TW201219053A - Targeting antigens to human dendritic cells via DC-asialoglycoprotein receptor to produce IL-10 regulatory T-cells - Google Patents

Abstract

Compositions and methods for targeting protein antigens to human DCs via DC-asialoglycoprotein receptor (DC-ASGPR) are disclosed herein. The DC-ASGPR carries an intracellular tyrosine-based and dileucine motif, resulting in the generation of such IL-10 Tregs both in vitro and in vivo. The methods of the present invention can be used for designing vaccines against autoimmune diseases where autoantigens are defined, such as type 1 diabetes and multiple sclerosis.

Description

201219053 VI. INSTRUCTIONS OF THE INVENTION: TECHNICAL FIELD OF THE INVENTION The present invention relates generally to immunomodulation, and more particularly to targeting a protein antigen via a DC-asial acid glycoprotein receptor (DC-ASGPR) Human dendritic cells (DC) are produced to produce IL-10 regulatory T cells. • [Prior Art] The prior art of the present invention is described in connection with modulating immune responses against autoantigens and foreign antigens and limiting immunopathology associated with infection and autoimmunity without limiting the scope of the present invention. U.S. Patent Application Publication No. 20080206262 (Banchereau et al., 2008) includes compositions and methods for making and using anti-DC-ASGPR antibodies which, for example, can activate DCs and other cells. The method of the invention comprises the steps of isolating and purifying a DC-ASGPR-specific antibody or a fragment thereof, which is ligated to an antigen to form an antibody-antigen complex, wherein the antigen is derived from an antibody-antigen The complex-contacted dendritic cells process and present ^ DC-ASGPR-specific antibodies or fragments thereof to bind half of the adhesion protein (Coherin)/dystin (Dockerin), and the antigen and the cyclin/anchor protein are Complementary half. Combines to form a complex. U.S. Patent Application Publication No. 2008 No. 241,170 (Zurawski and »

Banchereau, 2008) includes compositions and methods for enhancing the effectiveness of antigen presentation using DCIR-specific antibodies or fragments thereof that are ligated to an antigen to form an antibody-antigen complex, wherein the antigen is Dendritic cells that are in contact with the antibody-antigen complex are processed and present in 159403.doc 201219053 NO [Abstract] The present invention describes the targeting of protein antigens to human DCs via DC-asial acid glycoprotein receptor (DC-ASGPR) In compositions and methods, the DC-desialin receptor carries intracellular tyrosine-based and leucine-based motifs that cause the production of such IL-10 Tregs in vitro and in vivo. In one embodiment, the invention encompasses compositions and methods for producing one or more antigen-specific regulatory T cells (Tregs) comprising: isolating one or the other species of human dendritic cells (DC) from an individual; Or a plurality of antigen-binding or fused anti-DC- asialoglycoprotein receptor (DC_ASGpR)-specific antibodies or binding fragments thereof, the one or more antigens are loaded into one or more Dc to form an antigen-loaded DC; and The dc carrying the antigen is contacted with one or more original butyl cells, wherein the antigen-loaded DC stimulates proliferation of antigen-specific Tregs. In one aspect, the one or more antigens comprise a peptide or protein. In another aspect, the peptide is a foreign antigen or an autoantigen. In another aspect, the peptide triggers an allergic or asthmatic response. In another aspect, the one or more antigens comprise a bacterial protein, a viral protein, a fungal protein, a protozoal protein, or a cancer protein. In another aspect, the antigen comprises H, PSA, or a combination thereof and a modification. In another aspect, the antigen-specific Tregs secrete IL_1〇iTreg. In another aspect, the dendritic cell line is used to prevent or treat one or more autoantigens"guided autoimmune diseases, multiple sclerosis, influenza or cancer' to ameliorate the symptoms of the above diseases. In another aspect, the autoimmune disease is selected from the group consisting of alpha: allergic fistula, asthma, snoring disease, type 3 diabetes (IDDM), systemic lupus erythematosus (SLE), and Sjogren's syndrome. 159403.doc 201219053 (Sj6grenis syndrome), Churg Strauss Syndrome, Hashim〇t〇, s thyr〇iditis, Graves' disease, special Thrombocytopenic purpura, transplant rejection, multiple sclerosis, psoriasis and rheumatoid arthritis (RA) » In another aspect, the anti-system is made from fusion tumor cells contained in ATCC accession number ρτΑ_ 10248 Another embodiment of the invention includes a vaccine composition that is resistant to one or more autoantigen-mediated autoimmune diseases, comprising an anti-DC-sialic acid glycoprotein that binds or fuses with one or more autoantigens. An (ASGpR)-specific antibody or binding fragment thereof, and one or more pharmaceutically acceptable adjuvants, optionally produced, wherein the vaccine composition produces one or more autoantigen-specific secretions of E. coli Tune T cells (of Treg), to enhance the one or more kinds of T cells producing one or more of the cells of producing τ, or both to generate the one or more cells and enhanced butoxy. In one aspect, the autoimmune disease is selected from the group consisting of allergy, asthma, celiac disease, type 3 diabetes (IDDM), systemic lupus erythematosus (SLE), Hugh's syndrome, and singularity _ Siquan syndrome 'Hashimoto's thyroid gland, Graves' disease, idiopathic thrombocytopenic purpura, transplant rejection, multiple sclerosis, psoriasis and rheumatoid arthritis (RA). In another aspect, the Treg is an autoantigen-specific IL-10 Treg. In another aspect, the vaccine is administered orally, parenterally or intranasally. In another aspect, the one or more antigens comprise peptides, proteins, lipids, carbohydrates, nucleic acids, and combinations thereof. In another aspect, the composition binds to and activates dendritic cells that activate IL-10-secreting Treg. In another aspect, the anti-system is prepared from a 159403.doc 201219053 tumor-bearing cell contained in ATCC Accession No. 〇1〇248. Another embodiment of the present invention includes a method of treating, preventing, or improving cancer symptoms in an individual, comprising the steps of: identifying an individual in need of treatment, prevention or amelioration of symptoms for prostate cancer; administering a therapeutically effective amount of the medicament A composition or a vaccine sufficient to treat, prevent or ameliorate symptoms, wherein the composition comprises: an anti-DC- asialoglycoprotein receptor (ASGPR)-specific antibody or binding fragment thereof, which binds to one or more cancer-specific antigens Or a fused recombinant fusion protein and one or more pharmaceutically acceptable excipients or adjuvants as appropriate. In one aspect, the one or more antigens comprise a peptide or protein. In another aspect, the peptide is a foreign antigen or an autoantigen. In one aspect, the cancer-specific antigen is a peptide selected from the group consisting of a tumor-associated antigen selected from the group consisting of CEA, prostate, prostate specific antigen (PSA), HER-2/neu, BAGE, GAGE, MAGE 1 to 4, 6 and 12, MUC (mucin) (eg MUC-1, MUC-2, etc.), GM2 and GD2 neuroglycosmic lipids, ras, myc, tyrosinase, MART (melanoma antigen), MARCO -MART, cyclin B1, cyclin D, Pmel 17 (gpl00), GnT-V intron V sequence (N-ethyl glucosyltransferase V intron V sequence), prostate cancer psm, prostate Serum antigen (PSA), PRAME (melanoma antigen), β-sodium hydrocyclone, MUM-1-B (melanoma common mutation gene product), GAGE (melanoma antigen) 1, BAGE (melanoma antigen) 2 to 10, c-ERB2 (Her2/neu), EBNA (Epstein-Barr virus nuclear antigen) 1 to 6, gp75, human papillomavirus (HPV) E6 and E7, p53, lung resistance Protein (LRP), Bcl-2 and Ki-67. In another aspect, the resistance system is included in the ATCC Depository 159403.doc

S 201219053 PTA-10248 fusion tumor cells were prepared. Another embodiment includes a method of treating, preventing, or ameliorating a pathogen symptom in an individual comprising the steps of: identifying an individual in need of treatment for the pathogen, (d) or ameliorating the symptoms; administering a therapeutically effective amount of the pharmaceutical composition or A vaccine sufficient to treat, prevent or ameliorate the symptoms, "This composition comprises a recombinant fusion of an anti-Dc asialoglycoprotein receptor (ASGpR) specific antibody or binding fragment thereof or - or multiple pathogen antigens. Protein and/or a plurality of pharmaceutically acceptable excipients or adjuvants, optionally selected, in one aspect, the vaccine is administered orally, parenterally or intranasally. In another aspect' The vaccine produces one or more pathogen-specific regulatory T cells (Tregs), increases the content of the one or more tau cells, or both produces the one or more T cells and increases the content of the one or more T cells. The anti-system is prepared from a fusion tumor cell containing the SATCC accession number pta_1 248. Another embodiment of the invention includes treating and preventing autologous antigen-mediated autologous exemption in an individual. A method for discriminating the symptoms of an autoimmune disease, comprising the steps of: identifying an individual in need of treatment, preventing an autoimmune disease, or ameliorating the symptoms of an autoimmune disease; and administering a therapeutically effective amount of the vaccine, the vaccine comprising an antibiotic a recombinant fusion protein in which a DC-desial acid glycoprotein receptor (ASGpR)-specific antibody or binding fragment thereof binds or fuses with - or a plurality of autoantigens, and one or more pharmaceutically acceptable adjuvants, optionally selected, Wherein the vaccine composition produces - or a plurality of autoantigen-specific regulatory T cells (Tregs), enhances the production of the one or more tau cells, or both produces the one or more T cells and enhances the one or more T cells. Manufacturing. In the case of 159403.doc 201219053, the autoimmune disease is selected from the group consisting of allergies, asthma, celiac disease, type 3 diabetes (IDDM), systemic lupus erythematosus (SLE), and Hugh's disease. Symptoms 'Church _ Siquan syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, transplant rejection ^ sclerosis, psoriasis and rheumatoid arthritis (10)). In another case, the autoimmune disease is type j diabetes. In another aspect, Treg is secreted by IL, and the autoantigen is specifically called. In another aspect, the 9 seedlings are administered sigma, parenterally or intranasally. In another aspect, the - or antigens comprise peptides, proteins, lipids, carbohydrates, nucleic acids, and combinations thereof. In another aspect, the anti-system was prepared from fusion tumor cells containing the ACC accession number ΡΤΑ-1〇248. Another embodiment of the invention includes 吝ώ & =

^栝 Producing a self-antigen-specific regulatory T cell (D-(10) pharmaceutical composition comprising: an anti-sialic acid-protein receptor (ASGPR)-specific antibody or a binding fragment thereof, or a plurality of autoantigens Recombinant fusion protein; and - or a plurality of pharmaceutically acceptable excipients or adjuvants as appropriate, in one aspect, the composition is for the prevention, treatment of one or more autoantigen-mediated forms Autoimmune disease, multiple sclerosis, influenza or cancer, and a disease shield that improves the above diseases. In another aspect, the 'self antigen number is selected from the group consisting of anti-allergy, asthma, celiac disease, the first cause Type 2 diabetes (1), systemic lupus erythematosus (SLE), Hugh's syndrome, qiqi _ shi syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic such as small plate reduction 2. Transplant rejection, multiple sclerosis, psoriasis, and rheumatoid granules (RA). In another aspect, the anti-system is made from fusion tumor cells contained in ATCC-registered 159403.doc 201219053 PTA-10248. Another embodiment is included Compositions of antigen-specific Tregs prepared by the method of the present invention. [Embodiment] In order to more fully understand the features and advantages of the present invention, reference is now made to the embodiments of the invention and the accompanying drawings. The preparation and use of the various embodiments of the present invention, but it should be understood that the present invention is intended to be in various embodiments of the present invention, which may be practiced in various specific embodiments. The specific embodiments discussed herein are merely illustrative of the preparation and use of the present invention. The scope of the invention is not limited thereto. To help understand the invention, a number of terms are defined below. Terms defined herein have the meanings commonly understood by those skilled in the relevant art to which the invention pertains. Terms such as "one" and "the" It is intended that the singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular singular The term "antigen" or "rAg" as used herein refers to a substance that is capable of eliciting an immune response, such as For example, a T cell-mediated immune response elicited by an antigen on a major histocompatibility antigen (MHC) cellular protein and causing an antigen-specific tau cell reaction. Targeted in regulatory T cells (Treg) In the case of the original reaction, the immune response caused by other effector cells (such as helper T cells (Th) and/or cell sputum T cells (Tc) is reduced or improved. Proficiency immunologists will recognize that when discussing antigens When processed to present T cells, the term "antigen" refers to those antigenic portions (eg, peptide fragments) that are presented to the T cell epitope of the T cell receptor by MHC. When the antigen is passed through 159403.doc 201219053 Or autologous modification, which refers to an autoantigen or autoantigen that is normally present in the mhc molecule but also triggers a T cell response. When used in the context of a B cell-mediated immune response, the antibody is used in the form of an antibody that is specific for the antigen, ie, the antigenic portion that binds to the complementarity determining region of the antibody variable domain (light and heavy) It can be a linear or three-dimensional epitope. In some instances, the antigen delivered by the vaccine or fusion protein of the invention is rendered internalized and processed by the antigen prior to presentation, e.g., by cleavage of one or more portions of the antibody or fusion protein. The term "antigenic peptide" as used herein refers to a polymorphic antigen portion that is lined by B cells and/or T,, 'field-specific s lines. b cells are produced by antibody production to respond to foreign antigenic determinants, while T lymphocytes mediate cell immunity. Thus, the antigenic peptide in the T cell response is the portion of the antigen that is recognized by the antigen-specific T cell receptor in the case of Mhc. The term "antigenic determinant" as used herein refers to a T cell receptor capable of specifically binding to an immunoglobulin or capable of being expressed by a major histocompatibility complex (MHC) protein (eg, class I or class II). Any of the protein determinants. The antigenic determinant is usually a short of 5 to 30 amino acid lengths. It is assembled in the groove of the MHC molecule that presents certain amino acid side groups to the T cell receptor. And having some other residue in the groove, for example due to the specific charge characteristics of the groove, has a peptide side group and a T cell receptor. Generally, when the dissociation constant is 1 mM, 1 〇〇 nM or even 10 nM, the antibody specifically binds to the antigen. The term "antigen presenting cells" (APC) as used herein is a cell capable of activating a buty cell' and includes, but is not limited to, certain macrophages, B cells 159403.doc 201219053 and dendritic cells. "dendritic cells" (DC) refers to any member of a different population of cell types found in lymphoid or non-lymphoid tissues. These cells are characterized by their unique morphology, surface π-like MHC high expression (Steinman et al, Ann Rev. Immun〇l. 9:271 (1991); their description of these cells is incorporated herein by reference. in). As described herein, such cells can be isolated from a variety of tissue sources and conveniently from peripheral blood. Dendritic cell-binding protein refers to any protein to which the receptor is expressed on dendritic cells. Examples include GM-CSF, IL-1, TNF, IL-4, CD40L, CTLA4, CD28, and FLT-3 ligands. For the purposes of the present invention, the term "vaccine composition" means a composition which can be administered to a human or animal to induce an immune system response; this immune system reaction can cause the production of antibodies or cause only certain cells (especially antigen-presenting cells). Activation of T lymphocytes and b lymphocytes). The vaccine composition can be a composition for prophylactic purposes or for therapeutic purposes or for both purposes. As used herein, the term "antigen" refers to any antigen that can be used in a vaccine, whether or not it involves an entire microorganism or subunit, and regardless of its nature: peptides, proteins, glycoproteins, polysaccharides, glycolipids, lipopeptides, and the like. It may be a viral antigen, a bacterial antigen or an analogue thereof; the term "antigen" also encompasses a polynucleotide whose sequence is selected to encode an antigen that is expressed by an individual to be administered by such a polynucleotide' in immunological technology. In the case of DN A immunization, it can also be a group of antigens, especially in the case of multivalent vaccine compositions comprising antigens that are resistant to several diseases, which are generally referred to as vaccine combinations or in the presence of several different antigens. In the case of compositions that are resistant to a single disease, such as certain vaccines against pertussis or influenza. The term "antibody" refers to an immunoglobulin, whether naturally or partially artificially or completely artificially (e.g., recombinant), which may be a monoclonal antibody or a plurality of antibodies. In some cases, the antibody can be a member of an immunoglobulin class or a combination thereof, including: IgG, IgM, IgA, IgD, and IgE. The term "antibody" includes, but is not limited to, naturally occurring antibodies isolated and/or purified, and non-naturally occurring antibodies. Specifically, the term "antibody" includes a plurality of antibodies and monoclonal antibodies, and a binding fragment thereof that continues to bind to the antigen. Furthermore, the term "antibody" includes chimeric antibodies as well as fully synthetic antibodies and fragments thereof. Multiple strains of anti-system are derived from the serum of animals immunized with antigen. Monoclonal antibodies can be prepared using fusion cell technology (Köhler et al, Nature 256:495 (1975); Hammerling et al. 'M_cl〇nal Antibodies and T-Cell Hybridomas, Elsevier, Ν·Υ. pages 563 to 681 Page (1981)) 'The relevant portions of these references are hereby incorporated by reference. Antibodies also include polyclonal antibodies, affinity-purified polyclonal antibodies, monoclonal antibodies' and antigen-binding fragments, such as F(ab,)2 and Fab proteolytic fragments. Also included are genetically engineered intact antibodies or fragments that bind to DC-ASGPR, such as chimeric antibodies, Fv fragments, single chain antibodies and analogs thereof, as well as synthetic antigen binding peptides and polypeptides. Non-human antibodies can be grafted onto human frameworks and constant regions by non-human CDRs or by incorporation of entire non-human variable domains (as appropriate by replacing exposed residues to "cover" human-like surfaces, "antibody" is humanized. In some cases, humanized antibodies may retain non-human residues within the human variable region framework domain to enhance appropriate binding characteristics. The human half-life is prolonged by humanizing the antibody and the likelihood of a harmful immune response after administration to humans is reduced. Furthermore, as disclosed in, for example, 159403.doc 8 •12·201219053 WIPO Publication WO 98/24893, 乂α^ un-human antibodies can be engineered into non-human animals that have been transfected with human immunoglobulin genes. Manufacture, the relevant part of the case is incorporated herein by reference. The term "humanized antibody" refers to a chimeric antibody comprising a definitive region from a human antibody and most or all of the framework sequences from human variable "all or most of the CDRs are derived from a mixed variable region of a non-human variable region. Humanized antibodies, also known as chimeric antibodies or veneer antibodies, are described by recombinant techniques and readily available starting materials, and are described in, for example, British Patent Application GB 2,188,638 A. Incorporated herein by reference. The term "effective amount" or "therapeutically effective amount" means a biological or medical response that will result in a tissue, system, animal or human being sought by a researcher, veterinarian, physician or other clinician. The amount of the compound of the invention. The term "treating" as used herein refers to administration of a compound of the invention and includes (1) inhibiting the disease of an animal undergoing or showing the pathology or syndrome of the disease (ie, further curbing pathology and/or syndrome). Development) or (7) improvement of the disease of an animal that experiences or shows the pathology or symptom of the disease: (ie, reversal of pathology and/or syndrome). The term "control" includes the prevention, treatment, eradication, amelioration or otherwise reducing the severity of the condition being controlled. The term "fusion protein" refers to an expression product of two or more nucleic acid molecules that are not naturally expressed as a manifestation product. Fusion proteins can place proteins or peptides at the nucleic acid coding level by in-line and in the correct coding framework. Manufactured in two or more sequences. Fusion proteins are obtained by methods known to those skilled in the art (including, for example, solid phase protein synthesis) and 159403.doc -13-201219053 by allowing molecular manipulation of DN A in vitro (including polymerase chain reaction (PCR) ) and oligonucleotide-directed mutagenesis induced synthesis. The fusion protein used in the present invention is an immunotoxin including an antigen-binding portion, in this case, an anti-DC-ASGPR antibody or a fragment thereof and a toxin. The term "autoimmune disorder" as used herein refers to a disease condition in which a patient's immune system recognizes an autoantigen or autoantigen in a patient's organ or tissue as a foreign antigen and becomes activated. Activating immune cells directed against autoantigens or autoantigens can cause damage to the target organ or tissue or can damage other organs or tissues. The dysregulated immune cells secrete an inflammatory cytokine that causes systemic inflammation or can recognize the autoantigen as a foreign antigen, thereby promoting an immune response against the autoantigen. Non-limiting examples of autoimmune diseases include: rheumatoid arthritis, autoimmune or autoinflammatory disease of the skin, allergies, sclerosis, arteriosclerosis, multiple sclerosis, asthma, psoriasis, lupus, systemic erythema Lupus, diabetes, myasthenia gravis, chronic fatigue syndrome, muscle fiber pain, Crohn's disease, Hashimoto's thyroiditis, Graves' disease, Addison's disease, - Ba 2 syndrome (plus (1) is also like... syndrome) and scleroderma. "Autoimmune" refers to the allergic immune response to autoantigens. "Autoimmune disease" refers to a condition in which the immune system responds to the "self" antigen that is normally ignored and causes normal body tissue damage. Autoimmune disorders are thought to be caused, at least in part, by allergic reactions like allergies' because in both cases the immune system responds to substances that are normally ignored. Autoimmune disorders include, for example, Hashimoto's thyroiditis, pernicious anemia, Addison's disease, type 姨 (姨 素 Dependence) diabetes, phobia thirst 159403.doc -14- 201219053 arthritis, systemic lupus erythematosus , Hugh's syndrome, multiple sclerosis, myasthenia gravis, Reyter's syndrome (Rejter, s Syndr〇me) and Graves' disease, plaque alopecia, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Hemolytic anemia, autoimmune hepatitis, autoimmune lymphoproliferative syndrome (ALPS), autoimmune thrombocytopenic purpura (ατρ), Behcet's disease, bullous pemphigoid, cardiomyopathy, Celiac sprue-type dermatitis, chronic fatigue syndrome immunodeficiency syndrome (CFIDS), chronic inflammatory demyelinating polyneuropathy, scar pemphigoid, cold agglutinin disease, restrictive scleroderma ( CREST syndrome), Crohn's disease, Dege's disease (Deg〇, s(1) for phantom, dermatomyositis, discoid lupus, primary mixed cryoglobulinemia, muscle fiber pain-fibrosis Inflammation, Ge-Barth's syndrome, idiopathic pulmonary fibrosis, idiopathic thrombocytopenic purpura (ITP), IgA nephropathy, juvenile rheumatoid arthritis, Meniere's disease, Mixed connective tissue disease, pemphigus vulgaris, nodular polyarteritis, polychondritis, polyadenotrophic syndrome, rheumatic polymyalgia, polymyositis, primary no globulinemia, primary biliary Liver cirrhosis, psoriasis, Raynaud's phenomenon, rheumatic fever, sarcoma-like disease, scleroderma, ft-human syndrome, Takayasu arteritis, ankle arthritis/macrocellular arthritis, ulcerative colon Inflammation, uveitis, vasculitis, leukoplakia and Wegener, s granul〇mat〇sis. Certain autoimmune disorders are also chronic inflammatory diseases, which are usually defined as long-term cells (greater than 6 Months) Activating the process of inflammation that causes inflammation. Chronic inflammation can also cause damage to organs or tissues of patients. Many diseases are chronic inflammatory diseases 159403.doc 201219053 'but not known to have autoimmune basis For example, atherosclerosis, congestive heart failure, Crohn's disease, ulcerative colitis, nodule f, ectopic arteritis, Whipple's disease, and primary sclerosing cholangitis. Other antigenic peptides include cancer peptides selected from tumor-associated antigens, such as autologous cancer antigens obtained from patients. Non-limiting examples of cancer antigens include antigens from leukemias and lymphomas, neuro-tumors (such as stars) Astrocytoma or glioblastoma), melanoma, breast cancer, lung cancer, head and neck cancer, gastrointestinal cancer, stomach cancer, colon cancer, liver cancer, pancreatic cancer, genitourinary tumors (such as cervical cancer, uterine cancer, nest cancer, Vaginal cancer, testicular cancer, prostate cancer or penile cancer), skeletal tumor, aneurysm, or lip cancer, nasopharyngeal cancer, pharyngeal cancer and oral cancer, esophageal cancer, rectal cancer, gallbladder cancer, cholangiocarcinoma, laryngeal cancer, lung cancer and Bronchial, bladder, kidney, brain, and other parts of the nervous system, cancer, squamous cell carcinoma, Hodgkin's disease, non-Hodge Non-Hodgkiis lymphoma, multiple myeloma, and leukemia. In a specific aspect, the composition further comprises an antigenic peptide selected from the group consisting of a tumor-associated antigen selected from the group consisting of CEA, prostate specific antigen (PSA), HER-2/neu, BAGE, GAGE, MAGE 1 to 4, 6 and 12, MUC (mucin) (eg MUC-1, MUC-2, etc.), GM2 and GD2 neuroglycosides, ras, myc, tyrosinase, MART (melanoma antigen), MARCO -MART, cyclin B1, cyclin d, pmei 17 (gplOO), GnT-V intron V sequence (N-acetyl glucosyltransferase v intron V sequence), prostate cancer psm, prostate Serum antigen (psA), 159403.doc 16 201219053 PRAME (melanoma antigen), β_sodium hydrocarbon, (melanoma common mutation gene product), GAGE (melanoma antigen) 丨, BAGE (melanoma antigen) 2 to 10, c-ERB2 (Her2/neu), EBNA (Epstein-Barr virus nuclear antigen) 1 to 6, gP75, human papillomavirus (Hpv) E6 & E7, p53, lung resistance protein (LRP), Bcl-2 and Ki-67. The manufacture and use of Tr eg secreting IL-10 can also be used to treat conditions such as inflammatory bowel disease caused by butyl cells recognizing one or more autoantigens or autoantigens, such as ileitis and ulcerative colitis. And Crohn's disease's inflammatory lung disease, such as bronchitis, oxidant-induced lung injury and chronic obstructive airway disease; ocular inflammatory diseases such as corneal dystrophy, high intraocular pressure, trachoma, Jingwei silk Insect disease, retinitis, uveitis, sympathetic ophthalmia and endophthalmitis; chronic inflammatory gingival conditions, including periodontitis; chronic inflammatory joint disorders such as joints, septic arthritis and osteoarthritis, tuberculosis Sexual joints, leprosy arthritis, sarcoma-like arthritis; skin disorders such as sclerosing dermatitis, sunburn psoriasis and eczema; encephalomyelitis and viral encephalitis or autoimmune encephalitis; autoimmune diseases Including immune-complex vasculitis, and heart H disease, such as ischemic heart disease, heart failure and cardiomyopathy. The term "adjuvant" refers to a substance that enhances, potentiates or potentiates the host's immune response to a vaccine antigen. The term "gene" is used to refer to a unit that encodes a functional protein, polymorphism or form. Those skilled in the art will appreciate that this functional term includes genomic sequences, cDNA sequences or fragments and combinations thereof, as well as gene products, including those that may have been artificially altered. Purified genes, nucleic acids, proteins and their 159403.doc -17-201219053 analogs are used to refer to such entities when distinguished and separated from at least one contaminating nucleic acid or protein normally associated with a compound. The term "nucleic acid" or "nucleic acid molecule" as used herein refers to a polynucleotide, such as deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), an oligonucleotide, produced by polymerase chain reaction (PCR). Fragments and fragments produced by any of ligation, cleavage, nuclear Ss» endotransfer, and exonuclease. A nucleic acid molecule can be composed of a naturally occurring nucleotide (such as DNA and RNA) or an analog of a naturally occurring nucleotide (such as the alpha enantiomeric form of a naturally occurring nucleotide), or both The combination. The modified nucleotide may have a change in the sugar moiety and/or the screaming or quinone moiety. Sugar modifications include, for example, substitution of one or more hydroxyl groups with a hydroxyl group, an alkyl group, an amine, and an azide group, or the sugar can be functionalized as a hydrazine or an ester. In addition, the entire sugar moiety can be replaced by sterically and electronically similar structures such as saccharide and carbocyclic analogs. Examples of modifications in the base moiety include alkylated oxime and. A pyridine, hydrazine or pyrimidine or other well known heterocyclic substituent. The nucleic acid monomers may be linked by phosphodiester bonds or similar bonds of the bonds. Similar bonds of phosphodiester bonds include phosphorothioate linkages, phosphorodithioate linkages, selenophosphate linkages, diselenyl phosphate linkages, aniline phosphorothioate linkages, anilinophosphate linkages, amine scales Acid ester bonds and the like. The term "nucleic acid molecule" also includes so-called "peptide nucleic acids" which comprise naturally occurring or modified nucleobases linked to a polyamine backbone. The nucleic acid can be a single or double stranded nucleic acid. The term "amino acid" as used in this application means one of the aminocarboxylic acids constituting the natural presence of the protein. The term "polypeptide" as used herein refers to a polymer of amino acid residues linked by peptide bonds, whether natural -18 - 159403.doc

S 201219053 or synthetically produced. A polypeptide of less than about 10 amino acid residues is commonly referred to as a peptide. A "protein" is a macromolecule comprising one or more polypeptide chains. The protein may also comprise non-peptide components, such as carbohydrate groups. Carbohydrates and other non-peptide substituents can be added to the protein by the protein-producing cells and will vary depending on the cell type. The protein herein is defined in terms of its amino acid backbone structure; substituents such as carbohydrate groups are generally not specified, but they may still be present. The term "in vivo" as used herein refers to inside the body. The term "in vitro" or "in vitro" as used in this application shall be taken to mean an operation performed in an abiotic system or external to a biological system. The term "treatment" as used herein refers to administration of a compound of the invention and includes (1) inhibition of a disease in an animal that has undergone or shows the pathology or signs of the disease (ie, curbing pathology and/or symptomology). Further developments) or (2) improving the disease (ie, reversing pathology and/or syndrome) of an animal that experiences or displays the pathology or signs of the disease. The present invention describes compositions that produce one or more antigen-specific regulatory T cells (Tregs), increase the content of the one or more tau cells, or both the one or more tau cells and the one or more tau cells. Method for inducing in vitro and in vivo by targeting a protein antigen to a human DC14' via a DC_asial acid glycoprotein receptor (DC-ASGPR) carrying an intracellular tyrosine-based and leucine-based motif The IL-1() Treg is produced internally to achieve. In one embodiment, the invention includes an anti-DC-ASGPR antibody, which may be, for example, AB4-5.49C11.7 (HS4128), which has been deposited with the accession number ρτα·1〇248 in the American Type Culture Center (American Type Culture) I59403.doc • 19· 201219053

Collection), in accordance with the provisions of the Budapest Treaty on the International Recognition of the Deposit of Micro-organisms, the American Type Culture Collection (ATCC) 10801 University Boulevard, Manassas, Va. 20110-220 ° The cell (DC) can control the immune response 1G_13, 15 in part by expressing toll-like receptors and lectins. Human DC-lectins (such as Dectin-Ι, lectin-like oxidized LDL receptor (LOX-1) and DC-specific ICAM-3 binding non-integrin (DC-SIGN)) can transmit intracellular signals, causing T cell responses Alteration of I5, ". Human DC-ASGPR has a motif based on tyrosine and di-leucine 14 , but its biological function has not been fully investigated. Antibodies and other reagents: This study produces specificity for human DC lectin Monoclonal antibodies (mAbs), including LOX-1 and DC-ASGPR, and recombinant fusion proteins of HA1 and PSA. Use mAb labeled with Alexa fluor 488 or 568 (Molecular Probes). Antibodies for staining cell surfaces ( Anti-CD3, anti-CD4, anti-CDllc, anti-CD14, anti-CD19, anti-CD123, anti-CTLA-4 and anti-PD-1) lines were purchased from Biolegend (CA). To block IL-10, anti-IL-IO (BIIR) was used. And anti-IL-10R (R&D systems, MN). Antibodies and reagents for intracellular staining were purchased from BD Pharmingen (CA) and cells were stained according to the manufacturer's protocol. In general, for cell surface and Intracellular staining, both 0.5 x lO6 to 1χ1〇6 cells were combined with 〇·5 mg/ml to 1 mg/ml antibody. Incubate for 20-30 minutes anti TGFbl (lDll) and anti TGFbR purchased from R &. D antibody is used in the present study were purchased from BD Biosciences, and anti-anti-HLA-DR CDlc lines from.

159403.doc -20- a S 201219053

Biolegend, and the anti-LOX-1 line was purchased from Abcam (MA) recombinant IL-10 line from R&D. IFNa, IL-4 and GM-CSF were purchased from the Baylor University Medical Center (TX) pharmacy. IL-2 and CFSE were purchased from Peprotech (NJ) and Molecular Probes (CA), respectively. The HA1 and PSA peptide libraries are from Mimotopes (CA). Cells: IL-4DC (interleukin 4 activated or mature DC) and IFNDC (interferon gamma activated or mature dendritic cells) were prepared by culturing monocytes from healthy donors. Mononuclear cells were cultured in Cellgenix (UK) medium containing 100 ng/ml GM-CSF and 50 ng/ml IL-4 (IL_4DC) for 6 days or with 100 ng/ml GM-CSF and 500 units/ml IFNa. (IFNDC) was cultured for 3 days in Cellgenix (UK) medium. Total CD4+ T cells, CD14+ monocytes, and CD19+ B cells were purified by using a negative selection kit (StemCell, CA). The original CD4+ T cells (CD45RA + CD45RO-) (purity greater than 99.2%) were purified by FACS Aria (BD Biosciences, CA). CD4+ Τ cell response. 5χ103 IFNDCs loaded with 1 mg/ml to 2 mg/ml recombinant protein were co-cultured with 2χ105 purified and CFSE-labeled CD4+ T cells. Cell proliferation was tested by measuring CFSE dilution on day 6. On day 9 or day 10, CD4+ T cells were restimulated with APC loaded with 1 mM to 5 mM peptide for 5 hours in the presence of brefeldin A (1 ml/ml). The cells are then stained for intracellular cytokine expression. The cytokine content in the culture supernatant was measured using the BeadLyte Cytokine Assay Kit (Upstate, MA) according to the manufacturer's protocol. The allogeneic system was used in the inhibition assay, and CFSE low CD4+ T cells were selected from primary cultures and used as effector cells. Different numbers of effector cells from the same donor and newly purified CD4+ T cells in the presence of 1 〇mg/ml anti-IL-10 and anti-159403.doc 21 201219053 IL-10R antibody or the same concentration of control IgG (reaction) Cells) are co-cultured with heterologous DCs. At the last 18 hours prior to collection, CD4+ T cells were pulsed with 1 mL/well of 3[H] thymidine. The amount of 3[H] thymidine incorporation was measured by a 1450 Microbeta counter (Wallac, ΜΑ). Trans-well plates (96-well plate inserts, Nunc, PA) were used in the autoantigen-specific inhibition assay. CFSE low effector cells were generated with DCs loaded with anti-DC-ASGPR-HA1 or anti-LOX-1-HA1 and anti-DC-ASGPR-PSA or anti-LOX-1-PSA. The sorted effector cells were co-cultured with DCs loaded with 1 mM to 5 mM peptide in the upper wells. The CFSE-labeled purified CD4+ T cells (reactive cells) were co-cultured with DCs loaded with anti-LOX-1-HA1 in the lower wells. A blocking antibody or a control antibody is added to the lower well. On day 6, proliferation of CD4+ T cells in the lower wells was assessed by measuring CFSE dilutions using flow cytometry. On day 9, CD4+ T cells were restimulated with DC loaded peptide and stained for intracellular IFNg (IFNy) expression. In order to target antigens via DC-ASGPR, DCs produce monoclonal antibodies (mAbs) specific for human DC-ASGPR. An anti-LOX-1 mAb was generated as a control. Anti-LOX-l (IgG2a: pure 15C4) and anti-DC-ASGPR (1 Lu 02 &: pure line 49 (: 11) 111 eight 13 are associated with Zhao 〇 (: (1) 1 〇 (:) and € 014 + single Nuclear cells bind, but do not bind to plasmacytoid DCs (pDC) (Fig. 1A). In vitro cultured monocyte-derived IL-4DC and IFNDC also exhibit two lectins (Fig. 1B). In human skin, The cells expressing LOX-1 and DC-ASGPR are mainly localized in the dermis (Fig. 2A to Fig. 2D). HA1 (HA1 subunit of influenza virus A/PR/8/34, H1N1) and 159403.doc •22·

S 201219053 The gland-specific antigen (PSA) acts as a foreign antigen and an autoantigen, respectively. Recombinant mAbs fused to HA1 (anti-LOX-1-HA1, anti-DC-ASGPR-HA1 and IgG4-HAl) or PSA (anti-LOX-1-PSA, anti-DC-ASGPR-PSA and IgG4-PSA) were produced with two Mouse variable region of human site mutation (S228P and L235E) - human IgG4k chimera 17 (Fig. 3A). Both anti-LOX-1-HA1 and anti-DC-ASGPR-HA1 bind to IFNDC more efficiently than IgG4-HAl (Fig. 3B and Fig. 3C). Anti-LOX-1-HA1 and anti-DC-ASGPR-HA1 (1 mg/ml) induced more CD4+ T cell proliferation (greater than 66%) compared to IgG4-HAl (7%) or unloaded DC (1.7%) (Fig. 3D). Anti-LOX-l-PSA (25%) and anti-DC-ASGPR-PSA (18%) also induced more CD4+ T cell proliferation than IgG4-PSA (2.5%) (Fig. 3E). The antigen specificity of proliferating CD4+ T cells was first tested by measuring intracellular IFNg expression during restimulation of seven HA1-derived peptide clusters (Fig. 4). The individual peptides in cluster 4 were further analyzed (Fig. 5A). Both anti-LOX-1-HA1 and anti-DC-ASGPR-HA1 caused HA1250-266 specificity, HA1256-272 specificity and HA1262-278 specific CD4+ T cell reaction for the production of IFNg. Interestingly, anti-DC-ASGPR-HA1 produced a greater number of HA1-specific CD4+ T cells expressing IL-10 compared to anti-LOX-1-HA1 (Fig. 5B). HA1250-266-specific CD4+ T cells expanded with anti-DC-ASGPR-HA1 secreted higher levels of IL-10 (p<0.02) and lower compared to CD4+ T cells expanded with anti-LOX-1-HA1 Content of IFNg (p < 0.02) (Fig. 5F). Studies using cells from three other healthy donors also showed that anti-DC-ASGPR-HA1 enhanced the CD4+ T cell response to IL-10 production (Figure 6). Only lower levels of IL-2 and other cytokines were detected (IL-4 and IL-5 were less than 50 159403.doc -23-201219053 pg/ml, not shown). PMA/ionomycin stimulates immune-incompetent Thl cells to produce IFNg18. Figure 5D shows that most IL-10 producing CD4+ T cells exhibited IFNg when stimulated with PMA/ionomycin (Fig. 5D), confirming that it was derived from Thl6. It does not exhibit Foxp3 (Figure 7), but greater than 60% exhibits CTLA-4. The smaller portion (approximately equal to 30%) exhibits a lower level of PD-1. In conclusion, HA1 delivered to DC via DC-ASGPR caused an increase in the HA1-specific Foxp3-CD4+ T cell response from IL-derived IL-10. The HA1 derived peptides determined in this study are summarized in Table 1. Table 1: Predicted binding scores for the corresponding anthraquinone HLA of the HA-1 derived peptide and the tested healthy donor. Donor class II type Peptide binding fraction (ABR score) tested in this study Donor HLA-DRB01-13 HA 150-266 LEPGDTIIFEANGNLIA (SEQ ID NO: 1) HA 156-272 IIFEANGNLIAPWYAFA (SEQ ID NO: 2) HA 162-278 GNLIAPWYAFALSRGFG (SEQ ID NO: 3) (1000000) (9879.0) (83821.7) HLA-DRB3* NA ΝΑ HLA-DQB1*06 NA ΝΑ Donor 1 HLA-DRB01-07 HA 126-142 SSFERFEIFPKESSWPN (SEQ ID NO :4) HA 150-266 LEPGDTIIFEANGNLIA (SEQ ID NO: 5) (5796.6) (1000000) HLA-DRB0M5 HA 126-i42 SSFERFEIFPKESSWPN (SEQ ID NO: 6) HA 150-266 LEPGDTIIFEANGNLIA (SEQ ID NO: 7) (461455.1 (1000000) HLA-DRB5* HA 150-266 LEPGDTIIFEANGNLIA (SEQ ID NO: 8) (1000000) HLA-DRB4* NA ΝΑ HLA-DQB1* NA ΝΑ Donor 2 HLA-DRB01-01 HAl 186.202EKEVLVLWGVHHPPNIG (SEQ ID NO :9) HAl215.23i VSVVSSHYSRRFTPEIA (SEQ ID NO: 10) (261.9) (1109.7) HLA-DRB01-08 HA 1,86.202 EKEVLVLWGVHHPPNIG (SEQ ID NO: 11) HA 1215.231 VSVVSSHYSRRFTPEIA (SEQ ID NO: 12) (2634.8) (30673.0) HLA-DQB1* NA ΝΑ Donor 3 HLA-DRB01-07 HAl 126-i42 SSFERFEIFPKESSWPN (SEQ ID NO: 13) HA 1274-290 SRGFGSGIITSNAP MDE (SEQ ID NO: 14) (5796.6) (2723.8) HLA-DRB01-11 HAli2〇.136EQLSSVSSFERFEIFPK (SEQ ID NO: 15) HA 1274.290 SRGFGSGIITSNAPMDE (SEQ ID NO: 16) (18064.8) (85130.7) HLA-DRB3* NA ΝΑ HLA-DRB4* NA ΝΑ HLA-DQB1* NA ΝΑ 1. The peptide is predicted by a network algorithm: (http://tools.immuneepitope.org/ analyze/cgi-bin/mhc_II_binding.py). Maximum and minimum ABR score 159403.doc -24-

S 201219053 is 1000000 and 0. 2. NA: The algorithm is not available in the algorithm. The inventors then tested whether the autoantigen delivered to DC via DC-ASGPR would produce antigen-specific 1L_10 Treg. In fact, 'loading anti-DC-eight 8 〇卩 11-? 8 八 〇 (^ makes the production of 11 > -10 - 8 eight specifics 〇 4 +1 \ cell response enhancement (Figure 5E, Figure 5F and Figure 8 In contrast, 'CD4+ T cells induced with anti-LOX-1-PSA secrete higher levels of ΙΡΝγ than those induced with anti-DC-ASGPR-PSA (Fig. 5F). After using PMA/ionomycin During stimulation, most of the PSA-specific CD4+ sputum cells producing IL-10 exhibited IFNy (Fig. 5G). It also exhibited CTLA-4, but not Foxp3 (Fig. 9). The inventors further tested DC-SIGN12, 19 and Whether Dectin-12Q can also induce IL-10 Treg (Fig. 10A to Fig. 10C). Production of mAb (anti-Dectin-1: 15E2 IgG2a and anti-DC-SIGN: 20B3 IgG1) and fusion with PSA as anti-DC-ASGPR-PSA ( Not shown. Anti-Dectin-1-PSA and anti-DC-SIGN-PSA induced PSA-specific CD4+ T cells producing IFNY, but did not significantly increase CD4+ T cells producing IL-10. In summary, via DC-ASGPR target The PSA to DC induces antigen-specific IL_i〇Treg. The PSA-derived peptides determined in this study are summarized in Table 2. After loading anti-DC-ASGPR-HA1, DC (especially IFNDC) is associated with monocytes and b-cells. Than manufacturing iFNy CD4+ T cells were more efficient in both amplification of CD4+ T cells producing IL-1 ( (Fig. 11A to UD). DCs loaded with peptides were also more effective in stimulating T than other APCs loaded with the same peptide. The cells express cytokines (Fig. 12A and Fig. 12B).

The inhibitory function of CD4+ T cells was assessed in allogeneic and HA1 specific settings. CFSE low CD4+ T cells were sorted from co-cultures of T cells with IFNDC loaded with anti-DC-ASGPR-PSA or anti-LOX-1-PSA 159403.doc-25·201219053. A growing number of selected sputum cells are added to the culture of autologous IFNDC and allogeneic CD4+ sputum cells. CD4+ Τ cells induced by anti-DC-ASGPR-PSA inhibited allogeneic CD4+ Τ cell proliferation in a dose-dependent manner (Fig. 13Α left panel), compared with CD4+ Τ cells with anti-LOX-1-PSA Significant inhibition of allogeneic sputum cell proliferation (Figure 13 Α right panel). Neutralization of IL-10 reduces the inhibition of proliferation of allogeneic CD4+ T cells by anti-DC-ASGPR-PSA-induced T cells (Fig. 13B)»Initiated by anti-DC-ASGPR-HA1 but not by anti-LOX-1-HA1 CFSE low CD4+ T cells were also able to inhibit autologous T cell proliferation in trans-well (Fig. 13C). The IFNy expression induced by DCs loaded with HA1-derived HA1250-266 was also inhibited by CFSE low CD4+ T cells expanded with anti-DC-ASGPR-HA1 (left panel of Figure 13D). Table 2. Predicted binding scores for PSA-derived peptides and corresponding Class II HLAs of healthy donors tested in this study. Donor Type II Type The peptide/binding fraction tested in this study was restricted by class II MHC of the donor tested in this study. Peptide 1 HLA-DRB01- PSA30-44 HLA-DRB01-1501 Restriction Peptide: 1501 ECEKHSQPWQVLVAS (SEQ ID NO: 17) / (983.6) PSA102-116 DMSLLKNRPLRPGDD (SEQ ID NO: 18) / (7.3) (Clin Cancer Res 2005; 11 (8) 2853-2861) PSA] 62-176 PSA171., 9 〇lqcvdlhvisndvcaqvhpq PEEFLTPKKLQCVDL (SEQ ID NO: 19) / (1000000.0) (SEQ ID NO: 21) PSAs2-96 PSA221.240 GVLQGITSWGSEPCALPERP HPEDTGQVFQVSHSF (SEQ ID NO: 20) / (397.1) (SEQ ID NO; 22) HLA-DRB1 *14 NA/^A HLA-DQB1* NA/NA Donor 2 HLA-DRB01-11 PSA58.72 QWVLTAAHCIRNKSV (SEQ ID NO: 23) / (14859.9) -26- 159403.doc 201219053 HLA-DRB5*

HLA-DRB3* _HLA-DRB16 donor 3~HLA-DRB01-15XX

HLA-DRB01- 07XX HLA-DRB5* PSA78-92 HSLFHPEDTGQVFQV (SEQ ID NO: 24) / (3293.1) PSAi46_i6 〇 PALGTTCYASGWGSI (SEQ ID NO: 25) / (1000000.0) PSA 178-192 VISNDVCAQVHPQKV (SEQ ID NO: 26) /(130615.6) PSA226-240 ITSWGSEPCALPERP(SEQ ID NO:27)/(871249.4) PSA58-72 QWVLTAAHCIRNKSV(SEQ ID NO:28)/(146.1) PSA78.92 HSLFHPEDTGQVFQV(SEQ ID N0:29)/(10000_) PSA丨你(10) PALGTTCYASGWGSI(SEQ ID N0:30)/(1000000.0) PSA|78-|92 VISNDVCAQVHPQKV(SEQ ID NO:31)/(6707.6) PSA226-240 ITSWGSEPCALPERP(SEQ ID N0:32)/(1000000.0) NA/ NA _NA/NA_ PSAjo^ CGGVLVHPQWVLTAA (SEQ ID NO: 33) / (356.0) PSA62.76 TAAHCIRNKSVILLG (SEQ ID NO: 34) / (1.4) PSA66.8 〇 CIRNKSVILLGRHSL (SEQ ID NO: 35) / (1.4) PSAi46 ]60 PALGTTCYASGWGSI (SEQ ID NO: 36) / (1071.3) PSA166-180 LTPKKLQCVDLHVIS (SEQ ID NO: 37) / (438254.3) PSA50.64 CGGVLVHPQWVLTAA (SEQ ID NO: 38) / (266306. I) PSA62.76 TAAHCIRNKSVILLG (SEQ ID NO: 39) / (14.9) PSA66.8〇CIRNKSVILLGRHSL (SEQ ID NO: 40) / (14.9) PSAi46.|6〇PAUJTTCYAS(3WGSI(SEQ ID NO:41)/(798.0)PSAj66-180 LTPKKLQCVDLHVIS(SEQ ID NO: 42) / (183.7) PSA5 〇 ^ 4 CGGVLVHPQWVL TAA (SEQ ID NO: 43) / (6279.2) PSA62.76 TAAHCIRNKSVILLG (SEQ ID NO: 44) / (434.6) PSA^so CIRNKS VILLGRHSL (SEQ ID NO: 45) / (1.5) HLA-DRB4 expression donor (Clin Exp Immunol 1998;114:166-172) PSA49.63 ILLGRMSLFMPEDTG (SEQ ID NO:48) PSA64.78 QVFQVSHSFPHPLYD (SEQ ID NO:49) PSA95.i〇9 NDLMLLRLSEPAELT (SEQ ID NO:50) PSA 丨22]36PALGTTCVASGMGSI (SE(5 ID N0:51) PSA,34-I48 GSIEPEEFLTPDQMK(SEQ id NO:52) PSAI48.|6〇KKLQCVQLHVISM(SEQ id NO:53) PSA,94.2〇8GPLVCNGVLQGITSM(SEQ id NO:54) PSA200.214 GVLQGITSMGSEPCA (SEQ id NO: 55) 159403.doc •27· 201219053 PSAi46-160 PALGTTCYASGWGSI (SEQ ID N0:46)/(1000000.0) PSAi66-I80 LTPKKLQCVDLHVIS (SEQ ID N0:47)/(1408.0)

HLA-DRB4* NA/NA HLA-DQB1* NA/NA_ 1 · Peptides are predicted by a network algorithm: (http://tools.immuneepitope.org/ analyze/cgi-bin/mhc_II_binding.py). The maximum and minimum ABR scores were 1000000 and 0. 2. NA: The algorithm was not available in the algorithm. This inhibition was restored by blocking IL-10 (Fig. 13D). Therefore, IL-10 secreted from CD4+ T cells inhibits T cell proliferation and IFNy expression. Activation of mitogen-activated protein kinase (MAPK) is directly associated with IL-10 induction in DCs21,22. In addition, differences in IL-10 production by DC are associated with the intensity of extracellular signal-related kinase (ERK) activation22. DCs exposed to anti-ASGPR-PSA showed enhanced phosphorylation of ERK and p38 (Fig. 14A). Phosphorylation of ERK/p38 was further demonstrated by flow cytometry (Fig. 14B). Consistently, DCs exposed to anti-DC-ASGPR-PSA but not anti-LOX-1-PSA secreted IL-10 (Fig. 14C). Anti-DC-ASGPR-PSA does not induce IL-27 or ICOSL expression that promotes IL-10 secretion by T cells (not shown). Blocking IL-10 in co-cultures of CD4+ T cells with DCs loaded with anti-DC-ASGPR-PSA resulted in a decreased induction of IL-10 producing PSA-specific CD4+ T cells (Fig. 14D and Fig. 14E). In contrast, the addition of exogenous IL-10 to co-cultures of CD4+ T cells with DCs loaded with anti-LOX-1-PSA enhanced the CD4+ T cell response to IL-10 production (Fig. 14F and Fig. 14G). Neutralization of TGFp also slightly reduced the C〇4+ T cell response of IL-10, but exogenous TGFp did not enhance the CD4+ T cell response of IL-10 (Fig. 15A to Fig. 15C). In summary, we 159403.doc -28· 201219053 data show that PSA is delivered to DC-induced DCs in ERK/p38 phosphorylation and IL-10 via DC-ASGPR but not LOX-1, which promotes induction of IL-10 Treg . Interestingly, Figures 16A and 16B show that the biological function of DC-ASGPR can outperform LOX-1. The antigen delivered to DC via DC-ASGPR and LOX-1 simultaneously reduced the CD4+ T cell response to produce IFNy and enhanced the CD4+ T cell response to produce IL-10. In order to expand the in vitro observation of the present invention, cynomolgus macaques were immunized with the fusion protein. Both anti-LOX-1 and anti-DC-ASGPR mAbs bound to CDllc+ and CD14+ cells but not to CD3+ T cells (Fig. 17A). All animals (12 animals in total: 6 animals per group) were pre-immunized with live influenza virus (H1N1, PR8). Serum showed high levels of HA1-specific IgG (not shown). Four months after initial immunization, anti-LOX-1-HA1 (right arm) and anti-LOX-1-PSA (left arm) or anti-DC-ASGPR-HA1 (right arm) and anti-DC-ASGPR -PSA (left arm) immunizes animals. After three immunizations with recombinant fusion protein, blood was collected as indicated in Figures 17B and 17C. PBMCs from animals immunized with anti-LOX-1-HA1 reacted with the HA1 peptide pool to secrete significantly higher levels of IFNY compared to animals immunized with anti-DC-ASGPR-HA1 (Fig. 17B). In contrast, PBMCs from animals immunized with anti-DC-ASGPR-HA1 reacted with the HA1 peptide to secrete significantly higher levels of IL-10 when compared to those immunized with anti-LOX-1 -HA1. The same observations were made in animals immunized with the PSA fusion protein. The predominantity of the animals immunized with anti-DC-ASGPR-PSA showed a PSA-specific cellular response to IL-10, whereas the dominant ones were inoculated with anti-LOX-1-PSA immunized I59403.doc -29-201219053. Highly produced PS A-specific cellular responses of ΝγΝ (Fig. l7c). For both HA1 and PSA, the peak of the cellular response to IL-10 production was obtained in the first week, but the peak of the cellular response to IFNy production was obtained in the third week after the second booster immunization. Given the critical role of IL-10 Treg derived from Th1 in limiting host immunopathology, m.7, 8'25-29, these findings provide novel therapies for treating human diseases caused by non-recessive inflammatory responses. . The antigen-specific IL1〇Treg is established in vivo as an alternative to current therapeutic approaches such as repetitive peptide immunization 6,3〇 and in vitro produced IL-10 receptive transfer.29 The unique ability of Dc·ASGPR applies to itself. Antigens and foreign antigens, as well as primordial and memory CD4+ T cell responses. Therefore, DC_ASGpR also appears to be a common target for designing vaccines against autoimmune diseases (such as type 2 diabetes and multiple sclerosis) identified by autoantigens. Any of the embodiments discussed in the specification can be practiced with respect to any method, kit, reagent or composition of the invention, and vice versa. Furthermore, the compositions of the invention can be used to achieve the methods of the invention. The specific embodiments are not intended to limit the invention, and the main features of the present month can be used in various embodiments without departing from the scope of the invention. Those skilled in the art will recognize The equivalents of the specific procedures described herein are to be determined by the routine experimentation. The equivalents are considered to be the scope of the invention. All publications and patent applications mentioned in this specification are indicative of the technical skill of those skilled in the art of the invention. All publications and patents 159403.doc 201219053 They are incorporated herein by reference in the form of a reference to the ancients, and the extent of the reference is as follows: 疋 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固 固The scope of the patent and, or the use of the term "including" in this specification, the use of the word "-" may mean "-", but it is also associated with "one or more J, "at least one broadcast" and "king" And the meaning of "one or more" is used. The term "or" as used in the scope of the patent application is used to mean "and/or" unless the description is exclusive or exclusive. Case support refers to the definition of the only option A "and, or". In the whole case, the „, ,约” is used to indicate that the value includes the inherent deviation of the error of the device or method used to determine the value or the study individual. Intermittent storage The words "including", "having", "including" or "including" as used in this specification and the claims are intended to be inclusive or open and do not exclude other elements or methods that are not stated. The term "or a combination thereof" as used herein refers to all permutations and combinations of items listed above the term. For example, "A, B' C or a combination thereof" is intended to include at least one of the following: B, C, AB, AC, BC* ABC, and if the order is significant in a particular situation, it also includes BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continue this example 'clearly including duplicates Appearance - or a combination of multiple items or terms, such as BB, AAA, MB, BBC, AAABCCCC, CBBAAA, CABABBm, should be understood, unless otherwise obvious from the context, usually without limiting the terms or terms in any combination. number. 159403.doc -31- 201219053 In accordance with the present disclosure, all compositions and/or methods disclosed and claimed herein can be made and practiced without undue experimentation. Although the compositions and methods of the present invention have been described in accordance with the preferred embodiments, it will be apparent to those skilled in the art that Or a method, and in the sequence of steps or steps of the methods described herein, a certain variation can be applied. All such similar substitutes and modifications, which are apparent to those skilled in the art, are considered to be within the scope of the spirit, scope and concept of the invention as defined by the appended claims. REFERENCES US Patent Application Publication No. 20080206262: dgewb ί/ζαί Engage Antigen-Presenting Cells through Dendritic Cell Asialoglycoprotein Receptor (DC-ASGPR). US Patent Application Publication No. 20080241170: Facchw on Targeting Antigen to DCIR Expressed on Antigen-Presenting Cells o 1. Trinchieri, G. Interleukin-10 Production by Effector T Cells: Thl Cells Show Self Control. J Exp Med 204, 239- 43 (2007). 2. Groux, H et al. €€04+丁-匚61181^361;1111^1^3入111^611_ Specific T-Cell Responses and Prevents Colitis. Nature 389, 737-42 (1997) » 3. Moore, KW, De Waal Malefyt, R., Coffman, RL & O'Garra, A. Interleukin-1 0 and the Interleukin-10 Receptor. 159403.doc •32· 201219053

Annu Rev Immunol 19, 683-765 (2001) 〇 4. Haringer, B., Lozza, L., Steckel, B. & Geginat, J. Identification and Characterization of IL-10/IFN-Gamma-Producing Effector-Like T Cells with Regulatory Function in Human Blood. J Exp Med 206,1009-17 (2009) 0 5. Anderson, CF, Oukka, M., Kuchroo, VJ & Sacks, D. CD4(+)CD25(-)Foxp3 (-) Thl Cells are the Source of IL-10-Mediated Immune Suppression in Chronic Cutaneous Leishmaniasis. J Exp Med 204, 285-97 (2007) o 6. Gabrysova, L. et al. Negative Feedback Control of the Autoimmune Response through Antigen -Induced Differentiation of IL-10-Secreting Thl Cells. J Exp Med 206, 1755-67 (2009). 7. Jankovic, D. et al. Conventional T-Bet(+)Foxp3(-) Thl Cells are the Major Source of Host-Protective Regulatory IL-10 During Intracellular Protozoan Infection. J Exp Med 204, 273-83 (2007) ° 8.0'Garra, A. & Vieira, P. T(H)1 Cells Control Themselves by Producing Interleukin-10. Nat Rev Immunol 7, 425-8 (2007)〇9. Banchereau, J. & Steinman, RM Dendritic Nature 392, 245-52 (1998) ° 10. Brown, GD Dectin-1: A Signalling Non-TLR Pattern-Re cognition Receptor. Nat Rev Immunol 6,33-43 159403.doc -33 - 201219053 (2006). 11. Figdor, CG, Van Kooyk, Y. & Adema, GJ C-Type Lectin Receptors on Dendritic Cells and Langerhans Cells. Nat Rev Immunol 2, 77-84 (2002) ° 12. Geijtenbeek, TB & Gringhuis, SI Signaling through C-Type Lectin Receptors: Shaping Immune Responses. Nat Rev Immunol 9, 465-79 (2009) ° 13. Akira, S., Takeda, K. & Kaisho, T. Toll-Like Receptors: Critical Proteins Linking Innate And Acquired Immunity. Nat Immunol 2, 675-80 (2001) 〇 14. Valladeau, J. et al. Immature Human Dendritic Cells Express Asialoglycoprotein Receptor Isoforms for Efficient Receptor-Mediated Endocytosis. J Immunol 167, 5 767-74 (2001). 1 5. Delneste, Y. et al. Involvement Of LOX-1 In Dendritic

Cell-Mediated Antigen Cross-Presentation. Immunity 17, 353-62 (2002) ° 16. Geijtenbeek, T. B. SIGN to Suppress Dendritic Cell Function. J Exp Med 197, 7-17 (2003). 17. Reddy, Μ. P. et al. Elimination of Fc Receptor-Dependent Effector Functions of A Modified IGg4 Monoclonal Antibody to Human CD4. J Immunol 164, 1925-33 (2000)° 159403.doc •34·201219053 18. Macian, F. et al. Transcriptional Mechanisms Underlying Lymphocyte Tolerance. Cell 109, 719-31 (2002). 19. Smits, Η. H. et al. Selective Probiotic Bacteria Induce IL-10-Producing Regulatory T Cells In Vitro by Modulating Dendritic Cell Function through Dendritic Cell-Specific Intercellular Adhesion Molecule 3-Grabbing Nonintegrin. J Allergy Clin Immunol 115, 1260- 7 (2005) ° 20. Dillon, S. et al. Yeast Zymosan, A Stimulus for TLR2 and Dectin-1, Induces Regulatory Antigen-Presenting Cells and Immunological Tolerance. J Clin Invest 116, 916-28 (2006). 21. Agrawal, A., Dillon, S., Denning, TL & Pulendran, B. ERK1 -/- Mice Exhibit Thl Cell Polarization and Increased Susceptibility to Experimental Autoimmune Encephalomyelitis. J Immunol 176,5788-96 (2006) o 22 Kaiser, F. et al. TPL-2 Negatively Regulates

Interferon-Beta Production in Macrophages and Myeloid Dendritic Cells. J Exp Med 206, 1863-71 (2009). 23. Ito, T. et al. Plasmacytoid Dendritic Cells Prime IL-10-Producing T Regulatory Cells by Inducible Costimulator Ligand. J Exp Med 204, 105-15 (2007).

24. Iwasaki, A., Torres, C. A. T., Ohashi, P. S., Robinson, H. L. & Barber, B. H. The Dominant Role of Bone Marrow-Derived Cells in CTL Induction Following Plasmid DNA 159403.doc -35- 201219053

Immunization at Different Sites 1. J. Immunol. 159, 1-4 (1997). 25. Gazzinelli, R. Τ· et al. In the Absence of Endogenous IL-10, Mice Acutely Infected with Toxoplasma gondii Succumb to A Lethal Immune Response Dependent on CD4+ T Cells and Accompanied by Overproduction of IL-12, IFN-Gamma and TNF -Alpha. J Immunol 157, 798-805 (1996) ° 26. Allan, SE et al. CD4+ T-Regulatory Cells: Toward Therapy for Human Diseases. Immunol Rev 223, 391-421 (2008). 27. Burkhart, C., Liu, GY, Anderton, SM, Metzler, B. & Wraith, DC Peptide-Induced T Cell Regulation of Experimental Autoimmune Encephalomyelitis: A Role For IL-10. Int Immunol 11, 1625-34 ( 1999). 28. O'Neill, EJ, Day, MJ & Wraith, DC IL-10 is Essential for Disease Protection Following Intranasal Peptide Administration in the C57BL/6 Model of EAE. J Neuroimmunol 178,1-8 (2006) ° 29. Roncarolo, MG & Battaglia, M. Regulatory T-Cell

Immunotherapy for Tolerance to Self Antigens and Alloantigens in Humans. Nat Rev Immunol 7, 585-98 (2007). 30. Sundstedt, A., O'Neill, E. J., Nicolson, K. S. & Wraith, D. C. Role for IL-10 in Suppression Mediated by -36- 159403.doc s 201219053

Peptide-Induced Regulatory T Cells In Vivo. J Immunol 170, 1240-8 (2003). [Simplified Schematic] Figure 1A and Figure IB: LOX-1 and DC-ASGPR in vivo and in vitro DC: Figure 1A Blood-medullary DC (mDC) (Lin-HLA-DR+CDllc+ CD123-), plasma cells The appearance of LOX-1 (top panel) and DC-ASGPR (bottom panel) on the surface of DCs (PDC) (Lin-HLA-DR+CDllc-CD123+), CD14+ monocytes and CD3+ T cells, Figure 1B single core Surface expression of LOX-1 (top panel) and DC-ASGPR (bottom panel) on cell-derived IFNDC and IL-4DC; Figure 2A to 2D: Immunofluorescence staining of healthy human skin sections: Figure 2A for fluorescence Labeled DAPI, anti-HLA-DR, anti-CDlc and anti-LOX-1 mAb stained the ice sheet (5 mm), Figure 2B with fluorescently labeled DAPI, anti-HLA-DR, anti-CDlc and anti-DC-ASGPR mAb, Figure 2C DAPI and control mAb, Figure 2D is a summary of data generated from skin from two healthy donors. Each point represents data from one section; Figure 3A to Figure 3E: Recombinant fusion protein of anti-DC surface receptor mAb and antigen: Figure 3A Reductive SDS-PAGE analysis of purified recombinant fusion protein (lane 1: molecular weight marker, Lane 2: anti-LOX-1 'lane 3: anti-DC-ASGPR, lane 4: control IgG4-HAl, lane 5 · anti-LOX-1-HA1, lane 6: anti-DC-ASGPR-HA1, lane 7: control IgG4 -PSA, Lane 8: anti-LOX-1-PSA and Lane 9: anti-DC-ASGPR-PSA, Figure 3B and Figure 3C HA1 and PSA fusion protein binding to IFNDC, Figure 3D and Figure 3E recombined by loading 1 mg/ml CD4+ T cell proliferation induced by fusion protein, 1 mg/ml mAb or unloaded IFNDC. Data represent five independent experiments using 159403.doc -37-201219053 cells from different healthy donors; Figure 4: via LOX Influenza virus HA1, which is delivered to DC by DC-ASGPR, causes HA1-specific CD4+ T cell response to produce IFNg (interferon gamma). Use 1 pg/ml anti-LOX-1-HA1 or anti-DC-ASGPR-HA1 The fusion protein was loaded with 5xl03 IFNDC and subsequently incubated overnight with CFSE-labeled purified autologous CD4+ T cells (2χ105) Raised for 7 days. In the presence of BFA, the peptide pool (0.1 μΜ for each peptide, 11 to 12 peptides per cluster, and 17 amino acid overlapping 17-mers) was used to re-stimulate CD4+ sputum cells, followed by CD4 and intracellular IFNg stained cells. Two independent studies showed similar results; Figure 5A to Figure 5G: Antigen-specific CD4+ T cells from IL-producing IL-10 produced by DC-ASGPR-targeted DCs: Figure 5A by load 1 The frequency of HA1-derived peptide-specific CD4+ T cells expressing IFNy caused by IFNDC of pg/ml recombinant fusion protein, and Figure 5B shows the frequency of HA1-derived peptide-specific CD4+ T cells of IL-10. In Figures 5A and 5B The peptides HA1280, 296 were tested as negative controls. Four independent studies in Figures 5A and 5B showed similar results, and the cytokine content in the culture supernatant of Figure 5C. Each line represents data from a single operation, and Figure 5D uses PMA ( 50 ng/ml)/inonomycin (1 pg/ml) stimulated CD4+ T cells expanded by DCs loaded with anti-LOX-1-HA 1 or anti-DC-ASGPR-HA1 for 4 hours, followed by Intracellular IFNy and IL-10 staining, Figure 5E is derived from PSA derived from IFNDC loaded with 1 pg/ml recombinant fusion protein The frequency of CD4+ T cells producing IL-10, and the cytokine content in the culture supernatant of Fig. 5F. The error bars represent the mean soil standard deviation of the three replicates. Three independent studies were performed using 59 peptides derived from PSA. Only show positive reactions in all three experiments

159403.doc -38- S peptide of 201219053. Control peptides indicating each donor, Figure 5G was stimulated with PMA (50 ng/ml)/ionomycin (1 pg/ml) by DCs loaded with anti-LOX-1-HA1 or anti-DC-ASGPR-HA1 CD4+ T cells were stained for 4 hours, followed by intracellular IFNy and IL-10 staining. More than four independent studies in Figures 5D and 5G showed similar results; Figure 6: HA1 delivered to DC via DC-ASGPR enhanced IL-10-specific CD4+ T cell response to IL-10: 1 mg/ml anti-LOX- 1-H1 or anti-DC-ASGPR-HA1 was loaded with 5 x 103 DCs and incubated overnight. Autologous CD4+ T cells (2χ105 cells) were co-cultured for 7 days. On day 9, CD4+ T cells were restimulated for 48 hours with IFNDC loaded with 1 mM peptide. The IL-10 content in the culture supernatant was measured by Luminex. Three studies were performed using the same peptide bundle in three replicate assays. The value is the average of 12 data points. Gray indicates the mean soil standard deviation obtained without peptide; Figure 7: Performance of Foxp3, PD-1 and CTLA-4 on HAI250-266 specific CD4+ T cells producing IL-10; Figure 8: Via DC - APSPR-delivered PSA to enhance the PSA-specific CD4+ T cell response of IL-10: loading with 1 mg/ml anti-LOX-1-?3 VIII or anti-00 八80?1^8 八5)< 103 0 (:, and incubated overnight. Autologous CD4+ T cells (2x105 cells) were co-cultured for 7 days. On day 9, CD4+ T cells were restimulated with mM DCs loaded with 1 mM peptide for 48 hours. The contents of IFNg and IL-10 in the culture supernatant. The error bars represent the mean standard deviation of the three replicates. Three independent studies were performed with 59 peptides derived from PS A. Only the results were positive in all three experiments. Peptide of reaction. Control peptide indicating each donor; 159403.doc -39· 201219053 Figure 9: Production of 1!^10? 8 八 3 0-44 specific (8) 匸〇 4+1\ cells on Foxp3, PD-1 and CTL A-4 performance. Three independent studies yielded similar data; Figure 10A to Figure 10C: DC-ASGPR produces specific work for the production of IL-10 antigen-specific CD4+ T cells 5χ103 IFNDCs were loaded with 1 mg/ml PSA recombinant fusion protein or mixed peptide (10 mM). After incubation overnight, CFSE-labeled autologous CD4+ T cells (2χ105 cells) were co-cultured for 7 days. Figure 10Α Load 1 mM IFNDC of peptide PSA102_ii6 or control peptide (PSA82.96) re-stimulated CD4+ T cells. After 48 hours, IL-10 and IFNg in culture supernatants were measured by Luminex. Data from five independent studies were pooled. 10B Re-stimulates CD4+ T cells with 1 mM peptide PSA102_116 or control peptide (PSA82-96) for staining intracellular IL-10 in the presence of BFA, a summary of three independent studies of Figure 10C Figure 10B. Student's t-test calculates the P value; Figure 11A to Figure 11D: IFNDC produces antigen-specific CD4+ T cells secreting IFNg and IL-10 with higher potency than other APCs. Recombinant fusion protein with 1 mg/ml HA1 Load 5χ103 APCs. After incubation overnight, CFSE-labeled autologous CD4+ T cells (2><105) were co-cultured for 7 days. On day 9, in the presence of brefeldin A, load 1 was used. IFNDC re-stimulated CD4+ T cells with mM HA 1 250_266 or control peptide HA 1 280_296. The cells were subsequently stained for intracellular IL-1 0 and IFNg. Data from four independent studies were compiled. P values were calculated using the Sturden t test; Figures 12A and 12B: Peptide-loaded IFNDCs were more effective in inducing intracellular IFNg and IL-expression of CD4+ T cells than other APCs loaded with the same peptide.

159403.doc - 40 - S 201219053 10. 5 χ 103 IFNDCs were loaded with 1 mg/ml anti-LOX-1-HA1 (Fig. 12A) or anti-DC-ASGPR-HA1 (Fig. 12B). After incubation overnight, CFSE-labeled autologous CD4+ sputum cells (2 x 105) were co-cultured for 7 days. On day 10, CD4+ T cells were restimulated with 5 mM ΗΑ 1 250-266 or the control peptide HAl28〇-296 iAPC in the presence of brefeldin. The cells were subsequently stained for intracellular IL-10 and IFNg. Data from four independent (Figure 12A) and three (Figure 12B) independent studies were summarized. P value was calculated by the Sturden t test; Figure 13A to Figure 13D: CD4+ T cells produced by anti-DC-ASGPR-PSA and anti-DC-ASGPR-HA 1 inhibited the production of IFNg allogeneic and HA 1 specific CD4+ T Cellular Reaction: Figure 13A shows different numbers of FACS-selected CFSE low-effect cells and loaded anti-DC-ASGPR-PSA produced with anti-DC-ASGPR-PSA (left panel) or anti-LOX-1-PSA (right panel) IFNDC was co-cultured with newly purified allogeneic CD4+ T cells. The proliferation of allogeneic CD4+ T cells was assessed by measuring 3[H] thymidine uptake, and Figure 13B was subjected to FACS with anti-DC-ASGPR-PSA in the presence of anti-IL-10 and anti-IL-10R antibodies. The sorted CFSE low effector cells were co-cultured with anti-DC-ASGPR-loaded IFNDC and newly purified allogeneic CD4+ T cells. Effector cells: The ratio of responding cells is 1:2. The proliferation of allogeneic 0〇4+!\ cells was assessed by measuring the amount of 3[H] thymidine uptake. Three independent studies of four replicates in Figure 13 and Figure 138 showed similar results. Representative data from one run are shown, and Figure 13C co-cultures FACS-selected CFSE low effector cells produced with anti-DC-ASGPR-HA1 with loaded HAl 250_266iIFNDC in the upper well. The newly purified CFSE-labeled CD4+ T cells were co-cultured with anti-LOX-1-HA1-loaded IFNDC in the lower trans-well plate 159403.doc-41 _ 201219053. On day 6, proliferation of CD4+ T cells in the lower wells was assessed by measuring CFSE dilutions. Anti-IL-10/IL-10R or control IgG was added to the lower wells, and on day 10, Fig. 13D, the frequency of CD4+ T cells expressing ΙΡΝγ was measured. Data from five independent studies are summarized in the right panel; Figure 14A to Figure 14G: Induction of IL-10 in DC via DC-ASGPR-targeting DC antigen, and this IL-10 promotes antigen specificity for IL-10 production Production of CD4+ T cells· Figure 14A after loading IFNDC 1 with 1 mg/ml recombinant protein (anti-DC-ASGPR-PSA and anti-LOX-1-PSA) or 10 mg/ml zymosan (Zymosan) for 5 minutes, The cell lysate ERK, phospho-ERK, p38, and phospho-p38 were analyzed by Western blotting. Figure 14B ERR/p38 phosphorylation by flow cytometry analysis, Figure 14C loading 1 mg/ml anti-LOX-1-PSA or IL-10 content in culture supernatant of IFNDC (lxl05) against DC-ASGPR-PSA. The cells were incubated for 24 hours. Four independent studies showed similar results, Figure 14D IL-10 blockade: in the presence of control IgG or anti-IL-10/IL-10R antibodies, the purified CD4+ T cells were co-administered with anti-DC-ASGPR-PSA-loaded IFNDC to cultivate. On day 9, cells were re-stimulated with the indicated peptide (1 mM) and stained for intracellular IL-10, Figure 14E is a summary of data from four independent studies of Figure 14D, Figure 14F exogenous IL-10: at 100 CD4+ T cells were co-cultured with IFNDC loaded against LOX-1-PSA in the presence or absence of ng/ml IL-10. On day 9, cells were stained for intracellular IL-10, and Figure 14G is a summary of data from five independent studies of Figure 14F. Each point represents data from an independent study. P values were obtained by the Studden t test; Figures 15A to 15C: Blocking TGFbl reduced the antigen-specific CD4+ T cell response of IL-10 production, but the addition of exogenous TGFb 1 did not enhance the anti-

159403.doc -42- S 201219053 should be. 5χ103 IFNDCs were loaded with 1 mg/ml anti-DC-ASGPR-PSA. After overnight incubation, CFSE-labeled autologous CD4+ T cells (2χ105) were co-cultured for 7 days in the presence of 20 mg/ml anti-TGFbl and anti-TGFb receptors. On day 9, in brefeldin A In the presence of CD4+ T cells, the IFNDC loaded with 1 mM peptide was re-stimulated. Figure 15A then stains cells for intracellular IL-10. Figure 15B summarizes data from three independent studies. Figure 15C co-cultures anti-LOX-1-PSA-loaded IFNDC with CFSE-labeled CD4+ T cells in the presence of 1 ng/ml TGFbl. Cells were stained for intracellular IL-10. Figure 16A and Figure 16B: DC-ASGPR signal can be stronger than LOX-1 signal: Figure 16A was co-cultured with purified CD4+ T cells (2χ105) with 5χ103 IFNDC loaded with 1 mg/ml fusion protein for 10 days. CD4+ T cells were restimulated with 1 mM ΗΑ 1250-266 or control peptide ΗΑ128〇-296. IFNg and IL-10 levels were measured by Luminex. Figure 16B co-cultures 5 x 103 IFNDCs loaded with 1 mg/ml PSA fusion protein with purified CD4+ T cells (2 χ 105 cells) for 1 day. CD4+ T cells were restimulated with 1 mM PSA30-44 or a control peptide. Each point represents data from an independent study. P value was calculated by the Studden's t test; and Figures 17A to 17C: Non-human primates immunized with antigens (HA1 and PSA) carried by anti-DC-ASGPR mAb caused the production of IL-10 antigen Data of specific cell reaction. Figure 17A shows the amount of expression of 1 > 0 and -1 and 0 (: - 八80?11) in CDllc+, 0014+ and 003+ cells of cynomolgus monkey PBMC. Figure 17B and Figure 17C will be used. Live influenza virus (H1N1, A/PR8/38) intradermal initial immunization and booster immunization of 12 animals were divided into two groups (6 159403.doc •43·201219053 animals/group). One group of animals used 250 mg Anti-LOX-1-HA1 (right arm) and 250 mg anti-LOX-1-PSA (left arm) were immunized intradermally. Another group of animals used 250 mg anti-DC-ASGPR-HA1 (right arm) and 25 0 mg anti- DC-ASGPR-PSA (left arm) was immunized intradermally. After a second booster immunization with recombinant fusion protein, PBMC was re-stimulated with a peptide pool of 25 mM HA1 (Fig. 17B) or PSA (Fig. 17C), respectively. 2χ105 cells/200 ml medium/well in the well plate. IFNg and IL-1 0 in the culture supernatant were measured by ELISA. The peptide pool of HIVgag was used as a control. Figure 1 7B and Figure 1 7C Displayed value According to the value after the value is also used staphylococcal enterotoxin B (staphylococcal enterotoxin B) (1 0 mg / ml) stimulated cells (FIG. 1 7B, and FIG. 17C, right). Statistically significant by ANOVA test.

159403.doc -44- S 201219053 Sequence Listing <110> American Baylor Research Association <120> Targeting human dendritic cells via DC·sialyl glycoprotein receptor to produce IL-10 regulatory T cells

<130> BHCS: 2458TW <140> 100137170 <141> 2011-10-13 <150> 61/392,910 <151> 2010-10-13 <160> 55 <170> patentin version 3.5 <;210> 1 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 1

Leu Glu Pro Gly Asp Thr lie lie Phe Glu Ala Asn Gly Asn Leu lie 15 10 15

Ala <210> 2 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 2 lie lie Phe Glu Ala Asn Gly Asn Leu lie Ala Pro Trp Tyr Ala Phe 15 10 15

Ala <210> 3 <211> 17

<212> PRT 159403 - Sequence Listing.doc 201219053 <213> Artificial Sequence <220><223> Synthetic Peptide <400>

Gly Asn Leu lie Ala Pro Trp Tyr Ala Phe Ala Leu ser Arg Gly Phe 1 5 10 15

Gly <210> 4 <211><212><213> 17 PRT artificial sequence <220><223> Synthetic peptide. <400> 4

Ser ser Phe Glu Arg Phe Glu lie Phe Pro Lys Glu ser ser Trp Pro 1 Asn 5 10 15 <210><211><212><213> 5 17 PRT artificial sequence <220><223> Synthetic peptides. <400> 5

Leu Glu Pro Gly Asp Thr lie lie Phe Glu Ala Asn Gly Asn Leu lie 15 10 15

Ala <210><211><212><213> 6 17 PRT artificial sequence <220><223> Synthetic peptide. <400> 6 ser ser Phe Glu Arg Phe Glu lie Phe Pro Lys Glu ser ser Trp Pro 15 10 15 159403 - Sequence Listing.doc 201219053

Asn <210> 7 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 7

Leu Glu Pro Gly Asp Thr lie He Phe Glu Ala Asn Gly Asn Leu lie 15 10 15

Ala <210> 8 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 8

Gly Asn Leu lie 15

Leu Glu Pro Gly Asp Thr lie lie Phe Glu Ala Asn 1 5 10

Ala <210> 9 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide <400>

Glu Lys Glu val Leu val Leu Trp Gly val His His Pro Pro Asn lie 15 10 15

Gly <210> 10 <211> 17

<212> PRT 159403. Sequence Listing.doc 201219053 <213> Artificial Sequence <220><223> Synthetic peptide. <400> 10 val Ser Val val ser ser His Tyr Ser Arg Arg Phe Thr Pro Glu lie 15 10 15

Ala <210> 11 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide e <400>

Glu Lys Glu Val Leu val Leu Trp Gly val His His Pro Pro Asn lie 15 10 15

Gly <210> 12 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 12 val ser val Val ser ser His Tyr ser Arg Arg PheThr pro Glu lie 15 10 15

Ala <210> 13 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 13

Ser Ser Phe Glu Arg Phe Glu lie Phe pro Lys Glu ser ser Trp Pro 15 10 15 -4- 159403 - Sequence Listing.doc s 201219053

Asn <210> 14 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide e <400> 14 ser Arg Gly Phe Gly ser Gly lie lie Thr ser Asn Ala Pro Met Asp 15 10 15

Glu <210> 15 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 15

Glu Gin Leu Ser Ser val ser Ser Phe Glu Arg Phe Glu lie Phe Pro 15 10 15

Lys <210> 16 <211> 17 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 16 ser Arg Gly Phe Gly ser Gly lie lie Thr ser Asn Ala pro Met Asp 1 5 10 15

Glu <210> 17

<211> 15 <212> PRT 159403 - Sequence Listing.doc 201219053 <213> Artificial sequence <220><223> Synthetic peptide. <400> 17 Glu cys Glu Lys His ser Gin Pro Trp Gin val Leu val Ala 15 10 ser 15 <210> 18 <2U> 15 <212> PRT <213> Manual sequence <220><223> Synthetic peptide. <400> 18 Asp Met Ser Leu Leu Lys Asn Arg Phe Leu Arg Pro Gly Asp

Asp 15 <210> 19 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 19 pro Glu Glu Phe Leu Thr Pro Lys Lys Leu Gin cys val Asp 1 5 10

Leu 15 <210> 20 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 20

His Pro Glu Asp Thr Gly Gin val Phe Gin val ser His ser 15 10

Phe 15 <210> 21 <211> 20 <212> PRT <213> Artificial sequence <220><223><400> 21 159403 - Sequence Listing.doc · 6 - 201219053

Leu Gin cys Val Asp Leu His val lie Ser Asn Asp Val cys Ala Gin 1 5 10 15 val His Pro Gin 20 <210> 22 <211> 20 <212> PRT <213> Manual Sequence <220><223> Synthetic peptide. <400> 22

Gly val Leu Gin Gly lie Thr Ser Trp Gly ser Glu Pro Cys Ala Leu 15 10 15

Pro Glu Arg Pro 20 <210> 23 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 23

Gin rrp Val Leu Thr Ala Ala His Cys lie Arg Asn Lys Ser Val 1 5 10 15 <210> 24 <211> 15 <212> PRT <213> Artificial Sequence <220><223> Synthetic Peptide . <400> 24

His ser Leu Phe His pro Glu Asp Thr Gly Gin Val Phe Gin val 15 10 15 <210> 25 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 25 159403 - Sequence Listing.doc 201219053 pro Ala Leu <3ly Thr Thr cys Tyr Ala ser Gly Trp Gly ser lie 15 10 15 <210> 26 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 26 vall lie ser Asn Asp val cys Ala Gin val His Pro Gin Lys val 5 10 15 <210><211><212><213><220><223><400&gt 27 15 PRT artificial sequence synthetic peptide 27 lie Thr ser Trp Gly ser Glu Pro Cys Ala Leu Pro Glu Arg Pro 15 10 15 <210><2U><212><213><220><223><400> 28 15 PRT artificial sequence synthetic peptide 28

Gin Trp val Leu Thr Ala Ala His cys lie Arg Asn Lys ser val 15 10 15 <210> 29 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 29

His ser Leu Phe His Pro Glu Asp Thr Gly Gin val Phe Gin val 15 10 15 <210> 30 <211> 15 159403 · Sequence Listing, doc s 201219053 <212> PRT <213> Manual Sequence <220>;<223> Synthetic peptide. <400> 30

Pro Ala Leu Gly Thr Thr cys Tyr Ala ser Gly Trp Gly ser lie 15 10 15 <210> 31 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide e <400> 31

Val lie ser Asn Asp val cys Ala Gin val His Pro Gin Lys val 10 15 <210> 32 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide <400> 32 lie Thr ser Trp Gly ser Glu Pro Cys Ala Leu Pro Glu Arg Pro 15 10 15 <210> B3 <211> 15 <212> PRT <213> Manual sequence <220><223> Synthetic peptides. <400> 33 cys Gly Gly val Leu Val His Pro Gin Trp Val Leu Thr Ala Ala 1 5 10 15 <210> 34 <211> 15 <212> PRT <213> Manual Sequence <220>;223> Synthetic peptide. <400> 34 159403 - Sequence Listing.doc 201219053

Thr Ala Ala His cys lie Arg Asn Lys Ser val lie Leu Leu Gly 15 10 15 <210> 35 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 35 cys lie Arg Asn Lys ser Val lie Leu Leu Gly Arg His ser Leu 15 10 15 <210> 36 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 36 pro Ala Leu Gly Thr Thr cys Tyr Ala ser Gly Trp Gly ser lie 15 10 15 <210> 37 <211> 15 <212> PRT <213> Artificial sequence Synthetic peptide. <220><223><400> 37

Leu Thr Pro Lys Lys Leu Gin cys val Asp Leu His val lie ser 15 10 15 <210> 38 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 38 cys Gly Gly Val Leu Val His Pro Gin Trp val Leu Thr Ala Ala 15 10 15 <210> 39 <211> 15 -] 0· 159403 - Sequence Listing.doc s 201219053 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 39

Thr Ala Ala His cys lie A "g Asn Lys ser val lie Leu Leu Gly 15 10 15 <210> 40 <211> 15 <212> PRT <213> .Artificial sequence <220><223> Synthetic peptide. <400> 40 cys He Arg Asn Lys ser val lie Leu Leu Gly Arg His ser Leu 15 10 15 <210> 41 <211> 15 <212> PRT <213> Artificial sequence <220>;<223> Synthetic peptide. <400> 41

Pro Ala Leu Gly Thr Thr cys Tyr Ala ser Gly Trp Gly ser lie 1 5 10 15 <210> 42 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide . <400> 42

Leu Thr Pro Lys Lys Leu Gin cys Val Asp Leu His val lie ser 15 10 15 <210> 43 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 43 159403 - Sequence Listing.doc -11- 201219053

Cys Gly Gly val

Leu val His Pro Gin Trp val 5 10

Leu Thr Ala Ala 15 <210> 44 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 44

Thr Ala Ala His cys lie Arg Asn Lys Ser val lie Leu Leu Gly 15 10 15 <210> 45 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 45

Cys lie Arg Asn Lys ser val Leu Leu Gly Arg His ser Leu 15 10 15 <210> 46 <2U> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 46

Pro Ala Leu Gly Thr Thr cys Tyr Ala Ser Gly Trp Gly ser lie 1 5 10 15 <210> 47 <211> 15 <212> PRT <213> Artificial Sequence <220><223> Synthetic Peptide . <400> 47

Leu Thr Pro Lys Lys Leu Gin cys val Asp Leu His Val lie ser 15 10 15 <210> 48 <211> 15 •12· 159403 - Sequence Listing.doc 201219053 <212> PRT <213> Artificial Sequence <;220><223> Synthetic peptide. <400> 48 lie Leu Leu Gly Arg Met ser Leu Phe Met Pro Glu Asp Thr Gly 15 10 15 <210> 49 <211> 15 <212> PRT <213> Manual sequence <220><223> Synthetic peptide. <400> 49

Gin val Phe Gin val ser His ser phe Pro His Pro Leu Tyr Asp 15 10 15 <21〇> 50 <211> 15 <212> PRT <213> Artificial sequence <220><223> Peptide. <400> 50

Asn Asp Leu Met Leu Leu Arg Leu ser Glu Pro Ala Glu Leu Thr 15 10 15 <210> 51 <211> 15 <212> PRT <213> Artificial sequence Synthetic peptide. <220><223><400> 51

Pro Ala Leu Gly Thr Thr cys val Ala ser Gly Met Gly ser lie 1 5 10 15 <210> 52 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide . <400> 52 -13- 159403 - Sequence Listing.doc 201219053

Gly ser lie Glu Pro Glu Glu Phe Leu Thr Pro Asp Gin Met Lys 15 10 15 <210> 53 <211> 13 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 53

Lys Lys Leu Gin cys val Gin Leu His val lie ser Met 15 10 <210> 54 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 54

Gly Pro Leu val cys Asn Gly val Leu Gin Gly lie Thr ser Met 15 10 15 <210> 55 <211> 15 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide. <400> 55

Gly Val Leu Gin Gly lie Thr ser Met Gly ser Glu Pro cys Ala 15 10 15 159403 - Sequence Listing.doc 14-

Claims (1)

201219053 VII. Patent Application Range: 1. A method for producing one or more antigen-specific regulatory T cells (Treg), comprising: isolating one or more human dendritic cells (DC) from an individual; using one or more antigens Binding or fused anti-DC- asialoglycoprotein receptor (DC-ASGPR)-specific antibody or binding fragment thereof, the one or more antigens are loaded into the one or more DCs to form an antigen-loaded DC; The antigen-loaded DC is contacted with one or more original T cells, wherein the antigen-loaded DC stimulates proliferation of antigen-specific Treg. 2. The method of claim 1, wherein the one or more antigens comprise a peptide or a protein. 3. The method of claim 2 wherein the peptide is a foreign antigen or a self antigen. 4. The method of claim 2 wherein the peptide triggers an allergic reaction or an asthma response. 5. The method of claim 2, wherein the one or more antigens comprise a bacterial protein glycoprotein, a fungal protein, a protozoal protein or an oncoprotein. Method of Moon Length Item 1 wherein the antigens comprise H A-1, PSA or a combination and modification thereof. 7. The method of claim 1, wherein the antigen-specific Tregs are IL-10-secreting Treg 〇8. The method of claim 1 &gt; + a, wherein the dendritic cell lines are used in one or more Autoantigen-mediated autoimmune disease, multiple sclerosis, influenza or cancer; π prevent, treat, and ameliorate symptoms. 159403.doc 201219053 9. The method of claim 8, wherein the autoimmune diseases are selected from the group consisting of: allergy, asthma, celiac disease, type III diabetes (IDDM), systemic lupus erythematosus (SLE) ), Sj0grenis syndrome, Churg Strauss Syndr〇me, Hashimoto's thyroiditis, Graves, disease, idiopathic Thrombocytopenic purpura, transplant rejection, multiple sclerosis, psoriasis, and rheumatoid arthritis (RA). 10. The method of claim 1, wherein the anti-system is produced by the fusion tumor cells contained in ATCC Accession No. 10248. 11. A vaccine composition against one or more autoantigen-mediated autoimmune diseases comprising an anti-DC- asialoglycoprotein receptor (ASGpR) specificity for binding or fusion to one or more autoantigens An antibody or a binding fragment thereof, and one or more optionally pharmaceutically acceptable adjuvants, wherein the vaccine composition produces one or more autoantigen-specific IL i〇 secreting regulatory T cells (Treg) production, enhanced production , or generate and enhance production. 12. The composition of claim ' wherein the autoimmune diseases are selected from the group consisting of: allergy, asthma, celiac disease, diabetes, 13th 4 diabetes (IDDM), systemic lupus erythematosus (SLE) , Hugh's syndrome, Cherokee's Syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, transplant rejection, multiple sclerosis, psoriasis and rheumatoid arthritis RA). 13. The composition of π 求 丨 , , , , , 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 〇 Intestinal or intranasal administration. 15. A composition as claimed, wherein the one or more autoantigens comprise peptides, proteins, lipids, carbohydrates, nucleic acids, and combinations thereof. 16. The composition of claim </RTI> wherein the composition binds to and activates dendritic cells which activate IL-10 secreting Treg. 17. The composition of claim 11 wherein the anti-system is made from a fusion tumor cell contained in ATcc Accession No. PTA-10248. 18. A pharmaceutical composition for use as a medicament for preventing cancer, improving cancer symptoms, or preventing cancer and improving cancer symptoms, comprising: an anti-DC- asialoglycoprotein receptor (ASGPR)-specific antibody or a combination thereof A recombinant fusion protein that binds or fused to one or more cancer-specific antigens; and one or more pharmaceutically acceptable excipients or adjuvants as appropriate. 19. The composition of claim 18, wherein the composition is administered orally, parenterally or intranasally. 20. The composition of claim 18, wherein the composition produces, increases, or produces and increases levels of one or more regulatory tau cells (Tregs) specific for prostatic serum antigen (PSA). 21. The composition of claim 20 wherein the antigen is a prostate cancer antigen and the Treg is prostate cancer specific il-10 Treg. 22. The composition of claim 20, wherein the cancer-specific antigens are peptides selected from the group consisting of tumor-associated antigens selected from the group consisting of CEA, prostate, prostate specific antigen (PSA), HER-2/ Neu, BAGE, 159403.doc 201219053 GAGE, MAGE 1 to 4, 6 and 12, MUC (mucin) (eg MUC-1, MUC-2, etc.), GM2 and GD2 gangliosides, ras, myc, tyramine Acidase, MART (melanoma antigen), MARCO-MART, cyclin B1, cyclin D, Pmel 17 (gpl00), GnT_V intron V sequence (N-ethyl glucosyltransferase V intron) V sequence), prostate cancer psm, prostate serum antigen (PSA), PRAME (melanoma antigen), β-sodium catenin, MUM-1-B (melanoma mutated gene product), GAGE (melanoma) Antigen) 1, BAGE (melanoma antigen) 2 to 10, c-ERB2 (Her2/neu), EBNA (EB (Epstein-Barr) virus nuclear antigen) 1 to 6, gp75, human papillomavirus (HPV) E6 And E7, p53, lung resistance protein (LRP), Bcl-2 and Ki-67. A composition according to claim 18, wherein the anti-system is prepared from a fusion tumor cell contained in ATcc Accession No. ΡΤΑ-1〇248. 24. A pharmaceutical composition for use as a medicament for preventing pathogens, improving symptoms of pathogens, or preventing pathogens and improving symptoms of pathogens, comprising: an anti-DC- asialoglycoprotein receptor (ASGPR)-specific antibody or binding thereof A recombinant fusion protein that binds or fused to one or more pathogen antigens; and one or more pharmaceutically acceptable excipients or adjuvants as appropriate. 25. The composition of claim 24, wherein the composition is administered orally, parenterally or intranasally. 26. The composition of claim 24, wherein the composition produces, increases, or produces and increases levels of - or a plurality of pathogen-specific regulatory T cells (Tregs). 159403.doc -4· S 201219053 .24; Composition of 24 wherein the anti-system is prepared by the fusion tumor cells contained in ATCC Accession No. ΡΤΑ-10248. 28. A pharmaceutical composition, which is used as a drug for the prevention of autoimmune diseases mediated by one or more autoantigens, enemies, and sputum L. ^ 药物 drugs for improving symptoms, or preventing and improving symptoms , comprising: a recombinant fusion protein that binds or fuses with an anti-DC- asialoglycoprotein receptor (ASGpR)-specific antibody or binding fragment thereof and/or a plurality of autoantigens; and one or more pharmaceutically Accepted adjuvants wherein the vaccine composition produces, enhances, produces, and enhances production of one or more autoantigen-specific regulatory tau cells (Tregs). 29. The composition of claim 28, wherein the autoimmune disease is selected from the group consisting of: allergy, asthma, celiac disease, diabetes, type 1 diabetes (IDDM), systemic lupus erythematosus (SLE) , Hugh's syndrome, Cherokee's syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, transplant rejection, multiple sclerosis, psoriasis and rheumatoid arthritis (RA). The composition of claim 28, wherein the autoimmune disease is type 1 diabetes (IDDM). The composition of claim 28, wherein the Treg is an autoantigen-specific Treg that secretes IL_1〇. 32. The composition of claim 28, wherein the composition is administered orally, parenterally or intranasally. 33. The composition of claim 28, wherein the one or more autoantigens comprise 159403.doc 201219053 Peptides, proteins, lipids, carbohydrated human anti-aqueous compounds, nucleic acids, and combinations thereof. 34. The composition of claim 28, which is prepared from a fusion cell containing AT&apos;s accession number PTA-10248. 35. A pharmaceutical composition for producing a self-antigen-specific regulation of fine mail (IV), comprising: an anti-DC-sialic acid glycoprotein into &amp; white body (ASGPR)-specific antibody or a binding fragment thereof One or more self-humans, body antigens, ., sigma or fused recombinant fusion proteins; and one or more opt-outs, such as the composition of claim 35, 36. pharmaceutically acceptable excipients or An adjuvant, wherein the composition is for use in the prevention, treatment, and amelioration of one or more autoantigen-mediated autoimmune diseases, multiple sclerosis, or cancer. 37. The composition of claim 35, wherein the self The antigen is selected from the following antigens: allergies, asthma, celiac disease, type I diabetes (IDDM), systemic lupus erythematosus (SLE), Hugh's syndrome, Qiqi-si syndrome, Hashimoto's thyroiditis , Graves' disease, idiopathic thrombocytopenic purpura, transplant rejection, multiple sclerosis, psoriasis and rheumatoid arthritis (RA) 38. The composition of the remedy 35, the anti-system By atc pta-10248 The hybridoma cell containing system. Sub. No. flat I59403.doc