Anti-ASGR1 polypeptides for immune tolerance and methods of use

Abstract

Disclosed herein are binding polypeptides that bind to asialoglycoprotein receptor 1 (ASGR1) and methods of use thereof for inducing immune tolerance against an antigen of interest, e.g., for treating, ameliorating, inhibiting, or preventing a disease or disorder associated with an undesired immune response against an antigen of interest, such as an autoimmune disease.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 268,183 filed on February 17, 2022, U.S. Provisional Patent Application No. 63 / 268,190 filed on February 17, 2022, U.S. Provisional Patent Application No. 63 / 483,456 filed on February 6, 2023, and U.S. Provisional Patent Application No. 63 / 483,466 filed on February 6, 2023.

[0003] References to sequence listings

[0004] This application is submitted with a sequence listing in electronic format. The sequence listing is provided in the form of an XML file named ANOK.048WO_ST26, which was created and last modified on February 16, 2023, and is 649,053 bytes in size. The information in the electronic sequence listing is hereby incorporated by reference in its entirety. Technical Field

[0005] Aspects of the present disclosure relate to binding polypeptides that bind to ASGR1 and methods of use thereof for inducing immune tolerance to a desired antigen, such as for the prevention or treatment of a disease. Background Art

[0006] The liver is involved in a variety of tolerogenic processes. For example, it plays a role in the development of immune tolerance to non-self antigens absorbed into the blood discharged from the intestine or to newly formed antigens generated by liver metabolic activities. Targeting the liver with therapeutic compositions may be beneficial for the induction of antigen-specific immune tolerance. Summary of the invention

[0007] Antigens such as non-self antigens absorbed into the blood discharged from the intestine or newly formed antigens generated by metabolic activities of the liver are unable to induce immune responses in healthy individuals. Antigen-specific immune tolerance and cross-tolerance induction for CD4+ and CD8+ T cells, respectively, have been attributed to various cell types in the liver, including hepatocytes and liver sinusoidal endothelial cells (LSECs). Hepatocytes are the main cell type constituting the liver parenchyma, and they can process and present antigens on MHC-I and MHC-II to signal to CD8+ and CD4+ T cells, respectively. LSECs efficiently clear, process and present soluble antigens on MHC-I and MHC-II in the bloodstream to circulating lymphocytes, usually resulting in the induction of CD4+ regulatory T cells or anergic CD8+ T cells.

[0008] Disclosed herein are asialoglycoprotein receptor 1 (ASGR1) binding polypeptides. In some embodiments, the ASGR1 binding polypeptide comprises a heavy chain variable region. In some embodiments, the heavy chain variable region comprises one or more of HCDR1, HCDR2, and HCDR3. In some embodiments, the ASGR1 binding polypeptide comprises or further comprises a light chain variable region. In some embodiments, the light chain variable region comprises one or more of LCDR1, LCDR2, and LCDR3.

[0009] Also disclosed herein are tolerogenic compounds comprising an ASGR1 binding polypeptide conjugated or fused to an antigen to which tolerance is desired. The ASGR1 binding polypeptide and the antigen can be conjugated or fused using a linker. The ASGR1 binding polypeptide and the antigen can be chemically conjugated with a chemical conjugation linker. The ASGR1 binding polypeptide and the antigen can be recombinantly fused.

[0010] Also disclosed herein are compositions comprising any one of the tolerogenic compounds disclosed herein and a pharmaceutically acceptable excipient.

[0011] Also disclosed herein are methods of inducing tolerance to an antigen to which a subject is capable of producing an undesirable immune response. In some embodiments, the method comprises administering to the subject any one of the tolerogenic compounds or compositions disclosed herein.

[0012] Also disclosed herein are the tolerogenic compounds or compositions disclosed herein for use in inducing tolerance to an antigen in a subject to which the subject is capable of producing an undesirable immune response, or for use in the preparation of a medicament.

[0013] In some embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, comprising a heavy chain variable region (VH), the heavy chain variable region comprising a first heavy chain complementary determining region (HCDR1), a second heavy chain complementary determining region (HCDR2), and a third heavy chain complementary determining region (HCDR3). In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:9. In some embodiments, HCDR2 comprises the sequence of SEQ ID NO:10. In some embodiments, HCDR3 comprises the sequence of SEQ ID NO:11. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:9, HCDR2 comprises the sequence of SEQ ID NO:10, and HCDR3 comprises the sequence of SEQ ID NO:11. In several embodiments, the VH comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:15.

[0014] In several embodiments, the ASGR1 binding polypeptide further comprises a light chain variable region (VL) comprising a first light chain complementarity determining region (LCDR1), a second light chain complementarity determining region (LCDR2), and a third light chain complementarity determining region (LCDR3). In several embodiments, LCDR1 comprises the sequence of SEQ ID NO: 25. In several embodiments, LCDR2 comprises the sequence of SEQ ID NO: 26. In several embodiments, LCDR3 comprises the sequence of SEQ ID NO: 27. In several embodiments, LCDR1 comprises the sequence of SEQ ID NO: 25, LCDR2 comprises the sequence of SEQ ID NO: 26, and LCDR3 comprises the sequence of SEQ ID NO: 27. In several embodiments, the VL comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:31.

[0015] In several embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO:15, and the light chain variable region comprises the sequence of SEQ ID NO:31.

[0016] In several embodiments, the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb), or a scFv. In several embodiments, the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

[0017] In several embodiments, a polynucleotide encoding an ASGR1 binding polypeptide is provided, the polynucleotide comprising one or more of the sequences of SEQ ID NOs: 12-14 or sequences having at least 95% identity thereto. In several embodiments, the polynucleotide comprises the sequence of SEQ ID NO: 16. In several embodiments, the polynucleotide comprises one or more of the sequences of SEQ ID NOs: 25-27. In several embodiments, the polynucleotide comprises the sequence of SEQ ID NO: 32. In several embodiments, the polynucleotide further comprises the sequence of SEQ ID NO: 16.

[0018] In several embodiments, a tolerogenic compound is provided, comprising an ASGR1 binding polypeptide as disclosed herein, wherein the ASGR1 binding polypeptide is conjugated or fused to an antigen to which tolerance is desired. In several embodiments, the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker. In several embodiments, the linker is a cleavable linker. In several embodiments, the linker comprises glycine and / or serine, optionally wherein the linker comprises a sequence of SEQ ID NO: 37 or a sequence having at least about 80%, 85%, 90% or 95% identity thereto. In several embodiments, the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide.

[0019] In several embodiments, the antigen comprises a food antigen. In several embodiments, the food antigen is associated with diarrheal celiac disease. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to SEQ ID NO: 41. In several embodiments, the antigen comprises SEQ ID NO: 41. In some embodiments, the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), phytocystatin, thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin, and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitor protein (Sin a 4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenain, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens, and a mimetic of any one of the antigens. In several embodiments, the food antigen is selected from the group consisting of high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenain, a portion of any one of the antigens, and a mimetic of any one of the antigens. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

[0020] In some embodiments, the antigen comprises an autoantigen. In some embodiments, the autoantigen is selected from thyroglobulin, thyroid peroxidase, thyroid stimulating hormone receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

[0021] In several embodiments, the antigen comprises an antigen associated with an autoimmune disease. In several embodiments, the autoimmune disease is selected from the group consisting of multiple sclerosis, type 1 diabetes, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, diarrheal celiac disease, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and anti-neutrophil cytoplasmic antibody-associated vasculitis. In several embodiments, the autoimmune disease is multiple sclerosis. In several embodiments, the antigen comprises a sequence or fragment thereof having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 100, 71-99, 101-106, or 157-159. In several embodiments, the autoimmune disease is type 1 diabetes. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0022] In several embodiments, tolerogenic compounds are provided, comprising an ASGR1 binding polypeptide conjugated or fused to an antigen to which tolerization is desired, wherein the ASGR1 binding polypeptide comprises a heavy chain variable region comprising one, two, or all three of HCDR1, HCDR2, and HCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO: 9; wherein HCDR2 comprises the sequence of SEQ ID NO: 10; and / or wherein HCDR3 comprises the sequence of SEQ ID NO: 11. In several embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 15. In several embodiments, the ASGR1 binding polypeptide further comprises a light chain variable region comprising one, two or all three of LCDR1, LCDR2 and LCDR3; wherein LCDR1 comprises the sequence of SEQ ID NO:25; wherein LCDR2 comprises the sequence of SEQ ID NO:26; and / or wherein LCDR3 comprises the sequence of SEQ ID NO:27. In several embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:31. In several embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO:15, and the light chain variable region comprises the sequence of SEQ ID NO:31. In several embodiments, the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb), or a scFv. In several embodiments, the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced. In several embodiments, the antigen is a polypeptide.

[0023] In several embodiments, the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker. In several embodiments, the linker is a cleavable linker. In several embodiments, the linker comprises glycine and / or serine, optionally wherein the linker comprises the sequence of SEQ ID NO: 37. In several embodiments, the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide. In several embodiments, the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced, and the antigen is conjugated or fused to the Fc domain, optionally wherein the antigen is conjugated or fused to the C-terminus of the Fc domain.

[0024] In several embodiments, the antigen comprises a food antigen. In some embodiments, the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), plant cysteine ​​proteinase inhibitor (phytocystatin), thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin, and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitor protein (Sin a 3), 1S globulin (Sin a 4), 1S globulin (Sin a 5), ​​1S globulin (Sin a 6), 1S globulin (Sin a 7), 1S globulin (Sin a 8), 1S globulin (Sin a 9), 1S globulin (Sin a 11), 1S globulin (Sin a 2), 1S globulin (Sin a 3), 1S globulin (Sin a 12), 1S globulin (Sin a 13), 1S globulin (Sin a 14), 1S globulin (Sin a 15), 1S globulin (Sin a 16), 1S globulin (Sin a 17), 1S globulin (Sin a 18), a4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens, and a mimetic of any one of the antigens. In several embodiments, the food antigen is selected from the group consisting of high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, a portion of any one of the antigens, and a mimetic of any one of the antigens. In several embodiments, the food antigen is associated with diarrheal celiac disease. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

[0025] In some embodiments, the antigen comprises an autoantigen. In some embodiments, the autoantigen is selected from thyroglobulin, thyroid peroxidase, thyroid stimulating hormone receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

[0026] In several embodiments, the antigen comprises an antigen associated with an autoimmune disease. In several embodiments, the autoimmune disease is selected from the group consisting of type 1 diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, diarrheal celiac disease, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and anti-neutrophil cytoplasmic antibody-associated vasculitis. In several embodiments, the autoimmune disease is type 1 diabetes. In several embodiments, the antigen comprises a sequence or a fragment thereof having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156. In several embodiments, the autoimmune disease is multiple sclerosis. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 71-106 or 157-159, or a fragment thereof.In several embodiments, the autoimmune disease is rheumatoid arthritis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 107-116, 160-239 or 272-288, or a fragment thereof; Sjogren's syndrome, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 117-118 or 240-245, or a fragment thereof; rheumatic heart disease, autoimmune myocarditis, viral myocarditis or inflammatory cardiomyopathy, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 117-118 or 240-245, or a fragment thereof; NO: 119 or any one of 251-270 having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity or a fragment thereof; Parkinson's disease, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity or a fragment thereof to any one of SEQ ID NO: 120 or 247-250; anti-neutrophil cytoplasmic antibody-associated vasculitis (ANCA vasculitis), wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity or a fragment thereof to any one of SEQ ID NO: 121-129; primary biliary cholangitis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity or a fragment thereof to any one of SEQ ID NO: 121-129; NO:130-132 or 271 having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity, or a fragment thereof; or autoimmune hepatitis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NO:133-140, or a fragment thereof.

[0027] In several embodiments, the antigen comprises an alloantigen. In several embodiments, the alloantigen is selected from the group consisting of subunits of MHC class I and MHC class II haplotype proteins and their complexes with the antigens they present, and minor blood group antigens RhCE, Kell, Kidd, Duffy, Diego, and MNSs.

[0028] Also provided are compositions comprising a tolerogenic compound according to the present disclosure and a pharmaceutically acceptable excipient.

[0029] Provided herein are methods of inducing tolerance to an antigen to which a subject is capable of producing an undesirable immune response, the method comprising administering to the subject a tolerogenic compound or composition provided herein. In several embodiments, the compound or composition is administered before the subject is exposed to the antigen, after the subject is exposed to the antigen, or in both cases. In several embodiments, the undesirable immune response is associated with an allergic reaction to a food, animal, plant, or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

[0030] In several embodiments, a tolerogenic compound or composition as disclosed herein is provided for inducing tolerance to an antigen in a subject to which the subject is capable of producing an undesirable immune response. In several embodiments, the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or a graft-versus-host disease. In several embodiments, the tolerogenic compound or composition is used to prepare a medicament. In several embodiments, the composition is used to induce immune tolerance in a subject in need thereof.

[0031] Also provided herein is a method of inducing tolerance to an antigen to which tolerance is desired, the method comprising administering to a subject a compound comprising: (i) an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region comprising HCDR1, HCDR2, and HCDR3; wherein HCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO:9; wherein HCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO:10; and wherein HCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO:11; and (ii) an antigen to which tolerance is desired, wherein the ASGR1 binding polypeptide is conjugated or fused to the antigen to which tolerance is desired.

[0032] In several embodiments, the ASGR1 binding polypeptide further comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises a sequence at least 90% identical to SEQ ID NO:25, wherein LCDR2 comprises a sequence at least 90% identical to SEQ ID NO:26, and wherein LCDR3 comprises a sequence at least 90% identical to SEQ ID NO:27.

[0033] In several embodiments, the antigen to which tolerance is desired is associated with diarrheal celiac disease. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 41, 40, 43-54, or a fragment thereof. In several embodiments, the antigen to which tolerance is desired is associated with MS. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 100, 71-99, 101-106 or 157-159, or a fragment thereof. In several embodiments, the antigen to which tolerance is desired is associated with type 1 diabetes. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0034] In several embodiments, administration is via an intravenous route.

[0035] In several embodiments, a method for delivering an antigen to liver tissue of a subject is provided, the method comprising: conjugating or fusing the antigen to an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide, thereby generating an ASGR1 binding polypeptide-antigen complex; and contacting the ASGR1 binding polypeptide-antigen complex with the subject's liver tissue in situ, wherein the contact allows the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, thereby delivering the antigen to the liver tissue.

[0036] In an additional embodiment, a method of delivering an antigen to liver tissue of a subject is provided, the method comprising: contacting an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide conjugated or fused to an antigen with the liver tissue of the subject in situ, thereby allowing the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, wherein the ASGR1 binding polypeptide comprises a heavy chain variable region comprising HCDR1, HCDR2, and HCDR3, wherein HCDR1 comprises a sequence at least 90% identical to SEQ ID NO:9, wherein HCDR2 comprises a sequence at least 90% identical to SEQ ID NO:10; and wherein HCDR3 comprises a sequence at least 90% identical to SEQ ID NO:11; and wherein the ASGR1 binding polypeptide comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises a sequence at least 90% identical to SEQ ID NO:25, wherein LCDR2 comprises a sequence at least 90% identical to SEQ ID NO:26; and wherein LCDR3 comprises a sequence at least 90% identical to SEQ ID NO: NO: 27 has a sequence with at least 90% identity. In several embodiments, delivering the antigen in situ to the liver tissue of the subject induces tolerance to the antigen in the subject. In several embodiments, the method is used to treat an autoimmune disease.

[0037] In several embodiments, the method is used to treat allergic reactions.

[0038] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:297; wherein HCDR2 comprises the sequence of SEQ ID NO:298; wherein HCDR3 comprises the sequence of SEQ ID NO:299; wherein LCDR1 comprises the sequence of SEQ ID NO:313; wherein LCDR2 comprises the sequence of SEQ ID NO:314; and wherein LCDR3 comprises the sequence of SEQ ID NO:315. In several embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 303. In several embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 319. In several embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO:303, and the light chain variable region comprises the sequence of SEQ ID NO:319.

[0039] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:329; wherein HCDR2 comprises the sequence of SEQ ID NO:330; wherein HCDR3 comprises the sequence of SEQ ID NO:331; wherein LCDR1 comprises the sequence of SEQ ID NO:345; wherein LCDR2 comprises the sequence of SEQ ID NO:346; and wherein LCDR3 comprises the sequence of SEQ ID NO:347.

[0040] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:361; wherein HCDR2 comprises the sequence of SEQ ID NO:362; wherein HCDR3 comprises the sequence of SEQ ID NO:363; wherein LCDR1 comprises the sequence of SEQ ID NO:377; wherein LCDR2 comprises the sequence of SEQ ID NO:378; and wherein LCDR3 comprises the sequence of SEQ ID NO:379.

[0041] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:393; wherein HCDR2 comprises the sequence of SEQ ID NO:394; wherein HCDR3 comprises the sequence of SEQ ID NO:395; wherein LCDR1 comprises the sequence of SEQ ID NO:409; wherein LCDR2 comprises the sequence of SEQ ID NO:410; and wherein LCDR3 comprises the sequence of SEQ ID NO:411.

[0042] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:425; wherein HCDR2 comprises the sequence of SEQ ID NO:426; wherein HCDR3 comprises the sequence of SEQ ID NO:427; wherein LCDR1 comprises the sequence of SEQ ID NO:441; wherein LCDR2 comprises the sequence of SEQ ID NO:442; and wherein LCDR3 comprises the sequence of SEQ ID NO:443.

[0043] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:457; wherein HCDR2 comprises the sequence of SEQ ID NO:458; wherein HCDR3 comprises the sequence of SEQ ID NO:459; wherein LCDR1 comprises the sequence of SEQ ID NO:473; wherein LCDR2 comprises the sequence of SEQ ID NO:474; and wherein LCDR3 comprises the sequence of SEQ ID NO:475.

[0044] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:489; wherein HCDR2 comprises the sequence of SEQ ID NO:490; wherein HCDR3 comprises the sequence of SEQ ID NO:491; wherein LCDR1 comprises the sequence of SEQ ID NO:505; wherein LCDR2 comprises the sequence of SEQ ID NO:506; and wherein LCDR3 comprises the sequence of SEQ ID NO:507.

[0045] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:521; wherein HCDR2 comprises the sequence of SEQ ID NO:522; wherein HCDR3 comprises the sequence of SEQ ID NO:523; wherein LCDR1 comprises the sequence of SEQ ID NO:537; wherein LCDR2 comprises the sequence of SEQ ID NO:538; and wherein LCDR3 comprises the sequence of SEQ ID NO:539.

[0046] In several embodiments, an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide is provided, which comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3; wherein HCDR1 comprises the sequence of SEQ ID NO:553; wherein HCDR2 comprises the sequence of SEQ ID NO:554; wherein HCDR3 comprises the sequence of SEQ ID NO:555; wherein LCDR1 comprises the sequence of SEQ ID NO:569; wherein LCDR2 comprises the sequence of SEQ ID NO:570; and wherein LCDR3 comprises the sequence of SEQ ID NO:571.

[0047] In several embodiments, provided herein are asialoglycoprotein receptor 1 (ASGR1) binding polypeptides comprising a heavy chain variable region comprising HCDR1, HCDR2, and HCDR3, wherein HCDR1 comprises the sequence of SEQ ID NO: 9; wherein HCDR2 comprises the sequence of SEQ ID NO: 10; and wherein HCDR3 comprises the sequence of SEQ ID NO: 11. In several embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 15.

[0048] In several embodiments, the ASGR1 binding polypeptide further comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises the sequence of SEQ ID NO:25, wherein LCDR2 comprises the sequence of SEQ ID NO:26; and wherein LCDR3 comprises the sequence of SEQ ID NO:27. In several embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:31. In several embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO:15, and the light chain variable region comprises the sequence of SEQ ID NO:31.

[0049] Depending on the embodiment, the ASGR1 binding polypeptide is optionally an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb), or a scFv. In several embodiments, the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

[0050] Also provided herein is a polynucleotide encoding an ASGR1 binding polypeptide comprising one or more of the sequences of SEQ ID NOs: 12-14 or 25-27. In several embodiments, the polynucleotide comprises the sequence of SEQ ID NOs: 16 or 32 or both. In several embodiments, the polynucleotide encodes an ASGR1 binding polypeptide according to the disclosure herein.

[0051] In several embodiments, tolerogenic compounds are also provided, comprising an ASGR1 binding polypeptide conjugated or fused to an antigen to which tolerance is desired. In several embodiments, the ASGR1 binding polypeptide comprises a heavy chain variable region comprising HCDR1, HCDR2, and HCDR3, wherein HCDR1 comprises the sequence of SEQ ID NO:9, wherein HCDR2 comprises the sequence of SEQ ID NO:10, and wherein HCDR3 comprises the sequence of SEQ ID NO:11. In several embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:15.

[0052] In several embodiments, the tolerogenic compounds provided herein further comprise a light chain variable region comprising LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises the sequence of SEQ ID NO: 25, wherein LCDR2 comprises the sequence of SEQ ID NO: 26, and wherein LCDR3 comprises the sequence of SEQ ID NO: 27. In several embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 31. In several embodiments, wherein the heavy chain variable region comprises the sequence of SEQ ID NO: 15, and the light chain variable region comprises the sequence of SEQ ID NO: 31.

[0053] In several embodiments of the tolerogenic compounds provided herein, the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb), or a scFv. In several embodiments of the tolerogenic compounds provided herein, the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

[0054] In several embodiments of the tolerogenic compounds provided herein, the antigen is a polypeptide. In several embodiments, the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker. In some embodiments, the linker is a cleavable linker. In several embodiments, the linker comprises glycine and / or serine, optionally wherein the linker comprises the sequence of SEQ ID NO: 37. In several embodiments, the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide. In some embodiments of the tolerogenic compounds provided herein, the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced, and the antigen is conjugated or fused to the Fc domain, optionally wherein the antigen is conjugated or fused to the C-terminus of the Fc domain.

[0055] In several embodiments of the tolerogenic compounds provided herein, the antigen comprises a food antigen. In some embodiments, the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), phytocystatin, thaumatin-like protein (Act d2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin, and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitor protein (Sin a 4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pena 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens and a mimetic of any one of the antigens.

[0056] In several embodiments, the food antigen is selected from the group consisting of high molecular weight glutenins, low molecular weight glutenins, α-, γ- and Ω-gliadins, hordeins, secalins, avenins, a portion of any of said antigens, and a mimetic of any of said antigens.

[0057] In several embodiments, the food antigen is associated with diarrheal celiac disease.In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

[0058] In several embodiments of the tolerogenic compounds provided herein, the antigen comprises an autoantigen. In several embodiments, the autoantigen is selected from thyroglobulin, thyroid peroxidase, thyroid stimulating hormone receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

[0059] In several embodiments of the tolerogenic compounds provided herein, the antigen comprises an antigen associated with an autoimmune disease. In several embodiments, the autoimmune disease is selected from the group consisting of type 1 diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, celiac disease with diarrhea, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and anti-neutrophil cytoplasmic antibody-associated vasculitis.

[0060] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is type 1 diabetes. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0061] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is multiple sclerosis. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 71-106 or 157-159, or a fragment thereof.

[0062] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is rheumatoid arthritis. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 107-116, 160-239 or 272-288, or a fragment thereof.

[0063] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is Sjogren's syndrome. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 117-118 or 240-245, or a fragment thereof.

[0064] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is rheumatic heart disease, autoimmune myocarditis, viral myocarditis, or inflammatory cardiomyopathy. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 119 or 251-270, or a fragment thereof.

[0065] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is Parkinson's disease. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 120 or 247-250, or a fragment thereof.

[0066] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is anti-neutrophil cytoplasmic antibody-associated vasculitis (ANCA vasculitis). In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 121-129, or a fragment thereof.

[0067] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is primary biliary cholangitis. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 130-132 or 271, or a fragment thereof.

[0068] In several embodiments of the tolerogenic compounds provided herein, the autoimmune disease is autoimmune hepatitis. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 133-140, or a fragment thereof.

[0069] In several embodiments of the tolerogenic compounds provided herein, the antigen comprises an alloantigen. In several embodiments, the alloantigen is selected from the group consisting of subunits of MHC class I and MHC class II haplotype proteins and their complexes with the antigens they present, and minor blood group antigens RhCE, Kell, Kidd, Duffy, Diego, and MNSs.

[0070] In several embodiments, provided herein are compositions comprising a tolerogenic compound according to the present disclosure and a pharmaceutically acceptable excipient.

[0071] Also provided herein is a method of inducing tolerance to an antigen to which a subject is capable of producing an undesirable immune response, the method comprising administering to the subject a tolerogenic compound according to the disclosure or a composition according to the disclosure.

[0072] In several embodiments, the tolerogenic compound or composition is administered prior to exposure of the subject to the antigen, after exposure of the subject to the antigen, or both.

[0073] In several embodiments, the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

[0074] Also provided herein are tolerogenic compounds of the present disclosure and / or compositions of the present disclosure for inducing tolerance to an antigen in a subject to which the subject is capable of producing an undesirable immune response. In several embodiments, the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

[0075] Also provided herein are tolerogenic compounds of the present disclosure and / or compositions of the present disclosure for use in the preparation of a medicament.Additionally, provided herein are compounds of the present disclosure and / or compositions of the present disclosure for use in inducing immune tolerance in a subject in need thereof.

[0076] In several embodiments, a method of inducing tolerance to an antigen to which tolerance is desired is provided, the method comprising administering to a subject a compound comprising (i) an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region comprising HCDR1, HCDR2, and HCDR3, wherein HCDR1 comprises a sequence at least 90% identical to SEQ ID NO:9, wherein HCDR2 comprises a sequence at least 90% identical to SEQ ID NO:10, and wherein HCDR3 comprises a sequence at least 90% identical to SEQ ID NO:11; and (ii) an antigen to which tolerance is desired, wherein the ASGR1 binding polypeptide is conjugated or fused to the antigen to which tolerance is desired.

[0077] In several embodiments, the ASGR1 binding polypeptide further comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises a sequence at least 90% identical to SEQ ID NO:25, wherein LCDR2 comprises a sequence at least 90% identical to SEQ ID NO:26, and wherein LCDR3 comprises a sequence at least 90% identical to SEQ ID NO:27.

[0078] In several embodiments of the provided methods, the antigen to which tolerance is desired is associated with diarrheal celiac disease. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

[0079] In several embodiments of the provided methods, the antigen to which tolerance is desired is associated with multiple sclerosis (MS). In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 71-106 or 157-159, or a fragment thereof.

[0080] In several embodiments of the provided methods, wherein the antigen to which tolerance is desired is associated with type 1 diabetes. In several embodiments, the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0081] In several embodiments of the provided methods, administration is via an intravenous route.

[0082] Also provided herein is a method for delivering an antigen to liver tissue of a subject, the method comprising: conjugating or fusing the antigen to an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide, thereby generating an ASGR1 binding polypeptide-antigen complex; and contacting the ASGR1 binding polypeptide-antigen complex with the subject's liver tissue in situ, wherein the contact allows the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, thereby delivering the antigen to the liver tissue.

[0083] Additionally provided is a method of delivering an antigen to liver tissue of a subject, the method comprising: contacting an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide conjugated or fused to an antigen with the liver tissue of the subject in situ, thereby allowing the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, wherein the ASGR1 binding polypeptide comprises a heavy chain variable region comprising HCDR1, HCDR2, and HCDR3, wherein HCDR1 comprises a sequence at least 90% identical to SEQ ID NO:9, wherein HCDR2 comprises a sequence at least 90% identical to SEQ ID NO:10, and wherein HCDR3 comprises a sequence at least 90% identical to SEQ ID NO:11; and wherein the ASGR1 binding polypeptide comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises a sequence at least 90% identical to SEQ ID NO:25, wherein LCDR2 comprises a sequence at least 90% identical to SEQ ID NO:26, and wherein LCDR3 comprises a sequence at least 90% identical to SEQ ID NO:27. NO:27 has a sequence with at least 90% identity. In several embodiments, the antigen is delivered in situ to the liver tissue of the subject to induce tolerance to the antigen in the subject. In several embodiments, such methods are used to treat autoimmune diseases. In several embodiments, such methods are used to treat allergic reactions. BRIEF DESCRIPTION OF THE DRAWINGS

[0084] In addition to the features described herein, additional features and variations will become apparent from the drawings and the following description of the embodiments.It should be understood that these drawings depict various embodiments and are not intended to limit the scope.

[0085] Figure 1A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0086] Figure 1B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0087] Figure 1C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0088] Figure 1D Non-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0089] Figure 1ENon-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0090] Figure 1F Non-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0091] Figure 2A Depicted are the sequences of human asialoglycoprotein receptor 1 (ASGR1) (SEQ ID NO:33), its extracellular domain (ECD) spanning amino acids Q62-L291 (SEQ ID NO:34), and a His-Avi tagged variant (SEQ ID NO:35) that can be used for antigen production and purification.

[0092] Figure 2B Depicted are sequences of p31 mimotope peptides that induce T1D pathology in the BDC2.5 model, conjugatable p31 variants with a cysteine ​​linker for attachment of an ASGR1 binding polypeptide, non-limiting examples of glycine-serine linkers, and glycine-serine linker p31 variants that can be used for antibody fusions.

[0093] Figure 3 Survival curves are depicted, showing that liver-targeted antigens protect mice from T1D in the pre-activated BDC2.5 model. NOD.SCID mice received 3 x 10 5 Adoptive transfer of activated BDC2.5 T cells and treatment with intravenous injection of 50 pmol / g αASGR1-p31, 50 pmol / g p31 or saline on days 0 and 4. Blood glucose was measured 2-3 times per week for 105 days. Mice were considered diabetic after two consecutive blood glucose measurements ≥ 250 mg / dL. 7-8 animals were used in each group.

[0094] Figure 4 Depicted are diabetes onset survival curves for NOD.SCID mice with adoptively transferred BCD2.5 splenocytes treated with p31 tolerogen (819-p31) fused to anti-ASGR1 antibody mAb-60819, p31 peptide alone, or saline control.

[0095] Figure 5 Depicted are EAE disease severity curves for EAE model mice injected with encephalitogenic T cells and a liver-targeted tolerogen of MOG10 fused to mAb-60819 (819-MOG10). 819-MOG10 was administered at 2 pmol / g ("dose level 1") or 10 pmol / g ("dose level 2").

[0096] Fig. 6A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0097] Figure 6B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0098] Figure 6C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0099] Fig.6DNon-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0100] Fig. 6E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0101] Fig. 6F Non-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0102] Figure 7 Depicted are diabetes onset survival curves for NOD.SCID mice with adoptively transferred BCD2.5 splenocytes treated with p31 tolerogen fused to anti-ASGR1 antibody mAb-60856 (856-p31), p31 peptide alone, or saline control.

[0103] Fig. 8A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0104] Figure 8B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0105] Figure 8C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0106] Fig.8D Non-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0107] Fig. 8E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0108] Figure 8FNon-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0109] Fig.9A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0110] Fig. 9B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0111] Fig. 9C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0112] Fig.9DNon-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0113] Fig.9E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0114] Fig.9F Non-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0115] Fig.10 Depicted are diabetes onset survival curves for NOD.SCID mice with adoptively transferred BCD2.5 splenocytes treated with p31 tolerogen (869-p31) fused to anti-ASGR1 antibody mAb-60869, p31 peptide alone, or saline control.

[0116] Fig.11A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0117] Fig. 11B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0118] Fig. 11C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0119] Fig.11D Non-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0120] Fig.11E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0121] Fig.11FNon-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0122] Fig. 12A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0123] Fig. 12B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0124] Fig. 12C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0125] Fig.12DNon-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0126] Fig.12E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0127] Fig.12F Non-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0128] Fig.13A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0129] Fig. 13BNon-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0130] Fig. 13C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0131] Fig.13D Non-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0132] Fig.13E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0133] Fig.13FNon-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0134] Fig.14A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0135] Fig. 14B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0136] Fig. 14C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0137] Fig.14DNon-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0138] Fig.14E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0139] Fig.14F Non-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0140] Fig.15A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0141] Fig. 15BNon-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0142] Fig. 15C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0143] Fig.15D Non-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0144] Fig.15E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0145] Fig.15FNon-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0146] Fig.16A Non-limiting heavy chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0147] Fig. 16B Non-limiting heavy chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0148] Fig. 16C Non-limiting heavy chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0149] Fig.16DNon-limiting light chain framework sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0150] Fig.16E Non-limiting light chain complementary determining region (CDR) sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated.

[0151] Fig.16F Non-limiting light chain variable region sequences that can be used for the anti-ASGR1 antibodies provided herein are depicted. The provided DNA sequences encode the provided peptide sequences, but other DNA sequences that can encode the same peptide sequences by virtue of codon degeneracy are also contemplated. Peptide sequences and DNA sequences that have at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to the sequences provided therein are also contemplated. DETAILED DESCRIPTION

[0152] Immune responses to various antigens may be an important source of morbidity and mortality. An immune response may occur in an individual, which has an adverse effect on the health and well-being of the individual, reduces the efficacy of the treatment received by the individual, and even reacts to endogenous molecules that occur or exist naturally in the individual. Although in some cases, extensive immunosuppression is used to solve certain types of immune responses, these may lead to general susceptibility to infection and disease. Therefore, more tailored schemes (such as those described herein) are advantageous because antigen-specific immune responses can be targeted. Several embodiments disclosed herein utilize the role of the liver and its various types of cells in the development of immune tolerance to specific antigens. For example, in several embodiments, a specific antigen, its immunogenic fragment and / or its mimics (collectively referred to as antigens, unless otherwise indicated as a specific type, such as a fragment) are connected or coupled to a molecule configured to target the liver (or a specific cell in or associated with the liver), thereby allowing specific antigens, its immunogenic fragments and / or its mimics to be processed, and allowing the immune system to be recalibrated to reduce, improve or otherwise eliminate the immune response to the antigen (or a part of an antigen, or multiple antigens). For example, in several embodiments, the compositions provided herein are targeted for delivery to (and uptake by) the liver, such as hepatocytes or other cells expressing scavenger receptors (eg, asialoglycoprotein receptor (ASGPR), etc.).

[0153] Some embodiments disclosed herein demonstrate that hepatocytes can be manipulated using synthetic constructs (such as those compositions disclosed herein) to actively induce immune tolerance of antigen-specific CD4+ or CD8+ T cells, for example, by presentation or cross-presentation of extracellular antigens. Hepatocytes are considered "non-professional antigen presenting cells" and are promising cell candidates for antigen presentation to induce tolerogenic T cell responses. Hepatocytes account for 80% of the entire liver and are in direct contact with circulating T lymphocytes. Hepatocytes do not express immune co-stimulatory molecules. Therefore, it was tested whether hepatocytes promote peripheral tolerogenic production by presenting and cross-presenting blood-borne antigens. It is demonstrated herein, and according to several embodiments, hepatocytes can be manipulated in situ (e.g., by targeting hepatocytes with constructs according to embodiments disclosed herein) to promote peripheral tolerogenic production by presenting and cross-presenting antigens to T cells.

[0154] Unlike other organs, where circulating lymphocytes only extravasate and enter parenchymal cells under conditions of inflammation, the liver microvasculature has a unique fenestrated endothelium without any basement membrane, allowing direct physical contact between circulating T lymphocytes and liver MHC+ parenchymal cells, including hepatocytes. Hepatocytes have a poor cross-presentation capacity in vitro compared to other liver cells, especially LSECs. Nevertheless, due to the lack of CD28 co-stimulation that leads to T cell clonal exhaustion, direct antigen presentation obtained by transduction of transgenes and / or viral vectors in vitro and in vivo, and subsequent MHC-I-dependent antigen presentation by hepatocytes, can generate immune tolerance mainly through suboptimal activation of antigen-specific CD8+ T lymphocytes. Upon lentiviral-mediated hepatocyte-dependent antigen presentation, induction of CD4+CD25+FoxP3+Treg cells also occurred, indicating that other antigen-presenting cells (APCs) may be involved in hepatocyte-driven tolerogenic mechanisms because hepatocytes express low levels of MHC-II to interact with CD4+ T cells.

[0155] Hepatocytes outnumber other cellular components of the liver and are in close contact with components of the blood. In some embodiments disclosed herein, hepatocytes are used to establish peripheral tolerance of CD4+ and CD8+ T cells by mechanisms of extracellular antigen uptake and presentation or cross-presentation. In other embodiments, alone or in combination with antigen presentation or cross-presentation, the constructs and compositions disclosed herein are used to induce tolerance by other mechanisms. Hepatocytes possess lectin receptors (among other receptors), including asialoglycoprotein receptors (ASGPR). The apoptotic process activates neuraminidase, which desialylates glycoproteins to expose terminal N-acetylgalactosamine residues, which bind to ASGPR. Hepatocyte-dependent antigen presentation or cross-presentation (as well as other mechanisms induced by liver resident immune cells after administration of the constructs disclosed herein [including the induction of regulatory T cells] and related methods) can be more generally used in methods of inducing immune tolerance via T cell exhaustion and / or anergy. In some embodiments, hepatocytes can be used as target cells for tolerogenic prevention or therapeutic intervention.

[0156] In general, the compositions provided herein comprise an antigen of interest (e.g., an antigen to which immune tolerance is desired, including antigenic fragments of larger molecules, or in some embodiments, multiple antigens / fragments thereof), a targeting moiety (e.g., a molecule, particularly a binding polypeptide or antibody, which specifically targets the liver, a cell type within the liver, or another target organ or cell or is recognized by them, such as in lymph nodes and / or spleen), wherein the antigen of interest and the targeting moiety are connected, such as using a linker. In some embodiments, mimetics of these antigens can be used instead of antigens or antigen fragments.

[0157] The scheme for connecting the antigen of interest and the targeting moiety is generally known in the art. For example, when the antigen of interest and the targeting moiety are proteins (for example, when the targeting moiety is an antibody), the antigen of interest and the targeting moiety can be fused together in different configurations, such as the end of any protein. In some embodiments, the antigen of interest and the targeting moiety can be fused together recombinantly in different configurations. In some embodiments, the antigen of interest and the targeting moiety can be fused using traditional recombinant cloning techniques. The antigen of interest and the targeting moiety can be connected using a joint, as generally understood in the art. In several embodiments, the joint is advantageously designed and / or configured to release antigen (or its antigen fragment or its mimetics) in vivo, and the released antigen (or its antigen fragment or its mimetics) is in its natural or substantially natural form (for example, it is conjugated or fused to the form before the joint). In some embodiments, the antigenic part of the molecule is attached to the joint via a degradable bond. Thus, in several embodiments, the antigen of interest is released at, in, or near the liver (or other target site) and processed and presented to the immune system in a manner that allows the immune system to self-recognize the native antigen (or antigenic fragment or mimetic thereof) and reduce or eliminate the immune response against that antigen.

[0158] In several embodiments, the antigen can be endogenous (e.g., self-antigen or autoantigen) or exogenous (e.g., foreign antigen), including but not limited to: foreign alloantigens, to which the transplant recipient produces an undesirable immune response (e.g., graft-versus-host disease or transplant rejection); foreign food, animal, plant or environmental antigens, to which the patient produces an undesirable immune response (e.g., allergic reaction or hypersensitivity); therapeutic agents, to which the patient produces an undesirable immune response (e.g., hypersensitivity and / or reduced therapeutic activity); self antigens, to which the patient produces an undesirable immune response (e.g., autoimmune disease); or a tolerogenic portion (e.g., fragment or epitope) thereof. The compositions provided herein can be used to induce tolerance to antigens, and can be used to treat undesirable immune responses, e.g., graft-versus-host disease, transplant rejection, immune responses to therapeutic agents, autoimmune diseases and / or allergic reactions, depending on the embodiment.

[0159] Also provided are pharmaceutical compositions containing a therapeutically effective amount of a compound of the present disclosure. In some embodiments, the compound is mixed with at least one pharmaceutically acceptable excipient. In another aspect, the present disclosure provides methods for treating an undesirable immune response, such as graft-versus-host disease, transplant rejection, a response to a therapeutic agent, an autoimmune disease, or an allergic reaction.

[0160] the term

[0161] In the following detailed description, reference is made to the accompanying drawings, which form a part thereof. In the accompanying drawings, similar symbols generally identify similar components unless the context dictates otherwise. The illustrative embodiments described in the detailed description, drawings, and claims are not intended to be limiting. Other embodiments may be utilized, and other changes may be made, without departing from the spirit or scope of the subject matter presented herein. It will be readily understood that the various aspects of the present disclosure, as generally described herein and illustrated in the accompanying drawings, may be arranged, substituted, combined, separated, and designed in a variety of different configurations, all of which are expressly contemplated herein.

[0162] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the subject matter belongs. The terminology used in the description of the subject matter herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the subject matter.

[0163] The articles "a" and "an" are used herein to refer to one or to more than one (e.g., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0164] By "about" is meant a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by up to 10% from a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length.

[0165] In several embodiments, the antigen-binding polypeptide comprises one or more complementary determining regions ("a CDR" or "multiple CDRs"). As used herein, the term "CDR" should be given its ordinary meaning and should also refer to the complementary determining region (also known as the "minimum recognition unit" or "hypervariable region") within the antibody variable region sequence. CDRs allow antigen-binding proteins to specifically bind to a particular antigen of interest. There are three heavy chain variable region CDRs (CDRH1, CDRH2, and CDRH3) and three light chain variable region CDRs (CDRL1, CDRL2, and CDRL3). The CDRs in each of the two chains are usually aligned by the framework region to form a structure that specifically binds to a specific epitope or domain on the target protein. From the N-terminus to the C-terminus, the naturally occurring light chain variable region and the heavy chain variable region generally follow the following order of these elements: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. A numbering system has been designed for assigning numbers to the amino acids that occupy the positions in each of these domains.

[0166] The precise amino acid sequence boundaries of a given CDR or FR can be readily determined using any of a number of well-known schemes, including those described by Kabat et al. (1991), "Sequences of Proteins of Immunological Interest," 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD ("Kabat" numbering scheme); Al-Lazikani et al, (1997) JMB 273, 927-948 ("Chothia" numbering scheme); MacCallum et al, J. Mol. Biol. 262:732-745 (1996), "Antibody-antigen interactions: Contact analysis and binding site topography," J. Mol. Biol. 262, 732-745." ("Contact" numbering scheme); Lefranc MP et al, "IMGT unique numbering for immunoglobulin and T cell receptor variable domains and Ig superfamily V-like domains,"Dev Comp Immunol,2003Jan;27(l):55-77 ("IMGT" numbering scheme); Honegger A and Pliickthun A,"Yet another numbering scheme forimmunoglobulin variable domains:an automatic modeling and analysis tool,"JMol Biol,2001Jun8;309(3):657-70, ("Aho" numbering scheme); and Martin et ah,"Modelingantibody hypervariable loops:a combined algorithm,"PNAS,1989,86(23):9268-9272, ("AbM" numbering scheme).

[0167] The boundaries of a given CDR or FR can vary depending on the scheme used for identification. For example, the Kabat scheme is based on structural alignments, while the Chothia scheme is based on structural information. The numbering of the Kabat and Chothia schemes is based on the most common antibody region sequence lengths, with insertions accommodated by inserted letters (e.g., "30a"), and deletions in some antibodies. These two schemes place certain insertions and deletions ("indels") in different positions, resulting in different numbering. The Contact scheme is based on the analysis of complex crystal structures, and is similar to the Chothia numbering scheme in many respects. The AbM scheme is a compromise between the Kabat and Chothia definitions, based on the scheme used by Oxford Molecular's AbM antibody modeling software.

[0168] Throughout this specification, unless the context requires otherwise, the words "comprises", "comprises" and "comprising" will be understood to imply the inclusion of the stated steps or elements or groups of steps or elements, but not to exclude any other steps or elements or groups of steps or elements. The so-called "consisting of" is meant to include and be limited to whatever follows the phrase "consisting of". Thus, the phrase "consisting of" indicates that the listed elements are required or mandatory, and indicates that other elements may not be present. The so-called "consisting essentially of" is meant to include any element listed after the phrase, and is limited to other elements that do not interfere with or contribute to the activity or effect specified for the listed elements in the present disclosure. Thus, the phrase "consisting essentially of" indicates that the listed elements are required or mandatory, but indicates that other elements are optional and may or may not be present, depending on whether they substantially affect the activity or effect of the listed elements.

[0169] As used herein, the terms "individual", "subject" or "patient" have their simple and ordinary meanings as understood in accordance with this specification, and refer to humans or non-human mammals, for example, dogs, cats, mice, rats, cows, sheep, pigs, goats, non-human primates or birds (e.g., chickens), as well as any other vertebrate or invertebrate. The term "mammal" is used in its usual biological sense. Therefore, it specifically includes, but is not limited to, primates (including apes (chimpanzees, apes, monkeys) and humans), cattle, horses, sheep, goats, pigs, rabbits, dogs, cats, rodents, rats, mice, guinea pigs, and the like.

[0170] As used herein, the term "isolated" has its plain and ordinary meaning as understood in light of this specification, and refers to (1) a substance and / or entity that is separated from at least some of its associated components when originally produced (whether in nature and / or in an experimental setting), and / or (2) a substance and / or entity that is produced, prepared and / or manufactured by the hand of man. An isolated substance and / or entity may be separated from other components with which they were originally associated by equal to, about, at least, at least about, no more than, or no more than about, 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 98%, about 99%, substantially 100%, or 100% (or a range including and / or spanning the foregoing values). In some embodiments, the isolated preparation is, is about, is at least, is at least about, is no more than, or is no more than about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, substantially 100%, or 100% pure (or includes and / or spans a range of the above values). As used herein, an "isolated" material can be "pure" (e.g., substantially free of other components). As used herein, the term "isolated cell" can refer to a cell that is not contained in a multicellular organism or tissue.

[0171] As used herein, "in vivo" is given its plain and ordinary meaning as understood in light of this specification, and refers to methods performed in living organisms, typically animals, mammals (including humans) and plants, rather than tissue extracts or dead organisms.

[0172] As used herein, "ex vivo" is given its plain and ordinary meaning as understood in light of this specification, and refers to methods performed outside a living organism with little change in natural conditions.

[0173] As used herein, "in vitro" is given its plain and ordinary meaning as understood in light of the present specification and refers to methods performed outside of biological conditions (eg, in a petri dish or test tube).

[0174] As used herein, the term "nucleic acid" or "nucleic acid molecule" has its simple and common meaning as understood according to this specification, and refers to polynucleotides, such as deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), oligonucleotides, polynucleotides naturally occurring in cells, fragments generated by polymerase chain reaction (PCR), and fragments generated by any one of ligation, severance, endonuclease action, and exonuclease action. Nucleic acid molecules can be composed of monomers of naturally occurring nucleotides (such as DNA and RNA) or analogs of naturally occurring nucleotides (e.g., enantiomeric forms of naturally occurring nucleotides) or a combination of the two. Modified nucleotides can have changes in sugar moieties and / or pyrimidine or purine base moieties. Sugar modifications include, for example, replacement of one or more hydroxyl groups with halogens, alkyl groups, amines, and azido groups, or sugars can be functionalized as ethers or esters. In addition, the entire sugar moiety can be replaced with spatially and electronically similar structures (such as azasugars and carbocyclic sugar analogs). The example of modification in the base part includes alkylated purine and pyrimidine, acylated purine or pyrimidine or other well-known heterocyclic substitutes. Nucleic acid monomers can be connected by phosphodiester bonds or analogs of such bonds. The analogs of phosphodiester bonds include thiophosphates, dithiophosphates, selenophosphates, diselenphosphates, anilinothiophosphates, anilinophosphates or aminophosphorophosphates. The term "nucleic acid molecule" also includes so-called "peptide nucleic acids", which include naturally occurring or modified nucleic acid bases attached to a polyamide backbone. Nucleic acids can be single-stranded or double-stranded. "Oligonucleotide" can be used interchangeably with nucleic acids, and can refer to double-stranded or single-stranded DNA or RNA. One or more nucleic acids can be contained in a nucleic acid carrier or nucleic acid construct (e.g., a plasmid, a virus, a retrovirus, a lentivirus, a phage, a cosmid, a fosmid, a phagemid, a bacterial artificial chromosome (BAC), a yeast artificial chromosome (YAC), or a human artificial chromosome (HAC)), which can be used to amplify and / or express the one or more nucleic acids in various biological systems. Typically, the carrier or construct will also contain elements including, but not limited to, a promoter, an enhancer, a terminator, an inducer, a ribosome binding site, a translation start site, a start codon, a stop codon, a polyadenylation signal, an origin of replication, a cloning site, a multiple cloning site, a restriction enzyme site, an epitope, a reporter gene, a selectable marker, an antibiotic selectable marker, a targeting sequence, a peptide purification tag, or an auxiliary gene, or any combination thereof.

[0175] Nucleic acid or nucleic acid molecule can comprise one or more sequences encoding different peptides, polypeptides or proteins. These one or more sequences can be adjacently connected in the same nucleic acid or nucleic acid molecule, or this same nucleic acid or nucleic acid molecule has extra nucleic acid therebetween, for example, a joint, recombinant sequence or restriction enzyme site or any other sequence, the length of this any other sequence is, is about, is at least, is at least about, is no more than or is no more than about 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,25,30,35,40,45,50,55,60,65,70,75,80,85,90,95,100,150,200 or 300 bases, or any length within the range limited by any two of the aforementioned lengths. As used herein, the term "downstream" on a nucleic acid has its simple and ordinary meaning as understood in accordance with this specification, and refers to the sequence located after the 3' end of the previous sequence on the strand containing the coding sequence (the sense strand) (if the nucleic acid is double-stranded). As used herein, the term "upstream" on a nucleic acid has its simple and ordinary meaning as understood in accordance with this specification, and refers to the sequence located before the 5' end of the latter sequence on the strand containing the coding sequence (the sense strand) (if the nucleic acid is double-stranded). As used herein, the term "grouping" on nucleic acids has its plain and ordinary meaning as understood in light of this specification, and refers to two or more sequences that appear in proximity, either directly in proximity or with additional nucleic acids in between, e.g., linkers, repeat sequences, or restriction enzyme sites or any other sequence that is, is about, is at least, is at least about, is no more than, or is no more than about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases in length, or any length within the range defined by any two of the foregoing lengths, but typically does not have a sequence encoding a functional or catalytic polypeptide, protein, or protein domain in between.

[0176] Nucleic acids described herein comprise nucleobases. Main, classical, natural or unmodified bases are adenine, cytosine, guanine, thymine and uracil. Other nucleobases include, but are not limited to, purine, pyrimidine, modified nucleobases, 5-methylcytosine, pseudouridine, dihydrouridine, inosine, 7-methylguanosine, hypoxanthine, xanthine, 5,6-dihydrouracil, 5-hydroxymethylcytosine, 5-bromouracil, isoguanine, isocytosine, aminoallyl bases, dye-labeled bases, fluorescent bases or biotin-labeled bases.

[0177] As used herein, the terms "peptide", "polypeptide" and "protein" have their simple and ordinary meanings as understood in light of this specification, and refer to macromolecules composed of amino acids linked by peptide bonds. The various functions of peptides, polypeptides and proteins are known in the art, and include, but are not limited to, enzymatic, structural, transport, defense, hormone or signal transduction. Peptides, polypeptides and proteins are usually, but not always, produced biologically by ribosome complexes using nucleic acid templates, although chemical synthesis may also be used. By manipulating nucleic acid templates, peptide, polypeptide and protein mutations, such as substitution, deletion, truncation, addition, duplication or fusion of more than one peptide, polypeptide or protein, can be performed. These fusions of more than one peptide, polypeptide or protein can be contiguously linked in the same nucleic acid or nucleic acid molecule, or the same nucleic acid or nucleic acid molecule has additional nucleic acids therebetween, e.g., linkers, repeat sequences, epitopes or tags, or any other sequence that is, is about, is at least, is at least about, is no more than, or is no more than about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases in length, or any length within the range defined by any two of the foregoing lengths. As used herein, the term "downstream" on a polypeptide has its simple and ordinary meaning as understood in light of this specification, and refers to a sequence that is located after the C-terminus of the previous sequence. As used herein, the term "upstream" on a polypeptide has its plain and ordinary meaning as understood in light of the present specification, and refers to a sequence located before the N-terminus of a latter sequence.

[0178] As used herein, "antigen" should have its simple and common meaning, and should refer to any substance used as a target of a receptor for an innate or adaptive immune response, such as a T cell receptor, major histocompatibility complex class I and class II, a B cell receptor or an antibody. In some embodiments, the antigen can be derived from in vivo (e.g., "self," "autologous" or "endogenous"). In additional embodiments, the antigen can be derived from in vitro ("non-self," "foreign" or "exogenous"), such as by inhalation, ingestion, injection or transplantation, percutaneous entry, etc. In some embodiments, exogenous antigens can be biochemically modified in vivo. Foreign antigens include, but are not limited to, food antigens, animal antigens, plant antigens, environmental antigens, therapeutic agents, and antigens present in allografts.

[0179] As used herein, the term "epitope", also known as an antigenic determinant, shall have its simple and common meaning, and shall refer to a segment of a macromolecule (such as a protein) recognized by the immune system, such as by an antibody, a B cell, a major histocompatibility complex molecule, or a T cell. An epitope can be recognized by, for example, an antibody or a B cell, and may include a portion or segment of a macromolecule that can bind to an antibody or an antigen-binding fragment thereof. In this context, the term "binding" particularly relates to specific binding. In the context of several embodiments of the present invention, it is preferred that the term "epitope" refers to a segment of a protein or a polyprotein recognized by the immune system. In several embodiments, the "antigen" used for the constructs disclosed herein may include one or more epitopes. In some embodiments including more than one epitope, the additional epitopes may be from the same or different antigens.

[0180] A peptide that specifically binds to a particular target is called a "ligand" for that target.

[0181] As a commonly used term in the field of biology, "specific binding" refers to a molecule that binds to a target with a relatively high affinity compared to non-target tissues, and generally involves a variety of non-covalent interactions, such as electrostatic interactions, van der Waals interactions, hydrogen bonds, etc. Specific binding interactions characterize antibody-antigen binding, enzyme-substrate binding, and certain protein-receptor interactions; although such molecules may sometimes bind to tissues other than their specific targets, to some extent, such non-target binding is unimportant because high-affinity binding pairs can still fall within the definition of specific binding.

[0182] As used herein, the term "conservative changes" shall have its simple and common meaning and refers to changes that can be made to an amino acid sequence generally without changing the activity. These changes are referred to as "conservative substitutions" or mutations; that is, an amino acid belonging to an amino acid group with a specific size or characteristic can be substituted for another amino acid. Substitutes for an amino acid sequence can be selected from other members of the class to which the amino acid belongs. For example, non-polar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, methionine, and tyrosine. Polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine. Positively charged (basic) amino acids include arginine, lysine, and histidine. Negatively charged (acidic) amino acids include aspartic acid and glutamic acid. It is expected that such substitutions will not significantly affect the apparent molecular weight as determined by polyacrylamide gel electrophoresis or isoelectric point. Conservative substitutions also include replacing optical isomers of a sequence with other optical isomers, particularly replacing L-amino acids with D-amino acids for one or more residues of the sequence. In addition, all amino acids in the sequence can be subjected to D- to L-isomer substitutions. Examples of conservative substitutions include, but are not limited to, replacing Arg with Lys, and vice versa, to maintain a positive charge; replacing Asp with Glu, and vice versa, to maintain a negative charge; replacing Thr with Ser, so as to maintain a free -OH; and replacing Asn with Gln, to maintain a free -NH2. Yet another type of conservative substitution constitutes the following situation: if chemical derivatization is required to occur, an amino acid with the desired chemical reactivity is introduced to confer a reactive site for a chemical conjugation reaction. Such amino acids include, but are not limited to, Cys (to insert a thiol group), Lys (to insert a primary amine), Asp and Glu (to insert a carboxylic acid group) or specialized non-classical amino acids containing ketones, azides, alkynes, alkenes, and tetrazine side chains. Conservative substitution or addition of amino acids with free -NH2 or -SH can be particularly advantageous for chemical conjugation with a linker. In addition, in some cases, point mutations, deletions and insertions of a polypeptide sequence or corresponding nucleic acid sequence can be performed without loss of function of the polypeptide or nucleic acid fragment. Substitutions can include, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50 or more residues (including any number of substitutions between the listed substitutions).Variants useful in the present invention may exhibit a total of up to 200 changes (e.g., exchanges, insertions, deletions, N-terminal truncations, and / or C-terminal truncations) in the amino acid sequence (e.g., up to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, or 200, including any number between the listed numbers). In several embodiments, the number of changes is greater than 200. Additionally, in several embodiments, variants include polypeptide sequences or corresponding nucleic acid sequences that exhibit a degree of functional equivalence to a reference (e.g., an unmodified or native sequence). In several embodiments, variants exhibit about 80%, about 85%, about 90%, about 95%, about 97%, about 98%, about 99% functional equivalence (and any degree of functional equivalence between the listed functional equivalence) to an unmodified or native reference sequence. The amino acid residues described herein are represented by single-letter amino acid designators or three-letter abbreviations to be consistent with standard polypeptide nomenclature. In this article, all amino acid residue sequences are represented by formulas with left-right orientation in the conventional direction from the amino terminus to the carboxyl terminus.

[0183] The term "sequence identity" is used in relation to polypeptide or nucleic acid sequence comparisons. This expression refers in particular to the percentage of sequences identical to the corresponding reference polypeptide or nucleic acid, e.g., at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, after aligning the sequences and introducing spaces (if necessary) to achieve the maximum sequence identity percentage, and without taking into account any conservative substitutions as part of the sequence identity. Alignment for the purpose of determining the percentage of amino acid sequence identity can be accomplished in a variety of known ways, e.g., using publicly available computer software. Appropriate parameters for aligning sequences can be determined, including any algorithm required for achieving maximum alignment over the full length of the compared sequences.

[0184] As used herein, the term "purity" of any given substance, compound or material has its simple and common meaning as understood according to this specification, and refers to the actual abundance of the substance, compound or material relative to the expected abundance. For example, the substance, compound or material can be at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% pure, including all decimals therebetween. Purity may be affected by undesirable impurities, including but not limited to nucleic acids, DNA, RNA, nucleotides, proteins, polypeptides, peptides, amino acids, lipids, cell membranes, cell debris, small molecules, degradation products, solvents, carriers, carriers or pollutants or any combination thereof. In some embodiments, the substance, compound or material is substantially free of host cell proteins, host cell nucleic acids, plasmid DNA, contaminating viruses, proteasomes, host cell culture components, process-related components, mycoplasmas, pyrogens, bacterial endotoxins and exogenous agents. Purity can be measured using techniques including, but not limited to, electrophoresis, SDS-PAGE, capillary electrophoresis, PCR, rtPCR, qPCR, chromatography, liquid chromatography, gas chromatography, thin layer chromatography, enzyme-linked immunosorbent assay (ELISA), spectroscopy, UV-visible spectroscopy, infrared spectroscopy, mass spectrometry, nuclear magnetic resonance, gravimetric analysis, or titration, or any combination thereof.

[0185] As used herein, the term "yield" of any given substance, compound, or material has its simple and ordinary meaning as understood in light of this specification, and refers to the actual total amount of the substance, compound, or material relative to the expected total amount. For example, the yield of a substance, compound, or material is, is about, is at least, is at least about, is more than, or is no more than about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the expected total amount, including all decimals therebetween. Yield may be affected by the efficiency of a reaction or process, undesirable side reactions, degradation, the amount of substance, compound, or material input, or the loss of a desired substance, compound, or material during any step of production.

[0186] As used herein, the term "effective amount" or "effective dose" has its simple and common meaning as understood in accordance with this specification, and refers to the amount of the recited composition or compound that results in an observable effect. For example, an effective amount can refer to an amount of a composition or compound that improves the condition of a subject by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 100%. The actual dosage level of the active ingredient in the active composition of the present disclosure subject can vary so that the amount of the active composition or compound administered can effectively achieve the desired response for a particular subject and / or application. The dosage level selected will depend on a variety of factors, including but not limited to the activity of the composition, formulation, route of administration, combination with other drugs or treatments, severity of the condition being treated, and physical condition and prior medical history of the subject being treated. In some embodiments, a minimum dose is administered, and the dose is escalated to the minimum effective amount in the absence of dose-limiting toxicity. The determination and adjustment of the effective dose, as well as the assessment of when and how to make such adjustments, are contemplated herein.

[0187] As used herein, the terms "function" and "functional" have their plain and ordinary meaning as understood in light of the present specification, and refer to a biological function, an enzyme function, or a therapeutic function.

[0188] As used herein, the term "inhibit" has its simple and common meaning as understood according to this specification, and can refer to the reduction or prevention of biological activity. The percentage of reduction can be, about, at least, at least about, not more than or not more than about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100%, or an amount within the range defined by any two of the aforementioned values. As used herein, the term "delay" has its simple and common meaning as understood according to this specification, and refers to slowing down, delaying or postponing a biological event to a time that would otherwise be later than expected. Delay can be the percentage of the following delay: for, about, at least, at least about, not more than or not more than about 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80% or 90%, 100%, or an amount within the range defined by any two of the aforementioned values. The terms inhibition and delay may not necessarily mean 100% inhibition or delay. Partial inhibition or delay may be achieved.

[0189] As used herein, "pharmaceutically acceptable" has its simple and common meaning as understood according to this specification, and refers to carriers, excipients and / or stabilizers that are not toxic to cells or mammals exposed to them at the dosages and concentrations employed, or have an acceptable level of toxicity. As used herein, "pharmaceutically acceptable", "diluents", "excipients" and / or "carriers" have their simple and common meanings as understood according to this specification, and are intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents and absorption delaying agents, etc., compatible with administration to humans, cats, dogs or other vertebrate hosts. Typically, pharmaceutically acceptable diluents, excipients and / or carriers are diluents, excipients and / or carriers approved by federal regulatory agencies, state governments or other regulatory agencies, or listed in the U.S. Pharmacopeia or other recognized pharmacopeias for animals, including humans and non-human mammals, such as cats and dogs. The term diluent, excipient / " carrier" can refer to a diluent, adjuvant, excipient or vehicle used together with a pharmaceutical composition. Such pharmaceutical diluents, excipients and / or carriers can be sterile liquids, such as water and oil, including oils from petroleum, animals, plants or synthetic sources. Water, saline solutions and aqueous dextrose and glycerol solutions can be used as liquid diluents, excipients and / or carriers, particularly for injectable solutions. Suitable pharmaceutical diluents and / or excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glyceryl monostearate, talcum, sodium chloride, skimmed milk powder, glycerine, propylene glycol, ethylene glycol, water, ethanol etc. The non-limiting example of physiologically acceptable carriers is an aqueous pH buffer solution. Physiologically acceptable carriers may also include one or more of the following: antioxidants such as ascorbic acid; low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin, immunoglobulins; hydrophilic polymers such as polyvinyl pyrrolidone; amino acids; carbohydrates such as glucose, mannose or dextrin; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt-forming counterions such as sodium; and nonionic surfactants such as Polyethylene glycol (PEG) and If desired, the composition may also contain a small amount of a wetting agent, bulking agent, emulsifier or pH buffer. These compositions may take the form of solutions, suspensions, emulsions, sustained-release formulations, etc. The formulation is generally suitable for the mode of administration.

[0190] Cryoprotectants are additives used to improve the efficiency and yield of cryopreservation by preventing the formation of large ice crystals. Cryoprotectants include, but are not limited to, DMSO, ethylene glycol, glycerol, propylene glycol, trehalose, formamide, methylformamide, dimethylformamide, 3-phosphoglycerol, proline, sorbitol, diethylene glycol, sucrose, triethylene glycol, polyvinyl alcohol, polyethylene glycol, or hydroxyethyl starch. For example, it can be found that the concentration of at least one cryoprotectant is, is about, is at least, is at least about, is no more than, or is no more than about 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, 80% or 90%, or any percentage within a range defined by any two of the foregoing numbers.

[0191] Additional excipients having desired properties include, but are not limited to, preservatives, adjuvants, stabilizers, solvents, buffers, diluents, solubilizers, detergents, surfactants, chelating agents, antioxidants, alcohols, ketones, aldehydes, ethylenediaminetetraacetic acid (EDTA), citric acid, salts, sodium chloride, sodium bicarbonate, sodium phosphate, sodium borate, sodium citrate, potassium chloride, potassium phosphate, magnesium sulfate, sugar, dextrose, fructose, mannose, lactose, galactose, sucrose, sorbitol, cellulose, serum, amino acids, polysorbate 20, polysorbate 80, sodium deoxycholate, sodium taurodeoxycholate, magnesium stearate, octylphenol ethoxylate, benzethonium chloride, thimerosal, gelatin, esters, ethers, 2-phenoxyethanol, urea, or vitamins, or any combination thereof. Some excipients may be present in residual amounts or contaminants from the manufacturing process, including, but not limited to, serum, albumin, ovalbumin, antibiotics, inactivators, formaldehyde, glutaraldehyde, β-propiolactone, gelatin, cell debris, nucleic acids, peptides, amino acids, or growth medium components or any combination thereof. For example, it can be found that the amount of excipient in the composition (as a percentage) is, is about, is at least, is at least about, is not more than, or is not more than about 0%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 100% w / w, or any percentage (by weight) within the range defined by any two of the foregoing numbers.

[0192] The term "pharmaceutically acceptable salt" has its simple and common meaning as understood in this specification, and includes relatively non-toxic inorganic and organic acid or base addition salts of compositions or excipients, including but not limited to analgesics, therapeutic agents, other materials, etc. Examples of pharmaceutically acceptable salts include salts derived from mineral acids such as hydrochloric acid and sulfuric acid, and salts derived from organic acids such as ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, etc. Examples of suitable inorganic bases for forming salts include hydroxides, carbonates and bicarbonates of ammonia, sodium, lithium, potassium, calcium, magnesium, aluminum, zinc, etc. Salts can also be formed by suitable organic bases, including organic bases that are non-toxic and strong enough to form such salts. For example, the class of such organic bases can include, but are not limited to, mono-, di- and trialkylamines, including methylamine, dimethylamine and triethylamine; mono-, di- or trihydroxyalkylamines, including mono-, di- and triethanolamine; amino acids, including glycine, arginine and lysine; guanidine; N-methylglucamine; N-methylglucosamine; L-glutamine; N-methylpiperazine; morpholine; ethylenediamine; N-benzylphenethylamine; trishydroxymethylaminoethane.

[0193] Appropriate formulations depend on the selected route of administration. The technology for the preparation and administration of the compounds described herein is known to those skilled in the art. A variety of techniques for administering compounds exist in the art, including but not limited to enteral, oral, rectal, external, sublingual, oral, intraauricular, epidural, applied skin, aerosol, parenteral delivery, including intramuscular, subcutaneous, intraarterial, intravenous, intraportal, intraarticular, intradermal, peritoneal, intramedullary injection, intrathecal, direct intraventricular, intraperitoneal, intranasal or intraocular injection. Pharmaceutical compositions can usually be customized according to specific expected routes of administration.

[0194] As used herein, "carrier" has its plain and ordinary meaning as understood in this specification, and refers to a compound, particle, solid, semisolid, liquid or diluent that facilitates the passage, delivery and / or incorporation of a compound into cells, tissues and / or organs of the body.

[0195] As used herein, "diluent" has its simple and common meaning as understood in this specification, and refers to an ingredient in a pharmaceutical composition that lacks pharmacological activity but may be pharmaceutically necessary or desirable. For example, a diluent may be used to increase the volume of a potent drug whose mass is too small for manufacture and / or administration. It may also be a liquid that is used to dissolve a drug to be administered by injection, ingestion, or inhalation. A common form of diluent in the art is an aqueous buffer solution, such as, but not limited to, phosphate buffered saline that simulates the composition of human blood.

[0196] As used herein, the terms "treat," "treating," and "treatment" shall have their plain and ordinary meanings and refer to any type of action that imparts a modulatory effect, which may be, for example, a beneficial effect on a subject suffering from a disorder, disease, or condition, including improving the subject's condition (e.g., improving one or more symptoms), delaying or reducing the progression of the condition, and / or changing clinical parameters, diseases, or conditions, curing the patient, etc. In some embodiments, treatment may include one or more of preventing or prophylaxis of a disease or condition, not developing clinical symptoms, inhibiting a disease or condition, arresting or inhibiting the development of clinical symptoms, alleviating a disease or condition, and / or causing regression of clinical symptoms. In certain embodiments, treating a subject achieves one, two, three, four or more of the following effects, including, for example: (i) reducing or ameliorating the severity of a disease condition or symptoms associated therewith; (ii) reducing the duration of symptoms associated with a disease or immune response; (iii) preventing the progression of a disease or symptoms associated therewith; (iv) regression of a disease or symptoms associated therewith; (v) preventing the development or onset of symptoms associated with a disease; (vi) preventing the recurrence of symptoms associated with a disease; (vii) reducing the subject's hospitalization; (viii) reducing the length of hospitalization; (ix) increasing the survival rate of subjects with the disease; (x) reducing the number of symptoms associated with the disease; (xi) enhancing, improving, supplementing, complementing or increasing the preventive or therapeutic effect of another therapy.

[0197] As used herein, the term "tolerogen" refers to a molecule, composition or substance that induces an individual's immune tolerance to the molecule, composition or substance or its antigenic portion. Typically, when a substance (e.g., an allergen, a foreign peptide, an organism or its antigenic portion) is introduced into an individual, an immune response to the substance is elicited. This has an impact on diseases (e.g., autoimmune diseases (e.g., type 1 diabetes, multiple sclerosis)) in which the individual's immune system produces an inappropriate response to self-proteins. Administration of tolerogens ("tolerogen therapy") is intended to reduce or alleviate this undesirable immune response, thereby serving as treatment or prevention. The liver plays an important immunomodulatory role because hepatocytes and other resident liver cells show the ability to take up molecules that may be antigenic under normal circumstances, or may be abnormally antigenic in the case of autoimmune diseases, to induce immune tolerance.

[0198] As used herein, the term "liver targeting moiety" refers to a moiety that has the ability to direct an agent (e.g., an immune tolerance inducing construct, a polypeptide, etc.) to the liver. The liver contains different cell types, including but not limited to hepatocytes, sinusoidal epithelial cells, Kupffer cells, stellate cells, and / or dendritic cells. Typically, the liver targeting moiety directs the polypeptide to one or more of these cells. On the surface of each liver cell, there are receptors that recognize and specifically bind to the liver targeting moiety. Liver targeting can be achieved, for example, by chemically conjugating an antigen or ligand to a galactosylated or glucosylated moiety, desialylating an antigen or ligand to expose a potential galactosyl or glucosyl moiety, or specifically binding an antibody to an antigen or ligand associated with the liver, e.g., ASGPR.

[0199] As used herein, the term "operably linked" should be given its ordinary meaning. In some embodiments, by way of illustration, when two groups are operably linked, the groups are attached so as to provide one or more of the linked groups without substantial loss of their native reactivity or activity. In some embodiments, an antigen disclosed herein is operably linked to a linker and a targeting agent.

[0200] As used herein, the term "undesirable immune response" refers to a reaction by a subject's immune system that is undesirable in a given situation. Typically, if the immune system's reaction is directed to an inappropriate target, it can cause, enhance, or worsen a disease. For example, undesirable immune responses include, but are not limited to, transplant rejection, immune responses to therapeutic agents, autoimmune diseases, and allergic or hypersensitivity reactions.

[0201] The term "variant" is understood to be a protein (or nucleic acid) that has one or more changes in length, sequence or structure compared to the protein (or nucleic acid) from which it is derived. The polypeptide derived from a protein variant is also referred to as a parent polypeptide or polynucleotide. The term "variant" comprises a "fragment" or "derivative" of a parent molecule. Typically, the length or size of a "fragment" is less than that of a parent molecule, while a "derivative" exhibits one or more differences in its sequence or structure compared to a parent molecule. Modified molecules are also encompassed, such as, but not limited to, post-translationally modified proteins (e.g., glycosylated, phosphorylated, ubiquitinated, palmitoylated or proteolytically cleaved proteins) and modified nucleic acids (such as methylated DNA). The term "variant" also encompasses a mixture of different molecules, such as, but not limited to, RNA-DNA hybrids. As used herein, naturally occurring and artificially constructed variants are understood to be encompassed by the term "variant". Further, variants that can be used in the present invention can also be derived from homologues, orthologues or paralogues of the parent molecule, or derived from artificially constructed variants, as long as the variant exhibits at least one biological activity (e.g., functional activity) of the parent molecule. The variant is characterized in that it has a certain degree of sequence identity with the parent polypeptide from which it is derived. More precisely, protein variants in the context of the present disclosure can exhibit at least 80% sequence identity with its parent polypeptide. Preferably, the sequence identity of protein variants is on a continuous stretch of 20, 30, 40, 45, 50, 60, 70, 80, 90, 100 or more amino acids. As discussed above, in several embodiments, variants exhibit about 80%, about 85%, about 90%, about 95%, about 97%, about 98%, about 99% functional equivalence (and any degree of functional equivalence between the listed functional equivalences) with unmodified or natural reference sequences.

[0202] As used herein, the term "% w / w" or "% wt / wt" has its plain and ordinary meaning as understood in light of this specification, and refers to the percentage expressed as the weight of an ingredient or agent to the total weight of the composition multiplied by 100. As used herein, the term "% v / v" or "% vol / vol" has its plain and ordinary meaning as understood in light of this specification, and refers to the percentage expressed as the liquid volume of a compound, substance, ingredient or agent to the total liquid volume of the composition multiplied by 100.

[0203] The disclosure herein generally uses affirmative language to describe numerous embodiments. The disclosure also includes embodiments in which subject matter is excluded in whole or in part, such as substances or materials, method steps and conditions, schemes or procedures.

[0204] Liver-specific binding peptide

[0205] Disclosed herein are compositions of binding polypeptides that bind to an asialoglycoprotein receptor (ASGPR) or a component thereof. In some embodiments, the component of the ASGPR is asialoglycoprotein receptor 1 (ASGR1) or asialoglycoprotein receptor 2 (ASGR2).

[0206] Additional anti-ASGPR antibodies are contemplated in US Patents 9,771,427 and 10,358,497, each of which is expressly incorporated herein by reference in its entirety.

[0207] Additional liver-targeted compositions for immune tolerance, which may or may not be polypeptides, are explored in U.S. Patent Nos. 10,046,056, 10,821,157, 10,940,209, 10,946,079, and 10,953,101, each of which is expressly incorporated herein by reference in its entirety. Further liver-targeted compositions for immune tolerance, which may or may not be polypeptides, are explored in WO 2021 / 053589, which is expressly incorporated herein by reference in its entirety.

[0208] ASGR1 binding polypeptides are provided herein. In some embodiments, the ASGR1 binding polypeptide comprises a heavy chain variable region. In some embodiments, the heavy chain variable region comprises a heavy chain complementary determining region 1 (HCDR1). In some embodiments, the heavy chain variable region comprises HCDR2. In some embodiments, the heavy chain variable region comprises HCDR3. In some embodiments, the heavy chain variable region comprises HCDR1, HCDR2, and HCDR3. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:9. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:297. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:329. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:361. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:393. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:425. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:457. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:489. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:521. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:553. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:10. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:298. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:330. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:362. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:394. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:426. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:458. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:490. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:522. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:554. In some embodiments, HCDR3 comprises a sequence of SEQ ID NO:11. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 299. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 331. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 363. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 395.In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 427. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 459. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 491. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 523. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO: 555. In some embodiments, the HCDR1 comprises the sequence of SEQ ID NO: 9, 297, 329, 361, 393, 425, 457, 489, 521, or 553, the HCDR2 comprises the sequence of SEQ ID NO: 10, 298, 330, 362, 394, 426, 458, 490, 522, or 554, and the HCDR3 comprises the sequence of SEQ ID NO: 11, 299, 331, 363, 395, 427, 459, 491, 523, or 555. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 15. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 303. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 335. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 367. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:399.In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 431. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 463. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 495. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 527. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 559. In some embodiments, the ASGR1 binding polypeptide comprises (or further comprises in combination with the heavy chain variable region) a light chain variable region. In some embodiments, the light chain variable region comprises a light chain CDR1 (LCDR1). In some embodiments, the light chain variable region comprises LCDR2. In some embodiments, the light chain variable region comprises LCDR3. In some embodiments, the light chain variable region comprises light chain CDRs LCDR1, LCDR2 and LCDR3. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 25. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 313. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 345. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 377. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 409. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 441.In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:473. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:505. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:537. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:569. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:26. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:314. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:346. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:378. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:410. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:442. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:474. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:506. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:538. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:570. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:27. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:315. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:347. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:379. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:411. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:443. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:475. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:507. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:539. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:571. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:25, 313, 345, 377, 409, 441, 473, 505, 537 or 569, LCDR2 comprises the sequence of SEQ ID NO:26, 314, 346, 378, 410, 442, 474, 506, 538 or 570, and LCDR3 comprises the sequence of SEQ ID NO:27, 315, 347, 379, 411, 443, 475, 507, 539 or 571.In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 31. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 319. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 351. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 383. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 415. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 447. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 479. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 511.In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 543. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 575. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 15, and the light chain variable region comprises the sequence of SEQ ID NO: 31. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 303, and the light chain variable region comprises the sequence of SEQ ID NO: 319. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 335, and the light chain variable region comprises the sequence of SEQ ID NO: 351. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 367, and the light chain variable region comprises the sequence of SEQ ID NO: 383. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 399, and the light chain variable region comprises the sequence of SEQ ID NO: 415. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 431, and the light chain variable region comprises the sequence of SEQ ID NO: 447. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 463, and the light chain variable region comprises the sequence of SEQ ID NO: 479. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 495, and the light chain variable region comprises the sequence of SEQ ID NO: 511. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 527, and the light chain variable region comprises the sequence of SEQ ID NO: 543. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 559, and the light chain variable region comprises the sequence of SEQID NO: 575. In some embodiments, the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb) or a scFv. In some embodiments, the ASGR1 binding polypeptide comprises an Fc domain. In some embodiments, the Fc domain is silenced. In some embodiments, the Fc domain is partially silenced.

[0209] Also disclosed herein are polynucleotides encoding ASGR1 binding polypeptides. In some embodiments, the polynucleotides comprise one or more of the sequences of SEQ ID NOs: 12-14 or 25-27. In some embodiments, the polynucleotides comprise one or more of the sequences of SEQ ID NOs: 300-302 or 313-315. In some embodiments, the polynucleotides comprise one or more of the sequences of SEQ ID NOs: 332-334 or 345-347. In some embodiments, the polynucleotides comprise one or more of the sequences of SEQ ID NOs: 364-366 or 377-379. In some embodiments, the polynucleotides comprise one or more of the sequences of SEQ ID NOs: 396-398 or 409-411. In some embodiments, the polynucleotides comprise one or more of the sequences of SEQ ID NOs: 428-430 or 441-443. In some embodiments, the polynucleotide comprises one or more of the sequences of SEQID NO:460-462 or 473-475. In some embodiments, the polynucleotide comprises one or more of the sequences of SEQID NO:492-494 or 505-507. In some embodiments, the polynucleotide comprises one or more of the sequences of SEQID NO:524-526 or 537-539. In some embodiments, the polynucleotide comprises one or more of the sequences of SEQID NO:556-558 or 569-571. In some embodiments, the polynucleotide comprises the sequence of SEQID NO:16, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO:15. In some embodiments, the polynucleotide comprises the sequence of SEQ ID NO:304, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO:303. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 336, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 335. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 368, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 367. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 400, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 399. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 432, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 431.In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 464, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 463. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 496, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 495. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 528, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 527. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 560, which is a non-limiting example of a polynucleotide sequence encoding a heavy chain variable region sequence of SEQ ID NO: 559. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 32, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 31. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 320, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 319. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 352, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 351. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 384, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 383. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 416, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 415. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 448, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 447. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 480, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 479. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 512, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 511. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 544, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 543. In some embodiments, the polynucleotide comprises a sequence of SEQ ID NO: 576, which is a non-limiting example of a polynucleotide sequence encoding a light chain variable region sequence of SEQ ID NO: 575.In some embodiments, the polynucleotide encodes any one of the ASGR1 binding polynucleotides disclosed herein.

[0210] When applied to the ASGR1 binding polypeptides disclosed herein and the polynucleotides encoding the same, the ASGR1 binding polypeptides may further comprise an immunoglobulin framework. These immunoglobulin frameworks may be commonly known in the art. For example, in several embodiments, the heavy chain variable region framework and the light chain variable region framework used each have four framework sequences (FW-1, FW-2, FW-3, FW-4), wherein three CDRs are provided.

[0211] A non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 1 and may be encoded by the polynucleotide provided in SEQ ID NO: 5. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 289 and may be encoded by the polynucleotide provided in SEQ ID NO: 293. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 321 and may be encoded by the polynucleotide provided in SEQ ID NO: 325. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 353 and may be encoded by the polynucleotide provided in SEQ ID NO: 357. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 385 and may be encoded by the polynucleotide provided in SEQ ID NO: 389. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 417 and may be encoded by the polynucleotide provided in SEQ ID NO: 421. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 449 and may be encoded by the polynucleotide provided in SEQ ID NO: 453. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 481 and may be encoded by the polynucleotide provided in SEQ ID NO: 485. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 513 and may be encoded by the polynucleotide provided in SEQ ID NO: 517. Another non-limiting example of a heavy chain framework 1 (H-FR1) is provided in SEQ ID NO: 545 and may be encoded by the polynucleotide provided in SEQ ID NO: 549. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 2 and may be encoded by the polynucleotide provided in SEQ ID NO: 6. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 290 and may be encoded by the polynucleotide provided in SEQ ID NO: 294. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 322 and may be encoded by the polynucleotide provided in SEQ ID NO: 326. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 354 and may be encoded by the polynucleotide provided in SEQ ID NO: 358.A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 386 and may be encoded by the polynucleotide provided in SEQ ID NO: 390. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 418 and may be encoded by the polynucleotide provided in SEQ ID NO: 422. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 450 and may be encoded by the polynucleotide provided in SEQ ID NO: 454. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 482 and may be encoded by the polynucleotide provided in SEQ ID NO: 486. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 514 and may be encoded by the polynucleotide provided in SEQ ID NO: 518. A non-limiting example of a heavy chain framework 2 (H-FR2) is provided in SEQ ID NO: 546 and can be encoded by the polynucleotide provided in SEQ ID NO: 550. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 3 and can be encoded by the polynucleotide provided in SEQ ID NO: 7. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 291 and can be encoded by the polynucleotide provided in SEQ ID NO: 295. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 323 and can be encoded by the polynucleotide provided in SEQ ID NO: 327. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 355 and can be encoded by the polynucleotide provided in SEQ ID NO: 359. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 387 and may be encoded by the polynucleotide provided in SEQ ID NO: 391. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 419 and may be encoded by the polynucleotide provided in SEQ ID NO: 423. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 451 and may be encoded by the polynucleotide provided in SEQ ID NO: 455. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 483 and may be encoded by the polynucleotide provided in SEQ ID NO: 487. A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 515 and may be encoded by the polynucleotide provided in SEQ ID NO: 519.A non-limiting example of a heavy chain framework 3 (H-FR3) is provided in SEQ ID NO: 547 and can be encoded by the polynucleotide provided in SEQ ID NO: 551. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 4 and can be encoded by the polynucleotide provided in SEQ ID NO: 8. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 292 and can be encoded by the polynucleotide provided in SEQ ID NO: 296. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 324 and can be encoded by the polynucleotide provided in SEQ ID NO: 328. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 356 and can be encoded by the polynucleotide provided in SEQ ID NO: 360. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 388 and may be encoded by the polynucleotide provided in SEQ ID NO: 392. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 420 and may be encoded by the polynucleotide provided in SEQ ID NO: 424. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 452 and may be encoded by the polynucleotide provided in SEQ ID NO: 456. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 484 and may be encoded by the polynucleotide provided in SEQ ID NO: 488. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 516 and may be encoded by the polynucleotide provided in SEQ ID NO: 520. A non-limiting example of a heavy chain framework 4 (H-FR4) is provided in SEQ ID NO: 548 and can be encoded by the polynucleotide provided in SEQ ID NO: 552. A non-limiting example of a light chain framework 1 (L-FR1) is provided in SEQ ID NO: 17 and can be encoded by the polynucleotide provided in SEQ ID NO: 21. A non-limiting example of a light chain framework 1 (L-FR1) is provided in SEQ ID NO: 305 and can be encoded by the polynucleotide provided in SEQ ID NO: 309. A non-limiting example of a light chain framework 1 (L-FR1) is provided in SEQ ID NO: 337 and can be encoded by the polynucleotide provided in SEQ ID NO: 341. A non-limiting example of a light chain framework 1 (L-FR1) is provided in SEQ ID NO: 369 and can be encoded by the polynucleotide provided in SEQ ID NO: 373.Non-limiting examples of light chain framework 1 (L-FR1) are provided in SEQ ID NO: 401 and can be encoded by the polynucleotide provided in SEQ ID NO: 405. Non-limiting examples of light chain framework 1 (L-FR1) are provided in SEQ ID NO: 433 and can be encoded by the polynucleotide provided in SEQ ID NO: 437. Non-limiting examples of light chain framework 1 (L-FR1) are provided in SEQ ID NO: 465 and can be encoded by the polynucleotide provided in SEQ ID NO: 469. Non-limiting examples of light chain framework 1 (L-FR1) are provided in SEQ ID NO: 497 and can be encoded by the polynucleotide provided in SEQ ID NO: 501. Non-limiting examples of light chain framework 1 (L-FR1) are provided in SEQ ID NO: 529 and can be encoded by the polynucleotide provided in SEQ ID NO: 533. A non-limiting example of a light chain framework 1 (L-FR1) is provided in SEQ ID NO: 561 and can be encoded by the polynucleotide provided in SEQ ID NO: 565. A non-limiting example of a light chain framework 2 (L-FR2) is provided in SEQ ID NO: 18 and can be encoded by the polynucleotide provided in SEQ ID NO: 22. A non-limiting example of a light chain framework 2 (L-FR2) is provided in SEQ ID NO: 306 and can be encoded by the polynucleotide provided in SEQ ID NO: 310. A non-limiting example of a light chain framework 2 (L-FR2) is provided in SEQ ID NO: 338 and can be encoded by the polynucleotide provided in SEQ ID NO: 342. A non-limiting example of a light chain framework 2 (L-FR2) is provided in SEQ ID NO: 370 and can be encoded by the polynucleotide provided in SEQ ID NO: 374. Non-limiting examples of light chain framework 2 (L-FR2) are provided in SEQ ID NO: 402 and can be encoded by the polynucleotide provided in SEQ ID NO: 406. Non-limiting examples of light chain framework 2 (L-FR2) are provided in SEQ ID NO: 434 and can be encoded by the polynucleotide provided in SEQ ID NO: 438. Non-limiting examples of light chain framework 2 (L-FR2) are provided in SEQ ID NO: 466 and can be encoded by the polynucleotide provided in SEQ ID NO: 470. Non-limiting examples of light chain framework 2 (L-FR2) are provided in SEQ ID NO: 498 and can be encoded by the polynucleotide provided in SEQ ID NO: 502. Non-limiting examples of light chain framework 2 (L-FR2) are provided in SEQ ID NO: 530 and can be encoded by the polynucleotide provided in SEQ ID NO: 534.A non-limiting example of a light chain framework 2 (L-FR2) is provided in SEQ ID NO: 562 and can be encoded by the polynucleotide provided in SEQ ID NO: 566. A non-limiting example of a light chain framework 3 (L-FR3) is provided in SEQ ID NO: 19 and can be encoded by the polynucleotide provided in SEQ ID NO: 23. A non-limiting example of a light chain framework 3 (L-FR3) is provided in SEQ ID NO: 307 and can be encoded by the polynucleotide provided in SEQ ID NO: 311. A non-limiting example of a light chain framework 3 (L-FR3) is provided in SEQ ID NO: 339 and can be encoded by the polynucleotide provided in SEQ ID NO: 343. A non-limiting example of a light chain framework 3 (L-FR3) is provided in SEQ ID NO: 371 and can be encoded by the polynucleotide provided in SEQ ID NO: 375. Non-limiting examples of light chain framework 3 (L-FR3) are provided in SEQ ID NO: 403 and can be encoded by the polynucleotide provided in SEQ ID NO: 407. Non-limiting examples of light chain framework 3 (L-FR3) are provided in SEQ ID NO: 435 and can be encoded by the polynucleotide provided in SEQ ID NO: 439. Non-limiting examples of light chain framework 3 (L-FR3) are provided in SEQ ID NO: 467 and can be encoded by the polynucleotide provided in SEQ ID NO: 471. Non-limiting examples of light chain framework 3 (L-FR3) are provided in SEQ ID NO: 499 and can be encoded by the polynucleotide provided in SEQ ID NO: 503. Non-limiting examples of light chain framework 3 (L-FR3) are provided in SEQ ID NO: 531 and can be encoded by the polynucleotide provided in SEQ ID NO: 535. A non-limiting example of a light chain framework 3 (L-FR3) is provided in SEQ ID NO: 563 and can be encoded by the polynucleotide provided in SEQ ID NO: 567. A non-limiting example of a light chain framework 4 (L-FR4) is provided in SEQ ID NO: 20 and can be encoded by the polynucleotide provided in SEQ ID NO: 24. A non-limiting example of a light chain framework 4 (L-FR4) is provided in SEQ ID NO: 308 and can be encoded by the polynucleotide provided in SEQ ID NO: 312. A non-limiting example of a light chain framework 4 (L-FR4) is provided in SEQ ID NO: 340 and can be encoded by the polynucleotide provided in SEQ ID NO: 344. A non-limiting example of a light chain framework 4 (L-FR4) is provided in SEQ ID NO: 372 and can be encoded by the polynucleotide provided in SEQ ID NO: 376.Non-limiting examples of light chain framework 4 (L-FR4) are provided in SEQ ID NO: 404 and can be encoded by the polynucleotide provided in SEQ ID NO: 408. Non-limiting examples of light chain framework 4 (L-FR4) are provided in SEQ ID NO: 436 and can be encoded by the polynucleotide provided in SEQ ID NO: 440. Non-limiting examples of light chain framework 4 (L-FR4) are provided in SEQ ID NO: 468 and can be encoded by the polynucleotide provided in SEQ ID NO: 472. Non-limiting examples of light chain framework 4 (L-FR4) are provided in SEQ ID NO: 500 and can be encoded by the polynucleotide provided in SEQ ID NO: 504. Non-limiting examples of light chain framework 4 (L-FR4) are provided in SEQ ID NO: 532 and can be encoded by the polynucleotide provided in SEQ ID NO: 536. A non-limiting example of a light chain framework 4 (L-FR4) is provided in SEQ ID NO: 564 and can be encoded by the polynucleotide provided in SEQ ID NO: 568. It is contemplated that any of the frameworks disclosed herein can be substituted with an alternative framework, as will be appreciated by those skilled in the art.

[0212] In some embodiments, the ASGR1 binding polypeptides disclosed herein bind to ASGR1. By virtue of binding to ASGR1, these ASGR1 binding polypeptides may also bind to the asialoglycoprotein receptor complex (ASGPR) composed of ASGR1 and ASGR2 subunits. In some embodiments, the ASGR1 binding polypeptides provided herein do not bind to ASGR2 (e.g., they are specific for ASGR1).

[0213] Tolerogenic compounds and compositions

[0214] Embodiments disclosed herein relate to compounds, compositions (e.g., pharmaceutical compositions), or constructs for immune tolerance. Based on the disclosure provided herein, immune tolerance can be induced against a variety of antigens. For example, antigens can be endogenous (e.g., self-antigens) or exogenous (e.g., foreign antigens or alloantigens), including but not limited to: alloantigens, to which a transplant recipient produces an undesirable immune response, such as graft-versus-host disease or transplant rejection; foreign food, animal, plant, or environmental antigens, to which a patient produces an undesirable immune response, such as an allergic reaction or a hypersensitivity reaction; therapeutic agents, to which a patient produces an undesirable immune response, such as a hypersensitivity reaction and / or a reduction in therapeutic activity; self-antigens, to which a patient produces an undesirable immune response, such as an autoimmune disease; or a tolerogenic portion of any such antigen, such as a fragment or epitope.

[0215] Disclosed herein are tolerogenic compounds. In some embodiments, the tolerogenic compound comprises an ASGR1 binding polypeptide conjugated or fused to an antigen to which tolerance is desired. In some embodiments, the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally the linker is a polypeptide linker or a chemical conjugation linker. In some embodiments, the ASGR1 binding polypeptide is chemically conjugated to the antigen. In some embodiments, the ASGR1 binding polypeptide is recombinantly fused to the antigen. In some embodiments, the ASGR1 binding polypeptide is any one of the ASGR1 binding polypeptides disclosed herein. In some embodiments, the ASGR1 binding polypeptide comprises a heavy chain variable region. In some embodiments, the heavy chain variable region comprises one or more of the heavy chain CDRs HCDR1, HCDR2, and HCDR3. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO: 9. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO: 297. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO: 329. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO: 361. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:393. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:425. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:457. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:489. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:521. In some embodiments, HCDR1 comprises a sequence of SEQ ID NO:553. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:10. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:298. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:330. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:362. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:394. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:426. In some embodiments, HCDR2 comprises a sequence of SEQ ID NO:458. In some embodiments, HCDR2 comprises the sequence of SEQ ID NO: 490. In some embodiments, HCDR2 comprises the sequence of SEQ ID NO: 522. In some embodiments, HCDR2 comprises the sequence of SEQ ID NO: 554. In some embodiments, HCDR3 comprises the sequence of SEQ ID NO: 11. In some embodiments, HCDR3 comprises the sequence of SEQ ID NO: 299.In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:331. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:363. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:395. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:427. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:459. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:491. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:523. In some embodiments, the HCDR3 comprises the sequence of SEQ ID NO:555. In some embodiments, HCDR1 comprises the sequence of SEQ ID NO:9, 297, 329, 361, 393, 425, 457, 489, 521, or 553, HCDR2 comprises the sequence of SEQ ID NO:10, 298, 330, 362, 394, 426, 458, 490, 522, or 554, and HCDR3 comprises the sequence of SEQ ID NO:11, 299, 331, 363, 395, 427, 459, 491, 523, or 555. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 15. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 303. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 335. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 367.In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 399. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 431. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 463. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 495. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 527. In some embodiments, the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 559. In some embodiments, the ASGR1 binding polypeptide comprises (or further comprises in combination with the heavy chain variable region) a light chain variable region. In some embodiments, the light chain variable region comprises one or more of the light chain CDRs LCDR1, LCDR2, and LCDR3. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 25. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 313. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 345. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO: 377.In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:409. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:441. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:473. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:505. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:537. In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:569. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:26. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:314. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:346. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:378. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:410. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:442. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:474. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:506. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:538. In some embodiments, LCDR2 comprises the sequence of SEQ ID NO:570. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:27. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:315. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:347. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:379. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:411. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:443. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:475. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:507. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:539. In some embodiments, LCDR3 comprises the sequence of SEQ ID NO:571.In some embodiments, LCDR1 comprises the sequence of SEQ ID NO:25, 313, 345, 377, 409, 441, 473, 505, 537 or 569, LCDR2 comprises the sequence of SEQ ID NO:26, 314, 346, 378, 410, 442, 474, 506, 538 or 570, and LCDR3 comprises the sequence of SEQ ID NO:27, 315, 347, 379, 411, 443, 475, 507, 539 or 571. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 31. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 319. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 351. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 383. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 415. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 447.In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 479. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 511. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 543. In some embodiments, the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 575. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 15, and the light chain variable region comprises the sequence of SEQ ID NO: 31. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 303, and the light chain variable region comprises the sequence of SEQ ID NO: 319. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 335, and the light chain variable region comprises the sequence of SEQ ID NO: 351. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 367, and the light chain variable region comprises the sequence of SEQ ID NO: 383. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 399, and the light chain variable region comprises the sequence of SEQ ID NO: 415. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 431, and the light chain variable region comprises the sequence of SEQ ID NO: 447. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 463, and the light chain variable region comprises the sequence of SEQ ID NO: 479. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO:495, and the light chain variable region comprises the sequence of SEQ ID NO:511.In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 527, and the light chain variable region comprises the sequence of SEQ ID NO: 543. In some embodiments, the heavy chain variable region comprises the sequence of SEQ ID NO: 559, and the light chain variable region comprises the sequence of SEQ ID NO: 575. In some embodiments, the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb), or a scFv. In some embodiments, the ASGR1 binding polypeptide comprises an Fc domain. In some embodiments, the Fc domain is silenced. In some embodiments, the Fc domain is partially silenced.

[0216] In some embodiments of the tolerogenic compounds disclosed herein, the ASGR1 binding polypeptide and the antigen are conjugated or fused. In some embodiments, the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker. In some embodiments, the ASGR1 binding polypeptide and the antigen are chemically conjugated, such as using a chemical conjugation linker. In some embodiments, the ASGR1 binding polypeptide and the antigen are recombinantly fused. In some embodiments, the ASGR1 binding polypeptide and the antigen are recombinantly fused using a linker. In some embodiments, the linker is a polypeptide linker (e.g., including but not limited to when the ASGR1 binding polypeptide and the antigen are both polypeptides). In some embodiments, the linker is a cleavable linker. In some embodiments, the linker comprises glycine and / or serine. The use of glycine and serine linkers is generally known in the art. In some embodiments, the linker comprises the sequence of SEQ ID NO: 37. In some embodiments, when the ASGR1 binding polypeptide is in the form of an antibody, the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide, for example, the N-terminus or C-terminus of the heavy chain, heavy chain variable region, light chain, or light chain variable region of the ASGR1 binding polypeptide. In some embodiments, the ASGR1 binding polypeptide comprises an Fc domain. In some embodiments, the Fc domain is silenced or partially silenced, for example, such that the Fc domain exhibits significantly reduced or no binding to FcRn or other FcRγ receptors. In some embodiments, silencing of the Fc domain eliminates recycling of antibodies exiting the cell after uptake. In some embodiments, the antigen is conjugated or fused to the Fc domain. In several embodiments, the antigen is conjugated or fused to the C-terminus of the Fc domain.

[0217] In some embodiments, the antigen of any of the tolerogenic compounds disclosed herein is an agent capable of inducing an undesirable immune response in a subject. In some embodiments, the antigen may be a protein, peptide, or polypeptide. In some embodiments, the antigen may be a complete or partial therapeutic agent, a full-length autoantigen or a portion thereof, a full-length alloantigen or a portion thereof, a full-length allergen or a portion thereof, or a mimetic of any of the foregoing antigens. Nucleic acids are also contemplated that may encode any of the foregoing antigens, or act as an antigen itself (e.g., acting as a nucleic acid autoantigen). Depending on the embodiment, a combination of multiple antigens or portions or fragments may also be used. For example, if a longer peptide identified as P has antigenic regions A, B, C, and D, the composition disclosed herein for inducing tolerance to P may include any one or more of A, B, C, and D and / or any one or more of A, B, C, and D. Any particular antigen is included in a category or associated with any particular disease or reaction, and does not exclude that antigen from being considered part of another category or associated with another disease or reaction.

[0218] In several embodiments, the antigens to which tolerance is desired include one or more foreign antigens, such as food, animal, plant, and environmental antigens, to which the patient may experience an undesirable immune response. Although therapeutic proteins may also be considered foreign antigens due to their exogenous origin, such therapeutic agents are described as a separate group for the purpose of clearly describing the present disclosure. Similarly, plant or animal antigens may be edible and considered food antigens, and environmental antigens may be derived from plants. However, they are considered foreign antigens. For simplicity, no attempt will be made to describe, distinguish, and define all such potentially overlapping groups, as those skilled in the art will understand the antigens that may be used in the compositions of the present disclosure, particularly in light of the detailed description and examples.

[0219] In several embodiments, the antigen comprises one or more therapeutic agents, which are proteins, peptides, antibodies and antibody-like molecules (including antibody fragments and fusion proteins with antibodies and antibody fragments) and gene therapy vectors. These include human proteins, non-human (such as mice) and non-natural (e.g., engineered) proteins, antibodies, chimeric antibodies, humanized antibodies, viruses and virus-like particles, and non-antibody binding scaffolds, such as fibronectin, DARPin, knottin, etc. In several embodiments, human allogeneic transplant antigens are used, and the transplant recipient produces an undesirable immune response to it. In several embodiments, the antigen comprises one or more autoantigens, which cause an undesirable autoimmune response. Although autoantigens are endogenous sources in patients with autoimmune diseases, according to several embodiments, the polypeptides used in the compositions of the present disclosure are synthesized exogenously (rather than purified and concentrated from the source), depending on the embodiment.

[0220] In several embodiments, the antigens to which tolerance is desired include one or more foreign antigens, such as food, animal, plant, and environmental antigens, to which the patient may experience an undesirable immune response. Although therapeutic proteins may also be considered foreign antigens due to their exogenous origin, such therapeutic agents are described as a separate group for the purpose of clearly describing the present disclosure. Similarly, plant or animal antigens may be edible and considered food antigens, and environmental antigens may be derived from plants. However, they are considered foreign antigens. For simplicity, no attempt will be made to describe, distinguish, and define all such potentially overlapping groups, as those skilled in the art will understand the antigens that may be used in the compositions of the present disclosure, particularly in light of the detailed description and examples.

[0221] In some embodiments, the antigen is selected from the group consisting of insulin, proinsulin, preproinsulin, gluten, gliadin, myelin basic protein, myelin oligodendrocyte glycoprotein and proteolipid protein, desmoglein 3, desmoglein 1, alpha-synuclein, acetylcholine receptor, factor VIII, factor IX, asparaginase, uricase, adeno-associated virus (AAV) and a fragment of any of the foregoing. In some embodiments, the antigen is not a full-length protein. For example, in some embodiments, the antigen is not a full-length gliadin, insulin or proinsulin. In some embodiments, the antigen is not a full-length myelin basic protein, is not a full-length myelin oligodendrocyte protein or is not a full-length proteolipid protein. In some embodiments, the antigen is not a fragment of a protein. As discussed in more detail below, there are a variety of antigens that may be expected to be tolerated. These may include, but are not limited to, exogenous antigens that cause adverse immune responses when subjects are exposed to antigens. In some embodiments, adverse immune responses may be the result of ingesting antigens (e.g., orally, nasally or via some other mucosal routes). For example, these approaches can be the case with food antigens. In some embodiments, antigens can be purposefully administered to a subject, for example, by administering a therapeutic composition to treat a disease or condition suffered by the subject. In other additional embodiments, antigens can be produced by a subject, for example, an autoimmune antigen. For example, in several embodiments, the antigen comprises an exogenous transplant antigen or a tolerogenic portion thereof, to which the transplant recipient produces an undesirable immune response. In several embodiments, the antigen comprises an exogenous food, animal, plant or environmental antigen or a tolerogenic portion thereof, to which the patient produces an undesirable immune response. In several embodiments, the antigen comprises an exogenous therapeutic agent or a tolerogenic portion thereof, to which the patient produces an undesirable immune response. In several embodiments, the antigen comprises a synthetic self-antigen or a tolerogenic portion thereof, to which the patient produces an undesirable immune response to an endogenous form of the synthetic self-antigen.

[0222] In further detailing the above, compounds are provided in several embodiments wherein the antigen is a food antigen. In some such embodiments, the antigen is one or more of the following: conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), alpha-lactalbumin (ALA), lactotransferrin, Pen a 1 allergen (Pen a 1), allergen Pen m2 (Pen m2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, alpha-gliadin, gamma-gliadin, omega-gliadin, hordein, secalin, and avenin. In several embodiments, fragments of any of these antigens and / or mimetic epitopes of any of these antigens are also used. In several embodiments, the antigen is selected from the group consisting of gluten, high molecular weight gluten, low molecular weight gluten, α-gliadin, γ-gliadin, Ω-gliadin, hordein, secalin and avenin and fragments thereof. In several embodiments, the antigen is selected from the group consisting of gluten, high molecular weight gluten, low molecular weight gluten, α-gliadin, γ-gliadin and Ω-gliadin and fragments thereof. In several embodiments, the antigen is gluten or a fragment thereof. In several embodiments, the antigen is gliadin or a fragment thereof.

[0223] In several embodiments, compounds are provided in which the antigen is a therapeutic agent. In several embodiments, the antigen is selected from the group consisting of: factor VII, factor VIII, factor IX, asparaginase, uricase, adeno-associated virus (AAV) and a fragment of any one thereof. In several embodiments, the antigen is a therapeutic agent selected from the group consisting of: factor VII and factor IX and fragments thereof. In several embodiments, the antigen is a therapeutic agent selected from the group consisting of: factor VIII or a fragment thereof. In several embodiments, when the antigen is a therapeutic agent, the compound can be used to treat, prevent, reduce or otherwise improve the immune response produced against a therapeutic agent for hemophilia. As discussed herein, mimicking epitopes of any antigenic portion of the above-mentioned antigens can be used in several embodiments.

[0224] In several embodiments, the antigen comprises asparaginase or a fragment thereof. In several embodiments, the antigen comprises uricase or a fragment thereof. In several such embodiments, the compounds can be used to treat, prevent, reduce or otherwise improve the immune response generated against an anti-tumor agent. As discussed herein, mimotopes of any antigenic portion of the above antigens can be used in several embodiments.

[0225] In several embodiments, the antigen is associated with an autoimmune disease. For example, in several embodiments, the associated autoimmune disease is one or more of: type 1 diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, celiac disease with diarrhea, myasthenia gravis, and neuromyelitis optica.

[0226] In several embodiments, the autoimmune disease is type 1 diabetes and the antigen comprises insulin or a fragment thereof. In several embodiments, the autoimmune disease is type 1 diabetes and the antigen comprises proinsulin or a fragment thereof. In several embodiments, the autoimmune disease is type 1 diabetes and the antigen comprises preproinsulin or a fragment thereof. As discussed herein, mimotopes of any antigenic portion of the above antigens can be used in several embodiments. In several embodiments, combinations of these antigens can be incorporated into tolerogenic compounds, which can help reduce immune responses to autoantigens at multiple points along the insulin pathway.

[0227] In several embodiments, the autoimmune disease is multiple sclerosis, and the antigen comprises myelin basic protein or a fragment thereof. In several embodiments, the autoimmune disease is multiple sclerosis, and the antigen comprises myelin oligodendrocyte glycoprotein or a fragment thereof. In several embodiments, the autoimmune disease is multiple sclerosis, and the antigen comprises proteolipid protein or a fragment thereof. As discussed herein, mimotopes of any antigenic portion of the above antigens can be used in several embodiments. In several embodiments, combinations of these antigens can be incorporated into tolerogenic compounds (e.g., mixtures of antigens or fragments of MOG, MBP and / or PLP), which can help reduce immune responses to autoantigens at multiple points along the pathway that controls myelination or myelin repair.

[0228] As discussed herein, mimotopes of any antigenic portion of the above-described autoantigens may be used in several embodiments.

[0229] In several embodiments, a pharmaceutically acceptable composition consists of or consists essentially of a compound wherein the antigen is a food antigen, a therapeutic agent, an autoantigen or fragment thereof, an optional linker, and a liver targeting moiety as disclosed herein.

[0230] The antigen may be a complete protein, a portion of a complete protein, a peptide, etc., and may be derivatized for, e.g., conjugation with a linker. In some embodiments, the antigen may be a variant and / or may contain conservative substitutions, particularly maintaining sequence identity, relative to the antigen found in nature, and / or may be desialylated (or otherwise modified).

[0231] In some embodiments, the antigen comprises a food antigen. In some embodiments, the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), plant cystatin, thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin, and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitory protein (Sin a 4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens and a mimetic of any one of the antigens.

[0232] In some embodiments, the food antigen is selected from the group consisting of gluten, high molecular weight gluten, low molecular weight gluten, α-gliadin, γ-gliadin, Ω-gliadin, hordein, secalin, avenin, a portion of any of the antigens, and a mimetic of any of the antigens. In several embodiments, the food antigen is selected from gluten, high molecular weight gluten, low molecular weight gluten, α-gliadin, γ-gliadin, and Ω-gliadin, a portion of any of the antigens, and a mimetic of any of the antigens. In some embodiments, the food antigen is gluten or a portion or mimetic thereof. In general, these antigens may be associated with gluten intolerance, gluten-sensitive enteropathy, and / or diarrheal celiac disease. In some embodiments, the antigen is associated with the HLA-DQ2 serotype group.

[0233] In diarrheal celiac disease, major antigens include, but are not limited to, tissue transglutaminase and native and deamidated forms of gluten or gluten-like proteins, such as α-, γ- and Ω-gliadins, glutenins, hordeins, secalins and avenins. Those skilled in the art will appreciate that, for example, antigens associated with gluten intolerance may be converted to be more immunogenic in vivo by deamidation, which is achieved by converting the glutamine of the antigen to glutamate by tissue glutaminase. Thus, while antigens associated with gluten intolerance may initially be considered foreign food antigens, they may also be considered self-antigens after being modified by the body.

[0234] In some embodiments, the sequences of wheat gluten and other proteins associated with gluten intolerance and / or diarrheal celiac disease are generally known in the art, for example from Bromilow et al. "A curated gluten protein sequence database to support development of proteomics methods for determination of gluten in gluten-free foods" J. Proteomics (2017) 163: 67-75, which is hereby expressly incorporated by reference in its entirety.

[0235] In some embodiments, peptides or epitopes useful in the tolerogenic compounds disclosed herein for preventing undesirable immune responses against proteins associated with gluten intolerance and / or diarrheal celiac disease include some or all of the following sequences, either alone or in combination:

[0236] HLA-DQ-2.5 associated, native α-gliadin "33-mer":

[0237]

[0238] HLA-DQ-2.5-associated, deamidated α-gliadin "33-mer":

[0239]

[0240] HLA-DQ-8 related, α-gliadin:

[0241]

[0242] HLA-DQ-8 associated, Ω-gliadin (wheat, U5UA46):

[0243]

[0244] α-gliadin "15-mer" fragment:

[0245]

[0246] Gliadin with linker:

[0247]

[0248] Gliadin with linker and cysteine ​​conjugated moieties:

[0249]

[0250] Extended Gliadin:

[0251]

[0252] Extended gliadin with cysteine ​​conjugated moieties:

[0253]

[0254] Extended deamidated gliadin:

[0255]

[0256] Extended deamidated gliadin:

[0257]

[0258] Dq-8, α-gliadin:

[0259]

[0260] Dq-8, α-gliadin with a cysteine ​​conjugated moiety:

[0261]

[0262] Dq-8, extended α-gliadin:

[0263]

[0264] Dq-8, extended α-gliadin with cysteine ​​conjugated moiety:

[0265]

[0266] For any of the peptide or epitope sequences provided herein for use as tolerogens (e.g., SEQ ID NOs: 40-54 or fragments thereof), embodiments may also include sequences having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity, which are contemplated to have the same or similar efficacy. Some of the sequences provided herein may include a cysteine ​​conjugated moiety for chemical conjugation. Alternative moieties for chemical conjugation are also contemplated. In alternative embodiments, the sequences provided herein may be fused to antibody sequences without additional conjugated moieties, e.g., by recombinant cloning techniques.

[0267] In some embodiments, the antigen is a foreign antigen associated with an animal, plant, or environmental allergen, toxin, or irritant. Some non-limiting examples include antigens, allergens, toxins, or irritants from cats, mice, dogs, horses, bees, dust, fungi, trees, and plants, including but not limited to:

[0268] Weeds (including ragweed allergens Amb a 1, 2, 3, 5, and 6 and Amb t 5; Chenopodium album Che a 2 and 5; and other weed allergens Par j 1, 2, 3, and Par o 1);

[0269] Grasses (including major allergens Cyn d 1, 7 and 12; Dac g 1, 2 and 5; Hol I 1.01203; Lol p 1, 2, 3, 5 and 11; Mer a 1; Pha a 1; Poa p 1 and 5);

[0270] Pollen from ragweed and other weeds (including curly dock, lambs quarters, pigweed, plantain, sorrel, and wormwood), grasses (including Bermuda grass, Johnson grass, Kentucky grass, orchard grass, sweetgrass, and timothy grass), and trees (including catalpa, elm, hickory, olive, pecan, sycamore, and walnut trees);

[0271] Dust (including major allergens from species of house dust mites such as Der p 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 14, 15, 18, 20, 21 and 23; major allergens from species of dust mites such as Der f 1, 2, 3, 6, 7, 10, 11, 13, 14, 15, 16, 18, 22 and 24; major allergens from species of tropical anaplasma mites such as Blo t 1, 2, 3, 4, 5, 6, 10, 11, 12, 13, 19 and 21; in addition allergens Eur m 2 from the mycophagous mite, allergen Tyrp 13 from the putrescentia putrescentia, and allergens Bla g 1, 2 and 4; Per a 1, 3 and 7 from cockroaches);

[0272] Pets (including cats, dogs, rodents and farm animals; major cat allergens include Fel d 1 to 8, cat IgA, Bla g 2 and cat albumin; major dog allergens include Can f 1 to 6 and dog albumin);

[0273] Bee stings, which include major allergens Api m 1 to 12; and

[0274] Fungi, including allergens derived from Aspergillus species and Penicillium species, as well as Alternaria species, Davidiella tassiana species, and Trichophyton rubrum species.

[0275] In some embodiments, the antigen comprises an autoantigen. In some embodiments, the autoantigen is selected from thyroglobulin, thyroid peroxidase, thyroid stimulating hormone receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, myelin oligodendrocyte glycoprotein, proteolipid protein, desmoglein, desmoglein-1, desmoglein-3, desmoglein-4, alpha-synuclein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38, chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and fragments or portions of any of the antigens and mimetics of any of the antigens. In some embodiments, the antigen comprises an antigen associated with an autoimmune disease. In some embodiments, the autoimmune disease is selected from the group consisting of type 1 diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, celiac disease with diarrhea, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and antineutrophil cytoplasmic antibody-associated vasculitis.

[0276] In several embodiments, the antigen to be tolerated is a viral antigen, for example, a viral antigen derived from a therapeutic viral vector, such as an adeno-associated viral vector (AAV). In several embodiments, the antigen to be tolerated comprises or is derived from an immunogenic fragment of AAV serotype 2 capsid protein 1 (SEQ ID NO: 141). In several embodiments, the antigen to be tolerated comprises or is derived from an immunogenic fragment of AAV serotype 2 capsid protein 2 (SEQ ID NO: 142). In several embodiments, the antigen to be tolerated comprises or is derived from an immunogenic fragment of AAV serotype 2 capsid protein 3 (SEQ ID NO: 143). In several embodiments, the antigen to be tolerated comprises or is derived from an immunogenic fragment of AAV serotype 9 capsid protein 1 (SEQ ID NO: 144). In several embodiments, the antigen to be tolerated comprises or is derived from an immunogenic fragment of AAV serotype 9 capsid protein 2 (SEQ ID NO: 145). In several embodiments, the antigen to be tolerated comprises or is derived from an immunogenic fragment of AAV serotype 9 capsid protein 3 (SEQ ID NO: 146).

[0277] An exogenously obtained form of MART1 (Melanoma antigen recognized by T cells 1, Melan-A) that may be used in the compositions of the present disclosure has the following sequence (Uniprot #Q16655):

[0278]

[0279] Tyrosinase, including exogenously obtained forms useful in the compositions of the present disclosure, has the following sequence (Uniprot #P14679):

[0280]

[0281]

[0282] Melanocyte protein (gp100), including exogenously obtained forms useful in the compositions of the present disclosure, has the following sequence (Uniprot #P40967):

[0283]

[0284] An exogenously obtained form of aquaporin 4, including one useful in the compositions of the present disclosure, has the following sequence (Uniprot #P55087):

[0285]

[0286]

[0287] In uveitis, the major antigens include retinal S antigen or "S inhibitory protein" and interphotoreceptor retinoid binding protein (IRBP) or retinol binding protein 3.

[0288] The S inhibitory protein, including exogenously obtained forms useful in the compositions of the present disclosure, has the following sequence (Uniprot #P10523):

[0289]

[0290] An exogenously obtained form of IRBP, including one useful in the compositions of the present disclosure, has the following sequence (Uniprot #P10745):

[0291]

[0292]

[0293] In some embodiments, the autoimmune disease is type 1 diabetes. In some embodiments, the antigen comprises insulin, proinsulin, preproinsulin, glutamic acid decarboxylase 65 (GAD-65), GAD-67, glucose-6-phosphatase 2, islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP), insulinoma-associated protein 2 (IA-02), insulinoma-associated protein 2β (IA-2β), ICA69, ICA12 (SOX-13), carboxypeptidase H, Imogen 38, GLIMA 38, chromogranin A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-intestinal-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy kinase, and SST G protein-coupled receptor 1-5, or a fragment, portion, or mimetic thereof. In some embodiments, combinations of these antigens can be incorporated into tolerogenic compounds, which can result in a synergistic effect in reducing immune responses to autoantigens at multiple points along the insulin pathway. It should be noted that some embodiments of the present disclosure include recombinant forms of insulin and derivatives thereof for the treatment of diabetes. It should be noted that insulin is an example of an antigen that can be characterized as an autoantigen and a therapeutic protein antigen. For example, rHu insulin and bovine insulin are therapeutic protein antigens (are the object of undesirable immune attack), while endogenous human insulin is an autoantigen (is the object of undesirable immune attack). Because endogenous human insulin is not suitable for use in pharmaceutical compositions, a recombinant form is used in certain embodiments of the compositions of the present disclosure.

[0294] Human insulin, including exogenously obtained forms useful in the compositions of the present disclosure, has the following sequence (Uniprot #P01308):

[0295]

[0296] An exogenously obtained form of GAD-65, including one useful in the compositions of the present disclosure, has the following sequence (Uniprot #Q05329):

[0297]

[0298] An exogenously obtained form of IGRP, including one useful in the compositions of the present disclosure, has the following sequence (Uniprot #QN9QR9):

[0299]

[0300] In several embodiments, exogenously obtained forms of human proinsulin, including those useful in the tolerogenic compositions of the present disclosure, have the following sequence:

[0301]

[0302] In some embodiments, peptides or epitopes useful in the tolerogenic compounds disclosed herein for preventing an undesirable immune response to proteins involved in the insulin pathway and / or for treating type 1 diabetes include some or all of the following sequences (alone or in combination):

[0303] Human Proinsulin 1-70:

[0304]

[0305] Human Proinsulin 9-70:

[0306]

[0307] Human proinsulin 9-38:

[0308]

[0309] Human proinsulin 1-38:

[0310]

[0311] Human proinsulin 9-23:

[0312]

[0313] Human proinsulin 45-71 (C13-A6):

[0314]

[0315] Human proinsulin C24-A1:

[0316]

[0317] Human Proinsulin C19-A3:

[0318]

[0319] Human Proinsulin C13-32:

[0320]

[0321] Human Proinsulin B9-C4:

[0322]

[0323] Human proinsulin C22-A5:

[0324]

[0325] Human IA-2 654-674:

[0326]

[0327] Human IA-2 718-782:

[0328]

[0329] Human IA-2 785-819:

[0330]

[0331] Human IA-2 828-883:

[0332]

[0333] Human IA-2 943-979:

[0334]

[0335] For any of the peptide or epitope sequences provided herein for use as tolerogens (e.g., SEQ ID NOs: 55-70 or fragments thereof), embodiments may also include sequences having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity, which are contemplated to have the same or similar efficacy. It is contemplated that the sequence may also include a cysteine ​​conjugated moiety or any other alternative moiety for chemical conjugation. In alternative embodiments, the sequences provided herein may be fused to an antibody sequence without an additional conjugated moiety, e.g., by recombinant cloning techniques.

[0336] In some embodiments, the autoimmune disease is multiple sclerosis. In some embodiments, the antigen comprises myelin basic protein, myelin oligodendrocyte glycoprotein, proteolipid protein, or any fragment, portion, or mimetic thereof. In some embodiments, combinations of these antigens can be incorporated into tolerogenic compounds, which can result in a synergistic effect in reducing the immune response to autoantigens at multiple points along the pathological pathway to promote resolution of the disease.

[0337] In some embodiments, peptides or epitopes useful for the tolerogenic compounds disclosed herein are involved in myelination, myelin repair and / or for the treatment of multiple sclerosis, and the tolerogenic compounds disclosed herein are used to prevent undesirable immune responses against proteins. In multiple sclerosis, major antigens include, but are not limited to, myelin basic protein ("MBP"), myelin oligodendrocyte glycoprotein ("MOG"), and myelin proteolipid protein ("PLP").

[0338] An exogenously obtained form of MBP, including one useful in the compositions of the present disclosure, has the following sequence (Uniprot #P02686):

[0339]

[0340] MOG, including exogenously obtained forms useful in the compositions of the present disclosure, has the following sequence (Uniprot #Q16653):

[0341]

[0342] PLP, including exogenously obtained forms useful in the compositions of the present disclosure, has the following sequence (Uniprot #P60201):

[0343]

[0344] In several embodiments, peptides or epitopes useful in the tolerogenic compounds disclosed herein include some or all of the following sequences derived from myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), or myelin proteolipid protein (PLP), either alone or in combination:

[0345] MBP 1-39:

[0346]

[0347] MBP 13-32: KYLATASTMDHARHGFLPRH (SEQ ID NO:72);

[0348] MBP 76-106:

[0349]

[0350] MBP 83-99 (altered peptide ligand): ENPWHFFKNIVTPRTP (SEQ ID NO: 74); MBP 102-136:

[0351]

[0352] MBP 111-129: LSRFSWGAEGQRPGFGYGG (SEQ ID NO:76);

[0353] MBP 146-170: AQGTLSKIFKLGGRDSRSGSPMARR (SEQ ID NO:77);

[0354] MBP 146-170 with cysteine ​​conjugated moiety:

[0355]

[0356] MBP 83-99: ENPVVHFFKNIVTPRTP (SEQ ID NO:79);

[0357] MBP 82-98: DENPVVHFFKNIVTPRT (SEQ ID NO: 80);

[0358] MBP 82-99: DENPVVHFFKNIVTPRTP (SEQ ID NO:81);

[0359] MBP 82-106: DENPVVHFFKNIVTPRTPPSQGKG (SEQ ID NO: 82);

[0360] MBP 87-106: VHFFKNIVTPRTPPSQGKG (SEQ ID NO:83);

[0361] MBP 131-155: ASDYKSAHKGLKGVDAQGTLSKIFK (SEQ ID NO: 84); MBP 131-170 with a cysteine ​​conjugated moiety:

[0362]

[0363] MBP 76-136:

[0364]

[0365] MOG 1-20: GQFRVIGPRHPIRALVGDEV (SEQ ID NO:87);

[0366] MOG 1-27: GQFRVIGPRHPIRALVGDEVELPCRIS (SEQ ID NO:88);

[0367] Alternative MOG 35-55: MEVGWYRPPFSRWHLYRNGK (SEQ ID NO:89);

[0368] MOG 30-60: KNATGMEVGWYRSPFSRVVHLYRNGKDQDAE (SEQ ID NO:90);

[0369] MOG 34-56: GMEVGWYRSPFSRVVHLYRNGKD (SEQ ID NO:91);

[0370] MOG 35-55: MEVGWYRPPFSRVVHLYRNGK (SEQ ID NO:92);

[0371] MOG 35-55 (mouse): MEVGWYRSPFSRVVHLYRNGK (SEQ ID NO:93);

[0372] MOG 33-62:

[0373]

[0374] MOG 11-30: PIRALVGDEVELPCRISPGK (SEQ ID NO:95);

[0375] MOG 18-62:

[0376]

[0377] MOG 21-40: ELPCRISPGKNATGMEVGWY (SEQ ID NO:97);

[0378] MOG 64-86: EYRGRTELLKDAIGEGKVTLRIR (SEQ ID NO:98);

[0379] MOG 1-60:

[0380]

[0381] MOG 1-62:

[0382]

[0383] PLP 41-58:GTEKLIETYFSKNYQDYE(SEQ ID NO:101);

[0384] PLP 89-106:GFYTTGAVRQIFGDYKTT(SEQ ID NO:102);

[0385] PLP 95-116:AVRQIFGDYKTTICGKGLSATV(SEQ ID NO:103);

[0386] PLP 178-197:NTWTTCQSIAFPSKTSASIG(SEQ ID NO:104);

[0387] PLP 190-209:SKTSASIGSLCADARMYGVL (SEQ ID NO:105); and

[0388] PLP 139-154:HCLGKWLGHPDKFVGI (SEQ ID NO: 106).

[0389] For any of the peptides or epitope sequences provided herein for use as tolerogens (e.g., SEQ ID NOs: 75-106), embodiments may also include sequences having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity, which are contemplated to have the same or similar efficacy. Some of the sequences provided herein may include a cysteine ​​conjugated moiety for chemical conjugation. Alternative moieties for chemical conjugation are also contemplated. In alternative embodiments, the sequences provided herein may be fused to antibody sequences without additional conjugated moieties, e.g., by recombinant cloning techniques.

[0390] In some embodiments, the antigen is a therapeutic agent. In some embodiments, the antigen is selected from the group consisting of Factor VIII, Factor IX, asparaginase, uricase, adeno-associated virus (AAV) (i.e., for gene therapy), and mimetics, fragments, or portions of any of the above antigens. In some embodiments, the antigen is associated with hemophilia.

[0391] In embodiments where the antigen is a therapeutic protein, peptide, antibody or antibody-like molecule, the specific antigen may be selected from the group consisting of: abatacept, abciximab, adalimumab, adenosine deaminase, Ado trastuzumab drug conjugate, α-galactosidase, β-galactosidase, aldeslukin, arabinoside, α-albinosidase, α-1-proteinase inhibitor, anakinra, anistreplase (anisoylated plasminogen streptokinase activator complex), antithrombin III, antithymocyte globulin, alteplase, bevacizumab, Bivalirudin, botulinum toxin type A, botulinum toxin type B, C1-esterase inhibitors, canakinumab, carboxypeptidase G2 (Glucarpidase and Voraxaze), certolizumab pegol, cetuximab, collagenase, Crotalidae immune Fab, darbepoetin-alpha, denosumab, digoxin immune Fab, danase alfa, eculizumab, etanercept, factor VIIa, factor VIII, factor IX, factor XI, factor XIII, fibrinogen, filgrastim, galactosidase, golimumab, histrelin acetate, Hyaluronidase, idursulfase, imiglucerase, infliximab, insulin [including recombinant human insulin ("rHu insulin") and bovine insulin], interferon-α2a, interferon-α2b, interferon-β1a, interferon-β1b, interferon-γ1b, ipilimumab, L-arginase, L-asparaginase, L-methioninase, lactase, laronidase, lepirudin / hirudin, mecasermin, mecasermin-linfibeb, methoxynatalizumab, octreotide, ofatumumab, oprelleukin, pancreatic amylase, pancreatic lipase, papain, Pegylated asparaginase, pegylated doxorubicin HCl, pegylated epoetin-β, pegylated filgrastim, pegylated interferon-α2a, pegylated interferon-α2b, pegololac, pegvisomant, phenylalanine ammonia lyase (PAL), protein C, rasburicase (uricase), sucrasidase (Sacrosidase), salmon calcitonin, sargramostim, streptokinase, tenecteplase, teriparatide, tocilizumab (atlizumab), trastuzumab, type 1 alpha-interferon, ustekinumab, vW factor. Therapeutic proteins can be obtained from natural sources (e.g., concentrated and purified) or synthetically (e.g., recombinant), and include antibody therapeutics that are typically IgG monoclonal or fragments or fusions.

[0392] Particular therapeutic proteins, peptides, antibodies or antibody-like molecules include, but are not limited to, abciximab, adalimumab, α-galactosidase, β-galactosidase, aldesleukin, α-alglucosidase, factor VIII, factor IX, infliximab, insulin (including rHu insulin), L-asparaginase, laronidase, natalizumab, octreotide, phenylalanine ammonia lyase (PAL) or rasburicase (uricase) and generally IgG monoclonal antibodies in their various forms.

[0393] Some embodiments employ hemostatic agents (eg, Factors VIII and IX), insulin (including rHu insulin), and the therapeutic molecules uricase, phenylalanine ammonia lyase (PAL), and asparaginase (which may be of non-human origin).

[0394] In several embodiments, the therapeutic agent is delivered by using, for example, a gene therapy vector. In some such embodiments, an immune response can be generated against a portion of such a vector and / or its cargo (e.g., a therapeutic agent). Thus, in several embodiments, the antigen to which tolerance is desired comprises a gene therapy vector, which includes, but is not limited to: adenovirus and adeno-associated virus (and corresponding variants-1, -2, -5, -6, -8, -9 and / or other parvoviruses), lentivirus, and retrovirus.

[0395] In some embodiments, for autoimmune diseases of the thyroid gland (including Hashimoto's thyroiditis and Graves' disease), exemplary antigens for tolerogenic compounds include, but are not limited to, thyroglobulin (TG), thyroid peroxidase (TPO), thyrotropin receptor (TSHR), sodium-iodine symporter (NIS), or megalin. In some embodiments, for thyroid-related eye and skin diseases, insulin-like growth factor 1 receptor is another exemplary antigen. In some embodiments, for hypoparathyroidism, calcium-sensitive receptor is another exemplary antigen.

[0396] In some embodiments, for Addison's disease, exemplary antigens for tolerogenic compounds include, but are not limited to, 21OH hydroxylase, 17OH hydroxylase, cytochrome P450 side chain cleavage enzyme (P450scc) and / or P450c21 or P450c17, or adrenocorticotropic hormone receptor (ACTH receptor).

[0397] In some embodiments, for premature ovarian failure, exemplary antigens for tolerogenic compounds include, but are not limited to, FSH receptor and alpha-enolase.

[0398] In some embodiments, for autoimmune hypophysitis or pituitary autoimmune diseases, exemplary antigens for tolerogenic compounds include, but are not limited to, pituitary specific protein factor (PGSF) 1a and 2, or type 2 iodothyronine deiodinase.

[0399] In some embodiments, for rheumatoid arthritis, exemplary antigens for tolerogenic compounds include, but are not limited to, collagen I, collagen II, immunoglobulin binding protein, double-stranded DNA, fibrin, fibrinogen (including fibrinogen α, β or γ chains), vimentin, aggrecan, or α-enolase.

[0400] An exemplary sequence for collagen II is provided as Uniprot #P02458 (SEQ ID NO: 107). In some embodiments, the tolerogenic compound may comprise amino acids 1236-1249 (SEQ ID NO: 108), 662-678 (SEQ ID NO: 160), 459-478 (SEQ ID NO: 161), 461-473 (SEQ ID NO: 162), 471-485 (SEQ ID NO: 163), 450-470 (SEQ ID NO: 164), 595-603 (SEQ ID NO: 165), 455-474 (SEQ ID NO: 166), 498-511 (SEQ ID NO: 167), 1133-1150 (SEQ ID NO: 168), 1427-1435 (SEQ ID NO: 169), 378-440 (SEQ ID NO: 170), 1350-1362 (SEQ ID NO: 171), 1237-1249 (SEQ ID NO: 172), In some embodiments, the tolerogenic compound is post-translationally modified, for example, amino acids 461-473 of collagen II are carbamylated at K4 (SEQ ID NO: 272), amino acids 455-474 of collagen II are hydroxylated at P4 and P19 (SEQ ID NO: 273), amino acids 1237-1249 of collagen II are citrullinated at R4 (SEQ ID NO: 274), amino acids 511-525 of collagen II are citrullinated at R5 (SEQ ID NO: 275), or amino acids 756-764 of collagen II are hydroxylated at P6 (SEQ ID NO: 276).

[0401] An exemplary sequence of aggrecan II is provided as Uniprot #P16112 (SEQ ID NO: 109). In some embodiments, the tolerogenic compound may comprise amino acids 84-103 (SEQ ID NO: 110), 32-64 (SEQ ID NO: 199), 68-82 (SEQ ID NO: 200), 76-110 (SEQ ID NO: 201), 116-130 (SEQ ID NO: 202), 140-169 (SEQ ID NO: 203), 161-177 (SEQ ID NO: 204), 174-188 (SEQ ID NO: 205), 200-215 (SEQ ID NO: 206), 225-244 (SEQ ID NO: 207), 252-269 (SEQ ID NO: 208), 287-314 (SEQ ID NO: 209), 341-355 (SEQ ID NO: 210), 520-539 (SEQ ID NO: 211), 553-570 (SEQ ID NO: 214). ID NO:212), 568-583 (SEQ ID NO:213), 589-601 (SEQ ID NO:214), 620-636 (SEQ ID NO:215), 666-674 (SEQ ID NO:216), 714-725 (SEQ ID NO:217), 866-879 (SEQ ID NO:218), 988-998 (SEQ ID NO:219), 1056-1067 (SEQ ID NO:220), 1352-1366 (SEQ ID NO:221), 1645-1658 (SEQ ID NO:222), 1892-1904 (SEQ ID NO:223), 1785-1798 (SEQ ID NO:224), 1805-1819 (SEQ ID NO:225), 1885-1898 (SEQ ID NO:226), 1937-1950 (SEQ ID NO:227), 1961-1974 (SEQ ID NO:228), 2018-2031 (SEQ ID NO:229), 2104-2118 (SEQ ID NO:230), 2189-2201 (SEQ ID NO:231), 2282-2296 (SEQ ID NO:232), 2351-2368 (SEQ ID NO:233), 2507-2521 (SEQ ID NO:234), 2513-2528 (SEQ ID NO:235) or 2516-2530 (SEQ ID NO:236).In some embodiments, the tolerogenic compound is post-translationally modified, e.g., amino acids 161-177 of aggrecan are citrullinated at R3, R10, and R13 (SEQ ID NO: 282), amino acids 200-215 of aggrecan are citrullinated at R11 (SEQ ID NO: 283), amino acids 225-244 of aggrecan are citrullinated at R7 and R12 (SEQ ID NO: 284), amino acids 520-539 of aggrecan are citrullinated at R11 and R16 (SEQ ID NO: 285), amino acids 553-570 of aggrecan are citrullinated at R4 and R9 (SEQ ID NO: 286), amino acids 568-583 of aggrecan are citrullinated at R8 (SEQ ID NO: 287). NO:287), or amino acids 620-636 of aggrecan are citrullinated at R13 (SEQ ID NO:288).

[0402] An exemplary sequence for a fibrinogen alpha chain is provided as Uniprot #P02671 (SEQ ID NO: 111). In some embodiments, the tolerogenic compound may comprise amino acids 78-91 (SEQ ID NO: 112), 717-725 (SEQ ID NO: 113), 51-95 (SEQ ID NO: 175), 138-152 (SEQ ID NO: 176), 171-185 (SEQ ID NO: 177), 201-215 (SEQ ID NO: 178), 300-314 (SEQ ID NO: 179), 347-361 (SEQ ID NO: 180), 363-377 (SEQ ID NO: 181), 420-434 (SEQ ID NO: 182), 438-452 (SEQ ID NO: 183), 456-470 (SEQ ID NO: 184), 542-556 (SEQ ID NO: 185), 717-725 (SEQ ID NO: 186), 718-730 (SEQ ID NO: 187), 720-731 (SEQ ID NO: 188), 732-740 (SEQ ID NO: 189), 744-751 (SEQ ID NO: 190), 755-761 (SEQ ID NO: 191), 765-772 (SEQ ID NO: 192), 770-781 (SEQ ID NO: 193), 781-790 (SEQ ID NO: 194), 800-810 (SEQ ID NO: 195), 811-82 In some embodiments, the tolerogenic compound is post-translationally modified, for example, amino acids 138-152 of the fibrinogen alpha chain are citrullinated at R6 (SEQ ID NO: 277), amino acids 456-470 of the fibrinogen alpha chain are citrullinated at R3 and R4 (SEQ ID NO: 278), amino acids 717-725 of the fibrinogen alpha chain are citrullinated at R4 (SEQ ID NO: 279), or amino acids 737-751 of the fibrinogen alpha chain are citrullinated at R7 (SEQ ID NO: 280).

[0403] An exemplary sequence of a fibrinogen beta chain is provided as Uniprot# P02675 (SEQ ID NO: 188).In some embodiments, the tolerogenic compound may comprise amino acids 433-411 (SEQ ID NO: 189) or 69-81 (SEQ ID NO: 190) of a fibrinogen beta chain.

[0404] An exemplary sequence of vimentin is provided as Uniprot#P08670 (SEQ ID NO: 114). In some embodiments, the tolerogenic compound may comprise amino acids 66-78 (SEQ ID NO: 115), 447-455 (SEQ ID NO: 116), 26-49 (SEQ ID NO: 191), 51-88 (SEQ ID NO: 192), 116-122 (SEQ ID NO: 193), 130-138 (SEQ ID NO: 194), 226-242 (SEQ ID NO: 195), 371-387 (SEQ ID NO: 196), 415-443 (SEQ ID NO: 197), or 447-455 (SEQ ID NO: 198) of vimentin. In some embodiments, the tolerogenic compound is post-translationally modified, for example, amino acids 447-455 of vimentin are citrullinated at R4 (SEQ ID NO: 281).

[0405] An exemplary sequence of alpha-enolase is provided as Uniprot# P06733 (SEQ ID NO: 237).In some embodiments, the tolerogenic compound may comprise amino acids 4-40 (SEQ ID NO: 238) or 326-340 (SEQ ID NO: 239) of alpha-enolase.

[0406] In some embodiments, for autoimmune gastritis, exemplary antigens for tolerogenic compounds include, but are not limited to, H+ / K+ ATPase.

[0407] In some embodiments, for pernicious anemia, exemplary antigens for tolerogenic compounds include, but are not limited to, intrinsic factor.

[0408] In some embodiments, for vitiligo, exemplary antigens for tolerogenic compounds include, but are not limited to, tyrosinase or tyrosinase-related proteins 1 and 2.

[0409] In some embodiments, for myasthenia gravis, exemplary antigens for tolerogenic compounds include, but are not limited to, nicotinic acetylcholine receptor (AChR), muscle specific kinase (MuSK), or lipoprotein receptor-related protein 4 (LRP4).

[0410] In some embodiments, for pemphigus vulgaris and variants, exemplary antigens for tolerogenic compounds include, but are not limited to, desmoglein-1, desmoglein-3, desmoglein-4, desmocollin 1, plectin, placoglobulin, periplakin, desmoplakin I and II, envoplakin, and acetylcholine receptors.

[0411] In some embodiments, for bullous pemphigoid, exemplary antigens for tolerogenic compounds include, but are not limited to, BP180, BP230, plectin, or laminin-5.

[0412] In some embodiments, for dermatitis herpetiformis (Duhring's disease), exemplary antigens for tolerogenic compounds include, but are not limited to, endomysial or tissue transglutaminase.

[0413] In some embodiments, for epidermolysis bullosa acquisita, exemplary antigens for tolerogenic compounds include, but are not limited to, collagen VII.

[0414] In some embodiments, for systemic sclerosis, exemplary antigens for tolerogenic compounds include, but are not limited to, matrix metalloproteinase 1, matrix metalloproteinase 3, heat shock protein 47, fibrillin 1, PDGF receptor, Scl-70, U1 snRNP, Th / To, Ku, Jo1, NAG-2, centromere proteins, topoisomerase I, nucleolar proteins, RNA polymerase I, II or III, PM-Scl, fibrillin, or B23.

[0415] In some embodiments, for mixed connective tissue disease, exemplary antigens for tolerogenic compounds include, but are not limited to, U1 snRNP.

[0416] In some embodiments, for Sjogren's syndrome, exemplary antigens for tolerogenic compounds include, but are not limited to, nuclear antigens SS-A (Ro) and SS-B (La), cytoplasmic proteins, poly (ADP-ribose) polymerase, topoisomerase, muscarinic receptors, and Fc-γ receptor IIIb. An exemplary sequence for SS-A is provided as Uniprot # P19474 (SEQ ID NO: 117).

[0417] An exemplary sequence of SS-B is provided as Uniprot# P05455 (SEQ ID NO: 118). In some embodiments, the tolerogenic compound may comprise amino acids 19-33 (SEQ ID NO: 240), 46-72 (SEQ ID NO: 241), 151-180 (SEQ ID NO: 242), 211-228 (SEQ ID NO: 243), 244-258 (SEQ ID NO: 244), or 304-329 (SEQ ID NO: 245) of SS-B.

[0418] In some embodiments, for systemic lupus erythematosus, exemplary antigens for tolerogenic compounds include, but are not limited to, "Smith antigen," SS-A, high mobility group box protein 1 (HMGB1), nucleosomes, histones, and double-stranded DNA.

[0419] In some embodiments, for Goodpasture syndrome, exemplary antigens for tolerogenic compounds include, but are not limited to, glomerular basement membrane protein or collagen IV.

[0420] In some embodiments, for rheumatic heart disease, autoimmune myocarditis, viral myocarditis, and / or inflammatory cardiomyopathy, exemplary antigens for tolerogenic compounds include, but are not limited to, cardiac myosin and myosin 6. An exemplary sequence for cardiac myosin is provided as Uniprot #P13533 (SEQ ID NO: 119).

[0421] An exemplary sequence of myosin 6 is provided as Uniprot #P12883 (SEQ ID NO: 251). In some embodiments, the tolerogenic compound can include amino acids 1906-1923 (SEQ ID NO: 252), 1828-1845 (SEQ ID NO: 253), 1411-1493 (SEQ ID NO: 254), 1554-1584 (SEQ ID NO: 255), 1170-1181 (SEQ ID NO: 256), 1619-1636 (SEQ ID NO: 257), 1671-1714 (SEQ ID NO: 258), 450-464 (SEQ ID NO: 259), 1801-1819 (SEQ ID NO: 260), 1333-1363 (SEQ ID NO: 261), 1867-1897 (SEQ ID NO: 262), 1749-1814 (SEQ ID NO: 263), 1142-1156 (SEQ ID NO: 264), NO:264), 1372-1389 (SEQ ID NO:265), 1710-1727 (SEQ ID NO:266), 1645-1662 (SEQ ID NO:267), 689-703 (SEQ ID NO:268), 1593-1610 (SEQ ID NO:269) or 111-119 (SEQ ID NO:264) NO:270).

[0422] In some embodiments, for autoimmune polyendocrine syndrome type I, exemplary antigens for tolerogenic compounds include, but are not limited to, aromatic L-amino acid decarboxylase, histidine decarboxylase, cysteine ​​sulfenic acid decarboxylase, tryptophan hydroxylase, tyrosine hydroxylase, phenylalanine hydroxylase, liver P450 cytochromes P4501A2 and P4502A6, SOX-9, SOX-10, calcium sensing receptor protein, or type 1 interferon.

[0423] In some embodiments, for neuromyelitis optica, exemplary antigens for tolerogenic compounds include, but are not limited to, aquaporin-4.

[0424] In some embodiments, for uveitis, exemplary antigens for tolerogenic compounds include, but are not limited to, retinal S-antigen, S-trap, interphotoreceptor retinoid binding protein (IRBP), or retinol binding protein 3.

[0425] In some embodiments, for Parkinson's disease, exemplary antigens for tolerogenic compounds include, but are not limited to, alpha-synuclein. An exemplary sequence of alpha-synuclein is provided as Uniprot# P37840 (SEQ ID NO: 120). In some embodiments, the tolerogenic compound may comprise amino acids 2-23 (SEQ ID NO: 246), 32-46 (SEQ ID NO: 247), 56-85 (SEQ ID NO: 248), 81-110 (SEQ ID NO: 249), or 116-140 (SEQ ID NO: 250) of alpha-synuclein.

[0426] In some embodiments, for anti-neutrophil cytoplasmic antibody-associated vasculitis (ANCA-V, ANCA-vasculitis), exemplary antigens for tolerogenic compounds include, but are not limited to, myeloperoxidase and proteinase 3 / myelogenin. An exemplary sequence for myeloperoxidase is provided as Uniprot#P05164 (SEQ ID NO: 121). In some embodiments, the tolerogenic compound may comprise amino acids 447-461 (SEQ ID NO: 122), 435-454 (SEQ ID NO: 123), 409-423 (SEQ ID NO: 124), 409-474 (SEQ ID NO: 125), 279-341 (SEQ ID NO: 126), 341-409 (SEQ ID NO: 127), or 598-745 (SEQ ID NO: 128) of myeloperoxidase. An exemplary sequence for proteinase 3 is provided as Uniprot#PP24158 (SEQ ID NO: 129).

[0427] In some embodiments, for primary biliary cholangitis, exemplary antigens for tolerogenic compounds include, but are not limited to, pyruvate dehydrogenase complex E2 subunit (PDC-E2). An exemplary sequence of PDC-E2 is provided as Uniprot# P10515 (SEQ ID NO: 130). In some embodiments, the tolerogenic compound may comprise amino acids 163-176 (SEQ ID NO: 131), 159-167 (SEQ ID NO: 132), or 159-176 (SEQ ID NO: 271) of PDC-E2.

[0428] In some embodiments, for autoimmune hepatitis, exemplary antigens for tolerogenic compounds include, but are not limited to, cytochrome P450 2D6 (CYP2D6). An exemplary sequence for CYP2D6 is provided as Uniprot#P10635 (SEQ ID NO: 133). In some embodiments, the tolerogenic compound may comprise amino acids 245-254 (SEQ ID NO: 134), 217-260 (SEQ ID NO: 135), 305-348 (SEQ ID NO: 136), 193-212 (SEQ ID NO: 137), 305-325 (SEQ ID NO: 138), 313-322 (SEQ ID NO: 139), or 393-412 (SEQ ID NO: 140) of CYP2D6.

[0429] In some embodiments, the antigen comprises an alloantigen. In some embodiments, the alloantigen is selected from the group consisting of: subunits of MHC class I and MHC class II haplotype proteins and their complexes with the antigens they present (e.g., donor / recipient differences identified in tissue cross-matching), and minor blood group antigens RhCE, Kell, Kidd, Duffy, Ss, Diego, and MNSs (including single nucleotide polymorphisms), and mimetics, fragments, or portions thereof. In some embodiments, the antigen is associated with graft-versus-host disease, transplant rejection, or autoimmune aplastic anemia. Such compositions can be prepared separately for a given donor / recipient pair.

[0430] Other allergens with further descriptions can be found on the World Wide Web (www.allergen.org), and the nomenclature used herein as standardized by the WHO / IUIS Allergen Nomenclature Subcommittee (eg "Aca f 2").

[0431] In some embodiments, the antigens of the tolerogenic composition may comprise one or more of any antigen sequences disclosed herein, such as one or more of SEQ ID NOs: 40-288, or fragments thereof, or any other antigen generally known in the art.

[0432] Also disclosed herein are compositions comprising any one or more of the tolerogenic compounds disclosed herein, and a pharmaceutically acceptable excipient.

[0433] The pharmaceutical composition can be manufactured in a manner known per se, for example by conventional mixing, dissolving, granulating, dragee making, grinding, emulsifying, encapsulating, embedding or tableting processes. Additionally, the active ingredient is present in an amount effective to achieve its intended purpose. Many compounds for use in the pharmaceutical compositions disclosed herein can be provided as salts with pharmaceutically compatible counterions.

[0434] If desired, the composition may be present in a dispenser device that may contain one or more unit dosage forms containing an active ingredient. The dispenser device may be accompanied by instructions for use. The dispenser may also be accompanied by a notice associated with the container, the form of which is regulated by a government agency that manages the manufacture, use, or sale of the drug, which reflects the agency's approval of the drug form for human or veterinary administration. For example, such a notice may be a prescription drug label approved by the U.S. Food and Drug Administration, or an approved product instruction sheet. Compositions that may include compounds and / or salts described herein formulated in a compatible pharmaceutical carrier may also be prepared, placed in an appropriate container, and labeled for the treatment of the indicated condition.

[0435] Methods of inducing tolerance

[0436] Also disclosed herein are methods of inducing tolerance to an antigen to which a subject is capable of producing an undesirable immune response. In some embodiments, the method comprises administering any one of the tolerogenic compounds or compositions disclosed herein. In some embodiments, the tolerogenic compound or composition is administered before the subject is exposed to the antigen, after the subject is exposed to the antigen, or in both cases. In some embodiments, the undesirable immune response is associated with an allergic reaction to a food, animal, plant, or environmental allergen, an autoimmune disease, a therapeutic agent, or a graft-versus-host disease, or other diseases and conditions associated with any antigen disclosed herein.

[0437] Also disclosed herein are tolerogenic compounds and compositions disclosed herein for inducing tolerance to an antigen in a subject to which the subject is capable of producing an undesirable immune response. In some embodiments, the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease, or other diseases or conditions associated with any of the antigens disclosed herein.

[0438] Also disclosed herein are the tolerogenic compounds and compositions disclosed herein for use in the preparation of a medicament.

[0439] In some embodiments, provided herein are compounds or compositions for treating, preventing, reducing or otherwise changing the immune response to an antigen. In some embodiments, an immune response has occurred or is occurring in a continuing manner. In some embodiments, the treatment of a compound or composition and use are used in a preventive manner. In some embodiments, administration to a subject is performed before, after, or before and after the subject is exposed to an antigen. In some embodiments, administration is performed before exposure to play a preventive role, to substantially avoid or significantly reduce an undesirable immune response.

[0440] The administration of the composition can be carried out via a variety of methods, including but not limited to parenteral, intravenous, infusion, intramuscular, oral, rectal, pulmonary, external, aerosol, transdermal, intradermal or other administration routes. In several embodiments, the composition is delivered in a therapeutically effective amount, for example, by a systemic or local route (e.g., intravenous, intraarterial, local, intramuscular, subcutaneous, intramedullary, intrathecal, intraventricular, intraperitoneal, intranasal, intraocular, etc.). Administration can be carried out at a less frequent time point or substantially equal to a time point per year, per month, per day, per week, multiple times per day, or as needed (e.g., before expected exposure).

[0441] In some embodiments, compound or composition is applied with the amount sufficient to cause the clonal exhaustion and / or anergy of induced T cells, and the T cells have specificity to the antigen of interest.In some embodiments, compound or composition is configured to target hepatocytes and / or LSEC.In some embodiments, compound or composition is configured to induce the expansion of certain colonies or sub-populations of regulatory T cells (such as CD4+CD25+FOXP3+ regulatory T cells).In some embodiments, compound or composition is applied with the amount that can effectively reduce the concentration of antibody, and the antibody has a causal relationship with any one of the diseases disclosed herein or disorders (such as graft-versus-host disease, transplant rejection, immune response for therapeutic agents, autoimmune diseases, hypersensitivity and / or allergies), and the amount reduces the concentration of the antibody in the patient's blood by at least 10%, 20%, 30%, 40% or 50%, or any percentage within the range defined by any two of the above-mentioned reductions.

[0442] The compounds or compositions described herein may be administered to a patient per se, or in combination with other active ingredients, such as in combination therapy.

[0443] In several embodiments, the compound or composition is provided as a unit dose. In several embodiments, the methods and uses disclosed herein include administering a unit dose to a patient or subject. In several embodiments, the unit dose includes a tolerogenic compound of 1 μg / kg to 10 mg / kg relative to the subject's body weight. In several embodiments, the tolerogenic compound administered each time relative to body weight in a single dose is equal to or less than: about 10 μg / kg, about 50 μg / kg, about 75 μg / kg, about 0.1 mg / kg, about 0.2 mg / kg, about 0.5 mg / kg, about 0.75 mg / kg, about 1.0 mg / kg, about 1.5 mg / kg, about 2.0 mg / kg, about 2.5 mg / kg, about 4.0 mg / kg, about 5.0 mg / kg, 10.0 mg / kg or a range spanning and / or including the aforementioned values. In some embodiments, the amount of tolerogenic compounds administered is less than or equal to or equal to a unit dose of 10, 5, 2, 1, 0.5, 0.1, 0.05, 0.01, 0.005, 0.001 or 0.0005 mg per kg body weight. In several embodiments, a dosing regimen is provided in which a subject receives at least one dose of a composition according to an embodiment disclosed herein. In several embodiments, a subject receives at least two, at least three, at least four, at least five or more doses of a composition according to an embodiment disclosed herein. In several embodiments, a given subsequent dose is provided at a concentration less than or equal to the previous dose. For example, when receiving a second dose, if the concentration of the first dose is 0.5 mg / kg, a second dose can be provided at about 0.25 mg / kg. In additional embodiments, the dose remains constant over time. Depending on the severity of the potential immune response (or possible immune response), the dose is optionally increased over time.

[0444] Non-limiting embodiments

[0445] 1. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region (VH), wherein the heavy chain variable region comprises a first heavy chain complementarity determining region (HCDR1), a second heavy chain complementarity determining region HCDR2 and a third heavy chain complementarity determining region HCDR3;

[0446] wherein the HCDR1 comprises the sequence of SEQ ID NO: 9, 297, 329, 361, 393, 425, 457, 489, 521 or 553;

[0447] wherein the HCDR2 comprises the sequence of SEQ ID NO: 10, 298, 330, 362, 394, 426, 458, 490, 522 or 554; and

[0448] wherein the HCDR3 comprises the sequence of SEQ ID NO: 11, 299, 331, 363, 395, 427, 459, 491, 523 or 555.

[0449] 2. The ASGR1 binding polypeptide of embodiment 1, wherein the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:15, 303, 335, 367, 399, 431, 463, 495, 527 or 559.

[0450] 3. The ASGR1 binding polypeptide according to embodiment 1, further comprising a light chain variable region (VL), wherein the light chain variable region comprises a first light chain complementarity determining region (LCDR1), a second light chain complementarity determining region (LCDR2), and a third light chain complementarity determining region (LCDR3);

[0451] wherein the LCDR1 comprises the sequence of SEQ ID NO: 25, 313, 345, 377, 409, 441, 473, 505, 537 or 569;

[0452] wherein the LCDR2 comprises the sequence of SEQ ID NO: 26, 314, 346, 378, 410, 442, 474, 506, 538 or 570; and

[0453] wherein the LCDR3 comprises the sequence of SEQ ID NO: 27, 315, 347, 379, 411, 443, 475, 507, 539 or 571.

[0454] 4. The ASGR1 binding polypeptide of embodiment 3, wherein the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:31, 319, 351, 383, 415, 447, 479, 511, 543 or 575.

[0455] 5. The ASGR1 binding polypeptide of embodiment 1, wherein the heavy chain variable region comprises the sequence of SEQ ID NO: 15, 303, 335, 367, 399, 431, 463, 495, 527 or 559, and the light chain variable region comprises the sequence of SEQ ID NO: 31, 319, 351, 383, 415, 447, 479, 511, 543 or 575.

[0456] 6. The ASGR1 binding polypeptide according to embodiment 1, wherein the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb) or a scFv.

[0457] 7. The ASGR1 binding polypeptide according to embodiment 1, wherein the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

[0458] 8. A polynucleotide encoding the ASGR1 binding polypeptide according to embodiment 1, comprising one or more of the sequences of SEQ ID NO: 12-14, 300-302, 332-334, 364-366, 396-398, 428-430, 460-462, 492-494, 524-526 or 556-558.

[0459] 9. The polynucleotide of embodiment 8, wherein the polynucleotide comprises the sequence of SEQ ID NO: 16, 304, 336, 368, 400, 432, 464, 496, 528 or 560.

[0460] 10. A polynucleotide encoding the ASGR1 binding polypeptide according to embodiment 3, comprising one or more of the sequences of SEQ ID NO: 25-27, 313-315, 345-347, 377-379, 409-411, 441-443, 473-475, 505-507, 537-539 or 569-571.

[0461] 11. The polynucleotide according to embodiment claim 10, wherein the polynucleotide comprises the sequence of SEQ ID NO: 32, 320, 352, 384, 416, 448, 480, 512, 544 or 576.

[0462] 12. The polynucleotide of embodiment 11, wherein the polynucleotide further comprises a sequence of SEQ ID NO: 16, 304, 336, 368, 400, 432, 464, 496, 528 or 560.

[0463] 13. A tolerogenic compound comprising the ASGR1 binding polypeptide according to any one of embodiments 1 to 12, wherein the ASGR1 binding polypeptide is conjugated or fused to an antigen to which tolerance is desired.

[0464] 14. The tolerogenic compound according to embodiment 13, wherein the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker.

[0465] 15. The tolerogenic compound of embodiment 14, wherein the linker is a cleavable linker.

[0466] 16. The tolerogenic compound of embodiment 15, wherein the linker comprises glycine and / or serine, optionally wherein the linker comprises the sequence of SEQ ID NO: 37.

[0467] 17. The tolerogenic compound according to embodiment 13, wherein the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide.

[0468] 18. The tolerogenic compound according to embodiment 13, wherein the antigen comprises a food antigen.

[0469] 19. The tolerogenic compound according to embodiment 18, wherein the food antigen is associated with diarrheal celiac disease.

[0470] 20. The tolerogenic compound of embodiment 19, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to SEQ ID NO:41.

[0471] 21. The tolerogenic compound according to embodiment 19, wherein the antigen comprises the sequence of SEQ ID NO:41.

[0472] 22. The tolerogenic compound according to any one of embodiments 13 to 19, wherein the food antigen is selected from conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), plant cystatin, thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin, and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sina 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitory protein (Sin a 4), inhibitory protein (Api g4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens and a mimetic of any one of the antigens.

[0473] 23. The tolerogenic compound according to any one of embodiments 13 to 19 or 22, wherein the food antigen is selected from the group consisting of high molecular weight glutenins, low molecular weight glutenins, α-, γ- and Ω-gliadins, hordeins, secalins, avenins, a portion of any of said antigens and a mimetic of any of said antigens.

[0474] 24. The tolerogenic compound according to any one of embodiments 13 to 19, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

[0475] 25. The tolerogenic compound according to any one of embodiments 13 to 17, wherein the antigen comprises a self-antigen.

[0476] 26. The tolerogenic compound according to embodiment 25, wherein the autoantigen is selected from thyroglobulin, thyroid peroxidase, thyrotropin receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

[0477] 27. The tolerogenic compound according to any one of embodiments 13 to 17 or 25 to 26, wherein the antigen comprises an antigen associated with an autoimmune disease.

[0478] 28. The tolerogenic compound of embodiment 27, wherein the autoimmune disease is selected from the group consisting of multiple sclerosis, type 1 diabetes, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, celiac disease with diarrhea, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and anti-neutrophil cytoplasmic antibody-associated vasculitis.

[0479] 29. The tolerogenic compound according to embodiment 27 or 28, wherein the autoimmune disease is multiple sclerosis.

[0480] 30. The tolerogenic compound of embodiment 29, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 100, 71-99, 101-106 or 157-159, or a fragment thereof.

[0481] 31. The tolerogenic compound according to embodiment 27 or 28, wherein the autoimmune disease is type 1 diabetes.

[0482] 32. The tolerogenic compound of embodiment 31, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0483] 33. A tolerogenic compound comprising an ASGR1 binding polypeptide conjugated or fused to an antigen to which tolerance is desired, wherein the ASGR1 binding polypeptide comprises a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3;

[0484] wherein the HCDR1 comprises the sequence of SEQ ID NO: 9, 297, 329, 361, 393, 425, 457, 489, 521 or 553;

[0485] wherein the HCDR2 comprises the sequence of SEQ ID NO: 10, 298, 330, 362, 394, 426, 458, 490, 522 or 554; and

[0486] wherein the HCDR3 comprises the sequence of SEQ ID NO: 11, 299, 331, 363, 395, 427, 459, 491, 523 or 555.

[0487] 34. The tolerogenic compound of embodiment 33, wherein the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 15, 303, 335, 367, 399, 431, 463, 495, 527 or 559.

[0488] 35. The tolerogenic compound of embodiment 33 or 34, wherein the ASGR1 binding polypeptide further comprises a light chain variable region comprising LCDR1, LCDR2 and LCDR3;

[0489] wherein the LCDR1 comprises the sequence of SEQ ID NO: 25, 313, 345, 377, 409, 441, 473, 505, 537 or 569;

[0490] wherein the LCDR2 comprises the sequence of SEQ ID NO: 26, 314, 346, 378, 410, 442, 474, 506, 538 or 570; and

[0491] wherein the LCDR3 comprises the sequence of SEQ ID NO: 27, 315, 347, 379, 411, 443, 475, 507, 539 or 571.

[0492] 36. The tolerogenic compound of embodiment 35, wherein the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:31, 319, 351, 383, 415, 447, 479, 511, 543 or 575.

[0493] 37. A tolerogenic compound according to any one of embodiments 33 to 36, wherein the heavy chain variable region comprises the sequence of SEQ ID NO: 15, 303, 335, 367, 399, 431, 463, 495, 527 or 559, and the light chain variable region comprises the sequence of SEQ ID NO: 31, 319, 351, 383, 415, 447, 479, 511, 543 or 575.

[0494] 38. The tolerogenic compound according to any one of embodiments 33 to 37, wherein the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb) or a scFv.

[0495] 39. The tolerogenic compound according to any one of embodiments 33 to 38, wherein the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

[0496] 40. The tolerogenic compound according to embodiments 34 to 39, wherein the antigen is a polypeptide.

[0497] 41. The tolerogenic compound according to any one of embodiments 33 to 40, wherein the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker.

[0498] 42. The tolerogenic compound of embodiment 41, wherein the linker is a cleavable linker.

[0499] 43. The tolerogenic compound of embodiment 41, wherein the linker comprises glycine and / or serine, optionally wherein the linker comprises the sequence of SEQ ID NO: 37.

[0500] 44. The tolerogenic compound according to any one of embodiments 33 to 43, wherein the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide.

[0501] 45. The tolerogenic compound of any one of embodiments 33 to 44, wherein the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced, and the antigen is conjugated or fused to the Fc domain, optionally wherein the antigen is conjugated or fused to the C-terminus of the Fc domain.

[0502] 46. ​​The tolerogenic compound according to any one of embodiments 33 to 35, wherein the antigen comprises a food antigen.

[0503] 47. The tolerogenic compound of embodiment 46, wherein the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), plant cystatin, thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sina 2), lipid transfer protein (Sin a 3), inhibitory protein (Sin a 4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens and a mimetic of any one of the antigens.

[0504] 48. The tolerogenic compound according to embodiment 46 or 47, wherein the food antigen is selected from the group consisting of high molecular weight glutenins, low molecular weight glutenins, α-, γ- and Ω-gliadins, hordeins, secalins, avenins, a portion of any of said antigens and a mimetic of any of said antigens.

[0505] 49. The tolerogenic compound according to any one of embodiments 46 to 48, wherein the food antigen is associated with diarrheal celiac disease.

[0506] 50. The tolerogenic compound of embodiment 49, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

[0507] 51. The tolerogenic compound of any one of embodiments 33 to 45, wherein the antigen comprises a self-antigen.

[0508] 52. The tolerogenic compound of embodiment 51, wherein the autoantigen is selected from thyroglobulin, thyroid peroxidase, thyrotropin receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

[0509] 53. The tolerogenic compound of any one of embodiments 33 to 45 or 51 to 52, wherein the antigen comprises an antigen associated with an autoimmune disease.

[0510] 54. The tolerogenic compound of embodiment 53, wherein the autoimmune disease is selected from the group consisting of type 1 diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, celiac disease with diarrhea, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and anti-neutrophil cytoplasmic antibody-associated vasculitis.

[0511] 55. The tolerogenic compound according to embodiment 53 or 54, wherein the autoimmune disease is type 1 diabetes.

[0512] 56. The tolerogenic compound of embodiment 55, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0513] 57. The tolerogenic compound according to embodiment 53 or 54, wherein the autoimmune disease is multiple sclerosis.

[0514] 58. The tolerogenic compound of embodiment 57, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 71-106 or 157-159, or a fragment thereof.

[0515] 59. The tolerogenic compound according to embodiment 53 or 54, wherein the autoimmune disease is:

[0516] Rheumatoid arthritis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 107-116, 160-239 or 272-288, or a fragment thereof;

[0517] Sjogren's syndrome, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 117-118 or 240-245, or a fragment thereof;

[0518] Rheumatic heart disease, autoimmune myocarditis, viral myocarditis, or inflammatory cardiomyopathy, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 119 or 251-270, or a fragment thereof;

[0519] Parkinson's disease, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 120 or 247-250, or a fragment thereof;

[0520] Anti-neutrophil cytoplasmic antibody-associated vasculitis (ANCA vasculitis), wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 121-129, or a fragment thereof;

[0521] Primary biliary cholangitis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 130-132 or 271, or a fragment thereof; or

[0522] Autoimmune hepatitis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 133-140, or a fragment thereof.

[0523] 60. The tolerogenic compound of any one of embodiments 33 to 45, wherein the antigen comprises an alloantigen.

[0524] 61. The tolerogenic compound according to embodiment 60, wherein the alloantigen is selected from the group consisting of subunits of MHC class I and MHC class II haplotype proteins and their complexes with the antigens they present, and the minor blood group antigens RhCE, Kell, Kidd, Duffy, Diego and MNSs.

[0525] 62. A composition comprising a tolerogenic compound according to any one of embodiments 33 to 61 and a pharmaceutically acceptable excipient.

[0526] 63. A method of inducing tolerance to an antigen to which a subject is capable of producing an undesirable immune response, the method comprising administering to the subject a tolerogenic compound according to any one of embodiments 33 to 61 or a composition according to embodiment 62.

[0527] 64. The method of embodiment 63, wherein the tolerogenic compound or the composition is administered before the subject is exposed to the antigen, after the subject is exposed to the antigen, or both.

[0528] 65. The method of embodiment 63 or 64, wherein the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

[0529] 66. A tolerogenic compound according to any one of embodiments 33 to 45 or a composition according to embodiment 63 for use in inducing tolerance to an antigen in a subject to which the subject is capable of generating an undesirable immune response.

[0530] 67. The tolerogenic compound or composition for use according to embodiment 66, wherein the undesired immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

[0531] 68. A tolerogenic compound according to any one of embodiments 33 to 45 or a composition according to embodiment 62 for use in the preparation of a medicament.

[0532] 69. A compound according to any one of Embodiments 33 to 45 or a composition according to Embodiment 62 for use in inducing immune tolerance in a subject in need thereof.

[0533] 70. A method of inducing tolerance to an antigen to which tolerance is desired, the method comprising:

[0534] A compound comprising:

[0535] (i) an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3;

[0536] wherein the HCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO: 9, 297, 329, 361, 393, 425, 457, 489, 521 or 553;

[0537] wherein the HCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO: 10, 298, 330, 362, 394, 426, 458, 490, 522 or 554; and

[0538] wherein the HCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO: 11, 299, 331, 363, 395, 427, 459, 491, 523 or 555; and

[0539] (ii) the antigen to which tolerance is desired,

[0540] The ASGR1 binding polypeptide is conjugated or fused to the antigen to which tolerance is desired.

[0541] 71. The method of embodiment 70, wherein the ASGR1 binding polypeptide further comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3;

[0542] wherein the LCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO: 25, 313, 345, 377, 409, 441, 473, 505, 537 or 569;

[0543] wherein the LCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO: 26, 314, 346, 378, 410, 442, 474, 506, 538 or 570; and

[0544] wherein the LCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO:27, 315, 347, 379, 411, 443, 475, 507, 539 or 571.

[0545] 72. The method of embodiment 70 or 71, wherein the antigen to which tolerance is desired is associated with diarrheal celiac disease.

[0546] 73. A method according to embodiment 72, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 41, 40, 43-54, or a fragment thereof.

[0547] 74. The method of embodiment 70 or 71, wherein the antigen to which tolerance is desired is associated with MS.

[0548] 75. A method according to embodiment 74, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 100, 71-99, 101-106 or 157-159, or a fragment thereof.

[0549] 76. The method of embodiment 70 or 71, wherein the antigen to which tolerance is desired is associated with type 1 diabetes.

[0550] 77. A method according to embodiment 76, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

[0551] 78. The method of any one of embodiments 70 to 77, wherein the administration is via an intravenous route.

[0552] 79. A method of delivering an antigen to liver tissue of a subject, the method comprising:

[0553] conjugating or fusing the antigen to an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide, thereby generating an ASGR1 binding polypeptide-antigen complex; and

[0554] contacting the ASGR1 binding polypeptide-antigen complex with liver tissue of a subject in situ,

[0555] wherein the contacting allows the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, thereby delivering the antigen to the liver tissue.

[0556] 80. A method of delivering an antigen to liver tissue of a subject, the method comprising:

[0557] contacting an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide conjugated or fused to an antigen with liver tissue of a subject in situ, thereby allowing the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue,

[0558] wherein the ASGR1 binding polypeptide comprises a heavy chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3;

[0559] wherein the HCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO: 9, 297, 329, 361, 393, 425, 457, 489, 521 or 553;

[0560] wherein the HCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO: 10, 298, 330, 362, 394, 426, 458, 490, 522 or 554; and

[0561] wherein the HCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO: 11, 299, 331, 363, 395, 427, 459, 491, 523 or 555; and

[0562] wherein the ASGR1 binding polypeptide comprises a light chain variable region, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3;

[0563] wherein the LCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO: 25, 313, 345, 377, 409, 441, 473, 505, 537 or 569;

[0564] wherein the LCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO: 26, 314, 346, 378, 410, 442, 474, 506, 538 or 570; and

[0565] wherein the LCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO:27, 315, 347, 379, 411, 443, 475, 507, 539 or 571.

[0566] 81. The method of embodiment 70 or 80, wherein delivering the antigen in situ to liver tissue of the subject induces tolerance to the antigen in the subject.

[0567] 82. The method of any one of embodiments 80 to 81 for treating an autoimmune disease.

[0568] 83. The method of any one of embodiments 80 to 82, for treating an allergic reaction.

[0569] Example

[0570] Additional embodiments are disclosed in further detail in the following examples, which are not intended in any way to limit the scope of the claims.Methods and Materials represent non-limiting embodiments of how to make, screen, test, or otherwise evaluate the compounds disclosed herein.

[0571] Example 1: Materials and Methods

[0572] An antibody was generated against the extracellular domain (ECD) of human ASGR1 (SEQ ID NO: 34). The ECD corresponds to amino acids Q62-L291 of the full-length wild-type ASGR1 sequence (SEQ ID NO: 33).

[0573] ASGR1 preparation

[0574] ASGR1 was biotinylated using the EZ-Link Sulfo-NHS-Biotinylation Kit (Thermo Scientific, Cat. No. 21425).

[0575] The ASGR1 protein was concentrated to approximately 1 mg / mL and the buffer was exchanged to PBS before adding the biotinylation reagent at a 1:7.5 molar ratio. The mixture was kept at 4°C overnight before another buffer exchange to remove free biotin in solution. Biotinylation was confirmed by binding of the streptavidin sensor of the labeled protein on the ForteBio.

[0576] Naive library selection

[0577] Eight naive human synthetic yeast libraries, each with approximately 109 diversity, were amplified as previously described (see, e.g., Y. Xu et al, PEDS 26(10), 663-70 (2013); WO2009036379; WO2010105256; and WO2012009568, each of which is expressly incorporated herein by reference in its entirety.)

[0578] For the first two rounds of selection, magnetic bead sorting using the Miltenyi MACS system was performed as described previously (see, e.g., Siegel et al, J Immunol Methods 286(1-2), 141-153 (2004), which is expressly incorporated herein by reference in its entirety). Briefly, yeast cells (approximately 10 10Cells / library) were incubated with 50nM biotinylated human ASGR1-His in wash buffer (HEPES buffered saline (HBS) / 0.1% bovine serum albumin (BSA) / 1mM CaCl2) at 30°C for 30 minutes. After washing once with 40mL ice-cold wash buffer, the cell pellet was resuspended in 20mL wash buffer, and streptavidin microbeads (500μl) were added to the yeast and incubated at 4°C for 15 minutes. Next, the yeast was pelleted, resuspended in 5mL wash buffer, and then loaded onto a Miltenyi LS column. After loading 5mL, the column was washed 3 times with 3mL wash buffer. The column was then removed from the magnetic field, and the yeast was eluted with 5mL growth medium and then grown overnight.

[0579] The following rounds of selection were performed using flow cytometry (FACS). To obtain species cross-reactivity, yeast was pelleted, washed three times with wash buffer, and incubated with 50 nM biotinylated human ASGR1-His, 50 nM biotinylated cynomolgus ASGR1-His, 50 nM biotinylated mouse ASGR1-His, or 50 nM biotinylated rat ASGR1-His at 30 ° C, or incubated with a multi-specific reagent (PSR) to remove non-specific antibodies from the selection. As previously described, for PSR depletion, the library was incubated with a 1:10 dilution of biotinylated PSR reagent (see, e.g., Y.Xu et al, PEDS 26 (10), 663-70 (2013)). Yeast was also incubated with 50 nM biotinylated human ASGR1-His without CaCl2 to enrich for calcium-dependent antibodies. After various incubations, the yeast was washed twice with wash buffer and stained at 4°C for 15 minutes with goat F(ab')2 anti-human κ-FITC (LC-FITC) (Southern Biotech, catalog number 2062-02) diluted 1:100 and streptavidin-AF633 (SA-633) (Life Technologies, catalog number S21375) diluted 1:500 or Extravidin-phycoerythrin (EA-PE) (Sigma-Aldrich, catalog number E4011) diluted 1:50. After washing twice with ice-cold wash buffer, the cell pellet was resuspended in 0.3mL wash buffer and transferred to a sorting tube covered with a filter. Sorting was performed using a FACS ARIA sorter (BD Biosciences), and a sorting gate was determined to select antibodies with desired characteristics. Selection rounds were repeated until a population with all desired characteristics was obtained. After the last round of sorting, yeast was plated and single colonies were selected for characterization.

[0580] Light chain batch disruption diversification

[0581] The light chain diversification libraries for additional selection rounds were prepared using heavy chains from the naive output. As described above, these libraries were selected, i.e., using one round of MACS and four rounds of FACS. In different FACS selection rounds, the libraries were evaluated by ASGR1 titration for, for example, PSR binding, species cross-reactivity, calcium dependency, and affinity pressure. Sorting was performed to obtain a population with the desired characteristics. Single colonies from each final FACS selection round were selected for sequencing and characterization.

[0582] Antibody production and purification

[0583] Yeast clones were grown to saturation and then induced with shaking at 30°C for 48 hours. After induction, yeast cells were pelleted and the supernatant was harvested for purification. IgG was purified using a protein A column and eluted with acetic acid at pH 3.5.

[0584] ForteBio Octet KD Measurement

[0585] As previously described, ForteBio affinity measurements are typically performed on Octet HTX (see, e.g., Estep et al, Mabs 5(2), 270-278 (2013)). In brief, ForteBio affinity measurements are performed by loading IgG online onto the AHC sensor. The sensor is equilibrated offline in assay buffer for 30 minutes and then monitored online for 60 seconds to establish a baseline. The sensor loaded with IgG is exposed to 100 nM ASGR1 for 3 minutes, then transferred to assay buffer for 3 minutes for off-rate measurements. All kinetics are analyzed using a 1:1 binding model.

[0586] ForteBio Epitope Binning

[0587] Epitope binning was performed using a standard sandwich format cross-blocking assay. A control anti-ASGR1 IgG was loaded onto the AHQ sensor, and unoccupied Fc binding sites on the sensor were blocked with an irrelevant human IgG1 antibody. The sensor was then exposed to 100 nM human ASGR1-His followed by exposure to a secondary anti-ASGR1 antibody. Additional binding of the secondary antibody after ASGR1 association indicated the presence of an unoccupied epitope (non-competitor), while the absence of binding indicated epitope blocking (competitor).

[0588] Cell binding assay

[0589] Approximately 200,000 cells overexpressing human ASGR1 or parental cells were washed with wash buffer and incubated with 100 μL of 100 nM IgG at room temperature for 15 minutes. The cells were then washed twice with wash buffer and incubated with 100 μL of 1:100 anti-human IgG R-PE (Southern Biotech, catalog number 2040-09) on ice for 15 minutes. The cells were then washed twice with wash buffer and analyzed on a FACS Canto II analyzer (BD Biosciences).

[0590] Example 2: Non-limiting examples of anti-ASGR1 antibodies

[0591] Several non-limiting examples of anti-ASGR1 antibodies were identified by this process. The antibodies comprised sequences as detailed in Table 2 and are also depicted in Figures 1A-1F , 6A-6F, 8A-8F, 9A-9F, 11A-11F, 12A-12F, 13A-13F, 14A-14F, 15A-15F and 16A-16F. Based on routine experimentation, it can be assumed that a certain degree of sequence difference is tolerated without significantly affecting ASGR1 binding. The anti-ASGR1 antibodies are referred to as mAb-60819, mAb-60856, mAb-60881, mAb-60869, mAb-83198, mAb-83226, mAb-83237, mAb-83245, mAb-83257, mAb-83256, and are referred to elsewhere herein by their terminal three digits, e.g., "819".

[0592] Table 2: Anti-ASGR1 antibody sequences

[0593]

[0594]

[0595] mAb-60819 was biochemically characterized and its apparent KD (KDapp) was found to be 9.29 x 10- 9 M. mAb-60856 was biochemically characterized and its apparent KD (KDapp) was found to be 9.29 x 10- 9 M. mAb-60881 was biochemically characterized and its apparent KD (KDapp) was found to be approximately 3 x 10- 9 M. mAb-60869 was biochemically characterized and its apparent KD (KDapp) was found to be 3.22 x 10- 9 M. ASGR1 is dependent on calcium (Ca) for carbohydrate binding 2+ ions). Proteins have active sites where carbohydrates bind to the active site calcium as well as calcium ions at the proximal and distal sites, all of which stabilize the protein structure. Therefore, the pH and Ca binding of anti-ASGR1 antibodies were evaluated by surface plasmon resonance using the Biacore system, bio-layer interferometry using the Octet system, and using a cell-based assay. 2 + dependence. mAb-60819 was found to have pH and Ca 2 + dependence, such that a significant reduction in binding was detected in the absence of calcium. mAb-60856, mAb-60881, and mAb-60869 were found to have no pH and Ca dependence. 2 +-dependency, such that the binding of these antibodies to the ASGR1 target is not affected by the absence or removal of calcium ions.

[0596] Example 3: Induction of immune tolerance in a type 1 diabetes model

[0597] Alternative forms and novel liver-targeted immune tolerance platforms of different capabilities have been previously described in U.S. Patent No. 10821157, U.S. Publication No. 2020 / 0101146, and PCT Publication No. WO 2021 / 053589, in which antigen-conjugated sugar-containing polymers are specifically targeted to the liver to induce antigen-specific immune tolerance, each of which is expressly incorporated by reference in its entirety. This liver-targeted technology induces immune tolerance by deleting antigen-specific T cells, reducing antigen-specific inflammatory cytokine production, and enhancing regulatory T cells (Treg). A novel antigen delivery platform was envisioned and developed, which uses a monoclonal antibody (mAb) specific for ASGR1 (αASGR1) expressed by hepatocytes, including hepatocytes. It was examined whether antigens targeted to the liver using αASGR1 antibodies induce immune tolerance in the BDC2.5 mouse model of type 1 diabetes (T1D). In this model, the non-limiting example of using αASGR1 antibodies (mAb1) to target antigens to the liver delayed the onset of diabetes, which indicates the induction of antigen-specific immune tolerance in this model.

[0598] The BDC2.5 TCR transgenic mouse model of T1D was used (NOD.Cg-Tg(TcraBDC2.5,TcrbBDC2.5)1Doi / DoiJ [Jackson Laboratories]). BDC2.5 TCR transgenic mice contain diabetogenic CD4+ T cells specific for a region of chromogranin A (ChgA) that is post-translationally associated with the c-peptide fragment of proinsulin. BDC2.5 CD4+ T cells also recognize major histocompatibility class II (MHC-II) I-Ag 7The restricted peptide p31 is one of several non-natural mimotope peptides recognized by BCD2.5 T cells. 7 is a mouse major histocompatibility complex that predisposes non-obese diabetic mice to autoimmune diabetes. When adoptively transferred into NOD.SCID mice, BDC2.5 transgenic T cells induce T1D pathology marked by insulitis and hyperglycemia. As described herein, we evaluated whether p31 delivered to the liver using αASGR1 mAb induces antigen-specific tolerance to prevent and / or delay disease in this very aggressive model of T1D.

[0599] The BDC2.5 mouse model of T1D was used to evaluate the efficacy of recombinantly expressed αASGR1-p31 fusions as tolerogens. In this model, activated BDC2.5 TCR transgenic splenocytes were adoptively transferred into non-obese diabetic (NOD) mice carrying the severe combined immunodeficiency (SCID) mutation. In control animals that received administration of vehicle (normal saline), the onset of diabetes was observed 10 to 14 days after adoptive transfer.

[0600] NOD.SCID(NOD.Cg-Prkdc <scid> / J [Jackson Laboratories]): Due to a mutation in the protein kinase PRKDC, these immunodeficient mice do not develop mature T and B lymphocytes, which are required for VDJ recombination of antigen receptor genes in germ cells. NOD.SCID mice do not spontaneously develop diabetes unless they receive adoptive transfer of diabetogenic T cells.

[0601] The primary goal was to test whether delivery of p31 peptide to the liver using αASGR1 mAb could prevent the onset of diabetes. On day 0, 3x 10 5 BDC2.5 splenocytes were adoptively transferred into NOD.SCID mice by intravenous injection (iv). On days 0 and 4 after cell transfer, NOD.SCID mouse recipients received intravenous administration of αASGR1-p31, free p31 peptide, or saline. Mouse blood glucose levels were monitored 2-3 times per week. Diabetes onset was defined as two consecutive blood glucose measurements ≥ 250 mg / dL.

[0602] Antigen design and tolerogen preparation: The tolerogen used in this study was designed to include a p31 mimetic epitope sequence (YVRPLWVRME (SEQ ID NO: 36)) recognized by BDC2.5CD4+ T cells. The αASGR antibody-p31 tolerogen (mAb1-p31) includes a glycine-serine linker (e.g., SEQ ID NO: 37) between the heavy chain C-terminus and the p31 antigen (e.g., SEQ ID NO: 39).

[0603] mAb1-p31: p31 was recombinantly expressed and fused downstream to the end of the heavy chain of the exemplary αASGR1 mAb (mAb1) using a short glycine-serine (GS) linker. Other exemplary antibodies specific for ASGR1 and / or ASGR2 can be found in U.S. Patents 9771427 and 10358497, each of which is expressly incorporated herein by reference in its entirety. An example of a GS linker-p31 peptide sequence that can be fused to the heavy chain of the αASGR1 antibody (or other permitted antibody region) is depicted in SEQ ID NO:39. DNA sequences corresponding to the GS linker and p31 mimetic epitopes were synthesized and cloned into modified pcDNA3.4 vectors containing DNA corresponding to the αASGR1 antibody. αASGR1-p31 was produced by transient transfection at a scale of 480 mL for 5 days in Expi293 cells using the manufacturer's protocol. Cell supernatants were harvested by centrifugation and filtered through a 0.2 μm aPES membrane filter to remove cells and debris prior to antibody purification.

[0604] Animals and in vivo procedures: These studies followed animal guidelines approved by the Institutional Animal Care and Use Committee. Upon arrival, recipient NOD.SCID mice were randomly removed from shipping containers and assigned to cages.

[0605] Isolation and Activation of Splenocytes for Adoptive Transfer: BDC2.5 spleens were collected and processed into cell suspensions, which were then activated with p31 peptides. Briefly, spleens were collected in 5 mL of Iscove's modified Dulbecco's medium (IMDM), and the spleens were gently ground using a syringe plunger and broken through a 70 μm cell strainer. The cell strainer was then washed with 5 mL of IMDM, and the cell suspension was centrifuged at 500 x g for 5 minutes at 4 ° C. The supernatant was aspirated, the precipitate was broken and resuspended in 1 mL of potassium ammonium chloride (ACK) red blood cell lysis buffer, and incubated at room temperature for 2 minutes. The cells were washed twice with IMDM, transferred to a new 15 mL conical tube through a 30 μm filter, and counted. The cells were centrifuged again and resuspended in IMDM culture medium. The spleen cells were then treated with 0.5 μM of p31 peptide at 1 x 10 6 The cells were cultured at a cell density of 10 cells / mL at 37°C with 5% CO2 for 4 days. The activated BDC2.5 splenocytes were washed in IMDM medium and cultured at 3x 10 6 The cells were resuspended at a density of 10 cells / mL and 0.1 mL of the cell suspension was injected intravenously into mice (adoptive transfer).

[0606] Blood Glucose Measurement: Blood was collected by making an incision at the tip of the mouse tail with an insulin syringe. Blood glucose levels were determined using a handheld blood glucose meter (Lifescan, Inc.) and blood glucose test strips (Gen Ultimate). The day of diabetes onset was recorded as the first of two consecutive blood glucose measurements ≥ 250 mg / dL. Diabetic mice were euthanized.

[0607] Program overview of the BDC2.5 model: Twenty-one (21) NOD.SCID female mice were used as recipients and one (1) BDC2.5 female mouse was used as a spleen cell donor. The recipient mice were divided into three (3) experimental groups (Table 3). The mice were monitored 2-3 times per week using a handheld blood glucose meter. The following procedures were performed during the in vivo portion of this study:

[0608] Day 4: Splenocytes were isolated from BDC2.5 mice and treated with 0.5 μM p31 peptide at 1 x 10 6 The cells were cultured at a cell density of 10 cells / mL at 37°C with 5% CO2 for 4 days.

[0609] Day 0: Recipient NOD.SCID mice were divided into 4 groups (Table 3). Cultured splenocytes were harvested, washed and prepared into single cell suspensions. 3 x 10 5 Cells / mouse were adoptively transferred into all recipients by intravenous injection.

[0610] Day 0 and Day 4: Mice were injected intravenously with mAb1-p31, p31 or saline (Table 3). Doses were calculated based on the average body weight of all recipient mice in this study (21.3 g). Baseline body weight and blood glucose were measured 2 days before the start of this study.

[0611] Days 4-105: Mice were monitored for T1D by twice weekly blood glucose measurements. Mice were considered diabetic after two consecutive blood glucose measurements ≥ 250 mg / dL.

[0612] Table 3: Experimental groups of the BDC2.5 model

[0613]

[0614]

[0615] result:

[0616] Liver-targeted p31 antigen induces immune tolerance and prolongs diabetes-free survival in the BDC2.5 model of T1D: Animals treated with mAbl-p31 showed prolonged diabetes-free survival ([MST] = 91 days) when compared to p31- and saline-treated controls (MST 9 and 8 days, respectively) (p = 0.0001, Figure 3 , Table 4; means were compared with p31-treated controls using the Mantel-Cox log-rank test). These data demonstrate that αASGR1 mAb1-p31 protects mice from T1D by inducing p31-specific immune tolerance even when diabetogenic T cells were highly preactivated prior to adoptive transfer.

[0617] Table 4: Survival statistics of the BDC2.5 model for T1D

[0618] Group Tolerogen Median diabetes-free survival time (days) p-value 1 mAb1-p31 91 0.0001 2 p31 9 n / a 3 Normal saline 8 0.093

[0619] Example 4: Immunization in the BDC2.5 Mouse T1D Model Using a Non-Limiting Embodiment of an Anti-ASGR1 Antibody Tolerance

[0620] Anti-ASGR1 antibodies mAb-60819, mAb-60856, mAb-60869 as described herein (e.g., VH having the sequence of SEQ ID NOs: 15, 303, 367, and VL having the sequence of SEQ ID NOs: 31, 319, 383, respectively) were used to prepare p31 tolerogens for use in BDC2.5

[0621] The p31 peptide was fused to the C-terminus of the antibody heavy chain using a glycine-serine linker.

[0622] Optionally, the effector functions of the Fc region of the antibody may be silenced so that it does not bind to FcRn or other FcRy receptors.The Fc portion of the antibody is not required for the desired immune tolerance effect.

[0623] According to the method described in Example 2, the anti-ASGR1 antibody tolerogen was administered to the NOD.SCID T1D mouse model adoptively transferred with BCD2.5 splenocytes. Figure 4 , 7 As depicted in Figures 10 and 10, mAb-60819, mAb-60856, and mAb-60869 significantly delayed the development of T1D in the NOD.SCID model compared to unconjugated p31 peptide or saline controls.

[0624] Example 5: Induction of immune tolerance in the EAE mouse model of multiple sclerosis

[0625] The effects of the tolerogens disclosed herein on immune tolerance in a mouse model of human multiple sclerosis (MS) were also investigated.The animal model was an experimental autoimmune encephalomyelitis (EAE) adoptive transfer mouse model.

[0626] Readouts of immune tolerance include changes in the following parameters: 1. Disease severity as measured by disease score (paralysis), 2. Weight loss, 3. Inflammatory cytokine production by myelin-specific CD4+ T cells.

[0627] EAE is an established mouse model of MS that has been widely used in preclinical development of other MS therapies, in which the transfer of activated autoreactive CD4+T cells induces MS-like disease. Given that the transferred cells treated with this therapy are highly activated, the model closely mimics the therapeutic environment in which patients have circulating activated T cells that recognize self-antigens. The findings herein are believed to be applicable to other autoimmune diseases (such as diarrheal celiac disease) in which immunopathology is driven by autoreactive T cells.

[0628] In brief, in the EAE adoptive transfer model, donor mice are immunized to generate antigen-specific encephalitogenic T cells. Then, splenocytes from these mice are activated and amplified in cell culture and transferred intraperitoneally (ip) into recipient mice, which induces symptomatic EAE. The antigen used for vaccination in this model is an immunodominant peptide fragment of myelin oligodendrocyte glycoprotein (MOG), amino acid sequence 35-55. The similar region of human MOG is highly homologous to mouse MOG, and is an identified T cell epitope in human MS, which spans amino acids 34-56 of human MOG, which is the same as in mice. As disclosed herein, other fragments of human MOG can be used in several embodiments, such as MOG 1-60, MOG 33-62, or any other MOG fragment disclosed herein or otherwise known in the art.

[0629] The efficacy of the EAE model was measured by the reduction of several clinical parameters including:

[0630] 1) EAE incidence, which is the percentage of mice that became ill at any time during the study, regardless of disease severity or subsequent remission.

[0631] 2) Mean Maximum Score (MMS), which is calculated as the average of the highest clinical score achieved by each individual subject at any time during the study. This is considered the primary measure of disease severity. The theoretical range is 0 to 5, depending on the degree of mobility.

[0632] 3) Weight loss, which is expected to be positively correlated with an increase in disease scores.

[0633] 4) Median day to disease onset, which is the median day that a group of animals exhibited a clinical score greater than 0.

[0634] 5) Mean Final Score, which is the mean clinical score for a group at the end of the study. Because disease scores spontaneously decreased later in the study, this measure was considered less relevant to overall efficacy.

[0635] Antigen design: The EAE induction model involves inoculating mice with an immunodominant peptide domain of mouse MOG (MOG 35-55; SEQ ID NO: 79); inoculation induces encephalitogenic CD4+ T cells that are subsequently expanded ex vivo and adoptively transferred into recipient mice and mediate disease. The tolerogen may include the immunodominant peptide domain of MOG fused to a linker, such as a cysteine-containing linker for conjugation to an ASGR1 binding polypeptide, or other linkers for fusion to antibodies, such as linkers containing glycine and serine residues, as contemplated herein. The homologous immunodominant peptide domain of human MOG (MOG 35-55; SEQ ID NO: 78) may be used.

[0636] EAE was induced in C57BL / 6 recipient mice by intraperitoneal injection of encephalitogenic T cells derived from B6.SJL donor mice immunized with MOG 35-55 on day 0. Test condition mice were also treated with liver-targeted MOG 35-55 antigen.

[0637] The negative control group for treatment was mice treated with vehicle (saline), where the mice were expected to develop EAE.

[0638] EAE model mice administered with encephalitogenic T cells were treated with MOG 33-62 ("MOG10") conjugated to mAb-60819 (819-MOG10). 819-MOG10 was administered at a dose level of 2 pmol / g or 10 pmol / g. Figure 5 As seen in Figure 2, administration of 819-MOG10 at 10 pmol / g resulted in >90% efficacy within 20 days after induction of EAE. This efficacy was dose-dependent. These data suggest that myelin-derived autoantigens (MOG) delivered to the liver by αASGR1 mAb-60819 induce antigen-specific immune tolerance and prevent undesirable immune responses associated with autoimmune diseases (here, as a non-limiting example, MS).

[0639] As disclosed herein, in several embodiments, targeting an antigen to which tolerance is desired (such as an antigen associated with MS, celiac disease, or type 1 diabetes, as non-limiting examples) to the liver by an anti-ASGR1 antibody results in antigen-specific immune tolerance to prevent undesirable immune responses associated with a disease (such as MS, celiac disease, or type 1 diabetes, as non-limiting examples) to which tolerance is desired. As disclosed herein, in several embodiments, targeting an antigen to which tolerance is desired to be ...

[0640] In at least some previously described embodiments, one or more elements used in an embodiment may be interchangeably used in another embodiment unless such replacement is technically infeasible. It will be appreciated by those skilled in the art that various other omissions, additions and modifications may be made to the methods and structures described herein without departing from the scope of the claimed subject matter. All such modifications and changes a...

Claims

1. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region (VH), wherein the heavy chain variable region comprises a first heavy chain complementarity determining region (HCDR1), a second heavy chain complementarity determining region (HCDR2) and a third heavy chain complementarity determining region (HCDR3); wherein the HCDR1 comprises the sequence of SEQ ID NO:9; wherein the HCDR2 comprises the sequence of SEQ ID NO: 10; and Wherein the HCDR3 comprises the sequence of SEQ ID NO:

11.

2. The ASGR1 binding polypeptide of claim 1, wherein the heavy chain variable region comprises a sequence that is at least 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:

15.

3. The ASGR1 binding polypeptide of claim 1, further comprising a light chain variable region (VL), wherein the light chain variable region comprises a first light chain complementarity determining region (LCDR1), a second light chain complementarity determining region (LCDR2), and a third light chain complementarity determining region (LCDR3); wherein the LCDR1 comprises the sequence of SEQ ID NO:25; wherein the LCDR2 comprises the sequence of SEQ ID NO: 26; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

27.

4. The ASGR1 binding polypeptide of claim 3, wherein the light chain variable region comprises a sequence that is at least 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:

31. 5 . The ASGR1 binding polypeptide of claim 1 , wherein the heavy chain variable region comprises the sequence of SEQ ID NO: 15, and the light chain variable region comprises the sequence of SEQ ID NO:

31. 6 . The ASGR1 binding polypeptide of claim 1 , wherein the ASGR1 binding polypeptide is an antibody, a Fab′ fragment, a F(ab′) 2 fragment, a domain antibody (dAb), or a scFv.

7. The ASGR1 binding polypeptide of claim 1, wherein the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

8. A polynucleotide encoding the ASGR1 binding polypeptide according to claim 1, wherein the polynucleotide comprises one or more of the sequences of SEQ ID NOs: 12-14.

9. The polynucleotide of claim 8, wherein the polynucleotide comprises the sequence of SEQ ID NO:

16. 10 . A polynucleotide encoding the ASGR1 binding polypeptide according to claim 3 , wherein the polynucleotide comprises one or more of the sequences of SEQ ID NOs: 25-27.

11. The polynucleotide of claim 10, wherein the polynucleotide comprises the sequence of SEQ ID NO:

32.

12. The polynucleotide of claim 11, wherein the polynucleotide further comprises the sequence of SEQ ID NO:

16.

13. A tolerogenic compound comprising an ASGR1 binding polypeptide according to any one of claims 1 to 12, wherein the ASGR1 binding polypeptide is conjugated or fused to an antigen to which tolerance is desired.

14. The tolerogenic compound of claim 13, wherein the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker.

15. The tolerogenic compound of claim 14, wherein the linker is a cleavable linker.

16. The tolerogenic compound of claim 15, wherein the linker comprises glycine and / or serine, optionally wherein the linker comprises the sequence of SEQ ID NO:

37.

17. The tolerogenic compound of claim 13, wherein the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide.

18. The tolerogenic compound of claim 13, wherein the antigen comprises a food antigen.

19. The tolerogenic compound of claim 18, wherein the food antigen is associated with diarrheal celiac disease.

20. The tolerogenic compound of claim 19, wherein the antigen comprises a sequence having at least 95%, 96%, 97%, 98%, 99% or 100% sequence identity to SEQ ID NO:

41.

21. The tolerogenic compound of claim 19, wherein the antigen comprises SEQ ID NO:

41.

22. The tolerogenic compound according to any one of claims 13 to 19, wherein the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mald 1.03D (Mal d 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), plant cystatin, thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin, and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitory protein (Sin a 4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens and a mimetic of any one of the antigens.

23. The tolerogenic compound according to any one of claims 13 to 19 or 22, wherein the food antigen is selected from the group consisting of high molecular weight glutenins, low molecular weight glutenins, α-, γ- and Ω-gliadins, hordeins, secalins, avenins, a portion of any of said antigens and a mimetic of any of said antigens.

24. The tolerogenic compound of any one of claims 13 to 19, wherein the antigen comprises a sequence having at least 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

25. The tolerogenic compound of any one of claims 13 to 17, wherein the antigen comprises a self-antigen.

26. The tolerogenic compound of claim 25, wherein the autoantigen is selected from the group consisting of thyroglobulin, thyroid peroxidase, thyrotropin receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

27. The tolerogenic compound of any one of claims 13 to 17 or 25 to 26, wherein the antigen comprises an antigen associated with an autoimmune disease.

28. The tolerogenic compound of claim 27, wherein the autoimmune disease is selected from the group consisting of multiple sclerosis, type 1 diabetes, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, celiac disease with diarrhea, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and antineutrophil cytoplasmic antibody-associated vasculitis.

29. The tolerogenic compound according to claim 27 or 28, wherein the autoimmune disease is multiple sclerosis.

30. The tolerogenic compound of claim 29, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 100, 71-99, 101-106 or 157-159, or a fragment thereof.

31. The tolerogenic compound of claim 27 or 28, wherein the autoimmune disease is type 1 diabetes.

32. The tolerogenic compound of claim 31, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

33. A tolerogenic compound comprising an ASGR1 binding polypeptide conjugated or fused to an antigen to which tolerance is desired, wherein the ASGR1 binding polypeptide comprises a heavy chain variable region comprising one, two or all three of HCDR1, HCDR2 and HCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO:9; wherein the HCDR2 comprises the sequence of SEQ ID NO: 10; and / or Wherein the HCDR3 comprises the sequence of SEQ ID NO:

11.

34. The tolerogenic compound of claim 33, wherein the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:

15.

35. The tolerogenic compound of claim 33 or 34, wherein the ASGR1 binding polypeptide further comprises a light chain variable region comprising one, two or all three of LCDR1, LCDR2 and LCDR3; wherein the LCDR1 comprises the sequence of SEQ ID NO:25; wherein the LCDR2 comprises the sequence of SEQ ID NO: 26; and / or Wherein the LCDR3 comprises the sequence of SEQ ID NO:

27.

36. The tolerogenic compound of claim 35, wherein the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:

31.

37. The tolerogenic compound according to any one of claims 33 to 36, wherein the heavy chain variable region comprises the sequence of SEQ ID NO: 15, and the light chain variable region comprises the sequence of SEQ ID NO:

31.

38. The tolerogenic compound according to any one of claims 33 to 37, wherein the ASGR1 binding polypeptide is an antibody, a Fab' fragment, a F(ab')2 fragment, a domain antibody (dAb) or a scFv.

39. The tolerogenic compound of any one of claims 33 to 38, wherein the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced.

40. The tolerogenic compound according to claims 34 to 39, wherein the antigen is a polypeptide.

41. The tolerogenic compound according to any one of claims 33 to 40, wherein the ASGR1 binding polypeptide and the antigen are conjugated or fused using a linker, optionally wherein the linker is a polypeptide linker or a chemical conjugation linker.

42. The tolerogenic compound of claim 41, wherein the linker is a cleavable linker.

43. The tolerogenic compound of claim 41, wherein the linker comprises glycine and / or serine, optionally wherein the linker comprises the sequence of SEQ ID NO:

37.

44. The tolerogenic compound according to any one of claims 33 to 43, wherein the antigen is conjugated or fused to the N-terminus or C-terminus of the ASGR1 binding polypeptide.

45. The tolerogenic compound of any one of claims 33 to 44, wherein the ASGR1 binding polypeptide comprises an Fc domain, optionally wherein the Fc domain is silenced, and the antigen is conjugated or fused to the Fc domain, optionally wherein the antigen is conjugated or fused to the C-terminus of the Fc domain.

46. ​​The tolerogenic compound according to any one of claims 33 to 35, wherein the antigen comprises a food antigen.

47. The tolerogenic compound of claim 46, wherein the food antigen is selected from the group consisting of conarachidin (Ara h 1), allergen II (Ara h 2), peanut agglutinin, lupin globulin (Ara h 6), 31 kda major allergen / disease resistance protein homolog (Mal d 2), lipid transfer protein precursor (Mal d 3), major allergen Mal d 1.03D (Mald 1), alpha-lactalbumin (ALA), lactotransferrin, actin (Act c 1, Act d 1), plant cystatin, thaumatin-like protein (Act d 2), kiwellin (Act d 5), ovomucoid, ovalbumin, ovotransferrin and lysozyme, yolk globulin, apovitillin, vosvetin, 2S albumin (Sin a 1), 11S globulin (Sin a 2), lipid transfer protein (Sin a 3), inhibitor protein (Sin a 4), inhibitory protein (Api g 4), high molecular weight glycoprotein (Api g 5), tropomyosin (Pen a 1), arginine kinase (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, α-, γ- and Ω-gliadins, hordein, secalin, avenin, pathogenesis-associated protein from strawberry (Fra a 1), inhibitory protein (Mus a 1), a portion of any one of the antigens and a mimetic of any one of the antigens.

48. The tolerogenic compound according to claim 46 or 47, wherein the food antigen is selected from the group consisting of high molecular weight glutenins, low molecular weight glutenins, α-, γ- and Ω-gliadins, hordeins, secalins, avenins, a portion of any of said antigens and a mimetic of any of said antigens.

49. The tolerogenic compound according to any one of claims 46 to 48, wherein the food antigen is associated with diarrhoeal celiac disease.

50. The tolerogenic compound of claim 49, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 40-54, or a fragment thereof.

51. The tolerogenic compound of any one of claims 33 to 45, wherein the antigen comprises a self-antigen.

52. The tolerogenic compound of claim 51, wherein the autoantigen is selected from the group consisting of thyroglobulin, thyroid peroxidase, thyrotropin receptor, glutamate decarboxylase (GAD), 21OH hydroxylase, 17OH hydroxylase, H+ / K+ ATPase, intrinsic factor, transglutaminase, tyrosinase, tyrosinase-related protein 2, myelin basic protein, proteolipid protein, desmoglein, acetylcholine receptor, 2-oxoacid dehydrogenase complex, insulin, proinsulin, preproinsulin, insulinoma-associated protein 2 (IA-2), insulinoma-associated protein 213 (IA-213), ICA69, ICA12 (SOX-13), carboxypeptidase H, imogen 38, GLIMA 38. Chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-entero-pancreatic / pancreas-associated protein, S100β, glial fibrillary acidic protein, regeneration gene II, pancreatic duodenal homeobox 1, myotonic dystrophy protein kinase (DMPK), islet-specific glucose-6-phosphatase catalytic subunit-associated protein, SST G-protein coupled receptor 1-5, myeloperoxidase (MPO), proteinase 3 / myeloblastin, and a portion of any of the above antigens and a mimetic of any of the above antigens.

53. The tolerogenic compound of any one of claims 33 to 45 or 51 to 52, wherein the antigen comprises an antigen associated with an autoimmune disease.

54. The tolerogenic compound of claim 53, wherein the autoimmune disease is selected from the group consisting of type 1 diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphigus vulgaris, neuromyelitis optica, Goodpasture's disease, Parkinson's disease, myasthenia gravis, celiac disease with diarrhea, primary biliary cholangitis, Sjogren's syndrome, autoimmune hepatitis, myocarditis, inflammatory cardiomyopathy, and anti-neutrophil cytoplasmic antibody-associated vasculitis.

55. The tolerogenic compound of claim 53 or 54, wherein the autoimmune disease is type 1 diabetes.

56. The tolerogenic compound of claim 55, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

57. The tolerogenic compound according to claim 53 or 54, wherein the autoimmune disease is multiple sclerosis.

58. The tolerogenic compound of claim 57, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 71-106 or 157-159, or a fragment thereof.

59. The tolerogenic compound according to claim 53 or 54, wherein the autoimmune disease is: Rheumatoid arthritis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 107-116, 160-239 or 272-288, or a fragment thereof; Sjogren's syndrome, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 117-118 or 240-245, or a fragment thereof; Rheumatic heart disease, autoimmune myocarditis, viral myocarditis, or inflammatory cardiomyopathy, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 119 or 251-270, or a fragment thereof; Parkinson's disease, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 120 or 247-250, or a fragment thereof; Anti-neutrophil cytoplasmic antibody-associated vasculitis (ANCA vasculitis), wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 121-129, or a fragment thereof; Primary biliary cholangitis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 130-132 or 271, or a fragment thereof; or Autoimmune hepatitis, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 133-140, or a fragment thereof.

60. The tolerogenic compound of any one of claims 33 to 45, wherein the antigen comprises an alloantigen.

61. The tolerogenic compound of claim 60, wherein the alloantigen is selected from the group consisting of subunits of MHC class I and MHC class II haplotype proteins and their complexes with the antigens they present, and minor blood group antigens RhCE, Kell, Kidd, Duffy, Diego and MNSs.

62. A composition comprising a tolerogenic compound according to any one of claims 33 to 61 and a pharmaceutically acceptable excipient.

63. A method of inducing tolerance to an antigen to which a subject is capable of producing an undesirable immune response, the method comprising administering to the subject a tolerogenic compound according to any one of claims 33 to 61 or a composition according to claim 62.

64. The method of claim 63, wherein the tolerogenic compound or the composition is administered before the subject is exposed to the antigen, after the subject is exposed to the antigen, or both.

65. The method of claim 63 or 64, wherein the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

66. A tolerogenic compound according to any one of claims 33 to 45 or a composition according to claim 63 for use in inducing tolerance to an antigen in a subject to which the subject is capable of generating an undesirable immune response.

67. The tolerogenic compound or composition for use according to claim 66, wherein the undesirable immune response is associated with an allergic reaction to a food, animal, plant or environmental allergen, an autoimmune disease, a therapeutic agent, or graft-versus-host disease.

68. A tolerogenic compound according to any one of claims 33 to 45 or a composition according to claim 62 for use in the preparation of a medicament.

69. A compound according to any one of claims 33 to 45 or a composition according to claim 62 for use in inducing immune tolerance in a subject in need thereof.

70. A method of inducing tolerance to an antigen to which tolerance is desired, the method comprising: A compound comprising: (i) an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3; wherein the HCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO:9; wherein the HCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO: 10; and wherein the HCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO: 11; and (ii) the antigen to which tolerance is desired, The ASGR1 binding polypeptide is conjugated or fused to the antigen to which tolerance is desired.

71. The method of claim 70, wherein the ASGR1 binding polypeptide further comprises a light chain variable region comprising LCDR1, LCDR2, and LCDR3; wherein the LCDR1 comprises a sequence having at least 90% identity to SEQ ID NO:25; wherein the LCDR2 comprises a sequence having at least 90% identity to SEQ ID NO:26; and wherein the LCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO:

27.

72. The method of claim 70 or 71, wherein the antigen to which tolerance is desired is associated with diarrheal celiac disease.

73. The method of claim 72, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 41, 40, 43-54, or a fragment thereof.

74. The method of claim 70 or 71, wherein the antigen to which tolerance is desired is associated with MS.

75. The method of claim 74, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 100, 71-99, 101-106 or 157-159, or a fragment thereof.

76. The method of claim 70 or 71, wherein the antigen to which tolerance is desired is associated with type 1 diabetes.

77. The method of claim 76, wherein the antigen comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs: 55-70 or 153-156, or a fragment thereof.

78. The method of any one of claims 70 to 77, wherein the administration is via an intravenous route.

79. A method of delivering an antigen to liver tissue of a subject, the method comprising: conjugating or fusing the antigen to an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide, thereby generating an ASGR1 binding polypeptide-antigen complex; and contacting the ASGR1 binding polypeptide-antigen complex with liver tissue of a subject in situ, wherein the contacting allows the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, thereby delivering the antigen to the liver tissue.

80. A method of delivering an antigen to liver tissue of a subject, the method comprising: contacting an asialoglycoprotein receptor 1 (ASGR1) binding polypeptide conjugated or fused to an antigen with liver tissue of a subject in situ, thereby allowing the ASGR1 binding polypeptide to bind to ASGR1 on the liver tissue, wherein the ASGR1 binding polypeptide comprises a heavy chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3; wherein the HCDR1 comprises a sequence that is at least 90% identical to SEQ ID NO:9; wherein the HCDR2 comprises a sequence that is at least 90% identical to SEQ ID NO: 10; and wherein the HCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO: 11; and wherein the ASGR1 binding polypeptide comprises a light chain variable region, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3; wherein the LCDR1 comprises a sequence having at least 90% identity to SEQ ID NO:25; wherein the LCDR2 comprises a sequence having at least 90% identity to SEQ ID NO:26; and wherein the LCDR3 comprises a sequence that is at least 90% identical to SEQ ID NO:

27.

81. The method of claim 79 or 80, wherein delivering the antigen in situ to liver tissue of the subject induces tolerance to the antigen in the subject.

82. A method according to any one of claims 80 to 81 for the treatment of an autoimmune disease.

83. A method according to any one of claims 80 to 82 for treating an allergic reaction.

84. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2, HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO: 297; wherein the HCDR2 comprises the sequence of SEQ ID NO: 298; wherein the HCDR3 comprises the sequence of SEQ ID NO: 299; wherein the LCDR1 comprises the sequence of SEQ ID NO:313; wherein the LCDR2 comprises the sequence of SEQ ID NO: 314; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

315.

85. The ASGR1 binding polypeptide of claim 84, wherein the heavy chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:

303.

86. The ASGR1 binding polypeptide of claim 84, wherein the light chain variable region comprises a sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO:

319.

87. The ASGR1 binding polypeptide of any one of claims 84 to 86, wherein the heavy chain variable region comprises the sequence of SEQ ID NO: 303, and the light chain variable region comprises the sequence of SEQ ID NO:

319.

88. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2, HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO: 329; wherein the HCDR2 comprises the sequence of SEQ ID NO: 330; wherein the HCDR3 comprises the sequence of SEQ ID NO: 331; wherein the LCDR1 comprises the sequence of SEQ ID NO: 345; wherein the LCDR2 comprises the sequence of SEQ ID NO: 346; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

347.

89. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2, HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO: 361; wherein the HCDR2 comprises the sequence of SEQ ID NO: 362; wherein the HCDR3 comprises the sequence of SEQ ID NO: 363; wherein the LCDR1 comprises the sequence of SEQ ID NO: 377; wherein the LCDR2 comprises the sequence of SEQ ID NO: 378; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

379.

90. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO: 393; wherein the HCDR2 comprises the sequence of SEQ ID NO: 394; wherein the HCDR3 comprises the sequence of SEQ ID NO: 395; wherein the LCDR1 comprises the sequence of SEQ ID NO:409; wherein the LCDR2 comprises the sequence of SEQ ID NO:410; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

411.

91. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2, HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO:425; wherein the HCDR2 comprises the sequence of SEQ ID NO:426; wherein the HCDR3 comprises the sequence of SEQ ID NO: 427; wherein the LCDR1 comprises the sequence of SEQ ID NO:441; wherein the LCDR2 comprises the sequence of SEQ ID NO:442; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

443.

92. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO: 457; wherein the HCDR2 comprises the sequence of SEQ ID NO: 458; wherein the HCDR3 comprises the sequence of SEQ ID NO: 459; wherein the LCDR1 comprises the sequence of SEQ ID NO:473; wherein the LCDR2 comprises the sequence of SEQ ID NO: 474; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

475.

93. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2, HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO: 489; wherein the HCDR2 comprises the sequence of SEQ ID NO: 490; wherein the HCDR3 comprises the sequence of SEQ ID NO: 491; wherein the LCDR1 comprises the sequence of SEQ ID NO:505; wherein the LCDR2 comprises the sequence of SEQ ID NO:506; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

507.

94. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO:521; wherein the HCDR2 comprises the sequence of SEQ ID NO:522; wherein the HCDR3 comprises the sequence of SEQ ID NO: 523; wherein the LCDR1 comprises the sequence of SEQ ID NO:537; wherein the LCDR2 comprises the sequence of SEQ ID NO:538; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:

539.

95. An asialoglycoprotein receptor 1 (ASGR1) binding polypeptide comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2, HCDR3, and the light chain variable region comprising LCDR1, LCDR2, and LCDR3; wherein the HCDR1 comprises the sequence of SEQ ID NO:553; wherein the HCDR2 comprises the sequence of SEQ ID NO: 554; wherein the HCDR3 comprises the sequence of SEQ ID NO: 555; wherein the LCDR1 comprises the sequence of SEQ ID NO:569; wherein the LCDR2 comprises the sequence of SEQ ID NO: 570; and Wherein the LCDR3 comprises the sequence of SEQ ID NO:571.

Images