiments were performed in order to analyze the mechanism by which thymus-independent antigens (nonspecific B-cell mitogens) can induce specific immune responses to antigenic determinants present on the same molecule. The hapten NNP was coupled to the B-cell mitogen, lipopolysaccharide (LPS). The conjugate retained full mitogenic activity and bound specifically to NNP-reactive cells. NNP-LPS activated polyclonal as well as specific anti-NNP antibody synthesis, but the optimal concentrations for induction of specific anti-NNP cells were several orders of magnitude lower than the concentrations required for polyclonal activation. These low concentrations failed to activate nonspecific cells, but they induced specific thymus-independent responses of high-avidity NNP-specific cells with the typical kinetics of antigenic responses in vitro. Furthermore, hapten-specific cells were paralyzed by NNP-LPS concentrations that were optimal for induction of polyclonal activation. Specific activation and paralysis could be abolished by free hapten indicating that selective binding of NNP-LPS to hapten-specific cells was responsible for the specificity of the response. However, the triggering signal lacked specificity, since high-avidity specific anti-NNP cells could still be activated by stimulating concentrations of NNP-LPS in the presence of free hapten, even though the Ig receptor combining sites were presumably occupied by NNP. The findings show that B cells with specific Ig receptors for the antigenic determinants on mitogen molecules preferentially bind these molecules and become activated at concentrations still unsufficient to trigger other B cells that lack specific receptors. It is suggested that activation for primary IgM responses in B cells is the result of "one nonspecific signal." This nonspecific signal is provided by the mitogenic properties of some antigens (highly thymus independent or, alternatively, by nonspecific T-cell factors (for highly T cell-dependent antigens), or both, and the surface structures responsible for triggering are not the Ig receptors. The specific Ig receptors only act as passive focusing devices for nonspecific stimuli, entitling the cell to be selectively activated, even though both the signal and the receptors for the triggering are nonspecific. PMID:4128449

Vaccination and the TAP-independent antigen processing pathways.

PubMed

López, Daniel; Lorente, Elena; Barriga, Alejandro; Johnstone, Carolina; Mir, Carmen

2013-09-01

The cytotoxic CD8(+) T lymphocyte-mediated cellular response is important for the elimination of virus-infected cells and requires the prior recognition of short viral peptide antigens previously translocated to the endoplasmic reticulum by the transporter associated with antigen processing (TAP). However, individuals with nonfunctional TAP complexes or infected cells with TAP molecules blocked by specific viral proteins, such as the cowpoxvirus, a component of the first source of early empirical vaccination against smallpox, are still able to present several HLA class I ligands generated by the TAP-independent antigen processing pathways to specific cytotoxic CD8(+) T lymphocytes. Currently, bioterrorism and emerging infectious diseases have renewed interest in poxviruses. Recent works that have identified HLA class I ligands and epitopes in virus-infected TAP-deficient cells have implications for the study of both the effectiveness of early empirical vaccination and the analysis of HLA class I antigen processing in TAP-deficient subjects.

Immune responses in dogs with cutaneous adverse food reactions.

PubMed

Veenhof, E Z; Knol, E F; Willemse, T; Rutten, V P M G

2012-06-01

Adverse food reactions (AFR) in dogs are reactions due to apparently harmless food antigens, with an unknown aetiology, i.e. immunopathogenesis. Despite the entry of food allergens via the intestinal tract, in the majority of dogs with AFR, clinical symptoms are only associated with the skin (CAFR). In the present review, factors are presented of relevance in triggering the differentiation of naive T cells into effector T cell types and the role of these T cell types in allergy. More specifically, the allergic immune responses in intestine and skin are discussed in this article as well as the potential pathways, e.g. homing of antigen presenting cells or allergen-induced T cells to the skin, of induction of cutaneous symptoms.

A two-step non-flowcytometry-based naïve B cell isolation method and its application in Staphylococcal enterotoxin B (SEB) presentation.

PubMed

Chokeshai-u-saha, Kaj; Buranapraditkun, Supranee; Jacquet, Alain; Nguyen, Catherine; Ruxrungtham, Kiat

2012-09-01

To study the role of human naïve B cells in antigen presentation and stimulation to naïve CD4+ T cell, a suitable method to reproducibly isolate sufficient naïve B cells is required. To improve the purity of isolated naive B cells obtained from a conventional one-step magnetic bead method, we added a rosetting step to enrich total B cell isolates from human whole blood samples prior to negative cell sorting by magnetic beads. The acquired naïve B cells were analyzed for phenotypes and for their role in Staphylococcal enterotoxin B (SEB) presentation to naïve CD4+ T cells. The mean (SD) naïve B cell (CD19+/CD27-) purity obtained from this two-step method compared with the one-step method was 97% (1.0) versus 90% (1.2), respectively. This two-step method can be used with a sample of whole blood as small as 10 ml. The isolated naive B cells were phenotypically at a resting state and were able to prime naïve CD4+ T cell activation by Staphylococcal enterotoxin B (SEB) presentation. This two-step non-flow cytometry-based approach improved the purity of isolated naïve B cells compared with conventional one-step magnetic bead method. It also worked well with a small blood volume. In addition, this study showed that the isolated naïve B cells can present a super-antigen "SEB" to activate naïve CD4 cells. These methods may thus be useful for further in vitro characterization of human naïve B cells and their roles as antigen presenting cells in various diseases.

Sarcoidosis Th17 Cells are ESAT-6 Antigen Specific but Demonstrate Reduced IFN-γ Expression

PubMed Central

Richmond, Bradley W.; Ploetze, Kristen; Isom, Joan; Chambers-Harris, Isfahan; Braun, Nicole A.; Taylor, Thyneice; Abraham, Susamma; Mageto, Yolanda; Culver, Dan A.; Oswald-Richter, Kyra A.; Drake, Wonder P.

2013-01-01

Rationale Sarcoidosis is a granulomatous disease of unknown etiology. Many patients with sarcoidosis demonstrate antigen-specific immunity to mycobacterial virulence factors. Th-17 cells are crucial to the immune response in granulomatous inflammation, and have recently been shown to be present in greater numbers in the peripheral blood and bronchoalveolar lavage (BAL) fluid (BALF) of sarcoidosis patients than healthy controls. It is unclear whether Th-17 cells in sarcoidosis are specific for mycobacterial antigens, or whether they have similar functionality to control Th-17 cells. Methods Flow cytometry was used to determine the numbers of Th-17 cells present in the peripheral blood and BALF of patients with sarcoidosis, the percentage of Th-17 cells that were specific to the mycobacterial virulence factor ESAT-6, and as well as to assess IFN-γ expression in Th-17 cells following polyclonal stimulation. Results Patients with sarcoidosis had greater numbers of Th-17 cells in the peripheral blood and BALF than controls and produced significantly more extracellular IL-17A (p=0.03 and p=0.02, respectively). ESAT-6 specific Th-17 cells were present in both peripheral blood and BALF of sarcoidosis patients (p<0.001 and p=0.03, respectively). After polyclonal stimulation, Th-17 cells from sarcoidosis patients produced less IFN-γ than healthy controls. Conclusions Patients with sarcoidosis have mycobacterial antigen-specific Th-17 cells peripherally and in sites of active sarcoidosis involvement. Despite the Th1 immunophenotype of sarcoidosis immunology, the Th-17 cells have reduced IFN-γ expression, compared to healthy controls. This reduction in immunity may contribute to sarcoidosis pathogenesis. PMID:23073617

Preparation of protein loaded poly(D,L-lactide-co-glycolide) microparticles for the antigen delivery to dendritic cells using a static micromixer.

PubMed

Wischke, Christian; Lorenzen, Dirk; Zimmermann, Julian; Borchert, Hans-Hubert

2006-04-01

The cellular immune response against tumors, viruses, or intracellular bacteria requires adequate antigen delivery to professional phagocytes, their processing and the presentation of antigenic peptides to T-cells. Biodegradable microparticles to enhance antigen phagocytosis and the response of cytotoxic lymphocytes have been proposed. The aim of the present study was to formulate poly(lactide-co-glycolide) (PLGA) microparticles using a w/o/w solvent evaporation procedure in order to obtain suitable vehicles for vaccination. Bovine serum albumin bearing fluorescein isothiocyanate (FITC-BSA) was used as a model antigen. For microparticle preparation a static micromixer was employed. Microparticles of 2-3 microm can be produced with good reproducibility by applying high flow rates at the micromixer. Microparticles with a smooth surface and only one pore were observed using scanning electron microscopy (SEM). Confocal laser scanning microscopy (CLSM) allowed localisation of the FITC-BSA near the surface of the microparticle. Microencapsulation of FITC-BSA did not altered the polymer characteristics, as determined by measuring the glass transition temperature. Additionally we could determine residual methylene chloride, employed as solvent in microparticle preparation, to be less than 1/1000 of the USP and Ph. Eur. limit. The microparticles described herein were able to deliver the model antigen to human dendritic cells (DC).

Cis-Acting Pathways Selectively Enforce the Non-Immunogenicity of Shed Placental Antigen for Maternal CD8 T Cells

PubMed Central

Tay, Chin-Siean; Tagliani, Elisa; Collins, Mary K.; Erlebacher, Adrian

2013-01-01

Maternal immune tolerance towards the fetus and placenta is thought to be established in part by pathways that attenuate T cell priming to antigens released from the placenta into maternal blood. These pathways remain largely undefined and their existence, at face value, seems incompatible with a mother's need to maintain a functional immune system during pregnancy. A particular conundrum is evident if we consider that maternal antigen presenting cells, activated in order to prime T cells to pathogen-derived antigens, would also have the capacity to prime T cells to co-ingested placental antigens. Here, we address this paradox using a transgenic system in which placental membranes are tagged with a strong surrogate antigen (ovalbumin). We find that although a remarkably large quantity of acellular ovalbumin-containing placental material is released into maternal blood, splenic CD8 T cells in pregnant mice bearing unmanipulated T cell repertoires are not primed to ovalbumin even if the mice are intravenously injected with adjuvants. This failure was largely independent of regulatory T cells, and instead was linked to the intrinsic characteristics of the released material that rendered it selectively non-immunogenic, potentially by sequestering it from CD8α+ dendritic cells. The release of ovalbumin-containing placental material into maternal blood thus had no discernable impact on CD8 T cell priming to soluble ovalbumin injected intravenously during pregnancy, nor did it induce long-term tolerance to ovalbumin. Together, these results outline a major pathway governing the maternal immune response to the placenta, and suggest how tolerance to placental antigens can be maintained systemically without being detrimental to host defense. PMID:24391885

Lymphocyte apheresis for chimeric antigen receptor T-cell manufacturing in children and young adults with leukemia and neuroblastoma.

PubMed

Ceppi, Francesco; Rivers, Julie; Annesley, Colleen; Pinto, Navin; Park, Julie R; Lindgren, Catherine; Mgebroff, Stephanie; Linn, Naomi; Delaney, Meghan; Gardner, Rebecca A

2018-06-01

The first step in the production of chimeric antigen receptor T cells is the collection of autologous T cells using apheresis technology. The procedure is technically challenging, because patients often have low leukocyte counts and are heavily pretreated with multiple lines of chemotherapy, marrow transplantation, and/or radiotherapy. Here, we report our experience of collecting T lymphocytes for chimeric antigen receptor T-cell manufacturing in pediatric and young adult patients with leukemia, non-Hodgkin lymphoma, or neuroblastoma. Apheresis procedures were performed on a COBE Spectra machine using the mononuclear cell program, with a collection target of 1 × 10 9 total mononuclear cells per kilogram. Data were collected regarding preapheresis and postapheresis blood counts, apheresis parameters, products, and adverse events. Ninety-nine patients (ages 1.3-25.7 years) and 102 apheresis events were available for analysis. Patients underwent apheresis at a variety of absolute lymphocyte cell counts, with a median absolute lymphocyte count of 944 cells/μL (range, 142-6944 cells/μL). Twenty-two patients (21.6%) had absolute lymphocyte counts less than 500 cells/μL. The mononuclear cell target was obtained in 100% of all apheresis harvests, and chimeric antigen receptor T-cell production was possible from the majority of collections (94%). Mononuclear cell collection efficiency was 65.4%, and T-lymphocyte collection efficiency was 83.4%. Ten patients (9.8%) presented with minor adverse events during the 102 apheresis procedures, with one exception of a severe allergy. Mononuclear cell apheresis for chimeric antigen receptor T-cell therapy is well tolerated and safe, and it is possible to obtain an adequate quantity of CD3+ lymphocytes for chimeric antigen receptor T-cell manufacturing in heavily pretreated patients who have low lymphocyte counts. © 2018 AABB.

Direct Delivery of Antigens to Dendritic Cells via Antibodies Specific for Endocytic Receptors as a Promising Strategy for Future Therapies

PubMed Central

Lehmann, Christian H. K.; Heger, Lukas; Heidkamp, Gordon F.; Baranska, Anna; Lühr, Jennifer J.; Hoffmann, Alana; Dudziak, Diana

2016-01-01

Dendritic cells (DCs) are the most potent professional antigen presenting cells and are therefore indispensable for the control of immunity. The technique of antibody mediated antigen targeting to DC subsets has been the basis of intense research for more than a decade. Many murine studies have utilized this approach of antigen delivery to various kinds of endocytic receptors of DCs both in vitro and in vivo. Today, it is widely accepted that different DC subsets are important for the induction of select immune responses. Nevertheless, many questions still remain to be answered, such as the actual influence of the targeted receptor on the initiation of the immune response to the delivered antigen. Further efforts to better understand the induction of antigen-specific immune responses will support the transfer of this knowledge into novel treatment strategies for human diseases. In this review, we will discuss the state-of-the-art aspects of the basic principles of antibody mediated antigen targeting approaches. A table will also provide a broad overview of the latest studies using antigen targeting including addressed DC subset, targeted receptors, outcome, and applied coupling techniques. PMID:27043640

Using X-ray Crystallography, Biophysics, and Functional Assays to Determine the Mechanisms Governing T-cell Receptor Recognition of Cancer Antigens.

PubMed

MacLachlan, Bruce J; Greenshields-Watson, Alexander; Mason, Georgina H; Schauenburg, Andrea J; Bianchi, Valentina; Rizkallah, Pierre J; Sewell, Andrew K; Fuller, Anna; Cole, David K

2017-02-06

Human CD8+ cytotoxic T lymphocytes (CTLs) are known to play an important role in tumor control. In order to carry out this function, the cell surface-expressed T-cell receptor (TCR) must functionally recognize human leukocyte antigen (HLA)-restricted tumor-derived peptides (pHLA). However, we and others have shown that most TCRs bind sub-optimally to tumor antigens. Uncovering the molecular mechanisms that define this poor recognition could aid in the development of new targeted therapies that circumnavigate these shortcomings. Indeed, present therapies that lack this molecular understanding have not been universally effective. Here, we describe methods that we commonly employ in the laboratory to determine how the nature of the interaction between TCRs and pHLA governs T-cell functionality. These methods include the generation of soluble TCRs and pHLA and the use of these reagents for X-ray crystallography, biophysical analysis, and antigen-specific T-cell staining with pHLA multimers. Using these approaches and guided by structural analysis, it is possible to modify the interaction between TCRs and pHLA and to then test how these modifications impact T-cell antigen recognition. These findings have already helped to clarify the mechanism of T-cell recognition of a number of cancer antigens and could direct the development of altered peptides and modified TCRs for new cancer therapies.

Engineering Chimeric Antigen Receptor T cells to Treat Glioblastoma.

PubMed

Choi, Bryan D; O'Rourke, Donald M; Maus, Marcela V

2017-08-01

Immunotherapy has emerged as a promising strategy for glioblastoma (GBM), a disease that remains universally fatal despite currently available standard-of-care. Adoptive T cell therapy has been shown to produce potent antitumor immunity while obviating the need for traditional antigen presentation and primary immune responses. Chimeric antigen receptors (CARs) are specialized molecules that can be expressed on the surface of T cells allowing for redirected cytotoxicity against tumor antigens of interest. To date, the application of CAR T cells for GBM has been relatively limited, in large part due to a dearth of well-described tumor specific antigens that are both homogenously and frequently expressed. A mutated version of the epidermal growth factor receptor, EGFRvIII, is a constitutively activated tyrosine kinase that is expressed on the surface of GBM and other common neoplasms, but completely absent from all normal tissues. We have recently generated CAR T cells directed against EGFRvIII and reported results from a Phase I clinical trial investigating this platform in patients with EGFRvIII-expressing GBM. Our study showed that despite conventional notions of central nervous system "immune-privilege," EGFRvIII CAR T cells trafficked to intracerebral tumors, leading to successful targeting and eradication of this antigen in the brain. Here, we review our experience with EGFRvIII CAR T cells and highlight important considerations for the clinical translation of this therapy in patients with GBM.

Antigen tissue distribution of Avian bornavirus (ABV) in psittacine birds with natural spontaneous proventricular dilatation disease and ABV genotype 1 infection.

PubMed

Wünschmann, Arno; Honkavuori, Kirsi; Briese, Thomas; Lipkin, W Ian; Shivers, Jan; Armien, Aníbal G

2011-07-01

Tissues of 10 psittacines from aviary 1 ("case birds") and 5 psittacines from different aviaries were investigated for the presence of Avian bornavirus (ABV) antigen by immunohistochemistry using a polyclonal serum specific for the viral nucleocapsid (N) protein. Seven of 10 case birds had clinical signs, and necropsy findings consistent with proventricular dilatation disease (PDD) while 3 case birds and the 5 birds from other aviaries did not exhibit signs and lesions of this disease. In birds with clinical signs of PDD, ABV antigen was largely limited to neuroectodermal cells including neurons, astroglia, and ependymal cells of the central nervous system, neurons of the peripheral nervous system, and adrenal cells. ABV antigen was present in the nuclei and cytoplasm of infected cells. In 2 case birds that lacked signs and lesions of PDD, viral antigen had a more widespread distribution and was present in nuclei and cytoplasm of epithelial cells of the alimentary and urogenital tract, retina, heart, skeletal muscle, and skin in addition to the mentioned neuroectodermal cells. ABV RNA was identified by reverse transcription polymerase chain reaction (RT-PCR) in tissues of all 7 case birds available for testing from aviary 1, including 4 birds with PDD lesions and the 3 birds without PDD lesions. Sequencing and phylogenetic analysis indicated the presence of ABV genotype 1 in all cases. Findings further substantiate a role of ABV in PDD of psittacine bird species.

Antigen tissue distribution of Avian bornavirus (ABV) in psittacine birds with natural spontaneous proventricular dilatation disease and ABV genotype 1 infection

PubMed Central

Wünschmann, Arno; Honkavuori, Kirsi; Briese, Thomas; Lipkin, W. Ian; Shivers, Jan; Armien, Aníbal G.

2014-01-01

Tissues of 10 psittacines from aviary 1 (“case birds”) and 5 psittacines from different aviaries were investigated for the presence of Avian bornavirus (ABV) antigen by immunohistochemistry using a polyclonal serum specific for the viral nucleocapsid (N) protein. Seven of 10 case birds had clinical signs, and necropsy findings consistent with proventricular dilatation disease (PDD) while 3 case birds and the 5 birds from other aviaries did not exhibit signs and lesions of this disease. In birds with clinical signs of PDD, ABV antigen was largely limited to neuroectodermal cells including neurons, astroglia, and ependymal cells of the central nervous system, neurons of the peripheral nervous system, and adrenal cells. ABV antigen was present in the nuclei and cytoplasm of infected cells. In 2 case birds that lacked signs and lesions of PDD, viral antigen had a more widespread distribution and was present in nuclei and cytoplasm of epithelial cells of the alimentary and urogenital tract, retina, heart, skeletal muscle, and skin in addition to the mentioned neuroectodermal cells. ABV RNA was identified by reverse transcription polymerase chain reaction (RT-PCR) in tissues of all 7 case birds available for testing from aviary 1, including 4 birds with PDD lesions and the 3 birds without PDD lesions. Sequencing and phylogenetic analysis indicated the presence of ABV genotype 1 in all cases. Findings further substantiate a role of ABV in PDD of psittacine bird species. PMID:21908314

Microenvironmental stresses induce HLA-E/Qa-1 surface expression and thereby reduce CD8(+) T-cell recognition of stressed cells.

PubMed

Sasaki, Takanori; Kanaseki, Takayuki; Shionoya, Yosuke; Tokita, Serina; Miyamoto, Sho; Saka, Eri; Kochin, Vitaly; Takasawa, Akira; Hirohashi, Yoshihiko; Tamura, Yasuaki; Miyazaki, Akihiro; Torigoe, Toshihiko; Hiratsuka, Hiroyoshi; Sato, Noriyuki

2016-04-01

Hypoxia and glucose deprivation are often observed in the microenvironment surrounding solid tumors in vivo. However, how they interfere with MHC class I antigen processing and CD8(+) T-cell responses remains unclear. In this study, we analyzed the production of antigenic peptides presented by classical MHC class I in mice, and showed that it is quantitatively decreased in the cells exposed to either hypoxia or glucose deprivation. In addition, we unexpectedly found increased surface expression of HLA-E in human and Qa-1 in mouse tumor cells exposed to combined oxygen and glucose deprivation. The induced Qa-1 on the stressed tumor model interacted with an inhibitory NKG2/CD94 receptor on activated CD8(+) T cells and attenuated their specific response to the antigen. Our results thus suggest that microenvironmental stresses modulate not only classical but also nonclassical MHC class I presentation, and confer the stressed cells the capability to escape from the CD8(+) T-cell recognition. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Preventing vaccinia virus class-I epitopes presentation by HSV-ICP47 enhances the immunogenicity of a TAP-independent cancer vaccine epitope.

PubMed

Raafat, Nermin; Sadowski-Cron, Charlotte; Mengus, Chantal; Heberer, Michael; Spagnoli, Giulio C; Zajac, Paul

2012-09-01

Herpes simplex virus protein ICP47, encoded by US12 gene, strongly downregulates major histocompatibility complex (MHC) class-I antigen restricted presentation by blocking transporter associated with antigen processing (TAP) protein. To decrease viral vector antigenic immunodominance and MHC class-I driven clearance, we engineered recombinant vaccinia viruses (rVV) expressing ICP47 alone (rVV-US12) or together with endoplasmic reticulum (ER)-targeted Melan-A/MART-1(27-35) model tumor epitope (rVV-MUS12). In this study, we show that antigen presenting cells (APC), infected with rVV-US12, display a decreased ability to present TAP dependent MHC class-I restricted viral antigens to CD8+ T-cells. While HLA class-I cell surface expression is strongly downregulated, other important immune related molecules such as CD80, CD44 and, most importantly, MHC class-II are unaffected. Characterization of rVV-MUS12 infected cells demonstrates that over-expression of a TAP-independent peptide, partially compensates for ICP47 induced surface MHC class-I downregulation (30% vs. 70% respectively). Most importantly, in conditions where clearance of infected APC by virus-specific CTL represents a limiting factor, a significant enhancement of CTL responses to the tumor epitope can be detected in cultures stimulated with rVV-MUS12, as compared to those stimulated by rVV-MART alone. Such reagents could become of high relevance in multiple boost protocols required for cancer immunotherapy, to limit vector-specific responsiveness. Copyright © 2011 UICC.

Choice and Design of Adjuvants for Parenteral and Mucosal Vaccines

PubMed Central

Savelkoul, Huub F. J.; Ferro, Valerie A.; Strioga, Marius M.; Schijns, Virgil E. J. C.

2015-01-01

The existence of pathogens that escape recognition by specific vaccines, the need to improve existing vaccines and the increased availability of therapeutic (non-infectious disease) vaccines necessitate the rational development of novel vaccine concepts based on the induction of protective cell-mediated immune responses. For naive T-cell activation, several signals resulting from innate and adaptive interactions need to be integrated, and adjuvants may interfere with some or all of these signals. Adjuvants, for example, are used to promote the immunogenicity of antigens in vaccines, by inducing a pro-inflammatory environment that enables the recruitment and promotion of the infiltration of phagocytic cells, particularly antigen-presenting cells (APC), to the injection site. Adjuvants can enhance antigen presentation, induce cytokine expression, activate APC and modulate more downstream adaptive immune reactions (vaccine delivery systems, facilitating immune Signal 1). In addition, adjuvants can act as immunopotentiators (facilitating Signals 2 and 3) exhibiting immune stimulatory effects during antigen presentation by inducing the expression of co-stimulatory molecules on APC. Together, these signals determine the strength of activation of specific T-cells, thereby also influencing the quality of the downstream T helper cytokine profiles and the differentiation of antigen-specific T helper populations (Signal 3). New adjuvants should also target specific (innate) immune cells in order to facilitate proper activation of downstream adaptive immune responses and homing (Signal 4). It is desirable that these adjuvants should be able to exert such responses in the context of mucosal administered vaccines. This review focuses on the understanding of the potential working mechanisms of the most well-known classes of adjuvants to be used effectively in vaccines. PMID:26344951

Renaissance in tumor immunotherapy: possible combination with phototherapy (Conference Presentation)

NASA Astrophysics Data System (ADS)

Hamblin, Michael R.

2016-03-01

Photodynamic therapy (PDT) uses the combination of non-toxic dyes and harmless visible light to produce highly toxic reactive oxygen species that destroy tumors. The ideal cancer treatment should target both the primary tumor and the metastases with minimal toxicity. This is best accomplished by educating the body's immune system to recognize the tumor as foreign so that after the primary tumor is destroyed, distant metastases will also be eradicated. PDT may accomplish this feat and stimulate long-term, specific anti-tumor immunity. PDT causes an acute inflammatory response, the rapid induction of large amounts of necrotic and apoptotic tumor cells, induction of damage-associated molecular patterns (DAMPS) including heat-shock proteins, stimulates tumor antigen presentation to naïve T-cells, and generation of cytotoxic T-cells that can destroy distant tumor metastases. By using various syngeneic mouse tumors in immunocompetent mice, we have studied specific PDT regimens related to tumor type as well as mouse genotype and phenotype. We have investigated the role of tumor-associated antigens in PDT-induced immune response by choosing mouse tumors that express: model defined antigen, naturally-occurring cancer testis antigen, and oncogenic virus-derived antigen. We studied the synergistic combination of low-dose cyclophosphamide and PDT that unmasks the PDT-induced immune response by depleting the immunosuppressive T-regulatory cells. PDT combined with immunostimulants (toll-like receptor ligands) can synergistically maximize the generation of anti-tumor immunity by activating dendritic cells and switching immunosuppressive macrophages to a tumor rejection phenotype. Tumors expressing defined tumor-associated antigens with known MHC class I peptides allows anti-tumor immunity to be quantitatively compared.

Encryption of agonistic motifs for TLR4 into artificial antigens augmented the maturation of antigen-presenting cells.

PubMed

Ito, Masaki; Hayashi, Kazumi; Minamisawa, Tamiko; Homma, Sadamu; Koido, Shigeo; Shiba, Kiyotaka

2017-01-01

Adjuvants are indispensable for achieving a sufficient immune response from vaccinations. From a functional viewpoint, adjuvants are classified into two categories: "physical adjuvants" increase the efficacy of antigen presentation by antigen-presenting cells (APC) and "signal adjuvants" induce the maturation of APC. Our previous study has demonstrated that a physical adjuvant can be encrypted into proteinous antigens by creating artificial proteins from combinatorial assemblages of epitope peptides and those peptide sequences having propensities to form certain protein structures (motif programming). However, the artificial antigens still require a signal adjuvant to maturate the APC; for example, co-administration of the Toll-like receptor 4 (TLR4) agonist monophosphoryl lipid A (MPLA) was required to induce an in vivo immunoreaction. In this study, we further modified the previous artificial antigens by appending the peptide motifs, which have been reported to have agonistic activity for TLR4, to create "adjuvant-free" antigens. The created antigens with triple TLR4 agonistic motifs in their C-terminus have activated NF-κB signaling pathways through TLR4. These proteins also induced the production of the inflammatory cytokine TNF-α, and the expression of the co-stimulatory molecule CD40 in APC, supporting the maturation of APC in vitro. Unexpectedly, these signal adjuvant-encrypted proteins have lost their ability to be physical adjuvants because they did not induce cytotoxic T lymphocytes (CTL) in vivo, while the parental proteins induced CTL. These results confirmed that the manifestation of a motif's function is context-dependent and simple addition does not always work for motif-programing. Further optimization of the molecular context of the TLR4 agonistic motifs in antigens should be required to create adjuvant-free antigens.

Encryption of agonistic motifs for TLR4 into artificial antigens augmented the maturation of antigen-presenting cells

PubMed Central

Hayashi, Kazumi; Minamisawa, Tamiko; Homma, Sadamu; Koido, Shigeo; Shiba, Kiyotaka

2017-01-01

Adjuvants are indispensable for achieving a sufficient immune response from vaccinations. From a functional viewpoint, adjuvants are classified into two categories: “physical adjuvants” increase the efficacy of antigen presentation by antigen-presenting cells (APC) and “signal adjuvants” induce the maturation of APC. Our previous study has demonstrated that a physical adjuvant can be encrypted into proteinous antigens by creating artificial proteins from combinatorial assemblages of epitope peptides and those peptide sequences having propensities to form certain protein structures (motif programming). However, the artificial antigens still require a signal adjuvant to maturate the APC; for example, co-administration of the Toll-like receptor 4 (TLR4) agonist monophosphoryl lipid A (MPLA) was required to induce an in vivo immunoreaction. In this study, we further modified the previous artificial antigens by appending the peptide motifs, which have been reported to have agonistic activity for TLR4, to create “adjuvant-free” antigens. The created antigens with triple TLR4 agonistic motifs in their C-terminus have activated NF-κB signaling pathways through TLR4. These proteins also induced the production of the inflammatory cytokine TNF-α, and the expression of the co-stimulatory molecule CD40 in APC, supporting the maturation of APC in vitro. Unexpectedly, these signal adjuvant-encrypted proteins have lost their ability to be physical adjuvants because they did not induce cytotoxic T lymphocytes (CTL) in vivo, while the parental proteins induced CTL. These results confirmed that the manifestation of a motif’s function is context-dependent and simple addition does not always work for motif-programing. Further optimization of the molecular context of the TLR4 agonistic motifs in antigens should be required to create adjuvant-free antigens. PMID:29190754

Immunization with recombinantly expressed glycan antigens from Schistosoma mansoni induces glycan-specific antibodies against the parasite

PubMed Central

Prasanphanich, Nina Salinger; Luyai, Anthony E; Song, Xuezheng; Heimburg-Molinaro, Jamie; Mandalasi, Msano; Mickum, Megan; Smith, David F; Nyame, A Kwame; Cummings, Richard D

2014-01-01

Schistosomiasis caused by infection with parasitic helminths of Schistosoma spp. is a major global health problem due to inadequate treatment and lack of a vaccine. The immune response to schistosomes includes glycan antigens, which could be valuable diagnostic markers and vaccine targets. However, no precedent exists for how to design vaccines targeting eukaryotic glycoconjugates. The di- and tri-saccharide motifs LacdiNAc (GalNAcβ1,4GlcNAc; LDN) and fucosylated LacdiNAc (GalNAcβ1,4(Fucα1-3)GlcNAc; LDNF) are the basis for several important schistosome glycan antigens. They occur in monomeric form or as repeating units (poly-LDNF) and as part of a variety of different glycoconjugates. Because chemical synthesis and conjugation of such antigens is exceedingly difficult, we sought to develop a recombinant expression system for parasite glycans. We hypothesized that presentation of parasite glycans on the cell surface would induce glycan-specific antibodies. We generated Chinese hamster ovary (CHO) Lec8 cell lines expressing poly-LDN (L8-GT) and poly-LDNF (L8-GTFT) abundantly on their membrane glycoproteins. Sera from Schistosoma mansoni-infected mice were highly cross-reactive with the cells and with cell-surface N-glycans. Immunizing mice with L8-GT and L8-GTFT cells induced glycan-specific antibodies. The L8-GTFT cells induced a sustained booster response, with antibodies that bound to S. mansoni lysates and recapitulated the exquisite specificity of the anti-parasite response for particular presentations of LDNF antigen. In summary, this recombinant expression system promotes successful generation of antibodies to the glycans of S. mansoni, and it can be adapted to study the role of glycan antigens and anti-glycan immune responses in many other infections and pathologies. PMID:24727440

Regulation of allergic airway inflammation by adoptive transfer of CD4+ T cells preferentially producing IL-10.

PubMed

Matsuda, Masaya; Doi, Kana; Tsutsumi, Tatsuya; Fujii, Shinya; Kishima, Maki; Nishimura, Kazuma; Kuroda, Ikue; Tanahashi, Yu; Yuasa, Rino; Kinjo, Toshihiko; Kuramoto, Nobuyuki; Mizutani, Nobuaki; Nabe, Takeshi

2017-10-05

Anti-inflammatory pharmacotherapy for asthma has mainly depended on the inhalation of glucocorticoids, which non-specifically suppress immune responses. If the anti-inflammatory cytokine interleukin (IL)-10 can be induced by a specific antigen, asthmatic airway inflammation could be suppressed when individuals are exposed to the antigen. The purpose of this study was to develop cellular immunotherapeutics for atopic diseases using IL-10-producing CD4 + T cells. Spleen cells isolated from ovalbumin (OVA)-sensitized mice were cultured with the antigen, OVA and growth factors, IL-21, IL-27 and TGF-β for 7 days. After the 7-day culture, the CD4 + T cells were purified using a murine CD4 magnetic beads system. When the induced CD4 + T cells were stimulated by OVA in the presence of antigen-presenting cells, IL-10 was preferentially produced in vitro. When CD4 + T cells were adoptively transferred to OVA-sensitized mice followed by intratracheal OVA challenges, IL-10 was preferentially produced in the serum and bronchoalveolar lavage fluid in vivo. IL-10 production coincided with the inhibition of eosinophilic airway inflammation and epithelial mucus plugging. Most of the IL-10-producing CD4 + T cells were negative for Foxp3 and GATA-3, transcription factors of naturally occurring regulatory T cells and Th2 cells, respectively, but double positive for LAG-3 and CD49b, surface markers of inducible regulatory T cells, Tr1 cells. Collectively, most of the induced IL-10-producing CD4 + T cells could be Tr1 cells, which respond to the antigen to produce IL-10, and effectively suppressed allergic airway inflammation. The induced Tr1 cells may be useful for antigen-specific cellular immunotherapy for atopic diseases. Copyright © 2017 Elsevier B.V. All rights reserved.

Identification of Inhibitors of the Association of ZAP-70 with the T Cell Receptor by High-Throughput Screen

DOE PAGES

Visperas, Patrick R.; Wilson, Christopher G.; Winger, Jonathan A.; ...

2016-12-13

ZAP-70 is a critical molecule in the transduction of T cell antigen receptor signaling and the activation of T cells. Upon activation of the T cell antigen receptor, ZAP-70 is recruited to the intracellular ζ-chains of the T cell receptor, where ZAP-70 is activated and colocalized with its substrates. Inhibitors of ZAP-70 could potentially function as treatments for autoimmune diseases or organ transplantation. In this work, we present the design, optimization, and implementation of a screen for inhibitors that would disrupt the interaction between ZAP-70 and the T cell antigen receptor. Finally, the screen is based on a fluorescence polarizationmore » assay for peptide binding to ZAP-70.« less

Systemic RNA delivery to dendritic cells exploits antiviral defence for cancer immunotherapy

NASA Astrophysics Data System (ADS)

Kranz, Lena M.; Diken, Mustafa; Haas, Heinrich; Kreiter, Sebastian; Loquai, Carmen; Reuter, Kerstin C.; Meng, Martin; Fritz, Daniel; Vascotto, Fulvia; Hefesha, Hossam; Grunwitz, Christian; Vormehr, Mathias; Hüsemann, Yves; Selmi, Abderraouf; Kuhn, Andreas N.; Buck, Janina; Derhovanessian, Evelyna; Rae, Richard; Attig, Sebastian; Diekmann, Jan; Jabulowsky, Robert A.; Heesch, Sandra; Hassel, Jessica; Langguth, Peter; Grabbe, Stephan; Huber, Christoph; Türeci, Özlem; Sahin, Ugur

2016-06-01

Lymphoid organs, in which antigen presenting cells (APCs) are in close proximity to T cells, are the ideal microenvironment for efficient priming and amplification of T-cell responses. However, the systemic delivery of vaccine antigens into dendritic cells (DCs) is hampered by various technical challenges. Here we show that DCs can be targeted precisely and effectively in vivo using intravenously administered RNA-lipoplexes (RNA-LPX) based on well-known lipid carriers by optimally adjusting net charge, without the need for functionalization of particles with molecular ligands. The LPX protects RNA from extracellular ribonucleases and mediates its efficient uptake and expression of the encoded antigen by DC populations and macrophages in various lymphoid compartments. RNA-LPX triggers interferon-α (IFNα) release by plasmacytoid DCs and macrophages. Consequently, DC maturation in situ and inflammatory immune mechanisms reminiscent of those in the early systemic phase of viral infection are activated. We show that RNA-LPX encoding viral or mutant neo-antigens or endogenous self-antigens induce strong effector and memory T-cell responses, and mediate potent IFNα-dependent rejection of progressive tumours. A phase I dose-escalation trial testing RNA-LPX that encode shared tumour antigens is ongoing. In the first three melanoma patients treated at a low-dose level, IFNα and strong antigen-specific T-cell responses were induced, supporting the identified mode of action and potency. As any polypeptide-based antigen can be encoded as RNA, RNA-LPX represent a universally applicable vaccine class for systemic DC targeting and synchronized induction of both highly potent adaptive as well as type-I-IFN-mediated innate immune mechanisms for cancer immunotherapy.

Impairments of Antigen-Presenting Cells in Pulmonary Tuberculosis

PubMed Central

Sakhno, Ludmila V.; Shevela, Ekaterina Ya.; Tikhonova, Marina A.; Nikonov, Sergey D.; Ostanin, Alexandr A.; Chernykh, Elena R.

2015-01-01

The phenotype and functional properties of antigen-presenting cells (APC), that is, circulating monocytes and generated in vitro macrophages and dendritic cells, were investigated in the patients with pulmonary tuberculosis (TB) differing in lymphocyte reactivity to M. tuberculosis antigens (PPD-reactive versus PPD-anergic patients). We revealed the distinct impairments in patient APC functions. For example, the monocyte dysfunctions were displayed by low CD86 and HLA-DR expression, 2-fold increase in CD14+CD16+ expression, the high numbers of IL-10-producing cells, and enhanced IL-10 and IL-6 production upon LPS-stimulation. The macrophages which were in vitro generated from peripheral blood monocytes under GM-CSF were characterized by Th1/Th2-balance shifting (downproduction of IFN-γ coupled with upproduction of IL-10) and by reducing of allostimulatory activity in mixed lymphocyte culture. The dendritic cells (generated in vitro from peripheral blood monocytes upon GM-CSF + IFN-α) were characterized by impaired maturation/activation, a lower level of IFN-γ production in conjunction with an enhanced capacity to produce IL-10 and IL-6, and a profound reduction of allostimulatory activity. The APC dysfunctions were found to be most prominent in PPD-anergic patients. The possible role of APC impairments in reducing the antigen-specific T-cell response to M. tuberculosis was discussed. PMID:26339660

Application of an enzyme-labeled antigen method for visualizing plasma cells producing antibodies against Strep A, a carbohydrate antigen of Streptococcus pyogenes, in recurrent tonsillitis.

PubMed

Onouchi, Takanori; Mizutani, Yasuyoshi; Shiogama, Kazuya; Inada, Ken-ichi; Okada, Tatsuyoshi; Naito, Kensei; Tsutsumi, Yutaka

2015-01-01

Streptococcus pyogenes is the main causative pathogen of recurrent tonsillitis. Histologically, lesions of recurrent tonsillitis contain numerous plasma cells. Strep A is an antigenic carbohydrate molecule on the cell wall of S. pyogenes. As expected, plasma cells in subjects with recurrent tonsillitis secrete antibodies against Strep A. The enzyme-labeled antigen method is a novel histochemical technique that visualizes specific antibody-producing cells in tissue sections by employing a biotin-labeled antigen as a probe. The purpose of the present study was to visualize plasma cells producing antibodies reactive with Strep A in recurrent tonsillitis. Firstly, the lymph nodes of rats immunized with boiled S. pyogenes were paraformaldehyde-fixed and specific plasma cells localized in frozen sections with biotinylated Strep A. Secondly, an enzyme-labeled antigen method was used on human tonsil surgically removed from 12 patients with recurrent tonsillitis. S. pyogenes genomes were PCR-detected in all 12 specimens. The emm genotypes belonged to emm12 in nine specimens and emm1 in three. Plasma cells producing anti-Strep A antibodies were demonstrated in prefixed frozen sections of rat lymph nodes, 8/12 human specimens from patients with recurrent tonsillitis but not in two control tonsils. In human tonsils, Strep A-reactive plasma cells were observed within the reticular squamous mucosa and just below the mucosa, and the specific antibodies belonged to either IgA or IgG classes. Our technique is effective in visualizing immunocytes producing specific antibodies against the bacterial carbohydrate antigen, and is thus a novel histochemical tool for analyzing immune reactions in infectious disorders. © 2014 The Authors. Microbiology and Immunology Published by The Societies and Wiley Publishing Asia Pty Ltd.

Molecular characterization of neoplastic and normal "sister" lymphoblastoid B-cell lines from chronic lymphocytic leukemia.

PubMed

Lanemo Myhrinder, Anna; Hellqvist, Eva; Bergh, Ann-Charlotte; Jansson, Mattias; Nilsson, Kenneth; Hultman, Per; Jonasson, Jon; Buhl, Anne Mette; Bredo Pedersen, Lone; Jurlander, Jesper; Klein, Eva; Weit, Nicole; Herling, Marco; Rosenquist, Richard; Rosén, Anders

2013-08-01

Chronic lymphocytic leukemia (CLL) B-cells resemble self-renewing CD5 + B-cells carrying auto/xeno-antigen-reactive B-cell receptors (BCRs) and multiple innate pattern-recognition receptors, such as Toll-like receptors and scavenger receptors. Integration of signals from BCRs with multiple surface membrane receptors determines whether the cells will be proliferating, anergic or apoptotic. To better understand the role of antigen in leukemogenesis, CLL cell lines producing monoclonal antibodies (mAbs) will facilitate structural analysis of antigens and supply DNA for genetic studies. We present here a comprehensive genotypic and phenotypic characterization of available CLL and normal B-cell-derived lymphoblastoid cell lines (LCLs) from the same individuals (n = 17). Authenticity and verification studies of CLL-patient origin were done by IGHV sequencing, fluorescence in situ hybridization (FISH) and DNA/short tandem repeat (STR) fingerprinting. Innate B-cell features, i.e. natural Ab production and CD5 receptors, were present in most CLL cell lines, but in none of the normal LCLs. This panel of immortalized CLL-derived cell lines is a valuable reference representing a renewable source of authentic Abs and DNA.

Maternal cell traffic bounds for immune modulation: tracking maternal H-2 alleles in spleens of baby mice by DNA fingerprinting.

PubMed

Wan, Wenhan; Shimizu, Shoji; Ikawa, Hiromichi; Sugiyama, Kiyosh; Yamaguchi, Nobuo

2002-10-01

We have previously reported that the immunization of pregnant mice with T-dependent antigens successfully induced suppression of the antigen-specific plaque-forming cell (PFC) response to the relevant antigens in the offspring. This suppression was not caused by the administered antigens, the antibodies produced by the pregnant mother, or lactational transfer, but was dependent on the presence of the intact maternal T cells. It was major histocompatibility complex (MHC)-restricted manner tolerance, which continued for at least one-sixth of the murine life. Traditionally, the placenta acts as a natural barrier, not allowing the cells to pass through. However, the results presented strongly suggested that maternal T cells pass through the placenta and subsequently induce tolerance. In this present study, we attempted to substantiate the presence of maternal cells in the fetal circulation through the use of molecular techniques. We found that a highly polymorphic microsatellite sequence within the class II Eb gene of the H-2 complex is useful for the molecular detection of various H-2 alleles. DNA polymorphic analysis was used for tracking maternal H-2 alleles in the spleens of baby mice. The main procedure involved polymerase chain reaction amplification and restriction fragment length polymorphism analysis of the DNA sequence encompassing the H-2-specific microsatellite from the genomic DNA of baby mice. The results indicated that maternal T cells of immunized pregnant mice cross the placenta into the fetus, eventually inducing antigen-specific immunological tolerance in the offspring.

The Basics of Artificial Antigen Presenting Cells in T Cell-Based Cancer Immunotherapies.

PubMed

Neal, Lillian R; Bailey, Stefanie R; Wyatt, Megan M; Bowers, Jacob S; Majchrzak, Kinga; Nelson, Michelle H; Haupt, Carl; Paulos, Chrystal M; Varela, Juan C

2017-01-01

Adoptive T cell transfer (ACT) can mediate objective responses in patients with advanced malignancies. There have been major advances in this field, including the optimization of the ex vivo generation of tumor-reactive lymphocytes to ample numbers for effective ACT therapy via the use of natural and artificial antigen presenting cells (APCs). Herein we review the basic properties of APCs and how they have been manufactured through the years to augment vaccine and T cell-based cancer therapies. We then discuss how these novel APCs impact the function and memory properties of T cells. Finally, we propose new ways to synthesize aAPCs to augment the therapeutic effectiveness of antitumor T cells for ACT therapy.

The Basics of Artificial Antigen Presenting Cells in T Cell-Based Cancer Immunotherapies

PubMed Central

Neal, Lillian R.; Bailey, Stefanie R.; Wyatt, Megan M.; Bowers, Jacob S.; Majchrzak, Kinga; Nelson, Michelle H.; Haupt, Carl; Paulos, Chrystal M.; Varela, Juan C.

2017-01-01

Adoptive T cell transfer (ACT) can mediate objective responses in patients with advanced malignancies. There have been major advances in this field, including the optimization of the ex vivo generation of tumor-reactive lymphocytes to ample numbers for effective ACT therapy via the use of natural and artificial antigen presenting cells (APCs). Herein we review the basic properties of APCs and how they have been manufactured through the years to augment vaccine and T cell-based cancer therapies. We then discuss how these novel APCs impact the function and memory properties of T cells. Finally, we propose new ways to synthesize aAPCs to augment the therapeutic effectiveness of antitumor T cells for ACT therapy. PMID:28825053

Boosting immunity to small tumor-associated carbohydrates with bacteriophage qβ capsids.

PubMed

Yin, Zhaojun; Comellas-Aragones, Marta; Chowdhury, Sudipa; Bentley, Philip; Kaczanowska, Katarzyna; Benmohamed, Lbachir; Gildersleeve, Jeffrey C; Finn, M G; Huang, Xuefei

2013-01-01

The development of an effective immunotherapy is an attractive strategy toward cancer treatment. Tumor associated carbohydrate antigens (TACAs) are overexpressed on a variety of cancer cell surfaces, which present tempting targets for anticancer vaccine development. However, such carbohydrates are often poorly immunogenic. To overcome this challenge, we show here that the display of a very weak TACA, the monomeric Tn antigen, on bacteriophage Qβ virus-like particles elicits powerful humoral responses to the carbohydrate. The effects of adjuvants, antigen display pattern, and vaccine dose on the strength and subclasses of antibody responses were established. The local density of antigen rather than the total amount of antigen administered was found to be crucial for induction of high Tn-specific IgG titers. The ability to display antigens in an organized and high density manner is a key advantage of virus-like particles such as Qβ as vaccine carriers. Glycan microarray analysis showed that the antibodies generated were highly selective toward Tn antigens. Furthermore, Qβ elicited much higher levels of IgG antibodies than other types of virus-like particles, and the IgG antibodies produced reacted strongly with the native Tn antigens on human leukemia cells. Thus, Qβ presents a highly attractive platform for the development of carbohydrate-based anticancer vaccines.

In Silico Prediction Analysis of Idiotope-Driven T–B Cell Collaboration in Multiple Sclerosis

PubMed Central

Høglund, Rune A.; Lossius, Andreas; Johansen, Jorunn N.; Homan, Jane; Benth, Jūratė Šaltytė; Robins, Harlan; Bogen, Bjarne; Bremel, Robert D.; Holmøy, Trygve

2017-01-01

Memory B cells acting as antigen-presenting cells are believed to be important in multiple sclerosis (MS), but the antigen they present remains unknown. We hypothesized that B cells may activate CD4+ T cells in the central nervous system of MS patients by presenting idiotopes from their own immunoglobulin variable regions on human leukocyte antigen (HLA) class II molecules. Here, we use bioinformatics prediction analysis of B cell immunoglobulin variable regions from 11 MS patients and 6 controls with other inflammatory neurological disorders (OINDs), to assess whether the prerequisites for such idiotope-driven T–B cell collaboration are present. Our findings indicate that idiotopes from the complementarity determining region (CDR) 3 of MS patients on average have high predicted affinities for disease associated HLA-DRB1*15:01 molecules and are predicted to be endosomally processed by cathepsin S and L in positions that allows such HLA binding to occur. Additionally, complementarity determining region 3 sequences from cerebrospinal fluid (CSF) B cells from MS patients contain on average more rare T cell-exposed motifs that could potentially escape tolerance and stimulate CD4+ T cells than CSF B cells from OIND patients. Many of these features were associated with preferential use of the IGHV4 gene family by CSF B cells from MS patients. This is the first study to combine high-throughput sequencing of patient immune repertoires with large-scale prediction analysis and provides key indicators for future in vitro and in vivo analyses. PMID:29038659

In Silico Prediction Analysis of Idiotope-Driven T-B Cell Collaboration in Multiple Sclerosis.

PubMed

Høglund, Rune A; Lossius, Andreas; Johansen, Jorunn N; Homan, Jane; Benth, Jūratė Šaltytė; Robins, Harlan; Bogen, Bjarne; Bremel, Robert D; Holmøy, Trygve

2017-01-01

Memory B cells acting as antigen-presenting cells are believed to be important in multiple sclerosis (MS), but the antigen they present remains unknown. We hypothesized that B cells may activate CD4 + T cells in the central nervous system of MS patients by presenting idiotopes from their own immunoglobulin variable regions on human leukocyte antigen (HLA) class II molecules. Here, we use bioinformatics prediction analysis of B cell immunoglobulin variable regions from 11 MS patients and 6 controls with other inflammatory neurological disorders (OINDs), to assess whether the prerequisites for such idiotope-driven T-B cell collaboration are present. Our findings indicate that idiotopes from the complementarity determining region (CDR) 3 of MS patients on average have high predicted affinities for disease associated HLA-DRB1*15:01 molecules and are predicted to be endosomally processed by cathepsin S and L in positions that allows such HLA binding to occur. Additionally, complementarity determining region 3 sequences from cerebrospinal fluid (CSF) B cells from MS patients contain on average more rare T cell-exposed motifs that could potentially escape tolerance and stimulate CD4 + T cells than CSF B cells from OIND patients. Many of these features were associated with preferential use of the IGHV4 gene family by CSF B cells from MS patients. This is the first study to combine high-throughput sequencing of patient immune repertoires with large-scale prediction analysis and provides key indicators for future in vitro and in vivo analyses.

Loss of T Cell Antigen Recognition Arising from Changes in Peptide and Major Histocompatibility Complex Protein Flexibility: Implications for Vaccine Design

DOE Office of Scientific and Technical Information (OSTI.GOV)

Insaidoo, Francis K.; Borbulevych, Oleg Y.; Hossain, Moushumi

Modification of the primary anchor positions of antigenic peptides to improve binding to major histocompatibility complex (MHC) proteins is a commonly used strategy for engineering peptide-based vaccine candidates. However, such peptide modifications do not always improve antigenicity, complicating efforts to design effective vaccines for cancer and infectious disease. Here we investigated the MART-1{sub 27-35} tumor antigen, for which anchor modification (replacement of the position two alanine with leucine) dramatically reduces or ablates antigenicity with a wide range of T cell clones despite significantly improving peptide binding to MHC. We found that anchor modification in the MART-1{sub 27-35} antigen enhances themore » flexibility of both the peptide and the HLA-A*0201 molecule. Although the resulting entropic effects contribute to the improved binding of the peptide to MHC, they also negatively impact T cell receptor binding to the peptide {center_dot} MHC complex. These results help explain how the 'anchor-fixing' strategy fails to improve antigenicity in this case, and more generally, may be relevant for understanding the high specificity characteristic of the T cell repertoire. In addition to impacting vaccine design, modulation of peptide and MHC flexibility through changes to antigenic peptides may present an evolutionary strategy for the escape of pathogens from immune destruction.« less

Expression, Polyubiquitination, and Therapeutic Potential of Recombinant E6E7 from HPV16 Antigens Fused to Ubiquitin.

PubMed

de Oliveira, Liliane M Fernandes; Morale, Mirian G; Chaves, Agtha A M; Demasi, Marilene; Ho, Paulo L

2017-01-01

Ubiquitin-proteasome system plays an essential role in the immune response due to its involvement in the antigen generation and presentation to CD8 + T cells. Hereby, ubiquitin fused to antigens has been explored as an immunotherapeutic strategy that requires the activation of cytotoxic T lymphocytes. Here we propose to apply this ubiquitin fusion approach to a recombinant vaccine against human papillomavirus 16-infected cells. E6E7 multi-epitope antigen was fused genetically at its N- or C-terminal end to ubiquitin and expressed in Escherichia coli as inclusion bodies. The antigens were solubilized using urea and purified by nickel affinity chromatography in denatured condition. Fusion of ubiquitin to E6E7 resulted in marked polyubiquitination in vitro mainly when fused to the E6E7 N-terminal. When tested in a therapeutic scenario, the fusion of ubiquitin to E6E7 reinforced the anti-tumor protection and increased the E6/E7-specific cellular immune responses. Present results encourage the investigation of the adjuvant potential of the ubiquitin fusion to recombinant vaccines requiring CD8 + T cells.

Recognition of Live Phosphatidylserine-Labeled Tumor Cells by Dendritic Cells: A Novel Approach to Immunotherapy of Skin Cancer

PubMed Central

Shurin, Michael R.; Potapovich, Alla I.; Tyurina, Yulia Y.; Tourkova, Irina L.; Shurin, Galina V.; Kagan, Valerian E.

2014-01-01

Dendritic cells (DC) loaded with tumor antigens from apoptotic/necrotic tumor cells are commonly used as vaccines for cancer therapy. However, the use of dead tumor cells may cause both tolerance and immunity, making the effect of vaccination unpredictable. To deliver live tumor “cargoes” into DC, we developed a new approach based on the “labeling” of tumors with a phospholipid “eat-me” signal, phosphatidylserine. Expression of phosphatidylserine on live tumor cells mediated their recognition and endocytosis by DC resulting in the presentation of tumor antigens to antigen-specific T cells. In mice, topical application of phosphatidylserine-containing ointment over melanoma induced tumor-specific CTL, local and systemic antitumor immunity, and inhibited tumor growth. Thus, labeling of tumors with phosphatidylserine is a promising strategy for cancer immunotherapy. PMID:19276376

Phenotypic and Functional Properties of Human Steady State CD14+ and CD1a+ Antigen Presenting Cells and Epidermal Langerhans Cells.

PubMed

Fehres, Cynthia M; Bruijns, Sven C M; Sotthewes, Brigit N; Kalay, Hakan; Schaffer, Lana; Head, Steven R; de Gruijl, Tanja D; Garcia-Vallejo, Juan J; van Kooyk, Yvette

2015-01-01

Cutaneous antigen presenting cells (APCs) are critical for the induction and regulation of skin immune responses. The human skin contains phenotypically and functionally distinct APCs subsets that are present at two separated locations. While CD1ahigh LCs form a dense network in the epidermis, the CD14+ and CD1a+ APCs reside in the dermal compartment. A better understanding of the biology of human skin APC subsets is necessary for the improvement of vaccine strategies that use the skin as administration route. In particular, progress in the characterization of uptake and activatory receptors will certainly improve APC-targeting strategies in vaccination. Here we performed a detailed analysis of the expression and function of glycan-binding and pattern-recognition receptors in skin APC subsets. The results demonstrate that under steady state conditions human CD1a+ dermal dendritic cells (DCs) were phenotypically most mature as measured by the expression of CD83 and CD86, whereas the CD14+ cells showed a higher expression of the CLRs DC-SIGN, mannose receptor and DCIR and had potent antigen uptake capacity. Furthermore, steady state LCs showed superior antigen cross-presentation as compared to the dermal APC subsets. Our results also demonstrate that the TLR3 ligand polyribosinic-polyribocytidylic acid (pI:C) was the most potent stimulator of cytokine production by both LCs and dDCs. These studies warrant further exploration of human CD1a+ dDCs and LCs as target cells for cancer vaccination to induce anti-tumor immune responses.

How Do CD4+ T Cells Detect and Eliminate Tumor Cells That Either Lack or Express MHC Class II Molecules?

PubMed Central

Haabeth, Ole Audun Werner; Tveita, Anders Aune; Fauskanger, Marte; Schjesvold, Fredrik; Lorvik, Kristina Berg; Hofgaard, Peter O.; Omholt, Hilde; Munthe, Ludvig A.; Dembic, Zlatko; Corthay, Alexandre; Bogen, Bjarne

2014-01-01

CD4+ T cells contribute to tumor eradication, even in the absence of CD8+ T cells. Cytotoxic CD4+ T cells can directly kill MHC class II positive tumor cells. More surprisingly, CD4+ T cells can indirectly eliminate tumor cells that lack MHC class II expression. Here, we review the mechanisms of direct and indirect CD4+ T cell-mediated elimination of tumor cells. An emphasis is put on T cell receptor (TCR) transgenic models, where anti-tumor responses of naïve CD4+ T cells of defined specificity can be tracked. Some generalizations can tentatively be made. For both MHCIIPOS and MHCIINEG tumors, presentation of tumor-specific antigen by host antigen-presenting cells (APCs) appears to be required for CD4+ T cell priming. This has been extensively studied in a myeloma model (MOPC315), where host APCs in tumor-draining lymph nodes are primed with secreted tumor antigen. Upon antigen recognition, naïve CD4+ T cells differentiate into Th1 cells and migrate to the tumor. At the tumor site, the mechanisms for elimination of MHCIIPOS and MHCIINEG tumor cells differ. In a TCR-transgenic B16 melanoma model, MHCIIPOS melanoma cells are directly killed by cytotoxic CD4+ T cells in a perforin/granzyme B-dependent manner. By contrast, MHCIINEG myeloma cells are killed by IFN-γ stimulated M1-like macrophages. In summary, while the priming phase of CD4+ T cells appears similar for MHCIIPOS and MHCIINEG tumors, the killing mechanisms are different. Unresolved issues and directions for future research are addressed. PMID:24782871

[Regulation of immune responses by exosomes derived from antigen presenting cells].

PubMed

Maravillas-Montero, José Luis; Martínez-Cortés, Ismael

2017-01-01

Cells release several biomolecules to the extracellular environment using them as a communication alternative with neighbor cells. Besides these molecules, cells also release more complex elements, like vesicles; structures composed of a lipidic bilayer with transmembrane proteins that protect a hydrophilic content. Exosomes are a small subtype of vesicles (30-150 nm), produced by many cell types, such as tumor cells, neurons, epithelial cells and immune cells. Included in this last group, antigen presenting cells produce exosomes that contain different types of molecules depending on their activation and/or maturation state. In recent years there has been an exponential interest in exosomes due to the recent evidences that show the immunomodulatory properties of these vesicles and therefore, their great potential in diagnostic approaches and development of therapies for different inflammation-associated pathologies.

pDC therapy induces recovery from EAE by recruiting endogenous pDC to sites of CNS inflammation

PubMed Central

Duraes, Fernanda V.; Lippens, Carla; Steinbach, Karin; Dubrot, Juan; Brighouse, Dale; Bendriss-Vermare, Nathalie; Issazadeh-Navikas, Shohreh; Merkler, Doron; Hugues, Stephanie

2016-01-01

Plasmacytoid dendritic cells (pDCs) exhibit both innate and adaptive functions. In particular they are the main source of type I IFNs and directly impact T cell responses through antigen presentation. We have previously demonstrated that during experimental autoimmune encephalomyelitis (EAE) initiation, myelin-antigen presentation by pDCs is associated with suppressive Treg development and results in attenuated EAE. Here, we show that pDCs transferred during acute disease phase confer recovery from EAE. Clinical improvement is associated with migration of injected pDCs into inflamed CNS and is dependent on the subsequent and selective chemerin-mediated recruitment of endogenous pDCs to the CNS. The protective effect requires pDC pre-loading with myelin antigen, and is associated with the modulation of CNS-infiltrating pDC phenotype and inhibition of CNS encephalitogenic T cells. This study may pave the way for novel pDC-based cell therapies in autoimmune diseases, aiming at specifically modulating pathogenic cells that induce and sustain autoimmune inflammation. PMID:26341385

Manufacture of Clinical-Grade CD19-Specific T Cells Stably Expressing Chimeric Antigen Receptor Using Sleeping Beauty System and Artificial Antigen Presenting Cells

PubMed Central

Singh, Harjeet; Figliola, Matthew J.; Dawson, Margaret J.; Olivares, Simon; Zhang, Ling; Yang, Ge; Maiti, Sourindra; Manuri, Pallavi; Senyukov, Vladimir; Jena, Bipulendu; Kebriaei, Partow; Champlin, Richard E.; Huls, Helen; Cooper, Laurence J. N.

2013-01-01

Adoptive transfer of T cells expressing a CD19-specific chimeric antigen receptor (CAR) is being evaluated in multiple clinical trials. Our current approach to adoptive immunotherapy is based on a second generation CAR (designated CD19RCD28) that signals through a CD28 and CD3-ζ endodomain. T cells are electroporated with DNA plasmids from the Sleeping Beauty (SB) transposon/transposase system to express this CAR. Stable integrants of genetically modified T cells can then be retrieved when co-cultured with designer artificial antigen presenting cells (aAPC) in the presence of interleukin (IL)-2 and 21. Here, we reveal how the platform technologies of SB-mediated transposition and CAR-dependent propagation on aAPC were adapted for human application. Indeed, we have initiated clinical trials in patients with high-risk B-lineage malignancies undergoing autologous and allogeneic hematopoietic stem-cell transplantation (HSCT). We describe the process to manufacture clinical grade CD19-specific T cells derived from healthy donors. Three validation runs were completed in compliance with current good manufacturing practice for Phase I/II trials demonstrating that by 28 days of co-culture on γ-irradiated aAPC ∼1010 T cells were produced of which >95% expressed CAR. These genetically modified and propagated T cells met all quality control testing and release criteria in support of infusion. PMID:23741305

TIL 2.0: More effective and predictive T-cell products by enrichment for defined antigen specificities.

PubMed

Schober, Kilian; Busch, Dirk H

2016-06-01

Adoptive transfer of in vitro-expanded T cells derived from tumor-infiltrating lymphocytes (TILs) in melanoma patients started the era of tumor immunotherapy three decades ago. The approach has demonstrated remarkable clinical responses in several studies since. Reinfusion of TIL-derived T cells represents a highly personalized form of immunotherapy, taking into account the enormous interindividual tumor heterogeneity. However, despite its successes, TIL therapy does not lead to objective clinical responses in all cases. It is thus crucial to find out which tumor antigens are particularly valuable targets and to develop strategies to enhance the reactivity of T-cell products toward them. In this issue of the European Journal of Immunology, Kelderman et al. [Eur. J. Immunol. 2016. 46: 1351-1360] present a platform for the generation of antigen-specific TIL therapy. Combining recently developed technologies for clinical identification and enrichment of antigen-specific CD8(+) T cells, such as MHC Streptamers and UV-mediated peptide exchange, the authors could enrich T-cell populations with defined antigen specificities from melanoma-derived TILs. This T-cell product showed higher reactivity against autologous tumor cell lines than bulk TIL-derived T cells. The novel platform might enable the generation of more effective and predictable TIL-derived T-cell products for future clinical applications. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Understanding original antigenic sin in influenza with a dynamical system.

PubMed

Pan, Keyao

2011-01-01

Original antigenic sin is the phenomenon in which prior exposure to an antigen leads to a subsequent suboptimal immune response to a related antigen. Immune memory normally allows for an improved and rapid response to antigens previously seen and is the mechanism by which vaccination works. I here develop a dynamical system model of the mechanism of original antigenic sin in influenza, clarifying and explaining the detailed spin-glass treatment of original antigenic sin. The dynamical system describes the viral load, the quantities of healthy and infected epithelial cells, the concentrations of naïve and memory antibodies, and the affinities of naïve and memory antibodies. I give explicit correspondences between the microscopic variables of the spin-glass model and those of the present dynamical system model. The dynamical system model reproduces the phenomenon of original antigenic sin and describes how a competition between different types of B cells compromises the overall effect of immune response. I illustrate the competition between the naïve and the memory antibodies as a function of the antigenic distance between the initial and subsequent antigens. The suboptimal immune response caused by original antigenic sin is observed when the host is exposed to an antigen which has intermediate antigenic distance to a second antigen previously recognized by the host's immune system.

Oxidative bioactivation of abacavir in subcellular fractions of human antigen presenting cells.

PubMed

Bell, Catherine C; Santoyo Castelazo, Anahi; Yang, Emma L; Maggs, James L; Jenkins, Rosalind E; Tugwood, Jonathan; O'Neill, Paul M; Naisbitt, Dean J; Park, B Kevin

2013-07-15

Human exposure to abacavir, a primary alcohol antiretroviral, is associated with the development of immunological drug reactions in individuals carrying the HLA risk allele B*57:01. Interaction of abacavir with antigen presenting cells results in cell activation through an Hsp70-mediated Toll-like receptor pathway and the provision of T-cell antigenic determinants. Abacavir's electrophilic aldehyde metabolites are potential precursors of neoantigens. Herein, we have used mass spectrometry to study the oxidative metabolism of abacavir in EBV-transformed human B-cells. RNA and protein were isolated from the cells and subjected to transcriptomic and mass spectrometric analyses to identify the redox enzymes expressed. Low levels of isomeric abacavir carboxylic acids were detected in subcellular fractions of EBV-transformed human B-cells incubated with abacavir. Metabolite formation was time-dependent but was not reduced by an inhibitor of Class I alcohol dehydrogenases. Relatively high levels of mRNA were detected for several redox enzymes, including alcohol dehydrogenase 5 (Class III), aldehyde dehydrogenases (ALDH3A2, ALDH6A1, and ALDH9A1), CYP1B1, CYP2R1, CYP7B1, and hydroxysteroid dehydrogenase 10. Over 2600 proteins were identified by mass spectrometry. More than 1000 of these proteins exhibited catalytic activity, and 80 were oxido-reductases. This is the first proteomic inventory of enzymes in antigen presenting cells. However, neither of the hepatic alcohol dehydrogenases of Class I which metabolize abacavir in vitro was expressed at the protein level. Nevertheless the metabolic production of abacavir carboxylic acids by B-cell fractions implies abacavir-treated immune cells might be exposed to the drug's protein-reactive aldehyde metabolites in vivo.

CD3+ CD8+ NKG2D+ T Lymphocytes Induce Apoptosis and Necroptosis in HLA-Negative Cells via FasL-Fas Interaction.

PubMed

Ivanova, Olga K; Sharapova, Tatiana N; Romanova, Elena A; Soshnikova, Natalia V; Sashchenko, Lidia P; Yashin, Denis V

2017-10-01

An important problem in cellular immunology is to identify new populations of cytotoxic lymphocytes capable of killing tumor cells that have lost classical components of MHC-machinery and to understand mechanisms of the death of these cells. We have previously found that CD4 + CD25 + lymphocytes appear in the lymphokine-activated killer (LAK) cell culture, which carry Tag7 (PGRP-S) and FasL proteins on their surface and can kill Hsp70- and Fas-expressing HLA-negative cells. In this work, we have continued to study the mechanisms of killing of the HLA-negative tumor cells, focusing this time on the CD8 + lymphocytes. We show that after a tumor antigen contact the IL-2 activated CD8 + lymphocytes acquire ability to lyse tumor cells bearing this antigen. However, activation of the CD8 + lymphocytes in the absence of antigen causes appearance of a cytotoxic population of CD8 + NKG2D + lymphocytes, which are able to lyse HLA-negative cancer cells that have lost the classic mechanism of antigen presentation. These cells recognize the noncanonical MicA antigen on the surface of HLA-negative K562 cells but kill them via the FasL-Fas interaction, as do cytotoxic T lymphocytes. FasL presented on the lymphocyte surface can trigger both apoptosis and necroptosis. Unlike in the case of TNFR1, another cell death receptor, no switching to alternative processes has been observed upon induction of Fas-dependent cell death. It may well be that the apoptotic and necroptotic signals are transduced separately in the latter case, with the ability of FasL + lymphocytes to induce necroptosis allowing them to kill tumor cells that escape apoptosis. J. Cell. Biochem. 118: 3359-3366, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Chimeric Antigen Receptor T-Cell Therapy for the Community Oncologist

PubMed Central

Levine, Bruce L.

2016-01-01

The field of cancer immunotherapy has rapidly progressed in the past decade as several therapeutic modalities have entered into the clinic. One such immunotherapy that has shown promise in the treatment of cancer is the use of chimeric antigen receptor (CAR)-modified T lymphocytes. CARs are engineered receptors constructed from antigen recognition regions of antibodies fused to T-cell signaling and costimulatory domains that can be used to reprogram a patient’s T cells to specifically target tumor cells. CAR T-cell therapy has demonstrated sustained complete responses for some patients with advanced leukemia, and a number of CAR therapies are being evaluated in clinical studies. CAR T-cell therapy-associated toxicities, including cytokine release syndrome, macrophage activation syndrome, and tumor lysis syndrome, have been observed and effectively managed in the clinic. In patients with significant clinical responses, sustained B-cell aplasia has also been observed and is a marker of CAR T-cell persistence that might provide long-term disease control. Education on CAR T-cell therapy efficacy and safety management is critical for clinicians and patients who are considering this novel type of treatment. In the present report, the current landscape of CAR T-cell therapy, the effective management of patients undergoing treatment, and which patients are the most suitable candidates for current trials are discussed. Implications for Practice: The present report describes the current status of chimeric antigen receptor (CAR) T lymphocytes as an immunotherapy for patients with relapsed or refractory B-cell malignancies. CAR T cells targeting CD19, a protein expressed on many B-cell malignancies, typically induce high complete response rates in patients with B-cell leukemia or lymphoma who have very limited therapeutic options. Recent clinical trial results of CD19 CAR T-cell therapies and the management of CAR T-cell-associated adverse events are discussed. The present report will therefore inform physicians regarding the efficacy and safety of CAR T cells as a therapy for B-cell malignancies. PMID:27009942

A cell-ELISA for the quantification of adherent murine macrophages and the surface expression of antigens.

PubMed

Nibbering, P H; Van de Gevel, J S; Van Furth, R

1990-07-20

The present study was performed in order to establish whether a cell-ELISA could be used to determine the expression of antigens by adherent murine peritoneal macrophages and also quantify the numbers of such macrophages. Accurate determination of the number of adherent macrophages proved to be possible with a cell-ELISA designed to assess complement receptor type III (CRIII) expression. Expression of CRIII was considerably more sensitive than determination of the cell-protein or DNA content as a measure of the number of adherent macrophages. For the calculation of the expression of CRIII, Ia antigen, and antigen F4/80 by resident and activated macrophages, use was made of the linear part of the curve obtained when the numbers of macrophages were plotted against the absorbance values for each of the antigens. The values for CRIII expression did not differ significantly between resident macrophages, macrophages activated with recombinant interferon-gamma (rIFN-gamma) and macrophages activated with BCG/PPD. IFN-gamma-activated and BCG/PPD-activated macrophages expressed Ia antigen significantly more intensely than did resident peritoneal macrophages. In contrast the activated macrophages expressed F4/80 significantly less intensely than resident peritoneal macrophages.

Chemoselective ligation and antigen vectorization.

PubMed

Gras-Masse, H

2001-01-01

The interest in cocktail-lipopeptide vaccines has now been confirmed by phase I clinical trials: highly diversified B-, T-helper or cytotoxic T-cell epitopes can be combined with a lipophilic vector for the induction of B- and T-cell responses of predetermined specificity. With the goal of producing an improved vaccine that should ideally induce a multispecific response in non-selected populations, increasing the diversity of the immunizing mixture represents one of the most obvious strategies.The selective delivery of antigens to professional antigen-presenting cells represents another promising approach for the improvement of vaccine efficacy. In this context, the mannose-receptor represents an attractive entry point for the targeting to dendritic cells of antigens linked to clustered glycosides or glycomimetics. In all cases, highly complex but fully characterized molecules must be produced. To develop a modular and flexible strategy which could be generally applicable to a large set of peptide antigens, we elected to explore the potentialities of chemoselective ligation methods. The hydrazone bond was found particularly reliable and fully compatible with sulphide ligation. Hydrazone/thioether orthogonal ligation systems could be developed to account for the nature of the antigens and the solubility of the vector systems. Copyright 2001 The International Association for Biologicals.

CD1d expression by hepatocytes is a main restriction element for intrahepatic T-cell recognition.

PubMed

Agrati, C; Martini, F; Nisii, C; Oliva, A; D'Offizi, G; Narciso, P; Nardacci, R; Piacentini, M; Dieli, F; Pucillo, L P; Poccia, F

2005-01-01

The liver has specific mechanisms to protect itself from infectious agents and to avoid autoimmunity, indicating an important role of the hepatic tissues in antigen presentation and tolerance induction. Since intrahepatic lymphocytes may contribute to the innate immunity and to the liver pathology, it is of interest to analyze the expression of antigen presenting molecules and of the related T cell recognition in liver, and how these change in relation to different diseases. We analyzed the expression of MHC class I, and of CD1-a, -b, -c, and -d proteins on liver tissues from patients with different hepatic diseases. Moreover, in the same patients we studied the intrahepatic and peripheral NKT cell recognition of alpha-galactosyl ceramide antigen in the context of CD1d. Unlike in other tissues, classical MHC class I molecules were poorly expressed in the hepatic compartment, suggesting that inflamed hepatocytes may trigger weak MHC-restricted T cell responses. Nevertheless, we observed a prevalent expression of HLA class I-like CD1d isoform on the hepatocyte surface, indicating that CD1d is the main restriction element in the liver. In patients with viral hepatitis, the intrahepatic CD1d expression parallels the recruitment of CD56+Valpha24Vbeta11+ NKT cells in the liver which recognize CD1d presenting glycolipids such as alpha-galactosyl ceramide, suggesting that the intrahepatic T cell immunity may focus on glycolipid antigens.

Turned on by danger: activation of CD1d-restricted invariant natural killer T cells

PubMed Central

Lawson, Victoria

2012-01-01

CD1d-restricted invariant natural killer T (iNKT) cells bear characteristics of innate and adaptive lymphocytes, which allow them to bridge the two halves of the immune response and play roles in many disease settings. Recent work has characterized precisely how their activation is initiated and regulated. Novel antigens from important pathogens have been identified, as has an abundant self-antigen, β-glucopyranosylcaramide, capable of mediating an iNKT-cell response. Studies of the iNKT T-cell receptor (TCR)–antigen–CD1d complex show how docking between CD1d–antigen and iNKT TCR is highly conserved, and how small sequence differences in the TCR establish intrinsic variation in iNKT TCR affinity. The sequence of the TCR CDR3β loop determines iNKT TCR affinity for ligand–CD1d, independent of ligand identity. CD1d ligands can promote T helper type 1 (Th1) or Th2 biased cytokine responses, depending on the composition of their lipid tails. Ligands loaded into CD1d on the cell surface promote Th2 responses, whereas ligands with long hydrophobic tails are loaded endosomally and promote Th1 responses. This information is informing the design of synthetic iNKT-cell antigens. The iNKT cells may be activated by exogenous antigen, or by a combination of dendritic cell-derived interleukin-12 and iNKT TCR–self-antigen–CD1d engagement. The iNKT-cell activation is further modulated by recent foreign or self-antigen encounter. Activation of dendritic cells through pattern recognition receptors alters their antigen presentation and cytokine production, strongly influencing iNKT-cell activation. In a range of bacterial infections, dendritic cell-dependent innate activation of iNKT cells through interleukin-12 is the dominant influence on their activity. PMID:22734667

Carbon nanotubes' surface chemistry determines their potency as vaccine nanocarriers in vitro and in vivo

PubMed Central

Hassan, Hatem A.F.M.; Smyth, Lesley; Rubio, Noelia; Ratnasothy, Kulachelvy; Wang, Julie T.-W.; Bansal, Sukhvinder S.; Summers, Huw D.; Diebold, Sandra S.; Lombardi, Giovanna; Al-Jamal, Khuloud T.

2016-01-01

Carbon nanotubes (CNTs) have shown marked capabilities in enhancing antigen delivery to antigen presenting cells. However, proper understanding of how altering the physical properties of CNTs may influence antigen uptake by antigen presenting cells, such as dendritic cells (DCs), has not been established yet. We hypothesized that altering the physical properties of multi-walled CNTs (MWNTs)-antigen conjugates, e.g. length and surface charge, can affect the internalization of MWNT-antigen by DCs, hence the induced immune response potency. For this purpose, pristine MWNTs (p-MWNTs) were exposed to various chemical reactions to modify their physical properties then conjugated to ovalbumin (OVA), a model antigen. The yielded MWNTs-OVA conjugates were long MWNT-OVA (~ 386 nm), bearing net positive charge (5.8 mV), or short MWNTs-OVA (~ 122 nm) of increasing negative charges (− 23.4, − 35.8 or − 39 mV). Compared to the short MWNTs-OVA bearing high negative charges, short MWNT-OVA with the lowest negative charge demonstrated better cellular uptake and OVA-specific immune response both in vitro and in vivo. However, long positively-charged MWNT-OVA showed limited cellular uptake and OVA specific immune response in contrast to short MWNT-OVA displaying the least negative charge. We suggest that reduction in charge negativity of MWNT-antigen conjugate enhances cellular uptake and thus the elicited immune response intensity. Nevertheless, length of MWNT-antigen conjugate might also affect the cellular uptake and immune response potency; highlighting the importance of physical properties as a consideration in designing a MWNT-based vaccine delivery system. PMID:26802552

Preservation of Antigen-Specific Functions of αβ T Cells and B Cells Removed from Hematopoietic Stem Cell Transplants Suggests Their Use As an Alternative Cell Source for Advanced Manipulation and Adoptive Immunotherapy.

PubMed

Li Pira, Giuseppina; Di Cecca, Stefano; Biagini, Simone; Girolami, Elia; Cicchetti, Elisabetta; Bertaina, Valentina; Quintarelli, Concetta; Caruana, Ignazio; Lucarelli, Barbarella; Merli, Pietro; Pagliara, Daria; Brescia, Letizia Pomponia; Bertaina, Alice; Montanari, Mauro; Locatelli, Franco

2017-01-01

Hematopoietic stem cell transplantation is standard therapy for numerous hematological diseases. The use of haploidentical donors, sharing half of the HLA alleles with the recipient, has facilitated the use of this procedure as patients can rely on availability of a haploidentical donor within their family. Since HLA disparity increases the risk of graft-versus-host disease, T-cell depletion has been used to remove alloreactive lymphocytes from the graft. Selective removal of αβ T cells, which encompass the alloreactive repertoire, combined with removal of B cells to prevent EBV-related lymphoproliferative disease, proved safe and effective in clinical studies. Depleted αβ T cells and B cells are generally discarded as by-products. Considering the possible use of donor T cells for donor lymphocyte infusions or for generation of pathogen-specific T cells as mediators of graft-versus-infection effect, we tested whether cells in the discarded fractions were functionally intact. Response to alloantigens and to viral antigens comparable to that of unmanipulated cells indicated a functional integrity of αβ T cells, in spite of the manipulation used for their depletion. Furthermore, B cells proved to be efficient antigen-presenting cells, indicating that antigen uptake, processing, and presentation were fully preserved. Therefore, we propose that separated αβ T lymphocytes could be employed for obtaining pathogen-specific T cells, applying available methods for positive selection, which eventually leads to indirect allodepletion. In addition, these functional T cells could undergo additional manipulation, such as direct allodepletion or genetic modification.

Preservation of Antigen-Specific Functions of αβ T Cells and B Cells Removed from Hematopoietic Stem Cell Transplants Suggests Their Use As an Alternative Cell Source for Advanced Manipulation and Adoptive Immunotherapy

PubMed Central

Li Pira, Giuseppina; Di Cecca, Stefano; Biagini, Simone; Girolami, Elia; Cicchetti, Elisabetta; Bertaina, Valentina; Quintarelli, Concetta; Caruana, Ignazio; Lucarelli, Barbarella; Merli, Pietro; Pagliara, Daria; Brescia, Letizia Pomponia; Bertaina, Alice; Montanari, Mauro; Locatelli, Franco

2017-01-01

Hematopoietic stem cell transplantation is standard therapy for numerous hematological diseases. The use of haploidentical donors, sharing half of the HLA alleles with the recipient, has facilitated the use of this procedure as patients can rely on availability of a haploidentical donor within their family. Since HLA disparity increases the risk of graft-versus-host disease, T-cell depletion has been used to remove alloreactive lymphocytes from the graft. Selective removal of αβ T cells, which encompass the alloreactive repertoire, combined with removal of B cells to prevent EBV-related lymphoproliferative disease, proved safe and effective in clinical studies. Depleted αβ T cells and B cells are generally discarded as by-products. Considering the possible use of donor T cells for donor lymphocyte infusions or for generation of pathogen-specific T cells as mediators of graft-versus-infection effect, we tested whether cells in the discarded fractions were functionally intact. Response to alloantigens and to viral antigens comparable to that of unmanipulated cells indicated a functional integrity of αβ T cells, in spite of the manipulation used for their depletion. Furthermore, B cells proved to be efficient antigen-presenting cells, indicating that antigen uptake, processing, and presentation were fully preserved. Therefore, we propose that separated αβ T lymphocytes could be employed for obtaining pathogen-specific T cells, applying available methods for positive selection, which eventually leads to indirect allodepletion. In addition, these functional T cells could undergo additional manipulation, such as direct allodepletion or genetic modification. PMID:28386262

Identification of a dendritic cell receptor that couples sensing of necrosis to immunity

PubMed Central

Sancho, David; Joffre, Olivier P.; Keller, Anna M.; Rogers, Neil C.; Martinez, Dolores; Hernanz-Falcón, Patricia; Rosewell, Ian; Reis e Sousa, Caetano

2009-01-01

Injury or impaired clearance of apoptotic cells leads to the pathological accumulation of necrotic corpses, which induce an inflammatory response that initiates tissue repair1. In addition, antigens present within necrotic cells can sometimes provoke a specific immune response2-4 and it has been argued that necrosis could explain adaptive immunity in seemingly infection-free situations, such as after allograft transplantation or in spontaneous and therapy-induced tumour rejection5, 6. In the mouse, the CD8α+ subset of dendritic cells (DC) phagocytoses dead cell remnants and crossprimes CD8+ T cells against cell-associated antigens7. Here, we show that CD8α+ DC utilise CLEC9A (DNGR-1), a recently-characterised C-type lectin8-10, to recognise a preformed signal that is exposed on necrotic cells. Loss or blockade of CLEC9A does not impair uptake of necrotic cell material by CD8α+ DC but specifically reduces crosspresentation of dead cell-associated antigens in vitro and decreases the immunogenicity of necrotic cells in vivo. The function of CLEC9A requires a key tyrosine residue within its intracellular tail that allows recruitment and activation of the tyrosine kinase Syk, which is also essential for crosspresentation of dead cell-associated antigens. Thus, CLEC9A functions as a Syk-coupled C-type lectin receptor to mediate sensing of necrosis by the principal DC subset involved in regulating crosspriming to cell-associated antigens. PMID:19219027

An approach to the unification of suppressor T cell circuits: a simplified assay for the induction of suppression by T cell-derived, antigen-binding molecules (T-ABM).

PubMed

Chue, B; Ferguson, T A; Beaman, K D; Rosenman, S J; Cone, R E; Flood, P M; Green, D R

1989-01-01

A system is presented in which the in vitro response to sheep red blood cells (SRBC) can be regulated using antigenic determinants coupled to SRBC and T cell-derived antigen-binding molecules (T-ABM) directed against the coupled determinants. T suppressor-inducer factors (TsiF's) are composed of two molecules, one of which is a T-ABM and one which bears I-J determinants (I-J+ molecule). Using two purified T-ABM which have not previously been shown to have in vitro activity, we produced antigen-specific TsiF's which were capable of inducing the suppression of the anti-SRBC response. Suppression was found to require both the T-ABM and the I-J+ molecule, SRBC conjugated with the antigen for which the T-ABM was specific, and a population of Ly-2+ T cells in the culture. Two monoclonal TsiF (or TsF1) were demonstrated to induce suppression of the anti-SRBC response in this system, provided the relevant antigen was coupled to the SRBC in culture. The results are discussed in terms of the general functions of T-ABM in the immune system. This model will be useful in direct, experimental comparisons of the function of T-ABM and suppressor T cell factors under study in different systems and laboratories.

Immunosuppression and induction of anergy by CTLA4Ig in vitro: effects on cellular and antibody responses of lymphocytes from rats with experimental autoimmune myasthenia gravis.

PubMed

McIntosh, K R; Linsley, P S; Drachman, D B

1995-11-01

The pathogenic antibody response to acetylcholine receptor (AChR) in experimental autoimmune myasthenia gravis (EAMG) is T cell dependent. Therefore, it should be possible to design specific immunotherapeutic approaches to treat EAMG (and human MG) by interfering with AChR-specific helper T cells. Productive T cell activation by antigen requires at least two signals: one signal delivered through the T cell receptor by antigen and a second costimulatory signal delivered through the CD28 receptor via the B7 counterreceptor expressed on antigen-presenting cells. Here we show that interference with the B7 costimulatory signal, using a soluble CD28 analogue, CTLA4Ig, resulted in a profound decrease in IL2 production and significantly decreased lymphoproliferative responses and antibody responses by primed lymph node cells from rats with EAMG, when stimulated with AChR in vitro. Nonclonal AChR-specific T cell lines, when stimulated with AChR in the presence of CTLA4Ig, were also inhibited in their ability to proliferate and to produce the cytokines IL2 and IFN-gamma. They remained deficient in their ability to produce IL2 when restimulated with AChR plus fresh antigen-presenting cells and showed variable inhibition of proliferation. The induction of hyporesponsiveness was accompanied by the expression of functional IL2 receptors, as shown by vigorous proliferative responses to addition of exogenous IL2. These results indicate that specific antigen stimulation in the presence of CTLA4Ig can induce certain features typical of anergy. CTLA4Ig provides a promising approach for the immunomodulation of MG and other antibody-mediated autoimmune diseases.

Designing oral vaccines targeting intestinal dendritic cells.

PubMed

Devriendt, Bert; De Geest, Bruno G; Cox, Eric

2011-04-01

Most pathogens colonize and invade the host at mucosal surfaces, such as the lung and the intestine. To combat intestinal pathogens the induction of local adaptive immune responses is required, which is mainly achieved through oral vaccination. However, most vaccines are ineffective when given orally owing to the hostile environment in the gastrointestinal tract. The encapsulation of antigens in biodegradable microparticulate delivery systems enhances their immunogenicity; however, the uptake of these delivery systems by intestinal immune cells is rather poor. Surface decoration of the particulates with targeting ligands could increase the uptake and mediate the selective targeting of the vaccine to intestinal antigen-presenting cells, including dendritic cells. In this review, current knowledge on dendritic cell subsets is discussed, along with progress in the development of selective antigen targeting to these cells, in addition to focusing on data obtained in mice and, where possible, the pig, as a non-rodent animal model for humans. Moreover, the potential use and benefits of Fcγ receptor-mediated targeting of antigen delivery systems are highlighted. In conclusion, dendritic cell targeting ligands grafted on antigen carrier systems should preferably bind to a conserved endocytotic receptor, facilitating the design of a multispecies vaccine platform, which could elicit robust protective immune responses against enteric pathogens.

Sibling rivalry: competition between MHC class II family members inhibits immunity.

PubMed

Denzin, Lisa K; Cresswell, Peter

2013-01-01

Peptide loading of major histocompatibility complex (MHC) class II molecules in the endosomes and lysosomes of antigen-presenting cells is catalyzed by human leukocyte antigen-DM (HLA-DM) and modulated by HLA-DO. In a structural study in this issue, Guce et al. show that HLA-DO is an MHC class II mimic and functions as a competitive and essentially irreversible inhibitor of HLA-DM activity, thereby inhibiting MHC class II antigen presentation.

Blocking Blood Supply to Breast Carcinoma With a DNA Vaccine Encoding VEGF Receptor-2

DTIC Science & Technology

2006-03-01

recognize antigens in the form of 8 to 10 amino acid long peptides, presented to T- cell receptors (TCRs) on the cell surface as complexes with major... receptor , and providing tumor- associated antigens , our DNA vaccine can efficiently activate DCs, NK cells , and CTLs, presumably in Peyer’s patches. The... immunoreceptor in immune cell activation and natural killing. Immunity. 2002;17:19-29. (5) Snyder MR, Weyand CM, Goronzy JJ. The double life of NK receptors

Ontogeny and function of murine epidermal Langerhans cells.

PubMed

Kaplan, Daniel H

2017-09-19

Langerhans cells (LCs) are epidermis-resident antigen-presenting cells that share a common ontogeny with macrophages but function as dendritic cells (DCs). Their development, recruitment and retention in the epidermis is orchestrated by interactions with keratinocytes through multiple mechanisms. LC and dermal DC subsets often show functional redundancy, but LCs are required for specific types of adaptive immune responses when antigen is concentrated in the epidermis. This Review will focus on those developmental and functional properties that are unique to LCs.

High-throughput identification and dendritic cell-based functional validation of MHC class I-restricted Mycobacterium tuberculosis epitopes

PubMed Central

Nair, Smita K.; Tomaras, Georgia D.; Sales, Ana Paula; Boczkowski, David; Chan, Cliburn; Plonk, Kelly; Cai, Yongting; Dannull, Jens; Kepler, Thomas B.; Pruitt, Scott K.; Weinhold, Kent J.

2014-01-01

Emergence of drug-resistant strains of the pathogen Mycobacterium tuberculosis (Mtb) and the ineffectiveness of BCG in curtailing Mtb infection makes vaccine development for tuberculosis an important objective. Identifying immunogenic CD8+ T cell peptide epitopes is necessary for peptide-based vaccine strategies. We present a three-tiered strategy for identifying and validating immunogenic peptides: first, identify peptides that form stable complexes with class I MHC molecules; second, determine whether cytotoxic T lymphocytes (CTLs) raised against the whole protein antigen recognize and lyse target cells pulsed with peptides that passed step 1; third, determine whether peptides that passed step 2, when administered in vivo as a vaccine in HLA-A2 transgenic mice, elicit CTLs that lyse target cells expressing the whole protein antigen. Our innovative approach uses dendritic cells transfected with Mtb antigen-encoding mRNA to drive antigen expression. Using this strategy, we have identified five novel peptide epitopes from the Mtb proteins Apa, Mtb8.4 and Mtb19. PMID:24755960

Endogenous antigen processing drives the primary CD4+ T cell response to influenza

PubMed Central

Miller, Michael A.; Ganesan, Asha Purnima V.; Luckashenak, Nancy; Mendonca, Mark; Eisenlohr, Laurence C.

2015-01-01

By convention, CD4+ T lymphocytes recognize foreign and self peptides derived from internalized antigens in combination with MHC class II molecules. Alternative pathways of epitope production have been identified but their contributions to host defense have not been established. We show here in a mouse infection model that the CD4+ T cell response to influenza, critical for durable protection from the virus, is driven principally by unconventional processing of antigen synthesized within the infected antigen-presenting cell, not by classical processing of endocytosed virions or material from infected cells. Investigation of the cellular components involved, including the H2-M molecular chaperone, the proteasome, and gamma-interferon inducible lysosomal thiol reductase revealed considerable heterogeneity in the generation of individual epitopes, an arrangement that ensures peptide diversity and broad CD4+ T cell engagement. These results could fundamentally revise strategies for rational vaccine design and may lead to key insights into the induction of autoimmune and anti-tumor responses. PMID:26413780

MR1 antigen presentation to mucosal-associated invariant T cells was highly conserved in evolution

PubMed Central

Huang, Shouxiong; Martin, Emmanuel; Kim, Sojung; Yu, Lawrence; Soudais, Claire; Fremont, Daved H.; Lantz, Olivier; Hansen, Ted H.

2009-01-01

Several nonclassical major histocompatibilty antigens (class Ib molecules) have emerged as key players in the early immune response to pathogens or stress. Class Ib molecules activate subsets of T cells that mount effector responses before the adaptive immune system, and thus are called innate T cells. MR1 is a novel class Ib molecule with properties highly suggestive of its regulation of mucosal immunity. The Mr1 gene is evolutionarily conserved, is non-Mhc linked, and controls the development of mucosal-associated invariant T (MAIT) cells. MAIT cells preferentially reside in the gut, and their development is dependent on commensal microbiota. Although these properties suggest that MAIT cells function as innate T cells in the mucosa, this has been difficult to test, due to the (i) paucity of MAIT cells that display MR1-specific activation in vitro and (ii) lack of knowledge of whether or not MR1 presents antigen. Here we show that both mouse and human MAIT cells display a high level of cross-reactivity on mammalian MR1 orthologs, but with differences consistent with limited ligand discrimination. Furthermore, acid eluates from recombinant or cellular MR1 proteins enhance MAIT cell activation in an MR1-specific and cross-species manner. Our findings demonstrate that the presentation pathway of MR1 to MAIT cells is highly evolutionarily conserved. PMID:19416870

Cationic liposomes as vaccine adjuvants.

PubMed

Christensen, Dennis; Korsholm, Karen S; Rosenkrands, Ida; Lindenstrøm, Thomas; Andersen, Peter; Agger, Else Marie

2007-10-01

Cationic liposomes are lipid-bilayer vesicles with a positive surface charge that have re-emerged as a promising new adjuvant technology. Although there is some evidence that cationic liposomes themselves can improve the immune response against coadministered vaccine antigens, their main functions are to protect the antigens from clearance in the body and deliver the antigens to professional antigen-presenting cells. In addition, cationic liposomes can be used to introduce immunomodulators to enhance and modulate the immune response in a desirable direction and, thereby, represent an efficient tool when designing tailor-made adjuvants for specific disease targets. In this article we review the recent progress on cationic liposomes as vehicles, enhancing the effect of immunomodulators and the presentation of vaccine antigens.

Engineering antigens for in situ erythrocyte binding induces T-cell deletion.

PubMed

Kontos, Stephan; Kourtis, Iraklis C; Dane, Karen Y; Hubbell, Jeffrey A

2013-01-02

Antigens derived from apoptotic cell debris can drive clonal T-cell deletion or anergy, and antigens chemically coupled ex vivo to apoptotic cell surfaces have been shown correspondingly to induce tolerance on infusion. Reasoning that a large number of erythrocytes become apoptotic (eryptotic) and are cleared each day, we engineered two different antigen constructs to target the antigen to erythrocyte cell surfaces after i.v. injection, one using a conjugate with an erythrocyte-binding peptide and another using a fusion with an antibody fragment, both targeting the erythrocyte-specific cell surface marker glycophorin A. Here, we show that erythrocyte-binding antigen is collected much more efficiently than free antigen by splenic and hepatic immune cell populations and hepatocytes, and that it induces antigen-specific deletional responses in CD4(+) and CD8(+) T cells. We further validated T-cell deletion driven by erythrocyte-binding antigens using a transgenic islet β cell-reactive CD4(+) T-cell adoptive transfer model of autoimmune type 1 diabetes: Treatment with the peptide antigen fused to an erythrocyte-binding antibody fragment completely prevented diabetes onset induced by the activated, autoreactive CD4(+) T cells. Thus, we report a translatable modular biomolecular approach with which to engineer antigens for targeted binding to erythrocyte cell surfaces to induce antigen-specific CD4(+) and CD8(+) T-cell deletion toward exogenous antigens and autoantigens.

Targeting Alpha-Fetoprotein (AFP)-MHC Complex with CAR T-Cell Therapy for Liver Cancer.

PubMed

Liu, Hong; Xu, Yiyang; Xiang, Jingyi; Long, Li; Green, Shon; Yang, Zhiyuan; Zimdahl, Bryan; Lu, Jingwei; Cheng, Neal; Horan, Lucas H; Liu, Bin; Yan, Su; Wang, Pei; Diaz, Juan; Jin, Lu; Nakano, Yoko; Morales, Javier F; Zhang, Pengbo; Liu, Lian-Xing; Staley, Binnaz K; Priceman, Saul J; Brown, Christine E; Forman, Stephen J; Chan, Vivien W; Liu, Cheng

2017-01-15

The majority of tumor-specific antigens are intracellular and/or secreted and therefore inaccessible by conventional chimeric antigen receptor (CAR) T-cell therapy. Given that all intracellular/secreted proteins are processed into peptides and presented by class I MHC on the surface of tumor cells, we used alpha-fetoprotein (AFP), a specific liver cancer marker, as an example to determine whether peptide-MHC complexes can be targets for CAR T-cell therapy against solid tumors. We generated a fully human chimeric antigen receptor, ET1402L1-CAR (AFP-CAR), with exquisite selectivity and specificity for the AFP 158-166 peptide complexed with human leukocyte antigen (HLA)-A*02:01. We report that T cells expressing AFP-CAR selectively degranulated, released cytokines, and lysed liver cancer cells that were HLA-A*02:01 + /AFP + while sparing cells from multiple tissue types that were negative for either expressed proteins. In vivo, intratumoral injection of AFP-CAR T cells significantly regressed both Hep G2 and AFP 158 -expressing SK-HEP-1 tumors in SCID-Beige mice (n = 8 for each). Moreover, intravenous administration of AFP-CAR T cells in Hep G2 tumor-bearing NSG mice lead to rapid and profound tumor growth inhibition (n = 6). Finally, in an established intraperitoneal liver cancer xenograft model, AFP-CAR T cells showed robust antitumor activity (n = 6). This study demonstrates that CAR T-cell immunotherapy targeting intracellular/secreted solid tumor antigens can elicit a potent antitumor response. Our approach expands the spectrum of antigens available for redirected T-cell therapy against solid malignancies and offers a promising new avenue for liver cancer immunotherapy. Clin Cancer Res; 23(2); 478-88. ©2016 AACR. ©2016 American Association for Cancer Research.

A modern approach for epitope prediction: identification of foot-and-mouth disease virus peptides binding bovine leukocyte antigen (BoLA) class I molecules

USDA-ARS?s Scientific Manuscript database

Major histocompatibility complex (MHC) class I molecules regulate adaptive immune responses through the presentation of antigenic peptides to CD8positive T-cells. Polymorphisms in the peptide binding region of class I molecules determine peptide binding affinity and stability during antigen presenta...

Immunoglobulins and Disease.

DTIC Science & Technology

1987-02-06

MICROCOPY RESOLUTION TEST CHART NA7 UNAL hu AT ’N Ad . . 0 0 10 PROCEEDINGS THE TWENTY-SIXTH MIDWINTER CONFERENCE OF IMMUNOLOGISTS January 17-20, 1987...monoclonal BA5 has also been used to study the abnormal regulation of expression of BCGF receptors in common variable ilmmunodeficiency, SLE , and B cell...antigen, we have shown that membrane Ig-mediated antigen presentation by small B cells to rabbit globulin - specific T cell lines results in a vigorous

Simultaneous targeting of tumor antigens and the tumor vasculature using T lymphocyte transfer synergize to induce regression of established tumors in mice.

PubMed

Chinnasamy, Dhanalakshmi; Tran, Eric; Yu, Zhiya; Morgan, Richard A; Restifo, Nicholas P; Rosenberg, Steven A

2013-06-01

Most systemic cancer therapies target tumor cells directly, although there is increasing interest in targeting the tumor stroma that can comprise a substantial portion of the tumor mass. We report here a synergy between two T-cell therapies, one directed against the stromal tumor vasculature and the other directed against antigens expressed on the tumor cell. Simultaneous transfer of genetically engineered syngeneic T cells expressing a chimeric antigen receptor targeting the VEGF receptor-2 (VEGFR2; KDR) that is overexpressed on tumor vasculature and T-cells specific for the tumor antigens gp100 (PMEL), TRP-1 (TYRP1), or TRP-2 (DCT) synergistically eradicated established B16 melanoma tumors in mice and dramatically increased the tumor-free survival of mice compared with treatment with either cell type alone or T cells coexpressing these two targeting molecules. Host lymphodepletion before cell transfer was required to mediate the antitumor effect. The synergistic antitumor response was accompanied by a significant increase in the infiltration and expansion and/or persistence of the adoptively transferred tumor antigen-specific T cells in the tumor microenvironment and thus enhanced their antitumor potency. The data presented here emphasize the possible beneficial effects of combining antiangiogenic with tumor-specific immunotherapeutic approaches for the treatment of patients with cancer. ©2013 AACR.

Carbon nanotubes as vaccine scaffolds

PubMed Central

Scheinberg, David A.; McDevitt, Michael R.; Dao, Tao; Mulvey, Justin J.; Feinberg, Evan; Alidori, Simone

2013-01-01

Carbon nanotubes display characteristics that are potentially useful in their development as scaffolds for vaccine compositions. These features include stability in vivo, lack of intrinsic immunogenicity, low toxicity, and the ability to be appended with multiple copies of antigens. In addition, the particulate nature of carbon nanotubes and their unusual properties of rapid entry into antigen-presenting cells, such as dendritic cells, make them especially useful as carriers of antigens. Early attempts demonstrating carbon nanotube-based vaccines can be used in both infectious disease settings and cancer are promising. PMID:23899863

A β-Lactam Antibiotic Dampens Excitotoxic Inflammatory CNS Damage in a Mouse Model of Multiple Sclerosis

PubMed Central

Torres-Salazar, Delany; Bittner, Stefan; Zozulya, Alla L.; Weidenfeller, Christian; Kotsiari, Alexandra; Stangel, Martin; Fahlke, Christoph; Wiendl, Heinz

2008-01-01

In multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE), impairment of glial “Excitatory Amino Acid Transporters” (EAATs) together with an excess glutamate-release by invading immune cells causes excitotoxic damage of the central nervous system (CNS). In order to identify pathways to dampen excitotoxic inflammatory CNS damage, we assessed the effects of a β-lactam antibiotic, ceftriaxone, reported to enhance expression of glial EAAT2, in “Myelin Oligodendrocyte Glycoprotein” (MOG)-induced EAE. Ceftriaxone profoundly ameliorated the clinical course of murine MOG-induced EAE both under preventive and therapeutic regimens. However, ceftriaxone had impact neither on EAAT2 protein expression levels in several brain areas, nor on the radioactive glutamate uptake capacity in a mixed primary glial cell-culture and the glutamate-induced uptake currents in a mammalian cell line mediated by EAAT2. Moreover, the clinical effect of ceftriaxone was preserved in the presence of the EAAT2-specific transport inhibitor, dihydrokainate, while dihydrokainate alone caused an aggravated EAE course. This demonstrates the need for sufficient glial glutamate uptake upon an excitotoxic autoimmune inflammatory challenge of the CNS and a molecular target of ceftriaxone other than the glutamate transporter. Ceftriaxone treatment indirectly hampered T cell proliferation and proinflammatory INFγ and IL17 secretion through modulation of myelin-antigen presentation by antigen-presenting cells (APCs) e.g. dendritic cells (DCs) and reduced T cell migration into the CNS in vivo. Taken together, we demonstrate, that a β-lactam antibiotic attenuates disease course and severity in a model of autoimmune CNS inflammation. The mechanisms are reduction of T cell activation by modulation of cellular antigen-presentation and impairment of antigen-specific T cell migration into the CNS rather than or modulation of central glutamate homeostasis. PMID:18773080

Cytotoxic T cell clones isolated from ovarian tumor-infiltrating lymphocytes recognize multiple antigenic epitopes on autologous tumor cells.

PubMed

Ioannides, C G; Freedman, R S; Platsoucas, C D; Rashed, S; Kim, Y P

1991-03-01

CTL clones were developed from tumor infiltrating lymphocytes (TIL) from the ascites of a patient with ovarian carcinoma by coculture of TIL with autologous tumor cells and subsequent cloning in the presence of autologous tumor cells. These CTL clones expressed preferential cytolytic activity against autologous tumor cells but not against allogeneic ovarian tumor cells and the NK-sensitive cell line K562. The cytolytic activity of these CTL against autologous tumors was inhibited by anti-TCR (WT31 mAb), anti-HLA class I, and anti-CD3 mAb but not by the NK function antibody Leu 11b. Cloning of the autologous tumor cells in vitro revealed that the CTL clones of the ovarian TIL expressed differential abilities to lyse autologous tumor cell clones. The specificity analysis of these autologous tumor specific CTL suggested that they recognize several antigenic determinants present on the ovarian tumor cells. Our results indicate the presence of at least three antigenic epitopes on the tumor cells (designated OVA-1A, OVA-1B, and OVA-1C), one of which (OVA-1C) is unstable. These determinants are present either simultaneously or separately, and six types of ovarian clones can be distinguished on the basis of their expression. These results indicate that CTL of the TIL detect intratumor antigenic heterogeneity. The novel heterogeneity identified within the ovarian tumor cells in this report may be of significance for understanding cellular immunity in ovarian cancer and developing adoptive specific immunotherapeutic approaches in ovarian cancer.

Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors

PubMed Central

Karlsson, Hannah; Svensson, Emma; Gigg, Camilla; Jarvius, Malin; Olsson-Strömberg, Ulla; Savoldo, Barbara; Dotti, Gianpietro; Loskog, Angelica

2015-01-01

CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G) CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G) CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs. PMID:26700307

A Safe Bacterial Microsyringe for In Vivo Antigen Delivery and Immunotherapy

PubMed Central

Le Gouëllec, Audrey; Chauchet, Xavier; Laurin, David; Aspord, Caroline; Verove, Julien; Wang, Yan; Genestet, Charlotte; Trocme, Candice; Ahmadi, Mitra; Martin, Sandrine; Broisat, Alexis; Cretin, François; Ghezzi, Catherine; Polack, Benoit; Plumas, Joël; Toussaint, Bertrand

2013-01-01

The industrial development of active immunotherapy based on live-attenuated bacterial vectors has matured. We developed a microsyringe for antigen delivery based on the type III secretion system (T3SS) of P. aeruginosa. We applied the “killed but metabolically active” (KBMA) attenuation strategy to make this bacterial vector suitable for human use. We demonstrate that attenuated P. aeruginosa has the potential to deliver antigens to human antigen-presenting cells in vitro via T3SS with considerable attenuated cytotoxicity as compared with the wild-type vector. In a mouse model of cancer, we demonstrate that this KBMA strain, which cannot replicate in its host, efficiently disseminates into lymphoid organs and delivers its heterologous antigen. The attenuated strain effectively induces a cellular immune response to the cancerous cells while lowering the systemic inflammatory response. Hence, a KBMA P. aeruginosa microsyringe is an efficient and safe tool for in vivo antigen delivery. PMID:23531551

A and B antigen levels acquired by group O donor-derived erythrocytes following ABO-non-identical transfusion or minor ABO-incompatible haematopoietic stem cell transplantation.

PubMed

Hult, A K; Dykes, J H; Storry, J R; Olsson, M L

2017-06-01

ABO-incompatible haematopoietic stem cell transplantation (HSCT) presents a challenge to blood component transfusion. The aim of this study was to investigate the weak blood group A or B antigen expression by donor-derived group O red blood cells (RBC) observed following transfusion or minor ABO-incompatible HSCT. In addition, in vitro experiments were performed to elucidate possible mechanisms underlying this phenomenon. A sensitive flow cytometry assay for the semi-quantification of RBC A/B antigen levels was used to assess patient samples and evaluate in vitro experiments. Analysis of blood samples from patients, originally typed as A, B and AB but recently transplanted or transfused with cells from group O donors, revealed the A antigen expression on donor-derived RBC, ranging from very low levels in non-secretor individuals to almost subgroup A x -like profiles in group A secretors. The B antigen expression was less readily detectable. In vitro experiments, in which group O donor RBC were incubated with (i) group A/B secretor/non-secretor donor plasma or (ii) group A/B donor RBC in the absence of plasma, supported the proposed adsorption of A/B antigen-bearing glycolipids from secretor plasma but also indicated a secretor-independent mechanism for A/B antigen acquisition as well as direct cell-to-cell transfer of ABO antigens. The in vivo conversion of donor-derived blood group O RBC to ABO subgroup-like RBC after transfusion or minor ABO-incompatible HSCT raises the question of appropriate component selection. Based on these data, AB plasma should be transfused following ABO-incompatible HSCT. © 2017 British Blood Transfusion Society.

Immune complexome analysis reveals the specific and frequent presence of immune complex antigens in lung cancer patients: A pilot study.

PubMed

Ohyama, Kaname; Yoshimi, Haruka; Aibara, Nozomi; Nakamura, Yoichi; Miyata, Yasuyoshi; Sakai, Hideki; Fujita, Fumihiko; Imaizumi, Yoshitaka; Chauhan, Anil K; Kishikawa, Naoya; Kuroda, Naotaka

2017-01-15

Cancer immunotherapies such as antibodies targeting T cell checkpoints, or adaptive tumor-infiltrating lymphocyte (TIL) transfer, have been developed to boost the endogenous immune response against human malignancies. However, activation of T cells by such antibodies can lead to the risk of autoimmune diseases. Also, the selection of tumor-reactive T cells for TIL relies on information regarding mutated antigens in tumors and does not reflect other factors involved in protein antigenicity. It is therefore essential to engineer therapeutic interventions by which T cell reactivity against tumor cells is selectively enhanced (i.e., "focused cancer immunotherapy") based on tumor antigens that are specifically expressed in the tumor of a certain cancer and in many patients with this cancer. Immune complexes (ICs) are the direct and stable products of immunological recognition by humoral immunity. Here, we searched for tumor-specific IC antigens in each of five cancers (lung (n = 28), colon (n = 20), bladder (n = 20), renal cell (n = 15) and malignant lymphoma (n = 9)), by using immune complexome analysis that comprehensively identifies and profiles the constituent antigens in ICs. This analysis indicated that gelsolin and inter-alpha-trypsin inhibitor heavy chains were specifically and frequently detected (at a frequency higher than 80%), and that phosphoproteins (VENTX, VCIP135) were also specifically present in the ICs of lung cancer patients. Immune complexome analysis successfully identified several tumor-specific IC antigens with high detection frequency in lung cancer patients. These specific antigens are required to validate the clinical benefit by further analysis using a large number of patients. © 2016 UICC.

CD8 Memory Cells Develop Unique DNA Repair Mechanisms Favoring Productive Division.

PubMed

Galgano, Alessia; Barinov, Aleksandr; Vasseur, Florence; de Villartay, Jean-Pierre; Rocha, Benedita

2015-01-01

Immune responses are efficient because the rare antigen-specific naïve cells are able to proliferate extensively and accumulate upon antigen stimulation. Moreover, differentiation into memory cells actually increases T cell accumulation, indicating improved productive division in secondary immune responses. These properties raise an important paradox: how T cells may survive the DNA lesions necessarily induced during their extensive division without undergoing transformation. We here present the first data addressing the DNA damage responses (DDRs) of CD8 T cells in vivo during exponential expansion in primary and secondary responses in mice. We show that during exponential division CD8 T cells engage unique DDRs, which are not present in other exponentially dividing cells, in T lymphocytes after UV or X irradiation or in non-metastatic tumor cells. While in other cell types a single DDR pathway is affected, all DDR pathways and cell cycle checkpoints are affected in dividing CD8 T cells. All DDR pathways collapse in secondary responses in the absence of CD4 help. CD8 T cells are driven to compulsive suicidal divisions preventing the propagation of DNA lesions. In contrast, in the presence of CD4 help all the DDR pathways are up regulated, resembling those present in metastatic tumors. However, this up regulation is present only during the expansion phase; i.e., their dependence on antigen stimulation prevents CD8 transformation. These results explain how CD8 T cells maintain genome integrity in spite of their extensive division, and highlight the fundamental role of DDRs in the efficiency of CD8 immune responses.

Antigen Presentation Keeps Trending in Immunotherapy Resistance.

PubMed

Kalbasi, Anusha; Ribas, Antoni

2018-04-19

Through a gain-of-function kinome screen, MEX3B was identified as a mediator of resistance to T-cell immunotherapy not previously identified using CRISPR-based screens. MEX3B is a posttranscriptional regulator of HLA-A, validating the critical role of tumor-intrinsic antigen presentation in T-cell immunotherapy and indicating a new putative molecular target. Clin Cancer Res; 24(14); 1-3. ©2018 AACR. See related article by Huang et al., p. xxxx . ©2018 American Association for Cancer Research.

Fc receptors for mouse IgG1 on human monocytes: polymorphism and role in antibody-induced T cell proliferation.

PubMed

Tax, W J; Hermes, F F; Willems, R W; Capel, P J; Koene, R A

1984-09-01

In previous studies, it was shown that there is polymorphism in the mitogenic effect of mouse IgG1 monoclonal antibodies against the T3 antigen of human T cells. This polymorphism implies that IgG1 anti-T3 antibodies are not mitogenic for T cells from 30% of healthy individuals. The present results demonstrate that this polymorphism is caused by polymorphism of an Fc receptor for mouse IgG1, present on human monocytes. The Fc receptor for murine IgG1 could be detected by a newly developed rosetting assay on monocytes from all individuals responsive to the mitogenic effect of IgG1 anti-T3 antibodies. This Fc receptor was not detectable on monocytes from those individuals exhibiting no mitogenic responses to IgG1 anti-T3 monoclonal antibodies. Cross-linking of T3 antigens appears to be essential for antibody-induced mitosis of T cells, because mononuclear cells that did not proliferate in response to WT 31 (an IgG1 antibody against T3 antigen) showed a proliferative response to Sepharose beads coated with WT 31. The Fc receptor--if functionally present--may be involved in the cross-linking of T3 antigens through anti-T3 antibodies. Further evidence for the involvement of this Fc receptor in antibody-induced T cell proliferation was provided by inhibition studies. Immune complexes containing IgG1 antibodies were able to inhibit the proliferative response to IgG1 anti-T3 antibodies. This inhibition by immune complexes appears to be mediated through the monocyte Fc receptor for mouse IgG1. These findings are important for the interpretation of previously described inhibitory effects of anti-T cell monoclonal antibodies on T cell proliferation, and show that such inhibitory effects may be monocyte-mediated (via immune complexes) rather than caused by a direct involvement of the respective T cell antigens in T cell mitosis. The Fc receptor for mouse IgG1 plays a role in antibody-induced T cell proliferation. Its polymorphism may have important implications for the therapeutic use of IgG1 monoclonal antibodies.

Tomatine adjuvantation of protective immunity to a major pre-erythrocytic vaccine candidate of malaria is mediated via CD8+ T cell release of IFN-gamma.

PubMed

Heal, Karen G; Taylor-Robinson, Andrew W

2010-01-01

The glycoalkaloid tomatine, derived from the wild tomato, can act as a powerful adjuvant to elicit an antigen-specific cell-mediated immune response to the circumsporozoite (CS) protein, a major pre-erythrocytic stage malaria vaccine candidate antigen. Using a defined MHC-class-I-restricted CS epitope in a Plasmodium berghei rodent model, antigen-specific cytotoxic T lymphocyte activity and IFN-gamma secretion ex vivo were both significantly enhanced compared to responses detected from similarly stimulated splenocytes from naive and tomatine-saline-immunized mice. Further, through lymphocyte depletion it is demonstrated that antigen-specific IFN-gamma is produced exclusively by the CD8(+) T cell subset. We conclude that the processing of the P. berghei CS peptide as an exogenous antigen and its presentation via MHC class I molecules to CD8(+) T cells leads to an immune response that is an in vitro correlate of protection against pre-erythrocytic malaria. Further characterization of tomatine as an adjuvant in malaria vaccine development is indicated.

Designer dendritic cells for tolerance induction: guided not misguided missiles.

PubMed

Hackstein, H; Morelli, A E; Thomson, A W

2001-08-01

Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that play crucial roles as initiators and modulators of adaptive immune responses. Although DC-based vaccines have been utilized successfully to generate cytolytic T-cell activity against tumor antigens (Ags), evidence has accumulated that DCs also have potent capabilities to tolerize T cells in an Ag-specific manner. DCs cultured in the laboratory can suppress auto- or alloimmunity. Current and prospective strategies to promote this inherent tolerogenic potential of DCs might prove to be important for the therapy of transplant rejection and autoimmune diseases.

B cells and TCR avidity determine distinct functions of CD4+ T cells in retroviral infection1

PubMed Central

Ploquin, Mickaël J-Y; Eksmond, Urszula; Kassiotis, George

2011-01-01

The T-cell-dependent B-cell response relies on cognate interaction between B cells and CD4+ Th cells. However, the consequences of this interaction for CD4+ T cells are not entirely known. B cells generally promote CD4+ T-cell responses to pathogens, albeit to a variable degree. In contrast, CD4+ T-cell responses to self or tumor antigens are often suppressed by B cells. Here we demonstrated that interaction with B cells dramatically inhibited the function of virus-specific CD4+ T cells in retroviral infection. We have used Friend virus (FV) infection of mice as a model for retroviral infection, in which the behavior of virus-specific CD4+ T cells was monitored according to their TCR avidity. We report that avidity for antigen and interaction with B cells determine distinct aspects of the primary CD4+ T-cell response to FV infection. Virus-specific CD4+ T cells followed exclusive Th1 and T follicular helper (Tfh) differentiation. High avidity for antigen facilitated expansion during priming and enhanced the capacity for IFN-γ and IL-21 production. In contrast, Tfh differentiation was not affected by avidity for antigen. By reducing or preventing B-cell interaction we found that B cells promoted Tfh differentiation, induced programmed death 1 (PD-1) expression and inhibited IFN-γ production by virus-specific CD4+ T cells. Ultimately, B cells protected hosts from CD4+ T-cell-mediated immune pathology, at the detriment of CD4+ T-cell-mediated protective immunity. Our results suggest that B-cell presentation of vaccine antigens could be manipulated to direct the appropriate CD4+ T-cell response. PMID:21841129

Immunological Properties of Hepatitis B Core Antigen Fusion Proteins

NASA Astrophysics Data System (ADS)

Francis, Michael J.; Hastings, Gillian Z.; Brown, Alan L.; Grace, Ken G.; Rowlands, David J.; Brown, Fred; Clarke, Berwyn E.

1990-04-01

The immunogenicity of a 19 amino acid peptide from foot-and-mouth disease virus has previously been shown to approach that of the inactivated virus from which it was derived after multimeric particulate presentation as an N-terminal fusion with hepatitis B core antigen. In this report we demonstrate that rhinovirus peptide-hepatitis B core antigen fusion proteins are 10-fold more immunogenic than peptide coupled to keyhole limpet hemocyanin and 100-fold more immunogenic than uncoupled peptide with an added helper T-cell epitope. The fusion proteins can be readily administered without adjuvant or with adjuvants acceptable for human and veterinary application and can elicit a response after nasal or oral dosing. The fusion proteins can also act as T-cell-independent antigens. These properties provide further support for their suitability as presentation systems for "foreign" epitopes in the development of vaccines.

GS143, an I{kappa}B ubiquitination inhibitor, inhibits allergic airway inflammation in mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hirose, Koichi; Wakashin, Hidefumi; Oki, Mie

2008-09-26

Asthma is characterized by airway inflammation with intense eosinophil infiltration and mucus hyper-production, in which antigen-specific Th2 cells play critical roles. Nuclear factor-{kappa}B (NF-{kappa}B) pathway has been demonstrated to be essential for the production of Th2 cytokines and chemokines in the airways in murine asthma models. In the present study, we examined the effect of GS143, a novel small-molecule inhibitor of I{kappa}B ubiquitination, on antigen-induced airway inflammation and Th2 cytokine production in mice. Intranasal administration of GS143 prior to antigen challenge suppressed antigen-induced NF-{kappa}B activation in the lung of sensitized mice. Intranasal administration of GS143 also inhibited antigen-induced eosinophil andmore » lymphocyte recruitment into the airways as well as the expression of Th2 cytokines and eotaxin in the airways. Moreover, GS143 inhibited antigen-induced differentiation of Th2 cells but not of Th1 cells in vitro. Taken together, these results suggest that I{kappa}B ubiquitination inhibitor may have therapeutic potential against asthma.« less

Immunotherapy Plus Cryotherapy: Potential Augmented Abscopal Effect for Advanced Cancers

PubMed Central

Abdo, Joe; Cornell, David L.; Mittal, Sumeet K.; Agrawal, Devendra K.

2018-01-01

Since the 1920s the gold standard for treating cancer has been surgery, which is typically preceded or followed with chemotherapy and/or radiation, a process that perhaps contributes to the destruction of a patient’s immune defense system. Cryosurgery ablation of a solid tumor is mechanistically similar to a vaccination where hundreds of unique antigens from a heterogeneous population of tumor cells derived from the invading cancer are released. However, releasing tumor-derived self-antigens into circulation may not be sufficient enough to overcome the checkpoint escape mechanisms some cancers have evolved to avoid immune responses. The potentiated immune response caused by blocking tumor checkpoints designed to prevent programmed cell death may be the optimal treatment method for the immune system to recognize these new circulating cryoablated self-antigens. Preclinical and clinical evidence exists for the complementary roles for Cytotoxic T-lymphocyte-associated protein (CTLA-4) and PD-1 antagonists in regulating adaptive immunity, demonstrating that combination immunotherapy followed by cryosurgery provides a more targeted immune response to distant lesions, a phenomenon known as the abscopal effect. We propose that when the host’s immune system has been “primed” with combined anti-CTLA-4 and anti-PD-1 adjuvants prior to cryosurgery, the preserved cryoablated tumor antigens will be presented and processed by the host’s immune system resulting in a robust cytotoxic CD8+ T-cell response. Based on recent investigations and well-described biochemical mechanisms presented herein, a polyvalent autoinoculation of many tumor-specific antigens, derived from a heterogeneous population of tumor cancer cells, would present to an unhindered yet pre-sensitized immune system yielding a superior advantage in locating, recognizing, and destroying tumor cells throughout the body. PMID:29644213

Comprehensive Analysis of Cytomegalovirus pp65 Antigen-Specific CD8+ T Cell Responses According to Human Leukocyte Antigen Class I Allotypes and Intraindividual Dominance.

PubMed

Hyun, Seung-Joo; Sohn, Hyun-Jung; Lee, Hyun-Joo; Lee, Seon-Duk; Kim, Sueon; Sohn, Dae-Hee; Hong, Cheol-Hwa; Choi, Haeyoun; Cho, Hyun-Il; Kim, Tai-Gyu

2017-01-01

To define whether individual human leukocyte antigen (HLA) class I allotypes are used preferentially in human cytomegalovirus (CMV)-specific cytotoxic T lymphocyte responses, CD8 + T cell responses restricted by up to six HLA class I allotypes in an individual were measured in parallel using K562-based artificial antigen-presenting cells expressing both CMV pp65 antigen and one of 32 HLA class I allotypes (7 HLA-A, 14 HLA-B, and 11 HLA-C) present in 50 healthy Korean donors. The CD8 + T cell responses to pp65 in the HLA-C allotypes were lower than responses to those in HLA-A and -B allotypes and there was no difference between the HLA-A and HLA-B loci. HLA-A*02:01, -B*07:02, and -C*08:01 showed the highest magnitude and frequency of immune responses to pp65 at each HLA class I locus. However, HLA-A*02:07, -B*59:01, -B*58:01, -B*15:11, -C*03:02, and -C*02:02 did not show any immune responses. Although each individual has up to six different HLA allotypes, 46% of the donors showed one allotype, 24% showed two allotypes, and 2% showed three allotypes that responded to pp65. Interestingly, the frequencies of HLA-A alleles were significantly correlated with the positivity of specific allotypes. Our results demonstrate that specific HLA class I allotypes are preferentially used in the CD8 + T cell immune response to pp65 and that a hierarchy among HLA class I allotypes is present in an individual.

Upregulation of Immunoproteasome Subunits in Myositis Indicates Active Inflammation with Involvement of Antigen Presenting Cells, CD8 T-Cells and IFNγ

PubMed Central

Ghannam, Khetam; Martinez-Gamboa, Lorena; Spengler, Lydia; Krause, Sabine; Smiljanovic, Biljana; Bonin, Marc; Bhattarai, Salyan; Grützkau, Andreas; Burmester, Gerd-R.

2014-01-01

Objective In idiopathic inflammatory myopathies (IIM) infiltration of immune cells into muscle and upregulation of MHC-I expression implies increased antigen presentation and involvement of the proteasome system. To decipher the role of immunoproteasomes in myositis, we investigated individual cell types and muscle tissues and focused on possible immune triggers. Methods Expression of constitutive (PSMB5, -6, -7) and corresponding immunoproteasomal subunits (PSMB8, -9, -10) was analyzed by real-time RT-PCR in muscle biopsies and sorted peripheral blood cells of patients with IIM, non-inflammatory myopathies (NIM) and healthy donors (HD). Protein analysis in muscle biopsies was performed by western blot. Affymetrix HG-U133 platform derived transcriptome data from biopsies of different muscle diseases and from immune cell types as well as monocyte stimulation experiments were used for validation, coregulation and coexpression analyses. Results Real-time RT-PCR revealed significantly increased expression of immunoproteasomal subunits (PSMB8/-9/-10) in DC, monocytes and CD8+ T-cells in IIM. In muscle biopsies, the immunosubunits were elevated in IIM compared to NIM and exceeded levels of matched blood samples. Proteins of PSMB8 and -9 were found only in IIM but not NIM muscle biopsies. Reanalysis of 78 myositis and 20 healthy muscle transcriptomes confirmed these results and revealed involvement of the antigen processing and presentation pathway. Comparison with reference profiles of sorted immune cells and healthy muscle confirmed upregulation of PSMB8 and -9 in myositis biopsies beyond infiltration related changes. This upregulation correlated highest with STAT1, IRF1 and IFNγ expression. Elevation of T-cell specific transcripts in active IIM muscles was accompanied by increased expression of DC and monocyte marker genes and thus reflects the cell type specific involvement observed in peripheral blood. Conclusions Immunoproteasomes seem to indicate IIM activity and suggest that dominant involvement of antigen processing and presentation may qualify these diseases exemplarily for the evolving therapeutic concepts of immunoproteasome specific inhibition. PMID:25098831

Upregulation of immunoproteasome subunits in myositis indicates active inflammation with involvement of antigen presenting cells, CD8 T-cells and IFNΓ.

PubMed

Ghannam, Khetam; Martinez-Gamboa, Lorena; Spengler, Lydia; Krause, Sabine; Smiljanovic, Biljana; Bonin, Marc; Bhattarai, Salyan; Grützkau, Andreas; Burmester, Gerd-R; Häupl, Thomas; Feist, Eugen

2014-01-01

In idiopathic inflammatory myopathies (IIM) infiltration of immune cells into muscle and upregulation of MHC-I expression implies increased antigen presentation and involvement of the proteasome system. To decipher the role of immunoproteasomes in myositis, we investigated individual cell types and muscle tissues and focused on possible immune triggers. Expression of constitutive (PSMB5, -6, -7) and corresponding immunoproteasomal subunits (PSMB8, -9, -10) was analyzed by real-time RT-PCR in muscle biopsies and sorted peripheral blood cells of patients with IIM, non-inflammatory myopathies (NIM) and healthy donors (HD). Protein analysis in muscle biopsies was performed by western blot. Affymetrix HG-U133 platform derived transcriptome data from biopsies of different muscle diseases and from immune cell types as well as monocyte stimulation experiments were used for validation, coregulation and coexpression analyses. Real-time RT-PCR revealed significantly increased expression of immunoproteasomal subunits (PSMB8/-9/-10) in DC, monocytes and CD8+ T-cells in IIM. In muscle biopsies, the immunosubunits were elevated in IIM compared to NIM and exceeded levels of matched blood samples. Proteins of PSMB8 and -9 were found only in IIM but not NIM muscle biopsies. Reanalysis of 78 myositis and 20 healthy muscle transcriptomes confirmed these results and revealed involvement of the antigen processing and presentation pathway. Comparison with reference profiles of sorted immune cells and healthy muscle confirmed upregulation of PSMB8 and -9 in myositis biopsies beyond infiltration related changes. This upregulation correlated highest with STAT1, IRF1 and IFNγ expression. Elevation of T-cell specific transcripts in active IIM muscles was accompanied by increased expression of DC and monocyte marker genes and thus reflects the cell type specific involvement observed in peripheral blood. Immunoproteasomes seem to indicate IIM activity and suggest that dominant involvement of antigen processing and presentation may qualify these diseases exemplarily for the evolving therapeutic concepts of immunoproteasome specific inhibition.

Engineered outer membrane vesicle is potent to elicit HPV16E7-specific cellular immunity in a mouse model of TC-1 graft tumor.

PubMed

Wang, Shijie; Huang, Weiwei; Li, Kui; Yao, Yufeng; Yang, Xu; Bai, Hongmei; Sun, Wenjia; Liu, Cunbao; Ma, Yanbing

2017-01-01

Currently, therapeutic tumor vaccines under development generally lack significant effects in human clinical trials. Exploring a powerful antigen delivery system is a potential approach to improve vaccine efficacy. We sought to explore engineered bacterial outer membrane vesicles (OMVs) as a new vaccine carrier for efficiently delivering tumor antigens and provoking robust antitumor immune responses. First, the tumoral antigen human papillomavirus type 16 early protein E7 (HPV16E7) was presented on Escherichia coli -derived OMVs by genetic engineering methods, acquiring the recombinant OMV vaccine. Second, the ability of recombinant OMVs delivering their components and the model antigen green fluorescent protein to antigen-presenting cells was investigated in the macrophage Raw264.7 cells and in bone marrow-derived dendritic cells in vitro. Third, it was evaluated in TC-1 graft tumor model in mice that the recombinant OMVs displaying HPV16E7 stimulated specific cellular immune response and intervened the growth of established tumor. E. coli DH5α-derived OMVs could be taken up rapidly by dendritic cells, for which vesicle structure has been proven to be important. OMVs significantly stimulated the expression of dendritic cellmaturation markers CD80, CD86, CD83 and CD40. The HPV16E7 was successfully embedded in engineered OMVs through gene recombinant techniques. Subcutaneous immunization with the engineered OMVs induced E7 antigen-specific cellular immune responses, as shown by the increased numbers of interferon-gamma-expressing splenocytes by enzyme-linked immunospot assay and interferon-gamma-expressing CD4 + and CD8 + cells by flow cytometry analyses. Furthermore, the growth of grafted TC-1 tumors in mice was significantly suppressed by therapeutic vaccination. The recombinant E7 proteins presented by OMVs were more potent than those mixed with wild-type OMVs or administered alone for inducing specific cellular immunity and suppressing tumor growth. The results indicated that the nano-grade OMVs might be a useful vaccine platform for antigen delivery in cancer immunotherapy.

Development of Epstein-Barr virus-specific memory T cell receptor clonotypes in acute infectious mononucleosis

PubMed Central

1996-01-01

The importance of cytotoxic T lymphocytes (CTLs) in the immunosurveillance of Epstein-Barr virus (EBV)-infected B cells is firmly established, and the viral antigens of CTL recognition in latent infection are well defined. The epitopes targeted by CTLs during primary infection have not been identified, however, and there is only limited information about T cell receptor (TCR) selection. In the present report, we have monitored the development of memory TCR-beta clonotypes selected in response to natural EBV infection in a longitudinal study of an HLA-B8+ individual with acute infectious mononucleosis (IM). By stimulating peripheral blood lymphocytes with HLA-B8+ EBV-transformed B lymphoblastoid cells, the primary virus- specific CTL response was shown to include specificities for two HLA-B8- restricted antigenic determinants, FLRGRAYGL and QAKWRLQTL, which are encoded within the latent EBV nuclear antigen EBNA-3. TCR-beta sequence analysis of CTL clones specific for each epitope showed polyclonal TCR- beta repertoire selection, with structural restrictions on recognition that indicated antigen-driven selection. Furthermore, longitudinal repertoire analysis revealed long-term preservation of a multiclonal effector response throughout convalescence, with the reemergence of distinct memory T cell clonotypes sharing similar structural restrictions. Tracking the progression of specific TCR-beta clonotypes and antigen-specific TCR-V beta family gene expression in the peripheral repertoire ex vivo using semiquantitative PCR strongly suggested that selective TCR-beta expansions were present at the clonotype level, but not at the TCR-V beta family level. Overall, in this first analysis of antigen-specific TCR development in IM, a picture of polyclonal TCR stimulation is apparent. This diversity may be especially important in the establishment of an effective CTL control during acute EBV infection and in recovery from disease. PMID:8920869

Toxocara canis adult worm antigen induces proliferative response of healthy human peripheral blood mononuclear cells.

PubMed

Inuo, G; Akao, N; Kohsaka, H; Saito, I; Miyasaka, N; Fujita, K

1995-02-01

The proliferative response of human peripheral blood mononuclear cells (PBMC) from healthy donors to Toxocara canis adult worm antigens (TcA) was examined. PBMC from all donors examined (n = 7) strongly responded to TcA in a dose-dependent fashion after six days of culture, irrespective of their serological reactivity. In contrast, cord blood mononuclear cells did not react to TcA. The proliferation of PBMC in response to TcA was completely inhibited by anti-HLA-DR antibody. Purified CD4+ T cells reconstituted with autologous irradiated antigen presenting cells (APC) vigorously proliferated in response to TcA, but this was abrogated by pretreatment of APC with paraformaldehyde. Significant IL-2, IL-3, IL-4, IL-5 and IFN-gamma mRNA expression was detected in PBMC stimulated with TcA, with expression peaking at 72 h after stimulation. IL-1 beta, IL-6, IL-10 and GM-CSF mRNA expression was also upregulated, peaking at 24 h after stimulation. Taken together, these results suggest that adult T. canis-derived antigens have the ability to activate human PBMC as conventional antigens, possibly due to their cross-reactivity, which may be involved in the host defence against helminth infection.

Paracrine release of IL-2 and anti-CTLA-4 enhances the ability of artificial polymer antigen-presenting cells to expand antigen-specific T cells and inhibit tumor growth in a mouse model.

PubMed

Zhang, Lei; Wang, Limin; Shahzad, Khawar Ali; Xu, Tao; Wan, Xin; Pei, Weiya; Shen, Chuanlai

2017-09-01

Accumulating evidence indicates that bead-based artificial antigen-presenting cells (aAPCs) are a powerful tool to induce antigen-specific T cell responses in vitro and in vivo. To date, most conventional aAPCs have been generated by coupling an antigen signal (signal 1) and one or two costimulatory signals, such as anti-CD28 with anti-LFA1 or anti-4-1BB (signal 2), onto the surfaces of cell-sized or nanoscale magnetic beads or polyester latex beads. The development of a biodegradable scaffold and the combined use of multiple costimulatory signals as well as third signals for putative clinical applications is the next step in the development of this technology. Here, a novel biodegradable aAPC platform for active immunotherapy was developed by co-encapsulating IL-2 and anti-CTLA-4 inside cell-sized polylactic-co-glycolic acid microparticles (PLGA-MPs) while co-coupling an H-2K b /TRP2-Ig dimer and anti-CD28 onto the surface. Cytokines (activating signal) and antibodies (anti-inhibition signal) were efficiently co-encapsulated in PLGA-MP-based aAPCs and co-released without interfering with each other. The targeted, sustained co-release of IL-2 and anti-CTLA-4 achieved markedly enhanced, synergistic effects in activating and expanding tumor antigen-specific T cells both in vitro and in vivo, as well as in inhibiting tumor growth in a mouse melanoma model, as compared with conventional two-signal aAPCs and IL-2 or anti-CTLA-4 single-released aAPCs. These data revealed the feasibility and importance of the paracrine release of multiple costimulatory molecules and cytokines from biodegradable aAPCs and thus provide a proof of principle for the future use of polymeric aAPCs for active immunotherapy of tumors and infectious diseases.

Impairment of the humoral and CD4+ T cell responses in HTLV-1-infected individuals immunized with tetanus toxoid

PubMed Central

Souza, Anselmo; Santos, Silvane; Carvalho, Lucas P.; Grassi, Maria Fernanda R.; Carvalho, Edgar M.

2016-01-01

T cells from HTLV-1-infected individuals have a decreased ability to proliferate after stimulation with recall antigens. This abnormality may be due to the production of regulatory cytokine or a dysfunctional antigen presentation. The aims of this study were to evaluate the antibody production and cytokine expression by lymphocytes before and after immunization with tetanus toxoid (TT) and to evaluate the immune response of monocytes after stimulation with TT and frequency of dendritic cells (DC) subsets. HTLV-1 carriers (HC) and uninfected controls with negative serology for TT were immunized with TT, and the antibody titers were determined by ELISA as well as the cell activation markers expression by monocytes. The frequencies of DC subsets were determined by flow cytometry. Following immunization, the IgG anti-TT titers and the frequency of CD4+ T cells expressing IFN-γ, TNF and IL-10 in response to TT were lower in the (HC) than in the controls. Additionally, monocytes from HC did not exhibit increased HLA-DR expression after stimulation with TT, and presented low numbers of DC subsets, therefore, it’s necessary to perform functional studies with antigen-presenting cells. Collectively, our finding suggests that HC present an impairment of the humoral and CD4+ T cell immune responses after vaccination. PMID:27282836

Antigen-Conjugated Human IgE Induces Antigen-Specific T Cell Tolerance in a Humanized Mouse Model

PubMed Central

Baravalle, Günther; Greer, Alexandra M.; LaFlam, Taylor N.; Shin, Jeoung-Sook

2015-01-01

Dendritic cells (DCs) play an important role in immune homeostasis through their ability to present Ags at steady state and mediate T cell tolerance. This characteristic renders DCs an attractive therapeutic target for the induction of tolerance against auto-antigens or allergens. Accordingly, Ag-conjugated DC–specific Abs have been proposed to be an excellent vehicle to deliver Ags to DCs for presentation and tolerance induction. However, this approach requires laborious reagent generation procedures and entails unpredictable side effects resulting from Ab-induced crosslinking of DC surface molecules. In this study, we examined whether IgE, a high-affinity, non–cross-linking natural ligand of FcεRI, could be used to target Ags to DCs and to induce Ag-specific T cell tolerance. We found that Ag-conjugated human IgE Fc domain (Fcε) effectively delivered Ags to DCs and enhanced Ag presentation by 1000- to 2500-fold in human FcεRIα-transgenic mice. Importantly, this presentation resulted in a systemic deletion of Ag-specific T cells and prevented these mice from developing delayed-type hypersensitivity, which is critically dependent on Ag-specific T cell immunity. Thus, targeting FcεRI on DCs via Ag-Fcε fusion protein may serve an alternative method to induce Ag-specific T cell tolerance in humans. PMID:24610015

Mannose-pepstatin conjugates as targeted inhibitors of antigen processing.

PubMed

Free, Paul; Hurley, Christopher A; Kageyama, Takashi; Chain, Benjamin M; Tabor, Alethea B

2006-05-07

The molecular details of antigen processing, including the identity of the enzymes involved, their intracellular location and their substrate specificity, are still incompletely understood. Selective inhibition of proteolytic antigen processing enzymes such as cathepsins D and E, using small molecular inhibitors such as pepstatin, has proven to be a valuable tool in investigating these pathways. However, pepstatin is poorly soluble in water and has limited access to the antigen processing compartment in antigen presenting cells. We have synthesised mannose-pepstatin conjugates, and neomannosylated BSA-pepstatin conjugates, as tools for the in vivo study of the antigen processing pathway. Conjugation to mannose and to neomannosylated BSA substantially improved the solubility of the conjugates relative to pepstatin. The mannose-pepstatin conjugates showed no reduction in inhibition of cathepsin E, whereas the neomannosylated BSA-pepstatin conjugates showed some loss of inhibition, probably due to steric factors. However, a neomannosylated BSA-pepstatin conjugate incorporating a cleavable disulfide linkage between the pepstatin and the BSA showed the best uptake to dendritic cells and the best inhibition of antigen processing.

Exploration of graphene oxide as an intelligent platform for cancer vaccines

NASA Astrophysics Data System (ADS)

Yue, Hua; Wei, Wei; Gu, Zonglin; Ni, Dezhi; Luo, Nana; Yang, Zaixing; Zhao, Lin; Garate, Jose Antonio; Zhou, Ruhong; Su, Zhiguo; Ma, Guanghui

2015-11-01

We explored an intelligent vaccine system via facile approaches using both experimental and theoretical techniques based on the two-dimensional graphene oxide (GO). Without extra addition of bio/chemical stimulators, the microsized GO imparted various immune activation tactics to improve the antigen immunogenicity. A high antigen adsorption was acquired, and the mechanism was revealed to be a combination of electrostatic, hydrophobic, and π-π stacking interactions. The ``folding GO'' acted as a cytokine self-producer and antigen reservoir and showed a particular autophagy, which efficiently promoted the activation of antigen presenting cells (APCs) and subsequent antigen cross-presentation. Such a ``One but All'' modality thus induced a high level of anti-tumor responses in a programmable way and resulted in efficient tumor regression in vivo. This work may shed light on the potential use of a new dimensional nano-platform in the development of high-performance cancer vaccines.We explored an intelligent vaccine system via facile approaches using both experimental and theoretical techniques based on the two-dimensional graphene oxide (GO). Without extra addition of bio/chemical stimulators, the microsized GO imparted various immune activation tactics to improve the antigen immunogenicity. A high antigen adsorption was acquired, and the mechanism was revealed to be a combination of electrostatic, hydrophobic, and π-π stacking interactions. The ``folding GO'' acted as a cytokine self-producer and antigen reservoir and showed a particular autophagy, which efficiently promoted the activation of antigen presenting cells (APCs) and subsequent antigen cross-presentation. Such a ``One but All'' modality thus induced a high level of anti-tumor responses in a programmable way and resulted in efficient tumor regression in vivo. This work may shed light on the potential use of a new dimensional nano-platform in the development of high-performance cancer vaccines. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr04986e

Potentiation of pH-sensitive polymer-modified liposomes with cationic lipid inclusion as antigen delivery carriers for cancer immunotherapy.

PubMed

Yoshizaki, Yuta; Yuba, Eiji; Sakaguchi, Naoki; Koiwai, Kazunori; Harada, Atsushi; Kono, Kenji

2014-09-01

Cationic lipid-incorporated liposomes modified with pH-sensitive polymers were prepared by introducing 3, 5-didodecyloxybenzamidine as a cationic lipid to egg yolk phosphatidylcholine liposomes modified with 3-methylglutarylated hyperbranched poly(glycidol) (MGlu-HPG) as a pH-sensitive polymer. These liposomes were stable at neutral pH, but were destabilized below pH 6.0 because MGlu-HPG changed its characteristics from hydrophilic to hydrophobic in response to the pH decrease. Cationic lipid inclusion improved their pH sensitivity at weakly acidic pH and association of liposomes with murine dendritic cell (DC) lines. Cationic lipid-incorporated liposomes delivered entrapped ovalbumin (OVA) molecules not only to cytosol but also to endosome/lysosome. Treatment with cationic lipid-incorporated liposomes induced up-regulation of antigen presentation-involved molecules on DCs, the promotion of cytokine production, and antigen presentation via both major histocompatibility complex (MHC) class I and II molecules. Especially, antigen presentation via MHC class II was promoted by cationic lipid inclusion, which might correspond to efficient endosome/lysosome delivery of OVA. Subcutaneous administration of OVA-loaded cationic lipid-incorporated liposomes induced antigen-specific antibody production in serum and Th1-dominant immune responses in the spleen. Furthermore, administration of the cationic lipid-incorporated liposomes to mice bearing E.G7-OVA tumor more significantly reduced the tumor volume than liposomes without cationic lipids. Therefore, cationic lipid inclusion into pH-sensitive polymer-modified liposomes, which can achieve both efficient antigen intracellular delivery and activation of antigen presenting cell, is an effective approach to develop antigen carriers for efficient cancer immunotherapy. Copyright © 2014 Elsevier Ltd. All rights reserved.

Enhanced Vaccine-Induced CD8+ T Cell Responses to Malaria Antigen ME-TRAP by Fusion to MHC Class II Invariant Chain

PubMed Central

Spencer, Alexandra J.; Cottingham, Matthew G.; Jenks, Jennifer A.; Longley, Rhea J.; Capone, Stefania; Colloca, Stefano; Folgori, Antonella; Cortese, Riccardo; Nicosia, Alfredo; Bregu, Migena; Hill, Adrian V. S.

2014-01-01

The orthodox role of the invariant chain (CD74; Ii) is in antigen presentation to CD4+ T cells, but enhanced CD8+ T cells responses have been reported after vaccination with vectored viral vaccines encoding a fusion of Ii to the antigen of interest. In this study we assessed whether fusion of the malarial antigen, ME-TRAP, to Ii could increase the vaccine-induced CD8+ T cell response. Following single or heterologous prime-boost vaccination of mice with a recombinant chimpanzee adenovirus vector, ChAd63, or recombinant modified vaccinia virus Ankara (MVA), higher frequencies of antigen-specific CD4+ and CD8+ T cells were observed, with the largest increases observed following a ChAd63-MVA heterologous prime-boost regimen. Studies in non-human primates confirmed the ability of Ii-fusion to augment the T cell response, where a 4-fold increase was maintained up to 11 weeks after the MVA boost. Of the numerous different approaches explored to increase vectored vaccine induced immunogenicity over the years, fusion to the invariant chain showed a consistent enhancement in CD8+ T cell responses across different animal species and may therefore find application in the development of vaccines against human malaria and other diseases where high levels of cell-mediated immunity are required. PMID:24945248

Expanding and characterizing esophageal epithelial cells obtained from children with eosinophilic esophagitis.

PubMed

Sayej, Wael N; Foster, Christopher; Jensen, Todd; Chatfield, Sydney; Finck, Christine

2018-06-12

The role of epithelial cells in eosinophilic esophagitis (EoE) is not well understood. In this study, our aim was to isolate, culture, and expand esophageal epithelial cells obtained from patients with or without EoE and characterize differences observed over time in culture. Biopsies were obtained at the time of endoscopy from children with EoE or suspected to have EoE. We established patient-derived esophageal epithelial cell (PDEEC) lines utilizing conditional reprogramming methods. We determined integrin profiles, gene expression, MHC class II expression, and reactivity to antigen stimulation. The PDEECs were found to maintain their phenotype over several passages. There were differences in integrin profiles and gene expression levels in EoE-Active compared to normal controls and EoE-Remission patients. Once stimulated with antigens, PDEECs express MHC class II molecules on their surface, and when co-cultured with autologous T-cells, there is increased IL-6 and TNF-α secretion in EoE-Active patients vs. controls. We are able to isolate, culture, and expand esophageal epithelial cells from pediatric patients with and without EoE. Once stimulated with antigens, these cells express MHC class II molecules and behave as non-professional antigen-presenting cells. This method will help us in developing an ex vivo, individualized, patient-specific model for diagnostic testing for causative antigens.

Species-Specific Elements in the Large T-Antigen J Domain Are Required for Cellular Transformation and DNA Replication by Simian Virus 40

PubMed Central

Sullivan, Christopher S.; Tremblay, James D.; Fewell, Sheara W.; Lewis, John A.; Brodsky, Jeffrey L.; Pipas, James M.

2000-01-01

The J domain of simian virus 40 (SV40) large T antigen is required for efficient DNA replication and transformation. Despite previous reports demonstrating the promiscuity of J domains in heterologous systems, results presented here show the requirement for specific J-domain sequences in SV40 large-T-antigen-mediated activities. In particular, chimeric-T-antigen constructs in which the SV40 T-antigen J domain was replaced with that from the yeast Ydj1p or Escherichia coli DnaJ proteins failed to replicate in BSC40 cells and did not transform REF52 cells. However, T antigen containing the JC virus J domain was functional in these assays, although it was less efficient than the wild type. The inability of some large-T-antigen chimeras to promote DNA replication and elicit cellular transformation was not due to a failure to interact with hsc70, since a nonfunctional chimera, containing the DnaJ J domain, bound hsc70. However, this nonfunctional chimeric T antigen was reduced in its ability to stimulate hsc70 ATPase activity and unable to liberate E2F from p130, indicating that transcriptional activation of factors required for cell growth and DNA replication may be compromised. Our data suggest that the T-antigen J domain harbors species-specific elements required for viral activities in vivo. PMID:10891510

Toxicological and pharmacological assessment of AGEN1884, a novel human IgG1 anti-CTLA-4 antibody

PubMed Central

Gonzalez, Ana; Manrique, Mariana; Chand, Dhan; Savitsky, David; Morin, Benjamin; Breous-Nystrom, Ekaterina; Dupont, Christopher; Ward, Rebecca A.; Mundt, Cornelia; Duckless, Benjamin; Tang, Hao; Findeis, Mark A.; Schuster, Andrea; Waight, Jeremy D.; Underwood, Dennis; Clarke, Christopher; Ritter, Gerd; Merghoub, Taha; Schaer, David; Wolchok, Jedd D.; van Dijk, Marc; Buell, Jennifer S.; Cuillerot, Jean-Marie; Stein, Robert; Drouin, Elise E.

2018-01-01

CTLA-4 and CD28 exemplify a co-inhibitory and co-stimulatory signaling axis that dynamically sculpts the interaction of antigen-specific T cells with antigen-presenting cells. Anti-CTLA-4 antibodies enhance tumor-specific immunity through a variety of mechanisms including: blockade of CD80 or CD86 binding to CTLA-4, repressing regulatory T cell function and selective elimination of intratumoral regulatory T cells via an Fcγ receptor-dependent mechanism. AGEN1884 is a novel IgG1 antibody targeting CTLA-4. It potently enhanced antigen-specific T cell responsiveness that could be potentiated in combination with other immunomodulatory antibodies. AGEN1884 was well-tolerated in non-human primates and enhanced vaccine-mediated antigen-specific immunity. AGEN1884 combined effectively with PD-1 blockade to elicit a T cell proliferative response in the periphery. Interestingly, an IgG2 variant of AGEN1884 revealed distinct functional differences that may have implications for optimal dosing regimens in patients. Taken together, the pharmacological properties of AGEN1884 support its clinical investigation as a single therapeutic and combination agent. PMID:29617360

Linking innate to adaptive immunity through dendritic cells.

PubMed

Steinman, Ralph M

2006-01-01

The function of dendritic cells (DCs) in linking innate to adaptive immunity is often summarized with two terms. DCs are sentinels, able to capture, process and present antigens and to migrate to lymphoid tissues to select rare, antigen-reactive T cell clones. DCs are also sensors, responding to a spectrum of environmental cues by extensive differentiation or maturation. The type of DC and the type of maturation induced by different stimuli influences the immunological outcome, such as the differentiation of Thl vs. Th2 T cells. Here we summarize the contributions of DCs to innate defences, particularly the production of immune enhancing cytokines and the activation of innate lymphocytes. Then we outline three innate features of DCs that influence peripheral tolerance and lead to adaptive immunity: a specialized endocytic system for antigen capture and processing, location and movements in vivo, and maturation in response to an array of stimuli. A new approach to the analysis of DC biology is to target antigens selectively to maturing DCs in vivo. This leads to stronger, more prolonged and broader (many immunogenic peptides) immunity by both T cells and B cells.

CD8+ T cell recognition of an endogenously processed epitope is regulated primarily by residues within the epitope

PubMed Central

1992-01-01

Cytotoxic T lymphocytes (CTL) recognize short antigenic peptides associated with cell surface class I major histocompatibility complex (MHC) molecules. This association presumably occurs between newly synthesized class I MHC molecules and peptide fragments in a pre-Golgi compartment. Little is known about the factors that regulate the formation of these antigenic peptide fragments within the cell. To examine the role of residues within a core epitope and in the flanking sequences for the generation and presentation of the newly synthesized peptide fragment recognized by CD8+ CTL, we have mutagenized the coding sequence for the CTL epitope spanning residues 202-221 in the influenza A/Japan/57 hemagglutinin (HA). In this study over 60 substitution mutations in the epitope were tested for their effects on target cell sensitization using a cytoplasmic viral expression system. The HA202- 221 site contains two overlapping subsites defined by CTL clones 11-1 and 40-2. Mutations in HA residues 204-213 or residues 210-219 often abolished target cell lysis by CTL clones 11-1 and 40-2, respectively. Although residues outside the core epitope did not usually affect the ability to be lysed by CTL clones, substitution of a Gly residue for Val-214 abolished lysis by clone 11-1. These data suggest that residues within a site that affect MHC binding and T cell receptor recognition appear to play the predominant role in dictating the formation of the antigenic complex recognized by CD8+ CTL, and therefore the antigenicity of the protein antigen presented to CD8+ T cells. Most alterations in residues flanking the endogenously expressed epitope do not appreciably affect the generation and recognition of the site. PMID:1383384

Origin and Role of a Subset of Tumor-Associated Neutrophils with Antigen-Presenting Cell Features in Early-Stage Human Lung Cancer.

PubMed

Singhal, Sunil; Bhojnagarwala, Pratik S; O'Brien, Shaun; Moon, Edmund K; Garfall, Alfred L; Rao, Abhishek S; Quatromoni, Jon G; Stephen, Tom Li; Litzky, Leslie; Deshpande, Charuhas; Feldman, Michael D; Hancock, Wayne W; Conejo-Garcia, Jose R; Albelda, Steven M; Eruslanov, Evgeniy B

2016-07-11

Based on studies in mouse tumor models, granulocytes appear to play a tumor-promoting role. However, there are limited data about the phenotype and function of tumor-associated neutrophils (TANs) in humans. Here, we identify a subset of TANs that exhibited characteristics of both neutrophils and antigen-presenting cells (APCs) in early-stage human lung cancer. These APC-like "hybrid neutrophils," which originate from CD11b(+)CD15(hi)CD10(-)CD16(low) immature progenitors, are able to cross-present antigens, as well as trigger and augment anti-tumor T cell responses. Interferon-γ and granulocyte-macrophage colony-stimulating factor are requisite factors in the tumor that, working through the Ikaros transcription factor, synergistically exert their APC-promoting effects on the progenitors. Overall, these data demonstrate the existence of a specialized TAN subset with anti-tumor capabilities in human cancer. Copyright © 2016 Elsevier Inc. All rights reserved.

Essential differences in ligand presentation and T cell epitope recognition among HLA molecules of the HLA-B44 supertype.

PubMed

Hillen, Nina; Mester, Gabor; Lemmel, Claudia; Weinzierl, Andreas O; Müller, Margret; Wernet, Dorothee; Hennenlotter, Jörg; Stenzl, Arnulf; Rammensee, Hans-Georg; Stevanović, Stefan

2008-11-01

Human leukocyte antigens (HLA) have long been grouped into supertypes to facilitate peptide-based immunotherapy. Analysis of several hundreds of peptides presented by all nine antigens of the HLA-B44 supertype (HLA-B*18, B*37, B*40, B*41, B*44, B*45, B*47, B*49 and B*50) revealed unique peptide motifs for each of them. Taking all supertype members into consideration only 25 out of 670 natural ligands were found on more than one HLA molecule. Further direct comparisons by two mass spectrometric methods--isotope labeling as well as a label-free approach--consistently demonstrated only minute overlaps of below 3% between the ligandomes of different HLA antigens. In addition, T cell reactions of healthy donors against immunodominant HLA-B*44 and HLA-B*40 epitopes from EBV lacked promiscuous T-cell recognition within the HLA-B44 supertype. Taken together, these results challenge the common paradigm of broadly presented epitopes within this supertype.

Mismatch in epitope specificities between IFNγ inflamed and uninflamed conditions leads to escape from T lymphocyte killing in melanoma.

PubMed

Woods, Katherine; Knights, Ashley J; Anaka, Matthew; Schittenhelm, Ralf B; Purcell, Anthony W; Behren, Andreas; Cebon, Jonathan

2016-01-01

A current focus in cancer treatment is to broaden responses to immunotherapy. One reason these therapies may prove inadequate is that T lymphocytes fail to recognize the tumor due to differences in immunogenic epitopes presented by the cancer cells under inflammatory or non-inflammatory conditions. The antigen processing machinery of the cell, the proteasome, cleaves proteins into peptide epitopes for presentation on MHC complexes. Immunoproteasomes in inflammatory melanomas, and in antigen presenting cells of the immune system, are enzymatically different to standard proteasomes expressed by tumors with no inflammation. This corresponds to alterations in protein cleavage between proteasome subtypes, and a disparate repertoire of MHC-presented epitopes. We assessed steady state and IFNγ-induced immunoproteasome expression in melanoma cells. Using epitope specific T-lymphocyte clones, we studied processing and presentation of three NY-ESO-1 HLA-Cw3 restricted epitopes by melanoma cell lines. Our experimental model allowed comparison of the processing of three distinct epitopes from a single antigen presented on the same HLA complex. We further investigated processing of these epitopes by direct inhibition, or siRNA mediated knockdown, of the immunoproteasome catalytic subunit LMP7. Our data demonstrated a profound difference in the way in which immunogenic T-lymphocyte epitopes are presented by melanoma cells under IFNγ inflammatory versus non-inflammatory conditions. These alterations led to significant changes in the ability of T-lymphocytes to recognize and target melanoma cells. Our results illustrate a little-studied mechanism of immune escape by tumor cells which, with appropriate understanding and treatment, may be reversible. These data have implications for the design of cancer vaccines and adoptive T cell therapies.

[Detection of antigen structures in blood cells in various prepared plasma transfusions].

PubMed

Barz, D

1994-01-01

We investigated the content of antigen-bearing cells and cell fragments in Fresh Frozen Plasma (FFP) from blood centers, in Octaplas (virus-inactivated fresh plasma produced with the solvent/detergent technique by the Octapharma Company) and in MB-plasma (virus-inactivated fresh plasma after photodynamic treatment with methylen blue coming from the German Red Cross in Springe, Lower Saxony). With the aid of an immunoassay (MAIPA-test) these plasmas were tested regarding Rhesus-D-antigen, HLA-class-I- and HLA-class-II-antigens, platelet specific antigens HPA-1a/HPA-1b and granulocyte specific antigens NA1/NA2. In Octaplas (n = 10) we did not find cells or cell fragments and no antigen-bearing blood cell structures. In FFP (n = 28) there were platelet specific antigens in 27 cases (96.4%) and HLA-class-I-antigens in 4 cases (14.3%). In MB-plasma (n = 14) we found platelet specific antigens in all cases, HLA-class-I-antigens in 4 cases (18.6%), HLA-class-II-antigens and granulocyte specific antigens in 1 case (7.1%) and Rhesus-D-antigen in 3 cases (21.4%). Plasma derived from whole blood showed lower levels of cells and antigens than plasma which was produced with the aid of the cell separator.

Antigen-specific, CD4+CD25+ regulatory T cell clones induced in Peyer's patches.

PubMed

Tsuji, Noriko M; Mizumachi, Koko; Kurisaki, Jun-Ichi

2003-04-01

Since intestine is exposed to numerous exogenous antigens such as food and commensal bacteria, the organ bears efficient mechanisms for establishment of tolerance and induction of regulatory T cells (T(reg)). Intestinal and inducible T(reg) include T(r)1-like and T(h)3 cells whose major effector molecules are IL-10 and transforming growth factor (TGF)-beta. These antigen-specific T(reg) are expected to become clinical targets to modify the inflammatory immune response associated with allergy, autoimmune diseases and transplantation. In the present study, we characterized the antigen-specific T(reg) induced in the intestine by orally administering high-dose beta-lactoglobulin (BLG) to BALB/c mice. Seven days after feeding, only Peyer's patch (PP) cells among different organs exerted significant suppressive effect on antibody production upon in vitro BLG stimulation. This suppressive effect was also prominent in six BLG-specific CD4(+) T cell clones (OPP1-6) established from PP from mice orally administered with high doses of BLG and was partially reversed by antibodies to TGF-beta. Intravenous transfer of OPP2 efficiently suppressed BLG-specific IgG1 production in serum following immunization, indicating the role of such T(reg) in the systemic tolerance after oral administration of antigen (oral tolerance). OPP clones secrete TGF-beta, IFN-gamma and low levels of IL-10, a cytokine pattern similar to that secreted by anergic T cells. OPP clones bear a CD4(+)CD25(+) phenotype and show significantly lower proliferative response compared to T(h)0 clones. This lower response is recovered by the addition of IL-2. Thus, antigen-specific CD4(+)CD25(+) T(reg), which have characteristics of anergic cells and actively suppress antibody production are induced in PP upon oral administration of protein antigen.

Integrin-mediated interactions between B cells and follicular dendritic cells influence germinal center B cell fitness1

PubMed Central

Wang, Xiaoming; Rodda, Lauren; Bannard, Oliver; Cyster, Jason G.

2014-01-01

Integrin-ligand interactions between germinal center (GC) B cells and antigen-presenting follicular dendritic cells (FDCs) have been suggested to play central roles during GC responses but their in vivo requirement has not been directly tested. Here we show that while integrins αLβ2 and α4β1 are highly expressed and functional on mouse GC B cells, removal of single integrins or their ligands had little effect on B cell participation in the GC response. Combined β2-integrin deficiency and α4-integrin blockade also did not affect the GC response against a particulate antigen. However, the combined integrin deficiency did cause B cells to be outcompeted in splenic GC responses against a soluble protein antigen and in mesenteric lymph node GC responses against gut-derived antigens. Similar findings were made for β2-deficient B cells in mice lacking VCAM1 on FDCs. The reduced fitness of the GC B cells did not appear to be due to decreased antigen acquisition, proliferation rates or pAKT levels. In summary, our findings provide evidence that αLβ2 and α4β1 play overlapping and context-dependent roles in supporting interactions with FDCs that can augment the fitness of responding GC B cells. We also find that mouse GC B cells upregulate αvβ3 and adhere to vitronectin and milk fat globule EGF-factor-8 protein. Integrin β3-deficient B cells contributed in a slightly exaggerated manner to GC responses suggesting this integrin has a regulatory function in GC B cells. PMID:24740506

Dendritic cells produce eicosanoids, which modulate generation and functions of antigen-presenting cells.

PubMed

Harizi, H; Gualde, N

2002-01-01

Eicosanoids have been shown to be potent immunoregulatory arachidonic acid (AA) metabolites. AA is the precursor of prostaglandin E(2) (PGE(2)) and leukotriene B(4) (LTB(4)) which are able to modulate both inflammation and the immune response. Dendritic cells process and present antigens to T lymphocytes. They are highly specialized antigen-presenting cells (APC) and usually considered as 'professional APC'. In the present paper, we report some data on the biosynthetic capacity of murine APC from the bone marrow (BM-DCs) to produce AA metabolites. Using an ELISA we have observed that BM-DCs spontaneously produce both PGE(2) and LTB(4) whose production increased in response to bacterial lipopolysaccharides (LPS). In addition we found that LTB(4) production was twice as high when both COX pathways were blocked with selective COX-inhibitors. We have also investigated the effect of PGE(2) and LTB(4) on the in vitro generation of the so-called BM-DCs. Exogenous PGE(2) and LTB(4) added to bone marrow cultures inhibit and promote, respectively, BM-DC generation. PGE(2) added to the maturing BM-DCs reduces their MHC class-II expression.

Differential requirement for cathepsin D for processing of the full length and C-terminal fragment of the malaria antigen MSP1.

PubMed

Tulone, Calogero; Sponaas, Anne-Marit; Raiber, Eun-Ang; Tabor, Alethea B; Langhorne, Jean; Chain, Benny M

2011-01-01

Merozoite Surface Protein 1 is expressed on the surface of malaria merozoites and is important for invasion of the malaria parasite into erythrocytes. MSP1-specific CD4 T cell responses and antibody can confer protective immunity in experimental models of malaria. In this study we explore the contributions of cathepsins D and E, two aspartic proteinases previously implicated in antigen processing, to generating MSP1 CD4 T-cell epitopes for presentation. The absence of cathepsin D, a late endosome/lysosomal enzyme, is associated with a reduced presentation of MSP1 both following in vitro processing of the epitope MSP1 from infected erythrocytes by bone marrow-derived dendritic cells, and following in vivo processing by splenic CD11c+ dendritic cells. By contrast, processing and presentation of the soluble recombinant protein fragment of MSP1 is unaffected by the absence of cathepsin D, but is inhibited when both cathepsin D and E are absent. The role of different proteinases in generating the CD4 T cell repertoire, therefore, depends on the context in which an antigen is introduced to the immune system.

Differential Requirement for Cathepsin D for Processing of the Full Length and C-Terminal Fragment of the Malaria Antigen MSP1

PubMed Central

Raiber, Eun-Ang; Tabor, Alethea B.; Langhorne, Jean; Chain, Benny M.

2011-01-01

Merozoite Surface Protein 1 is expressed on the surface of malaria merozoites and is important for invasion of the malaria parasite into erythrocytes. MSP1-specific CD4 T cell responses and antibody can confer protective immunity in experimental models of malaria. In this study we explore the contributions of cathepsins D and E, two aspartic proteinases previously implicated in antigen processing, to generating MSP1 CD4 T-cell epitopes for presentation. The absence of cathepsin D, a late endosome/lysosomal enzyme, is associated with a reduced presentation of MSP1 both following in vitro processing of the epitope MSP1 from infected erythrocytes by bone marrow-derived dendritic cells, and following in vivo processing by splenic CD11c+ dendritic cells. By contrast, processing and presentation of the soluble recombinant protein fragment of MSP1 is unaffected by the absence of cathepsin D, but is inhibited when both cathepsin D and E are absent. The role of different proteinases in generating the CD4 T cell repertoire, therefore, depends on the context in which an antigen is introduced to the immune system. PMID:22053177

The nanoscale spatial organization of B-cell receptors on immunoglobulin M- and G-expressing human B-cells.

PubMed

Lee, Jinmin; Sengupta, Prabuddha; Brzostowski, Joseph; Lippincott-Schwartz, Jennifer; Pierce, Susan K

2017-02-15

B-cell activation is initiated by the binding of antigen to the B-cell receptor (BCR). Here we used dSTORM superresolution imaging to characterize the nanoscale spatial organization of immunoglobulin M (IgM) and IgG BCRs on the surfaces of resting and antigen--activated human peripheral blood B-cells. We provide insights into both the fundamental process of antigen-driven BCR clustering and differences in the spatial organization of IgM and IgG BCRs that may contribute to the characteristic differences in the responses of naive and memory B-cells to antigen. We provide evidence that although both IgM and IgG BCRs reside in highly heterogeneous protein islands that vary in size and number of BCR single-molecule localizations, both resting and activated B-cells intrinsically maintain a high -frequency of single isolated BCR localizations, which likely represent BCR monomers. IgG BCRs are more clustered than IgM BCRs on resting cells and form larger protein islands after antigen activation. Small, dense BCR clusters likely formed via protein-protein interactions are present on the surface of resting cells, and antigen activation induces these to come together to form less dense, larger islands, a process likely governed, at least in part, by protein-lipid interactions. © 2017 Lee, Sengupta, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Alterations in dendritic cell function in aged mice: potential implications for immunotherapy design.

PubMed

Paula, Carine; Motta, Adriana; Schmitz, Carla; Nunes, Claudia P; Souza, Ana Paula; Bonorino, Cristina

2009-02-01

It is known that immune system functions decrease with age, and that adaptive immune responses, especially CD4+ T cell function, seem to be the main affected point in immunity with aging. Dendritic cells (DC) are the major antigen presenting cell (APC), and at least part of the defects observed in adaptive immunity of aged individuals could be due to diminished potential of bone marrow to generate new DC, or defects in DC function. In this study, we investigated if the ability of aged bone marrow (BM) to generate new DC in vitro, as well as aged BM-derived DC responses to lypopolysaccharide (LPS). Because DC are important tools in newly developing anti-tumor therapies, we also studied the ability of aged DC to phagocytose and present antigen from necrotic tumor cells. We found that aged BM generated fewer DC in vitro compared to young BM. While LPS-induced DC maturation is reduced in DC of aged mice, a high TNF-alpha production is observed in aged DC even without LPS stimulation. While phagocytosis of tumor cells is not affected by age, and DC derived from aged BM show a higher TNF-alpha production in response to phagocytosis, presentation of tumor antigens was decreased in aged DC. Because class II upregulation in response to phagocytosis was similar between aged and young DC, this could indicate an age associated processing defect in the exogenous pathway. These findings suggest that age of BM used to generate DC does not impair their phagocytic ability or TNF-alpha production, however leads to a decreased yield in mature DC, reduced response to LPS, and diminished antigen processing/presentation potential. Our results are relevant to optimization DC-based vaccine design for aged populations.

Spirochetal antigens and lymphoid cell surface markers in Lyme synovitis. Comparison with rheumatoid synovium and tonsillar lymphoid tissue.

PubMed

Steere, A C; Duray, P H; Butcher, E C

1988-04-01

Using monoclonal antibodies to spirochetal antigenes and lymphoid cell surface markers, we examined the synovial lesions of 12 patients with Lyme disease, and compared them with rheumatoid synovium and tonsillar lymphoid tissue. The synovial lesions of Lyme disease patients and rheumatoid arthritis patients were similar and often consisted of the elements found in normal organized lymphoid tissue. In both diseases, T cells, predominantly of the helper/inducer subset, were distributed diffusely in subsynovial lining areas, often with nodular aggregates of tightly intermixed T and B cells. IgD-bearing B cells were scattered within the aggregates, and a few follicular dendritic cells and activated germinal center B cells were sometimes present. Outside the aggregates, many plasma cells, high endothelial venules, scattered macrophages, and a few dendritic macrophages were found. HLA-DR and DQ expression was intense throughout the lesions. In 6 of the 12 patients with Lyme arthritis, but in none of those with rheumatoid arthritis, a few spirochetes and globular antigen deposits were seen in and around blood vessels in areas of lymphocytic infiltration. Thus, in Lyme arthritis, a small number of spirochetes are probably the antigenic stimulus for chronic synovial inflammation.

Urushiol (poison ivy)-triggered suppressor T cell clone generated from peripheral blood.

PubMed Central

Kalish, R S; Morimoto, C

1988-01-01

Allergic contact dermatitis to Toxicodendron radicans (poison ivy) is mediated by the hapten urushiol. An urushiol-specific, interleukin 2 (IL-2)-dependent T cell clone (RLB9-7) was generated from the peripheral blood of a patient with a history of allergic contact dermatitis to T. radicans. This clone proliferated specifically to both leaf extract and pure urushiol. Although the clone had the phenotype CD3+CD4+CD8+, proliferation to antigen was blocked by anti-CD8 and anti-HLA-A, B, C, but not by anti-CD4, suggesting that CD4 was not functionally associated with the T cell receptor. Furthermore, studies with antigen-presenting cells from MHC-typed donors indicated that the clone was MHC class 1 restricted. RLB9-7 was WT31 positive, indicating it bears the alpha beta T cell receptor. The clone lacked significant natural killer cell activity and produced only low levels of IL-2 or gamma-interferon upon antigen stimulation. Addition of RLB9-7 to autologous peripheral blood mononuclear cells in the presence of urushiol inhibited the pokeweed mitogen-driven IgG synthesis. This suppression was resistant to irradiation (2,000 rad) and was not seen when RLB9-7 was added to allogeneic cells, even in the presence of irradiated autologous antigen-presenting cells, suggesting that suppression was MHC restricted and not mediated by nonspecific soluble factors. However, RLB9-7 cells in the presence of urushiol inhibited the synthesis of tetanus toxoid-specific IgG by autologous lymphocytes, indicating that the suppression, although triggered specifically by urushiol, was nonspecific. PMID:2458387

Complete responses of relapsed lymphoma following genetic modification of tumor-antigen presenting cells and T-lymphocyte transfer.

PubMed

Bollard, Catherine M; Gottschalk, Stephen; Leen, Ann M; Weiss, Heidi; Straathof, Karin C; Carrum, George; Khalil, Mariam; Wu, Meng-fen; Huls, M Helen; Chang, Chung-Che; Gresik, M Victoria; Gee, Adrian P; Brenner, Malcolm K; Rooney, Cliona M; Heslop, Helen E

2007-10-15

Epstein-Barr virus (EBV)-associated tumors developing in immunocompetent individuals present a challenge to immunotherapy, since they lack expression of immunodominant viral antigens. However, the tumors consistently express viral proteins including LMP2, which are immunologically "weak" but may nonetheless be targets for immune T cells. We previously showed that a majority of cytotoxic T lymphocytes (CTLs) reactivated using EBV-transformed B-lymphoblastoid cells lines (LCLs) contained minor populations of LMP2-specific T cells and homed to tumor sites. However, they did not produce remissions in patients with bulky disease. We have now used gene transfer into antigen-presenting cells (APCs) to augment the expression and immunogenicity of LMP2. These modified APCs increased the frequency of LMP2-specific CTLs by up to 100-fold compared with unmodified LCL-APCs. The LMP2-specific population expanded and persisted in vivo without adverse effects. Nine of 10 patients treated in remission of high-risk disease remain in remission, and 5 of 6 patients with active relapsed disease had a tumor response, which was complete in 4 and sustained for more than 9 months. It is therefore possible to generate immune responses to weak tumor antigens by ex vivo genetic modification of APCs and the CTLs so produced can have substantial antitumor activity. This study is registered at http://www.cancer.gov/clinicaltrials (protocol IDs: BCM-H-9936, NCT00062868, NCT00070226).

Expansion of natural (NK1+) T cells that express alpha beta T cell receptors in transporters associated with antigen presentation-1 null and thymus leukemia antigen positive mice

PubMed Central

1996-01-01

Thymic selection of natural killer-1+ natural T cells that express alpha beta T cell receptors requires a conserved beta 2-microglobulin- associated molecule, presumably CD1d, displayed by CD4+8+ thymocytes. Here we demonstrate that positive selection of natural T cells occurs independent of transporters associated with antigen presentation-1 (TAP- 1) function. Moreover, natural T cells in TAP-1o/o mice are numerically expanded. Several H-2 class Ib molecules function in a TAP-independent manner, suggesting that if expressed in TAP-1o/o thymocytes, they could play a role in natural T cell development. Of these class Ib molecules, H-2TL is expressed by TAP-1o/o thymocytes. Moreover, we find that thymi of TL+ mice congenic or transgenic for H-2T18 also have a numerically expanded natural T cell repertoire compared with TL- mice. This expansion, as in TAP-1o/o thymi, is evident in each of the limited T cell receptor V beta chains expressed by natural T cells, suggesting that TL and CD1d impact similar repertoires. Thus TL, in addition to CD1d, plays a role in natural T cell development. PMID:8879233

Saponin-based adjuvants induce cross-presentation in dendritic cells by intracellular lipid body formation

PubMed Central

den Brok, Martijn H.; Büll, Christian; Wassink, Melissa; de Graaf, Annemarie M.; Wagenaars, Jori A.; Minderman, Marthe; Thakur, Mayank; Amigorena, Sebastian; Rijke, Eric O.; Schrier, Carla C.; Adema, Gosse J.

2016-01-01

Saponin-based adjuvants (SBAs) are being used in animal and human (cancer) vaccines, as they induce protective cellular immunity. Their adjuvant potency is a factor of inflammasome activation and enhanced antigen cross-presentation by dendritic cells (DCs), but how antigen cross-presentation is induced is not clear. Here we show that SBAs uniquely induce intracellular lipid bodies (LBs) in the CD11b+ DC subset in vitro and in vivo. Using genetic and pharmacological interference in models for vaccination and in situ tumour ablation, we demonstrate that LB induction is causally related to the saponin-dependent increase in cross-presentation and T-cell activation. These findings link adjuvant activity to LB formation, aid the application of SBAs as a cancer vaccine component, and will stimulate development of new adjuvants enhancing T-cell-mediated immunity. PMID:27819292

IT-25DEVELOPMENTALLY REGULATED ANTIGENS FOR IMMUNOLOGIC TARGETING OF MEDULLOBLASTOMA SUBTYPES

PubMed Central

Pham, Christina; Flores, Catherine; Pei, Yanxin; Wechsler-Reya, Robert; Mitchell, Duane

2014-01-01

INTRODUCTION: Medulloblastoma (MB) remains incurable in one third of patients despite aggressive multi-modality standard therapies. Immunotherapy presents a promising alternative by specifically targeting cancer cells. To date, there have been no successful immunologic applications targeting MB. Emerging evidence from integrated genomic studies has suggested MB variants arise from deregulation of pathways affecting proliferation of progenitor cell populations within the developing cerebellum. Using total embryonic RNA as a source of tumor rejection antigens is attractive because it can be delivered as a single vaccine, target both known and unknown fetal proteins, and can be refined to preferentially treat distinct MB subtypes. METHODS: We have created two transplantable, syngeneic animal MB models recapitulating human SHH and Group 3 variants to investigate the immunologic targeting of different MB subtypes. We generated T cells specific to the developing mouse cerebellum (P5) and tested their reactivity to target cells pulsed with total RNA from two MB subtypes and the normal brain. Immune responses were evaluated by measuring cytokine secretion following re-stimulation of activated T cells with both normal and tumor cell targets. In vivo antitumor efficacy was also tested in survival studies of intracranial tumor-bearing animals. RESULTS: We generated T cells specific to the developing cerebellum in vitro, confirming the immunogenicity of developmentally regulated antigens. Additionally, we have shown that developmental antigen-specific T cells produce high levels of Th1-type cytokines in response to tumor cells of two immunologically distinct subtypes of MB. Interestingly, developmental antigen specific T cells do not show cross reactivity with the normal brain or subsequent stages of the developing brain after P5. Targeting developmental antigens also conferred a significant survival benefit in a treatment model of Group 3 tumor bearing animals. CONCLUSIONS: Developmental antigens can safely target multiple MB subtypes with equal effectiveness compared to previously established total tumor strategies.

ImmunoChip Study Implicates Antigen Presentation to T Cells in Narcolepsy

PubMed Central

Kornum, Birgitte Rahbek; Kenny, Eimear E.; Trynka, Gosia; Einen, Mali; Rico, Tom J.; Lichtner, Peter; Dauvilliers, Yves; Arnulf, Isabelle; Lecendreux, Michel; Javidi, Sirous; Geisler, Peter; Mayer, Geert; Pizza, Fabio; Poli, Francesca; Plazzi, Giuseppe; Overeem, Sebastiaan; Lammers, Gert Jan; Kemlink, David; Sonka, Karel; Nevsimalova, Sona; Rouleau, Guy; Desautels, Alex; Montplaisir, Jacques; Frauscher, Birgit; Ehrmann, Laura; Högl, Birgit; Jennum, Poul; Bourgin, Patrice; Peraita-Adrados, Rosa; Iranzo, Alex; Bassetti, Claudio; Chen, Wei-Min; Concannon, Patrick; Thompson, Susan D.; Damotte, Vincent; Fontaine, Bertrand; Breban, Maxime; Gieger, Christian; Klopp, Norman; Deloukas, Panos; Wijmenga, Cisca; Hallmayer, Joachim; Onengut-Gumuscu, Suna; Rich, Stephen S.; Winkelmann, Juliane; Mignot, Emmanuel

2013-01-01

Recent advances in the identification of susceptibility genes and environmental exposures provide broad support for a post-infectious autoimmune basis for narcolepsy/hypocretin (orexin) deficiency. We genotyped loci associated with other autoimmune and inflammatory diseases in 1,886 individuals with hypocretin-deficient narcolepsy and 10,421 controls, all of European ancestry, using a custom genotyping array (ImmunoChip). Three loci located outside the Human Leukocyte Antigen (HLA) region on chromosome 6 were significantly associated with disease risk. In addition to a strong signal in the T cell receptor alpha (TRA@), variants in two additional narcolepsy loci, Cathepsin H (CTSH) and Tumor necrosis factor (ligand) superfamily member 4 (TNFSF4, also called OX40L), attained genome-wide significance. These findings underline the importance of antigen presentation by HLA Class II to T cells in the pathophysiology of this autoimmune disease. PMID:23459209

Prolactin, dendritic cells, and systemic lupus erythematosus.

PubMed

Jara, Luis J; Benitez, Gamaliel; Medina, Gabriela

2008-01-01

Dendritic cells (DC) play a central role in the induction of autoimmunity in T and B cells. DC express a high level of the major histocompatibility complex that interact with the receptors on T cells. Immature DC present antigens efficiently. Prolactin (PRL) participates in DC maturation. Systemic lupus erythematosus (SLE) is characterized by a loss of tolerance to self-antigens and persistent production of autoantibodies. Serum from SLE patients induces normal monocytes to differentiate into DC in correlation with disease activity depending on the actions of interferon-alpha, immune complexes, PRL, etc. High serum PRL levels have been found in a subset of SLE patients associated with active disease and organ involvement. It is possible that PRL interacts with DC, skewing its function from antigen presentation to a proinflammatory phenotype with high interferon-alpha production. Therefore, SLE is characterized by deficiency of DC functions and abnormal PRL secretion. The relationships between PRL and DC may have a role in the pathogenesis of SLE.

CD1c presentation of synthetic glycolipid antigens with foreign alkyl branching motifs

PubMed Central

de Jong, Annemieke; Arce, Eva Casas; Cheng, Tan-Yun; van Summeren, Ruben P.; Feringa, Ben L.; Dudkin, Vadim; Crich, David; Matsunaga, Isamu; Minnaard, Adriaan J.; Moody, D. Branch

2009-01-01

Summary Human CD1c is a protein that activates αβ T cells by presenting self antigens, synthetic mannosyl phosphodolichols and mycobacterial mannosyl phosphopolyketides. To determine which molecular structures of antigens mediate a T cell response, we measured activation by structurally divergent M. tuberculosis mannosyl-β1-phosphomycoketides as well as by synthetic analogs produced by two methods that yield either stereorandom or stereospecific methyl branching patterns. T cell responses required both a phosphate and a β-linked mannose unit, and showed preference for C30–34 lipid units with methyl branches in the S-configuration. Thus, in all cases T cell responses were strongest for synthetic compounds that mimicked the natural branched lipids produced by mycobacterial polyketide synthase 12. Incorporation of methylmalonate to form branched lipids is a common bacterial lipid synthesis pathway that is absent in vertebrates, so the preferential recognition of branched lipids may represent a new type of lipid-based pathogen associated molecular pattern (PAMP). PMID:18022562

Non-equivalent antigen presenting capabilities of dendritic cells and macrophages in generating brain-infiltrating CD8 + T cell responses.

PubMed

Malo, Courtney S; Huggins, Matthew A; Goddery, Emma N; Tolcher, Heather M A; Renner, Danielle N; Jin, Fang; Hansen, Michael J; Pease, Larry R; Pavelko, Kevin D; Johnson, Aaron J

2018-02-12

The contribution of antigen-presenting cell (APC) types in generating CD8 + T cell responses in the central nervous system (CNS) is not fully defined, limiting the development of vaccines and understanding of immune-mediated neuropathology. Here, we generate a transgenic mouse that enables cell-specific deletion of the H-2Kb MHC class I molecule. By deleting H-2K b on dendritic cells and macrophages, we compare the effect of each APC in three distinct models of neuroinflammation: picornavirus infection, experimental cerebral malaria, and a syngeneic glioma. Dendritic cells and macrophages both activate CD8 + T cell responses in response to these CNS immunological challenges. However, the extent to which each of these APCs contributes to CD8 + T cell priming varies. These findings reveal distinct functions for dendritic cells and macrophages in generating CD8 + T cell responses to neurological disease.