ABSTRACT
We report the preparation of gold nanoparticle (AuNP)-based vaccine candidates against the tumor-associated form of the mucin-1 (MUC1) glycoprotein. Chimeric peptides, consisting of a glycopeptide sequence derived from MUC1 and the T-cell epitope P30 sequence were immobilized on PEGylated AuNPs and the ability to induce selective antibodies in vivo was investigated. After immunization, mice showed significant MHC-II mediated immune responses and their antisera recognized human MCF-7 breast cancer cells. Nanoparticles designed according to this report may become key players in the development of anticancer vaccines.
Subject(s)
Cancer Vaccines/immunology , Epitopes, T-Lymphocyte/immunology , Glycopeptides/immunology , Gold/chemistry , Metal Nanoparticles/chemistry , Mucin-1/immunology , Amino Acid Sequence , Animals , Cancer Vaccines/chemistry , Epitopes, T-Lymphocyte/chemistry , Genes, MHC Class II , Glycopeptides/chemistry , Humans , Immunization , MCF-7 Cells , Mice , Molecular Sequence Data , Mucin-1/chemistry , Neoplasms/immunology , Neoplasms/prevention & controlABSTRACT
In studies within the realm of cancer immunotherapy, the synthesis of exactly specified tumor-associated glycopeptide antigens is shown to be a key strategy for obtaining a highly selective biological reagent, that is, a monoclonal antibody that completely differentiates between tumor and normal epithelial cells and specifically marks the tumor cells in pancreas tumors. Mucin MUC1, which is overexpressed in many prevalent cancers, was identified as a promising target for this strategy. Tumor-associated MUC1 differs significantly from that expressed by normal cells, in particular by altered glycosylation. Structurally defined tumor-associated MUC1 cannot be isolated from tumor cells. We synthesized MUC1-glycopeptide vaccines and analyzed their structure-activity relationships in immunizations; a monoclonal antibody that specifically distinguishes between human normal and tumor epithelial cells was thus generated.
Subject(s)
Antibodies, Monoclonal/biosynthesis , Breast Neoplasms/pathology , Breast/cytology , Cancer Vaccines/administration & dosage , Glycopeptides/immunology , Pancreatic Neoplasms/diagnosis , Female , HumansABSTRACT
Coevolution of ticks and the vertebrate immune system has led to the development of immunosuppressive molecules that prevent immediate response of skin-resident immune cells to quickly fend off the parasite. In this article, we demonstrate that the tick-derived immunosuppressor sialostatin L restrains IL-9 production by mast cells, whereas degranulation and IL-6 expression are both unaffected. In addition, the expression of IL-1ß and IRF4 is strongly reduced in the presence of sialostatin L. Correspondingly, IRF4- or IL-1R-deficient mast cells exhibit a strong impairment in IL-9 production, demonstrating the importance of IRF4 and IL-1 in the regulation of the Il9 locus in mast cells. Furthermore, IRF4 binds to the promoters of Il1b and Il9, suggesting that sialostatin L suppresses mast cell-derived IL-9 preferentially by inhibiting IRF4. In an experimental asthma model, mast cell-specific deficiency in IRF4 or administration of sialostatin L results in a strong reduction in asthma symptoms, demonstrating the immunosuppressive potency of tick-derived molecules.
Subject(s)
Cystatins/pharmacology , Immunity, Innate/drug effects , Immunosuppressive Agents/pharmacology , Interferon Regulatory Factors/immunology , Interleukin-9/immunology , Mast Cells/drug effects , Animals , Asthma/genetics , Asthma/immunology , Asthma/pathology , Binding Sites , Cell Degranulation/immunology , Cystatins/immunology , Gene Expression Regulation , Host-Parasite Interactions/immunology , Interferon Regulatory Factors/deficiency , Interferon Regulatory Factors/genetics , Interleukin-1beta/genetics , Interleukin-1beta/immunology , Interleukin-6/genetics , Interleukin-6/immunology , Interleukin-9/antagonists & inhibitors , Interleukin-9/genetics , Mast Cells/immunology , Mast Cells/pathology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Promoter Regions, Genetic , Protein Binding , Receptors, Interleukin-1/genetics , Receptors, Interleukin-1/immunology , Signal Transduction , Transcription, GeneticABSTRACT
The development of selective anticancer vaccines that provide enhanced protection against tumor recurrence and metastasis has been the subject of intense research in the scientific community. The tumor-associated glycoprotein MUC1 represents a well-established target for cancer immunotherapy and has been used for the construction of various synthetic vaccine candidates. However, many of these vaccine prototypes suffer from an inherent low immunogenicity and are susceptible to rapid in vivo degradation. To overcome these drawbacks, novel fluorinated MUC1 glycopeptide-BSA/TTox conjugate vaccines have been prepared. Immunization of mice with the 4'F-TF-MUC1-TTox conjugate resulted in strong immune responses overriding the natural tolerance against MUC1 and producing selective IgG antibodies that are cross-reactive with native MUC1 epitopes on MCF-7 human cancer cells.
ABSTRACT
The quality of the adaptive immune response depends on the differentiation of distinct CD4(+) helper T cell subsets, and the magnitude of an immune response is controlled by CD4(+)Foxp3(+) regulatory T cells (Treg cells). However, how a tissue- and cell type-specific suppressor program of Treg cells is mechanistically orchestrated has remained largely unexplored. Through the use of Treg cell-specific gene targeting, we found that the suppression of allergic immune responses in the lungs mediated by T helper type 2 (TH2) cells was dependent on the activity of the protein kinase CK2. Genetic ablation of the ß-subunit of CK2 specifically in Treg cells resulted in the proliferation of a hitherto-unexplored ILT3(+) Treg cell subpopulation that was unable to control the maturation of IRF4(+)PD-L2(+) dendritic cells required for the development of TH2 responses in vivo.
Subject(s)
Casein Kinase II/immunology , T-Lymphocytes, Regulatory/immunology , Th2 Cells/immunology , Animals , CD4-Positive T-Lymphocytes/enzymology , CD4-Positive T-Lymphocytes/immunology , Cell Differentiation/immunology , Cell Growth Processes/immunology , Cell Line , Dendritic Cells/enzymology , Dendritic Cells/immunology , Forkhead Transcription Factors/immunology , Humans , Hypersensitivity/blood , Hypersensitivity/immunology , Interferon Regulatory Factors/immunology , Leukocytes, Mononuclear/immunology , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Transgenic , Receptors, Cell Surface/immunology , T-Lymphocytes, Regulatory/enzymology , Th2 Cells/enzymologyABSTRACT
Self-adjuvanting antitumor vaccines by multifunctional cationic nanohydrogels loaded with CpG. A conjugate consisting of tumor-associated MUC1-glycopeptide B-cell epitope and tetanus toxin T-cell epitope P2 is linked to cationic nanogels. Oligonucleotide CpG complexation enhances toll-like receptor (TLR) stimulated T-cell proliferation and rapid immune activation. This co-delivery promotes induction of specific MUC1-antibodies binding to human breast tumor cells without external adjuvant.
Subject(s)
Adjuvants, Immunologic/pharmacology , Cancer Vaccines/immunology , Glycopeptides/immunology , Hydrogel, Polyethylene Glycol Dimethacrylate/chemistry , Nanoparticles/chemistry , Oligodeoxyribonucleotides/chemistry , Amino Acid Sequence , Animals , Cations , Enzyme-Linked Immunosorbent Assay , Glycopeptides/chemistry , Humans , Hydrogel, Polyethylene Glycol Dimethacrylate/chemical synthesis , Mice, Inbred BALB C , Molecular Sequence Data , Nanoparticles/ultrastructureABSTRACT
For antitumor vaccines both the selected tumor-associated antigen, as well as the mode of its presentation, affect the immune response. According to the principle of multiple antigen presentation, a tumor-associated MUC1 glycopeptide combined with the immunostimulating T-cell epitope P2 from tetanus toxoid was coupled to a multi-functionalized hyperbranched polyglycerol by "click chemistry". This globular polymeric carrier has a flexible dendrimer-like structure, which allows optimal antigen presentation to the immune system. The resulting fully synthetic vaccine induced strong immune responses in mice and IgG antibodies recognizing human breast-cancer cells.
Subject(s)
Antigens, Neoplasm/immunology , Cancer Vaccines/immunology , Glycerol/chemistry , Glycopeptides/chemical synthesis , Polymers/chemistry , Animals , Antibodies/immunology , Antigen Presentation , Breast Neoplasms/drug therapy , Cancer Vaccines/therapeutic use , Click Chemistry , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/immunology , Female , Glycopeptides/immunology , Humans , MCF-7 Cells , Mice , Mucin-1/chemistry , Mucin-1/metabolism , Tetanus Toxoid/chemistry , Tetanus Toxoid/metabolismABSTRACT
Highly decorated: Tumor-associated MUC1 glycopeptide and tetanus toxoid T-cell epitope P2 can be attached to water-soluble poly(N-(2-hydroxypropyl)methacrylamide) carriers by orthogonal ligation techniques. Fully synthetic vaccine A with additional nanostructure-promoting domains induced antibodies that exhibit high affinity to tumor cells.
Subject(s)
Cancer Vaccines/chemistry , Epitopes, T-Lymphocyte/chemistry , Glycopeptides/chemistry , Polymethacrylic Acids/chemistry , Amino Acid Sequence , Animals , Cancer Vaccines/immunology , Epitopes, T-Lymphocyte/immunology , Glycopeptides/immunology , Humans , MCF-7 Cells , Mice , Molecular Sequence Data , Mucin-1/chemistry , Mucin-1/immunology , Solubility , T-Lymphocytes, Helper-Inducer/immunology , Vaccines, Synthetic/chemistry , Vaccines, Synthetic/immunology , Water/chemistryABSTRACT
Ticks developed a multitude of different immune evasion strategies to obtain a blood meal. Sialostatin L is an immunosuppressive cysteine protease inhibitor present in the saliva of the hard tick Ixodes scapularis. In this study, we demonstrate that sialostatin L strongly inhibits the production of IL-9 by Th9 cells. Because we could show recently that Th9-derived IL-9 is essentially involved in the induction of asthma symptoms, sialostatin L was used for the treatment of experimental asthma. Application of sialostatin L in a model of experimental asthma almost completely abrogated airway hyperresponsiveness and eosinophilia. Our data suggest that sialostatin L can prevent experimental asthma, most likely by inhibiting the IL-9 production of Th9 cells. Thus, alternative to IL-9 neutralization sialostatin L provides the basis for the development of innovative therapeutic strategies to treat asthma.
Subject(s)
Asthma/immunology , Cystatins/immunology , Interleukin-9/immunology , Ixodidae/immunology , T-Lymphocyte Subsets/immunology , Animals , Asthma/metabolism , Asthma/prevention & control , Cell Separation , Cystatins/pharmacology , Cytokines/immunology , Cytokines/metabolism , Disease Models, Animal , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , Interleukin-9/biosynthesis , Male , Mice , Mice, Inbred BALB C , Mice, Knockout , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , T-Lymphocyte Subsets/metabolismABSTRACT
A shot in the arm for cancer treatment: two MUC1 tetanus toxoid vaccines were synthesized and induced a strong immune response in mice. The antibodies elicited by the vaccines show a high selectivity for the tumor cells in mammary carcinoma tissues and also distinguish between tumor tissues at different stages.
Subject(s)
Antineoplastic Agents/pharmacology , Breast Neoplasms/drug therapy , Cancer Vaccines/pharmacology , Glycopeptides/immunology , Mammary Neoplasms, Experimental/drug therapy , Mucin-1/immunology , Vaccines, Synthetic/pharmacology , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/immunology , Breast Neoplasms/immunology , Breast Neoplasms/pathology , Cancer Vaccines/chemical synthesis , Cancer Vaccines/immunology , Drug Screening Assays, Antitumor , Female , Glycopeptides/chemistry , MCF-7 Cells , Mammary Neoplasms, Experimental/immunology , Mammary Neoplasms, Experimental/pathology , Mice , Mice, Inbred BALB C , Molecular Conformation , Mucin-1/chemistry , Vaccines, Synthetic/chemistry , Vaccines, Synthetic/immunologySubject(s)
Antigens, Tumor-Associated, Carbohydrate/chemistry , Cancer Vaccines/chemistry , Fluorine/chemistry , Mucin-1/chemistry , Tetanus Toxoid/chemistry , Animals , Antibodies/metabolism , Breast Neoplasms/prevention & control , Breast Neoplasms/therapy , Cancer Vaccines/immunology , Cancer Vaccines/therapeutic use , Cell Line, Tumor , Female , Flow Cytometry , Glycopeptides/chemistry , Humans , Mice , Vaccines, Synthetic/chemistry , Vaccines, Synthetic/immunology , Vaccines, Synthetic/therapeutic useABSTRACT
Interferon-regulatory factor 4 (IRF4) is essential for the development of T helper 2 (Th2) and Th17 cells. Herein, we report that IRF4 is also crucial for the development and function of an interleukin-9 (IL-9)-producing CD4(+) T cell subset designated Th9. IRF4-deficient CD4(+) T cells failed to develop into IL-9-producing Th9 cells, and IRF4-specific siRNA inhibited IL-9 production in wild-type CD4(+) T cells. Chromatin-immunoprecipitation (ChIP) analyses revealed direct IRF4 binding to the Il9 promoter in Th9 cells. In a Th9-dependent asthma model, neutralization of IL-9 substantially ameliorated asthma symptoms. The relevance of these findings is emphasized by the fact that the induction of IL-9 production also occurs in human CD4(+) T cells accompanied by the upregulation of IRF4. Our data clearly demonstrate the central function of IRF4 in the development of Th9 cells and underline the contribution of this T helper cell subset to the pathogenesis of asthma.
Subject(s)
Interferon Regulatory Factors/immunology , Interleukin-9/immunology , T-Lymphocytes, Helper-Inducer/immunology , Animals , Asthma/genetics , Asthma/immunology , Cell Differentiation , Cells, Cultured , Humans , Interferon Regulatory Factors/deficiency , Interferon Regulatory Factors/genetics , Interferon Regulatory Factors/metabolism , Interleukin-9/biosynthesis , Interleukin-9/genetics , Mice , Mice, Inbred C57BL , Mice, Knockout , Promoter Regions, Genetic , Protein Binding , RNA, Small Interfering/genetics , T-Lymphocytes, Helper-Inducer/cytologyABSTRACT
In humans and mice naturally occurring regulatory T cells (nTregs) are crucial for the maintenance of peripheral tolerance by controlling not only potentially autoreactive T cells but virtually all cells of the adaptive and innate immune system. Here we show that co-culture of murine dendritic cells (DC) and nTregs results in an immediate increase of cAMP in DC, responsible for a rapid down-regulation of co-stimulatory molecules (CD80, CD86). In addition, the inhibitory surface molecule B7-H3 on DC is up-regulated. Subsequently, nTreg-derived IL-10 inhibits the cytokine production (IL-6, IL-12) of suppressed DC therewith preserving their silent phenotype. Hence, our data indicate that nTregs effectively control exuberant immune responses by directly limiting the stimulatory capacity of DC via a sophisticated chronologic action of inhibitory signals.